1a. Objectives (from AD-416)
Objective 1: Strategically expand the genetic diversity in the ARS Culture Collection and improve associated information for priority microbial genetic resources. Sub-objective 1.A. Acquire from diverse sources samples of food-borne pathogenic bacteria, actinobacteria from equine sources, basidiomycetous yeasts, plant pathogenic fungi, and grain storage molds to fill current gaps in the ARS Culture Collection for these priority microbial strains. Sub-objective 1.B. In consultation with the microbial research community, identify microbial genetic resources associated with discontinued research programs, or held by researchers who are nearing retirement, and attempt to acquire those of strategic importance to current or future agricultural research programs. Objective 2: Conserve priority microbial genetic resources efficiently and effectively, and distribute them and associated information worldwide. Sub-objective 2.A. Conserve more than 90,000 accessions of priority microbial genetic resources and associated information, emphasizing food-borne pathogenic bacteria, actinobacteria from equine sources, basidiomycetous yeasts, plant pathogenic fungi, and grain storage molds, as well as microbes of biomedical and biotechnological importance. Sub-objective 2.B. Back-up at the National Center for Genetic Resources Preservation (NCGRP) approximately 15,000 strains (emphasizing Fusarium) that are currently preserved under liquid nitrogen vapor only at the National Center for Agricultural Utilization Research (NCAUR). Sub-objective 2.C. Continue to improve data management and technology transfer procedures, emphasizing improvements in user interface for the public access catalog system. Sub-Objective 2.D. Distribute on request microbial accessions and information that meet the specific needs of agricultural, biomedical, and biotechnological researchers. Objective 3: Strategically characterize (“genotype”) and evaluate (“phenotype”) priority microbial genetic resources through multigene analyses, and with key morphological, physiological, and biochemical descriptors. Sub-objective 3.A. Develop and apply multigene markers for phylogenetic and genetic diversity analyses of priority microbial genetic resources. Incorporate characterization data into GRIN and/or other databases, and apply the data to providing accurate taxonomic identifications, as well as to predicting the agricultural and biotechnological utility of newly discovered taxa. Sub-objective 3.B. Determine the phenotypic diversity and elucidate the population genetic structure for the Fusarium Head Blight (FHB) species Fusarium graminearum and F. asiacticum. Map their worldwide distribution, as a first step of establishing a molecular surveillance system for the early detection of Fusarium populations introduced to North America.
1b. Approach (from AD-416)
New species and novel strains of known species of plant pathogens and mycotoxigenic fungi, food-borne pathogens, actinobacteria important to animal health and biotechnology, and yeasts will be isolated from nature or acquired from reports in the literature and from cooperators worldwide. New strain accessions will be cataloged in the collection database, preserved by lyophilization and/or freezing in liquid nitrogen vapor where appropriate, and information related to well characterized strains will be made publicly available on the Collection website. Information provided by ARS Program Leadership, national and international microbiology societies and Culture Collection organizations will identify microbial collections in danger of being lost and important collections will be acquired and accessioned where existing resources permit. The entire collection is secured in a limited access room and records for strain inventory and distribution are maintained on the collection database system. High priority microbial strains held only as frozen preparations under liquid nitrogen vapor phase will be duplicated and shipped to the National Center for Genetic Resources Preservation. NCGRP for backup in a liquid nitrogen freezer dedicated for microbial germplasm. Strains will be freely distributed to the scientific community worldwide but requestors must provide required documentation or permits before animal or plant pathogenic strains or those requiring Biosafety Level II confinement are distributed. Through phylogenetic analysis of sequences from multiple gene loci, evaluate the diversity and systematics of actinomycetes, Bacillus, Aspergillus, and yeasts of importance to agriculture, food safety, and biotechnology. A multilocus genotyping assay previously developed for identification of FHB species and trichothecene chemotypes will be applied to a global collection of FHB isolates to determine the current distribution and trichothecene chemotype diversity of Fusarium graminearum and F. asiacticum populations. Population diversity and relatedness will then be assessed using a published panel of variable number tandem repeat markers. Differences in pathogen fitness and aggressiveness in individual populations will be evaluated by determining a range of phenotypic characteristics, such as growth, reproduction, and toxin production.
3. Progress Report
The overall goals of this project are to enhance the Agricultural Research Service (ARS) Culture Collection through acquisition of novel microbial germplasm and to characterize this germplasm genetically through gene sequence analysis. A total of 811 strains have been accessioned into the general collection and 149 deposits made into the Patent Collection. The Patent Culture Collection has distributed 337 strains including 179 to scientists in the United States, and 158 to foreign scientists. Strain distributions from the general collection totaled approximately 7,828, including 836 strains to the Institute of Genomic Biology, University of Illinois, to complete the ARS contribution to a collaborative research project, 1,501 to ARS scientists, 3,648 to other non-ARS scientists in the United States, and 1,843 to foreign scientists. The ARS Culture Collection website now has 12,425 strain records available in the public access catalog, had over 24,252 visitors from over 120 different countries and has been indexed by at least 79 different search engines. The online strain request module for the Culture Collection database system/website has facilitated the processing and tracking of strain requests using the collection staff. These activities provide for continued preservation of agriculturally and biotechnologically significant microbial germplasm and distribution to researchers in ARS as well as throughout the world. Continued collaboration with scientists at the Institute of Genomic Biology, University of Illinois, through a Material Transfer Agreement and cooperative agreement has been leveraged to provide raw sequence data for the house-keeping genes atpD, rpoB, and trpB for approximately 1,400 strains of actinomycetes of the genus Streptomyces, including both type of strains as well as uncharacterized strains in support of project objectives. The data are being assembled and organized into a web-based online identification database which will be made available to the scientific community worldwide. The identification database is being tested continually as the data are processed and entered and several new species have been detected within the strain collection. Two new methanol-utilizing yeast species, Ogataea saltuana and Ogatae parapolymorpha, were discovered within the holdings of the Yeast Culture Collection. Methanol-utilizing yeasts have been genetically modified in the past to produce large quantities of recombinant proteins and this discovery provides molecular biologists with novel microorganisms for biotechnological exploitation. In regard to the Fusarium Head Blight (FHB) research component of the project, a population of nivalenol-producing Fusarium cereal pathogens was discovered to be present in the U.S. for the first time. Nivalenol is of greater public health concern than deoxynivalenol, which is produced by most FHB populations in the U.S. The identification of a nivalenol-producing population in the U.S. indicates the need for additional monitoring, and for the development and use of rapid mycotoxin test kits capable of detecting nivalenol and differentiating it from the more common deoxynivalenol.
1. Molecular characterization of priority microbial genetic resources. The biodiversity of much of the microbial germplasm held in the Agricultural Research Service (ARS) Culture Collection in Peoria, IL, is not known because of limited characterization for many strains, making it difficult to assess its real or potential value by customers involved in agricultural and biotechnology research. The diversity of various groups of microbes within the Collection was estimated, particularly for poorly characterized isolates, based on the sequences of various genes. The identification system being developed for the actinomycete genus Streptomyces, whose species produce many of the medically important antibiotics, has already demonstrated the presence of unrecognized new species. In collaboration with scientists at the Institute of Genomic Biology, University of Illinois, scientists in the ARS Bacterial Foodborne Pathogens and Mycology Research Unit, National Center for Agricultural Utilization Research (NCAUR), Peoria, IL, discovered over 400 bacterial strains with the genes necessary to produce novel natural products, such as antibiotics. These research accomplishments contribute to the characterization of germplasm held in the Collection and provide knowledge regarding potential uses in biotechnology, crop production, and food safety. As a result of the efforts to preserve and characterize the strains within the Collection, during the past year, 8,165 strains from the ARS Culture Collection were sent out in response to requests from industry, academic, and government scientists in 57 countries, which included requests from 464 different scientists in the U.S.
2. Identification of a new cereal pathogen population in the United States. Fusarium head blight [FHB] is a disease of cereal crops, caused by a fungal pathogen, which results in significant economic and health problems throughout the world. In collaboration with scientists at the Agricultural Research Service (ARS) Cereal Disease Laboratory in St. Paul, MN, scientists in the Agricultural Research Service (ARS) Bacterial Foodborne Pathogens and Mycology Research Unit, National Center for Agricultural Utilization Research, Peoria, IL, identified the first significant U.S. population of FHB pathogens that can contaminate grain with the mycotoxin nivalenol (NIV). This is significant because NIV can be more toxic to humans and livestock than the toxin deoxynivalenol, or DON, which is produced by previously recognized FHB populations in the U.S. These results alert grain producers and processors to the possible presence of NIV in grain, and indicate the need to utilize, as appropriate, mycotoxin detection kits that are capable of detecting NIV. In addition, by providing plant breeders with accurate information about the spectrum of FHB pathogens present in U.S. fields, these data will be of use to cereal breeding programs aimed at developing broad-based resistance to fungal pathogens.
Kurtzman, C.P. 2011. Babjeviella Kurtzman & M. Suzuki (2010). In: Kurtzman, C.P., Fell, J.W., Boekhout, T., editors. The Yeasts, a Taxonomic Study. Volume 2, 5th edition. New York, NY: Elsevier. p. 329-331.