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Arthur Pierson Kelley was born in tlie village of Malvern,
in the hills suburban to Philadelphia, on August 15, 1897. He
studied at the University of Pennsylvania, graduating with the
degree of B.S. in Biol, in 1920. At the same University he ac-
quired the degrees of M.A. (1921) and Ph.D. (1923). Later he
studied plant physiology under Dr. Livingston at The Johns
Hopkins University. Commencing as a teacher, he was suc-
cessively instructor and assistant professor of botany at Rutgers
University. Thereafter he joined the United States Forest
Service hoping to find more time for research on mycorrhizae.
In an effort to devote himself more fully to this work he later
developed his private biological station and herbarium at Landen-
berg, Pennsylvania. From here he produced his digest of mycor-
rhizal literature which has become widely known. Dr. Kelley is
now chiefly engaged in farm life and the reconstruction of his-
torical farm houses, pursuits which, he writes, often leave too
little time for the study of mycotrophy. Dr. Kelley's early
publications dealt with soil acidity in relation to plant distri-
bution, later papers with other ecological problems and the
various aspects of mycotrophy.

A NEW SERIES OF PLANT SCIENCE BOOKS •

edited by Frans Verdoorn
Volume XXII

MYCOTROPHY

in

PLANTS

Frontispiece. — Development of an Odontoglossum as figured by Noel Bernard in 1909.
Figure 1 is a sectional view of seed; the remaining figures are habit sketches ot emoryos and
seedlings, the last two indicating regions ot tungal infection by shading. Figure 3 is of special
Interest because it shows neatly coils of the fungus (considered Rhizoctonia lanuginosa), which
has entered by an "absorbing hair". It shows also amoeboid condition of nucleus of infected
cells. Figure 2 shows by vivid contrast the condition of an uninfected embryo at 4 months of
age, whereas the embryo of figure 3 is but one month old.

MYCOTROPHY

in

PLANTS

Lectures on the Biology

of Mycorrhizae
and related structures

hy

Arthur P. Kelley, ph.d,

Landenberg, Pennsylvania

1950

WALTHAM, MASS., U.S.A.

Published by the Chronica Botanica Company

First published MCML

By the Chronica Botanica Company

of Wahham, Mass., U. S. A.

Copyright, 1950, by the Chronica Botanica Co.

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PREFACE

Since this little volume was started more work has been done on
mycotrophy. Yet the status of the subject remains unchanged. It
is still considered by most botanists— who are the only people to whom
even the word is familiar — as a specialty which should be listed
under the general heading of pathology. Mostly it is not listed at all.
Mycorrhizae are still considered abnormal structures occurring chiefly
on pine roots; while by some curious aberration the mycotrophic
structures found in prothallia and in rhizomes are likewise termed
mycorrhizae. Fungus-roots are found in rootless plants! There is,
moreover, a common impulse to lump all symbiotic phenomena
together, as odds and ends and thrown together in a heap, so that
the mycotrophist (if such a term may be used) is expected to be
interested in cases of algae growing in higher plants, or of small
insects found living in plant tissues. It is hoped that this book, in
spite of its obvious faults, may serve to show how mycotrophy is
separated from other phenomena, and how widespread is the
mycotrophic habit.

Europe continues to be the center of mycotrophic study. In spite
of war, a considerable number of papers has appeared in this part
of the world in the last few years. Only a few papers on the subject
have been published on the other continents.

Tree mycorrhizae continue to attract attention. Bjorkman
(1944) has made important studies in Sweden. In Poland, Dominik
(1946) has published interesting papers on fruit-tree mycorrhizae
which were produced on all trees studied. Endotrophic mycorrhizae
which were stimulated by applications of farmyard manure to the
soil were found by Sabet (1946) on Citrus in Egypt, while Smith
(1944/45) in Queensland found that "decline" in mandarin was
associated with depletion of the soil and consequent interference with
mycorrhizal activity. These results lend support to the idea that
internal processes in at least certain trees can be controlled by appro-
priate manipulation of the habitat.

Kelley — viii — Mycotrophy

Forest tree mycorrhizae receive continued attention from two of
the principal investigators of mycotrophy. Rayner (1941) has
summarized her own and related researches on the effect of composts
on tree growth. Melin's (1946) attention has been turned especially
to growth and anti-bacterial substances with respect to mycotrophy.
Recently fallen leaf-litter was found to contain water-soluble sub-
stances that promoted the growth of litter-decomposing and mycorrhi-
zal fungi. Cognate studies have been made by Harley but his latest
paper was not available at this writing.

Synthesis of Boletus with pine seedlings to form synthetic
mycorrhizae was performed by Ferreira (1941), thus confirming
the discoveries of a number of earlier investigators. What is said
to be the first synthesis of alder nodules was accomplished by Plotho
(1944).

Obligate symbiosis is still questioned, but it is claimed by Bose
(1947) for Casnarina under the name of hereditary symbiosis.
Mycelium extended into every organ of the tree, spreading from the
seed-coats.

Among herbaceous plants, orchid mycotrophy claims the attention
of Burgeff (1943), and Sprau (1939). The Burmannias and their
mycotrophy were investigated by Ciferri ; and at his laboratory at
Pavia there is active interest in our subject. To the several papers
on mycotrophy in halophytes was added one by Fries (1944) : nine
out of ten halophytes examined were mycotrophic. Again, in game-
tophytes of the fern Botrichium, Naryana (1939) described endo-
phytic infection. So, too, in hepatics described by Peyronel (1942)
from Italy: double infestations occur and the symbiosis is much
affected by edaphic conditions, but in general the relation seems to
be of mutual benefit. A noteworthy contribution to the study of
herbaceous mycotrophs was made by Barrett (1947), who suc-
ceedeed in isolating Rhizophagus in pure culture.

There is a continually increasing emphasis on the physiology of
mycotrophy, and several contributions to this subject have been
made recently. From a yellow Corticiiim a pigment called "corticro-
cin" was isolated (Erdtman, 1947), "the first n-polyneoid diacid
found in Nature." Another paper reverted to the Stahlian concept
of stomata in relation to mycotrophy. It will be recalled that Stahl
believed mycotrophs had a reduced number of stomata and a limited
transpiration stream, mycotrophy supplying the nutrients otherwise
supplied by photosynthesis. A recent investigation (Linsbauer &
Ziegenspeck, n.d.) concludes that amongst mycotrophs there is a
significant reduction in number and formation of stomata. Extreme
mycotrophs resemble holoparasites. A third physiological paper

Mycotrophy — ix— Preface

(Prat, 1945) treats of gradients in mycotrophic plant tissues.
Resistance to parasites is an important function in plants. In myco-
trophy the gradient of resistance to the mycelium is slowly progres-
sive. Mycelium penetrating toward the apex is always checked, for
the axial gradient of resistance is directed toward the point of the
root and is progressive ; but the radial gradient of resistance is
abruptly corrected at the level of the endodermis. Chemical barriers
are more efficacious in developing resistance than physiological
barriers. The gradients vary according to season. Still other papers
(Magrou, 1944, 1946) deal with tuberisation and the factors which
control it.

Acclimatisation has been proved aided by symbiosis. At Angers
(Blaringhem, 1937), building a well-aerated humus layer favourable
to growth of symbiotic fungi aided the acclimating of 400 spp. of
conifers and 150 forms of oak, which proceeded to make 3 to 5
times the growth of the finest specimens of these species in their
native haunts.

Two general papers on mycotrophic symbiosis have appeared re-
cently. In one (Owen, 1947), symbiosis is examined with reference
to the true character of the symbionts. It is concluded that true
mutualism exists between nodule bacteria and legumes, and perhaps
with mycorrhizae. But orchids are considered dependent on fungi,
since the fungi can live apart saprophytically. With characteristic
thoroughness Burgeff (1943) has analyzed mycotrophic phenomena
and presented a new classification of them. Realizing that their most
important function is material exchange which is dependent on the
union of tissues of both components, he classifies "mycorrhizae" as :
(1) Tolypophagous, in which rhizoctonial fungi form pelotons that
are digested with release of fat, glycogen and nitrogenous material
into the orchid plant that harbours them. (2) Thamniscophagous, in
which arbuscles are digested, leaving sporangioles as excreta. Very
widely distributed in green plants, including ferns and liverworts.
In certain ferns and colorless saprophytes the arbuscles go over to
the preceding form and are hence termed thamniscotolypophagous{ !)
The fungi appear to be Endogonaceae, hence the mycorrhizae are
called phycomycetoid. (3) Ptyophagous. Found in such plants as
Gastrodia and Monotropa, these "mycorrhizae" show resorption of
materials released into cell by fungal hyphae. Free fungal bodies
called "phytosomes" are formed which, being resorbed, leave
"Exkretkorper". (4) Chylophagoiis. Sap resorption occurs in sub-
terranean colorless and saprophytic prothallia of Lycopodium. There
is no digestion and there are no excretion bodies. Sap is exuded by
guttation of hyphae into intercellular spaces. (5) Halmophagous.

Kelley — x — Mycotrophy

Included are the ectotrophic mycorrhizae which have mantle and
Hartig net. There is said to be resorption of nutrient salts. These
mycorrhizae are obligatory for many forest trees. With some species,
ectendotrophic forms are found.

Selected chapters of the manuscript were read by Dr. G. R. Bisby,
of the Imperial Mycological Institute; by Dr. D. T. MacDougal;
by Dr. M. C. Rayner, of Bedford College ; and by Dr. H. E. Young,
Dept. of Agriculture and Stock, Queensland. None of these is
responsible for views expressed by the author. Above all, the author
is indebted to Dr. Frans Verdoorn, whose unflagging patience has
brought the book through difficult times.

The Author

Dr. J. A. HijNER of the Netherlands, who worked at the California
Institute of Technology, during 1948, just sent us a preliminary report
in which he claims, on the basis of experimental data, that vitamins
are essential for growth of Rhizoctonia isolated from Cymbidium;
specifically, folic acid or para-aminobenzoic acid with thiamin. He
considers that in nature these growth substances are suppHed by ger-
minating orchid seeds, which are known to contain them. He also
points out localization of fungi in tissues of the host which he believes
due to antibiotics produced by the host.

Attention might still be drawn to the small symposium edited by L.
Blaringhem at the occasion of the Exposition Internationale, Paris
(1937) : — Symbiose et Parasitisme, I'oeuvre de Noel Bernard. 89 pp.
Paris : Masson. Other very recent publications which have not been
referred to in the book: — Bahme, R. B. (1949) : Nicotinic acid as a
growth factor for certain orchid embryos. Science 109 :522-3. —
Baumgartel, T. (1940) : Mikrobielle Symbiosen im Pflanzen- und
Tierreich. 132 pp. Braunschweig (Lithoprint ed. 1946. Ann Arbor,
Mich.: Edwards Bros.). — Kramer, P. J. & K. M. Wilbur (1949) :
Absorption of radioactive phosphorus by mycorrhizal roots of pine.
Science 110:8-9. — Magrou, J. (1943) : Des orchidees a la pomme de
terre. 203 pp. Paris. — Peyronel, B. (1939-40) : Luce, humus e mi-
corrizia. Atti d. R. Ace. Sc. di Torino 75:13 pp. — Schaede, R.
(1948) : Die pflanzlichen Symbiosen. ed. 2. 187 pp. Jena.

CONTENTS

Preface vii

Contents xi

List of Text Illustrations and Plates xv

Lecture I

The Rise of Mycotrophic Study:- The beginning of root study

— Early studies on root hairs — Early study of nodules —
Nutrition of Monotropa — Nutrition of orchids — -My-

COTROPHY IN FERNS AND FERN ALLIES MyCOTHALLISM IN

LIVERWORTS EARLIEST OBSERVATIONS OF MYCORRHIZAE DIS-
COVERY OF THE HaRTIG NET — NaTURE OF MYCOTROPHISM

MyCORRHIZAE DEFINED CONFLICTING CLAIMS OF DISCOVERY

Frank's antagonists — Bernard and orchid symbiosis —
Tuber formation and perennism — Symbiosis in Lolium

— Obligate mycotrophy in heaths — Identity of the my-
corrhizal fungi — Isolation of mycorrhizal fungi — The

NITROGEN theory OF MYCOTROPHISM ThE StAHLIAN HY-
POTHESIS— Carbon hypotheses — Growth promoting sub-
stances — Phagocytosis — Phycomycete mycorrhizae —
Forest tree mycorrhizae — Ecology of mycorrhizae • —
Trends 1

Lecture II

The Occurrence of Mycorrhizae:- Reasons for studying my-
corrhizal occurrence — ^The occurrence of root-hairs —
Generality of mycorrhizal occurrence — Symbiosis among

ALGAE MyCOTHALLI AMONG LIVERWORTS FUNGAL SYMBIOSIS

with Pteridophytes — Symbiotic fungi among Arthro-
PHYTES — Mycorrhizae and mycothalli of Lycopsida —
Gymnospermous mycorrhizae — Mycorrhizae of Cycads —
Mycorrhizae of Ginkgo — Mycorrhizae amongst the Tax-
aceae — Mycorrhizae in Pinaceae — Mycorrhizae in
Gnetaceae — The method of opportunism — Mycorrhizae
in Apetalae 14

Kelley — xii — Mycotrophy

Lecture III
The Fungal Endophytes:- Nature of the mycorrhizal fungi

PhYCOMYCETE mycorrhizal fungi ASCOMYCETOUS MY-
CORRHIZAL FUNGI HeMIBASIDIOM YCETES HyMENOM YCETOUS

MYCORRHIZAL FUNGI GaSTEROMYCETOUS MYCORRHIZAL FUNGI

PhALLOMYCETOUS MYCORRHIZAL FUNGI FoRM GENERA OF

MYCORRHIZAL FUNGI — FuSARIUM RhIZOCTONIA — PhOMA

— Mycelium radicis — Conclusion Z7

Lecture IV

Fossil Mycorrhizae:- Limitations of the fossil record —
Sources of material — Fossil Phycomycetes — Fossil
hepatics — Fossil ferns and their endophytes — Mycor-
rhiza of fossil Lycopod — Mycorrhizae in a seed fern —
Mycorrhizae in Cordaites — Summary 47

Lecture V
Distribution of Mycotrophic Plants:- General — Germany —

France and the Iberian Peninsula — British Isles — Low-
lands AND Scandinavia — Baltic and Russian States —
The Arctic — The Alps — Central Europe — The Balkans

— Italy — Africa — Asia — Java — Japan — New Zealand

— Australia — South America — West Indies and Central
America — North America — North-eastern U.S.A. —
Southern U.S.A. — Central U.S.A. — Rocky Mountains —
Pacific Coast 53

Lecture VI

Mycotrophic Plants and Their Environment:- Soil as a myco-
trophic HABITAT — Mycorrhizae and soils — Humus — •
Mycorrhizae as soil indicators — Mycorrhizae in the soil
profile — Soil texture — Soil moisture — Soil solution • —
Soil reaction — The use of free nitrogen — Soil tempera-
ture — Altitude — Light — Phenology — Mycorrhizae in
relation to habitat — Salt marsh — Prairie — Soil In-
oculation — Compost studies 68

Lecture VII
Mycothalli and Mycorrhizomes:- General character — Mv-
cothalli in liverworts — Infection of mycothalli — ■

Mycotrophy — xiii — Contents

Limitation of endophyte — Digestion of endophyte —
Mycothalli in fern gametophytes — Mycothalli in lyco-
POD gametophytes — Mycorrhizomes in ferns — Mycor-

RHIZOMES in orchids MyCOCARYOPSES AND INFECTION OF

aerial organs 91

Lecture VIII

Mycodomatia:- Significance of the term — Psilotum —

Cycads — PoDOCARPus — Casuarina — Myrica — Alnus —

Polygonum — Raphanus — Tribulus — Legumes — Ailan-

THUS — Ceanothus — Elaeagnus — Hippophae — Coriaria

— Eucalyptus — Daucus — Ericads — Solanum — Melam-

PYRUM OrOBANCHE — CoMPOSITES — JUNCUS MOLINIA

— Cyperus — Asparagus — Allium — Orchids . . 103

Lecture IX
Structure of Mycorrhizae:- The kinds of mycorrhizae — My-
corrhizal compared with non-mycorrhizal roots — Exter-
nal form — Pseudomycorrhizae — The colours of mycor-
rhizae— The exterior surface — The mycorrhizal apex —
Renewed growth — Ectotrophic vs. endotrophic — Ecto-
trophic mycorrhizae — Endotrophic mycorrhizae — Pel-
OTON mycorrhizae — Vesicular-arbuscular mycorrhizae

— Arbuscles — Ericaceous mycorrhizae — Coralloid my-
corrhizae — Tuberous mycorrhizae — Outer cortex and

PASSAGE cells WaLL TUBULES HaRTIG NET ThE STELE

116

Lecture X
Obligate Symbiosis:- Fungi and trees — Ecological influences

— Special cases — Fungi and herbs — Obligatism and
NUTRITION — The ericads — The problem of Calluna —
Other Ericaceae — The orchids — Germination of orchid
SEED — Necessity of orchid fungus — Degeneration of
orchid fungus — Hydrolysis of starch — Sugars and asym-
biotic germination — Fungus supplies sugar — Impotence
of unaided small seeds — Carbon supply to embryo — Oblig-
atism IN LOWER PLANTS SUMMARY 136

Lecture XI
Theories of Mycotrophy:- Contrasted concepts — Parasitism

— Curbed pathogens — Schools of mycotrophism — Mycor-

Kelley — xiv — Mycotrophy

RHIZA REPLACES ROOT-HAIRS — MyCOPHAGY — RoMELLIAN HY-
POTHESIS — Nitrogen theory — Chemical studies of the

NITROGEN theory IMPORTANCE OF P StAHLIAN HYPOTHESIS

— Hatchian hypothesis — Transpiration and mycotrophy

— Objections to the Stahlian hypothesis — Bjorkman's
hypothesis — Carbonaceous theories — Hydrocarbon hypo-
thesis — Carbohydrate hypothesis — Growth-promoting
substances — Limitation of mycotrophic hypotheses —
The intaking mechanism — Ectotrophic intake — Root-
hairs versus setae — Endotrophic intake — Hyphae as
nutrient conveyors — Teleology in mycotrophism . 148

Lecture XII
Mycotrophic Phagocytosis :• Significance of the term — Older
descriptions of phagocytosis — Phagocytosis in peloton
mycorrhizae — Phagocytosis in arbuscular- vesicular
mycorrhizae — Phagocytosis in ericaceous mycorrhizae —
Phagocytosis in ectotrophic mycorrhizae — Limitation of
endophyte — Limitation in orchids and other herbs ■ — ■
Limitation in hepatics — Limitation in root apices —
Limitation in long roots — Limitation in green tissues —
Summary of limitation — The starch relation — Con-
clusion 164

Bibliography 183

Subject Index 216

Index of Plant Names 219

LIST of TEXT ILL USTRA TIONS and PLA TES

Frontispiece. — The development of Odontoglossum iv

Fig. 1. — Longitudinal section through mycothallus of P^//ia ^/>i/'/i;y//a .. 18

Fig. 2. — Mycorrhizae in Piniis virginiana 27

Fig. 3. — Mycorrhizae in Sugar Maple, Acer saccharum 32

Fig. 4. — Renewed growth of mycorrhiza-bearing mother-root of Pinus

Strobus 82

Fig. 5. — Section through an older mycothallus of Botrychium obliqimm 94

Fig. 6. — Portion of mycothallus of Lycopodium obscumm shown in sec-
tion 96

Fig. 7. — Longitudinal section through apex of a mycorrhiza of Pinus

rigida 120

Fig. 8. — Renewed growth of a mycorrhiza of Pinus virginiana 122

Fig. 9. — Cross-section of an ectotrophic mycorrhiza of Qucrcus montana 124

Fig. 10. — Cross-section of endotrophic mycorrhiza of Acer Negundo. . . . 126

Fig. 11. — Section of a mycorrhiza of Abies balsamea 133

Fig. 12. — Portion of a cross-section of the ectendotrophic mycorrhiza of

Cornus florida 166

Fig. 13. — Portion of a section through mycorrhiza of Abies balsamea. . . . 168

Fig. 14. — A cell from mycorrhiza of Allium sphacrocephalus 170

Fig. is. — Portion of a longitudinal section through a mycorrhiza of

Pteridium aquilinum 171

Fig. 16. — Some cells from mycorrhizal cortex of Fraxinus americana. . . 173

Pl. 1. — One of the earliest illustrations of mycotrophic infection, by

Edouasd Prillieux, 1856 209

Pl. 2. — Mycorrhizae in Scot's Pine, Pinus sylvestris 210

Pl. 3. — Mycorrhizae of Car pinus betulus and Fagus sylvatica 211

Pl. 4. — Effect of mycorrhizae on plant growth 213

Pl. 5. — Photomicrograph of a portion of a fossil mycorrhizome of Sclero-

pteris illinoiensis 215

"Das Wort Symhiose hedeutet sundchst gans allgemein das re-
gelm'dssige Zusammenvorkovimen von Lehenszvesen unter denselben

dussern Faktoren Die beiden Organismen bilden nach der

Vereinigung eincn neuen Organisnnts, der ah einheitlich su betrachten
ist und unter neuen Bedingungen den Kampf urns Dasein aufnimmt."
(H. BURGEFF, 1909).

"... der Pils, als der alleinige Zufuhrer alles fiir den Baum
erforderlichen Wassers und Ndhrniaterials aiis dem Boden erscheint."
(A. B. Frank, 1885).

"... the similar {tropical) conditions that prevailed during the
Carboniferous mitigate our surprise at finding symbiosis occurring so
far back as this." (E. W. Berry, 1904).

"Mit Anwesenheit oder Abwesenheit von Bautnhumus die My-
corhiza entstcht oder verschwindet." (A. B. Frank, 1888).

"... the host plant may, as it were, gain the upper hand and cause
the fungus to enter into a mutually beneficial partnership." (Cavers,
1903).

"Quoique I'existcnce pour ainsi dire normale d'un parasite dans les
tissus d'une plante soit un fait tres singulier, on pent observer asses
frequemment dans la famillc des Orchidees." (E. Prillieux, 1856).

"On any hypothesis, the evolution of an obligate relation with a
parasitic or facultatively parasitic fungus is difficult to explain."
(M. C. Rayner, 1927).

" . . . ce me semble, inferer que la maficre brune sert a I'alimenta-
tion de la plante. . . " (E. Prillieux, 1856).

"La symbiose est a la frontiere de la maladie." (N. Bernard, 1909).

Lecture I
THE RISE OF MYCOTROPHIC STUDY

The Beginning of Root Study:— After Herbalists had done
their work and, by means of wood-cut and description, had made
known the flora of Europe, inquiry began to be made into physiology
of plants. It is said that Major, in 1665, directed attention to
circulation of sap. Four years later, Ray proved ascent and descent
of sap and its lateral movement in certain plants. At the same time,
Woodward demonstrated by experiment that roots take in, not merely
water, but also materials dissolved in the water. Both Ray and
Woodward published their papers in the Philosophical Transactions
of the Royal Society. Beginning in this manner, with study of roots
in water culture, plant physiology was turned to root-hair study,
from which it has not seriously deviated to this day.

Early Studies on Root Hairs: — Early observers were perhaps
influenced by knowledge of circulation of blood in animal bodies and
were doubtless expecting to find vessels in plants. When Malpighi
found root-hairs on elm, black poplar, and willow roots, he assumed
that these structures took up crude sap and passed it on to vessels.
Grew, publishing about the same time (1682) had decided that
spongy ends of roots served admirably for absorption of water and
food from the soil; and Hales in 1727 and de la Baisse in 1733
tried to show experimentally that the greater quantity of water
used by the plant was taken up through ends of the root-tips and
that root-hairs were only incidental phenomena. To these hypotheses
was added in 1768 that of S. Simon, who stated that roots, at least
the noduliferous, are merely excretionary organs which serve to
eliminate excess elaborated sap from the plant. In the earliest years
of the nineteenth century, Garradori showed that root-hairs are
wanting in water, from which fact he concluded that root hairs serve
for absorption of moisture from the air and not for absorption of
liquid water, which, he concluded, must be taken up by the spongy
ends of roots. But, according to Moldenhawer, root-hairs may be
compared to druse-hairs of leaves : they secrete a liquid which serves

Kelley — 2 — Mycotrophy

to dissolve food materials somewhat as saliva does in animals. It was
F. Meyen, in 1838, who came to the modern view that root-hairs
serve merely to increase the outer surface area of the root.

By such studies attention was focused on root-hairs until Botany
was firmly moulded to the view that higher plants are nourished by
a root-hair mechanism. So positive had Botany become that by 1883,
Frank Schwarz (from whom we have quoted much of the pre-
ceding paragraph) was able to state without exciting contradiction:
"From my researches it may be stated that root-hairs are present on
most plants, and when a plant fails to produce root-hairs it may be
counted an exception." He listed as exceptions : water and swamp
plants, and those the water and salt requirements of which are met in
a special way, as in conifers, noduHferous plants and in part by
parasites.

Early Study of Nodules: — Thus it was, not by extended ob-
servation or study of plants in nature but by sheer dogmatism that
root-hairs came to be regarded as the predominate root structures of
higher plants. Hairless roots were considered exceptional, but they
were constantly being noted. Malpighi, early microscopist that he
was, had described and figured nodules while du Hamel du Monceau
in 1758 had stated that such structures were generally found on
leguminous plants. Even Meyen, in 1829, who has been accredited
with discovery of mycorrhizae, simply described nodules of the
alder. Alder nodules were more carefully studied by Woronin in
1867 but his inadequate facilities led him to confuse bacterial strands
with fungal hyphae. Even to this day there is confused thought
about root-nodules for some investigators assert them to be purely
bacterial while others consider them fungal.

Nutrition of Monotropa: — Besides nodulous roots there were
obviously other exceptional kinds. The waxy Monotropas that appear
in deciduous woods have no apparent root-hairs and were long con-
sidered parasites. It was thought that they must be attached to
tree roots for they always grew under the trees and in a thick mat of
humus and intertangled rootlets, although as early as 1832 Fries
had noticed a fungus connected to Monotropa. Several investiga-
tors reported on it in that short-lived journal, The Phytologist, and
came to the conclusion that, whatever else this plant might be, it was
certainly not a parasite. One of the observers, Luxford, in 1844,

Lecture 1 — 3 — Rise of Mycotrophy

hazarded the suggestion that Monotropa secures nutriment from
the surrounding humus. Drude, in 1873, conformed to tradition
by stating that Monotropa starts life as a parasite but later, he asserted,
the plant becomes a saprophyte on soil humus, a fungus being present
in its tissues. The true nature of Monotropa was first made clear
by Kamienski in 1881, who carefully described the structure of its
mycorrhiza and indicated that the plant is supplied with nutriment
by a fungus which derives its materials from the soil humus. His
papers (for there were two) have long since become forgotten history
but they were considered important in their day.

Nutrition of Orchids: — Orchids, like Monotropas, also proved
exceptional in their root structures. Indeed, many orchids have no
roots ! In place of roots they have branched stems that form coral-
structures that anchor the plant in the rich humus soil in which it
grows. It was in 1842 that Schleiden described what were later
recognized as fungal hyphae in Neottia (for Neottia has been as
necessary to orchid students as Drosophila to geneticists), but
Schleiden confessed he did not know what the "tubes" were which
he had observed in the rhizomes. Unless, he said, they were like the
ones which Gottsche had found in liverworts. Five years later
Reissek identified true fungal hyphae in rhizomes of many orchids
but he oddly concluded that these hyphae developed from starch
grains. But Schacht in 1854 showed that the starch in reality was
utilized by the fungus, which forms a weft of hyphae about the
starch grains. Just how the starch was digested (by the process we now
call phagocytosis) was described by Prillieux in an excellent paper
that appeared in 1856. He found in orchid cortical cells (needless to
say, of Neottia), a yellowish-brown matter, and he noted that these
cells retained their nuceli which were of great size and provided with
two nucleoli. The matter seemed to be nitrogenous and was woven
about with septate hyphae but as phagocytosis proceeded the matter
dwindled ; and Prillieux concluded that this matter served as nutri-
ment for the orchid. He observed, too, that cells filled with granular
matter at flowering time gradually lost the matter as anthesis ad-
vanced. The granules were apparently absorbed and they probably
nourished the orchid. But Prillieux's work was little regarded and
later papers by other authors were farther from the truth. Thus
Reinke in 1873 suggested that this yellowish matter which he called
slime, acted as a pumping organ, swelling up as water was taken in

Kelley — 4 — Mycotrophy

and forcing the water on through the tissues. Mollberg in 1884
questioned whether the fungal endophyte brought any nutriment to
the orchid; while Eidam in 1879 attempted to culture the fungus by
allowing fungi to develop on orchid "roots" placed in damp air.

Mycotrophy in Ferns and Fern Allies: — ^^A yellow matter similar
to that found in orchids was found in lycopods by van Tieghem in
1871. He described this substance, and it was further described by
Bruchmann in 1874, who found it free from starch. Bruchmann
noticed, too, that in older tissues the fats and nitrogenous substances
dwindled in amount while a quantity of chrome-yellow granules ap-
peared. Ten years later (cf. Treub, 1884) it had become established
that endophytes are generally distributed in lycopods and that their
presence is not harmful. Indeed, Treub regarded them as commensals.
In ferns, especially the Ophioglossaceae and Marattiaceae, fungal
infection had been reported, and also presence of yellow matter.
Since the infection was present in fern stems, these structures were
appropriately termed fungal-rhizomes or mycorrhizomes. The term
mycorrhizome was coined by Dangeard in 1891, in his study of
Tmesipteris.

Mycothallism in Liverworts: — In various liverworts a foreign
substance was observed in the form of large brown cask-shaped
structures (Milde, 1851), the significance of which could not be
discerned. But Gottsche in 1843, as earlier stated, had observed
a system of branching tubes in Aneura; and these tubes were definitely
described as hyphae by Leitgeb in 1874. Since the thallus of the
liverwort harbours a fungal endophyte, it is known as mycothallus.

Earliest Observations of Mycorrhizae: — The association of
fungi with roots of higher plants has long been known. In Theo-
iHRASTUs' Enquiry Into Plants (according to Hort's translation)
we read: "For as for the fungi which grow from the (oak) roots
or beside them, these occur also in other trees." Theophrastus (or
Tyrtamus, to use his proper name) may have been walking in a
woodland and observed sporophores of fungi which he seems to have
traced to tree roots. It is a long step from the third century B.C. to
A.D. 1829, but the next reference to mycorrhizae was made in the
latter year by Meyen. In a short paper he called attention to peculiar
structures which he found on beech roots, which he believed were
beginnings of parasitic plants. Perhaps they were mycorrhizae,
perhaps not. But attention was being directed to root structures and
little by little knowledge increased. In 1837 Link stated that most

Lecture I — 5 — Rise of Mycotrophy

roots are formed in humus; Tulasne in 1841, that tree roots were
found frequently surrounded by mycelium of the truffles fungus;
while Gasparini in 1856 stated that a fungal mantle was found about
roots of chestnut, hazel and pine. Hairs were considered so inevitably
present on roots that it was heresy to speak of anything else, hence
ScHACHT in 1860 cautiously stated that, while root-hairs are present
on such trees as oak and beech, they were less abundant on pine and
fir. ScHWARZ, from whom we have already quoted, presented a
list of conifers from which they were lacking.

Discovery of the Hartig Net: — When coniferous rootlets
were examined in section it was seen that the wall possessed what
was termed a peculiar cell-wall thickening; and it is an odd fact
that morphologists beheld fungal mycelium in such rootlets for a
long time without being aware of its nature. Thus, Nicolai in 1865
gave a tolerable description of what is now known as Hartig net
without realizing that he was describing a foreign organism in the
conifer. Van Tieghem in 1871 also described these "thickening
bands" ; and he noticed furthermore that the penultimate layer of
root cortical cells was filled with a solid deposit, a fact of significance
in mycotrophic nutrition of these conifers. It remained for Reinke
in 1873 to call attention to the similarity of these supposed thickenings
to mycelial strands which Gottsche had found in the liverwort,
PelUa. With realization that tree rootlets were characteristically
invaded by hyphae, specific infections were described. Thus, Boudier
in 1876, described Elaphomyces on birch, oak and chestnut roots; and
he noticed furthermore that such roots were found in acid but not
in alkaline soils.

Nature of Mycotrophism : — The nature of the fungus-host
relation was next considered. That the fungus in the root was a
harmless parasite was the opinion of Resa, expressed in 1878; and
GiBELLi in 1873 had the same opinion. But Reess, publishing in
1880, questioned whether the fungus was a parasite on the tree-root
or a saprophyte on soil humus. The supposed parasitism of some
plants which live in rich humus, such as the Burmannias, had been
questioned by Cruger in 1848; and clear recognition of saprophytes
as distinct from parasites had been made by Solms-Laubach in
1868. By a shrewd guess, Pfeffer in 1877 came to the conclusion
that saprophytes actually make use of the materials of humus. With-
out experimental evidence, he inferred that mycorrhizal fungi ob-
tained nutrient material from the humus and transferred it to the host
plant which, of itself, was incapable of utilizing the otherwise un-

Kelley — 6 — Mycotrophy

available materials of the humus. At the same time Pfeffer realized
that the fungus was essentially a parasite which was no more than
kept in check by the host plant. Since the picture of mycotrophy pre-
sented by Pfeffer is so close to the actual phenomenon, he may
perhaps be considered the true discoverer of the mycorrhiza.

Mycorrhizae Defined: — But, important as Pfeffer's paper
now appears, it attracted little attention and it was not until 1885 that
world-wide attention was suddenly drawn to fungus-roots. Just why
Albert Bernhard Frank's dissertation "Ueber die auf Wurzel-
symbiose beruhende Ernahrung gewisser Baume durch unterirdische
Pilze" should have had such a profound effect is for others to de-
termine : Suffice to say, modern mycorrhizal study dates from this
paper. In it, Frank had invented and defined the term in these words :
"Der ganze Korper ist also weder Baumwurzel noch Pilz allein,
sondern ahnlich der Thallus der Flechten, eine Vereinigung zweier
verschiedener Wesen zu einem einheitlichen morphologischen Organ,
welches vielleicht passend als Pilzwurzel, Mycorhiza, bezeichnet
werden kann." As the word comes from the Greek and good usage
requires doubling the letter r in compounding, we now write it,
mycorrhiza.

Conflicting Claims of Discovery: — Immediately after publica-
tion of Frank's epochal work there was a rather entertaining flurry
of papers. Some people wished to call attention to their own work,
published earlier than Frank's, while others desired the world to
know they had often seen exactly what Frank had described.
Several pressed the claims of Kamienski as the discoverer of the
mycorrhiza, but as we read his 1884 paper we find only this vague
statement: "I suppose, moreover, without being able to confirm it,
that the fungus which grows on Monotropa is the same which lives
parasitically on roots of conifers and other trees. This fungus de-
forms the root and occasions their dichotomy. I have found indeed,
among the roots of Monotropa, a great quantity of other roots which
were very fine, deformed, and belonging to trees which grew all there
about : they were so interlaced that the mycelium which webbed them
on touching might be said to be interblended."

Frank's Antagonists: — Then, too, there was a persistent effort
to label mycorrhizal fungi as mere harmless parasites like the leaf-
spot fungi. Robert Hartig was a particularly active opponent of
Frank ; and Hartig's views were held in later years by the American,
McDougall. Frank spent little time in advancing mycorrhizal

Lecture I — 7 — Rise of Mycotrophy

study for he turned to other studies, but he returned to the defence
of his hypothesis of mycotrophy from time to time. At first he
thought that mycorrhizal fungi are concerned especially with nitrogen
nutrition of the higher plant, bringing nitrogen salts into the
mycorrhiza. But later he taught that the higher plant is actually a
parasite on the fungus, drawing it into the root, tending and finally
devouring it.

Bernard and Orchid Symbiosis: — At the turn of the century
a new phase of mycotrophic study was developed. Hitherto the
interest had been chiefly with forest plants, following the initiative
of Frank. But when Noel Bernard began to publish on myco-
trophism of orchids an important new branch of science was opened
up. Bernard isolated from orchid tubers fungi which he classified
into three groups and placed in the genus Rhizoctonia. These fungi,
he found, were able to cause orchid seed germination and, lacking
presence of a fungus, there was no germination. It is interesting
to note that Salisbury, when he described orchid seed germination
in 1804, failed to note presence of fungus. But Bernard did not
assert that a fungal symbiosis was an inevitable necessity, for he
induced asymbiotic germination of orchid seed with salep, a poly-
saccharide derived from dried orchid tubers. Sterile cultures were
grown by Knudson, who showed that in greenhouse propagation
of orchids sugars may be used in germination of the seeds in place
of fungi ; but in nature it is of course the fungus that is responsible
for the germination.

Even before Bernard's untimely death m 1911, Burgeff had
been publishing on orchid mycotrophism. He called the endophytes
Orcheomyces, but later made use of the less convenient designation
of Mycelium radicis.

Tuber Formation and Perennism: — Bernard was convinced
that tuber formation in orchids and other plants was due to their
symbiotic life with fungi, and that after many generations of such
symbiotic life the "habit" of forming tubers was acquired so that
tubers were still formed by the plant even in the absence of fungal
aid. Thus the potato, native of Andean highlands, formed tubers in
the Andes in symbiosis with a species of Phoma, but in northern
latitudes, as in France, tubers were still formed without presence
of a fungus. He advanced the idea that influence of a cold climate
parallels the action of the fungus, and pointed out that in a hot
climate such as that of Algiers, tuber formation "degenerated" and
tubers were no longer formed in absence of a fungus. Using this

Kelley — 8 — Mycotrophy

principle, Bernard's successor, Magrou, and his compatriot, Cos-
TANTiN, have developed an hypothesis of perennism by which they
attempt to account for existence of perennial plants on a symbiotic
basis.

Symbiosis in Lolium: — Another phase of mycotrophic study
dealt with fungal infection of seeds, particularly of the seed-fruits
or caryopses of Lolium, a genus of grasses. Discovered by Vogl
in 1897, the constant presence of hyphae in the hyaline layer of the
caryopsis and typical stages of digestion, are curious phenomena that
have nevertheless been established by repeated researches, especially
at the hands of McLennan in Australia. The endophyte also occurs,
according to Neill (1940) in leaves of Lolium but not in those of
other pasture grasses.

Obligate Mycotrophy in Heaths: — Infection of seeds of
heaths, especially of Calhina, has likewise been reported, particularly
by Rayner; indeed, practically all tissues of the plant were said to
be invaded by the otherwise mycotrophic fungus. Rayner claimed
that mycelium present in the seed-coat of Calluna grows into the
developing plantlet so that a sterile culture of this heath can not be
obtained. This assertion has been challenged by Knudson and by
Freisleben. According to Freisleben (1935), the fungus causes
an amelioration of soil conditions in which the heath grows and does
not directly affect life of the heath.

Identity of the Mycorrhizal Fungi: — The specific identity of
mycotrophic fungi has proved more puzzling than might at first be
suspected. It would seem that sporophores directly attached to tree
roots might be safely considered the mycorrhizal fungi of these hosts.
Constant association of truffles with tree roots furnished the incen-
tive for Frank's study of the mycorrhiza, and there are many other
such associations, as, for example, Heheloma with birch. In many
cases rhizomorphs have been traced from sporophore to mycorrhiza
and the associated fungus has been termed mycorrhizal, but such as-
sociations may actually be the result of a secondary infection. Hence
the numerous citations of mycorrhizal fungi based on connection
of sporophore with mycorrhiza are not necessarily valid.

Isolation of Mycorrhizal Fungi: — Isolating the fungus directly
from the mycotrophic organ has proved, in most cases, impracticable ;
and the usual method of identifying the fungus in question is to
grow a suspected fungus in pure culture, inoculate it into a sterile

Lecture I — 9 — Rise of Mycotrophy

seedling and if a mycorrhiza results, to consider the fungus mycor-
rhizal. The pioneer in synthesizing mycorrhizae was Josef Fuchs,
and one of the most prominent investigators of such syntheses is
Melin. Still more recently, Modess has reported many such
syntheses ; and Fries has formed synthetic mycorrhizae with mono-
spore mycelia.

The Nitrogen Theory of Mycotrophism: — As to the nature
of the mycorrhizal symbiosis, numerous hypotheses have been ad-
vanced. It has already been noted that R. Hartig and others con-
sidered the symbiosis to be a case of harmless parasitism, and that
Kamienski maintained that there is a beneficial symbiosis only in
the case of Monotropa. Frank had concluded that the parasitism
was just the reverse, — that the higher plant was a parasite on the
fungus ! He had found that "the tissues of a mycorrhizal tree are
nitrate free". As it was known that the fungus could readily make
use of ammonia and organic nitrogen compounds, he considered it
self-evident that in this way such compounds were taken up from
nitrate-free or nitrate-poor soils and liberated in the mycorrhiza.
The nitrogen nutrition hypothesis which Frank originated was
elaborated by subsequent investigators, von Tubeuf, Moller, Mul-
LER, Weiss, to mention a few ; and especially by Melin, who has
conducted many researches into the nature of mycorrhizae of forest
trees. Melin appears to consider that, in Swedish forests, the prob-
lem of the mycorrhiza is above all a nitrogen problem.

The Stahlian Hypothesis: — On the other hand, Stahl in 1900
emphasized the mycorrhizal intake of all minerals used by the higher
plant. Trees being brought into competition with fungi and bacteria
for nutrient materials contained in the soil are seriously limited in
their food supplies ; and in the mycorrhizal fungi find provisioners
that bring water and dissolved salts into the roots. It is not merely
nitrogen that is brought into the root, according to this hypo-
thesis, but all the minerals the soil solution contains. As a corollary
the supposed relation of transpiration to mycotrophy was pointed
out. Plants possessing a large transpiration stream bring a
considerable quantity of mineral salts into the higher plant and
deposit them in its tissues. Plants with a smaller transpiration stream
have a smaller salt intake and are presumably compensated with
the salts provided by the mycorrhizae. But the Stahlian hypothesis
ran into difificulties which are detailed later; and it met with little
favour until supposedly revived by Hatch in 1937. In reality Hatch
originated a new hypothesis which states among other things that

Kelley — 10 — Mycotrophy

mycorrhizae greatly increase the absorbing surface of the rootlet.
The Hatchian hypothesis quietly drops the Stahlian conception of
transpiration streams. Still more recently, in 1943, Routien and
Dawson stated that "mycorrhizae increase the salt-absorbing capa-
city of the roots primarily by adding to the supply of exchangeable
hydrogen-ion derived in part at least from carbonic acid."

Carbon Hypotheses: — Besides the nitrogen- and mineral-nutri-
tion hypotheses, there are carbon hypotheses which have been advanced
by two investigators. McLennan in 1926 stated that the more
generally accepted ideas connecting mycotrophism with nitrogen
nutrition were insufficiently founded ; and she concluded on the basis
of her researches on Lolium that "a. metabolic exchange takes place
from the fungus to the higher plant, with the result that
the later obtains a supply of fat or oil." McLennan believed that the
researches of Knudson and of Bernard lent support to this con-
ception. Another carbon hypothesis was advanced by Young in
1940, in these words : "It is thought that as well as providing a more
efficient absorptive system on the tree roots so that mineral salts and
nitrogen compounds are more readily available, the mycorrhizas also
furnish a means of augmenting the carbohydrate supply ... In the
author's conception the fungus manufactures its own carbohydrate
supply from the available soil organic matter, and a portion of this
is transferred to the higher plant by means of the intimate associ-
ation existing in the mycorrhizal structures."

Growth Promoting Substances: — Complex substances of the
humus suggested still further hypotheses. Link, Wilcox and Link
in 1937 had suggested that "heteroauxin applied to a plant can
either substitute in part for its autoauxones or augment their ac-
tion, and the well-known fact that soil fungi and bacteria produce
heterauxin suggests that some of the beneficial effects of humus soil
may be due to the auxones of decaying plant debris or soil flora."
Still more specifically, Magrou in 1939 reported a more luxuriant
development in Arum as a result of supplying it with aneurin
(Vitamin B^). Melin in 1939 and 1940 found increased growth
in seven mycorrhizal fungi when these organisms were given aneurin
or yeast filtrate. In mycorrhizae of Monterey pine, MacDougal and
DuFRENOY reported in 1943 that the "independent growth of isolated
segments of mycorrhizal roots makes it obvious that through these
hyphal branches the root receives from the soil not only the C, N,
O, P necessary to build up the nucleus, the cytoplasm and its inclusions
(mitochondria and plastids), cell-walls, but also the mineral com-

Lecture I — 11 — Rise of Mycotrophy

pounds ordinarily taken in by root-hairs, and growth promoting sub-
stances, thiamin, nicotinic acid . . ."

Phagocytosis : — Whatever may be finally decided as to the nature
of the mycorrhizal symbiosis, it is observed that something is released
in the tissues of the higher symbiont by the fungus. It is becoming
increasingly evident that in what are termed "digestion cells" of the
host the hyphae break down and disgorge their contents, the matter
being digested and assimilated by the host. Since it is a cellular diges-
tion, Bernard called it a phagocytosis; but the whole process had
been pictured and to some extent described long before Bernard.
In 1943 it was suggested by Kelley that the whole mycotrophic rela-
tion depends on a balancing of ionic concentrations between fungus
and higher plant.

Phycomycete Mycorrhizae: — While the majority of mycor-
rhizal fungi are basidiomycetes, it is now recognized that in many
mycotrophic symbioses the fungus is a phycomycete. Such symbioses
were studied in earlier years especially by Peyronel. Rayner, in

1935, commented on the "remarkably wide geographical distribution
of this 'Phycomycete type' of mycorrhizal association, its prevalence
in plant species of the most diverse affinities, together with its re-
corded appearance in certain crop plants. . ." Other more recent
studies of the phycomycete mycorrhizae include those of Biraghi in

1936, on cereal grains. Bain in 1937 on cranberry, Sabet in 1939 on
cotton, and Ruggieri in 1937 on Amygdahts. Butler in 1939 pre-
sented a paper summarizing what was then known of this sort of
mycorrhiza, terming it the vesicular-arbuscular mycorrhiza and
grouping the endophytes under the generic name of Rhizophagus.

Forest Tree Mycorrhizae: — Although so much work has been
done on forest tree mycorrhizae, knowledge concerning them is still
defective. Considerable is known of those that occur on pine, spruce,
larch, beech and some others, but researches upon them have been
done by a few individuals working with limited material. The
extent of mycorrhizal occurrence, both taxonomic and geographical,
is still a matter of conjecture and, as most of mycorrhizal research
has been done in Europe, the forests of the other continents are for
the most part still uninvestigated by students of this science. Moreover,
the natural difficulties in the way of isolating the mycorrhizal fungi
make for ignorance of the symbiosis, for the syntheses of seedlings
and fungi in pure cultures show merely what man can achieve and
not what occurs naturally in the forest. Then, too, the nutrition of

Kelley — 12 — Mycotrophy

forest trees is largely conjectural. From information now extant
one cannot say with accuracy whether trees have root-hairs or mycor-
rhizae, much less the exact mode of nutrition of any particular kind
of tree. Even were it concluded that the tree was nourished with
the aid of mycorrhizae, the precise nature of mycotrophism is still
in doubt. Hence the foresters' dealings with trees are akin to the
mediaeval doctor's treatment of his patients.

Ecology of Mycorrhizae*: — The earlier mycotrophic problems
in ecology involved the relation of mycorrhizae to sandy soils and
to humus. More recently there has been discussion regarding the
presence or absence of mycorrhizae in prairie soils with reference
to establishment of trees on such areas. It has been claimed that the
endophytes are absent from prairie soils and that such soils must be
inoculated with suitable fungi before mycorrhizae will be formed.
But investigators have not been careful to distinguish between prairie
soils and the more arid steppe soils. Then, too, several investigators
report abundant endophytes in these soils, and recent observations
show that there is a rapid spread of trees into certain prairie areas.

As to soil reaction, there is more nearly an unanimity of opinion,
for it is evident that mycorrhizal fungi thrive best in acid media.
Hence, mycotrophic structures are more likely to be found in acid
soils while root-hairs may be expected in mull soils. But in regard
to soil solution, little can be said, for in spite of the number of in-
vestigations into nutritional and soil problems and the multiplicity of
papers on salts in soil solution, the actual connection of the plant
with the soil has been almost completely ignored. To say that a plant
has mycorrhizae and is nourished by mycotrophy has been regarded
as sufficient, and whether materials get from the soil into the plant
by mechanical means or by black magic is left to the imagination of
the reader. All the detailed studies of soil solution in the B horizon
have no necessary connection with a large proportion of plants in
native habitats. And thus the mechanism for the intake of materials
into mycorrhizae is a subject of research awaiting investigation. Yet
some attention has been paid to the microhabitat of the fungus-root
and its community of organisms, the rhizosphere as it has been appro-
priately called.

*A valuable paper on the ecology of ectotrophic mycorrhizae, by Dr. J. L.
Harley, of Oxford, has recently appeared in Biological Reviews.

Lecture I — 13 — Rise of Mycotrophy

Trends: — Early interest in mycotrophic study was taxonomic,
and a sufficient number of plants was examined to show that the
mycotrophic habit was widespread in the plant kingdom. Then there
were sporadic collections which showed that mycotrophy existed in
each of the continents. Today biology inclines toward philosophical
dissertations which place the writer in an impregnable position since
if anyone demurs it rests with him to disprove the postulates. Myco-
trophic study still requires much work in taxonomy and morphology.

Lecture II

THE OCCURRENCE OF MYCORRHIZAE

Reasons for Studying Mycorrhizal Occurrence: — The occur-
rence of mycorrhizae is a subject of the first importance in mycor-
rhizal study. It is important for two reasons : first, to determine the
extent of mycorrhizal occurrence, and, second, to determine mycor-
rhizal importance. It must be confessed that little is known of the
extent of mycorrhizal occurrence. As we examine the history of the
subject it is evident that human interest in mycorrhizae has followed
a usual pattern : first there has been a flurry of interest, then an
haphazard and eager collecting of various material from casual
sources, and finally a settling down to solving problems of isolated
detail that may or may not be important to the subject as a whole.

It would seem to be more logical to examine first of all the plant
kingdom to determine whether mycorrhizae actually occur widely in
that kingdom. We assume that they do but the assumption is not
based on research. Two or three dozen investigators have gone into
the woods and fields, they have sunk their digging tools into the earth,
and whatever came up was made the subject of study. There are a
few good papers on mycorrhizal occurrence, but very few. The classic
paper is Janse's account of the mycorrhizae occurring about Buiten-
zorg in Java, and after half a century we still consult the paper. Janse
consciously limited his research to chosen representatives of various
plant families and as a preliminary study it is excellent, but it should
be followed by similar papers on other members of the same, and of
other, families.

In consequence of the fortuitous method which has hitherto been
employed in mycorrhizal study, we have the following summary of
what is now on record : Considerable good work on some of the hepat-
ics ; nothing on mosses or arthrophytes ; a little on pteridophytes ;
good studies on the gametophyte generation of some lycopods ; a very
unequal emphasis on members of the Gymnospermae, with most mem-
bers unknown as to mycorrhizal condition (all attention must be de-
voted to a few pines and spruce!) ; and a scattering of information
about some angiosperms. On this slender evidence scientists assert
the importance of mycorrhizae. They may be important and probably
are important; but it is scarcely scientific to jump at conclusions. We

Lecture II — 15 — Occurrence of Mycorrhizae

would be in a much better position to go forward with research if our
research were founded on a considerable number of papers like that
of Janse's, or even such simple lists as that of Klecka and Vukolov
(1935).

But if we know little of mycorrhizal occurrence, we know even less
of root-hair occurrence. Morphologists have never paid much atten-
tion to roots except to study the vascular systems of older roots. Roots
are in the ground, it takes considerable work to get them out, and the
botanist pulls off a twig or a leaf or a flower and goes onward. An
herbarium in which a "specimen" always included the root would be a
curiosity. A thorough-going study of root structure has yet to be
made, especially of the "absorbing" system of smaller rootlets: we
still await a Systematic Morphology of Root-endings.

This statement leads to a consideration of the second reason for
studying occurrence of mycorrhizae, namely, their importance. It is
obvious that plants take in their nutrient materials through root-hairs
or mycorrhizae except for a comparatively small number that are able
to live without either. If root-hairs predominate in nature, then
physiological research should be directed chiefly to root-hair plants;
but if mycorrhizae predominate, then plant physiology should be con-
cerned chiefly with mycorrhizae. Botany will some day be forced to
a decision in the matter. At present botanists are in a position of
ignorance, for they do not know what sort of root endings exist on
the majority of plants in their natural haunts. They assume that root-
hairs are the usual organs for intake of nutrient materials into plants,
but their assumption cannot be substantiated from the records of
research. Moreover, there is little prospect that research will be done
on such structures, for the ruling motive in botanical science today
appears to be a subservience to the authority of tradition.

The Occurrence of Root-Hairs: — According to Frank
ScHWARZ (1883), the first mention of root-hairs is found in Mal-
piGHi's Opera Omnia (1681), having observed them in elm, black
poplar, and willow and he believed that in their tiny tubes he had be-
fore him that in which crude sap ascended and was later led into the
vessels. He found them especially in those places where earth was
not immediately in contact with roots. When the root hairs then
pushed out into neighbouring soil, they grew around individual soil
particles and surrounded them, so that they formed a span between
the roots and soil particles. A similar clinging of the hairs was de-
scribed by Malpighi in the roots of ivy. Almost simultaneously N.
Grew (1682), in his "Anatomy of plants", advanced the idea that the
spongy ends of roots served admirably for provision with food and

Kelley — 16 — Mycotrophy

water; while Hales (1727), no less than the Father of Plant Physi-
ology !, decided that root-hairs are only incidental phenomena in intake
of materials from the soil, the chief intake being through the root tips.
Among the seemingly countless authors of the first 20 years of the
19th century (yet producing nothing new), may be mentioned Garra-
DORi and MoLDENHAWER. The former noted that root-hairs are
wanting in water, from which he concluded that they serve, not for
absorption of liquid water, but moisture from the air, while liquid
water is taken up by spongy root-ends. According to Moldenhawer,
root-hairs may be compared tO' druse-hairs of leaves : they secrete a
liquid which serves to dissolve the food materials, being comparable
in a way to saliva of animals.

The first description that was given right direction was by F.
Meyen {in Neues System der Pflanzenphysiologie, 1838. Bd. 2, p.
6), who proceeded from a description of absorptive hairs of moss and
characeous plants, showing incidentally that in these cases the root-
hairs completely take the place of roots. He called attention further-
more to the supposed universal distribution of root-hairs in higher
plants, investigated their development, and what is more important,
attributed to them the direct intake of liquid water. He came to the
view that root-hairs serve merely to increase the outer surface of the
root, and he showed that the number of root-hairs formed is dependent
upon external conditions. Next we may cite the work of G. Gaspar-
RiNi, the "Richerche sulla natura dei succiatori e la escrizione delle
radici" (1856), which is a most comprehensive work on root-hairs
but it offers in general nothing new. Gasparrini had investigated
quite a large number of plants and found them with few exceptions to
have root-hairs ; he did not go into a study of the conditions of root-
hair formation but satisfied himself with describing their form, con-
tent, etc. The finest portions of earth, embedded in a gummy sort of
a mass which clung to the root-hairs he considered to be excretionary
products of the hairs. He even designated as such roots which had
an evident root-cap. Schacht incorporated Gasparrini's work in
his text of 1859. Much more precise than his predecessors, Sachs
(1860) made clear the significance and function of root-hairs; and
from him the more modern phases of such study may be dated.

Generality of Mycorrhlzal Occurrence: — In spite of our
comparative ignorance of root structure in particular, it is known that
in all major groups in the plant kingdom there are fungi living with
other plants in mutualistic relation. No major group from "Thallo-
phyte" to Spermatophyte is excepted. In the lower groups we speak
of mycothalli or lichen bodies while in higher groups are mycorrhizo-

Lecture 11 — 17 — Occurrence of Mycorrhizae

mata or mycorrhizae, but in all cases the relationship appears essen-
tially the same.

Symbiosis among Algae: — Mutualistic symbiosis of fungi with
algae, so far as known, is confined to lichens. While it is still main-
tained by some people that the lichen symbiosis is a parasitism of the
fungus upon the alga, the balance of favour is with the Schwenden-
erian theory of mutualism. The algae, principally Cyanophytes and
protococcid Chlorophytes, supply organic material, presumably sugary
carbohydrates, to the fungi which take in water and dissolved salts
from the exterior into the lichen body and thus to the enmeshed algae.

The lichen body is a thallus but it differs radically from the myco-
thallus of the liverwort, which is a tissue containing hyphal strands.
In the lichen thallus the chlorophyll-bearing thalloid cells (algae) are
discrete or loosely massed together (gonidia), not forming a tissue as
in the liverwort ; and the lichen thallus is for the most part a specially
and characteristically formed mycelium. Then, too, the fungal sym-
biont of the lichen thallus produces reproductive structures (spores
and soredia) while in a true mycothallus the fungus does not produce
reproductive bodies.

Mycothalli among Liverworts: — Widespread occurrence of
fungal symbiosis amongst liverworts has been demonstrated by the
twenty-six investigators who, in the course of history, have studied
mycothalli. It is to be assumed that anything so lacking in obvious
utilitarian interest as a liverwort should attract but little general regard.
It was apparently Schleiden who in 1839 first described what we
know as fungal infection of a liverwort (Pellia), — not Leitgeb (1879)
as has been erroneously stated ; but as Schleiden did not realize what
he had seen, Gottsche (1858) may be termed the real discoverer of
mycothallism. In old thalli of Pellia epiphylla and of Preissia com-
mufafa he found a branched system of threads going from cell to cell
which he at first considered as an individual vascular system but later
recognized as fungal. Earlier in the history of mycorrhizal study it
was supposed that fungi are commonly associated with the Junger-
manniaceae (Leafy liverworts) but absent from the Marchantiaceae
(Thalloid liverworts). Such at least was the opinion of Ncmec
(1899), who supposed that the Marchantiaceae, being starch pro-
ducers, could not have endophytes ; and Stahl (1900) seized upon
this erroneous suggestion and wove it into his ingenious hypothesis
of mycotrophism. But it was soon made clear that symbiotic fungi are
constantly found in many of the Thalloid liverworts.

Kelley

18 —

Mycotrophy

Four orders of Hepatics are recognized, of which two, the Ric-
ciales and the Anthocerotales have received virtually no attention from
our workers. It is not likely that the Ricciales, which are mostly
aquatic, should have endophytes ; and there is but one report for these
two orders, made by an early worker named Milde in 1851. He found
what he termed "Kugeln" or little barrel-shaped spore-like bodies in
the thallus of Anthoceros, Riccia, and other frondose hepatics, and he
found that these "Kugeln" were made of little "cells" united in strings
and that they never left the thallus voluntarily: "neither am I able",
he said, "to make any observation as to their significance".

Fig. 1. — Longitudinal section through mycothallus of PeIHa epiphylla. A
fungal hypha, having entered through a rhizoid, ramified through tissues of the
mycothallus and produced intracellular vesicles. (^Redrawn from Ridler, Ann.
Bot. 36:198, 1922).

Most of the work on mycothalli has been done on the Marchanti-
aceae and Jungermanniaceae, the Favourite Four species for study
being Conocephalus (Fegatella) coniciis, Marchantia polymorpha,
Lunularia criiciata, and Pellia epiphylla. A greenhouse in Holland, a
mountain area in India, a region in South Africa, the botanical garden
of Java, and a few other spots all in Europe except one in Morocco —
and none at all in the Americas — give us the rest of our information
regarding the mycothalli of liverworts. Apart from the four species
mentioned, only 37 other species have been reported on for their myco-
thallism and some of these reports are from the vague early days
while others are mere casual mentions.

As to the other Bryophytes, we know virtually nothing of their
possible mycothallism. The Sphagnums, being aquatic mosses for the

Lecture II — 19 — Occurrence of Mycorrhizae

most part, would not be suspected of harbouring endophytes although
it is true that a fungus has been reported from a Sphagnum capsule ;
the Andreales are not mentioned; and but one report comes to us of
the higher mosses. Servattz (1913) stated that white filaments of a
fungus (the size of Streptothrix) formed dense intertangled masses
which grew well on agaricized peptone bouillion, a fungus which
seemed to be an Oospora. This organism exerted a particularly ac-
tivating action on Phascum cuspidatum, and in culture the moss plants
and mycelium made normal growth when together whereas without
the mycelium the moss developed only a protonema. This favourable
action was negated later when the fungus covered over the gelose and
gained ascendancy over the moss.

Fungal Symbiosis with Pteridophytes : — Fungal symbiosis
seems to occur commonly with Pteridophytes and certain species,
especially of the OphiogJossaceae, have been studied intensively. On
the other hand, the families of Matoniaceae, Hymenophyllaceae and
Schisaeaceae have never been examined for symbiotic fungi, so far as
literature records ; while many species in the remaining families are
yet tO' be investigated. It is scarcely to be expected that the Parkeri-
aceae, Marsiliaceae and Salviniaceae should harbour mycorrhizal endo-
phytes since these plants are aquatics; and we find that Asm (1934)
states that Ceratopteris and Marsilea are not mycorrhizal, thus con-
firming Stahl, who had also found Pilidaria globulifera non-mycor-
rhizal.

Of all the Pteridophytes, the Ophioglossaceae have been most
studied for symbionts. Janse, who worked at Buitenzorg, had found
branched hyphae and sporangioles in the third layer of cortex only,
in Ophioglossum pendulum; while a few years later Campbell (1907),
working in the same place, found the same form of endophyte in both
gametophyte and sporophyte, infection of the sporophyte occurring
chiefly from the gametophyte. O. moluccanum and O. simplex have
also been studied carefully and found to be characteristically mycor-
rhizal. Helminthostachys, at first reported to be without endophytes,
was studied later (Lang, 1902) and found tO' be essentially similar
in its symbiotic relationships to Ophioglossum. Fourteen species of
Botrychium have been examined for endophytes and proved to be
mycorrhizal: of these, twelve were studied by Grevilltus (1895)
who stated that in these species hyphal formation always occurred in
the roots. Both generations of Botrychium are mycorrhizal, infection
taking place through the rhizoids.

All of the five genera of Marattiaceae have been studied and found
mycorrhizal. The tree-like Angiopteris of the Orient tropics is re-

Kelley — 20 — Mycotrophy

ported by several workers, the latest (Stark, 1925) finding the plant
with endophyte in the Leningrad Gardens. In Archangiopteris, as in
several others ferns, West (1917) found a new fungus that produced
under natural conditions distinct reproductive bodies other than
vesicles. Marattia itself, although reported non-mycorrhizal by Stahl,
is attested by several later workers. Campbell (1908) states posi-
tively the presence of endophyte in green prothallia of M. Douglassi
besides those of several other Marat tiaceae, including Kanlfussia aes-
culifolia which West (1917) confirms for the sporophyte plant. West
also describes and figures infection for Danaea alata and D. nodosa,
neatly demonstrating apparent phagocytosis within the outer layers of
cortex.

The leptosporangiate ferns have been less studied but are not with-
out their endophytes. Two genera of the Osmundaceae have been
studied, van Tieghem as early as 1870 reporting mycelial hyphae of
a parasite coiled about dark masses in large cells of the inner zone
of cortex of Osmunda regalis and several other ferns. Strangely
enough, Stahl asserts that this species is not mycorrhizal and no one
else has made a later statement. Campbell, in his studies of fern
prothallia, found that many cells in O. cinnamonea and 0. Claytoniana
contain an endophyte which consists of large non-septate hyphae that
are strictly intracellular. For the sporophyte of the last species, Loh-
MAN (1927) says that an endophyte is absent. In an excellent paper
on Todea harhara, Cribbs (1920) notes that an endophytic fungus
was found to occur frequently in the cortical tissues of the root exter-
nal to the endodermis and internal to the sclerenchymatous cells of the
peripheral region. It was found to gain entrance by root hairs and also
by dissolving its way through the epidermal cell-wall at the edge of
the root-cap. Cribbs gives us neat figures which beautifully delineate
apparent digestion stages.

For the Gleicheniaceae our only author is Campbell, who mentions
five species of Gleichenia that have mycothallic prothallia. Two au-
thorities sponsor the Cyathaeaceae, those Tree ferns of the tropics:
Janse found Cyathaea mycorrhizal in Java, pelotes and sporangioles
being found in 3-4 layers of cortex; while Asai (1934) reports C.
spimdosa as mycorrhizal, and also Alsophila formosana and A.
pusttdosa.

The Polypodiaceae offer a hopeful although little touched field, for
most of these ferns live in humus soil and might be expected to har-
bour endophytes ; yet we suspect that lack of economic utility of ferns
accounts for aversion to their study.

Amongst wildlings of the American prairies, Lohman (1927)
gathered some casual specimens of fern and reported briefly as to

Lecture II — 21 — Occurrence of Mycorrhizae

their being mycorrhizal or the reverse; and two species are thus re-
ported by him for the first and only time, namely, Cystopferis fragilis
(mycorrhizae occasional) and Onoclea sensihilis (ectotrophic) ; and
he records Adiantum pedatum and Pteridium aquilinum also mycor-
rhizal. DoAK (1927) finds the Adiantum species endotrophic while
Asm (1934) reports on A. flabeUuIatum. Pteridium is undoubtedly
mycorrhizal although the poll as it now stands is tied: Stahl (1900)
and Takamatsu (1930) insisting that P. aquilinum is not mycorrhizal
while LoHMAN (1927) and Rayner (1927) state that it is or appears
to be, and Rayner clinches the matter by presenting a photomicro-
graph of the apparent endophytic fungus within the root tissues.

The large genus Aspidium is reported non-mycorrhizal by the
four who have reported upon it, — Frank, Stahl, Hoeveler, and
Lohman; yet surely these humus-dwellers deserve a reconsideration.
So, too, with Aspleniiim which Stahl and Janse reported as without
endophytes, and Polypodiinn which Stahl negated although Faber
(1925) seems to indicate mycorrhizae for P. Feei. Then Nephrolepis
and Blechnmn are recorded very casually only by Asai (1934) and
deserve more study. Last of all in this brief list of studied Poly-
podiaceae is Cliciropleuria, which is of special interest because occur-
rence of endophytic fungi in prothallia of C. hicuspis var. integri-
folia is added evidence in the author's opinion (Nakai, 1933) that the
genus should be removed from the Polypodiaceae and be placed in a
separate family, the Cheiropleuriaceae. This is the only case known
to the writer where mycorrhizae are made of service as a taxonomic
criterion.

Symbiotic Fungi among Arthrophytes: — There is almost no
information extant regarding symbiotic fungi of the Equisetums.
Sadebeck in 1875 described browning of the prothallia of E. arvense
and E. palustre in culture, due tO' infection by a species of Pythium
which was named P. equiseti, but this was apparently a case of para-
sitic attack rather than of mycothallism. Janse said that forest dwell-
ing Eqiiisctae in Java appear never to have endophytic fungi in the
roots; Hoeveler (1892) found E. hie^nale and E. silvaticum not
mycorrhizal ; and Stahl found no trace of infection in Equisetum.
LoHMAN, in Iowa, lists E. arvense as containing an endophytic
Phycomycete while E. kansanum was lacking in mycorrhizae. Detailed
investigations of the Equisetums are yet to be made.

Mycorrhizae and Mycothalll of Lycopsida: — Lycopodium has
received much attention. Following Treub's discovery of fungal in-
fection of a Javan lycopod, Bruchmann (1885) described similar

Kelley — 22 — Mycotrophy

infection for L. annotinum, the mycelium being both inter- and intra-
cellular. Next, GoEBEL (1887) told how that in L. inmindatum the
lower non-meristematic part of the prothallium is always inhabited by
a fungus. Janse followed with reports on 8 Javan species, several
being reported for the first time. Holloway (1920) has added much
to our knowledge of Australasian species, hence Europe and the
Austral region have been partly covered but America has offered but
two papers on lycopodiaceous fungal symbionts, — by Spessard (1922)
and by Stokey and Starr (1924). Americans have produced six
papers on mycorrhizae of ferns, two on Lycopods, and none on the
hepatics.

Sixteen species of Lycopodiiim are reported mycorrhizal in the
sporophyte while as to the gametophyte, the long-sought gametophytes
of Lycopodiiim, discovered by Fankhauser in 1873 at Emmenthal,
have likewise received attention and found to contain an endophyte.
Whether this endophyte is a Pythium or an Ascomycetous fungus as
Spessard claims, or of different sorts in different prothallia remains
to be determined. The endophyte appears to be a mutualist.

Selaginella has received slight attention by students of symbiosis.
Bruchmann (1897) found 5. spinulosa mycorrhizal in the Alps while
6". helvetica was not mycorrhizal. Janse said that Selaginella in Java
possesses a fungus in the hairless roots. American species of Selagin-
ella have never been reported upon for mycorrhizae.

The little family of the Psilotaceae which is segregated from the
Lycopods and with similarities to the fossil Sphenophyllineae, has
attracted a number of investigators most of whom agree that these
plants are mycorrhizal; yet Costantin (1925, 1936) maintains it has
been found without endophyte, not alone by himself but also by Noel
Bernard. Solms-Laubach, Janse and Bernatsky were earlier
students of the mycorrhizal condition, the last trying to isolate the
fungus ; while Shibata described cytological detail and called attention
to phagocytosis occurring in the tissues and the similarity of the
process to that occurring in the orchids. All of this work was done
on the sporophyte but the gametophyte — a small colourless tuber
embedded in humus — is likewise infected. A gametophyte supposed
to be that of Psilohnn was described by Lang but a detailed report was
given by Darnell-Smith (1917). Sporelings of Psilotum are pene-
trated by an endophytic fungus after a comparatively few cell divi-
sions and soon almost all of the cells of the prothallium are filled
with a skein of hyphae, reports the author. Presence of the fungus
does not cause a change in form of cell but the nucleus is frequently
obliterated by its mycelium. Infection occurs near the growing point :
hyphae are non-septate and two may occur at once in a rhizoid.

Lecture II — 23 — Occurrence of Mycorrhizae

Hyphae have also been observed in antheridia and in canal cells of
the archegonium but never in the egg-cell.

The work on Psilotiini has presumably been done on P. trique-
trum from which we may turn to the other genus of the family,
Tmesipteris, a genus of Australian herbs. In the first of several papers
on mycorrhizae, Dangeard (1891) described endotrophic mycorrhizae
from five species of Tmesipteris, telling of the fungi and their appear-
ance in the root ; and it would seem that he may have been dealing with
both parasitic and mycorrhizal fungi. But he describes and figures
Hartig net, hyphal coils, and notes disappearance of starch from the
infected region. It is curious that the useful term, "mycorrhizome",
should have been invented for the service of these plants so little
known to the general botanical public; yet Dangeard said that as
roots are wanting in these plants, they may be said to possess mycor-
rhizomes. So came into being a designation for all endophytic creep-
ing stems, especially amongst ferns and orchids.

The gametophyte of Tmesipteris, like that of Psilotum, contains
an endophyte ; for Lawson (1917) in a paper complementary to that
of Darnell-Smith described the infection in T. tannensis. Structu-
rally the prothallium of Tmesipteris does not resemble that of Ly co-
podium but it does that of Psilotuin.

Gymnospermous Mycorrhizae : — In all classes of Gymncsperms
there are found mycorrhizal fungi occurring as endophytes. Among
all the branches of the plant kingdom, none has attracted more re-
search than that of the conifers ; and especially have the pines been
investigated. As early as 1865 Nicolai had unwittingly described the
mycorrhizal character of pine rootlets although it was not until 1873
that Reinke remarked the similarity of the cortical "thickenings" of
the pine rootlets to those of the liverworts known to be due to fungi.
Several reasons may be adduced for predilection for pine mycorrhizal
research : first, modern mycorrhizal research began with the pines ;
second, the pines are easily studied ; third, they are of great economic
importance. About one-fifth of all mycorrhizal research in the last
decade has been done with Gymnosperms, and of these principally
pine, spruce and fir.

Mycorrhizae of Gycads: — Tubercles of the Cycads appear to
harbour both fungi and bacteria and are modified rootlets. For them
the name of "consortium" has been proposed, a name first suggested
by Grisebach according to Reinke (1871) who quaintly observed
that the term is "sehr zutreffend". Life, who made an extended study
of these "consortia", declared that "In reference to the symbiotic rela-

Kelley — 24 — Mycotrophy

tions which exist between these various organisms it is difficult to
speak with any certainty . . . the tubercles of Cycads may be said to
have at least two functions, that of aerating and that of assisting nitro-
gen assimilation." But whatever their structure or function, they can
be considered only as a very special case of mycorrhiza, and the same
may be said for the "nodules" of Encephalartos; while those of
Macrosamia are reported by McLuckie (1922) to be purely bacterial.
Spratt (1915), whose work on Cycadean nodules comes nearest to
being monographic, states that all Cycadean genera produce root-
nodules which are perennial modified lateral roots, repeatedly branched
and forming large coralloid masses. They are primarily produced, he
says, by infection with Bacillus radicola ; and he asserts that the Cyca-
deaceae are the only nodule-bearing plants known in which four
organisms are associated together symbiotically, viz. two nitrogen-
fixing bacteria, an alga, and the cycad.

Mycorrhizae of Ginkgo: — Only one living member of this genus
occurs and this member, the Maidenhair tree, has long since ceased to
exist in a wild state. It is thus in the nature of an exotic wherever it
grows, and its rooting conditions and structures are in a sense anoma-
lous. Perhaps no other plant, the lone representative of its order,
presents such an unique case ; yet Ginkgo is reported mycorrhizal. Its
earliest observer was Reinke (1873) who noted "thickening strips"
in its root cortex ; its latest observers were Klecka and Vukolov
(1935) who state that the mycorrhizae are racemose, slightly furcate.
Yet ScHWARZ (1883) and von Tubeuf (1896) reported abundant
root-hairs for this species. An ecological study of Ginkgo roots in the
native haunts of the species, so far as China could provide "native
haunts", would be desirable.

Mycorrhizae amongst the Taxaceae: — The curious tubercles
and necklace rootlets of various Podocarpi have proved fascinating
to students of root structure. There is something which arouses
curiosity in them : the roots are excavated, some lumpy excrescences
appear, and forthwith the botanist hurries to his laboratory to see what
meaneth this strange thing! A mere ordinary rootlet is passed by as
commonplace : for example, the possible mycorrhizae of Torreya are
virtually unknown, apparently because there is nothing about them
to attract curiosity. But thanks to studies of the curious we have much
on record about the nodules of ten species of Podocarpus. It appears
that these nodules are called forth by bacterial action as well as by
fungal invasion ; but the consensus of opinion seems to be that they
are often true mycorrhizae, being developed usually by a symbiotic

Lecture II — 25 — Occurrence of Mycorrhizae

fungus although in cases a fungus is lacking. Reinke (1873), who
saw much and described well, noted "thickening strips" in Podocarpus
cortex; Berggren (1887), with the meticulous exactness of a
Scandinavian, described in detail the pearl-necklace rootlets of the
Podocarpineae, seeing in them a similarity in function to the velamen-
covered roots of orchids. The similarity cannot be drawn too closely,
however, for Hiltner (1899), in conjunction with Nobbe, demon-
strated fixation of atmospheric N by roots of Podocarpus, which he
considered as true endotrophic mycorrhizae ; yet Hiltner suggests
that Heaths and Orchids likewise may fix nitrogen. McLuckie
(1923) also found the Podocarpineae active in N fixation, stating
that the process was accomplished by bacteria present in the cortical
cells. On the other hand, Saxton (1930) was unable to find bacteria
in Tasmanian material of Podocarpus : "No trace of bacteria could be
found but unmistakable and well-preserved mycorrhizal filaments."

Hiltner had considered these nodules as unformed roots but
McLuckie (1923) says that the nodules are modified lateral roots
and arise from the pericycle, their normal growth being checked before
they emerge from the cortex of the main root. Root-hairs, he says, are
commonly present as von Tubeuf had already stated. Yet it is neces-
sary to be careful about accepting reports of root-hairs on mycorrhizal
roots too readily, for setae of the fungus often simulate root-hairs ; and
McLuckie himself says that the surface of the nodule and the main
root is frequently invested with a loose tangle of fungal hyphae.

It is interesting to note that the term "prosporidi" of Petri (1903)
was originated from a study of species of Podocarpus growing at
Florence in Italy. These spore-like bodies produced by the fungus, the
sporangioles of Janse, he called "prosporidi" on account "al loro
significato morphologico piu probabile". Shibata (1902) described
in some detail the fungal structure and reaction, and reputedly demon-
strated an enzyme in the mycorrhiza.

In addition to the nodules of Podocarpus are the mamillate or
pearl-necklace rootlets described so well by Janse (1897). In P.
cupressus he found intermittent growth : "En general, apres une
courte interruption, la croissance reprend pour s'arreter encore une
fois des qu'il s'est forme en second mamelon spherique au sommet du
premier. Cette croissance intermittente pent se repeter ainsi plusieurs
fois de suite, mais au plus tard apres le developpement du cinquieme
mamelon I'arret est definitif ." Janse continues with a detailed descrip-
tion of the histological structure and origin of these mamelons or
pearl-necklace mycorrhizae which are so widely found amongst coni-
fers, casuarinas, Liquidamhar, Acer, Celtis, and others.

Kelley — 26 — Mycotrophy

The genus Cephalotaxus is virtually uninvestigated, for aside from
notes by Reinke and von Tubeuf, there is only an observation by
Prat (1926) that plants of this genus were abundantly mycorrhizal
in the Arboretum at Angiers in France. Torreya has exactly the same
record ; and there is nothing whatever on record of the American
Torreya which lingers in the Appalachicola hills of Florida. Taxus,
being of more familiar presence, is better known as to its mycorrhizae :
the older generation of mycorrhizal students noted it and in more re-
cent days several have described it, particularly Prat (1926, 1934),
who has made rather thorough studies of, first, the European T. bac-
cata, and, second, the Canadian T. canadensis. The mycorrhizae in
Taxus appear to be endotrophic mamelons or pearl-necklace beaded
rootlets, and phagocytosis occurs in them. In his later paper Prat con-
cludes that there is not a true mutualism but that the tree is a parasite
on its parasite! Kle6ka and Vukolov (1935) record for T. baccata
endotrophic mycorrhizae comparable to those of Ginkgo.

The genus Pherosphaera, sometimes doubtfully included in the
Taxaceae, was studied by Saxton (1930), who found both species
provided with nodules, but the Tasmanian species produced nodules
more freely.

Mycorrhizae in Pinaceae: — Pines, orchids and heaths, — these
are the mycorrhizal plants par excellence ! Frank brought mycor-
rhizal study to the fore by his studies on pines and much of recent
research has been concerned with these important economic trees. The
first genus in the family for our consideration is Juniperus, the com-
mon juniper which, like the Yew, is of familiar presence. Its mycor-
rhizae are endotrophic (vide KleCka and Vukolov, 1935) and neck-
lace-beaded but as Janse (1897) observed: "Les mamelons sont en-
core plus allonges et plus rares que chez le Cupressus. Au demeurant
ils leur resemblent beaucoup". Sarauw (1903) observed that in this
species (the common juniper) an endotrophic mycorrhiza exists in con-
junction with an Hartig net, which he says is the only case of the sort
known, except that in Cedrus Deodara there is an Hartig net without
a mantle. In recent days the mycorrhizae of /. communis have been
monographed by Lihnell (1939) in an extended and well-illustrated
paper. The American species of Juniperus are very little studied :
McDouGALL and Jacobs (1927) state that /. monosperma is endo-
trophically mycorrhizal in the Central Rocky Mountains ; Henry
(1936) states that no mycorrhizae occur in /. sibirica and in /. utahen-
sis. J. sibirica Burgsd. is the same as /. communis L. var. montana Ait.,
and /. communis is well known to be mycorrhizal in Europe.

Lecture II

— 27 —

Occurrence of Mycorrhizae

The genus Cupressus has attracted no modern investigator except
that Birch (1937) says that in New Zealand the fungus Rhisopogon
rubescens appears to be a mycorrhizal symbiont of C. macrocarpa. For
the Italian cypress, C. sempervirens, mycorrhizae were described by
Janse and by Kirchner (1908), the former describing necklace-
beads and the latter simple coralloid mycorrhizae with endotrophic
mycelium. Berggren (1887) had stated that Hartig net is lacking;

Fig. 2. — Mycorrhizae in Pinus virginiana. A
"long-root" is beset with mycorrhizal short-roots
or mycorrhizae, which in the older portion exhibit
beginning of coral branching by dichotomy. The
mycorrhizal sheath or mycoclene over the apex has
split by renewed growth.

Yeates (1924) that the fungus is similar to that in Taxads. One
other species, C. Lindleyi, was reported to have no root hairs by
SCHWARZ (1883).

Chamaecyparis is even less studied than Cupressus, having no
modern investigator except that Kle5ka and Vukolov (1935) list
C. Lawsoniana as having endotrophic mycorrhizae. Noelle (1910)

Kelley — 28 — Mycotrophy

reports for Thuopsis dolobrata and for the Incense cedar, Lihrocedrus,
which last Yeates (1924) says contains a fungus similar to that in
taxads. Thuja stands in a better position, being a more abundant
tree in the cool temperate zone where most mycorrhizal students have
lived. T. occidentalis and T. orientalis are both well studied while T.
plicata and T. Standishii are reported as mycorrhizal. Two recent
papers have cited the genus, — Klecka & Vukolov (1935) and
DoMiNiK (1937). Taxodium distichum has been listed as having en-
dotrophic mycorrhizae but Sequoia — more attractive to curiosity — has
been more studied. Both species of Sequoia possess endotrophic
mycorrhizae, it would appear, and according to Strasburger, root-
hairs are entirely wanting. Oddly enough, it is only European material
of Sequoia that has been investigated while Calif ornians neglect their
most famous tree. Crypfoineria is reported mycorrhizal in Europe :
VON TuBEUF found root-hairs wanting in C. japonica although he
notes that Klebs found sparse hairs on seedlings, which hairs were
sloughed ofif with the outer cell layer, Mimura (1933), working at
TokyO', states that mycorrhizae are wanting on this species when
planted at the Experiment Station but were found on roots that had
grown from the pots into the ground. Cunninghamia, the China fir,
is reported mycorrhizal by Noelle (1910) and by Yeates (1924) ;
Sciadopitys, the monotypic Umbrella pine, by Noelle and by Laing
(1923), the last describing the histology in some detail.

Four species of Araucaria are termed mycorrhizal. Janse com-
pared its rootlets to those of Podocarpus but thought they were rather
larger. Of modern writers we may note Rayner (1938) who in a
review states that A. Cunninghamii grew in Nyassaland without inocu-
lation of the soil ; Young (1938) found that lime-induced chlorosis in
this Hoop-pine was eliminated from some Queensland nursery beds
by sulphur applications, and the same author found by pure culture
experiments that its seedlings produced endotrophic mycorrhizae when
grown in association with the fungus Boletus elegans and failed to
develop in the absence of a mycorrhizal fungus.

The genus Abies has not proved attractive to our students although
it appears to be mycorrhizal and material is abundant. Fifteen species
of the genus are cited as mycorrhizal but without detailed description
and with no details of physiological relationship. Of recent workers
we may cite Dominik (1937) who notes three exotic species mycor-
rhizal in Poland; Colla (1931) who found three Basidiomycetes on
A. alba in Italy; Tazoye (1940) who cites A. Mayriana as mycor-
rhizal in Japan; Henry (1936) who says that dwarfed A. lasiocarpa
in the mountains of Utah is an excellent mycorrhizal host (Mc-

Lecture II — 29 — Occurrence of Mycorrhizae

DouGALL, 1927, had cited the same species from Idaho) ; and Klecka
and VuKOLOv (1935) who Ust A. alba as ectotrophic.

Pseudotsuga is not popular with mycorrhizal students : but one of
the four species has been investigated at aU for mycorrhizae and this
species only in a cursory way. In its native haunts it was described as
both ecto- and endo-trophic by McDougall (1927) in Utah, and as
having an endophyte in Canada by Lewis (1924) ; while Laing
(1923) insists that this species does not form mycorrhizae readily.
Klecka (1935) and Dominik (1937) find the species mycorrhizal in
Europe while Birch (1937) records its fungi in New Zealand. Tsuga
has received passing attention : five species are noted as mycorrhizal,
of which one species, T. heterophylla is said by Laing to have semi-
ectotrophic mycorrhizae the hyphae being found only between the
cortical cells, and there is no mantle.

Twelve of the 39 species of Picea are cited as mycorrhizal and one
of them, P. Abies, the Norway spruce, has been studied in detail by
several investigators, especially Melin (1925). Using Melin's cul-
ture methods, Modess (1939) synthesized various Hymenomycetes
with seedlings of P. Abies, a study further reported in 1941 when he
Usted 8 species of Hymenomycetes and one Gasteromycete that formed
mycorrhizae with this spruce. Melin's methods were likewise used
by Fries (1942) in synthesizing monospore mycelia of Scleroderma
aurantiiim with spruce whereby mycorrhizae were formed but not as
abundantly as with pine. Also in Sweden, Lindquist (1939) did his
work on spruce and wrote philosophically on the physiology of myco-
trophism; and Rom ell (1938) reports on his trenching experiments
with spruce, and their bearing on the problems of mycotrophy. In
another cultural study, Bjorkman (1940) reports on the ecology
of the mycorrhizae of this spruce, while Thomas (1941) presents a
plot study of young spruce plantations in the Rhine Valley. Besides
the Norway spruce studied in Europe, various other spruces have been
studied or noted in America and Japan.

Larix shares with Picea the attention accorded by Melin. Five
species of Larix are recorded as mycorrhizal by various authors ; and
more recently How (1942) has made a monographic study of the
mycorrhizal relations of L. decidua. Colla (1931) records Hypho-
lonia fasciculare with the same species of larch; and Thomas (1941)
notes larch plantations in the Rhine Valley. Pseudolarix is recorded
onlyby Noell (1910).

Finally, in the Coniferales, we come to Pinus, the most studied
genus of Gymnosperms. Thirty-seven species and varieties of Pinus
are recorded mycorrhizal but of these only four have been studied
in detail, namely, Strobus, sylvestris, pinaster and montana.

Kelley — 30 — Mycotrophy

Mycorrhizae in Gnetaceae: — There are but two references to
the possible mycorrhizal condition of these plants : ( 1 ) Strasburger
said that root hairs are exceptional in Ephedra, while (2) Kirch ner
remarked that root fungus was not observed by von Tubeuf and that
root-hairs are not exceptional but found covering the roots for a dis-
tance of 2-3 mm.

The Method of Opportunism: — To summarize what is known
of the mycorrhizae of Gymnosperms, therefore, one must say that
much is known of a few pines and spruce and larch but that there is
no general research upon the occurrence of mycorrhizae in the class as
a whole. The same method of opportunism rules with the Angio-
sperms : there have been few scientific approaches to the mycorrhizae
of higher plants through a systematic investigation of their occurrence.
A few papers such as those of Janse and of Schwarz point the way
to a more thoroughgoing study of the rooting structures of Angio-
sperms ; and meanwhile one pieces together the isolated papers to
form the following picture.

Mycorrhizae in Apetalae: — First as to that collection chiefly of
trees which has been called the Apetalae one notes that many are re-
corded mycorrhizal ; indeed, the oaks and beeches are, with the pines,
much studied plants. Of the poplars and aspens, Popiilus, seven
species have been studied although not in much detail but their my-
corrhizal character is established: KleCka and Vukolov (1935) are
their only modern students. Sixteen species of Salix are given a
similar character by various reporters, Klecka and Asai being the
most recent. The Garryaceae are unreported, for Mexico and the
West Coast are almost untouched mycorrhizal fields. But the Myri-
caceae are much investigated because of their root-nodules which are
true consortia (or mycodomatia), being occupied by bacteria and
fungi simultaneously; and they are present in all members of the
genus that have been studied — which are five of the 35 listed for the
genus. Most of the work on Myrica has been done in Europe and
Asia, almost none in America ; but the American Comptonia is listed,
by Kellerman. Leitneria, monotypic genus of the Corkwood family
is unreported, and so, too, are the Asiatic Platycarya and Pterocarya;
but the walnuts (Juglans) are recorded mycorrhizal. It is to be
observed that Frank and Stahl both stated that /. regia is not
mycorrhizal, KleCka (1935) calls it ectotrophic, while the few
reports on the two American species term these latter endotrophic.
No detailed study of the walnuts is in print, nor of the hickories
(Carya) except for that of the pecan (C pecan) by Woodroof

Lecture II — 31 — Occurrence of Mycorrhizae

(1933). It is curious that trees so abundant as the hickories and so
comparatively important should have escaped attention. Likewise the
Betulas, for birches flourish in countries where mycorrhizal workers
live ; yet there is not much to report on them. The two more recent
studies, of Laitikari (1934) and of Bjorkman (1941) are con-
cerned with the root system in general and with the concomitant fungi.

But with alder the case is different, for alder has "root excres-
censes" that attract curiosity; and two European species attracted
much attention in earlier days, Klecka (1935), Plotho (1941)
and Cernik (1937) being their modern students. Harshberger
was the only student of American alder while brief citations have
come for Japanese species. Plotho (1941) tried the synthesis of
alder nodules, which appears to be the only experimentation of the
sort on record. Two species of Carpinus are reported mycorrhizal,
the American species of Ostrya, and three species of Corylus. Com-
ing, then, to the Fagaceae we meet with beech which, for some rea-
son, has ever been popular at least in Europe, the latest study being
by Harley (1939). It was on beech that Meyen (1829) observed
his "pseudomorphose" of the roots which may have been an unwitting
discovery of mycorrhizae. Of chestnut, mycorrhizae were described
on the European species by earlier students who thought to find in
them a cause of disease of that economic tree. Chestnut provided
Kelley (1940) with his material for discovering the essential simi-
larity between blight and mycorrhizal infection. But the Californian
Castanopsis and Lithocarpus, with many species in Asia, are yet
untouched. The oaks {Qiiercus) include "more than 200 species"
of which 23 have been noted as mycorrhizal, one species only
(,Q. robur) having received some careful attention. Since oaks are
preeminently American they offer a splendid opportunity for study
of a vital function in important timber trees, especially open to those
who say they have the interest of forests at heart, — a field of re-
search that is virtually untouched.

Elm (Ulmus) was noted by one of our earliest students,
Duhamel (1758), and since his time 5 species have been listed but
no detailed study of any member of this genus exists. A couple of
reports of "fungus-free" may be covered by Stahl's statement:
"Wenn auch feineren Ulmenwurzeln des Oefteren sich pilzfrei
erweisen, so trifft man doch hie und da innere Verpilzung."
McDouGALL (1928) and Janse (1897b) say that Celtis is mycorrhi-
zal, and AsAi reports the same for Zelkova (Abelicea hirta) but
otherwise nothing more is known of the mycorrhizae of the Ulmaceae.
Similarly, 3 species of Morus are reputed mycorrhizal, being, like
Ulmus, endotrophic; but we are as innocent of exact knowledge of

Kelley

32 —

Mycotrophy

the mulberries as of the elms. Several Ficus have been termed
mycorrhizal.

As to the herbaceous members of the Apetalae : Asai says that
Boehmeria is not infected ; Peyronel states that Urtica is : we can
say no more for the Urticaceae. Asariim appears to be mycorrhizal
in Europe, America and Japan by a single report in each case ; Rumex
by two reports ; Polygonum by several. P. viviparum was said by
Stahl to have "innere Verpilzung" while Asai and Takamatsu both
state that certain Japanese species lack mycorrhizae. Phytolacca
decandra also does not have any according tO' Asai. Two species of
Atriplex are said to be mycorrhizal, the woody species wxre not in-

FiG. 3. — Mycorrhizae in Sugar Maple, Acer saccharum. A race-
mose system of mycorrhizae which are "beaded", due to periods
of quiescence and renewal of growth. This mode of growth is
characteristic of Acer, Ilex and other genera.

vestigated ; Beta vulgaris, the common beet, is mycorrhizal but not
Salsola, Amaranthus sylvestris was termed mycorrhizal by Trotter
and Scleranthus annuus by Schlicht and by Stahl, while Chenopo-
dium is mycorrhizal (Schlicht) or not mycorrhizal (Asai).
Portulaca is positive while of the CaryopJiyllaceae Gypsophila,
Arenaria, Stellaria, and Cerastium are in the plus column while
Dianthus, Silene and Sagina are negative. This is the record of the
herbaceous Apetalae.

Amongst the numerous Apopetalae and Gamopetalae there is a
similar scantiness of information, the larger families showing a
number of genera that have been examined casually for mycor-

Lecture II — 33 — Occurrence of Mycorrhizae

rhizae while smaller families are entirely ignored. Thus, one dozen
genera of Ranuncnlaceae are reported mycorrhizal and twelve of
Cruciferae; twenty-six genera of Papilionaceae are reported which
is the record for the Dicotyls ; but as many or perhaps most of
these cases are bacterial they are dubiously considered as mycorrhi-
zal. The next largest record is of the Compositae with 25 genera
reported.

There are conflicting reports : Thus, Podophyllum is termed
mycorrhizal by Lohman and non-mycorrhizal by MacDougal;
Benzoin is mycorrhizal according to Henry but non-mycorrhizal ac-
cording to MacDougal; Ailanthus is non-mycorrhizal according
to Stahl and to Duthie but endotrophic according to KleCka and

VUKOLOV.

Then there are families that are listed on the basis of single
reports, as the Calycanthaceae (Asai), Menispermaceae (McDougall
& Liebtag), Sarraceniaceae (MacDougal), Pittosporaceae
(Asai), Sterculiaceae (Asai), Tamaricaceae (Stahl), Cistaceae
{Vtrotta) , Nyssaceae (Henry), Diapensiaceae (Asai), Myrsinaceae
(Asai), Phmibaginaceae (no mycorrhizae according to Costantin) ;
and these reports are not confirmed nor amplified.

In a number of families there is an amazingly large number of
genera yet to be examined : Thus, in the Borraginaceae with 85
genera and 1500 species but 4 genera and 6 species have been stud-
ied for mycorrhizae; in the Labiatae but 10 genera and 14 species
have been studied amongst the total of 160 genera and 3000 species ;
in the Scrophiilariaceae but 6 genera and 18 species have been
studied amongst 180 genera and 3000 species; while the Bignonaceae
with 100 genera and 600 species is entirely untouched. In the
great family of Compositae with its thousands of species there are
but 54 studied for mycorrhizae.

In many of the Dicotyls we would expect to find phycomycete
mycorrhizae as in the Violaceae of which 12 species are termed
mycorrhizal but we have no detailed studies upon them. Two species
of Liniim, 3 of Oxalis, 3 of Hypericum, 7 of Epilohium, five of
Primula, six of Campanula are listed as mycorrhizal ; and we await
further information as to whether the concomitant fungi are phyco-
or basidiomycetes.

Several special cases may be noted : Thus the insectivorous
plants have received some attention, the Droseras at the hands
of Frank, Hoeveler, Schlicht, and Peyronel; Sarracenia, of
MacDougal. Stahl was interested in the Polygalas, terming them

Relley — 34 — Mycotrophy

endotrophic. Four species of Euphorbia are cited but Asclepias
which would seem to be of equal interest as a lactiferous plant has
escaped observation except for two reports on A. syriaca.
D'Angremond and Hell (1939) describe endotrophic mycorrhizae
for Hevea. Three species of Cactaceae are cited, by Johansen (1931)
except that Asm also cites N eomammillaria. Monotropa was long a
focal point of interest. Twenty-one species of Gentiana are cited,
and Oholaria is noted.

The woody Dicotyls have fared better, and we may run briefly
through the list: The Magnolias and Liriodendron, and the tropi-
cal Talaiima, Manglietia and Michelia (according to Janse) ; Merati
of the Calycanthaceae (Asai) ; Asimina of the Anonaceae ; Sassa-
fras, Benzoin and Ocotea of the Lauraceae ; Pittosporiim (Asai) ;
Liquidamhar , Altiugia and Haniamelis; Platamis; 9 genera of the
Malaceae, four of the Rosaceae, nine species of Prunus; two of the
Mimosaceae, 3 of the Caesalpinaceae and 7 of the Leguminosae ;
four of the Riitaceae (including 3 spp. of Citrus) ; Picrasma and
Ailanthus; Melia and Dysoxylum, two species of Buxus; 3 of the
Anacardiaceae, 3 species of Ilex, 2 genera of Celastraceae; 3 of
Staphyleaceae; 8 species of Acer; 7 of Aesculus; 7 genera of Sapin-
daceae; 4 of Rhamnaceae; 2 of Vitaceae; 5 species of Tilia; Firniiana
of the Sterculiaceae ; Thea; Tamarix; 8 species of Daphne, 3 genera
of Elacagnaceae ; Nyssa sylvatica (Henry) ; 3 spp. of Eucalyptus;
3 genera of Araliaceae, 2 of Cornaceae; Clethra (Asai) ; 21 genera
of Ericaceae; including 13 spp. of Vaccinium; Diapensia (Asai) ;
Ardisia (Asai) ; Diospyros (Asai) ; Symplocos (one species out of
the 290 spp. in this monotypic family !) ; Styrax (Asai) ; 5 genera of
Oleaceae; 2 species of Nerium; Gardenia (Asai), 4 genera of
Caprifoliaceae.

For more recent work we may note: Milanez (1940) records
root fungi for Citrus aurantifolia, said to be the first record for
South America, but he considers them as parasites; Muller (1936)
reports on mycorrhizae of citrus in the Netherlands Indies; Reed
& Fremont (1935) and Rayner (1933) describe a phycomycete
mycorrhiza for Citrus and regard it as beneficial under certain con-
ditions. Berkeley (1936) states that raspberry roots (in Canada)
show a phycomycetous infestation similar to that in strawberry, as
recorded by Richards & McKIay (1936). Bouwens (1937) con-
sidered the strawberry endophyte to be a Rhisoctonia, which generic
fungus was alsO' responsible for mycorrhizae in quince (Cydonia).
A phycomycete mycorrhiza is described likewise for almond {Amyg-

Lecture II — 35 — • Occurrence of Mycorrhizae

dolus) by RuGGiERi (1937). In Cacao in Trinidad, mycorrhizae
also occur (Pyke, 1935; Laycock, 1945), although not invariably.
Sabet (1939) describes mycorrhizae for cotton (Gossypium), and
TuNSTALL (1940) for Thea.

For other plants, Heath and Luckwill (1938) report my-
corrhizae in Potentilla and several other heather-land plants ; while
Malan (1938) studied mycorrhizae of alpine legumes (finding
them phycomycetous). For Ericaceae, Barrows (1936, 1941)
studied Epigaea, and Freisleben (1933, 1934) particularly Vac-
cinium; Gordon (1937), Rhododendron; Bain (1937), after study-
ing Oxycoccus, comes to the conclusion that there is no obligate
symbiosis while Rayner & Levisohn (1940) contradict him;
MoLLiARD (1937) after studying Calluna, concludes that mycor-
rhizae are not essential. For potato (Solamim), Costantin (1935,
1936) and Joseph (1935) present data. Kurbis (1937) and
Kelley (1943) have described mycorrhizae for Fraxinus;
ScHiMMLER (1937) for 12 spp. of Gentiana.

Since Monocotyls are not woody, less interest can be expected
in them. It is true that there are some monocotyledonous trees which are
reported mycorrhizal, — the palms Phoenix and Livistona, the screw-
palm, Pandanus, and the banana "tree", Musa. But most Mono-
cotyls are herbs, and many are aquatic plants in which no mycor-
rhizae are found, as Typha (Asai) ; Alisma (Asai) ; Calla palus-
tris; Acorus, 2 spp. ; and 10 spp. of Juncus which, however, pro-
duce root swellings that do contain a fungus according to Magnus.
No mycorrhizae are reported for the Cyperaceae; vis. 2 spp. of
Cyperus, 3 spp. of Eriophorum, and 14 spp. of Carex; but numer-
ous species of grasses are reported mycorrhizal. For the Gramin-
eae, Asai (1934) reported 23 species mycorrhizal and 4 not my-
corrhizal, the latter all hygrophylls; while 58 spp. were reported
mycorrhizal by other observers. The latest researches on grasses are
by BiRAGHi (1936) on cereals, and Neill (1940) on Lolium. Of
the aroids, Arisaema is mycorrhizal (Lohman. 1927), while Magrou
(1937, 1939) used Arum for isolation of the endophyte.

The Liliales seem richly mycorrhizal : Veratrum in the Melan-
thaceae, Allium, Lilium, Tulipa, Erythronimn, Ornithogalum,
Muscari, Hemerocallis, Yucca, Fritillaria, Scilla and Aloe in the
Liliaceae; Asparagus, Smilicina, Maianthemum, Uvidaria, Poly-
gonatum, and Convallaria in the Convallariaceae; and Trillium in
the Trilliaceae. Oddly, there is no report for Smilax. Then Nar-
cissus, Galanthus, Leucojum (Stahl) and Agave (2 spp.) of the

Kelley — 36 — Mycotrophy

Amaryllidaceae (but no mycorrhizae in Aletris [Takamatsu] ) ;
Dioscorea (Asai) ; 4 genera of Iridaceae; Zingiber and Musa of
the Scitaminaceae ; Ananas of the Bromelidaceae ; while the Bur-
mannias have attracted much interest, the latest record by Ciferri
(1946). The orchids would require a separate section to do them
justice, for no less than 85 genera are described as mycorrhizal while
20 papers on orchid mycorrhizae have appeared in the last decade.

Lecture III

THE FUNGAL ENDOPHYTES

Nature of the Mycorrhizal Fungi: — It scarcely needs to be
said that mycorrhizal fungi are not a separate taxonomic unit in the
classification of fungi. They are the ordinary soil fungi of forest
and woodland, of meadow and cultivated field. Neither are they
special members amongst the congeries of soil fungi in the sense that
one, and only one, member can achieve a mycorrhiza. One fungus or
another can produce it, and ordinarily there may be several fungi par-
ticipating, forming what has been called a "multiple mycorrhiza". In
other words, the fungi living in the soil grow into plant roots as into
a part of their environment, and, if the host plant is able to check the
fungus in its rootlet cortex and break down the fungal hyphae, the
association is said to be mycorrhizal. Presence of the fungus, re-
gardless of its taxonomic identity, has little to do with the form of
the mycorrhiza, which is characteristic for a given host plant and is
determined by the host. In an informing paper by Magrou, Douchez
& Segretain (1943), it is shown that mycorrhizae are formed with
potato by various endophytes some normally present with monocoty-
ledonous, some with dicotyledonous plants. The endophytes present
in various soils simply grew into the potato roots and gave the stimulus
to the production of characteristic tubers. It was the potato plant that
determined tuber form, not the fungus.

The mycorrhizal association, therefore, appears more as a casual
thing than as an occult and premeditated action that can be achieved
only by special, designated actors. It is true that certain fungi do seem
more or less confined to certain mycorrhizal hosts, although specifi-
city cannot be said to be absolutely proved ; but a certain amount of
specificity could be posited on the grounds of chemical affinities.
The emphasis that has been placed on mycorrhizal fungi would seem,
therefore, to be somewhat exaggerated because in so many cases
the identity of the fungus seems a relatively inconsequential thing.
It is nutrient that the higher plant requires and in many cases it seems
of little moment whether the particular fungus which supplies the
nutrient happens to be a Russula or an Amanita, a Boletus or a Tri-
choloma. These are the fungi of the forest floor and naturally have to
be the mycorrhizal fungi of the trees that grow there. It would seem

Kelley — 38 — Mycotrophy

logically deducible that the only fungi available to forest trees for
formation of mycorrhizae would be those of the forest soil ; while
phycomycetes of cultivated ground are available to crop plants. Never-
theless, there is a physiological separation possible amongst soil fungi,
according to Melin (1925), whO' recognized three groups of these
fungi, vis., symbiophiles, saprophytes and parasites. All "mycorrhi-
zal fungi" are considered as symbiophiles.

Some investigators, wishing to prove that sporophores of Russula,
etc., which appear on the forest floor are actually part of the mycor-
rhizal mycelium, have laboriously traced that mycelium from the
sporophore to the mycorrhiza and thereby established, so they said,
the identity of that particular mycorrhizal fungus. But their success
was denied by other investigators who asserted that attachment of a
sporophore to a mycorrhiza is no proof whatever that the fungus
concerned is mycorrhizal ; for who can say but that this sporophoric
fungus is not a secondary parasite? Therefore, say these later stu-
dents, the only thing to do is to grow the fungi in pure culture, in-
oculate them into sterile seedlings, and if a mycorrhiza results there
is positive proof of the identity of the mycorrhizal fungus. But is
there positive proof ? Laboratory experiments show what can happen
in the laboratory but not what happens in nature. A laboratory syn-
thesis of Boletus granulatus with pine shows by its production of a
mycorrhiza that this fungus is capable of such production but it does
not prove that mycorrhizae produced on pine in nature were pro-
duced by B. granulatus. They might have been produced by another
fungus growing on the same area. When only a single fungal species
has formed sporophores over the roots of pine and when that species
is shown by synthesis-experiment to be able to produce mycorrhizae,
then it can be said with justice that this species is the mycorrhizal
fungus in question ; but one could have come to that conclusion with-
out experiment. Or, to use Romell's (1939) illustration: Lacta-
rius delicosus has been grown on pine in the laboratory but in nature
it rarely if ever is found on pine. In other words, the various lines
of research used with reference to mycorrhizal fungi all help to iden-
tify the fungi ; but the question of identity is after all not of major
importance.

In earlier days of mycorrhizal research, it was thought that my-
corrhizae were produced on trees by basidiomycetes and that herbs
in general lack mycorrhizae ; but with greater development of micro-
scope and technique it is known that all major fungal groups furnish
mycorrhizal fungi. We shall consider them in the usual systematic
order.

Lecture III —39-- Fungal Endophytes

Phycomycete Mycorrhizal Fungi : — The records for these fungi
before 1920 are somewhat uncertain because it was not until recent
years that phycomycetous mycorrhizae were regarded as constant
features of nature. It is true that Treub, Bruchmann, and Goebel
had independently found Pythium in prothallia of lycopods; while
Jeffrey had assigned the endophyte of Botrichium to the same
genus. Dangeard had found a chytridiaceous fungus on Tmesipteris
which he regarded as mycorrhizal ; and there are a few other records
of the same sort.

It was Peyronel who brought the "Phycomycete mycorrhiza"
to our attention, commencing in 1922 with a study of cereal grains
that were brought to his station for a study of diseased condition.
Peyronel found that these cereals, instead of being autotrophic,
possessed mycorrhizal infection, — the infection being considered
mycorrhizal because the hosts were "perfectly normal". From this
study Peyronel continued : He saw quickly that endotrophic fungi
are of two major sorts, — the first possessing arbucles and vesicles
and the second only mycelial pelotons (found chiefly in orchids ex-
cept that Mollberg found vesicles in certain orchids). Later (1924)
Peyronel described three species of Endogyne involved in forma-
tion of endotrophic mycorrhizae on herbaceous phanerogams, the
first characteristic of peaty, swampy soils, the second exclusively
hydrophilous, and the third found on Euphorbia dulcis. Other species
of Endogyne were reported in 1937 from the Val Valdesi, producing
endotrophic mycorrhizae on Viola and other herbs. In the same
year, he published on endotrophic mycorrhizae of the Alps at Kleinen
St. Bernhard, and noted that conditions in a cultivated garden were
markedly less favourable for growth of the mycorrhizal fungi than
in the natural habitat.

Interest in the phycomycetous mycorrhizae had been stimulated
by Jones (1924) in a publication in which he recorded the discovery
"that the roots of nearly all our common leguminous crops, wherever
grown, are extensively invaded by a characteristic fungus which has
previously been known as a mycorrhizal fungus. So abundant is this
fungus that it appears unlikely that many plants of alfalfa, clover,
peas, and other legumes ever reach maturity without having their
roots more or less invaded. . . The taxonomic position of the fungus
has not been determined but it appears to belong among the Phy-
comycetes." Jones gave a list of other plants besides legumes in
which this same sort of mycorrhizal invasion had been found.

This paper of Jones' inspired Samuel (1926) to work in South
Australia, and he reported the same sort of infection in 27 legumes,
30 Gramineae, and in herbs of the families Liliaceae, Ranunculaceae,

Kelley — 40 — Mycotrophy

etc. Other workers continued the reports, and, in 1935, Rayner re-
marked on "the remarkably widespread geographical distribution of
this 'Phycomycete type' of mycorrhizal association, its prevalence in
plant species of the most diverse affinities (and) its recorded appear-
ance in certain crop plants." Biraghi (1936) confirmed Peyronel
on the frequence of endophytic infection of roots of cereals, finding
Asterocystis radicis in a majority of cases. Ruggieri (1937) reported
endotrophic mycorrhizae common on fruit trees, a Phycomycete being
constant in root cortex of almond. Berkeley (1936) found a phy-
comycetous mycorrhizal fungus on raspberry in Canada; Richards
& McKay (1936), on strawberry in Utah; and Reed & Fremont
(1935), on Citrus.

In 1939, Butler published a paper devoted to a study of "the
distribution and morphological characters of the vesicular-arbuscular
or Phycomycetoid endophytes which commonly occur in cultivated
and probably other soils forming mycorrhizal associations in the
roots of many flowering plants and cryptogams, including prothalli
of liverworts and of some ferns. The regularity of their occurrence in
some annual field crops is believed to be merely the result of the
greater opportunity to persist indefinitely, by passing from the older
to later developed roots, offered to the organism in perennial plants."
Believing with Peyronel that these fungi belong to the Endogyn-
aceae, Butler cites Dangeard's (1898) name of Rhizophagus for
their genus, and describes the species as R. populinus, R. theae and
R. marratiaceum. Sabet (1939) promptly placed on record the
presence of Rhizophagus sp. as the mycorrhizal fungus of cotton in
the Sudan.

The first reputed synthesis of a Phycomycete mycorrhiza is said
to have been that of the unnamed endophyte of Arum with roots
of A. italicum (Magrou, 1936).

Ascomycetous Mycorrhizal Fungi: — Various Ascomycetes
have been cited in connection with mycorrhizae, as Aspergillus and
PenicilUmn (Ternetz, 1907) ; Terfesia (Pirotta, 1900) ; MoUisia
(Nemec, 1899) ; and Humaria (Nicolas, 1929) ; but Elaphomyces
and Tuber are the most frequently reported of the group. Very
early, Boudier (1876) had noted presence of Elaphomyces on low
ground with Molinia, a grass ; or on higher grounds where Leuco-
bryuni moss was growing. Still earlier (1837), Berkeley had
cited the association of E. miiricatus with beech roots in mountainous
woods. TuLASNE (1841) remarked that E. granulatns is confined
to roots of one sort of tree (not named) and "flourishes when tree
is active." This same species Boudier had found on birch, oak and

Lecture III —41— Fungal Endophytes

chestnut at Nancy in France. Reess (1880) observed coralloid
clusters of mycorrhizae on pine bound with mycelium of Elaphomyces,
while Lewton-Brain (1901) described mycorrhizae of pine formed
in conjunction with E. variegatus. Since then interest in Elaphomyces
has languished.

The genus Tuber also attracted observers, e.g. Frank (1888) who
observed T. aestivum on beech; and more latterly Costantin (1924)
who found that ascospores can be formed by the fungi apart from
mycorrhizal symbiosis. Mattirolo wrote a number of papers on truf-
fles, finding (1934) the mycorrhizal fungus of the introduced Popu-
lus canadensis to be T. Borchii; and he suggested the possible intro-
duction of fungus with the tree when the latter was brought to Italy
from California.

Hemibasidiomycetes : — Of the Hemibasidiomycetes may be
noted the following: Weber (1884) assigned the fungus responsible
for tuber formation in Juncus to Entorhiza of the Tilletiaceae. Lag-
erheim (1888) described a new species of Entorhiza from roots of
Juncus articulatus obtained in Switzerland. The fungus had caused
the roots to form into galls, and within was an abundance of yellow
"spores". In the Black Forest similar nodules were found on the
same species of rush, and similarity to leguminous nodules was pointed
out. Formation of nodules on several species of Juncus was
noted by Schwartz in 1910.

Hymenomycetous Mycorrhizal Fungi: — These are the chief
mycorrhizal fungi. Upwards of 50 genera of Hymenomycetes have
been reported as forming mycorrhizae (or perhaps it should be said,
incriminated in their formation) ; but in most of these cases there
are only one or two species cited in one or two reports. The principal
"mycorrhizal fungi", if numbers of reported species mean anything,
are Boletus, Amanita, Lactarius, Cortinarius, Russula, and Tricholoma.
Commencing with Frank's (1888) observations on Boletus bovinus
with spruce, later observers — almost all since 1920 — have shown by
field observation and synthetic experiment the connection of about
30 species of boletes with various trees. In a few cases a bolete has
failed to form mycorrhizae in synthesis, as B. edulis with pine and
spruce (MoDESs, 1941) ; while B. parasiticus is a parasite as the name
indicates (Smotlacha, 1911). Smotlacha believed that certain
boletes are confined to the neighbourhood of certain trees, as B.
rufus with aspen and B. rugosus with beech.

The Agaricaceae are much investigated mycorrhizal fungi, al-
though Agaricus itself provides few members that are endophytes.

Kelley — 42 — Mycotrophy

Amanita, so common in woodlands of Europe and eastern America,
has been studied, commencing with Boyer's (1915) observation that
the mycelia of many mushrooms, especially of Amanitas and boletes,
extend to mycorrhizae of neighbouring trees. A. muscaria seems the
principal mycorrhizal fungus of this genus, and was shown by
MoDESs (1941) to form mycorrhizae with pine and spruce. No less
than 17 species of Lactarius are said to be mycorrhiza-formers, and of
these L. delicosus and L. rufus are the chief, being confirmed by
synthetic experiment. All of these reports come from Europe, except
for three citations by Hatch (1937) for American material. Amer-
icans have listed three species of Clitocybe as mycorrhiza-formers,
but MoDESS, in synthesis experiment, reports none of the six species
he investigated as forming mycorrhizae. A considerable number of
species of Cortinarius are said to be mycorrhizal but detailed studies
are lacking in almost all cases ; and the same may be said for the 18
species of Russula that are alleged to form mycorrhizae. Tricholoma
has fared better, especially at the hands of Melin and Modess; but
the latter reports 4 species of the genus that failed to form synthetic
mycorrhizae.

Some special cases among the Hymenomycetes may be cited.
The polypore Strohilomyces strobilaceus, a widely distributed
woodland species, was stated by Peyronel to be connected with
Coryliis Avellana. The Hydnums and most polypores one thinks of as
bracket fungi on wood, but Masui (1927) states that H. affine "was
determined" as a mycorrhiza-former with Pinus densiflora; Poly-
ucomelas was mycorrhizal also on this pine; while Long (1913)
stated that Polyporus Berkeleyi had been found on larch in Montana,
the fungus securing food from the forest humus, — which may or
may not have meant that the species was mycorrhizal. Amongst the
agarics one would suppose that Lepiota would surely be a mycorrhiza-
former, but MoDESS (1939) obtained uniformly negative tests in
attempting synthesis with species of this genus. Amanitopsis vag-
inata is mycorrhizal in Europe according to Peyronel and Modess.
The species is common also in America but is not reported mycorrhi-
zal. On the other hand, Cautharellus cibarius, which also occurs
both in Europe and America, is reported mycorrhizal only by Amer-
ican workers (Doak, 1934; Thomas, 1941.) Hygrophorus, having a
viscid cap, includes H. virgineus, which is mycorrhizal on spruce
(Frank, 1888), and H. Bresadolae and H. lucorum, on larch
(Peyronel, 1922). Omphalia, which we think of as tiny fungi of
damp leaf-mold, is mycorrhizal on Nothofagus in New Zealand ; and
the Fairy-ring fungus, Marasmius oreades, is mycorrhizal with Pinus

Lecture III — 43 — Fungal Endophytet,

ponderosa (Birch, 1937). Armillaria is mycorrhizal only in Japan,
so far as records go.

Whatever spore-colour may, or may not, have to do with it,
the great majority of mycorrhiza-forming agarics are white-spored,
Cortinarius being the only important exception.

Gasteromycetous Mycorrhizal Fungi: — The record for the
Gasteromycetes is much shorter. For Lycoperdon, McArdle (1932)
stated that L. gemmatmn formed mycorrhizae in synthesis vv^ith Pinus
Sfrobus and Picea nigra, and he implicates L. pulcherrimum also in
mycorrhiza-formation. Birch (1937) found L. /'^r/a^i«w mycorrhizal
on P. laricio; but Modess (1939, 1941) said that this fungus failed
to enter into synthesis; also L. pyriforme. Similarly, McArdle re-
garded Calvatia saccata as mycorrhizal, but Modess says that this
species did not enter into synthesis. Again, Noack (1889) implicates
Geaster fimbriatus and G. fornicatns, but Modess says that G.
minimus did not enter into synthesis; and Melin (1925) also failed
to secure synthesis. Scleroderma has a better record since three
species, — aurantium, hovista, and vulgare, — are fully attested as
mycorrhizal, with even Modess (1941) agreeing on the first. In
South Africa, Polysaccum crassipes is mycorrhizal on Eucalyptus,
and shows phagocytosis unusually well (Smith & Pope, 1934).

Phallomycetous Mycorrhizal Fungi : — Only one record appears
for the Phallomycetes, vie. that offered by Barsali (1922): My-
corrhizal-like mycelia on roots of Robinia Pseudo-Acacia were seen
in fruit to be Clathrus cancellatus; and in the same way the fungus
was found in gardens on roots of Phyllostachys bambusoides and
P. nigra.

Form Genera of Mycorrhizal Fungi: — The "form genera" of
mycorrhizal fungi have yet to be considered. These homeless waifs
of mycological taxonomy have been adopted by ardent mycorrhizolo-
gists and given cognomens which do not relate them to any other fungi
but do enable the student to talk about them conveniently. That is,
convenience with some reservations, for, confronted with such scienti-
fic names as Mycelium radicis Walyczvi or Mycelium radicis Didymo-
plexis pallentia, one wonders whether taxonomy may not have reverted
to pre-Linnaean habits. Melin goes still further and speaks of
M.r. abietis, alpha, beta, gamma, etc.

Fusarium: — The form genus Fusarium, established by Link in
1809, is the longest cited form-genus in connection with mycorrhizae.

Kelley — 44 — Mycotrophy

As early as 1847, Reissek was isolating a fungus from the "root" of
Orchis Morio which he assigned to this genus and named F. endor-
rhisinn; while, in 1890, Vuillemin cites a Fusariiim from O. mascula;
in 1900, Bernard, from Ophioglossum vttlgatmn. In 1901, Bernard
said that tuber-formation in the potato is called forth by an endophy-
tic fungus, F. solani. The fungus is now generally distributed in
European soil and potatoes form freely, but at first potatoes grown
from seed did not form tubers until the soil was inoculated with
fungus. The next year Bernard stated that the fungi concerned in
all tuber-formation are Fusarium spp., conidial forms of which are
near to the related genera of Nectria and Hypomyces except that
the fungus of potato is F. solani. But, in 1904, Bernard decided that
the Fusaria often obtained from orchids are not the specific fungi
since they do not cause germination ; and the endophyte, he decided,
as obtained from Cattlyea is a fungus described by Bernard as
"Mucedinee oosporee." The following year he said that, while the
endophyte of Cattlyea has structures similar to those of Oospora,
that from Odontoglossum grande is similar to RJiisoctonia; and to
Rhisoctonia Bernard adhered during the rest of his brief life.

Rhizoctonia : — The sterile fungus, Rhisoctonia, which in one case
at least (Sprau, 1937) is identified with Corticinin, has been much
talked of since the days of Bernard ; indeed, many botanists had the
idea that study of mycorrhizae was largely the study of these fungi.
Most of the fungi isolated from orchids in those days were identified
as species of Rhisoctonia, for example: R. languinosa (Bernard,
1909), R. Goodyerae repentis (Costantin, 1920), etc. More recently
other species have been cited, as R. repens (Knudson^ 1925), R.
mucoroides (Porter, 1942).

Phoma: — The genus Phoma, with conidiospores in pycnia in-
stead of on conidiophores as in the Rhizoctonias, has been cited a
number of times. Ternetz (1907) studied five species assigned to
this genus, which she isolated from native German Ericaceae; while
Rayner (1915) found a fungus in Calluna which she placed in a new
genus, Phyllophoma, since it occurred not alone in the root but
throughout the whole plant. From Vaccinium Oxycoccos, Addoms
(1931) isolated Plwma radicis. But in his study of root fungi of Vac-
cinium, Freisleben (1934), who isolated the mycorrhizal fungi, said
that they were apparently not to be referred tO' the genus Phoma, to
which other authors had assigned the endophytes of the Ericaceae. As
to other plants: P. R. White (1929) separated several fungi from
mycorrhizae of Fragaria and thought that a Phoma was responsible

Lecture III — 45 — Fungal Endophytes

for the mycorrhizae. Auret (1930) found a Phoma sp. in Lunularia
in South Africa; Ridler (1922) in Pellia and (1923) in Lunularia
in England, but was not certain in the latter case that Phoma was
the true endophyte.

Mycelium Radicis : — The older names of Fusarium and Rhizoc-
tonia were supplanted in 1909 by Burgeff's new name of Orcheomy-
ces which he applied to fifteen orchid fungi. The name of "Orcheomy-
ces" is attractive : it is short and expressive, but apparently only
Nobecourt (1923) adopted it; and in 1911 Burgeff had abandoned
the name and adopted Mycelium radicis in its stead. This name
is of more general application but it is awkward, even though abbrevi-
ated to M. r., and it violates the Linnaean principal of binomialism.
Melin adopted the designation for his isolates, M. r. abietis from
spruce and M. r. silvestris from pine. Most of these fungi are basidio-
mycetes but M. r. atrovirens is a phycomycete and a parasite that
forms pseudomycorrhizae (Melin, 1921). Associated with this
fungus may be another distinguished by a mycelium of coarse, lus-
trous, jet-black hyphae that radiate from the mantle of a mycorrhiza,
a fungus which was named M. r. nigrostrigosum by Hatch (1934).
This fungus was apparently figured by Gibelli (1898) and is de-
scribed by Mangin (1899). Bjorkman (1941) found both these
fungi in Sweden, under stands of spruce, pine and birch.

As the designation Mycelium radicis usually (but not always)
refers to Basidiomycetes, so the recently prominent Rhizophagus
refers to Phycomycetes. Butler's (1939) paper on this genus had
already been referred to in an earher paragraph.*

Conclusion: — In conclusion, we may say that there seems to be
an unnecessary emphasis laid on the fungal endophyte. If it were
shown that one fungus is more capable of proteolysis than another
and therefore better able to invade tissues of a plant ; or if another
fungus had a greater supply of diastasic enzyme and was conse-
quently better fitted to be an orchid symbiont ; or if yet another fungus
had rich provision of N cation or phosphorus-complex and was
therefore a richer "booty" for the "mycorrhiza to capture", there
would seem to be some point in the emphasis laid on fungal identifica-
tion. But in all cases it is simply a case of : A occurs on B, or C
occurs with D ; when, as a matter of fact, we know that A and C —
and E and G, for that matter — can all occur in the mycorrhiza of B
at the same time.

*LiHNELL finds that M.r. nigrostrigosum is the same as Cenoccoccum
graniforme (Symbol.bot.UpsaUens. 5(2), 1942).

Kelley — 46 — Mycotrophy

Even though one were to say it is necessary to know fungal
identity to distinguish between beneficial and parasitic species, the
argument breaks down before the realization that, strictly speaking,
there are no mycorrhizal fungi : there is only a mycorrhizal state.
Apparently almost any fungus can be a non-pathogenic symbiont ;
but nature of the symbiosis depends on a complex of physiological
and ecological conditions or influences, and not necessarily upon any
specific fungus, and the non-pathogen under diflferent circumstances
may become a pathogen, or the reverse. Since there is apparently no
specificity in mycorrhizal endophytism, and since no analyses of my-
celia are made to determine specific differences, the identifying of the
mycorrhizal endophytes must be regarded somewhat in the nature of an
hobby. It is important, just as every scientific discovery is important,
but its importance would seem to consist chiefly in allaying our
curiosity as to what fungi can enter into mycorrhizal symbiosis. It
is something like discovery of mountains in the Antarctic, — very
interesting but of no obvious utility.

Lecture IV

FOSSIL MYCORRHIZAE

Limitations of the Fossil Record: — Since 1904, when the first
paper on fossil mycorrhizae appeared, enough information has been
gathered to outline the fossil record of our subject. Yet this record
has grave limitations, imposed not alone by scantiness of the in-
vestigations but by the nature of all palaeobotany. We have become
so accustomed to thinking in the terms of Historical Geology that
ofttimes we forget the "geological time table" was created a century
ago, when knowledge was far more deficient than it is today, and that
later discoveries have been pieced into the Lyellian system, the re-
sultant table being far from convincing. It is of interest to observe
that Historical Geology is one of the few sciences, perhaps the only
science, that has not undergone major revision in the current cen-
tury; and, whereas Newtonian Physics has been supplanted by Ein-
steinian Physics and other sciences have been critically reworked,
Historical Geology continues unrevised. Indeed, no thought of re-
vision seems entertained or desired. When the terms of Historical
Geology are used, therefore, it is simply an act of convenience, as
the writer pointed out in an earlier paper (Kelley, 1939). It can
scarely be conceded that the terms "Carboniferous", etc., have any
definite time value, yet they are convenient terms since they are in
general acceptance and convey some idea at least of the stratum or
strata from which the material is derived.

Sources of Material: — The most hopeful source of material
for fossil mycorrhizae is in the Coal-balls which have been found
and described from Europe and America. Harder fossilizations in
the midst of the coal, they preserve in often intimate detail the
structure of root and contained fungus from an extinct flora. Where
the fungus is present in actual tissues of the host and shows struc-
ture similar to that of living material, we may feel assured that we are
dealing with a mycorrhiza; but, where fungi are found in peat or
otherwise, it is not so clear that they are mycorrhizal.

Fossil Phyconiycetes : — Butler (1939) described "the vesic-
ular-arbuscular or Phycomycetoid endophytes which commonly occur

Kelley — 48 — Mycotrophy

in cultivated and probably other soils, forming mycorrhizal associa-
tions in the roots of many flowering plants and cryptogams, including
prothalli of liverworts and some ferns." After describing these
fungi, he notes the "fossil records" of fungi "of this type". Thus
KiDSTON and Lange found a fungus, Palaeomyces Asteroxyli, very
regularly in inner cortex of Asferoxylon Mackeii and of basal region
of stems having transitional structure between rhizome and stem,
all from Rhynie Chert assigned to Early Devonian. It is not
clear that evidence is afforded of any mycorrhizal structure in this
fossil material, and we may note that Palaeomyces Gordonii is found
on decaying stem of Rhynia major. Butler cites still further the
Protomycitis protogens described by Smith in 1884 from rootlets
of Lepidodendron, assigned to Lower Coal Measures of Yorkshire;
but again we do not know that there is positive evidence for consider-
ing this material mycorrhizal. Seward says that Peronosporites
antiquarius is found in scalariform tracheids of Lepidodendron from
Coal Measures. The supposed reproductive bodies may be oogonia or
sporangia, or merely vesicular enlargements of hyphae. Similar
swellings are seen in cells, probably of cortex of Lepidodendron or
Stigmaria, from the Halifax Coal Measures. Such material would
be questionably assigned to mycorrhizae.

Still more recent material comes from peat bogs, locally known
as "muskegs", in Alberta through Prof. Lewis of Edmonton; but
here again there is no positive evidence of a mycorrhizal nature.
Butler (1939) described this interesting material and decided that
the fungus is the same as the "well-known vesicular-arbuscular
endophyte of modern plants and with the fungus described by
OsBORN and Halket". Again, Rosendahl (1943) reports the same
sort of fungus from three Pleistocene sites in Minnesota and refers
the fungus to the genus Rhisophagiis. The fossils came from a depth
of more than 80 feet in well-borings, and after sand was washed
from the matrix the material was examined. From the excellent
photographs, one would think that he was looking at mould fungi;
and it is stated in the paper that the fossil fungi had grown on moss
leaves and coniferous needles, which are scarcely the organs in which
one would naturally look for mycorrhizal fungi. Incidentally, it may
be mentioned that according to Ellis (1917) there are 15 species of
fossil Phycomycetes known, and of these he mentions Paleomyces
hacilloides as a saprophyte on fossil leaf mould.

Fossil Hepatics: — So far as we are aware, there is no record
of fossil endophytes in hepatics. It would doubtless be a difficult
study of a rare specimen were fossil mycothalli to be described.

Lecture IV — 49 — Fossil Mycorrhizae

Then, again, it must be realized that few students of fossils have any
keen interest in mycorrhizae, and many examples of our science may
languish in slide-boxes as in sarcophagi of our science. When it is
realized that morphologists looked at the Hartig net in roots of woody
plants for many years before realizing that they dealt with anything
more than "curious thickening strips", and even today are inclined to
ignore or at best to tolerate mycorrhizae, it is not to be wondered that
palaeobotanists, who have even less interest, should have succeeded
so well in ignoring them.

Fossil Ferns and their Endophytes: — There are three papers
dealing with fossil mycorrhizal ferns. The first paper, by Seward
(1924), deals with the fern Tempskya from Montana. "Some roots
have lost the x)'lem, and the centre is occupied by a group of dark
brown bodies that may be coprolites of a small insect or, in some
cases, possibly escaped cell contents. Entomologists whom I have
consulted have not been able to identify the oval bodies with the
activities of any known boring animal : no trace of any insect has
been discovered. Attention has elsewhere been called to the resem-
blance of these bodies to the supposed coprolites frequently found in
tissues of Carboniferous plants." Seward figures the same sort of
bodies as those figured by Janse (1897) for Celtis, and the writer
found similar structures in Juglans collected at Mont Alto, Pennsyl-
vania.

In Osmundites Dozvkeri, "The ground tissue cells contain traces
of distinct fungal hyphae, and in many of the parenchymatous ele-
ments the cavity is completely filled with spherical vesicles ; in other
cases one finds hyphae in the center of the cell while vesicles line
the walls. Carruthers refers to these bladders as starch grains, and
this may be their true nature ; their appearance and abundant oc-
currence in the parenchyma certainly suggest vesicular cell-contents
rather than fungal cells. I could detect no proof of any connection
between the hyphae and bladders, and the absence of the latter in
the cavities of the tracheids, favoured the view of their being either
starch grains or other vacuolated contents similar to that in the cells
of the Portland cycad referred to." (Seward, 1898).

The third paper on fossil ferns is by Andrews and Lenz (1943),
who describe a petrified Coenopterid fern stem from the Middle
Pennsylvanian of Illinois, which contains an abundant mycelium in
the cortex. The fern stem, or possibly a rhizome, has been described
by Andrews as Scleroptcris illinoiensis, and the mycelium is found
within host cells throughout the cortex although it is somewhat more
abundant in the middle and inner regions. Hyphae were also found

Kelley — 50 — Mycotrophy

in tracheids of the stele although they do not assume typical mycor-
rhizal form in these cells. Whether or not all this mycelium belongs
to the same fungus cannot be stated positively. Most of the mycelium
appears to be intracellular and typically endotrophic but there is some
evidence that it may be intercellular as well. A considerable number
of host cells contain a very dense aggregation of mycelium, while in
many of the host cells infected in this way the mycelium tends to
assume a nearly spherical form until finally the hyphae lose their
identity as individual strands and in some cells the entire mycelial
body appears as a nearly uniform amber-coloured sphere. This action
may have resulted from a plasmolysis of the entire contents of the
host-cell, or there is a possibility that phagocytosis has occurred. In
a few of the cortical cells there may be noted a number of larger
bodies varying from 15 to 33 /x in diameter, which are considered
tentatively as vesicles.

Mycorrhiza of Fossil Lycopod: — One of the best known in-
stances of fossil mycorrhizae was described by Weiss (1904). A
mycorrhiza or perhaps a mycorrhizome was found in the Lower Coal
Measures, the root not being associated with the plant which bore it ;
but the plant was possibly Lycopodiaceous and was referred to the
form genus Rhisonium of Corda. Hyphae were found in root-hair
and in epidermis but for the most part in the inner cortex, where
hyphal swellings were found. The vesicles are usually empty but
sometimes contain homogenous contents. "The obvious resemblance
between these clumps in the fossil plant and those of recent mycor-
rhiza, together with the close agreement in the structure and behaviour
of the Fungus in the outer layers of the cortex with those of the
Fungus in recent mycorrhiza will, I think, be regarded as sufficient
evidence for the conclusion that we are dealing in the case of this
fossil plant with a mycorrhiza or a mycorrhizome. The Fungus differs
materially in its manifestations from other cases of endotrophic
mycorrhiza so far observed in fossil plants and in no case suggests
that it was living either saprophytically or parasitically upon the host
plant. The excellent preservation of both the Fungus and the host
plant and the specialization of the cortex into two layers comparable
with the Tilzwirtzellen' and 'Verdauungszellen' of recent mycorrhiza
would suggest that, as in the case of the latter, the host plant is deriv-
ing some benefit from presence of the Fungus."

Mycorrhizae in a Seed Fern: — For the Pteridosperms or
Seed Ferns there is one record of possible mycorrhiza by Ellis
(1917). According to Ellis, in the fossilized vegetable remains of

Lecture IV — 51 — Fossil Mycorrhizae

the Lower Coal Measures, it is not unusual to meet with fragments of
fungal threads. Peronosporites gracilis is very widely distributed in
this horizon, the hyphae occurring in cortex of young rootlets of
Lygiuodendron Oldhamium and are not wanting in the stele, in which
both hyphae and vesicles were found. The fungus was probably a
parasite according to Ellis. He says further that vesicles, both
terminal and intercalary, were found, and tuberous swellings. While
in older plants the cortex alone is invaded, in young plants stelar
cells are also infected.

Mycorrhizae in Gordaites: — One tree at least of the Palaeozoic
was provided with mycorrhizae, for these structures in Cordaites
have been described in some detail. Osborn (1909) described the
roots of Amyclon radicans, which has been shown to belong to this
group. It bears such remarkable and irregularly arranged bunches of
lateral roots that Osborn examined them to discover if these bunches
might correspond in any way with the root tubercles of recent plants.
These lateral roots are found to have a thick cortex divisible into
two regions, the inner of which contains dark cells that show evident
fungal hyphae. The fungus occurs in knots of non-septate hyphae
that sometimes bear terminal vesicles but there was no trace of spore
formation. The conclusion was reached that this tree probably in-
habited saline swamps and had bunches of coralline roots such as are
known to occur in many recent plants under similar conditions.
Osborn considered the relation of fungus to Amyelon to be in the
nature of a mutualistic symbiosis.

In another study of Amyelon, Halket (1930) made sections of
a British coal-ball and found in longitudinal sections of rootlets of
Amyelon that they "not only showed the structure of the root-cortex
but also had fungal hyphae present in its cortex, and forming a definite
'fungal zone' round the stele". The description and excellent photo-
micrographs indicate that the structure of those ancient Carboniferous
rootlets was very similar to that of coniferous rootlets of today. Root-
hairs were not as a rule developed. The diarch rootlets, which
branched laterally as a result of division of cells in the pericycle, had
apices which indicated that many of the rootlets had "limited growth".
The (septate) hyphae were mainly intercellular but formed vesicles
and arbuscles intracellularly. The author mentions, and the illustra-
tions would seem definitely to> indicate, digestion stages in cortical
cells; but the vascules were never invaded. Halket considered the
symbiosis to be of mutual benefit.

Kelley — 52 — Mycotrophy

Summary: — There are no records of mycorrhizae in fossil Angio-
sperms because, so far as we are aware, there are no descriptions of
fossil angiospermous root structures. Impressions of aerial organs in
clay beds give us no clue to the subterranean organs ; but since mycor-
rhizae were so well developed in the lower plants, it would perhaps
not be an unwarranted assumption that they occurred in higher plants
also. The general picture of ancient life that the fossil record gives
us is a duplicate of the one we see today : There were forests and on
the forest floor was leaf litter and mould in which saprophytic fungi
lived ; and the rootlets and other subterranean structures of ferns,
lycopods and trees were invaded by fungal hyphae as they are today,
and these hyphae produced swellings and vesicles that give the pre-
pared sections a modern appearance. Then, too, there are "digestion
stages" that indicate phagocytosis occurred in those old mycorrhizae.
Mycotrophism is by no means a new process, for it appears coinciden-
tally with the appearance of rooted plants. The explanation of myco-
trophism on any developmental basis involves serious problems.

"The antiquity of fungi also raises again the question of their
origin, whether they came from the Algae or from one or more
separate and distinct phylogenetic lines. The sum of geological
evidence appears to favor the conclusion that they have been distinct
from the beginning and should not be placed in the same phylum
with the algae." Wolf and Wolf, The Fungi, vol. 2, p. 488, 1947.

Lecture V
DISTRIBUTION OF MYCOTROPHIC PLANTS

General: — It is unknown whether plants in nature have root-
hairs or mycorrhizae,— -or neither; but there is enough evidence at
hand to indicate that mycorrhizae predominate over root-hairs in the
majority of cases. That many plants can produce root-hairs when
grown under artificial conditions of greenhouse or laboratory control
has been amply demonstrated, yet it is also demonstrated that these
same species of plants when in their native haunts may produce mycor-
rhizae. Hence, almost exclusive study of root-hair plants in botanical
classwork is questionably scientific, and some day Botany must revise
its programme; for the attitude of traditionalism that has fastened
itself upon Science is unfortunate. In the future, the Geograph-
ical Distribution of root structures will doubtless be better known;
but at present something of a picture of mycorrhizal distribution may
be gained from incidental references made in various papers. There
are very few researches that deal directly with the subject.

Two general observations may be noted before the geographical
data are presented. First, Costantin & Magrou (1926) thought
that geographical distribution of symbiotic plants depends on distri-
bution of mycorrhizal fungi: Thus, mountain plants rest ephemerally
on the plains because of absence of appropriate fungi. But this idea
is not very well established and awaits further evidence. Second,
WiLKiNs & Patrick (1939) thought that "there is a possibility that
the phanerogamic species may influence fungus distribution." This
idea is perhaps better grounded than the former.

Since more than one half the students of our science have lived
in western Europe, more is naturally known of mycorrhizae in this
region than in the rest of the world.

Germany: — Botanists of Germany, earliest and chief center of
mycorrhizal study, have given us records of a large part of the German
flora. Chief among these studies are those of Schlicht (1888), a
student of Frank. He was led to investigate herbs of his region by
finding mycorrhizae on Rammculus acris and he came to the con-
clusion that mycorrhizae are "distributed over a great range of our
flora." His reports form an almost unique model for in each case he

Kelley — 54 — Mycotrophy

lists the species, its habitat, and locaHty. Thus, he found Lotus corni-
ctilattis in sandy soil at Halensee, Fragaria vesca in forest at Negast
in Pomerania, and Myosxirus minimus in humus-rich soil at Putbus.
It is refreshing to find such precision when so often a paper states in
its title that it recounts "The Occurrence of Mycorrhizae in Pine",
for example, when actually the paper merely tells about a few samples
of one sort of pine collected in an unnamed locality. May all students
of mycorrhizae pay close attention to the place and conditions of
growth of their material !

A somewhat similar list of mycorrhizal and non-mycorrhizal herbs
was published by Hoeveler (1892) that follows Schlicht's
statements rather closely. Many of the species these investigators
listed as non-mycorrhizal are now known to possess phycomycetous
endophytes, but 41 out of 68 investigated species were placed in this
category. About an hundred other German authors tell us of various
other mycorrhizal plants, and sometimes a locality is given, as, trees
and herbs from East Prussia, pine from the Brandenburg Marshes, or
alder from Breslau.

From all these studies, Frank^s early (1888) conclusion seems
justified that most German plants are mycorrhizal. Frank said that
in all the many hundreds of cases of cupulifers examined in forests
throughout Prussia mycorrhizae were never lacking, and he said that
the "Umstand, dass diese Symbiose an den natiirlichen Standorten
eine allgemein verbreitete, iiberall und an jedem Individium constant
auftretende Erscheinung ist, gibt derselben den Charakter einer
Anpassung der Pflanze an die Pilzthatigkeit, wobei diese von der
letzteren einem bestimmten Nutzen zieht." And this conclusion was
emphasized by Stahl (1900) who said that "die Mehrzahl der
hoheren Pflanzen, wenigstens gelegentlich, in diese Symbiose mit
Pilzen eingeht."

France and the Iberian Peninsula: — France, although it stands
second to Germany in number of mycorrhizal students, gives us less
information about the native flora since French students have been
more concerned with the problems of mycotrophy. There are few
citations of locality in French accounts, and no list of French mycor-
rhizal plants. BouDiER in 1876 found Elaphomyces about Mont-
morency; Lecomte in 1887 noted beech, chestnut, oak and hazel
mycorrhizal in the Vosges; Dangeard in 1896 noted poplar, about
Poitiers; Mangin (1910) collected Castanea in woods at St. Cloud
and Viroflay; Boyer (1915) found Trametes connected with tree-
roots at Vallon ; Dufrenoy in 1920 had collected Adenostyles in the
beech woods of the Pyrenees at 3700' A.T. ; Nicolas (1924) collected

Lecture V —55— Distribution

mycothalli of Lunidaria at Toulouse; while Costantin has collected
in the forest at Fontainebleau.

With these records, we may go on to the Iberian Peninsula where
chestnut is found mycorrhizal in Portugal (Camara, 1907) ; but
otherwise we know nothing of mycorrhizal conditions in these lands.
Mendes d'Almeida in 1908 presented a general account of mycor-
rhizae in Portuguese.

British Isles: — Crossing the Channel to the British Isles, we find
little information on mycorrhizal distribution although a considerable
number of papers on mycorrhizae have been published, especially in
England. The earlier papers on the subject were published in The
Phytologist, years 1842-1844, and localities of collection were noted,
as Cotswold Hills (Lees), Lancashire coast at Southport (W. Wil-
son), and Southport, Kent and Sussex (Somerville) ; but this
praiseworthy habit was not continued by later investigators. Rayner
(1911)^ said that her Calluna was common on chalk downs of the
south of England where collections were presumably made; and
Harley (1937) made his collections of beech in the Chiltern Hills.
Only two English papers deal at all with distribution of endophytic
structures: Ridler (1922) cites various localities where Pellia myco-
thalli grow, while Paulson (1923) cites birch from Epping Forest
and in the following year listed certain trees as mycorrhizal in wood-
lands of south-eastern England, viz. Quercus Rohur, Fagiis sylvatica,
Carpimis Betulus, Betula alba, Castanea sativa, Pinus syhestris and
■ Taxus baccata.

For the north-east of Scotland, I. Gordon (1936) cites 16 species
of broad-leaved trees as mycorrhizal and 8 species as having no mycor-
rhizae; but as these eight are oaks, maples, etc., one would suppose
they might be reinvestigated with profit. Strawberry plants in the
Clyde Valley are mycorrhizal according to O'Brien (1928). The one
Welsh paper (Sampson, 1935) deals with Loliiim, the one Irish
paper (Jennings, 1898) with Corallorhiza from the eastern
Alps. The British Isles offer an almost virgin field to the student of
mycorrhizal distribution.

Lowlands and Scandinavia: — Crossing back to the Lowland
countries, we find little information about mycorrhizae. Hesselink
(1924) wrote on mycorrhizae of pines in afforestation of the
Netherlands dunes, and there is a paper on hepatics; but future
studies must tell of mycorrhizal structures in woods of Limburg or
in plantings of The Bosch. In Denmark, pine is mycorrhizal in the
brush-lands of Jutland (P. E. Muller, 1902) ; and so is the im-

Kelley — 56 — Mycotrophy

portant tree, Alntis (Bornebusch, 1914). Certain fungi are always
associated with certain trees in Denmark, according to Lange (1923) :
thus, Amanita muscaria grows under conifers, also under birch, but
never under beech. At Oslo in Norway, Horn (1933) found fairy-
rings formed by Heheloma about the bases of ten young trees of
Betula lenta, and examination proved the tree roots to be abundantly
mycorrhizal. This fungus is generally found in Norway to the limits
of birch distribution. Birch, aspen, and conifers are the chief trees
of Scandinavia and naturally are most studied by mycorrhizal investi-
gators of these countries. The studies of Melin on pine, spruce,
larch, aspen, and birch are justly well known; and the ecological
studies of Romell involving Swedish trees and their fungi. Lihnell
(1939) made an extended study of the mycorrhizae of Juniperus
communis; Lindquist (1939) made cultural studies of spruce.
Hammarlund (1923) studied the association of Boletus with Larix.
An elaborate study of root development in Betula was made by
Laitakari (1934) with ecological emphasis on soils, mycorrhizae
being most plentifully developed on moorland soils and least on sandy
soils. In 1920 Thesleff presented a study of Basidiomycetes of
Finland ; and that about completes our knowledge of mycorrhizal
distribution in Scandinavian countries. Since beech forest finds its
northernmost limit in Sweden, it would form an interesting study to
investigate the woodlands of Skane and compare the mycorrhizal
structures with those, let us say, of French woodlands.

Baltic and Russian States: — Of the small Baltic States we
know nothing of their mycorrhizae ; but Voss & Ziegenspeck (1929)
have made valuable studies of ericads and other native plants of East
Prussia, about Konigsberg. They conclude that the xeromorphy of
these moorland plants is due to mycotrophy. Arcularius (1928)
studied nodules of Hippophae collected from the Baltic region, while
Endrigkeit reported on Allium, Molinia and several trees from E.
Prussia. One of the earliest students of mycorrhizae in Poland,
Bonicke (1910), found that several members each of three families,
Ophioglossaceae, Orchidaceae and Pyrolaceae are endotrophically
mycorrhizal and that germination stages and cell structures may be
used as distinguishing characters. The hepatic Haplomitrium is
mycothallic according to Lilienfeld (1911). A number of exotic
conifers in Poland are mycorrhizal (Dominik, 1937), and native
members of Viola (Zabloca, 1936).

In Russia, in the Gov. Cherson among dry arid sand vegetation,
fungal nodules were found on the herb Tribulus terrestris (Issat-
CHENKO, 1913) ; while in the Gov. Ekatinerinoslaw it was thought

Lecture V — 57 — Distribution

that oak seedlings had failed because of destruction of mycorrhizae in
a very wet summer (Nadson, 1908). Ganeshin (1923) found my-
corrhizal connection between pine and larch, and Boletus luteus and
B. elegans. That is all we can say for mycorrhizal plants in the vast
extent of the Soviet Union.*

The Arctic: — Looking northward to the Arctic, one learns that
perennial plants which inhabit these frigid areas are likewise mycor-
rhizal. In the one paper for the Arctic region, by Hesselman (1900),
there are described plants collected on the Swedish Nathorst Expedi-
tion, and we learn that Arctic species of Salix are constantly mycor-
rhizal while the herbaceous Polygonmn viviparum is thoroughly in-
fected in both its bulbils and countless adventive roots. For the
Antarctic, Johow (1889) observed that Arachnites from Antarctic
South America is the only humus plant known from polar lands.

The Alps : — Coralloid mycorrhizae were described by Hesselman
for Dryas octopetala both in arctic and alpine situations ; and this
description was confirmed by Colla (1931) for the Alps at the
laboratory of La Linnaea. As early as 1888, Ebermayer had
observed roots of spruce, fir and beech only in the humus layer of
forests in the Bavarian Alps; while Stahl (1900) noted Populus
tremtila as mycorrhizal in alpine as well as in lowland situations, and
he included a section of a couple of pages on alpine mycorrhizae.
Tubeuf (1903) observed that Pinus Cembra lives with root fungi
in alpine humus. In the Vanoise, Costantin & Magrou (1926)
found structures like those reported by Hesselman for the Arctic.
They say that Salix in Savoy has a structure identical with that in the
Arctic; and from several studies they derive the generalization that
mycorrhizal symbiosis is found not only in a single species in all
stations of its range, but in numerous species of a genus or even
genera of a family (as the Ericaceae) disseminated throughout the
vast domain of arctic and alpine regions. They conclude that mycor-
rhizae play "a great role in alpine flora as well as in the arctic", and
they list both ecto- and endotrophic forms. Peyronel (1937) also
generalizes about distribution of alpine mycorrhizae, having studied
them on the Italian side of the Alps, and at Kleinen St. Bernhard.
He regarded endotrophic mycorrhizae as universally distributed in
the alpine plant world and believed that members of a plant associa-
tion are most closely bound to each other through symbiosis with a
common mycorrhizal fungus. Malan (1938) worked with legumes
in the Alps and his "results showed that in all the Leguminosae

♦There is an article on a bolete of Russia, as a mycorrhizal fungus, by
Vasiklov. Sovetsk. Bot. 1944(2) :21-27. 1944.

Kelley — 58 — Mycotrophy

studied endotrophic mycorrhiza with hyphae of the Phycomycetoid
type . . . predominated", as a Review stated ; or, as the original has
it: "In tutte le leguminose studiate prevalgno micorize ectotrofiche
con ife de tipo ficomicetoide". Orchids have also been observed in the
Alps : Beau ( 1920) found that in a grotto of the Maritime Alps the
orchids CephaJanthera and Epipactis alone of green plants penetrated
to depths of the grotto, being able to grow in subdued light by the
aid of symbiotic fungi. Jennings (1898) studied Corallorhiza in
the eastern Alps.

Central Europe: — In Bohemia, Ncmec wrote of mycothallic
hepatics ; Peklo, of various mycorrhizae ; and more latterly Klecka
& VuKOLOV (1935), of numerous congeries of trees and shrubs.
Detailed investigations of one hundred eleven woody species were
made in which mycorrhizae occur as constant phenomena independent
of soil properties, and "it follows that mycorrhizae are a generally
distributed phenomenon in woody plants." The species studied com-
prised most if not all the woody plants of Central Europe and a num-
ber of exotics such as Cedriis atlantica, Thuja occidenfalis, and Cor-
nus florida. It is one of the best modern studies extant. The same
authors (1937) studied salt-marsh plants collected from saline soil
about Neusiedler See and from Auschitz and Louny in Bohemia. The
roots of Suaeda maritima, Salicornia herhacea, Plantago maritima,
and six other species showed mycorrhizae which were identical in
structure with endotrophic mycorrhizae found by the authors in
woody plants. These observations coincide with those of Mason
( 1928) except for Salicornia, which was not mycorrhizal in England.
Another Bohemian study, by Smotlacha (1911), indicates that
certain boletes are confined to the neighbourhood of certain trees,
as B. riifus with aspen, and he infers that mycorrhizae are oftentimes
formed on a certain tree only by a certain fungus.

In an early Austrian paper, Henschel (1887) wished to upset
any idea of a beneficial symbiosis and he stated very positively that
presence of mycorrhizal fungi is "absolutely injurious" to spruce.
Another Austrian paper deals with endotrophic mycorrhizae of
Asdepiadaceae (Busich, 1913), an unusual group for mycor-
rhizal study, but as the material came from a botanical garden it tells
us nothing of Austrian plants except that A. syriaca is not mycor-
rhizal. In lower Austria, Pyrola is endotrophic and its mycorrhizal
association is obligatory (Furth, 1920). In Hungary, Bernatsky
(1900) wrote on exotics and philosophized on mycotrophy.

The Balkans: — For the Balkans, we learn that Daphne is mycor-
rhizal in the land of the Croats, at the northernmost edge of the Balkan

Lecture V — 59 — Distribution

peninsula; and Skoric (1925) comments on the curious fact that
both ecto- and endotrophic forms should be found in the same genus.
From nearby Istria were collected the mycorrhizomes of the orchid
Centrosis, used by Arcularius (1928) in his studies. The shrub
Forsythia, which is native to the Balkans, is known to be mycor-
rhizal.

Italy: — For the Mediterranean region there are papers only from
Italy, except for those by Rivett (1924) and by Dufrenoy (1917)
on Arbutus. The Italian papers deal almost exclusively with northern
Italy and leave the maqui vegetation of southern Italy for future
study. Since mycorrhizae occur in chaparral of California (Cooper,
1922), a similar plant formation, it is to be presumed that they may
occur also in the maqui. A considerable number of wild and culti-
vated plants of northern Italy have been investigated, particularly
by Peyronel, who tells of the general localities of his collections, as
the Val Germanasca and the Valli Valdesi in Piedmont, forests about
Pisa, etc. Peyronel (1922&) concludes: 'L'estinza di micorize in
un grandissimo numero, verosimilimente la maggiore parte, della
pliante vascolari e un fatto accertato da tempo della osservazioni di
numerosi riceratori."

One paper, by Ruggieri (1937), records mycorrhizae for almond
in the province of Syracuse in Sicily; and Reed & Fremont (1935)
say that citrus is mycorrhizal in this island. For the future, we may
expect studies of root structures in scrub vegetation of Mediterranean
shores, a comparison of those of the desert flora of the Mediterranean
area with those in America, and studies made in the numerous islands
and in the Balkans where forests still await students of our science.
Several papers have appeared in recent years from the University of
Pavia, notably by Ciferri and by Elisei.

Africa: — Crossing to African shores, we find endophytes in
Morocco. Emberger (1924) tells of hepatics collected in this land,
and Miege (1936) has a paper on potato. As for the Atlas Moun-
tains and their Cedrus forests, we know nothing of possible mycor-
rhizae, nor do we know anything of the alpine flora of Africa. Ac-
cording to Stefansson, there is probably more permanent snow in
equatorial Africa than in all of the Arctic lowlands, and it will be
interesting to learn what effect it has on vegetation, in comparing
root-structures of the Arctic and the Alps with those of the Kili-
manjaro and the Ruwenzori Ranges. As for the rest of equatorial
Africa, we are in entire ignorance for no botanical Livingstone has
invaded dripping forest of the lowlands nor arid plateaus to learn for
us what the root structures may be. Yet there are some notes pre-

Kelley — 60 — Mycotrophy

served by Rayner (1938) who brought together data from several
African forestry stations on growth of gymnosperms with or without
soil inocula, — notes from Taganyika, Nyasaland, and Rhodesia. In
South Africa, Auret (1930) wrote on the hepatic Lunularia, and
Smith & Pope (1934) on the exotic Eucalyptus. For Madagascar
there is a paper by Heim (1937), who says that clove trees in the east
of Madagascar and on the island of St. Mary possess a Pythium-like.
mycorrhizal fungus. Three papers come from Egypt, two of them
being on cotton while the third deals with several garden plants, —
all mycorrhizal. Africa offers a great opportunity for original work
in this field.

Asia: — Continuing with Asia, we find that the forests of this
greatest of all continents, whether tropical or temperate, are unex-
plored by students of our science : nothing is known of possible
mycorrhizae on the high plateaus or in the vast taiga, in arctic lands
or in the high mountains. All the reports that come to us from the
mainland of Asia are from India except that Reed (1935) says Citrus
is mycorrhizal in Malaya. From India come two papers on Casuarina,
which is of course not a native; from Tocklai in N. E. India Tun-
stall (1925) reports on tea mycorrhizae; while Chaudhuri (1925-
35) tells of the hepatics. Butler (1939) found phycomycetoid in-
fection in a number of cultivated plants in Indian plantations. And
it may be noted that Litchi chinensis of China was found possessing
short roots and intracellular infection when imported into the U.S.A.
(Coville, 1921).

So much for the continent. In Ceylon, mycorrhizae are also
found on tea roots (Park, 1928) ; while Parsons (1938) gives us
notes on orchid cultivation and orchid rhizoctonial fungi from the
island. For Sumatra there is a paper by Palm (1930) who said
that a Boletus, probably B. palUdus, was observed to grow in associa-
tion with Pimts Merkusii in forests of Sumatra where ground vegeta-
tion was sparse and needle litter deep and compact. From Borneo
a paper by Posthumus (1937) tells us that Legitmiuosae, often in
symbiosis with bacteria, are frequent in the dry savannas of the
Padang Loewai in E. Borneo, taking the place of mycorrhizae of
acid soils.

Java: — It is Java, however, that is the principal seat of mycor-
rhizal study in these great islands, for in Java are found the Buiten-
zorg Botanical Gardens where some of the best known students of
our science have worked. Chief of all was Janse (1897), whose
classic paper records presence of endophytes in selected cases through-

Lecture V — 61 — Distribution

out the whole range of that tropical flora. In the course of a study
of parasites of the coffee-tree, Janse's attention was drawn to fungi
on roots, and from that beginning he was led on to make an extensive
study of roots of tropical plants. It seemed preferable to study plants
from native haunts and hence almost all material was taken from the
forest at Tjibodas, which belongs to the Botanical Garden and is
situated on the flanks of the Gedeh volcano at an altitude of 1400-1800
m.A.T. The flora of E. Java is of an extraordinary richness and, as
it was impossible to study the roots of all the plants, he decided to
omit ectotrophic sorts entirely and to devote his attention to the
endotrophic, mostly of large forest trees. In general, only a single
species of each family represented at Tjibodas was studied ; and in this
logical way Janse built up his excellent study, which nevertheless is
only a preliminary one. He summarized his results in a graphic table
which is here reproduced, showing the numbers of plants studied in
each taxonomic group, with and without mycorrhizae :

Tabular summary of endotrophic mycorrhizae in some Javanese plants: —

Trees

H]

JRBS

Total

Plus

M

jnus

Plus

Minus

Plus

Minus

Cryptogams
Gyninosperms
Monocotyledons
Dicotyledons

1

5

2

38

0
0
0
0

5

0

12

6

2
0
3

1

6
5

14
44

2
0
3

1

Total 46 0 23 6 69 6

Several years before Janse published his paper, Goebel had
written on hepatics (1891) and Lycopodimn (1888) at Tjibodas and
their fungal infection. Miehe (1911) had called attention to vegeta-
tion on volcanic soil in Java, the pioneer plants being provided with
root symbionts. He suggested that there is a significant relation
between occurrence of these plants and soil conditions, naming Casiia-
rina, Myrica, Alhizsia, and two ericads as particularly involved.
Faber (1925) confirms these suggestions, stating that all the investi-
gated solfatara plants are mycorrhizal, the root symbiosis apparently
serving for N assimilation since the soil is very poor in N. He notes
among these plants two groups, one xeromorphic as the ericads, and
the other more nearly hygromorphic.

Some special studies of Java plants are to be noted : Treub (1885)
reported a Pythium in roots of sugar cane; Figdor (1897) on the
gentianaceous Cotylauthera tenuis; Campbell (1907) on Ophioglos-
siim; Steinmann (1929) on mycorrhizae of Cinchona which, he
said, is the first report for this tree; Pijl (1934) on mycorrhizae of
Burmannia and Epirrhisanthes.

Kelley — 62 — Mycotrophy

From the Dutch East Indies in general come these reports : Casua-
rina equisetifolia growing on coral islands of the bay of Batavia
possesses root nodules like those of legumes (Kamerling, 1911). A
paper on non-symbiotic germination of orchids by La Garde (1939).
Hevea rubber trees in the D.E.I, are endotrophically mycorrhizal
(d'Angremond, 1939). A method of mycorrhizal staining by
Frahm-Leliveld (1941).

Japan: — This country has produced a third of the students of our
science in the Orient. Earliest among them was Kusano (1911),
whose study of the orchid Gastrodia attracted much attention; while
later Hamada (1939) studied Galeola from the Kyoto district, these
being the two orchids studied in Japan. Nodulous plants have attracted
more attention, Coriaria having been studied by Katakoa (1930) and
Shibata (1917); Podocarpus by Kondo (1931), Mimura (1933),
and Shibata I.e.; Alnus by Masui (1926) and previously by Shi-
bata (1902) ; Myrica by Shibata I.e. Conifers, another of the my-
corrhizal favourites, attracted Masui (1926), Shimizu (1930), and
Tazoye (1940), the last describing rootlets of coniferous seedlings
but saying nothing of mycorrhizae. Nakai (1933) wrote on the fern,
Cheiropleuria. Two papers only give insight into geographical distri-
bution of mycorrhizal plants in Japan.

Takamatsu (1930) wrote on the solfatara plants in the region
of Hakkoda, studying all the 28 species that existed there, a number
limited by the high acidity of such soils; and he found 6 of the 28
fungus-free. These six were grasses and sedges, hydrangea and
Aletris, and Pteridiiim which is elsewhere known to be mycorrhizal.
The mycorrhizal species are Piniis, Betula, Salix, various ericads with
other shrubs, and some herbs. Asai (1934) presented a well-organ-
ized paper in which he reported presence or absence of mycorrhizae
from many plants in various habitats. He stated that mycorrhizae are
absent from Polygonaceae, Centrospermae and close relatives : ecto-
trophic mycorrhizae are limited to a few families and most mycor-
rhizae are endotrophic. Collections were made in a tropical island,
an alpine mountain of Japan, on the seacoast, fields and cultivated soil.
Many grasses were found to be mycorrhizal, and a Drosera. Specific
localities, as given by Schlicht, are not given by Asai ; but he does
give a good idea of root structures of a considerable cross-section of
the Japanese flora.

New Zealand: — The first New Zealand paper (Cavers, 1903)
on fungal symbionts was appropriately on a liverwort, Monoclea
Forsteri, described from these pleasant islands of the South Pacific;

Lecture V — 63 — Distribution

while succeeding papers are in a chance taxonomic order. Holloway
(1920) wrote on fungal symbiosis in epiphytic prothallia of several
New Zealand lycopodia, describing the infected thalli in some detail.
Next may be noted some remarks attributed by Prof. Cockayne
(1923) to Prof. E. H. Wilson, who thought that slow growth of
pine in garden soil was due to climate rather than to any lack of
microorganisms. But Walker (1931) presented a study of the
mycorrhizae of Pinus radiata, "one of the chief exotic timber trees
of New Zealand", collections of which were made in various localities
in the Nelson and Canterbury districts. Miss Walker stated that
no difficulty in establishing this pine had been experienced in N.Z.
McKee (1941) gives us a paper on growth of spruce at Conical Hill,
a mycorrhizal explanation. And from Birch (1937) came a paper
on "forest fungi of significance in New Zealand" which records my-
corrhizal symbiosis proved or suspected in several exotic pines, in
Betula alba and Nothofagus Solanderi. Neill (1940) wrote on
endophytic infection of Lolinm, which however was not mycorrhizal
since the infection was confined to the leaves. Invasion of roots was
found in field grown plants but the hyphae differed from those of
the leaves.

Later, Neill (1944) recorded mycorrhizae caused by Rhiso-
phagus in virtually all vascular components of the New Zealand
flora except in exotic pines, where it has not "been identified with
certainty."

Australia: — This country has attacked the mycorrhizal problem
especially from an economic standpoint. McLennan has made in-
tensive studies of Lolinm while Young has studied exotic conifers,
the former in Victoria and the latter in Queensland. From Queens-
land comes also a note by Simmonds (1936) which records mycor-
rhizae on exotic pines and the rapid infection following acidification
of the soil with sulphur. The first published mention of mycorrhiza
in Queensland is said to refer tO' Finns taeda and to date from 1928.
New South Wales gives us McLuckie and a series of papers on my-
corrhizae.— of Dipodinm, an orchid which grows under Eucalyptus;
Gastrodia, another orchid ; of Macrozamla, Podocarpus, Casuarina,
and Eriostemon, — in which last study Alan Burges participated.
Victoria provides, in addition to the work of McLennan, a paper by
Coleman (1936) on Sarcosiphon, a rare Thismiaceous plant asso-
ciated with hazel. In the adjacent island of Tasmania, Saxton (1930)
studied Pherosphaera, one of the Podocarpineae. At Penola, in South
Australia, Samuel (1926) found an oat disease associated with
typical endotrophic mycorrhizae and this discovery led him to a

Kelley — 64 — Mycotrophy

further study : he found 27 species of legumes infected in the same
way as Jones (1924) had described; roots of other crops and fodder
plants, weeds and native plants were examined and a large majority
were found to be infected to some extent with fungi. Piniis insignis
and Eucalyptus rubida had ectotrophic mycorrhizae. All species of
legumes (27) and grasses (30) were found to be mycorrhizal. Like-
wise in South Australia, Eardley (1932) described mycorrhizae of
P. radiata. In Western Australia, according to Kessell (1938), "it
appears to be part of the standard practice to inoculate nurseries . . .
with the appropriate fungi, thus obtaining normal growth of the tree
plants; the infected plants when put in the forest are said to infect
the soil quite satisfactorily."

Besides these papers which give us information on mycorrhizae
by states, there are several general Australian papers. Three papers
tell us about mycorrhizae of pine : Cromer (1935) on those of planted
P. radiata; Burbidge (1936) on root development of P. pinaster and
the seasonal variation of its mycorrhizae; Ludbrook (1940) on a
correlation between mycorrhizae and boron deficiency in plantation
soils. PiTTMAN (1929) described mycorrhizomes in the orchid
Rhizanthella, and listed "mycorrhizae" for about a dozen other sorts.
Fraser (1931) presented an unusual study on the genus Lobelia, of
which two species are said to maintain an obligate relationship with
mycorrhizal fungi.

Thus, in the splendid series of Australian papers much is given in
regard to mycorrhizae of exotics, and something of the native flora;
and there are some excellent detailed studies. Yet the student of my-
corrhizal distribution finds almost a blank page for Australian native
flora, for there are no records for the lower plants and a very limited
representation of the higher plants. Indeed, practically all of the
Australian flora is yet to be investigated for occurrence of mycor-
rhizae ; and the same may be said for that of Africa, Asia, South
America and North America apart from the U.S.A. The Hawaiian
and other Pacific islands are yet to be studied for root-structures, ex-
cept that for the Philippines one paper is reported (Hatch, 1937).

South America: — Several papers treat of South American mycor-
rhizae : They were described from several Brazilian species of
SciapJiila by Poulsen (1886) ; while Macfarlane (1897) described
a mycorrhiza from Philesia, a liliaceous plant of western Patagonia.
Mycorrhizae were recorded for Citrus by Milanez (1940), for the
first time for South America, it is said. Marchionatto (1940) has
a preliminary note on the endophyte of Lolium in Chile; while from
Chile also' came the Solanum Maglia used by Bernard (1911). Cacao

Lecture V — 65 — Distribution

is mycorrhizal in Venezuela (Laycock, 1945). In addition, a paper
by Berggren (1887) treated of austral conifers, including Arancaria,
but no localities were given. An article has been published recently
by Ca STELLA NOs On nodules of alder in mountains of Argentina
(Lilloa 10:413-416, 1944). To sum up the matter, there is not yet
a single paper devoted to mycorrhizal distribution in South America.

West Indies and Central America: — For the West Indian
islands four studies may be cited: Johow (1885) published a paper
on West Indian saprophytes belonging chiefly to the genera Burman-
nia and Apteria. Mycorrhizae of sugar-cane in San Domingo were
studied by Ciferri (1928), who has also published on mycorrhizae
of the Burmanniaceae (1946). A fuller description of cacao mycor-
rhizae is given by Laycock & Dale (1945). A brief note by Palm
(1930) on pine in Guatemala is all that can be said for mycorrhizal
distribution in Central America. With botanical facilities available in
the Panama Canal Zone and in Puerto Rico and with the example of
the Buitenzorg Gardens before them, it would seem that the Ameri-
cans might match the splendid contributions from the Dutch East
Indies with some good studies of mycorrhizae in the American tropics.

North America: — In North America there are no mycorrhizal
studies whatever to report from Mexico or from Alaska, while from
Canada come three papers, — a neat study of Taxus from Quebec
(Prat, 1934) ; a short note by Lewis (1924) on Picca of Alberta;
and a citation of raspberry (Rtibus) by Berkeley (1936). In other
words. North America, apart from the U.S.A. is yet to be explored
for root structures of plants.

North-eastern U.S.A.: — There is not very much known of
mycorrhizal distribution in the U.S.A. New England, oldest center
of learning in the country, has told us nothing of the subject, except
that Ames (1921 et seq.) described "mycorrhizae" for some orchids
while Stokey (1924) reported fungal infection of Lycopodium
prothallia in western Massachusetts. Epigaea from Connecticut pro-
vided Barrows (1936) with material for her studies. For the Middle
Atlantic States there are two papers by Henry that tell us of the
Wading River region of Long Island (1934) and of Butler County
in western Pennsylvania (1933). For Long Island he lists all the
pines and junipers, birch, chestnut, oaks, maples and ericads, showing
that a cross-section of a pine-barren area exhibits all the woody plants
as mycorrhizal. In western Pennsylvania, in a deciduous forest area,
he reported 60 species of woody plants as mycorrhizal, — a large

Relley — 66 — Mycotrophy

proportion of the native flora. Myrica carolinensis from the coastal
area of New Jersey was described as mycorrhizal by Harshberger
(1903), In an unpublished paper prepared in 1930, Kelley listed
as mycorrhizal 160 out of 172 spp. of woody plants investigated in
the Middle Atlantic States.

Southern U.S.A.: — There is only one paper on mycorrhizal
distribution in the Southern States of the American Union, a paper
by McDouGALL (1928) on the 16 spp. which he observed in the
Great Smoky Mountains of North Carolina and eastern Tennessee.
Three of these species he considered non-mycorrhizal, namely Leio-
phyllum and Rhododendron which are ericaceous, and Sassafras, which
Kelley finds to be mycorrhizal. His other species are conifers,
cupulifers, magnolias and hickory. Besides this paper there are
isolated observations: Pine is mycorrhizal in North Carolina (Ashe,
1915; Cobbe, 1916) ; while southern pine (presumably P. palustris)
is termed mycorrhizal by Pessin (1939) and Huberman (1940),
both authors saying that the mycorrhizae are abundant. Pessin in 1928
reported 4 spp. of pine mycorrhizal at Bogalusa, Mississippi, and
mycorrhizae abundant on seedlings grown at McNeill, in the same
state. The orchid Tipularia collected in South Carolina contains an
endophyte (Clifford, 1899) ; the valuable pecan (Hicoria) bears
mycorrhizae in Georgia (Woodroof, 1933) ; the exotic Casiiarina
is mycorrhizal in Florida (Mowry, 1933) ; while the orchid Zeuxine
strateumatica of S.E. Asia is now established in peninsular Florida
(Porter, 1942), Atkinson (1892) noticed galls on Ceanothus
collected in Alabama.

Central U.S.A.: — Turning next to the mid-portion of the U.S.A.,
active centers of mycorrhizal study are found. In Indiana, Doak
(1927) presented a list of 21 mycorrhiza-bearing species of plants
collected about Lafayette, mostly trees but with a few herbs, including
the fern, Adiantmn. In lUinois, Lessman (1928) hsted "a new form
of ectotrophic mycorrhiza" on Quercus hicolor. At Urbana, Illinois,
McDougall & Liebtag (1928) examined 145 of the 183 spp. occur-
ring in the university woods and found that mycorrhizal fungi oc-
curred on roots of 93 spp. Pfeiffer (1914) found Thisniia with
endophyte on prairie near Chicago. For Michigan, Duthie (1908)
presented a list of mycorrhizal tree species but the list cannot be fol-
lowed since no scientific names are given. A detailed study of ten
species of native plants growing in bogs of the Huron River Valley
led Transeau (1906) to consider their "mycorrhiza" to be detri-
mental, but as they were bog plants perhaps the structures were

Lecture V — 67 — Distribution

actually pseudomycorrhizae. Prothallia of the fern Botrychium vir-
ginianum collected in Grosse Isle contained an endophyte according
to Jeffrey (1898). Two papers on connection of sporophores to
tree roots come from Michigan (Kauffman, 1906; Pennington,
1908), Kauffman noting mycorrhizae on oak, sugar maple and
Celastrus at Ann Arbor. For cultivated plants, A. H. Smith (1930)
described endotrophic mycorrhizae for various fruit trees about Ann
Arbor. Freeman (1904) made studies of Loliiim at the University of
Minnesota. In an unpublished paper, Kelley described root-endings
for woody plants of the Kawishiwi Ranger District of the Superior
National Forest (northern Minnesota), all the species proving my-
corrhizal. In Iowa, Lohman (1927) made a valuable study of the
"occurrence and nature of mycorrhizae in Iowa forest plants", collec-
tions having been made in central and northwestern Iowa. Seventy
mdividual plants were studied (16 being ferns) in 40 species of which
20 had a constant root endophyte, 5 an occasional, while 15 were
fungus-free. For Missouri, there is a paper from a forest nursery
where Miller (1938) studied influence of mycorrhizae on growth
of short-leaf pine seedlings (presumably P. echinata). From St.
Louis come several papers on exotic orchids.

Rocky Mountains: — In the Rocky Mountains, mycorrhizae are
found on 8 spp. of trees and 3 spp. of Cercocarpiis (McDougall &
Jacobs, 1927) ; and ectotrophic mycorrhizae on certain trees of the
Uinta Basin of Utah are described by Henry (1936), who also re-
corded absence of mycorrhizae from 7 spp. A survey of northern
Colorado flora bearing mycorrhizae was made by Thomas (1943),
listing 21 families.

Pacific Coast: — On the Pacific Coast, studies come only from
California except that there is one paper from Oregon that deals with
nursery trees. The three principal species of the scrub vegetation or
chaparral of Jasper Ridge, vis. Adenostoma, Quercus and Arcto-
sfaphylos are mycorrhizal in sand and clay as well as in humus
(Cooper, 1922). Sarcodes was collected at San Bernardino by
Oliver (1890), and presumably MacDougal's (1900) collections
were Calif ornian since the plant occurs nowhere else. More lately
(1944), MacDougal & Dufrenoy report on Aplectrum, Coral-
lorhiza and Pinus Torreyana. Smaller roots of celery were heavily
infected with fungus in delta peat soil (Rawlins, 1925) ; while Reed
& Fremont (1934) found mycorrhizae generally present on Citrus
in California.

LECTURE VI

MYCOTROPHIC PLANTS AND THEIR
ENVIRONMENT

Soil as a Mycotrophic Habitat: — There is perhaps no satis-
factory definition of soil. If soil is defined as the "unconsolidated
upper few feet of the earth's crust", then some mycorrhizae do not
occur in soil at all, for one may find them in pockets of humus formed
by decaying stubs on trunks of living trees. Such occurrences are not
uncommon with Acer ruhriim growing in swamps of the eastern
U.S.A. where the living tree will develop "necklace-beaded" mycor-
rhizae in pockets of humus on its own trunk from a root-branch de-
veloped by the trunk. Or, seedlings of other species, which have been
termed "pseudoepiphytes", may develop in such situations. Cordemoy
(1904) found that aerial roots of Vanilla form mycorrhizae in rotting
supports that are supposed to hold the plant up ; and the same thing
may be seen even better with pepper vines. Again, in a forest one may
find dozens of spruce or Tsuga seedlings growing on a partly decayed
log or stump ; and occasionally one finds a sapling that started in such
a situation and later extended its roots down into the soil, the stump
or log meanwhile rotting, leaving the sapling standing as it were on
stilt roots. All of these examples show that plants can grow for some
years in a flourishing condition without any contact with mineral
soil. If it is necessary for trees to have mycorrhizae in order to gain
inorganic salts from the soil, they must win the salts vicariously
for they have no direct contact with the soil, yet the seedlings flourish.

In contrast, there are plants growing and producing mycorrhizae
in pure mineral soil which, as in Holler's (1902) sand, showed no
trace of humus. Or, mycorrhizal plants are found in agricultural soil
where humus and mineral portions are mixed together, although in
such situations the vesicular-arbuscular mycorrhiza is more likely to
be found. It may be said, then, that mycorrhizae are formed wherever
rootage organs grow in contact with appropriate fungi.

Mycorrhizae and Soils: — Consequently, mycorrhizae are found
in a variety of soils. Von Tubeuf (1903) said: "Die Mycorhiza
findet sich auf Moorboden, im Waldhumus, auf nahrstoffreichen
Lehm — und gediingten Ackerboden und selbst im Bleisande des

Lecture VI — 69 — Environment

Eberswalder Kiefernbodens" ; and Sarauw (1904), after reviewing
the evidence, said that in general : "la formation des mycorhizes n'est
influencee que d'une maniere quantitative et non qualitative par les
differentes sortes de sols." But Prat (1926), in a study of Taxus,
concluded that the nature of the soil seems to exercise an influence
on branching of the roots. Harley (1937) agrees in that he says
the form of mycorrhizae in beech and extent of infection is correlated
with soil type. Kelley (1941) studied mycorrhizae of Pinus vir-
giniana in four soil series, — Chester (granitic), Conowingo (serpen-
tine), Dekalb (sandstone), and Sassafras (Cretaceous gravel); and
he found characteristic differences in each of the soil series. The my-
corrhizae were coral-branched in sandy soils and racemose or elongate
in clay soils, while pseudomycorrhizae predominated in wet clay soils.
In droughty sands were found necklace-beaded mycorrhizae, caused
by intermittent growth. A. Moller is said to have found different
sorts of mycorrhizae in different soils about Berlin (Henry, 1903).

Frank had supposed that mycorrhizae are found only on humus
soils but are absent from sands and sandy soils. Moller (1902)
thought, on the contrary, that pines produced mycorrhizae in sand
rather than in humus. In France, Vuillemin (1890) "a constate aussi
que le terre sabloneuse de bruyeres est favorable aux mycorhizes."
Pessin (1928) records pine mycorrhizae from Norfolk sandy loams
and Orangeburg fine sandy loam in the southern U.S.A.

Calcareous soils are usually considered unavailable to mycotrophic
plants since most mycorrhizal fungi prefer an acid substratum. But
Calhina is rather an anomaly since it grows on chalk downs which are
rich in mineral constituents but poor in lime. Rayner ( 1921 ) found
that the mycorrhizal fungus grew well even in a strongly alkaline
extract of pH 8.0; but Calluna developed normally in presence of
fungus (but with exclusion of bacteria) only on Ca-poor soils.
RuGGiERi (1937) found almond mycorrhizal on calcareous soils of
Sicily. Pine does not thrive on alkaline soils, and it is possible that
some of the difficulty with establishment of pine on prairie soils may
have been due to the alkalinity of the limestone soils of the region.
Young (1938) corrected alkalinity in nursery beds by S applications.

Solfatara soils of volcanic regions present special conditions.
MiEHE (1918) had drawn attention to the fact that mycotrophic and
bacteriophagic plants occur in numbers on solfatara soils, and sup-
posed that they were excellent pioneers inasmuch as they are able
to fix atmospheric N. Faber (1925) came to the same conclusion:
he said that these soils are characterized by Al-content, N-poverty,
acid content, and high temperature ; and that solfatara plants are
adapted to these factors. He found both xeromorphic and hygromor-

Kelley — 70 — Mycotrophy

phic plants included but decided that there was no "physiological
dryness" of the habitat, nor was there any lessening of transpiration
in these plants. They assimilate Al so greatly that they could be termed
"Al-plants". All sorts of mycotrophic conditions were described by
Takamatsu (1930) for the solfatara soils at Hakkoda, and the
mycorrhizal structure was similar to that of mycorrhizae from forest
soils.

Humus: — Whatever name is placed on the complex of substances
usually denominated "humus", it is manifest that these substances
are determinate in the distribution of mycorrhizae. Two principal
forms of humus, — raw-humus and mull — are associated with "ecto-
trophic" and "endotrophic" mycorrhizae respectively; and these in
turn are related to certain edaphic conditions, especially water. Hence
there is an interplay amongst organic detritus, microorganisms and
moisture that determines existence of mycorrhizae. This organic
detritus is undecomposed ; and when it is broken down with forma-
tion of mineral salts, it ceases to be humus. It is futile, therefore, to
speak of absorption from humus of water and mineral salts. It is
equally futile to say that mycorrhizae make use of humus : what mycor-
rhizae use is the protoplasm of invading fungi. These invading fungi
utilize humus through the soil portion of their mycelium, either directly
or through the mediation of microorganisms ; and the humus has
ceased to be humus when these organisms have made use of it . . .
Hence mycorrhizae never take in or "absorb" mineral salts ; and how
they acquire water is, to the best of our knowledge, still a matter of
conjecture.

Humus is naturally formed by the partial breaking down of
organic matter, chiefly vegetable, by the action of microorganisms.
Earlier workers, such as Nikitinsky (1902), had shown that humic
substances could not be used directly by higher plants but that they
were broken down by bacteria and fungi into simpler products that
could be used. In his well-known studies of humus-formation,
Falck (1923) used the term "Mykokrinie" to describe the chemical
changes involved in the decomposition of forest duff; and by this
process fallen branches, dead leaves, etc., are transformed into humus
and finally into "mineral salts" that the higher plant can utilize. In
Sweden, where Melin (1925) has studied so intensively, the forest
soils may be divided into a mull type wherein there is a more or less
rapid destruction of plant residues resulting among other things in
nitrate formation ; and this type has a rich herbaceous ground-cover ;
or a raw-humus type resulting from a less rapid destruction with
scarcely any nitrification while the organic matter remains in an

Lecture VI — 71 — Environment

ammoniacal state: this type has a moss and Hchen ground cover. In
raw-humus Melin visuaHzed a struggle between fungi and trees for
N, and the tree provided with mycorrhizae could compete with soil
organisms for that N. In mull soils, he found mycorrhizae (i.e.,
obviously ectotrophic) poorly developed. Yet Engler (1903) had
supposed that mull but not raw-humus is available to mycorrhizal
fungi. The nature of the soil plays an important part in distribution
of these fungi (Peyronel, 1921) ; and also on fungal form since in
soils poor in organic materials rhizomorphs develop while in humus
a disperse mycelium is produced. Then, not only fungal but mycor-
rhizal form is influenced by humus (Bjorkman, 1940), for Type
C of Melin, formed by Boletus spp., was found on occasional pine
plants in "mor" and sand (humus mixed with sand in a volume ratio
of 1 :2). An interesting line of work is brought out by Magrou: "In
the cultivated field, manure is capable of elevating the osmotic pres-
sure of the soil solution and, in consequence, that of the sugar of the
plant, beyond the level at which tuberisation might commence." (Ann.
d. Sci. nat., Bot, XI, 4:97-102, 1943).

Two ideas about humus have governed students of mycorrhizae,
vis., (1) that humus by its decomposition provides salts to the plants
growing in it, and (2) that humus buffers the soil by absorbing
deleterious substances. In other words, the influence of humus is
considered to be either chemical or physical.

The first idea is the older. Frank, in his experimental work
recorded in 1888 and 1889, indicated that fungi living in humus obtain
their nutrient from it and change over the humus into directly assimil-
able N compounds or ammonia. For beech, Muller (1886) concluded
that the tree lives on the remains of its own vegetative activity, a
peat being built up under the tree in which mycorrhizae live. Not
only beech but spruce and fir live thus in the Bavarian Alps (Eber-
MAYER, 1888), the tree using ammonia salts directly and also mineral
salts derived from the humus layer. Of these salts, coniferous litter
was found to contain more N, deciduous litter more K and P. (Ferry,
1887). By removal of leaf-litter, the fungi are deprived of their
normal food-supply and are transformed from mycorrhizal into
parasitic fungi (Delacroix, 1897).

The physical influence of humus on mycorrhizal development has
been adverted to by several earlier workers, as e.g. Hoeveler (1892)
who noted a rich branching of the mycorrhizal root system in humus,
while Shimizu (1930) thought that humus determines mycorrhizal
form in pine. But more recently the idea has been current that in
humus there are inhibitory or deleterious substances which interfere
with growth of plants, and that possession of mycorrhizae enables

Kelley — 72 — Mycotrophy

certain plants to grow in these soils. According to Freisleben
(1935), "The beneficial action of fungi on the growth of Ericaceae . . .
does not rest on a direct influence, as through the excretion of growth-
promoting substances, but on an inactivation, destruction or absorption
of the inhibiting materials. It is tO' be supposed that in natural soils
also the root fungi and the soil fungi which enter as components of a
peritrophic mycorrhiza, have a similar significance for the Ericaceae."
Again, Rayner's (1944) Wareham experiments indicate presence
of a toxic substance in this infertile soil and in a note, Neilson-
JoNES (1940) adduced some experimental evidence to prove the
hypothesis that emerged from Dr. Rayner's experiments in Wareham
forest : "The hypothesis was put forward that the local infertility on
the area is due to toxic residues formed during decomposition of
organic detritus by micro-organisms ; and that the effect of the
compost is to provide a substrate which, by altering the serial activities
of the different soil micro-organisms, modifies the chain of reactions
constituting soil decomposition ; with the result that the final residues,
instead of being toxic, are favourable to the growth of the trees, tO' the
mycelial growth of the fungi associated with them as mycorrhiza-
formers, and to the establishment and free-functioning of a normal
and balanced mycorrhizal relationship."

Mycorrhizae as Soil Indicators: — Bernatsky (1900) con-
sidered mycorrhizae as indicators of poor although well-aerated soils
except in the case of Alnus. Certain generalizations can perhaps be
made : A coralloid mycorrhiza is associated with a raw-humus forest
soil ; racemose mycorrhizae of light colour mostly come from mineral
soil ; bushy-branched rhizothamnia are characteristic of sands ; while
pearl-necklace mycorrhizae are found in droughty soils. In mulls
the preceding sorts are not likely to be found, but endotrophic struc-
tures prevail.

Mycorrhizae in the Soil Profile: — Mature natural soils present
a profile that is considered to exist in three "horizons", viz. the "A"
horizon, or zone of leaching and extraction of salts and colloids by
percolating waters ; the "B" horizon, or zone of concentration of
these materials ; and the "C" horizon, or zone of subsoil and rock
fragments where there is no visible concentration of leached materials.

There is not enough data extant to formulate any comprehensive
picture of the occurrence of mycorrhizae in the soil profile but certain
cases are known. In northern coniferous forests where raw-humus
develops, mycorrhizae are developed chiefly near the surface in the
uppermost A horizon. Masui (1927) figured a soil profile for "woody

Lecture VI — 73 — Environment

plants" in which mycorrhizae are located in a layer above the con-
ducting roots. For forest trees of central Europe, Klecka &
VuKOLOV (1937) state that mycorrhizae are developed most richly
and best in "the middle root depth". Scully (1942) found greatest
concentration of small roots in the A^ horizon and greatest numbers
of dead roots in a lower horizon. But trees may form mycorrhizae
at some depths in the soil : Pecan mycorrhizae may be formed at 30
inches depth (Woodroof, 1933), while mycorrhizal roots of Piniis
densiflora were found by Mimura (1933) at 10 m. depth. On shallow
infertile soils certain nut-trees have the roots confined to the upper
levels and the rootlets are almost entirely turned into mycorrhizae,
whereas on deep fertile soils the mycorrhizae are widely and deeply
distributed (Schuster, Stephenson & Evenden, 1944). Frank
(1887) had stated that in German forests the mycorrhizae occur in
the uppermost 1.5 cm. of humus while at lower depths there are
fewer although they may be found at ^ m. depth. Yeates (1924)
also found, in the New Zealand Podocarpus, that the roots are mostly
at the surface in an organic layer, a condition which, he said, is pos-
sible only in a rain forest. In Finland, Laitakari (1934), in an
elaborate study of root development in Bctiila, found that horizontal
roots occurred at depths of 2.8 cm. to 31.1 cm.; on moranic soils the
depths were greatest (av. 20 cm.) while on water-logged soils they
were least (ca 8 cm.). Vertical roots penetrated to depths of over
2 m., being deepest in clay. Fagus in England, according tO' Harley
(1937), shows variations in root systems with depth of soil: In
shallow soils on chalk the whole substrate is colonized while in deep
plateau soils the roots are fairly evenly distributed in upper layers
of mineral soil. In podsols and semi-podsols, fine roots are restricted
to litter and humus layers.

Herbs and dwarf shrubs apparently have superficially placed
mycorrhizae. Calluna root system (Rayner, 1911) is confined to the
first 12 inches of soil; while Burgeff (1932) stated that hemi-
saprophytic organs of orchids are found in the uppermost layers of
soil.

Soil Texture: — Effect of soil texture on root development was
neatly shown by Ter-Sarkisow {cf. Kirchner, 1908) for Pinus
sylvestris, 4 month old seedlings in pots showing the following root
development :

Number Length

Sand 363 713

Loam 181 420

Humus 54 179

Kelley — 74 — Mycotrophy

Pine is successful when planted in grey sand of Australia (Cromer,
1935) ; but Laitakari, already cited, found mycorrhizae most
plentifully developed in moorland soils and least in sands. Pecan forms
spreading clusters of mycorrhizal short-roots in light or sandy soils
but fan-like clusters in firm-textured soils such as the heavy red clay
subsoils of northern Georgia (Woodroof, 1933) . Influence of tex-
ture in humus on mycorrhizal form v^as neatly described by Mangin
(1910) : In soil formed of leaves which are superimposed and com-
pressed the mycorrhizae are often distichous with their branches in
the same plane ; while in contact with debris of cupules and fruits
their form is more or less complicated and branches of the mycorrhiza
are oriented in all directions and more or less pelotonized, dependent
upon size of the space in which it develops. In duff the mycorrhizal
short-roots of Populus were found to be clustered into nodules while
in sand they were betuloid in type, being ordinarily dark in coloui
except where growth is renewed (Kelley, 1937). Where a layer of
humus overlaid clay, Kelley (1941) found seedling pine with mycor-
rhizal short-roots in the humus but rootlets that had penetrated into
the clay beneath were transformed into pseudomycorrhizae. A some-
what similar case was reported by Frank (1888) in that a beech
seedling had mycorrhizae in the upper layers of soil (to 20 cm. depth)
while deeper occurring roots were fungus-free. Frank also noticed
that roots were much more richly branched in humus while in poorer
soil layers the roots assumed an elongate form. Perhaps the latter
were pseudomycorrhizae, but that term and concept had not been
formulated in Frank's day.

Soil Moisture: — Mycorrhizae are found only under optimum
conditions of moisture ; that is, optimum conditions during the period
of growth. Thus, mycorrhizae are absent from aquatic plants so far
as known, yet they may be found on marsh plants (Mason, Osborn),
or on semi-aquatics like Orysa (Peyronel, 1922). They seem absent
or less abundant under bog conditions where pseudomycorrhizae are
more frequently found but are common enough in meadows and in
moist ground in general. On the other hand, they may be found in
soil that is very dry and even on desert cacti (Asai, 1934, Johansen,
1931) but they are present under arid conditions as reserve organs
and were formed while moisture was ample for growth. But too much
drying may result in death of the mycorrhizae, and Paulson (1923)
described an interesting case from Epping Forest, England, where
an unusually dry summer killed the mycorrhizae, thus cutting off
supplies of nutrient to the trees and paving the way for entrance of
secondary parasites and saprophytes by which many of the birch trees

Lecture VI —75 — Environment

that composed the forest were destroyed. Yet in the case of pine,
Cromer (1935) found that, while drought caused collapse of the
cortex of absorbing roots, it did not affect that of the mycorrhizae.
Besides physical dryness of the soil, physiological dryness must also
be considered; yet Faber (1925), in a study of volcanic soils of Java,
decided that occurrence of the same association in the crater of the
volcano as well as lower down on the sides in wet volcanic soil may
not be explained through the hypothesis of "physiological dryness"
of solfatara soil but it is the result of individual nutrient conditions on
young solfatara soils.

Certain mycorrhizal plants such as Obolaria and Orchis spectahilis
are found only in moist shady places and thus indicate a relationship to
soil moisture. This relationship was noted as early as 1889 by Johow.
Dependence is probably on the mycorrhizal fungi which can extract
nutrient materials from moist humus and duff but not from dry mate-
rials. On the basis of soil moisture one might separate the mesic
mycorrhizal plants from the xeric ones such as ericads, conifers and
certain cupulifers, the former being chiefly endotrophs and the latter
ectotrophic. However, it must be noted that Voss & Ziegenspeck
(1929) have shown that xeromorphy in ericads may be due to physio-
logical conditions resulting from mycotrophy rather than to dryness
of soil.

It must be observed that mycorrhizae are chiefly developed in the
uppermost A horizon in what is naturally the driest portion of the
soil, at least on well-drained sites. For this reason there is a selective
action on mycorrhizal plants, the mesic species being confined to sites
where minimum soil moisture for mycorrhizal development is higher
than for xeric species. Certain anomalies in plant distribution can
thus be explained: Thus, Pimis virginiana grows on hills of Penn-
sylvania and Maryland but is absent from adjacent sand flats of the
coastal plain of Delaware where oak woodlands flourish. It was found
by experiment that seedlings of the pine transplanted to open sands
of the Delaware area could not withstand desiccation of the summer
dry season ; but where pine is watered or grows in favourable lowland
(there is a colony of P. Taeda near Newark), the tree is able to grow
in spite of dry seasons. In the same way, Orchis spectahilis grows
on shaded north slopes of Pennsylvania woodlands but never on sunny
south slopes where xeric ectotrophs thrive. Boudier (1876) had
noticed that Elaphomyces is found on south and east slopes more than
on north and west slopes at Montmorency in France.

Another fact that must be taken into account is that rainfall on an
area is by no means uniform in distribution nor regular in occurrence ;
and these irregularities have a consequent influence on mycorrhizal

Kelley — 76 — Mycotrophy

development. After a rain there is a rapid root growth but as available
soil moisture lessens in amount growth slows and may cease, to be
renewed with the next rainfall. Mycorrhizae therefore are not neces-
sarily structures of a steady growth that finally comes to a definite
end, but growth can be renewed. After a rain there is rapid mycor-
rhizal growth but as available moisture lessens, growth is retarded,
to recommence at the next rainfall. Every student of mycorrhizae has
seen old brown or even black mycorrhizae that have split their mantle
and protruded a new white mycorrhizal apex. With some plants the
periods of growth and quiescence are marked by constrictions or
rings, and the mycorrhiza assumes in consequence a beaded appear-
ance. Even in winter a warm rain starts new mycorrhizal growth
and one finds abundant white mycorrhizal root-tips.

Soil Solution: — Much is known of the composition and physics of
the soil solution but its relationship to actual plants growing in native
habitats is problematical. For, if plants are nourished through a mycor-
rhizal apparatus located in the uppermost A horizon, of what particu-
lar interest to them is a soil solution in the mineral B horizon?
Scientists, with that habit so ingrained in the human race, have gone
into the utmost minutia of research regarding the soil solution, but
they have never gone to the trouble to find out whether roots of
naturally growing plants actually come into contact with this solution.
Even for the mycorrhizae that do occur in the B horizon it is not
known what significance the soil solution has for them because the
mycorrhiza is buffered, so to speak, by fungal structures that more
or less isolate the mycorrhiza from the soil. Hence the whole question
of soil solution and mycorrhiza is largely conjectural, and because
of its character the various theories of mycotrophy are conditioned.

Some of the more recent studies have thrown incidental light on
the soil solution-mycorrhizal relationship. For some years Melin
has emphasized the importance of N salts to mycorrhizal plants as
indicated by laboratory tests. He has found that the simpler N com-
pounds, such as asparagin, can be utilized but that more complex com-
pounds, as peptone and nucleic acid, are used with difficulty. Harley
(1937) found with reference to beech that if the fungus supplies N
to the tree, it does not overcome low N content of the soil, and vigour
of the beech tree is more attributable to soil variations than to varia-
tions in infection of roots; yet infected roots had a greater N content
than uninfected. In the case of "fairy rings", Guinier (1937) found
that ammonia content of the soil was markedly higher within the ring
and grass found here was dark green. Also, in coniferous forests the
ammonia content of the soil was greater immediately beneath the

Lecture VI —11— Environment

sporophores of certain hymcnomycetes which form ectotrophic mycor-
rhizae with roots of trees. Guinier supposed that benefit of the
fungi to the higher symbiont consisted in accumulation of ammonia
in the soil. Mitchell (1937), in experimental studies with coniferous
seedling beds treated with various N, P, and K combinations found
that the benefits attributable to mycorrhizae, like their distribution in
nature, vary inversely as the concentration of readily available mineral
nutrients in the soil. As to what these mineral nutrients may be, a
paper by Chandler (1941) indicates that decay of leaf litter in a
central New York hardwood forest returns Ca to the soil in greatest
amount, N in the second greatest, followed by P, K and Mg. McComb
(1944) indicates P rather than N as incentive to good growth in
conifers.

Soil Reaction: — Anyone who has worked wdth soil reaction tests
knows the difficulties of securing an accurate reading for that highly
buffered colloidal thing called soil ; and if he has studied the relation
of plants to soil /jH, knows further the wide tolerance most plants
show for h.i.c. He is not surprised that Biraghi (1936) reports
cereals growing in soils of /)H 5 to 8, and he has sympathetic under-
standing for the report that Pinus radiata was planted in grey sand of
pU 6.18.

Long ago Melin (1925) stated that optimum conditions for fungi
of pine and fir are provided by pH values between 4.0 and 5.0 and he
noted with interest that this observation accorded well with observed
pH values for middle and northern Europe recorded by Hesselman.
Henry (1933) found trees and shrubs mycorrhizal on soils of pU
5.0 in Butler County, Pennsylvania. For Pinus Strohus, McArdle
(1932) reports a /)H of 6.0, a little lower for spruce. Germinating
seed and young seedlings of P. echinata cannot survive in culture
media having a soluble Ca content of approximately 500 p.p.m. or
more and a pH value of approximately 6.5 or more, or having either
of these characteristics. This condition was evidenced by behaviour
of seed in greenhouse cultures and of seedlings in nursery beds. With
P. caribaea in Australia, Young (1938) concluded from experiment
that "The efficiency of the mycorrhiza is increased with increasing
acidity up to pH 4.7 and thereafter is adversely affected." For orchids,
Burgeff (1932) found the optimum values lying between pH 5.0 and
6.0; while LaGarde (1929) said that h.i.c. is of the greatest impor-
tance in germination, growth being best at pK between 4.8 and 5.2;
and above 6.0 no germination took place.

It is evident that soil reaction affects the fungal symbiont rather
than the higher plant because the latter is virtually isolated from the

Kelley — 78 — Mycotrophy

soil. The soil is merely an anchorage medium for the higher plant
and the fungus is its body servant that makes contact with the soil.
It is the fungus that benefits from acid reaction and is limited in its
pH range. Modess (1941), in his investigations of mycorrhizal
fungi, found that all investigated fungi produced acid solutions.
Optimum grovi^th occurred with the Amanita spp. at the pH range of
3.5-4.5 ; with Paxillus Primuhis and the Boletus spp. (with the excep-
tion of B. variegatus) at pH 5.0 or somewha loove this value, relative
to Lactarius delicosus and Rhizopogon roseolus at pYi 5.5-6.0. A
species of Mortierella isolated from Empetrum made best growth at
pB. 2.77-4.0 (Hasselbaum, 1931).

It may be observed, however, that Henry (1936) records five
ectotrophs growing in Utah above the aspen zone where soil is neutral
or slightly acid; and Auret (1930) found the mycothalli of Lnnu-
laria growing in slightly alkaline soil of South Africa. Ridler (1923)
also reports Pellia in England on soils of pH 6.8-7.0.

The Use of Free Nitrogen: — The several studies relative to
fixation of atmospheric N by mycorrhizae may be summarized by stat-
ing that if such fixation occurs it is in too small amounts to be of
consequence to the mycorrhizal plant. Melin (1922) found that
fungi associated with mycorrhizae of Pinus sylvestris and Picea Abies
can in no case fix atmospheric N ; and in 1925 he stated that there is
no fixation of free N in mycorrhizae of trees examined. Moller
(1906) had found that dichotomons mycorrhizae of Pinus montana
are of no use to the tree in fixing free N ; the fungus of Empetrum is
likewise unable to use atmospheric N (Hasselbaum, 1931) ; the same
is true of the fungus of Monotropa (Francke, 1934) ; and also of
Mycelium radicis Fagi A (Aali, 1923). But Rayner (1922a) claimed
that certain strains of Phoma isolated from ericaceous plants can use
atmospheric N and she said that Aspergillus and Penicillium are
similarly capable but in varying degrees. She backed Ternetz (1907)
who had published similar statements. Furthermore, Neilson-Jones
(1928), experimenting with a fungus isolated from Calluna, decided
that the "plant can obtain nitrogenous supplies from the air, probably
in the form of molecular N, in sufficient amount to prevent the advent
of any symptoms of N starvation." The volumes of culture solution
tested were 50-100 cc, and the amounts of N fixed, from 0.00004-
0.00384 gm. But Addoms (1931) decided that if atmospheric N were
fixed by Phoma radicis isolated from cranberry (Oxycoccus) plants,
it was in amounts too small to be of service to the higher plant.

As to composition of the soil air in general and its effect on mycor-
rhizae, the author knows of no studies except that Laing (1923)

Lecture VI -79- Environment

noted deficient aeration and oxidation of peat soils affects distribu-
tion of mycorrhizae. This is all the more remarkable because work of
LuNDEGARDH and others would seem to indicate that soil atmosphere
might have a profound effect on the mycorrhiza and associated or-
ganisms, especially through an heightened CO2 content. The great
importance of optimum CO2 supply in tuber formation has already
been indicated by Molliard (1920), tubers failing to form in its
absence. It may be that in the future the soil air will be shown of more
importance to the mycorrhiza than some of the soil influences which
are now stressed.

Soil Temperature : — As with soil air, there are no direct studies
on influence of soil temperature on mycorrhizae. But it is well known
that soil temperature does not fluctuate to the same extent as air
temperature ; and in woodlands where the soil is blanketed with a layer
of duff it is partially insulated from fluctuations of air temperature.
A woodland with heavy leaf litter is so well protected from frost that
the ground may not freeze all winter and in consequence the mycor-
rhizae are not destroyed as they often are on freezing. In contrast,
a woodland that lacks a protective leaf litter freezes and thaws re-
peatedly and only certain plants, especially deep-rooted ones, survive.
Again, through freezing and thawing many seedlings are heaved out
of the ground whereby certain species are prevented from establish-
ment in a habitat which would otherwise be suitable for them. Yet
freezing does not necessarily destroy mycorrhiza! plants for it has
already been seen that such occur in arctic and alpine situations.
Chaudhuri (1935) stated that the endophytes of hepatics studied
can withstand very low temperatures and even an exposure to 0° for
four weeks did not kill any of them. A great number of mycorrhizal
fungi seem benefitted by low temperature but in nature mycorrhizal
fungi exist at many varied temperatures (Melin, 1925). In the
Japanese orchid, Galeola, the fungus is dominant when its optimum
soil temperature of 25 °C prevails while the host is more active under
the more congenial conditions of the colder months (Hamada, 1939).
Yet high temperatures do not prevent mycorrhizal development, al-
though presence of mycorrhizae in the tropics does not necessarily
indicate toleration of high soil temperatures since these may be
comparatively low and steady in the rain-cooled, shaded forest. But
mycorrhizae of cactus are certainly exposed to extremes of soil
temperature and show the hardiness of the mycorrhizal association.

The temperature of the soil may undoubtedly be changed by action
of microorganisms, and a soil with a rich microflora should be a
warmer soil and more favourable to winter survival of seedlings than

Relley — 80 — Mycotrophy

a sterile one. Greaves & Jones (1944) have suggested that addition
of manure to soil may add new microorganisms and modify the soil
temperature.

Altitude: — The production of mycorrhizae at various altitudes
has been studied especially by Costantin and his associates, the effect
of altitude being considered due to air temperature and thus parallel-
ing effect of latitude. In 1926 Costantin & Magrou observed
similarity of mycorrhizae of Dryas octopetala in the Alps and in the
arctic as described by Hesselman. Hitherto only ectotrophic mycor-
rhizae had been observed in such situations but now endotrophic my-
corrhizae were found widely distributed in the Alps. Annual plants
are absent from the Arctic and rare in the Alps, but the annual
Gentiana campestris was found to have an endophyte which, however,
underwent a "brutal phagocytosis". The authors came to an important
tentative conclusion that if essentially mountain genera are found
sporadically on the plain, their stations rest ephemerally because the
mycelium which is transported accidentally at the same time as the
seed does not reproduce itself. Later (1934) these same investiga-
tors, with associates, grew potato seed at 1,400 m.A.T. and obtained
some plants with infection, some without; but at 550 m.A.T. all the
plants died without producing a tuber. Since potatoes are not ordi-
narily grown in these alpine places, the authors concluded that potato
can form mycorrhizae with fungi already present in such situations.
These results are aligned with the theory that the mycorrhizal habit
in alpine plants commenced with a chance association of fungus and
root, forced beneath ground by inclement weather. Whatever value
this theory may have, it seems better established that there is an
optimum altitude for mycotrophy. Bouget had studied potato since
1901, and in 1922, with Bonnier, discovered the law of optimum
altitude, which was not published until rediscovered and published by
Lebard in 1931 (Costantin, 1936). Lebard & Magrou (1935)
state that, through three seasons' experiments it was shown that there
is an altitude where yield of potato is maximum, decreasing above or
below.

Light: — Light can affect mycorrhizae only indirectly since they
are not ordinarily exposed out of the soil. But illumination does affect
the vigour of the host plant, and the production of ionizable substance
in the host tissues. Bjorkman {cf. Romell, 1944) has studied the
effect of light on seedlings and it is stated that mycorrhizae are formed
in weak light (1/16 or sometimes 1/8 full sunlight), but under
greater illumination there were better seedlings with more mycor-

I

Lecture VI — 81 — Environment

rhizae. Naturally under optimum light exposure photosynthesis is
carried on most favourably and the host is accordingly better pro-
vided with assimilate. Bjorkman finds a connection between assimi-
lation and mycorrhizal formation, for which the reason is given in the
last chapter of this book.

Phenology: — With reference to seasonal aspects of mycorrhizae
there are a number of observations made incidentally in the course of
other studies. Early in the history of our science, R. Hartig (1886),
who was no friend to the concept of mycotrophy, asserted that tree
roots are free from fungi in summer and that mycorrhizae are present
only in autumn and winter; while McDougall (1914), upon whom
Hartig's mantle has to some degree fallen, claimed that mycorrhizae
are annual, being formed in summer and persisting through the winter.
A. B. Frank (1888) countered Hartig's statement by saying that
"die Mykorhiza zu keiner Jahreszeit ihren Pilzmantel verliert". The
mycorrhiza, he said, is formed in the earliest youth of the plant and,
like all absorbing roots, dies when it has exhausted its soil locus. My-
corrhizae can exist at least two years, probably much longer. These
statements were an amplification of his 1885 assertion that mycor-
rhizae have a limited life-span, some being lost while others are being
replaced ; and it seems evident that mycorrhizae are able to live several
years. Moller confirmed Frank by stating ( 1890) : "Als Beweis
dafiir fiihre ich an, das ich bei Material, welches im Januar gesammelt
war, gleichwie bei solchem im Juni sammtliche Entwicklungszustande
und in gleicher Verteilung gefunden habe". The fungus grew out
simultaneously with the tuberous mycorrhiza of the pine studied, in
summer rapidly but at other seasons as the cold permitted.

For the rebuttal, McDougall & Jacobs (1927) stated that at
7,100' A.T. on Mt. Logan, Utah, only dead mycorrhizae of the pre-
ceding year were found on Pseudotsnga inncronata. Above 7,000'
on Mt. Logan and at 10,000' on Mt. Washburn in the Yellowstone
Park, only dead mycorrhizae were found. New mycorrhizae can be
formed only when new rootlets are being developed and mycorrhizal
fungi are active, and these conditions seem to obtain in the latter part
of the growing season.

It will be recalled that Busgen (1901) did some cultural work on
ash, beech, maple, oak, and spruce, to learn more of the disputed ques-
tion of periodicity of root growth. He found that in Germany best
growth occurs in June and October with little growth occurring in
July and August. In March there are numerous roots growing, also
in November and December. In conifers a winter rest is indicated by
browning of the root-tips. Goebel, in the Organography, calls at-

Kelley

— 82 —

Mycotrophy

tention to the fact that some trees, as Tilia europaea, have greatest de-
velopment of roots in fall while in oaks the greatest development is in
spring. Other observations indicate similar generic differences : Thus,
Prat (1926) found that in Taxiis the long-roots grow throughout
the whole season with varying rapidity although cold lessens activity
and causes modification of the apex. Most of the absorbing rootlets
cease growth completely in winter, at which time the cortex dies from
the apex and exposes a red surface, while growth recommences in

Fig. 4. — Renewed growth of mycorrhiza-
bearing mother-root of Piniis Strohus, new
white mycorrhizae being formed amongst
old dark ones, with some rhizomorphic in-
vesting mycelium also indicated. Collected
at Baltimore, 26 February 1930.

spring. In pine, Rayner ( 1934) says positively that the mycorrhizae
are annual and only in exceptional cases is growth renewed. P. Bank-
siana is completely dormant in winter in Minnesota, the roots growing
from April to October (Kauffman, 1945). Again, McArdle (1932)
stated that mycorrhizae of spruce and pine are formed mostly in
September to November inclusive and that they are usually dead by
spring. Yet Preston (1943) found that pine mycorrhizae did not
appear to be "strictly annual", and several instances were noted where

Lecture VI — 83 — Environment

they had achieved renewed growth at the beginning of the growing
season by bursting through the fungal sheath. With Pennsylvania
trees and shrubs, mycorrhizae are present every month of the year
but particularly in late summer and autumn (Henry, 1933). On
deciduous trees of Scotland mycorrhizae are present every month from
November to March (Gordon, 1936). Still other observations are
to be recorded : Pecan mycorrhizae are to be found at all seasons in
Georgia (Woodroof, 1933) ; in Vitis it is a mistake to speak of the
dying of all roots in autumn, for only those formed in spring die while
those formed in autumn persist through the winter into spring (Rives,
1923) ; in Citrus seasonal variation was found, mycorrhizae being
best developed in the spring growing season (Reed & Fremont,
1935). In beech in England, the time of most rapid root growth
(chiefly spring and autumn) is marked by appearance of numerous un-
infected roots (Harley, 1937). This period is followed by one of in-
fection of the new roots. The shallowest chalk escarpment soils are
characterized by a short spring period of growth and infection ; in
deepest escarpment soils the spring growth persists longer, root growth
and infection going on together and being interrupted only by drought.
Infection is never complete and many uninfected roots are present in
spring and summer. In very acid plateau soils, roots are formed near
the surface and growth occurs in an upward direction in spring, incom-
pletely decayed litter of the previous autumn being colonized by un-
infected roots. In April and May infection takes place rapidly, while
in early summer it is nearly complete.

For herbaceous plants there are various reports. In liverworts the
fungus was found occurring in autum (Schacht, 1854) ; in Pyrola
Stahl (1900) found mycorrhizae also in autumn but not in spring;
while Endrigkeit (1937) said that plants of Convallaria and Maian-
themum are almost completely fungus-free in spring. Orchid roots
collected in September were uninfected (Costantin, 1926) while
Beau (1913) stated that roots formed in Spiranthes at end of the
flowering season are infected from the soil but not from old roots. In
Galeola the symbiont invades the cortex during summer and autumn
and ingestion proceeds through the winter until the following summer
(Ham ADA, 1939).

Cromer (1935) had noticed that mycorrhizae of Finns radiata
renew their growth after rain. According to Paulson (1924) dur-
ing drought of even short duration mycorrhizae are desiccated and
thereby killed. "Mycorrhiza does not revive after being destroyed by
lack of moisture and does not reappear on the return of copious rain
until new rootlets have been developed and they in their turn have be-
come associated with a fungus. . . . Observation of roots after heavy

Kelley — 84 — Mycotrophy

rain, which followed dry weather, has been sufficient to . . . conclude
that new rootlets followed by a complete change to mycorrhiza have de-
veloped within ten days."

As to phenology of internal anatomy, several observations may be
cited: In Hippophae, Arcularius (1928) said the fungus grows
best in summer and there are new vesicles present in winter. In Vitis
(Rives, 1923), vesicles appear at the end of the season, in August
and September. In Orchis, fungal digestion occurs chiefly from
autumn into winter (A. Fuchs, 1924). In Fraxinus, Kelley (1943)
found phagocytosis occurring in April and May, in Pennsylvania.

Mycorrhlzae in Relation to Habitat: — Apart from influence
of soils on mycorrhizal form, more recent studies have been directed
to influence of habitat as a whole on mycorrhizae. It is obvious that
environmental influences of the habitat react first on the fungus, as in-
dicated by Curtis (1937) : "There is an apparent correlation between
ecological habitat and fungus type, rather than between orchid species
and fungus." In conifers the possession of mycorrhizae seems de-
pendent on edaphic conditions (Dominik, 1937), and the more my-
corrhizae are developed the better the growth. Naturally, conditions
that favour the fungus result in a greater development of mycorrhizae.

In his review of soil fungi and root infection, Surges (1939)
considered the soil flora with its microhabitats ; and the relative abun-
dance of fungi, which is one-thirtieth that of bacteria but greater in
numbers than that of any other group. The biological groups of fungi
present in the soil the author considers as (a) root parasites, (&)
casual parasites and mycorrhizal fungi, (c) facultative parasites and
primary saprophytes, and {d) true soil fungi. The last group com-
prises those of a "humus type", the second group being most difficult
to study and some seem to be obligate parasites.

The term "microhabitat" graphically expresses the situation of a
mycorrhiza, for it is in a little cosmos of its own. Here it is subject to
the inorganic and biological influences of the immediate neighbourhood,
the "rhizosphere" as it has been called. Hiltner is said to have origi-
nated this term for the space about a root which is subject to root ex-
cretion, in which he thought there is an aggregated microflora. But
KiJRBis (1937) pointed out that fungi live in and on tree roots and
separate the root from the purely rhizospheric fungi. Consequently
Jahn (1934) invented the term "peritrophic mycorrhiza" and defined
the peritrophic fungus as one that lives in an outer zone, mantling
the root, between soil-portion and root-epidermis. Ordinarily con-
sidered saprophytes, they bear a definite relation tO' the root. He
said that in many cases endo-, ecto- and peritrophic fungi are present

Lecture VI — 85 — Environment

in the mycorrhiza at the same time; or, the cortical hyphae may be
neither parasitic nor endotrophic but peritrophic ; and the peritrophic
fungi may become dependent on the root plant. Jahn supposed that
the function of rhizospheric fungi is to change the h.i.c. of the rhizo-
sphere that it will correspond to the most favourable concentration for
optimum permeability of the roots. In an experiment to determine
whether it is the H- or the Ca-ion that is active, he found that several
fungi cultured from the rhizosphere caused heightening of the h.i.c.
of the culture solution without addition of calcium carbonate, but
with such addition the pH changes were less but nevertheless were in
an acid direction. With calcarous fungi there was better development
on addition of CaCOg than without it. It had been early suggested
(KuNZE, 1906) that there is not a simple relation between root-
secretion and mycotrophy but that the higher plant makes use of the
decided soil "ausschliessenden" action of fungi. So Kurbis decided
that the fungal flora of Fraxinus are not necessarily mycorrhizal, but
surround the root with acidity. He found that microorganisms were
greater in numbers in the rhizosphere than in root-free soil ; also, that
seedlings of Fraxinus dwindled and died in sterile sand but waxed
in unsterilized or inoculated soil.

Salt Marsh: — Two special habitats are to be considered, the salt
marsh and the prairie. A salt marsh, with its high osmotic coefficient
because of relatively large salt content, one would not suppose to be
favourable to mycorrhizae, yet two papers record characteristic pres-
ence of these structures in it. According to Mason (1928), mycor-
rhizae were found in such common coastal plants as Plantago mari-
tima, Aster tripolium, Glaux maritima, Armeria maritima and Glyceria
marithna, but no mycorrhizae were found in Salicornia europaea,
Triglochin maritimmn and several others, including Juncus. But
Klecka & VuKOLOV (1937) found mycorrhizal symbiosis in the small
roots of Juncus Gerardi, Salicornia herhacea, Suaeda maritima and
Triglochin maritimmn which duplicated that of endotrophic mycor-
rhizae of woody plants. The material was collected from saline soil
about Neusiedler See and from Auschitz and Louny in Bohemia ; and
the authors thought it very interesting that the fungi endured an high
osmotic pressure in root cells of these species. We would like to be
assured that these salt-marsh soils were truly saline, for our experience
with the New Jersey marshes indicates that such soil is not necessarily
salty.

Of 14 halophytes collected on the west coast of Sweden by Fries,
six bore thamniscophagous mycorrhiza which contained arbuscles,
vesicles and hypertrophic nuclei. (Bot. Not. 1944:255-264).

Kelley — 86 — Mycotrophy

Prairie: — The other habitat to be considered is prairie. Strictly
speaking, prairie is a special sort of meadow once found in the central
United States but the term is now loosely applied to non-forested
lands throughout the more northern portion of the Mississippi Valley.
Virgin prairie is now virtually extinct throughout the area and the
soils have been changed by agricultural practice. Since trees were
absent in the prairie area, except along water-courses and on some
rougher lands at time of discovery by white men, it has been supposed
the mycorrhizal fungi were absent from the prairies. Yet it appears
obvious that prairie grasslands existed simply because tree growth
was excluded by fire and difficulties of ecesis, and recent studies in
Iowa indicate a rapid spread of oak-scrub over former prairie lands
to the annoyance of the farmer. "Harrison County (Iowa) vegetation
was used by Shimek ... to support his thesis of climax prairie in
Iowa, yet 30 years later Quercus macrocarpa is spreading so rapidly
on the less intensively farmed lands of the country as to constitute a
serious economic problem". (McComb & Loomis, 1944). Apparently
these trees have no difficulty in ecesis. The author, while living on the
Iowa prairies, has personally seen how readily bur-oak becomes estab-
lished wherever prairie sod is uprooted.

But Hatch (1936) stressed a reputed absence of mycorrhizal
fungi from prairie soils, meaning by "prairie" apparently what is
otherwise known as "dry prairie" or "steppe". He noted from the
literature that "16 nursery and plantation failures have occurred in
widely separated regions of the world" due to "lack of a biologic
factor in the soils". He secured, through friends, some "prairie soil"
from Wyoming for his experiments.

As a matter of fact, Wyoming is several hundred miles west of
the prairies ; it is five thousand feet higher in altitude ; and it has a
different climate. The name, Prairie, may not be applied indiscrimi-
nately to all grasslands. The plains of North America, the pampas of
South America, the steppes of Asia, and the veld of South Africa
are all grasslands ; but they are none of them prairies.

In this Wyoming soil Hatch grew seedlings of Piniis Strohus and
found growth poor and unthrifty when mycorrhizae were absent but
on inoculation with pure culture fungi of several species growth
became good. N, P, and K determinations of the seedlings were made
after 10 months growth, showing marked increase in the absorption
of N, K, and especially P by the mycorrhizal plants. Hatch believed
the evidence was conclusive in showing that the pine seedlings grown
in this soil did not obtain sufficient nutrients to support normal growth
when mycorrhizal fungi were absent ; and he repeated his conclusion
in a paper published the following year. Rayner (1937), ignoring

Lecture VI — 87 — • Environment

the question of "prairie", commented on Hatches work and remarked
that there is some doubt as to whether the greater acquisition of N
by the mycorrhizal seedhngs in Hatch's "prairie" soil experiment
is related solely with the more efficient absorption of nitrates and his
claim that peptone and nucleic acid can be absorbed directly by the
roots of pine seedlings is not discussed from this point of view. In a
paper by McComb (1938, also' 1943) the claim is made, based on
experimental data, that differences in pine seedling development in a
forest tree nursery on old agricultural land in the prairie province
(Iowa) are due to disparities in the amounts of available P, and that
mycorrhizae are the means of enabling the seedlings to absorb this
element at a sufficiently rapid rate for normal growth. Thus Hatch
and McCoMB stress P but a writer in the Annual Report of the
Wisconsin Agricultural Experiment Station (1942) said that inocula-
tion of evergreen seedlings with suitable mycorrhizal fungi, particu-
larly Boletus feUeiis, greatly improved growth and survival on prairie
soil. The evidence obtained indicated that the mycorrhizal fungi
rendered the K present in the soil more readily available to the
seedlings.

That mycorrhizal fungi are absent from at least certain prairie
soils is asserted by Rosendahl & Wilde (1942), who found such
fungi in cut-over forest lands of central Wisconsin but "invariably
absent" from adjacent prairie soils. McComb & Loomis (1944) also
report a sharp difference in microflora between forest and prairie soils.
Harvey (1908) asserts an absence of fungi from prairies; yet it must
be noted that Pfeiffer (1914) found Thisniia mycorrhizal on the
prairies at Chicago; and Wilkins & Patrick (1938) presented a
paper on the fungi found in grasslands about Oxford. White (1941)
regarded mycorrhizal fungi as beneficial, and suggested that mycor-
rhizae exert a specific growth-promoting effect upon forest seedlings,
the absence of this stimulus being a major factor in the poor growth
of trees on mycorrhiza-free prairie soils. But the majority of cases
of poor growth of pine in the U.S.A. are apparently not associated
with mycorrhizal deficiency (Latham, Doak & Wright, 1939) ; and
in Indiana a failure that was so associated was more easily corrected
by fertiliser than by inoculation. "Even in new conifer nurseries in
the Prairie States growth is usually satisfactory . . ." Again, it must
be noted that Jones (1924) said of endotrophic fungi of legumes in
western America that no field, no matter how recently reclaimed, is
free from infestation and that but few mature leguminous plants are
uninvaded by mycorrhizal fungi.

A great difficulty with the question of occurrence of mycorrhizal
fungi in prairie soils is, that the subject has never been investigated.

Kelley — 88 — Mycotrophy

There is not a single paper devoted to an analysis of the subject and
not a dozen references in the literature. The reputed absence of my-
corrhizal fungi from prairie soils is simply a dictum mundi that has
been adopted trustfully as an axiom; whereas there are several facts
against it. Thus, trees and presumably root fungi have occurred from
time immemorial along the numerous watercourses which traverse the
prairies ; trees and shrubs flourish along streets of innumerable prairie
towns and about tens of thousands of prairie farmsteads. These woody
growths have a hard battle against desiccating winds and temperature
extremes in the trying climate of the region, but hardier species thrive.
It is true that conifers often fail in prairie soils, but it is possible that
the failure may be due to alkalinity of the soil, an alkalinity that pos-
sibly may be connected with the fact that the prairies are very generally
underlaid by limestone, although prairie soils are not residual. And
then it must be noted that trees spread rapidly into the prairies when
the sod is broken. As to the Plains which lie west of the prairies,
aridity is a potent influence on plant development, and establishment
of trees in these droughty soils must always be conditioned by the
water supply as well as by other "soil factors".

Soil Inoculation: — On the assumption that necessary mycor-
rhizal fungi are absent from certain soils on which trees are to be
grown, the practice of soil inoculation with these fungi has arisen.
Thus Rayner (1934) found that seedlings which grew poorly on a
sterile heath were greatly benefitted by application of humus which
"must contain active mycorrhizas of the species free from any ab-
normalities of structure and from contaminations of such pseudo-
mycorrhizal fungi as can be identified". Probably there were "active
mycorrhizas" in the transplants made to various treeless regions in
those benighted days before the science of mycorrhiza flamed so
brightly. Thus, Leonard Flemming, a pioneer in afforestation in
South Africa, says nothing of inoculating the soil when he planted
thousands of seedling pines on the high veld where trees had never
grown before. It was water that the pine-trees craved and when he
supplied their need the trees flourished. But in Australia, exotic
conifers needed for softwood plantings in the Brisbane Valley often
failed to grow at the Yarraman nursery, and on examination it was
found that the roots lacked mycorrhizae (Young, 1938). (In passing
it may be remarked that Frank in 1894 had raised the question
whether the needful fungi were present in all soils used for planta-
tions.) On inoculation of the seed beds with the proper mycorrhizal
fungus, mycorrhizae were formed and the seedlings became thrifty.
Moreover, it appears to be part of the standard practice to inoculate

Lecture VI — 89 — Environment

nurseries in Western Australia with the appropriate fungi, thus obtain-
ing normal growth of the tree seedlings (Kessell, 1938). The inocu-
lated plants when put out in the forest are said to infect the soil quite
satisfactorily. Rayner (1938) gathered together various reports on
soil inoculation from nurseries and plantations, particularly from the
British Empire. In northern Rhodesia it was found that Pinus
halepensis only amongst several exotic pines made any growth beyond
the seedling stage without soil inocula, whereas inoculation with soil
from a southern Rhodesian P. radiata plantation caused remarkable
stimulation in growth in several spp. of pine. In Nyasaland, all species
of pine observed except P. longifolia and Araucaria failed to grow
without inoculation. At Buitenzorg in Java, P. Merkusii is completely
dependent on the presence of mycorrhizal infection for normal de-
velopment, inoculation resulting in vigorous growth and rapid spread
of infection from plant to plant. In New Zealand, inoculations of
P. radiata with Boletus-'miected soil gave positive results (whatever
that means), the control plots remaining free from infection. In India,
Casiiarina flourished after inoculation whereas controls died within
three years. Caragana became established in Canada after use of soil
inocula. At a new forest tree nursery in Iowa, pine seedlings failed
to grow unless they developed mycorrhizae (McComb, 1943). On the
other hand, S. A. Wilde remarked in a recent review that "99 percent
of all practicing foresters will not have to lose any sleep over the
problem of mycorrhizal inoculation."

Compost Studies: — But soil inoculation with mycorrhizal fungi
does not necessarily lead to mycorrhizal formation because the soil it-
self may be unfavourable to such formation even though the appro-
priate fungi are present. Thus, in Rayner's heath soils mentioned
in an earlier paragraph, it was the inhibitory effect of the soil that
prevented mycorrhizal formation. Rayner therefore initiated studies
of "organic composts" in relation to growth of young trees. Her
general conclusion after several years' experiments is that an increased
supply of nutrients plays a relatively insignificant part in improved
fertility of the soil studied, induced by addition of composts. In the
soil are substances deleterious to growth, but their action is obviated
by addition of compost although addition of the equivalent amount of
salts had no effect. Rayner considers that the striking effects on
tree growth brought about by composts on natural soils do not depend
to any extent upon the addition of nutrients, but are directly associated
with qualitative changes in the humus constituents and with the bio-
logical activities related with these changes. They may also, possibly,
be associated with the presence of growth-promoting substances in

Lecture VI — 90— Mycotrophy

individual composts or produced in the soil as the result of fungal
action. The chief biological change in relation to fertility of the soil
is production of toxins, according to Brian (1945), especially of
"fungistatic organic substances" produced by Penicillia. The chief
toxin appears to be gliotoxin, which has been found highly toxic to
mycorrhizal fungi. Brian suggests that the toxicity of Wareham soil
may be due to accumulation of gliotoxin and other antibiotic
substances.

Valuable as Dr. Rayner's (1944) studies on the Wareham area
undoubtedly are, the results will of course be applied with caution to
other areas. Results obtained with a very unusual soil existing at low
altitude but high latitude under an oceanic climate will not necessarily
be applicable to all other areas. As evidence in point, the paper by
LiNDQUiST (1945) may be cited, in which it is stated that "larger and
better-colored seedlings" of Pinus resin osa were grown on a duff-peat
mixture than on a compost-peat area. Again, composting often pro-
duced abnormally crooked roots (Muntz, 1945).

A study of the organic matter of forest soils led Romell (1938)
tO' a new theory of mycotrophy. In experimental work in a spruce
forest in Sweden he sank sheet-iron shielding, one foot high, in a poor
stand of spruce, surrounding two quarter-hectare plots. One plot,
being covered with blueberry bushes, was mowed with the scythe while
the other plot was untreated. The author states that a marked efifect
resulted, for the vegetation on the plots became thriftier and greener,
and retained its foliage longer in autumn. Romell considered the
effect due to killing of the tree roots or of mycorrhizae and their
associated fungi by trenching, the organic matter thus killed becoming
"green manure" for the remaining vegetation. Also, root competition
of the trees, and fungal competition, was stopped. Numerous sporo-
phores of the supposed mycorrhizal fungi were formed outside the
plots while practically none were formed within. Romell thought
that these experiments show a fundamental physiological difference
between litter-decomposing and mycorrhizal fungi, the latter being
practically unable to break down dead organic residues under condi-
tions prevailing in nature. He points out the value of trenching experi-
ments in mycotrophic studies, since laboratory experiments show
merely what is physiologically possible but not what is ecologically
important.

LECTURE VII

MYCOTHALLI AND MYCORRHIZOMES

General Character: — Mycothalli and mycorrhizomes are ordi-
nary liverwort gametophytes, fern stems and orchid rhizomes that
possess endophytes. Most of these structures in nature appear to be
invaded with fungi, for apparently most thalli and prothalli that are
not actually in water are mycotrophic, and most rhizomes likewise.
Here again the fortuitous character of the symbiosis is seen, since
apparently the fungi simply grow into these structures as into a part
of the environment ; and there is nothing evidently obligate about the
relationship.

Mycothalli in Liverworts: — Their structure is detailed for
Pellia by Ridler (1922): In Pellia no plants were found entirely
without infection and usually the endophyte occurs in a definite zone
along the thickened median portion towards the ventral surface of the
thallus and in the rhizoids. Infection from the soil is presumably
through the rhizoids. Within the thallus, penetration of the cell-walls
seems effected mechanically ; the hyphae are swollen where their
growth is arrested by cell-walls, and they are constricted by passage
through them. The liverwort seems to exert some control over the
fungus and limits its invasion as stated, to a definite region in the
thallus. Here the hyphae form arbuscles or bushy-branched struc-
tures which later degenerate into sporangioles or little rounded bodies
that are insoluble in usual reagents. Formation of arbuscles stops
further growth of the fungus and this phenomenon caused Bernard
(1909) tO'term it an "immunity humorale". The effect of the fungus
in Pellia is very marked for protoplasmic content of invaded cells of
the thallus is killed, chloroplasts disappear and cells ultimately become
brown in colour. Starch disappears from cells of the thallus on
entrance of the fungus and is replaced by oil. When the sporophyte
is invaded (the thallus is of course the gametophyte) the contents
of the cells are wholly or partially absorbed. The fungus invades the
region of the sexual organs but does not grow into them.*

♦According to Peyronel, the Jungermanniaceae are infected only by
mycomycetes. On poor soil, infestation dwindles with decrease of light (Nuovo
Gior.Bot.Ital. 49:362-382, 1942).

Kelley — 92 — Mycotrophy

Infection of Mycothalli: — Infection seems to take place always
through rhizoids and is thus reported by all workers. Kny (1879)
said that "In numerous root hairs (sic) of Lumilaria (from the uni-
versity greenhouse) it was observed that a great part harboured
thread structures. In a series of cases these were sterile fungal hyphae
which branched hither and thither". "Seven cultures of Calypogeia
from very different habitats about Hilversum showed almost all
rhizoids attached to substratum infested while aerial hyphae were
fungus-free." (Garjeanne, 1903) Fungal hyphae penetrate rhi-
zoids of Marchantia and Lumilaria, especially where plants grow in
humus (Cavers, 1903) : hyphae were found in rhizoids of Lunularia
in South Africa (Auret, 1930) : in Italy, Bergamaschi (1932) found
in Fegatella and in Lunularia that non-septate hyaline hyphae invaded
the rhizoids and passed into underlying cells ; while Chaudhuri
(1935) found hyphae in rhizoids of all Indian liverworts investigated.
An endophyte penetrates rhizoids in Sezvardiella of southern India
(Chalaud, 1932). In Zoopsis of Java, the rhizoids frequently har-
bour hyphae which form pelotons and refractory granular material,
perhaps albuminoid. Divers other hepatics from the same forest pre-
sent the same endophyte (Janse, 1897).

Limitation of Endophyte: — Limitation of the endophyte to
a definite portion of the thallus seems general. In New Zealand liver-
wort, Monodea Forsteri, every thallus possessed a sharply defined
mycorrhizal zone consisting of 2-4 layers of cells densely filled with
branching fungal hyphae (Cavers, 1903). This zone is confined to the
thicker median portion of the thallus and extends to within a short
distance of the growing point. Hyphae pierce the cell-wall and branch
out in the cell cavity, the nucleus of the infected cell grows in size and
often becomes enveloped by a tuft of short hyphal branches and some-
times the chloroplast becomes similarly enveloped, suggesting in ap-
pearance a lichen. On some of the hyphae are formed large spherical
vesicles. In Lunularia criiciata the fungus is confined to a definite
zone below the assimilating tissue (Auret, 1930) ; it occurs also in
the rhizoids and amphigastria but does not penetrate the gemmae-cups
and archegonia. The fungus consists of branched septate hyphae
with granular contents giving rise to vesicles, arbuscles and sporan-
gioles which conform with the general type of endophytic fungus found
in a great variety of higher plants. All plants, infected or uninfected,
are green and apparently healthy. Nicolas (1942) found in Lunu-
laria two sorts of infection: {!) confined to a band which runs the
length of the mid-nerve parallel to lower surface and removed from
it by several layers of immune cells rich in starch: (2) In other, male,

Lecture VII — 93 — Mycothalli and Mycorrhizomes

thalli the fungus is localized in cells throughout the thallus. Sterile
thalli were destitute of mycelium and Nicolas thought that presence
of fungus is necessary to fructification.

Emberger (1924) also found hyphae in Lumdaria cruciata oc-
cupying a large band separated from the lower surface by several
layers of cells ; the chloroplast tissue is never invaded. Inconstancy of
infection, he thought, negated the hypothesis that infection is neces-
sary to formation of sexual organs ; and the association seemed simply
accidental. It was supposed by Ncmec (1899) that mycothalli are
general in Jungermanniaceae but rare or absent from Marchantiaceae.
We have already seen that infection is common in Lumdaria; it also
occurs in Marchantia and a number of other European hepatics ac-
cording to GoLENKiN (1902), who found that in all cases the fungal
hyphae are confined to the compact ventral tissue ; and infected cells,
though they retain nuclei and protoplasm, never contain starch or
chlorophyll. Thalli of Marchantia nepalensis on sand and clay at
Lahore, India, contained a fungus limited to a zone beneath the air
cavities, and branched and interwoven in the cells (Chaudhuri,
1925). Chlamydospores were sometimes found. In this and other
Indian liverworts, infection is localized in regions definite for each
species (Chaudhuri, 1935). Conocephalus is similar to the preced-
ing, as described by Bolleter (1905) who found the thalli often
turned red upon infection; but in alpine situations the thalli turned
red without infection, — another fact in line with Costantin &
Magrou's idea that refrigeration parallels the action of mycotrophy.

Digestion of Endophyte: — In his description of mycothallism
in Pellia, Magrou (1925) said that the fungus degenerates about the
archegonia or the sporogonia, which organs seem to exert an inhibitory
influence on its growth. The endophyte exhibits all the structures
characteristic of mycorrhizal fungi, — large non-septate hyphae, ar-
buscles, sporangioles and multinucleate vesicles, the contents separat-
ing into uninucleate cells. Digestive structures were also described by
Garjeanne (1903) from thalli of Netherlands hverworts, — haustoria
and hyphal coils (Knauel) ; and under influence of the latter the
cells disorganized. Immersed clots were found in a number of liver-
worts by MiLDE (1851), while Ncmec (1904) found clots in Caly-
pogeia coincident with degeneration of mid-hyphae: they disappear
before death of the thallus. Many vesicles were formed in tissues of
Conocephalus but few in Lumdaria (Bergamaschi, 1932). Chalaud
(1932) figures vesicles and arbuscles in S ewardiella.

Tuberous-thickening of the stem of Fossornhronia with which a
fungus was always associated was noted by Humphrey (1906) ; while

Kelley — 94 — Mycotrophy

Chalaud (1932) found tuber-formation in Seivardiella "in all re-
spects like the stem of other Metzgerias, especially Petalophyllum and
Fossomhronia."

Denis (1919) found an endophyte in the chlorophylless thalli of
Aneura, and compared them to similar thalli known amongst lycopods.

Mycothalli in Fern Gametophytes: — A description of the my-
cothallus in Opioglossiim pendulum is given by Lang (1902). Tissues
of the young prothallus are parenchymatous throughout, cells of the
lower portion contain an endophytic fungus while those of the upper
portion are free from it. Infection is usually through a rhizoid.
Superficial cells of the prothallus contain only infecting hyphae, the

Fig. 5. — Section through an older mycothallus of Botrychium
obliquum. Shaded portion indicates the extent of region occupied by
endophyte, vicinage of reproductive organs being uninvaded {Redrawn
from Campbell, Ann. Bot. 35, pi. viii, fig. 12, 1921).

fungus otherwise being confined to internal tissue. In older regions
of a branch the fungus occupies all the cells except for a superficial
zone of 1-2 layers. A number of vesicles are formed in a cell, often
close to the nucleus, while other cells contain thick coiled hyphae.
Plastids and chloroplasts occur in cells occupied by the fungus. The
European Ophioglossum vulgatmn mycothallus was described by
Bruchmann (1904). Infection is directly through the epidermal
cell- wall and hyphae spread through the mid-portion of the prothallus
but the outer cells are always fungus-free. Innermost cells are also
fungus-free and contain starch. Nuclei of infected cells increase in
volume while hyphae coil in the cells and form an irregularly shaped
structure: vesicles occur in older portions of the prothallus. The in-
fected portion forms in effect a cylinder which is particularly well
developed about the sexual organs.

Campbell (1907) in general confirms Lang's description of
prothalli in Ophioglossum, and in the appendix of his Mosses and

Lecture VII — 95 — Mycothalli and Mycorrhizomes

Ferns gives a longer description ; he found infection also in prothallus
of Osmitnda cinnamomea. In 1908 he stated that he had found an
endophytic fungus normally present in green prothalli of several
Marattiaceae, Osmiindaccae and Gleicheniaceae. The endophyte con-
sisted of large, branched non-septate hyphae w^hich are strictly intra-
cellular. Vesicles and apparent digestion stages are described, and in
figure ten there is shown a vesicle near an intact nucleus. In 1921,
Campbell said than an endophytic fungus occupies a large part of
inner tissue of gametophyte of Botrychiuni ohliqimm, but in older
gametophyte it does not invade tissues about reproductive organs. The
fungus fills lumen of cells but nucleus remains intact. The myco-
thallus of B. lunaria is described by Bruchmann (1906), infection
taking place usually through rhizoids although it may occur directly
through surface of the prothallus. Outer cells are at first invaded but
later the fungus leaves them and is confined to middle and basal cells
which have large nuclei and fungal clots. Starch is present only in
meristem and in cells about reproductive organs while in those cells
which have no starch the hyphae are filled with oil and protein. The
advantage of mutualistic life of fungus and prothallus seems to con-
sist in a holding and storing of reserve, especially oil, which is of value
during summer heat and winter cold in protecting the prothallus from
drying. The endophyte is present in every prothallus, living in all
the inner and radially formed outer portion.

Stokey (1942) found no infection of gametophyte of Marattia
or of Macroglossum grown on sterilized peat, growth being vigorous
and "normal". The structure of Hehninthostachys zeylanica prothalli
is essentially similar to that of Ophioglossmn (Lang, I.e.) The cells
containing vesicles seem healthy but starch is usually absent from
them. The fungus is healthy until growth of the sexual portion of
the prothallus commences, whereupon the fungus dies and the pro-
thallus develops up to the extent of the amount of reserve material.
In prothalli of the fern Cheiropleuria, Nakai (1933) found fungal
hyphae which entered by way of brown rhizoids and filled the median
part of the prothallus where they branched and coiled to form a
nutritive layer. The median layer is stimulated by presence of fungus
to form 5-10 layers of cells. Uninfected prothalli were also observed
and Nakai thought these may have been sterile or male.

Mycothalli in Lycopod Gametophytes: — Following Treub's
discovery of endophytism in a Javan lycopod, Bruchmann (1885)
described fungal structures in prothallus of Lyeopodium; and in 1898
said that a Pythium sort of a fungus occurs in palisade and cortical
layers of L. elavatum and L. annotinum, and is also found in rhizoids

Kelley — 96 — Mycotrophy

from whence it comes in contact with the soil. Goebel (1887) stated
that the lower, non-meristematic, portion of the prothallus of L.
inundatum was always without exception inhabited by a fungus
which forms coils within the cell content without killing the cell, the
nucleus remaining plainly visible. From the coil a branch may go
through to the next cell. The fungus is limited to one or two cell
layers forming a zone separated from the exterior by several cell
layers, and it is unable to penetrate the meristem, or lobes of the
prothallus. The infected cells do not contain starch but drops of oil.

According to Holloway (1920), a fungal symbiont occurs in
epiphytic prothalli of the New Zealand Lycopodium Billardieri, L. B.

Fig. 6. — Portion of mycothallus of Lycopodium obsciirum
shown in section, with pelotons or fungal coils and oil
globules (Redrazvn from F. L. Barrows, in Contr. Boyce
Thompson Inst. 7:299, fig. 34, 1935).

gracile, L. varium, and in the epigeic species L. cernuum, L. laterale
and L. ramidosum. In the epiphytic species, the fungus occupies the
base of the prothallus even in epidermal cells at the prothallial point,
and grows forward with the prothallus, occupying a zone between the
epidermis and the central conducting cells. Fungal coils soon disap-
pear in many of the cells, their place being taken by clusters of darkly
staining oval "spores" which are probably used by the prothalli ; but
oil droplets were not seen. In the epigeic species, fungus is present
only in the epidermis of prothallus but occurs in inner tissues of pro-
tocorm where only "spores" were observed.

Lecture VII — 97 — Mycothalli and Mycorrhizomes

An endophytic fungus, thought to be an Ascomycete, is described
from prothalli of L. luciduluin and L. obscurum var. dendroideum by
Spessard (1922). It enters through rhizoids or through epidermal
cells (or, the author suggests that the fungus may be leaving the plant
through these structures) and spreads tO' within 2-3 cell layers of
meristem. The mycelium is coiled in lower part of prothallus (the
fourth layer from outside) and does not enter the palisade. Spore-
like bodies were found in L. luciduluin, sometimes with chromatin-
like content and sometimes with fine hyphae proceeding from pores
as though the body were germinating ; while in some cells were found
true spore-bodies. Stokey & Starr (1924) cite fungal infection of
L. complanatum, L. obscurum and L. clavatmn; and state that fungal
hyphae were usually found in great abundance in the soil in which
prothalli were growing. But in culture Barrows (1936) found that
an endophyte isolated from a Lycopodium did not aid development of
germinating spores of L. complanatum var. flabelliforme; in fact, it
proved impossible to grow the plantlets at all beyond a ten-cell stage.
In nature. Barrows (1935) found endophytic fungi in gametophytes
of Lycopodium sporophytes, including L. annotinum, L. clavatum,
L. complanatum var. flabelliforme, L. lucidulum, L. obscurum, and
L. tristachyum.

Mycorrhizomes in Ferns: — Originator of the term "mycor-
rhizome" was Dangeard (1891), who described such a structure from
several species of Tmesipteris, although not very clearly. He described
and figured an apparent Hartig net but said the infection seemed
typically endotrophic ; and he figured what appear to be vesicles. But
mycorrhizomes have long existed : Andrews describes a fossil mycor-
rhizome from a coal-ball; and doubtless there are innumerable fern
mycorrhizomes were there eyes to see. A few descriptions of them
come from particularly close observers, such as van Tieghem (1870)
who found mycelial hyphae of a parasite {sic) coiled about dark
masses in large cells of the inner zone of Osmunda regalis and several
other ferns. Rayner (1927) cited and figured infection of Pteridium.
LoHMAN (1927) showed vesicles in his illustration of Adiantum
pedatum and figured an apparent digestion stage for Botrichium. In
general, however, ferns remain for future investigation, because
they are not, like the pines, an economic group that commands
attention.

Mycorrhizonies in Orchids: — Aside from studies of ferns and
fern allies, the only other rhizomes to- be studied for endophytes are
those of orchids, except for the following: Kamienski (1881) de-

Kelley — 98 — Mycotrophy

scribed infection in rootstocks of Monotropa; Oliver (1890), in
Sarcodes; MacDougal (1900), in Pterospora; and Pfeiffer (1914),
in Thismia. An endophyte was discovered in an orchid rhizome by
Link in 1840; while an early student of orchid mycorrhizomes was
Prillieux (1856), who took his historical introduction back to
Tragus of 1552, confirmed Schacht that threads penetrating tubers
of Neottia are fungal hyphae ; and stated that at St. Germain he had
found sand grains agglutinated in a mass about the orchid. Within
the rhizome the 2-3 outermost cortical cell layers were filled with a
yellowish-brown material (a material which he found in a great many
genera) and the cells containing this material retain their nuclei.
These nuclei became very large and often had two nucleoli. The cells
having the brown matter regularly contained filaments wound without
order about the central mass in the cell ; and not infrequently the fila-
ments branched and penetrated through the cell-wall into another
cell. After a time this matter diminished in amount, from which fact
it may be inferred (said Prillieux) that it serves for nutrition of
the plant. Recall that this description was written 29 years before
publication of Frank's epochal paper.

Pfeiffer (1877) confirmed this report for Neottia, finding fungal
infection constant and supposing that the fungus takes the place of
root hairs which the orchid lacks. A most detailed study of this same
orchid was made by Werner Magnus (1900) : he found 3-4 of the
outermost cortical layers of cells infected, — sometimes even 6 layers.
In this paper Magnus distinguished between "host" and "digestion"
cells. In the host-cell the fungus never degenerates : "The cells here
pictured, which always possess that ring of thick-walled hyphae with
various modifications and the coil of fine median hyphae, — these cells
in which the fungus does not degenerate but remains living to the
last, shall be designated henceforth as host-cells (Pilzwirthzellen)"
(p. 216). Thick-walled hyphae run ring-formed, in various modifica-
tions out to the cell-wall and send out fine, thin-walled haustorial
hyphae which gain control of the whole cell, — haustoria which seem
fitted for passage of nutrient. These cortical ring-like hyphae remain
alive after death of the root.

In contrast tO' the host-cell region, in the digestive region the
fungus always degenerates. Thin-walled, protoplasm-rich hyphae
grow through the cells in thick coils but very soon die ; or after they
have formed protein (as Eiweisshyphen) their content is taken up by
the cell and the residue is pressed together while at the same place or
at a place mostly lying in the middle of the cell a clotting formation
takes place, which results in their separation with a portion of the
plant plasm as a clot, which is a dead unchangeable waste product con-

Lecture VII — 99 — Mycothalli and Mycorrhizomes

sisting of plant and fungal material. Of the fungus-inhabited layers,
the digestion cells take the outer and inner while the host-cells take
the middle. The digestion cells are defined by Magnus as follows :
"If, in Neottia, the fungus in a cell does not take on the modification
which characterizes the host-cell but branches again after the
'meristem condition' into thin-walled hyphae that inevitably encounter
a certain developmental process, — death, robbery of content and final
mantling into a clot, a development not less sharply delimited than in
the host-cell, — we shall designate such a cell a 'digestion-cell' (Ver-
dauungszelle)" (p. 223). Infested cortical cells are enlarged, and
later formed cells are also enlarged, causing a change in the whole
structure. The plasm continually surrounds the fungus in the diges-
tion cell and upon death of the fungus a copious formation of vacuoles
takes place. Vacuoles neighbouring the wall-layer which is free from
the fungus unite to form a large sap-vacuole and thereby separate
the clot, which either remains suspended in the sap-vacuole or is com-
pletely separated from the protoplast by formation of a new internally
lying plasm-layer. Plasm of the fungus-inhabited cell never dies before
death of the whole root. Plasm segregated in the clot becomes
changed into a cellulose sort of a substance. Upon migration of the
fungus there arises a fine-grained starch which soon dwindles but
after death of the fungus reappears in a modified form. The nucleus
becomes constricted or amoeboid and intensely chromatophilic, but
after phagocytosis is completed the nuclei return to their former
barrel-shape.

Bernard (1899) described the mycorrhizome of Neottia as ex-
hibiting three zones of cells: (i) a starch layer; (2) several layers of
cells filled with intertwined mycelial filaments ; (5) epidermis, without
starch or hyphae. Spiranthes autumnalis dififers from other Neottiae
(according to Beau, 1913) in being annual, but it has mycorrhizomes
which are the organs of reserve and at time of flowering of the orchid
are invaded by an endophytic mycelium as evidenced by a pronounced
yellow colour given the sections through bodies resulting from diges-
tion of mycelial coils. Towards the end of the flowering season new
"roots" are formed which must be infected from the soil.

Orchids other than Neottiae have received attention : Calypso has
a coralloid-branched mycorrhizome but Lundstrom (1889) failed to
find infection in plants of C. borealis collected in southern Sweden.
But in C. hidhosa, MacDougal (1899) found fungus living in outer
cortex but not passing out through nodal trichomes; its hyphae are
septate and form vesicles. Inner cortex and apex are free from infec-
tion. Corallorhiza arisonica, according to the same author, has the
coralloid rhizome represented by papillae which are infested early by

Kelley — 100 — Mycotrophy

a fungus which fills the mediocortex and grows forward with the
apex. The nucleus is seldom affected by hyphal invasion. Corallorhiza
innata of the Alps has a coralloid rhizome which bears papillae from
which tufts of hair arise, and at the tips of the hairs chemical changes
seem to take place. Hyphae pass directly from the soil through the
hair into the rhizome, going through the outermost layer of cells
(which are rich in starch) to a zone in which they coil within thin-
walled cells. There is a paucity of starch in this zone but within is a
third zone in which starch increases in quantity as hyphae become
less numerous, and all stages in degeneration of fungus may be seen
in cells of this region. The nucleus enlarges and contains bodies which
stain deeply with Hoffman's blue (Jennings & Hanna, 1898).

In the Australian orchid, i?/M>aMf/t^//a, PiTTMAN (1929) described
fungal infection of the rootless rhizomes to a depth of ten cell layers,
the epidermis being fungus-free. Clots are illustrated as chiefly in
the exocortex. Infection was through hairs borne on the mycor-
rhizome. No arbuscles, vesicles or sporangioles were seen, but the
hyphal clots degenerate into a golden-brown mass.

In Gastrodia the fungus inhabits superficial cells of the fleshy
succulent rhizome (McLuckie, 1923), the fungus being Armillaria
(KUSANO, 1911).

Various other orchid mycorrhizomes have been described, as in
Orchis, Cephalanthera, etc. The general structure is always the same
however, as summarized by Burgeff (1909) : The mycorrhizal
fungus enters through hairs into the most external cells of the mycor-
rhizome and penetrates the cortex even to the endodermis, dissolving
whatever starch is present as it goes. Then the hypha coils within the
cell, and the cell plasm digests it, the undigested remainder being sur-
rounded by a membrane that excludes it from the living portion. A
few hyphae, in many species of orchid, grow out of the rhizome again
and form spores. In German orchids. Ad. Fuchs (1924) found di-
gestion occurring chiefly from autumn into winter. Penetration of the
fungus is accompanied by solution of the starch in the plant cells. The
fungus follows the concentration gradient and only in such places
as react to the fungal passage. The so-called protein hyphae contain
an evident preponderance of glycogen, and the designation of protein
in connection with them is an error. As a result of living in a region
poor in N and P, the orchid undergoes modification (Fuchs &
ZiEGENSPECK, 1925) : There is an early cessation of root-develop-
ment ; the rhizome swells and takes over the root function as the roots
dwindle and disappear. There is lessening of the water intake, a
crumpling of the habdrome while the leptome is kept open.

In Thismia americana, Pfeiffer (1914) found underground struc-

Lecture VII — 101 — Mycothalli and Mycorrhizomes

tures which appeared like rhizomes with secondary branches, inhabited
by an endophyte just beneath the epidermis; and there were finer
hyphae internally in the cortex.

Mycocaryopses and Infection of Aerial Organs: — Since this
book is devoted to endophytic roots and infected rooting struc-
tures, it is not deemed advisable to enter into a discussion of
endophytic infection of other structures. Yet it is established that
mycotrophy exists in such plants as Lolium, a grass in which the
fungus lives symbiotically with the immature sporophyte. The "seeds"
of Lolium, which are strictly fruit and seed together and technically
known as "caryopses", harbour a fungus which is said to have been
discovered by Vogl in 1897. Described by Guerin, by Hanausek,
and by Nestler, in 1898 and by Hiltner in 1899, and by well-nigh
a dozen investigators since, the fact of endophytic infection of
L. tementulum is well established. Not only does it occur in recent
specimens of this grass but Lindau in 1904 described similar infec-
tion from grains recovered from an Egyptian tomb about 4,000 years
old. But other grasses, according to March al (1902), lack such
infection.

A detailed study of Lolium was made by McLennan (1920), who
described intracellular infection in the aleurone layer, hyphae pene-
trating also the scutellum wherever the two were in contact. Fungus
is present in embryo sac at or immediately after fertilisation, and the
ovum is infected before any divisions have taken place in it. Hyphae
sometimes extend from base of ovary into staminal filaments where
they become peculiarly knotted. In development of the embryo it is
seen that endosperm is formed by an "endospermic cambium", and
"if the fungus does not keep pace with the absorbing power of the
endosperm, no hyphal layer is formed in the ripe grain, but hyphae
can then be found in the scutellum and embryo". Endospermic cam-
bium persists as the aleurone layer, which receives a supply of nutrient
from the fungal system. McLennan concludes that "the association
of the fungus with Lolium tementulum and L. perenne is probably a
well-marked case of symbiosis, comparable in many respects with that
met with in Calluna vulgaris". She also says : L. perenne is unable
to fix nitrogen in the total absence of external supplies of combined
nitrogen."

Rayner (1915) had described a constant infection of seed-coats of
Calluna but stated that the embryo is never infected, a mycorrhizal
infection resulting by infection of the plantlet from the seed-coats.
Rayner asserted that the fungus grows through aerial organs of
Calluna but Freisleben (1934) decided that infection in this plant

Kelley ' —102— Mycotrophy

is not as general as thought by Rayner. Lewis (1924) described
stem infection of two other ericads and also of Picea and Larix.
Barrows (1941) found the endophyte of Epigaea widely distributed
in stem, flower, pollen, young ovules and on ripened fruit and seed;
and BosE (1943) reports a similar condition for Casuarina in India,
infection of seedlings occurring from the seed-coat.

In Lolium there is also infection of aerial portions. Neill (1940)
described an endophyte in the leaves, while Freeman (1904) said
that all organs of the plant except the pollen may be infected.

LECTURE Vin
MYCODOMATIA

Significance of the Term: — Literally, the word "mycodomatium"
means "fungus-chamber". Frank (1891) said that on the basis of
nutritional physiology, endotrophic mycorrhizae and root nodules may
be considered together : their morphological differences will be taken
care of if we call one "endotrophic mycorrhizae" while nodules of
alder and legumes are called "Pilzkammer" or mycodomatia. But as
nodules of legumes contain bacteria rather than fungi, we prefer to
limit the term mycodomatia to those hypertrophied structures found
on Alnus and a number of other plants which are caused in whole or
in part by fungi. In using the word "mycodomatium" in the essential
sense given it by Frank, we realize that we are not following the
original meaning as used by Lundstrom in 1887. Melin (1936)
has revived the term in its proper sense and applied it to all myco-
trophic structures, those in which the symbionts "in einem Verhaltnis
gegenseitiger Forderung stehen". Perhaps we should follow Melin's
lead, but we encounter two difficulties: {1) It is very uncertain that
all — or any — of the mycotrophic symbioses are true mutualisms, and
it would be extremely hard to sort them out in classes of mutualist
and non-mutualist. (2) If we gave up the word "mycodomatium",
we would have no term to apply to those hypertrophied structures
known as nodules, excrescenses, tubers, tubercles, etc. The term "my-
cocecidium" has been applied to them, but this term refers to galls and
is generally understood to refer to a parasitic structure. Perhaps
Frank's "Pilzkammer" should be the term used !*

All leguminous nodules caused by bacteria are ruled out of our
study. Known from the days of duHamel duMonceau in the middle
of the 18th century, nodules of legumes have attracted much attention
and their nutritional processes are of related interest to those of my-
codomatia. It is true that endophytic infection of leguminous roots
is widespread but vesicular-arbuscular mycorrhizae are not nodules.

In the present state of our knowledge it is impossible to state
positively the exact character of mycodomatia ; but it is evident that —
regardless of how they are formed — ^they are outgrowths of a larger
sort than mycorrhizae. The latter are swollen side-branches ("short-

*Baas Becking has called attention to the mistake of making a false analogy
between symbiosis in leguminous nodules and the symbioses occurring in leaf-
nodules and mycorrhizae {cf. Naturwet. Tijdschr. v. Nederl. Ind. 102:120,
1946).

Kelley — 104 — Mycotrophy

roots") while mycodomatia are often enlargements of the larger
("mother") root, either of the root as a whole or in part, or of a sub-
terranean stem. Smaller excrescences are termed tubers, tubercles,
bulbs, etc., although the actual differences between them are not great
unless we consider the former as primarily lateral outgrowths. As to
the exact nature of nodules, there is obvious disagreement, but per-
haps it will be found that they are chiefly the result of a bacterial
stimulus and the fungi which are often if not always associated may
be associate commensals, if such an expression may be used. Again, the
exact nature of tubers, etc., is not altogether clear, but it seems that
they are ordinarily called forth by a fungal stimulus, although other
influences acting on the cell-plasm may equally well produce the tuber.
Of course a tuber is strictly a stem (as in tuber of potato) and bears
scale-leaves ; but the word is also freely applied to tuberous roots as
in the dahlia, and in both cases the enlarged growth appears due to
a fungus, hence the justice of the term, mycodomatium. In orchids,
the mycodomatia are sometimes true tubers, as in Aplectrum or
Tipularia, or are tubercles or enlarged roots. Bulbs and corms are
apparently also to be classed as mycodomatia.

Psilotum: — Our information on Psilotiim infection is meagre,
but it is certain that Gallaud (1905) listed this plant in "Series 4"
with the orchids, and said it was similar in its mycotrophy with Tamus.
Psilotum is an "humus saprophyte" in which rootage organs are much
branched rhizomes that bear small gemmae on the subterranean shoots.
Yet Psilotum can grow asymbiotically as discovered by Bernard and
reaffirmed by Costantin (1925, 1936), who said that the plant had
been grown asymbiotically at the Museum at Paris for 132 years.

Gycads: — The root tubercles of Cycas, according to Bottomley
(1907), are morphologically lateral roots showing a central vascular
cylinder with a well-marked endodermis completely surrounded by
"bacteroid" tissue. These tubercles are dichotomous and perennial,
and they differ from leguminous nodules, which are of limited growth.
Kellerman in 1910 isolated N-fixing bacteria from Cycas nodules,
while Life (1901) found in C. revoluta both bacteria and hyphae of
a fungus which resembled Rhizohium. These organisms, he said, are
confined to the mediocortex. Life thought the functions of the
tubercles were aeration and N-assimilation but decided it is difficult
to speak with certainty of the symbiotic relations of the various organ-
isms within. Spratt (1915) said that the tubercles are formed pri-
marily by Bacillus radicicola, and noted that in them four symbionts
are concerned, — two bacteria, an alga and the cycad.

Lecture VIII — 105 — Mycodomatia

Podocarpus: — Nodules on Podocarpus chinensis were noted as
early as 1893 by Kellner, and were described in 1896 by v.Tubeuf.
They were examined in detail by Nobbe & Hiltner (1898), who
found a fungus growing throughout the root, and forming nodules
from within the roots, hence they concluded that the nodules or myco-
domatia are true endotrophic mycorrhizae. Plants were grown in
pure quartz sand for 5 years and watered with non-nitrogenous cul-
ture solution, the plants growing luxuriantly and presumably securing
N from the air through the mycodomatia. Later, Hiltner (1903)
said that N-fixation was shown for Podocarpus but not with the same
certainty as with Alnus. Kondo (1931) also wrote of N-relations of
these plants but his work is hidden in the Japanese language.

Processes occurring within the Podocarpus mycodomatium were
described by Shibata (1902), who found a large hyphomycete which
by branching filled the whole cell : the host nucleus increased in volume
and assumed amoeboid form, dividing amitotically until as many as
8 nuclei are formed which become distributed in the cell, then becom-
ing amoeboid once more. When the fungus has attained its full growth
it is digested by the host-cell and the nuclei may then resume their
normal condition and divide mitotically. A proteolytic enzyme capable
of digesting fibrin was demonstrated in the tubercle. Shibata corro-
borated Magnus and Frank, that the fungus is subservient to the
host-cell. Hiltner (1903) said that the host-cell digests and absorbs
not only the plasm but the chitinous wall of the fungus ; and he also
noted nuclear activity in these cells. "Blasen" in Podocarpus, he said,
are equivalent to Janse's sporangioles, being partly formed and then
digested. McLuckie (1923) described nodules of P. spinidosa and
P. elata as of dual character. He said that these species, like other
species of the Podocarpineae, are actively engaged in N-fixation by
virtue of bacteria present in cortical cells. The nodules are modified
lateral roots and arise from the pericycle, their normal growth being
checked before they emerge from the cortex of the main root. Root-
hairs, he said, are commonly present. Nodules and cortex of main
root frequently contain fungal hyphae and peculiar spore-like bodies
belonging to the fungus ; the surface of the nodule and main root is
frequently invested with a loose tangle of fungal hyphae, some of
which enter the root tissues. The nodules of P. chinensis and P. nubi-
gena are occupied by a fungus (Schaede, 1943) which is considered
a harmless parasite since it has so slight a connection with the soil ;
but the arbuscular structure is markedly developed.

Gasuarina: — Nodules filled with a gummy mass were found by
Janse (1897) on C. muricata in Java. Kamerling (1911) described

Kelley — 106 — Mycotrophy

Casuarina nodules from coral islands in the bay of Batavia, and said
that in section it is seen the nodules possess a small central strand while
larger or smaller groups of cortical cells are filled with protein-reacting
bodies resembling leguminous bacteroids. Kamerling supposed that
the nodules were responsible for N-fixation, and the same function
was ascribed to them by Adinarayana (1924), Mowry (1933), and
Narashimhan (1918), the last isolating bacteria that fixed N.
MiEHE (1918) tacitly inferred the same function, and asserted that
these nodules are mycodomatia, the symbiont being a small hyphal
fungus which heavily infests the cortex, passing directly from cell to
cell but never invading the vascular bundles.

Myrica: — Brunchorst in 1887 had mentioned tubercles in M.
Gale which were described by Bottomley (1912) as modified lateral
roots. Three branches arise from the end of each primary nodule and
afterwards the stele grows out through the apex of the nodule into a
hair-like root. In each mature nodule four regions may be recognized,
vis.: apical meristem, "infection thread" area, "bacterial zone" which
includes most of the cortex, and basal zone devoid of bacteria but
with the cells containing oil drops. At maturity the bacteria disappear
and basal zone encroaches until it finally replaces all the others. In
old nodules, filling a majority of cortical cells and sometimes the base
of young nodules, mycorrhizal fungi are found. Fungal hyphae, said
Bottomley, were earlier thought to be responsible for nodule forma-
tion and it is possible that they may be of mycorrhizal nature and of
benefit to- the Myrica plant. Bottomley caused nodules to develop
by inoculation; he also showed fixation of free N. Schaede (1938)
considered the causal organism of this Myrica to be Actinomyces and
he gives a well illustrated account of the infection.

In M. rubra, Shibata (1902) found the "fungus" (which he
believed to be Actinomyces) confined to a definite "ring" in the cortex.
In M. carolinensis, the author found beneath a cuticularized epidermis
a cortex of 10-11 layers of rounded cells, larger internally and full of
protoplasmic content. A zone of 2-3 layers commencing about the
fourth from the outside of cortex is a "bacterial layer" containing
comparatively large "rods" which are densely clustered and deeply
stained. About the outside of the domatium there is more or less a
weft of branched septate and geniculate hyphae, dark in colour. It is
difficult to demonstrate infection but neverheless in the outer cortex
there is the appearance of intracellular hyphae ; while many of the
cells have content suggestive of partially digested protoplasm which
takes a reddish stain while the bacteria stain blue. The latter divide
transversely to form rosettes. Under oil immersion, strands can be

Lecture VIII — 107 — Mycodomatia

seen connecting adjacent "masses" through cell-walls, tangentially.
The cell nucleus is not hypertrophied. Rosette formation by Actino-
myces is mentioned by Youngken in a study published in 1915 on
the Myricaceae ; and he said that this organism later penetrates
tracheae and grows out into the seed. In "M. cerifera," Harsh berger
(1903) described mycodomatia inhabited by Frankia. He supposed
that these structures were intermediate between ectotrophic mycor-
rhizae as in Monotropa and endotrophic forms as in Thismia.

Alnus: — In alders occur the earliest described root excrescences
which may be termed mycodomatia. Meyen in 1829 gave the first
description of tubercles in alder (so far as we are aware), and con-
sidered them as "pseudomorphosed roots" in the ends of which there is
a parasitic growth comparable to that of Lathrea, etc. Meyen was sure
that he had shown them to be "ganz vollkommene parasitische Ge-
bilde" and that they were formed by "gleich anderen vollkommenen
Organismen". Since then there have been numerous other descrip-
tions given but their exact nature and symbionts still remain unsettled.
Apparently alder nodules are not caused by one organism nor do they
always have the same physiology, for some investigators describe
them as bacterial, others as fungal. Thus Cernik (1937) lists the
"fungi" of alder nodules as Frankia, Schinsia, and Actinomyces, all
of which are presumably bacterial; while Pieschel (1929) cites
Lactarius lilacinus and L. cyathula as always associated with alder
and presumably the mycodomatial symbionts, with Gyrodon rubescens
a probable third symbiont. Two investigators report synthetic my-
codomatia for alder, — Plotho (1941) and Roberg (1938). Roberg
grew seedlings of four species of alder in a synthetic nutrient solution
with a suspension of ground root-nodules isolated from each of the
species. Only healthy seedlings reacted to inoculation by nodule
production; and in all cases the symbiont was Actinomyces alni. Be-
cause of the frequent presence of this organism, Shibata (1902)
termed the alder mycodomatia cases of vegetable actinomycoses.

A number of papers describe nutritional processes of alder my-
codomatia. Shibata, already mentioned, tells of the "blaschen" or
small bodies formed by the "fungus" in mycodomatial cells and their
subsequent digestion ; he also described clots which contained, besides
some fungal hyphae, a number of little rounded drop-like or oval
structures which he termed "sekretkorper". But Zach (1908) did
not find these bodies in A. glutinosa (Shibata worked with A.
japonica), but considered the broken threads or "Stabchen" of Shi-
bata as concentrated cell-content of the hyphae while spore-like knots
and bacteria-like threads are degenerate forms of hyphae which, he

Kelley — 108 — Mycotrophy

claims, absorb much water and fill the entire cell lumen. Terminal
swellings of the hyphae are also degeneration stages of the fungus
which are ultimately digested by the host-cell, during which process
the fungal masses pass through various degenerative stages. Spherical,
oval and other shaped bodies of an oily consistency appear during the
digestive process and to these he applied the name "Exkretkorper".
Shibata had described a proteolytic enzyme from alder mycodomatia.
The author has seen numerous yellow clots in outer cortical cells of
Alnus riigosa, which alder has coralloid mycorrhizae in addition to
mycodomatia. Klecka & Vukolov describe fungal digestion in alder
and other nodules and regard the fungus as provider of starch and
protein. Hiltner (1896) claimed that alder nodules assimilate free
nitrogen; and he also found that CaCOg stops their growth. Borm
(1931) said that in Alnus it has been found possible to prove that the
bacteria fix N, but that it is not certain the nodules formed only by
fungi can perform this process.

Polygonum: — Ectotrophic mycorrhizae are constant in P. vivi-
parum, not only in the countless adventive roots but in the bulblets
(Hesselman, 1900), which must then be considered as mycodomatia.

Raphanus: — Molliard (1920) stated that radish produces
tubers perfectly well under sterile conditions when supplied with
sugar and CO2 in sufficient quantity. The presumption is that in
nature radishes are "fungus-chambers" called forth by infection.

Tribulus: — In sandy places of the Gov. Cherson in Russia, among
dry arid sand vegetation, Issatschenko (1913) found fleshy green
specimens of T. terrestris that bore nodules on their roots, — small
white ones on thin roots and larger dark nodules that recalled legu-
minous nodules. In section, dark septate hyphae were evident, cloth-
ing outside of the nodule and penetrating into it in places while within,
the hyphae were thinner and lighter in colour, and proceeded from
cell to cell. Disappearance of starch from the nodules was observed.
Issatschenko thought that these mycodomatia were true mycorrhizae
and agreed with Bernard that using of the starch increases osmosis of
the cell and with it the water intake.

Legumes : — In addition to bacterial nodules and endotrophic m}'-
corrhizae, legumes possess mycodomatia. Janse (1897) described
fungal nodules in Pithecolobium montanum, a member of the Mimo-
saceae: the cortex contains 2 layers of tannin cells separated by 2
layers of parenchyma and in the latter the fungus is found, but it never

Lecture VIII —109— Mycodomatia

enters tannin cells. The nodules recall those of Casuarina. Fungal in-
vasion of Orobus was discovered by Frank (1879), and he figured
arbuscles and vesicular swellings in the nodules of O. vermis and
O. tuherosus; and Frank considered that the hyphae were trans-
formed into "Sproszellchen".

Ailanthus: — In the Erlangen Botanic Garden, Andreae (1894)
found that sturdy side-roots of Ailanthus had irregular tuberous out-
growths of 5-40 mm. diameter placed directly on the root cylinder and
composed of smaller bodies in a grape-like cluster. Their structure
was thought due to the higher plant and not induced by the fungi
(mostly Pyrenomycetes) found in the nodules. Further studies are
awaited on these structures.

Geanothus: — Nodules were noted on C. americanus by Beal in
1890 and were described by Atkinson (1892). While resembling
in form leguminous nodules, he found the causal organism was a
"fungus" which he named Frankia ceanothi. Material was collected
from Alabama and Michigan, and similar nodules were found on
Alnus serrulata. In a more extended study, Arzberger (1910) said
that infection is through epidermis or root-hair and the mycodomatium
consists of 3 systems of tissues : an outer corky layer, a middle cortical
tissue, and an inner vascular bundle. In the cortical layer are infested,
hypertrophied cells, the nuclei being enlarged. He noted also three
stages in development of the fungus, — mycelial, "sporange" and
digested. No "Exkretkorper", as described by Zach, were found, but
an enzyme capable of digesting fibrin was found. He said that sym-
biosis exists but both host-cell and fungus dies. A very different de-
scription was given by Bottomley (1915), who considered the
nodules purely bacterial, formed by bacteria of the Bacillus radicicola
group. As no nodules were formed on C. americanus in England, he
imported material from America, securing nodules also of C. velu-
tinus. The bacteria, when isolated, grew in pure culture and fixed N.

Elaeagnus: — The same discrepancies must be noted in descrip-
tions of Elaeagnus nodules. Brunchorst and Frank agreed at first
for their fungal nature; then Frank (1887) withdrew to the posi-
tion that the nodules are merely reserve organs, containing no sym-
biont. Zach (1908) described them as fungal and similar to those of
Alnus; Nobbe (1892) believed that he had demonstrated N-fixation
with them. On the other hand, Spratt (1912) identified the causal
organism as Pseudomonas radicicola, but stated that it does fix free
N. The author found coralloid mycorrhizae but no nodules on Shep-
herdia argentea in northern Minnesota.

Kelley — 110— Mycotrophy

Hippophae:— In 1887 Brunchorst spoke of the "well-known"
nodules of Hippophae and considered their possible action in fixation
of free N. These nodules were rediscovered by Arcularius (1928)
who gives a detailed description of their structure and ascribes their
formation to a fungus. There is no infection of the vegetative apex
nor is starch present in cells infested by the fungus, though abundant
elsewhere. The fungal hyphae swell near the cell-nucleus and form
"little heads" which gradually swell with a fine, deeply-staining mate-
rial. This material is then apparently digested and finally disappears,
with coincident nuclear changes. The relation of fungus to host is
not obligate, and the author supposed that soil must be inoculated in
order to have mycodomatia produced. But Borm (1931) ascribes the
nodules to bacteria which, he said, multiply in an enormous way until
they fill the whole cell (the nucleus remaining intact) and then diges-
tion occurs.

Gorlaria: — In C. japonica, Katakoa (1930) ascribed N-fixation
to the nodules, saying that plants with nodules make vigorous growth
while without them growth is retarded. Shibata (1917) said that the
endophyte, a typical Actinomycete, forms a rich weft separate from
the cortical tissue, the colonies of which in the host-cells have consecu-
tive partitions with centripetal pectinate hyphae arranged club-wise
about the vacuoles which are filled with cell-sap. The root-nodules of
Coriaria in respect to anatomical dififerences surpass all others and
its characteristic symbiosis-tissue is quite similar to that of legumes.

Eucalyptus: — According to Dufrenoy (1922), swellings are
found on axes of young Eucalyptus plants, of which the origin is
unknown.

Daucus: — Sterile achenes of carrot were germinated in a mineral
gelatine with some addition of glucose (5.0-7.5 parts per 100) and
plants were grown in large tubes plugged with cotton; but with
glucose there was poor growth. In sterile humus soil with addition
of mineral salts, in tubes plugged with rubber stoppers, the air had
5 parts per 100 of COg, and the plants grew well, in 40 days forming
a tuber 1 cm. in diameter (Molliard, 1920).

Ericads:— VON Tubeuf (1903) said that on the Chiemsee Moors
at Bernau in Bavaria, the largest of the Vaccineae is V. uliginosum,
which has its root system deeply sunk in sphagnum. If whole stocks
are drawn out of the sphagnum, a portion of the attached roots is
obtained with a tender root-work ; while on thin rootlets are found

Lecture VIII — 111 — Mycodomatia

copious nodules of various sizes and forms which appear as little
clubs although they may have a very delicate continuation as a thin
rootlet. In section they are seen to have a central cylinder with pro-
nounced water tissue and peripherally a normal cortical tissue, von
TuBEUF thought that neither fungi nor bacteria are responsible for
these structures. He found them on all woody plants of the moor
except pine, that is, on six ericads including Calluna and Andromeda.

But for Arbutus, true mycodomatia caused by a fungus are de-
scribed by RiVETT (1924) and by Dufrenoy (1917). Inoculated
rootlets developed into small pear-shaped tubercles, said Dufrenoy,
on which nearly all the epidermal cells develop into root-hairs, around
which algae and bacteria collect and form a mucous. The fungus in-
vades external layers of cortex which stores large quantities of re-
serve material as "tannin" while medullary tract and rays are crowded
with starch grains. Rivett, in describing the endophyte in old tu-
bercles, said that infection by fungus keeps pace with production of
new cells by growing point, and digestion and reinfection proceed
successively. Thus peripheral cells, except at growing points, are to
be found filled with partially digested hyphae. Digestion proceeds
all the time that tubercles are growing and even in winter it is hard
to find clearly defined hyphae. In a great majority of the cells cavities
are filled with a granular mass of deeply staining material in midst
of which persists a large nucleus. Endodermal sheath becomes densely
filled with reserve, and the conducting tissue itself becomes blocked
with deeply staining material. Tubercles persist in this condition
throughout winter and early spring.

Pyrola rotundifolia possesses tubers formed by inordinate radial
increase in size of epidermal cells as result of fungal infection. At
first the hyphae are intercellular but later they penetrate the cells and
fill them ; the nuclei become hypertrophied and then disappear. A
mantle is finally formed about the root (Kramai', 1899).

Solanum: — Tuber formation of the potato, according to Ber-
nard's early work (1901) is called forth by an endophytic fungus,
Fusarhwi solani (later called Rhisoctonia solani). In pure culture
with the fungus, tubers were freely produced while in soil that was
little infested tubers were sparse. He said that, according to state-
ments made by de l'Ecluse in 1601, when potato seed was first
planted in Europe, flowering but not tuber-forming plants were pro-
duced, so that to secure a crop of tubers, older tubers rather than seed
had to be planted. Today, plants grown from seed produce tubers the
first year because with general cultivation of the potato the fungus is
widely distributed in the soil. Bernard noted still further (1902a)

Kelley — 112 — Mycotrophy

that tuber formation is not dependent on the fungus per se but to a
certain sap concentration, for cuttings of potato plants placed in
aqueous sugar solutions produced tubers. This discovery he at-
tributed to Marchal. a critical concentration exists for each plant ;
and Bernard thought that tuberisation in all cases depends directly
upon a certain degree of concentration of cell sap. But ordinarily
the habitual provider of this sap concentration is a fungal parasite
{\902b), which produces the optimum concentration for diastasic
ferments. Bernard grew the fungus, Fusarimn solani, in a macera-
tion of potato sap and found it increased the sap concentration as
indicated by a lowering of the freezing point.

That tuber formation is connected with sap phenomena was in-
dicated further by the work of Rolfs (1901), who found that small
tubers were formed on the stem when a stricture was placed about the
stem, either by the fungus or by artificial girdling, and the sap was
prevented from flowing to the region of tuber formation. It may be
noted incidentally that presence of glucose in concentrations of 1/100
to 1/10 mol. is termed essential to cell division and elongation in wheat
roots (Burstrom, 1941). But Molliard (1915) found that even
when the plant is placed in a sugar solution there was no tuber forma-
tion until gaseous interchange (which increased sugar absorption) was
suppressed. Magrou finds that normal tuberisation may be obtained
glucose (also glycerine), in combination with action of light (Ann.
d. Sci. Nat., Bot., XI, 5:135-136, 1944).

Believing that tuber formation is always induced by fungi, Bernard
(1911) investigated other plants and found that 6". Dulcamara and
S. Maglia (the latter from Chile) also contained an endophyte. Janse
(1897) had found the same for S. verbascifolium in Java. Magrou
(1914) found furthermore that the endophyte of S. Dulcamara could
induce tuber-formation in S. tuberosum, hence there is no necessarily
specific endophyte ; and this observation was confirmed by Costantin
(1935). Yet under cultivation the endophytic fungus is lost and the
potato plant produces tubers without it (Magrou, 1921; Castan,
1941) ; and the suggestion is made that dunging destroys the fungus, —
which still lives in the wild form, ^S. Maglia of Chile . With further
study it was concluded that tuber formation in the potato is an "ac-
quired habit" of the plant in cold climates, the climate having the same
sort of action in tuber formation as the fungus ; for the potato in
cold climates, either in high latitudes or in high altitudes, produces
tubers normally whereas in warm climates this power is lost (Costan-
tin, 1922). Potatoes grown at 1400 m. produces more tubers than
those at 560 m. (Costantin, 1935a), a result confirming Lebard
& Magrou ( 1935), who found that there is an altitude where the yield

Lecture VIII — 113 — Mycodomatia

is maximum. Miege (1936) found that refrigeration for not longer
than 4-5 months restored vitality to the potato quite as well as a change
in altitude. Loss of the endophyte in this fashion explains Jumelle's
(1905) problem of why the isolated Fusariiim seemed to have little
importance in tuberisation of 5". tuberosum and 6". Commersonii. In-
deed, Castan (1941) concluded that a symbiotic fungus is not neces-
sary to tuberisation, at least at low altitudes.

These statements were modified somewhat by Costantin (1936).
Thus, while tubers of cultivated potato do not contain symbiotic
fungi, certain varieties contain mycorrhizal fungi just as the wild
forms. Furthermore, while infestation is usually abundant, it may
be sparse or completely lacking in certain individuals. Again,
in late summer at high altitudes, small ("microscopic") tubers
were formed in conjunction with symbiotic fungi that, left in the soil,
spontaneously reproduce the plant ; but at lower altitudes the sym-
biotic fungi are lacking and the tubers perish during the winter. This
action was confirmed by Joseph (1935), who notes also that "micro-
scopic" tubers dilTer in colour.

Melampyrum: — M. prate use utilizes the humus of the moss, or
grass, tussocks in which it lives through delicate protuberances or ab-
sorptive organs produced from the roots. These protuberances were
found actually growing intO' dead objects (Koch, 1887).

Orobanche: — Henfrey (1849) suggested that the whole tuberous
base of the plant is concerned in absorption, just as in orchids. Further
studies of mycotrophy in this plant are awaited.

Composites: — Molliard (1920) has been mentioned already for
his work on radish and carrot : under similar sterile conditions he was
able to induce tuber-formation in Dahlia, that is, under optimum con-
ditions of sugar and COo supply. Swollen adventive roots were formed
within 6 weeks, and Molliard concluded that "under favourable
conditions" micro-organisms are not necessary to tuber formation, —
although it is not explained how plants in nature are to secure flasks,
sugar solutions and rubber stoppers as substitutes for the aforemen-
tioned endophytes. An actinomycosis is described by Dufrenoy
(1920) for Adenostyles, but he does not state definitely that there is
an enlargement of the tissues.

Juncus: — Tubers on Jimciis were mentioned by Chatin in 1856,
also by Cameron in 1886, who found root-swellings likewise on
Ruppia maritinia, R. rostella and Eriophorum vaginatiim. Weber

Kelley — 114 — Mycotrophy

(1884) made a detailed study of various J uncus plantlets and found
them inhabited by a fungus and swollen into tubers the size of which
depends on activity of the rush. The fungus is present only in radially
enlarged cells of the periblem where it forms coils of reagent-resisting
hyphae, and surrounds the nucleus; or hyphae penetrate to other
cells. As in rust fungi, the hyphae are surrounded by a cellulose layer
that is continuous with the membrane of the penetrated cell. Spores
are formed by the fungus, which assume a barrel-shape and become
surrounded with a thick dark wall. In winter, the mass of the former
tuber in wet earth is full of ochre-yellow spores which germinate
naturally in February. The fungus is considered to be Entorrhisa
cypericola, placed in the Tilletiaceae. Grutter said that in /. Tenageia
the fungus encloses tip of root, penetrates epidermis and forms
special structures in it. The stele is much reduced. Lagerheim
(1888) described E. digitata from /. articulatus in Switzerland. The
roots were deformed into root galls and contained an abundance of
yellow spores, and the fungus was extracted with difficulty. In the
Black Forest, /. articulatus bore mycodomatia in very sandy and not
too wet soil but they were absent from moor and loam soils, occurring
in the uppermost soil horizon.

Molinia: — This grass forms a "molinetum" on sterile sands of
northern Germany and elsewhere, its rhizomes and interlaced roots
acting as sand-binders. It overwinters as swollen basal nodes while
the roots are endotrophic, never ectotrophic. A line drawing indicates
fungal coils in inner cortical cells and a possible "sporangium", per-
haps a vesicle. Data are presented on N content of tuberous rhizomes
and seeds. Plants were grown for three months in culture solution
and sand, and one plant at the end of the experiment was found with
fungus-free root system, while its rhizome-base was filled with starch.
Hence grasses must be examined in considerable numbers to determine
true extent of infection for some, like Molinia, may be facultatively
mycotrophic (von Tubeuf, 1903).

Gyperus: — Magnus (1879) described a fungus, Schinsia cyper-
icola, living in roots of C. flavescens. Through its activity the root
swells into a simple tuber or, if the roots branch, into a branched
tuberous body. In Schoenus ferrugineus there are mycodomatia
containing normal fungal hyphae (Renner, 1935).

Asparagus: — Root nodules of Asparagus have been described in
Japanese by Fujita (1940).

Lecture VIII — 115 — Mycodomatia

Allium: — In an extended study of A. roseum, Capelletti (1931)
isolated a fungal endophyte which he referred to Rhizoctonia.

Orchids: — Tuber formation of orchids was described by Fabre
in 1855 but interest in its significance dates from Bernard (1902).
The latter found, as in potato, the causal fungus is Fusarium {Rhizoc-
tonia), and that tuber formation takes place very early in develop-
ment of the plant ; yet the tuber itself, at least the parenchymatous in-
terior, remains fungus-free. Beau (1914) also said that in the adult
plant, tuberisation may take place without fungal invasion. Bernard
found a retardation of development in the orchid plant which goes
hand in hand with nodule formation and storing of food-stuff, and to
him it seemed the result of a sort of poisoning caused by the endo-
phyte. So long as the plant is free from infection there is active growth
of leaves, flowers and fruits ; but tubers are formed only after en-
trance of fungus. Later (1909), Bernard classified orchids as
facultatively mycotrophic (as in epiphytic members) and constant
mycotrophs. According to Burgeff (1910), the fungus is found in
roots and protocorms of almost all orchid plants. Gallaud (1905)
classified orchid tubers in Series Four, and stated that the endophyte
is intracellular and produces coils (pelotons) which sometimes remain
inactive (host-cells) or are digested (digestion cells). A curious
condition was described by Barsali (1921) in which two horizontal
tubers are formed in addition to the ordinary ones, the former tubers
supposedly making use of humus of the top-soil.

LECTURE IX
STRUCTURE OF MYCORRHIZAE

The Kinds of Mycorrhizae: — With insight which characterized
his work, Frank early stated that there are two principal sorts of
mycorrhizae, the coralloid sort found with forest trees, and the endo-
trophic which he illustrated from ericads. That general distinction
into basidio- and phycomycete types is found to hold generally good.
Yet it must be remembered that mycorrhizae are formed primarily
by the higher symbiont and that their form is determined by the vas-
cular plant producing it: the fungus is of secondary significance. One
fungus or another, or several fungi together, may invade the root,
but the mycorrhizal form will be essentially the same in all cases : its
form is characteristic for the higher symbiont rather than for the
fungus. This fact is emphasized by Woodroof (1933) whO' says:
". . . it is seen that the influence of the fungus in the gross morphology
of mycorrhizal roots is slight. The presence of the fungus is to all
outward appearances merely incidental."

Mycorrhizal Compared with Non-Mycorrhizal Roots: —

Not all the rootlets of a mycorrhizal plant are necessarily mycorrhizal,
and a distinction must be made in the case of woody plants between
long-roots or "roots of extension" that grow rapidly through the soil,
and short-roots or small laterals which serve principally for intake of
materials from the soil. Woodroof (I.e.) calls attention to the fact
that not all short-roots are infected and that short-roots are not
started by the endophyte but, being formed, are invaded. The struc-
ture, whether invaded or not, is the same in both cases as regards gross
morphology. Long-roots are considered to be fungus-free, and when
short-roots are likewise uninfected they usually bear root-hairs ; but if
infected by symbiotic fungi, the short-roots become shortened and
swollen in their development. Yet Hatch (1937) presents some
evidence to indicate that mycorrhizal fungi stimulate their growth
and thereby increase the absorbing surface areas. The rootlet that
bears mycorrhizae is called a mother-root (following Noelle, 1910) :
the mother-root may renew its apical growth and extend out into
the soil as a pioneer-root. To illustrate, a simple mycorrhiza is a
mother-root bearing a very few elongate laterals ; a coralloid mycor-

Lecture IX — 117— Structure

rhiza is a mother-root plus small coral-branched short-roots. But
in the case of annual plants or biennials, any of the secondary roots
may be infected, or even the adventitious roots ; while in several plants
with aerial roots, these are turned into mycorrhizae.

External Form: — The form of a mycorrhiza is characteristic
for each species of plant and generally the form is constant for a
genus and even for a family. Thus, all cupulifers have a coralloid
sort of mycorrhiza ; Jitglans have simple, and Acer have necklace-
beaded mycorrhizae. Yet it must be noted that Melin (1925) states
that mycorrhizal form is greatly influenced by the sort of salts present
in the soil. The least complex form is the simple mycorrhiza which
consists of elongate monopodia! rootlets such as occur in Liriodendron,
Coriuis and Fraxinus. As described by Melin from pine heaths,
simple mycorrhizae may grow to 10 mm. length and 0.2 mm. diameter,
and are ordinarily without root-hairs. Apparent root-hairs on simple
mycorrhizae may on inspection turn out to be fungal setae, which often
simulate epidermal outgrowths.

The coralloid mycorrhiza is said by Ulbrich (1924) to have been
first described by Hartig in 1851. It is branched freely like coral,
the "mother-root" bearing numerous short branches that, in compari-
son with the simple mycorrhiza, may grow to 1 mm. or more length
and 0.4 mm. diameter; i.e., they are short and thick. They are well
seen in pines, oaks, birches, and in the German are called "Gabel-
mykorrhizen", a shrub-like sort of structure. The racemose
mycorrhiza, as found in spruce and other forest trees, is formed by
lateral rootlets branching monopodially in two rows upon a main
axis. When coral-branches cluster thickly at one place to form a sort
of "witch's broom", the cluster is called a rhisothamnion; or, in the
German, a "Buschel". Rhizothamnia are seen on pine (Muller,
1902) and oak, and are said to be characteristic for Casuarinas
(MiEHE, 1918). Or, the cluster of short dense branches formed by
dichotomy may be weft about with mycelium to form a nodulous
lump called a tuberous mycorrhiza, or in the German, a "KnoUen-
mykorhiza". It is not truly a nodule, neither a tuber ; and is said to
have been first observed by Muller on Pinus montana.

Pearl-necklace mycorrhizae are formed in yet a different way.
They commence as ordinary racemose mycorrhizae or perhaps as
widely spaced coral-branches but through intermittent growth succes-
sive additions are made and a constriction is left between each two
additions. Thus are developed the "pearl-necklace" beads so charac-
teristic of Acer, and found in various other plants. Such mycorrhizae
may be found on Pinus virginiana when the latter grows in droughty

Kelley — 118 — Mycotrophy

soil, and doubtless in most if not all cases this sort of mycorrhiza is
associated with intermittent growth.

Pseudomycorrhizae : — Infection of a short-root by a fungus does
not necessarily result in formation of a mycorrhiza, for there are many
cases in which the infecting fungus is a parasite. Such "false mycor-
rhizae" had long been observed but were named "pseudomycorrhizae"
by Melin (1917), who observed them on pine and spruce growing
in Swedish moors. The pseudomycorrhiza is thinner and simple, or
sometimes monopodially branched, in pine ; the hyphae are intracellu-
lar and penetrate even the meristem, and must be considered para-
sitic. Melin thought that Holler's "ectotrophic mycorrhiza" was the
same as a pseudomycorrhiza. Latham, Doak & Wright (1939) said
that under field conditions most non-mycorrhizal short-roots of pine
become pseudomycorrhizae, thus reducing the absorbing surface of
the roots and their ability to take up mineral nutrients.

Pseudomycorrhizae are thin and lack the basal constriction that
marks the mycorrhiza; then, too, mycorrhizae are usually lighter
in colour than the mother root, at least when young, whereas the
pseudomycorrhiza is dark in colour.

The Colours of Mycorrhizae: — In earlier days some attention
was paid to colours of mycorrhizae: Thus, Mangin (1910) cites
Querciis with white and rose-coloured ones, Fagus with yellow and
blue. McDouGALL (1914) presented a classification of mycorrhizae
based in part on colour, vis.: bright yellow, brown, white. Masui
(1926) said there are three types of ectotrophic mycorrhizae on roots
of Alnus firma var. Sieboldiana, — white, yellow and dark. A yellow
colour of the root is characteristic of mycorrhizae of potato (Magrou,
BouGET & Segretain, 1943).

Two things influence mycorrhizal colour, viz. age, and the fungal
symbiont. In general, young mycorrhizae are light in colour, often
a pure glistening white ; and they become darker as they grow older
until they usually turn brown, although very old mycorrhizae may be
black. But a black pine mycorrhiza may split its sheath and produce
a white tip of renewed growth under favourable conditions. Or, a
black colour may be given the mycorrhiza by a fungus long known but
more recently described as Mycelium radicis nigrostrigosum, which
usually develops strands of hyphae from the surface. Other fungi may
cause other colours, as yellow, reddish or pale violet ; but as the
mycorrhiza grows older these colours tend to disappear.

Lecture IX —119— Structure

The Exterior Surface: — Phycomycete and simple mycorrhizae
are usually smooth of surface and lack visible mycelial coating, while
coralloid mycorrhizae are often shaggy with hyphae. In the latter,
when the hyphae are densely interwoven they form a mantle that is
weft as closely as a tissue and when young may have a white satiny
surface, but when older becomes "fuzzy" with free hyphal ends.
Peyronel (1922) termed this mantle a micoclena, or it would perhaps
be better written "mycoclena", Greek for fungus-mantle; while
ZiEGENSPECK (1929) Called it a mycoderm, when it is a pseudotissue.
In both the smooth and the shaggy mycorrhizae there are doubtless
numerous hyphae that extend into the soil, passing from the soil cer-
tain materials into the interior of the root. Such hyphae have been ap-
propriately called Communication-hyphae. Being delicate, they are
inevitably broken in removing the mycorrhiza from the soil and could
be observed directly, if at all, only by some "glass-plate" method.
Communication-hyphae are the "root-hairs" of a mycorrhiza.

But oftentimes the fungus produces short setose hyphae that
project evenly from surface of the mycorrhiza and simulate root-
hairs, — except that of course root-hairs are not developed from a
mycorrhiza, neither are they formed on a root-cap! Yet Gordon
(1936) describes and figures "root-hairs" over tip of a mycorrhizal
short-root. Presence of setae over apex of the rootlet or mycorrhiza
is indicative of their hyphal origin, and close examination with high-
power stereoscopic binocular microscope shows continuation of the
seta with a close-weft mantle hypha. Woodroof (1933) describes
pecan mycorrhizae with three sorts of setae on the surface, — flask-
shaped with long necks, with short necks, or stellate with intermixed
spines. Mangin (1910) also described and figured setae, as hairs
dilated at the base and tapered regularly to a point, — length 100-150
/x, diameter 5-6 jn at base.

Or, hyphae may coalesce on or near the surface of the mycorrhiza
to form strands known as rhizomorphs which are similar to, but
usually smaller than, the rhizomorphs found in soil or under bark of
dead trees.

It is quite possible that root-hairs and setose hyphae are present
simultaneously on a root, and fungal infection may be through root-
hairs; or it may occur directly through the epidermal wall. Root-
hairs are not developed, usually, to any extent on a plant provided with
mycorrhizae; yet their presence depends to a considerable extent
upon the soil in which the plant is growing, for in a forest the roots
will be mostly turned into mycorrhizae whereas in cultivated soil
root-hairs are more to be expected. Acid humus is not favourable to
growth of root-hairs, and when such are formed in so unfavourable

Kelley

120-

Mycotrophy

an environment they are apt to be crumpled, shortened and otherwise
indicative of an untoward environment. These considerations again
show the casual nature of the mycorrhizal relationship.

The Mycorrhizal Apex: — The effect of habitat on rootlet is
perhaps never better shown than in the root-cap region. In aquatic
roots where there is little resistance to apical growth, the apex may be
freely exposed ; or, if there is a root-cap as in Eichhornia, it grows to

"'^'^:^-^!:

■■^•':-r--ii,-n;<tii

Fig. 7. — Longitudinal section through apex of a mycorrhiza of Pinus
rigida. r.c., root-cap; me, meristem; m, mantle; h, Hartig net; e, endo-
dermis, filled with "tannin".

a relatively enormous size and fits loosely over the apex. Aesculiis,
grown in water culture, produces roots that "keine Wurzelhaube haben
und diese ihnen von ihrer ersten Entwicklung an fehlt". (Klein &
SzABO, 1880). In ordinary root-hair roots (as in Zea), there is a
close-fitting root-cap formed from a definite zone of proliferation in
the proximal portion of the apical meristem. Pressure of rootlet

Lecture IX —121— Structure

against soil causes the cap to fit closely while friction of growth
causes sloughing of external cells. But contrast with the mycorrhizal
apex is decided, for in the mycorrhiza the root-cap is small, densely
filled with content and is closely covered over by a mantle of firmly
weft hyphae. At first white or light-coloured, it becomes dark with
age and the apical meristem finally ceases activity. In healthy mycor-
rhizae, said Mangin (1898), the root-cap instead of being lost is
conserved although invaded by mycelial filaments; and in part the
cells are detached from each other. The only change caused by pres-
sure of the mantle is a regular hemispheric form given to the root-
cap. The next year he pointed out that the mycorrhizal root-cap is
never exfoliated but persists throughout the life of the mycorrhiza
between the mycelial mantle and the more or less hypertrophied ex-
ternal cortical cells. And, as already noted, the apex may be covered
over with "root-hairs", as described by Gordon (1936) for broad-
leaved trees, by Muller (1886) for beech, and by Zach (1909) for
Sempervivum.

The mycorrhizal apex in short-root of Corsican pine is neatly
illustrated by Aldrich-Blake in Oxford Forestry Memoirs : Details
of meristem, cap, and infected tissues are well-shown and are con-
formable to those of other species of pine. But in 1874 it was sup-
posed (by Janczewski) that gymnospermous roots have no cap nor
true epidermis although the primary cortex is exceedingly voluminous
over the root-apex, replacing the cap. MacDougal (1900) found the
root-cap little developed in Monotropa but many-layered in Sarcodes
and Pierospora: but in all cases the tip is covered with mycelial mantle
that crushes the cap-cells.

Under favourable conditions of growth, as during a rainy season
that succeeds a drought, the apical meristem may renew its activity
and again cut off cells. As a result of pressure thus set up, mantle
covering the apex is split, exposing a new mantle that has developed
beneath, and the mycorrhizal short-root continues its growth in length.

Renewed Growth: — This is such a common phenomenon it is
strange that it is not more generally remarked. Masui (1926) devoted
a paper to the subject, finding that "the mantle-clad root-apex of the
completed mycorrhiza in Abies firma (Form A and Form B), Abies
Mayriana, Alnns japonica and Finns densiflora can renew its growth
breaking through the mantle." The mantle splits in various directions,
hyphae grow out from the split and later cover the exposed root-tip.
Or, the quiescent meristem renews its growth after bark and mantle
have been sloughed off.

Kelley

— 122-

Mycotrophy

Normal renewal of growth in mycorrhizae of Taxus was described
and figured by Prat (1926): "'Mamelons" formed during one
season are quiescent over winter but "Au printemps, les cellules du
meristeme central se remettent a proliferer, formant un massif qui
traverse les assises externes comme fait une radicelle pour les tissus
de la racine-mere. A I'interieur de ce massif apparait un nouvel
endoderme qui se raccorde a I'ancien. L'ecorce du nouveau segment
de racine est done separee de I'ancienne ecorce par des assises de tissus

Fig. 8. — Renewed growth of a mycorrhiza of
Pinus virginiana, showing splitting of mycoderm
and extension of root-tip (Collected near Balti-
more, 22 February 1930).

morts et pigmentes." This is the method of formation of "pearl-
necklace" beaded mycorrhizae.

Ectotrophic vs. Endotrophic: — Mycorrhizae have been con-
ventionally classified as ecto- and endotrophic, the distinction originat-
ing with Frank (1887) who said: "We (may) designate all those
forms as 'ectotrophic' which have the nourishing fungus external
to itself, and as 'endotrophic' where it (the fungus) penetrates into
the interior of certain root-cells." But with passing years doubt was

Lecture IX —123- Structure

expressed whether any such absolute distinction could be made between
the two. Kamienski (according to Grosglik, 1885) had early said
that in cupulifers hyphae penetrate into inner tissues of the root and
extract nutrient; and Melin (1922) said that intracellular infection
does occur in ectotrophic mycorrhizae of Larix, suggesting that
earlier the intracellular hyphae may have been overlooked (Melin,
1923a). Melin called these mycorrhizae which combined characters
of both the preceding sorts, ectendotrophic. Intracellular hyphae in
ectotrophic mycorrhizae were described by Masui (1926a, h) for
Alnus and Abies; by Johansen (1931) for the cactus, Neomammil-
laria, which has a white mantle about the root with hyphae penetrat-
ing between epidermal cells and into cortical cells where apparently
vesicles were formed; and by Klecka & Vukolov (1935), who
found individual hyphae enter cells as short branches or barrel-shaped
structures.

But Endrigkeit (1937), in a study of both sorts, came to the
conclusion that in ectotrophic mycorrhizae of Tilia, Quercus and
Pinus in East Prussia there is no such infection : "The opinion re-
cently expressed as to the endophytic character of the ectotrophic
forest tree mycorrhiza finds no support in the writer's investigations".
He thought that no nutritional or physiological significance can be
attributed to the occasional observation of rudimentary intracellular
infection or the common intensification of the "Hartig net" on a
decayed primary cortex. In pine, Young (1938) observed that "My-
corrhizal development varies from strictly ectotrophic in the case of
untreated controls, through the typical ectendotrophic in the 1^ lb.
of S treatment to the almost purely endotrophic in the 3 lb. S treat-
ment." Both ecto- and endotrophic mycorrhizae were found on
Corsican pine by Aldrich-Blake (1930).

Ectotrophic Mycorrhizae: — In cupulifers, according to Frank
(1885), the root is surrounded by a fungal mantle that is mostly
many-layered; now colourless, now a light to dark brown pseudo-
parenchyma which lies close upon the true epidermis. It sends hyphae
in between the cells but never quite into the innermost layers of root
cortex, growing always in the membrane of the cell only which they
thickly weave about ; but they never enter the cell lumen. The oute*-
surface of the mycorrhiza is not infrequently smooth but root hairs
are never formed, their place being taken by a felt of loose hyphae
that extend out into the surrounding soil.

Mangin (1910) described the ectotrophic mycorrhiza of Castanea
as follows: The diameter of the root augments and the piliferous
cells, covered over by mycelial mantle, are never permitted to form

Kelley

— 124 —

Mycotrophy

hairs ; they elongate in an obHque direction at an angle of 45° with
the axis of the root and their length attains the double or the triple
of their diameter. He notes that, whereas in cupulifers only the
epidermis is involved, in beech, etc., two layers of cells are hyper-
trophied ; while in pine, fir, larch, a great number of layers are affected.
In any case, hyphae intrude between walls of the cells, — dissolving
out the cementing pectate and forming "palmettes" which may cover

Fig. 9. — Cross-section of an ectotrophic mycorrhiza of Quercus
montana, indicating mantle or mycoclena and cortical cells filled
with endophyte. {Original draiving from a slide prepared by Dr.
K. D. Doak).

the radial face of the cell. MacDougal & Dufrenoy (1944) state
that in pine the middle lamella of the outer cortex is traversed. Seen
in section the intruded hyphae between the cortical cells appear as a
netted structure which has been called the "Hartig net" after Th.
Hartig who early observed it, although he supposed the structure
was composed of anastamosing canals.

Besides in cupulifers, ectotrophic mycorrhizae occur in ericads
and some other plants. Hesselman described such structures from
the arctic Salix herbacea and in the herbaceous Dryas octopetala and

Lecture IX — 125 — Structure

Polygonum viviparum. According to Christoph (1921), these my-
corrhizae occur in ericads only in such species as Arctostaphylos Uva-
ursi which form coralloid mycorrhizae in humus, the fungal mantle
living at the expense of epidermal cells. Again, he says that such my-
corrhizae occur only in such species as have subterranean organs dif-
ferentiable into rhizome and root, and especially when, in a humus-
rich location, a copious branching of short-roots has been produced.

Ectotrophic mycorrhizae are, accordingly, distinguished by an
exterior mantling of hyphae that have been called a "mycoclena" or,
when tightly weft into a pseudotissue, a "mycoderm" ; from which
extend hyphae into the surrounding soil. The endophyte penetrates
intercellularly into the epidermis, or in some sorts of plants into the
cortex, in the latter case forming the structure which appears as the
Hartig net. If intracellular infection occurs, then the mycorrhiza is
not strictly ectotrophic. These mycorrhizae occur on roots of woody
plants, rarely on herbs.

Endotrophic Mycorrhizae: — It is evident that a diverse series
of mycorrhizae have been grouped under the term "endotrophic".
Frank (1887) included here those of ericads, which have a mantle
and approach the ectotrophic condition ; mycodomatia have been in-
cluded also; and Gallaud (1905) included mycothalli. Endotrophic
mycorrhizae are typically developed in all Pinaceae except the Abie-
tineae (which areectrotrophic) {cf. Noelle, 1910) ; in orchids, and in
herbs in general.

Gallaud (1905) distinguished endotrophic mycorrhizae into
"series", viz. (1) Series of Arum maculatum: Mycelium at first intra-
cellular in the protective layer of the root, then intercellular and lodged
in the plasm ; arbuscles and sporangioles generally simple and without
very precise localization. Examples are cited from monocotyls, dico-
tyls, and Angiopteris. {2) Series of Paris quadrifolia: Mycelium
always intracellular, arbuscles or sporangioles generally composite,
not terminal and harboured in a definite region of the root. Certain
angiosperms are cited as examples ; also Araiicaria, Podocarpus,
Sequoia and Ophioglossum. (4) Omitting the hepatics, which are
his third series, the Series of orchids: Mycelium always intracellular,
taking the form of coils (pelotons) which sometimes remain inactive
(host-cells), sometimes are digested (digestion-cells). Examples:
orchids, Psilotum and Tamus.

In other words, in various endotrophic mycorrhizae there may be
simple hyphal coils formed as in the orchids, or arbuscles and sporan-
gioles are developed,— the nature of which will be detailed shortly.
Or, it might be said that there are basidial endotrophic mycorrhizae

Kelley — 126 — Mycotrophy

(orchids), and phy corny cetous endotrophs as in the vesicular-arbus-
cular sorts. Since the terms "vesicle", "arbuscle" and "sporangiole"
have been adopted from the French, there seems no valid reason why
the term "peloton" should not also be adopted, to describe the endo-
trophic mycorrhizae of orchids and some other plants.

Peloton Mycorrhizae: — In Neottia Nidus-avis, according to
W. Magnus (1900), the root-inhabiting fungi possess very few and
irregular connections with the outside. The 3-4 outermost layers

Fig. 10. — Cross-section of endotrophic mycorrhiza of Acer
Negundo. Invading hyphae are coiled in the cortex.

of cells beneath the epidermis are completely and without exception
inhabited by the fungus while in rhizome and stem even six layers
may be infested. Infected cortical cells are enlarged and later formed
cells are also enlarged, causing a change in the whole structure. Within
the cortex two sorts of cells were distinguished, "Pilzwirthzellen"
and "Verdauungszelle". According to Magnus, after the hyphae had
formed protein (Eiweisshyphen) their content was taken up by the
cell and the residue was pressed together, while at the same place or
at a place mostly lying in the middle of the cell there begins a local
clotting formation. They then become separated, with a portion of

Lecture IX — 127 — Structure

the plant plasm, as a clot (Klumpe) which is dead, unchangeable waste
product. On death of the fungus a copious formation of vacuoles
takes place and by union of vacuoles a large sap-vacuole is formed
in which the clot remains suspended. A new cell membrane may be
formed about this body: Thus are formed the clots, the "gelbliche
Stoffe", which puzzled earlier observers.

BuRGEFF (1909) observed formation of hyphal coils external to
the mycorrhiza: The hyphae excrete a drop of water into which an
hyphal branch grows, and this branch, hindered by the outer surface
tension from growing out of the drop, is consequently rolled into a
spiral inside. Thus might form the coils (pelotons) of the mycorrhiza.

Three sorts of orchid mycorrhizae were distinguished by Burgeff
{I.e.), viz. {!) Neottid (including most sorts) ; (2) Coralloid {Coral-
lorhisa and Epipogon) ; {2) Sporangiole (including a tropical genus
only). Later (1931) he also described vesicle formation which, how-
ever, is not common in orchids.

Hypertrophy of the nucleus occurs during fungal digestion {cf.
Magnus, Arcularius).

The peloton mycorrhizae, therefore, are characterized by intra-
cellular infection whereby hyphae coil within the cells to form "pelo-
tons" which are afterwards to be digested : vesicles and sporangioles
are rarely developed in this sort.

Vesicular- Arbuscular Mycorrhizae: — The "phycomycete my-
corrhiza" is distinguished by possession of arbuscles and vesicles.
Neill (1944) records as a further distinctive feature a constant anas-
tamosis of the hyphae which sometimes results in formation of a closed
system of intercommunicating passages traversed by moving proto-
plasm.

Vesicles were first recorded by Mollberg in 1884 {cf. Groom,
1894) but were described by Janse (1897) : *T give the name of
'vesicles' to the spherical or ovoid swellings which occupy extremities
of the hyphae. In the young state these organs contain only a small
amount of protoplasm and their cavity is occupied almost entirely
by large vacuoles. Little by little the quantity of protoplasm augments,
nutritive reserves accumulate and finally they are filled with a granu-
lar mass in which are mixed oily droplets. I have found analogous
bodies in a great number of plants. May they be compared to the
cysts which are present in other fungi, and a role in the asexual multi-
plication of the endophyte be attributed to them? I incline to that
belief." It may be noted that vesicular swellings were figured by
Frank (1879) in nodules of Orohiis while they were described still
earlier (1847) by Reissek for certain monocotyls and for Cochlearia.

Kelley — 128 — Mycotrophy

Vesicles may be inter- or intracellular, or even formed outside the
root. Gallaud (1905) regarded the vesicles as "organe de reserve
souvent temporaire" (an opinion which Demeter, 1923, shared) ; and
he described their formation as almost always terminal: the hypha
ceases to grow in length and swells at the extremity while a very
dense protoplasm accumulates there and the nucleus multiplies rapidly
by division. Groom (1894) thought that the vesicles merely appeared
terminal but actually were intercalary, a tip growing into another
cell and there forming another or several more vesicles. Similarly,
Pyke (1935) said that in Cacao vesicles form in the epidermis and
from there a stout hypha grows into cortical cells adjacent ; but
Laycock & Dale (1945) state that vesicles were never seen in my-
corrhizal roots of Cacao, and arbuscles but rarely ; and they would
prefer not to use the term "vesicular-arbuscular" to describe these
mycorrhizae. In Vinca, the vesicles form either terminally or inter-
calary ; and in age, in addition to a number of nuclei they have great
fat vacuoles with protoplasm between them like cross-walls.

Vesicles were described in detail by Peyronel (1923), who con-
sidered them to be reproductive bodies of the endophyte ; and he
figured spores within them, — a view that was held by a number of
earlier authors {cf. Rives, 1923) ; but "most authors consider them
cysts which survive death of the organ and reinfect the new root."
Bernard (1911) figured "germination of a vesicle" isolated and
placed in a hanging drop; while Johansen (1931) described and
figured "spores" (vesicles?) which, he stated, germinate and renew
the infection, in certain cacti. In Stapdia variegata, Busich (1913)
figured a germinating vesicle. Butler (1939) attempted to grow the
fungi from external mycelium and vesicles and, though fine hyaline
new hyphae were sometimes obtained, they did not grow extensively.
The vesicles of Vitis were figured as thick-walled and containing four
rounded bodies like vacuoles (Rives, 1923) ; while in Gossypium
they are terminal, round, oval, or irregular (Sabet, 1939). In
Lolium (McLennan, 1926), vesicles are oval (65x45 fi), usually
terminal but sometimes intercalary and mostly intercellular. At
maturity they have thickened walls but later lose their content and
collapse. They appear to be "an attempt towards spore formation",
while for the higher plant they exist as a "temporary reserve organ".
At an early stage they are packed with fat which is later given up
to the host plant.

No vesicles were formed in synthetic mycorrhizae although such
occurred in nature (Bouwens, 1937).

Vesicles are also formed by the endophyte of Conocephalus, both
in nature and in culture (Beauverie, 1902; Bergamaschi, 1932):

Lecture IX —129— Structure

they are large and spherical in Monodea (Cavers, 1903). They occur
in older prothalli of Ophioglossum (Bruchmann, 1904; Campbell,
1908) while in prothalli of Botrychium thin-walled vesicles are often
so abundant as to fill the cells with a botryose mass (Jeffrey, 1898).
Baas-Becking (1923) said of Botrychium: "The so-called vesicles
occur, thin-walled, apparently osmotic products, filling sometimes the
whole cell ; or two vesicles may occur in a cell."

Arbuscles : — These are said to be even more constant than vesicles
but more delicate and difficult to observe. It was the privilege of
Gallaud (1905) to describe these organs which had escaped earlier
observers. He said that a branch of an intercellular hypha enters
the cell-wall and within the cell gives off 3-4 new branches and then
dichotomises until it forms what looks like a floccose mass. These
ramifications enter into the host's protoplasm. "A cause de leur forme
generale rappelant en petit celle d'un arbre tres chevelu j'appelerai
arbuscles ces formations singulieres tres importantes sur lesquelles
j'aurai a revenir plus longuement." An arbuscle terminates the ex-
tension of each hypha. Gallaud thought that arbuscles are absorbing
organs (organe absorbant ou sugoir). They are simple or composite
(arbuscles composes) when a complex of arbuscles, sporangioles and
hyphae. Vuillemin thought that arbuscles are less a characteristic
production of the endophyte and more a result of the reaction of the
host-cells to invasion by a foreign body.

Digestion of arbuscles by the host results in formation of the
sporangioles of Janse, otherwise called "prosporidi" by Petri. Janse
said : "These sporangioles, as I call them, owe their embossed aspect
to the relief which the spherical bodies that are contained in the in-
terior cast under the extremely thin membrane, bodies which I desig-
nate by the name of 'spherules'. The spherules are filled with a
quantity of little 'granules'. In spite of the name of sporangioles which
I give to these organs, I am far from affirming that they have a part
in the propagation of the endophyte. It must be stated, however, that
they are found, with only two or three exceptions, among all the plants
which I have studied." Gallaud said that transformation of arbuscles
into sporangioles is almost always very rapid, and Janse noted in a
number of the plants he investigated that the sporangioles later freed
their content to form a gummy mass, or at maturity, they formed very
fine granules that diffused through the cell. And Endrigkeit (1937)
decided that intracellular arbuscles cannot be interpreted as assimila-
tory organs since they are digested as they are formed and show no
indication of hyphal development from their terminal branches, but

Kelley — 130 — Mycotrophy

rather as proliferations induced by the growth promoting stimulus
of the cell-sap.

Ericaceous Mycorrhizae : — As described by Kamienski (1884),
Monotropa's epidermis is covered with a fungal mycelium of septate
hyphae which form a compact pseudoparenchymatous mantle 2 or 3
times as thick as the epidermis. The fungus lives on the surface and
never penetrates living cells, but sometimes does so in older portions
where these cells are filled with a brown content. In older parts of
Monotropa roots the epidermis disorganizes at the same time the
mycelium develops. But Francke (1934) said that in epidermal but
never in deeper layers of cells, hypae are found entering and surround-
ing the nuclei, becoming filled with reserves that are emptied into the
host-cells as "plasmoptyse". The reserve is now absorbed but no
excretion was observed, and no significant change in nuclear condi-
tion was seen. After digestion has occurred, vesicular swellings of
hyphae dwindle. Only one hypha enters an epidermal cell. Tannin
is no protection from the fungus for hyphae are found in tannin cells
as well as in tannin-free. MacDougal (1900) said that the mycelium
of Monotropaceae consists of an external absorbing system and an
internal one which fills the epidermal cells. In the Monotropas,
vesicles, sporangioids, and sporangioles fill the cells, and "probably
serve as organs of interchange."

In Arbutus (Dufrenoy, 1917) the roots are clothed with a dense
mantle of hyphae protected by a thick greyish membrane. The fungus
penetrates external layers of cortex and sends haustoria towards starch
grains. Late in the season (Rivett, 1924), hyphae penetrate more
deeply in the cortex, but digestion continues all the time the host is
growing and the reserve storing tissues packed with content. In
Calluna (Rayner, 1927), in young roots the cortex consists of a
single layer of large cortical cells each of which encloses a dense
branch system of mycelium that is continuous with hyphae upon the
external surface of the root. Intracellular hyphae are of relatively
large and uniform diameter with abundant oily content.

Vaccinium is infected by hyphae passing through the cell- walls
(Coville, 1910) and the epidermal cells are completely filled with
coiled hyphae; and in Andromeda polifoUa also the epidermal cells
are infected by very fine hyphae (Frank, 1887) ; while in Vaccinium
Oxycoccos connection was found between external mycelium and
intracellular hyphae. Voss & Ziegenspeck (1929) found digestion
stages ("clumps") in Andromeda and curious mamillae in the cran-
berry. In Ledum these authors found that in winter roots the fungal
hyphae roll together to form a brown mass; in Erica there was less

Lecture IX — 131 — Structure

infection; while in all ericads studied the youngest roots had some
portions fungus-free.

The Pyrolas studied by Furth (1920) had the mycelium spread
over the whole length of the root but confined to the epidermal cells
which were hypertrophied and gradually filled completely with hyphae
that cause the death of the cell and result in development of "clumps".

Goralloid Mycorrhizae: — The various pine mycorrhizae are
described with difficulty, largely because there is so little information
available about them. Although so many papers have been published
on pine, there are very few anatomical studies about them extant.
That is especially true of recent times, for the trend at present is
toward philosophical disquisitions on the nature of mycotrophy, a
branch of learning that conveniently eliminates the drudgery of sec-
tioning. Hence knowledge of pine mycorrhizal anatomy is largely
confined to two European species, — P. montana and P. sylvestris;
and there is almost nothing on American pines. But coralloid mycor-
rhizae are not confined to pines since they occur on other plants, —
on how many others no one could say.

Knowledge of coralloid mycorrhizae (Gabelmykorrhizen) dates
back at least to Reess (1887) although they are said to have been
first described by Hartig in 1851. Moller (1902) said that they
were well known in P. sylvestris while in 1908 he recorded
them from sand of the Brandenburg Marches but absent from humus.
Infection in P. sylvestris and Picea Abies (Melin, 1921) is through
root hair or epidermal cell and hyphae grow at first intracellularly
in outer cortical cells where they form a pseudoparenchyma, but later
form an Hartig net and mantle. Similarly, Abies firma mycorrhizae
(Masui, 1 926^7 ) have not only a mantle and Hartig net but intra-
cellular infection. In Larix the fungus penetrates intracellularly into
the roots (Melin, 1923) and forms individual hyphae or knots which
in time are digested, after which the fungus penetrates intercellularly
and the mycorrhiza becomes more strictly ectotrophic.

In synthesis experiments with pine (Melin, 1923), only ectendo-
trophic mycorrhizae were obtained.

Tuberous Mycorrhizae: — "Knollenmykorrhizen" were brought
into the limelight by Muller (1902) who regarded them as similar
to the coral clusters of cycads and some legumes. They are formed
by a dichotomy which is rare amongst roots, yet not called forth here
by a fungus: "We stress the fact that the dichotomous branched
tubercle is not invaded until after its dichotomy has been manifested".
Both racemosely and dichotomously branched mycorrhizae occur

Kelley — 132 — Mycotrophy

mixed on the same root. From the tubercle extend strands of hyphae
into the soil, as Kirchner said (1908) : "auch hier strahlen von der
Pilzscheide formliche Hyphen-Perriicken in den Boden aus, der zu
einer dichten Masse verflochten wird." Ordinarily these tubercles
live but one year but sometimes they are persistent and continue to
branch, forming a rhizothamnion.

On P. mugho (Laing, 1923), conspicuous nodular bodies, often
over y2 inch diameter, are found on trees not more than 12-13 years
old. The nodules are sessile or stalked and frequently found detached
in the soil. Branching of the rootlets is considered checked by the
fungus which is purely ectotrophic and rarely penetrates cortical
cells. NoELLE (1910) also failed to find the intracellular infection
claimed by Kirchner ( 1908) . In P. sylvestris nodules were described
by Laing (1923) with endophytic infection, while Melin (1922)
also described them, stating that they are as large as a pea. Nodulous
roots on P. Cenihra were described by von Tubeuf (1888), who
quoted from Reess (1887), who seems also to have described them.
Nodules of P. montana are mentioned by Somerville (1911).
Muller (1902) thought that they are of service in fixation of at-
mospheric nitrogen.

Outer Cortex and Passage Cells: — In plants that develop a
thickened outer layer of cortex, or exodermis, in the rootlet, there are
left some thin-walled cells in the layer that do' not develop the wall-
thickening; and these thin-walled exodermal cells form a con-
venient means of access to the cortex beneath for the invading hyphae.
When so used by the endophyte these thin-walled cells are termed
passage-cells. They were described by Janse from Spadiciflorae and
named "cellules de passage", and he said that they are found in all
orchids. Burgeff called them "Durchlasszellen" (Demeter, 1923).
Passage-cells were described from Tipularia by Clifford (1899),
from Aphyllorchis by Groom (1894), from Dipodium by McLuckie
(1922), from Vanilla by Cordemoy (1904), from Vinca and As-
clepias Cornuti by Demeter (1923), who found passage-cells also
in the endodermis, from Gentiana by Schimmler (1937) ; and they
are figured for Hoja carnosa by Busich (1913), who considered
the presence of passage-cells as an inherited character of the Asclepi-
daceae that aids mycotrophy and adapts the plants to their habitat.

Wall Tubules : — A hypha, passing through a living cell-wall, may
stimulate the cell to form a growth about the hypha which has been
called a wall-tubule or, in German, a "Rohrentiipfel". Jeffrey
(1898) had observed in Botrychium mycothalli that a thick sheath

Lecture IX

— 133 —

Structure

surrounds the hypha for 10 or more micra but only when the wall
is cuticularized. Arcularius (1928) found that the host-plant sur-
rounds the hypha with a cellulose layer; Francke (1934) reported
that the haustorial hyphae are very early surrounded by cellulose ;
while Magnus (1900) had found the same in Neottia, and Shibata
& Tahara (1917) had described an analogous relation. The sheath
separates the living hypha from the host plasm. Kusano (1911)
said that the cell-wall formed papillae where hyphae passed through,
which were often branched or formed a "tubular sheath" that he
thought was lignified. Burgeff (1932) said that the wall-tubules are
composed of lamellate cellulose which would indicate by its structure
that there is a diffusion current preventing a regular layering of the
wall, and forming a teet-like structure.

Hartig Net: — Hyphae, on entering the cortex, do not always
penetrate the cell-walls. In certain plants, particularly well seen in the

Fig. 11. — Section of a mycorrhiza of Abies balsamea, showing Hartig net
which has formed about the cortical cells.

conifers, penetrant hyphae weave about cortical cells until they form
a basket-weave structure in which the cortical cells are literally em-
bedded. Seen in section it appears as a netted structure and is called

Kelley — 134 — Mycotrophy

the Hartig net, after Theodor Hartig who early described it from
pine. Hartig considered the deHcate net to consist of anastamosing
intercellular canals (Mangin, 1910:245): it had been described by
NicoLAi (1865) as "thickening strips", and he said that they are
found not only on radial cell walls but in all places where the cells
are not pressed together, so that a peculiar net is formed. He found
them in conifers and the apple-tree. Reinke (1873) also described
such a structure from a number of conifers, and van Tieghem &
DuLioT (1888), as a "reseau de soutien". In pines, Hartig-net is
formed of 2-3 rows of hyphae, as in Moller's P. sylvestris (Shimizu,
1930); Hartig net also occurs in spruce (Melin, 1921), in Podo-
carpineae (Berggren, 1887), and in the fern Tmesipteris (Dan-
GEARD, 1891). Because of mutual pressure of cortical cells, hyphae
penetrating between them become flattened and, branching, are
formed into a digitate appressed structure which Mangin (1910)
called a palmette. But Mangin (1898) said that there is no network
in the cortex but, because of pressure exerted upon them, the hyphae
are flattened into regularly branched layers between the cortical cells.
Voss & ZiEGENSPECK (1929) Said that in older plants of Betula
pubescens the Hartig net is digested ; and in pine it also breaks down
(Lewton-Brain, 1901).

The Stele: — In general, mycelial infection of the central cylinder
does not occur, yet exceptions have been noted. Thus, Dufrenoy
(1917) found infection of all tissues in Arbutus Unedo, and in 1920
recorded heavy infection of pericyclic tissues of Adenostyles albi-
frons. Lewis (1924) found the endophytic fungus growing through
all the tissues of all vegetative organs of Picea and Larix; and
Rayner (1927:p.l00) found similar growth in Calluna stems. Masui
(1926&) found outer layer of pericycle in old roots invaded and in
rare cases the whole central cylinder, which was subsequently de-
stroyed. The stele of Cupania is also sometimes invaded (Waage,
1891). Infection of the stele was found in young roots of Lygino-
dendron but not in older roots by Ellis (1917); while Schacht
(1854) had found vascular infection of a leguminous wood from the
London clay as well as in living ferns and other plants ; although in
these cases the infection may well have been parasitic: indeed, in all
cases of vascular infection the endophyte may be parasitic.

That the endophyte influences structure of the stele is a fact not
widely recognized, according to Noelle (1910), who said that "es
ist bisher nicht bekannt das ein ektotrophes Myzel die Struktur nicht
nur der Rindenschichten, sondern haufig auch das Zentralzylinder zu
beeinflussen scheint". The "normal" (uninfected) root is diarch but

Lecture IX — 135 — Structure

under influence of infestation becomes monarch : the phloem area be-
comes reduced while the xylem area is proportionately increased,
suggesting an increased water intake through fungal influence. But
Hatch (1937, p. 131) says that in pure culture, short-roots of Pinus
Strobus are monarch in vascular structure, and the reduction in
stelar tissues is accordingly not due to invasion by the endophyte. In
holosaprophytes (Johow, 1889), the vascular portion is always much
reduced, or xylem and phloem are diversely arranged. The stele is
much reduced in J uncus Tenageia following infection (Grutter,
1886). Later researches will have to determine further the influence
of endophytism on stelar anatomy.

LECTURE X
OBLIGATE SYMBIOSIS

Fungi and Trees: — That fungi are confined in their symbiosis
to one species of plant was stated as early as 1841 by Tulasne, who
said that Elaphomyces granulatus was confined to one species of tree ;
and GiBELLi in 1883 suggested that the mycorrhizal condition is a
necessary one in the cupuliferae. In more recent years the confine-
ment of certain fungi to the neighbovirhood of certain trees has been
remarked. Thus, Barsali (1922) found that in forests about Pisa
there were always the following fungi associated with the trees named :
Lactarius delicosus under Pinns Pinea on sandy soils (but Romell,
1939, said that this fungus is rarely found with pine) ; L. volemus and
L. oedomatopus under P. pinaster; Boletus boviiius under P. pinaster
and Jiiniperus macrocarpa; B. granulatus under P. pinaster; B.
edidis under Quercus and Castanea; B. corsicianus under Q. ilex
(where this species grows with Q. Suher) ; Rnssida grisea and R.
emetica together with Lactarius under P. pinaster. Lange (1923)
gives a comparable list for Denmark, and Lidl (1939) for Germany.
Palm (1930) had noted a Boletus growing in association with P.
Merkusii in forests of Sumatra and another bolete under P. cubensis
in the Guatemalean highlands. Young (1937) found both Rhiao-
pogon luteolus and Boletus granulatus under the same pine tree in
Queensland, and thought it possible that both are symbionts of the
same tree at the same time. Reess (1885) had concluded that Elapho-
myces is dependent for its occurrence on presence of pine. Again,
Hammarlund (1923) noted association of Boletus with Larix in
Sweden, and proved by synthesis that the fungus is mycorrhizal.

Ecological Influences: — An interesting ecological study of such
occurrences was made by Peyronel (1917) : (i) in Larix decidua
woods, boleti predominate; (2) in cupuliferous woods agarics pre-
dominate, especially polypores ; (J) with Salicaceae, Populus treniula
has a rich fungal coterie while Salix alba has none; (4) Betida alba
is accompanied by a discrete number of humus fungi, while Alnus
glutinosa is found with Lactarius; but A. viridis lacks characteristic
humus fungi ; (5) most other ligneous species are never accompanied
by humus-dwelling hymenomycetes ; (6) meadows are characterized

Lecture X — 137 — Obligate Symbiosis

by many species of Hygrophorns and Agaricaceae. In all cases the
variations in the florula seem correlated with the sort of humus
present.

Special Gases: — Such examples could be multiplied but the
objection is raised that finding of sporophores under certain trees
is not a priori proof that these fungi are mycorrhizal symbionts of
the trees ; neither does tracing of mycelial connection between the two
constitute proof. But Romell (1930) offers the interesting observa-
tion that Lactarius delicosus not only constantly occurs with Picea in
the region of Stockholm, but that this species disappeared upon re-
moval of the few spruce trees that stood in a mixed pine stand. On
the other hand, Dittrich (1923) said that L. delicosus is found in
great quantities under thick-set spruce trees when the trees are young,
but disappears completely after the trees have reached a certain age.
An interesting case is cited by Mattirolo (1934&) in that Populus
canadensis was introduced into Italy from America about 100 years
ago : Tuber Borchii is found on this poplar and, as the fungus is
recorded also from California, may have been introduced into Italy
on introduced Amentaceae. Further, Melin (1922) said that Boletus
elegans appeared in Sweden only after introduction of larch.

Fungi and Herbs: — Associations of fungi with herbs are like-
wise noted. Thus Colla (1931) found two species of fungi associated
with Dryas ocfopetala. Fraser (1931) made "the first report of an
obligate association of an annual herb with a mycorrhizal fungus"
in two species of Lobelia. In a study of German species of Polygala,
Heinricher (1900) concluded that some species at least of this
genus can scarcely be considered as obligate mycorrhizal plants, for
he was able to grow the plants in culture without mycorrhizae. For
the orchids, Kusano (1911) thought that Gasfrodia was unable to
flower without infection as indicated by pot experiments. Obligate
symbiosis is indicated by Porter (1942), who reports that Rhisoc-
tonia mucoroides is probably the specific endophyte of Zeuxinc
stratcumatica since this fungus is found both in Florida and in Java.

Obligatism and Nutrition: — Without multiplying examples, the
argument is the same in all cases : Certain fungal fruiting bodies are
found habitually in the neighbourhood of certain higher symbionts,
and by careful examination a mycelial connection can be traced be-
tween the two. The objection is raised that here there is no necessary
obligate relation since it is possible, perhaps, to grow the two apart
in sterile culture. The fact remains, however, that the two symbionts

Kelley — 138 — Mycotrophy

do grow in union, and are thus found in association. The relationship
is probably not obligatory since the fungus, e.g., which grows on Finns
Taeda in America might grow equally well on P. sylvestris in Europe ;
but in actuality the two are separated by the Atlantic ocean and have
no means of coming together. In other words, the association is
obligatory only in the sense that the symbionts have no other means
of livelihood except in association. The humus-dwelling hymenomy-
cetes cannot live in arid mineral soil and cannot therefore associate
with dune grass or beach plum ; and it may be that some are limited
to certain specific compounds just as human individuals are limited
in metabolism to certain polypetides. At the same time it is perfectly
true that the otherwise "obligate fungi" can be grown in culture apart
from their usual symbionts ; yet the laboratory and the woodland are
two different things, and without being too cynical we may say that
our experience with natural woodlands is that they are notably deficient
in supplies of Erlenmeyer flasks and culture media.*

The Ericads: — The ericads offer a much-mooted relationship.
At least three cases of obligate symbiosis have been recorded for these
plants: {1) Francke (1934), in a study of Monotropa Hypopitys,
found that a fungus-free protocorm was never seen which, he thought,
indicated an obligate mycotrophisni. Kamienski in 1884 had raised
the question whether the relationship were obligatory. (2) Paula
FiJRTH (1920) made the unconfirmed report of obligate symbiosis
in Pyrola in Lower Austria. (J) Rayner (1915) claimed to have
proved an obligate relationship in Calluna vulgaris. A fungus identi-
fied as Phyllophoma was found to grow not only in roots but through-
out the whole plant — stems, leaves, flower and seed-coat but not grow-
ing into the embryo. Infection of the developing plantlet was from
the seed-coats ; while sterile seedlings did not live long enough to
develop roots. But Christoph (1921) reported that Calluna in
nature is facultatively mycotrophic since the fungus is entirely absent
from habitats lacking humus, indicating that an obligate symbiosis
does not exist.

The Problem of Calluna: — Calluna was further studied by
Knudson ( 1929) . He used seed grown in the U.S.A., and germinated
the seed on Rayner's solution A to which was added 1.5% of stand-
ardized agar. The seed was sterilized in a Ca hypochlorite filtrate,
usually for 30 min., and transferred by a looped platinum wire

♦Systemic infection is also claimed by BosE for Casuarina. He says that
only the resting embryo is free. Seed infection was also found in tomato, apple
and several kinds of cereal grains (Nature 159:513-514, 1947).

Lecture X — 139 — Obligate Symbiosis

directly to the culture tubes without rinsing. In the first experiment,
no effort was made to isolate seed from the floral tissues but the whole
mass was placed upon the agar. In every case an Alteniaria was found
contaminating the culture but owing to lack of sugar fungal growth
was slight. The roots of many of the plants were examined micro-
scopically and in no case was any fungus-root infection found. The
seedlings developed normally without fungal infection. In the second
experiment, because of contaminations in the preceding, sound seeds
were selected and the experiment was repeated. No one h.i.c. was
found most favourable for growth but availability of iron was limited
in more acid reactions. No root infection was found and the roots
developed normally. "The conclusion is inevitable from this and the
preceding experiment that the fungus is not essential for normal
germination of the seed." Knudson supposed that Rayner's unin-
fected seedlings had behaved "abnormally" either because of toxic
action of a possible excess of iron in the culture solution, or through
injury by mercuric bichloride in sterilisation.

Rayner (1929) rebutted these arguments by stating first, that she
never used more than a trace of iron in solution (3-4 drops of 0.1%
solution per litre), and second that Knudson's sterilisation method
(with calcium hypochlorite) was inadequate and that he had there-
fore not destroyed the fungus in the seed-coats.

Freisleben (1933), finding the problem still unsettled, made
culture experiments that demonstrated much better growth of Calluna
seedlings with fungus than without, thus confirming Rayner in respect
to benefit of mycotrophy ; but he also found it possible to grow sterile
seedlings, indicating the symbiosis is not obligate and that mycelium
does not penetrate seed. Indeed, he found much less general infection
of the plant than indicated by Rayner. Continuing his studies (1934) ,
he found that endophytes of various Vaccineae could be exchanged,
indicating that there was no obligate symbiosis in these plants.
Lumiere (1919) had called attention to the fact that Stahl had
grown Vaccinium in sterile soil without difficulty. Molliard (1937)
studied Calluna still further and came to agreement with Knudson
and Christoph, that presence of mycorrhizae is not necessary to de-
velopment of heather.

Other Ericaceae: — In Rhododendron, infection by the endophyte
is not an obligate condition of development of the higher plant, which
in fact can form roots and establish itself in the total absence of
micro-organisms (Gordon, 1937). Neither is there obligate symbiosis
in Azalea mollis: Melin (1921) experimented with this species and
concluded that mycorrhizal fungi are not found in the aerial parts.

Kelley — 140 — Mycotrophy

Freisleben's statements about Vaccinium have already been noted :
he showed that in this genus there is no general shoot and seed in-
fection, and accordingly no cyclic symbiosis can exist. In V. macro-
carpon, according to Bain (1937), "Systemic infection of the type
attributed tO' Phoma radicis by some investigators could not be found
either in seedlings or in prepared slides from field grown material.
The hypothesis of systemic infection by mycorrhizal fungi and its
obligate relationship to root formation in the heath family was
examined critically and it was shown that the hypothesis fails to con-
form to observed facts in some important respects, for example, in
100,000 cultures of cranberry fruits made by the U.S.D.A. in the
last 30 years not even a single culture developed Phoma radicis."

The casual nature of the mycorrhizal symbiosis is made evident
once again by these studies. Apparently there is no obligate symbiosis
in ericads in the sense that the ericad could not live without the fungus,
neither that the two are habitually associated ; but yet the fact remains
that in the ericads' usual habitat, conditions exist which favour mycor-
rhizal association with an endophyte, and such exists. Perhaps in
unusually favourable habitats there is a general systemic infection.

The Orchids: — Next may be considered the orchids, in reference
to which there has been much argument as to their possible obligate
mycotrophy. As to the possibility of its occurrence in roots and
rhizomes of orchids, a denial was made a century ago by Reissek
(1847) : "The regularity and Constance with which fungal formation
occurs in orchids must be considered a characteristic and vital pheno-
menon, but just as phanerogams can be propagated without seed, so
can orchids be produced without root-fungi."

In the orchid Gastrodia, Kusano (1911) concluded that the plant
is dependent on its fungal endophyte because only in association did
the orchid thrive and bloom well. But Curtis (1937) denies that
there are any specific endophytes with orchids. He said : "There is
an apparent correlation between ecological habitat and fungus type,
rather than between orchid species and fungus." Bog orchids all had
the same fungus indiscriminately but some orchids had dilTerent
fungi dependent upon whether they grew in bog or prairie. Several
strains of RJiisoctonia were isolated from the same orchid. Hence
Curtis concluded that there is no specific mycorrhizal fungus, but
any species found in the habitat may form mycorrhizae. This is the
same principle of fortuitous mycorrhizal formation that seems gener-
ally to be true.

While speaking of orchids as a unit, it is evident that, like other
sorts of biological phenomena, they consist of various discrete entities.

Lecture X — 141 — Obligate Symbiosis

BuRGEFF (1932), who is so well qualified to speak on the subject,
distinguished amongst the orchids, holosaprophytes (plants with
normal roots but of a very limited number), hemisaprophytes (in-
cluding only Hcllehorine amongst orchids), and root saprophytes (in
which the root alone has taken over the saprophytic function of
acquiring carbon-compounds). Root saprophytes, he said, seem less
adapted to mycotrophy than rhizome saprophytes : Gastrodia is an
extreme example of a root saprophyte. Saprophytism of the germinat-
ing plantlet, independence of the embryo of its nutrient tissue, the
smallness and great number of the seeds, must go together with fungal
infection and the antagonistic phenomena of phagocytosis. But de-
terminate of results of infection is quality of the fungus together
with the amount of its transmitted organic material. The fungi are
of different sorts and lead by steps from Rhizoctonia to the cellulose-
splitting Hymenomycetes and thence to the wood-fungi; and with
increase in amount of nutrient translocated by the fungus there is
development of saprophytism and increase in size of the saprophytic
organ.

Germination of Orchid Seed: — Mention of the germinating
plantlet leads to a consideration of obligate endophytism of orchid
seed germination. NoiiL Bernard, the gifted French botantist who
did so much for our science during his brief life, first made known
the fungal symbiosis which exists with orchid seeds. It had long
been a mystery why seeds germinated with such difficulty, although
germination had been observed as early as 1804 by Salisbury, who
described it from OrcJiis Morio and Limodorum. Bernard (1899),
by a chance observation of germinating seeds of Neottia. a stalk of
which had been accidentally buried in soil, saw that fungi (which he
called "mycorrhizes") were associated, and jumped to the conclusion
"that the mycorrhizae are indispensable to the plant at the time of
its germination". This conclusion he then set himself to verify, with
brilliant results. In 1900 he said: "The known difficulty in germinat-
ing orchid seed is due to the fact that presence of a fungus, normally
present in orchid roots, is necessary for their development." In 1902
he stated: "In orchids the mycorrhizal fungus penetrates even into the
germinating seed and apparently they are able tO' germinate only when
infected." Next year he continued : "Thanks to the help of M.
Magne, I am able to recount observations on germination of Cattlyea
and Laelia. The seeds of these species and their hybrids are the most
easily germinated of their series. . . . these experiments show that
penetration of the fungus is a necessary and sufficient supplementary
condition for their germination."

Kelley — 142 — Mycotrophy

Necessity of Orchid Fungus: — Here was a thesis around which
a battle was waged. Bernard continued to develop his idea (1903) :
An orchid seed germinated aseptically, he said, will swell to a large
diameter but will then still be stationary after 100 hours of culture,
but transported to a pure culture of the hyphomycete will germinate
at once. "This case is, I think, the first certain example of an organism
which is not normally able to get past an erabyronic state without
penetration of a parasite, just as an egg, e.g., cannot pursue its de-
velopment without fertilisation. Using a term which has been applied
to lichens, we may say that we have made the synthesis of a plantlet
of the orchid." These plantlets are not comparable to those of the
majority of plants, which are derived from an egg, but are complex
forms of the value of mycocecidia {i.e., mycodomatia). Bernard
(1904) proceeded to isolate the orchid fungi, finding a number of
which some induced germination and others not. In 1905 he recorded
experiments showing orchids exceptionally difficult to germinate de-
pend on endophytes different from those already isolated from
Cattlyea and Cypripediiim : thus there is a differential power amongst
the fungi. He said (1909a) : RJiizoctonia repens affects the majority
of orchids but R. languinosa and R. mucoroides infect them with
comparative rarity. (The orchid fungi are generally basidiomycetes :
cf. Derx, 1937; Sprau, 1937).

Degeneration of Orchid Fungus: — Then came a statement that
added a fresh discussion : The activity of each species of fungus is
not fixed but dwindles rapidly when the Rhizoctoneae live apart
from the orchids : it does not require more than 2-3 years to cause
their activity to become unappreciable. Burgeff (1909), recounting
his own experiments, stated, however, that while Bernard found that
orchid fungi, which are essential to germination of the seed, soon
lose their virulence if cultivated for some time apart from the orchid,
he himself found that the fungus of Hahenaria psychodes and others,
in spite of over 2 years culture on starch-agar exhibited the same
infection-power which the fungus had earlier possessed. He felt
that any idea of a decadence in infective power of the fungus should
be abandoned. (It may be noted here that Wolff, 1923, found that
the age of orchid seed determines its power of germination,
vitality dwindling rapidly after maturity and at the end of 38 months
there was no germination.) Burgeff isolated a considerable number
of orchid fungi which he at first termed Orcheomyces but later
called Mycelium radicis. That it is the orchid root fungi that cause
germination of orchid seed was further demonstrated by Sprau
(1937), who isolated a Rhizoctonia from Orchis masculus that stimu-

Lecture X

143 Obligate Symbiosis

lated seed germination of the same species. The same sort of relation-
ship was supposed by Wibiral, 1910, to hold good with respect to the
Gentians,

Hydrolysis of Starch: — In germination of an orchid seed
(BuRGEFF, 1910), at first some oil is transformed into starch; then
the fungus grows in by way of dead suspensor cells, changes starch
to sugar and thus increases osmotic energy and greatly accelerates
growth. The fungus is now digested by enzymes produced by the
orchid cell and a clot is left. With most orchid seeds, said Burgeff,
there is never germination without a fungus.

Sugars and Asymbiotic Germination: — In stating that the
fungus changes starch to sugar another major thesis was enunciated.
Bernard had initiated asymbiotic orchid seed germination by causing
the seed to germinate asymbiotically with salep, a product of dried
orchid tubers rich in bassorin, a polysaccharide (cf. Ballion, 1924).
Beau (1920) had further experimented with seeds of Spiranthes and
Orchis, causing them to- germinate by transfer of salep material from
gelatine to seed via a fungus. Ballion & Ballion (1924) secured
strictly asymbiotic germination of Cattlyea with a medium half mineral
and half organic. Knudson (1922) extended these experiments by
trying the effect of various sugars on germination of orchid seeds.
He sterilised seeds in calcium hypochlorite and cultured them on agar
slants; and found that seeds of Cattlyea, Laelia and related forms
germinate when sugar is supplied, fructose appearing more favourable
than glucose. In glucose, chlorosis resulted, the concentration of
glucose appearing important. Later (1924) he germinated seeds of
various hybrids of Cymhidium, Odontoglossum, Phalenopsis and
Ophyrs, finding "almost 100%" germination in sugar solution with-
out fungal aid. He said: "These experiments lend further support
to the hypothesis that the germination of orchid seed is dependent on
an outside source of organic matter." Failure to germinate without
sugar suggests necessity of an internal factor to activate chlorophyll, as
indicated by Briggs. After the process is initiated, then seedlings
develop without aid either of fungus or of sugar. In still further
experiments, Knudson (1925) isolated orchid fungus resembling
Rhisoctonia repens from Cattlyea, Cypripedium and Epipactis; and
grew it in culture solution with added "0.5% starch". This fungus
induced germination in Cattlyea "but not 100%", but there was no
germination without it. The h.i.c. was increased by the fungus due
tO' organic acids excreted by it, best growth being between pH 4.7-5.2.
I'lie fungus digests starch and changes it to sugar, while some of the

Kelley — 144 — Mycotrophy

sugar is changed to organic acid. With less starch, most of the seeds
were killed by the fungus. With starch in the culture solution there
is no germination unless fungus is supplied. With sugar and fungus,
plants made much better growth, the beneficial effect being attributed
to the fact that the fungus changes the h.i.c. to a more favourable con-
centration. La Garde (1929) found maltose the most efficacious sugar,
and the best h.i.c. between 4.8 and 5.2, no germination taking place
above pH 6.0. Clement (1924&) found that for Odontoglossum
the pYi was best held at 6.5-6.8, which he said possibly influenced
solution of phosphates. For Goodyera, Downie (1940) said that
levulose and dextrose are better than sucrose. But the same author
said that seeds of Goodyera on mineral nutrient solutions adjusted
to range of 3.6-7.6 failed to germinate without the fungus, but with
the fungus no other aid was necessary. And Porter (1942) found
the results of symbiotic tests were generally far superior to those
secured on the asymbiotic substratum.

Fungus Supplies Sugar: — These experiments of Knudson's
confirm earlier discoveries and further elucidate the fungus-orchid
relation. It seems evident that the crucial action in germination of an
orchid seed is provision of that seed with a sugar solution, although
recent work indicates that something more than a carbohydrate is
necessary for germination and development of the orchid, namely
a "growth-factor" which is supplied by the fungus. Furthermore,
Curtis (1943) thinks that vernalization is necessary for germination
of Cypripediiim seed: he secured some (20%) germination in an
adjusted environment with a covering of agar to prevent a down-
ward diffusion of oxygen. But provision of sugar can come only
through aid of a fungus, or by artificial supply by man : if there
is a third alternative we do not know it. Apparently the orchid
seed is incapable of supplying its need through any autolytic action
since practically all orchid seeds fail tO' germinate unless stimulated
by an external aid as so early indicated by Bernard. Knudson fails
to meet this situation when he asserts that fungi are not responsible
for germination of orchid seeds ; and his only suggestion is that in
nature other fungi than the "orchid fungi" may provide the seed
with sugar. But what other fungi ?

Downie (1943) suggested that the symbiotic fungus of Good-
yera repens, which cannot survive in compact humus, may live in
other ways; for he found it on bi foliar spurs of Piiius syh'estris
before the spurs were shed. He thought that his discoveries "contra-
dict Knudson's hypothesis that orchid seed germination under
natural conditions is effected by the action of non-symbiotic sapro^
phytic fungi on the organic substratum".

Lecture X —145 — Obligate Symbiosis

Impotence of Unaided Small Seeds: — Ceillier (1912) pre-
sented a suggestive paper on the germination of small seeds. In
certain cases, such as in Juncaceae, the seeds are small and little
differentiated but as they possess chlorophyll they are able to begin
photosynthesis immediately on sowing. Small seeds with much re-
duced embryO' also occur in parasitic forms such as Cuscuta, Oro-
hanche, etc. ; no fungus is present in these genera but apparently
germination is not successful unless contact is made with organs of
the requisite host. It may be, said Ceillier, that the stimulus neces-
sary in these cases is analagous to that requisite to bring about root-
formation in those plants with "obligate mycorrhizae". Here is a sug-
gestion of interest : Cuscuta can germinate unaided but only as it
develops parasitic connections can it be nourished. By an apparently
casual meeting with a host is its vigorous life assured. So it is ap-
parently with the orchids ; they have no specific fungi (as stated in
some detail by Curtis, 1939; also by Derx, 1937) ; but without any
fungi at all they appear as helpless as Cuscuta without an host. This
is more or less the conclusion of Downie (1940) who used Knudson's
methods with Goodyera repens and stated that "The experiments . . .
tend to support the original contention of Bernard that endophytic
fungi play a large, if not the whole, part in germination of orchid
seeds in the field."

Germination of fungus-spores, seeds and pollen-grains has been
shown to depend on external supplies of activators, dependence on a
particular activator being due to a failure to synthesize this or a
similar substance (Brown, Nature 157:65-69, 1946).*

Carbon Supply to Embryo: — Another angle of the problem
was presented by Pollacci & Tredici (1936) : It is not the fungus
per se that causes germination but something produced by the fungus.
Using seeds of Cymbidium, Cattlyea and Phalcnopsis, he found that
germination was accomplished by filtrate from Rhizoctonia; also from
mycelium isolated from Phalcnopsis; and perfect plants were grown
to flowering age. What these substances are that produce germination
remain to be discovered; but Beau (1920) suggests that they are
carbonaceous. He grew the orchid fungus on a gelatine to which
salep was added and placed seeds of Spiranthes and Orchis on the
glass but not in contact with the gelatine. When hyphae of the fungus
grew up and into the seeds, germination was accomplished and growth
continued until the hyphae were destroyed that connected the seed

*ScHAFFSTEiN "insists that seeds of the genera Vanda and Phalenopsis, for
germination, must be supplied with the vitamin, vandophytin, which they lack"
(Jahrb.wiss.Bot. 68:720-752, 1938).

Kelley — 146 — Mycotrophy

with the gelatine ; then growth stopped. This experiment indicated
that sugar within the seed was not sufficient to support growth but
that the embryo was dependent on an external carbonaceous supply.
By further experiment it was found that the orchid fungus could
grow "perfectly" on cotton moistened with non-carbonaceous mineral
solution, the cellulose being dissolved. These facts fit into a carbon-
theory of mycotrophy, and throw further light on the use of sugars
in germination of orchid seeds.

Obligatism in Lower Plants: — The so-called lower plants are
yet to be considered. In the hepatics, obligate symbiosis was posited
by Chaudhuri (1925) for the Indian liverwort, Marchantia nepalen-
sis. This species was found to contain an endophyte characteristically,
but sterile thalli were easily raised, developing normally for a time
but soon drying up without forming spores. Infested plants both
vegetated and reproduced ; from these facts it was concluded that the
hepatic cannot develop to maturity without a fungus. Goebel's Or-
ganography, 3d Ed., remarks laconically : "This requires further
proof". Nicolas (1924), working with Luniilaria cruciata found
likewise that fruiting occurred only in infected thalli, the sterile
thalli being destitute of mycelium. This observation, together with
Golenkin's, suggests the possibility, said Nicolas, that presence of
a symbiotic fungus is necessary for fructification.

Chalaud (1932) considered the fungal endophyte necessary to
tuberisation in the Indian hepatic, Sewardiella. Each year the succes-
sive gametophytes are infested but the endophyte is checked in the
tissues by activity of meristematic cells that are incited to form an im-
mune tuber. These facts, thought Chalaud, are in conformity with
Bernard's discoveries about orchids ; and he considered Sezvardiclla
as the most nearly perfect adaptation of gametophyte to fungus
amongst the liverworts, presenting a chart in which Lunularia infec-
tion is listed as accidental, Marchantia and Pellia as habitual, and in
Fossombronia leading to a tardy tuber-formation and little change
in vegetative character.

As to fern prothalli, Nakai (1933) supposed that uninfected
thalli of Cheiroplenria would be sterile or male, implying a need for
infection to form archegonia.

Infection of Lycopods is very general, but we do not know that
the symbiosis has ever been claimed as obligate.

Summary: — From a review of what has been thought and dis-
covered about obligate mycotrophy it is evident that there are two
view-points in regard to the question, a theoretical and a practical view-

Lecture X —147— Obligate Symbiosis

point. To illustrate them one may cite the following : A frog is killed
and the heart bathed in an isotonic salt solution. One may now say
the frog is not dead because its heart is still beating. In regard to
Obligate Mycotrophy, there is an academic view of obligatism by
which we have to admit that no plant is so united to its fungal partner
that the plant cannot be forced to hve in some other way ; and there is
a natural view of obligatism by which we perceive that the vast
majority of plants in nature are obligatorily mycotrophic through
their physiological requirements in a limited environment.

Added evidence for this view is afforded by the work of Dominik
and Jagodzinski (Diary Trees & Forest Res. Inst. Kornik. 1 :48-73,
1946). Working with fruit trees, they find that certain spp. are con-
fined to a mycorrhizal nutrition. Since the soils of Kornik garden
contain less than a minimum required for ordinary plant nutrition,
plants growing in it are necessarily dependent on fungi for N supply.

LECTURE XI

THEORIES OF MYCOTROPHY

Contrasted Concepts: — There are two contrasted concepts of
the endophytic relation, first, that the fungus is a parasite on its host,
and second, that the two symbionts live in some sort of a mutualism.
Of 118 papers in which the writers expressed a definite opinion about
the endophytic relationship, fourteen voted for parasitism. To some
extent these opinions were based on personal bias rather than on ex-
periment, and it remains evident that most investigators who have
worked experimentally with endophytic structures conclude that the
relationship of fungus to host is beneficial to at least the higher plant.
But the views of the others deserve consideration, especially since in
a number of cases the divergence in opinion is after all a divergence
in viewpoint, and in the last analysis both sides are found in essential
agreement.

Parasitism: — Those who concluded from a morphological study
that the relationship is a parasitism include : Cavara ( 1893) , Chodat
& Lendner (1896), GiBELLi (1883), Goebel (1888), McDougall
(1914 et seq.), Masui (1926), Prat (1934), and Robert Hartig
(1888), who was decidedly opposed tO' Frank's ideas of mycotro-
phism. He based his opinion chiefly on the alleged observation that
trees in nature have young roots that are entirely immune to the
fungus, and that mycorrhizae are more or less exceptional. "I have
shown above", said Hartig (p. 118) "That mycorrhizae are not
always present, that not a trace of a mycorrhiza was to be found on 10-
year old oak, beech, hornbeam, hazel, in the experimental garden of
the Forest Institute, which were carefully excavated, that a relatively
large number of roots of native trees which were carefully studied
were found to be fungus free, and that there is not a single fact
which would lead to the belief that cupulifers, conifers, ericads, etc.,
have so remarkably different a mode of nutrition from that of other
trees. I see in mycorrhizal fungi nothing more than parasites which
live on the tree but do not kill it, just as there are countless parasites
which live on the leaves without injuring them." Much of the preju-
dice against mycotrophy which has existed to this day comes from
this positive statement of Hartig. Such is the power of dogmatism.

Lecture XI — 149 — Theories

A number of investigators have concluded that the endophytic
relation is a parasitism after experimental study. Thus, F. Fuchs
(1911), who originated synthetic experiment in mycorrhizal study,
came to the conclusion after studying synthetic mycorrhizae of coni-
fers, that there is no symbiosis here of the sort in which the host
plant is benefited. Where fungal hyphae penetrated into cells they
were deformed and killed while the infested cells turned brown and
were thrown off by the root. This holds good for endotrophic roots
as well. The wide distribution of mycorrhizae indicates that it is
an endured parasitism (ertragbares Parasitismus) in which the host
plant suffers no injury because it is able to render the fungus harm-
less. With our present knowledge we can see that Fuchs was partly
right and partly wrong.

H. Gordon (1937) was led to the view that the endophyte of
Rhododendron spp. is a relatively feeble parasite and of no specific
importance to the higher plant by the observation that in culture the
higher plant can form roots and establish itself in the total absence of
any micro-organisms. Needless to point out, Gordon worked only
under the highly artificial conditions of culture and did not deal with
actual plants in nature. Christoph (1921), like Gordon, concluded
in the case of other ericads, that the fungus is simply a harmless para-
site that does not injure the plant, because he was able to raise sterile
plants in culture, and in nature he says that Erica, Calhina, the Pyrolas
and Monotropae are only facultatively mycotrophic.

Curbed Pathogens: — Similar opinions are expressed regarding
orchids. Bernard himself was no mycotrophist, but regarded the
endophytes as invaders that are checked by an humoral substance in a
phagocytosis that immunizes the remaining tissues of the plant.
CosTANTiN (1926) confirms Bernard and regards the orchids as
pathological, being hereditarily accommodated to their disease. It was
perhaps with such precedents in mind that Burges (1936) wrote:
"The presence of a fungus in a mycorrhizal association is to be
regarded as an example of controlled parasitic attack and has no
mutualistic significance. The fungi are weak pathogens whose activity
is curbed by the reactions of the host-cells. . . . One seems justified
in concluding that the mycorrhizal fungi, both ectophytic and endo-
phytic, are potential parasites controlled by reactions of the host-
cells." Curtis (1939) likewise votes for parasitism, influenced by
a cultural study of possible specificity amongst orchid fungi. From
the fact that orchid seed can be germinated asymbiotically and that
the orchid-fungus relation is non-specific, Curtis concluded "that
the symbiotic relationship is one of parasite and host, with the orchid

Kelley — 150 — Mycotrophy

deriving no benefit from the fungus in its roots." But MacDougal
& DuFRENOY (1944) say that "The non-necessity of the fungus for
germination of terrestrial orchids has been wrongly taken as a proof
of parasitism by J. F. Curtis."

A similar comment could be made of Renner's (1935) assertion
that because seeds of Salix and Acer germinate without fungal aid,
also because soil-grown plants grew equally well without fungi while
weaker plants in water-culture were killed by the fungi, that the
symbiosis is a tolerated parasitism.

Magrou (1921) took the position that mycorrhizal symbiosis is
on the border of disease. In perennial plants, as the potato, the fungus
is in part phagocytized while in annual plants (as Orobus) it was
completely destroyed. Limitation of the fungus was thought due to
toxic constituents of the cell-sap, while tubers appear as symptoms
of disease. In 1928, Magrou inclined to the view that the fungi are
mdififerent parasites which, if not harmful in all cases, are without any
use.

Schools of Mycotrophism: — Those who reject the parasitic
view of the mycorrhizal symbiosis and believe in mycotrophism are
divided amongst themselves into several divergent schools of thought.
They all agree that the mycorrhizal fungi are useful to the higher
plant, but they disagree as to what the mycorrhizal fungi afford to
the higher symbiont.

Mycorrhiza Replaces Root-Hairs: — One of the earliest views
was that the mycorrhizal fungi, through the hyphae that connect
mycorrhiza with soil, take the place of root-hairs and provide thS
higher plant with nutrient. Pfeffer (1877) said that in rhizome and
roots of saprophytic orchids there frequently appears a fungus the
mycelium of which, at least in Neottia, is found in living cells and
sends out strands to the outside which, like root-hairs, take up
organic and inorganic soil portions and with them make growth. Per-
haps Pfeffer borrowed the idea from Drude (1873) who had said:
"This constant entrance of a parasite into definite layers of the sub-
terranean organs requires closer consideration. We have never found
it in the epidermis : the question arises whether the basis thereof is
not to be sought in the circumstances connected with the nutrition,
whether perhaps the mycelial threads in these cell layers might not
yield a rich nutrient." Marcuse (1902), after a study of representa-
tive mycorrhizae of a number of plants, came to the conclusion that
"in most endotrophic mycorrhizae the communication-hyphae have a
physiological role comparable to root-hairs, as was first hypothesized

Lecture XI — 151 — ■ Theories

by Pfeffer." Kamienski (1884), finding Monotropa roots felted
with hyphae which prevented any direct contact of the plant with soil,
concluded that the fungus is its nutrient provider. "There is no other
way by which nutritive solutions may pass and provide the roots of
Monotropa, except the way of the mycelium." Fluid intake was also
posited by Issatschenko (1913) : "Disappearance of starch from
the nodule (of Trihulns) was observed which, according to Noel
Bernard, increases the osmosis of the cell and with it the water in-
take ; and in spite of Pavillard's recent criticism, the author con-
siders the relation between mycorrhizae and intake of water as un-
questionable."

This was the idea which Frank originally adopted. As reported
by Klebs, the fungus, which is to be regarded as a parasite in its
colonisation and entrance, causes the host-tree no more damage than
that an organ is developed which supplants the regular roots of the
tree in taking in water and inorganic salts from the soil. As water
cultures indicate, the tree is not dependent for its existence upon the
fungus although it is evident that with fungal aid the tree grows all
the more thriftily. Yet the fungus seems in its development confined
to the tree, since efforts were not successful to grow the fungus on
culture media. Frank regarded his discovery of mycotrophy in
cupulifers as "ein neues Beispiel von Symbiose im Pflanzenreiche"
and stated (1885a) that "der Pilz als der alleinige Zufuhrer alles
fiir den Baum erforderlichen Wassers und Nahrmaterials aus dem
Boden erscheint." This statement is simply an amplification of
Pfeffer's earlier statement about Neottia.

Miehe (1918) adopted the same idea when he said that myco-
trophy is a more or less developed modification of nutrient salt
acquisition.

Mycophagy: — But as Frank continued to study mycorrhizae,
he saw digestion of the mycelium and was led to develop another
idea which may be termed "mycophagy". According to this concept,
the "fungus-eating plants" are able to draw their victim into the
protoplasm, there to tend it and make it large, and finally to digest
it, and thus the rich protein production of the fungus is made use of.
One of the two symbionts seems to have the advantage of the other
in that it appears as the raw material for the other. Stripped of its
fattening-pen implications, this concept has persisted. It was adopted
and modified by Magnus (1900) for Neottia: So far as purely
anatomical structures indicate, the physical significance of the
digestion-cells consists in an exclusive use for the higher plant by
which the substance-rich fungus is killed, digested and excreted ; the

Kelley — 152 — Mycotrophy

significance of the host-cell is an exclusive use for the fungus which
grows there purely as a parasite, injures the protoplasm, forms
"closed" organs which apparently serve to overwinter the plant. It
was adopted by Kusano (l^^O who said that the fungus, although
it acts as a parasite at times, becomes a victim of the orchid so that the
reciprocal exchange is not equal, for "Gastrodia is parasitic on the
fungus". And Cortesi (1912) believed the relation between fungus
and orchid is a case of helotism in which the fungus plays a subor-
dinate role. The orchid supports and nourishes the endophyte so long
as its presence is beneficial but it finally kills the fungus when flower-
ing and seed-production time arrives.

Prat (1934) came to the same conclusion about Taxus canaden-
sis. Symbiosis, he said, is scarcely the term to be used but disease,
for the invading fungus is limited by the host; and thus the tree
becomes a parasite on its parasite ! For Empetriim, Hasselbaum
(1931) concluded that there is no question of a mutual symbiosis,
but rather an attraction and destruction of the endophyte ; and Fraser
(1931) regarded two species of Lobelia studied as parasitic upon
their mycorrhizal fungus. For Vinca, Demeter (1923) saw an ad-
vantage to the host plant, according to Frank's concept, in myco-
phagy by which the host makes a certain gain in N ; but he did not
regard the symbiosis as ideal since the host-plant may be badly in-
jured. Besides, under cultural conditions the fungus-free plants
made decidedly better growth than the infected, indicating the my-
corrhizal relation was more of a parasitism.

Romellian Hypothesis: — In contrast to mycophagy is Romell's
(1939) hypothesis: that "the obligate mycorrhizal fungi associated to
conifers are not saprophytes decomposing soil organic matter, and
that they are energetically parasites on their host trees." MacDougal
& Dufrenoy (1944) take an opposite view-point, stating that "The
absorption of inorganic phosphorus from the soil by the fungus and
the stages of its metabolism terminating in the stele of the root, identi-
fiable origination in hyphae and translocation of auxin, vitamins and
amino compounds to the root tissues, together with the capacity of
isolated segments of mycorrhizal roots to survive and grow, like a
chlorophylless plant, establishes the non-parasitic character of the
fungus." Bjorkman (1944), furthermore, found that on so treat-
ing pine seedlings that they ceased to form carbohydrate in the root,
the fungus did not become a parasite : it simply ceased to "attack"
the root.

Lecture XI — 153 — Theories

Nitrogen Theory: — It is Frank's nitrogen theory that has been
received with most attention, a theory that is briefly stated by Lind-
QUiST (1939) : The nitrogen theory was first clearly formulated by
Frank (1894), [in a short discussion of the various possibilities of
significance of mycorrhizae, in which he supposes the fungus makes
available the N-compounds contained in forest humus and duff. More
probably, it was said, the fungus aids intake of humic compounds of
calcium], who considered that the beneficial influence of the mycor-
rhizal fungus consists mainly, for the higher symbiont in the pro-
vision of organic N-compounds for the latter. As basis of this con-
ception, Frank, etc., pointed out that the tissues of a mycorrhizal
tree are nitrate free. As it is known that the fungus can readily take
up NH4 and organic N-compounds, he considered it self-evident that
such compounds were taken up from nitrate-free or nitrate-poor
forest soils and into the mycorrhiza-bearing tree. The N-nutrition
hypotheses were extended later by numerous successors, as e.g. von
TuBEUF, MoLLER, MuLLER, Weiss, and especially Melin, who in the
years 1917-1927 conducted numerous investigations into the mycor-
rhizae of forest trees and their role as N-absorbing organs. . . . With-
out wholly renouncing the mineral nutrition hypothesis, he assigned
to the mycorrhiza no great role in the intake of mineral nutrients,
showing in this connection that the trees have ability to obtain rich
nutrition from the mineral soil by their deeply penetrating roots.

A similar reciprocity was claimed by Rayner (1927) for Calluna,
and she said: "it seems probable that Melin's conclusions respecting
the beneficial effects resulting from the presence of the mycorrhiza
in acid humus may be extended to the more specialized case of Calluna.
Under such conditions, the mycorrhiza of this species, and doubtless
of other cricoids, probably functions in a similar way to that of trees,
conferring on the host plant the power of drawing upon the organic
food reserves locked up in humus. . . . Like certain conifers and other
trees, Calluna and its allies are not strictly autotrophic in respect to
their nitrogen metabolism, and they are singularly well equipped for
successful competition in the struggle to obtain the requisite nitro-
genous food materials, whether in sandy soils, poor in organic con-
stituents, or in acid humus soils deficient in nitrates. They have
solved the problem of growth upon the poorest and most unpromising
soils, but they have solved it at the price of their independence."

Chemical Studies of the Nitrogen Theory: — We come next to
a chemical study concerning presence of NH^ salts in plants by
Weevers (1916), who found NH^ salts present in all species in-
vestigated except mycotrophic and insectivorous sorts. He said that

Kelley — 154 — Mycotrophy

mycotrophs apparently make use of organic N compounds through aid
of their mycorrhizae, this action taking place best on acid soils, the
occurrence of mycotrophs on alkaline soils therefore becoming
impossible.

Weyland (1912) initiated micro-chemical studies of the occur-
rence of salts in tissues of mycotrophic plants, and concluded from
his studies that, as a result of a limited transpiration stream which
results in part from activity of the fungus, P and K would be de-
veloped in concentrated form in the plants. The plants could thus
satisfy their Ca-requirement only by living on a calcareous soil from
which they evidently take the Ca. The theory of winning nutrient
salts through fungal aid, which the author (Weyland) had learned
from his teacher, Stahl^ broke down at this point. Nitrogen (Harn-
stoff) in orchid tubers was regarded as a metabolic product of the
root fungi. N-assimilation is to be considered as an essential func-
tion of the root-fungus.

An extended micro-chemical study of mycorrhizae was made by
Rexhausen (1920) who, seeking particularly to test Weyland's
conclusions, found that mycorrhizae were as rich in P and Ca as
were the fungus-free roots; also in protein content, although it
seemed somewhat greater in infected plants. He said that mycor-
rhizae are to be seen as isosmotically acting individuals which provide
the plants with all nutrient salts, apparently not merely P and Ca.
He thought that the fungus takes N from the higher plant because
of the ease of acting as a parasite in comparison with difficulty of se-
curing nutrient salts from the humus. Only in soils rich in nutrient
for the fungus can the higher plant throw off the fungus. But he
was sure of the nutrient salt provision for he repeats: "In the other
plants studied it may be stated positively that nutrient salts are brought
into the root through the hyphae."

Finn (1942) found in culture studies of white pine that seedlings
provided with mycorrhizae took in more N and K per seedling than
those of uninoculated controls. Earlier, Mitchell, Finn & Rosen-
DAHL (1937) had reported that their observations "indicate that the
benefits attributable to mycorrhizae, like their distribution in nature,
vary inversely as the concentration of readily available mineral nu-
trients in the soil. Seedlings lacking mycorrhizae are unable to exist
in very infertile substrates."

Importance of P: — In some cases at least, P seems of greater im-
portance than N. Thus McComb (1943) states that with pines,
especially P. Banksiana, good growth followed P fertilisation while
little or no response was obtained with N. "Without mycorrhizae

Lecture XI — 155 — Theories

pines acquired P with difficulty." "It is suggested that the stimulating
effect of mycorrhizal fungi on conifer seedlings is due to heightened
metabolism, associated in this instance with transfer of phosphorus
and growth stimulators from fungus to seedling" (McComb &
Griffith, Plant Physiol. 21:11-17, 1946). Young (1940) attributes
the benefit to the fungus, for "With the presence of adequate P for
fungus growth and the availability of a phosphatide supply, the
fungus is enabled to carry out the breakdown of raw organic matter
and transfer the products to the higher plant." Young presents a
diagram to show the effect of phosphate applications on the nutritional
cycle of the mycorrhizal fungus and the pine tree. Intake of P is
important to the higher symbiont in formation of phosphoric com-
plexes as seen in the next lecture.

Stahllan Hypothesis: — "The mineral hypothesis was first defi-
nitely formulated by Stahl (1900). He considered Frank''s con-
ception of a close relationship between mycotrophy and N-supply to
be false and claimed that, in soils poor in mineral nutrients, trees are
brought into competition with fungi and bacteria; and through my-
corrhizal symbiosis the tree is benefitted. Stahl^s hypothesis has not
met with favour amongst modern investigators except with Hatch,
who goes beyond Stahl in claiming the mycorrhizal relation to be
chiefiy a 'physical relationship', i.e., the chief significance of the
mycorrhiza is in its increase in absorbing surface."

Hatchian Hypothesis: — Hatch (1937), however, said his
hypothesis was "tentative" and applied only to "ectotrophic" mycor-
rhizae, and stated : "The mycotrophic relationship in pine, and pre-
sumably in other plants possessing ectotrophic mycorrhizae, is a
symbiotic mechanism which increases, chiefly by physical and therefore
by relatively non-selective means, the absorption of soil nutrients. . . .
The greater absorption capacity of mycorrhizal seedlings is brought
about by, and is proportional to, increases in the effective absorbing
surface areas of short-roots resulting from fungal invasion. . . . Trees
are dependent on symbiotic association with mycorrhizal fungi for
their soil nutrients and therefore for their existence in all but the
most fertile agricultural soils."

RouTiEN & Dawson (1943) sought to amplify the Hatchian
hypothesis, and after experimenting concluded that mycorrhizae
increase the salt absorbing capacity of the roots primarily by adding
to the supply of exchangeable H-ion derived in part at least from
carbonic acid. They found that development of mycorrhizae increased
the average rate of aerobic COg production of each short root from

Kelley — 156 — Mycotrophy

nearly 2-4 times the normal amount depending upon the degree of such
development.

The Hatchian hypothesis suffers from two grave disabilities: (1)
it fails to take into consideration the rooting medium of the myco-
trophic plant. The loci of mycorrhizae are most diverse and in many
cases are disintegrating organic residues, and it is rather puzzling to
understand how there can be a total intake of water and mineral salts
from a medium consisting chiefly of organic compounds. (2) The
Hatchian hypothesis (and all other hypotheses of mycotrophism for
that matter) does not make provision for an intaking mechanism.
Advocates of the root-hair hypothesis of plant nutrition have con-
sidered the root-hair in detail, and in so far as root-hair nutrition oc-
curs the process is fairly understood ; but advocates of mycotrophy
blithely ignore the structure which transfers materials from the soil
to the interior of the mycorrhiza, and leave one to assume that in some
way the substances jump in. The mycorrhiza apparently says : "Abra-
cadabra", and the deed is done!

Transpiration and Mycotrophy: — It must not be lost sight of
that the essential concept in Stahl's hypothesis, especially in the eyes
of his contemporaries, lay in the relation of mycotrophy to transpira-
tion. To Stahl, mycotrophy was most necessary to those plants which
have a limited transpiration stream and accordingly obtained less
nutrient salt in a "normal" way. A large stream of water passing
through the plant and being transpired from the appropriate organs
would presumably leave large quantities of salts in the tissues. Recent
work indicates that plants which transpire for long periods with
little elongation of the roots take in water through lenticels, breaks
around branch-roots and wounds (Addoms, Plant Physiol. 21 : 109-
111, 1946). Plants which have a limited transpiration stream and
hence more difficulty in securing nutrient salts are those which have
"sugar leaves" rather than "starch leaves". The advantage of starch
formation lies in the lessened trouble in assimilation and the greater
ease of transpiration, while addition of sugar increases the difficulty
of the latter. Thus, plants which secure nutrient salts with difficulty
are those which do not excrete liquid water from the leaves ; and of
these Stahl listed a number. Stahl perhaps got his idea from F.
ScHWARz (1883), who said that conifers and cupressineae have the
leaf reduced and cuticularized, and hence have a lesser transpiration
stream: in these trees root-hairs are lacking and the water and salt
requirements are met by parasites (Schmarotzern).

BuRGEFF (1909) adopted Stahl's views on transpiration in
orchids, apparently without much thought ; while he found feminine

I

Lecture XI — 157 — Theories

supporters in Busich (1913) for asclepiads, and Kerkichu (1930)
and Stajanow (1916) for orchids. Stahl's idea in regard to orchids
was that the orchids can take in and transpire only a small amount
of water but the symbiotic fungi have a greater osmotic power and
the orchids owe to this property a greater inflow of water and the
salts dissolved therein.

Objections to the Stahlian Hypothesis: — Opposition to Stahl's
view developed. Tubeuf (1903) pointed out that Stahl's view
could not apply to endotrophic mycorrhizae because, he said, there
is no connection of the endotrophic mycorrhiza with the surrounding
soil, meaning apparently that individual hyphae are insufficient to
meet the needs of the plant. Stahl was at fault, too, in regard to
liverworts; for Ncmec (1899) had asserted that Marchantiaceae,
being starch producers, could not have endophytes, which were to be
expected only with the sugar-producing Jungernianniaceae; and
Stahl had seized upon this suggestion as fitting into his hypothesis.
But actually, endophytes are commonly present in the Marchantiaceous
liverworts. Then, as to orchids, Fuchs & Ziegenspeck (1925)
pointed out that in orchid mycorrhizae the transformation of cortical
cells into digestion-cells interferes with water-transport, hence
lessened transpiration is not compensated by mycotrophy but is
rather caused by it.

Opposition to Stahl's hypothesis developed also in regard to
his ideas in relation to soil sterilization. Existence of a struggle for
nutrient salts was posited amongst roots of all plants, and myco-
trophic plants were supposed to gain an advantage over concurrent
fungi through aid of mycotrophic fungi. The existence of such a
struggle was thought by Stahl to be demonstrated by the fact that
autotrophic plants do better in sterilized than in unsterilized soil.
But Neger (1903) conducted experiments that led him to conclude
that thriftier growth of plants in sterilized soil is to be attributed
mainly to the increase in nutrient salts caused by sterilization and
not (or only to a minor degree) to the misconceived battle against
soil fungi.

Bjorkman's Hypothesis : — Coming still closer to an explanation
of mycotrophy, Bjorkman found a connection between the products
of photosynthesis in the host and mycorrhizal formation. A vigor-
ously growing plant in an environment moderately deficient in N and
P, or both, and exposed to optimum illumination forms abundant
mycorrhizae because there is abundant reserve of assimilate in the
root tissues for the mycorrhizal fungus to use. The author's only

Kclley —158— Mycotrophy

acquaintance with this hypothesis is through Romell's (1944) review,
from which the following is quoted: "the mycorrhizae seek soluble
carbohydrates in the root, and they consume them for energy, but
they find them only if there is a surplus of carbohydrate in the root.
Whether or not there is any surplus depends on the head-start photo-
synthesis (carbohydrate formation) has attained in the plant before
the formation of albuminous substances. This advantage will be
small or nil if there is an abundance of all plant foods including
nitrogen and phosphorus, so that there will not need to be a shortage
of materials when albuminous substances are built up. Then there
will be little for the fungi to seek after in the root, and mycorrhizae
will be formed sparingly or not at all. On the other hand, should
there be a deficiency of nitrogen or phosphorus foods, there can
easily arise a surplus of carbohydrates so that the formation of al-
buminous substances falls behind photosynthesis. With a moderate
lack of nitrogen or phosphorus or both, mycorrhizae can form
abundantly, but if there is a serious lack of nitrogen or phosphorus,
sooner or later photosynthesis will become weak also (phosphorus
especially strongly influences carbohydrate formation in plants) so
that the surplus of assimilated carbohydrate becomes less for that
reason, and mycorrhizae are formed more rarely. Similar reactions
occur if light becomes weak, Mycorrhisal formation is, expressed
briefly, a result of and a sign that there is a certain surplus of energy-
giving nourishment in the host plant."

Carbonaceous Theories: — Besides the theories of mycotrophy

which posit the intake of inorganic salts, there are not lacking

theories that connect it with carbon. The idea that roots take in carbon

has been familiar from the days of Liebig and his "humus theory"

which posited a normal intake of carbonic acid by rootlets of seedling

plants. The same idea appears in a paper by Breal (1894) who, in an

experimental study concerned chiefly with cereals, concluded that

these plants are able in efifect to take up organic carbon materials

through their roots. Beauverie (1902) studied the Hverwort, Cono-

cephalus, and found that endophytic infection is most pronounced

where humus is most abundant, and where humus is lacking there is no

mycelium and thallus is small sized. By experiment it was indicated

that the plant secures part of its C from the humus. For the orchid

Gastrodia, McLuckie (1923) came to the same conclusion: "Gastro-

dia is an elaborate example of symbiosis in which an Angiosperm is

associated with a fungus and a bacterium ; and is directly or indirectly

dependent upon its endophytes for its carbonaceous and nitrogenous

foods." Other examples are given by Young (1940).

Lecture XI — 159 — Theories

Hydrocarbon Hypothesis: — That nutrient exchange in mycor-
rhizae is carbonaceous rather than nitrogenous was advocated by
McLennan (1926). "A critical discussion of [some of] the earlier
work on mycorrhizae, more particularly that dealing with the physical
relations between the two forms, discloses the fact that the most
generally accepted ideas of this relation are those connecting it with
nitrogen fixation without any real evidence that such is the case. . . .
The demonstration of many infecting strands [in Lolium mycor-
rhizae], together with the appearance of fat, firstly in the conducting
and travelling hyphae of the root, with its subsequent removal to the
sporangiole, and then to the host-cell, accompanied by collapse and
shrivelling of the fungal mechanism, have led to the conclusion that
a metabolic exchange takes place from the fungus to the higher plant,
with the result that the latter obtains a supply of fat or oil." Knud-
son's results were thought to favour this hypothesis and "The idea
that the exchange is carbonaceous rather than nitrogenous is also
compatible with Bernard's suggestion" of the relation between
tuberisation in plants and the presence of endotrophic mycorrhizae.

As a postscript we may add: "Although these homogenous
globules do not stain black with osmic acid after bichromate, they
have, nevertheless, been proved to be fat globules."

Carbohydrate Hypothesis: — Another carbonaceous hypothesis
was proposed by Young (1940) as a "mycorrhizal theory regarding
the cause of fused needle disease" in pine. "According to this hypothe-
sis, normal mycorrhizas supply the tree with an essential part of their
carbohydrate supply, and it is to the inefficient functioning of the
mycorrhizas in this respect that the fused needle condition is due. The
supplying of additional phosphorus to soils low in this element results
in a more abundant phosphatide excretion from the pine roots,
thus stimulating normal mycorrhiza formation and bringing about a
satisfactory balance of conditions for correct mycotrophic activity.
The amount of vegetable detritus present is important in this respect,
as it is from this source that the carbohydrate supplied to the higher
plant by the mycorrhizal fungus is obtained. The addition of phos-
phates to the soils in question stimulates the growth of natural
vegetation, and thus aids the development of the necessary supply
of vegetable detritus."

This explanation was not accepted by Neilson-Jones (1941),
who ascribes development of needle fusion to a sudden shortage of
water in the plant as the leaves start to expand, due to a failure to
produce mycorrhizae at the critical juncture. But the hypothesis of
carbon intake through mycorrhizae remains to be experimentally

Kelley — 160 — Mycotrophy

evaluated. Young (1941, p. 91) makes it clear that it is not merely
C that the plant receives from the soil through its endophyte but
"Inorganic salts and perhaps nitrogen compounds are probably also
supplied to the plant."

Growth-Promoting Substances: — In more recent years atten-
tion has been turned to growth-promoting substances, and Lindquist
(1939) presented a Growth-substance Hypothesis, based on his
studies which indicated an excretion of a substance from mycorrhizal
mycelia that stimulated growth of spruce in pure culture. "The
mycelia . . . influenced so markedly the nutrient liquid into which they
were drawn during these experiments that a decided increase in
growth of needles, as well as of stem and root, could be marked."
Lindquist considered it established that "the nutrient materials
derived from fungi are of essential, indeed of vital, significance."
BuRGEFF (1934) also found the higher plant stimulated by the fungus,
in this case specialized orchids of the Vanda group, which always
develop slowly in absence of their natural symbionts. In promoting
growth, the dead fungus was found to be just as effective as living
hyphae, because of presence of a "growth- factor" resembling "Bios
11" although its chemical nature was otherwise unknown. In signi-
ficant experiments, Noggle & Wynd (1943) tried the effect of
various growth-promoting substances, and found "good germination
and excellent development of the seedlings when nicotinic acid
(P - P factor) was supplied in the nutrient medium."

But RoMELL (1939) rejected Lindquist's hypothesis that "the
chief function of mycorrhiza formation would be the exchange of
growth-promoting substances between the symbionts", and "returned
to Frank's original ideas". Yet it is established that growth-promot-
ing substances do influence mycorrhizal fungi. Melin (1939) found
increased growth with aneurin in synthetic (glucose-containing)
medium; yeast filtrate gives even better results while biotin and
inosit failed to improve growth except in combination with aneurin.
Aneurin (vitamin B^) also gave better growth when added to cul-
tures of endophyte of Arum (Magrou, 1939). Later, Melin (1942)
found that aneurin is replaceable by its pyrimidin and thiazol compo-
nents, and equimolar quantities in synthetic cultures of certain mycor-
rhizal fungi. The development of M. r. atrovirens, which is aneurin-
autotrophic, is retarded by addition to the medium of aneurin or its
components, especially pyrimidin alone or mixed with thiazol. Ex-
perimenting with orchids, Meyer (1944) concluded that thiamin
(vitamin B^) substantially aided growth of seedlings.

Lecture XI — 161 — Theories

Working with aqueous extracts of fall-litter, Melin found that
"all types of litter contained substances soluble in water which
favourably influenced mycelial growth of thiamin-heterotrophic fungi
(mycorrhizal Hymenomycetes and Gasteromycetes, as well as litter-
decomposing Hymenomycetes) in a nutrient medium containing
sugar, salts and thiamin. The litter extracts likewise exercised a
favourable influence on the development of thiamin-autotrophic soil
fungi." These results apply only to tree-litter examined and not to
litter from the grass, Glyceria. The litter extracts appear to contain
growth substance like Robbins' "Z" factor (Symbol. Bot. Upsaliensis
8(3) :1-116, 1946).

Limitation of Mycotrophic Hypotheses: — It must be realized
in any study of mycotrophic hypotheses that the protagonists of each
hypothesis have been specialists in some one field of mycorrhizal
study, and their hypothesis naturally reflects experiences with their
own material. It may be that each investigator is correct, as un-
doubtedly he is so far as his understanding of his own data is con-
cerned, but no one person has yet had a broad enough grasp of all
phases of mycotrophy to develop an explanation which will fit all
cases.

Again, it must ever be remembered that a mycorrhiza is not a
static thing: Endrigkeit (1937) has neatly expressed the case:
"Since the parasitic acquisition of nutrients by the fungus is of a
very restricted order in both mycorrhizal groups, the higher plant is
evidently the chief gainer by the association until the activity of the
roots begins to decrease with age, the balance inclines in favour
of the fungus. Comparative membrane and permeability observations
on plants in a colonized and uncolonized condition revealed a pro-
gressive loss of independent assimilatory capacity of the roots with
increasing fungal activity."

The Intaking Mechanism: — The point at which current theories
of mycotrophy break down is with respect to the mechanism for
intake of liquid materials from the habitat. It is after all of little
consequence whether the plant is to receive N or P or any other ion
if there is no means of transport for these substances. Of course
the plant does receive them, but not much attention is paid to the
method or the apparatus by which the substances enter the mycor-
rhiza or mycotrophic organ. The presumption is that they enter
through communication-hyphae, but so far as we are aware there is
not a single research devoted to the structure and functioning of these
hyphae. They seem to have been forgotten by the protagonists of

Kelley — 162 — Mycotrophy

the various mycotrophic theories ; yet they are utterly essential to any
theory of mycotrophic nutrition. To formulate a theory of nutrient
intake and ignore the intaking apparatus is like baking a cake — with
the baking-powder omitted.

Ectotrophic Intake: — It is evident that there are two very
distinct sorts of mycotrophy, as has been indicated from early days.
In the first sort, the ectotrophic (if there is indeed a true ectotrophic
mycorrhiza), the surface of the mycorrhiza is covered with a fungal
mantle from which extend out numerous hyphae that assume various
shapes, sizes and forms. This mechanism would seem to be suited
to an intake of liquid materials, — water and dissolved salts, — just as
Frank originally observed. Yet even among these mycorrhizae there
is a great difference between various sorts as briefly but well shown
by WooDROOF (1933) for pecan. Here the surface of the mycorrhiza
is in some forms covered with short setae that seem tO' have the
qualities of root-hairs and may function as such.

Root-Hairs Versus Setae: — -In any report of root-hairs on an
otherwise mycorrhizal root, a due scepticism must be maintained
until there is positive proof that the structure is a root-hair and not
a fungal hypha. Short-roots are frequently to be observed with
what appears to be a firm epidermis from which extend trichomes
in the form of root-hairs, but on closer inspection it is to be seen that
the "epidermis" is composed of closely appressed hyphae from which
setae extend. Thus, MacDougal & Dufrenoy (1944) remark that
the mycelium of the endophyte of Corallorhica sends "branches out-
wardly through the epidermal cells of underground coralloid branches
in simulation of the arrangement of root-hairs." In cases of doubt
the material must be embedded and sectioned ; and lacking such
demonstration we remain unconvinced in reading any report of the
presence of root-hairs on mycorrhizal roots.

Endotrophic Intake: — For the endotrophic mycorrhiza there is
a very dift'erent structure. There is no mantle of mycelium, no myco-
derm, no setae; and the only hyphae that connect the mycorrhiza to
the soil surrounding are the so-called communication-hyphae. If water
and salts are to enter the mycorrhiza they must come through these
hyphae or through the uncuticularized portions of the exposed root.
But could substances come through these hyphae? Recall that the
endotrophic mycorrhiza is the prevalent sort, the one by far the
most common, especially if we consider the functioning of the ectendo-
trophic mycorrhiza as in part included here. Recall also that the

Lecture XI — 163 — Theories

hypha is in effect a capillary tube of relatively great length. Now it
is true that water passes readily through a capillary tube, but only
if it is empty. The fungal hypha is not empty but filled with a more
or less viscous colloid protoplasm. Furthermore, in the hymenomyce-
tous hypha there are septae at intervals, in the soil portion of the
mycelium, at least, although cross-walls are rare inside the mycorrhiza.
(It is to be noted that MacDougal & Dufrenoy [1944] , states : "Pads
of material at the pores of cross-walls of hyphae . . . were observed.
Possible similarity of composition and functioning with sieve plates
of higher plants is noted.") Here there are two obstacles to the
passage of liquids through the "communication-hyphae", — colloid
and cross-walls ; and unless different physical principles can be ad-
duced than those which ordinarily obtain, we can only conclude that
the "communication-hypha" is not an efficient apparatus for transfer
of water and mineral salts, if it is used in that way at all.

"Individual hyphae, as the demonstrations of Frank indicate, can
not suffice for extensive transportation paths" (von Tubeuf, 1903).
"The endotrophic mycorrhizae have only a slight independent capacity
for absorption as compared with the autotrophs." (Endrigkeit,
1932).

Hyphae as Nutrient Conveyors: — Contrast the "communica-
tion-hypha" with the root-hair. The latter is constructed for intake
of liquids : It is a thin-walled cell-extension, relatively short and of
considerable diameter as compared with its axial length. It has a
minimum of protoplasm and contains a large sap vacuole that is so
constituted as to aid osmotic action. The communication-hypha is
the opposite to all this : Its wall is not of cellulose ; it is tremendously
long in comparison with its diameter ; it is densely filled with proto-
plasm with a minimum of vacuole, and it may even be boxed off at
intervals by septae.

Obviously, the communication-hyphae function otherwise, namely
in bringing in of elaborated organic substance to the mycorrhiza where
it is dealt with in an ordinary biological way.

Teleology in Mycotrophism: — In spite of scientists' sensitive
denials of teleology, the current theories of mycotrophy are decidedly
teleological. The mycorrhizal apparatus is represented in some sort as
an automat wherein the mycorrhiza drops a starch-grain in the slot
and takes out a dish of nitrogen compounds or of phosphorus salts.
It is the author's opinion that ordinary processes of biology are
sufficient to explain the mycotrophic reactions without attributing needs
of salt absorption, etc., which the higher plant seeks to satisfy ; and this
opinion is elaborated in the next lecture.

LECTURE XII
MYCOTROPHIC PHAGOCYTOSIS

Significance of the Term: — In 1905 Bernard had arrived at
the conclusion that orchid "symbiosis is in some sort a serious and
prolonged disease, intermediate between a fatal malady and complete
immunity." This idea was elaborated in 1909: Bernard sought to
show, as VuiLLEMiN said, the common characters between infectious
diseases of animals and those of plants, and the narrow bonds which
unite the states of symbiosis and disease. He transported to botany
the medical terms of vaccination, immunity, phagocytosis, etc. The
word "phagocytosis" connotes three distinct actions, vis. (1) Attrac-
tion of foreign bodies, (2) their capture or active penetration in the
element into which they are drawn, (3) their intracellular digestion
and assimilation. But for Bernard the word "phagocytosis" implied
nothing more than intracellular digestion ; and he dissociated the
first two acts from the third. He regarded the Rhisoctoniae and their
orchids as two antagonists (1909&), developing their means of
attack and defense; while symbiosis {i.e., mutualism) represents an
immunity attained by phagocytosis. The plant makes use of all its
means of defense in order to preserve its essential tissues.

After this vitalistic statement, the mechanics of the reputed de-
fensive action are related. The formation of mycelial coils in orchids,
of Gallaud's arbuscles in mycorrhizae of Allium, of Janse's spor-
angioles in roots of plants in Java, are all considered phenomena of
agglutination due to a humoral property of phagocytotic origin. But
Bernard did not overlook the fact that clotting of hyphae took place
in cultures of Rhisoctoniae where there could be nO' question of lytic
action induced by an orchid tissue. Bernard's untimely death put a
stop to these significant studies.

It should be noted that there is a distinction between phago-
cytosis, which is probably a proteolysis in large part, and toxicity.
MacDougal & Dufrenoy (1944) neatly contrast this difference:
"The action of the 'humoral' secretion ... is to be distinguished from
toxicity by the fact that in the first case the secretion simply blocks
some of the processes of cell metabolism but does not destroy the
mechanism; in toxic action (as in pseudomycorrhizae) a theoretical
secretion causes irreversible and fatal changes in the cytochemical set-

Lecture XII — 165 — Mycotrophic Phagocytosis

up of living cells of the organism affected. In parasitism, in addition
to possible anatomical and cytological damage, a fatal extraction of
nutritive material may ensue."

Older Descriptions of Phagocytosis : — New names do not make
new processes, and that of phagocytosis had long been known, although
not under that name. Reissek (1847) observed ellipsoid bodies in
Neottia that later turned yellow, — in other words, clot-formation;
and in Orchis Morio he noted the process was especially active in
autumn and early winter. He described and figured peloton forma-
tion with remarkable clearness. Schacht (1854) saw that the fungus
uses starch in orchid rhizome and root-cells; but Prillieux (1856)
differed with Schacht: The 2-3 outermost cortical layers of cells
in Neottia were filled, he says, with a yellowish-brown matter which
Schacht had considered as an index to death of cell. Prillieux
found it in a great number of orchid genera, in cells which retain their
nuclei ; but he noted that the matter seemed to diminish in roots which
have vegetated a long while, from which he inferred that the brown
matter served for nutrition of the plant. The material, he said, is
probably nitrogenous. The infected orchid cell nuclei are of great
size and often have 2 nucleoli. The cells having brown matter regu-
larly contain filaments wound without order about the central mass
in each cell. Reinke (1873), noting the matter or "slime" in orchid
cells, ingeniously supposed that it is a "Schwellkorper" or apparatus
for maintaining sap concentration.

Phagocytosis in Peloton Mycorrhlzae : — The process of phago-
cytosis in the peloton mycorrhizae, found chiefly in orchids, is accord-
ingly as follows : Hyphae penetrate in part through root-hairs or in
part directly through cell-walls, and enter the cortex, finding entrance
through passage cells where an exo-dermis is present. Hyphae pass
freely through cell walls, a marked constriction sometimes being
evident in the hypha at the point of passage of cell- wall (Dangeard,
1898) which grows up about the hypha (Burgeff, 1932) to form a
wall-tubule or "Rohrentiipfel." This organ Burgeff considered one
which, by differences of suction force, passes water or solutions of
low sap concentration through the hypha. Passage of hyphae through
cortical cell- walls is sometimes prevented by thickening of the walls
to 6 or 7 ji, by undergoing an excessive cuticularization (Burges,
1939).

At a certain distance from the central cylinder, depending on the
sort of orchid, the hypha ceases to grow forward and begins to coil.
Burgeff (1909) observed coiling on aerial hyphae when a drop of

Kelley

— 166 —

Mycotrophy

water was excreted : a hypha being drawn into the drop would be
prevented by surface tension from growing out of it again, and
would thus continue its growth by coiling within the water drop.
Perhaps something of the sort occurs in orchid cells, those cortical
cells nearer to the central cylinder having an higher concentration of
ions in the cytolymph and accordingly a greater surface tension in
the protoplast. Regardless of explanation, the hypha coils in cells
of the cortex, the nucleus of the cell remaining intact but becoming
enlarged, hypertrophied and even dividing amitotically. According
to most authors, hyphae never penetrate raphide cells but Busich
(1913) says that the fungus is not warded off by calcium oxalate but
on the contrary forms it.

Later the hyphal coils degenerate and release their content into
the host-cell. "As with many other orchids, the process of ingestion
of the hyphal coils is preceded by a turning point at which the h.i.c.

Fig. 12. — Portion of a cross-section of the ectendotrophic
mycorrhiza of Cornus jlorida, showing hyphal coils or
pelotons (p) and vesicles (v).

of the hyphal clumps reached a maximum of /'H 6.2." (Hamada,
1939). The actual breaking down of the hypha seems due to the
action of a proteolytic enzyme (Burges, 1939) and results in the
formation of a more or less homogeneous yellowish mass in the
centre of the host-cell. This is the "yellow body" or "matiere
brunatre" or "gelbliche Stoffe" of earlier investigators. The yellow

Lecture XII — 167 — ■ Mycotrophic Phagocytosis

clots are highly refractive, of irregular form, and are very resistant to
acids and alkalis. In concentrated H2SO4 they dissolve after a few
days, w^hile in KOH they swell somewhat, and on treatment yellow
drops appear on the periphery which with osmic acid turn dark brown,
indicating presence of fat or oil (Wahrlich, 1886).

The material brought into the orchid cells has been termed pro-
teinaceous, and the hyphae were called "protein hyphae" (Eiweiss-
hyphen), but according to A. Fuchs (1924) they are actually glyco-
gen hyphae. The breaking down of these hyphae, according to Burges
(1939) is due to a loss of vitality. The addition of small quantities
of extracts from tubers, stems, leaves and roots of orchids tO' young
cultures of the endophytic fungus resulted in all cases in an inhibition
of fungal growth and complete decomposition of hyphae within 3-4
days, root extracts being much less toxic than those prepared from
tubers or stems. Sap of host-cells, withdrawn by means of a micro-
pipette and added to blocks of agar smear-cultures produced visible
changes within 24 hours, and at the end of four days some empty
hyphae could be seen.

Phagocytosis in Arbuscular- Vesicular Mycorrhizae: — My-

cotrophy in this sort of mycorrhiza was described by Boulet (1910)
from cultivated fruit trees. He said the habit of the endophyte is
very constant: Mycelium traverses the piliferous layer, penetrates
into cortical cells, ramifies, but seldom penetrates further than %
width of the cortex. The endophyte apparently lives upon the starch
reserve of the cells which harbour it, for the starch reserve disappears
from these cells. In the most internally placed layers part of the
hyphae continue development in the cells while another part insinuate
themselves into the intercellular spaces, filling the cavities. The my-
celium frequently contains reserve. In the region of the endodermis,
which is never penetrated, the hyphal wall is partially dissolved and
inclusions extravasated. On certain filaments vesicules are abundant
and some are intercellular (size 100 /x x 54 jx) while others are
intracellular (57 /x x 36 /x). The intercellular hyphae, by a more
or less regular dichotomy, form dense coralloid branches called
"arbuscles" by Gallaud. The branches often terminate in sporan-
gioles which at times are so numerous as to fill the cell cavities. They
disorganize rapidly to a granular, somewhat floccose, mass and finally
into scattered granules ; or, even more frequently a solid mass is
formed and degeneration seems checked.

Janse, who invented the name of "sporangiole", speaks of them
as present in a number of the plants he describes ; e.g., in Dysoxyhim,
a member of the Meliaceae, he said the fungus penetrates to the cor-

Kelley

— 168 —

Mycotrophy

tex, showing swellings before it passes the cell wall and later branch-
ing to form terminal sporangioles which at first are filled with
spherules, and finally to a gummy mass. Nuclei of cells which contain
them are much larger than those of other cells nearby. Again, in
Begonia 7'obusta, the fungus produces sporangioles which at maturity
contain very fine granules which finally diffuse through the cell. In
Helicia (of the Proteaceae) large cortical cells are infected by hyphae
that form sporangioles and later free their content to form a gummy
mass.

Fig. 13. — Portion of a section through mycorrhiza of
Abies balsamea indicating some of the mantle or myco-
clena to the right, and within, fungal hyphae and arbuscles.

In Vitis (Petri, 1907) the intracellular mycelium shows the fol-
lowing development: (-Z) A net of fine hyphae is formed about the
starch grains, which are soon dissolved. (2) Nuclear elements in
the ends of these fine hyphae undergo a differentiation which recalls
the synkarion-phase of the basidia during karyogamy. (3) A great
quantity of proteinaceous material accumulates in the ends of these
hyphae. (4) By chemical transformation, these hyphal branches and
their content are gradually transformed into prosporidia.

In asclepiads, Busich (1913) found a special sort of vesicle
termed a "glomerule" ; and she also found formation of vesicles out-
side the root. In Vinca (Demeter, 1923), penetration of the epider-

Lecture XII — 169 — Mycotrophic Phagocytosis

mis is direct and the fungus passes through "Kurzzellen" or passage
cells intO' the cortex where, in consequence of presence of fungus, the
starch dwindles. Inter- or intra-cellular hyphae become swollen or
wurst-shaped, either terminal or intercalary. In age, in addition to a
number of nuclei, they have great fat vacuoles with protoplasm be-
tween, like cross-walls. When intracellular, vesicles are always
terminal, and the small glomerules described by Busich were never
found by Demeter. The content of the vesicle may be resorbed into
the hypha as there is seldom a cross-wall evident; and the function
of the vesicle is apparently to serve as a temporary storage organ.
"Germination" of the vesicles has been reported, said Demeter, by
Bernard and Busich only; and the very infrequence bespeaks a
slight significance for the germination. But more important in the
lives of both symbionts are the arbuscles, and both the simple and the
compound sorts described by Gallaud are found in the Apocynaceae
and Asclepiadaceae. Simple arbuscles are always terminal, formed
at the end of hyphae which have penetrated cells. In the end branches
in young stages of arbuscles there are little granules arranged in
nebulae, and staining deeply with haematoxylin. From their origin
these nebulae appear to be protein precipitates. As a result of action
of free H-ion in the cell-sap, the tips of the arbuscles burst and empty
their content into the host-cell. It is possible, said Demeter, to form
these "plasmoptyse" of the endophyte in pure culture, the action oc-
curring in vitro at an optimum acidity of 0.025 N HCl, and on the
basis of this observation, the name of "Plasmoptysen mycorrhizae"
was chosen. "Sporangioles" are merely the last structureless residue
of the arbuscles which have been made harmless, a residue which is
finally resorbed.

Just as the hyphae can be broken down in HCl and the content
extruded as in the "plasmoptyse", so in pure culture Demeter found
that by use of different concentrations of sugars he could produce
peculiar stuntings of growth which recalled arbuscle formation.

Phagocytosis in Ericaceous Mycorrhizae: — Kamienski
(1884), in his pioneer work on Monotropa, saw that the roots of this
plant are much branched and interlaced, and fragile; the epidermis,
being covered by a fungal mycelium of septate hyphae that form a
pseudoparenchymatous mantle two or three times as thick as the
epidermis. The fungus lives on the surface and never penetrates liv-
ing cells except sometimes in older portions where those cells are
filled with a brown "tannin" content. In older parts of Monotropa
roots the epidermis disorganizes at the same time as the mycelium
develops. It is evident that the epidermal cells play an important

Kelley

170 —

Mycotrophy

role, for all the interior cells are cut off from the exterior by a fungal
mantle : "Consequently, there is no other way by which the nutritive
solutions may pass and provide the roots of Monotropa except the
way of the mycelium. This is composed of vegetative filaments of
which those that neighbor the epidermis are applied so closely to
these cells that diffusion seems not only possible but absolutely exists.
Monotropa", said Kamienski, "is thus able to nourish itself by the
mediation of this fungus."

But MacDougal (1900) said that in the Monotropas, vesicles,
sporangioids and sporangioles fill the cells, and "probably serve as
organs of interchange". Francke (1934) said that infection is

Fig. 14. — A cell from mycorrhiza of Allium sphaerocephalus showing an
arbuscle which is breaking down to form sporangioles {Redrarvn from Gallaud,
Rev. gen. Bot., 1905).

limited to the epidermis, one hypha only penetrating a cell, going to
the nucleus where it forms a plasmoptyse, followed by phytophagy
in which there is no evidence of excretion. No great changes were
observed in the host nucleus.

In Empetrum (Hasselbaum, 1931) digestion also begins next
the nucleus and proceeds outward. During digestion the cell nuclei
become amoeboid, its nucleoli are split up and yielded to the plasm as
producers of ferment. A distinction between host- and digestion-
cells is observable, the cells being of different morphological origin:
in the host-cells the fungus forms thick coils of hyphae which are
not digested while in the digestion-cells there is a sporangiole mycor-
rhiza "destined to digestion".

In Calluna, Rayner (1927) described intracellular digestion of
mycelium. "Throughout the growing season the mycorrhiza cells
exhibit active intracellular digestion of mycelium with disappearance
of the resulting — and presumably soluble — products. The nearer to
the apical meristem, the more rapidly is digestion initiated. Its onset
is marked by the usual signs of cell activity — increase in size and

Lecture XII

— 171-

Mycotrophic Phagocytosis

chromatin content of the nuclei often accompanied by deformation,
'clumping' of the mycelium about the nucleus, disappearance of the
sharp outlines of individual hyphae, and the gradual conversion of
the hyphal constituents from the region of the nucleus outwards, to
a structureless mass possessing strong stainability. The last stages
in the process are marked by shrinkage of the nuclei and disappearance
of the stainable contents. This intracellular digestion of mycelium is
a continuous process observable throughout the vegetative season from
early spring to> late autumn. The proportion of cells in the active
mycelial condition or undergoing digestion at any given moment
varies with the time of year, the age of the root, and possibly also
with the season and other external factors."

Phagocytosis in Ectotrophic Mycorrhizae: — Laing (1923)
makes the statement that there is no evidence of digestion in ecto-

pic. 15. — Portion of a longitudinal section through
a mycorrhiza of Pferidiitm aquilinum, showing a
plasmoptyse stage in breaking down of hyphae (Re-
drawn from LoHMAN, Univ. Iowa Studies in Nat.
Hist, V. 9, pi. II, fig. 10).

trophic mycorrhizae of conifers. But the whole question of ecto-
trophism remains in doubt for if it is true, as Melin (1923) re-
marked, that in older researches the delicate intracellular hyphae may

Kelley — 172 — Mycotrophy

have been overlooked, then there is nO' such thing as a true ectotrophic
mycorrhiza with the mycehum closely surrounding the rootlet but
not penetrating its cells. Were such a mantled root to exist as in-
dicated, then mycotrophy in such a case would inevitably consist in a
provision of the higher plant with materials taken directly from the
soil (since the higher plant is otherwise isolated from the soil by
felted hyphae) while the fungus would gain nothing except a con-
genial site for mantling its hyphae. But if the "ectotrophic" mycor-
rhiza actually has hyphae extending into the host-plant's cells, then
its mycotrophism is the same as for other sorts of mycorrhizae,
namely a mycotrophic phagocytosis. In the absence of definite in-
formation our judgment must remain suspended ; yet we can make
one incidental observation, that free-hand sections are useless for my-
corrhizal study and researches based on this method are necessarily
invalidated.

If we distinguish a mycorrhiza as "ectotrophic" when it possesses
an Hartig net and ignore the question of infection or non-infection,
then the cases described by Melin as ectendotrophic may be utilized
for this category. In Larix (Melin, 1922&), three phases of mycor-
rhizal formation are distinguished: {!) The fungus penetrates in-
tracellularly into the roots and forms individual hyphae or knots;
(2) then the intracellular hyphae are digested and the mycelium pene-
trates intercellularly, while {3) finally the fungus lives almost ex-
clusively externally and the mycorrhiza becomes mainly "ectotrophic".
In Pinus sylvestris and Picea Abies (Melin 1922a) the hyphae grow
principally in the interior of the cortical cells where they form a
pseudoparenchyma of the same appearance as in the fungal mantle of
the completely developed mycorrhiza. Later the Hartig net and the
fungal mantle are formed.

In Betula and Populus, Melin (1923) described the mycorrhiza
as consisting of (i) an hyphal mantle; (2) a "palisade" layer in
which there is an Hartig net and intracellular hyphae of two sorts :
(a) Haustorial hyphae which are very thin (1 /*) and grow in a
tortuous course: they are seldom septate, are plasm-poor and some-
times fragment while at other times they form grape-like bodies.
{h) Protein (Eiweiss) hyphae may attain 10 /u, thickness. They
extend longitudinally in the palisade cells and grow into neighbouring
digestion cells or penetrate several palisade cells. They are at first
very rich in plasm and protein and contain several (up to 8) large
(3 /i) nuclei which have apparently 12 chromosomes. They seldom
branch, (i) Digestion layer, which is bounded by an endodermis
provided with tannin and starch wherein is no infection.

Lecture XII

173 —

Mycotrophic Phagocytosis

Melin concluded: "The anatomical structure shows that the
higher symbiont suffers no injury from the fungal hyphae. Quite the
contrary, some of the hyphae are later digested, whereby the higher
symbiont obtains some nutrient while the fungus, through its haus-
torial hyphae, derives some nutrient-material from the higher sym-
biont. Finally, a nutrient-interchange takes place between the Hartig-
net and the palisade layer which long keeps both tissues alive".

These descriptions inform us of the ectotrophic (or ectendo-
trophic, if we choose) mycorrhizae in Sweden. From the other polar

Fig. 16. — Some cells from mycorrhizal cortex of
Fraxinus americana, in which the fungal reserve,
which overwintered, is largely broken down and
partaking of a plasma stain. Note enlarged nuclei.

extreme, from the Cape of South Africa, comes an exactly similar
report. Smith & Pope (1934) state with reference to mycorrhizae
of Eucalyptus: All the main internal features (the layered mantle,
the palisade-like epidermal cells with "Hartig-net" mycelium) are
paralleled in Melin's descriptions of other tree mycorrhizae. The
fungus is usually present inside cells of epidermal layer and the
outermost cortical layer but rarely occurs in any deeper layer. Intra-

Kelley — 174 — Mycotrophy

cellular digestion of hyphae is exhibited with a clarity unusual in tree
mycorrhizae.

Endrigkeit (1937) says that at no time is there intracellular
digestion in Pinus. In the monograph on mycotrophy in Pinus
(Hatch, 1937), we learn nothing of the method of intake of nutrient,
the mechanism of intake, or of possible phytophagy.

Limitation of Endophyte: — Confinement of the endophyte to
a certain region of the mycorrhiza is a common observation. It was
the basis of the early distinction between ecto- and endotrophic my-
corhizae, the former having the endophyte supposedly confined to the
epidermis of the host. In those mycorrhizae in which hyphae pene-
trate internally, Frank (1885) observed that they never go beyond
the innermost cortex of cupulifers that are invaded. In fruit-trees
the hyphae penetrate three-fourths of the distance through the cortex
(BouLET, 1910). In Olea the "prosporidi" are localized in an inter-
nal zone of large cells of cortical parenchyma (Petri, 1908). Mc-
DouGALL (1914), in studying forest trees of Illinois, found that the
central cylinder is never invaded ; while Taxus in France is said
(Prat, 1926) to keep the fungus out of stelar tissues by a layer of
"tannin" in the endodermis. In Eucalyptus the fungus is found in
epidermis and outer cortex but rarely deeper (Smith & Pope, 1934).
Of particular interest, remarked Noell (1910), are cases like Cun-
ninghamia in which hyphae penetrate only a few certain cell-layers
without any reason being apparent why they should not invade all
the cortical cells. Even the fossil tree, Ainyelon, shows the central
cylinder never penetrated (Halket, 1930).

Such phenomena were freely recorded by Janse (1897), whose
work is characterized by so much admirable detail : In Ophioderma,
sporangioles are found in third layer of cortex only while in Lecan-
orchis it is the second layer that is invaded, and in Dendrobium all
layers except the last are penetrated. In Burmannia the layer next
the endodermis is exempt while in Aronychia the hyphae never invade
the innermost cortical cells, which are filled with "tannin". In Elaeo-
carpus, invasion is to the mediocortex only while in Michelia invasion
is confined to certain points in the cortex, and resin canals are never
penetrated. So, too, in Dysoxylon the secretory canals are never
invaded.

Limitation in Orchids and other Herbs: — Besides the orchids
named by Janse, the following may be cited: In Ccntrosis it is the
mid-cortex to which the endophyte penetrates and the inner cortex
and the central cylinder are always free from infestation (Arcu-

Lecture XII — 175 ^ Mycotrophic Phagocytosis

LARius, 1928). In Pogonia the fungus seems never to penetrate
deeper than the inner cortical cells (Carlson, 1938). In Neottia the
3-4 outer layers of cortex are infested (Magnus, 1900) while the
fungus never penetrates the central cylinder which, said Magnus,
''seems a remarkable localization". Pittman (1929) found that the
fungus never penetrated Rliisantliclla tubers. Ames, who saw (1922)
that the vascular tract of Goodyera is never invaded, remarked (1921)
that certain areas of the orchid root {sic) seem able to repel advance
of the fungus; and "it is as if there were some fungicidal capacity
in the cells of the root structure that restricts the fungus to a limited
area."

Some other herbs may be mentioned : Thus, O'Brien & Naughton
(1928) found the fungus in localized patches of inner cortex in
Fragaria; and Treub (1885) said that in Sacchariim the central
cylinder is never invaded. For the ferns the same condition obtains :
In the sporophyte of Botrychium, at a definite distance from the
epidermis, the fungus branches copiously and forms sporangioles
while the vascular tissue is free. In the gametophyte the outer cells
are at first invaded but become fungus-free, which is the condition
of the apex and reproductive organs at all times. In Ophioglossum
prothalli the inner cells are fungus-infested while the outer are free
(Bruchmann, 1904; Lang, 1902). In Lycopodium the fungus is
present in epidermis of prothallus only (Hollo way, 1920), or at
most 1-2 outer cell layers (Goebel, 1887).

Supposed limitation of endophytic invasion by what are called
tannin deposits does not occur, for the endophyte can freely invade
such cells. Incidentally it may be remarked that Woodroof (1933)
found tannin formed in cold weather and present in both mycor-
rhizal and non-mycorrhizal roots.*

Limitation in Hepatics : — Every report on the hepatics indicates
a definite localization of the endophyte. Thus, in Conocephalus the
fungus is limited to a zone of central tissue (Beauverie, 1902), while
GoLENKiN (1902) reports that in a number of liverworts the hyphae
are confined to a compact ventral tissue. In the New Zealand liverwort
Monoclea the fungus is found in a sharply defined zone and does not
occur in the growing point (Cavers, 1903). In Marchantia the fungus
is limited to a zone beneath the air cavities (Chaudhuri, 1925), while
in Lumdaria the endophyte is present in a band of tissue (Emberger,
1924; Nicolas, 1924) along the midrib (Ridler, 1923). Ridler

*MacDougal & DuFRENOY State that decompensated respiration results in
polymerization of the quinoids into gummy masses, the presence of which forms
a barrier to tlie extension of hyphae (Plant Physiol. 21 :1-10, 1946).

Kelley — 176 — Mycotrophy

(1922) said that the fungus occurs in a definite zone along the ventral
midrib of Pellia and that the hepatic seems to exercise control over
the fungus. According to Magrou (1925) the fungus degenerates
about the archegonia and sporogonia, which seem to exert an inhibitory
influence upon growth. Auret (1930) found further that the
endophyte does not penetrate gemma-cups and archegonia of Liinu-
laria. Moreover, in Seivardiella (Chalaud, 1932) the fungus is
checked by active meristematic cells and the bulb is immune.

Limitation in Root Apices: — From all published accounts the
mycorrhizal apex is free from infection. To give some examples :
No hyphae were found in the vegetative point of Hippophae (Arcu-
LARius, 1928) ; the root tip of Vitis is never invaded (Petri, 1907),
nor the apical meristem of Taxus (Prat, 1934) ; while in pecan only
occasionally does the fungus enter cells of the growing tip (Wood-
ROOF, 1933), In Neottia the fungus is always found a short distance
back of the growing point (Drude, 1873), while in Philesia the
fungus penetrates tO' within 10-12 zone cells behind the apex (Mac-
farlane, 1897). Young roots of Paris are fungus-free 1.5 cm. from
the root-apex (Schlicht, 1889). In Monotropa the fungus
diminishes toward the apex (Kamienski, 1884) ; it does not enter
the meristematic zone of Dipodium (McLuckie, 1922) ; and the root-
tip of Thismia is fungus-free (Pfeiffer, 1914). The fungus is sel-
dom closer than 3-4 mm. of the root-tip of Corallorhiza (Thomas,
1893) ; in Angiopteris and other ferns the endophyte is absent from
the root-tip (West, 1917).

Limitation in Long Roots : — It is well-known that the long roots
of extension are fungus-free. Thus Gibelli (1883) said that in
Castanea the long, rapidly growing roots are free from infection, while
in Cacao the long roots are specifically stated by Pyke (1935) to be
fungus-free, and they are rarely infected in Taxus (Prat, 1926).

Limitation in Green Tissues: — Magrou (1925) noticed that
when hyphae invade cells of Pellia containing chlorophyll, the latter
is destroyed; and Ridler (1922) also observed that chloroplasts dis-
appear in Pellia on fungal invasion. Conversely, where chloroplasts
exist there are no fungi: Thus, Bolleter (1905) found that green
plants of Conocephalus showed no infection while neither starch nor
chlorophyll bodies occurred in infested cells. Again, in Lunidaria,
chlorophyll tissue is never invaded (Emberger, 1924) ; and indeed,
GoLENKiN (1902) had said that in liverworts infested cells never
contain starch or chlorophyll. Moreover, where Orchis incarnata roots

Lecture XII — 177 — Mycotrophic Phagocytosis

were exposed to light, chlorophyll developed on the upper (lighted)
portion and here there was no infection, but in the lower (shaded)
portion chlorophyll was absent and the endophyte was present
(BuRGEs, 1939). This observation had been anticipated by Janse
(1897), who noted that Lecanorchis cells were fungus-free when
they contained chlorophyll. Mollison (1943) suggested a loss of
fungal vitality after a length of time, to explain failure of fungus to
penetrate where chlorphyll is developed.

Summary of Limitation: — The sum of all these observations is
as follows : The invading endophyte is kept out of mycotrophic plant
tissues (-?) at a definite distance from stelar tissues of vascular plants ;
(2) from the growing apex of the root, never occurring in a meriste-
matic tissue; (i) from all chlorophyll tissues, which of course con-
tain active plastid bodies; and (4) from reproductive bodies such as
gemmae-cups, or archegonia of liverworts. Or to sum up these
categories into a single one, the endophyte is kept from all places
where active physico-chemical processes occur. They are kept out by
what has been aptly called a brutal phagocytosis.

The Starch Relation: — One more link in a chain of evidence
must be presented, namely, the fungus-starch relation. Briefly, fungus
and starch stand in inverse relationship, for where the fungus is
present no starch exists, for the fungus utilizes the starch as it pro-
gresses. Many examples may be cited: Boulet (1910) found that
starch disappears from fruit tree mycorrhizae when fungus is present ;
RuGGiERi (1937), that starch vanishes from sporangiole cells of
Amygdalus; Endrigkeit (1937) noted similar disappearance of
starch from Rhamnus; Figdor (1897), from Cotylanthera; Issat-
schenko (1913), from Tribulus; and Jennings (1898), from Coral-
lorhiza. Starch disappears from Dipodiiim mycorrhizae soon after
penetration of hyphae (McLuckie, 1922) ; the fungus uses starch
in Centrosis (Arcularius, 1928) ; penetration of hyphae in Orchis
is followed by dissolution of the starch (Fuchs, 1924) ; on the
entrance of the fungus into Pogonia the starch begins to disappear
(Carlson, 1938). Burgeff (1909) had said that the orchid fungi
dissolve out starch as they go, a statement anticipated by Schacht
in 1854.

On the other hand, Kusano (1911) stated that in Gastrodia starch
disappears from all mycorrhizal cells of the cortex but reappears in
the inner (third region) after cessation of metabolic activity. In the
innermost cells of Ophioglossurn prothallus the fungus is alasent and
cells are full of starch. In Botrychium the apex and reproductive

Kelley — 178 — Mycotrophy

organs (being fungus-free) are full of starch, which occurs nowhere
else. In Lycopodium, infected cells contain oil rather than starch
(Bruchmann, 1906), while in Pellia (Ridler, 1922) the fungus
uses starch "which is replaced by oil after entrance of fungus". Again,
in higher plants it is found that "Whereas in non-infested roots the
starch is deposited indiscriminately, in those colonized by mycorrhizae
it preponderates in the cells free from mycelium" (Endrigkeit,
1937). Added evidence that carbohydrates are used by the fungus
is provided by Bjorkman (1944) whose experiments show that
pine on being "strangled" by a wire placed 5 cm. above the ground
level formed almost no mycorrhizae while the amount of soluble car-
bohydrate in the roots dwindled. Bjorkmann believed that mycor-
rhizal form is largely conditioned by an excess of soluble carbohy-
drate in the roots. The fungus can use only a readily soluble carbo-
hydrate like glucose, as shown by Melin & Norrkrans (1942).
Magrou has found that formation of potato tubers is conditioned by
the osmotic pressure of sugar within the cell. In nature, it is the
mycorrhizal fungus which ordinarily changes starch of the plant cell
into sugar. "Ce processus de dislocation des parties colloidales du
protoplasme a ete designe par Errera sous le nom d'anatomose"
(Ann. d. Sci. nat. Bot., XI, 4:97-102, 1943).

Rexhausen (1920) has summed up the matter by saying that
the fungus takes carbohydrates from the plant in the form of sugar.
Thus sugar is obviously obtained by use of the plant's reserve starch.
Or, as MacDougal & Dufrenoy (1944) state : "Hydrolyzation prod-
ucts of polyuronides, of starch, and of other dififusible compounds
may be absorbed by the fungus."

At the same time Young's (1940) objection must be taken into
account : "The concept of the higher plant obtaining carbohydrate
from its fungus symbiont is in direct contradiction to the unsupported
but generally assumed theory that the mycorrhizal fungi obtain carbo-
hydrate from the tree roots as their share of the symbiotic relation-
ship. The hymenomycetous fungi which form the mycorrhizas are,
however, quite capable of obtaining their own carbohydrate supply
from the breakdown of organic matter. This is evidenced by their
vigorous growth on raw organic substrata and is supported by the ex-
perimentally proved fact that one of the major functions of the fungi
associated with orchid roots is to supply carbohydrate to the higher
plant." The solution of this problem would seem to lie in this, that the
fungus "dissolves" starch as it invades the tissues of the higher plant,
and releases later to the higher plant whatever it has brought from the
soil on phagocytosis. The action would seem to be in both cases
mechanical.

Lecture XII — ^179— Mycotrophic Phagocytosis

Conclusion: — The explanation of these phoenomena remains for
the future. Certainly there is an underlying cause. Meanwhile, certain
facts may be pointed out.

(1) It is already established that there is a difference between the
included content of stele and cortex. MacDougal & Dufrenoy
(1943) state: "The pericycle and endodermis layers in the root
mark the boundary between two contrasted t}'pes of tissues ; those
in the stele rich in phosphorus linkages which may be described
as energy-rich, and able to counterbalance the oxidase activity,
and those in the cortex, relatively poor in such linkages, and
rich in catechol and in catechol oxidase. Fungi . . . never transgress
beyond the endodermis into the stele." Further, these authors state :
"Oxidase activity seems to be higher in the cortex of the pine root
(whether previous to mycorrhizal infestation or after) than it is in
the stele. Such a difference should play a role in controlling selective
permeability : anions, with their negative charge, should be carried
from the site of higher activity, to that of the lower. The tissues of
the stele, from their meristematic stage, maintain a low oxidase level,
by retaining a high level of phosphoric complexes, acting as dehydro-
genases. This condition enables them to trap such anions as (H0PO4)
or (HPOJ".

(2) Another fact is, that the hypha which penetrates into the cor-
tex develops branches at a certain point to form an arbuscle. Such
proliferation is ordinarily the result of introducing an hypha into
an hypertonic solution of ions. We may note that Burges (1939)
had already noticed that during early stages of infection hyphae are
capable of further growth but that as histological changes become
apparent the fungus gradually loses its vitality. "The intracellular
arbuscles cannot be interpreted as assimilatory organs, since they
are digested as they form and show no indication of hyphal develop-
ment from their terminal branches, but rather as proliferations
induced by the growth-promoting stimuli of the cell-sap" (End-
RiGKEiT, 1937). VuiLLEMiN, in reviewing Galea ud's work, said
that arbuscles are less a characteristic production of the endophyte
and more a result of the reaction of the host-cells to invasion
by a foreign body. Magrou (1939) saw incipient arbuscle for-
mation with endophyte of Arum, on addition of aneurin. Demeter
(1923) had found with endophyte of Vinca that peculiar stuntings of
growth, called forth by different concentrations of sugar and special
sorts of sugar, recall arbuscle formation.

Kelley — 180 — Mycotrophy

(3) The hyphae break down. By dissociation of complexes in the
cell-sap, free H-ions are left in solution. These ions, acting on the
fungal arbuscle or the undifferentiated hypha, cause it to break down
and extrude its plasm into the host-cell. Demeter (1923) had shown
the breaking down of hyphae in vitro at an optimum acidity of
0.025N HCl. This concept is in agreement with Magrou (1921),
who said that the fungus is limited to certain parts of the plant
through toxic constituents of the cell-sap. It is also indicated by
Hatch's (1937) statement that susceptibility to infection by mycor-
rhizal fungi is apparently controlled indirectly by the internal concen-
tration of nutrient elements in short-roots. These "toxic substances"
are apparently ions normally present and not special humoral bodies.
RouTiEN & Dawson (1944) suggest an increased H-ion output in the
mycorrhiza, arising from carbonic acid, but leave unsettled the ques-
tion of its origin.

(4) The fungal material is digested. The presence of proteolytic
enzymes enables the host's digestion-cell to utilize the extrav-
asated plasm of the fungus. Hitherto the hypha was utilizing the
host's substance; but now the host gets back not only what it had
previously lost but all that the fungus brought in from the soil. In this
sense there is a total intake of mineral salts, organic substances and
water by the mycorrhiza, but all combined as protoplasm of the fungus.

(5) The digestion-area is strictly localized. Since the ionizable sub-
stances which pass from the stele to the cortex are subject to definite
physical laws, the rate of diffusion is specific for a given sort of plant,
being conditioned by the nature of the substances through which
diffusion must take place. For this reason, phagocytosis must neces-
sarily be limited to a certain region of the cortex. "I think," said
Emberger (1924), "that localization of infection is conditioned by
differences of osmotic pressure."

The apical meristem and other growing points are richly supplied
with ionizable substance by the flow of liquid materials into such
regions. Through these rich supplies, actively growing tissues can
readily repel the endophyte by breaking it down at a distance from
the meristem to which the ionizable substances extend. Chloroplasts
in green tissues and perhaps leucoplasts in tubers probably exert
a similar influence.

(6) The mechanism of phogocytosis is apparently ionic. A plant
is not static, and the more active its growth the more ionizable material
it will have at its disposal, and the more certainly will the fungus
be destroyed in its tissues. Hence it may be understood what Reed

Lecture XII

181 —

Mycotrophic Phagocytosis

& Fremont (1935) discovered when they applied stable manure or
cover crops to plots of Citrus and found that the trees seemed to
develop resistance to the fungus, a resistance which untreated trees
seemed to lack. In the treated trees they found that the cytoplasm of
the host-cell enveloped arbuscles of the fungus with apparent active
proteolysis. It is evident that with better conditions of growth the
Citrus trees had more H-ion at their disposal for breaking down of
the fungus.

In this connection likewise may be cited the writer's (1944)
studies of chestnut, sprouts of American chestnut having little
resistance to blight whereas seedlings have decided resistance. More-
over, seedlings under better conditions of growth in a natural wood-
land are more resistant than seedlings in the open. Resistance to
the fungus is once again, in these observed cases, correlated with
vigorous growth.

"One seems justified in concluding that the mycorrhizal fungi, both
ectophytic and endophytic, are potential parasites controlled by
reactions of the host-cells" (Burges, 1936). Lacking sufficient ioniz-
able substance, the tissue is parasitized and progressively destroyed.
Possessing requisite ions the tissue breaks down the fungus.

4

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PLATES

Plate 1. — This plate, prepared by Edouard Prillieux and
published in 1856, is one of the earliest illustrations of mycotro-
phic infection. It shows the habit of the orchid, Neottia, and
its fungal infection in figures 7-10. In fig. 7, the outer cortical
cells are shown filled with a mycotrophic content ; in fig. 8, a
septate mycelium ; in fig. 9, a mass of yellowish matter sur-
rounded by hyphae; in fig. 10, young cellular tissue containing
pelote and intact nucleus. Further stages, and development of
coralloid mycorrhizome, were illustrated in Plate 18. {Repro-
duced from Ann. d. sci. nat. : Bot., 4 me. ser. vol. 5, pi. 17, 1856)

;: (/:■.'• i-i-:/i'\ /if/,' . ^'JcJU'

/liK' ■ J',i,;i . ,'> / / il

\j

b^'i

J , i-

v\ \ 1.

J

///;

-i^;

20

V-

i^

.•"•V/. /'ri'^^er/.T Je/ .

^-?r-' /■,■./'.../« j-<-

Jjei'e/tYY>c//<i-/i/' (/rs y7,7,-//if.y ,///' Xeottia niihis-avjs.

i

Plate 2. — Mycorrhizae in Scot's Pine, Pinus syl-
■ucstris. — Dr. Frank started modern mycorrhizal study
with a description of sucIt structures in pine and
beech. This illustration shows both coralloid and
nodulous forms of mycorrhizae. and was originally
published by Meux. {Reproduced from Rayner's
Mycorrhiza, Phitc VI).

■<' M\

/■},y /

.'• , "I,

_..x

7;//:/'

/I

/-/^ '/

/''f>T O-

/'''!/ J

/'}J-'r<'.'i/< :',.'

Fnj.h'

Plate 3. — The plate which accompanied Frank's epochal paper. All of the figures
except 4 and 7 are to illustrate mycorrhizae of Carpinits Befitlus, the other two being
of Fagns sylvatica. The mycotrophic enlargement of the rootlets, and mantle of
hyphae are well shown; and the sections neatly indicate ectotrophic structure. (Ber.
deut. hot. Gesell., vol. 3, pi. 10).

Plate 4. — Effect of mycorrhizae on plant growth. "Two beds
of seedlings of a Himalayan species of pine, Pinus longifolia,
from the same sowing in northern Rhodesia. Further bed inocu-
lated with soil containing a mycorrhiza-forming fungus for the
species from a vigorous plantation of this pine at a station in
Southern Rhodesia 1000 miles distant; nearer bed untreated."
(From a photograph kindly loaned by Dr. R.wxf.r).

Plate 5.— Photomicrograph of a portion of a fossil mycorrhizome of
Sclera ptcris UUnnicnsis. In the large cortical cell are hyphae and bodies that
arc considered as vesicles. {After Andrews and Lexz in Torr. Bull. 70:122,
1943).

INDICES

Subject Index

Acid soil 5,62,63,69,78,83,85

Actinomycosis 113

Adventive roots 57,108,117

Aeration 104

Aerial organs 68,101,102,117.130

Africa 59

Alder 31,54,56,72

Algae 17,24

Alkaline soil 5,69,73,78,83.85

Alpine 57,93

Altitude 80,86,112

.\luminium plants 69.70

Ammonia content of soil 70

Anatomy 84

.\neurin 10,160,180

Annuals 80,82,117,137

Antheridia 23

Apetalae 30

Apex 22,82,92,99,100.110.119.120.170.170

Arbuscle 3,9,85,93,109,125,129,107,179

Archegonia 23,92,93,170

Arctic 57

Aridity 74,88

Arthrophytes 14,21

Ascomycete 40,97

Asia 60

Asparagin 76

Aspen 74

Asymbiosis 104.10S.110

Asymbiotic germination 7,142.143

Australia 63

Austria 58

Autotrophic 39,153

.Vuxonc 10

Bactf,ri.\ 23,24,25.30.70.103.104,105,106,107,

108,109
Baltic States 56
Basidiomycete 38,116,142

Beech 31,40,55.56.57.69.71.73.70.81.83,121,148
Bjciikmann's hypothesis 157
Bohemia 58
Brazil 64
British Isles 55
Bryophytes 18
Bulb 104

Cacti 74

Calcareous soil 69

Calcium salts 154

Canada 65

Carbohydrate 144,152,158,178

Carbon dioxide 79,155,158

Carbon hypothesis 10,146,158,1.^9.178

Carbon supply to embryo 145

Carbonic acid 10

Carvopsis 8,63,101

Central U.S.A. 66

Ceylon 60

Chaparral 59,67

Chile 64

Chlorophyll tissue 92,93,176

Chlorosis 28

Clay 67,69,74,93

Climatic influence 7

Clof 3,18,93,98,100,127,105

Coal-ball 47

Cold, effect of 82

Colours of mycorrhizae 118

Communication hyphae 119,150,156,163

Composts 89

Conifer 23,25,56,62,63,76,81,88,149,171.177

Consortium 23,30

Coralloid 24,41,09,72,99,108,109.110,117,127,

131
Corm 104
Cortex 20,23,29,49,50,51,75,82,83,95,99,101,

104,105,106,108,109,110,123,130,132,157,

167,174
Croatia 58
Cryptogams 40,61
Culture experiment 139,149,160

Degeneration 7,142

Denmark 55

Diarch bundle 51

Dichotomy 6

Dicotyledons 33,61,125

Digestion : See Phagocytosis

Digestion cells 98,99,115,151,157,170,172

Digestion stages 8,93,95,97

Disease 150,152,164

Distribution 53,75

Ecology 5,12,24,29,137
Ectendofrophic 123,172
Ectotrophic 29,30,59,61,02,64,67,70.71.75,78,

80,84,108,122,123,155,171
Edaphic 70

Eiweisshvphen 98,126,157
Endophyte 4,7,12,18,23,29,37,39,41 .44,54,60,

60,80,91,93,97,101,102,103,111.112,115,

116,123,140,145.149,157
Endophytism 46

Mycotrophy

!17

Subject Index

Endotrophic 21,23,25-28,32,34,40,5(5,58,59,61-
62,70,72,75,80,84,85,87,97,103,105,108,
114,116,122,125,149,157,162
Enzyme 25,105,108,109,143,166
Epidermis 50,94,97,109,114,119.123,130,168,

169,170
Epiphytic 63,68
Ericads 56,61,62,65,66,72,78,110,116,125,1.30,

138,139,148,149,169
Exkretlcorper 109

Facultative mycotrophy 114,115.138,149

Fairy rings 76

Fern prothallia 40,66,146,177

Ferns 4,20,49,52,94,97

Finland 56

Forest fungi 63,136

Forest tree mycorrhizae 7,11,38,55,61,73,87,

136,153
Form genera 43,45
Fossil mycorrhizae 47
France 54
Freezing 79
Fungal identity 8,41
Fungistatic substances 90
Fungus 23,30,48,68,77,99,104,107,109,110

Gametophyte 14,19,22,23,94,175

Gasteromycete 43

Germany 53

Germination 44,56,77,139,141,149

Gliotoxin 90

Glomerule 168

Glycogen 100

Grasses 64

Growth-promoting substances SO.lOO.UiO

Guatemala 165

Gymnosperm 14,23,60,61

Habitat 39,54,84,132

Halophyte 58

Hartig net 5,26,27,49,119,123,134,133.172

Hatchian hypothesis 9,155

Heather 35

Helotism 152

Heminasidionivcete 41

Hepatic 14,17,22,48,56,58,60,61,157,158,175

Herb 32,38,53,54,57,60,66,73,83,125,137,175

Heteroauxin 10

Historical geology 47

Host-cell 50,98,105,115,152

Humus 2,3,5,12,42,54,67,68,70,71,73,89,92,104,

113,115,119,136,144,153
Hungary 58

Hydrocarbon hypothesis 159
Hydrogen ion 3,9,99,152,169,180,182
Hygromorpliic 61,69
Hygrophyll 35
Hypertrophy 85,92,99,103,105,109,111,121,124,

168
Hyphae 17,20,22,25,49,51,91,94,105,119,123,

128

Identity mycorrhizal fungi 8,38,67

Immunity 182

India 60

Inoculation of soil 12,88,89

Insectivorous plants 33

Intaking mechanism 161

Intercalary vesicles 51

Intercellular 22,111,125,128,129,134,167

Intracellular 20,22,50,95,101,106,115,118,123,

127,168,170,173
Ions, balancing of 11,181
Isolation 8
Istria 59
Italy 59

Japan 43,62
Java 60

Law optimum altitude 80

Legumes 39,57,60,62,64,103

Lichen symbiosis 17

Light, effect of 80

Limitation endophyt.e 91,92,94,174,177

Liverwort 3,4,14,17,40,62,91,146,157,158,175

Loam 73,114

Long-root 82,116,176

Lycopod 14,94.95,146

Lycopsida 21,39,50,52,61,177

Madagascar 60

Mamelon 25,26

Mamillae 130

Mantle 29,111,119,121,123,130,169,172

Manure 71,80,90

Maqui 59

Marsh plants 74

Meristem 22,120,121,170,170

Mesic species 175

Microflora 79

Microhabitat 12.84

Mineral intake 9,154,155

Mineral nutrients 9,11,70,77,153,155,100

Monarch bundle 135

Monocotyledons .35,61,125,127

Mother-root 104,116

Mull 70,72

Multiple mycorrhiza 37

Muskeg 48

Mutualism 17,26,51,95,103,148,149.104

Mycocaryopses 8,101

Mycocecidium 103,142

Mycoclena 119,124,125

Mycoderm 119

Mycodomatia 30,103,142

Mycophagy 151

Mycorrhiza, definition 6

Mj'corrhizae, early observers 5

Mycorrhizal form 39,71,74,110,117

Mycorrhizome 4,23,49,50,59,64,91,97,99.100

Mycothalli 4.17,21,22.58,91,132

Mykokrinie 70

Netherlands 55

New Zealand 42,62

Nicotinic acid 160

N assimilation 24,61,87,104

N atmospheric 78

N fixation 25,69,78,104,105,106,108,109,110,

132
N nutrition 7,9,61,153
N salts 7,9,45,71,76,77,86,153

Kelley

218-

Mycotrophy

N theory 9,153

Nodules 2,24,25,26,31,41,62,103-105,108-111,

114,132
Northeastern U.S.A. 65
Nonvaj' 56
Nucleic acid 76
Nucleus 22
Nursery 86 -SS
Nutrient conveyors 163
Nutrient salts 15,68,154
Nutrition, mycorrhizal 98.137,151

Oblig.vte mycotrophy 8,91,110.136.138.140
Occurrence 14,16

Orchid 3,7,36.39,58,60,62,64-66,77,79,83,97,
104.115,125,140,149,157,158,160,164,174
Orchid fungi 7,141,142
Orchid seed germination 44,141
Oxidase level 180

Pacific co.\st 67

Palaeobotany 47

Palmette 124,134

Parasitism 2,5,6,9,21,23,38,50,51,71,74,84,105,

107,112,118,149,156
Passage cell 132,169
Pathogen 149

Pearl-necklace mycorrhiza 25-27,68,09,70,117
Peat 67,90,95

Pecan mycorrhizae 66,73,74,83,119
Peloton 20,39,92,96,114,115,120,165
Peptone 76
Perennism 7,8,80
Peritrophic 84,85
Phagocytosis 3,8,11,26,50,51,93,99.100,105.107,

110,111,127,129,143,164,170.177,180
Phallomycete 43
Phenology 81-83,114,122,171
pH reactions 77,85.143,166
Phycomycete 11,33-35,38,39,47,54,57,116,119
Phytophagy 7,170
Pilzwirthzellen 50,98,126
Pinaceae 26

Pine 23,26,29,38,42,45,54,55,57,63-65,67,69,71,
74,77,82,83,86,89,121,131.154,174

Pioneer-root 116

Plasmoptyse 130,169

Phosphoric complexes 179

Podsol 73

Portugal 55

Potassium salts 45,77,87,154.158

Potato 37,44,59,80,104,111,1.50

Prairie 12,86-88

Prosporidi 25,148,174

Protein liyphae 98,100,167,172

Proteolysis 45,164,182

Prothallia 20,21,40,48, 63,91, 146, 17.-.

Pteridophyte 14,19

Pteridosperm 50

Pseudomycorrhizae 66,69,74,1 IS. 104

Pure culture 28

Racemose 24,69,117

Rain 75,76,83,121

Raw humus 70,72

Renewed growth 69,74,76,82,83,110.121

Rhizoids 22,91,92,95,97

Rhizomorph 71,119

Rhizosphere 12,84,85

Rhizothamnion 72,117,132

Rocky Mountains 67

Romellian hypothesis 152

Root-cap 20,120,121

Root competition 70

Root -hair 1,5,12,15.20,24,25,27,28,50,53.98,

109,119,150,156,162
Rosette formation 107

Salt absorptiox 155

Salt marsh 51,58,85

San Domingo 65

Sand 12,56,67,69,71-75.93,105.108.114

Sap concentration 112.105

Saprophyte 5,38,50,52,74,84,101,104,140

Seed coat 101

Seed infection 8,139

Seed sterilisation 139,143

Seedlings 76,79,80,85,86.88-90.107.139,142.154,

155
Seeds, impotence 145
Sekretkorper 107
Setose hyphae 117,119.121,102
Short-root 116,118
Sicily 59

Simple mycorrhiza 117
Slope exposure 75
Soil 12,68,69,72,73,74,76-78
Soil inoculation 28,60,64,88,89
Solfatara 62,69,75
South America 64
Southern U.S.A. 66

Sporangiole 20,91,93,105.125,127.129,107
Spore colour 43
Sporophore 4,8,38,67,70
Sporophyte 19,22
Spruce 23,29,56
Stahlian hypothesis 19,155
Staining 62
Starch 3,17,91.95,99,100,108.110.114,130.151,

156,167,177
Starch, hydrolvsis 143.178
Stele 48,49,51,100,109,111,114,120,134,174,177
Sugar 7,71,78,108,110,112,113,139,143,144,150,

169,178
Sugar-cane 65
Symbiophile 38
Symbiosis 109,136,164

Synthesis 9,29,31.38,40-43,107,131,130,142,149
Synthetic mycorrhizae 128
Systemic infection 44,140

Tannin 108,111,172,174,175
Taxonomic identity 21
Thamniscophagous 85
Theories of mycotrophy 9,148
Thiamin 160,166
Toxic residues 72,90
Toxin 115,139,149,164,180
Transpiration stream 9,70,154,156
Tree mycorrhizae 77,83,90,136
Trenching experiments 29,90
Trinidad 65
Tropical 60,65
Truffles 40

Mycotrophy

•219

Index of Plant Names

Tuber 93,103,104,108,111,113,115
Tuber formation 7,79,80,115.14(3
Tuber, microscopic 113
Tubercle 23,24,51,104,106,107,111
Tuberous mycorrhizae 93,109,113.117,131

Vascul/Vr structure 51
Venezuela 65

Verdauungszellen 50.99.12G
Vesicle 39,48-51,85.92-94,97.99.109.123,128,
130,169

N'esitular-arbuscular mycorrhiza 11,47,48,103,

126,127,167
\'itaniin 10

Wall tubule 114,132,165
Water intake 70
West Indies 65

Xeric species 75
Xeromorphy 61,69

Index of Plant Names

Abies 28,121,123,131,133

Abietineae 125

Abelicea 31

Acer 2.5,34,60,117,150

Acorus 35

Actinoiiijjces 106,107

Adenostoma 67

Adenostylcs 54,113,134

Adiantum 21,66,97

Aescubi:i 34,120

Agariciircue 41,137

Agaricus 41

Agave 35

Atlantlnis 33,34.109

Albizzia 61

Aletris 36,62

Alisma 35

Allium 35,56,115.104,170

Almis 31,.56, 62,72. 105, 107, 108, 109. 118, 121, 123,

130
Aloe 35
Alsophila 20
Alternaria 138
Altingia 34
A7nanita 37,41,42,56,78
Arnanitopsis 42
Arnaranthus 53
Amaryllidacrae 36
AmeiUaceae 137
Amyelon 51
Aviygdalus 11,34.69,177
Anacardiaceae 34
Ananas 36
Andreales 19
Andromeda 111,130
Aneiira 4,94
Atigiopteris 19,125,136
A7ionaceae 34
Anthoceros 18
A-plectrum 67,104

Apocynaceae 169
Apteria 65

Araclinites 57
Araliaceae 34
Araucaria 28,89,125
Arbutus 59,111,130,134
Archangiopteris 20
Arctostaphylos 67,125
Ardisia 34
Arenaria 32
Arisae77ia 35
Armeria 85
Armitlaria 42,100
Aronychia 174
vlnmt 10,35,40,125,160
.4sar!^ni 32
Asclepiadaceae 58,169
Asclepias 34,58,132
Asimina 34
Asparagus 35,114
Aspergilhis 40,78
Aspidium 21
Asplenium 21
.4s<er 68
Asterocystis 40
Asteroxylon 48
.l/ri'p/ex 32
.4ra/ea 139

Bacillus radicicola 24,104,109
Begonia 168
Benzoin 33,34
J5e<a vidgaris 32
fie<!i/a 56,62,73,134
.8. aZ6a 55,63,136,172
Bignonaceae 33
Blechnum 21
Boehmeria 32

Boletus 28,37,38,41,56,57,58,60,71,78,87,89,136.
137

Kelley

220

Mycotrophy

Borraginaceae 33

Cyathea 20

Botrychium 19,39,66,94,95,97,129,132,175,177

Cycadaceae 24

Bromelidaceae 36

Cj/cas 104

Burmannia 5,36,61,65

Cydonia 34

Buxus 34

Cymbidium 143,145
Cyperaceae 35

Cacao 35,65,128,176

Cyperus 35,114

Cactaceae 34

Caesalpinaceae 34

Da/i/io 113

Calla 35

Danaca 20

Calluna 8,35,44,55,69,73,78,100,102,111,130,

Daphne 34,58

134,138,139,149,153,170

Daucus 110

Calycanthaceae 33,34

Dendrobium 174

Calypogeia 92,93

Dianthus 32

Calypso 19,99

Dlapensia 33,34

Calvatia 43

Dicotyledons 61

Campanula 33

Dioscorea 35

Canthardlus 42

Diospyros 34

Caprifoliaceae 34

Dipodium 63,132,176,177

Carex 35

Drjscm 33,62

Caragana 89

Drj/as 57,80,124,137

Carpinus 31,55

Dysoxylon 34,167,174

Caryophyllaceae 32

CosfOHea 54,55,123,136,176

Eichlwnna 120

Castanopsis 31

Elaeagnaceae 34

Casuarina 60,61,63,66,89,105,109

Elaeagnus 109

Cattleya 44,142,143

Elacocarpus 174

Ceanotlms 66,109

Elaphomyces 40,41,54,75,136

Cedrws 26,58,59

Empetrum 78

Celasfrus 34,67

Encephalartos 24

Centrospermae 30

Endogyne 39,40

Ce/iis 25,31,49

Entorrhiza 41,114

Centrosis 59,174,177

Ephedra 30

Cephalanthera 58,100

Epigaca 35,65,102

Ceplialofaxus 26

Epilobium 33

Cerastium 32

Epipactis 58,143

Ceratopteris 19

Epipogon 127

Cercocarpiis 67

Epirrhizanthes 61

Chatnaecyparis 27

Bnca 130,149

Cheiropleuria 21,62,95,146

Ericaceae 34,35,44,57,72

Chenopodium 32

EriophoTum 35,114

Cinchona 61

Eriostemon 63

Cistaceae 33

Erythronium 35

Citrus 34,40,60,67,83

Eucalyptus 34,43,60,63,64,110.173,174

Clathrus 43

Euphorbia 34,49

Clethra 34

Clitocybe 42

Fagaceae 31

Cochlearia 127

J^o.vMS 55,69,71,73,118

Coiiipositae 33

FegateUa 18,92

Comptonia 30

F?f«s 32

Conorephalus 18,93,128,158,17.5.176

Firmiana 34

ConvaUaria 35,83

Forsythxa 59

Corallorhiza 55,58,67,99,100,127,162,176,177

Fossombronia 93,94,146

Cordaites 50

Fragaria 44,54,175

Coriaria 62,110

Frankia 107,109

Cornaceac 34

Fraxinus 35,84,85,117,173

Corticium 44

Fritillaria 35

Cortinarhis 41,42,43

Fusarium 43,44,45,111,115

CoryZtw 31,42

Cotylanthera 61.177

Galanthus 35

Cruciferae 33
Cryptogams (il
Cryptoincria 6S
Cunninghamia 28
Cupania 34
Cupressus 27
Cuscuta 145

Ga/fo/a 62,79

Gardenia 34

Garryaceae 30

Gastrodia 62,63,100,137,140,141,153,158,177

Geaster 43

Gentiana 34,35,80,132

Gw/cffo 24,26

Mycotrophy

221 —

Index of Plant Names

Glaux So

Gleicheniaceae 20,95
Glyceria 85,161
Oonyanthes 174
Goodyera 144,145,175
Gossypium 35,128
Gramineae 35,39
Gyrnnosperms 61
Gypsophila 32
Gyrodon 107

Habenaria 141
Hamamelis 34
Hoplomitrium 56
Hebeloma 3,56
//ehVio 168
Helleborine 141
Helminthostachys 19,95
Hemerocallis 35
Hemibasidiomycetes 41
Hevea 34
Flicoria 30,66,73,83
Hippophae 56,84,110,176
i/oio 132
Humaria 40
Hydnurn 42
Hygrophorus 42,137
Hymenopliyllaceae 19
Hypericum 33
Hypholoma 29
Hypomyces 44

7Zex 34
Iridaceae 36

Juglans 30,49,117
Juncaceae 145
ywici/s 35,41,85,113,114,135
Jungermanniaceae 17,18,93,157
Juniperh 26,56

Kaulfussia 20

Labiatae 33

Lactarius 38,41,42,78,136,137

Laelia 143

/^oni 29,102,123,134,136,172

Lafhraea 107

Lauraceae 34

Lecanorchis 174,177

Ledum 130

Leguminosae 34,160

Leiophyllum, 66

Leitneria 30

Lepidodendron 48

Lepiota 42

Leucobryum 40

Leucojum 35

Librocedrus 28

Liliales 35,39

Limodorum 141

Linum 33

Liquidambar

Liriodendron

Litchi 60

Lit/iDcarpus

Livistona 35

Lobelia 64,152

25,34
34,117

31

Lo^mm 8,10,35,55,63,64,67,101,102,128

Lotus 54

Lumdaria 18,45,60,78,92,93,146,175,176

Lycoperdon 43

Lycopodium 22,61,65,95,96,97,175,176

Macroglossum 95

Macrozainia 24,63

Magnolia 34

Maianthemum 35,83

Malaceae 34

Mangiletia 34

Marasmius 42

Marattiaceae 4,19,20,95

Marchantia 18,92,93,146

Marchantiaceae 17,18,93,157,175

Marsilea 19

Matoniaceae 19

Melampyrum 113

Melanthaceae 35

MeZza 34

Menispermaceae 133

Merati 34

Michelia 34,174

Mimosaceae 34,108

Molinia 40,56,114

Mollisia 40

Monoclea 62,92,129,175

Monocotyledons 61

Monotropa 2,3,6,9,34,78,98,121,130,138.149,

151,169,170,176
Mortierella 78
Morua 31
Mwso 35,36
Muscari 35

Mycelium radiris 7,45,141
M.r. abietis 45
M.r. atrovirens 45,160
M.r. J-affi .4 78
Af.r. nigrostrigosum 45,118
Af.r. sylvestris 45
Myosurus 54
Mi/ncffl 30,61,62,65,106,107
Myrsinaceae 33

Narcissus 35

Nectria 44

Negundo 34,126

Neottia 2,3,6,9,34,98,99,126,141,150,151,165,

175,176
Nephrolepis 21
Neomammillaria 34,123
Nerium 34
Nothofagus 42,63
A^j/sm 33,34

Obolaria 34,75

Ocotea 34

Odontoglossum 44,143,144

OZea 174

Oleaceae 34

Omphalia 42

Onoclea 21

Gospora 19,44

Ophioderma 174

Ophioglossaceae 4,19,56

Ophioglossum 19,44,61,95,125,129,177

Kelley

99?

Mycotrophy

Ophyrs 143

Orchcomyces 7,45,142

Orchidaceae 56

Orchis 44,75,84,100,141,145,1(15,177

Ornithogalum 35

Orobanche 113,145

Orobus 109,127

Oryza 74

Osmundaceae 20,95

Osrnundites 49

Ostrya 31

Oxalis 33

Oxycoccus 35,78

Palaeomyces 48

Pandanus 35

Papilionaceae 33

Pam 125,176

Parkerinceae 19

Paxillus 78

PeHi'a 17,18,45,55,78,91,93,146,175,176,177

PeniciUium 40,78

Peron isporites 48,51

Pctallophyllum 94

PImlenopsis 143,145

Phascum 19

Phcrosphaera 26,63

Philesia 64,176

Phoenix 35

P/iomo 7,44,45,78,140

Pliyllophoma 44,138

PliyUostachys 43

Phytolacca 32

Pfem 29,43,65.78,102

Picrasma 34

Pilularia 19

Pinaceae 125

P/7n(S 42,43,57,62,64.66,07

132,136,154,174
/\ dcnsiflora 42.73,121
P. Merkusii 60,89,136
P. montana 29,78,117,131,
P. pinaster 29,64,136
P. rarfw^a 63,64,77,83,89
P. S^robus 29,43.77,82,86,135
P. sylvestris 29,55,73,78,131,132,134,138,144,

172
P. Taeda 63,75.1.38
P. virginiana 27,68,75,122
Pithecolobium. 108
Pittosporum 33,34
Plantaqo 58,85
Platanus 34
Platycarpa 30
Phunhaginaceuc 33
Podocarpineae 134
Podocarpus 24,25,28.62,63,73,105.125
Podophyllum 33
Pogoniu 175,177
Polygala 33,137
Pnlyg maceae 62
Potygonatum 35

Polygonum viviparimi 32,57.108,125
Polypodiaceae 20,21
Polyporus 42
Polysaccum 43
Populus 30,41,57,136,137,172

,131,134,137,172

,77.82,89,90.120,123,

,132

Portulaca 32
Potentilla 35
Preissia 17
Pr;>?i i/?a 33
Proteaccae 168
Protomycitis 48
Prwnis 34
Pseudolarix 29
Pseudomonas 109
Pseudotsuga 29,81
Psilotaceae 22
Psilotum 23,104,125
Pteridium 21,62,97,171

Pterocarya
Pterospora
Pyrolaceae
Pytliium

30

98,121
56,111
21,39,60,61,95

Quercus 31,55,66,67,86,118,123,124,136

Ranunculaceae 33,39

Ranunculus 53

Raphanus 108

Rhamnaceae 34,177

Rhizanthella 64.100,175

Rhizobiuvi 104

Rhizoctonia 7.34,44,45.111.115.137.140.141,

143,164
Rhizonium 50
Rhizophagus 1 1 .40,45,48,63
Rhizopogon 27,78,136
Rhi>dode7tdron 35,66,139,149
Riccia 18
Robinia 43
Rosaceae 34
Rubus 65

Saccharum 175
Sagina 32
Sallcornia 32.58,85
Soiw 30,57,62,124,136,150
Sn/«o/a 32
Salviniaceae 19
Sapindaceae 34
Sarforfes 67,98,121
Sarcosiphon 63
Sarraceniaceae 33
Sassafras 34,66
Scliinizia 107,114
Schizaeaceae 19
Sciadopitys 28
Sciaphila 64
Sc27?o 35
Scitarninaceae 36
Schoenus 114
Scleranthus 32
Scleroderma, 29,43
Scrophulnriaceae 33
Selaginella 22
Seinpcrviviim 121
Sequoia 28,125
Sewardiella 92,93,145,176
Shepherdia 109
Silene 32
Smilacina 35
Synilax 35
So/aHM?« 35,64,112

Mycotrophy

99

Z^O —

Index of Plant Names

S. tuberosum 80,111,112.113
Sphagnuvi 19
Spiranthes 83,99,143,145
Stapelia 128
Stapliyleaccae 34
Stellaria 32
Sterculiaceae 33,34
Stigmaria 48
Streptothrix 19
Strobilomijces 42
Styrax 34
Suaeda 58,85
Symploros 34

Talauma 34

Tamaricaccae 33,34

J'a??i!;s 104,125

Tax odium 28

rax7« 26,55,69,82,122,152,174,176

Terfezia 40

T/ieti 34,35

Thismja 66,87,98,100,176

r/«i;n 28,58

Thuopsis 28

T/fe 34,82,123

Tilletiaceae 41,114

TipuJaria 66,104,132

Tmesipteris 4,23,39,97,134

Torfea 20

Torreya 24,26
Trametes 54
Tribulus 56,108,151.177
Tricholoma 37,41,42
Trigolochin 85
Trillium 35
Tjiber 40,41,147
Tw/ipa 35
T/zp/ia 35

[/?)m(S 31,32
LViira 32
Uvularia 35

FofT;;(/(/»( 34,35,110.130,13!)

Vaitda 160

Vanilla 68

FerofruHi 35

Fmca 128,132,152,168,176

Fio^a 33,39,56

Fiiaceae 34

Fiffs 83,84,128,168,176

Yucca 35

Zelkova 31

Ze.uxine 66,137

Zingiber 36

Zoopsis 92

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Verdoorn et al.: Plant Science and
Plant Scientists during the Second
World War (in press) ca. 3.75

Verdoorn (ed.): 21st World List of
Plant Science Institutions and
Societies (in press) ca. 4.00

Wyman: Arboretums of North Amer-
ica 1.50

* * *

Imports which tve handle

for the .Americas only: —

Lindquist: Genetics in Swedish For-
estry Practice ,3. .50

Meyer- Abich (ed.): Biologic der

(ioethe-Zeit 5.50

Nelson: Introductory Botany 3.75

\ erdoorn et al.: Manual of Brjology '♦..^O
\ erdoorn (ed.) : Manual of Pti-ridology 1 1 .00
\N eevers: Fifty Years of Plant Physi-
ology 5-00

Catalogue and Book Department List on Request

llw ("Jtranica lintanira Cn.. M altliarti. Massachusetts, V.S..4.

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