人工湿地净化典型OCPs过程中的植物-微生物耦合降解机制研究

The contamination of organochlorine pesticides (OCPs) in the water body released from agricultural nonpoint sources is not only an urgent environmental problem but also an eco-security problem which relates to the public health. Constructed wetlands (CWs) is considered to be a cheap and effective technology for removing OCPs from water. Our previous study found that the combined purification effect of plant-microorganism for OCPs in wetland is better than that of single plant or microbial treatment. But the coupling mechanism is lack of relevant research and cognition, which play a key knowledge for the potential purification of OCPs in constructed wetland. In this proposal, the community structural characteristics of microbial in different plants of CWs and its degradation ability of typical OCPs will be studied by the molecular ecology methods of PCR-DGGE, Q-PCR, and high throughput sequencing (HTS). The degradation intermediate of typical OCPs in CWs will be identified by using GC-MS analysis. The interaction between plant and rhizosphere microorganisms, the key factors impacting on synergistic purification of β-HCH in system will be identified. The mechanism of the plant-microbial coupling degradation typical OCPs in CWs will be illuminated. The results make a contribution to understand the biodegradation mechanism of OCPs removed from water by CWs. This research will provide scientific evidence for improving the efficiency of OCPs in water using CWs.

农业面源水体有机氯农药（OCPs）污染关系到公众健康的生态安全问题，是亟待解决的环境问题，人工湿地有望成为解决该问题的有效手段。课题组前期研究发现湿地中植物-微生物联合净化OCPs的效果优于单独的植物或微生物处理，但其耦合作用机制缺乏相关研究和认知，这是充分发挥人工湿地OCPs净化潜力的关键。基于前期研究基础，本项目拟运用PCR-DGGE、Q-PCR和高通量测序（HTS）等分子生物学技术探究人工湿地不同植物生境微生物群落结构特征及其对典型OCPs的降解性能；通过GC-MS弄清湿地典型OCPs降解的中间产物；查明水生植物与根际微生物的相互关系，识别驱动其协同净化典型OCPs的关键因子，阐明人工湿地植物与微生物耦合净化水体有机氯农药的降解机理与过程。研究结果将为揭示人工湿地净化OCPs的生物降解机理奠定基础，为提高人工湿地修复OCPs污染水体的效率提供科学依据。

农业面源水体有机氯农药（OCPs）污染关系到公众健康，是亟待解决的环境问题，人工湿地是解决该问题的有效手段之一。本项目构建垂直潜流式人工湿地微宇宙系统，研究人工湿地植物–微生物耦合净化β-HCH的效果、季节性差异和降解路径，识别人工湿地不同植物生境微生物群落特征及其对β-HCH的降解性能，采用GC-MS/MS分析人工湿地β-HCH降解中间产物，基于修复实验查明湿地β-HCH降解的关键因素。项目研究表明人工湿地基质、植物和微生物对水体β-HCH去除贡献率分别约为38%、34%和10%，基质吸附是人工湿地去除β-HCH的主要途径，植物具有促进作用，微生物抑制剂明显影响了基质中β-HCH的去除。人工湿地去除β-HCH无明显季节性差异，美人蕉人工湿地去除β-HCH的性能更稳定。高通量测序发现人工湿地不同植物生境微生物的群落结构和数量存在差异（P<0.05），其中拟杆菌门和放线菌门、芽孢杆菌属和鞘氨醇单孢菌属更具耐受性，可提高人工湿地去除β-HCH的能力。基于富集培养法从稳定处理β-HCH的人工湿地土壤中分离纯化出6株β-HCH降解菌，经鉴定B、F、A1、M1菌株均为Ochrobactrum，J菌株为Pseudomonas，J1菌株为Microbacterium。菌株的降解适宜条件均为pH=7、温度为30℃以及5%接种量，在此条件下，单一菌株和混合菌对10~100 μg/L β-HCH的降解率分别为50.28%~67.38%和24.84%~69.57%；不同混合方式的混合菌可呈现协同和拮抗作用。经GC-MS/MS确认人工湿地β-HCH降解中间产物为四氯环己烯（β-TCCH）等。水培实验研究发现，不同降解菌联合湿地植物对水体中β-HCH降解率在8.9%~99.06%之间，呈现出协同和拮抗等2种不同的耦合效应，且受β-HCH胁迫浓度影响。基于β-HCH降解菌和美人蕉构建的人工湿地对水体β-HCH的去除率可达89.6%，其中微生物降解和植物吸收约占73.2%。外源单一菌和混合菌对湿地土壤β-HCH的降解结果表明，降解菌和根系分泌物具有良好的耦合作用，可明显促进β-HCH的降解；好氧条件、革兰氏阴性菌、生物酶（S-POD、sDHA和S-PPO）可能是湿地土壤中β-HCH降解的关键因素。本研究成果为提高人工湿地修复农业OCPs面源污染水体的效率提供科学依据。