Multilocus sequence typing as an approach for population analysis of Medicago-nodulating rhizobia

Abstract

Multilocus sequence typing (MLST), a sequence-based method to characterize bacterial genomes, was used to examine the genetic structure in a large collection of Medicago-nodulating rhizobial strains. This is the first study where MLST has been applied in conjunction with eBURST analysis to determine the population genetic structure of nonpathogenic bacteria recovered from the soil environment. Sequence variation was determined in 10 chromosomal loci of 231 strains that predominantly originated from southwest Asia. Genetic diversity for each locus ranged from 0.351 to 0.819, and the strains examined were allocated to 91 different allelic profiles or sequence types (STs). The genus Medicago is nodulated by at least two groups of rhizobia with divergent chromosomes that have been classified as Sinorhizobium meliloti and Sinorhizobium medicae. Evidence was obtained that the degree of genetic exchange among the chromosomes across these groups is limited. The symbiosis with Medicago polymorpha of nine strains placed in one of these groups, previously identified as S. medicae, ranged from ineffective to fully effective, indicating that there was no strong relationship between symbiotic phenotype and chromosomal genotype.

FIG. 1.

Linkage distance among 231 Medicago-nodulating rhizobia derived from allelic variation among 10 chromosomal loci. A matrix of the strain identification and the ST followed by the allele labels for each was used in the START (Sequence Type Analysis and Recombinational Tests, version 1.05) program to generate the UPGMA dendrogram. The program was written by Keith Jolley, University of Oxford (14). MLST group affiliation is indicated. Singletons are identified by an “s”; ST numbers followed by a blank belong to group 1.

FIG. 2.

Population snapshot of 231 Medicago-nodulating rhizobia derived from the allelic variation of 10 chromosomal loci. A matrix of the ST followed by the allele labels for each was used in eBURST (11) to generate a diagram of the evolutionary patterns among the strains. The snapshot was produced by setting the group definition to 0/10 genes. The sizes of the circles are related to the numbers of strains within each ST. The founder and cofounder genotypes are colored blue and red, respectively. Distances between STs indicated by connecting lines are arbitrary.