High Genetic Heterogeneity Of Giardia Duodenalis In Pre-weaned Dairy Calves In Shanghai And Xinghua,Jiangsu China

Giardia duodenalis is a zoonotic pathogen of waterborne infectious disease.Clinical manifestations of diarrhea,decreased appetite,malnutrition,growth retardation and other symptoms can present in humans and animals,even cause negative effects on human health and animal husbandry.Despite the convenience of single-locus genotyping,multilocus genotyping is better reflected the genetic heterogeneity of this pathogen.Epidemiological and molecular data are available from dairy calves in many industrialized countries,but information on genetic diversity at multiple genetic loci is limited,especially in pre-weaned dairy calves.818 and 334 fecal specimens were collected from five dairy calves farms located in suburbs of Shanghai,China,with 2-5 samplings per farm,and from one farm in Xinghua,Jinangsu province,respectively.All the specimens were screened for G.duodenalis by nested PCR(polymerase chain reaction)analysis of the bg(β-giardin)gene,the gdh(glutamate dehydrogenase)gene and tpi(triosephosphate isomerase)gene,followed by sequencing to differentiate the genotypes and subtypes,and further analyze transmission characterization.Among the 818 specimens analyzed in Shanghai,492(60.1%)pre-weaned dairy calves were positive for G.duodenalis.Three genotypes were observed,including the prevalent genotype Assemblage E(n=482),Assemblage A(n=5),Assemblage B(n=1),mixed infection of Assemblage A and E(n=4).High genetic diversity of Assemblage E were observed at each locus,showing 12,13 and 17 subtypes at the bg,gdh and tpi locus,respectively,including 4,6 and 8 novel subtypes.The dominant subtypes E3,E2,E8 at the bg locus,E3 and E1 at the gdh locus,E3 and E11 at the tpi locus shared among five farms on multiple sampling points,while no obvious different transmission characterization was noticed.Altogether,58 Assemblage E MLGs were identified,all of which had not been reported before,and seven(MLG-E1-MLG-E7)were each seen on multiple farms.These results indicate a high occurrence of G.duodenalis Assemblage E in dairy calves,the existence of high genetic heterogeneity of Assemblage E on five farms,and frequent exchange of parasite populations among farms within a small geographic area.Among the 334 fecal specimens from dairy calves in Xinghua,Jiangsu China,216(64.7%)were positive for G.duodenalis.Only the prevalent Assemblage E was detected in this study with high gentic diversity,showing 9,8 and 10 subtypes at the bg,gdh and tpi locus,respectively,including one-two novel subtypes at each locus,and 7,6,3 subtypes were identified with the subtypes observed from Shanghai,respectively.The dominant subtypes E3,E2,E6 at the bg locus,E3 and E1 at the gdh locus,E3 and E11 at the tpi locus shared among all groups.Limited differentiations of the subtype distribution were observed among all specimen groups.By MLG technique,61 Assemblage E MLGs were successfully genotype,11 had been detected in the above study in Shanghai,and five(MLG-E1-MLG-E4 and MLG-E6)were each seen on multiple farms.The characterization of high genetic diversity of Assemblage E was both observed in two studies.The common occurrence of 11 Assemblage E MLGs revealed the potential of exchange of parasite populations among all farms,while the remaining different 98 Assemblage E MLGs showed the existence of distribute differences between two regions.In conclusion,the results of the two studies improved our knowledge of the prevalent characterizations and transmission of G.duodenalis in pre-weaned dairy calves,and provided basis for preventing and controling G.duodenalis diseases.