摘要

       [目的]抗生素在畜禽养殖生产中的大量使用,使得动物体及养殖场周围环境中的抗生素耐药菌日益增加,对人类健康和公共卫生安全造成威胁。氮循环是生态系统中重要的物质循环之一,抗生素在环境中的残留一定程度上阻碍氮循环进程,进而影响氮素平衡,扰乱环境微生物群落的功能代谢。通过监测抗生素进入土壤后对氮循环微生物及其耐药性造成的影响,有助于明确抗生素对菜地土壤微生物造成的危害。[方法]本文构建了氟苯尼考(FFC)-粪便-土壤模型,以FFC为胁迫因子,构建室内粪便-土壤模型,分别设空白组(0μg/kg)、低浓度药物组(0.167μg/kg)和高浓度药物组(16.67μg/kg),每组重复三次。于第1、3、7、14和30 d进行样品采集。采用宏基因组学和qPCR等技术分析FFC对土壤微生物耐药性和氮循环进程的影响。[结果]结果表明,低浓度FFC对土壤氮循环功能基因抑制范围广,而高浓度FFC抑制程度较大。此外,高浓度FFC明显提高土壤中floR、fexB、fexA耐药基因的丰度。FFC诱导了土壤微生物呈现不同的耐药图谱,表现在初期(1～3 d)部分优势ARGs类型丰度发生显著变化。研究还发现了高浓度FFC显著改变变形菌、放线菌、拟杆菌和厚壁菌菌群丰度,其中明显降低鞘脂单胞菌科中各类菌属丰度。值得注意的是,土壤理化性质是影响土壤优势菌属分布的主导因素。FFC还可降低蛋白质代谢功能,并影响大部分氮代谢功能;在氮代谢通路中,NO还原酶在FFC的胁迫下较敏感于其它酶,相关酶包括硝酸盐还原酶、亚硝酸盐还原酶和一氧化二氮还原酶亦呈现轻度响应。此外,FFC对携带氮循环基因丰度较高的菌属的影响随孵育时间延长而发生变化。[结论]总的来说,这些研究结果为养殖场合理使用兽用抗生素提供理论指导,也为评估抗生素引发微生物耐药性导致的环境生态风险提供参考数据。

       [Objective] The extensive use of antibiotics in livestock and poultry breeding has caused an increase in antibiotic-resistant bacteria in the animal body and the surrounding environment of the breeding farm, posing a threat to human health and public health safety. Nitrogen cycle is one of the important material cycles in the ecosystem. The residue of antibiotics in the environment hinders the nitrogen cycle process to a certain extent, thereby affecting the nitrogen balance and disturbing the functional metabolism of the environmental microbial community. By monitoring the impact of antibiotics on the nitrogen cycle microorganisms and their drug resistance after entering the soil, it is helpful to clarify the harm caused by antibiotics to the soil microorganisms in the vegetable field. [Method] In this paper, a florfenicol (FFC)-fecal-soil model was constructed. Using FFC as a stress factor, an indoor fecal-soil model was constructed. The blank group (0μg/kg) and the low-concentration drug group (0.167μg/ kg) and high-concentration drug group (16.67μg/kg), each group was repeated three times. Samples were collected on the 1, 3, 7, 14 and 30 days. Metagenomics and qPCR were used to analyze the effects of FFC on soil microbial resistance and nitrogen cycle processes. [Result] The results showed that low-concentration FFC has a wide range of inhibition of soil nitrogen cycle functional genes, while high-concentration FFC has a greater degree of inhibition. In addition, high concentrations of FFC significantly increased the abundance of floR, fexB, and fexA resistance genes in the soil. FFC induced different resistance profiles of soil microorganisms, which showed that the abundance of some dominant ARGs types changed significantly in the initial period (1～3 d). The study also found that high concentrations of FFC significantly changed the abundance of Proteobacteria, Actinomyces, Bacteroides and Firmicutes, and significantly reduced the abundance of various species in the Sphingomonas family. It is worth noting that the physical and chemical properties of soil are the dominant factors affecting the distribution of soil dominant bacteria. FFC can also reduce protein metabolism and affect most of the nitrogen metabolism. In the nitrogen metabolism pathway, NO reductase is more sensitive to other enzymes under the stress of FFC. Related enzymes include nitrate reductase, nitrite reductase and Nitrous oxide reductase also showed a mild response. In addition, the effect of FFC on bacteria with higher abundance of nitrogen cycle genes changes with the increase of incubation time. [Conclusion] In general, these research results provide theoretical guidance for the rational use of veterinary antibiotics in farms, and also provide reference data for evaluating the environmental and ecological risks caused by antibiotics-induced microbial resistance.

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