Bacterial Population Characteristics And Nitrogen-degrading Bacteria Screening In Tianjin Coastal Constructed Wetland

The Coastal wetlands have been the last gateway for the control of pollutants entering the sea and the prevention of eutrophication in offshore waters due to its unique geographical location and characteristics of water purification,It also makes the research on population characteristics of bacteria which plays a key role in removing nitrogen in constructed wetlands and denitrification performance of nitrogen-degrading bacteria more practical and useful.Based on it,selecting the Tianjin Lingang coastal constructed wetland as research area,which is located in the shallow sea area on the south side of the Haihe River Estuary,and using high-throughput sequencing technology to explore bacterial population characteristics in the whole and analyze the temporal and spatial differences.The bamo A and nir(nir K and nir S)genes were further used as molecular markers to analyze the population characteristics of nitrifying and denitrifying bacteria.In addition,the dominant nitrogen-degrading strains were enriched,isolated and selected from wetland water in the laboratory,and they were identified by morphological observation,physiological and biochemical index determination,16S r DNA sequence analysis and comparison,and the degradation efficiency of strains on wetland raw water were studied.The main results of the above research are as follows:The bacterial distribution characteristics in overlying water and sediment of reed roots in different seasons were different,and there was higher bacterial diversity and richness in sediments comparing with overlying water in all seasons.In four seasons,Proteobacteria was the first dominant bacteria phyla inoverlying water and sediments,but the first dominant bacteria genera in overlying water were different,namely norank f norank o Chloroplast,norank f Anaerolineaceae,norank f norank o Chloroplast and Flavobacterium,the main environmental factors affecting the distribution of bacterial populations were temperature,concentration of NO₃-N,concentration of NH₄-N and p H.And in sediments,the first dominant bacterial genera in four seasons were norank f Anaerolineaceae,norank f norank o norank c Subgroup 6,norank f norank o norank c Subgroup 6,and norank f A4b respectively,the main environmental factors were concentration of NH₄-N,COD,COD and p H.The main dominant bacterial genera of bamo A-type nitrifying bacteria in sediments were unclassified k norank d Bacteria and Nitrosomonas,the environmental factors that had a significant impact on their richness were temperature and PO₄-P concentration.For the nir K and nir S denitrifying bacteria,the latter had higher value of diversity and abundance both in overlying water and sediments,and there was higher value of diversity in overlying water but higher value of abundance in sediments.In overlying water,the main dominant genera of nir K and nir S denitrifying bacteria were unclassified k norank d Bacteria and unclassified p Proteobacteria,but the value of abundance were different.In sediments,the main dominant genera of nir K denitrifying bacteria were unclassified k norank d Bacteria and unclassified d Unclassified,but the main dominant genera of nir S denitrifying bacteria were unclassified p Proteobacteria and unclassified k norank d Bacteria.The environmental factors in overlying water had greater influence on the population distribution of nir K and nir S denitrifying bacteria and the dominant nir K-type denitrifying bacteria were more closely related to environmental factors.Four dominant nitrogen-degrading strains were screened from overlying water of reed roots and identified as Pseudomonas aeruginosa,Ensifer adhaerens,Serratia marcescens and Bacillus pacificus.Cultivating four strains in raw water from wetlands at 5?,13?,20?,27?and 34?,the growth of bacteria was weak at 5?.Ensifer adhaerens and Bacillus pacificus showed high degradation efficiency for NH₄-N in raw water,which could reach 83.9%and 74.8%in the most optimal temperature of 34?,for Pseudomonas aeruginosa,Ensifer adhaerens and Serratia marcescens,the most optimal temperature for NO₃-N were 27?,but for Bacillus pacificus was 34?,the degradation rates at the optimal temperature were 65.5%,70.2%,67.8%and 55.9%,respectively,and for Pseudomonas aeruginosa,Ensifer adhaerens and Serratia marcescens,the most optimal temperature for TN were27?,but for Bacillus pacificus was 20?,the degradation rates at the optimal temperature were 23.6%,71.7%,34.4%and 89.1%,respectively.