Pesticide contaminants, such as chlorpyrifos (CP) and its more toxic metabolite 3,5,6-trichloro-2-pyridinol (TCP) in farmland drainage have attracted extensive concerns around the world, and are in urgent need of treatment. In this work, three groups of anaerobic reactors packed with zero valent iron (mono-ZVI system), ZVI coupled with anaerobic sludge (coupled system) and anaerobic sludge (mono-cell system) were continuously operated to investigate their performances on CP, TCP and chemical oxygen demand (COD) removal, as well as the reaction mechanisms of ZVI coupling with anaerobic microorganisms. Results showed that the removal efficiencies of CP and COD in coupled system were both around 95%. The corresponding results were 80% and 82% in mono-cell system, and the unstable values in mono-ZVI system were around 86% and 70%. The TCP residual concentration was the lowest in coupled system. The results of X-ray photoelectron spectrometer (XPS) and the photo images of ZVI at the end of reaction implied that the presence of sludge and the interaction between microorganisms and ZVI protected iron shavings from excessive corrosion. Moreover, the addition of ZVI promoted the electron transfer in the anaerobic system, which was proved by the higher value of electron transport system (ETS) activity and Cytochrome C (Cyt C) content of sludge with 250 mg (g h)(-1) and 20.71 nmol L-1 in coupled system, compared to 200 mg (g h)(-1) and 17.56 nmol L-1 in mono-cell system. The improvement of extracellular polymeric substances (EPS) and anaerobic sludge granulation in the coupled system were on account of the existence of ZVI. It was obvious that the key enzymes and microbial species functioned in COD, CP and TCP degradation were enriched in coupled system.