Aims <p>Soil organic carbon (SOC) persistence is essential for sustaining agroecosystem productivity and mitigating carbon loss. However, the influence of carbon input type and application rate on microbial carbon use efficiency (CUE) and SOC persistence remains unclear. This study examined how straw, manure, and biochar, applied at two rates, affect SOC persistence and soil functional microbiota.</p> Methods <p>To address this knowledge gap, a short-term pot experiment was conducted with three types of carbon input (straw, manure, and biochar), at full and half doses under uniform fertilization conditions. SOC content, chemical composition, enzyme activity, microbial biomass, microbial necromass carbon (MNC), and gene abundances related to carbon degradation pathways were analyzed to reveal microbial-mediated SOC persistence mechanisms.</p> Results <p>Straw and manure enhanced SOC persistence primarily via microbial-driven processes, including increased microbial activity, CUE, and MNC accumulation. Manure achieved the highest SOC accumulation due to balanced nutrient availability. Biochar stabilized SOC mainly through physic chemical protection afforded by its recalcitrant structure and porous matrix. Higher carbon input rates generally promoted SOC persistence, although excessive straw or manure reduced CUE. Fungal necromass carbon (FNC) was critical for long-term SOC persistence.</p> Conclusions <p>Straw inputs increase SOC persistence through ‘fungal-chemical reorganization’, whereas manure increases it through ‘bacterial-rapid turnover’ and biochar through ‘physico-chemical dominance’. Moreover, SOC persistence increases with higher application rates. Furthermore, FNC plays a pivotal role.</p>