Multi-Trait Screening Identifies Cold-Adapted Soybean Rhizobia from Northeastern China as Candidate Inoculant Strains

Developing effective rhizobial inoculants for high-latitude soybean production requires strains that are both well-adapted to local soil conditions and capable of sustaining symbiotic nitrogen fixation under abiotic stress. In this study, 66 indigenous rhizobial strains isolated from four ecological regions of Heilongjiang Province, China, i.e., the Songnen Plain, Sanjiang Plain, Northwest Arid Sandy Region, and Daxing'anling-Xiaoxing'anling mountainous area, were characterized for phenotypic diversity, abiotic stress tolerance, symbiotic nitrogen fixation capacity, and plant growth-promoting (PGPR) traits. Carbon and nitrogen source utilization, physiological and biochemical properties, and tolerance to salinity, pH extremes, temperature, and drought were assessed and analyzed by hierarchical cluster analysis. The strains showed habitat-associated phenotypic differentiation, with stress-tolerance traits accounting for the greatest proportion of variation. Isolates from the Northwest Arid Sandy Region displayed the broadest stress tolerance, while Sanjiang Plain isolates showed more conservative phenotypic profiles. Eight representative strains were subsequently evaluated for symbiotic performance and PGPR activity. Nodule dry weight, nitrogenase activity, and total plant nitrogen content were strongly intercorrelated and appeared more reliable indicators of nitrogen-fixation efficiency than nodule number alone. Songnen Plain strains SN1 and SN8 showed the highest symbiotic performance, while Northwest Arid Sandy Region strain FS1 exhibited the greatest ACC deaminase activity. A two-dimensional ranking framework integrating symbiotic and non-symbiotic traits ordered the strains as SN1 > SN8 > SJ1 > SJ11 > DX5 > DX1 > FS7 > FS1, identifying SN1 and SN8 as the most promising candidates requiring field validation for inoculant applications. These findings suggest that multi-trait evaluation frameworks may offer a practical and reproducible approach to screening indigenous rhizobia for region-specific biofertilizer applications in cold-region soybean agriculture.