日本血吸虫线粒体基因序列的研究
摘要
[目的]研究云南省三个自然隔离群日本血吸虫线粒体CO1、cytb、ND1、ND6、ND4L基因差异,探讨日本血吸虫种内遗传特点,为阐明我国日本血吸虫的种群遗传结构及进一步研究日本血吸虫生物学特性奠定基础。
[方法]采集云南省大理古城、大理喜洲、永胜三地阳性钉螺及购买江苏省实验室传代阳性钉螺,逸出尾蚴,经腹部攻击感染健康家兔(雄性),每兔感染尾蚴200条～250条,攻击感染后42天剖杀家兔,利用灌注法取成虫。试剂盒抽提基因组DNA,以NC_002544为参考序列设计引物,采用PCR技术对云南三地以及江苏实验室传代的日本血吸虫的CO1、Cytb、tb、ND1、ND6、ND4L全基因进行扩增、测序。用DNAStar软件及Mega3.1软件进行相似性比较和进化树分析。
[结果]分别获得云南三地及江苏省实验室传代日本血吸虫全长CO1、Cytb,ND1、ND6、ND4L基因片段,全长分别为1527bp、1116bp、891bp、459bp、264bp。相似性分析,各序列有微小差异,分别有10、6、6、6、2个碱基位点不同,同源性分别为:99.5%～100.0%、99.6%～100%、99.5%～100.0%、99.1%～100%、99.2%～100%。除ND4L基因外其CO1、Cytb均有一个错义突变,ND1、ND6各有两个错义突变。进化树显示云南三地及江苏省实验室传代日本血吸虫明显聚集在一起,同曼氏血吸虫、埃及血吸虫、湄公血吸虫、马来血吸虫有明显的分支,但与所检获的以往发表的日本血吸虫线粒体相同基因序列比较不构成独立的分支。
[结论]各序列有微小差异,同源性较高;CO1、cytb均有一个错义突变,ND1、ND6各有两个错义突变,ND4L无错义突变;本研究所选云南三地及一江苏省实验室传代的样本均属于日本血吸虫,且亲缘关系密切,无明显分支,不能构成种内分类。
[Objective]To study genomic diversity among the three natural isolates of Schistosoma japonicum in Yunnan province.To investigate intraspecific genetic characteristic about Schistosoma japonicum.To establish foundation for elucidate the population hereditary constitution of Schistosoma japonicum in China and do further study about biologic characteristics of Schistosoma japonicum.
[Methods]Cercariaeum of Schistosoma japonicum were obtained from masccline Oncomelania hupensis from Dali,Xizhou,and Yongsheng in Yunnan Province and one laboratory maintained isolates in Jiangsu Province.Then every male rabbit is infected with 200～250 cercariae of Schisosoma japonicum via abdominal skin. Rabbits were killed on the 42nd after infection.Schistosome japonicurn adults were collected from rabbits through perfusion method.Genomic DNA was isolated with kit, and the complete gene fragments of the mitochondrial cytochrome C oxidase subunit 1(CO1)and cytochrome b(Cytb)and NADH dehydrogenase subunit 1(ND1)and NADH dehydrogenase subunit 6(ND6)and NADH dehydrogenase subunit 4L (ND4L)were amplified by PCR using specific primers and sequenced.Program DNAStar and Mega3.1 were used for sequence analysis.
[Results]The complete gene fragments of the mitochondrial cytochrome C oxidase subunit 1(CO1),cytochrome b,NADH dehydrogenase subunit 1(ND1),NADH dehydrogenase subunit 6(ND6)and NADH dehydrogenase subunit 4L(ND4L)of Schistosoma japonicum amplified from different geographic strains were respectively obtained,the length of complete gene fragments of the mitochondrial cytochrome C oxidase subunit 1(CO1),cytochrome b,NADH dehydrogenase subunit 1(ND1), NADH dehydrogenase subunit 6(ND6)and NADH dehydrogenase subunit 4L (ND4L)of Schistosoma japonicum amplified from different geographic strains were 1527bp,1116bp,891bp,459bp and 264bp,respectively.Comparison of sequences representing the isolates revealed small inter-isolate sequence variations of 10,6,6, 6 and 2 basic group sites,respectively.The identity were 99.5%～100.0%,99.6 %～100%,99.5%～100.0%,99.1%～100%and 99.2%～100%, respectively.There was one missense mutant in the gene fragments of the mitochondrial cytochrome C oxidase subunit 1(CO1)and cytochrome b respectively, and two missense mutant in the gene fragments of the mitochondrial NADH dehydrogenase subunit 1(ND1)and NADH dehydrogenase subunit 6(ND6) respectively,except NADH dehydrogenase subunit 4L(ND4L).The phylogenetic tree demonstrate:Schistosoma japonicum from three natural isolates in Yunnan Province and one laboratory maintained isolates in Jiangsu Province obviously assemble,have obvious branch with Schistosoma mansoni,Schistosoma haematobium,Schistosoma mekongi and Schistosoma malay,but can't be constituted independent branch with the published sequences of the same mitochondrial gene were obtained from GeneBank.
[Conclusion]The sequences have small difference and higher autoploidy;There was one missense mutant in the gene fragments of the mitochondrial cytochrome C oxidase subunit 1(CO1)and cytochrome b,and two missense mutant in the gene fragments of the mitochondrial NADH dehydrogenase subunit 1(ND1)and NADH dehydrogenase subunit 6(ND6),and no missense mutant in the gene fragment of the mitochondrial NADH dehydrogenase subunit 4L(ND4L);The samples still belong to Schistosoma japonicum in this study and have intimate genetic relationship,but no obvious branch,so can't be constituted intraspecies categorization.
[方法]采集云南省大理古城、大理喜洲、永胜三地阳性钉螺及购买江苏省实验室传代阳性钉螺,逸出尾蚴,经腹部攻击感染健康家兔(雄性),每兔感染尾蚴200条～250条,攻击感染后42天剖杀家兔,利用灌注法取成虫。试剂盒抽提基因组DNA,以NC_002544为参考序列设计引物,采用PCR技术对云南三地以及江苏实验室传代的日本血吸虫的CO1、Cytb、tb、ND1、ND6、ND4L全基因进行扩增、测序。用DNAStar软件及Mega3.1软件进行相似性比较和进化树分析。
[结果]分别获得云南三地及江苏省实验室传代日本血吸虫全长CO1、Cytb,ND1、ND6、ND4L基因片段,全长分别为1527bp、1116bp、891bp、459bp、264bp。相似性分析,各序列有微小差异,分别有10、6、6、6、2个碱基位点不同,同源性分别为:99.5%～100.0%、99.6%～100%、99.5%～100.0%、99.1%～100%、99.2%～100%。除ND4L基因外其CO1、Cytb均有一个错义突变,ND1、ND6各有两个错义突变。进化树显示云南三地及江苏省实验室传代日本血吸虫明显聚集在一起,同曼氏血吸虫、埃及血吸虫、湄公血吸虫、马来血吸虫有明显的分支,但与所检获的以往发表的日本血吸虫线粒体相同基因序列比较不构成独立的分支。
[结论]各序列有微小差异,同源性较高;CO1、cytb均有一个错义突变,ND1、ND6各有两个错义突变,ND4L无错义突变;本研究所选云南三地及一江苏省实验室传代的样本均属于日本血吸虫,且亲缘关系密切,无明显分支,不能构成种内分类。
[Objective]To study genomic diversity among the three natural isolates of Schistosoma japonicum in Yunnan province.To investigate intraspecific genetic characteristic about Schistosoma japonicum.To establish foundation for elucidate the population hereditary constitution of Schistosoma japonicum in China and do further study about biologic characteristics of Schistosoma japonicum.
[Methods]Cercariaeum of Schistosoma japonicum were obtained from masccline Oncomelania hupensis from Dali,Xizhou,and Yongsheng in Yunnan Province and one laboratory maintained isolates in Jiangsu Province.Then every male rabbit is infected with 200～250 cercariae of Schisosoma japonicum via abdominal skin. Rabbits were killed on the 42nd after infection.Schistosome japonicurn adults were collected from rabbits through perfusion method.Genomic DNA was isolated with kit, and the complete gene fragments of the mitochondrial cytochrome C oxidase subunit 1(CO1)and cytochrome b(Cytb)and NADH dehydrogenase subunit 1(ND1)and NADH dehydrogenase subunit 6(ND6)and NADH dehydrogenase subunit 4L (ND4L)were amplified by PCR using specific primers and sequenced.Program DNAStar and Mega3.1 were used for sequence analysis.
[Results]The complete gene fragments of the mitochondrial cytochrome C oxidase subunit 1(CO1),cytochrome b,NADH dehydrogenase subunit 1(ND1),NADH dehydrogenase subunit 6(ND6)and NADH dehydrogenase subunit 4L(ND4L)of Schistosoma japonicum amplified from different geographic strains were respectively obtained,the length of complete gene fragments of the mitochondrial cytochrome C oxidase subunit 1(CO1),cytochrome b,NADH dehydrogenase subunit 1(ND1), NADH dehydrogenase subunit 6(ND6)and NADH dehydrogenase subunit 4L (ND4L)of Schistosoma japonicum amplified from different geographic strains were 1527bp,1116bp,891bp,459bp and 264bp,respectively.Comparison of sequences representing the isolates revealed small inter-isolate sequence variations of 10,6,6, 6 and 2 basic group sites,respectively.The identity were 99.5%～100.0%,99.6 %～100%,99.5%～100.0%,99.1%～100%and 99.2%～100%, respectively.There was one missense mutant in the gene fragments of the mitochondrial cytochrome C oxidase subunit 1(CO1)and cytochrome b respectively, and two missense mutant in the gene fragments of the mitochondrial NADH dehydrogenase subunit 1(ND1)and NADH dehydrogenase subunit 6(ND6) respectively,except NADH dehydrogenase subunit 4L(ND4L).The phylogenetic tree demonstrate:Schistosoma japonicum from three natural isolates in Yunnan Province and one laboratory maintained isolates in Jiangsu Province obviously assemble,have obvious branch with Schistosoma mansoni,Schistosoma haematobium,Schistosoma mekongi and Schistosoma malay,but can't be constituted independent branch with the published sequences of the same mitochondrial gene were obtained from GeneBank.
[Conclusion]The sequences have small difference and higher autoploidy;There was one missense mutant in the gene fragments of the mitochondrial cytochrome C oxidase subunit 1(CO1)and cytochrome b,and two missense mutant in the gene fragments of the mitochondrial NADH dehydrogenase subunit 1(ND1)and NADH dehydrogenase subunit 6(ND6),and no missense mutant in the gene fragment of the mitochondrial NADH dehydrogenase subunit 4L(ND4L);The samples still belong to Schistosoma japonicum in this study and have intimate genetic relationship,but no obvious branch,so can't be constituted intraspecies categorization.
引文
[1]汪世萍主编.医学寄生虫学[M].北京:高等教育出版社,2004:121-122.
[2]王陇德.纪念血吸虫病在中国发现100周年[M].2005,10,28.
[3]赵慰先.人体寄生虫学[M].2版.北京:人民卫生出版社,1992:326.
[4]Saccone S,Giorgi CD,Gissi C,et al.Evolutionary genomics in Metazoa:the mitochondrial DNA as a model system[J].Gene,1999,238:195-209.
[5]肖武汉.银鲴自然群体线粒体DNA的遗传分化[J].水生生物学报,2000,24(1):1-9.
[6]王存芳,曾庆勇,杜立新,等.猪线粒体DNA(mtDNA)及其在遗传育种中的应用[J].黑龙江畜牧兽医,2001,(9):5-7.
[7]胡文革,王金富.动物线粒体DNA多态性研究及其在鱼类群体遗传结构上的应用[J].石河子大学学报,2001,5(3):253-258.
[8]徐庆刚,花保祯.线粒体DNA在昆虫系统学研究中的应用[J].西北农林科技大学学报(自然科学版),2001,29(增):79-83.
[9]张方,米志勇.动物线粒体DNA的分子生物学研究进展[J].生物工程进展,1998,18(3):25-31.
[10]夏德全,王文君.动物线粒体DNA研究及在鱼类种群遗传结构研究中的应用[J].水产学报,1998,22(4):364-370.
[11]苟德明,王智新.动物线粒体DNA特性及其多态性研究进展[J].遗传育种·黄牛杂志,1999,22(4):7-10
[12]Boore JL.Animal mitochondrial genomes[J].Nucleic Acids Res 1999,27:1767-1780.
[13]Telford,MJ.,Hemiou,EA.,Russell,R.B.,Littlwood,D.T.J.,2000.Changes in mitochondrial genetic codes as phylogenetic characters:two examples from the flatworms[J].Proc.Natl.Acad.Sci.USA,97,11359-1364.
[14]Le,T.H.,Blair,D.,McManus,D.P.,2001b.Complete DNA sequence and gene organizationof the mitochondrial genome of the liverfluke,Fasciola hepatioa L.,(Platyhelminthes;Trematoda)[J].Parasitology 123,609-621.
[15]Le,T.H.,Pearson,M.S.,Blair,D.,Dsi,N.,Zhang,L.H.,McManus,D.E,2002b.Complete mitochondrial genomes confirm the distinctiveness of the horse-dog and sheep-dog strains of Echinococcus granulosus[J].Parasitology 24,97-112.
[16]Thanh H,Le,David Blair,et al.Phylogenies inferred from mitochondrial gene orders-A cautionary tale from the parasitic flatworms[J].Mol.Biol.Evol,2000;17(7):1123-1125.
[17]Thanh H,Le,Pierre-Francois Humair,et al.Mitochondrial gene content,arrangement and composition compared in African and Asian schistosomes[J].Mol.Biol.Evol.2001;11(7):61-71.
[18]Iwin DW,Kocher TD,Wilson AC.Evolution of the cytochrome b gene of mammals[J].J Mol Evol,1991,32:128-144.
[19]Esposti MD,Vries SD,Crimi M,Chelli A,Patamello T,Meyer A.Mitochondrial cytochrome b:evolution and structure of the protein[J].Biochem Biophys Acta,1993,143:243-271.
[20]李伟,杨毅梅,等.日本血吸虫成虫灌注收集法的改进.大理学院学报.2005,4(5).
[21]Fletcher M,Woodruff DS,Loverde PT,et al.Genetic differentiation between Schistosoma mekongi and S.japonicum:An electrophoretic study[J].Malacol Rev,1980;Supp 12:113-122.
[22]Woodruff DS,Merenlender,AM Upatham ES,et al.Genetic variation and differentiation of three Schistosoma specites from the Philippines,Laos and Peninsular Malaysia[J].AM J Trop Med Hyg,1987,36:345-354.
[23]Merenlender AM,Woodruff DS,Upatham ES,et al.Large genetic distance between Chinese and Philippine Schistosoma japonicum[J].J Parasitol,1987,73:861-863.
[24]俞小淙,吴观陵,等.血吸虫遗传变异的研究进展[J].中国寄生虫学与寄生虫病杂志.1999,17(3):184-187.
[25]何毅勋.中国大陆日本血吸虫品系的研究-Ⅻ总结[J].中国寄生虫学与寄生虫病杂志,1993,11(2):93-97.
[26]谢觅,何毅勋,裘丽株,等.我国与日本山梨不同地域品系日本血吸虫DNA的限制性片段长度多态性分析[J].中国寄生虫学与寄生虫病杂志,1997,15(4):216-219.
[27]冯友仁,牛安欧,方正明.血吸虫种株随机扩增多态性DNA差异研究[J].中国人兽共 患病杂志,2001,17(4):49-53.
[28]Robin BG,Qian BZ,Peter N,et al.Use of RAPD for the detection of genetic variation in the human blood fluke,Schistosoma japonicum,from mainland China[J].Mol Cell Probe,1996,10:353-358.
[29]Rubinsztein DC,Amos W,Leggo J,et al.Microsatellite evolution-evidence for directionality and variation in rate between specie[J].Nat Genet,1995,11:360-362.
[30]何平.真核生物中的微卫星及其应用[J].遗传,1998,20(4):42-47.
[31]Shrivastava J,Gower CM,Balolong EJr,et al.Population genetics of multi-host parasites-the case for molecular epidemiological studies of Schistosoma japonicum using larval stages from naturally infected hosts[J].Parasitology,2005,131:617-626.
[32]郭凯文,牛安欧.日本血吸虫线粒体DNA两个分子的遗传变异[J].中国寄生虫学与寄生虫病杂志,2004,22(5):300-302.
[33]郭凯文,牛安欧.日本血吸虫线粒体DNA的遗传变异[J].热带医学杂志,2004,4(4):357-360.
[34]郭凯文,牛安欧.PCR-SSCP研究中国日本血吸虫线粒体DNA的遗传变异[J].中国人兽共患病杂志,2004,20(6):529-532.
[35]Bogh HO,Zhu XQ,et al.Scanning for nucleotide variations in mitochondrial DNA fragments of Schistosoma japonicum by single-strand conformation polymorphism [J].Parasitology,1999,118,73-82.
[36]邱持平,Chris Spolsky,夏明仪,等.中国日本血吸虫地域株基因差异的研究[J].中国寄生虫学与寄生虫病杂志.2002,20(2):79-82.
[1]陈名刚.世界血吸虫病流行情况及防治进展[J].中国血吸虫病防治杂志,2002,2:5-7.
[2]李雍龙.人体寄生虫学[第六版].人民卫生出版社.
[3]汪世萍主编.医学寄生虫学[M].北京:高等教育出版社,2004:121-122.
[4]王陇德.纪念血吸虫病在中国发现100周年[M].2005,10,28:
[5]Rollinson D,Kaukas A,Johnston DA,Simpson AJG,Tanaka M.Some molecular insights into schistosome evolution[J].Int J Parasitol 1997,27:11-28.
[6]Avise JC,Wollenberg K.Phylogenetics and the origin of species[J].Proc Natl Acad Sci USA 1997,94:7748-7755.
[7]Saccone S,Giorgi CD,Gissi C,et al.Evolutionary genomics m Metazoa:the mitochondrial DNAasamodelsystem[J].Gene,1999,238:195-209.
[8]胡文革,王金富.动物线粒体DNA多态性研究及其在鱼类群体遗传结构上的应用[J].石河子大学学报,2001,5(3):253-258.
[9]徐庆刚,花保祯.线粒体DNA在昆虫系统学研究中的应用[J].西北农林科技大学学报(自然科学版),2001,29(增):79-83.
[10]肖武汉.银鲴自然群体线粒体DNA的遗传分化[J].水生生物学报,2000,24(1):1-9.
[11]王存芳,曾庆勇,杜立新,等.猪线粒体DNA(mtDNA)及其在遗传育种中的应用[J].黑龙江畜牧兽医,2001,(9):5-7.
[12]张方,米志勇.动物线粒体DNA的分子生物学研究进展[J].生物工程进展,1998,18(3):25-31.
[13]夏德全,王文君.动物线粒体DNA研究及在鱼类种群遗传结构研究中的应用[J].水产学报,1998,22(4):364-370.
[14]苟德明,王智新.动物线粒体DNA特性及其多态性研究进展[J].遗传育种黄牛杂志,1999,22(4):7-10.
[15]Boore JL.Animal mitochondrial genomes[J].Nucleic Acids Res 1999,27:1767-1780.
[16]Telford,MJ.,Hemiou,EA.,Russell,R.B.,Littlwood,D.T.J.,2000..Changes in mitochondrial genetic codes as phylogenetic characters:two examples from the flatworms. Proc. Natl. Acad. Sci.USA, 97, 11359-11364.
[17] Le, T.H., Blair, D., McManus, D.P., 2001b. Complete DNA sequence and gene organizationof the mitochondrial genome of the liverfluke.Fasciola hepatioa L. , (Platyhelminthes;Trematoda) . Parasitology 123, 609-621.
[18] Le, T.H., Pearson, M.S., Blair, D., Dsi, N., Zhang, L.H., McManus, D.P., 2002b. Complete mitochondrial genomes confirm the distinctiveness of the horse-dog and sheep-dog strains of Echinococcus granulosus[J]. Parasitology, 24, 97-112.
[19] Skelly PJ, Stein LD, Shoemaker CB. Expression of Schistosoma mansoni genes involved in anaerobic and oxidative glucose metabolism during the cercaria to adult transformation[J]. Mol Biochem Parasitol, 1993; 60: 93-104.
[20] Franco GR, Adams MD, Soares MB, Simpson AJ, Venter JC, Pena SD. Identification of new Schistosoma mansoni genes by the EST strategy using a directional cDNA library[J]. Gene, 1995; 152: 141 - 147.
[21] Bowles J, Hope M, Tiu WU, Liu X, Mcmanus DP. Nuclear and mitochondrial genetic markers highly conserved between Chinese and Philipine Schistosoma japonicum [J]. Acta Trop, 1993; 55: 217-229.
[22] Bowles J, Blair D, McManus DP. A molecular phylogeny of the human schiatosomes[J]. Mol PhylogenetEvol, 1995; 4: 103-109.
[23] Sorensen E, Drew AC, Brindley PJ, et al. Variation in the sequence of a mitochondrial NADH dehydrogenase 1 gene fragment among six natural populations of Schistosoma japonicum from China[J]. Int J Parasitol, 1998; 28: 1931 - 1934.
[24] Sorensen E, Bogh HO, Johansen MV,McManus DP. PCR-based identification of individuals of Schistosoma japonicum representing different subpopulations using a genetic marker in mitochondrial DNA[J]. Int Parasitol 1999; 29: 1121 - 128.
[25] Thanh H, Le, David Blair, et al. Phylogenies inferred from mitochondrial gene orders-A cautionary tale from the parasitic flatworms[J]. Mol.Biol.Evol.2000; 17(7): 1123-1125.
[26] Thanh H.Le, Pierre-Francois Humair, et al. Mitochondrial gene content, arrangement and composition compared in African and Asian schistosomes[J]. Mol.Biol.Evol.2001: 11(7): 61 -71.
[27]Despre's L,Imbert-Establet D,Combes C,Bonhomme F,Monnerot M.Isolation and polymorphism in mitochondrial DNA from schistosoma mansoni[J].Mol Biochem Parasitol,1991;47:139-142.
[28]Blair D,Ver Herwerden L,Hirai H et al.Relationship between Schistosoma malayensis and other Asian schistosomes deduced from DNA sequences[J].Mol Biochem Parasitol,1997;85:259-263.
[29]Bogh HO,Zhu XQ,et al.Scanning for nucleotide variations in mitochondrial DNA fragments of Schistosoma japonicum by single-strand conformation polymorphism[J].Parasitology,1999,118,73-82.
[30]Zhu,XQ,Henrik Bogh,et al.Dideoxy fingerprinting of low-level nucleofide variation in mitochondrial DNA of the human blood fluke,Schistosoma japonicum[J].Electrophoresis,1999,20:2830-2833.
[31]T.H.LE,D.BLAIR,D.P.McMANUS.Revisiting the question of limited genetic variation within Schistosoma japonicum[J].Annals of Tropical Medicine Parasitology,2002,96(2):155-164.
[32]邱持平,Chris Spolsky等.中国日本血吸虫地域株基因差异的研究[J].中国寄生虫学与寄生虫病杂志,2002,20(2):79-82.
[33]Kane RA,Southgate VR,Rollinson D,et al.A phylogeny based on three mitochondrial genes supports the division of Schistosoma intercalatum into two separate species[J].Parasitology,2003,127(2):131-137.
[34]刘伟,朱兴全,等.湖南省日本血吸虫线粒体nad4基因的多态性研究[J].论文摘要·吸虫和绦虫.
[35]郭凯文,牛安欧.PCR-SSCP研究中国日本血吸虫线粒体DNA的遗传变异[J].中国人兽共患病杂志,2004,20(6):529-532.
[36]郭凯文,牛安欧.日本血吸虫线粒体DNA的遗传变异[J].热带医学杂志,2004,4(4):357-360.
[37]谢觅,何毅勋.我国与日本山梨地区不同地域品系日本血吸虫DNA限制性片段长度多态性分析[J].中国寄生虫学与寄生虫病杂志,1997;15(4):321-324.
[38]冯友仁,牛安欧,方正明,等.血吸虫种株随机扩增多态性DNA差异研究[J].中国人 兽共患病杂志,2001;17(4):48-52.
[39]熊衍文,牛安欧.微卫星锚定PCR研究日本血吸虫的遗传变异[J].中国人兽共患病杂志,2002;28(6):7-9.
[40]郭凯文,牛安欧.日本血吸虫线粒体DNA的遗传变异[J].热带医学杂志.2004,4(4):357-360.
[41]郭凯文,牛安欧.日本血吸虫线粒体DNA两个分子的遗传变异[J].中国寄生虫学与寄生虫病杂志,2004,22(5):300-302.
[42]刘伟,戴荣四,等.湖南省日本血吸虫线粒体coxl基因部分序列多态性的研究[J].热带医学杂志,2007,7(10):939-942.
[2]王陇德.纪念血吸虫病在中国发现100周年[M].2005,10,28.
[3]赵慰先.人体寄生虫学[M].2版.北京:人民卫生出版社,1992:326.
[4]Saccone S,Giorgi CD,Gissi C,et al.Evolutionary genomics in Metazoa:the mitochondrial DNA as a model system[J].Gene,1999,238:195-209.
[5]肖武汉.银鲴自然群体线粒体DNA的遗传分化[J].水生生物学报,2000,24(1):1-9.
[6]王存芳,曾庆勇,杜立新,等.猪线粒体DNA(mtDNA)及其在遗传育种中的应用[J].黑龙江畜牧兽医,2001,(9):5-7.
[7]胡文革,王金富.动物线粒体DNA多态性研究及其在鱼类群体遗传结构上的应用[J].石河子大学学报,2001,5(3):253-258.
[8]徐庆刚,花保祯.线粒体DNA在昆虫系统学研究中的应用[J].西北农林科技大学学报(自然科学版),2001,29(增):79-83.
[9]张方,米志勇.动物线粒体DNA的分子生物学研究进展[J].生物工程进展,1998,18(3):25-31.
[10]夏德全,王文君.动物线粒体DNA研究及在鱼类种群遗传结构研究中的应用[J].水产学报,1998,22(4):364-370.
[11]苟德明,王智新.动物线粒体DNA特性及其多态性研究进展[J].遗传育种·黄牛杂志,1999,22(4):7-10
[12]Boore JL.Animal mitochondrial genomes[J].Nucleic Acids Res 1999,27:1767-1780.
[13]Telford,MJ.,Hemiou,EA.,Russell,R.B.,Littlwood,D.T.J.,2000.Changes in mitochondrial genetic codes as phylogenetic characters:two examples from the flatworms[J].Proc.Natl.Acad.Sci.USA,97,11359-1364.
[14]Le,T.H.,Blair,D.,McManus,D.P.,2001b.Complete DNA sequence and gene organizationof the mitochondrial genome of the liverfluke,Fasciola hepatioa L.,(Platyhelminthes;Trematoda)[J].Parasitology 123,609-621.
[15]Le,T.H.,Pearson,M.S.,Blair,D.,Dsi,N.,Zhang,L.H.,McManus,D.E,2002b.Complete mitochondrial genomes confirm the distinctiveness of the horse-dog and sheep-dog strains of Echinococcus granulosus[J].Parasitology 24,97-112.
[16]Thanh H,Le,David Blair,et al.Phylogenies inferred from mitochondrial gene orders-A cautionary tale from the parasitic flatworms[J].Mol.Biol.Evol,2000;17(7):1123-1125.
[17]Thanh H,Le,Pierre-Francois Humair,et al.Mitochondrial gene content,arrangement and composition compared in African and Asian schistosomes[J].Mol.Biol.Evol.2001;11(7):61-71.
[18]Iwin DW,Kocher TD,Wilson AC.Evolution of the cytochrome b gene of mammals[J].J Mol Evol,1991,32:128-144.
[19]Esposti MD,Vries SD,Crimi M,Chelli A,Patamello T,Meyer A.Mitochondrial cytochrome b:evolution and structure of the protein[J].Biochem Biophys Acta,1993,143:243-271.
[20]李伟,杨毅梅,等.日本血吸虫成虫灌注收集法的改进.大理学院学报.2005,4(5).
[21]Fletcher M,Woodruff DS,Loverde PT,et al.Genetic differentiation between Schistosoma mekongi and S.japonicum:An electrophoretic study[J].Malacol Rev,1980;Supp 12:113-122.
[22]Woodruff DS,Merenlender,AM Upatham ES,et al.Genetic variation and differentiation of three Schistosoma specites from the Philippines,Laos and Peninsular Malaysia[J].AM J Trop Med Hyg,1987,36:345-354.
[23]Merenlender AM,Woodruff DS,Upatham ES,et al.Large genetic distance between Chinese and Philippine Schistosoma japonicum[J].J Parasitol,1987,73:861-863.
[24]俞小淙,吴观陵,等.血吸虫遗传变异的研究进展[J].中国寄生虫学与寄生虫病杂志.1999,17(3):184-187.
[25]何毅勋.中国大陆日本血吸虫品系的研究-Ⅻ总结[J].中国寄生虫学与寄生虫病杂志,1993,11(2):93-97.
[26]谢觅,何毅勋,裘丽株,等.我国与日本山梨不同地域品系日本血吸虫DNA的限制性片段长度多态性分析[J].中国寄生虫学与寄生虫病杂志,1997,15(4):216-219.
[27]冯友仁,牛安欧,方正明.血吸虫种株随机扩增多态性DNA差异研究[J].中国人兽共 患病杂志,2001,17(4):49-53.
[28]Robin BG,Qian BZ,Peter N,et al.Use of RAPD for the detection of genetic variation in the human blood fluke,Schistosoma japonicum,from mainland China[J].Mol Cell Probe,1996,10:353-358.
[29]Rubinsztein DC,Amos W,Leggo J,et al.Microsatellite evolution-evidence for directionality and variation in rate between specie[J].Nat Genet,1995,11:360-362.
[30]何平.真核生物中的微卫星及其应用[J].遗传,1998,20(4):42-47.
[31]Shrivastava J,Gower CM,Balolong EJr,et al.Population genetics of multi-host parasites-the case for molecular epidemiological studies of Schistosoma japonicum using larval stages from naturally infected hosts[J].Parasitology,2005,131:617-626.
[32]郭凯文,牛安欧.日本血吸虫线粒体DNA两个分子的遗传变异[J].中国寄生虫学与寄生虫病杂志,2004,22(5):300-302.
[33]郭凯文,牛安欧.日本血吸虫线粒体DNA的遗传变异[J].热带医学杂志,2004,4(4):357-360.
[34]郭凯文,牛安欧.PCR-SSCP研究中国日本血吸虫线粒体DNA的遗传变异[J].中国人兽共患病杂志,2004,20(6):529-532.
[35]Bogh HO,Zhu XQ,et al.Scanning for nucleotide variations in mitochondrial DNA fragments of Schistosoma japonicum by single-strand conformation polymorphism [J].Parasitology,1999,118,73-82.
[36]邱持平,Chris Spolsky,夏明仪,等.中国日本血吸虫地域株基因差异的研究[J].中国寄生虫学与寄生虫病杂志.2002,20(2):79-82.
[1]陈名刚.世界血吸虫病流行情况及防治进展[J].中国血吸虫病防治杂志,2002,2:5-7.
[2]李雍龙.人体寄生虫学[第六版].人民卫生出版社.
[3]汪世萍主编.医学寄生虫学[M].北京:高等教育出版社,2004:121-122.
[4]王陇德.纪念血吸虫病在中国发现100周年[M].2005,10,28:
[5]Rollinson D,Kaukas A,Johnston DA,Simpson AJG,Tanaka M.Some molecular insights into schistosome evolution[J].Int J Parasitol 1997,27:11-28.
[6]Avise JC,Wollenberg K.Phylogenetics and the origin of species[J].Proc Natl Acad Sci USA 1997,94:7748-7755.
[7]Saccone S,Giorgi CD,Gissi C,et al.Evolutionary genomics m Metazoa:the mitochondrial DNAasamodelsystem[J].Gene,1999,238:195-209.
[8]胡文革,王金富.动物线粒体DNA多态性研究及其在鱼类群体遗传结构上的应用[J].石河子大学学报,2001,5(3):253-258.
[9]徐庆刚,花保祯.线粒体DNA在昆虫系统学研究中的应用[J].西北农林科技大学学报(自然科学版),2001,29(增):79-83.
[10]肖武汉.银鲴自然群体线粒体DNA的遗传分化[J].水生生物学报,2000,24(1):1-9.
[11]王存芳,曾庆勇,杜立新,等.猪线粒体DNA(mtDNA)及其在遗传育种中的应用[J].黑龙江畜牧兽医,2001,(9):5-7.
[12]张方,米志勇.动物线粒体DNA的分子生物学研究进展[J].生物工程进展,1998,18(3):25-31.
[13]夏德全,王文君.动物线粒体DNA研究及在鱼类种群遗传结构研究中的应用[J].水产学报,1998,22(4):364-370.
[14]苟德明,王智新.动物线粒体DNA特性及其多态性研究进展[J].遗传育种黄牛杂志,1999,22(4):7-10.
[15]Boore JL.Animal mitochondrial genomes[J].Nucleic Acids Res 1999,27:1767-1780.
[16]Telford,MJ.,Hemiou,EA.,Russell,R.B.,Littlwood,D.T.J.,2000..Changes in mitochondrial genetic codes as phylogenetic characters:two examples from the flatworms. Proc. Natl. Acad. Sci.USA, 97, 11359-11364.
[17] Le, T.H., Blair, D., McManus, D.P., 2001b. Complete DNA sequence and gene organizationof the mitochondrial genome of the liverfluke.Fasciola hepatioa L. , (Platyhelminthes;Trematoda) . Parasitology 123, 609-621.
[18] Le, T.H., Pearson, M.S., Blair, D., Dsi, N., Zhang, L.H., McManus, D.P., 2002b. Complete mitochondrial genomes confirm the distinctiveness of the horse-dog and sheep-dog strains of Echinococcus granulosus[J]. Parasitology, 24, 97-112.
[19] Skelly PJ, Stein LD, Shoemaker CB. Expression of Schistosoma mansoni genes involved in anaerobic and oxidative glucose metabolism during the cercaria to adult transformation[J]. Mol Biochem Parasitol, 1993; 60: 93-104.
[20] Franco GR, Adams MD, Soares MB, Simpson AJ, Venter JC, Pena SD. Identification of new Schistosoma mansoni genes by the EST strategy using a directional cDNA library[J]. Gene, 1995; 152: 141 - 147.
[21] Bowles J, Hope M, Tiu WU, Liu X, Mcmanus DP. Nuclear and mitochondrial genetic markers highly conserved between Chinese and Philipine Schistosoma japonicum [J]. Acta Trop, 1993; 55: 217-229.
[22] Bowles J, Blair D, McManus DP. A molecular phylogeny of the human schiatosomes[J]. Mol PhylogenetEvol, 1995; 4: 103-109.
[23] Sorensen E, Drew AC, Brindley PJ, et al. Variation in the sequence of a mitochondrial NADH dehydrogenase 1 gene fragment among six natural populations of Schistosoma japonicum from China[J]. Int J Parasitol, 1998; 28: 1931 - 1934.
[24] Sorensen E, Bogh HO, Johansen MV,McManus DP. PCR-based identification of individuals of Schistosoma japonicum representing different subpopulations using a genetic marker in mitochondrial DNA[J]. Int Parasitol 1999; 29: 1121 - 128.
[25] Thanh H, Le, David Blair, et al. Phylogenies inferred from mitochondrial gene orders-A cautionary tale from the parasitic flatworms[J]. Mol.Biol.Evol.2000; 17(7): 1123-1125.
[26] Thanh H.Le, Pierre-Francois Humair, et al. Mitochondrial gene content, arrangement and composition compared in African and Asian schistosomes[J]. Mol.Biol.Evol.2001: 11(7): 61 -71.
[27]Despre's L,Imbert-Establet D,Combes C,Bonhomme F,Monnerot M.Isolation and polymorphism in mitochondrial DNA from schistosoma mansoni[J].Mol Biochem Parasitol,1991;47:139-142.
[28]Blair D,Ver Herwerden L,Hirai H et al.Relationship between Schistosoma malayensis and other Asian schistosomes deduced from DNA sequences[J].Mol Biochem Parasitol,1997;85:259-263.
[29]Bogh HO,Zhu XQ,et al.Scanning for nucleotide variations in mitochondrial DNA fragments of Schistosoma japonicum by single-strand conformation polymorphism[J].Parasitology,1999,118,73-82.
[30]Zhu,XQ,Henrik Bogh,et al.Dideoxy fingerprinting of low-level nucleofide variation in mitochondrial DNA of the human blood fluke,Schistosoma japonicum[J].Electrophoresis,1999,20:2830-2833.
[31]T.H.LE,D.BLAIR,D.P.McMANUS.Revisiting the question of limited genetic variation within Schistosoma japonicum[J].Annals of Tropical Medicine Parasitology,2002,96(2):155-164.
[32]邱持平,Chris Spolsky等.中国日本血吸虫地域株基因差异的研究[J].中国寄生虫学与寄生虫病杂志,2002,20(2):79-82.
[33]Kane RA,Southgate VR,Rollinson D,et al.A phylogeny based on three mitochondrial genes supports the division of Schistosoma intercalatum into two separate species[J].Parasitology,2003,127(2):131-137.
[34]刘伟,朱兴全,等.湖南省日本血吸虫线粒体nad4基因的多态性研究[J].论文摘要·吸虫和绦虫.
[35]郭凯文,牛安欧.PCR-SSCP研究中国日本血吸虫线粒体DNA的遗传变异[J].中国人兽共患病杂志,2004,20(6):529-532.
[36]郭凯文,牛安欧.日本血吸虫线粒体DNA的遗传变异[J].热带医学杂志,2004,4(4):357-360.
[37]谢觅,何毅勋.我国与日本山梨地区不同地域品系日本血吸虫DNA限制性片段长度多态性分析[J].中国寄生虫学与寄生虫病杂志,1997;15(4):321-324.
[38]冯友仁,牛安欧,方正明,等.血吸虫种株随机扩增多态性DNA差异研究[J].中国人 兽共患病杂志,2001;17(4):48-52.
[39]熊衍文,牛安欧.微卫星锚定PCR研究日本血吸虫的遗传变异[J].中国人兽共患病杂志,2002;28(6):7-9.
[40]郭凯文,牛安欧.日本血吸虫线粒体DNA的遗传变异[J].热带医学杂志.2004,4(4):357-360.
[41]郭凯文,牛安欧.日本血吸虫线粒体DNA两个分子的遗传变异[J].中国寄生虫学与寄生虫病杂志,2004,22(5):300-302.
[42]刘伟,戴荣四,等.湖南省日本血吸虫线粒体coxl基因部分序列多态性的研究[J].热带医学杂志,2007,7(10):939-942.