循环水养殖异养硝化-好氧反硝化反应器脱氮性能及微生物群落代谢特性研究

Nitrate accumulation is an important issue in the recirculating aquaculture, as denitrification is thought hardly occurred in aerobic conditions of recirculating aquaculture system. Recently, heterotrophic nitrification-aerobic denitrification bacteria have been thought as a feasible method to solve the problem. The objective of this proposal is to illustrate the microbial community metabolism and nitrogen removal characteristics in heterotrophic nitrification-aerobic denitrification reactor. First of all, heterotrophic nitrification-aerobic denitrification bacterium community were cultured and inoculated to set up a heterotrophic nitrification-aerobic denitrification reactor using bioaugmentation technology. Secondly, we will investigate the effects of carbon source, C/N ratio, temperature, DO and pH on nitrogen removal efficiency in the heterotrophic nitrification-aerobic denitrification reactor. Thirdly, we will figure out the bacteria diversity and microbial community metabolism, and illustrate the relationship between the environmental factors and microbial community structure of the reactor by high-throughput DNA sequencing technology. We hope to provide the scientific and theoretical basis for applying the heterotrophic nitrification-aerobic denitrification technology to the recirculating aquaculture system. The research has great significance for improving the development of the recirculating aquaculture.

硝酸盐积累是海水循环水养殖面临的重要问题，异养硝化—好氧反硝化细菌的发现和应用为有氧条件下去除硝酸盐提供了可能。针对目前国内外海水循环水养殖系统异养硝化—好氧反硝化研究较少的现状，本研究从生态学和微生物学角度对海水异养硝化—好氧反硝化反应器的脱氮性能及其内部微生物群落结构及代谢特性进行探讨。首先采用纯菌构建复合菌群的生物强化技术，将高效的耐盐异养硝化—好氧反硝化细菌构建复合菌群，接种建立异养硝化—好氧反硝化反应器，实现有氧条件下硝酸盐的去除；其次探讨不同碳源、碳氮比、温度、溶解氧、pH等环境因子对异养硝化—好氧反硝化反应器脱氮性能的影响机制；最后采用分子生物学技术揭示海水异养硝化—好氧反硝化反应器中的微生物多样性，阐明反应器内微生物群落结构与环境因子之间的关系，明确反应器内微生物群落代谢特性。本研究结果将为异养硝化—好氧反硝化技术在循环水养殖中的应用提供科学依据和理论基础。

循环水养殖具有环境可控、高效、环保和产品优质安全等优点，是当前国际上先进养殖模式的代表，其核心环节生物过滤依靠硝化细菌的硝化作用将对养殖物有害的氨氮转化为毒性较小的硝酸盐，而硝酸盐浓度积累过高对养殖鱼类具有潜在的胁迫和毒性，成为海水循环水养殖面临的重要问题。本项目首先采用复合菌群的生物强化技术，将高效的耐盐异养硝化—好氧反硝化细菌构建复合菌群，接种建立异养硝化—好氧反硝化反应器，其次探讨不同环境因子对反应器脱氮性能，构建反硝化动力学特性模型，最后采用分子生物学技术揭示海水异养硝化—好氧反硝化反应器中的微生物多样性及碳源代谢特性，明确反应器内微生物群落功能。研究结果显示将3种反硝化细菌接种构建的反应器具有良好的脱氮性能，好氧反硝化反应器接种后挂膜成熟需要15d；当C/N大于10，温度20-35℃，pH为7，盐度2-5mg/L，在有氧条件下反应器的硝酸盐去除率均达到90%以上；Monod方程的动态模型能很好的拟合该好氧反硝化的动力学过程，相应的参数分别为：V_(max,"NO3" )为25.32 mg/(L•h), K_s",NO3" 为0.11mg/L, V_(max,"NO2" )为21.26 mg/(L•h), K_s",NO2" 为0.04 mg/L；反应器内微生物群落隶属于19个主要的科，发现接种盐单胞菌Halomonas venusta是优势种，说明盐单胞菌在这个反硝化过程中起主要作用；采用Biolog-ECO板来分析好氧反硝化菌的群落代谢特征发现进水硝酸盐浓度保持不变，HRT越长，微生物代谢活性越强，反应器内微生物对碳源的代谢利用强弱顺序是：多聚物＞氨基类＞碳水化合物＞羧酸类＞胺类＞酚酸类。本项目研究结果为异养硝化—好氧反硝化技术在循环水养殖中的应用，解决循环水养殖系统氮的积累以及海水养殖污水排放问题提供有力的科学依据和数据基础。