Gaurav Rawal

In this study, we developed and validated (1) singleplex real-time RT-PCR assays for specific detection of five PRRSV-2 MLV vaccine viruses (Ingelvac MLV, Ingelvac ATP, Fostera, Prime Pac, and Prevacent) and (2) a four-plex real-time RT-PCR assay (IngelvacMLV/Fostera/Prevacent/XIPC) including the internal positive control XIPC for detecting and distinguishing the three most commonly used vaccines in the USA (Prevacent, Ingelvac MLV, and Fostera). The singleplex and 4-plex vaccine-like PCRs and the reference PCR (VetMAXTM PRRSV NA&EU, Thermo Fisher Scientific, Waltham, MA, USA) did not cross-react with non-PRRSV swine viral and bacterial pathogens. The limits of detection of vaccine-like PCRs ranged from 25 to 50 genomic copies/reactions. The vaccine-like PCRs all had excellent intra-assay and inter-assay repeatability. Based on the testing of 531 clinical samples and in comparison to the reference PCR, the diagnostic sensitivity, specificity, and agreement were in the respective range of 94.67–100%, 100%, and 97.78–100% for singleplex PCRs and 94.94–100%, 100%, and 97.78–100% for the 4-plex PCR, with a CT cutoff of 37. In addition, 45 PRRSV-2 isolates representing different genetic lineages/sublineages were tested with the vaccine-like PCRs and the results were verified with sequencing. In summary, the vaccine-like PCRs specifically detect the respective vaccine-like viruses with comparable performances to the reference PCR, and the 4-plex PCR allows to simultaneously detect and differentiate the three most commonly used vaccine viruses in the same sample. PRRSV-2 vaccine-like PCRs provide an additional tool for detecting and characterizing PRRSV-2.

Porcine respiratory coronavirus (PRCV), a mutant of the transmissible gastroenteritis virus (TGEV), was first reported in Belgium in 1984. PRCV typically replicates and induces mild lesions in the respiratory tract, distinct from the enteric tropism of TGEV. In the past 30 years, PRCV has rarely been studied, and most cited information is on traditional isolates obtained during the 1980s and 1990s. Little is known about the genetic makeup and pathogenicity of recent PRCV isolates. The objective of this study was to obtain a contemporary PRCV isolate from US pigs for genetic characterization. In total, 1245 lung homogenate samples from pigs in various US states were tested via real-time PCR targeting PRCV and TGEV RNA. Overall, PRCV RNA was detected in five samples, and a single isolate (ISU20-92330) was successfully cultured and sequenced for its full-length genome. The isolate clustered with a new group of variant TGEVs and differed in various genomic regions compared to traditional PRCV isolates. Pathogens, such as PRCV, commonly circulate in pig herds without causing major disease. There may be value in tracking genomic changes and regularly updating the diagnostic methods for such viruses to be better prepared for the emergence of variants in ecology and pathogenicity.