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The relationship between photosynthetic rhizobia that nodulate 10 Aeschynomene species (Aeschynomene afraspera, Aeschynomene denticulata, Aeschynomene evenia, Aeschynomene indica, Aeschynomene nilotica, Aeschynomene pratensis, Aeschynomene rudis, Aeschynomene scabra, Aeschynomene schimperi, and Aeschynomene sensitiva) and reference strains of the genera Bradyrhizobium, Rhizobium, and Azorhizobium was investigated by analyzing cellular fatty acid methyl esters (FAME) and 16S rRNA sequences. The members of each genus produced very distinct FAME patterns, and the photosynthetic rhizobia formed a subcluster in the Bradyrhizobium cluster. The absence of the cyc C19:0 type of fatty acid in all of the photosynthetic rhizobium strains isolated from 10 Aeschynomene species distinguished these microorganisms from other known rhizobia, including strain BTAi 1, a photosynthetic symbiont of A. indica. We sequenced a 264-base segment of the 16S rRNA genes of selected strains after amplification by the PCR and compared the results with previously published sequences for species of rhizobia and related photosynthetic bacteria. Photosynthetic strains IRBG 2 (from A. afraspera), IRBG 230 (from A. nilotica), and ORS 322 (from A. afraspera) had identical sequences but were distinct from strain BTAi (from A. indica) and from strain IRBG 231 (from A. denticulata), which is similar to the type strain (DNA homology group Ia) of Bradyrhizobium japonicum. Nonphotosynthetic strain IRBG 274 (from A. afraspera) was closely related to Bradyrhizobium elkanii (DNA homology group II). All of the photosynthetic rhizobia clearly fell into the Bradyrhizobium cluster. Although the results of the FAME and 16S rRNA analyses were in excellent agreement, our placement of the photosynthetic rhizobia is in apparent conflict with phenotypic data, as determined by numerical taxonomy (Ladha and So, Int. J. Syst. Bacteriol., in press) which placed the photosynthetic rhizobia in a coherent cluster that is as far from the genus Bradyrhizobium as the genera Rhizobium and Azorhizobium are. While the FAME and 16S rRNA data probably provide a more reliable indication of phylogeny, the degree of phenotypic divergence observed raises questions concerning the polyphasic approach to bacterial systematics.

Photosynthetic Symbionts of Aeschynomene spp. Form a Cluster with Bradyrhizobia on the Basis of Fatty Acid and rRNA Analyses