大熊猫粪便菌群ERIC-PCR指纹图谱的分析及优势菌群的鉴定
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摘要
探讨1只野生大熊猫和不同年龄段圈养大熊猫粪便菌群的多样性和相似性,并鉴定野生大熊猫圈养之后,其粪便中的优势菌群。对来自3个年龄段9只(3只亚成体、3只成年体、3只老年体)圈养大熊猫及1只野生大熊猫圈养前、后的11份粪便细菌的DNA进行ERIC-PCR指纹图谱分析,并利用16S rRNA基因序列分析对优势菌条带进行鉴定。结果显示:该只野生大熊猫粪便菌群多样性比圈养大熊猫丰富;圈养大熊猫粪便菌群多样性:成年体>亚成体>老年体;16s rRNA和ERIC-PCR指纹图谱优势条带鉴定的优势菌结果有差异性。结果表明:大熊猫的肠道菌群多样性易受生长环境、年龄因素的影响;其次,不同年龄段的圈养大熊猫,其肠道菌群的相似性与年龄因素不相关;利用ERIC-PCR指纹图谱优势条带鉴定优势菌的方法有一定的局限性。
In the current study,the diversity and similarity of the gut were investigated for a wild giant panda and 9 captive giant pandas of different age groups,and the dominant bacteria of the intestinal flora was identified for the wild giant after the captivity.Fecal samples of captive pandas were collected from 3 subadult,3 adult and 3 senile pandas,and 1 wild panda before and after at Bifengxia zoological institutions in Ya'an.Bacteria DNA was extracted from fecal bacteria and analyzed by ERIC-PCR.An advantage bacteria DNA band on the fingerprint of the wild giant panda in captivity was purified,cloned,sequenced and alignmented.Then,the predominant bacteria were checked again via 16S rRNA method and compared with the results of identification of ERIC-PCR.The results revealed that the diversity of intestinal flora of the wild giant panda was more abundant than that of captive giant pandas and the diversity of feces intestinal flora of adult pandas group was most abundant than that of the other pandas in captive populations groups.In addition,the predominant flora results via ERIC PCR fingerprinting and 16S rRNA method were not related,implyed that the ERIC-PCR fingerprinting method was limited.No significant similarity of the intestinal flora regarding to the ages factor was seen in all giant pandas.Our results indicated that the diversity of intestinal flora of the giant panda was susceptible to the growth environmental and age factors.
In the current study,the diversity and similarity of the gut were investigated for a wild giant panda and 9 captive giant pandas of different age groups,and the dominant bacteria of the intestinal flora was identified for the wild giant after the captivity.Fecal samples of captive pandas were collected from 3 subadult,3 adult and 3 senile pandas,and 1 wild panda before and after at Bifengxia zoological institutions in Ya'an.Bacteria DNA was extracted from fecal bacteria and analyzed by ERIC-PCR.An advantage bacteria DNA band on the fingerprint of the wild giant panda in captivity was purified,cloned,sequenced and alignmented.Then,the predominant bacteria were checked again via 16S rRNA method and compared with the results of identification of ERIC-PCR.The results revealed that the diversity of intestinal flora of the wild giant panda was more abundant than that of captive giant pandas and the diversity of feces intestinal flora of adult pandas group was most abundant than that of the other pandas in captive populations groups.In addition,the predominant flora results via ERIC PCR fingerprinting and 16S rRNA method were not related,implyed that the ERIC-PCR fingerprinting method was limited.No significant similarity of the intestinal flora regarding to the ages factor was seen in all giant pandas.Our results indicated that the diversity of intestinal flora of the giant panda was susceptible to the growth environmental and age factors.
引文
[1]邹兴淮,曾鲁军,孙中武,等.大熊猫疾病死亡因素分析及其保护对策[J].东北林业大学学报,1998,26(1):53-56.
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[10]Wei G F,Lu H F,Zhou Z H.The Microbial Community in the Feces of the Giant Panda(Ailuropoda melanoleuca)as Determined by PCR-TGGE Profiling and Clone Library Analysis[J].Microbial Ecology,2007,54:194-202.
[11]谢浩,刘斌,罗瑜,等.野化培训大熊猫的采食动态及食物利用率[J].野生动物,2011,32(4):186-190.
[12]丁俊荣,张秋香,刘小鸣,等.江苏无锡健康与肠病人群肠道脱硫弧菌数量及肠道菌群多样性[J].微生物学报,2012,52(8):1033-1039.
[13]杜惠敏,陈颖虹,陈俊仪,等.轮状病毒感染与健康儿童肠道菌群结构的比较[J].实用儿科临床杂志,2006,21(18):1237-1249.
[14]鲍楠,刘成君,张和民,等.大熊猫肠道微生态的研究进展[J].畜牧与兽医,2005,37(4):57-59.
[15]王燚,何廷美,钟志军,等.不同季节亚成体大熊猫肠道菌群ERIC-PCR指纹图谱分析[J].中国兽医科学,2011,41(8):778-783.
[16]Villers L M,Jang S S,Lent C L,et al.Survey and comparison of major intestinal flora in captive and wild ring‐tailed lemur(Lemur catta)populations[J].American Journal of Primatology,2008,70(2):175-184.
[2]孙飞龙,刘敬贤,席丹,等.大熊猫肠道疾病致病菌[J].经济动物学报,2002,6(2)20-23.
[3]高卫科.应用ERIC-PCR方法对不同日龄猪肠道菌群分布规律的研究[D].郑州:河南农业大学,2008.
[4]鲁海峰,魏桂芳,李仲逵,等.ERIC-PCR分子杂交技术分析大熊猫肠道菌群结构[J].中国生态学杂志,2005,17(2):81-84.
[5]Brock T D.The study of microorganisms in situ:progress and problems[M].Symposia of the Society for General Microbiology.1987:96.
[6]di Giovanni G D,Watrud L S,Seidler R J,et al.Fingerprinting of mixed bacterial strains and BIOLOG gram-negative(GN)substrate communities by enterobacterial repetitive intergenic consensus sequence-PCR(ERIC-PCR)[J].Current microbiology,1999,38(4):217-223.
[7]李亮,魏桂芳,童耕雷,等.用PCR指纹图技术分析焦化废水接触氧化池中悬浮污泥和生物膜的微生物种群组成[J].中国微生态学杂志,2004,16(1):8-12.
[8]胡锦矗.从野外大熊猫的粪便估计年龄及其种群年龄结构的研究[J].兽类学报,1987,7(2):81-84.
[9]崔婷婷.基于16S rDNA-RFLP技术对圈养成年大熊猫秋季肠道菌群多样性的研究[D].四川省雅安市:四川农业大学,2011.
[10]Wei G F,Lu H F,Zhou Z H.The Microbial Community in the Feces of the Giant Panda(Ailuropoda melanoleuca)as Determined by PCR-TGGE Profiling and Clone Library Analysis[J].Microbial Ecology,2007,54:194-202.
[11]谢浩,刘斌,罗瑜,等.野化培训大熊猫的采食动态及食物利用率[J].野生动物,2011,32(4):186-190.
[12]丁俊荣,张秋香,刘小鸣,等.江苏无锡健康与肠病人群肠道脱硫弧菌数量及肠道菌群多样性[J].微生物学报,2012,52(8):1033-1039.
[13]杜惠敏,陈颖虹,陈俊仪,等.轮状病毒感染与健康儿童肠道菌群结构的比较[J].实用儿科临床杂志,2006,21(18):1237-1249.
[14]鲍楠,刘成君,张和民,等.大熊猫肠道微生态的研究进展[J].畜牧与兽医,2005,37(4):57-59.
[15]王燚,何廷美,钟志军,等.不同季节亚成体大熊猫肠道菌群ERIC-PCR指纹图谱分析[J].中国兽医科学,2011,41(8):778-783.
[16]Villers L M,Jang S S,Lent C L,et al.Survey and comparison of major intestinal flora in captive and wild ring‐tailed lemur(Lemur catta)populations[J].American Journal of Primatology,2008,70(2):175-184.