Molecular Detection of Porcine Parainfluenza Viruses 1 and 5 Using a Newly Developed Duplex Real-Time RT-PCR in South Korea

SIMPLE SUMMARY: In the present study, we first developed a highly specific and sensitive TaqMan probe-based duplex real-time reverse transcription polymerase chain reaction (dqRT-PCR) assay for simultaneous and differential detection of porcine parainfluenza virus 1 (PPIV1) and PPIV5 in a single reaction. The developed dqRT-PCR, with high sensitivity, specificity, and accuracy, was useful for the detection of PPIV1 and PPIV5 in clinical pig samples, and the diagnostic sensitivity was higher than that of the previous qRT-PCR for PPIV1 and consistent with that of the previous qRT-PCR for PPIV5. Using the dqRT-PCR, we demonstrated that PPIV1 and PPIV5 are co-circulating in Korean pig herds, and subsequent phylogenetic analysis suggested that genetically diverse PPIV1 strains are presented in Korean pig herds. The new dqRT-PCR developed in this study will be a valuable tool for diagnosis and further studies for PPIV1 and PPIV5. ABSTRACT: Two species of porcine parainfluenza viruses (PPIV), PPIV1 and PPIV5, are globally distributed in pig herds and associated with porcine respiratory diseases, and a diagnostic tool for the simultaneous detection of the two viruses is required. In this study, a TaqMan probe-based duplex real-time reverse transcription polymerase chain reaction (dqRT-PCR) assay was first developed for the differential detection of PPIV1 and PPIV5 nucleocapsid protein (NP) genes in porcine clinical samples. The dqRT-PCR assay was highly sensitive, its limit of detection was approximately 10 RNA copies/reaction, it specifically amplified the targeted NP genes of PPIV1 and PPIV5 without cross-reacting with other porcine pathogens, and their clinical detection rates were 15.2% and 0.7%, respectively. The results from 441 clinical samples taken from 278 Korean domestic pig farms showed that the prevalence of PPIV1 and PPIV5 was 11.2% and 1.1%, respectively, and co-infection of both viruses was confirmed in a farm, suggesting that PPIV1 and PPIV5 are co-circulating in current Korean pig herds. Phylogenetic analysis based on the partial NP genes suggested that genetically diverse PPIV1 strains are circulating in Korean pig herds. The developed dqRT-PCR assay was found to be an accurate, reliable, and quantitative detection tool for PPIV1 and PPIV5 RNA in clinical pig samples and will be useful for etiological and epidemiological studies and the control of viral infections in the field.