一株霍氏肠杆菌对四环素的降解作用及其降解产物的毒性评估

Biodegradation of tetracycline by an Enterobacter hormaechei strain and toxicity of degradation products

摘要:

【目的】抗生素作为新兴污染物,已经引起社会的极大关注。针对四环素有效降解菌株缺乏这一现状,本研究旨在筛选和鉴定具有降解四环素功能的菌株,分析其降解特性和降解作用类型、初步探讨其降解活性物质定位并评估其降解产物的生理毒性。【方法】以四环素为唯一碳源,从受四环素污染的猪场污泥中筛选四环素降解菌株;结合菌落形态学特征、生理生化特征、扫描电镜观察和16S rRNA基因测序鉴定菌株,通过不同外源碳、pH及去除动力学阐明菌株对四环素的降解特性,提取菌株不同成分探讨其去除四环素的作用类型,并进一步从细胞内液和细胞外液开展生物降解的活性物质定位,最后评估降解产物的生理毒性。【结果】筛选鉴定得到一株霍氏肠杆菌(Enterobacter hormaechei) MEH2305,pH为7.0和添加10 g/L外源碳胰蛋白胨是其发挥降解作用的最适条件。MEH2305通过非生物降解和生物降解的共同作用,在培养第7天对四环素总去除率达到68% (对土霉素和盐酸强力霉素去除率分别为53%和56%),其分泌的细胞内液和细胞外液对四环素的去除率分别为40.77%和31.18%。同时,与未经MEH2305处理的四环素对照组比较,MEH2305降解四环素的产物对革兰氏阴性大肠杆菌(Escherichia coli) K88和革兰氏阳性枯草芽孢杆菌(Bacillus subtilis) 168的生理毒性作用显著降低。【结论】MEH2305可以作为一株潜在的有效且安全的四环素降解菌株,应用于抗生素的环境治理领域。

Abstract:

【Objective】 Antibiotics as emerging pollutants have aroused wide concern. In view of the shortage of effective tetracycline-degrading strains, this study aims to screen and identify the strains for tetracycline degradation, analyze degradation properties and type, pinpoint the localization of active substances for bio-degradation, and evaluate the physiological toxicity of degradation products. 【Methods】 Tetracycline was used as the sole carbon source to screen out the target strain from tetracycline-contaminated pig sludge. The strain was identified based on colony morphology, physiological and biochemical characteristics, scanning electron microscopy images, and the 16S rRNA gene sequence. Different carbon sources, pH, and removal kinetics were employed to characterize the degradation process of the strain. Different components of the strain were extracted to determine the degradation type of tetracycline by the strain. Furthermore, the intracellular and extracellular fluids of the strain were used to degrade tetracycline, so as to determine the location of the active substance for degradation. Finally, the toxicity of the degradation products was assessed. 【Results】 The strain MEH2305 was screened out and identified as Enterobacter hormaechei, which showed the best degradation performance at pH 7.0 and with tryptone as the carbon source. Strain MEH2305 showed a total tetracycline removal rate of 68% on the 7th day of culture via abiotic degradation and bio-degradation, and the removal rates of oxytetracycline and doxycycline hydrochloride were 53% and 56%, respectively. The tetracycline removal efficiency by the intracellular and extracellular fluids of MEH2305 was 40.77% and 31.18%, respectively. Compared with tetracycline control without MEH2305, the tetracycline degradation products of MEH2305 had reduced physiological toxicity on Gram-negative Escherichia coli K88 and Gram-positive Bacillus subtilis 168. 【Conclusion】 The strain MEH2305 can be used as an effective and safe tetracycline-degrading strain for the treatment of antibiotics in the environment.