Location: Mycotoxin Prevention and Applied Microbiology Research
2024 Annual Report
Objectives
The major goals of this project will be most effectively achieved through strengthening the relationship between the ARS Culture Collection and the user community and fostering a synergistic relationship. To this end, we will implement a “virtuous cycle” of collection management (Graphical Project Summary) to achieve the project goals and enhance our connection to the user community.
Objective 1: Acquire, distribute, back-up, and maintain the safety, genetic integrity, health, and viability of priority microbial genetic resources and associated descriptive information in the ARS Culture Collection.
Sub-objective 1.A: Maintain the safety, genetic integrity, viability and accessibility of priority microbial genetic resources and associated information in the ARS Culture Collection.
Sub-objective 1.B: Enhance the diversity and utility of the ARS Culture Collection by acquiring germplasm representing type material or novel strains important to ARS and other microbial research programs.
Objective 2: Increase the value of priority microbial resources in the ARS Culture Collection for research and development by conducting integrated phylogenetic, physiological, phenotypic, and metabolomic analyses. Connect these multiple data types to accession records in the ARS Culture Collection, and deposit in appropriate public databases.
Sub-objective 2.A: Characterize 2,500 priority strains of bacteria and fungi.
Sub-objective 2.B: Update the metadata on strains in the ARS Culture Collection using the Ag Data Commons website to facilitate research by ARS and other scientists worldwide.
Approach
More than 98,000 accessions consisting of microbial cultures of agriculturally, industrially, and medically important bacteria and fungi will be maintained in a secure centralized limited-access facility. These microbial genetic resources will be effectively preserved and secured, characterized to improve their utility, taxonomic accuracy, and made available to support agricultural and other research projects worldwide. In addition to maintaining and distributing currently held strains, high priority microbial genetic resources will be acquired and safeguarded so that these critical resources are widely available for microbial research and development. Microbial cultures will be characterized with a combination of genetic, genomic, metabolomic, and phenotypic approaches and analyses to elucidate their phylogenetic relationships, secondary metabolite profiles, ensure accurate species identification, and enhance their utility for research and development.
Progress Report
The ARS Culture Collection, also known by the original acronym for the Northern Regional Research Lab (NRRL) in Peoria, Illinois, is one of the largest publicly accessible collections of microorganisms in the world. The ARS Culture Collection consists of a collection of strains that are open for public distribution and a collection of strains that are distributed in accordance with an international treaty on recognition procedures for patents involving microorganisms. The open collection currently maintains more than 98,000 strains of fungi and bacteria together with more than 7,500 strains held in the ARS Patent Culture Collection. The ARS Culture Collection is one of only two International Depositary Authorities recognized by the World Intellectual Property Organization that curate fungi and bacteria in the United States. In this capacity, The ARS Culture Collection facilitates technological innovation by enabling scientists to simultaneously fulfill microbial culture deposition requirements in association with patent applications in the United States and internationally. The major goals of the current project are to conduct and support microbiological research that advances food safety, public health, economic development, and agricultural production. In-house research in pursuit of these goals is focused on improved understanding and utilization of microbiological diversity together with efforts to enhance the value and accessibility of microbial germplasm in the ARS Culture Collection. Strains from the collection have contributed significantly to advances in agriculture, medicine, and biotechnology, and are cited in approximately 70,000 patents and scientific publications. Goals of the first objective include acquiring, safeguarding, and distributing priority microbial genetic resources to advance scientific discovery. In response to ARS research programs and customer requests, 461 new strains were accessioned into the open collection and 88 new strains were accessioned into the patent collection. During the last year the ARS Culture Collection distributed 5,879 microbial strains to 561 unique recipients across the United States and 42 other countries. These strain distributions conservatively represent an in-kind contribution of ~ $1 million in support of microbiological research and biotechnological innovation. The utility and security of the collection was improved by adding or updating 5,200 strain history or inventory records.
Research under the first objective is also focused on maintaining the integrity of the existing collection and acquiring material to improve the taxonomic representation and utility of the collection. To this end the ARS Culture Collection acquired and accessioned 215 isolates of the rice pathogen Magnaporthe oryzae collected by a scientist at Purdue University that represent the global distribution (15 countries in North and South America and Asia) of this important fungal pathogen. These represent the first members of the Magnaporthe genus in the ARS Culture Collection, so ARS researchers in Peoria, Illinois, also optimized a preservation protocol for these accessions to ensure their long-term viability. These strains are currently being used to understand the genetic diversity and host range adaptation of this species to various grass host including rice.
As part of the goal to improve website functionality through user feedback, a survey was developed in fiscal year (FY) 2023-2024 that included questions on how the layout of the website can be improved and topics related to customer service and/or the strains and related metadata in the database. This survey was posted on the NRRL website and was made available to the public between December 01, 2023 – January 31, 2024. The survey received 62 responses. Respondents included scientists from academia, industry, and the federal government. A majority of respondents found the website easy to navigate, but there were some difficulties navigating the online catalogue. Respondents specifically requested greater search functionality.
In response to this feedback, a new site was designed and migrated to the AZURE cloud system to avoid further security concerns. The new site provides the user with the option to download a spreadsheet with either the fungal or bacterial public catalog. Using this spreadsheet format the user can now design their own searches depending on their research needs. The spreadsheet catalog allows the user to sort by Genus, country or origin, substrate, host, cross-reference number in other collections, etc. The new site retained the same URL https://nrrl.ncaur.usda.gov/ minimizing any potential disruptions to the user community. We have received positive feedback from the user community, and orders have continued to be placed at a similar rate as received from the previous version of the website.
Identification and classification of fungi and bacteria increasingly depends on DNA sequences of standardized genetic markers or entire genomes. The greatest value associated with strains in the ARS Culture Collection is this additional metadata generated by project scientists and collaborators including the DNA-based data for identification as well as metabolomic, phenotypic and physiological data. Research under the second objective is focused on generating and utilizing DNA-sequence based analyses of genetic variation and phenotypic data to characterize agronomically important microbes in NRRL and to make these strains and associated metadata publicly available to promote agricultural production, biotechnological development, and food safety. During FY 2024 we initiated a DNA-based taxonomic assessment of 314 Pseudomonas strains in the ARS Culture Collection. The genus Pseudomonas includes both human and plant pathogens as well as species used in biological control and plant growth promotion. This variation in lifestyle between species demonstrates the importance for accurate species-level identification. We were able to accurately identify 212 strains to species and these strains have been added to the public-facing catalog. An additional 35 strains could not be revived or consisted of mixed cultures and were removed from the collection. Finally, 67 strains could not be accurately aligned to a known reference strain based on DNA sequence data (>98% sequence similarity) and may represent putative novel species. Further analyses are being conducted to determine the appropriate taxonomic classification of these strains. This year, we also worked with another ARS researcher in Peoria, Illinois, to complete genome sequencing on 1,155 agronomically important strains from the genus Bacillus. This information was used to update the taxonomy of these strains in the NRRL database. In addition, the GenBank accession number for the genome assemblies of 84 isolates was added to their corresponding accession record and is now available to the public. The remaining Genbank accessions will be added in FY 2025.
Under objective 2, the NRRL is also working to make more of the -omics data generated on strains from the collection available to the public. In a collaboration with the National Agricultural Library (NAL), the ARS Program Page host on the NAL Ag Data Commons was moved to a FigShare formatted website https://agdatacommons.nal.usda.gov/ARS_Culture_Collection. Since going live in January 2024, the site has received 78,000+ views and 1,000+ downloads of datasets. Data curators are regularly checking National Center for Biotechnology Information Genbank for new genome and transcriptome datasets generated from NRRL strains. There are currently 1,278 datasets available on the NAL ARS Program Page.
Accomplishments
1. Development of a search tool to determine microbial origin of metabolites. Microorganisms drive global carbon cycles and establish beneficial relationships with their hosts, influencing health, aging, and behavior. These microbes interact and manipulate their environment through the production and use of secondary metabolites. The vast metabolic potential of these communities is demonstrated by the fact that microbial communities inhabiting humans, plants, animals, and insects contain 100 times more genes than the host genome. However, this metabolic potential remains hidden in modern microbiome experiments, where typically less than 1% of the metabolites can be classified as microbial. ARS researchers in Peoria, Illinois, collaborated with a global network of scientists from over 20 academic institutions led by University of California-San Diego to develop microbeMASST which uses a mass spectrometry (MS) search tool that is able to annotate and discover the microbial origin of known and unknown metabolites in untargeted metabolomics experiments. Researchers anticipate that microbeMASST will be a key resource to improve our understanding of microbial metabolites across a wide range of ecosystems, including plants, soil, insects, animals, and humans. Moreover, microbeMASST holds immense potential for various applications, ranging from aquaculture and agriculture to biotechnology and the study of microbial-mediated human health conditions.
2. Natural genetic diversity allows yeast fungi to utilize a variety of carbon resources. Organisms both big and small show an extensive variation in their ability to exist and grow in a range of specific environmental conditions or niches. Specialist organisms can only occupy a specific niche, whereas generalists are able to occupy a range of niches. Theories proposed to explain this variation include trade-offs between efficiency of resource use by specialist and breadth of resource use by generalist, or they suggest underlying internal genetic or external environmental forces are driving this evolution. ARS researchers in Peoria, Illinois, collaborated with scientists at the University of Wisconsin and Vanderbilt University to assemble genomic, metabolic, and ecological data from nearly all known species of yeast fungi to examine niche range evolution. The study found that closely related species of yeast had large differences in the ability to utilize various carbon resources that is the result of internal differences in genes encoding specific metabolic pathways. There was no evidence of trade-offs between resource use efficiency vs breadth, and a limited role of external environmental factors on ability to utilize carbon resources. These comprehensive data strongly suggest internal genetic factors are driving microbial resource use variation. These results regarding resource utilization by yeast will be of particular interest to fermentation scientists and the fermentation industry. The evolution of genetic and metabolic pathways and genome resources will be of interest to the synthetic biology industry.
3. Developed a new website and a webpage to make ARS Culture Collection strains and associated genome data available to the public. The ARS Culture Collection has a long history of open science including making microbial strains available to the scientific public at no charge. In response to a survey of the user community, the ARS Culture Collection website was updated and now includes greater accessibility to strain data. Through this website the ARS Culture Collection distributes on average 5,500 microbial strains each year to scientists representing the United States and 42 other countries. ARS researhers from the ARS Culture Collection in Peoria, Illinois, collaborated with the National Agricultural Library to launch ARS Culture Collection program page (https://agdatacommons.nal.usda.gov/ARS_Culture_Collection) on its open data repository, Ag Data Commons. This digital version of the ARS Culture Collection contains genetic data generated on microbial strains from the collection. Since January 2024, the site has received 110,000+ views and 2,000+ downloads of datasets. The collection will continue to grow as data curators are regularly checking National Center for Biotechnology Information GenBank for new genome and transcriptome datasets generated from the collection. There are currently 1,278 datasets available to the public from the collection.
Review Publications
Queiroz, C.A., Caniato, F.F., Siqueira, V.K.S., de Moraes Catarino, A., Hanada, R.E., O'Donnell, K., Laraba, I., Rehner, S.A., Sousa, N.R., Silva, G.F. 2023. Population genetic analysis of Fusarium decemcellulare, a guaraná pathogen, reveals high genetic diversity in the Amazonas state, Brazil. Plant Disease. 107(5):1343-1354. https://doi.org/10.1094/PDIS-01-22-0083-RE.
Heiden, N., Broders, K.D., Hutin, M., Ortiz Castro, M., Roman-Reyna, V., Toth, H., Jacobs, J. 2023. Bacterial leaf streak diseases of plants: Symptom convergence in monocot plants by distant pathogenic Xanthomonas species. Phytopathology. 113(11):2048-2055. https://doi.org/10.1094/PHYTO-05-23-0155-IA.
Opulente, D.A., Labella, A.L., Harrison, M.-C., Wolters, J.F., Liu, C., Li, Y., Kominek, J., Kurtzman, C.P., Rokas, A., Hittinger, C.T., et al. 2024. Genomic factors shape carbon and nitrogen metabolic niche breadth across Saccharomycotina yeasts. Science. https://doi.org/10.1126/science.adj4503.
Zuffa, S., Schmid, R., Bauermeister, A., Gomes, P.P., Caraballo-Rodriguez, A., El Abiead, Y., Aron, A.T., Gentry, E.C., Zemlin, J., Dorrenstein, P., Broders, K.D., et al. 2024. microbeMASST: A taxonomically informed mass spectrometry search tool for microbial metabolomics data. Nature Microbiology. https://doi.org/10.1038/s41564-023-01575-9.
Jung, T., Milenkovic, I., Balci, Y., Janousek, J., Kudlacek, T., Nagy, Z.A., Baharuddin, B., Bakonyi, J., Broders, K.D., Cacciola, S.O., et al. 2024. Worldwide forest surveys reveal forty-three new species in Phytophthora major Clade 2 with fundamental implications for the evolution and biogeography of the genus and global plant biosecurity. Studies in Mycology. https://doi.org/10.3114/sim.2024.107.04.