Stress and Immunity of Nodule Bacteria Sinorhizobium Meliloti: Localization, Polymorphism and Phylogeny of Genetic Determinants

Abstract

Sinorhizobium meliloti are agriculturally valuable species of soil bacteria that form nitrogen-fixing symbiosis with alfalfa plants. Global climate changes lead to an increase of agricultural areas subjected to salinity. Current knowledge about about high-salt stress impact on soil saprophitic root nodulated microsymbionts of legumes is weakly studied and rhizobia gene pool responsible for salt tolerance are fragment and far from clear. An increase of bacteria nonspecific resistance (immune status) to unfavorable stress factors can occur through the induction of defense mechanisms like restrictionmodification systems and CRISPR/cas systems which are aimed to protect bacteria cells from bacteriophages widespread in soil microbiome. The aim of this research was to evaluate the role of the megaplasmid pSymA in the formation of ecological genome of S. meliloti, which is related to stress tolerance and to determine the location of elements of adaptive immune systems protecting root nodule bacteria against external foreign DNA. The analysis was done on 11 genes, products of which involved in response to ion stress and synthesis of osmoprotectors. It was found that 6 out of 11 genes were found in the genomes of all analyzed S. meliloti strains, while it was not a case for other 5 genes. It was found that, unlike chromosome, megaplasmid I of S. meliloti accumulated copies of 4 from 5 genes, except kdpA gene, which is represented by a single copy and localized on megaplasmid I in all so far studied strains. It was predicted that closest phylogenetic relatives of genes whose products are involved in response to ion stress as well in synthesis of osmoprotectors are homologous genes of closely related S. medicae species. The exception was for betI2, for which the closest phylogenetic relative was homologous gene of Klebsiella pneumonia, and another exception is kdpA gene introduced onto megaplasmid-I from actinobacteria. Regarding elements of immune systems it was revealed that nonsymbiotic plasmids of S. meliloti harbored incomplete elements of RMS-I, -II, and - III systems, while the 4 complete RMS-IV systems were detected on a single plasmid. It was found out that corresponding methylases had similarities with similar enzymes detected in nitrogen-fixing strains of Agrobacterium tumefaciens, Mezorhizobium sp., Bradyrhizobium sp. CRISPR sequences were not detected on megaplasmid-I, while they were on chromosome, megaplasmid-II and on cryptic plasmids. So, it was concluded that megaplasmid-I of S. meliloti are enriched in copies of genes related to osmotic stress tolerance, but it role in immune status of rhizobia is requested further elucidation.