W147: Managing Plant Microbe Interactions in Soil to Promote Sustainable Agriculture

(Multistate Research Project)

Status: Inactive/Terminating

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SAES-422 Reports

Annual/Termination Reports:

[09/01/2002] [07/29/2003]

Date of Annual Report: 09/01/2002

Report Information

Annual Meeting Dates: 12/09/2001 - 12/10/2001
Period the Report Covers: 01/01/2001 - 12/01/2001

Participants

James Borneman (Chair), UC Riverside, Riverside, CA;
Jenifer H. McBeath (Secretary) University of Alaska, Fairbanks, AK;
George Abawi, Cornell University, Geneva, NY;
J. Ole Becker, UC Riverside, Riverside, CA;
Carol Lewis, University of Alaska Fairbanks, Fairbanks, AK;
John A. Menge, UC Riverside, Riverside, CA;
L. S. Pierson, University of Arizona, Tucson, AZ;
Peter Gay, University of Alaska Fairbanks, Fairbanks, AK;
Charity Gitschel, University of Alaska Fairbanks, Fairbanks, AK;
Margaret Ma, University of Alaska Fairbanks, Fairbanks, AK;
Takako Yokogi, University of Alaska Fairbanks, Fairbanks, AK;
Mingyuan Cheng, University of Alaska Fairbanks, Fairbanks, AK

Brief Summary of Minutes

Dr. Jenifer H. McBeath, the Local Arrangements Chair, welcomed the group to Fairbanks. Dr. Carol Lewis, the Interim Dean for the School of Agriculture and Land Resource Management at the University of Alaska Fairbanks (UAF) and a new W-147 committee member, gave a welcome address. She outlined important aspects of the virus-free seed potato program at UAF and gave a brief history of agriculture in interior Alaska.

Following greeting addresses, Dr. James Borneman, Chair, started the individual research progress report presentations. The Saturday morning session adjourned at noon, re-convened at 7 pm and was completed at 10:30 pm.

On Sunday, the business meeting was brought to order by Dr. Borneman at 9:30 am. Primary topics discussed were as follows:

1) Development of a regional website. Discussion was conducted related to developing a website to promote the western regional project. It was also mentioned that other regions including NC 125 and S267 have websites. The purpose of the website was to promote biological control as well as an outreach project for W-147. Ole Becker would look into this.

2) Recruitment of new members and encouragement of participation of current members. It is important to tap new members, especially from western states not currently represented. Potential members include Jeff Miller (University of Idaho) Carolee Bull and Frank Martin (USDA-ARS, Salinas CA), Tom Gordon (UC-Davis), Hank Williamson (Univ. of Illinois) and others. Drs. McBeath and Borneman will compile a list of contact people.

3) Project renewal and timing of submission. Renewal of W-147 is slated for 2003. It was agreed that by Feb 2002 current members would be contacted to contribute ideas. John Menge will look for the disc version of the last submitted proposal and will forward it to each member electronically. George Abawi will search the USDA website for a version of the last proposal submitted. Sandy Pierson will help compile the proposal. The draft will be brought to next years meeting for final comments and revisions before submission in April 2003. It was agreed that the next submitted version will be a revision rather than a rewrite. Promotion of soil health and soil quality will be a primary focus.

4) The next meeting will be December 7 and 8, 2002 in Riverside, CA. The UC Riverside group will organize the meeting.

Jenifer McBeath, this years secretary, will become Chairperson next year. Sandy Pierson will be next years Secretary. The meeting was adjourned.

Action Items/Assigned Responsibilities/Deadlines/Target Dates:

Jenifer McBeath will invite Jeff Miller (University of Idaho) to formally participate in the W147 project and attend the annual meetings.

Intensify efforts to recruit new members (all members)

Inquire about setting-up a W-147 web page on the University of California Riverside server (Ole Becker)

Time schedule and work distribution were developed for the renewal of W-147 in 2003.

Next Meeting Information:

Location and Date: Dec 6 and 7, 2002 in San Diego, California.

Responsible Individual(s): Ole Becker, James Borneman and John Menge

Non-Committee Member to be Invited: Jeff Miller (Idaho University), Linda Hanson (USDA-ARS), Chris Lawrence (Colorado State University).

Accomplishments

Objective 1: To identify and characterize plant microbe interactions that provide suppression of diseases caused by soilborne pathogens. Members of the W147 group are utilizing a diverse set of approaches to address the goals of identifying and characterizing important plant microbe interactions. Below is a summary of the progress made this past year. Conidia of Hirsutella rhossiliensis readily attach to the surface of nematodes (Figs. a, e). Subsequently, the nematode is invaded by the germinating conidia. It was established from transmission electron microscopy, SEM and LSC-Microscopy that attachment of H. rhossiliensis conidia is instantaneous upon contact, and that germination begins within 18 hours. The mucilaginous layer that surrounds the spores of H. rhossiliensis attaches the conidia to the nematode surface. In doing so, it flattens onto the nematode surface. In vitro, attached spores of H. rhossiliensis develop germ tubes, apparently in any direction, either in intimate contact with the surface of the nematode or into the surrounding environment. By 24 hours many of the spores have germinated and a few have penetrated the nematode cuticle. Infected nematodes often live and exhibit vigorous movements up to 40 hours after attachment of the conidia (and, approximately 20 hours after being invaded by the fungus). By 96 hours, the nematodes are fully occupied by hyphae of the pathogen. A survey of California avocado groves has been initiated to identify local groves which have soils which are suppressive to Phytophthora cinnamomi. Two criterions are used to identify a suppressive soil. It is a soil which degrades P. cinnamomi hyphae or chlamydospores, or one which has high populations of Phytophthora but the trees continue to thrive. Greenhouse tests with autoclaved soil indicated that autoclaving will destroy the suppressiveness of the Vanoni soil, indicating microorganisms are responsible for the suppressiveness. Other soils thought to be suppressive to P. cinnamomi did not appear to be suppressive in pot tests, indicating that drainage or some charactersitic of the field soil must be responsible for the suppressiveness. Twenty-four groves have been surveyed with four showing suppressiveness to Phytophthora. Individual trees in other groves also show suppressive characteristics. We have identified two microorganisms which are directly pathogenic to P. cinnamomi. Rozella sp. and Lytobacter mycophilus are known parasites of fungi and will attack and kill P. cinnamomi under laboratory conditions. Work is ongoing in the laboratory to test the biocontrol ability of these microorganisms. Epidemics of Phytophthora in individual groves have been studied. Populations of P. cinnamomi appear to decline precipitously immediately behind the leading edge of the epidemic in some groves. Biocontrol fungi are being isolated from this region. We postulate that microorganisms which fill a niche similar to that of the pathogen or that compete for chemical substrates important to the pathogen, such as root exudates, will be effective biological control agents. To identify such organisms, we have developed an in situ, culture-independent strategy to identify bacteria and fungi that rapidly grow in response to specified chemical substrates in environmental samples. To test this approach, we examined soils from a southern Californian avocado grove where a Phytophthora cinnamomi epidemic has produced three distinct zones. In Zone 1 where the pathogen epidemic has occurred, the trees are dying or dead but there is very little recoverable Phytophthora. Zone 2 is where the advancing margin of the Phytophthora epidemic is located and where the trees are relatively healthy but the roots are becoming infected and Phytophthora populations are high. In the region ahead of the advancing margin (Zone 3), the trees are healthy and the soil is conducive to avocado root rot. We postulate that in Zone 1, microorganisms are reducing the populations of P. cinnamomi and the soil has become suppressive. We used this new experimental approach to identify six bacteria whose population levels correlated with the differing levels of suppressiveness. These bacteria have varying similarities to several Bacillus species and Arthrobacter globiformis. We are currently in the process of testing these organisms in greenhouse trials. Trichoderma aureovirde, Trichoderma harzianum, Gliocladium virens and Hyphodontia alutacea, which were recovered behind the leading edges of Phytophthora epidemics, greatly damage Phytophthora chlamydospores. Other new potential biocontrol organisms, which have been isolated, include Pseudomonas alcaligenes and Erwinia cypripedii. Using the new species specific DNA probes, many potential biocontrol agents were identified from decomposing mats of P. cinnamomi hyphae. These organisms, which could not be cultured, included an unidentified protozoa-like organism, Trichosporon sp., which is a biocontrol agent of a corn disease, Tritirachium sp., which is closely related to parasites of insects, Arthrobotrys dactyloides, which is a parasite of nematodes, and Hypomyces chrysospermus, which is a parasite of many fungi. Attempts are now being made to isolate these fungi. The rain forest of Papua New Guinea is thought to be the center of origin for P. cinnamomi. We have visited Papua New Guinea and verified that P. cinnamomi appears to be endemic to the Highlands region of New Guinea. This area has a rainfall of several hundred inches per year. Flowing water is plentiful and would be ideal to disseminate P. cinnamomi. Nearly all crops are grown on mounds to prevent the roots from standing in water. Many endemic plants susceptible but tolerant to P. cinnamomi were located in the highlands region including Pandanus, Nothofagus, Acacia, Eucalyptus, Melanoleuca and Casuarina. Many avocados were found growing and surviving in standing water in which P. cinnamomi was present. Either the avocado varieties in New Guinea are resistant to P. cinnamomi or there are natural microorganisms in New Guinea which suppress the activities of P. cinnamomi. We have enlisted the aid of Bob Tombe, University of Goroka, Papua New Guinea, to cooperate with us and send samples every two months for the next two years. Locations and plant associates were identified during the visit. Preliminary data indicates that many of the samples from New Guinea are extremely suppressive to P. cinnamomi. Samples of soil were also brought back to the US from Northern Australia during the Australian Avocado Symposium. Seventeen out of 40 samples were highly suppressive to P. cinnamomi. Nearly all of the suppressive soil samples were from rainforest areas. Soil samples from arid regions or avocado groves were not suppressive to P. cinnamomi. Efforts are now being made to isolate biocontrol microorganisms from these Australian soils. One of the first steps in characterizing an ecosystem is to describe the organisms inhabiting it. For microbial studies, experimental limitations have hindered the ability to depict these diverse communities. To address these methodological deficiencies, we developed a new method termed oligonucleotide fingerprinting of ribosomal RNA genes (OFRG). This method permits the identification of arrayed ribosomal RNA genes (rDNA) through a series of hybridization experiments using small DNA probes. To demonstrate this strategy, we examined the bacteria inhabiting two agricultural soils possessing differing abilities to suppress the plant parasitic nematode Heterodera schachtii. Analysis of 1536 rDNA clones revealed 766 clusters grouped into 5 major taxa: Bacillus, Actinobacteria, Proteobacteria and two undefined assemblages. Soil specific populations were identified and then independently confirmed through sequence specific PCR of the original soil DNA. Near species level resolution was obtained by this analysis as it resolved bacterial clones with an average sequence identity of 96%. The pathogen suppressive soil was shown to contain greater species richness and diversity than the non-suppressive soil, when examined by Chao1 and Shannon analyses and by summing the branch lengths from UPGMA trees. A comparison of these OFRG results with those obtained from a denaturing gradient gel electrophoresis (DGGE) analysis of the same two soils demonstrated the significance of this methodological advance. OFRG provides a cost effective means to extensively analyze microbial communities and should have application in medicine, biotechnology and ecosystem studies. Accomplishments for Objectives 2 and 3 are not included due to limited space. The full report is available from the administrative advisor. Objective 2: To understand how biological and environmental factors regulate microbial populations and the expression of genes responsible for disease suppression. Objective 3: To develop and implement economic biological control systems to achieve sustainable agriculture.

Publications

ALASKA Cheng, M., Gay, P. A., and McBeath, J. H. 2000. Determination of chitinolytic activity in Trichoderma atroviride under differing environmental conditions. In Biocontrol in a New Millenium, Proceedings of the Third Joint National Biocontrol Conference, Estes Park Center, CO. D. M. Huber, ed. pp. 57-61. Gay, P. A., Cheng, M., and McBeath, J. H. 2001. Induction of proteins in Trichoderma atroviride in association with biological control of Botrytis cinerea, Phytophthora erythroseptica and Rhizoctonia solani. Phytopathology 91:S31 Gay, P. A. and McBeath, J. H. 2000. Development of an autofluorescent molecular marker system in Trichoderma atroviride. In Biocontrol in a New Millenium, Proceedings of the Third Joint National Biocontrol Conference, Estes Park Center, CO. D. M. Huber, ed. pp. 109-111. Gay, P. A. and Tuzun, S. 2000. Involvement of a novel peroxidase isozyme and lignification in hydathodes in resistance to black rot disease in cabbage. Canadian Journal of Botany 78:1144-1149. Gay, P. A. and Tuzun, S. 2000. Temporal and spatial assessment of defense responses in resistant and susceptible cabbage varieties during infection with Xanthomonas campestris pv. campestris. Physiological and Molecular Plant Pathology 57:201-210. McBeath, J. H. 2001. Biocontrol and growth promotion with cold tolerant Trichoderma. The IPM Practitioner 23: 1-6. McBeath, J. H. 2000. Effects of Trichoderma atroviride on snow mold of turfgrasses in interior Alaska. In Biocontrol in a New Millenium, Proceedings of the Third Joint National Biocontrol Conference, Estes Park Center, CO. D. M. Huber, ed. pp. 97-100. McBeath, J. H. and Kirk, W. W. 2000. Control of Seed-borne late blight on pre-cut potato seed with Trichoderma atroviride. In Biocontrol in a New Millenium, Proceedings of the Third Joint National Biocontrol Conference, Estes Park Center, CO. D. M. Huber, ed. pp. 88-96. McBeath, J. H., Mao, W., and Nguyen, C. 2000. Evaluation of in-furrow soil application of Trichoderma atroviride on the germination and growth of cotton seedlings. In Biocontrol in a New Millenium, Proceedings of the Third Joint National Biocontrol Conference, Estes Park Center, CO. D. M. Huber, ed. pp. 84-87. McBeath, J. H., Parent, T., and Kreuger, R. 2000. Effects of Trichoderma atroviride on the root system of Panax quinquefolius. In Biocontrol in a New Millenium, Proceedings of the Third Joint National Biocontrol Conference, Estes Park Center, CO. D. M. Huber, ed. pp. 101-109. McBeath, J. H., Gay, P. A., and Yokogi, T. 2001. Biological control of pink rot by Trichoderma atroviride. Phytopathology 91:S59. ARIZONA Pierson, L.S. III. 2000. Expanding the club: engineering plants to talk to bacteria. Trends in Plant Sciences 5:89-91. Pierson, L.S. III. 2000. Bacterial signaling: Identification of N-acyl-homoserine lactone-producing bacteria. The Plant Health Instructor DOI:10.1094/PHI-I-2000-1214-01. Pierson, L.S. III and Ishimaru, C.A. 2000. Genomics of plant-associated bacteria: a glimpse of the future that has become reality. American Phytopathological Society, Feature story, August 2000 (http://www.apsnet.org/online/feature/genomics/Top.html). Pierson, L.S. III. 2000. Disease decline. In: Encyclopedia of Plant Pathology, Maloy, O.C., and Murray, T.D., eds. John Wiley and Sons, New York, NY. Zhang, Z, and Pierson, L.S. III. 2001. A second quorum sensing system regulates cell surface properties but not phenazine antibiotic production in Pseudomonas aureofaciens. Applied Environ. Microbiol. 67:4305-4315. CALIFORNIA Becker, J.O. 2000. Evaluation of Nematode-suppressive soil. California Ornamental Research Federation News 4 (2), 14-15 (abstract). Borneman, J. and R.J. Hartin. 2000. PCR primers that amplify fungal rRNA genes from environmental samples. Applied and Environmental Microbiology 66:4356-4360. Borneman, J., M. Chrobak, G. Della Vedova, A. Figueroa, and T. Jiang. 2001. Probe selection algorithms with applications in the analysis of microbial communities. Bioinformatics 17(Suppl. 1):S39-S48. Downer, A. J., J.A. Menge and E. Pond. 2001. Association of cellulytic enzyme activities in Eucalyptus mulches with biological control of Phytophthora cinnamomi. Phytopathology 91: 847-855. Downer, A. J., J. A. Menge and E. Pond. 2001. Effects of cellulytic enzymes on Phytophthora cinnamomi. Phytopathology 91: 839-846. Gao, X., and J.O. Becker 2001. Biological soil suppression affects both sexes of Heterodera schachtii. Phytopathology 91:S134 (abstract). Kim, D.H., Stanghellini, M.E., Waugh, M.M., and Mayberry, K.S. 2001. Vine-decline of melons caused by Monosporascus cannonballus: II. Postplant disease management strategies. Phytopathology 91:S48. Menge, J. A. 2001. Mulches. P. 650-651. In Encyclopedia of plant pathology. Maloy, O. C. and T. D. Murray. (Eds.). John Wiley and Sons, Inc., New York. Menge, J. A. and L. J. Marais. 2001. Soil environmental factors and their relationship to avocado root rot.. Subtropical Fruit News 8: 11-14. Radewald, K.C., Stanghellini, M.E., Kim, D.H., Waugh, M.M., Mayberry, K.S., and Turini, T. 2001. Vine-decline of melons caused by Monosporascus cannonballus: III. Postharvest disease management strategies. Phytopathology 91:S74. Stanghellini, M.E., Kim, D.H., Waugh, M.M., Radewald, K.C., Sims, J.J., Ohr, H.D., Mayberry, K.S., Turini, T., and McCaslin, M.A. 2001. Vine-decline of melons caused by Monosporascus cannonballus: I. Preplant disease management strategies. Phytopathology 91:S84. Steddom, K. and J. A. Menge. 2001. Evaluation of continuous application technology for delivery of the biocontrol agent Pseudomonas putida 06909-rif/nal. Plant Dis. 85: 387-392. Steddom, K., J. A. Menge and J. Borneman. 2001. The effect of continuous application of a biocontrol bacterium on root rot of citrus and the soil community. Proceedings Biocontrol in a New Millenium: Building for the Future on Past Experience. Joint Reg. Biocontrol Conf. Estes Park, Colorado. Yang, C.-H., D. E. Crowley and J. A. Menge. 2001. 16S rDNA fingerprinting of rhizosphere bacterial communities associated with healthy and Phytophthora infected avocado roots. FEMS Microbial Ecology 35: 129-136. Yin, B., J. A. Menge, E. Pond and J. Borneman. 2001. An in situ, culture independent approach to examine substrate competition of soil miicroorganisms for discovery of biocontrol agents. Phytopathology 91:S98. (Abst.) Waugh, M., D.H. Kim and Stanghellini, M.E. 2000. Scanning electron microscopy of germinated ascospores of Monosporascus cannoballis in the rhizosphere of cantaloupe roots. Mycological Research 105:745-748. MONTANA Callan, N.W. and Mathre, D.E. 2000. Bio-Priming Seed Treatment. Encyclopedia of Plant Pathology, Wiley and Sons. NEW YORK Abawi, G. S., and J. W. Ludwig. 2001. Root rot control for peas and beans. NYS Vegetable Conference, pp. 38 40, Cornell Cooperative Extension, Ithaca, NY. Abawi, G. S., and J. W. Ludwig. 2001. Update on beet diseases and their management. NYS Vegetable Conference, pp. 122-126, Cornell Cooperative Extension, Ithaca, NY. Abawi, G. S., Ludwig, J. W. and Widmer, T. L. 2000. Root-knot nematode and control options for carrots. NYS Veg. Conf., pp 53-56, Cornell Cooperative Extension, Ithaca, NY. Padgham, J.L., J.M. Duxbury, G.S. Abawi, J.G. Lauren, and S. Parvin Banu. Diagnosing poor crop performance using solar-oven soil pasteurization. Presented at the 2000 Agronomy Society of America meetings in Minneapolis, MN. Nov. 7-11, 2000. Westphal, A., and J.O. Becker. 2001. Components of soil suppressiveness against Heterodera schachtii. Soil Biology & Biochemistry 33:9-16. Westphal, A., and J.O. Becker 2001. Impact of soil suppressiveness on various population densities of Heterodera schachtii. Annals of Applied Biology 138:371-376. Widmer, T. L., and G. S. Abawi. 2000. Mechanism of suppression of the northern root-knot nematode by sudangrass incorporated as a green manure. Plant Disease 84: 562 568. Widmer, T. L., and G. S. Abawi. 2001. Relation between levels of cyanide of sudangrass hybrids incorporated into soil and suppression of Meloidogyne hapla. J. of Nematology 33: (Accepted). Widmer, T. L., N. A. Mitkowski, and G. S. Abawi. 2001. Impact of organic matter management on plant-parasitic nematodes, their damage to host crops, and soil health: A review. J. of Nematology 33: (submitted, symposium paper). Yin, B., J. O. Becker, J.A. Menge, E. Pond, and J. Borneman 2000. In situ analysis of substrate utilization for identification of potential biological control micro-organisms. In: Proceedings Biocontrol in a new millennium: Building for the future on past experience (ed. D.M. Huber), Estes Park Center, Colorado, 27-30.

Impact Statements

Members of this research group seek to find environmentally friendly solutions for management of plant pests. Towards this goal, we are examining both basic and applied research areas. Below is a summary of the usefulness of our findings.
Trichoderma atroviride is a versatile, aggressive hyperparasite that can parasitize a wide spectrum of pathogenic fungi. It has also been found to inhibit Phytophthora erythroseptica under laboratory conditions. Plant Helperb, a commercial product containing T. atroviride spore suspension @ 106cfu/g, has been found to be effective in the control of damping off of cotton, rusty root, and early season disease on ginseng under commercial field conditions and snow mold diseases on golf courses.
T. atroviride produces a coordinated biochemical response in the presence of different plant pathogens. Characterization of these enzymatic responses has revealed an induced and constitutive response by T. atroviride. Timing of response seems to be important as well. Production of chitinases seemed to play a significant role in the hyperparasitism of T. atroviride in the suppression of diseases.
Further studies are necessary to determine specific roles of each enzymatic group and specific isozyme involvement in biocontrol activity. T. atroviride and T. virens (GL3) are compatible in disease control.
The research at University of Arizona is a continuation of a long-term effort at elucidating the molecular mechanisms involved in bacterial sensing. These regulatory systems determine the patterns of competitive gene expression and therefore have a direct effect on the success or failure of a biological control agent in the field.
One exciting aspect of the work with the negative signaling strains is that they may result in the identification of genes that encode signals that interfere with AHL systems. Thus, these genes may be of usefulness in engineering plants to produce signals to knockout pathogenic bacteria that utilize AHL regulation for virulence gene expression.
The study on the parasitism of root-knot nematode larvae by Hirsutella rhossiliensis investigates the basic cell biology of fungal-nematode interactions. The findings are useful in that they set time lines for the infection process as well as for the mode on fungal ingress into the nematode body.
Knowledge on the effects of cultural practices and production inputs on the incidence and damage of soilborne pathogens is critical in the development of control options of root diseases that are compatible with sustainable management of soil health and productivity. It is also important to validate the efficacy of available commercial preparations in different soils and production systems, as their activities might well be different.
It appears that Phytophthora cinnamomi may have originated in or near Papua New Guinea. Preliminary evidence from soil samples gathered in New Guinea indicate that potent biocontrol agents against P. cinnamomi may be found in New Guinea.
Mulch is an effective inhibitor of Phytophthora root rot of avocado. Cellulose in the mulch is degraded by saprophytic fungi which produce abundant cellulase and laminarinase. Application of mulch to avocado trees growing in P. cinnamomi infested soil provides a viable control method for the devestating root rot of avocado caused by P. cinnamomi.
We believe that the EcoSoils field fermenter effectively produced and distributed bacterial biocontrol organisms. Continuous application of biocontrol bacteria have tremendous promise and our bacterial biocontrol agent applied in this way gave increased populations in the soil over the growing season. It appears that the EcoSoils field fermenter is an effective delivery method for biocontrol agents.
The beet cyst nematode is one of the most important pests of beets and crucifers in the United States and Europe. Understanding the basis of nematode control in suppressive sites may lead to future exploitation for crop management systems. This study has revealed the biological nature of the suppressiveness, its potential to be transferred by small amounts of soil or cysts into fumigated, conducive sites, and its activity against both sexes of H. schachtii.
Biological seed treatment with Pseudomonas aureofaciens AB254 protected supersweet corn seed from Pythium seed decay at a level equivalent to metalaxyl treatment, but other commercially available biological seed treatments did not increase seedling stand. Our results demonstrate the variability in effectiveness of biological seed protection treatments.
Results indicate that management of cantaloupe vine decline can be accomplished by inhibiting pathogen reproduction on melon roots left in the field following crop termination.

Date of Annual Report: 07/29/2003

Report Information

Annual Meeting Dates: 12/14/2002 - 12/15/2002
Period the Report Covers: 01/01/2002 - 12/01/2002

Participants

Alaska: J.H. McBeath;

Arizona: L. S. Pierson III;

California: J.O. Becker,
J. Borneman,
J. A. Menge,
M. Stanghellini;

Montana: N. W. Callan;

Washington: T. C. Paulitz;

Administrative Advisor: D. Cooksey

Brief Summary of Minutes

Minutes Attachment:

Attachment

Accomplishments

Objective 1: A survey of California avocado groves has been initiated to identify local groves which have soils which are suppressive to Phytophthora cinnamomi. Two criterions are used to identify a suppressive soil. It is a soil which degrades P. cinnamomi hyphae or chlamydospores, or one which has high populations of Phytophthora but the trees continue to thrive. Greenhouse tests with autoclaved soil indicated that autoclaving will destroy the suppressiveness of the Vanoni soil, indicating microorganisms are responsible for the suppressiveness. Other soils thought to be suppressive to P. cinnamomi did not appear to be suppressive in pot tests, indicating that drainage or some charactersitic of the field soil must be responsible for the suppressiveness. Twenty-four groves have been surveyed with four showing suppressiveness to Phytophthora. Individual trees in other groves also show suppressive characteristics. We have identified two microorganisms which are directly pathogenic to P. cinnamomi. The goal of studies by Borneman (UCR) was to identify bacteria and fungi involved in soil suppressiveness against the plant-parasitic nematode Heterodera schachtii. Since H. schachtii cysts isolated from the suppressive soil can transfer this beneficial property to nonsuppressive soils, analysis of the cyst-associated microorganisms should lead to the identification of the causal organisms. Our experimental approach was to identify bacterial and fungal rDNA associated with H. schachtii cysts obtained from soil mixtures with varying levels of suppressiveness. We hypothesized that we could identify microorganisms involved in the suppressiveness by correlating population shifts with differing levels of suppressiveness. Bacterial rDNA associated with H. schachtii cysts were identified by Borneman using a culture-independent method termed oligonucleotide fingerprinting of ribosomal RNA genes (OFRG). Five major taxonomic groups were identified: Actinobacteria, Cytophaga-Flexibacter-Bacteroides, a-Proteobacteria, b-Proteobacteria, and c-Proteobacteria. Three bacterial rDNA groups contained more clones from the highly suppressive soil treatments than the less suppressive treatments, indicating a potential involvement in the H. schachtii suppressiveness. When these three groups were examined with specific PCR analyses performed on H. schachtii cysts that developed in soils treated with three biocidal compounds, only one bacterial rDNA group with moderate to high sequence identity to rDNA from several Rhizobium species and uncultured a-Proteobacterial clones was consistently associated with the highly suppressive treatments. Borneman also identified fungi through an rDNA analysis termed oligonucleotide fingerprinting of ribosomal RNA genes (OFRG). Cysts obtained from soil mixtures consisting of 10% and 100% suppressive soil predominantly contained fungal rDNA with high sequence identity to Dactylella oviparasitica. The dominant fungal rDNA in the cysts isolated from the soil mixtures comprised of 0.1% and 1% suppressive soil had high sequence identity to Fusarium oxysporum). Pink rot of potato (caused by Phytophthora erythroseptica), potato late blight (caused by Phytophthora infestans), gray mold (caused by Botrytis cinerea), and damping-off (caused by Rhizoctonia solani) are all economically destructive diseases. For many years, control of these diseases relied primarily on the use of chemical fungicides. Increasing awareness of the development of fungicide-resistant strains of pathogens, chemical residues in the food chain and the adverse effects of pesticides on human health are having profound impacts on the management of plant diseases. With a diminishing number of means to control disease, growers are seeking alternatives that are both safe and environmentally benign. T. atroviride is a versatile, aggressive hyperparasite that can parasitize a wide spectrum of pathogenic fungi, including Phytophthora erythroseptica, Rhizoctonia solani, Phytophthora infestans and Botrytis cinerea. T. atroviride has a temperature range of 4 to 33 0C and prefers high humidity. Results of compatibility studies by McBeath (AK) show that T. atroviride growth is not adversely affected by common fungicides and/or herbicides, even at field applicable levels. However, performances of T. atroviride are limited by high soil temperatures and sub-optimal dosages. A coordinated biochemical response has been observed in T. atroviride during biocontrol of plant pathogenic fungi. Production of chitinases, chitosanases, and glucanases, seemed to play a significant role in hyperparasitism involved in the suppression of diseases. In Pacific Northwest cereal cropping systems, little is known about the populations and virulence of Pythium and Rhizoctonia spp. Pythium spp. that cause root rot and significant yield reductions in wheat and barley. ARS scientists at Pullman, Washington sequenced the rDNA of the ITS 1 region of Pythium isolates from an extensive collection in eastern Washington. In addition to identifying 11 known species, a totally new species was discovered, named Pythium abappressorium sp. nov. Paulitz et Mazzola, which is widespread and pathogenic. Knowledge of the diversity of Pythium spp. is critical for plant breeders, who will utilize these isolates to screen for resistant wheat and barley germplasm. Rhizoctonia oryzae along, with R. solani AG-8, causes the disease of wheat and barley know as Rhizoctonia root rot, but Rhizoctonia oryzae was not considered to be a problem in previous surveys. ARS scientists at Pullman, Washington conducted an extensive survey of wheat and barley fields. They showed that Rhizoctonia oryzae is widespread on cereals in eastern Washington, it causes damping-off and stunting of barley and wheat, it attacks peas and other broadleaf rotation crops, some isolates are more virulent than R. solani AG-8, variation in virulence exists among isolates, and there is a significant cultivar X isolate interaction. Rhizoctonia oryzae causes chronic root rot and stunting of wheat and barley. ARS scientists at Pullman, Washington in conjunction with a scientist at Washington State University adapted spatial generalized linear mixed models to detect spatial correlation and develop disease maps. Using GPS-located sites on a 90-acre farm over two years, an aggregated, overdispersed distribution of the pathogen and the disease was detected. These tools will have applications for disease mapping in precision agriculture. For the past few years, Ole Becker and cooperators have investigated a beet cyst nematode-suppressive soil at a field station near UCR. After the initial nematode infestation of this field 9E in the 1970s and a high population build-up caused by frequent cropping to susceptible hosts, the population declined and has remained fairly constant at a low level. Although previous research had investigated the progression of the suppressiveness during a cropping season, several data gaps required a more in-depth analysis of the beet cyst nematode population dynamics. Consequently, the population development of Heterodera schachtii on Swiss chard (Beta vulgaris L. subsp. cicla (L) cv. Large White Ribbed) in suppressive and conducive soil was monitored for two generations in greenhouse trials. Staining of the second-stage juveniles in the roots 75 degree days (DD) after soil infestation with the pest revealed no significant differences between the number of nematodes grown in suppressive soil and conducive soil. After the first nematode generation, the beet cyst nematode populations in the suppressive and the conducive soil were not significantly different as indicated by the number of males and cysts as well as by the number of eggs and second-stage juveniles within those cysts. However, after two beet cyst nematode generations, the number of all monitored life stages were significantly lower in the suppressive soil than in the conducive soil. Objective 2: Potato is an economically important crop to Alaska as well as in the US. Identification of molecular markers associated with disease resistance and adaptability to extreme Northern climates may help molecular breeders in the development of cultivars more suitable to these conditions. Osmotins represent a multigene family that has been implicated in disease resistance and cold tolerance. Isolation and characterization of osmotin genes may help in understanding host-pathogen interactions as well as in the development of enhanced biological control agents for the control of plant diseases. Research by Leland Pierson (AZ) involving further characterization of the molecular mechanisms responsible for phenazine gene regulation in Pseudomonas aureofaciens 30-84 is providing additional evidence for how a bacterium senses its environment and response to it by altering patterns of gene expression. In addition to the molecular analyses, ecological approaches are being used to quantify the role of cross-communication among rhizobacteria on phenazine gene expression in situ. Wheat genes that govern disease suppression can be identified and mapped using classical genetics. To examine genetic variation in root colonization, ARS scientists at Pullman, Washington compared wheat root colonization by the aggressive P. fluorescens strain Q8r1-96 with that of the weaker colonizer Q2-87 for 28 Pacific Northwest (PNW) cultivars. Five cultivars supported higher root populations of Q8r1-96, and although PNW cultivars differed in root morphology and growth, no single factor was correlated with preference for Q8r1-96. Genetic variation in root traits of PNW wheat cultivars can be exploited for genetic, biochemical and molecular approaches to host-mediated disease suppression. Strains of Pseudomonas fluorescens producing the antifungal metabolite 2,4-diacetylphloroglucinol (DAPG) are important biocontrol agents against dampingoff, root rot and wilt diseases, and are responsible for the natural suppressiveness of some soils against root diseases. ARS scientists at Pullman, Washington used DNA fingerprinting methods to identify genetic groups or subspecies within populations of DAPG producers. Seventeen distinct genetic groups were identified, and certain groups were shown to have superior abilities to colonize the roots of wheat or pea or both crops, and control root diseases. Objective 3: Research by Mike Stanghellini (UCR) is directed at developing a disease management strategy for the control of vine-decline of melons caused by the root-infecting fungus Monosporascus cannonballus. Ascospores of M. cannonballus, the primary inoculum for root infection and survival of the fungus in soil, play a major role in vine decline. Our previous studies indicate that the development of an effective disease management strategy in known pathogen-infested fields is dependent upon (i) reducing the amount of primary inoculum in soil with a preplant soil fumigant and then (ii) maintaining low population densities by inhibiting pathogen reproduction on melon roots left in the field after harvest. Strategies identified in 2000 for inhibiting pathogen reproduction included (i) the application of metam sodium or (ii) soil cultivation to expose and dry the roots immediately after the final harvest or collapse of the vines. These postharvest strategies, if successful, would reduce the necessity for yearly applications of the more costly preplant soil fumigants. Based on this, we initiated a three-year study to evaluate the efficacy of the above strategy for management of vine-decline of melons. The Spring 2002 crop constituted the second of three consecutive spring crops (subject only to postharvest treatments) following a single application of a preplant soil fumigant in the Spring of 2001.

Publications

ALASKA Gay, P. A., and McBeath, J. H. 2002. Cold tolerant Trichoderma atroviride, biotype CHS 861, ATCC 74015, 42 kD endochitinase gene, partial sequence. Genbank accession number AY162405, submitted. Gay, P. A., and McBeath, J. H. 2002. Cold tolerant Trichoderma atroviride, biotype 453, ATCC74016, 42 kD endochitinase gene, partial sequence. Genbank accession numberAY162406, submitted. Gay, P. A., and McBeath, J. H. 2002. Cold tolerant Trichoderma atroviride, biotype 901C, ATCC 74017, 42 kD endochitinase gene, partial sequence. Genbank accession number AY162407, submitted. Gay, P. A., and McBeath, J. H. 2002. Cold tolerant Trichoderma atroviride, biotype 603, ATCC 74018, 42 kD endochitinase gene, partial sequence. Genbank accession number AY162408, submitted. McBeath, J.H. 2002. Snow Mold-Plant-Antagonist Interactions: Survival of the Fittest under the Snow. APSnet Feature Article, March Issue. McBeath, J.H. 2002. Snow Mold-Plant-Antagonist Interactions: Survival of the Fittest under the Snow. Plant Health Instructor. DOI:10.1094/PHI-1-2002-1010-01. McBeath, J.H. Snow Mold: Nemesis of Turfgrasses. Golf Course Management. (in press) ARIZONA Chancey, S.T., Wood, D.W., Pierson, E.A. and L.S. Pierson III. 2002. Survival of GacS/GacA mutants of the biological control bacterium Pseudomonas aureofaciens 30-84 in the wheat rhizosphere. Appl. Environ. Microbiol. 68: 3308-3314. Loh, J., Pierson, E.A., Pierson, L.S. III, Stacy, G., and Chatterjee, A. 2002. Quorum sensing in plant-associated bacteria. Curr. Opin. Plant Biol. 5:1-5. Zhang, Z. and Pierson, L.S. III. 2001. A second quorum sensing system regulates cell surface properties but not phenazine antibiotic production in Pseudomonas aureofaciens. Appl. Environ. Microbiol. 67: 4305-4315. CALIFORNIA Dirac, M. and J.A. Menge. 2002. High temperatures are not reponsible for lack of infection of citrus roots by Phytophthora citrophthora during the summer, but supressive microorganisms may inhibit infection by P. citrophthora. Plant and Soil 241:243-249. Dirac, M. and J.A. Menge. 2002. Comparison of seasonal infection of citrus roots by Phytophthora citrophthora and Phytophthora nicotianae. Plant Disease 86: (In Press). Marais, L.J., J.A. Menge, G.S. Bender and B. Faber. 2002. Phytophthora root rot. AvoResearch, California Avocado Commission. 4p. Marais, L. J, J.A. Menge, G.S. Bender and B. Faber. . 2002. Avocado stem canker or collar rot. AvoResearch, California Avocado Commission. 4p. Menge, J.A. A.J. Downer, K. Steddom and J. Borneman. 2002. Biocontrol of Phytophthora cinnamomi. Proceedings Calif. Conf. Biological Control. Aug15-16. Davis, CA. Menge, J.A. and R. Ploetz. 2002. Diseases of avocado. In Diseases of Tropical and Subtropical Fruit Crops. R. Ploetz (ed.) CABI Publishing, Oxford. 50 pp. (In Press) Menge, J.A. and L.J. Marais. 2002. Strategies to control Phytophthora cinnamomi root rot of avocado. Subtropical Fruit News 10: (In Press). Steddom, K, O. Becker and J.A. Menge. 2002. Repetitive applications of the biocontrol agent Pseudomonas putida 06909-rif/nal and effects on populations of Phytophthora parasitica in citrus orchards. Phytopathology 92:850-856. Steddom, K., J.A. Menge, D. Crowley and J. Borneman. 2002. Effect of repetitive application of the biocontrol bacterium Pseudomonas putida 06909-rif/nal on citrus soil microbial communities. Phytopathology 92:857-862. Yang, C.-H., D. E. Crowley and J. A. Menge. 2001. 16S rDNA fingerprinting of rhizosphere bacterial communities associated with healthy and Phytophthora infected avocado roots. FEMS Microbial Ecology 35: 129-136. Yin, B, Valinsky, L., Gao, X., Becker, J.O., and J. Borneman. Bacterial rDNA associated with soil suppressiveness against the plant-parasitic nematode Heterodera schachtii. Appl. Environ. Microbiol. (accepted with revisions). Valinsky, L., Della Vedova, G., Jiang, T., and Borneman, J. 2002. Oligonucleotide fingerprinting of ribosomal RNA genes for analysis of fungal community composition. Appl. Environ. Microbiol. 68:5999-6004. Waugh, M.M., D.H. Kim, D.M. Ferrin, and M.E. Stanghellini. 2003. Reproductive potential of Monosporascus cannonballus. Plant Dis. 87: 45-50. Valinsky, L., Della Vedova, G., Scupham, A. J., Alvey, S., Figueroa, A., Yin, B., Hartin, J., Chrobak, M., Crowley, D. E., Jiang, T., and Borneman, J. 2002. Analysis of bacterial community composition by oligonucleotide fingerprinting of rRNA genes. Appl. Environ. Microbiol. 68:3243-3250. WASHINGTON Cook, R.J., Weller, D.M., El-Banna, A.Y., Vakoch, D., Zhang, H. 2002. Yield responses of direct-seeded wheat of rhizobacteria and fungicide seed treatments. Plant Disease. 86: 780-784. Landa, B.B., de Werd, H.A.E., McSpadden Gardener, B.B., Weller, D.M. 2002. Comparison of three methods for monitoring populations of different genotypes of 2,4-diacetylphorlglucinol-producing Pseudomonas fluorescens in the rhizosphere. Phytopathology. 92: 129-137. Landa, B.B., Mavrodi, O.V., Raaijmakers, J.M., McSpadden Gardener, B.B., Thomashow, L.S., Weller, D.M. 2002. Differential ability of genotypes of 2,4-diacetylphloroglucinol-producing Pseudomonas fluorescens strains to colonize the roots of pea plants. Applied and Environmental Microbiology. 2002. 68: 3226-3237. Mavrodi, D.V., Bonsall, R.F., Delaney, S.M., Soule, M.J., Phillips, G., Thomashow, L.S. Functional analysis of genes for biosynthesis of pyocyanin and phenazine-1-carboxamide from Pseudomonas aeruginosa PAO1. 2001. Journal of Bacteriology. 183: 6454-6465. Mavrodi, O.V., Mavrodi, D.V., Weller, D.M., Thomashow, L.S. 2002. Genetic basis for the unique root-colonizing activity of Pseudomonas fluorescens. Phytopathology. 92(Suppl.). p. S53. (abstract). McSpadden Gardener, B.B., Weller, D.M. 2001. Changes in populations of rhizosphere bacteria associated with take-all disease of wheat. Applied and Environmental Microbiology.67: 4414-4425. McSpadden Gardener, B.B., Weller, D.M. 2002. Specificity, population dynamics, and biocontrol of take-all by DAPG-producing Pseudomonas spp. in a long-term wheat field. Phytopathology. 92(Suppl.). Abstract p. S54. Okubara, P.A., Landa, B.B., Madsen, B., Kornoely, J.P. 2002.Wheat cultivar-dependent root architecture and root colonization by P. fluorescens Q8r1. Phytopathology 92(Suppl.). p. S61. Okubara, P.A., Madsen, B., Landa, B.B. 2002. Is root colonization by P. fluorescens Q8r1 cultivar-dependent in wheat? Plant Biology 2002, American Society of Plant Biologists. 2002. p. 136. Abstract number 589. Paulitz, T. C. 2002. Biological control of plant pathogens (fungi). D. Pimentel, editor. Marcel Dekker, NY. Encyclopedia of Pest Management. p. 64-67. Paulitz, T. C. 2002. Biological webs of plant disease. New Phytologist.154: 272-773. Book Review. Paulitz, T. C. 2002. Impacts and management of soil acidity under direct seed systems: Effects on soilborne crop pathogens. Proceedings of the Northwest Direct Seed Cropping Systems Conference, Spokane WA. p. 41-46. Paulitz, T. C. First report of Rhizoctonia oryzae on pea. 2002. Plant Disease. 2002. 86: 442. Paulitz, T. C. Pseudomonas aureofaciens strain 63-28: A new weapon in the biocontrol arsenal. Proceedings of the International Symposium on Biological Control of Plant Diseases for the New Century- Mode of Action and Application Technology, Taichung, Taiwan. 2002. p. 97-106. Paulitz, T. C. 2002. New insights into the make-up and management of soilborne crop pathogens under direct seeding: Rhizoctonia. Proceedings of the Northwest Direct Seed Cropping Systems Conference, Spokane WA. p. 131-139. Paulitz, T. C., Adams, K. and Mazzola, M. 2002. Pythium abappressorium- a new species from eastern Washington. Mycologia: in press. Paulitz, T. C., Smiley, R. and Cook, R. J. 2002. Insights into the prevalence and management of soilborne cereal pathogens under direct seeding in the Pacific Northwest, U.S.A. Can. J. Plant Pathol. in press. Paulitz, T. C., Smith, J. and Kidwell, K. 2002. Virulence of Rhizoctonia oryzae on wheat and barley cultivars from the Pacific Northwest. Plant Disease: 87: 51-55. Paulitz, T.C., Adams, K., Mazzola, M., Livesque, C.A. 2002. A new species of Pythium from apple and wheat in eastern Washington. Phytopathology 92(Suppl.). Abstract p. S64. Schroeder, K.L., Paulitz, T.C. 2002. Development of Rhizoctonia root rot of barley in soils from conventional and no-till fields. Phytopathology 92(Suppl.). Abstract p. S74. Smiley, R., Cook, R. J. and Paulitz, T. 2002. Seed treatments for small grain cereals. Oregon State University Extension Publication EM 8797. Smiley, R., Cook, R. J. and Paulitz, T. 2002. Controlling root and crown diseases of small grain cereals. Oregon State University Extension Publication EM 8799. Weller, D.M., Raaijmakers, J.M., McSpadden Gardener, B.B., Thomashow, L.S. 2002. Microbial populations responsible for specific suppressiveness to plant pathogens. Annual Review of Phytopathology. 40: 309-348.

Impact Statements

It appears that the EcoSoils field fermentor is an effective delivery method for biocontrol agents.
The studies by Borneman (UCR) have led to the identification several bacteria and fungi that positively correlated with suppressiveness against the plant-parasitic nematode Heterodera schachtii., The experimental design utilized in these studies will provide a new investigative approach for soilborne biological control research.
Information on optimal spore concentrations and stickers for seed treatments will benefit the commercialization of Trichoderma atroviride and other biocontrol agents.
The findings on pathogen diversity conducted by ARS scientists at Pullman, Washington, will be useful for plant breeders who are selecting for resistant or tolerant germplasm.
Research by Pierson (AZ) is a continuation of a long-term effort at elucidating the molecular mechanisms involved in bacterial sensing and its effects on bacterial communication that influences gene expression patterns and therefore community structure. Studies on the occurrence of spontaneous gasA two component mutants in rhizosphere populations indicates that global regulatory mutations may exist as a normal component of a rhizosphere population.