In addition to rhizobia, other non-symbiotic endophytic bacteria also have been simultaneously isolated from the same root nodules. The existence of non-symbiotic endophytic bacteria in leguminous root nodules is a universal phenomenon. The vast majority of studies have detected endophytic bacteria in other plant tissues. In contrast, little systemic observation has been made on the non-symbiotic endophytic bacteria within leguminous root nodules. The present investigation was carried out to isolate plant growth-promoting endophytic non-symbiotic bacteria from indigenous leguminous Sphaerophysa salsula and their influence on plant growth. A total of 65 endophytic root nodule-associated bacteria were isolated from indigenous legume S. salsula growing in the northwestern arid regions of China. When combining our previous work with the current study, sequence analysis of the nifH gene revealed that the strain belonging to non-nodulating Bacillus pumilus Qtx-10 had genes similar to those of Rhizobium leguminosarum Qtx-10-1. The results indicated that horizontal gene transfer could have occurred between rhizobia and non-symbiotic endophyties. Under pot culture conditions, out of the 20 representative endophytic isolates, 15 with plant growth-promoting traits, such as IAA production, ACC deaminase, phosphate solubilization, chitinase, siderophore, and fungal inhibition activity showed plant growth-promoting activity with respect to various plant parameters such as chlorophyll content, fresh weight of plant, shoot length, nodule number per plant and average nodule weight per plant when co-inoculated with rhizobial bioinoculant Mesorhizobium sp. Zw-19 under N-free culture conditions. Among them, Bacillus pumilus Qtx-10 and Streptomyces bottropensis Gt-10 were excellent plant growth-promoting bacteria, which enhanced the seeding fresh weight by 87.5% and the shoot length by 89.4%, respectively. The number of nodules grew more than 31.89% under field conditions. Our findings indicate the frequent presence of these non-symbiotic endophytic bacteria within root nodules, and that they help to improve nodulation and nitrogen fixation in legume plants through synergistic interactions with rhizobia.
Keywords: Non-symbiotic endophytes; Phylogeny; Plant growth promotion; Root nodules; Sphaerophysa salsula.