| PRRSV is currently one of the most important pathogens threatening global pig production and became endemic in Europe and the Americas in the 1990 s.The virus was first isolated in China in 1996,and since then PRRSV has become widespread in China.2006 saw the emergence of highly pathogenic PRRSV in China and it quickly became the dominant epidemic strain in China.2012 saw the emergence of NADC30-like strain and recombinant strain in China,and NADC30-like strain spread rapidly and caused serious losses to the Chinese pig industry.In 2017,the NADC34-like strain was isolated for the first time in China.Currently,the NADC34-like strain is mildly or moderately pathogenic to piglets and mainly affects sows,often leading to severe abortions in pregnant sows,which affects the healthy development of China’s pig farming industry.To understand the current epidemic and recombination of PRRSV in Shandong Province,samples of suspected PRRSV infection cases were collected from pig farms of different sizes in different cities of Shandong Province,and the genetic evolution,major amino acid mutations and homology of GP5 and Nsp2 genes were analyzed by clinical symptom observation,dissecting lesion observation,pathological histological examination,PRRSV virus isolation and purification,RT-PCR pathogen detection.The recombinant strains of GP5 and Nsp2 were analyzed for whole-genome sequence determination and recombination.In addition,sera from clinically manifest normal pigs in different sizes of pig farms in Shandong Province were collected in this study and tested for RRSV antibodies by ELISA.From 2021 to 2022,a total of 1860 samples with suspected PRRSV infection were collected from all over Shandong,and the RT-PCR results showed that 426 samples were positive for PRRSV,with a positive rate of 22.90%,and the highest rate of mixed infection was found of PRRSV and PCV2.The highest PRRSV positivity rate was found in autumn and winter,and the overall PRRSV positivity rate in 2022 was slightly higher than that in2021.GP5 genetic evolutionary analysis revealed that the predominant strain type of PRRSV was NADC30-like in Shandong Province.NADC34-like was also prevalent in Shandong Province.NADC34-like was second only to the NADC30-like strain in terms of prevalence.The ORF5 nucleotide homology between the isolated strains and the reference strains of CH-1a,VR2332,JXA1,NADC30,NADC34 and QYYZ in this study were 88.3%-99.2%,85.8%-99.3%,85.3%-99.5%,79.4%-96.0%,87.1%-96.0%,76.9%-84.9%,and the GP5 amino acid homology was 86.1%-98.5%,80.6%-99.1%,82.1%-98.5%,79.6%-95.0%,88.1%-96.0%,and 75.7%-85.6%,respectively.Some isolates had mutations in amino acids13,151 and the neutralizing epitope(aa36-aa52)of ORF5,which may lead to changes in virulence and the ability of the virus to escape immunity.Genetic evolutionary analysis of Nsp2 revealed that 35.19% of the PRRSV strains prevalent in Shandong Province underwent recombination of Nsp2 with GP5.The Nsp2 nucleotide homology of the isolates in this study and the reference strains CH-1a,VR2332,JXA1,NADC30,and NADC34 were75.3%-87.5,74.8%-88.6%,79.5%-87.5%,74.1%-85.0%,76.5%-89.1%,and 76.9%-84.9%,respectively.Nsp2 amino acid homology was 76.3%-86.4%,73.7%-83.5%,79.7%-88.5%,78.6%-86.6%,and 76.1%-88.8%,respectively,with a greater degree of mutation in the Nsp2 region.The full-length gene sequence determination and recombination analysis of GP5 and Nsp2 recombinant strains resulted in the isolation of two strains,SDHY-DZ037 and SDHY-WF138,respectively.The recombination analysis revealed that both strains were based on Lineage 1 as the primary parent,SDHY-DZ037 was recombinant with NADC30-like strain and NADC34-like strain,SDHY-WF138 was based on NADC30-like as the primary parent and NADC34-like and VR2332-like as the secondary parents.There were complex restructuring regions and diverse restructuring models.ELISA antibody determination was performed on 4535 normal pig sera collected in2021-2022,and the antibody positivity rate was 81.01%,and the antibody positivity rate in different stages of pigs was also above 75%.In summary,the pressure of PRRSV infection in pig farms in Shandong Province is high,although most pig farms use PRRSV vaccine for high-intensity immunization,but due to the many types of strains,the commercial vaccines on the market today have poor protection against non-homologous strains,coupled with frequent recombination of PRRSV and various types of recombination,making the immunization effect of the vaccine unsatisfactory.Some immunized pig farms still have cases of disease that cause serious economic losses,so it is necessary to strengthen the epidemiological investigation of PRRSV and improve the biosafety system.This study further enriches the epidemiological data of PRRSV in Shandong Province and lays a theoretical foundation for the vaccine development and immunization prevention and control of the disease. |
