基于凝集素基因对部分隐孢子虫种类的分子鉴定  

Molecular identification of some species of Cryptosporidium based on Lectin gene

作　　者：刘珍珍[1,2] 胡苏辉 张璐 陈凯丽[1,2] 张龙现 宁长申 菅复春[1,2] Rongchang YANG LIU Zhen-zhen;HU Su-hui;ZHANG Lu;CHEN Kai-li;ZHANG Long-xian;NING Chang-shen;JIAN Fu-chun;YANG Rong-chang(College of Animal Science and Veterinary Medicine, Henan Agricultural University, Zhengzhou 450002, China;International Joint Research Laboratory for Zoonotic Diseases of Henan Province, Zhengzhou 450002, China;School of Veterinary and Life Sciences, Murdoch University, Murdoch, Western Australia, 6150, Australia)

机构地区：[1]河南农业大学牧医工程学院,郑州450002 [2]河南省人兽共患病国际联合实验室,郑州450002 [3]澳大利亚莫道克大学生命与兽医学院,澳大利亚6150

出　　处：《中国人兽共患病学报》2018年第4期290-295,共6页Chinese Journal of Zoonoses

基　　金：国家自然科学基金国际合作交流项目(No.31310103012)和河南省基础与前沿技术研究项目(No.162300410166)联合资助

摘　　要：目的为了解隐孢子虫凝集素基因在不同分离株的序列差异及其遗传进化关系,以SSU rRNA基因作参照,评价Lectin基因位点作为隐孢子虫基因分型和进化关系分析的可行性。方法采用巢式PCR,分别在Lectin和SSU rRNA位点处对本实验室分离保存的多个隐孢子虫分离株进行PCR的扩增。用Clustalx对扩增序列与参考序列进行比对,用MEGA5中的邻近法(Neighbor joining method NJ),进行进化树的构建。结果基于Lectin基因位点,在C.parvum、C.hominis、C.cuniculus及其在SSU rRNA处与人源C.hominis极为相近的horse genotype、驴源C.hominis均成功扩增出了大小在450bp左右的目的条带,并进行了进化树的分析,不同种类和同种不同动物来源的隐孢子虫分布在不同的分支上。结论 Lectin基因位点可很好的区分SSU rRNA序列极为相似的几种隐孢子虫,有望为人兽共患隐孢子虫分类和遗传研究提供新的基因靶标。In order to understand the sequence differences and their genetic evolution of Lectin gene in different Cryptosporidium isolates, the Lectin gene locus was evaluated as the feasibility of Cryptosporidium gene typing and evolutionary relationship with SSU rRNA gene as reference. The nested PCR was used to amplify the Cryptosporidium isolates isolated from the laboratory at Lectin and SSU rRNA loci. ClustalX was used to compare the amplified sequence with the reference sequence, and the phylogenetic tree was constructed by the Neighbor joining method NJ in MEGA5. Lectin gene locus was used as a typing tool to amplify C. parvum, C. hominis, C. cuniculus and horse genotype, C. hominis from donkey which were very similar to C. hominis from Human. These isolates at Lectin gene was amplified successfully（about 450 base pair）and then was carried out a phylogenetic tree analysis. Results indicated that, different Cryptosporidium species and the same species derived from different animals were in different branches. Lectin gene locus is very likely to be a new Cryptosporidium classification tool which provides a reference basis for the further development of zoonotic Cryptosporidium classification and genetic research. Therefore, it is worthy to do more study on the gene. The possibility of using Lectin gene as a genotyping tool was explored by amplifying the SSU rRNA and Lectin loci of Cryptosporidium species, respectively. The study is to provide new molecular tools for the epidemiological investigation of Cryptosporidium, analysis of genetic structure and the origin of epidemic disease. The results of sequence alignment and phylogenetic tree show that Lectin gene can be used to identify the species of Cryptosporidium which similar to the SSU rRNA sequences. It is expected to add a new molecular tool for molecular epidemiology and population genetic analysis of Cryptosporidium, and it is a useful supplement to the classification tool of Cryptosporidium.

关 键 词：隐孢子虫 Lectin基因 SSU RRNA基因 基因分型 

分 类 号：R382[医药卫生—医学寄生虫学]