In this study, two pyridine-degrading strains namely Enterobacter cloacae complex sp. BD17 and Enterobacter sp.BD19 were isolated from the aerobic tank of a pesticide wastewater treatment plant. The mixed bacteria H4 composed of BD17 and BD19 at a ratio of 1:1 was immobilized by Solidago canadensis L. stem biochar with a dosage of 2 g·L−1. The highest pyridine removal rate of 91.70% was achieved by the immobilized H4 at an initial pyridine concentration of 200 mg·L−1, pH of 7.0, temperature of 28 °C and salinity of 3.0% within 36 h. The main intermediates of pyridine degradation by BD17 were pyridine-2-carboxamide, 2-aminopropanediamide, and 2-aminoacetamide, while 2-picolinic acid, isopropyl acetate, isopropyl alcohol, and acetaldehyde were identified with BD19 by adopting GC-MS technique. Interestingly, there was a possibility of totally mineralization of pyridine and the corresponding degradation pathways of BD17 and BD19 were revealed for the first time.