Chickpeas, lentils, and peas are the oldest grain legume species that spread to other regions after their first domestication in Fertile Crescent, and they could reveal the rhizobial evolution in relation to the microsymbionts of wild species in this region. This study investigated the phenotypic and genotypic diversity of the nodule-forming rhizobial bacteria recovered from Pisum sativum subsp., Cicer pinnatifidum, and Lens culinaris subsp. orientalis exhibiting natural distribution in the Gaziantep province of Turkey. PCA analyses of rhizobial isolates, which were tested to be highly resistant to stress conditions, showed that especially pH and salt concentrations had an important effect on these bacteria. Phylogenetic analysis based on 16S rRNA determined that these wild species were nodulated by at least 7 groups including Rhizobium and non-Rhizobium. The largest group comprised of Rhizobium leguminosarum and Rhizobium sp. while R. pusense, which was previously determined as non-symbiotic species, was found to nodulate C. pinnatifidum and L. culinaris subsp. orientalis. In recent studies, Klebsiella sp., which is stated to be able to nodulate different species, strong evidences have been obtained in present study exhibiting that Klebsiella sp. can nodulate C. pinnatifidum and Pseudomonas sp. was able to nodulate C. pinnatifidum and P. sativum subsp. Additionally, L. culinaris subsp. orientalis unlike other plant species, was nodulated by Burkholderia sp. and Serratia sp. associated isolates. Some isolates could not be characterized at the species level since the 16S rRNA sequence similarity rate was low and the fact that they were in a separate group supported with high bootstrap values in the phylogenetic tree may indicate that these isolates could be new species. The REP-PCR fingerprinting provided results supporting the existence of new species nodulating wild ancestors.
Keywords: 16S rRNA; Genotypic diversity; Nodulation; Phenotypic diversity; Rhizobium; Wild legume.