Isolation, identification, and pathogenicity comparison of three NADC30-like PRRSV variants with distinct mutational patterns

Porcine reproductive and respiratory syndrome virus (PRRSV) remains a major challenge to disease control in the swine industry. Since NADC30-like PRRSV was first detected in China in 2013, it has become a particular concern because of its widespread circulation, frequent recombination, and variable pathogenicity. Here, three PRRSV isolates, SCABTC-202503, SCABTC-202505, and SCABTC-202506, were isolated and characterized. Whole-genome sequencing and phylogenetic analyses placed all three isolates within the NADC30-like L1C sublineage and identified characteristic discontinuous amino acid deletions in Nsp2. Recombination analyses indicated a shared NADC30-like genomic backbone, but distinct putative recombination patterns. Comparative GP5 analysis showed that SCABTC-202503 and SCABTC-202505 each harbored a single-amino-acid deletion at position 37 and multiple additional substitutions, whereas SCABTC-202506 encoded a full-length GP5 protein. In piglet challenge experiments, the three isolates showed different patterns of disease-associated changes, with variation in clinical scores, viral loads, cytokine levels, and the extent of lung lesions among the infected groups. However, because the isolates differed at multiple genomic sites, the association between the GP5 deletion and the observed phenotypic variation remains correlative. Establishing the specific contribution of the GP5 deletion will require infectious clone-based mutagenesis in a controlled genetic background. Collectively, these findings highlight the genetic and phenotypic heterogeneity of the NADC30-like PRRSV isolates examined and underscore the importance of continued surveillance of emerging variants.IMPORTANCEPorcine reproductive and respiratory syndrome virus (PRRSV) remains a major pathogen affecting the swine industry because of its rapid evolution, frequent recombination, and variable pathogenicity. NADC30-like PRRSV has circulated widely in China and continues to diversify genetically. In this study, we obtained three recently circulating NADC30-like PRRSV isolates belonging to the L1C sublineage from clinical samples collected in China. We characterized their whole-genome sequences, phylogenetic relationships, putative recombination patterns, GP5 and Nsp2 sequence variations, and disease-associated changes in piglets. Under the conditions tested, the isolates exhibited genetic heterogeneity and varying degrees of clinical, virological, pathological, and inflammatory changes. These findings expand current knowledge of NADC30-like PRRSV diversity and underscore the importance of continued genomic surveillance for tracking emerging variants and informing future prevention and control strategies.