| Healthy aquaculture is the key factor in the ecological environment situation in aquaculture water. The diversity of the microbial community structure and the physicochemical factors play important roles in the healthy aquaculture. Nowadays, the accumulation of nitrogen is harmful to aquatic organisms in aquaculture ponds. Microorganisms play a crucial role in nitrogen circulation, the study of pond microbial abundance and structure is important to adjust the nitrogen balance. In this study, the abundance and diversity of bacteria in two types of ponds were investigated by quantitative PCR and 16S rRNA-based Illumina MiSeq Sequencing. In the meantime, the seasonal variation of abundance of ammonia-oxidizing bacteria (AOB) was investigated. The relationship between microbial community structure and environmental factors, the abundance of AOB and environmental factors was studied respectively by using relevant statistical methods, which provide basic reference to control pond water quality. The main results are as follows:1. The comparison between the two types of feeds in the bacterial abundance and community structure(1) There were significant differences in the concentration of ammonia and nitrite between the two types of ponds, and no significant difference was observed in nitrate. The concentration of ammonia in the bottom layer of green fodder ponds (0.83±0.18 mg/L) was significantly lower than the bottom layer of commercial feed ponds (1.51±0.99 mg/L). The concentration of nitrite in the surface (0.06±0.05 mg/L) and bottom (0.06±0.05 mg/L) layer of green fodder ponds were significantly lower than the surface (0.13±0.08 mg/L) and bottom (0.13±0.09 mg/L) layer of commercial feed ponds (1.51±0.99 mg/L), respectively. There were no significant differences in the concentration of ammonia, nitrate and nitrite between the surface and bottom layer, respectively.(2) Results showed the abundance of bacterial 16S rRN A genes was ranged from (2.81±0.64)×109～(8.63±1.01)×1010 copies/mL. There was significant difference in the abundance of bacterial 16S rRNA genes between the two types of ponds.The abundance of bacterial 16S rRNA genes in green fodder ponds was significantly lower than that in commercial feed ponds in September. There was significant difference in the abundance of bacterial 16S rRNA genes between the surface and bottom layer. In green fodder ponds, the abundance of bacterial 16S rRNA gene was significantly higher in the surface water layer than in the bottom layer in August. Similar results were obtained for commercial feed ponds.(3) The microorganism in the ponds supplied with green fodder found to have a more diverse microbial community. The dominant phyla in pond water were Proteobacteria (24.28%-48.22%), Cyanobacteria (5.9%-37.61%), Bacteroidetes (5.4%-34.82%) and Acidobacteria (8.2%-22.37%). There were no significant differences in the four dominant phyla between the two types of ponds, but there were still some differences between them. Proteobacteria and Actinobacteria were more abundantly found in green fodder ponds, while Bacteroidetes were more abundantly found in commercial feed ponds. At genus level, Comamonadaceae unclassified were significantly enriched in green fodder ponds compared to commercial feed ponds; while Bacillus, Acinetobacter and Aeromonas were significantly enriched in commercial feed ponds compared to green fodder ponds.(4) Similarity analysis and cluster analysis results showed that the water samples from different types of feeds were different, while the samples from same types of pond were fairly similar, which suggested the microbial communities were affected by the different types of feeds. It was shown that the pond microbial community was featured by strong temporal shifts and seasonal clustering. The water samples from the same sampling month were fairly similar, while the samples from different months were different.(5) The relationship between the microbial community structure and environmental factors was analyzed by using Canonical Correlation Analysis (CCA). Dissolved oxygen, temperature, and total organic carbon were the most environmental factors explaining the water community composition, were positively correlated with Axis 1.Axis 2 had a positively correlation with temperature nitrite, and nitrate, but negatively correlated with ammonia.2. The effect of different feeds on nitrogen cycling bacteria in pond waterN-functional bacteria and nitrogen removal characteristics were analyzed in two types of ponds. Our physicochemical demonstrated that significantly lower NH4+-N, NO2--N, and N03--N concentrations were observed in the D ponds (with grass carp fed sudan grass) compared with those in E ponds (with grass carp fed commercial feed). Differences in N cycle groups among the two types of ponds water samples were identified with Illumina MiSeq Sequencing. The result revealed nitrite-oxidizing bacteria (Nitrospira), ammonia-oxidizing bacteria (Nitrosomonas), all sequences related to heterotrophic denitrification bacteria, all sequences related to anammox bacteria, and autotrophic denitrification bacteria (genera Paracoccus and Thiobacillus) had much higher abundances in the E ponds, whereas more sequences related to autotrophic denitrification bacteria (genera Dokdonella and Lysobacter) were detected in the D ponds. These results suggested the nitrogen cycle might be changed by supplying with sudan grass to the aquaculture ponds. And it could effectively to improve the pond water quality by supplying with sudan grass instead of commercial feed to the ponds.3. Correlation analysis between the seasonal variations of abundance of ammonia-oxidizing bacteria and environmental factors in aquaculture water columnTo investigate the seasonal variations of abundance of ammonia-oxidizing bacteria (AOB) and the relationship between the abundance of AOB and environmental factors in different water columns by using the functional gene amoA as a molecular marker, we investigated grass carp ponds in Chonghu fishing ground, Gong’an County, Hubei province. Results showed that the different water columns found the similar results, significantly higher copy numbers of the AOB amoA gene were observed in summer, while no significant differences were detected in spring and autumn. There was no significant difference between the AOB abundance at the surface and bottom layer in the same season. The AOB abundance at the surface and bottom layer were positively correlated with temperature and total phosphorus, but negatively correlated with DO. Furthermore, the surface AOB abundance was significant negatively correlated with total organic carbon. The principal component analyses (PCA) showed that the eight environmental factors were grouped into three PC As. At the surface layer, PCA1 included temperature, DO, ammonia nitrogen and total phosphorus, PCA2 included nitrate nitrogen and total organic carbon, PCA3 included total nitrogen and nitrite nitrogen. At the bottom layer, PCA1 included temperature, DO, ammonia nitrogen, total phosphorus and total organic carbon, PCA2 included nitrate nitrogen and nitrite nitrogen, PCA3 included total nitrogen. |
PDF Full Text Request