Antibiotics and antibiotic resistance genes (ARGs) in sewage sludge can cause high ecotoxicological risks in the environment and public health concerns. The aims of this study were to establish enzymatic integrated in-situ advanced anaerobic digestion (AAD) by adding cellulase and papain as well as the two enzymes combined with zero valent iron (ZVI) directly into the anaerobic digesters to explore the removal of antibiotics and ARGs under the mesophilic condition (35 °C). The methane production potential during in-situ AAD was effectively improved. Papain and cellulase at 30 mg/gTSS were most effective in improving antibiotic removal. The removal of sulfamerazine (SMZ) and sulfadiazine (SMR) could reach 89.10 % and 71.75 %. Combined enzymes with ZVI also enhanced the removal of all target antibiotics, especially roxithromycin (ROX), SMZ and SMR most significantly. Except for sul1, tetA and tetB, the removal of ARGs by papain reached 6.33 %-82.15 %. The addition of cellulase effectively improved tetA removal. The combination of biological enzymes further enhanced the removal of qnrS and ermX. The tetG, tetB, sul3, ermX, ermT, qnrS, and aac(6')-IB-CR by combined enzymes with ZVI could even not be detected after digestion. Addition of papain, cellulase, and ZVI caused variations in the dominant bacteria. All target antibiotics presented significant positive correlations with the genera norank_f__Bacteroidetes_vadinHA17, norank_f__norank_o__SJA-15, norank_f__norank_o__Aminicenantales. Redundancy analysis showed archaea Methanosaeta and Candidatus_ Methanoacidiosum genera greatly contributed to antibiotics removal with the combination of enzymes and ZVI. Co-occurrence network analysis indicated the removal of ARGs was mainly based on the changes of existence of host bacteria.
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