Annual/Termination Reports:

[01/09/2007] [01/07/2008] [02/16/2009] [11/14/2009] [12/20/2010] [02/06/2012] [07/10/2013]

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Brief Summary of Minutes

Accomplishments

Publications

Impact Statements

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Participants

Participants:

Benson, Mike (mike_benson@ncsu.edu)  North Carolina State University
Canaday, Craig (ccanaday@utk.edu)  University of Tennessee
Cubeta, Marc (marc_cubeta@ncsu.edu) - North Carolina State University
Dick, Warren (dick.5@osu.edu) - Ohio State University
Elliott, Monica (melliott@ufl.edu)  University of Florida
Graham, Peter (graha019@umn.edu)  University of Minnesota
Jimenez Gasco, Maria (jimenez-gasco@psu.edu) - Pennsylvania State University
Keinath, Tony (tknth@clemson.edu)  Clemson University
Loynachan, Thomas (teloynac@iastate.edu) Iowa State University
Ownley, Bonnie (bownley@utk.edu)  University of Tennessee
Padgett, Boyd (bpadgett@agcenter.lsu.edu) - Louisiana State University
Rothrock, Craig (Rothrock@comp.uark.edu)  University of Arkansas
Westphal, Andreas (Westphal@purdue.edu)  Purdue University
Williams, Mark (markwill@uga.edu)  University of Georgia

Brief Summary of Minutes

Accomplishments

Accomplishments:<br /> <br /> Objective 1. Examine commercial and non-commercial biocontrol agents for use as seed treatments, in-furrow treatments or as potting mix amendments.<br /> " Cooperative Broccoli Field Trial Project  (Canaday, Keinath, Ownley, Rothrock, Seebold). Canaday - Delays in seed germination were observed with bioactive Monarda herbage incorporated into potting soil, however, differences in growth were not observed at 41 days after seeding. Keinath  After transplantation to field soil, and artificial inoculation with Rhizoctonia solani, Rhizoctonia wirestem was greatest with the Monarda treatment. <br /> " Benson  Binucleate Rhizoctonia (BNR) isolates BNR621 and P023, and Trichoderma hamatum isolate 382 induced resistance to the foliar fungal pathogen Botrytis cinerea in geranium. <br /> " Canaday  The combination of BioYield (plant growth-promoting rhizobacteria; PGPR) and fungicide treatments with and without application of Actigard (induces systemic acquired resistance in plants) had no effect on the incidence of timber rot (Sclerotinia sclerotiorum) and buckeye rot (Phytophthora nicotianae) affecting four tomato cultivars (Mountain Fresh Plus, Nico, Amelia, Red Defender).<br /> " Canaday  The incidence of Tomato Spotted Wilt Virus was greater in Mountain Fresh Plus tomato than in Nico, Amelia, and Red Defender; however, fungal foliar diseases (early blight and Septoria leaf spot) were lowest in Mountain Fresh Plus.<br /> " Canaday  The percentage of healthy soybean plants was greatest with a seed treatment combination of the PGPR strain Bacillus subtilis MBI 600 and the fungicides mefenoxam and fludioxinil.<br /> " Canaday  The PGPR product BioYield Flowable increased head weight of broccoli cultivars, Arcadia and Packman, but had not effect on Premier Crop. <br /> " Keinath - A non-pathogenic (binucleate) Rhizoctonia isolate and BioYield Flowable significantly increased the percentage of healthy broccoli plants and reduced the number of plants with wirestem 30 days after transplanting.<br /> " Ownley  Beauveria bassiana 11-98 endophytically colonized tomato and cotton following seed treatment. The extent of colonization was dependent on seed treatment rate and host plant; i.e. the fungus was found in roots, stems and leaves of tomato but primarily in above ground tissues of cotton.<br /> " Ownley - New ITS primers were developed for detection of B. bassiana 11-98 DNA within cotton tissues without amplification of ITS sequences in cotton. <br /> " Ownley - Application of Beauveria bassiana 11-98 to seedling roots induced systemic resistance in cotton leaves against the bacterial pathogen Xanthomonas axonopodis pv. malvacearum. <br /> <br /> Objective 2. Examine the effect of cultural practices on soilborne pathogens and plant growth.<br /> " Keinath - To maximize biomass production in coastal South Carolina, Cahaba White vetch should be seeded in October or November and incorporated in January or February.<br /> " Keinath - Cahaba White vetch reduced the number of wilted seedless watermelon plants in one of two experiments.<br /> " Keinath - The plant defense activator Actigard increased the number and weight of seedless watermelons.<br /> " Rothrock  The rate of brassica green manures had a greater effect on disease severity of Rhizoctonia in impatiens and petunia, and Meloidogyne incognita in cucumber, than the source of the green manures.<br /> " Rothrock  In soils containing varying amounts of sand, infested with Meloidogyne incognita and/ or Thielaviopsis basicola, height of cotton plants was greatly reduced in the sandiest soil with both pathogens present. Thielaviopsis basicola reduced plant growth over all soil textures.<br /> " Rothrock - Pythium species were the most important pathogens in stand establishment of rice in cool/wet environmental conditions in field and controlled environmental studies based on stand response to metalaxyl seed treatment. <br /> " Westphal  In soybean monoculture no-tillage soil infested with Fusarium solani f. sp. glycines (causes sudden death syndrome; SDS) and soybean cyst nematode (Heterodera glycines), foliar symptoms of SDS were less severe in soil that had not been fumigated than in fumigated plots three years after pathogen infestation, suggesting that microbially-based soil suppressiveness had developed to these pathogens in the non-fumigated soil.<br /> " Westphal  Foliar symptoms of SDS in soybean were greater with chisel and moldboard plow tillage than with ridge tillage; yields were also improved with reduced tillage compared to intensive tillage.<br /> <br /> Objective 3. Examine the genetic diversity of Rhizoctonia solani between natural ecosystems and agricultural ecosystems.<br /> " Cubeta  A field population of R. solani anastomosis group 3 (AG-3) consisting of 59 isolates was examined to determine the genetic diversity and evolutionary history of the genetic element M2 dsRNA This genetic element is hypothesized to be associated with altering the expression of metabolic pathways involved in parasitic and saprobic activity of the fungus. Results suggest that differential selective forces of mutation and recombination have contributed to the evolution of the M2 dsRNA genome. The distinct lineages and patterns of evolution inferred with coalescent analyses were unique for the M2 dsRNA genome.<br /> " Cubeta - The M2 dsRNA genetic element was detected in representative isolates belonging to three anastomosis groups (AG) of R. solani (AG-1-IA, AG-4, and AG-6; teleomorph = Thanatephorus) and four AG of binucleate Rhizoctonia (AG-A, AG-F, AG-R, and AG-U; teleomorph = Ceratobasidium) using reverse-transcription polymerase chain reaction (PCR). Phylogenetic analysis of M2 dsRNA sequence data resulted in seven inferred haplotypes and there was no unique association with AG to support co-evolution of the M2 dsRNA haplotype within the fungal host. Based on the rooted gene genealogy inferred from coalescent analyses, the ancestral M2 dsRNA haplotype most likely evolved in R. solani AG-1-IA and has recently been acquired by isolates of Ceratobasidium. This is likely the first report of a dsRNA occurring in isolates of binucleate Rhizoctonia and other AG of R. solani.<br /> " Cubeta - Horizontal transmission of the 3.57 kb M2 dsRNA between mycelia of somatically incompatible isolates of Rhizoctonia solani AG-3, an economically important pathogen of cultivated plants in the family Solanaceae, was investigated. Nine donor isolates of R. solani AG-3 containing the M2 dsRNA were paired on potato dextrose agar with each of three different recipient isolates where the M2 dsRNA was absent. Reverse-transcription PCR (RT-PCR) was used to detect horizontal transmission of the M2 dsRNA via hyphal anastomosis from donor to recipient isolates by examining hyphal explants taken 3-cm from the hyphal interaction zone. PCR-RFLP genetic-based markers of two nuclear loci and one mitochondrial locus were used to confirm identity and transmission between donor and recipient isolates of R. solani AG-3. The frequency of transmission observed between 72 pairings of the eight donor and three recipient isolates was approximately 4% of the total pairings and differences in the phenotype of the recipient isolates after acquisition of the M2 dsRNA via horizontal transmission were observed. To our knowledge, this represents the first demonstration of transmission of dsRNA between genetically different individuals of R. solani confirmed with nuclear and mitochondrial markers. These results suggest that transmission can occur between somatically incompatible isolates of R. solani AG-3, but that maintenance of the dsRNA in the recipient isolates was not stable following repeated sub-culturing on nutrient medium.<br /> " Cubeta - The nuclear intergenic spacer (IGS) and a portion of the mitochondrial large subunit (mtLSU) ribosomal DNA (rDNA) genes from Sclerotinia minor and closely related species were amplified with PCR. The amplified products were sequenced to identify regions of polymorphism to develop oligonucleotide primers for specific amplification of S. minor. One primer (Sm-F) was designed and used with the IGS-12a primer previously developed by Carbone and colleagues for specific amplification of the IGS rDNA region of S. minor. Two additional primers (MtSm-F and MtSm-R) were developed for specific amplification of the mtLSU rDNA region of S. minor. The IGS primers amplified a 260-bp fragment from all isolates of S. minor examined, while the mtLSU primers amplified a 153-bp fragment from all but one isolate of S. minor. Both sets of primers did not amplify DNA of peanut (Arachis hypogaea), Botrytis cinerea, S. homeocarpa, S. sclerotiorum, and S. trifoliorum. The minimum concentration required for amplification of DNA of S. minor with the IGS primers ranged between 50 to 100 pg, while the minimum concentration required for amplification with the mtLSU primers ranged from 1 to 10 ng. DNA of S. minor was detected from asymptomatic and symptomatic leaflets, lateral branches, and pegs of peanut with both sets of primers, but in higher frequency with the IGS primers. Detection of S. minor with the IGS primer set was concordant with results obtained by direct isolation of the fungus from infected peanut tissue.<br /> " Rothrock  Rhizoctonia solani from anastomosis groups 4, 7, and 11, and binucleate Rhizoctonia spp. were isolated from cotton seedlings and soil from cotton-producing states participating in the National Cottonseed Treatment Program, suggesting that a diversity of Rhizoctonia isolates is present across the country. <br /> <br /> Grant Funding Relevant to Objectives<br /> " Canaday, C., A. Mengistu, M. Newman, and P. Donald. Screening of Roundup Ready Soybean Varieties and Breeding Lines for Charcoal Rot, SCN and other Yield Limiting Diseases. Amount: $26,000. Sponsor: Tennessee Soybean Promotion Board. Time: 01/01/2007  03/31/2008. <br /> " Dick, W. Graduate Fellowships in Soil Microbial Ecology and Environmental Science: Focus on Bioremediation, Biosecurity and Biogeochemical Cycling. Amount: $207,000. Sponsor: CSREES-USDA. Time: 09/01/2005 - 08/31/2008.<br /> " Ownley, B., and K. Gwinn. Integration of Beauveria bassiana and Trichoderma harzianum for Control of Soilborne Pathogens in Tomato. Amount: $6,529. Sponsor: IR-4 Biopesticide Research Program. Time: 06/30/2007  06/29/2008. <br /> <br /> <br />

Publications

Impact Statements

1. " Two biocontrol products (binucleate Rhizoctonia isolate and BioYield Flowable) were as effective as the soil-applied fungicide Terraclor in reducing wirestem on broccoli at 30 days after transplanting, i.e. during the period when plants are most susceptible to stem girdling.
2. " Four root-colonizing biocontrol fungi effectively induced resistance in host plants against foliar pathogens, including induction of resistance in cotton by Beauveria bassiana 11-98 against Xanthomonas axonopodis pv. malvacearum and induction of resistance in geranium by binucleate Rhizoctonia isolates BNR621 and P023, and Trichoderma hamatum 382 against Botrytis cinerea. These are the likely the first reports of induction of plant resistance for the BNR isolates and Beauveria bassiana.
3. " A seed treatment combination of the biological control PGPR strain Bacillus subtilis MBI 600 and the fungicides mefenoxam and fludioxinil significantly increased healthy plant stand of soybean.
4. " Foliar symptoms of SDS and yields in soybean were reduced with reduced tillage compared to intensive tillage.
5. " The genetic element M2 dsRNA, which likely evolved in R. solani AG-1-IA, was detected in isolates of binucleate Rhizoctonia spp. and other AG of R. solani; this is likely the first report of dsRNA in binucleate Rhizoctonia and other AG of R. solani.

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Brief Summary of Minutes

Accomplishments

Accomplishments/Outcomes:<br /> <br /> Objective 1. Examine commercial and non-commercial biocontrol agents for use as seed treatments, in-furrow treatments or as potting mix amendments.<br /> <br /> The first regional trial of biocontrol agents and amendments for control of wire stem of broccoli (Rhizoctonia solani) was conducted at five sites in four states in 2007. At Jackson, TN (Canaday), all materials reduced the incidence of Rhizoctonia root rot or Pythium root rot compared to the untreated control and plant growth-promoting rhizobacteria (PGPR) were equal to the fungicide standard for disease control with no adverse effects on growth or yield. In eastern TN (Ownley), broccoli seed treated with Beauveria bassiana 11-98 (1,000,000 spores/seed), followed by planting into potting mix containing 2% bioactive dried Monarda herbage resulted in significantly greater head size and yield weight than the standard fungicide treatment (etridiazole soil treatment and thiram seed treatment) or other treatments evaluated. In South Carolina (Keineth), application to potting mix of Monarda herbage (2% w/w) and BioYield Flowable increased broccoli plant height at 50 days after transplanting compared with the pathogen-infested control. Terraclor increased the number of marketable heads per plot compared with the infested control. In Arkansas (Rothrock), no differences were found in number of healthy plants, plant height or leaf length. The regional trial was planted again in the fall of 2008 at the five sites. The eight treatments had no effect on transplant growth; an untreated control, a fungicide standard, PGPR, a binucleate Rhizoctonia sp., a commercial preparation of Streptomyces lydicus (WYEC 108), a seed treatment with Beauveria bassiana (Bb11-98), and Monarda herbage alone and in combination with Bb11-98. After transplanting in TN (Canaday), the Monarda herbage plus Bb11-98 combination increased plant growth more than any single biocontrol agent. In South Carolina (Keineth), wirestem incidence averaged 7% in noninfested plots and over 36% in plots infested with 15 sclerotia of Rhizoctonia solani AG-4 per kilogram soil. In infested plots, only azoxystrobin fungicide, which was applied to soil at transplanting, reduced wirestem compared with the nontreated infested control. Wirestem incidence did not differ among treatments in noninfested plots. Sites in Kentucky (Seebold) and Arkansas (Rothrock) in 2008 were infested, but incidence of disease was low and no significant differences were found among any of the treatments. <br /> <br /> Other biological control studies were conducted by individual project members. A trial conducted in KY (Seebold) found Bioten, a commercial formulation containing Trichoderma harzianum and T. viride, significantly reduced Phytophthora blight, caused by Phytophthora capsici, on summer squash when applied to soil via drip irrigation (60% reduction over the untreated control), while Ridomil Gold reduced incidence by at least 90% compared to the untreated control. In TN (Ownley), integration of Beauveria (B. bassiana 11-98 or BotaniGard) seed treatments and RootShield (Trichoderma) potting mix treatment provided mixed results on improvement of growth variables of tomato seedlings in soil infested with Rhizoctonia solani or Pythium myriotylum. Growth was greater with thiram seed treatment, followed by B. bassiana 11-98 seed treatment in one Rhizoctonia trial for most variables, but shoot weight was improved by the combination of Beauveria seed treatments and RootShield in the second trial. In the first Pythium trial, shoot height, and leaf number were greater than the untreated control for seed treatments with BotaniGard, B. bassiana 11-98 and thiram. Stem diameter was greatest for plants from BotaniGard seed treatment. In the second Pythium trial, shoot weight was greatest with the individual treatments of Root Shield and thiram seed treatment. In LA (Padgett), the efficacy of several commercial fungicides and one biological fungicide was evaluated on foliar, pod, and stem diseases on soybean. Foliar treatments were applied during R3 or R3 and R5. Foliar diseases did not develop to appreciable levels during the growing season, however, pod diseases were prevalent late season. Percent defoliation and pod diseases did not differ among treatments. Yields were greater for the treatments Ballad Plus at 2.0 qt/A tank mixed with 3.0 fl oz of Headline and 0.5 qt/A tank mixed with 6.0 fl oz of Headline than the non-treated control. In FL (Elliot), container-grown mexican fan palm (Washingtonia robusta) and queen palm (Syagrus romanzoffiana) transplanted into a field nursery having phosphorus (P)-sufficient and P-deficient soils were treated at the time of planting with four commercial microbial inoculants (each containing arbuscular mycorrhizal fungi, either alone or with other microbial components or fertilizers), two fertilizers, or nothing (control). None of the treatments improved growth over the control in the P-deficient soil. In the P-sufficient soil, none of the microbial inoculants improved growth over that of similarly fertilized non-inoculated palms. Discrepancies were observed regarding non-mycorrhizal fungi and bacteria present in the microbial inoculant products from the product labels. In TN (Canaday), response to Bacillus subtilis (MBI600) when added to fungicide seed treatments for control of seedling diseases of spring-planted snap beans varied with the cultivar used. Adding B. subtilis to a seed treatment of fludioxonil + mefenoxam increased stand loss and lowered yield with Carlo but reduced stand loss and increased yield with Tapia. Adding B. subtilis to a seed treatment of trifloxystrobin + metalaxyl increased stand loss and lowered yield of Carlo but decreased stand loss and increased yield of Tapia. Ashy stem blight of summer-planted snap beans was best controlled and highest yields obtained when an in-furrow spray of boscalid was combined with seed treated with Captan or Captan plus B. subtilis. Potash fertilizers, seed treatments, and in-furrow fungicide sprays were evaluated for control of charcoal rot of soybean (Macrophomina phaseolina). The highest soybean yield was obtained with a seed treatment of Captan plus B. subtilis.<br /> <br /> <br /> Objective 2. Examine the effect of cultural practices on soilborne pathogens and plant growth.<br /> <br /> In FL (Elliot), bermudagrass root (2 sites, one each in SC and FL) and bentgrass root (2 sites, one each in NC, AL) microflora in sand-based putting greens were examined by analyzing over 9000 randomly selecting bacterial colonies using gas chromatography for analysis of fatty acid methyl ester profiles (GC-FAME). The two dominant genera in both bentgrass and bermudagrass rhizospheres were Bacillus and Pseudomonas, with Bacillus dominant in bermudagrass and Pseudomonas dominant or equal to Bacillus in bentgrass. Other genera that comprised at least 1% of the isolates at all four sites were Clavibacter, Flavobacterium and Microbacterium. Arthrobacter also comprised a significant portion of the bacterial isolates in the bentgrass rhizosphere, but not the bermudagrass rhizosphere. Overall, there were 40 genera common to all four sites. At the species level, there were five that comprised at least 1% of the isolates at each location; B. cereus, B. megaterium, C. michiganensis, F. johnsoniae and P. putida. In IA(Loynachan), a large diversity of mycorrhizal fungi was documented in 8 soils of 4 soybean fields. Large variability exists within a single soil within a field. The composition of individual species varied within soil samples collected 3 m apart. Five species (Glomus. claroideum, G. etunicatum, G. mosseae, G. viscosum, and Paraglomus occultum) were detected in all samples collected.<br /> <br /> Cultural practices, including raised planting beds composed of wood chips or pine bark mulch, and soil amendment of cow compost to promote biological activity or sulfur to lower pH, were evaluated at five sites for disease management of Phytophthora root rot of Fraser fir (Abies fraseri) grown for Christmas tree production on disease conducive, infested soils in four western NC counties (Benson with Brantlee Ricther, PhD Thesis, and Kelly Ivors, Asst. Prof.). Mulches established increased levels of bacterial and fungal populations, total microbial activity, and cellulase activity, relative to underlying soil. Total microbial activity and cellulase activity remained elevated in mulches, relative to soil treatments, through the first two growing seasons. Mulched plots showed significantly reduced disease at three of five sites through the third growing season, but mortality rates were high across treatments. Possible mechanisms of control in the raised beds include improved drainage and suppression of sporangia production by Phytophthora spp., as well as biological control by cellulose-degrading microbes in the wood chips that will produce cellulase and degrade Phytophthora propagules surviving in soil. Laboratory assays are being conducted to determine if cellulase levels in mulches similar to those observed in the field trials is sufficient to inhibit growth of P. cinnamomi. <br /> <br /> Cover crops were examined for disease suppression in South Carolina and Arkansas. An on-farm trial was done in SC (Keineth) to evaluate the effects of cover crops and fumigants on Fusarium wilt and yield of seedless watermelon. The two fumigants, 67% methyl bromide-33% chloropicrin and Telone-C35, both combined with hairy vetch (Vicia villosa), were more effective than hairy vetch or rye alone or fallow at increasing yield. Methyl bromide-chloropicrin was more effective at reducing symptoms than Telone C-35. In AR (Rothrock), a Brassica winter cover crop of Indian mustard cv. Fumus was compared with the chemical Telone II and a fallow control in cotton fields having seedling diseases and severe reniform or root-knot nematode infestations. Brassica amendments were observed to reduce seedling root and hypocotyl disease symptoms. Brassica amendments also reduced early season galling from the root-knot nematode similar to the Telone treatment. Brassica treatments improved cotton yield over the control and were similar or greater than the Telone treatment in both the root-knot and reniform nematode infested locations. Brassica amendments including biomass of Brassica juncea cv. Fumus, specifically bred for high levels of glucosinolates, B. napus cv. Jetton, and B. juncea cv. Bionute also significantly reduced disease symptoms and recovery of Rhizoctonia solani from petunia and impatiens. Brassica crops were not significantly different from each other. Rate of Brassica application had the greatest impact on disease symptom reduction and reducing Rhizoctonia solani isolation, with a rate of 3000 g fresh above-ground biomass/square m being more effective than lower rates. <br /> <br /> In TN (Canaday), the effects of reduced rate fungicides combined with acibenzolar-S-methyl (a SAR inducer) on the incidence of timber rot (Sclerotinia sclerotiorum) and buckeye rot (Phytophthora nicotianae) affecting eight tomato cultivars were evaluated. No significant effects on these soilborne diseases were noted. However, it was shown that routine fungicide sprays may be applied at 1/3 the normal rate in combination with the SAR inducer without significant yield loss or loss in disease control efficacy. The percentage of culled fruit was lowest with 1/3 rate fungicides plus the SAR inducer. <br /> <br /> Objective 3. Examine the genetic diversity of Rhizoctonia solani between natural ecosystems and agricultural ecosystems.<br /> <br /> Rhizoctonia solani was isolated from seedlings and soil from cotton producing states participating in the National Cottonseed Treatment Program in 2008. Isolates were characterized using the anastomosis technique. AG 4, AG 7, and AG 11 were found on cotton from different states suggesting a diversity of R. solani is present across the country. Binucleate Rhizoctonia spp. were found in many of the samples. <br /> <br /> <br /> Outputs:<br /> <br /> Papers were published on the taxonomic diversity of rhizosphere bacteria associated with golf course putting greens, double stranded RNA viruses in Rhizoctonia solani, the biological control agent Beauvaria bassiana, and the use of Brassica amendments. <br /> <br /> Reports were published on the regional studies using biological control agents for the control of wirestem, caused by Rhizoctonia solani, on broccoli. <br /> <br /> A new peer-reviewed video (Soil Fungi) has been made available through the MicrobeLibrary from the American Society of Microbiology. A soil biology educational website containing video of important soil organisms including mycorrhizae and their activities has been maintained (http://www.agron.iastate.edu/~loynachan/mov/).<br /> <br /> Activities:<br /> <br /> Biological control of wirestem on broccoli, caused by Rhizoctonia solani, was evaluated at five locations (KY, TN-Knoxville, TN-Jackson, AR, and SC) in 2007 and 2008. Field trials are being planned for the 2009 season based on the results from previous tests. Biocontrol trials on impatiens and tomato are being discussed to see if regional evaluations can be expanded to other vegetables and an ornamental crop in 2009. <br /> <br /> A regional evaluation of techniques for isolation of Rhizoctonia solani from soil with different cropping histories is being planned for 2009. Analyses of recovered isolates of R. solani by anastomosis groups (AG) and other techniques will allow the diversity of the pathogen to be examined based on geography and cropping history.<br /> <br /> <br />

Publications

Books/Book Chapters<br /> <br /> Charlton, N.D., Tavantzis, S.M. and Cubeta, M.A. 2008. Detection of double<br /> stranded RNA viruses in the soil fungus Rhizoctonia solani, In Plant<br /> Pathology Techniques and Procedures, Chapter 14, pp. 171-182. Eds. R. Burns,<br /> Humana Press, 2nd Edition, Tocawa, NJ.<br /> <br /> Referred Journal Articles<br /> <br /> Charlton, N.D., Carbone, I., Tavantzis, S.M., and M.A. Cubeta. 2008.<br /> Phylogenetic relatedness of the M2 double stranded RNA in Rhizoctonia fungi.<br /> Mycologia 100:555-564.<br /> <br /> Elliott, M.L., McInroy, J.A., Xiong, K, Kim, J.H., Skipper, H.D., and Guertal, E.A. 2008. Taxonomic diversity of rhizosphere bacteria in golf course putting greens at representative sites in the southeastern United States. HortScience 43:514-518.<br /> <br /> Ferrucho, R.L., Zala, M., Zhang, Z., Cubeta, M.A., Garcia-Dominguez, C., and<br /> Ceresini, P.C. 2008. Highly polymorphic in silico-derived microsatellite<br /> loci in the potato-infecting fungal pathogen Rhizoctonia solani anastomosis<br /> group 3 from the Colombian Andes. Molecular Ecology Resources (In Press).<br /> <br /> <br /> Leckie, B. M., B. H. Ownley, R. M. Pereira, W. E. Klingeman, and C. J. Jones. 2008. Effects of Beauveria bassiana (Balsamo) Vuillemin mycelia and metabolites incorporated into synthetic diets on larval Helicoverpa zea. Biocontrol Sci. Technol.18: 697-710.<br /> <br /> Ling, K. S., Kousik, C. S., and Keinath, A. P. 2008. First report of Southern blight on bottle gourd (Lagenaria siceraria) caused by Sclerotium rolfsii in South Carolina. Plant Dis. 92:656.<br /> <br /> Njoroge, S. M. C., Riley, M. B., and Keinath, A. P. 2008. Effect of incorporation of Brassica spp. residues on population densities of soilborne microorganisms and on damping-off and Fusarium wilt of watermelon. Plant Disease 92: 287-294.<br /> <br /> Ownley, B. H., M. R. Griffin, W. E. Klingeman, K. D. Gwinn, J. K. Moulton, and R. M. Pereira. 2008. Beauveria bassiana: endophytic colonization and plant disease control. J. Invertebra. Pathol. 98: 267-270.<br /> <br /> <br /> Abstracts<br /> <br /> Bartz, F.E., Danehower, D.A., Tavantzis, S.M., and Cubeta, M.A. 2008. Carbon<br /> metabolism and plant growth regulation: the influence of quinic acid on<br /> phenylacetic acid production and pathogenic activity of Rhizoctonia solani<br /> AG-3. Proc. 4th International Rhizoctonia Symposium, Berlin, Germany<br /> (Abstract).<br /> <br /> <br /> Bartz, F.E., Tavantzis, S.M., Danehower, D.A., and Cubeta, M.A. 2008.<br /> Influence of quinic acid catabolism on the production of the plant growth<br /> regulator phenylacetic acid by Rhizoctonia solani AG-3. Inoculum 59:19 (Supplement to Mycologia, Abstract).<br /> <br /> Bartz, F.E., Tavantzis, S.M., Danehower, D.A., and Cubeta, M.A. 2008.<br /> Influence of quinic acid catabolism on the production of the plant growth<br /> regulator phenylacetic acid by Rhizoctonia solani AG-3. Phytopathology<br /> 98:S18 (Abstract).<br /> <br /> Charlton, N.D., Carbone, I., Tavantzis, S.M., and Cubeta, M.A. 2008.<br /> Evolutionary history and population genetics of the M2 double-stranded RNA<br /> of Rhizoctonia solani anastomosis group 3. Proc. 4th International<br /> Rhizoctonia Symposium, Berlin, Germany (Abstract).<br /> <br /> Copes, W.E., Rinehart, T. A., Toda, T., and Cubeta, M.A. 2008. Rhizoctonia<br /> species associated with bark media and plant strata of container-grown<br /> azalea. Proc. 4th International Rhizoctonia Symposium, Berlin, Germany<br /> (Abstract).<br /> <br /> <br /> Cubeta, M.A., Dean, R. A., Bayman, P., Jabaji, S., Neate, S., Nolte, P.,<br /> Tavantzis, S.M., Toda, T. Vilgalys, R., Federova, N., and Nierman, W.C.<br /> 2008. Whole genome sequencing of the soil fungus Rhizoctonia solani AG-3.<br /> Inoculum 59:21 (Supplement to Mycologia, Abstract).<br /> <br /> Gomaa, N. M., Rothrock, C. S., and Jia, Y. 2008. Sequence variation of the rDNA internal transcribed spacer (ITS) region among isolates of Rhizoctonia solani. Proceedings of the 4th International Symposium on Rhizoctonia.<br /> <br /> Greenberg, W., L. Hartley, T. Loynachan, D. Lindbo, and S. Schultz. 2008. Maintaining and updating the SSSA K-12 education website. 531-7 Agron. Abstracts, Madison.<br /> <br /> Kaye, A., Parks, L., Kennedy, G., Shew, B.B., Carbone, I., Cubeta, M.A., and<br /> Moyer, J.W. 2008. Population genetics of Tomato Spotted Wilt Virus on peanut<br /> in North Carolina and Virginia. Phytopathology 98:S79 (Abstract).<br /> <br /> Ownley, B. H., M. M. Dee, and K. D. Gwinn. 2008. Effect of conidial seed treatment rate or entomopathogenic Beauveria bassiana 11-98 on endophytic colonization of tomato seedlings and control of Rhizoctonia disease. Phytopathology 98:S118. (Abstract.)<br /> <br /> Richter, B. S., Benson D. M., and Ivors, K. L. 2008. Cellulase activity and microbiology of cultural systems for Phytophthora root rot control in Fraser fir. Phytopathology 98:S132 (Abstract).<br /> <br /> Toda, T., Upchurch, R.G., and Cubeta, M.A. 2008. Development of an<br /> Agrobacterium-based transformation system for Rhizoctonia solani. Proc. 4th<br /> International Rhizoctonia Symposium, Berlin, Germany (Abstract).<br /> <br /> <br /> Toda, T., Strausbaugh, C., Vilgalys, R., and Cubeta, M.A. 2008.<br /> Characterization of a basidomycete fungus from sugarbeet. Inoculum 59:59 (Supplement to Mycologia, Abstract).<br /> <br /> Peer-Reviewed Journal Reports<br /> <br /> Canaday, C.H., 2008. Effects of cultivar, a SAR inducer, and PGPR on foliar diseases and yields of staked-tomato, 2007. Plant Disease Management Reports 2:V029.<br /> <br /> Canaday, C.H. and A. Mengistu, 2008. Effects of seed treatments, in-furrow sprays, and herbicide timing on charcoal rot of soybean, 2007. Plant Disease Management Reports 2:FC014.<br /> <br /> Canaday, C.H. and A. Mengistu, 2008. Effects of directed fungicide sprays and potash form on charcoal rot of soybean, 2007. Plant Disease Management Reports 2:FC015.<br /> <br /> Canaday, C.H., Ownley, B.H., Gwinn, K.D., Rothrock, C.S., Keinath, A.P., and Seebold, K.W. 2008. Effects of biocontrol agents versus a fungicide on growth, yield, and diseases of broccoli in Tennessee, 2007-2008. Plant Disease Management Reports 2:V145.<br /> <br /> Keinath, A.P., V.B. DuBose, A.W. Lassiter, C.H. Canaday, C S. Rothrock, B.H. Ownley, 2008. Evaluation of biological controls against wirestem of broccoli in South Carolina, 2007. Plant Disease Management Reports 2:V152.<br /> <br /> Ownley, B.H., Gwinn, K.D., Dee, M.M., Canaday, C.H., Keinath, A.P., Rothrock, C.S., and Seebold, K.W. 2008. Effects of biocontrol agents on growth, yield, and root disease of broccoli in East Tennessee, 2007-08. Plant Disease Management Reports 2:V155.<br /> <br /> Seebold, K.W., Holdcroft, A.M., and Dixon, E. 2008. Effect of potassium phosphite and fungicides on Phytophthora crown and fruit rot of summer squash, 2007. Plant Disease Management Reports 2:V005.<br /> <br /> <br /> Other<br /> <br /> Loynachan, T. E. 2007. Endomycorrhizae [Online]. Available at http://www.microbelibrary.org/asmonly/details.asp?id=2504&Lang=. MicrobeLibrary. American Society for Microbiology.<br /> <br /> Loynachan, T. E. 2007. Nematode-trapping fungi [Online]. Available at http://www.microbelibrary.org/asmonly/details.asp?id=2506&Lang=. MicrobeLibrary. American Society for Microbiology.<br /> <br /> Loynachan, T. E. 2007. Ectomycorrhizae [Online]. Available at http://www.microbelibrary.org/asmonly/details.asp?id=2508&Lang=. MicrobeLibrary. American Society for Microbiology.<br /> <br /> Loynachan, T. E. 2008. Soil fungi [Online]. Available at http://www.microbelibrary.org/asmonly/details.asp?id=2801&Lang=. MicrobeLibrary. American Society for Microbiology.<br />

Impact Statements

1. The cash value of vegetable crops was $20.5 billion in the U.S., with estimated losses of up to 10% due to soilborne plant pathogens. For growers intent on not using pesticides, production of pesticide-free or organic crops can increase crop value by 30%. Effective biopesticide treatment identified in the regional broccoli experiment, and the integrated treatments evaluated in the tomato experiments have not been previously tested. A commercial formulation of Trichoderma viride and T. harzianum, applied to soil by drip irrigation, was shown to suppress the incidence of Phytophthora blight on summer squash. Although not as effective as commercially available fungicides, using the biocontrol agents in conjunction with commercial fungicides may permit fewer applications or allow for reduced rates of the fungicide products. These treatments have the potential to increase yield of field grown vegetable crops and greenhouse grown transplants.
2. Four commercially-available mycorrhizal/bacterial/fungal inoculants for improving growth of palms (Syagrus romanzoffiana and Washingtonia robusta) were evaluated for use in nurseries.
3. Hairy vetch was adopted by a large watermelon grower in South Carolina as a winter cover crop in fields with a history of losses to Fusarium wilt. Acreage planted to hairy vetch increased three-fold from 2007 to 2008. Brassica winter cover crops were shown to reduced damage from soilborne pathogens on cotton and increase cotton yields. Brassica cover crops appear to be a viable alternative to the application of Telone II in cotton.
4. Changing potash form from muriate of potash to sulfate of potash reduced soybean preemergence damping-off and increased yield by over 180 kg/ha (2.68 bu/A). Use of biological agents with fungicide seed treatments was shown to be dependent on cultivar for snap bean seedling diseases and yield.
5. Grant Funding Relevant to Objectives: Ownley, B., and K. Gwinn. Integration of Beauveria bassiana and Trichoderma harzianum for Control of Soilborne Pathogens in Tomato. Amount: $6,529. Sponsor: IR-4 Biopesticide Research Program. Time: 06/30/2007 to 12/31/2008. Cochran, K. and C. S. Rothrock. Importance of Brassica soil amendments for managing soilborne disease in ornamentals and vegetables. Amount $9,944. Sponser: Graduate student Project, Southern Region SARE Program.

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Report Information

Participants

A. Keinath (SC), M. Elliott (FL), C. Canaday (TN), C. Rothrock (AR), C. Garzon (OK), K. Seebold (KY), B. Ownley (TN), K. Gwinn (TN, visitor), and Andrea Vu (TN, visitor).

Brief Summary of Minutes

The meeting, which was held in Room 410 of the Plant Biotechnology Building at UTK, was called to order by A. Keinath, Chair, at approximately 9:10 a.m. B. Ownley, local arrangements host, welcomed the attendees and visitors were introduced.

Objective 1. The first order of business was to discuss the cooperative project to evaluate new biocontrol agents and materials for control of wirestem on broccoli. Eight treatments were evaluated at five locations (KY, TN-Knoxville, TN-Jackson, AR, and SC) in the fall 2008-winter 2009 test. Three treatments were incorporated into the potting mix: millet seed infested with binucleate-Rhizoctonia (BNR) at 0.56% dry wt., Monarda herbage at 2% dry wt., and BioYield Flowable, a source of plant growth-promoting rhizobacteria, at 0.75 fl oz/1000 plants. Two treatments included seed treated with Beauveria bassiana isolate 11-98 at 1 x 10 7 cfu/seed in a 2% methyl cellulose solution. An untreated control, a fungicide standard (Quadris 2.08F or Terrachlor 75WP), and a treatment of two applications of Actinovate AG (one 1 day after seeding trays and again 1 day before transplanting) were also included.

Results were a little disappointing. In SC, only 50% of the plots were infested with Rhizoctonia solani and no effects on disease or yield were noted. In KY, rodents destroyed the test before any data was recorded. In AR, only yield data was recorded. In TN at Knoxville, plants were lost to winter injury before data collection. In TN at Jackson, there was also no disease, but a treatment effect on plant growth was noted: Plants with Beauveria bassiana 11-98 seed treatment + 2% Monarda herbage appeared to grow faster than most other plants and yielded significantly more than all other treatments except the fungicide standard. In subsequent discussions, C. Canaday and B. Ownley reported that changes in the research focus of UT's branch research stations will make continued participation in the broccoli regional research tests more difficult. No test was planned for fall 2009. C. Canaday will see if there's sufficient horticultural data for a journal article in HortTechnology on the effects of the biocontrol treatments on broccoli growth.

Meeting attendees then reported on their research related to Objective 1. C. Canaday reported on his research on the effects of potash treatments and seed treatment biocontrol agents on plant stand and yield of soybean. K. Gwinn reported on her research on the volatile components of Monarda herbage, their percentages of total extracted oils, and their EC50 values for R. solani. C. Rothrock reported on his research on Brassica biofumigation comparing use of Brassica napus cv. Bionute (broadcast over cotton fields) to the commercial fumigant Telone. A. Keinath reported on his research on the use of root grafts to control Fusarium oxysporum f. sp. niveum on seedless watermelon.

Objective 3. Committee members reported on the results of their evaluation of treatments for recovery of anastomosis groups (AG) of Rhizoctonia solani from soil using two techniques (soil pelleting and toothpick baits) and three media (modified ethanol-potassium nitrate medium [Plant Disease 71:1098-1100; substitute 300 ppm streptomycin and 100 ppm rifampicin for 100 ppm tobramycin and use 2% ethanol instead of 5%], Ko and Hora medium [Phytopathology 61:707-710], and water agar + benomyl + chloramphenicol). M. Elliott found the poorest growth on the water agar and best growth on Ko and Hora medium. B. Ownley also reported best results with Ko and Hora medium. C. Rothrock and A. Keinath reported that the ethanol-potassium nitrate medium was best. Ko and Hora medium was acceptable, but Pythium and Trichoderma contaminants were a problem. C. Rothrocks Ph.D. student plans additional tests with toothpick baits. A. Keinath reported that toothpick isolation was better than soil pellets and better than bean or broccoli seedlings. He found that prochloraz slowed the growth of R. solani from soil and that rifampicin was a suitable substitute for tobramycin. C. Garzon reported that she plans to compare soil pellets versus molecular techniques in her course on soilborne diseases of plants.

Members then discussed protocol details for Objective 3. Final protocol details included:

1. Collect soil to a depth of 7.5 cm from five locations per site. Ideally, collect soil from agricultural field site about two weeks after planting, from within the row.

2. Don't store soil. Refrigerate no more than two weeks before assay.

3. Uniformly mix soil and remove large pieces of organic debris (no need to screen soil). Save enough soil for physical (textural) and chemical analysis; air dry this sub-sample.

4. For each sample, fill four 10-cm-diameter plastic, square greenhouse pots 7.5 cm deep with soil.

5. Water soil with tap water until it drains. Wait 24 hours.

6. For three of the pots, place nine white birch toothpicks (flat) vertically in soil to a depth of 5 cm (1 cm remaining above the soil line) about 2.5 cm apart in a 3 x 3 grid in soil in each pot. Do not autoclave toothpicks.

7. With the fourth pot, remove 10 g of soil, dry it for 24 h at 105 C and reweigh for gravimetric water analysis.

8. Place pots with toothpicks in the dark at 22±1 C.

9. After 48 h, remove toothpicks and place on modified ethanol-potassium nitrate medium (see above), 3 toothpicks per plate. Place plates in the dark at 22±1 C. Agri-Mek 0.15EC (abamectin) (5 ppm [0.275 ml per L]) may be added to inhibit mites.

10. Obtain about 500 g of soil from these three pots, spread thin in a tray and autoclave for 20 minutes. Then freeze at -20 C for future shipment to C. Garzon (but do not ship until Garzon makes the request).

11. After 24 to 72 h, record the number of colonies of Rhizoctonia growing from the toothpicks. Collect every suspect Rhizoctonia colony and transfer to full-strength PDA.

12. Save 10 colonies per sample for genetic analysis and anastomosis group determination. Store isolates as colonized agar plugs in sterile deionized water (minimize amount of agar added to storage vials). Store at room temperature.

Plans for work under Objective 2 not already mentioned were covered in state reports:

Arkansas. C. Rothrock plans to do more R. solani AG-group diagnoses.

Oklahoma. C. Garzon reported that she plans to do DNA diversity studies and will use diagnostic primers to test for the presence of Phymatotrichopsis omnivora in asymptomatic hosts. She also will use her Pythium irregulare primers to determine if the pathogen is infesting different commercial nurseries. She will continue her work on the population biology of Sclerotinia minor and is trying to identify the peanut genes for resistance to S. minor. She also will continue to investigate the microbial populations in suppressive and non-suppressive Ecuadorian soils for Phytophthora infestans.

Business Meeting. A. Keinath, chair, called the business meeting to order at 5:40 p.m. C. Garzon was nominated as secretary for 2010. The nomination was seconded and passed unanimously. C. Garzon, who had to depart the meeting earlier in the day, subsequently accepted the secretary nomination via email correspondence with A. Keinath. C. Rothrock offered to host the next meeting in Fayetteville, AR, in November 2010. The offer was accepted by all those present. Members were asked to send state reports to A. Keinath before Christmas break. The meeting was adjourned at 5:50 p.m. Members traveled home on November 15.

Respectfully submitted,
C. Canaday, Secretary

Accomplishments

Publications

Impact Statements

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Report Information

Participants

Backman, Paul (pbackman@psu.edu)  Pennsylvania State University;
Baird, Richard (RBaird@plantpath.msstate.edu)  Mississippi State University;
Benson, Mike (mike_benson@ncsu.edu)  North Carolina State University;
Canaday, Craig (ccanaday@utk.edu)  University of Tennessee;
Cubeta, Marc (marc_cubeta@ncsu.edu) - North Carolina State University;
Dick, Warren (dick.5@osu.edu) - Ohio State University;
Elliott, Monica (melliott@ufl.edu)  University of Florida;
Garzon, Carla Domenica (carla.garzon@okstate.edu)  Oklahoma State University;
Jimenez Gasco, Maria (jimenez-gasco@psu.edu) - Pennsylvania State University;
Keinath, Tony (tknth@clemson.edu)  Clemson University;
Loynachan, Thomas (teloynac@iastate.edu)  Iowa State University;
Lu, Shien (sl332@msstate.edu)  Mississippi State University;
Ownley, Bonnie (bownley@utk.edu)  University of Tennessee;
Padgett, Boyd (bpadgett@agcenter.lsu.edu) - Louisiana State University;
Rothrock, Craig (Rothrock@uark.edu)  University of Arkansas;
Seebold, Kenneth (kwseebold@uky.edu)  University of Kentucky;
Williams, Mark (mw452@msstate.edu)  Mississippi State University;

Brief Summary of Minutes

Attendees: Craig Canaday (TN), Craig Rothrock (AR), Carla Garzon (OK), Terry Spurlock (AR, visitor), Boyd Padgett, and Shouan Zhang (University of Florida, TREC, Homestead - visitor).


The meeting was hosted by University of Arkansas at Fayetteville, AR, in the Rosen Alternative Pest Control Center Conference Room beginning at 8:30 AM. Attendance was sparse due in part to restrictions in interstate travel and the difficulty in finding reasonable airline connections to the meeting site.


Craig Rothrock, local arrangements host, welcomed the attendees and visitors were introduced. Craig Canaday, Chair, opened the official meeting and welcomed reports on project objectives. Carla Garzon, Secretary, recorded the presentations and comments.

Objective 1. Examine commercial and non-commercial biocontrol agents for use as seed treatments, in-furrow treatments or as potting mix amendments.


The third regional trial of biocontrol agents for control of wire stem of broccoli (Rhizoctonia solani) was cancelled due to funding restrictions and changes in resource allocation. A report on the work to date is still in preparation. Shouan Zhang expressed his interest in future collaborations regarding objective 1. Dr. Zhangs research focuses on Biorational management of vegetable diseases including Phytophthora blight of squash, cause by Phytophthora capsici.

Objective 2. Examine the effect of cultural practices on soilborne pathogens and plant growth.


Craig Canaday passed out reprints of the recently published report: Canaday, C. H., and Schmitthenner, A. F. 2010. Effects of chloride and ammonium salts on the incidence of Phytophthora root and stem rot of soybean. Plant Dis. 94:758-765. He presented a brief pictorial history of this work and mentioned that he was continuing to investigate the potential changes in the micro-partitioning of root calcium with chloride salts using energy-dispersive X-ray analysis.


Craig Rothrock reported progress done on research on strawberry treated with green manure. The objective of the study is to determine the effect of green manure on the microbial diversity (bacteria and fungi). A metagenomics approach was used analyzing ribosomal DNA sequences by Denaturing Gradient Gel Electrophoresis (DGGE). The results presented suggest green manure treatment produced evidence of changes in the microflora of two experimental sites, over two years. A brief discussion followed about the diverse soilborne problems that affect strawberries, including Phoma, Rhizoctonia and Pythium, possible sources of inoculum, and management strategies such as soil solarization.

Objective 3. Examine the genetic diversity of Rhizoctonia solani between natural ecosystems and agricultural ecosystems.

Craig Rothrocks graduate student, Terry Spurlock, presented progress reports on comparisons between two baiting protocols (toothpick and soil pellets) and selective media. He modified the toothpick protocol by using twice as many toothpicks as initially recommended in the protocol and wetting the soil. The toothpick protocol seemed the most effective to capture Rhizoctonia solani. It was especially effective to recover AG11 and AG1A. The pellet protocol was more effective to recover R. oryzae. The toothpick protocol involves saprofitic activity and requires high humidity. Ethanol-potassium nitrate medium and TS medium worked better than Ko and Horas. Contamination was a problem with the latter medium. The best baiting/selective media combination reported were toothpick baiting combined with either ethanol-potassium nitrate medium or TS medium. In the discussion which followed it was noted that the toothpick method may work best, since large amounts of soil can be assayed as compared with the pellet protocol.


Craig Rothrock reported on the spatial distribution of R. solani AG1-IA in a soybean-rice rotation. Areas with higher levels of humidity had higher R. solani AG1-IA recovery rates. Isolates were recovered from asymptomatic seedlings (10/site). Epiphytic growth was observed up to 60 cm on the canopy under high humidity conditions. Aerial mycelium and abundant sclerotia were produced. The pathogen was homogenously distributed in the field, and the high incidence hides any spatial patterns. It is apparent that AG1-IA is associated with plant residue and organic matter. Periodic tilling was discussed as possible management practice but its implementation seems unlikely due to its high cost.


Carla Garzon reported that DNA fingerprinting protocols (AFLP, ISSR, SSR) were standardized for analysis of Rhizoctonia solani populations.


Boyd Padgett examined the Rhizoctonia diversity in Louisiana agricultural soils. Toothpick baiting was done in 8 locations (2 corn, 2 cotton, 2 grain sorghum, 2 soybean). A minimum of 31 isolates (from soybean, location 2) and a maximum of 139 isolates (from grain sorghum) of Rhizoctonia were recovered.

Plans for next year.


Rhizoctonia diversity will be compared between AR fields (vegetables, cotton, rice and prairie) and LO (corn, cotton, sorghum, soybean).


Craig Rothrock and Terry Spurlock will write the methods paper.


Carla Garzon will compare R. solani AG1-IA genotype diversity between methods using molecular tools.

Business Meeting.

After the research reports were finished, Craig Canaday, Chair, called the business meeting to order. C. Rothrock was nominated as secretary for 2011. The nomination was seconded and passed unanimously.


C. Garzon, secretary and next years Chair, offered to host the next meeting in Oklahoma City, OK, in November 2011. The offer was accepted by all those present.


The future of the S-1028 project was discussed. C. Canaday and C. Garzon will open discussion to identify new objectives for the project for the next 5 years. It was proposed to encourage team work to produce preliminary data that could be used for NIFA funded research.


C. Garzon will contact Mark McLellan for guidance in the organization of a writing group for the new project. C. Canaday asked members to send him state reports before Thanksgiving. The meeting was adjourned. Members traveled home on November 7.


Respectfully submitted,
Carla Garzon, Secretary

Accomplishments

Accomplishments/Outcomes (by Project Objective and State)<br /> <br /> <br /> Objective 1. Examine commercial and non-commercial biocontrol agents for use as seed treatments, in-furrow treatments or as potting mix amendments.<br /> <br /> <br /> Kentucky: A trial was conducted in the summer of 2010 to evaluate Bioten, a commercial formulation containing Trichoderma harzianum and T. viride, for suppression of Phytophthora blight, caused by Phytophthora capsici, on summer squash when applied to soil via drip irrigation, alone or in conjunction with fungicide programs. In 2009, results of the trial showed that Bioten-containing treatments reduced the severity of Phytophthora blight by 25% over the untreated control.; however, fungicide programs that contained Ridomil Gold applied pre-plant reduced incidence by over 60% compared to the untreated control. The 2010 trial was lost due to flooding of the field. It will be conducted again in 2011.<br /> <br /> <br /> South Carolina: The commercial biofungicides RootShield and Actinovate were tested for prevention of damping-off of arugula on a noncertified organic vegetable farm in Charleston County, South Carolina. Biofungicides were applied immediately after seeding over raised beds with a backpack sprayer at 32 oz/A (RootShield) or 12 oz/A (Actinovate). Water was applied as a control. Emergence and percentage damping-off did not differ significantly among the three treatments, even though Pythium spp. were isolated from 43-53% and Rhizoctonia solani was isolated from 39-78% of the plants sampled. Soil populations of R. solani (measured as percentage of organic matter colonized) were higher in the Actinovate-treated plots (51%) compared to the water control (36%). Fresh weight of arugula harvested 1 month after seeding did not differ among treatments. <br /> <br /> <br /> Tennessee: Three commercial biological seed treatment products, Kodiak Concentrate (Bacillus subtilis GB03), Subtilex (B. subtilis MBI600), and Yield Shield (B. pumilus GB34), were evaluated under a variety of environmental conditions for control of seedling diseases of snap bean and soybean in field tests conducted in soils natural infested with R. solani, Pythium spp., Macrophomina phaseolina, and Fusarium spp. GB03 or MBI600, used alone or as supplements to standard seed treatment chemicals, failed to increase snap bean stand or yield. Similarly, supplementing soybean seed treatment chemicals with GB03 or GB34 failed to increase health plant stand or yield. <br /> <p>Studies were initiated to address the manipulation of a biological control agent, Pasteuria nishizawae, to improve disease management of the soybean cyst nematode (SCN). In order to determine the level of P. nishizawae needed to act as an effective biological control agent, the initial objectives were (1) to determine the best method for detection of P. nishizawae levels in soil and (2) to evaluate methods for quantification of P. nishizawae in SCN cysts. Progress on both objectives was fraught with technical difficulties and more work is needed on methodology development.<br /> <br /> <br /> Project Modification: The third multi-state trial of biocontrol agents for control of wire stem of broccoli (caused by R. solani) was cancelled due to funding restrictions and changes in resource allocation. A publication on combined data from the first and second trials is being developed.<br /> <br /> <br /> Objective 2. Examine the effect of cultural practices on soilborne pathogens and plant growth.<br /> <br /> <br /> Arkansas: Research was established to characterize changes in plant pathogen and general microbial populations in soils following a summer brassica cover crop, mustard seed meal amendment, solarization or a combination of the cover crop and solarization compared to no soil treatment prior to establishing an annual strawberry crop at two different locations in Arkansas. General microbial and suspect pathogen populations from soils were quantified by plate count methods. Additional soil samples were taken after cover crop incorporation to generate denatured gradient gel electrophoresis (DGGE) profiles for bacterial and fungal populations. Roots of strawberry plants, including the initial transplants, were also analyzed for the isolation frequency of plant pathogens. Soil treatments did affect the level of bacterial, fungal and actinomycete populations in the soil at the time of treatment and at strawberry transplant. In all cases, there was a trend for higher bacterial, fungal and actinomycete populations in brassica, brassica plus solarization and mustard seed meal amended soils compared to solarized only and control soils. Total culturable, bacterial populations were significantly higher in soils that had been planted with a brassica cover crop followed by solarization and soils receiving mustard seed meal amendments at both locations at the time of strawberry transplanting. From soil samples taken at 7 and 25 days after the brassica cover crop was incorporated into the soil, DGGE produced unique profiles of bacteria and fungi compared to that of control soils. At the time of strawberry transplant, all bacterial DGGE profiles of soils from both locations receiving different treatments were still distinct and grouped separately in dendograms, while fungal DGGE profiles were not as consistently distinct among treatments. This project has successfully proven how including soil treatments such as a brassica cover crop, solarization or mustard seed meal application as a practice in annual strawberry production can enhance the soil microflora, especially the bacterial community. Since changes could be observed in both the bacterial and fungal communities throughout the sampling times, this system has the potential to produce a soil that is more diverse and possibly reduce populations or colonization of roots by soilborne pathogens. The impacts of these shifts in the soil microflora for soilborne diseases need to be compared to chemical fumigants in soil with a history of strawberry production to examine their value in developing a sustainable strawberry production system. <br /> <br /> <br /> Iowa: Biofuels are being carefully evaluated for their efficacy to replace hydrocarbons. Pyrolysis of plant materials leaves behind biochars that may impact soil cultural practices and affect soilborne organisms and plant growth. We studied corn stover and three biochars (pyrolysis products made from corn stover) and their decomposition in soil. The percentage of carbon lost from the amendments after eight weeks was greatest for corn stover (44 wt%), followed by the two less completely pyrolyzed biochars (9 and 10%), and least (4%) from the biochar made by fast pyrolysis at 500°C. Thus, all biochars contained at least some readily bioavailable carbon that could be measured as CO2 emissions over 8 weeks. Microbial availability roughly correlated inversely with the degree of pyrolysis completeness. There was an apparent shift in microbial communities from bacteria and actinomycetes to fungi with increasing pyrolysis. This work will be submitted for publication shortly.<br /> <p>A soil biology educational website containing video of important soil organisms including fungi, ectomycorrhizae, endomycorrhizae, and their activities in the soil food web is maintained at (http://www.agron.iastate.edu/~loynachan/mov/). Two presentations were given in October at Hoover High School in Des Moines on living organisms in the soil. <br /> <br /> <br /> Louisiana: Foliar-applied biological fungicides were evaluated. Fungicide field studies evaluating Ballad Plus (Bacillus pumilus (Strain QST 2808)) for managing fungal diseases of wheat, corn, and soybean were conducted during 2010. Foliar applications were made when each crop reached a selected growth stage. Diseases were monitored if present and quantified in each plot during the growing season. Yields and test weights were collected when possible. Ballad Plus was applied at rates of 0.5 and 1.0 qt/A on wheat during growth stage F8 (flag leaf emergence). These products were applied alone or in combination with Headline fungicide (pyraclostrobin) (3.0 and 6.0 fl oz/A). Leaf rust (Puccinia triticina) severity (except for one rating), test weights, or grain yield did not differ among the non-treated wheat and solo applications of Ballad Plus. The addition of Headline fungicide resulted in less disease severity and higher yields than the non-treated. Ballad Plus applied alone at 0.5 or 1.0 qt/A or in combination with Headline at 3.0 or 6.0 fl oz/A was evaluated in field corn. Foliar applications were made to corn during the R1 growth stage (tasseling). Diseases did not develop to damaging levels in this test. Yields did not differ among treatments. Ballad Plus applied alone at 0.5 or 1.0 qt/A or in combination with Headline at 3.0 or 6.0 fl oz/A was evaluated in soybean. Foliar applications were made when soybean was in the R3 growth stage (pod initiation). Diseases did not develop to damaging levels and did not differ among treatments. Test weights and yield did not differ among any treatments.<br /> <br /> <br /> Mississippi: Mississippi is the third largest sweetpotato producer in the United States after California and North Carolina (USDA National Agricultural Statistics Service, 2009). Sweetpotato storage rots in Mississippi have increased annually since 2005 and are occurring earlier each year. Although increasing in incidence, the rot seldom damages enough of the crop to cause serious economic loss. Until 2008, the most common rots were Rhizopus and several Fusarium rots, circular spot, charcoal rot, punky rot, and Java black rot. However, during 2008-2009 growing season, an unknown rot complex dramatically impacted stored roots and coupled with other rots caused substantial economic losses. These losses would represent 1,334,313 boxes of sweetpotatoes or ca. $16,775,000 for the 2008 production year.<br /> <p>Repeated random isolations from sweetpotatoes symptomatic of the new rot and drawn from different bins of multiple growers, failed to obtain a consistent pathogen, usually resulting in one of eight different Fusarium spp., many of which are common soil inhabitants and M. phaseolina. These findings illuminated the need for baseline data of the associate root microorganisms of sweetpotato roots that could support determination of a causal agent(s), or be used to systematically assess biologically based management schemes or evaluate crop rotations. Therefore, the objectives of the initiated studies were: (1) to utilize selective media to determine the diversity and densities of fungi and bacteria associates from sweetpotato tissues, including roots and stems from key production stages in the field and storage; (2) to co-sample the same tissues and utilize molecular cloning and sequence data to compare organism(s) diversity to results from objective (1); and (3) to conduct pathogenicity screening of the putatively important bacterial and fungal isolates (ca. 100 to 150 total species expected) in sweetpotato tissues to confirm parasitism using Kochs postulates.<br /> <p>Two fields with a history of sweetpotato rot, with 1-2 years of continuous sweetpotato production were selected. Both fields were planted with Beauregard varieties. The cultural practices from bedding to harvesting followed the recommendations of Mississippi State University Extension Services and Thompson et al. (2002). Tissue sampling was divided into eight periods during sweetpotato production. The sampling periods were: (1) sampling of seed stock roots prior to bedding to include four pieces each from eyes at the distal and proximal ends of the potato, as well as eight samples drawn from the internal storage parenchyma obtained by slicing the root horizontally down the middle, (2) at six weeks after seed bed establishment [roots and attached slips were dug and tissues processed as in period 1 except tissue at the proximal end were sampled instead of the proximal eyes]; (3) 1 cm long replicate tissue pieces removed from the base, central node, and top of each slip [sampling as in period 2]; (4) roots dug prior to herbicides application(s) ca. 45 days after slip transplanting [sampling as in period 2]; (5) roots were dug immediately prior to mechanical vine removal [sampling as in period 2]; (6) except roots were dug immediately after vine removal or during harvest [sampling as in period 2]; (7) at ca. 60 days after harvest [sampling as in period 1 except mother roots were selected from four replicate storage bins per location for tissue assay]; (8) at ca. 90 days after harvest [sampling as in period 7]. Four isolation media were used to culture bacteria and fungi from the tissues sampled in each of the 8 sample periods. Nutrient glucose agar (NGA) and Kings B agar (KB) media were used for bacteria and potato dextrose agar (PDA) and water agar (WA) for fungi. Identification of bacteria and fungi used fatty acid methyl esters analyses for bacteria and morphological features for fungi. Molecular techniques will be used to confirm identification. Representative tissues from each sample are currently stored in -80°C for further processing (i.e., DNA extraction). Approximately 2400 isolates of fungi and 2500 isolates of bacteria were collected from sampling period 1-6. Samples from period 7 are currently being processed. Identification of bacteria and fungi is currently under way. Several fungal genera which tentatively have been identified included Fusarium (several species), Candida, Curvularia, Cercosporella, Phoma-like, Aspergillus, and Macrophomina. Following identification, representative isolates of each taxon (fungi and bacteria) will be stored in -80°C as sweetpotato end/tip rot study culture collections.<br /> <p>Recent research at Mississippi State University has focused on understanding the mechanisms of antifungal bacteria present in soil-borne disease systems. More than 50 bacteria obtained from the local soils of Mississippi exhibited significant antifungal activities against pathogenic fungi. Strain MS14 possesses a very broad range of antifungal activities against economically important fungi, including plant and animal pathogens. It was revealed that a set of nonribosomal peptide synthetase genes, which are harbored in a 56-kb gene cluster in the MS14 genome, are required for the antifungal activities and the antifungal compound was an oligopeptide (Gu et al. Biochem. Biophys. Res. Commun. 380:328-332). Two key regulatory genes ambR1 and ambR2 positively regulate expression of antifungal activities of strain MS14 (Gu et al. FEMS Microbiol. Lett. 297:54-60). The genes involved in modification and secretion of the oligopeptide have been characterized from the right and left boards of the gene cluster. In collaboration with Dr. James L. Smith, the chemical structure of the oligopeptide (named occidiofungin) was characterized as a glycopeptide containing eight amino aids and may target glucan biosynthesis of fungal cell walls (Lu et al. Biochemistry, 48:8812-8321). Analyses of the antifungal spectrum of activity and the minimum inhibitory concentration revealed occidiofungin is of great potential in both plant disease management and animal disease treatment. <br /> <br /> <br /> North Carolina: Wood-based mulches are used in avocado production and are being tested on Fraser fir for reduction of Phytophthora root rot, caused by Phytophthora cinnamomi Rands. Research with avocado has suggested a role of microbial cellulase enzymes in pathogen suppression, through effects on the cellulosic cell walls of Phytophthora. This work was conducted to determine whether cellulase activity could account for disease suppression in these systems. A standard curve was developed to correlate cellulase activity in mulches with concentrations of a cellulase product. Based on this curve, cellulase activity in mulch samples was equivalent to a cellulase enzyme concentration of 25 U/ml or greater of product. Sustained exposure of P. cinnamomi to cellulase at 10-50 U/ml significantly reduced sporangia production, but biomass was only reduced with concentrations over 100 U/ml. In a lupine bioassay, cellulase was applied to infested soil at 100 or 1000 U/ml with three timings. Activity diminished by 47% between one and 15 days after application. Cellulase applied at 100 U/ml two weeks before planting yielded activity of 20.08 µmol glucose equivalents per gram soil water (GE/g aq) at planting, a level equivalent to mulch samples. Activity at planting ranged from 3.35 to 48.67 µmol GE/g aq, but no treatment significantly affected disease progress.<br /> <p>Phytophthora root rot of Fraser fir, caused by several Phytophthora spp., is a severe problem in Christmas tree production. Since fungicides are not economically viable for disease management in field plantings and host resistance is not available, cultural control methods were investigated. Mulches, dairy compost, and soil pH adjustment were tested at five field sites in North Carolina. Treatments included wood chips, wood chips plus compost, or pine bark as raised beds, and compost or sulfur tilled into soil. Soil and mulch microbial populations were characterized by dilution plating and calculation of a log series diversity index, and by enzyme analyses at 5, 12, 17, and 24 months after planting. Bacterial and fungal counts, microbial activity, and cellulase activity were higher in mulch than in soil at all sites and times (P<0.01), and generally did not differ among mulch types or among soils. Treatments significantly affected disease ratings and tree survival at three of five sites, with one or more mulch treatments yielding lower disease ratings and greater survival than controls. Tree mortality at each time point varied significantly with cellulase activity in the upper root zone (P=0.005). Other biological variables did not show significant relationships with disease ratings or mortality.<br /> <br /> <br /> Ohio: A biochemical assay was developed to detect Phytophthora sojae infestation in soil. The approach was based on profiling total cellular fatty acid methyl esters (FAME) of P. sojae in pure culture. A total of 12 fatty acids (14:0, 16:0, 18:0, 16:1 É7, 18:1 É9, 18:2 É6, 18:3 É6, 20:1 É9, 20:3 É6, 20:4 É6, 22:1 É9 and 24:1 É9) were identified in the FAME profiles of P. sojae (races 1, 4 and 7) pure cultures. The predominant fatty acids in the FAME profiles are the unsaturated 18C fatty acids (18:1É9 and 18:2 É6) followed by the saturated and unsaturated 16C fatty acids (16:0 and 16:1 É7). The ratio of some of the individual fatty acids significantly changed due to growth conditions, and to a lesser extent due to pathogen race. Zoospores of P. sojae additionally contained the long-chain saturated fatty acids (20:0 and 22:0), which were not detected in the mycelium of this organism. The potential of using FAME profiles of P. sojae for detecting the pathogen in soil was evaluated by adding a known number of zoospores of P. sojae to soil. The results showed that fatty acids such as 18:1w9, 18:2w6, 20:1w9, 20:4w6 and 22:1w9 could be detected and quantified against the background levels of fatty acids present in soil. This outcome is significant because it offers the potential for a simple and rapid method for determining P. sojae infestations in soil. <br /> <p>Work was also conducted to investigate extracellular proteins belonging to class-I elicitins in Phytophthora. It is known that elicitins are involved in sterol acquisition, but very little is known about the relationship between sterols and elicitin gene expression. The objective was to determine the pattern of class-I elicitin gene expression in P. sojae, when its growth medium contains different types of sterol (fungal, plant or animal). It was discovered that the growth of P. sojae was stimulated by nanomolar concentrations of all the sterols tested. This also resulted in a differential regulation of class-I elicitin gene expression compared to controls when monitored over-time using real time Reverse Transcription Polymerase Chain Reaction (RT-PCR). Generally, class-I elicitin genes became down-regulated over time which also coincided with a reduction in elicitin biosynthesis when any of the sterols was present in the growth medium. However, kinetics of down-regulation varied as a function of sterol structure, which may be related to binding efficiencies for sterols with elicitins. Also, using Elemental Analysis-Isotopic Ratio Mass Spectrometry (EA-IRMS), it was discovered that P. sojae rapidly assimilated 15N-labeled extracellular proteins (which predominantly constitute class-I elicitins) into its mycelium. This happened when stigmasterol was added to the growth medium, suggesting that elicitins are involved in sterol sequestration by this organism. This study is the first to show that sterols regulate the expression of class-I elicitin genes in Phytophthora and provides strong evidence for the involvement of elicitins in sterol acquisition. <br /> <br /> <br /> Pennsylvania: The ecology and diversity of endophytic and soilborne fungi and bacteria are poorly understood. Verticillium dahliae causes wilt diseases in hundreds of economically important crops worldwide. Populations of this pathogen are highly structured and composed of Vegetative Compatibility Groups (VCGs) that define clonal lineages. Over sixty isolates, representing all VCGs (1A, 1B, 2A, 2B, 4A, 4B, and 6), numerous hosts and geographic origins were typed based on IGS sequences and six anonymous genomic regions containing single nucleotide polymorphisms. Results showed that IGS sequences display a very complex structure with numerous large indels with varying copy numbers between isolates. Maximum parsimony analysis indicated that certain subgroups (e.g., 1A and 1B) are indeed closely related and placed in the same clade; however, other subgroups (e.g., 4A) are related more closely to members of a different VCG (2B) than to subgroups of the same VCG (4B). MP analysis that VCG2B is polyphyletic and comprises at least three distinct phylogenetic lineages. Endospore-forming bacterial endophytes were isolated from Theobroma cacao. Cacao leaves, pods, branches, and flower cushions were removed from cacao trees escaping disease and tissue was surface sterilized, heat treated (75°C for 15 min), and plated. 69 endophytic endospore-forming bacteria were isolated and the 16S rRNA gene was sequenced to determine species identity. Isolates were further classified for characteristics of biological control activity. Sixteen isolates were chitinolytic and in antagonism studies against cacao pathogens, 42% were antagonistic to Moniliophthora roreri, 33% to M. perniciosa, and 49% to Phytophthora capsici. 25% of isolates inhibited the growth of both Moniliophthora spp., while 22% of isolates inhibited the growth of all three pathogens. Isolates that were chitinolytic and tested negative on Bacillus cereus (Bc) agar were tested in planta. All 14 isolates were capable of endophytic colonization; while 8 isolates significantly inhibited P. capsici lesion formation in detached leaf assays. ARISA was used to estimate the diversity of total bacteria and bacilli within cacao leaves. Inundative application of a single bacterial species to cacao leaves did not impact native bacterial communities. Bc is a common inhabitant of soil as well as the endophytic portions of plant tissue. Although common in the environment, some isolates cause foodborne illness through the production of enterotoxins. A collection of endophytic Bc isolates were obtained from several crops including apple, cacao, tomato, and potato. The presence of several genes for enterotoxins was tested by PCR. Of the 35 endophytic Bc isolates, the majority had one or more of the genes required for enterotoxin production and 14% of isolates tested lacked any enterotoxin genes. The endophytic host had no impact on the presence of enterotoxin genes, as they were detected in isolates from all tested plants. The enterotoxin genes from endophytic isolates were sequenced and compared to Bc isolates known to cause foodborne illness. While Maria del Mar Jimenez-Gasco and Paul A. Backman are the principal investigators of this project, other participants included Anissa Poleatewich (former graduate student), Rachel L. Melnick (former graduate student), and Glenna Malcolm (postdoctoral researcher).<br /> <br /> <br /> South Carolina: The rootstocks Emphasis, Macis, and WMXP 3945 (Lagenaria siceraria), Shintosa Camel and Strong Tosa (Cucurbita moschata x Cucurbita maxima), and Ojakkyo (Citrullus lanatus var. citroides) were evaluated in a field experiment. The rootstocks were grafted with seedless watermelon Tri-X 313 (Citrullus lanatus var. lanatus); Tri-X 313 not grafted or grafted to itself served as the controls. Plants were transplanted to a field in which the transplanting holes had been infested with a 1:1 mixture of races 1 and 2 of Fusarium oxysporum f. sp. niveum, the causal agent of Fusarium wilt of watermelon. Wilt incidence and plant death was highest in the self-grafted Tri-X 313. Ojakkyo citron had more wilted plants than plants grafted to Lagenaria rootstocks. Yield (weight and number of fruit) were greater with Lagenaria rootstocks than with Tri-X 313 (mean of not grafted and self-grafted) or with Cucurbita rootstocks. <br /> <p>Oilseed radish (Raphanus sativus), mustard (Brassica juncea cv. Pacific Gold), and winter rapeseed (B. napus cv. Dwarf Essex) were grown in the spring for seven weeks before incorporation as biofumigation cover crops. Plots were covered with black virtually impenetrable film (VIF) after incorporation in late May. Control treatments were fallow with and without VIF. Five weeks after incorporation, populations of Pythium did not differ among any treatments. Populations of R. solani were higher in fallow without VIF than in all mulched treatments, indicating that the VIF increased soil temperatures enough to reduce populations of R. solani. Pepper plants transplanted into the plots were stunted due to Pythium infection of the roots in all plots but particularly in the fallow plots without VIF. Yields of pepper over 10 weeks were greater in the winter rapeseed, mustard, and fallow plus VIF treatments than in the fallow plots without VIF. The lack of an effect of the cover crops may be due to the relatively low biomass obtained with spring-planted brassica cover crops in coastal South Carolina.<br /> <br /> <br /> Tennessee: Interactions between two fertilizers (muriate of potash and sulfate of potash), two biological control agents (GB03 and MBI600), and seed treatment fungicides were evaluated for control of snap bean seedling diseases in an April-planted field test. Fertilizers were applied at 135 kg K2O/ha. The biological agents were evaluated both with and without supplemental seed treatment fungicides. Over 4 cm of rain were received as the snap bean seedlings were beginning to emerge. Another 36 cm of rain were received during the next 10 days. Under these very disease-conducive conditions, GB03 and MBI600 failed to increase snap bean stand or yield without the use of seed treatment fungicides. MBI600 reduced plant stand and yield when combined with captan + metalaxyl and increased plant stand and yield when combined with fludioxonil + mefenoxam. Highest plant stands and yield were achieved with the use of trifloxystrobin + metalaxyl seed treatment fungicides without a supplemental biological agent. Use of sulfate of potash instead of muriate of potash increased plant stands by over 12% and increased snap bean yield by over 50%. <br /> <p>Potash fertilizer form, fertilizer application timing, and seed treatment materials were evaluated for control of charcoal rot of soybean (caused by M. phaseolina) in both May-planted and June-planted field tests. Potash fertilizer form (sulfate of potash versus muriate of potash) and application timing (March or at planting) had no effect on soybean stand or yield in the 2010 tests. Adding biological agents (MBI600 or GB34) to a soybean seed treatment of fludioxonil + mefenoxam had no effect on plant stand or yield. <br /> <p>A laboratory study on the effects of chloride salts on the micro-partitioning of root Ca was initiated using energy-dispersive analysis of x-rays. After 136 separate EDX analyses of root tissues from six different plants, detectable Ca levels in the outer cells of taproots were notably lower when plants were grown in soil treated with MgCl2 than when grown in untreated soil. There were no detectable treatment effects on lateral roots. <br /> <br /> <br /> Objective 3. Examine the genetic diversity of Rhizoctonia solani between natural ecosystems and agricultural ecosystems. <br /> <br /> <br /> Arkansas: R. solani populations were examined by the protocol developed by the regional project which included two selective media, ethanol-potassium nitrate and Ko and Hora, and two sampling techniques, multiple-pellet soil sampler and toothpick method. In 2010, soils used in evaluations in Arkansas were one soybean field in rotation with rice and a prairie soil in Prairie County, three cotton fields in Ashley and Poinsett Counties, and a vegetable production field in Sebastian County. The diversity of Rhizoctonia species in these cropping systems included R .solani AGs 11, 1-1A, 4 and 2-1, R. oryzae and bi-nucleate Rhizoctonia species. Ethanol or Ko and Hora media both allowed the isolation of R. solani. However, Ko and Hora medium had more problems with fungal contaminants. Anastomosis groups detected were similar for the two selective media, two techniques, and seedlings. The toothpick method was an effective method for the recovery of R. solani from soils with diverse cropping histories because no special tools and few units were needed to detect the population. By comparing the soil pellet weight assayed to toothpick isolation frequency, the volume of soil assayed by the toothpick method can be estimated. <br /> <br /> <br /> Kentucky: Samples were collected from three locations (histories of vegetable, tobacco, and soybean production) in May and June, 2010 and were assayed using the toothpick method chosen by the S1028 group. Isolates have been recovered and stored for analysis. They will be sent to C. Rothrock for complete identification.<br /> <br /> <br /> Louisiana: A study was conducted at the Macon Ridge Research Station near Winnsboro, LA to assess the diversity of Rhizoctonia sp. in corn, cotton, grain sorghum, and soybean fields. A standardized protocol developed by the S1028 group was used. Soil samples were taken from fields (two per crop) and placed in plastic containers. Moisture was supplemented to soil to saturation and toothpicks were placed in soil and left for a predetermined time according to protocol. Toothpicks were plated onto ethanol-potassium nitrate medium. Plates were monitored for growth of Rhizoctonia at 24, 48 and 72 hours. Fungal growth was minimal at 24 and 48 hours; however, colonies were counted 72 hours after toothpicks were placed on plates. There were 142, 158, 233, and 113 colonies counted for corn, cotton, grain sorghum, and soybean, respectively.<br /> <br /> <br /> Oklahoma: DNA fingerprinting protocols (AFLP, ISSR, SSR) were standardized for analysis of R. solani populations. Peanut populations of the Sclerotinia minor from Oklahoma were characterized using AFLP and ISSR. Multiple strains of Pythium irregulare species complex from diverse locations and hosts were characterized. Phylogenetic analyses of this clade showed evidence that multiple cryptic species exist. The genetic fingerprinting of multiple isolates of Phymatotrichopsis omnivora from Oklahoma, New Mexico, Texas and Arizona was completed. The genetic fingerprinting was completed and population genetic analyses performed of multiple isolates of Fusarium oxysporum f. sp. palmarum in collaboration with Monica Elliot, University of Florida. It was demonstrated that sublethal doses of two fungicides, Cyazofamid and Propamocarb, as well as ethanol induce stimulation in Pythium aphanidermatum and that hormesis is involved in the stimulation processes. It was also demonstrated that low doses of ethanol induce stimulation in Rhizoctonia zeae and that hormesis is involved in the stimulation process, while Propiconazole at sub-lethal doses do not have hormetic effects on either R. zeae or on R. solani. Diagnostic primers were designed for multiple Pythium, Rhizoctonia, and Sclerotinia spp.<br /> <br /> <br /> Tennessee: Previous studies had shown that populations of Rhizoctonia solani were greatest in soil with a current crop of fall-planted broccoli, followed by fallow soil with a previous spring broccoli crop, and lowest in non-agricultural soil; detection was based on incubation of a toothpick bait in soil, and then plating onto Ko and Hora culture medium. To support the results of these studies, and to determine if populations of Rhizoctonia detected were pathogenic, disease assays with broccoli seedlings as the bait plant, were conducted. In the disease assays, the incidence of damping-off of broccoli seedlings was 0% in the wooded non-agricultural soil, 40% in soil with a previous crop of spring broccoli, and 56% in the soil that was currently cropped to fall broccoli. The rates of damping-off were consistent with the population levels of R. solani detected in the three soils. Rhizoctonia solani was re-isolated from all damped-off broccoli seedlings. This confirmed previous studies on detection methods and microbiological culture media for identification of pathogenic R. solani from soil. <br /> <br /> <br /> Grant Funding Relevant to Objectives<br /> <br /> <br /> Canaday, C. H. Elemental Analysis of Tap Root Tissues Using Energy-Dispersive X-ray (EDX) Analysis. University of Tennessee Professional Development Award, $4,955. <br /> <br /> <br /> Cox, M. and Rothrock, C. S. Microbial changes associated with use of brassica cover crops compared to traditional production systems for strawberry. Graduate student Project, Southern Region SARE Program, $9,971.<br /> <br /> <br /> Donald, P., Biggerstaff, J., and Canaday, C. Advisory Program for Management of Soybean Cyst Nematode. Tennessee Soybean Promotion Board, $46,198.<br /> <br /> <br /> Ivors, K. and Benson, D.M. Evaluating integrated strategies for management of Phytophthora root rot on Fraser fir Christmas trees in North Carolina. NC Agricultural Foundation. $21,073.<br /> <br /> <br /> Mengistu, A., Arelli, P., Canaday, C. Screening of soybean varieties, breeding lines for charcoal rot, frogeye leaf spot and Phomopsis seed decay resistance, Tennessee Soybean Promotion Board, $30,000.<br /> <br /> <br /> Seebold, K.W. and Ji, P.J. 2010. Managing Phytophthora capsici on Pepper and Summer Squash with Combinations of Bioten and Conventional Fungicides. IR-4 Biopesticide Program, $20,000.

Publications

Refereed.<br /> <br /> <br /> Bartz, F. E., M. A. Cubeta, T. Toda, S. Naito, and K. Ivors, 2010. An in planta method for assessing the role of basidiospores in Rhizoctonia foliar disease of tomato. Plant Disease 94:515-520. <br /> <br /> <br /> Berbegal, M., A. Ortega, M. M. Jimenez-Gasco, C. Olivares-Garcia, R. M. Jimenez-Diaz and J. Armengol. 2010. Genetic diversity and host range of Verticillium dahliae isolates from artichoke and other vegetable crops in eastern-central Spain. Plant Disease 94: 396-404.<br /> <br /> <br /> Canaday, C.H. and A.F. Schmitthenner, 2010. Effects of chloride and ammonium salts on the incidence of Phytophthora root and stem rot of soybean. Plant Dis. 94:758-765. <br /> <br /> <br /> Gwinn, K.D., B.H. Ownley, S.E. Greene, M.M. Clark, C.L. Taylor, T.N. Springfield, D.J. Trently, J.F. Green, A. Reed, and S.L. Hamilton. 2010. Role of essential oils in control of Rhizoctonia damping-off in tomato with bioactive Monarda herbage. Phytopathology 100:493-501.<br /> <br /> <br /> Gu, G., L. Smith, N. Wang, H. Wang, and S.-E. Lu., 2009. Biosynthesis of an antifungal oligopeptide in Burkholderia contaminans strain MS14. Biochemical and Biophysical Research Communications. 380: 328-332.<br /> <br /> <br /> Gu, G., N. Wang, N. Cahney, L. Smith, and S.-E. Lu. 2009. AmbR1 is a key transcriptional regulator for antifungal activity of Burkholderia contaminans strain MS14. FEMS Microbiology Letter. 297:54-60.<br /> <br /> <br /> Jangid, K., M. A. Williams, A. J. Franzluebbers, J. M. Blair, D. C. Coleman, and W. B. Whitman, 2010. Development of soil microbial communities during tallgrass prairie restoration. Soil Biology & Biochemistry 42:302-312<br /> <br /> <br /> Kaye, A.C., Moyer, J.W., Parks, E.J., Carbone, I., and Cubeta, M.A., 2011. Population genetic<br /> analysis of Tomato spotted wilt virus on peanut in North Carolina and Virginia. Phytopathology 101:147-153.<br /> <br /> <br /> Keinath, A. P., Hassell, R. L., Everts, K. L., and Zhou, X.-G. 2010. Cover crops of hybrid common vetch reduce Fusarium wilt of seedless watermelon in the eastern United States. Online. Plant Health Progress doi:10.1094/PHP-2010-0914-01-RS.<br /> <br /> <br /> Kim, D.-G, T. M. Isenhart, T. B. Parkin, R. C. Schultz, and T. E. Loynachan. 2010. Methane flux in cropland and adjacent riparian buffers with different vegetation covers. J. Environ. Qual. 39:97-105.<br /> <br /> <br /> Lu, S.-E., J. Novak, F. W. Austin, G. Gu, D. Ellis, M. Kirk, S. Wilson-Stanford, M. Tonelli, and J. L. Smith. 2009. Occidiofungin, a unique antifungal glycopeptide produced by a Strain of Burkholderia contaminans. Biochemistry 48: 8312-8321. <br /> <br /> <br /> Njoroge, S. M. C., Toler, J. E. and Keinath, A. P. 2010. Soil solarization on populations of Pythium spp., fluorescent Pseudomonas, and damping-off of broccoli and cucumber. Int. J. Veg. Sci. 16:15-31.<br /> <br /> <br /> Sullivan, M. J., Parks, E. J., Cubeta, M. A., Gallup, C. A., Moyer, J.W., and Shew, H.D. 2010. An assessment of genetic diversity from a field population of Phytophthora nicotianae with a changing race structure. Plant Disease 94:455-460.<br /> <br /> <br /> Abstracts<br /> <br /> <br /> Berbegal, M., C. D. Garzon, A. Ortega, J. Armengol, R. M. Jimenez-Diaz and M. M. Jimenez-Gasco. 2009. Development and application of new molecular markers for the analysis of genetic diversity in Verticillium dahliae populations. In 10th International Verticillium Symposium Book of Abstracts, November 16-20, 2009, Corfu, Greece. p. 29.<br /> <br /> <br /> Jimenez-Gasco, M. M., M. Berbegal, G. M. Malcolm, J. Armengol and R. M. Jimenez-Diaz. 2009. New insights on the phylogenetic relationships of vegetative compatibility groups in Verticillium dahliae. In 10th International Verticillium Symposium Book of Abstracts, November 16-20, 2009, Corfu, Greece. p. 28.<br /> <br /> <br /> Spurlock, T. N., Rothrock, C. S., and Monfort, W. S. 2010. Evaluation of selective media and selective chemicals on the isolation of Rhizotonia spp. from soil. (Abstr.) Phytopathology 100(6) Supplement S121. <br /> <br /> <br /> Book Chapters.<br /> <br /> <br /> Benson, D.M., and B.H. Ownley. 2010. Exercícios de laboratório com fitopatógenos do solo. Pages 255-270 in: Fitopatologia: Conceitos e Exercícios de Laboratório. 2a Edição. R. Trigiano, M. Windham, and A. Windham, eds., Taylor and Francis, CRC Press, Boca Rotan, FL. (Translation in Portuguese).<br /> <br /> <br /> Ownley, B.H., and D.M. Benson. 2010. Fitopatógenos do solo. Pages 237-254 in: Fitopatologia: Conceitos e Exercícios de Laboratório. 2a Edição. R. Trigiano, M. Windham, and A. Windham, eds., Taylor and Francis, CRC Press, Boca Rotan, FL. (Translation in Portuguese).<br /> <br /> <br /> Ownley, B.H., and M. Windham. 2010. Controle biológico de fitopatógenos (Revised). Pages 447-460 in: Fitopatologia: Conceitos e Exercícios de Laboratório. 2a Edição. R. Trigiano, M. Windham, and A. Windham, eds., Taylor and Francis, CRC Press, Boca Rotan, FL. (Translation in Portuguese).<br /> <br /> <br /> Ownley, B.H., K.D. Gwinn, and F.E. Vega. 2010. Fungal entomopathogens with activity against plant pathogens: ecology and evolution. Pages 113-128 in: The Ecology of Fungal Entomopathogens. H.E. Roy, F.E. Vega, D. Chandler, M.S. Goettel, J. Pell, and E. Wajnberg, eds. Springer-Verlag, NY.<br /> <br /> <br /> Other Publications.<br /> <br /> <br /> Canaday, C.H., 2010, Effects of seed treatments, potash formulation and application time on plant height, stand loss, and yield, 2009. Plant Disease Management Reports (online), Report 4:ST012. doi:10.1094/PDMR04.<br /> <br /> <br /> Canaday, C.H., 2010, Evaluation of seed treatments, in-furrow sprays, and biological agents for control of seedling diseases of snap bean, 2009. Plant Disease Management Reports (online), Report 4:V084. doi:10.1094/PDMR04.<br /> <br /> <br /> Francis, R., and Keinath, A. 2010. Biofungicides and chemicals for managing diseases in organic vegetable production. Clemson Cooperative Extension Information Leaflet 88.<br /> <br /> <br /> Malcolm, G. M., and M. M. Jimenez-Gasco. 2010. Associations of Verticillium dahliae with host and non-host plants in agricultural fields. In 95th Ecological Society of America Annual Meeting, August 1-6, 2010, Pittsburgh, PA. p COS4.<br /> <br /> <br /> Yousef, L.F. 2010. Class-I elicitins in relation to sterol acquisition and lipid profiling of Phytophthora sojae. PhD Dissertation. The Ohio State University, Columbus, OH.<br />

Impact Statements

1. A strawberry production system that used a summer brassica cover crop (mustard seed meal) or a combination of the cover crop and solarization was successful in changing soil microbial populations in both the bacterial and fungal communities. This system has the potential to produce a soil that is more diverse and possibly reduce populations or colonization of roots by soilborne pathogens.
2. A efficient effective protocol developed for the assay of Rhizoctonia solani from soils should allow better detection of the pathogen in soil and the examination of the diversity of this important pathogen across different native and agricultural systems increasing our understanding of the role of cropping history and transport of the pathogen in disease development and diversity.
3. Results from tests will be used to develop a database for fungicide recommendations in wheat, corn, and soybean. Based on preliminary results, Ballad Plus did not result in an economical benefit in wheat, corn, or soybean. However, more testing will be necessary to develop a stronger database.
4. Understanding the genetic diversity of populations of Rhizoctonia solani in Oklahoma agricultural soils compared to those in natural soils, can contribute information about the origin of inoculum that can be useful for disease management and prevention. Microsatellite markers and highly sensitive PCR and real-time PCR specific for anastomosis groups are being developed and evaluated on Oklahoma isolates and on collections of isolates obtained through collaborations with Clemson University and the University of Arkansas.
5. Lagenaria rootstocks appear to be more promising than interspecific hybrid Cucurbita or citron rootstocks for seedless watermelon grown in fields affected with Fusarium wilt. Brassica cover crops should be planted in the fall in coastal South Carolina to obtain sufficient biomass to reduce populations of soilborne plant pathogens.
6. In terms of planted acreage, snap beans are Tennessees number one vegetable crop. Farm receipts for this crop typically total over $9,000,000 per year. Instead of using untreated seed and muriate of potash, growers could potentially double their snap bean yield and gross returns if they used a seed treatment of trifloxystrobin + metalaxyl coupled with use of sulfate of potash.
7. The forecasted value of U.S farm cash receipts for vegetable crops in 2010 is more than $21 billion, with $96 million projected in Tennessee. Annual losses due to soilborne plant pathogens are estimated to be as high as 10% and are greatest in transplant production. Soilborne pathogens like Rhizoctonia solani have a wide host range, including many vegetable crops with no genetic resistance. Development and improvement of methods for detection of Rhizoctonia prior to planting will provide information that will impact crop management decisions for vegetable production.
8. A commercial formulation of Trichoderma viride and T. harzianum, applied to soil by drip irrigation in 2009, was shown to suppress the incidence of Phytophthora blight on summer squash. Although not as effective as commercially available fungicides, using the biocontrol agents in conjunction with commercial fungicides may permit fewer applications or allow for reduced rates of the fungicide products.

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Report Information

Participants

Backman, Paul (pbackman@psu.edu) - Pennsylvania State University;
Baird, Richard (RBaird@plantpath.msstate.edu) - Mississippi State University;
Benson, Mike (mike_benson@ncsu.edu) - North Carolina State University;
Broders, Kirk (kirk.broders@unh.edu ) - University of New Hampshire;
Canaday, Craig (ccanaday@utk.edu) - University of Tennessee;
Cubeta, Marc (marc_cubeta@ncsu.edu) - North Carolina State University;
Dick, Warren (dick.5@osu.edu) - Ohio State University;
Elliott, Monica (melliott@ufl.edu) - University of Florida;
Garzon, Carla Domenica (carla.garzon@okstate.edu) - Oklahoma State University;
Gentry, Terry (tgentry@ag.tamu.edu ) - Texas AgriLife Research;
Jimenez Gasco, Maria (jimenez-gasco@psu.edu) - Pennsylvania State University;
Keinath, Tony (tknth@clemson.edu) - Clemson University;
Loynachan, Thomas (teloynac@iastate.edu) - Iowa State University;
Lu, Shien (sl332@msstate.edu ) - Mississippi State University;
Ownley, Bonnie (bownley@utk.edu) - University of Tennessee;
Padgett, Boyd (bpadgett@agcenter.lsu.edu) - Louisiana State University;
Rothrock, Craig (Rothrock@uark.edu) - University of Arkansas;
Seebold, Kenneth (kwseebold@uky.edu ) - University of Kentucky;
Williams, Mark (mw452@msstate.edu) - Mississippi State University;

Brief Summary of Minutes

Minutes of the Annual Meeting of the Technical Committee of MRP Project S-1028

Nov. 4 to 5, 2011, Oklahoma City, Oklahoma


Attendees: C. Garzon (OK), C. Canaday (TN), C. Rothrock (AR), K. Broders (NH), H. Scherm, (Adm. Advisor). Other participants Luisa Santamaria (OR)


The meeting was held at the Hilton Garden Inn Quail Springs in Oklahoma City, Oklahoma. The meeting was called to order by Carla Garzon, Chair, at approximately 8:30 A.M. Carla welcome the participants. Harald Scherm introduced himself as the new administrative advisor and talked about multi-state projects and the status of the current project. He also reviewed the steps involved in rewriting the project and his feeling regarding a multi-state projects and information exchange groups.


The members discussed their interests in developing a new project and the focus of the project. The general consensus of the participants was that there were advantages to developing a collaborative research project and that we should explore a new multi-state project. The project would focus more on specific pathogens. The genera suggested were Pythium, Rhizoctonia, and/or Verticillium. Areas of research suggested were diversity, IPM, metagenomics, and development of methodologies for sampling, detection, statistical data analysis, and education.


Presentations relating to the projects research objectives and current research of participants were then started.


Craig Rothrock presented research on Brassica cover crops as part of objective 2 (examining the effect of cultural practices on soilborne pathogens and plant growth). Cover crop establishment in the winter of 2010 was poor and results did not show much benefit in growth or disease suppression. The greatest effects on pathogen suppression in past years had been on nematodes. In addition, he presented evidence of shifts in microbial populations from the use of Brassica cover crops using DGGE analysis. Research examining the genetic diversity of Rhizoctonia solani between natural ecosystems and agricultural ecosystems (objective 3) included surveys in 2011 to collect samples from diverse crops in the Southeast. Isolates are being characterized. The sampling protocol involves baiting populations of Rhizoctonia from soil using toothpick baits and selective media, as well as seedling isolations


Craig Canaday presented data on the role of chloride in enhancing Phytopthora sojae on soybean and recent parallel research on snap bean production and potassium fertilizers. He showed significant yield response from the use of K2SO4 versus KCL. He also presented information for soybean and snap bean on mechanisms of plant response from Cl amendments on Ca micropartitioning.


Carla Garzon gave an overview of her research on diversity of Pythium spp. and species characterization (objective 3). Additional research presentations involved graduate students in her laboratory. Andres Espindola presented information on development of specific primers for identification of Sclerotinia spp. and a novel massive parallel sequencing based diagnostic tool. Francisco Flores presented research on chemical hormesis caused by subinhibitory levels of fungicides and methods of modeling hormetic effects.


Kirk Broders presented research related to the project on diversity (objective 3). He presented a review of a large project characterizing Pythium spp. on soybean. In addition to characterizing species associated with soybean he presented information collected to examine the role of soil physical or chemical factors on importance of Pythium seed and root rot in fields and species composition. He also presented information on his recent research on the diversity of Butternut pathogen, Ophiognomonias clavigignenti-juglandacearum. He gave an overview of his current research interests.


Luisa Santamaria introduced her research with the nursery industry in Oregon on Verticillium dahliae. Her research has focused on sources of inoculum of the pathogen as influenced by cultural practices used in the industry. In addition, she discussed her education efforts with workers in the horticultural industry to increase awareness of plant pathology and improve management practices.


Business Meeting. C. Garzon, chair, called the business meeting to order following a break after the presentations by participants.


The members present discussed a location for next year's meeting and Carla volunteered to host the group in Oklahoma City again in 2012. Participants agreed that the location was convenient and with Carla's role in rewriting the project the site would be best. Craig Rothrock will become chairman for next year's meeting. Kirk Broders was nominated for secretary for 2012, and the nomination was seconded and passed unanimously. The writing committee for a new project will consist of Carla Garzon and Kirk Broders and will be chaired by Carla Garzon. Research objectives will be identified by present and potential members of the project. The committee with then involve participants in writing the new project. A possible timeline for activities to receive approval was discussed. Craig Canaday put forth a resolution thanking Carla for organizing the meeting and her efforts as host during the meeting. The resolution was endorsed by all present. The meeting was adjourned at 5:00 p.m.


Respectfully submitted,
C. Rothrock, Secretary

Accomplishments

Objective 1. Examine commercial and non-commercial biocontrol agents for use as seed treatments, in-furrow treatments or as potting mix amendments.<br /> <br /> Kentucky: Biological control of Phytophthora Blight of Summer Squash with Trichoderma was studied. A trial was conducted in the summer of 2011 to evaluate Tenet, a commercial formulation containing Trichoderma harzianum and T. viride, for suppression of Phytophthora blight, caused by Phytophthora capsici, on summer squash when applied to seedlings prior to transplanting or to soil via drip irrigation, alone or in conjunction with fungicide programs. Earlier results showed that Tenet-containing treatments reduced the severity of Phytophthora blight by 25% over the untreated control; however, results from the 2011 study differed. Tenet, when applied to squash seedlings one week prior to transplanting, reduced overall severity of Phytopthora blight compared with the untreated control. Treatments applied in the field had no effect  a departure from the trial conducted in 2009.<br /> <br /> <br /> Mississippi: Recent research in the Plant Bacteriology laboratory of Mississippi State University continued understanding the mechanisms of antifungal bacteria present in soil-borne disease systems. Further characterization was conducted to understand functions of the genes in the 56-kb occ gene cluster that is essential for production of occidiofungin production by Burkholderia contaminans strain MS14. A biosynthesis pathway was proposed based on genetic and biochemical analysis. Occidiofungin was evaluated regarding its stability and efficacy as a antifungal compound. Interestingly, disruption of the ocfC gene in the ocf gene cluster by the insertion of the nptII cassette resulted in xylose-free occidiofungin production and reduction of antifungal activity against the indicator fungus Geotrichum candidum. However, the antifungal activity of the xylose-free occidiofungin was significantly increased against Candida species as compared with the wild-type occidiofungin. These results are significant for development of agricultural biofungicides and medical parametrical drugs. In addition, more than 60 bacteria obtained from Mississippi soils exhibited significant antifungal and/or antibacterial activities against pathogenic fungi and bacteria. The mechanisms of antifungal and antibacterial activities of the isolates are under investigation.<br /> <br /> <br /> Oklahoma: A study of bacterial communities present in Andean soils from Ecuador suppressive to Phytophthora infestans was conducted. Serial dilutions of soils were performed and plated on differential and selective media following standard protocols. Approximately 3,000 bacterial strains of Bacillus, Pseudomonas, and Actinobacteria (including Streptomyces) were isolated, evaluated for their ability to inhibit P. infestans and Rhizoctonia solani. Strains with stronger inhibitory effects were selected for further characterization, and 200-300 strains of each bacterial group were sequenced (16S rDNA). Currently, the microbial communities in these soils are under further examination using metagenomic methods. Finally, analyses of the physical and chemical properties of the soils are being assessed, since the soils studied preserved suppressive qualities (about 30% of the observed suppression) after removal of microbial populations by tyndallization at 80ºC. <br /> <br /> <br /> South Carolina: A new commercial formulation of Bacillus subtilis (Serenade Soil) was evaluated for control of root diseases on cabbage and spinach. Two field experiments are in progress in which three rates of Serenade Soil are compared with water-treated and a fungicide-treated controls.<br /> <br /> <br /> Objective 2. Examine the effect of cultural practices on soilborne pathogens and plant growth.<br /> <br /> <br /> Arkansas: Research was repeated to characterize changes in plant pathogen and general microbial populations in soils following a summer Brassica cover crop, mustard seed meal amendment, solarization or a combination of the cover crop and solarization, compared with no soil treatment prior to establishing an annual strawberry crop at two locations in Arkansas. General microbial and suspect pathogen populations from soils were quantified by plate count methods. Additional soil samples were taken after cover crop incorporation to generate denatured gradient gel electrophoresis (DGGE) profiles for bacterial (including actinobacteria) and fungal populations. Soil treatments affected the level of bacterial and fungal populations in the soil at the time of treatment and at strawberry transplant. In all cases, there was a trend for higher bacterial and fungal populations in Brassica, Brassica plus solarization, and mustard seed meal-amended soils compared with solarized only and control soils. Total culturable bacterial populations were significantly higher in soils that had been planted with a Brassica cover crop followed by solarization and soils receiving mustard seed meal amendments at both locations at the time of strawberry transplanting. DGGE produced unique profiles of bacteria and fungi compared with that of control soils. Bacteria profiles were still present at the time of strawberry transplanting. This project has successfully proven how including soil treatments such as a Brassica cover crop, solarization or mustard seed meal application as a practice in annual strawberry production can enhance the soil microflora, especially the bacterial community. Cover crop establishment of Brassica crops in the winter of 2010 was poor and results did not show much benefit in growth or disease suppression for the subsequent summer cotton crop. The greatest effects on pathogen suppression in past years for the subsequent cotton crops have been on nematodes.<br /> <br /> <br /> Mississippi: The Sweet Potato Rot research project is a survey comparing microbial flora between damaged and control roots to identify possible pathogen or other cause such as specific management practices. The research is divided into three main areas of study including: 1) sampling of sweet potato plant and root tissues from fields with a history of the rot complex for culturing of fungal and bacterial strains present in plant and root tissues; 2) identification of each fungal and bacterial isolate based on both morphological characteristics and gene fragment (ITS, 16s rDNA, EF1-alpha) sequences; and 3) determining pathogenicity of fungal and bacterial isolates on storage root tissue in the laboratory and on intact plants in the greenhouse under field conditions. We hope to determine which of the pathogenic isolates are capable of infecting a plant under field conditions and, using this information, work with collaborating extension research groups to improve sweet potato resistance to the identified pathogen(s). Roots and stem tissues have been sampled throughout the year, beginning with seed stock, at the time slips are transplanted into the field, pre- and post-harvest, and during storage. The study has been repeated over two years, with the second year of sampling nearing completion. To date, over 4,500 fungal isolates and over 4,600 bacterial isolates have been cultured, and many have been identified using morphological techniques for fungi and molecular techniques for bacteria. The bacterial community is primarily composed of members of Bacillus spp., Paenibacillus spp., and Stenotrophomonas maltophilia. Predominant species of fungi include Fusarium spp., Macrophomina spp., Botryodiplodia theobromae, and Aspergillus niger. A portion of these isolates are being tested for pathogenic characteristics on sweet potato storage root tissue, and several potential fungal pathogens have been tentatively identified. Between management period 7 and 8, when end-rot symptoms become visible, the distribution and total number of fungal species changed significantly. Botryodiplodia theobromae, Fusarium spp. and Macrophomina spp. increased in number in the Site 1 field, while the dominant Aspergillus niger from the previous sampling period was not detected. Storage roots taken from the Site 2 field showed an increased occurrence of Fusarium spp. at management period 8. We also detected a greater variety of isolates in samples from management period 8 (Site 1 n=36; Site 2 n=25) than we did in samples from management period 7 (Site 1 n=16; Site 2 n=15). Fungal isolates were found in all tissue types that were sampled, and there was no statistical difference in the frequency of isolate occurrence between tissue types (parenchyma, stem end, root end). Currently, pathogenic trials are underway in the greenhouse to determine the effects of individual fungal pathogens on transplanted sweet potato slips.<br /> <br /> <br /> North Carolina: Phytophthora root rot of Fraser fir, caused by several Phytophthora spp., is a severe problem in Christmas tree production. Since fungicides are not economically viable for disease management in field plantings and host resistance is not available, cultural control methods were investigated. Mulches, dairy compost, and soil pH adjustment were tested at five field sites in North Carolina. Treatments included wood chips, wood chips plus compost, or pine bark as raised beds, and compost or sulfur tilled into soil. Soil and mulch microbial populations were characterized by dilution-plating and calculation of a log series diversity index and by enzyme analyses at 5, 12, 17, and 24 months after planting. Bacterial and fungal counts, and microbial activity were higher in mulch than in soil at all sites and times (P<0.01), and generally did not differ among mulch types or among soils. Treatments significantly affected disease ratings and tree survival at three of five sites, with one or more mulch treatments yielding lower disease ratings and greater survival than controls. Tree mortality at each time point varied significantly with cellulase activity in the upper root zone (P=0.005). Other biological variables did not show significant relationships with disease ratings or mortality. Wood-based mulches were tested on Fraser fir for reduction of Phytophthora root rot to determine whether cellulase activity could account for disease suppression in mulch systems. A standard curve was developed to correlate cellulase activity in mulches with concentrations of a cellulase product. Based on this curve, cellulase activity in mulch samples was equivalent to a cellulase enzyme concentration of 25 U ml-1 or greater of product. Sustained exposure of P. cinnamomi to cellulase at 10-50 U ml-1 significantly reduced sporangia production, but biomass was only reduced with concentrations over 100 U ml-1. In a lupine bioassay, cellulase was applied to infested soil at 100 or 1000 U ml-1 with three timings. Activity diminished by 47% between one and 15 days after application. Cellulase applied at 100 U ml-1 two weeks before planting yielded activity of 20.08 µmol glucose equivalents per gram soil water (GE g-1 aq) at planting, a level equivalent to mulch samples. Activity at planting ranged from 3.35 to 48.67 µmol GE g-1 aq, but no treatment significantly affected disease progress.<br /> <br /> <br /> South Carolina: Two research projects were conducted. The objective of the first project was to evaluate the susceptibility of Lagenaria, Cucurbita and Citrullus rootstocks to Fusarium wilt of watermelon.The rootstocks Emphasis, Macis, and WMXP 3945 (Lagenaria siceraria), Shintosa Camel and Strong Tosa (Cucurbita moschata x Cucurbita maxima), and Ojakkyo (Citrullus lanatus var. citroides) were evaluated in a field experiment. The rootstocks were grafted with seedless watermelon Tri-X 313 (Citrullus lanatus var. lanatus). Tri-X 313 not grafted or grafted to itself served as controls. Plants were transplanted to a field naturally infested with a mixture of races 1 and 2 of Fusarium oxysporum f. sp. niveum, the causal agent of Fusarium wilt of watermelon. Wilt incidence was highest in the nongrafted Tri-X 313 and moderate in the self-grafted Tri-X 313 and those grafted onto Ojakkyo citron. All five other rootstocks had less than 2% wilted plants and did not differ from each other, but differed from the other three treatments. Fruit weight was greater with Lagenaria and Cucurbita rootstocks than with Tri-X 313 (mean of not grafted and self-grafted). Lagenaria rootstocks also increased fruit number. In the second project oilseed radish (Raphanus sativus), mustard (Brassica juncea cv. Pacific Gold), and winter rapeseed (B. napus cv. Dwarf Essex) were sown in fall 2010 and spring 2011 before incorporation as biofumigation cover crops. After incorporation in February and April, plots were covered with white-on-black virtually impenetrable film (VIF). Control treatments were fallow with and without VIF. Isothiocyanate concentrations (ITCs) in soil were higher after incorporating fall-planted vs. spring-planted Brassicas. Mustard produced the most ITCs and oilseed radish produced the least. Five weeks after incorporation, populations of Pythium spp. and Sclerotium rolfsii did not differ among treatments. Populations of Rhizoctonia solani were higher in fallow without VIF than in all mulched treatments, indicating that the VIF reduced populations of R. solani. Pepper plants transplanted into the plots were stunted due to infection of the roots in all plots by Pythium aphanidermatum but particularly in the fallow plots without VIF. Pepper yields were consistently highest in Brassica treatments compared with fallow with VIF, and yields in fallow with VIF yields were higher than yields in fallow without VIF.<br /> <br /> <br /> Tennessee (Canaday): A laboratory study on the effects of chloride salts on the micro-partitioning of root Ca was conducted using energy-dispersive analysis of x-rays to measure the relative changes in the root nutrients. Detectable levels of calcium in the outer cell layers of 10 to 13 day-old soybean taproots were significantly lower when plants were grown in soil treated with potassium chloride, sodium chloride, or magnesium chloride than when the plants were grown in untreated soil or soil treated with potassium sulfate. <br /> <br /> <br /> Tennessee (Ownley): Anaerobic soil disinfestation (ASD) treatments were applied to soil in growth chamber pot studies to determine effects on populations of Rhizoctonia solani, and growth and disease of a subsequent broccoli crop. Cover crops were hairy vetch, arugula, wheat, cereal rye, crimson clover, and mustard. A fallow control was included; due to lack of organic matter incorporated into the control, it was anticipated that R. solani populations would be low. Cover crops were grown for 4 weeks, and then chopped and incorporated into soil. Inoculum of R. solani grown on cornmeal:sand was added to all pots at 2% w/v. Half the pots were irrigated (ASD treatment) after incorporation of the cover crops, and half received no water. After 4 weeks, toothpicks were inserted into soil as baits for R. solani and Packman broccoli seed were planted into the pots to determine pathogenicity of R. solani populations. The toothpick bait method was used to determine populations of R. solani per gram of soil. Two trials were conducted. Choice of cover crop incorporated into soil had a significant impact on plant growth, Rhizoctonia disease rating of broccoli, and number of propagules of R. solani in soil, with and without irrigation. Irrigation following cover crop incorporation is an important component of the ASD method in order to achieve anaerobic soil conditions. Without irrigation, the largest broccoli seedlings with lowest disease ratings and lowest numbers of R. solani propagules in soil were found in the fallow control for both trials. In trial 1, broccoli seedlings from pots treated with the hairy vetch cover crop were also large and not different from the fallow control. However, disease rating and populations of R. solani in soil were higher. In trial 2, there were no differences in number of R. solani propagules in soil or plant disease rating among the cover crops; but crimson clover resulted in larger broccoli seedlings than the mustard treatment. in trial 1 (with irrigation, ASD treatment), broccoli from soil mixed with chopped hairy vetch were twice as large (weight) as the fallow control, and were equally low in disease rating and propagule counts of R. solani in soil as the control. In trial 2 (with irrigation), crimson clover, wheat, and hairy vetch resulted in broccoli shoot height that was the same as the fallow control plants. Rhizoctonia populations were equally low in the fallow and wheat soil treatments, while disease rating of broccoli was lowest, and not different from the fallow control, with hairy vetch, wheat, crimson clover, and cereal rye ASD treatments.<br /> <br /> <br /> Texas: Three studies were conducted. In the first study, laboratory assays were conducted to determine the impact of oilseed meals and isothiocyanates (ITCs) on Phymatotrichopsis omnivora, the causal agent of cotton root rot. The effect of oilseed meals from both Brassicaceous plants, including mustard (Brassica juncea) and camelina, as well as non-Brassicaceous plants, including jatropha, flax, and Chinese tallow, on P. omnivora sclerotial germination and hyphal growth in Branyon clay soil were investigated. Also, the effects of four types of individual ITCs, including allyl, butyl, phenyl, and benzyl ITC, on P. omnivora growth on potato dextrose agar (PDA) were assessed. Oilseed meals were added to soil at rates of 0, 1, and 5% (w/w). The results showed that all tested brassicaceous and jatropha seed meals were able to inhibit P. omnivora sclerotial germination and hyphal growth at 5% and 1% application rates, respectively, with mustard seed meal being the most effective. Neither flax nor Chinese tallow showed any inhibitory effects on sclerotial germination. All tested ITCs inhibited P. omnivora OKAlf8 hyphal growth, although the level of inhibition varied with concentration. The IC50 values were 0.62 ± 0.19, 4.47 ± 0.08, 5.67 ± 0.10, and 20.48 ± 0.30 µg cm-3 for allyl, butyl, phenyl, and benzyl ITC respectively. In the second study, a laboratory study was conducted to examine the impact of various oilseed meals on soil C, N, bacteria, and fungi. Mustard (B. juncea) and flax seed meals and sorghum-sudangrass were added to Weswood loam soil at levels of 0, 1, 2.5, and 5% (w/w). Both the type of amendment and application rate affected soil organic C, total C & N, and C & N mineralization. Mustard meal amendment initially inhibited C mineralization as compared to flax, but >50% of mustard and flax organic C was mineralized within 51 days. Nitrogen mineralization was similar for flax and mustard, except for the 2.5% rate at which a lower proportion of mustard N was converted to nitrate. Soil bacterial and fungal community responses to the 2.5% amendment rate were characterized using 16S rRNA and fungal ITS gene sequences at four time points over the course of the experiment. Distinct bacterial and fungal communities occurred among amendment-type lines, with mustard inducing large increases in the abundance of bacterial groups known to include many fungal disease-suppressing bacteria (e.g., Bacillus, Pseudomonas, and Streptomyces spp.), while other amendments increased Actinobacteria or Bacteroidetes abundances. Shifts in bacterial community composition slowed after 14 days but all biomass treatments still contained unique communities after 28 days. Dramatic shifts were also seen among fungi, with fungal phylotype richness decreasing by > 60% following mustard seed meal addition. Changes among the fungi, especially for the mustard treatment, persisted throughout the entire study. Finally, a third laboratory study was conducted to determine soil microbial community responses to four types of ITCs (allyl, benzyl, phenyl, and butyl) in a microcosm study using Weswood loam soil. Microcosms were amended with 50 µg g-1 of an individual ITC and incubated at 25°C for 28 days. Community qPCR assays were used to evaluate relative abundances of soil microbial populations after 2, 7, 14, 21, and 28 days. Soil microbial community composition after 2, 7, and 28 days was determined through tag-pyrosequencing using 454 GS FLX titanium technology, targeting ITS and 16S regions for fungal and bacterial communities respectively. Our results showed that the application of ITCs altered soil microbial community composition by suppressing either soil fungal or bacterial abundance, depending upon the ITC applied. Allyl ITC greatly decreased soil fungal abundance by >6 times compared with the unamended control, and significantly changed both fungal and bacterial communities composition. In contrast, butyl ITC had a larger impact on bacterial rather than fungal abundance and composition.<br /> <br /> <br /> Objective 3. Examine the genetic diversity of Rhizoctonia solani between natural ecosystems and agricultural ecosystems. <br /> <br /> <br /> Arkansas: Populations of Rhizoctonia solani were examined using the protocol previously developed by the regional project members. The sampling protocol involved baiting populations of Rhizoctonia from soil using toothpick baits and the TS selective media, as well as seedling isolations. In 2011, isolations included surveys to collect samples from diverse crops in the Southeast. In addition, a long-term rotation study including rice, soybean, corn, and winter wheat was used to examine shifts in soil populations of Rhizoctonia spp. as a result of cropping history. Isolates are currently being characterized. Soybean and rice rotations are being assessed for spatial distribution of Rhizoctonia populations. The diversity of Rhizoctonia spp. in these cropping systems included R.solani (AGs 11, 1-1A, 4 and 2-1), Rhizoctonia oryzae, and bi-nucleate Rhizoctonia species. A new selective medium TS was developed for improved recovery of Rhizoctonia spp. from soil. The medium is an improvement over the previous selective media used as part of the project as a result of better suppression of Trichoderma spp. and Mucorales. <br /> <br /> <br /> Kentucky: Isolates recovered in 2010 were stored improperly and could not be shipped to C. Rothrock for identification. We were unable to work on this objective in 2011, but plan to re-do the 2010 sampling and isolation to provide comparisons of isolation methods for R. solani and for diversity of R. solani.<br /> <br /> <br /> Louisiana: A study was conducted at the Macon Ridge Research Station near Winnsboro, LA to assess the diversity of Rhizoctonia spp. in fields planted to cotton, grain sorghum, and soybean fields. A standardized protocol developed by the S-1028 group was used. Thirty random soil samples per crop were taken from two locations (15/location). Within each location five sample sites were randomly chosen. When possible, one location was in a field where conventional tillage practices were utilized and the other location was from a field where minimum tillage practices were utilized. Soil was transported to the lab for further processing. Soil was placed in plastic pots. Moisture was supplemented to soil to saturation and toothpicks were placed in soil and left for a predetermined time according to the protocol. Toothpicks were plated onto ethanol-potassium nitrate medium. Plates were monitored for growth of Rhizoctonia 24 hours after plating and colonies were quantified. Colonies were then transferred to antibiotic-amended water agar plates. Number of colonies per treatment is presented in the table below. Attempts were made to transfer Rhizoctonia colonies to water agar, but fungal contaminants interfered with this process. <br /> <br /> 2011 Rhizoctonia solani Diversity Study<br /> <br /> Tillage Crop # Colonies <br /> <br /> Minimum CT 15 <br /> <br /> Conventional GS 24 <br /> <br /> Conventional CT 16 <br /> <br /> Minimum SB 21 <br /> <br /> Minimum SB 36 <br /> <br /> Conventional GS 15 <br /> <br /> <br /> An additional study was conducted on the Macon Ridge and Red River Research Station in collaboration with Dr. Craig Rothrock at the University of Arkansas. Each station served as a sampling site. Soil and seedling samples of cotton and soybean were taken at these sites. Samples were then shipped to Dr. Rothrock's lab for further processing.<br /> <br /> <br /> Additional Ecological and Genetic Diversity of Soilborne Pathogens and Indigenous Microflora Outcomes<br /> <br /> Iowa: Activities included conducting and analyzing studies on the effects of biochar on soil respiration and soil microbial populations. Biofuels are being carefully evaluated for their efficacy to replace hydrocarbons. Pyrolysis of plant materials leaves behind residues (biochars) that may impact soil cultural practices and affect soilborne organisms and plant growth. We studied corn stover and three biochars (pyrolysis products made from corn stover) and their decomposition in soil. The study revealed that the percentage of carbon lost from the amendments after eight weeks was greatest for corn stover (44% by weight), followed by the two less completely pyrolyzed biochars (9 and 10%), and least (4%) from the biochar made by fast pyrolysis at 500C. Thus, all biochars contained at least some readily bioavailable carbon that could be measured as CO2 emissions over 8 weeks. Microbial availability roughly correlated inversely with the degree of pyrolysis completeness. There was an apparent shift in microbial communities from bacteria and actinomycetes to fungi with increasing degree of pyrolysis. A soil biology educational website containing video of important soil organisms including fungi, ectomycorrhizae, endomycorrhizae, and their activities in the soil food web is maintained at (http://www.agron.iastate.edu/~loynachan/mov/). <br /> <br /> <br /> Oklahoma: A study of the genetic diversity of populations of Sclerotinia minor in agricultural soils in Stillwater, OK, was completed. Sclerotia collected from five experimental peanut plots (two breeding, two variety disease resistance assessment, and one chemical trial) were sterilized, germinated, and strains were characterized to species morphologically. Species identification was verified by sequencing the ITS region (including the small subunit of the rDNA). Most of the strains recovered (n=130) were verified as S. minor, however a few (n=15) produced sequences with higher similarity to S. sclerotiorum than S. minor, but not 100% similar. The population structure of S. minor was examined and it was determined that strains from breeding and disease resistance assessment were not significantly different. However, significant differentiation was found between S. minor populations from chemical trial plot and all other plots. Currently, 15 strains representing the genetic diversity of the studied sample are being evaluated for pathogenicity and fungicide sensitivity. A second study is currently underway to evaluate the effects of sublethal fungicide concentrations on populations of oomycete (Pythium and Phytophthora) and fungal (S. homeocarpa, Botrytis cinerea, and R. solani), plant pathogens.

Publications

Peer-reviewed<br /> <br /> Aliferis, K.A., Jabaji, S., and Cubeta, M.A. 2011. Chemotaxonomy of fungi in the Rhizoctonia solani species complex using GC/MS metabolic profiling. Metabolomics (30 July), pp 1-11, doi:10.1007/s11306-011-0340-1.<br /> <br /> Bartz, F.E., Danehower, D.A., Glassbrook, N., and Cubeta, M.A. 2011. Elucidating the role of phenylacetic acid and hydroxy- and methoxy-phenylacetic acid derivatives in the pathogenic activity of Rhizoctonia solani AG-3. Mycologia 103: (In Press).<br /> <br /> Berbegal, M, Yanez, J.M, Garzon, CD, Armengol, J, Jimenez-Diaz, R.M, and Jimenez-Gasco, M.M. 2011. Development and application of new molecular markers for the analysis of genetic diversity in Verticillium dahliae populations. Plant Pathology. 60: 866877.<br /> <br /> Copes, W., Rodriguez-Carres, M., Toda, T., Rinehart, T.A., and Cubeta, M.A. 2011. Seasonal prevalence of species of binucleate Rhizoctonia fungi in growing<br /> medium, leaf litter, and stems of container grown azalea. Plant Dis. 95:705-711.<br /> <br /> Ellis, D., J. Gosai, C. Emrick, R. Heintz, L. Romans, D. Gordon, S. E. Lu, F. Austin, and L. Smith. 2011. Occidiofungin's chemical stability and in vitro potency against Candida species. Antimicrob. Agents Chemother. In Press.<br /> <br /> Garzon, CD, Molineros, J.E., Yanez, J.M, Flores, F.J., Jimenez-Gasco, M.M, and Moorman G.W. 2011. Sublethal doses of mefenoxam enhance Pythium damping-off of geraniums. Plant Disease 95:1233-1238. <br /> <br /> Gu, G. Y., J. L. Smith, A. L. Liu, and S.-E. Lu. 2011. A genetic and biochemical map for the biosynthesis of occidiofungin, an antifungal produced by Burkholderia contaminans strain MS14. Appl. Environ. Microbiol. 77:6189-6198.<br /> <br /> Hu, P., A.S. Wang, A.S. Engledow, E.B. Hollister, K.L. Rothlisberger, J.E. Matocha, D.A. Zuberer, T.L. Provin, F.M. Hons, and T.J. Gentry. 2011. Inhibition of the germination and growth of Phymatotrichopsis omnivora (Cotton Root Rot) by oilseed meals and isothiocyanates. Appl. Soil Ecol. 49:68-75.<br /> <br /> Kim, D.G, Isenhart T.M., Parkin T.B., Schultz R.C., and Loynachan T.E. 2010. Methane flux in cropland and adjacent riparian buffers with different vegetation covers. J. Environ. Qual. 39:97-105.<br /> <br /> Mengistu, A., Arelli, P. A., Bond, J. P., Shannon, G. J., Wrather, A. J., Rupe, J. B., Chen, P., Little, C. R., Canaday, C. H., Newman, M. A., and Pantalone, V. R. 2011. Evaluation of soybean genotypes for resistance to charcoal rot. Online. Plant Health Progress doi:10.1094/PHP-2010-0926-01-RS.<br /> <br /> Richter, B. S., D. M. Benson, and K. L. Ivors, 2011. Microbial profiling of cultural systems for suppression of Phytophthora root rot in Fraser fir. Plant Dis. 95:537-546.<br /> <br /> Richter, B. S., Ivors, K. I., Shi, W., and Benson, D. M. 2011. Cellulase activity as a mechanism for suppression of Phytophthora root rot in mulches. Phytopathology 101:223-230.<br /> <br /> Samuels, G., Ismaiel, A., McMahon, P., Guest, D., Rosmana, A., Junaid, M., Rodriguez-Carres, M., and Cubeta, M.A. 2011. Vascular streak dieback of cacao in Southeast Asia and Melanesia: in planta detection of the pathogen and a new taxonomy. Fungal Biology. (In Press).<br /> <br /> Sullivan, M. J., Parks, E. J., Cubeta, M. A., Gallup, C. A., Moyer, J.W., and Shew, H.D. 2010. Assessment of genetic diversity from a field population of Phytophthora nicotianae with a changing race structure. Plant Disease 94:455-460.<br /> <br /> Woodhall, J.W., Webb, K.M., Harper, G., Peters, J.C. Rodriguez-Carres, M., and Cubeta, M.A. 2011. First report of a new binucleate Rhizoctonia in UK potato tubers. New Disease Reports 23:31 [doi:10.5197/j.2044-0588.2011.023.031].<br /> <br /> Wang, A.S., P. Hu, E.B. Hollister, K.L. Rothlisberger, A. Somenahally, T.L. Provin, F.M. Hons, and T.J. Gentry. 2012. Impact of Indian mustard (Brassica juncea) and flax (Linum usitatissimum) seed meal applications on soil carbon, nitrogen, and microbial dynamics. Appl. Environ. Soil Sci. Article ID 351609; 14 p; doi:10.1155/2012/351609. <br /> <br /> Yin, J., Koné, D., Rodriguez-Carres, M. Cubeta, M.A., Burpee, L.L. Fonash, E.G. Csinos, A.S., and Ji, P. 2011. First report of root rot caused by binucleate<br /> Rhizoctonia anastomosis group F on Musa spp. Plant Disease 94:490.<br /> <br /> Abstracts<br /> <br /> Abd-Elmagid A, Garrido P, Hunger R, Melouk H, and Garzon, CD. 2011. Multiplex PCR for diagnosis of four Sclerotinia species. 8th Annual Graduate Research in Biological Sciences Symposium. Stillwater, OK, September 22-23, 2011.<br /> <br /> Canaday, C. H., Donald, P., and Mengistu, A., 2011, Increases in snap bean and soybean seedling diseases associated with a chloride salt and changes in the micro-partitioning of tap root calcium. Phytopathology101:S26.<br /> <br /> Dobhal, S, Garrido, P, Melouk, H, Garzon, CD. 2011. Strain discrimination and genetic diversity assessment of the fungal plant pathogen Sclerotinia minor from neighboring agricultural fields in Oklahoma. Chemical and Biological Defense Science and Technology Conference. Las Vegas, NV, November 14-18, 2011.<br /> <br /> Espindola A., Daniels J., Stobbe A., Schneider W., and Garzon, C.D. 2011. Design and validation of queries for the detection of Phytophthora ramorum in simulated metagenomes. American Phytopathological Society 2011 National Meeting, Honolulu, Hawaii. Phytopathology. 101: S50<br /> <br /> Flores, F., Garzon, C. D. Evaluation of hormesis on the response of soilborne plant pathogens to pesticides in vitro. 8th Annual Graduate Research in Biological Sciences Symposium. Stillwater, OK, September 22-23, 2011<br /> <br /> Flores, F., Garzon, C. D. 2011. Evaluation of hormesis on the response of soilborne plant pathogens to pesticides in vitro. 8th Annual Graduate Research in Biological Sciences Symposium. Stillwater, OK, September 22-23, 2011<br /> <br /> Garrido P., Melouk H, and Garzon, C.D. 2011. Population structure and genetic diversity of Sclerotinia minor from peanut research plots in Oklahoma. American Phytopathological Society 2011 National Meeting, Honolulu, Hawaii. Phytopathology. 101: S59<br /> <br /> Hollister, E.B., P. Hu, A.S. Wang, F.M. Hons, and T.J. Gentry. 2011. Soil bacterial and fungal community responses to oilseed meal addition. In Abstracts, 111th General Meet., ASM, New Orleans, LA. 21-24 May 2011. ASM, Washington, D.C.<br /> <br /> Hu, P., E. Hollister, A. Wang, F. Hons, and T. Gentry. 2010. Soil microbial community changes after oilseed meal application. In Abstracts, ASA-CSSA-SSSA Annual Meet., Long Beach, CA. 31 October - 3 November 2010.<br /> <br /> Orquera, G., Mogrovejo, C., Villamarin, D., Jarrin, F., Garzon, C.D., Molineros, J., Forbes, G., Koch, A., Benitez, M.S. 2011. Characterization of microbial populations from P. infestans suppressive Andean soils in Ecuador. American Phytopathological Society 2011 National Meeting, Honolulu, Hawaii. Phytopathology. 101: S133<br /> <br /> Spurlock, T., Rothrock, C., and Monfort, W. 2011. A new selective medium for isolation of Rhizoctonia spp. from soil. (Abstr.) Phytopathology 101:S170. <br /> <br /> Villamarin D, Orquera G., Mogrovejo, C., Garzon, C.D., Molineros, J., Forbes, G., Koch, A., Benitez, M.S. 2011. Supressiveness to Phytophthora infestans infection in potato tubers by Andean soils from three provinces of Ecuador. American Phytopathological Society 2011 National Meeting, Honolulu, Hawaii. Phytopathology. 101: S183<br /> <br /> Other Publications<br /> <br /> Canaday, C.H., 2011, Effects of seed treatment fungicides, biological agents, and potash fertilizers on snap bean seedling diseases, 2010. Plant Disease Management Reports 5:ST002. <br /> <br /> Dau, L. 2010. The promise of biochar. M.S. creative component, Iowa State University, Ames, IA.<br /> <br /> Flores, FJ. 2010. Effect of Low Doses of Pesticides on Soilborne Pathogens  An Approach to the Hormetic Response. M.S. Thesis. Oklahoma State University<br /> <br /> Hansen, Z. R. 2011. Potential of three brassica cover crops for biofumigation in the field and the relationship between soil myrosinase and biofumigation efficacy. M.S. Thesis, Clemson University. 136 pp.<br /> <br /> Hansen, Z. R., Keinath, A. P., Baccari, G. V., and DuBose, V. B. 2011. Evaluation of Brassica cover crops for control of root rot in peppers, 2010. Plant Disease Management Reports 5:V073. Online publication. doi: 10.1094/PDMR05.<br /> <br /> Keinath, A. P., Baccari, G. V., and DuBose, V. B. 2011. Evaluation of biofungicides to prevent damping-off of organic arugula, 2010. Plant Disease Management Reports 5:ST010. Online publication. doi: 10.1094/PDMR05.<br /> <br /> Grants awarded<br /> <br /> Backman PA, Stehouwer R, and Gugino B. Conservation Agriculture as a potential pathway to better resource management, higher productivity, and improved socio-economic conditions in the Andean Region. USAID SANREM CRSP, $340,000; 5 years. <br /> <br /> Backman PA and Gugino B. Integrated Pest Management: Science for Agricultural Growth in Latin America and the Caribbean. USAID IPM CRSP $190,000; 5 years. <br /> <br /> Baird, R. Participatory modeling and decision support for improving sweet potato production efficiency, quality, and food safety. USDA-CSREES-SCRI. $ 32,500; 1 year.<br /> <br /> Baird, R. Baseline data on sweetpotato root microorganisms and the determination of the causal agents associated with a newly emerging rot diseases complex. SRI $37,000; 1 year.<br /> <br /> Koch AR, Benitez MS, Garzon CD, Molineros JE. Population dynamics of Phytophthora infestans in potato producing soils in the Carchi and Chimborazo provinces, and inoculum infection potential. Politechnic School of the Army, Ecuador. $38,000; 1 year. <br /> <br /> Keinath, A. Development of Grafting Technology to Improve Sustainability and Competitiveness of the US Fruiting Vegetable Industry. USDA, NIFA, SCRI, $286,403; 3 years. <br /> <br /> Lu, S. Genetic and Chemical Characterization of Novel Antibiotics Produced by Plant Disease-Suppressive Bacteria. MAFES-SRI. $79,300; 1 year.

Impact Statements

1. A new selective medium was developed as part of the multi-state project to improve recovery of Rhizoctonia spp. A more effective selective medium will allow better detection of the pathogen in soil and the examination of the diversity of this important pathogen across different native and agricultural systems increasing our understanding of the role of cropping history and transport of the pathogen in disease development and diversity.
2. The potential of biochar as soil amendment was evaluated. New knowledge was shared at a conference and to high school students in Des Moines and college students in a class taught at Iowa State University. The sharing of knowledge in educational and conference settings allows the audience to make educated decisions on the use of biochars in agricultural settings.
3. Testing commercial a formulation of Trichoderma viride & T. harzianum (Tenet WP) as a soil drench or transplant treatment was shown to provide significant suppression of Phytophthora blight. Tenet has potential to serve as an adjunct for traditional fungicides used for management of Phytophthora blight that could help enhance the performance of these products and provide greater returns to vegetable producers.
4. Several potential fungi and bacteria that show pathogenic traits on sweet potato parenchyma tissue have been identified. After being subjected to greenhouse plant trials to determine field-scale effects of individual isolates, those determined to be pathogenic to both storage root tissue and intact sweet potato plants will be selected for field-level tests at the Pontotoc MAFES station.
5. It was determined that Lagenaria rootstocks were more promising than inter-specific hybrid Cucurbita or citron rootstocks for seedless watermelon grown in fields affected with Fusarium wilt.
6. It was shown that cover cropping with Brassica can increase yield of bell pepper affected with root rot, and that Brassica cover crops should be planted in the fall in coastal South Carolina to obtain sufficient biomass.
7. New evidence was found that seedmeals of several brassicaceous species, mustard particularly, have potential for inhibiting P. omnivora, thus possibly reducing cotton root rot. Also, that oilseed meal amendment and ITCs have the potential to substantially alter soil microbial community structure. This information could ultimately be used by plant breeders in the development and selection of crops producing specific isothiocyanates (and other biocidal compounds) for biofumigation.

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Report Information

Participants

Backman, Paul (pbackman@psu.edu) - Pennsylvania State University;
Baird, Richard (RBaird@plantpath.msstate.edu) - Mississippi State University;
Benson, Mike (mike_benson@ncsu.edu) - North Carolina State University;
Broders, Kirk (kirk.broders@unh.edu) - University of New Hampshire;
Canaday, Craig (ccanaday@utk.edu) - University of Tennessee;
Cubeta, Marc (marc_cubeta@ncsu.edu) - North Carolina State University;
Dick, Warren (dick.5@osu.edu)  Ohio State University;
Elliott, Monica (melliott@ufl.edu) - University of Florida;
Garzon, Carla Domenica (carla.garzon@okstate.edu) - Oklahoma State University;
Gentry, Terry (tgentry@ag.tamu.edu ) - Texas AgriLife Research;
Ghanem, Nina J Abou (ninaag@igbb.msstate.edu) -Mississippi State University ;
Graham, Peter (graha019@umn.edu)  University of Minnesota;
Jimenez Gasco, Maria (jimenez-gasco@psu.edu)  Pennsylvania State University;
Keinath, Tony (tknth@clemson.edu) - Clemson University;
Kurle, James E. kurle001@umn.edu - University of Minnesota;
Lamour, Kurt H. klamour@utk.edu - University of Tennessee;
Loynachan, Thomas (teloynac@iastate.edu) - Iowa State University;
Lu, Shien (sl332@msstate.edu ) - Mississippi State University;
Ownley, Bonnie (bownley@utk.edu) - University of Tennessee;
Padgett, Boyd (bpadgett@agcenter.lsu.edu) - Louisiana State University;
Rothrock, Craig (Rothrock@uark.edu) - University of Arkansas;
Santamaria, Luisa (luisa.santamaria@oregonstate.edu) - Oregon Cooperative Extension (ORE);
Seebold, Kenneth (kwseebold@uky.edu ) - University of Kentucky;
Westphal, Andreas (Westphal@purdue.edu) - Purdue University;
Williams, Mark (markwill@vt.edu)  Virginia Tech

Brief Summary of Minutes

Harald Scherm  Administrative Advisor oversaw the annual meeting and provided guidance and information on the renewal process. To date 15 members are signed up for the new S1053. As S1028 has come to a close a termination report will need to be prepared. When discussing future directions for the research group, emphasis on collaborative proposal development was suggested as an area the group could work together to improve on from the last project period.

Those in attendance included:

Sheng Yu - Mississippi State University

Kurt Lamour  University of Tennessee

Kirk Broders  University of New Hampshire

Craig Rothrock  University of Arkansas


Carla Garzon  Oklahoma State University

Craig Canaday  University of Tennessee WTREC

Louisa Santamaria  Oregon State University

After introduction, each member in attendance provided a presentation and update of current ongoing research and results from the past year associated with the project. At the conclusion of the presentation, the group came to a consensus to hold next years meeting of the S1053 multi-state group in Oklahoma City, OK around the first week of November. Kurt Lamour was nominated for Secretary and a unanimous vote confirmed his new post. As secretary, Kurt is also the Chair-elect. Kirk Broders will be the incoming Chair, and Craig Rothrock is the outgoing chair.

The final topic of discussion was how we can better integrate projects by individual investigators to tackle larger problems that require a multi-state effort to accomplish. Some of the ideas put forth included:

Chloride and relationship to Pythium

P. capsici distribution and management

Pythium taxonomy and describing non-spore forming species

Disease complexes symposium/workshop

Rhizoctonia population resources

International Potato Center for Rhizoctonia resources

Meeting adjourned.

Accomplishments

Objective 1. Examine commercial and non-commercial biocontrol agents for use as seed treatments, in-furrow treatments or as potting mix amendments.<br /> <br /> A regional trial of biocontrol agents and amendments for control of wire stem of broccoli (Rhizoctonia solani) was conducted at five sites in four states (KY, TN-Knoxville, TN-Jackson, AR, and SC) over multiple years. Eight treatments were evaluated; millet seed infested with binucleate-Rhizoctonia (BNR), Monarda herbage, BioYield Flowable, Beauveria bassiana isolate 11-98, Actinovate AG, an untreated control, and a fungicide standard. Positive responses included: Jackson, TN (Canaday) all materials reduced the incidence of Rhizoctonia root rot or Pythium root rot compared to the untreated control and plant growth-promoting rhizobacteria (PGPR) were equal to the fungicide standard for disease control and in another year the PGPR product BioYield Flowable increased head weight of some broccoli cultivars. In eastern TN (Ownley), broccoli seed treated with Beauveria bassiana 11-98 (1,000,000 spores/seed), followed by planting into potting mix containing 2% bioactive dried Monarda herbage resulted in equal or greater head size and yield weight than the standard fungicide treatment and greater than other treatments evaluated. In South Carolina (Keineth), application to potting mix of Monarda herbage (2% w/w) and BioYield Flowable increased broccoli plant height at 50 days after transplanting compared with the pathogen-infested control. In another year, Keinath, a non-pathogenic (binucleate) Rhizoctonia isolate and BioYield Flowable significantly increased the percentage of disease-free broccoli plants and reduced the number of plants with wirestem 30 days after transplanting. <br /> <br /> Additional biocontrol studies were conducted by individual participants. <br /> <br /> Benson (NC): Binucleate Rhizoctonia (BNR) isolates BNR621 and P023, and Trichoderma hamatum isolate 382 induced resistance to the foliar fungal pathogen Botrytis cinerea in geranium. <br /> <br /> Canaday (TN): The combination of BioYield and fungicide treatments with and without application of Actigard had no effect on the incidence of stem rot (Sclerotinia sclerotiorum) and buckeye rot (Phytophthora nicotianae) affecting tomato. <br /> <br /> Canaday (TN): In field tests in soils natural infested with R. solani, Pythium spp., Macrophomina phaseolina, and Fusarium spp., GB03 or MBI600 used alone or as supplements to standard seed treatment chemicals, failed to increase snap bean stand or yield. Additional evaluations found response to Bacillus subtilis (MBI600) when added to fungicide seed treatments for control of seedling diseases of spring-planted snap beans varied with the cultivar used. For soybean, the percentage of disease-free plants was greatest with a seed treatment combination of the PGPR strain Bacillus subtilis MBI 600 and the fungicides mefenoxam and fludioxinil. <br /> <br /> Ownley (TN): Beauveria bassiana 11-98 endophytically colonized tomato and cotton following seed treatment. The extent of colonization was dependent on seed treatment rate and host plant. Plant colonization by B. bassiana was essential for disease control. Application of Beauveria bassiana 11-98 to seedling roots induced systemic resistance in cotton leaves against the bacterial pathogen Xanthomonas axonopodis pv. malvacearum. Integration of Beauveria (B. bassiana 11-98 or BotaniGard) seed treatments and RootShield (Trichoderma) provided mixed results on tomato seedlings in soil infested with Rhizoctonia solani or Pythium myriotylum. <br /> <br /> Padgett (LA): Biological fungicides when applied alone and in combination with strobilurin fungicides on wheat, corn, and soybean were not effective for managing diseases or preserving yields when applied alone. <br /> <br /> Seebold (KY): Bioten, a commercial formulation containing Trichoderma harzianum and T. viride, significantly reduced Phytophthora blight, caused by Phytophthora capsici, on summer squash when applied to soil via drip irrigation (25% to 60% reduction over the untreated control), while Ridomil Gold reduced incidence by at least 60% to 90% compared to the untreated control. <br /> <br /> Elliot (FL): Container-grown Mexican fan palm (Washingtonia robusta) and queen palm (Syagrus romanzoffiana) treated at the time of planting with four commercial microbial inoculants (each containing arbuscular mycorrhizal fungi, either alone or with other microbial components or fertilizers), or two fertilizers did not improve growth over the control in the P-deficient soil or P-sufficient soil. <br /> <br /> Keinath (SC): The commercial biofungicides RootShield and Actinovate were tested for prevention of damping-off of arugula on a noncertified organic vegetable farm in Charleston County, South Carolina. Fresh weight of arugula harvested 1 month after seeding did not differ among treatments. <br /> <br /> Discovery of new biological control agents were examined as part of the project.<br /> <br /> Lu (MS): Occidiofungin from the bacterium Burkholderia contaminans strain MS14 was evaluated regarding its stability and efficacy as an antifungal compound. Functions for the genes in the 56-kb occ gene cluster that is essential for production of occidiofungin was examined. Interestingly, disruption of the ocfC gene in the ocf gene cluster by the insertion of the nptII cassette resulted in xylose-free occidiofungin production and reduction of antifungal activity against the indicator fungus Geotrichum candidum. However, the antifungal activity of the xylose-free occidiofungin was significantly increased against Candida species as compared with the wild-type occidiofungin. These results are significant for development of agricultural biofungicides and medical parametrical drugs. In addition, more than 60 bacteria from 900 isolates obtained from Mississippi soils exhibited significant antifungal and/or antibacterial activities against pathogenic fungi and bacteria. <br /> <br /> Garzon (OK): A study of bacterial communities present in Andean soils from Ecuador suppressive to Phytophthora infestans was conducted. Approximately 3,000 bacterial strains of Bacillus, Pseudomonas, and Actinobacteria (including Streptomyces) were isolated and evaluated for their ability to inhibit P. infestans and Rhizoctonia solani. Strains with stronger inhibitory effects were selected for further characterization, and 200-300 strains of each bacterial group were sequenced (16S rDNA). Currently, the microbial communities in these soils are under further examination using metagenomic methods. <br /> <br /> Backman (PA): Endospore-forming bacterial endophytes were isolated from Theobroma cacao. Cacao leaves, pods, branches, and flower cushions were removed from cacao trees escaping disease and tissue was surface sterilized, heat treated (75°C for 15 min), and plated. 69 endophytic endospore-forming bacteria were isolated. Sixteen isolates were chitinolytic and in antagonism studies against cacao pathogens, 42% were antagonistic to Moniliophthora roreri, 33% to M. perniciosa, and 49% to Phytophthora capsici. 25% of isolates inhibited the growth of both Moniliophthora spp., while 22% of isolates inhibited the growth of all three pathogens. All 14 isolates were capable of endophytic colonization; while 8 isolates significantly inhibited P. capsici lesion formation in detached leaf assays. <br /> <br /> Ojective 2. Examine the effect of cultural practices on soilborne pathogens and plant growth <br /> <br /> The value of mulch or cover crop amendments were examined for a number of cropping systems.<br /> <br /> Benson (NC): Phytophthora root rot of Fraser fir, caused by several Phytophthora spp., is a severe problem in Christmas tree production. Treatments included wood chips, wood chips plus compost, or pine bark as raised beds, and compost or sulfur tilled into soil. Bacterial and fungal counts, microbial activity, and cellulase activity were higher in mulch than in soil at all sites and times (P<0.01), and generally did not differ among mulch types or among soils. Treatments significantly affected disease ratings and tree survival at three of five sites, with one or more mulch treatments yielding lower disease ratings and greater survival than controls. Tree mortality at each time point varied significantly with cellulase activity in the upper root zone (P=0.005). <br /> <br /> Ownley (TN): Anaerobic soil disinfestation (ASD) treatments were applied to soil in growth chamber pot studies to determine effects on populations of Rhizoctonia solani, and growth and disease of a subsequent broccoli crop. Cover crops were hairy vetch, arugula, wheat, cereal rye, crimson clover, and mustard. Irrigation following cover crop incorporation is an important component of the ASD method in order to achieve anaerobic soil conditions. In trial 1 (with irrigation, ASD treatment), broccoli from soil mixed with chopped hairy vetch were twice as large (weight) as the fallow control, and were equally low in disease rating and propagule counts of R. solani in soil as the control. In trial 2 crimson clover, wheat, and hairy vetch resulted in broccoli shoot height that was the same as the fallow control plants. Rhizoctonia populations were equally low in the fallow and wheat soil treatments, while disease rating of broccoli was lowest, and not different from the fallow control, with hairy vetch, wheat, crimson clover, and cereal rye ASD treatments. <br /> <br /> Keinath (SC): Cahaba White vetch reduced the number of wilted seedless watermelon plants in one of two experiments. An on-farm trial was done to evaluate the effects of cover crops and fumigants on Fusarium wilt and yield of seedless watermelon. The two fumigants, 67% methyl bromide-33% chloropicrin and Telone-C35, both combined with hairy vetch (Vicia villosa), were more effective than hairy vetch or rye alone or fallow at increasing yield. <br /> <br /> Oilseed radish (Raphanus sativus), mustard (Brassica juncea cv. Pacific Gold), and winter rapeseed (B. napus cv. Dwarf Essex) were incorporated as biofumigation cover crops. After incorporation, plots were covered with white-on-black virtually impenetrable film (VIF). Control treatments were fallow with and without VIF. Isothiocyanate concentrations (ITCs) in soil were higher after incorporating fall-planted vs. spring-planted Brassicas. Five weeks after incorporation, populations of Pythium spp. and Sclerotium rolfsii did not differ among treatments. Populations of Rhizoctonia solani were higher in fallow without VIF than in all mulched treatments, indicating that the VIF reduced populations of R. solani. Pepper plants transplanted into the plots were stunted due to infection of the roots in all plots by Pythium aphanidermatum but particularly in the fallow plots without VIF. Pepper yields were consistently highest in Brassica treatments compared with fallow with VIF.<br /> <br /> Rothrock (AR): The winter cover crop Indian mustard cv. Fumus reduced seedling root and hypocotyl disease symptoms. Brassica amendments also reduced early season galling from the root-knot nematode similar to the Telone treatment. Brassica treatments improved cotton yield over the control and were similar or greater than the Telone treatment in both the root-knot and reniform nematode infested locations. In other studies, rate of Brassica application had the greatest impact on disease symptom reduction and reducing Rhizoctonia solani isolation. <br /> <br /> A summer brassica cover crop, mustard seed meal amendment, solarization or a combination of the cover crop and solarization were compared to no soil treatment prior to establishing an annual strawberry crop. In all cases, there was a trend for higher bacterial, fungal and actinomycete populations in brassica, brassica plus solarization and mustard seed meal amended soils compared to solarized only and control soils. Total culturable bacterial populations were significantly higher in soils that had been planted with a brassica cover crop followed by solarization and soils receiving mustard seed meal amendments. From soil samples taken at 7 and 25 days after the brassica cover crop was incorporated and at the time of strawberry transplant, bacterial DGGE profiles of soils receiving different treatments were distinct and grouped separately in dendograms. <br /> <br /> Gentry (TX): The effect of oilseed meals from both Brassicaceous plants, including mustard (Brassica juncea) and camelina, as well as non-Brassicaceous plants, including jatropha, flax, and Chinese tallow, on Phymatotrichopsis omnivora sclerotial germination and hyphal growth in Branyon clay soil were investigated. The results showed that all tested brassicaceous and jatropha seed meals were able to inhibit P. omnivora sclerotial germination and hyphal growth at 5% and 1% application rates, respectively, with mustard seed meal being the most effective. All tested ITCs inhibited P. omnivora OKAlf8 hyphal growth, although the level of inhibition varied with concentration. Distinct bacterial and fungal communities occurred among amendment-type lines, with mustard inducing large increases in the abundance of the bacterial genera Bacillus, Pseudomonas, and Streptomyces spp., while other amendments increased Actinobacteria or Bacteroidetes abundances. Dramatic shifts were also seen among fungi, with fungal phylotype richness decreasing by > 60% following mustard seed meal addition. <br /> <br /> Loynachan (IA): Biochars impact on soil cultural practices and soilborne organisms and plant growth were examined. The percentage of carbon lost from the amendments after eight weeks was greatest for corn stover (44 wt%), followed by the two less completely pyrolyzedbiochars (9 and 10%), and least (4%) from the biochar made by fast pyrolysis at 500°C. Microbial availability roughly correlated inversely with the degree of pyrolysis completeness. There was an apparent shift in microbial communities from bacteria and actinomycetes to fungi with increasing pyrolysis. <br /> <br /> Broders (NH): Soil amendments and cover crops on disease suppression of Verticillium dahliae on strawberry and mint are being evaluated initially by screening strawberry accessions and a variety of cover crops for their ability to resist infection and suppress inoculum levels of the soilborne pathogen V. dahliae.<br /> <br /> Tillage systems impact was investigates as part of the project.<br /> <br /> Westphal (IN): Soybean under soybean monoculture in soil having no-tillage and infested with Fusarium virguliforme (causes sudden death syndrome; SDS) and soybean cyst nematode (Heterodera glycines) had less severe foliar symptoms of SDS in soil that had not been fumigated than in fumigated plots three years after pathogen infestation, suggesting that microbially-based soil suppressiveness had developed to these pathogens in the non-fumigated soil. Foliar symptoms of SDS in soybean were greater with chisel and moldboard plow tillage than with ridge tillage; yields were also improved with reduced tillage compared to intensive tillage.<br /> <br /> Padgett (LA): The impact of no-tillage, minimum tillage or conventional tillage on overwintering populations of several pathogens and diseases were examined. The incidence of Cercospora blight or purple seed stain on soybean did not differ among three tillage systems. Black root rot incidence on cotton was lower in conventional and minimum tillage systems. Rhizoctonia populations were highest in fields planted to grain sorghum and soybean; however, tillage practices did not impact Rhizoctonia populations. <br /> <br /> The microflora of several cropping systems were examined.<br /> <br /> Elliot (FL): Bermudagrass and bentgrass root microflora in sand-based putting greens were examined using fatty acid methyl ester profiles (GC-FAME). The two dominant genera in both bentgrass and bermudagrass rhizospheres were Bacillus and Pseudomonas, with Bacillus dominant in bermudagrass and Pseudomonas dominant or equal to Bacillus in bentgrass. Other genera that comprised at least 1% of the isolates at all four sites were Clavibacter, Flavobacterium and Microbacterium. Arthrobacter also comprised a significant portion of the bacterial isolates in the bentgrass rhizosphere, but not the bermudagrass rhizosphere. At the species level, there were five that comprised at least 1% of the isolates at each location; B. cereus, B. megaterium, C. michiganensis, F. johnsoniae and P. putida. <br /> <br /> Loynachan (IA): A large diversity of mycorrhizal fungi was documented in 8 soils of 4 soybean fields. Large variability exists within a single soil within a field. The composition of individual species varied within soil samples collected 3 m apart. Five species (Glomus. claroideum, G. etunicatum, G. mosseae, G. viscosum, and Paraglomus occultum) were detected in all samples collected.<br /> <br /> Baird (MS): Sweetpotato storage rots in Mississippi have increased dramatically causing substantial economic losses. Identification of root microorganisms of sweetpotato roots could support determination of a causal agent(s), or be used to systematically assess biologically based management. Microflora appear to shift in relative abundance between the growing cycle and harvest. Bacillus spp., L. enzymogenes, and P. lentimorbus accounted for more than 50% of bacteria identified. The majority of fungi were Macrophomina phaseolina, Aspergillus spp., Trichoderma spp., and a large number of Fusarium spp. In early season samples from seed stock and bedding plants, the community is primarily composed of Fusarium sp., nearly 70%. Post-harvest samples show differences in relative abundance of the dominant species, with M. phaseolina increasing to an average of 6.5% in samples taken from storage, and Fusarium spp. decreasing to an average of 27% between 60 and 90 days post-harvest.<br /> <br /> The impact of fertility, fungicides, and heavy metals were also being investigated by project scientists.<br /> <br /> Canaday (TN): Effects of chloride and ammonium salts on the incidence of Phytophthora root and stem rot of soybean were found to change the micro-partitioning of calcium in the soybean roots using energy-dispersive X-ray analysis. Use of sulfate of potash instead of muriate of potash increased plant stands by over 12% and increased snap bean yield by over 50%. Potash fertilizer form, fertilizer application timing, and seed treatment materials were evaluated for control of charcoal rot of soybean (caused by M. phaseolina) in both May-planted and June-planted field tests. Detectable levels of calcium in the outer cell layers of 10 to 13 day-old soybean taproots were significantly lower when plants were grown in soil treated with potassium chloride, sodium chloride, or magnesium chloride than when the plants were grown in untreated soil or soil treated with potassium sulfate. <br /> <br /> Fungicide hormesis was assessed in vitro on Pythium irregulare, Sclerotinia homoeocarpa, and Botrytis cinerea, validating the stimulation of pathogen activity by low fungicide levels. . It was demonstrated that sublethal doses of two fungicides, cyazofamid and propamocarb, as well as ethanol induce stimulation in Pythium aphanidermatum and that hormesis is involved in the stimulation processes. It was also demonstrated that low doses of ethanol induce stimulation in Rhizoctonia zeae and that hormesis is involved in the stimulation process, while propiconazole at sub-lethal doses do not have hormetic effects on either R. zeae or on R. solani. <br /> <br /> Loynachan (IA): Six heavy metals (Cd, Co, Cr, Cu, Ni, and Pb) reduced plant weights, nodulation, and N uptake on the legumes Vicia faba, Trifolium alexandrium, and Glycine max with increasing heavy metal concentrations, from 0 to as high as 4.3 mmol per kg soil. From the slopes of these lines, the concentrations of each metal required to produce 50% reduction in the parameter were calculated. Results showed that values varied among the soils and legumes studied but, in general, the lowest metal concentrations for 50% reduction (i.e., the most toxic) heavy metals were Cd and Pb.<br /> <br /> The value of cultivar resistance was examined on watermelon.<br /> <br /> Keineth (SC): The value of the rootstocks Emphasis, Macis (Lagenaria siceraria),Carnivor and Strong Tosa (Cucurbita moschata x Cucurbita maxima) for control of Fusarium wilt in watermelon were evaluated in a field experiment in South Carolina. The rootstocks were grafted with seedless watermelon Fascination (Citrullus lanatus var. lanatus), a new cultivar which is resistant to Fusarium oxysporum f. sp. niveum race 1 and susceptible to race 2. Plants were transplanted in a field known to be naturally infested with race 2 of Fusarium oxysporum f. sp. niveum. Wilt incidence was highest in the nongrafted and self-grafted Fascination with a mean of 81%. The four rootstocks had 2 to 16% wilted plants and did not differ from each other but differed from the control treatments. The four rootstocks also significantly increased the number of marketable-sized fruit compared to the control treatments. <br /> <br /> Objective 3. Examine the genetic diversity of Rhizoctonia solani between natural ecosystems and agricultural ecosystems. <br /> <br /> Committee members evaluated protocols for recovery of anastomosis groups (AG) of Rhizoctonia solani from soil using two techniques (soil pelleting and toothpick baits) and three media. M. Elliott found the poorest growth on water agar and best growth on Ko and Hora medium. B. Ownley also reported best results with Ko and Hora medium. C. Rothrock and A. Keinath reported that the ethanol-potassium nitrate medium was best. A. Keinath and K. Seebold reported that toothpick isolation was better than soil pellets. Keinath found that prochloraz slowed the growth of R. solani from soil and that rifampicin was a suitable substitute for tobramycin. Contamination was a problem with the Ko and Hora medium. Anastomosis groups detected were similar for the two selective media, two techniques, and seedlings. The toothpick method was an effective method for the recovery of R. solani from soils with diverse cropping histories because no special tools and few units were needed to detect the population. B. Ownley found the rates of damping-off were consistent with the population levels of R. solani detected in the three soils. Rhizoctonia solani was re-isolated from all damped-off broccoli seedlings. In Arkansas, the diversity of Rhizoctonia spp. in rice-soybean cropping systems included R. solani AGs 11, 1-1A, 4 and 2-1, Rhizoctonia oryzae and bi-nucleate Rhizoctonia species. <br /> <br /> Cubeta (NC): A field population of R. solani anastomosis group 3 (AG-3) consisting of 59 isolates was examined to determine the genetic diversity and evolutionary history of the genetic element M2 dsRNA This genetic element is hypothesized to be associated with altering the expression of metabolic pathways involved in parasitic and saprobic activity of the fungus. The distinct lineages and patterns of evolution inferred with coalescent analyses were unique for the M2 dsRNA genome. The M2 dsRNA genetic element was detected in representative isolates belonging to three anastomosis groups (AG) of R. solani (AG-1-IA, AG-4, and AG-6; teleomorph = Thanatephorus) and four AG of binucleate Rhizoctonia (AG-A, AG-F, AG-R, and AG-U; teleomorph = Ceratobasidium) using reverse-transcription polymerase chain reaction (PCR). Phylogenetic analysis of M2 dsRNA sequence data resulted in seven inferred haplotypes and there was no unique association with AG to support co-evolution of the M2 dsRNA haplotype within the fungal host. Based on the rooted gene genealogy inferred from coalescent analyses, the ancestral M2 dsRNA haplotype most likely evolved in R. solani AG-1-IA and has recently been acquired by isolates of Ceratobasidium. This is likely the first report of a dsRNA occurring in isolates of binucleate Rhizoctonia and other AG of R. solani. Horizontal transmission of the 3.57 kb M2 dsRNA between mycelia of somatically incompatible isolates of Rhizoctonia solani AG-3, an economically important pathogen of cultivated plants in the family Solanaceae, was investigated. The frequency of transmission observed between 72 pairings of the eight donor and three recipient isolates was approximately 4% of the total pairings and differences in the phenotype of the recipient isolates after acquisition of the M2 dsRNA via horizontal transmission were observed. Maintenance of the dsRNA in the recipient isolates was not stable following repeated sub-culturing on nutrient medium. <br /> <br /> Rothrock (AR):The spatial distribution of Rhizoctonia spp. in fields undergoing rice and soybean rotations is being characterized. Rhizoctonia aerial blight of soybean is a disease caused by Rhizoctonia solani AG1-IA. This pathogen also causes sheath blight of rice. Populations and disease assessments were characterized in producers field on a spatial scale by using a number of GPS positions to represent the topography of the field intermittent of the rice levee positions. Directional distribution ellipses for distribution of R. solani AG1-IA using soil and plant samples indicated agreement with drainage. Across years, distribution of R. solani AG1-IA appears to be controlled by levee position. Where levees do not form logical areas of collection, the greatest concentration of inoculum appears to be in the lower elevations of the field. <br /> <br /> Detection techniques were developed for Rhizoctonia and other pathogens<br /> <br /> Cubeta (NC) - The nuclear intergenic spacer (IGS) and a portion of the mitochondrial large subunit (mtLSU) ribosomal DNA (rDNA) genes from Sclerotinia minor and closely related species were amplified with PCR to develop specific primers. Both sets of primers did not amplify DNA of peanut (Arachis hypogaea), Botrytis cinerea, S. homeocarpa, S. sclerotiorum, and S. trifoliorum. <br /> <br /> Dick (OH): A biochemical assay was developed to detect Phytophthora sojae infestation in soil. The approach was based on profiling total cellular fatty acid methyl esters (FAME) of P. sojae in pure culture. A total of 12 fatty acids (14:0, 16:0, 18:0, 16:1 É7, 18:1 É9, 18:2 É6, 18:3 É6, 20:1 É9, 20:3 É6, 20:4 É6, 22:1 É9 and 24:1 É9) were identified in the FAME profiles of P. sojae (races 1, 4 and 7) pure cultures. The potential of using FAME profiles of P. sojae for detecting the pathogen in soil was evaluated by adding a known number of zoospores of P. sojae to soil. The results showed that fatty acids such as 18:1w9, 18:2w6, 20:1w9, 20:4w6 and 22:1w9 could be detected and quantified against the background levels of fatty acids present in soil. This outcome is significant because it offers the potential for a simple and rapid method for determining P. sojae infestations in soil.<br /> <br /> Borders (NH): The diversity of Rhizoctonia solani isolates associated with wheat, canola, soybean, and drybean by phylogenetic analysis. <br /> <br /> Garzon (OK): DNA fingerprinting protocols (AFLP, ISSR, SSR) were standardized for analysis of R. solani populations. Peanut populations of Sclerotinia from Oklahoma were characterized using AFLP and ISSR. Multiple strains of the Pythium irregulare species complex from diverse locations and hosts were characterized. Phylogenetic analyses of this clade showed evidence that multiple cryptic species exist. The genetic fingerprinting of multiple isolates of Phymatotrichopsis omnivora from Oklahoma, New Mexico, Texas and Arizona was completed. The genetic fingerprinting was completed and population genetic analyses performed of multiple isolates of Fusarium oxysporum f. sp. palmarum in collaboration with Monica Elliot, University of Florida. <br />

Publications

Peer-reviewed<br /> <br /> Aliferis, K.A., Cubeta, M.A. and Jabaji, S. 2013. Chemotaxonomy of fungi in the Rhizoctonia solani species complex using GC/MS metabolic profiling. Metabolomics 9(Suppl.): 159-169.<br /> <br /> Bartz, F.E., Danehower, D.A., Glassbrook, N., and Cubeta, M.A. 2013. Modulation of the phenylacetic acid metabolic complex by quinic acid alters the disease-causing activity ofRhizoctoniasolanion tomato. Phytochemistry 89:47-52..<br /> <br /> Bartz, F.E., Danehower, D.A., Glassbrook, N., and Cubeta, M.A. 2012. Elucidating the role of phenylacetic acid and hydroxy- and methoxy- phenylacetic acid derivatives in the pathogenic activity of Rhizoctonia solani AG-3. Mycologia 104:793-803.<br /> <br /> Burchhardt, K. and Cubeta, M.A. 2012. Microsatellite marker development for the plant pathogenic fungus Monilinia vaccinii-corymbosi. Molecular Ecology Resources 12:(In Press).<br /> <br /> Copes, W., Rodriguez-Carres, M., Toda, T., Rinehart, T.A., and Cubeta, M.A. 2011. Seasonal prevalence of species of binucleate Rhizoctonia fungi in growing medium, leaf litter, and stems of container grown azalea. Plant Dis. 95:705-711.<br /> <br /> Hansen, Z. R., and Keinath, A. P. 2013. Increased pepper yields following incorporation of biofumigation cover crops and the effects on soilborne pathogen populations and pepper diseases. Appl. Soil Ecol. 63: 6777.<br /> <br /> Kaye, A.C., Moyer, J.W., Parks, E.J., Carbone, I., and Cubeta, M.A. 2011. Population genetic analysis of Tomato spotted wilt virus on peanut in North Carolina and Virginia. Phytopathology 110:147-153.<br /> <br /> Keinath, A. P., Hassell, R. L., and DuBose, V. B. 2012. Field evaluation of six cucurbit rootstocks to manage Fusarium wilt on triploid watermelon, 2011. Plant Disease Management Reports 6:V024. doi: 10.1094/PDMR06.<br /> <br /> McCormick, M.A., Cubeta, M.A., and Grand, L.F. 2013. Geography and hosts of the wood decay fungi Fomes fasciatus and Fomes fomentarius in the United States. North American Fungi 8(2): 1-53.<br /> <br /> McCormick, M.A., Grand, L.F., Post, J.B., and Cubeta, M.A. 2013. Phylogenetic relatedness and phenotypic characterization of Fomes fasciatus and Fomes fomentariussampled from the United States. Mycologia 76: (Accepted). <br /> <br /> Richter, B. S., D. M. Benson, and K. L. Ivors, 2011. Microbial profiling of cultural systems for suppression of Phytophthora root rot in Fraser fir. Plant Dis. 95:537-546.<br /> <br /> Richter, B. S., Ivors, K. I., Shi, W., and Benson, D. M. 2011. Cellulase activity as a mechanism for suppression of Phytophthora root rot in mulches. Phytopathology 101:223-230.<br /> <br /> Samuels, G., Ismaiel, A., McMahon, P., Guest, D., Rosmana, A., Junaid, M., Rodriguez-Carres, M., and Cubeta, M.A. 2012. Vascular streak dieback of cacao in Southeast Asia and Melanesia: in planta detection of the pathogen and a new taxonomy. Fungal Biology 116(1):11-23.<br /> <br /> Thomas, E., Pakala, S., Fedorova, N. D., Nierman, W. C., and Cubeta, M. A. 2012.Triallelic SNP-mediated genotyping of regenerated protoplasts of the heterokaryotic fungus Rhizoctonia solani. Journal of Biotechnology 158:144-150.<br /> <br /> Toda, T., Strausbaugh, C., Rodriguez-Carres, M., and Cubeta, M.A. 2011.Characterization of a basidiomycete fungus from sugarbeet. Mycologia 104:70-78.<br /> <br /> Woodhall, J.W., Webb, K.M., Harper, G., Peters, J.C., Rodriguez-Carres, M., and Cubeta, M.A. 2011. First report of a new binucleate Rhizoctonia in UK potato tubers. New Disease Reports 23:31 [doi:10.5197/j.2044-0588.2011.023.031].<br /> <br /> Yin, J., Koné, D., Rodriguez-Carres, M. Cubeta, M.A., Burpee, L.L., Fonash, E.G. Csinos, A.S., and Ji, P. 2011. First report of root rot caused by binucleate Rhizoctonia anastomosis group F on Musa spp. Plant Disease 94:490.<br /> <br /> Abstracts<br /> <br /> Hansen, Z. R., and Keinath, A. P. 2012. Increased pepper yields following incorporation of biofumigation cover crops and their effects on soilborne pathogen populations and pepper diseases. (Abstr.). Phytopathology 102:S4.49.<br /> <br /> Keinath, A. P., and Hassell, R. L. 2012. Grafting on hybrid squash and bottle gourd rootstocks to manage Fusarium wilt of watermelon. (Abstr.) Phytopathology 102:S4.153.<br /> <br /> Other publications<br /> <br /> Hansen, Z. R. 2011. Potential of three Brassica cover crops for biofumigation in the field and the relationship between soil myrosinase and biofumigation efficacy. Clemson University, M.S. Thesis. UMI Number 1505535<br /> <br /> Loynachan, T. E. 2012. Life in the soil: Who cares? pp. 28-30. In Deborah McDonough (ed.) Getting Into Soil and Water. Iowa Water Center, Ames, IA.<br /> <br /> Seebold, K., 2012. Evaluation of a Biopesticide and Conventional Fungicides for Management of Phytophthora Blight of Yellow Squash. Pp. 32-34 in: 2011 Fruit and Vegetable Crops Research Report (PR-626). T. Coolong, J. Snyder, and C. Smigell, eds. UK Cooperative Extension Service, College of Agriculture, 53 pp<br /> <br /> Target Audiences<br /> <br /> This project was focused on evaluating commercial and non-commercial microbial biological control agents for control of soilborne plant pathogens, examining the effect of cultural practices on soilborne pathogens and plant growth, and examining the genetic diversity of Rhizoctonia solani between natural ecosystems and agroecosystems. Findings were shared with researchers and companies that provided the biological control agents for testing, and with collaborators of the Multistate Regional Research Project S-1028 in annual meetings. Highlights of this project were included in the S-1028 annual report, which was disseminated via the National Information Management and Support System (NIMSS) website. Findings were also shared with other scientists and students through research presentations at the American Phytopathological Society and other scientific meetings. Target audiences include producers of a variety of commodities, agricultural consultants, agricultural industry representatives, extension specialists, and other agricultural scientists interested in plant disease management. Producers/consultants use this information for decisions concerning disease management (product selection/cultural practices) in row crops. <br />

Impact Statements

1. Root-colonizing and endophytic biocontrol fungi effectively induced resistance in host plants against foliar pathogens, including induction of resistance in cotton by Beauveria bassiana 11-98 against Xanthomonas axonopodis pv. malvacearum and induction of resistance in geranium by binucleate Rhizoctonia isolates BNR621 and P023, and Trichoderma hamatum 382 against Botrytis cinerea. These observations increased awareness of potential benefits from application of biocontrol agents.
2. The cash value of vegetable crops was $20.5 billion in the U.S., with estimated losses of up to 10% due to soilborne plant pathogens. For growers intent on not using pesticides, production of pesticide-free or organic crops can increase crop value by 30%. Effective biopesticide treatments were identified in the regional broccoli experiment. A commercial formulation of Trichoderma viride and T. harzianum, applied to soil by drip irrigation, was shown to suppress the incidence of Phytophthora blight on summer squash. Although not as effective as commercially available fungicides, using the biocontrol agents in conjunction with commercial fungicides as investigated with broccoli. tomato, watermelon, and squash may permit fewer applications or allow for reduced rates of the fungicide products. These treatments have the potential to increase yield of field grown vegetable crops and greenhouse grown transplants.
3. Results have lead to greater adoption of cover crops in the southeast. Hairy vetch was adopted by a large watermelon grower in South Carolina as a winter cover crop in fields with a history of losses to Fusarium wilt. Acreage planted to hairy vetch increased three-fold from 2007 to 2008. Brassica winter cover crops were shown to reduced damage from soilborne pathogens on vegetable and field crops and increase yields. Brassica cover crops appear to be a viable alternative to the application of fumigants.
4. Research on Rhizoctonia solani diversity has characterized a greater diversity in many crops that previously known. This diversity may lead to changes understanding the etiology of disease and impact growers, and industry and academic researchers as they move forward in developing chemical, biological, and cultural control strategies.
5. A efficient effective protocol developed for the assay of Rhizoctonia solani from soils should allow better detection of the pathogen in soil and the examination of the diversity and distribution of this important pathogen across different native and agricultural systems increasing our understanding of the role of cropping history and transport of the pathogen in disease development and diversity.
6. Techniques were developed to characterize Rhizocotonia solani and other soilborne pathogens, including microsatellite markers and highly sensitive PCR and real-time PCR specific for anastomosis groups. The genetic element M2 dsRNA, which likely evolved in R. solani AG-1-IA, was detected in isolates of binucleate Rhizoctonia spp. and other AG of R. solani. Diagnostic primers were designed for multiple Pythium, Rhizoctonia, and Sclerotinia spp.
7. In terms of planted acreage, snap beans are Tennessees number one vegetable crop. Farm receipts for this crop typically total over $9,000,000 per year. Instead of using untreated seed and muriate of potash, growers could potentially double their snap bean yield and gross returns if they used a seed treatment of trifloxystrobin + metalaxyl coupled with use of sulfate of potash. Changing potash form from muriate of potash to sulfate of potash reduced soybean preemergence damping-off and increased yield by over 180 kg/ha (2.68 bu/A).
8. Communities of microorganisms associated with multiple production systems have been characterized to identify beneficial microorganisms or pathogens and has lead to the identification of novel biocontrol agents and new antibiotics.
9. Grafting of watermelon is the most promising alternative to soil fumigation to control Fusarium wilt of watermelon. Grafting was demonstrated to be effective against Fusarium wilt race 2, the race for which there are no resistant cultivars. Grafting significantly increased marketable yield; fruit produced on grafted plants reached marketable size. Based on a cost of $1.00 per grafted transplant, grafted transplants are economically feasible, because of the yield increase with grafted plants, even in soil not heavily infested with Fusarium.
10. Human activities may contribute to the contamination of soils by heavy metals. Studies suggests that heavy metals in soil can have negative consequences on the nitrogen cycle: limiting plant growth, nodulation by beneficial bacteria, and uptake of the essential plant nutrient N. Results will allow informed decisions on the impact of heavy metals in the soil environment.
11. Results demonstrating the value of mulches for disease suppression of Phytophthora root rot should provide North Carolina Christmas Tree Growers options for disease suppression, since fungicides are not economically viable for disease management.

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