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TeCHNICAL BULLeTIN
No. 186
JULY,
1930
UNITED STATES DEPARTMENT OF AGRICULTURE WASHINGTON. D. C. THE BACTERIAL BLIGHT OF BEANS CAUSED BY BACTERIUM PHASEOLP By W. J.
ZAUMEYER 2
Assistant Pathologist, Office of Horticult-ural Orops alld Diseases, Bureau of
Pla·llt Industry
CONTENTS
Introduction____ . _______ .- __________ •______ _
History of the disoase________ •______________ _
I rost plants_________________________________ _
Distribution
and economic importance_____ __
Symptoms _________________________________
Moisture as a factor intIuencing infection __ _
Transmission of bacterial blight____________ _
Seed transmission ______________________ _
Overwintering on bean straw___________ _
[nscct transmission ______ .. ______________ _
De,,' as a factor in dissomlnation._. _____ _
Otl!er e!lYir.onmental factors Illfscting
dlSSOllll nation ____ .. ___ .... _________ .. _...... _
'rh~ pro~onk.ing 'Jf seed as u factor in
tlissonunntlOn. ___________ •___________ _
~
Page
1
2
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4
56
9
9
10
11
II
II
Relation of parasite to hosL_________________
Materials and methods__________________
Relation of parnsite to leaf tissne_________
Relation of parru:ito to stem tissue_______
disintegration
bacterial
Cell-wall
nction_ ______
_____ _______through
___ _______________
Helation of the parasite to pods and seeds_
Penetration of bacteria into the cotyledon_ __
Varietal resistance___________________________
:>[ethods ________ ______ ______________ ____
\"arietal tests____________________________
Summary_______.___________________________
r,iternture cited_____________________________
Page
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33
34
13
INTRODUCTION
Since the intensive culture of any crop results in the introduction
and increase of infectious diseases, a general survey of the occur­
rence of bean maladies was undertaken in 'Visconsin. From ob­
servations made it was evident that the principal diseafjes which oc­
cur in that State are the bacterial blights, caused by the widespread
Baotel'i:u1n phaseoli EFS. and Bact. 1nedicaginis val'. phaseolieola
Burk. and the relatively unimportant BMt. flaeaumfaeiens Hedges.
Anthracnose caused by Oolletot?'iclLu7n lindernuthiamurn (Sacc. et
Magn.) B. et C. and a fungous root rot caused by F1t8ariwln ·martii
phaseoZi Burk. are of little consequence except in certain years when
conditions are extremely fa\"orable for their development. Mosaic
is of common occurrence and often causes severe damage, particu­
larly to the Refugee varieties.
1 Presented in partinl fulfillment of the requirements for the degree of doctor of philosophy at the Univer­
sity of Wisconsin.
, The writer is indebted to T" R. Jones, at whose sug~estion the problem was undertaken, for the advice
and encouragement he !(nye throughout the course of the inYestigation; to J. C. Walker for mnny helpful
suggestions nnd criticisllls olfered during the progress of the work; and to L. L_ Harter and Florence Hedges
for C!lreful rending of the Illonnscri!,t.
106267-30-1
~t.p
12 1930
f ,
2
TECHNICAL BULLETIN 186, U. S. DEPT. OF AGRICULTURE
Because of its almost unh'ersal presence and the great loss it
occasions, 11. study of Bac{el'ilNn phaseol; was initiated, This or­
ganism anel the cli::ease it produces were described more than 30 years
ago. It was not until r('cl'ntly, howevcr, that bean blight was
separated into three dj~tinct bacterial diseases, as noted above.
Since tlH'Y no\\' oN'lIr mere or le~s illtermingled alld product' symp­
toms that on~rlap in tht'ir characteristics. it i:-: probable that in the
past thl''}' were ail gt'ollpec1 with the common beall hlight. caused by
Bact, lJhaseoli. Pl'e\'iolls investigators in working with Bact.
phwseori haY(' given chief attention to the bacteriological characters
of the pathogene. It is the purpose of this bulletin to re\Tiew briefly
the known fenturcs of thc disease :md to give a detailed account of
the writer's investigations dl'aling with the environmental Iactors
that influence dissemination, inIection, and "arietal resistance. and
especially with the relationship OI the parasite to the host.
.
HISTORY OF THE DISEASE
The disease of beans callfjed by Bacte7'iwln phaMoli is probably
of American origin. It was reported by Halsted (15L3 who stated
that the disease was first u""'lght to his attentioll uy a large ~eed
l'ompany in Xew .Jersey. Later, !:'imith (.J2) i~olated. named. and
described the organism. Delacroix (9) recorded a disease of beans
near Paris that may have been the same as that described by Smith.
.:\. further account OI the cultural characters of the orO'anism was
published in a second paper by Smith in 1901 (33). fn the same
year Halsted (16) descrihed the field symptoms and experimented
with possible preventive measures such as irrigation, shading, sprin­
kling, and crop rotation. Sackett (30) and Whetzel (39) each pub­
lished a brief account of the disease. Fulton (13) described the
field characteristics of the disease and discussed measures of control.
He experimented with the hot-water seed treatment and spraying,
and tested the comparntive resistance and susceptibility of a num­
ber of yarietie:;. E(lgerton and )[oreland (11) published an account
of bacterial blight from the standpoint of the resistance of the or­
ganism to drying. 'rltey experimented with seed treatment and
concluded that benetol and corrosive sublimate had given good re­
sults. They suggested also the use of home-grown seed.
That the organism can Ii\'(> over winter in diseased bean trash
ancl may also be carried to clean fields in manure was shown by
Muncie ·(125). His seed treatments with chemical solutions and with
moist and dry heat failed to gi\'e results. He recommended the use
of clean seed obtained' by pod selection. In a general surveyor
the disease Rapp (i!S) reportecl that seed 2 to 3 years old produced
blight-free plants. Seed treatments with chemicals and hot water
he found to be impracticable.
The vascular nature OI the disease was first reported by Barss (13),
who found by internal microscopic examination that the xylem
H'ssels of the affected plants were filled with bacteria not only toward
the base uut u~ually throughout the entire stem and even out into
the branches, petioles, and veins. He likewise traced the bacteria
'Italic numbers in parentheses refer to Literature Cited, p. 34.
BAOTERIAl, BLIGHT OF BEANS
3
ill lIlillly in~tnll(,l'~ 1'1'0111 the xylem vess('ls of the main stem to the
suture of the pod. He showed that the bacteria entered through
thc vascular system of the seed ,yithout producing any outward symp­
toms. He stated that the death of seedlings e:1I1 be aceounted for by
in\'H81011 illto the vnseulnr systelll frolll illfcdcli eotyledons.
Coincidently with Banis, llmkhol<.1eI' (5) likewisf:' reported the
blight of b(,Hns as a systemic disease alll1 showed that the organism
produced wilt symptoms by plllgging the yessels. He also statell
thnt since the disease is of a. syst('.mic nahn-e, the seeds may become
infected withollt producing lesiOlls 011 the pods, a fact which is of
considerable importancc from the standpoint of control.
A eomparisoll of the eau,w of SOybCUIl pnstule, Bacte)'iwn plw..seoli
sojell~e Hedges. with Bad. p/laseoli waS madc by Hedgcs (18) 1
who worked pl'illl'ipally with the cultlll'al dllll'ucteristics of both
OI·gUlljSll1~.
Hallds und Brotherton (27) cxpel'imentcc! with se\'cml of the
beall diseases und mac1~ tests for disease resistance with
liO va l'ieties of IJea ns fl'om the U nitec1 States and 493 varieties ob­
tai lied :from 23 fOI'eign conntries.
A Ilew blleterinl diseasl' of l)('ans enused Ly lJaeteJ'iwn flaccum­
jacit.:ns was replnted by Hedges (10), who named and desCl'ibcd the
organism and made Ii thOl'ough study of the cultural characteristics in
comparison with Bact. plu~eoli.
Burkholder (7') described a new bean disease caused by Bacterium,
1I1edicaginis yar. plwseolico/a. itS being widespread in New York. He
named and deserlbed the OI'gnnism and gave the general characters
of the disease in cOlllparison with those cuusec1 by Bact. plul-seoli and
Bact. f!acc'l.lm!acie)/8.
A report on the sel'Ologicul differentiation of Bacterium campestre
EFS., Bad. jlaccumfaciells, Bact. l)lw8eoli, and Bact. plw,8eoli sojense
wus published by Link and Sharp (22). They showed that these
four pnthogent's could be differentiated by agglutination tests and
that serologically Jjact. cam pestrc, although distinct from Baat.
plia8eo/ i and Bact. pll([,~eol i sojel/8e, was closely related to them and
mOre r('motely I'l.'latt'(l to B(lct. flacC'umjacie1l8. Sharp (31) reported
on ll1orphologieai, physiological, and sel'Ologicrl studies together with
virulence and acid agglutination studies of Bact. flac(JUmfaei.ens,
Bact. plw8C'o/i. and Bad. plwseoli 80jense and concluded that these
three species nil c1ifl'('I' and (':til be diff(,l'cntiated by the use of the
agglutination tN·t. Bact. flacr':llmfaciens serologically stands apart
from Bact. p!t(l,~eoli and Bact. pll.aseoli 8ojense, which are very
closely related.
,'omHlOll
HOST PLANTS
It is f[C'ncl'ally recognized that practically all the commercial va­
deties of the common bean, Plwseolus 1Y!uga1is L., are susceptible to
the bacterial blight caused by Bacte1'iwm pluuJeoli. The studies
of other ,,"orkel's have shown that the following are also hosts of
the org:mism: The Scadet Runner, P. coccineus L.; the civet bean,
P. lWlatus L.; the I.-imn. bean, P. lunat1l8 val'. 'nuwrocarpU8 Renth.;
the pinto bean, a variety of P. vulgmis/ the white tepary bean, P.
acutifolius latifoUU8 Gra),; the moth bean, P. aconitifolius Jacq.;
the adzuki bean, P. angulari8 (Willd.) W. F. 'Wight; the urd benn,
4
TEOHNICAL BULLETIN 186, U. S. DEPT. OF AGRIOULTURE
P. mJ/tngo L.; and the black-eyed cowpea, Vigna sinensis (L.) Endl.
Gardner (14) has reported that the trailing wild bean, St'l'oplwstyles
hel!vola (L.) Britton, is a wild host. In the writer's inoculation
work it was observed that the hyacinth-bean, Dol:iclws lahl.o;b L., is
also susceptible.
DISTRIBUTION AND ECONOMIC IMPORTANCE
The geogmphical distribution of bacterial blight is not completely
known. It probably occurs in every State in the Union, but little is
known about its occurrence in other parts of the world. Delacroix
(9) reported what appears to have been the same disease from
France, and Ideta (120) reported it from Japan in 1903. It was also
reported from the Philippine Islands by Reinking (!B9) in 1919,
and from South Africa in the same year by Doidge (10). Skoric,
formerly of the laboratory of plant pathology, University of Wis­
consin, in conversation with the writer, stated that the same disease
is widespread in Yugoslavia.
Next to anthracnose, bacterial blight is the most important disease
of beans. According to the Plant Disease Reporter,4 it was reported
from New York in 1918 that the blight was prevalent in 75 per cent
of the bean fields and caused very serious damage. In 1919 the
disease cansed an extremely high loss throughout the bean-growing
districts. In that year Colorado suffered a loss of from 40 to 60 per
cent of the crop, with It decrease in yield of about 35 per cent. In
New York, which produced about 1,660,000 bushels of dry beans in
1918, or nearly one-eleventh of the total yield in the United States,
it was estimated that from 5 to 10 per cent, with a possible average
of 7 per cent, or Itbout 125,000 bushels, WitS lost through the disease.
In 1921 this disease continued to be a very serious menaCe to the
bean industry. It was prevalent over the entire eastern United
States except in Vermont and New Ham.pshire. The most severe
loss was in Michigan, estimated to be 25 per cent of the crop. In
1922 the total loss throughout the United States was about 10 pel
cent, and the same was true in 1923. In 1924 New York reported
serious losses, even greater than in the previous year, and a reduction
in yi.eld estimated at 10 per cent. In 1925 the largest losses were
reported from New York, Louisiana, Indiana, and Iowa. Reduc­
tion in yield averaged about 15 per cent. The damage in 1926,
caused by the bacterial blight, was about the same as in the pre­
vious year, New York reporting the largest loss (10 to 15 per cent
damage), while New Jersey, Ohio, Colorado, and Arizona all re­
ported losses ranging from 5 per cent and lower.
In 1927 reports again showed serious losses from the blight,
Indiana and Louisiana reporting losses of 4 per ceni; of the crop;
Michigan and Montana, 3 per cent; Connecticut, Maryland, Virginia,
Wisconsin, Minnesota, Mississippi, and Texas, 1 to 1.5 per cent.
Linford, reporting from Utah, estimated a killing of from 5 to 95
per cent of the plants in numerous fields. The average loss through­
• l]:-;ITEO STATES DEPART~IENT OF AGRICULTURE, BUREAU OF PU:-;T !:-;OU3TRY.
PLANT
nrSEASFJ SURVEY nur.r.ETIN. v. 2. 1918; v. 3 1919; v. 4, 1920; v. 5. 1921; v. 6. 1922;
v. I, 1\)23; v. 8, 1924; v. 9, 1925; v. 10,
[Mlmcogl·npilcd.]
i 926;
\'. 11, 1927; \'. 12, 1928.
1918,1928.
5
BAOTEmAL BLIGHT OF BEANS
out the United States was estimated at 1.4 pI'!' cent, or a total of
approximately 226,000 bushels of beans. 5
SYMPTOMS
Although the symptoms of bacterial blight have been accurately
described by several investigators, there is included here a descrip­
tion of its appearance as observed on different parts of the bean plant
at various stnges of maturity.
Pr'obaoly the most striking evidence of the disease is on the leaves.
Here the lesions first appear on the lower side us ~mall, water-soaked
spots in the center of which, as they develop, a slight incrustntion
of bacterial exudate is immel. The lesion is surrounded by a yel­
lowish 1111101ike zone. These lesions gradually enlarge and may
coalesce with others, producing a brown scaldedlike area which in
time ea,uses a de foliation of the plant.
Bacteria from diseased seeds often produce ver'y peculiar lesions
on the first primary leaves. Here large angular water-soaked areas
appeal' on the opposite leaves at si mi1ar positions, indicating that
the initial infection took place while the leaves were still folded
between the cotyledons. These differ from the small water-soaked
circular secondary lesions in that they al'c larger and decidedly
:lngular·. On the yOllllg trifoliolate l(,Hves a wry unusual symptom
appears in many cases when infection is severe, the bacteria entering
the smltll veinlets, enusing a slight discoloration of the adjacent tis­
sues and a retardation 0-[ development in this region. 'With a. rapid
growth ,in the 1lllinfe~ted areas. a pllckered appparanee. \"C'ry similar
to mosalC symptoms, 1S prodnced.
'Vhl'lL the hactel'ia are found in the vaselllar tissues of the leaf,
another chamcteristic symptom can be seen. (PI. 1, C.) The
infe('tion uS\lally begins in the small veinlets, subsequently inyolving
the larger veins and finally the midrib. In the case of severe infec­
tion the bacteria IH'oclllce a"'Nldish dis('oloration with u water-soaking
of the ti~s\les immediately slH'l'ounding the wins. 'Vhen the leaf
illic('tion starts i'l'om the pctiole, the main vein and its hranches
fin;t appcal' water'-soaked, later taking on a brick-rcd discoloration.
'Vhell aisensed sceds nre planted they may produce seedlings that
('xhibit :t'characteristic wilting in the case of severe infection. The
first macroscopic appearance is a slight flagging 01' drooping of the
leaves at the region of the pulvinlls. (PI. 1, B.) From isolation
lind microscopic observation of such tissues bacteria nrc generally
found in large masses. During the night such leavcs appear quite
normal and turgid, but during the heat of the clay they again become
flaccid. In more acIYanced stages this drooping is followed by a
wilting that in some CHses may involve the entire seedling, whereas
in others only a portion of the plant mlty be affected.
On tire stem the pathogene may canse yarious types of lesions.
The young seedling- lesion begins ns a small water-soaked spot which
g'mcluaUy enlarges. appenring somewhat similar to pod lesions.
The ncel'otil' areas are som(·times sunken [mel later appear as red­
dish dashes, extendin~ longitudinally along the stem. (PI. 1, E.)
• UNIT~D &r.\TES DEI'AUTMm;T OF AGRICULTURE, BUIIE.\U OF PLANT INDUSTIIY.
Op. cit.
6
TEOHNICAL l~t'LLETfX 1 S 6, U. S. DEPT. OJ!' AGUICVLTUUE
The surface of these spots is often split (pt. 1, B), and baderial
exudate can sometimes be seen oozinf! ft'olll the lesions (pi. 1. A).
'When the plants al'e ill the initial stages of pod formation, It
lesion known liS stem girdle or joint rot appears (pI. 1~ D), which js
first manifest as a small watee-soaked spot either at the cotyledonary
node or at other nodes along the stem. Upon enlargement, the lesion
{i,nally encircles the stem. The girdling is usually completed a~ the
tune when the pods are half mature, and the affected plant IS so
wel1ken{'~l by the increasing weight of the top that the stem breal{s
at the dlsensed llode.
Bacterinl Llight causes much damage to the poels, The initial
lesions fit':;t appear as minute water-soakeel areas which gradu!tlly
enlarge. accompnnied by It discoloration nnd distinct 7.oning. Later
the spot become'S dry and sunken and takes on a brick-red eolol'.
Often a yellowish-white incl'lIstntion of baetel'iat pxuilate can be
Seen eovcl:ing the lesion. (PI. 1, F.)
The bacteria may also infect the pod by way of the vasculnr cle­
ments, Following the dorsal suture, they Clluse a discoloration of
this region and n water-soaking of the surrounding tissue. CPt 1, I.)
In n like mal1lH.'l' the organism may attack the seeds, producing sev­
ernl types of lesions, 'When infection occurs while. the pods are
young the secds may rot entirely, or they may become 80 8m'erely
infccte(l that only the shriveled seed coat remains. On the other
hand, the bactl'r.i!l enterinf! by way of the funiculus may cnU8e a dis­
colol'l1tioll nt til(' hilulll. (PI. 1. G and H.) On dark-seeded varie­
tip;;; these discolorations are oitell difHcult to detect, but on light­
sl't'cled Yarictil'S they arc very noticeable. (PI. 1, G.) On light­
eolored seeds when the infection is severe the lesions eoYer a con­
si(kl'nhle area. and have a varnishlike appearance.
i\lOIS'l'URE AS A FACTOR INFLUENCING
INFECTIO~
It is generally agreed by various investigators that the chief ex­
ternal faetors infinencing stomatalmo\'ement aI:c light and tempera­
ture. :4oIl1e belicyc that humidity greatly affects the degree of stom­
ahtl openings]., wlwreas others consider it of only minor importance.
,Yilson nnd \treenman (40) founel that the stomata, on plants of
J[elil()tH,~ aTh(£ L, thnt \\'el'l' left in a, satuL'llted atmo:-;phere -were well
open, but the stomata of the plants that remained in the dder open
an' (lxposed to approximately the same light were nearly aU closed.
Darwin (8) showed that stomata, werc very sensitive to changes in
humidity. dosing when taken from a high to a low humidity and
opening' tmder the opposite conditions. Lloyd (:2.3) believes that
there is but little evidence to show that a high relative humidity,
f:\\'oni the wide opening of the stomata, in the ocotillo, and in
JJfcntlla pipeJ'itc6 L., also ·a desert plant. Poole and :McKay (26), on
the otlwr hand, believe that while light may be considered a funda­
mental factor in the movement of the stomata of the beet, yet stoma­
tal closure is affected by low relative humidity even though the light
is active.
Difficulty has Gften been experienced in obtaining stomatal pene­
tration in the greenhollse with bactet'ial plant pathogenes when the
inoculnted plants were not placed un.der conditions of high humidity_
BACTERIAL BLIGHT OF BEANS
7
The ncc~ssitv of stich conditioll!> for infection with Baetaluln
plwseoli hns been known for some time. Smith (35) in working with
this and other bacterial plant pathogcl1l's always subjected the host
plants used to a moistlll'C treatment pt'evious to inoculation, in ordel'
to be insured o.f good infection. 'Vith this in mind, inoculation ex­
pCI'iments were performed by the writer to determine, if possible, the
relationship of moisture to stomatal mOW'lI1ent in bean It'aves. The
principnl varieties used Jor this -lVork were "'anlwell Kidney "'ax
and -Wells Red Kidney, both of which nre \'ery susceptible to the
blight.
'
'fhe cnltme8 of Ba.ctcl'itl1n plw,'tcoli used .for inoculntion 'were re­
ceived from It number of sourees. Two ('Hme from the United States
Department of AgricuJtl\l'e, 'VashingtoIl, D. C.; one from J. H.
Muncie, Michigan Agdcultural Expcrilllcllt Station; one ,,,ns iso­
lated by the writer from beans O'atherec1 at Racine l 1Vis.; and one was
isolated .from material coilectetl at Madison, 1Vis. These cultures aU
gave similar results .and will tilN'efol'c not be considered separately in
the following discussion. A culture of Bact. 1ncdicaginis vat'.
pl~aseolioola isolated frolll beans collected at Columbus, 'Vis., was
11180 used. The bacterin, were allowed to grow on potato-dextrose
agal' slants (pH 7) for it period of about a week at :), tC'mpemtul'c
or 28° C. They were then l'emo\'ed by washing tht:1Il into sterile
atomizen; cont.atning sterile distilled writel'~ nnd this suspension was
used as the inoculul1I.
Since the pathogenicity of Bactl''1'ill1n phaBeo7i has beC'n del11on­
strated by other inwstigatol's, results on this phasc of the problem
wi It not be reconled here.
'rho plants fOI' inoclllation purposes w('t'e ttTOWn in 4-inch pots
until they rcadH'tl a height of about 8 incl1C's. These seetHings were
gronped into thre(' lots. The first lot was ('on~recl completely with
U glass ('ontnil1C'r in 01'([('1' to protiuC'e a saturated al:ll1osphet·c about
tho plant and WIiS allowt'd to remain in place Jor :t period of 2-:1: 1Ioms,
dtet· which the plan/::; \\'('1'e spmyed with a :;uRpcnsion of the bean­
blight ol'p:nni:-;Ill and again <':oH'I'C'd for :2·111OIIrs. The s('('onc1 series
of plants WtlS not gin'n a pr('ll1oist trt'atment as wa~ the first lot,
but was otherwise treated the same. The plants werc covered
nfter inoculation. The third set was illoculatNl without a premoi8t
treatment Ot' a co\'cl'ing after inoculntion. All the plants in the
three series werc then placed under similar conditions ]n a green­
house. The plants Wl're examined daily for water-soaked lesions.
Both of the organisms lIsed in these tcsts, Bactel'i'Uln lJlwseoZi and
Bact. /I1cdica.r;inis val'. plwseolicola., pt'oduced numerous water-soaked
lesions, and in eHch 501'](>8 plnti11gs were mnde in order to be positive
that the lesions wel'e being caused by the respective pathogenes used.
The J'esults of these l'xpcl'iments iIl'e given in Table 1.
8
TECHNIOAL BULI,ETIN 186, U. S. DEPT. OF AGRIOULTUR~
of infeotioll causcd bV Bu·c/.('ri.uln IJhu.scoli (11HZ Bact. medica­
Yinis va,.. ph(lscolicola on bcan lJianls 8//lijcelell 10 1II0il<ttlrc trc(/tm.l'nl~ of
va"i(}l~s d.w·a·lion
1'ABLE I.-Dcyrce
[Tho let.tors II, IJ, nnd C cHeh fllprl'sents n single inoculutioll experin","t)
Degree of illfection
'I'rentmollt
Dnct. phnseoll
1. Covcrl'd ror ~H huurs, inoctllntNf, and c()\'crcd (or 2.t 'lours • 1
In, JJ,\:n.\:~~~ __________________
·ll>, 1l~1I\ ~ •••.•••
- •••••.••..••
___________________
(', SCVl\~n.
Dnct. 1llodicaginls
Vllr. phasoolicoln
a, ll~'nv.y.
b, .1Iell\
y.
c,
lIl'IlVY.
II, MedIum.................. }
2. Inoculnted lind covcn,tl for 24 hours................. { c,
b, lIellVy....................
___________.. _______ _ :\ot Pl'fforlllcc\.
il, \'l'ry little ____............ II, V(lrV little. 3. Inoculuted nnd not covcred .................... _____ b, Very littlo __•• _...... _.... b, Vcr)' little
c, Very littlc...__....... _••• c, Very liltle. ll~uvy
t
Little difliculty was experienced in o\.Jtaining stomatal penetl'lltion
with Bacte'riu7n l1wdicag-inis val'. phaseolicola. Numerous water­
soaked lesions were obtained in one case without placing the inocu­
lated plants in a moist chamber; however, the greenhouse in which
the plants were kept was rnaintnined at a high tempeL'·ature and
hUlllldity. ",Vhcn inOclllutions were made under controlled moisture
conditions ior 24 hours numerolls infections were always in evidence.
Burkholder (7) was unable to obtain stomatal penetmtion, even
though his plants were placed in :l moist duunber for 12 hours a.iter
they were inoculated. He,statcd, however, that the disease was wide­
spl'cad in the field, showing particularly after moist wcather, which
seems to indicate that stomatal penetration took place. It is difficult
to explain such n, widespread occurrence of the disense if the organ­
ism enters only through wounds.
It is evident from Table 1 that in those instances where plants re­
ceived It great amount of moisture before and after inoculation a
high percentage of infectioll resulted, whereas a decidedly lower
umOl!lIt WIiS observed when the plunts were not given the moist tt'cat­
nwnt. Since covering produced a saturated humidity about the
plant, It film of moisture forl1led on the exterior of the leaves and
tho substomatal cu.vities probably becnme well supplied with water.
It. seems reasonable to supposn that ii moisture is a factor in influ­
encing the movement of thc stomata, there may have been set up
n continuolls passage of water from the exteriol' to the interior of
the leaf; e. g., to the substomata.l Cll "ity. Thus, since the plants had
recei\'cd a pre\·jow; moist treatment by being co\'ered 24 hours before
inoculation, the bacterilt may have had a free swimming channel
from tho droplets of moisture that collected on the surface to the
interior of the leaf and thus pl'OdllCecl infection.
These experimC'nts iIHlicntC' that moisture is au important factor
in favol'ing the production of disease. In Group 1, in which a high
amount of moisture was present both before and after inoculation,
the amount of infection ranged from very heavy to severe, whereas
in Group 2, where abundant moisture was supplied only after inocu­
lation, the amount of inf('clioll \HIS sOl1wwhat reduced. In Group 3,
whet'e little moisture was present-that is, only that which was pres­
ent in the greenhouse-very little infection was noted. These experi­
BACTERIAL BLIGHT OF BEANS
9
ments were repeated several times with a variety of plants and bac­
terial pathogenes, with similar results, indicating that moisture seems
to be an impodant factor in producing infection by first 1nfluencing
the movement of the stomata, which in tUt'll allows a channel for the
bacteria to make their way into the interior of the leaf. 'l'hese
experiments were performed at 10 a. m. and 5 p. m., with comparable
results.
TRANSMISSION OF BACTERIAL BLIGHT
Considerable attention has been given to the problem of dissemina­
tion of bacterial blight because of the possible bearing it might have
on control measures. Most of the evidence presented is observa­
tional, having been gathered for the last three years from experi­
mental plots at Mndison, ·Wis., from bean fields' throughout that
State, and from certain of the bean-producing al'eas of the western
United States.
.
SEED TRANSMISSION
II; has been known for a long time that bacterial blight is carried
from year to year in and on the seed. In this manner the dis­
ease spreads into districts that had previously been fLee from the
malady. Beach (3) reported that probably the disease wintered over
in the seed. Halsted (16) also inferred that infected seed trans­
mitted the disease from one year to the next.
Both in the laboratory and in the field, little difliculty has been
experienced in proving the existence of bacteria in the seed. The
bacteria are harbored in the seed coats and also about and between
the cotyledons, and when germination takes place they may either
caUE'e the death of the young hypocotyl before emergence or enter
th~ cotyledons, causing vascubr invasion of the plant. Cotyledonary
lesIOns and the large water-soaked areas that appear on the first
simple leaves and stems may serve as the initial sources of the
secondary spread of the disease in the field.
During the summer of 1927 a study was conducted on the develop­
ment of the disease nnder field cond.itions. For this purpose diseased
seeds, alternated with healthy ones, were planted in several rows.
'Vhen the bean plants were still small it was not a difficult matter to
distinguish plants tl:at were grown from diseased seed. Usually the
plants were more spindling than normal ones, the cotyledons dried
and dropped off before those of the healthy plants, and in many in­
stances large angular water-soaked spots were to be seen on the first
simple leaves. After a short period, certain of the plants surround­
ing such an infected seedling showed secondary lesions on their
leaves. In the early part of the growing period only plants in the
same drill row manifested these lesions, but later as the leaves en­
larged the infection passed from row to row. It is evident that from
the single infected plants serving' as sources of inoculum the blight
pathogene may spread very easily and cause severe destruction of a
crop.
:Most of the severely infected seeds are gleaned out by hand after
threshing. It is difficult, however, to detect the slightly infected
ones, since little shriveling or discoloration of the seed can be seen.
'l'hese seeds when planted produce seedlings that serve as the initial
sources for much of the secondary spread of the disease in the field.
OVCl'
100267-30---2
10
TEOHNIOAL BULLETIN 186, U. S. DEPT. OF AGRIOULTUHE
Exudate is often seen oozing from stem or leaf lesions when condi­
tions of high humidity prevail, as often exist during the early part
of the growing season. From such sources rapid and widespread
blight dissemination may take place.
OVERWINTEIUNG ON BEAN STRAW
It was suggested by eady workers that the disease may live over
from year to year on infected vines and pods. Harrison and Barlow
(17) stated that the bacteria can live over at least one winter in
stems and leayes allowed to remain on the gt·olmd. l\:[cCready (134.),
without producing conclusi \'e proof. stated. that "the disease is
carried over from year to year in the seed from a diseased crop, in
the soil on which a. diseased crop has been gL'Own, or in straw fL'Om
infected fields, in bedding Ot· nHlIlUL'e." Muncie's (e5) observations
are that the disease o\'erwintel's on diseased straw, and his experi­
ments tend to prove this point, for the organism was isolated from
diseased bean stubble which had remained in the field oyer winter.
The writer has gathered data from field studies which tend to
lend eyidence that the organi$m may live over the winter in this
manner. 'l'here are, however, no experimental data to substantiate
these observations. In one of the large bean-growing districts of
'Wisconsin serious outbreaks of the bacterial blight were in evidence
during the summer of 1926. 'Vardwell Kidney 'Vax, one of the
most susceptible of the commercial canning Yarieties, was being
grown to a large extent that year. In a single field of about 8 acres
practically 100 per cent of blight was estimated. After the bacteria
had killecl these plants cattle were allowed to feed on the diseased
vines and the remaining stubble was plowed under. The following
year the Improved Kidney Wax, a variety slightly less susceptible
to bacterial blight than 'Wardwell Kidney 'Wax, was planted in this
field about May 25, and on August 4 considerable blight infection
began to appear. This circumstance would not have stimulated any
thought of the overwintering of the organism if the other bean fields
in the vicinity had shown any blig-ht symptoms. It would naturally
11llYe been concluded that the infection was caused from diseased
seed. Since, howeyer, none of the other fields of this variety showed
signs of the blight, although the seed was obtained from the same
SOUl"ce, It was suspected that the bacteria might have overwintered
on the diseascd stllbble and were then transmitted to beans planted
in the same field the following year.
Similar obseL"Yations were I'ecorded in a number of the western
bean-growing sections during the summer of 1928. During the sum­
mer of 1927 a large field of Full Measure beans was entirely de­
stroyed by the bactel'ial blight, and ab?ut September 1 thE clisea~ed
refuse was plowed under. In the sprmg of 1928 a large plantmg
of beans was made in this same field and practically every variety,
arown was severely infected by August 1, and by the end of the
~lonth very few individuals could be found free from blight. High
winds accompanied by rain and hail possibly may ha,'e accounted
for some of the infection. But. since there were no bean fields in the
immediate vicinity of the trials, and since the seed was grown in a
section free from blight in 1927, it was suspected that much of the
infection came from the diseased refuse that had been plowed under
the previous fall.
BACTERIAL BLIGHT OF BEANS
11'
INSECT TRANSMISSION
'"\Vhethet· or llot insects transmit bacterial blight from plant to
plant is unknown, but undoubtedly they play some part in the dis­
Heminatioll of th.e disease. Insects as spreaders of the blight have
been reported by It number of workers. Sackett (SO, p. :21;3) stated
thnt "insects play an important part in disseminating the trouble,
consequently uny measures which tend to check these pests will aid
in controlling bacteriosis." There is, however, no experimentul evi­
del1ee to substantiate these statements. In the present work the leaf
hoppers (E1npO(h~('a mali I.e B.), the 12-spottecl cucumber beetle
(Diab1'otica, duodec:i'l7lf[fu.nctata Oliv.), and a lttdyuircl beetle (species
not identified) ha,ye been partiCUlarly noticeable feeding 011 the
foliage of bean leaves.
It is possible that insects may cllrry the bacteria, on their legs,
bodieI', and mouth parts und so become factors in disseminating the
clispaHl'. At best they Can not be considered as playing a, major role
in the disl'l'mination of the disease, since its natur!!} spread during
favorable wcathel' conditions is of primary importance.
DEW AS A FACTOR IN DISSEMINATION
Dew as II possible factor in the dissemination of bacterial blight
was reported as early as 1901 by Habted (lU, p. 15), who stated that
" it is not unlikely that the germs were ('llrl'ied from the diseased
leaves to the pods by tlw dripping dews." Sackett (30) stated that
" ruin nnd dew nre doubtless agcnts in spI'efldin~ the germs from one
part of the plant to anothcr by wnshing them fl'om old lesions onto
unaif<'ded palts." That moisture is essential for -widesprellli infec­
tion beeanl(' t'vident ill both greenhollse and field studies. Heavy
dews (h'ipping from lca-f to leaf may easily carry the pathogene and
cause secondary spread of the blight. :L\Iany of the pod lesions found
along the dorsal suture nUt)" be clltu;ecl in' this same manner. Dew
which collcds in droplets may run down the petiole, thence along
the dorsal sutlll'e. cal'l'yin,g with it the bacteria that producc .in many
cases Ihe chamctel'ist;c yasculat· pod lesions. If the infection be­
COIl1CS (lstablished along the suture of the. yOllng pod, deVelopmental
!!l'o\\'th often ccases and the pod shrivels and dies. 'VlH'n infection
t,akes place after the beans have fOl'l:ned, ~hey may easily become in­
fcctt'd I:hrollJ!h the vasculat· connectIOn of the seeds to the poel. nnd
the pathogene CtIll be carried oYer until the follo\dng yeul' in this
IlHlIll1t'I·.
'"\Vhil(' tIl(' ]('a"e:: are \Yl't with ell'\\' til(' pathogene may sprctHl from
leaf to leaf if a thin film of \Yatel' connccts portiol1s of tIl(' two
Jean'l'. It is ror this reason that beans ::holllc1 not be picked Ol' culti­
vated eady in the morning or directly after a rain while they arc
still ('oYl'I'('(l with moisturl'. If this is done, infection can be spread
fl'ol1l plant to p1unt in the ::ame row and cven fl'om row to I'OW. in
SOI11t' case" cHusing (kstrlletion of the crop.
'
OTHER ENVIRONMENTAL FACTORS AFFECTING DlSSEJlIINATION
That rain might be a possible fnctor .in the dissemination of the
blight was reported by many investigators. Rapp (f38) states that
following a min accompanied by wind, bactel'ial blight spreads from
12
'rBCHNICAL BULLETIN 186, U. S. DEPT. OF AGlllCULTURE
the center of primary infection to the greatest extent in a southeast­
erly direction. This, he says, is accounted for by the fact that wind­
driven rain is blown in that direction and undoubtedly carries the
pathogene from row to 1'01Y, a.nd in some cases across a number of
rows. Observations made by the writer also indicate that splashing
rains accompanied by winds are responsible for a great deal of the
infection found in bean fields.
HlLil is also very important in disseminating the disease from plant
to plunt. This was well demonstrated in many bean fields of the
western bean-growing States. In the yicillity of the Greeley, Colo.,
project, much hail injury to crops was reported. Certain bean fields
that were stL'uck by hail showecl almost complete destruction, much
of the injury being caused by bacterial-blight infection. The dis­
semination apparently began from seedling-infected plants that sup­
l)lied tho SOHrce of inoculum. Because of the whipping of the leaves
and the injmy of the plants from the hail, the spread of the organ­
isms was vcry rapid. A short time thereafter practically all the
plants in the field were infected. Since comparable bean seed lots,
planted in regions of hail injury and also in hail-free sections,
showed a very decided difference in the amonnt of disease, it incli­
cated that the hajJ and wjnd were the limWng factors in the clissemi­
nation of the disease. Fields well protected by trees, preventing to a
great extent the whipping of lerwes, also showed less disease than
bean fields that were subject to high winds.
Some obsernltional eridence has been accumulated regarding sur­
face watel' as a means of spreading the bean-blight pathogene from
diseased to healthy plants. These data were collected in 11)28 at
the bean trial grounds at Madison, 'Vis., where a portion of the field
is sloping. During the early part of the growing season seedling
infection neal' thc 1I pper portion of the field was recorded. After a
series of hea "Y raim; the plots were again yisited, and the disease was
mOl'e ,ddrBprcnd. the direction of the spread being in many cases
from old d ist'tlsed centers. There was a tendency for the spread to
he in the downward direction of the slope, either along the row 01'
aero,;s the rows. In tracing these new infections it was observed that
the pathogene had been washed from the diseased seedlings and car­
l'ied down the small rivulets. 'Where this ,vater laden with the bac­
teritl e:unc in contact with the plants, ne,\" infections took place. At
least nino such j I1stances were recorded. indicating that sllrface­
c1:ainagc water very likely was responsible for carrying the organ­
isms from diseased seedlings to healthy plants.
That irrigation watet·, which is used almost exclusively in the
westel'll bean-growjng sections, may dissemjnate bacterial infection,
came to the writer's attention on several occasions in a suney of
many of. those scetions. It seems highly probable that the organisms
may' be C'arriecl down the small irrigation ditches and cause infection
to 'othet' plants in the same row. 'Where infected seed had been
plnnted, young lesions often extend to the ground level, and in many
instances bacterial oOze has been seen exuding from such necrotic
oreas. Ir6gation water in snch cases may carry the pathogene from
lesions to healthy plants in close proximity to the center of initial
infection. Diseased leaves that had ch'opped from infected bean
plants wno often seen in these ditches. In this mnnner the organisms
BACTERIAL BLIGHT OF
BEA~S
13
could also be carried down an irrigation ditch, causing plants along
the row to become infected.
TilE PRESOAKING OF SEED AS A FACTOR IN DISSEMINATION
That wuter applied to seed in the inoculation with Bacill1ts mdid­
cola Beij. spreads bacterial blight is very evident. Barss (18) remarks
that beam; shoultl not be soaked in a liquid culture of B. mdicioola
for nodule inoculation, since the soaking method results in a general
contamination of the entire seed lot, even if only very few seeds
are infected. Leonard ('Zl) hns reported that It slight application
of moisture will CtlUSe a stimulation of the bean-wilt disease, Bae­
t(!;)'hlilll. flacc/l'll//(nt'ieJlB.
This was also noted by the writer in 1925
aud 1927. ·Vllhiting, of the department of bacteriology at the Uni­
versity of 'Visconsin, npplied the wet-nodule seed treatment to a
Ilumbm' of samples of 'Vanlwell Kidney 'Vax variety. Apparently
only It small portiDn of the. original seeds was diseased, since few
blighted plants were produced in the check plots. The heated seeds,
however, produced plants manifesting appL'Oximately 100 per cent
blighj~,O which were .all killed before pod maturity. Since the checks
produced few blighted plants as compared with the complete b1ight­
iug of the treated seed, it was concluded that the dissemination was
brought about by the wet-seed treatment.
In 1928 at nerlin, 'Vis., where the Full Measure variety was
planted, a similar obs(,l'Yation was made by the writer. Seed PO'l'­
tions of this variety were treated with a liquid culture, whereas
the remaining portion ,yas planted without the treatment. A high
percentage of the plants grown from the heated s~ .::c1s were severely
:tfYected with typical cotyledonary symptoms, showing that the
disease must have come from the seed. The untreated seeds pro­
duced plants showing a slight amount 00£ infection, proving that
the wet treatment accounted for the widespread occurrence of the
disease.
R'ELA'fION OF PARASITE TO HOST
MATERIALS AND METHODS
The mat.erial fOt· the investigation of the parasitic relationship
of Baoterium pl/(/8(~oli to the bean consisted principally of the Ward­
well Kidney 'Vax variety. The material was collected both in the
greenhouse and in the field. Before killing, portions of the diseased
at'eas ·were plated out, in order to be positive that only Bact, phaseoli
was present. Formal-acet.ic alcohol was used throughout as a fixing
fluid. The sections were stained either with Giemsa stain (orange
G as a counterstain) or with safranin (licht griin in absolute alcohol
IlS the counterstain). These stains were used in a 2 per cent solution,
and the sections were allowed to remain in them for a period of 6
to 12 hoUl's, after which they were destained in absolute alcohol
and then cOl1l1terstained. These stains in the above dilutions were
f9.und to Le very 2fi'ective, because the host tissue took the stain
faintly, and the difi'erentiation. between the bacteria and the sur­
rounding tissues was very clear and distinct.
• Ullpubllshcd data from A. L. Whiting.
14
TUlOHNJOAL 13ULc,E'l'lN 186, D. S. DEPT. Ol!' AGl1fCULTUHE
RELATION OF PARASITE TO LEAF TISSUE
Microscopic exuminutiolls, It;,; previously reported by Smith (:J(J) ,
revealed that the organism gaills its entralll'e through the stomata.
Since stoma !"It II 1'(' more nWJ1()l'OUS on the ullder side of the leaves,
it is here tha~ the gl'~atest degree of infeetion takes place. (I!'ig. 1,
A ,and C.) r'.Jnh} Illight also be made through ,,:ounds in the tissues .
.Aftcr the bactenll cnter the stomata they pass JJ)to the sllbstoJllabLl
('(Lvity, multiply l'apillly, and when in sllflkient lIUm\)eL's penetrate
illto tlw lntereellulur spaces of the spongy pal'ellchYll1it.
'1'he bacterja nppear to produce nn enzyme which softens or pos­
sibLy dissolves the pectic materinls in advance of the pathoge'1e.
The middle I!Ltl1ella stains deep blue with tho Giemsa shun and can
be clearly clilterentinted from the primary walls of the adjacent
cells. 'rhe cells in the vicinity of the infected area usually show
abnormal chamcteristics. The lamellae of those ceUs take the stain
morc faintly than healthy ones, ancl in many cases it appears that
portions lLl'e dissolved out, since there is no regularity in the in­
tensity of Ule !;t,ain. Farther away from this region the celts are
nonnal, and the lamellae take the stain very uniformly. In the
region of SHere inteetion the bach~l·ja fill the intcl'celll1lal' spaces.
This bacterial mass later becomes emhedded ill the slimy matnx,
which caUSt!S all enlargement of the interc(>llular spaces owing to the
absorpti. ve powers of the slime. The epidermis remains intact, but
the underlying parenchyma tissues collapse, often forming large
bacterial pockets. When severe infection has taken place a large
brown scaldlike area, due to the death of the cells below, appears
on the leaf surface.
There appear to be two views as to how bacterial plant pathogenes
cause the death of the host cells. Bachmann (1) believes that the
cells nre killed by the extraction of liquids from the Pdtoplast fol­
lowed by plasmolysis owing to the high osmotic pressure set up in
the intei'cellular space;,; incident to bacteriaL invasion. Another view
by Steward (31) assumes that toxic products are secreted which dif­
fuse into and kill the cells. The writer's histological studies seem
to favoL' the theory of Bachmann. Staining reactions indicated that
when the bacteria occur in large numbers they urn n.lwl1Ys embedded
ill a slimy mass. The osmotic concentration of this material appears
to be greater than that of the cell ~ap itself, llml apparently an
~xo::lmosis takes place, causing the intercellular spaceH to enlarge,
and, as they become filled with It fluid, producing small water-soaked
lesions on the leaf surface, characteristic of initial blight symptoms.
Surrounding these small water-soaked spots thel'e develops a
yellowish halo. Upon microscopic examination few bacteria are
found in this discolored zone. In her study on the halo blight of
oats, Elliott (1B) found It similar condition. She statetl that it is
probable that the organism produces nl111l1onia, which is responsible
for the destruction or the chlorophyll about the Icf';i ow-; produced in
oat plants. The cause of this discoloration in be all blight has not yet
been tlrterminec1, but it is believed that a toxic substance secreted by
the organism di1fuses into the SllL'l'OllIHling tissue, causing the light­
yellow zone.
Microscopic examinations have shown that the bacteL'ilL are com­
monly pres<:llt in the xylem vesFP.ls of the leaf. (Fig. 2.) The
BAOTERIAL BLIGHT OF BEANS
FIGUItE l.-Stomatlll penetration by Jl(1ctc"i'm~ pll11.~coli: MORt of the P"Il!"
trntion of the lcn\"es. pods, nnd stems by this orgnnlsm In nature Is
stolllntni. A nnd C, Ilnct~rln penctrn ting the stomntn of the lenr. In
"neh cns'~ th" substollllltnl ('/wlty Is till",1 with a hnet(·,'lnl lIln~H, In C til<'
bnl'tcrill are following the Int,'rcellulnr spnc('s lenlllng from th" clI\'lty,
Il 1I11ll D, Bnctcrla In\'n(lIng stomntn of t:he stem. E. Ilnctet'in hn'ndlng n
stollln of the pod, X 1,200
15
16
TECIINICAL BULLETIN 186, U. S. DEPT. OF AGRICULTURE
organisms probably enter the large xylem vessels by first invading
the small veinlets, which in their initial stages of development consist
of undifferentiated tissue similar to that of the surrounding paren­
chyma. These tissues appear to be easily attacked! and after the
organism once gains entry it passes into the lnl'ge veinlets, which in
turn lead into the main veins of til<.' Ipat. After gaining access into
this tissue the bactCL'i!1 multiply rapidly anel wlipn in sufficient num­
bers cause It browning of the veins and veinlets (pl. 1, C) with a
grndual killing of the sllrroUlHling tissue, The pathogene might
also eHter the vessels of.the leaf by passing from infected petioles
FIGUItE
2,-VnsclIJllr Im'nslon of II belln lenf, Cross section of
bncterlll embedded In n. slimy mil trlx In the xylem vessels,
II
midrib showing
X 1,250
through the pulvinus and into the main vein. The bacteria seem to
become localized in the pulvinus, possibly because of the succulence
of this tissue, A severe invasion of these structures results in a
drooping of the leaves, a Yel'y characteristic symptom of primary
seed infection.
RELATION OF PARASITE TO STEM TISSUE
Burkholder (5) and Barss (~) observed that the disease was
systemic in 11Itture. Micl'oscopic study revealed that the vascular
~ystem of the stalks wn.s invaded by the bacteria, although no external
lesions appeal'f'd on the leaves or pods.
Tech. l3ul. 186, l!. S. Dept.
or Agriculture
PLATE 1
b
,
.,.,_1•••"
G
-
.• ­
~
H
ttl) lin b,'an . . tt·ru. If) 11·l)·... a(li'r irHH'ul:lliol1, X ~~; H, illf{'etl,d IlPlln Sll('dlin~
..howHlc dr{lt)ltlU~ nf h·~t\ (.... :ll pllh mil.. 'fP and .. ttont ('I",lt'kiu)! (/11 owiu~ tel 1t:H*tt'rinl fu(p('tiOll,
:< I:.!; ('I fhrhl<IH'd ,'pins :HJl) vpinlt'l'i HII flllhl\\ lJt~ 11)\":1"01011 uf t1l1 1 n'~~I'ls hr I Itt' Imdt'I'in. Hrnnll
\, ",ldl'thl l'\l1d:ltl'
W ltn·...tI:\h.(ld "'sIiln"'- dJI l'l'..;l1lr (lion !'>ltHll.Il:d Wfl'l'illlIl HId Iltl,\' I't·";1I1t In (IIrlh(,I' Vil~culnr irl\·u•
... jnlt wlit'n lit n1!lt:ll't \\ iIh \I'Wld .., "< I, Il, hal'tl'nn Itav!' I'IIt'irc'lt'd till' t'pi<'otyl and protlu('(lll n
!!lrdh: w' \\ hieh ~',lIhl'd till' ,t.,w II~ \\ j·:lk"t1. ;\;ltural Hl,(('('liol1, X Ii;, I';, illfl't'lPti IH'tlU stl'lll
..ho\\ 1IlJ.!' Inngiludmal r-('d"t"olutl'cj II,.. tlll1"i l"lIl,,;pII by h:tl'tt'T1:tI IIlfl'('tioll. ,S'ntllrnl Inftlc" iOIl, Xl;
VI l'fIIl h,'sillllS n':'llltln~ (rom "'11111I'1t'\1 fuftll'tltHI, .\n t1lt't'U~I;lLiou Ilr (Irit'tl 1':I('lt"'j:ll siilll{, i:;
"l't'll It) tilt' l't'utt'r IIf m:lIl) ..1'"1 .... , .\ lIunl mfl'I'liou, ~ .l,; n, db-liaS/It! !'Ol't'd tlf lIw Boulltiful
\.Inpt~ ..hO\\lt1~ ..llI'l\I'i.lll..! 1111\ dl',t'uim':lIiflll!-. (':uz"'I'd hy 1t:lt'lt'rial itl\':lsjol) o( IIlP .!-:l'pd (·onts.
~\.lllllli IUfl'dlflu. x "'I~ II. "'('I·d.., rrllm pod .... }town in f. Tlw thn'(' silrt\pil'd :-;'Pl'tb Of Ihp 141ft.
"t'rt, It'n1I1\t'd
rrllnt
Iht*
t'\III'01I' ru.::ht I,union of tlw pod, "< :t.,: I, pnd uf till' Boun1iful vnril'Ly
,Iol'''al "II till I" IltJll t .. "lH'I"!'ial iU\,:t-.IOU, :-< 'l.,
.... IHI\\ tUb dl!"l'olnt"ltion :t1ofl!-!; Ihl'
T«h. l3ul. 186. U. S. Dept. of Agriculture
PLATE 2
PHOTOMICROGRAPHS SHOWING BACTERIAL INVASION OF THE HYPOCOTYL
AND COTYLEDON
'\1 Il:w1I'1"I:1 ill UII' \) ielll \ l,:"...,·I.. . url,lIH·flJ~:P11("lll~ I.
In ~ulth' I':i"'l'~ lhl'~ h;l\p hl'Ohl'U ullt fl'oll1 t ht'
\'t's,wb :md an' 1';HI~in~ 1 dt~jtttl't!r:lllun o( tht· :trij:Wl'nt 11 ...... Ilt', (onlting It:ll'h'l'i:d Ptlt'J..ll~. ';.~ 1,It:H~
H. Int'{l<ri 1 ('tlll'riu~ ,III "pi'II'1 mal rin of th,' 1'(11 ylt'dllIl ",lu",,'d b~ <, ""p'll·hill)! flf tlH' ('l'lh dllriJl~
~t'J min:llil'l1.
'1'11('
II lthu~I'fll' ,';111
hI'
""'I'U
rol1owilJ!o!
lhl' 1Jtlf'rt'I'IIIII,IJ' "P"U't·S, ("ItI:-.illl!: I 1It'1I1
tu
t'uh,t'j!I' alit! til(> I'db bi bl' JltJ~llI'd lP:Ut
Till' .ulj:H't'lit n,ll ... an- III 1 dblOl'h·d j'undiliull rI'll III I
f'lt",,,,lln' "\1'1 bid b~ lllt" h:ldt'll.1I ~lllIh' 1lJ tlH' H1ku'plllll:lI "'1';U'l'!'o, ~.., I.t);il
B~CTEl1IAT~
BLIGHT OF BEANS
17
AI'tifiC'illl inoculations were per:iol'l1H'd by the writer by cutting off
the young t:otyledons befot,c the absl.'ission layt'l' had formed and
inserting tl drop Ot' two of bacterial ino(,lliultl into the cut. This
allowed the bacteria to enter the vascular systelll, and within a period
of 10 tla.ys the inoculated plants munifestt'd s.iight symptoms of wilt­
ing. rrhe check plants appeal'ed normal in all respects. Isolations
il'om such material and also from wilted seedlings grown frol11 dis­
eus('d se('(( d(,Inonstrnted the IH'esenl.'p of the baetel'ia in the tissues.
The pathogene was fourHI thl'Ollgh mierosl'opic examination t{) be
present ill great llurnbl'L's ill lIlall,)' of the xyll'lll ,'essets (figs. 3 and 4)
Ilnd to extend up Hnd down the stem frolll the cotyledonHry nodes.
In severe ('uses of infection the badpria apP(,llr to ureak through the
wnlIs of the im'adell vessPls alld to spI'en<l into the nellt'-by pllren­
r--..
Jo'IOVIIB
a,--Cross section of Ii benn stem showing invnsion of metnxylem ,'essels by
bllct"ria elllbedded in slime, X 1,!!(iO
chymn, C('lls. (PI. 2, A.) .As stated berore, in the growth of these
organisms much slime having the property of absorbing a large
amount or fluid is always produced. 'With this absorption a nlLtural
internal pres::;llre mny be set up, but whether this pressure is great
enough to cause a rupture of the cells is still undetermined. ,Vhen
the bacterin, break through the cell walls they enter the intercellular
spnces of the adjacent tissue (fig. Ii . .A, R and C), slowly dissolve
the middle lamella of these ('('Ib;, alld finally push them ai)art with.
a gradual disintegration of the tisi:iues. (PI. 2, A.) Bacterial cavi­
ties are not llIleonUllon in such l'cgions. 'l'hat the p:tthogene may
entH the stem from infected leuves appears to be vet'y probable,
In many cases where local infection begins through stomata in the
vicinity of smull veinleb; which urc roulld throughout the leaf, the
readily attacked xylem elements of these \'einleb; are occupied by
10020;-30--3
18
'n:CHX1CAT" ntJLLETfN
186,
U, S, DBT.'T, OF
AGmenf:runE
badet'ia, ..:.\1tet' (h(' bllctcl'ia entCt' the l:'mall y('inlet;.; ther Il111.kc their
way into UH' In r~('l' veins and tlH'nee i lJ'{O the Illich-ib, trll \'el ing down
the l)('tiol(' and into the ),;te!l1 of the plant. The bactel'iu may also
entl~l' thc xylem \'C!:ise!s of the stem by way of: the elluline stomata,
(Fig, 1, nand D,) The bacteria entering these openln~s multiply
in Lhe sub;;tomatal ('!l\'itieR and later PPlwtl'lltc the intercellular spaces
of tilt' ('ol'f:il'al erik ,Vithin II comparativrly short tim(! the patho­
gene may IIl1\'C' inV1Hle(1 Il1I1('h of tIl(' sllI'I'olll1c1ing tissue. l)l'olillcing
tL \\'utel'-soak(,d 1<>sioll which mani fpsts il:,('lf on the C'xt!'l'iol' of the
)-­
/
\
\
A
Flilnn1 .1.-·.·lIuc{(,r!n in x~'I'-1Il \'('s~('IH oC II hClln stPlll: A und n, Bnd,'riu f'lIlbcdo1,'o.l In
l\ slimy JIllltl'ix l!n-,\(Ilng ringed nH'l:nxrl~lll ,"esRels.
X l,:!(;O
W'!tNt in sufliticnt numbers the bacteria hegin to ClllISC" disin­
tegl'ntioll of the invaded cortical tissue, forming in many eases large
lysigenous Cll \'ities.
In nurnel'OllS sections in which penetration has b£'glln from the
extct'iol' of the :;tPlll tI\l' (,Ol-ticnl tisslles arc se\'erely invaded by the
puthc)lrcne. but within the endoc1ermis few hactct'in are in ('vidence,
stem.
The l:~v(,l' ()f eodieal ('ells contiguous to :lnd including the cndodcrmis
l\PP(,!ll~S to act as n bnrriPI' in partly pl'cventin:.r th(! hacteria from
penetrating into the \'asc1l1ar tissue. (Fig, 6.) These C(!US do not
completely sU!'l'()lUHI the stele of the stem, llnd it I:; through the
B.\CTEHIAL BLlG(LT OF BEANS
19
breaks that the bnch'ria mnkc their wav into the xykm y('sse\:.;. Suell
penetration takes plat'p only wlwll the plants :l1'e young-that is.
berore secondar,Y thiekcning has takcn place-siIH:p the l'ells or the
secondary xykm app('lu' to bl' liUle affl'eted by tIll' bnet('l'ia. The
cdl::; of thc endodermiH ure n'ry thitk walled, with Slll:lll inteeccllulnr
SplltCS. It is possible that bt'cauHc of this adaptive strlldure the
ol'gnnisms nre unable to pt'netmtc this tissue.
Bacteria lire seldom if ('yer round in the phloclll region; Il)(lst of
tlH'111 invade the protoxylelll and n1l'taxylclll C('US (fig. a) ; only occa­
sionally 11I'(> tlll'Y fonnd ill the Sl'(,OIHltu'y xylem. It is probable that
in th(1 fOl'Il1ution of the ('ell wnlls of the pl'il1lary ti:-'Slte the wall
lllU l.L'ri a Is are built up gradually. \\'hieh means thnt ligllifieat:ion is the
Ii'melt1-1 ;;.--1 Hh'I'("('llulnr Jl(\J\(\tr'a t Iuu uf pttl"louchYlIln l'(HIS hy hat1h'riu: A H IIti C.
]t]n1:tn,;pd intt're('lIulut' SPlll'PH d\w to iu\'usiOIi IJv bat:ttll'lil l"mb\ll.ltll,ld ill :l ;.;lillll'
Illlltrix; H, portion
ing bll\~t(,I'lII to
or
,'11 let·
II ,'\'11 Willi (Usintl'grll (,'<I 'through hlld"l'inl actloll, nllow_
:l!lj1lccnt c,'lIs,
X l,:!50
last process, The (,'('lIs in the ('ad.,' stagcs of growth contain l:ll'ge
amounts of cellulose or of h('micdluloses. These substances most
likely arc nUnekcel by baet('rial action, 1111<1 cntmnce may bl' gained
into thc young protoxyl('1U nnd metaxylcm cells. HowC\'cl', in the
forlllation of secondary xylem, which is laid down rapidly, the walls
possibly I'(~main in the cellulose state for 11 very shod pcl'iod. becom­
ing lig-nified yery r~picny. It.is })l'?baJ:>le th:~t the princi~)al !'(':lSon
thnt little pcnetratlOn occurs III thIS tissue IS bp('allse 01 tlH~ wall
composition of the ceUs.
That the hactel'ia togethel' with slime rcsulting from mpid mul­
tiplication of the organisms plug the vessels and cause It wilt of the
plnnt is not an pstnblished fact. Burkholder (4) belieycs that under
20
~r"CUSlC;\L BeLLT~TIS 186, t
T
•
S. DEPT. Oll' AGUICUL'rUllE
ccrtain conditions 1110 baetcl'ia may ('ntl'l' thc va~cl\lar tissuc ·without
cuusing a pel'~l1nnl'nt wilting, but ina~T pcrsist, causing dwarfing. and
on days of lugh enlpomtion may pl'odu('c a slight flagging of the
plant. According to the writer's investigation:.;. little of the wiltincr
of ~nntUL:e plnnts caused by Bacteriwn phaBeo{; appears to be du~
to plugglllg, but mOre likely it may be accounted Jor by It tlisinte­
gmtioll of the invaded tissue or the,' feet of toxie substances pro­
dueed by th(> pnl'!ll';itc. Microscopic"t>x!lminntionl'; havc ncyer re­
Ten led, that infected xylem "('sse Is Wl'l'P so compll'teJy filled with
bactcl'lal l11m,I';CS that c1l'ath mny haw' re~ultetl 1'1'0111 a plugging of
l~lGl'ln: (;,-B,\('t~rinl
Inytlsloll of the cortex of the hypocotyl. Cross section of the
IIJ'Pocotyl, showing lJllctcrlll InYlllllng the cortex where II Jllyer of cells (a) auja­
to tilt' L'll\lod<'rmls ll(l(1cllrs to IIct liS II barrier to hlYllSloll o( th~ Vllscular
l'l'lIt
ti~H\lP.
X 1,:.!GO
these cells, Since secondary xylem is formed rapidly in the ~tems
of oWer plants, the passage of water would not be sutliciently ham­
pered to cause wilting, even thouf!h the protoxylem and metaxylem
cells were filled with bacterial maBses.
The bacteria may enter the xylem vessels of the hypocotyl and
epicotyl from the infected cotyledons in the C!lse of diseased seed.
Under :fnvol'llble cOJ)(Htions the pathogene may increase with such
rapidity that the "essels become filled before ml1ch secondary thick­
enl11f! has taken place, In this case a plugging of the "essels may
bring about a wjltin~ of the young plant.
BACTERIAL BLIGHT OF BEANS
21
CELL-WALL DISINTEGRATION THROUGH BACTERIAL ACTION
Smith (34), working with Bactm'iwn campest1'e on turnip, stated
that the infected cells are crowded apart by the growth of the bac­
teria, and the middle lamella first disappears, bnt the cell walls
proper also become vague in outline and finally disappear. Micro­
scopic examinations have sh\ vn that in the case of seyere infection
of the hypocotyl, epicotyl, aI.t! funiculus with Baot. plwseoZi, dis­
integration of wall material becomes apparent. Careful observation
with serial sections indicated that the microtome knife was not the
cause of the disappearance of the wall or parts of it. (PI. 2, A, and
figs. 7 anti 8.) It appeared that the cellulose walls became dlsin­
tegmted, leaving only the lignified portions behind.
The protoxylem and metaxylem cells of the hypocotyl and epicotyl
in which most of the infection occllrs differ markedly from the
secondary xylem cells of thf> .~ame tissue in cell size and in wall
structure. Since the protoxylem cells are formed very early in the
FIGl'ItH i,-Cross H('('tion of Il belln stern, showing bnetel'ia im'lulIng nH'tnxylem cells
n 1111 JI"~slllg fro III e"11 to cpll thrOlI!:;h hrokNI WillI.
X 1,250
ontogeny of the tissue in which they lie, they are subject to tissue
stresses from increase in length and diameter. These pl'otoxylem cells
mature rapidly and arc not subject to growth changes. The stresses
brought about by elongation tend to stretch the already mature cells,
and in many cases a rifting results. These cells are 10nO' and slender,
with thin cellulose walls, reinforced by bands of lignified secondary
walls to prevent the collapse of the thin plastic walls. If bacteria
are pl'esent in such cells, it is probable that natural rifting of the
tissue would gi\'e the appearance of disintegration; but since this
occurs only in the Pl'otoxylem, it does not explain the partial disap­
pearance of the walls in many of the severely infected metaxylem
cells.
The water-conducting cells of the metaxylem are the characteristic
cells of this tissue because of the peculiar adaptation of their walls
to. the stretching that they normally undergo. The thin plastic
pl'lmary walls of these empty cells are also strenf,rthened by the addi­
22
TECHNICAL. BUL.LETIN 186, U. S. DEPT. OF AGRICULTUHE
tion of It lignified seconc1al'Y wall. The metaxylem cells first formed
have small amounts of secondary wall in the form of rings, whereas
cells fonned a little later possess spiral bands. 'The pl'oportionate
amount of secondary wall increases in the successively formed cells.
The fact that the greatest amount of infection OCClll;S in the pri­
mary xylem may possibly be explained by the chemical composition
of these tissues. Since the walls of the protoxylem and metaxylem
cells are composed mostly of cellulose, with only small amounts of
lignin in the fOl'm of spirals or rings, bacteria when in large masses
probably have the ability to pro(luce an enzyme (cellulase) with the
property of slowly c1issolvin~ the materials between the pits of the
vessels and finally other portIons of the cellulose walls, thus making
l~fGl!ln;
S.-Cross section of a benn fnnlculus, showing bac­
terlnl disin tcgrn tion of tissue. The bacterin h,we in vaded
the xylem cl('[nenls of the funiculus nnd disintegrated the
tissup wit h tlw fUloJluuiol1 I)f a hu'ge ly:-;ig(!lIolls ell \~ity
in the lower pnrt of which portions of the disintcgl'llted
\\'nlls may be Sl'<!n. X 1,:.!;;O
it possible for the pathogene to gain access directly into the xylem
elements or pass from one vessel to another. Little infection takes
place in the secondary xylem cells, probably because the walls ai'e
composed mostly of lignin, which the pathogene apparently is unable
to attack.
In the funicular region of infectecl pods other instances of cell­
wall clisintel;{ration were in evidence. .Microscopic examinations of
invaded fUlllculi many times revealed large lysi?,enous cavities show­
ing many stages of cell-wall disappearance. \Fig. 8.) The walls
of many of the invaded cells were thinner than the normal 'walls;
others were slightly visible, whereas mallY of them had disappeared
altogether. S('atterecl throughout these lnrge bacterial cavities were
straI1C1s of lignified wall material not attacked by bacterial action.
BAOTERIAL BLIGHT OF BEANS
23
UEI.ATION OF THE PAUASITE TO PODS AND SEEDS
It is a well-established fact that Bact81'i:il'n~ plwseoli is seed horne.
One of the most importa.nt points in the behavior of the organi::llll
is its ability to entol' the pods through the nlscular system alld
infect the seeds without ('llusing lesions 011 the surface of the pods.
In entering the spC'\l tll1"()llgh the vascular system the pathogene
fl'('(lUC'lItly eauses only a HIlHtll yellow discoloration at the hilum.
(PI. 1, G.) This sign of the disease is not readily detected in col­
ored seeds, as is also the case in white seeds, where normally there is
a slight yellow marking about the hilum. Seeds that possess only a
small amount of inf"dion without outward symptoms may cause
mueh <lamngn when planted the following year. From the stand­
point 1)£ control \'asclllar seNl infection is l·xtl·emely important. It
becomeH apparent that the seleetion of pods is not an adequate means
of eontl'ol aH ;.\ Ihp (,:1:';0 of hean anthra<:llo:,;c. where the infection
is IO(,lllizc(l.
be positive that only disease-free seeds are ob­
tained, only pOCo h'om healthy plants should be selected, and even
thclI, il' \,fI~clllnl' ini'C'ction is slight, n-ppal'ently healthy plants may
produce sC'ed!:i tlwt hnxbor the pathogene.
In causing this type 01' vascular infection, the bacteria travel up
the xyl.em vessels of the stem, through the vascular elements of the
pedico
<:nce they pass into the two sutures of the pod. The
lesions v u the sutures of severely infected pods are easily detected,
since they calise a discoloration of the vascular tissue, and ill many
cases water-soaked lesions extend along these sutures, particularly
the dorsal. (PI. 1, r.) The bacteria then travel from the xylem
vessels of this suture to those that pass into the funiculus and llre
then carried into the seed coats by way of the l'ap!le. The raphe
extencls only a short distance into the integuments and becomes un­
differentiated tissue, similar to that making up the third and fourth
nutritive cell layers of the seed coat. After the bacteria enter the
coats they make their way into the intercellular spaces, which are
extremely large and afford an easy passage for the spread of the
organism throughout the tissue. (Fig. 9, A.) ·When in sufficient
numbers the. organisms may destroy this tissue without the produc­
tion of any outward symptoms on the seed.
The first layer of the seed coat is composed of large palisadelike
cells, thick walled and upwards of 60,.,. in length. On both sides
of the hilum slit, which is found in the center of the hilum, two
layers of palisade cells are present, while immediately beneath the
slit is a group of sclerenchyma cells with reticulated" walls whi~hl
according to Tsrhirch and Oesterle (38), probably serve to prevent
the entrance of fungi into the seed. Below this palisadelike epider­
mal layer is found a. layer of cells known as the "I" layer, being
l111Hlc up of hOlll'gla:'.s-Hhapcd cells 18,.,. to 22,.,. in height, containing
calcium-oxalate crystals and possessing no intercellular spaces. Be­
low these two layers are found the nutritive spongy tissue with large
intercellular spaces into which the raphe enters. Bacteria passing
into the seed coats by means of the raphe traverse this region exten­
sively, as previously mentioned, but they seldom invade the first two
layers of the seed coat.
The old conception that direct penetration occurs through the
outer layers of the seed coat is difficult to explain, since in the first
<
24
TEOHNICAL BULLETIN 180, U. S. DEPT. OF AGRICULTURE
BACTERIAL
BJ~IGHT
OF BEANS
25
place the epidermal cells are covered with a layer of cutin or suberin;
and secondly, their cell walls are extremely thick and without inter­
cellular spaces so that penetration would be difficult. r.rhe symptoms
on mature seeds ,infected other than at the hilum, which are so often
illustrated, may be caused by some other parasite, possibly Oolleto­
tl'ic1wfJJlJ lind:eIJUlltlda?11li7n. :Much of the discoloration seen on matme
bacterial-blight infected seeds is probably due to the disintegration
of the cells below the palisadelike epidermal cells. This is especially
apparent in white-seeded varieties. In severely infected seeds,
sides this discoloration, a shriveling occurs, because of the collapse
of the third, fomth, and fifth layers of the first integument. In
cases where bacterial penetration is slight, llO discoloration is in
evidence at the hilar region, and it is often impossible to demonstrate
macroscopically the presence of the organism, although if severe
infection takes place a yellow or water-soaked discoloration is
evident.
.
"Vhen bacteria are found in large masses in the vascular elements
of the dorsal suture of the pod, they often cause disintegration of
the tissue and enter the surrounding parenchyma cells and thence pass
into the pod cavity where the young ovules are beginning to develop.
When funicular infection is severe, this tissue is destroyed, and the
seeds fail to undergo further development. It is a common occur­
rence to find one or two ovules of a pod decidedly shrunken, with the
remaining seeds developed to maturity.
.
When the pathogene enters the seed while in the milk stage the
bacteria in the seed coats may pass into the regions of the cotyledons
and under certain conditions entirely surrolmd these structures. The
bacteria appear to remain in this region as well as in the seed coats in
a dormant condition and do not cause the embryonic plant to become
infected until the time 0.£ germination.
This type of infection has been reported by both Barss (13) and
Burkholder (5) as being of a serious nature, since no external lesions
are noticeable on the pod. The writer's researches have shown that
a considerable nmount of infectiol1 takes place in this manner; how­
ever, bacterinl penetration through the micropyle of the seed is
cqunlly as importnnt ns vascular penetration and possibly more
widespread.
The bacteria mn,y enter the pod cavity, as stated above, by breaking
out, from the vascular tissue of the dorsal suture or the funiculus, or
by making their '.yay into the pod stomata. From here, as in the
leaf and stem, the pathogene fills the substomatal cavity (fig. 1, E),
pusses into the intercellular spaces, gradually becomes intracellular,
and later causes a disintegration of this tissue. The organisms have
likewise been found in the xylem vessels, which may distribute them
to all portions of the pod. In some cases they break out from the
vessels and when in large masses cause disintegration of the sur­
rounding cells. They may then pass into the pod cavity, where they
increase rapidly because of the favorable conditions for their develop­
ment. Numerous pods have been cut open, and in many instances
where only a slight amount of infection wp.s in evidence on the
exterior of the pod the inner cavities contained large masses of
bacteria embedded in It slime, surrounding, in many cases, each
individual seed. In such cases micropylar invasion is a simple
matter.
he­
26
TECHNIOAL BULLETIN 186, U. S. DEPT. OF AGIlICULTURE
The micropyle is a natural openiIlg in the palisade epidermis of
the seed coat, and the bacteritt may cuter here and reach the seed
coats with little difficulty. Below this point of entry a cavity is
found, and the surrounding cells, which are decidedly thick walled,
appear to be little affected by bacterial action. However, when the
organisms once make their way into the underlying tissues, little
opposition seems to be encountered. 'l'he bacteria pass readily
through the large intercellular spaces of the seed coat (fig. 9, A)
nnc1later break down this tissue, causing in severe cases a shriveling
of the developing ovnle.
It is in the immediate vicinity of the micropylar opening that the
young developing embryo is found. As the seed begins to germinate,
the young hypocotyl elongates and may push its way through this
diseased region as it emerges through the seed coat. Since the young
epidermal cells are very compact, with small intercellular spaces,
penetration into this tissue appears to be somewhat difficult.
In examining diseased germinated seeds it is not uncommon to
find the embryonic tip of the hypocotyl of many young embryos
killed before emergence. It seems likely that the pathogene present
in the seed coats in close proximity to this structure may cause the
death of these tissues, but the entrance of the bacteria into these
cells while they are still in the embryonic condition has not as yet
been actnally observed. If the bacteria are later found to enter
these cells, then much of the seedling wilt ClUJ be explained.
Stomata have been found on the hypocotyl above and below ground
after cell elongation has taken place. As this structure grows
through the diseased seed coat, it appears to be possible for the
pathogene to be carried aloIlg the surface of the cells, and as the
stomata are formed the adhering bacteria under favorable condi­
tions might enter these openings, ctwsing infection in this region.
It has likewise been obsel'Yecl that often in the case of diseased
seed the embryonic folds of the epicotyl. while still lying between
the cotyledons, are surrounded by bacteria. This fact may accOlmt
for the occurrence of the common initial water-soaked lesions often
found on the primary leaves of young diseased plants. These lesions
usually appear on the opposite simple leal'cs ill exactly the same posi­
tion, making it appear that the bacteria enter these embryonic struc­
ttu'es at the time when the primary letLyeS are still folded together.
The bacterin, that pass into the micropyle and invade the large
intercellular spaces of the seed coat may remain there in a dormant
condition until they penetrate into the cotylec1ona,ry tissue at germ­
ination time. On the other hau(l, if they enter the seed through the
vascular system they become well established in the seed coat also,
since they multiply therein ancI pass throughout by way of the inter­
cellular spaces. (Fig. 9, A.) In this way they may migrate into
the region of the micropyle, at which point they mny pass out of it
anel enter the cavity of the pod, where they multiply rapidly because
of the extremely high moisture conditions and abundant foo.d supply.
The organisms spreltd readily throughout this ca.\'ity and may infect
other young seeds by entering their micropyles. In thjs ml11l11el' it
can be seen thnt from a, single infected seed all the seeds of a pod
nmy become infected without the presence of 1I lesjon on the exterior
of the pod.
BACTERIAL BLIGHT OF BEANS
27
PENETRATION OF BACTERIA INTO THE COTYLEDON
The most important phase of seeel infection is the penetration of
the organism into the cotyledonaL'y tissues. For the study of this
phase of the problem, diseased seeds WHe surface sterilized in u solu­
tion of merclll'ic chloride 1-1,000 for 35 to 45 seconds, washed in
three cIlIwges of sterile water, and placed in sterile Pe'~ri dishes
between moistened sterilized filter paper. The seeds were allowed
to gel'lllinate, and niter the young hypocotyl had emerged through
the micL'Opyle the seed coats were removed and one of the cotyledons
of each seed was killed in formal-acetic acid fixative, the other being
used for the isolation of the organism. 'rhe killed cotyledons were
embedded in paraflin, <iectioned, and stained in the usual manner.
Since cotyledollaL'y penetmtion oecuL'L'ed only in germinated seeds,
it was suspected that the reason for the lack of. infection in ungermi­
nated olles was bt'cause of some protective covering, either cutin or
suoerin, over the cotyledonary epidermal cells of ungerminated seeds.
Microchemical tests were made for the presence of these substances
by treating freshly cut sections of cotyledons with a solution of
Sudan III. The sections were allowed to remain in this solution for
15 minutes, after which they were washed in 50 pel' cent alcohol,
placed in glycerill, and examined. The tests demonstrated that the
amount of suberin or clltin was extremely small and that there was
no difference in the amounts in the cotyledons at various stages of
deVelopment.
Microscopic examination of the embedded material revealed that
as the cotyledon absorbed water at the time of germination the sud­
den enlargement of the epidermal cells resulted in numerous in­
stanoes in the. pulling apart of their adjoining outer walls. .After
oosening these small r'ifts, and in mnny cases large tears, in the
cotyledonary tissues, measnrements of epidermal cells were made
before and after germination. These cells are disti nctly of two sizes,
those adjacent to the hilum being considembly longer than the cells
on the opposite side of the cotyledon. Measurements of 50 cells of
both types were made before and nfter germination. It was noted
that the a \'cr'age length of the former cells before germination was
46p., whereas these same cells after germination had enlarged to ap­
proximately 82.8p. in length. The other type of cells before germi­
nation averaged 8.6,l-p., "'hereas after germination they increased to
14:.25p. in lcngth. .As for increase in width before and after germi­
nation, little difference in size was noted.
It becollLes npparent that with the imbibition of water the :>eed
swells enormously. The mature cell walls apparently do not allow
for much expansion, and because of the enormous increase in length
a natural pulling apart of the cells results. (PI. 2, B, and fig. 10, B,
E, and F.) Some of these tissue tears are extremely small and are
often in the form of a small V (fig. 10, B), while others have been
found to measure as much as 9.8p..
With the pulling apart of the epidermal cells there is likewise a
stretching of the intercellular spaces. The bacteria on the outside
of the cotyledon may enter these rifts, pass into the intercellular
spaces, and in some cases cause them to swell to enormous size with
the distortion of the adjacent cells. (PI. 2, B, and fig. 9, B.) Dis­
integmtion then tukes place, and the bacteria rupidly traverse the
28
'rECIINICAL BULLETIN" 186, 1..'". S. DEPT. OF AGRICULTUHE
10.-Cross sections of b('an'sI'NI nlltt(,l'lnl showing normal nnd bacteria·
Infected tissues: A.-Normnl cotyledonary epidermal cells which show dark­
stained areas wbere the cells nrc pulled apart at ge~mlnation time. X 1,250.
B.-A natural rift In the epidermis enused by a stretching of the epidermal
cells. X 1,250. C.-Bacteria embedded in a slime in the intercellular space
of ('otyledollllI'Y cell8.
X 1.250. D.-Bacterin inmdlng the cells of th\!
funiculus. X ·S50. E. and I<'.-Bacterla entering the natural epidermal rifts
of the cotyledon and infecting the cells below the epidermis. X 1,:!50
FIOOlll)
BAOTERIAL BLIGHT OF BEANS
29
cotyledonary tissue, often forming cavities. It appears that the bac­
teria may then pnss into the vascular elements and thence enter the
xylem cells of the hypocotyl and epicotylat the cotyledonary node.
In connection witl~ epidennal penetration, microchemical tests
were made to detel'lnine the possible composition of the cell walls at
variou!; stages of development. Fresh sections of seeds in the milk,
mature, and germinating stages were made and placed in small vials
o~ether for the extmction of existing oils and fats. The material
was then treated with a 0.0;, per cent solution of ammonium oxalate
twd placed on a sand bath at 90° C. for a few hours. The vials were
allowed to stand for II short period at ordinary temperatures, and
upon cxamination .it was obscned that the cells of the geL'minated
cotyledons were not well intact and apparently had their middle
lamellae soml.'what diHsoh'cd by the ammonium oxalate, The epi­
dermal cells were likewise in It disintegrated condition, demonstrat­
ing thnt the walls were pl'Obably composed of a soluble substance,
pectinlike in nature. The walls of the cotyledons in the milk stage
lind also those in the matuL'e condition remained intact, demonstrat­
ing the presence of nn insoluble substance.
From this it cnn be assumed thnt penetration into the gprminated
cotyledon might be explained by a dissolution of the soluble mate­
rial in the walls of the cotyledonnry cells. At the time of germina­
tion the food materials in the seed are being changed to soluble
substances for the nourishment of the young developing embryo. If
the composition of the epidermal cells is likewise changed, ns the
foregoing experimcnt indicated, it nppears that the bacteria, after
once fraining entmnce into the epidermis through small ruptures that
arc formed during germination, make rapid progress into the inner
tisslles of the sced.
Furt.her to sllbstantiate the above results, similar material was
stnined with the GiemslL stain, which, ns stated before, has an affinity
for middle-lameila. material. staining these substances deeply. Cer­
tain regions of the epidermal waUs of uninfected cotyledons took this
stain very rendily. (Fig, 10, A.) These regions were found be­
tween the cells and extended a short distance to both sides of these
cells whc)'e bacteria I. penetratioll in the case of diseased cotyledons
took place. It seems reasonable to suppose that these dark-stained
arcas nre composed of a. soluble pectinlike substnnce: as the results
inclicated. If, as is believed, the bacteria sccrete an enzyme, pec­
tinase, these darkly stained areas call be broken down, espel~ia!ly after
the bacteria have made their way into the small epidc,"mal rifts
cnusec1 by an enlargement of the cells. (Fig. 10, F.)
Briefly, the cycle of development of the disease in the light of the
pathological histolog~r may be traced as follows: The bacteria in the
seed coats make their way into the region between and about the
cotyledons, and with th<; germination of th~ seed the pathogene en­
ters the cotyledonary tIss~le, follows the lI1tercellular spaces (fig.
10, C), and finally may gam access to the vascular elements. From
here it passes into the young seedling, traveling up the vessels of the
epicotyl ILnd part way down the hypocotyl. Burkholder (5) states
that the bacteria extend into the root system. He remarks that in
the xylem vesselH of the tap and laterall'oots great masses of bacteria
are found similar to those observed in that part of the plant above
30
TECIINlCAIi HULLB~'IN 18u, U. S. DEPT. OF AGHICULTURE
ground. Simil~r results hl1"e not been observed by the writer;
however, bacterllt have been delllollst1'llted in the vessels of the tl'lln­
sitioll stage of the hypocotyl, that portion in which the xylem de­
yelopment ch:wges from the exarch to the endarch condition.
After the bacteria. enter the vascular system the seedling often
wilts. The cause of this wilting has not been definitely established
as Jet. It might be owing to a plugging of the vessels by the bac­
teria and retarding the transpiration stream of the plant by caus1.ng
the invaded tissues to become disintegrated or as II. result of toxic
effects of the metabolic by-products of the organism. The bacteria
in the sUl'viving plnnts multiply rapidly nnd with this incrense pro­
duce considerable slime possessing high absorptive properties. They
break throngh the vessels and spread into the near-by parenchyma
cells, with a gradual disintegration of this tissue resulting in the
production of large bacterinl pockets.
The bact('ria travd up the st('m, enter the vessels of the petiole,
and mllss to a great extent in the xylem elcn1('nts of the pulvinus, this
tissue being more succul('nt than the tissues of the hypocotyl, epicotyl,
or petiole. FI'om here the pathogene passes into the main vein of
the leaf and thence into the smaller veins and veinlets. On the other
hanel, the bacterin may enter the stomata of the leaf, pass into the
intercellular spaces of the parenchyma, enter the vessels, and thence
pass into the vascular el('ments of the petiole and epicotyl.
From the :-..-vlem vessels of the stem the bacteria pass into those of
the pedieel anel peduncle and enter the sutures of the pod. From
the dorsal suture they enter the funiculus (fig. 10, D), either dis­
inte~rating this structure to such an extent that the seeds fail to
develop, or traveling through the raphe which leads into the seed
cOllts, where the organism overwinters. The bacteria entei'ing the
poel cavity either from the funicular region or directly from the
stomata ot the pod make their way thence into the seed coats.
The young plants may become intected at the time of germination,
and the cycle is then repeated.
VARIETAL RESISTANCE
For a number of years past considerable progress has been made
in t,h,e de,'eiopment Ot varieties of beans resistant to somc of the
common mtlladies. Mo~t of this development, however, has dealt
with bean anthracnose caused by Colletotriclnl1n lin (lel1wthiamtm ,
root rot caused by P'!l8a1'i1l7n '1IUll'tii pha.seoZi, and mosaic. Little
hus been accomplished in the breeding of varieties resistant to the
common blight caused by Bactmiu1n 7J1wseoli.
The works of BurldlOlder (fJ) and Ra.nds and Brotherton (~7) on
varietal resistance to bacterial blight are well known and need no
review. Since the"e te.r;:ts 'were carried on in New York and Michi­
gan, respectively) where climatic conditions are somewhat different
from those ill 1Vu:.consin, jt was deemed advisable to duplicate these
studies. The problem was carried on Tor a period of three summers,
practically all of the commercial .canning-bean varieties being used.
METHODS
Bacterial cultures tor all inoculation experiments were grown in
lllrge quantities, which necessitated i:u'ge surfaces upon which to
B,\CTERIAL BLIGHT OF BEANS
31
grow the organism. FOL' this purpose liter flasks were at first em­
ployed, but it wus latcr fonnel .that 5qO-c. c. culture bottles were
more advantageOll:i. ~\bout 1 lIleh of potato-dextrose agar was
placed in the f111:-;k, :;tcL'ilized. and then tipped upon its side to pro­
\'ide it larger surface.. ~\n onlinnry test-tube cultmc of .Bacterium
pltaseoli was washed 0/£ with ::terile water, and the suspensIOn pOUl'ed
into the flask containincr the agar. These translers were made under
a. hoodpd chamb('L·. pl'~\'i()usl.Y sterilized. to pre\'ent contamination.
TIll' Hasks containing the bacteL'ia. were allowed to incubate at 28°
C, from three to live days. a.fter which they were washed ofr with
sterile water, and the SlISlwlIsion was placed in a hanel spmy pump.
The beans inoculatcd in the prelimll1iu'y te~ts to establish It suit­
able. method wcrt' gl'own in L'OWS about ;3 feet apart. Boxes were
placed O\'Cr thc pI:lnts after they had been sprayed with the bac­
terinl suspension, and a slight spray of water was directed over
them. for a period of -:1:8 hours. The boxes were then removed and
the plants examined daily for symptoms. Check plants were run
in the same mannel', bllt instead of the bacterial su:;penRion distilled
wateL' was used. .At the end of l2 clays characteristic blight lesions
\\'l'l'l' l\oticed Oil the i \loculated plallts, wheL'eas the checks were free
from blight.
'l'IIe SlI('CPS!; of thi:; nH'thod of inoculation led to a large-scale ex­
periment in which ~O dill'crent nll'ieties of beans weL'e used, each
nn'iety consisting or Ol\e -lO-foot 1'0\\" and the varidal rows a feet
aptlL't. A similar plot. wail pitmtpd as a check, ,,:rith the liRe of a
3-gn.llon compressed-air spmyer the under side of the leaves could
be ('o\"cr('d ea::;ily with the inoculum, The plants were inoculated
at various times of the day, and the amount of infection was noted
in eHch case. SOllle we\'(' inoculated eady in the moming, others in
the !ail' a fternoon just i>1'j'OI'l' sunset., al1cl still othel's elirly in the
en'ning. The hp:,t rl'slllts Wl're noted on the plants that were in­
oculatl'(l in the latl' aftl'I'noon, The possible explanation for this is
that tbe leaf stomata wcre open wi(ler at that pcriocl than at other
times of the (hy, E\'cll though the SUIl ·was shininrr, it was not
illt:t'n:'c t'IH.lIlgh to ('a usc a ny a ppl'eciahle drying of ~the bacteria.
U\l(lpl' thes(' ('oIHlitions a high. percentage of infection resulted.
"'hpll the plnnts wpre inocl\lated at night thcre was, ,,,ithont doubt,
('on:iidl'l'nble moisture in the form of dew on the plants. The sto­
mata, howe\'pr, \\,pl'e not as wide open as during the clay, as shown
by gL't'('nho\l~e stllelies, and thi" pL'olmbly accounted To'r the small
amount of infection, The bacteria sprayed on the plants early in
the morning probably dried to a large degree because of sunlight,
and hpnce comparatively little infection reslllted.
In oedct, to produce a maximum amount of infection, the plants
wel'e inoclllated at "lll'iollS Btages of development, viz, before blos­
soming, during the blossoming period, and at the time of pod for­
mation. Checkrows similar to those inoculated were allowed to
grow normally. Observations on "ttL'ietal resis!:ance and suscepti­
bilit\' were made from time to time. A small amonnt of infection
was 'noted in the check plots, having come about through natural
$pread from the inoculated plots.
"
DUl'ing the summel' of 1927 a third type of inoculation was carried
on with fttirly good results. The two varieties used for this experi­
ment were the Refugee 'Vax, a fairly resistant variety, and Full
32
TECHNlCAL BULLETIN 186, U. S. DBPT. OF AGlUCULTUl!E
Measure, one qnite susceptible. A very thick water suspension of
the organism was made, and seeds of the two varieties mentioned
were soaked in this suspension for about ihe minutes, nfter which
they were planted. The suspension was then poured over the Refu­
gee 'Vax seeds after being placed in the {unow, while the Fun
~reusure seeds 'were cO\'ereel with soil without the last treatment.
The plunting was made July 1. On July 27 these two vllrieties
were examined for symptoms. The Rcfu!!l'e 'Yllx showed very little
inJection, whereas the Full Measure was henvily infected. Besides
having many leaf lesions, much infection was noted at the cotyledon­
ary node, indicating that the bacteria, after the seed coats had been
ruptured, had possibly made their way into the cotyledonary tissue
anel thence passed into the vascular clements of the epicotyl and hypo­
cotyl, producing lesions at the point of entry into the seedlings. E,'en
though both of these YHrieties were given the same soaking treatment,
many more of the seed coats of the Refugee 'Vax variety than of the
:Fulll\Ieas\Il'e \\'('t'e ruptured. r.rhis apparently had little effect on the
amount of infcetion. since the Refugee ''''ax had fewer lesions as
compared with the Full l\Ieasure, whieh show(>d a high dpgTPe of
susceptibility. Sincc this type of: inoculation is far more satisfactory
than the ordinary type mentioned pt'eviously, because of the fact
that weather conditions do not have to be taken into consideration
for abundant infection, it is entirely possible that it would be feasi­
ble for use in \'I1l'ietal-resistance work.
,
VARIETAL TESTS
Table 2 shows the results of three summers' work on the problem
of varietal rcsistance. All of the varieties listed were grown either
in 1!)25, 1926, 1!)27, or in each of the threc years. The amount of
inJection recorded as ,: yery light" corresponds to any infection up
to {; pel' cent of the crop; :; lirrht" indicates approximately 25 per
cent of the Ct'OP infected, "m~diuIl1" about 50 pCI' cent, "heavy"
65 pCI' cent, ,; severe " abont 80 per cent. and •. vcry severe" that all
of the plants were infected.
2,-Llmofllft of infection (m II I/l/lIIll('r Of ool/uncn:ial cannillg beans
illoolt/atoll 10ilh Bao/('rillln 1I/I(1S('ol; ';11. 1925, }926, II/Ill 1927, 'llmler fi('ld
TABLE
OOltl/i.tiOIlS
[:I:, Very Iil;ht inf~clion;
,
Average (Icgreo of
infection
+,
++. rnediurn infcetion; +++, heuvy infection; ++++,
+++++. ,'er~' se"ere infcction)
light infection;
se,'ere infection;
'I'ypc of
lH!On Ij
Vllriety
__ ""
lQ'~
'
_~~~___
!!
,
'{!logers Stringless Greell HCfugee. _______
±
' It
(1
Itefugeo 1,OO(J-L_________________________
±
cry I1& I -------.. oreen _____ Extra ~~arly nefugee ____________________
±
Keenc~' Stringless Groeu Rofugce_______
±
IFuIl },[easurc___________________________
1.lghL_____________ ---do•• _. ___ I.ow Champion ____•_____,.. __ .___________
1lJlurpee Fordhook Favonte bush bean, __ ' _________ ..
Burpee XO\\- Kidney \\'n"__.. _____ ._-. __
Hogers Improved Kidne~' Wa.... __ ..... _ ... _____ ._
'
IX
Pod Kidney wux·_· ___ ' _______ .. 1
1) 0.___________ \\ I, ------ ROUl!d
Poncll Pod Wax _______• _____ •___ ..... _.. ,
,[ Webber WUX .. ________________ .... ______ '_________
I
++
+
+
+
+
10211
!
102.
"__ ~" _ __
±'
±
±
±
±
±
±
+
__ ._+
__ .__ ..,____ ±+
_____ _
+!
+ ,
+ i
++
+
++
+
+
+
\..__ ._....
+++
++ , ---+------i
++
++
+
+++
+,:
+++++'
++
•
+
+++
H!,st p~oqf (:~I,(~en ~\:ux .... -.---.- ...... ------.---,
Glunt ::;tnn~l<ss Green POd ___ ._ .... ____1
Burpee il(ringless Green pOd ••• _______ ._\
Medium _________ ._ Grl'eu, ____ Dwarf IIorticulturIlL .. ___ .. ____________
{ LongfeIlow __________________________ .._
Improved nou/IIll'od ""Ientine..______ .... _.. _..
33
BAOTERIAL BLIGHT OF BEANS
TAOLEl
Average degreo at
Intoctlon
2,-Amoufl.t of ill.[ection, eta,-Continued
Ty pe at
boan
Variety
1925
1926
1927
1-----------·1--- -----­
sure
Crop Wax_________________________
+
+++
++
Olds rAlte Stringless Wax _______________ __________
++
+
Modlum__
Wax ______ Olds Early Stringless WIIX______________ __________
±
+++
Herugee
WI1X____________________________ __________
++
++
+++
++
Uodson Wax.___________________________
flllProved Golden WIIX__________________ +++++ ++++ +++
Ru"tproor Wax __________________ ---------- ++++
liea v y-_.________ ••• -- II 0.__ .... Ourrie
+++
DOlVls White WIlX_ .._____ .._____________ __________
++
+++
{ lIurpoo Dluck Wax______________________ ---------- +++ 1_________ _
IJOuntiruL _____________________________ ++++ ++++ ++++
Sevoro_____________ Orcon. ____ Illllck \'lIlentine_.____________________________ •____ ++++, ++
{ Wells Hcd "Kldney______________________ ---------- ++++ I ++++
D
WII
Style Wax __________________________ +++++ ++++ I' +++
0____________
o. ----.. {Old
Wanlwoll Wnx __________________________ ++++ ++++ ++++
Very sevo'o________, (l,ooll_____ 'renllllssee (lreen Pod ___________________ +++++ +++++,________ __
Do
WI
{(:rystlll Whi,le WnL ____ . _______________ ---------- +++++,________ __
._._ _ -----------0 x______ Koolloy Wluto-seeded wa~~--~~~~~~--~~:~~-=-_~~--------I+++++
~-------
l
From these results it can readily be seen that the different varie­
ties of beans show com;idcrable vari!ttion in their susceptibility to
baet(n'ial blight. Thel'(~ !IL'e no known varieties thnt show nbsolute
resistance to the pathogene, but there al'e a few that exhibit a high
deg-L'ee of resista.nce, The Refugee types, comprising Extra Early
Green Refugl'c, Refugee 1000-1, the Stringless Refugee, and Refugee
Wax show little evidence of. infection. Fortwlately, most of these
varieties are of an exeellent type and quality and are used to a great
extent by many of the liVisconsin canners, 'Vith the exception of
the Refugee 'Vax, they ;~re later than other varieties, and it has been
suggested that possibly they owe their resistance to their lateness
of maturity, Other varieties that show It medium degree of resist­
ance coupled with good cannillg qualities arc Giant Stringless Green
Pod, BUL'pee Str'illglcs;:; Green l)od, Full Measure, Burpee New Kid­
ney 'Vax, Round Pod Kidney 'Vax, and Rogers Improved Kidney
-Wax, Extremely susct'ptible "Varieties are Bountiful, Dwarf Horti­
culturul, Tennessee Green Pod, Improved Golden Wax, Old Style
Wax, 'Varclwell 'Wax, Currie Rustproof 'Wax, Crystal White 1Vf.x,
Keeney 'Vhite-seeded Wax, and 'Vells Red Kidney.
Table 2 shows that the rcsults in each case over the 3-year period
arc not at all comparable, This can be explained because similar
weather conditions did not prenlil .in the respective years which
gave some vaL'iation in the degree of infection,
SU1\mARY
B!lCteL'ia I blight of beans caused by Bacte1'iwm phaseoli is seed
borne and may cause characteristic lesions on stems, leaves, pods,
Ilnd seeds. In cases of severe infection the seedling may often
ms.nifest a wilting, resulting from disinte~ration, toxic effects of
the bacterial by-products, or plugging of the xylem vessels of the
stem,
Infection is markedly influenced by moisture, Plants placed in
a satul'Uted humidity with the proper temperature and light show
a high pel'centa(l'e of infection,
The bean-blight organism is widely distributed with infected seed.
Local di";:emination of the parasite may be brought about by dew:
34 CI.'l~CnNlCAL BULLBTIN 186, U. S. DEPT. OF AGRICULTUHE
rain, hail, ,rind, bean straw, insects, surface drainage, and irrigation
wuters. The impottance of seed transmission is increased when the
practice of inoculating the seed with a water suspension of the root­
nodule organism, Bacill'u8 l'adicicola, is employed, owing to the fact
thut this method of moistening the seed spreads the blight parasite
it'om !L few infected seeLIs 0\'01' the entire lot.
Leaf infeccioll1:i are stolltatal. Bacteria then invade the interce1­
lulnr spaces, causing a gradual dissolution of the middle lamella.
1../l1ter cell disintegration takes place, with the formation of bacterial
pockets. Stem infection occurs through the stomata of the hypo­
cotyl und epicotyl, through the vascular elements leading frolll the
leuf to the stem, or from infected cotyledons, Bacteria in the )..'YleIll
\'essels may cause a wilting of the plant either by plugging of the
vessels Ot' by disintegration of the cell walls. Little infection is
found in the secondary xylem becaus~ of the composition of the waU
material.
Experimcntal evidence shows that the pathogene is harbored below
the seed coats. The organisms pass into the sutures of the pods from
the va~cular s)'.\ih'lll of the pedicel and make their way into the
funiculu.\i and thence through the raphe leading into the seed coats.
Another method of enll'Y into the seed is through the micropyle. No
case of direct penetration through the seed coat has ever been
observed.
Thc bacteria in the seed coats either remain there or pass into the
region of the cotyledons and enter these structures when the seed
germinates. nifts in the epidermis of the cotyledon are formed after
germination because of the increllsed size of the cells, Bacteria make
their way through these tears, pass into the intercellular spaces of
the cells below, and finally invade the entire cotyledon. Entrance
might be made into the Yllscular elements whence infection of the
young plant takes place.
Microchemical tests have shown that aiter germination much of
the cotyledonary tissue becomes soluble, and bacterial action is prob­
ably influenced to a great extent through the solubility of material.
The data on varietal susceptibility to bacterial blight were col­
lected under field conditions for three successive summers. No
variety showed complete resistance i however: 4 varieties of the
Refugee type showed a high degree of resistance, 19 showed medium
resistance, and 12 sho;wed little or 110 resistance.
LI'f'ERATURE CITED
(1)
BACIDfANN, 1"'.
H1l3. TilE MIGRATIOX
m' BACILLUS AM"l'"LOVOI\CS IX
THE HOST TISSUES.
Phytopathology 3: [3]-13, ill US.
(2) HAnsS, H. P.
1021. BE:Az..- BLIGilT AND BEA..": MOSAIC. Oreg. Agr. Expt. sta. Crop Pest
and Hort. Rpt. (1915-20) 3: 192-194, illus.
(3) BE:Acn, S. A.
1802. SOlfE BE:AN DISEASES. N, Y. Stute Ab'T; Expt. Stu. Bul. (n. s.)
48, p. (308)-333, mus.
(4) BURKHOLDER, W. H.
1018. THE 1.'1I0DUCTION OF AN ANTHRACNOSE-RESISTANT WHITE MARROW
llt:AN. Phytopathology S: (353J-35H, ilJu;;.
(5)
1021. TlIE I1ACTERIAL BLIGUT OF THE BEAN: J\. SYSTEMIC DISEASE. Pllyto.
pathology 11: [61]-69.
35
BAOTERIAL BLIGHT OF BEANS
(6) rWIlKlIOl.DElt, W. H.
102-!. V.Ilnt:rAL susm-:t'TUm.l'J'Y .UIOXO IJEA;SS
Phytopathology 14: [1]-7, iIlus.
1026.
TO TUE BACTERIAL DLIGIIT.
NEW HAOTElUAL !lISEASE OP TIU} m:.AN. Phytopathology 16:
015-027, iIlus.
(S) DAUWIN, x'.
lS!JS. OHSI·:lt\'.ITW:"'1' o=" ST(J~l \T.\.. Ltor. Soc. [London] Phil. Trans. (B)
J
fi:n-ftll.
(0) DELAcRorx, [G.J
A
no :
1S[l0. lJA
(lJLI1SSEl,
~L\lJAIlIE HAC'I'(mIENXt, DES IU1tICOTS.
Compt. Rend.
Acad. Sci. [Paris] 12U: 6:>G-G5D.
(10) DOIDGE:, E. ~r.
1l>18-11>. 'I'llt; llACTt::lUAL llI.IOIIT OF lIR.\NS: IJACTE1tfU)[ PUASEOU Ell\\'. S~r.
SO. AfriCllll Jour. Sci. 15: 50a-505.
(11) EOOERTON, C.
/llld "fORELAND, C. e.
IH13. TIU: IIBAX HI,WIlT .INIl l·IW..s~-:It\'.ITION AND TREATMBNT OF nF_~N SEED.
La. Agl\ Expt Stl!. Bul. 13!), 43 p., iIlus.
(12) EU-lOTl" C.
1020. 1I.\.I.o-m.IGIIT m' OATS. Jom', Agt·. Resem'cll 10: 139-172, mus.
(13) FUVfON, II. R.
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