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655 "3 HE HARD ROT DISEASE OF GLADIOLUS

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A - THESIS

PRESENTED TO THE FACULTY OF THE GRADUATE SCHOOL

OF CORNELL UNIVERSITY FOR THE DEGREE OF

DOCTOR OF PHILOSOPHY

BY

LOUIS MELVILLE MASSEY

Published as Cornell University Agricultural Experiment Station Bulletin 380,
September 1916 ,

Byrajin’s

Sean i
Beene

THE HARD ROT DISEASE OF GLADIOLUS

A THESIS

PRESENTED TO THE FACULTY OF THE GRADUATE SCHOOL

OF CORNELL UNIVERSITY FOR THE DEGREE OF

DOCTOR OF PHILOSOPHY

BY

LOUIS MELVILLE MASSEY

Published as Cornell University Agricultural Experiment Station Bulletin 380,
September 1916

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CONTENTS

PAGE
ERPS EN LER TLC SA veer Acyn WS cks, Se SeES.e MNS Stee oben inves late ace ee Sys oeaas acme 153
Economic importance of the gladiolus industry........2.....4....---+:-- 153
TES CHISD SEs a hae ca aN ei cg a is RE TP ee ir rR eM ar ie ee 153
eaponmie tianioruarnice;or, the diseaseu..0 Ff ww ss oa. bob Be Mes ee eee es 155
SUPER TEVE CLAIR pena Oa ge eee ee Pe eR eS Gr 156
pune sla VeSee ley case: shrs pete ce ecole ine ees eres at GP ye Pe aR Reet tone eee 156
O@nigtiiercormis ee oer. ae wos he ae a eboiaess Loree ae Sn eee 156
Ie Ae elma foe Pec) xis alk nee iste Ss ok tise 2 OR Pal Pee cag Inne eg aot 9 157
ARAN OUND ene he oes OES. cosh a Siac ace ue, ee he cd Oe ra aes, ene aa 158
LENG (STAUIGUE a le beet ees cd a Re, aR aeerenly Sota neae Sot unites Reece eee rete c 158
IMirvn@e lintitraecs a. veetatackrda woe aise c.0 fiat Woe ariel tie shel Reeve. = ieee eis 159
Source orleat tmbechionhata oncck ck: cio ceke Eee eee oh ae ee eee 159
Bares ot Comm imbeehtotiwor, 1k 22. ods = Peseta eka ee, ese oe 161
Longevity of the organism on the foliage and in the soil............ 162
eS URRPIEMCL INC OVNI MMEM ost act eke yale Mie Seuens: oe ok aera iatel sian seus hag ere ger ae ened ee ai 163
In@ctilettOn Cxperinaenics pa ciestces seta or er eee weet te gee oar reer ae 163
Pe PR GM ICAlsISUOLOPY: =. ce. aunt 2 c-Si ee hove es ate cama oe ales Pare 167
IGE ES i aie ee ee AA Sen ne SH ee ear ee le SCL AN PS BERL 167
(CRONE wv Sh Pepe oa irae ed ie eg RAG aensa cas ©. ono icecany cc 168
Cultural ee ners Ohet ne Man etsy. 4 Ne eee Seek eas estat 168
(Clormitirall., Gach Rao ee es MIR TS Batt eon fer ie ned ol Ae 172
ee MMREURETNONEIIES «ccc a':u ssi eiecd + vosleleserem « Sieve) +)s leer gat rede yaa oyeiake! « 172
AG rHamnIBGRC AIM GMLS Hes tsar chats oucSeles StS Syedeis)oyave tes S1 ye ayact #2 sannileeateenpelsnecedetet - 173
Healthy corms in soil free from the pathogenes..........--+.-++-+++-- 173
Healthy corms in soil known to harbor the pathogenes..............-.. 175
Diseased corms in soil free from the pathogenes....................... 175
RapiGiiVey EC AEMICINES ck, ths pare’ eect aha ume ale BG Gelb eyalsi= cm ae etme bee a eee 175
/aNolmbbaaval “mesh calelaQesieey eine aati ts eaprer ee eer PGi D cpt Onc cyeiascee conn tee 176

Experiment 1. Treatment of corms with formalin and corrosive
Shislimmate: SOlmutHOnss tat coerulea = ecient eee RC 176
Experiment 2. Formaldehyde gas as a disinfectant........... ey

Experiment 3. Hot-water and hot-air treatments of diseased
CIR IAS is so Bice Oak pe koe Re ale Spare tas ane gt Ae eee 177
Scoville SATIS ae one Oe Ae ire Eg tests te rh a ere RRP od TUG Pn len ee. Pecan 178

Perpsnume nt F. OMEMUCAIS .-..5 Jie oats. eiseneren tte chee een ces nae es aes 107)

Experiment 2. Formalin as a soil disinfectant...............+.--.+--- 179
Experiment 3. Formalin as a soil disinfectant... .-. 05. ....52-655i25:- 180
ena eS dere ad Fence SRt Saat g ans, « eRe ee ud ot aesm nik) 5 ompar eC 180
aMMMMnIrE Aa MRT cr Uo es 0nts Soto scevaiatetaa t's Satter Sie ule asin ones Siero wecee 180

151

THE HARD ROT DISEASE OF GLADIOLUS!

L. M. Massey
THE HOST PLANT

The gladiolus is a cormous, summer-flowering plant. Pax (1889)?
classifies it as a member of the family Iridaceae, of the tribe Ixioideae,
subtribe Gladioleae, genus Gladiolus. The species of Gladiolus may be
grown from corms, from cormels (the grayish to black, hard-shelled bodies
formed on underground stems at the base of the new corm), or from seed.
The plants are indigenous to South Africa, where, according to Crawford
(Crawford and Van Fleet, rorr:3), about fifty species have been dis-
covered. This writer states: :

It is also a native of middle Africa, central and southern Europe, Persia, Caucasus,
and the country around the eastern end of the Mediterranean. About forty additional
species have been found in these localities, and one in Hampshire, England. These have

been hybridized and crossed until they are so mixed that it is impossible for the ordinary
grower to say what blood may have entered a given variety —nor does it matter.

ECONOMIC IMPORTANCE OF THE GLADIOLUS INDUSTRY

According to Hendrickson (rorr), there are from four hundred to five
hundred acres in the United States devoted to gladioli, the annual pro-
duction of corms being from 14,000,000 to 15,000,000 and the estimated
value of. the crops $250,000. In New York State, besides many small
growers there are two growers each having over one hundred acres devoted
entirely to the cultivation of gladioli. A list of the members of the
American Gladiolus Society which appeared in 1914 in the Modern Gladiolus
Grower (1:31-32) contains two hundred and twenty names, of which but
sixty-three are those of amateurs and twenty-eight those of foreign dealers
and growers. The output of these growers and dealers represents only
a portion of the total output of the United States. Almost every florist
is more or less interested in the production of corms and flowers of the
gladiolus, which appears to be increasing in popularity as a cut flower.

THE DISEASE

The name hard rot was given to the corm stage of the disease under
consideration by Wallace (1909:18), who makes no reference to a leaf
stage. This name was given to distinguish the disease from other corm

1 Also presented to the Faculty of the Graduate School of Cornell University, January, 1916, as a major
thesis in partial fulfillment of the requirements for the degree of doctor of philosophy.

ACKNOWLEDGMENT. The writer wishes to express his indebtedness to Professors Donald Reddick and
H. H. Whetzel, under whose immediate direction the work was conducted, for helpful criticisms and
suggestions.

2 Dates in parenthesis refer to bibliography, page 180.
153

154 BULLETIN 380

diseases, such as dry rot and soft rot, with which both Wallace and
Fitzpatrick® worked prior to 1912. The writer took up the investigation
of these diseases in 1912 and has given them constant study since that time.

Apparently the hard rot disease of gladiolus exists wherever gladioli
are grown. Specimens have been received from many of the largest
growers in the United States. Horticultural publications contain many
references to corm rots, and no doubt much of this injury is due to the
hard rot disease. Plants growing in the greenhouse at Cornell University
from corms received from Italy by A. C. Hottes bore the leaf stage of the
disease. The writer has received corms affected with hard rot from
Canada, Germany, and Holland.

Prillieux and Delacroix (1894) report having studied a disease of the
gladiolus in which the tissue was deeply corroded, but the writer is unable
to determine whether or not it is the same disease as the one considered
in this bulletin. Unpublished notes placed at the disposal of the writer
by Professor F. C. Stewart, of the New York: (Geneva) Agricultural
Experiment Station, mention the only distinction observed, prior to
Wallace’s thesis (1909), between two types of rots. Concerning specimens
of diseased corms received from a New York State grower, Professor
Stewart suggested the probability that they were affected with the bac-
terial disease described by Prillieux and Delacroix, since he was unable
to locate any trace of fungous hyphe in the diseased tissue. Wallace
(1909: 15) was of the opinion that the corms received by Professor Stewart
were affected with the hard rot disease.

In 1874 Passerini collected specimens of the leaf stage of the hard
rot disease, which he contributed to exsiccatz of Rabenhorst’s Fungi
Europaet.

Saccardo (1884) reports the occurrence of the leaf stage of the disease
on Gladiolus segetum at Parma, Italy, and on Gladiolus gandavensis at
Coimbra, Portugal. Allescher (1897), in addition to the occurrence
on hosts listed by Saccardo, reports the disease as occurring on the leaves
of Gladiolus palustris in Silesia. So far as known to the writer, the leaf
stage of this disease has never been reported in America. A “blight”
is frequently mentioned in horticultural publications, but the descriptions
of the injury are in all cases so indefinite that it is impossible to determine
what diseases the writers had under observation. Hicks (1907:35) and
Childs (1907) write of gladiolus leaf blight, but there is nothing in their
writings sufficiently definite to make it possible to determine the nature
of the injury. Halsted (1894-1901) reports having worked on gladiolus
diseases, but leaves the reader in doubt as to what the diseases were.

’ Unpublished notes of Professor H. M. Fitzpatrick, of Cornell University, covering his investigations
of gladiolus diseases, were kindly placed at the disposal of the writer.

THe Harp Rot DIsEAseE or GLabDIOLus 155

Undoubtedly the leaf stage of the hard rot disease occurs more generally
throughout the country than is indicated by an examination of literature.
This is due to importation of stock from Europe and exchange of stock
by growers in this country. On the other hand, the writer has observed
specimens of the disease on the foliage of plants grown by but three large
growers.

Foliage affected with the disease was first observed by the writer in
1912 in seedling beds, and later on plants grown from cormels. Not
until the season of 1915 did the writer find the disease on the foliage of
large plants, at which time six plants of flowering size were observed to be
affected. In many cases large flowering plants of different varieties
have been observed growing in seed beds or among plants from cormels,
all of which were badly diseased and yet the large plants showed no signs
of the leaf stage. Large plantations of cormels have been observed in
which fifty per cent of the plants bore the disease on the foliage.

Nothing has been found in literature that would indicate that there
is any relation between the leaf stage and the corm stage of the disease
under consideration. It has been the writer’s fortune to obtain conclusive
evidence that they are but different stages of the same disease, and the
experimental data leading to this conclusion are here set forth.

ECONOMIC IMPORTANCE OF THE DISEASE

No figures are available to show the economic importance of the hard
rot disease of the gladiolus, but it must be considerable as compared with
the extent of the industry. Many thousands of corms are discarded
during the winter and in the’ spring at planting time because of their
diseased condition. During the summer many thousands of corms fail
to reach maturity, due to the decay of the parent corms in the soil. While
there are other diseases of the gladiolus, it is the opinion of the writer,
based on observations made during the past four years, that. a large pro-
portion of this loss is due to the hard rot disease. Several varieties of
gladiolus that have been examined showed fifty per cent or more of
the corms to be affected by hard rot. So far as the writer knows, no
variety is immune.

While the loss caused by the leaf stage of the disease is materially less
than that caused by the corm stage, it is still of considerable importance
to the grower. It has been observed that when the foliage of seedlings
and of plants from cormels is affected by the disease, the corms are smaller
than those of plantings that were free from disease. Therefore the
decrease in size must be considered along with the total loss of many
thousands of corms. To this must be added the extra expense incurred
by the grower in sorting and selecting more or less healthy corms from

156 BULLETIN 380

diseased lots, either in filling orders or for his own planting. All in all,
the loss must take from the producer a yearly toll of surprising magnitude.

SYMPTOMS
On the leaves

The first signs of the disease on the leaf are minute brown or purplish
brown discolored areas more or less circular in outline. These lesions
usually appear in July or early August. The color of the diseased areas
deepens somewhat with age until a shade from reddish brown to almost
black is reached. Spots of a reddish brown color predominate. In the
older lesions there is a well-differentiated center, light gray in color and

dotted with numerous black bodies which are very apparent (Plate xv,1). ~

Surrounding the center is a prominent zone, varying from purplish brown
to black in color, which blends into the green of the healthy tissue.

The lesions are more or less circular but with straight sides where
they are limited by the midrib and the edge of the leaf. At first the dis-
coloration may be visible on only one side of the leaf, but it soon makes its
appearance on the opposite side, so that lesions appear practically identical
on either side. They are few or numerous, and vary in size depending
on conditions. The coalescence of several or many spots may occur,
causing the formation of a single large necrotic area along the entire
side of a leaf. Lesions on the tips of the leaves are usually larger and
less characteristic than those below. In some cases the ashen gray centers
of diseased areas drop out, giving a shot-hole appearance. This is more
likely to occur with spots on large flowering plants than with those on
seedlings.

On the corms

Hard rot lesions appear in the fall as minute water-soaked spots, of
a reddish brown to brownish black color, usually on the sides and the
lower half of the corm but not infrequently on the upper half as well
(Plate xv1). It is usually necessary to remove the husks (sheathing leaf
bases) from the corms in order to see the lesions, although in some cases
the husk also is diseased. The lesion on the husk serves as an indication
of the more important lesion underneath. There is no sharp line of
demarcation between the healthy and the diseased tissue.

As the spot increases in size, the center becomes sunken, the color
deepens to a distinct black, and the margin becomes more definite.
A narrow ring, water-soaked in appearance, still indicates the advancing
decay. The more definite margin of the older spots is due to the rapidity
with which the sunken condition follows the advancing water-soaked area,
due to drying of the tissue. The tissue gradually becomes hard, in

BULLETIN 380 PLATE XV

HARD ROT LESIONS ON LEAVES AND CORMS

1, Lesions on leaves of gladiolus seedlings. X 2

2, Lesions on corms. The two upper corms show lesions well advanced, with
the diseased area blending into the healthy tissue. At the bottom the corm on
the left has been cut in two in order to show the depth to which the disease has
progressed. Natural size

BULLETIN 380 PLATE XVI

HARD ROT LESIONS ON CORMS

Different stages in the destruction of acorm. 1}

THe Harp Rot DISEASE oF GLADIOLUS Lisi

some cases extremely so, making it difficult to cut the tissue with a
sharp knife. :

Many small lesions may coalesce into.one large lesion, in some cases”
leaving areas of more or less normal tissue insulated in a large sunken
area. Enough tissue not completely decayed may be left to indicate
the margins of the formerly separate lesions. Frequently the disease
advances so far that the corm is reduced to a hard, shriveled, and wrinkled
mummy.

Excepting in very late stages — and in some cases not even then —
the lesions do not extend deeply into the corm. The usual range is from
one to five or six millimeters (Plate xv, 2). If conditions are not favor-
able for the development of the rot, the active border disappears, soon
assuming the sunken, darkened aspect of the central part. When this
stage is reached the diseased tissue can be chipped out with the finger
nail, leaving the apparently healthy tissue beneath, as if the disease were
not now advancing and the plant had formed a callus over the affected
area.

Plants of more or less dwarfed, stunted appearance, which sometimes
fail to produce blossoms, are to be found throughout the fields during
the growing season. The leaves of these plants usually turn brown
and die, the plant having the appearance of having died from drought.
In a dry season the number of these plants is unusually large. At this
time there is no decay of the new corm which is being developed, but
rather the injury is caused by the premature decay of the parent corm
before the offspring has developed a sufficient root system to enable
it to supply its own moisture and food. This premature decay of the parent
corm is not necessarily due to the advancement of the hard rot disease,
but probably in most cases to the entrance of saprophytes which cause
a rapid disintegration of the corm.

ETIOLOGY

The hard rot disease of the gladiolus is caused by the fungous pathogene
Septoria Gladioli Passer. Passerini collected specimens of the leaf stage
of the disease on the foliage of Gladiolus segetum near Parma, Italy, in
June, 1874, which he contributed to Rabenhorst’s Fung: Europaet —
a collection of exsiccatz material. On this packet of exsiccatze material
is written the original description of the fungus.t Passerini noted the
occurrence of the disease only on the leaves. None of the other investi-

4 Rabenhorst, Fungi Europaei.
1956. Septoria Gladioli Passer. hb.
Perithecia punctiformia atra in macula cxarida fulvomarginata: sporae cylindricae subrectae continuae
hyalinae cirrose ejectae.
Ad folia G. segetum Vigheffio prope Parmam.
Junio 1874. leg. G. Passerini.

158 BULLETIN 380

gators found a sporulating stage of the fungus known to cause the hard
rot disease of corms, and consequently the septorial fungus on the leaf
was not associated with the organism causing the rot of the corms.

it Life history

Pycnidia (Plate xv, 1) of the hard rot fungus become visible usually
within four or five days after, or in some cases even simultaneously with,
the appearance of the lesion on the leaf. They are imbedded in the tissue,
but protrude sufficiently to form black papillae which are visible to the
naked eye.

The pycnidia arise from intercellular mycelium (fig. 38). They
measure from too to 160m in diameter by 60 to 130m high, the

———
————
—

—

iM)

Fic. 38. PYCNIDIUM OF SEPTORIA GLADIOLI
Section through the pycnidium showing how the spores are borne. (Outlined

with a camera lucida.) XX 333
average being 127m in diameter by orp high. The outer wall of the
pycnidium consists of pseudoparenchymatous tissue which is brown in
color.

From a more or less inconspicuous inner layer of thinner-walled pseudo-
parenchymatous tissue, hyaline conidiophores arise. From these conidio-
phores spores are cut off by constriction. In his description of the fungus
Allescher (1897) describes the spores as being unicellular and measuring
from 30 to 54m long by 2 to 2.5 u4in diameter. However, an examination
made by the writer of specimens contained in packet no. 1956 of Raben-
horst’s Fung: Europaet, as well as of fresh material, shows that the spores
are usually three-septate. As measured by the writer they are from 20 to
55 long by 2.25 to 4u in diameter, the average being about 4o by 3 uy.
The spores from fresh material are cylindrical, almost straight, and hyaline.

“su niy Seabed nite es

5 RE

Bie Ai)

THE Harp Ror Disease or GLADIOLUS 150

When placed in water containing small pieces of leaf tissue, germination
occurs in eighteen hours. From one to several germ tubes may develop
from a single spore (fig. 39).

Mycelium

The mycelium in the corm is intercellular (figs. 40 and 41). It usually
measures from 1.5 to 2.5 u in diameter, but is in some cases even double
this size. The mycelium is septate and varies from olive-brown to
black in color.

FIG. 39. SPORES OF SEPTORIA GLADIOLI

Some of the spores have germinated. (Outlined with a camera lucida.) XX 666

Source of leaf infection

No sexual stage of the fungus has been found. Old leaves bearing
pycnidia when exposed out of doors throughout the winter showed usually
only empty pycnidia when examined the following spring. From the
results of experiments subsequently discussed, apparently the mycelium
of the fungus is able to live over winter in the soil. This suggests the
possibility that infection is produced on the foliage by rain splashing
soil containing mycelium on to the plants, or by the plants being beaten
down on to the soil that harbors the pathogene. However, seedlings ©
around which rye straw was placed to keep them off the ground and to
prevent soil from being splashed on to them, were attacked by the fungus

160 BULLETIN 380

as early and as severely as those not so treated; and attempts to produce
infection on the foliage of large plants by bending them over on to the

Fic. 40. HISTOLOGICAL EFFECT OF SEPTORIA GLADIOLI

Section of gladiolus corm through diseased tissue. The presence of intercellular
mycelium, and the absence of starch in many cells, should be noted. (Compare
with figure 41.) X 300

soil have thus far failed. Not enough work has been done to either prove
or disprove these suggested sources of infection of the foliage.

Fic. 41. MYCELIUM OF SEPTORIA GLADIOLI

The intercellular mycelium is shown much magnified. (Camera lucida drawing.)
xX 600

In the greenhouse, the incubation period of the fungus on plants
that were sprayed with water containing spores in suspension was about
twenty days.

THE Harp Rot DISEASE oF GLADIOLUS 165

Source of corm infection

An examination of corms harvested from seed beds where the foliage
was badly diseased, has frequently shown from sixty to seventy per cent
of them to be affected with hard rot. This, together with the fact that
infection was produced on corms by placing in contact with them water
containing spores in suspension (page 167), suggests the probability that
infection is produced by spores being washed down from pycnidia formed
on the foliage to the soil, where they germinate and infect the corms. As
seeds are not planted very deeply, this could readily take place. It is
unusual, however, for the disease to appear on the foliage of large flowering
plants; and as pycnidia have not been observed to be formed on the corm,
it seems that the fungus is carried over the winter primarily, if not entirely,
in the mycelial stage, no spore form being necessary for the existence of
the pathogene.

The fungus can be isolated from lesions on the corms at any time’ during
winter or spring. This shows that the living organism is carried to the
soil along with the corm at planting time. The offspring from diseased
corms may or may not be diseased. As discussed under control (page 173),
selected healthy corms grown in soil in which gladioli have never been
grown have without exception given sound offspring. This indicates
that the fungus is not a natural inhabitant of the soil. Furthermore,
three hundred corms, all of which showed hard rot lesions, were planted
in soil in which gladioli had never been grown, and seventy-eight per
cent of the offspring bore hard rot lesions. Thus it seems that, in the
majority of cases at least, a diseased offspring may be expected from the
planting of a diseased corm.

The fungus does not grow directly from the old corm into the new
one. This has been determined both by observations and by making
numerous cultures from tissue at the juncture of parent and offspring.
The fungus must either grow through the sheathing leaf base from the
old corm to the new one, or else, as is probably the case, grow out into
the soil, from which it attacks the newly developing corm.

No observations have been made which would lead the writer to believe
that all infection does not occur in the field. However, it is conceivable
that if corms were stored under humid conditions either in contact with
one another or with moist soil, the fungus might possibly penetrate a
healthy corm from an infected one or from infected soil; or, if they were
stored with soil containing the pathogene around them, there is no doubt
that, under moist conditions, infection could occur in the storage house
as well as in the field.

Diseased corms were minced and placed in soil known to be free from
the pathogene, in which two hundred healthy corms were growing. The

162 BULLETIN 380

pieces of diseased corms were merely sprinkled in among the corms before
covering them with soil and no attempt was made to see that pieces were
or were not in actual contact with the healthy corms. Seven per cent
of the offspring were diseased.

Longevity of the organism on the foliage and in the soil

As indicated by the following experiment, the fungus is carried over
winter on diseased tops:

Two hundred corms which had been grown for three consecutive years
in soil that had never before been used for growing gladioli, were again
planted in similar soil in 1915. Previous to planting, the corms were
examined and found to be absolutely healthy. After setting the corms,
tops from cormels which had been badly affected by the disease the pre-
vious year and which had remained out of doors on the ground throughout
the winter, were scattered in the row. The tops and the corms were then
covered with soil. These plants were harvested in September and the
corms stored in a cool place. When examined early in December it was
found that eighty per cent of the corms showed hard rot lesions. _Prac-
tically all the diseased corms had many lesions on them, and the disease
was well advanced. Septoria Gladioli Passer. was isolated from many
of these lesions, proving that this fungus caused the disease. Healthy
corms around which no diseased tops were placed but which were other-
wise given the same treatment, showed no signs of disease.

The results of experiments indicate that the fungus is able not only
to live over winter on old tops on the ground, but also to live in the soil
throughout a period of at least four years. In r1g915 selected healthy
corms were planted in soil in which gladioli had been grown the previous
year, and also in soil in which no gladioli had been grown for (a) one
year, (b) two years, (c) three years, and (d) four years. During the
intervening time the soil had been planted respectively to (a) rye and a
crop of rye and vetch, (b) rye and timothy, (c) oats, hay, seeded to clover,
cover crop of rye and vetch, (d) grass. In each of the five plots of ground
two hundred and fifty healthy corms were planted, two hundred of them
being planted in a single row and the remaining fifty in lots of ten at five
different places in the field. The corms were harvested in September and
stored in a cool place.

Results of these experiments were recorded the following December.
Forty-seven per cent of the corms which were grown in the plot in which
gladioli had been grown the previous year, were diseased, fifty per cent
of the diseased corms showing characteristic hard rot lesions. The corms
from the other plots were diseased as follows: (a) twenty-four per cent,
forty per cent of which bore hard rot lesions; (b) twenty-three per

THe Harp Rot DIsEASE or GLADIOLUS 163

cent, thirty-seven per cent of which bore hard rot lesions; (c) fifty-two
per cent, eighteen per cent of which bore hard rot lesions; (d) forty-seven
per cent, ten per cent of which bore hard rot lesions. Care was taken
during the summer to avoid contaminating these plots by introducing
affected soil from other fields, and the location was such that it is extremely
doubtful that the wind could have entered as a factor. Since healthy
corms planted in soil in which no gladioli have been grown give healthy
offspring, it follows that the organism must be able to live for at least
four years without the presence of the living host. No doubt decaying
parts of plants were left in the soil when the last crop was harvested,
but it is probable that, at least in the case of plot d, these plant parts
were entirely decayed in the four years which intervened between the
harvesting of the last crop of gladioli and the planting of the healthy
corms used in this experiment.

Pathogenicity

The pathogenicity of Septoria Gladioli Passer. was established first
for the mycelial stage. The mycelium of the fungus was discovered by
Wallace (1909:33), who, after having observed its presence in thin
sections of diseased tissue of the corm, succeeded in obtaining the organism
in pure culture. He later (1910 a) suceeeded in producing the character-
istic lesions on experimentally inoculated corms, and reisolated the fungus.
Following Wallace, Fitzpatrick (see footnote, page 154) records having
produced the characteristic lesions on corms artificially inoculated in
moist chambers, from which the fungus was reisolated.

Besides noting the constant association of the mycelium with lesions
on corms through microscopical examinations of diseased tissue, the writer
has made numerous isolations of the organism from these diseased areas.
The growth of the mycelium was studied in pure culture and infection
was produced at will, not only in moist chambers in sterile sand, but
also in the greenhouse, and in the field under natural conditions.

Inoculation experiments

Corms were selected which after having been in the storehouse for four
months showed no signs of disease. This necessitated the removal of the
husks. The surface was sterilized by immersing the corm in fifty-per-
cent alcohol for three minutes, then in 1—1000 corrosive sublimate solution
for ten minutes, and finally rinsing in sterile water. These corms were
then planted, some in sterile sand in moist chambers, some in soil in the
greenhouse in which gladioli had never been grown, and some out of
doors in soil never before used for the growing of gladioli.

164 BULLETIN 380

For inoculation, mycelium growing in pure cultures on solid media
was used. A bit of the medium containing mycelium was removed under
sterile conditions, and in some instances smeared over a part of the unin-
jured surface of the corm; in other cases the corm was first injured by
needle punctures and the culture was then smeared on the surface. The
corms were permitted to remain in the soil for a period of two or three
weeks, when they were removed and the growth they had made was
cut off.

In practically all cases one hundred per cent infection was obtained.
Most of the corms showed the dark brown, water-soaked areas, char-
acteristic of the hard rot disease, when dug. The remainder showed the
lesions very soon afterward. Equally as abundant infection was obtained
on the uninjured corms as on those punctured by the needle. From
diseased areas of the affected corms the fungus was reisolated and grown
in pure culture, where its growth corresponded in every detail with the
organism used for the inoculation. Corms similarly treated but having
no mycelium placed in contact with them remained healthy in all instances.

In order to further test the ability of the fungus to produce disease,
sound corms were planted in soil in which gladioli had never been grown,
and permitted to grow to maturity. On August 15, 1914, as the offspring
were developing from the parent corms, the soil was inoculated with
mycelium of the fungus. The inoculum was prepared by grinding cul-
tures of the organism on oatmeal agar with cornmeal, and was applied
by placing a small handful of the mixture around each corm in immediate
contact with it. Of the one hundred corms thus inoculated, seventy-three
showed characteristic hard rot lesions when the results of the experiment
were recorded the following December. Reisolations of the fungus were
obtained from many of the diseased corms. Corms from plants which
had not been inoculated with mycelium remained absolutely healthy.

The above experiments prove the ability of the mycelial stage of the
hard rot fungus to attack gladiolus corms. The experiments given below
show that this mycelium is but a stage of Septoria Gladioli, which Passerini
described as occurring on the foliage of Gladiolus segetum in Italy.

A pure culture of the fungus Septoria Gladioli Passer. was obtained
from the germination of a single spore from a pycnidium formed on the
leaf of a gladiolus seedling. The resulting fungous growth was identical
with that obtained from isolations from small pieces of diseased corm
tissue. Mycelium thus obtained from a single spore was used to inoculate
healthy corms, some of which were planted in moist chambers in sterile
sand, others in soil in the greenhouse known to be free from the pathogene,
and still others out of doors in soil in which gladioli had never been grown.
Numerous experiments were performed, and in all cases one hundred

THE Harp Rot DISEASE OF GLADIOLUS 165

per cent infection was obtained by smearing a small quantity of an agar
culture of the mycelium from a single spore on the surface of the corms.
The fungus was reisolated from diseased areas on the corms, and its
growth in pure culture was found to be identical with the organism pre-
viously isolated from corms and from the germination of a single
pycnospore.

In order to test the ability of the fungus isolated from a lesion on a
corm to attack the foliage, a small piece of an agar culture of the organism
was mixed with a little sterile distilled water and painted on the foliage
of seedlings and flowering plants growing in the greenhouse. The seed-
lings were then inclosed in bell glasses lined with moistened filter paper,
while the parts of the large plants on which the mycelium was placed
were inclosed in a lamp chimney stoppered at both ends with cotton.
Seedlings and large plants were similarly treated with mycelium obtained
from the germination of a single pycnospore. Plants were similarly
treated, except for the omission of the mycelium, to serve as a check.

Inoculations were successful with both the mycelium from the germinated
spore and that from the diseased corm. Infection was evident on the seed-
lings within ten days. The lesions differed somewhat from those found
under natural conditions, infection manifesting itself in the form of large,
dark, water-soaked areas, with the early death of the entire area over
which the inoculum was painted. The lesions produced by mycelium
from the two different sources were similar.

On the large plants, infection was observed within fourteen days after
inoculation, the lesions being identical on the plants inoculated with
mycelium, from the two different sources. At first a dark area, water-
soaked in appearance, was formed, and then the lesions turned brown
due to the death of the tissue. The lesions in no case extended much
farther in area than that covered by the culture of mycelium painted
on the foliage. The most significant fact is that pycnidia developed in
these lesions on the leaf on practically all of the twenty-four plants inocu-
lated with the mycelium, regardless of whether the mycelium was from
a germinated spore or from a diseased corm. Although many pycnidia
failed to reach maturity, spores were formed in several of them. These
spores were germinated and the fungus was-obtained in pure culture.

In October, 1914, Septoria-like spores were found in a culture of the
organism isolated from a diseased corm. ‘These spores, together with
others obtained from pycnidia on seedling leaves, were used in the
following experiments:

Seeds were planted in three flats in the greenhouse and the plants were
permitted to grow until they were from two to four inches high. The
plants in one flat were sprayed with water containing, in suspension,

166 BULLETIN 380

spores from a culture of the fungus isolated from a diseased corm; the
plants in the second flat were sprayed with a suspension of spores from
pycnidia formed naturally on seedlings; the plants in the remaining flat
were sprayed with water containing no spores, for a check. The seed-
lings were then covered with bell glasses lined with moistened filter
paper, and the three flats were placed in a large moist propagating
chamber for seventy-two hours.

An examination of these seedlings twenty days after they had been
sprayed with the suspension of spores in water showed evidence of infec-
tion. Small yellowish brown areas were apparent and numerous pycnidia
appeared in these lesions a few days later. The lesions were characteristic
of-those found on the seedlings under natural conditions. The check
plants alone remained healthy, infection occurring on plants which were
inoculated either with spores from culture or with spores from pycnidia
formed under natural conditions. The fungus was again obtained in
pure culture from the germination of single spores from pycnidia formed
on both lots of infected plants.

It then seemed desirable to determine whether or not corms could
become infected by spraying spores upon them. The surfaces of thirty
healthy corms were sterilized by immersing them first in fifty-per-cent
alcohol for three minutes, then in 1—1ooo corrosive sublimate solution
for ten minutes, and finally rinsing in. sterile water. Ten of these corms
were then planted in each of three moist chambers containing moist sand
which had previously been subjected to steam at ten~ pounds pressure
for two hours. Corms that were particularly depressed at the crown
were selected for the experiment, in order that a cubic centimeter or
more of water could be held in each of these cavities. Water containing
spores in suspension was placed in the depressed areas of the corms in
two of the moist chambers. The spores for one chamber were obtained
from pycnidia formed naturally on the foliage of seedlings, while for
the other chamber the spores were obtained from a culture of the fungus
isolated from a diseased corm. The third chamber was used for a check,
water containing no spores being placed in the cavities of the corms.
One-half of the corms in each chamber were then pricked with a sterile
needle in the area covered by the water.

Observations made twenty days later showed most of the corms in
the two chambers which were inoculated with spores to be infected. Six
days later, when they were removed, all the inoculated corms showed the
characteristic hard rot lesions, while the check corms remained healthy.
Lesions were as abundant on corms that had not been injured as on those
punctured with the needle. About one-half of the corms showed hard rot
lesions on the sides, where evidently spores had been washed over from

THE Harp Rot DISEASE OF GLADIOLUS 167

the concave crowns. The fungus was isolated from many of these diseased
areas and again obtained in pure culture.

In the spring of 1915 healthy corms were planted in soil in which gladioli
had never been grown, and allowed to grow to maturity. On August 21
the soil was removed from around thirty of these plants and water con-
taining a suspension of spores was poured around the corms. At this
time the offspring were about one-half inch in diameter. The spores
for inoculating fifteen of the corms were obtained from pycnidia formed
naturally on the foliage of seedlings, while for the other fifteen corms
the spores were obtained from a culture of the fungus isolated from a
diseased corm. For a check, corms were given the same treatment
except that the water poured around them contained no spores. The
soil in which these plants were growing was kept moist for the follow-
ing three days.

The corms were harvested in the following September and stored in a cool
place. When examined in November it was found that ten of the fifteen
corms inoculated with spores from pycnidia showed hard rot lesions;
also, six of the fifteen corms inoculated with spores from culture showed
lesions characteristic of the hard rot disease. The check plants remained
healthy. This experiment is significant in showing not only that spores
from cultures and from naturally formed pycnidia are able to infect the
corms, but also that it is possible for infection to occur on the corms from
spores discharged from
pycnidia on the leaves.
The spores are washed
down into the soil,
where they germinate
and produce infection.

Pathological histology
- Leaf

An examination of
thin sections of leaves
bearing young diseased

Fic. 42. HISTOLOGICAL EFFECT OF SEPTORIA GLADIOLI

4 Camera lucida drawing of a free-hand section through a hard rot lesion
areas shows the lesion on the leaf of a seedling. The cells are beginning to shrivel and collapse.

X 200

produced by the fun-
gus to be necrotic. The cells turn brown, shrivel, and collapse (fig. 42).
Here and there a cell may be found filled with a yellow, granular or
oil-like substance the identity of which is undetermined. The diseased
area is usually but from one-third to one-half the thickness of the
healthy tissue.

168 BULLETIN 380

Corm

In order to study the histological changes that occur in the corm, com-
parative studies of healthy and diseased tissues have been made. Both
microtome and free-hand sections have been used, the former being less
satisfactory because of the difficulty encountered in sectioning prepared
material. Sections were stained with a weak solution of iodine in order
to study starch content of cells, and with Haidenheim’s iron-alum-heema-
toxylin and aniline blue for a general study of the tissue.

While the cells of healthy tissue are densely packed with starch, those
of diseased tissue show but very few starch grains or none at all (figs. 4o
and a1 [page 160], and 43). This, together with the deposit in the
diseased area of a yellow substance of undetermined composition, is the
most pronounced effect to
be noticed by comparing
sections of diseased and
healthy tissue. Espe-
cially in the early stages
of the disease, nuclei and
even the cytoplasm
appear but slightly dis-
turbed. The cell walls
retain their shape for
some time after the starch
has disappeared. Later,

i : ; “" shrinkage takes place and

Camera lucida drawing of a microtome section through medium
of healthy and diseased tissue. The layer of comparatively thin- the cells collapse, the
walled cork cambial cells separating the starch-filled healthy cells Ps P
from the diseased cells, which contain little or no starch, should walls becoming distorted
be noted. X 300 :

and broken. This last
effect is no doubt due to loss of moisture rather than to any direct
action of the fungus.

A layer of cork cambium is formed at the juncture of the diseased
and the healthy tissue (fig. 43). Young, actively advancing lesions do
not show this layer of thin-walled cells, but it is to be found in those
instances in which it appears that the advance of the disease has been
checked and the canker healed. In cases in which the diseased area
can be chipped out, the break is at this layer of cork cells.

Fic. 43. HISTOLOGICAL EFFECT OF SEPTORIA GLADIOLI

Cultural characters of the fungus

Pure cultures of Septoria Gladioli Passer. were obtained from isolation
plantings of diseased tissue from a corm. The surfaces of corms showing
hard rot lesions were disinfected by immersing them in fifty-per-cent
alcohol for three minutes, then in 1-1000 corrosive sublimate solution

THe Haxkp Rot DISEASE or GLADIOLUS 169

for ten minutes, and finally rinsing in sterile water. By means of a sterile
scalpel the surface of the corm was cut away and a small piece of the
tissue at the advancing margin of the lesion was removed to a sterile
medium. Comparatively few contaminations were obtained in the large
number of isolations made in this manner.

No marked difference was observed in the growth of the mycelium
on nutrient or on soil-extract agar, or on other solid media consisting of
agar and various plant decoctions, such as of gladiolus, potato, oats, corn,
and beans. On the
other hand, rolled-oat
agar® proved slightly
more favorable for
mycelial growth, and
spores were produced
by the fungus only
when growing on this
medium. For this
reason rolled-oat
agar was used almost
entirely for culturing
the organism during
the last year of study,
and the following cul-
tural characters of
the fungus are a rec-
ord of its growth on
this medium.

Macroscopically,
no growth from bits
of diseased tissue

Fic. 44. MYCELIUM OF SEPTORIA GLADIOLI

came lucida grains tes ES of he hard act fans growing
+ ium i on rolled-oat agar. , colorless strands of hyphz radiating from a bit
placed ie medi am of diseased viseue. ond ne cell Ben Cant fake Geena the cells
{ having assumed a globose form. , colorless hyphe to be found in old
previously piewre d cultures. E, an intermediate stage between A and B. X 600

petri dishes is evident

for from seven to fourteen days. However, if the plate is examined under
the low power of the microscope, mycelium radiating from the transferred
piece of tissue can be seen in about four or five days from the time of
making the culture. Frequently the first macroscopical evidence of growth
is the appearance of a black growth on the transferred piece of tissue,
which may be completely covered before the organism invades the

5The rolled-oat agar was prepared as follows: 50 grams of rolled oats, in 700 cubic centimeters of dis-
tilled water, was cooked in a double boiler for about an hour, or until the oats were thoroughly cooked
through. Most of the solid matter was then squeezed through cheesecloth. To this was added 15 grams
of agar and enough water to make one liter of medium.

170 BULLETIN 380

medium. Soon a dense, black colony spreads very slowly into the sur-
rounding medium. After growth of a month the colonies usually do not
exceed one or two centimeters in diameter. If portions of a colony are
transferred to flasks of media or to other plates, the resulting growth is
somewhat more rapid.

Some of the characters of the mycelium as grown in culture are shown
in figure 44. The first strands of hyphz to be seen radiating from the
piece of diseased tissue are hyaline (fig. 44, A). Color frequently makes
its appearance in streaks, which radiate from the piece of diseased corm,
where the hyphae seem to become gnarled. Cells of the much-septate
mycelium thicken, assuming a globose form (fig. 44, B). Well-defined
globular bodies, which appear to be oil drops, soon appear within the
cells. Osmic acid causes these to turn brown. The walls turn brown
with the appearance of these bodies, giving the colony its black color
when viewed macroscopically. The globose or subglobose cells of the
hyphe may remain attached, forming chains, or may separate into
individual cells (fig. 44, B).

Although the growth is usually confined beneath the surface of the
medium, small scant patches of white, aérial mycelium are found occa-
sionally. The hyphz of old cultures is of two kinds: one of compara-
tively long, colorless cells measuring from 1.5 to about 4m in diameter
(fig. 44, C, E); and one of short, thick, globose cells containing the oil
drops mentioned above, measuring from 3 to 6 or 7 uw, or sometimes even
12m, in diameter (fig. 44, B, D).

Scattered through the colonies are areas which under the microscope
appear denser and blacker than other areas. These seem to be caused
by a gnarling, or balling, of the hyphz at these points, together with
the anastomosing of cells of different hyphae, as if pycnidia or other
fruit bodies were to be formed. Cultures have been examined inter-
mittently throughout a period of over three years, and no further develop-
ment of these masses of hyphe has been observed.

Spores of Septoria Gladioli Passer. were first observed in culture in
October, 1914. The mycelium on which these spores were formed was
isolated in the preceding June, from a hard rot lesion on a corm. This
mycelium was allowed to grow in a tube of rolled-oat agar from June
until August 20, when a square block of the medium containing mycelium
was transferred to the slanted surface of about 200 centimeters of rolled-
oat agar contained in a 300-cubic-centimeter Erlenmeyer flask. The
medium contained in this flask was freshly prepared. There were about
10 cubic centimeters of water of condensation in the flask at the base of -
the slanted medium. The culture from which the transfer was made was
well dried at the time when the square of medium containing mycelium

THe Harp Rot DisEasE or GLADIOLUS yt

was removed, and this condition may have influenced spore formation
when the mycelium was placed on the freshly prepared medium.

By approximating the above conditions the writer has been able to
bring about the formation of spores in cultures of the mycelium from
other sources than the one above noted. Spores were formed in a culture
of the mycelium obtained from the germination of a single pycnospore
formed naturally on the leaf. Spores formed in cultures of mycelium
isolated from corms and from germinated pycnospores were identical
in shape and size, thus materially helping to establish the identity of
the two previously unconnected organisms.

Spores in culture have always appeared as minute pinkish white pustules
on the upper edge of the block of medium containing mycelium trans-
ferred from the old culture. Later these pustules may appear scattered
over the surface of the medium of the flask to which the transfer was
made. If at this time transfers are made from this flask to another, the
pustules are formed more readily and in greater abundance.

Owing to difficulties encountered in obtaining sections or mounts of
these pinkish white elevations, but little is known of their structure,
especially in reference to the formation of the spores. Normal pycnidia
such as those formed on the foliage are not produced. The spores are
formed in a very loose stromatic mass. There is an abundance of dense,
pinkish white mycelium, which is still in evidence after spores are no
longer to be found associated with the pustules.

Spores formed in culture are variable in size, ranging from 25 to 97 u
by 1.8 to 3.75 u, the average being 58 by 2.71. Dilution plates of these
spores were made in nutrient agar and practically one hundred per cent
germination was obtained within a period of eighteen hours. The result-
ing mycelial growth was not so brown in color as that isolated from corms.
Many minute, black dots appeared, which, when examined under the
microscope, proved to be aggregations of short, thick-walled cells formed
commonly and more abundantly in cultures of the fungus isolated from
corms. Transfers were made from these plates to tubes of rolled-oat
agar, where the resulting growth was identical with that obtained by
isolations made from diseased corms.

In order to correlate the growth of mycelium isolated from diseased
tissue of corms with that isolated from the leaf, dilution plates of spores
from naturally formed pycnidia were made. From these dilution plates
individual spores, which were so located that they could be removed
singly, were transferred to other plates where germination was observed
under the microscope. The resulting mycelial growth in all cases has
been identical with that isolated from corms when the two were growing
under similar conditions.

172 BULLETIN 380

CONTROL

The great need of some method of combating the organisms causing
rots of gladiolus corms was early impressed upon -the writer, and many
suggested methods of general application were tried for the control of
the rots collectively rather than separately. Another disease, designated
by Wallace (1909:61) as dry rot, was found to be present along with
the hard rot disease in stock which was used in all control experiments.
The lesions produced by the fungi causing these two diseases are so similar
that they can be distinguished only in the earliest stages, and not even
then with a great degree of accuracy. Cultural isolations of the organisms
will often show a lesion to have been caused by the hard rot fungus when
it was selected as being a dry rot lesion, or vice versa. Not only are the
lesions produced by the two fungi similar, but the life histories of the
organisms are not materially unlike except for the fact that no spore
form of the dry rot fungus has been found. From all indications, a treat-
ment applicable to the control of one disease should be of value in con-
trolling the other. At least fifty per cent of the corms used for experimental
purposes were affected with the hard rot disease. This estimate is based
on observations and cultural studies throughout a period of several years.
In practically all cases, after the corms were treated, the organisms have
been isolated from diseased areas in order to make it absolutely certain that —
both were present, and in no case has any treatment resulted, so far as
the writer was able to judge, in materially changing the ratio of the corms
affected by the two diseases.

In view of the fact that control experiments were conducted previous
to, and simultaneously with, life history studies, it is not surprising that
some treatments which at first seemed worthy of trial failed to bring
results. Many of the following treatments have given negative results.
This does not wholly deprive them of their value, for they serve to narrow
down the field of experimental possibilities of control. Many data have
been obtained from the treatments which will be valuable in a further
study of control measures.

SEEDLING TREATMENTS

The hard rot disease on the foliage of seedlings has been materially
reduced by spraying with bordeaux mixture used at the strength of five
pounds of copper sulfate and five pounds of lime to fifty gallons of water.
In ro1q4 the first spray was applied on July 17. This application was
followed by eight other treatments made at intervals of about seven days.
Because of the smooth surface of the foliage, it was necessary to use a
“sticker,” or adhesive, to cause the fungicide to adhere to the plants.
The “ sticker’ used consisted of resin two pounds, sal soda (crystals)

THe Harp Rot Disease oF GLADIOLUS 173

one pound, and water one gallon, which, after being boiled until a clear
brown color was obtained, was added to each fifty gallons of the bordeaux
solution. The seedling beds were sprayed twice the same day for each
application, the second spray being applied as soon as the first was dried
on the foliage. This was done in order to thoroughly cover the plants.
A hand sprayer was used, in which a pressure of from three and one-half
to five pounds could be maintained at all times.

H. H. Groff, of Simcoe, Ontario, informed the writer that he was
successful in controlling a disease of the foliage of seedlings by spraying
with a solution of copper sulfate in water. Specimens of the disease sent
by Mr. Groff to the writer proved to be the hard rot disease. From the
nature of the foliage of the gladiolus, it is probable that the plant is more
resistant to spray injury than are most plants and that a solution of copper
sulfate could be used without causing injury. However, no experiments
have been conducted by the writer using copper sulfate solution as a spray
for the control of this disease, and it is very unlikely that results could be
obtained, because the copper sulfate would be washed away with the
first rain.

Although spraying will greatly reduce the amount of disease on the
foliage, a simpler and more efficient method is to plant the seed in soil in
which gladioli have never been grown. When this was done, and care
was taken not to carry parts of diseased plants or soil bearing the fungus
to these seedlings, it was found that not a single diseased plant appeared
during the summer. The corms of these plants were materially larger when
harvested than the corms of plants whose foliage was attacked by the
hard rot fungus, and no evidences of disease on the corms were observed.
This is the logical way to control the hard rot disease, which causes so much
damage in seedling beds. It is doubtful whether any grower plants
such a large quantity of seed, or has such a limited area of ground, that
soil in which gladioli have never been grown cannot be obtained for this
purpose. If this plot is kept isolated and care is taken not to introduce
the pathogene into the soil, there appears to be no reason why seedlings
cannot be grown on the same area year after year, if necessary, at least
so far as the hard rot disease is concerned.

CORM TREATMENTS
- Healthy corms in soil free from the pathogenes

Selected healthy corms have been grown for the past four years in
soil in which no gladioli had ever before been planted, without a single
corm’s becoming diseased. The fact that these corms were stored through-
out each winter in a room containing diseased corms leads to the con-

174 BULLETIN 380

clusion that the fungi causing the hard rot and dry rot diseases are not
disseminated in storage. It is obvious that in order to obtain results from
the selection of healthy corms, rigid and painstaking care must be exercised
to select corms known to be absolutely free from disease. Any doubtfully
healthy corms must be rejected, for a single diseased corm may serve to
infect the soil in which healthy corms are planted.

To select healthy corms it is necessary to remove the husks and to be
sure there is no evidence of disease on the corms. It is best to do the
selecting in the spring, as near planting time as possible, for, whereas
a corm may be infected in the fall at digging time and still show no evidence
of being diseased, the lesion is sure to be noticeable by planting time.
Previously to planting these corms it is advisable to treat them with
a five-per-cent solution of formalin for thirty minutes, in order to kill
any parts of the pathogenes which may be clinging to them.

In 1912 from two thousand to three thousand healthy corms were selected
in the manner suggested above. They were planted each year in soil
that had not been under cultivation for about twenty years. A com-
mercial phosphate fertilizer was applied to the experimental plots at the
rate of about five hundred pounds to the acre, and the corms were planted
in the usual manner. Care was exercised to see that no foreign soil nor
diseased plant parts were introduced into these plots. The plants received
the usual amount of cultivation and were subjected to the same con-
ditions as commercially grown plants. Spikes of flowers were cut during
the blooming season, and the corms were harvested and stored each
autumn in the ordinary way.

This process of selecting healthy corms and growing them in soil free
of the pathogenes is the only means known that will give an absolutely
healthy crop. Of course the large amount of labor, the carelessness of
laborers, the need of a larger outlay of land, and the inability to procure
land on which gladioli have never been grown or at least not for many
years, are some of the important factors that will at once suggest themselves
to growers, especially the larger growers who produce many thousands
of corms annually. It is admitted that this is a slow and somewhat
undesirable method from many standpoints, yet it is a process that has
proved conducive to results, and undoubtedly can find some application
by all growers. Small growers can readily and with no great loss adopt
such a method for growing gladioli. Larger growers can adopt the process
in part.

Such a method could be begun on a small scale, by selecting as many
healthy corms the first year as conveniently possible and planting them
in soil in which gladioli had never been grown. More selected corms
could be added to this lot the second year, and so on until the grower

THE Harp Rot DISEASE OF GLADIOLUS 175

gradually worked away from diseased to healthy stock. The opportunity
for healthy corms to become diseased is thus lessened, and diseased con-
ditions are in general improved.

Healthy corms in soil known to harbor the pathogenes

When selected healthy corms were planted in soil in which gladioli
had been grown the previous year, the offspring were diseased. The
amount of disease varied from twenty-three to forty-seven per cent.
The possibility suggested itself that some treatment might be devised
which would protect the offspring of the sound corms that were planted,
from the pathogenes that must be in the soil.

An experiment was conducted in 1914 in which the corms of the various
plots were treated with different chemicals. A smal] handful of the
chemicals was placed over each corm previously to covering the corms
with soil. The chemicals used were: plot 1, sulfur; plot 2, air-slaked
lime; plot 3, acid phosphate; plot 4, soot. The soot was suggested by
a grower who claimed to have obtained good results through a liberal
application of this substance to the soil. The plants received ordinary
cultivation during the summer, and the offspring were harvested and stored
in the usual manner. In December, when the results of the experiments
were recorded, it was found that none of the treatments were of any value,
the percentages of disease in the treated plots being practically the same
as that in a check plot where no treatment was given. The experiment
was repeated in 1915 with the same results.

Diseased corms in soil free from the pathogenes

Spring treatments

When diseased corms that had received no treatment were planted in
soil free from the pathogenes, it was found that the offspring gave various
percentages of disease. Seventy-eight per cent of the offspring from
three hundred corms bearing typical hard rot lesions, which were planted
in soil free from the pathogenes, were diseased. In other instances, thirty-
three and fifty-seven per cent diseased offspring, respectively, were recorded
from the planting of two lots of three hundred corms affected with either
hard rot or dry rot, or both.

An experiment was conducted in 1914 to determine whether or not
some treatment could be given these corms at planting time which would
lessen the amount of disease in the offspring when the corms were grown
in soil free from the pathogenes. Corms that bore typical hard rot lesions,
and others that were affected with either the hard or the dry rot disease
or both, received the following treatments: (1) formalin at the rate of

176 BULLETIN 380

one pint of commercial formalin to fifteen gallons of water, for eighteen
hours; (2) corrosive sublimate, 1—1ooo solution, for eighteen hours;
(3) chemicals, in which the corms were rolled and with which they were
covered after being placed in the rows and before covering them with
soil. The chemicals used were sulfur, air-slaked lime, acid phosphate,
and soot. The corms were planted in soil in which gladioli had never
before been grown, and received ordinary cultivation during the summer.

When the corms were examined in December, 1914, the results obtained
indicated that none of the treatments were effective in reducing the
amount of disease. The experiment was repeated in 1915, with the
same results except that corms over which a handful of sulfur was placed
were injured severely by the chemical. Such treatments of diseased
corms have proved to be of no value in controlling the hard rot and dry
rot diseases.

Autumn treatments

Since the lesions on corms attacked by the hard rot and dry rot organisms
are materially smaller in the autumn when the corms are dug than in the
winter, it was thought that possibly the corms could be given some treat-
ment at digging time whereby the pathogenes within the tissues would
be killed. Consequently the following experiments were performed
with the hope of at least lessening the extent of injury to. the corms.

Experiment 1. Treatment of corms with formalin and corrosive sublamate
solutions— In 1914 one thousand corms, of which many had lesions
in various stages of advancement at digging time, were treated,
immediately after digging, with formalin at the strength of one pint of
commercial formalin to fifteen gallons of water, in which they were left
for eighteen hours. An equal number of corms were treated with 1—1000
corrosive sublimate solution for eighteen hours, and an equal number
were left untreated for a check. After treatment the corms were cured
out of doors and then stored as usual.

In the following December, when the results of these treatments were
recorded, it was found that neither had reduced the amount or the extent -
of the diseases. Thirty-five per cent of the offspring from the untreated
corms were diseased, while thirty-seven and thirty-eight per cent,
respectively, of the offspring from the corms treated with formalin and
corrosive sublimate solution were diseased.

The same experiment had been performed the previous season (1913),
with the result that about ninety per cent of the corms of both the treated
lots were diseased while the corms in the check were but seventy per:
cent diseased. This remarkable situation is difficult to explain. There
was a prolonged period of wet weather about the time the treatments

THE Harp Rot DISEASE OF GLADIOLUS 177

were made, so that the corms remained wet for about a week after they
were treated. The corms were either injured by being subjected to the
action of the reagents for so long a time, or else the increased per-
centage of diseased corms was due to some other abnormal condition
brought about by the wet condition of the corms. Many of the corms
bore lesions which were not characteristic of either the hard rot or the dry
rot disease, from which neither the dry rot organism nor Septoria Gladiolt
Passer. could be isolated.

Experiment 2. Formaldehyde gas as a disinfectant.— On the basis of
successful experiments performed for the control of potato scab by the
use of formaldehyde gas, diseased corms were subjected in 1913 to a
similar treatment. Obviously this would eliminate the humid condition
arising from the use of solutions.

In this experiment the gas was generated by the potassium permanganate
method. At harvesting time one thousand corms were placed, immediately
after digging, in shallow trays in a large air-tight box. The formaldehyde
gas was obtained by using enough potassium permanganate to generate
gas at the rate of three pints of formalin and twenty-three ounces of
permanganate crystals to five hundred cubic feet of space, it having
been previously determined that corms thus treated were unharmed.
The treatment extended over a period of twenty-four hours. The corms
were then thoroughly cured in the open air and stored as usual.

In January, ro14, when the results of this treatment were recorded,
it was found that sixty-nine per cent of the corms were diseased while
seventy per cent of the untreated corms from the same lot were diseased.
The hard rot and dry rot organisms were isolated from many lesions,
showing both organisms to be alive. The difference of one per cent in the
amount of disease can easily be explained on the basis of experimental
error, with the resulting conclusion that formaldehyde gas as used in
this experiment is of no value in controlling the corm rots of gladioli.

Experiment 3. Hot-water and hot-air treatments of diseased corms.— Ina
third experiment some means was sought whereby diseased corms could
be subjected to heat, which would kill the organisms within the tissue
without causing injury to the corms. After such treatment the corms
could be planted in soil known to be free of the pathogenes and be depended
on to yield a healthy crop.

Previously to conducting the experiments it was determined that
the thermal death point of the hard rot and dry rot organisms was about
50° C. when subjected to this temperature in a test tube culture for
a period of ten minutes. The tubes were immersed in the hot water as
soon as new growth appeared from pieces of medium containing mycelium
which were transferred to the tubes. It was also previously determined

178 BULLETIN 380

that corms of from three-fourths inch to one and one-half inches in
diameter, when subjected to dry heat at 50° C. for one and one-half hours
or to water at this temperature for one-half hour, were not materially
harmed.

Having thus obtained some idea of the relative resistance of both
corms and the two pathogenes to heat, corms were subjected in 1913
to dry heat and to water at 50° C. for one and one-half hours and one-half
hour, respectively, and the progress of the disease was noted. There
was enough difference between the length of time required to kill the |
fungi and that which caused no injury to the corms to warrant this treat-
ment. The corms used were of a single variety and showed considerable
disease when dug. They were treated on the same day that they were
harvested. For the hot-water treatment a half-bushel galvanized iron
measure was used, the heat being supplied by an oil-stove flame, and for
the dry-air heating a Freas electric oven was used. After treatment
the corms were cured as quickly as possible and then stored in a cool
place as usual. Wet weather lengthened the time necessary to thoroughly
cure the corms more than was desirable.

In the following January, when the results of this experiment were
recorded, it was found that seventy per cent of the untreated corms
bore lesions of either the hard rot or the dry rot disease, while of those
treated with dry heat and hot water eighty-five and ninety-five per cent,
respectively, were diseased. In both cases in which treatments were
given, the corms, besides containing a iarge percentage of disease, showed
the lesions to be more advanced than those in the check. Both the
hard rot and dry rot organisms were isolated from many diseased corms
of both lots, showing the pathogenes to be still alive.

In accounting for the increased percentages of disease in the treated
over the untreated corms, it was found that many of the lesions, besides
being different in appearance from those produced by the two fungi,
were identical with those produced on healthy corms which were sub-
jected to heat under the same conditions and from which neither the
hard rot nor the dry rot organism could be isolated. These lesions were
undoubtedly due to injury caused by the heat. However, a sufficiently
large number of corms bore characteristic lesions of the two diseases,
and the causal organisms were isolated from enough lesions, to prove that
the treatments were a failure in killing the fungi within the tissue at a
temperature that would not injure the corm.

SOIL TREATMENTS

Experiment 1. Chemicals— Since the organisms causing the hard rot
and dry rot diseases are able to live over winter in the soil, the possibility

Tue Harp Rot DisEasE OF GLADIOLUS 179

suggested itself that some chemical might be applied to the soil which,
either through its toxicity or by its ability to change the composition
of the soil thereby rendering it unsuited for the existence of the pathogenes,
would serve to eradicate them. Healthy corms could then be planted
safely in this soil.

For the experiment in 1912 a plot of land was used on which gladioli
had been grown for the past three years. The chemicals used and the
amounts per acre were as follows: air-slaked lime, 1200 pounds; sulfur,
tooo pounds; air-slaked lime 800 pounds, and sulfur tooo pounds; sulfate
of iron, 1800 pounds; acid phosphate, 1200 pounds; acid phosphate, 2100
pounds. The chemicals were applied by the use of a lime spreader,
in strips of ro by 136 feet, a strip of equal width being left between each
of the treated areas to serve as a check. The entire experiment was
conducted in triplicate. Across the strips and at right angles to them
were planted the rows of corms, each row consisting of a single variety.
During the growing season special care was taken to see that no soil
was carried from one treated area into another or into the checks. The
results of the treatments were based on corms removed from a center
seven-feet strip of each of the treated and the check areas.

In the following January, when the results of this experiment were
recorded, it was found that none of the treatments had been of any value.
No reduction whatever was obtained in the amount of disease in treated
as compared with untreated corms. Since the chemicals were applied
in as large amounts as is commercially practicable, if not larger, no
further soil treatments with chemicals have been tried on a large scale.

Experiment 2. Formalin as a soil disinfectant.— Soil in which seedlings
had been grown for the past two years was treated in 1912 with one gallon
of one-per-cent formalin solution per square foot. The plot was covered
with heavy burlap for two days after being treated. As soon as the
odor of formaldehyde could no longer be detected, seeds were planted
in the treated soil, other seeds being planted in untreated soil to serve
as a check.

During the summer the hard rot disease appeared on the foliage of
these seedlings. No doubt infection occurred from spores blown from
diseased seedlings growing near by in untreated soil. The corms were
harvested in the autumn and stored in a cool room.

In the following January, when the corms were examined, it was found
that seventeen per cent of those grown in treated soil were diseased, while
thirty-seven per cent of the corms from untreated soil showed disease.
Since the disease appeared on the foliage of these plants during the summer,
it was impossible to determine whether the source of infection was the
mycelium of the fungus in the soil, or spores that might have been washed

180 BULLETIN 380

down from the leaves to the soil where they would germinate and infect
the corms. Therefore it was impossible to determine from this experiment
whether or not the formalin treatment had been of value as a soil dis-
infectant. The experiment was repeated in rors, but no results were
obtained because the seed planted failed to germinate.

Experiment 3. Formalin as a soil disinfectant— The value of formalin
as a soil disinfectant for Septoria Gladioli Passer. and the dry rot fungus
was further tested by treating soil in which gladioli had been grown for
the past two years with formalin at the rate of one gallon of one-per-cent
solution per square foot. Healthy corms were planted in this soil. No
lesions of the hard rot disease appeared on the foliage during the summer.
In the following January, when the corms were examined, it was found
that the treatment had proved of no value in reducing the percentage
of disease, as compared with healthy corms growing in untreated soil.
However, the treated plat was not sufficiently isolated from other untreated
areas to preclude the possibility that infected soil might have been carried
from untreated soil to that which was treated, and hence the results
must be considered with that restriction.

SANITATION

It has been shown that the hard rot fungus is able to live over winter
on dead tops left lying about on the ground. It follows that these tops
should be raked up in the fall and burned. This suggestion applies
particularly to the tops of seedlings and cormels, since the disease has
been observed by the writer to occur on the foliage of but six plants
of flowering size. It has also been indicated that the fungi causing the
hard rot and dry rot diseases of gladioli will live in the soil for at least
four years. Care should therefore be taken that the soil does not become
infected with the pathogenes. Only healthy corms should be planted
in soil which it is desired to keep free from these fungi; at least more
care should be exercised at planting time to see that no corms badly
diseased are planted. Such corms should be discarded and burned,
for they will but decay in the soil and infect it with the disease-producing
organisms. Crop rotation should be practiced.

BIBLIOGRAPHY

ALLESCHER, ANDREAS. Septoria Gladioli Passer. In Kryptogamen-
Flora von Deutschland, Oesterreich, und der Schweiz [Rabenhorst]
130. 780 Teor:

Cuixtps, J. L. Leaf blight of gladiolus. Florists’ ex. 23: 541... £007,

CRAWFORD, MATTHEW, and vAN FLEET, W. The gladiolus, p. 1-98. rort.

Tue Harp Rot DIsEASE OF GLADIOLUS 181

HatsTeD, Byron D. Fungous diseases of ornamental bulbous plants.
In Report of the Botanist. New Jersey Agr. Exp. Sta. Rept.
14:392-396. 18094.

Experiments with gladiolus. Ju Report of the Botanist. New
Jersey Agr. Exp. Sta. Rept. 17:398-399. 1897.

Experiments with ornamental plants. Jn Report of the
Botanist. New Jersey Agr. Exp. Sta. Rept. 18:334-339. 1808.

Experiments with ornamental plants. Jz Report of the
Botanist. New Jersey Agr. Exp. Sta. Rept. 19:324-327. 1809.

Experiments with ornamental plants. Ju Report of’ the
Botanist. New Jersey Agr. Exp. Sta. Rept. 20:408-409. 1900.

Experiments with ornamental plants. Ju Report of the
Botanist. New Jersey Agr. Exp. Sta. Rept. 21:461-462. 19071.

Henprickson, I. S$. Gladioli. American Glad. Soc. Bul. 3:5-13. tort.

Hicks, D. C. Notes upon the gladiolus and its culture. Vermont State
Hort. Soc. Rept. 4:32-36. (Part of the 27th Ann. Rept. Vermont
Biacesbd. \Acr:) /1907. =

Pax, F. Ixioideae-Gladioleae. Ju Die nattirlichen Pflanzenfamilien
[Engler und Prantl] 2:5:153-154. “18809.

Prititizux, E. E., and DELAcRoIx, G. Pathologie végétale. Maladies
bacillaires de divers végétaux. Acad. Sci. Paris. Compt. rend.
118:668-671. 1894.

Roprnson, W. Gladiolus. Jn The English flower garden and home
rounds, Pp. 575-578. 1883.

Saccarpo, P. A. Septoria Gladioli Passer. In Sylloge fungorum 3: 574.
1884.

WALLACE, ERRETT. Some bulb rots of gladioli. Thesis for degree of
M. S. in Agr., Cornell University, 1909. (Unpublished.)

Diseases of gladioli. Rural New-Yorker 69:355. 1910 (a).
Gladiolus bulb rots. Cooperative experiments. Gardening
£S:308. 1910 (b).

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