现代非培养技术在反硝化微生物种群结构和功能研究中的应用

Review of modern culture-independent methods used to study community structure and function of denitrifying microorganisms

反硝化过程对于废水生物脱氮工艺的运行、土壤肥分的流失以及N2O的排放均具有重要意义,但参与反硝化过程的微生物种类繁多且多数不可培养,导致对自然环境中反硝化微生物的种群结构及功能的研究具有很大难度.现代非培养分子技术的发展使得对反硝化微生物进行原位、准确、全面的研究成为可能.对反硝化功能基因进行指纹图谱分析、定量PCR或者利用FISH等技术可以有效确定反硝化菌的组成和数量,通过检测反硝化酶和mRNA可将反硝化菌的种群鉴定与代谢活性联系起来,最近新出现的同位素底物标记技术甚至可直接确定反硝化菌的碳源利用情况.重点介绍了上述各种现代非培养技术对反硝化细菌种群结构和功能的研究现状,以期为深入了解反硝化微生物的多样性和功能特性提供参考.

Denitrification is a significant process for biological nitrogen-removal from wastewater, loss of nitrogen fertilizers from soils and emissions of N2O. However, the microorganisms involved in this process are taxonomically diverse and mostly uncultured, which bring considerable difficulties to investigate the community structure and function of denitrifiers in natural environments. The development of modern molecular approaches independent of culture enables the accurate and comprehensive study of denitrifying microbes in situ. Employing fingerprinting analysis, quantitative PCR or fluorescence in situ hybridization （FISH） allows efficient determination of tbe composition and abundance of denitrifiers. Detection of denitrifying enzymes and mRNA can link the community identification and metabolic activity of denitrifiers; and the newly developed isotope substrate labeling technique can even directly identify the denitrifying population of utilizing carbon resources dircetly. Recent researches that apply these techniques to the study of community structure and function of denitrifying bacteria are reviewed, which may help our understanding of the diversity and functional characteristics of denitrifying microbes.