THE STUDY OF NATURAL BIOFILM FORMATION FOR NITROGEN REMOVAL FROM AQUACULTURE WASTEWATER AND ANALYSIS ON MICROBIAL COMMUNITY IN BIOFILM
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摘要: 以弹性填料和流化床填料为硝化反应的生物挂膜材料, 聚羟基丁酸/戊酸共聚酯(PHBV)为反硝化反应的碳源和生物膜载体, 通过微生物自然挂膜处理低C/N比水产养殖废水, 去除水体中的氨氮、亚硝酸盐氮及总氮。应用Miseq高通量测序技术对生物膜的微生物群落组成和结构进行分析。结果表明: 温度25—30℃, 该处理系统首次挂膜成功需要4周, 启动后运行稳定, 对2种不同来源和氮污染程度的养殖废水均有较好的脱氮效果, 氨氮、亚硝酸盐氮及总氮的去除率均在90%以上。硝化生物膜(a)的优势菌分别归属变形菌门(Proteobacteria)、拟杆菌门(Bacteroidetes)和厚壁菌门(Firmicutes)。反硝化生物膜(b)微生物群落的多样性指数和丰度指数均远大于前者, 主要为变形菌门、厚壁菌门、拟杆菌门、螺旋体门(Spirochaetae)及绿菌门(Chlorobi)。其中, 归属于变形菌门β-变形菌纲(Betaproteobacteria)的丛毛单胞菌科(Comamonadaceae)和红环菌科(Rhodocyclaceae)在2种生物膜中占比均较高。由于所处环境(载体, 碳源、溶氧等)不同, 在属分类水平上, 2种生物膜的细菌群落结构表现出明显差异。生物膜a中属的种类仅为b的三分之二, 相对丰度>0.5%的优势菌属, a为8个, b为18个。其中, 隶属丛毛单胞菌科和红环菌科未知属的优势种群分别占到a、b总序列数的56.67%和45.51%。磁螺菌属(Magnetospirillum)和硝化螺菌属(Nitrospira)是a中特有的优势功能菌群, 梭菌属(Clostridium)、动胶菌属(Zoogloea)、管道杆菌属(Cloacibacterium)、脱硫弧菌属(Desulfovibrio)等具有反硝化功能的菌群为b的优势菌属。Abstract: This study was conducted to evaluate the applications of biological filter medias in low C/N ratio aquaculture wastewater. Elastic filler and fluidized bed packing materials were used as biofilm of nitration and PHBV (poly-3-hydroxybutyrate-co-3-hydroxyvalerate) were used as the carbon source and biofilm carrier of denitrification in this study. The water quality and the bacterial community compositions were determined by the method of Miseq high-throughput sequencing. The obtained results demonstrated that biofilm of the system formed after 4 weeks operation, at the temperature of 25—30℃. Our results also showed that the two kinds of media had great effects on removing total nitrogen (TN), ammonia nitrogen (NH3-N), nitrite (NO2-N). The removal rate of TN, NH3-N and NO2-N can reach up 90%. The sequencing results indicated that the predominant bacteria on the biofilm sample a coating onto elastic filler and fluidized bed packing materials were Proteobacteria, Bacteroidetes and Firmicutes. While the major microorganisms on the biofilm sample b attaching to the surface of PHBV granules were Proteobacteria, Firmicutes, Bacteroidetes, Spirochaetae and Chlorobi. The analysis revealed that both diversity index and abundance index of microbial community of biofilm sample b were far higher than the sample. Family of Comamonadaceae and Rhodocyclaceae belonging to Betaproteobacteria were highly enriched in both biofilms samples. However, there was a distinctive difference in phylotypes between the two biofilm samples at genus level, which was probably due to the diversities of biofilm carrier, carbon source and dissolved oxygen. The genera of biofilm sample a was only two thirds of b. A total of 8 and 18 genera with a relative abundance greater than 0.5% were identified from a and b, respectively. Among them, the unknown groups belonging to Comamonadaceae and Rhodocyclaceae accounted for 56.67% and 45.51% of the total sequence of a and b. Organisms from the genus Magnetospirillum and Nitrospira were only detected in a, while the genus Clostridium, Zoogloea, Cloacibacterium, Desulfovibro etc were identified in b, which probably contributed to denitrification performance.
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Keywords:
- Biofilm carrier /
- PHBV /
- Aquaculture wastewater /
- Biofilm /
- Microbial community
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图 1 本试验中使用的挂膜材料(a1-弹性填料, a2-流化床填料, b-PHBV)
Figure 1. The biofilm carriers used in this experiment (a1-elastic filler, a2-fluidized bed packing materials, b-PHBV)
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图 3 自然微生物挂膜阶段养殖废水NH4-N、NO2-N、TN及pH变化
Figure 3. Changes of aquaculture wastewater NH4-N, NO2-N, TN and pH during cultivation of biofilm
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图 4 甲鱼养殖废水NH4-N、NO2-N、TN及pH变化
Figure 4. Changes of Chinese turtle aquaculture effluent NH4-N, NO2-N, TN and pH
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图 5 南美白对虾养殖废水NH4-N、NO2-N、TN及pH变化
Figure 5. Changes of Penaeus vannmei aquaculture wastewater NH4-N, NO2-N, TN and pH
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图 8 生物膜样品a中主要细菌分类组成(属水平)
Figure 8. Bacterial communities of biofilm sample a at genus level
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图 9 生物膜样品b中主要细菌分类组成(属水平)
Figure 9. Bacterial communities of biofilm sample b at genus level
表 1 生物膜样品中细菌的丰度及多样性指数
Table 1 Richness and diversity indexes of bio-film samples (α=0.03)
样品编号
Sample ID序数
Number
sampled丰度指数
Abundance1
index多样性指数
Shannon
index测序深度
Coverage
indexa 25327 155 2.40 0.999 b 25327 216 3.24 0.999 
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