1a. Objectives (from AD-416)
Manage the genetic resources of the National Rhizobium Germplasm Resource Collection. This is a service component of this project that is part of "The Germplasm Resources Information Network (GRIN)" within the National Microbial Germplasm Program (NMGP). Use genomic and phylogenetic analyses to characterize microbes that interact symbiotically with higher plants, such as strains of Bradyrhizobium, that form symbioses with soybean, and the genetic interrelations of the rhizobia that infect Medicago species. Some parts of the technologies that have been developed will be applied to the genetic and phylogenetic analysis of phytopathogenic fungi with another laboratory at Beltsville.
1b. Approach (from AD-416)
Rhizobial cultures will be managed by their preservation, quality control, and disbursement to ARS customers upon request. In some cases inoculum will be manufactured by special request for research purposes. Technical information about rhizobia, their isolation, their culturing, the symbiosis, data analysis and research advice will be given. New rhizobial cultures will be isolated from soil samples collected within the USA and from abroad. Characterization of microbes that interact symbiotically with higher plants and crops will use approaches based on genomic and phylogenetic analyses, predominantly achieved through sequence and analyses of ribosomal and housekeeping gene loci. This will include the development of several new technologies, and analysis of strains of Bradyrhizobium that form symbioses with soybean, and two projects examining the genetic interrelationships of the rhizobia that infect Medicago species.
3. Progress Report
One component of this project is responsibility for curation of the USDA ARS National Rhizobium Germplasm Resource Collection and to provide rhizobial resources and technical guidance to stakeholders and users who depend upon this program for their business and research. Rhizobia is the common name for numerous bacterial species that form associations (known as symbiosis) with plants of the legume family (peas, beans, soybeans and alfalfa, for example) to “fix” gaseous nitrogen from the atmosphere that is then used for plant growth. Progress was made in providing rhizobial cultures and technical assistance to support African American institutions of higher education, 1890 Institutions and other State and Private Universities. Technological support and cultures were provided to small companies specializing in inoculant production and to non-profit organizations. Assistance was given to stakeholders who are developing bio-energy production resources through the cultivation of specific leguminous trees. Our staff spent many hours in telephone and email discussions with these companies, dispensing advice on recommended strains for agronomically important legumes, protocols on media preparation on which to grow rhizobia cultures and the proper growth of cultures, and methods for scaling-up rhizobial inoculant production. Progress was made on the research component of the project related to the use of genomic and phylogenetic analyses to characterize microbes that interact symbiotically with higher plants, such as strains of Bradyrhizobium, that form symbioses with soybean, and the genetic interrelations of the rhizobia that infect Medicago species. One of the technologies under development is the determination of the variation in the bacteria by measuring differences in the DNA of specific genes and is known as MultiLocus Sequence Typing (MLST). When this methodology was applied to bacteria forming a symbiosis with alfalfa we were able to identify differences in the DNA sequence of a number of genes. When these differences in DNA sequence of a number of genes were combined they allowed us to define the genotype of each individual that was analyzed. By then combining the different genotypes, it was possible to describe the genetic structure of the bacterial population that infects the roots of plants and then forms a beneficial symbiosis. The method proved to be far superior in providing this form of information about the genetic structure of the bacteria than the commonly used techniques that are based on DNA fingerprinting. When the method was used with collections of isolates obtained from Western Asia and North Africa it was possible to identify the genotypes of bacteria that normally form a symbiosis with alfalfa in its natural environment. The progress included the design of a database with the information obtained with the MLST technique. The database can then be used as a reference for subsequent projects that determine how biotic and abiotic factors in the environment affect the population of these bacteria.
1. Movement of bacterial genes necessary for the association with common bean from American to European bacterial species. Common bean is a very important crop world-wide and can be grown in many countries without adding specific rhizobial cultures to the soil to encourage the establishment of a nitrogen fixing symbiosis between the plant and rhizobia. This legume crop originally is from America, but there are four bacterial species of American origin that associate with common bean found in the soils of France, Spain and Portugal. These rhizobia were unintentionally transported with the seeds, but their invasive effects on local bacterial species are unknown. Working together with collaborators in Spain, a single bacterial species of European origin was found to have the DNA necessary for the association with common bean. This DNA was determined to be from two of the American bacterial species. This impact of this accomplishment is enhanced insight into how the practice of agriculture affects evolution of agriculturally important bacterial species within very short periods of time. Also, this discovery will potentially provide bacteria with different characters useful for adapting common bean cultivation to other and different climatic zones.
Zribi, K., Mhadhbi, H., Badri, Y., Aouani, M.E., Van Berkum, P.B. 2011. Sinorhizobium meliloti strains TII7 and A5 by Multilocus Sequence Typing (MLST) have chromsomes identical with Rm1021 and form an effective and ineffective symbiosis with Medicago truncatula line Jemalong A17, respectively. Canadian Journal of Microbiology. 59:996-1002.