Continuous Cropping Is Associated with Shifts in Strawberry Root-Associated Bacterial Communities and Predicted...

Long-term cultivation of strawberries can alter plant-associated microbial communities; however, the responses of root-associated bacteria within the combined rhizoplane and endophytic community remain insufficiently characterized. This study compared bacterial community composition, co-occurrence patterns, and predicted ecological functions in root-associated bacteria from short-term (4 months; 4 mon) and long-term (16 months; 16 mon) cultivation of Dandong 99 strawberry plants. Because the two groups differed simultaneously in soil cultivation history, plant age, and root developmental stage, all observed differences are interpreted as correlates of cultivation duration rather than as effects of continuous cropping per season. The 6 composite root samples were analyzed using Illumina NovaSeq sequencing of the bacterial 16S rRNA V4 region, followed by ASV-based diversity analysis, taxonomic profiling, LEfSe, FastSpar-based co-occurrence network analysis, and FAPROTAX functional prediction. Across all samples, 264,350 high-quality sequences and 11,442 ASVs were obtained. Long-term cultivation was associated with lower observed ASV richness and a marked shift in community composition (PERMANOVA R² = 0.935; ANOSIM R = 1). At the phylum level, Actinobacteria declined significantly (30.50% to 12.85%), while Proteobacteria and Bacteroidota were enriched. At the genus level, Streptomyces, Steroidobacter, Bradyrhizobium, and unidentified Rhizobiaceae decreased significantly. Network analysis identified Sphingobium, Bradyrhizobium, and Steroidobacter as highly connected genera by degree centrality, with the latter two also showing reduced relative abundance. Functional prediction indicated significantly lower predicted chemoheterotrophy, aerobic chemoheterotrophy, nitrogen fixation, nitrate reduction, and chitinolysis, alongside higher predicted methylotrophy, methanol oxidation, ureolysis, and aromatic compound degradation. These findings provide genus-level evidence that cultivation duration is associated with compositional and predicted functional shifts in strawberry root-associated bacteria. The depleted Actinobacteria and Bradyrhizobium partially align with microbial signatures of continuous cropping obstacles, warranting future validation in age-controlled, temporally replicated studies.