养殖场粪便好氧堆肥过程中抗性基因的分布与削减机制研究

Plenty of antibiotics were used as animal feed additive to avoid disease in the large-scale livestock and poultry farm. The manure discharged would carry drug-resistance bacteria and lots of antibiotic resistance genes ARGs when the livestock and poultry ate much feedstuff with antibiotics, which would bring great environmental risk to land if those manures were composted and used as fertilizer. Aiming for analyzing the distribution rule and reduction mechanism of drug-resistance bacteria and resistance gene during livestock manure composting process, the methods of chromatographic analysis, tradition microbial culture, molecular biology and metagenomics were used to monitor the antibiotics residue and variation of microbial community along with the full composting process. The classification and evaluation of ARGs, the mechanism of enhancing antibiotics degradation, the responding relationship and variation of residue antibiotics, drug-resistance bacteria and resistance gene, the variation of microbial community, the gene transfer and reduction of ARGs along with the composting process and the safe evaluation of compost product were carried out to establish a modified composting process based on antibiotics and ARGs reduction and achieve the safe land utilization of livestock manure. The research will have great theoretical significance and applicable value, which will be helpful to reduce ARGs diffusion in the environment, to supply safe manure compost and to improve china soil quality.

为防治畜禽病疫，规模化畜禽养殖场在饲料中添加了大量的抗生素药物，长久食用含药饲料的畜禽则产生含有大量耐药菌与抗性基因的粪便，这种粪便的堆肥农用将带来环境风险。本项目针对畜禽粪便堆肥工艺中耐药菌与抗性基因分布规律与去除机制不清晰的科学问题，采用色谱分析、传统微生物培养、分子生物学、高通量测序相结合的手段，建立堆肥全过程的药物残留及微生物菌群变化监控方法，开展堆肥过程中抗性基因的分类与鉴定、抗生素药物强化降解、抗生素残留及耐药菌及抗性基因响应与变化、微生物群落演替与抗性基因转移、抗性基因削减机制及堆肥产品的安全评估研究，探寻基于药物与抗性基因削减的改良粪便好氧堆肥处理工艺，达到规模化养殖厂粪便安全农用的目的。本课题的研究具有较高的理论意义和应用价值，其研究成果有助于降低环境中抗性基因的扩散，提供安全的畜禽粪便堆肥产品也将不断改善我国土壤环境质量。

规模化畜禽养殖场在饲料中添加了大量的抗生素药物，导致畜禽粪便中抗生素药物和抗性基因（ARGs）的残留。好氧堆肥则是去除和削减畜禽粪便中的抗生素残留物及其抗性基因有效方法。但是畜禽粪便好氧堆肥工艺中耐药菌与抗性基因分布规律与去除机制尚不清晰。因此，本项目研究了含抗生素的猪粪好氧堆肥对抗生素、耐药菌及ARGs的削减等。课题组发表论文10篇，完成研究的指标要求。.以含抗生素残渣好氧堆肥为预实验，建立抗生素和ARGs的检测方法结果显示抗生素可以有效的去除。随后开展关于外源添加生物菌剂、金霉素（CTC）和石灰石对猪粪好氧堆肥过程的影响。结果显示，微生物菌剂的添加降低了堆肥产生的CO2、甲烷、NH3；提高了N2O的释放；提高了NO3--N和NO2--N的含量。石灰的添加缩短了升温时间，同时使高温期的持续时间更长；石灰能加快堆体腐熟；石灰的添加促进了金霉素的降解；CTC对微生物的活性及堆体腐熟有抑制作用；堆肥过程可以有效的去除猪粪中残留的CTC和固化重金属，种子发芽指数也表明堆肥的生物毒性已经降到较低水平。.当粪便中CTC含量在20mg/kg、100 mg/kg时，堆肥可将CTC完全去除；在多重抗生素残留情况下，堆肥过程中温度对这些ARGs携带菌起到了筛选作用，堆肥可以有效削减多数ARGs、重金属Cu和Zn的抗性基因。对猪粪堆肥中可培养耐药菌的抗性研究表明，堆肥对于耐药菌的数量和种类削减非常有效。.本课题的研究成果有助于降低环境中抗性基因的扩散，提供安全的畜禽粪便堆肥产品也将不断改善我国土壤环境质量。