Biochar is useful for soil organic carbon (SOC) sequestration. However, the effects of biochar aging and addition rates on SOC stabilization are unclear. A field experiment with four biochar application rates (0% (control), 1% (LB), 2% (MB), and 4% (HB) of dry fluvo-aquic soil) was conducted. Soil samples were sampled after 8, 12, and 24 months of its application to clarify the question. In general, SOC gradually increased with the biochar application rate. SOC with HB was higher than that in other treatments, while the ratio of microbial biomass carbon (MBC)/SOC and readily oxidizable carbon (ROC)/SOC with HB was lower than that in other treatments (p < 0.05), indicating a positive effect of HB for C stabilization over time. The effects of biochar on the soil carbon pool management index (CPMI) changed from negative to positive after 8 and 24 months of biochar application. The activities of β-D-glucosidase (βG), cellobiohydrolase (CBH), and β-N-acetylglucosaminidase (NAG) under HB were higher than with other treatments after 12 and 24 months of biochar application (p < 0.05) and negatively correlated with the ratio of MBC/SOC and ROC/SOC over time. The CPMI was positively related with βG and CBH activities after 8 and 24 months of biochar application, respectively (p < 0.05). HB increased the relative abundance of oligotrophs, including Acidobacteria, Actinobacteria, and Chloroflexi, but decreased the relative abundance of copiotrophs, including γ-Proteobacteria and Bacteroidetes over time (p < 0.05). The ratio of dissolved organic carbon (DOC)/SOC was positively correlated with the bacterial oligotroph/copiotroph ratio and significantly affected the oligotrophic and copiotrophic bacterial communities, especially after 8 and 12 months of biochar application (p < 0.05). These findings reinforce that increasing the biochar application rate and time enhances SOC stabilization by decreasing the proportions of labile organic carbon and making oligotrophic/copiotrophic communities and enzyme activities more conducive to C sequestration.

This study aimed to investigate the legacy effects of prolonged brackish irrigation on rhizobacterial communities and agricultural productivity in wheat. Here, we conducted pot experiments to investigate the mechanisms through which different irrigation regimes (irrigation using brackish groundwater and normal water) regulate wheat production. We applied four irrigation treatments across different stages of wheat growth (early stages, seedling-to-jointing, and late stages, jointing-to-maturity). This included irrigation exclusively using normal water during both stages (RR), using normal water followed by brackish groundwater (RW), exclusively using brackish groundwater (WW), and using brackish groundwater followed by normal water (WR). Under the premise of retaining 10 seedlings per pot, the average number of effective spikes per 10 plants in the RR, RW, and WR treatments was approximately 1.3, 1.1, and 1.1 times that of WW (19 ± 1), respectively. The spike weight per 10 plants in the RR, RW, and WR treatments was approximately 1.8, 1.5, and 1.3 times that of WW (12.75 ± 1.74 g), respectively. Compared with brackish groundwater irrigation, the use of normal water during the early stages significantly reduced the relative abundance of Pseudomonadota and increased that of Chloroflexota (p < 0.05). The number of effective spikes was positively correlated with the relative abundances of Actinobacteriota, Acidobacteriota, Chloroflexota, and Bacteroidota, but negatively correlated with the abundance of Pseudomonadota (p < 0.05). Irrigation regimes altered the rhizobacterial community structure. However, the legacy effect of long-term irrigation using brackish groundwater resulted in the dominance of stochastic processes in bacterial community assembly and stability of the Shannon diversity across all treatments. The complexity of the rhizobacterial co-occurrence network was lower in the RW treatments than that in the WW treatments (p < 0.05). Structural equation modeling revealed that irrigation using normal water during early stages boosted the number of effective spikes in wheat. This enhancement was achieved by increasing rhizobacterial diversity, reducing rhizosphere sodium, and simplifying the microbial network. This study challenges the “legacy effect” of brackish water irrigation by demonstrating that optimal irrigation timing is key to enhancing crop yield.