典型抗生素菌渣肥引发土壤细菌耐药的剂量-效应关系及耐药菌环境行为研究

Antibiotic mycelial residues(AMRs) are Hazardous wastes in China, though they are rich in nutrients, trace amounts of antibiotics and metabolites residue they contained also lead tobacterial resistance and pose a threat to environment.Antibiotics with many different kinds have been widely used, but until to now ,we still lack of systematic research for thedose effect relationship between AMRs antibiotic residue and bacterial resistanceat home and abroad. so far thepollution control standards for antibiotic residue in AMRs do not exist in China or other countries, and the problem of AMRs harmless treatment and resource utilization can not be solved effectively.Based on the characteristics of AMRs (containing lots ofprotein, carbohydrates, amino acids and other nutrients) and previous research, the project intends to use high temperature treatment, combination of aerobic fermentation and high temperature treatment, microwave and other treatment means to remove the residues of antibiotics and its degradation products in AMRs, then to processed the organic fertilizer using the AMRs of penicillin ,cephalosporin, erythromycin and other typical antibiotics. Use the simulated planting experiment to study the effect of AMRs organic fertilizer on soil texture, soil microbial diversity and soil enzyme activity. Apply primers selection and design, PCR-DGGE and qPCR technologies to identify and quantify the resistance genes in resistant bacterium then build the dose effect relationship between antibiotic residue and bacterium resistance. Analysis the effects and mechanisms of heavy metals in soil to thebacterium resistance caused by antibiotics residue and discuss the environmental behavior of resistant bacterium in soil.The implementation of the project will have both theoretical significance and practical valueto establish AMRs pollution control standards and promote China pharmaceutical enterprise to develop healthily.

抗生素菌渣虽然富含大量营养物质，但因其含有微量抗生素及其降解产物残留，具有引发细菌耐药的环境风险，在我国属于危险废物。.基于抗生素菌渣富含大量营养物质这一特点，本项目在前期研究的基础上，拟采用高温、好氧发酵+高温、微波等处理手段去除菌渣中抗生素及其降解产物残留，构建青霉素、头孢菌素、红霉素等典型抗生素菌渣有机肥制备工艺。通过菌渣有机肥土壤模拟试验，深入研究菌渣肥对土壤土质、土壤微生物活性等影响。通过引物筛选与设计、并借助多聚酶链式反应-变性梯度凝胶电泳（PCR-DGGE）和实时荧光定量（qPCR）技术对抗生素残留所引发耐药菌的耐药基因进行鉴别和定量分析，建立抗生素残留与引发细菌耐药的剂量-效应关系。深入分析土壤中重金属对抗生素引发细菌耐药的影响与作用机制，探讨土壤中耐药菌的环境行为，以期解决目前我国抗生素菌渣有机肥利用存在的安全性问题，并为我国抗生素菌渣污染控标准标的制定提供依据。

我国是抗生素原料药的生产和出口大国，其生产过程会产生大量菌渣。抗生素菌渣如若不能妥善处理，其中的抗生素残留会通过引发细菌耐药而产生潜在的环境风险和人类健康风险，也严重制约我国抗生素制药行业的健康发展。.本研究基于抗生素理化性质差异，构建了水解法青霉素G菌渣、微波法头孢菌素C菌渣、高温水热法红霉素菌渣和高温水热法林可霉素菌渣中抗生素高效去除与肥料化生产工艺；通过实验室土壤施肥模拟试验，解析了四种抗生素菌渣肥的施入对土壤酶活、土壤生物群落结构、作物生长的影响规律，证实了其可有效提高土壤的肥力；利用高通量实时荧光定量PCR对菌渣肥和施肥后土壤进行抗性基因的检测，解析了残留抗生素剂量与土壤耐药基因及微生物群落多样性的关系，从分子生物学角度阐明了菌渣肥的使用安全性。