Study On Prevalence And Genotyping Of Enterocytozoon Bieneusi In Black-Boned Sheep And Goats In Partial Areas Of Yunnan Province

Enterocytozoon bieneusi,an important obligate intracellular parasitic eukaryote infecting humans and livestock,causes life-threatening diarrhea in immunocomprised hosts(AIDS patients).The infection of E.bieneusi has been detected in various breeds of sheep and goats in Spain,Sweden,Brazil,Iran,Egypt and some regions of China.Currently,more than 90 genotypes were identified in sheep and goats based on the sequence of the internal transcribed spacer(ITS)of the ribosomal RNA(rRNA)gene in E.bieneusi,with at least 30zoonotic genotypes,indicating the zoonotic potential of these animals transmitting E.bieneusi to humans.Black-boned sheep and goats are important local species originating from Nanping county of Yunnan province,with extremely high nutritional and medicinal values.However,no studies on E.bieneusi infection in black-boned sheep and goats were available prior to the present study.Here,a total of 325 and 336 faecal specimens were respectively collected from black-boned sheep and goats in Shilin,Yongsheng,Nanping,Meishui and Shanshu of Yunnan province.The prevalence and genotypes of E.bieneusi in these samples were determined using the molecular biological method based on the ITS gene,and the genetic diversity of E.bieneusi isolated in our study were analysed using the multi-locus genotyping tool(MLST)based on micro-and mini-satellite gene loci.1.The total prevalence of E.bieneusi in balck-boned sheep and goats was 10.59%(70/661).The prevalence of E.bieneusi in black-boned sheep(12.31%,40/325)was higher than that(8.93%,30/336)in black-boned goats,but without statistically significant difference(?~2=1.781,P>0.05).E.bieneusi was found in each sampling site,and the differences in prevalence were significant(?~2=9.629,P<0.05)among five investigated locations,with the highest in Nanping(14.79%,21/142),followed by Meishui(14.75%,9/61),Shilin(12.24%,24/196)and Shanshu(7.75%,11/142),and the lowest in Yongsheng(4.17%,5/140).E.bieneusi was also detected in all four age groups,with significant(?~2=18.59,P<0.001)differences,and the prevalence decreased with the age.2.Sequence analysis revealed 12 genotypes of E.bieneusi in balck-boned sheep and goats,including 10 known(BEB6,COS I,SX1,CM21,CHG3,CHG1,CHS5,EbpC,COS II,PigEb4)and two novel genotypes(named as YNS1 and YSM1),with the BEB6 as the predominant genotype in both black-boned sheep and goats.Phylogenetic analysis showed that the genotypes EbpC,CHS5 and PigEb4 were clustered into the zoonotic group 1 with zoonotic potential,while the remaining genotypes were clustered into the group 2.MLST analysis identified 12 distinct multilocus genotypes(MLG15~MLG26)in black-boned sheep and goats,with 11 MLGs in black-boned sheep and only one in a black-boned goat.Collectively,this study revealed the presence of E.bieneusi in black-boned sheep and goats,and the prevalence of E.bieneusi was closely related to the age and the sampling site.Furthermore.Additionally,zoonotic potential and genetic diversity of E.bieneusi were also observed in black-boned sheep and goats.These findings provided fundamental data for controlling E.bieneusi infection in balck-boned sheep and goats as well as other hosts in this province.