Molecular Epidemiological Investigation Of Cryptosporidium Spp. In Sheep And Goats In Midwest China And Development Of A Subtyping Tool For Cryptosporidium Xiaoi

Cryptosporidium spp.are important zoonotic protozoa.Cryptosporidiosis can cause diarrhea and even death in young animals,including lambs or goat kids.Cryptosporidium xiaoi is the dominant species infecting sheep and goats.However,little is known about the genetic characteristics of C.xiaoi.The present study aims at investigating the prevalence of Cryptosporidium spp.in sheep and goats in midwest region of China,developing a subtyping tool for C.xiaoi,and analyzing its genetic characteristics.We collected 646 fecal specimens from sheep on five farms in Anhui,Henan and Qinghai,and 402 fecal specimens from goats on four farms in Anhui.Nested PCR and sequence analyses of the SSU r RNA and gp60 genes were used for genotyping and subtyping of Cryptosporidium spp.,respectively.The overall infection rate of Cryptosporidium spp.in sheep and goats was 39.0%(409/1048).The infection rate of Cryptosporidium in goats(46.5%,187/402)was significantly higher than that in sheep(34.4%,222/646,P < 0.01).A comparison between different provinces found that the infection rates of Cryptosporidium spp.in Henan(55.4%,92/166)and Anhui(66.9%,87/130)were significantly higher than in Qinghai(12.3%,43/350,P < 0.01).In addition,results of statistical analyses showed that farms and ages were associated with variations in Cryptosporidium infection rates.Three Cryptosporidium species were detected,including C.xiaoi(n = 396),C.ubiquitum(n = 12)and C.parvum(n =1).The subtypes of the latter two species were identified as c XIIa and IId A19G1,respectively.As these two subtypes are common human pathogens,Cryptosporidium spp.in sheep and goats have zoonotic potential.Thus far,the whole genome sequence of C.xiaoi is not available.Based on the whole genome sequence of C.bovis,which is genetically related to C.xiaoi,we developed a gp60 gene-based subtyping tool for C.xiaoi based on sequences of conserved regions of the gene.By using this tool,a total of six subtype families(XXIIa～XXIIf)were identified,without obvious host and geographic segregation among them.In addition,some C.xiaoi-positive samples produced multiple bands with different lengths in gp60 PCR,suggesting that there were concurrent infections of different C.xiaoi subtype families in sheep and goats.In conclusion,data from this study revealed the epidemiological characteristics of Cryptosporidium spp.in sheep and goats in the midwest region of China,and the newly established subtyping tool for C.xiaoi should be useful in molecular epidemiological studies of this common Cryptosporidium species in small ruminants.