Abstract:[Background] The synthetic progestin levonorgestrel has attracted wide attention due to its endocrine-disrupting effects. Microorganisms demonstrate extensive potential in pollutant degradation and environmental remediation. However, the characteristics and mechanisms of levonorgestrel degradation by microorganisms remain poorly understood. [Objective] To isolate an efficient levonorgestrel-degrading strain and explore its degradation mechanism for the remediation of levonorgestrel pollution. [Methods] We isolated a strain capable of efficiently degrading levonorgestrel from aquaculture wastewater and identified the strain based on its morphological, physiological, and biochemical characteristics and the 16S rRNA gene sequence. The degradation characteristics of the strain for levonorgestrel were investigated by whole genome analysis, degradation kinetics, and analysis of the degradation product spectrum. [Results] Gordonia sp. H52 was isolated, which had 99.0% homology of the 16S rRNA gene sequence compared with Gordonia cholesterolivorans Chol-3. This strain completely removed 0.5 mg/L levonorgestrel within 18 h. Liquid chromatography-tandem mass spectrometry revealed that Gordonia sp. H52 produced five metabolites during the degradation process of levonorgestrel, primarily through the cleavage of the side chain and hydroxy group. Furthermore, the strain facilitated the removal of various synthetic progestins in water, increasing the removal rates of levonorgestrel from 0% to 91.06%, acetylene norethindrone from 37% to 100%, and gestodene from 0% to 88.87%. [Conclusion] Gordonia sp. H52 demonstrates high degradation efficiency for levonorgestrel. This study provides valuable microbial resources and theoretical references for the treatment and remediation of water contamination caused by synthetic progestins.