Preparation Of Fasciola Gigantica Diagnostic Antigen And The Establishment And Application Of Immunological Diagnosis Method For Bovine Fascioliasis

Research purpose and background:Fascioliasis is a serious hazard to animal husbandry,and the infection rate of bovine Fascioliasis is high in Guangxi.This study aims to use F22 and its component molecules as diagnostic antigens to establish an immunological diagnosis method for bovine Fascioliasis,applied to conduct an epidemiological investigation of bovine Fascioliasis in Guangxi,and preliminary evaluation of the effect of drug deworming,which will provide reference for the prevention and control of bovine Fascioliasis in Guangxi.In addition,considering the lack of bovine Fascioliasis ELISA diagnostic kits and colloidal gold rapid diagnostic test strips.This study can also lay a foundation for the future development of bovine Fasciolasis ELISA diagnostic kits and colloidal gold rapid diagnostic test strips.Research methods and results:1.In this study,F22 was prepared and identified by mass spectrometry,from which three diagnostic antigen candidate molecules Fg SAP-2,Fg Cat L2,and Fg LAP were screened and then expressed in prokaryotic cells.2.Among three molecules,r Fg SAP-2 was screened as a diagnostic antigen,and the reaction conditions were optimized together with F22to establish an indirect ELISA diagnosis method for bovine Fascioliasis.The sensitivity,specificity,stability,and early diagnostic value of the established method were tested and compared with the traditional indirect ELISA method of Fg ESP as the diagnostic antigen for bovine Fascioliasis.The results showed that the three diagnostic methods have no cross-reactivity with Dictyostelium and Babesia,with good stability.F22 showed the highest sensitivity as a diagnostic antigen,with a false negative rate of 0%.Fg ESP,F22,and r Fg SAP-2 as diagnostic antigens can all detect antibodies within 2 weeks post infection.F22 has the highest OD_450nm value at 2 weeks post infection and is extremely significantly higher than the OD_450nm value of Fg ESP and r Fg SAP-2（P<0.01）,suggesting that F22 has a higher early diagnosis value.3.In this study,F22 was successfully developed as a diagnostic antigen for bovine Fasciolasis colloidal gold rapid diagnostic test strip and proved that it has good sensitivity,specificity,and stability.The antibody can be detected 2 weeks post infection.It can be applied to the rapid diagnosis of bovine Fasciolasis.4.An indirect ELISA method using F22 and Fg ESP as diagnostic antigens to conduct an epidemiological investigation of bovine Fascioliasis in Guangxi area found that the positive rates were 68.37%and 57.30%,respectively.Using the above two indirect ELISA methods and colloidal gold test strips to test the bovine serum of the cattle ranch A the positive rates of the tests were only 6.15%,4.31%,and 2.00%,respectively.The established method was used to evaluate the effect of the fascioliasis medication regimen of the Buffalo Research Institute,which preliminarily showed that the medication regimen has a good deworming effect.Conclusion:In this study,an immunological diagnosis method for bovine Fascioliasis was established.F22 as diagnostic antigen indirect ELISA method and colloidal gold rapid diagnostic test strips have good sensitivity,specificity and stability,and can be used for early diagnosis of bovine Fascioliasis.It laid the foundation for the development of ELISA kits and colloidal gold rapid diagnostic test strips for bovine Fascioliasis in future.The established method was used to conduct epidemiological investigation of bovine Fascioliasis in Guangxi and to evaluate the effect of the medication plan of the cattle ranch A.The results showed that the infection of bovine Fascioliasis in Guangxi is relatively serious,and the medication program of the cattle ranch A has a relatively high level of efficacy.With a good deworming effect,this medication plan can be used as a reference for the herds in Guangxi.