Characterization of the pathogenicity of newly emerged NADC30-Like PRRSV strains causing severe brain infections via twice inter-lineage recombination.

Porcine reproductive and respiratory syndrome virus (PRRSV) poses a persistent and devastating threat to the global swine industry. In China, NADC30-like PRRSV strains (Lineage 1.8) have undergone extensive and continuous recombination with field isolates and vaccine-derived strains, resulting in variants with altered pathogenicity and tissue tropism that severely complicate disease control and prevention efforts. In the present study, two novel NADC30-like PRRSV strains (PRRSV01-2024 and PRRSV02-2024) were isolated from piglets exhibiting respiratory and neurological symptoms in two distinct pig farms in Fujian Province, China. Genetic and recombination analyses revealed that both isolates are Lineage 1.8 strains derived from at least two rounds of inter-lineage recombination, with GDsf1707 (a Lineages 1.8 and 8.7 recombinant strain) serving as the major parental virus, while VR2332-like vaccine strain RespPRRS MLV as the minor parental virus. Animal studies indicated that both strains exhibited more severe clinical symptoms in piglets, including 20%-40% mortality, persistent fever, significant weight loss, higher viremia, and severe interstitial pneumonia compared with the moderately virulent NADC30-like strain FJZ03. More importantly, the two strains possessed a novel neurotropic phenotype, inducing neurological symptoms and severe neurological lesions, with PRRSV antigens and high viral loads (up to 104.4-104.6 copies/mg) detected in brain tissues. Furthermore, qPCR and four-color multiplex immunofluorescence staining revealed that both strains downregulated the expression of tight junction proteins (CLDN5, OCLN, and TJP1), thereby enabling them to cross the blood-brain barrier (BBB) into the central nervous system and infect glial cells. Taken together, these findings suggest that multi-round recombination increases the tissue tropism and virulence of PRRSV, underlining the importance of intensified molecular surveillance and targeted control strategies.