上海地区不同动物源隐孢子虫种型鉴定及微小隐孢子虫CP15和P23基因的克隆和表达
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摘要
隐孢子虫是一种严重危害人和动物健康的寄生原虫,广泛寄生于人、哺乳动物、鸟类、两栖类和鱼类的消化道和呼吸道中,引起腹泻等症状。对免疫功能正常的动物,腹泻可自限,但免疫功能缺陷时,常表现水泻不止甚至死亡。
隐孢子虫不同种性状有别,寄生宿主的对象不一。本研究利用PCR-RFLP技术成功鉴定了本实验室收集的上海地区分离的鼠源、兔源、鸡源、猪源、牛源隐孢子虫虫种和基因型,共有3个虫种和3个基因型,其中鼠源隐孢子虫虫种为微小隐孢子虫鼠基因型(C.parvum Mouse genotype);兔源隐孢子虫虫种为微小隐孢子虫兔基因型(C.parvum Rabbit genotype);猪源隐孢子虫虫种为微小隐孢子虫猪基因型II(C.parvum Pig genotype II);鸡源隐孢子虫虫种为贝氏隐孢子虫(C.baileyi);牛源隐孢子虫虫种为安氏隐孢子虫(C.andersoni)。
成功克隆了上海地区分离的微小隐孢子虫鼠基因型、兔基因型、猪基因型II和安氏隐孢子虫的子孢子表面抗原CP15基因,其编码基因为大小为357bp,编码119个氨基酸残基,不同来源的CP15基因序列高度一致。其中安氏隐孢子虫的CP15基因成功克隆为本研究首次报道。
成功克隆了上海地区分离的微小隐孢子虫鼠基因型、兔基因型和猪基因型II的子孢子表面抗原P23基因,大小为342bp,编码113个氨基酸,不同来源的P23基因序列高度一致。成功构建了微小隐孢子虫鼠基因型的原核重组表达质粒pET-28a(+)-CP15和pET-28a(+)—P23,实现了CP15和P23基因在大肠杆菌中的表达。ELISA初步分析表明CP15和P23基因具有良好的抗原性。
上述工作的进展,为逐步查明上海地区目前流行的隐孢子虫病原种类、掌握隐孢子虫病流行状况和特点提供了基础,具有重要的公共卫生学意义。隐孢子虫子孢子表面抗原CP15和P23基因的成功克隆和表达,为进一步利用基因的表达产物进行免疫预防和诊断研究提供了条件。
Cryptosporidium spp. is a protozoan parasite and it is harmful severely to men and animals. It grows in epithelial cells and digestive organs of men, animals, birds, creeperes or fishes and it causes diarrhea. Cryptosporidium spp. is known to be ubiquitous and to have many hosts. The diarrhea can be cure by themselves about the normal immune people, but when people who had immune deficiency, the diarrhea maybe serious even take to death.
There were different characteristics and hostes about Cryptosporidium spp. The species and genotypes of Cryptosporidium spp. isolated from mouse, rabbit, chicken, pig and dairy calves in Shanghai district were identified by nested polymerase chain reaction linked restriction fragment length polymorphism (PCR-RFLP). The results showed that there were three species and three genotypes, Cryptosporidium spp. which comes from mouse was Cryptosporidium paravum mouse genotype, rabbit was Cryptosporidium paravum rabbit genotype, chicken was Cryptosporidium baileyi, swine was Cryptosporidium paravum pig genotype II and dairy calves was Cryptospori—dium andersoni.
The sporozoite surface antigen CP15 gene which came from C.paravum mouse genotype, C. paravum rabbit genotype, C. paravum pig genotype II and C. andersoni were successful amplified and encoded 119 amino acids. Cryptosporidium spp. came from different sources had a highly homoely and C. andersoni was first report in this expriment.
The sporozoite surface antigen P23 gene which came from C. paravum mouse genotype, C. paravum rabbit genotype and C.paravum pig genotype II were successful amplified and encoded 113 amino acids. Cryptosporidium spp. came from different sources had a highly homoely. The prokaryotic expressing vector pET-28a(+)-CP15 and pET-28a(+)-P23 were constructed and expressed in E.coli successfuliy. Both of them had a well antigenicity which analyzed by ELISA.
The development of our experiments we had done would much helpful to know the disease sources and epidemic conditions about Cryptosporidium spp. in Shanghai district, and had an important sense about public hygieology. The sporozoite surface antigen CP15 and P23 gene which were successful cloned might be much helpful for the next work of using these genes to develop a immunoprophylaxis or detection technique.
隐孢子虫不同种性状有别,寄生宿主的对象不一。本研究利用PCR-RFLP技术成功鉴定了本实验室收集的上海地区分离的鼠源、兔源、鸡源、猪源、牛源隐孢子虫虫种和基因型,共有3个虫种和3个基因型,其中鼠源隐孢子虫虫种为微小隐孢子虫鼠基因型(C.parvum Mouse genotype);兔源隐孢子虫虫种为微小隐孢子虫兔基因型(C.parvum Rabbit genotype);猪源隐孢子虫虫种为微小隐孢子虫猪基因型II(C.parvum Pig genotype II);鸡源隐孢子虫虫种为贝氏隐孢子虫(C.baileyi);牛源隐孢子虫虫种为安氏隐孢子虫(C.andersoni)。
成功克隆了上海地区分离的微小隐孢子虫鼠基因型、兔基因型、猪基因型II和安氏隐孢子虫的子孢子表面抗原CP15基因,其编码基因为大小为357bp,编码119个氨基酸残基,不同来源的CP15基因序列高度一致。其中安氏隐孢子虫的CP15基因成功克隆为本研究首次报道。
成功克隆了上海地区分离的微小隐孢子虫鼠基因型、兔基因型和猪基因型II的子孢子表面抗原P23基因,大小为342bp,编码113个氨基酸,不同来源的P23基因序列高度一致。成功构建了微小隐孢子虫鼠基因型的原核重组表达质粒pET-28a(+)-CP15和pET-28a(+)—P23,实现了CP15和P23基因在大肠杆菌中的表达。ELISA初步分析表明CP15和P23基因具有良好的抗原性。
上述工作的进展,为逐步查明上海地区目前流行的隐孢子虫病原种类、掌握隐孢子虫病流行状况和特点提供了基础,具有重要的公共卫生学意义。隐孢子虫子孢子表面抗原CP15和P23基因的成功克隆和表达,为进一步利用基因的表达产物进行免疫预防和诊断研究提供了条件。
Cryptosporidium spp. is a protozoan parasite and it is harmful severely to men and animals. It grows in epithelial cells and digestive organs of men, animals, birds, creeperes or fishes and it causes diarrhea. Cryptosporidium spp. is known to be ubiquitous and to have many hosts. The diarrhea can be cure by themselves about the normal immune people, but when people who had immune deficiency, the diarrhea maybe serious even take to death.
There were different characteristics and hostes about Cryptosporidium spp. The species and genotypes of Cryptosporidium spp. isolated from mouse, rabbit, chicken, pig and dairy calves in Shanghai district were identified by nested polymerase chain reaction linked restriction fragment length polymorphism (PCR-RFLP). The results showed that there were three species and three genotypes, Cryptosporidium spp. which comes from mouse was Cryptosporidium paravum mouse genotype, rabbit was Cryptosporidium paravum rabbit genotype, chicken was Cryptosporidium baileyi, swine was Cryptosporidium paravum pig genotype II and dairy calves was Cryptospori—dium andersoni.
The sporozoite surface antigen CP15 gene which came from C.paravum mouse genotype, C. paravum rabbit genotype, C. paravum pig genotype II and C. andersoni were successful amplified and encoded 119 amino acids. Cryptosporidium spp. came from different sources had a highly homoely and C. andersoni was first report in this expriment.
The sporozoite surface antigen P23 gene which came from C. paravum mouse genotype, C. paravum rabbit genotype and C.paravum pig genotype II were successful amplified and encoded 113 amino acids. Cryptosporidium spp. came from different sources had a highly homoely. The prokaryotic expressing vector pET-28a(+)-CP15 and pET-28a(+)-P23 were constructed and expressed in E.coli successfuliy. Both of them had a well antigenicity which analyzed by ELISA.
The development of our experiments we had done would much helpful to know the disease sources and epidemic conditions about Cryptosporidium spp. in Shanghai district, and had an important sense about public hygieology. The sporozoite surface antigen CP15 and P23 gene which were successful cloned might be much helpful for the next work of using these genes to develop a immunoprophylaxis or detection technique.
引文
[1] O Donoghue P J.Cryptosporidium and Cryptosporidiosis in man and animals.International Journal for parasitology,1995,25(2):139~195
[2]Rochelle RA,Jutras EM,Atwill E R et al.Pol ymorphisms in B-Tubulin gene of Cryptosporidium paravum differertiate between isolated based on animals hostes but not geographivorign.J Parasitol,1999,25(5):986~989
[3] Tyzzer E.A sporozoon found in the peptic glands of the common mouse.Proc Soc Exp Biol Med,1907,5:12~13
[4] Tyzzer E.Cryptosporidium parvum,a coccidium found in the small intestine of the common mouse.Arch Protisenkd,1912,26:394~412
[5] Vetterling J M,Jervis HR,Merrill T G,et al.Cryptosporidium wrairi sp. n. from the guinea pig Cavie poroellus with an emendation of the genus.J Protozool,1971,18:243~247
[6] Iseki M.Cryptosporidium felis sp. n. (Protozoa:Eimeriorina) from the domestic cat.Jpn J Patssitol,1979,28:285~307
[7] Lindsay D S,Upton S J,Owens D S,et a1.Cryptosporidium andersoni n sp(Apicomplexa: Cryptosporiidae)from cattle,Bos taurus.J Eukaryot Microbiol,2000,47:91~95
[8] Fayer R,Trout J M,Xiao L,et a1.Ctyptosporidium canis n sp from domestic dogs.J Parasitol,2001,87:1415~1422
[9] Morgan-Ryan U M,Fall A,U'ard L A,et al.Cryptosporidium hominis n sp(Apicomplexa: Cryptosporidiidae)from Homo Sapiens.J Eukaryot Microbiol,2002,49:433~440
[10] Slavin D.Cryptosporidiu meleagridis(sp Nov).J Comp Pathol,1955,65:262~270
[11] Cturent W L,Upton S J,Haynes T B.The life cycle of Cryptosporidium baileyi n sp (Apicomplexa:Cryptosporidiidae)infecting chickens.J Protozool,1986,33:289~296
[12] Ryan U M,Xiao L,Read C,et al.A tedescription of Cryptosporidium galli Pavlbsek, ( Apicomplexa: Cryptosporidiidae ) from birds.J Parasitol,1999,89:809~813
[13] Levine N D.Some corrections of coccidian (Apicomplexa:Protozoa)nomenclature.J Pacesitol,1980,66:830~834
[14] Koudela B D,Modry.New species of Cryptosporidium (Apicomplexa:Cryptosporidiidae) from lizards.Folia Parasitol,1998,45:93~100
[15] Alvare-Pellitero P,Sitj-Bobadills A.Cryptosporidium molnari n sp (Apicomplexa: Cryptosporidiidae) infecting two marine Gsh species,Sparus aurata L and Dicentrarchus labrax.Int J Parasitol,2002,32:1007~1021
[16] Xiao L,Irshad M S,Una M R,et al.Host adaptation and host parasite co-evolution in Cryptosporidium:implications for taxonomy and public health.Int J Parasitol,2002,32(14):1773~1785
[17] Szewzyk U,Szewyk R,Manz W.Microbiological Safety of drinking Water.Annu Rev Microbiol,2000,(54):81127
[18] 陈义民,李观娣,杜重波,等.兰州地区发现犊牛隐孢子虫病.中国兽医科技,1986,12(12):61
[19] 张敬伦.鸡的隐孢子虫病在广东首次发现.中国兽医科技,1988,1:63
[20] 韩范.南京地区人体隐孢子虫病 2 例报告.江苏医学,1987,13:692
[21] Naciri M,Lefay M P,Mancassola R,et al.Role of Cryptosporidium parvum in neonatal diarrhea complexa in sucking and dairy calves in France.V Para,1999,18(4):353~355
[22] 李培英,陈圣军,刘亚,等.安徽阜阳地区黄牛隐孢子虫感染情况的调查.畜牧与兽医,1999,31(3):19~20
[23] 李培英,廖圣法,陆凤琳等.安徽省奶牛隐孢子虫感染情况及分布特点.中国兽医学报,1999,19(3):275~277
[24] Wieler L H,Ilieff A,Herbst W,et al.Prevalence of enteropathogens in sucking and weaned piglets with diarrhea in Southern Germany.Zentralblatt fur Veterinarmedizin Reihe B,2001,75(2):123~142
[25] 刘毅,刘锌宁,李道解,等.猪隐孢子虫病调查.中国兽医杂志,1993,26(6):5~6
[26] 张龙现,宁长申,许兆孟,等.郑州市郊猪隐孢子虫感染情况的初步调查.中国兽医寄生虫病,1999,6(3):19～20
[27] GOODWIN M A.Cryptosporidiosis in birds a review.Av Pathol,1989,18(3):365~384
[28] 陈兆国,吴薛忠,史天卫,等.上海地区鸡鸭隐孢子虫病调查及人工感染试验.中国兽医科技,1996,26(10):17~19
[29] 刘毅,曹小明,李健夫,等.湖南鸡隐孢子虫的感染情况调查及人工感染试验.中国兽医科技,1994,20(7):5~6
[30] 周望平,肖兵南.湖南省部分地区禽隐孢子虫感染情况调查.中国兽医科技,2000,30(10):15~6
[31] 陆绍红,林爱芬,陈睿,等.浙江省儿童隐孢子虫感染的诊断及分析.中国人兽共患病杂志,2000,13(1):422~441
[32] 陈佩惠.人本寄生虫学(第 4 版).北京:人民卫生出版社,1995,75
[33] 崔巍,梁瑞文,王尊哲.人兽隐孢子虫在潍坊地区儿童中的感染和流行.中国优生与遗传杂志,2001,9(2):114~115
[34] 陈永忠,孙英,王海连,等.湟中县长毛兔隐孢子虫感染情况.青海畜牧兽医杂志,2001,31(5):24
[35] 李海峰,张守发,许应天,等.延边地区兔隐孢子虫感染情况调查.黑龙江畜牧兽医,2005,9:39~40
[36] Current W L,Garcia L S.Cryptosporidiosis.Clinic Microbiol Rev,1991,4(3):325~358
[37] 韩范,王荣生,许生.隐孢子虫病病原诊断方法的改进.中国寄生虫学与寄生虫病杂志,1990,3(1):73
[38] Macpherson D W,Mcqueen R.Cryptosporidiosis:Multi attribute evaluation of six diagnostic methods.J Clin Microbiol,1993,31(2):198~202
[39] 郭步平,连德润.隐孢子虫卵囊分离纯化方法的比较研究.中国人兽共患病杂志,1997,13(1):34~35
[40] 周吉礼,许春华.原虫性腹泻与对策.世界华人消化杂志,2000,8:93~95
[41] Rusnak J,Hadfield T L,Rhodes M M,et al.Detection of Cryptosporidium oocysts in human fecal specimens by an indirect immunofluorescence assay with monoclonal antibodies.J Clin Microbiol,1989,27(5):1135~1136
[42] Ungar B L P.Enzyme linked immunoassay for detection of Cryptosporidium antigens in fecal specimens.J Clin Microbiol,1990,28(11):2491
[43] Ortega L M,Troncoso J M,Rojo-Vazquez F A,et al.Indentification of Cryptosporidium parvum occyst/sporozite antigens recognized by infected and hyperimmuelambs.Vet Para,1994,53:159~167
[44] Dagan R,Frase D,El-On J,et al.Evaluation of anenzyme immunoassay for the detection of Cryptosporidium parvum in stool specimens from infants and young children in fieldstudies.Am J Trop Med Hyg,1995,52(2):134~138
[45] Farrington M,Lloyd S,Winter S,et al.Patterns of Cryptosporidium antigen And occyst excretion in calves studied by reverse passive haemag glutination and lightmicroscopy.Vet Para,,1995,60:7~16
[46] Arrowood M J,Hurd M R,Mead J R.A new method for evaluating experimental Cryptosporidial parasite load susing immune fluorescence flowcytometry.J Para,1995,81:404~409
[47] Valdez L M,Danghan,Okhuysen,et al.Flowcytometric detection of Cryptosporidium oocysts inhuman stool samples.J Clin Microbiol,1997,35(8):2013~2017
[48] Lappin M R,Ungar B,Brown-Hahn B,et al.Enzyme-linked immunosorbent assay for the detection of Cryptosporidium parvum IgG in the cats.J Para,1997,83(5):957~960
[49] 黄克和,李培英,廖圣法.应用间接ELISA方法检测动物隐孢子虫病抗体.中国兽医学报,1999,19(4):353~355
[50] 尹继刚,张西臣,陈建宝,等.双夹心-ELISA诊断牛隐孢子虫病.中国兽医寄生虫病,1999,7(20):1~2
[51] Laxer M A,Timblin B K,Patel R J.DNA sequence for the specific detection of Cryptosporidium.Am J Trop Med Hyg,1991,45:688
[52] 郭步平,连德润.聚合酶链反应技术在隐孢子虫检测中的应用研究.中国人畜共患病杂志,1998,14(2):33~35
[53] 陈雅棠,马良.聚合酶链反应结合非放射性标记探针检测粪便中微小隐孢子虫.中国寄生虫学与寄生虫病杂志,,1998,16(4):246~250
[54] Wieger H,Tarmara V G,Yuen Y K.Characterization of Cryptosporidium parvum in human and animal feces by single-tube nested PCR and restriction analysis.Parasitol Res,1999,85:707~712
[55] Timothy S,Anita M,Kate H,et al.Detection of single viable Cryptosporidium parvum oocyst in environmental water concentrates by RT-PCR.Applenvron Microbio,1996,62(9):3385~3390
[56] Paul A R,Donna M F,Troy J H,et al.An assay combining cell culture with RT-PCR to detect and determine the infectivity of waterborne Cryptosporidium parvum.Appl Environ Microbio,1997,63(5):2029~2037
[57] Anonymous.Update:treatment of cryptosporidiosis in patients with acquired immune deficiency syndrome(AIDS).Morbid Mortal Weekly Rep,1984,33:117
[58] Wittenberg D F,Miller N M,Vanden Ende J.Spiramycin is not effective in treating Crypotosporidium in infants:resulits of a double-blind randomized trial.J Infect Dis,1989,159:131
[59] Doumbo O.Nitazoxanide in the treatment of Cryptosporidial diarrhea and other intestinal parasitic infections associated with acquired immunodeficiency syndrome in tropical Africal.Am J Trop Med Hyg,1997,56:637
[60] Marshall R J,Flanigan T P.Paromomycinin hibits Crypotosporidium infection of human enterocyte .J Infect Dis,1992,165:772
[61] Fayer R,Ellis W.Paromomycinis effective asprophylaxis for cryptosporidiosis in dairy calves.J Parasitol,1993,79(5):771
[62] Mancassola R,Reperent J M,Naciri M,et al.Chemoprophlaxis of Crypotosporidium parvum infection with paromomycinin kids and immunological study.Antimicrob Chemother,1995,39(1):75
[63] 韩范,潭渭仙,周性兰.南京地区人体隐孢子虫病 2 例报告.江西医药,1998,7(13):692
[64] 苏庆平,陈达光,华雪玲,等.福州地区婴幼儿隐孢子虫病.中国人兽共患病杂志,1998,5:35~36
[65] 赵雪妮,张冬梅,张立宪,等.黑龙江地区婴幼儿隐孢子虫病调查.中国人兽共患病杂志,1998,14(1):73
[66] Sulaiman,Irshad M,Lihua Xiao,et al.Evaluation of Cryptosporidium parvum genotyping Techniques.Applied and Environment Microbiology,1999,65(10):44311~4435
[67] Xiao L,Escalante L,Yang C F,et al.Phylogenetic analysis of Cryptosporidium parasites based on the Small-Subunit rRNA gene locus.Appl Environ Microbiol,1999,65(4):1578~1583
[68] Arrowood M J and Sterling C R.Isolation of Cryptosporidium oocysts and sporozoites using discontinuous sucrose and isopycnic Percoll gradients.Parasitol,1987,73(2):314~319
[69] Xiao L H,John E,Moore,et al.Prevalence and Identity of Cryptosporidium spp. in Pig Slurry.App Envio Microbiol,2006,72(6):4461~4463
[70] Prichard R,Tait A.The role of molecular biology in veterinary parasitology.Vet Parasitol,2001,98(1):169~194
[71] Gasser R B.PCR-based technology in veterinary parasitolgy.Vet Parasitol,1999,84(2):229~258
[72] 杨月中,白会芬,和爱芬,等.野生鼠隐孢子虫对实验动物的感染试验.中国兽医科技,1998,28(7):29~30
[73] 郭敏,戈建军,沈京培,等.鼠隐孢子虫感染模型的建立.江苏预防医学,2005,16(1):58~60
[74] Ernest J A,Blagburn B L,Lindsay D S.Infection dynamics of Cryptosporidium parvum(Apicomplexa:Cryptospridiidae)in neonatal mice(Mus musculus).J Parasitol,1986,72:796~798
[75] Rehg J E.Characterization of a dexamethasone-treated rat model of cryptosporidial infection.J Infect Dis,1988,158(6):1406
[76] 杨建民,鲍嘉铭,陈廷和,等.畜禽隐孢子虫病的调查与人工感染试验.中国兽医杂志,1999,25(12):5~8
[77] 陈兆国,吴薛忠,史天卫,等.上海地区鸡鸭隐孢子虫病调查及人工感染试验.中国兽医科技,1996,26(10):17~20
[78] Jenkin M C , Fayer R . Cloning and expression of cDNA encoding an antigenic Cryptosporidium parvum protein.Mol Biochem Parasitol,1995,71(1):149~152
[79] Tilley M,Upton S J.Both CP15 and CP25 are left as trails behind gliding sporozoite of Cryptosporidium parvum (Apicomplexa). FEMS Microbiol Lett,1994,120:275~278
[80] Reperant J M, Naciri M,Chardes T,et al.Immunological characterization of 17-kDa antigen from Cryptosporidium parvum recognized early by mucosal IgA antibodies.FEMS Microbiol lett,1992,99:7~14
[81] Sagodira S,Iochmann S, Mevelec M N,et al.Nasal immunization of mice with Cryptosporidium parvum DNA induces systemic and intestinal immune responses.Present Immunol,1999,21:507~516
[82] Moss D M,Chappell C L,Okhuysen,et al.The antibody response to 27-,17-,and 15-kDa Cryptosporidium antigens following experimental infection in human.Infect Dis,1998,178:827~833
[83] Jenkin M C.Present and future control of Cryptosporidiosis in humans and animals.Expere Rev Uaccines,2004,3(6):669~671
[84] Sagodira S,Buzoni-Gatel D,Iochmann S,et al.Protection of kids against Cryptosporidium parvum infection after immunization of dams with CP15-DNA .Vaccine,1999,17(19):2346~2355
[86] 王妍,张守发,张乾坤,等.牛隐孢子虫卵囊分离和纯化方法的比较试验.黑龙江畜牧兽医,2003,8:9~11
[87] Arrowood M J,Sterling C R.Isolation of Cryptosporidium oocysts and sporozoites using discontinuous sucrose and isopycnic Percoll gradients .J. Parasitol,1987,73(2):314~319
[88] Perryman L E,Jamser D P,Riggs M W,et al.A cloned gene of Cryptosporidium parvum encodes neutralization-sensitive epitopes.Mol Biochem Parasitol,1996,80:137~147
[89] Theodos C M.Innate and cell-mediated immune responses to Cryptosporidium parvum.Adv Parasitol,1998,40:88-119
[90] Fayer R.Cryptosporidium in cryptosporidiosis in men and animals.CRC Press,1997,1~49
[91] Ortega L M,Wright S E.Age-related resistance in ovine cryptosporidiosis:patterns of infection and humoral immune response.Vet Parasitol,1994,(53):159~166
[92] L M Tatalick.Fatal cryptosporidiosis in quail.Vet Parasitol,1994,(55):11~13
[93] Schaefer D A,Auerhach-Dixon B A,Riggs M W.Characterization and formulation of multiple epitope -specific neutralizing monoclonal antibodies for passive immunization against cryptosporidiosis.Infect lmmun,2000,68(5):2608~2616
[94] Penyman L E,Kapil S J,Jones M L,et a1.Protection of calves against Cryptosporidiosis with immune bovine colostrum induced by a Cryptosporidium parvum recombinant protein.Vaccine,1999,17(17):2142~2149
[95] Brackett E J,Mason P H,Savidge J,et al.Excretion pattern of mucosally delivered antibodies to p23 in Cryptosporidium parvum infected calves.Vet Immunol Immunopathol,2000,76(324):309~317
[96] Eisenberg J N,Priest J W,Lammie P J,et a1.The Serologic response to Cryptosporidium in HIV-infected persons:implications for epidemiologic research.Emerg Infect Dis,2001,7(6):1004~1009
[97] 何宏轩,张西臣,,尹继刚,等.微小隐孢子虫子孢子表面蛋白质粒 DNA 接种诱导 Balb/c小鼠的免疫应答.免疫学杂志,2002,4(18):263~265
[98] 何宏轩,张西臣,欧阳红生,等.微小隐孢子虫子孢子表面抗原基因 gp23 的克隆和测序.中国寄生虫学与寄生虫病杂志,2002,20(3):161~163
[99] 何宏轩,张西臣,欧阳红生,等.微小隐孢子虫子孢子表面抗原P23融合蛋白在大肠杆菌的表达.中国人兽共患病杂志,2003,19(3):52~54
[100] Rosenblatt J E,Sloan L M.Evaluation of an enzyme linked immunosorbent assay for detection of Cryptosporidium spp. in stool specimens.J Clin Microbio1,1993,3l(6):1468~1471
[101] Baron E J,Schenone C,Tanenbaum B.Comparison of three methods for detection of Cryptosporidium oocysts in a 1ow-prevalence population.J Clin Microbio1,1989,27(1):223~224
[102] Gobet P,Buisson J C,Vagner O,et a1.Detection of Cryptosporidium parvum DNA in formed human feces by a sensitive PCR-based assay including uracil- N-glyeosylase inactivation.J Clin Microbio1,1997,35(1):254~256
[103] Dagan R,Fraser D,El-On J,et al.Evaluation of an enzyme immunoassay for the detection of Cryptosporidium spp..In stool specimens from infants and young children in field studies.Am J Trop Med Hyg,1995,52(2):134~138
[104] Parisi M T,Tiemo PM Jr.Evaluation of new rapid commercial enzyme immunoassay for detection of Cryptosporidium oocysts in untreated stool specimens.J Clin Microbiol,1995,33(7):1963~1965
[105] Doing K M,Harem J L,Jellison J A,et al.False-positive results obtained with the Alexon ProSpecT Cryptosporidium enzyme immunoassay.J Clin Microbiol.1999,37(5):1582~1583
[2]Rochelle RA,Jutras EM,Atwill E R et al.Pol ymorphisms in B-Tubulin gene of Cryptosporidium paravum differertiate between isolated based on animals hostes but not geographivorign.J Parasitol,1999,25(5):986~989
[3] Tyzzer E.A sporozoon found in the peptic glands of the common mouse.Proc Soc Exp Biol Med,1907,5:12~13
[4] Tyzzer E.Cryptosporidium parvum,a coccidium found in the small intestine of the common mouse.Arch Protisenkd,1912,26:394~412
[5] Vetterling J M,Jervis HR,Merrill T G,et al.Cryptosporidium wrairi sp. n. from the guinea pig Cavie poroellus with an emendation of the genus.J Protozool,1971,18:243~247
[6] Iseki M.Cryptosporidium felis sp. n. (Protozoa:Eimeriorina) from the domestic cat.Jpn J Patssitol,1979,28:285~307
[7] Lindsay D S,Upton S J,Owens D S,et a1.Cryptosporidium andersoni n sp(Apicomplexa: Cryptosporiidae)from cattle,Bos taurus.J Eukaryot Microbiol,2000,47:91~95
[8] Fayer R,Trout J M,Xiao L,et a1.Ctyptosporidium canis n sp from domestic dogs.J Parasitol,2001,87:1415~1422
[9] Morgan-Ryan U M,Fall A,U'ard L A,et al.Cryptosporidium hominis n sp(Apicomplexa: Cryptosporidiidae)from Homo Sapiens.J Eukaryot Microbiol,2002,49:433~440
[10] Slavin D.Cryptosporidiu meleagridis(sp Nov).J Comp Pathol,1955,65:262~270
[11] Cturent W L,Upton S J,Haynes T B.The life cycle of Cryptosporidium baileyi n sp (Apicomplexa:Cryptosporidiidae)infecting chickens.J Protozool,1986,33:289~296
[12] Ryan U M,Xiao L,Read C,et al.A tedescription of Cryptosporidium galli Pavlbsek, ( Apicomplexa: Cryptosporidiidae ) from birds.J Parasitol,1999,89:809~813
[13] Levine N D.Some corrections of coccidian (Apicomplexa:Protozoa)nomenclature.J Pacesitol,1980,66:830~834
[14] Koudela B D,Modry.New species of Cryptosporidium (Apicomplexa:Cryptosporidiidae) from lizards.Folia Parasitol,1998,45:93~100
[15] Alvare-Pellitero P,Sitj-Bobadills A.Cryptosporidium molnari n sp (Apicomplexa: Cryptosporidiidae) infecting two marine Gsh species,Sparus aurata L and Dicentrarchus labrax.Int J Parasitol,2002,32:1007~1021
[16] Xiao L,Irshad M S,Una M R,et al.Host adaptation and host parasite co-evolution in Cryptosporidium:implications for taxonomy and public health.Int J Parasitol,2002,32(14):1773~1785
[17] Szewzyk U,Szewyk R,Manz W.Microbiological Safety of drinking Water.Annu Rev Microbiol,2000,(54):81127
[18] 陈义民,李观娣,杜重波,等.兰州地区发现犊牛隐孢子虫病.中国兽医科技,1986,12(12):61
[19] 张敬伦.鸡的隐孢子虫病在广东首次发现.中国兽医科技,1988,1:63
[20] 韩范.南京地区人体隐孢子虫病 2 例报告.江苏医学,1987,13:692
[21] Naciri M,Lefay M P,Mancassola R,et al.Role of Cryptosporidium parvum in neonatal diarrhea complexa in sucking and dairy calves in France.V Para,1999,18(4):353~355
[22] 李培英,陈圣军,刘亚,等.安徽阜阳地区黄牛隐孢子虫感染情况的调查.畜牧与兽医,1999,31(3):19~20
[23] 李培英,廖圣法,陆凤琳等.安徽省奶牛隐孢子虫感染情况及分布特点.中国兽医学报,1999,19(3):275~277
[24] Wieler L H,Ilieff A,Herbst W,et al.Prevalence of enteropathogens in sucking and weaned piglets with diarrhea in Southern Germany.Zentralblatt fur Veterinarmedizin Reihe B,2001,75(2):123~142
[25] 刘毅,刘锌宁,李道解,等.猪隐孢子虫病调查.中国兽医杂志,1993,26(6):5~6
[26] 张龙现,宁长申,许兆孟,等.郑州市郊猪隐孢子虫感染情况的初步调查.中国兽医寄生虫病,1999,6(3):19～20
[27] GOODWIN M A.Cryptosporidiosis in birds a review.Av Pathol,1989,18(3):365~384
[28] 陈兆国,吴薛忠,史天卫,等.上海地区鸡鸭隐孢子虫病调查及人工感染试验.中国兽医科技,1996,26(10):17~19
[29] 刘毅,曹小明,李健夫,等.湖南鸡隐孢子虫的感染情况调查及人工感染试验.中国兽医科技,1994,20(7):5~6
[30] 周望平,肖兵南.湖南省部分地区禽隐孢子虫感染情况调查.中国兽医科技,2000,30(10):15~6
[31] 陆绍红,林爱芬,陈睿,等.浙江省儿童隐孢子虫感染的诊断及分析.中国人兽共患病杂志,2000,13(1):422~441
[32] 陈佩惠.人本寄生虫学(第 4 版).北京:人民卫生出版社,1995,75
[33] 崔巍,梁瑞文,王尊哲.人兽隐孢子虫在潍坊地区儿童中的感染和流行.中国优生与遗传杂志,2001,9(2):114~115
[34] 陈永忠,孙英,王海连,等.湟中县长毛兔隐孢子虫感染情况.青海畜牧兽医杂志,2001,31(5):24
[35] 李海峰,张守发,许应天,等.延边地区兔隐孢子虫感染情况调查.黑龙江畜牧兽医,2005,9:39~40
[36] Current W L,Garcia L S.Cryptosporidiosis.Clinic Microbiol Rev,1991,4(3):325~358
[37] 韩范,王荣生,许生.隐孢子虫病病原诊断方法的改进.中国寄生虫学与寄生虫病杂志,1990,3(1):73
[38] Macpherson D W,Mcqueen R.Cryptosporidiosis:Multi attribute evaluation of six diagnostic methods.J Clin Microbiol,1993,31(2):198~202
[39] 郭步平,连德润.隐孢子虫卵囊分离纯化方法的比较研究.中国人兽共患病杂志,1997,13(1):34~35
[40] 周吉礼,许春华.原虫性腹泻与对策.世界华人消化杂志,2000,8:93~95
[41] Rusnak J,Hadfield T L,Rhodes M M,et al.Detection of Cryptosporidium oocysts in human fecal specimens by an indirect immunofluorescence assay with monoclonal antibodies.J Clin Microbiol,1989,27(5):1135~1136
[42] Ungar B L P.Enzyme linked immunoassay for detection of Cryptosporidium antigens in fecal specimens.J Clin Microbiol,1990,28(11):2491
[43] Ortega L M,Troncoso J M,Rojo-Vazquez F A,et al.Indentification of Cryptosporidium parvum occyst/sporozite antigens recognized by infected and hyperimmuelambs.Vet Para,1994,53:159~167
[44] Dagan R,Frase D,El-On J,et al.Evaluation of anenzyme immunoassay for the detection of Cryptosporidium parvum in stool specimens from infants and young children in fieldstudies.Am J Trop Med Hyg,1995,52(2):134~138
[45] Farrington M,Lloyd S,Winter S,et al.Patterns of Cryptosporidium antigen And occyst excretion in calves studied by reverse passive haemag glutination and lightmicroscopy.Vet Para,,1995,60:7~16
[46] Arrowood M J,Hurd M R,Mead J R.A new method for evaluating experimental Cryptosporidial parasite load susing immune fluorescence flowcytometry.J Para,1995,81:404~409
[47] Valdez L M,Danghan,Okhuysen,et al.Flowcytometric detection of Cryptosporidium oocysts inhuman stool samples.J Clin Microbiol,1997,35(8):2013~2017
[48] Lappin M R,Ungar B,Brown-Hahn B,et al.Enzyme-linked immunosorbent assay for the detection of Cryptosporidium parvum IgG in the cats.J Para,1997,83(5):957~960
[49] 黄克和,李培英,廖圣法.应用间接ELISA方法检测动物隐孢子虫病抗体.中国兽医学报,1999,19(4):353~355
[50] 尹继刚,张西臣,陈建宝,等.双夹心-ELISA诊断牛隐孢子虫病.中国兽医寄生虫病,1999,7(20):1~2
[51] Laxer M A,Timblin B K,Patel R J.DNA sequence for the specific detection of Cryptosporidium.Am J Trop Med Hyg,1991,45:688
[52] 郭步平,连德润.聚合酶链反应技术在隐孢子虫检测中的应用研究.中国人畜共患病杂志,1998,14(2):33~35
[53] 陈雅棠,马良.聚合酶链反应结合非放射性标记探针检测粪便中微小隐孢子虫.中国寄生虫学与寄生虫病杂志,,1998,16(4):246~250
[54] Wieger H,Tarmara V G,Yuen Y K.Characterization of Cryptosporidium parvum in human and animal feces by single-tube nested PCR and restriction analysis.Parasitol Res,1999,85:707~712
[55] Timothy S,Anita M,Kate H,et al.Detection of single viable Cryptosporidium parvum oocyst in environmental water concentrates by RT-PCR.Applenvron Microbio,1996,62(9):3385~3390
[56] Paul A R,Donna M F,Troy J H,et al.An assay combining cell culture with RT-PCR to detect and determine the infectivity of waterborne Cryptosporidium parvum.Appl Environ Microbio,1997,63(5):2029~2037
[57] Anonymous.Update:treatment of cryptosporidiosis in patients with acquired immune deficiency syndrome(AIDS).Morbid Mortal Weekly Rep,1984,33:117
[58] Wittenberg D F,Miller N M,Vanden Ende J.Spiramycin is not effective in treating Crypotosporidium in infants:resulits of a double-blind randomized trial.J Infect Dis,1989,159:131
[59] Doumbo O.Nitazoxanide in the treatment of Cryptosporidial diarrhea and other intestinal parasitic infections associated with acquired immunodeficiency syndrome in tropical Africal.Am J Trop Med Hyg,1997,56:637
[60] Marshall R J,Flanigan T P.Paromomycinin hibits Crypotosporidium infection of human enterocyte .J Infect Dis,1992,165:772
[61] Fayer R,Ellis W.Paromomycinis effective asprophylaxis for cryptosporidiosis in dairy calves.J Parasitol,1993,79(5):771
[62] Mancassola R,Reperent J M,Naciri M,et al.Chemoprophlaxis of Crypotosporidium parvum infection with paromomycinin kids and immunological study.Antimicrob Chemother,1995,39(1):75
[63] 韩范,潭渭仙,周性兰.南京地区人体隐孢子虫病 2 例报告.江西医药,1998,7(13):692
[64] 苏庆平,陈达光,华雪玲,等.福州地区婴幼儿隐孢子虫病.中国人兽共患病杂志,1998,5:35~36
[65] 赵雪妮,张冬梅,张立宪,等.黑龙江地区婴幼儿隐孢子虫病调查.中国人兽共患病杂志,1998,14(1):73
[66] Sulaiman,Irshad M,Lihua Xiao,et al.Evaluation of Cryptosporidium parvum genotyping Techniques.Applied and Environment Microbiology,1999,65(10):44311~4435
[67] Xiao L,Escalante L,Yang C F,et al.Phylogenetic analysis of Cryptosporidium parasites based on the Small-Subunit rRNA gene locus.Appl Environ Microbiol,1999,65(4):1578~1583
[68] Arrowood M J and Sterling C R.Isolation of Cryptosporidium oocysts and sporozoites using discontinuous sucrose and isopycnic Percoll gradients.Parasitol,1987,73(2):314~319
[69] Xiao L H,John E,Moore,et al.Prevalence and Identity of Cryptosporidium spp. in Pig Slurry.App Envio Microbiol,2006,72(6):4461~4463
[70] Prichard R,Tait A.The role of molecular biology in veterinary parasitology.Vet Parasitol,2001,98(1):169~194
[71] Gasser R B.PCR-based technology in veterinary parasitolgy.Vet Parasitol,1999,84(2):229~258
[72] 杨月中,白会芬,和爱芬,等.野生鼠隐孢子虫对实验动物的感染试验.中国兽医科技,1998,28(7):29~30
[73] 郭敏,戈建军,沈京培,等.鼠隐孢子虫感染模型的建立.江苏预防医学,2005,16(1):58~60
[74] Ernest J A,Blagburn B L,Lindsay D S.Infection dynamics of Cryptosporidium parvum(Apicomplexa:Cryptospridiidae)in neonatal mice(Mus musculus).J Parasitol,1986,72:796~798
[75] Rehg J E.Characterization of a dexamethasone-treated rat model of cryptosporidial infection.J Infect Dis,1988,158(6):1406
[76] 杨建民,鲍嘉铭,陈廷和,等.畜禽隐孢子虫病的调查与人工感染试验.中国兽医杂志,1999,25(12):5~8
[77] 陈兆国,吴薛忠,史天卫,等.上海地区鸡鸭隐孢子虫病调查及人工感染试验.中国兽医科技,1996,26(10):17~20
[78] Jenkin M C , Fayer R . Cloning and expression of cDNA encoding an antigenic Cryptosporidium parvum protein.Mol Biochem Parasitol,1995,71(1):149~152
[79] Tilley M,Upton S J.Both CP15 and CP25 are left as trails behind gliding sporozoite of Cryptosporidium parvum (Apicomplexa). FEMS Microbiol Lett,1994,120:275~278
[80] Reperant J M, Naciri M,Chardes T,et al.Immunological characterization of 17-kDa antigen from Cryptosporidium parvum recognized early by mucosal IgA antibodies.FEMS Microbiol lett,1992,99:7~14
[81] Sagodira S,Iochmann S, Mevelec M N,et al.Nasal immunization of mice with Cryptosporidium parvum DNA induces systemic and intestinal immune responses.Present Immunol,1999,21:507~516
[82] Moss D M,Chappell C L,Okhuysen,et al.The antibody response to 27-,17-,and 15-kDa Cryptosporidium antigens following experimental infection in human.Infect Dis,1998,178:827~833
[83] Jenkin M C.Present and future control of Cryptosporidiosis in humans and animals.Expere Rev Uaccines,2004,3(6):669~671
[84] Sagodira S,Buzoni-Gatel D,Iochmann S,et al.Protection of kids against Cryptosporidium parvum infection after immunization of dams with CP15-DNA .Vaccine,1999,17(19):2346~2355
[86] 王妍,张守发,张乾坤,等.牛隐孢子虫卵囊分离和纯化方法的比较试验.黑龙江畜牧兽医,2003,8:9~11
[87] Arrowood M J,Sterling C R.Isolation of Cryptosporidium oocysts and sporozoites using discontinuous sucrose and isopycnic Percoll gradients .J. Parasitol,1987,73(2):314~319
[88] Perryman L E,Jamser D P,Riggs M W,et al.A cloned gene of Cryptosporidium parvum encodes neutralization-sensitive epitopes.Mol Biochem Parasitol,1996,80:137~147
[89] Theodos C M.Innate and cell-mediated immune responses to Cryptosporidium parvum.Adv Parasitol,1998,40:88-119
[90] Fayer R.Cryptosporidium in cryptosporidiosis in men and animals.CRC Press,1997,1~49
[91] Ortega L M,Wright S E.Age-related resistance in ovine cryptosporidiosis:patterns of infection and humoral immune response.Vet Parasitol,1994,(53):159~166
[92] L M Tatalick.Fatal cryptosporidiosis in quail.Vet Parasitol,1994,(55):11~13
[93] Schaefer D A,Auerhach-Dixon B A,Riggs M W.Characterization and formulation of multiple epitope -specific neutralizing monoclonal antibodies for passive immunization against cryptosporidiosis.Infect lmmun,2000,68(5):2608~2616
[94] Penyman L E,Kapil S J,Jones M L,et a1.Protection of calves against Cryptosporidiosis with immune bovine colostrum induced by a Cryptosporidium parvum recombinant protein.Vaccine,1999,17(17):2142~2149
[95] Brackett E J,Mason P H,Savidge J,et al.Excretion pattern of mucosally delivered antibodies to p23 in Cryptosporidium parvum infected calves.Vet Immunol Immunopathol,2000,76(324):309~317
[96] Eisenberg J N,Priest J W,Lammie P J,et a1.The Serologic response to Cryptosporidium in HIV-infected persons:implications for epidemiologic research.Emerg Infect Dis,2001,7(6):1004~1009
[97] 何宏轩,张西臣,,尹继刚,等.微小隐孢子虫子孢子表面蛋白质粒 DNA 接种诱导 Balb/c小鼠的免疫应答.免疫学杂志,2002,4(18):263~265
[98] 何宏轩,张西臣,欧阳红生,等.微小隐孢子虫子孢子表面抗原基因 gp23 的克隆和测序.中国寄生虫学与寄生虫病杂志,2002,20(3):161~163
[99] 何宏轩,张西臣,欧阳红生,等.微小隐孢子虫子孢子表面抗原P23融合蛋白在大肠杆菌的表达.中国人兽共患病杂志,2003,19(3):52~54
[100] Rosenblatt J E,Sloan L M.Evaluation of an enzyme linked immunosorbent assay for detection of Cryptosporidium spp. in stool specimens.J Clin Microbio1,1993,3l(6):1468~1471
[101] Baron E J,Schenone C,Tanenbaum B.Comparison of three methods for detection of Cryptosporidium oocysts in a 1ow-prevalence population.J Clin Microbio1,1989,27(1):223~224
[102] Gobet P,Buisson J C,Vagner O,et a1.Detection of Cryptosporidium parvum DNA in formed human feces by a sensitive PCR-based assay including uracil- N-glyeosylase inactivation.J Clin Microbio1,1997,35(1):254~256
[103] Dagan R,Fraser D,El-On J,et al.Evaluation of an enzyme immunoassay for the detection of Cryptosporidium spp..In stool specimens from infants and young children in field studies.Am J Trop Med Hyg,1995,52(2):134~138
[104] Parisi M T,Tiemo PM Jr.Evaluation of new rapid commercial enzyme immunoassay for detection of Cryptosporidium oocysts in untreated stool specimens.J Clin Microbiol,1995,33(7):1963~1965
[105] Doing K M,Harem J L,Jellison J A,et al.False-positive results obtained with the Alexon ProSpecT Cryptosporidium enzyme immunoassay.J Clin Microbiol.1999,37(5):1582~1583