Genotype Distribution Characteristics Of Giardia Duodenalis And Enterocytozoon Bieneusi Dogs And Cats In Guangdong

Giardia duodenalis and Enterocytozoon bieneusi are two important zoonotic pathogens causing diarrhea in humans and animals.Dogs and cats both are important hosts for G.duodenalis and E.bieneusi,but investigations of the two pathogens are very limited in dogs and cats in China.Therefore,with using molecular biology methods,this study studied the prevalence and genotype distribution characteristics of G.duodenalis and E.bieneusi in dogs and cats in Guangdong Province,where the number of dogs and cats is the largest in China.Based on the results,we assessed the risk factors and public health significance of G.duodenalis and E.bieneusi in dogs and cats in the province.Altogether,682 canine feces specimens and 424 feline feces specimens were collected from research center,shelter,pet shop,veterinary clinic,household and stray animals in Guangdong Province.In this study,nested PCR and sequencing methods were used to examine for the prevalence and genotype distribution of G.duodenalis and E.bieneusi in the collected specimens.Theχ~2 test was used to compare the infection rate of dogs and cats between different ages,genders,breeds and living conditions.At the same time,the multilocus sequence typing tool(MLST)and the analysis of population genetic structure were used to evaluate the genetic diversity and explore existence of host adaptability in the zoonotic E.bieneusi.The results showed that the infection rates of G.duodenalis and E.bieneusi in dogs were9.4%(60/641)and 31.5%(215/682),respectively.The infection rates of G.duodenalis and E.bieneusi in cats were 3.6%(15/418)and 3.5%(15/424),respectively.Theχ~2 test results showed that the infection rate of G.duodenalis in dogs in the research center(27/201,13.4%)was significantly higher than that in pet hospitals(3/109,2.8%)(χ~2=9.223,P<0.05).The infection rates of E.bieneusi was significantly higher in dogs from research center(116/222,52.3%)and shelters(71/140,50.7%)than domestic dogs(15/82,18.3%),dogs in pet shops(4/118,3.4%)and stray dogs(0/13,0.0%)(12.301≤χ~2≤80.549,P<0.05).The infection rate of E.bieneusi in cats≤6 months(6/75,8.0%)was significantly higher than that in cats>6 months(8/296,2.7%)(χ~2=4.624,P<0.05).Therefore,frequent contact and overcrowded environments in research centers and shelters are risk factors for E.bieneusi infection,and frequent contact environments are also risk factors for G.duodenalis infection.The genotype identification results showed that the genotypes of G.duodenalis detected in dogs were assemblages C(n=27),assemblages D(n=26),and 5 mixed infections of assemblages C and D.The genotypes of E.bieneusi in dogs were Pt Eb IX(n=212),WW8(n=2)and Type Ⅳ(n=1).The genotypes of G.duodenalis detected in cats were assemblages F(n=14)and assemblage A(n=1),and the genotypes of E.bieneusi were Type Ⅳ(n=12)and D(n=2).Assemblage A,genotypes Type Ⅳ and D are all zoonotic genotypes.In addition,few studies have found assemblages C,D,and F could cause human giardiasis occasionally.Therefore,dogs and cats,especially cats,have the risk of spreading G.duodenalis and E.bieneusi between humans and animals in Guangdong.The genetic diversity of the two zoonotic genotypes Type Ⅳ(n=13)and D(n=2)identified in the study was further assessed by multilocus sequence typing based one ITS and four microsatellite/minisatellite loci.Altogether,12 multilocus genotypes(MLGs)were identified.Further combined with the published MLST data of the E.bieneusi genotypes Type Ⅳ and D in humans,monkeys,foxes and raccoon dogs,population genetic analysis was performed.It was found that different subpopulations can be formed in the genotype Type Ⅳ and D isolates,and there is host adaptation among these subpopulations.The specimens from farmed monkeys and canine animals(foxes and raccoons)generated two separated subpopulations,which were both distinct from the subpopulation containing specimens from humans,cats and dogs.The above results indicate that there may be host-adapted subpopulations in the zoonotic genotypes Type Ⅳ and D.The results of the study are helpful to understand the prevalence and risk factors of G.duodenalis and E.bieneusi in dogs and cats in Guangdong Province.These will be helpful for making effective prevention and control measures for the related diseases.