Interactions between the vesicular-arbuscular mycorrhizal fungus Glomus fascicuhtum and Aphanomyces euteiches root rot of peas

Download Interactions between the vesicular-arbuscular mycorrhizal fungus Glomus fascicuhtum and Aphanomyces euteiches root rot of peas

Post on 03-Oct-2016

212 views

Category:

Documents

0 download

Embed Size (px)

TRANSCRIPT

<ul><li><p>Phytopath. Z., 114, 3 1 - 4 0 (1985) 1985 Verlag Paul Parey, Berlin und HamburgISSN 0031-9481 /InterCode: PHYZA3</p><p>Agricultural Research Department, Rts0 National Laboratory,DK-4000 Rosktlde, Denmark</p><p>Instttute of Thallophytes University of Copenhagen,0ster Fartmagsgade 2D, 1353 K, Denmark</p><p>Interactions between the vesicular-arhuscular mycorrhizalfungus Glomus hscicuhtum and Aphanomyces euteiches</p><p>root rot of peas</p><p>S. RoSENDAHL</p><p>With one figure</p><p>Received April 2, 1984</p><p>Abstract</p><p>Interactions between Glomus fasdculatum and Aphanomyces euteiches root rot of peas (Pisumsativum), were studied in pot experiments using irradiated soil.</p><p>Infections with the pathogen were suppressed by VAM when plants were challenge inoculatedafter two weeks. No reduction of the pathogen was detected when the plants were inoculated withboth fungi at the same time.</p><p>The suppression of the pathogen, obtained by preinoculation with G. fasciculatum, was notreduced when the inoculum level of the pathogen was increased thirty times.</p><p>The induced resistance to A. euteiches in VAM plants was partially a systemic effect. When rootsystems were split into two halves, one with mycorrhiza and one with A. euteiches, the oosporeproduction was reduced in both root systems. The infection with the pathogen was only suppressedwhen both fungi were present in the same pot. The background for the induced resistance is discussed.</p><p>Interaktionen zwischen dem vesikular-arbuskularen Mykorrhiza PilzGlomus fasciculatum und der Wurzelfaule der Erbsen, hervorgerufen</p><p>durch Aphanomyces euteiches</p><p>Wechselwirkungen zwischen Glomus fasciculatum und Aphanomyces euteiches Wurzelfaule derErbsen (Pisum sativum) wurde in Experimenten studiert, bei denen Erde verwendet wurde.</p><p>&gt;?v enn I ilanzen nacri zwei iVocnen geimprt wurden, Konnten intektioneii durch den I atnogenmit VAM unterdruckt werden. Keine Verminderung des Pathogens wurde jedoch festgestellt, wenndie Pflanzen mit beiden Pilzen gleichzeitig geimpft wurden.</p><p>U.S. Copyright Clearance Center Code Stacemem: 0 0 3 1 - 9 4 8 1/85/1 401 "003 1 $ 0 2 . 5 0 / 0</p></li><li><p>-L^ ie Unterdruckung des Pathogens, die durch die vorherige Impfung mit G-fasciculatumcrrcicht wurde, wurde nicht vermindert, wenn die Inokulumdichte des Pathogens 30maI erhoht</p><p>Die induzierte Resistenz gegenUber A. euteiches in VAM-Pflanzen war teilweise ein systemi-schcr Effekt. Wenn Wurzelsysteme in zwei Halften geteilt wurden, eine Halfte mit Mykorrhiza unddie andere Halfte mit A. euteiches, war die Oosporenproduktion in beiden Wurzelsystemen reduzien.Die Infektion durch den Pathogen war nur unterdriickt, wenn beide Pilze in demselben Topfvorhanucn waren. Die Orunulsge riir die mduzierte Resistenz wird besprochen.</p><p>Introduction</p><p>The significance of vesicular-arbuscular mycorrhiza (VAM) in the resistanceof plants to pathogens, has attracted great attention in the latest years (SCHENCKand KELLAM 1978, SCHENCK 1981, DEHNE 1982).</p><p>In most investigations VAM decreased the incidence of several root-pathogenic fungi (DEHNE und SCHONBECK 1975) and nematodes (KELLAM andSCHENCK 1980), whereas no clear effect was obtained in others (BAATH andHAYMANN 1983).</p><p>The resistance to pathogens, induced by VAM, seems to depened on thetime interval between the mycorrhizal inoculation and the challenge infectionwith the pathogen, hence a reduction of the disease is only obtained if themycorrhizal infection is well established at the time of challenge (BARTSCHI etal.1981).</p><p>The increased resistance in VAM plants is described by SCHONBECK (1979) asa strictly local effect occurring oniy in the mycorrhizal areas, whereas, otherauthors found increased resistance in the non-mycorrhizal parts of root systemsas well (DAVIS and MENGE 1980).</p><p>DEHNE (1982) describe the interactions between VAM and root-pathogensto be based on modifications of the disease since very virulent isolates or increasedinoculum levels of the pathogen may reduce or even eliminate the beneficialeffects of VAM.</p><p>The mechanisms behind the induced resistance in VAM plants remainunexplained, but many different mechanisms have been proposed (SCHONBECK1979).</p><p>Phosphorus may be important (DAVIS and MENGE 1980, GRAHAM andMENGE 1982) and several mechanisms could be indirect effects of increasedphosphorus content in the VAM plants. Besides, the VAM plants are usually inbetter nutrional condition than the smaller non-VAM plants, and thereby lesssusceptible to the pathogens (DEHNE 1982). It is, therefore, important to workwith plants of the same size, either by using isolates of mycorrhizal fungi that donot give any growth response, or by adding phosphate to the non-VAM plants.</p><p>Aphanomyces euteiches DRESCHL. is described as the most destructive patho-gen on peas in several countries (PAPAVIZAS and AYERS 1974), and is regarded asthe principal limiting factor for pea production in several districts in USA(PFENDER and HAGEDORN 1983), Norway (SUNDHEIM and WIGGEN 1972) and thesouthern part of Sweden (OLOFSSON 1967). The importance of the pathogen hasnot yet been investigated in Denmark.</p></li><li><p>Interactions between the vesicular-arbuscular 33</p><p>No resistant pea cultivars have been found, and chemical control seems to beineffective or very expensive. Losses can mainly be reduced by avoiding artasheavily infested with the pathogen (PFENDER et al. 1981).</p><p>Few papers describe interactions between A. euteiches and other microor-ganisms, and no antagonistic microorganisms have yet been found (PAPAVIZASand AYERS 1974).</p><p>The purpose of this study was to investigate the influence of Glomusfasciculatum (THAXTER) GERDEMANN &amp; TRAPPE, VA mycorrhiza onA. euteiches root of of peas, and to examine the nature of the mechanismsinvolved in the induced resistance in VAM plants.</p><p>Material and Methods</p><p>2.1 Host plants and treatments</p><p>Peas were grown in a 2 1 mixture of sand and clay loam (pH 7) containing 12 ppm phosphorus(Olsen P). The mixture was irradiated with 0.8 Mrad (10 MeV electron beam) in order to killmycorrhizal propagules.</p><p>Two seeds of Pisum sativum L. (cv. Bodil), pregerminated on moist filterpaper for 48 hours andsurface sterilized m 70 % ethanol for two minutes, were planted in each 10 cm pot holding 1 kg of thegrowing medium. At planting, the pots were inoculated with 20 ml of a dense suspension ofRhizobium leguminosarum (RiS0 stram 1 a).</p><p>with A.eutetches, 4) inoculated with both G. fasciculatum and A. euteiches, were included in allexperiments except ror the split-root experiment (Section 3.3.2). The treatments m tnis experiment aregiven in table 3.</p><p>Six replicates of every treatment were tnade in all experiments. The pots were placed in agreenhouse at 2025C and watered as necessary.</p><p>2.2 Mycorrhizal inoculum</p><p>The G. fasaculatum inoculum was produced on maize grown in autoclaved, expanded clayLeca* for three months in 2000 cm' pots (H.-W. DEHNE pers. comm.). The inoculum containedinfected roots, spores and expanded clay particles with external mycelium, ihe VAJVi plants wereinoculated with approximately 10 g inoculum per pot at planting, and similar amount of uninfectedmaize roots and expanded clay were added to the non-VAM plants.</p><p>2.3 Aphanomyces euteiches inoculum</p><p>The A. euteiches isolate was obtained from L Sundheim, As, Norway. The culture wasmaintaineu on corn mesl aga.r at 4 C Zoospore inoculum was obtained according to the method oi</p><p>100ml maltose-peptone brotn medium (MPB). Ine medium was decantated aiter rive days, andreplaced with 100 ml sterile tap-water which was again replaced by 50 ml sterile distilled water aftertwo hours. After seventeen hours at 21 C the mycorrhizal mats were strained off and the zoospore</p><p>The plants were inoculated by pouring 10 ml of the zoospore suspension on the soil surface.Control plants received 10 ml distilled water.</p><p>2.4 Determination of dry weight, phosphorus content and degree of infection</p><p>Shoot and root dry weight were determined at harvest. The phosphorus concentration wasmeasured by the molybdate-blue method (MURPHY and RILEY 1962).</p><p>Representative root samples of each of the six replicates, was cleared in KOH and stained inTryphanblue according to PHILLIPS and HAYMAN (1970), and the infection with both mycorrhiza and</p><p>Phylopaih.Z.,Bd. 114, Hett 1 3</p></li><li><p>A. eutctchcs were estimated as percent mfected root length by the grid-line intersect method (GIOVAN-</p><p>Infection with A. euteiches was confirmed by occurence of oospores in the cortex, and theni\ corrhizal infection was identified by occurence of vesicles, arbuscles or chlamydospores.</p><p>The number of oospores per mm^ infected root was determined microscopically by measuringthe length and the diameter of the examined root piece. At every intersection between an infected rootand the grid-line, used for determination of infections, the number of oospores was counted in a250//m long root cylinder. For each of the six replicates, thirty such root cylinders were examined.</p><p>Experiments and Results</p><p>3.1 Influence of different challenge timesThe influence of the time interval, between inoculation with the mycorrhizal</p><p>fungus and the pathogen, on the interaction between the two fungi, was assessed.Three groups of plants, including all four treatments, were planted at 14 daysintervals and then challenge inoculated simultaneously 28 days after the firstplantmg (see Table 1).</p><p>All plants were harvested two weeks after challenge inoculation and dryweight of shoots and roots were determined along with the infection withG. fasciculatum and A. euteiches (Table 1).</p><p>VAM suppressed the infection with the pathogen when the challengeinoculation was made two and four weeks after planting (Treatment 8 and 12),whereas the pathogen was not influenced by VAM when the plants wereinoculated with both fungi at the same time (Treatment 4).</p><p>Aphanor</p><p>No.</p><p>nt Plan'</p><p>G.faT</p><p>tum. A:</p><p>t age (days) AT:. with:A. eut. harvest</p><p>T.</p><p>:inocula</p><p>Dry</p><p>Shoo</p><p>uble 1</p><p>ted wit</p><p>weigh,(g)</p><p>h A. euteich</p><p>% Roo: VAM</p><p>itlength^ mjA. eut. V A M ' A.</p><p>md time ofl.fasacuU-</p><p>2M3A4M-(-A -5C 06M7A8M-t-A -9 C 0</p><p>lOMl l A12M-I-A -</p><p>P</p></li><li><p>IS between the vesicular-arbusc 35</p><p>Estimated as percentage infection, VAM infection reached maximum .ihcronly two weeks. A reduction of the mycorrhizal infection was observed onlywhen plants were inoculated with both fungi at the same time (Treatment 4).</p><p>In spite of different values of percent infections with A. euteiches in the non-VAM plants, infections expressed as mg infected root were similar at all harvestsand thus independent of the root weight.</p><p>3.2 Effect of increased pathogen inoculum levelTo determine the influence of increased inoculum, six different zoospore</p><p>concentrations were used. The experiments were carried out at different times butunder similar conditions. The plants were inoculated with G. fasciculatum fourweeks before challenge inoculation and harvested two weeks after challenge.</p><p>The infection with A. euteiches were determined in the VAM and the non-VAM plants, and plotted against the zoospore concentration (Fig. 1).</p><p>The infection with the pathogen increased with increasing inoculum level inthe non-VAM plants, whereas the infection in the VAM plants was almostindependent of the zoospore concentration. The slope of the lines were signific-antly different (P</p></li><li><p>Porccntagcfre,,</p><p>zoosp./pot.</p><p>A.</p><p>no</p><p>infection types</p><p>euteiches infec</p><p>infc. M</p><p>T.</p><p>both futs VAM</p><p>al obserA</p><p>able2s of VAM plants</p><p>ngi). The right haand non-VAM r</p><p>vedM + A</p><p>challenge inoi</p><p>ootreq^ally</p><p>TotalObsv.</p><p>:ulated with differen</p><p>ts the hypothesis tha</p><p>^.^-Values'</p><p>' All values are significant (P &gt; 0.05)</p><p>in Section 3.2. Roots of the plants inoculated with both fungi were examinedmicroscopically and the infection pattern was recorded for each intersectionbetween a root and the grid-line. The following categories were used: 1) noinfection, 2) infection with G.fasdculatum, 3) infection with A. euteiches,4) infection with both G.fascieulatum and A. euteiches (Table 2).</p><p>Significant (P &gt; 0.05) x' values were obtained at all six inoculum levels ofthepathogen, when the hypothesis that the pathogen infected VAM and non-VAMroots of the same plant equally was tested.</p><p>3.3.2 Infection pattern in a split-root experimentA split-root experiment was used in order to examine whether the mycorrhi-</p><p>zal infection had any influence on the degree of pathogen infection when the twofungi were in separate pots.</p><p>Seeds were pregerminated for three days, whereafter the tip of the radicleswere severed in order to promote formation of side roots. After one week m</p><p>Table JDry weights and infections with G.fascieulatum and A. euteiches in a split-root experiment (Section3.3.2). (0: uninoculated control, M: inoculated with G.fascieulatum, A: inoculated with A. euteiches.</p><p>The plants were inoculated with G.fascieulatum four weeks before challenge</p><p>Tr(</p><p>Left</p><p>MOMM</p><p>:atment</p><p>Right</p><p>oAAM + A</p><p>Shoots</p><p>1.96a1.95a2.D6a2.26b</p><p>Dry</p><p>left</p><p>0.44vO.38x0.47v0.45V</p><p>weights (fRoots</p><p>right</p><p>0.26w0.29w0.24w0.42v</p><p>;)</p><p>total</p><p>0.70z0.67y0.71yz0.87Z</p><p>G.fasleft</p><p>88.0a0</p><p>85.7a85.9a</p><p>/o Rootl:cicul.right</p><p>000</p><p>83.9a</p><p>ength mfeA.e</p><p>left</p><p>0000</p><p>cted'uteiches</p><p>right</p><p>038.4x38.1x10.6y</p></li><li><p>Phosphor</p><p>Left</p><p>MOMM</p><p>Values in</p><p>CO</p><p>t</p><p>Rig</p><p>oAAM-</p><p>thes</p><p>ahte</p><p>ntrol, M: inoculat.uptakeed witl</p><p>PhosphorusShoots</p><p>;ht left</p><p>0.23a0.15b0.26a</p><p>ame column follow</p><p>the side roots</p><p>0.57x0.16y0.57X</p><p>ved by</p><p>Table</p><p>Rootsright</p><p>0.28z0.16y0.24z</p><p>F-test</p><p>separated \</p><p>4n exper</p><p>Shoot</p><p>4.50a2.84b5.24a</p><p>iment (Section 3.3.2) (0:</p><p>Phosphoi</p><p>left</p><p>0.58y2.66x</p><p>halves ai</p><p>us uptake (mg/</p><p>Mght</p><p>0.72z0.48y0.55y</p><p>Ltly different (P</p><p>ad placed in</p><p>un.no.uLacd</p><p>'plant)Total</p><p>7.663.548.45</p></li><li><p>38</p><p>No difference was detected in the oospore production, between mycorrhizaland non-mycorrhizal roots of VAM plants, although the oospore production wasreduced in the VAM plants, compared with the non-VAM plants.</p><p>Discussion</p><p>These experiments show that VAM infection can reduce root rot of peascaused by A. euteiches. This effect was increased with increasing time lapsebetween the mycorrhizal inoculation and the challenge inoculation with thepathogen (Section 3.1). This resuk is in agreement with BARTSCHI etal. (1981),who found that VAM did not protect roots of Chamaecyparis lawsoniana toPhytophthora cinnamomi unless the mycorrhizal infection was well estabhshed atthe time of challenge.</p><p>Percent infected rootlength was used as an estimate of the infection withA. euteiches, in order to avoid subjective estimates as e. g. "disease index" used inother experiments with root rot. However, a clear correlation was found hetweenpercent infected rootlength and the "disease index" in several pilot tests.</p><p>In the first experiment (Section 3.1) (Table 1), infected root weight was usedin order to compare infections in plants of different ages. By using this estimatethe problems in comparing infections in root systems of unequal size were met.However, both G. fasaculatum and A. euteiches will probably infect the finelateral roots more easily than the more hgnified main roots, and the obtainedvalues might therefore be over estimated.</p><p>A. euteiches was able to suppress the VAM infection in the plants inoculatedwith both fungi at the same time (Table 1, treatment 4). A similar effect wasreported by K...</p></li></ul>