| Pseudorabies(PR)is a kind of acute contact infectious disease in many kinds of animals caused by pseudorabies virus(PRV).The main symptoms are nervous symptoms and death after infection in newborn piglets.Pregnant sows infected with PRV show abortion,stillbirth,mummified fetus and respiratory system symptoms,which has seriously restricted the development of pig industry at home and abroad for a long time.Since the widespread application of Bartha-K61 vaccine since the 1990 s,the epidemic situation of PR has been effectively controlled.However,since the second half of 2011,the occurrence of PR was continuously reported in pig farms with immune vaccines.In order to understand the epidemic situation of pseudorabies in some areas of Shandong province,this study carried out the pathogenic and serological detection on epidemic materials respectively,and analyzed the biological characteristics of the virus through exploring the genetic evolution analysis on the main virulence genes of the isolated strains and pathogenicity on BALB/c mice as well.From the first half of 2018 to the second half of 2019,the laboratory received 86 samples of pathogen detection and 348 samples of serum detection from farms in different regions of Shandong province.The test results showed that the nucleic acid positive rate of PRV was8.14%(7 of 86),which was lower than the PR positive rate of Shandong province reported in previous years,indicating the prevention and control measures of PR in Shandong province have been slightly effective in the past two years,and local farms pay more attention to the prevention and control of PR and scientific pig breeding.The results of serological detection showed that the positive rate of antibody against g E and g B was 22.13% and 93.68%respectively.The test of antibody agaist g E showed that there still exist wild virus infection in pig farms in Shandong province,while the test results of antibody againt g B showed that the vaccination coverage rate of PRV in pig farms from Shandong province was ideal in relative terms.In order to understand the wild virus situation of PRV in Shandong province,the positive pathogen materials from three different places were selected for isolation and identification by cell inoculation and IFA immunofluorescence test.The genetic variation of g E,g B and TKgenes of the three stains of viruses were analyzed,and the pathogenicity of the viruses were studied by inoculating them into BALB/c mice.The results showed that BHK-21 cells inoculated by three strains respectively began to show typical CPE about 24 hours after inoculation.That is,infected cells degenerated and contracted,and syncytia were observed,then cells gradually died and fell off.Compared with those in the blank control group in IFA test,the fluorescent labeled cells in the experimental group could be seen under the fluorescence,and the obvious mutations could be found in the white light,while the control group had no fluorescence or cell mutation,indicating that three PRV strains were isolated.It was found that the mice itched,licked and tore their noses for about 10 hours after challenge,and then suffered from serious depilation around the nose,followed by neurological symptoms,and died in succession after 72 hours.Among them,the mice in SD1803 strain group died first,then the mice in SD1801 strain group died,and the mice in SD1802 strain group died at the latest.In the control group,the mice were alive and asymptomatic.The virulence of the three strains were slightly different,but all of them could cause mouse death,indicating that the three strains of viruses had strong virulence.Sequence analysis showed that all of the three viruses were PRV variant strains.Genetic variation analysis showed that the mutation of g E gene was the largest,and the gene of g B and TK was relatively stable compared with that of g E gene.The genetic variation of g E gene was mainly related to the isolation time,followed by the isolation location.The genetic variation of g B gene was related to the isolation location,and the gene of TK was mainly related to isolation time. |
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