Isolation and characterization of Rhizobia isolates from Glycine max

Abstract

The soybean plant (Glycine max), gradually gaining popularity as a cash crop in Bangladesh, plays a crucial role in the country's agriculture. However, intensive chemical fertilizers are used in its cultivation each year. The symbiotic relationship of soybean with rhizobia bacteria is vital for biological nitrogen fixation, where rhizobia converts atmospheric nitrogen into a form that plants can absorb and utilize. The identification of beneficial nitrogen-fixing rhizobia specific to the host can allow growers to select the right inoculum, which can increase yield and reduce usage of chemical fertilizers. The objective of this research was to identify and isolate nodule forming rhizobial strains, analyze their diversity, and characteristics. 30 nodule-associated bacteria were isolated from 8 Soybean plants. The isolates were classified based on their nodule formation capacities in the nitrogen free Jensen’s media (Sucrose 20 g/L, Dipotassium phosphate 1g/L, Magnesium sulphate 0.5g/L, Sodium chloride 0.5 g/L, Ferrous sulphate 0.1 g/L, Sodium molybdate 0.005 g/L, Calcium carbonate 2g/L and agar 15 g/L).via an in-vitro infection test. A series of biochemical tests such as Glucose Peptone Agar test, Bromothymol Blue test, Nitrate Reduction, Oxidase, Catalase and Urease tests were performed. Additionally, stress tests were performed by observing growth of the isolates in a range of pH and salt concentrations. Among the isolates some were found to successfully nodulate the soybean plants in-vitro. During the infection test, significant effects of rhizobial strain in the growth of plants’ lateral and tap roots, shoot is observed among the nodulating and non-nodulating plants. The rhizobia spp. was identified as stress-tolerant, slow-growing rhizobia spp. with capacity to withstand extreme low pH and salinity. All isolates were found to be positive for oxidase and nitrate reduction tests, and the majority were positive for urease tests as well. Moreover, the isolates were found to be capable of withstanding up to 6% NaCl. They could also withstand lower pH up to 4, making them ideal for commercial inoculum development in coastal regions and highly acidic soil.