Biochar-assisted biotransformation of 2,4,6-trinitrotoluene by Pseudomonas sp. CMR5c T22: synergistic mechanisms and metabolic reprogramming.
2,4,6-Trinitrotoluene (TNT) contamination presents serious threats to ecological safety and human health due to its high toxicity, carcinogenicity, and environmental persistence. Although microbial bioremediation is eco-friendly and cost-effective, its efficiency is often limited by low removal rates and weak microbial tolerance under TNT stress. This study utilized biochar to facilitate microbial biotransformation of TNT and systematically elucidated its synergistic mechanisms. Four types of biochar from different waste sources were screened, among which the wood biochar (MBC) exhibited the optimal performance. With MBC amendment, the TNT removal efficiency increased from 46.2% (strain-only control) to 88.7% within 48 h at an initial TNT concentration of 100 mg/L. Meanwhile, the first-order kinetic rate constant rose from 0.01323 h-1 (strain-only) to 0.03987 h-1, revealing that MBC greatly accelerated the TNT removal rate. Biochar accelerated removal kinetics, alleviated TNT-induced growth inhibition of strain T22 via lowering aqueous TNT concentration, and stimulated extracellular polymeric substance secretion. Untargeted metabolomics revealed 330 up-regulated differential metabolites in the biochar-microbe system, indicating intensive metabolic reprogramming. Mechanistically, MBC directly enhanced TNT transformation via activation of the nitrotoluene degradation and cofactor biosynthesis pathways. The synergistic mechanisms were proposed to include: (1) weakened TNT exposure; (2) enhanced antioxidant metabolism; (3) activated core transformation pathways; and (4) supplemented basal metabolic supply. Although complete mineralization and soil-scale validation are beyond the present scope, our findings establish a mechanistic foundation for designing biochar-microbe systems for enhanced nitroaromatic remediation.