Bacterial Diversity in the Root Nodules and Rhizosphere of Chickpea (Cicer arietinum L.)

Abstract

The main objective of the present study was to study bacterial diversity in the root nodules and rhizosphere of chickpea varieties growing in different regions of Pakistan by cultivation on growth media as well as by using culture-independent DNA-based techniques. A total of 60 isolates, including symbiotic (10 isolates) as well as free-living bacteria (50 isolates), were purified from “Desi-type” and “Kabuli-type” chickpea varieties collected from 5 different localities. In pure culture, maximum IAA production was recorded in Kocuria sp. RTL99 (37.77 μg/mL) and maximum phosphate solubilization was recorded in Serratia sp. 5D (119.94 μg/mL). Among the bacterial inocula tested in pot and field experiments, co-inoculation of Mesorhizobium sp. NTY7 and Ensifer sp. NFY8 was found to be the most effective treatment at all localities and on both varieties of chickpea. To investigate bacterial diversity by culture-independent DNA-based technique, DNA extracted from root nodules and rhizospheric soil was used for pyrosequencing of 16S rRNA and nifH genes. 16S rRNA sequences originating from the nodules revealed occurrence of 10 bacterial phyla. At genus level, 16S rRNA sequences of 111 genera (70.78 % of the total sequences) of culturable bacteria were retrieved from nodule DNA along with 29.22 % sequences of “uncultured” bacteria. In the nodules, a significant fraction i.e., 52.77 % of 16S rRNA sequences and 88.83 % of the nifH sequences among the total sequences retrieved from all sites belonged to genus Mesorhizobium. The 16S rRNA sequences originating from the rhizospheric soil revealed enormous diversity of 22 bacterial phyla. At genus level, 16S rRNA sequences of 313 genera (29.72 % of the total sequences) of culturable bacteria were retrieved from rhizospheric soil DNA along with 70.28 % sequences of “uncultured” bacteria. Mesorhizobial 16S rRNA and nifH sequences retrieved from rhizospheric soil comprised 0.265 % and 16.68 % of the total recovered sequences, respectively. In the present study, sequences related to well-known plant growth promoting rhizobacteria Serratia spp. were frequently detected, which lead to targeted isolation of two Serratia strains from the nodules. Both the isolates showed growth improvement of chickpea when used as inoculants for chickpea grown at different localities.