Development of a quadruple PCR-based gene microarray for detection of vaccine and wild-type classical swine fever virus, African swine fever virus and atypical porcine pestivirus.

BACKGROUND: Classical swine fever (CSF), African swine fever (ASF), and atypical porcine pestivirus (APPV) are acute, virulent, and contagious viral diseases currently hampering the pig industry in China, which result in mummification or stillbirths in piglets and mortality in pigs. Diagnostic assays for the differentiation of infection and vaccination of CSFV, in addition to the detection of ASFV and APPV, are urgently required for better prevention, control, and elimination of these viral diseases in China. METHODS: A quadruple PCR-based gene microarray assay was developed in this study to simultaneously detect wild-type and vaccine CSFV strains, ASFV and APPV according to their conserved regions. Forty-two laboratory-confirmed samples, including positive samples of 10 other swine viral diseases, were tested using this assay to confirm its high specificity. RESULTS: This assay's limit of detections (LODs) for the wild-type and vaccine CSFV were 6.98 and 6.92 copies/µL. LODs for ASFV and APPV were 2.56 × 10 and 1.80 × 10 copies/µL, respectively. When compared with standard RT-PCR or qPCR for CSFV (GB/T 26875-2018), ASFV (MARR issue No.172), or APPV (CN108611442A) using 219 clinical samples, the coincidence was 100%. The results showed that this assay with high sensitivity could specifically distinguish ASFV, APPV, and CSFV, including CSFV infection and immunization. CONCLUSION: This assay provides a practical, simple, economic, and reliable test for the rapid detection and accurate diagnosis of the three viruses and may have good prospects for application in an epidemiological investigation, prevention, and control and elimination of these three diseases.

Serologic evidence of prevalent avian H3 subtype influenza virus infection in chickens.

H3-subtype influenza viruses are known to infect avian and mammalian species, including humans. However, little is known about the prevalence of H3 influenza virus infection in chicken populations in China. Therefore, a serologic survey of chickens was conducted in China to investigate the seroprevalence of avian H3-subtype influenza virus. Anti-H3 antibodies were assayed by using hemagglutination inhibition (HI) and confirmatory virus neutralization (VN) testing of 4598 serum samples, collected between July 2006 and June 2007, from 173 chicken flocks located in 18 areas that included 16 provinces and two municipalities. Seroepidemiologic results indicated that avian H3-subtype viruses were circulating in chickens in some regions of China, regions that included 12 of the 18 test areas, with an overall average prevalence rate of 2.83%. Samples from 44 of 173 flocks were HI/VN seropositive, including 15 flocks with levels that ranged from 10.00% to 41.94%. Significantly higher seroprevalence rates were observed in older chicken flocks and in those sampled in the cooler seasons. Standardized comparisons showed that Guangdong and Jiangsu, located in the south and east of China, respectively, had significantly higher levels of H3 seropositivity. For the first time, these results demonstrated serologic evidence for H3 avian influenza virus infection in chicken populations in several locations throughout China. These observations highlight the need for continued epidemiologic surveillance of the H3 subtype and for other low-pathogenic avian influenza viruses in China and other regions.

Genetic analysis of H3 subtype influenza viruses isolated from domestic ducks in northern China during 2004-2005.

The broad distribution and prevalence of H3 subtype influenza viruses in avian and mammalian hosts constitutes a global threat to both human and veterinary health. In this present study, six H3N8 influenza viruses isolated from domestic ducks during 2004-2005 in northern China were genetically and phylogenetically characterized. Sequence analysis showed that HA, NA, and M genes of all the six H3N8 isolates had a close relationship with those of Equine/Jilin/1/89 (H3N8) virus, which once caused outbreak in equine populations in northern China. The PB2 and PA genes of the viruses possessed the highest similarities with highly pathogenic avian H5N1 influenza viruses currently circulating in this region. These findings emphasize the importance of avian influenza virus surveillance in this region for understanding the genesis and emergency of novel reassortants with pandemic potential.