鲫鱼养殖池塘底泥菌群结构及常见可培养菌分析
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摘要
为了研究江苏地区鲫鱼养殖池塘底泥细菌菌群结构与多样性,通过高通量测序技术对2017年8～10月间采集的江苏省不同区域养殖池塘底泥中所有细菌的16S rRNA V3-V4基因进行扩增测定,分析了底泥菌群结构特征;并通过平板计数法和选择培养基法检测池塘底泥中细菌总数和主要常见菌。结果表明,49个样品高通量测序共获得有效序列2 152 956条,可归为17 735个分类操作单元(OTUs);物种注释结果显示,鲫鱼养殖池塘底泥中具有较高的菌群多样性,检测到的细菌归属于29个门类;所有池塘底泥都具有主要门类为变形菌门、绿弯菌门、厚壁菌门、拟杆菌门、水生古细菌门、酸杆菌门、放线菌门、硝化螺旋菌门、螺旋菌门、芽单胞菌门、蓝细菌门、浮霉菌门、疣微菌门等。在属的水平上主要以硫杆菌属、乳酸球菌属、硫氧化菌、甲烷菌、地杆菌属、假单胞菌属、厌氧粘杆菌属、甲烷绳菌属、芽孢杆菌属等为主;所有池塘底泥共有菌主要有乳酸球菌属、甲烷菌、盐单胞菌、芽孢杆菌、红杆菌属、链球菌属、肉杆菌属、微小杆菌属、土芽孢杆菌属等。通过细菌培养发现,池塘底泥中常见益生菌如乳酸菌、芽孢菌具有较高的比例。可见鲫鱼养殖池塘底泥菌群结构及组成具有多样性,不同养殖池塘中共有的细菌可作为益生菌开发的参考菌株。
In order to evaluate the bacterial community and structure of carp cultured pond's sediment,highthroughput sequencing of 16 S rRNA V3-V4 gene technique was used to analyzes all 49 sediment samples collected from August-October 2017 collected from different regions in Jiangsu Province. Total bacteria and common bacteria in sediments were also detected using selecting medium. The results show that 2 152 956 valid sequences and17 735 OTUs were obtained from the samples. Phylogenetic analysis showed that Proteobacteria was the predominant phylum in all samples,followed by Chloroflexi,Firmicutes,Bacteroidetes,Euryarchaeota,Acidobacteria,Actinobacteria,Nitrospirae,Spirochaetae,Gemmatimonadetes,Cyanobacteria,Planctomycetes and Verrucomicrobia. Thiobacillus,Lactococcus,Sulfurovum,Methanosaeta,Geobacter,Pseudomonas,Anaeromyxobacter,Methanolinea,Bacillus were the dominant microbial genus. The genus of Lactococcus,Methanosaeta,Halomonas,Bacillus,Rhodobacter,Streptococcus,Carnobacterium,Exiguobacterium,Geobacillus were shared in all sediments samples.With the bacterial culture method,Bacillus sp. and Lactobacillus sp. presented higher abundances in the sediments that Aeromonas sp. and Vibrio sp. It is concluded that the bacterial communities of carp culture pond sediments are diversity and the common bacteria in all the ponds can be used as autochthonous strains for the probiotics in aquaculture.
In order to evaluate the bacterial community and structure of carp cultured pond's sediment,highthroughput sequencing of 16 S rRNA V3-V4 gene technique was used to analyzes all 49 sediment samples collected from August-October 2017 collected from different regions in Jiangsu Province. Total bacteria and common bacteria in sediments were also detected using selecting medium. The results show that 2 152 956 valid sequences and17 735 OTUs were obtained from the samples. Phylogenetic analysis showed that Proteobacteria was the predominant phylum in all samples,followed by Chloroflexi,Firmicutes,Bacteroidetes,Euryarchaeota,Acidobacteria,Actinobacteria,Nitrospirae,Spirochaetae,Gemmatimonadetes,Cyanobacteria,Planctomycetes and Verrucomicrobia. Thiobacillus,Lactococcus,Sulfurovum,Methanosaeta,Geobacter,Pseudomonas,Anaeromyxobacter,Methanolinea,Bacillus were the dominant microbial genus. The genus of Lactococcus,Methanosaeta,Halomonas,Bacillus,Rhodobacter,Streptococcus,Carnobacterium,Exiguobacterium,Geobacillus were shared in all sediments samples.With the bacterial culture method,Bacillus sp. and Lactobacillus sp. presented higher abundances in the sediments that Aeromonas sp. and Vibrio sp. It is concluded that the bacterial communities of carp culture pond sediments are diversity and the common bacteria in all the ponds can be used as autochthonous strains for the probiotics in aquaculture.
引文
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2 黄德.水产养殖对环境的影响及其防治对策分析[J].黑龙江水产,2016(4):11-13Huang De. The influences of aquaculture on environment and the prevention strategy analysis[J]. Heilongjiang Fisheries,2016(4):11-13
3 Heuer H,Smalla K. Application of denaturing gradient gel electrophoresis(DGGE)and temperature gradient gel electrophoresis(TGGE)for studying soil microbial communities[M]//van Elsas J D,Trevors J T,Wellington E M H. Modern Soil Microbiology. New York:Marcel Dekker,Inc. 1997:353-373
4 Caporaso J G,Kuczynski J,Stombaugh J,et al. QIIME allows analysis of high-throughput community sequencing data[J]. Nature Methods,2010(7):335-336
5 Ozsolak F,Milos P M. RNA sequencing:Advances,challenges and opportunities[J]. Nature Reviews Genetics,2011(12):87-98
6 王兴春,杨致荣,王敏,等.高通量测序技术及其应用[J].中国生物工程杂志,2012,32(1):109-114Wang Xingchun,Yang Zhirong,Wang Min,et al. High-throughput sequencing technology and its application[J]. China Biotechnology,2012,32(1):109-114
7 田李,张颖,赵云峰.新一代测序技术的发展和应用[J].生物技术通报,2015,31(11):1-8Tian Li,Zhang Ying,Zhao Yunfeng. The next generation sequencing technology and its applications[J]. Biotechnology Bulletin,2015,31(11):1-8
8 Sandle T. Microbiological culture media:A complete guide for pharmaceutical and healthcare manufacturers[M]. River Grove:DHI Publishing,2018:405-406
9 李晓,李冰,董玉峰,等.精养团头鲂池塘沉积物微生物群落的结构特征及组成多样性分析[J].水产学报,2014,38(2):218-227Li Xiao,Li Bing,Dong Yufeng,et al. Analysis of sediment microbial communities in Megalobrama amblycephala intensive rearing pond[J]. Journal of Fisheries of China,2014,38(2):218-227
10 史丽娜,可小丽,刘志刚,等.罗非鱼-鱼腥草共生养殖池塘沉积物菌群结构与功能特征[J].中国农学通报,2015,31(14):64-73Shi Lina,Ke Xiaoli,Liu Zhigang,et al. Effect of Houttuynia cordata floating-bed on the structure and function of bacterial community in the sediments of Tilapia aquacultural system[J]. Chinese Agricultural Science Bulletin,2015,31(14):64-73
11 Hai N V. The use of probiotics in aquaculture[J]. Journal of Applied Microbiology,2015,119(4):917-935