Analysis Of The Epidemic Variation Of PCV2 And PRV In Jilin Province And The Mechanisms Underlying Their Induction Of Cell Apoptosis

Porcine circovirus 2 is the most widely distributed porcine circovirus.It is the pathogenic agent responsible for porcine circovirus diseases or porcine circovirus associated diseases.It has caused serious disease and economic losses worldwide.Pseudorabies is a highly contagious viral disease caused by pseudorabies virus and is also one of the most important diseases threatening the pig industry.It has been reported that PRV variation and recombination might contribute to the persistent prevalence of PR in China.The diseases caused by PCV2 and PRV infection were always in the top several of porcine virus diseases.PCV2 infection can cause subclinical symptoms,leading to immunosuppression,which creates good conditions for co-infection and/or secondary infection with other pathogens.Co-infection of PCV2 with multiple viruses poses a greater challenge for disease prevention and control.Research showed that mixed infections of PCV2 and PRV were increasing.The piglets with PRV and PCV2 mixed infection showed characteristic clinical symptoms such as "fever-wasting-neurological dysfunction-vomiting-death",and all the piglets died on the 14 th day after infection,which undoubtedly brought great difficulties to the diagnosis and control of PCV2 and PRV.Therefore,comprehending the prevalence and pathogenesis of PCV2 and PRV co-infection is of great significance for the prevention and treatment of related diseases.In order to clarify the prevalence of PCV2 and PRV in Jilin Province in recent years,clinical samples of pigs were collected for investigation and analysis of porcine viruses,genetic evolution and protein structure.The results showed that PCV2 was widely present in pig farms in Jilin Province and there was co-infected with PRV.The genetic evolution analysis showed that there were regional differences in the distribution of PCV2 genotype in China,and PCV2 d gradually surpassed PCV2 b and became the dominant genotype in China.However,PCV2 b was still the dominant genotype in Jilin Province from 2016 to 2021,followed by PCV2 e and 2d.Mutation was detected in PCV2,but no recombination occurred,indicating that the PCV2 genotype in Jilin Province was stable for many years.The 18 PCV2 included 9 PCV2 b strains,3 PCV2 d strains,and 6 PCV2 e strains.The nucleotide and amino acid sequences of the full-length PCV2 genome and ORF2 were more suitable for genotyping and evolutionary analysis.Compared with the three domestic commercially inactivated vaccines,18 PCV2 showed changes in the B cell epitopes of Cap and Rep proteins and T cell epitopes of Cap proteins,but did not affect their spatial conformation.Whether the mutation affected their function needs further analysis.The genetic evolution analysis of PRV revealed that the PRV strains mainly had two genetic clades,which further explained why the Bartha-K61 vaccine(belonging to clade 1)could not provide complete protection against the current domestic epidemic strain(belonging to clade 2).The PRV JL-CC strain isolated in this study belonged to an epidemic variant strain,and g D,g B and TK mutations had no effect on the spatial conformation,but might affect the recognition of antigen epitopes.Therefore,there was a need for continuous surveillance and the development of PRV vaccines against specific variants.Endoplasmic reticulum stress,autophagy and apoptosis are the mechanisms that maintain intracellular homeostasis and play an important role in virus infection.The transcriptome results of porcine kidney cells(PK-15)infected with PCV2 and PRV single and co-infection showed that PCV2 and PRV single and co-infection caused endoplasmic reticulum stress,autophagy and apoptosis.The effect of co-infection of PCV2 and PRV on the proliferation and cell activity after virus infection was similar to that of the group infected successively with PCV2 and PRV.PRV inhibited the proliferation of PCV2 at 36-72 hours after infection,while PCV2 enhanced the proliferation of PRV at 24-36 hours post infection.PRV played a leading role in the co-infection process.Through further analysis and verification of differential genes involved in transcriptome analysis,it was found that PCV2 and PRV infection alone and co-infection activated PERK-e IF2α-ATF4-CHOP endoplasmic reticulum stress pathway and promoted virus replication;PRV infection single and co-infection with PCV2 and PRV could activate the IRE1 pathway,but PCV2 and PRV infection did not activate the ATF6 pathway.In addition,PCV2 and PRV infection single and co-infection coule also activate autophagy and mitochondrial dominated cell apoptosis.These results suggested that endoplasmic reticulum stress,autophagy,and apoptosis were related to PCV2 and PRV infection alone and together.The PERK-ATF4-CHOP axis was one of the key pathways of endoplasmic reticulum stress,apoptosis,and autophagy.In summary,this study enriched the epidemiological data of PCV2 and PRV in Jilin Province,and provided scientific theoretical insights for the comprehensive prevention and control.Moreover,the investigation of potential links between endoplasmic reticulum stress,autophagy,and apoptosis during PCV2 and PRV infection of PK-15 cells revealed new targets and insights for future research on virus-host interactions.Lastly,this study laid a solid foundation for further antiviral research and applications.