Studies Of Population Structure And Molecular Epidemiologic Investigation Of Four Intestinal Protozoa In Diarrheic Pigs In Three Southern Provinces Of China

Enterocytozoon bieneusi,Cryptosporidium spp.,Entamoeba spp.,and Blastocystis sp.are four common zoonotic pathogens with a wide host range that can cause widespread epidemics in several countries and regions.Diarrhea is one of the common characteristic of these four diseases,but previous reports on the epidemiological and subtypes/genotypes of these four intestinal protozoa based on diarrhea were very limited.The aim of this study was to investigate the infection of these four intestinal protozoa in diarrheic pigs in Jiangxi,Hunan and Fujian provinces,and to analyze their epidemiological distribution in diarrheic pigs,which was important for the prevention and control of related diseases locally.In this study,the epidemiological characteristics and species/genotypes of Enterocytozoon bieneusi,Cryptosporidium spp.,Entamoeba spp.and Blastocystis sp.in 1254 fecal samples from pigs with diarrhea were identified based on nested PCR.The results showed that the total infection rate of Enterocytozoon bieneusi in pigs with diarrhea was 5.74%,Cryptosporidium was 4.55%,Entamoeba was 53.83% and Blastocystis was 31.42%.Sequence analysis of Enterocytozoon bieneusi revealed that the 72 Enterocytozoon bieneusi-positive samples belonged to 13 genotypes,including8 known genotypes and 5 newly identified genotypes.Cluster analysis revealed that all identified genotypes were grouped into the zoonotic Group1.A total of 57 samples were found to be Cryptosporidium-positive,belonging to Cryptosporidium suis and Cryptosporidium scrofraum.The 732 Entamoeba-positive samples were found to consist of Entamoeba suis and Entamoeba polecki.Genotyping of E.polecki revealed two genotypes,ST1 and ST3.In this study,394 positive samples of Blastocystis were found.Four subtypes with zoonotic potential(ST1,ST3,ST5 and ST14)were deceted.In this study,multilocus sequence typing(MLST)was also applied to genotype all samples identified as Enterocytozoon bieneusi-positive.The results showed that only29 of the 72 Enterocytozoon bieneusi-positive samples were successfully amplified and sequenced at MS1,MS3,MS4 and MS7,and forty-one multi-locus genotypes MLGs were detected.The statistical analysis software SPSS 26 was applied to analyze the prevalence of four intestinal protozoa with respect to the risk factors of age groups and regions.The results showed that geographic and age factors had a significant effect on the prevalence of Enterocytozoon bieneusi and Blastocystis,while for Cryptosporidium spp.and Entamoeba spp.