Establishment Of A Method For Testing The Relative Efficacy Of Highly Pathogenic Porcine Reproductive And Respiratory Syndrome Inactivated Vaccines

Porcine reproductive and respiratory syndrome(PRRS)is an acute,highly contagious infectious disease that infects pigs of all ages,especially pregnant sows and piglets.It has become a major epidemic affecting the global pig industry,and this disease has also become one of the main infectious diseases threatening the development of China’s pig industry in recent years.PRRSV belongs to the genus Arteritis virus of the arteriitis family Arteritis Virus family,which is a single-stranded positive-stranded RNA virus with capsule and no segments,which is a huge challenge to prevention and control due to the continuous mutation of the virus.Since porcine reproductive and respiratory syndrome virus only infects pigs,the evaluation method of inactivated vaccine is mainly based on the animal(pig)challenge and ELISA antibody determination,and there is no alternative to experimental animals,resulting in a long evaluation period of inactivated vaccine efficacy,and the results are also easily affected by the quality of test pigs.The efficacy evaluation of vaccines is mainly through in vivo animal challenge protection test,immune animal antibody determination and in vitro substitution test,but the in vivo efficacy test of animals has the disadvantages of long test cycle,large impact of individual differences on test results,and difficulty in finding negative pigs.The aim of this study is to establish an accurate and efficient method for evaluating the relative efficacy(RP)of inactivated vaccines.PRRSV-RP19 strain was inoculated on Marc-145 cells to prepare PRRSV inactivated vaccine;The quality inspection,safety evaluation and efficacy evaluation of the tested vaccine were carried out,and the reaction conditions of the test were screened and optimized by the established method on the basis of the double antibody sandwich ELISA method,and finally determined that when the capture antibody was diluted 10 μg/m L,the detection antibody was diluted 40 times,the secondary antibody was diluted 1:8000,and the color development temperature was 37 °C,the reaction was the best,and the established method was used to control swine fever virus(CSFV),Japanese encephalitis virus(JEV),Porcine epidemic diarrhea virus(PEDV),porcine circovirus type II(PCV2),porcine pseudorabies virus(PRV),seedling adjuvant,etc.were tested,and the results were all negative,indicating that the established method had good specificity.Three batches of antigens were selected and diluted to 1:128 by 2-fold ratio to detect their sensitivity,and the results showed that the established method had good sensitivity.Three batches of inactivated vaccines with different antigen content that passed quality inspection and safety inspection were selected for reproducibility test,and the OD450 nm value was used as the reaction index,and the relative efficacy(RP value)of the inactivated vaccine was evaluated after statistical analysis by Relpot4.0 software,and the results showed that the intra-batch variation coefficient was 4.8% and the between-batch coefficient of variation was 2.0%,indicating that the method had good stability.The correlation between the relative efficacy RP value and antigen content of the vaccine was analyzed,and the relationship with animal immunization tests was analyzed,and the results showed a positive correlation,in order to replace animal experiments for rapid evaluation of the immune efficacy of porcine reproductive and respiratory syndrome vaccine as soon as possible.