3, 5, 6-Trichloro-2-pyridinol (TCP) is a metabolite of the insecticide chlorpyrifos and the herbicide triclopyr, and it is higher toxic than the parent compounds. Microbially-mediated mineralization appears to be the primary degradative pathway and the important biological process of detoxification. However, little information is available on TCP complete metabolic pathways and mechanisms. In this study, the degradation of TCP was studied with a novel strain Micrococcus luteus ML isolated from a stable TCP degrading microbiota. Strain ML was capable of degrading 61.6% of TCP (50 mg/L) and 35.4% of chlorpyrifos (50 mg/L) at 24 h and 48 h under the optimal conditions (temperature: 35 °C; pH: 7.0), respectively. It could also degrade 3, 5-dichloro-2-pyridone, 6-chloropyridin-2-ol, 2-hydroxypyridine and phoxim when provided as sole carbon and energy sources. Seven TCP intermediate metabolites were detected in strain ML and two possible degradation pathways of TCP were proposed on the basis of LC–MS analysis. Both the hydrolytic-oxidative dechlorination pathway and the denitrification pathway might be involved in TCP biodegradation by strain ML. To the best of our knowledge, this is the first report on two different pathways responsible for TCP degradation in one strain, and this finding also provides novel information for studying the metabolic mechanism of TCP in pure culture.