The Study Of Molecular Ecology Of Aerobic Ammonium-oxidizing Bacterium In Sediments Of Changjiang Estuary And Its Adjacent Waters

Nitrogen cycle in material circulation of marine ecosystem is a key process, whichplays an important role in maitaining marine ecological balance. The nitrosificationprocess promoted by aerobic ammonium-oxidizing bacterium (AOB) is therate-limiting step of nitrification. Studying community characteristics of AOB and therelationship between AOB and environmental factors are very significant tounderstand the nitrogen cycle. In this reseach, molecular biology techniques wereused in the community structure and diversity of AOB in the sediments of Changjiangestuary and its adjacent waters. The community diversity of AOB was revealed byestablishing clone libraries for key functional gene amoA, and the communitydistribution characteristics of AOB was also showed by amoA gene copy numbers.The results were summarized as below:1. Through establishing clone libraries for β-AOB amoA gene from station G6and33, multiple sequences alignment and phylogenetic analysis, it was found that theseclone sequences were similar to the ones from several waters around the world in theGenbank. The results showed a higher diversity of β-AOB in the sediment of stationG6offshore closer than station33offshore further. The dominated β-AOB populationof the two stations were obviously different.2. The quantitative real-time PCR (qRT-PCR) is easily affected negatively by theinhibitors especially when complex sedimental samples are analyzed, which will leadto low quantitation accuracy. In this study, two methods of diluting template andadding bovine serum albumin (BSA) were used to optimize the qRT-PCR assay inorder to reduce inhibition when dealing with the sedimental samples. The result showed that the goal of reducing inhibition was achieved with the two methods, butthe latter was better when the accuracy and convenience were taken into account. Theoptimum concentration of BSA was0.1-0.5μg/μL.3. Because the amoA gene copy can represent AOB abundance in a sense, throughdetecting sediments of several cruises in different seasons with qRT-PCR, thetemporal and spatial distribution of amoA gene were analyzed in Changjiang Estuaryand its adjacent waters. The results indicated that β-AOB abundance was the largest insummer, smaller in autumn and smallest in spring. Temperature might be an importanteffecting factor on the growth of β-AOB. β-AOB abundance was the largest near theChangjiang estuary, and gradually decreased along with farther offshore. From thevertical distribution of amoA gene numbers, it could be found that the variation ofβ-AOB abundance was complex in G6, and it might be strongly influenced byChangjiang diluted water.In this study, the distribution and diversity of β-AOB and the information ofenvironmental regulations were obtained from the perspective of the molecularmicrobial ecology, which provided reliable basis for recognizing the biologicaldrivering factors in nitrogen cycle in estuaries and coastal waters, understanding thenitrogen cycle rules and enriching the biogeochemical theory in estuary.