不同养殖模式下罗氏沼虾肠道菌群结构特征及其与环境因子的关系
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摘要
通过高通量测序技术分析不同养殖模式对罗氏沼虾(Macrobrachium rosenbergii)肠道菌群结构的影响,冗余分析(RDA)肠道菌群与水环境因子的关系。实验设置6种养殖模式:罗氏沼虾单养(MP组)、罗氏沼虾+浮萍(Lemna minor)(PP组)、罗氏沼虾+鲢(Hypophthalmichthys molitrix)(PF组)、罗氏沼虾+背角无齿蚌(Anodonta woodiana)+鲢(PMF组)、罗氏沼虾+背角无齿蚌+浮萍(PMP组)、罗氏沼虾+背角无齿蚌+浮萍+鲢(PMPF组)。养殖64 d,测定水环境因子(浊度、DO、pH、Chl.a、COD、BOD、NO_3-N、NO_2-N、NH_3-N、TN、TP、PO_4-P、TOC)及肠道菌群结构。结果表明:罗氏沼虾不同养殖模式对水体浊度、PO_4-P、TN和Chl.a具有显著影响(P<0.05),MP组PO_4-P浓度最大且显著大于其他组(P<0.05)。肠道细菌多样性指数MP组最大(4.08),最低为PMF组(1.27)。变形菌门(Proteobacteria)、软壁菌门(Tenericute)和厚壁菌门(Firmicutes)在虾肠道中相对丰度较大。不同养殖模式最大优势菌存在较大差异,其中MP、PMP和PMPF组为气单胞菌属(Aeromonas)、PP组为柠檬酸杆菌属(Citrobacter)、PF和PMF组为Candidatus Hepatoplasma。肠道细菌与环境因子相关性分析表明:TP对罗氏沼虾肠道菌群具有显著影响(P<0.05);肠棕气单胞菌(Aeromonas enteropelogenes)、有益菌肠球菌(Enterococcus)和格氏乳酸菌(Lactococcus garvieae)与NO_3-N和TN呈正相关,红细菌属(Rhodobacter)和假单胞菌(Pseudomonas vranovensis)与TP呈正相关。可见,养殖模式可通过影响水体营养盐尤其是氮磷含量影响罗氏沼虾肠道微生物群落结构。
Microbial communities in the intestine of Macrobrachium rosenbergii under six cultural patterns were evaluated by high-throughput pyrosequencing technology. The interactions between microbial and water environment were also analyzed by RDA(Redundancy analysis). The cultural patterns included prawn monoculture group(MP), prawn with aquatic plant(Lemna minor) group(PP), prawn with silver carp(Hypophthalmichthys molitrix) group(PF), prawn with mussel(Anodonta woodiana) and H. molitrix group(PMF), prawn with mussel and aquatic plant group(PMP), prawn with A. woodiana, L. minor and H. molitrix group(PMPF). Four replicates of each treatment were set up. After 64 days of culturing, the microbial communities and physico-chemical factors were assessed. There were significant differences in turbidity, soluble phosphorus(PO_4-P), total nitrogen(TN) and chlorophyll-a(Chl.a) among groups, and PO_4-P concentration in MP group was significantly higher than those in other groups(P< 0.05). Shannon diversity index of intestinal bacteria was the highest in the MP group(4.08), and the lowest in the PMF group(1.27). Proteobacteria, Tenericute and Firmicutes were the major dominant phyla in intestine of M. rosenbergii in the six groups. The major dominant genus was different in six groups, which was Aeromonas in MP, PMP and PMPF groups, and Citrobacter in PP group, and Candidatus and Hepatoplasma in PF and PMF groups. The interaction analysis between intestinal bacteria and environmental factors showed that total phosphorus(TP) had significant effects on the intestinal bacterial community of M. rosenbergii(P<0.05), Adaptation of microorganisms to the environment factors was different, and the abundance of Aeromonas enteropelogenes, beneficial Enterococcus and Lactococcus garvieae were positive correlation with NO_3-N and TN. Rhodobacter and Pseudomonas vranovensis were positive correlation with TP. Conclusively, cultural patterns can affect the intestinal bacterial community of M. rosenbergii by affecting the nutrient concentration in the water, especially nitrogen and phosphorus.
Microbial communities in the intestine of Macrobrachium rosenbergii under six cultural patterns were evaluated by high-throughput pyrosequencing technology. The interactions between microbial and water environment were also analyzed by RDA(Redundancy analysis). The cultural patterns included prawn monoculture group(MP), prawn with aquatic plant(Lemna minor) group(PP), prawn with silver carp(Hypophthalmichthys molitrix) group(PF), prawn with mussel(Anodonta woodiana) and H. molitrix group(PMF), prawn with mussel and aquatic plant group(PMP), prawn with A. woodiana, L. minor and H. molitrix group(PMPF). Four replicates of each treatment were set up. After 64 days of culturing, the microbial communities and physico-chemical factors were assessed. There were significant differences in turbidity, soluble phosphorus(PO_4-P), total nitrogen(TN) and chlorophyll-a(Chl.a) among groups, and PO_4-P concentration in MP group was significantly higher than those in other groups(P< 0.05). Shannon diversity index of intestinal bacteria was the highest in the MP group(4.08), and the lowest in the PMF group(1.27). Proteobacteria, Tenericute and Firmicutes were the major dominant phyla in intestine of M. rosenbergii in the six groups. The major dominant genus was different in six groups, which was Aeromonas in MP, PMP and PMPF groups, and Citrobacter in PP group, and Candidatus and Hepatoplasma in PF and PMF groups. The interaction analysis between intestinal bacteria and environmental factors showed that total phosphorus(TP) had significant effects on the intestinal bacterial community of M. rosenbergii(P<0.05), Adaptation of microorganisms to the environment factors was different, and the abundance of Aeromonas enteropelogenes, beneficial Enterococcus and Lactococcus garvieae were positive correlation with NO_3-N and TN. Rhodobacter and Pseudomonas vranovensis were positive correlation with TP. Conclusively, cultural patterns can affect the intestinal bacterial community of M. rosenbergii by affecting the nutrient concentration in the water, especially nitrogen and phosphorus.
引文
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[19] WU S G,WANG G T,ANGERT E R,et al.Composition,diversity,and origin of the bacterial community in grass carp intestine[J].PLoS One,2012,7(2):e30440.
[20] ROESELERS G,MITTGE E K,STEPHENS W Z,et al.Evidence for a core gut microbiota in the zebrafish[J].ISME Journal,2011,5(10):1595-1608.
[21] 郁维娜,戴文芳,陶震,等.健康与患病凡纳滨对虾肠道菌群结构及功能差异研究[J].水产学报,2018,42(3):399-409.YU W N,DAI W F,TAO Z,et al.Characterizing the compositional and functional structures of intestinal micro-flora between healthy and diseased Litopenaeus vannamei[J].Journal of Fisheries of China,2018,42(3):399-409.
[22] MIAO S Y,ZHU J Y,ZHAO C Z,et al.Effects of C/N ratio control combined with probiotics on the immune response,disease resistance,intestinal microbiota and morphology of giant freshwater prawn (Macrobrachium rosenbergii)[J].Aquaculture,2017,476:125-133.
[23] 蔡仲石.三种不同环境中鲢鳙肠道微生物研究[D].上海:上海海洋大学,2014.CAI Z S.The research of intestinal microbial in Hypophthalmichthys molitrix and Aristichthys nobilis in three different environment[D].Shanghai:Shanghai Ocean University,2014.
[24] HENNERSDORF P,MROTZEK G,ABDUL-AZIZ M A,et al.Metagenomic analysis between free-living and cultured Epinephelus fuscoguttatus under different environmental conditions in Indonesian waters[J].Marine Pollution Bulletin,2016,110(2):726-734.
[25] LI J J,NI J J,LI J L,et al.Comparative study on gastrointestinal microbiota of eight fish species with different feeding habits[J].Journal of Applied Microbiology,2014,117(6):1750-1760.
[26] LIU H,GUO X W,GOONERATNE R,et al.The gut microbiome and degradation enzyme activity of wild freshwater fishes influenced by their trophic levels[J].Scientific Reports,2016,6:24340.
[27] 车建锋、李志斐、王广军,等.混养池塘生物膜微生物群落功能多样性特征分析[J].上海海洋大学学报,2017,26(6):862-871.CHE J F,LI Z F,WANG G J,et al.Microbial metabolic characteristics of biofilm communities in polyculture pond[J].Journal of Shanghai Ocean University,2017,26(6):862-871.
[28] SCHMIDT V T,SMITH K F,MELVIN D W,et al.Community assembly of a euryhaline fish microbiome during salinity acclimation[J].Molecular Ecology,2015,24(10):2537-2550.
[29] SULLAM K E,RUBIN B E R,DALTON C M,et al.Divergence across diet,time and populations rules out parallel evolution in the gut microbiomes of Trinidadian guppies[J].The ISME Journal,2015,9(7):1508-1522.
[2] RAWLS J F,SAMUEL B S,GORDON J I.Gnotobiotic zebrafish reveal evolutionarily conserved responses to the gut microbiota[J].Proceedings of the National Academy of Sciences of the United States of America,2004,101(13):4596-4601.
[3] CHAIYAPECHARA S,RUNGRASSAMEE W,SURIYACHAY I,et al.Bacterial community associated with the intestinal tract of P.monodon in commercial farms[J].Microbial Ecology,2012,63(4):938-953.
[4] ZHANG M L,SUN Y H,CHEN K,et al.Characterization of the intestinal microbiota in Pacific white shrimp,Litopenaeus vannamei,fed diets with different lipid sources[J].Aquaculture,2014,434:449-455.
[5] LIU H D,WANG L,LIU M,et al.The intestinal microbial diversity in Chinese shrimp (Fenneropenaeus chinensis) as determined by PCR-DGGE and clone library analyses[J].Aquaculture,2011,317(1/4):32-36.
[6] TZENG T D,PAO Y Y,CHEN P C,et al.Effects of host phylogeny and habitats on gut microbiomes of oriental river prawn (Macrobrachium nipponense)[J].PLoS One,2015,10(7):e0132860.
[7] MENTE E,GANNON A T,NIKOULI E,et al.Gut microbial communities associated with the molting stages of the giant freshwater prawn Macrobrachium rosenbergii[J].Aquaculture,2016,463:181-188.
[8] RUNGRASSAMEE W,KLANCHUI A,MAIBUNKAEW S,et al.Characterization of intestinal bacteria in wild and domesticated adult black tiger shrimp (Penaeus monodon)[J].PLoS One,2014,9(3):e91853.
[9] RUNGRASSAMEE W,KLANCHUI A,CHAIYAPECHARA S,et al.Bacterial population in intestines of the black tiger shrimp (Penaeus monodon) under different growth stages[J].PLoS One,2013,8(4):e60802.
[10] ZHANG M L,SUN Y H,CHEN K,et al.Characterization of the intestinal microbiota in Pacific white shrimp,Litopenaeus vannamei fed diets with different lipid sources[J].Aquaculture,2014,434:449-455.
[11] ZHENG Y F,YU M,LIU Y,et al.Comparison of cultivable bacterial communities associated with Pacific white shrimp (Litopenaeus vannamei) larvae at different health statuses and growth stages[J].Aquaculture,2016,451:163-169.
[12] 孙振丽,宣引明,张皓,等.南美白对虾养殖环境及其肠道细菌多样性分析[J].中国水产科学,2016,23(3):594-605.SUN Z L,XUAN Y M,ZHANG H,et al.Bacterial diversity in the Penaeus vannamei Boone intestine and aquaculture environment[J].Journal of Fishery Sciences of China,2016,23(3):594-605.
[13] 刘其根,杨洋,唐永涛,等.罗氏沼虾与三角帆蚌、鲢和鳙混养模式优化[J].水产学报,2014,38(11):1855-1864.LIU Q G,YANG Y,TANG Y T,et al.Experimental study on structure optimization for polyculture of giant freshwater prawn with triangle sail mussel,silver carp and bighead carp[J].Journal of Fisheries of China,2014,38(11):1855-1864.
[14] 张皓.养殖环境微生物群落的动态变化及与水环境的互作[D].苏州:苏州大学,2015.ZHANG H.The dynamic changes of microbial communities in aquaculture environment and interactions with water environment[D].Suzhou:Soochow University,2015.
[15] 尹军霞,沈文英,郦萍.水温对南美白对虾肠道菌群影响的研究[J].海洋科学,2004,28(5):33-36.YIN J X,SHEN W Y,LI P.Study on the influence of water temperature on the intestinal microflora of Penaeus vannamei[J].Marine Sciences,2004,28(5):33-36.
[16] 金相灿,屠清瑛.湖泊富营养化调查规范[M].2版.北京:中国环境科学出版社,1990:160-188.JIN X C,TU Q Y.The investigation standard of lake eutrophication[M].2nd ed.Beijing:China Environmental Science Press,1990:160-188.
[17] 章宗涉,黄祥飞.淡水浮游生物研究方法[M].北京:科学出版社,1991:345-347.ZHANG Z S,HUANG X F.Methods for the plankton study in freshwater[M].Beijing:Science Press,1991:345-347.
[18] 中华人民共和国国家质量监督检验检疫总局,中国国家标准化管理委员会.GB 11607—1989 渔业水质标准[S].北京:中国标准出版社,1989.General Administration of Quality Supervision,Inspection and Quarantine of the People’s Republic of China,Standardization Administration of the People’s Republic of China.GB 11607-1989 Water quality standard for fisheries[S].Beijing:Standards Press of China,1989.
[19] WU S G,WANG G T,ANGERT E R,et al.Composition,diversity,and origin of the bacterial community in grass carp intestine[J].PLoS One,2012,7(2):e30440.
[20] ROESELERS G,MITTGE E K,STEPHENS W Z,et al.Evidence for a core gut microbiota in the zebrafish[J].ISME Journal,2011,5(10):1595-1608.
[21] 郁维娜,戴文芳,陶震,等.健康与患病凡纳滨对虾肠道菌群结构及功能差异研究[J].水产学报,2018,42(3):399-409.YU W N,DAI W F,TAO Z,et al.Characterizing the compositional and functional structures of intestinal micro-flora between healthy and diseased Litopenaeus vannamei[J].Journal of Fisheries of China,2018,42(3):399-409.
[22] MIAO S Y,ZHU J Y,ZHAO C Z,et al.Effects of C/N ratio control combined with probiotics on the immune response,disease resistance,intestinal microbiota and morphology of giant freshwater prawn (Macrobrachium rosenbergii)[J].Aquaculture,2017,476:125-133.
[23] 蔡仲石.三种不同环境中鲢鳙肠道微生物研究[D].上海:上海海洋大学,2014.CAI Z S.The research of intestinal microbial in Hypophthalmichthys molitrix and Aristichthys nobilis in three different environment[D].Shanghai:Shanghai Ocean University,2014.
[24] HENNERSDORF P,MROTZEK G,ABDUL-AZIZ M A,et al.Metagenomic analysis between free-living and cultured Epinephelus fuscoguttatus under different environmental conditions in Indonesian waters[J].Marine Pollution Bulletin,2016,110(2):726-734.
[25] LI J J,NI J J,LI J L,et al.Comparative study on gastrointestinal microbiota of eight fish species with different feeding habits[J].Journal of Applied Microbiology,2014,117(6):1750-1760.
[26] LIU H,GUO X W,GOONERATNE R,et al.The gut microbiome and degradation enzyme activity of wild freshwater fishes influenced by their trophic levels[J].Scientific Reports,2016,6:24340.
[27] 车建锋、李志斐、王广军,等.混养池塘生物膜微生物群落功能多样性特征分析[J].上海海洋大学学报,2017,26(6):862-871.CHE J F,LI Z F,WANG G J,et al.Microbial metabolic characteristics of biofilm communities in polyculture pond[J].Journal of Shanghai Ocean University,2017,26(6):862-871.
[28] SCHMIDT V T,SMITH K F,MELVIN D W,et al.Community assembly of a euryhaline fish microbiome during salinity acclimation[J].Molecular Ecology,2015,24(10):2537-2550.
[29] SULLAM K E,RUBIN B E R,DALTON C M,et al.Divergence across diet,time and populations rules out parallel evolution in the gut microbiomes of Trinidadian guppies[J].The ISME Journal,2015,9(7):1508-1522.