In vitro evaluation of porcine reproductive and respiratory syndrome virus (PRRSV) ORF5 sequences in samples containing PRRSV modified-live vaccine and wild-type strains.

Porcine reproductive and respiratory syndrome (PRRS) causes significant economic losses and is a major challenge to the swine industry. The PRRS virus (PRRSV) has high rates of mutation and evolution. We examined the influence on ORF5 Sanger sequencing outcomes of various concentrations of a wild-type (WT) PRRSV (lineage 1A RFLP 1-7-4) and a modified-live vaccine (MLV) virus (lineage 5 RFLP 2-5-2) in the same sample. Vaccine-like sequences were detected more frequently than the WT virus when the MLV virus was present at equal or higher concentrations than the WT virus. This result suggests that ORF5 Sanger sequencing may preferentially detect the dominant virus in samples containing more than one virus, potentially masking WT viral infections in vaccinated herds. Although our findings highlight a limitation in identifying co-circulation of strains, Sanger sequencing is still widely used as an accessible tool to characterize PRRSVs. Advanced sequencing techniques, such as NGS or CLAMP-based approaches, would complement Sanger sequencing results and allow improved detection of co-circulating variants by minimizing consensus sequence bias and selectively blocking vaccine-like sequences.