The taxonomic position of isolates of a new genomic species (designated genomic species 2) obtained from several annual Medicago species and originating from different geographical locations was established through the results of phenotypic tests (including the of auxanographic and biochemical tests and symbiotic properties) and 16S rRNA phylogenetic inferences. A comparison of the complete 16S rRNA sequence of a representative of genomic species 2 (strain A 321 [T = type strain]) with the 16S rRNA sequences of other members of the and closely related taxa showed that genomic species 2 was phylogenetically related to , and . The levels of sequence similarity and observed numbers of nucleotide substitutions in strains indicated that A 321 and exhibited the highest level of sequence similarity (99.7%), with four nucleotide substitutions and one deletion. The results of a numerical analysis based on data from 63 auxanographic and biochemical tests clearly separated genomic species 2 isolates from . Genomic species 2 isolates nodulated and fixed nitrogen with , whereas isolates were ineffective and formed rudimentary nodules on this host plant. On the basis of phenotypic and 16S sequence analysis data, genomic species 2 isolates cannot be assigned to a previously described species. We propose that these isolates belong to a new species, .


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