Whole-genome sequencing of Mesorhizobium huakuii 7653R provides molecular insights into host specificity and symbiosis island dynamics.

BACKGROUND: Evidence based on genomic sequences is urgently needed to confirm the phylogenetic relationship between Mesorhizobium strain MAFF303099 and M. huakuii. To define underlying causes for the rather striking difference in host specificity between M. huakuii strain 7653R and MAFF303099, several probable determinants also require comparison at the genomic level. An improved understanding of mobile genetic elements that can be integrated into the main chromosomes of Mesorhizobium to form genomic islands would enrich our knowledge of how genome dynamics may contribute to Mesorhizobium evolution in general. RESULTS: In this study, we sequenced the complete genome of 7653R and compared it with five other Mesorhizobium genomes. Genomes of 7653R and MAFF303099 were found to share a large set of orthologs and, most importantly, a conserved chromosomal backbone and even larger perfectly conserved synteny blocks. We also identified candidate molecular differences responsible for the different host specificities of these two strains. Finally, we reconstructed an ancestral Mesorhizobium genomic island that has evolved into diverse forms in different Mesorhizobium species. CONCLUSIONS: Our ortholog and synteny analyses firmly establish MAFF303099 as a strain of M. huakuii. Differences in nodulation factors and secretion systems T3SS, T4SS, and T6SS may be responsible for the unique host specificities of 7653R and MAFF303099 strains. The plasmids of 7653R may have arisen by excision of the original genomic island from the 7653R chromosome.

[Cloning and identification of the plasmid replication gene repC in Mesorhizobium huakuii].

OBJECTIVE: The repC gene is the principal initiation protein gene for plasmid replication. We identified the repC-like sequences from Mesorhizobium huakuii strain HN3015 and its derivatives of plasmid curing. METHODS: Primers of RC1 and RC3 were used to amplify the repC-like sequences by a polymerase chain reaction, the PCR products were obtained and cloned into plasmid vector pMD-18T and then sequenced. Location of the repC sequences on different plasmids in the strains tested was carried out by Southern blotting. The nucleotides homology analysis of the repC gene was carried out using the BLAST. Amino acid sequences were deduced using the ExPASy. Multiple sequences alignments were performed using the ClustalW. Analysis of protein secondary structure was carried out using the PredictProtein. RESULTS: The repC-like sequences were obtained from the strains tested. The sizes of the PCR products were about 750 bp. The results of Southern blotting showed that the repC-like sequences were only associated with a plasmid in the stains tested. CONCLUSIONS: The repC sequences of the strains tested showed 100% sequence similarity, but were obviously different from that of other rhizobia strains.

RNA-Seq and microarrays analyses reveal global differential transcriptomes of Mesorhizobium huakuii 7653R between bacteroids and free-living cells.

Mesorhizobium huakuii 7653R occurs either in nitrogen-fixing symbiosis with its host plant, Astragalus sinicus, or free-living in the soil. The M. huakuii 7653R genome has recently been sequenced. To better understand the complex biochemical and developmental changes that occur in 7653R during bacteroid development, RNA-Seq and Microarrays were used to investigate the differential transcriptomes of 7653R bacteroids and free-living cells. The two approaches identified several thousand differentially expressed genes. The most prominent up-regulation occurred in the symbiosis plasmids, meanwhile gene expression is concentrated to a set of genes (clusters) in bacteroids to fulfill corresponding functional requirements. The results suggested that the main energy metabolism is active while fatty acid metabolism is inactive in bacteroid and that most of genes relevant to cell cycle are down-regulated accordingly. For a global analysis, we reconstructed a protein-protein interaction (PPI) network for 7653R and integrated gene expression data into the network using Cytoscape. A highly inter-connected subnetwork, with function enrichment for nitrogen fixation, was found, and a set of hubs and previously uncharacterized genes participating in nitrogen fixation were identified. The results described here provide a broader biological landscape and novel insights that elucidate rhizobial bacteroid differentiation, nitrogen fixation and related novel gene functions.