Southern Regional Project S302
Biological Control of Soilborne Plant Pathogens for Sustainable Agriculture

ANNUAL REPORT OF COOPERATIVE REGIONAL PROJECT S-302
Supported by Allotments of the Regional Research Fund,
Hatch Act, as Amended August 11, 1955

January 1 to December 31, 2003

  Cooperating Agencies and Principal Leaders:

* Voting member in state

PERIOD COVERED: January 1 to December 31, 2003

DATE OF REPORT: January 2, 2004

MINUTES OF ANNUAL MEETING

The 2003 Annual Meeting of the Technical Committee S-302 Southern Regional Project
Sunday & Monday, November 2-3, 2003, Ft. Lauderdale, FL.

Chair: Don Huber

Secretary: Monica Elliott

Members Present: Mike Benson (NC), Craig Canaday (TN), Monica Elliott (FL), Charles Howell (USDA ARS, TX), Anthony Keinath (SC), Don Huber (IN), Boyd Padgett (LA), Craig Rothrock (AR), and Kenneth Seebold (GA). Robert Nowierski (CSREES Representative) joined us the morning of November 3. Administrative advisor, Craig Weidemann, could not attend.

The meeting was hosted by Monica Elliott at the University of Florida-IFAS Fort Lauderdale Research and Education Center. It convened at 8:00 AM on November 2. Minutes of last years' meeting were approved. Members were asked to submit annual reports electronically to Don Huber for compilation of the project annual report. Hard copies of state annual reports were available to those in attendance.

Summary data for the 2003 S-302 Regional Project evaluation of biological control agents for cotton seedling diseases were reviewed, with a discussion led by Craig Rothrock. Bill Batson was unable to attend in person because of a family illness, but he had forwarded summary tables to Florida for our review. The experiment evaluated six biological control agents, with and without fungicides, as seed treatment applications for two cotton cultivars. Stand count was determined as a measurement of disease pressure. Significant effects were observed due to cultivar at 5 locations (AR, GA, LA, MS, NC), and due to treatment at the same locations. However, significant interactions between cultivar and treatment were only observed in NC. A potential negative impact was observed for treatments with Trichoderma virens isolate G-6 (AR, LA) and YieldShield (Bacillus pumilus GB34) (LA, NC). The biological plus fungicide combinations were usually not significantly different from the fungicide alone treatment.

Craig Canaday, coordinator of the S-302 Regional Project evaluation of biological control agents for snap bean seedling diseases, led the discussion on the 2003 results. The experiment evaluated four biological control agents, with and without fungicides, as seed treatment applications for two snap bean cultivars, Bronco and Carlo. This was a repeat of the 2002 experiment, except that different cultivars were evaluated. Stand count was determined as a measurement of disease pressure. Significant treatment differences were observed in IN and TX. Significant cultivar effects were observed at 4 locations (AR, IN, SC and TN) and biological treatment at one location (TX). Unlike last year, a significant interaction was observed for chemical and biological treatments only in TX.

Monica Elliott discussed results from the seed assays associated with the cotton and snap bean experiments. As observed previously, the greatest variability in populations of biocontrol agents on seed between delivery and planting was observed with the commercial fungal treatment used on cotton. The addition of fungicides significantly increased populations of bacterial agents, but decreased populations of the fungal agents on both snap bean and cotton.

Plans were made for the 2004 snap bean trial. Two changes were proposed. First, a cultivar from the 2002 trial (Hialeah) and one from the 2003 trial (either Bronco or Carlo) would be the cultivars compared. Second, two additional chemical treatments would be added - one specific for Rhizoctonia and one specific for Pythium - as one means of determining pathogen pressure. The biological treatments and locations would remain the same.

Individual members made presentations of state reports under objectives 1 and 2 of the S-302 Regional Project. Kenny Seebold (GA) reported on results using various bacterial strains and Actigard to induce resistance in yellow squash to Phytophthora capsici. Mike Benson (NC) reported on the use of composted swine waste for suppression of Pythium root rot on poinsettia. Craig Rothrock reported on potential management of post-emergence damping-off (R. solani and Pythium spp.) with binucleate Rhizoctonia strains applied as in-furrow treatments and a Trichoderma strain applied as a seed treatment. Charles Howell (TX) reported that Trichoderma appears to be effective in preventing pre-emergence damping-off caused by Pythium spp. by inactivating seed exudates that stimulate Pythium germination. Boyd Padgett (LA) reported that there was an increase in disease due to Rhizoctonia when wheat was planted as a cover crop to prevent soil loss and conserve soil moisture, but that fungicides were useful in disease management. Tony Keinath reported that Fusaria were the most sensitive of the fungal groups assayed to biofumigation. Craig Canaday reported on evaluation of a PGPR product used in combination with Beauveria bassiana.

The business meeting was held the morning of November 3. Boyd Padgett was elected Secretary for 2004, to become chair in 2005. If acceptable to our companion projects in the North-Central (NC-125) and Western (W-147) regions, a joint meeting will be held in 2004 at a time and location to be determined, but most likely in October or November.

Following the business meeting, our CSREES representative, Robert Nowierski made a presentation regarding his areas of responsibility and programs of special interest to our group. Of concern to the group was the elimination of the biopesticide-based management program in the up-coming NRI 2004 program. The group expressed our concern with the new rules associated with APHIS permits and their negative impact on the ability of scientists to conduct research.

Meeting was adjourned at 12:00 PM November 3, 2003.

PROGRESS OF THE WORK AND PRINCIPAL ACCOMPLISHMENTS:

Objective 1. Selection, optimization and regional evaluation of biological control agents to include application techniques and enhancement of biological control agents to control diseases caused by soilborne plant pathogens.

Ten states (Nine test sites) were involved in the Regional Cotton Trials (AL, AR, FL, GA, IN, LA, MS, NC, TN, and TX). The fourteen treatments evaluated on two cotton cultivars (Deltapine 451B/RR and Paymaster 1218 BG/RR) included: Bacillus subtilis MBI 600 (Subtilex), B. subtilis GB03 (Kodiak), B. pumilus GB34 (YieldShield), B. subtilis GB03+ B. amyloliquefaciens GB99 (Bio Yield), Trichoderma virens G6, and T. virens/T. koningii fusant TV-117, alone and in combination with Vitavax-PCNB+ Allegiance, and the fungicide and non-treated controls. Significant treatment effects were observed at 5 locations (AR, GA, LA, MS, NC); however, significant interactions between cultivar and treatment were only observed in NC. A potential negative impact was observed for treatments with Trichoderma virens isolate G-6 (AR, LA) and YieldShield (Bacillus pumilus GB34) (LA, NC). The biological plus fungicide combinations were usually not significantly different from the fungicide alone treatment.

Ten states (AL, AR, FL, GA, IN, MS, NC, SC, TN, and TX) participated in the Regional Snap Bean Trials coordinated by Craig Canaday (TN). Four biological control agents (Bacillus subtilis MBI 600 (Subtilex), B. subtilis GB03 (Kodiak), Bacillus licheniformis SB3086 (710-145F), and Trichoderma harzianum T-22 HC), with and without captan + streptomycin were evaluated as seed treatments on two snap bean cultivars (Bronco and Carlo). A significant increase in stand was observed overall only with the chemical seed treatment. Significant cultivar effects were observed at 4 locations (AR, IN, SC and TN) and biological treatment at one location (TX). A significant interaction for chemical and biological treatments was observed only in TX.

The cooperative effort of Southern Regional Project S-302 also determines the stability, population, and purity of the biological agents applied to the seed from time of seed delivery to test cooperators until the seed is actually planted. Populations of most bacterial agents on seed were stable for the 4-8 weeks from delivery to planting. The greatest variability in populations of biocontrol agents on seed was observed with the fungal treatment on cotton. The addition of fungicides significantly increased populations of bacterial agents, but decreased populations of fungal agents on both snap bean and cotton. No significant differences between cultivars were observed when treatments were averaged across all locations.

Other Research under Objective 1.
Various techniques for enhancing efficacy of biocontrol agents are being evaluated. Actigard, an induced systemic-resistance activator (ISR) was evaluated with three new bacterial strains antagonistic to Phytophthora crown and fruit rot (P. capsici) (GA) and with binucleate Rhizoctonia (BNR) against R. solani seedling disease of cotton (AR). Infestation of soil with nonpathogenic strains of BNR significantly suppressed seedling disease caused by R. solani in both greenhouse and field tests (AR). There was no significant benefit of the ISR, but the three strains of BNR were as effective as the fungicide treatment used for comparison. Placement and timing of the biocontrol agents were important enhancements for R. solani control. BioYield, another induced systemic resistance activator, with and without two Plant Growth-Promoting Rhizobacteria (PGPR), increased tomato yield and reduced the number of culls from buckeye rot depending on the harvest time and cultivar (TN). A phosphite compound (FNX-100) reduced Phytophthora crown and fruit rot in the greenhouse, but did not increase yields of field-grown squash (GA). Combinations of Trichoderma virens and T. harzianum effective against Pythium damping-off (preemergence) with fungicides active against Rhizoctonia solani (post-emergence damping-off) increased cotton stands 72 to 80 % (TX). The addition of abiotic factors such as Al and composted animal wastes to potting mixes improved biological control of Phytophthora diseases (NC). Fourteen of over 912 Bacillaceae isolates screened in vitro for activity against R. solani were selected for further testing in vivo (FL). A reduction in post-emergence damping-off of marigolds was observed when the bacteria were applied just prior to transplanting, but not at seeding time (FL).

Objective 2. Improve understanding of mechanisms and applicability of biological control agents across different cultivars, environments, and cropping systems.

Two Bacillus isolates selected for biocontrol of post-emergence damping-off also restored the normal color to "iron chlorosis" Vinca plants independent of disease suppression (FL). As little as 50-60 ppm ferrous iron was found to inhibit germination of Exserohilum rostratum conidia and there was no inhibition with 200 PPM or more of ferric iron (FL). Glyphosate reduced Mn uptake and physiological efficiency, but increased take-all of wheat following glyphosate-resistant soybeans was indirect through modification of the soil microflora (IN). Combinations of PGPR with Beauveria bassiana biocontrol agent were evaluated to improve biocontrol efficacy (TN). Trichoderma appears to prevent pre-emergence damping-off caused by Pythium spp. by inactivating seed exudates that stimulate Pythium germination (TX). Increased Rhizoctonia observed when a wheat cover crop was used to prevent soil and moisture loss may necessitate fungicide applications to manage subsequent disease on vegetables (GA). Crop sequence and tillage influenced the availability of Mn for a subsequent crop and could account for the yield benefit of crop rotation for soybeans in a corn -soybean-wheat rotation and yield decline in no-till compared with fall chisel tillage programs (IN). A greater number of jumbo-grade onions were produced in Pyrenocheata terestris (pink root) infested soil by biofumigation with turnip and solarization, but solarization alone increased bacterial rots (sour skin and center rot) (GA). Soilarization decreased populations of Sclerotium rolfsii and Rhizoctonia solani, but not Pythium in soil (SC). Fusaria were the most sensitive fungal group to solarization. Populations of Rhizoctonia solani AG-3 from potato seed tubers are genetically diverse and research is determining the association of the M2 dsRNA and other dsRNA elements with population dynamics and virulence (NC). The Mn-oxidizing factor associated with virulence of Gaeumannomyces graminis var tritici (take-all) and Magnaporthe grisea (rice blast) is an extracellular protein approximately 50 kd size, but not manganese peroxidase or a common laccase (IN). Competition for organic matter is a possible mechanism for the interaction between the biocontrol fungus Laetisaria arvalis and Sclerotium rolfsii (OK).

IMPACT:

Pesticides in the environment, and public concern over food safety, have led scientists in Southern Regional Research Project S-302 to look at biological control as a central component of an ecologically based approach to Integrated Pest Management for plant diseases. A major deterrent to widespread adoption of biological control for soilborne plant diseases, however, has been the limited knowledge regarding activity and stability of potential biological control agents on multiple crops in diverse locations. This information is limited because biological control agents are initially discovered by university or USDA researchers and not by the agricultural industry. Development of large scale, multi-state evaluations of biocontrol agents are difficult for a single researcher or small company. This problem was solved through Cooperative research conducted by researchers in Alabama, Arkansas, Florida, Georgia, Indiana, Louisiana, Mississippi, North Carolina, South Carolina, Tennessee, and Texas under the auspices of Southern Regional Research Project S302. This project has demonstrated that several bacteria and fungi, either individually and in combination with standard seed treatment chemicals, are effective in reducing damping-off and seedling diseases of cotton and green beans across diverse environments and soil types. These "early season" crop diseases cost cotton growers between $9.78 (2001) and 19.46 (2002) million dollars annually in lost stand and replant costs. In addition to providing efficacy data on biocontrol agents, this research includes an analysis for stability and purity of the seed populations of these biological agents from time of seed delivery to planting in order to insure product quality and performance. The evaluation of potential biological agents for disease control in this manner provides new incentives for alternative controls of poorly managed soilborne plant diseases and helps understand inconsistent performance problems encountered with biological agents in the past.

WORK PLANNED FOR NEXT YEAR:

Work will continue on both project objectives. Regional testing of biological control agents on cotton and snap bean will continue to be the primary focus of the regional research effort. The modified protocol will evaluate the interactions between genotype, biological control agents (both bacterial and fungal), and chemical seed treatments. Researchers in several states will work together to better understand the mechanisms of biocontrol and the complex interactions between host, pathogen, biocontrol agent, and the environment.

PUBLICATIONS ISSUED AND MANUSCRIPTS APPROVED DURING THE YEAR:

Refereed Journal Articles:
Broschat, T.K. and Elliott, M.L. 2003. Effects of iron source on iron chlorosis and Excerohilum leaf spot severity in Wodyetia bifurcata. Hort Sci. (In Press).

Bush, B.J., Carson, M.L., Cubeta, M.A., Hagler, W.M., and Payne, G.A. 2003. Infection and fumonisin production by Fusarium verticillioides in developing maize kernels. Phytopathology 93:(In Press).

Ceresini, P.C., Shew, H.D., Vigalys, R., Rosewich-Gale, U.L., and Cubeta, M.A. 2003. Detecting migration in populations of Rhizoctonia solani AG-3 from potato in North Carolina using multilocus genotype probabilities. Phytopathology 93:610-615.

Diab, H., Hu, S., and Benson, D.M. 2003. Suppression of Rhizoctonia solani on impatiens by enhanced microbial activity in composted swine waste amended potting mixes. Phytopathology 93:1115-1123.

Elliott, M.L., Guertal, E.A., and Skipper, H.D. 2003. Rhizosphere bacterial population flux in golf course putting greens in the Southeastern United States. HortTechnol.: (In Press).

Hanson, L.E. and Howell, C.R. 2003. Elicitors of plant defense responses from biological control strains of Trichoderma virens. Phytopathology 93:(In Press).

Hollowell, J.E., Shew, B.B., Wilcut, J.W., and Cubeta, M.A. 2003. Weed species as hosts of Sclerotinia minor in peanut fields. Plant Dis. 87:197-199.

Howell, C.R. 2003. Mechanisms employed by Trichoderma species in the biological control of plant diseases: The history and evolution of current concepts. Plant Dis. 87:4-10.

Hwang, J. and Benson, D.M. 2003. Expression of induced systemic resistance in poinsettia cuttings against Rhizoctonia stem rot by treatment of stock plants with binucleate Rhizoctonia. Biol. Control 27:73-80.

Keinath, A.P., Harrison, H.F., Marino, P.C., Jackson, D.M., and Pullaro, T.C. 2003. Increase in populations of Rhizoctonia solani and wirestem of collard with velvet bean cover crop mulch. Plant Dis. 87:719-725.

Books, Book Chapters and Review Articles:
Cubeta, M.A., Mozley, S.E., and Porter, D. 203. Laboratory Exercises with Zoosporic Fungi. Pp. 99-110. In: Trigiano and Windam (eds.). Zoosporic Fungi. CRC Press, Boca Raton, FL.

Mozley, S.E., Leander, C.E., Porter, D., and Cubeta, M.A. 2003. Concepts: Zoosporic fungi. Pp. 91-98. In: Trigiano and Windam (eds.) Zoosporic Fungi. CRC Press, Boca Raton, FL.

Payne, G.A. and Cubeta, M.A. 2003. Biology and Detection of Human and Plant Pathogenic Fungi. Elsevier (In Press).

Symposia and other published works:
Bates, G.D., Rothrock, C.S., and Rupe, J.C. 2003. Possible mechanism for resistance to Pythium spp. in the soybean cultivar Archer. Proc. 8th Intern. Cong. Plant Pathol. 2:330.

Huber, D.M. 2003. Nutrient-disease interactions in the take-all disease of cereals. Proc. Symposium on take-all, April 2003, Temuco, Chili.

Huber, D.M. 2003. The role of plant nutrition in disease. Proc. Symposium Relacao entre Nutricao de Plantas e Incidencia de Doencas, March, 2003, Piracicaba, SP, Brazil.

Keinath, A.P., DuBose, V.B., Cantrell, J.P., and May, W.H. III. 2003. Evaluating foliar-applied fungicides to control wirestem on heading Brassica vegetables. Fungic. Nematic. Tests. 58:V010.

Rosskopf, E.N., Yandoc, C.B., and Lamb, E.M. 2002. Evaluation of PCC1210 for control of Phytophthora capsici on pepper. Pp. 85.1-85.3. In: Proc. 2002 Ann. Intern. Res. Conf. Methyl Bromide Alternatives Emissions Reductions..

Abstracts:
Bates, G.D., Rothrock, C.S., and Rupe, J.C. 2003. Possible mechanism for resistance to Pythium spp. in the soybean cultivar Archer. Proc. 8th Intern. Cong. Plant Pathol. 2:330.

Benson, D.M. and Parker, K.C. 2003. Suppression of Pythium root rot in poinsettia with swine waste and binucleate Rhizoctonia fungi. Phytopathology 93:S8.

Cubeta, M.A., Ceresini, P.C., Shew, H.D., Cody, B.R., Boerema, M.E., Lakshman, D.K., and Tavantzis, S.M. 2003. Occurrence of the M2 dsRNA in populations of Rhizoctonia solani AG-3 from potato and soil. Proc. Intern. Cong. Plant Pathol. Pp. 110.

Fichtner, E.J., Benson, D.M., Diab, H., and Shew, H.D. 2003. Suppression of Phytophthora parasitica in composted swine waste. Phytopathology 93:S26.

Gonzalez, D., Cubeta, M.A., and Vilgalys, R. 2003. Molecular systematics of Rhizoctonia phylogenetically integrates anamorphs into Ceratobasidiaceae. Proc. Intern. Cong. Plant Pathol. Pp. 29.

Howell, C.R. 2003. Pre-emergence damping-off of cotton seedlings. Workshop: Cotton Seedling Diseases. Proc. Beltwide Cotton Conf., National Cotton Assoc. America.

Howell, C.R., Puckhaver, L.S., and Stipanovic, R.D. 2003. Biocontrol of cotton pre-emergence seedling disease by metabolism of pathogen germination inducers. Phytopathology 93:S37.

Jia, Y., Singh, P., Cartwright, R.D., Lee, F.N., Rothrock, C.S., Eizenga, G.C., and Rutger, J.N. 2003. Characterization of isolate diversity in the rice sheath blight pathogen Rhizoctonia solani. Phytopathology 93:S40.

Keinath, A.P. and Smith, J.P. 2003. Effects of winter cover crops on populations of Rhizoctonia and Fusarium in bulk soil. Phytopathology 93:S43.