Investigation Of Toxoplasma Gondii,neospora Caninum Of Pigs And Genotyping Of T.gondii In Hunan Province

Both Toxoplasma gondii and Neospora caninum can infect pigs,resulting the farrowing rate of pigs and seriously threatening the health of humans and animals.To ascertain the prevalence of T.gondii and N.caninum in Hunan Province and to determine the genotype of T.gondii.In this study,indirect hemagglutination test(IHAT)and enzyme-linked immunosorbent assay(ELISA)were used to detect Toxoplasmosis and Neosporidiosis of pigs in Hunan province,and nested PCR was used for molecularly detected of T.gondii and N.caninum,and the genotype of T.gondii in Hunan province was identified by PCR-RFLP.The results are as follows:(1)Serum infection of T.gondii and N.caninum in pigs in Hunan Province: A total of1500 pigs serum samples were collected from January 2017 to December 2018 in Hunan Province.Detection of pigs serum by IHAT and ELISA.A total of 87 and 28 positive samples of T.gondii antibody and N.caninum antibody were detected,and the overall positive rates were 5.8%(87/1500)and 1.9%(28/1500).Among them,The seroprevalence of T.gondii and N.caninum in different regions were 2.1% ～ 7.7% and 0.3% ～ 4.6%,respectively.The risk of T.gondii infection was highest in pigs in central Hunan(OR=3.9,95%CI=1.5-10,P<0.01).The risk of N.caninum infection was highest in pigs in Southern Hunan(OR=19.1,95%CI=2.4-148.7,P<0.01).The seroprevalence of T.gondii and N.caninum in different seasons were 1.9% ～ 9.7% and 0% ～ 2.6%,respectively.The risk of T.gondii infection was highest in pigs in winter(OR=5.5,95%CI=1.6-18,P<0.01).The seroprevalence of T.gondii and N.caninum in different production stages were 2.3% ～11.8% and 0.6% ～ 2.8%,respectively.The risk of T.gondii infection was highest in sows(OR=5.7,95%CI=2.8-11.7,P<0.01).The risk of N.caninum infection was highest in sows(OR=4.7,95%CI=1.1-20.9,P<0.05).(2)Molecular identification of T.gondii and N.caninum in pigs in Hunan Province: A total of 339 pigs brain tissue samples were collected from April 2015 to December 2017 in Hunan Province.A total of 34 positive DNA samples were detected from the DNA of pig brain plasma samples after PCR detection of T.gondii B1 gene,and the positive rate was about 10%(34/339).A total of 5 positive DNA samples and 8 positive DNA samples were detected from the DNA of pig brain plasma samples after PCR detection of N.caninum NC-5 coding gene and ITS-1 sequence,the positive rates were 1.8%(5/339)and 2.4%(8/339),respectively.The NC-5 coding gene and ITS-1 sequence sequencing results showed that all positive sequences were the target fragment sequence of N.caninum.(3)Genotype of T.gondii in pigs in Hunan Province: This study is based on the detection results of T.gondii B1 gene in pig brain plasma DNA samples.The 34 positive DNA samples were typed at 12 genetic markers(SAG1、5’-SAG2、3’-SAG2、SAG3、BTUB、GRA6 、 C22-8 、 C29-2 、 L358 、 PK1 、 SAG2 and Apico)by polymerase chain reaction-restriction fragment length polymorphism(PCR-RFLP)technology to confirm genetic characterization.The results showed that only two samples successfully amplified all the genetic markers,one of which was finally identified as Toxo DB#61,and the other one was finally identified as Toxo DB#10.In conclusion,through the serological and molecular investigations of T.gondii and N.caninum in Hunan Province,the two parasite infections existed in pigs in Hunan Province,but the infection rate was low.Through the identification of T.gondii genotype,Toxo DB#61was identified for the first time in pigs,indicating that this genotype T.gondii can also infect pigs.These results not only fills the blank of the research on T.gondii and N.caninum in pigs in Hunan Province,but also provided the theoretical basis for the prevention and control of Toxoplasmosis and Neosporosis.