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‘BULLETIN No. 1038
Contribution from the Bureau of Plaft Industry
~WM. A. TAYLOR, Chief
| Washington, DC. | ¥ March 20, 1922
PECAN ROSETTE.
ITS HISTOLOGY, CYTOLOGY, AND RELATION
TO OTHER CHLOROTIC DISEASES
. By
4 _ FREDERICK V. RAND
Pathologist, Laboratory of Plant Pathology
CONTENTS
‘ Page .
Types of Chlorotic Plant Diseases .... 1 Studies of Pecan Rosette—Continued.
Chloroses Due to Soil or Atmospheric Histological and Cytological Studies . 19
riniGONE oot nstawel ra cae els) 2 Subsidiary Experiments ....... 30
t hi Infectious Chloroses ..........-. 6 Probable Nature of Pecan Rosette .... 31
gis Studies of Pecan Rosette. ........ Rai ceremmimariens eu ki co Pa aw ere Sra). ay 36
Results of Previous Work ...... 13, Pp interatore Cited) o6 2 2)5) oo she, ele’ a: eh aie 37
External Signs of Rosette ......
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meGAN ROSETTE
ITS HISTOLOGY, CYTOLOGY, AND RELATION TO
OTHER CHLOROTIG DISEASES
BY
Frederick V. Rand
Pathologist, Laboratory of Plant Pathology
United States Department of Agriculture
Submitted in Partial Fulfilment of the Requirements for the Degree of
Doctor of Philosophy, in the Faculty of Pure Science
Columbia University, New York City
UNITED STATES DEPARTMENT OF AGRICULTURE
, BULLETIN No. 1038 4%
Contribution from the Bureau of Plant Industry y a |
WM. A. TAYLOR, Chief
Washington, D. C. Vv March 20, 1922
PECAN ROSETTE: ITS HISTOLOGY, CYTOLOGY,
AND RELATION TO OTHER CHLOROTIC DIS-
EASES.*
By FREDERICK V, RAND, Pathologist, Laboratory of Plant Pathology.
CONTENTS.
: Page. Page.
Types of chlorotic plant diseases____ 1 | Studies of pecan rosette—Continued.
Chloroses due to soil or atmospheric Histological and cytological
CORGIEIONS Seed she. rel ee pe 2 Rbudieg=4) A264 St Bee th 19
infectious, chloroses!= o-22—=_ = =.= 6 Subsidiary experiments _______ 30
Studies of pecan rosette___________ 13 | Probable nature of pecan rosette____ 31
Results of previous work_____~_ LS>|' SUMMA rye = ee 36
External signs of rosette______ iM BLCGEaALITe *CltCG i= = eae a ee 37
TYPES OF CHLOROTIC PLANT DISEASES.
The chlorotic group of plant diseases to which pecan rosette be-
longs has long been recognized and has presented to the investigator
some of the most baffling problems in plant pathology. The potato-
mosaic group began to assume alarming proportions in the British
Isles and on the Continent toward the end of the eighteenth century
and at the beginning of the nineteenth century (27, 45, 59).2 Peach
yellows was known and much written about in the United States
near the beginning of the nineteenth century (69). Tobacco mosaic
was first described by Mayer in 1886 (52), but more fully treated by
Beijerinck in 1898 (17). Other well-known chloroses will readily
come to mind in addition to those recently discovered or not so
generally recognized.
It is not the purpose of this paper to consider in detail all types
of plant variegation. Chloroses have to do with the reduction or
1The present study was largely carried out under the direction of Dr. R. A. Harper, of
Columbia University, and was completed under the direction of Dr. Erwin FE. Smith, of
the Laboratory of Plant Pathology, Bureau of Plant Industry.
2The serial numbers in parentheses refer to “ Literature cited” at the end of this
bulletin,
76289—22 1
2 BULLETIN 1038, U. S. DEPARTMENT OF AGRICULTURE.
total suppression of chlorophyll, but since a yellowing or bleaching
of normally green parts may result from a wide variety of causes _
chlorosis in itself is a symptom rather than a disease. There are,
for example, the chloroses of etiolation and of the normal autumnal
ripening of leaves. Moreover, a yellowing of chlorophyll frequently
follows upon some types of insect injury, such as that of root aphids
on the peach and the grape, and it is a constant accompaniment of
certain diseases caused by parasitic bacteria and fungi, such as bac-
terial black-rot of cabbage and Fusarium wilts of cabbage and potato.
Again there are the more or less general chloroses due to unfavorable
soil or climatic conditions, and finally those infectious chloroses of
obscure origin which present a fairly regular sequence of pathologi-
cal signs, including fundamental derangements in both metabolism
and morphogenesis.
Some of these chloroses are true diseases in the restricted sense.
Others are not diseases except under a broad application of the
term, and certain forms of chlorophyll restriction are clearly not
diseases at all.
There are, however, two fairly well marked types of chlorophyll
disturbance which are usually included under the chlorotic group of
plant diseases. These are (1) the infectious chloroses which are
communicable through expressed plant juices or through those juices
as directly transmitted within the living plant tissues, and (2) the
noninfectious chloroses due to unfavorable soil or atmospheric con-
ditions.
The present study of pecan rosette deals with the histology and
cytology together with the sequence of gross symptoms of the dis-
ease. In order to place the results of this study in proper relation
to other diseases of this type it is necessary to review briefly some of
the work of other investigators.
CHLOROSES DUE TO SOIL OR ATMOSPHERIC CONDITIONS.
With respect to those chlorophyll changes due to physical or
chemical conditions of soil or atmosphere it is difficult to say just at
what point the normal state ends and chlorosis begins.
Certain plants prefer an acid condition of the soil, others tolerate
it, others are restricted for their optimum development to neutral
or alkaline situations. Nevertheless, neither macroscopic nor micro-
scopic examination of such a plant as field sorrel (Aumewx acetosella
Linn.), for example, would give a clue to its acid-soil toleration. In
response to certain environmental changes, however, Transeau (78)
has shown that this species does develop anatomical changes. In
moist situations, with soil and air temperature approximately iden-
tical, the leaves of this species are relatively large, with a loose
PECAN ROSETTE. 3
arrangement of the tissues, a poorly developed palisade tissue of
one-cell layer, three layers of sponge cells, and an epidermis of
large thin-walled cells with delicate cuticle. On the other hand,
when grown in dry sand or in undrained sphagnum bogs where the
soil temperature was several degrees below that of the air the leaves
were thickened, reduced in size, and revolute margined. The meso-
phyll tissue was more compact, with two or three layers of palisade,
and two layers of sponge cells. The epidermal cells were smaller,
with outer walls and cuticle thickened. In addition to the develop-
- ment of these other xerophilous characters, drops of oil or resin,
characteristic of bog plants, were formed on the epidermis and cells
adjacent to the bundles; these are absent under moisture conditions
more favorable for this species.
Warming (82) states that in acid soils intimately associated with
high water content, in a cold or temperate climate, the tendency of
plants is toward the development of leaf coatings of hairs, papille,
or wax; thickened cuticle; mucilage; erect and ericoid, terete, or
filiform leaves; with bilateral internal structure. Since these char-
_acters develop on wet, moor soils the world over, he considers that
there must be a connection between these soils and the xeromorphic
structure, and that consequently these soils must be “ physiologically
dry.” These facts also account for the xeromorphic structure of
plants in the extreme north or at high altitudes.
The experimental results obtained by Mrs. Clements (23) show
that the xerophyte tendency is toward the development of a diplo-
phyll palisade (bilateral) tissue with restricted air spaces and with
or without water-storage cells. This prolate closely packed type of
cell tends to reduce transpiration. The mesophyte type, on the other
hand, approximates an equal development of palisade and sponge
cells with moderate looseness of structure. The hydrophyte type
consists in the development of simple globose cells and large air
spaces. She found that decreased light and increased water absorp-
tion caused an increase of leaf surface but a decrease in thickness,
while increased light and decreased water absorption brought about
a reduction in leaf surface but an increased thickness. Extremes
of any factors not at the optimum tended toward dwarfing.
Hanson (39) found differences in total thickness between leaves
from the south periphery and the center of the same tree usually
greater than the differences hitherto reported between leaves of meso-
phytic and xerophytic forms of a species. Leaves from the south
periphery, as a whole, developed more palisade, greater compactness
of structure, and thicker epidermis and cuticle than leaves from
within the crown.
~ Halophytes, or “ salt-loving plants,” usually develop thick, fleshy
leaves which are more or less translucent, owing partly to the abun-
4 BULLETIN 1038, U. S. DEPARTMENT OF AGRICULTURE. |
dant cell sap and poverty of chlorophyll and partly to the smaller
intercellular spaces. The thickness of the leaves is caused by the
enlarged roundish sponge cells and by the massive, often transversely
divided palisade. Sodium chlorid thus appears to act morphologi-
cally, Warming says, in much the same way as sunlight (82).
In the cell sap the solution of salt. is more concentrated than in the
soil. Side by side with this increased salt content goes a decrease in
the development of chlorophyll, due to a reduction both in size and
number of chloroplasts. Succulent halophytes usually show at first
a dark-green color, later passing over into a yellowish green or red.
Wax coatings are characteristic of many salt-loving plants. Some
‘species, such as Solanum duleamara Linn., are dimorphous, exhibit-
ing halophytic forms and also inland forms with thin leaves.
Such general environmental factors as those enumerated may and
do have appreciable developmental results. Plants thrive or fail to
thrive and may even die, or again various general adaptive morpho-
logical changes are brought about, but there are no fundamental
derangements in morphogenesis or in metabolism which could prop-
erly characterize these conditions as disease.
In this same category of general environmental effects are to be
placed the calciphilous or lime-loving plants; and the calcifugous
or lime-avoiding plants. Plants grown upon a lime soil, Warming
says, tend to a greater pubescence and to a bluish green color and a
more divided condition of the leaf (82). Moreover, not only are the
chemical but also the physical characters of lime soils to be taken
into consideration. é
It is but a step, now, from these general lime relations to some of
the more specific lime effects usually considered as diseases. In the
pineapple chlorosis of Porto Rico sometimes the plant becomes almost
ivory white. In other cases the leaves are yellowish white with
streaks or patches of green, or the outer leaves may be green while
the later developing leaves at the center are white from the start.
In still other instances the leaves, normally green for several months,
may gradually develop a light-colored mottling until finally the
whole leaf becomes blanched. The plants are dwarfed and the red-
dish or pinkish fruit finally cracks open and decays. Gile (35-37)
has definitely shown that this pineapple trouble is primarily caused
by a lowering of the availability of iron to plant absorption due to
calcium carbonate in the soil. A chlorosis of pineapple in Hawaii
(46) appears to be caused by a similar depression of the availability
of iron due to the high manganese content of the soil.
Similar lime chloroses of rice and of sugar cane in Porto Rico have
been shown by Gile and Carrero (38) to be caused by a lack of suffi-
cient iron absorption. In all these cases spraying with a solution of
PECAN ROSETTE. 5
iron sulphate evokes a development of chlorophyll and at least a
temporary resumption of growth.
Grape chlorosis may also result from this same set of causes and
in such cases responds to the ferrous-sulphate spray treatment.
According to Mazé (53), grape chlorosis may also result directly
from excessive absorption of lime, and in other cases from a defi-
ciency of sulphur in the soil due to lowering of its availability by the
action of lime. Furthermore, as a result of water-culture experiments
he found chloroses of maize due to lack of both iron and sulphur.
Chlorosis in maize was also experimentally induced by the addition
to the solutions of various toxic substances, such as lead or methyl
alcohol.
The overabsorption of lime was reported ays Clausen (22) as caus-
ing a chlorosis of oats in Europe.
These plant relations to various salt constituents of the soil bring
up the questions of plant absorption, antagonism, and changes in per-
meability investigated more recently by Loeb, Osterhout, Brooks,
Waynick, and others. As the result of a careful series of experiments
Waynick (83) concluded that no “optimum calcium-magnesium
ratio” appears to exist and that a balance between all ions present
in a solution appears to be far more important than any single ratio.
He found by chemical analysis that the composition of plants in inor-
ganic constituents may be enormously altered by variations in the
surrounding medium. The permeability of the plasma membrane
appeared to be changed by the nature and balance of the solution
around the roots. The same salt was found to act differently at differ-
ent concentrations, preserving the normal permeability at certain con-
centrations but at other strengths allowing a large penetration. With
regard to each salt tested, its presence in toxic concentration always
resulted in increased permeability of the plant tissues to calcium and
magnesium. On the other hand, normal growth was always accom-
panied by an approximately equal percentage of calcium and magne-
sium in the plants tested; and in nearly all cases where growth was
markedly decreased the amounts of calcium and magnesium were
greatly increased in the tissues. The degree of absorption of any
salt seemed, over a wide range, to be independent of the concentra-
tion present; and growth was the same under widely varying ratios
of calcium and magnesium. The findings of Loeb, Osterhout, and
Brooks were confirmed in that antagonistic salt action tends toward
the preservation of normal permeability.
It will readily be seen that there are many closely intergrading
steps or degrees of environmental effects. These are—
(1) The adaptations without visible changes in structure or metabolism.
(2) The general adaptive changes in anatomy or physiological processes in
response to physical or chemical stimuli from without.
6 BULLETIN 1038, U. S. DEPARTMENT OF AGRICULTURE.
(a) The dwarfing effects of unfavorable soil or climatic conditions.
(b) Starvation phenomena due to insufficiency or absence of essential
nutrients.
(c) Chloroses caused by the absorption of toxic amounts of mineral or
organic soil constituents.
(d) The general chloroses due to insufficient or to overabundant water supply.
(e) Restrictions in chlorophyll development due to reduction of light.
(f) Finally there are chloroses due to lowering of temperature.
All these reactions, however, are rather general effects which are
more or less comparable to starvation, overfeeding, or direct poison-
ing. There are no profound or strictly localized derangements in
both metabolic and anatomical development, and it is a question
whether in the restricted sense some of these phenomena should be
regarded as diseases at all.
INFECTIOUS CHLOROSES.
As opposed to the general chloroses caused directly by soil or
climatic conditions are those specific chlorotic diseases of infectious
nature and obscure origin in which are simultaneously brought about
fundamental derangements in both physiological and structural de-
velopment. Concomitant with the rise of plant pathology as a
science there have come to light an increasing number of diseases of
this type until now it seems apparent that almost every plant group
may have one or more infectious chloroses.
Reports of early scientific investigations upon two of the pencil
types of infectious chlorosis appeared at nearly the same time—those
upon tobacco mosaic by Mayer (52) and Beijerinck (17) and upon
peach yellows by Penhallow (58) and Erwin F. Smith (69, 70).
In peach yellows the sign often first to appear is a red blotching
of the fruit on one or more branches, with the color extending
through the flesh to the pit. A yellowing of the foliage always oc-
curs at some stage of the disease. Another characteristic feature con-
sists in the premature development of the buds of several series into
spindling depauperate shoots with dwarfed and linear and often
curled or inrolled leaves. A premature ripening of the fruit also
usually takes places. The disease ordinarily affects one or more
branches at first, but may develop signs at once over the whole tree.
Penhallow (58) found an abnormally loose cellular structure in the
bark, but a reduction in the size of the cells and an abnormally dense
structure of the wood. Assimilation is profoundly affected and trans-
location of starch is delayed. The leaves become gorged with starch,
and excessive storage occurs in the cortex rather than in the inner
bark and the wood, as in normal trees. The oxidizing enzyms are
increased in the diseased leaves, and a larger tannin content has been
found in the diseased fruit. In one instance (18) delayed starch
translocation was also found in an apparently healthy branch con-
PECAN ROSETTE, q
tiguous to a diseased branch, showing that external signs may be
preceded by deep functional disturbances. Conditions unfavorable
to growth tend to intensify the signs of yellows but do not cause
the disease (71). Yellows is transmitted by budding or grafting
from diseased trees to healthy stock, and infections through the
roots as well as the stems may take place in this way. However, all
attempts to infect with the expressed plant juices have thus far
failed, nor have insect relations been discovered.
Peach rosette (70, 72) and little peach (73) differ in the charac-
ter and sequence of the signs, but are similar in type to peach yel-
lows. All three diseases induce deep changes in assimilation, trans-
location, structure, and development. All may appear first in one
branch, and they are transmitted by budding or grafting. The dis-
ease progresses gradually from the point of infection, requiring
longer for the development of external signs in the top when infected
by root grafting than when grafted into the branches. That rosette
may not affect the whole tree at once was shown in one case where
- buds from branches showing external signs transmitted the disease,
whereas those from the apparently healthy side failed to give infec-
tion. The normal side of this tree, however, developed rosette the
following season. The outer leaves of rosettes fall early.
In spike disease of the parasitic sandalwood we find a close re-
semblance to the peach-yellows group. The spikelike appearance of
the leaves standing out stiffly from the branches suggested the name
“spike disease.” The entire tree is not attacked at once over the
whole top, but symptoms appear first on one, then on several branches,
and gradually spread over the tree. The internodes become short-
ened and the leaves reduced in size and narrowed. The continuous
development of buds into new leaves and branches throughout the
year produces a growth closely resembling the “ witches’-brooms” of
the peach, and with the progress of the disease the leaves become
smaller and more chlorotic. No blossoms or fruit are borne in the
later stages, though sometimes flowers and fully developed fruit are
formed on portions of trees still retaining their normal appearance.
Death of the haustoria and fine root ends keeps pace with the prog-
ress of the disease.
Spiked leaves and branches contain a marked accumulation of
starch (24). In the diseased leaves this starch is distributed through-
out the parenchyma, especially in the sheaths of the fibrovascular
bundles, and no marked difference in quantity is found at different
periods of the day. In the diseased twigs the starch occurs as grains
of considerable size in the pith, in the medullary rays, in the wood,
and in the bast fibers; whereas in the normal twig such grains are
rarely found except when the leaves are fully matured. This accumu-
lation in the twigs precedes external signs of the spike disease both
8 BULLETIN 1038, U. S. DEPARTMENT OF AGRICULTURE.
in natural and artificial infections. Diastase activity in the healthy
leaves was found to be almost double that in the spiked leaves. The
mesophyll tissues are hypertrophied and the leaves increased in
thickness and the vascular tissues become reduced in advanced stages
of the disease. Chemical analysis (24) shows a higher percentage of
nitrogen and of most of the ash constituents in healthy leaves. Since
young, healthy leaves pass through a stage comparable in chemical
composition to that of spiked leaves, it seems probable that in the
latter case development has for some reason been checked in the early
stages of growth.
In a large percentage of Coleman’s grafting experiments (24)
spike was successfully transmitted to originally healthy stocks, and
in almost every case the disease first appeared in the stock on branches
closest to the point of grafting and spread from these regions to the
other parts of the stock. A considerable time always elapsed before
external signs of disease appeared. In all cases examined, infection
had spread to the roots and resulted in the death of the root ends
and haustoria.
The occurrence of spike disease is not dependent on the fertility
of, the soil, nor does injury to the roots have any relation to the dis-
ease. Venkatarama (81) found experimentally that isolation of the
trees from all possible hosts by digging trenches did not cause signs
of spike even after two years under observation. Cutting the root
connections and removing the haustoria, injecting the lateral roots
with strong sulphuric acid, and girdling to the heartwood did not
cause the disease. Thousands of trees previously growing under a
heavy covering of vines were exposed to the light, but the increased
loss of water resulted in nothing resembling spike disease. Fischer
(34) states that the spike disease spreads from a center, not appear-
ing simultaneously over considerable areas.
In tobacco mosaic the yellow and green mottling of the leaves is a
prominent sign. Not only leaves but also the calyx may be mottled,
and the corolla becomes flecked with red and white blotches instead
of exhibiting the normal even red or white color. The light areas of
the leaves are usually slower growing than the green areas, thus often
resulting in distortions which may become extremely marked in
young leaves. Often, however, such leaves almost recover from these
malformations as they mature (1). Sometimes the lamine are al-
most suppressed, and in other cases a long, sinuous, ribbonlike leaf
is produced. In many cases abnormally dark-green blisters develop
on the immature leaves.
Koning (47) and Heintzel (40) reported a separation of the cells
in diseased foliage which often leaves spaces nearly as large again as
the cell itself. The chlorophyll bodies become distributed in irregular
groups, chlorophyll disappears, the cell walls disintegrate, and finally
PECAN ROSETTE. 9
complete disorganization follows. Woods (84, 85) found that in the
lighter areas of badly diseased leaves the palisade parenchyma had
not developed at all, but the tissue consisted entirely of a respiratory
parenchyma with cells packed together rather more closely than nor-
mal. In healthy leaves the palisade cells were four to six times as
long as broad, whereas in the moderately diseased leaves these cells
were almost as broad as long. The leaf surface becomes depressed.
in the light areas and raised in the green areas, thus giving a rough-
ened appearance to the lamina.
As first shown by Woods (84), the oxidizing enzyms are greatly
increased in the diseased areas. He also found more starch in the
form of granules in the yellow areas than in the green areas of the
same leaf. The cells were often completely gorged with starch. Ex-
amination in the early morning showed only a slight decrease, while
healthy tissue at the same time was empty or contained only a trace.
Starch translocation in the diseased leaf is greatly delayed in spite
of the fact that diastase is present often in larger amount than in the
normal leaf, and Hunger (41) from experiments in vitro concluded
that the retarding effect upon diastase action is caused not by the
oxidizing enzyms, but by reducing substances including tannin.
Mayer (52) first showed that transmission of tobacco mosaic
could take place through the expressed juices of diseased plants.
Iwanowsky (42), and Beijerinck (17) independently demonstrated
that the infective principle would pass through the pores of a Cham-
berland filter, though such a filtrate was less infective than the un-
filtered juice. Allard (4) proved that infection fails to result after
the juice from diseased plants has been passed through a Livingston
porous-clay cup filter. Transmission of the disease by an infective
principle in the expressed juice was thoroughly demonstrated, and it
was shown by Allard and others that oxidizing enzyms do not con-
stitute this infective principle (4). Such plant juices diluted to 1 to
1,000 in water were quite as infective as the undiluted juice; attenua-
tion was indicated at 1 to 10,000, while at greater dilutions infection
was found unlikely to take place (2). The virus is infectious to all
susceptible plants, but such plants never develop mosaic so long as
chances for infection are excluded, and this regardless of soil and
climatic conditions. The infective principle may be present in all
parts of a diseased plant except within the seed and has been demon-
strated even in the trichomes (3, 6). Furthermore, infection may
occur through inoculation of the trichomes alone. Cutting the mid-
rib at the base or severing the larger veins on one or both sides does
not prevent the final dissemination of the infective principle to all
parts of the leaf and to other leaves of the plant. Environmental
conditions may partly or even wholly mask the external signs for a
time, but can neither cause nor cure the disease.
76289°—22 2
10 BULLETIN 1038, U. S. DEPARTMENT OF AGRICULTURE.
Nishimura (55) found that the bladder cherry (Physalis alkekengt
Linn.), after inoculation with tobacco-mosaic virus is capable of act-
ing as a carrier without itself showing external signs of the disease.
Cross-inoculation experiments (5) have shown that mosaics in to-
bacco, tomato, pepper, and petunia are caused by the same infective
principle. On the other hand, the mosaic of pokeweed (Phytolacca
decandra Linn.), though readily transmitted by expressed plant juices
within the species, was not found to be cross inoculable on tobacco or
vice versa (8). Tobacco mosaic is transmissible by insects.
Another type of infectious mosaic is that carefully worked out by
Baur (11-16) in the Malvacee. He showed that the Abutilon
mosaic is transmitted only by grafting and not by inoculation with
the expressed juices, as occurs in tobacco mosaic. As in the latter
disease, however, seed transmission does not occur. He found that
when scions of the immune Abutilon arboreum Sweet are grafted
on the variegated A. thompsoni Hort. they grow vigorously and re-
main apparently normal. However, if scions of the green but sus-
ceptible A. indicum Sweet are now grafted upon the immune 4.
arborewm they become infected and develop the typical mottling. On
the other hand, the contagium passing through the immune A. arbo-
reum is not capable of remaining there and giving infection if this
portion of the shoot is subsequently grafted into a susceptible stock.
In the case of the immune Lavateria arborea Linn., however, there
is no transmission at all when double grafted between the mottled
Abutilon thompsoni and the green susceptible A. indicum. Baur
succeeded in transmitting this mosaic by grafting to about 50 species
and varieties of Abutilon and related plants.
Tt was found that if the leaves of variegated plants were removed,
or if the shoots were cut back so that no leaves remained and the
new shoots developed in darkness, only the first two or three leaves
were mottled. If these mottled leaves were then removed the plants
remained permanently green in the light unless they were grafted
with scions from other variegated plants. However, if axillary buds
on old parts were forced into growth these produced shoots with
mottled leaves which in turn infected all the newly formed leaves
on the plant. Furthermore, when scions of a green but susceptible
variety were grafted upon defoliated, mottled plants the scions re-
mained green; but here again, if a mottled shoot was allowed to de-
velop from the stock it rapidly infected the whole plant. The con-
tagium is,’ therefore, capable of infecting only the embryonic leaves,
and in the buds it may be stored up for months in inactive form.
2'The term “ contagium ” has been suggested by Dr. H. M. Quanjer as synonymous with
any infective principle, whether of known or unknown origin. Its use in the place of
“virus” with reference to the so-called “ filterable-virus ’ diseases does away with ob-
jectionable connotations and leaves nothing to be taken back.
PECAN ROSETTE. 11
In varieties where the size and distribution of the yellow spots
made it possible, Baur found that by carefully cutting out all yellow
spots and continuing this process on all newly developing leaves for
one or two weeks, finally green leaves only were formed. From
this result he considered it certain that the contagium is present in
the yellow spots but only in sufficient quantity to infect, about three
or four newly developing leaves at the growing point. After this it
is apparently used up, and leaves subsequently formed remain green.
Darkening the assimilating leaves of a mottled plant led to a similar
-result. Here the first leaves to develop thereafter were yellow
spotted, but if these also were darkened before they began to assimi-
late, the subsequently developed leaves were all green.
Girdling experiments demonstrated that the contagium is carried
only through the bark. Different species and varieties of Malvacez
were found to vary widely in susceptibility and also in the incubation
period, but several days at least are required. Baur also demon-
strated infectious chloroses in Fraxinus, Laburnum, Sorbus, Ptelea,
Euonymus, and Ligustrum.
In potato mosaic the mottling is irregularly distributed irrespec-
tive of the venation; and, moreover, profound dwarfing, with curling,
erinkling, and further distortions of the foliage occur in the more
severe attacks of the disease. The parenchyma tissues are less per-
fectly developed in the light-colored areas, the palisade cells tend to
shorten up and chlorophyll development is restricted. Potato mosaic
(65, 67) is transmitted by the tubers, by grafting, and by inoculation
with the expressed juices of diseased plants; and it is also dissemi-
nated by aphids.
In potato leaf-roll the leaves become inrolled from the margin,
reduced in size, and of a paler green to yellowish cast. A necrosis
of the phloem (60-62) seems to be characteristic of the disease, but
whether this condition is specific for leaf-roll is still a moot point.
The disease is transmitted by means of the tubers, by grafting, and by
insects (66).
In both potato mosaic and leaf-roll portions of the healthy plants
growing near diseased plants contract the disease, but not all tubers
from such secondarily infected plants necessarily develop diseased
progeny. Affected leaves in both these diseases exhibit: deep-seated
assimilatory derangements, including increase in activity of the oxi-
dizing enzyms and gorging of the leaves with starch, with greatly
delayed starch translocation. In both these diseases tuber forma-
tion is greatly restricted. In neither case are these diseases induced
or cured by soil or climatic conditions. However, certain environ-
mental factors may temporarily mask the external signs though such
diseased plants still retain their power to infect.
12 BULLETIN 1038, U. S. DEPARTMENT OF AGRICULTURE.
The infectious chlorosis of the sugar beet known as curly-top is not
transmitted by expressed plant juices nor by way of the seed, but is
disseminated by the sugar-beet leafhopper (HL utettia tenella Baker).
This insect is capable of producing infection only aftetf a definite
incubation period subsequent to feeding upon diseased plants. <Ap-
parently there is no other agent of transmission for this disease
(10, 68).
Some extremely interesting insect relations of spinach blight have
recently been worked out by McClintock and Loren B. Smith (49).
Not only were healthy plants successfully inoculated by needle,
pricks with the contagium from diseased plants and with the crushed
juice of aphids fed upon diseased plants but the potato aphid
(Macrosiphum solanifolii Ashmead) and the spinach aphid (ho-
palosiphum persicae Sulzer), free from infection at first, were demon-
strated to transfer the blight to healthy spinach after feeding upon
diseased plants. Control plants invariably remained healthy. Later,
these two species were obtained from four different States where
spinach blight did not occur, and they failed to induce the disease
on healthy plants until after they had fed on blighted spinach. The
same two species collected locally and tested at the same time pro-
duced the disease. These investigators demonstrated that the con-
tagium may be carried from spring to fall by a direct line of aphids.
Transmission tests with several other species of insects gave nega-
tive results, thus also tending to show that the insect relation to
spinach blight is not that of a purely mechanical disseminator.
Sugar-cane mosaic has been shown by Brandes (19-21) to be
transmitted by cuttings, by expressed juices from diseased plants,
and by certain insects (A phis maidis Fitch) fed upon infected plants.
No evidence of seed transmission was found. Insect transmission
of corn mosaic has also been demonstrated by Brandes.
That cucurbit mosaic is transmitted by the expressed plant juices
and by insects has been definitely proved.by Doolittle (28), by
Jagger (43, 44), and by Doolittle and Gilbert (31); and the latter
investigators have apparently shown that at least in some cases the
disease is carried over by the seed (30). In this disease both foliage
and fruit become yellow mottled and distorted, and growth of.the
entire plant is seriously checked. The dark-green portions of dis-
eased leaves are slightly thicker than normal, thus accounting for
their blistered and distorted appearance. The yellow areas, though
thinner than contiguous dark-green parts, are of about the same
diameter as in the normal leaf. The palisade cells of the green areas
are crowded closely together and are somewhat longer and narrower
than in the normal leaf. In the yellow parts these cells are more
nearly isodiametric and less in number than normal per unit of area.
PECAN ROSETTE. ; 13
The spongy parenchyma of these parts is also more compact, and
the intercellular spaces are smaller than in the green areas. The
chloroplastids are decidedly smaller and often are pressed so closely
against the cell wall as to be almost invisible. In the fruit the
structural derangements are similar in general to those occurring in
the diseased leaves.
Dilutions of the virus up to 1 to 1,000 were found to be just as
potent as the undiluted juice expressed from diseased plants, but at
dilutions greater than 1 to 10,000 no infections took place. Where
infections took place with the higher dilutions, the incubation period
was no longer than when undiluted juice was used, thus showing a
rapid reproduction of the virus within the plants. This virus was
found to be entirely removed by passage of the expressed juices
through porcelain filters of the finer grades (29).
Taubenhaus (76) reported experiments in which mosaic of sweet
peas was transmitted by insects and by needle inoculations with plant
juices.
Reddick and Stewart (63, 64, 75) found mosaic of beans trans-
missible by rubbing the young leaves of normal seedlings with
crushed leaves from mosaic plants and obtained a high percentage
of mosaic by sowing seeds from diseased plants. In cases of inocu-
lation external signs usually appeared after about four weeks.
Many other chlorotic diseases such as aster yellows, cassava leaf-
curl, mulberry dwarf, cotton leaf-curl, little-leaf of the vine, citrus
mottle-leaf, raspberry leaf-curl, apple rosette, and mosaics of peony
and sweet potato present characters suggesting a possible relation to
the group of infectious diseases. However, they have scarcely re-
ceived sufficient study for any final statements regarding infectivity
or causes. With this brief general review the particular ‘disease
under investigation may now be considered.
STUDIES OF PECAN ROSETTE.
RESULTS OF PREVIOUS WORK.
Pecan rosette was recognized by orchardists as far back as 1900,
but no early published references to the disease have been found.
Field investigations by W. A. Orton were undertaken in 1902 and
continued about four years. During the years 1910 to 1913 field
studies were carried out independently by the writer. The results
of these two sets of field studies were brought together and published
as a joint paper (56). With the exception of a few brief references
to thesdisease, this was the first published account of pecan rosette.
The disease is fairly well distributed over the pecan-growing
regions of the Southern States, but has not been reported from the
northern limits of pecan culture.
14 BULLETIN 1038, U. S. DEPARTMENT OF AGRICULTURE.
TRANSMISSION.
In the investigation by Orton and Rand (56) negative results
were obtained with inoculations using bits of diseased buds or tissue
taken from beneath the bark of rosetted shoots, inserted into slits
in the terminal branches of healthy nursery trees. Cultural methods
and microscopical examination likewise gave negative results, show-
ing the apparent absence of fungi or bacteria in still living rosetted
twigs and branches.
Normal buds or scions worked on rosetted stocks all developed
rosetted shoots except in one case where the stock itself recovered.
On the other hand, rosetted buds and scions worked on apparently
healthy stocks usually developed into normal shoots; in the cases
where rosette did develop in such buds or grafts the percentage was
no greater than in adjacent stocks worked with supposedly normal
buds or grafts in the commercial propagation.
ENVIRONMENTAL RELATIONS.
The observations and orchard records by Orton and Rand (56)
showed that pecan rosette is not absolutely limited to any soil type,
topography, or season. The disease was found at least to some ex-
tent in practically all kinds of soils where pecans were observed,
with the single exception of parts of some orchards where the land
tended to be swampy. In the latter case very little growth was
made, and the trees finally developed a diffuse general chlorosis,
but no signs of any phase of rosette. The disease, however, was
observed to be particularly prevalent under poor soil or cultural
conditions for the species, such for example as in the dry uplands
of Texas, or the washed-out hillsides of the Southern States. The
disease was found to be comparatively rare in the alluvial river bot-
toms of Texas, Louisiana, and Mississippi, where the tree is under
native environmental conditions.
In most cases where rosetted trees were transplanted into appar-
ently better local soil conditions, the larger percentage of such trees
and often all of them resumed normal development; and all rosetted
nursery trees recovered when shipped from the South and trans-
planted in the open or in potted garden soil at Washington.
Nearly all healthy trees used to replace rosetted orchard trees
subsequently developed the disease ;-whereas only about half of those
replacing healthy trees later contracted the disease.
In a three-year fertilizer test on level uniform soil cases of rosette
developed in 9 out of 11 plats where lime was used; and the largest
number of cases and severest attacks occurred on two plats each
receiving lime* and acid phosphate, in one case combined with
4 Fertilizers were applied at the following rates: Lime (CaO acted on jointly by air
and water), 1 bushel per tree; nitrate of soda, 8 pounds; cottonseed meal, 32 pounds;
muriate and sulphate of potash, 8 pounds; acid phosphate, Thomas phosphate, and
ground bone, 24 pounds; stable manure, a liberal application.
PECAN ROSETTE. 15
muriate of potash, in the other with nitrate of soda. The two limed
plats free from rosette received in addition cottonseed meal and
Thomas phosphate, respectively. In the five plats without lime no
rosette at all developed with the exception of doubtful signs in two
trees immediately contiguous to a limed plat. The four lime-free
plats showing no traces of rosette were the control, untreated, and
three plats treated respectively with muriate of potash and acid
phosphate, stable manure alone, and stable manure with ground
bone. During this period no other cases of rosette developed in the
vicinity of the experimental block, though cases appeared in other
parts of this orchard of 700 acres. In two other fertilizer tests where
the disease was already present at the start, it increased somewhat
in severity of attack or in the number of new cases in the plats
receiving lime.
Analyses of the subsoil around normal pecan trees in parts of an
orchard free from rosette gave 0.5 to 9.5 per cent of calcium. It
appears then that the disease is not caused by the presence of lime
alone, since more lime occurred here than in parts of the orchard
where rosette was present. Ash analyses of normal and diseased
Jeaves and twigs showed only slight or highly variable differences.
Apparently, however, the percentage of potassium is greater in the
diseased leaves and twigs.
In one spray test with Bordeaux mixture on rosetted trees nega-
tive results were obtained.
It is evident from numerous orchard records covering periods of
2 to 12 years that pecan rosette fluctuates from year to year without
any variation in fertilization or cultural methods. The diseased
trees may apparently make a complete recovery and remain normal
for an indefinite period, or after one or more years may again con-
tract the disease. However, in the majority of cases of recovery
observed, the trees had not reached the stage where the branches
were dying back. It seemed thus (56, p. 165, 169) highly probable
that seasonal climatic changes, such as variations in precipitation or
moisture content of the soil, might have at least an indirect relation
to the prevalence of rosette. In large orchards the more or less
simultaneous appearance of rosette in patches, and its usual limita-
tion to these areas, suggested some connection with soil phenomena.
From the apparent nontransmissibility through the seed, negative
results in attempts at isolation of organisms, the apparently nega-
tive results of budding and grafting between normal and diseased
trees, and the results of transplanting tests it was concluded that
the disease was probably nonparasitic.
From pruning and “ dehorning ” tests, transplanting and fertilizer
experiments, dynamiting of soil around rosetted trees, and from
orchard records of disease fluctuation it was at that time considered
16 BULLETIN 1038, U. S. DEPARTMENT OF AGRICULTURE.
highly probable that pecan rosette belonged to the group of non-
transmissible chlorotic diseases caused by improper nutritive supply
or injurious physical conditions. The possibility of the presence of
some parasitic organism was not entirely precluded, but it was
thought highly probable that the ultimate cause would be found in
some lack of balance in nutritive supply, or possibly in some toxic
substance or substances in the soil. These conclusions were based ~
entirely upon the results of field experimentation.
Miller (54) observed that buds from rosetted trees worked upon
healthy stock in most instances grew into normal trees, but that
when a tree was decidedly rosetted its buds would sometimes develop
the disease when worked upon healthy stock. He is of the opinion
that rosette is due to soil relations and observed that in*dry seasons
the disease is more prevalent. Rosette and a proper amount of
moisture, he says, do not go together. Impoverished soil, lack of
humus, overstimulation of growth, and use of improper fertilizers
all favor rosette. He states that the severest cases are incurable or
at least not curable by practical means.
Fawcett (33) refers to pecan rosette in a short paragraph, and
Crittenden (26, pp. 44-45) briefly summarizes the work of Orton
and Rand.
Matz (51, pp. 139-141) in a bulletin relating to various pecan dis-
eases and insects devotes a short section to rosette. He states that
the disease apparently occurs on all types of soil and at all seasons,
but that wherever it occurs it is most abundant during late summer.
He also says that rosette is more abundant on higher and more ex-
posed soils than in low and more protected situations. Aiter
briefly describing the signs of the disease he adds that many of the
dead leaves adhere to the branches throughout the winter unless
blown off by strong winds. The disease is favored by planting in
open, sandy soil or where a hardpan exists near the surface.
MecMurran (50) in a paper dealing with field experiments and
observations states that rosette is generally considered to be the most
serious pecan disease. It is found upon a wide range of soil types
and under various conditions of culture and fertilization; but one
factor, he says, appears to run through it all. On the river flood
plains of Southern Louisiana the disease is practically unknown.
Here the soil is deep and black and of high fertility and presumably
of high water-holding capacity as compared with the typical sand,
sand-clay, and clay soils of the Atlantic and Gulf Coastal Plains
where the disease is so prevalent. In Georgia and Florida probably
90 per cent of the affected trees are on hilltops and slopes. All
cases on the bottoms that have been examined were found to be in
deep sand or in a clay or sand-clay soil underlain at 2 or 3 feet by
sand. It was noted further that large healthy trees 5 to 10 years old
Bul. 1038, U. S. Dept. of Agriculture. PLATE I.
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DIFFERENT STAGES OF PECAN ROSETTE.
[ILLUSTRATION BY J. F. BREWER. |
Fie, 1.—Enlargement of one of the yellow spots on a pecan leaf in the secondary stage of rosette showing
the distortion of vein islets and their radial expansion around a focal center. X about 30. Fre. 2.—
Pecan leaflet in the secondary stage of rosette, showing the distribution of yellow spots between the side
veins and the crinkling and roughening of the leaf surface. Natural size. Ftc. 3.—Pecan leaflet
showing the red-brown stage of rosette which often follows either the primary or the secondary stage.
Natural size. F1e@. 4.—Pecan leaves in the primary stage of rosette, showing no external signs of the
disease except the yellow mottling between the side veins. Natural size.
RP AT Ce
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Bul. 1038, U. S. Dept. of Agriculture. PLATE II.
NORMAL PECAN LEAF, FROTSCHER VARIETY.
Collected on August 25, 1919, at Thomasville, Ga. Photographed by transmitted light to show
the texture and opacity of the leaf. x 3.
Bul. 1038, U. S. Dept. of Agriculture. PLATE III.
fee
SSS LESSOR,
Mee 5G PE ,
ROSETTED AND MOTTLED LEAVES OF PECAN, FROTSCHER VARIETY.
Collected on August 25, 1919, at Thomasville, Ga. Photographed by transmitted light to show
reduction in size, yellow mottling, and abnormalities in form and texture of the leaves. X 3.
Bul. 1038, U. S. Dept. of Agriculture. PLATE IV.
MOTTLED LINEAR LEAVES OF PECAN, FROTSCHER VARIETY.
Collected on August 25, 1919, at Thomasville, Ga. Photographed by transmitted light to show
abnormal shape and texture. x 4.
Bul. 1038, U. S. Dept. of Agriculture.
PLATE V.
FREEHAND VERTICAL SECTIONS OF PECAN LEAVES, FROTSCHER
VARIETY.
Semidiagrammatic camera-lucida drawings of diseased and healthy leaves, showing tissue
changes due to rosette. Collected on August 25, 1919, at Thomasville, Ga. All of same mag-
nification. The small side diagrams show the relative size and shape of the leaves and the
locations of the sections. A.—Section through the center of a thin, yellow area, showing
close packing of the cells and lack of differentiation. B.—Section at the margin of the
vellow spot in the green portion of a leaflet, showing the close packing of the cells and
the partial decrease in the long axis and increase in the short axis of palisade cells. C.—Sec-
tion through a healthy leaflet, showing palisade and spongy tissue well developed and the
looser arrangement of the spongy cells, the narrower openings being intercellular spaces.
D.—Section of a mottled, linear leaflet, showing the close packing of the cells and only partial
differentiation of the palisade cells. Drawings by the writer.
PECAN ROSETTE. d af
often showed marked signs of rosette the first year after trans-
planting. Trees in low situations where humus and fertility ac-
cumulate from year-to year were almost always found to be uni-
formly vigorous and free from disease. Briefly stated, 90 per cent
of the cases of rosette were found under conditions indicating lack
_of humus, plant food materials, and moisture.
Fertilizer tests (50) showed in two years a marked improvement.
in rosette cases and also many cases of [apparent] recovery. Stable
manure, particularly, gave excellent results. Rosetted trees in the
plat that received ground limestone at the rate of 3 tons to the acre
not only failed to improve but were more severely attacked at the
end of the third season following its application than at the be-
ginning.
Examination of large numbers of trees (50) showed that the feed-
ing roots are distributed through the surface soil, and in proportion
as this is deep and fertile do pecan trees usually attain their normal
development and vigor. Long hot, dry periods often kill many of the
feeding roots in the shallow surface soils; and deep sand, clays under-
lain by sand, and eroded hillsides were found particularly to favor
rosette. An acid soil, according to McMurran, is probably not the
cause, since river flood plains nearly all exhibit an acid soil, and
pecan rosette under these conditions is a rarity.
EXTERNAL SIGNS OF ROSETTE.
Every phase of the disease is observed on trees of all ages, from
young seedling or budded and grafted stock in the nursery row to
trees of long-established maturity.
In every distinct case the constant sign of rosette consists in the
final development of undersized, more or less crinkled and yellow-
_ mottled leaves (Plate I, fig. 2; Pls. If to IV), particularly at the ends
of one or more branches. This phase may be properly designated as
the secondary stage of the disease. The chlorotic areas are situated
between the principal veins, while portions adjoining these veins and
along the margins of the leaflets are green. In severe cases these in-
tervascular chlorotic areas are thinner than in healthy leaves, while
along the midrib and principal veins the blade is often somewhat
thicker than normal. This condition gives the leaf a peculiarly rough
and furrowed appearance and causes the veins to stand out char-
acteristically: Such leaves do not attain their normal size, are
often linear (Pl. IV) and otherwise malformed, and present a
crinkled or undulated appearance of the lamine. Parts of the
lamine are often suppressed ; sometimes the leaflet consisting merely
of the midrib bordered by an edging of ragged tissue. In lamine
otherwise fairly normal in general form, portions of the mesophyll
76289°—22——_3
18 BULLETIN 1038, U. S. DEPARTMENT OF AGRICULTURE.
tissue occasionally fail to develop; as a result of subsequent growth
stretching in continguous tissues the blade becomes dotted with
smooth-margined holes suggestive of healed insect punctures.
During the early course of the disease, or in cases of very slight
attack, yellow mottling may be the only external sign (PI. I, fig. 4),
the size, shape, and texture of the leaves appearing normal. This
primary stage is less characteristic than the secondary stage, but
here also the chlorotic mottling is confined to areas between the prin-
cipal side veins. The regions along the veins and leaf margins
remain green. With advance of season the chlorotic areas of both
primary and secondary stages often turn a dark reddish brown.
(Pl. I, fig. 3.) Signs may appear over the whole tree at once, but
frequently only one or more branches on a tree are affected at first.
Early indications of rosette often consist in the appearance late in
the season of a few mottled leaves near the tip of one or more
branches, the remainder of the tree appearing normal. Leaves de-
veloping signs earlier in the season often present a general bronzed
appearance during late summer and fall, and particularly is this
true under conditions of drought.
Later, where the branches also become affected, there is considerable
reduction in growth, so that the aborted leaves become clustered
together on a shortened axis, giving the characteristic bunched appear-
ance of the foliage at this stage (Pl. III, figs. A and C). It was
this close bunching of the leaves that led Orton originally to apply
the name “rosette” to the disease (56, p, 151). A few nuts are
often borne on branches not too severely attacked, but they are
usually malformed and reduced in size.
Affected trees may continue thus for several years, or they may
appear to recover completely after showing moderate signs for
one or more seasons. However, in severe cases where the signs have
spread over the whole tree and in some instances where only one or
more branches are severely attacked, the affected branches begin
to die back from the tip during the latter half of the growing season.
At first brownish spots and streaks develop in the chlorophyllous
inner bark, and these dead areas increase in size until the bark and
cambium are disorganized and the end of the twig or branch dies.
This staghorn phase is followed during the current and subsequent
seasons by development of abnormal numbers of shoots from dor-
mant and adventitious buds. Usually in young rosetted trees the
first shoots of the season are abnormally large and succulent, and the
leaves are dark green and larger than normal. This is probably in
part due to the severe pruning induced by the staghorn phase, since
‘similar results are obtained by severe artificial pruning during
earlier phases of the disease. Toward midseason, however, the mot-
tling begins to appear and the later developed leaves present the
PECAN ROSETTE. 19
dwarfed, mottled, and roughened appearance typical of the second-
ary phase. Dormant and axial buds of one to several series may and
usually do prematurely develop into depauperate shoots, and toward
the end of the season clusters of dwarfed branches are usually put
out from dormant and adventitious buds farther back on the branches
or main trunk. With each repeated sequence of premature abnormal
growth and subsequent dying back of the branches, the new twigs
and leaves tend to become more and more depauperate, so that a well-
marked case of several years standing presents a characteristically
gnarled appearance.
HISTOLOGICAL AND CYTOLOGICAL STUDIES.
No less striking than the external changes brought about by
rosette are the internal abnormalities of structure and metabolism
in the leaf. The abnormal histological characters develop with the
development of the leaf, and the most far-reaching internal derange-
ments are found in cases of the greatest external malformation.
Within the yellow thin areas between the larger side veins (second-
ary stage) the tissue is usually less developed than normal, and the
cells are more closely packed together (Pl. V, figs. A and B; Pl. VI,
figs. E and F). In less severe cases the palisade tissue is well de-
veloped, but the intercellular spaces become almost obliterated (PI.
VI, figs. A, B, and D). In other instances the palisade cells are dif-
ferentiated, but the individual cells are greatly shortened vertically
(Pl. V, figs. B, C, and D) and may be only two or three times as
long as broad, instead of seven to ten times, as in the normal leaf.
In the most severely affected leaves with extreme variations in leaf
diameter there is no differentiation into palisade and sponge tissue
at all in the center of these yellow spots (Pl. VI, figs. E and F), but
the tissue within these parts of the leaf consists entirely of closely
packed, more or less isodiametric cells without conspicuous air spaces.
Under these conditions the number of cell divisions may be somewhat
increased, the cells remaining smaller than normal (PI. VI, figs. E
and F'). Usually, however, the number of divisions is reduced, and
in some instances a parenchyma tissue only three cells deep has been '
found. (Pl. V, fig. A.) Occasionally the entire tissue between the
margin and the midrib consists of undifferentiated cells. That this
lack of differentiation is not due to immaturity was shown by ex-
amination of young healthy leaves just after their emergence from
the bad. Even at this young and only partially expanded stage the
palisade tissue of healthy leaves was found to be well differentiated
(PLVEE exc }.
In the cmeened portions along the veins, or r sometimes throughout
the lamine in the aborted nonmottled fee of depauperate rosettes,
the size of cells may become increased in all three dimensions Souiantt
20 BULLETIN 1038, U. S. DEPARTMENT OF AGRICULTURE.
essential change in ratio between spongy and palisadetissue. (PI. VI,
figs. G and H; Pls. VIII and IX.) In some cases a slight increase
in number of cell divisions also takes place (Pl. VI, fig. E). The
individual cells appear abnormally healthy, and increase in leaf
thickness here is due largely to increase in size of the individual cells
(Table I). Along with this increase in cell size there is a reduction
in size of intercellular spaces within the spongy parenchyma, and in
the severely attacked leaf an almost complete obliteration of air
spaces results (Pl. VI, figs. B to F).
TABLE I.—Tissue and cell measurements of normal pecan leaves compared with
those of leaves hypertrophied with rosette.
Tissue and cell measurements (microns).
Variety and leaf tissue.
Rosette, about Rosette
Normal. haifcva, Rosette, aborted. acd
—_——[—$— EE
piceige variety, Thomasville,
Palisade cells...dimensions. .| 50 to 52 aby 5.5 to 6.) 57 oe Ay 6.5t09.5 Fee es St oy B.5 tolost! Fe
Spongy tissue. ...thickness..| 87 to 100........... 16:10,116 52 ce cenis| ES TONG 2a. 22 | eee
Spongy tissue where palisade mare Getase Spice sae ee 33 to oe LEG ESS Seba Gp él ye oO pick wt S2UE See. ete
tissue is lacking:
ip Weep POT 9.4 to 12.6. .-...... 10:to 12.2555 25<5 .22 11.2 to 14:40.03- 2418 2s285 5
Bee ors cine 6 thickness.
Lower epidermis...do....] 10.3 to ll.......... 10 to.ll-.s. sachs 2 9.4: to 11s. . 3g58..]2-,-832 yi.
Van Deman variety, Cairo, Ga.
Palisade tissue... .thickness..| 53..........-.-...- 69. sce scccdegee tenes 3. Gst ae ces tbe= sod 57
Spongy tissue......... GOo on] S8see sien sane ace 1 OTe kde BASSE Sobel case sense ee eS 94
Linear leaves may or may not show a differentiation into palisade
and spongy tissue, but where the palisade cells are developed they
are usually shortened vertically and the spongy cells of the paren-
chyma are more closely packed together than the cells in healthy
leaves (Pl. V, fig. D). The average thickness of these leaves tends
to be less than normal and this reduction is due partly to decrease
in the number of cell divisions and partly to the shortened vertical
axis of the palisade cells. The elongated shape of the leaves, how-
ever, is not due to variations in cell shape but rather to a decrease
in the number of cell divisions in which the central spindles are
perpendicular to the midrib. This would tend to keep the cells closer
to the main water supply of the leaf.
Amelung (9) working with plants and Conklin (25) working
with animals have shown that normal tissue cells of corresponding
organs or parts of organs within a species or variety are in general
of the same size and that the size of organs is primarily due to
differences in the number rather than in the size of cells. Mrs.
Tenopyr (77), as a result of investigations in several species of
plants, found that difference in the shape of leaves of the same
plant or related species is not correlated with difference in the shape
of their cells. Linear leaves are not composed of longer, narrower
Bul. 1038, U. S. Dept. of Agriculture. PLATE VI.
*VERTICAL SECTIONS OF PECAN LEAVES KILLED WITH CARNOY’S FLUID.
Fleming’s triple stain used. A, C, and E to H, Van Deman variety. Collected in July, 1913, at
Belleview, Fla. A.—Section of healthy leaflet at the margin, showing well-defined palisade
and spongy parenchyma and large intercellular spaces. B.—Section of a yellow, thin area in
a large rosetted leaflet, showing a differentiated palisade, but close packing of the cells. C.—
Section through the yellow area of a much-aborted leaflet where the tissues have partially col-
lapsed. There is no well-defined palisade tissue. D.—Section of a much-aborted leaflet at the
margin, showing the close packing of the cells. E and F.—Sections through the yellow, thin
portions of a badly mottled but nearly full-sized leaflet, showing the lack of differentiation
into palisade and spongy parenchyma, almost complete obliteration of intercellular spaces, and
difference in the leaf diameters in the center ( F) and toward the margin (E) of a yellow spot.
G.—Normal leaflet, showing palisade cells well differentiated and typical loose arrangement of
the spongy parenchyma. H.—Section through the green tissue around a yellow spot of a
mottled leaflet, showing slight hypertrophy of the palisade and spongy parenchyma cells and
the reduction in size of the intercellular spaces in the spongy tissue. A to F, x 220; G and H,
420. Photomicrographs by the writer.
Bul. 1038, U. S. Dept. of Agriculture. PLATE VII.
EPIDERMIS AND SECTIONS OF PECAN LEAVES.
A.—Epidermis of a healthy pecan leaflet collected before sunrise and stained with iodin to show
the retention of starch in the guard cells. X 540. B.—Vertical section of a pecan leaflet, show-
ing a calcium oxalate crystal aggregate. X< 420. C.—Vertical section of a young pecan leaflet
collected just after emergence from the bud, showing the differentiation of the palisade tissue at
this early stage of development. Killed in Carnoy’s fluid; Fleming’s triple stain used. X 540.
D.—Horizontal section of spongy parenchyma at the margin of a yellow spot, secondary stage,
stained with iodin to show the presence of starch in the green periphery but a smaller quantity
“ total absence of starch toward the center of the yellow area. X 540. Photomicrographs by
1e writer.
Bul. 1038, U. S. Dept. of Agriculture. PLATE VIII.
VERTICAL SECTIONS OF NORMAL AND MOTTLED PECAN LEAFLETS, FROTSCHER
VARIETY.
Collected in August, 1919, at Thomasville, Ga. Killed with Carnoy’s fluid; Fleming’s triple stain
used. 540. A.—Section of normal leaflet. B.—Green portion of mottled leaflet, showing the
enlargement of the cells, the reduction in size of the intercellular spaces in the spongy parenchyma,
and the unevenness in the arrangement of the pa'isade cells. Photomicrographs by the writer.
Bul. 1038, U. S. Dept. of Agriculture.
PLATE IX.
HORIZONTAL SECTIONS OF NORMAL AND MOTTLED PECAN LEAFLETS.
A and B.—Horizontal sections through the palisade tissue illustrated in figures A and B of Plate WES
showing the enlargement of the short diameter of the palisade cells in the mottled leaflet (B). C
and D.—Horizontal sections through the spongy parenchyma illustrated in figures A and B of
Plate VIII, showing the enlargement of the cells and the reduction in size of intercellular spaces in
the green area surrounding a yellow spot in the mottled leaflet (D). All figures X 940. Photo-
micrographs by the writer.
PECAN ROSETTE. a)
cells than rounded leaves of the same plant, but the narrower form
of such leaves is due to a larger number of cell divisions with spin-
dles parallel to the long axis.
In pecan rosette the general shape of the leaf seems to follow this
rule, depending upon the orientation of the cell divisions rather than
upon differences in the size or the shape of the cells. That is, the
linear shape is not due to the development of cells elongated parallel
to the midrib, but to a difference in the number of cell divisions in
the two axes; nor are reduction in both length and breadth of leaf
caused by a decrease in the size of the cells, but rather to a decrease
in the number of cell divisions in both axes.@ On the other hand,
there is often a large and localized change in both the size and the
shape of the diseased cells, but not as related to leaf shape nor neces-
sarily to leaf diameter. In the linear type of leaf the cells are as
likely to be enlarged parallel to the short as to the long axis of the
blade, and in portions of leaves profoundly reduced in both length
and breadth the palisade and the spongy cells are at the same time
often considerably enlarged in all three dimensions.
As a result of a considerable number of measurements of the
thickness of leaves from healthy and rosetted trees of like age and
variety, striking differences were found associated with the disease.
(Table II.) Comparisons were made between leaves collected from
the north and from the south sides of trees, but no constant differ-
ences were found which could be referred to situation, since all
leaves were taken from the lower, outstanding branches, and under
these conditions those on the north received nearly or quite as much
light as those on the south periphery.
In each case the figures are based on 10 to 15 measurements of
the thickness of each of several sections from comparable parts of
each of 10 or more leaves. From such measurements it was found
that the average variation from the greatest thickness in normal,
individual leaves was 18 per cent, with extremes varying between
10 and 22 per cent. In the various types of rosetted leaves the ex-
' treme differences in thickness varied between 11 and 62 per cent
of the greatest thickness. The least variation was found in the
nonmottled linear or aborted leaves, while the greatest differences
occurred in mottled leaves. Extreme variations in thickness of
normal leaves of the Frotscher variety were 131 to 187 microns,
while in diseased leaves of the same variety the range was from 70
to 234 microns. The smallest measurements were taken at the thin
places in the leaves where tissue differentiation was lacking. The
Van Deman specimens examined had slightly thicker leaves, but the
same relations in thickness were found to hold between the healthy
and the rosetted leaves of this and several other varieties.
76289°—22—_4
22, BULLETIN 1038, U. S. DEPARTMENT OF AGRICULTURE.
TABLE II.—Variation in the thickness of normal and rosetted pecan leaves, using
the thickest part of each leaf as the standard of comparison.
Measurements of thickness.
Description of material.
s Variation
Average.| Extremes. average.
NORMAL LEAVES.
Frotscher variety, Thomasville, Ga.: Microns Microns. | Per cent.
North sidewittreess: 22 Cans Se Tees Eee ek ee ay 162 131-187 18
South sideortreesseceaiss- cae. ae eetans piieicbene coe eae eae ee 147 122-173 18
Yan Deman variety 1Cairo, Ga. 2: Seeeges basal ite. Oe 168 154-187 18
ROSETTED LEAVES.
Frotscher variety, Thomasville; Ga.:
Mottled leaves, about half size—
WNorthiside: 524 Seek Se eee se eee MELE ontles 158 108-206 38
Southside. sess eet ee ee seer eee eee riccee sen aeee cee ee 167 80-210 } 37
Much-aborted leaves, south side—
Nonmottled. 5. ° 35-25 eee ee es eee oe amano. cemabes 152 70-215 26
Mottled'\. 62250908 52 ce sry - eee ee SAE So oe 158 72-229 42
Linear leaves, south side—
Nonmottledrw 222-82) 2 et edaseh eae enemies kha dabtatcese 118 103-141 26
Mottled sti 3. 2 east seer ne renee teas tes ask east 113 70-173 50
Mottled leaves, about halfsize, northeast side.............--..--.- 139 75-234 55
Variety not stated, Cairo, Ga.:
Mottled, red-brown stage, about halfsize.-...........--.--------- 151 94-187 50
Van Deman Variety:
Mottled ‘about fll size 2st. ht eceser tet eae eet e tans sais cobain 198 140-281 45
IMGT He sineGAne. sa- 0h. -5 > Cease eee eee eeeene hn ao ane ee emer 140 | 78-195 47
The average thickness of linear leaves was always less than that
of healthy or of other types of rosetted leaves. That this difference
is not normally related to leaf size was shown by measurements
comparing the thickness of the large juvenile leaves of normal young
seedlings with that of the first type leaves above and that of large
tip leaflets with small basal leaflets on single juvenile leaves. (Table
III.) In the case of large juvenile leaves as compared with type
leaves above there was a 67 per cent variation in the area of the
laminee, but only a 7 per cent variation in thickness. In the vascular
portion of the large side veins, however, a 72 per cent variation in
the area of the cross section was correlated with this increase in leaf
(Table IV.)
size.
Tarte IlI.—Leaf thickness and vein diameter as related to size in normal
young pecan seedlings.
Thickness (microns). Leaf variation (per
cent).
Description of material.
Leaf | Leafex-| Large | Thick- nee
average. | tremes. Veins. ness. " .
Large juvenile leaves. ........--.-2-------2--+-+----- 103 | 94 to 117 515 \ 67
First type leaves above. .......-.--.--+-------+--+----- 96 | 89 to 103 388 7
Large juvenile tip leaflets.......--...---------------- 112 | 98 to 126 538 \ 10 94
Small juvenile basalleaflets on same leaves. -...--..- 101 | $8 to 112 243
Dee ee een ee ee ee ee SS SS ESE ES
PECAN ROSETTE. 23
In the large and small leaflets of single leaves a 94 per cent varia-
tion in leaf area gave only 10 per cent variation in leaf thickness;
but here again there was a 92 per cent variation in the area of the
cross section of the vascular tissue, corresponding to the 94 per cent
increase in the area of the leaf blade. “(Table IV.) It will be readily
seen that the differences in leaf thickness are at the lower range of
variation found in normal leaves. In these juvenile leaves the varia-
tion in area of the total cross section of veins was high, but not
quite as great as the difference in leaf area; the area of the vascular
part, however, increased in direct proportion with the size of the
leaf, as would be necessary to carry an adequate supply of water and
nutrients to and from the larger leaf blade. In these leaves very
little variation was to be found in the vertical diameter of either
palisade or spongy tissue.
TABLE [V.—Tissue measurements as related to size in normal young pecan
seedlings.
Thickness of Diameter of ee oe Variationin | Areaof
tissue. large side Veins. ida ee areas. cross
section,
; asa relation
Description of material. | : Cross ee
Pali- Vas- Vas- |section, petits
sada Spongy-| Total. | cular | Total.| cular | vas- Leaf. Eta
Sia part. part. | cular veins
parts. 3
Mi- Mi- Mi- Mi- | Square} Square| Per Per Per
crons. | crons. | crons. | crons. |microns.)microns.| cent. | cent. | cent.
Large juvenile leaves........ 39 41 515 351 |207,460 | 96,211 | 46
First typeleafaboveon same 72 67
lentes ee tees soe 40 30 388 187 |118,215 | 27,172 | B
Large juvenile tip leaflets. ... 35 43 538 351 |227,285 | 96, 211 | 42
Small juvenile basal leaflets 92 94
on same leaves........-....| 35 39 243 103 | 45,987 | 8,171 18
The total area of the cross section of comparable veins in healthy
leaves and in aborted leaves of approximately half size (PI. ITI,
fig. D) was nearly the same for a given variety and set of external
conditions. However, the area of the vascular portion of these veins
was reduced from 43 per cent of the total area of the normal to
about 33 per cent of the total area of the diseased veins. In the
greatly aborted leaves (Pl. III, figs. A and C) the area of the total
vein cross section was about half that in the normal leaves, while
the cross section of the vascular portion of these veins was only 10
per cent of the total vein cross section as opposed to 48 per cent in ©
the veins of normal leaves. (Table V.) While the area of total
vein cross section, except in thé most aborted leaves, tended to remain
the same as in normal leaves, the development of vascular tissue
within the vein became greatly reduced with the reduction in the
24 BULLETIN 1038, U. S. DEPARTMENT OF AGRICULTURE.
size of the leaf. In other words, the total size of vein in rosetted
leaves of reduced size tends to be as great as that normally developed
to support the full-sized leaf, but the development of vascular tissue
within the vein becomes reduced with the severity of attack and
consequent reduction in the size of the leaf blade.
TABLE V.—-Relation of vascular tissue in leaves to rosette.
Diameter oflarge | Area ofcrosssection, | Area of
side veins. large Side veins. cross sec-
RMI Di EA [Se FE a a as tine. rele-
anti : tion o
Description of material. bien woe aaseetee
a ar ascular | part to
Total part. Total. part. ‘total
veins.
ree : Square Square
Frotscher variety, Thomasville, Ga.: Microns.| Microns.| microns. microns. | Per cent.
Normal leaf, south side. ....-....------------ 347 227 94, 006 40, 107 43.
Rosetted leaf, about halfsize—
NOME SOD 20s Ao nn Sosy = cinta Sowre mini oe eee 359 202 100, 641 32, 041 32
SOT MEIG OS SSee on aoe 5 Seep Seeboaeeacas 346 202 94, 006 32, 041 34
Abarted leat south Side: ..<-2-=22---2-= s22-22-2- 259 83 52, 269 5, 280 10
Linear leaf, south side (Gearts halt 0) PSR Banner oceans 487 | 180 85, 472 25, 442 30
An examination of the vein islets of healthy and diseased leaves:
(secondary stage) has revealed striking differences in size, shape,
and arrangement. Over the entire normal leaf blade these tiny areas
bounded by the small, anastomosing veinlets tend to be isodiametric
and of uniform size (Pl. X, fig. E). In the yellow areas, on the
contrary, great differences in size and shape are the rule (Pl. X, figs.
AtoD). At the center of these spots the vein islets are smallest and
become larger and larger with increasing distance from the center
until often in the neighboring green parts they are considerably
larger than in the healthy leaf. Their appearance suggests an in-
hibitory influence generated from the center, which largely prevents
normal growth an differentiation there, but acts as a poison more
and more feebly with receding distance avi the focal center until in
the neighboring green parts it has become sufficiently attenuated to
function as a stimulant rather than as an inhibitory factor. This
theory is also borne out by the writer’s histological studies.
Not only are the vein islets highly variable in size, but they are
often greatly distorted in shape. In many cases they are linear in
outline, and with reference to the spot they approximate the arrange-
ment ai spokes in a wheel (PI. I, fig. 1; Pl. X, fig. B). syn it
will be seen that the direction of their greatest expansion may or
may not parallel the direction of greatest expansion in the leaf blade
as a whole. That is, the size, shape, and arrangement of the vein
islets in these chlorotic spots of the secondary stage are controlled
from the focal center of the spot rather than by the normal mor-
phogenic forces of the leaf.
Bul. 1038, U. S. Dept. of Agriculture. PLATE X.
VEIN ISLETS OF UNSECTIONED AND UNSTAINED PECAN LEAVES.
A, and C to E are from photomicrographs of leaves preserved in alcohol. B is from a photomicro-
graph of a dry herbarium specimen. All at the same magnification. X about 60. The white
spots are calcium-oxalate crystal aggregates. A to D.—Yellow spots of rosetted leaflets in the
secondary’stage, showing the distortion in shape and the variation in size of the vein islets. E.—
Similar view of vein islets in a normal leaflet, showing approximately uniform size and regular
arrangement. Photomicrographs by the writer.
Bul. 1038, U. S. Dept. of Agriculture. PLATE XI.
VERTICAL SECTIONS OF HEALTHY AND DISEASED PECAN LEAFLETS,
FROTSCHER VARIETY.
Collected on August 25 and 26, 1919, at Thomasville, Ga. All figures at the same magnification.
% 540. A.—Healthy pecan leaflet collected at sundown, killed in Carnoy’s fluid, and stained
with iodin to show the presence of starch in the palisade and spongy cells. B.—Healthy
pecan leaflet collected in the morning, killed in Carnoy’s fluid, and stained with iodin to show
the absence of starch. C and D.—Pecan leaflets aborted with rosette, collected and treated
in the same way, respectively, as in the preceding two normal leaflets. Chloroplasts in both
night (C) and morning (D) specimens were gorged with starch, and apparently no trans-
location had taken place during the night. Note also the enlargement of the cells in the dis-
eased material. Photomicrographs by the writer.
Bul. 1038, U. S. Dept. of Agriculture. PLATE XII.
FRESH LIVING CELLS OF PECAN LEAVES.
Camera-lucida drawings of freehand vertical sections, mounted in water. All at the same mag-
nification (x about 1,200) and oriented the same as in the leaf. A.—Healthy palisade cell,
showing well-defined nucleus and plump livid-green chloroplasts. B.—A disorganization stage
of apalisade cell in a leaf affected with rosette; chloroplasts disintegrated and nuclear outline
vague. C.—Healthy cell of the spongy parenchyma. D.—Slightly diseased cell of the spongy
parenchyma in which the chloroplasts have lost a part of their green color. E.—Spongy paren-
chyma cell at the margin of a yellow area. The protoplasmic structures on the side toward the
spot (right) are disorganized and the nucleus is fragmenting. Chloroplasts next to the green
periphery of the spot (left) are still green and unfragmented, though a part of them are smaller
than normal. F and G.—Spongy cells at further stages in disorganization. H.—Spongy cell
showing entire disorganization of contents. I—Tannin degeneration products gathering into
flocules at a later stage of the disease in a spongy cell. J.—Spongy cell at the red-brown stage
with more or less homogeneous reddish brown contents plasmolized. K.—Spongy cell at the
margin of a yellow area (right) showing chloroplasts red-brown, but unfragmented on the side
toward the spot. This form of injury is rather uncommon. Drawings by the writer.
PECAN ROSETTE. 25
A comparison of the size and shape of the vein islets in large and
small mature leaflets on the pecan leaves and in large (mature)
juvenile (undivided) and type leaves on the same normal seedlings
showed very little variation. Ensign (32) found in healthy citrus
leaves that the shape varied in different parts of the leaf, but the size
was independent of the shape or location on the leaf. The size and
shape of the vein islets in citrus were approximately the same in
normal leaves and in chlorotic leaves of plants dwarfed from starva-
tion. A comparison of the vein islets in large and small leaves on the
same plant showed that the size was constant irrespective of the size
of the leaf. A comparison of the leaves of all ages on the same plant
- showed that the youngest leaf had the smallest vein islets and with in-
creasing maturity a corresponding increase in the size of the islets
took place. Ensign concluded that the main skeleton of the vascular
system is laid down very early in the history of the leaf and that but
little differentiation later takes place. It is clear that in pecan rosette
the’size, shape, and arrangement of the vein islets are considerably
altered from the normal. In fact, there is in the mottling of the
leaves in pecan rosette much that appears comparable in origin to
Liesegang’s diffusion patterns, for example, those obtained with
drops of silver-nitrate solution on a layer of solidified gelatin in
which potassium bichromate had been dissolved. These “ Liesegang
phenomena ” have also been compared by Kiister (48) to the formation
of growth rings by Armillaria, Penicillium, and other fungi, to the
alternative lighter and darker areas of some leaf-spot diseases of
fungous or bacterial origin, and to a great variety of pattern phe-
nomena in both plant and animal kingdoms. The end results in all
these cases of crganic origin are similiar in appearance to those:
obtained by diffusion experiments carried out in vitro. Furthermore,
while great caution should be exercised in the interpretation of such
experiments the results are extremely suggestive that diffusion in
colloidal systems plays a prominent role in the development of the
pattern phenomena in both health and disease.
Profound derangements take place in starch assimilation and trans-
location. In healthy leaves collected at sundown the starch content
of the chloroplasts is uniform for each tissue with usually the
greatest accumulation in the palisade cells (Pl. XI, fig. A). Stained
with iodin the starch is seen in a more or less irregular mass at the
center of the chloroplasts but in no case completely filling the plastid.
Starch also occurs in the guard cells and in occasional plastids in the
bundle sheaths. Healthy leaves collected before sunrise the following
morning from similar situations on the same trees in general,showed
no starch at all (Pl. XI, fig. B). In these healthy leaves the iodin
gave merely a yellow stain to the cell contents. An extremely rare
26 BULLETIN 1038, U. S. DEPARTMENT OF AGRICULTURE.
palisade or spongy cell and many of the guard cells were, however,
filled with starch as in leaves collected at sundown (PI. VII, fig. A).
Mottled leaves of all sizes collected at night showed the green por-
tions gorged with starch (Pl. XI, fig. C). The plastids of the pali-
sade and spongy cells were not only filled to the periphery but swollen
as if almost bursting with their accumulation of starch. Toward
the center of the yellow spots (secondary stage) the chlorophyll
bodies are of less and less frequent occurrence until at the center only
an occasional starch-filled plastid is to be found (PI. VII, fig. D).
In these areas where the effects of the disease have been in operation
from the early stages of bud growth the inhibitory influence has
affected not only tissue differentiation but has also prevented normal
development of the plastids.
Mottled leaves collected before sunrise the following morning ap-
peared the same as those collected at night. (Pl. XI, fig. D). As far
as could be determined from the iodin stain, no eratslbentiod at all
had taken place during the night. Palisade cells, spongy cells, and
guard cells of the green areas were full of starch, and the occasional
plastids in the bundle sheaths of the veins were also gorged.
The nonmottled, but greatly aborted leaves, showed this gorging
of the plastids over the whole lamina. The starch sheaths around
the midveins were also black with starch. As in the other diseased
leaves the plastids appeared as full of starch in the early morning as
at the end of a sunny day.
Wherever plastids are present in either stage of leaf mottling the
first sign of disease in these bodies consists in a gradual loss of the
green chlorophyll. In the healthy living cell of both palisade and
‘spongy parenchyma the plastids are plump and of a livid green color,
while the nucleus is plainly visible as a grayish, more or less cen-
trally located body with definite outline (Pl. XI, figs. A and C).
As the disease progresses these chlorophyll bodies first lose their
green color (PI. XII, fig. D), then both nucleus and plastids begin to
break down (Pl. XII, figs. B and E to H), first losing their definite-
ness of outline, or becoming fragmented or appearing as if eaten away
at the periphery. Later all the visible remains of the cell structures
consist of globules, probably fatty, and darker-colored granules of
various shapes and sizes irregularly distributed throughout the cell,
with all appearance of disorganization. With ferrous salts many of
the brownish granules gave the reaction for tannin. In cases of
severe attack the entire tissue within these yellow areas later becomes
reddish brown and collapses (Pl. I, fig. 3). In reaching this end
stage the tannin degeneration products here described gather into
larger and larger floccules (Pl. XII, fig. 1) until finally the cell may
be filled with a more or less homogeneous reddish brown matrix,
PECAN ROSETTE. 27
which later recedes from the cell wall (Pl. XII, fig. J), and at last
the whole cell collapses and shrivels.
The deposition of crystal aggregates of calcium oxalate is char-
acteristic of pecan leaves (PI. VII, fig. B). These crystals begin
to form in giant binucleate cells of the palisade just after the young
leaf emerges from the bud. Finally the protoplasmic contents dis-
appear, and the crystal aggregate nearly fills the cell. These crystal
ageregates are distributed with considerable regularity in the healthy
leaf and in the majority of cases one to each vein islet (PI. X, fig. E).
In the yellow spots of the secondary stage of the disease, on the other
hand, their formation and distribution vary widely. At the focal
centers in severe cases few or no crystals at ail are formed, whereas
in the surrounding green parts they are often far more numerous
and sometimes larger than in the comparable healthy leaf.
Averaging a large number of counts in cross sections of green
parts of diseased and normal leaves of the Frotscher variety col-
lected at Thomasville, Ga., it was found that in the normal leaves
20 crystal aggregates occurred to every 100 vein islets observed in
section, while in the diseased leaves 60 crystal aggregates were found
to every 100 vein islets. In Van Deman leaves collected at Baconton,
Ga., there were 16 crystal aggregates to every 100 vein islets in the
healthy leaves as compared with 54 in the diseased leaves. Further-
more, after all due allowance for differences in cortical area of the
diseased and healthy leaves compared, a much larger number of
these crystals was found in the cortex of petioles and midveins in
rosetted leaves. In view of the fact that waste organic acids usually
accompany carbohydrate formation (57, p. 173) these results are
significant. Since growth at the periphery of the yellow spots
in the secondary stage is often abnormally active, as is shown by
the size of the cells, an unusually large accumulation of such or-
ganic acids would naturally be expected to take place in that region.
Conversely, with the reduction of chlorophyll formation and assimi-
lation in the centers of the spots a smaller production of such acids
would occur.
Development of the shield-shaped resin glands occurring mostly
on the lower surface of the leaves is also profoundly affected by the
disease. On the normal leaf these glands are rather regularly and
sparsely distributed over the surface of the blade and contiguous
to the veins and veinlets. In diseased leaves of the secondary stage,
on the contrary, the focal centers of the yellow spots are often
_ thickly covered with these resin glands both contiguous to the vein-
lets and well within the vein islets themselves. The more severe
the general effects of the disease the more numerous were these glands
found to be, until in places where the tissues were practically de-
28 BULLETIN 1038, U. S. DEPARTMENT OF AGRICULTURE.
funct or bordering wounds where such tissues had fallen out, they
presented a continuous, rough layer of glandular shields. Fixam inte
tion of ordinary leaf wounds such as those made by insects showed
no such abnormal development of resin glands. These conditions
have been frequently observed in the general examination of leaves
collected during several seasons and in different varieties and locaii-
ties. Furthermore, using a simple binocular microscope, observa-
tions have been checked up by exact counts. For example, counts
were made of the numbers occurring in a single field in different
parts of 10 normal leaves of the Frotscher variety collected at
Thomasville, Ga. (1916), and in the yellow areas of 10 comparable
rosetted leaves. The average for normal leaves was 8 to a field,
while in the diseased leaves there were 92 to a field. In a lot of the
Schley variety collected at Orangeburg, S. C. (1920), the average for
healthy leaves: was 43 and that for comparable yellow areas of
diseased leaves 212 to a field. Similar results were found in material
collected at Belleview, Fla., and at Baconton and Cairo, Ga. In some
older spots of the secondary stage these resin glands were so closely
packed together that counting was impossible.
This increased development of resin glands is characteristic of
many halophytes and xerophytes and in these cases apparently bears
some relation to the condition of physiological dryness. Further-
more, Tschirsch (80) has demonstrated that the secretion of resin
is produced within the cell wall itself just below the cuticle. This
region he calls the “resinogenous layer.” Nutrient substances and
water pass out of the protoplast into this resinogenous layer, to be
there further molded into the final product, resin. This process,
then, is participated in by both protoplast and cell wall and necessi-
tates loss of material from both and a final breaking down of the
cell wall itself. In pecan rosette this abnormal development of resin
glands is then probably to be connected in some way with the gradual
and general degeneration of the cells involved.
In order to determine whether new spots may be formed after the
full expansion of the leaf and whether yellow spots already formed
may increase in size, resort was made to careful field observation and
microscopical study of fresh living material. On July 20, 1920, at
Orangeburg, S. C., the outlines of several hundred spots on 56 dif-
ferent leaves of both primary and secondary stages were carefully
traced with India ink on the upper surface of the blade. At the
same time the outline of one leaflet on each leaf was traced on paper
for comparison with its size at the end of the observational period.
Furthermore, nonmottled areas of diseased leaves were marked in a
similar way; and finally also certain areas of normal leaves were so
marked as a check on possible injury by the India ink. These leaves
were located on one normal and five rosetted trees in a 45-acre
PECAN ROSETTE. 29
pecan orchard of the Schley variety at Orangeburg, S. C. Final
notes were taken on a portion of these leaves at the end of 10 days,
while the remainder were left until September 9, 51 days later.
In leaves of the primary stage, after 10 days, 17 yellow spots had
developed within 170 previously marked green areas. Of 130 yellow
spots already present, 11 in all had increased in size but only 2 con-
spicuously so. In slightly affected leaves of the secondary stage, 36
green areas had developed 13 yellow spots; and out of 26 spots already
present 19 had increased in size, 15 of them decidedly. In the consid-
erably aborted leaves, 109 green areas had developed 137 new spots;
and out of 270 spots present at the start 193 had increased in size, 70
of them by at least 50 per cent of their original diameter. These
leaves were all fully expanded at the beginning, since no appreciable
increase in size of blade could be discerned on comparing the leaflets
with their original tracings. The check leaves remained normal and
evidenced no ill effects from the presence of the India ink.
Four uniformly pale leaves recently out of the bud and not fully
expanded at the first observation, at the end of 10 days, had de-
veloped a conspicuous green color around the margins and along the
veins, but had failed to develop further chlorophyll in the centers of
the areas between the lateral veins. These leaves were now typical
of the secondary mottled stage.
In those leaves left for 51 days, 33 green areas in the primary stage
leaves had developed only 2 yellow spots. Of 180 spots already pres-
ent, 86 had increased in size, but only 15 decidedly so. In aborted
leaves of the secondary stage, 50 green areas had developed 17 spots
and of 66 spots present at the beginning 50 had increased in size, and
37 of them decidedly so.
Most of the leaves of the primary stage were at the beginning older
and fully matured. The leaves of the secondard stage were fully ex-
panded, but for the most part soft and immature. It is thus apparent
that these yellow spots are not necessarily laid down in the bud stage
of the leaf or in a definite and unchangeable pattern, as may be the
case in certain heritable variegations. It is clear also that spots may
develop de nevo even after expansion of the leaf blade and that those
already formed may increase in size, though less rapidly and fre-
quently after the leaf has fully matured.
That yellow spots may increase in size was also shown by a study
of free-hand sections of fresh, living material. Here on the margins
of the spots an occasional cell was found which showed disorganiza-
tion of its internal structure on the side bordering the yellow area,
while that part of the cell contents toward the green periphery ap-
proximated the normal. (PI. XII, figs. E and K). Here again ap-
pearances suggest the outward diffusion of some toxin from a focal
center of its production.
30 BULLETIN 1038, U. S. DEPARTMENT OF AGRICULTURE.
These changes in mature cells affect the protoplasmic structures
rather than the cell size, shape, or other so-called histological char-
acters. They consist in such changes as loss of chlorophyll, breaking
up of the nucleus, and degeneration of the cytoplasm. The situation
seems to be that the toxic substances which in the embryonic stages
may produce the profound alterations in morphogenetic processes of
leaf formation described above, in the mature leaf cells are simply
destructive of the still plastic protoplasm. What the ultimate ex-
pression of the disease will be depends, then, upon the ontogenetic
stage of the plant organ at which the cause becomes effective.
SUBSIDIARY EXPERIMENTS.
In order again to test the effect of subjecting pecan trees to vary-
ing environmental conditions, further transplanting and fertilizer
experiments were conducted on a small scale at the New York
Botanical Garden. .
In one of these experiments 32 large nursery trees showing severe
attack of rosette during the summer of 1913 at Monticello, Fla.,
were transplanted late in the fall to large pots of garden soil in one
of the botanical garden greenhouses. On account of the length of
the tap roots these trees were so severely root pruned that only a
part of them survived. However, the 18 remaining trees were under
observation for two seasons, and at no time during this period did
any sign of rosette develop although before transplanting all were
badly mottled and dying back from the tip. |
In another test several pecan nuts were germinated in fine quartz
sand in each of 32 glazed crocks and in 4 similar crocks of garden
soil. All were uniformly watered by the porous clay cup autoirriga-
tion method. The crocks containing sand were divided into 8 lots
receiving the following different fertilizer treatments:
(1) Liberal application of a commercial fertilizer containing muriate of
potash, ammonium sulphate, and acid phosphate.
(2) Liberal application of a fertilizer made up of sulphate of potash, slaugh-
ter-house tankage, and Thomas slag.
(3) An excess of stable manure.
(4), (5), and (6) An excess, respectively, of slacked lime, magnesium sul-
phate, and ferrous sulphate.
(7) Equal quantities of slaked lime and magnesium sulphate.
(8) Control, untreated.
Lot 9 consisted of the 4 crecks with garden soil alone. All seed-
lings germinated normally and were under observation during one
season. All those in lots 1, 2, 3, and 9 made good growth and ap-
peared dark green and healthy throughout the experiment. Seed-
lings in the other lots started out well but after using up the nutrients
in the seeds they became stunted and finally developed a general
PECAN ROSETTE, 31
yellowing of the foliage. However, none of the mottling or other
signs of rosette appeared at any time during the season.
Two large, healthy seedling pecans were transplanted to a plat
of garden soil containing a great excess of lime where mortar had
previously been mixed for building purposes. These trees have
been under observation for three seasons, and no signs of any type
of chlorosis have at any time become evident.
In another small experiment run for two months, 10 seedlings
potted in garden soil were given liberal and fairly uniform appli-
cations of water throughout the period, while 10 similar potted seed-
lings were given only sufficient water to prevent wilting. In the
latter case but little growth was made, and a part of the leaves
developed a general chlorosis. At no time, however, were any signs
of rosette to be seen.
PROBABLE NATURE OF PECAN ROSETTE.
In the writer’s opinion the pathogenic picture of pecan rosette as
shown in the preceding pages, including histological and cytological
features, is much more in agreement with the infectious type of
chloroses, including the yellows and mosaic groups, than with those
chloroses known to be caused directly by soil or climatic conditions.
It is not considered, however, that adequate proof has yet been given
on either side, and the present study is offered merely as one more
step in the study of this baffling group of diseases and as a sugges-
tion for further research.
In the environmental type of chlorosis structural changes may
occur. The xerophytic tendency is toward a bilateral palisade with
restricted air spaces reducing transpiration. Hydrophytic condi-
tions produce large, globose cells and loose arrangement of tissues.
Excess of sodium chlorid develops enlarged, rounded, spongy, and
massive palisade cells and causes reduction in the number of chloro-
plastids. Wax coatings are also characteristic of many salt-loving
plants. Lack of sufficient water or soil nutrients results in the
dwarfing and hardening of the tissues, often followed by general
chlorosis but without morphogenic changes. The dimorphism of
certain plants when grown under differing environmental conditions
is well known, but is in no sense comparable to a diseased or other
deranged condition. Changes in metabolism also may be caused
directly by soil or climatic conditions. All these changes, however,
in the main, are general changes affecting the plant tissues as a
whole or tending to affect all similar tissues of the plant alike.
In pecan rosette profound alterations and derangements in metab-
olism, anatomy, and morphogenesis occur together in great com-
plexity. Different types of tissue derangement are found in the same
Jeaf. Reduction and increase in the number of cell divisions, tissue
32 BULLETIN 1038, U. S. DEPARTMENT OF AGRICULTURE.
differentiation and lack of tissue differentiation, hypotrophy and
hypertrophy of cells may all occur together within an area only a
few millimeters in diameter. There seem to be focal centers out from
which these alterations spread, and the resulting abnormal develop-
ment appears to be controlled from these centers rather than by the
normal morphogenic forces. The factors controlling morphogenesis
as related to cell differentiation, to development of tissues in dis-
eased areas, and to general pattern of leaf are profoundly altered.
No less far-reaching are the derangements in metabolism evidenced
by the altered assimilation and translocation of starch.
Pecan rosette frequently appears first on the tip leaves of a single
branch and seems as likely to affect a part of the tree first as to occur
at once over the whole top.
Careful field observations have given no evidence of varietal dif-
ferences in susceptibility or resistance to pecan rosette. This same
statement would apply equally well to most infectious chloroses.
Fertilizer and transplanting experiments and field observations
indicate that rosette is affected, at least indirectly, by soil and cli-
matic conditions, but similar relations are exhibited by chloroses of
known infectious nature as well as by many diseases of bacterial or
fungous origin. Furthermore, great reliance is not to be placed upon
the results of plat soil experiments unless such results are unusually
definite, or unless in a large number of similar tests the data all
point in the same direction.
In view of the refractory attitude exhibited toward grafting and
cross-inoculating experiments by certain of the infectious chloroses it
is considered that this type of experimentation has not yet given a
conclusive answer to the question of possible infectivity in pecan
rosette. In the early experiments a small portion of the diseased
buds developed the disease, though in no larger percentage than in
contiguous nursery trees worked with supposedly healthy buds in
the ordinary commercial propagation. These experiments need repe-
tition on a large scale under controlled conditions and with extreme
care in selection both of diseased and healthy buds and stocks. More-
over, the possibility of insect transmission has not been touched upon
from the standpoint of experiment.
The disease has not been definitely and experimentally caused by
a set of known conditions. Though it is more prevalent and severe
under certain environmental conditions, it occurs to some extent in
practically every soil type where the tree has been observed, with
the possible exception of swamp land: Under these conditions the
tree makes very little growth and presents a starved and stunted
appearance followed by a general form of chlorosis bearing no
resemblance to rosette. Ultimate proof of the cause must account
for all cases of the disease.
PECAN ROSETTE. 33
It is difficult to explain on the soil hypothesis why only a part of
a tree may be diseased and why when two trees of the same age stand
within a few inches or a few feet of each other the one may remain
perfectly normal and vigorous while the other is stunted and dying
back with rosette. The recovery of diseased trees when transplanted
in the north on the other hand is also difficult to explain, but no
more so than the apparent recovery of potatoes from mosaic when
carried from Maine to Colorado or tobacco mosaic under certain rela-
tions of light or temperature. As previously mentioned, Baur has
clearly demonstrated that the contagium of abutilon mosaic is readily
killed by subjection to certain environmental conditions such as the
withdrawal of light. He found that cutting out the yellow areas
as they developed also finally brought recovery. Here, as in many
parasitic diseases both of plants and of animals, though the conta-
glum may be carried to remote parts of the body, it is only in certain
definite locations and under certain definite conditions that it can
reproduce itself and initiate lesions in the host.
If pecan rosette is due to some chemical compound brought in from
the soil in harmful quantities one would expect the tissues along the
veins to be first and most profoundly: affected and that the result
would be evident over the whole tree at about the same time. Fur-
thermore, if the yellow mottling is interpreted as due not to a cause
operating from the focal centers of the spots but to a lack of suffi-
cient soil nutrients or water brought in by the veins, how are to be
explained the lack of chlorosis along the leaf margins and the abnor-
mally increased growth at the periphery of the spots?
Tt is true that the local application of purely physical or chemical
~ stimuli may locally cause cells to enlarge or proliferate, and their
application in lethal quantities may result in injury and final death
without the intervention of any parasitic organism, as witness, for
example, the more recent experiments of Dr. Erwin F. Smith (74)
in the production of plant overgrowths without the intervention of
any parasite. However, as in any cytological or embryological study,
it is not the single section taken by itself that tells the story, but the
sequence of one following the other, the whole series fitting together
in an orderly manner to build up the complete picture, so in the
chloroses of plants it is not the mere fact of chlorophyll disintegra-
tion that will show the type of disturbance present, or that will
eventually lead to the determination of the cause in any particular
case,-but the whole series of events and appearances concerned: in the
production and manifestation of the derangement.
It is probable that all effects of parasites upon their hosts, when
reduced to their ultimate reactions, may be explained in terms of
physics and chemistry. It is in the regulation of these physical
34 BULLETIN 1038, U. S. DEPARTMENT OF AGRICULTURE.
and chemical reactions and stimuli, and in their application in a
particular manner and at a particular time and place, that the com-
plexity of the final manifestations of infectious diseases differs from
the more simply induced changes in metabolism and structure. It
is this regulatory effect and this extreme complexity of reaction
which make many infectious diseases so difficult to induce by arti-
fi¢ial means.
If causes may in any measure be judged from their effects, the
histological and cytological evidence points to the cause of pecan
rosette as being similar in its general nature to the ultimate causes.
of the infectious chloroses. Whether in this particular disease the
factors responsible for alterations in the normal structure and metab-
olism must be introduced into the plant from without, or whether
they originate within the plant itself, is a question yet to be answered;
but whatever the ultimate solution of the problem may be, the cause
will undoubtedly not be found in any simple soil or water relation.
So far as worked out, the infectious chloroses in general exhibit,
like pecan rosette, a simultaneous and deep-seated derangement both
in morphogenesis and metabolic processes. Structural derangements
in diseased leaves may show ‘abrupt local change. That is, in many
cases, entirely different types of tissue derangement, such as reduc-
tion in size of cells and number of cell divisions or enlargement of
cells and increased number of cell divisions, may occur side by side,
not only in the same leaf but even in the same part of the leaf.
Along with these structural changes occur fundamental functional
derangements concerned with the assimilation and translocation of
carbohydrates and with nitrogen metabolism. As is well stated by
True and Hawkins (79) in considering spinach blight:
It would seem to be indicated that the cause of carbohydrate accumulation
should be sought in the deeper lying metabolic processes in connection with
which carbohydrates are utilized. . . . Accumulation is due not to a breakdown
of digestion but to some partial failure in the subsequent metabolic processes
in connection with which carbohydrates are used.
Moreover, not only restriction in chlorophyll development occurs,
but often abnormal and irregular groupings and a reduction in size
and number of chloroplasts. In the later stages disintegration of
the plastids and other cell contents often follows, and finally a
shriveling of the entire cell ensues.
It is typical of the group of infectious chloroses that only meriste-
matic tissues are morphogenetically affected. There is always a
rather definite incubation period for each disease, and external signs
of the disease gradually progress from the point of entry of the
contagium. In some instances the contagium appears not to reach
all parts of the plant, as in the case of potato plants secondarily
infected with mosaic or leaf-roll where often only a part of the
PECAN ROSETTE. 35.
tubers give rise to diseased progeny. Furthermore, the fact that
many of these diseases are transmissible by insects shows them to be
entirely different in nature from those chloroses due to soil or cli-
matic conditions. In some of these diseases where the insect rela-
tions have been most carefully worked out it is indicated, that a
definite incubation period must also elapse after feeding upon a
diseased plant before an insect becomes infective.
Except in the most general way, the infectious chloroses exhibit no
pattern as related to the leaf. The factors controlling morpho-
genesis of the leaf in many-cases seem to have run riot. In the
mosaic type the spots, irregular in themselves, are also irregularly
distributed over the surface of the leaf. In many cases, it is true,
they avoid the larger veins, but in other instances they are distributed
irrespective of venation.
There is nothing to show a difference in kind between those mosaics:
transmissible by expressed plant juices and those transmitted only
by grafting, such as the infectious mosaic of Abutilon. The causal
contagium in the latter case may be compared to a more obligate
parasite which is greatly restricted in the conditions necessary to its:
life activities and reproduction.
The fact that later investigation has shown some of the filterable
contagium diseases to be due to microorganisms is at least presump-
tive evidence in favor of the organism theory of infectious chloroses.
Furthermore, Allard has shown that with a fine enough clay cup it
is possible entirely to filter out the infective principle from the
expressed juices of tobacco plants affected with mosaic.
The crucial difference, however, lies in the power of self-reproduc-
tion possessed by the contagia of all the infectious chloroses. To
give a concrete example, starting with a mosaic-diseased tobacco
plant and in each case using a drop of the expressed plant juice di-
luted 1 to 1,000 in water,.it 1s possible to cause the disease in an in-
definite series of plants successively inoculated one from the other at
proper intervals, and the juice from the last of the series will possess
as high a power of infection as that from the first. In a case like
this it might be mathematically shown that if reproduction had not
taken place a drop from the last plant of the series must contain less
than one molecule of the originally injected material. No such power
of self-reproduction hag ever been demonstrated for a definite chemical
compound. The true enzyms are formed by living organisms in re-
sponse to certain stimuli and have never been shown to be capable
of self-reproduction.
Assuming the contagia of infectious chloroses to be nonliving sub-
stances, their effects may be compared up to a certain point with
various phenomena of bacterial toxins and immunization. However,
the fact of reproduction in these contagia is in no way elucidated by
36 BULLETIN 1038, U. S. DEPARTMENT OF AGRICULTURE.
any such comparison. Diphtheria toxin, for instance, if injected into
a susceptible host may cause all essential signs of the disease, but it
takes living bacteria to produce more of the toxin so that the disease
may be transmitted down the line through a successive series of in-
dividuals.
On the other hand, the “Contagium vivum fiuidum” theory of
Beijerinck (17) in some modification is at least worthy of serious
consideration as a working hypothesis. It is not impossible that de-
composition processes may be propagated by methods entirely unlike
reproduction by division as known in living organisms and so as to
imitate self-reproduction. If some such course of events could be
demonstrated it would perhaps explain the known facts, including
the gradual progress of the disease from the point of entry, equally
as well as the organism theory. However, though it is well to keep
an open mind on all questions still in the realm of theory, no such
course of events is yet known to chemistry. On the other hand, most
of the known facts concerning the so-called filterable virus diseases,
so it seems to the writer, conform to the known results of invasion by
parasitic organisms.
SUMMARY.
As a class, the chloroses due to soil or atmospheric conditions are
rather general effects which are more or less comparable to starva-
tion, overfeeding, or direct poisoning. There are not the profound
or strictly localized derangements in both metabolic and anatomical
development, and it is a question in regard to many of these phe-
nomena whether, in the restricted sense, they should be regarded as
diseases at all.
In the specific chlorotic diseases of an infectious nature funda-
mental derangements in both physiological and structural develop-
ment are simultaneously brought about. Although all effects of
parasite upon host when reduced to their ultimate reactions are
probably to be explained in terms of physics and chemistry, it is
in the regulatory effect and in the extreme complexity that these
and many other infectious diseases SHES from the direct effects of
nonliving substances.
The histological and cytological bain suggests that pecan
rosette in its specific sequence of signs and in the complexity of
the structural and physiological derangements bears far more simi-
larity to the known infectious chloroses than to those caused
by soil or climatic conditions. Whether in this particular disease
the factors responsible for alterations in the normal structure and
metabolism must be introduced into the plant from without, or
whether they originate within the plant itself, is a question yet to
be answered; but whatever the ultimate solution of the problem the
cause will undoubtedly not be found in any simple soil or water
relation.
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(54) Miter, H. K. ;
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(57) PALLADIN, VLADIMIR L.
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sian ed. (1914), edited by Burton Edward Livingston. xxv,
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(58) PENHALLOow, D. P.
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(59) PETHYBRIDGE, GEORGE H.
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PECAN ROSETTE. 41
(62) QUANJER, H. M., and others.
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Litertuur, p. 67-70. Summary [in English], p. 71-74.
REpDpICK, DONALD, and STEWART, VERN B.
1918.
Varieties of beans susceptible to mosaic. In Phytopathology,
v. 8, no. 10, p. 530-534.
1919. Transmission of the virus of bean mosaic in seed and observa-
tions on thermal death-point of seed and virus. Jn Phyto-
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1921.
1919.
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Leaf-roll, net-necrosis, and spindling-sprout, of the Irish potato.
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VITA.
FREDERICK VERNON RAND:
Born at Barnet, Caledonia County, Vermont, March 16, 1883.
Early /education in the public schools of Cummington, Brimfield,
Pelham and Norwich, Massachusetts. Graduate of Franklin Academy,
Malone, New York, in 1904. Recipient of the degrees of Bachelor
of Science, 1908, and Master of Science, 1911, both from the Uni-
versity of Vermont. Graduate study, Johns Hopkins University,
1912-1913.
Elected to the Green Mountain Chapters of Phi Beta Kappa and
Alpha Zeta in 1908. Graduated from the University of Vermont with
“cum laude” and “Honorable mention tor thesis of conspicuous merit.”
Fellow in Botany, Columbia University, 1913-1914. Fellow, American
Association for the Advancement of Science, 1920. Charter Member,
American Phytopathological Society. Member of Botanical Society
of America and Botanical Society of Washington.
Undergraduate Instructor in Botany, University of Vermont, two
years, 1906-1908. Student Assistant in Botany, Vermont Agricultural
Experiment Station, summer of 1907. Scientific Assistant in Plant
Pathology, U. S. Dept. of Agriculture, June, 1908, to Sept., 1913.
Collaborator and Scientific Assistant, U. S. Dept. of Agriculture and
Assistant in Botany, Columbia University, Sept., 1913, to May, 1914.
Scientific Assistant, May to June, 1914; Assistant Pathologist, July,
1914, to June, 1920; Pathologist, July, 1920, to date, U. S. Dept. of
Agriculture.
Author of the following publications :
The shrubs and woody vines of Vermont. Vt. Bot. Club Bull. 3,
Apr., 1908.
Direct color photography. Vt. Cynic, Burlington, Apr. 22, 1908.
Vermont shrubs and woody vines. Joint author with L. R. Jones.
Vt. Agric. Exp. Sta. Bull. 145, 1909.
The botanical work of the National Department of Agriculture
Bull. Vt. Bot. Club, 1910.
A pecan leaf blotch. Phytopathology 1:133-138, 3 figs., 1911.
Further studies of pecan “rust.” Science 35:1004, 1912.
The practical in science. The Student, Malone, N. Y., Feb., 1913.
Some diseases of pecans. Jour. of Agric. Research 1 :303-337, 8
figs., pls. 33-37 (1 col.), 1914. é.
Pecan rosette. Joint author with W. A. Orton. Jour. of Agric.
Research III :149-174, 1 fig., pls. 24-28, 1914.
Dissemination of bacterial wilt.of cucurbits. Jour. of Agric. Re-
search V :257-260, pl. 24, 1915.
Transmission and control of bacterial wilt of cucurbits. Joint
author with Ella M. A. Enlows. Jour. of Agric. Research VI:417-
434, pls. 53-54, 1916.
Leaf spot-rot of pondlilies caused by Helicosporium nymphaearum,
n. sp. Jour. of Agric. Research VIIT :219-232, pls. 67-70, 1917.
A competence in water-lilies. The Rural New Yorker 78 :1637-1639,
2 figs., 1919.
Some insect relations of Bacillus tracheiphilus EFS. Joint author
with Lillian C. Cash. Phytopathology X :133-140, 1 fig., 1920.
Bacterial wilt of cucurbits. Joint author with Ella M. A. Enlows.
U. S. Dept. of Agric. Bull. 828 :1-43, 2 tab., 10 figs., pls. 1-4, 1920.
A coordination of our knowledge of insect transmission in plant and
animal diseases. Joint author with W. Dwight Pierce. Phytopath-
ology X :1-43, 1920.
A lotus leaf-spot caused by Alternaria nelumbii sp. nov. Joint
author with Ella M. A. Enlows. _ Phytopathology XI:135-140, 1 fig.
and 1 pl., 1921.
Stewart’s disease of corn. Joint author with Lillian C. Cash. Jour.
of Agric. Research XXI:263-264, 1921.
Insect dissemination ot plant diseases from the viewpoint of past
endeavor. Phytopathology X11, 1922.
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