Investigation Of Giardia Duodenalis,Cryptosporidium And Blastocystis Infection In Some Areas Of Henan Province

Dogs are intermediate and final hosts for a variety of intestinal parasites,and the number of dogs in China is about 1 billion,with the huge number of dogs posing a potential risk to public health safety.Giardia duodenalis,Cryptosporidium and Blastocystis are common human-animal parasites that cause outbreaks of waterborne and foodborne diarrheal diseases worldwide and can infect humans and many mammals.Currently,there are no effective drugs or vaccines for these three diseases.There are few reports on intestinal parasitic infections of canine origin in China,and data on the germline development of important parasites such as Giardia duodenalis,Cryptosporidium and Blastocystis are relatively limited.In this study,we investigated intestinal parasite infections in pet hospital dogs,stray dogs and breeding dogs in six regions of Henan Province to investigate the infection and germline development of three important intestinal parasites in dogs,and to provide a theoretical basis for the prevalence and transmission of intestinal parasites of canine origin in Henan Province.In this study,1393 dog fecal samples were collected from six regions of Henan Province(Zhengzhou,Kaifeng,Anyang,Luohe,Nanyang and Puyang),and the identification of Giardia duodenalis,Cryptosporidium and Blastocystis species and genotypes was performed using PCR technique combined with sequencing and sequence analysis.The positive results detected for the three intestinal protozoa mentioned above were analyzed using statistical analysis of the data for eight factors:region,source,sex,month of age,season,diarrhea,different immunizations and deworming status to assess the variability of infection rates under different influencing factors.Neighbor-Joining(NJ)was used to perform genetic evolutionary analysis of the identified gene sequences to assess the kinship between genotypes and potential public health risks.A total of 1393 dog fecal samples were collected from 6areas and microscopic examination showed an overall infection rate of 35.39%(493/1393),8 species of parasite eggs,oocysts or encysts were detected,namely,Cysticercus isosporus 4.88%(68/1393),Strongylus 22.83%(318/1393),Toxocara4.31%(60/1393),Trichuris 3.52%(49/1393),Entamoeba 1.79%(25/1393),Giardia3.37%(47/1393),Blastocystis 1.15%(16/1393)and Cryptosporidium 0.50%(7/1393),with Cysticercus isosporus,Strongylus,and Toxocara as the dominant species.Nest PCR amplification of Giardia duodenalis based on the β-giardin(bg)gene resulted in an overall infection rate of 2.37%(33/1393);three Giardia duodenalis assembalges were identified,assembalges C(11/33),D(13/33)and E(9/33),of which assembalges C and D were at risk for zoonoses.The total infection rate was 0.72%(10/1393)based on nested PCR amplification of the SSU r RNA gene for Cryptosporidium,all are zoonotic species C.canis.The total infection rate was 0.86%(12/1393)based on PCR amplification of SSU r RNA gene for Blastocystis;DNA sequence analysis identified two Blastocystis genotypes: ST1(3)and ST3(9),both of which are zoonotic genotypes.In this study,six test puppies(five in the test group,each infected with 1×105cysts;one in the blank group)artificially infected with Giardia duodenalis assembalges D encysts were investigated by fecal examination,and the results showed that Giardia cysts were expelled intermittently with an interval of 5 days;the expulsion of cysts began on the second day of infection and reached a peak on the10 th day after Infection,and two peaks were examined,both of which were maintained for 3-4 days,with the first peak expulsion being higher than the second;the duration of cyst expulsion was 32 days,and the test dogs showed signs of depression,diarrhea and anorexia during the infection.According to the density gradient centrifugation method,the envelopes in fresh fecal samples were separated and purified using 33% Zn SO4 solution,0.85 M sucrose solution and removable filters.The purified Giardia duodenalis cysts had normal morphology,high purity and75% recovery,which is not technically demanding,low cost and simple to operate.In conclusion,dog intestinal parasite infections were more common in six regions of Henan Province;the infection rates of Giardia duodenalis,Cryptosporidium and Blastocystis were relatively low,but all identified zoonotic species or genotypes,suggesting the possibility of mutual transmission of intestinal parasites between dogs and humans.In this study,Giardia duodenalis cysts of dog origin could successfully infect the test pups,and the cysts were expelled intermittently,suggesting that cysts infection may be one of the reasons for the high infectivity of Giardia duodenalis in dogs.This study provides data to support the assessment of Giardia duodenalis,Cryptosporidium and Blastocystis infections in dogs and the risk of human-animal infections.