Microorganisms play an important role in removing pollutants from constructed wetlands. We investigated the microbial
characteristics in a novel integrated vertical-flow constructed wetland (IVCW), which has been in operation in Wuhan, China since
1998. We used phospholipid fatty acid (PLFA) and amoA gene to analyze the structure and diversity of the microbial community
within the IVCW. PLFA results suggested that the amount of bacterial PLFA was significantly higher than that of fungal PLFA, but
the total microbial biomass represented by PLFA index was low in the system. Microbial spatial distribution showed significantly
higher bacterial (both G+ and G−) and fungal biomass in the surface than in the subsurface layers. The ratios of monounsaturated to
branched PLFA demonstrated that an anaerobic layer sandwiched by two aerobic layers existed in the IVCW, consistent with the redox
potential results. Analysis of the amoA revealed the presence of Nitrosomonas-like sequences in the surface substrate of the downflow
chamber and apparent diversities of ammonia-oxidizing bacteria in the system. These results suggest that microorganisms, despite their
relatively low biomass, have inhabited the IVCW, and the results will offer some valuable information on microbe to system designers
and managers.