Studies On Anammox Bacterial Abundance、Community Structure And The Influence Of Environmental Factors In Pearl Estuary

Anaerobic ammonium-oxidizing (anammox) process plays an important role inmarine nitrogen cycle, anammox bacteria can generate N2through NH4+oxidationcoupled with NO2reduction under anoxic condition and they may be responsible for30%to70%of the total N2production in the ocean. Pearl Estuary, a typicalsubtropical estuary, has complex environmental conditions due to the intense mixingof freshwater with seawater and the strong influence of anthropogenic activities.Excessive nutrients and the environment of hypoxia caused by eutrophication is themost typical environmental characteristics of Pearl Estuary. As the ideal ecosystemconditions of researching various kinds of microorganisms, this area has receivedextensive attentions of various subjects during decades. Rich nutrients and thehypoxia environmental conditions provided suitable conditions for anammox bacterialgrowth. The aim of this paper is to reveal the diversity, distribution and abundance ofanammox bacteria through investigating the community structure, spatial distributionand abundance of anammox bacteria. Additionally, we aim to evaluate the influenceof various environmental factors on anammox bacteria through the correlationanalysis between anammox bacterial community structure, abundance andenvironmental factors.The sediment and water samples of this research were collected by voyage ofPearl Estuary in summer during16July to12August,2012. Totally nine sites weretaken along the salinity gradient from freshwater to the open ocean. Sampling sitesincluding Three typical aquatic environment: freshwater (P1, P3, P7), ecotonesbetween freshwater and seawater(A4, A8, C2, C3) and seawater(F412, F414). Surfacesediment, surface and bottom water were collected.Anammox bacterial16S rRNA gene and hydrazine oxidoreductase (HZO) gene clone libraries were constructed and482and455nucleic acid sequences wereobtained respectively. The455hzo gene sequences were translated into amino acidsequences. The phylogenetic analysis were performed based on16S rRNA genesequences and HZO amino acid sequences of anammox bacteria. The results ofnucleic acid sequences and amino acid sequences phylogenetic analysis indicated theobvious community structure shift that anammox bacterial dominant genus in surfacesediment changed from “Candidatus Brocadia” or “Candidatus Anammoxoglobus” to“Candidatus Scalindua” along the salinity gradient from freshwater to the open ocean.This result indicated the evident geographical distribution of anammox bacteria inPearl Estuary. The correlation analysis between anammox bacterial communitystructure in surface sediment and sedimentary environmental factors indicated thesalinity, temperature, C/N and pH were the major factors contributing to differentdistribution of anammox bacterial community structure in surface sediment. Theresults of phylogenetic analysis also revealed the rich diversity of anammox bacteriain Pearl Estuary including four novel clusters.In addition, abundance of total bacteria and anammox bacteria in both water andsurface sediment of Pearl Estuary were investigated by quantitative PCR targeting thetotal bacterial and anammox bacterial16S rRNA gene. In sediment, bottom water andsurface water, the copy numbers of total bacterial16S rRNA gene ranged from2.66×1011to6.68×1011copies/g,8.64×1010to5.45×1011copies/L,1.18×1011to4.73×1011copies/L, respectively. The copy numbers of anammox bacteria in surfacesediment, bottom water and surface water ranged from4.22×105to2.55×106copies/g,1.24×104to1.01×105copies/L and8.07×103to8.86×105copies/L respectively. Thecorrelation analysis between anammox bacterial abundance and environmental factorsrevealed that the abundance of total bacteria16S rRNA gene in both sediment andwater samples did not show significant correlations with any environmental factorsmeasured except the bottom water that showed positive correlation with NO2-concentration. Additionally, the abundance of anammox bacterial16S rRNA gene inwater samples were positively correlated with NO2-and negatively correlated withDO. The results indicated the rich diversity, high abundance and new assemblages ofanammox bacteria in Pearl Estuary. Additionally, environmental factors contributedsignificantly to anammox bacterial spatial distribution and abundance. This study willbe of great significance to the knowledge of anammox bacteria in estuarineecosystem.