Distribution Characteristics Of ANAMMOX Bacteria And DAMO Bacteria In Integrated Wetland Aquaculture System

Anaerobic ammonium oxidizing( anammox) and denitrification dependent anaerobic methane oxidation(DAMO)are two kinds of important anaerobic denitrifying bacteria, the required carbon source are respectively for CO2 and CH4 which are two categories of greenhouse gases. ANAMMOX bacterias are belong to planctomycetes that is deep branch in the evolutionary tree, and DAMO bacterias are belong to NC10. The application of anammox bacteria and DAMO bacteria can not only solve the problem of high nitrogen load and nitrate accumulation in the traditional pond culture system, but also can consume the greenhouse gas and protect the environment. At present, there is a lack of research on anaerobic denitrification in recirculating aquaculture system consist of composite wetland. The research to systematically understand and master the anaerobic bacteria species, groups, distribution, and related environmental factors, however, can provide recirculating aquaculture system consist of composite wetland with the theory of artificial regulation of micro ecological environment. Besides, more efficient ecological aquaculture systems can be targeted designed and optimized. With builded 16 S r RNA clone library and typical correspondence analysis between physical and chemical factor and community structure, we study key environment factors and community structure and distribution characteristics of anammox bacteria and DAMO bacteria in recirculating aquaculture system consist of composite wetland which include pond(P), ecology ditch(ED), ecology pond(EP), subsurface flow constructed wetland(SSFCW).With high throughput sequencing, we study the abundances of Planctomycetes and NC10 among the microbial communities in recirculating aquaculture system consist of composite wetland. As a result, the dominant species of anammox bacteria and DAMO bacteria in this system were identified. the distribution regularities of anammox bacteria and DAMO bacteria in this system and key environmental factors were realized during one year. the abundances of Planctomycetes and NC10 among the microbial communities in this system were verified. Meanwhile, spatial distribution pattern of the abundances of anammox bacteria in SSFCW. The main research results are as follows:1) The dominant species of anammox bacteria and DAMO bacteria in recirculating aquaculture system consist of composite wetland were identifiedWith 16 S r RNA clone library technology, two kinds of known bacteria which are brocadia and kuenenia and most unknown anammox bacteria were detected in recirculating aquaculture system consist of composite wetland, showed that anammox bacteria are widely distributed in this system. Brocadia is a dominant anammox bacteria for this system.By 16 S r RNA clone library technology, many of specific fragment of DAMO bacteria were amplified. Part of OTU of DAMO bacteria divided by 3% differences are belong to Group B, while most of it have a far distance in the evolutionary tree with Group A and Group B. None of OTU highly similar to Methylomirabilis oxyfera was found.2) Community structure characteristics of anammox bacteria and DAMO bacteria in recirculating aquaculture system consist of composite wetland were exploredThe results showed that brocadia was existed in four seasons in recirculating aquaculture system consist of composite wetland, but kuenenia was only existed in summer and winter in this system. RDA and CCA analysis showed that DO, NO2--N and NH4+-N of overlying water,TOC of sediment are key environment factors to affect community structure of anammox bacteria in this system.The numbers of OTU kinds of DAMO in this system were most in autumn and were least in spring and in winter. RDA and CCA analysis showed that T, ORP, NO2--N, NH4+-N, NO3--N of overlying water are key environment factors to affect community structure of DAMO bacteria in this system.3) Diversity characteristics of anammox bacteria and DAMO bacteria in recirculating aquaculture system consist of composite wetland were exploredGenerally, the diversity of anammox bacteria in EP is higher than that of other three types wetland, and the diversity of anammox bacteria in SSFCW is higher than that of ED and P. In addition, diversity order of each function unit in recirculating aquaculture system consist of composite wetland is not the same.Compared with natural ecosystem, the diversity of DAMO in recirculating aquaculture system consist of composite wetland is higher than it. compared with other types wetlands in this system, the diversity of DAMO in SSFCW is higher.4) Spatial distribution patterns of the abundances of anammox bacteria in SSFCW were exploredThere is a great difference in anundance of ANAMMOX bacterium in SSFCW during different seasons. The anundance of outlet at lower layer in Mar. during spring was the lowest, while in Jun. during summer was the highest, and the difference is more than 102 times. The anundance of anammox bacterium in Sep during autumn was that the part of upper layer and lower layer of inlet was lower than the outlet at upper and lower layer, and the anundance of inlet at medium layer was significantly higher than outlet at medium. In the vertical direction of SSFCW, the anundance of anammox bacterium in the medium layer of wetland were higher than the upper and lower layer part both in Mar during spring and in Sep during autumn, while the result of anundance in Jun during summer was lower layer > medium layer > upper layer. Thus, there was an obvious spatial difference in the anundance of ANAMMOX bacterium in SSFCW, including horizontal distribution and vertical distribution difference, but no seasonal change rule.5) The abundances of Planctomycetes and NC10 among the microbial communities in this system were verifiedPlanctomycetes content of 9P,9ED,9EP,9SSFCW is 0.368%,0.146%,0.137%,0.696%, respectively. NC10 content of 9P,9ED,9EP,9SSFCW is 0.027%,0.031%,0.028%,0.050%, respectively. The bacterial abundance of NC10 was lower one order of magnitude than that of the Planctomycetes.6) Structure and diversity of microbial community in recirculating aquaculture system consist of composite wetland were exploredThe main bacteria in recirculating aquaculture system consist of composite wetland were Proteobacteria(41.93%), Firmicutes(12.09%), Acidobacteria(7.63%), Bacteroidetes(5.38%). Gemmatimonadetes(4.14%), Chloroflexi(3.91%), Nitrospirae(3.00%). The dominant bacteria are Proteobacteria and Firmicutes. Among them, Proteobacteria, Bacteroidetes and Chloroflexi are widely exist in four types wetland. In this study, we found that Betaproteobacteria-Rhodocyclales- Rhodocyclaceae is a OTU type that contains the largest number of sequences.In microbial diversity level, 9SSFCW is significantly different than other three types wetlands.