Serological Investigation And Genetic Variation Analysis Of Swine Japanese Encephalitis Virus In Liaoning Area

Background:The Japanese encephalitis virus(JEV)is mainly distributed in southeast Asia and the western Pacific Ocean region.About tens of thousands of people are infected with Japanese encephalitis virus every year.The mortality rate is about 25%.Japanese encephalitis is an acute infectious disease of the central nervous system,which is a serious threat to the health of human beings and animals,mainly transmitted by Culex tritaeniorhynchus.Swine is an important storage host and proliferative host of Japanese encephalitis virus.It is the main source of infection of Japanese encephalitis.Japanese encephalitis can cause reproductive disorders such as abortion,stillbirth or mummification of pregnant sows.It can also cause acute orchitis or infertility in boars.The virus usually circles in "mosquito-swine-mosquito" between animals.The research on the Japanese encephalitis in Liaoning province is not thorough enough.Therefore,it is necessary to carry out serological detection of Japanese encephalitis virus and research on molecular biology.Objective:This study was conducted to understand the infection of Japanese encephalitis virus in host animal swine in some parts of Liaoning province by serological detection.Based on the isolation and identification of Japanese encephalitis virus,the genotype,major genetic variation and genetic evolution of Japanese encephalitis virus strains from swines were analyzed,providing a theoretical basis and technical support for further prevention and control of Japanese encephalitis.Methods:The serum Ig G antibody of Japanese encephalitis virus was detected by enzyme linked immunosorbent assay(ELISA).Detection of antibody negative swine serum samples for cell culture(antibody positive indicates that the body has been infected,after infection,the virus is suppressed and neutralized,and the virus can no longer be isolated).The virus RNA was extracted from the diseased cells and RT-PCR was identified by specific primers of Japanese encephalitis virus,and the positive PCR products were sequenced.The results of sequencing compare with those of reference genes in Japanese encephalitis virus in Genbank and construct the genetic evolution tree of Japanese encephalitis virus.Result:1.In 2018,a total number of 347 swine serum samples were collected from parts of Liaoning province for the detection of Japanese encephalitis virus serum Ig G antibody.195 of them were positive for Japanese encephalitis virus serum Ig G antibody,with a positive rate of56.2%.It provides a data base for determining the infection status of Japanese encephalitis virus in swine herds.2.A type GI Japanese encephalitis virus was isolated from 152 negative swine serum Ig G antibody samples.Gene sequencing of Japanese encephalitis virus products identified by RT-PCR and analyzed by using biological software such as DNAStar.The nucleotide homology of the isolate was 77.3%-99.3% and amino acid homology was 91.0%-99.8% with the representative strains of Japanese encephalitis virus E gene in Genbank at home and abroad.The nucleotide sequence homology of the E gene of the vaccine strain SA14-14-2 was87.8%,and the coding amino acid homology was 97.2%.There were 13 amino acid mutations in Domain region and 1 amino acid mutation outside Domain region of E gene of this strain and SA14-14-2 vaccine strain.This is the first time that the Japanese encephalitis virus was isolated from swine blood in our province.It laid the foundation for the research on the molecular biological characteristics of the Japanese encephalitis virus,and is beneficial to the development of Japanese encephalitis vaccine,the protection of human and livestock health and the economic benefit of the swine industry.Conclusion:The infection rate of Japanese encephalitis is higher in swine herd in Liaoning province.The serological detection of the swine herd should be strengthened,and the infection of swine encephalitis should be grasped in time.Compared with the E gene of the vaccine strain SA14-14-2,the GI Japanese encephalitis virus isolated has a total of 14 amino acid site mutations,it is prompted that some of the nucleotides of the Japanese encephalitis virus isolated from this isolate are mutated.It is of great significance to study the molecular biological characteristics of Japanese encephalitis virus,to prevent the occurrence and epidemic of Japanese encephalitis,to ensure human and animal health,and to promote public health and safety.