Role of Soil Microbial Community in Suppression of
Rhizoctonia Stem Rot of Cauliflower
Final Report - December 1998
Principal Investigator:
Joseph Hancock
(Formerly) Department of Environmental Science, Policy and Management
Division of Insect Biology
201 Wellman Hall
University of California
Berkeley, CA 94720
(510) 643-9223, jchanc@aol.com
Other Investigators:
Andrew Magyarosy, Division of Insect Biology, UC Berkeley
J.O. Becker, Department of Nematology, UC Riverside
Steve Koike, Farm Advisor, UCCE Monterey County
Don May, Farm Advisor, UCCE Fresno County
Eva Poinar, Division of Insect Biology, UC Berkeley
Location of project: Fresno County
Commodities: Cauliflower
Funding:
FY 1996-97: $9,200
FY 1997-98: $9,200
Table of Contents:
Summary
Objectives
Accomplishments
Literature Cited
Soils from sample sites 100 feet apart in a field at the West Side Research and Extension Center (Fresno Co.) were consistently either suppressive (DS) or conducive (DC) to Rhizoctonia wirestem (= stem rot, RSR) disease of cauliflower (RWS) in September 1996 and March and June 1997. Populations of general bacteria and fluorscent pseudomonads were similar in magnitude in the DS and DC soil samples in both Sept. 1996 and Mar. 1997 when the differences in suppressiveness were greatest between the two sites. Actinomycetes were up to 8-fold higher in the DS soil than in the DC soil. Microbial "metabolic fingerprints" of DS and DC soils were similar. When seedlings of cauliflower were grown in transplant-plug trays, RWS was less severe when transplants grown in DS soil were planted into Rhizoctonia infested UC mix (very disease conducive) then transplants grown in DC or heat-treated DS soil. Protection against RWS by plug-transplants did not seem to be a consequence of physical interference with the pathogen because when plants were grown in the heat-treated DS soil or vermiculite the plug-transplants in these soils were more susceptible to RWS than transplants grown in DS soil. The degree of disease suppressiveness of soil from the suppressive site declined progressively over the course of the year from Sept. 1996 to Sept. 1997. In Sept. 1997 the soils were conducive at both sites. Disease suppressiveness was destroyed when soil was heated at 40 C or higher for an hour, which suggested that suppressivness had a biological basis. When soil was washed from roots of plug-transplants, plants grown in DS soil were as susceptible to RWS as those grown in DC soil. This indicated that suppressiveness was protective rather than an enhancement of plant resistance to RWS. In support of this interpretation, we found that protection against RWS was lost when efficacious biological control agents isolated from DS soil were washed out of the UC mix plugs to which they had been applied. Screening of a large number of isolates of actinomycetes from DS soils resulted in the recovery of a few isolates of Streptomyces species that protected cauliflower seedlings from RWS when seedlings had been grown in plug trays in UC mix drenched with inoculum of these microbes. It was found with the plug-transplant system, that some commercial and locally formulated potting mixes suppressed RWS without amendment. Results of this work showed that the suppressiveness of certain soils to RWS is transferrable to plug-transplants, that certain actinomycetes from DS soils can infer suppressiveness to RWS and that potting mixes that are DS could offer a practical means of avoiding seedling diseases after transplanting into pathogen infested soils.
- To establish microbial community profiles or "signatures" (based on growth on different nutrient sources) in Rhizoctonia stem rot (RSR) (= Rhizoctonia wirestem disease, RWS) suppressive and conducive soil field sites at the West Side Research & Extension Center, Fresno Co.
- To determine changes in the microbial community profiles associated with loss of RWS suppressiveness associated with mild heat treatments.
- To construct microbiological media selective for microbes present in RWS suppressive soils but not in RWS conducive soils (natural soils and in those where suppressiveness is lost with mild heat treatments) for the isolation of these microbes and for measuring the quantitative and qualitative differences in individual microbes between suppressive and conducive soils.
- To measure RWS in soils (sterilized soil, conductive soil, and potting mixes) amended with microbes most affected by mild heat treatment of Rhizoctonia suppressive soils.
Objectives 1 and 2: Two parallel transects (transect-A and transect-C) were laid out in adjacent fields (Fields A & C) bisected by a parallel access road at the West Side Research and Extension Center (WSREC). Transects were 85 feet apart and each was 30 feet from the shoulder of the access road. Soil samples were taken from 4 square meter areas from 3 areas 100 feet apart on each transect (total of 6 samples per sample date) in Sept. 1996 and Mar. 1997. Samples were also taken from transect -C in June and Sept. 1997. A single soil type encompassed all of the transect sample sites (Panoche silty clay loam). The field with transect-A was cropped continuously to tomatoes for 10 years whereas the field with transect-C was cropped to barley, chickpea, and tomatoes in 1995, 1996, and 1997, respectively. It was found in the September 1996 sampling that the westerly sample areas (A1 & C1) in both transects were disease conducive (DC) whereas the soil samples from the other four sample areas were moderately (A2 & C3) to highly disease suppressive (DS) (A3 & C2) to the RWS disease. In the March sampling, the soil in C2 sample area was highly DS whereas the degrees of suppressiveness in other sample areas ranged from DC to moderately DS. The C1 and C2 sample areas were DC and DS, respectively, in June 1997 but were similar in degrees of suppressiveness in September 1997. It was found in a number of tests that suppressiveness in DS soils was expressed at higher inoculum densities then that found in DC soils or UC mix.
We measured the actinomycete populations in RWS-suppressive and conducive soils (Rowbotham and Cross, 1977), and we measured total bacteria as well as the nonfluorescent and fluorescent pseudomonad populations. The actinomycete and fluorescent pseudomonad populations were unusually high in both the DS and DC soils from WSREC. The populations were somewhat higher in the C2 (DS) soil then the C1 (DC) soil but these differences were less then an order of magnitude. Nutrient-use "metabolic signatures" in soil samples were determined by measuring the consumption of a wide variety of simple organic nutrients using the BIOLOG method. Dendrograms allowed a comparison of C1 and C2 soils at different sample periods on their "relatedness." There were significant differences between sample periods in "metabolic signatures" but there was greater relatedness between C1 and C2 "metabolic signatures" during the periods when differences in suppressiveness between C1 and C2 were greatest! Indeed, when differences in suppressiveness between C1 and C2 did not occur in September 1997, the differences in the "metabolic signatures" were greatest.
Objectives 3 & 4: Changes in microbial indicator populations (general molds, Fusarium and Pythium spp., and fluorescent pseudomonads) in soil were undetected after temperature treatments of 45 C or less for 1 hr - even though disease suppressiveness was measurably reduced. There also were no significant changes in populations of bacteria or species of fungi on feeder roots of plants grown in this soil after this heat treatment. However, we found incipient changes in "metabolic signatures" after DS soil was heated to 45 C but these changes were insignificant.
Protection against RWS by DS soils (C2) did not seem to be a consequence of a physical barrier against the pathogen because we found that when plants were grown in autoclaved soil, UC mix, or vermiculite that the plug-transplants were more susceptible to RWS than those plug-transplants grown in nonautoclaved soil. Washing suppressive soil from roots of plug-transplants increased their susceptibility to RWS when transplanted into the Rhizoctonia-infested UC mix. Plug-transplants grown in conducive soil, autoclaved soil or vermiculite were equally susceptible to RSR when roots were washed or unwashed, which indicated that root wounding or soil nutrients were not the basis for changes in susceptibility or differences in growth. Apparently, the loss of soilborne organisms from untreated DS soil plugs when they were washed was responsible for increased susceptibility to RWS Moreover, loss of protection of transplants against RWS by root-washing indicated that surviving plants were not rendered resistant to RWS by changes in the plants immune system. These investigations demonstrated that organisms in the raw, suppressive soil were transmitted with the plugs and afforded protection against RSR even in a normally conducive environment. This finding supports the feasibility of using plug-transplants as a biological means of controlling wirestem and damping-off diseases of a variety of crops caused by Rhizoctonia solani.
We screened nearly a hundred isolates of actinomycetes from the C2 site and a soil from the Armstrong experimental site at UC Davis for biological control activity against RWS. We found several isolates that showed promise as biological control agents. Figure 9 shows data from three experiments with nine of the most promising isolates and several mixtures (cocktails). There was variation between experiments with some strains such as with isolates 05 and 081. Other isolates, such as D11 and 071, showed greater consistency in protection of cauliflower seedlings against RWS. The GC-FAME method was applied in their identification and compared with the actinomycete GC-FAME database used by Microbe Inotech Laboratories, Inc., (St Louis, MO). It was determined that D11 most closely resembled Streptomyces halstedii scabies and 071 most closely resembled S. violaceusniger violaceusniger. The similarity coefficents for D11 and 071 were 0.702 and 0.131, respectively. It is concluded that 071 is an unknown species of Streptomyces and that further work is needed to insure the correct identity of D11.
We found that locally prepared and commercial potting mixes (also called soilless potting mixes) differed markedly in their natural DS to RWS. We found that UC mix varied in its DS, which was a problem for the standard pathogenicity test using plug transplants. However, we discovered that reproducible results were attained when we used UC mix within a few days of its preparation. (Similar results were found in tests with Pythium damping-off.) In further tests, we compared UC mix with other potting mixes. Here we found that UC mix was the most DC potting mix toward RWS but that certain commercial mixes were also DC. The UC Davis mix (coarse sand, compost, peatmoss1:1:1 v/v, unpasteurized) was very DS. Of the commerical oils tested, the "Original Super Soil" was the most DS.
Inspite of the success of recovery of suppressive isolates of microbes from DS soils, it is unlikely that these isolates alone account for the disease suppressive qualities found at certain sites at the WSREC. However, these results indicate that plug-transplant delivery of biologicals is a practical means of protecting transplants from RWS and, coupled with the use of DS potting mixes, plug-transplant systems could offer a practical means of reducing disease losses after transplanting seedlings into pathogen infested DC fields.
Rowbotham, T.J., and Cross, T. 1977. Ecology of Rhodococcus coprophilus and associated actinomycetes in fresh water and agricultural habitats. J. Gen. Microbiol. 100:231-240.