零换水条件下益生菌组合在凡纳滨对虾育苗生产中的应用
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摘要
为探索应用益生菌的凡纳滨对虾零换水育苗技术,选择具有不同生态学功能的四株益生菌进行配伍形成两个益生菌组合PBE和PBN,并应用于凡纳滨对虾无节V期幼体(N5)至仔虾第五天(PL5)的育苗实验。每个实验组包含6个平行,同时设置对照组。整个实验过程不换水,PBE和PBN组的水体中连续施用益生菌组合。当幼体发育至蚤状幼体Ⅲ期(ZⅢ)、糠虾幼体Ⅲ期(MⅢ)和PL5时,检测幼体变态率、幼体体内和水体中可培养细菌总数(total culturable bacterial count,TBC)和假定弧菌总数(total presumptive Vibriobacteria,TPVB);在实验结束时统计各实验组幼体成活率,并用Illumina测序PL5幼体体内的微生物群落。结果显示:在整个育苗周期内,未发生疫病;与对照组相比,益生菌组合显著降低了(P <0. 05) TPVB在幼体体内和育苗水体中的相对含量,并显著提高了(P <0. 05) ZⅢ幼体和仔虾PL5的变态率和成活率(各增加15%)。各实验组的PL5体内核心微生物群的多样性和结构并不存在显著性差异(P> 0. 05),红杆菌科是微生物群落中最具优势的菌群,在各实验组中的相对丰度为40%～86%。研究表明,零换水条件下,益生菌组合能够有效抑制弧菌在育苗水体和凡纳滨对虾幼体体内的过度增殖,提高幼体的成活率,实现育苗的健康生产。
In order to explore breeding technology with the application of probiotics combinations for Penaeus vannamei under zero-water exchange conditions,four probiotic bacteria with different ecological functions were formulated to two combinations( i. e.,PBE and PEN),which were applied in the breeding experiment from nauplius larval phase V( N5) to postlarvae phase V( PL5). Each experimental group contained six parallels and the control group was set at the same time. Culturing water was not exchanged throughout the experiment,and probiotics combinations were continuously applied to the water in the PBE and PBN groups. The larval metamorphosis rate,total culturable bacterial count( TBC) and total presumptive Vibrio bacteria( TPVB) in larva and water were counted at the phases of zoea Ⅲ( ZⅢ),mysis Ⅲ( MⅢ) and PL5. At the end of the experiment,the survival rate of postlarvae in each experimental group was determined,and the microbial communities from postlarvae were sequenced using an Illumina platform. The results showed that there was no disease outbreak in each group,and that probiotics combinations significantly decreased( P < 0. 05) relative abundances of TPVB in postlarvae and culturing water in the groups of PBE and PBN compared to that in the control group,and significantly increased( P < 0. 05) the larval metamorphosis rate( 15%) and survival rate( 15%) at the phases of ZⅢ and PL5,respectively. There was no striking difference of microbial diversities and structures of the core populations in the microbial communities from postlarvae in the three groups,and every microbial community was dominated by Rhodobacteriaceae with the relative abundance ranging from40% to 86%. Taking together,probiotics combinations can effectively inhibit the overabundance of Vibrio in rearing-water and postlarvae and improve the survival rate of postlarvae,thereby achieving healthy breeding of P. vannamei under zero-water exchange conditions.
In order to explore breeding technology with the application of probiotics combinations for Penaeus vannamei under zero-water exchange conditions,four probiotic bacteria with different ecological functions were formulated to two combinations( i. e.,PBE and PEN),which were applied in the breeding experiment from nauplius larval phase V( N5) to postlarvae phase V( PL5). Each experimental group contained six parallels and the control group was set at the same time. Culturing water was not exchanged throughout the experiment,and probiotics combinations were continuously applied to the water in the PBE and PBN groups. The larval metamorphosis rate,total culturable bacterial count( TBC) and total presumptive Vibrio bacteria( TPVB) in larva and water were counted at the phases of zoea Ⅲ( ZⅢ),mysis Ⅲ( MⅢ) and PL5. At the end of the experiment,the survival rate of postlarvae in each experimental group was determined,and the microbial communities from postlarvae were sequenced using an Illumina platform. The results showed that there was no disease outbreak in each group,and that probiotics combinations significantly decreased( P < 0. 05) relative abundances of TPVB in postlarvae and culturing water in the groups of PBE and PBN compared to that in the control group,and significantly increased( P < 0. 05) the larval metamorphosis rate( 15%) and survival rate( 15%) at the phases of ZⅢ and PL5,respectively. There was no striking difference of microbial diversities and structures of the core populations in the microbial communities from postlarvae in the three groups,and every microbial community was dominated by Rhodobacteriaceae with the relative abundance ranging from40% to 86%. Taking together,probiotics combinations can effectively inhibit the overabundance of Vibrio in rearing-water and postlarvae and improve the survival rate of postlarvae,thereby achieving healthy breeding of P. vannamei under zero-water exchange conditions.
引文
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[2]宫春光,于清海,陈福杰.南美白对虾育苗中弧菌病和丝状细菌病的防治对策[J].科学养鱼,2008(9):50-51.GONG C G,YU Q H,CHEN F J.Prevention and treatment of vibrio and filamentous bacterial disease in Litopenaeus vannamei[J].Scientific Fish Farming,2008(9):50-51.
[3]刘建勇.复合微生物制剂在凡纳滨对虾育苗中的应用[J].海洋科学,2005,29(4):36-40.LIU J Y.Effect of microecological compound agent on breeding of Litopenaeus vannamei Boon[J].Marine Sciences,2005,29(4):36-40.
[4]姚晓东.抗生素在水产养殖中应用存在的问题及对策[J].农业与技术,2016,36(24):103.YAO X D.The problems and countermeasures of antibiotics in aquaculture[J].Agriculture&Technology,2016,36(24):103.
[5]CHIU C H,CHENG C H,GUA W R,et al.Dietary administration of the probiotic,Saccharomyces cerevisiae P13,enhanced the growth,innate immune responses,and disease resistance of the grouper,Epinephelus coioides[J].Fish&Shellfish Immunology,2010,26(5):691-698.
[6]LIU K F,CHIU C H,SHIU Y L,et al.Effects of the probiotic,Bacillus subtilis E20,on the survival,development,stress tolerance,and immune status of white shrimp,Litopenaeus vannamei larvae[J].Fish&Shellfish Immunology,2010,28(5/6):837-844.
[7]NIMRAT S,SUKSAWAT S,BOONTHAI T,et al.Potential Bacillus probiotics enhance bacterial numbers,water quality and growth during early development of white shrimp(Litopenaeus vannamei)[J].Veterinary Microbiology,2012,159(3/4):443-450.
[8]吴翔宇,于明超,单洪伟,等.两株益生菌在凡纳滨对虾育苗生产中的应用[J].科学养鱼,2014(2):54-55.WU X Y,YU M C,SHAN H W,et al.The application of two probiotics in the production of prawn seedlings in Litopenaeus vannamei[J].Scientific Fish Farming,2014(2):54-55.
[9]王春迪.凡纳滨对虾防病益生菌的筛选和效果评价[D].上海:上海海洋大学,2016.WANG C D.Screening and evalution of potential probiotics against disease in Litopenaeus vannamei[D].Shanghai:Shanghai Ocean University,2016.
[10]李玉宏.养殖对虾肠道微生物区系的特征与功能[D].上海:上海海洋大学,2013.LI Y H.The characteristic and function research on the intestinal microflora of shrimp[D].Shanghai:Shanghai Ocean University,2013.
[11]王春迪,宋晓玲,张晓静,等.养殖水体中添加蜡样芽孢杆菌PC465对凡纳滨对虾抗病力的影响[J].中国水产科学,2016,23(1):146-155.WANG C D,SONG X L,ZHANG X J,et al.Effects of adding Bacillus cereus PC465 to rearing water on disease resistance of Litopenaeus vannamei[J].Journal of Fishery Sciences of China,2016,23(1):146-155.
[12]张盛静,宋晓玲,赵小金,等.饲料中添加益生菌对凡纳滨对虾抗感染和5种免疫基因表达的影响[J].水产学报,2015,39(6):899-907.ZHANG S J,SONG X L,ZHAO X J,et al.Effects of adding probiotics to the feed on anti-infection and five kinds of immune gene expression of Litopenaeus vannamei[J].Journal of Fisheries of China,2015,39(6):899-907.
[13]LIVAK K J,SCHMITTGEN T D.Analysis of relative gene expression data using real-time quantitative PCR and the2-ΔΔCtmethod[J].Methods,2001,25(4):402-408.
[14]隋昶生,王利华.崂山奶山羊瘤胃微生物总DNA提取方法的优化[J].中国农学通报,2009,25(5):18-21.SUI C S,WANG L H.Extraction method optimizing of total DNA in rumen microorganism of Laoshan dairy[J].Chinese Agricultural Science Bulletin,2009,25(5):18-21.
[15]PEIFFER J A,SPOR A,KOREN O,et al.Diversity and heritability of the maize rhizosphere microbiome under field conditions[J].Proceedings of the National Academy of Sciences of the United States of America,2013,110(16):6548-6553.
[16]朱鹏飞.大菱鲆养殖环境细菌多样性分析及水产病原菌基因芯片检测体系的构建[D].青岛:国家海洋局第一海洋研究所,2015.ZHU P F.Analysis of bacterial communities from turbot(Scophthalmus maximus)aquaculture system and construction of DNA microarray detection system for aquaculture pathogenic bacteria[D].Qingdao:The First Institute of Oceanography,2015.
[17]邱德全,周鲜娇,邱明生.氨氮胁迫下凡纳滨对虾抗病力和副溶血弧菌噬菌体防病效果研究[J].水生生物学报,2008,32(4):455-461.QIU D Q,ZHOU X J,QIU M S.Study on anti-disease ability of Litopenaeus vannamei and the biological control of Vibrio parahaemolyticus bacteriophage under stresses of ammonia nitrogen[J].Acta Hydrobiologica Sinica,2008,32(4):455-461.
[18]徐纪萍,谈小兰,钱辉仁.凡纳滨对虾高密度零换水高产养殖技术[J].水产科技情报,2012,39(1):26-29.XU J P,TAN X L,QIAN H R.The high-density zeroexchange and high-water aquaculture technology for shrimp[J].Fisheries Science&Technology Information,2012,39(1):26-29.
[19]潘康成,杨金龙,王振华,等.枯草芽孢杆菌制剂在南美白对虾育苗上的应用[J].饲料研究,2004(12):33-34.PAN K C,YANG J L,WANG Z H,et al.Application of Bacillus subtilis in the breeding of Litopenaeus vannamei[J].Feed Research,2004(12):33-34.
[20]于鹏,王腾,于明超,等.三株潜在益生菌在凡纳滨对虾苗种培育过程中的应用[J].河北渔业,2018(1):12-16,45.YU P,WANG T,YU MC,et al.Application of three potential probiotics in the cultivation of Litopenaeus vannamei larva[J].Hebei Fisheries,2018(1):12-16,45.
[21]周丽,宫庆礼.海水鱼虾蟹贝病害防治技术[M].青岛:青岛海洋大学出版社,1998:83-86.ZHOU L,GONG Q L.Prevention and treatment of Marine fish,shrimp,crab and shellfish[M].Qingdao:Qingdao Ocean University Press,1998:83-86.
[22]ZOKAEIFAR H,BABAEI N,SAAD C R,et al.Administration of Bacillus subtilis strains in the rearing water enhances the water quality,growth performance,immune response,and resistance against Vibrio harveyi infection in juvenile white shrimp,Litopenaeus vannamei[J].Fish&Shellfish Immunology,2014,36(1):68-74.
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