紫菜养殖对养殖水体中细菌多样性分布及环境因子的影响
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摘要
采用基于16S rDNA的高通量测序技术对山东长岛紫菜养殖区的细菌多样性分布进行了研究,并与环境因子进行关联分析,以探究紫菜养殖对环境的影响。结果显示,长岛海区水环境细菌多样性较丰富,呈现由近岸区(CDCNS)、养殖区(CDPF)、外海区(CDCOS)递减的趋势。分别基于所有细菌组成以及相对丰度最高的前20个属/种的聚类结果均显示,CDCNS和CDCOS区聚在一起,明显区别于CDPF区。CDPF区的特异优势菌包括鼠尾菌属(Muricauda)、假单胞菌属(Pseudomonas)、盐单胞菌属(Halomonas)、赤杆菌属(Erythrobacter)、海杆菌属(Marinobacter)。3个区域均未检测到河豚毒素假交替单胞菌(Pseudoalteromonastetraodonis)、柠檬假交替单胞菌(Pseudoalteromonas citrea)等疑似紫菜致病菌。溶解氧(DO)和pH呈现由近岸向外海逐渐降低的趋势,总颗粒悬浮物(TSS)反之。CDPF区氨氮(NH_4~+-N)含量最低,盐度最高。关联分析发现,环境因子与环境微生物之间具有一定的相关性,如嗜氨菌属(Ammoniphilus)的丰度与NH_4~+-N浓度具显著正相关性,CDPF区最低,CDCOS区最高;盐单胞菌属的分布与盐度具显著正相关性,CDPF区盐单胞菌属含量显著高于其他2个海区。研究表明,紫菜养殖海区细菌群落结构特征与环境因子具有关联性,其间的互作机制尚需进一步深入研究。
Pyropia is one of the most economically important seaweeds, which is mainly cultured in China. In recent years, water quality deterioration caused by long-term and intensive culture have led to problems for Pyropia aquaculture, such as disease outbreaks, decreased yields, and lower product quality. Microorganisms are important components of the environment and have important effects on the growth of other organisms. In this study, the microbial community structure in a Pyropia yezoensis aquafarm in Changdao, China, was studied using 16S rDNA-based high-throughput sequencing technology. Whether there were correlations between different environmental factors and the abundance of some predominant bacterial taxa was also tested to explore the environmental impacts of Pyropia farming. The results showed that microbial diversity was rich in the surveyed marine area. The abundance of Proteobacteria was the highest among all of the identified bacterial phyla. When the Pyropia farming area was compared with nearby non-farming areas, it was found that the highest abundances of microbial species occurred in the near-shore area(CDCNS), followed by the Pyropia farming area(CDPF), and the microbial diversity was the lowest in the offshore area(CDCOS). Cluster analysis of the samples was carried out based on Bray-Curtis similarity coefficient values calculated among them. The microbial community structure of CDCNS and CDCOS formed a clade that was separated from CDPF. Furthermore, the twenty genera with the highest abundances were used for a further cluster analysis, and the results showed that CDCNS and CDCOS clustered together, with more genera being especially abundant in CDPF. The predominant bacteria from CDPF included Muricauda, Pseudomonas, Halomonas, Erythrobacter, and Marinobacter. Suspected pathogenic bacteria of Pyropia, such as Pseudoalteromonas tetraodonis and Pseudoalteromonas citrea, were not identified in the surveyed areas. Indeed, the abundance of the genus Pseudoalteromonas was lower than 0.1%, suggesting that the environment was safe for Pyropia farming. Dissolved oxygen(DO) and pH decreased gradually from CDCNS to CDCOS, and TSS increased gradually from CDCNS to CDCOS. The content of NH_4~+-N was the lowest in CDPF and the highest in CDCOS, while the salinity was the highest in CDPF. Correlation analyses showed that there were relationships between environmental factors and environmental microorganisms. The abundance of Ammoniphilus was positively correlated with the content of NH_4~+-N, which was the lowest in CDPF and the highest in CDCOS. The distribution of Halomonas was related to salinity, and the abundance of Halomonas was significantly higher in CDPF than in the other two areas. Halomonas functions in denitrification, and thus can promote the conversion of N from an oxidized state to reduced forms that are more easily absorbed by algae. In conclusion, this study represented the first report of the structure of the microbial community in a Pyropia aquafarm and its correlation with environmental factors. The results showed that farming Pyropia had significant influences on the environmental microbial community, which were closely related to variations in environmental factors caused by Pyropia farming. Such changes in environmental factors and microbial structure had corresponding effects on Pyropia growth. The mechanisms of these interactions between environmental microorganisms and Pyropia growth should be further investigated in the future.
Pyropia is one of the most economically important seaweeds, which is mainly cultured in China. In recent years, water quality deterioration caused by long-term and intensive culture have led to problems for Pyropia aquaculture, such as disease outbreaks, decreased yields, and lower product quality. Microorganisms are important components of the environment and have important effects on the growth of other organisms. In this study, the microbial community structure in a Pyropia yezoensis aquafarm in Changdao, China, was studied using 16S rDNA-based high-throughput sequencing technology. Whether there were correlations between different environmental factors and the abundance of some predominant bacterial taxa was also tested to explore the environmental impacts of Pyropia farming. The results showed that microbial diversity was rich in the surveyed marine area. The abundance of Proteobacteria was the highest among all of the identified bacterial phyla. When the Pyropia farming area was compared with nearby non-farming areas, it was found that the highest abundances of microbial species occurred in the near-shore area(CDCNS), followed by the Pyropia farming area(CDPF), and the microbial diversity was the lowest in the offshore area(CDCOS). Cluster analysis of the samples was carried out based on Bray-Curtis similarity coefficient values calculated among them. The microbial community structure of CDCNS and CDCOS formed a clade that was separated from CDPF. Furthermore, the twenty genera with the highest abundances were used for a further cluster analysis, and the results showed that CDCNS and CDCOS clustered together, with more genera being especially abundant in CDPF. The predominant bacteria from CDPF included Muricauda, Pseudomonas, Halomonas, Erythrobacter, and Marinobacter. Suspected pathogenic bacteria of Pyropia, such as Pseudoalteromonas tetraodonis and Pseudoalteromonas citrea, were not identified in the surveyed areas. Indeed, the abundance of the genus Pseudoalteromonas was lower than 0.1%, suggesting that the environment was safe for Pyropia farming. Dissolved oxygen(DO) and pH decreased gradually from CDCNS to CDCOS, and TSS increased gradually from CDCNS to CDCOS. The content of NH_4~+-N was the lowest in CDPF and the highest in CDCOS, while the salinity was the highest in CDPF. Correlation analyses showed that there were relationships between environmental factors and environmental microorganisms. The abundance of Ammoniphilus was positively correlated with the content of NH_4~+-N, which was the lowest in CDPF and the highest in CDCOS. The distribution of Halomonas was related to salinity, and the abundance of Halomonas was significantly higher in CDPF than in the other two areas. Halomonas functions in denitrification, and thus can promote the conversion of N from an oxidized state to reduced forms that are more easily absorbed by algae. In conclusion, this study represented the first report of the structure of the microbial community in a Pyropia aquafarm and its correlation with environmental factors. The results showed that farming Pyropia had significant influences on the environmental microbial community, which were closely related to variations in environmental factors caused by Pyropia farming. Such changes in environmental factors and microbial structure had corresponding effects on Pyropia growth. The mechanisms of these interactions between environmental microorganisms and Pyropia growth should be further investigated in the future.
引文
Bai J,Li HY,Zhao YG.Bacterial distribution at different stations in the Northern Yellow Sea.Acta Microbiologica Sinica,2009,49(3):343-350[白洁,李海艳,赵阳国.黄海北部不同站位海洋细菌群落分布特征.微生物学报,2009,49(3):343-350]
Bronk DA,Glibert PM,Ward BB.Nitrogen uptake disolved organic nitrogen release and new production.Science,1994,265(5180):1843-1846
Chen GY,Shen HS,Zhu MX,et al.Preliminary investigation on attaching bacteria and disease bacteria in thalli of Porphyra yezoensis.Journal of Aquaculture,1999(1):17-19[陈国耀,沈怀舜,朱庙先,等.条斑紫菜叶状体附生菌及病原菌的初步调查.水产养殖,1999(1):17-19]
Fisher MM,Wilcox LW,Graham LE.Molecular characterization of epiphytic bacterial communities on Charophycean green algae.Applied and Environmental Microbiology,1998,64(11):4384-4389
Fu XH,Liu GN,He JL,et al.Analysis of microbial community diversity in the Bohai Sea marine protected areas of the Shandong Province.Marine Sciences,2017,41(1):39-47[付新华,刘国宁,何健龙,等.山东省渤海海洋保护区典型海域表层海水微生物群落多样性分析.海洋科学,2017,41(1):39-47]
Gai SS,Zhang WD.Microbiota of shell-boring conchocelis of Pyropia yezoensis determined by the next-generation sequencing.Oceanologia et Limnologia Sinica,2016,47(5):990-996[盖珊珊,张伟东.基于第二代测序技术的条斑紫菜(Pyropia yezoensis)贝壳丝状体附生菌群研究.海洋与湖沼,2016,47(5):990-996]
Hu XJ.Analysis on microbial community characteristics in the typical sea areas in Guangdong Province.Master￠s Thesis of Jinan University,2013,46-49[胡晓娟.广东典型海域微生物群落特征分析.暨南大学硕士研究生学位论文,2013,46-49]
Li XL,Wang WJ,Liang ZR,et al.Antioxidant physiological characteristics of wild Pyropia yezoensis under desiccation stress.Progress in Fishery Sciences,2017,38(5):156-163[李晓蕾,汪文俊,梁洲瑞,等.野生条斑紫菜叶状体对干出胁迫的抗氧化生理响应特征.渔业科学进展,2017,38(5):156-163]
Li W,Mamat O,Xu JL,et al.Comparative study of volatile components from tow strains of Pyropia haitanensis in different culture areas.Progress in Fishery Sciences,2016,37(5):147-156[李微,阿曼尼萨·买买提,徐继林,等.不同海域不同品种坛紫菜(Pyropia haitanensis)挥发性成分的比较分析.渔业科学进展,2016,37(5):147-156]
Liu JH,Long C,Lu XL,et al.Identification and bioassays of a strain of Halomonas sp.from the East China Sea.Chinese Journal Marine Drugs,2009,28(6):5-10[刘军华,龙聪,卢小玲,等.一株来源于东海的盐单胞菌的鉴定及活性筛选.中国海洋药物杂志,2009,28(6):5-10]
Patrick DS,Sarah LW,Thomas R,et al.Introducing mothur:Open-source,platform-independent,community-supported software for describing and comparing microbial communities.Applied and Environmental Microbiology,2009,75(23):7537-7541
Shen ML,Yang R,Luo QJ,et al.Microbial diversity of Pyropia haitanensis phycosphere during cultivation.Acta Microbiologica Sinica,2013,53(10):1087-1102[沈梅丽,杨锐,骆其君,等.坛紫菜养殖周期中的藻际微生物多样性.微生物学报,2013,53(10):1087-1102]
Wang HB,Li XS,Xia YM,et al.Isolation,identification and biological pathogen of yellow spot disease in conchocelis of Porphyra yezoensis.Marine Environmental Science,2011,30(3):361-364,408[王洪斌,李信书,夏亚明,等.条斑紫菜丝状体黄斑病病原体分离鉴定及生物学特性研究.海洋环境科学,2011,30(3):361-364,408]
Wu HQ.Epibacterial community structure of several macroalgae.Doctoral Dissertation of University of Chinese Academy of Sciences,2012,50-51[武洪庆.不同养殖海藻表面附着细菌多样性分析.中国科学院大学博士研究生学位论文,2012,50-51]
Xiao T,Wang R.Distribution of Synechococcus in the Bohai Sea in Autumn and Spring.Acta Ecologica Sinica,2002,22(12):2071-2078[肖天,王荣.春季与秋季渤海蓝细菌(聚球蓝细菌属)的分布特点.生态学报,2002,22(12):2071-2078]
Yan Y,Ma JH,Xu P,et al.Pseudoalteromonas citrea,the causative agent of green-spot disease of Porphyra yezoensis.Journal of Fishery Sciences of China,2002,9(4):353-358[闫咏,马家海,许璞,等.1株引起条斑紫菜绿斑病的柠檬假交替单胞菌.中国水产科学,2002,9(4):353-358]
Yang R,Fang WY,Shan YY,et al.Genetic diversity of epiphytic bacteria in Porphyra yezoensis.Acta Oceaologica Sinica,2008,30(4):161-168[杨锐,方文雅,单媛媛,等.条斑紫菜外生细菌的遗传多样性.海洋学报,2008,30(4):161-168]
Zaitsev GM,Tsitko IV,Rainey FA,et al.New aerobic ammonium-dependent obligately oxalotrophic bacteria:Description of Ammoniphilus oxalaticus gen.nov.,sp.Nov.and Ammoniphilus oxalivorans gen.nov.,sp.nov.International.Journal of Systematic Bacteriology,1998,48(1):151-163
Zhang DY,Zhang TX,Wang GX.Development and application of second-generation sequencing technology.Environmental Science and Technology,2016,39(9):96-102[张丁予,章婷曦,王国祥.第二代测序技术的发展及应用.环境科学与技术,2016,39(9):96-102]
Zhao GC,Zhang T,Gao Y,et al.Identification and bioactivities analysis of Halomonas sp.from seawater.Academic Journal of Second Military Medical University,2011,32(8):813-817[赵贵成,张韬,高云,等.海洋盐单胞菌属微生物的鉴定及生物学活性分析.第二军医大学学报,2011,32(8):813-817]
Bronk DA,Glibert PM,Ward BB.Nitrogen uptake disolved organic nitrogen release and new production.Science,1994,265(5180):1843-1846
Chen GY,Shen HS,Zhu MX,et al.Preliminary investigation on attaching bacteria and disease bacteria in thalli of Porphyra yezoensis.Journal of Aquaculture,1999(1):17-19[陈国耀,沈怀舜,朱庙先,等.条斑紫菜叶状体附生菌及病原菌的初步调查.水产养殖,1999(1):17-19]
Fisher MM,Wilcox LW,Graham LE.Molecular characterization of epiphytic bacterial communities on Charophycean green algae.Applied and Environmental Microbiology,1998,64(11):4384-4389
Fu XH,Liu GN,He JL,et al.Analysis of microbial community diversity in the Bohai Sea marine protected areas of the Shandong Province.Marine Sciences,2017,41(1):39-47[付新华,刘国宁,何健龙,等.山东省渤海海洋保护区典型海域表层海水微生物群落多样性分析.海洋科学,2017,41(1):39-47]
Gai SS,Zhang WD.Microbiota of shell-boring conchocelis of Pyropia yezoensis determined by the next-generation sequencing.Oceanologia et Limnologia Sinica,2016,47(5):990-996[盖珊珊,张伟东.基于第二代测序技术的条斑紫菜(Pyropia yezoensis)贝壳丝状体附生菌群研究.海洋与湖沼,2016,47(5):990-996]
Hu XJ.Analysis on microbial community characteristics in the typical sea areas in Guangdong Province.Master￠s Thesis of Jinan University,2013,46-49[胡晓娟.广东典型海域微生物群落特征分析.暨南大学硕士研究生学位论文,2013,46-49]
Li XL,Wang WJ,Liang ZR,et al.Antioxidant physiological characteristics of wild Pyropia yezoensis under desiccation stress.Progress in Fishery Sciences,2017,38(5):156-163[李晓蕾,汪文俊,梁洲瑞,等.野生条斑紫菜叶状体对干出胁迫的抗氧化生理响应特征.渔业科学进展,2017,38(5):156-163]
Li W,Mamat O,Xu JL,et al.Comparative study of volatile components from tow strains of Pyropia haitanensis in different culture areas.Progress in Fishery Sciences,2016,37(5):147-156[李微,阿曼尼萨·买买提,徐继林,等.不同海域不同品种坛紫菜(Pyropia haitanensis)挥发性成分的比较分析.渔业科学进展,2016,37(5):147-156]
Liu JH,Long C,Lu XL,et al.Identification and bioassays of a strain of Halomonas sp.from the East China Sea.Chinese Journal Marine Drugs,2009,28(6):5-10[刘军华,龙聪,卢小玲,等.一株来源于东海的盐单胞菌的鉴定及活性筛选.中国海洋药物杂志,2009,28(6):5-10]
Patrick DS,Sarah LW,Thomas R,et al.Introducing mothur:Open-source,platform-independent,community-supported software for describing and comparing microbial communities.Applied and Environmental Microbiology,2009,75(23):7537-7541
Shen ML,Yang R,Luo QJ,et al.Microbial diversity of Pyropia haitanensis phycosphere during cultivation.Acta Microbiologica Sinica,2013,53(10):1087-1102[沈梅丽,杨锐,骆其君,等.坛紫菜养殖周期中的藻际微生物多样性.微生物学报,2013,53(10):1087-1102]
Wang HB,Li XS,Xia YM,et al.Isolation,identification and biological pathogen of yellow spot disease in conchocelis of Porphyra yezoensis.Marine Environmental Science,2011,30(3):361-364,408[王洪斌,李信书,夏亚明,等.条斑紫菜丝状体黄斑病病原体分离鉴定及生物学特性研究.海洋环境科学,2011,30(3):361-364,408]
Wu HQ.Epibacterial community structure of several macroalgae.Doctoral Dissertation of University of Chinese Academy of Sciences,2012,50-51[武洪庆.不同养殖海藻表面附着细菌多样性分析.中国科学院大学博士研究生学位论文,2012,50-51]
Xiao T,Wang R.Distribution of Synechococcus in the Bohai Sea in Autumn and Spring.Acta Ecologica Sinica,2002,22(12):2071-2078[肖天,王荣.春季与秋季渤海蓝细菌(聚球蓝细菌属)的分布特点.生态学报,2002,22(12):2071-2078]
Yan Y,Ma JH,Xu P,et al.Pseudoalteromonas citrea,the causative agent of green-spot disease of Porphyra yezoensis.Journal of Fishery Sciences of China,2002,9(4):353-358[闫咏,马家海,许璞,等.1株引起条斑紫菜绿斑病的柠檬假交替单胞菌.中国水产科学,2002,9(4):353-358]
Yang R,Fang WY,Shan YY,et al.Genetic diversity of epiphytic bacteria in Porphyra yezoensis.Acta Oceaologica Sinica,2008,30(4):161-168[杨锐,方文雅,单媛媛,等.条斑紫菜外生细菌的遗传多样性.海洋学报,2008,30(4):161-168]
Zaitsev GM,Tsitko IV,Rainey FA,et al.New aerobic ammonium-dependent obligately oxalotrophic bacteria:Description of Ammoniphilus oxalaticus gen.nov.,sp.Nov.and Ammoniphilus oxalivorans gen.nov.,sp.nov.International.Journal of Systematic Bacteriology,1998,48(1):151-163
Zhang DY,Zhang TX,Wang GX.Development and application of second-generation sequencing technology.Environmental Science and Technology,2016,39(9):96-102[张丁予,章婷曦,王国祥.第二代测序技术的发展及应用.环境科学与技术,2016,39(9):96-102]
Zhao GC,Zhang T,Gao Y,et al.Identification and bioactivities analysis of Halomonas sp.from seawater.Academic Journal of Second Military Medical University,2011,32(8):813-817[赵贵成,张韬,高云,等.海洋盐单胞菌属微生物的鉴定及生物学活性分析.第二军医大学学报,2011,32(8):813-817]