Surviving and thriving in terms of symbiotic performance of antibiotic and phage-resistant mutants of Bradyrhizobium of soybean [Glycine max (L.) Merrill].

Rhizobial inoculation plays an important role in yielding enhancement of soybean, but it is frequently disturbed by competition with bacterial population present in the soil. Identification of potential indigenous rhizobia as competitive inoculants for efficient nodulation and N(2)-fixation of soybean was assessed under laboratory and field conditions. Two indigenous bradyrhizobial isolates (MPSR033 and MPSR220) and its derived different antibiotic (streptomycin and gentamicin) and phage (RT5 and RT6)-resistant mutant strains were used for competition study. Nodulation occupancy between parent and mutant strains was compared on soybean cultivar JS335 under exotic condition. Strain MPSR033 Sm(r) V(r) was found highly competitive for nodule occupancy in all treatment combinations. On the basis of laboratory experiments four indigenous strains (MPSR033, MPSR033 Sm(r), MPSR033 Sm(r) V(r), MPSR220) were selected for their symbiotic performance along with two exotic strains (USDA123 and USDA94) on two soybean cultivars under field conditions. A significant symbiotic interaction between Bradyrhizobium strains and soybean cultivar was observed. Strain MPSR033 Sm(r) V(r) was found superior among the rhizobial treatments in seed yield production with both cultivars. The 16S rRNA region sequence analysis of the indigenous strains showed close relationship with Bradyrhizobium yuanmingense strain. These findings widen out the usefulness of antibiotic-resistance marked phage-resistant bradyrhizobial strains in interactive mode for studying their symbiotic effectiveness with host plant, and open the way to study the mechanism of contact-dependent growth inhibition in rhizobia.

Genotypic characterization of phage-typed indigenous soybean bradyrhizobia and their host range symbiotic effectiveness.

Analysis of genetic diversity among indigenous rhizobia and its symbiotic effectiveness with soybean cultivar is important for development of knowledge about rhizobial ecology. In India, little is known about the genetic resources and diversity of rhizobia nodulating soybean. Indigenous bradyrhizobia isolated from root nodules of soybean plants, collected from traditional cultivating regions of two states (Madhya Pradesh and Uttar Pradesh) of India, were screened for bacteriophage sensitivity to identify successful broad host range symbiotic effectivity. Of 172 rhizobial isolates, 91 showed sensitivities to eight lytic phages and form ten groups on the basis of sensitivity patterns. The genetic diversity of 23 isolates belonging to different phage groups was assessed along with that of strains USDA123 and USDA94 by the restriction fragment length polymorphism (RFLP) analysis of 16S rDNA, intergenic spacer (IGS) (16S-23S rDNA), and DnaK regions. RFLP analysis of 16S rDNA formed 5 groups, whereas 19 and 9 groups were revealed by IGS and the DnaK genes, respectively. The IGS regions showed many amplified polymorphic bands. Nine isolates which revealed high RFLP polymorphism in the abovementioned regions (16S rRNA, IGS, DnaK) were used for 16S rRNA sequence analyses. The results indicate that taxonomically, all isolates were related to Rhizobium etli, Bradyrhizobium spp., and Bradyrhizobium yuanmingense. The doubling time of isolates varied from 9 h (MPSR155) to 16.2 h (MPSR068) in YM broth. Five isolates which did not show cross infectivity with isolated phage strains were studied for symbiotic efficiency. All isolates showed broad host range symbiotic effectiveness forming effective nodules on Vigna mungo, Vigna radiata, Vigna unguiculata, and Cajanus cajan. The present study provides information on genetic diversity and host range symbiosis of indigenous soybean rhizobia typed by different phages.

Identification and assessment of symbiotic effectiveness of phage-typed Rhizobium leguminosarum strains on lentil (Lens culinaris Medik) cultivars.

Symbiotic effectiveness of 19 indigenous and two exotic (USDA 2426 and USDA 2431) strains of lentil Rhizobium belonging to different phage-sensitive and phage-resistant groups was compared under axenic condition. Four strains (USDA 2431, BHULR 104, BHULR 113, and BHULR 115) sensitive to different phages were found significantly superior over others in terms of nodule number, acetylene reduction activity, and total dry weight per plant. Inoculation response of these strains was then evaluated on six lentil cultivars under field condition. A significant symbiotic interaction between rhizobial strains and lentil cultivars was observed. Grain yield enhancement was noticed by the compatible interaction of lentil cultivars HUL-57, L-4147, K-75, and PL-4/DPL-15/DPL-62 with rhizobial strains USDA 2431, BHULR 104, BHULR 113, and BHULR 115, respectively. The authentication of rhizobial strains was accomplished through 16S rDNA sequence analysis. All rhizobial strains had close matching with R. leguminosarum bv. viciae strains. The results have shown that phages can trustfully help selecting out the symbiotically efficient most rhizobial strains for advantageous use with lentil cultivars, in order to strengthen the BNF-based future lentil breeding programs.

Differential symbiotic response of phage-typed strains of Bradyrhizobium japonicum with soybean cultivars.

In this study, native Bradyrhizobium strains were isolated from the host plant, Glycine max, harvested from fields in Madhya Pradesh, India, and were typed by lytic rhizobiophages. Eight indigenous (Soy2, ASR011, ASR031, ASR032, MSR091, ISR050, ISR076 and ISR078) and two exotic strains (USDA123 and CB1809), all of which evidenced a distinct reaction with six phages, were employed in this study. The symbiotic interaction of these strains was studied initially using soybean cultivar JS335 in a sand culture in a controlled environment, and the efficiency was assessed based on the nodule number, nodule dry weight, plant dry weight, nitrogenase activity, and total accumulation of N per plant. Symbiotic effectiveness was found to be highest with the native phage-sensitive isolate ASR011, whereas it was at a minimum with the phage-resistant isolates, ISR050 and ISR078. Additionally, the effectiveness of these strains was evaluated using six soybean cultivars belonging to different maturity groups; namely, Bragg, Lee, Pusa20, PK416, JS335 and NRC37. Analysis of variance data evidenced significant differences due to both symbionts, for the majority of the tested parameters. The CB1809, USDA123, and ASR011 strains evidenced relatively superior symbiotic effectiveness with soybean cultivars Bragg, Lee and JS335. Strain ISR078 evidenced no significant responses with any of the cultivars. The ASR031 strain performed moderately well with all tested cultivars. The symbiotic response of all the strains was quite poor with cultivar PK416. Our studies showed that a significant relationship existed between the phage sensitivity and symbiotic efficiency of the bacterial strains with the host-cultivars.