Progress 11/19/07 to 11/18/12
Outputs
Progress Report Objectives (from AD-416): Objective 1: Optimize delivery and evaluate performance of chemical, cultural, biological, and genetic alternatives to methyl bromide for crops/pathogen combinations currently benefiting from the use of methyl bromide. a. Evaluate alternative fumigants. b. Identify genes associated with pathogenicity of Verticillium dahliae based on a Verticillium comparative genomics study to support the development of alternative control procedures. c. Identify emerging diseases and their etiology and evaluate resistant germplasm for diseases of strawberry, lettuce, and vegetables. d. Evaluate the influence of crop rotation on pathogen populations and beneficial microbial community dynamics in the soil and severity of Verticillium wilt and other diseases. Develop tools to evaluate and understand the role of myxobacteria in agricultural and natural ecosystems in an effort to elucidate their potential for pathogen control. Objective 2: Develop molecular diagnostic tools for the identification of emerging diseases of vegetables and strawberries, and use these tools in the development of management strategies as alternatives to methyl bromide. Approach (from AD-416): Develop integrated management approaches including crop rotation, biological control, selection of disease resistant varities, organic production, other biological practices, and combinations of biological practices with reduced concentrations of chemical fumigants to control diseases. Identifying useful commercial biological agents as well as new agents from the strawberry rhizosphere that will help to improve plant growth and disease management. Molecular tools will be developed for evaluation of the fficacy of pathogen management and modeling population dynamics of beneficial microbes. Replaces 5305-22000-009-00D(11/07). This project expired 11/18/12, approximately seven weeks into the reporting period so additional details on accomplishments may be found in the report for the bridging project (5305-22000-014-00D) as the transition into NP303 takes place.
Impacts
(N/A)
Publications
|
Progress 10/01/11 to 09/30/12
Outputs
Progress Report Objectives (from AD-416): Objective 1: Optimize delivery and evaluate performance of chemical, cultural, biological, and genetic alternatives to methyl bromide for crops/pathogen combinations currently benefiting from the use of methyl bromide. a. Evaluate alternative fumigants. b. Identify genes associated with pathogenicity of Verticillium dahliae based on a Verticillium comparative genomics study to support the development of alternative control procedures. c. Identify emerging diseases and their etiology and evaluate resistant germplasm for diseases of strawberry, lettuce, and vegetables. d. Evaluate the influence of crop rotation on pathogen populations and beneficial microbial community dynamics in the soil and severity of Verticillium wilt and other diseases. Develop tools to evaluate and understand the role of myxobacteria in agricultural and natural ecosystems in an effort to elucidate their potential for pathogen control. Objective 2: Develop molecular diagnostic tools for the identification of emerging diseases of vegetables and strawberries, and use these tools in the development of management strategies as alternatives to methyl bromide. Approach (from AD-416): Develop integrated management approaches including crop rotation, biological control, selection of disease resistant varities, organic production, other biological practices, and combinations of biological practices with reduced concentrations of chemical fumigants to control diseases. Identifying useful commercial biological agents as well as new agents from the strawberry rhizosphere that will help to improve plant growth and disease management. Molecular tools will be developed for evaluation of the efficacy of pathogen management and modeling population dynamics of beneficial microbes. Research is ongoing to examine the localization of the fungus Verticillium dahliae in spinach seeds, flowers, pollen, and other plant parts, in collaboration with the UC,Davis. Research was undertaken to characterize the functions of genes identified in the comparative analyses of the genomes of fungal vascular wilt pathogens. Research was initiated to evaluate the utility of a DNA-based assay for the quantification of disease resistance in spinach. DNA sequence analyses of lettuce gene homologs associated with plant resistance or genetic responses to the fungus, V. dahliae, and collaboration with the UC Davis is ongoing to further characterize the potential role of three of these plant genes in disease resistance. Research was conducted to assess the involvement of a gene from V. dahliae in pathogenicity, and experiments to assess plant gene expression in response to colonization by this particular mutant strain. Analyses of transposable elements in the fungal pathogen, V. dahliae is ongoing. Research was initiated on detection of Peronospora effusa, the downy mildew pathogen of spinach. Completed pathogenicity and host range testing needed to expand Pseudomonas cannabina to include Pseudomonas syringae pv. coriandricola and P. syringae pv. philadelphi. Began in situ testing of QPCR methods with molecular markers for S. suberifaciens. Bacterial leaf spot of radicchio (Cichorium intybus) is caused by isolates of Xanthomonas hortorum related to the bacterial leaf spot pathogen from lettuce which presents implications for disease management. Analyzed the comparative genomics of closely related pathogens in the species P. cannabina to provide insights into mechanisms of host-pathogen interaction, differential virulence factors, and pathogen evolution and find target genes for developing PCR- based detection and quantification protocols. Identified a novel species causing bacterial blight on garlic in Brazil and identified potential sequences for the development of PCR-based detection and quantification protocols. Identified novel pathovars of Pseudomonas syringae causing bacterial blight and leaf spots on cucurbits. Demonstrated that leaf spot of Australian Cedar is caused by Xanthomonas axonopodis. Completed preliminary experiments that suggest that there are interactions between Xanthomonas campestris pv. vitians strains and lettuce cultivars for disease severity and pathogen growth/survival. Demonstrated that P. syringae pv. apii has a broader host range than previously reported and documented the occurrence of this pathogen causing disease in cilantro fields. Correlations between the mitochondrial haplotype and nuclear AFLP and SSR analysis of 100+ isolates of Phytophthora cinnamomi representing a worldwide collection is near completion. Finishing up analysis of field samples collected as part of the national survey for Phytophthora ramorum conducted through the USDA-FS using our Phytophthora real time PCR diagnostic markers. A single tube nested amplification that can be done directly from infected tissue is under development and validation for identification of Phytophthora ramorum mitochondrial haplotype. Significant Activities that Support Special Target Populations: One Scientist won the Non-supervisory 2012 Agricultural Research Station Adminstrator�s Outreach, Diversity, and Equal Opportunity Award �For innovative program development and outstanding mentoring provided to minority and women undergraduate researchers in the Salinas Valley of California.� One Scientist won the Inaugural Mentor of the Year Award from the Undergraduate Research Opportunities Center at California State University Monterey Bay, one Scientist was recognized in the City of Champions program along with her students by Mayor of Salinas, California, for outstanding service to the community in growing scientists. One Scientist mentored four women and Hispanic interns from Hartnell College and California State University Monterey Bay (two largely Hispanic serving institutions) as part of a United States Department of Agriculture funded program to increase the exposure of minority (specifically Hispanic) students to science. This Scientist mentored two women students from University of California, Davis, one local high school student and a faculty member from Hartnell College. One Scientist developed a training program for student interns at the United States Department of Agriculture/ Agriculture Research Station One Scientist presented �Growing Scientists in the Salad Bowl of the United States.� Science-Technology-Engineering-Mathematics education series at the Carnegie Science Center and the Pittsburgh Regional Center for Science Teachers. One Scientist serves as the Director of the Advisory Board for the Hartnell College Math and Science Institute. One Scientist serves as an Advisory Board for the Undergraduate Research Opportunities Center for California State University Monterey Bay. Two Scientists planned innovative programming for fifth graders from Los Padres Elementary School (an underperforming mostly Hispanic serving school) to introduce the students to the scientists in their neighborhood. This year the program included 12 visits by as many scientists and students and an end of year field trip to the United States Department of Agriculture/ Agriculture Research Station in Salinas, California. Two Scientists Participated in the Health and Science Fair organized by Hartnell College and Alisal School District (largely Hispanic serving institutions). One Scientist lead visits to the research station for Hartnell College Biology Class, 2 Middle School Career Camps; and 2 High School Equivalency Program Classes. One Scientist and current and former students planned the first ever session on plant pathology for the Society for the Advancement of Chicanos and Native Americans in Science national meeting. The American Phytopathological Society provided a booth used to promote agricultural research in general and plant pathology specifically. One Scientist served as the Agriculture Research Station representative to The United States Department of Agriculture Science, Technology, Engineering, and Mathematics (STEM) Professionals event at Cabrio College designed to expose students to career options in STEM fields within United States Department of Agriculture. One Scientist serves on the American Phytopathological Society Committee on Diversity and Equality. Two Scientists mentored students (one minority student) for a science fair project, for local and state competition. Accomplishments 01 �Light�- tagged reporter-phage detects and quantifies living bacterial plant pathogens on plants. ARS researchers from Salinas, California, and South Carolina in collaboration with a Cooperative Research and Development Agreement partner demonstrated proof of concept for a novel approach for detecting and quantifying bacterial plant pathogens. This method can be adapted to other plant pathogens. The phage will only infe and produce light in living cells. Thus, this method is highly specific and allows for detection and quantification of living cells of specific bacterial plant pathogens. 02 Quantification of Verticillium dahliae in spinach seeds. The movement o the fungal plant pathogen in spinach seeds from seed production areas introduces pathogen inoculum and exotic strains to crop production areas An ARS researcher in Salinas, in collaboration with researchers at the University of California, led the research for the development of an ass for the quantification of Verticillium dahliae in spinach seeds. The ass will be useful to quickly assess the level of seed infection in commerci seed lots and be helpful to limit the spread of the pathogen from seed production areas, to areas where susceptible crops are grown in rotation with spinach. The assay may also be useful as an additional tool to asse the level of plant resistance to V. dahliae. 03 Quantification of the pathogen Verticillium dahliae in the soil. V. dahliae causes vascular wilt in a number of plant species and crops like strawberry which are very sensitive to low levels of this pathogen, so having the ability to rapidly and accurately determine inoculum levels i the soil would help growers make planting decisions. In collaboration wi a researcher at the University of California, ARS researchers in Salinas completed validation of a real time PCR TaqMan quantification assay for determining soil populations of V. dahliae. To help provide access to th assay for the growers, the specifics of the technique and materials need to validate results were provided to labs that perform diagnostic servic for the strawberry industry prior to manuscript publication. Prior techniques for soil quantification involved plating soil on culture medi but did not always provide accurate quantification and took between 6-8 weeks, so the development of this molecular assay should improve the ability of the growers to identify risk before planting. 04 Organic Strawberry Production Manual produced. Collaboration by ARS researchers in Salinas, CA, Co-Editors from the University of California and authors from various institutions led to the production of this usef manual. Producers had little scientific data to use for improving organi strawberry production. This manual provides a snapshot of organic strawberry research findings. The manual will allow growers to improve production and highlights gaps in knowledge as targets for future resear projects. 05 Analyses of the function of a glucosyl transferase in Verticillium dahli The plant pathogenic fungus Verticillium dahliae causes disease in ove 200 species of plants, including high value crops such as strawberry. An ARS researcher in Salinas, California, along with a team of researchers, deleted a gene encoding a glucosyl transferase in Verticillium dahliae a observed that the mutant strain was not pathogenic on one plant host. Th glucosyl transferase is widely conserved among bacteria, but present in only a few genera of pathogenic fungi, suggesting that the gene may have been acquired through a mechanism involving a horizontal genetic transfe from bacteria to fungi. The bacterial glucosyl transferase is involved i responses to osmotic stress, and the fungal homolog of the gene may function similarly. Identification of the genetic basis for pathogenicit or host range expansion may lead to alternative strategies for their control in high value crops such as lettuce. 06 Pseudomonas cannabina pv. alisalensis caused blight on radish in Germany Pseudomonas cannabina pv. alisalensis had not previously been reported from continental Europe. Bacterial blight caused by P. cannabina pv. alisalensis causes significant economic damage. ARS researchers in Salin California, in collaboration with collaborators from The Netherlands expanded the known geographic range of this disease to continental Europ by documenting the first occurrence of this disease there. This information demonstrated the need for management practices and detection practices for this pathogen world-wide. 07 Pseudomonas cannabina pv. apii causes bacterial streak in fennel (Foeniculum vulgare). ARS researchers in Salinas, CA in collaboration wi scientists from the University of California determined that bacterial streak in fennel was caused by an important pathogen of celery. Bacteria streak symptoms spread down the stems leading to unmarketable plants due to bulb decay. This pathogen also causes disease in parsley. This information was incorporated into recommendations for crop rotation practices to avoid other hosts of this pathogen. 08 Examination of a hybrid Phytophthora species. The ability of Phytophtho species to form interspecific hybrids and the impact these have on causi new disease has been of significant interest in the research and regulatory community. Phytophthora infestans has a significant impact o potato and tomato production worldwide and there is a new species, P. andina, that was believed to be a hybrid between P. infestans and a another unidentified species. A collaboration between ARS, University o California at Riverside and Franklin and Marshall College researchers examined a collection of these and closely related species using 11 genetic markers in an attempt to identify the other parental isolate and better understand the genetic background of the hybrid isolates. The results provided insight to better understand the process of species hybridization in the genus. 09 Development of web site to support Phytophthora research. The genus Phytophthora is responsible for many diseases of crop plants worldwide a consists of approximately 117 species that are difficult to identify. AR researchers at Salinas, California, and other ARS locations, and university researchers developed a web-based database for Phytophthora research to enhance the understanding of the genus, simplify species identification and stimulate further research on the genus. The database includes complete morphological descriptions, information on host range and geographical distribution, a comprehensive molecular phylogeny using seven nuclear and four mitochondrial genes, a sequence database with ove 6,900 entries of that is searchable by Basic Local Alignment Search Tool analysis, and a section on molecular identification and detection. This database will serve as a resource for researchers working on the genus, means to simplify sequence based identification of species, as well as a repository for future work.
Impacts
(N/A)
Publications
- Atallah, Z.K., Maruthachalam, K.K., Vallad, G.E., Davis, M., Klosterman, S. J., Subbarao, K.V. 2011. Analysis of Verticillium dahliae suggests a lack of correlation between genotypic diversity and virulence phenotypes. Plant Disease. 95:1224-1232.
- Martin, F.N., Coffey, M. 2012. Mitochondrial haplotype analysis for differentiation of isolates of Phytophthora cinnamomi. Journal of Phytopathology. 102:229-239.
- Bilodeau, G.J., Koike, S., Uribe, P., Martin, F.N. 2012. Development of an assay for rapid detection and quantification of Verticillium dahliae in soil. Phytopathology. 102:331-343.
- Jardini, T.M., Koike, S.T., Bull, C.T. 2011. First report of bacterial streak of fennel (Foeniculum vulgare) in California caused by Pseudomonas syringae pv. apii. Plant Disease. 96:285.
- Duressa, D.O., Rauscher, G.M., Koike, S.T., Mou, B., Hayes, R.J., Maruthachalam, K., Subbarao, K.V., Klosterman, S.J. 2012. A real-time PCR assay for detection and quantification of verticillium dahliae in spinach seed. Phytopathology. 102:443-451.
- Blair, J.E., Coffey, M.D., Martin, F.N. 2012. Species tree estimation for the late blight pathogen, Phytophthora infestans, and close relatives. PLoS One. 7(5):e37003.
- Bull, C.T., Martin, F.N. 2012. Cultivar Selection for Pest Management in Organic Strawberry Production. In: Koike, S. T., Bull, C. T., Bolda, M., Daugovish, O., editors. Organic Strawberry Production Manual. University of California Division of Agriculture and Natural Resources. p. 15-24.
- Bull, C.T., De Boer, S.H., Denny, T.P., Firrao, G., Fischer-Le Sau, M., Saddler, G., Scortichini, M., Stead, D.E., Takikawa, Y. 2012. List of New Names of Plant Pathogenic Bacteria (2008-2010). Journal of Plant Pathology. 94:21-27.
- Klosterman, S.J., Subbarao, K.V., Kang, S., Veronese, P., Gold, S.E., Thomma, B.P.H.J, Chen, Z., Henrissat, B., Lee, Y., Park, J., Garcia- Pedrajas, M.D., Dez, B.J., Anchieta, A.G., De Jonge, R., Parthasarathy, S., Maruthachalam, K., Atallah, Z.K., Amyotte, S., Paz, Z., Inderbitizin, P., Hayes, R.J., Heiman, D.I., Young, S., Zeng, Q., Engels, R., Galagan, J., Cuomo, C., Dobinson, K.F., Ma, L-J. 2011. Comparative genomics yields insights into niche adaptation of plant vascular wilt pathogens. PLoS Pathogens. 7(7): e1002137.
- Klosterman, S.J. 2012. Real-time PCR for the quantification of fungi in planta. In: Bolton, M.D., Thomma, B.P.H.J., editors. Plant Fungal Pathogens: Methods and Protocols. Series: Methods in Molecular Biology. Volume 835. New York, NY: Humana Press. p. 121-131.
- Martin, F.N., Abad, Z., Balci, Y., Ivors, K. 2012. Identification and detection of Phytophthora: Reviewing our progress, identifying our needs. Plant Disease. 96(8):1080-1103.
- Rubio, I., Hiddink, G., Asma, M., Bull, C.T. 2012. First report of the crucifer pathogen Pseudomonas cannabina pv. alisalensis causing bacterial blight on radish (Raphanus sativus) in Germany. Plant Disease. 96:904.
- Schofield, D.A., Bull, C.T., Rubio, I., Wechter, W.P., Westwater, C., Molineux, I.J. 2012. Development of an engineered �bioluminescent� reporter phage for the detection of bacterial blight of crucifers. Applied and Environmental Microbiology. 78:3592-3598.
- Koike, S., Bull, C.T., Bolda, M., Daugovish, O. 2012. Organic strawberry production manual. Organic Strawberry Production Manual. 148 p.
|
Progress 10/01/10 to 09/30/11
Outputs
Progress Report Objectives (from AD-416) Objective 1: Optimize delivery and evaluate performance of chemical, cultural, biological, and genetic alternatives to methyl bromide for crops/pathogen combinations currently benefiting from the use of methyl bromide. a. Evaluate alternative fumigants. b. Identify genes associated with pathogenicity of Verticillium dahliae based on a Verticillium comparative genomics study to support the development of alternative control procedures. c. Identify emerging diseases and their etiology and evaluate resistant germplasm for diseases of strawberry, lettuce, and vegetables. d. Evaluate the influence of crop rotation on pathogen populations and beneficial microbial community dynamics in the soil and severity of Verticillium wilt and other diseases. Develop tools to evaluate and understand the role of myxobacteria in agricultural and natural ecosystems in an effort to elucidate their potential for pathogen control. Objective 2: Develop molecular diagnostic tools for the identification of emerging diseases of vegetables and strawberries, and use these tools in the development of management strategies as alternatives to methyl bromide. Approach (from AD-416) Develop integrated management approaches including crop rotation, biological control, selection of disease resistant varities, organic production, other biological practices, and combinations of biological practices with reduced concentrations of chemical fumigants to control diseases. Identifying useful commercial biological agents as well as new agents from the strawberry rhizosphere that will help to improve plant growth and disease management. Molecular tools will be developed for evaluation of the fficacy of pathogen management and modeling population dynamics of beneficial microbes. Replaces 5305-22000-009-00D(11/07). Comparative analyses of the genomes of fungal vascular wilt pathogens V. dahliae, Fusarium oxysporum, and V. albo-atrum and those of other fungi revealed candidate genes to pursue in functional analyses. Further analyses of some candidate genes identified were initiated in a collaborative effort with other researchers. Research is ongoing to quantify V. dahliae in commercial spinach seed lots. Research is ongoing to examine the localization of the fungus in spinach seeds, in collaboration with the University of California, Davis. An alternative and rapid method to prepare targeted gene deletion constructs for fungi was analyzed and verified. We continued the DNA sequence analyses of lettuce gene homologs associated with plant resistance to the fungus, V. dahliae, and initiated collaboration with the University of California, Davis to characterize the role of one these genes in disease resistance. We completed analyses of a method to distinguish between two races of V. dahliae, in collaboration with the University of California, Davis. Two genes important for colonization of lettuce were identified in V. dahliae, by insertional mutagenesis and pathogenicity assays. Research was initiated for comprehensive analyses of transposable elements in the fungal pathogen, V. dahliae. We demonstrated that P. cannabina pv. alisalensis strains from Australia were the causal agents for bacterial blight of mustard, although the pathogen was previously identified as P. syringae pv. maculicola. We completed host range and phage sensitivity tests that supported the hypothesis that bacterial blight of radish in France was caused by P. cannabina pv. alisalensis in collaboration with INRA. We demonstrated that lettuce cultivars influence population dynamics of Xanthomonas campestris pv. vitians on lettuce leaves. We identified a third pathogen from parsley as a close relative of Pseudomonas viridiflava. We identified a pathogen of fennel as Pseudomonas syringae pv. apii, thus expanding the known host range of this pathogen. We documented genomic changes in P. syringae pv. apii populations. We isolated bacteriophage specific to Xanthomonas campestris pv. vitians. A comprehensive, multigene, phylogenetic analysis of four Phytophthora species and species complexes was completed in collaboration with three other labs. A comprehensive, mitochondrial, multigene phylogeny of the genus Phytophthora was concluded. A new diagnostic marker system for Phytophthora was developed and validated with field samples from the USDA-FS P. ramorum national survey. A real time PCR soil quantification assay for Verticillium dahliae was developed and validated with field soils. Significant Activities that Support Special Target Populations One SY mentored four women (two minority) and one Hispanic student from Hartnell College and California State University-Monterey Bay (both Hispanic serving institutions), and one woman student from University of California, Davis through grant funded internship programs. Mentored two visiting women scientists (Brazil for one year and Egypt), one woman PhD student (Brazil for 1 year) and two women (faculty and staff member) from our local community college (Hartnell College). The other two SYs mentored one minority intern student each. Two SYs organized displays and participated in the Hartnell College Health and Science Day May 2011. All SYs coordinated and participated in presentations made to our adopted 5th grade classes at a local minority serving elementary school (Los Padres Elementary) to provide science programming and field trips. One SY also gave science presentations at 5th and 7th grade classes in the Spreckels School District. Participated in WISE - Women In Science Exploration of Hartnell Community College as a mentor and sponsor. Advisory Committee Member to the Undergraduate Research Opportunities Center (UROC) of California State University at Monterey Bay (a minority serving institution). Advisory Board Chair for the Hartnell College Science & Math Institute Advisory Board. Serves on the Committee for Equality and Diversity for American Phytopathological Society. Developed a scientific program for the 2011 Annual Meeting of Society for the Advancement of Chicanos and Native Americans in Science (SACNAS). Proposed and organized a booth for the American Phytopathological Society at the 2011 Annual Meeting of Society for the Advancement of Chicanos and Native Americans in Science (SACNAS). Attended the 2010 Annual Meeting of Society for the Advancement of Chicanos and Native Americans in Science (SACNAS) as a mentor. Accomplishments 01 Insight into niche adaptation of fungal vascular wilt pathogens. Two soilborne, fungal plant pathogens, Verticillium dahliae and V. albo-atru cause vascular wilt diseases on over 200 plant species worldwide, and costly soil fumigants are sometimes deployed to kill these fungal pathogens in the soil. To gain insight into niche adaptation, ARS researchers in Salinas, California, along with a team of international researchers, completed analyses of these genomes and extended the comparison with other fungi. The research discovered a set of proteins shared among three wilt pathogens, and an abundance of enzymes that degrade pectin, that may enable the two fungi to infect their host crops and allow ecological adaptation. Identification of the genetic basis for pathogenicity or host range expansion may lead to alternative strategies for their control in high value crops such as lettuce. 02 Molecular diagnostic technique for detection of Phytophthora genere and species. The fungal pathogen phytophthera causes disease on many crops. new, real time PCR molecular diagnostic technique developed by ARS researchers at Salinas, CA, for detection and identification of Phytophthora spp. exploits a gene order difference between Phytophthora and other related genera and plants. The technology allows in one reacti to determine if a Phytophthora spp. has infected the plant, and if it is the species of interest (not possible with techniques currently used). Genus-specific probes were developed for 14 economically important Phytophthora spp., including several invasive and quarantine species, using sequence data from 500+ isolates representing all species in the genus. Validation is with close to 400 field samples provided by 3 labs participating in the national survey for Phytophthora ramorum. This technique will provide a new tool for diagnosticians to determine not on if a particular species is associated with plant infection, but if any member of the genus is present in the sample as well as a systematic approach for developing new species specific markers. 03 Rapid identification of more than 60 plant pathogens demonstrated. Identification of pathogens of specific plants, also called pathovars, within the Pseudomonas syringae group has been difficult or impossible since the inception of this taxon in the 1980s. These bacterial pathogen cause diseases on thousands of plants world-wide in a pathogen/host specific manner. An ARS researcher at Salinas, California, and colleague demonstrated that MultiLocus Sequence Typing (MLST) can be used to rapid identify members of the genus Pseudomonas to the genus and pathovar leve Four genes were sequenced for all 61 known pathotypes from the P. syring group and related strains and were used to rapidly identify two pathogen from parsley. Additionally, a significant correlation between DNA/DNA hybridization values and % similarity was demonstrated. Using the method and tools described, the time required to make identifications of plant pathogenic bacteria was reduced from several months to less than one wee The sequences are publicly available and will reduce the number of strai to be moved among laboratories for identification purposes as well as speed identification of these pathogens. 04 Identification of genes associated with Verticillium wilt symptom development. Verticillium wilt disease on lettuce causes severe symptom that can render an entire field unmarketable. The objective of this research was to identify genetic responses of lettuce during wilt sympto development. ARS researchers in Salinas, California, identified sets of lettuce genes, including those associated with symptom development, whic are repressed or induced in response to infection by the plant pathogeni fungus, Verticillium dahliae. Insights on the regulation of symptom development may contribute to the development of plants that are more tolerant to symptoms associated with Verticillium wilt and other fungal diseases. 05 Reclassification of Pseudomonas syringae pv. alisalensis as P. cannabina pv. alisalensis. Appropriate and specific management strategies could n be applied to manage bacterial blight and pepper spot diseases of crucifers because the pathogens causing these diseases had been indistinguishable. MultiLocus Sequence Typing (MLST) and phenotype based analyses of P. syringae pv. alisalensis and related strains resulted in the transfer of this pathogen to a new species and the demonstration tha a well studied pathogen previously identified as P. syringae pv. maculicola is P. syringae pv. alisalensis. MLST data also provides information needed for rapid identification of the pathogens. Correct an rapid identification of these pathogens leads to more rapid appropriate disease management responses for crucifer production. This is significan for these pathogens because P. cannabina pv. alisalensis causes a severe bacterial blight on crucifers while the disease caused by P. syringae pv maculicola is significantly milder. 06 A new tool for genetic studies of Verticillium dahliae and other fungi. Targeted gene deletion in fungi enables researchers to examine the function of genes of interest. Existing methods for generating DNA constructs for gene deletions in fungi are cumbersome, often involving multiple steps. The objective of this research was to develop an alterna approach for the creation of gene deletion constructs. An ARS researcher in Salinas, California, along with University colleagues, developed and verified an alternative method to quickly generate DNA constructs for targeted deletion of genes. Thus, the tool will enable rapid preparation of constructs for the deletion of genes associated with pathogenicity of Verticillium dahliae and other fungi important to US Agriculture. 07 Web site to support Phytophthora research. The fungal genus Phytophthor is responsible for many diseases of crop plants worldwide and consists o approximately 117 species that are difficult to identify. ARS researcher at Salinas, CA, and other ARS locations, and university researchers developed a web-based database for Phytophthora research to enhance the understanding of the genus, simplify species identification and stimulat further research on the genus. The database includes complete morphological descriptions, information on host range and geographical distribution, a comprehensive molecular phylogeny using seven nuclear an four mitochondrial genes, a sequence database with over 6,900 entries of that is searchable by Basic Local Alignment Search Tool analysis, and a section on molecular identification and detection. This database will serve as a resource for researchers working on the genus, a means to simplify sequence based identification of species, as well as a reposito for future work. 08 Molecular quantification of Verticillium dahliae from soil. The pathoge V. dahliae has a broad host range and can survive for long periods of ti in the soil. Although soil plate assays can be used to quantify the pathogen populations in the soil, they are not always accurate and can take six to eight weeks to complete. The real time polymerase chain reaction (PCR) assay that was developed by ARS researchers is able to accurately determine population densities as low as 1 to 2 propagules/g soil within a day or two. This assay has the potential to significantly reduce the time needed for growers of susceptible crops to get the resul they need with greater accuracy than current techniques.
Impacts
(N/A)
Publications
- Bull, C.T., De Boer, S., Denny, T., Firrao, G., Saux, M., Saddler, G., Scortichini, M., Stead, D.E., Takikawa, Y. 2010. Comprehensive list of names of plant pathogenic bacteria, 1980-2007. Journal of Plant Pathology. 92:551-592.
- Bull, C.T., Mauzey, S.J., Koike, S.T. 2010. First report of bacterial blight of Brussels sprouts (Brassica oleracea L. var. gemmifera) caused by Pseudomonas cannabina pv. alisalensis in California. Plant Disease. 94:1375.
- Grunwald, N.J., Martin, F.N., Larsen, M.M., Sullivan, C.M., Press, C.M., Coffey, M.D., Hansen Everett, M., Parke, J.L. 2011. Phytophthora-ID.org: a sequence-based Phytophthora identification tool. Plant Disease. 95(3):337- 342.
- L�vesque, C., Brouwer, H., Cano, L., Hamilton, J.P., Holt, C., Huitema, E., Raffaele, S., Robideau, G.P., Win, J., Zerillo, M.M., Beakes, G.W., Boore, J.L., Busam, D., Dumas, B., Ferriera, S., Fuerstenberg, S.I., Gachon, C.M. , Gaulin, E., Govers, F., Grenville-Brigg, L., Horner, N., Hostetler, J., Jiang, R., Johnson, J., Krajaejun, T., Lin, H., Meijer, H., Moore, B., Morris, P., Phuntmart, V., Puiu, D., Shetty, J., Stajich, J., Tripathy, S., Wawra, S., West, P., Whitty, B.R., Coutinho, P., Martin, F.N., Thines, M., Thomas, P.D., Tyler, B.M., Vries, R., Kamoun, S., Yandell, M., Tisserat, N., Buell, C. 2010. Genome sequence of the necrotrophic plant pathogen Pythium ultimum reveals original pathogenicity mechanisms and effector repertoire.Genome Biology. 11:R73.
- Mammella, M., Cacciola, S., Martin, F.N., Schena, L. 2011. Genetic characterization of Phytophthora nicotianae by the analysis of polymorphic regions of the mitochondrial DNA. Fungal Biology. 115:432-442.
- Paz, Z., Garcia-Pedrajas, M.D., Andrews, D.L., Klosterman, S.J., Montanez, B., Gold, S.E. 2011. One step construction of agrobacterium recombination- ready plasmids (OSCAR), an efficient and robust tool for ATMT based gene deletion construction in fungi. Fungal Genetics and Biology. 48:677-684.
- Bull, C.T., Clarke, C.R., Cai, R., Vinatzer, B.A., Jardini, T.M., Koike, S. T. 2011. Multilocus sequence typing of Pseudomonas syringae sensu lato confirms previously described genomospecies and permits rapid identification. Journal of Phytopathology. DOI: 10.1094/PHYTO-11-10-0318.
- Mauzey, S., Koike, S.T., Bull, C.T. 2011. First report of bacterial blight of cabbage (Brassica oleracea var. capitata L.) caused by Pseudomonas cannabina pv. alisalensis in California. Plant Disease. 95(1):71.
- Bull, C.T., Rubio, I. 2011. First report of bacterial blight of crucifers caused by Pseudomonas cannabina pv. alisalensis in Australia. Plant Disease. 95(8)1027.
- Klosterman, S.J., Anchieta, A.G., Garcia-Pedrajas, M.D., Maruthachalam, K., Hayes, R.J., Subbarao, K.V. 2011. SSH reveals a linkage between a senescence-associated protease and Verticillium wilt symptom development in lettuce (Lactuca sativa). Physiological and Molecular Plant Pathology. 76:48-58.
- Martin, F.N. 2010. Mitochondrial haplotype analysis as a tool for differentiating populations of Verticillium dahliae. Phytopathology. 100(11):1231-1239.
- Maruthachalam, K., Klosterman, S.J., Kang, S., Hayes, R.J., Subbarao, K.V. 2011. Identification of pathogenicity-related genes in the vascular wilt fungus verticillium dahliae by agrobacterium tumefaciens-mediated t-DNA insertional mutagenesis. Molecular Biotechnology. PMID:21424547.
- Maruthachalam, K., Atallah, Z.K., Vallad, G.E., Klosterman, S.J., Hayes, R. J., Davis, R., Subbarao, K.V. 2010. Molecular variation among isolates of verticillium dahliae and PCR-based differentiation of races. Phytopathology. 100(11):1222-30.
|
Progress 10/01/09 to 09/30/10
Outputs
Progress Report Objectives (from AD-416) Objective 1: Optimize delivery and evaluate performance of chemical, cultural, biological, and genetic alternatives to methyl bromide for crops/pathogen combinations currently benefiting from the use of methyl bromide. a. Evaluate alternative fumigants. b. Identify genes associated with pathogenicity of Verticillium dahliae based on a Verticillium comparative genomics study to support the development of alternative control procedures. c. Identify emerging diseases and their etiology and evaluate resistant germplasm for diseases of strawberry, lettuce, and vegetables. d. Evaluate the influence of crop rotation on pathogen populations and beneficial microbial community dynamics in the soil and severity of Verticillium wilt and other diseases. Develop tools to evaluate and understand the role of myxobacteria in agricultural and natural ecosystems in an effort to elucidate their potential for pathogen control. Objective 2: Develop molecular diagnostic tools for the identification of emerging diseases of vegetables and strawberries, and use these tools in the development of management strategies as alternatives to methyl bromide. Approach (from AD-416) Develop integrated management approaches including crop rotation, biological control, selection of disease resistant varities, organic production, other biological practices, and combinations of biological practices with reduced concentrations of chemical fumigants to control diseases. Identifying useful commercial biological agents as well as new agents from the strawberry rhizosphere that will help to improve plant growth and disease management. Molecular tools will be developed for evaluation of the fficacy of pathogen management and modeling population dynamics of beneficial microbes. Replaces 5305-22000-009-00D(11/07). Comparative analyses of the predicted protein sets from the genomes of fungal vascular wilt pathogens V. dahliae, Fusarium oxysporum, and V. albo-atrum, revealed a conserved set of proteins that may be important for niche adaptation and pathogenesis in these fungi. Further analyses of these genes were initiated in a collaborative effort with other researchers. Research was initiated to quantify the fungus V. dahliae in commercial spinach seeds and to examine the localization of the fungus in spinach seeds. Analyses of a method to distinguish between two races of V. dahliae, was conducted, and nearly completed, in collaboration with the UC Davis. We continued with the analyses of genes that may have a role in plant resistance to fungi. Analyses of a publicly available plant gene database revealed a lettuce gene that shares homology with a gene that confers race 1 resistance to Verticillium wilt disease of tomato and potato. Additional insertional mutants of the fungus V. dahliae were generated and analyses of the mutants are ongoing using pathogenicity tests. We described six novel diseases from the Western US and made tentative identifications of the pathogens from four of the diseases. We completed Koch�s postulates on three pathogens causing bacterial leaf spot on parsley, identified two of the pathogens and demonstrated the usefulness of MLST/MLSA for allocating strains to pathovars of Pseudomonas syringae. We completed the evaluation of rifampicin resistant variants of bacterial plant pathogens for virulence and growth characters and began to use them in population experiments. We began data analysis of FAME results from an experiment measuring the impact of 1 � 5 years vegetable rotation on organic strawberry production in collaboration with UCSC. We transferred P. syringae pv. alisalensis to P. cannabina, resulting in a proposal for a novel species and an additional pathovar, P. cannabina pv. cannabina. We characterized phage pBS1 to which P. cannabina pv. alisalensis is specifically sensitive. Dose response studies are in progress to evaluate the efficacy of alternative fumigants and what concentrations are needed to manage specific soilborne pathogens (Pythium ultimum and Rhizoctonia solani) A comprehensive multigene phylogenetic analysis of 4 Phytophthora species and species complexes has been completed in collaboration with 3 other labs and the results are currently being written up. A comprehensive mitochondrial multigene phylogeny of the genus Phytophthora has been concluded, the data analysis is in the final stages and the manuscript will be written up shortly. A new diagnostic marker system for Phytophthora has been developed and is currently being validated with field samples from the USDA-FS P. ramorum national survey (this work will support the mission of the USDA- FS). Significant Activities that Support Special Target Populations � Mentored five students (4 women and one minority) from Hartnell College and CSUMB (both minority serving institutions) through a grant funded internship program. � Organized and participated in 5 field trips or outreach activities by the USDA for Hartnell College or CSUMB classes or students. � Initiated a project in which our research station is �adopting� with four 5th grade classes at a local minority serving elementary school to provide science programming and field trips. � Organized USDA participation and participated in a Teacher Externship Day Thirty teachers from the Salinas valley attended. � Participated in WISE Women In Science Exploration of Hartnell Community College as a mentor and sponsor. � Serves on the Committee for Equality and Diversity for American Pathological Society. � Mentored one minority intern student � Worked with administrators and teachers at Spreckels Union School District on their science curriculum. Gave classroom presentations in two 5th grade and two 7th grade classes. � Contributed to two classroom presentations (lead responsibility on one) in four 5th grade classes at a local minority serving elementary school the has been �adopted� by our research station to provide science programming and field trips. Accomplishments 01 Gene sets in two fungal pathogens that enable infection of lettuce. ARS researchers in Salinas, California along with a team of international researchers analyzed the genomes of two soilborne, fungal plant pathogen Verticillium dahliae and V. albo-atrum. These two fungi cause vascular wilt diseases on over 200 plant species worldwide, and costly soil fumigants are sometimes deployed to kill these fungal pathogens in the soil. The research discovered gene sets that enable the two fungi to infect their host crops and permit their ecological adaptation. Identification of the genetic basis for pathogenicity or host range expansion may lead to alternative strategies for their control in high value crops such as lettuce. 02 International movement of the Verticilium wilt plant pathogen on spinach seed. ARS researchers in Salinas, California collaborated with a team of university researchers to assess the possibility that Verticillium dahli the soilborne fungus that incites Verticillium wilt disease of lettuce and a wide range of other crop species, can be transmitted over long distances on spinach seed. Verticillium wilt is a relatively new and devastating disease of lettuce, and the pathogen can persist in the soil for many years. The results indicate that there has likely been intra- a intercontinental dissemination of the fungus on spinach seeds from domestic and foreign seed production areas to the major vegetable production area of central, coastal California. Movement of V. dahliae seed from a seed production area into a vegetable production area has troubling implications for Verticillium wilt disease control and mitigation, especially with potentially decreased use of expensive soil fumigants to control this pathogen in U.S. agriculture. 03 Development of the 2010 List of names of Bacterial Plant Pathogens. The nomenclature of bacterial plant pathogens changes as our understanding o relationships among bacteria is advanced. Confusion arises when there a two or more proposals for classification, each with a different nomenclature, for the same organism. A clarified nomenclature is essential to the identification of plant pathogens and other bacteria. A ARS researcher at Salinas, CA convened the International Society of Plan Pathology, Committee on the Taxonomy of Plant Pathogenic Bacteria to develop an official list of names to be used by researchers, regulators, quarantine officials world-wide. 04 Genetic analysis of an important soilborne pathogen of lettuce and other important crops. Verticillium wilt, incited by the soilborne fungus Verticillium dahliae, is a serious disease of lettuce and many other important crop species. Current techniques for identifying sub-populatio such as different pathogenic races, of the pathogen are slow, and as a result may adversely affect management and cropping decisions by farmers ARS researchers at Salinas, CA sequenced parts of the mitochondrial DNA from diverse isolates of the pathogen. They also developed an analytical technique for (haplotype) analysis of mitochondrial DNA that complements current techniques for identification of sub-population groups using vegetative compatibility groupings and nuclear DNA analysis. The mitochondrial technique is currently under evaluation using a much large number of field isolates of the pathogen that have been fully characterized by nuclear DNA analysis to evaluate the utility of mitochondrial haplotype analysis for population studies of this importan pathogen. 05 Web site to support Phytophthora research. The fungal genus Phytophthor is responsible for many diseases of crop plants worldwide and consists o approximately 106 species, which are difficult to identify. ARS researchers at Salinas, CA and other ARS and university researchers with support of NRI Plant Biosecurity grants developed a web-based database f Phytophthora research to enhance the understanding of the genus, simplif species identification and stimulate further research on the genus. The database includes complete morphological descriptions, information on ho range and geographical distribution, a comprehensive molecular phylogeny using seven nuclear genes (four mitochondrial genes will soon be added), and a section on molecular identification and detection. This database will serve as a resource for researchers working on the genus as well as repository for future work. Expansion of the database web portal to include the related pathogens Pythium, downy mildews and Albugo was initiated.
Impacts
(N/A)
Publications
- Almeida, N., Yan, S., Cai, R., Morris, C.E., Schaad, N.W., Schuenzel, E., Lacy, G.H., Sun, X., Jones, J.B., Castillo, J.A., Bull, C.T., Leman, S., Gutman, D.S., Setubal, J.C., Vinatzer, B.A. PAMDB, 2010. A Multilocus Sequence Typing & Analysis Database and Website for Plant-Associated Microbes. Phytopathology. 100:208-215.
- Bull, C.T., Lesaux, M., Manceau, C., Lydon, J., Kong, H.N., Vinatzer, B.A. 2010. Pseudomonas cannabina pv.cannabina pv. nov., and Pseudomonas cannabina pv. alisalensis(Cintas Koike and Bull 2000)comb. nov., are members of the emended species Pseudomonas cannabina(ex �utic & Dowson 1959)Gardan et al., 1999. Systematic and Applied Microbiology. doi:10. 1016/j.syapm.2010.02.001.
|
Progress 10/01/08 to 09/30/09
Outputs
Progress Report Objectives (from AD-416) Objective 1: Optimize delivery and evaluate performance of chemical, cultural, biological, and genetic alternatives to methyl bromide for crops/pathogen combinations currently benefiting from the use of methyl bromide. a. Evaluate alternative fumigants. b. Identify genes associated with pathogenicity of Verticillium dahliae based on a Verticillium comparative genomics study to support the development of alternative control procedures. c. Identify emerging diseases and their etiology and evaluate resistant germplasm for diseases of strawberry, lettuce, and vegetables. d. Evaluate the influence of crop rotation on pathogen populations and beneficial microbial community dynamics in the soil and severity of Verticillium wilt and other diseases. Develop tools to evaluate and understand the role of myxobacteria in agricultural and natural ecosystems in an effort to elucidate their potential for pathogen control. Objective 2: Develop molecular diagnostic tools for the identification of emerging diseases of vegetables and strawberries, and use these tools in the development of management strategies as alternatives to methyl bromide. Approach (from AD-416) Develop integrated management approaches including crop rotation, biological control, selection of disease resistant varities, organic production, other biological practices, and combinations of biological practices with reduced concentrations of chemical fumigants to control diseases. Identifying useful commercial biological agents as well as new agents from the strawberry rhizosphere that will help to improve plant growth and disease management. Molecular tools will be developed for evaluation of the fficacy of pathogen management and modeling population dynamics of beneficial microbes. Replaces 5305-22000-009-00D(11/07). Significant Activities that Support Special Target Populations Continued analyses of the genomes of V. dahliae and V. albo-atrum and comparison to other genomes have revealed sets of genes that are unique to V. dahliae. For additional in-depth analyses, a research proposal was written with 7 other researchers from the US and abroad and submitted to the competitive grants program of the US Agriculture and Food Research Initiative for functional genomics analyses of V. dahliae. Preliminary screens of insertional mutants of V. dahliae have identified two candidate mutant strains of V. dahliae that are potentially nonpathogenic on lettuce. We examined the expression of two defense-related genes of lettuce in response to fungal infection in an additional lettuce cultivar. The experiments revealed that both putative antifungal genes are upregulated in the susceptible interaction with V. dahliae, in leaf tissues. Results of independent differential screening also reveal that these same genes are expressed at higher levels in lettuce leaves infected with V. dahliae versus the non-inoculated control plants. The development of a PCR method in collaboration with the UC Davis to distinguish between two races of V. dahliae is ongoing, using published primer sequences. Our results indicate that this marker system may be useful for rapidly distinguishing between races of V. dahliae pathogenic on lettuce and tomato grown in California. The development of an assay for use in growth chambers that enables a rapid assessment of pathogenicity of V. dahliae on lettuce was completed. This technique employs the use of lettuce seedlings contained in tubes. This technique offers advantages over conventional assays in greenhouses for Verticillium wilt since it is contained, rapid, the environment is controlled, and reduces labor costs associated with lengthy greenhouse assays. We described six novel diseases from the Western US and made tentative identifications of the pathogens from four of the diseases. We also determined that epidemics of bacterial blight on parsley in California and Arizona since 2003 have been caused by two and possibly three different pathogens and are currently evaluating the breadth of the host range of these pathogens. We continue to monitor these emerging diseases. Rifampicin resistant variants of nine bacterial plant pathogens and environmental organisms (needed for myxobacterial enrichment studies) were generated. We are currently comparing these variants to the wild type strains for growth characteristics and virulence on plants. We concluded field trials that identified commercially available strawberry cultivars that are resistant to the pathogen causing Strawberry Blossom Blight. We have shown that the pathogen is similar but substantially different from a know plant pathogen, Pseudomonas marginalis and are concluding the identification. We have completed FAME analysis to measure changes in microbial communities in an experiment measuring the impact of 1 � 5 years vegetable rotation on organic strawberry production in collaboration with a scientist. Data analysis is in progress. We collaborated with ARS Scientists at two other locations in California and university researchers on Significant Activities that Support Special Target Populations � An ARS scientist mentored three students (2 minority and one woman) from Hartnell College and CSUMB (both minority serving institutions) through a grant funded internship program. � An ARS scientist mentored a former minority student intern in preparation graduate school application, a USDA graduate student proposal, and personally provided support and travel for this student to the American Phytopathological Society Meeting in Portland, OR. This student was accepted to the Plant Pathology Department at the University of Wisconsin and will begin a Ph.D. program this fall. � An ARS scientist organized and participated in 5 field trips or outreach activities by the USDA for Hartnell College or CSUMB classes or students. � An ARS scientist initiated a project in which our research station is �adopting� with four 5th grade classes at a local minority serving elementary school to provide science programming and field trips. � An ARS scientist organized USDA participation and participated in a Teacher Externship Day Thirty teachers from the Salinas valley attended. � An ARS scientist participated in WISE Women In Science Exploration of Hartnell Community College as a mentor and sponsor. � An ARS scientist provided additional outreach activities to classes or administrators from 2 additional elementary schools in Salinas. � An ARS scientist provided a field trip to the station for Girl Scout Troup 3197 and their parents. � An ARS scientist serves on the Joint Committee for Women in Plant Pathology and Cultural Diversity for APS � An ARS scientist organized and delivered one summer workshop for Salinas High School teachers at the USDA-ARS station in Salinas, CA. � An ARS scientist has been working with administrators and teachers at Spreckels Elementary School on their science curriculum. Technology Transfer Number of New Germplasm Releases: 1 Number of Web Sites managed: 4
Impacts
(N/A)
Publications
- Njoroge, S., Kabir, Z., Martin, F.N., Koike, S.T., Subbaro, K.V. 2009. COMPARISON OF CROP ROTATION FOR VERTICILLIUM WILT MANAGEMENT IN CONVENTIONAL AND ORGANIC STRAWBERRY PRODUCTION. Plant Disease 93:519-527.
- Martin, F.N., Coffey, M., Berger, P., Hamelin, R.C., Tooley, P.W., Garbelotto, M., Hughes, K., Kubisiak, T. 2009. Evaluation of molecular markers for Phytophthora ramorum detection and identification; testing for specificity using a standardized library of isolates. Phytopathology 98:390-403.
- Klosterman, S.J., Hayes, R.J. 2009. A soilless Verticillium wilt assay using an early flowering lettuce. Plant Disease. 93:691-698,2009.
- Klosterman, S.J., Atallah, Z.K., Vallad, G.E., Subbarao, K.N. 2009. Diversity, Pathogenicity And Control of Verticillium Species.. Annual Review of Phytopathology 2009, 47:39-62.
- Bull, C.T., De Boer, S.H., Denny, T.P., Firrao, G., Fischer-Le Saux, M., Saddler, G.S., Scortichini, M., Stead, D.E., Takikawa, Y. 2008. Demystifying the Nomenclature of Bacterial Plant Pathogens. Journal of Plant Pathology. 90:403-417.
- Bull, C.T., Du Toit, L. 2009. First Report of Bacterial Blight on Conventionally and Organically Grown Arugula in Nevada Caused by Pseudomonas syringae pv. alisalensis. Plant Disease 93:109.
- Vinatzer, B., Bull, C.T. 2009. The impact of genomic approaches on our understanding of diversity and taxonomy of plant pathogenic bacteria. Pages 37-61. In Plant Pathogenic Bacteria: Genomics & Molecular Biology, R. W. Jackson Ed. Caister Academic Press, Norfolk, UK.
|
Progress 10/01/07 to 09/30/08
Outputs
Progress Report Objectives (from AD-416) Objective 1: Optimize delivery and evaluate performance of chemical, cultural, biological, and genetic alternatives to methyl bromide for crops/pathogen combinations currently benefiting from the use of methyl bromide. a. Evaluate alternative fumigants. b. Identify genes associated with pathogenicity of Verticillium dahliae based on a Verticillium comparative genomics study to support the development of alternative control procedures. c. Identify emerging diseases and their etiology and evaluate resistant germplasm for diseases of strawberry, lettuce, and vegetables. d. Evaluate the influence of crop rotation on pathogen populations and beneficial microbial community dynamics in the soil and severity of Verticillium wilt and other diseases. Develop tools to evaluate and understand the role of myxobacteria in agricultural and natural ecosystems in an effort to elucidate their potential for pathogen control. Objective 2: Develop molecular diagnostic tools for the identification of emerging diseases of vegetables and strawberries, and use these tools in the development of management strategies as alternatives to methyl bromide. Approach (from AD-416) Develop integrated management approaches including crop rotation, biological control, selection of disease resistant varities, organic production, other biological practices, and combinations of biological practices with reduced concentrations of chemical fumigants to control diseases. Identifying useful commercial biological agents as well as new agents from the strawberry rhizosphere that will help to improve plant growth and disease management. Molecular tools will be developed for evaluation of the fficacy of pathogen management and modeling population dynamics of beneficial microbes. Replaces 5305-22000-009-00D(11/07). Significant Activities that Support Special Target Populations We prepared three V. dahliae EST libraries, one grown in complete medium, one with a limited nitrogen source, and one in the presence of lettuce root extract. These libraries were shipped to the Broad Institute of MIT and Harvard, and used in the annotation of the V. dahliae genome. In addition, the EST libraries are currently being analyzed to assess overall V. dahliae gene expression under the three different growth conditions. Because the initial screens of lettuce cDNAs for differential expression were unsatisfactory, an approach was employed to examine the expression of candidate lettuce antifungal genes. The expression of antifungal pathogenesis-related genes, such as a class IV endochitinase gene and a thaumatin-like gene were examined in the lettuce-V. dahliae interaction. The experiments revealed tissue-specific expression patterns of both genes in the susceptible interaction with V. dahliae, with higher levels of the thaumatin-like gene expressed in leaf tissues and reduced expression of the class IV endochitinase in root tissues infected with V. dahliae. A PCR method developed in collaboration with the UC Davis to distinguish between two variants, or races, of V. dahliae infecting lettuce was further tested for validity and applicability. This method was invalidated by testing isolates using the PCR method, coupled with virulence assays in greenhouses. Other methods are being pursued to distinguish races of V. dahliae. An assay was developed for use in growth chambers that enables a more rapid assessment of pathogenicity of V. dahliae on lettuce. This technique employs the use of lettuce seedlings contained in tubes. This technique offers advantages over conventional assays in greenhouses for Verticillium wilt in that it is contained, rapid, and the environment is controlled. The assay also reduces labor costs associated with lengthy greenhouse assays. Insertional mutants of V. dahliae were analyzed and preliminary analyses revealed a coalescence of filaments and reduced production of microsclerotia in one of these mutants. Currently, analyses of the DNA from this strain by thermal asymmetric interlaced (TAIL) PCR are being performed to identify the gene associated with this phenotype. We tentatively identified a novel pathogen of strawberry flowers as Pseudomonas marginalis. We conducted laboratory and field evaluations to determine if there is resistance to the pathogen in currently used commercial varieties. Indications from the first field experiment indicate that there is some level of resistance available. This work is supporting NP308 Problem Statement 1C: Identification and Mitigation of Emerging Problems. Significant Activities that Support Special Target Populations � One of the SYs organized a field trip of University of California, Santa Cruz students, including minorities and women, to meet and discuss research and careers with researchers at the USDA-ARS station in Salinas, CA. � One of the SYs has trained student interns from Hartnell College and CSUMB as part of a USDA funded program to increase the exposure of minority (specifically Hispanic) students to science. � Dr. Bull works with Hartnell College on the development of a Science Institute designed to develop minority student interest in Science Careers and to increase internship possibilities for underrepresented populations. This resulted in a Science Institute Visioning Workshop held April 22, 2008 where she served as a Workshop Organizer, MC and breakout session leader. � March 5, 2007; Dr. Bull provided a pre-print of the Ruth Allen Chapter to Charmaine Scardina for Women�s history month. She provided it to all locations in the PWA. � August 2008: Dr. Bull organized and Moderated a session on Diversity for the American Phytopathological Society Centennial meeting. To prepare for this session Dr. Bull completed and began to analyze a survey and special session on the status of women and minority plant pathologists. � July 2008: Dr. Bull gave a talk about research opportunities and careers a high school bridging program at Hartnell community College designed to increase math and science skills of incoming Freshmen. � Dr. Bull serves on the Joint Committee for Women in Plant Pathology and Cultural Diversity for APS
Impacts
(N/A)
Publications
|
|