Isolation,Identification And Genetic Evolution Analysis Of Porcine Teschovirus In Yunnan Province

Porcine teschovirus（PTV）belongs to a member of the enterovirus genus of Picornaviridae,containing 13 serotypes,which is one of the important diseases that infect pigs.PTV infection can cause neurological related symptoms such as ataxia,limb paralysis and muscle twitching by damaging the central nervous system of pigs,It also causes many symptoms,such as reproductive disorders,pneumonia,myocarditis and diarrhea.Currently,PTV has been widely spread to Asia,the Americas,Africa,and Oceania.PTV is often mixed with other diarrhea viruses,and it is latent,which should be paid attention to by the pig-raising industries.This study on PTV infection in Yunnan province by designing PTV specific primers,and the results showed that the PTV positive rate was 34.09%,and PTV infection with different genotypes existed in the same pig farm,and the regional prevalence was not obvious.Meanwhile,triple RT-PCR detection methods for PTV,PSV and PEDV were established to investigate the mixed infection of these three virus.The results of mixed infection in clinical samples found that,PTV+PSV was 8.33%（11/132）,PEDV+PTV was 6.82%（9/132）,PEDV+PSV was 3.79%（5/132）,and PTV+PSV+PEDV was1.51%（2/132）.To analyze the genetic variation and evolutionary relationship of PTV in Yunnan Province,the PTV-positive samples were inoculated in PK-15 cells,a PTV strain was successfully isolated after serial passage.The results showed that the TCID₅₀of YN01-2023 virus strain was 3.16×10⁷/m L.Meanwhile,seven primer pairs were designed to amplify the whole gene sequence of YN01-2023 strains,and the sequencing results were analyzed by genetic evolution and recombination with the whole gene sequence of 34global PTV referencestrains.The results showed that the whole genome nucleotide sequence of YN01-2023 strain was 6987 bp and encoding 2329 aa.Phylogenetic tree analysis showed that this strain is on the same clade as PTV-4 type and is closely related.Recombination analysis showed that the main parent of YN01-2023 was the Hunan strain,PTV-9 Hu N 1,and the minor parent was PTV-4 Vir 918-19/85.The length of the capsid protein P1 gene in the YN01-2023 strain is 2576bp,encoding 859 aa,Compared with the four reference strains on the same branch,the results showed that the P1 protein of YN 01-2023 strain has the highest nucleotide sequence identity with the domestic PTV-4Hu N 21 strain（MF170925）.The protein secondary structure prediction showed that P1protein is a moderately flexible,low hydrophilic protein,and the dominant epitope of capsid protein is in the VP1 fragment.