陕西部分地区山羊毕氏肠微孢子虫多位点基因分型研究
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摘要
为了阐明山羊毕氏肠微孢子虫的遗传多样性,笔者采用基于微卫星(MS1、MS3、MS7)和小卫星(MS4)位点的多位点序列分型技术首次对不同用途山羊的122个毕氏肠微孢子虫分离株进行了多位点序列分型研究。结果发现,在MS1、MS4和MS7位点的扩增效率依次为27.9%(34/122)、18.0%(22/122)、50.8%(62/122),而在MS3位点所有样品均未获得有效扩增。核苷酸序列分析表明,所有样品的MS1、MS4和MS7位点分别具有16、9和18个基因型,共形成了14个MLGs。其中,50份绒山羊阳性样品3个位点扩增效率分别为10.0%(5/50)、14.0%(7/50)和90.0%(45/50),分别具有3、3和10个基因型,共形成了5个MLGs;56份奶山羊阳性样品3个位点的扩增效率依次为28.6%(16/56)、19.6%(11/56)和8.9%(5/56),分别组成4、4、1个基因型,共组成7个不同的MLGs;16份黑山羊阳性样品在3个位点的扩增效率依次为81.3%(13/16)、25.0%(4/16)、75.0%(12/16),分别具有9、2、7个基因型,构成2个MLGs。
To indicate the genetic variability of Enterocytozoon bieneusi in goats,the multi-locus sequence genotypes(MLGs)of 122 E.bieneusi isolates from different production categories were studied using the multi-locus sequence genotyping(MLST)technique based on micro-(MS1,MS3,MS7)and mini-satellite(MS4)loci.The results showed that,the respective amplification efficiencies in loci MS1,MS4 and MS7 were 27.9%(34/122),18.0%(22/122),50.8%(62/122),while no amplifications were detected in the locus MS3.The nucleotide sequences analysis indicated that,16,9and 18 genotypes were found in all positive samples for loci MS1,MS4 and MS7,respectively,forming fourteen MLGs.Among them,the amplification efficiencies of three loci among 50 positive samples of cashmere goats were 10.0%(5/50),14.0%(7/50)and 90.0%(45/50),respectively,belonging to three,three and ten genotypes,and five MLGs.The ampli-fication efficiencies of 56 positive samples of dairy goats in three loci were 28.6%(16/56),19.6%(11/56)and 8.9%(5/56),grouping into four,four,one genotypes,and seven different MLGs.The amplification efficiencies of three loci of 16 positive samples of black goats were 81.3%(13/16)and 25.0%(4/16),75.0%(12/16),clustering into 9,2,7genotypes,and two MLGs.
To indicate the genetic variability of Enterocytozoon bieneusi in goats,the multi-locus sequence genotypes(MLGs)of 122 E.bieneusi isolates from different production categories were studied using the multi-locus sequence genotyping(MLST)technique based on micro-(MS1,MS3,MS7)and mini-satellite(MS4)loci.The results showed that,the respective amplification efficiencies in loci MS1,MS4 and MS7 were 27.9%(34/122),18.0%(22/122),50.8%(62/122),while no amplifications were detected in the locus MS3.The nucleotide sequences analysis indicated that,16,9and 18 genotypes were found in all positive samples for loci MS1,MS4 and MS7,respectively,forming fourteen MLGs.Among them,the amplification efficiencies of three loci among 50 positive samples of cashmere goats were 10.0%(5/50),14.0%(7/50)and 90.0%(45/50),respectively,belonging to three,three and ten genotypes,and five MLGs.The ampli-fication efficiencies of 56 positive samples of dairy goats in three loci were 28.6%(16/56),19.6%(11/56)and 8.9%(5/56),grouping into four,four,one genotypes,and seven different MLGs.The amplification efficiencies of three loci of 16 positive samples of black goats were 81.3%(13/16)and 25.0%(4/16),75.0%(12/16),clustering into 9,2,7genotypes,and two MLGs.
引文
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[9]AKINBO F O,OKAKA C E,OMOREGIE R,et al.Molecular epidemiologic characterization of Enterocytozoon bieneusi in HIV-infected persons in Benin City,Nigeria[J].Am J Trop Med Hyg,2012,86(3):441-445.
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[11]NKININ S W,ASONGANYI T,DIDIER E S,et al.Microsporidian infection is prevalent in healthy people in Cameroon[J].J Clin Microbiol,2007,45(9):2841-2846.
[12]ZHANG X,WANG Z X,SU Y,et al.Identification and genotyping of Enterocytozoon bieneusi in China[J].J Clin Microbiol,2011,49(5):2006-2008.
[13]SAK B,BRADY D,PELIKNOVM,et al.Unapparent microsporidial infection among immunocompetent humans in the Czech Republic[J].J Clin Microbiol,2011,49(3):1064-1070.
[14]WIDMER G,AKIYOSHI D E.Host-specific segregation of ribosomal nucleotide sequence diversity in the microsporidian Enterocytozoon bieneusi[J].Infect Genet Evol,2010,10(1):122-128.
[15]SHI K,LI M J,WANG X X,et al.Molecular survey of Enterocytozoon bieneusi in sheep and goats in China[J].Parasit Vectors,2016,9:23.
[16]SANTN M,FAYER R.Enterocytozoon bieneusi genotype nomenclature based on the internal transcribed spacer sequence:A consensus[J].J Eukaryot Microbiol,2009,56(1):34-38.
[17]FENG Y Y,LI N,DEAREN T,et al.Development of a multilocus sequence typing tool for high-resolution genotyping of Enterocytozoon bieneusi[J].Appl Environ Microbiol,2011,77(14):4822-4828.
[18]NAGPAL A,PRITT B S,LORENZ E C,et al.Disseminated microsporidiosis in a renal transplant recipient:Case report and review of the literature[J].Transpl Infect Dis,2013,15(5):526-532.
[19]DENGJEL B,ZAHLER M,HERMANNS W,et al.Zoonotic potential of Enterocytozoon bieneusi[J].J Clin Microbiol,2001,39(12):4495-4499.
[20]LEE J H.Prevalence and molecular characteristics of Enterocytozoon bieneusi in cattle in Korea[J].Parasitol Res,2007,101(2):391-396.
[21]GHOSH K,WEISS L M.Molecular diagnostic tests for microsporidia[J].Interdiscip Perspect Infect Dis,2009,2009:926521.
[22]BREITENMOSER A C,MATHIS A,BRGI E,et al.High prevalence of Enterocytozoonbieneusi in swine with four genotypes that differ from those identified in humans[J].Parasitology,1999,118(Pt 5):447-453.
[23]FAYER R,SANTIN-DURAN M.Epidemiology of microsporidia in human infections[M]//WEISS L M,BECNEL J J.Microsporidia:Pathogens of Opportunity.UK:John Wiley&Sons,Inc.Press,2014:135-164.
[24]PREZ-LOSADA M,CABEZAS P,CASTRO-NALLAR E,et al.Pathogen typing in the genomics era:MLST and the future of molecular epidemiology[J].Infect Genet Evol,2013,16:38-53.
[25]KARIM M R,DONG H J,YU F C,et al.Genetic diversity in Enterocytozoon bieneusi isolates from dogs and cats in china:Host specificity and public health implications[J].J Clin Microbiol,2014,52(9):3297-3302.
[26]KARIM M R,WANG R J,DONG H J,et al.Genetic polymorphism and zoonotic potential of Enterocytozoon bieneusi from nonhuman primates in China[J].Appl Environ Microbiol,2014,80(6):1893-1898.
[27]LI N,XIAO L H,WANG L,et al.Molecular surveillance of Cryptosporidium spp.,Giardia duodenalis,and Enterocytozoon bieneusi by genotyping and subtyping parasites in wastewater[J].PLoS Negl Trop Dis,2012,6(9):e1809.
[28]LI W,DENG L,YU X M,et al.Multilocus genotypes and broad host-range of Enterocytozoon bieneusi in captive wildlife at zoological gardens in China[J].Parasit Vectors,2016,9:395.
[2]DENG L,LI W,ZHONG Z J,et al.Multi-locus genotypes of Enterocytozoon bieneusi in captive Asiatic black bears in southwestern China:high genetic diversity,broad host range,and zoonotic potential[J].PLoS One,2017,12(2):e0171772.
[3]PRASERTBUN R,MORI H,PINTONG A R,et al.Zoonotic potential of Enterocytozoon genotypes in humans and pigs in Thailand[J].Vet Parasitol,2017,233:73-79.
[4]YUE D M,MA J G,LI F C,et al.Occurrence of Enterocytozoon bieneusi in donkeys(Equus asinus)in China:apublic health concern[J].Front Microbiol,2017,8:565.
[5]TEN HOVE R J,VAN LIESHOUT L,BEADSWORTH M B J,et al.Characterization of genotypes of Enterocytozoon bieneusi in immunosuppressed and immunocompetent patient groups[J].J Eukaryot Microbiol,2009,56(4):388-393.
[6]LIGUORY O,SARFATI C,DEROUIN F,et al.Evidence of different Enterocytozoon bieneusi genotypes in patients with and without human immunodeficiency virus infection[J].J Clin Microbiol,2001,39(7):2672-2674.
[7]SADLER F,PEAKE N,BORROW R,et al.Genotyping of Enterocytozoon bieneusi in AIDS patients from the north west of England[J].J Infect,2002,44(1):39-42.
[8]SANTN M,FAYER R.Microsporidiosis:Enterocytozoon bieneusi in domesticated and wild animals[J].Res Vet Sci,2011,90(3):363-371.
[9]AKINBO F O,OKAKA C E,OMOREGIE R,et al.Molecular epidemiologic characterization of Enterocytozoon bieneusi in HIV-infected persons in Benin City,Nigeria[J].Am J Trop Med Hyg,2012,86(3):441-445.
[10]METGE S,VAN NHIEU J T,DAHMANE D,et al.A case of Enterocytozoon bieneusi infection in an HIV-negative renal transplant recipient[J].Eur J Clin Microbiol Infect Dis,2000,19(3):221-223.
[11]NKININ S W,ASONGANYI T,DIDIER E S,et al.Microsporidian infection is prevalent in healthy people in Cameroon[J].J Clin Microbiol,2007,45(9):2841-2846.
[12]ZHANG X,WANG Z X,SU Y,et al.Identification and genotyping of Enterocytozoon bieneusi in China[J].J Clin Microbiol,2011,49(5):2006-2008.
[13]SAK B,BRADY D,PELIKNOVM,et al.Unapparent microsporidial infection among immunocompetent humans in the Czech Republic[J].J Clin Microbiol,2011,49(3):1064-1070.
[14]WIDMER G,AKIYOSHI D E.Host-specific segregation of ribosomal nucleotide sequence diversity in the microsporidian Enterocytozoon bieneusi[J].Infect Genet Evol,2010,10(1):122-128.
[15]SHI K,LI M J,WANG X X,et al.Molecular survey of Enterocytozoon bieneusi in sheep and goats in China[J].Parasit Vectors,2016,9:23.
[16]SANTN M,FAYER R.Enterocytozoon bieneusi genotype nomenclature based on the internal transcribed spacer sequence:A consensus[J].J Eukaryot Microbiol,2009,56(1):34-38.
[17]FENG Y Y,LI N,DEAREN T,et al.Development of a multilocus sequence typing tool for high-resolution genotyping of Enterocytozoon bieneusi[J].Appl Environ Microbiol,2011,77(14):4822-4828.
[18]NAGPAL A,PRITT B S,LORENZ E C,et al.Disseminated microsporidiosis in a renal transplant recipient:Case report and review of the literature[J].Transpl Infect Dis,2013,15(5):526-532.
[19]DENGJEL B,ZAHLER M,HERMANNS W,et al.Zoonotic potential of Enterocytozoon bieneusi[J].J Clin Microbiol,2001,39(12):4495-4499.
[20]LEE J H.Prevalence and molecular characteristics of Enterocytozoon bieneusi in cattle in Korea[J].Parasitol Res,2007,101(2):391-396.
[21]GHOSH K,WEISS L M.Molecular diagnostic tests for microsporidia[J].Interdiscip Perspect Infect Dis,2009,2009:926521.
[22]BREITENMOSER A C,MATHIS A,BRGI E,et al.High prevalence of Enterocytozoonbieneusi in swine with four genotypes that differ from those identified in humans[J].Parasitology,1999,118(Pt 5):447-453.
[23]FAYER R,SANTIN-DURAN M.Epidemiology of microsporidia in human infections[M]//WEISS L M,BECNEL J J.Microsporidia:Pathogens of Opportunity.UK:John Wiley&Sons,Inc.Press,2014:135-164.
[24]PREZ-LOSADA M,CABEZAS P,CASTRO-NALLAR E,et al.Pathogen typing in the genomics era:MLST and the future of molecular epidemiology[J].Infect Genet Evol,2013,16:38-53.
[25]KARIM M R,DONG H J,YU F C,et al.Genetic diversity in Enterocytozoon bieneusi isolates from dogs and cats in china:Host specificity and public health implications[J].J Clin Microbiol,2014,52(9):3297-3302.
[26]KARIM M R,WANG R J,DONG H J,et al.Genetic polymorphism and zoonotic potential of Enterocytozoon bieneusi from nonhuman primates in China[J].Appl Environ Microbiol,2014,80(6):1893-1898.
[27]LI N,XIAO L H,WANG L,et al.Molecular surveillance of Cryptosporidium spp.,Giardia duodenalis,and Enterocytozoon bieneusi by genotyping and subtyping parasites in wastewater[J].PLoS Negl Trop Dis,2012,6(9):e1809.
[28]LI W,DENG L,YU X M,et al.Multilocus genotypes and broad host-range of Enterocytozoon bieneusi in captive wildlife at zoological gardens in China[J].Parasit Vectors,2016,9:395.