Download Experimental and conceptual plant pathology

Chapter3
Are Bacteria-PlantCell Interactioris
Specific?
YOAV BASHAN
I. INTRODUCTION
Certain bacterialspeciesor pathovarcan-attack only one speciesof plant
(compatiblecombination) with different virulences-frorn high virulenceto
avirulence-while plant reactionsgo from highly resistantthrough various
degreesof tolerance,to completesusceptibility.Therefore,.thetype of relationship betweenthe host and the parasite,host-specificity,is difficult to
define. It can be a generalrelationshipbetweenbacterialspeciesand plant
species,or a narrowerone betweena bacterialstrain and a plant cultivap.
Nevertheless,
eventhis last definition is stil.ltoo largeasmany bacterialstrainplant cultivar systemswill haveseveralkinds of specificand somenon-specific
combinationsas well.
Severalf'actorscan determinespecificity.The main onesare: recognition
or non-recognitionbetweenthe pathogenand the plant, ability of the invading bacteriato multiply in the plant tissueand responseof the tissueto
the presenceof bacteria.
The term 'recogpition'also hasno absolutedefinition: Clarke and Knox
(1978)definedit as an 'initial eventin ceH-cellcommunicationthat elicits
a definedbiochemical,physiologicalor morphologicalresponse',in whicb
'initial event' and 'communication'
are not defined in terms of plant
pathology; while Sequeira(1978)restrictedthe meaningof recognitionto
'an early specificeventthat triggers
a rapid overt responseby the host, either
facilitating or impedingfurther growth of the pathogen'.
Plantsareliving in a hostileenvironmentof bacterialpathogensand support a largepopulationof microorganisms
on eachof their parts.Flowever,
asinfectionsand diseases
arenot the mostcommonphenomena,plantsmust
somehowget rid of the potentialpathogensby severalmechanisms,which
aresupposedto dependon recognitionbetweenthe plant and the pathogen.
Severaltheoreticalmodelsdescribecell-cellrecognitionin plant pathology
as the sequenceof eventsgiven lelow. A sensingsystem,prabably the
patible pathogen.
includeattachmentto, and encapsula
The plant defencemechanisms
by the plant cell wall, or inhibition by plant metabolites.The virulence
a phytopathogenicbacteriumis definedasits ability to escapefrom the pla
cell attachmentor its insensitivityto plant toxic materials.However,if t
pathogenis incompatible,the bacteriausually induce a hypersensitivere
tion in the plant.
This chapter has severalpurposes:to describethe interactions betw
plants and bacterial pathogen at the cell level, to summarizethe curre
knowledgeon cell surfacerecognition and multiplication of pathogenica
of th
non-pathogenicbacteriain plant tissue,to assessthe consequences
a
different
interactionsinsideand outsidethe host cell and to evaluatethe
time-changingconceptsin this subject.
BETWEEN BACTERIAAND PLAN
2. RECOCNITION PROCESSES
2.I
ADHERENCE
OFBACTERIATO SUNTECNS
NoN-SpBCTTIC
A bacterium,eitherpathogenicor saprophytic,reaches,at random,its tar
plant with the help of living vectorsor by its own motility as a resul
chemotaxis(Chet e/ al., 1973;Raymundoand Ries, 1980;Venette,19
Onepossiblemechanism,by whichthe bacteriumcan adhereto plant s
faces, dependson the balancebetweenmutual repulsion forces-as b
bacteriaand plant cell walls are negativelycharged(Fletcheret ql., 1980
and van der Waals' attractionforces(Pethica,1980).Marshal(1980)de
'the phaseof reversibleso
ed this temporarilynon-specificinteractionas
conditionsbacterialcells,very clos
tion'. In thesevery looseadherence
plant
with
their
flagellar action. Flagellarmoti
can
move
away
the
cells,
play
pathogenic
role in 'successful'inf
seems
to
an
essential
of
bacteria
tion (Panopoulosand Schroth, 1974).The different degreesof attachm
betweenbacteriaand plant cell wall seemto be dueto extracellularpolym
materialsbridging to and interacting with complementarystructures(M
shall, 1980).The specificityof the interaction,if thereis any at this sta
dependson the mutualaffinitiesof the cellstructures.Theseinteractions
be betweenlocal sitesof oppositecharges(ionic concentration),by mut
exclusionof incompatiblemoleculesto give an increasedlocal concentra
of both polymers,or by energeticallyfavorableassociationof structur
compatible chain segments(Fletcher et al., 1980)
Although, adherehcestudiesof phytopathogenicbacteriaassuchare qu
rare, a good examplecan be found. Lebenand whitmoyer (1979)sho
that seven pathogenic and non-pathogenic bacterial genera adhered
approximatelythe sameextentto young cucumberleaves.Even Escheri
coli and Serrotiamarcescens,which do not belong to the natural cucum
2.2 SpBctpIcRpcocNlrroN BETwEEN
aNo PleNrs
BACTERTA
Specificcontact betweenbacteria and plants can occur at thp extracellu
level or later at the intercellularlevel, and is in generala prerequisitefor c
cell recognition(Sequeira,1978;Stall and Cook 1979;Vance, 1983).T
interactioncanbe eithercompatibleor incompatible.A compatibleinteract
resultsin successfulpenetration,establishmentof the bacteriainsidethe h
tissue and multiplication of the bacteria to massivepopulations. A
incompatible interaction results in hypersensitiveresponse,phytoale
accumulation, pathogen immobilization or reduction of bacte
multiplicationto a minimum. This happens4sa reactionof hostplantsw
the most incompatibleplant pathogenicbacteria.
One of the most specificinteractionsis betweenRhizobiumsp. and th
wh
legumeplants,wherespecificrecognitionmechanisms,
corresponding
promotehost infectionby bacteria,havedeveloped(seeVolume2, Chapte
In general,thereis relativelylittle evidenceindicatingthat suchrecognit
mechanismsalso occur betweensusceptibleplants and their compat
pathogens;one known exampleis tumor induction in dicotyledonouspla
by Agrobacteriumtumefaciensin which positivehost recognitionis associ
with compatibility.On the other hand, specificrecognitionof pathoge
bacteriaby plantseventuallyleadsto incompatibleinteraction.Plant-bact
recognitionprocesses
occurwithin a few hoursafter inoculation.The cellu
events which actually restrict pathogen multiplication in incompati
interactionoccur much later, as it takestime for the tissueto changet
basic metabolism of the infected cells by manufacturing products wh
should respondto the presenceof the pathogen.
Compatibility betweenpathogenicbacteria and their hosts is indica
mainly by production of extracellularpolysaccharides,toxins, indoleac
acid and commonantigenicrelationships.Many other additionalfactorsw
different degreesof importancecontributeto the developmentof the disea
Currently, all the mechanismsknown are constitutive requirementsfor t
pathogenicityof a certain isolate on a plant species.However, they are n
directly relatedto cultivar specificpathogenicity(Keenand Holliday,198
Pathogenic bacteria lacking one or more factors required for this ba
compatibilityareconsideredto be avirulent.Theseavirulentstrainswill indu
several disease-resistance
mechanismsin the plant against the invadi
bacteria.
Plant defence mechanisms which may control specific recognit
mechanismsare describedas follows:
(a) Protectiveresponsewhich occursonly in solanaceousspeciespreve
multiplication of compatible or incompatible bacteria in the tissue af
plants againstincompatible bacterial pathogens(seeSection 3,Chapter
(c) Immobilization and encapsulationof bacteriaare plant reactionsagai
saprophytic 4nd avirulent strains. It is assumedthat the encapsulation
bacteria inside host tissuepreventstheir multiplication. Thesemechanis
will be discussedin details later.
(d) The phytoalexinsare antibiotic compoundsproducedby the plant,
a reactionto the presenceof pathogens.Only slight evidenceof phytoale
production in bacterial-plantsystemsexists.The role of thesecompound
discussedin Chapter 5 of Section 3.
The importanceof cell-surfacecarbohydratesdeterminingrecognitionh
increasedconstantlyduring the last decade.More structural variations c
be obtained from complex carbohydratesthan from any other molecu
suchas proteinsand nucleicacidswhich are availablein the plant cell. On
a small changein the structureof cell-wall carbohydratesmay resultin la
effects on recognition specificity. Phytopathogenicbacteria and plant ce
wall interaction can be used as an extraordinary model, becausetheir fi
attachmentor contactis at cell-walllevel. Thus, it can be generallypropos
that surfacecarbohydrateson the cell surfacesof both plant and bacte
may functionally participate in the recognition phenomenon.
OF BACTETABY PLANTPRODUCTS
2.3 AGGLUTINATION
The first step of the plant reaction against the invading bacteria
agglutinationof the bacteriausually to the plant cell wall. Latet, structu
.h*g.r, not clearly defined and perhaps associatedwith plant activiti
the bacteria(Goodmanet al., 1976a, b; Sequ
immobilizeand encapsulate
et al.,1977). The first agglutination factor, discoveredby Berridge (192
was later characterizedas potato lectin which agglutinatesonly avirul
Pseudomonassolanaceorumcells(Grahamand Sequeira,1976;Sequeiraa
to be an hydroxyproline-richprotein w
Graham, lg77).ltis now suggested
high,agglutination activity ([.each et al.,1982 a, b; Duvick and Seque
1984a, b).
Sincethen, various other agglutinins, of different chemicaland phys
have been discovered.Agglutination of pathogenicbacte
c'haracteristics
in apple tissue was first reported by Fluang et ol. (1975). Inoculation
virulJnt and avirulent strains of Erwinia amylovora resultedin localiza
of the avirulent strain by small granules of plant'origin, in the xyl
parenchymacells..The granulesstuck the avirulent bacteriain clusterswh
iysed rn situ, anditopped further multiplication of the bacteria.At the sa
time, the virulent strain multiplied freely and eventuallycausedsympto
of diseaseto appear in the apple tissue. Agglutinatlon in vivo was a
observed Uy Hoiino (19?6) in rice leaf xylem vessels,inoculated w
T
alsobind non-specificallyto polyanionssuchascarboxymethylcellulose.
may suggestthat polyanions of avirulent cell wall surfacesare acting
receptorsfor the apple agglutinin.
2.3. I Extracellular polysaccharide
In addition, Hsu and Goodman (1978 b) have discoveredextracellu
polysaccharideagglutininsof bacterialorigin. Apple-cell-suspension
cultu
inoculated with virulent p. amylovoraproduced a factor(s) which agglutina
(1979),sugge
cellsof avirulentE. smyloyora.Goodmanaia his associates
that the appleagglutinin and amylovorin, a toxin producedby the pathog
were similar, becausethe toxin also has an agglutinatingcapability. Deta
of this extracellular polysaccharidesystem will be provided later in th
chapter.
2.3.2 Lectin and hydroxyproline-rich glycoprotein bacterial agglutinin
Convincing evidencefor the presenceof the agglutination phenomen
was shown by Sequeira and Graham (1977) investigating potato lec
agglutination of many virulent and avirulent strains which representt
variability and sourcesof the pathogenP. solanacearum.Good correlat
betweenavirulenceand agglutination was achieved.Thirty-four avirule
strains were agglutinatedby. the lectin while 55 virulent strains were on
slightly agglutinatedor not at all. The virulent strains of E. amylovora a
P. solanacearumarenot agglutinated,probably becausethey havediffere
amounts of extracellularpolysaccharidesin their capsule.Virulent stra
have thicker capsules,which preventagglutination, whereasthe absenc
extracellularpolysaccharides
in the avirulent strainstriggersthe agglutinat
activity of the plants. Immunofluorescentstaining revealedthe presenc
lectin in the mesophyllcell walls in potato and tobacco. It seems,therefo
that a lectin-binding attachmentoccurs in the P. solanaceorurndiseas
potato. A further purification of the potato lectin (Leach et al., 1982a
leadto the discoveryof hydroxyproline-richglycoproteinbacterialaggluti
(HRGP), which, however, did not have the haernagglutinin activ
characterizinglectins. Mellon and Helgeson(1982)isolatedthe sameHRC
from a suspensionof tobacco cells,which also agglutinatethe avirulent
solanocearumisolatebut not the virulent one.Theseresultssuggestthat bin
ing is not hapten mediatedas in lectins, and sinceHRGP is widely pres
in higher plants it cannot be expectedto play a major role in determin
high specificity.
2.3.3 Pectic-polysaccharideagglutinin
Another type of agglutinin, which definitely doesnot determinespec
recognition, was found by Slusarenkoand Wood (1981).They isolated
of the virulent strains.Extraction of susceptiblebeancotyledonsdid not yi
the samefraction of agglutinatingmaterial. Pecticpolysaccharides,
wh
arewidelydistributedin plant cell walls,may indicatethe possibilityof no
specificattachmentin vivo of incompatiblepathogensto the plant. T
materialcould be the substance
observedby Hildebrandet al. (1980),wh
dissolvedfrom cell walls and trappedthe bacteria.Anyway, it is likely th
the pectic polysaccharideagglutinin is not responsible for the spec
resistanceof the resistantbean cultivar to the pathogen.
2.3.4 Other agglutinin-like compounds
The agglutinin-likecompoundsfrom beanleaves(Andersonand Jasalav
1979), can be of three kinds:
(a) Water homogenatesof leaveswhich causeagglutinationof saproph
pseudomonadsbut not of pathogenicP. syringae pv. phaseolicola.
(D) NaCl-wall releasedagglutinin which causedvariableagglutination
either pathogenicor saprophyticpseudomonads.
(c) Plant compoundssuchaspectin, galacturonicacid and severallecti
which agglutinatedpseudomonadscells non-specifieally.
The data presentedabove, are hardly connectedto the specificity of
agglutininsto the bacterial cells, as it is not numerical, and the agglutin
wereonly partially purified. Thus, someof the resultsmentionedaboveco
be relatedto residuesof other substances
which wereshownin other pla
bacteriainteractions.Andersonand Jasalavich's(1929;interpretationis u
conventionaland statesthat preliminary binding of bactetial cells to pla
cell walls would be initiated non-specificallyby the pectin compounds
a secondaryphase,tictin UinAng would determinewhetheror not the bacte
remainedbound. The failure of virulent strains to bind to cell walls is d
to production of materials that cancelthe attachment effect of the lect
2.3.5 Conclusion
No definitive conclusionscan be drawn from the limited data availa
so far. The mode of action of leaf agglutinins remains highly speculat
They may possiblyrecognizespecificsurfacepolysaccharides,
but no sim
sugarhas beenshown to preventthe activity of the agglutinins. It could
that more complex carbohydratesmay be neededfor recognition to occ
Furthermore, none of the specificity-determiningagglutinins were hig
purified and most of the conclusionswere basedon crude preparation
different degreesof purification. On the other hand, someagglutininsse
to be abundantand, logically,thesecommonmaterialscannotdetermineh
degreesof specificity. Nevertheless,the possibility that someagglutinins
determine some degreeof specificity should not be discarded.
of the virulent strains.Extraction of susceptiblebeancotyledonsdid not yield
the samefraction of agglutinating material. Pectic polysaccharides,which
arewidelydistributedin plant cell walls,may indicatethe possibilityof nonspecificattachmentin vivo of incompatiblepathogensto the plant. This
materialcould be the substance
observedby Hildebrandet at. (tl8O;, wtrich
dissolvedfrom cell walls and trappedthe bacteria.Anyway, it is likely that
the pectic polysaccharideagglutinin is not responsible for the specific
resistanceof the resistantbean cultivar to the pathogen.
2.3.4 Other agglutinin-like compounds
The agglutinin-likecompoundsfrom beanleaves(Andersonand Jasalavich,
1979),can be of three kinds:
(a) Water homogenatesof leaveswhich causeagglutinationof saprophytic
pseudomonadsbut not of pathogenicP. syringae pv. phaseolicolo.
(b) NaCl-wall releasedagglutinin which causedvariable agglutinationof
either pathogenicor saprophytic pseudomonads.
(c) Plant compoundssuchas pectin, galacturonicacid and severallectins,
which agglutinatedpseudomonadscells non-specifically.
The data presentedabove,are hardly connectedto the specificityof the
agglutininsto the bacterial cells, as it is not numerical, and the agglutinins
wereonly partially purified. Thus, someof the resultsmentionedabovecould
be relatedto residuesof other substanceswhich were shown in other plantbacteriainteractions.Andersonand Jasalavich's(1979)interpretationis unconventionaland statesthat preliminary binding of bacterial cells to plant
cell walls would be initiated non-specificallyby the pectin compounds. In
a secondiryphase,le'ctinbinding would determinewhetheror not the bacteria
remainedbound. The failure of virulent strains to bind to cell walls is due
to production of materials that cancelthe attachmenteffect of the lectiil.
2.3.5 Conclusion
No definitive conclusionscan be drawn from the limited data available
so far. The mode of action of leaf agglutininsremains highly speculative.
They may possiblyrecognizespecificsurfacepolysaccharides,but no simple
sugarhas beenshown to prevent the activity of the agglutinins. tt could be
that more complex carbohydratesmay be neededfor recognition to occur.
Furthermore, none of the specificity-determiningagglutinins were highly
purified and most of the conclusionswere basedon crude preparationsat
different degreesof purification. On the other hand, someagglutininsseem
to be abundantand,,logically,thesecommonmaterialscannotdeterminehigh
degreesof specificity. Nevertheless,the possibility that someagglutininsdo
determine some degreeof specificity should not be discarded.
of specificsugarstructures.Somerecognition systemshave beenattribute
to lectin recognition;the best exampleis the rhizobia-legumesymbiosis
2.4.1 Legume-rhizobiuminteraction
Selectiveinfection of certain legumespeciesby certainRhizobium speci
canleadto the formation of nodulesin the plant root system.The transform
ed bacterialcells(bacteroids),are fixing atmosphericnitrogen for the benef
of the two participantsof this symbiosis.Various degreesof positivehos
specificityare known in theseplant-bacteriacombinations.
After the establishmentof rhizobia cell in the legumerhizosphere,infec
tive rhizobia adhereto the root-hair surface. Severalselectiveattachmen
of infective rhizobial strains (compatible),but not of non-infectivestrains
have beenproposedto explain infection specificityin the symbiotic interac
tion. The attachmentwas first attributed to specificbinding of bacteriato
the root lectins.
Bohlool and Schmidt(1974)were the first to show a strong correlatio
betweenbinding in vitro of soybeanseedlectin to R. japonicuz cells an
the infectivity of thesecells.Lectin-mediatedattachmentwassimilarly show
in R. trifulii-clover interaction in which there was perfect correlation be
weenlectin binding in vitro and infectivity. Someof the symbiosishasa hig
degreeof host specificity, so that restrictedgroups of legumesare infecte
only by a singlespeciesof Rhizobium,e.g. R. melilotiinfectsand nodulate
alfalfa roots but not clover, soybeanor chickpearoots. Host specificityin
this caseis the earliesteventoccurringin this symbiosis.
Lectin recognitionmodel. A significantcontribution to the understandin
working with R
of the symbiosiswas made by Dazzo and his associates
trifolii-clover symbiosis.They showedthat surfaceroot lectinsbind rhizobi
by complementarysurfacemoleculesasan essentialand specificprimary ste
of the associatioi(Dazzoand Hubbel, 1975;Dazzoet al., 1916).The lecti
recognitionmodel they suggest,survivedvery intensiveinvestigations,bu
as yet is still unproven.They simply proposedthat specificlegumelectin i
bindingto a uniquecarbohydratestructurefound exclusivelyon the surfac
of the compatiblerhizobialsymbiont.Their later resultsare consistentwit
this hypothesis.They discoveredtrifoliin-A, a unique clover root-hair lec
tin, which seemsto be a cell recognitionmolecule.This glycoproteinspecifica
ly binds to, and agglutinates,the clover symbiont R. trifolii. The lecti
accumulateson the surfaceof the root-hair regionof clover, beingin greate
quantity at tlie growingroot-hair tips (Dazzoet at., 1978).The bacterium
which caninteractspecifica
R. trifulii, producesa capsularpolysaccharide
ly with the host lectin. Adsorption of cellsof non-infectivestrainsof th
compatibleR. trifulii or infectiveincompatibleR. melilotito cloverroot hair
was four or five timeslessthan adsorptionof compatibleinfectiveRhizobium
receptorsite.The biologicalexpressionof hostspecificityaffectsthe fre
of root-hair infections (Dazzo et al., 1984).
Receptor sites.The very important discoveryof the receptorsites
legumeroot, which specificallyrecognizethe compatiblerhizobia, was
by testingthe binding site of bacterialpolysaccharides.Theserecept
aresituatedonly on the root-hair, wherethe host lectin is accumulatin
are saturable, specific for lectin-binding polysaccharidesand spec
blocked by the appropriatehaptenof the host lectin (Dazzoand Brill,
1979;Kamberyu,1979; Gatehouseand Boulter, 1980;Kato et al., 1980
Dazzoand Truchet, 1983).
Bocterial polysaccharide structure. The exact molecular struct
bacterialpolysaccharidesshould be establishedto understandtheir s
interactionwith the host lectin. Thesecompoundsare extremelynon-u
and contain many different subcompounds(Dudman, 1984).For ex
the completestructureof any of the Rhizobium-hpopolysaccharide
is
unknown, perhapsdue to their exceedinglycomplexsugarcompositio
tracellular polysaccharideof R. trifoliiis built from carbohydratean
carbohydrate components such as acetate, pyruvate, succina
3-hydroxybutanoicacid (Hollingsworth et al., 1984).The pyruvate c
tuents are strong immunodeterminantsof rhizobial polysaccharide
man and Heidelberger,1969).It seemsthat the direct interactionbe
lectin-bindingsurfacepolysaccharides
of rhizobia and root hairs of the
legume is a characteristiccellular recognition on the molecular leve
Evidencesogairct the lectin-polysaccharidemodel, Although the
polysacchariderecognitiontheory seemswidely supported,someres
not fit the proposedmodel, due to the following difficulties mentio
Bauer(1982)and Vance(1983):(a) lack of a perfectcorrelationbetwee
ing to host lectin, and nodulatign (Schmidt, 1979);(D) availability of
in legumeroots is well documented(Dazzoet al., 1978 Gadeet al.,
although some studieswere performed with lectins from seedswhich
not presentin roots, but, somesoybeanvarietieswhich were found t
lectin, neverthelessnodulated very efficiently; (c) binding of incomp
rhizobia to roots is a not infrequentand rare phenomenon(Bauer,
The strongest evidenceagainst the role of lectin in determining s
recognition (Pull ef a/., 1978)is that infectiveR. japonicun nodulate
of soybeanlines lack leqtin. Pueppkeand Hymowitz (1982)screened5
of the subgenusGlycine to detect soybeanseed lectin, and showe
although the roots nodulated, none of the lines containedthe specific
which was supposedto determinerecognition.Thus, it seemsthat lect
not essentialin the recognition of rhizobia. This non-acceptancewas
out by Tsien et a/. (i983) who showedsmall quantitiesof lectin in se
in developingseedlingroots of the samesoybeanlines. Furthermore
bean roots containedadditional lectin which can bind R. japonicun s
cells by lectin may be only one of the specificity signalstransmitted from
the bacteriumto its plant symbiont. In his opinion, Dily signalsof differing
naturesmay be requiredfor the developmentof the complexlegumenodule
Bauer (1981) proposedthe idea that plant receptorsother than lectin
recognizea specific component on the bacterial cell surface.
Conclusions.Dazzo et al. (1984) explain these contradictory results a
follows: (a) somerhizobia strainsproducemore than one polysacchari
which bind the lectin (Hrabak et ol., l98l) and sorneof thesepolysaccharid
changetheir compositionwith the ageof the culture(Mort and Bauer, 1980
Cadmuset al., 1982);(D)the ability of the rhizobiato bind to the lectin is
a transientevent(Mort and Bauer, 1980;Hrabak et al., l98l); (c) the lecti
is availableon the root surfacefor a transientperiod of developmentof the
attachment(Paauel a/., l98l); (d) the environmentalconditions,prevailin
during recognition, affect the expressionof the lectin receptor on the
bacterium(Bhuvaneswari
and Bauer, 1978).In summary,failure of one o
lectinsto interactspecificallywith the homo
the severallegume-produced
logousrhizobiadoesnot automaticallyrejectthe lectin recognitionhypothes
which remainsa reasonableexplanationfor the specificrecognitionbetwee
rhizobia and legumes. Despite the undetermined role of cell-wa
carbohydratesin specificity, the basic bacterial cell surface architectureof
Rhizobium is'now better known.
2.4.2 Lectin-phytopathogenicbacteria interaction
In contrastto the relativelyvastamountof knowledgeconcerningthe role
of lectinsin the legume-rhizobia
interaction,very little has beenpublishe
on the interactionsof lectinsin plant-pathogenic
bacteriainteractions.Sin
and Schroth(1977)showedagglutinationof saprophyticbacteriaby a possib
lectin. Sequeiraand Graham (1977)suggestedthe involvement of lectins in
agglutination of P. solanacearumby potato. Sequeira(1979)showedtha
tobacco and potato lectins precipitated bacterial lipopolysaccharid
Precipitation was inhibited by chitin oligomers.This phenomenonindicate
inside the LPS. Thi
that lectins may recognizeN-acetylglucosamine
recognition activity of lectin was refuted after the discovery o
hydroxyproline-rich agglutinin within the lectin fraction which has a
remarkably strong agglutinating activity (Leach et al., 1982 a, b)
Additionally, Anderson and Jasalavich(1979)showed that bean lectin
agglutinateboth pathogenicand saprophyticpseudomonadsto roughly the
sameextent
Ghanekarand Perombelon(1980)disagreewith the hypothesisof lecti
participation in recognition. They reported that about 25 different Erwinia
spp. isolateswereagglutinatedwith purified potato lectin irrespectiveof thei
origin and of their pathogenicityto potato. Similarly,no clearpatternswer
found when other pathogenicor non-pathogenicbacteriaof severalgener
plartt diseases.
However,severalother pathogenicsystemswhich are hig
specific such as P. syringaepv. tomato on tomato, and X. campestrisp
vesicatoriaon pepper should be testedfor the possibility that lectin wou
play a role in determining their specificity.
2. 5 ExTnacELLULARPor-vsnccnARrDEANDLrpopoLysAccHARIDE
INtpnncrroNs
Although, extracellularpolysaccharides(EPSs) and lipopolysaccha
(LPS$ weresuggested,
in the seventies,
asplayinga role in cell-cellrecognit
processes,
their importancehas becomesignificantonly recently.They a
Neverthel
now assumedto play a major role in recognitionmechanisms.
the mode of action of some interactionsis not understood,most of t
proposedsystemshave not been firmly establishedand most of the ba
questionsare still open.The possiblenatureof the cell-cellactivitiesof the
complexcompoundswill be despribedbelow.
2.5.1 Definition and characteristicsof EPSand LPS
It can adh
EPS is generallydefinedasa bacterialcapsulepolysaccharide.
firmly or looselyto the bacterialcell, and presenta great variety both
compositionand in structureat the bacterialcell surface.The exactfunct
of EPSsin Gram-negativebacteriais clearerin animal tissue-whereit he
bacterial survival and adherence-thanin plant tissue. LPS, although a
complexbut easierto isolate,is a more solid structurewhich is contain
in the bacterialcell wall.
One of the best examplesof the complexity of definition of the role
EPS in the recognitionprocessand in virulence,is the.E amylovora-a
They discove
systemdescribedmainly by Goodmanand his associates.
amylovorin, a todc low-molecular-weight,host-specificgalactose-richEP
affectingcellularmembranes(Huang and Goodman,1976;Hsu and Goo
man, 1978a).Amylovorin was proposedas beingresponsiblefor the init
tissue.But Sjulin a
ultrastructuralchangesobservedin infectedrosaceous
Beer(1978)did not confirm theseresults:they detectedno increasein el
trolyte leakagefrom amylovorin-wiltedtissue,indicatingthat no signific
changesin membranepermeability had occurred. They concludedth
amylovorininducedwilt by restrictingthe movementof waterin the xyle
amylovorinappearsto be an ess
and it did not act by toxicity. Nevertheless,
tial pathogenicfeature, since non-pathogenicisolatesare nof able to pr
duceit. Otherpolysaccharides
alsoproducedby the bacteriumwereisolat
but wereantigenicallyide
they weredistinguishedby immunoelectrophoresis
tical to lipopolysaccharides(Sijam et al., 1983)
The isolationof EPSsopeneda new approachto research:the possib
which arecapsularmaterials,areinvolvedin virule
that polysaccharides,
EPS and virulenpe.Sijam el al. (1983\showeddoubtson
they showedthe production of severalpolysa
correlation;nevertheless,
charidesby E. amylovora,both in culture and in plemttissue,which ha
the sameproperties,but were not identical
2.5.2 The EPS theory
electronmicroscopicpreparations,following freez
By usingvery detricate
cells,Politis and Goodman(1980)detectedthe fin
E.
amylovora
of
drying
pathogen
EPS. They proposeda model consistingof tw
of
the
structure
polysaccharides:
looselytied and watersoluble,the other tigh
one
of
tlpes
They explainedthe failure to agglutina
water.
in
insoluble
ly bound and
(whichwasmentionedearlier)asfollow
E.
amylovora
of
virulent
strains
the
by the agglutinin,and thus preven
of
bacteria
binding
the
the EPS blocks
of the plant. A similar e
mechanism
induced
defence
the
of
the activation
planation wasprovided for the escapeofvirulent strainsof P. solanacear
from agglutination.All wild{ype virulent strainsproduceda greatamou
of EPS both in cultureand in the plant which seemsto determinepathogene
in this pathogen.Spontaneouschangefrom smooth (with EPS) to roug
(without EPS) bacterialcoloniesresultedin lossof virulence.Addition o
EPS to avirulent strainspreventstheir agglutination'and removalof EP
from virulent bacteriaresultsin their agglutination.Sequeiraand Graha
(1977)and sequeiraet al. (19'17)tried to explainthe lack of attachmen
'preventsbindi
productionof EPS by the pathogen,by sayingthat EPS
of the bacteriato a specificreceptorsite on the host cell wall'. This EP
becausethe
shouldhavespecificqualitativeand quantitativecharacteristics,
are slime-producingstrainswhich are incompatible.
The EPStheoryis stronglysupportedby Smithand Mansfield(1982)wh
studiedthe interaction betweenoat leavesand P. syringaepv. coronafacie
(compatible),P. syringae pv' coronafaciensvar. atropurpurea and P' sy
ingaepv. tabaci(incompatible)and P. fluorescerc,the saprophyticbacteriu
Quantitativeanalysisindicatedaccumulationof EPS around the compatib
pathogen but not around the incompatible pathogen, suggestingthat t
bacterialproductionof EPS preventsthe bacterialattachmentto plant c
walls. Theseresultssuggestthat EPS playsa significantrole in prevent
bacterial cell attachment.
2.5.3 Rolesof EPS and LPS
The role of bacterialLPS seemscomplementaryto the role of EPS.
isolatedfrom cell-freeculturesof P. syringaep
lipomucopolysaccharide
water
soakingof cucumberleaves(Keenand William
lachrymansinduced
Whatleyet al. (1976)showedthe i
Agrobocterium,
l97l). Working with
in specificrecognitionbetweent
portanceof bacteriallipopolysaccharides
plant.
pathogenand the
attachedrapidly to the walls of tobaccomesophyllce
while the virulent strainscontinuedto multiply in the intercellularspac
The avirulent strain was also capableof rapid aftachment to suspens
culturedtobaccocells.Bacteriawerestronglyattachedend-onand only e
haustivewashingswereable to significantlyreducetheir attaChment.to
t
plant cells.The maximum numberof bacteriaattachedto a singletobac
cell was as high as 8700.Attachmentwas inhibited by high ionic streng
favoured by low ionic strengthand supposedto be ion mediated.Thus, t
ionic conditionof the intercellularfluids seemsto be crucialin determin
the attachmentof bacteriato the cell wall. The attachmehtmechanismswe
affectedby high temperature,azide,EDTA and someantibiotics,sugges
an active metabolismplayrng a role in the attachmentprocess(Duvick a
Sequeira,1984a,b). The strong attachmentbetweenP. solanacearuma
tobacco cells may be due to pili binding!,sincethq avirulent strain is hi
piliated. The low piliated avirulent strain of the pathogenattachedloos
(Stemmerand Sequeira,l98l). Additional information on bacterialattac
ment to plant cells is given by hydroxyproline-rich glycoprotein o
tained from severalplants cell-walls. This agglutinin strongly agglutina
the avirulent strain of P. solanacearumand thereforeits highly basicnatu
hasthe capabilityof ionic interactionwith the negativelychargedEPSsa
LPSs, and maybeother negativelychargedmoleculesof the bacterialce
which can result in agglutinationinhibition of virulent strains (Leachet a
1982a, b; Baker et al., 1984).
The agglutininprecipitatesLPS from rough strainsand EPS from smoo
strains.Therefore,it seemsthat EPS producedby virulent strainsbindi
the.agglutinin, preventsthe recognition responseof the plant, characte
ed as LPS-agglutininbinding, allowing virulent bacteriato escape.
Hendrick and Sequeira(1984)selectedmutantsof virulent strain whi
have defectsin LPS and EPS structures.Even if the LPS composition
parts of the mutants was similar to that of the avirulent strain, they faile
to induce hypersensitivereaction. They were agglutinatedin the samema
ner as the avirulent strain. According to thesedata they concludedthat th
'hypersensitive
reaction-inducingpropertiesof the avirulint strainmust repr
sentalterationsin factors other than or in addition to thoseinvolved in EP
and LPS synthesis.'
In summary,Sequeira(1984)presentedthe following points:
(a) Bacterial attachmentmay be mediatedby the interaction of bacter
LPS with hydroxyproline-richglycoproteinson plant cell walls. Fromstudi
in vitro of the binding, attachmentappearsto be a result of charge-cha
interaction.
(b) EPSswhich inhibit the interaction in vitro probably do not have th
effectin the plant tissue,asinhibition occursonly at low ionic strengthswhic
are not found in the intercellular fluid.
(d) Pili seemto play a role in strong irreversibleattachments'
Different information about the role of LPS wasgiven by Romeiro et
(1981).E. amylovora-LPS-core-carbohydrate
structure,but not the O-cha
werespecificallyagglutinatedby plant products.The coreregionof LPS
Gram-negativebacteria is highly polyanionic, giving further confirmat
agglutinationof avirulentstrains.Howev
to the theory of charge-charge
they explainthe non-agglutinationof virulentstrainsby maskingtheir co
LPS With EPS.
The theory that LPS participatesin race-specificrecognition of comp
ble bacteriain certainplant speciesand in non-specificrecognitionof
compatiblebacteriain other plantsis not yet clear.The examplesgiven
at leasttwo different.tinds
this reviewindicatethat plants may possess
LPSsW
receptorsfon bacterialsurfacecarbohydrates,and bacteriapossess
multifo
to
the
due
and structures.Therefore,
different typesof appearance
appearanceof all surface macromolecules,both recognition or n
can be achieved.
recognitionprocesses
2.5.4 llrater soaking and EPS
Another possibleinterpretationof the role of EPS was suggeste
(1979-81).They relatedpathogenicit!'of so
Rudolph and his associates
phytopathogenicpseudomonads
and xanthomonadsto their productio
plants(
EPS, which specificallycauseswatersoakingonly in susceptible
Banobyand Rudolph,1979b).The EPS wasinitially extractedfrom liq
cultures of P. syringae pv. phaseolicolaor from halo-blight-infectedb
stemsor leaves(El-Banoby and Rudolph, 1979a).The EPS was partia
purified (El-Banobyand Rudolph, 1980)and some of its biological a
physicalpropertiesweredeterminedbut it did not help understandingits m
of action (El-Barrobyet al., 1980).They suggestedthat the specific ef
of EPS may explainthe different virulencesshownby bacterialisolates.Th
was also a differencein plant reaction towards EPS from different sou
of the samepathogen.Therefore,they concludedthat the induction of wa
soakingby purified EPS explainsthe specificityofthe bacteriaat the spe
and cultivar levels, although the mode of action is still a mystery.
Bacteriil EPS and the visible specificwater soakingdisappearedfrom
intercellularspacesof resistantcultivar 12hoursafter inoculation,while E
remainedfor three daysin the intercellularspacesof susceptiblebeanlea
(El-Banoby et al., l98l). A probable explanationis that EPS was degra
enzymaticallyin the lesistantplants and not in the susceptibleones,and t
lost its primary capability to induce water soaking.
Theseeventsindicate the activation of the plant defencemechanism
molecularlevels(El-Banobyel o/.,1981).Ultrastructuralstudiesof the ef
of EPS within plant tissueshoweda closerelationshipbetweenwater-soa
cell prganelle.However,in this case,the possibilitythat EPS may preve
the attachmentof pseudomonads
to plant cell walls was not estimated
The understandingof the exactmodeof action of EPS is preventedmain
by the difficult task of determiningthe molecqlarand the sphericalstructur
of thesemolecules.This leadsto all kinds of assumptionswhich cann
satisfactorily settle whether or not EPSs have any relation to virulence o
specificity.
2.5.5 Minor ddferent evidencesof bacteria-pluntinteraction
Minor points of differencein the interactionbetweenbacterialfraction
and plants are presentedbelow. They are few, and their mode of action
not known. A necrosis-inducing-factor
of non-hostplant leaves,possib
glycoprotein,wasextractedfrom cellsof P. syringaepv. tomato.It did no
show pectolytic or cellulolytic activity and was destroyedby pronaseor b
hydrolysiswith HCl. Ilalso causedgreatlossof electrolyteswithin 48 hou
of infiltration which is too short time for compatiblecombinationsand to
long for hypersensitivity
reactions.Its role and significanceremainobscu
(Bashanet al.,1982b).
An 'endotoxin' from P. syringae pv. phaseolicola was also isolated.
inducedcellcollapseon non-hosttissue(Crosthwaiteand Patil, 1978).Som
protein-lipopolysaccharide
complexes(Mazzuchiet al., 1979),inhibit th
hypersensitive
reactionand causeslight chlorosis.Hypersensitive
reactio
inducingsubstances
from the bacterialcellswerealsoisolated(Sequeiraan
Ainslie, 1969;Gardnerand Kado, 1972;Sequeira,1976).
2.5.6 Conclwion
Despitethe new charge-charge
hypothesisexplainingthe role of EPS an
LPS in bacterialrecognition,sugar-specific
binding shouldnot be ignore
as ionizedgroups can influencelectin affinity for a receptor.
3. MULTIPLICATION OF PATHOGENIC AND NON-PATHOGENI
BACTERIA ON THE SURFACE AND WITHIN LEAF TISSUE
Nearly everyimportant bacterial pathogenhas beenmonitored for chang
in bacterial multiplication, before, during and after diseasedevelopme
(Henis and Bashan,1986).To clarify the phenomenonof multiplication on
representativedata on severalbacterialdiseases
will be presentedhere.Som
interpretationsof eventsare commonlyacceptedby most investigators;thos
which are not, will also be described.
In general, bacterial multiplication in plant tissue has the followin
characteristics:
(a) Most bacteria, either pathogenicor saprophytic, can multiply on th
surface of plant leaves,if available nutrients are supplied externally or b
the host plant.
can multiply freely both outsideand insidethe leaf, whereassaprophytic
bacteriaare usually restrictedto the externalside only.
(d) The population of pathogenicbacteriausually increasesmassively
during diseasedevelopment.After partial degradationof host tissue,or its
death, saprophyticbacteriacan colonizethe infectedarea.
(e)Both compatibleand incompatiblestrainsof pathogenhave,to a varying extent,the ability of endophyticmultiplication. In most cases,the incompatiblestrainsform smallerpopulations.The endophyticpopulation can
developin the air-filled intercellular spacesonly if theseare filled up with
water.
(fl Resistanttissuecannottotally preventpathogenmultiplication, but can
lower it by severaldegleesof magnitude.
(g) Saprophyticor non-compatiblepathogensappliedbeforethe pathogenic
strain, can inhibit its multiplication and preventdiseasedevelopment.
3.I
EpIpgyTIc Gnowrrr PHASEoF BACTERIA
Although leaf surface colonization by phytopathogenicbacteria has been
known from the beginningsof phytobacteriology,Crosse(1959)wasthe first
to show that the massive epiphytic population of P. syringae pv.
morsprunorum (the causalagentof bacterialcankerof stonefruit trees)was
presenton the surfacesof apparentlyhealthycherry leaves,and supposedly
suppliedan inoculumfor'initiation of the disease.Englishand Davis(1960)
also isolatedunidentifiedpathogenicfluorescentPseudomonasfrom healthy
peach and almond leaves,fruit twigs and weedsin the field.
3.1.1 Conceptof residentpopulation
Basinghis concepton theseresultsand on his studiesof thg 'basic' role
of epiphyiicpopulationsof pathogerricbacteriain plant disease,Leben(1965)
suggestedthat the pathogenlives and multiplies on the upper parts of the
apparentlysymptomlessplant. Thesepopulationsare referredto as 'resident'.
Under optimal conditions,suchasappropriatetemperatures,presenceof free
water, high relativehumidity and plant physiologicalconditions,the inoculum
developingon apparentlyhealthyplantsmay start a bacterialdisease,without
any further external supply of inoculum.
Some of the many studies supporting this concept are the following.
X. campestrispv. vesicatoriawasfound residenton tomato seedlings(Leben,
1963)and pepperplants (Bashanet al., 1985iBashanand Okon, 1986a,b).
Analyzing P. syringae pv. syringaein bean leavesrevealedresident phase
in nature (Leben et al.,1970). A highly virulent strain of P. solanacearum
had a residbntphaseon hairy vetchwhich is a common weedin beancultivation in Wisconsin. The pathogenwas found to be the main component of
the Gram-negativeepiphyticmicroflora throughout the year (Ercolani ef a/:,
3.L2 Non-residentpathogens
It seems,however, that not all foliar pathogenshave a residentpopulation on their host. Thomson et al. (1975)indicated that E. omylovora, the
causalagent of fire-blight in applesand pears,was not detectbdin healthy
buds from healthy or diseasedtrees.It wasdetectedon leavesonly after fireblight had spreadall over the orchard. Independentlyand at about the same
time, Sutton and Jones(1975),basingtheir resultson very extensivedata
suggestedthat the diseasebroke out from only temporary epiphytic population and not from a residentpopulation.Thesedataand their conclusionshave
a major drawback,as they could not monitor small enoughpopulations
3.1.3 Exchangesof epiphytic and endophytic bocteria
Epiphytic bacteria can be either pathogenicor saprophytic, whereasendophytic bacteria(definedas bacterialpopulation presentin the intercellula
spaces or in substorhatal cavities) are usually only pathogenic. These
characteristicsare, however,only indicative sincedynamic exchangesoften
occur betweeninternal and externalleaf tissue(Henis and Bashan,1986)
For example: epiphytic populations of P, syringae pv, alboprecipitan
penetrateinto sweetcorn leavesthrough closestomata and multiply inside
(Gitaitiset al.,l98l); epiphyticP. syringaepv.tomato prefersto penetrat
through open stomata and leaf trichomes (Bashanet al., l98lb; Schneide
and Grogan, 1977),whereasepiphyticX. campstrtspv. vesicatoriapenetrat
through leaf veins (Sharon et al., 1982a;Bashanet al., 1985b).Theseexchangesare highly dependenton the microclimaticconditionsinsideand outside the leaf.
Once the exchangehas taken place, the total bacterialpopulation is composedof epiphytic and endophytic populations, making it difficult technically
to distinguish between the two populations. Two of the techniques are
ultraviolet irradiation (Barnes, 1965) and.chemical surface disinfections
(Sharon et al,, 1982b).Hirano and Upper (1983)prefer to adopt a loose
functional definition of epiphytic bacteria as 'those bacteria that can be
removed from above ground plant parts by washing'. But, as they explain
'This definition doesnot distinguishbetweenresidentsand causals'(aswas
distinguishedby Leben, 1965)'nor doesit count all of the bacteria.However
it providesrelativelyrapid, quantitativeresultsthat can be usedasan estimate
of total populdtion size.'
3.tr.4 Diseasedevelopmentdue to epiphytic or endophytic bicteria
There is almost no doubt that the presenceof an epiphyticpathogeni
population can resultin initiation of disease.But is it an essentialprerequisit
for diseasedevelopment?To answerthis questionthree criteria were sug
gestedby Hirano and Upper (1983):
size.
(c) Quantitativediseaseassessments
should be done.
In general,during diseasedevelopment,particularlyafter artificial inoc
tion, the pathogenicpopulationtendsto increase.This growth is depen
on plant and pathogentypes and varies from lff to l0 colony form
units (CFU) per leaf or gram of tissue.
Someexamplesof foliar diseases
causedby bacteriaare: the incidenc
fire-blight diseaseof stonefruits which is correlatedto E. amylovora epiph
destructive population (Thomson et al., 1975);likewise the world's m
destructiveabiotic diseasecausedby ice nucleationbacteria(Lindow, 19
brown spot on snap beans to P. syringae pv. syringae (Lindeman
al ., l98l) and halo-blight on oats to P. syringaepv . coronafociens(Hira
et al., 1982).
Theseresultsshowedan importantfactor: the bacterialnumberthresh
With a populationnumberof pathogensbelowthe threshold,there are
visible symptomsof disease,abovethis number, symptomsappear.
weller and saettler (1980)showedthat, under field conditions,vis
diseasesymptomswere detectedwhen the population level of eithe
campestrk pv, phaseoli or X. tampeslns pv. phoseoli var. fuscans reac
5 x ls cFU per leaflet. They also found a good correlationbetweendis
severityand number of bacteriaper leaflet. Smitley and McCarter (19
presenteddata showinga relationshipbetweenthe populationnumbero
syringaepv . tomato and bacterialspeckdevelopmentin tomato plants. Th
data werevalid only whenthe environmentalconditionswerefavorable
diseasedevelopment.The major importanceof the climatic condition
bacterialspeckdevelopmentis well known and wasdemonstrated
in se'r
studies(Bashanet al., 1978;Schneider
and Grogan,1978;yunis et al.,19
3. 1.5 Statistical model
A statisticalmodelwasdevelopedto explainthe quantitativerelation
betweenepiphyticpopulationsof pathogenicbacteriaand diseaseincide
(Rouseet al., l98l). The model is affectedby many environmentalfact
susceptibilityof the host, bacterialnumbersand different ratesof bacte
multiplication factorsin a givenfield. As sucha modelrequiresa largeamo
of data (not yet availablein the literature),it shouldbe treatedin the me
time with some caution. Another drawback of this model is thar so
bacterialsystemsdo not follow the descriptionfor bacterialmultiplica
in plantsgiven in this subsection.under field conditions,no positivec
relationwasfound betweenthe numberof bacteriain pear flowersinfec
with E'. amylovoraand diseaseincidence(Miller and schroth, r972);no re
tionship was found betweenx. campestrispv. vesicatorianumbersin p
per plants and symptom expression(Bashanet al., l915a\.
asa resultor tn" ."..li[;:
;;ilfi
.o.pr.it"ntive
not Yet be made'
IN PLANTTISSUE
PHASEOTBECTNNTA
GROWTH
3.2 ENDOPHYTIC
of data
Maybeduetothetechnicaldifficultiesindistinguishing.theinternaland
t*li;;; t*"llt1 il"-:*
0"il;;;;mtnt'.?1*
e*ernalbacteriar
r *,,n"ur.;-;;;"i'4oY:';
poililrliil",
aboutendopnv,i.
l?oulation
ri't'" .ry fi 1t;li:1'3;:il;t'::'H;;:l
o"r
*' ouiJi'
|ll theinterceuur
"
Acompatibr.putnoJ.]'il;;t'hastheuniiueauiritytomultiplyfreelyi
;ttl*i**milili,l*?t$5i""ll:'"ru;iJ'"'*ilu'sh
'fi:
someexampleswilrJ"io"#?i..iJ"nnr,,.*itiin'oiiioiohel.o
-P'
ingthedeveropmeniii'uJirr.,.*,T,i*,{;}#i*,:;ll*i
the leaf' Bashan'1-1';-tT"*"iJi""".t,
syringoepv
bacteriawithin
;;;il
:1,:^:lllT; Xl;Kliliil;;;;'
hchrYmansrn cuc
pv
x-'campes
"na
vesicotoriaino.ool,'iJu".J.'r"ro,.*:Tt.ffii
(lh?:* jt^
1':'; ;;lroucation of ^thepathogen
, r" l,' tga,Danopepper
"
:ili:1#""':;{;ilh;ft
o
surraces
1l;lnlinru;:$ffi
tt ":r.'::*"Du
"t"'ht-o,l:'J
o*n ;;;;.",
111ii -1t;,1-*"*"i
ation,
i ncub
:r;tnn*n'"i*ti*Xlfixil
ponulauol-iii;;;,.til;*cktptg* ot"'
ontomato'th
or crn?leafarea.Epiphvtic
"dti'ts atntro.9"i1of bacterial
i-nil"-"tti
tcpcFu/e "'to'ii'
,:
il,::#f*:f.fii?Xf;iJi;'*pend
o"o''iiliJ;;fi
endcphytic
x. ,o^p"'rn1Pl;
;il;;;d
the#;:
on"i,
either
ry
bac
l"'"ai"::':i1::::"fi',h"r,.
thatw
"nv"ip"'ent
-e;
*:ll':::S$i(ffii'#ii*
Howevttl!?l
it ont sidetoJheother'
soakingoftheo*ot'leavesi'"titll"l;;;;;;;q"ttittfot'endophvtic
multiPlication'
di:'f': svmfloms'
o:!':::'o'once of
population
3.2.1 Endophvtic
'"',ll:i#'ry?;;il:,;',''!ffi
Endophvtico"o"'-"iii''mavn:t1*"1vJil''"'y1
weeninoculum
iffiH
Massive*a"ot''ircn"oJ;1"."":::1:$:li
di
Nevertheless'^t]1il"r";;JJ
scab
theleaves'
itretvpical
cauDesuv
was similar to tnal
""
population
aopt'vtic
;;;
1985a)'
iguttton et al"
host. After a certain time, the inhibiting effect of the incOmpatibleh
becameapparent, when the bacterial population increasedlessrapidly
decreasedmarkedly.Bacterial population within the compatible host c
tinued to increaseuntil destructionof the tissueoccurred. lheir last ob
vation was on the seventhday after inoculation. Leben et al. (1968 a
prolonged their observation of population dynamics of P' syringoe
glycinea. They showedthat the population reachedits maximum betw
ihe seventhand thg fifteenth day after inoculation. Later; the popula
declined as lesionsbecamenecrotic and leavesdried. Substantialnum
ofthe pathogenwerepresentbeforesymptomappearance'and low inocu
concentration resultedin lower population maxima and delayedsymp
expression.Theseresultswere confirmed for P. syringaepv. tomato an
compestris pv.'vesicatoric (Bashanet al,, 1978 Diab et al.' 1982\,
t iUen and his associatesspeculatedthat in older drying leavesand nec
lesions,baCteriaare 'glued dgwn' and serveas a slow releasereservoi
future bacterial outbreak. This was tested in bacterid speck and sca
tomato and pepper.usinga scanningelectronmicroscopytechnique.A
developmentof microscopicalnecrosis,bacteriawerenot found in the m
necrotic sitesexaminedvery carefully. Bacteriain both diseaseswere dete
only in apparently healthy infectpd tissuesurroundingthe necrotic atea. T
presumabiybacteriasimply diedwith the plant cellsin the lesionitself (Ba
et sl., l98lb; Sharon et al,,1982a).
3.2:3 Leqf tissueas o protective environment
Evidencethat the leaf tissueservesas a protectiveenvironmentfor th
dophyte bacterial population can be drawn from the following exam
Endophytic population of X. campesfnbpv. phaseolivar. soiensisin
bean ieavesshowedthat the ultraviolet irradiation destructiveto bact
cellseither in water or on the leaf surfaceshaveno effect on the interce
population (Barnes, 1965).Surfacedisinfestation of pepperwith the bleac
pv. vesica
compound,Naocl, which usuallykills most of x. campesfnls
(Sh
bacteria
endophytic
in water suspensiondid not reducethe number of
which
elimi
pepper
leaves,
et al., 1982b).Only lyophilization of diseased
nearly all the epiphytic microflora including the bacterialpathogen,ca
reduction in the internal population (Bashanet al.' 1982a).
3.2.4 Conclwion
The endophyticmultiplication of phytopathogenicbacteriashowsthat
ly all compatible pathogenstested can multiply inside plant tissue
vironmental factors have little influence, but water soaking of leaves
least non-wilting conditions axenecessaryfor proper multiplication in
tissue.Plant tissueseemsto protect the endophyticpopulation from the h
of the defencemechanismsof the plants, they do not seemto be assoc
with visible symptoms.
3.3 Mur.Ttpr.rcanroNoF BAcTERTA
IN RELATToN
ro Corr{p,lrrBrl-rr
Plantsare not alwaysdiseased,and diseases
comefrom different bac
species:
theremust,therefore,be somekind of specificitybetweenthe p
and the bacteriaat the tissuelevel which determineswhether or not dis
processes
will develop.Most studieson bacterialmultiplicationin plan
not distinguishbetweenthe epiphyticand endophyticbacterialpopulat
but concentrateon the effects of compatibility and non-compatib
measurements
which are easilycarriedout with a precisesamplingprog
homogenization device, selective media (if available) and very sim
techniques.
The main drawbackof thesestudies,especiallyin field conditions,is
multiplication is affectednot only by the cornpatibility of the host, but
by many environmental,agrotechnicaland methodologicalfactors, lea
to contradictoryresultsevenon the samebacteria-plantcombination.De
all the problems,these.studies
show,with not too many exceptions,that e
patible interaction favoursmultiplication of the pathogenin the tissue,w
incompatibilityrestrictsor slowsthe rate of multiplicationof the bac
in the plant.
The following examplespicked from.the bulk of literature,illustrat
aboveremarks:multiplication studiesof X. campestrispv . vesicatoriain
per by stall and cook (1966)showedthat after severaldaysthe pathog
population was alwayslower in the resistantcultivar. similarly, p. syrin
pv. tomato showedthe sametrends in bacterial population (Bashanet
I 98lb). P. syringaepv . p haseolicola grew fasterand to hilher total num
in the susceptiblethanin the tolerant beancultivar leaves(Stadtand s
tler, l98l); intensivemultiplication occurredalso in the resistantcul
(omer and wood, 1969).In both studiesbacterialpopulation reache
apex three to four daysafter inoculation, and no differencecould be fo
between the cultivars. working with p. 'syringae pv, glycinea in soyb
leaves,Mew and Kennedy (l9zl) found a normal pattern of bact
multiplication in three pathogenicisolatesin susceptibleand resis
cultivars.They reportedthat greatnumbersof isolatescould be washedfr
symptomlesssoybeanleaves.
3.3,1 Plant resis,tance
influence on pathogenicpoputation
The degreeof resistanceof the infected plant has an effect on the l
of bacterialpopulation in the plant. Testingp. syringaepv. tabacimultip
tion on susceptible,
resistantand hybridswith variousdegreesofresista
hosts,but not in thoseof the resistantgenotypes.Systemiccolonizatio
plants throughout the growing seasondecreasedwith the increaseof
resistanceof the genotype,to zero in highly resistantcultivars (Cafati
Saettler,1980).
3.3.2 Multiplication of incompatible bacterid'
When incompatiblebacterialpathogenis inoculatedto the correspon
plant tissue,its ability to multiply is usually not affected, but sloweddo
significantly. Comparativestudieson the multiplication of bacteriain b
plants have shown that both compatible (P. syringoepv. phasealicola
incompatible (P. syringaepY. morsprunorum)pathogenscan multiply,
at different rates and to different final populations. The growth of the
compatiblebacteriastoppedone to three daysafter inoculation, wherea
compatiblepathogencontinuedto multiply until five daysafter inoculat
when diseasesymptomsappeared.Bacterial population was alwayssm
in the incompatiblethan in the compatibleinteraction (Ercolani and Cro
1966).
Incompatibility seemsto be due to a reaction of every individual
Studiesof multiplication on tobacco uniform callus tissueshowedthat
compatiblebacterium, P. syringaepv . tabaci, multiplied rapidly and colon
the calluswithin two daysof inoculation, while the incompatibleP. syri
pv. pisi multiplied relatively slowly and remained in the inoculation
(Huang and Van Dyke, 1978).
3.3.3 Saprophytic multiplication
Inoculationof a saprophyticisolatein to a plant tissueusuallyresu
nearly tot;l inhibition of bacterialmultiplication, if tlie plant tissuerem
ing under more or less normal. growth conditions. X. campestn
vesicatoriaand P. fluorescensinoculated onto pepper plants revealed
the saprophytemultipliedon the leaf surfacemorethan the pathogeniciso
but, within the leaf, it wastotally inhibited. Using this phenomenon,Sh
et al. (1982b)developeda highly sensitivemethod for detectingvery s
numbersof bacteriain seedsand symptomlessleaves,by artificially inc
ing the pathogenpopulation insidethe respectivesusceptiblehost and d
media.The me
ting it, whenits numberis high enough,in semiselective
was found to be very preciseand efficient and was adoptedby the see
dustry quality control units (Bashanand Assouline,1983).
Another approach to the problem of saprophyticmultiplication in
tissuewas suggedtedby Young and Paton (1972)who mixed P. syringa
phaseolicolawith saprophytesand showedthat the saprophyticpopul
was stimulatedby the presenceof the pathogenand multiplied to the
extent. In the absenceof the pathogen the initial saprophytic popu
decreasedslowly.
its maximum when severesymptoms of the disorder were observed.All bacteria (40 different isolates)obtained from this lesion failed to induce the disorder in carrot after various
natural methods of inoeulation, indicating secondary muitiplication of
saprophytes(Soroker et al., 1984)
3.3.4 Other factors affecting multiplication
Even in experimentsespeciallydesignedto define the effect of compatibility
on multiplication, factors such as inoculum levels,tissqeconditioning and
others which control the growth of the bacterium itself, had an important
effect. By studying detailedmultiplication of p. syringae pv. phaseolicolo,
P. syringae pv. morsprunorum and a pear strain of p. syringae pv. syringae,
Ercolani and brosse (1966)cameto the conclusionthat host specificity in
the field wasassociatedwith factorscontrd[ing growth of the organismitself
in vivo. They found that in compatible combinationspathogenpopuration
increasedlogarithmically during at least four days following inoculation,
whereas,in incompatiblecombinations,logarithmic growth endedabruptly
after two or three days due to an unknown reaction of the plant's defence
mechanisms.
Growth kineticsstudiesof compatiblep. syringaepv. syringaeand incompatible P. syringaepv. coronafaciensbyDoub and Hagedorn (1980)in susceP
tible and resistantbeansshowedno differencein bacterialgrowth ratesand
final bacterial populations in both types of host when a large dose of inoculum was used.But, for a low doseof inoculum, growth ratesin resistant
plants was much slower. surprisingly, growth rate and bacterialpopulation
of the incompatiblepathogenwere only a little lower than for the compatible pathogen;but, nevertheless,
higherthan expectedin plantswhich responded to the incompatiblepathogenby a typical hypersensitivereaction. Thus,
at least in this case,plant resistanceto the incompatible bacteria does not
see4 due to an abrupt death of the bacteria as was suggestedby Ercolani
and Crosse(1966)but to a slower rate of bacterial growth.
Nearly all the data on this subjectare of a numericalnature and very little
descriptiveinformation can be found. usually multiplication of a compati.
ble pathogenreachesl0p-10?CFLJ/gtissue (orlcmz leaf area); multiplication of an incompatiblepathogenin non-host reacheslOr-ICI cFU/g, and
in saprophytes0-10 without any appaxentmultiplication.
The maximum multiplication level of pathogenicbacteria inside a plant
whether susceptibleor resistantbelongsto x. campesfnispv. malvacearum,
Brown (1980)showedthe closerelationshipbetweenthe amount of pathogen
and the severeness
of diseasesymptoms.The growth pattern of the pathogenic
bacteriain the plant resembledthe typical growth curveof bacteriain broth
culture, in which, at the end of the growth period, the bacterial number reached a maximum of l0.per plant. Essenberget al, (Wg a,b) found even
3.3.5 Conclusion
Exceptfor severalcases,which can be relatedto abnormalor unusua
ditions of the plant tissue,generalconclgsionson the relationshipbet
compatibility and bacterialmultiplication can be drawn. Cgmpatibility
key factor for controlling multiplication in the tissue. It induces
multiplication while incompatibility limits to different extentsthe dev
ment of the bacterialpopulation. Of secondaryimportanceis the degr
plant resistancewhich showsthat, the more resistantthe genotype'the m
limited the pathogenpopulation is. The incompatible pathogenmulti
until stoppedor sloweddown by the plant's defencemechanisms;the c
patiblepathogenmultipliesuntil diseasesymptomsaPPetr,and thenit re
a stationary phase.Finally, saprophytes,under normal conditions of p
gfowth do not multiply in the plant tissue. They can follow a patho
interaction or be encouragedby the presenceof the pathogenin the ti
Environmental factors, inoculum dose and no competition from o
pathogenshave little effect. The inhibition effect is probably at cell or
cluster level.
4. ULTRASTRUCTURE OF PLANT-BACTERIA INTERACTION
Ortcea bacteriumpenetratesinto the intercellularspaces'eitherby its
motility or by artificial infiltration, the plant reactsmainly in two w
(a) Incompatibleresponse:the plant forms variousstructuralbarriersw
the invaders.This usuallyeliminatesor stro
irnmobilizeand encapsulate
reducesthe multiplying ability of the pathogen.The visible effec
reactionin somecellsor regions.This responseis trigg
hypersensitive
only by incompatiblepathogensor saprophytes.
(b) Compatible response:no effe6t is observedin a normal infection
aregra
tern, until severalhoursafter inoculation.Later, cell organelles
v
becomes
and
ly destroyed,and eventualtythe whole tissuecollapses
necrotic.
4,I
RESPONSE
OFTHEINCOMPATIBLE
UITNASTRUCTURE
Ultrastructuralevidencegivessupportto the hypothesisthat immobiliz
or attachmentof incompatiblepathoge9icand saprophyticbacteria,bu
of compatible pathogens,by fibrillaf and granular substancesat the
cell wall, is a specific active defencemechanismof the plant. The pr
is highly localiied and happensas bacterial cells come in contact wit
host cell wall during the first hours after inoculation.
4.1.1 Immobilization or encapsulationhypothesis
The following examplessupport this hypothesis.Goodman et al. (1
b) and Politis and Goodman(1978)showedthat the incompatiblebacte
fragments.This plant-createdenvelopethen immobilizedthe bacterialcells
which migratedto the plant cell wall surface.However,this did not affect
all the bacterialcellsinsidethe tissue,and the saprophyteP. fluorescensinducedthe phenomenononly slightly.Exceptfor a few.unimportantchanges
in plant cell ultrastructure,it seemsthat the plant 'ignores'the injectionof
compatibleP. syringaepv. tabaciinto the tissue,suggestihg
that an 'active'
immobilization occurs in responseto bacterial invasion. Sequeiraand
coworkers(1977)reportedalmostthe samephenomenon(with a slightlydifferent terminology), using incompatible and avirulent strains of P.
solonoceorunon tdbaccotissues.A slightly different observationwasmade
by Singand Schroth(1977)who showedthat fibrillar structuresoriginating
from the plant cell wall of beansimmobilized'actively' the saprophyteP.
putida, but not the compatiblepathogen,P. syringaepv. phaseolicoldor the
incompatible pathogenP. syringae pv; tomoto.
The sequence
of eventswhich can be observedis describedby Goodman
(1978).'Initially, the plant cell wall that is in closeproximity to the bacteria
appearsto ''blister" and the cuticularlayeron the outersurfaceof the plant
cell wall rupjures. This loosenedcuticular layer initiates the bacteriaenvelopingprocess.With time the cuticularlayer becomesthicker with the
integrationof fibrils and small vesicles,and firmly ensheaths
and localizes
the bacteria'Roebucket at. (1978)detailedthesecellularchangesduring incompatibleinteractionof P. syringaepv.phaseolicolain resistantbeanplants:
four hours after inoculationmost bacteriawereattachedto the cell wall which
was partially erodedat the siteof attachment.Aggregationof host cytoplasm
adjacentto the invading bacteriacauseda localizedcytoplasmicinvasion of
the vacuole,accumulationof osmophilicdropletsof unknown naturein the
cytoplasm, vesiculation,degenerationof organellesand finally collapseof
the cell, The adjacentcellsshowedsimilar ultrastructuralchanges,indicating
'transmission'of an unknown factor
betweenthe cells.Casonet al. (1978)
studied the infection of immune ,cotton cotyle{ons with X. campestrispv.
malvacearum.Twenty-four hours after inoculation, the outer cuticle of the
surfacesof host mesophyll cell loosenedand detached,and envelopedthe
adjacent bacteria. The envelopecontainedone or two bacteria and a mass
of fibrillar material. The plasmalemmafrequentlybroke and host cytoplasm
clustered. Thesephenomenawere not observedin inoculated susceptible
plants.
Using scanningelectronmicroscopy,Huang and Van Dyke (19?8)showed quick immobilization of incompatibleP. syringaepv. pisi on the surface
of incompatible tobacco callus by a network of fibrillar material, whereas
the compatiblepathogenP. syringaepv. tabocimultiplied freely without interaction with the plant cells. P. fluorescenscells formed sphericalbodies
apparentlyas a result of damageto the bacterial cell wall, which eventually
4.1.2 Evidenceagainst the immobilization hypothesis
phenomenonis
some contradictoryinformation on the immobilization
of a .surformation
the
givenby Sigeeand Epton (1975,1976)who showed
face membrane'over bacierialcells of both compatibleand incompatible
bean'
strainsof P. syringaepv. phaseolicolain the intercellularspaces'of
inand
saprophytic
of
However,all other stuiiesieported only attachment
imthe
on
Doubts
wall.
co-putiut" strainsof the pathogento the beancell
bacteriawithin the leaf and
portanceof the fibrillar materialencapsulating
podtissuewereformulatedbyDoubandHagedorn(1980),whod
after inbir.on.. strongevidenceof an immobilizationdefencemechanism
pods,
and
in
spaces
oculation. Bacteriamultiplied and filled the intercellular
could
and
rarely
encapsulationof bacteriain leaf tissuewas observedonly
or incombe found unspecificallyin the.interactionsof either compatible
pathogens'
patible
'
P' syrThe ultrastructur'alstudy of Fett and Jones(1982)working with
They
theory.
ingaepv. gtycineain soybeanalso resiststhe immobilization
'electron-dense
founOttraiUacterialcellswereoccasionallyenvelopedby an
activeor
material' of uncertainorigin. They could not prove that it was an
'"specificreactionof the host, sincesimilar cell structureswere found crossthere
Ltiaging adjacentleaf mesophyllcelisat cell junctions. Furthermore,
wasneitherevidenceof immobilizedbacteriain incompatiblecombinations
of other strains of P. syringde pv. glycinea, even if a typical hypersensitive
accumulareactionwasinduced,itbr aisruptim of the host plasmalemmaand
which were
tion of vesiclesat the site of bacterial encapsulation'events
describedby Politis and Goodman (1978)'
found
Resultscompletelycontradictoryto the immobilization theory were
byMazztlchietat.(1982),whoinducedprotectionagainstP.syringa
complexes
t ibaciinfectionin tobaccoleavesby proteinJipopolysaccharide
susceptible
in
both
encapsulated
was
case
pathogen
this
in
ih. .o,npurible
in the inmaterial
fibrillar
of
network
a
by
tibsue
unA in piotectedtobacco
to be a critical positivephase
tercelluiarspaces.This encapsulationis suggested
which enablesthe bacterii to survive until active growth is established'
the encapsulaHildebrand et al. (1980)take the strongeststand against
'active'.or passive
between
no
connection
is
tion theory, claimingthat there
fibrillar
immobilization of bacteriain the plant cell wall and resistance,asthis
that
suggest
plant.
They
the
in
phenomenon
physical
material is a result of a
possibly
an
is
micrographs
electron
on
the immobilization theory basid
inthe
into
infiltration
Water
interpretation.
artefact or at leastan incoirect
wall surtercellular spacesof bean leavesdissolvematerials from the cell
films
forming
water,
the
of
planf
transpiration causesevaporation
faces.
Comintelfaces'
air-water
at
material
consistingof amoiphous and fibrillar
patible (P. syrinsae pv. syringae),incompatible(P' syringaepv ' marginalis'
(P'
P. syringae pr. pii and P. syringae pv' tomato) and saprophyte
4.1.3 Adsorption of bacteria to plant suffices
Attachment of non-compatiblepathogensand saprophytesdid not alwa
occur, but adsorptionof bacteriato plant surfacesor the intercrllular spac
of plant tissuedid. Evidencethat the adsorptionin the intercellularspac
of tobacco leaf tissue of saprophytic bacterium p. fluoresceizsis strong
than for the pathogenicbacteriaP.syringaepv.plsl and p. syringaepv. taba
was shownby Atkinson et al. (1981).The adsorptionwasfast and two hou
after inoculation bacterialrecoveryfrom the tissuewas inefficient. Im
rnob_ilizationof P. syringae pv. pisi cells by tobacco cell walls, as reporte
by Goodmanet al. (1976a)was not.observedand a very small amount o
fibrillar material was found.
The numbersof the pathogenP. syringaepv. tolaasii, attachedto the su
face of the cultivated mushroom Agaricus bisporus was twice the nurnb
of its common saprophytesP. reactansand of p. putida, which inducesth
sporophoreformation in the mushroom.Firm attachmentof bacteriao
curred when either pathogenicor saprophyticbacteriawereappliedto th
hyphae of A. bispora,s(Preeceand Wong, 1982).
4.1.4 Influence of the inoculum concentration
The influenceof the inoculum concentrationon the ultrastructuralchang
was clearlydemonstratedin the caseof cotton infection by x. campes
pv. malvacearum.cason et ol. (1978)inoculatedimmunecotton plant wit
a high concentration'ofinoculum,which resultedin the total encapsulat
of bacteriaby fibrillar material connectedto the mesophyllcell walls.
Relativelylow inoculum levelsof this pathogenusedby Al-Mousawi
al. (1982b)showedthat bacteriawerefound in the intercellularspacesne
collapqedmesophyllcellsbut, althoughfibrillar materialwaspresent,it di
not completelyenvelopthe bacteria.This looseness
of the fibrillar materi
could not direcflyrestrictbacterialmultiplicationassuggested
by Goodma
et al. (1976a). Nevertheless,
this evidencewasnot sufficientto decidewheth
the attachmentobservedplays a role in the cotton resistance.
A drasticexampleof ultrastructuralchangesrelatedto inoculumconce
tration of P. syringaepv.phaseolicolawascarriedout by Sigeeand Epto
(1975,1976)inresistantand susceptible
beanplants.Markeddifferences
we
observedin the pathogen.Surfaceprotuberances
appeared,and later disa
peared.In resistantplants, the bacterial'nuclearregion' broke down an
ribosomalaggregationdeveloped,whereasin the susceptible
piant no suc
changesoccurred.Later, during diseasedevelopment,somebacteriain th
susceptibleplants producedlarge surfacevesiclesnot found in any othe
phytopathogenicbacteria.some vesiclesrupturedliberatingtheir contentint
the intercellularspace.Major changesalsooccurredin the plants.Howeve
which definitely is not typical of this disease.
4.L5 Problems in interpretation of ultrastructural evidence
Many unclear and contradictory eventsmake the interpretation of t
ultrastructural evidenceof immobilization of bacteriain planttissue and t
evaluation of its validity, importance or contribution to the plant defen
mechanismsa very problematictheory. Data givenin thebacterialmultiplic
tion sectionseemto rule out this theory. Hildebrand et al. (1980)wonder
if bacterialmultiplication is possiblebecausesomeor all bacteriaescapee
capsulation,or becausetheyare'strong' enoughto break out ofthe relativ
fragile binding structure.
Furthermore, both resistant and susceptibleplants were shown to im
mobilizeboth compatibleand incompatiblebacteria.Additiqnally, mainta
ing the leaveswater soakedtotally preventedthe attachment,suggestingth
if attachmentwas an activgprocessit could not be preventEdby the prese
of only water (Sigeeand Epton, 1976;Stall and Cook, 1979:'Mazntchi
al., 1982).
Additionally, somestructuresare formed in the tissue,not only in respo
to bacterial inoculation, but as a consequenceof water evaporation fro
plant tissue even in the absenceof bacteria, and act as non-specifictra
for bacteria.All this evidencetendsto prove that encapsulationalonecou
not be a generaldefencemechanism.
4.1.6 Conclusion
Bacterial attachmentto plant cellsby ultrastructural studiesis so uncle
that interpretation is subjective and highly speculative,and can be on
qualitative.The micrographsare subjectedto possibleartefactsand depe
on plant tissue conditions, inoculation method and the quality of t
micrographs.
4.2 UlrnnsrnucruRE oF THECorapettsI-E RESPoNSE
Little information is availableon compatibleinteractions.Contrary to
compatibleinteraction,most of thesestudiesagreewith eachother and d
fer only in detailsabout the diseases.
Attack of susceptiblecotton plants by X. compestrispv. molvacear
showedpathogenmultiplicationresultingin severedamageto the membra
of all organellesand to the wall structureof mesophyllcells.Early chan
includedformation of vesiclesbetweenthe plasmalemmaandceil walls,dis
pearanceof the granaland stromalmembranesof the chloroplasts,follo
to unde
of mitochondria.The last cellularorganelles
ed by degeneration
structural degenerationwerethe cell nucleusand the plasmalemma.Fibril
material was found at the externalcell surfaces,near to bacteria, 120hou
after inoculation.The pathogeniccellsthen enteredthe mesophyllcells,a
materialof the rentainingcell wall dissolvedthroughthe cell.The ultrastru
tural changesof the hostmembraneresembled
changesoccurringduringle
(Butlerand Simon,l97l); theymay, therefore,be dueto autoly
senescence
of the host in responseto pathogen(Al-Mousawi et al., 1982a;Cason
at.,1977).
4.2.1 Effect of inoculum concentration
The effect of inoculum concentrationson the ultrastructurewas show
by Goodmanand Burkowicz(1970)who infiltrated high concentrationso
virulent and avirulent E. amylovora into apple leavestissue.Twelve hou
later, spreadingof grana of the chloroplast,degradationof mitochond
and microbodiesand aggregationof stromain chloroplastsand cytoplas
were observed.Later most of the cellularorganelleshad lost their norm
structure.The interestingfact is that thesechangesoccurredboth in virule
and avirulentstrainsand wereprobablyrelatedto the high bacterialconce
plantsinste
reactionin susceptible
tration, and produceda hypersensitive
of a susceptiblereaction.
However, when relatively smaller numbers (iS CFU/cm2 leaf) of E
amylovora and P. syringaepv. pisi were infiltrated into tobaccotissue
widespre
moderateeffecton cellconstituentswasobserved.Nevertheless,
plasmalemma,
an
bounding
tonoplast,
damagewascausedwithin the cells:
mem
internal membranesof chloroplastsand mitochondria,the external
branesof microbodies,cytoplasm,and free ribsosomeswereall highly a
of bacter
fectedand/or destroyed.All theseeffectswerea consequence
(Goodman
and Plura
reaction
actionsresultingfinally in hypersensitive
l97l).
4.2.2 Ultrastructural changesdue to wilting bacterial diseases
Detailedhistopathologyof tomato plantsinfectedwith P. solanqceor
by Wallisand Truter (1978)showedthat someof the invadingbacteriamigra
into the tyloseswhich bulgeand later rupture, Iiberatingthe pathogensan
non-cellularmaterialsinto the plant vessels.Bacterialmultiplication th
rapidly producedlarge amountsof bacterialEPS. Later, a thick layer o
materialwasdepositedon the primary walls of many inva
electron-dense
Host degradationproductswereapparentlymin
ed rootsand stemvessels.
importance
in diseas.e
development.
and
in amount
disease,
caused by Corynebacter
vascular
system
In another
degrad
(1977)
the vessels,
disrupting
Wallis
detected
bacteria
michiganense,
collaps
and
plant
separate
to
cells
the
the middle lamellas,and causing
pathog
pv.
the
campestris,
by
X.
campestris
In cabbageleaves,infected
degradedthe primary wall. After completedissolutionpf the wall, the bacte
wereableto passin adjacentvessels.Shreddingof the primary wall resul
in swellingof wall residuesand releasedmassesof fibrillar material,whi
sepedonicumshowedthat cell organelles
and all the membranesystemswere
affected.Excessive
amountsof abnormalmembraneswerepresentin some
cells.Severityof the darnagewasrelatedto susceptibility.Organelledisruption was not correlatedto the presenceof bacteria,and cells far from the
site of bacteriain the intercellularspaceswerehighly affected.Thi'smay be
due to the action of a diffusible toxin (Hessand Strobel, 1970).
4.2.3 Conclusion
In compatibleinteractions,the destructionof plant cells,resultingin the
appearance
of visiblesymptomsat the end of the diseasesyndrome,could
previouslybe detectedat the ultrastructurallevel.The cellorganelleand particularly the membraneswere the first to be destroyed,indicating the
biologicaldeathof.the cells.
'Browning'of the deadtissuecan be relatedto othermechanisms,
such
asphenolaccumulationoccurringin the tissue.The relativelysmalldifferences
in the plant tissue,at the ultrastructurallevel,do not meanthat the disease
mechanismsinvolved in induction of theseeventsare similar.
5. CONCLUSION
Currentknowledgeon plant-bacteria
interactiondoesnot clearlydemonstrate
a generalspecifichost-pathogeninteractionat the cell level.Modelssuggested
do not explaingeneralrecognitionin plant pathologywell enough,but only
thesephenomenaoccurringbetweencertainspeciesof bacteriaand plants,
which unfortunatelycannot be generalizedto other pathogenicsystems.
One fact generallyacceptedis that surfacemoleculesof both bacteriaand
plant are involvedin determiningspecificrecognition.But somemajor questions remain:
(a) How canplant cell surfacesdeterminerecognitionand non-recognition
with the samemoleculararchitectureon its surface?
(b) Are the O-chainsof LPS, which is the most variablepart of the LPS,
responsiblefor pathogenicspecificity?
(c) Do the different pathovarsof P. syringaecontainuniquecarbohydrates
on their surfaces?
One possibleexplanationis that plantsmight havetwo differentreceptors
for bacterialcell wall carbohydrates.One of them-perhaps the core LPS
or certainEPSs-would be non-specificin order to respondto the general
characteristics
of incompatiblebacterialsurfacemolecules.These-bacteria.
kind of
receptorsallow the plant to recognizea largenumberof incompatible
Compatiblepathogenswould haveto developalteredstructuralsurfaceto
achievenon-recognitionby the plant generalreceptors,or to maskthe plant
recognitionsystemby producingbig amountsof EPSs.Pathogenswhich
escapedetectionby the recognitionsystemcan theoreticallyinducedisease
in their hostplant. The secondtype of receptormight be more specific.The
are in fact theseproposedreceptors.Despitethe amount of data, eviden
mediatespecificrecognitionby signaltrans
that surfacemacromolecules
is asyet quite limite
interfacesin bacterialplant disease,
at host-pathogen
incornpatiblerea
the
that
Howevei. most studiesacceptthe assumption
recognition.
specific
to
lead
tion, and not the compatiblereaction,
'a
In a recentreview,Daly (1984),claimedthat thereis still needto sea
involvinguniquemolecu
for an initial event(recognition)in pathogenesis,
species,that furnishesa signal(communication)for a biochemicalrespo
that in itselfmay be quitegeneral'.Criticalevaluationof all proposedmod
for plant bacteriarecognitiondo in fact needmore chemicaldata on t
bacteria.Intens
ofmany strainsof phytopathogenic
surfice carbohydrates
knowledgein molecularbiologyin termsof the differentrecognitionsyste
of specificity
wouldindicatewhetherthereis any uniquedetermination
interactio
whetherspecificityoccursthroughmanydifferenthost-pathogen
ACKNOWLEDGEMENTS
to the memoryof lateMr. Avner Bashanwho h
This reviewis dedicated
during
of inspirationand encouragement
source
immense
an
been
preparation.
This review was partially supportedby a Sir CharlesClore fellowsh
I wishto thankMrs VeroniqueLevyfor carefuleditingof thismanusc
during its preparation.Y. Bashanis incumbentof the william T. Hog
and Winifred T. Hogan Careerf)evelopmentChair.
REFERENCES
pathog
ALLINGTON, W.B. and D.W. CHAMBERLAIN. Trendsin the populationof
Phytopatholog
bacteriawithinleaftissuesofsusceptibleandimmuneplantspecies.
t
656-660,1949.
AL-MOUSAWI,A.H.,P'E.RICHARDSON,M'ESSENBERGandW'M'JO
Xanthomonascampestr
Ultrastructuralstudiesof a compatibleinteractionbetweerr
m alvacesrwn and cotton. Phytopt hology, 12, 1222-1230,1982a'
AL-MOUSAWI,A'H',P.E.RICHARDSON,M.ESSENBERGandW.M.Jo
cottonline inoculatedw
of a bacterialblight-immune
Cotyledonand leaf ultrastructure
Phytopathology,l2,1230-1
pv.
malvaceorum.
campestris
a low levelof Xanthomonas
l 982b.
ANDERSON,A.J. and C. JASALAVICH. Agglutinationof Pseudomonadcells bY P
products.Physiol.Plant Patho!., 15,149-159,1979'
ATKINSON.M.M., J.S. HUANG and C.G. VAN DYKE' Adsorptionof Pseudomonad
tobaccocell wallsand its significanceto bacterium-hostinteractions' 'Physiol. Plant Path
lE, l-5,1981.
of Erw
AYERS. A.R., S.B. AYERS and R.N. COODMAN. Extracellularpolysaccharide
1
659-666'
Microbiol.,38'
Environ.
Appl.
amylovoru:a correlationwith virulence.
Che
BAKER. C.J., M.J. NEILSON, L' SEQUEIRA and K'C' KEEGSTRA'
BASHAN, Y. and I. ASSOULINE. Complementarybacterialenrichmenttechniquesfor
detectionof Pseudomonassyringae pv. tornato andXanthomonascampestrispv. vesicat
in infested tomato and pepper seeds.phytoparasitica, ll, lg7-193, 19g3.
BASHAN, Y.' Y. oKoN and Y. HENIS. Infectionstudiesof pseudomonastomato, ca
agent of bacterial speckof tomato. phytoparasitica, 6, l3S-144, lg,lg:
BASHAN, Y., c. KRITZMAN, E. SHARON, y. OKON, and y. HENIS. A note on
detectionof phytopathogenic
bacteriawithin the leaf by a differentialstainingproced
J. Appl. Bacteriol.,50, 315-317,l98la.
BASHAN, Y., E. sHARoN, Y. oKoN and y. HENIS, scanningelecrronand light microsc
of infectionand symptomdevelopment
in tomato leavesinfxtedwith Pseudomonas
tomo
Physiol.Plant Pathol.,19, 139-144,
l98tb.
BASHAN' Y., S. DIAB and Y. OKON. Survival of Xanthomonascampestrispv. vesicot
in pepperseeds,in symptomless
and dry leavesand in soil. ptant soit,6E, 16l-l':.0,lgg
BASHAN, Y., Y. OKON and Y. HENIS. Detectionof a necrosis-inducing
factor of nonh
plant leavespro ducedby Pseudomo nassyringaepv . tomato. Can. J. Bot., 60, 24Si-24
1982b.
BASHAN, Y.,M. AZAIZEH, S. DIAB, H. yUNISand y. OKON. Croptossofpepperpl
artificially infected with xanthomonas campestrblv. vesicatoriain relation tq sympt
expression.Crop Protec.,4,77-84, l9g5a.
BASHAN' Y. and Y. oKoN. Internal infectionsof fruits and seeds.ofpeppercaused
Xanthomonas campestrb pv. vesicatoria.Can. J. Bot. 64,2865-28'll, 1986a.
BASHAN, Y. and Y. oKoN. Diseased
leaf lyophilization:a methodfor long-termpreven
of lossof virulencein phytopathogenic
bacteria.J. Appt. Bacteriol.61,163-16g,l9g
BASHAN' Y.' Y. OKON and Y. HENIS. Morphologyof leaf surfacesof tomato cultivar
relation to possibleinvasioninto the leaf by pseudomonassyringaepv. tomato.Ann, Bo
5 5 , 8 0 3 - 8 0 9t ,9 8 5 b .
BAUER, W.D. Infectionof legumes
by Rhizobia.Annu. Rev.plant physiol.,32,40't484,lg
BAUER, W'D. The initiation of infectionsin legumesby Rhizobia.ln Plant Infection. T
Fhysiologicaland BiochemicalBasr, editedby y. Asada,w,R. Bushnell,s, ouchi and c.
Vance,pp. 6l-78, JapaneseScienceSocietyPress,Tokyo, 1982.
BENNETT, R.A. and E. BILLING. Origin of the polysaccharide
componentof oozefrom pla
infectedwith Erwinia amylovora.J. Gen. Microbiol., 116,34l-349,19g0.
BERRIDCE, E.M. Studiesin bacteriosis.XVL The agglutinatingand plasmolyticacrion
the sapof the potato on variousparasiticand saphrophyticspeciesof bacteria.Ann. App
Biol., 16, 567-577,1929.
BHUVANESWARI, T. and w.D. BAUER. The role of lectins in plant-microorgan
interactions.III: Influenceof rhizosphere/rhizoplan
cultureconditionson the soybe
lectin-bindingpropertiesof rhizobia.ptant physiol,, 62,j1-74, lg:il,
BoHLooL, B.B. and E.L. SCHMIDT. kctins: a possiblebasis for specificityin t
Rhizobium-legumeroot nodulesymbiosis.Science,lES, 269-2'll, 1974.
BOHLOOL, B.B. and E.L. SCHMIDT. Immunofluorescent
polar tipsof Rhizobiumjaponicu
possiblesite of attachmentor lectin binding. J. Bacteriol.,125, I lgg-l 194, 1976.
BROWN, S.J. The relationshipbetweendiseasesymptomsand parasitegrowth in bacterialblig
of cotton.J. Agric. Sci.Camb.,94,305-312,1980.
BUTLER, R.D. and E.W. SIMON. Ultrastructuralaspectsof senescence
in plants.Adv. Gero
t ol. Res.,3, 7 3-129,197l.
CADMUS' M'c.' K.A. BURTON and M.E. sLoDKI. Growth-related
substituentchange
exopolysaccharidds
of fast-growingrhizobia.Appl. Environ. Miuobiot., 44,u2-24s,lg12
CAFATI, C.R. and A.W. SAETTLER. Effectsof host on multiplicationand distributiono
bean.commonblight bacteria. P hytopot hology, 70, 6j 5 -679, l9BO.
CASON,E:T., Jr., P.E. RICHARDSON,M.K. ESSENBERG,
L.A. BRINKERHOFF.W.M
malvacearum.Phytopathology,61, 195-198,1977.
lachryman
of Pseudomonas
and Y. HENIS. Chemotaxis
CHET, I., Y. ZILBERS',TEIN,
and susc
of resistant
plantextractsandto walerdropletscollectedfrom the leafsurfaces
tible plants.Physiol.Plant Pathol.,3,473-479,1973.
CLARKE, A.E. and R.B. KNOX. Cell recognitionin floweringplants.QuarL Rev.Biol.,5
3-28,1978.
on responseloXanthomonasvesicat
COOK, A.A. and R.E. STALL. Effectsof water-soaking
in pepperleaves.Phytopathology,67, I 101-l103, 1971.
of a methodfor measu
lV. Investigation
CROSSE.J.E. Bacterialcankerof stone-fruits.
the inoculumpotentialof cherry :rees.Ann' Appl. Biol., 4'1,306-317' 1959'
cRosTHwAITE, L.M. and S.S. PATIL. lsolation of an endotoxin from Pseudomo
phaseolicolawhich inducescellcollapsein bell pepper.Phytopathol.2,91,80-90, 19
Annu. Rev.Phytopathol.,22,273
in plantdisease.
DALY, J.M. The roleof recognition
1984.
DAZZO, F.B. and W.J. BRILL. Receptorsiteon cloverandalfalfarootsfor Rhizobium'App
Environ. Microbiol., 33, 132-136,1977.
which bindsRhizobiumtrifolii
DAZZO, F.B. and W.J. BRILL. Bacterialpolysaccharide
1919.
clover root hairs. ./. Bacteriol.,131,1362-1373,
receptor
DAZZO, F.B. and R.E. HOLLINCSWORTH.Trifoliin A and carbohydrate
mediators of cellular recognitionin Rhizobium trifolii-clover symbiosis.Biol. Cell, 5
267-274.1984.
and lectinsas determinan
antigens
DAZZA, F.B. and D.H. HUBBELL. Cross-reactive
Miuobiol., 30' l0l7-10
Appl.
association.
in
Rhizobiunr-clover
the
specificity
symbiotic
I975.
to rootsasrela
DAZZO, F.8., C.A. NAPOLI andD.H. HUBBELL.Adsorptionof bacteria
to host specificityin the Rhizobium-cloversymbiosis,Appl. Environ. Microbiol., 3
t66-171, 1976.
recepto
of lectinsand theirsaccharides
DAZZO, F.B. and C.L. TRUCHET.lnteractions
1983'
13,
1-16'
Biol.,
Membr.
J.
symbiosis'
Rhizobium-legume
the
DAZZO, F.8., G.L. TRUCHET, J.E. SHERWOOD,E.M. HRABAK' M' ABE and S'
PANKRATZ. Specificphasesof root hair attachmentin theRhizobiumtrdolii-cloversy
1984.
biosis.Appl. Environ.Microbiol.,4E'1140-1150'
DAZZO, F.B., W. YANKE and W.J. BRILL. Trifoliin: ARhiZobiumrecognitionproteinfro
white clover.Biochint.Biophys.Acta,539,276-286' 1978.
DIAB, S., Y. BASHAN and Y. oKoN. studiesof infection with Xanthomonascampe
pv. vesicatoria,causalagentof bacterialscabof pepperin lsrael.Phytoparasitica
183-19r,1982.
tabaciinleavesof bur
DIACHUN, S. and J. TROUTMAN. Multiplicationof Pseudomonas
186-187
44,
' 1954'
hybrids.
Phytopathology,
and
longdlora,
tobacco, Nicotiana
DOMBRINK-KURTZMAN,M.A.,W.E.DICK,JT,K.A'BURTON,M'C'CADMUS
acid specificity.Bioch
SLODKI. A soybeanlectin having 4-O-methyl-D-glucuronic
Biophys.Res.Commun, 111' 798-803'1983.
DOUB, M.E. and D.J. HACEDORN. Growth kineticsand interactionsof Pseudomonos
ingae, with susceptibleand resistanttissues.Plylopathology, 70,429-436,1980'
of Rhizobiumandt'hetrole in
andoligosaccharides
DUDMAN, W.F. The polysaccharides
infection process.ln Advancesin Nitrogen Fixation Research,edited by c. Veegersa
MaritinusNijhoff/Junk, The Hague,Netherlands'1984
W. Newton, pp. 39'1-404,
of newlyfound substitu
DUDMAN, W.F. andW.F. HEIDELBERCER.Immunochemistry
1969'
954-955,
164'
Science,
species.
of Rhizobiurr
of polysaccharides
lipopoly
solanacearum
DUVICK, J.P. and L. SEQUEIRA. Interactionof Pseudomonas
Appl. E
potato
tubets.
from
polysaccharide
with
agglutinin
charide and extracellular
viron. Miuobiol., 4E, 192-198,1984a'
in beanleaves(phoseolusvu
L.). phytopothol. 2., 95, 38-50,1979a.
EL-BANOB', F.E. and K' RUDOL'H.
Inductionof warer-soaking
in plant leavesb
tracellularpolysaccharides
from phytopathogenicpseudomonads
and xanthom
Physiol. plant pathol., 15,341_34g,l*SA.
EL-BANOBY' F'E' and K' RUDoLPH. Purification
of extracellularpolysaccharid
Pseudomonas
phaseoricora
which inducewater-soakingin bean reaves.
physior.
Pathol., 16, 425_437,1980.
EL-BAN'BY, F'E', K. RUD.L'H andA.
HUTTERMANN.Biological
andphysicalpr
ties of an extracellularpolysaccharidefrom pseudomonas
phaseoricota.physiot.
pathol., 17, 29lg0l,
1980.
EL.BANOBY, F.E., K. RUDOLPH ANd
K. MENDCEN. ThC fAtCOf CXITACEIIUIA
PO
charidefrom Pseudomonas
phaseolicoloin leavesand leaf extractsfrom
halo-blightsu
resisranrbeanplants(phaseorus
vurgarisL.). phvsior.plant pathot.,18, 9
l;1;.""0
ENCLISH' H' and J 'R' DAVIS' The source
of inoculumfor bacterialcankerand blastof
trees.phytopathology,S0,634(abstr.), 1960.
__ _fruit
ERCOLANI, G.L. and J.E. cRossE. rne gro*iir
of pseudomonas
phaseoricotaand
re
plant pathogensin vivo. J. Gen. Microbiot.,
45,42g_43g,1966.
ERCOLANI, C,L., D.J' HAGEDORN, A. KELMAN
ANdR,E. RAND. EPiPhYtiC
SUTV
Pseudomonas
syringaeon hairy vetch in relation 1o epidemiology
of bacteriatbrown
of bean in Wisconsin.phytopatftology,64,
l33}_133g,1974.
ESSENBERC,
M., E.r. cs^gf,,J.,e. HAMir_rbN, L.n.
sruNxpRHoFF, R.K. GHOL
and P'E' RI.HARDS'N. Singlecell colonies
of xanthomonasmarvacearuminsu
tibleand immunecottonleavesand the local
resistance
response
to coloniesin imm
teaves.physiol. plant pathol., 15, 53_6g,
1979a.
ESSENBERG,
M., B. HAM_ILTON,E.T. C^SON,fr,-r_.e.
BRINKERHOFF,R.K. .HOLS
and P'E' RICHARDSoN. Localizedbacteriostasis
indicateduv *ur.. jlrp.rsal of
oniesof Xanthomonasmarvacearuntwithin immune
physior. prant path
cotton reaves.
t5,69_78,1979b.
FETT' w'F' and s'B' JoNES' Roleof
bacterialimmobilizationin race-specific
resrstan
soybeanro pseudomonassyringae pv. glycinea. phytopathotogy,
i2,
4gg_4g2,
lgg
FLETCHER, M., M'J. LATHAM, r.'{.-r-vr.r-cn
una p.n. RUTTER. The characrerist
interfacesand their role in microbialattachment.
rn Microbial Attachmentto surfo
editedby R.C.W.Berkeley,
J.H. Lynch,J. Melling,p.R. RutterandB. Vincent,pp.6
Ellis Horwood, Chichester,19g0.
GADE, W., M.A. JACK, J.B. DAHL, E.L,
SCHMIDT ANdF. WOLD. ThEiSOIAtiO
characrerization
of u
from soybean(Gtycinemax. (L.),cultivar Chippe
:::t^].:tin
J. Biol. Chem.,256,12905_12910,
1981.
GARDNER, J'M. and c.I. KADo. rnduction
of the hypersensitive
reactionin tobaccow
specifichigh-morecular
weightsubstances
derivedfrom the osmoticshock fruid of.
winia ru brifaciens. phy topt hoIogy, 62, 759 (Abst.),
I 972.
GATEHOUSE, J'A. and D. BouLTiR. tsotatron
ano propertiesof a lectin from the ro
of Pisum sativum (garden pea).physiol. plant.,
49, 43i-442, lgg0.
^__
.HANEKAR, A' and M.c.M. 'ER.MBEL.N.
Interactionsbetweenpotarorecrinand so
phytobacteriain relationto potafo tuber
decaycausedby Erwinia cirotovora. phytopath
2., 9E, 137_149,
1980.
GITAITIS, R'D', D.A. 'AMUELS.N and
J.o. 'T'RANBERC. Scanningelectronmrcrosc
of the ingressand establishmentof pseudomonas
arboprecipilars in sweetcorn lqav
Phytopathology,71, 17l-17S, lg8l.
cooDMAN, R.N. Inducible resistanceresponses
in prants to plant pathogenicbacte
Mycopathologia, 65, t\j -t 13. 1979.
hypersensitiver.".t,on.uu,.dbyplantpathogenicbacteria.Physiol.PtantPatho
1971.
GOoDMAN,R.N.,J.S.HUANGandP,-Y.HUANC.Host-specificphytotoxicpolysacc
1E3' l08l-1082' 1974'
from appletissueinfectedby Erwinia amylovora'Science'
for immobiliza
evidence
Ultrasrructural
cooDMAN, R.N., p.-y. HUANe and J.A. WHITE.
leaf tissu
tobacco
prsr,
in
Pseudomonas
tion of an incompatible bacterium,
PhYto7atholog!, 66, 7 54-764,1976a'
GoODMAN,R'N',D.J'POLITISandJ'A.wHlTE.Ultraslructuralevidenceofan.a
immobilizationprocessofincompatiblebacteriaintobaccoleaftissue,ares
tion.lnCelt||atlBiochemistryRelatedtoSpecifici'tyinHostPlant-Pathogenlnt
/ions,editedbyB.SolheimundJ'Ruu'pp'423-438'ColumbiaUniversity
York, 1976b.
asinduce
Bacteriallipopolysaccharides
cRAHAM, T.L., L. SEQUEIRAand T.R. HUANG.
197'7'
34'424-432'
Microbiot"
Environ'
ih tobacco'Appl'
of diseaseresistance
HENDRICK,C'A.andL.SEQUEIRA.Lipopolysaccharide.defectivemutantsof
pathogenPseudomonas'ol-ono"o'u*'App!'Environ'Microbiol"4E'94-l0l'1984
gpiphytic survivalof bacterialleaf pathogens.ln Microbiolog
HENIS, y. and y. sasF;N.
van den Hevrel' pp' 252-268' Can
oJ thephyttosphere,editedby N'J' Fokkemaand J'
1986'
biidge'UniversityPress,Cambridge,
HESS,w.M.andc.A.srnonnr-.UltrastructureofpotatostemsinfectedwithCorynebacte
1' I 970'
epedonicum'Phytopathology,60, 1428-143
si;
SCHROTH. Phvsicalentrapmento
M.N.
and
ALOSI,
inl.C.
D.C.,
HILDEBRAT{D,
air-water interfaces.Phltopatholog
at
formed
films
pseudomonadsin ue.anieau., by
1
9
8
0
.
70,98-109,
HlRANo,s.S.andC.D.UPPER.Ecologyandepidemiologyoffoliarbacterialplantp
Annu. Rev.Phytopathot',21,243-269'1983'
of bacterialicenucleion snapbe
Frequency
HIRANO, S.S.,D.I. ROUSEand c.D. uppER.
prediStorof b".t.ti"l brown spot' Phytopatholog
(phaseolusrrtsor*";.li;ufl*r1r
"
72, 1006(Abstr.)' 1982'
and K' HALLENCA' ldentificatio
HOLLINCSWOTH, R'I., M. ABE, F'B'DAZZO
3.hydroxybutano,.u.ioasacomponentoftheacidicextracellularpolysac
c7'cll' 1984'
Rhizobium trifolii 0403'Carbohydr'Res'' 134'
HoRlNo,o.Inductionofbacterialleafblightresistancebyincompatiblestrains.ofXonlf
Piant-ParasiteInteraction, edited by
oryzaein rice.ln Biochemistryand cytotogy of
S' Ouchi' pp' 43-55'ElsevierScient
and
Tomiyama,J.M. Daly, l' Uritani, H' Okv
1976'
Pub., New York,
HRABACK,E.M.,M.R'URBANOandF'B'DAZZO'Growth-phasedependentimmu
which bind trifoliin A, a white clo
minantsof nnizotiiun rry'olii tipopotvsaccharide
lectin. ,/. Bacteriol.,l4E, 697-7ll' 1981'
wilt-inducingtoxin in ap
productionof a host-specific,
HSU, S.T. and R.N. CoOoMeN.
withErwiniaamylovora.Phytopathology,6t'351
cellsuspensionculturesinoculated
1978a.
culture
in applecell suspe^nsion
Hsu, s.T. and R.N. GooDMAN. AgglutinatinSactivity
19
Phytopathology,6E,355-360,
amylovora.
oculatedwith aniruf.ni rtran of;lrwinta
HUANC,J'S.andc.G.VANDYKE.InteractionoftobaccocallustissuewithPseudo
Phlsiot' Plant Pathol'' 13' 65-'12'1918'
tabaci,P. pisi andP.ifuo'""'^'
modificationsin applestemsindu
p.-y
Ultrastructural
GOODMAN.
R.N.
and
HUANC,
66' 269-216'19'1
Phytopathology'
toxin'
by Erwi4ia amylovoraand the fireblight
of applesh
mechanisms
Resistance
HUANG, P.-Y., J.S. HUANG and R.N. GOODMAN.
toanavirulentstrainofErwiniaamylovora.Physiol.PtantPathot',6,283-28.
inthe Rhizobium-peasymbiosis'FE
KAMBERGER, w. RoIeof cell surfacepolysaccharides
Microbiot. Lett., 6, 361-365,19'19'
Cell Physiol., 22,759-771,198L.
KEEN, N.T. and M.J. HOLLIDAY. Recognitionof bacterial pathogensby plants. In
Phytopathogenic
Prokaryotes,editedby M.S. Mount andC.H. Lacy.Vol. 2, pp. 179-21
AcademicPress,New York, 1982.
KEEN, N.T. and P.H. WILLIAMS. Chemicaland biologicalpropertiesof a lipopucopolysac
charide from Pseudomonaslachrymans.Physiol. Plant Pathol., 1,24'1-264, 1971.
LEACH, J.E., M.A. CANTRELL and L. SEQUEIRA.Hydroxyproline-rich
bacterialagglutini
from potato;extraction,purificationand characterization.
Plant Physiol.,10,1353-135
1982a.
LEACH, J.E., M.A. CANTRELL and L. SEQUEIRA. A hydroxyproline-rich
bacterialagglutinin from potato: its localizationby immunofluorescence.
Physiol.Plant Pathol.,2l,
319-325,1982b.
LEBEN, C. Multiplication of Xonthomonas vesicatorioon tomato seedlings.Phytopathology
5 3 , 7 7 8 - 7 8 11,9 6 3 .
LEBEN, C. Epiphyticmicroorganisms
in relationto plant disease.
Annu. Rev. Phytopathol.
3,209-230, 1965.
LEBEN, C. and R.E. WHITMOYER. Adherenceof bacteriato leaves.Can. J, Microbiol,,
2 5 . 8 9 5 - 9 0 11,9 7 9 .
LEBEN, C., G.C. DAFT andA.F. SCHMITTHENNER.Bacterialblightof soybeans:
popula
glycineain relationto symptomdevelopment.Phytopstholog:t
tion levelsof Pseudomonas
5 E , l l 4 3 - 1 1 4 61, 9 6 8 a .
LEBEN, C., V. RUSCH and A.F. SCHMITTHENNER. The colonizationof soybeanbuds
glycineaand other bacteria.Phytopathology,SE, 1677-1681,
by Psuedomonas
1968b.
LEBEN, C., M.N. SCHROTH and D.C. HILDEBRAND. Colonizationand movementol
Pseudomonassyringaeon healthy bean seedlings.Phltopathology, 60, 677-680,1970.
LINDEMANN, J., D.C. ARNY and C.D. UPPER. EpiphyticPseudomonas
syringoepopul^
tion sizegreaterthan a thresholdlevelis predictiveof brown spotincidenceon snapbeans
Phytopathology,71, 890 (Abstr.), 1981.
LINDOW, S.E. The role of bacterialice nucleationin frost injury to plants,Annu. Rev
Phytopathol.,21, 363-384,1983.
MARSHALL, K.C. Bacterialadhesionin naturalenvironment.In MicrobialAdhesionto Sur
faces, editedby R.C.W. Berkeley,J.H. Lynch, J. Melling, P.R. Rutter and B. Vincent
pp. 187-196,Ellis Horwood, Chichester,1980.
and hypersen
MAZZIJCCHL U., C. BAZZI andP. PUPILLO. The inhibitionof susceptible
sitive reactions by protein-lipopolysaccharidecomplexes from phytopathogeni
pseudomonads:relationshipto polysaccharide
antigenicdeterminants.Physiol. Plant
Pathol.,14, l9-30, l9?9.
of Pseudomon
MAZZUCCHI,IJ.,C.RAZZI andP. MEDEGHINI BONATTL Encapsulation
syringaepv. tabaci in relationto its growth in tobaccoleavesboth pretreatedand not
Physiol.Plant Pathol.,2l,l05-112,
pretreated
with protein-lipopolysaccharidecomplexes.
1982.
glycoproteinfrom
MELLON, J.E. and J.P. HELGESON.Interactionof a hydroxyproline-rich
tobaccocalluswith potentialpathogens.Plant Physiol.T0' 401-405'1982.
MILLER, T.D. and M.N. SCHROTH. Monitoring the epiphytic population of Erwinia
amylovoraon pear with selectivernedium.Phytopathology,62, lL'l5'1182,1972'
glycineaon the surfaceof soy
MEW, T.W. and B.W. KENNEDY. Growth of Pseudomonas
bean leaves.Phytopathologl, 61, 7 15-716,1971.
MEW, T.W. and B.W. KENNEDY. Seasonalvariation in populations of pathogen
Pseudomonason soybeanleaves.Phytopathology, 12, 103'105' 1982.
MORT, A. and W. BAUER. Compositionof the capsularand extracellularpolysaccharid
of Rhizobiumjaponicum:changeswith cultureageand correlationwith bindingof soy
1980.
beanseedlectinto the bacteria.Plant Physiol.,66,158-163,
and nodulationby Rhizobiummeliloti. Science,213'1513-1514'1981'
PANOPOULOS,N.J. and M.N. SCHROTH. Roleof flagellarmotility in the invasionof bea
leavesby Pseudomonosphaseolicola.Phltopathology, 64, L389-1397,L9'74.
PETHICA, B.A. Microbial and cell adhesion.In Microbial Adhesion to Surfaces,edited b
R.C.W. Berkeley,J.H. Lynch, J. Melling, P.R. Rutter and B. Vincent,pp' l9-45' Elli
Horwood, Chichester,1980.
POLITIS, D.J. and R.N. GOODMAN. Localizedcell wall appositions:incompatibilityrespon
pisi. Phytopathology,6E,309-316,1978'
of tobaccoleaf cellsto Pseudomonas
of Er
POLITIS, D.J. and R.N. GOODMAN. Fine structureof extracellularpolysaccharide
wina amylovora.Appl. Environ. Microbiol.,40, 596-607,1980'
PREECE, T.F. and W.C. WONC. Quantitativeand scanningelectronmicroscopeobserv
tions on the attachment of Pseudomonastolaasii and other bacteria to the surface o
Agaricus bisporus. Physiol. Plont Pathol., 21,251-257, 1982.
PUEPPKE, S.G. and T. HYMOWITZ. Screeningthe genusGlycine subgenusClycine for th
120,000dalton seedlectin. Crop 9ci,22,558-560, 1982.
PULL, S.P., S.G. PUEPPKE, T. HYMOWITZ and J.H. ORF. Sovbeanlines lacking th
120,000-daltonseedlectin. Science,200, 1277-12'19,1978.
RAYMUNDO, A.K. and S.M. RIES. Chemotaxisof .Erwiniaamylovora. Phytopathology' 70
1980.
1066-1069.
on th
ROEBUCK, P., R. SEXTON and J.W. MANSFIELD. Ultrastructuralobservations
developmentof hypersensitivereaction in leavesof Phaseolusvulgariscv. Red Mexica
phaseolicola(race1). Physiol.Plant Pathol.,12, l5l-157
inoculatedwith Pseudomonas
1978.
ROMEIRO, R. oS., A.L. KARR and R.N. COODMAN. Erwlria amylovoracell wall rece
tor for appleagglutinin.Physiol. Plant Pathol,,19, 383-390'1981.
ROUSE,D.L, S.S.HIRANO, J. LINDEMANN, E.V. NORDHEIM and C.D. UPPER.Acon
with pathogenpopulationmultiplicationindependeno
ceptualmodelfor thosediseases
diseasedevelopment.P h! t opat hology, 7 l' 90I (Abstr'), I 98l.
SCHMIDT, E.L. Initiation of plant root-microbeinteractions.Annu. Rev.Microbiol,, 3
355-376,1919.
SCHNEIDER,R.W. andR.G. GROGAN. Tomatoleaftrichomes,a habitatfor residentpopul
tion of Pseudomonast omato. Phttopnt hology, 61, 898-902,1977.
SCHNEIDER, R.W. and R.G. CROCAN. Influenceof temperatureon bacterialspeckof tomat
Phytopathol. News,12,204 (Abstr.), 1979.
SEQUEIRA, L. Indugtion and. suppiessionof the hypersensitivereaction caused b
phytopathogenic
bacteria:specificand non-specificcomponents.ln Specdicityin Pla
Diseases,
editedby R.K.S. Wood and A. Graniti, pp. 289-306,PlenumPress,New York
t976.
specificity.Annu. Rev.Phytopatho
SEQUEIRA, L. Lectinsand their role in host-pathogen
1 6 , 4 5 3 - 4 8 11,9 7 8 .
SEQUEIRA, L. Recognitionbetweenplant hosts and parasites.In Host-parasiteInterface
editedby B.B. Nickol, pp.7l-84, AcademicPress,New York, 1979.
in plants,Annu. Rev.Microbiol., 37,51-7
of inducedresistance
SEQUEIRA, L. Mechanisms
1983.
interactions.Biol. cell,51; 281-286'198
SEQUEIRA, L. Recognitionsystemsin plant-pathogen
preparationsthat induceor-preventth
cell-free
Bacterial
U.
AINSLIE.
and
L.
SEdUEIRA,
reactionin tobacco.I lth Int Cong. Bol., p. 195(Abstr'), 1969'
hypersensitive
SEeUilRA, L. and T.L. CRAHAM. Agglutinationof avirulent strainsof Pseudomon
by potato lectins.Plyslol. Ptant Pathol', 11,43-51, 19'17'
solanacearum
C.
GAARD and c.A. DE zoETEN. Interactionof bacteriaand host ce
L.,
SEQUEIRA,
Physiol.Plant Pathol" l0'43'50,197
of induceresistanee.
walls:its relationto mechanisms
f or detectingPseudomonassyringaepv . tomato andXonthomonascomrystris pv , vesicator
inseedsandsymptomlessleaves.oftomatoandpepper.
J.Appl.Bacteriol.,53,37l-j77,
1982b.
SIGEE, D'C. and H.A. S. EPTON. Ultrastructureof Pseudomonasphaseolicotainresistan
and susceptibleleavesof French bean. Physiol, Plant Pathol., 6,29-34;1975.
SICEE, D.C. and H.A.S. EPTON. Ultrastructuralchangesin resistantand stsceptiblevarietie
of Phaseolus vulgaris following artificial inoculation with Pseudomonasphaseolicela
Physiol. Pldnt Pathol,9, l-8, 1976.
SIJAIU, K., A.L. I(ARR and R.N. COODMAN. Comparisonof the extracellular:polysaccharid
produced by.E'rwiniaamylovora in apple tissueand culture medium, Physiol, Plont Pathol.
22,22t-23t, t983.
SINC' V.O. and M.N. SCHROTH. Bacteria-plantcell surfaceinteractions:activeimmobiliza
tion of saprophytic bacteria in plant leaves,Scrence,191, 7 59-761,lg77.
SJULIN, T.M, and S.V. BEER. Mechanismof wilt induction by amylovorin in cotoneast
shootsand its relationto wilting of shootsinfectedbyErwinio amylovora.Phytopatholog
6E,89-94,1978.
SLUSARENKO, A.J. and R.K.S. WOOD. Differential agglutination of races I and 2 o
Pseudomonasphaseolicola by a fraction from cotyledons of Phoseolus vulgaris cv. Re
Mexican.Physiol. Plant Pothol., lt, lE7-193,1981.
SLUSARENKO,A.J. and R.K.S. WOOD. fuglutination of Pseudomonaspha,seoticotabyp
polysaccharide
from leavesof Pl aseolwvulgaris,Phlsiol, Plant Pathol,,23,2l1-2n , 198
SMITH, H. Microbial surfacesin relation to pathogenicity,Bacterial Rev.,47S-5A0,1917
SMITH, J.J. and J.W. MANSFIFLD. Ultrastructureof interactionsbetweenpseudomona
and oat leaves.Physiol. Plant pothol,, 2l,259-2ff, 1982.
SMITI-EY, D.R. and S.M. McCARTER. Spreadof Pseudomonossyringae pv. tomato an
role of epiphyticpopulationsand environmentalconditionsin diseasedevelopment, Plan
Dis., 66; 7 l3-7 14, 19E2.
SOROKER, E., Y. BASHAN and Y. OKON. Reproducibleinduction of cavity spot in carrot
and physiologicaland microbial changesoccurring during cavity formation. Soil Biol
Biochem.,16, 541-548,1984.
STADT' S.J. andA.W. SAETILER. Effectof hostgenotypeonmultiplicationofPseudomon
p haseolicola.Phytopathologll 71, I 307-1310, I98|.
STALL' R.E. and A.A. COOK. Multiplication o f Xonthomonasvesicotoriaandlesiondevelop
ment in resistantand susceptible
pepper.Phytopathology,56,ll52-1154, 1966.
STALL, R.E. and A.A. COOK. Evidertcethat bacterialcontact with the plant cell is necessa
for the hypersensitivereaction but not the susceptiblereaction. Physiol. Plant Pothol,
14,77-84,lg7g.
STEMMER, P. and L. SEQUEIRA. Pili of plant pathogenicbacteria. Phytopathology, Tl
906 (Abstr.),l9El.
SUTTON, T.B. ahd A.L. JONES. Monitoring of Erwinia amylovorapopdations on applein
relation to diseaseincidence.Phytopathology,65,1009-1012,
1975,
THOMSON, S.V., M.N. SCHROTH, W.J. MOLLER and W.O. REIL. Occurrenceof fire
blight of pears in relation to weather and epiphytic population of Erwinia amylovora
Phytopathology,65, 353-358,1975.
TSIEN, H.C., M.A. JACK, E.L. SCHMIDT and F. WOLD. Lectinin five soyaeancultivar
previouslyconsideredto be lectin-negative.
Planta, 15t, 128-133,1983.
VANCE, C,P. Rhizobiun infection and nodulation: a beneficial plant disease?Annu. Rev
Microbiol., 37, 39942/' 19E3.
VENETTE, J.R. How bacteria find their hosts. In PhytopathogenicProkaryotes, edited by
M.S. Mount and G.H. Lacy, Vol. 2, pp.4-30, AcademicPress,New York, 1982.
WALLIS, F,M. Ultrastructuralhistopathologyof tomato plantsinfectedwith Corynebacteriu
michiganense.Physiol. Plant Pothol., 11,333.342, 1977.
Pothol., t, 37l-378, lgn.
WELLER, D.M. and A.W. SAETTLER. Colonizationanddistributionof Xanthomonas
and Xanthomonasphaspoli var. Juscansin field-grown navy beans.Phytopatholo
500-s06,1980.
WHATLEY, M.H., J.S. BODWIN, B.B. LIPPINCOTT ANdJ.A. LIPPINCOTT. R
Agrobacterium cell envelopelipopolysaccharidein infection site attachment.Infe
mun., 13, 1080-1083,1976.
.. YOLING, J.M. andA.M. PATON. Developmentof pathogenicand saprophyticbacteria
. tions in plant tissue. In Proceedingsof Intemational Conference,Plant Patholo
Bacteriology, edited by H.P. Maas Geesteranus,pp: 77-80, Wageningep,1972
YUNIS, H., Y. BASHAN, Y. OKON and Y. HENIS. Weatherdependence,
yield los
control of bacterial speck of tomato causedby Pseudomonostomoto. Plant D
937-939,1980.