Chloramphenicol acetate

The risk of environmental pollution caused by chloramphenicol has necessitated special attention. Biodegradation has tremendous potential for chloramphenicol removal in the environment. Six chloramphenicol-degrading consortia were acclimated under different culture conditions to investigate their chloramphenicol biodegradation behaviors, and the bacterial community structures were comprehensively characterized. The enriched consortia CL and CH which utilized chloramphenicol as their sole carbon and energy source could thoroughly degrade 120 mg/L chloramphenicol within 5 days, and the mineralization rate reached up to 90%. Chloramphenicol biodegradation kinetics by different enriched consortia fit the modified Gompertz model or the first-order decay model (R2≥0.97). Consortia CL could almost completely degrade 1-500 mg/L CAP with a final mineralization rate of 87.8-91.7%. Chloramphenicol 3-acetate was identified to be a major intermediate of CAP biodegradation by metabolite analysis and enzyme activity assay. 16S rRNA sequencing analysis revealed that the diversities and abundances of the main genera in the enriched consortia were distinct from each other. Forty-one core OTUs belonging to 18 genera were the core bacteria which might be related to chloramphenicol biodegradation. Among them, the genera Sphingomonas, Chryseobacterium, Cupriavidus, Bradyrhizobium, Burkholderia, and Afipia with high abundance may play potential roles for chloramphenicol biodegradation.

Chloramphenicol and florfenicol are broad-spectrum antibiotics. Although the bacterial resistance mechanisms to these antibiotics have been well documented, hydrolysis of these antibiotics has not been reported in detail. This study reports the hydrolysis of these two antibiotics by a specific hydrolase that is encoded by a gene identified from a soil metagenome. Hydrolysis of chloramphenicol has been recognized in cell extracts of Escherichia coli expressing a chloramphenicol acetate esterase gene, estDL136. A hydrolysate of chloramphenicol was identified as p-nitrophenylserinol by liquid chromatography-mass spectroscopy and proton nuclear magnetic resonance spectroscopy. The hydrolysis of these antibiotics suggested a promiscuous amidase activity of EstDL136. When estDL136 was expressed in E. coli, EstDL136 conferred resistance to both chloramphenicol and florfenicol on E. coli, due to their inactivation. In addition, E. coli carrying estDL136 deactivated florfenicol faster than it deactivated chloramphenicol, suggesting that EstDL136 hydrolyzes florfenicol more efficiently than it hydrolyzes chloramphenicol. The nucleotide sequences flanking estDL136 encode proteins such as amidohydrolase, dehydrogenase/reductase, major facilitator transporter, esterase, and oxidase. The most closely related genes are found in the bacterial family Sphingomonadaceae, which contains many bioremediation-related strains. Whether the gene cluster with estDL136 in E. coli is involved in further chloramphenicol degradation was not clear in this study. While acetyltransferases for chloramphenicol resistance and drug exporters for chloramphenicol or florfenicol resistance are often detected in numerous microbes, this is the first report of enzymatic hydrolysis of florfenicol resulting in inactivation of the antibiotic.

Chloramphenicol-3-acetate esterase activity was detected in cell-free extracts of strains of Streptomyces venezuela, Streptomyces sp. and Streptosporangium viridogriseum var. kofuense which produced chloramphenicol and also Corynebacterium hydrocarboclastus which produced chloramphenicol analogs (corynecins). None of the cell-free extracts of chloramphenicol- or corynecin-producing strains possessed chloramphenicol acetyltransferase activity under conditions which avoided the influenced of the esterase activity. Among 20 strains examined that did not produce chloramphenicol, chloramphenicol acetyltransferase was detected in cell-free extracts of one strain of Streptomyces coelicolor Müller and one strain of S. fradiae ISP5063.