Soybean Root Nodule Occupancy: Competition Between Bradyrhizobium and Sinorhizobium Strains Inoculated at Different Plant Growth Stages

ABSTRACT Soybean is frequently nodulated by species from the Bradyrhizobium (BR) and/or Sinorhizobium (SR) genera. Several factors, such as soil pH, host genotype, geographic location, and other environmental variables, are reported to influence the preferential selection between BR or SR species within soybean root nodules. However, it remains unclear whether the age of the host plant at the time of inoculation affects preferential rhizobial selection. To investigate this, we inoculated soybean plants with different cell densities of BR and SR strains at three time points: at sowing (T0), two weeks after germination (T2), and four weeks after germination (T4). We used 16S rRNA gene amplicon sequencing of root nodule and rhizosphere samples to assess the relative abundance of BR and SR in nodule and rhizosphere. We observed a clear shift in nodule occupancy that favored BR at the time of seed sowing (T0) but increasingly favored SR when plants were inoculated at T2 and T4 stages. Specifically, at T4, SR dominated in nodules across all treatments, representing 88–99% of total sequences, regardless of applied inoculum ratio. In contrast, a similar number of sequences for both strains were detected in the rhizosphere at the time of the final harvest. These results highlight host age as an important ecological driver in legume–rhizobium interactions and suggest that inoculation time strongly influences microsymbiont selection. This information is important in understanding rhizobial competition and optimizing the timing of inoculation for soybean. IMPORTANCE Soybean is one of the world’s most valuable crops and fulfills most of its nitrogen requirements by developing symbiotic associated with nitrogen-fixing rhizobia. This reduces the need for chemical fertilizers by converting atmospheric nitrogen into a plant useable form of nitrogen. Multiple species from four rhizobial genera can nodulate soybean, and the plant’s choice of rhizobial partner is reported to change depending on environmental conditions such as pH, host genotype, geographic location, and other environmental factors. This study explores how the age of the soybean plant affects its preference for two frequently reported beneficial rhizobial species (B. diazoefficiens and S. fredii). By testing inoculation at different growth stages, we discovered that at early growth stages plants favored BR, while older plants increasingly selected SR for nodule formation. These findings highlight the level of complexity in plant–microbe interactions and could help optimize bioinoculant strategies for improving sustainability and crop yields.