To explore the enhancement effect of humic acid-modified biochar carrier on the removal of thiamethoxam by microorganisms, the removal effect and enhancement mechanism of thiamethoxam were evaluated by Enterobacter cloacae TMX-6 immobilized on humic acid-modified biochar in the liquid system. The results showed that humic acid modification successfully grafted oxygen-containing functional groups such as carboxyl, phenolic hydroxyl and quinonyl groups onto the surface of biochar. Scanning electron microscopy results indicated that TMX-6 exhibited high adhesion density and intact cell morphology on the surface of HBC. Under the optimal immobilization conditions (biochar concentration of 10 g/L, incubation time of 12 h), the 5-day removal rate of 10 mg/L thiamethoxam by HBC@TMX-6 reached 51%, which was 13% and 28% higher than that by the unmodified biochar inoculant and free bacteria, respectively. This system could significantly promote the secretion of extracellular polysaccharides and proteins by microorganisms, and exhibited high extracellular electron transfer activity. This process might be attributed to the fact that humic acid-modified biochar constructs a dual-channel electron transfer system, effectively accelerating the reduction and degradation of thiamethoxam. The results of this study provide a theoretical basis for the further application of carbon-based modified material-immobilized microorganisms in the remediation of pesticide-contaminated soil.