Proceedings of the Indian Academy of Sciences Plant Science Section B Vol_91

Proceedings of the Indian Academy of Sciences

Volume 91, 1982

CONTENTS (Plant Sciences)

Thaxteriellopsis lignicola and its Moorella anainorph

C V Sitbramanian and G Sekar 1

A contribution to the embryology of Alysicarpus monillfer B.C.

V Seshavatharam 9

Noa-inheritance of isomerism in cocoyams T Venkateswarlu 17

Apomixis in Cenchrus glaucus Mudaliar ct Sundaraj C Shanthamma 25

Regeneration of plantlets from callus of Elettaria cardamomum Maton

TV K Srinivasa Rao, S Narayanaswamy, E KChako and R Dore Swamy 37

Studies in Cyperaceae : XVII Novelties in Finibristylis (L.) Vahl and their
vegetative anatomy E Govindamjalu 43

Embryological studies in three species of Cymbopogon Spreng (Poaceae)

S P Choda, Harsh Mitter and Ravfnder K JBhanwra 55

Reproductive efficiency of secondary successional herbaceous populations
subsequent to slash and burn of subtropical humid forests in north eastern
India K G Saxena and P S Ramakrishnan 61

Vesicular arbuscular mycorrhiza in subtropical aquatic and marshy plant
communities R Chaubat, G D Sharma and R R Mishra 69

Chandrasekharania : A new genus of Poaceae from Kerala, India

V J Nair, V S Ramachandran and P V Sreekumar 79

Chromosome relationships of spinous solanums

P £ Kirti and £ G S Rao 83

Groundnut rust-its survival and carry-over in India

P Subrahmanyam and D McDonald 93

Correlated promotion of ray-floret growth in chrysanthemum by potassium
chloride, gibberellic acid and sucrose

P Pardha Saradhi and H Y Mohan Mam 101

Nuclear behaviour during heartwood formation in Acacia auriculiformis

A. Cann K V Bhat and J D Patel 107

Identity of Ficus macrocarpa Wt. ex King (= F. amplocarpa nom. nov.)
and F. guttata (Wt.) King-A reinvestigation with anatomical evidence

E Gonndarajalu and f Masilamoney 115

Tke gentts JdcJdefia in Sottth India Ram Udar and Adanh Ktfaidr 131

ii Contents

Geocalyx Nees-a rare marsupial genus from India

Udar, S C Srivastava and Dhirendra Kumar k

Ontogeny of the paracytic stoma: Variations and modifications

Parveen Farooqui (nee Kiawai) 14

Growth response of some thermophilous fungi at different incubation
temperatures S Singh and D K Sandhu If

Studies on Beggiatoa : Distribution and growth in aquatic habitats of
Visakhapatnam M R R Mohan and A Narayana Rao 15

Photoperiodic control of extension growth, bud dormancy and flowering of
Nerium indicum Mill, and Thevetia pemviana Schum.

Kushal Singh, Surinder Kumar and K K Nanda 11

Interaction of kinetin with B group vitamins on the seedling growth of
green gram (Phaseolus radiatus L.)

P Gopala Rao and J Kodandaramaiah 18

Leaf architecture of apocynaceae / S S Mohan and J A Inamdar 1 £

Impact of extension growth and flowering on the cambial 'activity of
Delonix regia Rafin A K M Ghouse and Shamima Hashmi 2C

Pharrnacognostic studies on the flower of Mesua ferrea L.

Usha Shome, Shanta Mehrotra and H P Shanna 2J

Effect of C03 in overcoming self-incompatibility barriers in Brassica
campestris L. var. toria A S Dhaliwal and C P Malik 21

Pharmacognosy of the stems of Portulaca quadrifida L. and Portulaca
oleracea L. / Lai and A M Khan 23

Structure and function of a sub-tropical humid forest of Meghalaya I. i i
Vegetation, biomass and its nutrients Jasbir Singh and P S Ramakrishnan 2^

Structure and function of a sub- tropical humid forest of Meghalaya II.
Litter dynamics and nutrient cycling

Jasbir Singh and P S Ramakrishnan j 2i

Structure and function of a sub-tropical humid forest of Meghalaya III.
Nutrient flow through water Jasbir Singh and P S Ramakrishnan 2(

Anatomy of the seedling of the Leguminosae — I

Umavathi Hegde and V D Tilak 2£

Cork-warts in Eucalyptus species Parveen Farooqui (nee Kidwai) 2£

Pericarpial sclereids in some Mimosaceae

S Rangaiah, I L Kothari and G L Shah 25

Viability and infectivity of zoospores of Sclerospora graminicola (Sacc.)
Schroet in the soil C R Ramesh and K M Safeeulla 3(

Initiation, development and structure of root nodules in some members

of the tribe Tnfolieae (Papilionaceae) G L Shah cttld M Gopala Fao 3(

Contents iii

Turnera ulmifolia var. elegans x T. ulmifolla var. angustlfolia crosses and
its bearing on the taxonomy of the species

K Rajeev, P I Kwiachan and C A Nlium 319

Airborne pollen grains of Visakhapatnam : A combined field and air samp-
ling study A Janaki Bai and C Subba Rcddl 329

The floral anatomy of Kniphofia uvaria Hook. (Liliaceae : Knipholieae)

N P Vaikos and R M Pal 351

Transmission of seed-borne inoculum of Macrophomma phaseolina from

seed to plant Tribhuwan Singh and Dalblr Singh 357

Effect of water stress on opening and longevity of flowers in Gladiolus

/ V Ramanuja Rao and H Y Mohan Ram 371

Petal venation in Trigonella (Papilionaceae) Mohlnl Gupta 379

Responses of cotton-cultivars to long day conditions

J G Bhatt and M R K Rao 389

Seed germination and seedling establishment of two closely related Schiina
species Ram £oojh and P S Ramakrishnan 397

Anther and pollen development in cotton liaploids and their parent,

S S Mehetre 409

Changes in proteins, amino and keto-acids in different seedling parts of
Cyamopsis tetragonolobus Linn, during' growth in light and darkness

Prem Gupta and D Mukherjee 417

Effect of ridge gourd pollen on zoospore germination of Pseudoperonospora
cubensis and its significance in epidemiology

Amarnatha Shetty, H S Shetty and K M Safeeulla 427

Leaf proteinase and nitrate reductase activities in relation to grain protein
levels and grain yield in four species of grain amaranth

K Ramamurthy Naidu, Y Seethambaram and VSR Das 433

Cell division in Staurastrum gracile Ralfs. under the scanning electron
microscope Vidyavati 443

Leaf surface studies of some medicinal salvias H P Sharma and Usha Shomc 449

Morphological and metabolic changes in the egg and zygote of Lagers-
tromeia speciosa, I. Cell size, vacuole and insoluble polysaccharides

P Raghavan ami V J Philip -465

The floral anatomy of Pity a spathacea Mexz- (Promeliacene) with ;p cial
refereuce to nectaries R A Rulkarni and R M Pai 473

Cytological studies on certain acanthaceae from Central India

MIS Saggoo and S S Sir 479

Heterotrophic bacteria associated with seaweed

R Lakshmctnapemmalsamy and A Ptirwhothaman 487

IV

Contents

Association of chlorophyll content., phyllotaxy^ photosynthesis and B-group
vitamins in some €3 and C4 plants

P Gopala Rao and J Kodandaramaiah

Effect of morphactin, AMO-1618 and DPX 1840 on the endogenous levels of
hormones and its implication on apical domnance in Glydne max. Linn.

/ S Duai.U K Kohli and K S Chark

Taxonomic importance of epidermal characters in the Indian Thespesia
Corr. (Malvaceae) S Raja Shanmukha Rao and N Ramayya

Embryological studies in Launaea midicaulis Hook.

P S Chikkannaiah and B S Hiremath

Quantitative profile structure of certain forests in the Kmnaun Himalaya

A K Saxena and J S Singh

Contributions to our knowledge of Indian Algae — HI. Euglenineae-Part I.
The genus Euglena Ehrenber M T Philipose

fittc. Indian Acai Sci. (Plant Sci.j, Vol. 91, Number 1, February 1982* pp. 1-1
© Printed in India.

Thaxteriellopm lingicota and its Moorelta ariamorph*

C V SUBRAMANIAN and G SEKAR

Centre of Advanced Study in Botany, University of Madras, Madras 600 005, India

MS received 14 October 1981 ; revised 18 December 1981

Abstract. A. Moorelht anamorph referable to M. spedosa Rao and Rao, is described
for Tliaxtertetlopsis ligtiicola Sivaixesan, Panwar and Kaur. The connection is
established from a s-tudy of single ascosporc cultures of the fungus established
for several (6) South Indian collections. Both the teleomorph and anamorph are
described and brief notes are given of characteristics of this fungus in culture.

Keywords. Tltaxtenellojpsis Itgnicola ; Moorella spedosa ; toleomorph ; anamorph.

1. Introduction

During the survey of mierofuagi under the project ' Fungus $lora of South
India \ several collections of a Loculoascomyeete with setose ascomata were
made. Single ascospore cultures of the fuaigu* produced aoi anamorph referable
to the hyphomycctc genus Moorella. The Moorella anamorph was also found in
association with the Loculoascomyeete on the natural substrate. The teleomorph
was identified as Thaxlerltllopsti lignlcola Sivanesan, £*acnwar and Kaur, and the
anamorph as MooreJUi spedosa Rao ajid Rao.

The monotypic genus T/iaxtertellopsis was erected by Sivanesan et at (I9*t6)
with T. lignicola collected from Mount Abu, India as the type. It is characterized
by non-ostiolatc, setose, cupulatc ascomata, superficial on a subiculum, with
bitunicate asci containing hyaline or subhyaline ascospores, which are traaisversely
multiseptate. Some of the ascosporcs, in addition, develop one longitudinal or
oblique septum, rarely two in one or two of its cells.

In all the collections; studied, the fimgus agrees in all details with the type descri-
ption of T. lignlcola except for the difference in the number of the septa in ascos-
pores. The transversely multi septate asco.xpores, with one or two longitudinal
and obliquo septa in sonic., typical of T. lignlcola were seen only in one collection
from the Silent Valley, Kerala (Herb. FSI 3363). In the remaining collections
the ascospores were predominantly transversely 5-septate. The present study
shows that in T. l/gnicola the ascospores are initially 5-septate ; the centrum is
hyaline to white, as seen in most of the collections. As the ascospores mature,
additional transverse., longitudinal and oblique septa develop ; the centrum becomes
light brown, as seen in the single collection from the Silent Valley. The additional

* Memoir No, 352 from Centre of Advanced Study in Botany,

2 C V Subramanian and G Sekar

transverse, longitudinal and oblique scptc. develop in any sequence aond are forme
within any cell, without any regular pattern, in the originally 5-septate ascospore

2. Description of the fungus

Thaxteriellopsis lignlcola Sivanesan, Pan war and Kaur, Kavaka 4 : 39, 197
Aaamorph : Moorella spedosa Rao and Rao, Mycopath. Mycol. appl. 22 : 5
1964. (figures 1-19.)

Colonies on the substrate conspicuous, superficial, widely effused, black, consi
ting of velvety growth of anamorph, interspersed with scattered to gregarious asc<
mata of teleomorph. Surface mycelium composed of septate, branched, dai
brown, creeping hyphae 4-7/j wide, constricted at septa when cells are short an
swollen, smooth when the cells are long cylindrical. Immersed mycelium consi
ting of septate, branched, light brown or dark brown hyphae ca. 5p wide.

Ascomata superficial, connected to the creeping mycelium by septate, bro\\
pendant hyphae ca. 5p wide, originating from the lower half of the ascoma, son
seated on a stroma directly on the substrate, black, cupulate, in water becomir
spherical to broadly spherical or obconic-spherical, 20(WOO// high, 250-37(
diam., non-ostiolate, with numerous dark brown, septate, uabraached setae, wil
rounded ends, mostly on the upper half of the ascoma. Setae up to 250/z Ion
4-5-6-0/* wide. Wall of the ascomata somewhat fleshy, 35-50^ thick, pseud<
parenchymatous, composed of 6^9 layers, made up of polygonal cells, 5-20 :
5^13/j ; cells of 'ostiolar' region comparatively smaller, 5-10 x 3-5/j ; eac
cell with a large oil droplet, oozing out in teased mounts ; outer layers darkbrowj
gradually becornbag light brown to hyaline towards the inner layers ; cells of tl
inner layers flattened.

Asci in a basal hym^ium, bitunicate, long-cylindrical tocylindric-clavats, shor
pedicellate, 90->150 x 15-30/f, generally 8-spored ; sometimes fewer-spored.

P^eudoparaphyses present, attached both to the roof and the basal hymeniun
septate, hyaline, branched, ca. 1-5^ wide.

Ascospores irregularly tristichous, 23-50 x 5-5-13 (-^16)^, initially with
transverse septa. Mature ascospores with additional transverse, longitudin*
and oblique septa, up to 6-12-transversely septate aoid often with up to 3 long
tudinal and/or oblique septa, clavate to clavate-fusiform or fusiform \ general]
the second and thir doells from apex broader, tapering towards the rounded end
curved to straight, thin-walled, slightly constricted at septa in large spores, faint]
striate, hyaline to smoky brown in mass.

Conidiophores erect, straight or slightly bent, arising directly from creepin
mycelium, septate, smooth, up to 420/z long, blackish brown and 7-11/z wide 2
base, gradually becoming pale brown and 4^7/j wide at apex, bearing up to si
whorls of branches at intervals, with one to six branches per whorl ; branchc
1-4-celled, 4-5-7-5/J wide. Conidiophore often terminating in an apical who:
of branches or a conidiogenous cell. Terminal cell of each branch conidiogenou;
Conidiogenous cells hyaline, light brown to brown, flask-shaped to globose o
cylindrical, often proliferating, polyblastic, denticulate. Conidia helicoid, tight!
coiled Hi times, borne on short cylindrical denticles ca. I// long, 3-.10 septafc
generally 6-septate, 1 W5/j diam. ; filaments 4-6/* wide, hyaline, finally becomin
smoky brown with faint str-j^ttons,.*- • <-....,.

Thaxteriellopsis liguicola and its Moordla

II

Figures 1-11. tfiaxteriellopsis lignlcola and its Moordla anamorph. t. Vertical
soctiaa o,f ascoma (Kerb. FSI 3359) x 200. 2. Predominaatly 5-septatc ascos-
p^res (Kerb. FSE 3151) X 700. 3. M'Uure ascosporcs showing longitudinal and
oblique septa (Herb FSI 3363) x 480. 4. Part of a conidiopliorc with ccmidiogenous
colls and a coiidium initial (Herb. FS[ 3151) x 1200. 5. Conidia from natural
substrate (Herb. FSI 3363) x 750. 6. Genniii'-Ued ascosporc (Herb. FSI 21 2»)
X 375. 7. A conidiogenous cell with a developing -co ntdium (from culture, Herb.
FSt 3l5l)"x 1200, 8. A short conidiophore with a solitary terminal conidium (from
culture, Herb. FSt 3151) x 1065. 9, 10, 11. Mature brown conidia from a.
6 moatli's old culture (Herb, FSI 3151) X 1600,

Thaxteriellopsis ligniaola and its Moorella anamorph

Figures 12-19. Thaxteriellopsis lignicola and its Moorella anamorph. 12. Ascus
(Herb. FSI 3363). 13. Ascospores (Herb. FSI 3363). 14. Young conidia deve-
loping directly on, a hypha (from culture, Herb. FSI 1423). 15. A conidium borne
on a single conidiogenous. cell produced laterally on a hypha (from culture, Herb.
FSI 1423). 16. A conidiophore with several conidia produced from its apical part
(from culture, Herb. FSI 1423). 17. Conidia from natural substrate (Herb. FSI
3151). 18. Conidia (from culture, Herb. FSI 3151). 19. Brown conidia from ^
$ month's old culture (Herb. FSI 3151).

6 C V Subfamanian and G Sekar

Collections examined :

All collections by G. Sekar.
Kamataka State

Ch indst. wood, Ksmphole Forest, near Sakleshpur, Hassan District, 31 October
1977, Herb. FST 1423 ; on indet. wood ; Muadwar Pimpley Forest, near Londa
Belgaum District, 2? November 1980, Herb. FST 3204 ; on indet. bark, Karwar,
North Kanara District, 18 November 1980, Herb. FSI 3151.
Kerala State

On indet. wood, Silent Valley, Palghat District : Kumattanthod, 5 December
1980, Herb, FSI 3225, 3230 ; Dam site, 6 December 1980, Herb. FSI 3257 ; camp
site, 7 Dxember 1980, Herb. FSI 3261 ; near Valaiyampara Estate, 8 December
1980, Hfeib. FSI 3271 ; Mathrithod, 11 December 1980, Herb. FSI 3321 ;. Valajya^
parathod, 13 December 1980, Herb. FSI 3356, 3359. On wood and bark of
Syzygium cumini (Linn.) Skeels, Valaiyaparathod, Silent Valley, Palghat District,
13 December 1980, Herb. FSI 3363. On indet. wood, Karapara Forest, Palghat
District, 21 December 1980, Herb. FSI 3413, 3417 ; 24 December 1980, Herb.
FSI 3461.
Tamil Nadu State

Oa wood of Syzygium cumini S:egaltheri, Kalakad, Tirunelveli District, 13
February 1979, Herb. FSI 2048. On indet. wood, Muadanthurai Forest, Tirune-
lveli District, 16 February 1979, Herb. FSI 2128. On indet. bark, Muadanthurai
Forest, Tiruaelveli District, 18 February 1979, Herb. FSI 2166. On wood
of Mango, Kakkanalla bridge, Mudumalai Forest, Nilgiri District, 12 January 1981,
Herb. FSI 3480.

3. Cultural studies

Ascospores germinate on potato dextrose agar within 8 h at 25-30° C. Germ
tubes are produced from any one or all the cells of ascospores (figure 6), more
frequently from the end cells. Single spore isolations on potato dextrose agar
are slow growing, attaining 1cm. diameter in 10 days. The colonies are olive
green to olivaceous brown, velvety, with compact margin, restricted in growth
with aerial mycelium and dark brown submersed mycelium. The conidiogenous
cells are either intercalary or terminal on hyphae, or on a simple conidiophore
(figures 8 and 16). Typical dark brown erect conidiophores with whorls of
branches bearing conidiogenous cells are produced after three months in culture.
Conidia are produced on narrow, cylindrical denticles, ca. lju long and are loosely
or tightly coiled or juit bent and curved. Conidium size is comparable to that
on the natural substrate, but soim may be a little larger. In a long standing
culture, conidia (figures 9-^11 and 19) become brown, up to 12-septate and measure
up to 23/j diameter with filaments 9/j wide, constricted at the septa.

4. Discussion

Thaxteriellopsis is a member of the Loculoascomycetes and belongs to the Pleos-
porales ; in having Pleospora type of centrum. It is closely related to Thaxtertella,
as pointed out by Sivanesan et al (1976), aoid possibly to Tubeufia. The anainorpl}

ThaxterieUopsis Ugnicola and its Moofella anamofph 7

of Thaxteriellopsis, as shown here, is Moorella and that of Thaxterlelta and Tubeufia
are Helicorna and Helicosporiitm respectively. It is interesting to note thet all the
three teleomorphs mentioned above have somewhat fleshy ascornata, vrith usually
hyaline ascospores and their anamorphs are dematiaceous hyphomycetes producing
helicoid conidia on denticles. This is the first time a Moorella aaiamorph is con-
nected with a teleomorph.

Barr (1980), notes that the illustration and description of Thcateriellopsis ligni-
cola are suggestive of Boerlagiomyces Butzin. However, even if this v/ere to be
confirmed by further study of the types, the generic name ThaxterieUopsis \vill have
to be retained according to the rules of priority. The name ThaxterieUopsis
Ugnicola is therefore used here for the teleomorph of our fungus.

Acknowledgements

One of the authors (GS) is grateful to University Grants Commission for the award
of Junior Research Fellowship, under the project * Fungus Flora of South India *.
They also thank Mr V Kaviyarasan for photographic assistance.

References

Barr M E 1980 On the family Tubeufiaeeae (Pleosporales) ; Mycotaxon 12 137-167
Sivanesan A, Panwar K S and Kaur S J 1976 Thaxterieljopsis Ugnicola gen, et sp. nov., a new
Locmloascomycete from India ; KqvcikQ 4 39-42

l?roc. Indian Acad. Sci, (Plant Sci), Vol. 91, Number 1, February 1982, pp. 9-15,
© Printed in India.

A contribution to the embryology of Alysicarpus monilifer D.C.

V SESHAVATHARAM

Department of Botany, Andhra University, Waltair 530003, India

MS received 25 April 19S1 ; revised 4 February 1952

Abstract. Embryology of Alysicarpus monilifer D.C. is described. The anther wall
comprises the epidermis, the fibrous endothecium, two middle layers and uniseriate
glandular tapetum. Pollen is shed at the 2-celled stage. The ana-campylotropus
ovule is crassinucellate and bitegmic. The archesporium in the ovule is hypodermal
and cuts off a parietal cell. Embryo sac development conforms to the Polygonum
type. Endosperm development is nuclear. A chalazal part of the endosperm
remains free nuclear and is haustorial in function. Embryogeny conforms to
Period I Megarchetype IV and Series Ba. The embryogenic classification of the tribe
is discussed in the light of the present observation and earlier reports.

Keywords. Embryology ; Alysicarpus.

1. Introduction

Tne tribe Hedysareae of the Papilionaceae includes 47 genera and 7000 species
distributed in the tropical and subtropical regions of the world (Rendle 1925). Of
these only 14 genera are known embryologtcally. The researches of Soueges (1947,
1953a, 1953b, 1955, 1956), Johansen (1950), Rau (1951, 1953, 1954), Smith (1956),
Goursat (1961), Kaprokar (1964) and Dsshpande et al (1976) reveal the cmbryogeny
of 15 species belonging to 11 genera of this tribe. The embryology of Alysicarpus
monilifer D.C. is described here.

2. Material and methods

Buds, flowers and fruits of varying ages were collected from the plants growing
in the university campus, during the rainy season and were fixed in FAA. Custo-
mary methods of dehydration and embedding, etc. were followed. Sections cut
between 8 and 10 microns thick were stained with safranin and fast green. The
endosperm was observed in whole mounts after making suitable dissections.

3. Observations

3-1. Microsporogertesis and the development of pollen

The archesporium in the anther is hypodermal represented by a single row of 4-^5
cells. The primary parietal layer by further divisions forim 2-3 parietal layers
of which the innermost forim the secretory tapetum (figure 1). The tapetal cells
remain uninu^leate throughout. Some of the cells on the connective side adjoining

10

V Seshavatharam

the tapetum also behave like the tapetal cells in assuming a dense cytoplasm and
deeply stained nuclei. The hypodermal parietal layer forms the fibrous endo-
thecium (figure 3). The two middle layers are crushed in the mature anther.
Division of the microspore mother cell is simultaneous a»nd cytokinesis takes place
by furrowing (figure 2). The pollen is shed at the 2-celled stage.

Figures 1-9. 1. T.S. anther lobe showing wall layers and sporogenous tissue ;
2. Same showing cytokinesis in pollen mother cells ; 3. Same showing fibrous
endothecium and two celled pollen grains ; 4. L.S. portion of ovule showing a
linear tetrad of megaspores ; 5. LS ovule showing the integuments endosperm and
barrier tissue at the globular stage of embryo ; 6. Mature embryo sac ; 7. Nuclear
endosperm showing chalazal elongation ; 8. LS young seed showing the embryo
and endosperm ; 9. LS portion of the ovule at the chalazal regions showing the
incipient integumentary tapetum and thick walled barrier tissue,

Embryology of Alysicarpus monilifer D.C. 1 1

3*2. Megasporogenesis and female gametophyte

The ovule at maturity is campylotropous bitcginic and crassinueellate (figure 5).
The single celled archesporium in the ovule is hypoderinal and cuts off a
parietal cell. Tlie mogaspore mother cell undergoes the usual meiotic divisions
resulting in a linear tetrad of magaspores of which the chalazal is functional
(figure 4). Tne development of the mature embryo sac conforms to the Polygonum
type. Tnc mature megagametophyte is 8-nucleate with an egg apparatus, two
polar nuclei and three antipodal cells (figure 6). The antipodal cells are ephemeral
and degenerate prior to fertilization.

3*3. Endosperm

Tlie primary endosperm nucleus divides much earlier than the zygote and the
development of the endosperm is of the nuclear type. The endosperm remains
free nuclear until the embryo reaches the early dicotyledonous stage (figure 8).
During the course of its development the embryo sac enlarges enormously and
encroaches on the surrounding nucellar tissue. This is more prominent at the
micropylar region where it comes in direct contact with the innermost layer of the
inner integument. At the chalazal region the tubular free nuclear part assumes a
haustorial role (figure 7). The active growth of the endosperm haustorium at the
chalazal end is arrested by the development of thick walled barrier tissue (figure 5).
The cells of the innermost layer of the inner integument have dense contents and
arc prominent (figure 9).

3-4. Embryo

The developmental sequence in the embryogeny is illustrated in figures 10 to 23.
The zygote divides transversely resulting in a two celled proembryo. The apical
cell (ca) divides by an obliquely vertical wall resulting in two unequal cells
(figure 11). Another oblique division in the larger derivative cell'of the tier (cal)
cuts off an epiphyseal initial (e) (figure 12). The basal cell (cb) undergoes a trans-
verse division resulting in m and ci. Further divisions in the apical quadrant
demarcate the tiers / and /'. Periclinal divisions in both the tiers differentiate the
dermatogen. The epiphyseal initial undergoes a vertical division and by further
divisions contributes to the stem tip in the mature embryo.

The cell m divides by a vertical wall and both the cells divide transversely. Their
upper derivatives contribute to the root tip and the root cap, while the deriva-
tives of the lower cells together with those of tiers /* and n' contribute to the long
and massive suspensor. A schematic representation of the zygote derivatives
and their destinations in the mature embryo is given below :

Stem tip
•i Cotyledons

-I Hypocotyl and radicle
Zygote

Root tip and root cap
Suspensor

12

V Seshavatharam

21

3 mm

Figures 10-23. Stages in the development of embryo.

Thus the enibryogeny according to Soueges scheme (Crete 1963) falls under
Period I, Megarchetype IV and Series B2.

4* Discussion

The archesporium in the ovule is hypodermal in all the species investigated so far
in the tribe the sole exception being Desmodium pmiculatum where a subhypo-
dermal archesporium was reported by Rembert (1969). However, this needs
verification in view of its rare occurrence in the family Papilionaceae.

In Alysicarpus morilifer wall formation in the endosperm is initiated after the
differentiation of the cotyledonary lobes in the embryo and is further restricted
to the upper half of the embryo sac, while the chalazs 1 part functions as the haus-
torium. A similar feature has earlier been reported by Rau (1953) in Desmodium
tfiflomm, D. tortulosum, D. pulchellum, Eleiotis soraria and Aeschynomene indica ;
by Kapuskar (1964) in Aeschynomene aspera and by Deshpande et al (1976) in
Zornia diphylla. Stylosanthes mucronata (Rau 1953) is so far the only member
of the tribe where a cellular endosperm is not organised at all in the developing
seed.

The tribe Hedysareae shows considerable variation in the mode of embryo
development and in the nature and organisation of the smpensor. The variation
met with in the embryogeny of the tribe is summarized in table 1.

Embryology of Alystearpus monilifer D.C.

13

Table 1. Variation in the embryo development among the members of the tribe
Hcdysarcac.

Species investigated

Author

Embryo typo, afler Embryo type, after
Johanscn 1950 Soueges 1948

Coronilla minima

Desmodium canescens
D. canadense
Hcdysaruni coronarium
Hippocrepis comosa
lespcdeza violacca

Jrnithopus jpcrpusittus
}ttobrychis sativa
korpiurus vermicutata
lornia diphylla

ieschynoment indica
i. aspcra

Ittevigatum

Soueges 1947 Coroniiia variation Period I, Megarchc-

of the Oaugrad type VI, series A
type

Rau 1954 do. do.

Ran 1954 do. do.

Soueges 1956 ' do. do.

Soueges 1955 - do. do.

Rau 1954 do. do.

Soueges 1953a do. do,

Soueges 1953b do. do.

Goursat 1961 do. do.

Dcshpande and do. do.
Bhasin 1976

Rau 1951 do. do.

Kapuskar 1964 do. do.

Rau 1954 do. Period I, Megarche-

type VI, Series B

tlysicarpus monilifer

Present study

Alysicarpus varia-
tions of the
Onagrad type

Period I, Megarche-
type IV, Series B

Irackis kypogca

Smith 1956

Solanad type

Period I, Megarche-
type V, Series C

fedystrum nutats

Johansen 1950

Caryophyllad type

Period II, Megarche-
type IV, Series A

The cmbryogeny in a majority of the species follows a more or less uniform
pattern agreeing with that origii\3lly described by Soueges (1947) in Coronilla
minima and belongs to the first embryogenic group under Period I, Megarchetype
n and Series A. According to the system of Johaoiseji (1950) the embryogeny
a all these species conforms to the Onagrad type, but does not fit into any, of
he variations proposed by him. Hence it is now proposed to erect a new varia-
ion designated as * Coronilla variation * under the Onagrad type to accommodate
11 these genera of the tribe Hedysareae, belonging to Period I, Megarchetype VI
nd Series A, where cb = s.

14 V Seshavatharam • . .

Rau (1954) observed that Desmodium laevigatnm differs in its embryogeny from
the rest of the members of the tribe which necessitates further investigations to
evaluate the significance of this variation.

Alysicarpus monilifer shows further deviation from the fundamental type, where
the embryogeny falls under the second group along with Desmodium laevigatum
but belongs to the fourth megarchetype in period I where cb = iec + co + s. In
this respect Alysicarpus differs from the rest of the Hedysareae and resembles
members of Phaseoleae and Galegeae.

According to Johansen's system, the embryogeny in Alysicarpus rnonilifer
conforms to the Onagrad type but docs not fit into any of the variations proposed
under that type. The erntryogeny shows resemblence to the Trifolium variation
of the Onagrad type in the possession of the epiphyseal initial, but differs in the
destination of the derivatives of the basal cell cb. In Trifolium variation, based
on the embryogeny in Trifolium minus (Soueges 1927) the entire basal cell cb
contributes to the suspensor (cb = s) whereas in Alysicarpus the derivatives of
the basal cell contribute to the root tip and root cap in addition to suspensor
(cb = iec -f co 4- s).

Hence, a new variation designated as Alysicarpus variation is proposed to accom-
modate the type, having an epiphyseal initial and where the derivatives of the tier
cb contribute to the root tip and root cap in addition to the suspensor. Alysi-
carpus is so far the only genus in this tribe where an epiphyseal initial is demar-
cated in the tier ca and cb = iec + co + s. In this respect it resembles Rhynchosia
suaveolens of Phaseoleae, and Tephrosia strlgosa of Galegeae (Seshavatharam 1969)
suggesting a relationship between these tribes and Hedysareae embryogenicaliy. •

Acknowledgements

The author is grateful to late Professor J Venkateswarlu, for suggesting the
problem and for his encouragement during the progress of this work.

References

Crete P 1963 Embryo. In tycent advances in Embryology of angiosperms (ed.) £. Maheswari
Intl. Soc. Plant Morphologists, New Delhi pp. 171-222

Deshpande P K and Bhasin R K 1976 A contribution to the life history of Zornia diphylla Pers;
/. Indian JBot. Soc. 55 115-124

Goursat M J 1961 Etnbryogenie des Legumineuses. Developpement de Pembryon chezl e Scor-
piurus vermiculata L. ; C.R. Acad. Sci. Paris 253 307-309

Johausen D A 1950 Plant Embryology., Waltham Mass . \ •

Kapuskar A T 1964 Studies in the embryology of Legumtnosae II. Embryo development in '
Aesckyrtomene aspera Linn. ; Proc. Indian Acad. Sci. B60 87-94

Rati MA 1951 Development of embryo in Aeschynomene indica Linn. ; New Phytol 50 124-126

Rau M A 1953 Some observations on the endosperm in Papilionaceae ; Phytomorphology 3
209-222

Rau M A 1954 The development of embryo of Cyamopsis, Desmodium and Lespedeza, with
a discussion on the position of Papilionaceae in the system of embryogenic classification;
Phytomorphology 4 418-430 ' .

Embryology of Atysicarpus monilifer D.C. 15

Rembert D H (Jr) 1969 Comparative megasporogenesis in Papilionaceae ; Am. J. Bot. 56

584-591

Rendle A B 1925 Tfie classflcation of Flowering plants, Cambridge University Press 2

Seshavatharam V 1969 A contribution to the embryology of Papilionoideae and Caesalpinioideae ;
Ph.D. Thesis, Andhra University.

Smith B N 1956 Arachis hypozaea. Embryogeny and the effect of peg elongation upon embryo
and endosperm growth ; Am. J. Bot. 43 233-240

Soueges R 1927 Embryogenie des Legumineuses. Developprncnt du proembryon Chez le Tri-
folium minus Rehl ; C.R. Acad. Sci. Parts 184 1018-1019

Sou6ges R 1947 Embryogenie des Papilionacces. Developpement de 1'embryon Chez le Coronllla
minima L. ; C.R. Acad. Sci. Pans 224 1254-1256

Sou6ges R 1948 Embryogenie ct classification 3 fasciculac Paris

Sou6ges R 1953a Embryogenie des Papilionacecs. Developpement de T cmbryon Chez T Orni*
tropus perpusillm L. ; C.R. Acad. Sci. Paris 237 1199-1201

Soueges R 1953b Embryogenie des Papilionacees. Developpement de 1'embryon Chez 1' Onobry*
chis sativa Lam. C.R. Acad. Sci. Paris 237 450-452

Soueges R 1955 Embryogenie des Papilionacees. Developpment de 1'embryon Chez T Hippo-
crepis comosa L. ; C.R. Acad. Set. Paris 24§ 2100-2103

Sou6ges R 1956 Embryogenies des Papilionacees. Developpement dc 1'cmbryon Chez 1* Hedysa*
rum coronarium L. ; C.R. Acad. Sci. Paris 242 704-707

Proc. Indian Acad. Sci. (Plant Sci.), Vol. 91, Number 1, February 1982, pp. 17-23.
© Printed in India.

Non-inheritance of isomerism in cocoyams

T VENKATESWARLU

Central Tuber Crops Research Institute, Trivandrum 695017, India

MS received 16 July 1981

Abstract. The present communication describes the non-inheritance of isomerism
in prefoliation and contortion of spathes in cocoyams. Importance of isomerism
from the point of taxonomy and agronomy has been discussed. It is expected that
a study on the orientation of leaf promordia at shoot apex and their spiral
descendance would help in unravelling the hidden laws in shoot morphogenesis
of cocoyams.

Keywords. Isomerism ; cocoyams ; dextro ; levo ; contortion ; prefoliation ;
taro ; taunia.

1. Introduction

Isomerism in biological materials was described by many workers under different
names such' as asymmetry (Davis 1962) bioisomerism (Meyen 1973 ; Venkates-
warlu and Hrishi 1977) enantiomorphism (Davis 1974), isomerism (Bahadur and
Venkateswarlu 1976 a,b), chinlity (Venkateswarlu 1978) and radtel symmetry
(Davis 1978).

Isomerism in prefoliation and contortion of spathes in cocoyams was described
in detail (Venkateswarlu and Hrishi 1977, Venkateswarlu 1978). In some crops
this phenomenon is correlated with yield (Davis 1972). The present study is taken
up to see if the corm yield is associated with prefoliation and if so whether this
trait is transmitted through cormels and true seeds. Kasinov (1969) described
that this character is transmitted through fronds in Lemna gibba L. The present
paper describes the investigations carried out on the pattern of prefoliation in tuber
and seedling progenies in taro and only tuber progenies in taunia.

2. Materials and methods

All the specimens used were taken from the collection of germplasm of edible
aroids maintained at the Central Tuber Crops Research Institute, Trivandrum.
The method of scoring a plant into levo and dextro was the same as that reported
earlier (Venkateswarlu and Hrishi 1977 ; Venkateswarlu 1978). Observations on
transmission of prefoliation through cormels in taro and tannia were recorded
in 1978. The seed tubers were selected from labelled plants and sown. Observa-
tions on prefoliation were recorded $t weekly intervals1. Based o» the prefoliation

17

18 T Venkateswarlu

pattern, plants were classified as levo, dextro and bichin 1. Observations were
also made on prefoliaticm pattern on the main pseudostem and on the suckers
developing from the main clump.

For inheritance studies in taro seedlings, the nature of contortion in spathe \vas
recorded at the time of effecting crosses and labelled as C95 L x C 271 D etc.
as the case may be. Seeds obtained from the cross were sown and observations
were recorded from the spathes; produced in the seedlings, Observations were
also made on prefoliation in. taro seedlings,

3. Results and discussion

3*1. Observations on Frefoliatfon

The phyllotaxy in cocoyaim is alternate. Unfurling of leaves is either clockwise
(levo) or anticlockwise (dextro) (figures 1-6). Plants were labelled as levo, dextro
and bichiral based on their morphology during the growth period. The corm
and cormels of labelled plants were sown during 1978 to see whether the nature
of prefoliation is transmitted vegetatively. The prefoliation pattern of seed setts
and the resulting new pseudostems in both taro and tannia are presented in table 1 .

Out of 60 levo tubers planted in taro, 28 grew into levo, 28 as dextro and 4
bichiral types i.e., levo and dextro in 1:1 ratio. Total dextro tubers planted
in taro were 21, out of which 9 developed as levo, 12 as dextro. Interestingly,
no bichiral plants were produced when dextro tubers were planted.

In tannia, bichiral plants were produced from both levo and dextro tubers. In
all, 38 levo tubers were planted, from which 11 levo, 21 dextro and 6 bichiral
plants were produced. From 42 dextro tubers sown, 17 behaved as levo, 21 as
dextro and 4 as bichiral. There seems to be no association between the seed sett
and the preudostems in cocoyams similar to that of Costus speciosus (Davis 1978).

Observations were also made to know whether the new suckers developing from
the seed sett differ in the nature of isomerism in prefoliation in taro. The data
are presented in table 2. From the data presented in tables 1 and 2, it is obvious
that the isomerism in 'prefoliation of cocoyams is not transmissible vegetatively
which is contradictory to the results of Kasinov (1969) in Lemna gibba. further
the whole phenomenon is complicated by an irregular pattern the suckers show.

Similar observations were noticed in the seedlings of taro also. Levo and dextro
seedlings were produced almost in a 1 : 1 ntio indicating the non-genetic nature
of the character. This is similar to the findings of Davis (1962) in coconut.

3-2. Contortion of spathes in taro seedlings

An attempt was also made to study the inheritance pattern of spathe contortion
in taro seedlings. Crosses were made in all possible combinations (L x L,
L x D, D x D and D x i). At the time of flowering, the nature of contortion
in spathes was recorded. Seedlings of all the crosses flowered except of D >< D.
The data on the asymmetry of spathe are presented in table 3. It is clear from
the data that both levo and dextro spathes were produced in almost ecjuai numbers
the same plant. Davis(1962) also reported similar 1 ; 1 ratio of left

Non-inheritance of isomerism in cocoyams

19

Figures 1-6. 1. Levo prefoliation in taro, 2. Dextro prefoliation in taro, 3. Levo
contortion of spathe in taro, 4. Dextro contortion of spath© in taro, 5. Levo
prefoliation in tannia, 6. Dextro prefoliation in tannia,

20

T Venkateswarlit

Figures 7-8. 7. Levo clextro prefoliation in the same clump of taro, 8. Lcvo
and doxtro prefoliation in the same clump of tannia.

Non-inheritance of isorncrlsm in coco yams
Table 1. Isomerism in prefoliation of seed setts and new pseiidostems.

21

Name of the Nature of No. of

crop seed sett seed setts

planted

Isomerism in aew pseuclostems

Levo

Dexlro

Bichiral

Total

Taro

Levo

60

28

28

4

60

Dextro

21

9

12

...

21

Total

81

37

40

4

31

Tannia

Levo

n

11

21

6

38

Dextro

42

17

21

4

42

Total

80

28

42

10

80

right spiralled plants in coconut in all the four combinations indicating that this
character is not genetically controlled.

Furthermore, the fact that both levo and dextro rotatory leaves and spathes are
produced on the same plant suggests that this phenomenon is non-genetic
(figures 7,8). At the same time, it is interesting to see some plants behaving
either as levo or dextro throughout their growth period with respect to both
prefoliation and contortion (Venkateswarlu 1978). Such, plants were termed
unichiral.

Isomerism, a fairly common mechanism in biological materials has attracted
the attention of several workers. This is exhibited in different characters like
prefoliation in leaves, contortion in floral parts, twining in stems and coiling of
shells in molluscs.

This phenomenon was widely used over the years by taxonomists, phylogene-
ticists and agronomists . Hutchinson(1964) used twisting of keel petals and styles
for taxonornic purposes. Smartt (1976) pointed out that the Asiatic species of
Phaseolus show clock-wise contortion of style and the American species show
counter clock-wise contortion. More recently, Jos and Venkateswarlu (1978)
used twining nature in Asian yams to describe their distribution. It was suggested
that the phylogeny of dextral types might be basically different from that of
sinistral types even among the Asian yams.

In palms (coconut, arecanut and toddy palm) the nature of leaf spiral determines
the yield. Right spiralled plants were found to be higher than their counterparts
(Davis 1972). Bahadur et al (1978) pro-bed into the causes resulting in higher
yield in right spiralled coconut. Their study revealed that the right spiralled
coconuts were physiologically superior to the left spiralled ones and therefore the
higher fruit production in right handers was correlated to their efficient metabolism.

In Dioscorea etsculenta, the levo plants were found to give more yield than the
dextro plants under artificially controlled conditions. But the vericais were better
than lefts Ijy 42-63% and the rights by 55*57% (Davis 1972).

22 T Vertkateswarlu

Table 2. Isomerism in prefoliation of main stem and the suckers in taro.

Nature of

No. of

Prefoliation in suckers

main stem

tubers

Levo

Dextro

Total

planted

Levo rotation

8

4

15

19

Dextro rotation

3

3

3

6

Total

11

7

18

25

Table 3. Isomerism in spathes of seedling families in taro.

SI. Cross particulars Levo Dextro Total

No.

1. 95LX

271 L

1

2

3

2. 271 L x

251 L

1

1

2

3. 271 L X

135 1,

4

3

7

4. 271 L X

268 D

3

3

6

5. 271 L X

211 D

1

2

3

6. 271 D X

Kovvur L

2

1

3

Total

12

12

24

Similar studies conducted in cocoyams did not show any convincing results.
Moreover, since this character is, not even transmitted vegetatively, the results will
not have any bearing on its utility as selection criterion.

At the moment, we are not in a position to understand the factors responsible
for the expression of levo, dextro, and bichiral nature in the prefoliation and con-
tortion of spathes in cocoyams. Recent study by Davis (1978) on Costus spetiosns9
suggests the involvement of one or more important laws governing plant growth
and. he hopes that a study on the orientation of leaf primordia at shoot apex and
their spiral descendance may help in unravelling the hidden law in shoot morpho-
genesis.

.'Similar study in cocoyams may also reveal some interesting facts responsible
for the mechanism as the new leaves and spadices arise from shoot apex.

Non-inheritance of isomerism in cocoyams 23

Acknowledgements

The author is highly grateful to Prof. T A Davis, FAO, Coconut Expert to
Indonesia and Dr J S Jos for the constructive comments on the manuscript. The
author is also thankful to Shri P K Thomas, Director of CTCRJ for providing
facilities and encouragement.

References

Bahadur B and Venkateswarlu T 197(>a Isomerism in flowers of four species of Jaiwp\\a\ /.

Indian Hot. Soc. 55 30-37

Bahadur B and Venkateswarlu T 1976b /. Indian Bot. Soc. 55 $9-94

Bahadur B, Madusudhana Rao M, Chandraiah M and Lokendar Rao K 197$ The physio-
logical basis of handedness in relation to yield in Cocos nucifera, L — A proposal ; Incomp.

Newslett. 9 108-111

Davis T A 1962 Non-inheritance of asymmetry in Cocos nucifera ; /. Genet. 58 42-50
Davis T A 1972 Effect of foliar arrangement on fruit production in some tropical crop plants ;

Tropical Ecology with an emphasis on organic production Athens (U.S.A.) pp. 147-164
Davis T A 1974 Enantiomorphic structures in plants ; Proc. Indian Nat. Set. Acad. B40 424-429
Davis T A 1978 Radial symmetry in Costus speciosus ; Phytomorphology 28 373-378
Hutchinson J 1964 The Genera of Flowering plants (Angiospermae) Dicotyledons (London :

Oxford University Press)
Jos J S and Venkateswarlu T 1978 Twining in relation to distribution among Asian yams;

J. Root Crops 4 63-64
Kasinov V S 1969 Inheritance of left and right handedness in Lemnaceae and other organisms ;

Genetika 5 22-29

Meyen S V 1973 Plant morphology in its nomethetical aspects ; Bot, Rev. 39 205-260
Smartt J 1976 Tropical pulses (London : Longmans)

Venkateswarlu T 197S Unichirality in Edible Aroids ; /. Root Crops 4 15-18
Venkateswarlu T and Hrishi N 1977 Bioisomerism in Edible Aroids ; /. Roof Crops 3 29-32

Proc. Indian Acad. Sci. (Plant Sci.), VoL 91, Number 1, February . 1988; -pp--' 25^35.
© Printed in India,

Apomixis in Ceitchms glaucus Mudaliar et Suudaraj

C SHANTHAMMA

Department of Post-Graduate Studies and Research in Botany, University of
Mysore, Manasagangatri, Mysore 570006, India

MS received 18 July 1980 ; revved 29 December 1JMI1

Abstract. A detailed cytoembryological investigation in Cenchrus gfaucus revealed,
that it is ail obligate apomict producing only aposporous embryosacs. Microsparo-.
genesis disclose chromosomal irregularities and mcgasporogeiiesis occasionally occur
and the development of sexual embryosac is completely absent. Otv the other hand,
aposporous initials develop into 4-nuclcatc cmbryosacs. Autonomous development
of tlie embryo is of common occurrence.

Keywords. Cenchms glaucus ; obligate ; apospory.

1. Introduction

Cenchrus glaucus Mudaliar et Sundaraj, a pentaploid perennial tropical species
belongs to the tribe Paniccac of Panicoideae. The occurrence of apomixts in this
genus was first investigated by Fisher et al (1954) in Pennisetum clitoris and Cenr
chrus setigerus. Later Snyder et al (1955) outlined a detailed study on the forma-
tion of aposporous embryosacs in C. ciliaris, a polymorphic facultative apornict.
Prelimiixary investigation has disclosed that another species of Cemchrus, C. glaucus
an aneuploid with 2/i = 45, where in microsporogeaesis is characterized by
abnormalities such as uaivalents, muJtivalents and lagging chromosomes, repro-
duces by gametophytic apomixis. The present study reports the results of cytoent.
bryological studies revealing the formation of only aposporous embryosac (obligate
apomict) and the complete absence of sexual embryosac.

2. Material and methods

Clones were collected from Agricultural College, Coimbatore and were grown in
the Departmental Botanical Garden, Manasagangotri, "University of Mysore,
Mysore. Identification of the material was confirmed by the Botanical Survey
of India, Coimbatore. A voucher specimen of the material is deposited in the
Herbarium, Department of Botany, University of Mysore, Mysore, India.

Florets at appropriate stages of development "were collected between 10 a.m.
and i p.m. and fixed in a mixture of 3 : 1 absolute alcohol.aavd acetic acid, later
on stored in 70% alcohol. Pollen mother cells were smeared in 2% acetocannine
for the study of meiosis. For mitotk studies, root tips squashes were Cdfule follow-

25

2f C Shartthamma

ing Tijo and Levan's (1950) technique. Embryological studies were carried out
following conventional methods of dehydration, infiltration and embedding.
Sections were cut at 10-44 microns in thickness and stained in Heidenhain's iron
alum haematoxylin.

3. Results

3*1. Mttrosporogenesis

During diakincsis and metaphase I, tetravalents, univalents in addition to biva-
lents are formed (figures I, 5). Chromosome number was confirmed in root-tip
(figure 11). Occasionally, a bridge and a fragment configuration indicating struc-
tural changes were seen at anaphase I (figure 2). Anaphase distribution is marked
by irregularities such as irregular distribution (figure 6), laggards (figure 7) and
micronuclei formation at dyad stage (figure 8). Second division irregularities
arc also pronounced with precocious movement of chromosomes at anaphase
(figure 9). Figure 3 reveals a dyad in division stages with 2 micronuclei in one

Figures 1-4. Microsporogcnesis. 1. diakinesis stage showing tetravalents, tri
valents, urtivaleats and bivalents, 2. anaphase I showing bridge fragment configu-
ration, 3. dyad in division showing single micronucleus at each pole in one of the
dyad cells, while in the other, one ratcroouicleus, 4. tetrad with chromosomal
material being left behind.

Apomlxis in Certchrus glaucus Mudaliar et Sundaraj

27

Figures 5-11. Microsporogenesis. 5. metaphase I with multivalents, bivalents
and umvaleiits, 6. anaphase with irregular distribution of chromosomes, 7. late
ariaphase with lagging chromosomes, 8. dyad showing micronuclei, 9. dyad in
division with micronucleus in one of the cells and also showing precocious movement
of chromosomes, 10. polyad formation, 11. somatic cell with 45 chromosomes
(magnification x 750).

Apontfxis in Certchrus glauaus Mudaliar et Sundaraj

29

(For figure captions sec page 35)

of the dyad cells, \vhile in the other a single micronucleus at one pole. On the
other hand, figure 4 reveals chromosomal material being left behind in a tetrad.
All these above irregularities lead to polyad formation (figure 10). Pollen sterility
is about 51%.

30 C Shanthamma

3*2. Megasporogeitesis and megagametogenesis

Megasporogcncsis begins with the differentiation of an hypodermal arehesporial
cell \\hich Junctions as the megasporo mother cell (figure 27) and proceeds through
the us ua I meiotic divisions. Figures 16 and 17 reveal degenerating dyad and tetrads.
In addition, several ovular sections showed the inception of the mciotic divisions
(figures 13, 14, 15), The presence of either sexual embryosacs or the stages leading
to their formation wcie not observed in any ovule.

3 • 3 . Aposporons embryosac

Aposporous initials numbering from 1 to 8 in most of the ovules differentiated
in the vicinity of rnegaspore mother cell (figures 12, 13, 14) or its meiotic products
(figures 15, 16, 17) within the nuccllus. In the early stages of development
aposporous initial which consists of a single nucleus located at one end of the
cell with a prominent vacuole developing below it (figures 18, 29) undergoes the
first mltotic division resulting in 2 nuclei and both the nuclei are located at the
same pole (figures 19, 28). The next division would result in a 4-nucleate embryo-
sac and all the 4-nuclei remaining at the same pole (figures 20, 30). Later on,
organization of the four nuclei takes place, 3 nuclei contribute to form an egg
apparatus of 2 synergids and an egg and a solitary polar (figures 21, 31).
However, synergid nuclei degenerate very early, leaving an embryosac with an
egg and a polar nucleus (figures 22, 32). Occasionally, the 4-nuclei without
organization undergo further divisions resulting in the formation of 6 to 8 nuclei
all grouped together (figure 25). The development of aposporous embryosac
may be simultaneous (figures 19, 22, 29) or may be non-simultaneous (iigures 20,
21, 31).

In addition to the aposporous cells 2 or 3 nuccllar epidermal, cells at the micro-
pylar end become conspicuous with dense cytoplasm and enlarged nuclei (figure 30).
In extreme cases aposporous smbryosacs are found at the attachment of the ovule
to the ovary wall (figure 32). The average value of embryosacs per ovule is indi-
cated in table 1.

3-4. Embryo and endosperm

The association of embryo and endosperm varies in different embryosacs of the
same ovule. Embryosacs containing well developed embryo and multinucieate
endosperm occur along with embryosacs containing * n imdivjdcd egg cell and a

Table 1. Average value of embryosacs per ovule.

No. of embryosacs per ovule

1234567

No. of of ovules examined 15 16 30 15 IS 16 10 10

Apomixis in Cenchrus glmuus Miulaliar cf Sumlar

31

'i-Wft.M.

**ev^..*ifSS

-Y

>*.

«S

}'*'*{*,

m ''ASi.

29?ft¥-.;75£.®» so V;. "& & y$JKS:: *

'

^'P0™^'81^' aposponms cmbryosac development and embryo
endosperm reIat,on,Wp, 27. MMC, 28. 2-binuclca.e aposporoas cmbryosac,
l o'rZr 6 ap°Sp0rOUS einb'-y° sacs' 30" 4-iwIcate aposporous cmbryosac
^ , , aposporou$ «"*ryosac, 32. aposporous cmbryosao developing at

n P°ti0n °f th° °VU°

X 150)

° "y0>

th°

many Ce"C<l cmbry° w;th
Sh°wing disturbed P°larily (magnification

Apomms in Cenchrus glauctts Mudaliar et Sundaraj 33

single polar nucleus in the same ovule (figure 23). Embryosacs containing embryo
with a single polar nucleus coexist with cmbryosaes containing undivided egg cell
with endosperm in the same ovule (figure 24). Sometimes embryos lack normal
polarity (figures 26, 34) and the polar nucleus is found lateral to the embryo.
Precocious development of the embryo is frequently encountered as in figure 33
and a polar nucleus is found towards one side. Although, the number of embryos
varies from 3 to 4, twin seedlings arc only 4%. Those seedlings ako reproduced
apomictically and triple seedlings were absent.

4. Discussion

Cettetmts gluucus is an obligate pcntaploid apomict with chromosome number
2n = 45. Microsporogenesis proceeds with chromosomal irregularities resembling
those found in Ccnclirus ciliaris (Fisher et at 1954) resulting in about 51% sterile
pollen. The presence of high percentage pollen viatility in C. glaucus may iridiocte
pseudogamous seed formation. The chromosomal abnormalities in this taxon
are similar to Pemiisetum setaceum, P. villosum (Narayan 1951) which arc also
obligate apomicts. According to Weimarck (1972) such aberrations in Hierochloe
species seem to be associated with the occurrence of apomixis.

Mcgasporogencsis sometimes occurs and embryosac formation is. rarely cnoouru
tered. Jn aposporous apomicts irregularity of meiosis at the time of megasporc
formation may affect the ability of the cmbryosac formation. Degeneration of
all the megaspores sometimes observed in C. glaucus may be due to irregular
meiosis as reported in Hieracium auraittiacum (Skalinska 1971) and in Poa praten*
sis (Qrazi et al 1961). However, 4-nucleate unreduced embryosac is of common
occurrence. On the other hand, the megaspore mother cell or its products degene-
rate autonomoulsy even before the origin of aposporous initials. Such autonomous
degeneration even before the origin of aposporous initials is reported in Fertilise turn
villosum, P. setaceum (Narayan 1951), Paspalum secans (Snyder 1957), Po a grant-
tica (Skalinska 1959), Heteropogon coittortus (Emery and Brown 1958), Sotfoiochlofl
schaemum (Brown and Emery 1957), Fenitisetum ciliare (Snyder et al 1955),
Puniciim maximum (Warmkc 1954) and in Boutdouva curtipertdula (Mohamed
and Gould 1966).

The presence of 4-nucleate aposporous embryo^ac in this genus was reported
first in Ceitchrus clliaris (Snyder et al 1955). The 4-nucleate embryosac develop*
ment has been described in detail in nearly 14 species in Poaccae. Brown aa\d
Emery (1958) have reported the presence of 4-nucleate aposporous cmbryosacs
in 43 of 153 species.

A striking feature of this species, however, is apart from embryosacs present
in the nucellar portion, embryosacs are ako found in such extra-nucellar places,
the broad base of the ovule where it is attached to the pericarp wall. Such nucellar
embryosacs have been reported in Pertnisetum villosum and P. sctaceum (Nafayan
1951), P. meziajtifm (Shaaithamma 1974j, Themeda iriandra (Brown and Emery
1957), Hierochloe odorata (Weimarck 1967), Hieracium wrantiacum (Skalinska
1971) and Hieracium pratense (Skalinska and Kubien 1972).

Embryos in C. glaucus arise exclusively from unreduced aposporous embryo-
sacs, Poiyembryony as studied by Norstog (1957) is high in Poaceae. Armstrong

P.CB)-3

34 C Shanthamma

(1W7) reported 42% polyembryony in a particular strain of Poa. Snyder et al (1955)
reported 20% of polyembryony in Peimlsetum etliare, but in Cemhrus glaucus only
4% of the seeds produced twin seedlings. (All the seedlings rcporduced apomicti-
cally. Triple seedlings were totally absent although in oviilar sections apparently
2 to 3 embryos per ovule were commonly observed). The low frequency of twin
seedling produced in C. glaucus may be attributed to the lack of endosperm for-
mation in cinbryosacs of a single ovule.

Acknowledgement

The author is deeply indebted to Dr K N Narayan for his guidance and encoura-
gement throughout the investigation.

References

Armstrong J M 1937 A cytological study of the genus Poa L. ; Can. f. Res. (Bot. Sci.) 15

281-297

Bar N L 1960 The grasses of Burma, Ceylon, India and Pakistan (London : Pcrgamun Press)
Brown W V and Emery W H P 1957 Apamixis in. the Gramincac, Tribe Andrapogoneuc ;

Themeda triandra and Botkr'wchloa ischaemiun ; Bot. Gaz. 118 246-253
Brown W V and Emery W H P 195$ Apamixis in the Gramineac, Panicoidcuc ; Am. J. Bot. 45

253-263
Emery W H P and Brown W V 195& Apomixis in the Gramincac , Tribe Andropogoneue :

Heteropogoii contonus ; Madrono 14 238-246
Fisher W D, Bashaw E C and Holt E C 1954 Evidence for apomixis in Pennisctum ciliare and

Cenchrus setigems ; Agrori. J. 46 401-404
Gildfitthuys P J and Brix K 1959 Apomixis in Pennisetiun dubiuni ; S. Afr. J. Ayrte. Sci. 2

231-245
Grazi F, Umaerus M and Akerber^ E 1961 Observations on the mode of reproduction and

embryology of Poa pm tens is ; Hereditas 47 4iJ9~541
Moharaed A H and Gould F H 1966 Biosystematic studies in the Boittdoua ctirtipeitdula complex

V., Megasporogenesis and embryosac development ; Am. J. Bot. 53 166-169
Mudaliar et Sundaraj 195& Cenchrus glaucus ; /. Boftibay Nat. Hist. Soc. 54 926
Narayan K N 1951 Cytogeitetic studies in apomictic Pennisetum species ; Ph.D. Thesis, University

California', Berkeley.

Norstog K J 1957 Polyembryony in Hierochloe odomta (L.) Beauv ; Ohio J. Sci. 57 315-320
Shanthamma C 1974 Studies in Poaceae Unpublished thesis
Skalinska M 1959 Embryological studies in Poa granitica Br. B1.9 an apomictie species of the

Carpathian range ; Acta Biol. Crac. Ser. Bot. 2 91-112
Skaiinska M 1971 Experimental and embryological studies in Hieradum auranliacum Z. ; Acta

Biol. Crac. Ser. Bot. 14 139-155
Skalinska M and Kubien 1972 Cytological and embryological studies in Hieracium pratcme

Tausch ; Acta Biol. Crac. Ser. Bot. 15 39-55
Snyder L A, Hernandez H E and Warmke H E 1955 The mechanism of apomixis in

Pennisetum ciliare ; Bot. Gaz. 116 209-221

Snyder L A 1957 Apomixis in Pasjpalum secans ; Am. J. Bot. 44 318-324
Tjio J H and Levan A 1950 The use of oxyquinoline in chromosome analysis ; Am. Estec.

Exp. Anta. Dei. 2 21-64
Warmke H E 1954 Apomixis in Panicum maximum ; Am. J. Bot. 41 5-11

Apomixis in Certchrus gluitcus Mudaliar ct Similar aj 35

Weimarck G 1967 Apomixis and sexuality in Hierochloe austral? s and in Swedish Hierochloe

odomta on different polyploid levels ; Bot. Not. 120 209-235
Weimarck G 1972 Male meiosis in some Ainphimictic and Apomiclic Hierochloe (Gramineae) ;

Bot. Not. 126 7-36

Figures 12-26. Mcga^porogcncsis, aposporous embryo-sac development, and embryo
and endosperm relationship. 12. LS of nuccllus showing megaspore mother cell
and 3 conspicuous nucellar aposponnis. cells (x 3000), 13. MMC at diakinesis
and 2 aposporous cells, ( x 3000), 14. metaphasc I of MMC with a single apos-
sporom coll initial (x 3000), 15. dyad and 3 prominent nucellar cells (x 3000),
16, 17- degenerated dyad and tetrad with nuccllar cells (adjoining) prominent
(x 2000), 18. a group of 4 aposporous (• mi nucleate) crnbryosacs (x 2000), 19. a
group of 3 aposporous cmbryosacs., 2 cmbryosacs at binuclcate stage and the other
one is at mctaphase, 20. 3 aposporous. cmbryosac?, one at 4-nucleate stage while
the third one at anaphase (x 2000), 21. a group of 3 aposporaus embryosacs. at
dilTercat stages of development and one of the embryosacs is at 4-nucleate stage
(x 1200), 22. 3 apo.sporo.us. embryosacs showing 2 degenerated synergids, an egg
and a polar nucleus ( X 2000), 23. two apo&porous embryosacs, one showing many
colled embryo and free nuclear endosperm and the other embryosac showing
an egg and a polar nucleus, (x 1200), 24. ovule showing 2 embryosacs, one with
many celled embryo and a single polar nucleus while the other embryosac with
a single egg cell and 4 endosperm nuclei (x 1200), 25. a group of 4-aposparous
embryosacs, 3 embryosacs arc binuclcate and the remaining one showing unorga-
nized 6-nucieate condition (x 1 200), 26. an aposporous embryosac with disturbed
polarity of embryo (x 1200).

Proc. Indian Acad. Sci. (Plant Sci.), Vol. 91, Ho. 1, February 1982, pp. 37-41.
(0) Printed in India.

Regeneration of plantlets from callus of Elettaria cardamonwm Mat on

N K SR1NI.VASA RAO, S NARAYANASWAMY*, E K CHACKO

and R DORE SWAMY

Division of Plant Physiology and Biochemistry, Indian Institute of Horticultural

Research. (ICAR), Bangalore 5(50080, India

*Emeritus Scientist, Council of Scientific and Industrial Research, New Delhi

MS received .3 November 1980 ; revised 1 October 1981

Abstract. Embryo callus and callus of rootstocks of in w'/ro-raigod seedlings of
Elettaria canlamomtwi were grown on MS medium supplemented with C\V -1-2,
4-D -1- BAP. Differentiation of Shoot buds, roots and leaves leading to tho develop-
ment of plantlets could be induced irt callus by withdrawing 2, 4-D or substituting
it by IAA or NAA in low concentrations.

Keywords. Callus culture ; cardamom ; regeneration.

1. Introduction

Reports on induction of shoot buds and whole plants from tissue cultures of both
monocotyledonous and dicotyledon or s plants have been numerous in ree en t years
as evident from the spate of publications or the subject (see reviews by Murashige
1974 ; Narayanas\vamy 1977). Clonal propagation through tissue culture has
been successful with many spice and condiment plants such as Foenicttlum vulgcrc
(Maheshwari and Gupta 1965), Anerhum graveolens (Ratnambs and Chopra 1974),
Carwn corvi (Ammirato 1974) and Capsicum annuum (Ounay and Rao 1978),
Spectacular rate of multiplication of turmeric (Curcuma longa) plants have been
reported in cultures of young vegetative buds isolated from the root stock (Nada-
guada et al 1978). This prompted us to investigate the potential for organogenesis
in tissue cultures of the cardamom (Elettaria car damomurn Maton of Zingiberaceae)
widely used as a condiment. This paper reports the successful regeneration of
shootbuds and plantlets from seedling callus of the herbaceous perennial species,

2. Methodology and results
2 • 1 . Seed germination

Dry seeds of cardamom were surface sterilised by 0-lJj mercuric chloride solution
to which a few drops of the detergent Teepol had been added. After washing

Contribution number 944 of Indian Institute of Horticultural Research.

37

38 N K Srinivasa Rao et al

thoroughly in sterilized water, the seeds were sown on White's (1963) nutri
agar. Slender seedlings -were obtained in three weeks on incubation at 26C
Whole seedlings bearing tke first sheathing leaf and plumule were transferred
Murashige and Skoog's (1962) medium (MS) to which auxins, cytokinins t
coconut water (CW) as specified (table 1) and sucrose (2%) had been added.
EDTA was used as the iron source. Each treatment comprised 12 replicai
Embryo callus was also obtained directly from seeds sown on the medium (8gur<
containing an auxin such as 2, 4-D (2,4-dichlorophyenoxy acetic acid).

2-2. Callus induction

Five-week-old 3 cm long seedlings bearing the first sheathing leaf and plum
were transferred to MS medium to which CW (18% v/v) H- 2, 4-D(2mg/l)
indole-3 -butyric acid (IB A 2mg/l) or naphthalene acetic acid (NAA 2mg/l) a
benzylaminopurine (BAP 2mg/l) had been added in combinations as need
Proliferation of cells from the rootstock was observed in 75% of the cultures res
ting in the formation of exuberant callus in 6 weeks after incubation (figinc
MS supplemented with CW (18% v/v) 4- 2, 4-D (2mg/l) + BAP (0-5mg/0 \
conducive for callus initiation and growth which couJd be augmented by the ad
tion of casein hydrolysate (CH, lgm/1.) in the medium. Neither yeast cxtr
250mg/l nor malt extract (250mg/l) proved favourable for callus growth; 2, 4
could, however, be replaced by IBA for callusing. Propicmocarmine squasl
of the proliferating callus showed cells of diverse sizes and shapes (figure
Trachcidal differentiation of cells was marked.

2-3. Regeneration of shoot buds

Callus growing on. MS + CW + 2, 4-D \vas subcultured on medium devoid
2, 4-D but containing 1AA. (2mg/l) or NAA (lmg/1). Three weeks after trans
green nodular structures developed in the callus indicating the initiation of organic
growth centres. CalJus grown on medium with higher concentrations of the an?
became friable and was not conducive for shoot bud induction. But induct!

Table 1. Responso of cardamom calli to growth regulators in vilro oit soqucn
transfer.

SI. Media composition Nature of

No. (Hormone concentrations in mg/1) response

1 . MS -I- CW (18% v/v) 4- 2, 4-D (2) Callusing good

2. MS -h CW(18% v/v) -f 2, 4-D (2) -1- CH(1g/l) -h BAP CaHusiag exuberant
(0-5)

3. MS -f CW(18% v/v) -f- lAAor IBA(l) -hNA\(2) Callus grew as vascular nodul

4. MS-hCW(18%v/v)4-lAA(l)^BAP(2) Shoot bud initiation in call

(80%)

5. MS 4-CW(tO% v/v) 4-BAP(2~5) -f-UA (1) 4-6 jjhoot buds per subcultu

Regeneration in Elettarla aardarnoinum

39

Figures 1-5. Callus induction and shoot bud regeneration in cardamom.
1. Calhtiiitg of seedlings in vitro on MS ~f GW (1 8 %v/v) 4- 2,4-D (2 mg/1) -{-BAP
(ft P5 mg/1) 6 weeks after incubation, X 1 *5. 2. Nodular callus derived from root
stock of seedling transferred to MS medium -h 1. AA (t mg/l) ~h NAA (2 mg/l)
x 1-5. 3. Propionocatmine squash preparation from root-stock callus piece
showing free cells and cell aggregates x 800. 4. Regeneration of a shoot bud
in primary callus grown on MS -f BAP (2 mg/l) + 1 AA (mg/l), x 1 • 5. 5. Cluster
of shoot buds regenerated from callus Subcultures ort MS 4- CW (10 %v/v) -I- BAP
(2mg/l)-HAA (I mg/l) x 1-5.

Regeneration m Elettaria cardamomum 41

scurrcd if MS medium was supplemented with CW (10% v/v) 4- BAP
1-5 mg/1) with or without the addition or 1AA and incubated for 6 weeks under
00 lux (figures 4,5). Four to six shoot buds could be obtained from each callus
ibeulture of uniform size, Rooting occurred at the base of individual shoot
uds on prolonged incubation in the same medium (aged cultures) or vrhen indivi-
ual shoottets were isolated and grown on White's medium to which NAA (2mg/l)
nd sucrose (1%) had been added.

» Discussion

.egeneration of plantlets in callus culture is an alternate means of propagation
i cardamom. Callus subcultures could develop 4-6 rcgcncnmts, each of which
as capable of rooting, when isolated and grown, forming a whole plant. Plant-
is propagated thus might not conform to parent genotype, .having been obtained
om seedling calii. Nevertheless, tissue culture provides a method by which a large
umber of strains could be obtained for selection of desirable variants. Also, it
tTers a method of rapid multiplication of elite varieties through multiple shoot
duction. Preliminary studies have shown that test tiibc plants could be success-
illy transferred to soil.

cknowledgements

he authors thank J>r G S Randhawa, the then Director, Indian Institute of
horticultural Research, Bangalore, for his keen interest and for providing faci-
:ics for tissue culture work. Thanks are also due to Dr I> Oundu Rao, Senior
ant Pathologist, for the supply of cardamom seeds.

ifcmicos

.mnirato I* V 1974 The affects of ab&igio ucid ott the development of Somatic embryos: from
colls of caraway (Canwt carvi L.) ; Bat. Gaz. 135 328-337

iitay A L and Rao P S 1978 In vitro regeneration from liypocotyl and cotyledon. ox?lanttf of
rod pepper (Capsicum annum) ; Plant Sci. Lett. 11 3(55-372

ahoshwari S C and Oupta R. P G 1965 Production of adventitious cmbryo'idtf in vitro from

stem callus of Foenicutuni vttlgare ; Planta 67 384-386

urasliigo T 1974 Plant propagation through tissue culture ; Ann. Rev, Plant Physio 1 . 25
135-166

urasbuge T and Skoog'F 1962 A revised medium for rapid growth and bioassays with tobacco"
tissue cultures ; Physhl. Plant. 15 473-497

idaguada R S, Mascarcnhas A F, Hendrc R R and Jaganathan V 1978 Rapid multipli-
cation of turmeric (Curcuma longa Iiirtit.), plants by tissue culture ; Indian /. Exp. Bio I. 16
120-122

urayanaswamy S 1977 Regei\cratioit of plants from tissue cultures; In Fundamental and Applied
Aspects of Plant Cell, Tissue ami Organ Culture (ed$.) J Reinort and Y P S Bajaj (Berlin :
Springer-Verlag) pp. 179-250

>tnamba S P and Chopra R N 1974 In vitro induction of embryoids from liypocotyl and
cotyledons of Anelhwn graveotens seedlings; Z. Pflanzenphysiol. 73 452-455

ute P R 1963 The Cultivation of Animal and Plant Cells (New York : Ronald Press)

Proc. Indian AcadL Set. (Plant Sci.)» Vol. 91, Number 1, February i982, pp. 43-53.
© Printed in India.

Studies in Cyperaceae : XVII Novelties in Fimbristytis (L.) Vahl and
their vegetative anatomy

EGOVINDARAJALU

Department of 'Botany, Presidency College, Madras 600005, India

MS' received 27 March 1 980

Abstract. T\vo novelties of Fimbristylis collected from Tamil Nadu (Madras Stale)
arc described and illustrated. One of them belong* to tli« section Fuscac and the
other one to Cyinosae. The vegetative anatomical characters of these two novelties
not only fall within1 the ambit of the range of anatomical variability of the genus
as a whole but their respective anatomical features in combination appear to be
characteristic and different from those of the species already known anatomically .

Keywords. Novelties \n Fimbristylis ; vegetative anatomy.

Fimbristylis scabnsquama Govind $/>. nov. — Sect Fuscac Ohwi (figure 1)

Peremis. Rhizoma btevisshna, Lignosa. Cu/mj 1-2, pentaquetri, valdo costati,
sulcati, glabri, rigidi, erecti, straminei-atrobrunnei, scabridi in dimidio superiore,
tecta ovato-lanceolatis sq.i\amis et denique fibro&is sq.uarnis, 15-25 cm x 0-6-
0-8 mm. Folia multa, spiraliter disposita, canaliculata cum marginc incurva
rccurva, brunnea-atrobrunnea, demxini evadentia nigra, eligiilata, aci\mir.aia
(acuta), incrassata et in margine dimidio superiore scabrida, culmo quam angus-
tiora, 10-15 cm x J-2 mm ; vaginae omnes lammiferae, glabrae, oblique trim-
catae, nitideac vel pallidae, briunae aliquando evadcn& nigtae, pi\\smimisve cor-
ncac. Inflorescentia simplex, contracta, capitata, consistens (2-) 3-8 (-10) spiculis,
l-2-l-8cm longa et hta. Bracteae anguste ovatae, aristatae, plusminusve
scabridae ia margine aristae, glabrae, inflorescentia multo breviores, 0- 5-1 -Ocm
longae. Radii primari O, raro cum-unus radius adest, triquetroi^s, laeves, usque
1cm longus, Spiculae elliptico ovatae, acutae, castaneo^bmnnae, plerumquc
fasicutatae, aliquando soiitariae, sessiles, subteretes, 10-16 florae, 6-7 x 2-3 mm^
Glumae late ovatae, acutae-stibacutae, distichae in dimidio inferiore vel plus
minusve per totum proprie oblique et lineariter scabridae in nudatis partibus,
anguste scariosae ad marginem, pleramque ciliolatae in dimidio superiore margine
cum lateribus inervis, cymbiformes, charatceae, nitidae, adpressae, mi\cronatae,
4-0-4'2 (incluso mucrone) x 3-0-3;2ram ; carina aliquando valida 3 (-5) nervia
ncrvis excurrentibus in mucronem ; mucro plerumque recurvata, 0*5 mm longa ;
nervis lateralis levior. Rhachilla pannose alata, excavata. Stamina 3 ; filainenta
longe flexuosa, tenues, tortuosa, 2-5-2'8min longa ; anthera brun^ea, apwiculata

43
P.(B)-4

44

E Govindarajalu

Figure la-g. Fimbmtylis scabrisquama Govind. sp. uov. a. habit x \\ b. style
and stigma x 11; c. spikelet x 5; d. glume, lateral view x .10; c. stamen
x 8; f. nut x 23; g. glume, spread out x 10; (from Govindarajalu,
12,110, type).

cum plus minusve apice setacea, Uneares, distincte calcarata adbasim, 1-9-2*0 mm
pnga. Stylus triqueter, vix dilatata ad basim, distincte fimbriata in superiore
tertia parte vel dimidia, 2«4-2-5 mm longus ; stigmata 3, brevis ciliata per totum§
stylo breviora, 1-7-1' 8 mm longa, Nux angusta oblongo ovata, triquetra, tricos-
tulata, alba stramiaea cum leviter lateralis convexa> stipitata, non vimbonulata
cum triangulf riter apice plana, aliquantulum aspera ob minutas transverse elon-
gates tuberculas in dimidio superiore, 0-6-0 -8 xO'3-0-4mm; cellulae epicar-
picae in dimidio superiore circulares (plus minusve hexagonales), minutae, reticu-
latae ; stipes 0-12^0- 15 mm longae.

Perennial. Rkizome short, woody. Claims 1-2, pentaquetrous, strongly ribbed,
sulcate, glabrous, rigid, erect, stramineous-dark brown, scabrid in upper half,

Novelties in Fimbristylis (L.) Vahl 45

covered by ovate-lanceolate scales and ultimately by the fibrous remains of the
scales, 15-25 cm x 0-6-0 -8 mm. Leaves many, spirally arranged, canaliculate
with incurved margin, recurved, brown-dark brown finally tending to become
black, cligulate, acuminate (acute), thickened and scabrid in upper half margin,
shorter than culms, 10-< 15 cm x 1-2 mm ; sheaths all laminiferous, glabrous,
obliquely truncate, shining or dull, brown sometimes becoming black, more or
less horny. Inflorescence simple, contracted, capitate consisting of (2-^) 3-8 (-10)
spikelets, 1-2^1 -&cm long and broad. Bracts narrowly ovate, aristatc, more or
less scabrid in the aristatc margin, glabrous, much shorter than inflorescence,
0-5-4'Ocm long. Primary rays O, rarely when 1 present triquetrous, smooth
up to 1 cm long. Spikelets elliptic ovate, acute, castaneous brown, usually clus-
tered, sometimes solitary, sessile, sub terete, 10-16 flowered, 6-7 x 2-3 mm.
Glumes broadly ovate, acutc-subacutc, distichous in lower half or more or less
throughout, characteristically obliquely and linearly scabrid in uncovered parts,
narrowlys carious at margin, ususlly ciliolate in upper half margin with nerve-
less sides, cymbiform, chartaceous, shining, adpressed, mucronate, 4 -0-4 -2 (inch
mucro) x 3 -0-3 -2 mm ; keel rather strong, 3 (-5) nerved, nerves excurrcnt into
mucro; mucro usually recurved, 0-5 mm long; lateral nerves rather faint.
Rhachllla raggedly winged, excavated. Stamens 3 ; filaments long, flexuous,
slender, tortuous, 2-5-2-8 mm long ; anther brown, apiculatc 'with somewhat
setaceous apex, linear, distinctly spurred at base, 1-9-2- Omm long. Style tri-
quetrous, hardly dilated at base, distinctly fimb riately hairy in upper 1/3 or 1/2,
2-4-2-5 mm long ; stigma 3, shortly hairy throughout, shorter than style, 1-7-
1-8 mm long. Nut narrowly oblong ovate, triquetrous, tricostulatc, whitish
stramineous with slightly convex sides, stipitate, non umbonulate with triangularly
flat apex, somewhat rough due to minute transversely elongated tubercles in upper
half, 0-6-0-8 x 0-3-0*4 mm; epicarpic cells in upper half circular (somewhat
hexagonal), minute, reticulate ; stipe 0*12-0- 15 mm long.

Govindarajalu 12,110, Venniyar toVaraiyattumottai, Highwayys Mts., Maciurai
Dt.,Tamilnadu occurring in open grassy slopes (type : PCM) ; Isotypcs: 12,1 10 A
(CAL) ; 12,110 B (MH) ; 12,110 C (PCM).

Related to F. eragrostis (Nees) Hance but differs in having culms covered
at the base by ovate lanceolate scales and ultimately by their fibrous remains,
much narrower canaliculate acuminate (acute) recurved leaves, simple contracted
capitate short inflorescence consisting of lesser number of spikelets, primary rays
when present triquetrous, clustered smaller spikelets, obliquely and linearly scabrid
glumes, longer recurved mucro, slender tortuous very long •'•Maminal filaments,
anthers with somewhat setulose apex, distinctly fimbriately hairy style, stigmas
shorter than style, narrowly oblong ovate triquetrous tricostulate non umbonu-
late smaller nuts with triangular flat apex and rough surface marked by somewhat
transversely elongated tubercles in upper half and reticulately arranged circular
epicarpic cells.

Note ,; Easily observable field and herbarium characters of this novelty are the
presence of ovate lanceolf te scales at the base of culms which are ultimately becom-
ing fibrous, involute canaliculate leaves recurving towards the soil with a tendency

46

E Govindarajalu

to become ultimately black, contracted capitate inflorescence and scabrid sub-
distichous-distichous glumes. Not common in the said locality. This is named
after its characteristic obliquely and linearly present scabrid condition of the
glumes.

Fimbristylis tortifolia Govind. sp. nov.— Sect Cymosae Ohwi— {figure 2)

Perennis. Rliizoma crassa, lignosa, horizontaliter vel oblique rcptans (interdum
verticaliter crescens) sine ullis obviis internodis. Culmi aliquantum tcnues, pcnta-
goai, incrassati et foliati ad basim, plerumque fbxuosi (erecti), solitari, lacvc*.,
glabri, costati, sulcati, dense tectis multis basibus foliatis et demum fibrosis fila-
mentis, plerumque curvati ad basim (15-) 20-35 cm x 0-75-1-0 mm. Folia

Figure 2a-h. Fimbristylis tortifolia Govind. sp. nov. a. habit x £; j>. styjc
and Stigma x 11 ; c. spikelet x 5; d. glume, lateral view x 6;'e. stamen
x 10; f. glume, spread out x 6; g. outer epicarpic cells, diagrammatic'
h. nut x 15; (from . Govindarajalu 12,000, type). *

Novelties in Fimbristylis (L.) Vahl 47

multa, ftexuosa, omnes recurva vel varie tortuosa vel saltern infima folia recurva,
glabra, canaliculata, involuta, longinque et aliquantulum scabrida, laevis ad in-
fimum dimidium marginem eligulata, leviter lata, acuta, culmo breviora, multi-
nervia, non catinata, 8-15 cm x 1-0-1 -5 mm. Vaginae brunnae, valde nerviae,
glabrae, omnes laminiferae, infimae denique evadentes filamenta fibrosa, non
corneae cum lateribus angustis rnembranaceis stramineis, oblique truncatae.
Irtflorescentia simplex, capitzta (raro cum uno radio addita), consister.s 3-8 (-10)
spiculis, 8-10 cm longa et lata. Bracteae late ovato lanceolatae, atrobrunneae,
late ad basim cum arista aliquantulum scabrida, rigidae, erectae-oblique erectae,
aequilongae vel leviter longiores, in (foreseen tia 8-15 mm longae. Radii O. Spiculae
ovato lanceolatae, acutae, teretes, castaneobrunaae, multiftorae, sessiles, 6-9 x
2-5-^3 'Omm. Olumae late ovatae, acutae, glabrae et laeves iid apicem, patens
cum distincte margins hyalina et cum lateribus uno nervis in dimidio una quoque,
eglandulosae, non nitidae, distincte mucronatae, 4-5-«5-5 (iticluso mucrone) x
4-0 mm ; carina distincta, 5 nervia, excurrentes in mucronem ; mucro recurvata
vel erecta, 0-6-0-7 mm longa. Rhachilla pannose alata, excavata. Stamita3;
anthera Unearo oblonga, lutea ad apicem rotundata, non setacea, ad basim cal-
carata (lobata), 0-75-0 -8 mm longa. Stylus triqueter cum leviter dilatata pyra-
midalibasi, glaber vel leviter fimbriatus post trifurcationem, 1-75-2-0 mm longus ;
stigmata 3, papilhta, stylo longiora (aequilonga), 2-25-2 -5 mm longa. Nux
late obovata triquetra, tricostulata cum lateribus convexis, atrobrunnea demum
evadens nigra, distincte stipitata, umbdnulata, distincte tuberculata in dimidio
superiore 1- 1-1-25 (inclusis stipitibus) x 0-75-0-8 mm ; stipes 0-1-0-2 mm
longus ; cellulae epicarpicae extimae in dimidio superiore transverse elongatae,
hexagonales, tessellatae, occurrentcs in 10-^12 regularibus verticalibus seriebus
in una quoqae facie.

Perennial. Rhizome thick, woody, horizontally or obliquely creeping (sometimes
vertically growing) without any obvious internodes. Culms rather slender, penta-
gonous, thickened and leafy at base, usually flexuous (erect), solitary, smooth,
glabrous, ribbed, sulcate, densely covered by many leaf bases and ultimately by
their fibrous strands, usually curved at base (15-*) 20-3 5 cm x 0-75-4 -Omm.
Leaves many, flexuous, all recurved or variously tortuous or at least the lowermost
leaves recurved, glabrous, canaliculate, involute, distinctly ard somewhat scabrid
in the margins, smooth in lower half margins, eligulate, slightly broad, acute, shor-
ter than culms, many nerved, non keeled, 8-15 cm x 1-0-1 -5 mm; Sheaths brown,
strongly nerved, glabrous, all laminiferous, lowermost ultimately becoming fibrous
strands, non horny with narrow membranous stramineous sides, obliquely trun-
cate. Inflorescence simple, capitate (rarely 1 ray added), consisting of 3-^8 (-40)
spikelets, 8-10 mm long and broad. Bracts broadly ovate lanceolate, dark brown,
broad at base with somewhat scabrid arista, stiff, erect-«obliqu,ely erect, as long as
or slightly longer than inflorescence, 8-15 mm long. Rays O. Spikelets ovate
lanceolate, acute, terete, castaneous brown, many flowered, sessile, 6-9 x 2*5-
3-0 mm. Glumes broadly ovate, acute, glabrous and smooth at apex, patent with
distinct hyaline margin and 1 lateral nerves in each half, eglandular, not shining,
distinctly mucronate, 4- 5-5- 5 (incl. mucro) x 4-0 ram; keel distinct, 5 nerved,
excurrent into mucro j mucro recurved or erect, 0'6-0*7 mm Itfng. Rhachilla

48 E Govindarajalu

raggedly winged, excavated. Stamens 3 ; anther linear, oblong, yellow, rounded
at apex without bristles, spurred at base (lobed), O'75-O' 8 mm long. Style trique-
trow; with slightly dilated pyramidal base, glabrous or slightly hairy behind tri-
furcation, 1*75-2- Omm long ;. stigma 3, papillose, (as long as) longer than style,
2 -25-2 -5 mm long. Nut broadly obovoid, triquetrous, tricostulate with convex
sides, dark brown ultimately tending to become black, distinctly stipitate, unibonu-
late, distinctly tubercled in upper half, 1-1-1 -25 (incl. stipe) x 0-75-0-8 mm ;
stipe 0-1-^0 -2 mm long ; outer epicarpic cells in upper half transversely elonga-
ted hexagonal, tessellated, occurring in UM2 regular vertical rows in each face.

Govindarajalu 12,000, Kulikad, Highwavys Mts., Madurai Dt., Tamilnadu,
commonly occurring in wet open grassy rocks (type : PCM) ; Isotypes : 12,000 A
(CAL) ; 12,000 B (MH) ; 12,000 C (BLAT) ;. 12,000 D (BSI- ; 12,000 E-V (PCM).

Related to F. rigidiuscula Govind. (sect. Cymosae Ohwi) but differs in having
usually flexuous culms usually covered at base ultimately by fibrous strands of
leaf bases, usually all or at least the lowermost leaves flexuous recurved or variou-
sly becoming tortuous, less scabrid or more or less smoooth margined leaves,
slightly broader non keeled many nerved acute leaves, non horny brown sheaths
with stramineous sides, ovate lanceolate spikelets, broadly ovate patent acute dis-
tinctly mucronate and distinctly hyaline margined glumes with 1 lateral nerve in
each half, long recurved or erect nxucro, shorter anthers with non bristly rounded
apex, shorter style, stigma usually longer than style, broadly obovoid triquetrous
tricostulate distinctly stipitate smaller nuts with distinct tubercles in Tipper half
and transversely elongated hexagonal epicarpic cells occurring in lesser number of
rows in each face.

Note : The following are the outstanding characters of this novelty that can
easily be recognized in the field as well as in the herbarium. The flexuous culms
are curving at the base and covered by fibrous strands of several leaf bases ; hori-
zontally or obliquely creeping rhizomes ; flexuous or variously tortuous non
keeled leaves with more or less smooth margins ; simple capitate inflorescence
consisting of 3-<6 ovate lanceolate spikelets. The name is based on the tortuous
condition of the leaves.

Vegetative anatomy

Materials and methods

The materials for the anatomical investigation were taken from their respective
isotypes. The methods that were followed in earlier works are adopted here
(Govindarajalu 1966, 1968 a,b, 1975 et seq). Most of the descriptive terms used
here are those that have been recommended by Metcalfe and Gregory (1964). The
typological terminologies proposed by Chaedle and Uhl (1948 a,b) for designa-
ting the types of vascular bundles and metaphloem are followed.

Fimbristylfs scabrisquama Govind

Lwf-^Abaxial surface : Intercostal cells moderately long, axially elongated ; cell
walls tbin, smooth ; end walls straight. Stomata (L. 45-49-5 /on ; W. 27*0-

Novelties in Fimbristylis (L.) Vahl 49

i-.S //m), elliptic oblong, thick-walled ; subsidiary cells low dome-shaped ; inter-
omatal. cells long with concave ends. Silica cells short, broad, thin-walled
rcumng in 3 continuous rows, each one of them containing 2-3 cone-shaped
[ica bodies surrounded by satellites.

Adaxial surface : Cells isodiametric, somewhat hexagonal ; cell walls thin,
nooth. Stomata wanting. Other details as in al axial surface.

T.S. lamina : Lamina examined 2 -2 mm wide. Outline *W* shaped, assymm-
ric£ I (figure 3,d). Keel inconspicuous. Margins dissimilar, one margin acute
id the other one somewhat rounded (figure 3,d). Cuticle uniformly thick,
alliform cells not differentiated. Adaxial epidermal cells uniform in size and
tape and larger than those of the abaxial. Sclerenchyma strands : adaxial 3
It. 72-90 jum ; W. 45-^54 /*m), pentagonal ; abaxial associated with each vl
It. and W. 54-90 /(m), rounded (puiviniform) ; submarginal strands (Ht. 3 6 /on;
r. 157-5 /*in), pulviniform. Adaxial hypodermis consisting of 2-3 layers of in-
ited radially elongated or rounded colorless perenchyma cells. Chlorenchyma
diating restricted to a axial one half of lamina. Vascular bundles c. 20 in
imber comprising large, medium and small vb's and not showing any regular
ternation with each other ; large vb's (type III A) oval in outline while medium
id small vb's rounded (type I) ; metaxylem vessel elements (D. 27 /«n in dia-
eter) ; metaphloem of 'regular type*. Bundle sheaths 3 layered ; M.S. in
I vb/s fibrous, complete ; in large vb's O.S. and I.S. parenchyinatous, the former
mplete \vhile the latter incomplete and in the case of small vb's both complete,
icumvasdular sclerenchyma absent. Tannin idioblasts common.

Ciilm--Epidermis9 surface view : Cells axially elongated with thin somewhat
tucms walls, Stomata (L. 31-5-36-0 /on ; W. 27-0-31-5 jum, more or less
borbicular, thick-called ; subsidiary cells dome-shaped ; in terstomatal cells long
th concave ends. Silica cells, short, rather narrow, thin-walled, occurring in
more or less continuous rows and each cell containing usurlly 2-3 cone-shaped
ica bodies surrounded by satellites. Silica particles very commonly and charac-
ristically present in cells in between the ribs and interstomatal cells ; likewise
unded silica bodies characteristically present in subsidiary cells (figure 3,f).

T.S. Culm; Long diameter of cv 1m examined 0-8 mm. -Outline subcircular
th ribs and furrows (figure 3,e). Cuticle uniformly thick (thickness 13 '5/jm)
roughout. Epidermal cells thin-walled, similar in size and shape except at the
PS. Stomata in line with epidermis ; guard cells with outer ledges only ; sub-
>matal chamber very narrow and small. Hypodermjs consisting of 3-4 (-5)
fers of radiating chlorenchyma cells, the continuity of which interrupted by jeri^
,eral ring of vb's (figure 3,e). Sclerenchyma strands (Ht. 45-0-67-5 /on j W/
•0-45*0 /«a), variable, pulviniform, rounded, triangular. Ground tissue made
> of Large parenchymatous cells having. intercellular spaces, vascular bundles
28 in number comprising I oth large and small vb's, arranged in two peripheral
igs, the former forming; the inner ring and the latter outer ring (figure 3te) ;
:ge vb's 6 in number, oval shaped (type HI B), the remainder rounded (type I) j,

50 E Govindarajatu

large vb's containing protoxylem lacunae ; metaxylem vessel elements (D.c.
22*5 //min diameter); metaphloern of Regular type'. Bundle sheaths, see
lamina. Circumvascular-selerenchyma little or rot developed. Tannin idioblasts
abundant in hypodermis.

Rhizome. Transverse section: Diameter of the rhizome examined 3 -2 mm.
Epidermal cells thick-walled, isodianntric. Cortex very broad, organized by
thick-walled parenchyma cells ; cells compactly arranged without intercellular
spaces and containing abundant starch. Subepidermal fibrous strands (Ht. 174-0-
290-0 /on ; W. 92-8-174-0 /on), triangular. Endodermoid layer made up of
single layer of cells with *U' shaped thickenings and forming a wavy limiting layer.
Stele central containing many vb's more or less rounded in outline, amphivasal
(type V); metaxylem elements (D. c. 9-0 urn in diameter). Central ground
tissue sim'.iar to that of cortex. Tannin idioblasts large, very common throughout
ground tissue.

Root. Transverse section : (figure 3,c). Diameter of the root examined 0-7 mm.
Exodermis single layered consisting of cells similar in size and shape with suberized
cell walls. Cortex : recognizable into 2 zones ; outer zone narrow consisting
of 4-5 layers of compactly arranged parenchyma cells ; inner broad, lacunose
due to concentrically arranged air-cavities ; air-cavities separated by radiating
row^ of parenchyma cells. Endodermis single layered consisting of cells with
*U' shaped thickenings and broad lumina. Pericycle of single layer of thick-walled
cells. Metaxylem vessel central, solitary, rounded in outline (D. c. 45-0 /on
in diameter). Protoxylem units 7-8 in number. Metaphloem not easily disting-
uishable. Central ground tissue parenchymatous.

Fimbristylis tortifolia Govind.

Lezf-Abaxial surface : Intercostal cells axially elongated, thin-walled, smooth 'j
end walls straight. Stomata (L. 45-54 /mi; W. 18-0-22-5 //m), narrowly
oblong ;. subsidiary cells low dome-shaped ; interstomatal cells short with concave
ends. Silica cells rather broad, short, moderately thick-walled, present in 2-3
continuous rows ; each cell possessing (1-) 2-3 cone-shaped silica bodies with
satellites.

Adaxial surface : Cells broad, isodiametric ; cellwalls thin, smooth, end walls
straight Stomata absent. Silica cells occurring in a single more or less dis-
ontinuous row ; other details see abaxialc surface.

T.S. lamina : Lamina examined 1-6 mm wide. Outline flatly triangular (fig-
ure 3, a), asymmetrical with abaxial ribs and furrows. Keel slightly developed,
somewhat rounded or bilobed (figure 3, a). Margins dissimilar; one margin
sloping towards abaxial side, the other more or less truncate with depression
(figure 3,a). Cuticle thick on either surface, Adaxial epidermal cells slightly
larger than those of the abaxial. Bulliform cells not distinctly differentiated, (fig-
ure 3,a). Sclerer.chyma strands : adaxial 5 in number (Ht. 40-5-45-0 //m;
W, 54-0-67*5 //in), pentangular ; abaxial strands .(Ht. and W. 36-0-49-5 /<m),.

Novelties In Fimbrisiylis (L.) Vahl

51

more or less rounded or pulviniform and associated with vb's (figure 3, a). Adaxial
hypodermis consisting of 4-5 layers of compactly arranged inflated translucent
polygonal parenchyma celts. Chlorenchyma restricted to abaxial one third of
lamina. Air-cavities absent except at substomatal cavities. Vascular bundles
18-20 in number of two different sizes out of which 3 large and the rest small ;
both belonging to type III A and arranged without any regular alternation ;
median vb. adaxially capped by radiating rows of large translucent parenchyma
cells (figure 3, a) ; metaxylem vessel elements (O. 18 /on in diameter) ; meta-
phloem of 'regular type '. Bundle sheaths in all vb's 3 layered ; O.S. parenchy-
ittatous, complete ; M.S. fibrous, complete ; I.S. parenchymatous., incomplete
in all vb's . Tannin idioblasts not observed.

T.C.

CU.

Figure 3 a f. a, b. Flmbristylis torti folia Oovind. a. transcction of lamina
x 44; b. transection of culm x 33 ; <?-/. F. scabrtequama Oovind. c. Transtection:
of root x 44; d, translection of lamina x 33; e. traitscction of culm
x 44; f. surface view of epidermal cells (note pressewce of silica bodies in
subsidiary cells and silica particles in epidermal and interstom^tal cells) x 226.
(All pasted on their respective isotypes).

52 E Govindarajalu

Culm. Epidermis, surface view : Cells axlally elongated ; cell walls thin, smooth,
Stoma.ta (L. 38-0 /mi ; W. 27-0-28-8 /(in), oblong elliptic ; subsidiary cells low,
dome-shaped ; interstomatal cells long with concave ends. Silica cells present
in a single discontinuous row ; cells long, and each one of them containing 5-6
cone-shaped silica bodies without satellites.

T.S. Culm : Long diameter of culm examined 1-2 mm. Outine polygonal or
somewhat subhexagonal with ribs and furrows (figure 3,b). Cuticle uniformly
thiclc (thickness c. 9 jj,m) throughout. Epidermal cells thin-v^alled, . similar ,
in.size-a,nd shape. Hypodermis consisting of 6-8 layers of non radiating chloren-:j
chyma cells interrupted by fibrous strands and vb's (figure 3,b). Air-cavities repre-;j
sented by substomatal spaces. Sclerenchyma strands (Ht. 54-0-112-5 //m ;
W. 99*0-135-0 /zm),. usually pulviniform (triangular ovate). Ground tissue con-
sisting of large parenchyma cells arranged \vith intercellular spaces. Vascular
bundles c. 43 in number comprising both large (type III A) and small vVs (type I)
large vb's 7 in number somewhat oval or subcircular in outline containing pro-
toxylem lacunae ; small vb's circular in outline and arranged peripherally in the
form of undulating ring following the contour of the culm while large vb's
forming the inner ring (figure 3,b) j metaxylem vessel elements (D. 18-0-22-5/zm
in diameter); metaphloem of 'regular type'. Bundle sheaths in all vb's 3
layered ; O.S. and I.S. parenchymatous, the former complete and the latter in-
complete ; M.S. fibrous, complete in all vb's. ;;••/

Root. Transverse section : Diameter of the root examined 0-6 him; Exodermis :
single layered consisting of cells uniform in size and shape ; cell' walls suberized,
Cortex : recognizable into two zones, the outer zone narrow consisting of Slayers
of compactly arranged thick-walled parenchyma cells ; inner cortex rather broad,
lacunose ; air-cavities concentrically arranged and separated by radiating rows
of parenchyma. Endodermis of single layer made up of slightly tangentially
elongated cells with *U' shaped thickenings. Pericycle consisting of a single
layer of thick-walled cells. Metaxylem vessel solitary, central, circular in outline,
thick-walled (D. 67-5 /jm in diameter) ; protoxylem units 8 in number alternating
with as many metaphloem units ; each unit of the mstaphloem consisting of a
group of 4 cells of which 2 being large sieve tube elements and 2 companion
cells. Ground tissue made up of thick-walled parenchyma.

References

Cheadle V I and Uhl N W 1948a Types of .vascular bundles in the Monocotyledoneae and
their relations to the late metaxylem conducting elements ; Am. J. Bat. 35 486-496

Cheadle V I and Uhl N W 1948b The relation of metaphloem to the types of vascular bundles
in the Monocotyledons; Am. J. Bot. 35 578-583

Govindarajalu E 1966 Systematic anatomy of South Indian Cyperaceae : Bulbostylis Kunth
/. Linn, Soc. (Bot.) 59 289-304

Govindarajalu E 1968a Systematic anatomy of south Indian Cyperaceae: Fuirena Rottb.; /,
Linn. Soc. (Bot.) 62 27-40

Novelties in Fimbristylis (L.) Vahl

53

Govindarajalu E 1968b Systematic anatomy of south Indian Cyperaceae: Cyperus L. subgen.
Kyllinga (Rottb.) Suringar; /. Linn. Soc. (Bot.) 62 41-58

Govindarajalu E 1975 The systematic anatomy of South Indian Cyperacene: Eleocharis R.Br.,
Rhynchospora Vahl and Selena Berg.; Adansonia 14 581-632

Metcalfe C R and Gregory M 1964 Some new descriptive terms for Cyperaceae with a dis-
cussion1 of variations in leaf form noted in the family; Notes Jodrell Lab. I 1-11

Key to figure lettering and text abbreviations

A.B.E. abaxial epidermis:

A.C. air-cavity

AD.E. adaxial epidermis

CH. chlorenchyrna

CU. cuticle

E. epidermis
B.C. epidermal cell
EN. encloderrris
EX. exodermis

F. fibres

G.T. ground tissue

I.C. interstomatal cell

1,CO. inner cortex

M.V. metaxylem vessel

O.C. outer cortex

PH. metaphloem

P.I*. protoxylem lacuna

P,X. protoxylem

R.P. radiating parenchyma

S. sclerenchyma

S.B. silica body

S,P. silica particle

S.S. sclcrenchyma strand

ST. sttoma

T.C. translucent cell

T.i. tannin idioblast

V.B. vascular bundle

XY. metaxylem

Text abbreviations :

c. circa

D, diameter

Ht. height

l.S. inner sheath

1'. length

M.S. middle sheath

O,S. outer sheath

vb*s vascular bundles (sing, vb.)

W. width

Procf. Indian Acad. Sci. (Plant Sci.), Vol. 91, Number I, February 1982, pp. 55-60.
© Printed in India.

Embryological studies in three species of Cymbopogon Spreng
(Poaceae)

S P CHODA, HARSH MITTER and RAVINDER K BHANWRA

Department of Botany, Panjab University, Chandigarh 160 014, India

MS received 17 September 1980 ; revised 24 September 1981

Abstract. The embryology of Cymbopogon nardua var. confetti floras, C. martinii
var. Motia and C. par kef i has been studied. C. /utrdus and C. martinii have been
observed to be seed-sterile owing to failure of fertilization while in C. parkeri, the
seed-setting is only about 8-00%.

Keywords. Seed-sterility ; Cymbopogon ; Andropogoneac ; Poaccac.

1. Introduction

The genus Cymbopogon belongs to the tribe Andropogoneae of the subfamily
Panicoideac and it is represented by 24 species in the Indian sub-continent (Bor
1960). A number of species of Cymbopogon yield essential oils and are used in
perfumery. The embryology of C. mart mil and C. nervatus has been worked out
by Brown and Emery (1958) although of a preliminary nature. The present paper
deals with the embryology of Cymbopogon nardus (L.) Rendle var. Confertiflorus
(Steud.) Stapf ex Bor, C. martinii (Roxb.) Wats var. Motia and C. parkeri Stapf,
to find the nature of seed sterility observed in different species.

2. Materials and methods

The material of C. nardus and C. martinii was collected from the Botanical Gardens
of the Panjab University while that of C. parkeri from Shiwalik hills in the months
August to November. Conventional methods of dehydration and embedding
were used. The sections were cut at 5-10 /an and stained with safranin and fast
green. For studying the growth of the pollen tube in the style, the method given
by Khoshoo and Vij (1963) has been followed. For this purpose the gynoecia were
fixed in 1 : 3 acetic alcohol, after 3, 6, 9 and 24 hrs of pollination. After half an
hour the styles were transferred . to 30% ethyl alcohol for preservation. The
ovaries were transferred to 1 % solution of acid fuchsin for- about 20 min. Subse-
quently, the styles were cleared in lactic acid at 60° C and the whole mounts
were made in pure lactic acid.

55

56 S P Choda, Harsh Mitter and Ravftider K Shanwra

3. Observations and discussion

3.1. Microspofogenesis and male gametophyte

Anthers are tetrasporangiate (figure 1) and the anther wall consists of an epidermis,
followed on the inner side by the endothecial layer, a single middle layer and the
tapetum (figures 2, 3). The epidermal cells in mature anthers show deposition of
oil droplets in C. nardus and endothecial cells develop fibrous thickenings
(figure 10). The middle layer is ephemeral. The tapetum is of the glandular
type and its cells become binucleate in C. nardus while they remain uninucleatc
in C. martinii and C. parked. As seen in transections the MMC are disposed in
4 or 5 rows (figures 2, 3). Meiosis in MMC is normal and it is of the successive
type leading to the formation of isobilateral microspore tetrads (figures 4-7). The
development of the male gametophyte occurs as described in other members of
the family and the pollen grains are shed at the 3-celled stage (figures 8, 9). The
pollen grains are monocolpate with a thick smooth exine and slightly thinner
intine. The pollen fertility is about 90% in C. nardus and C. parked and about
70% in C. martinii.

3.2. Ovary and ovule

The ovary contains a sessile, bitegmic, pseudocfassinucellar hemianatropous ovule
(figures 16, 17). The inner integument is composed of cells 2-3 layers thick but
it usually fails to grow over the nucellus so that a definite micropyle is lacking
(figure 16). In Pennisetwn typhoideum (Narayanaswami 1953) and Capillipediutn
huegelii (Choda and Bhanwra 1980) the inner integument does not grow over the
nucellus but in most other species of the family the micropyle is formed by the
inner integument (Narayanaswami 1954, 1955, 1956). In Cymbopogon parkeri
where some seed-setting is observed, the inner integument is found to degenerate
after fertilization unlike that in Saccharum offidnarum (Artschwager et al 1929)
and Sorghum vulgar e (Artschwagei? and McGuire 1949), belonging to Andropogo-
neae where they persist in mature caryopsis.

The outer integument on the upper side of the ovule is 2-3 cell layered but it
covers only about a third of the ovule (figure 17), a feature characteristic of the
subfamily Panicoideae (Chandra 1963 ; Venkateswarlu and Devi 1964). The
outer integument on the lower side of the 6vule shows about the same growth as
the inner integument and is 2-5 cell layered (figures 16-18).

The nucellar epidermis undergoes 2-3 periclinal divisions in the region of the
micropyle (figures 16, 18) by the time the megaspore tetrads are formed. In
C. nardus, the nucellar cells in this region become conspicuous owing to their large
size, dense contents and prominent nuclei (figure 16). The formation of a parietal
tissue due to periclinal divisions in the nucellar epidermis near the micropyle has
been reported in many species belonging to the subfamily Panicoideae (Chandra
1963 ; Venkateswarlu and Devi 1964). The nucellar tissue is absorbed by the
developing embryo and endosperm and is hardly traceable in the mature caryopsis
of C. parkeri (figures 26, 27).

Embryology of Cymbdpogon Spreng. (Paaceae)

'•51

19

Figures 1-19. Microsporangium, microsporogenesis, male gametophyte, megasporo-
genesis and female gametophyte. 1, 3, 11, 16-19. Cymbopogon ncrdus. 2, 12,
14, 15. C. maninii. 4-10, 13. C. parkeri. 1. TS of the anther, 2. TS anther
lobe showing periclinal division in the inner secondary parietal layer, 3. TS anther
lobe showing wall layers and sporogenous cells, 4-7. Stages in microsporogenesis,
8, 9. 2- and 3 -celled pollen grains respectively, 10. Portion of wall layers showing
epidermis and fibrous thickenings of the endotheciura, 11. LS ovule primordium
at archesporial cells stage, 12. LS ovule at megaspore mother cell stage, 13, 14.
Megasporogenesis, 15, 16. 2- and 4-nucleate embryo sac stages respectively,
17. VS of the ovary and ovule, 18. Embryo sac showing egg cell, two synergids,
two polar nuclei and antipodal complex of several cells, 19. Embryo sac showing
egg apparatus, polar nuclei and degenerating antipodal cells. (See explanation of
abbreviation in p. 58).

58

S P Choda, Harsh Uitier aitd Ravinder K Bhaitwra

Figures 20-27. Post-pollination development. 20, 22. C. tfd/Y/ws ; 21, 23-27-

C. parkcri. 20-22. Degenerating embryo sac and shrivelled ovule ; 23-27.
Some stages in caryopsis development.

ANT— antipodal ; CO— co4eoptilc ; COZ— coleorhiza ", E— egg ; EL—embryomc

leaf ; END— endodermis ; EPI— epidermis ; IP— inner parietal layer ; MES-—

mesocotylc ; PN— polar nucleus ; R— radicle ; RC— root cap ; S— -sporogcnous
tissue ; SO-scutelluni ; SYN— synergid.

3.3. Embryo sac development

The single hypodermal archesporial cell is differentiated in the nucellus, which
increases in size and functions as the megaspore mother cell (figures 11, 12), It
divides meiotically so as to form a linear tetrad of megaspores (figure 13). In
C martinii^ however, the upper dyad cell degenerates without undergoing division
(figure 14). The chalazal megaspore functions and develops into the polygonum
type of embryo sac having aft egg cell, two synergids, a central cell with its two

Embryology of Cymbopogon Spreng. (Poaceae) 5J>

polar nuclei and three antipodal cells. The latter proliferate further and form
9-26 cells in C. nardus, 12-15 cells in C maninll and 9-16 cells in C. parkerl The
multiplication of the 3 antipodal cells is commonly reported in grasses
(Venkateswarlu and Devi 1964 ; Maze and Bohm 1973).

In C. nardus and C. martini!, the pollen grains germinate on the stigrnatic hairs
but the pollen tubes fail to reach the embryo sac due to some unknown factor.
The embryo sac and the ovule eventually shrivel and undergo disintegration
(figures 20-22). Occasionally the ovules become enlarged and they contain endo-
sperm nuclei formed probably due to autonomous divisions of the secondary
nucleus, but there is no embryo formation. In C. parkeri, however, about 8%
seed-set has been noticed. The primary endosperm nucleus starts dividing earlier
than the zygote. The endosperm is of the nuclear type (figure 23) as is reported
in other grasses. The endosperm becomes completely cellular at globular stage
of the proembryo. Figures 23-27 show some of the stages in the development
of embryo in the species. The sequence of early development of the embryo could
not be traced but the structure of the mature embryo is similar to that described
by Reeder (1957) in other members of the tribe Andropogoneae. Seed-sterility
in family Poaceae has been previously reported in Helaria belangeri and H. mutica
by Brown and Coe (1951) and in Digitaria decumbens by Sheth et al (1956).

In H. belangeri and H. mutica, the degeneration of the female gametophyte may
occur any time after megaspore formation and this has been suggested to be the
cause of seed sterility. In the present species, however, the degeneration of the
female gametophyte occurs only after it has attained maturity.

Poor seed-set as in C. parkeri and complete sterility as exhibited by C. nardus
and C. martinii appear to be compensated by the predominance of vegetative
propagation as a means of survival ; consequent on perturbation of sexuality.

References

Artschwager E, Brandos E W and Starrett R C 1929 Development of flower and seed of some

varieties of sugarcane ; /. Agric. Res. 39 1-30
Artschwager E and McGuire RC 1949 Cytology of reproduction in Sorghum vulgare ; J. Agric.

Res. 78 659-673
Bor N L 1960 Grasses of Burma, Ceylon, India and Pakistan (Excluding Bambuseae) ; (London,

New York, Paris : Pergamon Press)
Brown W V and Coe G E 1951 A study of sterility in Helaria belangeri (Stcud.) Nash and

H. mutica (Buckl.) Benth ; Am. J. Bat. 38 823-830
Brown W V and Emery W H P .1958 Apomixis in the Gramineae : Panicoideae ; Am. J. Bot.

45 253-263

Chandra N 1963 Som-3 ovule characters in the systeraatics of Gramineae ; Curr. Sci. 32 271-279
Choda S P and Bhanwra R K 1980 Apomixis in Capillipedium huegelll (Hack.) Stapf

(Gramineae) ; Proc. Indian Natl. Sci. Acad. 46 572-578
Khoshoo T N and Vij S P 1963 Biosystematics of Citmttus vulgaris var fislulosus ; Caryologia

16 541-552
Maze J and Bohrn L R 1973 Comparative embryology of Stijpa elmeri (Gramineae) ; CanJ.

Bot. 51 235-247
Narayanaswami S 1953 The structure and development of the caryopsis in some Indian millets.

1. Pennisetum typhoidewn ; Phytomorphology 3 98-112

60 S P Choda, Harsh Mitter and Ravinder K Bhanwra

Narayana-swami S 1954 The structure and development of the caryopsis in some Indian millets.

2. Plaspalum scrobiculatum ; Bull. Torrey Bot. Club. 81 288-299
Narayanaswami S 1955 The structure and development of the caryopsis in some Indian millets.

4. Echinochloa frumentaceae ; Phytomorphology 5 161-171
Narayanaswami S 1956 The structure and development of the caryopsis in some Indian millets.

6. Setaria itdica\ Bot. Gaz. 118 112-122

Reeder J R 1957 The grass embryo in systematics ; Am. J. Bot. 44 756-768
Sheth A A, Yu L and Edwardson J 1956 Sterility in pangola grass Digitaria decumbens ; Agron.

J. 48 505-507
Venkateswarlu J and Devi P I 1964 Embryology of some Indian grasses ; Curr. Sci. 33 104-106

Proc. Indian Acad. Sci. (Plant Sci.), Vol. 91, Number 1, February 1982, pp. 61-tfg.
© Printed in India,

Reproductive efficiency of secondary successional herbaceous
populations subsequent to slash and burn of sub-tropical humid
forests in north-eastern India

K G SAXENA and P S RAMAKRISHNAN

Department of Botany, School of Life Sciences, North Eastern Hill University,
Shi Hong 793014, India

MS received 10 January 19&1 ; revised 5 December 1981

Abstract. Three categories of secondary successional herbaceous communities
subsequent, to slash and burn, viz., early success-ional non-sprouting, early succes-
sional sprouting and late suceessional populations were investigated for tlieir
reproductive efficiency considering leaf component since it is the chief organ of
photosynthesis. Early successional non-sprouting populations were found to be
rcproductively the most efficient whereas the early successional sprouting popu-
lations allocated more to vegetatively reproducing organs. While the high reproduc-
tive potential of early successional non-sprouting species was. associated with vigour
and production efficiency of the species, this relationship was stronger with the
latter characteristic. On the other hand, early successional sprouting populations
showed inverse relationship between vegetative and sexual reproductive effort. The
strategy of late successional species seems to be to maximize vegetative growth in a
closed habitat. The significance of these strategies is discussed in the paper.

Keywords. Growth strategies ; leaf area ratio ; reproductive effort ;
communities ; adaptation.

1, Introduction

Slash, and barn agriculture, locally known as s Jhum ', is the most prevalent form of
cropping in the hill regions of north-eastern India. The early successional herba-
ceous communities constitute an important phase in the fallow development during
secondary succession subsequent to .cropping. This community, which holds the
ground for about 5-6 years, often gets arrested at this stage due to the shortening
of the Jhum cycle, the intervening period before the cropping is done on the same
site (Ramakrishnan et al 1981).

Optimization of reproductive output in plants is attained through a favourable
partitioning of the available resources for various life-activities such as mainte-
nance, growth and reproduction (Abrahamson and Gadgil 1973). Much is known
about the resource allocation pattern in relation to the reproductive strategy of
different plant species in an attempt to explain the ecological success of a species
in a given environment (Harper and Ogden 1970; Ogden 1974; Abrahamson
1975, 1979 ; Newell and Tramer 1978). While such an approach has yielded

61

62

K G Saxena and P S Rantakrishnart

valuable information, little effort has been made to relate the reproductive gfowff i
strategy with the leaf growth (McNaughtoa 1975; Bazzaz and Harper 1977 ;
Primack 1979). This approach for evaluating the reproductive strategy of plants
is more relevant because, leaf as an organ is the chief region of photosynthetic
activity. In the present paper, early successional non-sprouting and sprouting,
and late successional herbaceous populations have been compared for their growth
and reproductive characteristics considering leaf as the sole organ responsible for
energy capture and its overall distribution. The non-sprouting species, obviously,
are all established through seeds alone. Sprouting species, though they may also
come through seeds, are those that are established through sprouts alone.

2. Study area and climate

The study was earned out in Burnihat (26° N latitude and 91-5°E longitude) in
the Khasi Hills about 90 km north of Shillong, on precambrian rocks which are
represented by gneiss, schists and granites. The soil is red sandy loam and of
laterite origin. The pH ranges from 5 to 7. The angle of slope generally ranges
from 20° to 40°.

Climatically the year can be divided into three distinct seasons ; the dry summer
runs from mid-February to May and the rainy season extends from May to
September with an annual rainfall of 2200 mm. The latter is a warm period with
high humidity. The mild winter which is practically rainless except for a few
showers, extends from November to February. The annual maximum and
miaimum temperatures are 33° C and 7° C respectively (figure 1).

3. Methods of study

Four fallows which were slashed in January 1978 and freshly burnt in Mar&h
1978 and two 40 year old forested fallows were selected for this study. While

0 maximum temperature
* minimum temperature

A rainfall

500

400

300

200

100

0

2

Figure

JFMAMJJASONO

Time period (months )
1. Ombrothermic diagram of the study area (1978).

Sncaesslortal herbaceous communities 63

mixed cropping is done normally at least for one year (Ramakrishnan et al 1981),
for the purpose of the present study, the site was left directly as a fallow after
the burn instead of being cultivated. The fire was a high intensity burn, since
the slash burnt was derived from a 20 year regtowth. For each herbaceous
species, phonological observations were made throughout the year using three
permanent quadrats (50 cm x 50 cm) which were harvested at the fruiting stage.
Those species which did not flower, were harvested at the end of the growing
season. The samples were taken from uniformly monospecific patches of each
species in order to minimize the variation due to micro-environment. Sample
size ranged from 7 to 25 individuals. Below ground parts were carefully
washed and leaf blade and seed components were detached. In situations where
senescence started at flowering or fruiting stage, the fallen leaves and fruits or
seeds were also included. Different components were dried at 80 ± 5°C in a
hot air oven, for 48 hours and then weighed.

Leaf area (by planimeter) and leaf dry weight were estimated using three repli-
cates ith 20 leaves per replicate. Total leaf area per plant was obtained by
dividing total leaf biomass by dry weight per unit leaf area. Leaf area ratio was
calculated as leaf area (cm2) per unit (g) biomass.

Density values for different species were estimated by using 50 randomly placed
1m2 quadrats in each fallow (Misra 1968). Twenty quadrats at random were
harvested for estimating the average biomass per plant. This alongwith the
density values were used for calculating biomass per m2. Each shoot was consi-
dered as a separate individual, in the case of the rhizomatous species. Community
analysis was done at the end of the growing season.

4. Results

Table 1 shows the density and biomass values of different species in the early and
late successional communities, Amongst the early successional annuals, Engeron
linifolius was numerically the most dominant followed by Panicum maximum and
Cassia tora. However, C. tora having the lowest density, contributed maximum
to the herbaceous biomass. Six other annuals were present in a small proportion
and therefore are considered together. Amongst the early successional non-
sprouting perennials, Eupatorium odoratum was the most dominant. About 64%
of the herbaceous biomass was contributed by the sprouting species in the early
successionai communities. Thysanolaena maxima, though having higher relative
density than Saccharum arundinaceum, contributed lesser in terms of biomass
compared to the latter. In the late successional herbaceous communities,
Oplisrnenus compositus was the most dominant component.

'Species like Engeron linifolius, Eupatorium odoratum, Saccharum amndinaceum
and Thysanolaena maxima which were the most dominant component in the early
successional communities, had lower leaf area ratio compared to the less frequent
s pecies like Euphbrbia hirta, Borreria articularis, Digitaria adscendens and Mimosa
pudica. Leaf area ratio of the late successional species was, generally much
higher than the early successional species. Even £. odoratum and JP. maximum*,
which are common in the early and late successional stages, exhibited higher leaf
r&io in the l^e successiowal communities. Reproductive effort which

64 KG Saxena and P S Ramakrishnan

Table 1. Density and biomass of different species in tlie early and late successional
herbaceous communities.

Density Relative Biomass Biomass

, (individuals/ density (%) (g/m2) contribution
m2) (JQ

Early successional non-sprouting
populations :

Annuals :

Erigeron linifoliua 3-45 10-11 6-38 0-19

Panicum maximum 1-96 5-75 13-52 0-40

Cassia tora 1-10 3-22 16-61 0-49

Others (TJ = 6) 2-40 7-05 7-05 0-50

Perennials :

Eupatorium odaratum 3-75 10-99 1087-35 32-03

Panicum khasianum 2-45 7-18 15-44 0-45

Others (n « 4) 1-60 5-57 6-35 0-.19

Early successional sprouting
populations

Thysanolaena maxima 9-05 26-53 728-53 21-46

Saccharum arundinaceum 6-20 18-18 1305-10 38-45

Imperata cylindrica 1»50 4-40 87 -38 2-58

Others (/* = 2) 0-35 1-02 111-61 3-26

Late successional populations

Annuals :

Panicum maximum

5-62

16-11

3-37

3-59

Oryza granulata

1-85

5-30

12-21

13-02

Perennials :

Oplismenus compositus

21-46

61-53

27-84'

28-82

Centotheca lappacea

2-35

6-74

11-16

11-90

Others (# = 7)

3-60

10-32

40-07

42-67

n is the number of species,

worked out in relation to leaf growth (seed (mg)/10 cm2 leaf) indicates much higher
values for species like E. odoratum and E. linifolius in early successional commu-
nities. Amongrt the early successional sprouting perennials, Imperata cylindrica
and Grewia elastica did not flower in the first post fire year and M. pudica had
much higher reproductive effort than the others, The late successional species,
on the other hand, had comparatively very low values for reproductive effort ;
in Hedychium coccineurn and Curculigo recwvata flowering was not observed
during the year of study (figure 2).

Succession^ herbaceous communities

65

200

100

20

O

?

? 40

o

UJ

ft GO

Figure 2. Leaf area ratio and reproductive effort (seed mg/10 cm2 leaf) of secon-
dary successional herbaceous populations. Non-sprouting early successional
populations, (a) Annuals, (b) perennials, (c) sprouting early succes&ional perennials,
late successional populations, (d) annuals, (e) perennials. From left to right the
different columns are : Erigeron limifolius Willd., Rottboelia goalparensis Bor.,
Cassia torn L., Crossocephalum crepidioides (Benth.) S., Brachiaris distachya (L.)
Stapf., Panicum maximum Facq. , Euphorbia hirta L.,Borreira articular is (L.f.) Wild.,
Mollugc stricta L., Eupatorium cdoratum L,, Setaria palmifolia (Koen.) Stapf..
Paspatidium putictatum (Burm.) A. Camus., Panicum khasianum Munro., Manisuria
granularis L.f., Digitaria adscenderts (H,B,K.) Henr., Saccharum arundinaceum
Hook f., Thysanolaena tnaxima (Roxb.) O. Ketze., Grewia elastica Royle
Irnperata cylindrica Beauv,, Mimosa pudica L., Oryza granufata Nees et Arn.
Panicum maximum Facq., Rumex itepatensis Spreng., Eupatorium odoratum L.
Hedychium cocclneum Ham. Carex cruciata Wahl., Oplismenus compositus Beauv.
Centotheca tappacea Deov., Curculigo recurvata Dryand., Cypems globosus Allioni.
Chlorophytum arundinaceum Baker.

These population characteristics were compared to assess their ecological
importance. Pairwise comparison was made by Mann-Whitney's two sample
rank test. Early succes&ional non-sprouting category had significantly (I5 < 0-05)
lower leaf area ratio but higher reproductive effort than the late successional
category. Early successional sprouting species showed significantly (P < 0-05)
lower leaf area ratio than those of the early successional non-sprouting and late
successional types. However, its reproductive effort was not significantly
different (P >0-05) from both the categories.

Regression analysis showed that while, reproductive effort was negatively
correlated with leaf area ratio in the early successional non-sprouting category
(r = — 0-72, P < O'Ol), positive correlation existed in the early successional
sprouting category (r =0-86, P < 0-05). Further, ir the early successional non-
sprouting category only, reproductive effort was positively correlated with leaf
area per plant (figure 3). In the late successional category, no significant relation-
ship could be detected between leaf characteristics and reproductive effort.

K G Saxenci and P S Ramakrishnan

• Annuals
O Perennials

<-> 0

Q i
UJ '

*" 2 ^ 6 6 10

LOG LEAF AREA (cm2 )

Figure 3. Relationship between leaf area (cm2)/plant and reproductive effort (seed
mg/10cm2 leaf) in the non-sprouting early success-ional category.

Early successional non-sprouting populations when considered separately as
annuals and perennials did not differ significantly between themselves for their
leaf area ratio or reproductive effort. However, the negative correlation obtained
for these categories, between leaf area ratio and reproductive effort, was more
significant for the perennials (r = 0-93, P < 0-01) compared to the annuals
(r =0-65, P < 0-05). Reproductive effort was found to he positively correlated
with absolute leaf area in the perennials only (r = 0-80, P < 0-05) and not in
the annuals (figure 3).

5. Discussion

Leaf area ratio is an important structural concept as it expresses the proportion
of as^imilatory surface to respiratory mass (Evans 1972). Though the different
species exhibit a range of variation, late successional populations which occupy
the habitat of a low light regime, have higher leaf area ratio than the early succes-
sional ones as the adaptation in the former is to synthesize and maintain the
maximum light interception surface whereas the latter occupying a productive and
open environment divert their resource budget to other life purposes as growth
and reproduction. Higher leaf area ratio in shaded environments compared to
that in the open was also reported by Myerscaugh and Whitehead (1977) and
Bazzaz and Harper (1977). Lower leaf area ratio of early successional sprouting
species compared to that of the early successional non-sprouting types may be
accounted as due to the preferential allocation of photosyntbates to the under-
ground organs of the farmer. The somewhat lower reproductive effort of the. early
successional sprouting populations compared to the non-sprouting ones, though
not statistically significant (P > 0-05), might have evolved due to the failure of
their regeneration through seedlings (Wilson 1971),

Successioftat herbaceous, communities 67

Absolute leaf area of a plant gives an idea about its capacity of light inter-
ception and vigour while Leaf area ratio, the ratio of light interception surface and
total biomass (cm8 leaf area/g biornass) indicates the efficiency of dry matter
production on leaf area basis. Significantly positive correlation of leaf area and
negative correlation of leaf area ratio with reproductive effort in the early succes-
sionai non-sprouting category show that high reproductive potential is associated
with the vigour and also the production efficiency of the species. Comparatively
stronger correlation of leaf area ratio with reproductive effort tlau that of the
absolute leaf area with the reproductive effort in the early successional non-sprout-
ing species indicates that reproductive success here depends more upon the pro-
duction efficiency rather than the overall vigour of the plant. In contrast, early
successional sprouting populations exhibited positive correlation of leaf area ratio
with sexual reproductive eifort. Thus a species like Mimosa pudlca which has
a high leaf area ratio allocates more for sexual reproductive effort. Also it so
happens that this species is less vigorous in its vegetative regeneration compared
to others like Sac char wn arundinaceum and Thysanolaena maxima., and thus
compensates more through sexual reproduction. Complete paucity of flowering
in the first post-fire year, as in Imperata cylindrica and Grewia elastica has been
shown for a number of shrub species (Gill 1975). This aspect of the problem is
receiving our attention. Within the late successional group no signitican
(jP>0-05) relationship between leaf characteristics and reproductive effort wa
found, suggesting that sexual reproduction is not related with production effici-
ency or vigour of the plant ; the strategy seems to be to maximize vegetative-
growth in a closed habitat.

Considering the annuals and perennials of the early successional non-sprouting
category separately, certain differences are apparent. While reproductive effort
seems to be dependent upon production efficiency in both the cases, it was positively
correlated with absolute leaf area in the case of perennials alone indicating that
the vigour of the plant is less critical for the annuals having a single possibility
of flowering during their life-span.

MacArthur and Wilson (1967) pointed out that organisms in open environ-
ments are selected for greater reproductive capacity (r-strategy) while those in
closed environments are selected for greater ability to compete for resources, though
at the cost of lower reproductive potential (it-strategy). Grime (1974, 1977) has
extended this argument by describing three primary strategies in plants which.
are related to their ability to withstand competition, stress and disturbance. Here,
ruderal and stress tolerant strategies correspond to the extreme of r- and jf-selec-
tion while highly competitive species of productive environments occupy an inter-
mediate position. The findings of the present study clearly indicate that early
successional non-sprouting populations are of ruderal type as they are equipped
with the strategy to maximize seed production in order to colonize a disturbed
habitat whereas early successional sprouting and late successional populations
direct their synthetic capacity for competition and stress tolerance respectively by
economizing on the reproductive growth. Vegetative reproduction has been looked
merely as a growth in a horizontal plane (Harpeu 1977) and, therefore, is not
considered in the present study.

6S J? G Saxena anct P S Rcmakrishnm

Acknowledgement

This research was supported by the Department of EBvirenmcjit, Government of
India, under the ' Man and Biosphere ' programme.

References

Abrahamson W G 1975 Reproductive strategics in dewberries ; Ecology 56 721-726
Abrahamson W G 1979 Patterns of resource allocation in wildflower populations of fields and

woods ; Am. J. Bot. 66 71-79
Abrahamson W G and Gadgil M 1973 Growth and reproductive effort in golden rods (Solidago,

Compositae) ; Am. Nat. 107 651-661
Bazzaz F A and Harper J L 1977 Demographic analysis of the growth of Linwn itsitatissimwn ;

New Phytol. 78 193-208
Evans G C 1972 The quantitative analysis of plant growth (Oxford : Blackwell Scientific

Publications)

Gill A 3Vf 1975 Fire and the Australian flora : a review ; Aust. For. 38 4-25
Grime J P 1974 Vegetation classification by reference to strategies ; Nature (London) 250

26-31
Grime J P 1977 Evidence for the existence of three primary strategies in plants and its relevance

to ecological and evolutionary theory ; Am. Nat. Ill 1169-1194
Harper J L 1977 The population biology of plants (San Fransisco : Academic Press)
Harper J L and Ogden J 1970 The reproductive strategies of higher plants I. The concept of

strategy with special reference to Scnecio vnlgans L. ; /. Ecol 58 681-698
MacArthur R H and Wilson E O 1967 The theory of island biogeography (Princeton NJ |

Princeton University Press)

Misra R 1968 Ecology workbook (New Delhi : Oxford and IBM Publication)
McNaugaton S J 1975 r- and ^-selection in Typfa ; Am. Nat. 109 251-261
Myerscough P J and Whitehead F H 1967 Comparative biology of Tussilago far far a L,,
Chaiwerteriott angustifoliuni (L.) Scop. , Epilobium montanum L. and Epilobium adenocaulon
Hauskon. II. Growth and ecology ; New Phytol. 66 785-823
Newell S J and Tramcr E J 197S Reproductive strategies in herbaceous plant communities during

succession ; Ecology 59 228-234
Ogden J 1974 The reproductive strategy of higher plants. II, The reproductive strategy of

Tussilago farfara L. ; /. Ecol 61 291-324
Primack R B 1979 Reproductive effort in anuual and perennial species of Plantago (Plantagi-

naceae) ; Am. Nat. 114 51-62

Ramakrishnan P S, Toky O P, Mishra B K and Saxena K G 19SI Slash and burn
agriculture in north-eastern India ; In Fire regimes and ecosystem properties (eds.) H A
Mooney, J M Bonnicksen, N L Christensen, 3 E Lo-tan and W A Reincrs. USDA For.
Serv. Gen. Tech. Rep. pp. 560-584

Wilson M F 1971 Life history consequences of death rates ; Biologist (Phil, Sigma Sac,) 53
49-56

Proe. Indian Acad. Sci. (Plant Sci.), Vol. 91, Number 1, February 1982, pp. 69-77.
(g) Printed in India.

Vesicular arbusculai mycorrhiza in subtropical aquatic and marshy
plant communities

R CHAUBAL, G D SHARMA and R R MJSHRA

Department of Botany, North-Eastern Hill University, Shillang 793 0149 India

MS received 20 November 1980 ; revised 4 December 1981

Abstract. Occurrence of vesicular arbu&cular mycorrhiza in live subtropical ponds,
i.e. , cutrophic (jPl, P2 and P3), running water (T4), oligoiropluc lake (PS) and marshy
plant community (M) was studied. It was. observed that the plants growing in P\ , P$
and M habitats exhibited the vesicular arbuscular mycorrhizal association, whereas
the fungal association was. lacking in plants of jP2, P3 and P4 ponds. The cndo-
gonaceous spore population was estimated from water and sediments of the different
ponds and it was. found that cnxlophytcs in sediments arc less in terrestrial
habitats and completely absent from water samples. The bioassay studies revealed
that plants without mycorrhizal association grew poorly and all the cndophytcs
isolated could establish vesicular arbu&cular mycorrhizal associations in pot cultures.

Keywords. Vesicular arbuscular mycorrhiza ; subtropical aquatic community ;
endophytes ; bioassay.

L Introduction

Vesicular arbusciilar mycorrbiza (VAM) is universal in occurrence (NicoJson 1967;
Mosse 1973) and is useful to the host plant in various ^ays ; in enhancing the
uptake of nutrients (Kayman 1975) and water (Safir etal 1972), in resisting against
pathogen (Marx 1975) and in increasing the effective absorption surface of roots
(H&yman and Mosse 1971). Most of the rushes and sedges, however, are reported
to be non-mycorrhizal (Powell 1974 ; Khan 1974 ; Harley 1969 ; Gerdemann
1975). Recently, a few temperate (Soaderg&ard and Laeg&ard 1977) and tropical
(Bagyaraj et al 1979) aquatic species were reported to be mycorrhizal. In the
present study five aquatic and one marshy sub-tropical plant community were
•examined for occurrence of VAM and endophytic fungi. To test the ability and
efficiency of these endophytes a bioassay \*as also developed to test its potential
use in propagation of VAM in suecessiona communities of aquatic systems.

2. Materials and methods
2*1. Site selection

Five fresh water bodies and one marshy habitat in ShiUong (altitude 1450 m,
latitude 25-34°N and longitude 91*5i60E) \vere selected for the present study.

69

P.CJB-7

70 R Chaitbal, Q D Sharma and R R Mishra

The ponds, are designated as PI, P2, P3, P4, P5 and M (marshy habitat). Pl#
P2and P3 are eutrophic poads, P4 is a running stream and P5 is an oligotrophic
lake. PI remains dry in summer and receives water during the rainy season.

2-2. Collection of root samples and assessment of VAM

Intact plants with roots from different localities were collected in containers. The
roots were washed with tap water and cut into segments of approximately 1 cm in
length (100 segments from five plants). Further, the root segments were processed
and stained for VAM infection by the Phillips and Hayman (1970) technique.
Percentage of root infection was calculated in the presence of either vesicle,
arbuscules or both by counting the infected segments by the slide method (Mishra
et at 1981).

2-3. Estimation of endogonaceous spores

Fifty ml of water and 30 g of sediment were collected by water sampler from each
site in five replicates. The sediment was wet sieved and decanted (Gcrdemann
and Nicolson 1963) and water was filtered through. Whatman No. 1. filter paper,
tne spores retained on the sieves and filter paper were examined under a binocular
microscope.

2«4. Sioassay study for infection efficiency of endogonaceous spores In pot cultures

Sterilized maize seeds were germinated in sterilized moist chambers. Five seed-
lings (2 cm radicle stage) were transplanted to pots (11 x 10") containing steri-
lized soil (soil 4- sand in equal amount w/w). The plants were inoculated with
endophytes isolated from the sediments ; uninoculated pots received soil with
microflora but were devoid of mycorrhizal propagules. All the pots were regularly
watered. Plants were harvested 15, 30 and 45 days after transplanting. Root
infection, shoot height, dry weight and leaf production were recorded at each
harvesting.

2.5. PkysiGQ«chemical analysis of water and soil

pH of soil and water was measured by electronic digital pH meter. Organic
carbon (Walkey method), nitrate (phenol disulphonic method) and phosphorus
(Bray's method) were estimated as outlined by Jackson (1967).

3. Results

Five subtropical aquatic species, viz. Rotala rotwdifolia, Paspalum dilatatumt
Polygonum hydropiper, Nymphaea alba and Hydrilla verticellata have been observed
for the first time as mycorrhizal (table 1, figures 1-4). Vesicular arbuscular my-
corrhiza (VAM) was observed in JP1, P5 and M plant communities. Plants
growing in F2, P3 and P4 ponds \vere devoid of VAM infection. Percentage
infection was highest in PS plants followed by PI and was least in plants from
the marshy (M) habitat (table 2). Vesicles and hyphae were regularly observed.
The population of endogonaceous spores in general was low in all the sediments
from different sites and the number did not differ significantly (table 2). No

Vesicular arbusculaf mycorrhiza

71

Table 1. Percentage occurrence of vesicular arbuscular mycorrhiza in root* of
different plant species of sub-tropical aquatic and marshy communities.

Aquatic Marshy
commu- Plant species VAM (%) commu- Plant species
nity nity

VAM %
(root)

PI Rotala rotundifolia

10-00

Impatiens chinensis

100-00

Paspalum dilatatum

60-00

Drosera sp.

64-00

Polygonum hydropiper

56-00

Utricularia sp.

73-00

Cyperus distans

0-00

Soncfius sp.

52-00

Polygonum cajpitatum

51-00

PZ Cardamine hirsuta

0-00

Drymaria cordata

33-00

C. macrophylla

0-00

Plantago major

19-00

Scirpus articulate

0-00

Nasturtium indica

0-00

S. juncoides

0-00

Anemone rivalaris

0-00

Eleocharis congesta

0-00

Steudnera colocasioides

0-00

P3 Rotala rotundifolia

0-00

Oen&thera javinaca

0-00

Cyperus distans

0-00

Brassica juncea

12-00

Eleocharis congesta

0-00

Rumex nepalensis

28-00

Spargonium ramoswn

0-00

Galium rotundifolium

16-00

Panicum brevifolium

46-00

P4 Hydrilla vertidllata

0-00

Alternanthera philoxeroides

0-00

Monochoria hastata

0-00

Hydrocotyle sibthorpioides

0-00

Lasia spinosa

0-00

Pa Rotala rotundifolia

3-00

Hydrilla vertidllata

16-00

Nymphaea alba

12-00

Table 2. Eridogonaceous spore population and frequency of YAM jmycorrbizal
plants in different localities.

Mean spore

population/30 g/rnl

soil/water

Sites

- Frequency (%) of

Water

Soil

mycorrhiza

(per 30 ml) (per 30 g)

PI

0

27

75

P*

0

33

X)

P.

0

6

0

P*

0

7

0

A

0

11

100

Mr

a

34

71-3

72 R Chanhal, G D Sharma and R R Mishra

endogoiiaceous spores were found in \vater. The pot culture studies on infection
efficiency of endogonaceous spores from different sites revealed that they may
infect and establish in roots growing in soil and that their efficiency may differ
(table 3). TMnoculated seedlings did not grow well, whereas the seedlings with
mycorrhizal association gre^ better.

Soil and water from all the sites were acidic in nature. Organic carbon was
very lew in water samples and highest in soil of marshy land. Nitrate and phos-
phate were highest in P2 and Fl and lowest in P5 and M sites (table 4),

4. Discussion

The study reveals that VAM occur rarely in aquatic subtropical plant communities,
but they are not completely absent. The results are, therefore, contrary to the
vieus of Harley (1969), Gerdemann (1975), Powell (1974) and Khan (1974), but
support the findings of Sondergaard and Laegaard (1977) and Bagyaraj et at
(1979). Apart from environmental factors like light, temperature and aeration,
the occurrence of VAM and its intensity may be regulated by the nutrient status
of the aquatic system. Gerdemann (1968) observed that mycorrhiza may tie low
in rich soil. The amount of infection in a fe\v species, i.e., Rotala rotundifolia
and Hydrilla verticellata differed insignificantly in different \vater systems depending
on the nutrient status and other physical factors of these systems. In general,
jpl, P5 and M sites favoured VAM establishment but P2, P3 and P4 ponds
did not possess any mycorrhizal association. It seems that temporary drying of
PI and M habitats in summer may initiate the VAM establishment. A similar
trend was also observed in other environmental conditions (Read et al 1976).
The high percentage occurrence of VAM in, PS may be attributed to its eligotro-
phic nature (Sonderga?rd and Laeg§ard 1977), as Hydrilla veniddlata was my-
corrhizal in P5 community but not in P4. Besides the root system, the shoot
also helps the aquatic plants in absorbing the nutrients from water and this may
be one of the reasons for the absence or less frequent mycorrhizal association in
certain plants. Sutcliffe (1962) suggested that in aquatic plants, the roots primarily
act as anchors ; on the other hand, I>enny (1972) concluded that the nutrients
may enter through roots and shoots. Therefore, it seems that phosphorus uptake
may depend oa the efficiency of the root system, i.e.9 some root systems may be
adaptive enough to draw the phosphate at low level even in the absence of VAM
association (Powell 1975). VAM may not be of much importance in plants grow-
ing in rich medium (P29 P3 and P4 system) but it may help the hosts in absorp-
tion of the nutrients growing in low level nutrient systems (PS). Therefore, two
conditions, i.e., temporary drying of the aquatic system and the oligotrophic nature
seem to be more favourable for VAM development. The presence of endophyte
spores in all aquatic, systems suggests that these spores probably enter into the
water system through run off from terrestrial ecosystem, their subsequent develop-
ment, however, is governed by water regime, light, aeration and other factors
(Mosse 1973). The presence of endophyte spore in P2 community may further
indicate that either these species are not capable of causing infection or plants may
not be susceptible in such systems (Reeves et al 1979 ; Miller 1979). The bio-

Vesicular arbuscular mycorrhiza

Figures 1-4. 1. Spore, Glom:is sp. infecting foot tissuo of Panicitm brevlfolium
(x 1,000), 2. Vesicles and hyplue m root tissue of l*np atiens balsamia (x 100),

3. Obovato vesicles of Ghtnus sp. in tho root tissue of Rotala rotundifolia (X 400)^

4. Round vogiclos with oil globule and degenerating stage of laypha in root tissue
of Nyrnphaea alba (x 400),

Vesicular arbuxcular myaarrhiza

75

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76 R Ckauhal, G D Sharrna and R R Mishm

Table 4. Physico-chemical characters of soil and water in different water bodies .

Site

pH

Org. Carbon (%) Nitrate (ppm) Phosphate (ppm)

water

soil

water

Soil

Soil

water water

A

6-5

8-3

0-05

4'9

2-2

0-20

3-6

6-62

p*

6-4

6-3

Q'07

7-6

1-6

0-17

3*8

7-8

Pa

6-0

6-3

0-05

5-3

1-6

0-07

1-07

3-18

Pi

6-6

6-5

0*06

5-6

1-8

0-09

0-1

2-01

p*

6-1

6-3

0-04

3-9

1-4

0-05

o-oi

1-96

M

6-2

0*00

8-0

0-1

•-

1*42

assay studies using maize (Zea mays) as test plant suggested that these endo-
phytes may develop VAM and enhance the growth of plants, when placed into
sterilized soil. It is of great ecological importance to study the establishment,
germinatioa and entrance of these endophytes into the hosts under diverse
environmental conditions, in understanding of the developmental pattern of aquatic
and terrestrial communities. Baylis (1959) suggested that VAM play a significant
role in the evolution of plant communities. Therefore, studies in controlled
conditions in aquatic habitats on the esta lishment of VAM will provide infor-
mation towards further understanding of succession in aquatic systems and <would
aid management of aquacuitural systems.

References

Bagyaraj D J, Manjunath A and Patil R B 1979 Occurrence of vesicular arbuscular mycorrhiza
in some tropical aquatic plants ; Trans. Br. Mycol Soc. 72 164-165

Baylis GTS 1959 Effect of VAM on growth of Gnselinia littoralis (Cornaceae) ; New Phytol.

58 274-280

Denny P 1972 Sites of nutrient absorption in aquatic macrophytes ; /. Ecol. 60 819-329
Gerdemann J W 1975 Vesicular arbuscular mycorrhiza. In The development and function of roots

(eds.) J G Torrey and Clarkson (London : Academic Press)
Gerdemann J W 1968 Vesicular arbuscular mycorrhiza and plant growth; Ann. Rev. Phyto-^

pathol 6 397-410

Gerdemann J W and Nicolson T H 1963 Spores of mycorrhizal Endogone species extracted from
soil by wet sieving and decanting ; Trans. Br. Mycol. Soc. 46 235-243

Harley J L 1969 The biology of mycorrhiza (London : Leonard Hill)

Hayman D A 1975 Phosphorus cycling by soil microorganisms and plant roots. In soil Micro-
biology (ed.) N Walker (London : Butterworth)

Hayman D A and Mosse B 1971 Plant growth responses to VAM I. Growth of Endogone
inoculated plants in phosphate deficient soils ; Neb Phytol. 70 19-27

Vesicular arbuscutar mycorrhizd 1i

kson M L 1967 Soil chemical analysis (New Delhi : Prentice Hall)

.an A G 1974 The occurrence of mycarrhizas la Halophyfcos, hydrophytes and xerophytes and

of Endogow spores in the adjacent soils ; /. Gen. Microbiol. 81 7-14
irx D H 1975 Mycorrliizae of exotic trees in tlxe Peruvian Andes and synthesis of ectomyoor-

rhizae on niexican pines ; For. Sci. 21 353-35$
Her RM 1979 Some occurrence of vesicular arbuscular mycorrhizae in natural and disturbed

ecosystems of Red desert ; Can. J. Bat. 57 619-623
shra R R, Sharma G D and Kharsyntiew IB 1981 Response of inoculum density in maize ;

Experientia 37 568-569
:>sse B 1973 Advances in the study of vesicular arbuscular mycorrhiza ; Ann. Rev. Phytopathol.

11 171-195
colson T H 1967 Vesicular arbuscular mycorrhiza a universal plant symbiosis ; Sci. Prog.

(Oxford) 55 551-581

illips J M and Hayman D A 1970 Improved procedures for clearing roots and staining para-
sitic and vesicular arbuscular mycorrhizal fungi for rapid assessment of infection ; Traits.

Br. Mycol Sac. 55 158-161
wel C L 1974 Effect of P-fertilizer on root morphology and P-uptake of Carex coriacea ;

Plant Soil 41 651-667
wel C L 1975 Plant growth responses to VAM VIII Uptake of P by Onion and clover

infected with different Endogorte spore types in 32 P-labelled soils ; jNew PhytoL 75 563-566
?ad D J, Kovchiki H K and Hodgson J 1976 Vesicular arbuscular mycorrhiza in natural

vegetation systems ; New PhytoL 77 641-653
;cves B F, Wagner D, Moorman T and Kiel J 1979 The role of endomycorrhiza in rcvcgc-

tation practices in semi arid west. L A comparison of incidence of mycorrhizae in severe y

disturbed W. natural environments ; Am. J. Bot. 66 6—13
tir G R, Boyer J A and Gerdemann J W 1972- Nutrient status and mycorrhizal enhancement

of water transport in Soybeans ; Plant Physiol. 49 700-803
ndcrgdard M and Laegaad S 1977 Vesicular arbuscular mycorrhiza in sonic aquatic vascular

plants ; Nature 268 232-233
tcliffc J F 1962 Mineral salts absorption in plants (Oxford : Perfiamon)

Proc. Indian Acad. Sci. (Plant Sci.), Vol. 91, Number i, April 1982, pp. 79-82.
© Printed in India.

Chandmsekhamnia : A new genus of Poaceae from Kerala, India

V J NAIR, V S RAMACHANDRAN and P V SREEKUMAR

Botanical Survey of India, Caimbatore 641003, India

MS received 26 June 1981 ; revised 30 March 1982

Abstract. A new genus, Chandrasekharania and a new species Chandrasekharania
keralensis under it are being described from Cannauore District, Kerala.

Keywords. Chandrasekharania keralensis ', Poaceae ; new genus; new species.

1. Introduction

Intensive explorations are being conducted in various parts of Kerala to clearly
understand the flora of this botanically rich area. During these tours a number
of very interesting plants were collected. One of these belonging to the family
Poaceae., on critical study with reference to available literature (see Bor 1960 ;
Ved Prakash et al 1978) and the specimens available in the Herbarium of Southern
Circle, Botanical Survey of India, Coimbatore (MH) and the Central National
Herbarium, Calcutta (CAL), turned out to be quite distinct. So a specimen along
with a detaile^l description and analytical sketches was sent to Dr Thomas A
Cope, Herbarium, Royal Botanic Gardens, Kew, for expert opinion. He confirmed
that our collection belongs to a new genus. This along with its type species is
described here.

2. Latin description

Chandrasekharania V J Nair, V S Ramachandran et P V Sreekumar gen. nov.
Gramen annuum. Culmi erecti vel decumbentes. Folia lanceolata vel ovato-
lanceolata, basibus cordatis. Ligulae obscurae. Inflorescentia contracta, racemi-
flora, spiculae omnibus similaribus, lateraliter compressis, floribus duobus, pedi-
cellatis, pedicellis numquam articulatis. Flosculi similes, hennaphroditi. Glumae
inaequales, coriaceae, ovatae vel ovatolanceolatae, 7-nervatae, aristatae, aristis
curtis et rectis. Glumae infernae breviores in dorsi parte superiore pilis basi
tuberculatis. Lemmata 5-nervatae, coriacea, elliptico-lanceolata, apicibus
breviter emarginatis, aristatis ad sinus. Aristis curtis et rectis. Paleae ovato-
ellipticae, hyalinae, binervatae, bicarinatae, carinis ciliatis, apicibus bilobatis.
Ovarium glabrum. Styli 2, distincti. Stigmata plumos.a. Stamina 3, filamentis
brevissimis. Lodiculae 2. Grana ellipsoidea.

79

80 V J If air 9 V S Ramachandran and P V Sreekumar

Species typica sequens :

Chandrasekliarania keraknsis V J Nair, V S Ramachandran et P V Sreekumar
sp. nov.

Gramen annuum usque 40 cm altum. Culmi graciles, striati leaves et glabri.

Nodis inferioribus radicantes. Folia 2-4 -5 cm longa,, 5-8 mm lata, marginibus

et paginis ambabus pilis densis vel sparsis, basi tuberculatis. Vaginae striatae,

glabrae, internodiis breviores. Inflorescentia, contracta, ovoidea vel oblonga,

spiciformis; racemiflora, 1 • 5-2- 5 cm longa, 1-1 • 5 cm Uta. Spiculae 5-6 mm longae.

Pedicelli 0-5-1 -5 mm longi. Glumae infernae 7-nervatae, nervi duo alternati et

mediani prominentes, nervi cateri inconspicui, ovatae, 3-5-4 mm longae, ari&tatae,

arista ca. 3 mm longa, costa et arista scabridus, inargines ciliolati, dimidia supera

paginarum dorsalia dense tecta pilis, basibus tubercularibus. Glumae superae

5 • 5-6 mm longae, ovato-lanceolatae acuminatae, aristatae, 7-nervatae, inargines

ciliolati nervi scaberuli, arista scabrida, ca. 3mm longa. Lemmata elliptico-

lanceolata, 4-5 mm longa,, apicibus breviter emarginatis, aristatis ad sinus, coriacea,

glabra praeter scaberula costas, arista scabrida, 1-5-2 mm longa. Paleae ca.

4 mm longae, subtiles hyalinae, binervatae, dorsiis dimidiis infernis pilis longiore,

cariniis ciliatis, apicibus bilobatis, lobi acuti. Gynoecium ca. 2 mm longum.

Ovarium 0-5 mm longum, oblongum. Antherae ca. 1-25 mm longae. Fila ca.

0-25 mm longa. Lodiculae cuneatae, apicibus emarginatis. Grana ellipsoidea,

ca. 1 mm longa, fusca et extremis distalibus maculae ateris.

Holotypus : Kerala, Cannanore District, Kannoth, ± 175 m, 18-2-1978,
V S Ramachandran 54064 (CAL). Isotypi in K et MH.

3. Description

Chandrasekharania V J Nair, V S Ramachandran et P V Sreekumar gen. nov.
Annual grass. Culms erect or decumbent. Leaves flat, lanceolate or ovate-
lanceolate, base cordate. Ligules obscure. Inflorescence terminal, solitary,
contracted, racemose. Spikelets ail alike, laterally compressed, two flowered,
pedicelled, pedicels never jointed. Florets similar, bisexual. Glumes unequal,
coriaceous, ovate or ovate-lanceolate, 7-nerved, awned, awns short and straight.
Lower glume smaller with the dorsal surface with tubercle based hairs in the upper
half. Lemma 5-nerved, coriaceous, elliptic-lanceolate, minutely notched at the
apex with an awn in the sinus. Awns short and straight. Palea ovate-elliptic,
hyaline, 2-nerved, 2-keeled, keels ciliate, apex two lobed. Ovary glabrous.
Styles 2, distinct. Stigmas feathery. Stamens 3. Filaments very short. Lodi-
cules 2. Grain ellipsoid.

Type species followsi :

Chandrasekharania keralensis V J Nair, V S Ramachandran et P V Sreekumar
sp nov.

Annual grass up to 40 cm tall. Culms rooting at the lower nodes, slender,
striate, smooth, glabrous. Leaves 2-4, 5 cm long, 5-8 mm broad, both surfaces
and margins with dense or sparse tubercle based hairs. Sheaths shorter than the

Chandrasekharania, new genus of Poaceae

81

12mm

10 \JJ

Figures 1-13. Chandrasekharania kemlemis gzn. et sjp. nov. 1. Habit. 2. Spikclet.
3. Lower glutne. 4. Upper glutxxe. 5. Lower lemma. 6. Lower palea— dorsal view.
7. Same— ventral view, 8. Upper lemma 9. Upper palea— dorsal view, 10. Same—
ventral view. 11. Lodicules and stamens. 12. Gyaoecium, 13. Grain.

internodes, striate, glabrous, Inflorescence a contracted ovoid or oblong spikate
raceme, 1-5-2 -5 cm long, 1-1 -5 cm broad. Spdkelets 5-6 mm long. Pedicels
0-5-1 -5 mm long, scabrid. Lower glume 3 -5-4 mm long, ovate, awned, awn
ca. 3 mm long, densely hairy on the upper half of the dorsal surface with tubercle
based hairs, the median and the alternating two nerves prominent, the midrib
and the awn scabrid, margins ciliolate. Upper glume 5 -5-6 mm long, ovate-
lanceolate, acuminate, the tip with an awn ca. 3 mm long, scaberulous on the
nerves, scabrid on the arista, margins ciliolate. Lemma elliptic-lanceolate,
4-5 mm long, shortly notched at apex with an awn in the sinus, coriaceous, glabrous
except for the scaberulous midrib, awn scabrid, 1 • 5-2 mm long. Palea ca. 4 mm

82 V J Nair, V S Ramachandran and P V Sreekumar

long, delicate, hyaline, long-ciliate on the lower half of the outer surface, keels
ciliate, apex bilobed, lobes acute. Gynoecium ca. 2 mm in length ; ovary ca.
0-5 mm long, oblong. Anthers ca. 1-25 mm long, filaments ca. 0-25 mm long.
Lodicules cuneate and shallowly notched at apex. Grains ellipsoid, ca. I mm
long, brown coloured with a black spot at the distal end.

Holotype : Kerala, Cannanore District, Kannoth, ± 175 m, 18-2-1978,
V S Ramachandran 54064 (CAL). Isotypes in K and MH.

4. Affinities

Affinities of this new genus are not very clear. According to Dr T A Cope, Kew
Herbarium, anatomical studies are required to clearly understand its affinities
(personal communication).

5. Etymology

The genus is named after Dr N Chandrasckharaa Nair, the first author's teacher
and present Joint Director, Botanical Survey of India, Coimbatore, in recognition
of his outstanding contributions to Indian botany. The specific epithet is after
'Kerala' the state from where the plant has been collected

Acknowledgements

Authors are thankful to Dr Thomas A Cope, The Herbarium, Royal Botanic
Gardens, Kew, for examining the specimen and giving his valuable opinion and
R Suudararaghavan, Regional Botanist, (Kew) for help. Rev. Fr. Zachaeus,
CMI, Principal, Lisieux Higher Secondary School, Coimbatore, has kindly corrected
the Latin description.

References

Bor N L 1960 The Grasses of Burma, Ceylon, India and Pakistan (Oxford, London, New York, Paris :
Pergamon Press), p. 1-767 : Repr.ed. 1973 (Otto. Koeltz Antiquariat, Koeningstein, BRD)

Ved Prakash, Shukla U, Pal D C and Jain S K 1978 Additions to the Indian grass flora in
last two decades ; Bull. Bot. Surv. India 20 143-147

Prac. Indian Acad. Sci. (Plant Sci.), Vol, 91, Number 2, April 1982, pp. 83-91,
© Printed in India.

Chromosome relationships of spinous solanums

P B KIRTI* and B G S RAO

Department of Botany, Andhra University, Waltair 530003, India

* Present address : JARl-Regional Station, Jlajendranagar, Hyderabad 500030,

India

MS recdved 25 April 19S1 ; revised 29 March 1982

Abstract. Chramasome pairing wa.s studied in reciprocal hybrids of S. integnfolium
and S. indlaim and the FI S. integnfolium X 5. surattense. Pairing was generally
close and meiosis regular with higher chromosome associations. AH hybrids were
highly sterile. Such sterility could be due to the formation of unbalanced gametes
following pairing and exchange between partially homeologo-us chromosomes,
S. integrifolium, S. irtdictitn, S. surattense along with S. melongena and its wild forms
form a closely related group of taxa.

Keywords. Solarium indicum ; 5. surattense ; S. integnfolium ; Fj hybrids ; sterility ;
chromosome relationships.

1. Introduction

Chromosomal studies on species hybrids are of considerable importance in
elucidating species relationships and evolutionary trends. Such studies on
spinous Solanums, which are important medicinally as well as vegetables, were
meagre because of the difficulty in producing interspecific hybrids. Magoon
et al (1962) cited the earliest attempt^ of species hybridization in solaminis
including those of Sarvayya (1936), Hagiwara and lida (1939) and Tatebe (1939).
Bhadhuri (1951) first discussed the interrelationships of spinous solanums
including the origin of S. melongena. Further attempts in this direction were
made by Zutshi (1967), Rajasekaran (1969, 1970a, b, 1971), Rajasekaran and
Sivasubramanian (1971), Rangaswamy and Kadambavanasundaram (1974),
Veerabhadra Rao (1977), Veerabhadra Rao and Rao (1977b, c), Kirti and
Rao (1978, 1981a, b, c). Most of the cultivars of the brinjal are susceptible to
various diseases and pests (Krishnaiah and Vijay 1975). Wild species suet as
S. integnfolium and S. indicum are resistant to some of the diseases and pests.
However, these species cannot be utilized in egg plant improvement unless the
interrelationships of the whole group are unravelled. First attempts of crossing
«S. integnfolium and S. melongena were made by Hagiwara and lida (1939), Tatebe
(1939), Khan et al (1978). Kirti and Rao (1981c) discussed at length the chromo-
some relationships between these two species as well as S. integrifoUum and

84 ? 5 Kirti and 3 G S Rao

S. indicum var. multiflora. The chromosome relationships S. mtegrifolwm,
S. indicum, S. surattense and other species are discussed in some detail here.

2. Materials and methods

Seeds of S. indicum L. and S. surattense Burm. F. (=S. xanthocarpum Schrad
and Wendi) were collected from plants occurring wild while seeds of S. integrifolium
were kindly provided by Prof P V Bhiravamurthy of the Andhra University, who
obtained them from Denmark.

Method of crossing was the one employed by Veerabhadra Rao and Rao
(1977a).

Standard propionic carmine schedule was used for PMC smears.

3. Results and discussion

3-1. Crossability

An on-the field screening for functional pistillate flowers was done since stylar
heteromorphism is prevalent in spinous solanums. Only flowers with long and
exserted styles can be used as pistillate flowers in crossing experiments. Rao
and Veerabhadra Rao (1976) have studied this phenomenon in some detail in
S. surattense. Shamim Baksh etal (1978) described this in S. integrifolium. In
S. indicum, the inflorescence is a 3-4 flowered cyme and only the basal most flower
having the long an.d exserted style sets fruit. Others are generally with shorter
and inserted styles and do not set fruit. So this phenomenon should be taken
as an important criterion in the consideration of crossability.

The ease with which any two species are crossed reflects to some extent the
nearness of the species concerned with the absence of prezygotic incompatibility
"barriers. In the present study, S. integrifolium and S. indicum could be crossed
in reciprocal directions and the percentage of hybrid seed obtained was very
high (table 1). S. integrifolium could be crossed with S. surattense in only one
direction and only a solitary hybrid could "be realised. Thus it seems S. integri-
folium is more closely related to S. indicum on the basis of this observation than
to S. surattense even though other factors are to be taken into consideration.

3-2. Morphology of parental species and hybrids

S. indicum and S. surattense conformed to the classical descriptions of Gamble
(1957). S. integrifolium is an erect herb with small white flowers, and scarlet
red coloured and lobed berries.

Reciprocal hybrids of S. integrifolium and S. indicum resembled each other and
were intermediate between the parents in some features and resembling either
of the parents in some respects. Similarly the FI S. integrifolium x S. surattense
was intermediate.

Chromosome relalioriships of spinous xoiainmis

Figures 1-4. Cytology of hybrids ; Diakirwsia-Metaphase-I in S, indicum X
S. integrifoUutn 1. 10n -h 1J?. 2. 12n. 3. 10n -flir

Chromosome relationships of spinous s olanums

87

Table 1. Results? on crossabihty relationships of S. integrifolium, S. indicum and
S. Surattense.

S. integri folium

S. indicum

5. integrijolium

X

X

X

S. indicum

S. integrifolinni

S. surattense

umber of pollinations made

15

70

105

umber of fruits set

1

1

1

umber of healthy Seeds per fruit

165

174

1

Tcentagc of seed g[errmnation

97-6

94-2

100

>rcentage of hybrid seed obtained in

relation to the number of * target '

ovules* per ovary

78-9

56-5

0-48

Approximated from the average number of seeds per fruit in parental species, 209 in
i'rttegrifoliurn and 308 in 5. indicum.

Table 2. Frequencies of nuclei with different chromosome associations observed
in hybrids of S. integrifolium, S. indicum and S. surattense.

integrifoliujn S. integrifolium X Kinds of chromosome 5. indicum x
S. surattense S. indicmi associations per nucleus S. integrifolium
requency of Frequency of Frequency of nuclei

nuclei

nuclei

Makinesis

Diakinesis

Metaphase-I IV III

II

I Diakiriesis

Motaphase-I

9

27

15 1

10

€8

28

(40-9)

(44-3)

(50-0)

(63-0)

(73-7)

3

7

1

9

2 5

(13-6)

(11-5)

(4-6)

« *>

t

..

1

8

4 2

• •>

(1-8)

. .

4

2 .. 1

10

1 4

1

(6-6)

(6'7)

(3-7)

(2-6)

2

_

1

9

3 1

(9-1)

(0-9)

4

IS

12

12

23

S

(19-2)

(29-5)

(40-0)

(21-3)

(2M)

4

5

1

11

2 5

1

(18-2)

(8-2)

(3-3)

(4'6)

(2-6)

22

61

30 Total number

of nuclei

analysed 108

38

gures in parentheses indicate percentage frequency),
ytetapha$e-I data not available,

88 P B Klrtl and B G S Rao

3-3. Cytological observations

Meiosis proceeds regularly in the parents with the formation of twelve bivalents
(2n = 24) and regular chromosome separation at anaphase I and II.

Various types of chromosome associations (figures 1-3) and their frequencies
observed in the hybrids are summarised in table 2. Higher chromosome associations
in hybrids indicate chromosomal structural repatterning in the divergence of the
concerned taxa.

Mean frequencies of chiasmata in parents and hybrids are summarized in table 3.
Mean values in hybrids were significantly lower than in parents indicating some-
what reduced chromosome homeologies.

Later stages of meiosis followed normally with regular chromosome segregation
at anaphases I and II. Laggards, bridges without fragments and micronuclei were
rarely encountered. Despite regular meiotic divisions, all the hybrids were over
95% pollen sterile. Stebbins (1950) advocated that cryptic chromosomal structural
differences are responsible for sterility in hybrids with regular meiotic events.
It is also possible that segregational events, following pairing and exchange of
segments between partially homeologous chromosomes of the two genomes leading
to the formation of unbalanced gametes, eventually lead to lethality (Kirti 1978).

— — Hybrid obtaine-d

- Fruits set, seed sel variable from many

to none -a II shrivelled
--- Cross unsuccessful

Figure 4. Crossability relationships of some spinous

Chromosome relationships of spinous solanums

Reproductive behaviour of hybrids

_ Fertile

_ — — Partially fertile
Sterile

Figure 5. Fertility relationships of some spinous solanums.

Table 3. Average chiasma frequencies observed in 5. surattense, S. integrifolium,
S. indicum and their hybrids.

S.Surat- S.integri- S.indicum S.integri- S.indicurnx S.inte-

tense folium folium x S.integri" grifolium

S. indicum folium X S. surat-
tense

DiaJc* Diak Meta-I Diak Meta-I Diak Meta-I Diak Meta-I Biak*

Slumber of PMCst
analysed 30 41

32 27

61

30 108 38 22

Average chiasma 18-17 19-24 19-04 20'06 19-90 17-65 17-56 17-83 17-20 16-41
frequency per ±0-20 ±0-37 ±0-61 iO'23 ±0-40 iO-16 ^0-39 ±0-20 ±0-34 ±0-26
nucleus

Per bivalent

1-Sl 1'60 1-59 1-67 1-66 1-47 1-46 1-49 1'43

1-32

Diak = Diakinesis Meta-I = Metaphasc~I
* Metap^a$e-I data not available.

90 PS Ktrti and B G S Rao

3-4* Species relationships

Thus on the basis of the above cytogenetic observations on crossability, chromo-
some pairing and meiotic events, S. integnfolium is closely related to 5. indicum
and S. surattense (figures 4 and 5). The relationship with the former species
seems to be much cloeer since they can be crossed in reciprocal directions and the
recombination potential of the reciprocal hybrids is approximately the same as
in the parents. The hybrid, S. integnfolium x S. surattense, could not be main-
tained for long and extensive analyses on this hybrid could not be done. But
to the extent possible, it was observed that univ^lents were more frequent and
mean chiasma frequencies were comparatively lower. Also the cross was
achieved with greater difficulty and only a solitary hybrid could be realised. The
reciprocal cross was unsuccessful. But as far as possible, S. integrifoliurn and
S. surattense are also closely related. While full bivalent pairing has been reported
in hybrids of S. surattense and S. indicum, even though the hybrids are sterile
(Bhadhuri 1951 ; Rajasekaran 1969), Raju etal (1981) observed higher chromo-
some associations and a good amount of fertility in the hybrid, S. indicum x
S. surattense. Thus it can be concluded that the three species S. integnfolium,
S. indicum and S. surattense form a closely related group. With the reports of
hybridization in spinous solanums of Bhadhuri (1951), Zutshi (1967), Rajasekaran
(1969, 1970a,b, 1971), Rangaswamy and Kadambavanasundaram (1974), Veera-
bhadra Rao and Rao (1977b), Khan et al (1978), Kirti and Rao (1982),
S. integrifoliurn, S. rnelongena and its wild forms, S. surattense, S. indicum
form a group of closely related taxa.

Acknowledgements

The first author is thankful to the CSIR and UGC, New Delhi, for the award of
fellowships.

References

Baksh SJqbal M and Jamal A 197& Breeding systems of Solatium integrifoliurn with an emphasis
on sex potential and inter-crassability ; Euphytica 27 SI 1-8 15

Bhadhuri P N 1951 Interrelationships of non-tuberiferous species of solanum with some consi-
deration on the origin of brinjal, S. melongena ; Indian J. Genet. 11 75-32

Gamble G S 1957 « Solaaaceae '. In Flora of the Presidency of Madras Bat. Survey of India,
Calcutta, Vol. II, pp. 654-661

Hagiwara T and lida H 1939 Interspecific crosses between Solanum integnfolium and the egg
plant and the abnormal individuals which appeared in F2. Sot. Mag. (Tokyo} 50 (cited by
Magoon et al 1962)

Khan Reayat, Rao G R. and Baksh S 197S Cytogenetics of Solanum integrifoliurn and its

possible use in egg-plant breeding; Indian J. Genet, 33 343-347
Kirti P B 1978 Studies on the cytogenetic aspects of some spinous solanums. Unpublished Ph.D

thesis submitted to. the Andhra University, Waltair.
Kirti P B and Rao B G S 1978 Meiasis in the interspecific hybrid of two spinous

and its bearing on their affinities ; Curr. $ci, 47 696-697

Chromosome relationships of spirtous solanums 9\

Karti P B and Rao B G S 1981a Cytogenetic studies on fi hybrid Solatium indicum X S. torvum ;

Theor. Appl Genet. 59 303-306
KLirti P B and Rao B G S 1981b Chromosome pairing in reciprocal hybrids of Solatium

integri 'folium and S. indicum var. multi flora ; Caryologia 32 289-296
Kirti P B and Rao B G S 1982 Cytological studies on Fx hybrids of Solanum integrifolium with

S. melongena and S. melongena var. insanum ; Genetica (Accepted).
ECrishnaiah K and Vijay O P 1975 Evaluation of brinjal varieties for resistance to brinjal shoot

and fruit-borer, Leucinodes orbonalis ; Indian J. Hort. 32 84-86
Magoon M L, Ramanujam S and Cooper D G 1962 Cytogenetical studies in relation to the

origin and differentiation of species in the genus Solanum L.; Caryologia 15 151-252,
lajasekaran S 1969 Cytogenetic studies on the interrelationships of some common Solanum

species occurring in South India ; Annamalai Univ. Agric. Res. Annual 1 49-60
lajasekaran S I970a Cytogenetic studies o.f the Fi hybrid, S. indicum x S. melong-ena and its

arnphidiploid ; Euphytica 19 217-224
fcajasekaran S 1970b Cytology of hybrid, S. indicum X S. melongena var. insanum ; Curr. Sci.

3922
lajasekaran S 1971 Cytological studies on the Fx hybrid Solanum xanthocarjpum Schrad and

Wendl x s. melongena L. and its amphidiploid ; Caryologia 24 261-267
lajasekaran S and Sivasubramanian V 1971 Cytology of the FI hybrid of Solanum zuccagnfanum

Dunn x S. melongena L. ; Theor. Appl. Genet. 41 S5-S6
^aju D S S R, Moorty K V, Kirti P B and Rao B G S 1981 Studies on the origin of Solanum

melongena L. (The egg plant) : I. Cytogenetics of interspecific hybrids between S. indicum

L. and S. surattense Burm. F ; Indian J. Bot. 4 91-98
langaswamy P and Kadambavanasundaram M 1974 A Cytogenetic analysis of sterility in

interspecific hybrid, £. indicum L. x S. melangena L. ; Cytologia 39 645-654
lao B G S and Veerabhadra Rao S 1976 Some observations on the reproductive biology of

a few spino.us solanums in relation to their crossability relationships; In Physiology of sexual

reproduction in flowering plants (ed.) C P Malik (New Delhi : Kalyani Publishers)
larvayya J 1936 The first generation of an interspecific hybrid cross in solanums, between

Solanum melongena and S. xanthocarpum ; Madras Agric. J. 24 132-142
Itebbins G L 1950 Variation and evolution in plants ; (New York : Columbia University Press)
fatebe T 1939 Genetic and cytolagical studies OB the Fx hybrid of scarlet or tomato egg plant

(Solanum integrifolium Poir) X the egg plant (Solanum melongena L.) ; Bot. Mag. (Tokyo)

50 457-462
/eerabhadra Rao S 1977 Studies on the crossability relationships of some spinous solanums ;

Unpublished Ph.D. thesis submitted to the Andhra University, Waltair.
^eerabhadra Rao S and Rao B G S 1977a Screening of flowers in interspecific crosses of some

spinous solatium species ; Curr. Sci. 46 123-124
Veerabhadra Rao S and Rao B G S I977b Chromosome pairing in the Solanum surattense X

S. melongena F± heterozygote ; Curr. Sci. 46 124-125
Veerabhadra Rao S and Rao B G S 1977c Chromosomal repatterning ia the differentiation of

two spinous solanums ; Curr. Sci. 46 458-459
ftitshi U 1967 Interspecific hybrids in solanum I. Solanum indicum L. and S. incanum

L. Proc. Indian Acad. Sci. 65 111-113

Proc. Indian Acad. Sci. (Plant Sci.), Vol. 91, Number 2, April 1982, pp. 93-100.
© Printed in India.

Groundnut rust— its survival and carry-over in India4'

P SUBRAHMANYAM and D McDONALD

Groundnut Pathology, International Crops Research Institute for the Semi-Arid
Tropics, ICRISAT, Patancheru 502 324, India

MS received 27 December 1980

Abstract Groundnut rust has become an important disease in India, particularly
in the South, probably because of extensive and continuous cultivation of the crop.
Uredospores present on crop debris in the field, and on pods or seeds in storage
at ambient temperatures, lost viability within 6 weeks. They retained viability for
long periods when stored at — 16° C. Neither teliospores nor any collateral or
alternate hosts were found. Seeds heavily contaminated with viable uredospores
and sown in sterile soil gave rise to disease-free seedlings. There should be no
risk of spread of rust from properly treated seed samples.

Keywords. Groundnut rust ; survival ; carry-over ; Puccinia arachidis Speg. ;
Arachis hypogaea L.

1. Introduction

Rust of groundnut (Arachis hypogaea L.), caused by Puccinia arachidis Speg., was
reported from Punjab, India, in 19619 (Chahal and Chohan 1971) and now occurs
in most groundnut-growing Indian States (Snbrahmanyam et al 1979). The
disease has become particularly important in South India, where groundnuts are
grown for nrich of the year and where conditions favour development and spread
of the pathogen. This paper deals with the survival of the rust fungus and presents
results of investigations on possible carry-over of the disease in crop debris, on
seeds, and on weeds. The biology of the fungus is discussed in relation to distri-
bution of rust and groundnut cropping seasons.

2. Materials and methods

2.1. Survival of uredospores in crop debris

Dried haulms of groundnut collected from rust-infected rain-fed and irrigated
crops (cv. TMV-2) during 1976-78 were immediately exposed to weather by
spreading them in shallow layers in the field at ICRISAT Centre farm. At
intervals, uredospores were collected from the crop debris (dried haulms), suspended
in sterile distilled water on glass slides, and incubated in the dark at 25° C. After
6hr, 1,000 spores were checked for germination.

* Submitted as Journal Article No. 125 by the International Crops Research Institute for the
Semi-Arid Tropics (ICRISAT).

93

94 P Subrahmanyam and D Me Donald

2-2. Effect of temperature on uredospore longevity

Uredospores, freshly collectee from infected plants, were placed in glass vials and
stored at temperatures of - 16, 6, 25 and 40° C. At intervals, they were sampled
and tested for viability as described above.

2-3. Presence of uredospores on pods and seeds

Pods were collected from a crop with severe rust and separated into those with
no shell damage and those with shells broken during threshing. Undamaged
pods were shaken in distilled water to which Tween 80 (1 : 1000) had been
added, and washings were centrifuged at 2,000 rev/min for 1 hr. The pellet
obtained was examined microscopically for uredospores. t>amaged pods were
carefully opened and seeds were removed with minimal contact with the outside
of the shells. The seeds were washed and the washings examined as. described
for undamaged pods.

2-4. Longevity of uredospores on stored seed

Seeds were dusted with freshly collected uredospores and stored in cloth bags in
the laboratory at 25 to 30° C. Samples were removed at 5-day intervals and
uredospores washed off the seeds and their viability tested as described above.

2-5. Carry-over of rust on seed

Seeds of rust-susceptible cultivar TMV-^2 were surface-sterilised by immersion for
5 min in a 0-1% aqueous solution of mercuric chloride to which a small
amount of Tween-80 had been added. They were then washed in repeated changes
of sterile tap water. Isolation plant propagators (Burkard Manufacturing
Company, England) were prepared containing steam-sterilised garden soil in
pots which could be watered from below with sterile tap water. Into the pots
in one unit, 200 seeds, were aseptically sown. In another unit, 200 seeds liberally
coated with freshly collected uredospores were sown. A further 200 seeds were
aseptically sown in a third unit, and after germination, the seedlings were dusted
with freshly collected uredospores. Seedlings were checked for rust infection.

2-6. Germination of uredospores on germinating seeds

Two-day-old seedlings of the cultivar TMV-2 were carefully washed, testas
removed, and 100 cotyledons and 50 radicles excised. These organs were surface*
inoculated with a suspension of uredospores and placed in moist chambers for
incubation in the dark at 25° C. Samples were removed after 24 hr, stained,
and examined under the microscope. In another test, artificially-contaminated
seeds were sown in sterile soil, and resulting seedlings were carefully removed
and examined at intei-vals.

2.7, Search for teliospores and collateral hosts

A large number of specimens of rust-infected groundnut from different parts of
the country were examined for the presence of teliospores. Some 2,000 entries
from the ICRISAT groundnut germplasm collection were also examined at various

Groundnut rust — survival in India 95

stages of development under severe rust infection. Attempts were also made to
induce telial production by growing rust-infected plants of the susceptible TMV-2
cultivar under the following combinations of temperature and day length in plant
growth chambers.

Treatment Day temperature Night temperature Day length
(°C) (°C) (hr)

1

20

10

8

2

30

10

8

3

30

20

10

4

35

25

12

5

40

30

12

6

25

25

12

7

15

15

12

Various common crop and weed plants growing in or near fields of rust-infected
groundnuts on the ICRISAT farm and farmers* fields were examined for rust.
Some were also subjected to inoculation with uredospores in greenhouse tests ;
the groundnut cultivar TMV-2 was used as a susceptible check.

3. Results and discussion

3.1. Survival of uredospores in crop debris

The high initial viability of uredospores decreased rapidly with exposure to weather
(table 1). This was most marked in uredospores from irrigated crops, probably
because of the higher temperatures experienced in May than in the November-to-
January period following the rain -fed crop. Invariably, uredospores on exposed
crop debris lost all viability within 30 days. Similar work in other parts of India
also indicates that uredospores are short-lived in crop debris under field condi-
tions (Lingaraju etal 1979 ; Mallaiah and Rao, personal communication).

3-2. Effect of temperature on uredospore viability

Spores remained viable for several months when stored at low temperature (— 16° C)
while at 40° C they lost viability within 5 days (table 2). At the intermediate
temperatures, viability decreased with time of storage and was completely lost
within about 2 months. Mallaiah and Rao (personal communication) found that
uredospores remained viable for up to 4 weeks when stored at temperatures
below 30° C but lost viability within 2 weeks when stored at temperatures above
35° C. It would thus appear that temperature is an important factor influencing
viability and longevity of rust uredospores.

3.3. Carryover and distribution on seed

Garry-over and dissemination of uredospores on groundnut pods and seeds have
been suggested. Peregrine (1971) indicated -that movement of contaminated

96 P Subrahmanyam and D McDonald

Table 1. Viability of uredospores after various periods of exposure to weather on
infected crop debris.

Percentage of uredospores viable*

T> * J f «•»

jrenouL 01 exposure • —

GO

Rainy-season crops

Post-rainy-season crops

1976 1977

1976-77

1977-78

0

65 90

82

89

6

36 74

9

0

14

1 42

1

1

20

0 26

0

0

22

0 10

0

0

26

0 0

0

0

Period of test

13-12-1976 7-11-1977

4-5-1977

2-5-1978

to to

to

to

7-1 -1977 2-12-1977

30-5-1977

28-5-1978

RH% 0714 hr

80-7 83-5

60-7

60-7

1414 hr

26-0 46-6

26-9

23-9

Temp. °C : Max.

28-3 28-0

37-6

39-7

Min.

13-4 19-5

24-9

25-6

* 1,000 spores per sample.

Figures to nearest whole number

•

Table 2. Effects

of storage temperature on viability of uredospores.

Storage

Percentage* of uredospores viable after storage for

<°C)

Days

5

13 28 40 48 60

70 78 99

110 120

—16 88

82 89 90 98 88

92 93 92

94 93

6 84

85 82 35 15 4

0 00

• •• »« .

25 81

88 80 24 0 0

000

• •• .* *

40 0

00000

000

* 1,000 spores per sample. Figures to nearest whole number.

seed may have been involved in the spread of ru&t to Brunei . Pods from a rust-
infected crop would be contaminated with spores during threshing and any
damage to shells could well lead to contamination of seeds. Seeds could alsobe
contaminated during shelling. Examination of pods from a severely rusted crop
showed presence of uredospores on the shells. Where shells were broken, uredo*
spores were found on the seed surfaces.

Groundnut rust — survival in India 97

Table 3. Effects of storage at room temperature (25-30° C) on viability of uredosporcs.

Percentage* of uredosporcs viable after storage for :
Days

0 5 10 15 20 25 30 35 40 45 50 55
95 72 30 28 25 28 30 29 39 10 0 0

*1,000 spores per sample. Figures to nearest whole number.

The viability of uredospores on seed stored at room temperature for varying
engths of time is shown in table 3. Viability decreased rapidly with storage
:ime from an initial 95% to zero after 45 days.

Surface-sterilised seeds of cultivar TMV-2 sown in sterile soil in isolation plant
>ropagators gave rust-free seedlings. Seeds similarly treated, but coated with
viable uredospores prior to sowing, also gave rise to rust-free seedlings. A ' check '
xeatment where the foliage of seedlings was dusted with uredospores resulted in
severe rust disease within 25 days of sowing. This supports the argument that
,urface contamination of seeds with uredospores is unlikely to result in rust
nfection of seedlings.

When excised cotyledons and radicles of germinating seedlings, were surface-
noculated with uredospores and incubated in the dark, the spores germinated
ind appressoria were produced, but there was no development of disease. Exami-
lation of seedlings from seeds heavily contaminated with uredospores and sown
n sterile soil again showed germinated uredospores with appressoria, but no rust
leveloped.

There would appear to be little danger of rust disease developing, from uredo-
;pores carried on sown seed. Also, there is no authenticated report of the
'ust fungus being internally seed-^borne.

Although rust has spread rapidly to most parts of the world in recent years
Hammons 1977 ; Subrahmanyam etal 1979), there are still some groundnut-
growing areas where it is not present. Plant quarantine authorities and those
Concerned with distribution of groundnut germplasmare understandably concerned
vith the possible spread of the disease to these areas through contaminated seed
amples. However, the practice of dressing seed with fungicides, the rapid loss
>f viability of uredospores at ordinary temperatures and their inability to infect
eeds or germinating seedlings below ground all indicate that disease spread
hrough properly treated and handled seed samples is extremely unlikely. To
>btain successful spread, viable uredospores would have to be carried to the
urface of foliage of the susceptible plant under environmental conditions
Conducive to infection. This is more likely to happen due to long-distance air
lispersal or contamination on clothes and baggage of air travellers than on
>roperly treated seed samples.

J .4. Biology of the rust fungus

Oie pathogen is known almost exclusively by its uredial stage. There are a few
:ecords of the occurrence of the telial stage on cultivated Arachis hypogaea in

P.(B)-2

"38 P Subrahmanyam and D McDonald

South- America (Spegazzini 1884 'J Hennen etal 1976) and on wild Arachis spp.
(Guarch 1941 ; Bromfield 1971). In India, Chahal and Chohan (1971) recorded
the occurrence of teliospores on groundnut leaves but gave no details of spore
morphology and the disease has not recurred in Punjab. There has been no
other authenticated report of the occurrence of teliospores of groundnut rust.

We have examined many specimens of rust-infected groundnuts from different
parts of India but have found only uredospores. Some 2,000 entries from the
ICRISAT groundnut germplasm collection were examined at various stages of
development under severe rust infection, but again only the uredial stage of the
rust was found.

Attempts were made to induce teliospore production by growing rust-infected
plants under various combinations of temperature and day length but were
unsuccessful It is not known if the fungus can produce pycnia and aecia or if
any alternate host is involved in the life cycle, It would appear that uredospores
are the main, if not the only, means of dissemination of the groundnut rust
' fungus.

Table 4. Plant species examined as possible collateral hosts of rust.

"Leguminous crop plants

• Cajdnus cajan (L.)-Millsp.

Canavalia gladiata DC.

Cicer arietinum L.

Crotalaria juncea L.

Cyamopsis tetragonoloba (L.) Taub.
' Gly cine max (L.) Men.

Lablab purpureus (L.) Sweet

Lens culinaris Medik.

Phaseolus lunatus L.

P. vulgaris L.

Sesbania sp.

Vidafaba L.

Vigna mungp (L.) Hepper
, y. radiata (L.) Wilcz.

Leguminous weeds

Aeschynomene aspera L.

>A. Mica L. .

Alysicarpus monilifer (L.) DC.

.Cassia torn L.

Ihdigofera hirsute L.

Stylosanthes fruticosa (Retz.) Alston

Tephrosia- hirta Ham. ... , .

T. purpurea (L.) Pers.

Zornia diphylla (L.) Pers.

Non-legumes

Acanthospermum hispidum DC.

Achy rant lies asp era L.

Aerva monsoniae (L.F.) Mart.

Amaranthus viridis L.

Anisomeles indica (L.) O. Ktze.

Boerhaavia diffusa L.

Cathamnthus pusillus (Murr.) G. Don

Cor chorus aestuans L.

Cypems compressus L.

C. rotundus L.

Dactyloctenium aegyptium (L.) Beauv.

Digitaria ciliaris (Retz.) Koeler

Eclipta alba (L.) Hassk.

Euphorbia hirta L.

Evolvulus alsinoides (L.) L.

Ipomoea tridentata Roth

Lactuca hastata DC.

Lagascea mollis Cav.

Leucas lavandulifolia Sm.

Micrococca mercurialis Bth.

Mollugo pentajphylla L.

Ocimunt americanum L.

Panicum sp.

Phyllanthus niruri L.

Portulcica oleracea L. ' •

P. quadrifida L.

Sida sp.

Trianthema portulacastrum L.

Tridax procumbens L,

Groundnut rust— survival in India

99

There is no record of the occurrence of any collateral hosts of groundnut rust
outside the genus Arachis, and in India wild Arachis spp. occur only in research
centres and can hardly be involved in perpetuation of the disease. The possible
occurrence of other hosts was considered, and various common crop and weed
plants growing close to or within fields of rust-infected groundnuts (table 4) were
regularly examined for the presence of rust, but no case of infection was found
Some of these plants were also subjected to inoculation with rust urcdospores in
greenhouse tests, but again no case of infection was recorded.

3.5. Cropping seasons and rust survival and spread

There is no uniform groundnut growing season in India. In some of the southern
Sttates, particularly Andhra Pradesh, Tamil Nadu and Rarnataka, groundnuts
are grown in some areas throughout the year (figure 1), presenting excellent
opportunity for survival of rust. About 90% of the crop is grown in the rainy
season, most of the rest is gjcown in the post-brainy dry season under irrigation.
In some places a summer crop is grown.

Rust attack is most severe on the rainy-season crops but can still be noticeable
on dry^season crops. The disease has been seen on the summer crop in parts of
Andhra Pradesh, but pustules developed very slowly and did not sporulate until
the coming of the monsoon rains, when the disease developed rapidly on the
maturing crop.

On the rainy-season crop, the disease appears in July and August in South India,
in September in Central India, and in October in North India (May^e et al 1977).
In Central and North India normally only a rainy-season crop is grown, and it is
thought that the groudnut crops in South India may act as a reservoir of rust
disease from which spores are carried by the monsoon winds to infect the crops
in the north. The present trend towards increased cultivation of groundnuts in
southern India, particularly the irrigated dry-season crops, could result in more
effective carry-over and spread of rust disease within the country.

Figure 1. Groundnut cropping seasons in India.

100 P Subrahmanyam and D McDonald

References

Bromfield K R 1971 Peanut rust— A review of literature ; J. Am. Peanut Res. Educ. Assoc

3 111-121

Chahal D S and Chohan J S 1971 Puccinia rust on groundnut ; FAO Plant Prot. Bull. 19 90
Guarch A M 1941 Communicaciones fitopatologicas ; Rev. Fac. Agron. Univ. Montevideo,

23 14-16 (Abst. in Rev. Appl. Mycol. 1942 21 129-130)

Mammons R O 1977 Groundnut rust in the United States and the Caribbean ; PANS 23 300-304
Hennen J F, Figucredo M B, Riberio I J A and Soave J 1976 The occurrence of tcliospores

of Puccinia arachidis (Uredinales) on Arachis hypogaea in Sao Paulo State, Brazil ; Sununa

Phytopathol 2 44-46
Lingaraju S, Siddaramaiah A L and Hegde R K 1979 Viability and survival of urcdosporcs

of Puccinia arachidis Speg. in Dharwad ; Curr. Res. 8 68-69
Mayee C D, Godbole G M and PatiJ F A 1977 Appraisal of groundnut rust in India : problems

and approach ; PANS 23 162-165

Peregrine W T J 1971 Groundnut rust (Puccinia arachidis) in Brunei ; PANS 19 318-319
Spegazzini C L 1884 Fungi guaranitici I ; Anal Soc. Ci. Argentina 17 90
Subrahmanyam P, Reddy D V R, Gibbons R W, Rao V R and Garrcn K H 1979 Current

distribution of groundnut rust in India ; PANS 25 25-29

Proc, Indian Acad. Sci. (Plant Sci.), Vol. 91, Number 2, April 1982, pp. 101-106.
© Printed in Indif>.

Correlated promotion of ray-floret growth in chrysanthemum
by potassium chloride, gibberellic acid and sucrose

P PARDHA SARADHI and H Y MOHAN RAM

Department of Botany, University of Delhi, Dellii 110007, India

MS received 7 October 1981

Abstract. The role of 10~2 M, 2 X 10~2 M, 4 x 1Q-2 M patassium chloride, gibberellic
acid(10-GM; GA3) and sucrose (5 x 10"2M) (used individually and in various combi-
nations) in the elattgatio,n growth of excised ray-florets of Chrysanthemum mori-
folium var. Jyothsna, was investigated. KC1 (10~2M) caused 33-3% increase in
elongation as compared to control (16-7%). With GA3 and sucrose the percentage
of elongation recorded was 39-8 and 28-9 respectively. Maximal growth response
(£2-8%) was recorded in KC1 (4 X 10~2M) -f GA3(10-BM) -i-sucrose (5 X 10~2M).
When used in combination either with GA3 or sucrose, KC1 showed an almost
additive effect, whereas in the presence of bath it acted synergistically. It is
inferred that the increased turgor resulting from sucrose-promoted potassium uptake
along with GA3-caused tissue extensibility accounts for enhanced floret growth,

Keywords. Chrysanthemum ; cell elongation ; flower growth ; gibberellic

potassium chloride ; sucrose,

1. Introduction

Studies on the opening of flowers harvested at the immature stages of development
(bud-cut flowers) have gained importance owing to several commercial advantages
(Marousky 1971 ; Halevy and Mayak 1974). Sucrose and gibberellins have been
used for promoting flower bud opening in chrysanthemums and gladioli (Marousky
1971, 1972 ; Bravdo etal 1974 ; Rao and Mohan Ram 1979). In several bud-cut
flowers, however, further opening is a serious problem. It is envisaged that the
difficulties faced in causing the opening of bud-cut flowers can be overcome after
the basic processes controlling petal growth have been understood.

There has been a good amount of physiological work on flower initiation and
senescence. In comparison, literature on flower growth is scanty. The investi-
gations on cell and organ expansion have been largely confined to vegetative parts.
These events are known to be under the control of turgor pressure, viscoelastic
properties of the wall and cell wall synthesis. Ions such as K"1* are crucial in
regulating osmotic potential of the cell sap (Haschke and Luttge 1975 ; Parrish
and Davies 1977 ; Stuart and Jones 1977, 1978). Sugars have been shown to be
involved in the synthesis of wall precursors, besides regulating osmotic potential
and providing energy (Siegelman etal 1958). Gibberellins regulate the visco-
elastic properties of cell wall (Kamisaka et al 1972 ; Adams etal 1975 ; Nakamura

101

102 P Pardha Saradhi and H Y Mohan Ram

et al 1975 ; Kawamura et al 1976 ; Coartney arid Morre 1980) beside.s promoting
the influx of solutes (Katsumi and Kazama 1978). With this information in the
background, a study of the role of KC1, gibberellic acid and sucrose on ray-floret
elongation in chrysanthemums was taken up.

2. Material and methods

Stocks of Chrysanthemum morifolium var. " Jyothsna " (Asteraceae) were procured
from the National Botanical Research Institute, Lucknow, and were grown in the
Botanical Garden of the Department. The plants flowered profusely in December.
Capitula (measuring approximately 14 mm in diameter) in which the ray-floret
would become visible the next day were used for experimentation. Ray-florets
belonging to the outer two whorls and measuring 9 or 9-5 mm were excised from
the capitula. For each treatment 20 ray-florets, in groups of five, were floated in
petri plates containing 30 ml of the test solution and kept under continuous light
(cool-white fluorescent tubes ; 1200 Lux) at 20 ± 2° 0. The test solutions con-
sisted of distilled water (control) ; potassium chloride (KC1) at 10~2 M, 2 x
10~2 M and 4 x 10~2 M ; gibberellic acid (GA3) at 10~5 M ; sucrose (S) at 5 x
10-2M ; KC1(10-2 M) + S (5 x 10~2 M) ; KC1 (2 x 10~2 M) + S (5 x 10~2M) -
KC1 (4 x 10~2 M) + S (5 x 10~2 M) ; KC1 (10~2 M) + GA3 (1Q-5 M) ; KC1
(2 x 10-2M) + GA3 (lO^M) ; KCl (4 x 10~2 M) + GA3 (10~5M) ; KC1
(10-2 M) + S(5 x 10~2M) + GA3(10-5M) ; KC1 (2 x 10~2M) -H S (5 x 10~2M)
+ GA3 (10-5 M) ; KC1 (4 x 10~2 M) + S (5 x 10~2 M) 4- GA3 (10~5 M). Strepto.
Aiycin (25 ppm) was added to all the solutions to prevent microbial infection.
The length of the floret was measured every 24 hr to the nearest 0-1 mm. Each
experiment lasted 10 days.

3. Observations

In all the treatments (including the control) the florets showed an increase in
length with- time (table 1). Among the three concentrations of KCl used, the
florets showed the maximum response with 10~2 M (33 • 3 %), followed by 2 x 10""2 M
(22-2%) and 4 x 10"2 M (20-0%) by day 5.

• The floret length increased by 28-9% with sucrose and by 49-8% with GA8,
the -maximum length having been attained on day 6. When KCl was used in
combination with GA3 or sucrose, greater elongation of the florets than in the
treatments with only KCl, sucrose or GA3 was recorded. When KCl (all the
three concentrations) and sucrose were present together, the elongation ranged
from .46 -.2%. to 49-5%. In the presence of GA3, KCl (2 x 10~2M) showed as
high as 57 % increase in length. With KCl (1Q-2 M) 4- GA8 and KCl (4 x 1Q-2 M)
rf- GAS the corresponding values were 50-0 and 47-3% respectively.

- The florets exhibited highest elongation when they were kept in a mixture
containing -KC1, sucrose and GA3. The increase in length observed over the
control was 82r8% when the concentration of KCl in the combination was
4 x. iJOrfM. However, 10~2M KCl and 2x 10-2M KCl also showed large
increases up to 65 -6 and 72%, respectively. With the exception of KCl (10~2M)

Promotion of ray-floret growth in chrysanthemum

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104 P Pardha Saradhi and H Y Mohan Ram

-t- GA3, in which the florets showed an increase in length till day 8, in the other
combinations floret elongation continued until the termination of the experiment.
It is interesting that the effect elicited by KC1 in combination with either sucrose
or GA3 was nearly additive and that in combination with both was synergistic.

4. Discussion

In the present work it was noted that KC1 caused greater elongation at low rather
than high concentration. Potassium has been shown to play an important role
in the elongation of vegetative tissues by altering the osmotic potential (Stuart
and Jones 1977, 1978), by effecting wall loosening (Tagawa and Bonner 1957 ;
Haschke and Luttge 1975), by acidification of the incubation medium (Thimann
and Schneider 1938 ; Ordin etal 1956 ; Tagawa and Bonner 1957 ; Haschke and
Luttge 1975) or by acting as a co-factor for stimulating certain enzymes (Mahler
1961 ; Purves 1966).

In the presence of sucrose the florets showed 12-2% higher increase in length
than that observed in the control. Besides acting as a source of energy and in
providing building blocks for cell wall synthesis, sucrose is probably involved in
osmoregulation, providing the necessary force for the maintenance of turgidity
in the elongating ray-florets of chrysanthemums (present work) as has been shown
for other flowers (Winkenbach and Matile 1970 ; Dilley and Carpenter 1975).
Additionally, in the present investigation, sucrose showed better response in combi-
nation with KC1. There is evidence that the energy for K4 ion uptake and for
the probable production of carbon skeletons for organic anions that move with
K+ ions is derived from sucrose (Satter et al 1976). Setter etal (1976) have
suggested that the rhythmically controlled sucrose permeation in Samanea pulvini
could regulate sucrose-H+ transport and thus, in order, membrane potential, salt
flux and water flux resulting in increased turgor.

The enhanced ray-floret elongation observed in the present study in the presence
of GA3 could emanate from its effect on the viscoelastic properties of the cell
(i.e., cell wall extensibility) (Kamisaka etal 1972 ; Adams etal 1975 ; Coartney
and Morre 1980), osmoregulation (Kazama and Katsumi 1973) or the synthesis
of cell wall material (McComb 1966; Srivastava etal 1975) as demonstrated
in several other systems.

In the present work a greater elongation of the ray-florets was noted in response
to KC1 + sucrose + GA3 over that with KC1 + GA3 or KC1 + sucrose. When
used in combination either with GA3 or sucrose, KC1 showed an almost additive
effect, whereas in the presence of both it acted synergistically. A combined effect
of sucrose and GA3 has also been noted in the stimulation of elongation of
bypocotyl segments (Purves and Hillman 1958 ; Kazama and Katsumi 1973),
in the linear growth of staminal filaments (Murakami 1973) and in flower growth
and opening (Rao and Mohan Ram 1979).

GA3 perhaps enhances ATPa.se activity. This could regulate K+ ion and
sucrose influx (Katsumi and Kazama 1978) thereby regulating osmotic potential
and turgor pressure leading to ray-floret elongation in chrysanthemum,

Promotion of ray-floret growth in chrysanthemum 105

Acknowledgements

The authors thank Dr I V Ramanuja Rao for his critical comments. Financial
assistance provided by the Council of Scientific and Industrial Research, New
Delhi, to one of the authors (PPS) is gratefully acknowledged.

References

Adams P A, Montague M J, Tepfer M, Raylc D C, Ikuma H and Kaufman P B 1975

Effect of gibberellic acid on plasticity and elasticity of Avena stem segments ; PL PhysioL

56 547-554
Bravdo B, Mayak S and Gravrieli Y 1974 Sucrose and water uptake from concentrated sucrose

solutions by gladiolus shoots and effect of these treatments on floret life ; Can. J. Sot. 52

1271-1281
Coartney J S and Morre D J 1980 Studies on the role af wall extensibility in the control of

cell expansion ; Bat. Gaz. 141 56-62
Dilley B R and Carpenter W J 1975 The role of chemical adjuvants and ethylcne synthesis on

cut flower longevity; Acta Horticultiirae 47 117-132
Halevy A H and Mayak S 1974 Improvement of cut flower quality, opening and longevity by

pre-shipment treatments; Acta Horticultwae 43 335-347

Haschke H R and Liittge U 1975 Stoichioraetric correlation of malate accumulation with auxin-
dependent IC^-H*1 exchange and growth in Avena coleoptile segments ; PL Physiol. 56

696-698
Kamisaka S, Sano H, Katsurni M and Masuda Y 1972 Effects of cyclic AMP and gibberellic

acid on lettuce hypocotyl elongation and mechanical properties of its cell wall ; PL Cell

Physiol. 13 167-173
Katsumi M and Kazarna H 1978 Gibberellin control of cell elongation in cucumber hypoco-tyl

sections : Bot. Mag. Tokyo Special Issue 1 141-158
Kawanmra H, Kamisaka S and Masuda Y 1976 Regulation of lettuce hypocotyl elongation

by gibberellic acid. Correlation between cell elongation, stress relaxation properties of the

cell polysaccharide content ; PL Cell Physiol. 17 23-34
Kazama H and Katsumi M 1973 Auxin-gibberellin relationship in their effects on hypocotyl

elongation of light-grown cucumber seedlings : Responses of sections to auxin, gibberellin

and sacro.se ; PL Cell Physiol. 14 449-458
Mahler H R 1961 Interrelationships with enzymes ; in Mineral metabolism (eds.) C L Coxnar

and F Bronner (New York : Academic Press) IB 743-879
Marousky F J 1971 Handling and opening cut-chrysanthemum flowers from bud stage with

8-hydroxyquinoline citrate and sucrose ; USDA Agr. Mktg. Res. Rept. p. 905
Marousky F J 1972 Water relations, effects of floral preservatives on bud opening and keeping

quality of cut flowers; ffortsci. 1 114-116
McComb A J 1966 The stimulation by gibberellic acid of cell wall synthesis in the dwarf pea

plant: Ann. Bot. 30 155-163
Murakami Y 1973 The role of gibberellins in the growth of floral organs of Pharbitis nil; PI.

Cell Physiol. U 91-102
Nakamura T, Sekine S, Arai K and Takahas-hi N 1975 Effects of gibberellic acid and indole-

3-acetic acid on stress-relaxation properties of pea hook cell wall ; PL Celt Physiol. 16

127-138
Ordin L, Applewhite T H and Bonner J 1956 Auxin induced water uptake by Avena coleoptile

sections ; PL Physiol. 31 44-53
Parrisli D J and Davies P J 1977 On the relationship between extracellular pH and the growth

of excised pea stem segments ; PL Physiol. 59 574-573

106 p Pardha Saradhi and H Y Mohan Ram

Purves W K 1966 Monovalent cations and growth regulation. I. Growth responses in cucumber

hypocotyl segments ; PL Physiol. 41 230-233
Purves W K and Hillman W S 1958 Response of pea stem sections to indoleacetic acid, gibberellic

acid and sucrose as affected by length and distance from apex ; Physiologia PL 11 29-35
Rao. I V Ramanuja and Mohan Ram H Y 1979 Interaction of gibberellin and sucrose in flower

bud opening in gladiolus ; Indian J. Exptl. BioL 17 447-448
Satter R L, Applewhite P B and Galston A W 1976 Pfr phytochrome and sucrose requirement

for rhythmic leaflet movement in Albizzia ; Photochem. Protobiol. 23 107-112
Siegelman H W, Chow C T and Biale J B 1958 Respiration of developing rose petals ; PL

Physiol. 33 403-409
Srivastava L M, Sawhney V K and Taylor I E P 1975 Gibberellic acid induced cell elongation

in lettuce hypocotyls ; Proc. NatL Acad. Set., USA 72 1107-1111

Stuart D A and Janes R L 1977 The roles of extensibility and turgor in gibberellin- and dark-
stimulated growth; PL Physiol, 59 61-68
Stuart D A and Jones R L 197& The role of acidification in gibberellic acid and fusicoccin

induced elongation growth of lettuce hypocotyl sections; Planta 142 135-145
Tagawa T and Bonner J 1957 Mechanical properties of the Avena coleoptile as related to auxin

and to ionic interactions ; PL Physiol. 32 207-212
Thimann K V and Schneider C L 1938 Differential growth in plant tissues ; Am. J. Bot. 25

627-641
Winkenbach F and M.itile Pit 1970 Evidence for de novo synthesis of an invertase inhibitor

in senescing petals of Ipomoea ; Z, Pfl. Physiol. 63 292-295

Prac. Indian Acad. Sci. (Plant Sci.), Vol. 91, Number 2, April 1982, pp. 107-114
© Printed in India.

Nuclear behaviour during heartwood formation in Acacia
auriculiformis A. Cann.

K V BHAT and J D PATEL

Department of Biosciences, Sardar Patel University, Vallabh Vidyanagar 3S8 120,
India

MS received 15 September 1930 ; revised 23 March 1982

Abstract. Nuclear behaviour is studied in Acacia auriculiformis A. Cann. with
reference to aging in both axial and ray parenchyma cells (contiguous to vessels
and away from the vessels). The size of the nucleus and nucleolus reduces gradually
towards the inner sap wood and at last they disappear at the heartwood boundary.
Nuclei show lobing, fissuring, fragmentation and contraction in this zone prior to
their disintegration. Thus a gradual loss of vitality of parenchyma cells during
aging is noted. The parenchyma cells contiguous to vessels seem to be more actively
involved in formation of heartwood extractives.

Keywords. Nuclear disintegration ; nucleolus ; nucleus ; slenderness ratio ;
transition wood ; Acacia auriculiformis.

I,'. Introduction

Ths transformation of sapwood into heartwood during aging involves loss of proto-
plasm of living cells and concomitant accumulation of the extractives. The
parenchyma cells of the sapwood are known to play a major role in heartwood
formation as in the majority of the species they are the only living cells in the
sapwood (exceptionally living fibres occur in the sapwood). The physiological
status of the living parenchyma cells during different stages of aging is still a matter
of controversy. Evaluation of the vitality of the parenchyma cells and the rate
of metabolic activities is very important and can be done only by adequate -methods.
Bosshard (1966) considers that the form and dimensions of the nuclei are very
valuable indications for tracing the activity of a cell. Alteration of nuclear con-
figurations, in conifers, as a part of necrobiosis of storage tissue can be of great
use in studying the heartwood formation. This can be studied in two ways : by
determining the degree of slenderness ratio, and by calculating the nuclear surfaces
(Bosshard 1965). Although nuclear behaviour in ray cells has been studied in
detail, information on changes in shape and size of the nucleus in the parenchyma
contiguous to the vessels is not available. As there is a remarkable difference in
the nuclear, size and shape in, the cells contiguous to vessels and those away from
the vessels, the nuclear behaviour is studied in relation to the* above situations in
both axialand ray parenchyma cells of Acacia auriculiformis A/Cann. Thenuclear
disintegration has also been studied.

107

108 K V Rhat and J D Patel

2, Material and methods

The wood material of A. auriculiformis A. Gann. was collected from the University
botanical garden and was flxed in FAA (Sass 1958). The fixed material was
divided into blocks of suitable sizs for microtomy. No intermediate block was
discarded so that continuity was maintained from cambium to heartwood. The
radial strips of wood were divided into outer sapwood, middle sapwood, inner sap-
wood and sapwood-heartwood boundary or transition. Radial sections of 15 /mi
thichness were cut from each of the blocte and stained with toluidine blue *O*
(O'brien et al 1964), and pyron in -methyl green (Brachet 1953). Camera lucida
drawings of 100 nuclei each from axial and ray parenchyma cells (contiguous as
well as away from the vessels) from each zone of the wood were made on a piece
of paper using an oil immersion object The length, width and sectional area of
the nucleus were measured using area calculating device (ACD) (Chavan et al
1979). The slenderness ratio of the nuclei was obtained by dividing the average
length by average width of the nucleus.

3. Observations

The ray and axial parenchyma cells of sapwood contain nuclei but in fibres the
nuclei disappear in the outermost sapwood. For brevity we avoid the terms axial
and ray parenchyma cells in this paper, instead they are referred to as cells at many
places except where the two types of cells are to be specifically mentioned. The
chromaticity of the nucleus is comparatively higher in parenchyma cells contiguous
to vessels (figures 1-3, 7-11) as compared to those away from the vessels (figures
4-6, 12-14). However, the chromaticit} of the nuclei in general increases in the
cells of the inner sapwood (figures 3, 6, 11, 14). At the heartwood boundary
the nuclei show the highest chromaticity (figures 15-25).

Usually the nuclei are uninucleolate, but occasionally bimicleolate nuclei are
also observed (figure 1). The nucleolus is very prominent in parenchyma of the
outer and middle sapwood, especiall} in the cells contiguous to vessels (figures
1, 7- 10). The size and distinctness of the nucleolus are reduced towards the inner
sapwood (figures 3, 6, 11, 14), and it becomes totally indistinct in the cells of
heartwood boundary (figures i5-25). The nucleolus is larger in the nuclei of cells
contiguous to vessels (figures I, 2, 7-10) than those of the cells which are not
contiguous to vessels (figures 4, 5, 12, 13).

The nuclei of the cells contiguous to vessels and those away from the vessels
show a great difference in their cross-sectional area and slenderness ratio (SR)
during various stages of transformation of sapwood into heartwood.

3.1. Nuclear area

The size of the nucleus as revealed by its sectional area differs from zone to
zone. Cells contiguous to vessels possess larger ilplei (figures 1-3, 7-H) than
the cells away from the vessel (figures 4-6, 12-14).

Heartwood formation in A. auriculiformis

.a. ............. a, : i|(jj|jiiBHilL

Figures 1-14. 1-3. Nucleus in ray cells contiguous to vessels. 4-6. Nuclei in
ray cells away from vessels. 7-11. Nuclei in axial parenchyma cells contiguous to
vessels. 12-14. Nuclei in axial parenchyma cells away frora vessels. 1, 4, 7, 12.
Outer sapwood. 2, 5, 8-10, 13. Middle sapwood, 3, 6, 11, 14. Inner sapwoo<t
Inserted scale, 12 *5 microns,

11.0

K V Shat and J D Paid

Figures 15-25. Nuclei at the heartwood boundary. 15-22. In ray cells. 23-25.
In axial parenchyma cells. 15-18. Lobed nuclei. 19, 20. Fissuring of the nucleus.
21. Condensed nuclei. 22. Nuclear fragments. 23. Lob ing of the nucleus. 24, Nuclear
fragmentation. 25. Contracted nucleus. Inserted scale, 12 -5 microns.

Heart-wood formation in A. auriculiformis

111

3. la. Nuclear area in parenchyma cells contiguous to vessels : Among the cells
contiguous to vessels the nuclei are larger in axial parenchyma (figures 7-10)
than those in the ray cells (figures 1, 2; table 1). There is a little and gradual
reduction in the nuclear size of ray cells as traced from outer sapwood to the
middle sapwood, but then the nuclei show sharp reduction in their size as traced
towards the inner sapwood (table 1). Further reduction in nuclear size from inner
sapwood to heartwood boundary is very little. On the other hand the nuclear
size in axial parenchyma gets reduced slightly as traced towards the middle
sapwood, and from middle sapwood to heartwood boundary through the inner
sapwood the nuclear size diminishes rapidly and rather uniformly (table 1).

3. Ib. Nuclear area in parenchyma away from vessels : The changes in the nuclear
size are not prominent in these cells (table 1). However, as traced towards the
heartwood boundary after a slight enlargement of the nucleus in ray cells from
outer to middle sapwood a slight reduction in its size follows from middle to inner
sapwood. Again at the heartwood boundary a slight increase in the nuclear size is
observed (table 1). In the ray cells of the heartwood boundary region the nuclei
are much lobed and slightly enlarged (figures 15-.18), On the other hand the size
of the nucleus in axial parenchyma cells decreases from outer sapwood to middle
sapwood. As traced from middle to inner sapwood the axial parenchyma cells
do not show appreciable reduction in their nuclear size, but from inner sapwood
to heartwood boundary there is appreciable reduction in nuclear size (table 1).
Thus, reduction in the nuclear size is observed from outer sapwood to heartwood
boundary except in ray cells away from the vessels. But, invariably there is a
reduction in the size of the nuclei from middle to inner sapwood.

3.2. The slenderness ratio (SR)

The nuclear SR (length/width) differs with the increasing radial distance from the
cambial zone. Slendcraess ratio serves as an index of the changes in nuclear shape
occurring simultaneously with the changes in nuclear size.

Table 1. Average sectional area (NA) in /nn2 and slenderness ratio (SR) of
nucleus in ray (RP) and axial parenchyma (AP) of various zones of wood.

Zone Parenchyma contiguous to vessels

nf

Parenchyma not contiguous to. vessels

wood Axial parenchyma Ray parenchyma

Axial parenchyma Ray parenchyma

NA

SR

NA

SR

NA

SR

NA

SR

OS

78

3-0

70

3-2

23

2-7

16

4-0

MS

75

4-3

58

4-4

19

2-2

20

3-9

IS

51

5-0

40

4-2

17

2-7

15

3-7

HB

32

4-9

37

4-2

13

3-0

20

3-4

HB, heartwood boundary ; IS, inner sapwood ; MS, middle sapwood ; OS, outer sapwood.
Data based an measurements of 8,000 nuclei.

112 K V Bhat and J D Fatel

3-2a. SR of nutfei in cells contiguous to vessels : A gradual elongation accom-
panied with a reduction in width is noted in the nuclei of ray (figures . 1,. 2) and
axial parenchyma (figures 7-40) from outer to middle sapwood. Hence, the SR
increases from outer to middle sapwood in ray and axial parenchyma cells (table 1).
The SR in ray cells does not change appreciably thereafter, but in axial parenchyma
it continues to increase up to the inner sapwood and then remains more or less
constant (table 1).

. 3.2b. SR in parenchyma celts away from the vessels : The SR of nuclei in ray
parenchyma cells gets reduced gradually towards the heartwood boundary (table 1).
Nonetheless, in the axial parenchyma the SR of the nucleus reduces abruptly as
traced from outer to middle sapwood, and from middle sapwood. to heartwood
boundary it increases in uniform and moderately steep manner (table 1).

3.3. Nuclear disintegration

In both ray and axial parenchyma cells the nuclei disintegrate at the heartwood
boundary. The nuclei in axial parenchyma disappear slightly earlier than those
of the ray cells. Prior to the disintegration the nuclei show some morphological
variations. In some ray cells they appear variously lobed (figures 15-il8). Con-
strictions appear along the length of the nucleus at one (figures 15, 16) or more
loci (figures 17, 18). Thus, the nuclei appear enlarged and convoluted. Nuclei
in some ray cells show longitudinal flssuring (figures 19, 20). whereas in others they
appear dense, spherical and highly contracted (figure 21). A few cells atthe heart*
wood boundary contain only fragments of nuclear material (figure 22).

The nuclei in axial parenchyma rarely show lobing prior to their disintegration
(figure 23), Fragmentation of the nucleus can be observed in some cells (figure
24). In a few axial parenchyma cells the nuclei appear very small and highly
wrinkled (figure 25).

4. Discussion

The cells with high activity and high vitality possess large nuclei and nucleoli
(Bosshard 1966). The nuclei in sapwood near cambium have larger surface than
in the sapwood adjacent to the heartwood (Bosshard 1965), Earlier studies on
nuclei of storage tissue of wood have considered slenderness ratio of the nucleus
and nuclear surfaces in cells in relation to their radial position from the cambium
(Frey-Wyssling and Bosshard 1959; Bosshaid 1965, 1966). But, these are not
studied in relation to their radial position from cambium, as well as in relation to
their contiguity to the vessels. The present study clearly indicates that the nuclei
of the cells contiguous to vessels are quite different in their size and morphology
from those belonging to the cells away from the vessels. The axial and ray paren-
chyma cells which are contiguous to vessels in the investigated species possess
larger nuclei and nucleoli as compared to those which are ^way from the vessels*
Further the chromaticity of these larger nuclei in cells contiguous to vessels is
comparatively high. A similar situation was found in other angiosperm species
investigate dearlier (Bhat and Patell980). It reveals that the cells in the contiguity
of vessels are more active than other living cells of the wood.

Heartwood formation in A. auriculifofmis • • • • -J-J3

* From outer sapwood to the heartwood boundary through the middle and inner
sapwood a gradual reduction in the size of the nuclei is observed, except in ray
cells away from the vessels. The reduction in nuclear size is more prominent in the
cells contiguous to vessels. The reduction in nuclear size is also accompanied by
a gradual reduction in nucleolar size and loss of its distinctness. These facts
suggest that the activity and vitality of the cells gradually dimmish during aging.
In exceptional cases nuclear enlargement in ray cells away from the vessels at the
heartwood boundary is not accompanied by a nucleolar enlargement. Bosshard
(1966) considered the dimension ofnucleolusas one of the indications of cell vita-
lity and reported increase in dimension of nucleoius in the transition zone between
the sapwood and heartwood. Fufcazawa and Higuchi (1966) observed a gradual
reduction in the RNA content from cambium to negligible amount at the inter-
mediate wood. As we could not Snd either increase in nucleolar size or RNA
content (as judged by staining reaction) at the heartwood boundary we consider
the nucleolar enlargement at the heartwood boundary in ray cells away from the
vessels to be merely a stage of its disorganisation and it is attributed to the intensive
lobing of the nucleus. Enlargement of the nucleus at the heartwood boundary,
prior to nuclear disorganization, was observed by us earlier in Ougeinia oojeinensis
and Garuga pinnata (Bhat and Patel 1980).

The data show that the degree of slenderness varies in different zones of the wood
with the decreasing size of the nucleus. A more prominent increase in the slen-
derness ratio is observed with the decreasing nuclear size in cells contiguous to
vessels, while in cells away from the vessels the change in the SR was not very pro-*
minent. In the ray cells away from the vessels the SR gradually decreases. In
our earlier observations also we had noted that the SR either increases or decreases
towards the heartwood boundary (Bhat and Patel 1980). The form of the nucleus
does not appear to be significant for tracing the activity of the cell. It should be
regarded together with the size (Bosshard 1966). Higuchi et al (1967) found a
gradual elongation of the nucleus in conifers with the increasing distance from the
cambial zone, and in angiosperms the variation in the shape of the nucleus was
found to be more or less dependent on the species. In the present species varia-
tion in nuclear shape appears to depend on the type of the cell and/or contiguity
of the cell to the vessels.

Histochemical observations in other angiosperm species have revealed that the
extractives are formed earlier in the cells contiguous to vessels than the others
(Bhat 1981). Thus, it appears probable from the data that the cells in contiguity
with the vessels are more actively involved in the metabolic activities involved in
the formation of heartwood extractives .

Acknowledgements

This work is supported by the University Grants Commission by a departmental
research project. KVB acknowledges a fellowship.

References

Bhat K V 19S1 Studies on heartwood formation in some Angiosperm trees ; Ph.D. thesis, Sardar
Patel University, Gujarat, India

•114 K V Mat and J D Patel

Bhat K V and Patel J D 1980 Nuclear studies in relation to heartwood formation in Ougeinia

oojeinensis and Garuga pinnata ; Caryologia 33 519-526
Bosshard H H 1965 Aspects of the aging process in cambium and xylem ; Holzforschmg 19

65-69
Bosshard H H 1966 Notes on the biology of heartwood formation ; Int. Assn. Wood Anatomists,

News Bull. 1 11-14
Brachet J 1953 The use of basic dyes and ribonuclease for the cytochemical detection of ribo-

nucleic acid ; Q. J. Micros c. Sci. 94 1-10
Chavan R R, Kothari I L and Patel J D 1979 A simple area calculating device (ACD) for

biological systems ; Curr. Sci. 48 792-793

Frey-Wyssling A and Bosshard H H 1959 Cytology of the ray cells in sapwood and heart-
wood ; Holzforschung 13 129-137
Fukazawa K and Higuchi T 1966 Studies on the mechanism of heartwood formation IV. RNA

content in the ray parenchyma cell ; /. Jpn. Wood Res. Soc. 12 221-226
HigucaiT, Fukazawa Kand Shimada M 1967 Biochemical studies on the heartwood formation ;

Res. Bull. Coll. Expt. Forests 25 167-192
O'brien T P, Fedder N and McCully M E 1964 Polychromatic staining of plant cell wall by

"toluidine blue 4O' ; Protoplasma 59 367-373
Sass J E 1958 Botanical microtechnique ; Iowa State Univ. Press, USA

>roc. Indian Acad. Sci. (Plant 3ci.), Vol. 01, lumber 2, April 1932, pp. il5-12$.
0) Printed in India.

[dentity of Ficus macrocarpa Wt. ex King (== F. amplocarpa nom. nov.)
md JF. guttata (Wt.) King— A reinvestigation with anatomical

evidence

E GOViNDARAJALU and P MASILAMONEY

Department of Botany, Presidency College, Madras 600005, India

MS received 4 April 1981

Abstract. The two interesting and endemic but hitherto taxonomically indistin-
guishable and confused south Indian species of Ficus (F. guttata and F. macrocarpa)
were reinvestigated both exomorphologically and anatomically. With the help of
data thus obtained their original specific status instead of recently reduced ranks
has been restored, better and dependable distinguishing characters have been blocked
out and the existing confusions, inaccuracies and inconsistencies in literature have
all been rectified. Revised descriptions, illustrations and a workable key are
presented. Vegetative anatomy and the descriptions and illustrations of male
flowers are given for the first time. F. amplocarpa is proposed as a new name
for F. macrocarpa in view of its being a later homonym. It is established that
these two taxa belong to section Neomorphe King and not to Rhizocladus EndL
Anatomically F. amplocarpa is considered to be less specialized than F. guttata.

Keywords. Ficus guttata \ Ficus amplocarpa; morphology; anatomy ; revised
descriptions ; taxonomy.

L. Introduction

the south Indian species of Ficus, F. guttata and F. macrocarpa are two
iteresting taxa which are rather uncommon and restricted in distribution even
a the localities of their occurrence. They resemble each other so closely that
heir respective identity and taxonomic status have hitherto been confused and
aisunderstood. This unfortunate situation in the first place seems to have
temmed from the circumstances that the establishment of these two taxa was
originally based on just one or two complete, incomplete or badly preserved
pecimsns eventually resulting in recording wrong observations and conclusions
y earlier authors. Secondly the subsequent treatments of these taxa in all
ndian Floras [King, Ann. R. Hot. Qard. Calc. 1 (1888) 166, 167 pi. 208, 209 ;
•t in Hoot/. Fl. Br, Ind. 5 (1888) 534 ; Brandis, Ind. Tr. (1921) 621 ; Fischer in
Jamble'sFl. Pres. Madras 3 (1928)951 ; Fyson, Fl. s. Ind. Hill Stn. 1 (1932)
41 et 2 (1932) /. 473] perpetuate what is contained in the protologue without
dding any new points for further improvement Thirdly the illustrations as
provided by Wight [Ic. 6 (1853) t. 1965, 1966] and King (I.e.) reflect not only few
aaccuracies in respect of the delineation of certain characters but both their illus*

US
P.(B)-3

116 E Govindarajalu and P Masilamone'y

trations are not comparable. Lastly the differentiating key characters between
these two taxa have not been sufficiently emphasized but instead more number
of overlapping and common characters camouflaging their respective identity is
repeatedly given.

King (/.c.) has placed these two taxa under his section Neomorphe. Corner
[Gard. Bull Sing. 18 (I960) 7, 32] on the other hand has transferred them to
altogether a different section Rhizocladus Endl. to which F. laevis belongs as they
happen to be root climbers and possessing leaves similar to those of F. laevis
but differing from the latter by the cauliflorous condition only. Furthermore
he has reduced F. macrocarpa as a variety of F. laevis [F. laevis var. macrocarpa
(Miq.) Corner] and F. guttata as synonym of the variety.

The aim of the present investigation in the first place is to wipe out all the existing
wrong and inconsistent details of these two species and to restore their original
specific status and section utilizing new and overlooked exomorphic characters
combined with anatomical data hitherto unavailable, and secondly to provide
revised descriptions and illustrations based on more and better samples and liquid
preserved materials.

The information on general and wood anatomy is meagre in general for the
family as a whole and still less in particular for this large tropical genus Ficus
(Solereder 18.98 ; Metcalfe and Chalfc 19-50). Furthermore no anatomical infor*
mation is available on F. amplocarpa and F. guttata. In the present work the
vegetative anatomy of both these species is undertaken from this standpoint and
also to apply the anatomical data thus obtained as supplementary evidence towards
the elucidation of the taxonomtcal problems mentioned above.

2. Materials and methods

For exomorphological studies both liquid preserved and herbarium materials
cited under each species were used. Likewise both dried and materials pickled
in FAA were used for the anatomical investigations. The dried materials were
revived by boiling in 2 : 1 mixture of glycerol and ' Det ' which is a commercial
reagent (Govindarajalu 1966). After washing in water the revived materials were
stored in FAA. Epidermal peelings were prepared by treating the laminal bits
taken from the midregions of the lamina with 5% Jeffrey's maceration reagent
for about 6 hr at room temperature. When the appropriate stage is reached
the abaxial and adaxial epidermal layers were separated carefully and the unwanted
mesophyll cells gently removed with the help of a fine brush. Finally they were
stained with safranin and mounted in glycerine. Serial microtome sections were
prepared following the conventional methods of dehydration, clearing and
embedding (Johanson 1940). The old in tern odes were repeatedly boiled in water
and later softened with hydrofluoric acid of commercial strength for 4 hrs. After
thorough washing in running water the sections at a thickness of 14 /m were
taken with the sledge microtome and they were stained with safranin and permanent
slides prepared following the customary methods (Johanson 1940). The early
secondary xylem was macerated using Jeffrey's maceration reagent (10%) for
about 3 hrs at 60° C for talcing the measurements of vessels, librifonn and septate
fibres.

identity of Ficus macrbc'ar/a 117

& Observations
3.1. Descriptions
Ficus amplocarpa, Horn. nov.

F. macrocarpa Wt. ex King, Aan. R. Rot. Qard. Calc. 1 (1888) 166, pi. 208 et in
Hook. /. Fl. Br. IndL 5 (1888) 534 • Brandis, Ind. Tr. (1921) 610 ; Fischer in Gamble's
Fl. Madras 3 (1928) 951,955 ; Fyson, Fl. s. Ind. HillStn. 1 (1932) 541 et 2 (1932)
t. 473 ; nan Bl. Cat. Gewass Buitenz. 36 (1823) et Beijdr. (1825) 459, nee L6vill<§
and Vaniot in Mem. Acad. Barcelona 6 (1907) 152 — Pogonotrophe macrocarpa
Miq. in Book Lond. J. Bot. 7 (1848) 74 ; Wight, Ic. 6 (1853) t. 1965 ; King
in Hook /. Fl. Br. Ind. 5 (1888) 534 ; Brandis, Ind. Tr. (1921) 610 ; Fischer in
Gamble's Fl. Pres. Madras 3 (1928) 955. — Ficus vagan$v&x. macrocarpa Miq. Ann.
Mus. Bot. Lugd. Bat. 3 (1867) 293.— F. laevis var. macrocarpa (Miq.) Corner,
Gard. Bull. Sing. 18 (1960) 7 [excl. Coveltia guttata Wt. and F. guttata (Wt.)
King]— figure 1 N, 2 A-G.

Tall scandent tree. Twigs of first few internodes densely hairy, ftstular, rooting
at nodes. Leaves broadly elliptic ovate, rounded at base with entire margin^
membranous (subcoriaceous), usually trinerved (quintinerved), densely or sparsely
tomentose beneath, glabrous above, abruptly acuminate, 10-45 x (5-) 7-9 cm ;
tomentum over the veins brown or rusty brown and the remainder colourless ;
lateral veins 3 (-4) pairs, slightly raised beneath, curvipinnate, arcuate; petiole
subterete, tomentose, ultimately becoming glabrous, 3 -0-4 -5 cm long ; stipules
ovate lanceolate, acute (glabrous) 1 • 5-2 • 0 cm long. Receptacle ramiflorous, in fas-
cicles on naked pendent and/or horizontal cable like branches, globose, purp-
lish red with white streaks and patches, densely pubescent, later becoming glabrous
containing all 3 kinds of flowers without hispid hairs in the interior, 3-0-<6-5cm
across ; basal bracts absent ; peduncles up to I cm long with few small bracts at
base. Male flowers dark brown, orange yellow (when fresh), 4-gonous, pedicellate,
closely arranged behind ostiolar scales in 4 rows, 1-8-2 -Omm long ; pedicels
flattened, membraneously winged, conspicuously 1 nerved, widening towards and
hairy at base, up to 2 mm long; tepals 4-5, tannin punctate, dissimilar in size
and shape, concave ; outer pair narrowly elliptic oblong, 2-4-2-5 x 1-4 mm ;
inner pair elliptic ovate, 2-0-2-4 x 1-0-1-4 mm ; stamens 2, free, almost sessile,
surrounded by a dense tuft of stiff erect, brownish hairs, up to 1-5 mm long ;
anthers fleshy, more or less falcate or erect, mucronate, distinctly 4 lobed,
trigonous, more or less basifixed or basally adnate, up to 1-3 mm long ; filament
0-2 mm long. Female flowers few, intermingled with male, 2-2 mm long (excl.
pedicel, incl. style), pedicellate, rare ; pedicel nearly 1-5 mm long, hairy at base
and top, membraneously winged, flattened, strongly 1 nerved ; tepals 4, elliptic
ovate, inflated, purplish red, obtuse, tannin punctate, equalling or shorter than
ovary, concave, 1-5-1-6 x 1 mm ; ovary subglobose, up to 1-5 mm long; style
excentric, exserted, glabrous, slightly widening towards top, 0-5- 0-6 mm long;
stigma slightly dilated. Gall flowers abundant, 2- 5 mm long (excl. pedicel, incl.
style), pedicellate; pedicel hairy at base, 3-0-6-5 mm long; tepals 6 as long as or
longer than ovary, black, narrowly spathulate, attenuating towards base, acute-
sutjacute, 2-8r-3-0 x 0«5 mm J ovary subglobose containing pupa, 2 mm long f

118 E Govindarajalu and P Masitamoney

style short, lateral, included, erect, adherent with ovary wall, black, 0-5 mm
long, widening into discoid stigma. Acherte sub spherical.

Specimens examined : Govindarajalu 8161, on the way to Thandikudi, Kodai-
fcanal, Madurai Dt. ; Govindarajalu 15894, on the river banks* of Amngallar,
Pannaikadu, Kodaikanal, Madurai Dt., alt. 1300-1400 m (PCM) ; Joseph 12312,
Waverly estate, Anamalai, Coimbatore l>t. ; Vajravelu 37026, Kanakarai R.F.,
Nilgiris Dt. (MH).

Distribution : Coonoor ; Lamb's Rock Road, Pulneys ; Sholas on Church
Cliff, Kodaikanal. Seems to be an endemic to south Indian Hill stations.
F. guttata (Wt.) King

F. guttata (Wt.) King, Ann. R. Bot. Card, Calc. 1 (1888) pi. 209 ; King in Hook.
/. Fl. Br. Ind. 5 (1888) 534 (sub F. guttata Kurz, sphalm.) ; Fischer in Gamble's
Fl. Pres. Madras 3 (1928) 951 and 955 ; Brandis, Ind. Tr. (1921) 610 ; Fyson,
Fl. s. Ind. Hill Stn. 1 (1932) 542; Covellia guttata Wt, Ic. 6 (1853) 8, /. 1966,
figures 1 A-M.

Tall scandent tree. Twigs densely hairy, sometimes glabrous, solid, rooting at nodes.
Leaves coriaceous, (subcoriaceous), petiolate, broadly cordiformis, cordate or
subcordate at base with entire margin, usually trinerved (quintinerved), densely
tomentose throughout beneath and glabrous above, abruptly acuminate, (8-1)
10-15 x 6-tl2cxn; lateral veins 3 (-4) pairs, raised beneath, curvipinnate, arcuate ;
petiole subterete, tomentose ultimately becoming more or less glabrous, 2-4 (-^8)
cm long; stipule ovate-lanceolate, acute, rusty brown tomentose, 1 -0-1 -5cm long.
Receptacle short peduncled occurring in fascicles from the tubercles on the bran-
ches of main stem, globose-sub globose, densely rusty tomentose later becoming
glabrous, 2-5-3*5cm across ; interior of receptacles with hispid hairs ; basal
bracts 3, broadly ovate. Male flowers dark brown, trigonous, tannin punctate,
sessile, closely arranged behind ostiolar scales more or less in 2 rows, 2- 0 x 1 • 5~« 1 • 6
inm ; tepals brownish yellow, 2+2, similar, elliptic ovate, 2 -Ox 1-8-2 -Omni.
Stamen 2, basally united; up to 2 mm long; anthers fleshy, obcuneate, trigonous,
erect, mucronate, basally surrounded by a dense tuft of dark reddish brown stiff
erect hairs, 1-5 x 0-8^0-9 mm ; filament 0-5 mm long. Female flowers few, inter-
mixed with male, 2 • 7->3 - 0 (incl. pedicel) x I • 5 mm; pedicellate ; pedicel 0 • 8-1 -0
inm long, hairy at base, flattened ; tepals 5-6 (-7), narrowly elliptic ovate or oblong,
purplish red, subequal, obtuse, shining, longer than ovary, concave, tannin punc-
tate, 1-5-1-6 x 0-6-0 -7 mm ; ovary obovoid,up to 1 mm long ; style excentric,
erect, distantly hairy behind stigma, exserted, l'0->l-2 mm long ; stigma peltate,
or discoid with crenulate margta (sometimes oblique). Gall flowers abundant, pedi,
ceilate, 2*5mmlong(excl. pedicel, incl. style) ; pedicel hairy at base, <S*5~i7-Omm
long, rest as in female flowers ; tepals 6, dissimilar, narrowly obovate or spathu-
late attenuating towards base, acute-subacute , 2 • 5^3 -0x0- 6-0 • 8 mm ; ovary sessile
globose-subglobose containing pupa, 2' 0-^2-2 mmlong; style short, glabrous, in-
cluded, lateral, curved, adherent with ovary ending in dilated stigma, 0- 5~0 • 6 mm
long.

Specimens examined : Fyson 6448, sine loco ; Fyson, $.n., Shembaganur, Kodai-
kanal, Madurai Dt. ; Fischer, s.n.9 Coonoor, Nilgiris ; Govindarajalu 15720,
Tiger shola, Kodaikanal, Madurai Dt. et 15930, Thandikudi, Kodaikanal, Madurai
Dt. (all PCM) J Joseph 12312, Waverly estate, Anamalai, Coimbatore Dt

Identity of Fiats macrocarpa 119

(1333m) ; Sebastine 18343, Lockhart gap, Kottayam Dt. (1700m) et 24988,
Perumalmalai, Kodaikanal, Madurai Dt. (1700m) ;Vajravelu 26131, Silent Valley
R.F., Palghat Dt (950 m) et 35010, Curzon estate, Nilgiris (1925 m) et 37026,
KonakaraiR.K, Nilgiris ; Shetty 37613, Avalanche, Nilgiris (2000m) ; Vajravelu
39645, Kodanad, Nilgiris (1850m)— all MR.

Distribution : Districts of Coimbatore, Madurai, Nilgiris and Palghat seems
to be restricted to south Indian Hill stations,

3.2. Anatomy

Ficus amplocarpa

Lamina— Abaxial surface : Cells hexagonal, variable in size and shape ;. cell walls
thin, straight (undulate). Stomata (L. 18-22- 5 /im ; W. 9-13 -5 /mi), thin-walled,
anomocytic, elliptic (figure 2M). Trichomes (L. 348-512 /on), unicellular (2-3
celled), erect or reclinate, acute (figure 21), the basal cells of which containing
colourless nodular deposits abundant throughout (figures 3 B i 4 G) ; unicel-
lular microhairs (figure 4 B)and bicellular microhairs (L. 18-0-22-5 jton) with basal
rosette of cells common in the interveinal areas as in F. guttata (figure 3 E) ; shortly
stalked multicellular capitate or peltate glandular trichomes common over the
veins (figure 2 J); erect, unbranched, pointed, multicellular hairs (L. 232-406 //m)
abundant (Qgure 3 C). Idioblasts containing deposits of calcium carbonate of
variable shapes and sizes usually occurring in groups of few excessively thick-
walled cells commonly present as in F. guttata (figure 2 K, L).

Adaxial surface : Cell walls excessively thick-walled, straight. Stomata, cal-
cium carbonate containing idioblasts absent. Other details, see abaxial surface.
T.S. Lamina : Width of lamina examined 0*4-0-45 mm. Cuticle thin over
adaxial and thick with undulations over abaxial surface. Keel "U'-shapcd, adaxi-
ally grooved in the midrib ; margin subacute sloping, down wards containing 3-4
layers of collenchyma and one small submarginal vb. Adaxial epidermal cells
Isodiametric with a tendency to become subdivided in certain places. Abaxial
epidermal cells tangentially elongated, dissimilar in size, some of them containing
granular materials in the form of nodules. Palisade 2-3 layered ; spongy meso-
phyll conspicuously lacunose with large intercellular spaces and abundant tannin ;
rnesophyll cells tangentially elongated, lobed, reticulately arranged. Cyatoliths
common beneath abaxial epidermis (figure 4 B). Abaxial hypodermis 4-5
layered, collenchyma tous. Midrib : vascular strands deeply crescentiform adaxially
closed by strap-shaped single strand (figure 3 G) ; central ground tissue paren-
chymatous containing few phloem bundles ; vascular strands subtended by 4-5
discontinuous layers of gelatinous fibres (figures 2H ; 3 G) j nests of sclereids
with rounded outline and gelatinous wall layers belonging to convolute type
(Type 'A', Hoster and Liese, 1966) less common. Girders adaxia], incomplete,
collenchymatous, 8-10 celled in height and 4-5 celled in width present for all
vb's. Laminal vb's somewhat circular in outline surrounded by a single layer of
parenchymatous bundle sheath. Tannin abundant throughout.

T.S. Petiole (Distal) ; Outline circular with adaxial median groove (figure
4 A). Diameter of petiole examined 2-8-3-0 mm. Epidermal cells isodiametric
containing tannin. Hypodermis single layer of isodiametric tannin free cells.
Qijter cortex consisting of 10-12 layers of lamellar collenchyma ; inner cortex

120 E Govindarajalu and P Masitamoney

Figure 1. 809 for caption page 138.

Identity of Ficus macracarpa

121

Figure 2. See for caption page 128f

122

Govindarajalu and P Masilamoney

Figure 3. See for caption page 128,

Identity of Ficus macrocarpa

CQ

123

H.

PH.

Figure 4, See for caption page 129,

124 E Govtndarajalu and P Masilamoney

broad consisting of thin-^walled polygonal parenchyma cells arranged with inter*
cellular spaces. Vb's 16-18 in number of different sizes, discrete arranged in a
ring with a gap confronting adaxial groove (figure 4 A). Central ground tissue
parenchymatous containing 7-8 phloem bundles in abaxial half (figure 4 A). Tri-»
chomes (L. 348-928 /no), erect, multicellular, brown (due to tannin), unbranched,
pointed, thick-walled present ;; club shaped unicellular trichomes (L. 58-11 6 /on),
excessively thick-walled containing tannin also present ; peltate or spherical
multicellular, shortly stalked glandular trichomes present and all of them inter-
mixed. Cubical crystals and irregularly shaped crystalline bodies less common
in ground and cortical parenchyma cells. Basal and mid-regions of petiole
differing by containing 25 vb's subtended by discrete units of gelatinous fibres
(Type A). Basal and midregtons of petiole, see distal region of petiole.

T.S. Node : Pentalacunar, each one of three adaxial gaps confronted by a
single trace except two abaxial gaps containing split traces in each (Howard 1974).

TJS. Internode (early secondary xylem) : Diameter of internode examined
7-8 mm. Phellem superficial. Phelloderm broad. Sclereids tangentially elon-
gated or elliptic with lumen and ramiform pits (figure 4 H), 2-3 layered forming
a continuous ring present beneath phellem ; brachysclereids with reduced lumina
and gelatinous wall layers occurring in several tangential units through out secon-
dary cortex and secondary phloem (figure 4 E). Growth rings present. Librifora
fibres thin-walled (L. 1160-1508 //m) J septate fibres not common (L. 580-812
/on). Parenchyma paratracheal banded, 2-3 layered (figure 4 E). Pores solitary,
circular-oval or in short radial multiples of 2-3. Vessels (L. 348- 580 /an), oblique
porous (transverse) with alternate intervascular pittings, non-storied. Tannin
rare in secondary cortex. Starch grains abundant in cortex, xylem parenchyma
and rays.

F. guttata

Lamina— -Abaxial surface ; Cells polygonal, thin^walled J cell walls straight.
Stomata (L. 21-2-26- 5 /on ; W. 15 -9-21- 2 /*m), thick-walled, anomocytic,broadly
elliptic (figure 3 F). Trichomes (L. 337- 5-540 0m), unicellular, erect or reclinate,
thick-walled, pointed, mounted on dilated calcified base abundant over veins as
in F. amplocarpa (figures 3 B, 4 C) ; trichomes surrounded by 1 concentric row of
cells, the latter usually containing smooth (figure 4 F) or unevenly striated calca-
reous, deposits and groups of such cells variable in number and arrangement abun-
dantly present independent of trichomes also (figures 2 K, L). Calcareous de-
posits usually of elongated form (smaller rounded ones) occurring in discontinuous
rows over larger veins also present (figure 3 A) j shortly stalked multicellular
capitate or peltate glandular trichomes common as in F. amplocarpa (figure 2 J) ;
multicellular trichomes (L. 348-928 /^m), see F. amplocarpa j prickles (L. 22-5-
27-0 /an), 2 celled, excessively thick-walled, pointed, present usually in crypts
opposite to cystoliths (figure 3E),

Adaxial surface : Cells variable in size with moderately thick walls. Trichomes
less common. Other details, see abaxial surface.

T.S. Lamina ; Width of lamina examined 0-4-^0 -47 mm. Adaxial epidermis
usually tending to become 2-3 layered due to subdivisions. Midrib vasculature
as in F. amplocarpa but adaxially confronted by (2-) 3 (-4) discrete strands and
enclosing within variously oriented 3-5 additional strands (figure 3 H), Sclereids

Identity of Ficus rnacrocarpa 125

with gelatinous wall layers as in F. amplocarpa (figure 2 H) characteristically and
abundantly present in regular radial rows subtending the strands (figure 3 H).
Central ground tissue heterogeneous containing parenchyma, sclerenchyma and
few groups of gelatinous fibres. Phloem bundles absent. Cubical crystals pre-
sent in phloem parenchyma and in central ground tissue. Other details as in
F. amplocarpa.

T.S. Petiole (distal) : Diameter of petiole examined c. 4mm. Vb's 18-20
in number of different sizes, discrete, arranged in a ring without adaxial gap, en*
closing within phloem bundles (figure 4C). Trichomes (L. 290-464 ^m), 1-2
celled, erect, thicJowalled, uribranched, pointed, containing tannin abundant J
trichomes of other types found in F. amplocarpa absent. Basal and mid regions
differing by way of reduction in the number of phloem bundles and for other
details see distal. Other details as in F. amplocarpa.

T.S. Node : See F. amplocarpa.

T.S. Internode (early secondary xylem) : Pores usually in radial multiples of
(2-) 3-4. Vessels (L. 174-324/zm), storied. Tyloses common. Secondary
phloem rather broad and radially traversed by dilated rays (figure 4 D). Other
details, see F. amplocarpa.

4. Discussion

The present study clearly indicates that Ficus amplocarpa and F. gut tat a are two
distinct species and in this respect the former is characterized by usually membra*
nous broadly elliptic ovate leaves with rounded base, fistular internodes, rami-
florous larger receptacles on naked pendent and/or horizontal cable like branches,
ebracteate receptacles tetragonous pedicellate male flowers, flattened membra^
nously winged 1 nerved pedicel widening and hairy towards base, unequal tepals
in male flowers, almost sessile free stamens with somewhat falcate anthers, smaller
female flowers with longer pedicels and 4 tepalled perianth, sub globose ovary,
glabrous style slightly widening towards top with slightly dilated stigma, gall
flowers with elliptic linear tepals and discoid stigma. With reference to the
above mentioned characters JP. guttata differs from F. amplocarpa by coriaceous
(subcoriaceous) leaves, broadly cordiform solid internodes, usually smaller
receptacles developing from tubercles of stem and branches, bracteate receptacles,
sessile trigonous male flowers with similar tepals, stamens more or less united at
base with longer filaments., obcuneate anthers, larger female flowers with shorter
pedicels and perianth of 5-6 (-7) tepals, obovoid ovary, style distinctly hairy
behind stigma, peltate or infundibuliform stigma with crenulate margin, gall
flowers with narrowly ob ovate or spathulate tepals and dilated stigma.
Wight (1853) has given the illustrations of both F. amplocarpa ( = F. macro*
carpd) and F. guttata under Pogonotrophe rnacrocarpa Miq. (Ic. 1. 1965) and Covellia
gutiataVJt.Qc. t. 1966) respectively, the former without the analysis of floral
parts and the latter accompanied by them. Furthermore the green colour of the
receptacles, glabrous condition of young twigs, petioles and stipules, emarginate,
subacute or attenuating base mentioned by Wight (1853) for F. macrocarpa&tQ
all quite contrary to the present observations *

126 E Govindarajalu and P Masilamoney

King (Ann. Roy. Bot. Gard. Calc. 2 (1888) pi. 209) has adopted the figures of
female flowers of F.guttata as given by Wight (I.e.) with some modifications. But
the figures of vegetative organs and the receptacles as given by Wight on the one
hand and those of King on the other show no agreement at all. Wight has shown
the infundibuliform stigma to be hairy at the margin and also said that the
receptacles do not contain any male flowers which are not correct

The present observations which are based on more number of herbarium speci-
mens and liquid preserved materials differ from those of King (1888) in respect of
F. amplocarpct ( = F. macrocarpd). King reports the absence of male and gall
flowers in the receptacles but in fact the former are present in fewer numbers just
behind the ostiolar scales and the latter are abundant in the rest of the receptacles.
The perianth of female flowers are said to be made up of 6 tepals instead of only
4 tannin punctate tepals. He says fertile female flowers are sessile or pedicellate
and the style hairy. On the contrary they are always pedicellate and style glabrous.
The stigma is distinctly entire and dilated and not bilobed as mentioned by King
(1888). The ovary is not stipitate as shown in pi. 208, f. 5, 7 and 9. Ficus macro-
carpa as illustrated by King shows more number of primary lateral nerves, glabrous
twigs, leaves and stipules, receptacles without white patches, hairy style, bilobed
stigma and elliptic ovate tepals which prove to be otherwise according to our obser-
vations. In the legend for figure 1 it is stated that it is part of fascicle of recep-
tacle from the stem below the leaves but actually they are arranged in an elongated
cable like pendent and/or horizontal branches. It is shown that fertile female
flowers are pedicellate (figures 5 and 7) and sessile (figure 6) but the present
observation reveals only pedicellate condition.

King (Ann. Roy. Bot. Gard. Calc. 2 (1888) 167) has said that in F. guttatahz
could not observe the male flowers in the only receptacle that was available to
him and also as in the case of F. macrocarpa he could not conie across any gall
flowers since all the flowers in the single receptacle appeared to be fertile female
flowers. In fact, the receptacles are found to contain all the three kinds of
flowers. According to King (I.e.) the fertile female flowers are said to be sessile
which is contrary not only with reference to his own figure (pi. 209, f.6) but our
observations. Eachtepalis shown to be strongly medianly 1 nerved instead of the
nerveless condition. The ovary is obovate and the style is distinctly hairy behind
stigma unlike those of the figures in which the former is shown as subglobose and
the latter perfectly glabrous. Another interesting situation in this respect is that
Wight's figure 4 (Ic. t 196161) representing the female flowers of F.guttata
( = Covellia guttatd) is given as such with slight enlargement by King (i.e. pi.
209, figure 6) but the stigmatic margin shown to be hairy (Wight's figure 4) has
been shown as perfectly glabrous (King's figure 6). Furthermore King's figures
6 and 7 (pi. 209) are labelled as female flowers under different stages of develop-
ment but they all look so different that one has got nothing to do with the other.

Corner (Gard. Bull. Sing. 18 (I960) 7) has reduced F. macrocarpa as a variety of
F. laevis Blunder sect Rhizocladus Endl. and F. guttata as a synonym of this
variety. The reasons given by him for this procedure are the existence of root
climbing habit and the shape of the leaves which are similar to those of F. laevis.
Nevertheless F. laevis differs from F. amplocarpa (= F. macrocarpd) by its climbing
habit? dentate margin and deeply cordate base of the teaf? axillary solitary

Identity of Ficus macro carp a 127

greenish yellow receptacles on long peduncles subtended by 3 basal bracts with
densely hispid hairy interior, 5 linear lanceolate tepals in male and female flowers,
glabrous stamtnal filament, dorsifihced n on fleshy anthers with sub sagittate base and
acute apex, terminal style equalling the length of achene, bifid stigma and elongated
ovoid achene. Furthermore when the characters of both F. amplocarpa and
F. guttata agree well with those of the section Neomorphe King in that the flowers
are unisexual, male and gall flowers in one set of receptacle, fertile female flowers
in another set of receptacle, male flowers with 2 stamens, inflated perianth with

3 or 4 membranous tepals, fertile female flowers smaller than male or gall flowers*
receptacles often very large in fascicles from tubercles on the stem and longer
branches, trees rarely scandent shrubs, never epiphytal, they are allowed to retain
their berth in the sect. Neomorphe itself. Furthermore as the specific differences
between K amplocarpa and F. guttata are many and clear cut (see above) and each
one of them in turn widely differs from F. laevisin respect of several characters
the original specific status of the former two taxa is restored.

As mentioned by King (Ann. Roy. Bot. Gard. Calc. 2 (1888) 129), Miquel
(Ann. Mus. Lugd. Bat. 3 (1867) 278) has considered Pogonotraphe macrocarpa
(Wt. Ic. t. 1965) as referable to F. vagans Roxb. but Roxburgh's manuscript
drawing of the latter in Hb. CaL shows that it is clearly identical with authentic
specimens of F. laevis BL

Fyson (PL s. Ind. Hill. stn. i. (19-32) 541) distinguishes F. amplocarpa from
F. guttata only by the glabrous young parts of the former and the hairy young
parts of the latter and not on the basis of any other characters. Although the
peduncle bearing the receptacle and leafy branch as illustrated by Fyson I.e. 2
(1932) appears to be satisfactory the shape of the leaves has greater resemblance
to F. guttata than to F. amplocarpa. Likewise the female flowers illustrated by
him differ from the present observations.

The anatomical differences between F. amplocarpa and F. guttata are so signifi-
cant and convincing that these two taxa can be maintained undoubtedly as two
distinct species on this ground also. F. amplocarpa differs from K guttata by the
absence of 2-celled thick-walled prickles in the leaf, midrib with crescentiform
vascular strand adaxially closed by single strap-shaped strand, occurrence of gela-»
tin/oils fibres in 4-5 discontinuous layers subtending the midrib vascular strands,
central ground tissue of midrib containing phloem bundles, less common occur-
rence of sclereids with gelatinous wall layers subtending the midrib strands,
presence of club shaped and peltate or spherical glandular trichomes in the petioles,
non-storied vessels, longer vessels with oblique porous perforations, absence of
tyloses, nondevelopment of radially traversing dilated rays in the secondary
phloem, longer libriform and septate fibres. Incidentally it is also interesting to
observe that F. amplocarpa is found to be less specialized than F. guttata since the
vessels of the former are longer with oblique porous perforations and non-storied,
and septate and libriform fibres are longer than those of the latter.

Leaves broadly elliptic, membranous, rounded at base; receptacle large 3-0-
6' 5 cm across with white blotches, ebracteate and without hispid hairs in the
interior ; stamens free with somewhat falcate anthers ; tepals of female flowers

4 ; equalling or shorter than ovary ; ovary subglobose ; style glabrous, stigma
dilated F. amplocarpa (=F. macrocarpa)

128 E Govindarajalu and P Masilamoney

Leaves broadly cordiform, coriaceous, cordate or subcordate at base ;
receptacles small, 2 -5-3 -5 cm across without white blotches, bracteate with hispid
hairs in the interior ; stamens basally united with obcuneate anthers ; tepals of
female flowers 5-6-(-7) ; longer than ovary ; ovary obovoid ; style hairy behind
stigma ; stigma usually peltate or discoid F. guttata.

Acknowledgement

We are highly thankful to Dr N C Nair, Botanical Survey of India, Southern
Circle, Coimbatore, for loaning the specimens cited here as M H.

References

Goviadarajalu E 1966 The systematic anatomy of South Indian Cyperaceae : BulbostylisKunth;

jr. Linn. Soc. (Bat.) 59 289-304
Howard R A 1974 The stem node-leaf continuum of the Dicotyledoneae ; /. Arnold. Arb. 55

125-173
Hoster H R and Liese W 1966 Ober das Vorkommen von Reaktionsgewebc in Wurzeln und

Asten der Dikotyledonen ; Holforsch. 20 $0-90
Johanson D A 1940 Plant microtechnique (New York : McGraw-Hill)
Metcalfe C R and Chalk L 1950 Anatomy of the Dicotyledons 2 vols. (Oxford)
Solereder H 1908 Systematic anatomy of the dicotyledons 2 vols. (Oxford)

Figures 1 A-N. Ficus guttata. A. abaxial surface of leaf x 1/5 ; B. male flower
bud x 8 ; C. tepal of male flower x 14 ; D. stamen x 16; E. and F. different forms
of stigma (diagrammatic) ; G. female flower with tepals removed showing one form
of stigma x 12 ;H. gall flower x 7 ; L ovary of gall flower x 8 ; J, tepal of female
flower x 16 ; K. female flower (entire) with another form of stigma x 11; L. and M,
tepals of gall flower (two types) x 13 ; N. F. amplocarpa abaxial surface of
leaf x i.

Figures 2 A~M. A-J. F. amplocarpa. A. male flower bud X $ J B. outer tepal
of male flower x 11 ; C. inner tepal of male flower x 13 ; D. staminal pair of a
single male flower x 16 ; E. gall flower x 5 ; F. female flower X 10 ; G. another
type of stamen X 14 ; H. cortical gelatinous fibres (type A) x 170 ; I. unicellular
hair with a basal rosette of cells x 107 ; J. peltate glandular hair X 170 ; K. and L.
F. guttata leaf epidermal cells containing different forms of calcareous deposits
X 170 ; M. F. amplocarpa surface view of stoma x 450.

Figures 3 A~H. A. Ficus guttata portion of lateral vein showing different forms of
calcareous deposits x 170 ; B. and C. F. amplocarpa : B. part of leaf epidermal hairs
showing calcareous deposit containing basal cell x 170 ; C. rnulticellular hair aver
veins x 100 ; D-F. F. guttata : D.cystolith containing idioblast x 170 ; E. 2-celled
microhair with cystolith containing basal idioblast x450; F. surface view of stoma
x 450 ; G. F. amplocarpa transection of midrib x 22 ; H. F. guttata transectioa of
midrib X 22.

Identity of Ficus macro carpa 129

Figures 4 A-H. A. and B. Ficus amplocarpa : A. trans ectioni of petiole at distal end
(semi diagrammatic) ; B. cystolith opposed by 1 -celled microhair x 170 ; C. and D.
jp. guttata: C. transection of petiole at distal end (semidiagrammatic); D. transection
of oldinternode (semi diagrammatic) ; E. F. amplocarpa transection of old internode
(semiciiagrammatic) ; F. F. guttata leaf epidermal hair base encircled by calcareous
deposit containing cells x 170; G. and H. F. amplocarpa : G. surface view of cystolith
containing hair base x 170 ; H. brachysclereid from cortex x 65.

Abbreviations:. C. cystolith CO. cortex; D. calcareous deposit; E. epidermis ;
G. gelatinous fibres G.C guard cell ; H. hair; L. lenticel; P. xylem parenchyma;
PD. phelloderrn; PH. phloem ; PM. phellem ; R. rays ; S. sclereids ; T- tannin
idioblast ; VB. vascular bundle ; XY. xyiem ; c. circa ; D. diameter ; Ht. height ;
L. length ; vb. vascular bundle (pi. vb's) ; W. width.

Proc. Indian Acad. Slci. (Plant Sci.), Vol. 91, Number 2, April 1982, pp. 131-137.
© Printed in India.

The genus Jackiella in South India

RAM UDAR and ADARSH KUMAR

Department of Botany, University of Lucknow, Lucknow 226007, India

MS reoeived 8 April 1981

Abstract. Jackiella ceylanica Schiffn. ex St. and /. javanica var, cordifolia SchifFn.
are being reported for the first time from India. The taxonornic details of the
two taxa together with a discussion of the tenabiiity of two varieties of J. javanica
SchifFn. namely var. cordifolia and var. cavifolia have been given.

Keywords. Bryophyta; hepaticae; Genus Jackiella; South India; taxonomic details*

1. Introduction

In a recent contribution (Udar and Kumar 1981) the range of distribution and
diagnostic characteristics of the marsupial genus Jackiella together with the taxo-*
nomic details of J. javanica Schiffn. from eastern India had been given. However,
during a survey of south Indian liverworts, which have not yet received proper
attention, plants resembling J. javanica in all characters, except in habit of plant,
siz3 and shape of leaf, degree of development of trigones in cells of leaf and stem
and in structure of capsule wall, were discovered. Among all these characters,
the development of trigones seems to be ecologically influenced but the habit of
the plant, the shape of the leaves and the structure of capsule wall show more or
less stabilised features which merit recognition of varietal ranks for the east Indian
and south Indian plants. Schiffner (1900) had already recognised two varieties of
Jackiella javanica Schiffn, primarily based on vegetative characters viz., J. javanica
var. cavifolia Schiffn. from Java, Sumatra and /. javanica var. cordifolia Schiffn.
from Java (see also Bonnet 1966). The former variety is stated to be characterised
by small and creeping plants with leaves broader than long and the latter by more
elongated, uprising to almost erect plants with leaves longer than broad. The
plants of east Indian territory could be referred to as J". javanica var. cavifolia and
those from south Indian territory as J. javanica vat. cordifolia. Our study based
on east Indian (from Sikfcim and Shillong) and south Indian (from Wellington
and Dodabetta) plants reveals that sporophytic differences are also present
between these two varieties hitherto not recognised.

A survey of plants from Kodaikanal (Palni Hills) iu the Indian peninsular region,
nearer to Ceylon, revealed plants which resemble 3. ceylanica Schiffn. ex St.— a
taxon which has so far been considered endemic to Ceylon. The discovery of this
species from India constitutes a transoceanic disjunct distribution for this taxon

131

132 Ram Udar and Adarsh Kumar

In the present state of our knowledge the genus Jackiellais represented in India
by /. ceylanica, J. javanica var. cavifolia and /. javanica var. cordifolia. The
taxonomic details of /, ceylanica and /. javanica var. cordifolia, occurring in south
India, are being described in the present paper. The remainin g taxon from eastern
India has already been reported elsewhere (Udar and Kumar 1981).

2. Materials and methods

The plants of J. javanica V&T. cordifolia were collected from Wellington and Doda-
betta and /. ceylanica from Kodaikanal. The taxonomic details of the former
taxon are based on plants collected from Wellington except those relating to the
capsule wall which are drawn from the plants of Dodabetta. The plants were
stretched in water and placed in incubator for 24 hr at 40° C. The slides were
prepared in 70% glycerine.

3. Key to the Indian species of Jackie lla

1. Plants erect, deep brown. Leaves with curved margin. Male inflorescence
with 7-15 (or more) pairs of male bracts ; male bracts oblong with flat dorsal
lobe /. ceylanica

1. Plants prostrate^suberect, light green. Margin of leaves not curved. Male
inflorescence with 3-10 pairs of male bracts; male bracts globose- subglobose
with saccate dorsal lobe. . • /< javanica 2

2. Plants prostrate. Leaves broader than long, concave, with decurrent antical
margin. Inner layer of capsule wall in surface view with incomplete
thickening ban da /. javanica var. cavifolia

2. Plants suberect. Leaves longer than broad, without decurrent an tical margin.
Inner layer of capsule wall usually with complete thickening bands
/. javanica var. cordifolia

4. Observation

4.1. Jackiella ceylanica Schiffn. ex St. Species Hepaticarum 3 :272, (1908).
Figures 1-48.

Plants dioecious, dark brown, up to 15 mm long, rigid, erect, in dense mat.
Branches ventraHntercalary, large, ascending upward, brownish green to deep
brown. Stem up to 8(-10) cells across, 140-210 fan in diam., cortical cells isodia-
metric or variable deep brown, thick-walled, 18-33 x 18-25 jam ; medullary
cells usually isodiametric, yellowish brown, thin to thick-walled, 12-03 x 18-25
//m. Leaves succubous* entire, ovate to oblong, usually broader than long, or
occasionally longer than broad, 0-65-0*97 (-1*2) x 0-8-1 -Omm, deep brown,
sub-opposite, insertion oblique, apex obtuse, base broad J antical margin slightly
decurrent, curved ; postical margin arched, sometimes curved, marginal cells
longer than broad, broader than long or as long as broad, 15-30 x 15-30 //m J
middle and basal cells longer than broad, occasionally broader than long, (18-

The genus Jackiella in South India

133

Figures 1-18. Jackiella ceylanlca 1. Female plant with marsupium. 2-4. T. S.
of axes (2, 3 axes of vegetative plant ; 4. axis of male inflorescence). 5-7. Leaves:
8. Marginal cells of leaf. 9. Middle cells of leaf. 10. Underleaf . 11. Portion
of male plant with male inflorescence. 12. Male inflorescence. 13-16. Male
bracts. 17. Young marsupium. 18. Female bract.

21-44 (-51) x 18-44 /mi ; cell walls thick, sometimes thin in younger leaves, tri*
gones prominent and bulging. Gemmae not seen. Underleaves highly reduced,
only near apex of stem, (1-) 2-3 cells long, 2-3 cells broad, connate at base with the
leaf of one side, with 1-celled marginal teeth ; cells thin-walled, trigones feebly
developed. Rhizoids scarcely present on axis, usually restricted at base of under-
leaves, with swollen tips harbouring mycorrhiza. Male plant with short, spicate,
ventral branches, androecia terminal or intercalary consisting of 7-J5 or more pairs
of male bracts ; male inflorescence usually with 5-celled thick axis having indis-
tinct cortical and medullary regions ; male bracts bilobed, oblong, 180-260 x
105-155 im : ventral lobe oblong, comparatively larger, saccate : dorsal lobe
oblong, smaller, flat, enclosing single anthericjium ; male bractegles yestrjjctecl

134 Ram Udar and Adarsh Kumar

only at base of male inflorescence. Female plant with 2 or more short, ventral,
female inflorescence having 1-2 pairs of female bracts at mouth of marsupium ;
female bracts saccate, apex obtuse, sometimes subacute to acute j perianth absent ;
marsupium deep brown, cylindrical, 0' 5-1 -Omm long, with numerous rhizoids
on its surface ; archegonia up to 3->5 at mouth of the marsupium with short neck.

4- la. Specimens examined : LWU 79/66, 95/66 ; Coll. R. Udar and S. C. Sri-,
vastava ; Loc. Kodaikanal (Palni Hills, South India), alt ca 1200m ; t>ats
January 6, 1966 ; Det. R, Udar and A. Kumar.

4-lb, Ecology : The plants grow on rocky soil in association with Pogortatum
sp. on moist and exposed surface.

4. Ic . Discussion : J. ceylanica Schiffn . ex St. was originally described by Stephani
(1906-« 1909) from Ceylon, who provided a short Latin diagnosis only along with
some illustrations in his unpublished Icones No. 2024. Later Abeywickrama
(1959) reported it from the same island with a meagre description and wrong
illustration (see fig, I6c, p. 48). The adequate taxonomic details of this plant do
not seem to have been published so far. The characters of this species from Kodai-
fcanal reveal a great deal of diversity and plasticity in vegetative and reproductive
characters which tend to approach J.javanica. However, /. ceylanica differs from
the latter in larger, ascending to erect and deep brown plants, and with curved
margins in leaves. The androecia are very characteristic in being terminal or
intercalary and consist of 7-il,5 or more pairs of oblong male bracts with flat
dorsal lobe. In /. javanica, however, the plants are smaller, prostrate and light
green with more or less flat leaves. The androecia are terminal with 3~» 10 pairs
of male bracts which are globose to subglobose with saccate dorsal lobe.

Considerable variation occurs in the leaves in /. ceylanica which may be ovate
to oblong, broader than long (figures 6, 7) or longer than broad (figure 5) but
characteristically with curved margin. The uaderleaves are highly reduced
(figure 10) and apparently confined near the apex of the young shoot. The rhizoidal
tipsi are swollen and harbour mycorrhiza. They may perform similar function of
nitrogen fixation as reported for the east Indian plants by Udar and Kumar (1981).

4.2. Jackiella javanica var. cordifolia Schiffn.

Denkscher. Mat. Nat. Cl. Kais. Acad. Wiss. Wien 70 :217 ( : 115), (1900).
Figures 19-38, Plants dioecious, light green, occasionally yellowish brown, up to
10mm long, delicate, prostrate to suberect, in dense mat. Branches ventral inter-
calary, small, ascending, light green. Stem prostrate or with ascending apex,
up to 6-7 cells across, 100-160 /*m in diam., cortical cells isodiametric, light to deep
brown, thick-walled, 7-28 x 7-28 //m ; medullary cells more or less isodiametric,
light brown to yellowish brown, thick to thin-walled, trigonous, 10-25 x 10-23 /*m,
Leaves succubous, entire, ovate or cordate or subquadrate to slightly oblong, usually
longer than broad or as long as broad or occasionally broader than long, light
.green to light brown, 0-45-0 -62 x 0' 44-0 -54 mm at middle; at plant apex
usually broader than long, light green, (0-5-) 0-64-0-75 x (0-6-) 0-75-0 -9
(-1-0) mm, sub-opposite, insertion oblique, apex obtuse, occasionally slightly retuse,
entire (to wavy), not ctecurrent ; marginal cells usually as long as broad,

The genus Jackiella in South India

135

Figures 19-38. Jackiella Javanica var. cordifolia. 19. Female plant with marsupium.
20, 21. Vegetative plants. 22. T.S. of axis. 23-25. Leaves. 26. Marginal
cells of leaf showing attached gemmae. 27. Gemmae. 28. Marginal cells of leaf.
29. Middle cells of leaf. 30, 31. Underleaves. 32. Marsupium with young
sporophyte. 33, 34. Female bracts. 35. Outer layer of capsule wall. 36. Inner
layer of capsule wall. 37. Spores. 38. Elater.

longer than broad or broader than long, 18-28 (-38) x 18-28 (-33) //m ; middle
and basal cells isodiametric, more or less longer than broad, 20-44 x 15-30 /on ;
cell walls thin, trigones prominent, bulging ; base broader in younger leaves.
Gemmae occasional, at apical margin of young leaf, hyaline to yellowish brown,
1 -celled, spherical, 10-15 jum in diam., 2-celled, 18-35 x 12-15 //m. Underleaves
reduced, up to 4-5 cells long, 4-5 cells broad, bifid, lobes uniseriate, margin
dentate, connate at base with leaves of one side, cell walls thin, trigones feebly
developed. Rhizoids numerous on prostrate axis, particularly near the base of
Underleaves, with swollen tips harbouring mycorrlvza. Male plants absent,

136 Ram Udar and Adarsh Kumar

Female plant with 1-2 short, ventral, female inflorescence having 1-2 pairs of
female bracts at mouth of marsupium ; female bracts saccate, apex obtuse to
subacute with entire to wavy margin ; perianth absent ; marsupium light to deep
brown, cylindrical to sickel-shaped, narrowed at base, 1-2 mm long, 0-4-0 -55 mm
wide, with some rhizoids at surface, archegonia up to 6 at mouth of marsupium
with short neck. Sporophyte young, enclosed within marsupium. Seta small.
Capsule cylindrical, deep reddish brown, 540-760 /mi long, 2 15-325 /on wide,
with obtuse apex on 1-celled thick capsular disc, dehiscence in two valves each
with a small cleft at apex; wall bis-tratose ; cells of outer layer quadrate to
subquadrate to elongated, 23-44 x 12-24 /mi with nodular to confluent thickenings
on radial walls, occasionally also on transverse walls ; cells of inner layer sub-
quadrate to elongated, 25-50 (-60) x 20-25 jum with complete, sometimes in-
complete thickening bands on inner tangential wall, sometimes semiannular bands
bifurcate on inner tangential wall. Spores spherical, light yellowish-brown,
6-9 /mi in diam., exine with small papillae. Elaters 75-210 jum long, 11-1 6 /on
broad at middle, reddish brown, bispiral.

4-2a. Specimens examined: LWU 375/71 ; Coll. R. Udar and party ; Loc.
Wellington (Nilgiri Hills, South India), alt. ca 1350m ; Date December 31, 1971.
LWU 184/72 ; Coll. R. Udar and party ; Loc. Dodabetta (Nilgiri Hills, South
India), alt. ca 2670m ; Date January 5, 1972 ; Det. R. Udar and A. Kumar.

4-2b. Ecology : The plants grow on laterite soil on rock surface in dense mats
at moist places in association with Jungermannia sp. and Pogonatum sp.

4»2c. Discussion : J. javanica var. cordifolia Schiffn. was instituted by SchifFner
(1900) from Java and is being described from the bryoflora of Wellington and
Dodabetta (South India) for the first time from India. The plants differ from
/. javanica var. cavifolia in habit and in shape of leaves (see Schiffner 1900). How-
ever, the capsule wall structrae — a sporophytic character, earlier not known, also
provides additional differentiating features for the two varieties. The former
variety has prostrate plants with ascending apex (figure 21) or ascending branches
(figure 20) which are prostrate in the latter. The leaves in var. cordifolia, similar
to J. ceylanica, show plasticity in size and shape as they are longer than broad,
flat, hardly decurrent and comparatively smaller at middle part of the axis whereas
in var. cavifolia these are broader than long, concave and antically decurrent. In
addition to these vegetative features, the development of thickening bands on inner
tangential wall of the inner layer of capsule wall is usually complete in var.
cordifolia but normally incomplete in var. cavifolia.

The leaves show a great deal of variation in their size and shape. These are
longer than broad and cordate to ovate or sometimes subquadrate to slightly oblong
at middle and basal part of the axis but broader than long towards apex. The
margin may be entire or wavy with obtuse to retuse apex (figure 24) which is due to
formation of gemmae. The gemmae are 1-2 celled (figure 27) and formed exo-
genously from the marginal cells of the leaves (figure 26). The rhizoids are mostly
restricted on prostrate axis and also on the surface of marsupium (figure 19) with
swollen tips harbouring mycorrhiza. The marsupium is either cylindrical or sickel-

The. genus Jackiella in South India 137

shaped and the female bracts are with truncate to obtuse apex and entire
margin (figure 32) or with acute to subacute apex and wavy to dentate margin
(figures 33, 34).

Acknowledgements

The authors wish to acknowledge their gratitude to the late Dr Herman Persson
for Stephani's unpublished Icones and to the Department of Science and Techno-
logy (SERC), Government of India, for financial support.

References

Abeywickrama B A 1959 The genera of the liverworts of Ceylon ; Ceylon /. Sci. {Bio. Sc.)

2 33-81

Banner C E B 1966 Index Hepatlcarum VI (Germany : Verlag Von. J. Cramer) 4S1-739
Schiffner V 1900 Die Hepaticae der flora von Buitenzorg (Leiden : E J Brill) 1-220
Stephana F 1906-1909 Species Hepaticarum III (Geneve : Georg and C*° Libraires-Editeurs)

1-693
Udar R and Kumar A 1981 Jackiella javanica SchifTn. a rare and interesting taxon from India ;

/. Indian Bot. Soc. 60 105-111

Proc. Indian Acad. Scj. (PJant Sc/.), Vol. 91, Number 2, April 19&2, pp. 139-143
© Printed in India.

Geocalyx Nees-a rare marsupial genus from India

RAM UDAR, S C SRIVASTAVA and BHlRENt)RA KUMAR
Department of Botany, University of Lucknow, Lucknow 226 007, India

VCS received 11 Miy 1981; revised 23 April 1982

Abstract Taxonamic details of Geocalyx graveolens Nees recently discovered from
tlvj Valley of Flty.vers in western Himalayas (altitude ca 4670 meters) have been
provided. Tiie discovery of this genus in the above area not oaly extends its range of
distribution in the Himalayas but also constitutes a new record of this taxon In Indian
bryoflora. The plants are monoecious and characterized "by undulating stem, bifid
leaves which are flat or ascending, and bifid underleaves. The androecial and
g/aoccial branches are ventral and axillary. The marsupium arises initially as tuber-
like structure (often 2-S in number on the ventral surface of a plant) and, at matu-
rity becomes cylindrical with numerous rhizoids studded on its surface.

Key wards. Bryophyta; Hepaticae ; Jungermannialcs ; Geocalycaceae ; Geocdyx\
now record for India,

1. Introduction

Geocalyx Nees is a small genus represented by only seven species, viz., (?. cakdo*
nicus St. from New Caledonia, G. cantor tuplicatus Mont, et Nees from San
Domingo, G. graveolens (Schrad.) Nees from Europe, Siberia, Japan and North
America, G. orientals Besch.ef Spr., G. borbonicus St. (c.f. orientals) from Re-
union, G. novazelandiae Efcrz. from New Zealand and G. yakusimensis Hatt. from
Japan (Stephaui 1906-1909 ; Macvicar 1926 ; Herzog 1935 ; Hattori 1948 ;
Hodgson 1958;Bonner 1965).

This genus is remarkable in having the young sporophyte deeply sunk in a fleshy,
cylindrical sac-like, pendulous, marsupium arising from the postical surface of
the stem. Although several marsupial taxa are known from Indian sub-cor.ti-
nentbutthe details of the marsupium are available only in Jackiella javanica
Schiffn. and /. ceylanica Schiffn.. luxuriantly distributed in eastern Himalayas and
south India (Udarand Kumar 1981, 1982). The present paper provides details of
Geocalyx graveolens, another marsupial taxon , recently collected from the Valley
of Flowers. The discovery of this genus in the above area not only extends its
rangfc of distribution in the Himalayas but also constitutes a new record of this
taxon in Indian flora. Evans (1939) treated this genus under the family Harpan*
thaceae (see also Mtiller 1951-1958 ; Hodgson 1958). Schuster (1972), on the
other hand, erected a new suborder Geocalycinae to accommodate Geocalycaceae
to which this plant belongs, along with other families.

139

P.(B)-5

140 Ram Udar, $ C Srivastava and Dhirendra Kumar

2. Taxonomic description

Geocalyx graveolens (Schrad.) Nees, Eur. Lab. II. p. 397 1836 (figures 1-25)

Plants yellowish -green to green, sparingly branched, creeping with distinct undula-*
tions. Stem l-l-5(-i2)cm long, 9-11 cells across, cortical and medullary cells
alike. Leaves succubous, sub-opposite or alternate, obliquely inserted, flat or
ascending, lamina up to 24 cells broad, bifid, occasionally tri-tetrafid, sinus broad,
extending 1/4 to 1/3 of leaf length, lobes ovate, ICM4 cells long, 9-14 cells broad*
682^770 x 561-605 jum, apex acute to subacute (1-celled or 2-celled long), antical
margin decurrent, postical margin straight, leaf cells polygonal with inconspicuous
trigones, 15-23 x 10-18 /on at the margins, 19-29 (39) x 21-33 (45)^m towards
the middle and the basal region. Underleaves bifid, with narrow sinus extending
to 1/2 or more of length, lobes 5- 10 cells long and 5-7 cells broad, 275-407
(605) x 198-275 /mi, with smooth margin, apex acuminate, usually uniseriate,
cells polygonal, without conspicuous trigones. Rhizoids numerous, usually in
fascicles at the bases of the underleaves and also scattered on the stem ventrally.
Monoecious. Antheridial shoot ventral, in the axil of underleaf, 495-715/un long,
spiiate occasionally showing proliferation; antheridial bracts in 4-61 pairs, imbri-
cate, shortly bilobed, saccate, ventral lobe withlaciniate margin, incurved ; brae-
teoles bifid, one per pair of brae ts, usually 1/2 bifid with 3-celled uniseriate acuminate
apices ; antheridium single in the axil of each bract, body sub-globose, 56-67
(141) x 64-67 (l47)/mi with irregularly arranged jacket cells, stalk biseriate,
21-33 /jm long. Archegonial branches ventral, in the axil of underleaves, arche-
gonia 3-4 at the apex surrounded by small perichaetial leaves. Marsupium ini-
tially small, tuber lifce, later becoming fleshy and cylindrical, 2-2- 5 mm long, with
narrow base, studded with numerous rhizoids. Sporophyte with an anchor-shaped
multicellular foot buried at the base of the marsupium ; seta cylindrical, 7-8 cells
across, up to 2cni long, hyaline ; capsule cylindrical, deep brown, dehiscing in
four equal valves extending up to the base of the capsule, capsule wall bistratose,
outer layer of cells quadrate to elongated, with 2-phase development, thickenings
nodulose on the radial walls, extending slightly on the tangential walls, marginal
cells of each valve lacking any such thickenings ; inner layer of quadrate to elon-
gated cells with complete-incomplete bands on the inner tangential walls usually
connecting the radial bands at both ends. Spores yellowish to reddish-brown,
globose to sub-globose 8-12 #m in diameter, exine minutely papillose. Elaters
reddish-brown, 9 6- 192 /on long, 7-8 ^m broad, bispiral to occasionally trispiral
with blunt to tapering ends.

2.1. Distribution

Europe, America, Russia, Japan and India.

2 • 2. Specimens examined :

LWU 4279, 4390 Coll. S. C. Srivsstava, Dinesh Kumar and D. K. Singh : Loc :
Ou way to Hemkund from Ghangaria (Valley of Flowers), alt. ca 4670 meters
Date 22 May 1980. E>et. R. Udar, S. C. Srivastava and D. Kumar.

Geocalyx. — a rare marsupial genus

141

Figures 1-25. Geocalyx, graveolens. 1. Plant showing a male branch, a mature
marsupimn (with exserted sporophyte) and several tuberous young marsupia.
2. A male branch showing proliferation. 3. T.S. stem. 4, 5. Bifid leaves.
,6, 7. Apices of the le?f lobes. 8. Leaf cells from middle and base. 9-12.
Underleaves. 13. Male bract. 14. Antheridium. 15. Male bracteole. 16. L.S.
marsupium with sporophyte. 17. T.S. seta. 18. Base of capsule. 19. T.S.
capsule wall. 20. Cells of the outer layer of capsule wall. 21, 22. Cells of the
inner layer of capsule wall. 23, Spore. 24, 25. Elaters,

142 Ram Udar, S C Srivastava and Dhinndra Kumar

3. Remarks

This taxon grows in the alpine or subalpine zone of the western Himalayas (ca
46?a m) with low temperature and long seasonal periods of snow cover. It exhi"
bits a variety of habitat preferences as well as liverwort associates because of consi"
' derable variation in the microclimate where these plants grow. Growth of the
plants is favourable under diffused sunlight on soil covered rocks, on decaying
forest litter or at the base of small bushy trees at different locations along with
Lophocolea minor Nees, Lopkozia incisa (Schrad.) Dam., Haplomitrium hookeri
(Sm.) Nees, Blepharostoma trichophyllum (L.) l>um., Jungermannia (Plectocolea)
limbatifolia Amak. and species of CalypogeiaR&ddi, Plagiochila Dum. and Jame*
sonlella (Spruce) Schiffn. A more or less similar habitat preference as well as the
liverwort associates at the generic level have been reported by Schuster (1953) in
American population. Haplomitrium hookeri has. been found for the first time
growing in association with Geocalyx graveolens.

The stem, bearing a number of marsupia on the ventral surface, is highly
undulated and internally un differentiated (figures 1,2). Succubously arranged
leaves are obliquely inserted and slightly raised above the substratum forming a
continuous, gutter (channel) over the dorsal surface rather characteristic of this
tax: on. Both leaves and underleaves are typically bifid and the margin is usually
entire except in few underleaves where a tendency of developing additional lobe
on one or both sides is present.

The sexual branches are usually short and arise from the postical surface In the
axil of underleaves (figure 1) as also in Jackiella javanica. The antheridial branches
som'tiims proliferate (figure 2). The bracteoles are similar to amphigastria and
are relatively smaller, sometimes bearing rhizoidal outgrowth (figure 15). Owing
to the absence of perianth the archegonia having short necks are surrounded by
small pcrichaetial leaves which persist till the capsule remains embedded within
the marsupium. In the early stage of development there is a rapid elongation of
the lower side of the stem of the archegonial branch and eventually the apex bends
upwards. After fertilization the tissue beneath the archegonial group undergoes
rapid msristematic activity causing the formation of initially a sm? 11 ventral tubei>
like marsupium (figure 1). According to Schuster (1966) the development of the

marsupium takes place " due to an auxin secreted by the embryo or at least

by something derived from the embryo — the embryo factor/' However, there is
no experimental proof regarding this contention. Simultaneously with the forma-
tion of marsupium the development of sporophyte also takes place. As the growth
continues further the marsupium becomes pendulous and cylindrical, enclosing the
developing sporophyte. The mar&upium grows downwards into the substratum
and remains studded with dense rhizoids of the simple type. The sporophyte even
up to a late stage of development remains embedded within the marsupium. When
the sporophyte is mature the seta elongates considerably and the capsule is pro-
truded out of the calyptra as well as the marsupium. The foot is niulticellular,
anchor-shaped and remains embedded in the basal tissue of the marsupium (figure
16) unlike that of Jackiella which has a large haustorial collar consisting of nume*
rous uniseriate septate filaments (coiinate at base) arising from the junction of
foot and seta. The outer layer of capsule wall shows biphasic development with
nodulose thickenings on radial walls slightly extending over the tangential walls

Geo calyx — a rare marsupial genus 143

(figure 20). The cells of the inner layer have incomplete or semiannular bands
formed by the fusion of thickenings on the radial walls and the tangential walls
(figure 21). Such thickenings appear T-shaped or very rarely cL'-shaped in the
outer layer and cU'-shaped in the inner layer of cells in a transverse section (figure
19).

The plant has been discovered from high altitude zone of the western Himalayas
— the liverwort flora of which still remains unexplored. A cursory survey of the
entire collect! on of liverworts revealed that the area is extremely rich in rare
taxa which are either poorly known or still not described in Indian bryology.

Acknowledgements

The authors are thankful to the University Grants Commission, New l>ethi, for
financial assistance. Contribution, from the Department of Botany, University
of Lucknow, Luc know, New Series (Bryophyta) Ho. 140.

References

Banner C E B 1965 Index Hepaticanim VI Germany

Evans A W 1939 Classification; of the Hepaticae ; JBot. Rev. 5 45-96

Hattori S 1948 Contributio. ad floram hepaticarum yakusimcnsem III ; J. Hattori JBot. Lab. 3

1-35

Hferzog Th. 1935 Description of new species of N,Z. Hepaticae; Trans. R. Soc. N.Z. 65 365
Hodgson E A 1958 New Zealaad Hepaticae (Liverworts) X-Marsupial Genera of N.Z. ; Trans.

R. Soc. N.Z. 85 656-6S4

Macvicar S M 1926 The students handbook of British hepatics (London: Eastbourne)
Miiller K 1951-58 Die Lebermoose Europas-An Rabenhorst's Kryptagamen-Flora, ed. 3

(Leipzig)
Schuster R M 1953 Boreal Hepaticae ; A manual of the liverworts of Minnesota and adjacent

regions ; The Am. Middle Nat. 49 257-684
Schuster R M 1966 The Hzpaticae and Anthocerotae of North America Vol. I (New York and

London : Columbia University Press)
Schuster R M 1972 Phylogenetic and taxonomic studies on Jungermanniidae ; J. Hattori Bot.

Lab. 36 321-405

Stephani F 1906-1909 Species Hepaticarum III (Geneve)
Udar R and Kumar A 1981 Jackiella javanica SchifTn. A rare and interesting taxon ; J. Indian

Bot. Soc. 60 105-111
Udar R and Kuimr A 19S2 The Genus Jackiella in. south India; Proc. Indian Acad. Sci. 91 131-137

Proc. Indian Acad. Sci. (Plant Sci.), Vol. 91, Number 2, April 1982, pp. 145-152.
© Printed in India.

Ontogeny of the paracytic stoma: Variations and modifications

PARVEEN FAROOQUI (nee KIDWAI)

Department of Botany, The University, Allahabad, India

Present address : Regional Forest Research Centre, Jabalpur 482 020, India

MS received 6 March 19S1 ; revised 2 April 1952

Abstract. It was generally believed that the topography of the cells surrounding
the guard cells in the mature condition indicate their mode of development. How-
ever, it has now been established that more than one ontogenetic type may corres-
pond to a single mature type, or it may lead to the development of varied stoma tal
types. The paracytic stoma was studied from this viewpoint. It was found that
it may be formed through one of at least eight different modes. These are classified
and reviewed. The need to undertake studies on. the ontogeny of this type of stoma
in various groups of plants has been emphasized.

Keywords. Paracytic ; stoma ; ontogenetic types ; variations.

1. Introduction

Metcalfe and Chalk (1950) defined the paracytic stoma as a stoma "accompanied
on either side by one or more subsidiary cells parallel to the long axis of the pore
and guard cells ". Earlier, Vesque (1881, 1889) named such stomata as " Rubia-
ceous " and cited the family Rubiaceae as a typical example. Florin (1931, 1933,
1934), based on his studies of gymnosperms alone, proposed the term " syndeto-
cheilic " for such stomata, but the name also implied a mesogenous development
of the subsidiary cells. Developmental studies on stomata of the Rubiaceae by
Tognini (1897) and Pant and Mehra (1965) confirmed that the parallel subsidiaries
in these plants were formed from the same mother cell as the guard cells.

It was, therefore, generally believed that paracytic stomata always developed in
a mesogenous or syndetocheilic manner. However, it has been well established
that the developmental type should not be inferred from a study of the mature
stomatal complex alone but actual developmental studies should be carried out.

Recent work (Stebbins and Jain 1960 ; Stebbins and Khu&h 1961 ; Pant and
Kidwai 1966 ; Tomlinson 1974) on monocotyledons has brought to light the fact
that here the paracytic stomata develop perigenously and the subsidiary cells are
formed from lateral cells adjacent to the guard cells.

Indeed, the paracytic stomata, formed by these two entirely different modes, viz,
mesogenous and perigenous, look so similar that it has aroused a great deal of

145

146 Parveen Farooqui (nee Kidwai)

controversy and various authors have described their formation by different onto-
genetic pathways in one and the same plant. In Equisetum, Duval-Jouve (1864)
and Chatterjee (1964) mentioned that the stomatal meristemoid first divides by a
periclinai wall, each of these segments then divides by an anticlinal wall to form
two overlapping subsidiaries from the outer cell and two sunken guard cells from
the inner cell. However, other authors (Strasburger 1866-1867 ; Johnson 1933 ;
Hauke 1957 ; Pant and Mehra 1964a ; Pant and Kidwai 1968) found no periclinal
division and the stomata according to them are of the ordinary paramesogenous
type.

In Gnetum, tie development of stomata was described as haplocheilic or peri-
genous by some authors (Maheshwari and Vasil 1961a, b ; Inamdar and Bhatt
1972) whereas, others on the basis of division figures have concluded that the two
parallel subsidiary cells are formed inesogenously (Takeda 1913b : Kaushik 1974 ;
Nautiyal et al 1976 and others), but as they fall short of the poles, the stomata may
even be termed mesoperigenous.

The paracytic stoma is perhaps the most common among plants, occurring in
pteridophytes, gymnospenns, monocotyledons and dicotyledons. Dilcher (1974)
has pointed out that " more than one ontogenetic type may correspond to a single
mature stomatal type ". This is perhaps nowhere better illustrated than in the
paracytic stoma and a detailed study of its development based on published litera-
ture and illustrations has brought to light a number of different ontogenetic path-
ways which ultimately result in the formation of a paracytic stoma in the adult
condition, the three main types being the perigenous, the mesoperigenous and the
mesogenous types. Some of these types have not actually been reported but as
the possibility of their discovery at a later date remains, they have been included
here. Some types have been reported only as variations of other predominant
types and not as characteristic of any particular taxon. The terminology according
to any particular author is indicated by appropriate reference. A few new terms
are also being introduced for the first time here. The following main categories
have been found (figure 1).

2. Ontogenctic pathways leading to the formation of pamcytic

2-1. Perigenous type (Pant and Mehra 1964b)

Here a protoderrn cell becomes directly converted into the guard cell mother cell
and divides once to form the two guard cells. The parallel subsidiary cells are
formed from cells on the side of the guard cells (figure 1-1).

(a) Hemipam-perigenous type : Only one lateral subsidiary cell is formed from a
perigene cell on the side of a guard cell and the stoma is of the hemiparacytic
type in the mature condition (figure Ma). Fryns Classens and Van Cotthem
(1973) named it the mono-perigenous type but here it has been termed the hemipara-
perigenous type in keeping with their hemipara mesoperigenous type. Although
this type has not been described so far, Fryns Classens and Van Cotthem (1973)
suggested that some stomata on Polygonum lanigerum (Inamdar 1969b) may follow
this developmental pathway.

Ontogeny of the paracytic stoma

147

ONTOGENETIC MODES OF PARACYTIC STOMA FORMATION

FIRST
DIVISION

SECOND

DJVIfilOk

THIRD
DIVJSIOJM

FOURTH flFTH MATURE
DIVISION

D

c

(GO

D

1-2

D-

(D

ccn

a

CD

CDj

(IDS

SO

D

(D

o

am

1-3

a

OB

a

CD

GB

a

D

00

Figure i. Diagrammatic representation of the different ontogenetic modes of
paracytic stoma formation. The perigene cells and their divisions are represented
by dotted lines. 1-1. Perigenous. a. Hemi-para perigenous. b. Paraperigenous.
1-2. Meso perigenous. a. Herni-para-mesapcrigenotis. b. Para-meso perigenous
(Type I), c. Para-mesoperigenous (Type II). 1-3. Mesogenous. a. Para-msso-
genous. b. Hemi-parallelo-mesogenous. c. Parallelo-mesogenous.

(b) Para-perigenous type : The stoma is formed in the perigenous manner and the
two parallel subsidiaries are formed by asymmetric divisions in the cells lying on
the two lateral sides of the guard cells (figure 1 . Ib). This has been termed the
biperigenous (Paliwai 1969) or the diperigenous (Fiyns Classens and Van Cotthem
1973) type but for the sake of uniformity the term paraperigenous is preferred here-
This type is common in the monocotyledons and has been described in the Musa-
ceae, Gramineae, Juncaceae, Cyperaceae, Alismataceae, Marantaceae (Strasburger
1866-1867 ; Porterfield 1937 ; Flint and Moreiand 1946; Stebbins and Jain 1960 ;
Metcalfe 1960), twenty-four families of monocotyledons (Stebbins and Khush 1961),
Araceae (Pant and Kidwai 1966), Commelinaceae, Zingiberaceae (Tomlinson
1969), Centrolepidaceae and Phydraceae (Paliwai 1969) and others.

2.2. Mesoperigenous type (Pant and Mehra 19646)

The cells surrounding the guard cells in the adult condition have a dual origin,
some being formed from the same meristemoid as the guard cells while the others
are perigenous in origin (figure 1.2).

P.(B)-6

148 Parveen Farooqui (nee Kidwai)

(a) Hemipara-mesoperigenous type (Fryns Classens and Van Cotthem 1973) : The
single lateral subsidiary cell is formed from the stomatal meristemoid (figure 1 -2a)
before the formation of the two guard cells, e.g., Lophosoria (Kondo and Toda
1956, 1959), some Rubiaceae (Pant and Mehra 1965), Nyctaginaceae (Inamdar
1968), Bigonia (Inamdar 1969a), Crotalaria (Shah and Gopal 1969), Kalanchoe
(Inamdar and Patel 1970), Polemoniaceae, Boraginaceae and Solanaceae (Patel and
Inamdar 1971) and some stomata of Rauwolfia and Tabernae montana (Trivedi and
Upadhyay 1976) and Annona (unpublished observations).

(b) Para-mesoperigenous type I or Para-eumesoperigenous type : The adult para-
cytic stonia is surrounded by two parallel subsidiaries, one of which is formed from
the stomatal meristemoid and the other by a division in the perigene cell on the
other side of the guard cells (figure l-2b). This type is the true mesoperigenous
paracytic type (s/type l-2c) and although it has not been described so far, it has
been included here as the possibility of its occurrence exists. According to Payne
(1970), some stomata in Liriodendron tulipidera may be formed in this manner.

(c) Para-mesoperigenous type II or Para-pseudomesoperigenous type : The adult
stoma is paracytic and its two lateral subsidiary cells are formed from the same
initial as the guard cells (mesogenous). However, they do not meet at one or
both poles of the guard cells and the neighbouring perigene cells abut on the polar
ends of the guard cells (figure l-2c). Due to this falling short of the mesogene
lateral subsidiaries towards the poles, the stoma is considered as mesoperigenous.
This view has been expressed by Pant and Mehra (1964), Pant (1965), Fryns Clas-
sens and Van Cotthem (1973) and Nautiyal et al (1976), e.g., Wdwitschia (Takeda
1913a), Gnetum (Takeda 1913b; Nautiyal et al 1976), Drimys (Bondeson 1952),
Linum (Paliwal 1961), Borreria and Oldenlandia (Pant and Mehra 1965), Bignonia
(Inamdar 1969a), Polygonaceae (Inamdar 1969b), Jasminwn (Inamdar et al 1970),
Claytonia (Payne 1970), Casuarina (Pant et al 1975), Xylonymus and Brassianthus
(Den Hartog and Baas 1978), and some stomata in Zornia (Kannabiran 1975a),
Abrus precatorius (Kannabiran 1975b), Dipteria (Khare 1978) and Annona (un-
published observations).

If the mesogene subsidiaries meet at only one of the poles, the stoma is frequently
surrounded by three cells and appears aniso- or tricytic, eg. some stomata in Rumex
(Verma 1975), Zornia (Kannabiran 1975a), Brassiantha (Den Hartog and Baas
1978) and Annona (unpublished observations).

2-3. Mesogenous type (Pant and Mehra 1964ft)

The lateral subsidiary cells are mesogenous in origin and they completely flank
the guard cells so that no other cells immediately surround the guard cells (figure
1.3).

(a) Para-mesogenous type (Fryns Classens and Van Cotthem 1973) : There are
only two lateral subsidiary cells, parallel to the guard cells and these are meso-
genous (figure l-3a). Recorded from Cheiropleuriaceae, Dipteridaceae and
Dicksoniaceae, Phorbitis, Basella and Opuntia (Strasburger 1866-1867), Convol-
vulus, Euphorbiaceae, Impatiens and C<^a(Tognini 1897). Magnoliaceae (Paliwal

Ontogeny of the paracytic stoma 149

and Bhandari 1962 ; Pant and Gupta 1966), Cheiroplueria and Cibotium (Kondo
and Toda 1959), Convolvulaceae (Pant and Banerjee 1965), Rubiaceae (Pant and
Mehra 1965), Strapleonema (Stace 1965), Portulaceceae, Trianthema (Ramayya and
Rajagopal 1968), Dicksoniaceae, Dipteridaceae and Lopho&oriaceae (Van Cotthem
1970), Nyctaginaceae, Polygonales, Centrospermae, Zygophyllaceae, Simerubaceae,
Salvadoraceae, Bigonia and Ipomea (Inarndar 1968, 1969a, b, c, d, e), Leguminosae
(Shah and Gopal 1970; Bora and Baruah 1979; Farooqui 1979), Zornia (Kannabiran
1975a), Abrus precatorius (Kannabiran 1975b), Rauwolfia and Chatharanthus
(Trivedi and Upadhyay 1973, 1976, 1977), Bhesa sp. and Hedraianthera (Den
Hartog and Baas 1978), Dipteris (Khare 1978) and many others.

(b) Hemiparallelo-mesogenous type : Three cells are cut off from the stomatal
meristemoid in an alternate fashion so that the stoma is surrounded by 2 inner
subsidiary cells and an outer encircling cell (figure l*3b). This, was first described
by Payne (1970) under the parallelocytic type. However, as only one additional
cell is formed, it has been separated here, e.g., some stomata of Cinchona
succimbra (Pant and Mehra 1965), Magnoliaceae (Pant and Gupta 1966) and
Rauwolfia serpent ina (Trivedi and Upadhyay 1976).

(c) Parallelo-mesogenous type (Fryns Classens and Van Cotthem 1973): The meriste-
moid, instead of forming only two parallel subsidiary cells divides further in the
same alternate manner and forms two or more additional cells (figure 1 . 3c) which
surround the subsidiaries and axe parallel to them (encircling cells), e.g., Cactaceae,
Convolvulaceae, Euphorbiaceae, Leguminosae, Portulacaceae, Rubiaceae and
Umbelliferae (Payne 1970), Euonymus globularia (Den Hartog and Baas 1978).

The paracytic stoma may also show variations by further divisions in its sub-
sidiary cells. If one of the subsidiary cells divides the stoma may appear tricytic
or anisocytic in the adult condition. If both the subsidiaries divide, the stoma may
appear tetracytic or anomocytic. All these variations have been noticed in
Strychnos (unpublished observations). Therefore, although their adult form may
differ, they begin their development in essentially the same manner.

3. Discussion

The paracytic stoma may be formed by many different ontogenetic pathways. It
is, therefore, essential that actual developmental studies should be carried out
before their ontogeny can be determined.

Of the eight categories described, the most common are the para-perigenous,
para-pseudomesoperigenous and the para-mesogenous types. The others repre-
sent variations of these and their occurrence is limited or not reported so far.
A study of development in varied groups of plants may lead to their discovery at a
later date.

Metcalfeand Chalk (1950) have listed 105 dicotyledonous families in which the
paracytic type of stoma is predominant. In addition a number of monocotyledons'
pteridophytes and gymnosperms also have the paracytic stoma. There is an
urgent need to work out the exact mode of stomatal development in all these taxa.

P.(B)-7

150 Parveen Farooqui (nee Kidwai)

There are other paracytic stomata, like those of the Cycadeodales, whose develop-
ment will probably never be determined because they occur only as fossils.

According to Takhtajan. (1969) the paracytic type of stoma is the most likely
basic type of stomatal apparatus in the evolution of flowering plants. Its occur-
rence in such diverse groups as the pteridophytes, gymnosperms, monocotyledons
and dicotyledons may support such an assumption.

References

Bofa N and Baruah R 1979 Contiguous stomata in Destnodiwn Desv. (Papilionaceae) ; Curr.

Sa. 48 27-28
Bondeson W 1952 Entwicklungsgeschichte und Ban der Spaltoffnungen bci den Gattungen

Trochodendron Sleb. et Zucc., Tetracentron Oliv. mid Drimys J. R. ctg. Forst. ; Acta Hort.

berg. 16 169-217
Chatterjee J 1964 Stomata in Equisetum ramosissimu/n Desf, sub.sp. ramosissimwn. ; Phyto*

morphology 14 451-457
Den Hartog R M and Baas P 1975 Epidermal characters of the Celastraceae sensu lato ; Acta

Bot. Neerl 27 355-388
Dilcher D L 1974 Approaches to. the identification of angiosperm leaf remains ; Bat. Rev. 40

1-1 57

Duval-Jouve J 1864 His tor ie naturelle dcs Equisetum de France, Paris
Farooqui P 1979 Sequence of stomatal meristemoid formation in some Leguminosao ; Curr. ScL

48 489-490

Flint L H and Moreland C F 1946 A study of stomata in sugarcane ; Amer. J. Bot. 33 80-82
Florin R 1931 Untersuchungen zur Starnrnens— geschichte der Coniferales und Cordaitales ;

K. Svensk. Vet. Akad. Handl. 10 1-588
Florin R 1933 Studien iiber die Cycadales des Mosozoikums nebst Erorterungen iiber die

Spaltoffnungsapparate der Bennettitales ; K. Svensk. Vet. Akad. 12 1-134
Florin R 1934 Die Spaltoffnungsapparate von Welwitschia miraUlis Hook. f. ; Svensk. Bat.

Tidskr. 28 264-289
Fryns Classens E and Van Cotthem W 1973 A new classification of the ontogenetic types of

stomata ; Bot. Rev. 39 71-13S

Hauke R 1957 The stomatal apparatus of Equisetum ; Bull. Torrey bat. Club 84 178-181
Inamdar J A 1968 Epidermal structure and ontogeny of stomata in some Nyctaginaceae ; Flora*

158 159-166
Inamdar J A 1969a Structure and ontogeny of foliar nectaries and stamata in Bignonia chamber

laynii Sims. ; Proc. Indian Acad. Sci. 70 232-240
Inamdar J A 1969b Epidermal structure and stomatal ontogeny in some Polygonales and

C^ntrospermae ; Ann. Bot. 33 541-552 t "

Inaradar J A 1969c Epidermal structure, stomatal ontogeny and relationship of some

Zygophyllaceae and Simarubaceae ; Flora 158 360-368
Inamdar J A 1969d The stomatal structure and ontogeny in Azima and Salvadora ; Flora 158

519-525
Inamdar J A I969e Development of stomata on foliar and floral organs of two species of Ijpomoea ;

J. Ind. Bot. Soc. 48 173-176
Inamdar J A and Bhatt DC 1972 Epidermal structure and ontogeny of stomata in vegetative

and reproductive organs of Ephedra and Gnetum; Ann. Bot. 36 1041-1046
Inamdar J A, Chauh.au A J and Patel R C 1970 Morphological studies in Jctsminum officiwle

Linn.; Vidya 13 10&-113
Inamdar J A and Patel R C 1970 Structure and development of stomata in vegetative and floral

organs of three species of Kalanchoe ; Ann. Bot. 34 965-974
Johnson M A 1933 Origin and development of tissues in Equisetum sdrpoides ; Bot. Gaz. 94

469-494.
Kannabiran B 1975a Epidermal structure and stomatal o-ntogeny in Zornia Gmel ;

Aust.J.Bot. 23 327-333 •< .........

Ontogeny of the paracytic stoma 151

Kannabiran B 1975b A note on the stomatal ontogeny and systematic position of Abrus

precatorius ; Curr. Sci. 44 675-677

Kaushik S B 1974 Ontogeny of the stomata in Gnetum ula Brongn. ; Bot. J. Linn. Sac. 68 143-152
Khare P K 197& Epidermal structure and ontogeny of stomata in Dipteris wallachii ; Phyto-

morphology 28 400-405
Kondo T and Toda H 1956 A contribution to the study of fern stomata, with special reference

to their development and structure ; Rec. Bull. Fac. Educ. Siiizuoka Univ. 5 60-80
Kando T and Toda H 1959 Some notes on the stomata of Woodwardic, ; J.Jap. Bot. 34 18-22
Maheshwari P and Vasil V 1961a The stomata of Gnetum ; Ann. Bot. 25 313-319
Maheshwari P and Vasil V 1961b Gnetum, Bot. Monograph No. 1, C.S.I.R., New Delhi
Metcalfe C R 1960 Anatomy of the monocotyledons. I. Gramineae (ed.) Metcalfe C R (Oxford •

Clarendon Press)

Metcalfe C R and Chalk L 1950 Anatomy of the dicotyledons I and II (Oxford : Clarendon Press)
Nautiyal D D, Singh S and Pant D D 1976 Epidermal structure and ontogeny of stomata in

Gnetum gnemon, G. montanum and G. ula ; Phy to morphology 26 282-296
Paliwal G S 1961 The development of stomata in Llnum usitatissimum L. ; Curr. $ci. 30 269-271
Paliwal G S 1969 Stomatal ontogeny and phylogeny. I. Monocotyledons ; Acta Bot Neerl 18

654-668
Paliwal G S and Bhandari N N 1962 Stomatal development in some Magnoliaceae ; Phyto-

morphology 12 409-412
Pant D D 1965 On the ontogeny of storaata and other homologous structures ; p\. Sci Ser

1 1-24
PantD D and Banerjee R 1965 Epidermal structure and development of stomata in some

Convolvulaceae ; Senck. biol. 46 155-173

Pant D D and Gupta K L 1966 Development of stomata and foliar structure of some Magno-
liaceae ; /. Linn. Soc. (Bot.) 59 265-277
Pant D D and Kidwai P 1966 Structure of leaves and stomatal ontogeny in some Pandanales

and Spathiflorae ; Senck. biol. 47 309-333

Pant D D and Kidwai P 1968 Development of stomata in Equisetum ; Ann. Bot. 32 601-608
Pant D D and Mehra B I964a Development of stomata in some fern allies ; Proc natn

Inst. Sci. India 3 0 92-98

Pant D D and Mehra B 1964b Ontogeny of stomata in Ranunculaceae ; Flora 155 179-188
Pant D D and Mehra B 1965 Ontogeny of stomata in some Rubiaceae ; Phytomorphologv 15

300-310
Pant D D, Nautiyal D D and Singh S 1975 Cuticle, epidermis and stomatal development of

Casuarina equisetifolia Forst ; Ann. Bot. 39 1117-1123
Patel R C and Inamdar J A 1971 Structure and ontogeny of stomata in some Polemoniales •

Ann. Bot. 35 389-409
Payne W W 1970 Helicocytic and allelocytic stomata; unrecognized patterns in the Dicotyle-

donae ; Am. J. Bot. 57 140-147
Popterfield W M 1937 Histogenesis in the bamboo with special reference to the epidermis •

Bull. Torrey Bot. Club 64 421-432

Ramayya N and Rajagopal T 1968 Foliar epidermis as taxonomic aid in ' The flora of Hydera-
bad'; Journ. Osmania Univ. (Sc) Hyderabad 147-160
Shah G L and Gopal B V 1969 Stomatal ontogeny of the vegetative and floral organs of some

Papilionaceae ; Aust. J. Bot. 17 31-&7
Shah G L and Gopal B V 1970 Structure and development of stomata on the vegetative and

floral organs of some Amaryllidaceae ; Ann. Bot. 34 737-749

Stace C A 1965 Cuticular studies as an aid to plant taxonomy ; Bull. Brit. Mus. (N.H.) 4 1-78
Stebbins G L and Jain S K 1960 Developmental studies of cell differentiation in the epidermis

of monocotyledons. I. Allium, Rhoeo and Commelina ; Dev. Biol. 2 409-426
Stebbins G L and Khush G S 1961 Variation in the organization of the stomatal complex in the
leaf epidermis of monocotyledons and its bearing on their phylogeny; Am. J. Bot. 48 51-59
Strasburger E 1866-1867 Bin Beitrag Zur Entwicklungsgescichte der Spaltoffnungen ; Jahrb wiss

Bot. 5 297-342

Takeda H 1913a Some points in the anatomy of the leaf of Welwitschia mirabilis : Ann Bot
27347-357

152 Parveen Farooqui (rtee Kidwf)

Takeda H 1913b Development of stomata in Gnetum gnemon\ Ann. Bot. 27 365-366
Takhtajan A 1969 Flowering plants— origin and dispersal (English translation Jeffrey C) (Edin-
burgh : Oliver and Boyd)
Tognini F 1897 Contribuzione allo studio della organogenic cornparata deglistomi; An. tnst.

Bot. Univ. Pavia 4 1-42
Tomlinson P B 1969 Anatomy of the monocotyledons III Commelmales— Zingiberales (ed.)

Mctcalfe C R (Oxford : Clarendon Press)
Tomlinson P B 1974 Development of the stomital complex as a taxonomic character in the

monocotyledons ; Taxon 23 109-128
Trivedi B S and Upadhyay N 1973 Studies on stomata and trichomes of Rauwolfia serpentina

L.; Curr. Sd. 42401-403
Trivedi B S and Upadhyay N 1976 Ontogeny of stomata in some members of Apocynaceae ;

GeophytoL 6 92-97
Trivedi B S and Upadhyay N 1977 Morphological studies in Apocynaceae Epidermal

structures ; GeophytoL 7 29-37

Van Catthem W R 1970 A class ificatio.n of stomatal types ; Bot. J. Linn. Soc. 63 235-246
Verma B K 1975 Ontogeny of stomata in leaves of Rumex dentatus Linn,; GeophytoL 3 24-29
Yesque J 1881 L'anatomie des tissus appliqee a la classification des plantes ; Nauv. Arch.

Mus. Hist. Nat. Paris 4 1-56
Vesque J 1S89 De Femploi des caracteres aaatomiques dans la classification des vegetaux ;

Bull Soc. Bot. Fr. 36 41-77

Prac. Indian Acad. Sci. (Plant Sci.), Vol. 91, Number 2, April 1982, pp. 153-158.
© Printed in India.

Growth response of some thermophilous fungi at different
incubation temperatures

S SINGH and D K SANDHU

Department of Biology, Guru Nanak Dev University, Amritsar 143005, India

M3 received 4 September 19S1 ; revised 17 March. 19S2

Abstract. Growth response of 21 thermaphilous fungi at 10 different tempe-
ratures from 15-62° C has been studied. These fungi could be categorized into
three groups, i.e., microtkermophiles, thermotolerant and true thermophiles. The
temperature relations of 6 thermophilous fungi namely Asjpergiltiis tamarii, A. terreus
v&T.aureus (microtliennophiles), A. nidulansv&x.echinulatus, A. viridi-nutcMS, A.fumi-
gatus var. ellipticus and A. caespitosus (thermotolerant) are being reported for the
first time. The growth rates of different fungi varied from 0-19-1 -25 mm/hr at
their optimum temperatures. Penicittium sp. the slowest and Thermoascus sp. were
the fast growing fungi.

Keywords. Mtcrotherfnophiles ; thermotolerant ; thermophiles ; temperature ;
growth rates.

I. Introduction

Thermophilous fungi are those which show temperature optima in the range of
25 to 55° C (Apinis 1963), These fungi play a significant role in degradation of
plant material, composting and humification (Fergus 1971 ; Cailleux 1973 ; Tansey
and Brock 1978 ; Jain etal 1979). The occurrence of thermophilous fungi is
now known to be ubiquitous (Tansey and Brock 1978). In India these fungi
have also been reported from various substrates like compost (Maheshwari 1968 ;
Qureshi and Johri 1972), stored grains (Mehrotra and Basu 1975), coal mine
soils (Thakre and Johri 1976), nesting materials of birds (Satyanarayana etal 1977)
and from air (Thakur 1977). Recently, Sandhu et al (1980) and Sandhu and Singh
(1981) studied the occurrence and some ecological features of thermophilous fungi
associated with decomposing sugarcane bagasse, and forest soils. The present
study describes the effect of different incubation temperatures on the growth rates
and development of these fungi.

2. Materials and methods

Twenty-one fungi isolated from bagasse and forest soils were studied for their
response to different temperatures, The culture media used for the identification

153

154 S Singh and D K Saridku

of ^spergilU and Penicillium sp. were czapek's agar, malt extract agar and yeast
glucose agar respectively (Raper and Fennell 1973). Yeast starch agar (Cooney
and Emerson 1964) was used for the remaining fungi belonging to mucorales,
as«comycetes and hyphomycetes. Mycelial discs of 3 mm diameter from the peri-
phery of growing colonies were used as the inoculum and the plates were incu-
bated at 15° C, 20° C, 27° C and 32-62° C with 5° C interval. The petri plates
kept at 42-62° C were placed in polythene bags and a beaker of water was also
placed in each of these incubators to avoid desiccation. The bags were opened
twice daily to aerate the cultures. Growth response was recorded in terms of
colony diameter daily. Sporulation and cleistothecia production were noted
visually and confirmed by observing under the binocular and by slide preparations.
The growth rate of each fungus was calculated on yeast starch agar as follows :

— Average diameter in mm of four petri plates
rate "" Total time period in h

3. Results and discussion

Based on colony diameter the growth response of 21 fungi to 10 different incuba-
tion temperatures yielded 2 microthermophiles, 15 thermotolerant and 4 true
thermophiles (table 1). A comparison of our results with the literature revealed
that the temperature relations of 6 thermophilous fungi namely Aspergillus tamarii,
A. terreus var. aureus (micro thermophile), A. rtidularts var. echirtulatus, A. viridi-
nutans, A. fumigatus var. ellipticus and A. caespitosus are being reported for the
first time. The temperature relations of the other fungi studied, in general, fall
within the range described by Tansey and Brock (1978). Temperature responses
of Thermomyces lanuginosus, Humlcala grisea var. thermoidea and Mucor pusillus
are more close to those reported by Fergus (1964) and Evans (1971) respectively
than Apinis (1963) and Qureshi and Johri (1972). However, Humicola grisea var.
thermoidea differed from Evans (1971) since our isolates could not grow at the
minimum ^temperature of 20° C. The cardinal temperatures of Aspergillus fumi-
gatus and Sporotrichum thermophile varied widely from Qureshi and Johri (1972)
and almost fell within the range given by Evans (1971) and Tansey and Brock
(1978). In the case of Aspergillus nidulans and A. terreus the cardinal temperatures
were similar to those reported by Evans (1971) but had more variation of minimum
temperature than maximum. This may be due to the fact that in our study
minimum temperature was recorded up to 15° C only. The temperature variations
reported above may be due to strain variations, nutritional requirements and
temperature study at wider temperature intervals.

There was also variation in temperature regarding the optimum and the range
among the species and their varieties, e.g., Aspergillus fumigatus and A. fumigatus
var. ellipticus had their optimum temperature at 37° C and 42° C respectively
(figure 1). In the case of A. terreus and A. terreus var. aureus both had their
optimum temperature at 37° C. The former could grow up to 47° C while the
limit for the latter was at 42° C (figure 1 and table 1). There was a variation, in
optimum temperature for the development of the perfect and imperfect stage as
in the case of A. nidulans and A. nidulans var. echinulatus, maximum cleistothecia
formation was observed at 42° C while the sporulation was maximum at 37° Q,

Growth response of terhmophilous fungi 155

Aspergillus fumigatus r A. fumigatus var. ellipticus

A. mdulans var. echinuLatus
32 °C

24 48 72 95 120 U4 160 24 48 72 96 120 1U 168

HOURS OF INCUBATION

Figure 1. Growth response o.f ane microth.ermoplule (Aspergillus terreus var.
aweus) and 5 thermatolcramt fungi at different temperatures.

Similarly, abundant superficial cleistotkecia like bodies of A. caespitosus were
seen at 42° C while the optimum temperature for sporulation was .37° G The
temperature range for speculation and reproductive structures in all the groups
was narrower tha,n that for mycelial growth.

The growth rates of optimum temperature of 21 fungi varied from 0- 19-
l-25inm/hr (figure 2). Penicillium sp. was the slowest and Thermoapcus sp. the
fastest growing species. The fungi having growth rates of 0-19-0-39, 0-40-0 -75

156

Singh and D K Sctndhu

Table 1. Optimum temperature and raage for growth and sporulation of 21
thermopliilous fungi,

Group

Temperature Temperature Optimum

rarige (°C) for range (°C) temperature

mycelial for (° C)

growth sporulation

Fungi

Microthennophile 15-42

TJierrnotolerant I 15-47
(psyclirotoleraitt)

11 15-52

True thermophiles I 20-52

II 27-52

III 32-52

IV 32-57

20-37 32 Aspergillus taniarii(S*)

37 A. terreus var. aureus (B)

27-42 32 Paecilomyces varioti (B)

37 Aspergillus nidulans (S), A. nidulans

var. echinulatus (S), A. teneus (S),

A. caespitosus (B), Acrophialo-

phora fusispora (B), Penicilliurn

$P. (B)
42 Thielavia seped&nium (S), Absidia

carymbifera (B)
27-47 37 Aspergillus fumigatus (B), A. viridi-

nutans (S), Sporatrichum thermo~

phile (B)
42 Aspergillus fumigates var. ellipticus

(S), Absidia spinosa (B), Rhizopus

microsparus (S)

27-47 42 Mucor pusillus (B, S)

32-47 47 Humicola grisca var. thermoidea (B)

37-47 42-52 Thevmoascus sp. (S)

37-52 47 Thermornyces lanuginosus (Br S)

Source of fungi. B, bagasse and S, forest soil.

and 0-86-1-25 mm/hr were categorized as slow, moderate and fast growing
respectively. Comparison of growth rates at the optimum temperatures of these
fungi show that 8 are slow growing and out of these 2 are microthermophiles and
6 thermotolerant. Nine species belong to moderately growing group, 7 of which
are thermotolerant and 2 true thennophiles. In the fast growing group 2 are true
thermophiles and 2 thermotolerant.

The growth response of these fungi to diiferent temperatures was determined
on the basis of colony diameter. According to Hawker (1950) and Cochrane
(1958) the colony diameter alone does not account for density height and depth
of the colony. However, it has been justified for studies in which only one
environmental variable is studied for example temperature (Branc^to and Golding
1953 ; Cochrane 1958 ; Evans 1971 ; Trinci 1971 ; Tansey 1972).

(tnwth respvnxf <*f thrmvphihus fungi

I* growth rites at optimum temperature of 2!

fiiiiil fell, moderate and slow growing groups. Figures

llti bar* af inoubttion In hours.

Apiitti A I! t%l cif ili«r«itiplilliiiti micrafui^gi In certain alluvial soils near Notting-

ham ; /tow S 57-7*

I1 I1 und (folding H S i**ll Th« diametor of the mold colony as a reliable measure of
growth ; 4S

Caiileux R tf II Mtyaitlur^ du compo&i dnUin6 t It culture du champignon dc Couche ; jRev.

A(f>f*/, 17 14-35
C\«lifiiw V W t^H PhwMogv «ff«**l (Hew York : John Wiley)

l> Cl mid i'.morvm H IW4 fhtrmopkUtefitiig: An account of their biology, activities and
el*i*l/tr*tkm {San Frat»d«co and i*mdi»» : W H Frdwnan)
II C W7t Thermnphitoii« fungi of coal %poll tips* IL Occurrence, distribution and tempe-

relationship : Tram* Mr, Myp#L $®c> 57 255-266
C L 1971 Ttitrmtiphific tad thermotolerant molds and actinomycetes of mushioom

ttring ; W 267-284

Hawker L I! iWQ «//m«rf : Univ. of London Press)

Jain M K, K 1C m»d M M 1979 C^llul^e activity, degradation of cellulose and

httmu» formitl0n by ilttffii^tiilie funfi; Thvtf. Br. M^c<?/. *Soc. 72 85-89

S Singh and D K Sandhu

Maheshwari R 1968 Occurrence and isolation of thermophilic fungi ; Curr. Sci. 37 277-279
MTehrotra B S and Basu M 1975 Survey of the microorganisms associated with cereal grains

and their milling fractions in India. Part I. Imported wheat ; Int. JBiodeterior. Bull. 11 56-63
Qureshi A R and Johri B R 1972 Temperature relationship of some thermophilic fungi ; Bull.

Bat. Soc. Univ. Sagar 19 28-33

Raper K B and Fennell D I 1973 The genus Asjpergillus (New York : Robert E Krieger)
Sandha D K, Singh S and Waraich M K 1980 Thermophilous fungi of decomposing sugar-cane

bagasse; Can. J. Bot. 58 2015-2016
Sandhu D K and Singh S 19S1 Distribution of thermophilous microfungi in forest soils of

Darjeeling (Eastern Himalayas) ; Mycopathologia 74 79-86
Satyanarayana T, Johri B N and Sakessena S B 1977 Seasonal variation in mycoflora of nesting

materials of birds with special reference to thermophilic f ungi ; Trans. Br. Mycol. Soc. 68

307-309

Tansey M R 1972 Effect of temperature on growth rates and development of the thermophilic

fungus Cheetomium thermophile ; Mycologia 64 1290-1299
Tansey M R and Brock T D 1978 Microbial life at high temperatures : ecological aspects ; in

Microbial life in extreme environments (ed.) D J Kashner (London : Academic Press)
ThakreRP and Johri B N 1976 Occurrence of thermophilic fungi in coal mines soils of Madhya

Pradesh; Curr. ScL 45 271-273

Thakur S B 1977 Occurrence of spores of thermophilic fungi in the air at Bombay ; Mycologia
69 197-199

Trinci A P J 1971 Influence of the width of the peripheral growth zone on the radial growth
rate of fungal colonies an solid media ; /. Gen. MicrobioL 67 325-344

Proc. Indian Acad. Sci. (Plant Sci.), Vol. 91, Number 2, April 1932, pp. 159-174.
© Printed in. India.

Studies on Beggiatoa : Distribution and growth in aquatic
habitats of Visakhapatnam

M R R MOHAN and A NARAYANA RAO
Department of Botany, Andhra University, Waltair 530 003, India

MS received 19 June 1981

Abstract. Bzggiatoa is distributed in 11 of the 12 polluted aquatic habitats occurring
on the sediment and decaying leaves as a thick white scurn at 2, in detectable popu-
lation at 6, and in very less population at 3 habitats where H2S necessary for its
growth is produced. The Beggiatoa spp. present in these habitats are considered
as belonging to the 6 species described in Sergey's manual. J9. minima with less
than 1 -0, B. leptomitiformis with 1 - 66, B. alba with 3-32 and 4-9S ju wide trichomes
are distributed in 2, 6, 11 and 2 habitats respectively, which are fresh water, brackish
and marine habitats. B. arachn&idea with 9-96, B. mirabilis with 17-0 and
B. gigantea with 28- 22 & wide trichomes are present in only one brackish habitat.
B. alba (3-32 fji) occurs alone at 2 habitats, and in association with either one or
more of the other Beggiatoa species at the remaining 9 habitats. Except B. mirabilis
and B. gigantea, the others distributed in the 11 habitats could multiply in enrich-
ment culture media that contained extracted rice straw pieces and waters from
their respective habitats, and aggregate into visual white colonies, or loose clumps
in a thick film on the surface of medium, rice straw pieces and glass. B, mirabilis
and B. gigantea seem to be halophilic, truly autotrophic and more exacting than
the other species that exhibited differences in salt tolerance when grown in crude
enrichment culture media of differing salt content. It appears that physiological
strains or groups differing in salt tolerance may be existing in these species of
Beggiatoa.

Keywords. Beggiatoa ; distribution ; growth ; aquatic habitats ; Visakhapatnam *9
enrichment culture medium.

1. Introduction

Beggiatoa is a large, colourless, filamentous, gliding bacterium, and easily recogni-
zable. It is usually considered as carrying on chemolithotrophic nutrition, oxidizing
H2S, and depositing sulphur intracellularly as droplets. It is widespread in
distribution, occurring in lake bottoms, stream beds, ponds, sulphur springs,
brackish and marine habitats; even in the rice rhizosph ere (Pitts etal 1972) and
water-saturated soils of rice fields (Joshi and Hollis 1977), playing an important
role, the detoxification of H2S formed due to putrefaction, and activities of
anaerobic organisms such as Desulfovibrio spp. It is associated with, and also

159

160 M R R Mohan and A Narayana Rao

often described as indicative of strongly polluted polytrophic habitats. Ever
since the investigations, and the concept of chemolithotrophy by Winogradsky
(1887, 1888, 1889) attempts have been made (Keil 1912 ; Cataldi 1940 ; Faust
and Wolfe 1961 ; Lackey 1961 ; Scotten and Stokes 1962 ; Pringsheim 1964 ;
Kowallik and Pringsheim 1966 ; Jorgensen 1977) to increase the small number of
Beggiatoa trichomes present in samples from natural habitats into a rich popu-
lation by reliable enrichment culture techniques for isolation and cultivation in
pure culture media ; to study its distribution, morphology, growth and nutrition.
Pringsheim (1964) studied in detail the species differentiation of Beggiatoa, and
pointed out among other problems that the number and types of forms found
under a variety of ecological conditions are obscure.

The review of the literature showed the need for detecting and isolating as
many forms of Beggiatoa as possible for taxonomic differentiation, and studying
its nutrition ; and more so because there are no reports from India. Since
Visakhapatnam abounds with a variety of aquatic habitats that are continuously
polluted with urban refuse, sewage and industrial effluents, detailed investigations
are undertaken to study the distribution, growth, nutrition and species differen-
tiation of Beggiatoa. This paper presents 4 the observations and experimental
results on distribution and growth of Beggiatoa isolated from natural " polluted "
habitats, for the first time from India.

2* Materials and methods

Twelve sites of different aquatic habitats in Visakhapatnam were surveyed for
Beggiatoa. Their location and some relevant features are as follows :

Site 1 : The edge of a shallow one feet wide canal of a septic tank near the
Botany department garden, Andhra University campus. The entire edge is covered
with a thick white scum. The decaying leaves present in the canal and at the edge
are also covered with a white scum. The sediment is black silt. pH of water : 7,
salinity : 0 • 58 ppt.

Site 2 : The edge of a shallow sewage canal near Rama Krishna Mission,
Mahaianipet. The sediment is fine sand. pH of water : 7, salinity : 0-72 ppt.

Site 3 : The edge of a shallow sewage canal near Visakha women's college,
Maharanipet. The sediment is black silt. pH of water : 7, salinity : 1 • 07 ppt.

Site 4 : The edge of a stagnant pond receiving sewage water near " convent
junction". The sediment is black silt. pH of water: 7, salinity: 1-07 ppt.

Site 5 : The edge of a sewage stream near " naval coast battery ", Maharanipet.
The sediment is light black fine sand. pH of water : 7, salinity : 1-08 ppt.

Site 6 : The edge of a sewage stream near the wooden bridge, ferry road-old
post-office junction. The stream joins the harbour channel, and the sample site
is at about 200 metres from the channel. During high tide there will be a back
flow of sea water. The sediment is black silt. pH of water : 7, salinity : 1 • 4 ppt.

Site 1 : The edge of a stream adjacent to oil-loading tanks near naval head-
quarters. The bank is muddy, and covered with a thick asphalt-like layer mixed

Distribution and growth of Beggiatoa 161

with cakes and clumps of resin-like material, a conversion product of oil. pH
of water : 7-5, salinity : 4- 5 ppt.

Site 8 : The edge of the sea water canal carrying effluents from the Coromandel
fertilizers factory. The site is located near the Hindustan Petroleum Oil Refineries
entrance gate. The sediment is light red silt. pH of water : 5 and 2, salinity
19-0 ppt.

Site 9 : The edge of a stagnant brackish water pond near ' Chavulamadam '.
The sediment is black silt. pH of water : 8-5, salinity : 20 • 0 ppt.

Site 10 : The edge of one of the Harbour channel terminals near sulphur-
unloading berth of the harbour. The bank is fine sandy covered with a thick
asphalt-like layer mixed with cakes and clumps of a resin-like material, a conversion
product of oil. pH of water : 7-5, salinity : 28-0 ppt.

Site 11 : The edge of the sea water canal carrying effluents from Hindustan
Petroleum Oil Refineries. The bank is muddy and covered with a thick asphalt-
like layer mixed with cakes and clumps of a resin-like material like above, but
covered with a thick white scum. pH of water : 7, salinity : 31-25 ppt.

Site 12 : The edge of the sea coast near the fishing outer harbour. The sedi-
ment is light black fine sand. pH of water 7, salinity : 34-0 ppt.

At these sites the sediment samples along with some water were collected with a
thoroughly washed stainless steel spoon into 250 ml Erlenmayer flasks. Decaying
leaves, or decaying leaves covered with white scum were also collected into separate
flasks. The surface water samples were collected into 500 ml flasks. The flasks
were plugged with cotton.

The pH of surface water samples was noted using pH indicator papers (BDH)
supplied by the Chemicals Division, Glaxo Laboratories (India) Ltd., Bombay.

The salintiy of surface water samples was estimated after centrifuging at
4000 r.p.m. for 15 min by conductivity method using direct reading conductivity
meter-303 (Systronics, Ahrnedabad), and conductivity of different concentrations
of common salt solution as standard.

To detect the occurrence of Beggiatoa, the sediment, decaying leaf and water
samples on arrival at the laboratory, immediately and also after 24 hr incubation
in darkness at room temperature, ca 28 °C, were repeatedly examined micro-
scopically.

To ascertain the distribution of Beggiatoa the samples were subjected to crude
enrichment culture technique described by Faust and Wolfe (1961) with some
modifications. Instead of seasoned roadside winter grass, seasoned rice straw
was used. The procedure to extract and dry rice straw pieces of 2 cm in size
was the same as that of Faust and Wolfe. The enrichment medium for sample
of each site consisted simply of 0-6 g of extracted rice straw bits and 70 ml of
surface water collected at the site. The surface water used is referred to as site
water. The media were taken into 150 ml flasks, and plugged with cotton to
prevent evaporation. The media were not sterilized, because sterilization in the
initial studies proved to be less effective in increasing the population of Beggiatoa
trichomes. The unsterilized media were inoculated with sediment of about 1 ml
in volume, or 2 decaying leaf bits of about 1 cm2 in size, or sediment plus decaying
Jeaf bits. O$e set was left without adding either seciimei^t or decaying leaf bits,

162 M R R Mohan and A Narayana Rao

They were incubated at room temperature, ca 28° C, for 30 days in darkness.

To study their growth, and salt tolerance the same crude enrichment culture
technique was followed with some alterations. The enrichment medium for
Beggiatoa of each site consisted of the same quantity of rice straw bits, and 70 ml
of either respective unfiltered site water or filtered site water, or filtered and stored
sea water (pH 7-0 ; salinity 35-0 ppt), or tap water (pH 6-8 ; salinity 0-31 ppt),
or metal distilled water (pH 6-5 ; salinity 0-016 ppt). The media were taken in
150 ml flasks. These were also not sterilized. They were inoculated with sedi-
ment only, and incubated as above. The site and sea waters were filtered
through Whatman No. 1 filter paper.

The crude enrichment cultures were observed directly, and also examined micro-
scopically. The amount of growth was estimated visually because of the tendency
of the organisms to form aggregations of trichomes, fragmentation of trichomes,
and due to certain practical difficulties in estimating the exact amount of growth
by other methods in vogue

To test sulphur, the trichomes were transferred on to microscope slides, and
treated with aceto-carmine (Ellis 1932).

To identify the species of Beggiatoa, the width of the trichomes, which was
uniform, was considered and measured at x 970 magnification.

To observe the gliding movement on agar surface, the method of Faust and
Wolfe (1961) was followed using their modified Cataldi's agar medium dried at
60° C for about 10 min.

3. Results and discussion

The species differentiation in Beggiatoa is mainly based on the width of trichomes.
According to the usual nomenclature of Beggiatoa forms (Buchanan and Gibbons
1974), the forms with 1# or less wide are called B. minima Winogradsky ; those
between 1 and 2 /j, wide trichomes are called B. leptomitiformis Trevisan ; those
between 2-5 and 5 ju wide as B. alba (Va.uch.er) Trevisan ; those between 5 and
14 fi wide as B. arachnoidea (Agardh) Rabenhorst ; those between 15 and 21 p
wide as B. mirabilis Cohn ; and those between 26 and 55ju wide as B. gigantea
Klas. However, Pringsheim (1964) while discussing the species concept in
Beggiatoa concluded that after dropping for taxonomic purposes the use of the
arrangement of the sulphur droplets etc., one has to di!Op that of the width also,
and there is no feature to replace it. In the present study as there is no other
feature except width for species identification, width alone considered for identi-
fying the species of Beggiatoa, and the usual nomenclature is followed in mentioning
them.

In the present study, all the above mentioned 6 species of Beggiatoa are identi-
fied in the aquatic habitats of Visakhapatnam (figure 1). The trichomes of
B. minima and B. leptomitiformis are less than 1-0 and 1-66/f in width respec-
tively. Of B. alba, two distinct kinds or strains of consistent and uniform width
were observed. The trichomes of one are 3 • 32 jx, and those of the other are 4 • 98 //
wide. The trichomes of B. arachnoidea, & mirabilis and B. gigantea are 9 "96,
17 -Q and 28-22/J wide respectively,

Distribution and growth of Beggiatoa

163

e.arachnol«ka

Figure 1. The Beggiatoa spp. occurring in the aquatic habitats of Visakhapatnam.

Table 1. Distribution of Beggiatoa spp. in the sites.

Site No. B. minima B.lepta- B.alba B.arach-B.mirabilisB.gigantca

mitiformis noidea

(1-0^) (I -66ft) (3 -3 2/*) (4-98/1) (9*9^) (IV-O^) (28-22**)

1.

4- 4:

4*

_

2.

— _

4-

— — — —

3.

- 4-

4- '

_ . _ . _ —

4."

— —

4-

_L

5.

• +

H-

— — — —

6.

— _

4-

—

7.

— —

+

4- _

8.

— _

—

— — _ _

9.

4-

4-

— —. — . _.

10.

- 4-

4-

-4-4-4-

11.

• - 4-

4-

— — — _-.

12.

4.

4-

— M

- indicates presence and — indicates absence of Beggiatoa group.

The distribution of these 6 species in the 12 sites given in table 1 shows that
teggiatoa is distributed in all the sites, except site 8 of the stream carrying
[fluents from Coromandel Fertilizers Factory. In the sediment, the. decaying
iaf and the water samples *6f- this site, which is at a distance of about 2km
'om the factory, except a few minute unicellular bacteria, neither Beggiatoa nor
ay other associated organisms were found even when the samples were subjected
> enrichment culture technique (tables 1, 2, 3). Further, when the sediment was
isturbed for collection there was no odour of H2S, indicating the absence of
ay kind of organisms responsible for H2S production. It appears that the high
cidity of water (pH 5 to 2) <h*e to the efflueitfs released is the main cause for tfye

164

M R R Mohan and A Narayana Rao

Table 2. Growth of Beggiatoa spp. in, crude enrichment culture media containing
Site waters.

Site Beggiatoa Spp.
No.

Culture medium and inoculum

Site Site water Site water Site water
water + + +

Scdiment decaying sediment and

leaf decaying leaf

1. B. minima

B. leptomitiformis
B. alba (1- 12 p)

2. B. alba (!' 12 n)

3. B. leptomitiformis
B. alba (3'32ju.)

4. B. alba (1- 12 p)
B. alba (4- 98,*)

5. B. leptomitiformis
B. alba (1'12^)

6. B. alba (3 -32 ^)

7. B. alba (!' 12 p)

+ + +

+ + +
+ + + +

+ -H- +
4 +

4 + +

+ 4-

8.

9. B. leptomitiformis
B. alba (1- 12 &

10. B. leptomitiformis
B. alba (1'12 n)
B. arachnoidea
B. mirabilis

B. gigantea

11. B. leptomitiformis
B. alba (I* 12 n)

12. B. minima

B. alba (3 -32^)

+ + +

+ + + +
+ + + +

4- +

- =Nil, + = Very poor, ++ =Poor, +++ ==Fair, ++++ = Good.

* No growth, but survived for 7 days after irtcubatior,.

Distribution and growth of Beggiatoa

165

Table 3. Growth of Beggiatoa spp. in enrichment culture media containing
different waters.

Site

Site water
No. Salinity Beggiatoa spp.

Culture media with

Unfiltered Filtered Tap water Distilled Filtered
sits site (salinity water sea water

water water 0-31 ppt (salinity (salinity
pH 6-8) 0-016 ppt 35-0 ppt
pH 6-5) pH 7-0)

1. 0*58 B. minima

B. leptomitiformis
B.

2. 0-72 B.

3. 0-72 B. leptomitiformis
B.

4. 1-07 B.

B.alba (4 -98^)

5. 1 *08 B. leptomitiformis

B. alba (3-32fi)

6. 1-40 B.

7. 4-50 B.

B.

8. 19-0

9. 20-0 B. leptomitiformis
B.

10. 28-0 B. leptomitiformis

B. arachnoidea
B. rnirabilis
B. gigantea

11. 31-25 B. leptomitiformis

B.

12. 34 '0 B. minima
B.

+4-4-

+ 4+4
4-44

444 +
44++

+4++ _

4-4+

++++
++ +

++++
4-4-4-

+4++ _

++ + +
+++ +

+4-4-4-

444
44

+-++ -

4-4-4-4-

+++f-
++++•

++

+++
++

4-4-4-

— = Nil, + = Very poor, 4+ = Poor, 4 + + = Fair, 4444
* No growth, but survived for 7 days after incubation.
@ Not survived for even 1 day after incubation.

Good.

166 M R R Mohan and A Narayana Rao

absence of not only Beggiatoa but also other associated organisms. On the
contrary, the samples of the site 11 of the nearby stream carrying effluents from
Hindustan Petroleum Oil Refineries, when examined immediately or after 24 hr
incubation on arrival at the laboratory, were teeming with trichomes of B. lepto-
mitiformis followed, in number, by those of B. alba (3-32 //), in association with
some protozoans, Oscillatoria filaments and even nematodes. Further, the entire
bank of the stream is covered with a thick asphalt-like layer mixed with cakes
or clumps of resin-like material, a conversion product of oil, which in turn are
covered with a white scum in which the two Beggiatoa spp. are predominant. In
addition, when the scum was disturbed for collection there was a strong odour
of H2 S also. The same is almost the case with site 1 of the narrow and shallow
drain of a septic tank, where B. leptomitiformis trichomes followed, in population,
by those of B. alba (3-32 #) and B. minima are present in association with Thiospira
and some other minute bacteria, protozoans, Oscillatoria filaments, nematodes,
and even insect worms.' At the remaining sites, no such white scum was observed.
However, in the samples of the sitps 4, 5,, 6, 7,, 9 and 10 were found the Beggiatoa
spp. listed against each site in table 1. These are also associated with some
minute bacteria, protozoans, Oscillatoria filaments, diatoms, nematodes and
insect worms. The samples of site 10 contained, in addition, aggregations of
many actively gliding spiral trichomes of Spirulina sp. At these sites also, when
the sediment was disturbed for collection, there was a feeble odour of Ha S. On
the other hand, in the samples of the sites 2, 3 and 12, Beggiatoa spp. alone were
not detected in the initial microscopic examination. However, there was a feeble
odour of H2 S at the sites, when the sediment was disturbed for collection.

The trichomes of these Baggiatoa spp. present in the samples collected were
full of intracellularly deposited sulphur globules, and in active gliding condition.
In samples incubated for 24 hr in darkness, the trichomes present in the sediment,
or on the decaying leaves, or in the white scum aggregated into loose white clumps
on the surface of the glass, and as a white film at the surface of water. It appears
that the trichomes had moved out of the sediment, or decaying leaves, or white
scum, and aggregated on the surface of the glass, and water by gliding over the
surface of glass. However, in samples from sites 2, 3, 8 and 12 no such aggre-
gations were found. But, when the samples of these sites were subjected to
enrichment culture technique, B. minima, B. leptomitiformis and B. alba (3-32/0
trichomes had cropped up in considerable population in the case of samples from
sites 2, 3 and 12 only. This reveals that* even a thorough direct microscopic
examination of samples immediately and after 24 hr incubation on arrival at the
laboratory will not suffice to detect Beggiatoa, if they occur in very small numbers
between the detritus particles. However, ithis is not the case if they are subjected
to a reliable enrichment culture technique, or inoculated into a suitable enrichment
culture medium that is favourable for the small numbers of Beggiatoa trichomes
to multiply into a rich population^ '

The availability of a reliable enrichment culture technique is an important step
in dealing with any problem concerning Beggiatoa, either its natural distribution,
or isolation and cultivation in pure culture media, or taxonomic differentiation, or
nutrition, or tolerance to salinity, etc. Some of the techniques available are the
Butomus-ihizome technique, the technique, with decaying bay and sulphate, and
the sulphur spring technique of Winogradsky (1887) ; the macerated hay ami

Distribution and growth of Beggiatca 167

sulphate technique of Cataldi (1940) ; for isolation of marine strains of Beggiatoa
the hay medium with inorganic salts and artificial sea water technique of Pring-
sheim (1946) ; and the extracted roadside winter grass technique, a modification
of the method of Cataldi, and the weathered Corncob technique of Faust and
Wolfe (1961). Of these, as the extracted roadside winter grass technique of Faust
and Wolfe is reported as a reliable and the most satisfactory method by Pringsheim
(1964), and very simple, it was used to ascertain the occurrence of Beggiatoa spp.
in the sites. However, as the dried roadside winter grass is not available in our
place, dried roadside seasoned grass was used initially with either tap water or
site waters as the enrichment culture medium, and inoculated with sediment, or
decaying leaf bits. The media did not increase the population of Beggiatoa
trichomes. And even when green or dried Ulva bits, or dried dicot leaf bits were
used the result was same. As Beggiatoa occurs in rice rhizosphere (Pitts etal
1972) and water-saturated soils of rice fields (Joshi and Hollis 1977) seasoned rice
straw was next used with site waters as the enrichment culture medium. It has
promoted the growth of Beggiatoa fairly well (table 2).

Clear visual white colonies of the type described by Faust and Wolfe (1961)
appeared on 6, 8, 10 and 15 days after incubation in the media with site waters,
and inoculated with sediment or decaying leaf bits from the sites 1, 4, 6 and 7
respectively (figure 2). These colonies were of various sizes ; the average was
about 1-0 mm in diameter. They appeared both on rice straw bits and on the
surface of the flasks. Further, bundles of trichomes radiated from the centre of
each colony producing a starlike appearance. On the other hand, in the remaining
cultures of the samples from the other sites, instead of such colonies, a thick white
film was formed at the water surface, and on the rice pieces in all the cultures,
except in those of site of the stream carrying effluents from Coromandel Fertilizers
Factory. The microscopic examination of the colonies or the white film revealed
the Beggiatoa spp. listed in table 1. The amount of growth of species of each
site, estimated visually, was also recognizable (table 2). In general, the trichomes
of B. leptomitiformis were more in number than those of others present among
them. Of the remaining, the trend in density of population was first those of
B. alba (3-32/j), followed sequentially by those of B. alba (4-49//), B. minima,
B. arachnoidea, B. gigantea and B. mirabilis, when present in association with each
other.

In cultures of samples from sites 1, 7, 9 and 11, recognizable amount of growth
occurred within 5 to 8 days ; increased fairly well till 20 to 25 days after incu-
bation, and declined thereafter. However, in cultures of site 7, there was total
lysis of trichomes from the 24th day after incubation. On the contrary, in those
of sites 2 to 6 and 12, recognizable amount of growth was evident from 10 to 15
days ; increased fairly well till 21 to 23 days after incubation, and declined there-
after. Total lysis of trichomes, however, occurred after 28 and 25 days after
incubation in the case of cultures of sites 6 and 12 respectively. Whereas, in
cultures of site 10, in which B. alba (3 • 32 /*), B. leptomitiformis, B. arachnoidea,
B. mirabilis and B. gigantea were present the growth of B. alba and B. leptomitiformis
was evident from 14 days, and of B. arachnoidea from 7 days, and continued
to increase till the 30th day after incubation. On the other hand, in B. mirabilis
and B. gigantea even one day after incubation aggregations of trichomes as loose
appeared QIJ the serfage of glass, and in the film forraed at titte surface of

168 M R R Mohan and A Narayana Rao

water. It appears that the trichomes present in the inocula had crept on to glass
surfa.ce and into the film. Further, the trichomes started lysing from the 4th day,
and completely dissolved by the 7th day after incubation revealing that they
failed to multiply in the enrichment cultures.

The microscopic examination of colonies, or loose clumps, or white film placed
on microscope slides, clearly showed the gliding movement of the trichomes of
all the species from the centre of the aggregations or colonies ; and folding of the
trichomes back into the aggregate (figure 3). The dispersed trichomes of all the
species also exhibited active gliding movement. The speed of movement, however,
seems to be different in different species. The trichomes of B. alba, B. leptomitiformis,
ajad B. minima seem to be fast moving, whereas those of B. arachnoidea, B. mirabilis
and B. gigantea slow moving. The repeatedly washed colonies of site 9 cultures
containing a mixed population of B. alba and B. leptomitiformis trichomes on
Cataldi's agar medium solid surface exhibited not only the characteristic circular
flow pattern (figure 4) but also other patterns that resembled rivers. Such patterns
were also reported earlier by Faust and Wolfe (1961) and Pringsheim (1964).

The intracellular deposition of sulphur as droplets was also observed in the
trichomes of all the species of Beggiatoa multiplied in the crude enrichment culture
media. The treatment to trichomes with aceto-carmine resulted in dissolution of
sulphur droplets, and crystallisation of sulphur into flat octohedra outside the
trichomes (figure 5). The arrangement of sulphur droplets in the trichomes of
the different species of Beggiatoa was, however, different. It appears that the
arrangement of sulphur droplets is a characteristic feature of each species irrespective
of its distribution. But in enrichment cultures, sulphur-free trichomes were
also observed. These were in small numbers, and present in declining cultures
only.

It is evident from the above observations made on the samples collected, and
the growth of Beggiatoa spp. in enrichment culture media, that Beggiatoa is
distributed in 11 sites out of the 12 investigated, and favourable conditions for
its existence prevail ^t these sites, except at site 8. The pH of waters of these sites
containing Beggiatoa is not much varied, about 7-0 or above, the optimum for
most aquatic bacteria. The same wa,s also observed earlier by Lackey (1961) in
his studies on occurrence of Beggiatoa relative to pollution. However, with
regard to salt content there are considerable differences between the waters of
different sites, ranging between very, low (0-58 ppt) and very high (34ppt) salt
content. Sites 1 to 5 with waters having 0-58, 0-72, 1-07, 1-07 and 1-08 ppt
salt content respectively are fresh-water habitats. Sites 6 and 7 with waters having
1-4 and 4- 5 ppt salt content respectively are soft brackish water habitats. Sites
9 and 10 with waters having 19 and 20 ppt salt content respectively are hard
brackish water habitats, whereas sites 11 and 12 with waters having 31-25 and
34 ppt salt content respectively are truly marine habitats. As shown in tables
1 and 2, Beggiatoa of equal diameter exist in all these habitats. Further, there
is no difference between them in appearance. If any difference existed, it would be
physiological only, either in nutrition, or growth rate, or indifference to variation
in salt content etc. To study the growth and tolerance to low or high salt content
the sediment samples of all the 12 sites were, therefore, subjected to enrichment
culture technique using the rice straw medium with respective site waters, or tap
water of pH 6-8 ^d 0*31 ppt salinity, or met^l distilled. w^ter of pH 6-5

Distribution and growth of

169

and

3 2 Visual colony growth of Begglatoa spp. of site 7, in erwrichment

the aggregate (x 700), b Edge af the aggregate showing trichomes filled wUh
sulphur droplets also (x 860).

170 M R R Mohan and A Narayana Rao

Figures 4-6. 4. Circular gliding pattern of a mixed population of B. alba and B.
leptomltiformls trichomes on modified Cataldi's agar medium surface (x 860).
5. Octahedral crystals of sulphur outside the trichomes treated with aceto-caramine
(x 1940). 6. Visual colony growth &i Beggiatoa spp. of site 7 in enrichment culture
media containing a Unaltered site water (good growth), b Filtered site water
(good growth), c Tap water (poor growth), d Distilled water (poor growth),
e Sea water (nil growth) ; the white sctrm contained no Beggiatoa.

Distribution and growth of Beggiatoa 171

0-016ppt salinity, or sea water of pH 7 and 35 ppt salinity. The results are
shown in table 3 and figure 6.

In these cultures of samples from site 8 Beggiatoa was not detected, confirming
the above results that it is not occurring at this site. In the remaining cultures
of samples from 1 1 sites, the Beggiatoa spp. listed in tables 1 and 2 were again
observed. In general, by 6, 8 10 and 15 days after incubation clear visual
white colonies described above also appeared in the culture media favourable for
growth of Beggiatoa from the sites 1, 4, 6 and 7 respectively (figure 6). On the
other hand, in the remaining cultures, whichever was favourable for growth,
Beggiatoa trichomes except those of B. mirabilis and B. gigantea increased in
population between 8 and 15 days after incubation and aggregated into loose
clumps in the white film that was formed at the surface of water and glass. The
microscopic examination of colonies or loose clumps showed actively gliding
trichomes filled with sulphur droplets. It appears that enough H2S is produced
in the media due to putrefaction of sulphur containing proteins that were left in
the extracted rice straw pieces, and supported the growth of Beggiatoa. Further,
oxidation of H2S must have taken place, and probably organic nutrients were
also utilized for growth.

Regarding B. mirabilis and B. gigantea, the trichomes as in the previous enrich-
ment cultures aggregated by one day after incubation on rice bits, surface of glass,
and at the surface of medium with site waters or sea water only. They remained
for 3 days in active gliding state ; thereafter started declining, and completely
lysed and dissolved by 7th day after incubation revealing that they failed to grow
in the media. Whereas in tap water and distilled water media the trichomes did
not even appear either on rice bits, or on the surface of glass, or at the surface of
water ; indicating that they lysed within one day after incubation. Further,
when the trichomes of these two species were suspended IB tap water, or distilled
water, or sea water, or their site water for microscopic examination, those suspended
in tap water, or distilled water have bulged and burst releasing colloidal mass of
cytoplasm and sulphur droplets. This confirms the earlier observations by
Lackey (1961), Pringsheim (1964) and others that they are restricted to brackish
or marine habitats and appear to be strictly autotrophic and rather delicate and
exacting. These two largest species, hence, have not yet been grown successfully
either in crude enrichment cultures or in pure cultures by anybody.

The remaining Beggiatoa spp. of each site, as shown in table 3, could grow in
enrichment culture media, but their amount of growth was different in diiferent
media, ranging between good to nil growth. Further, the lag period was also
different. Where the amount of growth was good and fair, recognizable growth
occurred by 5 to 8 days ; increased steadily till 20 to 25 days after incubation
and thereafter declined. On the contrary, where it was poor and very poor recog-
nizable growth appeared by 10 to 15 days ; increased very slowly till 20 to 25
days after incubation ; and afterwards declined. Thus, these differences in the
growth of Beggiatoa spp. in different enrichment culture media suggest that
strains or groups differing in salt tolerance may exist within a species. B. minima
occurs in 2 sites, a. freshwater site 1 and a marine site 12. The nil growth in sea
water medium of that occurring in the fresh water habitat indicates that it has a
low salt tolerance. Whereas the growth, though very poor, in filtered site water,
sea water and tap water media of that occurring in the marine habitat shows that

1.72 M R R Mohan and A Narayana Rao

it is indifferent to variation in salt content, and has a wide salt tolerance. Further,
the nil growth of this marine form in distilled water medium may be due to lack of
enough mineral salts for its growth in the medium.

B. leptomitiformis is distributed in 6 sites ; 3 freshwater sites (1, 3 and 5),
2 brackish water sites (9 and 10), and 1 marine water site (11). The forms occurring
in freshwater habitats failed to grow in sea water medium, indicating that they
have a low salt tolerance ; and seem to be restricted to freshwater habitats only.
On the other hand, of the two occurring in brackish water habitats, the one from
site 9 of a stagnant brackish water pond failed to grow in sea water medium,
whereas the other from site 10 of one of the harbour channel terminals near
sulphur-unloading berth could grow in all the media indicating that the former
has a low salt tolerance and the latter has a wide salt tolerance. On the contrary,
that occurring at site 11 of the sea water canal carrying effluents from the Hindustan
Petroleum Oil Refineries by its good, fair, poor and nil growth in site waters, tap
water, sea water and distilled water media respectively, shows that it is indifferent
to variation in salt content, and can thrive well if enough mineral salts were
available for its growth. Thus, these B. leptomitiformis forms, though similar
IB appearance and morphology, may be different physiologically ; the 3 occurring
in sewage containing freshwater habitats formed one group ; the two occurring
in sewage containing brackish water pond, and oil containing and probably sulphur-
rich harbour channel terminal formed two distinct groups ; and the one occurring
in sea water canal carrying effluents from the oil refineries formed another group.

B. alba of 3 • 32 // wide is distributed in 1 1 sites, suggesting that it is ubiquitous
in distribution. The forms occurring in freshwater habitats, sites 1 1o 5, by their
nil growth in sea water seem to be restricted to fresh water habitat, and have a
low salt tolerance. It appears that physiologically these may belong to one group.
The one from site 6 of a sewage stream that joins the harbour channel receiving
sea water during high tides, by its nil growth in tap water or distilled water, and
poor growth in sea water media seems to be possessing high salt tolerance. Further,
it appears to belong to a separate group. On the other hand, those occurring in
sites 7 and 9 exhibited nil growth in sea water medium only. It appears that
these two could not tolerance high salt content and prefer lower levels, and may
belong to one group physiologically. Whereas that occurring in site 10, a hard
brackish water habitat, by its growth in all the media indicates that it is indifferent
to variation in salt content, and has a wide salt tolerance. Most likely it may
belong to another group. And those occurring in sites 11 and 12 truly of marine
habitats, showed growth in all the media, except in distilled water medium. It
shows that these two are also indifferent to variation in salt content with, however,
a preference for at least some amount of mineral salts for growth, and may belong
to one group. Thus, B. alba forms distributed in the 11 sites, though similar
in appearance and morphology are different in salt tolerance, and may belong
physiologically to 5 different groups.

The B. alba strain with 4-98 p wide trichomes iis distributed in two sites, 4 and 7*
Their nil growth in sea water medium indicates that they have a low salt tolerance.
Further, the poor and nil growth in tap water and distilled water respectively by
the one from site 4, and very poor growth in tap water and distilled water by the
other from site 7 indicate that the two may be different physiologically.

Distribution and growth of Beggiatod 173

' The remaining widest forms of Beggiatoa, viz., B. arachnoidea, B. mirabilis and
B. gigantea are detected in only one site (10). Of these, B. arachnoidea &lo&e could
grow both in tap water and sea water indicating that it is indifferent to variation
in salt content. However, its nil growth in distilled water may be due to lack
of enough mineral salts in the medium. On the contrary, B. mirabilis and
B. gigantea by their nil growth in any of the media seem to require more exacting
conditions for growth in culture ; halophilic, and strictly autotrophic.

4. Conclusion

The present study reveals that Beggiatoa is widespread in distribution ; B. minima,
B. leptomitiformis and B. alba are ubiquitous occurring in both freshwater and
marine habitats ; B. arachnoidea, B. mirabilis and B. gigantea are restricted to
brackish and marine habitats ; and conditions supporting the growth of these are
prevailing in 11 sites, out of the 12 investigated. Further, the enrichment culture
medium containing extracted rice straw bits and the respective site waters proves
to be a reliable and satisfactory medium for increasing the small number of Beggiatoa
trichomes into a rich population, which is a prerequisite for isolation and culti-
vation in pure cultures. Although the species of Beggiatoa identified are considered
as belonging to the six species described in Bergey's manual according to the
usual nomenclature, the occurrence of Beggiatoa of identical diameters both in
freshwater and marine habitats ; and the differences in growth in rice straw
medium with different waters of varying salt content suggest that width is not
the sole feature to be taken into consideration for species differentiation, substan-
tiating the views of Pringsheim (1964) and others. That physiological groups
exist within the smaller forms of Beggiatoa is also confirmed, though based only
on growth in crude enrichment culture media. However, there is every need for
isolating and cultivating these forms in pure culture media for further detailed
consideration of species differentiation, and their nutrition.

Acknowledgements

The award of a fellowship by the Council of Scientific and Industrial Research,
New Delhi, to one of the authors (MRRM) is duly acknowledged.

References

Buchanan R E and Gibbons N E 1974 Sergey's manual of determinative bacteriology (Baltimore :

William and Wilkins) pp. 1416
Cataldi M S 1940 Aislamiento de Beggiatoa alba en cultivo puro ; Rev. Inst. Bacteriol. (D.N.ff.)

9 393-423

Ellis D 1932 Sulfur bacteria (New York : Longmans, Green) p. 21
Faust L and Wolfe R S 1961 Enrichment and cultivation of Beggiatoa alba ; J. Bacteriol. 81

99-106
Jorgensen B B 1977 Distribution of sulfur bacteria (Beggiatoa spp.) in coastal marine sediment ;

Mar. Biol (Berlin) 41 19-28

174 M R R Mohan and A Narayana Rao

Joshi M M and Hollis J P 1977 Interaction of Beggiatoa and rice plant : Detoxification of

hydrogen sulfide in the rice rhizosphere ; Science 195 179-180
Keil F 1912 Beitrage Zur Physiologic der forblosen Schwefelbakterien ; Beitr. Biol. Pflanzen.

11 335-372
Kowallik U and Prmgsheirn E G 1966 The oxidation of hydrogen sulfide by Beggiatoa ;

Am.J. Bot. 53801-806
Lackey J B 1961 Occurrence of Beggiatoa species relative to pollution ; Water and Sewage

Works 729-31
Pitts G, Allarn A I and Hollis J P 1972 Beggiatoa : Occurrence in the rice Rhizosphere ;

Science 178 990

Pringsheim E G 1946 Pure cultures of algae (London : Cambridge University Press)
Pringsheim E G 1964 Heterorrophism and species concepts in Beggiatoa ; Am. J. Bot. 51

898-913
Scotten H L and Stakes J L 1962 Isolation and properties of Beggiatoa ; Arch. Mikrobiol 42

353-368

Winogradsky S 1887 Uber Schwefelbakterien ; Bot. Z. 45 489 tT. (Nos 31-37)
Wlnogradsky S 1888 Beitrage Zur Morphologic Und Physiologic der Bakterien Heft. 1.

Schwefelbakterien. Leipzig : Felix
Winogradsky S 1889 Recherches physiologiques surles sulfobacteries ; Ann. Inst. Pasteur.

349-60

Proc. Indian Acad. Sci. (Plant Sci.), Vol. 91, Number 3, June 1982, pp. 175-181.
© Printed in India.

Photoperiodic control of extension growth, bud dormancy and
flowering of Nerium indicum Mill, and Thevetia peruviana Schum.

KUSHAL SINGH, SURINDER KUMAR and K K NANDA

Department of Botany, Panjab University, Chandigarh 160 014, India

MS received 12 November 1981

Abstract. Plants of N. indicnm and T. peruviana grew taller and produced more leaves
under LD than under ND condition. While T. peruviana plants were taller and had
more leaves under ND than under SD, those of N. indicum did not differ under the two
photoperiods. In both cases LDS delayed the onset of bud dormancy but hastened the
initiation of floral buds. While in T. peruviana floral buds were not formed under SD
condition, in N. indicum floral buds were formed but they did not develop into
flowers. While in N. indicum more flowers were produced under LD than under
ND condition, in T. peruviana the number produced was higher under ND than under LD
condition.

Keywords. Photoperiod; extension growth; bud dormancy; flowering; Nerium indicum;
Thevetia peruviana.

1. Introduction

Studies on the effect of photoperiod on extension growth, bud dormancy and
flowering of woody species have shown that in general while long days prolong the
period of extension growth and therefore delay the onset of dormancy, short days
hasten the onset of rest (Nanda 1963; Whalley and Cockshull 1976; Bhatnagar and
Talwar 1978; Singh and Nanda 1981). The floral response, however, varies with
the plant species. Thus, the young seedlings of Cojfea arabica produce floral buds
under short day condition (Piringer and Borthwick 1955) while in Hydrangea
(Piringer and Stuart 1955) and Mains hupehensis (Zimmeraann 1971) floral buds
are induced under long day conditions. Davidson and Hamner (1957) have reported
that although long days induce floral buds in Rhododendron catawbiense, short days
are needed for their development into flowers. In contrast to this, Mirov (1956)
has reported that photoperiod does not affect the flowering response of 35 exotic
pines. This paper deals with the effect of photoperiod on extension growth, bud
dormancy and flowering of two garden plants, Nerium indicum and Thevetia
peruviana.

176 Kushal Singh, Surinder Kumar and K K Nanda

2. Materials and methods

Plants of Nerlum indicum Mill, were raised by planting one-year old stem cuttings
(15 cm each), while those of Thevetia peruviana Schum. were raised from the seed
collected locally from a healthy tree growing in the Panjab University Campus.
The seed was sown on February 1, 1980 in 3:1 mixture of field soil and sand in
earthenware pots (25 cm dia) under three photoperiodic regimes namely; long day
(LD) - consisting of continuous illumination which was provided by supplementing
the normal day-length by unfiltered 200 watt light from incandescent lamps which
provided light intensity of about 3000 lux at the level of the plants; normal day
ND - consisting of natural day-length conditions ; prevailing at Chandigarh
(figure 1) and short day (SD) - consisting of 8 hr daily light alternating with 16 hr
dark which was provided by covering the plants with thick tarpaulin sheets daily
from 1700 hr to 900 hr which led to the rise in temperature in the range of 3-6°C
throughout the course of experimentation. To ensure healthy growth, Hoagland's
nutrient solution (Hoagjand and Arnon 1939) was supplied to the plants twice a
week during the course of experimentation from April 1980 to June 1981.

£20-

«

o.

01

I—

to-

MAXIMUM TEMPERATURE

MI'MMUM TEMPERATURE

ISi

I*

10

I i I I ! i I

APRJMAYjJUNjjTlTlAUG^

1980* ' ' 1981 '

TIME (MONTH)

Photoperiodic control of extension growth ...

177

Observations of extension growth were recorded at 14-day intervals whereas the
number of leaves was counted daily. Records were also maintained of the dates of
initiation of floral buds. The number of floral buds produced and the time taken
for them to open into flowers were also recorded. The 'period of dormancy' in
this paper refers to the period during which the shoot apex remained apparently
inactive and no new leaves were produced on the plant.
3. Observations
The results are presented diagrammatically in figures 2 and 3 and table 1 .

150i

130-

120-

110-

100-

T
4-

T

o

S! 80-

en
1

:70-

«
50-

40-
30-

20-
10-

N. indicum

i

I I HEIGHT

NUMBER OF LEAVES

10 NO SO

CO NO SO

Figure 2. Effect of photoperiod on height of the main axis and number of leaves
produced on N. indicum and T. peruviana.

178

Kushal Singh, Surinder Kumar and K KNanda

glS23 PERIOD OF FLORAL euo INITIATION
I 1 PERIOD OF ACTIVE GROWTH
E23 PERIOD OF BUD DORMANCY

T. peruviano

SD

ND

^^

LD

»?^^vj

N- indicum

SD

HiMWWW^^

ND

*~vmw^^

LD

o : so: hoo j 150 : 200; ?so

APR jMAYljUNlJUL jAUGjSEPjOCT JNOVJOEC

19*0
TIME

360 ; 3$o i A%o i 4io

JANjFEBjMARiAPRiMAY; JUN

1981

•NUMBER OF DAYS

Figure 3. Effect of photoperiod on periods of active growth and bud dormancy in
N. indicum and T. pemviana.

Table 1. Effect of photoperiod on flowering of N. indicum and T. peruviana.

Photoperiod Days to floral Number of floral Days taken to Number of Number of plants
bud initiation buds flower opening flowers out of 10 that pro-
duced floral buds

N.vindicum

LD

190.5±2.50

70.8±2.81

314.4+2.81

17.5±1.71

10

ND

315.2±3.40

35.6±3.10

356.7±4.31

10.5±3,81

10

SD

340,0±4.21

3.0±1.05

—

—

10

T. peruviana

LD

340.5±3.42

6.1 ±2.40

360.8±4.21

4.5±2.85

10

ND

320.7±5.16

18.7±3.08

348.9±3.91

10.6±1.31

10

SD

—

—

—

—

—

± SE at 95% level of significance

3.1 Extension growth and number of leaves

Figure 2 shows that plants of both the species grew taller and produced more
leaves under LD than under ND and SD conditions. While in N. indicum the height

Photoperiodic control of extension growth .... 179

and number of leaves produced under SD condition did not differ from that
produced under ND condition, in T. peruviana plants remained significantly shorter
and produced fewer leaves under SD than under ND condition.

3,2. Periods of growth and bud dormancy

In N. indicum, plants continued growth till mid November under LD as compared
to that till early October and mid September under ND and SD conditions,
respectively. The period of rest that followed also lasted till early February under
LD but till mid February and early March under ND and SD conditions, respecti-
vely (figure 3).

In T. peruviana, growth continued till early February ui^der LD condition but lasted
till the end of December and October under ND and SD conditions, respectively.
The period of rest that followed and which lasted till mid-February and early
February under ND and LD conditions, respectively, continued till early March
under SD condition.

It may be noted that in both the species the period of rest was shorter under LD
but longer under SD than under ND condition (figure 3).

3.3. Days to floral bud initiation

In TV. indicum, floral bud initiation in plants under LD condition started in early
October prior to the onset of dormant phase. In contrast to this under ND
condition floral buds were initiated in mid February. Floral bud formation under
both these conditions continued till the end of June when the experiment was termi-
nated. In contrast to this, the formation of floral buds under SD condition was
delayed to early June. Floral bud initiation, thus, occurred earlier under LD but
later under SD than under ND condition (figure 3).

In T. peruviana floral bud initiation was observed immediately after the period of
rest in plants exposed to LD or ND conditions so that it occurred earlier under ND
than under LD condition (table 1). Plants maintained under SD condition did not
produce floral buds. It may be noted that the hastening effect of LDs was
markedly more in N. indicum than in T. peruviana.

3 .4. Days to floral bud opening

Although LDs hastened the initiation of floral buds, the initiated buds in N. indicum
took longer to develop into flowers under LD than under ND condition. Floral
buds produced under SD condition in this species did not develop into flowers at
all. In contrast to this in T. peruviana while the initiation of floral buds occurred
earlier under ND condition, their development was hastened under LD condition
(table 1). As stated earlier, plants of this species did not produce floral buds
under SD condition.

180 Kushal Singh, Surinder Kumar andK K Nanda

3.5. Number of floral buds and flowers

Table 1 shows that while in N. indicum the number of floral buds and flowers was
higher under LD than under ND condition, in T. peruviana it was higher under ND
than under LD condition. As stated earlier, in T. peruviana no floral buds were
produced under SD condition and in N. indicum a few floral buds which were
produced did not develop into flowers.

4, Discussion

The early onset of dormant phase under SD condition in plants of both the species
reported in this paper is in accord with the results reported earlier in some other
plants (Nanda 1963; Whalley and Cockshull 1976; Bhatnagar and Talwar 1978;
Singh and Nanda 1981). Hastened onset of rest period in plants under SD
condition may be due to accumulation of some growth inhibitory substance (s)
(Wareing and Saunders 1971) or to a decrease in photosynthates due to reduced
daily light period.

The fact that in N. indicum floral buds are produced even under SD condition,
while in T. peruviana they are not produced under this photoperiod shows that
while the former is quantitative, the latter is qualitative long day in its response to
photoperiod.

But the more interesting point is that in N. indicum although floral buds, are
produced, they fail to develop into flowers showing that the photoperiodic require-
ment for the completion of these two phases i. e.9 (i) induction of floral buds and
(ii) their development into flowers, may not be the same. That the requirement of
these two phases may vary is shown in soybean (Jindal and Nanda 1978) and
Bauhinia acuminata (Singh and Nanda 1981). It cannot, however, be ruled out that
failure of buds of this species to develop into flowers may be due to the limitation
of photosynthates under SD condition. This is particularly in the light of work of
Ramina et al (1979) and Even-Chen and Sachs (1980) who reported that photo-
synthetic availability influences the flowering intensity in Bougainvillea.

Acknowledgement

This work was supported by a grant (HCS/DST/2/76) from the Department of
Science and Technology, Government of India and the Council of Scientific and
Industrial Research, New Delhi.

References

Bhatnagar H P and Talwar K K 1978 Photoperiodic response of growth of Pinus caribaea
seedlings. I. Effect on stem height and diameter and tracheid characters. Indian For.
104 212-226

Davidson H and Hamner C L 1957 Photoperiodic responses of selected woody ornamental
shrubs. Mich. Exp. Stat. Quart. Bull. 40 327-343

Phot aperiodic control of extension growth .... 181

Even-Chen Z and Sachs R M 1980 Photosynthesis as a function of short day induction and

gibberellic acid treatment in Bougainvillea "San Diego Red". Plant Physio! . 65 65-68
Hoagland D R and Arnon D I 1939 The water culture method for growing plants without soil.

Calif. Agric. Exp. Stat. Cir. 347
Jindal R K and Nanda K K 1978 Effect of time of sowing on flowering of some varieties of

soybean (Glycine max) under varying photoperiods; in Physiology of Sexual Reproduction in

Flowering Plants, (eds) C P Malik, N C Bhattacharya, A K Srivastava and Rattan Singh

(New Delhi: Kalyani Publishers) pp. 148-151

Mirov N T 1956 Photoperiod and flowering in pines. For. Sci. 2 328-332
Nanda K K 1963 Studies on growth and development of forest trees. I. Effect of photoperiod

on annual growth cycle of seedlings of some Indian species. Indian J. Plant PhysioL 6 14-33
Piringer A A and Borthwick H A' 1955 Photoperiodic responses of coffee Turrialba 5 72-77
Piringer A A and Stuart N 1955 Responses of Hydrangea to photoperiod. Proc. Am. Soc.

Hortic. Sci. 65 446-454
Ramina A, Hackett W P and Sachs R M 1979 Flowering in Bougainvillea. A function of

assimilate supply and nutrient diversion. Plant PhysioL 64 810-813
Singh K and Nanda K K 1981 Effect of photoperiod on extension growth, rest period and

flowering of Bauhinia acuminata L. seedlings. Part I - Analysis of the first year growth.

Indian J. Exp. Biol. 19 337-340
Wareing P F and Saunders P F 1971 Hormones and dormancy; Ann. Rev. Plant PhysioL 22

261-288
Whalley D N and Cockshull K E 1976 The photoperiodic control of rooting, growth and

dormancy in Cornus alba L.; Sci. Hortic. 5 127-138
Zimmermann R H 1971 Flowering in crabapple seedlings. Methods of shortening the juvenile

phase. /. Am. Soc. Hortic. Sci. 96 404-411

Proc. Indian Acad. Sci. (Plant Sci.), Vol. 91, Number 3, June 1982, pp. 183-188.
© Printed in India

Interaction of kinetin with B group vitamins on the seedling growth
of green gram (Phaseolus radiatus L.)

P GOPALA RAO and J KODANDARAMAIAH

Department of Botany, Sri Venkateswara University, Tirupati 517 502, India

MS received 17 November 1981 ; revised 13 May 1982

Abstract. Kinetin (50 and 100 mg 1— 0 inhibited both the shoot and the root growth.
Inhibition of root growth by kinetin is considered to be mainly due to inhibition
of protein synthesis. Vitamins of the B group v/z., riboflavin, thiamin, niacin and
pantothenic acid are found to be antagonistic to kinetin in reversing the inhibition
of protein synthesis of the root. Vitamins, probably by acting as inducers of protein
synthesis, antagonized the action of kinetin. The response of kinetin to shoot protein
content is different from that of the root.

Keywords. Kinetin ; B-vitamins ; root protein ; green gram.

1. Introduction

Our knowledge of cytokinins and their possible functions in root growth
is extremely limited. That cytokinins inhibit root growth was shown by Gaspar
and Xhaufflaire (1967). But how they inhibit root growth was not categorically
explained. Hussain et al (1980) working on nutsedge with vitamins of B group
and kinetin have noticed that riboflavin and pyridoxin up to 100 mg I""1 promoted
root and shoot growth of the plantlets, whereas kinetin produced short, thick
shoots and inhibited root growth, with increasing concentration.

The present study has been designed to understand the interaction of vitamins
and cytokinins which might throw some light on the growth of root and shoot
systems of intact seedlings. Only inhibitory concentrations (50 and 100 mg I""1 )
of kinetin were used in order to find out the efficacy of vitamins to counteract the
influence of kinetin. Earlier report by Gopala Rao et al (1975) indicated that
riboflavin can effectively reverse the chloramphenicol inhibited growth of green
gram seedlings. It is a well-known fact that chloramphenicol is a potent inhibitor
of protein synthesis. Interaction of auxins and riboflavin in growth reactions in
plants was studied by Artamonov (1974). The present study is intended to
find out the capacity of vitamins to counteract the effect of kinetin in shoot or root
growth inhibition.

2. Materials and Methods

Seeds of green gram (variety G.G 525) were surface sterilized with 0.1% mercuric
chloride for 3 min, washed thoroughly with distilled water and allowed to grow

183

184

P Gopala Rao and J Kodandaramaiah

in petridishes. The seeds were subjected to presowing soaking for 24 hr with
kinetin (50 and 100 mg I-1) and vitamins of the B group viz., riboflavin,
thiamine, niacin and pantothenic acid, each at a concentration of 100 mg I""1
since it was found to be effective in reversing the kinetin inhibited protein
synthesis. Then the seeds were allowed to grow in distilled water in diffuse light
in the laboratory up to 8 days. Growth in length of the seedlings was measured
for shoot and root separately. Ten replications, each replications representing
five seedlings were maintained for growth measurements. The protein content
was estimated separately in the shoot and the root portions using the method
of Lowry et al (1951) at 2 day intervals. Three replications were maintained
for protein estimations.

3. Observations

3.1 Extension growth

With kinetin (50 and 100 mg I-1) treatment both the root and shoot growth were
inhibited (table 1). The results discussed pertain to 100 rng I""1 concentration
alone. On the eighth day, for example, the root growth of control seedlings was
8.0 cm while that of kinetin treated';; seedlings was 35cm with 50 mg I""1 and
1.03 cm with 100 mg I-1 respectively. The shoot growth of control seedlings was
17. 5 cm while that of kinetin treated seedlings was 16.1cm with 50 and 0.8cm
with 100 rng I-1 respectively (table 1).

Table 1. Effect of kinetin on growth (cm) and protein content (mg/g. dry wt.) of the
seedlings.

Treatment Observation

Part of
the
seedling

Age of seedling

2 days

4 days

*6 days

8 days

Root

2.55

6.46

760

8.00

Growth

±0.05

±091

±0.66

±0.65

Shoot

1.30

9.25

9.75

17.50

Control

±0.03

±0.27

±0.81

±0.41

Root

109.5

124.5

117.0

82.5

Protein

±7.6

±7.3

±9.1

±1.8

Shoot

124.5

172.5

127.0

90.0

±1.9

±3.8

±1.8

±2.6

Root

0.55

0.70

1.13

3.45

Growth

±0,03

±0.02

±0.04

±0.25

Shoot

0.63

253

2.63

16.15

Kinetin

±0.03

±0.07

±0.03

±029

(50 mg I-')

Root

153.0

96.5

90.0

70.5

Protein

±1.63

±1.51

±0.87

±0.88

Shoot

208.5

182.0

159.0

102.0

±2.91

±4.15

±1.35

±2.01

Root

0.43

0.6

0.8

1.03

Growth

±0.06

±0.08

±0.09

±0.04

Shoot

0.6

2.53

430

8.03

Kinetin

±0.09

±0.07

±0.46

±0.21

(100 mg l~i)

Root

109.5

60.0

51.0

45.0

Protein

±1.25

±0.62

±0.57

±0.69

Shoot

187.5

185.5

163.5

79.5

±3.56

±2.25

±1.79

±1.27

Seedling growth of green gram (Phaseolusradiatuis L.)

185

3.2 Protein content

The root protein content was reduced with both concentrations of kinetin (50 and
lOOmgl-1). There was only an initial increase with 50 mg I-1 kinetin on the
second day of seedling growth. On the eighth day, for example, the root pro-
tein content of control seedlings was 82.5 mg while that of kinetin treated seedlings
was 70.5 mg with 50 mg l^1 and 45.0 mg with 100 mg I-"1 respectively (table 1).

It is tempting to note that the shoot protein content was enhanced with kinetiu
(100 nig I-1) except on the eighth day (table 1).

3.3 Interaction of kinetin (100 mg 1~~*) with vitamins on protein content

There was about 50% reduction in root protein content with kinetin treatment
when compared to that of control seedlings (from 80 mg to 40 mg) on the eighth
day. Although there was an initial raise with riboflavin treatment up to fourth
day, it was followed by a steep fall during later stages. The interaction of
kinetin with riboflavin enhanced protein content of the root from 40 mg to about
140 mg on the eighth day (250%) or a 3 fold increase. Thiamine, niacin and
pantothenic acid could raise the protein level from 40 mg to 80 mg not exceeding
the control level when they interacted with kinetin (figure 2) .

5240

>s

C.

Q

160

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Ul
H-

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o

o

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80

40

SHOOT

O CONTROL
A RIBOFLAVIN
• KINETIN

RlBOFtAVtN-*-
KINETIN

O
A

•

A

SHOOT

CONTROL
TH/AMINE
KfNETlN
THIAMJNE-f

68
AGE OF THE

024
SEEDUNOS IN OAY$

8

Figure 1A.

186

P Gopala Rao and J Kodandaramaiah

$240

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U

80

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CL

40

SHOOT

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A NIACIN

O KINETIN

A NIACIN 4- KIN&TIN

SHOOT

CONTROL

NJACIN

KINETIN

PANTOTHENIC AGIO +
KINETIN

JL

AGE

68 024

OF THE SEEDLINGS IN DAYS

Figure

la the case of shoot, • riboflavin (figure 1A) increased the protein content from
a value of 124.5 mg to 172.5 mg (38%) on the second day and from "a value of
90 mg to 105 mg (16%) on the eighth day. Thiamine also increased the protein
content of the shoot. Niacin and pantothenic acid (figure IB) could not increase
the protein as effectively as the former two vitamins except on the second day.
As opposed to the response of the root, shoot protein content was not reduced by
kinetin, but instead, increased up to the sixth day at least. The interaction of
riboflavin with Jkinetin caused a significant increase in protein content of the shoot.
Thiamine interaction with kinetin caused an initial increase only, followed by a
gradual reduction. Interaction of pantothenic acid with kinetin was quite similar
to that <5f thiamine. Interaction of niacin with kinetin was almost similar to that
of riboflavin in that it caused an additive effect in increasing the protein content of
the shoot (figure IB). The behaviour of riboflavin and niacin on the one hand
and that of thiamine and pantothenic acid on the other hand were found to be
similar in their interaction with kinetin.

4. Discussion"

Kinetin at 50 and 100 mg I-1 is "inhibitory to both the shoot and the root growth.
One of the main causes of root growth inhibition (table 1) by kinetin is the reduc-
tion in protein content. Since all the B vitamins used in the present study were
able to effectively reverse the inhibition of protein synthesis by kinetin, it might be

Seedling growth of green gram (Phaseolus radfatus L)

187

O
cr*

Ul
I—

z
o
(J

z

UJ
I—

o
a
a

200

160

120

80

40J

o!

160
120
80
40

ROOT

® CONTROL

& KINETIN
A RIBOFLAVIN
- O RIBOFLAVI.M 4 KINETIN

CONTROL

A NIACIN

O WIACIN + KINETIN

ROOT

® CONTROL

4 KINETIN

A THIAMINE

O THIAMINE + KINETIN

• CONTROL
A KINETIN

A PANTOTHENIC ACID
O PANTOTHENIC ACID +
K'lNETIN

2 4 6 8 0 24 6
AGE OF THE SEEDLINGS IN DAYS

Figure 2.

assumed that vitamins increase protein synthesis by operating either at transcription
or translation level. This assumption can be corroborated by the observation made
by Srivastava (1967) that cytokinins can also inhibit RNA synthesis. A consequence
of the inhibition of RNA synthesis is inhibition of protein synthesis. Gopala Rao
et al (1975) earlier reported enhanced protein synthesis by B vitamins In green
gram. It is a well-known fact that chlorarnphenicol is an eflfective inhibitor
of protein synthesis. Gopala Rao et al (1976) observed that riboflavin can effec-
tively reverse the chloramphenicol inhibited growth of green gram. Basing on
these observations it can be assumed that a probable site of action of vitamins of
the B group could be at transcription or translation level.

The present study also reveals that the response of kinetin to root protein might
be different from that of shoot protein since the root protein content was signifi-
cantly reduced and that of the shoot was significantly enhanced. The present
study forms the basis for future work on the role of vitamins at molecular level.

1&8 P Gopala Rao and J Kodandaramaiah

Acknowledgements

The authors are grateful to Prof. V S Rama Das for his encouragement and
suggestions. One of the authors (JK) is highly thankful to the UGC for providing
financial assistance.

References

ArtamonovV 1 1974 Interact ion of auxins and riboflavin on growth reactions of plants Dokl

AkadNauk SSSR Ser Biol 210 978-981, Biol.Abst. 57 68993
Gaspar T and Xhaufflair A 1967 Effect of kinetin on growth, auxin catabolism, peroxidase and

catalasc activities Planta. 72 252-257
Gopala Rao P, Nagi Reddy A and Raja Kumar N 1975 Reversal of chloramphenicol inhibited

growth by riboflavin in green gram Curr. Sci. 44 399-400
Gopala Rao P, Nagi Reddy A and Raja Kumar N 1976 UC incorporation into amino acids in

chloramphenicol inhibited growth and its reversal by riboflavin in green gram

Z. Pflanzenphysiol. 80 279-282
Lowry O H, Rosenbrough N J, FarrAL and Randall R J 1951 Protein measurement with

folin phenol reagent ; Biol Chem. 193 265-275
Srivastava BIS 1967 Effect of kinetin on nucleic acid synthesis in barley leaf segments ;

Biochim. Biophys. Ada 145 166-169

Proc. Indian Acad. Sci, (Plant Sci,)» Vol. 91, Number 3, June 1982, pp. 189-20C).
(8) Printed in. India.

Leaf architecture of Apocynaceae

J S S MOHAN and J A INAMDAR

Department of Biosciences, Sardar Patel University, Vallabli Vidyanagar 388 120,
Gujarat, India.

MS received 16 May 1981 ; revised 12 April 1982

Abstract. Leaf architecture including venation pattern has been studied in 19 genera
and 29 species of the Apocynaceae. The leaves are simple, alternate, opposite or
\vhorled with entire margin and a simple midrib. The major venation pattern
conforms to pinnate camptodromous type with festooned brochidodromous seconda-
ries. The qualitative and quantitative features are charted. The leaf size, areolc
size, number of vein endings entering the areoles and number of vein terminations
entering the areoles vary from species to species even within the same species. The
highest degree of vein order is observed up to 7°. Isolated tracheids, isolated vein
endings, isolated free vein endings and tracheoidal elements are noticed. Bundle
sheath cells ensheaths all category of veins.

Keywords. Anatomy ; leaf architecture ; venation pattern ; Apocynaceae.

1. Introduction

Only sporadic information is available on the venation in some members of the
Apocynaceae (Chandra et al 1969, 1972; Kapoor et al 1969; Sharma et al 1970).
The leaf architecture of the Apocynaceae has not been studied in detail. The
present investigation has been undertaken to give a comprehensive and detailed
account of leaf architecture of the Apocynaceae.

2. Materials and methods

The material of the 19 genera and 29 species of Apocynaceae was collected from
different places in Gujarat, Karnataka (Lalbagh, Bangalore) and Kerala States.
The mature leaves were cleared following the procedure of Rao et al (19 80).
Photomicrographs were taken with a Carl-Zeiss photomicroscope 1 using yellow
filter and OR WO NP 15 film. Leaf size was measured using graph paper. The
areole size, the number of veinlets entering in areole and the number of vein
endings entering in areole were taken in five different fields of different leaves, and
the average value was recorded. Terminologies to describe leaf architecture
were adopted from Hickey (1973), Hickey and Wolfe (1975) and Melville (1976).

189

190 / S S Mohan and J A Inamdar

Table 1. Showing the qualitative features and numerical data on the venation pattern
of some Apocynaceae.

SI. No. Name of the taxa
1 2

Locality
3

Shape

4

Apex
5

Base
6

Margin Texture
7 8

1.

Aganosma caryo-

local

ovate

acute

obtuse

entire charta-

phyllata G. Don.

ceous

2.

Allamanda cathartica

Bulsar,

ovate

acumi-

obtuse

entire coria-

Linn.

Gujarat

nate

ceous

3.

A. nerifolia Hook

Lalbagh

oblong

39

»>

„ chart a-

Bangalore

ceous

4.

A. violacea Garden

,,

ovate

acute

acute

,, coria-

& Field

ceous

5.

Alstonia scholaris

Dangs,

99

99

99

99 99

R. Br.

Gujarat

6.

Alyxia pubescens

99

»

99

obtuse

„ chart a-

Turril

ceous

7.

Carissa carandas

Kerala

9)

,,

acute

„ coria-

Linn.

ceous

8.

C. congesta Wight

Dangs,

,,

obtuse

obtuse

„ charta-

Gujarat

ceous

9.

C. spinarum Linn.

9t

99

»

99

»» 99

10.

Catharanthes major

Kerala

»>

19

cordate

" 99

Linn.

11.

C.pusillus GDon.

Bulsar,

»

acute

obtuse

99 »

Gujarat

12.

C. rosem G. Don.

Local

»

obtuse

9>

,, membra-

naceous

13.

C. variegata

99

J»

99

J9

„ charta-

ceous

14.

Cerbera manghas Linn.

Lalbagh,

oblong

acute

acute

„ coria-

Bangalore

ceous

15.

C. odollum Gaertn &

99

»»

obtuse

99

99 ff

Bunt

16.

Chonemorpha macro-

Lalbagh,

wide

obtuse

obtuse

entire coria-

phylla G. Don.

Bangalore

ovate

ceous

17.

Holanhena antidy-

Dangs,

ovate

acute

obtuse

„ charta-

sent erica G. Don.

Gujarat

ceous

18.

Kopsia fruticosa

Kerala

elliptic

acumi-

acute

» 9>

A. DC.

nate

19.

Nerium indicum

Local

narrow

acute

»»

„ coria-

Mill.

ovate

ceous

20.

Parsonsia spiralis

Kerala

ovate

acute

obtuse

charta-

Vidal

ceous

21.

Plumeria rubra Linn.

Local

ob ovate

obtuse

acute

i, coria-

ceous

22.

Rauwolfia serpentina

Lalbagh,

ovate

acute

obtuse

,t charta-

Benthe & Kurz

Bangalore

ceous

23.

R. tetraphylla Linn.

99

99

99

>9

" 99

24.

Tabernamantana

Local

99

acumi-

divaricata R. Br.

nate

"

25.

Thevetia peruviana

99

needle-

acute

acute

,, coria-

(Pers.) K. Schum.

shaped

ceous

26.

Trachelosp ermum
jasminoides Lem.

Lalbagh,
Bangalore

ovate

»

obtuse

» charta-
ceous

27.

Vallaris solanacea

Dangs,

oblong

acumi-

(Roth.) O. K.

Gujarat

nate

9t •»

28.

Wrtghtia tinctoria

Lalbagh,

ovate

9)

R. Br.

Bangalore

ft ft

29.

W. tomentosa

Dangs,

99

acute

Roem & Schutt

Gujarat

_C

» >»

Leaf architecture of Apocynaceae

191

Predomi- Marginal
nant ultimate
tertiary venation
vein
origin
angle
9 10

Leaf
area
in mm2

11

No. of
2° veins
along
one side
midrib

12

Range of
angle
between
1°&2°
veins

13

No. of No. of vein
areoles/ veinlets ending
mm2 entering termina*
in areole/ t ion/mm1
mm2

14 15 16

RR, RA incomplete

3870

8-12

50°-65°

1

11

21

AR, RO incomplete

3635

15-50

60°-85°

1

8

32

RA, OR,

2830

12- 15

50°-75°

1

8

27

00

RR, OR,

5290

12- 16

40°-65°

1

8

56

RA

absent ,,

3775

35- 38

65°-70°

absent

—

—

RA, RO, Firnbriate

2565

15- 18

45°-60a

1

11

18

OR

RO, AR, incomplete

125

8- 10

25°-40°

2

8

13

RR

AA, AR,

1170

7- 10

70°-85°

1

22

49

RA

AR, RA,

965

8- 10

60°-80°

1

20

37

AO

RR, RO, •

735

8- 10

45°-75°

1

9

13

RA

RO, RA,

815

9- 12

3S°-55*

1

15

22

RR

RR, RO

935

7-10

250.450

—

—

—

RR, RO

760

8 - 10

50°-75°

1

10

9

RR, RA,

4210

15- 18

45°-60°

1

10

25

RO

RA, RO,

4490

16-20

40°~55°

1

9

22

RR

RR, RA incomplete

12875

20-25

70°-85°

2

9

19

AR, RR, „

3180

12- 16

45°~65°

2

6

19

RA

RA, AA, fimbriate

8850

18-22

550.750

3

11

45

OA

RR, RO,

2460

110- 125

70°-85°

6

3

1

RA

RR, OR incomplete

5135

14-16

65°-80°

1

6

31

RO, RR,

12015

30-35

35°-55°

1

9

29

RA

OR, RR,

2615

12-15

45°-70°

1

10

40

AR

JR. A, jxibv i»

765

9-12

65°-75°

1

12

41

RR, AR,

2135

10-14

35°-65°

1

14

42

RO

AR, AO

775

15 -V20

30°-45°

2

14

10

RA, OR,

715

6-9

60°-80°

2

5

26

RR

RO, RR,

2850

12-15

70°-80°

1

9

33

RA

OR, RA,

5365

12- 15

70°-85°

2

8

17

RR

OR, RR,

5530

11-15

60°-80°

1

9

21

RA

192 J S S Mohan and J A Inamdar

3. Observations

3. 1 Morphological description

Leaves simple, alternate, opposite or whorled. Shape ovate, narrow to w;
ovate, obovate, oblong (figure 1 A), elliptic. Margin entire (figures 1 A, B, I
Apex acute obtuse (figure 1 A) or rounded. Base acute, obtuse (figure 1 A)
cordate. Texture chartaceous, membranaceous or coriaceous. The qualitative li
features of the species studied are given in table 1.

3 . 2 Major venation pattern

The venation pattern conforms to pinnate camptodromous type with festoor
brochidodrornous secondaries in which secondaries do not merge at the marj
but upturn and join together in a series of prominent arches forming brochic
dromous secondaries having a set of secondary loops outside the main brochic
dromous and form "festooned brochidodromous" type of Hickey and Wolfe (19'
or multiarcuate wherein secondary veins form a coarcuate infra-marginal v
and breaking up into a series of small arching loops forming a zone between 1
infra- marginal vein and the margin (Melville 1976) in all the species, except
Catharanthus roseus where the venation pattern seems to be eucamptodromo
In eucamptodromous venation pattern, secondaries upturned and gradua
diminishing apically inside the margin, connected to the super adjacent secom
ries by a series of cross veins without forming prominent marginal loc
(Hickey 1973) or simple curvipinnate. Here, secondaries curve gradually towa;
the margin and often form marginal or submarginal veins (Melville 1976). 1
primary vein or midrib is the thickest vein of the leaf and after its departure fr<
petiole it traverses straight or markedly curved. The thickness of the primi
vein gradually decreases towards the apex. The next smaller size class of ve
are the secondary veins (2° veins) whose origin may be on either side of 1
primary vein in an alternate or sub-opposite fashion. The number of 2° veins
either side of the primary vein varies from species to species irrespective of 1
leaf size. The secondary veins do not merge into the margin but turn upwaj
and form arches with super adjacent secondaries with acute, right angle or obti
angle. Composite intersecondary veins are observed in all species. Intramargii
vein is observed in Nerium indicum. Intramarginal vein closely paralleling 1
leaf margins and into which the secondary veins are fused; probably the result
the fusion and straightening of the exmedial brochidodromous secondary ai
segments into what appears to be am independent vein (Hickey 1973) or margit
vein simple and linear which is situated close to the leaf edge and without a
other veins extending beyond it formed by linking the ends of all of the excurr<
veins at the margin (Melville 1976) (figure 1 B).

The tertiary veins arise from the secondaries having no definite patterns of an
of origin. Predominant tertiary vein angles of origin are right angle right an,
(RR), right angle acute (RA), right angle obtuse (RO), acute acute (A A), aci
right angle (AR), obtuse right angle (OR), acute obtuse (AO) or obtuse obtuse (0
in all the species studied except in Alstmia scholaris, where the predoratu*
tertiary vein origin angle is absent. Species-wise details of angles of origin i

Leaf architecture of Apocynaceae

193

Figure 1. (A) direct photograph of cleared leaf showing venation pattern in Carissa
congesta ; (B) infra-marginal vein in Nerium indicum ; (C) incomplete marginal
ultimate venation in Rauwolfia serpentina ; (D) looped marginal ultimate venation
in Allamanda nerifolia ; (E) areolation and loop formation in Trachelospermum
jasminoides ; (F) loop formation and veinlets in Trochelospermum jasminoides ;
(G) free vein ending and bundle sheath in Wrightia tinctoria
A — 2.1 X ; B — 55 X ; C — 149 X ;

D — 115 X ; E — 43 X ; F — 270 x ;

G — 335 X

L — Loop T — tracheid

194 J S S Mohan and J A Inamdar

Figure 2. (A) vessel element at the terminal position of the veinlet in Catharanthus
major ; (B) undifferentiated tracheary element (extension cell) in multiseriate vein
in Catharanthus major ; (C, D, E, F) uniseriate or biseriate tracheids in Rauwolfia
serpentina and Vallaris solanacea ; (G) uniseriate elongated tracheid in Catharanthus
major ; (H, I) tracheoidal elements in Rauwolfia serpentina and JR.. tetraphylla.
A ' — 617 x ; B — 435 X ; C - 402 X ;

D - 335 X ; E — 371 X ; F — 672 X ;

G — 385 X ; H — 471 X ; I — 335 X.

Leaf architecture of Apocynaceae 1 95

given in table 1 . The pattern of the tertiary veins is either random or orthogonal
reticulate (Hickey 1973) or scalariform where inter-coastal areas are bridged at
regular intervals by transverse veins either at right angles or with a regular
orientation and having the appearance of rungs on a ladder; transverse ramified
(Hickey 1973) or dendroid - regularly or irregularly dichotomous veins occupying
an areole and attached to the areolar veins at one point (Melville 1976)
(figure 1 E). But in Alstonia scholaris admedial ramification of tertiary veins
branching into higher orders without rejoining the secondary veins and oriented
towards the leaf axis (Hickey 1973) or pendulous type with branching veins lying
free in an intercostal area or an areolus, attached at their distal ends and appear-
ing to be pendulous from a submarginal vein or costal vein.

§.3 Minor venation pattern

Ihe highest order of veins is identified up to 7°, but in Catharanthusroseus'u.p to
3° or 4°, in some cases up to 5° or 6°. The numerical data on the venation pattern
are charted in table 1 . Marginal ultimate venation is incomplete (Hickey 1973)
3r marginal vein simple and incomplete i.e. marginal vein broken, linking some
of the excurrent veins but leaving others free (Melville 1976) (figure 1 C),
fimbriate (Hickey 1973) or marginal vein simple and arcuate i.e. marginal vein
formed of arching veins linking the ends of the excurrent veins (Melville 1976),
looped (Hickey 1973) or marginal vein simple and irregular (Melville 1976)
(figure 1 D).

The areoles are the smallest areas of the leaf tissue surrounded by the major
veins which taken together form a contiguous field over most of the area of the
leaf. The areoles in most of the cases are either well developed or imperfect.
But in Alstonia scholaris the areolation is lacking. The arrangement of the
areoles is either random or oriented. The shape of the areoles may be quadran-
gular, pentagonal, polygonal or irregular. The size of the areole is not constant,
but varies in different species and even in the same species. Venation characters
show variations in areole size, number of veinlets entering per areole and the
organizations of terminal vein endings in different species.

3.4 Veinlets

The ultimate veins of the leaf are either simple or branched. Simple vein
endings may be linear or curved (figure 1 F) . The branched ones divide dichoto-
mously.once or twice symmetrically or asymmetrically (figure 1 F). The veinlets
may be uniseriate (figure 1 C), biseriate or multiseriate (figure 2 B). They may
be thin and long or thick and short. The veinlet number vary irrespective of
ireole size. The veinlets whether uni-, bi- or multiseriate without terminal
tracheids are known as free vein endings (figure 1 G). Occasionally a vessel
with a single scalariform perforation plate is present at the vein tip alongwith
the tracheids in Catharanthus major (figure 2 A at arrow). In most of the cases
where areoles are devoid of vein endings a loop-like structure is seen
(figure 1 A, E, F) which is formed either due to the union of tracheids, veins or
tracheids and veins. Rarely, in a multiseriate veinlet some of the elements fail
to differentiate into tracheary elements (figure 2 B at arrow).

196 J S S Mohan and J A Inamdar

3.5 Tracheids

The tracheids manifest extraordinary variation in size, shape and nature and
situated at the terminal position of the veinlets. Tracheids may be uniseriate
(figure 2 C, G) or biseriate (figure 2 F). They may be juxtaposed (figure 2 F) or
superimposed having «V, *TJ or club-shape (figures 2 C, D, E, F; 3 F). They
may be isodiametric grouped or elongated.

3.6 Tracheotdal elements

Tracheoidal elements are observed in most of the species. They lie lateral and
parallel to the veins and veinlets (figures 2 H, I; 3 A). The tracheoidal elements
may be isodiametric or elongated and arranged either scattered or in rov/s.
Probably the function of tracheoidal elements is to provide mechanical support
and also aid in retention of water for the leaf.

3.7 Isolated tracheids

Uniseriate or biseriate tracheids that lie free and disjunct in the mesophyll and
those connected with veinlets by extension cells are called isolated tracheids
(figure 3 B, C).

3.8 Isolated vein endings

Vein endings with terminal tracheids either uni- or biseriate lying free and
disjunct in the areole are known as isolated vein ending (figure 3 D, E). These
are common in most of the cases.

3.9 Isolated free vein endings

These are uniseriate or biseriate vein ending without terminal tracheids lying free
and disjunct in the areole. These vein endings are seen in Catharanthus roseus
(figure 3 I).

3.10 Extension cells

These are parenchymatous cells which either adjoin isolated tracheid with a vein
(figure 3 G) or a vein with another vein (figure 3 H). Extension cells have
failed to differentiate either into sieve or tracheary elements.

3.11 Bundle sheath

All the major and higher order veins are ensheathed by parenchymatous bundle
sheath ceils. The thickness of bundle sheath varies from primary to higher order
veins. The shape of the bundle sheath cells may be either round, isodiametric
or rectangular (figure 1 G).

4. Discussion

According to Hickey (1973) and Melville (1976) the Apocynaceae leaves fall
under the pinnate camptodromous with festooned brochidodromous secondaries.

Leaf architecture of Apocynaceae

197

Figure 3. (A) iracheoidal element in Catharanthus major ; (B) isolated tracheid in
Rauwolfiu serpentina ; (C) grouped isolated tracheid in JRaitwolfia tctraphylla ;
(D, E) isolated vein endings in Rauwolfia serpentina and Allamanda neri folia ;
(F) isodiamctric tracheids in Rauwolfia letraphylla ; (G) extension cell between
tracheid and vein in Rauwolfia serpentina ; (H) extension cell between vein and
vein in Catharanthus roseus ; (I) isolated free vein ending in Catharanthus roseus.

A — 252 X

D
G

- 292 X

• 471 X
EC —
IFVe —
It —
IVe ~

B — 289 x ;

E — 303 X ;

H — 435 X :
extension cell
isolated free vein ending
isolated tracheid
isolated vein ending

C — 335 X ;
F — 429 x ;
I — 271 X.

Leaf architecture of Apocynaceae 1 99

rarely pinnate eucamptodromous or curvipinnate venation pattern with multi-
arcuate type, rarely simple curvi-pinnate respectively. Sehgal and Paliwal (1974)
classified the euphorbiaceous leaves as imi-, bi- or triveined on the basis of the
number of strands entering the base of the leaf. Apocynaceae leaves fall under
the univeined category.

Reports on the significant variation in the size, shape and number of vein
endings entering the areole are contradictory (see Nicely 1965; Sehgal and
Paliwal 1974). Sehgal and Paliwal (1974), Singh era/ (1976), Jain (1978) and
Inamdar and Murthy (1978) concluded that there is no direct relationship
between size of an areole and the number of vein endings and vein termination
in different species as well as in the same leaf.

Hickey (1973) classified the marginal ultimate venation into looped, fimbriate
and incomplete. In Nerium indicum the intramarginal vein is formed by secondaries
only and there are no higher order veins beyond it. Therefore, it becomes
extremely difficult to classify the marginal ultimate venation according to Hickey
(1973), however, it can well be classified into marginal and linear type of
Melville (1976).

Tracheids or tracheoidai idioblasts (Foster 1956) also regarded as peculiar
cells (Giibort 1881; Bierhorst and Zamura 1965), storage tracheids ~ speichert-
racheiden (Solereder and Meyer 1930), mechanical cells (Mangin 1882), water
storage cells (Kny and Zinimermann 1885), water cells (Pirwitz 1931). Tucker
(1974) referred to these elements as hybrid cells. According to Oltunji and
Nengim (1980) the occurrence of the tracheoidai elements in many unrelated
plants may be due to convergent adaptation to xerophytic habit. The occurrence
of tracheoidai elements along the lateral side walls of veinlets and rarely on the
4° and 5* veins are noticed in this family. These tracheoidai elements differ
from tracheids in form, arrangement and thickening.

Kasapligil (1951), Foster and Arnott (1960), Herbst (1972) reported the
occurrence of isolated veins in dicotyledonous leaves. The tracheidal elements
tfhich lie free in the areole are designated as "free vein ending'* by Sehgal and
Paliwal (1974). The isolated tracheids, isolated vein endings and free veins are
)bserved in most of the species. Terminologies such as vein endings, free vein
mdings, isolated vein endings and isolated tracheids as defined by Inamdar and
Vlurthy (1981) are used here. Sehgal and Paliwal (1974) termed the parenchy-
natous sheath cells which surrounds the veins, as 'ornamented'. Bundle sheath
s present in all the cases around the veins, vein ending and even tracheids.

Acknowledgements

Ve thank Dr (Sr) Avita for kindly supplying some material. Mr J S S Mohan
hanks the CSIR for awarding a junior research fellowship.

References

Jierhorst D W and Zamura P M 1965 Primary xylem elements and element association of
angiosperms ; Am. J. Bot. 52 567-710

200 / S S Mohan and J A Inamdar

Chandra V, Kapoor S L, Sharma P C and Kapoor L D 1969 Epidermal and venation studies in

Apocynaceae 1 ; Bull. Bot. Surv. India 11 286-289
Chandra V, Mitra R, Kapoor S L and Kapoor L D 1972 Epidermal and venation studies in

Apocynaceae IV ; Bull. Bot. Surv. India 14 76-82
Foster A S 1956 Plant idioblasts, remarkable examples of cell specialisation ; Protoplasma 46

184-193
Foster A S and Arnott H J 1960 Morphology and dichotomous vasculature of the leaf of

Kingdonia uniflora ; Am. J. Bot. 47 684-698
Gilbortt W H 1881 Notes on the histology of picher plants ; /. Quekett Microscopical Club 6

151-164

Herbst D 1972 Ontogeny of foliar venation in Euphorbia forbesii ; Am. J. Bot. 59 843-850
HickeyLJ1973 Classification of the architecture of dicotyledonous leaves ; Am. J. Bot. 60

17-35
HickeyL land Wolfe J A 1975 The bases ofangiosperm phylogeny vegetative morphology;

Ann. Misso. Bot. Gard. 62 538-589

Inamdar J A and Murthy GSR 1978 Leaf architecture in some Solanaceae ; Flora 167 265-272
Inamdar J A and Murthy GSR 1981 Vein endings in some Solanaceae ; Proc. Indian Acad. ScL

(Plant Sci.) 90 53-58

Jain D K 1978 Studies in Bignoniaceae III Leaf architecture ; /. Indian Bot. Soc. 57 369-386
Kapoor S L, Sharma P C, Chandra V and Kapoor L D 1969 Epidermal and venation studies in

Apocynaceae II ; Bull. Bot. Sury. India 11 372-376
Kasapligil B 1951 Morphological and ontogenetic studies of Umbellularia calif arnica Nutt. an

Laurus nobilis L.
Kny L and Zimmermann A 1885 Die Bendentung der spiralzelien von Nepenthes ; Ber. Dtsch.

Bot. Ges. 3 123-128
Mangin L 1882 Sur le development des cellules spiralis ; Ann des Sciences Naturelles Paris

(Botanique) 13 208

Melville R 1976 The terminology of leaf architecture ; Taxon 25 549-561
Nicely K A 1965 A monographic study of calycanthaceae ; Castanea 30 38-81
Olatunji O A andNengimRO 1980 Occurrence and distribution of tracheoidal elements in

the Orchidaceae ; Bot. J. Linn. Soc. 80 357-370
Pirwitz K 1931 Physiologische und anatomische untersuchengen an spechertracheiden und

velamen ; Planta 14 19-76

Rao VS, ShenoyK N and Inamdar J A 1980 Clearing and staining technique for leaf archi-
tectural studies ; Microsc. Acta 83 307-311
Seagal L and Paliwal G S 1974 Studies on the leaf anatomy of Euphorbia. II Venation patterns;

Bot. J. Linn. Soc. 68 173-208
Sharma P C, Chandra V, Kapoor S L and Kapoor L D 1970 Epidermal and venation studies in

Apocynaceae III, Bot. J. Linn. Soc. 12 92-96
Singh V, Jain D K and Meena Sharma 1976 Leaf architecture in Salix; J. Indian Bot. Soc. 55

140-150

Solereder H and Meyer F 1930 Systematische anatomic der Monocotyledonen Stuttgart 6 92-242
Tucker S C 1974 DedirTerentiation of guard cells in Magnoliaceous leaves ; Science 185 445-447

»roc. Indian Acad.Sci. (Plant Sci.), Vol. 91, Number 3t June 1982, pp. 201-209,
*\ Printed in India.

mpact of extension growth and flowering on the cambial
ictivity of Delonix regia Rafin.

A K M GHOUSE and SHAMIMA HASHMI

Department of Botany, Aligarh Muslim University, Aligarh 202 001, India

MS received 3 July 1981; revised 18 May 1982

Abstract. Shoot growth in Delonix regia takes place in three distinct flushes, the
first commencing in mid-March, the second in late May and the third in October.
The cambial reactivation starts in April after the initiation of the first flush of shoot
growth, but the addition of new vascular derivatives does not take place until the
second flush of shoot growth has occurred. Heavy flowering which ensues following
first flush of shoot growth seems to delay cell divisions in cambial initials in April.
Once the formation of cambial derivatives starts, it continues till mid-November
whence the cambium enters the dormant phase. Xylogenesis begins from July and the
formation of phloem in October, while the precursor phloem differentiates in early
April.

Keywords. Tree growth; extension growth; flowering cambial activity; vascular deri-
vatives; Delonix regia.

.. Introduction

Foster as early as 1927-28, considered that bud bursting and new leaf emergence
re highly significant for the initiation of cambial activity. The later works on
everal tropical and temperate trees have supported Coster's contention (Chow-
ihury 1958, 1969; Chowdhury and Tandon 1950; Wareing et al 1964; Waisel and
<ahn 1965; Waisel et al 1966; Fahn et al 1968; Ghouse and Hashmi 1979a). It
5 believed that an intricate relationship exists between bud burst, leaf emergence
nd initiation of cambial activity, the first two preceding the last. The present
eport deals with the relationship between shoot growth, flowering and the cambial
ctivity in Delonix regia, a flowering tree which originated in West Africa and is
presently grown in many parts of India. It has a deciduous habit and diffuse
torous wood. The study has been undertaken at Aligarh which is located at
;7° 53' N lattitude and 78° 4' longitude in the monsoon belt of the great gangetic
>lain of North India.

1. Materials and methods

"hirtysix trees of Delonix regia Rafin. planted 30 years earlier at the University
Uampus of Aligarh were used for the present investigation. To study the phenology

201

202

A K M Ghouse and Shamima Hashmi

of the selected trees, 40 current year branches were tagged with aluminium labe!
in each tree, at the rate of 10 branches facing east — west, north and south side;
Observations were recorded on leaf fall, bud burst, leaf emergence, flowering
fruit setting, fruit persistence and bud burn for 3 consecutive calenda
years commencing from 1974.

The daily data on atmospheric temperature, relative humidity and rainfa
were collected from the local meteorological unit maintained at the Universit
Campus. The monthly average of physical factors was calculated and represents
in figure 1 .

Samples of cambial strips of 2 cm sq, together with the inner bark and som
sapwood were collected at fortnightly intervals for 3 consecutive calen

* Re!. hum,(%
Temp* ( °C)
Rain fall (cm)

JFMAMJ JASO NO

I

J'F'M'A'M'J'J'A'S'O'N'D

ZZ Bud bursting

m Pre- cursor phloem

El Flowering

S Cambial activity

ED Leaf fall

^ Phloem production

S3 Extension growth.

ID Xylem production

Figure 1. Graphs showing the relationship between the atmospheric temperature,
relative humidity, rainfall, phenology and cambial activity in Delonix regta
•during a calendar year.

Extension growth and flowering on the cambial activity 203

dar years starting from 1974, Chisel and hammer were employed to take out the
blocks from the main trunks at chest height (1.5 in from ground level). Three
trees were sampled on each turn and from each tree four blocks were obtained,
one each from east, west — north and south side of the tree. The excised blocks
were fixed on the spot in FAA (formalin-acetic acid and ethanol mixture) and
aspirated after an hour. After fixation, they were softened in an alco-glycerol
1:1 mixture of 50% ethanol and 50% glycerol for 4 weeks. Sections were pre-
pared using a sliding microtome in transverse, tangential and radial longitudinal
planes at a thickness of 10 /"m. The sections were stained with tannic acid and
ferric chloride (Foster 1934) and with lacmoid combination (Cheadle et al 1953)
and mounted in Canada balsam, after dehydrating in ethanol-xylol series.

3. Observations

3.1 Extension growth

Dehnix regia, being a deciduous tree, starts shedding leaves from December and
becomes completely bare by early January and remains so till mid-March (figure 1).
In March- April, the weather becomes a little warm in day and large number of
buds emerge from the naked branches of the previous year in the axils of fallen
leaves. The bud grows rapidly and produces two or three bi-paripinnate leaves
within a fortnight. By mid-April buds of floral axes come up in large quantity
from the axils of new leaves on current year shoots. These buds develop into a
large corymbose inflorescence, each having a large number of conspicuously coloured
showy flowers. The trees bloom heavily in late April and early May and as
a result, the green crown appears as flaming red or orange.

The first flush of extension growth which begins in late March ends by late
April when reproductive growth becomes established. As a result of heavy
flowering, the vegetative growth of the shoots stop completely. In late Mayor
a little earlier, flowering declines and the fruits start setting. It may be noted
that the fruits remain on the tree for about 14-15 months.

The second flush of shoot growth starts in the second half of May and continues
up to August. It occurs at a rapid rate and accounts for the major part of the
year's growth product.

The third flush of shoot growth occurs in October in the same year, although
for a short duration because the low night temperature of November burns down
the apices of the branches. Leaf fall ensues in early December and the events
repeat once more (figure 1).

3 . 2 Cambial activity

The microscopic analysis of the cambial samples, collected at fortnightly intervals,
indicates that cambial reactivation commences in the first half of April soon after
bud burst and leaf emergence in first flush of extension growth. To start with

204 A KM Ghouse and Shamima Hashmi

the protoplasmic contents of the cambial cells stain lighter than before. Concur-
rently, the nuclei also lose their chromaticity. In late May a few cells of the
cambial zone enlarge in radial direction and later enter the active phase by under-
going cell division in the first week of June. The radial growth thus resuming in
early June continues up to the second week of November, stretching over a period
of five and a half months against the eight month extension, growth occurring in
three distinct flushes.

After cell division, the size of cambial zone population swells up (figures 2
C,D) touching its peak in August-September and declining later, as the derivatives
continue to differentiate at a rapid rate. In October the activity slowly declines
and stops by mid-November. The cambium thus enters its dormant phase in
late November when the cells develop dark protoplasmic contents (figures 2 A, B).
The walls of the cambial cells, especially the radial walls become thicker and
develop beaded appearance on account of unthickened primary pit-fields. The
cambium remains in this state till the following spring.

3.3 Formation of xylem and phloem

New xylem differentiates in trunks in July after the break of monsoon, although
the cell divisions start in cambial cells one month earlier. Xylem formation takes
place at a high rate throughout August and September and slows down in October
and later stops by mid-October (figure 1). The phloem production, on the other
hand, initiates in October, when the air temperature becomes somewhat moderate
and lasts for about a month.

Earlier to the initiation of cambial activity, but after the advent of summer in
early April, a few layers of cells differentiate into a narrow strip of new phloem
out of the overwintered mother cells. This precursor phloem measures about 75
to 100 [im in depth in transections.

In a growth year, as viewed in transections, about 800 /im of xylem and about
500 /tm of phloem are produced by D. regia at Aligarh conditions of weather.

4. Discussion

The majority of dicotyledons and gymnosperms show a sharp periodicity of shoot
growth including radial growth. However, in certain exceptional cases growth
may occur throughout the year without break, particularly in the tropical envi-
ronment (Alvim 1964; Fahn and Sarnat 1963; Fahn et al 1968).

The pioneering work of Chowdhury (1958, 1968, 1969) has shown that the
radial growth in certain Indian trees may extend up to 10 months in a calendar
year. Other works also indicate that the tree growth in tropics takes place for a
considerably longer duration than at the temperate regions (Fahn and Sarnat 1963;
Lawton 1972; Rao 1972; Chau and Chiang 1973; Lu and Chiang 1975; Khan and
Ghouse 1978, 1980; Ghouse and Hashmi 1979a). The present study on D. regia
also indicates the same, as far as the extension growth is concerned. However,

Intension growth and flowering on cainhlal activity

205

Figure 2. Photomicrographs showing the active (A and C) and dormant phases
(B and D) of vascular camlium in tangential (A and B) and transverse
(C and D) sections-Af B, C at X 372; D at X 160

Extension growth and flowering on the cambial activity 207

radial growth in D. regia takes place only for a period of five and a half months
in a year and it, therefore, does not resemble other trees which grow in tropics.
This may be due to the flowering habit of this species in which the cell division
in cambial cells is seemingly delayed by heavy flowering till June.

The direct relationship between extension growth and the cambial activity
established by earlier workers like Chowdhury and Tandon (1950), Chowdhury
(1958, 1969) and Ghouse and Hashmi (1979a) is further getting confirmed in the
present study, since it demonstrates that the occurrence of extension growth
acts as a prerequisite factor for the initiation of cambial reactivation in the inves-
tigated species. In Polyalthia longifolia the authors noted that the reactivation
of vascular cambium is invariably preceded by the swelling phenomenon in certain
cambial cells and this incidence is initiated, in turn, by the emergence of leaves
(Ghouse and Hashmi 1979a). The results obtained in the present study also indi-
cate the same for D. regia. In an earlier communication the authors further
brought to light that high temperature and high humidity accelerate the differentia-
tion of xylem and low humidity favours the formation of phloem in P. longifolia
(Ghouse and Hashmi 1978). A careful scrutiny of the data obtained in the
present study reveals that the requirements for differentiation of phloem and xylem
are different and they follow more or less the same trend in D. regia and P. longi-
folia (Ghouse and Hashmi 1978).

The decrease in chromaticity of the~protoplasmic contents, tanniferous substan-
ces and the cell wall characteristics have also been noticed in the past (Derr and
Evert 1967; Tucker and Evert 1969; Ghouse and Hashmi 1979a).

Differentiation of phloem preceding that of xylem has been recorded in a number
of tropical trees including those that grow in India (Lawton 1972; Ghouse and
Hashmi 1978, 1979b). In the present study xylem has been noticed to differentiate
first during a growth year. However, the precursor phloem differentiation
as noted in D. regia is more commonly observed in the Indian trees (Ghouse and
Hashmi 1979b, 1979c, 1980) than reported so far in the temperate trees (Evert
1960, 1963; Derr and Evert 1967; Davis and Evert 1968).

The differentiation of phloem two times in a year first in early April
and next in October, as found in D. regia invites special attention. The pheno-
menon appears to be controlled more by the environmental conditions than by the
internal make up of the species. A cursory look of the weather data provided
in figure 1 indicates that the temperature during the periods of phloem differentia-
tion happens to be almost the same in the present case. It appears that a slight
rise in temperature above this level does not seem to favour phloem differentiation
in this species. A situation of more or less similar nature has been observed in
Polyalthia longifolia by the authors (Ghouse and Hashmi 1978).

Acknowledgements

We are thankful to the Council of Scientific and Industrial Research, New Delhi
for financial assistance in the form of a Senior Fellowship awarded to
Miss Shamima Hashmi.

208 A KM Ghouse and Shamima Hashmi

References

Alvim P T 1964 Tree growth periodicity in tropical climate ; in The formation of wood in fores

trees (ed) M H Zimmerman (New York: Academic Press).
Chau T and Chiang S HT 1973 Seasonal changes of cambial activity in the young branch of

Psidium guajava Linn. ; Taiwania 18 35-41
Cheadle V I, Gifford E M and Esau K 1953 A staining combination for phloem and contiguous

tissues ; Stain Techno I. 28 49-53
Chowdhury K A 1958 Extension and radial growth in tropical perennial plants; in Modern deve

lopments in plant physiology (ed.) P Maheshwari (Delhi : Delhi Univ.)
Chowdhury K A 1968 History of botanical researches in India, Burma and Ceylon X WoOi

Anatomy (Aligarh : Muslim Univ. Press)

Chowdhury K A 1969 Cambial activity in temperate and tropical tree XI Int. Bot. Congr. Seatth
Chowdhury K A and Tandon K. N 1950 Extension and radial growth in trees ; Nature (London

165 732-733
Coster C 1927-28 Zur anatomic and physiologic der zuwachszonen and jahressingbildung in der

tropen ; Ann. Jard.]Bot. Buitenz. 37 49-160
Davis J D and Evert R F 1968 Seasonal development in the secondary phloem in Populu,

tremuloides ; -Bot. Gaz 129 1-8
Dcrr W F and Evert R F 1967 The cambium and seasonal development of the phloem in Robinit

pseudoacacia ; Am. J. Bot. 54 147-153
Evert R F 1960 Phloem structure in Pyrus communis L. and its seasonal changes ; Univ. Call)

Publ. Bot. 32 127-194
Evert R F 1963 The cambium and seasonal development of phloem in Pyrus malus; Am. J. Bot

50 149-159
Fahn A and Sarnat C 1963 Xylem structure and annual rhythm of development in trees an

shrubs of the desert IV Shrubs ; Bull. Res. Counc. Israel 11 198-209
Fahn A, Waisel Y and Benjamin L 1968 Cambial activity in Acacia raddiana SavL; Ann* Bot. 3^

677-686
Foster AS 1934 The use of tannic acid and iron chloride for staining meristernatic tissues

Stain TechnoL 9 91-92
Ghouse A K M and Hashmi S 1978 Seasonal cycle of vascular differentiation in Polyalthh

longifolia (Annonaceae); Beitr. Biol. Pflanzen 54 375-380

Ghouse A KM and Hashmi S 1979a Cambium periodicity in Polyalthia longifolia Benth <S

Hook; Phytomorpholo^y 29 61-67
Gbouse A K M and Hashmi S 1979b Longevity of phloem in Polyalthia longifolia Benth <S

Hook Bull. Torrey Bot. Club 106 182-184
Ghouse A K M and Hashmi S 1979c Longevity of phloem in Delonix regia Rafin ; Proc. Indiai

Acad. Sci. (Plant Sci.) 89 67-72
Ghouse A K M and Hashmi S 1980 Seasonal production of secondary phloem and its longevit]

in Mimusops elengi L. Flora 170 1 75-1 79
Khan M I H and Ghouse A K M 1978 Occurrence of intermittent growth waves in the shoots o

Psidium guajava L. ; in Environmental Physiology and Ecology of Plants (ed) D N Sen (Dehi

Dun : B S M P Singh) 351-355

Khan M I H and Ghouse A K M 1980 Studies on the impact of heavy flowering and fruiting 01
the extension growth ofguava (Psidium guajava L.); Flora 169 453-455

Lawton J R 1972 Seasonal variation in the secondary phloem of some forest trees from Nigeria
II. Structure of the phloem ; New Phytol. 71 335-348

Lu C Y and Chiang S H T 1975 Seasonal activity of the cambium in the young branches o
Liquidamber formosana Hance ; Taiwania 20 32-47

Rao A N 1972 Periodic changes in the cambial activity of Hevea brasiliensis ; /. Indian Bot. Soc
51 13-17

Extension growth and flowering on the cambial activity 209

Fucker C M and Evert R F 1969 Seasonal development of the secondary phloem in Acer negun-

do\ Am. J. Bot. 56 275-284
iVaisel Y and Fahn A 1965 The effects of environment on the wood formation and cambial

activity in Robinia pseudoacacia ; New Phytol. 64 436-442
tVaisel Y, Noah I and Fahn A 1966 Cambial activity in Eucalyptus cameldulensis Dehn. I. The

relation to extension growth in young sapling; La-Yaaran 16 103-108
Wareing P F, Hanney C E A and Digby J 1964 The role of endogenous hormones in cambial

activity and xylem differentiation; in The formation of wood in forest trees (ed)

M H Zimmerman (New York : Academic Press) 323-344

Proc. Indian Acad. Sci. (Plant Sci.), Vol. 91, Number 3, June 1982, pp. 211-226.
© Printed in India.

Pharmacognostic studies on the flower of Mesuaferrea L.

USHA SHOME, SHANTA MEHROTRA and H P SHARMA

Pharmacognosy Laboratory, National Botanical Research Institute, Lucknow 226
India

MS received 15 June 1981

Abstract. Stamens of Mesua ferrea L. constitute the genuine 'Nagkeshan
Ayurveda, a drug considered to be astringent, stomachic and expectorant. The pr
communication deals with detailed pharmacognosy of the drug and includes mo
logical, anatomical as well as certain phytochemical characters of the floral pai
Mesuaferrea. Some of the distinguishing characters are : cortical fibres, num
resin canals and calcium oxalate crystals in the cortex and pith of the pe<
anamocytic, anisocytic or paracytic stomata on sepals and petals ; and
zonocolporate pollen grains with reticulate exine surface. Fluorescence ans
behaviour of the drug with certain chemical reagents and thin layer chromatog
were also carried out.

Keywords. Mesuaferrea ; flower ; pharmacognosy ; cluseaccae.

1. Introduction

Mesua ferrea Linn, belonging to family Cluseaceae is considered the gen
'Nagkeshara' (Chunekar I960; Dymock ef #71885; Mudaliar 1957; Va
1971; Waring 1868). It is known as Nagpushpa champeya in Sanskrit; I
nagkeshara in Hindi ; Nageshwara in Bengali ; Nagchampa in Madhya Prad
Pilu-nagkeshara in Gujrati ; Viluha champkan in Tamil ; Iron wood tree
Cobras Saffron in English.

Stamens of M. ferrea constitute the actual drug (Sharma 1978). It :
mention in all the important ancient Ayurvedic literature (Bhavmishra li
Charaka 1949 ; Sushruta 1952) to be used as an astringent stomachic
expectorant. Flowers are given in bleeding piles, in the form of a paste n
with butter and sugar. These are also applied for relief in burning sensatic
the feet (with old and a hundred times washed 'Ghee') (Chakradatta, See Nadk
1937; Kirtikar and Basu 1933). A syrup of the flower buds (1 in 10) is sa
cure dysentery (Nadkarni 1937). The drug was also adopted by Arab
Unani physicians into their Materia Medica (Anonymous 1970) accordinj
which it has a depressent action on uterine muscles, is digestive, diaphor
antidysenteric and an emmenagogue. It is used as a constituent of aboul
compound Unani formulations (Anonymous 1970).

N.B.R.L Research Publication No. 139 (New Series)

212 Usha Shome, Shanta Mehrotra and H P Sharma

Mesua ferrea L. is a large, evergreen tree with a short trunk, often buttressed
at the base. It is found in the Himalayas from Nepal eastwards ascending to an
altitude of 1500m. Jt is also cultivated in the gardens for its beautiful flowers
and attractive foliage (Anonymous 1962). Detailed pharmacognostic work on
the flower of Mesua has not been carried out so far and hence the preient study.

1.1. Previous Work

Several workers have carried out market surveys of Nagkeshara. Satkopan and
Thomas (1967) surveyed Gujrat markets and found only one sample
from Surat as genuine 'Nagkeshara'. These authors have also tried to identify
other samples being sold as Nagkeshara (Satkopan and Thomas 1967a,b ;
1968). All (1967) surveyed the South Indian Markets. However, he found
none of the samples as Mesua ferrea L.

The presence of an essential oil and two bitter substances has been reported
in the flowers by Boorsma in 1904 (see Chakraborty et al 1959). Subramanyara
el al (1975) have isolated mesuanic acid, a new carboxylic acid, from the
acetone extract of Mesua ferrea stamens. Subramanyam and Subba Rao (1969)
had earlier isolated mammeisin from the seeds of Mesua ferrea L. Bala and
Seshadri (1971) isolated mammeigin and mesuol as the main phenolic components
from two different samples of seed oil. Dutt et al (1940), Chakraborty and Bose
(1960) and Chakraborty and Das (1966) isolated two crystalline antibiotic
principles mesuol and mesuone from the seed kernel oil. Chakraborty et al
(1959) investigated anti-bacterial activity of mesuol and mesuone. None of the
constituents, however, was found to be antifungal. Bhattacharya et al (1979)
have synthesized an antibiotic mesuagin from Mesua ferrea.

2. Material and methods

Fresh material was collected from the Forest Research Institute, Dehra Dun
in the month of May and preserved in form acetic-alcohol. Hand sections
stained with safranin were used for the present study. Phloroglucinol, iodine
and terric chloride were used to test lignin, starch and tannin respectively.
For whole mounts petals were treated with a dilute solution of nitric acid
followed by clearing in chloral hydrate. Physicochemical studies were
performed with the powdered, shade dried material.

3. External characters of the flower

The flowers are fragrant ; cream coloured ; ^ebracteate ; pedicellate ; pedicel
short, axillary or terminal ; solitary or in pairs and 2.5-7.5 cms in diameter
(figure 1). The buds are subglobose.

Sepals 4, large persistent, boat shaped, 2 outer slightly shorter than the
inner ones and depressed at the base. Petals 4, large, cream coloured, spreading,
cuneate, margins undulating, caducous ; stamens indefinite, golden-yellow, united

Pharmacognostic studies of Mesuaferrea L.

213

Figure 1. Mesuaferrea Linn : Two tuigs with flowers x 0.425.

214

Usha Shorn e, Shanta Mehrotra and H P Sharma

Figures 2-12. 2, 3. T. S. pedicel (Diagrammatic) 4. A portion of transverse section through
the pedicel region showing details. 5. Upper epidermis of a sepal showing stomata and
striations. 6. Upper epidermis of a sepal showing elongated narrow cells over the vein.
7. Lower epidermal cells of the sepal showing trichome bases and stomata. 8. Lower
epidermal cells showing trichomes only. 9 and 10. uni and multicellular uniseriate
trichomes. 11. T. S. sepal (Diagrammatic). 12. A portion of the same showing details.
Abbreviations: ASP, airspace; BS, bundle sheath; COL, collenchyma ; COR,
cortex; CR, crystal ; CU, cuticle ; EP, epidermis ; GT, ground
tissue ; HYP, hypodermis ; LEP, lower epidermis ; MXY,
metaxylem ; PH, phloem ; PI, pith ; PXY, protoxylem ; RC,
resin canal ; S, starch grains ; SCL, sclerenchyma, ST, stomata ;
STC, stone cells ; TR, trichome ; TRB, trichome base ; UEP, upper
epidermis ; YB, Vascular bundle ; XY, Xylern.

Pharmacognostic studies of Mesuaferrea L. 215

at the base and forming a fleshy basal sheath, filaments small, anthers oblong ;
ovary superior, bicarpellary, syncarpous, style twice as long as stamens, stigma
capitate, style and stigma persistent in young fruit but are shed away later on.

Fruit conical when young, ovoid to almost round with a prominent bea?
when mature, pericarp hard, warty, 2-valved after dehiscence. Seeds 1-4,
angular, smooth and chestnut brown.

4. Histology

4.1 Pedicel

A transverse section of the pedicel is almost circular in outline. The epidermis
is highly cuticularised and is formed by narrow elongated cells bearing one- to
five- celled, uniseriate hairs. The cortex is 19 or 20 layered and is composed
of rounded, collenchymatous cells. Cells of the outer cortical layers have
characteristic semi-lunar, lignified thickenings on their walls. The inner cortical
cells have fairly large starch grains as eragastic inclusions. As the pedicel
matures sclerenchymatous fibres develop in the form of radially elongated
patches outside the phloem. The phloem is well developed and forms a
concentric cylinder. It consists of sieve tubes and phloem parenchyma. The
xylem is endarch and comprises of vessels, fibres and xylem parenchyma.

The pith is well developed and collenchymatous like the cortex. Resin
canals (which are lined by numerous elongated secretory cells) and Ca-oxalate
crystals of rosette and prismatic types are abundant in both cortex and
pith (figures 2-4) .

4.2 Sepals

In surface view cells of the upper epidermis are larger as compared to those of
the lower one (figures 5-8). Stomata are anisocytic or paracytic (figures 5-7) and
those on the upper surface are sparse and larger in size. The guard cells have
parallel striations radiating out from the stomata (figures 5 and 6).

Cells over the veins are narrow and elongated (figure 6). Uni- to multicellular,
uniseriate trichomes (figures 9 and 10) are present on both the surfaces but are
more abundant on the lower one.

A transverse section of the sepal is semi-lunar in outline. Cells of the
upper epidermis have a thick cuticle on the outside ; are larger, and broader
than long and have lignified walls and sunken stomata (figures 11 and 12). The
epidermis is followed by a well demarcated hypodermis, again with lignified
large rounded cells. In contrast the lower epidermis comprises of small, narrow,
vertically elongated and unlignified cells.

The ground tissue is many layered and has rounded collenchymatous cells.
Up to four layers of these have greatly thickened cells similar to those in the pedicel
cortex. The vascular bundles are scattered in the middle of the ground tissue.
These are usually amphicribral and are surrounded by a bundle sheath formed
of two to three layered sclerenchymatous fibres. Resin canals, lined by a layer
of narrow epithelial cells are present throughout the ground tissue. The

216

Usha Shome, Shanta Mehrotra and H P Sharma

Figures 13-21. 13. Upper epidermal cells of a petal. 14. A portion from figure 13
enlarged to show the details. 15-17. Cells of the lower epidermis of a petal from apex,
middle and base respectively. 18. Epidermal cells of a petal showing stomata arranged at
right angle to the cells. 19 and 20. T. S. of petal from lower and middle portions. (Dia-
grammatic). 21. A portion from figure 19 magnified to show the details.
Abbreviations : CC, cell contents ; CU, cuticle ; GT, ground-tissue ; LEP, lower

epidermis ; PH, phloem ; RC, resin canal ; ST, stomata ; UEP,

upper epidermis ; XY, xylem.

diameter of the canals is variable, being sometimes even larger than the
vascular bundles. Disc shaped Ca-oxalate crystals are also present in
abundance.

Pharmacognostic studies of Mesua ferrea L. 217

4.3 Petals

The cells of the upper epidermis are uniform, laterally elongated, have
anisocytic or paracytic stomata and are larger than those of the lower side
(figures 13 and 14). The lower epidermal cells, on the other hand, vary
considerably in shape and size from base to apex (figures 15-18). Epidermal
cells in the apical region are squarish with wavy anticlinal walls (figure 15).
In the lower region, however, these are narrow elongated and thick walled
with comparatively smaller stomata (figures 16 and 17). The stomata are either
oriented in the same plane or may be arranged at right angles to the epidermal
cells (figures 17 and 18).

The transverse section of a petal near the basal region shows a thick
central portion and narrow curved margins (figure 19). The upper epidermis
is undulating and has a thick cuticle on the outside. Sometimes even the
anticlinal inner walls may also be thickened. A little below the epidermis
one or two layers of cells show conspicously brown tannin containing cells. This
layer is absent at higher levels (figure 20). The lower epidermis is smooth
but cuticularised and is formed of comparatively smaller cells. The ground
tissue is multilayered and is formed of parenchymatous cells with irregular
outlines (figure 21). Resin canals are very few at lower levels but at higher
levels these are quite abundant and are larger than the vascular bundles (figure
20). The vascular bundles are arranged almost in a row, some large and some
small. They are collateral and formed of a few xylem and phloem elements.

4.4 Androecium

The stamens are numerous, have short filaments and somewhat thick, elongated
anther lobes. These are tetra-sporangiate (figures 22 and 23). Some fused stamens
(figures 24 and 25), alongwith their respective vascular supplies, have also been
observed. Occasionally the fusion may involve the anther lobes as well. In such
cases, however, fusion of the anther lobes is not complete (figure 24). Another
stamen in which only connectives are fused in the middle can be seen in a
transverse section (figure 25).

A semi diagrammatic representation of a transverse section of dehisced anther
lobe can be seen in figure 26 and details of the vascular bundle in figure 27.
The cells of the connective are elongated and papillate (figure 28). They are
lignified and are positive to phloroglucinol — cone, hydrochloric acid stain.
The epidermis of the anther lobe is formed of large colourless highly cuticularised
cells followed by a fibrous endothecium (figures 26, 29).

4.5 Pollen grains

Pollen grains are 3-(4-) zono-colporate with average size 35.89 X 48.71 /"m (range
30.77 — 48.71 X 35.89 — 58.97^m) ; shape oblate to suboblate ; exine surface
reticulate ; thickness 1 — 2 /*m, muriduplibaculate, colpus measures 13 X 33 /"m
(figures 30-32).

218 Usha Shome, Shanta Mehrotra and H P Sharma

Table 1. Mesuaferrea Linn. : Behaviour of powder with different chemical reagents.

SI. No. Treatment

Observations

Flower

Stamen

L

Powder -f IN NaOH in methanol

Blackish brown

Brown with

yellow tinge

'2.

Powder -f Picric acid (saturated)

Yellowish

Yellowish

orange

orange

3.

Powder -f Acetic acid

Brown

Orange yellow

4.

Powder -f Cone. HC1

Brown

Brown

5.

Powder + Cone. HNOs

Reddish orange

Reddish orange

6.

Powder -f Iodine (5%)

Blackish brown

Blackish brown

7.

Powder -f- SeliwanoflPs reagent

Yellowish brown

Yellowish brown

8.

Powder + Ferric chloride (5%)

Dark blackish-

Dark brown

brown with

with green

green tinge

tinge

9.

Powder + 40% NaOH + a few drops of

Blackish-

Light brown

10% lead acetate

brown with

with yellowish

yellowish

white ppt.

white ppt.

10.

Powder + Sudan III (alcoholic)

Dark reddish

Reddish orange

orange

11.

Powder + Cone. HNOs 4- ammonia.

Orange with

Orange with

ye] low ppt.

whitish

yellow ppt

4.6 Gynaecium

The ovary is superior, bicarpellary and syncarpous (figures 33 and 34). Epidermal
cells of the ovary are small, hexagonal in surface view and have stomata measuring
21.09 X 11.59 (range 19.00 — 26.60 X 9.50 — 14.25 f^m (figure 35).

In transection the ovary is circular in outline (figures 36-38). The placentae
are swollen and united at the base (figure 36). But at higher levels they separate
(figures 36-39). The ovules arise one from each margin in each of the locales
(figure 36). The placentation is thus axile tending to become parietal at higher
levels. A transverse section passing through the style shows a stylar canal in the
centre and three bundles, a dorsal flanked by two laterals on either side (figure
40). Resin canals are abundant even at this level.

5. Powder study

The powder of the whole flower is greyish brown in colour, pleasant smelling and
slightly astringent in taste. The powder of stamens is golden brown in colour
pleasant smelling and astringent to taste. The powder of whole flower was sieved
through No. 40 mesh, cleared in chloral hydrate and mounted in glycerine. A
microscopic examination revealed the following elements, (i) Pieces of trichomes
(figures 41 A-F). (ii) Pieces of tissues showing stomata (figure 41 G), simple pits
(figure 41 H-I). (iii) abundant pollen grains in different planes (figures 41 J and K).
The behaviour of the powdered drug with different chemical reagents was also
studied and is presented in table 1 .

Pharmac agnostic studies of Mesua ferrea L.

219

Figures 22-41. 22. A stamen showing one antherlobe. 23. T. S. of the antherlobe showing
tetra sporangiate condition (Diagrammatic). 24. A stamen showing two anther lobes
partially fused. 25. T. S. of the fused antherlobe. (Diagrammatic). 26. T. S. of dehisced
amherlobe-(Semi-diaprammatic). 27. Vascular bundle from figure 26 magnified to show
the details. 28. Cells from the connective (surface view). 29, A portion of antherlobe
wall magnified showing cuticularised epidermis and endothecium. 30. A pollen grain
showing wall stratification and ornamentation. 31. A palynogram of pollen. 32. L O
pattern of pollen ornamentation. 33 and 34. Diagrammatic representation of very young
and slightly mature ovaries. 35. Epidermal cells from ovary showing stomata in surface
view. 36-38. Serial T. S. of ovary from base to apex (Diagrammatic) 39. L. S. of ovary
(Diagrammatic). 40. T. S. of style (Diagrammatic). 41. A- K Different tissues from the
drug powder. For details refer to the text.
Abbreviations : AL, antherlobe ; C, connective ; CO, Colpus ; CU, cuticle ;

epidermis ; EX, exine ; END, endothecium,

ovary ; OV, ovule ; OW, ovary wall ; PG,

phloem ; PL, placentum ; RC, resin canal ;

ST, stomata ; STI, stigma ; VB, vascular bundle ; VS, Vascular

supply ; XY, Xylem.

EP,

FI, filament ; O,
pollen grain, PH,
SC, stylar cavity ;

Pharmacognostic studies of Mesua ferrea L.
Table 2. Mesua ferrea L. : Fluorescence analysis

221

SI. No.

Treatment

Fluorescence under UV light

Stamen

Flower

1.

Drug as such

Purplish brown

Blackish brown

2.

Powder + Nitrocellulose in a myl acetate

Olive green

Purplish brown

3.

Powder + IN HC1

Reddish brown

Blackish brown

4.

Powder + IN HC1 -f Nitro-cellulose

Dirty green

Dark brown

in amyl acetate

with purple tinge

5.

Powder + aq. IN NaOH

Dark brown

Dark brown

6.

Powder + aq. IN NaOH + Nitro-

Blcckish brown

Blackish brown

cellulose in amylacetate

7.

Powder + 1 N NaOH in Methanol

Blackish brown

Blackish brown

8.

Powder + IN NaOH in Methanol -f

Greenish brown

Greenish brown

Nitrocellulose in amylacetate

9.

Powder + 50% Nitric acid

Reddish brown

Dark brown

10.

Powder + 50% Sulphuric acid

Greenish brown

Reddish brown

with red tinge

The fluorescence analysis was carried out according to the method described by
Chase and Pratt (1949) and Kokoski et <2/(l958). The observations are recorded
in table 2.

6. Phytochemical studies

Phytochemical characters of the drug were studied separately for the whole flower
and stamens respectively. The determination of ash values, sugar, tannins,
alcohol and water soluble extractives were made from air dried material. The
procedures recommended by Anonymous (1966) were followed for calculating
total ash, acid insoluble ash, alcohol and water soluble extractive percentages,
whereas for calculating tannins, total and reducing sugars Fohlin-Denis reagent
and Shaffer's Somogyi micromethods prescribed by AOAC (Anonymous 1965) were
followed.
(1) Ash Flowers Stamens

(a) Total ash 9.297% 2.370%

(b) Acid insoluble ash 3.730% 0.230%

(2) Tannins

(3) Sugars

(a) Total sugars

(b) Reducing sugars

(c) Non-reducing sugars

(4)
(5)

(6)

Volatile oil

Alcohol soluble extractive
(I.P. Method)

Water soluble extractive
(I.P. Method)

4.4%

1.244%
0.960%
0.350%

0.25%
21.57%

14.86%

3.76%

1.852%
1.160%
0.692%

1.75%
25.31%

1 5.40%

222

Usha Shome, Shanta Mekrotra and HP Sharma

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Pharmacognostic studies of Mesua ferrea L.

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BENZENE EXT

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Figures 42-47. A — Stamen extractive
B — Flower extractive

BR, brown ; BY, bright yellow ; D.BR, dark brown ; GY, greenish yellow ; LBR,
light brown ; LP, light purple ; LRY, light reddish yellow ; LY, light yellow ; P,
purple ; PY, purplish yellow; RO, reddish orange ; Y, yellow ; YB,Yellowish brown.

Beside the above 15 gms of air dried powdered flowers and stamens were
sxtracted separately in a Soxhlet apparatus with hexane, benzene, chloroform,
ilcohol and water successively and percentage of each extractive calculated after
evaporation of respective solvents. These were further screened for steroids,
and triterpenoids (L.B. test - Peach and Tracy 1955) ; flavonoids (Shinoda's test -
Loc.Cit.); alkaloids (Mayer's reagent - Loc.Cit.) ; and tannins (ferric chloride
test-Z0c.CzY.)- The results obtained are presented in table 3.

224

Usha Shome, Shanta Mehrotra and H P Sharma

The hexane and benzene extractives of both the parts show presence of
triterpenoids and resins. Percentage of reducing sugars in alcoholic extractive,
in the case of flower, was found to be quite high.

Water soluble portion of both the parts show presence of tannins and saponins.
Concentration of sapcnins, however, was quite high in the case of flower.

Thin layer chrornatography of the above extractives was also carried out
(figures 42-47). The hexane soluble extractive of stamens shows a larger number
of discrete spots as compared to that of the flower (figure 42). With hexane-ethyl
acetate (72 : 29) as solvent system the hexane extractive separates into spots with
shades of yellow and brown in the case of flower (figure 43b). Those of the
stamens are, however, purple (figure 43a).

The benzene extractives (figure 44) are quite comparable. Again, chloroform
extractive resolved into 4 spots each with toluene-acetone (35 : 15) as the solvent
system. Of these the two upper most and the lowermost on Rf 0.96, 0.82 and
0.35 are comparable. However, an oval yellow patch at Rf 0.43 and a brown spot
at Rf 0.65 in stamen and flower extracts respectively are diiferent. The same
extractive resolved into 5 spots in flower and 6 spots in stamens with solvent
system chloroform : acetone : methanol, 32.5 : 15 : : 2.5 (figure 46). It is interesting
to note that the alcoholic portion of the stamens gave 8 spots while the flower
portion gave only 2 spots (figure 47) (Solvent system : formic acid-ethyl formate-
toluene (1 : 4 : 5), The Rf values of different extractives have been recorded in a
tabular form (table 4).

Table 4. Mesuaferrea Linn : TLC Rf. Values

Extractive

Solvent system

Rf Values

Flower

Stamen

Hexane
soluble
extractive

Hexane : Benzene
(30 : 70)

0.067,0.12,0.19,0.82,
0.93

0.04,0.10,0.17,0.25,
0.64,0.89,0.95

Hexane
soluble
extractive

Hexane : Ethyl-
acetate (72 : 29)

0.76,0.83,0,91

0.79,0.83,0.91,0.99

Benzene
soluble
extractive

Benzene : Chloroform
Acetone
(3:1: 0.05 cc.)

: 0.46,0.63.0.80,0.95

0.50,0.63,0.79,0.99

Chloroform
soluble
extractive

Toluene : Acetone
(35 : 15)

0.35,0.65,0.83,0.96

0.35,0.43,0.81,0.97

Chloroform
soluble
extractive

Chloroform : Acetone
Methanol
(32.5 : 15 : 2.5)

0.28,0.43,0.59,

0.78,0.90

0.28,0.41,0.68,0.76,
0.00,0.93

Alcohol
soluble
extractive

Formic acid :
Ethyl formate :
Toluene
(1:4:5)

0.19,0.36,1.0

0.03.0.06.0.08,
0.23,0.29,0.35
0.4,0.48,0.99

Spraying reagent 2% H2$O4
Underlined Rfs denote spots which

are distinctive.

Pharmacognostic studies of Mesua ferrea L. 225

acknowledgements

"he authors are extremely grateful to Dr T N Khoshoo, Director, National
Jotanical Research Institute, for his continued and keen interest in the progress
>f this work. The late Mr K M Vaid was greatly helpful in procuring plant
naterial for us for which we are sincerely thankful to him. We are also thankful
o Dr Mithilesh Chaturvedi for her helpful suggestions regarding the pollen
tudy. Our thanks are also extended to Messrs A Jha, R S Ojha and M K Tandon
or technical assistance.

References

Vnonymous 1962 The wealth of India. A dictionary of Indian Raw Materials and Industrial Product

L-M (New Delhi : CSIR)
Anonymous 1965 Official methods of analysis of the AOAC (Benjamin Franklin Stn : Washington

DC 20044)

Vnonymous 1966 Indian pharmacopoeia 2nd ed (Delhi : Government of India)
Vnonymous 1970 Hamdard pharmacopoeia of Eastern Medicine (Karachi: The Times)
Ui U S 1967 Nagkeshara as known in South India : J. Res. Indian Med. 2 57-65
3ala K R and Seshadri T R 1971 Isolation and Synthesis of some coumarin components of Mesua

ferrea seed oil. Phy to chemistry 10 1131-1134
Bhattacharya P, Chatterjee D, Chakrabarti A and Chakrabarti D P 1979 Synthesis of mesuagin a

plant antibiotic from Mesua ferrea Linn. Indian J. Chem. B17 11 1-112
ihavmisra 1949 Sri Bhavprakash (Varanasi : The Chowkhamba)
rhakraborty O P, Purkayastha M and Bose P K 1959 On the antibiotic properties of some

constituents of Mesua ferrea L. Proc. NatL Inst. Sci. India B25 8-11
3hakraborty D P and Bose P K 1960 On the constitution of raesuol, the bitter antibiotic principle

of M. ferrea L. Proc. NatL Inst. Sci. India Pt. A26 1-11
^hakraborty D P and Das B C 1966 The structure of mesuol (an antibiotic from Mesua ferrea

L.) Tetrahedron Lett. 46 5727-5730

Hharaka 1949 The Charaka Samhita (Jamnagar : Gulab Kunverba Ayurvedic Society)
3iase C R and Pratt R J 1949 Fluorescence of powdered vegetable drugs with particular reference

to development of a system of identification ; /. Am. Pharrn. Assoc. 38 324-331
rhunekar K C 1960 Commentary on Bhavprakash Nighantu of Shri Bhavmishra (Varanasi : The

Chowkhamba)
Dutt P, Deb N C and Bose P K 1940 A preliminary note on mesuol, the bitter principle of Mesua

ferrea ; /. Indian Chem. Soc. 17 277-279
>ymock W, Warden C J H and Hooper D 1885 (reprinted in 1972) Pharmacographia indica vol.

History of the principle drugs of vegetable origin met within British India. (Delhi: Vivek

Vihar D-42)
Urlikar K P and Basil B D 1933 Indian Medicinal Plants 2nd ed (Allahabad : L N Basu and

Co,)
Cokoski J, Kokoski R and Slama F J 1958 Fluorescence of powdered vegetable drugs under ultra

violet radiation /. Am. Pharm. Assoc. 47 715
""Mudaliar Murugesa C S 1957 Gunapadam-vegetable, 1st ed. Section (in Tamil) (Government

Press) 318-320

^adkarni K M 1937 Indian Materia M edica (Bombay : Karnataka Printing Press) Vol. 7
Peach K and Tracy M V 1955 Modern methods of plant analysis (Heidelberg : Springer Verlag)

3rd and 4th Vol
Satkopan S and Thomas P J 1967a The identity of Ayurvedic Market Drugs. I-A. Nagkeshara

Mesua ferrea L. Nagarjun 10 461-467
Satkopan S and Thomas PJ 1967 The identity of Ayurvedic Market Drugs 1-B Nagkeshara

(Ochrocarpus longifolius} B and H, Nagarjun 11 67-72

226 Usha Shome, Shanta Mehrotraand H P Sharma

SatkopanS and Thomas P J 1968 The identity of Ayurvcdic Market Drugs Pt|l-C, Nagkeshara

(Cinnamomum sps. Nagarjun) 474-477

Sharma P V 1978 Dravyaguna vigyana 4th ed. (Varanasi : The Chowkhamba)
Subrarnanyam R M and Subba Rao N V 1969 Isolation of mammeisin from the seeds of Mesua

ferrea L, Indian J. Chem. Soc. 1 1278-1280
Subramanyam R M, Srimannarayana G and Subba Rao N V 1975 Structure of mesuanic acid ;

Indian J. Chem. 12 884-886

Sushruta 1952 Sushmta Samhita (Kanpur : Sri Saraswati Pustakalaya)

Vaidya B 1971 Some controversial Drugs of Indian Medicine II ; /. Res. IndiarftMed. 6 95-104
Waring E J 1868 Pharmacopoeia of India 1st ed. (London : W H Allen and Co.)

Proc. Indian Acad. Sci. (Plant Sci.), Vol. 91, Number 3, June 1982, pp. 227-234.
© Printed in India

Effect of COz in ovecroming self-incompatibility barriers in Brassica
carnpestris L. van toria

A S DHALIWAL* and C P MALIK

Department of Botany, Punjab Agricultural University, Ludhiana 141004, India
*Seed Technology Section, Department of Plant Breeding, Punjab Agricultural
University, Ludhiana 141 004, India

MS received 1 May 1981 ; revised 14 May 1982

Abstract. CO2 mediated induction of pollen adhesion, germination, tube pene-
tration, tube growth through the stylar tissue and seed set was studied in Brassica
campestris L. var. toria following incompatible pollination. This report stresses the
effect of COa not at a single step but at various levels which normally prevent and/or
hinder successful fertilization and seed set. The possible role of CO2 in overcoming
these barriers is discussed.

Keywords. Brassica campestris L. var. toria; incompatibility; COa; seed set.

1. Introduction

The incompatibility reaction in Brassica is sporophytically controlled with multiple
S-alleles (Bateman 1955 ; Thompson 1957). Double cross hybrids raised from
self-incompatible inbred lines would give low yielding crop because of high level of
cross-incompatibility with each other. In recent years, two developments have
taken place to overcome such problems. One is based on the manipulation of
genetic attributes (Thompson 1 978) and the other involves putting the self-incom-
patible plants in an atmosphere of 3 to 5% CO2 after selfing the flowers (Nakanishi
et at 1969, 1975). The latter technique is now successfully employed in obtaining
self-seeds.

Following incompatible pollination various check points exist at the level of
pollen germination, adhesion, germination, tube penetration, tube growth through
the stylar tissue and ultimately seed set. The present study describes the effect of
CC>2 on the aforementioned levels following illegitimate pollination.

2. Material and methods

Seeds of Brassica campestris L. var. toria were obtained from the Department of
Plant Breeding, Punjab Agricultural University, Ludhiana and the plants were

227

228 A S Dhaliwal and C P Malik

raised in the Botanical Garden. Experiments were performed in full flowering
season to avoid early or late season effect.

Excised pistil technique was employed to study the effect of CO2 on pollen
adhesion, germination, pollen tube penetration and growth through the stylar
tissue. Flower buds were emasculated in the evening before opening. Next
morning the flowers were removed and the pedicels with pistils were embedded in
1% agar medium in small glass vials. Self- and cross-pollinations were done with
a fine brush to obtain a thin and even coating of pollen on the stigmatic surface.
The vials containing self- and cross-pollinated pistils were put in a glass container
which was sealed with parafilm. CO2 was injected with the help of an air tight
syringe to obtain 4% concentration. After incubation for 6 hr at 22±2°C pollen
adhesion, germination and tube penetration into stigmatic papillae were studied by
staining pistils with cotton blue (0.1%) in lactophenol.

Growth of pollen tubes in cross-pollinated pistils without CO2 and self-pollinated
pistils with 4% CO2 was followed at times ranging between 6 to 30 hr after polli-
nation. The number of pollen tubes in 0.5 mm region alongwith the length of the
pistil were counted in the two situations (three replicates of six flowers each).

The emasculated flowers were pollinated with self- and cross-pollen and enclosed
within known volume of polythene chamber and 4% CO2 was injected with an air
tight gas syringe. After exposing the flowers for 6 hr the polythene chamber was
replaced by emasculating bags. The number of seeds formed per siliqua was deter-
mined 50-60 days after pollination.

3. Results

Due to full operation of self-incompatibility, adhesion of self pollen to the stigmas
was low. Those which made contact with the papillae showed poor germination.
Figure l(a, b, c) depicts the adhesion of pollen grains in three situations. Figure
la shows incompatible pollination with very few pollen grains adhering to the
stigmatic surface and without any germination. Figure Ic is that of compatible
pollination where most of the pollen grains are adhering to the papillae. Pollen
grains show germination and pollen tubes grow in the stigmatic and stylar tissues.
The firm attachment of pollen grains, following self-pollination, was observed when
they were enclosed in an atmosphere of CO2 (4%) (Figure Ib).

The data given in table 1 show that no pollen tube was recorded in the pistil
following self-pollination. In majority of the styles, the longest pollen tube
reached almost the same position in response to both self-pollination + COa
and cross-pollination styles. The data on percentage of pollen tubes present at
different levels of the pistil after 24 hrs of pollination are set in figure 2. A
gradual reduction in the number of pollen tubes from the top of the pistil down
into the ovary was noticed. A decrease in cross pollen tubes as well as self
pollen tubes (in the presence of CO2) is evident. This decrease was more
pronounced in the cross pollen tubes as compared with the self pollen tubes with
CO2.

Table 2 shows the number of seeds produced per siliqua after different pollination
treatments. The number of seeds per siliqua following bud pollination was nearly

Self -in compatibility barriers in Brassica campestris L.

229

Figure 1. Pollen adhesion, germination and tube penetration after 6 hr pollination. The
stigmas alongwith styles were stained with 0.1% cotton blue in lactophenol. (a) Noticed
very few pollen grains adhering to the stigmadc surface following self-pollination
(b) The number of pollen grains sticking to the stigmatic surface is more as compared
with figure (a) following CO2 treatment. Some pollen tubes are penetrating the
stigmatic papillae, (c) Most of the pollen grains germinate and penetrate following
cross-pollination (pollen tubes arrowed).

Self -incompatibility barriers in Brassica campestris L.

231

Table 1. Growth of pollen tubes in the pistil at different times after pollination in cross-,
self- and self 4- CO2 pollinations. Data are the mean of six flowers.

Time after Pollination
pollination treatment
(hr)

Maximum
pollen tube
length (mm)

Rate of pollen tube growth
mm/br

Max. rate

Mean rate

6 CP

0.66

0.11

0.05

SP +

C02

0.45

0.08

0.04

SP

0.03

0.01

0.00

11 CP

1 .40

0.13

0.07

SP 4-

C0a

1.35

0.12

005

18 CP

3.00

0.17

0.09

SP -4

C02

2.70

0.15

0.05

24 CP

3.50

0.15

0.08

SP-h

CO*

3.00

0.13

0.07

30 CP

4.00

013

0.07

SP +

CO*

3.60

0.12

0.05

Table 2. Effect

(4%) on seed production following self- and cross-pollination.

Pollination
treatment

Self seeds

Cross seeds

Number of
fruits

Number of
seeds per
fruit

Number of
fruits

Number of
seeds per
fruit

Bud
Open flower
— CO2
-hCOa

36

0

34

17.3

0.0
8.2

30

38
30

18.2

19.6
18.8

same in the two situations i. e, self- and cross-pollinations. No seed set was
recorded when the freshly opened flowers were self-pollinated. However, about 8
seeds per siliqua were formed when self-pollinated flowers were kept in 4% CO2
atmosphere for about 6 hr. CO2 did not affect seed set after cross-pollination.

4. Discussion

The strong self-incompatibility barrier exhibited by Brassica campestris var. toria
depends on the recognition and the rejection reaction. The events grouped in these
two stages constitute the various check points as mentioned earlier. The rejection
response is characterized by callose deposition.

The adhesion of the pollen grain to thestigmatic surface seems to be the initial
but most important event to ensure pollen germination and tube penetration. CC>2
plays an important role in the adhesion of the pollen grains to the self stigma
surface as is evident from the presence of more pollen grains on the self stigma
following CO2 treatment than control (without COa) even 6 hr after pollination.

232

A S Dhaliwal and C P Malik

JOO
in

•

A

5 80

c

I 60

.•Cross

0 0.5 i 2 3

Oistanc* from top of stigma(mm)

Figure 2. Percentage of pollen tubes at different levels from top of the stigma following
24 hr of pollination.

If pollination is compatible the tryphine is induced to 'gel' and promotes both
adhesion and hydration of the pollen grain (Dickinson et al 1980). The number
of pollen grains adhering to the stigma surface following cross-pollination is higher
than self-pollination. Stead et al (1980) proposed that differences in the degree/
extent of adhesion may result from physical or chemical alteration(s) taking place
in the components that bind the pollen to the stigma surface. They suggested the
involvement of stigma surface proteins in the pollen grain adhesion which have a
rapid turnover rate. CO2 has been shown to affect changes in the kinetic
behaviour of allosteric proteins (Mitz 1979). CO2 may be affecting pollen adhesion
through the above mentioned processes. CC>2 has been reported to affect protein
synthesis in maize roots (Splittstoesser 1966), Chlorella (Miyachi and Hogestu
1970) and germinating Amaryllis vittata pollen grains (Sharma et al 1981).

Dhaliwal and Malik (1980) and Dhaliwal et ah (1981) have suggested the role
of COa in pollen hydration and recognition rejection response which are essential
for the activation of enzymes and acceptance of pollen grain, respectively. Non-
specific esterases have been suggested to be involved in the active cutinase complex
essential for the breakdown of cuticle. C02 stimulates the leaching of these
esterases on to the stigma surface following illegitimate pollination (Dhaliwal and
Malik 1982).

COs increased both pollen germination and tube penetration following incom-
patible pollination. CO2 action was correlated with the metabolic changes of
pollen tubes and/or papilla cells at the time of their attachment (Nakanishi et al
1969). The growth rates of pollen tubes in self-pollination + COs and cross-
pollination styles indicate that the fastest self pollen tube (in presence of COs) grew
nearly as fast as the fastest cross pollen tube. The gradual reduction in the number
of pollen tubes from the top of the pistil down into the ovary indicate much
variation in the rate of pollen tube growth. Thus, the most rapidly growing tubes
registered two to three times faster rate than the average tubes. This decrease in
the number is more pronounced in the cross pollen tubes as compared to self pollen
tubes in the presence of CO2. The variation in the growth rate may be attributed
to competition between the pollen tubes for the nutrients. Clearly, single pollen

Self-incompatibility barriers in Brassica campestris L, 233

Pollen lands on the stigma

Adhesion

Hydratjon

Germination

Cutinase activation

Ptrwtration •*-•*— I

Orowth through
the stylar

•

Figure 3. Effect of COs (4%) on various check points that lead to self-incompatibility in
Brassica campestris L.

tube growing through the style will have less competition with each other than
several of them growing at the top of the stigma. The seed set increased when
the freshly opened flowers were self-pollinated and were kept in 4% COg atmos-
phere for about 6 hrs. CO2 did not affect seed set after cross pollination.

Talcing the present discussion in conjunction with our previous inferences on the
role of CO2 on various check points is indicated (figure 3). We believe that CO2
act at several stages promoting successful fertilization and seed set in incompatible
pollination.

Acknowledgements

Financial assistance in the form of Junior Research Fellowship to ASD from
Government of India, Department of Atomic Energy is gratefully acknowledged.

References

Bateman A J 1955 Self-incompatibility systems in angiosperms. III. Heredity 9 53-58

Dhaliwal A S and Malik C P 1980 Effect of relative humidity and COa on the shape, volume and

fresh weight of Brassica campestris L. pollen in vitro ; Indian J. Exp. Biol. 18 1522-1523
Dhaliwal A S and Malik C P 1982 Release of esterases from incompatible Brassica

campestris L. pollen in vitro and in vivo ; Indian J. Exp. Biol. 20 95-96
Dhaliwal A S, Malik C P and Singh M B 1981 Overcoming incompatibility in Brassica campestris

L. by carbon dioxide and dark fixation of the gas by self- and cross-pollinated pistils; Ann. Bot.

48 227-233
Dickinson H G, Roberts I N and Stead A D 1980 The role of exine held substances in the

development and behaviour of the pollen tube in Brassica : 5th International Palyrrological

Conference, Cambridge, Abs.
Mitz M A 1979 COa biodynamics: a new concept of cellular control; J. Theor. Biol. 80 537-551

234 A S Dhaliwal and C P Malik

Miyachi S and Hogestu O 1970 Effect of pre-illumination with light of different wavelengths or>

subsequent CO2 fixation in Chlorella cells ; Can. J. JBot. 48 1203-1207
Nakanishi T, Esashi Y and Hinata K 1969 Control of self- incompatibility by CO2 gas in

Brassica: Plant Cell Physiol. 10925-927
Nakanishi T and Hinata K. 1975 Self seed production by CO-2 gas treatment in self-incompatible

cabbage; Euphytlca 24117-120
Sharma S, Singh M B and Malik C P 1981 Dark CO2 fixation during germination of Amaryllis

vittata pollen in suspension cultures ; Indian. /. Exp. Biol 19 710-714
Splittstoesser W E 1966 Dark COa fixation and its role in the growth of plant tissue; Plant PhysioL

41 755-759
Stead A D, Roberts I N and Dickinson H G 1980 Pollen-pistil interaction in Brassica oleraceai

Events prior to pollen germination; Planta 146 21 1-216
Thompson K. F 1957 Self-incompatibility in narrow-stem kale— Brassica oleracea var acephala L

Demonstration of a sporophytic system; /. Genet. 55 45-60
Thompson K 1978 Application of recessive self-incompatibility to production of hybrid

rapeseed ; 5th International Rapeseed conference. Abstract 12.6-16.6 1978 Malmo, Sweden*

oc. Indian Acad. Sci., (Plant ScL), Vol. 91, Number 3, June 1982, pp. 235-240.
Printed in India

harmacognosy of the stems of Portulaca quadrifida L. and
ortulaca oleracea L.

J LAL* and A M KHAN

Pharmacognosy Research Unit, Central Council for Research in Unani Medicine >
Department of Botany, Aligarh Muslim University, Aligarh 202 001, India
*Dmg Standardization Research Circuit, Central Council for Research in Unani
Medicine, National Botanical Research Institute, Lucknow 226 001, India

MS received 3 December 1980 ; revised 5 September 1981

Abstract. Pharmaco gnostic details of the stems of P. quadrifida L, and P. oleracea L ,
are reported to distinguish one from the other.

Keywords. Pharmacognosy; Portulaca quadrifida L.; comparison with P. oleracea JL.

Introduction

wtulaca quadrifida and Portulaca oleracea, commonly known as Chhota Luniya
d Bara Luniya respectively, are succulent annual herbs and grow abundantly in
Id state throughout warmer regions of India (Anon 1969). It has been suggested that
)m the therapeutic point of view they are quite similar and one can be used as
mbstitute for the other by the drug dealers (Dymock et al 1980 ; Kirtikar and
LSU 1975). Detailed chemical analysis of leaf and stem of P. oleracea was
>rked out by Sadaaa and Ahmed (1947). Recently Lai (1980) described the
.armacogaostic features of the leaf of P. quadrifida. The pharmacognostic
tails of the stems of both species are presented in this paper.

Materials and methods

esh plants of P. quadrifida and P. oleracea collected from the Botanical Garden
Aligarh Muslim University were fixed in FAA. After usual processing, free
nd and microtome sections were cut and stained in safranin and fast green.
uorescerice analysis and extractive and ash values of the powdered mass of stems
:re carried out by Chase and Pratt (1949) and Indian Pharmacopoeia (Anon. 1966)
sthods respectively. • .

236

/ Lai and A M Khan

2.1 Macroscopic characters

The stem of P. quadrifida is succulent, diffuse, filiform, purple in colour at
maturity, less than a milimeter in diameter; on crushing mucilaginous; mucilage
slimy; rooting at the nodes; nodal appendages many, pilose, white; internodes
1.5 to 3 cm long; without any smell and taste acidic. The stem of P. oleracea
(figure 8) on the other hand, is about 2 mm in diameter; internodes 1.5 to 3.5 cm
long; nodal appendages less in number, minute, scarious. The other morphological
characters are more or less similar to P. quadrifida.

2.2 Microscopic characters

The cross-section of the stems of both the species are almost circular (figures 2
and 9). The epidermal cells are polygonal in shape in both species and are
surrounded externally by thick striated cuticle. The outer wall of some of the

Figures 1-7. 1. A twig of the plant Portulaca quadrifida L. (X 2.5). 2. T.S. stem
(diagrammatic) (X35). 3A, B. Anatomical details of a portion of figure 2 (X2973).
4. Epidermal cell in T.S. containing acicular crystals (X2973). 5A, B, C D
and E. Prismatic crystals and druses (X2973). 6 and 7. Macerated xylem
vessels (X2973).

Pharmacognosy of P. quadrifida L. and P. oleracea L.

237

epidermal cells of P. quadrifida slightly bulge out. Acicular crystals which appear
as crystals and in cross section are present m some of the epidermal cells of
P. quadrifida (figure 4). Epidermis is, followed by 2-3 layers of collenchyma cells
in the stem of P. oleracea (figures 9 and 10A), whereas it is parenchymatoiis in
P. quadrifida (figure 3A). The parenchyma in both species consists of thin-walled,
more or less isodiametric cells with large intercellular spaces. These cells are
loaded with starch grains, simple as well as compound. The compound starch
grains are usually 2 or 3 celled (figures 5F and 11). Druses, prismatics, acicular
crystals and colourless mucilage cells are commonly present in both the species.
The endodermis in both species is not well defined. Collateral vascular bundles
are arranged in a ring in both but the number of bundles in P. oleracea is almost
double or even more than those in P. quadrifida (figures 2 and 9). Pith consists of
thin-walled isodiametric cells some of which contain calcium oxalate crystals
(figure 3B). The macerated xylem consists mostly of helical and scalariform
vessel elements with simple perforation (figures 6, 7A> B, C and 12) and fibres
with intrusive growth.
The measurement of different tissues and cells is given in table 1.

Table 1. Measurement of different tissues and cells in microns.

P. quadrifida

P*oleracea

Cuticle M = 3.33 — 6.66 thickness
Epidermis M = 23.31 x 6.66 — 39.96

x 19.93 — 79.92 x 49.94

Parenchyma M = 6.66 — 13.32 — 93.24 diameter
Collenchyma Absent

Vessels M = 15.65 — 19.98 — 33.30 diameter
T = 79.92 X 23.31 — 123.21
x 63.27 — 404.0 X 31.10
Fibres M = 15.55 — 23.32 — 31.10 diameter
T = 155.50 X 23.32 — 233.25

X 31.10

Pith M = 16.65 — 23.31 — 66.60 diameter
Druse M = 33.30 — 39.96 — 49.96 diameter
Starch grains M = 3.33 — 9.99 — 13.32 diameter

3.66 — 6.89 thickness
39.6 x 26.4 — 42.9 x 29.7

— 49.6 x 36.4

46.8 — 124.8 — 156.0 diameter
M = 33.3 — 66.6 diameter
9.99 _ 26.24 — 29.97 diameter
90.0 x 28.54 — 223.31 x 68.27

— 532.98 X 35.30

46.8 — 124.8 — 153.50 diameter
46.8 — 78.0 — 124.8 diameter
1.66 — 3.33 — 16.50 diameter

M = measurement in cross-section; T = measurement of macerate

2.3 Macerate

Macerate consists of cuticle, parenchyma cells, xylem vessel elements, fibres, starch
grains and druses (figures 5, 6, 7, 11 and 12).

2.4 Extractive and ash values

Extractive and ash values were determined according tcf Anoja (1966) and the
results are given in table 2.

238

J Lai and A M Khan

Figures 8-12. 8. A twig of the plant Portulaca oleracea L. (Xl). 9. T.S.
stem (diagrammatic) (X20). 10A, B and C. Anatomical details of a portion
of figure 9 (X2973). 11. Starch grains and druses (x2973). 12. Macerates
(X2973).

Table 2. Extractive and ash values of the sterns of P. quadrifida and P. oleracea.

Extractive and ash values

P. quadrifida

P. oleracea

Water soluble extractive

19.73%

25.00%

(chloroform water)

Alcohol soluble extractive

10.32%

18.50%

Total ash

9.09%

25.18%

Acid insoluble ash

0.63%

3.18%

2.5 Fluorescence analysis of the powdered drugs

The stem powders prepared by drying fresh specimens at 60°C#were chemically
treated and exposed to ultraviolet light. The fluorescence observed is recorded
in table 3.

Pharmacognosy of P. quadrifida L. and P. oleracea L. 239

Table 3. Fluorescence analysis of the stem powders of P. quadrifida and P. oleracea.

Chemical treatments

Fluorescence

P. quadrifida

P. oleracea

>owder as such

Light green

Dark green

>owder mounted in IN NaOH

in methanol

Yellowish green

Green

*owder mounted 4n nitrocellulose

Orange

Brownish green

»owder treated with IN NaOH

in methanol and mounted in

nitrocellulose

Brown

Dark brown

1.6 Chromatographic studies

Vlcohol extracts of the stems of P. quadrifida and P. oleracea were subjected to
bin layer chromatography with the solvent system methanol : chloroform (3:7).
rhe plates were developed by iodine vapours. They showed the presence of four
pots (figure 13) with Rf Values 0.05, 0.65, 0.73, 0.90 and 0.05, 0.65, 0.76, 0.88
espectively. This indicates thai the two species have more or less the same
jhemica1. constituents.

Conclusion

two species differ considerably in their measurements of cells and tissues
[table 1), extractive and ash values (table 2) and fluorescence analysis of the
)owdered drugs under uv light (table 3). Little or no differences were obtained
n TLC studies as shown in the chromatogram (figure 13), Undoubtedly, the

Chromotogram

0

0 0
o o

o o

t-f

A - Alcoholic extract

of portulaca oleracea linn

B- Alcoholic extract of
portulaca quadrifida linn

Figure 13. Chromatogram.

240 / Lai and A M Khan

stems of P. quadrifida and P. oleracea differ morphologically and anatomically.
They also differ in extractive and ash values as well as in fluorescence analysis
under UV light; yet in view of the similarities in therapeutic properties the stem
of P. quadrifida can be used as a substitute for that of P. oleracea.

Acknowledgements

The authors are thankful to the Director, Central Council for Research in Unani
Medicine for his unceasing encouragement and for providing financial assistance.

References

Anon 1969 Portulaca Linn. (Portulacaceae); The Wealth of India (New Delhi: CSIR),

pp. 219-.221
Anon 1966 Determination of ash Pharmacopoeia of India (New Delhi: Govt. of India)

pp. 947-949
Chase C R and Pratt R 1949 Fluorescence of powdered vegetable drugs with particular

reference to development of system of identification; /. Am. Pharm. Assoc. (Sci. ed).

38 pp. 324-331
Dyrnock W, Warden C H J and Hooper D 1980 Portulacaceae; Pharmacographia Indica

(Dehra Dun : B S M P Singh), 1 pp. 158-159
Kirtikar K R and Basu B D 1975 Portulacaceae; Indian medicinal plants (Reprint Edition)

(Dehra Dun: B S M P Singh) 1 pp. 240-245
Lai J 1980 Foliar Pharmacognosy of Portulaca quadrifida Linn. Bull. Med. Ethnobot. Res.

I pp. 46-54
Sadana J C and Ahmad B 1947 Observations on the Carotenoid pigments of common Indian

vegetables; /. Sci. Ind. Res. B6 pp. 47-52

Abbreviations : CA : Cambium; ACO : Angular collenchyma; CR : Druses;
cs : Crystal sand; csx : Compound starch grains; cu : Cuticle; EP : Epidermis;
INV : Involucre; LF : Leaf; MA : Nodal appendages; PAR : Parenchyma;
PHI : Phloem; PIT : Pith; PR : Prismatic crystal; ST : Starch grains;
VB : Vascular bundle; XY : Xylem.

Proc. Indian Acad. Sci. (PJant ScL), Vol. 91, Number 3, June 1982, pp. 241-253.
O Printed in India.

Structure and function of a sub-tropical humid forest of
Meghalaya L Vegetation, biomass and its nutrients

JASBIR SINGH and P S RAMAKRISHNAN

Department of Botany, School of Life Sciences, North-Eastern Hill University,
Shillong 793 014, India

MS received 10 January 1981; revised I May 1982

Abstract. The peripheral disturbed zone of a 50-year old stand of the
forest at Lai lad was dominated by D emir o calamus hanriltonii, an early successional
bamboo characteristic of a secondary successional fallow of not more than
20-25 years while the central undisturbed zone had Schima wallichii, Castanopsis
indica and Shorea robusta as important components. The biomass of the forest
was computed as 137 X 10" kg/ha of which 64.7% was in the central zone. The
contribution by different species both in the central and peripheral zone of the
forest was worked out. Linear relationship between dbhfd^h and biomass was
worked out for different species. The standing crop had : N, 953 ; P, 284; K, 600;
Ca, 2281 ; and Mg, 450 kg/ha, of which 60% was in the central zone of the forest.

Keywords. Biomass ; nutrients; sub-tropical forest.

1 . Introduction

Understanding of the structural and functional attributes of a given forest
ecosystem is important for proper management of the environment and utilization
of the resource potential (Grantham and Ellis 1974). The present study on a
50-year old humid tropical forest ecosystem of north-eastern India is important
as it represents a stage in secondary succession after slash and burn agriculture
(locally known as jhum) which is a wide spread land use practice in this region
(Ramakrishnan et al 1981). The present series of three papers deal with forest
ecosystem from the point of view of its organization, biomass and nutrient flow
through litter and through water.

The structure of a vegetational unit depends upon the species composition and
their relative number (Gleason 1926). Biomass data form an important component
of the structure of any ecosystem (New Bould 1967). Though much infor-
mation on this and inventory of nutrients is available on different forest
types from the parts of the world, little is known on Indian forest types except for a
few studies (Dadhichi 1979; Agrawal 1980; Vyas et al 1980). This paper is related

241

242

Jasbir Singh and P S Ramakrishnan

to phytosociology and biomass analysis alongwith nutrient inventory of the forest
type mentioned above.

• Max. temperature
o Min. temperature
Q Rainfall

500
400

~ 300

*§ 200

TOO
0

n

10 1

J FMAMJ J ASONO
Months

Figure 1. Climate of Lailad based on the average of two years - 1977 and 1978 (data
obtained from Department of Silviculture, Government of Meghalaya).

2. Study area, geology and climate

The study area located at Lailad (25°45"~ 26°0" N Latitude and 91°45" - 92°OE
Longitude) at an altitude of 296 m, is tropozoidal shaped and is a part of a reserve
forest known as Nonghyllem reserve under the Meghalaya Forest Department
since 1910. The peripheral zone is subjected to biotic disturbances,
such as cutting of trees for firewood and removal of bamboo for fencing and house
building by the local tribal population. Therefore, only the central zone of the
forest is representative of a 50-year old stand. The soil is red, sandy loam and
is of laterite origin. ThepH ranged from 5.8 to 6.3. The climate is typically
monsoonic with most of the rainfall (84%) during May to September. The mon-
soon season is followed by a mild winter (mid November to mid February). March
and early April represent a brief dry summer period (figure 1).

2.1 Methods of study

Phytosociological studies were done in July 1976, at the peak of the growing
season both in the outer peripheral zone of about three hectare. The density,
frequency, basal area and importance value index were based on 20 quadrats of
10 X 10m for trees, 5 X 5m quadrats for shrubs and 1 X 1m quadrats
for herbs along a transect running from the periphery to the centre of the forest

Vegetation, biomass and its nutrients 243

(Misra 1968; Kershaw 1973). Species diversity was calculated using the formula
»iven by Shannon and Weaver (1949)

where H = Shannon index of general diversity, ni = importance value index of
ipecies /, N = Importance \alue index in the community,

The index of dominance of the community was calculated according to
Simpson (1949).

as C =

where C = concentration of dominance ; ni • importance value ofspscies /,
N = total importance value for the community.

Biomass estimation of tree species was done in a specified area during August at
:he time when the leaves were fully mature. Three different girth classes with 3 repli-
cates of 11 major tree species, namely, Shorea robusta, Schlma wallichii, Castono-
?sis indica, Artocarpus chaplasha, Gmelfne arbor ea, Garcinia cowa, MiUiusa roxbur-
%hiana9 Sterculia villosa, Dillenia indica, Vitex pedimcularls and Dendro calamus
hamiltonii were harvested. Various parameters like diameter of the bole at base,
niddle and top, total height of the tree, diameter and length of the branches and
;otal number of leaves were recorded after harvest. The fresh weight of all the
branches and leaves were determined in the field and sub-samples of branches,
small twigs and leaves were brought to the laboratory in polythene bags. Small
liscs (2-3 cm thick) from the base and top of the bole were taken for cornpula-
:ion of dry weight. All the sub-samples were oven dried at 80° - 85° C to cons-
ent weight. The regression equations were obtained relating the biomass
parameters to combinations of diameter at breast height (dbh) and diameter2 X
leight (d*h) . Regression equations were used to estimate the biomass of the
itanding crop.

Plant samples were ground and passed through a 20-mesh screen and chemically
maiyzed for nitrogen, phosphorus, potassium, calcium and magnesium using
rtandard methods as described by Peach and Tracy (1956) and Jackson (1958).
rhus, nitrogen was determined by micro-kjaldahl method and phosphorus was
estimated colorimetrically by molybdenum blue method. After dry ashing the
;amples, calcium and magnesium were analyzed by EOTA-filtration method aad
Dotassiurn by flame emission method.

I. Results and discussion

l.l Vegetation structure

Some of the most important component of the peripheral disturbed zone were
Dendrocalamus hamiltonii, Mesuaferrea, Millusa roxburghiana, Vitex peduncularis>
Schima wallichii, Castonopsis indica and Shorea robusta. It may be noted that
9. hamiltonii which is more dominant in the peripheral zone is an early successional

244

Jasbir Singh and P S Ramakrishnan

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Vegetation, biomass and its nutrients 245

species and reaches its peak in a 20-year old community after which it dec-
lines (Ramakrishnan and Toky 1978). On the other hand species like Schima
wallichii, Castonopsis indica and Shorea robusta were more dominant towards the
central zone of the forest with higher IVi values. In forest community as a
whole, dominant tree species were Dendrocalamut hamlltonii, Schima waUichii,
Castonopsis indica, Shorea robust a, Milusa roxburghiana and Artocarpus chaplasha
(table 1). M. roxburghiana had high density and frequency but low IVI values
due to low basal area whereas Artocarpus chaplasha and Vitex peduncular is had
high IVI values in spite of low density and frequency because of their greater
basal area.

Croton oblongifoliwn, Litsaea khasyana, Leea samhucina, Annona wallichli,
Randia demi flora and Micromelum pubescence were the important of the shrub
layer of the forest as a whole. It may be noted that species like Croton oblvngi-
folium had high IVI values towards the peripheral zone of the forest along with
species like Annona wallichi, Combretum decundrurn, Litsaea khusyana, etc. In the
forest as a whole, Panicum khasianum, Cyperus elegans, Hedychhim gracile, etc. are
predominant amongst herbaceous species (table 1). However, a number of these
species possess greater IVI in the peripheral zone in comparison to the central
zone of the forest. This may be due to lesser number of tree species with reduced
canopy cover which permitted greater light penetration and less competition in the
peripheral zone.

3.2 Biomass

The pattern of biomass distribution in the forest for the important species may be
related to the species diversity. Of the total biomass 64.7% was along the central
zone ducto greater species diversity whereas peripheral zone contributed about
35.3%. Schima wallichii, Castonopsis indica and Shorea robusta contributed maxi-
mum in the peripheral zone as well as central zone. Howqver, these three species
contributed more than 2-fold in the central zone of the forest compared to the
peripheral zone (table 2).

Leea stambucina, Annona wallichii, Sterculia cacclnia, Litsaea khasyana and
Croton oblongi folium account for the largest biomass contributed by the shrubs in
the forest. Of the total biomass contributed by shrubs, 54.3% was along the
peripheral zone and the remaining 45.7% was in the central zone of the forest.
Of the total plant biomass in the forest as a whole 64.5% was in the central zone
and only 35.5% along the peripheral zone. Along the periphery, shrubs and
herbs contributed larger share of the total biomass whereas trees contributed
mostly towards the central zone of the forest (table 3).

The average biomass on an unit area basis i'n the forest was 137 X 103 leg/ha,
the value being lower than those reported by other workers for the tropical
forests (Jordan and Kline 1972). Whittaker and Likens (1973 a, b) reported a
mean value of 35-45 X 104 kg/ha biomass for some tropical and seasonal forests.
However, the present value lies between 60~350m*lia reported by Whittaker
(1975) for some temperate evergreen and tropical seasonal forests and much higher
than that for temperate coniferous forest (Akai et al 1968; Smith et al 1971).

246 Jasbir Singh and P S Ramakrishnan

Table 2. Biomass contribution by major tree, and shrub species in the
central zones of the forest at Lailad (values in parentheses represents
of the total amount).

peripheral and
the percentage

Species

Biomass (kg/ha)

Peripheral zone

Central zone

Trees

Artocarpus chaplasha

3796.2

(1.4)

9124.2 (3.3)

Castonopsis indica

22484.1

(8.2)

45199.9 (16.4)

Dendro calamus hamiltonii

6914.8

(2.5)

3638.2 (1.3)

Dillenia indica

4594.6

(1.7)

2938.4 (J.7)

Grnelina arborea

2033.8

(0.7)

4589.2 (1.7)

Garcinia cowa

1967.3

(0-7)

51187 (1.9)

Miliusa roxburghiana

61857

(2.2)

4744.5 (1.7)

Schima wallichii

35113.9

(12.8)

74900.4 (27.2

Shorea robusta

8416.3

(3.1)

21395.9 (7.8)

Sterculia villosa

4674.8

(1-7)

2614.4 (0.9)

Vitex peduncularis

1057.1

(0.4)

1698.7 (0.6)

Shrubs

Anona wallfchti

196.2

(6.2)

65.4 (2,1)

Actinodaphne angustifolia

141.2

(4.5)

94.4 (30)

Croton oblongi 'folium

184.0

(5.8)

147.2 (4.6)

Combretum decandrum

127.4

(4.0)

72.8 (2.3)

Litsaea khasyana

186.0

(5.9)

248.0 (7.8)

Lfea sambiicina

265.0

(8.3)

371.0 (11.7)

Morinda umbel/ ata

151.2

(4.8)

108.0 (34)

Phlogacanthus thyrsiflorus

2030

(6.5)

124.8 (3.9)

Randia densiflora

73.6

(2.3)

92.0 (2.9)

Others

193.0

(6.1)

131.2 (4.1)

Table 3. Biomass contribution by major tree species, shrub and herbaceous layers
in the peripheral and central zones of the forest at Lailad (values in parentheses
represents the percentage of the total amount).

Different layers

Biomass (t/ha)

Peripheral zone

Central zone

Tree
Shrub
Herbaceous

97.234 (35.3)
0.172 (0.01)
0.008 (0.03)

177.984 (64.6)
0.146 (0.05)
0.004 (0.01)

Relationship between morphological growth parameters viz., diameter (dbh),
and diameter2 X height (d2h) on the one hand and biomass of individual tree
species as well as fractional plant parts on the other was found to be highly
significant for all the tree species except D. hamiltonii where significant correla-
tions were found with dbh (tables 4,5).

Vegetation, biomass and its nutrients

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Vegetation, biomass and its nutrients . 249

.3 Nutrient content in biomass

ji analysis of the concentration of M,P, K, Ca and Mg in different plant
omponents (bole, branches and leaves) showed that the leaf had higher pcrcen-
ige of N, P, K, and Mg while bole alongwith bark contained higher percentage
f Ca in all the species except Dendro calamus hamihonii where the concentration
f Ca was more in leaves. Next to the leaves, the branches had higher levels
f N, P, K and Mg and the bole had the least concentration. On the other hand, Ca
Dncentration was the least in branches with intermediate values from leaves
:able 6).

Table 6. Concentration of different nutrients in different compartments of
different tree species at Lai lad.

Species

Compart-
ment

N

P

K

Ca

Mg

Bole

0.41

0.12

0.31

3.00

0.14

Artocarpus chaplasha

Branches

0.61

0.23

0.32

0.82

0.49

Leaves

2.10

0.64

0.59

2.96

0.77

Bole

0.64

0.14

0.58

0.84

0.47

Dcndrocalamus hamiltonii

Branches

0.70

0.28

0.52

0,69

0.70

Leaves

1.76

0.59

0.83

1.99

0.88

Bole

0.85

0.14

0.26

2.90

0.18

Castonopsis indica

Branches

0.90

0.28

0.40

1.00

0.34

Leaves

1.90

0.49

0.66

2.40

0.79

Bole

0.58

0.15

0.26

2.80

0.29

Miliusa roxburghiana

Branches

0.75

0.28

0.65

1.00

0.83

Leaves

1.65

0.47

0.85

3.20

1.09

Bole

0.59

0.16

0.48

3.36

0.26

Schima wallichii

Branches

0.62

031

063

0.70

0.45

Leaves

1.86

066

1.19

260

0.67

Bole

0.42

0.11

0.34

2.40

0.30

Shorea robusta

Branches

0.82

0.26

0.44

0.88

0.68

Leaves

1.90

0.76

1.20

1.78

1.08

On hectare basis, the total amount of different elements contributed by different
pecies (table 7) showed that Schima wallichii, Castonopsis indlca and Shorea
riusta contributed maximum with respect to N, P, K, Ca and Mg. This could
e related to the large biornass in the standing crop of these tree species
i the same order as given above. Among other species a great variation
fas observed with regard to percentage contribution of different elements
'hich may be partly due to the concentration of nutrients. Thus, Dendro-
alamus hamiltonii stands next to Artocarpus chaplasha and Miliusa roxbur-
hiana in biomass but contributed more in terms of P, K and Mg due to
igher concentration of these three elements in the plant tissue. Because of low
'a concentration in the tissue coupled with low biomass the contribution of Ca
y Dendrocalamus hamiltonii was far less than that of Artocarpus chaplasha and
diliusa roxburghiana.

250

Jasbir Singh and P S Ramakrishnan

The distribution of nutrients in different tree compartments showed that bole
contained maximum amount of all nutrients followed by branches and leaves.
This is in spite of the higher concentration of N, P, K and Mg in the leaf tissue
This compartmentalization of nutrient is highly exaggerated for Ca due to the fact
that this nutrient also had the highest concentration in the bole (figure 2).

Table 7. Amount of different elements contributed by different species alongwith
percentage contribution of the total for the forest given in parentheses.

Species

Nutrient (kg/ ha)

N

P

K

Ca

Mg

Artocarpus chaplasha

34.7

11.3

21.2

135.8

17.2

(3-8)

(4.0)

(3.6)

(4.8)

(3.9)

Dendrocalamus hamiltcnil

?2.4

22.0

31.7

36.1

29.2

(3.5)

(7.9)

(5.4)

(i-4)

(>/)

Dillenia indica

23.7

H.I

21.1

74.6

10 3

(2.6)

(2.9)

(3.6)

(2.7)

(23)

Castanopsis indica

291,7

61.4

102.5

439.9

79.3

(32.0)

(22.0)

(17.4)

(25.7)

(17.9)

Gmelina arborea

18.8

5.2

16.6

63.8

12.7

(2.1)

(1.9)

(2.8)

(2.0)

(2.9)

Garcinia cowa

18.1

8.5

10.5

61.2

12.8

(2.0)

(3.0)

(18)

(2.2)

(2.9)

Miliusa roxburghlana

27,7

11.0

11.7

113.1

25.3

(3.0)

(3.9)

(2.0)

(4.7)

(57)

Schima wallichii

346.7

115.0

294.0

1086.6

174.0

(38.1)

(41.2)

(499)

(45.5)

(39.3)

Shorea robusta

83.7

25.2

58.1

181.7

61.0

(9.2)

(9.0)

(9.9)

(8.9)

(13.8)

Sterculia villosa

23.1

9.1

14.6

54.0

16.8

(2.5)

(3.2)

(2.5)

(1.9)

(3.6)

Vitex peduncularis

10.1

2.5

7.0

22.4

3.8

(I.I)

(0.1)

(1.2)

(0.9)

(0.7)

Figures indicates the percentage contribution (per hectare) by the different
elements in the different compartments of the standing crop. The quantities of
the different nutrients in the three compartments of the tree in a decreasing order
are as follows :

Ca>N>K>Mg>P.

The pattern of distribution of nutrients by trees, shrubs and herbs along the peri-
phery and the centre of the forest showed that about 60% of the total nutrient pool
was in the undisturbed central zone and the rest along the disturbed peripheral zone
(table 8), However, along the peripheral zone the contribution by shrub and
herb species was more (0.9 and 0 3% respectively) in comparison with the central
zone (0.7 and 0.2% respectively). Tree species contributed more towards the
nutrient pool of the living biomass (59.9%) in the central zone than along the
peripheral zone (37.9) of the forest.

Vegetation, biomass and its nutrients

251

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2100

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Figure 2. Pattern of compartmentalization of nutrients in trees at Lailad forest.
Open column bole; hatched column branches; closed column leaves.
Figure 3. Proportion of different nutrients in different components in living biomass of
trees (percentage of the total amount in kg/ha). Closed column N; strippled column P;
hatched column KL; open column Ca; cross hatched column Mg.

The total standing crop of nutrients as worked out in the present studies was
higher when compared with broad leaved temperate forests (Ovington 1958;
Grier el al 1974); DIvigneud et al (1968) but the values are lower when compared
with tropical forests of Ghana (Greenland and Kowa 1960) and some Indian
forest type (Deshbandhu 1970 ; Faruqi 1972)

Acknowledgements

The help given by Mr Ashish K Das in the preparation of this manuscript
is gratefully acknowledged. One of us (JS) was a recipient of a Fellowship of the
North-Eastern Hill University.

References

Agrawal S K 1980 Community structure of the deciduous forests at Prasad; Acta Ecol. 2 (3-4)
36-41

Akai T, Furono T, Ueda S and Soichi S 1968 Mechanisms of matter production in young loblo-
lly pine forest Kyoto Univ.; Japan For. Bull. 40 26-49

Vegetation., biomass and its nutrients 253

Dadhichi L K 1979 Studies on the productivity of dry deciduous trees at JharnarJcotra (i)

Anogeissus pendula Endew (ii) Wrightia tinctoria R.Br. ; Ada EcoL (1-2) 28-33
Deshbandhu A 1970 A study of the productive structure of northern tropical dry deciduous

forests near Varanasi; Trap. EcoL 11 90-104
Duvignaeud P S, Denaeyer-De Srnet and Marbaise J L 1969 Littere totale et restitution an sol

dcs polyelements biogens; Bull. Soc. Roy. Bot. Belg. 102 339-354
Faruqi Q 1972 Organic, mineral structure and productivity of Sal (Shorea robusta Gaertn.) and

teak ( Tectona grandis Linn). Ph.D. thesis. (Varanasi: Banaras Hindu University)
Grleason H A 1926 The individualistic concept of the plant association; Bull. Torrey. Bot. Club 53

7-26

Grantham J B and Ellis T H 1974 Potentials of wood for productivity energy; J. For. 72 552-556
Greenland D J and Kowal J M L 1960 Nutrient content of the moist tropical forest of

Ghana; Plant Soil 12 154-174
Grier C C, Cole D W, Dyrness C T and Fedreksen R L 1974 Nutrient cycling in 37 and 450-year

old Douglas fir ecosystem; In : Integrated research in the coniferous forest Biome (ed)

R H Waring and R L Edmunds 8-21

Jackson M L (ed) 1958 Soil chemical analysis (New Jersey: Prentice Hall. Inc. Englewood Cliffs)
Jordan C F and Kline J R 1972 Mineral cycling : Some basic concepts and their application

in a tropical rain forest; Adv. in EcoL and Syst. 33-50

Kershaw K A 1973 Quantitative and Dynamic Plant Ecology (London : Edward Arnold)
Misra R (ed) 1968 Ecology work Book. (New Delhi : Oxford and IBH Publication Co.)
STewbould P J 1967 Methods of estimating the primary production of forests. Black-well Sci.

Publ. Oxford - Edinburgh. IBP Hand Book No. 2
Dvington J D 1958 The sodium, potassium and phosphorus contents of tree species grown in

close stands; New Phytol. 57 273-284
Dvington J D 1959 The circulation of minerals in plantations of Finns sybestris L.; Ann. Bot.

(NS) 23 229-239
?aech K and Tracey M V (ed) 1956 Modern methods of plant analysis Vol. Ill (Berlin; Springer

Verlag), pp. 542

Ramakrishnan P S and Toky O P 1978 Preliminary observations on the impact of jhum (shift-
ing agriculture) in the forested ecosystem. In : Resource development and environment in the

Himalayan region', Dept. of Sci. and Tech. Govt. of India 343-354.
Ramakrishnan P S and Toky O P 1981 Soil nutrient Status of hill agroecosystem and recovery

pattern after slash and burn agriculture (Jhum) in north eastern India, Plant Soil 60 41-64
Ramakrishnan P S, Toky O P, Mishra B K and Saxena K G 1981 Slash and burn agriculture

in North-Eastern India. In: Fire regimes and ecosystem properties, (ed) H A Mooney,

T M Bonnickson, N JL Christensen, J E Lotan and W A Reiners, USDA Forest Service

general technical report WO-26 pp. 570-587

Shannon C E and Weaver W 1963 The mathematical theory of communication", (Urbana : Uni-
versity of Illinois Press), pp. 117

Simpson E H 1949 Measurement of diversity; Nature (London) 163 688
Smith W H, Nelson L E and Switzer G L 1971 Development of the shoot system of young

loblolly pine II Dry matter and nitrogen accumulation; For. Sci. 17 55-62
yyas L N, Garg R K, Vyas N L and Jindal K 1980 Community structure and above ground

biornass in dry deciduous forests around Udaipur (Rajasthan) India; Acta EcoL 2 25-32
Whittaker R H (ed) 1975 Communities and ecosystem. Second ed (London and New York:

MacMillan) pp. 385
Whittaker R H and Likens G E 1973a Carbon in the biota. In : Carbon in the biosphere (eds.)

G M Wood Well and E V Pecan Brookhaxen Symp. Biol. Spring field. Virginia
Whittaker R H and Likens G E 1973b Primary production the biosphere and man; Hum. EcoL

1 357-369

Proc. Indian Aca<L Sci. (Plant Sci.), Vol. 91, Number 3, June 1982, pp. 255-268.
© Printed in India.

Structure and function of a sub-tropical humid forest of
Meghalaya II- Litter dynamics and nutrient cycling

JASBIR SINGH and P S RAMAKRISHNAN

Department of Botany, School of Life Sciences, North-Eastern Hill University,
Shillong793 014, India

MS received 10 January 1981 ; revised 1 May 1982

Abstract. The litter production in a 50 year old humid sub-tropical forest at Lailad
was found to be 5.5 t/ha/yr, 77% of which was through leaf and the remainder
through \vood. Litter production was more in. the peripheral disturbed zone
compared to the undisturbed central zone which is related to the successional status
of the community. Species differences in the rate of decomposition of litter was
noted. Besides litter production and decomposition pattern were related to seasonal
differences in temperature and humidity. The present results have been discussed in
the light of the data from other studies done elsewhere.

Keywords. Litter dynamics ; nutrient cycling.

1. Introduction

Studies on litter production and its decomposition are important for understanding
of energy flow, nutrient cycling and primary production in the ecosystem. While
much is known on these aspects for temperate forests (Olson 1963; Bray and
Gorham 1964; Rodin and Bazilevich 1967; Gosz et al 1972) fewer studies
are available from the tropics (Jenny et al 1949; Loudelot and Meyer 1954;
Cornforth 1970 ; Singh and Gupta 1977 ; Edwards 1977). Little is known on the
litter dynamics of forests in India (Seth et al 1963 ; Singh 1969 ; Singh and Gupta
1977). Further, many of these studies pertain to leaf litter alone and that
derived from wood is often ignored.

In an earlier paper in this series (Part I), the phytosociology, biomass and
nutrient inventory of a 50 year old forest developed after slash and burn
agriculture (Ramalcrish.nan et al 1981) at Lailad in Meghalaya was considered.
The present study deals with the estimation of leaf and wood litter, its decom-
position and nutrient cycling in this forest stand.

2. Methods of study

For collection of litter, 20 permanent quadrats (1 X 1 m) made of wood (10
cms high) were randomly laid out in each site. Litter was collected at monthly

255

256 Jasbir Singh and P S Ramakrishnan

intervals, classified into leaves and wood and further sub-divided intc
different 'important tree species and a miscellaneous category. The litter was
oven-dried, weighed and passed through a 20-mesh screen for chemical analysis
The peripheral part of the forest was considered separated from the centra
zone (of Part I).

Decomposition of leaf litter was studied by litter bag technique (Shanks anc
Olson 1961 ; Singh and Gupta 1977). Freshly fallen leaves in April wen
collected in bulk from the forest and sorted out into five important tree
species, namely, Shorea robusta, Schima wallichn, Castonopsis indicci
Dendrocalamus hamiltonii and Artocarpus chaplasha and the remainder wen
treated as miscellaneous. The litter was brought to the laboratory and air
dried at constant temperature. Litter bags of nylon (10 X 14 cm) with 1 mn
mesh size containing 30 g samples were used. 50 bags of each species wen
prepared and decomposition of litter samples were evaluated by placing th<
bags on the surface of the soil. Three replicate samples of each categon
were recovered from the fields at monthly intervals. The material from th<
litter bags was washed with water using a 100^ mesh screen to remove all soi
particles. It was dried at 80°C, weighed and preserved for chemical analysis.

The rate of decomposition of wood litter was estimated using the methoc
given by Yoneda (1975). Freshly fallen wood pieces of moderate diamctc.
(3-4 cm) of the above-mentioned species were collected in April, air-dried a
constant temperature, cut into 12 cm long pieces and initial weight of end
piece was determined. 50 pieces for each species were randomly placed on tin
surface of the soil and three replicate of each species were picked at monthl;
interval. These were processed in the laboratory using similar procedures a;
used for leaf litter bags.

Chemical analysis of the litter was done following standard procedure
(Allen 1974). Thus, nitrogen was determined by the Kjeldahl method, anc
phosphorus was estimated color imetrically by the molybdenum-blue method
calcium and magnesium by EDTA method while potassium by flame ernissioi
method after dry ashing the samples in a muffle furnace at 450°C.

Soil respiration was measured by the alkali absorption method (Colemai
1973 ; Gupta and Singh 1977 ; Gupta and Singh 1981) using 16.6 cm diamete
metallic cylinder with 50 ml of 1 N KOH solution. The alkali solution wa
kept in the cylinders for 24 hours. Soil respiration was measxired in two type
of situations, (i) On the mineral soil after removing the litter layer, and (ii) Oi
the soil without removing the litter layer. A third set served as the blank for whicl
the beaker was placed on a wooden platform lined by a layer of polythene sheet
Three replicates were done for each treatment.

3. Results and Discussion

3.1 Litter production

Total litter production was estimated to be 5 5 t/ha/yr of which 77% wa
through leaves and the remaining through wood. Litter production was mor<

Litter dynamics and nutrient cycling 257

along the periphery than the central zone of the forest. Dendrocalamus hamiltonii
(bamboo) a comparatively early successional species was an important compo-
nent in the periphery due to frequent disturbances like felling of dicot trees for
timber and fuel wood and occurrence of accidental fire ; this species alone
contributed 19% of the total litter. This species has been shown to have a
high rate of turnover of biomass through leaves in the younger fallows up to 20
years (Ramakrishnan and Toky 1981). In the central zone of the forest contribu-
tion of litter was chiefly through Shorea rohusta, Schima wallichii and Castonopsis
indica with 13, 10 and 8% respectively of the total (table 1).

The average litterfall of the forest as a whole (5.5 t/ha/yr) was slightly lower
than the values obtained for other tropical forests. At one end of the range
are the values recorded by Mitchell (cited by Bray and Gorham 1964) for
Malaya (5.5-7.2), Klinge and Rodrigues (1968) for Amazonia (7.3) and
Edwards (1977) for lower Montane Rainforest in New Guinea (7.5). The values
reported on higher side are by Bernhard (1970) for Ivory Coast (8.3-13.4)
and Ewel (1976) for Guatemala (9,0). Toky and. Ramakrishnan (1980) reported
litterfall of 9.7 t/ha/yr in a successional forest of 20-year dominated by bamboo
(D. hamiltonii) in the same area where this study was done. This may be due
to the fast developing vegetation during the early successional stages and the
consequent rapid turnover of biomass. Since an early successional community
undergoes fast changes in species composition, often entire individual may con-
tribute to litter production resulting in an over shoot of litter production in
early successional phase (Toky and Ramakrishnan 1980 ; Ewel 1976).

Table 1. Distribution of litter along the peripheral and central zones of the sub-tropical
forest at Lailad (kg/ha/yr). (Figures in parentheses represent the percentage of the total
litter in each zone).

Species Peripheral zone Central zone

Leaf litter :-

Ar to car pus chap las ha

102.2 (2.3)

204.5 (3.1)

Castanopsis indica

245.8 (5.4)

491.8 (7.5)

Dendrocalamus hamiltonii

846.6 (18.7)

282.2 (4.3)

Dillenia indica

25.2 (0.6)

33.1 (0.5)

Garcinia cowa

48.8(1.1)

145.0 (2.2)

Machillus khasiana

4.0 (0.1)

5.0 (0.1)

Mesua ferrea

40.2 (0.9)

14.4 (0.2)

Miliusa roxburghiana

311.3(6.9)

140.7 (2.2)

Shorea rohusta

261.9(5.8)

849.5(13.0)

Schima wallichi

297.8 (6.6)

643.6(9.9)

Sterculia villosa

64.7(1.4)

26.3 (0.4)

Vitex peduncular is

36.6 (0.8)

43.4(0.7)

Other species (leaf)

1530.5 (33.9)

1796.0 (27.6)

(miscellaneous)

Total

3815.6

4675.5

Wood litter :-

Branches, twigs, barks.

701.0(15.5)

1833.8 (28.2)

(all species)

Total (leaf -f wood) litter

4516.6

6509.3

Mean of peripheral and central zone

5512.9

258

Jasbir Singh and P S Ramakrishnan

Wood litter (1.3 t/ha/yr) formed 23% of the total litterfall and the value is
comparable to 1.2 t/ha/yr (16% of the total) as reported by Edwards (1977) in
Lower Montane Rain forest in New Guinea. For temperate forests a range
of 22-78% of the total contribution through wood litter has been estimated
(Carlisle etal 1966; Anderson 1970).

The distribution of litterfall was markedly seasonal with a maximum leaf
litterfall (57%) during the dry months of February, March and April. Wood
litterfall showed a peak during April to July (figure 1). A similar seasonal trend
in litterfall was also found in other tropical rain forests (Nye 1961),Klinge
and Rodrigues (1968). Data of Laudelot and Meyer (1954) for young secondary
forests showed two periods of litterfall that come at the end of the drier season.
This may be related to the formation of abscission layer in leaves due to the
severity of drought stress during dry periods. During the present study, the peak
of the wood litterfall extended into the rainy season. This may be related to
storms prevailing during the rainy season, as also reported elsewhere (Edwards
1977).

3.2 Litter decomposition

Most of the workers who have considered the rate of decomposition of litter on
the forest floor (Jenny etal 1949; Olson 1963; Edwards 1 977) have assumed
that there is an exponential loss in weight as a result of decomposition, i.e.

Xlx0 = exp ( — kt)

where x0 is the initial weight, x is the weight at time r, and k is the annual
decomposition constant. This model expresses the loss as a negative exponen-
tial function of the fraction and calculation of k remains the most convenient
means of comparing forests.

During the present study the rate of decomposition was rapid in
D. hamfltonii, Schima wallichii and miscellaneous litter (high k values) than
other types of litter (table 2). The overall rate of decomposition in this
forest for wood and leaf litter was found to be lower than the values reported
by Laudelot and Meyer (1954) for Zaire and Singh (1968) for deciduous forest

Figure 1. Monthly litter production in a humid sub-tropical forest at Lailad
(1977-78).

Litter dynamics and nutrient cycling 259

at Varanasl (India) and was comparable to the values reported by Edwards
(1977) for New Guinea. Nitrogen content and the texture of the litter play a
great role in decomposition (Ewel 1976; Singh and Gupta 1980), however, Singh
(1968) correlated various chemical constituents and the rate of decomposition
in tropical tree species and found that not only nitrogen but numerous
chemicals interact to affect the rate of decomposition. The nitrogen content of
leaf litter (table 3) in S. rohusta, C. indica and A. chaplasha was quite high
but still a low rate of decomposition was observed in these species. This may
be due to decay resistant petiole and mid-rib of leaves due to high content of
lignin (Singh 1968). The lower decay rate may also partly be due to the
smaller mesh size of litter bags which do not allow larger fauna to enter
(Edwards and Heath 1963).

After a period of one year, the highest loss of litter was observed for
D. hamiltonii (78%) and least for S. rohusta (56%) (figure 2) 40-45% of litter
was lost during May to August due to higher temperature and humidity.
Subsequently the rate of decomposition slowed down (figure 3) due to low
temperature and moisture levels. This pattern of decomposition was also evident
from the evaluation of CO2 from litter layer on the mineral soil (figure 4).

Table 2. Decay constants and time required for the loss of one-half and 95% of the
original leaf and wood dry weight in different species.

Time parameter (years)

Species

Half-time
(0.693)

95%
(3)

K

K

Leaf

Wood

Leaf

Wood

Artocarpus chaplasha

1.99

0.79

1.50

3.78

Castanopsis indica

1.87

1.10

1.61

2.78

Dendro calamus hamiltonii

2.17

3.63

1.38

0.83

Shorea robusta

1.87

0.76

1.61

3.97

Schima wallichii

2.18

1.19

1.38

2.52

Other species

2.17

1.38

(miscellaneous)

Table 3. Chemical composition of leaf litter collected in April 1977 from Lailad forest.
Species N(%) P (%) K (%) Ca (%) Mg (%)

Artocarpus chaplasha

0.67

0.35

0.35

2.50

0.66

Castanopsis indica

0.82

0.46

0.36

1.88

0.59

Dendro calamus hamiltonii

0.68

0.32

0.22

1.15

.0.31

Shorea robusta

0.85

0.63

0.63

1.12

0.51

Schima wallichii

0.76

0.54

0.44

1.93

0.65

Other species

0.79

0.56

0.46

2.08

0.67

(miscellaneous)

260 Jasbir Singh and P S Ramakrishnan

Months

Figure 2." Rate'l of decomposition of leaf litter f expressed" as' percentage of the
original dry weight of leaves remaining'after^various"periods"of decomposition.

MJ JASONOJ F M A

Figure 3. Rate of decomposition of wood litter expressed as percentage of the
original dry weight remaining after various periods of decomposition.

Figure 4. Monthly pattern of CO2 evolution from the forest floor at Lailad.

3.3 Nutrient content in litter

The seasonal variations in concentration of nitrogen, phosphorus and potassium
was well marked in both leaf and wood litter (figures 5, 6). This may be explained

Litter dynamics and nutrient cycling

261

2-8-

0-2

M J S N J

Months

Figure 5. Monthly variation in concentration of different nutrients in leaf litter at
Lailad forest.

0-9

i 0-7
j

io-5

I 03

0-1

M J S N J M
Months

Figure 6. Monthly variation in concentration of different nutrients in wood litter
at Lailad forest.

as due to a number of factors like translocation of nutrients from leaves before
senescence, leaching of nutrients particularly potassium through rainfall and
the extent of decomposition of leaves and twigs before they fall (Nye 1961 ;
Tukey 1970; Goszetal 1972). The concentration of N, P and K was higher
in dry periods than during the rainy season ; this may be mainly due to leaching
of nutrients through rainfall.

Average concentration for the 12-months period, for various species showed
great variations. Nitrogen and calcium content was highest in Mesua ferrea,
Miliusa roxburghiana while phosphorus and magnesium concentration was found

262 Jasbir Singh and P S Ramakrishnan

Table 4. Nutrients content in leaf litter of different species and wood litter (Composite
sample for all species).

Species

N(%)

P(%)

K (%)

Ca (%)

Mg(%)

Leaf Utter :-

Artocarpus chaplasha

0.68

0.35

0.35

2.92

0.66

Castanopsis indica

0.85

0.46

0.36

2.28

0.59

Dendro calamus hamiltonii

0.74

0,32

0.22

1.39

0.36

Dillenia indica

0.64

0.24

0.54

1.86

0.28

Garcinia cowa

0.52

0.31

0.31

1.65

0.35

Machillus khasiana

0.85

0.40

0.30

1.98

0.65

Mesua ferrea

0.99

0.27

0.47

3.23

0.58

Miliusa roxburghiana

0.87

0.22

0.32

3.16

0.65

Shorea robusta

0.86

0.63

0.63

1.35

0.51

Schima wallichii

0.77

0.54

0.44

2.31

0.66

Sterculia villosa

0.66

0.30

0.40

1.33

0.42

Vitex peduncular is

0.57

0.33

0.23

1.81

0.35

Other species

0,79

0.56

0.46

2.51

0.67

(miscellaneous)

Wood litter :-

Branches, twigs and barks

0.60

0.38

0.73

0.45

(all species)

to be more in Shorea robusta, Artocarpus chaplasha and Schima wallichii. For
other tree species a great variety in concentration of different elements was
observed. In general calcium content was more followed by nitrogen in leaf and
wood litter (table 4). Ewel (1976) observed a reverse trend where nitrogen was
the predominant element followed by calcium.

3.4 Nutrient return through litter fall

The percentage contribution by different species not only depends upon litter
biomass contributed by them but also on the nutrient concentration in their
litter. Thus, even though Dendrocalamus hamiltonii had greatest litter biornass
in the forest as a whole, yet there were other species like S. robusta which
contributed higher amount of P through litter. In contrast a species like
M. ferrea which had high nutrient level in the leaf tissue for N and Ca, had
very low total contribution of these elements through litter because of lesser
litter production. Shorea .robusta, S. •wallichii and C. indica in that order
contributed high percentage of the nutrients both because of high litter production
and high nutrient concentration in their litter. The total contribution of nutrients
through litter was higher in the central zone of the forest than the periphery,
though the total production of litter was higher in the periphery than the
central zone (table 5), This maybe related to the difference in the species
composition in the zones due to difference in maturity of the forest community.
A comparison of nutrient returned through litterfall in the present study has
' been compared with other tropical and temperate forest ecosystems (table 6).
The values for all the elements were lower (except phosphorus) than the values
reported by Laudelot and Meyer (1954) for Zaire, and Nye (1961) for Ghana,

Litter dynamics and nutrient cycling

263

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Litter dynamics and nutrient cycling

265

but higher than that for (except nitrogen) Amazonian tropical Rain-forest Klinge
and Rodrigues (1968) and mixed deciduous forests (except calcium which is
comparable) at Varanasi (Singh 1969).

3.5 Nutrient release through decomposition

During the first 4 months after the placement of litter bags in the field, there
was a rapid fall in the concentration of all the elements ; potassium and nitrogen
were lost more rapidly. Loss of calcium and magnesium was comparatively
slower. A similar trend of decline in concentration of nutrients, was found
during wood decomposition too (figures 7, 8, 9). These results are similar to
the trends shown by Gupta and Singh (1977) and Ewel (1976).

Figures 7 a-d. a. Loss of nitrogen from leaf litter through decomposition. b>Loss of
nitrogen from wood litter through decomposition, c. Loss of phosphorus from leaf litter
through decomposition, d. Loss of phosphorus from wood litter through decomposition.

MJ J ASONOJ P M A

Figures 8 a-d. a. Loss of potassium from leaf litter-through decomposition, b. Loss of
potassium from wood litter through decomposition, c. Loss of calcium from leaf litter
through decomposition. d« Loss of calcium from wood litter through decomposition.

MJJASONOJFMA

MJJASONOJFMA
Months

Figures 9 a-b. a. Loss of magnesium from leaf litter through decomposition, b. Loss of
magnesium from wood litter through decomposition.

Litter dynamics and nutrient cycling 267

It may be concluded that (i) the turnover of litter is more in the peripheral
sturbed zone of the forest than in the central zone due to comparatively
triy successional stage of the community, (ii) litter production and decomposi-
3n was markedly seasonal related to the temperature and rainfall pattern. The
suits are compared with data on litter production and decomposition of
.her forest types.

cknowledgements

re are grateful to Dr O P Toky and Shri Ashesh K Das for their help in the
•eparation of manuscript. One of us (JS) was a recipient of a Junior Research
sllowship of the North - Eastern Hill University.

eferences

Hen S E 1974 (ed) Chemical analysis of ecological materials (Oxford : Blackwell Scientific

Publ.)-PP. 565
nderson F 1970 Ecological Studies in a Seaman woodlands. -and meadow area, Southern

Sweden II. Plant biomass, primary production and turnover of organic matter ; Bot. Not.

123 8-51
^rnhard P 1970 Etude de la Lirfere et de la contribution au cycle des 'elements mineraux

en foret ombrophile de cote dlvoire. Oecologia ; Plantarum 5 247-266
ray J R and Gorham E 1964 Litter production in forests of the World. In : Adv. Ecol. Res.

Vol. 2 (ed.) J B Cragg (London and New York : Academic Press) pp. 105-152
arlisle A, Brown A H F and White E J 1966 Litterfall, leaf production and the effect of defoli-
ation by Tortrix viridana in a Sessile oak (Quercus petrea) Woodland J. Ecol, 54 65-85
oleman D C 1973 Compartmental analysis of ''total soil respiration" an exploratory study,

Oikos 24 361-366

ornforth I S 1970 Leaf fall in a tropical rainforest ; /. AppL Ecol. 7 603-668
dwards C A and Heath G W 1963 The role of soil animal in breakdown of leaf material In

Soil Organisms (ed.) J Docksen and J van der Drift : (Amsterdam : North Holland )
iward P J 1977 Studies of mineral cycling in a montane rain forest in New Guinea. II.

Production and disappearance of litter ; /. EcoL 65 971-992
gunjobi J K and Fasehun F E 1972 Preliminary observations in the monthly litterfall and

nutrient contents of Pinus caribaea L Litter, Niger. J. Sci. 6 37-45
wel J J 1976 Litterfall and leaf decomposition in a tropical forest succession in eastern

Guatemala ; /. EcoL 64 293-308
osz J R, Likens G E and Bormann F H 1972 Nutrient content of litterfall on the Hubbard

Brook Experimental forest, New Hampshire, Ecology 53 769-784
upta S R and Singh J S 1977 Effect of alkali concentration, volume and absorption area on

the measurement of soil respiration in a tropical sword Pedobiologia 17 233-239
upta S R and Singh J S 1981 The effect of plant, weather variables and chemical composition

of plant material on decomposition in a tropical grassland. Plant Soil 59 99-118
;nny H, Gessel S P and Bringham FT 1949 Comparative study of decomposition rates of

organic matter in temperate and tropical region Soil Sci. 68 419-432
.linge H and Rodrigues W 1968 Litter production in an area of Amazonian terra firm forest

Amazonian 1 267-310
audelot H and Meyer J 1954 Les cycles d'elernents minerauz et de matiex organique and

forest equitarials congolaise Trans. Int. Conf. Soil Sci. (Comm. II) 267-272
i ye P H 1961 Organic matter and nutrient cycles under moist tropical forest. Plant Soil 8

333-346

268 Jasbir Singh and P S Ramakrishnan

Olson J B 1963 Energy storage and the balance of producers and decomposers in ecological
systems Ecology 94 322-333

Ovington J D 1959 The circulation of minerals in plantations of Pinus sylvestis L ; Ann. Bat.
(NS) 23 229-239

Ramakrishnan P S, Toky O P, Misra B K and Saxena KG 1981 Slash and burn agriculture in
north-eastern India. In Fire regimes and ecosystem properties, (ed) H A Mooney,
T M Bonnicksen, N L Christensen, J E Lotan and W A Reiners USDA Forest Service
general technical report WO-26, pp. 570-587

Ramakrishnan P S and Toky O P 1981 Soil nutrient status of hill agroecosystem and recovery
pattern after slash and burn agriculture (jhum) in north-eastern India Plant Soil 60 41-64

Rodin L E and Bazilevich N I 1967 (ed.) Production and mineral cycling in terrestrial vege-
tation (London : Oliver and Boyd) pp. 288

Seth S K., Kaul O N and Gupta A C 1963 Some observations on nutrition cycle and return of
nutrients in plantations at New Forests : Indian For. 89 90-98

Shanks R E and Olson J E 1961 First year breakdown of leaf litter in Southern appal achian
forests Science 134 370-376

Singh K P 1968 Litter production and nutrient turnover in deciduous forests of Varanasi. Proc.
Symp. Recent Adv. Trop. Ecol. pp. 655-665

Singh K P 1969 Nutrient concentration of leaf litter often important tree species of deciduous
forests at Varanasi ; Trop. Ecol. 10 83-95

Singh J S and Gupta S R 1977 Plant decomposition and Soil respiration in terrestrial eco-
systems Bot. Rev. 43 pp. 449-528

Tukey H B Jr 1970 The leaching of substances from plants. Ann. Rev. Plant Physiol. 21 305-324

Yoneda T 1975 Studies on the rate of decay of wood litter on the forest floor I. Some physical
properties of decaying wood ; Jpn. J. Ecol. 25 40-46

Proc. Indian Acad. Sci. (Plant Scu), Number 3, June 1982, pp. 269-280.
(P) Printed in India

Structure and function of a sub-tropical humid forest of
Meghalaya III. Nutrient flow through water

JASBIR SINGH and P S RAMAKRISHNAN

Department of Botany, School of Life Sciences* North-Eastern Hill University,
Shillong 793 014, India

MS received 10 January 1981; revised 1 May 1982

Abstract. This paper deals with water and nutrient flow through incident rainfall,
throughfall and stemflow through a 50- year old stand of forest at Lailad. A larger
proportion of water was accounted as throughfall and stemflow in March -April and
was related to canopy density and this was inversely related to interception loss which
was maximum in December. Many of the nutrient concentrations like that of
N, K, Ca, etc. increased with the maturation of the leaves during October-November.
Though concentration of nutrients was higher in stemflow, the total quantity was
more via throughfali because of larger quantity of water passing through this
compartment. More of Ca ind K was lost through run-off and percolation due to
greater release of these nutrients through litter. The significance of these results have
been discussed in the context of secondary succession after slash and burn agriculture
(jhum) of which this forms a later stage of community development.

1. Introduction

Atmosphere is a source of chemical inputs to terrestrial ecosystems which come
through direct fall, throughfall and stemflow (Ovington 1959; Carlisle et al
1965) Generalization of these aspects are chiefly based on temperate forests
and studies on tropical forests are meagre (Jackson 1971). In India the few
studies done on nutrient flow through water are on plantations (Debral and
Subba Rao 1968 ; George 1978) and no work has been done on natural
forests.

The earlier two papers in this series (Part I and II) pertain to a 50 year old
forest fallow at lailad in Meghalaya and deal with phytosociology, biomass and
nutrient inventory and cycling through leaf and wood litter. The present study
deals with the pattern of water and nutrient circulation through stemfiow and
throughfall and quantity interception, run-off and percolation losses in this
forested ecosystem. For the present study, only the central zone of the forest
representing the undisturbed 50-year stand has been considered.

269

270 Jasbir Singh and P S Ramakrishnan

2. Methods of study

Steinflow was sampled using a spiral polythene gutter of 6 cm diameter fitted in
each stem and sealed with paraffin wax. The gutter was fixed at a height
of 1.5m above the soil surface on the tree trunk. A plastic funnel was
attached to the two cut ends of the gutter and connected to a polythene container
of 5 litre capacity. A nylon filter 1 'mm mesh size was placed in the mouth of the
funnel to prevent entry of extraneous matter. Three replicates each for two girth
classes namely 30 and 90 cm were selected for each of the following important
species : (1) Shorea robusta (ii) Schima Wallichii, (iii) Castanopsis indica,
(iv) Gmelina arborea and (v) Artocarpus chaplasha.

Water of throughfall and incident rainfall was collected in polythene containers,
the mouth of each being fitted with 20 cm diameter funnel which was provided
with 1 mm mesh nylon filter to prevent entry of foreign matter. Three containers
were kept outside the forest in open places to collect the water from incident rain.
Twelve containers of the same size were kept under the forest canopy to measure
the throughfall. In order to measure the atmospheric precipitation two standard
rain gauges were kept in the open. All the containers were kept 50 cm above
the surface on a wooden platform to avoid splashing of soil particles into the
funnels. 2 ml toluene was added to the container to prevent microbial activity.
Sampling was done at intervals ranging from 2 to 7 days depending upon the
intensity and the frequency of rainfall during the monsoon. At the time of
sampling, 500 ml of well homogenised water from stemflow/throughfall/incident
rain was brought to the laboratory and the samples were filtered through a
Whatman no. 44 filter paper and chemically analysed for N, P, K, Ca and Mg.

For studies pertaining to run-off water and sedimentation, the loss from a
confined area of 1x10 m along the slope was collected in drums of 200 litre
capacity and periodically removed for analysis. Percolation studies were done
using a simple lysimeter of the Russian type (Buckman and Brady 1980). The soil
was cut out vertically to expose the profile. A small tunnel was excavated at a
depth of 40 cm (this is the depth at which root density is high) and a collector of
30 X 30 X 15 cm was placed inside the tunnel. The water percolating through
soil was tapped out from time to time and chemically analysed for N, P, K, Ca
and Mg using standard methods as described by Jackson (1958) and Allen (1974).
Thus, nitrogen was determined by micro-kjeldahl method and phosphorus
estimated colorimetrically by molybdenum-method. Calcium and magnesium
were analysed by EDTA titration method and potassium by flame emission
method.

3. Results and discussion

3,1 Throughfall

A summary of the seasonal distribution (table 1) shows that the percentage
of rainwater coming as throughfall is maximum in the month of March- April
(over 70% of the total) while in other months it was *mich less with minimum

Nutrient flow through water

271

percentage in December. The high proportion of throughfall in March-April
was due to reduced crown density at this time when 41 % leaf fall occurred. The
proportion of throughfall showed a decreasing trend with increase in canopy
density in subsequent months. Similar relationship between crown density and
throughfall was also reported by Szabo (1975). The average value of 52.4%
throughfall during the course of this study was higher than the values obtained
for other broad leaved forests (Debral and Subba Rao 1968 ; Aldridge and
Jackson 1973) but lower than a few others (Ovington 1959; Leonard 1961;
Szabo 1975). The quantity of throughfall is directly proportional to the gross
rainfall and the relationship is indicated in figure la (P < *05).

3.2 Stemflow

The average stemflow during the period of study was measured at 8% of the
precipitation (table 1). A low proportion of stemflow was measured in the month
of November due to lesser intensity and frequency of rain. On the other hand,

100 ?00 )00

Gross rainfall I mm '

Figure 1. Relationship between gross rainfall and (a) throughfall,
(b) stemflow and (c) interception.

272 Jasbir Singh and P S Ramkrishnan

reduced canopy density in the month of March and April allowed more water flow
through stem. The gradual decline in percentage stem flow from May-October
could be related to the high canopy density which in turn resulted in greater
proportion of interception losses. The present value of stemflow (8%) was found
to be higher than the values reported by Ovington (1959) and Szabo (1975) which
ranged from 0.12—3.10% but lower than the values reported by a few others
(Eidmam 1959 ; Aldridge and Jackson 1973). The relationship between gross
rainfall and stemflow was found to be highly significant which is expressed by
a linear regression line (figure Ib).

3.3 Interception loss

The proportion of the rainfall intercepted by the canopy was inversely related to
the proportion of throughfall and stemflow with maximum percentage values
recorded in December and minimum in March. This high percentage of
water transmission from the canopy during October-December may be attributed
to the fact that rainfall in these months was not regular in comparison to the
monsoon period (April-September) and the vegetation often remained dry. Thus
the greater quantity of water necessary to wet the vegetation accounted for the
high interception loss. The low interception values during March and April were
due to reduced crown density due to maximum leaf fall during this period.
Similar observation with low interception losses during heavier leaf fall was
made by Szabo (1975) in their study on Hungarian Oak forest ecosystem,
The relationship between gross rainfall and interception loss which follow the
same pattern as stemflow and throughfall is shown in figure Ic.

3.4 Water balance of the ecosystem

The total amount of water and percentage values given in table 1 are estimates
based considering forest canopy as entirely closed. But in order to explain the
total quantity of water in one hectare forest surface, the forest gaps are also to be
considered. The total canopy coverage in the present forest was 90% and the
rest 10% was accounted for gaps where the precipitation could reach the forest
floor almost directly without any interruption. After making allowances (Szabo
1975) for gaps in the forest canopy, the percentage distribution of the incident
rainfall in a forest of one hectare is shown in figure 2a. However, it may be
mentioned that the stemflow values would be on the higher side as only selected
important tree species of same girth classess were considered and assumed
contribute to the total cover of the forest.

3.5 Loss through run-off and percolation

Studies on surface run-off and percolation losses of rainwater at Lailad forest
showed that they represented 19.6% and 6.78% respectively of the total annual

Nutrient flow through water

273

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274

Jasbir Singh and P S Ramakrishnan

A S 0 N 0
Months

Figures 2a-b. a. Distribution of precipitation in the forest (per hectare). Hatched
columns, throughfall ; closed columns, stemtlow ; open column, intercepted
loss. b. Monthly pattern of surface run-off and percolation loss of Lailad
forest.

rainfall, during the year. The monthly pattern of losses of water could be related
to the rainfall pattern with maximum run-off and percolation losses occurring
during May-September (with 87% and 37% of the total run-off and percolation
losses respectively), with peak values in the months of July (figure 2b). Both, the
high frequency and intensity of rainfall during monsoon contribute to heavy
losses during this period.

3.6 Nutrient return by throughfall, stemflow and incident rainfall

The mean monthly concentration (mg/1) of the various elements throughout the
study period in throughfall, stemflow and incident rainfalls are shown in
figures 3a, b, c, d, e, The * concentration of total nitrogen in throughfall and
stemflow was low during March-May, followed by a steady rise reaching a
maximum towards October-November which could be related to the subsequent
maturity of the leaves. ' Similar increase in N concentration of throughfall with
increasing leaf maturity was shown by Tukey et al (1958) with subsequent
decrease after the formation of abscission layer. The concentration of nitrogen
in stemflow was more than that of throughfall which may be due to (i) release

Nutrient flow through wafer

275

J A S 0 N 0
Months

Figure3. Monthly variation in concentration of different elements, a,
nitrogen ; b, phosphorus ; c, potassium ; d, calcium and e, magnesium
in different water samples.

of nutrient during bark decay during the rainy season (ii) the wash out of
nutrients from leaves and . (iii) low quantity of water in stemflow and consequent
high concentration of nutrients. Monthly variation in the concentration of N in
incident rainwater could also be observed though the values were low compared
to that of the throughfall and stemflow.

276 Jasbir Singh and P S Ramakrishnan

The concentration of phosphorus in incident rain was very low during the rainy
season. A slightly higher level was noted during March-April which could be
due to (i) high level of dust particles in the atmosphere during the preceding dry
period, (ii) presence of partly burnt particles of organic matter in the air due to
burning of slash in the neighbourhood during this period due to shifting agri-
culture and (iii) low rainfall during this period with lesser dilution of this
element. In throughfall., the level of P gradually increased reaching a maximum
in August followed by a slow decrease in subsequent months. The gradual
increase in concentration up to August may be attributed to the presence of high
pollen grains of the tree species and also the production of new leaves which may
have higher concentration of phosphorus as was reputed by Carlisle et al (1967)
working on a sessile oak forest in England. A similar pattern of monthly
concentration changes was also observed for stemflow except that the concentration
of P on the average was two times higher than that for throughfall due to release
of this nutrient from the decayed bark.

The monthly pattern for potassium, calcium and magnesium concentration in
throughfall, stemflow and incident rainfall was similar (figures 3c, d, e) . The
gradual increase in concentration from the month of May to December is probably
related to gradual maturation of new leaves produced in April and the consequent
increase in release of some of these nutrients from more mature leaves, an
observation also made by Tukey et al (1958) and Deneyear-DeSmet (1966).
Amongst the three elements, the concentration of Ca was more in stemflow due
to its high level in the bark.

The total amount of nutrients (kg/ha/yr) contributed through stemflow,
throughfall and incident rain is shown in table 2. Throughfall contributed 98%
of all the nutrients. The low addition through stemflow is in spite of the high
concentration of nutrients in the stemflow water, as the total quantity of stemflow
is far less than that due to throughfall. Amongst the cations, calcium and potassium
were highly leachable with heavy washout through stemflow and throughfall.

Table 2. Nutrient return through stemflow, throughfall and rainwater at Lailad forests.

Nutrients (kg/ha/yr)

N

P

K

Ca

Mg

Stemflow

0.17

0.02

0.71

0.84

0.10

Throughfall

8.39

0.89

31.28

35.19

5.03

Total 9.56

0.91

31.99

35.93

5.13

Rainwater

4.33

0.43

7.80

9.96

4.77

Difference 5.23

0.48

24.19

24.97

0.63

Nutrient flow through water

277

AMJ J ASONOJ

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in different water samples.

andmcideat rain was in the order

J.7 Nutrient losses* through run-off and percolation of water

berlateto greater release of nutrients! from freshly decomposing

278

Jasbir Singh and P S Ramakrishnan

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Nutrient flow through water 279

The concentration of nutrients in percolation water also followed a similar pattern
as that in the run-off water, in that peak values were higher during July-
September with a minor peak during March-April (figures 4a, b, c, d, e).

Since the present forest represented a secondary successional fallow after jhum
(Toky and Ramakrishnan unpublished) a comparison of the pattern of losses with
a 0-year freshly burnt site would be interesting (table 3). The loss of Ca and K
from a 50-year old fallow was higher compared to other nutrients may be due to
greater release of these nutrients from the vegetation through litter decomposition
(Timmons et al 1977). In fact even though the quantity of water lost from a
deforested side througK run-off and percolation was not more than 2-3 fold
compared to the Lailad forest, nutrient losses increased as much as 6-7 times
compared to the forested site. This may be due to sudden release of nutrients
through burnt and lack of vegetation cover to hold the same (Ramakrishnan
et al 1980).

4. Conclusion

The three series of papers on the ecosystem structure and function of a 50-year
old secondary successional fallow is significant as it represents a comparatively
more stable forested ecosystem in the successional gradient. An early successi-
onal weed stage which dominates up to about 5 years of fallow development after
jhum is soon replaced by a bamboo dominated stand up to about 20 years beyond
which dicot tree species gain importance (Ramakrishnan et al 1981). The
peripheral zone of the Lailad forest (Part I in this series) represents such a stage
due to frequent disturbances in this zone. The central zone, however, represents
the 50-year old stand of a mixed broad leaved forest* Apart from the high species
diversity and biomass and nutrient stored in the living compartment alongwith
efficient cycling through litter (Ramakrishnan et al 1980), the loss of nutrients
from the system is also minimal compared to younger fallows (Toky and
Ramakrishnan 1981). In fact there is negligible loss from the system through
both run-off and percolation as seen from comparison presented in Part III
in this series. This presented in a dramatic way the damage done due to
deforestation during shifting agriculture. With the climate and steep topographic
conditions prevailing in the north-eastern hill region, the present study highlights
the significance of maintaining a forested ecosystem for environmental
stability.

Acknowledgement

We are grateful to Dr O P Toky and Dr Ashesh Kumar Das for their help in
the preparation of manuscript. One of us (JS) was a recipient of Junior Research
Fellowship of the North-Eastern Hill University.

References

Aldridge R and Jackson R J 1973 Interception of rainfall by bardbeach (Nothofagus truncata) at

Taita, New Zealand. N.Z. J. ScL 16 185-198
Allen S E 1974 (ed) Chemical analysis of ecological material. (Oxford: Blackwell Scientific Publ)

pp 565

280 Jasbir Singh and P S Ramakrishnan

Buckman H O and Brady N C 1960 (eds) The nature and properties of Solids. (New York :

The MacMillan) pp. 567
Carlisle A, Brown A H F and White E J 1965 The interception of precipitation by oak (Quercus

petraea) a high rainfall site ; Q. J. For. 59 140-143
Carlisle A, Brown A H F and White E J 1967 The nutrient content of tree stemflow and ground

flora litter and leachates in a sessile oak (Quercus petraea) Woodland. /. Ecol. 55 615-662
Debral B C and Subba Rao B K 1968 Interception studies in chir and teak plantations— New

Forest. Indian For. 94 541-551
Denaeyer-DeSmet S 1966 Bilan annuel des apports d'elements mineraux par les eaux dc

precipitation lous convert, forestier dans la foret caducifoliee de blamont (Birelessachimay).

Butt. Soc. R. Bot. Belg. 99 345-375
Eidmann F E 1959 The interception in Buchan und Fichtebestenden, Ergelomis mohrajah riger

untersuchengen in Rothaurgebirge (Samel and) Inst. Assn. Scientific Hydrology Publ.

48 5-25
George M 1978 Interception, stemflow and throughfal] in a Eucalyptus hybrid plantation.

Indian For. 104 719-726

Jackson M L (ed) 1958 Soil chemical analysis (New Jersey : Prentice Hall, fnc.)
Jackson I J 1971 Problems of throughfall and interception assessment under tropical forests.

/. Hydro!. 12 234-254
Leonard R E 1961 Interception of Precipitation by northern hardwoods. North Eastern Forests

Expl. Station Paper No. 159 1-16
Ovington J D 1959 The circulation of minerals in plantations" of Pinus sylvestris L. Ann. Bot.

(NS) 23 229-239
Ramakrishnan P S, Toky O P, Mishra B K and Saxcna K G 1981 Slash and burn agriculture in

North-Eastern India In : Fire regimes and ecosystem properties (ed) H Mooney, TM

Bonnicksen, N L Christensen, J E Lotan and W A Reiners, USDA Forest Service general

technical report WO 26, pp. 570-587
Szabo M 1975 Net precipitation under a forest (Quercetum petraea Gerris.) Canopy in 1974

Acta. Bot. Acad. Sci. Hung., 21 151-165
Toky O P and Ramakrishnan PS 1981 Run-off and infiltration losses related to shifting

agriculture (Jhum) in north-eastern India. Environmental conservation 8 313-321
Timmons D R , Very N S, Burwell R E and Hold R F 1977 Nutrient transport in surface run-off

and stemflow from an Aspen-Birch forest. Our Environ. Qual 6 188-192
TukeyJBJr, Tukey H B and Wittwer SH 1958 Loss of nutrients by foliage leaching as

determined by radioisotopes. Proc. Am. Soc. Hortic. Sci. 71 496-506

Proc. Indian Acad. Sci. (Plant Sci.), Vol. 91, Number 4, August 1982, pp. 281-237.
© Printed in India.

Anatomy of the seedling of the Legnminosae

UMAVATHI HEGDE and V D TILAK*

Department of Botany, KG College, Maharshi Karve Road, Bombay 400 020,

India

*Department of Botany, The Institute of Science, Madam Cama Road,

Bombay 400032, India

MS received 10 February 1981 ; revised 22 January 1982

Abstract. Anatomy of the juvenile nodes m the seedling of fourteen genera of
Legumtnosae is described. The cotyledonary node exhibits a two-trace, urailacunar
condition. The two traces show various degrees of approximation of the two traces
leading to one-trace, unilacunar condition in some genera. At the second node
level, only two genera show the one-trace, unilacunar pattern ; 11 of the remaining
genera have a three-trace trilacunar supply and one genus exhibits an intermediate
type. At the third node level all the genera show a three-trace, trilacunar pattern.
The present data suggest that the three-trace, trilacunar condition is derived by the
addition of a lateral trace on either side of the median trace and that the one-
trace, unilacunar condition appears to be a result of the approximation of the two
traces at the cotyledonary node.

Keywords. Nodal anatomy ; seedlings ; Lcguminosae.

1. Introduction

Anatomy of the seedling in the Leguminosae has been studied by Compton (1912),
Winter (1932), McMurray and Fisk (1936), Weaver (1960), Pillai and Sukumaran
(1969), Pillai et al (1970, 1974), Bairathi and Nathawat (1974) and Narang (1978).
Most of these studies are, however, restricted to the cotyledonary node and
root-stem transition region. The evolutionary sequence followed by the diffe-
rent types of nodal structures has always been of interest to plant anatomists.
The present investigation was, therefore, undertaken to find out if the anatomical
studies in the seedling of some dicots would be of help in this regard. For this
purpose the anatomy of the juvenile nodes of the seedling of 14 species belonging
to 14 genera of the Leguminosae were studied.

2. Materials and methods

Seedlings of the plants studied were raised in the garden of .the Institute of Science,
Bombay, from seeds collected locally or purchased in the market. Portions of the

281

282 Umavathi Hegde and V D Tilak

axis obtained by cutting a little below and above the first (cotyledonary), second
and the third nodes were fixed in FAA. The usual paraffin method was followed.
Sections of the paraffin infiltrated material were cut on a microtome at a thickness
of 15-20 microns. The sewere stained with crystal/gentian violet using erythrosin/
orange G as counter strain

3. Observations

Crotalaria juncea Linn. The hypocotyledonary axis has a ring of about a dozen
bundles (figure 1), Two traces depart from each of the two sides (figure 2). The
two traces corne closer to each other in their upward course but retain their sepa-
rate entity at the cotyledonary base (figure 3). Axillary buds are present in the
axil of the cotyledons (figure 3).

The epicotyledonary axis has a ring of about 15-20 bundles (figure 4). Three
traces, one median and a lateral on either side of it, emerge from the ring (figure 5),
The laterals in their upward course shift closer to the median and can hardly be
differentiated from each other in the petiole (figure 6).

The third node has, likewise, a three trace, trilacunar supply to. the leaf (figure 7).

The laterals after giving off a branch to the stipule shift closer to the median and
almost merge with it so as to become indistinguishable (figures 8-9). As a varia-
tion, some series cut off at the second and third nodes exhibit a four-trace, four
lacunar condition i.e. a median with two laterals on one side and one on the other
(figures 10-11).

The nodal structure at the first three nodes in Medicago sativa Linn., Tephorosia
purpurea (Linn.) Pers., Cassia tor a Linn, and Mimosa pudica Linn, is similar to
Crotalaria juncea except for minor differences regarding the degree of distinctiveness
of the two traces to the cotyledons as they extend upwards and that of the lateral
traces of the leaves of the second and third nodes from the median trace, as these
extend upwards into the petiole. In Mimosa pudica the cotyledons separate out
from the axis in the form of a ring which organizes into two cotyledons, and two
non-vascularized stipular structures (figure 12).

In Arachis hypogaea Linn., the cotyledonary node is two traced unilacunar with
the two traces remaining distinct in the cotyledons (figures 13-14). The leaves at
the second and the; third nodes are three-traced and trilacunar (figure 1 5) with the
laterals as well. as the median bundles dividing in their upward course (figure 16).
Some of the branches of the laterals extend into the stipules while the remaining
ones alongwith the branches of the median and the median itself, extend into the
petiole '(figure, 17).

The cotyiedonary node in Pisum sativum Linn, is two-traced, xmilacunar, traces
remaining distinct from one another in their upward course (figures 18-19). Tho
leaves of the second and the third node, are three-traced and trilacunar. This
plant is characterized by the precocious emergence of the lateral traces as compared
to the median trace of the same leaf. Thus, the lateral » tcaces of tb@, teaf of tbl?
second node emerge a little above the cotyledonary node and those of the leaves
of the third node emerge almost . alongwith the median bra.dk of the second ?Bodte
(figure 20)* These precociously emerged latoai traces, extend outwards from the

Anatomy of the seedling of the Legwninogae~I 283

vascular ring (figures 21 and 22). The median bundle and the lateral traces divide
in their upward course (figures 21-23). While some of the branches of the laterals
extend into the stipules, the remaining ones alongwith the branches of the median
and the median itself, extend into the petiole (figures 22,24).

The cotyledonary node in Clitoria ternatea Linn, is two-traced, unilacunar.
The two traces come together in their upward course. They ultimately lose their
distincti veness. The leaves of the second node are opposite in phyllotaxy. Each
leaf receives a single median trace and a common lateral only on one side of the
axis (figure 25). The single common lateral splits into two (figure 26). The
branches of the split lateral, after giving off branches to the stipule, shift closer
to the median in their upward course and ultimately merge with it forming a con-
centric bundle (figures 26-27). The stipules on the other side of the leaves receive
their vascular supply from the median bundle itself (figure 26). The phyllotaxy
at the third node is alternate. The leaves at this node receive a three-trace vas-
cular supply. The lateral traces, as usual, in their upward course, at first, give off
a branch to the stipule and later come closer to the median and merge with it.

The cotyledonary node in Abrus precatorins Linn, and Cicer arietinum Linn,
is two-traced, unilacunar. The two traces come closer to each other in their
upward course. The leaves of the second node are one-traced, unilacunar (figures
28,30). The single trace splits into many small strands, upwards (figure 29). The
leaf of the third node, is as usual three-traced, trilacunar.

The cotyledons in Leucaena glauca Linn, and Albizia kbbeck (Linn.) Benth
receive two traces, which in their upward course merge into one and divide into
three at the point of entry into the cotyledonary base (figures 31,32,33). The
cotyledons in the Tamarindus indica Linn, and Pithecellobium dulce, (Roxb.) Benth
are characterized by one-trace, unilacunar supply (figure 34). This single trace
divides into three right at the point of emergence (figures 35-36). The vascular
supply to the leaves of the second and the third nodes in the above mentioned plants
is of the three trace, trilacunar type with the lateral traces in their upward course,
at first giving off branches to the stipules and then shifting closer to median.

4. Discussion

Anatomists have treated different types of nodal structures in different ways, phylo-
genetically. Thus, Sinnott and Bailey (1914) suggest that the three-trace, trila-
cunar type is primitive as compared to unilacunar and multilacunar types. Ozenda
(1949) on the other hand treats the multilacunar type as more primitive. Marsden
and Bailey (1955) describe the so-called " fourth type " i.e., a two trace, unilacunar
type as the primitive type of node. They, further, consider that the unilacunar
node with one trace arose by the fusion of two traces. Bailey (1956) reexamined
the earlier viewpoint which he shared with Sinnott (Sinnott and Bailey 1914) and
reached a conclusion that the three-trace, trilacunar and multitrace, multilacunar
types in angiosperms were derived at some stage in their evolution from the one-
trace, unilacunar type. Canright (1955), Eames (1961), Esau (1960,1965) and
Carlquist (1961) have more strongly emphasized the primitiveness of the unila-
cunar node with two traces and consider it as basic in the evolution of angiosperm
nodal structure.

284

Umavatki Hegde and V D Tilak

Anatomy of the seedlin g of the Legummosae-l

285

.w Mfiil !

286 Umavathi Hegde and V D Tilak

Pant and Mehra (1964) and Benzing (1967) do not treat the two-trace, unilacunar
node as primitive. Instead, Benzing (1967) proposes that the three-traced condi-
tion is likely to be more primitive in the angiosperms than the two-trace, unilacunar
type. Takhtajan (1969) conceives of a hypothetical tc fifth type " of node with a
central gap haying two traces. This type is supposed to have given rise to all
other known types.

All the plants studied show a basically two traced, unilacunar nodal pattern at
the cotyledonary node. Some of these have a two-traced, unilacunar structure*
with varying degrees of independence or approximation of the two traces from the
point of emergence through their extention into the cotyledons. Other plants
have a one-traced, unilacunar type. Thus, there is a trend towards the approxi-
mation of the two traces leading to a one trace, unilacunar type of node.

At the second node level Abrus precatorius and Cicer arietinum have a one-trace
unilacunar condition. Clitoria ternatea presents an intermediate condition. It has
a median trace, each, for the two leaves of the opposite phyllotaxy and a common
lateral on one side of the axis. All the remaining plants have a three-trace, trila-
cunar supply to the leaves of the second node. Further, all the plants studied have
a three-trace, trilacunar structure at the third node. Thus, it is only in Abnis
precatorius and Cicer arietinum that the three trace, trilacunar pattern is attained
at the third node. Rest of the plants achieve this condition at the second node
itself.

The present data, therefore, shows that the three-trace, trilacunar condition
is a characteristic feature of the more mature second and third nodes while the
two-trace, unilacunar and one trace, unilacunar patterns are seen at the cotyledo-
nary nodes. Similar observations have been made during the course of a study of
- twentytwo genera of the Malvales (Rao 1980). Further, in no plant either of the
Leguminosae or of the Malvales could a condition be recorded wherein a three-
trace, trilaeunar condition of the cotyledonary or second node is succeeded by a
one-trace, unilacunar type at the second and third nodes. Bailey (1956) records
similar observations. These observations may be taken as an evidence in support
of the? primitiveness of the one trace, unilacunar type as compared to the three-trace,
trilacunar type.

In Abrus precatorius the single trace at the second node divides into many
small strands. Ifl Clitoria ternatea the branches of the split lateral give off a branch
to the stipule and shift closer to the median and merge with it. The stipule on the
other side receives its vascular supply from the median itself. In Arachis hypogaea
and Pisum sativum the median as well as the laterals divide into smaller strands.
While some of the branches of the laterals enter into the stipule, the remaining
ones alongwith the .branches of the median and the median itself extend into the
petiokf. In rest of the plants both at the second as well as the third nodes the
laterals at first give off a branch to the stipule and their remaining portions shift
closer to the median. These portions of the laterals may remain distinct from the
median in plants such as Medicago sativa and Tephrosia purpurea or merge with the
median as in Crotalaria juncea and Cassia tor a. Thus, the laterals, give vascular
supply to the stipules, however, when absent, the stipules receive branches from
the median itself.

Anatomy of the seedling of the Legumino$ae-I 287

Vascular bundles outside the main vascular ring have been recorded in the inter
node ofPisum sativum by Kuptcha (1975). He designates these as cortical bundles
but they appear to be precociously emerged lateral traces,

Acknowledgements

The authors sincerely thank Dr B C Haldar, Director, The Institute of Science
for laboratory facilities and Prof. R M Pai, Professor and Head of the Botany
Department, Marathwada University, Aurangabad for helpful suggestions and
keen interest. They thank Shri A M Siddiqui for his help in the preparation
of the illustrations.

References

Ba'ley I W 1956 Nodal anatomy in retrospect ; /. Arnold Arbor. Harv. Univ. 37 269-287
Bairathi H K and Nathawat G S 1974 Morphology and anatomy of polycotylous and twin

seedlings of Crotalaria juncea Linn ; /. Indian Bot. Soc. 53 196-201
Benztng DM 1967 Developmental patterns in stem primary xylem of woody Ranales ; Am. J.

Bot. 54 805-820
Ca'inght J E 1955 The comparative morphology and relationships of the Magnoliaceac IV. Wood

and nodal anatomy ; /. Arnold- Arbor. Harv. Univ. 36 1-50

Carlquist S 1961 Comparative Plant Anatomy (New York: Holt Reinhart and Winston)
Compton R H 1912 An investigation of the seedling structure in the Leguminosae ; /. Linn.

Bot. 41 1-122
Eamcs A J 1961 Morphology of Angiospenm (New York, Toronto and London : McGiaw-Hill

Co. Inc.)

Esau K 1960 Anatomy of the seed plants (New York and London : John Wiley and Sons, Inc.)
Esau 1965 Plant anatomy (2nd ed.) (New York amd London : John Wiley and Sons. Inc.)
Kupicha F K 1975 Observations on the vascular anatomy of the tribe Yicicae (Leguminosae) ;

Bot. J. Linn. Soc. 74 131-162
Marsden H P E and Bailey I W 1955 A fourth type of nodal anatomy in dicotyledons

illustrated by Clerodendron trichotomum Thumb. ; /. Arnold Arbon. Harv. Univ. 30 1-50
McMurray E B and Fisk E L 1936 Vascular anatomy of the seedling of Melilotus alba \ Bot.

Gaz. 98 121-134
Narang Arvind Kaur 1978 Seedling structure in Crotalaria and Tephrosia species ; J. Indian

Bot. Soc. 57 52-57
Ozenda P 1949 Recherches aur les Dicotyledones apocaspiques contribution aletude des Angiospermes

dites primitives Paris Publ. Las. Ecde. Norm. Sap.ser biol. II

Pant D D and Mehra B 1964 Nodal anatomy in Retrospect ; Phytomorphology. 14 384- 357
Pillai S K and Sukumaran K 1969 Histogenesis, apical meristem and anatomy of Cyamopsis

tetragonoloba (Linn) Taub, Phytomorphology ', 19 303-312
Piilai S K, Trivcdi M L ard Abraham K 1970 Certain aspects of the anatomy of Crotolaria

burhia Ham. ; /. Birla Inst. Technol. Sci. 2 153-164
Pillai S K, Ramasita K and Ishwar Dutt 1974 Embryology, histogenesis, apical meristem, seed

coat and seedling anatomy of Albizia lebbeck (Lirux) Benth. ; New Botany 1 23- 33
Rao Gmja K 1980 Anatomical studies in the seedling of some dicotyledons II Ph.D. thesis

University of Bombay
Sinnott E W and Bailey I W 1914 Investigations on the phyjogeny of angiosperms I. The

anatomy of the node as an aid in the classification of the ang'osperms ; Am. J. Bot. 1,

303- 332

Takktajan A 1969 Flowering plants— origin and dispersal (Edinburgh: Oliver and Boyd)
Weaver H L 1960 Vascular'zation of the root hypocotyl-cotyledon axis of Glydne max L ;

Phytomorphology. 10 82- 86
Winter C W 1932 Vascular system of young plants of Medicago sativa L.; Bot. Gaz. 94 156-16.

Proc. Indian Acad. Sci. (Plant Sci.), Vol. 01, Number 4, Aufiust 1982, pp. 289-195.
© Printed in India.

Cork-warts in Eucalyptus species

PARVEEN FAROOQUI (nee KIDWAI*) *
Department of Botany, The University, Allahabad. India
* Regional Forest Research Centre, Jabalpur 4#2 020, India

MS received 26 December 1980 ; revised 24 May 1932

Abstract. Four different types of cork-warts, the ordinary cork-warts, the d-scars,
wound cork and trichome scars have been described from E. citriodora and
E. torcllina. The differences between them are discussed.

Keywords. Eucalyptus ; cork-warts ; d-scars ; wound cork ; trichome scars ; d-
stomata.

1. Introduction

The occurrence of cork-warts or holes in the epidermis surrounded by radially
arranged concentric rows of suberised cells has been mentioned among others by
Solereder (1908), Metcalfe and Chalk (1950) and Stace (1965). Their presence has
been explained either as a diagnostic character or as a result of mechanical injuries.
During a general survey of epidermal characters of a number of Eucalyptus species,
cork-warts were noticed in some of them. As no detailed accounts of their struc-
ture and development is available it was thought worthwhile to investigate them
in detail.

2. Material and methods

Young and mature leaves of E. citriodora Hook, and E. torcllina F.VM. were obtained
from Tamil Nadu. Peels of the epidermis were obtained by scrapping and those
of the cuticle by maceration in a 1 : 1 mixture of nitric acid and chromic acid
(both 10%). The peels were mounted in Safranin glycerine jelly. Transparencies
of the leaves were also prepared by the method of Arnott (1959).

3. Observations

In both species, numerous black or brown spots are scattered over the upper and
lower epidermis (figures 1,12,13). A closer examination of these spots and study
of their early developmental stages show four distinct types of structures.

289
P.(B)~2

290 Parveen Farooqui (nee Kidwai)

3.1. d-stomata and d-scars

In the young epidermis, alongwith developmental stages of stomata, hairs etc., a
few large fully formed stomata are also present. They persist in the mature leaf
and occur isolated between groups of normal sized stomata (figure 2). Some
of these large stomata become darker in colour and a dark zone is demarcated
around them in the ordinary epidermal cells (figures 3,9). Others
persist as large stomata, without further development. The stomata with a dark
zone develop further. The surrounding cells divide concentrically around the
stoma (figure 4). The pore of the stoma enlarges and becomes thin walled
(figure 4). The guard cells appear to lose their contents. Finally the guard cells
are ruptured and lost and the surrounding cells become thick-walled and darker.
Similar stomata have been described in Ilex species by Korn and Fredrick (1973)
and have been named d-stomata (or developmentally important stomata) and the
replacement periderm tissue is termed the d-scar.

3 . 2. Ordinary cork-warts

Intermixed between the d-stomata and d-scars are other structures with similar
concentric rings of periderm like tissue which enclose either a single ordinary sized
stoma, or several ordinary stomata (figures 5,10) or even ordinary epidermal cells
without stomata. After the thickening of the walls of the periderm like tissue the
enclosed cells break down and disappear (figure 8).

3.3. Wound cork

Occassionally the epidermis also shows rounded or slit like gaps in it which appear
like punctures by insects or accidental wounds respectively. Later a periderm
like activity is seen around them (figure 6), the cells finally becoming thick walled
and cork-like with the hole in the centre.

3 . 4. Trichome scars

Young leaves of both these species show multicellular, multiseriate hairs or emer-
gences with a broad multiceliular base. In E. torellina, some of these trichomes
persist even on mature leaves especially over the margins and veins. When the
hair falls its base is surrounded by the thick-walled, somewhat papillate cells
(figure 7). A periderm like tissue develops around the hair base. These form the
tricliome scars (figure 11).

4. Discussion

Cork-warts have been mentioned by Solereder (1908), Haberlandt (1914),
Metcalfe and Chalk (1950) and others. Stace (1965) was perhaps the first to sus-
pect their composite nature and described them under " cork-warts and similar
structures ". The function of the cork-warts has invariably been attributed as

Cork-warts in Eucalyptus species

291

Figure 1-7. 1. £". torellina : Law power view of the leaf surface showing
distribution of cork warts (x 20). 2. E. torellina : Large d-stoma surrounded by
normal sized stomata f x 340). 3. E. citriodora: d-stoma showing a zone of darker
area around it (x 340). 4. E. citriodora : d-stoma and d-scar (x 340).
5. £'. torellina: Cork-wart showing three stomata within it (xl75). 6. E. torellina:
hole, presumably an insect puncture showing beginning of formation of wound-
cork around it (x 340). 7. E. torellina : Hair base with papillate cells and
beginning of periderm in lower right hand corner (x 340).

292

Parveen Farooqui (nee Kidwal)

mechanical and to heal and repair holes caused by insect punctures or mechanical
injuries. Stace (1965) has observed that cork-warts are not abundunt in nature and
that they are present in certain species irrespective of wounds or insect punctures
and, therefore, are of diagnostic value. He considers them as distinct from those
caused by mechanical or insect injury.

d-stomata and d-scars have been reported only from the leaves of some species
of Ilex (Korn and Fredrick 1973). They are formed around large stomata which
can be distinguished from the early developmental stages of the leaf. In some
mangrove genera, Stace (1965) had earlier mentioned cork formation around
some large stomata, which he suspected to be water stomata. As far as I am aware,
they have not been reported from any other plant. Hence, their presence in species
of Eucalyptus is interesting.

The large d-stomata in the young epidermis, and the few that persist in the
mature condition without the formation of a periderm like tissue, appear very
similar to the giant stomata reported in a number of plants (Stace 1965 ; Sitholey
and Pandey 1971 ; Farooqui 1979 and others). It is possible, that such giant
stomata at a later stage form a d-scar around them. It may be worthwhile to rein-
vestigate plants with giant stomata from such a stand point.

According to Korn and Fredrick (1973), the d-stomata produce a zone of inhib-
ition that prevents additional d-stomata from forming during subsequent growth.
Their scattered distribution and absence of any contiguous d-stomata either in
Ilex or Eucalyptus support,- such an interpretation.

Wound cork formed as a result of injury may also be distinguished into two
types. As pointed out by Stace (1965), purely mechanical accidental wounds may
be variously shaped but are usually long and scar like. On the other hand
insect punctures are usually more or less rounded. The size of the insect punctures
may sometime indicate the identity of the insect visitor as it will correspond to the
size of the stylet of the insect.

The formation of a trichome scar after the falling out of the trichome is also
interesting. Usually hair fall off without leaving any mark on the epidermis or
their bases are left as such or they may become slightly thick walled. Here,
periderm like tissue is formed around the trichome base.

In the mature condition, when the inner plug of tissue is completely formed
or has fallen off, the four types of structures described here, appear very similar
to each other (figures 12, 13). However, developmentally they are different and
may be distinguished on the following basis :

d-stoma and
d-scar

(a) d-stoma is

formed early in
development and
later forms the
d-scar. The posi-
tion of the d-scar
is therefore, pre-
determined.

Cork-wart

Formed towards
the beginning of
leaf maturation.
Position random.

Wound cork

Formed when
injury occurs.
Position deter-
mined by site of
injury.

trichome scar

Formed towards
•.leaf maturation.
Position deter-
mined by site of
the falling tri-
chome.

Cork-warts in Eucalyptus species

293

Figures 8-13. E. citriodora : Cork-wart with the central plug fallen (X 400). 9.
E. citriodom: d-stoma showing formation of a dark zone around it (x 400). 10.
E. citriodom : cork-wart showing four stonnta within it ( x 500). 11. E.
torellina : hair scar(X 10DO). 12, 13. E. torellina: cork-warts in different stages
of development (x600),

.Cork-warts in Eucalyptus species

295

) Only one large
stoma is involved.

) Cork forms
around stoma
first, stoma
breaks up later.

One, several or
no stomata are
involved. It may
include only
epidermal cells.

Cork formed
first and tissue
internal to it
breaks up later.

Tissue involved
is determined by
the size of the
wound.

Break up or
injury occurs
first and cork
formed later.

Only one tri-
chome base is
involved.

Trichome falls
off first. Cork
is formed later.
However, some
basal cells of the
trichome are
already thick-
walled.

.) May be of
diagnostic value
as they have
been reported in
rare cases only.

According to
Stace (1965) of
diagnostic value.

Of no diagnostic
value as they
are purely acci-
dental.

Falling hairs
leaving scars in
the epidermis
may be of diag-
nostic value.

The present study has clearly shown four distinct types of cork-warts. It is
:erefore, important that in all future studies where cork-warts are found, their
jvelopmental stages should also be studied in order to assign them to the correct
pe.

cknowledgements

hanks are due to Professor D D Pant, Head of the Botany Depart-
ent, Allahabad University for facilities and Shri K K Rao for help in
lotography.

eferences

rnott H J 1959 Leaf clearings ; Turtox News 39 192-194

irooqui P 1979 On the occurrence of abnormal stomata in plants; Curr. Sci.AS 841-849

aberlandt G 1974 Physiological Plant Anatomy Translated from the fourth German edition by

M Drummond, (London : Macmillan and Co.)
orn R W and Fredrick G W 1973 Development of D-type stomata in the leaves of Hex

crenata var. Convexa , Ann. Sot. 37 647-656

:etcalfe C R and Chalk L 1950 Anatomy of the Dicotyledons (Oxford : Clarendon Press,)
tholey R V and Pandey Y N 1971 Giant stomata , Ann. Sot. 35 641
rtereder H 1908 Systematic anatomy of the dicotyledons (Translated by L A Boodle and F E

Fritsch revised D H Scott ; Oxford)
ace C A 1965 Cuticular studies as an aid to plant taxonomy ; Bull Br. Mus. (N.H.) 4 1-78

3?roc.tidiati Acad. Sci. (Plant Scl), Vol.91, Nfutaber 4, August 1982, pp. 297-301.
© Printed in India.

Pericarpial sclereids in some Mimosaceae

S RANGAIAH, I L KOTHARI and G L SHAH

Department of Biosciences, Sardar Patel University, Vallabh Vidyanagar 388 120,
India

MS received 17 June 1981 ; revised 17 May 1982

Abstract. Structure, ontogeny and distribution of sclcrcids in the pericarp of seven
species of Mimosaceae are studied. Their occurrence is recorded in the epidermis
hypodermis, mesocarp and endocarp. They may be macrosclereids or brachy-
sclereids. Their structure and ontogeny are described and their taxonomic utility
is pointed out.

Keywords. Pericarp ; sclcreids ; Mimosaceae.

1. Introduction

Extensive Itierature is available on foliar sclereids (Metcalfe and Chalk 1979 ;
Rao 1951, 1980 ; Rao and Bhupal 1973 ; Rao and Bhattacharya 1978 ; Rao and
Das 1979), but it is very meagre on the occurrence and ontogeny of pericarpial scle-
reids (Gupta and Lamba 1981), especially in Leguminoseae (Halliburton et al 1975).
In this paper, therefore, an account of the distribution, structure and ontogeny of
sclereids in the pericarp of seven species belonging to five genera of Mimosaceae
is given and their use to delimit the investigated taxa is also indicated.

2. Materials and methods

Fresh fruits of Acacia auriculiformis Car., A. pennata Willd., A. torta Craib, Albizia
lebbeck Bth., Calliandra sp., Pithecellobium dulce Bth. and Samanea saman Merr.
were collected at different stages of their development from the University Bota-
nical Garden. The materials were processed through the usual procedures after
fixing in FAA. Voucher specimens are deposited in the Herbarium of Sardar Patel
University. Materials were macerated in Jane's fluid (Jane 1956). Histochemical
localization of lignin was done by Phloroglucinol-HCL method (Johansen 1940).

3. Observations

Sclereids occur at various places in the mature pericarp and are found scattered
among parenchyma. They are recognized by lignified cell walls, orderly pitting
and size. Their distribution is characteristic in the different species understudy.

297

298 S Aangata/i, /• L Kothari 'and G L Shah

There are two categories of sclereids in the investigated taxa : (a) non-idioblastic
tissue forming sclereid layers and (b) idioblastic sclereids. In all the investigated
taxa non-idioblastic tissue is present. In Acacia pennata and A. torta it consti-
tutes 2-3 hypodermal layers (figures 3,4, and 6) whereas in Calliandra almost an
equal number of layers occur below a hypodermal layer (figure 7). One to three
layers of sclereids are present in the endocarp of Albizia, Pithecellobium and
Samanea (figures 9-11). Idioblastic sclereids are present among the parenchyma
of mesocarp in small or large groups in A. aurlculiformis (figures 1,2) or isolated
and scattered in A. torta (figure 6 at arrow). Besides isolated sclereids they occur
in groups of 2-3 in the outer epidermis in the .latter species (figure 5 at arrow).
In Calliandra they are also present in small patches at the junction of sutural region
and the valve.

Sclereids vary in shape. Brachysclereids, which are more or less isodiametric
(figures 12, 16 at arrow), are more frequent than the other types of sclereids.
Other types of sclereids observed are the macrosclereids, which are rod-shaped
(figures 12 at dart, 13 at arrow, 14), fusiform, which are broader at the centre and
gradually taper at both ends (figure 16 at dart), spindle-shaped, which are also
broader at the centre but abruptly taper at the ends (figure 15 at arrow) and
kidney-shaped (figure 15 at dart),

In transection they may be spherical (figures 1,4,5,8), oval (figure 1), angular
(figure 7) or columnar and palisade like (figures 9-1 1). Sclereids have thick
lignified and stratified walls with either broad or narrow lumen. In the endocarpic
sclereids, however, the lumen is very much reduced and slit-like (figures 9-11).
Sometimes it is obliterated (figure 2). Usually mature sclereids are occluded with
tanniferous contents, as evidenced by ferric chloride staining. However, tanni-
ferous contents are absent or very rare in the endocarpic sclereids (figures 9-1 1).
Pit canals are visible in the epidermal, hypodermal and mesocarpial sclereids
(figures 1,3-8) but not in the endocarpial ones (figures 9-11). They may be simple
(figures 1,3,4) or branched (figure 8 at arrow).

Differentiation of sclereids is evident at the mature stage of fruit development.
In early stages a sclereid initial is distinguished by its size larger than the adjoining
cells, dense cytoplasm and prominent nucleus with a single nucleolus (figure 1 at
arrow). In the course pf differentiation the cell wall stratification increases with
the deposition of lignin. Consequently, the lumen size is reduced. Finally the
nucleus and the cytoplasm are autolyzed. In all the species, except endocarpic
sclereids in Albizia, Pithecellobium and Samanea, pit canals become prominent
and the lumen is filled up with tanniferous contents. Further mature sclereids
are devoid of cytoplasm and nucleus.

4. Discussion ..-.-.-.,...

As far as the authors are aware this is the first account of the presence of sclereids
in the pericarp of Mimosaceae (Metcalfe and Chalk 1979) though endocarpic

Figures 1-8. All transactions of fruits showing sclereids. 1, 2. Acacia auriculi-
formis. 3, 4. A. pennata. 5, 6. A. torta. 7, 8. Calliandra sp. (1,3,4,8 X
1080 ; 2, 5, 7 x 270 ; 6 x-430).

Pericarpial sclereids in some mimosaceae

299

309

S Rangaiah, f L Kothari and G L Shah

Figures 9-11. Transactions of fruits. 12-16. Macerated sclereids of fruits.
9. Albizia lebbeck. 10. Pithecellobium duke. 11. Sanwnea saman 12,16.
Acacia auriculiformis. 13-15. Calliandra sp. (9-11x270; .12, 13, 15x230;
14 x 340 ; 16 x 75).

Pericarplal sclefeicls! in some niimosacede 30 1

sclereids of three types -vermisosclereid (worm-like), calceusosclereids (boot-like)
and cruxosclereids (cross or T-like), are reported in Arachis of the Papilionaceae
(Halliburton et al 1975). The only other detailed report of pericarpial sclereids
is that of Gupta and Lamba (1981). They have accounted for the structure of
sclereids in the endocarp of Rauvolfia serpentina. In the species investigated by
us we have observed them in the epidermis of Acacia torta, hypodermal layers of
A. pennata and A. torta, and in the mesocarp of A. auriculiformis. In Calliandra
sp. sclereids occur at the junction of the sutural region and the valve in addition
to subhypodermal layers. Endocarpic sclereids occur in Albizia, Pithecellobiwn
and Samanea.

The position and type of sclereids can also be useful to delineate the taxa studied
as follows :

Sclereids only non-idioblastic :

Only brachysclereids present :

Sclereids endocarpial * Albizia, Samanea

Sclereids hypodermal Acacia pennata

Sclereids fusiform Pithecellobium

Sclereids idioblastic in addition to non-idioblastic ones :

Sciereids mesocarpial Acacia auriculiformis

Sclereids solitary or in

groups of 2-3 in the epidermis Acacia torta

Sclereids below a hypodermal layer * . * Calliandra

Acknowledgement

One of the authors (SR) thanks the Hoc for the award of a Teacher Fellowship.

References

Gupta V and Lamba L C 1981 Sclereids in the endocarp of Aauvolfia serpeiitina (L.) Benth. e#

Kurz. ; Proc. Indian Acad. Sci. 90 79-84
Halliburton B W, Glasser W G and Byrne J M 1975 An anatomical study of the pericarp

of Arachis hypogaea with special emphasis on sclereid component ; Sot. Gaz. 136 219-223
Jane F W 1956 The structure of wood (New York : Macmillan Company)
Johansen D A 1940 Plant Microtechnique (London : McGraw-Hill)
Metcalfe C R and Chalk L 1979 Anatomy of the Dicotyledons Vol. I, 2nd (ed.) (Oxford :

Clarendon Press)

Rao T A 1951 Studies on foliar sclereids. A preliminary survey ; J. Indian Bot. Soc. 30 2&-S9
Rao T A 1980 Aspects and prospects of foliar sclereids in angiosperms. Sym. Vol. : Current

trends in Botanical research, (eds.) M Nagaraj and C P Mallik, Kalyani Publications,

pp. 67-72
Rao T A and Bhattachary J 1978 A review on foliar sclereids in angiosperms ; Bull. Bot. Surv.

India 20 91-99

Rao T A and Bhupal O P 1973 Typology of sclereids ; Proc. Indian Acad. Sci. B77 41~55
Rao T A and Silpi Das 1979 Leaf sclereids— occurrence and distribution in the angio?perms.

Bot. Notiser 132 319-324

P.(B)~3

Proc. Indian Acad. Sci. (Plant Sci,), Vol. 91, Number 4, August 1982, pp. 303-308
® Printed in India.

Viability and infectivity of zoospores of Sclerospora graminicola
(Sacc.) Schroet in the soil

C R RAMESH* and K M SAFEEULLA

Downy Mildew Research Laboratory, University of Mysore, Mysore 570 006, Indja
* Scientist, Central Plantation Crops Research Institute, Kasargod 670 124, Icdia

MS received 24 July 19S1 ; revised 7 July 1982

Abstract. In the present study an attempt has been made to establish the fate of
sporangia of Sclerospora graminicola (Sacc.) Schroet deposited in the soil. A
technique has been standardised to demonstrate the germination of sporangia and
the viability and infectivity of zoospores in the soil under laboratory conditions.
For how long the zoospores remain motile in the soil is one of the many unanswered
questions in the zoospore biology. From the present study it is seen that, the
sporangia can germinate in the soil and liberate zoospores. The zoospores can
move against gravity, remain viable and infective for 5 hrs in the soil. Survival of
zoospores in the soil indicated that, they may serve as a potential secondary
source of inoculum through soil under field conditions.

Keywords. Sclerospora graminicola', pearl millet ; zcospoe ; soil ; green egr
disease.

1. Introduction

Data have accumulated in evidence of zoospore serving as inoculum through
water currents in soil-borne plant pathogenic phycomycetes like Phytophthora
(Bewley and Buddin 1921 ; Klotz et al 1949 ; Mehrotra 1961 ; Nolla 1928 ;
Mclntosh 1964 ; Zentmyer and Richards 1952). No studies have been
made on these lines in the host parasite relationship of the "green ear disease"
of pearl millet caused by Sclerospora graminicola. Turner (1960) found that
zoospores of Phytophthora palmivora remained viable for six months in the soil
at 50% water holding capacity. Royle (1963) established that, zoospores of
Pythiwn aphanidermatum, P. cryptogea9 P. fragariae and Aphaftomyces euteiches
retained motility 1-3 hrs in non-sterile soil. Following water column technique,
Mehrotra (1970) demonstrated that zoospores could move for a limited distance
through soil towards plant roots, such movement being largely dependent on
movement of water through the soil. Although infectivity of sporangia and
zoospores (Thakur and Kanwar 1977)have been established no attempt has been
made to study fate of sporangia and zoospores in the soil. In the present study
an attempt has been made to study the germination, viability and infectivity of
the sporangia of S. graminicola ip the soil . ; .

303

304 C R Rarnesh and K M Safeeulla

2. Materials and methods

2.1. Zoospore release and viability

Three corning glass funnels of 10 cm diameter were used in the experiment.
A cheese cloth with a 1 mm pore was inserted in the funnel and 3/4 of it was
filled with sterilised red loamy soil. A rubber tubing carrying a pinch-cock
was introduced at the narrower end of the two funnels, which were used for
porangial release. The third funnel was used for sowing surface sterilised seeds
of susceptible bajra cultivar HB-3. Distilled water was added to the funnels
and allowed to become saturated from the bottom until the soil surface was
covered with 2-4 mm of water. Inoculum was prepared by following the proce-
.dures given by Safeeulla (1976). Sporangial suspension was obtained by incubating
leaf bits collected from systemically infected bajra plants in petri dishes lined with
moist filter paper and scraping sporangia, thus obtained in distilled water. In
order to observe the release of zoosporcs from the sporangia in the soil,, freshly
collected sporangial suspension was added to the soil in the two funnels and
the water film on the surface of the soil was periodically observed for zoosporc
release. After the liberation of zoospores, water was periodically drained from
one of the funnels at regular interval of 1 hr for ten hrs to test the viability of
zoospores.

2.2. Infectivity and upward movement of zoospores

For testing the infectivity and upward movement of zoospores in the soil, the
following method was used. Three day old susceptible bajra seedlings were kept
in contact with the periodically drained water to test the infectivity of zoospores.
The seedlings thus inoculated were incubated for 12 hrs and transferred to pots
containing sterilised soil. To test the upward movement of zoospores, the second
funnel containing the zoospores was connected with the help of a rubber tubing
to the narrower end of the funnel containing 4-day old bajra seedlings and the
soil was saturated with water so that a continuous column of water was maintained
in the rubber tubing connecting the two funnels (figure 1).

3. Observations

3.1. Release and viability of zoospores

Zpospores were released from the sporangia 30 min after sowing in the funnels.
Observation of zoospores at hourly intervals revealed that they were motile for a
period of 5 hrs from the time of release.

3.2. Infectivity and upward movement of zoospores

Pearl millet seedlings kept in contact with drained off water, developed downy
mildew symptoms indicating the infectivity of zoospores up to 5 hts after their
release in the soil. However the percentage of infection decreased with the
increase in the duration of retention following their release. Retention for
1-2 hrs resulted in 60% infection. At 3rd, 4th .and 5th hrs it decreased to 36%,
and 9% respectively. Beyond 5 hrs no infection >as noticed. Seedling

Viability and infect ivity of zoo spores

305

Figure 1. Viability and upward movement of zoospores.

Viability and infectivity of zoospores 307

raised in the funnel connected to the zoospores source showed downy mildew
symptoms 6 days after inoculation indicating the upward movement of zoospores.

Discussion

Sclerospora graminicola produces a large number of asexual sporangia during
night under field conditions. Out of the large number of sporangia produced
and liberated, some get into the air, a few get deposited on the same or on the
neighbouring plants and a few fall to the ground. The significance of the sporangia
falling on the soil was not realised so far in Sclerospora graminicola. Sporangia
are produced during the humid night hours, which is congenial for the release
of zoospores from the sporangia. The present study has indicated that sporangia
can germinate in the soil liberating zoospores, which can thrive for 5 hrs without
loosing their viability. The earlier reports (Haensler 1925; Lockwood and
Ballard 1959 ; Esmarch 1927 ; Kuhlman 1964 ; Chupp 1917) have failed to
establish the distance travelled by zoospores in the soil unaided by water move-
ment. From the present study it is clear that the zoospores have moved up-
wards along the rubber tube, stem of the funnel and the soil profile to infect
the seedlings in the funnel. Since zoospores can thus remain viable and infec-
tive in the soil, they may serve as a potential source of secondary inoculum for
causing infection in the field. The texture and moisture content of the soil might
also influence the viability and infectivity of zoospores. The decline in the percen-
tage of infection of seedlings with the increase in the retention period might be
due to the reduction in the number of motile zoospores coming in contact with
the roots. This also indicates that, zoospore loose their motility in the soil when
they are retained for longer periods.

Acknowledgements

One of the author (CRR) acknowledges the Indian Council of Agricultural Research
New Delhi, for financial assistance.

References

Bewley W F and Buddin W 1921 On the fungus flora of gteen house water supplies, relation

to plant disease ; Ann. AppL Biol. 8 10-19
Chupp C 1917 Studies on club root of cruciferous plants ; New York State Agric. Exp. Stn.

Bull. 387 421-452

Esmarch F 1927 Untersuchungen zur biologic des kartofesl knebser ; Angew. Bot. 9 88-124
Haensler C M 1925 Studies on the root rot of peas (Pisum satium) caused by Aphanomyces

euteiches Drechsler. ; New Jersey Agric. Exp. Stn. Rep. 46 467-484
Klotz C J, Wong P P and De Wolfe T A 1949 Survey of irrigation water for the presence

of Phytophthora species pathogenic to citrus ; Plant Dis. Rep. Suppl 43 830-832
Kuhlman E G 1964 Survival and pathogenecity of Phytophthora cinnamomi in several Western

Oregon soils ; Forest ScL 10 151-158
Lockwood J C and Ballard J C 1959 Factors affecting a seedling test for evaluating resistance

of pea to Aphanomyces root rot ; Phytopathology 49 406-410
Mclntosh D L 1964 Phytophthora species in soils of the Okang^s and §imilankameen valleys

of British Cotyrnbi^ ; Can. J. Bot. 42 1411- 141?

308 C R Ramesh and K M Safeeulla

Mehrotra R S 1961 Phytophthora parasitica var piperina on pan piper betel Ph.D. thesis Univ.

Saugar India p. 181
Mehrotra R S 1970 Techniques for demonstrating accumulation of zoospores of Phytophthora

species on roots in soil ; Can. J. Bot. 48 890-892
Nolla JAB 1928 The black shank of tobacco in Puerto Rico ; /. Dept. Agric. Puerto Rico

12 185-215
Royle D J 1963 The behaviour of zoospores of Pythium aphanidermatwn in response to roots,

root substances and chemical compounds ; Ph.D. thesis, University of Western Ontario,

London, Ontario, Canada p. 194
Safeeulla K M 1976 Biology and control of downy mildews of pearl millet, sorghum and finger

millet ; Downy Mildew Research Laboratory, University of Mysore, Mysore, India. (Final

tech. report project CR 352, PL 4SO Grant No. FG-IN-414 1969-1975) XV + 304
Thakur D P and Kanwar Z S 1977 Infectivity of sporangia of Sclerospora graminicola on

pearl millet downy mildew ; Indian J. My col. Plant Path. 7 104-105
Turner P D 1960 Saprophytic activity of Phytophthora palmivora Annual Report West African

Cacao Res. last. 1958-59 25-26
Zentmyer G A and Richards S J 1952 Pathogenecity of Phytophthora cinnamomi to avocad,©

trees and the effect of irrigation on disease development ; Phytopathology 42 35-37

»roc. Indian Acad. Sci. (Plant ScL), Vol. 91, Number 4, August 1982, pp. 309-318.
g) Printed in India.

nitiation, development and structure of root nodules in
iome members of the tribe Trifolieae (Papilionaceae)

G L SHAH and M GOPALA RAO

Department of Biosciences, Sardar Patel University, Vallabh Vidyanagar 388 120,
India

MS received 16 September 1981 ; revised '14 June 1982

Abstract. Initiation and development of root nodules are studied in 7 species and
the structure in 4 species, belonging to 3 genera of the tribe Trifolieae. The shape
of the mature nodules may be spherical, cylindrical, fan-like or coral-like. The
bacterial threads enter the root through the intact epidermis and cause proliferation
in cortex by liberating the bacteria. The origin of nodules in the investigated taxa
is exogenous and they belong to the * apical * type in Kodama's classification. A
mature nodule comprises of nuristematic zone, cortex with vascular bundles and
the bacteroid zone. The bacteroid zone is heterogeneous and is composed of infected
and uninfected cells. •

Keywords. Trifolieae; toot nodule; exogenous origin ; bacterial thread; proli-
feration of cortex ; bacteroid zone.

L Introduction

fhe root nodules have been a subject of investigation because of nitrogen fixing
unction. Though extensive studies have been made on physiology, cytology
rid histology of root nodules in the leguminosae in general, only a few investi-
;ators have paid attention to the nodule anatomy in the tribe Trifolieae (Peirce
902; Thornton 1930; Nutman 1948; Dart and Mercer 1963, 1964; Jordan et al
.963; Mosse 1964; Munns 1968; Tu 1977). The present irivesligation is a
,upplement to the existing data based on the study of initiation and development
>f nodules in Medicago orbicularis All., M. scutellata Mill., M. truncatula Gaertn.,
\felilotus officinalis Pallas, M. wolgica Poir., Trigonella corniculata L. and
r. foenumgraecum L. and structure of nodules in Medicago sativa L., Melilotus
ilba Med. M. indica All. and Trigonella foenum-graecum L.

L Materials and methods

ieeds of Trigonella corniculata were obtained locally from a seedsman whereas
ully developed nodules of Melilotus indica were collected from the plants growing
rild in the University Campus. The rest of the species were raised from the
eeds obtained from Berlin in the University Botanical Garden.. Low viability

309

310 G L Shah and M Gopala Rao

of the seeds precluded the investigation of nodular structure of some species. The
roots and rootlets of seedlings and root nodules of different stages of develop-
ment were fixed in FAA (Johansen 1940) and stored in 70% alcohol after 48 hrs.
Longitudinal and transverse sections of roots and nodules (5-8 /on) were stained
with safranin O and fast-green FCF (Berlyn and Miksche 1976) and made perma-
nent in a customary way.

3. Observations

3«L Morphological description

The developing nodules are spherical, but the fully developed ones are commonly
cylindrical and variously lobed often becoming fan-like or coral-like and rarely
spherical. They occur on primary, secondary and tertiary roots. There is no
variation in size except that the smallest nodules are in Melilotus alba and the
largest in Trigonella foenum-graecum (figures 1-4).

3 .2. Infection., initiation and development

The entry of bacterial threads (BT) into the root cortex through the intact epidermis
(E) is observed only in Melilotus officinalis, Medicago truncatula and Trigonella
foenum-graecum (figure 5). The cortical cells through which the threads pass
are relatively larger than the remaining cells (figure 6, at arrows). The threads
are often found in close proximity or in contact with the host cell nucleus (N)
while passing through the cells (figures 5, 7). Further, they develop bulbous or
funnel-shaped swellings (s) adjacent to the cell wall in Medicago truncatula,
Melilotus officinalis and M. wolgica (figures 7, 8). The threads rupture and
liberate the bacteria into the middle of the cortex. The bacterial infected cells
contain dense cytoplasm and distinct nucleus, referred here as "proliferation
cortical initials" (PCI) (figure 9). By repeated divisions these initials produce a
mass of cells, eaph with dense cytoplasm and a nucleus (figure 10). Gradually,
the divisions become restricted to the distal end as a result of which the mass
of cells attains spherical shape (figure 11). The developing spherical nodule is
now distinguishable into 2-3 layers of peripheral tangentially elongated cells (PL)
around the inner mass of cells (MC) (figure 12). It is at this stage the inner mass

Figures 1-10. 1-4. Mature root nodules. 1. Melilotus alba. 2. Medicago
sativa. 3 and 4. Trigonella foenum-graecum. 5-10. T.S. of roots. 5. Meli-
lotfts officinalis showing the entry of bacterial thread, into the root. 6. Medicago
truncatula showing large cortical cells through which bacterial threads pass at arrows.

7. Melilotus wolgica showing the contact of bacterial thread with host cell nucleus.

8. Melilotus officinalis showing the swelling of bacterial thread near cell wall,
note the breaking of thread at arrow. 9. Melilotus wolgica showing proliferation
cortical initials. 10. Melilotus wolgica showing mass of proliferated cells.
1-4 line indicates 1 mm ; 5 x 260 ; 6 X 160 ; 7 x 650 ; 8 x 380 ; 9 X 430 ;
10 X 160.

Abbreviations : BT, bacterial thread ; N, host cell nucleus ; E, epidermis 5
s, -swelling; PCI, proliferation cortical initials.

Intiaition, development and structure of root nodules

311

312

G L Shah and M Gopala Rao

msm^mtK

initiation^ development and Structure of root nodules 313

of cells transforms into the bacteroid zone (BZ) and the peripheral layers into
nodule cortex (NC) with distinct apical meristematic zone (MZ) (figure 13). The
ruptured threads even after liberating the bacteria remain in the developing nodule
(figure 12 at arrow). The developing nodule now protrudes from the root with
a protective covering of a few layers of the root cortical cells (figure 13).

3.3. Structure

A mature nodule consists of meristematic zone (MZ), nodule cortex (NC) with
vascular bundles and the central bacteroid zone (BZ).

The meiistematic zone is situated at the apex of the nodule, composed of multi-
layered, thin walled, tangentially elongated cells with dense cytoplasm and promi-
nent nuclei, arranged compactly in regular rows (figure 14).

The nodule cortex is homogeneous comprising of 3 to 6 layers of compact
parenchymatous cells with vascular bundles (figures 15, 20). In Medicago sativa,
Melilotus alba and M. indica 2 vascular bundles enter the base of the nodule
(figures 16, 17 at arrows), but in Tngonella foenum-graecum there are 4 vascular
traces two of which supply to each side of the nodule (figures 18, 19 at arrows).
The vascular strands arise opposite to the protoxylem of the root stele (figure 17).
The vascular strands during their upward course, branch repeatedly within the
nodule cortex, but do not come in contact with the bacteroid zone (figure .20).
The vascular bundles are "inversely collateral" and conjoint, surrounded by an
endodermis (EN) (figure 21).

The bacteroid zone is heterogeneous composed of approximately 75% infected
cells (ic) and 25% uninfected cells (uc) (figure 22). The uninfected cells are
packed with spherical starch grains and interspersed within the tissue of infected
cells (figure 23 at arrow). The young infected cells are with distinct nucleus and
contain bacteria (figure 24). In the maturing infected cells small vacuoles (v)
appear (figure 25) and subsequently their fusion tend to form a large vacuole,
pushing the contents and the nucleus towards periphery of the cell (figures 26, 27)
and at this stage the mature infected cells are about four times larger than the
uninfected ones (figure 22). At a later stage the disappearance of the nucleus

Figures 11-19. 11-13. T.S. of roots. 11. Medicago orbicularis showing deve-
loping spherical nodule. 12. Tngonella corniculata showing differentiation of
peripheral layers and inner mass of cells in the spherical nodule. 13. Tngonella
foenum-graecum showing protrusion of nodule from the root. 14. L.S. of nodule
of Tngonella foenum-graecum showing meristematic zone. 15. T.S. of nodule of
Medicago sativa. 16. T.S. of the basalmost region of nodule of Medicago sativa
showing vascular bundles, at arrow. 17. T.S. of root with .nodule of Melilotus
indica showing vascular connections with root stele, at arrows. 18. -T.S. of the
basalmost region of nodule of Tngonella foenum-graecum showing vascular traces,
at arrows. 19. L.S. of the root with nodule of Tngonella foenum-graecum showing
vascular connections with root stele, at arrows.

11 x 532 ; 12 x 380 ; 13 x 43 ; 14 x 450; 15 x 32; 16 x 100; 17 x 48; 18 x 80;
19 x 38.

Abbreviations : PL, peripheral layers ; MC, mass of cells ; NC, nodule cortex;
BZ, bacteroid zone ; MZ, meristematic zone ; VB, vascular bundle ; RC, root cortex;
RS, root stele. .

314 G L Shah and M Gopala Rao

and clumping of the bacteroids of the infected cells lead to the senescence of
the nodule (figures 28, 29 at arrow).

4. Discussion

The mode of invasion of bacterial thread into the roots is considerably interesting.
Inmost of the legumes the infection takes place through root hairs (Thornton
1930 ; Bond 1948 ; Harris etal 1949 ; Arora 1956b, c ; Nutman 1959 ; Dart and
Mercer 1963, 1964 ; Narayana 1963 ; Narayana and Gothwal 1964 ; Kapil and
Kapil 1971), but in a few cases it is reported to enter through the intact root
epidermal cslh (McCoy 1929 ; Sshaede 1940), broken epidermal ceils (Bieberdorf
1938) and wounded and ruptured cortical cells during the emergence of lateral
roots (Allen and Allen 1940 ; Arora 1954). Narayana and Gothwal (1964) reported
the infection thread to enter through root hair in Ttigonella foenum-graecum,
but we have noted its entry through the intact root epidermis in Ttigonella foenum-
graecum and also in Medicago sativa and Melilotus officinalis.

From the data presented regarding the depth of penetration of infection thread
into the root cortex (Biebsrdorf 1938 ; Bond 1948 ; Arora 1956c ; Narayana
1963 ; Kapil and Kapil 1971) it appears that there is no correlation between the
depth of penetration of the thread and the structure (thickness) of the root cortex.
In the plants of present investigation also the penetration of the infection thread
is up to middle region of the cortex in primary, secondary and tertiary roots.

The dissemination of the bacteria can be either by the invasion of the infec-
tion threads of the newly produced cells (Bond 1948 ; Harris et al 1949 ; Nara-
yana 1963 ; Narayana and Gothwal 1964 ; Kapil and KapiJ 1971) or by the
division of the infected cells (McCoy 1929 ; Allen and Allen 1940 ; Arora 1954).
In our plants it is by the second method.

•Several view points have been put forth to explain the formation of the funnel
shaped swellings in the infection threads (McCoy 1929 ; Thornton 1930 ; Harris
etal 1949 ; Arora 1956c ; Narayana 1963 ; Narayana and Gothwal 1964 ; Dixon
1964). Such swellings are observed in the present investigation and may be due
to emaciation of the bacterial mass caused by the stretching of the thread during
the enlargement of the host cell harbouring it (Arora 1956c). The breaking of

Figures 2(1-29. L.S./T.S. of nodules. 20. L.S. of nodule of Tngonella foertum-
graecum showing branching of vascular strands. 21. Melilotus indica showing
vascular bundle. 22. Bacteroid zone of Medicago sativa showing infected and
uninfected cslls. 23. Melilotus indica showing starch grains in the uninfected
cells, at arrow. 24-26. Melilotus alba. 24. Young infected cells. 25. Maturing
infected cell showing vacuoles. 26. Fusion of vacuoles in the maturing infected
cell. 27. Miture infected cell of Melilotus indica showing large central vacuole
aid paripheral contents. 28-29. Trigondla foenum-graecum showing infected cells
at the senescence. 28. Early stage of bacterial clumping, at arrows. 29. Late
stage of bacterial clumping, at arrow.

20 x 38 ; 21 x 768 ; 22 X200 ; 23 x 380 ; 24 x 1040 ; 25 x 960 ; 26 X 640 •
27 x 350 ; 28 x 1600 ; 29 x 560.

Abbreviations : NC, nodule cortex ; EN, eadadermis ; XY, xylera ; PH, phloem ;
uc, uninfected cell ; ic, infected cell ; N, nucleus ; V, vacuole.

Tnitiation, development and structure of root nodules

315

Initiation, development and structure of roo-t nodules 317

the thread in the middle (Narayana 1963) further supports our conclusion (figure 8
at arrow).

The meristematic zone may be situated at the apex or at several places
surrounding the nodule and accordingly they may be " apical " or " spherical "
respectively (Kodama 1967). In the species of the present investigation the meri-
stematic zone occurs at the apex and thereby the nodules confirm to "apical"
type of Kodama (1967).

The structure of the nodule cortex is reported to be heterogeneous in some
legumes (McCoy 1929 ; Harris etal 1949 ; Arora 1954 ; Allen etal 1955 ; Nara-
yana and Gothwal 1964 ; Kapil and Kapil 1971), but it is homogeneous, compris*
ing of 3-6 layers of compact parenchyma cells surrounded by 3-6 layers of root
cortex in the species studied by us.

There is a great variation in the number and orientation of vascular strands
connecting the root stele in different leguminous nodules (Bieberdorf 1938 ; Bond
1948 ; Harris etal 1949 ; Arora 1954, 1956a-c ; Narayana 1963 ; Kapil and Kapil
1971). We have observed four vascular connections in Trigonella foenum-graecum
(see also Narayana and Gothwal 1964) and two in Medicago sativa, Melilotus
alba and M. indica.

The arrangement of phloem and xylem in the vascular bundles in different
leguminous nodules is variable (Allen and Allen 1940 ; Allen et al 1955 ; Kapil
and Kapil 1971). It is inversely collateral in the nodules of the investigated
species (see Kapil and Kapil 1971).

All the cells in the bacteroid zone contain bacteria (Allen and Allen 1940 ;
Arora 1954, 1956a) or some of them may be infected by bacteria and others may
not (Harris etal 1949 ; Arora 1956b, c ; Narayana 1963 ; Kapil and Kapil
1971). The latter situation is observed in our plants and the percentage of infected
and uninfected cells is 75% and 25% respectively.

We are in agreement with the observations of Allen and Allen (1958) who
report that the earliest indication of senescence is the change in colour of the
bacteroid zone from red to green and a change in the nodule surface from smooth
to wrinkled. Clumping of bacteria is quite common during the nodule senescence.

Acknowledgements

We thank the director of Botanischer Garten iind Botanisches Museum, Berlin
for the gift of seeds and the Government of Gujarat for awarding a research
fellowship to MGR (11-7-1977 to 10-7-1980).

References

Allen E K and Allen O N 1958 Ecological aspects of symbiotic uitiogen fixation ; Handb.

Pfi-Physiol. 8 48-118
Allen E K, Gregory K F and Allen O N 1955 Morphological development of nodules on

Caranaga arborescens Lam. ; Can. J. Bot, 33 139-14S
Allen O N and Allen E K 1940 Response of the peanut plant to inoculation with rhizobia,

with special reference to morpholcgical development of the nodules ; Bot. Gaz. 102 121-142
Aiora N 1954 Morphological development of the root and stem nodules of Aeschynomene indica

L. ; Phytomorphology 4 211" 21 6

318 G L Shah and M Gopala Rao

Arora N 1956a Morphological development of root nodules in Crotalaria juncea ; Proc. 43rd

Indian Sci. Congr. (Agra) 244
Arora N 1956b Morphological study of root nodules on Cajanus indicus ; Proc. 43rd Indian

Sci. Congr. (Agra) 244-245
Arora N 1956c Histology of the root nodules on Cicer arietinum L. ; Phy to morphology 6 367-

37$
Berlyn G P and Miksche JP 1976 Botanical microtechnique and cytochemistry; The Iowa State

University Press, Ames, Iowa.
Bieberdorf F W 1938 The cytology and histology of the root nodules of some Leguminosae ;

J. Am. Soc. Agron. 30 375-389
Bond L 1948 Origin and developmental morphology of root Modules of Piswn sativum ; Bot.

Gaz. 109 411-434
Dart P J and Mercer F V 1963 Development of the bacteroid in the root nodule of Barrel Medic

(Medicago tribuloides Desr.) and subterranean clover (Trifolium subterraneum L.) ; Arch.

Microbiol. 46 382-401
Dart P J and Mercer F V 1964 Fine structural changes in the development of the nodules of

Trifolium subterraneum L. and Medicago tribuloides Desr. ; Arch. Microbiol. 49 209-235
Dixon ROD 1964 The structure of infection threads, bacteria and bacteroids in pea and

clover root nodules ; Arch. Microbiol. 48 166-178
Harris J O, Allen E K and Allen ON 1949 Morphological development of nodules on

Sesbania grandiflora Poir. with reference to the origin of nodule rootlets ; Am. J. Bot.

36 651-661

Johansen D A 1940 Plant microtechnique New York, USA
Jordan D C, Grinyear I and Coulter W H 1963 Electron microscopy of infection threads and

bacteria in young root nodules of Medicago saliva ; /. Bact. 86 125-137

Kapil R N and Kapil N 1971 Root nodules of Cajanus cajan : Origin, structure and onto-
geny ; Phytomorphology 21 192-202
Kodama A 1967 Cytological studies on root nodules of some species in Leguminosae II ;

Bot. Mag. Tokyo 80 92-99
McCoy E F 1929 A cytological and histological study of the root nodules of the bean, Phaseolus

mlgaris L. ; Centr. Bakt. II 79 394-412
Mosse B 1964 Electron microscope studies on nodule development in some clover species ;

/. Gen. Microbiol. 36 49-66
Munns D N 1968 Nodulation of Medicago sativa in solution culture. I. Acid-sensitive steps ;

Plant Soil. 28 129
Narayana H S 1963 A contribution to the structure of root nodule in Cyamopsis tetragonoloba

Taub. /. Indian hot. Soc. 42 273-280
Narayaua H S and Gotbwal B D 1964 A contribution to the study of root nodules in some

legumes ; Proc. Indian Acad. Sci. B59 350-359
Nutmau P S 194& Physiological studies on nodule formation. I. The relation between nodu-

lation and lateral root formation in red clover; Ann. Bot. 12 SI -96
Nutrnan P S 1959 Some observations on root hair infection by nodule bacteria ; /. Exp. Bot.

10 250-263
Peirce G J 1902 The root tubercles of bur clover (Medicago denticulata Willd.) and of some

other leguminous plants ; Calif. Acad. Sci. Proc. 3rd Ser. Botany 2 295-328
Schaede R 1940 Die kn ollechen der advent; ven wasser wurzelen con Neptunia oleracea und

ihre bakterian symbiose ; Planta 31 1-21
Thornton H G 1930 The early development of the root nodule of lucerne (Medicago sativa) ;

Ann. Bot. 44 385-392
Tu J C 1977 Structural organization of the rhizobial root nodule of alfalfa ; Can J Bot 55

35-43

Proc. Indian Acad. Sci. (Plant Sci.)» Vol. 91, Number 4, August 19S2, pp. 319-328
© Printed in India,

Turnera ulmifolia var. elegans x T. ulmifolia var. angustifolia
crosses and its bearing on the taxonomy of the species

K RAJEEV, P I KURIACHAN and C A NINAN

Department of Botany, University of Kerala, Trivandrum 695581, India

MS received 17 November 1981

Abstract. The heterostylous taxon T. ulmifolia var. elegans Urb. (2/z = 20) and
the homostylous taxon T. ulmifolia var. angustifolia Willd. (2n — 30) were selfed and
intercrossed. Both pin and thrum forms of var. elegans are self incompatible while
the variety angustifolia is self compatible. Among the intervarietal combinations
only angustifolia x elegans (thrum) crosses were successful though the hybrid seeds
were inviable.

From an analysis of the results of in vivo pollen germination studies in the
incompatible crosses, it is concluded that angustifolia pollen are of the thrum type
and its pistil is of the pin type. It is suggested that var. angustifolia has evolved
by a rare crossing over within the super gene complex for heterostyly in the parent
taxon, which might be the hexaploid (2/z = 30) heterostylous T. ulmifolia Linn. The
failure of the apparently ' legitimate ' cross elegans (pin) x angustifolia is suggested
to be due to the ploidy difference between the two varieties. On morphological
cytological and biochemical grounds the separation of the elegans element from,
T. ulmifolia complex and assigning of species status to it is suggested.

Keywords. Turnera ; heterostyly ; intervarietal crosses ; incompatibility.

1. Introduction

Turnera ulmifolia L. is a polymorphic weedy species of the New World Tropics
with a fairly high caffeine content in the seeds (Raffauf 1970; Tarar and Patil 1974).
T. ulmifolia var. elegans Urb. and T. ulmifolia var. angustifolia Willd. occur as intro-
duced weeds in South India (Gamble 1915). The former variety is heterostylous
and is self incompatible while the latter is homostylous and self and cross compa-
tible (Barrett 1978). Earlier cytological studies have shown that T. ulmifolia
var. elegans has 2n= 20 chromosomes (Raman and Kesavan 1963; Barrett 1978)
and T. ulmifolia var. angustifolia has 2n= 30 chromosomes (Barrett 1978; Tarar
and Dnyanasagar 1976, 1979). Results of intervarietal crosses between these two
taxa are reported here.

2. Materials and methods

Plants of natural populations of the two varieties occurring in the Kariavattom
Campus of the University of Kerala were used in this study. The flowers of

319

320 K Rajeev, P I Kuriachan and C A Ninan

T. ulmifolia var. angustifolia open by about 6 am while those of T. ulmifolia
var. elegans open by 7-30 am. Flowers of the former taxon were emasculated on
the evening previous to blooming. Emasculation on the day previous to blooming
caused serious damage to the buds of T. ulmifolia var. elegans and hence these were
emasculated only about one hour before the flowers open. Flowers of both the
varieties were pollinated at 7-30 am.

White's (1954) culture medium solidified with 4% agar and supplemented with
100 ppm indole acetic acid was used to culture the hybrid seeds. In vivo germination
of pollen grains was studied by staining the styles and stigmas after treatment in
lactophenol for 17hrs at 70°C. Pollen tubes taken from 9 styles were studied.
Measurements of leaves, bracteoles, stamens, styles and pollen are given from 25
observations in each case.

3. Results

3-1. Intra- and inter- varietal crosses

Turnera ulmifolia var. elegans is a profusely branched, bushy, distylous herbaceous
perennial growing up to a height of 40 cm (figure 1). The leaves are 2-5 cm to
4 -25 cm long, roundish with crenate margin. The bracteoles are linear, up to
2 cm long. The flowers are cream coloured with dark violet spots at the base of
the petals (figures 2,3). The pin styles are on an average 10 -56 mm long while
the thrum styles measure only 5 -66 mm. The average length of pin and thrum
stamens were 6 -00 mm and 10 -74 mm respectively. The pollen also showed
dimorphism with an average measurement of P x E= 64-15/* x 58-93#na in pin
types and P x E = 70- 12/*jn x 63-35/*m in the thrums. Both the types showed
an average of 20 seeds per capsule.

Plants of T. ulmifolia var. angustifolia are sparcely branching erect shrubs, rea-
ching more than 1 in in height (figure 4). Leaves are elliptic-lanceolate, 10-12- 5 cm
long with irregularly serrate margin. Bracteoles are foliaceous up to 3- 1 cm long
and 0- 75 cm broad. Flowers are long-homostylous (figure 5) with bright yellow
petals. The styles are 19 -2 mm long and the stamens 20 -44 mm. The fruits are
larger than that of the former variety and contain about 70 seeds in each.

Self and cross pollinations in T. ulmifolia var. angustifolia and both pin and
thrum forms of T. ulmifolia var. elegans and the intervarietal crosses were made
(table 1). T. ulmifolia var. angustifolia was found to be self and cross com-
patible. In T. ulmifolia var. elegans only crosses between the thrum and pin forms
were compatible. Among the intervarietal crosses only the cross T. ulmifolia
var. angustifolia x T. ulmifolia var. elegans (thrum) was found to be successful.

The hybrid seeds obtained failed to germinate when sown on wet cotton in petri
dishes and also in soil. The hybrid seeds cultured in White's medium with 100
ppm indole acetic acid also failed to germinate.

3-2. In-vivo pollen germination

In vivo germination of pollen grains in the three unsuccessful intervarietal crosses
were studied with a view to elucidate the causes of their failure. In the successful

Intervdrietal crosses in Turnera ulmifolia

321

Figures 1-5. 1. vat. elegans, plant habit. 2. Dissected thrum flower of var.
elegans, arrow points to stigma x 1. 3. Dissected pin flower of var. elegans,
arrow points to stigma x 1. 4. Var. angustifolia, plant habit. 5. Dissected
flower of var. angustifolia, arrow points to stigma in a black background x 1,

322

K Rajeev, P I Kwlachan and C A Ninan

Figures 6-11. 6-$ and 11. x $0. lO. x 2. Arrow point to ends of pollen tubes.
6, Elegans (pin) stigma showing inhibition of elegans (pir>) pollen tubes. 7. elegans
(thrum) stigma showing inhibited angustifolia pollen tubes. 8, 9. elegans (pin)
style bases showing angustifolia pollen tubes. 10. elegans (pin) gynoecium after
(a) pollination with angustifolia pollen and (b) selfing. 11. Angustifolia style
showing inhibited elegans (pin) pollen tubes,

intervarietal crosses in "furnerct ulmifotid

Table 1. Details of inter and intravarietal pollinations in T. ultnifolia var. elegans
and T iilmlfolia var. angustifolia

No. of
Type of pollination flowers
pollinated

No. of Total Average
capsules no. of no. of
formed seeds seeds per
capsule

1.

Intravarietal

17

elegans (pin) x elegans (pin) (cross)

2.

efe#w,s(pin) x elegans (pin) (self)

15

. .

3.

elegans (thrum) x elegans (thrum) (cross)

17

. .

4.

elegans (thrum) x elegans (thrum) (self)

15

..

5.

elegans (pin) x elegans (thrum)

11

3. 67 22

6.

elegans (thrum) X elegans (pin)

11

1 13 13

7.

angustifolia x angustifolia (self)

10

7 518 73

8.

angustifolia x angusti folia (cross)

10

9 603 67

9.

Intervarietal

21

elegaus(v'm) x angustifolia

10.

elegans (thrum) x angustifolia

14

..

11.

angustifolia x elegans (pin)

39

..

12.

angustifolia x elegans (thrum)

22

9 117 13

cross T. ulmifolia var. angustifolia x T. ulmifoUa var. elegans (thrum type), the
elegans pollen tubes were found to reach the stylar base in 4 to 5 hrs after pollina-
tion. Therefore, the styles in the unsuccessful intra- and intervarietal crosses were
examined 7 hrs after pollination. In both pin x pin and thrum x thrum crosses
in elegans the pollen grains germinated, but failed to enter the style (figure 6).
The ends of pollen tubes got enlarged and burst at about the basal region of the
stigma. In the three unsuccessful intervarietal crosses also the pollen germinated
in vivo. In elegans (thrum) x angustifolia, the angustifolia pollen tubes were in-
hibited at the stigma style joint. The ends of most of the tubes got enlarged and
burst at this region (figure 7). In elegans (pin) x angustifolia crosses the angus-
tifolia pollen tubes were found to reach the stylar base in about 7 hrs after polli-
nation (figures 8,9). Pollinated flowers were invariably found to fall off, but only
after 5-7 days during which period the ovary showed some enlargement (figure lOa).
Long styled flowers of elegans emasculated and bagged for a day to prevent pollina-
tion took 5-6 days to fall off, but the ovaries in these were not enlarged (figure lOb).
In angustifolia x elegans (pin) crosses the elegans pollen tubes entered the style
and grew down ; but most of the pollen tubes stopped growth just below the
stigma, where their ends ballooned and burst opened (figure 11). The maximum
length attained by pollen tubes in 9 cross pollinated styles are given in table 2.
It is seen that the maximum length of pollen tubes observed was 95 % of the length
of the style.

K Rajeev, P / guriachan and C A tfinan

Table 2. In viva pollen tube growth in T. ulmifolia var. angustifolia x 7. ulmifolia
var. elegans cross

No. of
styles

Length of Maximum growth
style (mm) of pollen tubes
(mm)

Length of pollen
tube as percent-
taga of stylar
length

1

15-0

10-1

67-33

2

15-5

11-7

75-48

3

15-0

11-5

76-67

4

16-0

12'7

79-38

5

15-5

12-7

81-94

6

15*5

13-0

83-87

7

16*0

13-9"

86-88

8

15-5

13-9

89-68

9

16-0

15-2

95-00

4. Discussion

4-1. Causes of intervarietal incompatibility

Intervarietal crosses between angustifolia (female) and elegans thrum (male) were
successful with. 40-9% fruit set. There was no fruit set in the reciprocal cross :
though the angustifolia pollen grains germinated in vivo, the tube growth was
inhibited in the stigma itself. The nature of inhibition of pollen tubes in this cross
is similar to that in the incompatible intravarietal crosses of the heterostylous
elegans. Moreover, in elegans pin (female) x angustifolia (male) crosses, angusti-
folia pollen grew to the base of the styles in pin plants of elegans. These facts may
suggest that with regard to its incompatibility reaction to the pin and thrum forms
of elegans, angustifolia pollen are of the thrum type.

In the intervarietal cross using pin forms of elegans as males, the pin pollen readily
grew into the styles of angustifolia. But none of the pollen tubes was found to
grow beyond 95% length of the style and the vast majority of the pollen tubes
stopped growth much earlier. Their ends became bulged and were burst. It
is clear from this that this cross is incompatible. This may further indicate that
thr'long styled Angustifolia is, with regard to the incompatibility reaction of the
pistil, a pin type. This inference is supported by the observed compatibility of the
intervarietal cross when thrum plants of elegans were used as pollen parents.

Since angustifolia is a homostyle with ' thrum type ' behaviour of pollen grains,
the intervarietal cross elegans pin (female) x angustifolia (male) is an apparently
c legitimate * cross. In this cross the angustifolia pollen tubes could be traced
to the very base of pin styles of elegans. Though this together with the enlarge-
ment and the slightly delayed abscission of the cross pollinated ovaries might

Intervdrietal crosses in Turnera ulmlfolia

325

Figures 12-13. Photomicrographs of somatic chromosomes x 1500.
elegans ; 13. var. angusti 'folia.

12. var.

Intenarietal crosses in Turnera ulmifolia $27

suggest the possibility of fertilization in the cross, there was no fruit set. Normal
growth of pollen tube without fruit set have been reported in such * legitimate '
crosses between heterostylous (pin) Linum perenne and homostylous L. lewisii also
(Baker 1961). It is not clearly understood whether the failure of fruit set observed
in the present cross is related to the interaction between the heterostylous and
homostylous systems. However, L. perenne and L. lewisii are reported to have
the same chromosome number (Fedorov 1969) whereas elegans used in this study
is a tetraploid with 2n = 20 (figure 12) and angustifolia, a hexaploid with 2n = 30
(figure 13). It is known that plants differing in chromosome numbers may show
differences between reciprocal crosses and that in such cases greater success may
be met with when the taxon with larger chromosome number is used as the seed
parent (Thompson 1930). It is likely that the above cross is unsuccessful due to
the lower level of ploidy in elegans (pin), used as the seed parent, than in angusti-
folia.

4-2. The probable origin of var. angustifolia

The pistil of angustifolia exhibited incompatibility reactions characteristic of long
styled plants and its anthers exhibited incompatibility reactions characteristic of
short styled plants. This shows that this variety is a long homostyle. Lewis
(1954, 1979) and Baker (1961) have suggested that such long homostyles may be
formed as segregants in the progeny of polyploids of heterostylous species, as a
result of rare cross-overs between 1^ and /a genes in the super gene complex
GSIilaAP, controlling the heterostyly system. Instances of homostylous segre-
gants have been reported among the offsprings of polyploids of the heterostylous
species Fagopyrum esculentum (Esser 1953) and Primula obconica (Dowrick 1957),
It is highly probable that the long homostylous angustifolia is derived from a hetero-
stylous progenitor. The variety angustifolia is a hexaploid (2n = 30). The only
other hexaploid taxon so far known in this species complex is T. ulmifolia
(Hamel 1965). Apart from the fact that T. ulmifolia is heterostylous and T. ulmi-
folia var. angustifolia is a long homostyle, these taxa resemble each other very
closely in other morphological characters (Mudaliyar and Rao 1951). Therefore
it is likely that var. angustifolia is a segregant of T. ulmifolia.

4 '3. Taxonomic considerations

Crosses between T. ulmifolia var. angustifolia and short styled plants of T. ulmifolia
var. elegans yielded some seeds. But seed set per capsule in the cross was found
to be very low compared to seed set by selfing in the seed parent (table 1). More-
over, the hybrid seeds were inviable. These facts reveal the existence of a total
reproductive barrier between the two taxa. Turnera ulmifolia var. angustifolia
with 2n= 30 chromosomes (Tarar and Dnyanasagar 1976; Barrett 1978) is cytolo-
gically distinct from T. ulmifolia var. elegans with 2n = 20 (Raman and Kesavan
1963; Barrett 1978). T. ulmifolia var. elegans is endowed with a stable distyly
system having self incompatibility. The plants of this variety are of a spreading
type, with roundish leaves, 2- 5 to 4-25 cm long having crenate margin with promi-
nent scarlet spots in the basal parts of petals. Plants of the variety angustifolia

328 K Rajeev, P t Kuriackan and C A tfinart

on the other hand are erect and shruby, plants lack the corolla spots, have ctfaftge
yellow petals and elliptic-lanceolate leaves, 10 to 12- 5 cm long and with irregu-
larly serrate margin. Besides, with respect to their foliar phenolic constituents
these two varieties showed isolation values as high as 66-66% for elegans and
68 -18% for angustifolia, which denote significant difference between them (Rajeev,
Kuriachan and Ninan, unpublished). The foliar phenolic components of T. ulmi-
folia (s.s) are not known. Though it is heterostylous (Martin 1965) as T. ulmi-
folia var. elegans, in other morphological characters it resembles I7, ulmifolia var.
angustifolia more closely than T. ulmifolia var. elegans (Mudaliar and Rao 1951).
Cytologically also T. ulmifolia with 2n = 30 (Hamel 1965) is nearer to T. ulmifolia
var. angustifolia than T. ulmifolia var. elegans. Thus evidence from crossability,
cytology, morphology and foliar phenolic constituents support the separation
of the ' elegans ' element from Turner a ulmifolia complex and assignment of species
status to it.

References

Biker H G 1961 Rapid spsciation in relation to changes in the breeding system of plants, in

Recent Advances V« Botany Vol. 1 8S1~885 (University of Toronto Press : Toronto)
Barrett S C H 1978 Heterostyly in a tropical weed : the reproductive biology of Turnera ulmi-

folia complex (Turneraceae) ; Can. J. Bot. 56 1713~1725
Dowrick V P J 1957 Cited from Baker H G (1961)
Esser K 1953 Cited from Baker H G (1961)

Fedorov A 1969 Chromosome Numbers of Flowering Plants. (Komarov Botanical last. : Lenin-
grad)
Gamble J S 1915 Flora of the Presidency of Madras Vol. 1 2nd edn. (Botanical Survey of India.

Calcutta 1967)
Hamel J L 1965 Le noyan et les somatiques di Turnera ulmifolia L. ; Mem. Mus. Nat . Hist.

Natur. Now. Ser. B. Bot. 16 3-7
Lewis D 1954 Comparative incompatibility in Angiosperrns and Fungi in Advances in Genetics

6 235-285 (ed) M Demerce (Academic Press Inc. : New York)
Lewis D 1979 Sexual incompatibility in Plants (Edward Arnold Ltd. : London)
Martin F W 1965 Distyly and incompatibility in Turnera ulmifolia; Bull. Torrey Bat. Club 92

185-192
Mudaliar C R and Rao J S 1951 A contribution to the taxonomy of the genus Turnera ; Madras

Agric. J. 38 319-371
Raman V S and Kesavan P C 1963 Meiosis and nature of polyploidy in Turnera ulmifolia ; /.

Indian Bot. Soc. 43 495-497
Raffauf R F 1970 A Handbook of Alkaloids and Alkaloid Containing Plants. (Wiley Interscience :

New York)
Tarar J L 1974 Cited from Tarar J L and Patil K J Qualitative and quantitative estimation

of caffeine in Turnera ulmifolia L. ; Indian J. Bot. 2 118-119 (1979)
Tarar J L and Dnyanasagar V R 1976 Karyotype studies in Turnera ulmifolia Linn, var angusti*

folia wllld. Botan;que 7 217-222 cited from Tarar and Dnyanasagar, 1979
Tarar J L and Dnyanasagar V R 1979 Meiotic studies in Turnera ulmifolia Linn. var. angusti*

folia Willd. ; /. Indian Bot. Soc. 58 167-175
Thompson W P 1930 Causes of differences in success of reciprocal interspecific crosses ; Am.

Nat, 64 407-421
White P R 1954 Cultivation of Animal and Plant Cells (2nd edn) (The Ronald Press Co. : New

York)

Proc. Indian Acad. Sci. (Plant Sci.), Vol. 91, Number 4, August 1982, pp. 329-350.
© Printed in India.

Airborne pollen grains of Visakbapatnam : A combined field and
air sampling study

A JANAKI BAI* and C SUBBA REDDI

Department of Environmental Sciences, Andhia University, Waltair 530003, India
* Botany Department, Andhr? University, Waltair, India

MS received 9 December 1981 ; revised 21 June 1982

Abstract. Field assessments at regular intervals from April 1975 to March 1979
recorded 61 plant species comprising 13 grasses, 20 weeds and 28 trees and shrubs
as emitting appreciable amounts of pollen into the atmosphere of Visakhapatnam.
The data also showed relative prevalence of these taxa in different zones of the
city and their flowering periods. Pollen output in terms of mimbei per anther and
per flower was determined for 29 taxa. Air sampling with rod traps of 0-53 cm
diameter enabled the identification of 23 different pollen types in the atmosphere
with Poaceae accounting for Ca. 37% of the total pollen load. Casuarina contri-
buted to 13% followed by Cyperaceae 6%, Eucalyptus 5-8%, Dodonaea 3-8%,
Amaranth-Chenopod and Phoenix each 3-4%, Borassus 2-4% and Peltophorum
2%. Of the total identified pollen, ca 85% belonged to anemophilous taxa. There
was no pollen-free day. The total pollen and individual types displayed seasonally
quite closely corresponding with the blooming seasons of the source plants. Three
pollen peaks, two in the wet period (June -November) and one in the dry period
(December — May) were evident. Year to year variations in pollen abundance
occurred and urban growth affected pollen frequency pointing to the need for routine
[V; monitoring of the atmosphere.

Keywords. Pollination calendar ; .pollen pioductivity ; airborne pollen ; atmos-
pheric biopollutants ; Poaceae ; Casuarina.

1. Introduction

Allergic responses to airborne biogenic particles impose major adverse effects on
the physical and economic health of mankind (Davis 1972). Airborne pollen
grains belong to this group of particles and have long been recognised as the inci-
tants of rhinitis and asthma. In order to identify the offending pollen agent(s)
it is necessary to monitor the pollen particles in free air, their prevalence and
emission patterns through systematic air sampling. Field assessments recording
the relative abundance and blooming phenology of source plants complement
and greatly increase the value of such information. In areas of industrial atmos-
pheric pollution there is every likelihood that the airborne pollen and chemical
pollutants interact with each other and result in the aggravation of human dis-
comfort (Newmark 1970 ; Nilssow and Nybom 1978).

329

330 A Janaki Bai and C Subba Reddi

Visakhapatnam, situated in Andhra Pradesh on the east-coast of India is an
industrial area but which also has a rich vegetation (Venkateswarlu et al 1972).
It is thus likely that the air over the city is being charged with pollen from vegeta-
tion and gaseous emanations from industries that may cause suffering to the in-
habitants. The experience of clinicians in the King George Hospital and of private
practitioners support this supposition. The situation thus calls for serious efforts
at routine atmospheric monitoring so that necessary measures can be taken to
alleviate the human affliction. Efforts at trapping the airborne pollen were
begun a decade ago (Subba Reddi 1970), but the information at hand is still far
from complete. In this paper we present data describing the composition of
the airborne pollen, relative prevalence of the constituents and their seasonal emis-
sion patterns over a period of four years from April 1975 to March 1979 together
with the distribution, blooming phenology of source plants and their pollen pro-
ductivity per anther and per flower. We suggest these data serve as a basis for
local clinical strategy and as a comparison with pollen spectra established
elsewhere in India and abroad.

2. Materials and methods

2*1. Physiography and climate of the study area

The physiography of Visakhapatnam (latitude 17° 42' N and longitude 82° 18 ' E)
and its surroundings is shown in figure 1. On two sides the city is bounded by
hill ranges ; on the north-eastern side the famous pilgrimage centre Simhachalam
or Kailasa hill range with an average height of 300 m and about 16 km long and
on the southern side Yarada hill range running for about 8 km w ith an average
height of 300m and projecting prominently into the Bay, and popularly known
as Dolphin's nose. The shore of Bay of Bengal forms the eastern boundary and
the western side is bounded by the tidal basin called locally Vuppateru.

The most important feature of the climate is the alternation of monsoon seasons
classified by Indian Meteorologists as : (i) The northeast monsoon season
(December-February), (ii) The hot weather season (March-May), (iii) The
southwest monsoon season (June-September) and (iv) The retreating southwest
monsoon (October-November).

Figure 2 shows the mean distribution of temperature and rainfall for the period
when spore trapping was done. The mean monthly temperature was 27-4° C and
variation of the mean from month to month was approximately 2°C. The diffe-.
rence between mean monthly minima was no more than 9°C. The hottest weather
occurred in May, while the coolest in January. Most precipitation fell during.
June-November. The total precipitation during the .4-year period declined in
each successive year.
^l;-g:,

2«2. Field assessments

Visakhapatnam was divided into seven zones (figure 1). Zone-I constitutes the
thickly built up area ; zone-II a sparsely built up area with vacant land patches ;

Airborne pollen

331

332

A Janaki Bai and C Subba Reddi

220-

180-

140-

60-

20-

-15

N

M

252
o

Figure 2. Mean monthly, mean monthly maximum and mean monthly minimum
temperature and rainfall at Visakhapatnam for the period of the study, April 1975
to March 1979.

zone-Ill the areas adjacent to the Railway line ; zone-IV the industrial belt ;
zone-V a swampy area ; zone-VI the Yarada hill range and zone-VII the Kattasa
hill range. The last three zones were not included in the survey as the vegetation
of the two hill ranges is sparse scrub with Dodonaea occurring in considerable
frequency and the swampy area has mainly Avicennia plants of stunted growth.
Trips were made to the other four zones at approximately weekly intervals during
1975-1979 to record the prevalence of anemophilous and such entomophilous

Airborne

plants capable of liberating sizeable amounts of pollen into the ambient air.
Arbitrary units like sparse, common, more common and abundant were used to
indicate the prevalence. Concurrently observations were made on the onset,
intensity, duration and termination of flowering of these plants.

Taxonomic identification of the plants was done using the Flora of Visakhapatnam
by Venkater»warlu et al (1972) as well as comparison with the authenticated
herbarium specimens available in the Botany Department", Andhra University.

2-3. Pollen productivity

The output of pollen per anther was assessed by the method described by Subba
Reddi (1976). Mature and undehisced anthers were taken into clean, dry, stop-
pered tubes with the aid of forceps and allowed to dry. Care was taken not to
injure the anthers while transferring them into tubes. Afcer the completion of
dehiscence, a measured volume of 50% alcohol containing methyl green stain was
added to each tube with a small quantity of the wetting agent ' Tween 20 ' and
the contents shaken thoroughly to get a uniform suspension of pollen grains. From
the suspension thus obtained, one ml was pipetted out into a counting chamber and
the pollen counted. Three counts were made for each sample and from them was
calculated the number liberated by the anthers in that sample. Afterwards the
emptied anthers were mounted on a microscope slide and examined for remaining
pollen to make any necessary correction.

Knowing the number of pollen grains produced by the number of anthers in-
cluded in a particular sample, the pollen productivity was then estimated per anther
as well as flower.

2-4. Air sampling

This was done using glass rods of 0- 53 cm diameter and 23 cm long wrapped with
18mm square sticky cellophane strips. Each rod was supported vertically and
exposed for 24-hour periods (1700-1 600 hr) in shelters 'of the type used by Hyde
and Williams (1945) so as to protect the cylinder from rain other than that experi-
enced during violent storm. Airborne pollen grains are impacted by the wind
onto the traps.

The details of preparing, exposing cylinders, mounting of the trace and scanning
were the same as described by Ramalingam (1968) and Subba Reddi (1970). The
pollen counts are expressed as no/cm8 of the trap surface.

3* Results and discussion

3 - 1 . Plant species, their prevalence and flowering periods

A variety of plant taxa was reported to occur in the study area (Venkateswarlu
et al 1972). However, field observations on the mode of pollination indicated that
only a relatively small number would emit pollen into the ambient air in sizeable
amounts. Table 1 lists such plants with data on their relative prevalence in diffe-

334

A Janaki Bai and C Subba Reddi

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snt zones of Visakhapatnam and their periods of flowering. The pollination
alendar of some of the more important taxa is graphically presented in figure 3
iiich shows clearly that, unlike temperate regions of the world, one or the other
f the diverse plant taxa was in flower at all times of the year with consequent
beration of pollen into the ambient atmosphere. During two periods a maximum
umber of wind-pollinated plants were in bloom. Most Poaceae and other weeds
roe in bloom during late June-early December, while most of the trees bloom
uring January-April. Some of the weeds and grasses flowered in both periods

&6donaea

Eucalyptus

Artemisia

Xan.thlum

Amaronthac*

Cyperaceae

Pandanus

Gramineae

Syzygium

Peltophorum

Emblica

Madhuca

Borassus

Holoptelea

Casuarina

Phoenix

Mimusops

Crotoh

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Riclnus

P [ M { A | M ( J

f \ maxim at flowering - 'duration of flowering

Figure 3, Pollination calendar of some plant species at Visakhapatnam.

A Janaki Sai and C $ubba Reddi

but they were more, numerous during late Jute-early December, Thpse obser-
vations are in general agreement with the. earlier records of the blooming periods
of the plant specie of this area (Subba Reddi 1974), . .

3-2- Pollen productivity ^ •''•• ...

Table 2 giv&s the estimated pollen productivity .per antfcer .as well as .flower
determined for 29 of the -61 plant- taxa listed iti'table L The sample, size (the
number of anthers included in a sample, percent of pollen that failed to get
liberated into the medium together with the size of individual pollen types are
also included in the table.

Among these 29 plants, the highest pollen output per anther was for Phoenix,
whereas jnaximum number per flower was for Ricinus. The lowest production
both per anther and per flower was for Cyperus compressus. It might be expected
that pollen productivity in different plants would be influenced by the size of
grains as well as anthers, but this is not borne out by the results (table 2).

The pollen productivity data given in table 2 indicate the relative capability of
the different taxa to pollute the ambient air with their pollen. However, in some
cases all the pollen produced might not be emitted into the air because of the
hindrance offered by the plant habit. This is so in the case of Phoenix which was
first for pollen productivity but the thick crown of leaves obstructs the free dissemi-
nation of pollen.

Distinguishing plant species by light microscope examination of pollen is not
always possible, e.g., the pollen of stenopalynous groups like Poaceae and
Amaranthaceae-Chenopodiaceae are not readily separated. In such cases the
relative importance of the taxa contributing to the airspora might be determined
by a knowledge of their pollen productivity coupled with tfyeir distributional data.
For instance, the two species of Amaranthus greatly vary in' the quantity of pollen
produced (table 2) and by knowing their relative abundance in a region, one can
easily assess which species is important in the area. Assessed in this way, at
Visakhapatnam A. spinosus is more important than A. gracilis.

3 • 3 . Components of airborne pollen flora and their relative contributions

Over the 4-year period a total of 34976 pollen/cm2 of trap area were counted, Of
these, 30851 pollen were identified and assigned to 23 different pollen types which
are listed in table 3 alongwith the percent contribution of each to the total pollen
flora. Numerically Poaceae ranked first with a mean contribution of 37-32%
followed by Casuarina (13-11%), Cyperaceae (6-15%), Eucalyptus (5-83%),
Dodonaea (3-8%), Amaranth-chenopod and Phoenix each 3-44%, Borassus
(2- 45%) zn&Peltophorum (2-21 %). It may be noted here that the Cardiff aero-
biologists are of the opinion that the numbers alone are misleading as glides to
the relative importance of aeroallergens (Hyde and Adams 1960 ; Hyde and
Williams 1961). They stressed how the relationship between number and pollen
volume might be important in allergy, because some pollen although numerically
minor, might assume dominance in terms .of * bulk concentration'. However,
they did, not. stress enough that allergens that are superficial or migrate through

Airborne pollen

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340 A Janaki Bai and C Subba Reddi

Table 3, Components of the airborne pollen flora and their relative contributiont

4 year average

Pollen type seasonal Contribution

total

Gramineae

3263

37.15

Casuarina

1146

13.05

Cyperaceac

536

6.10

Eucalyptus

510

5.81

Dodonaea

33S

3.85

Phoenix

301

3.43

Araaranth-chenapod

300

3.42

Borassus

214

2.44

Peltophorum

193

2.20

Syzygium

178

2.03

*Muntingia

142

K62

Ricinus

130

1.48

Embliea

99

1.13

Asteraceae

98

1-12

Ooco&

73

0.83

Mimosoideae

56

0.64

Holoptelea

53

0.60

Tamarindus

29

0.33

Croton

25

0.28

Artemisia

25

0.28

MoruS

19

0.22

Cicca

15

0.17

Xanthiutn

9

0.10

Damaged and unidentified

pollen 1031 11.74

pollen surface might be more potent in unit weight of small rather than large
pollen (Hirst 1973). In this connection it is worth mentioning that the variety
of size was one of the features of the complete airspora that impressed Gregory
in 1951 (Gregory 1973) and led him to stress how size would influence dispersal.

3-4. Pollen groups counted

In allergy literature plant species are generally grouped into grasses, weeds and
trees (Duchaine 1959). Following this . tradition, the pollen recognised, were
differentiated into these three groups (table 4). Further, a division of these
pollen according to the mode of pollination ;pf the source plants was made. The
average contribution of grass, weecl and tree (including shrubs) pollein to th$ totaj

Airborne pollen 341

Table 4. Pollen group counts

% contribution to the total identified pollen

Name of group —

I II III IV

Gramineae

50-69

39-08

38-82

41-35

Weeds

9-11

17-26

10-89

13-33

Anemophilous

8-04

25-24

9-96

10-98

Entomophilous

1*06

2-02

0-93

2-35

Trees and shrubs

40-22

43-66

50-28

45-33

Anemophilous

30-57

27- lg

32-52

34-60

Entomophilous

9-63

16' 4S

17-77

10-73

Total anemophilous

81-31

81-50

81-30

86-93

Total entomophiloust

10-70

18-50

18-70

13-07

I » 1975-76 II =* 1976-77 HI = 1977-78 IV « 1978-79.

identified pollen were 42, 13, 45% respectively. The tree pollen were distributed
over 16 types and the weed pollen over 6 types. Of the four seasons, the 1st
year witnessed the highest incidence of grass pollen (50- 69%) ; it even excelled the
tree pollen count (40-22%) in this year. The 2nd year registered maximal inci-
dence of weed pollen (17-2%), and in the 3rd year, tree pollen (50-28%). As
expected, pollen from anemophilous taxa were predominant in the catches. On
average, 84-7% of the total identified pollen were of wind-pollinated taxa.

3 • 5 . Seasonal periodicity in airborne pollen

Table 5 and figure 4 give the trends in the seasonal emission of the different pollen
types recognised and of total pollen. While arriving at the data of table 5 each
month was considered to consist of a uniform 30 days and the monthly totals were
got by applying a correction factor. In figure 4 the counts of the same month over
the four-year period were averaged and plotted as a function of time. Additional
curves were also drawn through the higher and lower points. Such a graphical
representation of the data gives an indication of the most likely density of pollen
to be expected and the probable range about the mean (Ogden etal 1974 • .Solo-
mon 1976).

As was thought of from a consideration of the pollination calendar, no pieriod
altogether free from airborne pollen occurred. The total pollen frequency was
significant all through the season with three periods of pronounced occurrence*
These were July-September, November-December and February-March. Corres-
ponding to these periods of higher pollen incidence three peaks : two in the wet
period and one in the dry period, were evident. Of the two peaks in the wet period,
one was confined to the southwest monsoon. Qf the three peaks, the one in
August is the highest and, the February and November peaks follow in that

342 A Janaki Sal and G Subba Reddi

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346. A Janaki Sai and C Subba Reddi

Airborne poll&i • - - - , , - - - r.

f he. chief denors to the pollen geak in (August. were Poaceae, .^asuarina^ Cyperaceae
in that order. In February the chief donors were Casuarina, -.Rh'oenix and
Poaceae and to that in November, Poaceae, Dodonaea, Eucalyptus and Cyperaceae
contributed more.

The pollen of Poaceae, CyperacQae/.Amaranth-chenopod and Casuarina were
encountered all through the year. Poaceae pollen exhibited two distinct seasons :
one major and the other minor, the former practically started in early July with a
great uprush in the counts from late July to niid September folio wed' by1' a- gradual
decrease till the 2nd week of October. On the average, 50% of the-aiiimal catch
was recovered in this season. Peak incidence was recorded in the 2nd ;week of
August. The 'minor season can be said to have started by the 2nd \tfefck of
October and continued up to the 3rd week of December; maximal ntinrbe^'s- usually
occurred in November. The main Cyperaceae season occurred in' July-October ;
on average, 60 % of the annual datch was recovered here. Relatively 'high/numbers
of Amarantli-ChenopDd type were noticed during June-November peribd with
maximal incidence in September. With Casuarina, two periods of higher inci-
dence were apparent ; the main period extending from January to* April arid the
subsidiary from July to' October. Peak incidence occurred in February' during the
main and iti August during the subsidiary period ; on average, 80% and 19% of
the annual catch were recovered in the respective seasons. :

Pollen types like Asferaceae, Cocos and Ricinus were caught rather irregularly
all round the 'year. Types other than 'these were more or less confined to- a parti-
cular period of the year, thus the Syzigium pollen season was from late April
to early July 'with maximal incidence in June. Emblica pollen were caught from
early March to early June with higher numbers during April-May.' *Peliophomm
pollen were registered "from" March tb November (though not regularly) with
higher frequency from' April" to September. Tamarindus pollen were- -mainly
caught from May to August'1; greater frequency was from June rto 'July. -Muntingia
pollen season was from April* to- October ;. the numbers abounded in August-*
September. Mtmosoidcae pollen though encountered occasionally in. se^tfral-rftonths,
were rather more common from August to December. Eucalyptus pollen were
noticed from July to M^rch with maxima,! incidence during November-December,
The Dodonaea pollen season .extended from September until March, but was at
its maximum during November-January, Phoenix pollen began to appear in
air from late December, reached, a peak in February, decreased, through Mardj
$ndv, disappeared by April. Borassus pollen were trapped ia all months (though
not evejy year) except in 'December- January. They abounded during Marchr*
May with a ppak in Ajfril. ' ' / *.'.<•'•

The pollen of Holvptelea, Artemisia, Xanthium, Croton, Moms and Cicca were
of spasmodic occurrence over a major part of the year. '

3.6- Between-year' variation in 'pollen abundance ' ' ' ' ;

The different pollen types, recognised as well as total pollen counted, varied consi-
derably in their abundance froin year to ye.ar (tables 5 and 6)- The. catches 'of

' : :; Figure 4. Seasonal curves of maximum, minimuin abd avi&rage incidence -x>Jf major

.•,-;. :..' .•, pdlen.typeSasi wdi; as total pollen 'at Visakhapatnam, ' -^ ;•',' , / '

348 A Janaki Bai and C Subba Reddi

Table 6, The four-year average annual figure* and toe range of variation for the
airborne pollen

Pollen type

4-year average Range of
(Ko./sqom) variation (%)

Difference

SOYOSSUS

214

90-65-115*42

24-77

Ofcramineae

3263

84-71-115-35

30k64

Casuarina

1146

76-79-116-06

39-27

Phoenix

301

80-40-142-19

61-79

Amarantfrchenopod

301

58-80-125-25

66*45

feltophorum

193

72-54-145*60

73-06

MimoSoideae

56

67-86-142-86

75-00

Asteraceae

98

54-08-139-80

85-72

Cocos

73

67- 12~157- 53

90-41

Cyperaceae

536

74-81-171-64

96-83

Tamarindus

29

68-97-186-21

117-24

Dodonaea

338

39-64-162-13

122-49

Croton

25

52-00-188-00

136-00

Syzygiim

178

27-53-206-74

179-21

Eucalyptus

510

22-94-203-92

180-98

* Muntmgia

142

33-10-232-39

199-29

Emblica

99

40-40-275-76

235' 36

Artemisia

25

16-00-280-00

264-00

ffohptelea

53

15-09-320-75

30$- 66

Riciwts

130

12-31-327-70

315-38

Total pollen grains

8744

88-70-116-10

27-40

average of 3 years

Poaceae were higher in 1975-76 than in the other years monitored, and decreased
progressively in the subsequent years. These varied between 85% of 4-year
average in 1978-79 and 115% in 1975-76. The total weed pollen registered in
1976-77 were greater than in other years. They ranged from 68% to 158% of
average. The pollen of anemophilous weeds preponderated almost equally in
1976-77, and 1977-78. They ranged from 69% to 160% of average. The pollen
of entomophilous weeds of 1976-77 outnumbered others. They varied from
58% to 150% of average. The pollen of trees preponderated in almost equal
magnitude in 1976-77 and 1977-78. They ranged from 86-115% of average.
The total anemophilous as well as total entomophilous tree pollen were more in
1976-77 ; the former ranged from 95% to 104%, and the latter from 67% to
140% of respective averages. The total pollen preponderated in the 2nd year
of this study and ranged from 89% to 116% of average. From the range of
variations it becomes evident that group-wise, the pollen of trees and of grasses
remained relatively steady. Much more steady were the pollen of anemophilous
trees. This observation regarding tree pollen incidence is at variance with Dua

Airborne pollen

349

and Shivpuri (1962), Subba Reddi (1970), Mittre and Khandelwal (1973) from
India and Hyde (1972) from UK, Solomon and Buell (1969) from USA who
reported a high degree of annual variation.

Meteorological differences from year to year may account at least in part for
the recorded variation in annual pollen emission rates. This is especially so with
Poaceae and other weeds which grow and subsequently flower most abundantly
in response to the amount and distribution of precipitation. Vertical cylinder
traps are somewhat sensitive to wind speed and this may also account for some
of the observed variations, but the constancy of tree pollen counts suggest this
was not important.

3-7. Effect of urbanisation on airborne pollen

A comparison of the results of an earlier survey conducted during April 1966-
March 1968 (Subba Reddi 1970) and those of the present study revealed a consi-
derable decline in pollen abundance between the two study times (figure 5) indi-
cating that increasing urbanisation has had an effect on the airspora. Due to
increased industrialisation of the city the demand for residential houses increased
with the result the neighbouring suburban and rural areas have been intensively
used for extensive building construction. This has drastically curtailed the sites
available for plants especially of herbaceous flora.

12-

-

-

-

»_

O

-

-

0

o

9-

3

o

H

6-

CM

-

D

-

It

-•

-

D

*

•*"

-

-

6-

"

r~i

6-

~i

o

-

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-

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-

-

D

-

3-

3-

1-

i-

I-

n

nn

8-

4-

Totol pollen Gramineae.

Am-ch

Figure 5. The frequency of some pollen types at Visakhapatnam in six years from
1966 to 1979 (A, 1966-67 ; B, 1967-68 ; C, 1975-76 ; D, 1976-77 ; E, 1977-78; fr, 1978-79).

4. Conclusions

As is true of other tropical regions, no season was altogether free of pollen at
Visakhapatnam. Nor were any clear-cut tree, grass and weed pollen seasons
characteristic of spring, summer and autumn of temperate zones distinguished.
The different pollen types encountered, are to be considered as pernicious and
important pollutants of biological origin in the air of Visakhapatnam adding to

350 A Janaki Bal and C Subba Reddi

the overall daggers of atmospheric pollution of this growing industrial city. This
wan-ants an urgent inquiry into the clinical significance e of these biopollutants as
well as the interaction between these and the chemical pollutants, and the possible
resulting synergistic adverse effects on human health.

RefereaceS

Davis D J 1972rNIAID initiatives in allergy research ; /. Allergy Clin. ImmunoL 49 323

E>i*a K L £nd Shwpurj D N 1962 Atmospheric pollen studies in Delhi area in 1958-59 ;

/. Allergy 33 507-512
Duchaine J 1959 Allergy of the upper respiratory tract. In International Text-book of Allergy

(ed) J M Jamar Copenhagen pp. 154-195

Gregory P H 1973 The Microbiology of the Atmosphere 2nd (ed) Leonard Hill pp. * 377
Hirst J M 1973 A Trapper's line ; Trans. Br. Mycol. Soc. 61 205-213

Hyde H A 19-72 Atmospheric pollen and spores in relation to allergy I ; Clin. Allergy 2
' . 153-179 '

Hyde H A and Adams K F I960 Airborne allergens at Cardiff, 1942-59 ; Acta Allergol. 15

159-169

Hyde H A and. Williams I> A 1945 Studies in atmospheric pollen II. Diurnal variation in the
: incidence of grass pollen; New Phytol. 44 83-94

Hy^e H A and Williams D A 1961 Atmospheric pollen and spores as causes of allergic disease :
* hay fever, asthma and the aerospora ; Adv. Sci. (Lrondon) 11 525-533
Mittre V and Khandelmal A 1973 Airborne pollen grains and fungal spores at Lucknow during

1969-70 ; Palaeobotanist 22 111- 135

Newmark M F 1970 Recent developments in pollen Aerobiology; Ann. Allergy 28 149-152
Nilsson S and Nybom R 1978 Particles on pollen ; pp. 14-15, In International Aerobiology News

Letter (Special Issue) No. 8
Ogden E C, Raynor G S, Hayes J V, Lewis M D and Haines.J H 1974 Manual for sampling

airborne pollen ; (New York : Hafner Press) pp, 182

Ramalingam A 1962 The construction and use of a simple air sampler for routine aero-
biological surveys: Environ. Health 10 61-67
Solomon W 1976 Volumetric studies of aeroallergen prevalence I ; Pollens of weedy forbs

at a midwestern station ; /. Allergy Clin. ImmunoL 57 318-327
Solomon M A and Buell F M 1969 Effects of suburbanization upon airborne pollen ; Bull.

Torrey Bot. Club 96 435-444
Subba Rsddi C 1970 A comparative survey of atmospheric pollen and fungus spores at two

places twenty miles apart ; Acta Allergol. 25 189-215
Subba Reddi C 1974 A study of potentially allergenic pollen producing plants of Visakhapatnam ;

/. Palynol 1« 155-157
Subba Reddi C 1976 Floral mechanism, pollen productivity and pollen incidence in Madhuca

Mica Gmelin. with remarks on the mode of pollination ; New Botanist 3 11-15
Venkateswarlu J, Bhiravamurty P V and Narasimha Rao P 1972 The Flora of Visakhapatnam ;

Andhra Pradesh Akademi of Sciences, Hyderabad, pp. 260

Proc. Indian Acad, Sci. (Plant ScL), Vol. 91, Number 4, August 1982, pp 351-356
© Printed in India.

The floral anatomy of Kniphofia uvaria Hook. (Liliaceae: Kniphofieae)

N P VAIKOS and R M PAI

Plant Morphology Laboratory, Depaitment of Botany, Marathwada University,
Aurangabad 431 004, India

MS received 24 July 1981

Abstract. The floral anatomy of Kniphofia uvaria Hook, is described. The tepals
are anatomically similar and one-traced. The stamens are one-traced. The
outer whorl consists of shorter stamens. The placentation is parietal, nectary ovarian
and septal. The extension of the carpellary ventrals into the style is an important
anatomical feature. The trend towards development of an inferior ovary is noted.
Evidence from floral morphology and other disciplines is discussed and it is inferred
that the alleged affinity of Kniphofia and the Kniphofieae with the Aloineae and the
Hemerocallideae is rather remote.

Keywords. Floral anatomy ; Kniphofia uvaria.

I. Introduction

In earlier contributions, the floral anatomy of the Aloineae and Hemerocallideae
was presented (Vaikos, Markandeya and Pai 1978, 1981). The tribe Kniphofieae
is thought to chiefly comprise of the genus Kniphofia (cf. Stebbins 1971) although
two more genera Blandfordia and Notosceptmm are included in it by Hutchinson
(1959). The present paper deals with the vascular anatomy of the flower of
Kniphofia uvaria Hook.

2. Materials and methods

The flowering material was obtained from the Curator, Government Botanical
Gardens, Ootacamund. The flower buds were fixed in FAA. The usual paraflBn
method was followed. The microtome sections cut at 9-12/* were stained with
crystal violet using erythrosin as counter stain.

3. Observations

The pedicel contains a ring of three large bundles (figure 1). These are laterally
connected upwards to develop an anastomosis (figure 2). The six tepal strands

351

352 '• N--P Vaiko's and R M P<tir

emerge first (figure 2). The six staminal strands emerge quickly upwards
(figure 3). The remainder of the vascular tissue resolves into three carpellary
dorsals and six carpellary ventrals (figure 3).

The hypanthium is adnate to the ovary for a short length (figures 4, 5). The
tepals and stamens separate out simultaneously (figure 5). The tepals are united
into a tube for a considerable length (figures 5-11). Each of the tepals receives
a single vascular bundle (figures 5-11).

The stamens have cylindric filaments (figures 5-8). Those of the outer whorl are
antheriferous first and are also shorter (figures 9-10). The inner three stamens are
antheriferous at a much higher level (figures 10-11). Each of them receives a
single vascular bundle which continues upwards into the connective without a
division and ends beneath the tip of the anther (figures 5-10). The connective
splits and ends together with the anther lobes (figure 10). The anthers are introrse
and two-celled.

The ovary is trilocular at the base with the ovules arranged in two rows in each
loculus (figures 4-6). The carpellary ventrals of adjacent carpels are united to
form the composite placental bundles which are lodged on septal radii (figures 4-6).
These split into the constituent ventrals at the beginning of the ovuliferous zone
and bear traces to the ovules of adjacent carpels (figure 6). Upwards, the placen-
tae separate in the centre rendering the ovary unilocular (figure 7). The ventrals
of adjacent carpels extend at the inner end of the septa, continue to bear traces to
the ovules of adjacent carpels (figure 7) and enter the base of the style (figure 8).
The carpsllary ventrals end in the basal half of the style. .

The septal nectaries are developed from the base of the ovary (figure 4). These
open at the base of the style (figure 8). The ovarian loculus is continued upwards
into the style as a triradiate canal (figure 8). The stylar canal is closed in the centre
in some flowers to result in three cavities (figure 9). It is lined with transmitting
tissue. The carpellary dorsals extend into the style (figures 8-9) and end much
beneath its tip (figure 10). The style has three grooves along which it splits into
three non-vascular stigmatic lobes (figure 11).

3 • 1 . Abnormal flower

This flower has eight tepals, eight stamens and four carpels. The vascular supply
is derived in conformity with the tetramerous structure (figure 12). The septal
glands are four (figure 13). The style receives four carpellary dorsals and it is
four-lobed (figure 14).

4. Discussion

'. ' ' f . • ' ' • ' • "

The six tepals are arranged in two whorls and are united to develop a prominent
tube. Each of them receives a single bundle and, anatomically, both the whorls
are similar. This is a condition observed in many liliaceous genera, e.g., Ophiopogon,
Convallaria, Polygonatum, Maianthemum, Eucomis, Asphodelus, Urginea, etc. In
the allegedly allied Aloineae and Hemerocallideae (Vaikos, Markandeya and Pai
1978, 1981), the tepals are three-traced ; the three traces may arise separately

Floral anatomy of Knlphofia uvaria

353

from the stele or the laterals of the outer and inner tepals may.be derived through
a bifurcation of the common or commissural bundles.

The stamens are also one-traced ; this is. characteristic of most lilies studied.
The similarity in the vascular supply to the tepals and the stamens does not seem
to indicate a staminal origin for the perianth as is sometimes inferred (cf. Leinfellner
1963).

U

Kniphqfja uvaria Hook.

Figures 1-14. Knlphofia uvaria : 1-11. Transections of the flower from 'the base
upwards. 12-14. Transactions of the abnormal flower, showing teframerous
condition.

Abbreviations D, carpellary dorsal ; is, inner staminal strand ; L, lociile ; MIT*
median bundle of an inner tepal ; MOT, median bundle of an, outer tepal ; N,
nectary; os, outer staminal strand; PL? placental bundle; sc? stylar canal ;

' '' ' '

354 ff P Vaikos and R M Pal

The outer stamens are shorter than the inner ones. This condition is charac-
teristic of species of Aloineae, Allieae and some other genera (Markandeya 1978 ;
Vaikos 1974; Vaikos et al 1978, 1981). It represents a trend towards diffe-
rentiation of the two androecial whorls and ultimate reduction of one or more of
the shorter stamens or a whorl of stamens as occurs in Allieae (Markandeya
1978).

The outer floral whorls are adnate to the base of the ovary indicating a trend
towards the development of an inferior ovary. This is a trend which sporadically
occurs throughout the many genera and tribes of the family (Markandeya 1978 ;
Vaikos 1974 ; Vaikos et al 1978, 1981).

The gynoecium is tricarpellary and trilocular at the base and unilocular upwards-
The carpellary ventrals are lodged on the alternate septal radii and bear traces to
the ovules of adjacent carpels. The placentation is anatomically and morpho-
logically parietal (cf. Puri 1952).

The nectaries are ovarian and are typical septal glands. They develop at the
base of the ovary and open at the base of the style. The placental bundles which
lie close to the glands have to be associated with them in their function (Agthe
1951 ; Frei 1955 ; Pai and Tilak 1965).

The style receives the carpellary dorsals and the carpellary ventrals, although the
latter end early. The extension of the carpellary ventrals into the style is a less
advanced feature.

Tetramerous structure is noted in a few flowers. The vascular supply is derived
in conformity with tetramery. Tetramerous flowers occur normally in Aspidistra.

The genus Kniphofia forms a component of the Aloineae in the Englerian scheme
(cf Melchior 1964), whereas Hutchinson (1959) erects a tribe in its name. In the
old Bentham and Hooker's (1883) system it is placed under the tribe Hemerocalleae.

This paper demonstrates that the tepals in Kniphofia are one-traced while in
Aloineae and Hemerocallideae they are three-traced (Vaikos et al 1978, 1981). In
the studied plants of the tribes Aloineae and Hemerocallideae, the placentation
is axile, while it is parietal in Kniphofia.

In the taxa of the Aloineae studied, all or few stamens are adnate to the base
of the ovary and, as a variation, the stamens and the style may also be fused up to
the top to develop a prominent column-— the gynostemium (Vaikos and Markandeya
1976 ; Vaikos et al 1978). In Hemerocallis, the stamens are adnate to the
perianth (Vaikos et al 1981). However, in Kniphofia the stamens are neither
adnate to the ovary nor to the tepals.

Embryological evidence shows that Kniphofia is best treated distinct and not as
a component of the Hemerocallideae (cf. Di Fulvio and Cave 1964). The study
of vessel structure shows the vessels in Kniphofia are less specialised than those
in the Hemerocallideae, as also the Aloineae (Cheadle and Kosakai 1971).

Anthraquinones are detected in Aloe, whereas they are absent in Kniphofia (Van
Oudtshoorn and Van Rheede 1964).

The karyotype of Kniphofia is symmetric with n = 6 (Moffett 1932 ; Stebbins
1971). The Aloineae have the characteristic bimodal 4L + 3S karyotype (Brand-
ham 1971 ; Darlington 1963; Stebbins 1971), whereas Hemerocallis of the
Hemerocallideae has n = 11 and the karyotype is not very asymmetric and shows
rather a close similarity with Amaryllis (Sato 1942).

Floral anatomy of Kniphofia uvaria 355

Hutchinson (1959) considers the further development of the Kniphofieae to the
Aloineae. Cheadle and Kosakai (1971) consider this as a plausible suggestion
for they find the vessels in the Kniphofieae less specialised than those in the Aloi-
neae. Furthermore, " no member of the Aloineae has vessels less specialised than
those most specialised in the Kniphofieae ".

Stebbins (1971) infers a karyological affinity between the Kniphofieae and
Aloineae and suggests that, " increasing asymmetry and heterogeneity of the rela-
tively symmetrical karyotype of the Kniphofieae, consisting chiefly of the genus
Kniphqfia, together with the addition of a chromosome to the complement through
fixation of a centric fragment, would lead to the asymmetrical karyotype of the
Aloineae with n=7". This appears to be too speculative a statement at the
present stage of our knowledge and further karyological studies of more species
of the genus should be in order. It may be noted that the bimodal karyotype of
Aloineae is clearly marked with chromosomal markers for it. The present study
would demonstrate that Kniphofia is best placed distinct from the Aloineae as also
the Hemerocallideae. Further studies on more species of the genus are obviously
in order for a categorical conclusion.

Hutchinson's treatment of the tribe may also need a review. Di Fulvio and
Cave (1964) doubt the inclusion of Blandfordia in the tribe. Both Blandfordia
and Notosceptrwn merit a floral anatomical study.

Acknowledgements

The authors are grateful to Prof, K B Deshpande, ex-Head of the Botany
Department, for his encouragement. They thank Thiru G Kuppuswamy, Ootaca-
mund, for the supply of the material*

References

*Agthe C 1951 Uber die physiologische Horkunft des Pflanzennektars ; Ber. Schweiz. Bot. Ges.

61 240-277

Bentham G and Hooker J D 1883 Genera Plantamm Vol. HI Part 2, London
Brandham P E 1971 The chromosomes of the Liliaceae. II. Polyploidy and karyotype variation

in the Aloineae ; Kew Bull 25 381-399

Cheadle V I and Kosakai H 1971 Vessels in Liliaceae ; Phytomorphology 21 320-333
Darlington C D 1963 Chromosome botany and the origins of cultivated plants (London : Allen

and Unwin)
Di Fulvio T E and Cave M S 1964 Embryology of Blandfordia nobitis Smith. (Liliaceae), with

special reference to its taxonomic position ; Phytomorphology 14 487-499
*Fra E 1955 Die innerviernng der floralen Nektarien Dikotyler Pflanzenfamilien ; Ber. Schwiez.

Bot. Ges. 65 60-114

Hutchinson J 1959 The families of flowering plants H. Monocotyledons, Oxford
Leinfellner W 1963 Das Perigon der D'liacean ist Staminaler Herkunft ; Ost. Bot. Z. 110

448-467
Markandeya S K 1978 Morphological studies in the monocotyledons IV ; Ph.D. Thesis, Marath-

wada University

* Original not seen.

356 jf p Vaikos and R M Pal

Melchior H 1964 Liliaceae ; In Engler's Syllabus der Pflanzenfamilien II Bd. Gebruder Born-

traeger, Berlin
Moffett A A 1932 Studies on the formation of multiimclear giant pollen grains in Kniphofia ;

J. Genet. 25 315-336

Pai R M and Tilak V D 1965 Septal nectaries in the Scitamineae ; /. Biol ScL 81-3
Puri V 1952 PJacentation in angiosperms ; Bot. Rev. 18 603-651
Sato D 1942 Karyotype alteration and phytogeny in Liliaceae and allied families ; Jpn. J. Bot.

12 57-161

Stebbins G L 1971 Chromosomal evolution in higher plants (London : Edward Arnold)
Vaikos N P 1974 Morphological studies in the monocotyledons- HI. The Liliaceae ;Ph.D.Thesis

Marathwada University
Vaikos N P and Markandeya S K 1976 An incipient gynostemium in the Aloineae (Liliaceae);

Oar. Sci. 45 112-113
Vaikos N P, Markandeya S K and Pai R M 1978 The floral anatomy of the Liliaceae. The

tribe Aloineae ; Indian J. Bot. 1 61-68
Vaikos N P, Markandeya S K and Pai R M 1981 The floral anatomy of the Liliaceae. The

tribe Hemerocallideae /. Indian Bot. Soc. <50 222-231
Van Oudtshoorn M C B and Van Rheede 1964 Chemotaxonomic investigations in Asphodeleae

and Aloineae (Liliaceae); Phytochemistry 3 383-390

&:oc. loitau Acad. Sci. (Plant goi.), Vol. 91, Nfunibsr 4, AiigUst 19&», 0p. 3$7-37d
© Printed in India.

Transmission of seed-borne inoculum of Macrophomina phaseotina
from seed to plant

TRIBHUWAN SINGH and DALBIR SINGH

Department of Botany, University of Rajasth?n, Jaipur 302004, India

MS received 20 February 1981

Abstract. Macrophomina phaseolina is a serious pathogen which is externally as
well as internally seed-borne. It causes failure of seed germination and browning
and rotting of seedlings. The presence of pathogen in infected and healthy looking
seedlings was tested by clearing, sectioning and incubation techniques. After 8
weeks almost every surviving plant developed pale yellow to brown circular or oval
concentric spots on leaves, stem and capsules. Mycelium and micro sclerotia were
observed in the peripheral region of the lesions. Splitted root, stem and capsule
also showed the presence of microsclerotia. Cleared wholemounts of leaf and stem
and T.S. and L.S. of stem showed inter- and intracellular mycelium in cortex, xylem
and pith cells. Microsclerotia were also observed. In capsule, infection was
recorded on its inner wall, septum, placenta and seeds spreading from base to apex.

Keywords. Macrophomina phaseolina ; seed-borne transmission ; Sesamum indicum.

X. Introduction

Root, stem or charcoal rot caused by Macrophomina phaseolina (Tassi) Goid
is a serious seed-borne disease of sesame in India (Pearl 1923 ; Me Rae 1930 ;
Sundararamau 1931, 1932 ; Mehta 1951 ; Jain and Kulkarni 1965 ; Parasar
and Suryanarayana 1971 ; Mishra et al 1973 ; Gcmawatand Verma 1974 ; Verma
and Daftari 1974). It has been reported from many parts of the world and Meiri
and Solel (1963) regarded it as the most destructive disease of sesame in Israel.
It is soil as well as seed-borne (Noble et al 1958 ; Meiri and Solel 1963 ;
Richardson 1979). Singh and Singh (1979) have shown that in sesame M. phaseo-
lina is surface as well as internally seed-borne. The effects of seed-borne inoculum
of M. phaseolina on germination and the mode of disease transmission from
seed to plant have not been precisely investigated so far and therefore this study
was taken up.

2. Material and methods

Six samples ac. nos. 7, 9 (Ajmer), 15, 17, 18 (Chittorgarh) and 11 (Udaipur
investigated for deep location of M. phaseolina were selected for the study. Of

357

358 fribhuwan Singh and tidlbir $ingti

these ac . no. 11 was free from the infec ti on (figure 1) and used as c ontrol, whereas,
the remaining samples carrying various degrees of infection were categorized into
three groups (i) seeds without microsclerotia, (ii) seeds with moderate number
of microsclerotia, and (iii) seeds with abundant microsclerotia (figure 2). The
following three methods were used to investigate the transference of disease from
seed to plant. Seeds harvested in experimental plants were tested by standard
blotter method.

2-1. Petriplate method

One hundred seeds of each sample were treated with 2% available chlorine aqueous
solution of sodium hypochlorite and tested by standard blotter method. Obser-
vations were made on percentage of M. phaseolina infection, radicle emergence
germination, rotting and deformed seedlings, seedlings, with pycnidia andungerl
minated seeds at 24 hr intervals.

2-2. Seedling symptom test

Ten ml of 2% agar water was poured into each test tube and sterili23ed. One
hundred seeds of each sample after pretreatment with 1% available chlorine were
transferred aseptically to test tubes (one seed per test tube) and incubated for
8 days at 22° C under 12 hr of alternating cycles of day light and darkness.

2-3. Growth test

One hundred seeds of each sample untreated and pretreated with 2% and 5%
available chlorine for 5 min were sown in pots containing sterilized soil. Morta-
lity, seedling survival, symptoms and transmission of disease were recorded at
weekly intervals. Isolations were made from wilting and healthy looking seed-
lings and plants at regular intervals. For isolation, seedlings were carefully
uprooted, washed in running water, segmented and after surface washing with
1% available chlorine were spaced on blotters and incubated for 7 days. For
this study different parts, of seedlings (radicle, plumule, hypocotyl, cotyledon)
and plants (roots, stems, leaves, flower buds, flowers, green and dry capsules
and seeds) were incubated.

After 14 weeks dried plants were uprooted and the capsules were harvested
for further observations. The root system was separated, washed thoroughly and
splitted longitudinally. External and internal surfaces were observed by naked
eye and under stereobinocular microscope.

Stem portions were also split longitudinally and divided into three parts basal
middle and apical.

To detect the presence of M. phaseolina on the surface and inside the dried
capsules, they were kept in 70% ethanol overnight Capsule surface and its parts
showing microsclerotia were photographed while under 70% ethanol.

Clearing of plant parts carrying infection was made by using lactic acid,
potassium hydroxide and ethanol. Hand sections were also cut and stained with
safranin and fast-green combination. .

Seed-borne transmission in M. phaseolina

359

Figures 1-3. 1. Uninfected seed of sesame, 2. Seeds infected with Macrophomina
phaseolina ; note pin head like microsclero<ia on seed surface, 3. Leaves infected
with M. phaseolina ; note dark brown leaf spats with pale centre. Shot holes also
seen.

Seed-borne transmission in Af. pkaseotina 361

3. Results
3-1. Effect on germination and survival

In blotter test germination was only slightly affected (table 1). The radicle emerged
out in 19-29% seeds in five samples within 24 hr and in 55-75% seeds by 48 hr.
The germination ranged from 73-92%. Seedling abnormalities such as seedlings
without radicle and seedling with short radicle were observed and the abnorma-
lities ranged from 1-8%. Germination was found to be better in seeds with
superficial infection (ac. nos. 15,18) in comparison to other samples having a higher
percentage of embryonal infection (ac. nes. 7, 9 and 17). Pycnidia formation
was frequent on rotted seedlings and in ungerminated seeds.

The growth test using categorised seeds, seeds without microsclerotia and with
microsclerotia in five samples, showed that seedling emergence was considerably
affected (table 2) and ranged from 75-^82% in seeds without microsclerotia (untrea-
ted) and 56-78% in seeds with m'crosderotia. The germination was only 56-66%
in samples with deep infection and was 77 and 78% for remaining samples with
superficial infection. Survival of seedlings in samples with superficial infection
was also high. Seedlings obtained from seeds without nr.crosclerotia were more
vigorous to start with than those from mlcrocclcrotial seeds.

Chlorine pretreatment in growth test promoted emergence. Pretreatment with
2% Cl accelerated germination in both the categories and was 70-97% in seeds
without m'croselerotia and 60-93% in seed with microsclerotia. 5% Cl-pretrcat-
ment was phytotoxic and germination showed significant decline (61-88%) in seeds
without microsclerotia and was only 27-75% in seeds with microsclerotia. Seed,
ling survival was also higher in 2% Cl pretreatment (table 2). The Cl-pretreat-
nient (2%) brought about a slight improvement in germination even in the control
sample but the phytotoxic effects of higher concentration of Cl pretreatn^nt caused
36 and 37% decrease in germination and survival respectively as compared to
those of 2% Cl treatment. 5% Cl treatment probably inhibited root growth as
in petriplate experiment in 12-15% seeds. The embryo was observed to emerge
after rupturing the seed coat at the chalazal end. The cotyledons in such cases
turned green later on. The radicle remained lodged in the seed coat and its growth
was negligible.

3-2. Seedling symptom test

Three categories of seeds v/z., (i) seeds without m'croselerotia (ii) seeds with mode-
rate number of m'.cro:clerotia and (iii) seeds with abundant m'.croscltrotia were
used in the water agar seedling test (table 3). Seedling from first category showed
an initial vigorous growth but subsequently 20-25% of them became diseased.
Seeds of third category failed to germinate in sample ac. nos. 7,9,17 and gave
only 9 and 7 percent germination in seeds of ac. nos. 15 and 18 respectively,
Ungerminated seeds were covered with heavy mycelial growth of M. phaseolina^
In moderately infected seeds of second category the percentage of ungtrmiLattd
Seeds was 5-12. The seedlings were usually stunted as compared with those from
seeds without microsclerotia. The incidence of seedling infection of M. phaseo*
kna was high in ac. nos. 7,9,17 ranging from 51-70% and 37 and 19% in ac.
nos. 15 and 18 respectively. The percentage of deformed seedlings was also

P.(B)~S

362

Tribhuwan Singh and Dalbir Singh

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Seed-borne transmission In M. phaseollna

363

Table 2. Data an seedling emergence and seedling survival beyond 10 weeks in six
samples (I control and 5 infected) in pot experiment. 100 seeds used per treatment

Seeds without microsclerotia

Sample

number Emergence

Seeds with microsclerotia

Survival

Emergence

. , . Sur^iv&l

'

Un- Pretrcptcd

treated 2% 5%

Un- Pretreated
treated 2% 5%

Un- Pretreated
treated 2% ' 5% '

Un- iPrctrcated.
treated 2% 5%:

11 93

98

62

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(Control)

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93

75

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17 76

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61

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53

36

44

26

18 82

90

74

76

78

70

78

86

70

68

76

54

Table 3. Percentage of ungerminated seeds (US) and healthy (HS) deformed
(DS) aid infected seedlings (IS), infected with Mcicrophomina phaseollna^ from
nvw-mlaro *clerotiat and microsclerotial seeds having mild and heavy infection
(100 seeds/sample).

Sample
No.

Seeds

Non-microsclerotial

Microsclerotial

US HS DS IS

Mild

Heavy

US HS DS IS

US HS DS

IS

11
(Control)

... 100

7

H 60 9 20

12 26 11 51

100 00 00

00

9

9 56 13 22

10 10 10 70

100 00 00

00

15

7 62 9 22

8 53 6 37

91 3 00

6

: .17 .

10 . 50 15 25

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Tribhumn Singh and Dalbir Singh

in the former as

f the latter It may be mentioned
m and therefore, seeds.with micro-
of a high incidence of

in tho

test.

3-3. Symptoms and effect on host

• „ and rotting of seedlings in blotter

M . jtatffea caused seed rot, browiun^ ^^ ^^ at ^ root

(figuts 4) and water agar tests. The inxu u ^ rQot and fihoot tlps

St transition zone (fibres 5,6) ^ progress^ ito^ ^ ^ rf

(figure 7). Browning and damping of ^root toot pi ^^ ^ ^

kfshoot. ^^"Tf^Lwlpy in some cases when the seed coat
on the whole root turned bro™ ^* C0tyledons during germination and
having **<<f f t^^ downwards from

remain attached with for a longel peno ofmicrosclcrotia wasqmte

l-ral roots) system (^s 8, 9).

Figures 4-9. Diagrams to show symtoms of M. phaseolina on sesame ^

in blotter test. 4. Rotting of seed with pycmdia and imcrosderotia* 5, o,
Soedlings 'showing streaks and microsclerotia. 8,9. Seedlings showing disti
batioa of microsclcrotia ; note infection of primary w .yrolt as secondary toots m C*

Seed-borne transmission in M. phaseolina 365

In growth tests first disease symptoms were observed on the cotyledons after
a fortnight. In a few days (by 20th day after germination) these became well
marked as pale yellow to brown or blackish circular or oval and concentric spots.
Each spot had a whitish area in the centre. The spots enlarged, became irregular
and necrosis caused shot-holes. Similar spots were recorded on leaves after six
weeks of germination (figure 3). Infected plants were weak and had leaves smaller
than those on healthy plants. After 8 weeks almost every surviving plant from
untreated as well as Cl-pretreated seeds developed symptoms of M. phaseolina.
infection on stem appeared as yellowish brown discoloured patches. Their peri^
phery turned dark brown to black subsequently (figure 12). Pedicel and green
.capsules also revealed the occurrence of yellowish patches.

Under stereobin ocular microscope mycelial growth was observed on leaf, stemf
pedicel and capsule. Although no microsclerotia were seen on leaf surface, they
occurred in abundance in black peripheral region of the patches on stem. Scattered
microsclerotia were also seen on other parts of stem.

3-4. Isolation and observation of fungus

Isolations were made from seedlings and plants raised from untreated and
pretreated seeds at regular intervals. They wera classified into two categories
viz., rotted and healthy looking seedlings. The two types of seedlings, were treated
separately for incubation. For the latter, 25 seedlings were used but for the rotted
all the seedlings available on the day of setting the experiment were used which
were usually more than 10.

Ten-day old rotting seedlings yielded M. phaseolina from all the parts including
cotyledons. The incidence was more than 80% and usually higher in transition
zone than the other components. Rotted seedlings from pretreated seeds yielded
the pathogen in slightly low incidence in comparison to those of untreated seeds.
In healthy looking seedlings, M. phaseolina was recorded in not more than 41%
cases. Interestingly in such cases the incidence of pathogen recovery was higher

"in sample nos. 7 and 17 as compared to that of sample nos. 9, 15, 18.

Similar data were also collected on 50-day old plants which yielded M. phaseolina
in very high percentage. No significant difference in percentages was recorded
in plants from untreated and pretreated seeds in all the five samples. The in-
fection percentage on different plant parts were also comparable,

Mature but not dead dry plants (75-day old) yielded the fungus in 100% cases
from roots (figure 10) and transition zone in sample DOS. 7,9,17 and 18. It
was found to be 80 and 83 (roots) and 87 and 86 (transition zone) in plants
from untreated and pretreated seeds of sample no. 15. The recovery of fungus
was usually high from stem and leaves than the other plant parts i.e., flower buds*
flower, (figure 11) and capsules.
Af. phaseolina was also recovered in very high percentages from root,stem, whole

-capsule and its parts (Placenta, seed and septum) from dry plants in blotter test.
Visibility of microsclerotia was slightly difficult on the hairy surface of capsule,

.whereas, the placenta and septum turned black.

* Dry plant parts were also examined for the presence of pathogen. Dry plants
of all the .five samples were harvested, and examined with unaided eye or 10 x

*ha&dtens.. Microsclerotia were observed on and inside the tap root and stem from

366 Tfibhuwan Singh and Dalbir Singh

base to tip (table 4). Lateral roots could not be split. The incidence of micro*
sclerotia on roots and basal part of stem surface ranged from 78-100% and
7(MOO% respectively. In the in ten or part of root and basal stem it ranged from
9-^14% and 24-^53% respectively. Maximum infection (86%) was recorded on
the entire surface of the root and stem (figure 12) (base to apex). However, a
gradual decrease in its incidence was recorded inside the stem from its basal to
the apical part (figure 13), incidence being 53% (base), 33% (middle) and 22%
(apical part). In the remaining sample apical region yielded low percentage of
infection on the outside as well as inside.

The surface and the interior of the capsules showed abundant microsclerotia
on different parts including seeds (figures 14,15), A higher incidence of micro-
sclerotia was observed in the basal and distal region of capsule surface (outer and
inner), septa, placenta and seeds (table 5). Most of the seeds showed abundant
microsclerotia and a thick net of black mycelium on the micropylar region
(figure 15). The chalazal end was either free or showed mild infection. In septum
the incidence of fungus was rather low (19-67%).

3-5. Clearing and section cutting

Cleared wholemount preparations of infected leaves showed abundant dark brown,
branched and septate mycelium running across the leaf spot (figures 16,17). It
traversed mostly parallel to veins* The host cells in infected regions were small,
poor in pigments, cytoplasm and food materials. Microsclerotia were not
observed*

The wholemount preparations (figure 18), transactions and longisections

(figures 19,20,21 and 22) showed the presence of mycelium in different tissues

including xylem (figures 21,22) and pith ; inter and intracellular mycelium was

, observed to travel along the vessel length (figure 22). Microsclerotia were also

observed in the pith cells (figure 19) and vessels (figure 20).

Seeds harvested from experimental plants also showed the occurrence of micro-*
sclerotia and mycelium in seed coat, endosperm and embryo as revealed by
maceration and sections. .

4. Discussion

Water agar seedling test and the growth test have revealed the transmission of
M. phaseolina from seed to plant in all the five sesame seed samples tested pres-
ently. The fungus was isolated from rotting, wilting and healthy looking seed*
lings. The vegetative parts, floral btids, flowers and green fruits of experimental
plants also yielded the fungus on incubation. Careful examination of root, stem
and fruits from mature and dried plants showed the presence of mycelium and
microsclerotia. The fungus was recovered from pedicel, calyx, septum, placenta,
inner surface of capsule and seeds attached to the placenta. The infection usually
spread from nr.cropylar end onwards in the seeds of infected capsules. The
systemic transmission of Af. phaseolina from infected seeds is strongly supported.
The mycelium, running longitudinally as well as horizontally and the microsclero-
tia, were recorded in pith, cortex and vascular cells in stem. The pith became
hollow in infected plants probably due to the production of cellulolytic and pecto-*

Seed-borne transmission in M. phaseoih

ma

367

Figures 10-15. 10, 11. Root and flower after incubation on blotter. 10. Root
surface showing pycnidia and microsclerotia. 11. Bilabiate flower surface of sesame
showing microsclerotia. 12-15. Stem and capsule of sesame showing infection of
M. phaseolina in dried plants harvested at the end of growing season. 12. Surface
view of healthy and infected stem. 13. Longitudinally split stem showing micro-
sclerotia. 14. Partially dehisced capsule ; note the microsclerotia on surface.
15, Interior of an infected capsule. Note the infection of placenta and seeds.

368

Trlbhuwan Singh and D alb! r Singh

Figures 16-22. 16-18. Wholeniounts of cleared infected leaf and stem peeling. id-
Part of leaf surface showing mycelium in stamatal region. 17. Part of leaf surface
showing septate mycelium and its association with vein. 18. Cleared peeling,
from stem surface showing micro sclerotia and inter- as well as intracellular nrycelium
19-22. T.S. and L.S. of infected stem. 19. T-S. stem showing mycelium and
microsclerotia. 20-21. T.S. stem showing mycelium and micros clerotia in vessel,
22. L,S, stem ; note mycelium traversing in vessel,

Seed-borne transmission in M. phaseolina

369

Table 4. Percentage occurrence of raicrosclerotia of Macrophomina phaseolina
vcgetafwc parts of dried and harvested plants.

Vegetative
parts

Root

Stem

sample No.

tap

Basal

Middle

Apical

Surface

Inside

Surface

Inside

Surface

Inside

Surface

Inside

7

86

13

86

53

&6

33

86

2C

9

100

10

100

35

100

30

75

15

15

&2

9

76

26

41

5

30

5

17

7g

11

70

24

60

15

40

11

IS

100

14

92

40

82 ,

34

60

10

Table 5. Percentage occurrence of Macrophomina phaseolina on dried capsule parts
(after soaking in 70% ethanol for 24 hr)

Capsule

Placenta

Capsule

parts — -

sample Outer surface Inner surface Basel Distal Basal . Distal

Seeds Septum

No. Stalk

Basal Distal Basal Distal

J

84

82

73

80

40

80

33 •

93

33

26

9

80

70

70

65

50

70

42

70

40

30

15

69

67

• 67

,56

33

60

30

73

45 •

20

17

63

52

35

47

40

51

40

50

30

19

18

92

91

89

85

77 •

92

79

81

71

67

lytic enzymes in large quantities. This also accounts for easy breaking of stem
in these plants.

M. phaseolina is a well-known soil inhibitant and Cougnee (1963) reported
the disease to be soil borne, Mehta (1951) suspected the possibility of transmis-
sion of Sclerotiniabataticola by sesame seed in India. "Noble et al (1958) recorded
the seed-borne nature of M. phaseolina on the basis of investigation of Mehta in
their list of seed borne diseases, but were doubtful.1 Meiri and Solel (1963)
observed that infected seeds of < Ranner 15 > yielded diseased seedlings and
also observed the first site .of infection in the collar region.

Fakir et al (1976) working with sunflower kernels have shown that M. pha-
seolina is highly pathogenic and pathogen could be isolated from seeds of some
of the inoculated plants, but seeds of symptomless sunflower plants from naturally
infected seeds did not yield M. phaseolina. "This is in contrast to the present "study
and does not support the transmission of disease from seed to seedling. However*
in their description it is not clearly stated whether these symptomless plants grew
out of microsclerotial seeds. The present study not only provides the adequate

370 Tribhuwan Singh • and Dalbir Singh

evidence of seed transmission of M. phaseolina in sesame but also clearly reveals
that the seed-borne inoculum plays an important role in the spread as well as in
the manifestation of disease.

Pycnidia formation was rare in blotter as well as agar test. Only on rotted seeds
and seedlings, and in lesions on stem, pycnidia were frequently formed. Chidam-
baram and Mathur (1975) reported the production of pycnidia in 20 out of 58
non-sporulating isolates in water agar nr.dia. They tested 58 isolates of
M. phaseolina from 20 plants. Present isolate from Sesamum Indicum could be
described as a moderately sporulating one.

Acknowledgements

The authors thank the Council of Scientific and Industrial Research, Now Delhi
for providing a research fellowship to one of them (T.S.).

References

Chidambaram P and Mathur S B 1975 Production of pycnidia by Macrophomina phaseolina ;

Trans. Br. Mycol. Soc. 64 165-167
Cougnee M 1963 fitudes sur L' arrthraenase des Cotoniers ; G. barbadensc ; Cotton ct Fibres

Trop. 18 149,450
Fakir Golara A, Rao M H and Tliiruraalachar M J 1976 Seed transmission of Macrophomina

phaseolina in sunflower; Plant Dis. Rep. 44 221
Gemawat P D and Verma O P 1974 Root and stem rot cf Sesamum in Rajasthan. Evaluation

of varieties (Macrophomina phaseolina) Indian J. Mycol. Plant Pathol. 4 76-77
Jain A C and Kulkarni S N 1965 Root and stem rot of Sesamum ; Indian Oil Seeds J. 9

201-203
McRae W 1930 Report of the Imperial Mycologist Scient ; Rep. Agric. Res. Inst. Pusa

1928-29 51-56
Melita P R 1951 Observations on new and known diseases of crop plants of Uttar Pradesh;

Plant Prat. Bull New Delhi 3 7-12
Meiri Aliza and Solel Z 1963 Transmission of charcoal rot by sesame seeds ; Phytopathol.

Mediterr. 2 90-92
Mishra R P, Singh B P and Joshi L K 1973 Pod susceptibility of different varieties of till

(Sesamum orientale) to Macrophomina phaseoli (Maubl.) Ashby ; J.N.K.V.V. Res. J. 7

2S8-289

Noble Mary, De Tempo J and Neergaard P 1958 An annotated list of seed-borne diseases ;
-• C.M.L Kewl59
Richardson M J 1979 Annotated list of seed-borne diseases ; ?rd edition, Proc. Int. Sqed Test.

Ass. 23 1-320
Parasar R D and Suryanarayaaa D 1971 Polygalecturooase activity of Macrophomina phaseolina

the incitant root rot of Sesanum "indicum ; Indian Phytopath. 24 559-562
Pearl R J 1923 Report of the mycologist to the Govt. of the* Central Provinces and Berar.

Rept. D:ptt. A^ric. Cettral Provinces and Berar for the year ending 30th June 1922 19-20
Singh T and Singh D 1979 Anatomy of penetration of Macrophomina phaseolina in seeds of

sesame ; Recent researches in plant sciences (ed.) S S Bir (Ludhiana : Kalyani Publisher)

pp. 603-606
Sundarajraman S 1931 Administration report of the mycologist for the year 1929-30 ; De.pt. Agric.

Madras •

Suodararaman S 1932 Admbtstration report of the mycologist for the year- 1930-31 ; Dept.

Agric , Madras .
Verma O P and Daftari L N 1974 Amount of seed-borne inoculum of Macrophomina phaseolina

and its effect on mortality and growth of Sesamum seedlings ; Indian Phytopath. 27 130-131

Proc. Indian Acad. Sci. (Plant Sci.), Vol. 91, Number 5, October 198?, pp. 371-378.
© Printed in India.

Effect of water stress and sucrose on opening and longevity of
flowers in gladiolus

I V RAMANUJA RAO and H Y MOHAN RAM

Department of Botany, University of Delhi, Delhi 110007, India

MS received 20 January 1982 ; revised 9 August 1982

Abstract. The percentage of buds opening and flower longevity as affected by the
availability of water and sucrose to cut spikes of gladiolus were studied. Uptake
of sucrose solution and fresh weight changes in spikes were dependent on sucrose
concentration. Marked reduction in uptake and fresh weight occurred when poly-
ethylene glycol (PEG) was used as the stressing agent. In comparison, PEG failed
to induce any significant change in the percentage of flower buds op.en.ing.
Sucrose was essential for opening since the buds that failed to open in the control
were caused ta open in sucrose. Induced water stress did not curtail flower
longevity at any given concentration of sucrose. Thus flower opening and longe-
vity in gladiolus appear to be limited more by the availability of sucrose than
water.

Keywords. Flower longevity ; flower opening ; gladiolus ; polyethylene glycol ;
sucrose ; water stress.

1. Introduction

Studies have been carried out on the factors affecting water uptake and vascular
blockage in cut flowers on account of their crucial role in maintaining freshness
(Durkin and Kuc 1966 ; Marousky 1969 ; Oilman and Steponkus 1972 ; van Meeteren
1978 ; Rao and Mohan Ram 1982a). Water deficit causes early wilting of flowers
(Marousky 1969; Paulin 1972; Mayak et al 1974). Mayak et at (1974) noted a
sharp decline in the water potential of petal tissues in wilting cut roses but not in
intact flowers. Water stress has been identified as the cause of failure of flower
opening in the spikes of iris stored at low temperature for four days and
then for one additional day at 22° C (Mayak and Halevy 1971). Appreciable bud
opening has, however, been recorded in gladioli and chrysanthemums stored in cold
using sucrose alone or in combination with silver nitrate or gibberellic acid
(Kofranek et al 1975; Kofranek and Halevy 1976; Rao and Mohan Ram 1979,
1981, 1982b).

In spite of the development of successful techniques to handle cut flowers,
our understanding of water requirement of opening flowers and their ability to
withstand storage has remained incomplete. The property of sucrose to act as an
antidesiccant when supplied before storage, in addition to its metabolic role is

i

371

372 / V Ramanuja Rao and^H Y Mohan Ram - '

still unclear. For example, gladiolus spikes, given a pulse treatment with sucrose
before storage, open satisfactorily on subsequent transfer to water (Mayak et al
1973; Bravdo et al 1974). However if the spikes are first stored dry, it becomes
necessary to provide gibberellin plus sucrose subsequently to ensure full opening
(Rao and Mohan Ram 1979, 1982b). This paper discusses the importance of
water and sucrose in flower opening and longevity.

2. Material and methods

Spikes of Gladiolus natalensis Hort. were obtained from a commercial grower in
New Delhi at the green-bud stage (harvested one day before the corolla of the
lowermost bud emerged from the enveloping bracts and the tip just became visible).
The spikes were stored dry for 24 hr after harvest at 20° C to facilitate a larger
uptake of the pulsing solution (Rao and Mohan Ram 1981a). Twenty spikes
each were pulsed for 48 hr with sucrose solution (0'25 M and 0-5 M) to eliminate
any effects caused by a low amount of carbohydrates in the spike. These were
then transferred to (i) water (control), (ii) sucrose solution of the same concen-
tration as was used for pulsing and (iii) polyethylene glycol solution (PEG, MW
6000, Sigma Chemical Co., USA) having a water potential similar to the pulsing
solution (0 • 25 M sucrose = — 7 bars ; 0 • 5 M sucrose = —15 bars). In all six sets,
each with 20 spikes were set up in glass tubes (2*5 x 15*0 cm) containing 40 ml
of the test solution. The solutions were prepared using glass-distilled water.
One additional set of spikes held continuously in water served as the control.
The spikes were kept in a chamber at 20 ± 2° C with 14 hr photoperiod (under
cool-white daylight fluorescent tubes giving 500 lux).

The number of flowers opening and withering per spike were recorded daily. The
longevity of individual flowers on the spike (the period between flower opening
and withering) was recorded. The term 'uptake * used here refers to the volume
of the solution taken up and 4 fresh weight change ' to the differences in fresh
weight of a spike over a given period. Tubes containing different solutions but
without spikes served as controls to measure loss caused by evaporation,
Confidence intervals of the means were determined at P < 0*05.

3. Results

3.1. Rate of uptake

• During the period of pulsing a high initial uptake was recorded for the control
(table 1). Spikes kept in 0-25 M sucrose showed a greater uptake than those in
0-5 M during this period. By day 2 the spikes pulse-treated with sucrose and
transferred to water took up a larger volume of solution than those continuously
held in sucrose and the control.' A marked decrease was observed in the amount
taken up by the pulsed spikes on day 4 as compared to that on day 2. It was,
however, still higher than that for spikes kept continuously in sucrose. A decrease
in uptake was noted in the latter during the same period, although it exceeded

Effect of water stress and sucrose on flowers of Gladiolus 373

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374 / V Ramanuja Rao and H Y Mohan Ram

the amount in the pEG-treated spikes. Spikes transferred from sucrose (both
concentrations) to PEG showed practically no uptake during the first two days.
Subsequently, however, a very low uptake was recorded in these spikes ; the
uptake by spikes held in PEG at —7 bars was slightly higher than that
at —15 bars.

During the entire period of experimentation pulse-treatment with sucrose
(0*25 M) followed by transference to water resulted in a significantly greater
uptake over the control (table 1). However, in the spikes held continuously in
sucrose the magnitude of uptake was lower. Treatments with PEG markedly
curtailed uptake as compared to the control and other treatments,

3.2. Changes in fresh weight

As with uptake, the maximal fresh weight increment during the pulsing period
was noted in the control, followed by that in spikes treated continuously or pulsed
with sucrose at 0*25 M and 0'5 M, respectively (table 2). Whereas after two days
the control spikes attained a negative fresh weight change, the spikes which were
pulsed with sucrose and transferred to water or sucrose continued to show
a positive fresh weight change. Spikes transferred to PEG showed a negative
fresh weight value, much lower than that of the control. In general the spikes
which were held continuously in 0'5 M sucrose showed a lower fresh weight change
as compared to the spikes treated with 0-25 M. In all the treatments, the lowest
fresh weight change was recorded on days 6 and 8.

The overall highest fresh weight was observed in spikes pulsed with sucrose
at either concentration and transferred to water (table 2). Thus the control exhi-
bited the lowest fresh weight. Low fresh weight was also observed in spikes
treated with sucrose (0'5M) continuously or stressed with PEG at either con-
centration.

3.3. Percentage of flower buds opening

A significantly higher percentage of flower buds opened in all the treatments
over the control (table 3). Among the treatments the percentage of flower
buds opening did not vary except in spikes pulsed with sucrose at both the
concentrations and transferred to PEG which showed slightly lower opening as
compared with those pulsed with sucrose (0"25M) and transferred to water.

3.4. Flower longevity

A study of the longevity of individual flowers at different positions on the spike
showed differences between the control and those pulsed with sucrose and trans-
ferred to either water or sucrose (table 4). In spikes stressed with PEG at both
the concentrations there was a continuous increase in longevity from flowers 1-5.
Interestingly when the mean longevity of flowers 1-5 was compared, it turned
out that the lowest longevity was recorded for the control (2 '8 days). The longe-
vity of flowers in spikes pulsed with a particular concentration of sucrose and

Effect of wafer stress and sucrose on flowers of Gladiolus 375

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376 / V Ramamija Rao and H Y Mohan Ram

Table 3. Effect of' water stress on the percentage of flower buds opening in
gladiolus.

Treatments

Control SlW S2W SIC S2C SlP S2P ~~

x±cr x±ci x±cr x±ci x±ci x±cr x±cr

54-0 3-0 80*0 5*0 75-0 5'0 76*0 4-0 75'0 4-0 69'0 3'0 69-0 4-0

X Mean values.

CI Confidence interval calculated at P^O'05.

SlW, S2W Spikes pulsed with 0-25M and 0-5M sucrose for 48 hr and later transferred to

water.

S1C3 S2C Spikes held continuously in 0-25M and 0-5M sucrose.
SIP, S2P Spikes pulsed with 0'25M and 0-5M sucrose for 48 hr and later transferred to

PEG at. — 1 bars and — 15 bars, respectively.

Table 4. Effect of water stress on flower longevity* in gladiolus.

Treatments

Flower
number

Control

SlW

S2W

SIC

S2C

SIP

S2P

1

2-6

3'8

4-2

4-0

4-2

3-2

3-8

2

2*6

3"6

4-2

4-0

4-2

3'4

4-0

3

2-6

3-4

4-2

3'8

4-6

3'6

4-6

4

3-0 '

3-6

4-2

3-8

4-8

3-8

4'8

5

3-2

3'8

4.4

4-0

4-6

4-0

5-0

6

2-6

3-8

4-4

4-4

4-4

4-6

7

4-0

...

...

X

2- & .

3-6

4-2

3-9

4-6

3-6

4-4

* in days.

X Mean longevity of flowers (1-5).

SlW, S2W Spikes pulsed with 0*25 M and 0'5M sucrose far 48 hr and later transferred to

water.

SIC, S2C Spikes held continuously in 0'25M and 0* 5 M sucrose.
SIP, S2P Spikes pulsed with Q'25M and 0'5M sucrose for 48 hr and later transferred to

PEG at — 7 bars and — 15 bars, respectively.

transferred to water, sucrose or PEG was more or less similar. For example/ at
0*25 M of sucrose, it ranged from 3-6 to 3 -9 days and at 0'5 M of sucrose it
varied between 4 -2 and 4-6 days in different treatments.

Effect of water stress and 'sucrose on flowers of Gladiolus 377

4. Discussion

A study of the effect of water stress on gladiolus indicated that uptake of the
test solution and fresh weight change during the initial period of pulsing with
sucrose were related to the concentration of the solution. Whereas the lower
concentration of sucrose itself reduced initial uptake compared to the control,
doubling it did not result in a proportional decrease. A similar result has been
recorded by Bravdo et al (1974) who observed uptake even from a solution with
50% sucrose concentration. The subsequent absorption of liquid was dependent
not only on the water potential of the transfer solution but also on the nature
of the transfer osmoticum. Thus, when the spikes were held in PEG instead of
sucrose of similar water potential, there was a steep drop in uptake.

The fresh weight change of the spikes showed a direct relationship with the water
potential of the transfer solution. The sucrose-pulsed spikes which were transferred
to water maintained higher fresh weight. Halevy and Mayak (1974) have shown
that sucrose decreases the water potential of the petals and enhances their ability
to absorb water.

In comparison with water, the uptake of which was markedly curtailed by
PEG, the availability of sucrose was found to be a major factor in bud opening.
In all the treatments a higher percentage of opening over the control was ob-
tained. Stress pronouncedly affected uptake and fresh weight but not the per-
centage of flower buds opening. This is quite remarkable in the light of the
finding by Goldschrnidt and Huberman (1974) that citrus petals have a very large
water requirement during opening (highest fresh weight was recorded) and in
view of the reported failure of flower bud opening under water stress condi-
tions (Mayak and Halevy 1971), Thus, flower opening in gladiolus appears to
be limited more by the availability of sucrose than water, especially because of
the ability of the newly opening buds to draw out water from the older open
flowers and cause their premature withering (Rao and Mohan Ram 1982a). Our
recent study has also shown that green-bud spikes lack adequate reserves of carbo-
hydrates and that this is one of the principal causes of poor opening (Rao and
Mohan Ram 1981).

It is significant that in the present work induced water stress did not curtail
flower longevity at any given concentration of sucrose. In addition to its role
as a respiratory substrate (Coorts- 1973), -sucrose has been shown to enhance the
effect of cytokinins, and counter the deleterious effects of ethylene and abscisic
acid (Borochov et al 1976a; Mayak and Dilley 1976). Sucrose also reduced
the endogenous levels of abscisic acid in cut rose flowers (Borochov et al 1976b).
Spikes treated continuously with sucrose showed higher longevity than PEG-treated
spikes probably because of greater availability of sugar.

References

Borochov A, Mayak S and Halevy A H 1976a Combined effects of abscisic acid and sucrose
on growth and senescence of rose flowers ; Physiologia Plant. 36. 221-224

Borochov A, Mayak S and Halevy A H 197 6b Abscisic acid content of senescing petals on
cut rose flowers as affected by suerose and ; water stress ; PI. Physiol 58 175-178

378 / V Aamanuja Rao and H Y Mohan Ram

Bravdo B, Mayak S and Gravrieli Y 1974 Sucrose and water uptake from concentrated sucrose

solutions by gladiolus shoots and the effect of these treatments on floret life ; Can. J.

Bot. 52 1271-1281

Coorts G D 1973 Internal metabolic changes in cut flower? ; Hort. Sd. 8 195-198
Durkin D and Kuc R 1966 Vascular blockage and senescence of the cut rose flower ; Proc.

Am. Soc. Hortic. Sti. & 683-689
Oilman K F and Steponkus P L 1972 Vascular blockage in cut loses ; /. Am. Soc. Hortic.

Sci. 97 662-667
Goldschrnidt E E and Huberman M 1974 The co-ordination of organ growth in developing

citius flowers : a possibility for sink type regulation ; J. Exp. Bot. 25 534-541
Halevy A H and Mayak S 1974 Improvement of cut flower quality, opening and longevity

by pre-shipment treatments ; Acta Hortic. 43 335-347
Kofranek A M, Halevy A H and Kubota J 1975 Bud opening of chrysanthemums after long

term storage ; Hort. Sci. 10 378-380
Kofranek A M and Halevy A H 1976 Sucrose pulsing of gladiolus stems before storage to

increase spike quality ; Hort. Sci. 11 572-573
Marousky F J 1969 Vascular blockage, water absorption, stomatal opening and respiration

of cut "Better Times" roses treated with 8-hydroxyquinoline citrate and sucrose ; /. Am,

Soc. Hortic. Sd. 94 223-226
Mayak S and Halevy A H 1971 Water stress as the cause for failure of flower bud. opening,

in iris ; /. Am. Soc. Hortic. Sd. 96 482-483
Mayak S, Bravdo B, GuilU A and Halevy A H 1973 Improvement of opening o.f cut gladioli

flowers by pretreatment with high sugar concentrations ; Scientia Hortic. Amsterdam 1

35T-365
Mayak S, Halevy A H, Sagie S, Bar-Yoseph A and Bravdo B 1974 The water balance of

cut rose flowers ; Physiohgia Plant. 31 15-22
Mayak S and Dilley D R 1976 Effect of sucrose on responses of cut carnations U> kind in

ethylene and abscisic acid ; J. Am.Soc. Hortic. Sci. 101 583-585
van Meeteren U 1978 Water relations and keeping-quality of cut Gcrbera flowers. 1. The

cause of stem break ; Scientia Hortic. Amsterdam 8 65-74
Paulin A 1972 Influence d'un deficit temperaire en eau sur le metabolisms azote des flours

coupees d'/m gennanica ; C. r. Hebd. $eaitc. Acad. Sci.9 Paris 275 209-212
Rao I V Ramanuja and Mohan Ram H Y 1979 Interaction of gibberellin and sucrose in

flower bud opening in gladiolus ; Indian J. Exp. Biol. 17 447-448
Rao I V Rarcanuja and Mohan Ram H Y 1981 Nature of differences between green-bud and

tight-bud spikes of gladiolus : basis for a postharvest bud-opening treatment ; Indian J.

Exp. Blol 19 1116-1120
Rao I V Ramanuja and Mohan Ram H Y 1982a Prevention of compensatory withering and

stem-break in cut spikes of gladiolus by overcoming vascular blockage ; Proc. Indian Natl

Sci. Acad. Part B (in press)
Rao I V Ramanuja and Mohan Ram H Y J982b Specificity of gibberellin and sucrose promoted

flower bud growth in gladiolus ; Ann. Bot. 50 473 --479

fooc. Indian AcacL Sci. (Plant ScL), Vol. 91, Number 5, October I$g2, pp. 3t9-38&
© Printed in India.

Petal venation in Trigonclla (papilionaceae)

MOHINI GUPTA

Department of Botany, Institute of Advanced Studies, Meerut University,
Meerut 250001, India

MS received 7 March 1981 ; revised 12 June 1982

Abstract. Petal venation of nine species of Trigonella has been worked out. A
positive correlation has been found between length or area and the number of
dichotomies but no correlation is found with breadth. In all the species corolla is
of simple type except T. polycerata in which it is of medicagoid type. Among
the different types of anastomoses C and D types are of most frequent occurrence
and other types are species specific with a low range of variation.

Keywords. Petal venation ; papilionaceae ; trigonella.

1. Introduction

The analysis of petal venation received the attention of various workers in tracing
phylogeny after the remark that petal venation shows both simplicity and diver-
sity. Petal venation of regular corolla received the attention of various workers
(Arnott and Tucker 1963, 1964 ; Banerji and Mukherji 1970 ; Banerji 1972)
but that of irregular corolla received only the attention of Datta and Saha (1968)
and Subramanyam and Nair (1973). Datta and Saha (1968) reported important
differences at specific level in types of anastomoses and their pattern of distribution
on standard, wing and keel petals of four species ( Butea frondosa, Cajanus cajan,
Dolichos lablab and Erythrina indica) belonging to tribe phaseolac. Hence from
the perusal of the literature it appears that venation of petals may be significant
at specific level and since ho work is available on the species of Trigonella^ there-
fore, in this paper petal venation of nine species has been worked out.

2. Materials and methods

Flower buds for the present study were either collected locally or procured from
places .as mentioned below :—• .

Species Place of collection

1. Trigonella arabica Dehile Bet Dagan, Israel.

2. T. caemlea Ser, Ontario, Canada,

3. r. calllcerasoites Fish Ontario, Canada.

379
P.CW-2

380 -Afohini Gupta

Species Place of collection

4. T. corniculata Linn. Meerut, India.

5. T. cretica (L.) Boiss. Ontario, Canada.

6. T. gmciliSy Benth. Nainital, India.

7. T. polycerata, Linn. Meerut, India.

8. T. stellata Forsk. Jerusalem, Israel.

9. r. suavissima Lindl. Canberra, Australia.

Petals from fully mature flowers were cleaned in lactic acid, stained in, 1%
aqueous safranin and mounted in glycerine. Twenty-five petals of each, species
were studied. Drawing of each petal was subdivided equally into basal, central
and peripheral regions. Mean number of dichotomies and anastomoses was calcu-
lated in each sector. Area of the petal was measured by planimeter. For descri-
bing the types of anastomoses classification of Arnottand Tucker (1963) is mainly
followed.

3* Observations

Each flower consists of a standard, two keel arid two wing petals which vary in
shape, size and structure (figures 1-3). Standard petal is symmetrical while the
other petals are asymmetrical. Keel and, wing petals are clawed (figures 2-3,
5-6), but standard petal is generally nonclawed except T. arabica, T. gradlis
(figure 7) and T. callicerasoites. Each petal receives a single trace which branches
after the separation of the individual petal (figures 1-2).

Standard petal is generally obovate in shape (figure 4) except T. gmcilis,
T. cretica (figure 8) and T. callicerasoites in which it is elliptic. Apex is notched
in all the species. The bundle that enter the standard petal branches into one
median and two costal veins in the basal region (figures 4, 8). However, in
T. gracilis (figure 7) it branches at the base of the limb. Out of three veins, costal
vein branches further dichotomously but the median vein first branches trichoto-
mously and then dichotomously (figures 7-8). However, in few petals of
r. polycerata median vein shows first dichotomous branching then trichotomous
and further dichotomous branching.

The bundle of the keel and wing petals branches dichotomously at the junction
of limb and claw (figures 2-3, 5-9). In T. cderulea and T. arabica wing petals
possess ridges and grooves at the distal end and the veins end freely in the groove
region. The wing petals of T. polycerata (figure 5) and T. callicerasoites are serrated
at the distal end.

In all the species wing petals at the junction of the limb and claw possess a
spur like process which is generally tubular except T. stellata and T. cretica (figure 9),
in which it is flattened. This spur like process receives supply from the main
bundle of the petal (figure 3). Wing petal of T. polycerata has a tooth like process
also at the distal end. This tooth like process is nectariferous and vascular
bundles in this region are found to be inversely oriented with respect to the petal.
Keel petals of this speciei has a pocket Jike structure which receives the tooth
like process of the wing petal (figures 5-6). In all the types pf petals venation
is mainly open dichotomous (figures 1-9).

Petal venation' in fngomlia

381

Figures 1-9. Venation in petals of Trigonella. 1-3. standard, keel and wing petals
of T. caerulea, 4-6. standard, wing and keel petal of T, polycerata, 7. standard petal
of T. gracilis, 8. standard of T. suavissima, 9. wing 'petal of T, cfetica.

Mohini Gupta

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Petal venal ion in Trigonella

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384 Mohini Gupta

3.1. Dichotomies

Table 1 shows mean area, mean length and mean breadth of the different types
of petals and mean number of dichotomies in different regions with standard error.
A positive correlation has been found in all the species except T; caemka,
T. comiculata and T. callicerasoites between area and number of dichotomies
(figure 10). Similarly a positive correlation of dichotomies is also found with
length except T. comiculata^ T. gracilis and T. suavissima (figure 11). No corre-
lation is found with breadth (figure 12),

cr

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Figure 10. Histogram comparing the average area with mean number of dichoto-
mies of standard (s), wing (w), keel (k) and total corolla (T) in different species.

140

1202

,00

80
60
40
20

o

Figure 11. Histogram comparing the average length with the mean number of
dichotomies of standard (s), wing (w), keel (k) and total corolla (T) in different

species.

JOO o

Figure 12. Histogram comparing" the' average breadth with the mean number of
dichotomies of standard (s), wing (Mr), keel -Oc) and total corolla (T) *» '-different
species.

Petal venation in Tngonclla

385

Figure 13. Histogram comparing the average area with the mean number of anasto-
moses of standard (s), wing (w), keel (k) and total corolla (T) in different species.

Figure 14. Histogram comparing the average length with the mean number of
anastomoses of standard (s), wing (w), keel (k) and total corolla (T) in different
species.

Abbreviations : A = T. arabica , B = T. caerulea ; c . = T. callicerasoites ; D = T.
corniculata ; E = T. cretica ; * = T. gracilis ; o = T. polycerata ; H = T. stellata ;
i --= T. suavissima.

3 . 2. Anastomoses

The open dichotomous venation becomes complicated at places by vein fusions.
The percentage of petals showing vein fusions varies from 20-66-6% (table 2).
A slight positive correlation has been observed between the length or area of
petals and the number of anastomoses (figures 13-14). In most of the petals
anastomosing is generally found at one point. Among the different types of
anastomoses C and D types are of most frequent occurrence in all the species
(table 3), In all the species vein anastomoses are generally concentrated in the

386 Mohini Qupta

Table 2. Percentage of petals shown g

SI. Total

No. Name of the species (%)

Points of anastomosis

5-10

i.

T. arabica

40-0

66-6

33-34

2-

T. callicerassoites

53-4

37-5

25-00

25-0 12-5

3.

T. caerules

40-0

66-6

33*34

4.

Tm corniculata

20-0

66-0

33-0

. . . . .

5.

T. cretica

20-0

66-0

33-0

. .

6.

71 graeilis

71-4

80-0

10-0

to-o

7.

T. polycerata

66-6

20-0

30-0

10-0 40-0

S.

T. stellata

33-3

80-0

20-0

9.

T. suavlssima

6<5*6

60-0

20-0

10-0 10-0

Table 3. Percentage of different types of anastomoses.

SI.

No . ^atne of the species

D

h.

21 arabica

25-00

25-00

25-00

• • . *

25-00

2.

r. callicerasoites

66-60

..

33-34

3,

T. caerulea

11-11

77-78

11-11

4;

T. corniculatd

..

25-00

50-00

..

25-00

5.

T. cretica

..

25-00

75-00

6.

T. gracilis

11-11

44-44

ll'H 11-11

22-22

7.

T. polycerata

6-96

6-96

41-86

4*64 2-32

37' 21

8.

T. Stellata

57-14

..

42-86

9.

T.suavissima

11-76

35 '89

35-89

5-08

11-76

central region except the standard petal of T. polycerata in which they are more
in the peripheral region.

4. Discussion

The petal venation of this genus has shown some interesting features. Arnott
and Tucker (1964) reported significant correlation between size (length, width,
area) and number of dichotomies. In this paper a positive correlation is found
with the length and area only. They have also given importance to the position
of the dichotomies. In the present study maximum number of dichotomies is
found in the central region except the standard petal of T. cretica and T. corniculata
in which it is found in the basal region. Generally no correlation is found
between the frequency of anastomoses and length of the petals. HoweveF, the

Petal venation in TrigmiolUt 387

petals of T. arabica, T. polycerata and T. gmcilis have shown a positive correlation.
Just like the length no correlation is found with the area excepting the petals of
71 caemlea, T. comiculata and T. polycerata. Thus from above it is clear that
frequency of anastomoses is not dependent upon the area or the length of the
petals. In all the species highest frequency of anastomoses is found in the central
region but the wing petals of 7". ambica and keel petals of T. comiculata and
T. stellata possess highest frequency in the basal region. Datta and Saha (1968)
reported highest frequency in the peripheral region in the members of phaseolae.

Among the different types of anastomoses C and D types are of most frequent
occurrence in all the species studied except T. crelica in which C and C are
common. Earlier Dalta and Saha (1968) have also reported the frequent occur-
rence of these types for the members of phaseolae and Arnott and Tucker (1963)
for Ranunculus repens. In addition to these types only B type is found in
T. caendea and T. comiculata, A and B in T. arabica, A, B, C in T. suavissfma,
A, C, C" in T. gmcilis and all types in T. polycerata. Thus in this genus distri-
bution of anastomoses appears species specific. Among the different species
more points of fuisons are found in T. polycerata, T. callicerasolt.es, T. gracilis
and T. suavissima.

Foster and Arnott (l960),B£nerji and Mukherji (1970), and Subramanyam
and Nair (1973) are of opinion that open dichotomous venation is primitive while
on the other hand Chertek (1962, 1963) views that anastomosed venation is primitive.
The present study supports the former view and among the species of this genus
T. callicerasoites, T. gracilis, T. polycerata and T. suavissima appears to be more
advanced as these species have more points of fusions and more number of
petals showing vein fusions.

Earlier reports indicate that species of Trigonetta are characterized by presence
of both simple as well as medicagoid type of corolla. In the present investigation
simple corolla is found in all the species except T. polycerata in which it is of
inedicagoid type. The wing petal of T. polycerata possesses a tooth like structure
arising from the inner surface of the petal and receiving supply from the main
bundle of the petal. In this region vascular bundles are inversely oriented with
respect to the petal bundle. The same inversely oriented supply was reported
by Arber (1936) for the nectary of Ranunculus and for the corona by Narcissus
(Arber 1937). Thus this tooth like structure may be regarded as petalline nectary
or corona. Earlier Larkin and Graumann (1954) named this structure as horn
in Medicago sativa, Thus it can be concluded that among the different species
of this genus T. polycerata is the highest evolved species.

Acknowledgements

The author expresses her gratitude to Prof. Y S Murty for giving valuable
suggestions and facilities, to Dr R Loiselle, Prof. A H Gibson and Prof. C Heyn
for providing materials and to CSTR for giving Post-Doctoral Research
Fellowship.

388 Mohini Gupta

References

Arber A 1936 Studies in flower structure U, OB the vascular supply to the nectary in Ranun-
culus ; Ann. Bot. 50 305-319
Arber A 1937 Studies in flower structore HE. On the carcma and androecium in certain

Amaryllidaceae ; Ann. Bot. (NS) 1 293-304
Arnott H J and Tucker S C 1963 Analysis of petal venation in Ranunculus I. Anastomoses in

R. repens, V. plenlflorus ; Am. /. Bot. 50 821-830
Arnott H J and Tucker S C 1964 Analysis of petal venation in Ranunculus II. Number and

position of dichotomies in R. repens var. Pleniflorus ; Bot. Gaz. 125 13-25
Banerji M L 1972 Morphological studies on petal venation of Ranunculus diffusus DC. and its

affinities with fossil materials ; Biology Land Plants pp. 1-8
Banerji M L and Mukherji M 1970 Petal venation in Ranunculus scleratus Linn.; Castama 35

157-161
*Chertek J 1962 Die Verlauf der Nervatur in den Kronblattern bzul kronen der Dikotyledonen ;

Novitates Bot. Hort. Bot. Univ. Carolineae Pragensis pp. 3-10
*Chertek J 1963 Die Nevatur der Kronblattern bei den Vertretern der Ordnung Rosales s.L;

Acta Horn. Bot. Pragensis pp. 13-29
Datta P C and Sana N 1968 Specificity o.f distribution of venation anastomosis patterns in

petals of phaseoleae (Leguminosae) ; Ann. Bot. 32 791-801
Foster A S and Arnott H J 1960 Morphology and dichotomous va&culature of the leaf of

.Kingdonia umflora ; Am. J. Bot. 47 684-698
Larkin R A and Graumann, H O 1954 Anatomical structure of alfalfa flower an explanation of

tripping mechanism ; Bot. Gaz. 116 40-52
Subramanyam K and Nair N C 1973 Dichotomous venation and anastomosis in corolla of

an orchid ; Proc. Indian Acad. Sci. (Plant Sci.) 78 195-202

* Not consulted in original.

Proc. Indian Acad. Sci. (Plant Sci.)» Vol. 91, Number 5y October 1982', pp. 389-395.
© Printed in India.

Responses of cotton-cultivars to long day conditions

J G BHATT and M R K RAO

Central Institute for Cotton Research, Regional Station, Coirabatore 641 003, India

MS received 29 August 1981 ; revised 1 July 1982

Abstract. Flowering of cultivated varieties of cotton belonging to G. arboreum,
G. herbaceum and (?. hirsututn was delayed by over 14 hrs of daylength because of
increase in number of days for square formation. The long day treatment in
general increased height, production of fruiting branches, leaf area and dry weight
per plant. The number of fruiting forms, bolls retained, yield of seed cotton and
fruiting coefficient decreased under long day conditions. These characters were
affected more in upland varieties and short day Cambodia derivatives. The most
of G. arboreum and G. herbacewn varieties became more vegetative in growth but
their boll number and yield per plant increased.

Since the varieties 1998 F (G. hirsutum) and Gaorani 1111 (G. arboreum) were
tolerant to long photopejriod and grew satisfactorily, it is suggested that these
may be used as donor parents for improving the quality of cottons grown in northern
India.

Keywords. Photoperiod ; flowering ; long day ; boll ; square ; yield.

1. Introduction

The flowering of cultivated and wild species of cotton is governed by both, day-
length and temperature as shown by Waddle et al (1961) and Mauney and Phillips
(1963). They found that most of the varieties flower under short days and cool
nights. The importance of low night temperature in promoting flowering was
further stressed by Mauney (1966) using a non-photoperiodic upland cotton
variety. Bhatt (1977) and Bhatt et al. (1976) found that long days and high tem-
perature singly or in combination delayed flowering of upland cottons of northern
India whereas the photosensitive Cambodia varieties did not flower .udder long
days alone or long;day plus high temperature. The upland genotypes when gro.wn
at latitudes 29° N,21° and 11° N flowered progressively earlier and at lower nodes
at more southern latitudes because of reduction in daylength and temperature.
Though the cotton crops in the northern cotton zone in India are grown under
irrigation and thus give higher yields, they are of short and medium staple. An
attempt was therefore made to screen some of the promising varieties (belonging
to both G. hirsutum and G. arboreum) for tolerance to long days so. as to identify
donor parents for improving quality. , . :

389

390 / G Bhaii and M R K Rao

2. Materials and methods

The cultivated varieties in India representing
hybrid developed through genetic male sterile

Species

G. hirsutum
G. hirsutum
G. hirsutum
G. hirsutum

G. hirsutum

G. arboreum
G. arboreum
G. arboreum
G. arboreum
G. arboreum
G. herbaceum
G. herbaceum

Variety

LSS
320 F
CP 1998 F
170 Co 2

MCU 1

AK 235
G 27
K 9

Gaorani 1111
Gaorani 1187
V 797

Jayadhar
CPH 2

three species of cotton and an i\
line were taken as shown below :

Area

Upland type from North zone
Upland type from North zone
Grown in South zone

An Itfdo-Americart type grown
in Central zone

Photosensitive short day deriva-
tive of Cambodia cotton from
South (used as check)

From Central zone
From North zone
From South zone
From South zone
From South zone
Grown in Central zone

Grown in South zone

An intraspecific (G. hirsutum)

hybrid

The cotton varieties were raised in large (45 cm diameter and 105 cm depth)
pots adequately manured. There were six plants per treatment one in each pot.
The daylength around latitude 30°N in northern cotton zone in India when cotton
is sown and even thereafter until about 80 days is over 14 hours. The normal
dayteiigth in summer at Coimbatore (latitude 11 °Nf> varies from ll-50hrs to
12-20hrs. It was extended to 14-5€tes through 60 watt incandescent lamps.
The treatment were (i) Conti^l, i.e., normal dayleagth of 11-50 h*s to 12'201u?s,
and (H) long photopetiod of 14* 50 his. Under both the treatments the day and
night temperatures were the same. The ctey temperature was between 34-0° C
to 35- 5° C from germination to square formation for about 35 days and thereafter
until flowering did BOt rise above 3'6-8° C. The night temperature during this
period fluctuated between 17- 5° C to 21-8° C to 24- 6° C and subsequently remained
around 23>0°C. The long day treatment was discontinued after 75 days after
soi«g. Taking cotyledonary node a$ zero, the node on the main stem producing
first sympodiuin was taken as the first fruiting node. The fruiting coefficient is
defined as the yield of seed cotton produced pee lO&'gm of total dry matter
(Crowtter 1944).

Effect of long days on cotton

391

3. Results

The long days delayed formation of flower buds in all the varieties irrespective
of the species (table 1). The short day Cambodia derivatives MCU 1 and 170
Co 2 set squares just after the long day treatment was discontinued. The early
types CPH 2 and 1998 F took only 4 and 6 days more respectively to square
when compared with the delay of three weeks in LSS and 320 F. Among the
diploids both the Gaorani types were early whereas in the rest, delay in square
formation ranged from 15 to 20 days.

Most of the varieties took significantly more days to flower when photoperiod
was extended except CPH 2 and 1998 F with a delay of 4 and 9 days respectively
and of 6 days in Gaorani 1111. With a difference of only 2 days Gaorani 1187
appeared to be practically photoinsensitive. The photosensitive types MCU 1
and 170 Co 2 took maximum days to flower followed by the upland types LSS
and 320 F, and the Asiatic type AK 235. The square period, i.e., the number of
days from initiation of flower bud to opening of the flower remained more or less
the same between the treatments except that it increased by 3 days in 1998 F and
reduced by 3 days in G 27, 5 days in K 9, 6 days in Gaorani 1187 and 3 days
in Jayadhar.

Table 1. Flowering behaviour of cottoii-cultivars as affected by extended photo-
period.

.Species

Variety

Number of days
to square

Number of days
to flower

First fruiting
node

Days for first
boll bursting

G. hirsutum

LSS

34

56

52

75

4-

3

9-

0

us

150

G. hirsutum

320 F

33

54

56

75

4-

0

6-

0

120

145

G. hirsutum

CP1998F

25

31

46

55

4-

0

6-

0

S5

95

G. hirsutum

170 Co 2

35

72

54

39

3-

0

7-

6

120

147

G. hirsutum

CPH 2

22

26

42

46

6-

0

6-

0

86

84

G. hirsutum

MCU1

39

73

59

95

2-

3

10-

3

110

130

G. arboreum

AK235

33

52

53

72

3-

0

7-

0

122

140

G. arboreum

027

35

50

55

67

5-

3

7-

7

118

130

G. arboreum

*C9

34

54

59

74

2"

6

7-

0

125

142

G. arboreum

Gaorani 1H1

35

42

53

59

2-

0

6-

6

130

139

G. arkoreum

Gaorafli 1187

35

43

53

55

4-

6

7-

>7 .

125

132

G. herbaceum

V797

45

60

65

82

5-

•o

6

-o

130

150

G. herbaceum

Jayadhar

35

50

65

77

2-

6

5

•o

120

135

S.E. foe varieties

1-90

&-61

!••

40

4' 84

S.E. for treatment

1-10

3-51

o-

58

1-97

• Control T = Treatment

392 / G Sfiatt and M A K Rod

The node at which the iirst synipodial branch appeared was also higher, the
notable exceptions being CPH 2 and V 797. Other varieties flowered 2-8 nodes
higher . under .long day conditions.

An increase in daylength also increased boll maturation period, i.e., the days
from opening of the first flower to dehiscence of carpellary wall or boll bursting
thus adding to farther lateness. The trend was more or less similar to flowering.
The upland type LSS took the maximum number of days whereas CPH 2 was
unaffected. It was interesting to note that the Gaorani * cultures 1187, 1111
and 1998 F were late by 7, 9 and 10 days respectively.

Except for CPH 2 and Gaorani 1111 plant height increased significantly in all
the varieties reaching the maximum in 170 Co 2 and LSS (table 2). The number
of sympodia per plant also increased similarly except in CPH 2, V 797 and
CP 1998 F. The long day conditions reduced production of fruiting forms with
notable exceptions of G 27 and K 91 showing an increase whereas CP 1998 F
was not affected significantly. Maximum reduction was found in MCU 1, AK 235
and V 797.

There was a significant increase in leaf area per plant in all the varieties. It
was over three times in LSS and over twice the normal area in 320 F, MCU 1 and
G 27 followed by 170 Co 2. Increase in leaf area was comparatively less in
CPH 2, V 797 and CP 1998 F.

Table 2. Effect of extended photopcriod on Some growth characters.

Species Variety Height (cm) No. of sympodia No. of fruiting Leaf area per

per plant forms per plant plant (cm8)

G. hirsutum

LSS

60-

2

117-1

12

20

41-

6

30-7

2983

11581

G. hirsutum

320 F

65-

3

95-2

9

15

33-

3

39*0

2950

6170

G. hirsutum

GP 199$ F

40-

3

60-8

13

14

42-

0

39-0

2507

3575

G. hirsutum

170 Go 2

55-

3

120-7

12

22

34-

2

22-6

4210

8179

G. hirsutum

GPH2

54-

1

54-3

11

12

38-

0

30-0

2668

3548

G. hirsutum

MGU 1

51-

3

90-1

12

17

35-

6

17-0

6170

13217

G.arboreum

AK235

97-

3

144-2

22

32

59*

6

38-7

2850

4879

G. arbareum

G 27

108-

0

130-8

19

25

65

6

70' 0

2217

4717

G.arboreum

K9

72-

3

143-6

' 17

25

59-

3

73-3

2980

5316

G. arboreum

Gfaoraiti 1111

72-

6

77-0

16

18

51-

3

35-0

2950

4850

G.arboreum

Gaorani 11 87

52-

0

85-1

11

15

49.

0

31-0

2170

3950

G. herbaceum

V797

56-

0

83-3

12

15

62'

3

40-1

2007

2970

<j. herbaceum

Jayadhar

59

•1

101-2

14

20

54-

3

41-6

2350

' 4317

S.E.'for varieties

7-39

4-28

8-61

159-80

S.E. for treatments

3*02

1-75

'3-51

65-24

C «= Control T = treatment

Effect of long days on cotlari

At maturity dry matter per plant increased in ino&t of the varieties under long
day treatment (table 3). But CP 1998 F, AK 235 and Gaorani 11 87 were not
affected. It was, however, highly significant in CPH 2, LSS 320 F, MCU 1, K 9,
Gaorani 1111 ajad Jayadhar.

In G. hirsutum varieties LSS 170 Co 2 and MCU 1, the number of bolls per
plant at maturity decreased with consequent decrease in yield of seed cotton. Both
320 F and CP 1998 F were unaffected whereas CPH 2 recorded higher yield
under long days. Except Gaorani 1187, all G. arboreum varieties and V 797 had
more number of bolls and yielded more. Jayadhar remained unaffected. Fruiting
coefficient of most of the varieties decreased because of higher dry weights of their
vegetative parts when grown under long days.

4. Discussion

The enhanced photoperiod delayed flowering because of increase in the number
of days required to initiate square formation. The square period was more or
less unaffected in G. hirsutum varieties confirming the previous findings of Bhatt
(1977). G. arboreum varieties were similarly affected except Gaorani cultures
where flowering was delayed by 2 and 6 days only. The G. arboreum variety
used by Mauney and Phillips (1963) showed essentially no reaction to the environ-
ments in their study whereas flowering was delayed from 12 to 19 days under
long day conditions in the present study with the first fruiting node pushed up
significantly. Differences in flowering responses of different varieties reported
by Mauney and Phillips (1963) and in our experiments may be attributed to differ-
ences in day and night temperatures though long day conditions were similar.
Day temperatures in their experiments varied from 27° C to 32° C whereas the
night temperature was fixed at 15° C or 30° C, while in our experiments day tempe-
ratures varied from 34° C to 36° C and night temperatures from 17° C to
24° C.

An increase in leaf and stem dry weight and a decrease in yields of seed cotton
of most of the varieties under long days reduced their fruiting coefficients (Bhatt
1970). In G. hirsutwn group CP 1998 F responded exceptionally well in terms
of its vegetative and reproductive growth maintaining high fruiting coefficient.
This variety was late in flowering by 9 days under extended photoperiod and took
95 days for the first formed boll to burst as against 150 and 145 days taken by the
northern upland varieties LSS and 320 F respectively. Because of its superior
fibre characters, better yielding capacity and high degree of tolerance to long day
conditions, it may prove as a suitable donor parent for improving the quality of
northern hirsutums in India. Next in performance was the intm-hirsutum hybrid
CPH 2. But the short day Cambodia derivatives MCU 1 and 170 Co 2 failed
miserably in relation to flowering and growth (Hutchinson 1959; Bhatt 1977).

The G. arboreum varieties except Gaorani 1111 took 12 to 19 days more to flower
and then4 bolliAg period also increased by 12 to 18 days. The new culture Gaorani
1111 in this respect took only 6 to 9 days more for flowering and boll bursting
respectively. It compares well with tie northern arboreum variety G 27 in terms
of yield of seed cotton and similar fruiting coefficient. By virtue of its long staple,
it may prove useful for improving the quality of northern arboreums in India*

394

J G Bltatt and M R K &ao

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Effect of long days on cotton 395

Among the G. herbaceum varieties, V 797 though late in flowering and boll
bursting appeared tolerant to longer photoperiod as its growth and yield improved
But the fruiting efficiency of Jayadhar was reduced markedly.

Acknowledgement

We thank Dr M R H Qureshi, IARI Regional Station, Hyderabad, for kindly
supplying the Gaorani cultures developed by him.

References

Bhatt J G 1970 Yield capacity of cotton plant in relation to the production of dry matter ;
Indian J. Plant Physiol. 13 219-224

Bhatt J G 1977 Growth and flowering of cotton (Gossypium hirsutum L.) as affected by day-
length and temperature ; /. Agric. ScL (Camb.} 89 583-587

Bhatt J G, Shah R C, Patel B D and Seshadrinathan A R 1966 Responses of cotton genotypes
to latitudi nal differences ; Turrialba 26 247-252

Crowther F 1944 Studies, on growth analysis of the cotton plant under irrigation in the Sudan.
III. A comparison of plant development in Sudan Gezira and Egypt ; Ann. Bot. (NS)
8 213-257

Hutchinson J B 1959 The application of genetics to cotton improvement (Cambridge University
Press)

Mauncy J R 1966 Floral initiation of upland cotton Gossypium hirsutum L. in response to tempe-
ratures ; /. Exp. Bot. 17 452-459

Mauney J R and Phillips L L 1963 Influence of daylength and night temperature on flowering
of Gossypium ; Bot. Gaz. 124 278^283

Waddle B M, Lewis C R and Richmond T R 1961 The genetics of flowering response in
cotton. III. Fruiting behaviour of Gossypium hirsutum race lati folium in a cross with
a variety of cultivated American upland cotton ; Genetics 46 427-438

P. (B)-3

Proc. Indian Acad. Sci. (Plant Sci.), Vol. 91, Number 5, October 1982, pp. 397-407.
© Printed in India.

Seed germination and seedling establishment of two closely related
Schima species

RAM B.OOJH and P S RAMAKRISHNAN

Department of Botany, School of Life Sciences, North-Eastern Hill University,
Shillong 793 014, India

MS received 15 December 1981 ; revised 13 July 1982

Abstract Seed germination of S. khasiana from Upper Shillong and S. wallichii
from Shillong, Umsaw and Burnihat in Meghalaya, north-eastern India, and
seedling establishment and growth of these species/populations in reciprocal
cultivation were studied. Seeds lost viability and germinability gradually within
a year when stored at 5 cm below the soil surface under natural conditions or at
20° C in the laboratory. Storage at 0°C permitted retention of 15-25% viability.
Seeds germinated better on the surface layers (0-2 cm) of the soil. S. khasiana
had a lower temperature optimum (15° C) of germination while the populations
of S. wallichii had a higher temperature optimum (20/25° C). At a temperature
of 30° C, the lower altitude population of S. wallichii from Burnihat gave higher
germination than the high altitude population from Shillong. A given species/
population of Schima gave better seedling establishment and growth in its own
natural habitat as compared to the introduced populations from the other alti-
tudinal sites. This is indicative of the close adaptation of the natural populations
to their habitat and ecotypic differentiation in this species.

Keywords. Tree adaptation ; seed germination ; tree seedling establishment ; alti-
tudinal ecotype ; Schima.

1. Introduction

Germination and establishment represent two critical phases in the life-cycle of
a plant species and these two aspects have been related to adaptation and distri-
bution pattern of species in space (Koller et al 1962 ; Harper 1965 ; Cohen 1967 ;
Ramakrishnan 1972 ; Ross and Harper 1972 ; Thompson 1973 ; Boojh and
Ramakrishnan 1981a) and in time (Kapoor and Ramakrishnan 1973). However,
this aspect of the problem in relation to adaptive strategy of tree species has
received little attention (Kozlowski 1971 ; 1979). Although the size of a single
species population is to. some extent related to seed supply, it is ultimately deter-
mined by favourable conditions available for germination and establishment
(Harper et al 1970). Further, a large gap often exists between the seeding poten-
tial of a species and the actual number of seedlings established ' in that area,
depending upon environmental conditions.

397

398 Ram Boojh and P S Ramakrishnan

S. khasiana Dyer and S. wallichii (D C) Korth family Ternstroemiaceae are
two closely related and economically important timber tree species of north-
eastern hills of India. These species show a distribution pattern on an altitudinal
basis where S. khasiana is restricted to higher elevations (1800-1900 m), while
S. wallichii shows a wide distribution ranging from 100 to 1600m. These two
species are early successions and come up in the secondary fallows after slash
and burn agriculture (Jhum). These are light-demanding and regenerate pro-
fusely in the open, through light wind-dispersed seeds. The present study is a
comparative investigation of seed germination and seedling establishment of these
two species along an altitudinal gradient in the Khasi Hills of Meghalaya.

2. Climate

All the three sites are characterised by marked seasonal changes in climate. The
year could be divided into 3 distinct seasons : (i) Monsoon season of high tempe-
rature and humidity extending from May to October when over 80% of the rain-
fall occurs, (ii) Winter season (November to February) of lower temperature which
is comparatively dry except for a few winter showers, and (iii) A warm, dry and
windy summer in March-April (table 1).

3. Methods of study

Mature fruits of S. khasiana were collected from Upper Shillong and that of
5. wallichii from 3 sites at Shillong, Umsaw and Burnihat, in the months of
February-March, 1978. Seeds were separated out by air drying. The fruit and
seed weight measurements were based on 100 fruit/seed with 20 replications.

Table 1. Comparison of climatic data at study sites during 1978-79.

Upper Shillong Shillong Umsaw Burmhat

Location

Latitude (N)
Longitude (E)
Altitude (m)

25-34
91 '56
1900

25-34
91-56
1400

25-45
91-54
800

26-02
91-52
300

Temperature (° C)

Mean monsoon maximum 22 24 30 32

Mean monsoon minimum 15 16 22 24

Mean winter maximum 16 16 20 25

Mean winter minimum 3 6 10 12

Precipitation (mm) 2400 2000 1800 1600

Germination and establishment of Schima species 399

Seeds were stored in the laboratory at 20 ± 5° C and 0° C in BOD incubators
in tightly closed plastic bottles. In nature, seeds were similarly stored 5 cm below
soil surface. The moisture content of seeds at the time of storage was 10%.
Viability and germinability of stored seeds were tested at intervals of 3 months
with four replicates of 50 seeds of each species/population. For testing the viabi-
lity of seeds a freshly prepared 5% aqueous solution of 2, 3, 5-triphenyltetra-
zolium chloride (TZ) was used. Seeds were first soaked in water for 10-12 hrs,
then seed coats were punctured to facilitate entry of the xz solution and were left
in the dark at 30° C for upto 24 hrs. Seeds with completely stained (red colour)
embryos were scored as viable. Germinability was tested by placing seeds in petri-
dishes over moist filter-paper at a constant temperature of 15° C for S. khasiana
and 20° C for S. wdllichiL

Seeds were tested under two conditions, of continuous light under an incandes-
cent fluorescent tube (500-600 1 X ) or under continuous darkness by covering
the petri-dishes inside thick black paper, at a constant temperature of 20° C. Seed
germination in dark was counted under green light. Germination at different
constant (15, 20, 25, 30 and 35° C) and alternating (25/15 and 25/20° C) tempe-
rature regimes were tried in BOD incubators maintained at these temperatures.
The effect of different soil depths of 0, 2, 4, 6, 8 and 10cm on germination was
tested in pots filled with soil, by placing seeds at the appropriate depth.

All germination experiments were replicated 4 times with 50 seeds in each test.
The emergence of radicle was taken as an indicator of germination. Tests in all
cases were done for 20 days after the seeds were placed for germination.

Ten replicates of 100 viable seeds (viability was ascertained for a given seed-
lot on the basis of preliminary germination tests) of each species/population were
sown at a depth of 5 cm at all the 4 study sites both in the open and under
forested situations, in May 1978. The depth of 5 cm for sowing was chosen in order
to avoid washout of seeds under heavy rainfall. Observations on the seedling
emergence and establishment were taken at monthly intervals. Seedlings were
harvested at the end of one year period and after noting plant height and leaf
area using a planimeter, the root and shoot portions were separated and dried to
a constant weight at 85 ± 2° C.

4. Results

4.1. .Fruit and seed characters

The capsules and seeds of S. khasiana were heavier than that of the populations
of S. wallichii. While the fruit weight of the populations of S. wattichii were not
very different, significantly higher seed weight was noticed for the Burnihat popu-
lation of this species compared to that of the two other populations (table 2)t

4.2. Germination studies

4.2a. Effect of storage : When seeds were stored in the soil or in the laboratory
at a temperature of 20 ± 5° C, both viability and germinability of the seeds of all

400

Ram Boojh and P S Ramakrishnan

the populations decreased markedly with passage of time so that after one year,
seeds were totally non-viable or gave very poor germination. However, storage
at 0° C maintained better viability and germinability after 1 year storage (figure 1).

4.2b. Depth of burial : As seen from figure 2, the depth of burial affected both
the time and the final percentage of germination. Maximum germination was
found to occur at 2 cm depth and it decreased at the depths greater than this for
all the species and populations. At soil surface though faster germination
occurred the total percentage was lesser than at 2 cm depth.

Table 2. Fruit and seed weight of Schima species/populations.

Fruit weight
(g)

S. khasiana

1-6T±0-09

(± S,E. of the mean)

Seed weight
(mg)

M8±0-03

S. wattichil

Shillong

M3±0-07

0-46±0-06

Umsaw

1-04±0-03

0-44±0«02

Burnihat

1-07±0-05

0-53±0'01

80

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STORAGE PERIOD(MONTHS)

Figure 1. Viability (open columns) and germinability (hatched columns) of Schinm
seeds after different storage periods. First column, storage at 0° C ; second column
storage at 20 ± 5° C ; and third column, storage under soil.

Germination and establishment of Schima species

401

4.2c. The effect of light and darkness : There was germination both in the dark
and light and the results obtained were not significantly different under these two
conditions (table 3).

4.2d. The effect of temperature : Table 4 reveals the effect of various tempera-
ture regimes on the germination of the seeds of Schima species and populations

• S. knastana
o 5- walli

Shillong

• Urn saw and
Burnihat
Populations

6 8

DEPTH OF BURIAL (cm)

Figure 2. The relationship between seed depth, germination and emergence period
of seedlings of Schima species/populations over a period of 35 days after sowing.
Continuous lines represent % emergence of seedlings and broken lines represent
number of days taken for emergence.

Table 3. The effect of light and dark treatment on seed germination of Schima
species.

Continuous light Continuous dark
20° C 20° C

S. khasiana

50

48

± 5-4

± 3"4

S. wallichii

62

57

± 6-4

dz 1-3

(db S.E. of the mean)

The seeds of populations of 5. wallichii were pooled.

402 Ram Boojh and P S Ramakrishnan

Table 4. Germination (%) of S. khasiana and *S. wallichii seeds at various
constant and alternating temperatures.

Constant temperatures Alternating temperatures

(° C) (° C)

15 20 25 30 35 25/15 25/20

S. khasiana 55 46 44 42 0 46 45

±2-7 ±3-6 ±2*9 ±2'2 ±2-3 ±5-5

S. wallichti

Shillong 37 48 4S 33 20 48 50

±4-3 ± 6-0 ±6-8 ±4-1 ±7-5 ±4-1 ±4-6

Umsaw 42 65 58 40 ^1 61 70

±5-3 ±3-7 ±3-7 ±6-8 i 2*9 ±3-7 -H9-9

Burnihat 38 69 56 50 33 59 64

±2-2 ±3-7 ±5-4 ±4-8 ± 7'6 ±3-4 ± 3'6

(± S.E. of the mean)

S. khasiana showed maximum germination at constant 15°C, with a gradual
decrease with increase in temperature, so that at 35° C no seeds of this species
germinated. Populations of S. wallfchii showed maximum germination at 20
and 25° C with decrease in germination on either side. At 30° C, the lower alti-
tude population of S. wallichii from Burnihat gave higher germination than its
higher altitude population from Shillong. Two alternating temperature regimes
tried were favourable for germination for all the species/populations.

The rate of germination was faster at 15°C for S. khasiana and 20 and 25° C
or alternating (25/20° C) for S. wallichii populations (figure 3).

4.3 Seedling establishment

4. 3 a. Seedling emergence : Only a small proportion of seedlings could emerge
under field conditions at all the study sites. Further the differences in emergence
were not significant (at 5% level) between species/populations (table 5).

4.3b. Survivorship : No seedlings could survive under forested situations beyond
a period of 2 months. Under open grown situations, mortality was generally very
high resulting in a steep decline in population upto January-February, at all the
sites. At Upper Shillong, however, the rate of decline in population was slower
for S. khasiana with an ultimately large population size compared to the popula-
tions of S. wallichii. At the other 3 experimental sites, however, the pattern of
survivorship was not very different for the populations of Schima, though the local
populations showed better survivorship than the introduced ones. S. khasiana
gave the lowest final survival at these three sites (figure 4).

Germination and establishment of Schima species

403

70

60
50
40
3 0

: 20

£ 1 0

S.wa! iichii
(Shillong)

10

15 20 .. 5 10

DAYS OF GERMINATION

IS

Figure 3. Percentage germination of Schima species/population at different periods,
at constant and alternating temperature of 15°C(«&); 20°C(O); 25°C(H);
30°C (Q); 35°C (A,); 25/15° C (A) and 25/20° C (x).

Table 5. Seedling emergence (%) of S. khasiana and S. waltichii at different alti-
tudinal sites.

Field stations

S. khasiana

Species/Populations
S. wallichii

Shillong

Umsaw

Burnihat

Jpper Shillong

20
±4-6

10

d= 2-3

11
± 1-8

14
± 1-8

^hillong

19

± 2-4

21

± 3-9

10

± 3-3

10

± 2-1

Jmsaw

16
± 1-7

13

± 2-7

13
± 1'2

13

±2-5

lurnihat

16

± 3'9

12

± 1-7

11

±2-3

11
±0-6

'.(B)

404 Ram Boojh and P 5 Htnic?!s'inan

UPPER SHILLOKG

SHtlLCNG

J ASONDJ F W A M J JASOND J FMAMJ

Figure 4. Survivorship of Schima seedlings under field conditions (in open). No
seedlings could survive beyond 2 months under forested situations. S. khasiana
(•); S. wallichii, Shillong (O), Urnsaw (») and Burnihat (D) populations.

4 . 3c. Plant performance : The growth characteristics of the different species/
populations at different sites, given in figure 5, show that the naturalized popu-
lation for a given site was superior to the other introduced populations. Thus

5. khasiana gave better growth yield under Upper Shillong site, while the popula-
tions of S. wallichti from 3 different sites did better in their respective natural
habitats.

5. Discussion

Schima species being early successional colonizers depend for regeneration on the
availability of open sites which favour their seed germination, establishment and
growth. The most important factor limiting the ability of such species to colonize
disturbed sites is the availability of seed, which must come either from a stand
in close proximity or from storage in the soil. The latter is not possible for
Schima as seeds do not remain viable in the soil for an extended period of time
as seen from the present study where the viability of seeds is completely lost
after storage in soil for one year. Thus, the species is fugitive in nature
(Hutchinson 1951), where good dispersal mechanism would play an important role
permitting the species to colonize new habitats (Salisbury 1942). Schima due to

urerminunon ana

s

2
S

I 0

7 ...

2

6 <

x
3 2

2 5:

5-

•ce
o

Jo

Figure 5. Growth performance of Schima species/populations at different field
conditions, a = Upper Shillong ; b = Shillong ; c = Umsaw and d = Burnihat
site ; filled bars for 5. kjiasiana, and hollow bars for Shillong ; hatched bars for
Umsaw and stippled bars for Burnihat populations of S. wallichti.

its light, mobile (winged) seeds often invades highly disturbed areas after slash
and burn agriculture in the region. Similarly in temperate forests it has been
reported that light seeded species Fraxinus and Betula play an important role
in revegetation after clearcutting (Bormann and Likens 1979). The variation
in seed weight in between species/populations may partly be related to climate

406 Ram Boojh and P S Ramakrishnan

(Baker 1972 : Wearstler and Barnes 1977) and partly to ecotypic differences
related to altitude which is supported by growth studies of the different Schima
populations done at different altitudes discussed below.

The differences in germination behaviour in response to temperature as seen
in the present case where S. khasiana germinated at a comparatively lower tempe-
rature compared to S. wallichii populations, have often been correlated with
climatic conditions and seed source (Callaham 1970 ; Thomson 1973), whereby
seeds from colder areas germinate better at lower temperature than those from
warmer regions. Grose (1957) has demonstrated that montane species of Euca-
lyptus germinated best at a lower temperature of 16° C, in contrast to somewhat
higher temperatures for species of warmer areas. Though the total number of
seeds of a species which ultimately germinates at a given temperature is a good
indicator of that species potential, however, the time taken to germinate is of much
significance since the early germinating individuals enjoy a considerable competi-
tive advantage (Ross and Harper 1972). The germination rate which was higher
at 15° C for S. khasiana and at 20 or 25° C for S. wallichii populations is consis-
tent with temperature optima for their germination. The rapidity of germination
in this species without a dormancy mechanism is advantageous in colonizing new
areas by producing a profusion of seedlings and this has been reported for a
majority of tropical trees which has been termed as biological nomads (Ng 1978).

Schima seeds come under microbiotic category (Crocker and Barton 1953) as
they normally lose viability and germinability within a year. Small and light
seeds are reported to lose their viability faster (Quick 1961) and this has been
reported in species of Salix, Populus and Ulmus (Wareing 1963) and Alnus (Boojh
and Ramakrishnan 198 Ib). The better retention of viability and germinability
under lower temperature storage may be attributed to slow biological and bio-
chemical processes at such temperatures (Kamra 1967).

There exists a large gap between seeding potential of a species and the number
of seedlings emerged at a given site. The failure of survival of seedlings under a
forest canopy may be attributed to the shade intolerance of the seedlings. The
differences in survival pattern for different species/populations under field condi-
tions are suggestive of the adaptation of a given population to the natural clima-
tic conditions in which they grow. This is suggested by the relatively better survi-
val and performance of local species/populations to that habitat compared to
the introduced ones. Thus, the lower altitude population of S. wallichii which
is adapted to longer growing season, higher temperature and frost-free winter is
adversely affected at higher altitude.

Acknowledgements

This study was supported by a research grant from the Department of Science
and Technology, Government of India. RB acknowledges the Council of
Scientific and Industrial Research (CSIR), New Delhi also, for the partial support
in form of a Junior Research Fellowship during the preparation of this paper.

Germination and establishment of Schima species 407

References

Baker H G 1972 Seed weight in relation to environmental conditions in California ; Ecology

53 997-1010
Boojh R and Raimkrishnan P S 198 la Temperature responses to seed germination in two

closely related tree species of Schima Reinw. ; Curr. Sci. 50 416-418
Boojh R and Rarnikrishnan P S 1981b Germination behaviour of seeds of Alnus nepalensis

Don. ; Natl. Acad. Sci. Lett. 4 53-56
Bormann F H and Likens G E 1919 Pattern and process in a forested ecosystem (New

York : Springer-Verlag) pp. 253
Callaham R Z 1970 Geographic variation in forest trees ; In Genetic resources in plants —

Their exploration and conservation (eds) D H Frankel, E Bennett (Edinburgh : Oxford

Blackwell Scientific Publications) pp 43-48

Cohen D 1967 Optimizing reproduction in a randomly varying environment when a corre-
lation may exist between the conditions at the time a choice has to be made and the

subsequent outcome ; /. Theor. Biol. 16 1-14
Crocker W and Barton L V 1953 Physiology of seeds (Waltham Mass : Chronica Botanica)

pp. 267
Grose R J 1957 Notes On dormancy and effect of stratification on germination of some

eucalypt seeds ; Victoria Bull. For. Cotnmn. 3 1
Harper J L 1965 Establishment, agression and cohabitation in weedy species ; In The genetics

of colonizing species (eds.) H G Baker and G L Stebbins (New York : Academic Pi ess)

pp. 243-268
Harper J L, Lovel P H and Moore K G 1970 The shapes and sizes of seeds ; Ann. Rev.

Ecol. Syst. I 327-356
Kamra S 1C 1967 Studies on storage of mechanically damaged seeds of Scots pine (Pinus

sylvestris L.) ; Stadia Forestalia Suecica 42 1-19

Hutchinson G E 1951 Copepodology for the Ornithologist ; Ecology 32 571-577
Kapocr P and Ramakrishnan P S 1973 Differential temperature optima for seed germination

and seasonal distribution of two populations of Chenopodium album L. ; Curr. Sci. 42

838-839
Koller D, Mayer A M, Poljakoff-Mayber A and Klein S 1962 Seed Germination ; Annual

Review of Plant Physiology 13 437-464
Kozlowki T T 1971 Growth and development of trees Vol. 1 (New York : Academic Press)

pp. 444
Kozlowski T T 1979 Tree growth and environmental stresses (Seattle : Washington Univeisity

Press) pp. 192
Ng F S P 1978 Strategies of establishment in Malayan forest trees ; In Tropical trees as living

systems (eds.) P B Tomlinson and M H Zimmermann (Cambridge Univ. Press) pp. 129-162
Quick C R 1961 How long can a seed remain alive ? ; In Seeds, The Yearbook of Agri-
culture (Washington D.C. : U.S. Govt. Print. Off.) pp. 94-99
Rarnakrishnan P S 1972 Individual adaptation and its significance in population dynamics ;

In Biology of land plants (eds.) V Puri, Y S Murti, P K Gupta and D Banerji (India ;

Meerut : Sarita Prakashan) pp. 344-355

Ross M A and Harper J L 1972 Occuptation of biological space during seedling establish-
ment ; /. Ecol. 60 77-88
Salisbury E J 1942 The reproductive capacity of plants (London : G Bell and Sons Ltd.)

pp. 224
Thompson P A 1973 Geographical adaptation of seeds ; In Seed ecology (eds.) W Heydecker

(Penn. Univ. Park : Penn. State Univ. Press) pp. 31-58
Wearstler K A and Barnes B V 1977 Genetic diversity of yellow birch seedlings ; Can. J. Rot.

55 2778-2788
\V[areing P F 1963 The germination of seeds ; In Vistas in botany Vol. Ill (eds.) R C Rollins

and C Taylor (New York : The MacMillan Co.) pp. 195-227

Proc. Indian Acad. Sci. (Plant Sci.), Vol. 91, Number 5, October 1982, pp. 409-416.
© printed in India.

Anther and pollen development in cotton haploids and their parents

S S MEHETRE

Department of Botany, Mahatma Phule Agricultural University, Ralmri, District,
Ahmednagar (MS) 413722, India

MS received 29 October 1981 ; revised 29 July 1982

Abstract. Development of anther tapetum from premeiotic stages to pollen
formation was studied in six x-ray induced haploids of Gossypium hirsutum, three
interspecific F2 haploids, and one natural haploid of each of G. hirsutum and
G. barbadense, and the observations were compared with those of their respective
parents, a genetic male sterile, a male fertile and a cytoplasmic male sterile line of
G. hirsutum. Significant diffeiences were recorded for number of anthers per
flower, pollen size, pollen viability and number of microspores produced by PMC.
Anther development in haploids was normal. Anther dehiscence was also normal
in some haploids. Non-dehiscent anthers could be mostly attributed to the forma-
tion of immature pollen grains. Normal development of anthers and degeneration
of tapetum occurred in the parents and m the genetic fertile line. Contrastingly
no degeneration of tapetum was noticed in the cytoplasmic male sterile line.

Keywords, Gossypium spp. ; haploids ; anther tapetum ; male sterile.

1. Introduction

Haploids are characterized by significant decrease in size of vegetative plarrt
parts, vigour and fertility (Kostoff 1943) and in diameter of pollen mother cells
(Belling and Blakeslee 1923) by half of that in the diploid (Ivanov 1938), and by
smaller guard cells (Lamm 1938). Kimber and Riley (1963) indicated a
relationship between haploid and diploid guard cells of G. hirsutum and
G. barbadense by a factor of 1-26.

Partial or complete sterility due to halving of chromosome number and several
meiotic irregularities in haploids of G. hirsutum, G. barbadense, their F2 inter-
specific crosses and x-ray induced haploids have been reported (Mehetre and
Thombre 1981b, c, d). Studies were undertaken to investigate the development
of anther tapetum and its role in pollen sterility observed in these haploids. The
comparative observations made on haploids, their respective parents, genetic male
sterile and fertile lines and one cytoplasmic male sterile line have been reported

409

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2. Material and methods

The length of flower bud at various meiotic stages was determined by studying
meiosis in the fertile counterparts and haploids. Flower buds of different
haploids., their respective parents, genetic male sterile and fertile lines and
cytoplasmic male sterile line (table 1) were collected from premeiotic to
pollen formation stages and fixed in Randolph's Graf ; after dehydra-
tion, the anthers were embedded in paraffin. Sections of 12 JLCTR were
cut and stained with iron alum hematoxylin (Johansen 1940). Pollen viability
(fertility /sterility) was tested by differential staining with a solution comprising
among other organic components malachite green, acid fuschin, and orange G.
(Alexander 1969). The data collected from 25 observations for each of the
parameters mentioned in table 1 were analysed statistically (Panse and Sukhatme
1953) and standard deviations and significance of differences between means were
calculated.

3. Results and discussion

Significant differences were noticed between haploid and diploid ^plants in size of
flowers and and roecia, in number of anthers per flower (figure 1), tetrads per micro-
spore (figure 2) and in pollen size and pollen sterility (figure 3). In haploids
the average number of microspores resulting from a PMC ranged from 3 '11 to
4*78. The large variation observed in pollen size (figure 3) in all the haploids
indicated that the pollen grains contained varying number of chromosomes.
Although well-developed exine and spines were noticed on some pollen grains,
probably microspore mitosis and starch formation had not occurred in them,
thus resulting in pollen sterility.

In the- cytoplasmic male sterile line the tapetum was. well developed and its
cells were enlarged. There was normal differentiation of anther wall, but the
sporogenous tissue collapsed early during meiosis ; meiosis did not proceed
beyond prophase and hence the tapetum remained intact and enlarged (figure 6).
Similar observations were recorded by Murthi and Weaver (1974) and Mehetre
and Thombre (198 la) for the cytoplasmic and genetic male sterile (MS5 and MS6)
stocks of G. hirsutum. In the present study on the genetic male sterile stock
normal development of anther tapetum was noticed, but the microspores aborted
due to development of vacuoles in them (figure 7). In the male fertile counter-
parts (figure 8), in all the tetraploid parents and in all the haploids (figures 4, 5)
the tapetal cells begin to digenerate at the time of separation of microspores from
the tetrads.

In the fertile lines the microspores develop a thick exine and a thin intine. Spines
develop on the exine and the germ pores become distinct. The microspore
nucleus divides to produce the generative nucleus and the vegetative nucleus ;
at this stage the pollen grain is considered to be mature and ready for shedding.
Similar observations were reported by Murthi and Weaver (1974) and Mehetre
and Thombre (1981a) for male fertile anthers and Mehetre (1981) for triploid
(3x = 39) and tetraploid (4x = 52) anthers. In the haploids, however, after separa-
tion of the microspores from tetrads the exine may become well developed but the

Anther and pollen development

413

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Anther and pollen development 415

spines are not of uniform size, the germpores are not distinct and the pollen grains
do not mature because the mircospore nucleus does not undergo division, as was
also observed in the parents of the haploids, such underdeveloped immature pollen
does not contain sufficient starch grains.

Decrease in the radial dimensions of tapetal cells occurred in the fertile materials
between anaphase I and pollen formation, while in the haploid plants the decrease
vas observed between anaphase I and tetrad stage only. The extent of decrease
vas variable from anther to anther, flower to flower and plant to plant.

The data on measurements of radial width of anther tapetum in parents of
laploids and in the genetic fertile line indicated that in parents of haploids the
legeneration of anther tapetum continues progressively from meiotic anaphase
;o pollen stages and it ranged from 3 to 6 microns at pollen stage, while in the
genetic male sterile line the tapetum remained intact even after microspore tetrad
itage. Pollen abortion occurred due to vacuolation of pollen caused presumably
yy nutritional differences, while in the cytoplasmic male sterile line meiosis did
lot proceed and hence the tapetum remained intact. A similar behaviour of
mther tapetum was also reported by Brooks et al (1966) in anthers of different
genetic and cytoplasmic male sterile, make fertile and fertility restorer lines of
jorghum by Murthi and Weaver (1974) and Mehetre and Thombre (1981a) in
cotton.

In all the three groups of haploids a marked variation in the width of tapetal
;ells from pre-prophase to pollen stage was observed. In some individuals, the
legeneration of tapetum was rapid while in some individuals it was slow. Although
dgniflcant differences in tapetal cell width were noticed in tetraploid and diploid
)lants, it was not in a 1 : 2 ratio. The pollen abortion and sterility was mainly
iue to microspores containing variable number of chromosomes and pollen with
ligh variation in size and probably not due to the abnormal development of
;apetum.

References

Vlexander M P 1969 Differential staining of aborted and non-aborted pollens ; Stain Techno!.

44 117-122
Jelling J and Balakeslee A F 1923 The reduction division in haploid, diploid and tetraploid

Daturas ; Proc. Natl. Acad. Set. 60 106-111
Jrooks M H, Brooks J S and Chien I 1966 The anther tapetum in cytoplasmic genetic male

sterile sorghum ; Am. J. Bot. 53 902-908
vanov M A 1938 Experimental production of haploids in Nicotiana rustica L. ; Genetica 20

295-397

bhdnsen D A 1940 Plant Microtechnique, Tata McGraw-Hill Co. 2nd ed.
Cimber G and Riley R 1963 Haploid angiosperms ; Bot. Rev. 29 480-531
iCostoff D 1943 Haploicie Triticum vulgare and die Variabiliabat inrer diploiden Nachkommens

schaften ; Zuchter. 15 121-125

R 1938 Note on haploid potato hybrid ; Herediatas, 24 39
S S 1981 Anther and pollen development in traploid (3* = 39) and tetraploid (4* = 52)

plants in cotton (Gossypium spp.) Phytomorphology (in press)
vfehetre S S and Thombre M V 1981a Stages of pollen abortion in male sterile stocks of

Gossypium hirsutum L.; /. Maharashtra Agril. Universities 6 159-161
•lehetre S S and Thombre M V 1981b Cytomorphological studies in x-ray induced glandless

haploids in Gossypium hirsutum L. cotton ; Proc. Indian Acad. Sci. (Plant Sci.) 90 313-322

416 S S Mehetre

Mehetrc S S and TJiombre M V 19Slc Meiotic studies in the haploids (2n = 2x[= 26) of tetra-

ploid cottons (2n = 4x = 52) ; Proc. Nat 1. Sci. Acad. B47 516-518
Mehetre S S and ThombreM V 1981 d Microsporogenesis in interspecific F2 haploids of cotton ;

Phytomorphology (in press)
Murthi A N and Weaver J B 1974 Histological studies on the five male stcrik strains of

upland cotton ; Crop Sci. 14 658-662
Panse V G and Sukhatme P V 1953 Statistical methods for agricultural workers ICAR, New

Delhi

Proc. Indian Acad. Sci. (plant Sc/.), Vol. 9l, Number 5, October 1982, pp. 417-426.
© Printed in India.

Changes in proteins, amino and keto acids in different seedling
parts of Cyamopsis tetragonolobus Linn, during growth in light and
darkness

PREM GUPTA and D MUKHERJEE

Department of Botany, Kurukshetra University, Kurukshetra 132119, India

MS received 23 May 1981 ; revised 21 April 1932

Abstract. Comparative changes in protein, free amino and keto acids have been
studied in different seedling parts of Cyamopsis tetragonolobus plants in light and
dark. Endosperm recorded higher level of free amino acid? in darkness than in
light, while a low concentration of protein was exhibited both in light and dark.
The breakdown of soluble protein was more in darkened cotyledon due to higher
protease activity. The large increase in the free amino acids in the hypocotyl during
seedling growth in the dark may be due to its restricted capacity to incorporate all
the amino acids into proteins. Root samples from light recorded higher soluble
protein as well as a higher free amino acid pool. a-Oxoglutaric acid (a-OGA) was
recorded in low levels and at few growth stages in both light and dark. In light
raised cotyledon samples, the dominating keto acids are phosphoenolpyruvate and
pyruvic acid. Low levels of oxaloacetate in light, like a-OGA, indicate its rapid
utilization during growth, but its accumulation in the dark may suggest sluggish
protein synthesis thus sparing the utilization towards the synthesis of amino acids.
Utilization of asparagine and glutamine was also affected in dark.

Keywords. Seedling parts ; protein ; amino acids ; keto acids ; protease activity ;
Cyamopsis tetragonolobus.

1. Introduction

The correlative changes in amino and keto acids have been studied during seed
germination and seedling growth (Fowden and Webb 1955 ; Webb and Fowden
1955 ; Krupka and Towers 1958a, b; Mukherjee 1972 ; .Mukherjee and Laloraya
1974, 1979, 1980). Recently, studies have been carried out in our laboratory
(Gupta 1981 ; Afria and Mukherjee 1980, 1981) of the comparative changes in
aforesaid metabolites along with organic acids and soluble protein in different
seedling parts of various plants so that proper assessment could be made of their
mobilization and/or breakdown at various growth stages. Leguminous plants
can be divided into endospermic and non-endospermic ones depending upon
whether the endosperm has been retained into maturity or not. Metabolic

417
P.(B)-6

4i8 Prem Gupta and D Mukherjee

changes during development of endospermic legumes have received less attention
in comparison to the other group during seedling growth. For this reason,
various biochemical changes with growth in Cyamopsis tetragonolobus, an endo-
spermic legume, have been studied here. In this paper the comparative changes
in soluble proteins, free amino acids, keto acids and protease activity have been
described in endosperm, cotyledon, hypocotyl and root of this endospermic
legume, during the early stages of seedling growth in light and dark.

2. Material and methods

Seeds of Cyamopsis tetragonolobus Linn, were surface sterilized with 0- 1 % mercuric
chloride for 2 to 3 min followed by thorough washing. Acid treatment was given
thereafter and washed thoroughly again with sterilized distilled water. Seeds were
germinated on filter paper discs moistened with distilled water in Petri dishes and
grown in darkness or light (2910 lux provided by fluorescent tubes) in a growth
chamber maintained at 30 ±1° C. Three replicates of 30 seeds each were taken for
each experiment. Every care was taken to select morphologically uniform seeds
and to ascertain least variability, experiments on growth in light and darkness
(table 1) were repeated thrice. Protein, free amino and keto acids were determined
quantitatively in endosperm, cotyledon, hypocotyl and root 48 hr after sowing
(termed " initial ") as well as 48, 72, 96 and 120 hr after * initial ' of both light
and dark grown seedlings.

Soluble proteins from fresh plant material were measured according to the
method of Lowry et al (1951) using Folin phenol reagent. The plant material
was boiled in 80% ethanol for 2 rnin on a water bath. It was allowed to stand
for 15 rnin at room temperature, ground in the same ethanol and centrifuged at
6000-7000 rpm for 5 min. Supernatant was discarded and the residue was again
extracted with 80% ethanol. Supernatant was discarded again and the residue
was extracted with 5 % perchloric acid, followed by centrifugation at 6000-7000
rpm for 5 min. Supernatant was discarded and the residue was taken out in a
test tube containing IN NaOH and kept for 30 min. in warm water (40-50° C)f
0-5 to 1*0 ml of this clear solution was taken and 10 ml of reagent C, which was
prepared by adding reagent A (2% sodium carbonate in O'lN NaOH) and
reagent B (0-5% copper sulphate in 1% sodium-potassium tartarate) in the
ratio of 50 : 1 (v/v), was added to it and allowed to stand for 10 min at room
temperature. Then added 1 ml of Folin's reagent (diluted twice) rapidly with im-
mediate mixing and allowed to stand for 30 min. The OD was measured at 540 nm
in a Bausch and Lomb Spectronic-20 colorimeter. The amount of protein was
determined in terms of Bovine Serum Albumin.

The extraction procedure used for amino acids, their chromatographic separa-
tion and estimations were the same as recommended by Steward et al (1954).
For keto acids the extraction procedure of Kaushik (1966) which is a slight modi-
fication of the method described by Towers and Steward (1954) has been followed.
Free amino and keto acids were quantified as glycine and a-oxoglutaric acid equi-
valents, respectively.

Changes in C. tetragonolobus Linn, during growth

419

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Prem Gupta and D Mukherjee

Protease activity — The method of extraction of the enzyme was a slight modi-
fication of that described by Yomo and Varner (1973) and Ihnen (1976). 1%
casein solution was prepared in 0-1 N NaOH. lOOmg of each seedling part
(at least in 3 replicates) was homogenized in 10 ml of 100 mM phosphate buffer
(pH 6-0) and centrifuged at 5000 rpm for 15 min. After filtration the pellet was
homogenized with the 5 ml of buffer and the process repeated thrice for maximum
recovery. All supernatants were combined so as to make the final volume to
25 ml. Each reaction set received 1 ml of the enzyme extract and 1 ml of casein
solution and the pH was 10. The blank set received 1 ml each of enzyme extract,
casein solution and 10 % Trichloro acetic acid (TCAL) (cold). These sets were incu-
bated at a temperature of 37 ± 2° C for 2.5 hr. 1 ml of 10% TCA (cold) was added
to each reaction set after the incubation period was over and both the sets centri-
fuged. After discarding the residue 1 ml of filtrate was taken from each set and
2ml of 0-5 N NaOH and 1 ml of 1 N Phenol Folin's reagent were added with
immediate mixing. These sets were allowed to stand for 30 min and OD was
taken at 540 nm in a ECI Junior Spectrophotometer. Protease activity was expressed
in n mol of tyrosine equivalent lur* g~~* tissue.

3. Results and discussion

Results have been summarized in tables 1-2 and figures 1-3.
3.1. Seedling growth in light and darkness

Table 1 shows that the growing axis did not differentiate 48 hr after soaking (' Ini-
tial ' stage) but at 48 hr seedling stage roots and hypocotyls were noticed and the

Table 2. C. tetragonolobus showing protease activity (n mol) Tyrosine x 108 hr""1 g"*1
(fr. wt.) in different Seedling parts in light and darkness.

Stagos (hr) Endosperm

Cotyledon

Hypocotyl

Root

Light

Initial

2'772±0-036

2-592±-249

*l-726± -106

48

0-684±OOOO

2- 880 ±-252

2-558± -249

4-464±-200

72

0 • 900 i 0-253

7-668±-165

2-520± -259

3*096±'l57

96

Tn»ce

l-332±-259

l-296± -165

0-576±-072

120

Trace

1-54S±-190

1-440± -252

2'9l6±-225

Dark

Initial

34-128i -374

59-148±-655

*16'488± 1-590

48

H-592± -53S

11 -988 ±-124

35'100±0-561

40-068±-533

72

8-100± -272

8-532±-409

2'772±0'036

4- 068 ±-252

96

0-792± -060

l'224±-252

0'648±0-000

3-060±-095

120

0'432± -000

l'728zb'286

1-836±0'000

1'404±-124

Data represents growing axis as hypocotyl and root did not differentiate.

Changes in C. tetragonolobus Linn, during growth

421

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Soluble Proteins Total Free AmJno Acids
« o COTYLEDON a *

o a ENDOSPERM ^ a

a e HYPOCOTYL o o

, ROOT ® ®

LIGHT

Cyamopsis tetrogonotobus

Initial 48 72 9$ 120 hr*

Initial

Figure 1. Cyamopsis tetragonolobus : showing levels of soluble protein— N and
total free amino acid pool in light and darkness in different seedling parts at various
growth stages.

length of hypocotyls was greater in darkness than in light. At 120 hr stage the
hypocotyls length of dark grown seedlings were 1ST 60% more than those raised
in light. Root growth also exhibited the same pattern as recorded for hypocotyls
but the increase ranged between 15 to 25% in dark as compared with light.

As regards the fresh weight changes, the endosperm was of greater weight in
darkness than in light at ' initial ' stage. But with further seedling growth, at
120 hr stage, the value in dark was lower than in light. Changes in percent dry-
mass showed a different pattern in that the endosperm from seedlings raised in
dark had always a higher value than those in light irrespective of the seedling growth
stages (table 1). Cotyledons at 120hr stage also had more fresh weight in light
than in dark. Moreover, percent dry mass although initially more or less of the
same value in light and dark decreased much less with further growth in the dark.

Hypocotyls, after differentiation, show more than three-fold increase in fresh
weight in dark compared with those in light. However, the dry weight values
were slightly lower in the former. Roots exhibited a small decrease in their
fresh weight both in light and dark.

Growth data presented here illustrate the two common phenomena of photo-
morphogenesis and etiolation in light and dark. Dark-grown seedlings having
stimulatory effects on hypocotyl lengthening recorded a linear increase and direct
relationship with the fresh weight but percent dry mass exhibited inverse relation-
ship indicating the failure of translocation of the products of reserve hydrolysis
to keep pace with the extension growth,

Prem Gupta and D Mukherjee

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Changes in C. tetragonolobus Linn, during growth

423

A PEP
8 Pyr
CX-OGA
D OAA
E Urea
F GLY

Initial 48 hr 72 hr 96 hr 120 hr

Figure 3. C. tetragonolobus : Keto acid changes in different seedling parts at
various growth stages. Abbreviations: PEP, phosphaenolpyruvate ; PYR, pyruvic
acid ; GLY, glyoxylic acid ; «-OGA, a-oxoglutaric acid ; OAA, oxaloacetic acid.

3-2. Biochemical studies

The studies with this endospermic legume revealed that the endosperm had low
soluble protein values initially which further decreased with seedling growth in
both light and dark (figure 1). Endosperm samples from dark treatments recorded
a higher number and amount of free amino acids, when compared to corresponding
light samples. In light, levels of free amino acids remained low during early
stage but dark-raised endosperm samples recorded increasing values up to the
72 hi stage, whereafter they declined (figure 1). Glutamic acid, a-alanine, leucine-
phenylalanine, serine-glycine, glutamine and histidine dominated quantitatively
in dark-raised endosperm samples (figure 2).

The breakdown of proteins could also be detected in cotyledons of both light
and dark grown seedlings. However, the depletion was more in dark since light
causes a retention of proteins as mentioned earlier (Rai and Laloraya 1967 ;
Mukherjee and Laloraya 1979). Along with the protein depletion although an
enhancement in the free aminoacid pool was expected in endosperm and cotyledons,
their marked difference and a large increase in the latter suggests different rates
and pattern of accumulation in light and dark and a rapid translocation of these

424 frem Gupta and D Mukherjee

metabolites (figures 1-2). The hydrolysis of endosperm reserves by enzymes
released from the aleurone layer and their absorption by cotyledons followed by
the translocation to the growing axis has also been noted by Bewley and Black
(1978). Moreover, unequal rate of protein breakdown and free amino acid forma-
tion in relation to light and darkness will also influence the transport of amino
acids to the growing axes. Further, many-fold increase in the free amino acids
in the hypocotyl during seedling growth in dark (figures 1-2) may be due to restric-
ted capacity of dark grown seedlings to convert all the amino acids into proteins
as also noticed by Srivastava and Kooner (1972) in Phaseolus aureus L. Oota et al
(1953) while studying the changes in the content of various primary metabolites
in germinating Vigna sesquipedalis beans showed a decline of these metabolites in
the cotyledons while the hypocotyls and roots recorded an increase during early
germination for six days. It is also proposed that the growing axes of light grown
seedlings are capable of amino acid biosynthesis by animation of carbon skeleton
produced in photosynthesis and this process accounts for increase in amino acid
levels (Bewley and Black 1978). There is a differential concentration of amino
acids in the hypocotyls under the two situations (figures 1-2).

Root samples in the light showed higher protein content in comparison to dark
raised samples at all stages and a very marked increase was noticed in the total free
amino acid pool at 48 hr stage. Further growth gave a sharp increase in the free
amino acid pool in light whereas in dark the decline was more marked and could be
observerd from the beginning (figure 2).

Proline and methionine were not widely distributed in different seedling parts of
C. tetragonolobus. Proline was recorded in cotyledons in both light and dark
while endosperm and root contained the same at only few growth stages. Proline
may be converted to glutamic acid thus increasing the pool size of this amino acid
as reported by Bewley and Black (1978). Cysteic acid was not traced in endosperm
and root at any growth stage of light and dark while cotyledon and hypocotyl
recorded the same in very low concentrations at few growth stages. Methionine
was also traced at few stages. y-Aminobutyric acid was found in higher amounts
in cotyledons of C. tetragonolobus in comparison to other seedling parts although
it was of widespread occurrence in different organs. Its higher amount had been
reported earlier also (Altschul 1958).

Changes in protease activity in various seedling parts in the light and darkness
are shown in table 2. Dark-grown seedlings showed a significantly higher activity
of this enzyme than those in light in all parts during growth. Initial stages of all
seedling parts were unique in having maximum protease activity. The maximum
decline in the activity was noticed in the endosperm and cotyledon which corre-
lated with their protein depletion.

a-Oxoglutaric acid, the predominant keto acid, was recorded only at few growth
stages mostly in low levels in both light and dark seedling parts of C. tetragonolobus.
In cotyledons of light grown seedlings, the dominant keto acids were phosphoenol-
pyruvate and pyruvic acid (figure 3), the levels of which were maintained during
seedling growth. Dark raised cotyledon samples had low values which declined
further (figure 3). A characteristic feature of the keto acids was a rapid increase
in their concentrations followed by later decline. Hypocotyls of light-raised seed-

Changes in tetragonolobus Linn, during growth 425

liags maintained higher levels of phosphoenolpyruvate, pyruvic acid, oxaloacetate
and urea up to 72 hr stage followed by a decline and then a small increase, while
in dark although above-mentioned keto acids could be detected in higher concentra-
tions, levels of hydrazones recorded a gradual decline after 72 hr stage (figure 3).
Higher levels of keto acids could also be found in roots. Low levels, in most
of the samples of oxaloacetate, the keto analogue and precursor of aspartic acid
like a-oxoglutaric acid, can be explained by their rapid utilization during seedling
growth. The tendency for accumulation of oxaloacetate especially at 120 hr stage
in endosperm, cotyledon and hypocotyl of dark-raised samples may suggest that
this keto acid was rapidly utilized in the light-induced growth of seedlings. It has
been suggested by Webb and Fowden (1955) that accumulation of keto acids is
related to sluggish rate of protein synthesis thus sparing the utilization of keto
acids for the synthesis of amino acids.

Higher amounts of keto acid hydrazones of urea in light and dark in C. tetra-
gonolobus account for its active role in nitrogen metabolism during seedling
growth (figure 3). Asparagine and glutamine, the two common amides, which
store excess ammonia to get rid of the toxic compound, recorded higher amounts
from hypocotyl of dark grown seedlings in comparison to light while root samples
from dark contained no detectable glutamine and asparagine content declined with
seedling growth (figure 2). Cotyledons of 120 hr seedling stage were unique in
exhibiting accumulation of asparagine and glutamine in dark in comparison to
those in light thus sparing the utilization of these compounds in protein synthesis
which is affected by dark.

Acknowledgements

We are grateful to Dr H S Choudhary of the Chemistry Department for help
and many valuable suggestions during the protease assay. Thanks are also due to
Prof. R S Mehrotra for laboratory facilities and to csm, New Delhi, for giving a
SRF to one of us (Prcm Gupta) during the tenure of this work.

References

Afria B S and Mukhcrjee D 1980 Biochemical studies during seedling growth of sweet pea

(Lathyrus odoratus L-) ; Proc. Indian Natl. Acad. Sci. 46 490-494
Atria B S and Mukherjee D 1981 Metabolic studies in Sorghum vulgare Pers. and Zea mays L.

during seedling growth ; Proc. Indian Acad. Sci. (plant scl) 90 71-78

Altschul A M 1958 Processed plant protein food stuffs (New York : Academic Press Inc.)

Bewley J D and Black M 1978 Physiology and biochemistry of seeds in relation to germination.

I. Development, germination and growth (New York : Springei-Verlag, Berlin Heidelberg)

pp. 193, 221
Fowden L and Webb J A 1955 Evidence for the occurrence of y-methylene a-oxoglutaric acid

in groundnut plants ; Biochem. J. 59 228-234
Gupta P 1981 Ecophysiology of germination in certain plants ; Ph.D. Thesis, Kurukshetra Univ.,

Kurukshetra (India)
Ihjaen, J L 1976 Lung proteases and protease inhibitors ; M.S. Thesis, University of Iowa, Iowa

aty (USA)
Kaushik D D 1966 Studies on certain aspects of plant metabolism ; Thesis for D.Phil. Degree

of University of Allahabad (India)

426 Prem Gupta and D Mukherjet*

Krupka R M and Towers G H N 195Sa Studies on the keto acids of wheat. I. Behaviour during

growth ; Can. J. Bot. 36 165-177
Krupka R M and Towers G H N 1958b Studies on the keto acids of wheat- It. Glyaxylic acid

and its relation to allantoin ; Can. J. Bot. 36 179-186
Lo.wry O H, Rosebrough N J, Farr A L and Randall R J 1951 Protein measurement with

the folin phenol reagent ; /. Biol. Chern. 193 265-275
Muklierjee D 1972 Keto acid metabolism lit certain plants ; D.Phil. Thesis, University of

Allahabad (India)
Mukherjee D and Laloraya M M 1974 Metabolism of y-methyl-a-ketoglutaric acid, 7-methyleae-

a-ketoglutaric acid and other keto acids during the seedling growth in Tamarindus indica ;

Biochem. Physiol Pflanz. 166 429-436
Mukherjee D and Laloraya M M 1979 Nitrogen and free amino acid changes during seedling

growth in Banhinia purpurea ; /. Indian hot. Soc. 58 75-82
Mukherjee D and Laloraya M M 1980 Changes in the levels of keto acids in different parts

of seedlings of Bauhinia purpurea L. during growth ; Plant. Biochem. J. 7 120-125
Oota Y, Fuzii R and Osawa S 1953 Changes in content of protein nitrogen of embryonic

organs in Vigna sesquipedalis during germination ; /. Biochem. Tokyo 40 649
Rai V K and Laloraya MM 1967 Correlative studies an plant growth and metabolism.

II. Effect of light and gibberellic acid on the changes in protein and soluble nitrogen

in lettuce seedlings ; Plant Physiol. 42 440-444
Srivastava A K and Kooner N K 1972 Physio-logical and biochemical studies in seed germination

of moong (Phaseolus aureus Roxb.) ; Indian J. Exp. Biol. 10 304-306
Steward F C, Wetmore R H, Thompson J F and Nitsch J P 1954 A quantitative chromato-

graphic study of nitrogenous components of shoot apices ; Am. J. Bot. 41 123-134
Towers G H N and Steward F C 1954 The keto acids of Tulip (Tulipa gesneriana) with special

reference to the keto analog of y-methylene glutamic acid ; /. Am. Chcm. Soc. 76 1959-

1961
Webb J A and Fowden L 1955 Changes in oxo-acid concentrations during the growth of

groundnut seedlings ; Biochem. J. 61 1-4
Yomo Harugo-ro and Varner J E 1973 Control of the formation of amylases and proteases in

the cotyledons of germinating peas ; Plant Physiol 51 708-713

Proc. Indian Acad. Sci. (Plant Sd,)3 V°l* 91, Number 5, October 1982, pp. 427-431.
© Printed in India.

Effect of ridge gourd pollen on zoospore germination of
Pseudoperonospora cubemis and its significance in epidemiology

AMARNATHA SHETTY, H S SHETTY and K M SAFEEULLA
Downy Mildew Research Laboratory, University of Mysore, Mysore 570006, India

MS received 29 October 1981 ; revised 3 June 1982

Abstract. Ridge gourd pollen has a stimulatory effect on the germination of
Pseudoperonospora cubensis. The rate and percentage germination of zoospores
increased in the presence of pollen leachates. Spraying of leaves with a mixture
of pollen and sporangial suspension enhanced the development cf lesions. Early
germination of zoospores in the presence of pollen proved advantageous for infec-
tion as it provided a prolonged favourable infection period. The results are
discussed in relation to the epiphytotics o.f the disease during flowering period.

Keywords. Pseudoperonospora cubensis ; ridge gourd; pollen effect ; zoospore
germination ; epidemiology.

1. Introduction

Pseudoperonospora cubensis (Berk, and Curt.) Rostow, the incitant of cucurbit
downy mildew is one of the serious and production limiting diseases of Luffa
acittangula Roxb. in India. The disease attains serious proportions when the
plants start flowering and as a result the susceptible varieties of plants fail to
produce fruits. Bains and Jhooty (1975) reported that in Cantaloupes downy
mildew appeared during 1972-74, under field conditions only during the flowering
and fruiting stage. The effect of host pollen on stimulation of spore germination
of fungal pathogens has been worked out in different crops (Chou and Preece
1968 ; Fokkema 1976 ; Preece 1976 ; Meenakshi and Ramalingam 1979 ;
Suryanarayana and Ramalingam 1979). So far no report on the effect of host
pollen on germination of downy mildew pathogens has been made.

2. Materials and methods

2.1. In vitro effect of pollen on zoospore germination

One of the susceptible varieties of ridge gourd (Pusa Nasdar) was .grown in the
downy mildew experimental plots to obtain the sporangial inoculum and the host

428 Arnaniatha Shetty, H S Shetty and K M Safeeulla

pollen. Sporangial suspension was prepared by the following method : downy mildew
infected leaves were collected at 6 p.m. and the remnants of the downy growth
was washed off with moist cotton under running tap water. The leaves were air
dried and small bits of leaves with lesions were cut and placed inside petri plates
containing a wet blotter with the adaxial surface of the leaf in contact with the
blotter. A good crop of sporangia was obtained after incubating the leaf bits
for 12 hrs at 22° C in dark. The sporangia were scraped with a blade into a dish
containing distilled water. The concentration of sporangial suspension was
measured using a haemocytometer and was adjusted to about 10,000/ml. Host
pollen was collected and stored at 5° C. Five mg of the pollen was mixed in
10ml of the sporangial suspension. A suspension of the mixture was placed on
glass slides and incubated in moist chambers at room temperature (22-26° C).
In controls no pollen was added. Observations for zoospore germination were
made under binocular microscope and results were recorded at hourly intervals
after fourth hour.

2.2. Effect of pollen on infection of host leaves and lesion development

Pollen plus sporangial suspension was sprayed on the lower surface of the leaf
of 20-30 day old plants and retained inside a glass house. The leaves were covered
with moist polythene bags for about 24 hrs and observations for number and size
of lesions were made. The effect of pollen in reducing the infection threshold was
tested for moisture requirement by covering the leaves with polythene bags for
3, 4, 5 and 6 hrs. After removing the polythene bags the leaf was air dried and
left inside the glass house.

2.3. Disease development in the field in relation to age of the crop

Two varieties of ridge gourd viz. Pusa Nasdar, a highly susceptible variety and
long variety which is moderately resistant ware sown in the month of August
1980 in plots. Disease rating was made at weekly intervals using a 0 to 5 scale
as described by Thomas (1977). Fertiliser (NPK 17 : 17 : 17) was applied twice
at the age of 20 days and 45 days.

3. Observations

3.1. In vitro effect of pollen on zoospore germination

Sporangial suspension when incubated at room temperature (22-26° C) released
zoospores within H— 2 hrs. Maximum number of zoospores were observed after
2 hrs of incubation. Zoospores remained active in water for 90 to 120 min and
then encysted. The data with regard to percentage of zoospore germination and
germ tube length with and without pollen are recorded in figures. 1 and 2 respec-
tively. Zoospore germination started an hour earlier in the presence of pollen.
The percentage of zoospore germination and vigour of the germ tubes in the
presence of pollen was greater compared to the zoospore which germinated in the
absence of pollen.

Pseudoperonospora cubensis and its significance

429

80-

'£ -

£ 40-

D Treatment
E3 Control

678
Time (hr)

10

Figure 1. Effect of host pollen on the germination of zoospores of P. cubensis.

50

Z 30
o

10

Treatment
Control

20

60

Length (yxm)

100

140

Figure 2. Comparison of the germ tube length of zoospores in presence of pollen
and in distilled water and their relative frequency of occurrence.

3.2. Effect of pollen on infection of host leaves and lesion development

Difference in disease reaction was apparent in plants sprayed with the mixture
and the sporangial suspension alone. The number and size of lesions were more
on leaves sprayed with the pollen mixture. Leaves inoculated with sporangial
suspension required a minimum of four hrs of leaf wetness for successful infec-
tion under glass house conditions. Only 3 hrs of leaf wetness was needed for
infection in the presence of pollen under the same conditions of temperature
and inoculum concentration.

3.3. Disease development in the field in relation to age of the crop

Downy mildew of ridge gourd makes its appearance at the seedling stage. The
young true leaves are resistant to downy mildew. When the seedlings attain the

430

Amamatha Shetty, H S Shetty and K U Safeeiilla

age of 20 days (3-5 true leaf stage) symptoms appear on the true leaves as
greenish to yellow lesions. Disease appearance in " Long ' is delayed by a week
in comparison with Pusa Nasdar. Severity of disease in relation to age is plotted
in figure 3. Flowering in both the varieties start 35-40 days after planting. Till
then disease severity is less than stage 3 of the 0-5 scale. Soon after flowering
the disease reaches severe proportions and it ultimately results in the death of
vines in c Pusa Nasdar 9 whereas in c Long ' it is not very severe and the
vines continue to grow but the yield is significantly reduced.

4. Discussion

In saprophytic fungi and facultative pathogens the stimulating effect of pollen on
germination of conidia is attributed to carbohydrates (Suryanarayana and Rama-
lingam 1979 ; Fokkema 1976) but the aggressiveness of such fungi depends on the
pollen leachates, rather than the nutrients (Chou and Preece 1968). In P. cubensis
the zoospores germinate in distilled water thereby showing that it is not nutrient
dependent. Hence it is quite probable that pollen leachates provide a stimulatory
effect on zoospore germination.

Preece (1976) stated that the effect of 'pollen on leaf infection may be due to
(a) increase in the speed and rate of spore germination, (b) restoring the germina-
bility and infectivity of old spores and (c) reducing the infection threshold by
enhancing the speed and virulence of the pathogen. From the present study, it
is evident that there is an increase in the number of zoospores germinating and the
vigour of germination is enhanced. Under field conditions, sporangial liberation
occurs during morning hoars (Cohen and Rotem 1971 ; Thomas 1977 ; Bains
and Jhooty 1978). It starts at around 6 ajn. and reaches a peak at 8 a.m. For the
successful infection of a fresh host leaf a minimum of 4 hrs of leaf wetness is
needed. But when the zoospore germinates in presence of host pollen it needs
only a period of 3 hrs for infection.

cu

I

P 3

o

O)

5

• \/eqetatlve phase

Reproductive phase

Pusonasdar
Long var

20 40 60

Age of plant (days)

80

100'

Figure 3. Disease progress in two varieties of ridge gourd in relation to a$e of
the plant, " -.••»...-•

Pseudoperonospora cubensis and its significance 43 i

The sporangia liberated during morning hrs are subjected to a period of
unfavourable conditions which last until the next dew fall and infection of host
plant occurs during night hrs (Cohen and Rotem 1971 ; Cohen and Eyal 1980).
From our experiments under Mysore conditions, it is quite probable that those
sporangia liberated early in the morning get deposited on the leaves and they
start germinating by the production of zo»ospores. As dew persists at Mysore
condition till 9*30 to 10 a.m., infection of the leaf tissue in presence of host pollen
can occur successfully during the daytime. In addition, those sporangia which
are deposited in later hours of the day, i.e., those sporangia which fail to infect
during the morning hours due to the advent of unfavourable period for infection,
survive till the next dew fall with a considerable loss in viability of sufficient
number of sporangia. Those viable sporangia germinate and cause infection
during night hours. As a result, severity of the disease increases significantly
during flowering period. This is supported by the results of studies of lesion
development and zoospore infection under different leaf wetness periods.

Acknowledgements

The senior author is grateful to the Council of Scientific and Industrial Research,
New Delhi, for the award of a Junior Research Fellowship.

References

Bains S S and Jhoofcy J S 1975 Studies on the epidemiology of downy mildew of muskjnelon

caused by Pseudoperonospora cubensis (abstract) ; Indian J. of Mycol. and PL PathoL 5

46-47
Bains S S and Jhoaty J S 1978 Epidemiological studies on downy mildew of nmskmelon

caused by Pseudoperonospora cubensis ; Indian PhytopathoL 31 42-46
Chou M and Preece T F 1968 The effect of pollen grains on infection caused by Botrytis

cineria ; Ann. Appl. BhL 62 11-22
Cohen y and Rotem J 1971 Dispersal and viability of sporangia of Pseudoperonospora cubensis ;

Trans. Br. Mycol. Soc. 57 67-74
Cohen. Y and Eyal H 1980 Effects of light during infection on the incidence of downy mildew

(Pseudoperonospora cubensis) on cucumbers ; Physiol. PL PathoL 17 53-62
FokJkema N J 1976 Antagonism between fungal saprophytes and pathogens on aerial plant

surfaces in Microbiology of aerial plant surfaces (ed.) C H Dickinson and T F Preece

(London : Academic Press) pp. 487-506
Meenakshi M S and Ramalingam A 1979 The effect of sorghum pollen on the germination

of Drechslem turdca (Pass.) Subram. and Jain ; Curr. Sci. 48 447-448
Preece T F 1976 Some observations of leaf surfaces duiing the early stages of infection by fungi

in biochemical aspects of plant parasite relationships, (ed.) J Friend and D R Threlfall

(London : Academic. Press) pp. 1-10
Suryanarayana K and Ramalingam A 1979 Influence of pollen on the germination of conidia

of Drechslera turdca (Pass.) Subram. and Jain ; Curr. Sci. 48 1045-1047
Thomas C E 1977 Influence of dew on the downy mildew of cantaloupes in South Texas ;

Phytopathology 67 1368-1369

Proc. Indian AcadL Sci. (Plant Sci.), Vol. 91, Number 5, October 1982, pp. 433-441.
© Printed in India.

Leaf proteiuase and nitrate reductase activities in relation to grain
protein levels and grain yield in four species of grain amaranth

K RAMAMURTHY NAIDU, Y SEETHAMBARAM and
V S RAMA DAS*

School of Life Sciences, University of Hyderabad, Hyderabad 500 134, India
* Department of Botany, Sr-i Venkateswara University, Tirupati 517502, India

MS received 24 November 1981 ; revised 19 August 1982

Abstract. The relationship of leaf nitrate reductase (NR) and proteinase acti-
vities to the grain protein level and grain yield was investigated in four species of
grain amaranth (Amaranthus hypochondriacus, A. caudatus, A. cruentus and A. edulis).
A strikingly positive correlation between the leaf proteinase activity and the grain
protein content was found. A. edulis with higher gr?in protein level possessed
high leaf proteinase activity, while A. hypochondriacus, with relatively lower grain
protein content had lower leaf proteinase levels. Although there was no definite
correlation between the leaf proteinase levels and the grain yield, the integrated
leaf NR activity was positively correlated with the grain yield. The total nitrogen
content per plant seems to be dependent on the extent of rcot growth and the
levels of NR activity in leaves.

Keywords. Grian amaranth ; leaf proteinase activity ; leaf nitrate reductase acti-
vity ; grain protein ; grain yield.

1. Introduction

Grain amaranth is presently one of the under-exploited crop plants with a consi-
derable economic potential. The grain is regarded to be unique for its high
protein content. It is also known to be rich in lysine and sulfur containing amino
acids and can therefore be considered superior to the proteins of wheat, corn and
rice (Senfit 1980).

Significant correlations were found in the past between the integrated leaf
nitrate reductase (NR) activity and grain yield, reduced nitrogen levels of grain
and of whole plant in the case of wheat and maize (Abrol and Nair 1978 ;
Brunetti and Hageman 1976 ; Hageman 1979). On the other hand Dalling et al
(1975) found that wheat cultivars with similar levels of NR activity could accumulate
variable amounts of reduced nitrogen. Deckard et al (1973) identified one maize
genotype with relatively low NR activity but a high capacity to accumulate reduced

; 433

P.(B)-8

434 K Ramamurthy Naidu, Y Seethambaram and V S Rama £>as

nitrogen. However, several studies have indicated that NR assays could be a
useful predictive selection criterion for grain yield and grain protein levels (Croy
and Hageman 1970 ; Fakorede and Mock 1978). Not much work was done on
the role of proteinases of leaf in relation to the protein content on grain. That
leaf proteinase activities were correlated with grain nitrogen was evident in wheat,
rice and maize (Bailing et al 1976 ; Perez et al 1973 ; Reed et al 1980).

In view of meagre work on grain amaranths in general and its nitrogen meta-
bolism in particular, the present study was carried out, leading to an under-
standing of the relationship between the leaf proteinase activity, grain protein,
grain yield and secondly between the leaf NR activity, root growth and the accu-
mulation of reduced nitrogen in four species of grain amaranth.

2. Materials and methods

Seeds of Amaranthus hypochondriacus, L., Arnaranthus caudatus L., Amaranthus
cruentus and Amaranthus edulis L. were obtained from National Botanical
Research Institute, Lucknow. Plants were grown in 30cm diameter earthenware
pots on soil supplemented with manure (3 parts of red soil 4- 1 part of farm
yard manure) under natural (approximately 12 hr) photoperiod (temperature
about 35° C day and 22° C night). Three plants were retained in each pot.

The plants were harvested at three different stages, viz., vegetative stage (30 days
after sowing), flowering stage (45 days after sowing) and grain filling stage (60
days after sowing). At each stage, plants from 3 pots were collected and were
subdivided into leaf, stem, root and panicle. Fresh and dry weights of these
plant parts were determined and the shoot/root ratio was calculated.

Reduced nitrogen content in the dried samples was determined by kjeldahl method
using Tecator digestion and distilling systems (Tecator Manual). After anthesis
(90 days after sowing) plants were finally harvested to the ground level and the
dry weights of the panicle and stover were determined. Grain protein content
was calculated by multiplying the grain nitrogen by a factor of 6*25.

2.1. Nitrate reductase (NR) assay

Leaf NR activity was measured at the vegetative, flowering and grain filling stages.
Fully expanded young leaves from each pot were collected (at 10 a.m.) into a poly-
ethylene bag placed on an ice bath and were carried into the laboratory. The
leaves were deribbed, weighed and were then chopped into pieces. The in vitro
NR assay was as described by Hageman and Hucklesby (1971).

2.2. Proteinase activity

The proteinase activity was measured thrice at 15 day intervals after anthesis
(60, 75 and 90 days after sowing). The proteinase activity in the middle leaf of
the plant was assayed by the modified procedure of Peoples and Dalling (1978).
Leaves were homogenised by grinding in a mortar and with a pestle for 90 sec
with 5 ml/g extracting medium containing 23 mM sodium citrate; 155mM

Leaf proteinase and nitrate reductase activities

435

sodium phosphate ; 5mM L-cysteine; 5mM EDTA and 1% PVP, pH 6-8. After
straining through cheese cloth the homogenate was centrifuged at 25000 g for
10 min. The supernatant was dialysed at 4° C for 48 hr against 50 mM potassium
phosphate buffer, pH 7*0.

1% Bovine serum albumin solution was prepared in 0-05M Tris-HCl
(pH 7 -8). 0- 1ml of extract was incubated with 0"5ml of substrate for 2 hrs
at 37° C. The reaction was terminated by adding 0'7 ml of 15% trichloroacetic
acid and the soluble nitrogen in the mixture was determined by ninhydrin
(Spices 1957). Leucine was used as the amino acid standard.

3. Results

3.1. Shoot dry weight

There was a wide variation in the dry weight of shoot at the vegetative stage
between the four species of grain amaranth studied (table 1). Maximum dry
matter production during the vegetative stage was in A. hypochondriacus followed
by A. edulis, A. caudatus and A. cruentus. Though a similar trend was observed
at flowering stage, the variation in dry matter content was not significant suggesting
that the growth rate during different stages of growth period varied among four
species (table 2). The higher dry matter accumulation noticed in A. caudatus
than that in A. hypochondriacus during filling stage may be due to the faster
growth rate in A. caudatus from flowering stage (table 3). The greater dry matter
accumulation in shoots of A. hypochondriacus (table 4) than the other three
species, might be due to the bigger panicles in the former species.

Table I. Dry weights and leaf reduced nitrogen during vegetative stage (30 days
after sowing) of four species of grain amaranth.

Species

Total dry wt/plant

- S/R

Leaf Reduced N
reduced of stover

Shoot

Root

Total

N%

g/plant

8i

g

g

A. hypochondriacus L.

9-0

0-80

9*80

11-25

3-85

0-2S

± 0-4

± o-i

± 0-45

± 0-25

± 0-12

± 0-02

A. caudatus L.

3-5

0-25

3-75

14-00

4-27

0-13

± 0-2

± 0-05

± 0-26

± 0-36

± 0-15

± 0-02

A. cruentus L.

3-1

0-40

3*5

7-75

4-13

0-11

± 0-3

± 0-04

± 0-32

± 0-15

± 0-08

± o-oi

A. edulis L.

5-8

0-45

6-25

12-89

3-99

0-19

i 0-4

± 0-03

± 0-42

± 0-28

± 0*07

± 0-03

436 K Ramamurthy Naidu, Y Seethambamm and V S Rama Das

Table 2. Dry weights and leaf reduced nitrogen during flowering (45 days after
sowing) of four species of grain amaranth.

Total dry wt/plant

Leaf Reduced N
reduced of stover
N% g/plant

kjpcwaca

Shoot

Root

Total

• kJ/iV

g

g

g

A.

hypochondriacus L. 44*0

8-2

52-2

5*36

3-92 '

1-27

± 1-8

± 0-5

±2-1

± 0-25

± 0-06

± 0-07

A.

caudatus L. 41 '5

9-5

51-0

4'37

4-47

1-39

± 2-4

± 0-7

±2-0

i 0-28

± 0-08

± 0-06

A.

emeritus L. 35*5

8-0

43-5

4'44

3-71

0-09

± 1-6

± 0-3

± 1-8

±0-16

± 0-04

± 0'05

A.

eduKsL. 37-5

7-5

45-0

5-00

3'64

1-02

± 1*5

± 0-2

± 1'4

± 0-32

± 0-05

± 0-05

Table 3. Dry weights and leaf reduced

nitrogen

during grain

filling (60

days after

sowing) of four species

of grain amaranth.

Total dry wt/plant

Leaf

Reduced N

- S/R

reduced of stover

Shoot

Root

Total

N%

g/plant

g

g

g

A.

hypochondriacus L. 82 '0

12-0

94-0

6-80

3-50

2-86

± 4-2

± 0-8

± 4-5

± 0-4

± 0-06

± 0-21

A.

caudatus L. 87-0

15-0

102-0

5-80

3-57

3'29

± 3'6

± 1-2

± 4-8

± 0'26

± 0*04

± 0-18

A.

emeritus L. 66*5

12-0

78-5

5-54

3-44

2-41

± 3-8

± 1-4

± 4-2

± 0-32

± 0-07

± 0-18

A.

edulisl*. 69*5

11-5

81-0

6-04

2-94

2-65

' ±2-3

± 0-7

± 3-1

± 0-22

± 0-08

d= 0-14

3.2. Shootf Root (SfR) ratio

The S/R ratio in all the four species was maximum at vegetative stage and decreased
gradually during flowering and grain filling stages (tables 1, 2 and 3). A. caudatus
had the highest S/R ratio among the four species during vegetative phase of
growth, but the ratio decreased during flowering and grain filling stages due to
in rpot dry weight at later stages of growth* ....

Leaf proteinase and nitrate reductase activities

437

Table 4. Mean NR activity (calculated from figure i)9 dry matter at final harvest
(90 days after sawing) grain yield, grain protein and mean proteinase activity
(calculated from figure 2) of four species of grain amaranth.

Species

NR
activity

Dry matter

Grain

# mol NOJ Panicle Stover Total g/plant
g/plant

Grain Proteinase Total N
protein activity g/plant

A. hypochon-
driacus L.

26'2 75-0 51'0 126'0 23'6 12*5 21*6 2-42

±1-4 ±2-5 ±1-0 ± 3-4 ±1-8 ±0-4 ±0*8 ±0-12

A. caudatus L. 23'43 45-0 60" 0

± 0-8 ± 2-2 ± 1-8

105-0 16-8 14-0 31-5 2'96

± 4-0 ± 0-7 ± 0*3 ± 0-6 ± 0-18

A. cruentus L. 20*07 44*0 58'0 102*0 15'6 12'6 28-4 2-15

±1-2 ±1-9 ±2-1 ± 3-8 ±0-6 ±0-2 ±0-7 ±0-11

A. edulis L. 24' 17 69*0 46'0 115'0 21*8 15*0 35*0 2«24

± 0-9 ± 1-7 ± 1-4 ± 3-1 ± 0-7 ± 0-3 ± 1-2 ± 0-09

3.3, Levels of leaf reduced nitrogen (%) and grain protein (%)

Variation in the concentration of leaf reduced nitrogen was observed among the
four species of grain amaranth. Maximum protein content was in grain of
A. edulis while the maximum content of leaf reduced nitrogen was recorded in
A. edulis (table 4).

3.4. NR activity and grain yield

The greatest NR activity was at the flowering stage of all the four species of grain
amaranth and the activity decreased at grain filling stage (figure 1). The level
of NR in A. hypochondriacus was more than that in the other three species during
vegetative and flowering stages whereas, at grain filling stage, A. caudatus had
the greatest NR activity. There was a positive correlation between the mean NR
activity the three stages and grain yield (r = + 0-89) in all the four species of
grain amaranth.

3.5. Proteinase activity

The results of the mean proteinase activity calculated from figure 2 are given in
table 4. The level of leaf proteinase activity was in the decreasing order of
A. edulis followed by A. caudatus ; A. cruentus and A. hypochondriacus. The
leaf proteinase Activity was positively correlate^ with the percentage of proteiu

438

K Ramamurthy Naidu, Y Seethambaram and V S Rama Das

20

o A.hypochondriacus

® A. caudatus

A A.cruentus

A A- edulis

Vegetative Flowering Grain fillinq

Figure 1. The leaf in vitro nitrate reductase activity of four species of grain
amaranth during different stages of growth.

60

50

40

30

o

20

10

O A. hypochondriacs

• A. caudatus

A A. cruentus

A A. edulis

15 30

Days after anthesis

45

Figure 2. The leaf proteinase activity of four species of grain amaranth.

in grain (r = + 0'85) in all the four species. Leaf proteinase activity increased
during grain development (figure 2).

4. Discussion

The total reduced N per plant was positively correlated with leaf NR activity
(r= +0-86) during the vegetative phase. The leaf NR activity was positively
correlated with the total reduced N per plant (r = +0-86) but not with leaf
reduced nitrogen (r = +• 0'19). There was also significant correlation between
root growth and the total reduced N per plant (r = + 0'87) and NR activity
(r = + 0-65). These data in4ic^ tfeat total reduced nitrogen per plant

Leaf proteinase and nitrate reductase activities 439

influenced by NR activity as well as by the root growth. Therefore the higher NR
activity and root growth in A. hypochondriacus and A. edulis than those in
A. caudatus and A. cruentus resulted in more total reduced nitrogen per plant
in the former two species.

During the flowering stage also no positive correlations could be traced between
NR activity and leaf reduced nitrogen or (r = + 0*16) or total reduced nitrogen
per plant (r = +0*42) (table 2). A. caudatus had high N levels in leaf and in
total plant during flowering stage, but its NR activity was less than that of the
other three species. Reed and Hageman (1980) observed that accumulation of
reduced nitrogen was dependent more on nitrate uptake than on the relative
levels of NR activity per plant. Raper et al (1977a, b) suggested that
nitrogen uptake was positively related to the rate of root growth in tobacco,
cotton and soybean. The remarkable increase in root growth of A. caudatus
during the flowering stage might have resulted in its comparatively high reduced
nitrogen per plant (table 2).

A. caudatus retained more reduced nitrogen per plant during grain filling
stage. Again the NR activity was not related with either leaf reduced nitrogen
(r = +0-14) or total reduced nitrogen per plant (r = -f 0'34) in all the four
species (table 3). The high root growth of A. caudatus could have facilitated the
accumulation of more reduced nitrogen per plant either through mobilisation of
reduced N or by augmenting the uptake of nitrate and efflux. While there was
always a positive correlation between the root growth and total reduced nitrogen
per plant (r = -f 0*78), the NR activity was not correlated with the total reduced
nitrogen per plant even at the harvest (table 4). Thus, our observations confirm
that at any stage of growth period in the grain amaranths studied, the leaf NR
activity alone is not an index of total reduced nitrogen per plant but the extent
of root growth along with the NR levels would together influence the nitrogen
content per plant.

The average nitrate reductase activity in the leaf was positively correlated with
the grain yield (r = +0-89) as well as total dry matter accumulation at harvest
(r = 0'86) in all the four species of grain amaranth (table 4). These findings
confirm the observations of earlier workers where NR activity was correlated with
grain yield (Blackwood and Hallam 1979 ; Deckard et al 1977 ; Bailing and
Loyn 1977 ; Johnson et al 1976).

On the other hand, the protein levels in the grain were not related with the NR
activities (r = — 0*37), but were strongly correlated with proteinase activity
(r = -f 0*85) in all the four species (table 4). This agrees with the results of
Deckard et al (1977) who concluded that the NR activities were not correlated
with grain protein. Differences in nitrogen translocation efficiency could reduce
the correlation between NR activity and grain protein (Croy and Hageman 1970 ;
Eilrich and Hageman 1973). Leaf proteinase activities were related more closely
to accumulation of grain nitrogen than leaf NR activity (Reed et al 1980). In
the present study, A. edulis contained greater proteolytic activity in the leaf and
accumulated more protein in its grain than that in A. hypochondriacus in spite of
high NR activity in the latter species.

Although the number of observations are limited, the present investigation
nevertheless suggests that in grain amaranths the level of leaf proteinase activity

446 K Ramamurthy Naidu, Y Seethambaram and V S Rama Das

is an important determinant of the grain protein content whereas the leaf NR
activity and root growth modulate the total reduced nitrogen per plant.

Acknowledgements

K Ramamurthy Naidu acknowledges the award of Teacher Fellowship from
the University Grants Commission, New Delhi, and Y Seethambaram acknowledges
AP Agricultural University, Hyderabad, for providing study leave facility.
The authors thank Dr A S Raghavendra for reading the manuscript.

References

Abrol Y P and Nair T V R 1973 In Proceedings of National Symposium on Nitrogen assimila-
tion and crop productivity Hissar, India ; (eds.) S P Sen, Y P Abrol and S K Sinna

(New Delhi: Assoc. Publishing Co.) Pp. 113
Blackwood G C and Hallam R 1979 Nitrate reductase activity in wheat (Triticum aestivum L.)

II. The correlation with yield ; New PhytoL 82 417-425
Bmnetti N and Hageman R H 1976 Comparison of in vivo and in vitro assays of nitrate

reduction in wheat (Triticum aestivum) seedlings ; PL Physiol. 5 583-587
Croy L I and Hageman R H 1970 Relationship of nitrate reductase activity to grain protein

production in wheat ; Crop. $ci. 10 280-286
Dalling M J, Halloran G M and Wilson J H 1975 The relationship between nitrate reductase

activity and grain nitrogen productivity in wheat ; Aust. J. Agri. Res. 26 1-10
Dalling M J, Balland G and Wilson J H 1976 Relation between acid protejnase activity and

redistribution of nitrogen during grain development in wheat; Aust. J. PL Physiol. 3 721-

730
Dalling M J and Loyn R H 1977 Levels of activity of nitrate redutcase at the seedling stage as a

predictor of grain yield in wheat (Triticum aestivum L.) ; Aust. J. Agri. Res. 28 1-4
Deckard E L, Lambert R J and Hageman R H 1973 Nitrate reductase activity in corn leaves

as related to >ields of grain and grain protein : Crop. Sci. 13 343-350
Deckard E L, Lucken K A, Joppa L R and Hammond J J 1977 Nitrate reductase activity,

nitrogen distribution, grain yield and giain protein cf Tall and Semidwarf. Near-isogenic

lines of Triticum aestivum and T. turgidum ; Crop. Sci. 17 293-296

Eilrich G L and Hageman R H 1973 Nitrate reductase activity and its relationship to accumu-
lation of vegetative and grain nitrogen in wheat (Triticum aestivum L.) ; Crop. Sci. 13

59-66
Fakorede M A B and Mock J J 1978 Nitrate reductase activity and grain yield of maize

culiivar hybrids; Crop. Sci. 18680-682
Hageman R H and Hucklesby D P 1971 Nitrate icductase. In Methods in enzymology (ed.)

A San Pietro, Vol. XXIII, Part A (New York: Academic Press) pp. 491-503.
Hageman R H 1979 Integration of nitrogen assimilation in relation to yield. ID Nitrogen

assimilation of plants (eds) E J Hewitt, and C V Cutting (New York: Academic Press)

pp. 591-611
Jhonson C B, Wittingtan W T and Blackwood G C 1976 Nitrate reductase as a possible

predictive test of crop yield ; Nature (London) 262 132-134
Peoples M B and Dalling M J 1978 Degradation of ribulose 1, 5-aiphosphate carboxylase by

proteolytic enzymes from crude extracts of wheat leaves ; Planta 138 153-160
Perez C M, Cagampang G B, Esmana B V, Monserrate R U and Julliano B O 1973 Protein

metabolism in leaves and developing grains of rices differing in grain protein content ;

PL Physiol. 51 537-542
Raper CD, Patterson D T, Parsons L R and Kramer P J 1977a Relative growth in nutrient

accumulation rates for tobacco ; Plant Soil. 46 743-787

Leaf proteinase and nitrate reductase activities 441

Raper C D, Patterson D T, Parsons L R and Kramer P J I977b Relationship between growth
and nitrogen accumulation for vegetative cotton and soybean plants ; Bet. Gaz. 138
129-149

Reed A J and Hageman R H 1980 Relationship between nitrate uptake, flux and reduction
and the accumulation of reduced nitrogen in maize (Zea mays L.). II. Effect of nutrient
nitrate concentration; PL Physiol. pp. 1184-1189

Reed A J, Below F E and Hageman R H 1980 Grain protein accumulation and the relation-
ship between leaf nitrate reductase and protease activities during grain development of
maize (Zea mays L.) I. Variation between genotypes ; PL Physiol. 66 164-170

Senfit P J 1980 Protein quality of amaranth grain. Proc. 2nd Amaranth Conference ; Rod ale
Press Inc.

Spices J R 1957 Colorimetric procedures for amino acids ; Methods in EnzymoL 3 467-477

Proc. Indian Acatf. Sci. (Plant Sci.), Vol. 91 , Number 5, October 1982, pp. 443-447,
© Printed in India.

Cell division in Staumstrum gradle Ralfs. under the scanning
electron microscope

VIDYAVATI

Botany Department, Kakatiya University, Warangal 506 009, India

MS received 3 December 1981 ; revised 14 August 1982

Abstract. Staurastmm gradle Ralfs. was grown in Chu's No. 10 culture medium,
in a culture cabinet at 18-20° C with 16 hrs light and 8 hrs dark period. The
cells exhibited polymorphism. The cells were fixed and their division and growth
was examined under the scanning electron microscope.

Keywords. Desmids ; Staumstrum gracile Ralfs.

1, Introduction

The process of cell division is unique in desmid biology, with special reference
to placoderms, and differs from the other algal groups. Some of the problems
it solved, may help in providing a better understanding of some of the principles
of morphogenesis and the control of the shape of cells in general. With each
division of these, often elaborately-shaped desmids two daughter semicells are
produced, which then acquire the typical complex and symmetrical shape of the
parent semicells.

Having studied the morphological features, surface ornamentation and
polymorphism under the scanning electron microscope (Vidyavati 1981), it was
thought desirable to study the division of the cells also.

Previous work on cell division, under TEM and SEM, were mainly contributed
by Dodge (1963), Drawert and Kalden (1967), Drawert and Mix (1961), Pickett-
Heaps and Fowke (1969, 1970), Pickett-Heaps (1973, 1974, 1975), Schiille (1975)
and Brook (1981).

2. Material and methods

Staurastmm gracile Ralfs. 679/3 was obtained from the culture collection of
Algae and Protozoa, Cambridge, U K. The work was carried out at the Botany
Department, Royal Holloway College, University of London, U K.

From the cultures thus obtained, unialgal isolations were made following
Pringsheim's method and these cultures were maintained in Chu's (1942) 10
medium, at 1 8-20° C temperature, subjected to alternate light and dark conditions
for 16 and 8 hrs, respectively,

443
P.(B)-4Q

444 Vidyavati

Fixation from healthy cultures in exponential growth were made at hourly
intervals in order to study the cells at various stages of division and follow the
change in shape of the new semicells. The cells were fixed in 1% glutaraldehyde,
made up in the culture medium (Chu's 10) for about 1 hr at room temperature,
after washing in culture medium, they were then post-fixed for about 1 hr in 1%
Osmium tetroxide also made up in the culture medium. They were then washed
3 times in culture medium. The cells were dehydrated in acetone of 30%, 50%,
70%, 90% and 100 %. Fixation, washing and dehydration were all carried out
in the centrifuge tubes and each time the cells were centrifuged discarding the
supernatant. The cells were then passed through critical point drying procedure.
The dried specimens were moved from the CPD apparatus and were mounted on
specimen stubs, using transfer on double-sided sticky tape. These were then
coated quite heavily with carbon and gold. Specimens were examined at 15 KV
in a jeel-JSM-25 S scanning electron microscope.

3. Observations and results

Staurastrum gracile Ralfs. is known for its polymorphic -form, cells are variable,
medium in size (length 27-60 ji ; breadth, including processes, 44-1 10 /* ; breadth
of isthmus 5'5-13/f), constriction slight, usually an acute notch; .semicells
variable, upper angles produced to from long, slender processes of variable lengtl
each with ,3 or 4 minute spines and provided with denticulations. The vertica
view is usually triangular, sometimes quadrangular, angles are produced to font
long processes, chloroplasts are axile with a central pyrenoid in each semiceli
(West and West 1923).

For many placoderm desmids cell division seems to be the only means of repro
duction. Sexual reproduction is rarely observed in nature or under laboratory
conditions. During division, the cell symmetry is completely destroyed by 2
wall that grows around the narrow isthmus joining semicells. During the process
of division, the cell enlarges at the isthmus region, and elongates, as a result the
semicells are pushed farther apart. The median septum then forms and the wall!
push out to produce the new semicells. As the semiceli enlarges lobe formatioi
proceeds and finally the arms of the typical species will be formed by further wal
elongation.

When the primary wall is almost fully expanded, the outer secondary wall begin*
to be laid down with its pattern of ornamentation matching that already esta-
blished, in the primary wall. The secondary wall also acquires its system o:
mucilage pores, the position of which are indicated very early in wall," deposition
which penetrate the entire secondary wall.

The daughter cells remain joined to one another, with their apices, until thi
shedding of the primary wall. These newly formed daughter cells move apart
probably due to the extrusion of mucilage.

It was found that cell division always occurred at a definite time after the beginning
of the light period, when the light and dark periods were alternated regularly
This suggests that the onset of illumination triggers the events, which set eel
division in motion. Schiille (1975) reported that the total period of development

Cell division in Staurastrum gracile Ralfs.

445

Figure 1. Staurastrum gracile Ralfs. showing triradiate form ( X 980).

Figures 2-13. Staurastmjn gracile Ralfs. cell division. 2. Isthmus regie r
becoming elongated. 3 and 4. Semi cells becoming separated. 5, 6 and 7. The
young serai cell showing bulged and lobed condition. 8. The lobes elongating

9. The development of the typical ornamentation and shape of the semi cell

10. Mature cell, in side-view. 11 and 12. Development of the semicell in i
triradiate form. 13. Mature cell viewed from above. [(2, 3, 11 end 13 (x 840)
4-10 and 12 ( x 1120)]

Cell division in Staurastrwn gracile Ralfs. 447

!" newly-formed semicells was from 2-3 hrs. Their development was complete
"ter this interval of time, but the actual separation of the newly formed cells usually
>ok another 3 hrs. Under these conditions, cells divide only once every 24 hrs.
Scannnig electron microphotographs were taken at various stages of division,
igure 1 illustrates cells in a population, mostly, in a triradiate form. Figures 2,
and 4 show enlargement of the isthmus region. Figure 5 shows one smooth
>ung semicell. Figures 6, 7 and 8 show young semicells bulged and lobe forma-
on. Figure 9 shows the development of the typical ornamentation and shape
F the semicell. Figure 10 shows mature cell inside-view. Figures 11 and 12
iow development of the semicell in a triradiate form and figure 13 shows mature
>11 viewed from above. Thus figures 2-13 illustrate the various stages in the
;11 division of the species, under investigation.

cknowledgements

he author expresses her deep sense of gratitude and thankfulness to Prof, Jafar
[izam, Vice-Chancellor, Kakatiya University, for his constant encouragement,
hanks are also due to Prof. John D Dodge, Head, Department of Botany, Royal
[olloway College, London, for the guidance, where the present investigation was
irried out. The author is also thankful to the UGC and the British Council for
le award of a Commonwealth academic staff fellowship.

efereuces

rook and Alan J 1981 The Biology of Desmids, Botanical Monograph, Vol. 16, Blackwell

Scientific Publications
hu S P 1942 The influence of the mineral composition of the medium on the growth of

planktonic algae. I. Methods and culture media ; /. EcoL 30 284-325
odge J D 1963 The nucleus and nuclear division in the Dinophyceae ; Arch, Protistenk 106

442-452
irawert H and Kalden G 1967 Licht und elektron mikroskopische Untersuchungen an Desmidia-

ceen. XTK. Mitteilung : Der Gestaltwandel des Nucleolus in interphasekero von Micras-

terias rotata Mitt. ; Staatsinst. Allg. Bot. Hamburgl2 21-27
irawert H and Mix M 1961 Licht und elektron mikroskopische Untersuchungen an Desmidia-

ceen. HI. Der Nucleolus in Interphasekern von Micrasterias rotata Flora', 150 185-190
ickett-Heaps J D and Fowke L C 1969 Cell division in Oedogomum. I. Mitosis, cytokinesis

and cell elongation ; Aust. J. Biol Sci. 22 857-894
ickett-Heaps J D and Fowke L C 1970 Mitosis, cytokinesis and cell elongation in the

desmid Closterium littorale ; /. PhycoL 6 189-215

ickett-Heaps J D 1973 Cell division in Cosmarium— Botrytis ; /. PhycoL 8 343-360
ickett-Heaps J D 1974 Scanning electron microscopy of some cultured desmids ; Trans. Am.

Microsc. Soc. 93 1-23

ickett-Heaps J D 1975 The green algae, (Sunderland, Mass Sinaver Assoc.)
:hiille H M 1975 Untersuchungen zum Synchronen zellwachstum von Stauratrum pingue Telling

(Desmidiaceae) in Licht-Dunkel-Wechsel ; Heih. Nova Hedwigia 42 275-281
idyavati 1981 Polymorphism in Staurastrwn gracile Ralfs. under Scanning election microscope

Paper read at the IV Botanical Conference, from 28-30th December, 1981 at Calicut,
test W and West G S 1923 A Mouograph of the British Desmidiaceae Vol. V, 96-97

. Indian Acad. Sci. (Plant Sci.)» Vol. 91, Number 5, October 1982, pp. 449-461
g) Printed in India.

Leaf surface studies of some medicinal salvias*

H P SHARMA and US HA SHOME

Pharmacagnosy Laboratory, National Botanical Research Institute, Lucknow 226001,

India

MS received 18 May 1982 ; revised 18 August 1982

Abstract. Scanning election microscopic studies on the leaves of 8 medicinal salvias
comprising mainly surface ornamentation of the various epidermal cells and the
appendages, provide useful parameters to distinguish one species from another.
Some of the distinguishing features of the species studied arc : Sohia cabidica
Benth. — Striated lower epidermis, stomatal ledges "broad and smooth ; 5. Icnata
Roxb. — Ab axial side completely covered over by a thick coat of trichomes ;
S. macrosiphon Boiss. — verrucose trichomes with constricted joints ; S- moorcroftiana
Wall-— longitudinal folds an basal cells of trichomes ; S. officlnalis Linn.— curved
cylindrical trichomos, cells over veins with characteristic longitudinal ridges, gland
stalk very long ; S. plebeia R. Br. — basal cells of trichomes transversely striated ;
S. pratensis Linn.— verrucose trichomes and series of irregular folds on lower
epidermis ; S. spinosa Linn. — smooth collapsible hairs, folds on general surface
similar to S. pratensis.

Keywords. Salvia ; Lamiaceae ; leaves ; SEM studies.

.. Introduction

{ large number of species of the Lamiaceae are presently in medicinal use, parti-
ularly in the Indian Systems of Medicine mainly for their essential oils,. However,
ike most crude drugs these are subject to substitution and adulteration. Deter-
aination of reliable criteria for distinguishing the genuine drugs is, therefore,
;reatly important.

Very often the marketed drugs consist of small broken pieces of different
»rgans which are, therefore, difficult to identify. Ultra morphology of the surfaces
>f diffferent plant organs offers a useful, simple and reliable procedure for
Authentication and standardization of herbal drugs where often only minute
urfaces are available for study (Cappelletti and Casadoro 1977).

Salvia (commonly known as Sage in European countries) is one of the most
mportant genera in this respect, of which 9 species are medicinal (Chopra 'et a!

NBRI Research Publication No. 148 (New series).

449

450 H P Sharma and Usha Shoirtd

1956). The genus, comprising of ornamental herbs and shrubs, is distribute
mostly in the temperate regions. Some 24 species of Salvia are reported fror
the Indian subcontinent (Anonymous 1972).

2. Materials and methods

The present study deals with scanning electron microscopy of the leaf surface
of 8 Salvia species, viz., S. cabulica Benth. (Afganistan-Kunar Prov.
S. lanata Roxb. (Himachal Pradesh-Panjain), S. macrosiphon Boiss. (Afganistan
Kandhar), S. moorcroftiana Wall. (Afganistan-Maidan), S. officinalis Lini
(W. Germany-Garmisch), S. pkbeia R. Br. (Afganistan-Kunar Prov.), S. pratens
Linn. (West Germany-Bavaria), and S. spinosa Linn. (Iraq-Basra) collected froj
the Herbarium of the Institute for Systematic Botany and Botanical Garden
University of Munich, West Germany. Material of S. lanata Roxb. froj
Herbarium, NBRT, was also used for confirmation.
The dried leaves were first soaked in hot water and after thorough washing

1 cm square strips were cut from the middle portions of the lamina midws
between the midrib and the margin and dehydrated through ethyl alcohol serif
followed by critical point drying procedure using liquid CO^. This was part
cularly necessary as the oil glands presented a distorted picture on simple dry in

2 mm square pieces were cut from the dried material and one piece each of tl
adaxial and abaxial surfaces were mounted on to the specimen stubs usir
double sided adhesive tapes.

Gold coating of the specimens, about 200 A thick, was carried out in an ic
sputter coater (JFC-1100). The specimens were examined under a JEOL-JSM-35
scanning electron microscope at an accelerating voltage of 10 kV and tilt of 3<
incident to the electron beam at an aperture 100 pm. The image was observed
magnifications ranging from x 200- x 5000 and photographs recorded on ORVS
120 films.

3. Observations

3.1- Salvia cabulica Benth. (figures 1-3)

3. la. Adaxial surface ; Sparsely hairy, trichomes small, 1-2 celled, extremely thi
walled having narrow lumina, basal cells bulbous showing a distinct girdle
disc at the base and at the joint ; upper cell swollen just above the joint, curv
and pointed, more warty than the basal one, warts mammilate, irregularly arrang
(figure 1). Epidermal cells polygonal, mostly straight walled or smoothly curve
without epicuticular extrusions. Stomata few, mostly diacytic or with four su
sidiary cells and flush with the surface. Guard cells longitudinally striat<
parallel to the pore (figure 1). Glandular hairs very few.

3.1i. Abaxial surface : Hairs more profuse, up to 4-celled, otherwise simil
to the upper ones, stomata also profuse, raised above the surface, lips ridg

Leaf surface studies of some medicinal sahias

451

a stoma on the

2. Lower epKiernus skowmg d«taot

striations art of a
the right (x 600).

epicuticular folds

cell (x

Lcaj swjace miaies oj some meaicmai saivias 4o3

ipward forming a rim round the pore. Epidermal cells wavy in outline and
urface rough and profusely striated; striations on each cell sparse except for
lose over the veins (figures 2 and 3). Glandular hairs frequent, all of the same
fpe, thin walled, heads very large (figure 2).

.2. Salvia Janata Roxb. (figures 4-7)

.20. Aclaxlal surface : Thickly hairy, hairs of three types (besides the glandular
airs) : (i) in some of the 2-celled hairs the basal cells taper upward and appear
istinctly cuticularised with broad epicuticular folds. The apical cell is also
lickly cuticularised, has short longitudinal wavy wrinkles and pointed curved
r beaked tips (figure 4). (ii) Short, 2-3 celled, basal cells broad at the base
nd flattened at the top with edges forming a broad rim. From the centre of
lis arises the next upper cell which is distinctly narrower and gradually tapers
pward (figure 5). (iii) The other types are very long, thin, flattened, woolly and
ollapsible forming a thick mat which has to be removed to permit a glimpse
f the epidermal characters (figure 4).

The epidermal cells are wavy in outline and have raised cuticular rims all along
le periphery. The central part of the cell surface is again convex upwards*
'urther, parallel cuticular striations are also found on the epidermal surface
igure 6). The glands are stalked as usual and the heads are very small, one-
slled and capitate.

The lower epidermis is completely hidden under the woolly hairs (figure 7).

. 3 Salvia macrosiphon Boiss. (figures 8-9)

.30. Adaxial surface : Hairs upto 6- celled, long, thin-walled, cylindrical but
ollapsible on drying, closely tuberculate, tubercles laterally flattened, arranged
i continuous longitudinal ridges (figure 8), ground cell of the hair swollen and
used above the epidermal surface, basal cell shorter and broader than the
ibsequent one. Joints constricted with annulus type thickening on either side,
pidermal cells smaller than those of other species, outline wavy; intercellular
artitions depressed but the general surface raised and striated; striations longi-
idinal and discontinuous. Stomata on both surfaces equally profuse, distinctly
lised above in a broad dome-like manner. Stomatal ledges wavy but compara-
vely narrow (figure 8). Glandular hairs infrequent,

.36. Abaxlal surface : Hairs similar to those of the upper surface. Cell out-
nes marked by longitudinal folds; surface thrown into fine wrinkles and folds
11 over. Stomata on the lower side similar to those of the upper surface except
lat the subsidiary cells have broad longitudinal folds along their outer periphery
igure 9). Glandular hairs of two types, 1 and 8 celled.

A Salvia moorcroftiana Wall (figures 10-11)

Aa. Adaxial surface : Hairs sparse, verrucose, 2-6 celled, comparatively thinner;
parts minute, sparse, arranged on longitudinally flattened ridges ; surface wrinkled

454 H P Sharma and Usha Shame

with loose cuticular folds ; joints -constricted with upper and lower cells swollen
just above and just below the constriction respectively; tips narrow and pointed,
sometimes . curved (figure 10). Long, thin-walled collapsible type hairs also
present. Anticlinal walls of epidermal cells sinuous,, but sinuosities obscured by
irregular loose cuticular folds all over surface of cells (figure 10) ; stoniata scanty.
Glandular hairs scarce.

3.46. Abaxial surface : Hairs similar to the upper ones. Long thin collap-
sible hairs also present. Cell margins appear ridged: stomata almost similar to
5. plebeia but distinctly smaller and flush with the surface (figures 10 and 15);
ledges narrow, margins of the ledges wavy (figure 11). Glandular hairs are
of two types: (i) small, one-celled with one-celled stalk and (ii) larger 8-celled,
sessile.

3.5 Salvia officinalis Linn, (figures 12 and 13)

3.5a. Adaxial surface : Hairy, hairs 2- to several-celled, thin and cylindrical,
apex obtuse, usually psilate; basal cell bulbous at the lower end and psilate with
very faint transverse markings, upper ones with longitudinal ridges ; joints slightly
swollen. Upper cells of some hairs over midrib sometimes having a few sparsely
arranged warts. Two-celled glandular hairs having long stalks present. Anticlinal
walls of epidermal cells sinuate, sometimes sharply so; intercellular partitions
depressed; cell surface with broad depressions and epicuticular folds (figure 12).
Cells over the midrib elongated having characteristic prominent longitudinal ridges
involving even the stalk cells of the secretory hairs. Stomata diacytic, cuticular
ridges of subsidiary cells converging on stomata, outer rim of guard cells raised
but their central portion depressed ; inner ledges wavy ; surface with a series
of wavy cuticular folds parallel to the opening. Transverse cuticular folds also
prominent (figure 12).

3 . 5b. Abaxial surface : Densely hairy, hair ; similar to those of the upper epidermis,
those over the veins turned backward; characteristic minute, club-shaped hairs
also present in this region. Their basal cells are depressed laterally and the
upper cells are broader, blunt and flattened (figure 13). Epidermal cells over
the veins elongated and very deeply furrowed longitudinally (figure 13). Large
8 celled glandular hairs with small stalks present.

3.6. Salvia plebeia R.Br. (figures 14-15)

3.60. Adaxial surface : Hairs sparse, evenly distributed, stout, thick- walled and
verrucose, basal cell broad at the distal end and tapering upward; lower cells
polygonal; the apical one cylindrical, very narrow and elongated with pointed
tip ; joints greatly constricted ; tubercles sparse on the two lower cells but much
denser on the apical one. Horizontal striations on the basal cell of trichomes
noticeable (figure 14 A, B). Stomatal guard cells raised upward, ledges quite
prominent . and sinuous. Epidermal cells also striated; striations longitudinal.
Glandular hairs occur .on both surfaces, heads usually small, 1- and 4-celled; stalk
also 2-celled.

Leaf surface studies of some medicinal salvias

455

Figures 5-8. 5. Salvia to/w/o— Trichomes from the upper epidermis with a broad
rounded basal cell and much narrower upper'cell (X 600). 6. Upper epidermis
showing epicuticular folds and glandular hairs. Cell outlines indistinct (x 1000),
7. Lower epidermis covered with a thick mat of collapsible multicellular trichomei
(x 600). 8. Salvia macrosiphon— Upper epidermis showing cells with distinct
cell outlines, verrucose trichomes showing a constricted joint with annular type
thickening and stomata with wavy ledges (x 600).

456

H P Sharma and Usha Shome

Figures 9-12. 9. Salvia macrosiphon—LowQr epidermal cells showing broad
longitudinal folds and stomata with wavy ledges (x 2000). 10. Salvia moorcrof liana—
Upper epidermis showing wrinkled surface with loose epicuticular folds, trichome
with longitudinal folds on the basal cell and a glandular hair on the left (x 600).
11. Lower epidermal cells showing a stomawith wavy ledges (x 2000). 12. Salvia
officinalis— -Upper epidermis showing trichomes and a stoma (x 1000).

Leaf surface studies of some medicinal sahias

457

JfV

Figures 13-16. 13. Salvia offidnalis—Lwei epidermis showing cylindrical
trichomes over the vein cells having characteristic epiculicular ridges (x 260).
14 A Salvia plebia— Upper epidermis showing a trichome with its basal cell
having fine transverse stations (x 400). 14 B. A portion of trichome fiom
figure Ha enlarged to show transverse striations (x 960). 15. Lower epidermis
showing collapsible trichomes and astoma with wavy ledges (x 2000). 16. Salvia
pratensis—Uppu epidermis with distinct cell outlines, small glandular hairs,
stornata and a trichome (x 600),

458

H P Sharma and Usha Shome

Figures 17-20. 17. Salvia pratemis- -Lower surface showing wrinkled epidermis,
a stoma and glandular hair (x 2000). 18. Salvia spinosa— Upper surface of the
leaf showing wrinkles all over (x 660). 19. Lower epidermis showing a large
number of trichomes (x 600). 20. Lower epidermis showing stoma with fungal
mycelium coming out of one of the stomatal pores (x 860).

Leaf surface studies of some madicinal salvias

459

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Leaf surface studies of some medicinal saivias 46i

3 . 66. Abaxial surface : Beside the stout hairs, thin several-celled collapsible hairs
also present, particularly over the veins. Epidermal cells wavy and thick walled.
Stomata raised and characterised by cuticular ledges which are wavy and over-
hang the stoma on either side. Beside these the cuticle over the guard cells thrown
into two sets of folds, one set wavy and arranged more or less parallel to the
stomatal opening while the other set is represented by a fine series of wrinkles
at right angles to the long axis of the opening (figure 15). The cuticle of sub-
sidiary cells is also thrown into a series of folds perpendicular to and conver-
ging towards the guard cells.

3.7. Salvia pratensis Linn, (figures 16 and 17)

3.7#. Adaxial surface : Hairs on the lamina very sparse, short, cylindrical
thick-walled and sharply pointed at the tip, 1-3-celled ; basal cell very broad at the
lower end forming a disc ; upper one swollen just above the joint, verrucose, warts
denser on the upper cell and arranged in vertical rows (figure 16). Hairs on
midrib and petiole very long, more dense, thin walled and collapsible in dried
material. Ordinary epidermal cells sinuate and striated, striations very fine; anti-
clinal partitions depressed and forming a groove with edges of the adjacent cells
raised (figure 16). Stomata slightly raised, lips with raised longitudinal ledges.
Glandular hairs very few.

3.76. Abaxial surface : Hairs of two types : (i) longer collapsible type over
the midrib and (ii) thick cylindrical types elsewhere. The latter 1-4 celled, but
otherwise similar to those on upper surface, faint horizontal wrinkles present on
the hairs. Stomata diacytic, slightly raised, ledges on stomatal lips quite
prominent (figure 17). Epicuticular ornamentation in the form of irregular
wrinkles present particularly along the edges of the guard cells (figure 17).
Glandular hairs frequent, stalk 1-3 celled, gradually expanding upward, head
globular and composed of eight cells.

3.8. Salvia spinosa Linn, (figures 18-20)

3.8fl. Adaxial surface: General surface undulating and greatly wrinkled all over,
wrinkles short, irregular and wavy (figure 18). Peripheral margins of epidermal
cells raised, stomata diacytic, similar on both surfaces, outer and inner margins
of the guard cells raised. Outer margin broadly ridged, central part concave
and wrinkled, one set at right angles to the stomatal pore and the other dis-
continuous but more or less parallel to it (figure 19). Stomatal ledges prominent
(figures 18 and 19). Glandular hairs with large, smooth, 8-celled heads.

3. 86. Abaxial surface : Surface similar to the upper one but hidden under a thick
mat of multicellular hair. Hairs long, thin, smooth with faint longitudinal
cuticular markings and a few transverse wrinkles (figure 20); joints constricted.
Stomata similar to those on the upper surface but smaller and more dense.
A large number of fungal mycelia observed entering through storaatal pores
(figure 20).

46£ H P Shanna and Usha Shome

4. Discttssiqii

Comparative morphology of epidermal surfaces using SEM has proved valuable
in elucidating taxonomic problems (Atwood and Willams 1979; Ayensu 1974;
Cutler and Brandliam 1977; Dehgan 1980; Ehler 1974; Rollins and Banerjee,
1975).

Cappelletti and Casadoro (1977) distinguished Atropa bella-donna (family
Solanaceae) from its adulterants— Ailanthus altissima (Miller) Swingle (family — •
Simurubaceae) and Phytolacca amerlcana L. (family — Phytolaccaceae)— with the
help of ultra morphology of leaf surfaces.

The present study was undertaken with a view to determine such parameters
as could help characterise the different medicinal Sahia spp. even if the surfaces
available are too small to be of much diagnostic value otherwise.

The study has shown that ultra morphology of the leaf surface provides a useful
tool in authentication of vegetable drugs, at least in the case of salvias. The
epidermal surface on both sides have epicuticular ornamentation, in the form of
striations or loose folds in all species except for the adaxial surface in S. cabulica
and the abaxial one in S. qfficinalis. Of the remaining, S. plebeia has striations
on both surfaces while loose folds occur on both sides in S. moorcroftiana and

5. spinosa. In all others the two surfaces are dissimilar. Similarly, there are
differences in the stomatal characters. Another important feature which helps
to characterise the different species studied is the type of trichomes and surface
ornamentation (table 1). For instance, whereas only a single type of hair
(besides the glandular hair) were observed in three species, S. cabulica,
S. macrosiphon and 5. qfficinalis, all others have long, psilate, thin-walled collap-
sible type of hair as well- In case of S. lanata even the short hairs are of
two types. Again, except S. cabulica and S. spinosa all others have verrucose
to tuberculate ornamentation all over the trichome surface. However, of the
latter ones S. moorcroftiana has tubercles on the uppermost cell and S. officinalis
on the middle one.

Scanning electron microscopic studies have, thus., proved to be extremely
useful in characterisation of the different species of the same genus Salvia and
a positive advantage in standardization of herbal drugs.

Acknowledgements

The authors express their thanks to Dr T N Khoshoo, Director, National Botani-
cal Research Institute, for encouragement and keen interest in the progress of
the work, to Mr S L Kapoor, Incharge, Herbarium Section, NBRI, for sparing
the herbarium sheet of S. lanata Roxb. and to Dr M Yunus, for taking the
SEM photographs.

The senior author is particularly indebted to Prof- H Merxmuller, Director,
Institute for Systematic Botany, University of .Munich, for the excellent labora-
tory and herbarium facilities made available to him during his visit to the said
Institute and for permission to use herbarium material of the Staatsherbarium

Leaf surface studies of some medicinal salvias 46 3

for the present studies. He is also thankful to Prof. J Grau of the same
Institute for familiarising him with some techniques of Scanning electron micro-
scopy and to DAAD and the CSIR for arranging his visit to West Germany under
the exchange of Scientists Programme.

References

Anonymous 1972 The wealth of India. A dictionary of Indian raw materials and industrial

products. IX, Rh-So (New Delhi : CSIR)
Atwood J T and Williams N H 1979 Surface features of the adaxial epidermis in the con-

duplicatc leaved Cypripcdioideae (Orchidaceae) ; Bot. J. Linnean Soc. 78 141-156
*Ayejv>u E E 1974 Leaf anatomy and Systematic^ of New World Vellaziaceae. In Smithsonian

contributions 1o botany (15 City of Washington : Smithsonian Institution Press).
Cappelletti E M and Casadoro G 1977 Leaf surface morphology of Atropa hello donna and of

some adulterant species by Scanning electron microscopy ; Planta Medica 31 356-366
Chopra R N, Nayar S L and Chopra I C 1956 Glossary of Indian medicinal plants (New

Delhi : CSIR)
Cutler D F and Brandham P E 1977 Experimental evidence for genetic control of leaf surface

characters in hybrid Aloineae (Liliaceae) ; Kew Bull. 32 23-32
Dehgan B 1980 Application of epideimal morphology to taxonomic delimitations in the genus

Jatropha L. (Euphorbiaceae) ; Bot. J. Linnean Soc. 80 257-278
*Ehler N 1974 Die Feinskulpluren Madagassischer Euphorbien-Hockblatter und ihre taxonomiche

vertigkeit ; Feddes Repertorium 85 345-351
Rollins R C and Banerjee U C 1975 Atlas of the Trichomes of Lesquerella (Cruciferae) :

The gussy Institute, (Harvard University) pp. 1-48

* Not seen in original.

Proc. Indian Acad. Sci. (Plant Sci.), Vol. 91, Number 5, October -1982, pp. 465-472.
C) Printed in India.

Morphological and histochemical changes in the egg and zygote of
Lagerstroemia speciosa. I. Cell size, vacuole and insoluble
polysaccharides*

P RAGHAVAN and V J PHILIP**

Department of Botany, University of Calicut 673 635, Kerala, India
** P.O. Box 43844, Nairobi, Kenya

MS received 1 February 1980 ; revised 16 December 1980

Abstract. In Lagerstroemia speciosa, the decrease in size of the egg and its
micropylar vacuole immediately after fertilization is followed by a progressive
and marked expansion of the cell. The PAS-positive cell wall material in egg is
confined to the micropylar half. Soon after fertilization, but before completion
of decrease in size of the zygote, its cell wall grows in thickness. A complete wall
is not formed around the zygote. The bulk of the insoluble polysaccharides in the
cytoplasm is legalized at the chalazal pole of the egg and zygote. Following ferti-
lization, the size and number of starch gianules in the egg cytoplasm significantly
increased followed by a decrease and again an increase during zygote develop-
ment. The morphological changes in the egg following fertilization are probably
related to the osmolarity of the cell and of the vacuole which would account for
the change in cell size.

Keywords. Lagerstroemia speciosa ; variation in vacuolar size ; cell wall ; insoluble
pclysaccharides.

1. Introduction

The zygote is the fundamental structural and functional unit which constitutes
a new beginning and affords opportunities for investigating growth, development,
differentiation, assumption of form and functional activities. Ultrastructura*
changes have been described in the fertilized egg of a few plants such as cotton
(Jensen 1968), Capsella bursa-pastoris (Schulz and Jensen 1968), Zeamays (Diboll
1968), Petunia hybrida (van Went 1970b), flax (D'Alascio-Deschamps 1972),
barley (Norstog 1972) and Quercus gambelii (Mogensen 1972 ; Singh and
Mogensen 1975). But little is known about the metabolic changes accompanying
the formation of the zygote. The present study deals with the structural and
certain histochemical changes that result on fertilization of the egg of Lagerstroemia
speciosa and its subsequent development up to the first division of the zygote.

This paper is dedicated to. late Professor B. G. L. Swamy.

* part of tfye thesis of PR approved by the University of Calicut for the Ph.D. degree.

465

466 P Raghavan and V J Philip

2. Material and methods

Ovules and seeds of Lagerstroemia specicsa (Linn.) Pars, at different stages in
development were collected at weekly interval from areas in and around Calicut
University campus, fixed instantaneously in the field in formaim-acetic-alcoho
or Carnoy's fluid. Conventional method of dehydration through tertiary butyl
alcohol series was employed and serial microtome sections (10-1 5 /on) were
prepared. Insoluble polysaccharides were demonstrated by pAS-reaction (Jensen
1962). Tissue oxidation was carried out in 0'5% periodic acid in distilled water
for 20-30 min. Response to the stain was the same by sections fixed by either
fixatives. Treatment of tissues fixed in Carney's fluid with 1% aqueous mercuric
chloride solution gave a negative reaction with Schiffs reagent. Confirmatory
test carried out with potassium iodide-iodine stain indicates that the materials
stained with PAS is constituted of starch grains and that these occur in plastids.
Cellular, nuclear and vacuolar areas were calculated from camera lucida drawings
of the stained preparations.

3. Results and discussion

3.L Change in size of the cell and its vacuole

The pear-shaped egg cell with its broad chalazal and tapering micropylar poles
ranges from 457 ^m to 632 /tm in length. It contains a thin layer of cytoplasm
surrounding a large vacuole which reaches almost the cell wall at the micropylar
pole. The cytoplasm is concentrated in the chalazai half. The nucleus is confined
to either the chalazal pole (figure 1) or to a lateral position in the chalazal half
(figure 2).

Immediately after fertilization the cell shrinks to about 240 ^m and the vacuolc
also decreases (figure 3). This decrease in size is followed by a progressive and
marked expansion of the cell (figures 4-8), so that the maximal size attained is
about five-fold the initial size of the [zygote and about two and a half-fold the
size of egg cell (figure 8). Furthermore, a progressive flattening of the micropylar
region of the zygote occurs. In few preparations the nucleus was seen displaced
towards the centre of the cell due to the appearance of two smaller vacuoles at the
chalazal pole (figure 3). Subsequently these two apical vacuoles expanded
disproportionately to the remainder of the cell and displaced the nucleus to a
more central position (figure 7). In 75% of the zygotes examined, only the
micropylar vacuole was present which enlarged retaining the nucleus at the chalazal
pole itself.

The observed size changes in the egg cell and zygote of Lagerstroemia cannot
be accounted for merely by biological variation among the preparations examined.
In cotton also a prominent shrinkage of the zygote occurs (Jensen 1964, |1968),
but not in Capsella (Schulz and Jensen 1968) and barley (Norstog 1972). Cell
enlargement and cell elongation are considered as general features of the zygote
preparing for division. Assuming that cell shrinkage is due to loss of water,
the process is reversible because water from the exterior can be abstracted by
the maturing zygote. The underlying osmotic changes in the zygote and the

Morphological and hlstochemical changes in zygote

467

30JU

Figures 1*8. Lagerstroemia speci&sa- changes in size of egg, zygote and their
vacuole. All are longi sections ; micropylar pole towards bottom of page. 1. Egg
cell ; the major part of cytoplasm is confined to chalazal pok . 2. Egg cell showing
lateral disposition of nucleus and cytoplasm at chalazal half. 3. Zygote ; note
decrease in cell volume, and formation of two smaller vacuoles at the chalazal
pole. 4-8. Zygote at later stages in development. Note the progressive increase
in size and the flattening of micropylar region.

surrounding milieu result from metabolic changes at these two sites. One of
these changes is the reversible soluble carbohydrate/polysaccharide transformation
in zygote to be discussed in § 3.3.

3.2. Cell wall

Both in the egg and in the zygote the extent of cell wall present is variable.
Electron microscopic studies have shown that in mature eggs of cotton (Jensen

468 P Raghavan and V J Philip

1964, 1965), Torenia fournieri (van der Pluijm 1964), maize (Diboll and Larson
.1966) and Petunia hybrida (van Went I970a, b) the cell wall extends to only half
way up the micropylar pole. However, in the egg cell of Capsella bursci-pastoris
the wall extends almost over the entire cell ; at the chalazal pole the structure is
honey-combed with large gaps (Schulz and Jensen 1968). Thus, in all the species
investigated the egg cell shows regions of the plasma membrane at the chalazal
pole in direct contact with the embryosac, a feature which possibly enables the
egg to derive nutrition directly from the central cell. In general, in angiosperms
the micropylar region of the zygote is anchored to the wall of the embryosac, and
during development wall formation extends over the open chalazal region and
envelops the zygote all round. In Capsella bursa-pastorls simultaneously with
the deposition of the wall material in the gaps in the chalazal region, the wall in
the micropylar portion of the zygote becomes thickened (Schulz and Jensen 1968).
In barley the wall of the zygote is thicker at the micropylar region than elsewhere
(Norstog 1972).

In Lagerstroemia speciosa a positive periodic acid-Schiff reaction was obtained
both in the cell periphery and in the cytoplasm of the developing embryo. But
in the egg the reaction was confined to the micropylar half, resembling in ' this
respect, cotton, maize, Torenia and Petunia. But, unlike in cotton, in Lager-
stroemia a complete cell wall is not formed around the young zygote. In the egg
the reaction product is visible as a fine film extending to 60% of the perimeter
of the cell from the micropylar pole (figure 9). Soon after fertilization, but
before completion of decrease in size of the zygote, the cell, wall grows further in
length covering 70% of the cell perimeter measuring 56/wn (figure 10). Subse-
quently, however, the addition of wall material does not appreciably increase the
thickness, and the percentage of wall material to the perimeter of zygote remains
unchanged (figure 11). In a nearly mature zygote the wall extends to 112/xm
from the micropylar pole covering 80% of the cell perimeter (figure 12), and its
thickness is slightly less than that in the two earlier stages but more than that in
the egg.

Street and Opik (1970) have pointed out that during cell expansion (elongation),
the cell wall does not thin out, but there 'occurs a proportionate increase in cell
wall synthesis. It is not clear how a cell would respond to a decrease in cell size.
Because in both the egg and the zygote of Lagerstroemia speciosa the cell wall is
present only at the micropylar half, a decrease in size of the young zygote will not
presumably impose as much strain as it would have been if the zygote had an
entire cell wall.

3.3. Insoluble polysaccharides (starch granules)

Histochemical staining with periodic acid-Schiff reagent showed, besides cell wall,
numerous granules of insoluble polysaccharides in the cytoplasm of both egg and
zygote, especially localized in their chalazal pole. In all stages of development
beginning from egg, the starch granules are heterogeneous in size and varied
in number. The egg cell contains about 50 tiny starch granules (figure 9).
Following fertilization, there is a significant increase in their size and number. In
the young zygote, which showed a decrease in size, the number of starch granules
increased to 90-100 (figure 10). As the zygote enlarged, but before attaining

Morphological and hlstochemlcal changes In zygote

469

Figures 9-12. Lagerstroemia speciosa—mstian longisections of egg and zygote after
treatment with periodic acid-Schiff reagent. 9. Egg with PAS-positive wall covering
60% of its perimeter from the micropylar pole. Note the size and number of
starch granules at the chalazal pole. 10. Zygote as seen immediately after forma-
tion ; note further growth of wall in thickness, decrease in cell size, and number
of starch granules. 11. Zygote at a later stage, showing decrease in size and
number of starch granules but with no appreciable change in wall mateiial deposi-
tion. 12. Nearly mature zygote with the wall covering SO % of the cell peri-
meter. X 850.

Morphological and liistochemical changes in zygote 471

naturity, the number of granules decreased to about 60 ; their size also decreased
figure 11). In the nearly mature zygote, a second increase in number to about
100 granules can be noticed (figure 12), accompanied by an increase in size.

In cotton the egg cell contains one or two small starch granules per plastid and
:here is no specific association of the plastids with other organelles in the cell.
Following fertilization, when cell size is decreasing, the plastids accumulate along
with mitochondria, around the nucleus (Jensen 1968). At this stage, starch begins
to accumulate in the plastids. The close association of the nucleus, plastids and
ribosomes presumably facilitates the ready elaboration of the biosynthetic system (s)
.ind the transfer of the enzyme concerned to the granule or the cytoplasm, as the
:ase may be. During maturation of the zygote, when additional wall material
is being formed so as to complete the wall around the entire cell, the number of
plastids remains unaltered, but their size increases. Also, the number of starch
grains per plastid increases so that the plastids become filled with starch.

The occurrence of starch grains in the egg cell of Lagerstroemia speciosa, the
increase in their number soon after fertilization and their presence in large amounts
in the mature zygote, are in accord with the findings in cotton (Jensen 1968), but
the decrease in number and size of starch granules which occurs in the intermediate
stage is distinctive of Lagerstroemia. The increase in number and/or size of the
starch granules following fertilization is -accompanied by the decrease in the
vaculoar volume. Similarly, a decrease in the number and size of starch granules
is sometimes associated with increased vacuolar size, as when the early zygote
reaches the intermediate stage of development. Such a formation of starch
granules at the expense of soluble sugars stored in the vacuole would reduce the
osmotic pressure and result in a diminution of cell and vacuolar size. This would
account for the morphological changes, namely, a marked decrease in size, on
fertilization of the egg. The reverse process, namely starch degradation and
transfer of soluble sugars to the vacuoles, would increase the osmolarity of the
cell and vacuole. This would account for the increase in size of the early zygote
and its vacuole. Contrary to expectation, the second increase in number and
size of starch granules, which occurs in the mature zygote is associated with an
actual increase both in cell and vacuolar size.

A significant point is that cell wall synthesis in the zygote occurs at the micro-
pylar half, even though the protoplasm is concentrated at the chalazal half.
Evidently, the polarity of the cell does not extend to the metabolic activity of the
cell relative to wall synthesis.

Acknowledgements

The authors are grateful to Prof. P S Krishnan, Emeritus Professor of Bio-
chemistry, University of Calicut, for help in the preparation of this paper. One
of us (PR) thanks the University of Calicut and the University Grants Commission
for financial assistance in the form of fellowships.

References

D'Alascio-Deschamps R 3972 Le sac cmbryoniiaire du lin apr^s la fecondation ; Bctaniste 50
273-288

P Raghavan and V J Philip

Diboll A G 1968 Fine structural development of the mega-gametophyte in 2ea mays following

fertilization ; Am. J. Bet. 55 7S7-806
Diboll A G and Larson D A 1966 An electron microscopic study of the mature mega-gamato-

phyte in Zea mays \ Am. J. Sot. 53 391-402

Jensen W A 1962 Botanical histo chemistry (San Francisco ; W H Freeman) pp. 198-199
Jensen W A 1964 Cell development during plant embryogenesis in Meristems and differentiation

Brookhaven Symp. BioL 16 179-202
Jensen W A 1965 The ultrastructure and composition of the egg and central cell of cotton ;

Am. J. Bat. 52 781-797

Jensen W A 1968 Cotton embryogenesis : the zygote ; Planta 19 346-366
Mogensen H L 1972 Fine structure and composition of the egg apparatus before and after

fertilization in Quercus gambeUl : the functional ovule ; Am. J. Bot. 59 931-941
Norstog K 1972 Early development of the barley embryo : fine structure ; Am. J. Boi. 59

123-132
Schulz S R and Jensen W A 1968 Capsella embryogenesis : the egg, zygote and young embryo ;

Am. /, Bot. 55 807-819
Singh A P and Mogensen H L 1975 Fine structure of the zygote atfd early embryo in Quercus

gambelii; Am. J. Bot. 62 105-115
Street H E and Opik H 1970 The physiology of flowering plants : Their growth and development

(London : Edward Arnold) p. 16o
van der Plmjm J E 1964 An electron microscopic investigation of the filiform apparatus in the

embryosac of Torenia fournieri in Pollen physiology and fertilization (ed.) H E Linskens

(Amsterdam : North-Holland PubL Co.) pp. 8-16
van Went J L 1970a The ultrastructure of the egg and central cell of Petunia ; Acta Bot. Neerl.

19 313-322
van Went J L 1970b The ultrastructure of the fertilized embryo sac of Petunia ; Acta Bot.

NeerL 19 468-480

Proc. Indian Acad. Sci. (Plant Sci.), Vol. 91, Number 6, December 1982, pp. 473-478,
© Printed in India.

The floral anatomy of Puya spathacea Mez. (Bromeliaceae) with
special reference to nectaries

R A KULKARNI and R M PAI

Plant Morphology Laboratory, Department of Botany, Marathwada University,
Aurangabad 431 004, India

MS received 25 May 1981

Abstract. The floral anatomy of Puya spathacea Mez. is dcsciibed in detail. The
outer floral whorls are united to develop a short hypanthium which is adnate
to the base of the ovary. The sepals are five-traced and the petals, three-traced.
The placentation is axile. The occurrence of numerous ovules in more than two
rows as well as the extension of the carpellary ventrals into the style are less
advanced features. The ovarian nectary is extensively developed and shows a
transition between typical septal and epigynous nectaries of certain monocotv*
ledonous taxa.

Keywords, Puya spathacea ; floral anatomy ; nectaries.

1. Introduction

The Bromeliaoeae are a fairly large family with about sixty genera and about
2000 species. Hutchinson (1959) considers the family to be a homogeneous taxon
representing the * climax of a line of descent wherein the calyx and corolla have
remained distinct or fairly distinct from each other '. He treats it as related to,
but more advanced than the Commelinales.

Smith's extensive studies (1934) point out that the family has strongest affinities
with the Rapateaceae and that both families probably arose from a common
ancestral stock. Within the family, Puya is treated as probably the source of
ancestral types from which the other sub-families developed. According to
Pittendrigh (1948), Puya is the most primitive of living bromeliad genera.

The genus Puya is a native of thei Andes with unique habit and habitat. Some
species of the genus are considered to be the largest and most interesting of the
bromeliads, e.g., Puya raimondii Harms, a rare monument of the Peruvian Andes
threatened with extinction. Plants of this species attain a height of 9*5 m and
bear thousands of flowers with sugar-poor honey and pollinated by birds. The
tiny, winged seeds may number even a billion per plant. The plants are mono-

473
P. (B)-l

474 ... R A Kulkami and R M Pal . .

carpic and hapaxanthic (blooming once in their life and dying out thereafter,
and propagated exclusively by seed). Puya spathacea Mez. is a much smaller
plant attaining a height of 1 m.

Studies in the floral anatomy and morphology of this interesting group of
plants are meagre. An extensive investigation on the vascular anatomy of the
flower of the group is, therefore, undertaken in this laboratory, and the results
on one of the most .unique genera are presented in this paper. Amongst the
noteworthy earlier contributions on the family is a paper by Budnowski (1922) who
is of the opinion that probably no Bromeliaceae lack septal glands. Some work
on the group is in progress at Professor Rauh's laboratory in West Germany.

2. Materials and methods

The fixed flowering material was received from Prof. H Merxmuller, West
Germany, collected from his University Botanic Gardens. The usual paraffin
method has been followed. Serial transections (10-12 p, in thickness) and longi-
sections were stained with crystal violet using erythrosin as a counter stain.

3. Observations

The pedicel contains a ring of six to eight prominent bundles surrounded by
many discrete smaller strands (figure 1). All these bundles divide and form nume-
rous strands which resolve into a large number of centrally placed placental
bundles and six outer groups of strands from which the principal bundles of the
floral whorls emerge out" (figure 2), From each of the posterior and antero-lateral
groups of strands three LS strands a*e derived. While the laterals amongst the
three branch and extend into the sepals on either side, the median one bifur-
cates and branches into the margins of both the sepals (figure 3). The remainder
of these groups resolve into an inner is bundle and an outei? strand which splits
into an MP bundle and two LP strands (figure 4). The three groups in the postero-
lateral and anterior positions resolve into the MS, os and D strands (figure 4)

The nectary is developed from the very base of the ovary beneath the
level of the loculi (figure 4). It is extensively developed with many canal-
like passage ways and surrounded by the repeatedly dividing placental bundles
(figures 3, 5). This gives the appearance of ' processes ' of axile tissue of the
ovary lined by glandular cells. Upwards, the nectary is closed in the centre to
result in. three glandular clefts and some of the * processes ' persist in the form
of lobes of- carpellary tissue enclosed in septal cavities or canals. These open to
the outside towards the middle of the length of the ovary (figure 6).

The ovary is teilpcular with the placentation axile (figure 6). Upwards, the
margins of the carpels meet only in the centre to continue syncarpy (figures 6, 7).
Some of the placental bundles arrange themselves opposite to the loculi and these
bear traces into the placentae and the ovules (figures 6, 7). The ovules are
numerous and borne in many rows on each placenta. Most of the placental
bundles end in bearing traces to the ovules and towards the nectary. Six placental

Floral anatomy of Puya spathacea Mez.

475

Figures 1-5. Puya spathacea, serial transections of the flower from the base
upwards ; D, carpellary dorsal ; is, inner staminal strand ; LP, lateral bundle of
petal ; LS, lateral bundle of sepal ; MP, median bundle of petal ; MS, median bundle
of sepal ; N, nectary ; P, petal ; PL, placental bundles ; os, outer staminal strand ;
s, sepal.

Figures 6-11. Puya spathacea, serial transections of the upper part of the flower ;
QL, gland ; PL, placental bundles ; sc, stylar canal ; sx, stamen ; STY, style.

476 R A Kulkarni and R M pai

bundles (caipellaty ventrals) continue upwards into the base of the style (figure 8).
The ovarian loculi continue in the form of three canals which merge into a
triradiate canal in the style (figure 9). The carpellary dorsals continue up to the
tip of the style and into the three shallow stigmatic lobes.

The outer floral whorls form a short hypanthium which is adnate to the base
of the ovary. The sepals, petals and stamens separate out simultaneously
(figure 6). The sepals are twisted to the left and the petals to the right (figure 7).
At the level of insertion, the sepals and the petals receive 8 or 9 traces (figure 5).
The median bundles of the sepals and the petals bear lateral branches, some of
which may divide further.

The six stamens are one-traced. The staminal bundle extends upwards into
the connective and ends at the base of the short crest of the anther (figure 10).
It bears a lateral branch towards either anther lobe (figure 9). The anther is
two-celled. The outer stamens are longer (figure 10).

4. Discussion

The vascular anatomy of the flower of Puya presents several, features of interest
and importance. The outer floral whorls are united into a short hypanthium which
is adnate to the ovary for some length. This is a trend in the development of
an inferior ovary which is characteristic of the allegedly advanced bromeliads,
e.g., Aechmea, Billbergia, Cryptanthus, Neoregelia. The adnation is, however,
not of a marked degree which is also reflected by the absence of fusion of the
principal strands of the floral whorls. The placentation is axile. The occurrence
of numerous ovules borne in many rows is a primitive feature. The extension
of the carpellary ventrals into the style is also a less specialised condition.

The development and the structural details of the nectary are very significant.
Initially, the nectary is in the form of a central crater with canal-like passageways
which proliferate profusely. The placenta! bundles repeatedly divide and are lodged
in what appear as * processes ' of carpellary tissue lined by glandular cells. The
placental bundles have to be associated with the nectary in its function (Agthe
1951 ; Budnowski 1922 ; Frei 1955 ; Pai and Tilak 1965 ; Tilak and Pai 1974).
Upwards, the nectary is closed in the centre and the * processes' of carpellary
tissue appear in the form of distinct glandular outgrowths lodged in septal cavi-
ties. These are vascularised by the placental bundles. The glandular lobes
open to the outside. In Costus and Tapeinochilus of the Costaceae a more or
less similar initial condition is observed which probably prompted Brown (1938)
to describe the nectaries in Costus as septal. Rao etal (1954) describe them as
not septal nectaries but as vascularised outgrowths of carpellary tissue \vMch
extend upwards in ovarian canals along the septal radii and open at the top of
the ovary. In Kaempferia rosea (Pai 1966) these glandular lobes appear on. the
septal radii in similar but epiovatian canals at about the level where those in
Costus end, and extend above the ovary. In the majority of zingibers, the
nectaries are epigynous.

It may be noted that in monocotyledonous taxa with extensively developed
septal nectaries with canal-like passageways, vascularised * processes ' of carpellary
tissue dp occur, e.g., Muw and Ensete (Tilak and Pai 19?4). However, these

Floral anatomy of Puya spathacea Mez. 477

do not develop into glandular outgrowths. This would seem to indicate that
these ' processes ' may persist and extend upwards as glandular lobes or outgrowths
in some taxa as in Puya, Costus and Tapelnochilus. While they are short in Puya
and open to the outside, they proliferate and extend upwards and above the
ovary in the latter two genera. The function of nectar secretion is taken over
by the lobes or outgrowths so that the canals in which they appear lodged do
not have the secretory lining layer. The lining layer in Puya is secretory in the
basal half and the function of secretion is taken over by the glandular outgrowths
upwards.

Anatomical evidence is also significant in this context. The glandular out-
growths in Puya, Costus.. Tapelnochilus and Kaempferia rosea 2 re vasculaiised by
the ptacental bundles. As the glands ate elevated to an epigynous position, the
placental bundles are replaced by the vascular tissue derived from an anastomosing
vascular plexus which is generally developed at the top of the ovary in the zingibers
(rf. Pai 1966).

The condition in Puya may, therefore, be considered as transitory ; rather the
origin of epigynous nectaries of most zingiberaceous taxa may have to be sought
from extensively developed septal glands.

The sepals receive basically five traces while the petals are three-traced. The
differential twisting of the two whorls of the perianth is characteristic of many
bromeliads.

The stamens are one-traced and the outer whorl of the stamens is longer than
the inner. This is a feature observed in many petaloid monocotyledonous taxa
(Kulkarni 1973 ; Markandeya 1978 ; Vaikos 1974 ; Vaikos etal 1978) and demon-
strates the trend in differentiation of the two androecial whorls and the ultimate
reduction of either whorl. In the bromeliads, this has not made much headway
as is revealed by a study of many genera of the family (Kulkarni, unpublished
data). There is a short crest for the anther which is not described in most
taxonomic accounts. However, it does not appear to be of pertinent phylogenetic
significance.

Acknowledgements

The authors are grateful to Dr Merxmuller for his generous help with the flowering
material. Thanks are due to the University Grants Commission, New Delhi,
and Marathwada University, Aurangabad, for the award of a teacher fellowship
to one of them (RAK). RAK also thanks the Government of Maharashtra
and the Principal, Government College of Arts and Science, Aurangabad, for
permission to accept the same.

References

*Agthe C 1951 Uber die Physiologische Herkunft des Pflanzennektars ; Ber. Schweiz. Bot. Ges.

61 240-277
Brown W H 1938 The bearing of nectaries an the phytogeny of flowering plants ; Proc. Am,

Phil. Soc. 79 549-59?

478

R A Kulkami and R M fai

J V 1973

,

Pittendrigh C S 19k48HT1^^°Q^e K "954 The floral anatomy of some Scitatmneae I .;

^7. Indian bat. Soc. 33 118^47 the Unes of evolution in the Bromeliaceae ;

Smith L B 1934 Geographical evidence on m

""-*• <R"WCMam'i

M^rathwada Univ. I97g The flora, anatomy of tlie Liliaceae-the

s N P, Markandeya a *. ana. ." .
*** /- *'• 1 61-168

; ***
Not consulted in the original

Proc. Indian Adad. Sci. (Plant Sci.)> Vol. 91, Number 6, December 1982, pp. 479- 486.
© Printed in India.

Cytological studies on certain Acanthaceae from Central India

M I S SAGGOO and S S BIR

Department of Botany, Punjabi University, Patiala 147002, India

MS received 17 September 1981 ; revised 31 August 1982

Abstract Cytological studies have been made on 19 species of Acanthaceae from
Pachmarhi hills in Central India. Present studies reveal the first count of chromo-
some numbers for four species, namely, Dyschoriste depressa Nees, n = 30 ;
Lepidagathis fasciculata Nees, n = 10 ; L. hyalina Nees, n = 10 and Justkia diffusa
Willd. var. prostrata Roxb., n = 9. New cytotypes have been located in three
species as Hemi%mphis latebrosa Nees, n = 28 (4x) ; Rungia parviflora Nees, n = 13
(2x) and jR. pectinata Nees, n = 13 (2x). Diploid cytotypes of three speder, v/z.,
Blepharis maderaspaiensis (Linn.) Roth, n — 15 ; Justida betonica Linn., n = 17
and Thunbergia alata Bojer ex. Sims, n — 9 have been detected for the first time
from India. An analysis of the worked out species reveals the existence of only
10-53% polyploid species.

Keywords. Cytology ; polyploidy ; cytotypes ; Acanthaceae.

1. Introduction

Acanthaceae is a large pantropical family of about 250 genera and 2,500 species
(Airy Shaw 1973). The members are chiefly distributed in tropics and sub-tropics
but are also found in the Mediterranean regions. There is great diversity in
habit and habitat of the members of the family which are usually herbs or shrubs
and very rarely small sized trees. Economically, the family is important due
to the presence of large numbers of ornamentals and some medicinal plants.

In India the family is represented by about 427 species falling in 81 genera
(Santapau and Henry 1973). Although the members are widely represented in
our flora and are of economic importance yet very little attention has been paid
towards their cytological analysis. The only contributions are by Narayanan
(1951); De (1964; 1966); Joseph (1964) ; Kaur (1965a,b, 1966, 1970; Kaur and
Nizam (1970); Vermaand Dhillon(1967); Datta and Maiti (1968, 1970), Sareen
and Sanjogta (1976) and Krishnaswami and Menon (1974). In spite of these,
the members of the family from Pachmarhi hills, Central India by and large
have not been worked out. Therefore, as a part of our project of cytological
studies on the flora of Central India, the present work was taken up in 1978.

479

480 MIS Saggoo and S S Sir

Table 1. Ciuromosome numbers in members of Acanthaceae from Central India.

Name of Taxan

Locality PUN Chromosome Ploidy

Accession number level

number/s

1

23 4

5

Adhatoda vasica Nees

Pachmarhi (M.P.) 24240 n - 17
Jata Shankar,
1,000 m.

Diploid

Blepharis mademspatensis (Linn.) Pachmarhi
Roth (= B. boerhaaviaefolia Pers) Little Fall,

950m.

Crossandra infundibuliformis Nees Pachmarhi

Raj Bhawan,
1,000m.

Didiptera bupleuroides Nees

D. roxburghiana Nees

*Dyschoriste depressa Nees

Gendarussa vulgaris Nees
(= Justida gendarussa Linn.)

**Hemigraphis latebrosa Nees

Pachmarhi
Jata Shankar,
1,000m.

Pachmarhi .
Pathar Chata,
900m.

Pachmarhi :
Raj Bhawan,
1,000 m.

Pipariya (M.P.) :
200m.

Pachmarhi :
Jata Shankar,
1,000m.

Hygrophila auriculata (Schum.) Heine Pachmarhi :
(=*H. spinosa T. And.) 1,050m.

Justida betonica Linn.

/. diffuse* Willd.

*J. diffusa Willd. var.
prostrata Roxb.

Pachmarhi :
Bee Fall,
950 m.

Pachmarhi :
Mandadeo Caves,
900m.

Pachmarhi :
Mandadea Caves,
900m.

20847 n « 15 Diploici
(figure 11)

24235 n = 19 Diploid

24207 /i = 13 Diploid

(%ure 2)

24208 n = 13 Diploid

(figure 1)

20841 TI = 30 Tetraploid
(figure 12)

24223 n « 15 Dipioid

(figuic 3)

20832 n « 28 Tetraploid
(figure 13)

24228 n « 16 Diploid

(figure 4)

20840 TI « 17 Diploid
(rigure 5)

24232 n - 9 Diploid

(figure 7)

20839 n * 9 Diploid

(figure 6)

Cytological Studies on Acanthaceae

481

Lepidagathis cuspidata Nees

Pachmarhi :
Dhupgarh,
1,200m.

24239 H = 11
(figure 14)

Diploid

L. fasciculata Nees

Pachmarhi :
1,000m.

20830 n = 10
(figure 16)

Diploid

*L. hyalitiG Nees

Pachmarhi :
Jambu Dwip,
950 m.

20838 n = 10
(figure 15)

Diploid

Petalidium bralerioides Nees

Pachmarhi :
Pagara,
800m.

24237 n = 16
(figure 8)

Diploid

**Rungia parviflora Nees

Pachmarhi :
Jata Shankar,
1,000m.

20833 n = 13
(figure 17)

Diplcid

*'*jR. pectlnata (Linn.) Nees

Pachmarhi :
Polo garden,
950m.

20842 n - 13
(figure 9)

Diplcia

Thunbergia alata Bojer ex Sims

Pachmarhi :
1,050m.

24226 n = 9
(figure 10)

Diplcid

* Reported for the first time.

** Reports of new chromosome number. For previous reports reference is made to Darlington
and Wylie (1955), Love and Love (1961, 1964, 1975) Fedorov (1969). Index to Plant Chromo-
>ome numbers (1956-1974) and IOPB chromosome number reports (1964 onwards).

2. Material and methods

Materials for the present investigations were collected from different populations
>f wild as well as cultivated plants from Pachmarhi and its surroundings. The
specific locality and altitude of each taxon are indicated in table 1. For meiotic
itudies anthers from young flower buds were squashed in 1% acetocarmine after
ixing for 24 hrs in carnoy's fluid. Slides were made permanent in the usual
nanner and mounted in cuparal. Voucher specimens have been deposited in the
rlerbarium, Punjabi University, Patiala (PUN).

f. Observations

nformation about chromosome numbers of the presently worked out 19 species
>elonging to 13 genera of the family has been provided in table 1 (figures 1-17).
Dourse of meiotic division was found to be normal in all the worked out taxa.

L Discussion

'iemigraphis latebrosa Nees with n = 28 is a new record for the species and is
etraploid. Earlier Sareen and Sanjagta (1976) recorded a diploid species with

P (B)— 2

482 MIS Saggoo and S S Sir

n = 14 from North India. Both the presently worked out species of Rungia
namely, R. parviflora Nees' and R. pectinata Nees are diploid with n = 13
Record of n = 13 for R. parviflora Nees is new because the earlier reports are
of 7i = 8 (Baquar 1967-68), n = 15 (Datta and Maiti 1970) and n = 26 (Mehra
and Vasudevan 1972). Similarly, count of n = 13 for R. pectinata Nees is
different from that of 2n = 50 by De (1966) for the same species. So far only
3 species of Rungia are known cytologically. Datta and Maiti (1970) suggested
x = 10 to be the base number for the genus, but the possibility of polybasic
nature of genus with numbers x = 8, 10 13, 15, 25 cannot be ruled out although
x = 15, 25 would naturally be secondary basic numbers.

Cytotype of Blepharis maderaspatensis (Linn.) Roth with n = 15 has been
worked out for the first time from India. Earlier, Kaur (1966) recorded n = 13
from South India. From Africa, n = 15 had earlier been reported by Miege
(1960). Apparently, the present species is diploid. Cytological information about
Blepharis is meagre since only 5 out of a total of 100 species have been worked
out so far. Somatic members reported for the genus are In = 24, 26, 30, 34,
clearly indicating polybasic nature with base numbers x = 12, 13, 15, 17.

Dyschoriste depressa Nees (n = 30) has presently been worked out for the first
%ne and is a tetraploid. Base number x = 15 is well established in all the
cytologically worked out species. Justida diffusa Willd. var. prostrata Roxb.
with n = 9 has been worked out for the first time. Justida diffusa Willd. with
n = 9 and /. betonica Linn, with/* = 17 confirms the earlier reports of n = 9 by
Mehra and Vasudevan (1972) and Bir and Sidhu (1974) for the former and n = 17
by Ellis (1962) for the latter. Narayanan (1951), however, recorded In = 28 for
J. betonica Linn. A perusal of literature reveals the polybasic nature of the
genus with x = 9, 13, 14, 15, 16, 17. Base numbers x = 9 and 14 are of
common occurrence. Thus all the three presently worked out species of Justida
are at diploid level. Lepidagathis fasdculata Nees (n = 10) and L. hyalina
Nees (n = 10) have been worked out for the first time. The other species,
L. cuspidata Nees with n = 11 confirms the earlier counts by Verma and Dhillon
(1967). All the three present species are diploid.

The basic numbers in the family range from x = 8-25 but the commonest
numbers are jc = 9, 14, 16, 17. A number of genera show polybasic nature as
Barleria (x = 15, 16, 19, 20), Blepharis (x = 12, 13, 15, 17), Thunbergia (x « 9,
10, 14, 16), Rungia (x = 8, 10, 13, 15, 25), Justida (x = 9, 13, 14, 15, 16, 17)
and Strobilanthes (x = 8, 9, 10, 11, 13, 14, 15). The variability in the base
numbers and polybasic nature of several genera clearly indicate that cytologically
the family is a highly evolved one and all this could possibly be the result of
aneuploidy operative at generic level. This has led to the evolution of morpho-

Figures 1-10. Meiosis in pollen mother cells. 1. Didiptera roxburghiana, M-I
with 1311. 2. D. bupleitroide.-, n = 13 at M-I. 3. Gendarussa vulgam, M-I
with 1511. 4. Hygrophila auriculata, M-I with 1611. 5. Justida betonica, 1711
at M~T. 6. J. diffusa, n = 9 at diakinesis. 7. /. diffusa var. prostrata, h =* 9.
8. Patalidium barlenoides, 1611 at M-I. 9. Rungia pectinata, 13H at M-I.
10. Tliunbergia date, M-TI with n = 9.

Studies on Acanthaceae

483

FIGS. 3.8.l6

20nm
FIGS.1.2A.5.6.7.9

Figures 1-10.

Cytological studies on Acanthaceae 485

Figures 11-1 ;. Meiasis in pcllen mother cells. 11. Blepharis niaderaspatensis,
n— 15 at diakinesis. 12. Dyschoriste depressa, 3011 at M-1. 13. Hemigraphis latebrosa,
diakinesis showing 2811. 14. Lepidagathis cuspidata, n = 11. 15. JC. hyalina
showing 10 + 10 chromosomes, at M-II. 16. L. fasciculate* with 1011 at M-I.
17. Rungia parviflora, M-I showing n = 13.

ogical variations. Out of presently investigated 19 species of Acanthaceae only
wo (10*53%) are polyploids and both at tetraploid level only. Taking an overall
dcture on the basis of the world- wide cumulative data, it is seen that only 17*81%*
>f the worked out species are polyploids. Intraspecific polyploidy is so
ar reported in only eight species, namely, Crossandra infundibuliformis (2x, 6x) ;
7. nilotica (4#, 6#) ; Didiptera elagans (2x, 4x), Hemigraphis latebrosa (2x, 4x) ;
lungia parviflora (2x, 4x) and Thunbergia grandiflora (2x> 4x). It appers that
»olyploidy has not been as potent a factor of cytological evolution in the family
3 aneuploidy.

Out of 2500 species in Acanthaceae, only 219 are worked out and amonsst these 180 soecies
:e diploid and 39 polyploids (for details see footnote to table 1).

486 MIS Saggoo and S S Bir

Acknowledgement

One of the authors (MISS) is thankful to CSIR, New Delhi, for the award of , i

Research Fellowship. . j

References ;

Airy Shaw H K 1973 A dictionary of the flowering plants and ferns (ed.) J C Willis 8th ecL f

(Cambridge : Univ. Press) pp -ixxii. 1-1245 I

Baquar S R 1967-6$ Chromosome numbers in some vascular plants of East Pakistan ; Rev. J

BioL 6 400-448 (

Bir S S and Sidhu M 1974 In Love A : IOPB chromosome number repoits XLIV ; Taxon

23 373-380 j

Darlington C D and Wylie A P 1955 Chromosome atlas of flowering plants (London : George !

Allen and Unwin Ltd.) pp. i-xx, 1-519
Datta P C and Maiti R K 1968 Chromosomal biotypes of Adhatoda vasica Nees. growing in

the Eastern Part of India ; Cytologia 33 220-229
Datta P C and Maiti R K 1970 Relationships of Justicieae (Acanthaceae) based on cytology ;

Genetica 41 431-450
De A 1964 Cytological investigations in the family Acanthaceae and importance in the study J"

of phylogeny ; Proc. 51st and 52nd Indian Sci. Cong. Session Sect. Bot. Abst. pp. 354
De A 1966 Cytological investigations ir> the family Acanthaceae ; Sci. Cult. 32 198-199
Ellis J L 1962 Chromosome numbers in some members of Acanthaceae ; Sci. Cult. 28 191-192
Fedorov An A (ed) 1969 Chromosome numbers of the flowering plants ; Academy of Sciences

of the USSR Komarov Botanical Institute Leningrad (1974 reprint) pp. 1-928.
Joseph J 1964 Chromosome numbers and abnormalities observed in a few members of

Acanthaceae ; Curr. Sci. 33 56-57 |*

Kaur J 1965a Chromosome numbers in Acanthaceae J ; Curr. Sci. 34 295 i

Kaur J 1965b Chromosome numbers in Acanthaceae II ; Sci. Cult. 31 D3 1-532 ;

Kaur J 1966 Chromosome numbers in Acanthaceae III ; Sci. Cult. 32 142-143

Kaur J 1970 Chromosome numbers in Acanthaceae V ; Sci. Cult. 36 103-106 !

Kaur J and Nizam JT 1970 Karyotype analysis of some members of Acanthaceae ; Nucleus j

13 23-28 j

Krishnaswami S and Menon P M 1974 Cytomorpho logical studies on some species and an

interspecific hybrid of Barleria L. ; Cytologia 39 397-402 ' \

Love A anc L5ve D 1961 Chromosome number of Central and North-West European plant species ; I

Opera Botanica (Stockholm : Ainquist and Wiksell) pp. 1-581
Love A and Love D 1974 Cytotaxonomical Atlas of the Slovenian Flora ; J. Cramer. FL-3301

Lehre pp. i-xx, 1-1241 »,

Love A and Love D 1975 Cytotaxonomical Atlas of the Arctic Flora ; /. Cramer. FL-9490

Vaduz pp. i-xxiii, 1-598
Mehra P N and Vasudevan K N 1972 In Love A : IOBP chromosome number reports

XXXVI ; Taxon 21 333-346
Miege J 1960 Troisieme lists de nombres chiamosomiques despecas d' Afrique o> cidentale ;

Am. Fac. Sci. Univ. Daker 5 75-85

Naiayan?n C R 1951 Somatic chromosomes in the Acanthaceae; J.Madras Univ. B21 220-231 ;

Santapau H and Henry A N 1973 A Dictionary of the Flowering Plants in India (New Delhi CSIR)

pp. i-viii, 1-198
Sareen T S and Sanjogta, K 1976 Chromosome numbers in some species of Acanthaceae ;

Cytologia 41 283-290
Verma S C and DhUlon S S 1967 In Love A : IOPB chromosome number reports XI : Taxon :

16 146-157

Proc. Indian Acad. Sci. (Plant Sci.), Vol. 91, Number 6, December 1982, pp. 487-493.
© Printed in India.

Heterotrophic bacteria associated with seaweed

P LAKSHMANAPERUMALSAMY* and A PURUSHOTHAMAN

Centre of Advanced Study in Marine Biology, Annamalai University,
Parangipettai 608502, Tamil Nadu, India

* Present address : Department of Marine Sciences, Univeisity of Cochin,
Cochin 682016, India

MS received 30 March 1982 ; revised 30 August 1982

Abstract The heterotrophic bacterial population associated with seaweeds
(Enteromorp/ta sp., Chaetomorpha sp. and Hypnea sp.) and water of the Vellar
Estuary, Porto Novo, were estimated. Total heterotrophic bacteria associated
with the seaweeds were found to be more abundant than in water samples. Repre-
sentative cultures wore isolated and their morphological and biochemical charac-
teristics were studied. In addition, pi eduction of amylase, lipase and protdnase
of the isolates was also studied. Bacillus, Corynebacterium, Vibrio, Alcaligenes
and Pseudomonas woro the genera commonly encountered. The role of these
bacteria associated with seaweeds is discussed.

Keywords. Heterotrophic bacteria; seaweed; Enteromorpha sp. ; Chaetomorpha sp.
Hypnea sp.; Vellar estuary; Porto Novo.

1. Introduction

Available information, on mioro-o^gantsrm in an estuarins environment concerns
water and sediment predominantly. The epiphytic bacterial flora of seaweed
appears to have been neglected. Bacteria adhere to many types of solid surfaces,
probably by means of several mechanisms and in some cases in a selective manner
(Daniel 1972 ; Gibbons and van Houte 1975 ; Marshall 1976). Currently increasing
attention has be0n paid to bacterial adhesion and its ecological significance. Some
are adhesive to rocks in flowing streams (Geesey et al 1977), to suspended matter
and solid surfaces in an estuary (Goulder 1976, 1977 ; Austin et al 1979a), to
animal surfaces (Gibbons and van Houte 1975) and to sea weed (Sieburth 1962,
1968 ; Kong and Chan 1979 ; Sjoblad and Mitchell 1979 ; Shiba and Taga 1980).
Much interest has not been shown on the epiphytic bacteria associated with
seaweed in Indian estuaries, except by Chandramohan (1971). The purpose of
the present study is to describe hetepotrophic bacteria present in water and in
association with seaweed (Enteromorpha sp. ; Chaetomorpha sp. ; and Hypnea sp.)
at a particular location in a tropical estuary such as Vellar.

487
P. (Bh-3

488 P Lakshmanaperumalsamy and A Purushothaman

2. Material and methods

Seaweed samples were collected and transferred to sterile polyethylene bags after
draining completely. Water samples were collected in sterile glass bottles. The
samples were stored in a small insulation container (5° C) and brought to the
laboratory. All the samples were plated within an hour of collection.

A portion of the seaweed was washed separately in sterile water and trans-
ferred to 100 ml sterile estuarine water blank in a 500 ml flask shaken on a
reciprocal shaker (120 strokes per min) for 30 min at room temperature. The
bacterial population was estimated by serial dilution plate method using Estuarine
Peptone Yeast Extract Agar (EPYA; Bacto Peptone 1 %, Yeast extract Difco 0-3%,
Bacto Agar 2%) and the bacterial population was expressed as number s/g dry
weight of the seaweed and per ml of water. Bacterial strains were randomly
isolated, purified by repeated streaking and identified to various genera using
the taxonomic key of Simidu and Also (1962). Amylolytic, proteolytic and
lipolytic ability of the isolates were also tested (Harrigan and McCance 1972)«

3. Results

The total aerobic heterotrophic bacterial population from all the samples were
estimated and the results are presented in table 1. The epiphytic bacterial popu-
lation associated with Chaetomorpha sp. varied from 1*16 to 9*22 x 108/g,
Enteromorpha sp. from LI 3. to 18-63 X 106/g and Hypnea sp. from 1*14 to
13-75 x 106/g dry weight. Estimates of bacterial populations in the water sample
ranged from 3*9 x 103 to 1 -53 x 10*/ml. The maximum bacterial population
among the samples was recorded in October. Highet? (18-63 x 106/g) number
of epiphytic bacteria was found to be associated in Enteromorpha sp. followed
by Hypnea sp. and Chaetomorpha sp. Since this year experienced a heavy rain-
fall in monsoon season, no seaweed was found in the estuarine environ-
ment in November 1978 through February 1979, and this might be due to
heavy freshwater inflow and lower salinity. They appeared again in March. So,
no data could be collected between November 1978 and February 1979.

The total heterotrophic bacterial flora of water in Vellar estuary was found to
consist of Bacillus (13-33%), Corynebacterium (20%,) Micrococcus (10%), Vibrio
(16*67%), Pseudomonas (16*67%), Alcaligenes (10%), Flavobacterium-Cytophaga
group (10%), and Enterobacteriaceae (3*33%) as shown in table 2. In general,
gram-negative bacteria were more common (56*67%) than gram-positive. How-
ever gram-positive groups (Bacillus, Corynebacterium and Micrococcus) were
abundant in April than the gram-negative groups. Bacillus, Vibrio, Pseudomonas,
Alcaligenes, Flavobacterium-Cytophaga were recorded in all the samples. The
maximum number of Corynebacterium was recorded in March. The results
suggest that the dominant epiphytic flora associated with seaweed, Chaetomorpha
sp., Enteromorpha sp. and Hypnea sp., is Vibrio. Various genera associated were
Akaligenes, Flavobacterium-Cytophaga group, Pseudomonas, Enteiobacteriaceae,
Corynebacterium and Micrococcus. The domination of gram-negative bacteria
was observed in all collections. The epiphytic 'Vibrio "peak was recorded in
October in association with all three seaweeds and a reduction in number was

Heterotrophic bacteria associated with seaweed
Table 1. Total aerobic hetero trophic bacterial population.

489

Month Chacromorpha sp. Enteromorpha sp.
(106/g) (10ft/g)

Hypnea sp.
(108/g)

Water
(10* /ml)

September 1978
October
March 1979
April
May

3-51
9-22

3-81
1-76
1-16

3-19
18-63
2-93
1-36
1-13

4-32
13-75
4-10
1-49
1-14

4-9
15-3
15-1
3-9
4-2

Table 2.

Total number of

isolates assigned to

various genrra.

Choetomorpha sp. Enteromorpha sp.

Hypnea sp.

Water

Total nurr-ber of
isolates tested

72

74

73

60

Bacillus

...

...

...

8
(13-33)

Corynebacterium

2
(2-78)

3
(4-05)

1
(1'37)

YL
(20-00)

Micrococcus

2
(2-78)

1
(1*35)

...

6
(10-0)

Vibrio

42
08-33)

38
(51-35)

40
(54-8)

10
(16-67)

Pseudomonas

8
(1M1)

11
(14-86)

14

(19-18)

10
(16-67)

Alcallgenes

4
(5-56)

3
(4-05)

5
(6-85)

6
(10-0)

Fkvobacterium-Cytophaga 8
group (11*11)

10
(13-51)

9
(12-33)

6
(10-0)

Enterobacteriaceae

6
(8-33)

8
(10-81)

4
(5-48)

2
(3"33)

(Values in parentheses are percentages in total number of isolates tested).

noticed in March. Members of genus Vibrio could be isolated throughout the
period of study and formed major portion of the flora, followed by Flavobacterium-
Cytophaga group, Pseudomonas and Alcaligenes,

490 P Lakshmanaperumalsamy and A Purushothaman

The bacterial flora in water and in association with seaweed were assigned to
various physiological groups (table 3). Majority of heterotrophs in water appeared
proteolytic (63*33%), lipolytic (61*67%) and a low percentage (43 '33%) amylo-
lytic. Bacterial populations associated with Chaetomorpha sp., was dominated
by proteolytic (56*16%) followed by lipolytic (48.61%).

4. Discussion

Results of this study on the bacterial populations in water are comparable to
those reported for Long Island Sound, Chesapeake Bay and Tokyo Bay.
Altschular and Riley (1967) and Murchelano and Brown (1970) reported 103 to
104 bacteria pcsr ml in Long Island Sound and 8' 4 X 101 to 2*0 X 104/ml in
Chesapeake Bay and 1* 8 x 101 to 9" 1 X 104/ml in Tokyo Bay (Austin et al
1979b).

The maximum bacterial populations were recorded in October and minimum
in April, within the period of study. According to Velanker (1969), the numerical
magnitude of heterotrophs broadly parallels the distribution of other living orga-
nisms and the dissolved organic matter in the sea. Murchelano and Brown (1970)
suggested that the annual bacterial cycle in Long Island Sound coincided with
that of phytoplankton. Phytoplankton constitutes a locus for bacterial attach-
ment and produces organic substrates for bacterial utilization. Ecological para-
meters, viz., physical, chemical and biological can have a definite influence on the
bacterial population in an estuary. The annual bacterial cycle can be studied
by repetitive sampling at one geographic locus, along with environmental para-
meters. Since this study was done for only five months, no definite conclusions
could be reached regarding the annual cycle. The density of bacterial populations
associated with seaweeds varied considerably, from 1*3 to 18*63 x 106/g. The
epiphytic populations may result from availability of vitamins, growth factors or

Table 3. Physiological groupings of the bacterial isolates.

Sample

Total cumber
of isolates

Proteolytic

Amylolytic

Lipoly'ic

Chaetomorpha sp.

77

39
(54-16)

12
(16-67)

35
(45-61)

Enteromorpha sp.

74

42
(56-75)

9
(12-16)

37
(50-0)

Hypnea sp.

73

35
(47-94)

13
(17-81)

35
(47-94)

Water

60

3g
(63-33)

26

(43-33)

37
(61-67)

(Values in parantheses are percentages of isolates tested).

Heterotrophic bacteria associated with seaweed 491

other external metabolites extruded by actively growing algal filaments which
nourish the epiphytic flora (Fogg 1966 ; Sieburth 1968 ; Wetzel 1969 ; Wetzel
and Allen 1972 ; Bell etal 1974; Morishita etal 1978). Chandramohan (1971)
reported l"26xlOG/g bacterial population associated with Enteromorpha
mtest inalls in the Vellar estuary. Actively growing algal filaments were reported
to carry high numbers of epiphytic bacteria (Sieburth 1968 ; Ramsay and Fry
1976). Mary (1977) recorded high bacterial counts when mullets wefte fed on
large quantities of green filamentous alga Enteromorpha sp., which might account
for the epiphytic flora associated with the algae in the same area. Quantitative
data for bacteria were significantly high when there was an abundance of phyto-
plankton, Navicula sp.

The generic composition of the bacteria in water showed the presence of various
groups in Vellar estuary. In general, gram-negative bacteria were predominant
compared with gram-positives. On a generic basis, Corynebacteriwn were domi-
nant (20%), followed by Vibrio (16' 67%), Pseudomonas (16' 67%) and Bacillus
(13*33%). Murchelano and Brown (1970) reported that Pseudomonas was the
dominant bacterial group in Long Island Sound and. Pseudomonas, Achromobacter
and Flavobacterium composed of 92* 3% of the bacteria isolated. Vibrio, Bacillus,
Micrococcus and Cytophaga accounted for only 7* 3%. The various studies carried
out using water samples showed a domination of Vibrio and Achromobacter
in Chesapeake Bay, Vibrio in Kanagawa Bay, Pseudomonas in Long Island Sound
and Flavobacterium in Naragansett Bay (as cited by Murchelano and Brown 1970).
Austin etal (1979b), in their comparative study, found that the prominent aerobic
heterotrophic bacterial flora of the water column of Chesapeake Bay consisted of
Vibrio, Achromobacter, Pseudomonas and Corynebacterium, but in Tokyo Bay
predominantly Acinetobacter-Moraxella-tifo species, Caulobacter and Pseudo-
monas. It is difficult to quantify the bacterial genera diversities unless considerable
data are collected.

It may be inferred from the study reported here that members of seven genera
were found associated with seaweeds. Vibrio was found dominating over Pseudo-
monas, Flavobacterium and Alcaligenes. Abundance of Vibrio and Alcaligenes
during luxurious growth of algae (Enteromorpha sp.) suggested that these genera
were probably associated with algal bloom. The recovery of large population
of Vibrio throughout the study suggests that they form part of indigenous flora
of seaweed. Presence of epiphytic flora of marine algae has been reported by
Sieburth (1968) who examined species of Polysiphonia and Sargassum foi? possible
generic specificity of epiphytic flora and noticed Vibrio as the dominant species in
both. Kong and Chan (1979) reported seven genera to be associated with marine
algae and similar study was conducted by Shiba and Taga (1980) who found that
Flavobacterium was the dominant microflora. Vibrio were present in minimum
numbers, suggesting metabolites being toxic for Vibrio sp., a phenomenon not
observed in this investigation. No other taxonomic study has been carried out
to date on epiphytic bacteria associated with seaweed in Indian estuaries. The
generic composition, seasonal variation and interrelationships of associated hetero-
trophic bacteria remain to be defined.

The existence of bacterial populations on seaweed suggests that there exists a
beneficial relationship between the seaweed and the epiphytic bacteria. The
bacteria belonging to the genus Vibrio were predominant on the seaweed. It is

492 p Lakshmanaperumalsamy and A fiimshothaman

reported that the extract of seaweed acted as attractant for the marine bacterium,
Vibrio algmolyticus, by Sjoblad and Mitchell (1979). Hence it may be suggested
that a beneficial relationship exists between seaweed and their epiphytic bacteria
Vibrio.

Acknowledgements

Thanks are due to Dr R Natarajan, Director, for his encouragement and to
Dr D Chandramohan, Scientist, NIO, Goa, for his comments. One of the
authors (PL) thanks CSIR and UGC for financial assistance.

References

Altschuler S J and Riley G A 1967 Microbiological studies in Long Island Sound ; Bull.

Mnghain. Oceavogr. Coll. 19 81-88
Austin B, Allen D A, Zachaiy A, Belas M R and Colwell R R 1979a Ecology and taxonomy

of bacteria attaching to wood surfaces in a tropical harbour ; Can. J. Microbiol. 25 447-461
Austin B, Garges S, Conrod B, Harding E E, Colwell R R, Simidu U and Taga N 1979b

Compai stive study of the aerobic heterotrophic bacteiial flora of Chesapeake Bay and

Tokyo Bay ; Appl. Environ. MicrobioL 37 704-714
Bell W H, Lang J M and Mitchell R 1974 Selective stimulation of marine bacteria by algal

extracellular products ; Limnol. Oceanogr. 14 833-839

Chandramohan D 1971 Indole acetic acid synthesis in sea ; Proc. Indian Acad. Sci. L23 105-109
Daniel S L 1972 The adsorption of microorganisms onto solid surfaces: a review ; Dev. Indian

MicrobioL 13 211-253
Fogg G E 1966 Extiacellular products of algae ; In Octanogr. Mar. Biol Ann. Rev. (ed)

H Barnes (London : George Allen and Unwin Ltd.) 4 195-112

Geesey G G, Richardson W T, Yeomons H G, Irvin R T and Costerton J W 1977 Micro-
scopic examination of natural sessile bacterial populations from an alpine stream ; Can. J.

MicrobioL 23 1733-1736
Gibbons R J and van Houte 1975 Bacterial adherence in oral microbial ecology ; Ann. Rev.

MicrobioL 29 19-44
Gaulder R 1976 Relationship between suspended solids and standing craps and activities o^

bacteria in an estuary a neapspring-neaptide cycle ; Oecologia 24 83-90
Goulder R 1977 Attached and free bacteria in an estuary with abundant suspended solids ;

/. Appl. BacterioL 43 399-405
Harrigan W F and McCance M E (ed) 1972 Laboratory methods in microbiology (New York,

London : Academic Press) p. 362
Kong M K and Chan K 1970 A study on the bacterial flora isolated from marine algae ;

Bot. Mar. 22 83-97
Maishall K C fed) 1976 Interfaces in microbial ecology (Cambridge, Mass. : Harvard University

Piess) p. 156

Mary P P 1977 Studies in the gastrointestinal microflora of the mullet Liza dussumieri (Valen-
ciennes) (Mufcilifarms : Teleosti) ; Ph.D. thesir, Annamalai University, p. 122
Morishita H, Sano T, Kamiya N and Okuda M 1978 Growth stimulating substances for

Vibrio alginolyticus contained in Chloretta extract ; Bull. Jpn. Soc. Sci. Fish. 44 665-671
Murchelano R A and Brown C 1970 Heterotrophic bacteria in Long Island Sound ; Mar.

Bioll 1-6
Ramsay A J and Fry J C 1976 Response of epiphytic bacteria to the treatment of two aquatic

macrophytes with herbicide paraquat; Water. Res. 10 453-459

Shiba T and Taga N 1980 Heterotrophia bacteria attached to seaweeds ; J. exp. mar. Bid
7 251-258- . '• - -, - . .

Heterotrophic bacteria associated with seaweed 493

Sieburth J McN 1962 Biochemical warfare among the microbes of the sea ; Honors lecture,
University of Rhode Island, Kingston, 27 November 1962

Sieburth J McN 1968 The influence of algal antibiotics on the ecology of marine microorganisms ;
In Advances in the microbiology of the sea— I (eds) M R Droop and E J F Wood (New
York : Academic Prtss) pp. 63-94

Simidu U and Aiso K 1962 Occurrence and distribution of heterotrophic bacteria in seawater
from Kamogawa Bay ; Bull. Jpn. Soc. Fish. 28 1133

Sjoblad R B and Mitchell R 1979 Chaemotactic responses of Vibrio alginolyticus to algal
extracellular products ; Can. J. Microbiol 25 964-967

Velankar N K 1969 Bacteria in the ecology of the ocean with particular reference to Indian
Ocean ; Bull. Natl Inst. Sci. India No. 38 Part II 737

Wetzel R G 1969 Excretion of dissolved organic compounds by aquatic macrophytes ; Bio-
sciences 19 539-540

Wetzel R G and Allen H L 1972 Functions and interactions of dissolved organic matter
and the littoral zone in lake metabolism and eutrophication ; In Productivity problems in
freshwaters (eds) Z Kajak and A Hillbricht-IIkowaka (Warszawakrakowa) pp. 333-347

Proc. Indian Acad. Sci. (plant Sci.), Vol. $1. lumber 6, fiecember 1^8:2, pp.
© Printed in India.

Association of chlorophyll content, phyllotaxy, photosynthesis and
B group vitamins in some C3 and C{ plants

P GOPALA RAO and J KODANDARAMAIAH

Department of Botany, Sri Venkateswara University, Tirupati 51 7 502, India

MS received 17 November 1981 ; revised 21 October 1982

Abstract. The phatosynthetically efficient C4 plants viz Amamnthus viridis, Euphorbia
hirta and a C3 plant, Acalypha indica with mosaic leaf pattern showed the maximum
amount of B vitamins when compared to the other C3 plants. It is observed that
photosynthesis and vitamin synthesis go hand-in-hand showing close correlation.
The results also indicate that there is a close relation between chlorophyll content
and vitamin content. However, there appears to be no relation between phyllotaxy
and photosynthesis. Between the two C3 plants, viz., Acalypha and Carica* the
photosynthetic inefficiency of the latter might be due to more cf chlorophyll b and
less of chlorophyll a as seen from chlorophyll a] chlorophyll b ratios.

Keywords. Chlorophyll ; B vitamins ; phyllotaxy ; photosynthesis ; C% and C4
plants.

1. Introduction

The chlorophyll content of the cell must be closely associated with photosynthetic
activity because the photosynthetic rate is proportional to the chlorophyll concen-
tration (Maksymowych 1973), but there are contrary reports also. Black (1972)
concluded that the high rate of photosynthesis/mg chlorophyll is directly related
to the low chlorophyll content. The photosynthetic rates of different chlorophyll
mutants of pea, soybean, cotton and tobacco were studied by Benedict (1972).
The photosynthetic rate/mg of chlorophyll in mutants is 2-11 times faster than
in the wild type leaves. This phenomenon is compared to the photosynthetic
rate of the yellow sectors of variegated leaves. These yellow sectors although
containing a reduced chloropyhll content, have a much higher photosynthetic
rate/mg chlorophyll than the green sectors. Bonner and Bonner (1948) found
that thiamine synthesis in seedlings is often light-dependent and mature leaves of
full grown tomato plants are the centre of production. Gustafson (1948) sugges.
ted that light stimulates thiamine biosynthesis. It was stated that the photosynthe-
tic unit and not the chlorophyll content determines the rate of photosynthesis
(Black 1972).

495
P. (B)— 4

496 P Gopala Rao and J Kodandaramalah

The present investigation has been designed to understand the relationship
between chlorophyll content, vitamin content, phyllotaxy and photosynthesis in
some C3 and C4 plants. Work on the relationship between phyllotaxy and
photosynthesis is scanty. Earlier work (Evans 1975) indicates that canopies
with more vertically inclined leaves have a higher photosynthetic rate than those
with horizontal leaves.

2. Materials and methods

Young and fully expanded leaves from various plant species, viz., (1) Acalypha
indica, L., (2) Amaranthus viridis, L., (3) Carica papaya, L., (4) Commelina
benghalensis, L., (5) Euphorbia hirta, L., (6) Euphorbia pulcherrima, Willd., (7)
Ervatamia coronaria, Stapf., (8) Werium odorum, Soland., (9) Nyctanthus arbor,
tristis, L., (10) Petunia hybrida, L., (11) Sida acuta, Burn, (12) Tridaxprocumbens,
L., growing in the university Botanical Garden under natural photoperiod
constitute the experimental material.

Chlorophylls were extracted with 80% acetone and estimated according to the
method of Arnon (1949). The chloroplasts were isolated using N/15 phosphate
buffer (pH 7*3) containing sucrose (0-33M), disodiumsalt ofEDTA (2 X 10~3M)
dithiothreitol (5 X 10~3 M), MgCl2 (1 X 10~3 M), MgSO4 (1 ' 5 x 10~3 M) and 5% (w/v)
polyvinylpyrolidene as isolation medium following the procedure of James and
Das (1957). The method of Jagendorf and Evans (1957) was used to assay Hill
reaction activity of chloroplast preparation. 14CO2 fixation studies were done
using a technique similar to that described by Berry et al (1970), using 14 C
sodium bicarbonate (specific activity, 27 mci/m mole) and the net photosynthesis
was expressed as mg CO2 fixed dm~2hr~1.

The different vitamins of the B group, viz., thiamine (BJ9 riboflavin (J?2),
pyridoxin (#6), niacin and folic acids were extracted and estimated colorimetri-
cally following the methods given by Manzur-ul-Haque Hashmi (1973) and the
results were expressed as //g/g dry wt. All the results were averages of three
individual experiments.

3. Results and discussion

The results in table 1 indicate that Amaranthus viridis, Euphorbia hirta which are
C4 photosynthetic plants and Acalypha indica, a C3 plant with mosaic pattern
of 'phyllotaxy have maximum Hill activity, net photosynthesis and chlorophyll
coatent. Commelina benghalensis, Euphorbia pulcherrima, Ervatamia coronaria and
Sida acuta occupy the next position in order in this respect. Nerium odorum
shows the minimum activity.

It is tempting to note that there is a close correlation between the photosyn-
thetic parameters (table 1) and vitamins of the B group (table 2) viz., thiamine
(Bj), riboflavin (52), pyridoxin (56), niacin and folic acid in the plants mentioned
above. Nerium odorum shows the minimum amount of vitamins of the B group.
The data thus gives a circumstantial evidence to show that photosynthesis and
vitamin synthesis go hand-in-hand showing a close correlation (table 3).
The photosynthetically efficient C4 plants viz., Amaranthus veridis, Euphorbia hirta

B group vitamins and photosynthesis

497

Table 1. The pattern of chlorophylls and the rate of photosynthesis in different
plant species.

Plant species Phyllotaxy

Plant Total Chlorophyll Hill
type chloro- alb ratio activity
phylls

# * _i_

Net
photo
synthesis

**

1.

Acalypha indica L. mosaic

C3 4-

62

1-

Ib

163-

5

38-

4

±0-

'84

±o-

04

4- 6-

21

± 2-22

2.

Amaranthus viridis L. alternate

C4 3-

•58

1-

21

186-

2

45-

8

±0-22

±o-

12

± 10-

6

± 5-

16

3.

Carica papaya L. mosaic

C8 1-

90

o-

18

121-

1

26-

5

±0-31

±o-

01

± 7-

81

± 1-

76

4.

Commelina alternate

C3 2-

43

1-

17

148-

4

31-

6

benghalensis L.

±0

•14

±0

•06

± 3-

42

± 4-

32

5.

Euphorbia hirta L. opp-super-

C4 3

•96

1-

•25

169-

5

42'

0

imposed

±0

•86

±0

•22

± 12-

31

± 1-

78

6.

Euphorbia opp-decussate

C3 3

•47

!•

28

142'

6

36-

8

pWcherrima

±0

•86

±0'

•08

db 16-

2

± 6-

52

Willd.

7.

Ervatamia opp-decussate

C3 2

02

1-

25

136-

0

29-

1

coronoria Stapf.

±o

•10

±0

•12

d= 5-

61

± I'

•34

8.

Neriwn odorum Whcrled

C8 1

•69

1

63

84-

3

17-

2

So-land

±0

•21

±o-

•34

± 7-

52

± 2-

88

9.

Nyctanthes opp-super-

C3 1

•98

1

26

117-

5

21

•6

arbortristis L. imposed

±0

•28

±0

•07

± 11-

6

± 1-

06

10.

Petunia hybrida L. alternate

C3 1

•76

1

•31

104-

2

19

•o

-

±0

•11

±0

•26

± 18

•2

± 3

•82

11.

Sida acuta Burm. alternate

C3 3

•13

1

•17

126-

1

23

•2

±0

•16

±0-34

db 7-

•01

± 2

•17

12.

Tridax procumbens alternate

C3 1

•65

1

•24

100

•8

21

•8

L.

±0

•81

±0

•12

± 9

•44

± 4

•61

* mg gr1 fresh wt.
** mg CO2 fixed d-2m hr"1.
•f p moles of DCPIP reduced mg""1 chl hr"1.
^Values are means ±"S.E. of three individual experiments),

498 P Gopala Rao and J Kodandaramaiah

and Acalypha indica, a C3 plant with mosaic leaf pattern showed the maximum
amount of vitamins. Commelina benghalensis, Eurphorbia pulcherrima, Ervatamia
coronaria and Sida acuta form the second group in their vitamin contents quite
parallel to their photosynthetic parameters.

Tin results categorically indicate that there is a close relation between chloro-
phyll content and vitamin content (table 3). The maximum amount of chloro-
phyll (total) is seen in Acalypha indica (4-62mg/g fresh wt.). Nevertheless,

Table 2. Vitamin content 0*g g-1 dry wt.) in the leaves of different plant species.

Plant
$PP*

Thiamine
(Bj)

Riboflavin
(B3)

Pvridoxin
(B6)

Niacin

Folic acid '

Total

1.

369-0
± 12'1

124-0

± 8-9

215-0
± 16-2

152-0
± 20-4

134-0
± 6-2

994

2.

280-0
± 21-3

136-0
± 12-4

198-0

± 17-4

169-0
± 11-2

149-0
± 13-1

932

3.

114-0
± 16-2

82-0

± 3-2

128-0
db 15-8 •

97-0
±8-6

78-0

± 4-4

499

4.

232-0
± 31-0

64-0

± 5-7

176-0

± 2M

160-0

± 1M

52-0

± 2-3

684

5.

291-0
± 18-6

99-0

± 2-8

195-0

± 8-8

141-0

± 6-1

130-0

± 7-6

856

6.

207-0
i 26-2

76-0

± 4-9

180-0
± 11-2

122-0
± 10-1

104-0
± 11-2

689

7.

197-0
± 16-6

108-0
db 13-1

209-0

± 6-8

135-0

± 14-2

126-0
± 13-6

775

8.

74-0
± 8-1

42-0

± 3-6

96-0

± 2-4

28-0

± 3-8

49-0

± 6-4

289

9.

139-0
± 11*2

34-0

± 2-7

126-0

db 7-6

47-0
± 1-2

61-0

db 3-4

407

10.

98-0

± 14-1

51-0

± 2-6

113-0

± 3-2

62-0

± 5-6

93-0

± 2-8

417

11.

156-0
± 21-7

78-0
± 1-7

174-0
± 14-6

103-0

± 2-8

82-0
± 4-1

593

12.

124-0

± 6-4

46-0

± M

101-0
± 9-9

94-0
± 2-1

76-0

± 3-6

441

* Plant names as represented in table 1 serially.
(Values are means ± S.E. of three replications),

B group vitamins and photosynthesis
Table 3. Statistical analysis.

499

1. Correlation coefficient (for tables 1 and 2)
between a set of parameters

correlation coefficient

(a) Total chlorophylls x hill activity

-h 0-822(4-564)*

(b)

x photosynthesis

+ 0- 826 (4- 635)*

(c)

x thiamine (Bj)

•4-0-895(6* 346)*

(d)

x riboflavin (B2)

H- 0-730 (3 -378)*

(e)

x pyridoxin (B6)

+ 0-209 (0-676)

(0

x niacin

+ 0-676(2-901)*

(g)

x folic acid

+ 0-907(6-809)*

(h)

x total vitamins

4- 0-849 (5- 083)*

2. Analysis of variance (for table 2)

F calculated

F from table at 5% level

C.D. at 5% level

Vitami ns

26-765*

2-594

15-802

Plant species

10-515*
2-152
8-972

* Significant at 5% level.

Values in the parantheses represent the calculated values of f-statistic for testing the significance

of correlation coefficients.

chlorophyll a/b ratio is not the highest in Acalypha indica (table 1). Chlorophyll
alb ratio is maximum (1-63) in Nerium odorum, but the vitamin content is at a
minimum level indicating that neither chlorophyll a nor b has anything to do
with vitamin synthesis, but it is the total chlorophyll content that is associated
with vitamin content, both being at a lower level (tables 1 and 3).

It is noted that there is no relationship between photosynthesis and phyllotaxy.
Mosaic pattern of phyllotaxy is assumed to help light to fall directly on all the
leaves without any impediment, thus making the plant photosynthetically more
efficient. But, if a comparison is made between Acalypha and Carica (both show
mosaic pattern) the latter is not at all efficient (table 1). Similarly Amaranthus veridfs
tias alternate leaves, while Euphorbia hirta which is also a C4 plant possesses
3pposite superimposed leaf arrangement thus indicating that there is no relation
Detween phyllotaxy and photosynthesis. In the case of opposite decussate pattern
in Ervatamia and Euphorbia pulcherrima where overshadowing is avoided, the
photosynthetic efficiency is somewhat better than that of Carica papaya with
mosaic pattern.

On the 'basis of computations it can be inferred that there is a highly positive
correlation between total chlorophyll content and other parameters such as Hill
activity, net photosynthesis, thiamine, riboflavin, niacin, folic acid and the total
/itamins (table 3). The correlation coefficients of all the parameters are signi-
icant at 5% level except with pyridoxin. Hence there is an association of chloro-
phyll content and other parameters (except phyllotaxy).

SCO P Gopala Rao and J Kodandaramaiah

Even an analysis of variance (for table 2) reveals a close association among
the parameters. F-values calculated for vitamins and plant species, are signi-
ficant at 5% level.

Acknowledgements

The authors are highly thankful to Prof. V S Rama Das for his constant
encouragement and for providing facilities and to Sri P Balasiddamuni,
Lecturer in Statistics, SV University, Tirupati, for his help in statistical analysis.

References

Ainon D I 1949 Copper enzymes in isolated chloroplasts ; polyphenal oxidase in Beta viitgaris ;

Plant Physiol. 24 1-15

Benedict C R 1972 S. Sect. Am. Soc. Plant Physiol 7-25
Berry J A, Downton W J S and Tregunna E B 1970 The photosynthetic carbon rn.tabalism

o.f Zea mays and Gomphrena globosa : the location of COa fixation and the carboxyl transfer

reactions ; Can. J. Bot. 48 777-780
Bonner J and Bonner H 1948 The B vitamins as plant hormones ; Vitam. Horm. (New York)

6 225-275
Black C S 1972 Net carbon dioxide assimilation in higher plants ; S. Sect. Am. Soc. Plant

Physiol. 1-93

Evans L T 1975 Crop Physiology, Cambridge University Press, pp. 336
Gustafson F G 1948 Influence of light intersity upon the concentration of thiamine and ribo-

flavin in plants ; Plant Physiol. 23 373-378
James W O and Das VSR 1957 The organization of respiration in chlorophyllous cells ; New

Phytol 56 325-343
Jagendorf A T and Evans M 1957 The Hill reaction of red kidney bean chloroplasts ; Plant

Physiol. 32 435-443
Manzur-ul-Haque Hashmi 1973 Assay of vitamins m pharmaceutical preparations John Wiley and

Sons.
Maksyrnowych R 1973 Analysis of leaf development Cambridge University Press pp, 86

Proc. Indian Aead. Sci. (Plant ScL), Vol. 91, Number 6, December 1982, pp. 501~5o8.
© Printed in India.

Effect of morphactin, AMO-1618 and DPX-1840 on the endogenous
levels of hormones and its implication on apical dominance in
Glydne max Linn*

I S DUA, U K KOHLI and K S CHARK

Department of Botany, Panjab University, Chandigarh 160014, India

MS received 4 April 1981 ; revised 24 September 1982

Abstract. Application of morphactin, AMO-1618 and DPX-1840 to. 20 day old
plants of soybean caused the sprouting of almost all lateral buds. However, the
follow-up growth of newly ensued buds was manifested only with morphactin
and not with AMO-1618 or DPX-1840. The quantitative estimations of growth
promoters, carried out 20 days after the application of inhibitors, revealed that
the passible mechanism through which these substances exerted their influence on
apical dominance varied with the type of regulator. It appeared that morphactin
lifted the apical dominance mainly through the increase in endogenous levels of
cytakinin and partly through lowering the levels of auxins. AMO-161S checked the
gibberellin turnover while DPX-1840 suppressed significantly the levels of auxins.
The data show that apical dominance and not the subsequent growth of newly
evocated buds is regulated by the ratio of cytokinins (CK) to auxins (Au) plus gibberel-
lins (QA) and any factoi(s) which enhance(s) CK (like morphactin) or suppresses)
GA (like AMO-1618) or Au (like DPX-1840 or morphactin) would play a key role in
the abolition of apical dominance.

Keywords. Apical dominance ; hormones ; growth retardants ; soybean.

U Introduction

Sver since Thimann and Skoog (1934) showed that an application of auxin prepa-
ration from fungus Rhizopus onto the cut stump of decapitated Vicia faba
3revented the growth of lateral buds, many workers have put forward a plethora
3f chemicals which could be utilised for abolition or induction of apical dominance
n intact plants. In spite of these successful reports by such widely diverse sub-
itances like IAA (Thimann et al 1971), GA3 (Bradley and Crane 1960), kinetin
Ali and Fletcher 1970), B-995 (Brooks 1964), AMO-1618 (Ruddat and Pharis
1966), morphactin (Tognoni et al 1967 ; Schneider 1970), TIBA (Morey and Dahl
1975) and various ethylene releasing compounds (Morgan and Durham 1972),
here has hardly been an attempt to divulge the underlying mechanism through

501

502 / S Dua, U K Kohli and K S Chark

which these regulators might be controlling this phenomenon. Dua et at (1978)
and Dua and Dhuria (1980) reported that morphactin-induced growth of lateral
buds was associated with a concomitant increase in the levels of endogenous
cytokinins. However, a comprehensive hormonal appraisal of apical dominance
with regard to morphactia vis-a-vis a few other retardants, though unequivocally
linked with this process, is still lacking. The present investigation attempts to
study this aspect beside probing into the mechanism of apical dominance in
Glycine max Linn.

2. Materials and methods

Glycine max Linn, cv ' Bragg ' was raised in pots of 25 cm height and 22-5 cm
in diameter under optimum conditions and 20 day old seedlings were sprayed with
different inhibitors with an atomiser. The treatments consisted of morphactin
(methyl-2-chloro-9-hydtoxy-fluorene- (9)-carboxylate-chlorofluornol-methyIestei-li
3456), DPX-1840 (3, 3a-dihydro-2-[Jp-methoxyphenyl]8H-pyiazolo-[5,l]-a isoindol-
8-one) and AMO-1618 (ammonium (5-hydroxycaivacryl) iirimethyl chloride
piperidine carboxylate) of 200, 1000 and 1000 ppm icspectively. The three
regulators were selected on the basis of previous work and have been used fre-
quently in relation to studies on apical dominance (see introduction for references).
Twenty- days after the spray, the data for various morphological characters were
recorded. For extractions of different endogenous plant growth substances, the
samples from 20-day old treated or control seedlings were kept in methanol for
48 hrs at 0°C in a refrigerator. The extract was evaporated under suction to
remove ethanol and residue (pH 3*0) was utilised for the extraction of various
growth regulatory substances. Extraction procedures described by Nitsoh (1956)
for auxins, by Murakami (1966) for gibberellins and by Dua and Jandaik (1979)
for different cytokinins were adopted. Estimates for auxin activity were carried
out on the basis of growth test of ' Kent ' oat (Avena sativa Linn.) coleoptiles as
described by Mer et al (1962), gibberellins by the modified technique of Ogawa
(1963) using 4Tainan-3' rice (Oryza sativa Linn.) and cytokinin by Xanthium leaf
disc senescence method of Osborne and McCalla (1961). Standard series of
cytokinins (zeatin, 2iP, zeatin riboside, zeatin ribotide), gibberellins (GA3, GA4,
GAS, GA?) and auxins (LA A, IAN) were run separately for their Rf values. The
data were statistically analysed according to the variance method.

3. Results

3 • 1. Morphological changes

The application of various growth retardants alone or in combinations brought
a number of morphological changes within 20 days of treatment (table 1). The
plant height vis-a-vis internodal length was reduced drastically. The maximum
reduction in length (56-1% over control) was recorded when all the three regula-
tors were present together. The most conspicuous morphological change was
the growth of lateral buds in all treatments and 50-65% buds sprouted as compared

Hormonal regulation of apical dominance

503

Table I. Effect of morphactin, AMo-1618 and DPX-1840 on the various morpho-
logical characters in Glycine max Linn, after 20 days of treatment (average of
5 replications).

Plant

Inter nodal

Number of

Length of the

Treatment

height

length

sprouted

lateral buds

(cm)

(cm)

lateral buds

(cm)

Control

56-21

18-25

2-00

2-23

Morphactin

42-25

13-16

13-00

8-76

AMO-1618

34-65

11-25

10-00

3-52

DPX-1840

38- 75

12-30

11-00

3-89

Morphactin -1- AMO-16'18

30-25

11-03

16-00

3-25

Morphactin + opx-1840

34-25

11-32

16-00

3-25

AMO-1618 + DPX-1840

28-25

10-97

15-00

3-30

Moiphactin + AMO-1618 -I- DPX-1840

26-23

10-81

17-00

4-25

Critical difference at 5 % level of
significance

3-63

2-12

1-50

1-93

to control where only 10% buds were growing. The effect of each chemical, in
promoting the growth of lateral buds, was further augmented in the presence of
the second retardant (75-80% buds growing) and maximum effect was detec-
table when all the three regvlators were present simultaneously (85% buds ini-
tiated). Another striking revelation was the growth of lateral buds, following
their release from apical dominance (table 1). Though all the regulatory sub-
stances significantly abolished apical dominance, to almost the same degree, the
subsequent growth of the newly ensued buds was insignificant and poor in all the
treatments except morphactin. It was only in the later treatment (morphactin)
that a significant follow-up growth of the axillary branches occurred (approxi-
mately 2-8 times more than control or "other treatments). Surprisingly, in
the treatments where morphactin was combined with other regulators, the
morphactin's promotory effect on the growth of lateral branches was also nullified.

3.2. Behaviour of different endogenous hormones

3.2a. Auxins'. The estimates of auxins revealed the presence of three auxins at
Rfs 0' 1-0-2, 0-6-0'7 (IAA) and O'9-l- 0 while IAN (Rf 0'4-0'5) was not discernible.
Application of morphactin and DPX-1840 resulted in significantly lowering the total
auxins (table 2 and figure 1) whereas AMO-1618 was ineffective in bringing about
any significant change. In combined tieatments, DPX-1840 or morphactin also
checked the levels of auxins and minimum auxin activity was noticed when both
of them were present simultaneously.

3.2b. Gibberellins : The data (table 2 and figure 1) showed that two gibber ellins
at Rfs 0-4-0 '5 (GA3) and 0*7-0 '8 (GA6) were detectable and GA3 contributed
the major component (80%) of the total gibberellins. The only treatment to
significantly check the levels of endogenous gibberellips was AMO-1618 singly or
in combination with other retardants which themselves were ineffective in causing
any such change.

P.(B)-5

504

Duct, U K Kohti and K S Chark

3.2c. Cytokinins : Bioassays of different extracts deciphered the presence of
four different cytokinins, namely, zeatin riboside (Rf 0' 1-0' 2), zeatin (Rf 0-4-0-5),
zeatin ribotide (Rf 0'8-0'9) and an unknown cytokinin (Rf 0'7-0*8) The treat-
ment of morphactin led to an increase of total cytokinins (table 3 and figure 1).

Table 2. Effect of morphactin, AMO-1618 and DPX-1840 on the endogenous level
of auxins and gibberellins Og/100g fresh weight) (average of 3 replications).

Auxins (Rf)

Gibberellins (Rf)

Treatments

0-1-0-2 0-6-0-7 0-9-1-0 0'4-0'5 0-70-8

Control

2-53

12-51

1-52

1-20

0-30

Morphactin

0-85

7-2S

0-85

1-15

0-30

AMO-1618

2-12

10-87

1-35

0-51

Tr

DPX-1840

0-56

5-33

0-80

1-08

Tr

Morphactin -f AMO-1618

0-84

7-35

0-86

0-49

0-25

Morphactin 4- DPX-1840

0*51

3-84

0-60

1-02

Tr

AMO-161S -f DPX-1840

0-58

5-52

0-85

0'44

Tr

Mcrphactin + AMO-1618 -f DPX-1840

0-52

3-91

0-65

0-43

Tr

Critical difference at 5% level of

significance

0-47

1-81

0-56

0-39

0-J3

400

2°0 5

71

Control Morph. AMO

DPX
IO-4O

o

Morph. Morph. AMO
+ + +

AMO DPX DPX

Id

Morph.-f

AMO+

DPX

Figure 1. Effect of morphactin, AMO-1618 and DPX-1840 on the endogenous
levels of total CK and GA (small shaded blacks inside the bigger blocks) in ngm
and auxins (in /*g) per 100 g of fiesh weight (average of 3 replications).

Hormonal regulation of apical dominance 505

Tables. Effect of morphaotin, AMO-1618 and DPX-! 840 on the endogenous
levels of cytokinin //g/100g fresh weight of kiuetin equivalent (average of
3 replications).

Cytokinins
Rf

Treatment ~

ZR Z UC ZRT

0-1-0-2 0-4-0-5 0*7-0-8 0-8-0-9

Control 0-058 0-035 0-021 0-061

Morphactin 0-091 0-083 0-062 0-096

AMO-1618 0-053 0-056 C'023 0-053

DPX-1840 0-057 0-059 0-028 0-041

Morphactin ^ AMO-1618 0- 089 0- 081 0' 079 0- 073

Morphactin H- DPX-1840 0- 093 0- 082 0- 078 0- 076

AMO-1618 + DPX-1840 0-061 0*063 0*028 0-058

Morphactin + AMO-1618 -h DPX-1840 0-089 0- 084 0* 080 0* 079

Critical difference at 5% level

of significance 0-008 O'Ol 0-01 0*009

Both AMO-1618 and DPX-1840 were ineffective in multiplying the level of individual
or total cytokinins. In combined treatments, the cytokinin enhancing capacity
of morphactin was not impaired by the presence of other substances.

4. Discussion

The habit of growth displayed by herbaceous and woody plants or control of the
plagiotropic position of the lateral branches are examples of correlative effects
where the stem apex influences the growth and development of the other parts of
the plant The way in which this phenomenon is mediated has been a matter of
controversy and different postulations, for different species, have been envisaged.
Some of the early investigators (Goebel 1900 ; Loeb 1918 ; Dostal 1926) and a
few of the late reports (Mclntyre 1968 ; Wagner and Michael 1971) have empha-
sised the importance of inorganic and organic nutrition. However, a majority
of researches from mid-sixties were concentrated on the possibility that a corre-
lative signal from the apex was hormonal in nature (Phillips 1969 ; Kung Woo
?/ al 1974). Though endogenous auxins (Au) was linked with apical dominance
quite early (Wickson and Thimann 1960) and later on confirmed by more workers
(Phillips 1975) the evidences favouring gibberellins (GA) involvement with the same
have been very scanty and flare. In a variety of reports, gibberellins have been
Found to be stimulatory (Brain et al 1959 ; Marth et al 1956 ; Ruddat and
Pharis 1966), inhibitory (Bruinsma and Patil 1963) or indifferent (Sachs and
1964) to the growth of lateral bu4s. As compared to GA? there has been

506 I S Dm, U K Kohli and K S Chark

more convincing evidence linking cytokinin (CK) to apical dominance (Guern
and Usciati 1972 ; Thimann 1972). Unfortunately, most of the above correla-
tions were deduced by studying the growth of lateral buds following exogenous
application of some synthetic or natural regulators and there had been a lack of
effort to estimate quantitatively the endogenous hormones following the evoca-
tion of lateral buds. The present investigations show that lifting of apical
dominance, through growth retardants, coincided with a shift in the balance of
various hormones. The data revealed that in Glycine max this phenomenon is
probably regulated by a ratio of CK: to Au plus GA, The growth retardants
morphactin, opx-1840 or AMO-1618 lifted apical dominance by increasing CK or
decreasing Au or GA respectively. The role of these retardants in successfully
managing these changes was apparent from the studies on the changes of endo-
genous hormones under different treatments. This also draws support from the
work of Ruddat and Pharis (1966) and Baldev et al (1965) on AMO-1618 ; Morcy
and Dahl (1975) and Morey (1974) on DPX-184Q ; and Bednar and Linsmaier-
Bednar (1971), Dua et al (1978), Dua and Dhuria (1980) on morphactin.

The present findings further point that release from the apical dominance and
the follow-up growth of the newly sprouted buds are independent of each other
and probably have their own specific requirements. It was seen that while
inhibitors of auxins or gibberellins lifted the apical dominance by lowering the
denominator in CK/Au + GA ratio, the subsequent growth was impaired owing
to the reduction of the same. It seemed that for this latter growth, GA or An
presence or relatively higher levels were obligatory. This inkling was further
supported from the observation on the treatment where morphactin was in combi-
nation with DPX-1840 or AMO-1618. In this case though the number of buds
released from apical dominance was significantly higher (by virtue of increase in
CK), the follow-up growth of lateral buds was insignificant (probably due to the
non-availability of sufficient Au or GAS).

In essence, it seems that apical dominance in soybean is regulated by a balance
of cytokinins to auxin and gibberellin and any factor which can affect this balance
is likely to influence the manifestation of growth of lateral buds. On tha contrary,
the follow-up growth of newly ensued buds is independent of the above domag
and fo* this the presence of auxin and/or gibberellins seems mandatory.

Acknowledgements

The authors are thankful to Pi?of. K K Nanda for his critical suggestions and
to Late Dr H S Dhuria, Ex-Assistant Director General, ICAR, New Delhi, for
supplying DPX-1840.

References

All A A and Fletcher R A 1970 Xylem differentiation in inhibited cotyledonary buds af soy-
bean ; Can. J. Bot. 48 1139-1141

Baldev B, Lang A and Agatep A O 1965 Gibberellin production in pea seeds by developing
in excised pods ; effect of growth retardant AMO-1618 ; Science 147 155-157

Hormonal regulation of apical dominance 507

Bcdnar T W and Linsmaicr-Bednar R M 1971 Induction of cytokinin tobacoo tissues by

substituted fluorenes ; Proc. Natl. Acad. Sci. USA 68 1178-1179
Bradley V and Crane J C i960 Gibbereffln-induced inhibition of bud development in some

species of Primus ; Science 131 S25-826
Brain P W, Hemming H G and Lowe D 1959 The effect of gibberellic acid on shoot growth

of cupid seed peas ; Physiologia PL 12 15-29
Brooks H J 1964 Responses of pear seed seedlings to N-dimethylamino-succinamic acid, a

growth retardant ; (Nature) (London) 203 1303-1304
Bruinsma J and Patil S S 1963 The effects of 3-indoleacetic acid, gibberellic acid and vitamin E

on flower initiation in unvernaiized Petkus winter rye plants ; Naturwssenschaften 50 505
Dostal R 1926 Uber die wachstumsregulierende Wirkung des Laubblattes ; Act. Soc. Set. Nat.

Moravicae 3 83-209
I>ua I S and Dhuria H S 1980 Role of cytokinins during apical dominance release by morphactin

in Glycine max L. ; Proc. Indian Acad. Sci. (Plant Sci.) B89 375-379
3ua I S and Jandaik C L 1979 Cytokinins in two cultivated edible mushrooms ; Sci. Hortic.

10 301-304
Dua I S, Jindal K K, Srivastava L J, Dinabandhoo C L, Thakur J R and Jain R 1978

Correlation of endogenous cytokinins with apical dominance in response to morphactin

in soybean (Glycine max L.) ; Proc. Indian Acad. Sci. (Plant ScL) B87 319-324
3oebel K 1900 In Organography of plants especially of the Archequoniatae and Spermaphyta.

Part L General Organography (Oxford : Clarendon Press)
Iruern J and Usciati M 1972 The present status of the problem of apical dominance in Hormonal

regulation in plant growth and development (eds) H Kaldewey and Y Vardar (Weinheim :

Verlag Chemie) pp, 383-400

Cung Woo, Lee Kessler B and Thimann K V 1974 The effect of hadacidin on bud develop-
ment and its implication fcr apical dominance ; Physiologia PL 31 11-14
,oeb J 1918 Chemical basis of correlation. I. Production of equal masses of shoots by equal

masses of sister leaves in Bryophyllum calcyinum ; Bot. Gaz. 65 150-174
/Earth P C, Andia W V and Mitchel J W 1956 Effects of gibberellic acid on growth and

development of plants of various genera and species; Bot. Gaz. 118 106-111
flolntyre G I 1968 Nutiitional control of the correlative inhibition between lateral shoots in

the flax seedlings ; Can. J. Bot. 46 147-155

to C L, Choudhury S N, Dattary P and Hafeez U 1962 The influence of light and tempe-
rature on the estimation of auxin by a straight growth test ; Indian J. Plant PhysioL 5

97-116
forey P R 1974 Influence of 3«,3-dihydro-2-(/;-methoxyphenyl)-8H-pyrazolo-(5, 1-a) isoindol-8-

one on xylem formation in honey mesquite ; Weed Sci. 22 6-10
Corey P R and Dahl B E 1975 Histo logical and morphological effects of auxin transport

inhibitors on honey mesquite ; Bot. Gaz. 136 274-280

lorgan P W and Durham J I 1972 Abscission : potentiating action of auxin transport inhi-
bitors ; Plant PhysioL 50 313-318

furakami Y 1966 Bioassay of gibberellins, using rice endosperm and problems of its appli-
cation ; Bot. Mag. (Tokyo) 19 315-366

fitsch J P 1956 Methods for the investigation of natural auxins and growth inhibitors in
The chemistry and mode of action of plant growth substances (eds) R L Wain and
F Wightman (London : Butterworths Scientific Publications) pp. 3-31
>gawa Y 1963 Studies on the condition for gibberellin assay using rice seedlings ; Plant Cell

PhysioL (Tokyo) 4 227-237
>sborne D J and McCalla D R 1961 Rapid bioassay for kinetin and kinin using senescing

leaf tissue ; Plant PhysioL 36 219-221
hillips I D J 1969 Apical dominance ; in Physiology of plant growth and development (ed)

M B Wilkins (London : McGraw-Hill) pp. 163-202
hillips I D J 1975 Apical dominance ; Ann. Rev. Plant PhysioL 26 341-361
uddat M and Pharis R P 1966 Participation of gibberellin in the control of apical domi-
nance in soybean and redwood ; Planta (Berlin) 71 222-228

achs T and Thimann K V 1964 Release of lateral buds from apical dominance ; Nature
(London) 2« 939-940

508 / S Dua, U K Kohli and K S Chark

Schneider G 1970 Morphactins, physiology and performance ; Ann. Rev. Plant Physiol. 21

499-536
fhimann K V 1972 Apical dominance in Hormonal regulation in plant growth and development

(eds) H Kaldeway and V Vardar (Weinheim-Bergstr : Verlag-Chemie) pp. 391-398
Thimann K V, Sachs T and Mathur K N 1971 The mechanism of apical dominance in Coleus ;

Physiologia PL 24 68-72
Thimann K V and Skoog F 1934 On the inhibition of bud development and other functions

of growth substances in Vicia faba ; Proc. R. Soc. B114 317-339
Tognoni F, Heitogh de A A and Wittwer S H 1967 The independent action of morphactin

and gibberellic acid on higher plants ; Plant Cell Physiol. 8 231 -239
Wagner H and Michael G 1971 The influence of varied nitrogen supply on the production

of cytokinins in sunflower roots ; Biocliem. Physiol. Pflanzen. 162 141-158
Wickson M E and Thimann K V 1960 The transport of IAA in pea stems in relation to apical

dominance ; Physiologia PL 13 539-554

Prac. Indian A<Jad. Sci. (Plant Sci.), Vol. 01, Number 6, beceiriber Wttt PP-
© Printed in India.

Taxonomic importance of epidermal characters in the Indian
Thespesia Corr. (Malvaceae)

S RAJA SHANMUKHA RAO and N RAMAYYA*

Department of Botany, Saidar Patel College, Secunderabad 500025, India
* Department of Botany, Osmania University, Hyderabad 500 007, India

MS received 4 December 1981 ; revised 21 October 1982

Abstract In Thespesia lampas and T. populnea, the foliar stomata are anisocytic,
anomocytic and tetracytic, the first type being dominant. Further in Thespesia
lampas and T. populnea altogether eight trichome types are recognisable mostly an
the basis of structure. The two species can be distinguished from each other
by the presence of multiseriate aseptate stellate hair in the former and that of
rnultiseriate aseptate peltate hai, in the latter. T. populnea is also distinct from that
of T. lampas due to curved to wavy epidermal walls, striated surface, absence of
mucilaginous cells on the leaf abaxial. The present evidence also supports treat
mcnt of T. lampas under Thespesia rather than in Hibiscus.

Keywords. Epidermal characters ; taxonomy ; Thespesia ; Malvaceae.

1. Introduction

Taxonomic importance of epidermal characters in general and those of trichomes
in particular in angiospermous plants is now widely recognised (Prat 1932 ;
Tomlinson 1961 ; Stace 1965 ; Ghose and Davis 1973 ; Verhoeven and Schijff
1973). However, previous studies on the epidermis are limited to few taxa and
only to certain aspects in the Malvaceae (Solereder 1908 ; Metcalfe and Chalk
1950 ; Inamdar and Chohan 1969; Ramayya and Shanmukha Rao 1976 ; Shan-
mukha Rao and Ramayya 1977a, b). Hence, the present investigation was under-
taken which deals with foliar epidermal characters along with structure and
organographic distribution of trichomes in whole plant and their taxonomic
importance in the two Indian Thespesia.

2. Material and methods

The material of Thespesia lampas Dalz. & Gibs, was collected from Caramjol,
Goa, whereas T. populnea (L.) Sol. ex Corr. from plants growing at Sardar Patel

509

S Raja Shanmukha Rao and tf

College campus, Secunderabad. Mature trfohomes were, studied either from
epidermal peelings or those isolated by scraping the plant parts. These micro-
preparations were stained either with anilin blue in lactophenol or safranin,
hematoxylin and then mounted in 70% glycerine. Boiling the material with
5-10% glacial acetic acid was useful in obtaining the peels. Microtome sections
of shoot apices were cut at 10-14 jam thickness and stained with hematoxylin
and basic fuchsin. The foot of the different trichomes was determined in free-
hand and microtome sections of various parts of the species investigated. The
walls of the different trichomes were tested with phloroglucin and 2% HC1 for
lignin (Johansen 1940).

The terms describing stomatal types are after Metcalfe and Chalk (1950) and
as redefined by Shanmukha Rao and Ramayya (1977a) and for trichome types,
after Ramayya (1972) and Shanmukha Rao and Ramayya (1977b).

3. Observations

3.1. Structure and distribution of epidermal and stomatal complexes on the leaf

3 . la. Epidermal cell complex : Epidermal cells : 4-6-sided, anisodiametric ;
contents dense, brownish in T. populnea ; sides thin, straight (leaf abaxial and
adaxial of T. lampas and leaf adaxial of T. populnea} or curved to wavy (leaf abaxial
of T. populnea) ; surface smooth (T. lampas) or striated (r. pojpulnea), striations
wavy, prominent and continuous. Distribution : Confined to the interstices,
variously oriented (figures 1-4). Costal cells : 4-sided, linear ; contents scanty
or mostly brownish (T. populnea); sides moderately thick, straight or curved
(leaf abaxial of T. populnea) ; surface smooth (T. lampas) or striated (T. populnea).
Distribution : Oriented parallel to the veins, diffuse. Mucilaginous cells :
Similar to the epidermal cells but enlarged below with opaque contents ; sides
thin, straight or curved ; surface smooth. Distribution : Occasional either in
leaf abaxial (T. lampas) or adaxial (T. lampas and T. populnea) (figures 4 and 10).

3 • lb. Stomatal complex : Stomata mostly anisocytic, occasionally anomocytic
or tetracytic. Subsidiaries 3 or 4, monocyclic unequal, similar to the epidermal
cells except that cuticular striations are absent over stomata. Distribution :
Amphistomatic but adaxially stomata confined to margins of the midrib
(figures 1-6).

Data on the epidermal and stomatal complexes are given in table 1. In both
the species, single guard cells as stomatal abnormality are occasionally observed
in the leaf abaxial whereas they ate totally absent in the adaxial (figure 7).

3.2. Structure and distribution of trichome complex on vegetative and floral parts

Eight trichome types could be recognised in the two species, viz., (i) unicellular
conical Mar, (ii) uniseriate filiform clavate hair, (iii) uniseriate filiform pyrifoum
hair, (iv) multiseriate aseptate peltate hair, (v) multiseriate aseptate stellate hair,
(vi) multiseriate aseptate 4-armed stellate hair, (vii) multiseriate aseptate 3-armed
stellate hair and (viii) biseriate aseptate V-shaped hair. The details of the struc-
ture of the trichome types are as follows :

Taxonomic importance of epidermal characters in Thespesia 5.1 J

Figures 1-11. Epidermis of Thespesia. T. populnea: 1 and 2. Surface views of leaf
abaxial and adaxial respectively (cuticular stiiations not drawn). 7. Surface view
of stomatal abnormality with single guard % ell from leaf abaxial. 10. Mucilaginous
cell from leaf abaxial epidermis in T.S. 11. Oblipue seotion showing sunken
unisoriate filiform clavate hair from T.S. leaf (foot cell not visible). T. lampas :
3 and 4. Surface views of leaf abaxiial and adaxial respectively. 5. Semi-
diagrammatic representation of stomatal distribution confined to margins of midrib
from leaf adaxial. 6. Surface view of anomocytic stoxna from leaf adaxial. 8*
Uniseriate filifoirm clavate hair from T.S. pedicel. 9. Side view of emergence
from pedicel, (me = mucilaginous cell).

P.

512

S Raja Shanmukha Rao and N Ramayya

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Taxonomic importance of epidermal characters in Thespesia 513

8

Figures 12-28. Epidermis of Thespesia. T. populnea : 12. Multiseriate aseptate
peltate hair from leaf abaxial. 13-15. Intermediates between multi seriate aseptate
peltate hair and multiseriate aseptate stellate hair from petal abaxial. 16. Multi-
seriate aseptate stellate hair from petal abaxial. 17. Isolated unicellular conical
hair from sepal adaxial. 27. Uni seriate filiform pyriform hair of noctary from
sepal adaxial. T. lampas : 18. Unicellular conical hair from sepal adaxial.
19, 20. Biseriate V-shaped aseptate hair from sepal adaxial and petal abaxial
respectively. 21, 22. Multiseriate aseptate 3-armed stellate hair from petal
abaxial. 23. Surface view of uniseriate filiform clavate hair from pedicel.
24, 25. Multiseriate aseptate stellate hair from leaf abaxial and pedicel respectively.
26. Multiseriate aseptate 4-armed stellate hair from ovary. 2$.
filiform pyriform hair of nectary from sepal adaxial.

514 S Raja Shanmukha Rao and N Ramayya

3.2a. Unicellular conical hair : Foot consisting of the basal end of the body
cell, indistinct from the body, embedded in the epidermis ; neighbouring cells of
foot annular and striated ; contents absent ; wall thick. Body representing
extension of the foot, conical, apically pointed ; contents absent ; wall thick and
nonlignified ; surface smooth (figures 17 and 18).

3.2b. Uniseriate filiform clavate hair : Foot 1 -celled, square to linear or trape-
zoidal, embedded or projected above the epidermis ; contents absent ; wall thin.
Stalk 1-celled, rectangular to linear ; contents scanty ; wall thin ; surface smooth.
Head clavate, 2-6-tiered, each tier 2-6-oelled ; cells square to linear ; contents
dense; walls -thin ; surface smooth (figures 8, 11 and 23).

3.2c. Uniseriate filiform pyriform hair : Foot 1-celled, linear, embedded or
projected above the epidermis ; contents absent ; wail thin. Stalk 1-celled,
rectangular to lineai ; contents scanty ; wall thin ; surface smooth. Head pyri-
form, multiseriate, 4-10-tiered, each tier 2-6-ceUed, generally terminated by a
pair of rounded cells ; cells rectangular to linear ; contents dense ; walls thin ;
surface smooth (figures 27, 28).

3.2d, Multiseriate aseptate peltate hair: Foot narrow, consisting of the juxta-
posed basal ends of the 20-35-body cells, embedded in the epidermis, polygonal
in outline in peels mounted upside down, occasionally subtended by an emergence ;
contents absent ; walls thin, nonlignified. Body peltate, nearly circular or uneven,
serrate or fcrenulate at margin, 20-35-armed, representing continuation of tho
foot ; arms broadened near the centre of the body but tapering towards distal
end, parallel to the epidermis ; contents brown ; walls thin, nonlignified ; surface
smooth (figure 12). ,

3.2e. Multiseriate aseptate stellate hair : Foot as: in the above. Body stellate, !,

5-40-armed, representing continuation of the foot ; arms tapering towards the |

distal end, unequal in length, generally parallel to the epidermis, some obliquely 1

raised above ; contents absent ; walls thin to moderately thick, nonlignified ;
surface smooth (figures 9, 16, 24 and 25).

Multiseriate aseptate 4-armed stellate hair, multiseriate aseptate 3-armed I

stellate hair knd biseriate aseptate V-shaped hair types are similar to the multi- f

seriate aseptate stellate hair described above except for the difference in number
of arms of the body as indicated by the names of trichome types (figures 19-22
and 26). The details of distribution of the various trichome types are given
in table 2.

4. Discussion

Inamdar arid ChoKan (1969) recorded anisocytic and anomocytic stomata in
Thespesia , populnea^ which is presently confirmed. Further, tetracytic stomata
(as -defined by Shanmukha Rao and Ramayya 1977a) have also been presently
noted, in the leaves of the two species of Thespesia studied. Among the three
stomatal types, the anisocytic is dominant (table 1).

Taxonomic importance of epidermal characters in Thespesia 515

Table 2. Organographic distribution of trichome types in the Indian Thespesia.

Plant part

Thespesia lampas Thespesia populnea

^eaf abaxial

B, E

B, D

>af adaxial

B, E

B, D

^eaf margin

A, E

B, D

'etiole

B, E

B, D

Stipule abaxial

B, E

B, D

Stipule adaxial

B, E

B, D

Stipule margin

B, E

D

Stem

B, E

B, D

teduncle

B. E

B. D

Jracteale abaxial

B, E

B, D

iracteole adaxial

B

B, D

Jracteole margin

B, E

D

tepal abaxi?!

B, D

lepal adaxial

A, C, H

A, C

lepal margin

A

A

>etal abaxial

A, E-H

B, E-H

>etal adaxial

... •

*etal margin

A, E-H

A

Jtaminal tube

...

...

Style

...

...

Dvary

A, E-H

D

V, Unicellular conical hair ; B, Uniscriate filiform clavate hair ; C, Uniseriate filiform pyriform
lair ; D, Multiseriate aseptate peltate hair ; E, Multiseriate aseptate stellate hail ; F, Multiseriate
iseptate 4-arined stellate hair ; G, Muttiseriate aseptate 3-armed stellate hair ; H, Biseriate
iseptate V-shaped hair.

According to Youngman and Pande (1929) the following trichome types occur
n Thespesia : (i) single unbranched hairs, (ii) stellate hairs, (iii) peltate scales
ind (iv) club shaped bodies. The first and third trichome types quoted above
ire the same as the unicellular conical and multiseriate aseptate peltate hair types
•espectively, described by us. On the other hand, we consider the " stellate hair "
ype quoted above, to be a trichome complex rather than representing a specific
;richome type. On comparison with the trichome types delimited in the present
investigation, the stellate hair is resolvable into the following four types : (i) multi-
ieriate aseptate stellate hair, (ii) multiseriate aseptate 4-armed stellate hair,
Jii) multiseriate aseptate 3-armed stellate hair and (iv) biseriate aseptate V-shaped
hair. Similarly the club shaped body is distinguishable into two of the presently
described trichome types : (i) uniseriate filiform clavate hair and (ii) uniseriate
filiform pyriform hair. In the two species of Thespesia investigated thus in all
eight trichome types are presently recognisable as described in the text.

516 S Raja Shanmukha Rao and N Ramayya

Though, in T. lampas, the multiseriate aseptate stellate hair on pedicel and
sepal abaxial are 5 to many armed (figure 25), those on the leaf lamina, petiole
and stem are 20-40-armed with thin walls (figure 24). Likewise in T. populnea
the arms of the multiseriate aseptate peltate hair are 20-35 with thin walls on
all the parts of its occurrence (figure 12). Many intermediate forms connecting
the peltate and stellate types also occur on petals (figures 12-16). The characters
which distinguish the multiseriate aseptate stellate hair of T. lampas and the
multiseriate aseptate peltate hair of T, populnea from each other include the
separate and tapering nature of the arms in the former and the connate condition
and rounded ends of the arms in the latter (figures 12 and 24). As shown by
Ramayya and Shanmukha Rao (1976) the above differences in the two trichome
types are due to early onset of apical intrusive growth instead of symplastic growth
in the development of arms in the multiseriate aseptate stellate hair. Thus the
differences described are deep in origin and hence are significant in justifying
the separation of the two trichome types.

In the two species of Thespesia, all the vegetative parts are trichiferous whereas |

in the floral parts, the surface of the petal adaxial, staminal tube and style are |

non-trichiferous (table 2). Further, in each of the species, all the tiichiferous \

parts show at least two trichome types (viz., multiseriate aseptate peltate hair and [

uniseriate filiform clavate hair in T. populnea and multiseriate aseptate stellate \

hair and uniseriate filiform clavate hair in T. lampas) except the abaxial surface
of the petal and the ovary which possess several trichome types each (table 2).

Out of the eight trichome types now described, all of them occur in T. populnea
whereas in T. lampas there are seven types, the multiseriate aseptate peltate hair I

being absent (table 2). Thus the two species are distinguished from each other f

due to the presence of the multiseriate aseptate peltate hair in the former and j

that of the multiseriate aseptate stellate hair in the latter. Further, the leaves of |

T. populnea are distinct from those of T. lampas by curved epidermal cells with \

striations and absence of mucilaginous cells on the abaxiai surfaces (table 1).

In the systematic accounts, Thespesia lampas (Masters 1874 ; Borssum 1966 ; i

Rakshit and Kundu 1970 ; Saldanha and Nicokon 1976) is considered conspecific £

with Hibiscus lampas (Schumann 1890 ; Hochreutiner 1900 ; Gamble 1957) and I

Azanza lampas (Babu 1977). The tribe Gossypieae (includes Thespesia) differs |

from Hibisceae (includes Hibiscus) by the gossypol glands and conduplicate |

embryos (Fryxell 1968). The presence of gossypol glands on different parts of f

Thespesia lampas, T. populnea and other two species (Standford and Viehoever
1918 ; Lukefahi and Fryxell 1967) supports the separation of Thespesia from I

Hibiscus. Further, Thespesia lampas stands out distinct in other Malvaceae (with I

thick-walled non-lignified stellate hair) by the multiseriate aseptate thin-wailed f

stellate hair, thus providing further evidence to treat Thespesia lampas distinct t

from Hibiscus lampas. |

i

Acknowledgement !

\

One of the authors (SRR) is thankful to. the Principal, Sardar Patel College,
Secunderabad, for providing facilities. • • •

Taxonomic importance of epidermal characters in Thespesia 517

References

Sabu C R 1977 Herbaceous flora of Dehradun (New Delhi : csm) 84

Jorssum Waalkes J Van 1966 Malesian Malvaceae revised ; Blumea 14 1-251

7ryxell P A 1968 A redefinition of the tribe Gossypieae ; Bot. Gaz. 129 296-308

Gamble T S 1957 Flora of the Presidency of Madras (Reprinted ed.) (Calcutta : BSI) Vol. 1 p. 72

jhose M and Davis T A 1973 Stomata and trichomes in leaves of young and adult palms ;

Phytomorphology 23 216-229

lochreutiner B P G 1900 Revision du genera Hibiscus ; Ami. Cons. Jard. Bot. Geneve 4 23-91
namdar J A and Chohan A J 1969 Epidermal structure and stomatal development in some

Malvaceae and Bornbacaceae ; Ann. Bot. 33 865-878

ahansen D A 1940 Plat't microtechnique (New York : McGraw-Hill Book Co., Inc.) 194
^ukefahr M J and Fryxell P A 1967 Content of gossypol in plants belonging to genera i ela-
ted to cotton ; Econ. Bot. 21 128-131
Blasters M T 1874 Malvaceae in The flora of British India (ed) J D Hooker (London : Reeve

and Co.) Vol. 1 184
tfetcalfe C R and Chalk L 1950 Anatomy of the dicotyledons (Oxford : Clarendon Press) Vols.

1 and 2
>rat H 1932 L* Epiderme des Graminees ; Artnales des Sciences Naturelles Botanique et Biologie

Vegetate Series 10 165-258
Lakshit S C and Kundu B C 1970 Revision of the Indian species of Hibiscus ; Bull Bot. Surv.

India 12 151-175
lamayya N 1972 Classification and phylogeny of the trichomes of angiosperms in Research

trends in plant anatomy K A Chowdhury Commemoration Volume (eds) A K M Ghouse

and M Yunus (New Delhi : Tata-McGraw-Hill) 91-102
lamayya N and Shanmukha Rao S R 1976 Morpholo&y phylesis and biology of the peltate

scale stellate and tufted hair in some Malvaceae ; /. Indian Bot. Soc. 55 75-79
laldanha C J and Nicolson D H 1976 Flora of Hassan District Karnataka India (New Delhi ;

Amarind Publishing Company) Vol. 1» ppd 15^-156.
Ichumann K M 1890 Malvaceae In Die natur 'lichen pflanzenfamiUen (eds) H G A Engler and

K A E Prantl 3
Jianmukha Rao S R and Ramayya N 1977a Storoatogonosis in the genus Hibiscus L (Malvaceae) ;

Bot. J. Linn. Soc. 74 47-56
Shanmukha Rao S R and Ramayya N 1977b Structure distribution and taxonomic importance

of trichomes in the Indian species of Malvastrum ; Phytomorphology 27 40-44
olerede' H 1908 Systematic anatomy of the dicotyledons (Oxford : Clarendon Press) Vols. I and 2
•tace C A 1965 The significance of the leaf epidermis in the taxonomy of the Combretaceae I.

A general review of tribal, generic and specific characters ; Bot. J. Linn. Soc. 59 229-252
tanford E E and Viehoever A 1918 Chemistry and histology of the glands of the cotton

ylant with notes on the occurrence of similar glands in related plants ; /. Agric. Res. 13

419-435
'omlinson P B 1961 Anatomy of the monocotyledons. IL Palmae (Oxford : Clarendon Press)

pp. 30-31
rerhoeven R L and Schijff H P V D 1973 A key to the South African Combretaceae based on

anatomical characteristics of leaf ; Phytomorphology 23 65-74

roungman W and Pande S S 1929 The epidermal outgrowths of the genera Thespesia and Gassy-
plum ; Ann. Bot. 43 711-740

toe. tndian Acad. Sci, (Plant Sci.), Vol. 91, INtumW 6, December 1982, pp.
©Printed in India.

Embryological studies in Launaea nudicaulis Hook.*

P S CHIKKANNAIAH and B S HIREMATH

Department of Botany, Karnatak University, Dharwad 580 003, India

MS received 27 February 1981 ; revised 22 September 1982

Abstract. The ovule is anatropous, unitegmic and teruinucellate. Their funicular
vascular strands extend almost to the base of the ovule. A large hypodermal
archespoial cell functions directly as the megaspoje mother cell. It divides to
form a linear tetrad. The upper three megaspores degenerate while the chalazal
develops into an 8-nuckate embryo sac of the Polygonum type. Endothelium diffe-
rentiates at the megaspore tetrad stage. Fertilization is porogamous. Syngamy
and triple fusion take place almost simultaneously. The endosperm is nuclear
but it becomes cellular subsequently. The embryo is the Senetio variation of
Asterad type. Occurrence of polyembryony has been recorded only in one ovule.

Keywords. Launaea nudicaulis ; embryology ; Compositae,

1. Introduction

The Compositae is one of the largest families of flowering plants consisting of
more than 20,000 species, yet the embryological work in the family is scanty.
Davis (1966) has reviewed the previous embryological literature on the family
Compositae. Venkateswarlu and Maheswari Devi (1955a) have made embryo-
logical studies on Launaea nudicaulis, which is widely distributed and also has
cyto types, only a note has appeared so far (see Venkateswarlu 1939).

The cytotypes are diploid (2n = 18) and tetraploid (2n = 36). Attempt is
made here to present some aspects of comparative embryological studies of both
diploid and tetraploid taxa.

2. Material and methods

Young heads were fixed as such but old heads were cut symmetrically into two
halves before they were fixed in formalin-acetic alcohol. Following customary
methods, the material was dehydrated in ethyl alcohol series and embedded in

* Part of the Ph.D. thesis submitted by the Junior author (B S Hiremath) to the Karnatak
University, Dharwad.

519

520

P S Chikkannaiah and B S tiiremath

8 9

Figures 1-10. L.S. of Launaea nudicaulis ovules. 1. Showing anatropous condi-
tion (x 150). 2. Single hypodermal archesproial cell (x 350). 3. Megaspore
mother cell and supernumerary archesporial cells (x 350). 4, 5. Dyad and a
tetrad of megaspares. 6-8. Uni, four and six nucleate embryo sacs (x 350).
9, 10. Organised and mature ernbiyo sacs (x 350). ant, antipodal cells ; pel,
periendothelial zone ; sar, supernumerary archesporial cells ; ser, secondary
nucleus ; syn. synergid cells ; vt9 vascular traces.
Figures 1, 2, 4-7, 9-25 of 2n species. Figures 3, 8, 26 and 27 of 4/i species.

Embryological studies in Launaea nudicaulis Hook. 521

paraffin. Serial sections were cut from 6 to 15 microns and were stained in
Heidenhains iron-alum haematoxylin using sometimes erythrocin in 90% alcohol
as counter stain.

3. Observations

Detailed observations in the diploid taxon and only variations in the tetra-
ploid species are described. The inferior, bicarpellary and unilocular ovary
bears a single ovule at its base. The ovule is anatropous, unitegmic and tenui-
nucellate (figure 1). The funicular vascular strands extend almost to the base
of the ovule. There is generally a large hypodermal archesporial cell which
functions directly as the megaspore mother cell (figure 2). In tetraploid taxon
supernumerary archesporial cells are usually observed (figure 3). Their number
varies from 2 to 8. An archesporial cell undergoes meiotic divisions and forms
a linear tetrad (figures 4, 5). Further development of supernumerary archesporial
cells in tetraploid taxon is not observed. Subsequently, the upper three mega-
spores degenerate while the chalazal megaspore organises into an 8-nucleate
embryo sac of the polygonum type (figures 6, 7, 9). In tetraploid taxon the
divisions at both chalazal and micropylar ends are not synchronous. At the
chalazal end the third mitotic division has already resulted in four nuclei while
at the micropylar end the two nuclei are yet to divide (figure 8).

The organised embryo sac is spindle-shaped and its chalazal end is narrow.
The egg apparatus is at its micropylar and consists of two synergids and a centrally
placed egg cell (figure 9). Synetrgids are hooked (figure 10). There are three
antipodal cells which are placed in triangular or linear fashion in the narrow
chalazal end of the embryo sac. The two polar nuclei are in the centre and are
very near to each other (figure 9). Later the two polar nuclei, just below the
egg apparatus, form a secondary nucleus (figure 10).

Endothelium appears at the megaspore tetrad stage and presents its characteristic
glandular appearance (figure 11). It arises from the inner epidermis of the integu-
ment enclosing the nucellus (figures 7, 9 and 12). It consists of rectangular cells
and the cells are rich in cytoplasm (figure 11). Endothelium nourishes the
growing embryo sac. Gradually it starts losing its compactness (figures 12 to 15).
Its peripheral layer called periendothelial zone loses its cell contents finally forming
an empty layer (figures 13, 14). The endothelium persists as a thin layer around
mature embryo (figure 25).

Fertilization is porogamous. The pollen tube, after reaching the base of the
egg, bursts open at its base discharging the two male gametes. One male gamete
fuses with the egg and the other with the secondary nucleus (figure 12). Syngamy
and triple fusion take place almost simultaneously (figure 12).

The division of the endosperm nucleus precedes that of the zygote. The first
division of the primary endosperm nucleus results in the formation of two free
nuclei. As many as eight free nuclei have been observed around the two celled
proembryo (figure 13). Later it becomes cellular. Hence the endosperm is of
nuclear type. The developing embryo absorbs most of the endosperm tissue §Q

522

P S Chikkannaiah and B S Hiremath

ca

Figures 11-18. L.S. L. nudicaulis nucellus. 11. Endothelium at megaspore
tetrad stage. 12. Syngamy and double fertilization (x 350). 13. 8-nucleate
endosperm (x 350). 14, 15. Stages in the development of end c sperm and endo"
thelium (x 350). 16. Zygote (x 350). 17, 18. 2- and 4-ceUed embryos.

Embryological studies in Launaea nuaicaulis Hook.

523

27

Figures 19-27. L.S. ovules of Ir. nudicaulis. 19-24. Stages of development of
embryo (x 350). 25. L.S. of mature seed (x 20). 26. L.S. of globular embryo
showing long suspensor (x 350). 27. L.S. of seed showing two embryos A and B
(x 20). cot, cotyledon ; cot tr, cotyledonary traces ; de, dermatogen initials ; el9
endothelium ; en, endosperm ; pef periblem initials ; pl9 pleurome initials ; pvt,
stem tip ; re, root cap.

524 P S Chikkannaiah and B S Hiremath

that the mature seed has a thin layer of endosperm surrounding the embryo
(figure 25).

The zygote is pyriform (figure 16). The first division of zygote is transverse
resulting in the formation of a terminal cell ca and a basal cell cb (figure 17).
The next transverse division occurs in cb forming two tiers m and ci of one cell
each (figure 18) while the cell ca divides longitudinally forming two juxtaposed
cells resulting in a T-shaped proembryo. Further, the tier ca divides longitudi-
nally in a plane perpendicular to the first forming a quadrant q (figures
18-20).

Simultaneously, the tier m divides vertically into two cells while the tier ci
divides transversely into two tiers n and ri of one cell each (figures 19, 20).
Generally, the transverse division in the tier ci precedes that of the longi-
tudinal division in the tier m (figure 19). Thus, a proembryo of seven cells
disposed in four tiers is formed (figure 19). The tier ca consists of four cells,
m of one cell, n and ri of one cell each. The walls in the quadrant are placed
obliquely dividing them into central cells a and peripheral cells /J (figures 21, 22).
The peripheral cells j3 are very active and they form two cotyledons (pco) while
the central cells a are less active initially, but contribute to stem apex (pvt) (figures
22, 24). By this time m has undergone another vertical division to form four
circumaxial cells (figures 21, 22). Meanwhile the tier ri has divided transversely
forming two tiers o and p of one cell each (figures 21, 22) The cell p is highly
elongated and vacuolated. The proembryo consists of 14 cells arranged in five
tiers (figure 22). The terminal tier q consists of 6-cells, the tier m of four cells,
the tier n of two cells and the tiers o and p of one cell each. Subsequently the
periblem and pleurome initials are cut off from the tier q (figure 23).

Each one of the four circumaxial cells in the tier m divides periclinally giving
rise to dermatogen initials which become continuous with the dermatogen formed
in the apical tier (figure 23). Likewise, the periblem initials which are differenti-
ated in tier q also extend into tier m. Later, longitudinal and transverse walls
are laid down in this tier m which finally gives rise to hypocotyl (phy) and
radical (ice) (figure 24).

The two juxtaposed cells in the tier n undergo periclinal division cutting off
epidermal initials. The cells in the centre further divide both longitudinally and
transversely to contribute to root tip (iec). The tier o divides periclinally and
anticlinally to form a root cap (co). The tier p contributes to the suspensor (s).
In tetraploid taxon there is a long suspensor (figure 26). It perists till the heart-
shaped embryo stage and is totally absent in the mature embryo. The mature
embryo is dicotyledonous and somewhat straight (figure 25). It consists of an
elongated hypocotyl-root axis. The root apex is protected by the root-cap. The
stem apex is centrally placed between the two cotyledons having abundant
starch. The vascular strands run from the hypocotyl into the cotyledons. The
following sequence represents the development of the embryo of Launaea
nudicaulis.

A rare instance of polyembryony has been observed in tetraploid taxon based
on its position, the adventitious embryo seems to have been originated from one
pf the synergids (figure 27).

Embryoiogical studies in Launaea nudicaulis Hook, 525

Zygote

. ca • q ' — M"

phv + irr

U-f-

n iAr

n -rn

L,r

1

L o $

I. Discussion

ntegumentary vascular traces have been reported in some Compositae (Misra
965 ; Chopra and Singh 1976). In Launaea nudicaulis funicular vascular strands
ire present which extend only to the base of the chalaza. Although pluricellular
.rchesporium has been reported earlier (Harling 195 la ; Sehgal 1966), in Launaea
mdicaulis (2x) there is only one archesporial cell. However, in tetraploid plants
upernumerary archesporial cells are also observed. In most of the Compositae,
L linear tetrad of megaspores is formed in which the chalaza develops into an
dght nucleate embryo sac of the polygonuni type (Pullaiah 1977a, b ; Sharma
md Murthy 1978). T-shaped tetrads have also been reported in many members
Deshpande 1964a, b ; Kaul etal 1975). Invariably in Launaea nudicaulis only
inear tetrad of megaspores is observed.

There are many interesting features in the embryo sac development in the
Uompositae. Several divergences from its normal development occur (Davis
.966). An Allium type in Ammobium spp. (Davis 1962), in Chrysanthemum
pp. and Erigeron spp. (Harling 1951a,b) has bsen described. Three variations
>f the tetrasporic development of the embryo sac, a Fritillaria type in Gaillardia
ricta (Venkateswarlu and Maheswari Devi 1955b), Ratibida tagetes (Howe 1964),
Drusa type in Chrysanthemum spp., Erigeron spp. (Harling 195 la, b) and Adoxa
ype in Rudbeckia hirta (Palm 1934) have been reported. In Launaea nudicaulis
he Polygonum type of embryo sac has been recorded. Large haustorial synergids,
>ften extending into the micropyle, appears to be a common feature in the
Hompositae (Davis 1961b). Pullaiah (1977a, b) recorded hooked synergids in
Volidago canadensis and Achillea squarrosa. In Launaea nudicaulis, hooked
ynergids with blunt ends are observed. There is a great variation in the number
>f antipodal cells and in the number of nuclei in them, a feature found in many
nembers of the Compositae. Secondly, antipodal region has been found to be
laustorial persisting as vermiform appendage (Davis 1961a). Taigi and Taimni
1963) have also noted the haustorial antipodals in Vernonia sinerscens. Murthy
ind Sharma (1976) and Sharma and Murthy (1978) have observed basically three
intipodals in all the three species, Felicia bergariana, Conyza stricta and Erigeron
wnariensis while in Bellia perennis they have noticed an increase in the number
>f antipodal cells up to 21. In Launaea nudicaulis there are three antipodal cells
>laced in a triangular fashion and persist till the mature proembryo stage. But
hey do not show any haustorial nature.

526 P S Chikkannaiah 'and B S tfirertiath

Endothelium is present in the Compositae (Venkateswarlu and Maheswari Devi
1955a; Kaul et al 1975 ; Deshpande and Kothare 1976 ; Chopra and Singh 1976 ;
Sharma and Murthy 1978 ; Sehgal 1979). It appears in the early development of
the ovule at megaspore mother cell stage in Vernonia anthelmintica (Misra 1972)
or at megaspore tetrad stage in Guizotia abyssinica (Chopra and Singh 1976). In
the present study differentiation of the endothelium takes place at the megaspore
tetrad stage. It is usually unfseriate but sometimes biseriate. Nutritive activity
of the endothelium becomes evident during the development of the embryo sac.
The layers of cells surrounding the endothelium show some marked changes,
especially with respect to size, shape, form and stainability. This layer is called
periendotheliai zone (Misra 1972). In the material under study the nutritive
activity is at its peak at the time of fertilization.

Both cellular and nuclear types of endosperm are known to occur in the
Compositae, the former being more frequent (Davis 1966). Endosperm is of
nuclear type in the taxa under study. Persistence of the endosperm as a thin layer
around the embryo is in line with the observations of Misra (1965, 1972) , Kaul
et al (1975) and Sharma and Murthy (1978).

Deshpande (1961) has shown that in Eclipta prostrata obliquely oriented walls
are laid down in the octant which is characteristic of Onagrad type. He, further,
commented that these features constitute intermediate stages of the Onagrad type
and the Asterad type. However, the present study does not indicate any such
variations.

Acknowledgement

The authors thank Prof. M S Chennaveeraiah, Head of the Department of
Botany, Karnatak University, Dharwad, for critically going through the
manuscript.

References

Chopra S and Singh R P 1976 Effect of Gamma rays and 2,4-D en ovule, female gatnetophyte,

seed and fruit development ; Phytomorphology 26 240-249
Davis G L 1961a The life history of Podolepis jaceoides (Sims.) Voss. II. Megasporogcnesis,

female gametophyte and embryogeny ; Phytomorphology 11 206-219
Davis G L 1961b The occurrence of synergid haustoria in Cotula australis (Less) Hook. f.

(Campositae) ; Aust. J. Sd. 24 296
Davis G L 1962 Embryological studies in Australian Compositae. 2. Sporogenesis, gameto-

genesis and embryogeny in Ammobium alatum ; Aust. /. Bot. 10 65-67
Davis G L 1966 Systematic embryology of the angiosperms (New York : John Wiley)
Deshpande P K 1961 Fertilization and development of endosperm, embryo and seed coat in

Eclipta prostrata Linn., Bull Bot. Soc. Coll ScL Nagpur 21-8
Deshpande P K 1964a A contribution to. tir life history of Volutarella racemosa Roxb •

/. Indian Bot. Soc. 43 141-148
Deshpande P K 1964b A contribution to the embryology of Bidens biter ttata ; J Indian Bot

Soc. 43 149-157
Deshpande P K and Kathare M P 1976 Embryology of Cyathodine purpurea ; /. Indian Bot

Soc. 55 205-212

Embryological studies in Launaea nudicaulis Hook. 527

Hading G 195ia Embryological studies in the Compositae. II. Anthemideac-Chrysanthcminac

Acta Horn. Bergiani 16 1-56
Harliflg G 19515 Embiyologtcal studies in the Compositae. in. Astcreae ; Acta Horti.

Bergiani 16 73-120

Howe T D 1964 Developmert of embryo sac in Ratibida tagetes; Am. /. JBot. 51 678
Kaul V, Dathan A S R and Singh D 1975 Embryological studies on the genus Sonchus L. ;

J. Indian Bat. Sac. 54 238-247
Misra S 1965 Floral morphology of the family Compositae. III. Embryology of Siegesbeckia

orientalis L. ; Aunt. J. Bot. 13 1-10
Misra S 1972 Floral morphology of the family Compositae. IV. Vcrnonieae. Vemonia anthel-

mintica ; Bot. Mag. (Tokyo) 85 187-199
Murthy Y S and Sharma H P 1976 Embryol< gical studies in Compositae. II. Bellis perennis

Lima. A casual apomict. Recent trends and contacts between Cyto genetics and Embryo-
logy and morphology. All India UGC sponsored seminar. Nagpur pp. 251-261
Palm B 1934 Bin nour Embryosacktypus bei Rudbeckia hirta ; Bot. Notiser 87 423-427
Pulaiah T 1977a Embryological investigations in Solidago canader.sis Linn. (Compositae)

Astereae ; Proc. Indian Set. Cong. 65
Pullaiah T 1977b Embryology of Achillea squarrosa Ait (Compositae : Anthemideae) ; Proc.

Iiidiun Sci. Cong. 65
Sehgal C B 1966 Morphological and embryological studies on Erigeron bonariensis L.

(E. Uniforms Willd.). ; Beitr. Biol. Pflanz. 42 161-183
Sehgal C B 1979 Infrastructure of integumentary cells in Nicotic.ua rustica L. before and

after fertilization ; /. Cytol. Genet. 14 192-197
Sharma H P and Murthy Y S 1978 Embr>ok>gical studies in the Compcsitae, Astcreae — II.

Proc. Indian Acad. Set. B87 149-156
Tiagi B and Tainnii S 1963 Floral morphology and embryology of Vemonia cinerascens Sohultz.

and Y. cinera Less. ; Agra Univ. J. Res. (Sci.) 12 123-138
Vcnkateswarlu J 1939 A note on the structure and development of the ovule aad the embryo

sac in species of Launaea ; Curr. Sci. 8 556-557
Vcnkateswarlu J and Maheswari Devi H 1955a Embryological studies in the Compositae. I.

Launaea pintiatifida Cass. ; Proc. Indian Acad. Sci. 41 38—46
Venkateswarlu J and Maheswari Devi H 1955b Embryological studies in Compositae. If.

Helcnicae ; Proc. Natl. Inst. S>i. India B21 149-161

P (B)~8

Prac. Indian Acaci. S-si. (Plant Sci.), Vol. 91, Number 6, December 1982, pp. 529-549.
© Printed in India.

Quantitative profile structure of certain forests in the Kumaun
Himalaya

A K SAXENA and J S SINGH

Department of Botany, Kumaun University, Naini Tal 263 002, India

MS received 12 May 1982 : revised 30 September 1982

Abstract. The structure of forests occurring within the north-western catchment
of the river Gola in Kumaun Himalaya is quantitatively described. All the forestt
indicated a total of four strata ; two upper strata represented by trees, the third
stratum represented mainly by shrubs, and the fourth u-f herbs. The tree heighs
of the As (top most) stratum decreased with an increase in altitude. On the other
hand, the proportion of trees devoted to the canopy in the Al and A2 strata increased
with an increase in altitude. In all forests, the crowns of the A-± and A» strata
were more deep than wide. In general, the shrub layer in three oak forests was
comparatively dense and the crowns of the shrubs overlapped with each other. The
canopy index, a relative measiur of canopy coverage, of tree and shrub layers was
maximum for Quercus flonbunda forest and minimum for Pinus roxburghii forest.
Further, the cooler aspects developed a greater canopy index for these layers as
compared to the warmer aspects. Oak forests exhibited a poor development of
their herb layers. The trees in the Quercus lanuginvsa forest weere more stable,
while in Pinus roxburghii forest they were specially susceptible to wind effect. In
general the warmer aspects had more stable trees, while the cooler aspects showed
a lower tree stability. The different forest types, presently studied, could be graded,
as follows, in a decreasing order of potential for soil protection : Quercus flori-
buncLi > Quercus leucotrichophora > Quercus lanuginosa > mixed >P/;.w roxburghii f

Keywords. Himalayan forests ; profile structure ; canopy index.

1. Introduction

The Himalaya offers an array of forest types below the timber line, and is the
cradle of major rivers of India, harbouring (thus) a net work of catchment areas.
Growing human interference with the vegetation cover of the catchment of rivers
has generally led to substantial reduction of forest cover which in turn has led
to serious ecological disasters, such as, soil erosion, loss of soil fertility and cata-
strophic floods. As the catchment efficiency depends on the type, quantity and
stratification of vegetation, a quantitative evaluation of its vegetation is a pre-h
requisite. However, such data are few (Ralhan et at 1982; Saxena and Singh

529

530

A K Saxena and J S Singh

1982; Saxena etal 1982; Tewari and Singh 1981). Earlier studies, mainly quali-
tative, have been reviewed by Puri (1960) and Champion and Seth (1968). The
present study describes the structure of the forests occurring within the north-
western catchment of the river Gola in Kumaun Himalaya.

Due to the presence of a large variety of growth forms, forests are generally
highly stratified (Smith 1974). Tropical forests usually have a total of five
strata above the soil surface, while the temperate forests have only two or three
strata (Richards 1952). Upto some extent, light and moisture determine the
various strata which in turn modify the environment from the canopy to the forest
floor. The amount of light received in various strata of a community varies,
depending upon the density of different strata, type of vegetation, opening of
forest crown, etc. (Knight 1965). The canopy closure also plays a major role
n the regeneration of forests by conditioning the light intensity reaching the forest
floor. According to Richards (1952) and Holdridge (1968), an excellent visual
representation of the structure of a forest community can be communicated
by constructing a profile diagram. Several workers, including Beard (1941, 1955),
Burges and Johnston (1953), Dansereau (1957), Keay (1957), Webb (1959),
Fosberg (1961) and Legris (1961) have recognized the usefulness of profile
diagrams and their structural-functional information. Ashton and Brunig (1975)
and Whitmore (1975) recently reviewed the structural variation in the humid
tropical forests and in the forests of south-east Asia, respectively.

2. Study area and methods

2. 1. Location

A total of 14 sites, located in the north-western catchment of river Gola in
Kumaun Himalaya (29° 19' to 29° 27' N lat. and 79° 32' to 79° 42' E long.)
were selected for the present study. The sites are at different altitude, aspects
ands lope angles (table 1).

Table 1. Aspect, altitude and slope angle of the selected locations.

Forest type

Aspect

Altitude
(m)

Slope angle
(°)

Locality

Pinus
roxburghii

NE, E, SW

1280-1320

30 65

Champ Iii, Sattal

Mixed

E, NW, S

1320-1365

25-50

Sattal

Q. leucotrichophora

NE, E, SW

1950-2025

40-50

Maheshkhan

Q. lanuginosa

S

201C

55-65

Maheshkhan

Q. floribunda

N, NE, E, S

2100-2227

35-55

Maheshkhan

Profile structure of certain forests in Kumaun Himalaya 531

Under the forest settlement of 1911 to 1915, some of the new reserves were
grouped into 6 settlement blocks '. The right and concessions to the villagers
were given whenever possible for building-timber, wood for agricultural implements,
grazing for a limited number of cattle, lopping, collection of fuel, grasses, etc.
In 1921, the new reserve forests were divided into class I and class II forests
and the rules regarding rights were modified. Class I forests retained their
status as reserve forests but their management was under the civil authorities,
while the class II forests were kept under the direct management of the Forest
Department. According to the rules, in class I forests all bonqfied residents of
this region were allowed to graze cattle without any limit, fell and lop trees, cut
grasses, etc., but the felling or lopping of timber trees were rtstricted. In the class
II forests also, all ^onafied residents of this region could graze cattle, lop Quercus
and miscellaneous species, cut grasse and collect fallen fuel woods. The exceptions
were the regeneration areas, fuel and fodder reserves and plantation areas in which
some or all of these concessions were restricted. Timber trees were also reserved
and not allowed to be cut or lopped. In 1964, the management of class I forests,
except in those in which forest Panchayats had been formed, was transferred back
to the Forest Department. The present sites are located in the forests of * new
reserve' category notified after 1915 and represent the original class II forests.
Thus the rules regarding utilization weie liberal and were not related to the carry-
ing capacity of the area. The broad leaved trees, especially Quercus spp., were
frequently lopped. To stop this, in 1974, the Government banned the felling
by public, of all the trees of Quercus leucotrichophor&and Rhododendron arboreum
except for the dead, diseased and oveffsjctture trees.

2. 2. Climate, soil and geology

The sites are characterized by a climate which shows three distinct seasons, viz. ,
rainy (June to September), winter (October to February) and summer (March
to May). The average annual rainfall at Naini Tal is 2820 mm, 88% of which
occurs between June to September. The mean maximum temperature ranges
from 10 -0 to 30'0°C and the mean minimum from 0-2 to 19-8°C.

The soil has been derived from parent materials comprising mainly of quartzite,
quartz porphyry and schists (Raina and Dungrakoti 1975). The soil is dominated
by sand particles. Proportion of sand is lower in oak forests compared to
the mixed and Pinus roxburghii forests. The clay percentage differs little in
different soils but in contrast to the pattern shown by sand, silt percentage is
higher in the oak forests compared to other forests. The soil under all forests
is slightly acidic. Details of climate and soil are given in Saxena and Singh (1980).

2.3. Methods

Sampling was done on four topographic situations (viz., hill base, lower and
upper slopes and hill top) for each aspect (site). Each sample consisted of 12
randomly placed 10 x 10 m quadrats. The size and number of qnadrats were
determined, respectively by species area curve (Costing 1958) and the running
mean method (Kershaw 1973). Detailed phytosociology of the woody species
in these forests is described elsewhere (Saxena and Singh 1982).

532 A K Saxena and J S Singh

In each quadrat, diameter at breast height (dbh at 1 • 37 m from the ground)
of all trees > 10cm dbh was measured and recorded individually by species.
Each quadrat was subdivided into four 5 x 5 m plots for analysing shrubs, saplings
and seedlings. All individuals of 3-3 to 10*0 cm dbh were tallied either as
sapling or shrub, as appropriate. The herb layer was studied through tiller analy-
sis (Singh 1967, 1969) by using 50 x 50 cm harvest plots. The sampling was done
when the herbaceous vegetation was at its peak, i.e., during last week of September
to the first week of October. It is not intended to compare the density values for
herbs with those of the trees, however.

The other measurements included total tree height, height to first branch and
canopy width for each species. A rough sketch of the trees was made in the
field. Height of the tree and canopy depth were measured by a hypsometer
(Forbes 1961).

Profile diagrams were prepared for each topographic situations on each aspect
following Knight (1963). In order to include a maximum number of species
in the diagrams, an area equivalent to 20 x 10 m was found suitable. The number
of trees, saplings and shrubs to be included in the diagram was calculated on
the basis of their density (Curtis and Mclntosh 1950). Trees of each species were
selected for inclusion from among all trees of that species actually measured in
the stand by using a random table (Campbell 1974).

The canopy indexes for tree and shrub layers were calculated by dividing the
sum of the lengths of the strip covered by canopies by the total length of the
profile strip. The resulting value was then multiplied by 100. These calculated
values give only a relative measure of canopy coverage.

The quotient of slenderness (so) for the top canopy trees, in. each stand, was
calculated by the formula given by Brunig and Heuveldop (1976) :

so = h/d9

where, h = average height of the tree (m) ; d = average diameter of tree at breast
height (m).

Spatial distribution, abundance and stratification of vegetation are summarized
in vegetational formulae (Christian and Perry 1953). Letters and figures were
assigned to the trees, shrubs and herbs, to their component layers, and to the
density of each. Thus, Al was used for low trees and X2 for tall trees. The
letters, B and C were used for shrubs and herbs, respectively. Heights were
recorded for each stratum as mean values. Thus, Af represents the trees with
an average height of 20m. Density was sim'larly treated by prefixing x, y or z
for dense, average, or sparse and xx or zz for very dense and very sparse, respec-
tively. Thus, a stand with two tree layers, one shrub and one herb layer at vary-
ing densities is expressed as :

Af z, A^ y, BI-* xx, C°-5. x

This would indicate that the vegetation has two tree layers, one with average
height of 20 m, the other with average height of 10 m, one shrub layer with mean
Height of 1 -2 m and one herb layer with an average of 0- 5 m. Farther, the tall
trees are sparse, low trees have an average density, the shrub layer is very denset
and the herb layer is dense, The ranges for x, y, z\ xx and zz are given in table

Profile structure of certain forests in Kumaun Himalaya 533

Table 2- Ranges of plant density for density symbols used in vegetational formulae.

(Individuals 200 m~2)

Density
Symbols

A,} and AI
strata (trees -)-
saplings)

B stratum
(Shrubs)

C stratum (herbs + seedlings)

XX

81-100

801-1000

279033-347800

X

61-80

601-800

210265-279032

y

41-60

401-600

141497-210264

z

21-40

201-400

72729-141496

22

1-20

1-200

3960-72728

2. The range of the stratal height was set as : stratum A2 = 15 to 30m ;
stratum Al = 2 to 15m and stratum B = 0-5 to 2m.

The herbaceous stratum (C) is not shown in the diagrams although the informa-
tion is included in the vegetational formulae based on the earlier description by
Saxena and Singh (1980).

Results

The profile diagrams for various forests are given in figures 1-5 and the vegeta-
tional formulae in table 3.

3.1. Pinus roxburghii forest

Dn all aspects, the average height of the A2 stratum was higher on the hill top
:ompared to other topographic situations (table 3). The A^ stratum, on all topo-
graphic situations and aspects was represented by trees of P. roxburghii as illus-
trated by the profile diagram of this forest in figure 1. At the SW hill base
md NE lower slope, the Al stratum was represented only by P. roxburghii. On
;he other hand the A^ stratum at the hill base of east aspect consisted of Pyrus
lashia and Cocculus laurifolius, while the same at the upper slope of this aspect
lad Quercus leucotrichophora and Engelhardtia spicata.

The crowns of P. roxburghii in the A2 and Al strata were more deep than
vide, and were conical in shape. The canopy of the Az stratum was compara-
ively denser and almost continuous at the NE hill base, lower slope and hill top,
t the SW hill base and at the E lower slope. On other topographic situations
nd aspects, the canopy in this stratum was markedly broken. The canopy of
lie Al stratum on all topographic situations and aspects, was remarkably dis-
ontinuous. The trees in the A2 and Al strata were very sparse (table 3).

The total combined canopy index of the A^ and A2 strata was highest at the
•IE lower slope and lowest on the E upper slope and hill top (table 4).

A K Saxena and J S Singh

NORTH

SOUTH-WEST,(320

m

Figure 1. Profile diagram for Pinus roxburghii forest on north-cast, east and
south-west aspects. The scale on the .y-axis is for the height and the scale on
*-axis represents width of canopy and dbh. Each profile diagram is made up of
four sections as follows: HB »hill base, LS«lowr slope, us = upper slope,
HT = hilltop. Each section represents an area of 200m2. PR ^ Pinus roxburghii
Sarg., ES - Engdlwrdtia spicata Leschen ex B,. Var. Colebrookiana (LindL ex
Wall.) Ktze, si = Sapium insigne Benth., PP **Pyrus pashia Buch-Ham ex D.Don.,
QLe = Quercus leucotrichophora A. Camus, AC = Adina cordifolia (Roxb.) HK.f!
ex Brandis, CL^Coccutus laurifolius DC., LC^Lantana camara Linn.,jKE^=
Rubus ettipticus Smith., PC ^Pyracantha crenutata (D.Don) Roem., BAS « tierberis
astatica Roxb. ex DC,

Profile structure of certain forests in Kwnaun Himalaya

535

B*

EAST, (320m

SOUTH, 1340m

Figure 2. Profile diagram for mixed forest on east, north-west and south aspects.
For rest of the explanation see figure 1 . The additional species are : BR = JSauhinia
retusa Buch-Ham. ex DC., PO *= Per sea odoralissima (Nees) Kosterm., PI = Pistada
integerrima Stewart, CE = Celtis eriocarpa Decaisne, GO = Grewia oppositifolia.
Buch-Ham. ex Roxb., RT = Rhamnus triqueter Wall., cc = Cedrela ciliata Roem,
PS = Parthenodssus semicordata Wall, ER = Euphorbia royleana Boiss, scu =
Syzygiwn cumini (L.) Skeels., MP == Murray a paniculata (L.) Jack., BVr = JSauhinia
variegata Linn0 BVh = Bauhinia vahlii Wight & Arn., AO = Acer oblongum Wall,
ex DC, MAU = Millettia aumulata Baker ex Brand., MB = Myrica esculenta
Var. sapida Buch-Ham., GS = Grewia subinaequalis DC, CF = Cassia floribunda
car., MK-Murraya koetigii (L.) spren^., AT « AechnicmtMfa tameritote Nees,

A K Saxena and J S Singh

EAs

Figure 3. Profile diagram for Quercus lemotrichophora forest on north-east, east
and south- west aspects. For rest of the explanation see figure 1. The inset
diagram represents mainly the shrub layer magnified from the main diagram for
an area equivalent to 15'7ma. The additional species are:- RA= Rhododendron
arboreum Sm., co = Cornus oblonga Wall., LO == Lyonia ovallfolia (Wall) Drude,
QF = Quercus floribunda Rehder, sch == Symptocos chinensis (Lour) Druce, LQ «
Lonicera quinquelocularis Hardw., CT = Cupressus torulosa D.Don., CD « Cedrus
deodara (Roxb. ex Lambert) D.Don., MAf « Myrsine africana Linn., MS » Myrsine
semiserrata ,WalL, SH« Sarcococca hookeriana Baill., BA! » Boenninghausenia
albiflora (Hook.)" Reiclunb., sv = Smilax vagtnata Decaisne,, SA » Smilax aspera
Linn,, we = Wikstroemia canescens Meissn,, DC « Daphne cannablna Scmu Hook.
ff, BI, =Berberis lycium Royle,

Profile structure of certain forests in Kumaun Himalaya

537

QLO

GLa

SOUTH, 2010m

Figure 4. Profile diagram for Quercus lanuginosa forest on south aspect. For
rest of the explanation see figures 1 and 3. The additional species are : QLa =
Quercus lanuginosa D.Don., vc = Viburnum cotinifolium D.Don,

538

A K Saxena and J S Singh

OF

NORTH, ZIOOm

SOUTH, 2220m

Figure 5. Profile diagram for Quercm floribunda forest on north, north-east,
east and south aspects. For rest of the explanation see figures I and 3. The addi-
tional species is : AF = Arundinaria fakata Nees.

Profile structure of certain forests in Kumaun Himalaya

539

Table 3. Vegetational formulae for the stands examined. For explanation see
text.

Forest type Aspect

Position Vegetation formula

1 2

3 4

Pinus roxburghii North-east

Hill base A22l'Qzz, A^9zz, B^'-zz,

Cy

Lower slope A\im4tzz9 A\'* zz, B1'2zz)

Cy

Upper slope ^i°"szz, Al'Qzz, J5lt0zz,

Cy

Hili top Af'Qzz, A£szz9 B^'^zz,

Czz

East

Hill base A^zz9 A**zz9 B^zz,

Cx

Lower slope A^'^zz, Al'Qzz, B*'5zz9

Cy

Upper slope A%*'Qzz, A^zz, Blu*zz9

Cy

Hill top A?26"Gzz, A*'5zz, Bl'*zz,

Cz

South-west

Hill base 49*°zz, A^^zz, Bl'r>zz,

Cy

Lower slope A^'7zz9 A^'3zz9 B^'^zz,

Cxx

Upper slope A$*'2zz9 A^zz, BL"~zz,

Cx

Hill top A^'*zz9 A*'*zz9 Bl'*zz9

Cxx

Mixed East

Hill base A^^zz, A^zz, BI'Gzz.l

Cxx

Low^r slope AY^zz, A}*'5zz, Br*'*-zz,

Cy

Upper slope A¥'*zz, A7^Qzz, B^'^zz,

Cy

Hill top A\*m*239 Al'szz, B*'~zz,

Cy

North-west

Hill base A\9 A^zz, B^zz,

Cz

Lower slope /i|6'Tzz, A^SZZ, B^zz,

Czz

Upper slope Ar>']*2zz, Al°°zz, BQ>Qzz,

Cz

Hill top A\^zz, Al'Qzz9 B*'°zz,

Cz

South

Hill base A™'szz, A^zz, B*'*zz,

Cy

Lower slope A\**Qzz9 A*'7zz, B^'^zz,

Cz

Upper slope A\**zz, A™'*zzt Bl'&zz,

Cz

Hill top AlQ'*zz, A! zz, B , zz,

Cz

Quercus North-east

Hill base 4c'°zz, A* Gzz, I?1'0*,

Cz

leucotrichophora

Lower slope ^6"8zz, Al'Qz, B°'Dz,

Czz

Upp'.r slope 46<5zz, A»*zz9 B™x,

Czz

Hill top Al5'*zz, A^z, B^z,

• (^zz

East

Hill base A\**zz9 A^zz9 B^"yt

Czz

Lower slope ^Otlzz, ^i'1^ B1"1^,

Czz

Upper slope /^7*2zz, A^z, B°'sy}

Czz

Hilltop A*--*zz9 A*'Qzt B™x9

Czz

South-west

Hill base A%9 Aflz9 B**zz,

Czz

Lower slope A%9 Afzz, B*'Qzz,

Czz

Upper slope A°2, A™z9 B^zz9

Czz

Hilltop 49'9zz, A^z, B°'*zz9

Czz

Quercus South

"Hill Kncf A^Q'&77 A®*® XX B^*^Z
JdLHl DdaO /rig Z.<ij /^j_ ^\-^v» J-* ^.»

Cz

lanugin&sa

Lower slope ^J7"5zz, A^By, J51<8z,

Czz

540

A K Saxena and J S Singh

Upper slope

Al^zz, A***. *•»*, Czz

Hill top

Al^zz, Ai*y, Bl'2zz, Cz

Quercus North Hill base

A^'Qzz, A*'*y, a1'5*, Czz

flonbimda Lower slope

A\, A¥'°y, B^xx, Czz

Upper slope

A¥'*zz, Al'7y, B**z, Czz

Hill top

Al7'*zz, A^y, Bl'2z, Czz

North-east Hill base

A\*l2zz, Al'*xx, B**xx, Czz

Lower slope

A^>*zz9 A(±'lz, Bl'Qy, Czz

Upper slope

AlG'*zz, Af*x, jB1'*^:, Czz

Hill top

A™'\zz, A$*xx9 B™y, Czz

East Hill base

A92, A$*x, Bl'Qy, Czz

Lower slope

A2y AI x, B y, Czz

Upper slope

A™'*zz, A$*xx, B^x, Czz

Hill top

AlQ-*zz, Al*y, Bl'ixx, Czz

South Hill base

A%, Af*xx9 B^z, Czz

Lower slope

>1° /44'7YV W^-'lv C?7

flr>) ^1 **•*> •" y^ \*t+itt

Upper slope

A*>i /ii *C9 JO y^ C/2TIT

Hill top

A^zz, A£*xx, Bf'xx, Czz

The B stratum, being very sparse was not well defined on any topographic
situation and aspect (table 3). Lantana camara was the only occupant of this
stratum on most of the topographic situations and aspects. This stratum had
the highest canopy index at the B hill base and the lowest on the SW hill top
(table 4).

The herb layer in most stands indicated an average density and was composed
of Chrysopogon sermlatus-Desmodium polycarpum community (table 3).

The trees in the A% stratum, on the NE hill top were highly resistant to sway-
ing and bending by wind (so = 52-2) (table 4). On the other hand, tree stabi-
lity was lowest at the SW lower slope (so = 71-6).

3.2. Mixed forest

The profile $tructures of the mixed forest stand are shown in figure 2. On S
and E aspects, the hill base position indicated the highest average height of trees
in AI stratum (table 3). On NW aspect, the maximum average height of the
A% stratum was recorded on the lower slope, while at the hill base this stratum
was altogether absent. Except for the S hill base where Q. leucotrichophora
was the sole occupant of the A2 stratum and the NW upper slope where P. rox~
burghii alone formed the A% stratum, this stratum was constituted by a mixture
of species such as Bauhinia retusa, Persea odoratissima, Celtis eriocarpa, Bauhinia
variegata, Cedrela ciliata, Grewia subinaequalis> etc. The canopy of this stratum
was fairly irregular and discontinuous and trees were very sparse. The crowns

Profile structure of certain forests in Kumaun Himalaya

541

Table 4. Canopy depth, canopy index and quotient of slenderness for different
forests in noith- western part of Gola catchment.

Forest Aspect
type

Position

Average canopy
depth (m)

Canopy index (%) Q

Sl(

uotient of
snderness
(so)

tratum

strata

strata
A± -Mj. B

s
Total

1 2

3

4

5

6

*inus North-east

Hill base

12-7

3-5

128-6

37-6

166-2

57-9

ixbiirghii

Lower slope

10-5

4*4

190-6

14-9

205-5

61-9

Upper slope

9-3

4-4

144-3

40-4

184-7

62-3

Hill top

14-3

3-0

115"/

18-4

134-1

52-2

East

Hill base

13-1

2-0

98-8

40-8

139*6

56-3

Lower slope

12-0

3-1

110-6

10-6

121-2

60-9

Upper slope

14-3

5-9

68-2

13-7

81-9

62-6

Hill top

15-3

3-2

69-4

11-4

80-8

59'8

South-west

Hill base

11-2

3-5

108-6

14-5

123*1

66-4

Lower slope

13-5

3-6

81-6

18-4

100-0

71-6

Upper slope

13-4

2-8

110-2

18-4

128-6

57-6

Hill top

14-4

3-1

155*3

9-4

164-7

65-0

/fixed East

Hill base

13-6

6-0

116-9

256-0

372-9

39-5

Lower slope

9-8

6-2

91-S

151-4

243-2

50-3

Upper slope

11-6

5-1

126-7

397-3

524-0

46-6

Hill top

12-7

5-1

123-5

483-5

607-0

60-2

North-west

Hill base

0

6'3

82-0

16-9

98-9

0

Lower slope

17-3

5-0

169-0

27-5

196-5

48-3

Upper slope

13-4

3-8

100-8

60-8

161-6

57-9

Hill top

13-3

4-8

111*4

154-9

266-3

57-9

South

Hill base

13-6

5-1

147* 8

512-2

660-0

45-4

Lower slope

12*2

5-8

169-8

450-2

620"0

40-6

Upper slope

12-0

6-7

128*2

491-8

620-0

41-6

Hill top

12-0

8-4

139-2

408-2

547-4

56-6

2uercus North-east

Hill base

10-6

4-6

156-5

974-1

1130-6

45-6

mcotricho-

Lower slope

11-2

3-7

206*3

394-1

600-8

47-9

thorn

Upper slope

11-0

4-0

191-8

760-0

951-8

44*4

Hill top

10-4

4-6

167-0

398-8

565-8

47-7

East

Hill base

12-3

3-3

230-6

807-5

1038*1

41*3

Lower slope

13-1

4-6

335*7

896-0

1231-7

47*6

Upper slope

11-3

5-3

328-2

630-6

958-8

44-8

Hill top

15-0

2-6

168-6

1020-4

1189-0

42*5

South-west

Hill base

0

4-2

131-4

165-9

297-3

0

Lower slope

0

4-2

161-2

151*4

312-6

0

Upper slope

0

4-2

144-7

208-6

353-3

0

Hill top

13-6

3*2

142-0

116-5

258-5

49-6

542 A K Saxena and J S Singh

Table 4. (Contd.)

Qucrcus South

Hill base

14-

3

2-

6

333-

7

418-8

752-

5

30-2

lanuginosu

Lower slope

12-

7

3-

4

218-

8

368-6

587-

4

30-5

Upper slope

12-

7

2-

5

209-

0

255-3

464"

3

30-8

Hill top

17-

8

2-

8

182'

4

218-8

4CI*

2

35-7

Qucrcus North

Hill bast;

14-

1

4-

1

277-

3

777-6

1054-

9

52-8

fljribunda

Lower slope

0

3-

0

160-

8

344-7

1505"

5

0

Upper slope

14-

6

6-

0

350-

6

360-0

710-

6

58'0

Hill top

13-

5

4-

3

214-

9

316-0

530"

9

69"3

North-east

Hill base

15-

0

2-

8

237-

3

1165-5

1402-

S

42-3

Lower slope

13-

0

4-

6

176-

0

673*7

849-

7

56-0

Upper slope

13-

2

3-

4

260-

4

868-6

1129-

0

56-1

Hill top

14-

0

3-

4

255-

7

539-6

795"

3

56-2

East

Hill base

0

3-

9

205-

5

608-6

814-

1

0

Lower slope

0

4-

9

241-

6

556- 1

797-

7

0

Upper slope

13-

0

4-

9

320-

8

833-3

1154-

1

58'7

Hill top

13-

6

4-

7

275-

3

814-1

1089-

4

59-4

South

Hill base

0

3-

6

280-

0

402-7

682'

7

0

Lower slope

0

3-

8

247-

5

600-0

847'

5

0

Upper slope

0

4-

1

224-

7

896-5

1121-

2

0

Hill top

12'

•8

3-

4

256'

0

1574-0

1830'

0

53-0

of trees were usually deeper than wide. On S and E aspects the maximum
average canopy depth occurred at the hill base and on the NW aspect at the
lower slope (table 4).

The trees in the A± stratum, were also very sparse (table 3). However, the
canopy was comparatively better developed at the NW hill base than on other
topographic situations and aspects. Almost all trees in this stratum were young
individuals of species which reach the Az stratum upon maturity. Like the A^
stratum, the crowns tended to be more deep than, wide on all topographic situa-
tions and aspects.

On S and NW aspects, the canopy index (A% + Al strata) was highest at the
lower slope, while on E aspect it was highest on the upper slope (table 4).

With the exception of NW aspect, the B stratum was well developed with almost
a continuous canopy. This stratum was dominated by L. camara, which despite
a low density developed a spreading, close canopy, except for NW hill top where
it was dominated by Aechmanthera tomentosa. The canopy index of the shrub
layer was maximum at the S hill base and minimum at the NW hill base (table 4).

The herb layer on most of the positions of NW and S aspect was sparse, while
on E aspect it indicated an average density (table 3). This layer was composed
of Dicliptera bupleuroides—Oplismenus burmanii community.

The trees of the A2 stratum were more slender on the S and E hill tops
0-.2, respectively), ..and,. .on. thp JtfW .uppejc

Profile structure of certain forests in Kumaun Himalaya 543

(so = 57-9) (table 4). These results indicate lower stability of trees at these
positions, hence they are more susceptible to wind damage.

3.3. Quercus leucotrichophora forzst

The canopy in the A2 stratum was almost continuous at the E hill base, lower
slope and upper slope and at the NE lower slope (figure 3). At the SW hill base
and lower and upper slopes, the /42 stratum was absent. Where present, it was
composed of only Q. leucotrichophora with the exception of the E upper slope,
where one tree of P. roxburghii (per 200 m2) occurred with Q. leucotrichophora.
The crowns of the trees were more deep than wide in this stratum. The canopy
depth was maximum on the E hill top and minimum on the NE hill top (table 4).
Stratum A± was dominated by Q. leucotrichophora on all positions of E and
NE aspect (figure 3). On the other hand this stratum on all the four situations
of SW was dominated either by Myrica esculenta, Rhododendron arboreum or by
Lyonia ovalifolia. The E hill top had one tree of P. roxburghii per 200 m2. The
canopy in this stratum was more or less continuous on almost all positions and
aspects. Most of the gaps in the A% stratum were closed by trees in the stratum
A i, thus, strata A% and A± together formed a good cover on all positions and
aspects. The crowns in this stratum were more deep than wide.

The trees on all positions and aspects in the A2 stratum were very sparse,
while on most of the positions and aspects in AI stratum they were sparse (table 3).

The shrub stratum (B) was well defined on all positions and aspects (figure 3).
With the exception of NE and E upper slope, all other positions on different
aspects were dominated by the shrub Myrsine africana. On the E and NE upper
slope Boenninghausenia albiflora was the dominant shrub. The shrub layer on
all positions of SW was very sparse, while on the NE lower slope and hill top
the same was sparse (table 3). This layer on the E lower slope and hill top and
on the NE hill base and upper slope was dense. The canopy index of this stratum
was maximum on the E hill top (table 4),

On most topographic situations and aspects, the plants in the herb layer were
very sparse (table 3). This layer was comprised of Arundinella nepalensis—
Carex nubigena community.

Tree stability was lowest on the SW hill top (so =49.6) where only two trees
of Q. leucotrichophora per 200m2 occurred (table 4).

3 . 4. Quercus lanuginosa forest

At the S hill base and upper slope, the canopy of the A% stratum was fairly
dense and at places the crowns touched each other (figure 4). On the other hand
at the lower slope and hill top positions, canopies were broken ; only two trees
of Q. lanuginosa were present per 200 m3 on each of these positions. This
stratum consisted of only Q. lanuginosa trees on all positions. The crowns were
deeper than wide. The average canopy depth, was maximum at the hill base
(table 4).

The canopy of the A: stratum was more dense as compared to that of the
AS stratum on all positions (table 3). Though, this stratum was dominated
by Q. lanuginosa, other species such as R. arboreum^ M. esculenta, L. ovalifolia,
P, (B)— 9

544 A K Saxena and J S Singh

Cornus oblonga, etc., were also present. Like A2 stratum, the crowns of this
stratum were also deeper than wide.

The canopy index of A2 +• A2 strata was maximum at the hill base and
minimum on the hill top (table 4).

The plants in the stratum B were very sparse on the upper slope and hill top
positions, and sparse at the hill base and lower slope positions (table 3). The
dominant shrub on all positions was M. africana. The shrub canopy index was
maximum at the hill base and minimum on the hill top (table 4).

The herb layer on the hill base and hill top positions was sparse and on the
lower and upper slopes very sparse (table 3). This layer was dominated by the
Apluda mutica—Themeda anathera community.

The quotient of slenderness for the A2 stratum trees was highest on the hill
top and lowest at the hill base (table 4).

3 . 5. Quercus floribunda forest

The profile diagrams for the Quercus floribunda forest stands are illustrated in
figure 5. The A*, stratum was well defined only at the N hill base and upper
slope. The trees of this stratum on all positions and aspects belonged to
Q. floribunda.

The crowns of the A% stratum tended to be deeper than wide. The average
canopy depth was the highest at the NE hill base and the lowest on the S hill
top (table 4).

The stratum Al had a remarkably dense canopy, the individual crowns usually
touched each other. A majority of trees in this stratum was represented by young
individuals of Q. floribunda. Other species in this stratum were : R. arboreum*
L. ovalifolia, C, oblonga, Pyrus pashia, Q. leucotrichophora, etc. The crowns were
deeper than wide.

The trees in the A% stratum on all positions and aspects were very sparse,
while in /4: stratum the trees on most positions and aspects were either dense or
very dense (table 3). The canopy index of the upper two strata (A% H- A()
was the highest on the N upper slope and the lowest at the N lower slope (table 4)*

Below the two storeys of trees, the stratum B consisting chiefly of shrubs was
well defined in this forest. The plants were very dense at the NE hill base, N
lower slope and on the E and S hill top (table 3). On all S positions, at the
E hill base and at the N lower slope, M. africana was the dominant shrub while
M. 'semiserrata played the vicariant role on the N hill top. On rest of the posi-
tions and aspects Arundinaria falcata showed its dominance. The canopy index
of this stratum was on the whole higher than that of the corresponding stratum
in other forests of the study area (table 4).

The herb layer on all positions and aspects was very sparse (table 3) and was
composed of Muehlenbergia duthieana-Helictotrichon asperum community.

The tree stability in A2 stratum was the highest at the NE hill base (so = 42*3)
and the lowest on the N hill top (so = 69' 3) (table 4).

Profile structure of certain forests in Kumaun Himalaya 545

f. Discussion

fhe structure of the forests varied from stand to stand. Such inter-
tand variations in tree stature, crown geometry and canopy architecture are
;ommon (Anderson 1961 ; Ashton 1964 ; Brunig 1970, 1976 ; Brunig and
leuveldop 1976).

In these forests there was a total of four strata ; two upper strata represented
>y trees, the third stratum represented mainly by shrubs, and the fourth by herbs,
rhe maximum average tree height (across positions and aspects) in the A» stratum
vas recorded for P. roxburghii forest (22 -4m) "and the minimum for Q. flori-
wnda forest (17.1 m). Q. leucotrichophora and Q. lanuginosa forests had almost
;qual average tree height (about 18 -0 m) in this stratum. Further, with increasing
Jtitude the tree height of the X2 stratum decreased (Y = 27-3008— 0-0047 X ;

= —0-6799, P<0-001 ; where 7 = tree height in m and X = altitude in m).
Jrown (1919) suggested that the decrease in plant height with increasing altitude
5 due to the combined effects of decreased temperature and decreased illumina-
ion (due to increased cloudiness). Richards (1952) pointed out that this dwarf-
ng of the vegetation may be partly due to exposure to strong wind.

In all forests, the crowns of the A2 stratum were deeper than wide. On an
.verage, across positions and aspects, about 80% length of the trees in Q.flori-
unda, 72% in Q. lanuginosa, 68% in Q. leucotrichophora^ 67% in mixed and
.bout 57% in P. roxburghii forests was covered by the canopy. Contrary to
he tree height, the proportion of the tree devoted to canopy in the A^ stratum
ticreased with an increase in the altitude (Y = 36-6628 + 0-0185 X ; r = 0.8395,
3 < 0"001 ; where Y = percent length of the tree devoted to canopy and X =
Ititude in m). Thus the tree compensated for decrease in height by allocating
more of its length to development of photosynthetic canopy.

The canopies in the A^ stratum were comparatively denser than those in A^
tratum in all forests except those of P. roxburghii. The plants on most positions
,nd aspects in AI stratum were very sparse in P. roxburghii and mixed forests^
parse in Q. leucotrichophora forest, average in Q. lanuginosa forest and dense
o very dense in Q. floribunda forest. Most of the individuals in this stratum
Belonged to the species which constituted the A2 stratum. However, some other
rees such as, R. arboreum, L. ovalifolia, C. oblonga9 S. insigne, M. esculenta^
7. laurifolius. Rhamnus triqueter, etc., were confined only to this stratum.

The average tree height (across positions and aspects) AI stratum was the
jghest in the mixed forest (8 *9 m) and the lowest in Q. lanuginosa forest (4' 1 m).
n this stratum also, almost all trees in all forest types had their crowns deeper
bian wide. As the altitude increased, the percent of tree height covered by canopy
Iso increased (Y = 45-2850 + 0-0134^ ; r = 0-806, P< O'OOl ; where Y =
icrcent length of the tree devoted to canopy and X = altitude in m) ; maximum
78 • 8%) being in the Q. floribunda forest and minimum (62 • 2%) in the P. roxburghii
orest. The proportion of the tree devoted to canopy depth was greater in the A^
tratum compared to the AI stratum.

There was usually no clear vertical discontinuity between the canopies of A^
,nd AI strata because of the occurrence of a variable number of layers in each
The exceptions were tlje U hill b^se and hill top $nd SW upper slope of

546 A K Saxena and J S Singh

P. roxburghii forest, where a clear vertical discontinuity between the A*± and Al
strata occurred.

The shrub density on most positions and aspects in Q. floribunda forest was
average, while it was very sparse in P. roxburghii and mixed forests. In Q. leuco-
trichophora forest, SW aspect exhibited very sparse density, while on most positions
of NE and E, the plants were dense. In general, the shrub layer (B stratum-
in three oak forests was comparatively dense and the crowns of the shrubs over)
lapped each other. In the P. roxburghii forest, this stratum was not well deve.
loped.

The canopy index, a crude and relative measure of canopy cover, of both tree
and shrub layers was maximum for Q. floribunda forest and minimum for P. rox-
burghii forest. On categorizing the aspects into cooler (N, NE, E, and NW)
and warmer (S and SW), the cooler aspects developed a greater canopy index for
tree (X = 183-7%) and shrub (X = 431-5%) layers as compared to the warmer
aspects (canopy index for tree layer, X = 1 70 * 4% and for shrub layer, X = 318-9%).

About 50% of the stands each in P. roxburghii and mixed forests had, respec-
tively, an average and sparse plant density in the herb layer, while in Q. kuco-
trichophora and Q. floribunda forests the herbaceous plants were very sparse. The
plants in this layer were sparse to very sparse in g- lanuginosa forest. Poor
development of herbaceous plants under the oak forests may be because of a
tendency of inverse relationship between the canopy cover (of tree + shrub layers)
and the development of the herb layer (Richards 1952; Smith 1956 ; Zobel
et al 1976 ; Killingbeck and Wall 1978), Naturally, relatively open overhead
canopies would induce the development of the herb layer.

The data on the quotient of slenderness (so) indicates the stability of trees ;
the lower the SG value the higher is the stability (Brunig and Heuveldop 1976).
In the present area the trees in Q. lanuginosa forest were more stable, while in
the P. roxburghii forest trees were specially susceptible to wind effect. Since
P. roxburghii is a rich source of resin, trees are tapped heavily in this region
(Saxena 1977). As the tapping reduces the diameter, the resistence of the heavily
tapped trees to wind is greatly reduced (Assmann 1970). Because of the charac-
teristic low wind stability the tapping of this species for resin should be very
cautious and perhaps should be avoided in those forests which are exposed to
greater wind velocities or having poor stocking density.

A majority of trees in the present forests were more wind resistant as compared
to those in the humid tropical forests (Brunig and Heuveldop 1976). It may be
pointed out chat a mountainous country, due to varied nature of slopes, is charac-
terized by a higher degree of wind turbulence compared to a non-mountainous
region, and therefore, lower values of quotient of slenderness in the present
trees may be an adaptational feature.

In general the warmer aspects had more stable trees, i.e., lower so value
(X = 50-5), while the cooler aspects showed lower tree stability, Le.9 high SG value
(X = 55*1). In comparison to cooler aspects, the warmer aspects experience
greater wind velocities (Smith 1974). The low SG values of trees, thus growing
on warmer aspects may be an adaptation^! feature,

Profile structure of certain forests in Kumaun Himalaya 547

The canopies in the different strata also influence the soil condition of a site.
Packer 1951) observed that overland flow and erosion decrease with increase In
cover. A dense cover of vegetation is the most powerful weapon for reducing
erosion. According to Lull (1964), the drops that drip from the leaves are gene-
rally larger than the rain drops and their terminal velocity is reached by the
time they have fallen 7- 5m. Trimble and Weitzman '1954) concluded that a
high tree canopy has a limited value in reducing the erosion potential of rainfall
intensity, but a forest with a canopy that reaches close to the ground can effec-
tively reduce the erosion potential. Thus a site with trees confined only to the
A% stratum and having their canopies concentrated on the top will be relatively
less protective for the soil. But when the trees in the A2 stratum are supported
by deep and dense canopies in A± or B strata, the vegetation becomes more protec-
tive for the soil. A forest with a multilayered canopy with a high canopy index
and a well developed forest floor will, thus, have a greater protective value as com-
pared to a forest which has fewer layers and a lower canopy index (Kittredge 1948).

In this region, the rainfall is concentrated in a short monsoon period (June
to September). This is preceded by a long dry period (winter and summer
seasons) during which the herbaceous cover dries up and shatters. Additionally
during this dry period, grazing, herbage removal for animal feed and ground fires,
particularly in pine forests further decimate the herb-litter cover, leaving often a
semibare floor. At the culmination of this dry period, the monsoon breaks with
a high rainfall intensity. Under such situations, if the tree canopy is thin or
high or monolayered with little or no shrubby undergrowth, soil erosion and run-
off are remarkably accelerated. Keeping the above in mind the present forest
could be graded in a decreasing order of potential for soil protection : Q. flori-
bunda > Q. leucotric'iophora > Q. lanuginosa > mixed > P. roxburghii. In ' sensi-
tive catchment areas maintained for soil and water conservation, Q. floribunda
and Q. leucotrlchophora forests should, therefore, be encouraged.

Acknowledgement

The financial support from Indian Space Research Organisation, Bangalore is
gratefully acknowledged. Dr Uma Pandey and Mr O P Chaturvedi assisted
in the preparation of figures.

References

Anderson J A R 1961 The ecology and forest types of the peat swamp forests of Sarawak
and Brunei in relation to their silviculture. Ph.D. Thesis, Univ. of Edinburgh, Scotland

Ashtoa P S 1964 Ecological studies in the mixed Dipterocarp forests of the Brunei State.
Oxford Forestry Memoirs* 25, (Oxford : Clarendon Press)

Ashton P S and Brunig E F 1975 The variation of tropical moist forest in relation to environ-
mental factors as key to ecologically oriented land-use planning. Background— paper, FAO
— T. C.IT. M. F.9 Environment, item 3.3 Publ in Mitt. Bundesforsch. anst. f. Forst-Und
Holzwirtschaft, Reinbek, 109 59-86

Assmann E 1970 The principles of forest yield study. (Oxford : Perganion Press), p 59

geard J S 1941 Montane vegetation of the Antilles. Carib. For, 3 1-7Q

548 A K Saxena and J S Singh

Beard J S 1955 The classification of tropical America vegetation types. Ecology 36 89-100

Brown W H 1919 Vegetation of Philippine mountains. Manila.

Brunig E F 1970 Stand structure, physiognomy and environmental factors in some low land

forests in Sarawak. Trop. EcoL 11 26-43
Brunig E F 1976 Ecological studies in the Kerangas forests of Sarawak and Brunei. Borneo

Lit. Bur, Kuching
Brunig E F and Heuveldop J 1976 Structure and functk is in natural and man-made forests

in the humid tropics. XVI IUFRO World Congress. Procee dings- Refer at e-Exposes.

Norway, pp. 500-511
Surges A aud Johnston R D 1953 The structure of a New South Wales subtropical rain

forest. /. EcoL 41 72-83

Campbell R C 1974 Statistics for biologists (Cambridge University Press). 385 p
Champion H G and Seth S K 1968 A revised survey of the forest types of India.

(Delhi : Government of India Publications) p 404
Christian C S and Perry R A 1953 The systematic description of plant communities by the

use of symbols. J. EcoL 41 100-105

Curtis J T and Mclntosh R P 1950 The interrelations of certain analytic and synthetic
phytosociological characters. Ecology 31 434-455

Dansereau P 1957 Biogeography : an Ecological Perspective. (New York : Ronald Press)

Forbes R D 1961 Forestry hand book. (New York : Ronald Press)

Fosberg F R 1961 A classification of vegetation for general purposes. Trop. EcoL 2 1-28

Holdridge L R 1968 Classification and characterisation of tropical forest vegetation. R Misra
and B Gapal (eds) In Proceedings of the symposium on recent advances in Tropical Eco-
logy : Part II : (Faridabad : Today and Tomorrow's Printers and Publishers) pp 502-507

Keay R W J 1957 Wind-dispersed species in a Nigerian forest. /. EcoL 45 471-478

Kershaw K R 1973 Quantitative and dynamic plant ecology. (London : The English language
book society and Edward Arnold Ltd,,) 308 p

Killingbeck K T and Wali M K 1978 Analysis of a North Dakota gallery forest : nutrient,
trace element and productivity relations. Oikos 30 29-60

Kittredge J 1948 forest influences (New York : McGraw-Hill Book Company, Inc.)

Knight C B 1965 Basic concepts of Ecology. (New Yoik : The Macmillan Company) 468 p

Knight D H 1963 A distance method for constructing forest profile diagrams and obtaining
structural data. Trop. EcoL 4 89-94'

Legris P 1961 Forest classification. Trop. EcoL 2 85-88

Lull H W 1964 Ecological and silvicultural aspects, section 6. V T Chow (ed.) In Hand
book of applied hydrology. (New York : McGraw-Hill Book Company, Inc.), pp. 6-1 to
6-30

Costing H J 1958 The study of plant communities (San Francisco: W H Freeman and Company)

Packer P E 1951 An approach to watershed protection criteria. J. For. 49 639-644

Puri G S 1960 Indian Forest Ecology. Vol. I (New Delhi : Oxford Book and Stationery Co.),
318 p

Raina B N and Dungrakoti B D 1975 Geology of the area between NainiTal and Champa wat
Kumaun Himalaya, Uttar Pradesh, (eds) A G Jhingran and P K Verma In Himalayan
Geology, Vol. 5 (Delhi : Wadia Institute of Himalayan Geology) pp 1-27

Ralhan P K, Saxena A K and Singh J S 1982 Analyses of forest vegetation at and around
Naini Tal in Kumaun Himalaya Proc. Indian Natl. Sci. Acad. B 48 121-137

Richards P W 1952 The tropical rain forest. (Cambridge, Cambridge University Press) 450 p

Saxena A K 1977 On resin tapping of a Pirns roxburghii forest in Naini Tal. Uttarakhand
Bharati 2 59-69

Saxena A K, Pandey P and Singh J S 1982 Biological spectrum and other structural-func-
tional attributes of the vegetation of Kumaun Himalaya. Vegetatio 49 111-119

Saxena A K and Singh J S 1980 Analyses of forest grazing and vegetation in parts of Kumaun
Himalaya. Indian J. Range Mgmt. 1 13-32

Profile structure of certain forests in Kumaun tiimalayct 549

Saxena A K and Singh J S 1982 A phytasociological analyses of woody species in forest
communities of a part of Kumaun Himalaya. Vegetatio 50 3-22

Singh J S 1967 Seasonal variation in composition plant biomass and net community production in
the grassland at Varanasi. Ph.D. Thesis, Banaras Hindu University, Varanasi, India. 318 p

Singh J S 1969 Influence of biotic disturbance on the preponderance and interspecific associa-
tion of two common forbs in the grasslands at Yaranasi, India. Trap. Ecol 10 59-71

Smith R L 1956 An evaluation of conifer plantations, as wildlife habitat. Ph.D. dissertation
(Ithaca New \ork : Cornell University)

Smith R L 1974 Ecology and field biology. (New Yoik : Harper and Row Publishers, Inc.)
p 850

Tewari J C and Singh S P 1981 Vegetation analysis of a for;st lying in transitional zone
between lower and upper Himalayan moist temperate forest, (ed) G. S. Paliwal
In The vegetational wealth of Himalayas. (Delhi : Puja publishers) pp. 104-119

Trimble G R Jr and Weitzman S 1954 Effect of a hardwood forest canopy on rainfall inten-
sities. Trarts. Am. Geophys. Union 35 226-234

Webb L J 1959 A physiognomic classification of Australian rain forests. /. Ecol. 47 551-570

Whitmore T C 1975 Tropical rain forests of the Far-East (Oxford : Clarendon Press) p 282

Zobel D B, McKee A, Hawk G M and Dyrness C T 1976 Relationships of environment to
composition structure, and diversity of forest communities of the Central Western Cascades
of Oregon. EcoL Monogr. 46 135-156

Profile structure of certain forests in Kumaun Himalaya 543

(so == 57-9) (table 4). These results indicate lower stability of trees at these
positions, hence they are more susceptible to wind damage.

3« 3. Qiiercus leucotrichophora for^t

The canopy in the A» stratum was almost continuous at the E hill base, lower
slope and upper slope and at the N£ lower slope (figure 3). At the SW hill base
and lower and upper slopes, the A$ stratum was absent. Where present, it was
composed of only Q. leucotrichophora with the exception of the E upper slope,
where one tree of P. roxburghii (per 200 m2) occurred with g. leucotrichophora.
The crowns of the trees were more deep than wide in this stratum. The canopy
depth was maximum on the E hill top and minimum on the NE hill top (table 4).
Stratum AI was dominated by Q. leucotrichophora on all positions of E and
NE aspect (figure 3). On the other hand this stratum on all the four situations
of SW was dominated either by My r lea esculent a, Rhododendron arbor eum or by
Lyonia ovalifolia. The E hill top had one tree of P. roxburghii per 200 m2. The
canopy in this stratum was more or less continuous on almost all positions and
aspects. Most of the gaps in the A2 stratum were closed by trees in the stratum
A i, thus, strata A% and AI together formed a good cover on all positions and
aspects. The crowns in this stratum were more deep than wide.

The trees on all positions and aspects in the Az stratum were very sparse,
while on most of the positions and aspects in AI stratum they were sparse (table 3).

The shrub stratum (B) was well defined on all positions and aspects (figure 3).
With the exception of NE and E upper slope, all other positions on different
aspects were dominated by the shrub Myrsine africana. On the E and NE upper
slope Boennlnghausenia albiflora was the dominant shrub. The shrub layer on
all positions of SW was very sparse, while on the NE lower slope and hill top
the same was sparse (table 3). This layer on the E lower slope and hill top and
on the NE hill base and upper slope was dense. The canopy index of this stratum
was maximum on the E hill top (table 4).

On most topographic situations and aspects, the plants in the herb layer were
very sparse (table 3). This layer was comprised of Arundinella nepalensis—
Carex nubigena community.

Tree stability was lowest on the SW hill top (so = 49.6) where only two trees
of Q. leucotrichophora per 200 m2 occurred (table 4).

3 . 4. Quercus lanuginosa forest

At the S hill base and upper slope, the canopy of the A^ stratum was fairly
dense and at places the crowns touched each other (figure 4). On the other hand
at the lower slope and hill top positions, canopies were broken ; only two trees
of Q. lanuginosa were present per 200 m3 on each of these positions. This
stratum consisted of only Q. lanuginosa trees on all positions. The crowns were
deeper than wide. The average canopy depth was maximum at the hill base

(table 4).

The canopy of the A3 stratum was more dense as compared to that of the
A» stratum on all positions (table 3). Though, this stratum was dominated
by Q. lanuginosa, other species such as R. arboreum, M. esculenta, L. ovalifolia,

P. (B)— 9

544 A K Saxena and J S Singh

Cornus oblonga, etc., were also present. Like A% stratum, the crowns of this
stratum were also deeper than wide.

The canopy index of A2 -|- AI strata was maximum at the hill base and
minimum on the hill top (table 4).

The plants in the stratum B were very sparse on the upper slope and hill top
positions, and sparse at the hill base and lower slope positions (table 3). The
dominant shrub on all positions was M. africana. The shrub canopy index was
maximum at the hill base and minimum on the hill top (table 4).

The herb layer on the hill base and hill top positions was sparse and on the
lower and upper slopes very sparse (table 3). This layer was dominated by the
Apluda mutica — Themeda anathera community.

The quotient of slenderness for the A% stratum trees was highest on the hill
top and lowest at the hill base (table 4).

3 . 5. Quercus floribunda forest

The profile diagrams for the Quercus floribunda forest stands are illustrated in
figure 5. The A± stratum was well defined only at the N hill base and upper
slope. The trees of this stratum on all positions and aspects belonged to
Q. floribunda.

The crowns of the A2 stratum tended to be deeper than wide. The average
canopy depth was the highest at the NE hill base and the lowest on the S hill
top (table 4).

The stratum Al had a remarkably dense canopy, the individual crowns usually
touched each other. A majority of trees in this stratum was represented by young
individuals of Q. floribunda. Other species in this stratum were : jR. arboreum,
L. ovalifolia, C. oblonga, Pyrus pashia, Q. leucotrichophora, etc. The crowns were
deeper than wide.

The trees in the A2 stratum on all positions and aspects were very sparse,
while in AI stratum the trees on most positions and aspects were either dense or
very dense (table 3). The canopy index of the upper two strata (A% + AL)
was the highest on the N upper slope and the lowest at the N lower slope (table 4).

Below the two storeys of trees, the stratum B consisting chiefly of shrubs was
well defined in this forest. The plants were very dense at the NE hill base, N
lower slope and on the E and S hill top (table 3). On all S positions, at the
E hill base and at the N lower slope, M. africana was the dominant shrub while
M. semiserrata played the vicariant role on the N hill top. On rest of the posi-
tions and aspects Arundinaria falcata showed its dominance. The canopy index
of this stratum was on the whole higher than that of the corresponding stratum
in other forests of the study area (table 4).

The herb layer on all positions and aspects was very sparse (table 3) and was
composed of Muehleribergia duthieana-Helictotrichon asperum community.

The tree stability in A2 stratum was the highest at the NE hill base (so = 42 -3)
and the lowest on the N hill top (so = 69*3) (table 4).

Profile structure of certain forests in Kwnaun Himalaya 545

4. Discussion

The structure of the forests varied from stand to stand. Such inter-
stand variations in tree stature, crown geometry and canopy architecture are
common (Anderson 1961 ; Ashton 1964 ; Brunig 1970, 1976 ; Brunig and
Heuveldop 1976).

In these forests there was a total of four strata ; two upper strata represented
by trees, the third stratum represented mainly by shrubs, and the fourth by herbs.
The maximum average tree height (across positions and aspects) in the A» stratum
was recorded for P. roxburghii forest (22 -4m) and the minimum for Q. flori-
bunda forest (17. 1 m). g. leucotrichophora and Q. lanuginosa forests had almost
equal average tree height (about 18-0 m) in this stratum. Further, with increasing
altitude the tree height of the A$ stratum decreased (Y = 27-3008— 0-0047 X ;
r = —0-6799, P<0'001 ; where V = tree height in m and X = altitude in m).
Brown (1919) suggested that the decrease in plant height with increasing altitude
is due to the combined effects of decreased temperature and decreased illumina-
tion (due to increased cloudiness). Richards (1952) pointed out that this dwarf-
ing of the vegetation may be partly due to exposure to strong wind.

In all forests, the crowns of the A^ stratum were deeper than wide. On an
average, across positions and aspects, about 80% length of the trees in Q.flori-
bunda, 72% in Q. lanuginosa, 68% in Q. leucotrichophora, 67% in mixed and
about 57% in P. roxburghii forests was covered by the canopy. Contrary to
the tree height, the proportion of the tree devoted to canopy in the A> stratum
increased with an increase in the altitude (Y = 36-6628 + 0*0185 X ; r = 0.83955
P < 0*001 ; where Y = percent length of the tree devoted to canopy and X =
altitude in m). Thus the tree compensated for decrease in height by allocating
more of its length to development of photosynthetic canopy.

The canopies in the A^ stratum were comparatively denser than those in A2
stratum in all forests except those of P. roxburghii. The plants on most positions
and aspects in At stratum were very sparse in P. roxburghii and mixed forests^
sparse in Q. leucotrichophora forest, average in Q. lanuginosa forest and dense
to very dense in g. floribunda forest. Most of the individuals in this stratum
belonged to the species which constituted the A% stratum. However, some other
trees such as, R. arboreum, L< ovalifolia, C. oblonga, S. insigne, M. esculenta^
C. laurifolius. Rhamnus triqueter, etc., were confined only to this stratum.

The average tree height (across positions and aspects) Al stratum was the
highest in the mixed forest (8 -9 m) and the lowest in Q. lanuginosa forest (4* 1 m).
In this stratum also, almost all trees in all forest types had their crowns deeper
than wide. As the altitude increased, the percent of tree height covered by canopy
also increased (Y = 45*2850 + 0-0134 X ; r = 0-806, P < 0-001 ; where Y =
percent length of the tree devoted to canopy and X = altitude in m) ; maximum
(78*8%)being in the Q. floribunda forest and minimum (62 -2%) in the P. roxburghii
forest. The proportion of the tree devoted to canopy depth was greater in the A^
stratum compared to the AI stratum.

There was usually no clear vertical discontinuity between the canopies of A2
and AI strata because of the occurrence of a variable number of layers in each
Stratum. The exceptions were the E hill base and hill top and SW upper slope of

546 A K Saxena and J S Singh

P. roxburghii forest, where a clear vertical discontinuity between the A* and A^
strata occurred.

The shrub density on most positions and aspects in Q. floribunda forest was
average, while it was very sparse in P. roxburghii and mixed forests. In Q. leuco-
trichophora forest, SW aspect exhibited very sparse density, while on most positions
of NE and E, the plants were dense. In. general, the shrub layer (B stratum,
in three oak forests was comparatively dense and the crowns of the shrubs over)
lapped each other. In the P. roxburghii forest, this stratum was not well deve.
loped.

The canopy index, a crude and relative measure of canopy cover, of both tree
and shrub layers was maximum for Q. floribunda forest and minimum for P. rox~
burghil forest. On categorizing the aspects into cooler (N, NE, E, and NW)
and warmer (S and SW), the cooler aspects developed a greater canopy index for
tree (X = 183-7%) and shrub (T= 431 -5%) layers as compared to_ the warmer
aspects (canopy index for tree layer, J =170-4% and for shrub layer, X = 31 8 -9%).

About 50% of the stands each in P. roxburghii and mixed forests had, respec-
tively, an average and sparse plant density in the herb layer, while in Q. Icuco-
trichophora and Q. floribunda forests the herbaceous plants were very sparse. The
plants in this layer were sparse to very sparse in Q. lanuginosa forest. Poor
development of herbaceous plants under the oak forests may be because of a
tendency of inverse relationship between the canopy cover (of tree + shrub layers)
and the development of the herb layer (Richards 1952; Smith 1956 ; Zobel
et al 1976 ; Killingbeck and Wali 1978). Naturally, relatively open overhead
canopies would induce the development of the herb layer.

The data on the quotient of slenderness (so) indicates the stability of trees ;
the lower the so value the higher is the stability (Brunig and Heuveldop 1976).
In the present area the trees in Q. lanuginosa forest were more stable, while in
the P. roxburghii forest trees were specially susceptible to wind effect. Since
P. roxburghii is a rich source of resin, trees are tapped heavily in this region
(Saxena 1977). As the tapping reduces the diameter, the resistence of the heavily
tapped trees to wind is greatly reduced (Assmann 1970). Because of the charac-
teristic low wind stability the tapping of this species for resin should be very
cautious and perhaps should be avoided in those forests which are exposed to
greater wind velocities or having poor stocking density.

A majority of trees in the present forests were more wind resistant as compared
to those in the humid tropical forests (Brunig and Heuveldop 1976). It may be
pointed out chat a mountainous country, due to varied nature of slopes, is charac-
terized by a higher degree of wind turbulence compared to a non-mountainous
region, and therefore, lower values of quotient of slenderness in the present
trees may be an adaptational feature.

In general the warmer aspects had more stable trees, i.e., lower so value
(X = 50*5), while the cooler aspects showed lower tree stability, i.e., high so value
(jf = 55'l). In comparison to cooler aspects, the warmer aspects experience
greater wind velocities (Smith 1974). The low so values of trees, thus growing
on warmer aspects may be an adaptational feature,

Profile structure of certain forests in Kumaim Himalaya 547

The canopies in the different strata also influence the soil condition of a site.
Packer 1951) observed that overland flow and erosion decrease with increase in
cover. A dense cover of vegetation is the most powerful weapon for reducing
erosion. According to Lull (1964), the drops that drip from the leaves are gene,
rally larger than the rain drops and their terminal velocity is reached by the
time they have fallen 7*5m. Trimble and Weitzman '1954) concluded that a
high tree canopy has a limited value in reducing the erosion potential of rainfall
intensity, but a forest with a canopy that reaches close to the ground can effec-
tively reduce the erosion potential. Thus a site with trees confined only to the
A2 stratum and having their canopies concentrated on the top will be relatively
less protective for the soil. But when the trees in the A2 stratum are supported
by deep and dense canopies in A^ or B strata, the vegetation becomes more protec-
tive for the soil. A forest with a multilayered canopy with a high canopy index
and a well developed forest floor will, thus, have a greater protective value as com-
pared to a forest which has fewer layers and a lower canopy index (Kittredge 1948X

In this region, the rainfall is concentrated in a short monsoon period (June
to September). This is preceded by a long dry period (winter and summer
seasons) during which the herbaceous cover dries up and shatters. Additionally
during this dry period, grazing, herbage removal for animal feed and ground fires,
particularly in pine forests further decimate the herb-litter cover, leaving often a
semibare floor. At the culmination of this dry period, the monsoon breaks with
a high rainfall intensity. Under such situations, if the tree canopy is thin or
high or monolayered with little or no shrubby undergrowth, soil erosion and run-
off are remarkably accelerated. Keeping the above in mind the present forest
could be graded in a decreasing order of potential for soil protection : Q. flori-
bunda > Q. leucotriciophora > Q. lanuginosa > mixed > P. roxburghii. In sensi-
tive catchment areas maintained for soil and water conservation, Q. floribunda
and g. leucotrichophora forests should, therefore, be encouraged.

Acknowledgement

The financial support from Indian Space Research Organisation, Bangalore is
gratefully acknowledged. Dr Uma Pandey and Mr O P Chaturvedi assisted
in the preparation of figures.

References

Anderson J A R 1961 The ecology and forest types of the peat swamp forests of Sarawak
and Brunei in relation to their silviculture. Ph.D. Thesis, Univ. of Edinburgh, Scotland

Ashtdu I* S 1964 Ecological studies in the mixed Dipterocarp forests of the Brunei State.
Oxford Forestry Memoirs, 25, (Oxford : Clarendon Press)

Ashton P S and Brunig E F 1975 The variation of tropical moist forest in relation to environ-
mental factors as key to ecologically oriented land-use planning. Background— paper, FAO
— T. C.IT. M. F.9 Environment, item 3.3 Publ. in Mitt. Bundesforsch. anst. f. Forst-Und
Holzwirtschaft, Reinbek, 109 59-86 -

Assmann E 1970 The principles of forest yield study. .(Qxfcrd : Pergamon Press), p 59

geard J S 1941 Montane vegetation of the Antilles. Carib. For. 3 1-70

548 A K Saxena and J S Singh

Beard J S 1955 The classification of tropical America vegetation types. Ecology 36 89-100

Brown W H 1919 Vegetation of Philippine mountains. Manila.

Brunig E F J970 Stand structure, physiognomy and environmental factoi-'S in some low land

forests in Sarawak. Trap. EcoL 11 26-43
Brunig E F 1976 Ecological studies in the Kerangas forests of Sarawak and Brunei. Borneo

Lit. Bur, Kuching

Brunig E F and Heuveldop J 1976 Structure and functk is in natural and man-made forests
in the humid tropics. XVI IUFRO World Congress. Proceedtogs-Referate-Exposes.
Norway, pp. 500-511
Binges A aud Johnston R D 1953 The structure of a New South Wales subtropical rain

forest. J. EcoL 41 72-83

Campbell R C 1974 Statistics for biologists (Cambridge University Press). 385 p
Champion H G and Seth S K 1968 A revised survey of the forest type* of India.

(Delhi : Government of India Publications) P 404
Christian C S and Perry R A 1953 The systematic description of plant communities by the

use of symbols. /. EcoL 41 100-105
Curtis J T and Mclntosh R P 1950 The interrelations of certain analytic and synthetic

phytosociological characters. Ecology 31 434-455

Dansereau P 1957 Biogeography : an Ecological Perspective. (New York : Ronald Press)
Forbes R D 1961 Forestry hand book. (New York : Ronald Press)

Fosberg F R 1961 A classification of vegetation for general purposes. Trop. EcoL 2 1-28
Holdridge L R 196& Classification and characterisation of tropical forest vegetation. R Misra
and B Gopal (eds) In Proceedings of the symposium on recent advances in Tropical Eco-
logy : Part II : (Faridabad : Today and Tomorrow's Printers and Publishers) pp 502-507
Keay R W J 1957 Wind-dispersed species in a Nigerian forest. J. EcoL 45 471-478
Kershaw K R 1973 Quantitative and dynamic plant ecology. (London : The English language

book society and Edward Arnold Ltd.,) 308 p
Killingbeck K T and Wali M K 1978 Analysis of a North Dakota gallery forest : nutrient,

trace element and productivity relations. Oikos 30 29-60

Kittredge J 1948 forest influences (New York : McGraw-Hill Book Company, Inc.)
Knight C B 1965 Basic concepts of Ecology. (New Yoik : The Macmillan Company) 468 p
Knight D H 1963 A distance method for constructing forest profile diagrams and obtaining

structural data. Trop. EcoL 4 89-94
Legris P 1961 Forest classification. Trop. EcoL 2 85-88

Lull H W 1964 Ecological and silvicultural aspects, section 6. V T Chow (ed») In Hand
book of applied hydrology. (New York : McGraw-Hill Book Company, Inc.), pp. 6-1 to
6-30

Costing H J 1958 The study of plant communities (San Francisco : W H Freeman and Company)
Packer P E 1951 An approach to watershed protection criteria. /, For. 49 639-644
Puri G S 1960 Indian Forest Ecology. Vol. I (New Delhi : Oxford Book and Stationery Co.),

318 p

Raina B N and Dungrakoti B D 1975 Geology of the area between Nairn Tal and Champawat
Kumaun Himalaya, Uttar Pradesh, (eds) A G Jhingran and P K Verma In Himalayan
Geology, Vol. 5 (Delhi : Wadia Institute of Himalayan Geology) pp 1-27
Ralhan P K, Saxena A K and Singh J S 1982 Analyses of forest vegetation at and around

Naini Tal in Kumaun Himalaya Proc. Indian NatL Set. Acad. B 48 121-137
Richards P W 1952 The tropical rainforest. (Cambridge, Cambridge University Press) 450 p
Saxena A K 1977 On resin tapping of a Pinus roxburghii forest in Naini Tal. Uttarakhand

Bharati 2 59-69

Saxena A K, Pandey P and Singh J S 1982 Biological spectrum and other structural-func-
tional attributes of the vegetation of Kumaun Himalaya. Vegetatio 49 111-11^
Saxena A K and Singh J S 1980 Analyses of forest grazing and vegetation in parts of Kumaun
Himalaya. Indian J. Range Mgmt. 1 13-3?

Profile structure of certain forests in Kumaun Himalaya 549

Saxena A K and Singh J S 1982 A phytosociological analyses of woody species in forest
communities of a part of Kumaun Himalaya. Vegetatio 50 3-22

Singh J S 1967 Seasonal variation in composition plant biomass and net community production in
the grassland at Varanasi. Ph.D. Thesis, Banaras Hindu University, Varanasi, India. 318 p

Singh J S 1969 Influence of biotic disturbance on the preponderance and interspecific associa-
tion of two common forbs in the grasslands at Varanasi, India. Trap. EcoL 10 59-71

Smith R L 1956 An evaluation of conifer plantations, as wildlife habitat. Ph.D. dissertation
(Ithaca New 'Vork : Cornell University)

Smith R L 1974 Ecology and field biology. (New Yoik : Harper and Row Publishers, Inc.)
p 850

Tewari J C and Singh S P 1981 Vegetation analysis of a for:st lying in transitional zone
between lower and upper Himalayan moist temperate forest, (ed) G. S. Paliwal
In The vegetational wealth of Himalayas. (Delhi : Puja publishers) pp. 104-119

Trimble G R Jr and Weitzman S 1954 Effect of a hardwood forest canopy on rainfall inten-
sities. Trans. Am. Geophys. Union 35 226-234

Webb L J 1959 A physiognomic classification of Australian rain forests. /. EcoL 47 551-570

Whitmore T C 1975 Tropical rain forests of the Far-East (Oxford : Clarendon Press) p 282

Zobel D B, McKee A, Hawk G M and Dyrness C T 1976 Relationships of environment to
composition structure, and diversity of forest communities of the Central Western Cascades
of Oregon. EcoL Monogr. 46 135-156

?roc. Indian Acad, Sci. (Plant ScL), Vol. 91, Number S, December 19S2«, pp. 551-599
© Printed in India.

Contributions to our knowledge of Indian algae-IIL Buglenmeae-Part-I.

Fhe genus Euglena Ehrenberg*

M T PHILIPOSE

Nenagh Cottage, Coonoor 643101, Tamilnadu, India

MS received 25 May 1981

Abstract. An account of 29 taxa comprising 24 species and five varieties of the
genus Euglena collected from various localities in north-east, central and south
India during 1937-76 is given. Of these, two, viz., E. vaginicola and E.pseudo-
ehrenbergii have been considered as new species, one, viz., E. viridis var. maxima
a new variety and one, viz., E. tuba var. pseudotuba f. minima a new form of a
new combination variety. Four species, viz., E. vagans, E. hdicoideus, E. granulata
and E. hemichromata and three varieties, viz., E. tripteris var. klebsii, E. oxyuris
var. playfairii and E. caudata var. minor appear to be new resords for the Indian
region.

By studying the taxa from different ecological habitats it is shown that there
is considerable variation within species in E. acus and E. tuba and to a limited
extent in E. oxyuris. Since E. ehrenbergii as known at present is a composite species,
E. srinagari has been separated from it, following Huber-Pestalozzi in this respect.

E. tuba Carter which had been incompletely known so far and considered by
most authors as doubtful, was studied in detail and shown to be a well defined
species needing an emended description. E. tuba Johnson, which shows some
essential differences from Carter's species, is treated as a variety of E. tuba, viz.,
var. pseudotuba with an Indian form which is new. There is also a possibility of
polymorphism in E. tuba.

Meteorological and water conditions under which some cf the species dominated
are given. It is also shown that some species aic characteristic of particular habitats.

The need to give all relevant details including proper illustrations while describing
the taxa of this difficult genus is stressed. The author has made an attempt to fill
the gaps in existing descriptions of the Indian taxa givun in this paper to help in
proper identifications. A key to these taxa is also given.

A list of 29 other species reported from India by othei workers is given with
references and localities.

Keywords. Chromatophores; flagellum; haematochrome; paramylum; gullet; moat.
. Introduction

'hough, there have been records of species of Euglena from the Indian region
comprising India, Pakistan, Afghanistan, Nepal, Bangladesh, Burma and

First presented at the Symposium on " Fifty Years of Botany " at the Golden Jubilee
klebrations of the Univ. Bot. Lab,, Madras, during 28 Dec., 1980-2 Jan., 1981.

551
.(B)-40

552 M T

Sri (Lanka) from 1856 onwards, our information on the genus from this region is
still quite meagre. The earliest record of Euglena from the region appears to be
that of Carter (1856) who described E. agilis from Bombay. This species is
better known as E. pisciformis Klebs since Carter's description was considered
inadequate. He also recorded four other species, viz. E. acus, E. spirogyra
E. deses and E. viridis during the same year along with a fifth one E. texta (Duj.)
Huebner under the name of Crumenula texta Dujardin. In 1858 Carter again
referred to the occurrence of E. viridis and E. acus in his collections from Bombay.
The same author's (1859) E. fusiformis and E. zonalis are actually Lepodnclis
fusiformis (Carter) Lemm. and L. ovum (Ehr). Lemm. respectively.

In 1869, Carter described an interesting species, E. tuba, from Bombay. Since
his description was inadequate and the connection between the free living organism
and the characteristic encysted individual was not clearly established, subsequent
authors (see Kent 1881 ; Gojdics 1953 ; Pringsheim 1956) expressed doubts about
its real identity, the latter two authors considering it a& probably a E. sanguined.
Hansgirg (1908) recorded E. agilis again from Igatpuri, Bombay. Kashyap (1908)
reported a Woom of Euglena resembling E. tuba from Lahore, to which Walton
(1915) gave the name of E. orientalis. Since Kashyap's description was incomplete
and without accompanying figures, the real identity of his species is unknown.

Bhatia (1930) described four species of Euglena and a fifth under the name of
Amblyophis srinagari sp. nov., from Kashmir. Banerji (1936) reported E. viridis
from Lower Bengal. Skvortzov (1937) recorded four speci.es from Rangoon.
Philipose (1940) gave accounts of eleven species from Museum Pond, Madras, and
Gonzalves and Joshi (1943, 1943a, 1946) listed and illustrated five species from
Bombay. Skuja (1949) reported seven species and two varieties from Burma.
Biswas (1949) stated that E. viridis and E. sanguinea are quite common in Indian
and Burmese ponds. Suxena (1955) described four and Zafar (1959) recorded
three species from Hyderabad, Philipose (1960) enumerated a number of algae
including species of Euglena occurring commonly in Indian inland fishery waters
while Singh (1960) recorded two species from Uttar Pradesh.

lyengar (1962) described a new species, viz. E. pringsheimii in which there are
" inner " pyrenoids. He also emended the description of E. oblonga Schmitz
after detailed study of material from Madras.

Kamat (1961-62, 1963, 1964, 1967, 1968, 1968a, 1974 and 1975) and Kamat
and Frietas (1976) recorded twenty-seven species (including a new one) and a
number of varieties from Maharashtra, Gujarat, Rajasthan and Himachal Pradesh,
while Naidu (1962, 1966) gave accounts of nineteen taxa covering sixteen species
from Andhra Pradesh. HortoMgyi (1969) described seven species and two
varieties from three reservoirs on the banks of River Jamuna. Suxena et al (1973)
recorded three species from Kerala. Dodkundi et al (1973) observed six species
in a pond at Dharwar. Pandhol and Grover (1976) refer to one species from
Ludhiana. Hosmani (1977) and Hosmani and Bharati (1975) reported blooms
of E. sanguinea and E. elastica respectively from Dharwar. Venkateswarlu
(1976) observed four species in River Moosi, Hyderabad.

With the overlapping in various records, the total number of species (excluding
varieties and synonyms) so far recorded from the Indian region come to about
forty-seven. In spite of this sizeable number, including descriptions of some, it

the genus Euglena Ehrenberg 553

cannot be considered that due* attention has been given to the genus from the
Indian region.

According to Pringsheim (1956) quite a large number of species of Euglena have
been described inadequately or illustrated unsatisfactorily with the result that these
species cannot be recognised or they have to be treated as synonyms of well-known
species. Even records attributed to well-known species cannot always be considered
accurate when unaccompanied by important details. This, he states, is parti-
cularly applicable to groups other than " Rigidae " and " Lentiferae " (see under
Systematic Account) especially species of the " Sanguinea " group. Records from
the Indian subcontinent are no exception to this, quite a number of records being
limited to lists or with incomplete descriptions and figures. This is particularly
true of most taxonomic records other than those of Bhatia (1930) ; Skvortzov
(1937) ; Skuja (1949) ; Suxena (1955) ; lyengar (1962) and Hortobagyi (1969).
It has, therefore, been considered necessary to give full details of the taxa of this
difficult genus encountered by the author in a wide variety of habitats in India,
inlcuding considerable variation within species in several instances. Species of
Lepocindis, Phacus, Tmchelomonas and Strombomonas from these habitats will
be dealt with in later communications.

In this paper twenty-nine taxa belonging to twenty-four species, including those
from Museum Pond, Madras (Philipose 1940) are given making the total number
of species for the subcontinent to about fifty-three. These were collected during
year round ecological studies of some water bodies or during general surveys of
inland fishery waters of north-east, central and south India during 1937-76.

For a better understanding of the taxa discussed here, a key to the species and
varieties is given at the outset, following the classification into main groups
proposed by Pringsheim (1956). The class name Euglenineae as given by Fritsch
(1935), is adopted in preference to Euglenophyceae and Euglenoidina used by
some authors.

2. Localities and dates of collection

Plankton collections were made from a number of States covering north-east and
south India and a few from central India. Most of the collections were from
general surveys conducted by the staff of the Central Inland Fisheries Research
Institute, Barrackpore and Cuttack, including the author, and the rest from the
year round observations at weekly or fortnightly intervals by the author. The
water bodies involved were mostly fish ponds (nurseries of about 0' 04-0' 15 ha
and 1-li metres deep) and rearing and stocking tanks (about 0*2 to 0*5 ha or
more and li to 3 meters deep), some multi-purpose public tanks and small reservoirs,
shallow fishery bunds (used for fish breeding) a few swamps and moats and
two rivers (one of them in a polluted area) under lacustrine conditions. The
nursery ponds were invariably kept free of macro-flora while the rest had frequently
macro-vegetation at their margins.

2. la. Assam : Sibsagar : (1) Sibsagar Jamuna (8-6-51) ; Joyasagar :^Fish farm
(2) N.P. and (3) S.T. (8-6-51) ; (4) S.T. (6-4-55 and 31-5-55) ; (5) N.P. 2 and
(6) N.P. 14 and 18 (1/9-6-55) ; (7) N.P. 30 (15-11-65) ; (8) N.P. 38 (1-12-65) ;

554 Af T Ph&ipose

(9) N.P. 22 (15/16-3-66) ; (10) N.P. 32 (15-3-66) ; (11) N.P. 25 and (12) N.P.
27 (19-3-66) ; (13) N.P. 38 (21-3-66) ; (14) Gaurisagar tank (10-12-65) ;
(15) Beliaghat tank (16-3-66); (16) Teok: Rajabari tank (8-1-66) ; Jorhat :
(17) Municipal tank (25-10-59) ; Nazira : (18) S.D.O's tank and (19) S.T. 16
(6-4-55) ; Dibragarh : (20) S.T. 1 (26-5-55).

2.1b. West Bengal'. 24 Parganas : (21) Museum Pond, Calcutta (March 49-
Feb. 50) ; Dum Dum : (22) Mukherjee's Pond (4-10-49) ; (23) Dr. (Mrs.)
Ghosh's Pond (5-6-'50) ; Belgharia Fish Farm : (24) S.T. 11 and (25) S.T. 16
(23-5-50) ; (26) S.T. 10 (20-6-50) ; Kamarhati (27) Sagore Dutt Pond-1 (23-5-50) ;
Barrackpore : (28) Dhar's Pond (Mar. '49-Feb/50) ; (29) Central Fisheries
Res. Stn. Pond (Mar. '49-Feb.'50) ; (30) Lai Dighi's Pond, Sadar Bazaar (21-1-'50)
(31) Nundi's Pond (18-1- '50) ; (32) Palta Exptl. Filter bed (Mar. ,49-Feb/50) ;
Hooghly : (33) Choudhry Bagan pond, Serampore (Mar/ 49-Feb.'50) ;
Midnapore : Chandrakona Road : (34) Rangamati bund ; (35) Poddar bund
and (36) Kachahari bund (16/17-12-52).

2.1c. Bihar : Sone : (37) River Dehri at Darihat and Mahadewar Ghat down-
stream of polluted area (21-5-53) ; Patna : (38) Jail pond (Jui '49).

2. Id. Madhya Pradesh : Bhopal Fish Farm : (39) N.P.I and N.P. 25 (19-7-54)
(40) Nitora tank (20-7-54) ; (41) Tanks, Jetiose and (42) Nishatputa (24-7-54) ;
Raipur : (43) Turki nursery (21-4-56); Jabalpur : (44) Gangasagar (23-4-56).

2.1e. Orissa : Balasore : 20-12-52 (45) B.N.R. Tanks 2 and 11 ; (46) A.B.
Mohanty's Pond 1 ; (47) D.M.'s tank ; (48) Hafeezuddin tank (25-11-54) ; Dighi
Farm : (49) N.P. 12 (20-12-52) ; (50) N.P. 3,5 and 6 (26-11-54) ; Bhadrak:
(51) Parana Bazaar N.P. and (52) S.T. (28-11-54) ; Mayurbhanj : 21-12-52, Tanks
(53) Belgodia, (54) Chappal and (55) Himsagar ; Nilgiri (56) Raja's tank (28-11-
54) ; Keonjhar ; 19-2-57 : (57) Public tank and (58) N.P. ; Cuttack : Jenapur :
(59) N.P.2 and (60) S.T. ; (61) Jobra Fish Farm, Cuttack, N.P. 1-16 (1951-55) ;
(62) Cement tank (17-7-51) ; (63) Ella Fish Farm, N.P. 24-41 (1954-56) ;
(64) Killa Moat Sec. 5 (26-4-55) ; (65) Moat Extn. (9/11-4-56) ; (66) Office Pond,
Central Fisheries, 19, Cant. Rd (1965-66) ; (67) I.G.'s Pond, Cant. Rd (26-6-52) ;
(68) Pond, Bengali Sahi (1-3-61) ; (69) Pond, Dolmundi (13-1-66) ; (70) River
Mahanadi at Anicut (July '52-May '53) ; (71) Wolffia Pond, Satyabhamapur
(26-6-50) ; (72) Nuapara S.T. 5 and 6 (29/30-12-52) ; (73) S.T. 5 and 6 (25-2-57) ;
(74) Batai Ponds 1-3 and 9 and (75) Kujang Roadside Pond (30-1 2-52) ; (76)
Birbati pond (25-2-57) ; (77) Narasingpur S.T. 5 (17-12-54) ; (78) Chaudwar
Fish Farm S.T. 1 and 3 (2-1-53) ; Dhenkanal : (79) Kamakhyanagar Farm N.P. 1
and (80) N.P. 13 (18-12-54) ; Hindol (81) Faim N.P. 2 (82) Harihat Pukur and
(83) Subjail pond (20/21-12-54) (84) Talcher N.P. 6 and 10 (22-12-54) ; (85) Angul
Farm N.P. 5, 10 and 16 (22-12-54) ; (86) Golimoia Gundu Puku* (25-12-54) ;
Sundergarh : (87) Kunseri tank, Bonai Garh (11-12-54) ; Sambalpur : (88) Farm
ponds 2,11 and 26 and (89) Gorali tank (12-12-54) ; (90) Padampu^ Cota bund
(14-12-54) ; Bolangir : (91) Farm pond 10 (29-12-56) ; Puri : Kausalyagang faun :
(92) Swamp (10-4-51) ; (93) N.T. 4, S.T. 5, K and N (27-11-52) ; (94) S.T.,
Harikrishnapur, (95) S.T. 1 and 5, Attaranalla and (96) S.T., Brickfield farms
(11-12-52) ; Boudh-Phulbani : (97) Pond, Phulbani (19-12-54) : Ganjani :

The genus Euglena Ehrenberg 555

(98) S.T., Chatrapur and (99) Berhampur (20-12-54) ; Koraput : (100) Jeypore
nursery-1 (19-2-57).

2. If. Andhra Pradesh: Srikakulam : (101) Market Pond and (102) Pala bund
and Bokkara bund (3-12-54) ; (103) Jute wetting pond, Vizag Road (3-12-54) ;
(104) Yellamanchili market pond (5-12-54) ; Samalkot : (105) Amadalavalaru and
(106) Mothapuram ponds (6-12-54) ; Rajamundry : (107) Talkulwa pool, (108)
Tobacco Stn. pond and (109) Dwarapudi pond (6-12-54) ; (110) Venkam and
(111) Sesham Naidu ponds, Ellore (7-12-54) ; Kakinada : (112) Market pond and
(113) Ippur Farm pond-8 (8-12-54) ; (114) Chinnaravoor tank, Tenali and
(115) Ayithanagar tank, Tenali West (9-12-54); (116) A.R.P. Cement tank,
Vijayawada (9-12-54) ; Kurnool : (117) Sunkesala Farm ponds 9, 15 and 16 (10-
12-54) ; (118) Kutnool Fish. Office Pond and (119) Mariammakunta (11-12-54) ;
Hyderabad : (120) Kunta, Cess pool and (121) Pond 14, Hussainsagar (29-1-53) ;
(122) HayalnagaF, (123) Ibrahim Patan and (124) Sadyabag tanks (1-2-53) ;
(125) Jafar Charu (1-2-53).

2.1g. Karnataka : Bangalore : (126) Lalbagh Hort. Office pond (9-2-53) ; Mysore :
(127) Pool, Baratala, (128) Ishadbagh tank, (129) Hasalkot pond, (130) Dhobi Kote
and (131) Well-3, Nandi Hills (8-2-53) ; Cooig : (132) Ponnampet ponds 1 and
2 and (133) Beauvoir Farm pond, Mercara (9/10-2-53) ; Mangalore : (134) Fisheries
..College cement pond-1 (June-Nov. 1974) ; and (135) -do~(Jan.-Feb., 76).

2.1h. Kerala: Trichur : (136) Edasserikulam, Cranganore (10/27-2-49) : (136a)
Thandankulam. Azhicode (26-2-49) ; (137) Wolffia pond, Azhicode (29-9-51) ;
(138) Asarikulam, Madavana (9-10-51) ; (139) Kattukulam, (140) Pandarakulam
and (141) Pooppachira and (142) Temple tank, Cranganore (13/15-2-53) ; (143)
Vadakechira and (144) Padinjarekulam, Trichur (15-2-53) ; (145) Pond, Chalakudi
(13-2-53) ; (146) Pond and (147) Stream among rice fields, Chalakudi (17-2-53) ;
(148) Pond', Iringalakuda (13-2-53) ; Quilon : (149) Krishnapuram Res. Stn. pond
(24-2-53) ; Trivandrum (150) Aruvikkara reservoir.

2.1i. Tamilnadu : (151) Museum Pond, Madras (Dec. '37-Nov. '39) ; (152) Rect.
Pond, Chetput fish farm, Madras (11-11-42) ; (153) -do- (31-3-43) ; (154) -do-
(27-4-43) and (155) -do- (19-5-43) ; (156) Swamp, Chetput (22-11-44) ; Salem :
(157) Mettur Dam, N.P. 5 and 6 (21-12-54) ; (158) N.P. 2 (27-12-54) ; N. Arcot,
(159) Moat, Vellore (19-12-54) ; Nilgiris : (160) Muddy N.P., Fish Farm,
Ootacamund (11-6-52).

3. Estimation of plankton

Plankton were usually collected by straining a constant volume of water through
a plankton net made of No. 25 bolting silk (approx. 80 meshes per linear cm) and
the strained sample concentrated to a constant volume. Several drops from each
sample were examined and from fifty random fields of the mounts the organisms
were counted under the low power (about x 300) of the microscope and their
frequencies expressed as symbols, the numerical values for each symbol being
adopted from Howland (1931) as follows : i = 1-2 ; vr=3-5; r= 6-10;
re = 11-20 ; c = 21-50 ; vc = 51-100 ; a = 101-200 ; and va = over 200.

556 M T Philipose

When the organisms formed an appreciable scum, their frequencies were esti-
mated roughly as vc-va depending on the thickness and spread of the scum.

4. Ecological notes

Species of Euglena are usually found in large numbers in fish ponds, particularly
the smaller ones, and to a limited extent in multi-purpose tanks of small size and
in other shallow waters with decaying vegetation. Fish ponds are invariably
manured with cow dung and other organic manures, especially during May-June,
and are rich in organic matter, nitrates and sometimes phosphates. Public tanks
are usually polluted and in some of them there is pollution even from sewage.

Some of the species which were observed in large numbers in these water bodies
were E. tuba, E. pisciformis, E. oxyuris var. charkowiensis, E. proximo, and some-
times E. caudata and its variety minor. E. sanguined, E. viridis, E. acus, E. granu-
fata and E. tuba var. pseudotuba f. minima also occurred in some ponds in large
numbers. Of these, E. oxyuris var., E. acus and E. granulata usually occurred in
the general plankton while the rest formed fairly thick scums in addition
to their presence in the plankton.

E. tuba is one of the commonest species found all over north-east and south India
(also in west India-see Carter (1869) imparting to the water alternately a brick red
coloration during the bright hours of the day and dirty green at other times.
During bright periods the red haematochrome pigments are spread all over the
cell while in fading light they recede to the hind end. In encysted individuals
also the pigments usually remain at the hind end. In var. pseudotuba f. minima
the pigments are restricted to the axial region in fading light or when encysted.
E. tuba occurs also in the plankton and it has been found (see Philipose 1940) that
there is vertical stratification with its concentration gradually increasing towards
the surface by about 14hrs. on bright days and more uniform distribution at all
levels at night. The organism usually develops in the bottom silt wherefrom it
is recruited to the plankton.

In the filter beds at Barrackpore and ponds at Cuttack and Madras where year
round observations were carried out, E. tuba occurred abundantly throughout
the year except during periods of very heavy rainfall during July-September or
October-December. However, even during rainy months sometimes it occurred
in abundance during bright spells. E. pisciformis occurred abundantly in
Museum Pond, Madras, mostly during the heavy monsoon period of September-
November and at Cuttack in July. E. caudata dominated at Madras in October
after heavy rains accompanied by lowering of temperature whereas var. minor
occurred in two ponds at Barrackpore during the cold rainless month of January.
E. proxima was generally observed in ponds almost throughout the year with
greater development in January-February and December. E. oxyuris var.
charkowiensis usually occurred throughout the year (in Museum Pond, Madras)
with maxima during March-April and secondary maxima during June and August.
It also occurred in some ponds of Orissa in large numbers in August and December.
E. srinagari was common in February, June, August and December whereas
E, acus was more frequeitf in pebmaiy, May- June and November-December,

The genus Eugkna Ehrenberg

557

Text figure. Correlatian between meteorological conditions (av. air temp. av. daily
hrs of sunshine and total rainfall) and three species of Euglena in Museum Pond,
Madras (location 151) during 1938-39. (One year's data redrawn from Philipose
1940). !

Abbreviations : I— isolated ; VR— very rare ; R— rare ; \ RC— rather common ;

O— common ; VC— very common ; A— abundant ; VA— very abundant.

i

Text-figure 1 and table 1 give the meteorological and water conditions
respectively under which some of the species dominated. E. tuba occurred
abundantly when there was fairly long sunshine hours, little or no rainfall and
fairly high temperature in waters with fairly high total alkalinity, organic matter,
albuminoid ammonia, nitrates, chlorides, iron, variable phosphates, and moderate
pH. Its abundance in a large number of coastal waters probably suggests that
fairly high chlorides have a favourable influence though lower concentrations
apparently did not have any inhibiting effect as seen by its abundance in a few
interior waters with low chlorides. The fact that it was favourably influenced by
bright sunshine and probably high temperature was also borne out by its migra-
tion towards the water surface as the day advanced and its dispersal throughout
the water late in the afternoon and at night when sunlight was not operative and
the temperature was fairly uniform at all levels in small ponds. E. oxyuris var.
charkowlensis also appeared to be favoured by more or less similar conditions but
it showed its maximum in Museum Pond when the temperature was moderate,
sunshine hours high and there was very little rainfall. It {also appeared to be
favoured by fairly high nitrates. E. pisciformis, on the other hand, dominated
when there was lower sunshine hours, fairly good rainfall,) and maximum total
alkalinity, organic matter, nitrates and iron in Museum Pojid during September-
October. High temperatures seemed to bo unfavorable 'for its development.

M T Philipose

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The genus Euglena Ehrenberg

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560 M T Philipose

E. helicoideus, E. spirogyra and E. fusca seemed to be characteristic of shallow
waters with fairly low pH and total alkalinity and high organic matter caused by
decaying vegetation. E. granulata and E. vaginicola were quite common in a
rnoat at Cuttack during the bright hot month of April when macroflora were
decaying after chemical treatment.

Dodkundi et al (1973) observed marked stratification of six species of Euglena,
viz. E. acus, E. viridis, E. velata, E. gracilis, E. elastica and E. pisciformis (in the
order given) with increasing densities from bottom to surface after 9 a.m. in a
sewage stabilization pond at Dharwar. The zone of maximum density (surface)
was characterised by higher values for temperature, pH, dissolved oxygen, carbo-
nates and higher reductions in nutrients. They did not, however, study the varia-
tion, if any, in their densities at various levels in relation to sunlight and temperature
as the day advanced.

Fritsch and Rich (1913) and Lind (1938) reported maximum development of
Euglena, especially E. viridis, in British ponds when organic matter and sunshine
hours were high, with albuminoid ammonia and temperature as possible additional
factors. Lund (1943) observed abundance of E. viridis in the marginal silt of Clay
Pit Pond which had an organic content of 29-54%. In Kenwood Pond which
had a fairly low pH, high organic matter (30-50%) andiron compounds in the
marginal silt, Philipose (1948) observed abundance of E. viridis during February-
March and E. oblonga during May-August. The latter period corresponded to
long hours of sunshine, high temperature and decomposition of organic matter
resulting in reducing conditions in the silt accompanied by the release of fairly high
concentrations of ferrous compounds into the water. Pringsheim (1956) also
stated that most species of Euglena preferred water rich in nutritive substances
particularly readily available ferrous compounds and ammonium salts.

In Indian waters also a number of workers (Gonzalves and Joshi 1946 ; Zafar
1959 ; Singh 1960 ; Venkateswarlu I960, 1980 ; Munawar 1970 ; Hosmani
and Bharati 1980) correlated high densities of Euglonineae with high oxidisable
organic matter and sometimes dissolved iron. Other factors reported as favour-
able are high carbon dioxide, nitrates and temperature. Zafar (1959) also stated
that high nitrites and free ammonia higher than 0.104 ppm had an inhibiting
effect on Euglenineae. In Museum Pond, Madras, nitrites ranged from 0.005
to 0.02 ppm when E. tuba, E. oxyuris var. charkowiensis and E. pisciformis domi-
nated, which was quite low compared to the figures for the Hyderabad ponds
during certain periods. Free ammonia at average concentrations of 0.16-0.18
ppm. however, did not have any inhibiting effect on E. caudata and E. pisciformis
in Museum Pond. These two species were also favoured by lowering of tempera-
ture after heavy rainfall.

It was also frequently seen by the author in a series of ponds situated side by side
at Cuttack that E. tuba occurred abundantly in some ponds while it was absent
in the others. Apart from slight differences in the nutrient status of these ponds,
competition from other bloom forming algae, particularly Microcystis and
Anabaena and some Volvocales and Chloroooccales, appeared to be one of the
reasons for this difference. Ponds fertilized with inorganic fertilizers were also
usually free of this species at least for some tims while organically manured pon<Js
seemed to favour its |rowth,

The genus Euglena Ehrenberg 561

5. Systematic account

Genus Euglena Ehrenberg 1838

Single celled, usually free swimming, crawling or encysted (in resting condition) ;
fusiform to elongate-cylindrical with hind end frequently drawn out into a short
or long tail ; body fairly rigid, twisted or with varying degrees of " metaboly "
(change of shape) ; periplast striate ; vacuolar system a typical cystostome and
reservoir ; eye-spot by the side of the reservoir ; flagellum one and of varying
length ; chromatophores disc-shaped, band to ribbon like or trough-shaped, of
varying number and with or without pyrenoids; paramylum long or short rods or
plates, elongated Jinks, small discs or saucer-like sheathing the pyrenoids;
haematochrome present in some species ; usually in freshwater, rarely in brackish
water.

Key to the Indian species described :

I. Body fairly rigid with colourless tip ; chromatophores small, disc-to lens-
shaped and without pyi-enoids ; paramylum two or more solid rods or plates
or elongated Jinks ; nucleus usually ellipsoid and median ; flagellum shorter
than body Group Rigidae

(A) Paramylum in long rods or oblong solid plates, very rarely in links

(a) Paramylum more than two

(i) Body elongate-cylindrical, narrow and ending in a clear point ;
paramylum 3-16 rods, rarely links ; cells 65-220 x 4-8-25A
1 . E. acus

(ii) Body long and flattened with almost parallel sides,
twisted spirally ; paramylum 4-10 ; with a prominent tail ;
cells 240-530 X 25-40-60^ 2. E. helicoideus

(b) Paramylum usually two

(i) Body more or less spindle-shaped, rarely cylindrical

(1) Usually with a tail spine

+ Tail spine long ; paramylum one in front and the

other behind nucleus ; 52-150 X 6-8/j

3. E. acutissima

+ 4- Tail spine shorter ; paramylum in rods, rarely links,

and at same level as nucleus or one in front and

the other behind ; 51-82 x 7-1 Ip. . .4. E. hmnophila

Cells smaller and with additional elongate or

ovoid paramylum; 26.5-50 x 6-12^. . .var. minor

(2) With gradually tapering tail

+ Body small and not within a gelatinous envelope ;
striae longitudinal ; 50-53 X 6-8^ 5. E. vagans

562 M T Philipose

+ -I- Body larger and usually within a gelatinous envelope
striae spiral; 53-74x15-19.5^ with sheath;

41-63.5 X 11-16. 7/j without sheath

6. E. vaginicola sp. nov.

(ii) Body flattened 01 angular in cross-section ; with a prominent
tail piece

(1) Body flattened and grooved for part of its length ; usually
twisted during locomotion only ; 105-114 (-133) X 13-14
(-15. 5)/* 7. E. allorgei,

(2) Body angular in cross-section ; ' twisted even at rest
65-210 x 8-25/J 8. E. tripteris

Smaller form ; 48-52-63 x 9-15/* var. kkbsii

(B) Paramylum in elongated links and usually two, one in front and the
other behind the nucleus, rarely more.

(a) Body more or less cylindrical with short colourless tail ; striae
smooth ; (65-)85- 1 35 X 7 . 5-12 p 9. E. ignobilis

(b) Body more or less flattened or cylindrical ; striae with beads.

(i) Cell usually cylindrical ; pellicle yellowish ; striae beaded with
hemispherical excrescences ; tail distinct and more or less
straight and pointed ; usually 80-130 X 10-15/* . . . 10. E. spirogyra

(ii) Cell usually flattened ; pellicle brownish with square, rectangular
oi? L-shaped excrescences ; tail often gradually tapering and
oblique ; 153-225 x 117-27. 5 /* 11. E. fusca

(c) Body flattened in cross-section and twisted or grooved for
part of its length even when at rest.

(i) with two or more paramylum links ; body robust, about 10 times
as long as broad ; (280-) 375-490(^500) X 30-46 (-61-) JK
12. E. oxyuris

(ii) Paramylum only two

(+) Body about 12 times as long as broad and narrower than in
type; 201-270 (-290) X 16-22.5 (-30)/< var. playfairii

(++) Body about 6 (-8) times as long as broad and smaller than in
type (90-) 103-172 (-200) x 17-28 (-30)/* var. charkomensis

H. With marked metaboly by bulging, rarely by twisting ; tail when present
tapering gradually or short and stumpy ; chromatophores usually larger than
in " Rigidae " and lenticular, rarely small and discoid and without pyrenoids ;
paramylum usually many and granular to ovoid, rarely in additional long
rods ; nucleus ellipsoid to spherical ; flagellum 1/3 to li times the body
length — ........... r ........ f . , ........ , ...... T . , . . . .Group Lentifera^

The genus Eugtena Ehrenberg

(A) Body flat to cylindrical with both ends rounded ; chromatophotfes small
and discoid ; paramylum small, often in additional long rods ; nucleus
ellipsoid ; flagellum shorter than body.

(a) Without any tail ; pellicular striae fin© and close ; metaboly by
bulging or twisting ; without additional paramylum rods ; 110-200 X
15-40/z 13. E. srinagari

(b) With a short stumpy tail and uniformly prominent double striae ;
metaboly by twisting ; with additional paramylum rods ; 228-260 x
19-32-51// 14. E. pseudoehrenbergii sp. nov.

'(B) Body fusiform with gradually tapering tail ; metaboly by bulging ; chro-
niatophores lens-shaped and large ; paramylum large or small and short-
cylindrical to ovoid ; nucleus spherical ; flagellum 1-1£ times body
length ; cell (39-) 60-93 x (1 1-) 18-25/j 15. E. proximo.

I. Body fusiform with marked metaboly ; chromatophores in oblong, polygonal
or laminate plates or elongated bands with a " double pyrenoid " sheathed by
saucer-shaped paramylum caps ; with additional paramylum granules in

cytoplasm; nucleus spherical; flagellum body length or longer

Group Catilliferae

(A) Chromatophores 2 (rarely 3) parietal curved plates ; 16-35 x 5~12/j
16. E. pisciformis

(B) Chromatophores about 5-12 or more circular to angular plates ; without
haematochrome

(a) Body fusiform with conical anterior and double drawn out posterior
with a nearly cylindrical tail ; cturomatophores about 12 angular
plates ; 37-115 X 11-27.6 (-30> 17. E. granulata

(b) Body fusiform with narrowing posterior and drawn out or nearly
cylindrical anterior ; chromatophores circular to saucer-like, usually
lobed plates, often more than twelve; 60-110 x 15-38/*

18. E. caudata

Smaller form with 6-16 or more chromatophores ; 30-63 X 10-2 1/j
var. minor

(c) Body elongate fusiform to cylindrical; chromatophores 7-13 or more
circular to ovoid or triangular plates which are curved with the
pellicle and irregular due to close proximity ; striae faint ; 31-68 X
6-18-22.5/1 19. E. gracilis

(C) Chromatophores in elongated spindle-shaped bands ; flagellum usually
less than body length ; haematochrome present.

(a) Cysts like round-bottomed flasks with a stalk of varying length ending
in a funnel-like base ; chromatophores 5-17 or more ; cell 41-74-96
X 18-30-43/1 20. E. tuba

564 M T Philiposd

(b) Cysts like conical flasks with stalk almost absent and broad rim-like

base ; chromatophores numerous ; cell 87-130 x 17-27/j

'. var. pseudotuba comb. nov.

Smaller form with 5-10 ctoomatophores and dimensions, 45-82.5 x
17.5-31.5 (-37.% f. minima f. nov.

(D) Chromatophores numerous and in short or long bands.

(a) Chromatophores peripheral, 16-25, stellate with short bands punning
parallel to striae as well as radiating from centre ; paramylum two
and sheathing pyrenoid ; cell ovoid to ellipsoid ; flagellum 3/4 to
It times body length ; without haematochrome ; 50-110 x 16-37
(-40)M 21. E. oblonga

(b) Chromatophores in elongated bands in periphery ; cell ellipsoid to
fusiform or elongate cylindrical ; pellicle markedly striate ; flagellum
longer than body ; haematochrome present ; 50-150 (-200) x 22-55
(~72)/f 22. E. sanguined

IV. Metaboly less marked than in Groups II and III ; chromatophores ribbon-like
often breaking up into short lengths and radiating from 1-3 median " pyrenoid
centres ", rarely of two kinds (radial bands and discs) ; paramylum grains
usually massed round these centres ; nucleus spherical; flagellum about body
length Group Radiatae

(A) Chromatophores in bands radiating from one centre ; body fusiform
sometimes elongate-cylindrical ; 30-65 -73 (-89) X 9-22^. . .23. E. viridis

Larger form ; 69-94 x 28-40/j var. maxima var* nov.

(B) Chromatophores in elongated radiating bands as well as irregular discs ;
cells 62-86 (-128) x 12-23.5 (-29.5)0 24. E. hemichromata

(V) With marked metaboly ; cell cylindrical or with attenuated ends ; chromato-
phores large, disc-to trough-shaped, without pyrenoids ; flagellum easily

shed, locomotion by creeping Group Serpente,

(No species under this group observed by the author though E. deses,
E. guentheri and E. intermedia are reported by other workers).

I. Group RIGIDAE Pringsheim 1956

1. Euglena acus Ehrenberg 1956 1830 (figures 1 a-g)

Lemmermann 1913, p. 129, figure 209 ; Gojdics, 1953, pp. 99-102, Plate 11
figure 1 ; Huber-Pestalozzi 1955, p. 96, figure 15 ; Pringsheim 1956, p. 48
figure 2 ; Hortobigyi 1969 p. 30, figure 50 ; including var. minor Hansgiffg.

Body more or less rigid, rarely slightly twisted, elongate-cylindrical to spindle-
shaped with a slightly drawn-out neck and truncate anterior end ; gullet opening
slightly towards one side ; posterior end narrowing into a long of short hyaline

The genus Euglena Ehrenberg

Figures 1-5. 1 a-g. Euglena acus Ehr. (d and f slightly abnormal specimens,)
2 £". hdlcoldeus (Bern.) Lemm. 3 a-c. JS1. acutissima Lemm. 4 a-c. £*. limnophila
Lemin. 4 d-e. var. ra/>zor Drez. 5 a-b. £". v&gans Defl. [Figures of the same
magnification bracketed together :— (la) ; (lb;) (2) ; (Id-f, 4a-d, 5a»b) ; (Ig and
Ic) ; (3a-c, 4e)].

tail ; chroniatophores numetous, small and discoid ; nucleus usually ellipsoid
median or slightly behind ; paramylum 3-16 solid rods, rarely oblong plates, of
varying lengths ; eye-spot streak-like or nearly round ; flagellum usually up to 1/3
body length only ; striae when observed faint ; cells usually 85-170 x 7-12/f,
rarely as narrow as 4/x or as broad as 23/j, and as short as 64-74/L

566 M T Phitipose

Habitat : Planktonic ; common in locations 2, 14, 23, 27, 48, 61 (N.P.9.-May),
73, 82, 118 and 151 (February) ; rate in locations 1, 5, 17-19, 21, 26, 28, (29
(April, May, My, November), 33, 35, 44, 49, 57, 59, 63, 71-72, 75, 78, 80, 82-
84, 88-90, 93, 102, 105, 107, 113-18, 127, 134, 141-43, 154 and 156.

Considerable variation in shape and size of cell and in number and size of para-
mylum bodies was observed. The form from Chetput (location 154, figure Ic)
was very narrow in proportion to length (125 X 4//). An individual from Dum
Dum (location 23) was bulged out (figure If) towards the posterior end, had a
spherical nucleus and measured 88 X 23/j while another from Kamarhati (loca-
tion 27, figure Id) was intermediate in size (74 x 11.5/r) with slightly larger
chromatophores. A second individual from Kamarhati (figure le) measuring
64 x 8^ was slightly twisted. The Dum Dum and Kamarhati material had the
paramylum in broad plates or as a mixture of rods and plates. It was also not
uncommon to find long and very short rods together in normal narrow individuals.

Van Oye (as cited by Gojdics 1953) gives the range of size of the species as 65-
220 x 4-25/t and Chu (1947) as 60-160 x 7-15/*, whereas according to Pringsheim
(1956) the normal size comes within 80-150 X 7-12/j, if some of the varieties
are excluded. In Indian material Kamat (1961-1964) gives measurements of
100-180 X 6-15/f, Naidu (1962) 98-184 x 7/£ and Hortobagyi, I.e., 148 X 15.5/j,
as against 64-170 x 4-23/j in the author's material. Skuja's (1949) organism
from Burma measured 114-166 X 9-13/J.

The individual from Dum Dum resembled to some extent E. lata Swirenko (see
Gojdics 1953 p. 178 Plate 36, figure 2) which measured 93-95 X 33-39^.
Though Swirenko treated it as a distinct species, he himself suspected that it could
be an abnormal E. acus. Gojdics and Pringsheim also consider it as a E. aciis
or a doubtful species. Pringsheim also states that even Euglena with a fairly rigid
form might become deformed by the overcrowding of paramylum bodies under
certain unbalanced nutritional conditions. The individuals from Dum Durn and
Kamarhati, therefore, appear to be slightly abnormal.

Annular rings appearing as dark lines along the axes of paramylum rods (sec
Pringsheim 1956) or as links (see Hortobagyi 1969) were not observed in the
author's material.

Distribution in Indian region : Kashmir (Bhatia 1930) ; Gujarat (Kamat 1961-
62) ; Maharashtra (Cartel 1856, 1858 ; Gonzalves and Joshi 1946 ; Kamat 1963,
1964, 1974, ,1976) ; Uttar Pradesh (Hortobagyi 1969) ; Andhra Pradesh (Naidu
1962; Venkateswarlu 1976); Karnataka (Dodkundi et al 1973) ; Kerala
(Suxena et al 1973) Assam, W. Bengal, Madhya Pradesh, Orissa, Andhra Pradesh,
Karnataka, Kerala and Tamilnadu (!); Burma (Skvortzov 1937; Skuja 1949).

2. Euglena heUcoidem (Bern) .Lemm 1910 (figure 2)

Gojdics 1953, p. 119, Plate 18, figure 1 ; Rino, 1972 p. 149, Plate 5
figure 1 ; = Phacus helicoideus Bernard, 1908 p. 206 Pkte 16, figure 563 ;
= Euglena oxyuris var. helicoidea (Bern.) Playfair 1921 p. 119, Plate 3,
figure 18 ; = E. gigas Drezepolski 1925 p. 243, Plate 5 figure 159 ;
Huber-Pestalozzi 1955 p. 69, figure 44.

The genus Eitglena Ehrenberg 567

Body long and flattened with more or less parallel sides and twisted spirally ;
anterior end rounded ; posterior end with a pointed hyaline tail piece ; pellicle
fairly thick and with very fine close spiral striae ; paramylum (4-) 6-8 (-10) long
or short hyaline rods or plates. ; chromatophores small, discoid and numerous ;
eye-spot large ; flagcllum not observed ; Body (including tail) 319-330-368 x
25-36-37. 5/< ; tail, alone 33. 8-52. 5/<.

Habitat : Rare in plankton at locations 25, 34, 35, 92 and 160.

Though normally the paramylum (only 6-8 observed) were all in narrow rods,
in the Belgharia material (location 25) one individual had short (14 x 10.5//),
medium (24.5-28 X 10.5-12.3/0 and long (38.5-54.3 X 8.8-12.3/0 moderately
broad solid plates. Nucleus could not be observed clearly in most instances, but
in one individual (figure 2) from location 25 there were two small median
spherical bodies probably formed by the division of a single nucleus.

This species is distinguished from E. oxyuris with more than two paramylum by
the paramylum being in solid rods (rarely plates) and not in links. According to
Gojdics (1953 p. 120) the striae in E. oxyurls also appear as double lines with a
distance of 2/« between the striae. In E. helicoideus the striae are fine and close.
Pringsheim (1956) does not, however, place much reliance on the absence of para-
mylum links in E. helicoideus and considers it and E. gigas as possibly E. oxyurls.
In material of E. oxyuris (360-370 X 40-42/0 with more than two paramylum
observed by the author (1948) in a distinctly alkaline London pond (Highgatc Pond)
the paramylum were in distinct links when viewed along the broad side. In none
of the specimens from the five Indian localities this was the case. Because of this
and the close line striae the present species is retained here as E. helicoideus.

Huber-Pestalozzi (1955) observed E. gigas in highly eutrophic waters of Java.

•The water bodies from which the present species was collected were also shallow

with decaying vegetation and the water was fairly low in pH and total alkalinity.

Possibly E. oxyuris and E. helicoideus are characteristic of two distinct types of

waters viz. alkaline, and peaty and slightly acidic respectively.

Distribution in Indian region : — W. Bengal, Orissa and Tamiinadu ( ! ).

3. Eughnz &cutissim& Lemm. 1904 (figure 3 a-c)

Lemmerniann, 1910a, p. 496, figure 23 ; 1913, p. 129, figure 210

Body fairly rigid and spindle-shaped but sometimes slightly inflated by contraction
along the longitudinal axis. Differs from E. acus in having only two paramylum
rods, one in front and the other behind a median nucleus, the somewhat spiral
arrangement of the chromatophores and a proportionately long hyaline tail ;
flagellum not observed ; cells (including tail) 59.5-62 x 7-8 /r ; tail alone 18-19 /f.

Habitat : Rare in plankton of location 1, 29 (May), 33 (April), 70 (October) 72
and 151 (April, November)

Playfair (1921 pp. 120-21, Plate 4, figures 6-8) gives its dimensions as 110-

150 X 7-8/f with those of his var. parva as 54-94 x 6-8/z and states that the

*- Australian form is really E. acus. Gojdics (1953) and Huber-Pestalozzi (1955)

merge it with E. acus whereas Pringsheim (1956) states that " some species, for

P. (ft)— li

568 M T Philipose

instance E. acutissima Lemm. could be considered a variety of E. acus in the
wider sense". The species is retained here as distinct. The author could not
make out the nucleus clearly except in one specimen where it was small and
spherical (figure 3c). Johnson's (1944, p. 113, figure 13A-B) E. acutissima
(120-150 x 9-1 1/f) with more than two paramylum is obviously a E. acus.

Distribution in Indian region : Assam, W. Bengal, Orissa and Tamilnadu (!) ;
Burma (Skvortzov 1937).

4. EugknA limnophild Lemm, 1898 (figures 4 a-c)

Lemmermann 1913, p. 130, figure 205 ; Gojdics 1953 p. 103 Plate 11
figure 6 ; Huber-Pestalozzi 1955 p. 82 figure 59 ; Suxena 1955 p. 430
figure 1 ; Pringsheim 1956 pp. 50-51 figure 3 ; Hortobfigyi 1969 p. 30
Plate 4, figure 57

Body spindle-shaped with a slightly drawn out neck, truncate anterior end and
with a stiaight or slightly curved tail at posterior end ; body sometimes slightly
curved ; nucleus usually ellipsoid, large and median (rarely two small spherical
ones formed by division) ; chromatophores small and not arranged regularly ;
paramylum two rods, rarely flattened plates, one in front and the other behind the
nucleus or both just in front of nucleus at nearly the same level ; eye-spot prominent;
flagellum not observed ; cells (including tail) 51-70.5 x 7-7. Sjjt.

Habitat : Rare in plankton of location 33 (April-May).

There is some difference in the dimensions given by various authors. Lemmer-
mann gives it as 82 x 10/*, Skuja (1949) 55-69 X 7-13/1, Pringsheim 68-71 X
9-1 1/*, Suxena 60 - 80 X 7-10/*, Huber-Pestalozzi, 58-115 x 10/z and Horto-
b&gyi 68 x 9//. Lemmermann and Suxena noted a tail spine whereas Pringsheim's
specimens had a gradually tapering tail. Lemmermann also stated that the
paramylum rods were at the same level as the nucleus (also see Suxena 1955).
In the present material the tail was either gradually tapering or like a spine and
the paramylum just above the nucleus at nearly the same level or one in front and
the other behind as in Pringsheim's material. The nucleus is described as sphe-
rical by Lemmermann (1913) and Skuja (1948) and ellipsoid by Pringsheim. The
longitudinal dark line along the middle of the paramylum rods noted by Pring-
sheim and Skuja or the link-like paramylum seen by Pringsheim in some specimens
were not observed by the author. Nor was there any additional paramylum
(see Pringsheim 1956 figure 3A). The eye-spot was also quite small in contrast
to the large one mentioned by Huber-Pestalozzi.

Distribution in Indian region : Andhra Pradesh (Suxena I.e. ; Zafar 1959) ;
Maharashtra (Kamat 1975) ; Uttar Pradesh (Hortob&gyi 1969) ; Kerala
(Suxena et al 1973) West Bengal ( ! ) ; Burma (Skuja 1949).

rar. minor Drezepolski 1925 ( (figure 4 d-e)

Drezepolski 1925 p. 245 Plate 5, figure 162 ; Gojdics 1953 p. 103, Plate 11,
figure 7; Huber-Pestalozzi 1955 p. 83 figure 59B ; Hortobigyi 1969 p. 30
Plate 4, figure 55-56.

The genus Euglena Ehrenberg 569

Differs from the type in its smaller size and in having varying numbers of usually
small-sized cylindrical to ovoid paramylum bodies in addition to the normal
large ones. Cells 31-46 x 8-8. 5/z.

Habitat : Rare in plankton of locations 33 (July) and 151 (April).

Hortobagyi's specimens had an irregular outline and there were a number of
ring-like and cylindrical to ovoid paramylum bodies. In the author's material
rhere were 7-10 paramylum including two large ones but none of them were ring-
like. The two large ones in material of location 151 were also in the form of broad
plates.

Distribution in Indian region : Uttar Pradesh (Hcrtob&gyi 1969) ; West
Bengal and Tamilnadu ( ! ).

5. Euglena vagttns Deflandre 1932 (figure 5 a-b)

Gojdics 1953 p. 98, Plate 10 figure 8 ; Huber-Pestalozzi 1955 p. 94 figure 173

Body more or less cylindrical for most of its length, slightly tapered towards the
interior end which is obliquely truncate ; posterior end gradually tapering into a
short hyaline tail ; chi-omatophores small, disc-shaped and numerous ; paramylum
two and in the form of oblong solid plates, one in front and the other behind;
pellicle striated in a longitudinal manner ; nucleus, eye-spot and flagellum not
observed ; cells 49-55 X 7-8/j.

Neither Gojdics nor Huber-Pestalozzi refer to the striae
Habitat : Rare in plankton of locations 23 and 28 (May).

Distribution in Indian region : West Bengal ( ! ).
i. Euglen& vaginicofa sp. aov. (figure 6 a-b)

Drganismi intra cystam late fusiformem irregulariter serrulatam incolentes ;
systa antice late rotundata, aut colio brevi praedita, et aperturam parvam habens ;
sorpore fusiformi, antice rotundato, postice caudam gradatim attenuatam habente
lut in punctum abrupte terminante ; pellicula strias tenues spirales praebens ;
diromatophora parva, multa, disciformia, sine pyrenoidibus, aliquantulum spira-
iter ordinata ; paramylum ut duae viigulae solidae crassaeque, una anteriore,
ma posteriore, visum ; grana paramyli additica parva virguliformia ad disciforniia
nterdum praesentia ; fossa et stigma parva ; nucleus non observatus ; flagellum
;. 3/4 longitudinis corporis et per aperturam cystam anteriorem emergens ; cellulae
U-63.5 x lM6.7/im, vagina (cysta) 53-74 x 15-1 8.5 /mi.

Habitatio : Species rara in plancta locorum 23, 61, 63 et 73, frequens in loco 65.
Iconotypus : (figure 6 a-b)

Organism living inside a broadly spindle-shaped serrulate colourless gelatinous
mvelope (cyst) having a small opening at the anterior end and a gradually tapering
imooth posterior end. Anterior end of envelope broadly rounded or with a neck-
ike portion ; body spindle-shaped with the anterior end broadly rounded and the
posterior end with a gradually tapering, tail of abruptly ending in a point,
ktrferio* and posterior ends sfto^pg 'scpfe coBtoctiofo and expansion in living

570

M T Philip ose

Figures 6-12. 6a~b. Euglena vaginicola sp. nov. 7a-b. E.. allorgei Defl.
8a. E. tripteris (Duj.) Klebs. 8 b-c. var, klebsii Lemm. 9. £. ignobilis Johnson.
10.a-b. £*. spirogyra Ehr. 11 a- c. £. jfttscc- (Klebs) Lemm. 12 a-b. £". oxyuris
var. playfairii Bourr. 12c-d. J51. oxyuris var charkowiensis (Swlr.) Chu. [Same,
magnification :—(6a-b, 8b, 9); (7a) ; (7b) ; (8a) ; (8c) ; (lOa-b) ; (Ha, lie,
12a-c) ; (lib) ;

mat arial ; pellicle finely striated somewhat spirally with the spiral from left to right ;
chromatophores small, numerous, disc-shaped -and arranged somewhat spirally ;
paramylum two solid stout rods one in fronf and -the other behind ; with or
without additional rod-lik^to--discoid;^paramylum-,of . smaller size ; reservoir
and eye-spot* 'small, nucleus ^t^^bsdlved;: J^gettum-^^

The genus Euglena Ehrenberg 571

emerging through the anterior opening in the cyst ; cells 41-63.5 x 11-1 6. 7/z ;
cysts 53-74 xl5-18.5/<.

Habitat : Rare in plankton of locations 23, 61 (N.P.8,9,ll-May) ; 63 (N.P.25-
Janr.ary) and 73; common in location 65.

Figure 6 a-b to serve as the type.

The organism differs from all other known species of Euglena (except E. granulata
(Klebs) Schmitz-see Gojdics 1953 p. 138) in having a gelatinous cyst with an
aperture at the anterior end through which the flageHum emerges. In species
of Euglena with cysts, the cyst usually remains closed till the organism is liberated
by its breaking up or dissolution, and the flagellum does not develop inside the
cyst. In this respect the present organism resembled a Strombomonas, but the
essential organization of the cell was as in Euglena as when seen after removal of
the cyst with dilute potassium hydroxide solution. The cyst was also thinner
than the lorica of Strombomonas.

Though the liberation of the organism from the cyst was not observed, free
living individuals resembling in all respects those inside the cyst, except that the
cells were more cylindrical and always with a tail, were frequently observed in the
collections. In one such individual (57 x 10. 6ju) from location 23, distinct spiral
rows (12-13) of discoid chromatophores could be observed. There was alsopone
short stout paramylum (7-7.7 X S.5-4.4//) with a spherical (3.5-4.4/j) paramylum
at its inner end in each half of the cell with a distinct gap between the two ests.

Szabados (1950 as cited by Gojdics 1953) observed a bloom of E. granulata in
Hungary in which the organism was enclosed in flask-shaped cysts resembling the
shell of Trachelomonas similis with the flagellum emerging through an opening at
the end of a short neck, and the organism moved with the cyst. When the cysts
were exposed to strong light the organisms squeezed out of the cysts. Once free
they assumed the normal shape of E. granulata in a short time. The present
organism also moved with the cyst. It is quite possible that it was also in a condi-
tion of temporary encystment.

The organism inside the cyst showed- a certain degree of resemblance to
E. gaumei Allorge et Lefevre (1930, pp. 123-124, figures 9-12) in general shape,
size (60-70 X 10-12^) and in the presence of rectangular to oblong paramylum
(described as links but shown as solid ones in figures), but differs in having a thin
cyst in the encysted condition, in the chromatophores being more or less spirally
arranged, in the frequent presence of additional small paramylum bodies, a longer
flagellum and in the occasional absence of a tail. The striae in E. gaumei also
usually runs almost parallel to the long axis,, but cpuld be somewhat spiral
occasionally, whereas in the pfesent organism they appeared as Spiral. An eye-
spot and reservoir are' not Reported iff E. gatMetf'm the pteSetot organism they
could be seen faintly* in a few specimens.

It also shows some resemblance to E. vagans t>eflahdre in general appearance
and the gradually tapering tail, when present, but the anterior end of the present
organism is more rounded, tte cell ..is broader and usually within a cyst. As in
/?. gaumei, the "striae" in" & 1agiirts*:ftti5 affiriosTparaM to ' the Jong

572 M T Philipose

The organism under present consideration, is, therefore, treated here tenta-
tively as a new species under the name E. vagmicola. However, it has to be
observed whether it lives in a free state outside the cyst for any length of time.

Distribution in Indian region : West Bengal, and Orisssa (!).

7. Euglena allorgei Deflandre 1924 (figures 7 a-b)

Deflandre 1924 pp. 1116-1117, figures 1-2; Gojdics 1953, pp. 116-17,
Plate 16, figure 2 a-b ; Huber-Pestalozzi, p. 80, figure 56A.
Body nearly flattened in cross-section with the sides nearly parallel ; change of
shape only by curving during movement ; anterior end frequently with a slightly
curved furrow ; posterior end bent sharply towards one side or nearly straight
and ending in a hyaline tail piece ; pellicle striated more or less longitudinally ;
chromatophores small, discoid and numerous ; paramylum two long rods one in
front and the other behind a median ellipsoid nucleus ; eye-spot small and tri-
angular ; flagellum about one-third body length ; cell (including tail) 100-133 x
14.5-15.5/*.

Habitat : Rare in plankton of locations 26, 61 (December) and 151 (May).

Deflandre's organism is slightly smaller (105-114 x 13-14/f). He has also
shown the nucleus as spherical.

Distribution in Indian region : Maharashtra (Kamat 1975) ; West Bengal,
Orissa and Tamilnadu ( ! ).

(Kamat 1975, does not give any description or figures).

8. Euglena tripteris (Duj.) Klebs 1883 (figure 8a)

Lemmermann 1913 p. 130 figure 201 ; Skuja 1948 p. 198 figure 23, figures
12-13 ; Gojdics 1953, pp. 122-23, Plate 18, figures 3a-d ; Huber-Pestalozzi
1955 pp. 62-63, figure 39 ; Suxena 1955 p. 432 figure 14 ; Pringsheim 1956
pp. 57-59, figure 6.

Body elongate with nearly uniform breadth, rounded anteriorly with the canal
opening forming two lips, and ending posteriorly in a long hyaline tail piece ;
triangular in cross-section ; twisted markedly so that the body shows 2-3 sections
with the sides caved in; pellicle with dear striae which follow the twists ; chromato-
phores small, discoid and numerous ; paramylum two long rods or oblong plates;
(28-30 x 7-9//), one in front and the other behind an ellipsoid median nucleus ;
nucleus frequently not clear when the paramylum are in broad plates almost
touching each other ; eye-spot prominent ; flagellum about half body length ;
cells (including tail) 94-109 X 15-17/*.

Habitat : In plankton of locations 21 (March), 28 (April-May), 29 (October),
33 (February), 37, 45, 49-51, 56, 61 (N.P, 20-May), 63 (N.P. 32-My), 70 (July),
92, 113, 117, 134, 140 and 151 (November).

Distribution in Indian region : Andhra Pradesh (Suxena ; Zafar 1959 ;
Naidu, 1966 ; Venkateswarlu 1976) ; Maharashtra (Kamat 1975) ; West Bengal
Bihar, Orissa, Andhra Pradesh, Karnatoka, Kerala and Tamilnadu (!) • Burma,
(Skuja 1949). ' ' ?

The genus Euglena Ekrenberg 573

var. Klebsii Lemm. 1910 (figure 8 b-c)

Lemrnermaim 1910a p. 497 ; 1913, p. 130 ; Johnson 1944, p. 106,

figure 4 b-c; Huber-Pestalozzi 1955 p. 63 ; As E. frz>tera-Kamat 1961-62 p. 16

Smaller than the typical form ; cells 48-63 x 10-14/i (incl. tail) ; tail alone
10.5-14/* ; flagellum 18-20/f ; paramylum 10-15 X 3.5-5.3/f.
Habitat : In plankton of locations 13, 14, 23, 28 (December) 33 (April-June), 38,
61 (N.P. 20, 26, 28-29, February, March, May), 63 (N.P. 41-May), 73, 105, 114,
118, 151 (January-February) 152. Common in location 14 and rare in the rest.

Distribution in Indian region : Gujarat (Kamat 1961-62) ; Assam, West
Bengal, Bihar, Orissa, Andhra Pradesh and Tamilnadu ( ! ).

Gojdics (1953 pp. 122-23) suppressed all varieties of this species and merged
them with the typical form while Huber-Pestalozzi (1955) and Pringsheim
(1956 p. 59) retain some of them including var. klebsii. Though the size range
usually reported for the typical form is 70-80 X 8-14// (see Lemmermann 1913),
Chu (1947) gives it as 70-190 X 11-23// for his Chinese specimen whereas Skuja
(1948) gives it as (75-) 100-205 X 14-22/i with the large paramylum 17-40 x 7-1 1/z
in his Swedish material. Suxena (1955) gives dimensions of 100-210 x 15-230.
and Naidu (1966) 70-80 X 9-1 1//. Since the author's smaller form measures only
48-63 X 10-14 ju compared to Hortobagyi's Hungarian material with 58 x 9-10^
and Johnson's American one with 52-63 X 11-15/f, the variety is retained here.
Kamat's (1961-62) typical species measuring 25-30 X 5-7/r is much smaller than
even var. klebsii and could be a juvenile form. Kamat (1975) has not given the
dimensions of his typical species from Maharashtra.

Both Gojdics and Pringsheim refer to additional short rod-like, rectangular or
ovoid paramylum in the typical species. In the author's material two additional
very small rods were observed only in one individual of var. klebsii. Johnson
(1944) states that var. klebsii has less torsion than in the type. In the author's
material both well-twisted and individuals with only slight torsion were observed.

9. Euglena ignobilis Johnson 1944 (figure 9)

Johnson 1944 p. 118 figure 20 A-C

Body more or less cylindrical and curved, with the anterior end slightly narrowed
and rounded ; posterior end with a very short hyaline bent tail ; pellicle with promi-
nent continuous striae which are more or less longitudinal ; chromatophores small,
discoid to ovoid and numerous ; paramylum two oblong links, one in front and
the other behind a median ellipsoid nucleus ; with a few additional rod-like small
paramylum ; eye-spot fairly small ; flagellum not observed ; cell 65-69 x 7.5-8/«.
Habitat : Rare in plankton of locations 29 (February) 73 and 81.

The organism agreed in most respects with the American one, but was smaller,
the American form measuring 85-135 X 8-12/* and with flagellum i-J body length.
Also, there was Jess metaboly and the nucleus was ellipsoid compared to prolate
spherical in the American species. According to Pringsheim (1956 p. 57) this
species looks more like E. spirogyra minus papillae such as commonly occurs in
cultures.

Distribution in Indian legion : Mahansshtia (Kaxnat 1975 p. 466) ; West

Bengal and Orissa (!)
(Kamat 1975 does not give aiiy description ot figtrre.)

574 M T Philipose

10. Euglena spirogym Ehrenberg 1838 (figure 10 a-c)

Lemmermann 1910a pp. 488-89 figure 18 (p. 483); 1913 p. 131 figure 208 ;
Gojdica 1953 pp. 111-112 Plate 14, figure 2a-b ; Huber-Pestalozzi 1955
p. 101, figure 81 ; Suxena 1955 pp. 432-33 figure 5 ; Pringsheina 1956
pp. 53-56 figures 4-5 ; including var. marchica Lemm., 1913 p. 131 ; = E. fusca
wr. marchica Lemm., 1910a p, 498.

Body elongated and more or less cylindrical with the sides nearly parallel ; body
sometimes bent but not usually twisted ; anterior end slightly tapering with a
rounded end, with the canal opening frequently towards one side ; posterior end
also somewhat rounded and usually ending abruptly in a distinct hyaline tail piece ;
pellicle yellowish and with spiral rows of bead-like hemispherical excrescences
which are either uniformly developed (as in var. marchica Lemm.) or with 1-3 rows
of weakly beaded excrescences between two rows of strongly beaded ones ;
paramylum two fairly large Jinks one in front and the other behind an ellipsoid
median nucleus ; chromatophores small, discoid and numerous ; eye-spot fairly
large ; flagellum not observed ; body usually 62.5-86 X 10.5-17.5;*, rarely up to
160 X 16;*.

Habitat : Rare in plankton of locations 16, 28, 61 (N.P.5 and 15-December
May), 64, 65, 73, 92, 95, 132, 136, 144, 146, 148 and 157.

Bhatia (1930) gives its dimensions as 140 x 18-20;* and Suxena (1955) as 100-
110 X 13-14;*. Dimensions normally accepted for the species (see Pringsheim
1956 p. 55) are 80-130 x 10-15^. In the author's material they ranged from
62.5-86 x 10.5-17.5/4 with one individual from location 136 measuring (including
tail) 160 x I6p (see Plate III, figure lOc).

Distribution in Indian region : Kashmir (Bhatia 1930) ; Himachal Pradesh
(Kamat 1968a) ; Uttar Pradesh (Singh 1960); Gujarat (Kamat 1961-62); Bombay,
Maharashtra (Carter 1856); Andhra Pradesh (Suxena 1955; Zafar 1959); Assam,
W. Bengal, Orissa, Karnataka, Kerala and Tamilnadu (!) ; Burma (Skvortzov
1937 ; Skuja 1949).

11. Euglena fusca (Klebs) Lemm. 1910 (figure 11 a-c)

Lemmermann 1910a, p. 496 ; 1913 p. 130 figure 202 ; Gojdics 1953
pp. 112-14 Plate 15 figure la-f ; Huber-Pestalozzi 1955 pp. 64-65 figure 41 ;
Prowsel958 pp. 145-46, figure 1 e; = E.spirogyra var. fusca Klebs, 1883

Differs from E. spirogyra in the body being usually flat and ribbon-like with the
sides more or less parallel, twisted or slightly bent, posterior end usually gradually
tapering and ending in a tail which is sometimes slightly deflected towards one side '
and the pellicle being beaded with square, rectangular or L-shaped brownish
excrescences which are frequently closer and more marked ; paramylum links
quite large ; body 150-199 x 20,5-27/J.

Habitat : Rare in the plankton of locations 34, 92, 136 and 138.

Themost accepted size of the species is 170-225 x 23-27 . 5/i though Lemmermana
(1910a) gives it as 90-225 X 23-27-. Sfo Playfair (1921) as 160-250 X 18-3

The genus Englena Ehrenberg 575

Prowse (1958) as 150-230 X 15-20/j. Kamat (1963, 1964) gives dimensions of
125-200 X 22-30/j and 100-155 X 30-33^ respectively in his Kolhapur and
Bombay materials.

Lemmermann also makes a distinction on the basis of length of flagellum, that
of E. fusca being of body length and that of E. splrogyra up to only about one-
fourth body length. In the author's material the flagellum could not be observed
in E. spirogyra and in one instance of E. fusca where it was observed (Plate II,
figure 1 la) it was quite short. Gojdics (1953) states that not much reliance can
be placed on length of flagellum to distinguish between the two species. Prowse
(1958) and Riao (1972 p. 150) also report a small flagellum in E. fusca.

As in E. spirogyra the beading on the pellicle may be uniform or with 1-3 rows
of weakly beaded rows between two strongly beaded rows or sometimes the beads
in the same row may not be uniform (see Gojdics 1953). The author could not
find these in his material.

According to Pringsheim (1956 p. 55) " E. fusca Lemmermann (1913 p. 130)
is so near to E. spirogyra that it may be better to return to Klebs'' (1883 p. 77)
original suggestion of considering it as a variety of the latter ".

Distribution in Indian region : Maharashtra (Kamat 1963, 1964, 1976) ;
West Bengal, Orissa and Kerala (!).

12. Euglena oxyuris Schmarda 1846

Lemmermann 1910a p. 497 figure 16 (p. 483) ; 1913, p. 130 figure 207 ;
Gojdics 1953 pp. 120-21, Plate 20 figure 1 a-d ; Huber-Pestalozzi 1955,
p. 65 figure 42.

Body elongated and more or less flattened with 2-3 twists even when at rest ;
anterior end rounded or truncate ; posterior end with a hyaline tail piece ; pellicle
fairly thick and striated spirally parallel to the twists, the striae appearing as
double lines (see Gojdics 1953) ; chromatophores small, discoid and numerous;
nucleus ellipsoid and median ; paramylum usually two elongated links one in front
and the other behind the nucleus, rarely more (up to 20) distributed fairly uni-
formly ; eye-spot prominent ; flagellum up to about one-third body length. A fairly
robust species which is on the average about ten tiroes as long as broad with the
cell (280-) 375-490 (-500) X 30-46 (-61) jn.

The typical form was not observed in the author's collections. However, it
has been reported from Burma (Skvortzov 1937; Skuja 1949), Maharashtra
(Gonzalves and Joshi 1946 ; Kamat 1963, 1975) and Himachal Pradesh (Kamat
1968a). Kamat (1963) gives its dimensions as 280-300 X 30-35/*. From the
figure of Gonzalves and Joshi it is about 420 x 38-42/z. On the basis of size,
Bhatia's (1930) organism (148 x 14-18 /<), Suxena's (1955) measuring 160-200 x -
12-18/* and Kamat's E. estonica Mfllder (Kamat 1967) measuring 120-170 x
15-20/z could be included under E. oxyuris van charkowiensis (Swir.) Chu (see
below). Skuja's (1949) E. oxyuris measuring 152-231 X 20-26 /t could also be a
mixture of one or two varieties mentioned below rather than the typical form.

var. playfairii Bourrelly 1949 (figure 12 a-b)

Gojdics 1953 p. 122 ; Huber-Pestalozzi 1955 p. 66 ; = E. oxyuris var. $racilHm<*
.. . Playfair 1921 p. 11.9

576 M T Philipose

Diifers from the typical species in being markedly delicate and narrow in propor-
tion to its length, with the anterior end truncate or with prominent lips, posterior
end with a fairly long tail which is straight, curved or slightly deflected to one
side ; paramylum two links, one in front and the other behind the nucleus ; eye
spot fairly big and more or less rounded; flagellum not observed ; body on the
average about twelve times as long as broad with dimensions of 201-270 x 16-
22.5jLL ; tail alone 47-64/z.

Habitat : In plankton of locations 4, 6, 34, 61 (N.P. 27-rather common, August )
63 (N.P. 24, 26-28, 38-July, August, January, March, November), 73, 110, 116
and 132.

The organism in the author's collections was consistently narrow with dimen-
sions of three individuals which could be measured being 200. 6 x 16-17. 8/x, 206 X
17-20.6/Lc and 270 x22.5/t which was in agreement with Playfair's dimensions of
253 x 17/f.

In describing the variety gradllima Playfair stressed on the " gracile " (slender)
nature of the body. Though the name is probably appropriate, it is not clear
why Bourrelly preferred the name fa playfairii, the author having not consulted
Bourrelly in original.

Distribution in Indian region : Assam, West Bengal, Orissa, Andhra Pradesh
and Karnataka (!).

var charkowiensh (Swirenko) Chu 1947 (figure 12 c-d)

Chu 1947 pp. 125-28, figures 29-35 ; Gojdics 1953 p, 121, Plate 18, figure
2 a-b ; Prowse 1958 p. 146 figure If ; = Euglena charkowiensis Swir. 1913 ;
Huber-Pestalozzi 1955 p. 61 figure 37 ; Kamat 1961-62 p. 262 ; 1964 p. 9 ;
= E. oxyuris f. minor Defl., 1924, pp. 1117-1118, figure 9 ; Philipose 1940
p. 194 Plate 6 figures 107-108; Huber-Pestalozzi 1955, p. 65 ; Hortobigyi 1969
p. 30 ; = E. oxyuris fa charkowiensis Bourrelly, 1949.

Differs from the typical species due to its smaller size, the body being on the
average about 6 (-8) times as long as broad ; body 127-144 x 16.5-26/*, rarely
121 x ll.SjjL or 135 X 18.5-36/z, the difference in breadth in the last being due to
slight posterior inflation (figure 12c) ; tail alone 16-26/* ; paramylum two fairly
large links.

Habitat : Planktonic ; very common in location 46 ; very common to very
abundant (March-April), common (June, August), rather common (May, July
September) and stray to rare in other months in location 151 ; rather common to
common in locations 63 (N.P. 32-August), 89 and 127 ; stray to rare in locations,
4-6, 13, 18-20, 26, 29 (May-July), 33 (October), 34-35, 37-38, 40, 45, 51, 54-55
58, 61 (May, November) 63 (May, July-September), 72, 75, 81, 83, 86, 88,93-95
100, 103, 106, 115-17, 119, 121, 127-33, 134 (June), 135, 136, 154 and 157.
Distribution in Indian region : Kashmir (Bhatia 1930 as E. oxyuris, 148 x 14-
18 /x) ; Rajasthan (Kamat 1967 as E. estonica M61der 120-170 x 15-20 fjt)
Gujarat (Kamat 1961-62 as E. charkowiensis Swir., 130-170 x 24-27 /i) ; Maha-
rashtra (Kamat 1963, 1964, 125-1 50 X22-30/* ; 1974 ; 1975, as E. charkowiensis)
Andhra Pradesh (Suxenal955, 160-200x12-18/4; Naidu 1966, 170-180x24-25/4)
Uttar Pradesh, (Hortobdgyi 19fe9, as f. minor EfeflL, 143-156 x 20-220) ; Assam

The genus Euglena Ehrenberg 577

West Bengal, Bihar, Madhya Pradesh, Orissa, Andhra Pradesh, Karnataka, Kerala
and Tamilnadu (!).

Kamat's E. oxyuris (1963) from Maharashtra with dimensions of 75-300 X 15-30
(-50)jU could be a mixture of the typical species and a few other varieties.

There has been considerable confusion regarding the exact distinguishingfeatures,
particularly the dimensions, between E. oxyuris and its varieties, and in a number
of instances there have been overlapping dimensions. In 1949 Bourrelly made a
comparative study of the species and its varieties, and on the basis of size created
four forms, viz. fa minima Bourr. with dimensions of 74-86 X 6. 5-10/j (see Gojdics
1953 p. 122), fa charkowiensis (Swir.) Bourr. (which is synonymous to var.
charkowiensis (Swir.) Chu. 1947 with dimensions of 125-150 x 20-27// ; fa
estonica (Molder) Bourr. (= E. estonica Molder 1943) with measurements of 160-
230 X 16-22/j ; and fa play f aim Bourr. with dimensions of 247-290 X 20-30/j.

Gojdics (1953) is of the opinion that since Bourrelly created these forms on the
basis of size alone, the last variety is superfluous. Further Gojdics (p. 125)
retains E. estonica as a separate species on the basis that the paramylum is in the
form of two small round bodies.

In the opinion of the author, E. estonica is very similar to E. oxyuris. In r ich
collections of E. oxyuris var. charkowiensis the author has come across individuals
with the paramylum very much reduced in size or sometimes there is only one
paramylum. It is just possible that the smaller spherical paramylum in E. estonica
could be due to improper development. The author is in favour of suppressing
this species and merging it partly with var. charkowiensis and partly with var.
playfairii of E. oxyuris. Thus, three varieties may be recognised as follows :

(1) var. minima Bourrelly 1949 ( = E. charkowiensis Swir. f. minor Skvortz
1925 ; E. oxyuris var. minor Prescott 1945 (see Gojdics 1953 p. 121) with size
55.5-86 x 6-10 (15-18)^, the body being about nine times as long as broad

(2) var. charkowiensis (Swir.) Chu. 1947 with cells (90-) 103-172 (-200) x
16.5-28 (-30)/f , the body being about 6(-8) times as long as broad. Most common
size 125-150 X 16.5-27/r ;

(3) var. playfairii Bourrelly 1949 with cells 201-270 (-290) X 16-22.5 (-30/0
with a length-breadth ratio of about twelve (rarely up to fifteen). Tail (47-64;*)
also relatively longer in this variety.

The typical species (with two or more paramylum) will be left with dimensions
of (280-) 300-490 (-500) x 30-46 (-61)/i with a length to breadth ratio of about
ten. There would still be a little bit of overlapping, but it would be much less.

Kamat (1961-62) gives the dimensions of 80-90-100 x 12-13/* for his var.
minima Bourr. recorded from Ahmedabad.

n Group LENTLFERA Pringsheim 1956

13. Euglena SrinAgari (Bhatia) Huber-pest 1955 (figure 13 a— d)

Huber-Pestalozzi 1955pp. 58-59 figure 32 ; = Amblyophis srinagari Bhatiat
1930, p. 363 figure 1 ; Euglena amblyophis (Ehr.) Playfair 1921 p. 118 Plate 3)
figures 10-11; As E, ehrenber§ii Klebs-see Philipose 1940 p. 193 Plate 6?
figure 124. . . """"• ..

578

M T Philipose

I8e

Figures 10 c, 13-49. 10 c. Euglena spirogyra Ehr. 13 a-d. E. srinagari (Bhatia,
Huber-Pest. 14. E. pseudoehrenbergii sp. nov. 15a-d. E . proxima Dang.
16a-c. E. piscifor mis Klebs. 17. E. granulata (Klebs) Schmttz. In fignre 17
striae acfully from right to left. Cell accidentally reversed while transferring.
18 a-c. E. caudata Huebner. 18 d-f. var minor Defl. 19 a-b. E. gmcilis Klebs. [Same
magnification :-[(10c, 15b, 15d) J (13 ani, 14); (l5c, 16a) ; (18 d-f, 19 a~b:

Body flat to cylindrical and rounded at both ends with the anterior end frequently
narrower ; metaboly by bulging in the middle or rarely at the anterior end, often
shortening^ about half its normal length and also by twisting once along the longi-
axis } chromatophores ttumerbtiS small atrd discoid, without pyrenoids|-

The genus Euglena Ehrenberg 579

nucleus usually ellipsoid, rarely oblong, and median or sub-median ; paramylum
many, small and spherical to ovoid or small rod-shaped bodies ; eye-spot fairly
small and by the side of the reservoir ; flagellum up to 1/3-3/4 body length*
pellicle fairly thick and usually with fine close spiral striae which are sometimes
not visible ; body 116-160 X 15-49^ but in shortened individuals it could be
79 X 52/«.

Habitat : Rather common in plankton of locations 61 (N.P. 8 and 13-May and
August), 67, 82 and 106 ; stray to rare in locations 17, 34, 35, 53, 57, 61 (N.P.
6,8-May), 63 (N.P. 32-My), 68, 73, 76, 92, 127, 133 and 151 (January-
December).

Gojdics (1953 pp. 108-10) considers Bhatia's organism as synonymous to
E. ehrenbergii Klebs. Playfair's E. amblyophis, which is inadequately described
has also been considered by most authors (Gojdics 1953 ; Pringsheim 1956 p. 133
as synonymous to E. ehrenbergii.) While treating E. srinagari as a separate species
Huber-Pestalozzi (1955) states that it is either near or the same as E. ehrenbergii

However, E. ehrenbergii as reported from Europe, America and a number
of other areas, has invariably one to ten additional long straight or slightly curved
paramylum rods, or, as in Swedish material (see Skuja 1948 p. 192 Taf. 23
figures 7-8) stout solid rods, though occasionally these rods may be absent (see
Huber-Pestalozzi 1955 pp. 69-70). In Skuja's material the chromatophores were
fairly large and lenticular and the eye-spot fairly large (10-12 x 10/0 and in the
form of a curved plate. In material from a London pond observed by the author
(1948 Plate 2, figures 41, 48) the chromatophores were small, eye-spot small or
large and there was one additional paramylum rod by the side of the nucleus.
The range of size given by various authors (including Lemmermann 1913; Johnson
1944 ; Gojdics 1953 ; Huber-Pestalozzi 1955) fall within 107-400 x 13-48/«.
It would thus appear that E. ehrenbergii as conceived by most authors is an
extremely variable species. This species (with five paramylum rods) has been
reported only once from India (see Gonzalves and Joshi 1946 Plate 5, figure 7)
without any accompanying description.

Though the author observed the organism resembling E. ehrenbergii in several
respects from a number of localities in north-east and south India, surprisingly in
none of the individuals he could observe long paramylum rods. Chromatophores
and paramylum were always small and discoid, the eye-spot fairly small and the
striae when observed were fine and close. With usual dimensions of 116-160 x
15-40/4 it came near Bhatia's Amblyophis srinagari (114-37/0 and Prowse's (1958
p. 145) E. ehrenbergii from Malaysia (also without paramylum rods and 1 10-200 x
'15-20/0. Playfair's (1921) organism though slightly larger (200-300 x 20-25) /«
and described as without paramylum resembled E. srinagari better than £. ehren-
bergii. Though Playfair stated there was no flagellum his figure indicated a very
short one. Bhatia also figured a very short one. In the author's specimens
it was about 1/3 to 3/4 body length. " Metaboly " was also quite marked in the
author's material in contrast to limited metaboly in typical E. ehrenbergii.

The author is, therefore, of the opinion that organisms resembling E. ehrenbergii
but without long paramylum rods and some other associated characters are better
"treated as E. srinagari (Bhatte) Htibfcr-Pest. ....... •'••...

580 M T Philipose

Distribution in Indian region : Kashmir (Bhatia 1930) ; Assam, West Bengal,
Orissa, Andhra Pradesh, Karnataka and Kerala ( ! )

14. Euglena pseudoehrenbevgii sp. nov*

Corpus elongatum applanatumque, latitudine varians ; extremitas anterior mani-
feste augustata rotundataque, posterior necnon late rotundata, caudam conicam
brevern crassam habens ;metabole tortu effecta ; pellicula strias duplices acque
crassas praebens ; cauda necnon partim strias habens, estremitate ipsa, autem,
levi hyalinaque ; chromatophora parva, multa, disciformia ad lenticularia, sine
pyrenoidibus ; nucleus medius, ellipsoideus ad fere sphericum ; paramylum
parvum, virguliforme ad discoideum, in cytoplasmate dispersum ; duae additicae
virgulae paramyli longae solidaeque iuxta nucleum sitae ; fossa fere spherica,
canali brevissinio ; stigma flagellumque non observata ; cellula 228-260 x 19-51 /on,
cauda ipsa 8-10/mi long, striae 8-9 per 40 /mi.

Habitatio : Species in plancto loei 112 rara. Iconotypus : Plate III, figure 14.
Body elongate and more or less flattened with varying breadth ; anterior end
markedly narrowed and rounded ; posterior end broadly rounded and with a
short stout conical tail ; change of shape by twisting ; pellicle with uniformly
thick smooth striae which appear as double lines ; tail also partly with striae, the
very tip being hyaline and clear of striae; striae spiral from upper left to lower
right ; chromatophores numerous, fairly small, disc-shaped to lenticular and
without pyrenoids ; nucleus (18.5-27.5/0 ellipsoid to nearly spherical and
median ; paramylum small, rod-like to discoid distributed in the cytoplasm ; two
additional elongated solid rods (43-48 X4.5-5.5/*) by the side of the
nucleus ; reservoir nearly spherical and with a very short canal ; eye-spot and
flagellum not observed ; cell 228-260 X 19-32-51/* with tail alone 8-10/z,
long ; striae 8-9 in 40/z.

Habitat : Rare in plankton of location 1 12 (Market Pond, Kakinada), Plate III.
figure 14 to serve as the type.

The organism bears a close resemblance to E. ehrenbergii in shape, size and
general structure but differs in having a short stout conical tail and its uniformly
smooth double striae. It is also somewhat like E. subehrenbergii but in the
latter the hind end is truncate and hollowed out, the striae are fine and punctate
the paramylum is in ovoid-hexagonal bodies and there are no additional paramylum
rods. The chromatophores in the present organism are also slightly smaller.

In the presence of striae in double lines the organism resembles three other
species, viz. E. oxyuris Schmarda, E. heimii Lefevre and E. mesnili Defl. et Dusi.
Though the striae in E. oxyuris may be in double lines (see Gojdics 1953, p 120),
it is a well defined species with two or more paramylum links and a distinct hyaline
tail piece. E. heimii Lef. 1934 (see Gojdics, 1953 p. 177 Plate 35 figure 13 ;
Huber-Pestalozzi 1955 pp. 74-75, figure 52) has thick striae alternating with thin
ones, has the general appearance of a E. ehrenbergii, has one long straight or curved
paramylum rod and is 140-180^ in length. Though the hind end is described as
bluntly or obtusely rotmded, figures 52 d and g reproduced by Huber-Pestalozzi
suggest a very small conical point or tail ; Both Gojdics (1953) and Pringsheim
(1956) consider E. heimii as synonymous to E. ehreribergil The present organism

The genus Euglena Ehrenberg 581

is essentially different from both these species. E. mesnili Defl. et Dusi (see
Gojdics 1953 p. 88 Plate 7, figure 4) having thick striae alternating with delicate
ones or with a delicate striae after several marked ones, has been shown by
Pringsheim (1956) to be really a E. deses with which the present organism has
no resemblance.

The Kakinada organism is, therefore, considered a new species allied to
E. ehrenbergii under the name E. pseudoehrenbergil

Distribution in Indian region : Andhra Pradesh ( ! ).

Kamat (1963,1974) recorded E. heimii from Maharashtra. The dimensions of
the Kolhapur organism were 75-110 x 30-45//.

15. Euglena proximo, Dangeard 1901 (figure I5a-d)

Lemmermann 1910a pp. 498-99 figure 7 (p. 483) ; 1913, p. 129 figure 193 ;
Gojdics 1953 p. 90 Plate 7 figure 7 a-b ; Huber-Pestalozzi 1955 p. 86 figure 64
Pringsheim 1956 p. 62 figure 8 ; Prowse 1958 p. 143 figure Ic; Hortobagyi
1969 p. 30 figure 53.

Body fusiform, somewhat blunt towards the anterior end and tapering gradually
to a hyaline point at the posterior end ; changing shape markedly by contraction
or bulging without twisting and easily rounding up on irritation ; pellicle with
fine spiral striae ; chromatophores lenticular to ovoid and larger than in E. srinagari
and E. pseudoehrenbergii and without pyrenoids ; chromatophores generally absent
at the posterior and sometimes at the anterior ends ; paramylum numerous, ovoid
to short-cylindrical and almost the size of the chromatophores or sometimes
smaller ; nucleus nearly spherical and usually slightly below the median region ;
eye-spot elongate, disc-like or nearly triangular ; flagellum 1-1 Jr times body length ;
cells 39-83 x 11-22.5;*.

Habitat : One of the commonest species found in almost all collections, forming a
thin green scum when in abundance, or in the general plankton. Abundant at
locations 29 (February 61 (N.P. 9-May)) and 66 (January); Rather common to
common in locations 36, 39, 42, 43, 56, 77, 78, 88, 93, 100, 122, 143 and 151
(April and November and stray to rare in other months) ; stray to rare in other
localities.

The usual dimensions of this species is 60-93 x 18-25/j. However, Pringsheim
(1956) states that there are two size groups with averages of 50 and 6Qju in length,
though it is difficult to measure the organism on account of its marked ' metaboly ".
Hortob&gyi (1969) gives dimension of 80-90 X 14-16//, and Prowse (1958) 45-60 x
15-20/* in bis Malaysian material. The smaller paramylum are described by most
authors as links. However, the author could not make out links in his material.

Naidu (1962, p. 88, figure 1) describes and figures E. proximo, (85-90 x 26-30/0
as having two ovoid quite large paramylum bodies, one in front and the other behind
a small central spherical nucleus. Obviously this is not a E. proximo,. The figure
(Plate IV, figure 7) given by Gonzalves and Joshi (1946) for this species is also mis-
leading since the striae are shown as punctate and there is only just one paramylum
link.

Distribution in Indian region : Maharashtra (Gouzalves and Joshi 1943 a,
1946 : Kamat 1968, 1974) ; Uttar Pradesh (Horfob&gyi 1-969); Assam, West Bengal,

582 Af T Phitipose

Bihar, Madhya Pradesh, Orissa, Aadhra Pradesh, Karnataka, Kerala and Tamil-
nadu ( ! ).

Group II! CATiLLIFERAE Priagsheim 1956
16. Euglena piscifonnis Klebs 1883 (figure 16a-c)

Lemmermann, 1913 p. 125 figure 182 ; Huber-Pestalozzi 1955 p. 41 figure 15 ;
Priagsheim 1956 pp. 67-70 figure 12 ; including var. minor Hansgirg ; =
E. agilis Carter 1856 ; Gojdics 1953 pp. 133-36 Plate 1 figure 5 a-e ; Prowse
1952 p. 108 Plate 1, figure h-i ; Naidu 1966 p. 26 figure 7-8.

Body nearly cylindrical with the anterior end rounded and the posterior end
rather blunt and pointed ; changing shape by contraction and bulging and
rounding up on irritation ; chromatophores two (rarely three) elongated laminate
plates, each with a pyrenoid sheathed by paramylum caps ; additional ovoid to
short rodlike paramylum in the cytoplasm ; nucleus more or less spherical and
in the posterior half; eye-spot streak-like; flagelhim 1-2 times body length;
striae of pellicle not observed ; cells 16-35 X 7-1 1ft.

Habitat : In plankton or as a green scum when abundant, in locations 28 (April), 29
(common-February ; September-November ; rare-April-May), 33 (April), 62
(common), 125 (rare) and 151 (rather common-December, February, MarbtL;nil-
May-July or August ; stray to rare- January, August ; common to very common :
April ; very abundant-September to November). •

Though Carter was the first to describe the species as E. agilis his description
.was considered as inadequate. In 1883 Klebs gave proper description and
figures for the species under the name E. piscifonnis and ever since thfe name
was widely used. Gojdics (1953) revived the name E. agilis. However, according
to Pringsheim (1956) the long usage of the specific name piscifonnis and the wrong
use of the name agilis by Baker in 1926 for another species, viz E. gracilis coupled
with Carter's inadequate diagnosis warrant the continuation of the name "piscifor-
mis instead of agilis. :

Distribution in Indian region : Maharashtra (Carter 1856 ; Hansgirg 1902-
as E. agilis). ; Andhra Pradesh (Naidu 1966) ; Kerala (Suxena et al 1973), Karnataka
(Dodkundi et al 1973) ; West Bengal, Orissa, Andhra Pradesh and Tamilnadu
(!) ; Burma (Skuja 1949).

Naidu (1966) has also recorded var. piriformis (Szb.) Gojdics of £•• agilis from
Cuddapah, Andhra Pradesh. Since the name pisciformis is more acceptable, this
taxon will have to remain in its original name of £. pisciformis var. piriformis
Szabados (measuring 16 x 12ft for which Naidu gives dimensions of 25 x Up
compared to 21-28 x 9-1 0/^ in his typical species).

11. Euglena granulaia (Klebs) Schmitz 1884 (figure 17)

Gojdics 1953 pp. 137-39, Plate 24, 'figure 4 a-g ; Pringsheim 1956 pp. 72-74
figure 14 ; Prowse 1958 p. 148 figure 1m; = E. granulata (Klebs) Lemmermann,
19lOa p. 501 ; 1913 p. 131 figure 211 ; Huber-Pestalozzi 1955 p. 85 figure 63.

Body fusiform with changes in the ratio of length to breadth caused by bulging
' slightly .in front, or behind the ..middle ; ji#erior_end more or less conical ; posterior

The genus Eugkna Ehrenberg 583

>nd double-narrowed and ending in a short nearly cylindrical hyaline tail which
s frequently beat to one side ; pellicle with markedly spiral striae ; chromatophores
arge, about 6-12, and more or less polygonal with slightly lobed margin and
.ppearing somewhat spindle-shaped to elongate in side view ; anterior end gene-
ally free of chromatophores but at posterior end they may well extend to the
audal tip ; each chromatophore with a pyrenoid having double sheaths of saucer-
haped paramylum ; additional small ovoid to slightly elongated paramylum
>odies frequently present in the cytoplasm ; nucleus spherical and median to sub-
nedian ; cell contents highly granular ; eye-spot small ; flagellum less than body
ength ; cells 50-65 x 11-14/*.

habitat : Rare in plankton of locations 5, 29 (April), 39, 73, 109, 110, 134 and 151
April, August) and 154 ; common in location 65.

According to Pringsheim (1956), though the species is reported very often it is
lot certain whether the identification has always been correct. Further, he states
;hat apart from the shape of the cell with double narrowing at posterior end,the
nost reliable feature for identification of the species is the regular spiral rows of
jub-cuticular mucus bodies along the striae which do not always show without
;reatment with vital stains, though Klebs1 reason for giving the name "granulata"
Tor his E. velata var. granulata which is synonymous to E. granulata) may have
Deen due to the highly granular nature of the cytoplasm. Johnson (1944) has also
recorded sub-cuticular granules parallel to the striae in this species. Pringsheim
ilso states that the flagellum. is not more than body length though some authors
state it is longer.

In the author's material the flagellum when observed was shorter than the body.
The spiral rows of mucus bodies could not be observed clearly since no vital staining
sould be done. However, the shape of the cell together with the elongated chromato-
phores having sheathed pyrenoids and the granular nature of the cytoplasm pointed
to the organism as being an unmistakable E. granulata.

Pringsheim (1956) observed two size groups, viz. 80-105 x 16-24/« and 60-83 x
18-25/f in his strains. Sizes reported by other authors (including Chu 1947) vary
from 50-115 X 11-30/j. Pringsheim states that Szabados's organism (37-77 x
20-25/0 is either a smaller variety or it may be an instance of wrong identity. As
already stated under E. vaginicola, Szabados has reported flask-shaped cysts in this
species.

Distribution in Indian region : Assam, West Bengal, Madhya Pradesh, Orissa,
Andhra Pradesh and Tamilnadu (!).

18. Eugkna caudata Huebner 1886 (figure X8a~c)

Lemmermann 1913 p. 133 figure 198 ; Chu, 1947, pp. 110-12 figure 21 ;
Gojdics 1953 p. 137 Plate 24, figure 2 and Plate 29, figure 3 ; Huber-Pestaiozzi
1955 p. 88 figure 67 ;' Pringsheim 1956 pp. 74-75 figure 15; Prowse 1958
pp. 147-48 figure 1.

Body spindle-shaped with the anterior end drawn out or nearly cylindrical ;
posterior end narrowing strongly into a short tail (the name "caudata" being
derived from this feature) ; pellicle with fine spiral striae ; regular rows of coloured
granules or small spheres seen at wider intervals along the striae (usually along

P. B— 12

584 Af T Philipose

alternate striae) on vital staining ; chromatophores about 8-10-15 or more*
parietal and disc-like with a smooth or slightly lobed margin, the posterior ones
being usually elongated, and with a sheathed pyrenoid ; additional small para-
mylum bodies frequently present ; nucleus nearly spherical and median ; eye-
spot prominent ; flagellum f to 1£ times body length ; cells 50-73 x 19-26/f.
Habitat : As a fairly thick scum or in the plankton after overnight heavy rains in
location 151 (October 1938).

The organism from Madras agreed fairly well with the typical species (see Pring-
sheim 1956) but the tail was slightly shorter. Also, in contrast to Pringsheim's
material in which the chromatophores were saucer-shaped and lobed, they were
disc-like to saucer shaped (the posterior ones being elongated) with smooth or
slightly lobed margin in the author's material. Further, Pringsheim stated that
the chromatophores extended to the anterior and posterior ends giving the entire
organism a green appearance. In the Madras specimens the posterior elongated
ones extended to the hind end whereas the front end was usually free of them.
The number of chromatophores reported also vary. Huebner (as cited by Pring-
sheim) gave it as 50, Chu as 6-30 and Pringsheim as more than a dozen whereas
in the author's material the maximum number noted was fifteen. Dimensions
given by different authors also vary. The usual size is 80-120 x 20-38-50/* (see
Huber-Pestalozzi 1955 and Pringsheim 1956). Chu's organism measured 70-115 X
10-39/f, rarely 60-90 x 10-1 5-20/*, the latter measurements being more like those
of var. minor Defl. (63 x 18/f) which is not recognized by some authors. The
author's measurements were 50-73 x 19-26/* compared to 30-49 x 16-2 1/* in var.
minor (see below). Naidu (1966) gave the dimensions of E. caudata as 55-77 X
15-17/j.

Reports on flagellar length also vary. Huebner gave it as the body length, Chu
twice the body length and Pringsheim less than the body length. As already
stated, in the author's material it was f to li times body length.

Chu (1947) observed osmophilic granules arranged somewhat spirally under
the pellicle in his E. caudata, and he stated that E. granulata which has such
granules is really synonymous to the former. According to Pringsheim (1956),
though this feature is more characteristic of E. granualta, Chu's organism is really
a form of E. caudata with such granules well developed, and he retains E. granulata
and E. caudata as distinct species with well defined characteristics. The Madras
material also showed the granules in some specimens whereas they were not clear
in the majority of specimens. There is no doubt that the Madras form belongs
to E. caudata because of its characteristic tail, chromatophores and other features
as given by Pringsheim.

Distribution in Indian region : Andhra- Pradesh (Naidu 1966) ; Maharashtra
(Kamat 1975) ; Tamilnadu (!).

Var. minor Defiandre 1924 (figures 18d-f)

Deflandre 1924 p. 1119 figure 7 ; Huber-Pestalozzi 1955 p. 88 figure 67A.

Differs from the typical species in its smaller size ; posterior part ending in a
sharp point or a very short tail ; pellicle striated with fine smooth spiral striae ;

The genus Euglena Ehrenberg 585

hromatophores 6-16 and discoid to saucer-shaped, and they are larger when
mailer in number ; flagellum about J to \ body length ; cells 30-49 x 16-2 1/*.

'iabitat : As a green scum in locations 29 and 30 (January)

Chu (1947), Gojdics (1953), Pringsheim (1956) and Prowse (1958) do not
eparate this variety from the typical species. Since the author's specimens
.ppeared to be markedly different in size, it is retained under Deflandre's variety.

Distribution in Indian region : West Bengal (!).

,9. Euglena gradlis Klebs 1883 (figure I9a-b)

Lemmermann 1910a p. 502 figure 17 (p. 483) ; 1913, p. 133, figure 190 ;
Gojdics 1953 p. 141 Plate 26 figure 1 ; Huber-Pestalozzi pp. 71-72 figure 48 ;
Pringsheim 1956 pp. 81-83 figure 19

tody elongate fusiform to cylindrical with very little difference between anterior
,nd posterior ends, the former being somewhat rounded and the latter bluntly
jointed ; chromatophores shield-shaped to triangular, varying in number from
H-10 or 12 and with a sheathed pyrenoid ; with additional small paramylum bodies
n the cytoplasm ; nucleus spherical and usually below median region but some-
imes nearly median ; eye-spot streak-like ; pellicular striae very faint ; flagellum
tot observed (reported to be body length) ; cells 36-47 x 8-1 3/z.
riabitat : In plankton and scum of locations 30 and 31 (rare).

The dimensions given by various authors (including Prowse 1962) range from
•1-68 X 6-18, rarely up to 22. 5/*. Pringsheim states that the average length is
tot far from 50, with 40 and 70/* as the limits. Kamat (1961-62) gives it as 60-75 x
6-30//, the maximum breadth being particularly higher than in previous reports,
lowever, in the absence of figures it is not possible to say whether his organism
5 really a E. gradlis.

Distribution in Indian region : Gujarat (Kamat 1961-62) ; Maharashtra (Kamat
975) ; Karnataka (Dodkundi et al 1973 ; Hosmani and Bharati 1980, 1980a);
Vest Bengal (!).

0. Euglena tuba Carter X869 non Johnson 1944 emend PWlipose (figure 20a-rand)
aa-nn)

lody more or less cylindrical or fusiform to ellipsoid rarely nearly spherical with
tie anterior end broadly rounded and the canal opening slightly towards one side
iving it a lipped appearance ; posterior part ending in a sharp blunt point or with
very short tail or more often broadly rounded ; changing shape markedly by
ulging in the middle or curving at anterior end or by complete rounding up ;
ellicle with fine punctate spiral striae ; chromatophores about 5-16 or more and
sually in elongated broad bands or somewhat spindle-shaped ; in cysts and
reshly liberated individuals often markedly curved with the free ends almost
suching each other ; when inside the cyst or markedly crowded in free living
idividuals, sometimes appearing as spheroid to discoid ; each chromatophore
tith a double sheathed pyrenoid, the paramylum being saucer-like ; additional small
isc-like paramylum frequently present in the cytoplasm ; nucleus spherical and
ledian ; eye-spot fairly small ; flagellum about 1/3 to 1/2 body length, but easily
tied ; cell usually with haematochrome pigments spread all over the cell during

. (W-13

586

M T Philipose

Figures 20a-r and 2Gaa-nn. EugleiuitubaC&iiw. 20a~r. individuals with haemato-
chrome. 20ba-mm, Gieen individuals without haematocliromc. 20nn. a mixture
of green and partly re df individuals. 20a, 20aa and 20nn. Clusters of cysts 20b-f,
i-j, 20bb-dd and ff. Cysts of various size with stalks of varying length and shape
j and m with discoid cliromatophores, the farmer killed in iodine-formalin; ee.
liberation of cell from cyst. 20k, I-n, gg~kk. freshly liberated individuals ; o-q, and
11 -mm. broken remains of cysts. 20r. an abnormally long stalk found associated
with the organism, [same magnification :— -(20a and 20nn) ; (20b~i, k-m, o-q,
20aa-gg, jj, H-mm) ; (20j) ; (20ns hh, ii and rr) ; (20r)].

The genus Euglena Ehrenbetg 587

bright sunshine giving it a brick red appearance or receding to the hind
end at other times giving it a dirty green colour ; haematochrome at the
hind end in encysted individuals ; cyst in the form of a round-bottomed
flask with a tube-like prolongation (stalk) which ends in a funnel-like expansion
(base) ; tube frequently striated transversely, short or sometimes quite long, and
rarely irregularly jointed ; funnel also some-times with bristle-like hairs at the
rim ; a number of cysts may remain together either in a mucilaginous matrix
with the tubes projecting all round or attached to each other by muclaginous pads
on either side of the flask, forming an irregular linear row ; encysted organism
always with the hind end towards the stalk and the front end towards the bottom
of the flask ; very rarely haematochrome absent altogether in encysted and free
living state ; flagellum absent in encysted condition ; liberation of the organism
by the rupture of the flask near its middle or nearer the stalk or by gelatinization
of the flask wall, the flagellum (at first short) appearing shortly after liberation
and progressively becoming longer ; locomotion by slow swimming when with
flagellum or by slow motion accompanied by change of shape when without
flagellum ; cells usually 60-74-96 x 18-30/^ or larger, but in nearly spherical
ones 41.5-56 x 36-43^ ; cysts 34-98/* long with flasks 31-47 X 26-47/^, stalks
up to 34-1 Q/J, or longer and 13-18/j broad ; base 29-32^ in diameter.
Habitat : As a red or green scum (neuston) or in general plankton and bottom silt,
abundant to very abundant throughout the year except during periods of heavy
rainfall in locations 24, 32, 66, 151 and in a number of ponds in locations 61 and
63 ; also abundant in locations 7, 51, 60, 69, 72, 73, 80 and 116 ; common in
locations 84, 123, 135 and 139 ; also observed in locations 8, 9, 21 (December)
45, 50, 104, 113 and 149. It was frequently associated with E. sanguined (inclu-
ding E. rubra) in location 151, and with E. sanguinea, E. viridis var. and E. tuba var.
yseudotuba f. minima (see below), and rarely with E. proxima and E. hemichromata
in location 61.

Though the organism observed in various localities had haematochrome invari-
ably, in two collections from N.P. 12 (location 61) made on 5-12-53 and 14-12-53
it appeared as a pure green scum during the bright hours of the day after fairly
heavy rains overnight. In this material, except for very stray individuals with a
±ace of haematochrome, none of the individuals had the pigment so that the inter-
aal contents including the eye-spot could be seen clearly (see figures 20 aa-nn).
However, the stalk of the cyst was usually shorter (up to 33/0 and 8.5-14 rarely
up to 1 6/^ broad just above the flask, and with greater bulge below the base.
Fhe cysts (33.5-61 x22.5-34.5jLf with base 18-31/0 and the free living cells.
;31-70 x 17-27.5/0 were also on the whole smaller. However, they did not
ippear to be materially different from normal individuals.

According to Pringsheim (1956 p. 34) the general notion that species like
E. sanguinea having red pigments may occur in a green state when well supplied
flith nutritive substances is confirmed by observations in cultures. Green species
ike E. gracilis might also produce red granules in a state of nutritive exhaustion.
Probably this might apply to E. tuba also.

Even though E. tuba is one of the most common species occurring in a number of
States in north-east, south and west India, and was first described by Carter in 1869
Tom Bombay, it is still a controversial species, partly due to Carter's description
md figures being inadequate. Apart from the flask-shaped cyst with a tube-like

588 M T Philipose

prolongation ending in a trumpet-like structure characteristic of the species, size
of cell about 83 /^ largest cyst 41 /*, shape of cell spindle-shaped to cylindrical
flagellum about 1£ times body length (as seen in his figure 13) and presence of
haematochrome, he stated that the internal structure of cell was as in E. viridis
with which it was associated. He could not also establish the real connection
between the encysted organism and the free living individual Further, he
thought that the flagellum extended to the tube of the cyst, and with the tubes
projecting down into the water, it served for aeration of the encysted organism.
Kent (1881 p. 385) had gone to the extent of suspecting that the encysted organism
of Carter may not even be connected with the free living one, and the former may
not be a Euglena at all.

la 1908, Kashyap recorded a red species with flask-like cysts in a pond at Lahore,
which he stated was like Carter's E. tuba, but his description of the organism as
cylindrical to oval ending abruptly in a short tail, chromatophores small and
round, paramylum numerous and of various sizes, flagellum body length, eye-spot
large, cell 62 X 150 to 125 x 3 l/jt and pellicle faintly striated, did not help in
knowing the exact nature of the organism he observed (though it might to some
extent fit in with the structure of E. tuba), especially because it was unaccompanied
by figures. Walton (1915) gave Kashyap's organism the name of E. orientalis
based on his description. In all probability it could be a E. tuba.

Johnson (1944) described an encysted Euglena from Iowa which he wrongly
referred to E. tuba Carter. Since Johnson's organism is slightly different from that
of Carter, and the name cannot stand, Gojdics (1953 p. 159) renamed it as E.peduncu-
lata (Johnson) Gojdics. However, the cyst of E. pedunculata, as figured in original
by Gojdics, appears to be different from that of Johnson, and Johnson's and
Gojdics' organisms may not be the same (see below).

Regarding E. tuba Carter, Gojdics (1953 p, 190) states that it is a doubtful species
and it could very well be a E. sanguinea, E. haematodes or E. orientalis all of which
possess haematochrome and may have stalked cysts. Pringsheim (1956 p. 139)
is also of the opinion that E. tuba Carter is a E. sanguinea. Though Huber-Pesta-
lozzi (1955 pp. 43-44, figure 17A a-i) retains Carter's species, his description is
based on Johnson's E. tuba and mostly his figures. In fact, only figures 17A h-i
(cysts) are those of Carter (as reproduced by Kent 1881). Further, he states that
Carter's species is doubtful one and that Johnson's species being different, the
latter is better named as E. pseudotuba.

Hortob&gyi (1960) referred an organism occurring in the nenston of fish ponds
of Hungary to E. tuba Carter. It was olive green, with smooth spiral striae and
measured 55-81 x 20*5-32ju:. The chromatophores were discoid, 9-numerous,
of varying size and without pyrenoid or paramylum. However, ha has not shown
cysts in his figures.

After studying the organism described as E. tuba by Carter (1869) over a number
of years and from many localities, and by making undisturbed mounts of the surface
scums of the living organism, the author has been in a position to observe details
not noted by previous workers, and to study the liberation of the organism from
the cyst. Since the organism was invariably mixed with other red forms, viz. var.
pseudotuba f. minima (see below) and E. sanguinea, observations including measure-
ments were made only from individuals freshly liberated from cysts. As a
result, the author is inclined to consider that Carter had actually mixed up E. tuba

The genus Euglena Ehrenberg 589

and E. viridis, his figures 13 in plate 17 of an individual in active state having a
pointed tail, long flagellum, etc., pointing more to a E. viridis than to E. tuba. For
the reasons stated above, it appears that E. tuba Carter is actually a well defined
species needing an emended description while the organism of Johnson (1944),
which differs mostly in the shape of the cyst, may be considered a variety of
E. tuba under the name var. pseudotuba (Johnson-Huber-Pestalozzi) comb. nov.
Distribution in Indian region : Maharashtra (Carter 1869) ; Andhra Pradesh ?
(as E. orientalis Walton-see Naidu 1966-60-91 x 18-23/0 ; Assam, West Bengal,
Orissa, Andhra Pradesh, Karnataka, Kerala and Tamilnadu (!) ; Pakistan?
(as Euglena sp. — see Kashyap 1908 ; = £. orientalis Walton 1915).

var. pseudotuba (Johnson-Buber-Pest.) comb. nov.

= E. tuba Johnson 1944 p. 120, figure 22 A-H, non Carter 1869 ; =
E. pseudotuba (Johnson) Huber-Pest, 1955 p. 44

Structure of body and other details more or less as in E. tuba Carter with the
differences that : (1) the cyst is conical (like a conical flask) to nearly cylindrical
to vase-shaped with the stalk almost absent and with a broad rim-like base ;
(2) a number of cysts often remain together with their bases almost fused (see
Gojdics 1953, p. 139, based on Johnson's personal communication) ; (3) reser-
voir (spherical) and eye-spot relatively larger ; (4) nucleus below median region ;
(5) flagellum nearly body length ; (6) pellicular striae delicate, but not punctate,
and (7) cells large, viz. 87-130 X 17-27/r, with cysts 34-50/*.

In Johnson's organism also the chromatophores were frequently discoidal (in
old laboratory cultures) as in the typical species.

This variety is known only from N. America.

f. minima f. nov. (figure 20al-sl)

Forma a var. pseudotuba differens ut minor (cellula 45-82-5 x 17-5-37-5/rai,
atque cysta 35-60 x 24-39/mi, basi 29-44/on, diam.) ; extremitas posterior cellulae
puncto obtuso aut minuta cauda nodiformi, aut late rotundata terminans ;
chromatophora solum c. 5-10 ; paramylum numerosius, maius et in cellula
encystata grana paramyli prope extremitatem posteriorem in ordinibus transversis
uno, raro doubus, disposita ; flagellum c. dimidium longitudinis corporis ; cystae
in corymbis irregularibus aut ordinibus linearibus confertis saepe aggregatae, aut
per fila mucosa inter so lateraliter connexae ; basis cystae saepe transverse
striata.

Habitatio : Plantae ut spuma in stagnis 15 et 16 loci 61.

Differs from var. pseudotuba in its smaller dimensions (cell 45-56-82-5 x 17-5-
31 • 5-37 -5/1 and cysts 35-60 X 34-39/j with base 29-44/j in diameter) ; reservoir
and eye-spot correspondingly smaller ; hind end of cell ending in a blunt point
or a tiny knob-like tail or sometimes broadly rounded ; paramylum more in
number, larger and in encysted cell those near hind end arranged in one (rarely two)
transverse rows ; flagellum about half body length ; cysts frequently grouped
together in irregular clusters or close linear rows or connected laterally by mucila-
ginous threads ; base of cysts frequently striated transversely.

M T Phillpose

Figures 20-24. 20al~dl, fl-sl. Euglena tuba var pseudotuba (Johnson-Huber-Pest)
f. minima f. nov. 20el. E. tuba and var pseudotuba f. minima associated together ;
20al, ql. Cysts showing linear arrangement 20pl. same with lateral attachment
by mucilage threads ; 2Qfl-gl. Clusters of broken remains of cysts 20rl. One
cyst with bifurcation of muc'lage thread before attachment 20hl-il. Liberation of
organism from cyst ; jl-fcl, II. Freshly liberated cells ; si. anterior end of a cell
showing reservoir, eye-spot, flagellum, etc. ; ml-nl. two abnormal cysts dl, ol.
broken remains of cysts, 21. E. oblonga Schmitz. 22 a~d. E. sanguined Ehr. 22 c,
showing cyst. 22 b, showing haematochrome along axial region (note chromato-
phores not shown.) 23a-b. E. viridis Ehr. var* maxima var. nov. ; 24. E. hemi-
chromata Skuja. Same magnification — (20al-bl, dl-rl-kl, ml, sl.22a, b
and d, 23b) ; (20fl~gl, al-rl, nl ; 22c) ; (20cl, 11 ; 23a, 24, 21.)

The genus Euglenct Ehrenberg 591

Habitat : As a scum in ponds 15 and 16, location 61 (October-December and
January-February).

In the encysted condition the haematochrome was restricted to the axial region
in contrast to the hind end in E. tuba. The nucleus and pellicular striae could
not be made out due to the haematochrome. Liberation of the organism either
by a break near the base (the remains of the cyst showing a toothed appearance
just below the base-see figure 20 cl) or in the median region of cyst. Though the
cells usually measured 56-82*5 x 17'5-31'5/J, those which were slightly twisted
at anterior end and bulged in the middle measured only 45-55 x 33-5-37*5^. As
in E. tuba, the flagellum which appeared only after liberation of cell became pro-
gressively longer for some time.

The organism was invariably mixed with E. tuba, E. sanguinea, E. viridis var.
and sometimes with E. proxima and E. hemichromata. In one or two instances, a
cyst of the form was attached to the cyst of E. tuba (see figure 20 el). It could
not be ascertained whether this was accidental or otherwise. Gojdi.cs(1953 p. 140)
also observed one individual of E. pedunculata attached to a copepod.

From Johnson's figure 22f (of a broken cyst) and 22 g (of a cyst) there appears
to be no doubt that the Indian form is more or less identical to the American
organism except for differences in dimensions and other minor details.

Distribution in Indian region : Orissa(!).

It appears to the author that'j?. pedunculata Gojdics (1953 pp. 139-40 plate 25-
figure 1 a-e) which she created in the place of Johnson's E. tuba, is actually some-
what different from Johnson's organism. It has thin-walled cysts which are like
round-bottomed flasks with a fairly long uniformly broad stalk and a broader base
(as figured in original by Gojdics) ; cells are liberated by a break on the side of the
cyst and not by a break along the median region, and the free living cells are some-
what fusiform with a very gradually tapering posterior in contrast to the nearly
cylindrical cell with slight posterior extension or rounded hind end in Johnson's
material Probably/ it could be considered as another variety of E. tuba under
the name var. pedunculata (Gojdics) comb. nov. with dimensions 90-120 x 18-22//.

21. EugUna oblonga Schmitz 1884 emend. lyengar 1962 (figure 21)

lyengar 1962 pp. 329-31, figure 9-31 ; = E. oblonga Schmitz 1884 ; Lemmer-
mann 1910a, p. 494 ; 1913 pp. 127-28, figure 184 ; Gojdics 1953 pp. 64-66
plate 2, figure 2 a-e ; Skuja 1948 pp. 186-88, plate 21, figures 16-19 ; Huber-
Pestalozzi 1955 pp. 44-45 figure 18.

Body ovoid to ellipsoid to nearly oblong with the anterior end rounded and
lipped ; posterior end rounded or ending in a short point ; chromatophores
appearing as numerous short bands arranged peripherally in spiral rows ; pyrenoids
and the two sheathing cap-like paramylum could not be made out due to
dark cell contents particularly on either side of the nucleus ; nucleus large
and median or slightly below ; eye-spot large and irregular in outline ;
flagellum body length or slightly longer ; pellicular striae (reported as spiral) not
observed ; cells 65-70 x 35-40/j.

ffabitat : Rare in plankton of locations 6 and 32,

592 M T Philipose

There is difference of opinion regarding the shape of the cell and the nature of
the chromatophore in this species. The markedly ovoid shape reported by
Schmitz and several others has been considered (see Gojdics 1953) as due to the
organism being in the resting stage. lyengar (1962) gives the shape as fusiform.
In the author's material the cell was ellipsoid to nearly obloag. However, in foreign
material observed by the author (Philipose 1948) most specimens in the abundant
collection were distinctly ovoid with band-like peripheral chromatophores bearing
distinct sheathed pyrenoids.

Lemmermann (1910a, 1913) stated that the numerous small chromatophores
which lie peripherally continue radially. Later (1913) the same author referred to
the chromatophores as star-shaped, probably following Schmitz. By suitable staining
techniques, lyengar (1962) showed that the chromatophores in the species are
really stellate and saucer-shaped with 10-12 radiating arms and a central double-
sheathed pyrenoid. The chromatophores numbering 16-25 are distributed in the
periphery with their arms running radially and spirally along the surface. The
organism measures 70-85 X 16-32/f with nucleus 13'5-15/z and paramylum 4 -5-
6 X 3-4. 5/J, and has strong spiral pellicular striae. The dimensions of the orga-
nism given by various authors range from 50-110 x 16-37/*, the maximum breadth
of 57/i given by Huber-Pestalozzi (1955) being probably a printing error.

Though the author's organism appeared in several respects like a E. oblonga,
in the absence of all details, it is only tentatively assigned to that species. The
organism was also slightly broader in relation to the length.

Distribution in Indian region : Tamilnadu (lyengar 1962) ; Assam and West
Bengal (!).

22. Euglena wnguinea Ehrenberg 1830 (figure 22a-d)

Lemmermann 1910a p. 494, figure 19 (p. 483) ; 1913 p. 128 figure 185 ;
Gojdics 1953 pp. 154-55 plate 31, figure 3 ; Huber-Pestalozzi 1955 pp. 90-93
figure 70 ; Pringsheim 1956 pp. 89-94, figure 22 ; including E. rubra Hardy
1911 ; Lemmermann 1913 p. 128 figure 188 ; Philipose 1940 p. 192 plate 6,
figure 120.

Body ellipsoid to spindle-shaped with the anterior end broadly rounded and the
posterior part tapering gradually to a point or a short knob-like tail ; pellicle with
clear spiral striae ; chromatophores band-shaped (ribbon-like) but broken at
intervals and lying peripherally as well as radiating towards the inside, but less
regularly arranged than in E. oblonga ; with sheathed pyrenoids ; chromato-
phores and pyrenoids frequently obscured by haematochrome, but the former
invariably clear at the periphery and hind end ; additional small ovoid to discoid
paramylum bodies present in large numbers in the cytoplasm ; eye-spot prominent
and usually granulated ; nucleus spherical and median or slightly below, but
frequently obscured ; flagellum usually longer than body ; cells 60-91-140 X
24-52, rarely up to 72/j ; cysts large, spherical and mucilaginous and up to
130/j in diameter ; encysted cells 48-62/j in diameter.

Habitat : As a brick red scum or in plankton, common in location 15, rare in
locations 29 (October), 34, 35, 101, 151 and abundant in location 61 (N",Pt 15
and 16, November-December).

The genus Euglena Ehrenberg 593

Though this species has been reported by many workers from all over the world
there is still a lot of confusion regarding its exact structure. This is because the
cells are rarely free of haematochrome or the species is sometimes found in associa-
tion with other red species. According to Pringsheim (1956) the chromatophores
are not elongate spindles as described by Gojdics (1953) but actually ribbon-like
with gaps here and there, and the ribbons are connected laterally by fine strands.
However, the author could not make out these strands. Pringsheim also states
that E. rubra Hardy with cells 200 x 60/f and E. haematodes (Ehr.) Lemm. (1910a
p. 495) with cells 75-103 x 28-36^, are in all probability only E. sanguined.
Forms resembling E. rubra observed by the author in localities 61 and 151 are also
considered here along with E. sanguinea. The dimensions given by various authors
for this species range from 55-121-150-170 x 22-33-55;*.

Distribution in Indian region : Maharashtra (Kamat 1974) ; Himachal Pradesh
(Kamat 1968a) ; Karnataka (Hosmani 1977) ; Assam, West Bengal, Orissa and
Tamilnadu (!). According to Biswas (1949 p. 101) the red and green scums of
Euglena occurring in India and Burma are due to E. sanguinea and E. viridis
respectively. However, he has not taken into consideration E. tuba (red or green)
and other red and green species.

IV Group RADIATE Pringsheim 1956

23. Euglena viridis Ehrenberg 1830

Lemmermann 1910a pp. 491-92, figure 2 (p. 483) ; 1913 p. 127 figure 189 ;
Gojdics 1953 pp. 70-71 Plate 4 figure 1 a,b ; Huber-Pestalozzi 1955 pp. 45-47
figure 19; Pringsheim 1956 pp. 102-105 figure 26; Hortob&gyi 1969 p. 31
figure 54.

Body fusiform with the anterior end broadly rounded and the canal opening
slightly towards one side making it lipped ; posterior end more or less conical
and ending in a point or in a small narrow cylindrical tail ; pellicle with faint
spiral striae ; chromatophores ribbon-shaped and radiating towards the periphery
from a pyrenoid centre situated above the nucleus ; nucleus nearly spherical and
in the posterior part ; ovoid paramylum bodies found mostly around the central
area of the chromatophore group, but some are dispersed in the cytoplasm ; eye-
spot small, but bright red ; flagellum usually longer than body ; cells 30-65-73
(-89) X 9-22/f.

The typical species not observed by the author, but reported by Carter (1856,
1858, 1869) from Bombay, by Bhatia (1930) from Kashmir, by Banerji (1936)
from lower Bengal, Singh (1960) and Hortobdgyi (1969—30-40 X 9. 5-1 1/0 from
Uttar Pradesh and Dodkundi et al (1973) from Karnataka.

var. maxima var. nov. (figures 23 a-b)

Varietas a specie typica differens ut maior, cellulis 69-94 x 28-4Qwm

Habitatio : Varietas in spuma planctove locorum 51, 84, 85, 87-88, 101 et 157
rara ; in loco 61 frequens.

Differs from the typical species in its larger size, the cells measuring 69-94 X
28-40/r.

Habitat : Rare in the scum or plankton of locations 51, 84, 85, 87-88, 101 and 157 ;

594 M T Philtpose

common in location. 61 (N.P. 15-16, Nov.-Dec.) along with E. tuba, var. pseudotuba
f. minima and E. sanguinea.
Distribution in Indian region : Orissa, Andhra Pradesh and Tamilnadu (!).

24. Eugkna hemichromata Skuja 1948 (figure 24)

Skuja 1948 pp. 185-86 Taf. 21 figures 10-13 ; Gojdics 1953, pp. 130-31,
plate 21, figure 6 a-b ; Pringsheim 1956 pp. 107-8, figure 30.

Cell spindle-shaped to cylindric-spindle-shaped or somewhat bulged in the
middle due to " metaboly " while in motion ; anterior end truncated obliquely or
rarely, more or less rounded ; posterior end tapering to a blunt point or ending in
a small hyaline tail ; pellicle with fine faint spiral striae ; chromatophores numerous,
parietal, and in the form of spherical to irregular discs as well as elongated ones
radiating from the middle of the cell where occasionally a central paramylum
group is present ; chromatophores either restricted to the posterior two-thirds of
the cell or, sometimes, filling the whole cell and without pyrenoids ; paramylum
numerous, short-cylindrical to ovoid or as smaller granules distributed all over
or sometimes crowded at the anterior end ; eye-spot bright red ; flagellum 1/3 to
3/4 body length ; nucleus, reservoir and gullet not observed ; cells 63-86 X 17.5-
23.5, rarely up to 29.5//.

Habitat : Rare in the scum of location 61 (N.P. 15-February) along with E. tuba
and var. pseudotuba f. minima, and E. proxima.

Skuja gave the name hemichromata on the basis that the chromatophores are
mostly restricted to the posterior half or two-thirds of the cell. According to
Gojdics (1953) they may be found in the whole cell. Pringsheim. (1956) states that
the absence of chromatophores at the anterior end in Skuja's material is due to
the crowding of paramylum in that region. Both Skuja and Pringsheim place
this species near E. vtridfs because of the radial arrangement of the elongated
chromatophores from the middle of the cell.

The author's organism agreed in most respects with Skuja's. However, the cells
were slightly broader especially when changing shape. Skuja gives the dimen-
sions as 62-128 x 12-22ju, which according to Pringsheim (1956) shows an inhomo-
geneity in the material. Paramylum in links (as seen in Skuja's drawings) were not
observed. Eye-spot was also slightly smaller than in Skuja's organism.

Distribution in Indian region : Orissa (!).

Other species of Euglena recorded from the Indian region are as follows :

L E. anabaena Mainz 1926 var. minima Mainz 1927

K V Naidu 1966 p. 26 figure 12. Location Cuddapah, (AP)

2. E. brevicaudata Gojdics 1953

K V Naidu 1962 p. 90, figure 7. Location Cuddapah (AP)

According to Pringsheim (1956) though a good species, the use of the same

name by Schiller has priority

3. E. choretes Schiller 1952

Kamat 1975 p. 466 ; Kamat and Frietas 1976 p. 121. Location Vidarbh
Nagpur respectively

The genus Euglena Ehrenberg 595

4. E. deses Ehrenberg 1833

H J Carter 1856, E A Gonzalves and D B Joshi 1946 plate 5, figure 9.
Location Bombay

5. E. ehrenbergii Kiebs 1883

E A Gonzalves and D B Joshi 1946 plate 5, figure 7. Location Bombay

6. E. elastica Prescott 1951

K V Naidu 1962 p. 90 figure 4-6. location Cuddapah (AP) ; Dodkundi
ct al 1973 p. 141 ; S P Hosmani and S G Bharati 1975 p. 151 ; 1980
p. 32,198; location Dharwar

7. E. elongata Schewiakoff 1893

R K Pandhol and I S Grover 1976; location Ludhiana, Punjab

8. E. flava Dangeard 1901

K V Naidu 1966 p. 26 figure 11. Location Vijayawada (AP)

9. E. fundoversata Johnson 1944

K V Naidu 1966 p. 24 figure 1 ; Location Vijayawada (AP)

0. E. gaumei Allorge et Lefevre 1925

N D Kamat 1975 p. 466. Location Vidarbh, Maharashtra

1. E. gibbosa Schiller 1952

N D Kamat 1964 p. 9 ; N D Kamat and J F Frietas 1976. Location
Bombay and Nagpur respectively

2. E. guentheri Gojdics 1953 var. aJpina (Grandori) Gojdics 1953.

K V Naidu 1966 p. 26 figure 14. location. Vijayawada (AP)

3. £. heimii Lefevre 1934

N D Kamat 1974 p. 26; Kamat 1963 p. 263. location Marathwada and
Kolhapur respectively

Both Gojdics (1953) and Pringsheim (1956) consider this species as synony-
mous to E. ehrenbergii Klebs ;

4. E. intermedia (Klebs) Schmitz 1884

T Hortob£gyi 1969 p. 30 plate 4, figure 52, location UP near Delhi.
Pringsheim (1956) considers that this species (= E. deses var. intermedia
Klebs 1883) cannot really be maintained as a species distinct from E. deses

5. E. maharastrensis Kamat 1963

N D Kamat 1963 p. 263 plate 13 figure 68-69, location Kolhapur
A species with brown knobs on the periplast.

6. E. mucifera Mainz 1926

N D Kamat 1975 p. 466. Location Vidarbh, Maharashtra
1. E. multiformis Schiller 1952

N D Kamat 1963 1975. Location Kolhapur and Vidarbh respectively

8. E. obtusa Schmitz 1884

V Venkateswarlu 1976 p. 673 plate 4, figure 51. Location Hyderabad (AP)

9. E. polymorpha Dangeard 1901

Hortob^gyi 1969 p. 30 plate 4, figure 51; location UP near Delhi;
V Venkateswarlu 1976 p. 673 plate 4, figure 52, location Hyderabad (AP)
S P Hosmani and S G Bharati ; 1980a p. 33. location Dharwar
!0. E. pringsheimii lyengar 1962

MOP lyengar 1962 p. 325 figures l-8? location Madras

596 M T Philipose

21. E. pusilla Playfair 1921 var. longa Playfair 1921

N D Kamat 1964 p. 9, location Bombay. Both the type and var. consi-
dered doubtful by most authors due to inadequate descriptions ;

22. E. rustica Schiller 1952

N D Kamat 1963 p. 264, location Kolhapur

23. E. sacculiformis Schiller 1952

N D Kamat 1963 p. 264. location Kolhapur

24. E. splendens Dangeard, 1901

K V Naidu 1966 p. 26, figure 13, location Cuddapah (AP)

25. E. texta (Duj.) Huebner 1886

= Lepocindis texta (Duj.) Lemmermann 1901 ; as Crumenula texta
Duj. see H J Carter 1856 p. 119, plate 6, figure 53, 56, 60 and 60a.
Location Bombay

26. E. tuber culata Drezepolski 1925

N D Kamat 1961-62, 1963. Location Ahmedabad, Gujarat and Kolhapur,
Maharashtra respectively

27. E. spathirhyncha Skuja 1948

K V Naidu 1966 p. 24, figure 3, location Cuddapah and Hyderabad (AP)

28. E. subehrenbergii Skuja 1948

T Hortobagyi 1969 p. 31 plate 4, figure 49, location UP near Delhi

29. E. velata Klebs 1883

Dodkundi et al 1973 p. 141, location Dharwar

Quite a number of these species are only just listed, sometimes with measure-
ments, or when described and illustrated, complete details are lacking or the illustra-
tions are not satisfactory. However, lyengar (1962) and Hortobagyi (1969) have
given good descriptions and illustrations for the taxa recorded by them. E. texta
(Duj.) Huebner is also a well-known species. A good number of the remaining
records may have to be re-investigated.

As already stated elsewhere, E. estonica and E. charkowiensis have been consi-
dered in this account under E. oxyuris, E. orientalis under E. tuba and E. agilis
under E. pisciformis.

E. alata Thompson 1938 (see Kamat and Frietas 1976 p. 121) has been
considered by Pringsheim (1956) as synonymous to E. tripteris, the species
differing from the latter only in its paramylum being in links. Pringsheim also
treats E. archaeo-plastidiata Chadefaud 1937 (see Naidu 1966 p. 25 figure 2)
as synonymous to E. pisciformis while E. minima France 1894 and E. minuta
Prescott (1944) are treated by him as probably a variety of/ or related to
E. pisciformis respectively. Skuja (1949) and Kamat and Frietas (1976) have
reported E. minima and E. minuta respectively from Burma and Maharashtra.

Acknowledgements

The author wishes to express his gratitude to the Late Prof. MOP lyengar under
whose valuable guidance the ecology and systematics of the species relating to
Museum Pond, Madras, were studied ; the Director, Central Inland Fisheries

The genus fiugtena Ehrenberg 597

Research Institute, Barrackpore for kind permission to publish the data collec-
ted by the author while at the institute ; Sarva-Sri S M Banerjea and V Rania-
chandran, Fishery Scientists, CIFRI for some of the chemical analytical data ;
Dr Hannah Croasdale, Department of Biological Sciences, Dartmouth College,
Hanover, New Hampshire, USA., for the latin translations of diagnoses of new
taxa and for some valuable suggestions in naming new species ; and Sri T
Ramaprabhu, Fishery Scientist, CIFRI (now at University of Florida) for help
in securing a copy of L H Johnson's paper on Euglena.

References

Allorge P and Lef&vre M 1930 Algues de Sologne ; Bull. Soc. hot. Fr. 77 122-50, 132 figs.
Banerji J C 1936 Studies on the Myxophyceae of lower Bengal ; J. Indian hot. Soc. 15 285-302
Bernard C 1908 Protococcacees et Desmidi&es d'eau douce recoltees & Java. Dep. de Uagric. aux

Indes Necrlandaises, Batavia, 230 p. 16 pis
Bliatia B L 1930 On some freshwater rhizopods and flagellates from Kashmir ; Arch. Protistenk.

72 359-64
Biswas K 1949 Common fresh and brackishwater algal flora of India and Burma-I ; Rec. bot.

Surv. India 15 1-105, 10 pis
*Bourrelly P 1949 Euglena oxyuris Schmarda et formes affines ; Bull. Mus. Hist. nat. Paris Ser. 2

21 612-16, 2 figs
Carter H J 1856 Notes on the freshwater infusoria on the island of Bombay-I. Organisation ;

Ann. Mag. nat. Hist. Ser. 2, 18 115-32 ; 221-28, 3 pis
Carter H J 1858 On fecundation in Eudorim elegans and Crypfoglena ; Ann. Mag. nat. Hist.

Ser. 3, 2 237-53
Carter H J 1859 On fecundation in two Volvoces and their specific differences ; Ann. Mag.

nat. Hist. Ser. 3 3 1-20, 1 pi
Carter H J 1869 Notes on filiferous green Infusoria on the island of Bombay ; Ann. Mag.

nat. Hist. Ser. 4, 3 249-60 1 pi

Chu S P 1947 Contributions to our knowledge of the genus Euglena ; Sinensia 17 75-134
Deflandre G 1924 Additions £ la flare algologique des environs de Paris ; Bull. Soc. bot. Fr.

71 1115-1129 1 pi
Dodkundi G B, Patil H S and Rodgi S S 1973 Ecological aspects of stratification in sewage

stabilization pond ; /. Karnatak Univ. Set. 18 1 37-44
Drezepolski R 1925 Przyczynek do znajomosci polskich Euglenin (Supplement £ la connaissance

des Eugleniens de la Pologne) ; Kosmos (Lwow) 50 173-270
Fritsch F E 1935 Structure and Reproduction of the Algae ; Vol. I p. 791 (Cambridge :

University Press)
Fritsch F E and Rich F 1913 Studies on the occurrence and reproduction of British freshwater

algae in nature — 3. A four years observation of a freshwater pond ; Rec. Inst. zool.

Torley-Rousseau 6 31-115

Gojdics M 1953 The genus Euglena ; (Madison : Univ. Wisconsin Press) p, 268 pi 39
Gonzaives E A and Joshi D B 1943 The algal-flora of temporary waters around Bombay-I ;

/. Univ. Bombay NS 11 34-45

Gonzaives E A and Joshi D B 1943a /. Univ. Bombay 11 121-128
Gonzaives E A and Joshi D B 1946 Freshwater algae near Bombay ; J. Bombay nat. Hist.

Soc. 46 164-176 7 pis

Hansgirg A 1902 Algologische Schlussbemekungen ; S.B.K. bohm. Ges. Wiss. 14 6-15
Hortob£gyi T 1960 Az Euglena tuba H J Carter neuston vizviragzasa. Az Egri Pedag. Foisk.

Fuzetei 197 55 10 figs
Hortob£gyi T 1969 Phytoplankton organisms from three reservoirs on the Jamuna river, India ;

Stud. biol. Acad. Sclent, hung. Budapest pp. 7-75 36 pis
Hosmani S P 1977 Notes on the occurrence of Euglena sanguined Ehrenberg at Dharwar ;

/. Karnatak Univ. Sd. 22 122-124

598 M T Phitipose

Hosmani S P and Bharati S G 1975 Hydrobiological studies in ponds and lakes of Dharwar-

III. Occurrence of two Euglenoid blooms ; /. Karnalak Univ. Sci. 20 151-156
Hosmani S P and Bharati S G 1980 Algae as indicators of pollution ; Phykos 19 23-26
Hosmani S P and Bharati S G 1980a Limnological studies in ponds and lakes of Dharwar—

Comparative plankton ecology of four water bodies ; /. Karnatak Univ. Sci. 19 27-43
Rowland L 1931 A four years' investigation of a Hertfordshire pond ; New Phyt. 30 221-265
Huber-Pestalozzi G 1955 Das Phytoplankton des Susswassers— Systematik und Biologte ; Die

Binnengewasser 16 Euglenophycean 606 p, 114 Taf. Stuttgait
lyengar MOP 1962 Euglena studies from Madras ; Arch. MikrobioL 42 322-32
Johnson L P 1944 Euglenae of Iowa ; Trans. Am. Microsc. Soc. 63 97-135
Kamat N D 1961-62 The Euglenophyceae of Ahmedabad ; /. Univ. Bombay 30 15-21
Kamat N D 1963 The algae of Kolhapur, India ; Hydrobiologia 22 209-305
Kamat N D 1964 The Euglenophyceae of Bombay ; /. biol Sci. "/ S-14
Kamat N D 1967 The algae of Mount Abu ; Proc. Rajasthan Acad. Set. 11 49-54
Kamat N D 1968 Algae of Alibali, Maharashtra ; /. Bombay nat. Hist. Soc. 65 88-104
Kamat N D 1968a Algae of Simla ; /. Bombay nat. Hist. Soc. 65 27-77
Kamat N D 1974 Algae of Marathwada ; Phykos 13 22-32

Kamat N D 1975 Algae of Vidarbh, Maharashtra ; J. Bombay nat. Hist. Soc. 72 450-476
Kamat N D and Frietas J F 1976 A check list of Euglenophyceae and Chlorophyccae of

Nagpur, Maharashtra ; Phykos 15 121-125

Kashyap S R 1908 Notes on a peculiar form of Euglena ; Rec. Indian Mus. 2 111-112
Kent W S 1881 A Manual of Infusoria Vol. I, pp. 271-77 (London)

Lemmermann E 1910 Beitrage zurKenntnis der Plankton-algen-II ; Beih. hot. Zbl. 76 150-156
Lemmermann E 1910a Algen-I (Schizophyceen, Flagellaten, Pridineen) ; Kryptogamenflom

der Mark Brandenburg 3 1-712 (Euglena pp. 485-503) Leipzig
Lemmermann E 1913 *' Euglenineae " in A. Pascher's Die Susswasserflora Deutschlands,

Ostemichs und der Schweiz, Heft 2, Flagellaten -2, pp. 115-174, figs. 181-377 Jena
Lind E M 1938 Studies on the periodicity of the algae in Beauchieff ponds, Sheffield , /. Ecol.

26 257-274
Lund J W G 1943 The marginal algae of certain ponds with special reference to the bottom

deposits ; /. Ecol. 30 245-283
Munawar M 1970 Limnological studies on the freshwater ponds of Hyderabad, India-JI. The

Biocoenose. Distribution of unicellular and colonial phytaplankton of polluted and

unpolluted environments ; Hydrobiologia 31 105-138
Naidu K V 1962 Studies on the freshwater protozoa of South India ~L Euglenoidina ; /. zool.

Soc. India 14 88-92
Naidu K V 1966 Studies on the freshwater protozoa of South ludia — III. Euglenoidina-2 ;

Hydrobiologia 27 23-32
Pandhol R K and Grover I S 1976 Algal flora of Ludhiana and its adjacent area ; Phykos

15 23-32
Philipose M T 1940 The ecology and seasonal succession of algae in a permanent pool at

Madras ; M.Sc. Thesis, Univ. of Madras, 220 pp., 18 text figs., 8 pis
Philipose M T 1948 Problems in algal ecolagy-L The ecology of the marginal algae of two

Hampstead ponds with special reference to their bottom deposits ; Ph.D. Thesis, London

Univ. 134 pp, 18 text figs, 5 pis
Philipose M T 1960 Freshwater phytoplankton of inland fisheries In P Kachroo (ed.) Proc.

Symp. algol ICAR New Delhi pp. 272-291

Playfair G I 1921 Australian freshwater flagellates ; Proc. Linn. Soc. N.S.W. 48 206-228 20 figs
Pringsheim E G 1956 Contributions towards a monograph of the genus Euglena. Nova Acta

Leopoldina N.F. 18 1-168 44 figs

Prowse G A 1958 The Euglenineae of Malaya ; Gardens Bull. Singapore 16 136-204 8 figs
Prowse G A 1962 Further Malayan freshwater flagellata ; Gardens Bull. Singapore 19 105-40

5 pis
Rino J A 1972 Contribute para o conhecimento das algas de agua doce de Mosambique-III ;

Revista Ciencias Biol A 5 121-264 32 pis
Singh V P 1960 Phytoplankton ecology of the inland waters of Uttar Pradesh In P Kachroo

(ed.) Proc. Symp. algol. ICAR New Delhi pp. 243-271

The genus Euglena Ehreriberg 599

Skuja H 1948 Taxonomie des Phytaplanktons einiger seen in Uppland, Schweden Symb. Bot.

upsal 9 1-399, 39 Taf
Skuja H 1949 Zur Siisswasseralgen-flora Burmas : Nova Acta Reg. Sclent. UpsaL Ser 4 14

1-188 37 Taf.
Skvortzov B W 1937 Contributions to our knowledge of the freshwater algae from Rangoon,

Burma, India : Arch. Protistenk. 90 68-87
Suxena M R 1955 Freshwater Euglenineae from Hyderabad, India : /. Indian hot. Soc. 34

429-50, 59 figs
Suxena M R, Venkateswarlu V, Subba Raju N and Rao V S 1973 The algae and testacta of

Cranganore, Kerala State, India; /. Indian hot. Soc. 52 316-341
*Swirenko D O 1915 Zur Kenntnls dec russischen Algen-flora-II. Euglenaceae (excl. Trachelo-

mows) : Arch. Hydrobiol. 10 321-340
Venkateswarlu V 1960 An ecological study of the algae cf the River Moosi, Hyderabad (India)

with special reference to water pollution. Factors influencing the distribution of algae ;

Hydrobiologia 33 352-63
Venkateswarlu V 1976 Taxonomy and ecology of algae in the River Moosi, Hyderabad, India -I.

Ghlo.ro phyceac, Cyanophyceae and Euglenophyceae : Nova Hedwigia 27 661-688
Venkateswarlu V 1980 Ecology of algal blooms — a comparative study : Indian J. bat. 4 31-36
* Walton L B 1915 The Euglenoidina of Ohio. Ohio biol Surv. Bull. 419 343-450
Zafar A R 1959 Two years' observation on the periodicity of Euglenineae in two fish-breeding

ponds : /. Indian hot. Soc. 38 549-560

*"Ntot seen in original.

AUTHOR INDEX (Plant Sciences)

imar
a Udar

^avinder K
hoda S P

131

55

behaviour during heartwood farma-
Acacla auriculiformis A. Cann 107

r

ses of cotton-cultivars to ]« ag day
ms 3g9

(Wt.) King— A reinvestigation with anatomical

evidence 115
Gupta Prem

Changes in proteins, amina and keto-acids

in different seedling parts of Cyamopsis

telragonolobus Linn, during growth in light

and darkness 417

Hiremath B S
see Chikkannaiah

Jaggoo MIS

479

i K

o N K Srinivasa 37

S

a I S 501

R

ir arbuscular mycorrhiza in sub-
. aquatic and marshy plant com-
>s 69

Laiah P S •

logical studies in Launaea nudicaulis

519

P

^logical studies in three species of
pogon Spreng (Poacea) 55

P S

Inamdar J A
see Mohan J

S S

519

189

R

aidu K Ramamurthy 433

I A S .

of CO2 in overcoming self-incompatibi-
arriers "in Bmssica campestris L. var.

227

ra Kumar

im Udar 139

famy R

10 N K Srinivasa 37

of morphactin, AMO-1618 and DPX-
on the endogenous levels of hormones
ts implication on apical dominance in
\e max Linn,

AKM .

st of extension growth and flowering
he cambial activity of Delonix regia

Janaki Bai A
Airborne pollen grains of Visakhapatnam :

A combined field and air sampling study 329

Khan A M

see Lai J 235

Kirti P B

Chromosome relationships of spinous soia-

nums ^3

Kodandaramaiah J

see Rao P Gopala

see Rao P Gopala
Kohli U K

see Dua I S
Kothari I L

see Rangaiah S
Kuriachan P I

see Rajeev K

183

495

501
297
319

Lakshmanaperumalsamy P
Heterotrophic bacteria associated with sea-
weed «7

Lai J

Pharmacagnosy of the stems of Portidaca
quadnfida L. and Portuiaca oleracea L. 235

227

115

93

XVII Novelties in

55 All vxjrb'VA~'"— -

'istylis (L.) Vahl and their vegetative

ity of Ficus macrocarpa Wt. ex King
, amplocarpa nom. nov.) and F. guttata

Malik C P

see Dhaliwal A S
Masilamoney P

see Govindarajalu E
McDonald D

see Subrahiaanyara P
Mehetre S S

Anther and pollen development in Cotton

haploids and their parents 409

Mishra R R

see Chaubal R 69

Mittcr Harsh

see Chode S P 55

Mohan J S S

Leaf architects of apocyaaceae 189

Index

Mohan M R R

Studies on Beggiatoa: Distribution and growth
in aquatic habitats of Visakhapatnam 159

Mohini Gupta

Petal venation in Tngonetta (Papilionaceae)

379

Mohan Ram H Y

see Pardha Saradhi P 101

see Rao I V Ramanuja 371

Mukherjee D
see Prem Gupta 417

Naidu K Ramamurthy
Leaf proteinaso and nitrate reductase acti-
vities in relation to grain, protein levels and
grain yield in four species of grain amaranth

433

Nair V J

Chandrasekharaflia : A new genus of Poaceae
from Kerala, India 79

Nanda K K
see Singh Kushal 175

Narayanaswamy S
see Rao N K Srinivasa 37

Ninan C A

see Rajeev K 319

Pai R M
see Vaikos N P 351

Pardha Saradhi P

Correlated promotion of ray — floret growth
in chrysanthemum by potassium chloride
gibberellic acid and sucrose 101

Parveen Farooqui (nee Kidwai)
Ontogeny of the paracytic stoma : Varia-
tions and modifications 145
Cork- warts in Eucalyptus species 289

Patel J B
see Bhat K V 107

Philip V J
see Raghavan P 465

Philipose M T

Contributions to our knowledge of Indian
Algae-III. Euglenineae-Part I. The genus
Euglena Ehrenberg 551

Purushothaman A
see Lakshmanaperumalsamy P 487

Raghavan P

Morphological and metabolic changes in the
egg and zygote of Lagerstroemia speciosa. I.
Cell size, vacuole and insoluble polysaccha-
rides . 465

Rajeev K

Turner a ulmifolia var. elegans X T. ulmifolia
var. angustifolia crosses and its bearing
on the taxonomy of the species 319

Ramayya N
see Rao S Raja Shanmukha 509

Ramachandran V S
see Nair V J 79

Ramakrishnan P S

see Saxena KG 61

see Ram Boogh 397

see Singh Jasbir 241

see Singh Jasbir 255

see Singh Jasbir 269

Ram Boogh

Seed germination and seedling es;ablishment
of two closely related Schima species 397

Ramesh C R

Viability and infectivity cf zoospores ot
Sclerospora graminicola (Sacc.) Schroet in
the soil 303

Ram Udar

The genus Jackiella in South India 131
Geocalyx Nees— a rare marsupial genus
from India 139

Rangaiah S
Pericarpial sclerieds in some Mimosaceae 297

Rao A Narayana
see Mohan M R R 159

Rao B G S
see Kirti P B 83

Rao I V Ramanuja

Effect of water stress on opening and longevity
of flowers in Gladiolus 371

Rao M Gopaia
see Shah G L 319

Rao M R K
see Bhatt J G 389

Rao N K Srinivasa

Regeneration of plantlets from callus cf
Eleitaria cardamomum Maton 37

Rao P Gopaia

Interaction of kinetin with B group vitamins
on the seedling growth of green fcram
(Phaseolus radiatus L.) 183

Association of chlorophyll content, phyllo-
taxy, photosynthesis and B group vitamins
in some C3 and C4 plants 495

Rao S Raja Shanmukha
Taxonomic importance of epidermal charac-
ters in the Indian Thespesia Corr. (Malvaceae)

509

Reddi C Subba
see Janaki Bai A 329

Safeeulla K M

see Ramesh C R 303

see Shetty Amarnatha 427

Saggoo MIS

Cytological studies on certain acanthaceae

from Central Indi$ 47

Index

mdhu D K

see Singh S 153

ixena A K

Quantitative profile structure of certain forests
in the Kurnaun Himalaya 529

ixena K G

Reproductive efficiency of secondary succes-
sional herbaceous populations subsequent to
slash and burn of sub-tropical humid forests
in north-eastern India 61

jethambaram Y

see Naidu K Ramamurthy 433

>kar G

see Subramanian C V 1

^shavatharam V

A contribution to the embryology of
Alysicarpus monilifier D.C. 9

lah G L

see Rangaiah S 297

Initiation, development and structure of
root nodules in some members of the tribe
Trifolieae (Papilionaceae) 309

Lamima Hashmi

see Ghouse A K M 201

tanta Mehrotra

see Usha Shome 211

lanthamma C

Apomixis in Cenchrus glaucus Mudaliar et
Sundaraj 25

larma G D

see Chaubal R 69

larma H P

see Usha Shome 211

Leaf surface studies of some medicinal
salvias 449

letty Amarnatha

Effect of ridge gourd pollen on zoospore
germination of Pseudoperonospora cubensis
and its significance in epidemiology 427
tietty H S

see Shetty Amarnatha 427

ingh Dalbir

see Singh Tribhuwan 357

ingh Jasbir

Structure and function of a sub-tropical
humid forest of Meghalaya I. Vegetation,
biomass and its nutrients 241

Structure and function of a sub-tropical
humid forest of Meghalaya II. litter dynamics
and nutrient cycling 255

Sturcture and function of a subptrapfcal
humid forest of Meghalaya III. Nutr/ent
flow through water 269

Singh J S
see Saxena A K 529

Singh Kushal

Photoperiodic control of extension growth,
bud dormancy and flowering of Nerium
indicum Mill, and Thevetia pemviana Schunu

175

Singh S

Growth response of some thennophilous
fungi at different incubation temperatures

153

Singh Tribhuwan

Transmission of seed-borne inoculum of
Macrophomina phaseolina from seed to
plant 357

Sreekumar P V
see Nair V J 79

Srivastava S C
see Ram Udar 139

Subrahmanyam P

Groundnut rust— its survival and carry-over
in India 93

Subramanian C V

Thaxteriellopsis lignicola and its Moorella
anamorph 1

Surinder Kumar
see Singh Kushal 175

Tilak V D
see Umavathj Hegde

281

Umavathi Hegde

Anatomy of the seedling of the Legumi-

no?ae— I 281

Usha Shome

Pharmacognostic studies on the flower of

Mesua ferrea L. 211

see Sharma H P 449

Vaikos N P

The floral anatomy of Kniphofia uvaria Hook.

(Liliaceae : Kniphofieae) 351

Venkateswarlu T

Non-inheritance of isomerism in cocoyams

17
Vidyavati

Cell division in Staurastrum gradle Ralfs.

under the scanning electron microscope 44$

PROCEEDINGS OF THE

INDIAN ACADEMY OF SCIENCES

(Plant Sciences)

VOLUME 91, 1 982

INDEX

THE INDIAN ACADEMY OF SCIENCES

BANGALORE 56O O8O

feedings (Plant Sciences)

Volume 91, 1982

SUBJECT INDEX (Plant Sciences)

na auriculifonnis

iclear bchavio.ur during hcartwood forma-

»n ia Acacia anricidifornns A. Cann 107

ithaceae

Biological studies on certain acantliaceae

>m Central India 479

ptation

^productive efficiency of secondary succes-

>nal herbaceous populations subsequent to

ish and burn of sub-tropical humid forests

north-eastern India 61

iQrne pollen
rborne pollen grains of Visakhapatnam :

combined field and air sampling study

329
icarpus

contribution to the embryology of Alysi
rpus monllifer D.C, 9

no acids

aangesin proteins, amino and keto-acids
different seedling parts of Cyamopsis
iragonolobus Linn, during growth in light
id darkness 417

morph

ixterieUopsis lignlcola and its Moorella
.amorph 1

;omy

entity of Fie us macrocarpa Wt. ex King
= F. amplocarpa nom. nov.) and F. guttata
rt.) King— A rei nves tigation with anatomical
idence 115

af architecture of Apocynaceae 189

ropogoneae

ibryological studies in three species of
>mbopogon Spreng (Poaceae) 55

ter tapetum

ither and pollen development in cotton
ploids and their parents 409

a I dominance

ect of morphactin, AMO-1618 and DPX-
10 on the endogenous levels of hormones
I its implication on apical dominance in
ycine max Linn. 501

synaceae

af architecture of Apocynaceae 189

ipory

jomixis in Cenchrus glauctis Mudaliar et
rtdafaj 25

Aquatic habitats

Studies on Beggiatoa : Distribution and
growth in aquatic habitats of Visakhapatnam

159

Arachis hypogaea L.

Groundnut rust — its survival and carry-over
in India " 93

Atmospheric biopallutants
Airborne pollen grains of Visakhapatnam :
A combined field and air sampling study 329

Bacterial thread

Initiation, development and structure of
root nodules in some members of the tribe
Trifolieae (Papilionaceac) 309

Bactcroid zone

Initiation, development and structure of
root nodules in some members of the tribe
Trifolieae (Papilionaccae) 309

Beggiatoa

Studies on Beggiatoa : Distribution and
growth in aquatic habitats of Visakhapatnam

159

Bioassay

Vesicular arbuscular mycorrhiza in sub-
tropical aquatic and marshy plant communi-
ties 69

Biomass

Structure and function of a sub-tropical
humid forest of Meghalaya 1. Vegetation,
biomass and its nutrients 241

Boll

Responses of cotton-cultivars to long day
conditions 389

Brassica campestns L. var. taria
Effect of COjj in overcoming self-incompati-
bility barriers in Brassica campestns L. var.
tori a 227

Bryophyta

The genus Jackiella in South India 131
Geocalyx Nees — a rare marsupial genus
from India 139

Bud dormancy

Phctoperiodic control of extension growth,
bud dormancy an4 flowering of Nerium
indicum Mifl. arid TheVetia peruviana Schum

175

P.(B)-15

n

Index

B- Vitamins

Interaction of kinetin with B group vitamins
O,D the seedling growth of green gram
(Phaseolus radiatus L.) 183

Association of chlorophyll content, phyllo-
taxy, photosynthesis and B group vitamins
in some C3 and Q plants 495

Cardarnom

Regeneration of plantlets from callus of
Elettaria cardamomum Maton 37

Callus culture

Regeneration of plantlets from callus of
Elettaria cardamomum Maton 37

C3 and C4 plants

Association of chlorophyll content,' phyllo-
taxy, photosynthesis and B group vitamins
in some C3 and C4 plants 495

Canopy index

Quantitative profile structure of certain forest
in the Kumaun Himalaya 529

Carbondioxide

Effect of CO2 in overcoming self-incompati-
bility barriers in Brassica campestris L. var.
toiia 227

Carry-over

Groundnut rust— its survival and carry-over
in India 93

Casuarina

Airborne pollen grains of Visakhapatnam :
A combined field and air sampling study 329

Cell elongation

Correlated promotion of ray-floret growth
in chrysanthemum by potassium chloride,
gibbcrellic acid and sucrose 101

Cell wall

Morphological and metabolic changes in
the egg and zygote of Lager stroemia specie sa.
I. Cell size, vacuole and insoluble poly-
saccharides 465

Cenchrus glaucus

Apomixis in Cenchrus glaucus Mudaliar et
Sundaraj 25

Chaetomorpha sp.

Heterotrophic bacteria associated with sea-
weed 487

Chandrasekharania keralensis Poaceae
Chandrasekharania : A new genus of poaceae
from Kerala, India 79

Chlorophyll

Association of chlorophyll content, p hylic -
taxy, photosynthesis and B group vitamins
in some C3 and C4 plants 495

Chromctophores

Contributions to our knowledge of Indian
algaa-III. Euglenmeae-va.rt I- The . Euglena
Bhrenberg 551

Chromosome relationships
Chromosome relationships of spinous sola-
nums 83

Chrysanthemum

Correlated promotion of ray-floret growth
in chrysanthemum by potassium chloride,
gibberellic acid and sucrose 101

Cluseaceae

Pharmacognastic studies on the flower of
Mesua fenea L. 211

Cacoyams

Non-inheritance of isomerism in cocoyams

17

Compositae

Embryological studies in Launaea nudicaulis
Hook 519

Contortion

Non-inheritance of isomerism in cocoyams

17

Cork- warts
Cork-warts in Eucalyptus species 289

Cyamopsis tetragonolobus
Changes in proteins, amino. and keto-acids
in different seedling parts of Cyamopsis
tetragonolobus Linn, during growth in light
and darkness 417

Cymbopogon

Embryological studies in three species of
Cymbopogon Spreng (Poaceae) 55

Cytology

Cyto logical studies on certain acanthaceae
from Central India 479

Cyto types

Cytological studies on certain acanthaceae
from Central India 479

Delonix regia

Impact of extension growth and flowering
on the cambial activity of Delonix regia
Rafin. 201

Desmids

Cell division in Staurastrum gracile Ralfs
under the scanning electron microscope

4

Distribution

Studies an JBeggiatoa : Distribution and growth
in aquatic habitats of Visakhapatnam 159

d-scars
Cork-warts in Eucalyptus species 289

d-stomata
Cork-warts in Eucalyptus species 28

Embryology

A contribution to the embryology of
Alysicarpus monilifer D.C. 9

Embryological studies in Launaea nudicaulis
Hook. 51

Index

/tes

iar arbuscular mycorrhiza in sub-
il aquatic and mat shy plant commu-

69

cut culture medium

5 on Bzggiatoa: Distribution and growth
latic habitats of Visakhapatnam 159
wp/ia sp.

•trophic bacteria associated with sea

487

of ridge gourd pollen on zoospore
Lation of Pseudoperonospom cubensis
;s significance in epidemiology 427
al characters
)mic importance of epidermal charac-

the Indian Thespesia Corr (Malvaceae)

509
us

farts in Eucalyptus species 2,89

.is 01 i gin

an, development and structure of
>dules in some members of the tribe
sae (Papilionaccae) 309

i growth

medic control of extension growth,
ormincy and flowering of Nariwn

Mill, and T/ievetia peruvianu Schum.

175

of extension growth and flowering

cambial activity of Delonix regia

201

relationships of spinous
ns 83

ilocarpa

r of Ficus macrocarpa Wt. ex Kin^
implocarpa nom. nov.) and F. guttata
Cing — A reinvest! gation with anato-
ividencc 115

tata

r of Ficus macrocarpa Wt. ex King
implocarpa nom. nov.) and F. guttata
Cing— A TV investigation with anato-
ividence 115

'is

in Cyperaceae : XVII Novelties in
tylis L. Vahl and their vegetative
f 43

utions to our knowledge of Indian
I. Euglenineae-fa.it I. The genus
Ehrenberg 551

itomy

wral anatomy of Kniphofia uvarfa
(Lifiaeea'e : Kniphofie'ae) 351

Flower

Pharmacognostic studies on the flower of
Mesua ferrea L. 21 [

Flower growth

Correlated promotion of ray-floret growth
in chrysanthemum by potassium chloride,
gibberellic acid and sucrose 101

Flowering

Photoperiodic control of extension growth,
bud dormancy and flowering of Neriutn
indicum Mill, and Thevetia peruviana Schum.

175

Responses of cotton-cultivars to long day
conditions 389

Flowering cambial activity
Impact of extension growth and flowciing an
the cambial activity of Delonix regia Rafin. 201

Flower longevity

Effect of water stress on opening and longe-
vity of flowers in Gladiolus 371

Flower opening

Effect of water stress on opening and longe-
vity of flowers in Gladiolus 371

Genus Jackiella
The- genus Jackiella in South India 131

Gcocalycaceac

Geocalyx Nees— a rare marsupial genus from
India 139

Geocalyx

Geocalyx Nees— a rare marsupial genus from
India 139

Gibbercllic acid

Correlated promotion of ray-floret growth
in chrysanthemum by potassium chloride,
gibberellic acid and sucrose 101

Gladiolus

Effect of water stress on opening and longe-
vity of flowers in Gladiolus 371

Gossypium spp.

Anther and pollen development in cotton
haploids and their parents 409

Grain amaranth

Leaf proteinase and nitrate reductase activi-
ties in relation to grain protein levels and
grain yield in four species of grain amaranth

4'33

Grain protein

Leaf proteinase and n'trate reductase acti-
vities in relation to grain protein levels and
grain yield in four species of grain amaranth

433

Grain yield

Leaf proteinase and nitrate reductase activi-
ties in relation to grain protein levels and
grain yield in four spfccies of grain amaranth

433

IV

Index

Green ear disease

Viability and infectivity of zoospores of
Sclerospora graminicola (Sacc.) Schroet in
the soil 303

Green gram

Interaction of kinetin with B group vitamins
on the seedling growth of green gram
(Phaseolus radiatus L.) 183

Groundnut rust

Groundnut rust— its survival and carry-over
in India 93

Growth

Studies on Beggiatoa : Distribution, and
growth in aquatic habitats of Visakhapatnam

159

Growth characteristics

Seed germination and seedling establishment
of two closely related Schima species 397

Growth rates

Growth response of some thermophilous fungi
at different incubation temperature 155

Growth retardants

Effect of Morphactin, AMO-1618 and DPX-
1840 on the endogenous levels of hormones
and its implication on apical dominance in
Glycine max Linn. 501

Growth strategies

Reproductive efficiency of secondary succes-
sional herbaceous populations subsequent
to slash and burn of sub-tropical humid
forests in north-eastern India 61

Gullet

Contributions to our knowledge of Indian
algae-III. Etiglemneae-'Part-L The genus
Euglena Ehrenberg 551

Hematochrome

Contributions to our knowledge of Indian
algae-III. Englenineae-Part-I. The genus
Euglena Ehrenberg 551

Heplaids

Anther and pollen development in cotton
haploids and their parents 409

Hepaticae

The genus Jackiella in South India 131
Geocalyx Nees— a rare marsupial genus from
India 139

Heterostyly

Turner a ulmi folia var. elega?is x T. ulmifolia
van angustifolia crosses and its bearing on the
taxonomy of the species 319

Heterotrophic bacteria

Heterotrophic bacteria associated with sea-.
weed 487

Himalayan forests

Quantitative profile structure of certain
forests in the Ktimaun Himalaya 529

Hormones

Effect of Morphactin, AMO-1618 and
Dpx-1840 on the endogenous levels of hor-
mones and its implication on apical domi-
nance in Glycine max Linn. 501

Hypnea sp.

Heterotrophic bacteria associated with sea-
weed. 4&7

Incompatibility

Effect of CO2 in overcoming self-incompati-
bility barriers in Brassica campastris L. var.
toria 227

Turner a ulmifolia var. elegam x T. ulmifolia
var. angustifolia crosses and its bearing on
tht taxonomy of the species 319

Intervarietal crosses

Turnera ulmifolia var. elegam x T. ulmifolia
var. angustifolia crosses and its bearing on
the taxonomy of the species 319

Insoluble polysaccharides
Morphological and metabolic changes in
the egg and zygote of Lagerstroemia speciosa.
I. Cell size, vacuole and insoluble poly-
saccharides 465

Isomerism

Non-inheritance of isomerism in cocoyams 17

Jungermanniales

Geocalyx Nees-a rare marsupial genus
from India 139

Keto acids

Changes in proteins, amino and keto-acids
in different seedling parts of Cyamopsis
tetragonolobus Linn, during growth in light
and darkness 417

Kinetin

Interaction of kinetin with B group vitamins
on the seedling growth of green gram (Phaseo-
lus radiatus L.) 183

Kniphofia itvaria

The floral anatomy of Kmphofia uvaria Hook.
(Liliaceae ; Kniphofieae) 351

Lagerstroemia speciosa

Morphological and metabolic changes in
the egg and zygote of Lagerstroemia speciosa.
I. Cell size, vacuole and insoluble polysac-
charides 465

Lamiaceae

Leaf surface studies of some medicinal sal-
vias . 449

Launaea nudicaulis

Embryological studies in Launaea nudicaulis
Hook 519

Index

f architecture

>af architecture of apocynaceae 189

f area ratio

sproductive efficiency of secondary succes-
>nal herbaceous populations subsequent
slash and burn of sub-tropical humid
rests in north-eastern India 61

: nitrate reductase activity
>af proteinase and nitrate red'uctase acti-
ties in relation to grain protein levels
id grain yield in four species of grain
naranth 433

f proteinase activity

saf proteinase and nitrate reductase acui-
ties in relation to grain protein levels and
•ain yield in four species of grain amaranth

433
ves

aaf surface studies of some medicinal sal-
as 449

uminosae

.natomy of the seedling of the Leguminosae-I

281

er dynamics

tructtire and function of a sub-tropical
umid forests of Meghalaya II. Litter
ynamics and nutrient cycling 255

ig day

.esponses of co.ttan-cultiva,rs to long day
auditions 389

crophomlna phaseolina

'ransmission of seed-borne inoculum of
fiacrophomina phaseolina from seed to plant

357

le sterile

utfher and pollen development in cotton
aploids and their parents 409

Ivaceae

'axonomic importance of epidermal charac-
srs in the Indian Thespesia Corr. (Mal-
aceae) 509

sua ferrea

'harmacognostic studies on the flower of
desua ferrea L. 211

arothermophiles

Jrowth response of same thermophilaus
ingi at different incubation temperatures 153
nosaceae

'ericarpial sclereids in some Mimosaoeae 297
at

Contributions to our knowledge of Indian
Igae-IH- Euglenineae-V&rt I. The genus
faglena Ehrenborg 551

orella speciosa

liaxteriellopsis lignicola and its Moorella
namorph 1

Morphology

Identity of Ficus macrocarpa Wt. ex King
(F. amplicarpa nom. nov.) and F. guttata
(Wt.) King— A reinvestigation with anatomical
evidence 115

Nerium tndicum

Photoperiodic control of extension growth, bud
dormancy and flowering of Nerium indicum
Mill, and Thevetia peruviana Schum. 175

New genus

Chandrasekharania : A new genus of Poaceae
from Kerala, India 79

New species

Chandrasekharania : A new genus of Poaceae
from Kerala, India 79

Nodal anatomy

Anatomy of the seedling of the Leguminosae-
I 281

Nuclear disintegration

Nuclear behaviour during heartwood for-
mation in Acacia auriculif or mis A. Cann 107

Nuclec'Ius

Nuclear behaviour during heartwood for-
mation in Acacia auriculif or mis A. Cann 107

Nucleus

Nuclear behaviour during heartwood for-
mation in Acacia auriculiformis A. Cann 107

Nutrient cycling-
Structure and function of a sub-tropical
humid forest of Meghalaya. II. Litter dynamics
and nutrient cycling 255

Nutrients

Structure and function of a sub-tropical
humid forest of Meghalaya I. Vegetation,
biomass and its nutrients 241

Obligate

Apomixis in Cenchrus glaucus MudrJiar ct
Sundaraj 25

Ontogenetic types

Ontogeny of the paracytic storna : Varia-
tions and modifications 145

Papilionaceae
Petal venation in Trigonella (Papilionaceae) 379

Paracytic

Ontogeny of the paracytic stoma : Varia-
tions and modifications 145

Paramylum

Contributions to our knowledge of Indian
algae-III. Euglenineae Part-I. The genus
Euglena Enrenborg 551

Pearl millet

Viability and infectivity of zoospores of
Sderospora graminlcola (Sacc.) Schroet in
the soil ' 303

VI

Index

Pericarp
Pericarpial sclereids in sorns Mimosaceae 297

Petal venation

Petal venation in Trigonella (Papilionaceae)

379

Pharmacognosy

Pharmaco gnostic studies on the flower of
Mesua ferrea L. 211

Pharmacognosy of the stems of Portulaca
quadrifida L. and Portulaca oleracea L. 235

Photoperiod

Photoperiodic control of extension growth,
bud dormancy and flowering of Nerium
indicum Mill, and Thevetia peruviana Schum.

175

Responses of cotton-cultivars to long day
conditions 389

Photosynthesis

Association of chlorophyll content, phyllo-
taxy, photosynthesis and B gioup vitamins
in some C3 and C4 plants 495

Phyllotaxy

Association of chlorophyll content, phyllo-
taxy, photosynthesis and B group vitamins
in some C3 and C4 plants 495

Poaceae

Embryological studies in three species of
Cymbopogon Spreng (Poaceae) 55

Airborne pollen grains of Visakhapatnam :
A combined field and air sampling study 329

P. oleracea L.

Pharmacognosy of the stems of Portulaca
quadrifida L. and Portulaca oleracea L. 235

Pollen effect

Effect of ridge gourd pollen on zoospore
germination of Pseudoperonospora cubensis
and its significance in epidemiology 427

Pollen productivity

Airborne pollen grains ot Visakhapatnam :
A combined field and air sampling study

329

Pollination calendar

Airborne pollen grains of Visakhapatnam :
A combined field and air sampling study 329

Polyethylene glycol

Effect of water stress on opening and longe-
vity of flowers in Gladiolus 37j[

Polyploidy

Cytological studies on certain acanthaceae
from Central India 4?9

Porto Novo

Heterotropic bacteria associated with sea-
weed 487
Portulaca quadrifida L.

Pharmacognosy of the stems of Portulaca
quadrifida L. and Portulaca oleracea L. 235

Potassium chloride

Correlated promotion of ray-floret growth in
chrysanthemum by potassium chloride,
gibberellic acid and sucrose 101

Prefoliation
Non-inheritance of isomerism in coccyams 17

Profile structure

Quantitative profile structure of certain
forests in the Kumaun Himalaya 529

Proliferation of cortex

Initiation, development and structuie of
root nodules in some membcis of the tribe
Trifolieae (Papilionaceae) 309

Protease activity

Changes in proteins, ami no and kcto-acids
in different seedling parts of Cyamopsis teira-
gonolobus Linn, during growth in light and
darkness 417

Protein

Changes in proteins, ami no and keto-acids
in different seedling parts of Cyamopsis tetra-
gonolobus Linn, during growth in light and
darkness 417

Pseudoperonospora cubensis
Effect of ridge gourd pollen on zoospore
germination of Pseudoperonospora cubensis
and its significance in epidemiology 427

Puccinia arachidis Speg.
Groundnut rust— its survival and carry-over
in India 93

Regeneration

Regeneration of plantlets from callus of
Elettaria cardamommn Maton 37

Reproductive effort

Reproductive efficiency of secondary succcs-
sional herbaceous populations subsequent to
slash and burn of sub-tropical humid forests
in north-eastern India 61

Ridge gourd

Effect of ridge gourd pollen on zoospore
germination of Pseudoperonospora cubensis
and its significance in epidemiology 427

Root nodule

Initiation, development and structure of root
nodules in some members of the tribe Tri-
folieae (Papilionaceae) 309

Root protein

Interaction of kinetin with B group vitamins
on the seedling growth of green gram
(Phaseolus radiatus L.) 153

Salvia

Leaf surface studies of some medicinal salvias

449

Index

va

Scanning electron microscope
Leaf surface studies of some medicinal salvias

464

Sclereids
Pericarpial sclereids in some Mimosaeeac 297

Sclerospora graminicola
Viability and infectivity of zoospores of
Sclerospora graminicola (Sacc.) Schroet in
the soil 303

Seaweed

Heterotrophic bacteria associated with sea-
weed 487

Seed-borne transmission
Transmission of seed-borne inoculum of
Macrophomina phaseolina from seed to plant

357

Seed germination

Seed germination and seedling establishment
of two closely related Schima species 397

Seedling establishment

Seed germination and seedling establishment
of two closely related Schima species 397

Seedling parts

Changes in proteins, amino and keto-acids
in different seedling parts of Cyamopsis tetra-
gonolobus Linn, duiing growth in light and
darkness 41 7

Seedlings

Anatomy of the seedling of the Legumino-
sae-I 281

S. surattense

Chromosome relationships of spinous sola-
nums 83

Seed set

Effect of CO2 in overcoming self-incompati-
bility barriers in Bmssica campestris L. var.
toria 227

Seed-sterility

Embryological studies in three species of
Cymbopogon Spreng (Poaceae) 55

Sesamum indicium

Transmission of seed-borne inoculum of
Macrophomina phaseolina from seed to plant

357

S. integrifolium

Chromosome relationships of spinous sola-
nums 83

Slenderness ratio

Nuclear behaviour during heartwood formation
in Acacia auriculiformis A, Cann 107

Soil

Viability and infectivity of zoospores of
Sclerospora graminicola (Sacc.) Schroet in the
soil 303

Solanum indicum

Chromosome relationships of spinous
solanums 83

Soybean

Effect of mcrphactin, AMO-1618 and DPx-
1840 on the endogenous levels of hormones
and its implication on apical dominance in
Glycine max Linn. 501

Square

Responses of cotton-cultivars to long day
conditions 389

Staurastrum gracile Ralfs
Cell division in Staurastrum gracile Ralfs.
under the scanning electron microscope 443

Sterility

Chromosome relationships of spinous sola-
nums 83

Stoma

Ontogeny of the paracytic stoma : Varia-
tions and modifications 145

Subtropical aquatic community
Vesicular arbuscular mycorrhiza in sub-
tropical aquatic and marshy plant communi-
ties 69

Subtropical forest

Structure and function of a sub- tropical
humid forest of Meghalaya I. Vegetation.
biomass and its nutrients 241

Successional communities
Reproductive efficiency of secondary &UCGOS-
sio-nal herbaceous populations subsequent to
slash and burn of sub-tropical humid forests
in north-eastern India 61

Sucrose

Coi related promotion of ray-floret growth in
chrysanthemum by potassium chloride, gibbc-
rellic acid and. sucrose 101

Effect of water stress on opening and longevity
of flowers in Gladiolus 371

Survival

Groundnut rust — -its survival and carry-over
in India 93

Tannia
Non-inheritance of isomerism in cocoyams 17

Taro
Non-inheritance of isomerismin cocoyams 17

Taxo-nornic details
The genus Jackiella in South India 131

Taxonomy

Identity of Ficus macrocarpa Wt. ex King
(= F. amplocarpa nom. nov.) and F. guttata
(Wt.) King— A reinvestigation with anato-
mical evidence 11-

Taxonomic importance of epidermal character
in the Indian Thespesia Corr. (Malvaceae)

509

via

Index

Teleomorph

Thaxteriellopsis lignicola and its Moorella
anamorph *

Temperature

Growth response cf some thermophilous
fungi at different incubation temperatures 1 53
Thaxteriellopsis lignicola
Thaxteriellopsis lignicola and its Moorella
anamorph. *

Thermo philes

Growth response of some thermophilous
fungi at different incubation temperatures 153
Thermotolerant

Growth response of some thermophilous fungi
at different incubation temperatures 153
Thespesia

Taxonomic importance of epidermal characters
in the Indian Thespesia Corr. (Malvaceae) 509
Thevetia pemviana

Photoperiodic control of extension growth,
bud dormancy and flowering ofNerium indicum
Mill, and Thevetia pemviana Schum. 175
Transition wood

Nuclear behaviour during heartwood forma-
tion in Acacia auriculiformis A. Cann 107
Trigonella

Petal venation in Trigonella (Papilionaceae) 379
Tree adaptation

Seed germination and seedling establishment
of two closely related Schima species 397
Tree growth

Impact of extension growth and flowering
on the cambial activity of Delonix regia
Rafin 201

Trichome scars

Cork-warts in Eucalyptus species 289
Trifolieae

Initiation, development and structure of root-
nodules in some members of the tribe Trifolieae
(Papilionaceae) 309

Turnsra

Turnera ulmifolia var. X T. ulmifolia var.
angustifolia crosses and its bearing on the
taxonomy of the species 319

Variations

Ontogeny of the parley tic stonia : Variations
and modifications 145

Variation in vacuolar size
Morphological and metabolic changes in the
egg and zygote of Lager sir oemia specie sa. I.
Cell size, vacuole and insoluble pojysf.ccha-
rides 465

Vascular derivatives

Impact of extension growth and flowering on
the cambial activity of Delonix regia Rafin 201

Vegetative anatomy

Studies in Cyperaceae : XVII. Novelties in
Fimbristylis (L.) Vahl and their vegetative
anatomy 43

Vellar estuary

Heterotrophic bacteria associated with
seaweed 487

Venation pattern
Leaf architecture cf apocynaceae 189

Vesicular arbuscular mycorrhiza
Vesicular arbuscular mycorrhiza in subtropical
aquatic and marshy plant communities 69

Viability of seeds

Seed germination and seedling establishment
of two closely related Schima species 397

Water stiess

Effect of water stress on opening and longevity
of flowers in Gladiolus 371

Wound coik
Cork-warts in Eucalyptus species 289

Zoospore

Viability and infectivity of zoospores of
Sclersoopora graminicola (Sacc.) Schroet in the
soil 303

Zoospore germination

Effect of ridge gourd pollen on zoospore
germination of Pseudoperonospora cubensts
and its significance in epidemiology 427

Yield

Responses of cotton-cult ivais to long day
conditions 389