Occurrence of two endophytic associative nitrogen-fixing Caulobacter spp., from three non-nodulating endemic legumes based on nifH gene analysis

Abstract

Legumes are cosmopolitan plants, and nodulating legumes are well known for their symbiotic nitrogen-fixing ability by rhizobia-legume interaction. Biological nitrogen fixation in non-nodulating endemic legumes by associative or endophytic symbiotic bacteria requires much needed attention. Three non-nodulating legumes, namely, Humboldtia brunonis Wall., Kunstleria keralensis C.N. Mohanan and N.C. Nair, and Bauhinia phoenicea Wight and Arn., endemic to the Western Ghats regions of Karnataka state, were studied. Employed techniques of selective culture media to understand diazotrophic diversity inside the roots of these plants. The isolates that can grow in a nitrogen-free semisolid agar medium have been considered positive for nitrogen-fixing ability. nifH gene is taken as the marker gene to ascertain the nitrogen-fixing ability of the bacteria. The qualified bacterium in the previous steps is identified using 16s RNA sequencing and the Sanger sequence method. The results obtained showed B. phoenicea Wight and Arn. and H. brunonis Wall. as Caulobacter segnis, and in K. keralensis C.N. Mohanan & N.C. Nair, it is Caulobacter crescentus. The presence of the nifH gene is demonstrated through molecular methods. This work adds to the diverse works of Caulobacter as a successful plant growth-promoting endophyte even in the nitrogen-deficient, slopy soils of the Western Ghats.