Department of Parasitology, Harbin Medical University, Harbin, Heilongjiang 150081, China.
National Institute of Parasitic Diseases, Chinese Center for Disease Control and Prevention, Chinese Center for Tropical Diseases Research, WHO Collaborating Centre for Tropical Diseases, National Center for International Research on Tropical Diseases, Ministry of Science and Technology, Key Laboratory of Parasite and Vector Biology, MOH, Shanghai 200025, China.
Infect Genet Evol. 2018 Oct;64:249-253. doi: 10.1016/j.meegid.2018.07.006. Epub 2018 Jul 5.
Multilocus sequence typing (MLST) tools have been used widely to characterize population genetic structure of some Cryptosporidium species. To understand MLST subtypes and population genetic structure of Cryptosporidium cuniculus from rabbits in Heilongjiang Province, China, 34 C. cuniculus DNA specimens were collected including VbA21 (n = 6), VbA28 (n = 2), VbA29 (n = 18) and VbA32 (n = 8). They were analyzed by nested-PCR amplification and sequencing at seven microsatellite, minisatellite and polymorphic loci including CP47, CP56, ML2, DZ-HRGP, MSC6-5, MSC6-7 and RPGR. The CP47, CP56, MSC6-5 and MSC6-7 loci were monomorphic. The remaining loci were polymorphic, with two, three and two subtypes being found at ML2, DZ-HRGP and RPGR loci, respectively. Six MLST subtypes were obtained based on sequence information of 29 DNA specimens successfully amplified at all eight loci including gp60 locus. Linkage disequilibrium (LD) analysis showed a clonal population structure of C. cuniculus in the investigated areas. STRUCTURE, neighbor-joining and network analyses indicated the presence of two distinct groups, corresponding to VbA21 subtype and VbA28, VbA29 and VbA32 subtypes. This is the first report of MLST analysis of C. cuniculus. A clonal population structure of C. cuniculus suggested the prevalence of C. cuniculus in Heilongjiang Province is not attributed to the introduction of rabbits. Thus, prevention and control strategies should be focused on making stricter measures to avoid occurrence of cross-transmission and re-infection among rabbit individuals. Based on the previous findings of VbA21 subtype only in rabbits and VbA28, VbA29 and VbA32 subtypes both in rabbits and humans, the results of subpopulation analyses might be used to assess zoonotic potential of C. cuniculus subtypes in Vb family. These data will be helpful to explore source attribution of infection/contamination of C. cuniculus and understand its transmission dynamics in humans and rabbits in the investigated areas.
多位点序列分型（MLST）工具已被广泛用于描述一些隐孢子虫物种的种群遗传结构。为了解黑龙江省兔源隐孢子虫 C. cuniculus 的 MLST 亚型和种群遗传结构，共采集 34 份 C. cuniculus DNA 标本，包括 VbA21（n=6）、VbA28（n=2）、VbA29（n=18）和 VbA32（n=8）。采用巢式 PCR 扩增和 7 个微卫星、小卫星和多态性位点（CP47、CP56、ML2、DZ-HRGP、MSC6-5、MSC6-7 和 RPGR）测序进行分析。CP47、CP56、MSC6-5 和 MSC6-7 位点为单态性。其余位点呈多态性，在 ML2、DZ-HRGP 和 RPGR 位点分别发现了 2、3 和 2 种亚型。基于 29 个 DNA 标本在所有 8 个位点（包括 gp60 位点）成功扩增的序列信息，获得了 6 种 MLST 亚型。连锁不平衡（LD）分析显示，所研究地区兔源隐孢子虫呈克隆种群结构。STRUCTURE、邻接和网络分析表明，存在两个不同的群体，分别对应于 VbA21 亚型和 VbA28、VbA29 和 VbA32 亚型。这是首次报道兔源隐孢子虫的 MLST 分析。兔源隐孢子虫的克隆种群结构表明，黑龙江省隐孢子虫的流行并非归因于兔子的引入。因此，预防和控制策略应集中于采取更严格的措施，避免兔子个体之间的交叉传播和再感染。基于先前发现的 VbA21 亚型仅在兔子中存在，以及 VbA28、VbA29 和 VbA32 亚型均在兔子和人类中存在的结果，亚种群分析的结果可用于评估 Vb 家族中兔源隐孢子虫亚型的人畜共患潜力。这些数据将有助于探索兔源隐孢子虫感染/污染的来源归因，并了解其在研究地区人类和兔子中的传播动态。
