Hydrolysis of the neonicotinoid insecticide thiacloprid by the N2-fixing bacterium Ensifer meliloti CGMCC 7333

• A N2-fixing Ensifer meliloti CGMCC 7333 degrades the neonicotinoid thiacloprid.
• En. meliloti CGMCC 7333 shows the most rapid thiacloprid degradation.
• En. meliloti CGMCC 7333 Hydrolyzes thiacloprid to its amide metabolite.
• A nitrile hydratase mediates the hydrolysis of thiacloprid.

A thiacloprid (THI)-degrading bacterium CGMCC 7333 was isolated from soil and identified as the N2-fixing bacterium Ensifer meliloti. The major metabolite was identified as THI amide derived from the cyano moiety by hydrolysis of THI, using liquid chromatography-mass spectrometry and nuclear magnetic resonance analysis. En. meliloti CGMCC 7333 degraded 86.8% of 200 mg/L THI in 60 h with a half-life of 20.9 h and 90.9% of the reduced THI was converted to THI amide. CGMCC 7333 can convert THI to THI amide in the soil. Hydrolysis of THI by En. meliloti CGMCC 7333 is mediated by a nitrile hydratase (NHase) and the NHase gene cluster codes a cobalt-type NHase composed of an α-subunit, β-subunit, and accessory protein with lengths of 213, 219, and 128 amino acids, respectively. Whole cells of Escherichia coli Rosetta overexpressing NHase degraded 80.7% of THI (0.63 mmol/L) in 10 min and formed 0.58 mmol/L of THI amide, and the half-life of THI degradation was 5.2 min. The purified NHase degraded 80.6% of THI (0.70 mmol/L) and formed 0.60 mmol/L of THI amide in 5 min with a molar conversion rate of 85.7%, and the half-life of THI degradation was 6.9 min.