New insights into the testicular tropism of porcine reproductive and respiratory syndrome virus.

Porcine reproductive and respiratory syndrome virus (PRRSV) has a restricted host specificity, primarily infecting porcine macrophages. Notably, an exception to such macrophage-restricted tropism has been observed in sexually active boars, where the virus infects and induces apoptosis in the germinal epithelium, resulting in viral dissemination in the ejaculate. Whether this phenomenon occurs in prepubertal animals remains unclear. In this study, we isolated spermatogonia stem cells (SSCs) from neonatal pigs and cultured them in vitro. These SSC cultures formed morula-like colonies, exhibited alkaline phosphatase activity-a characteristic of stem cells-and expressed protein gene product 9.5, a marker of SSCs. Notably, the SSC cultures supported PRRSV replication with kinetics similar to that observed in porcine alveolar macrophages. To assess the testicular tropism of PRRSV in prepuberal animals, 28-day-old male pigs were infected with a virulent PRRSV strain. Testicular tissues were sequentially analyzed using a combination of in situ hybridization for PRRSV RNA and immunohistochemistry for specific cellular markers. Unlike in sexually active boars, PRRSV did not infect the spermatogonia cells within the seminiferous tubules of prepubertal pigs. Instead, the virus primarily infected macrophages and myoid cells located in the interstitium and peritubular areas. It appeared that the anatomical separation of spermatogonia from the basal membrane of the seminiferous tubules in prepubertal pigs prevents these cells from being infected by PRRSV. Overall, our findings offer valuable insights into the age-dependent testicular tropism of PRRSV.IMPORTANCEContaminated boar semen used in artificial insemination has significantly contributed to the global spread of porcine reproductive and respiratory syndrome virus (PRRSV), a virus that typically infects only cells within the monocyte and macrophage lineages. Our study reveals that spermatogonia stem cells (SSCs) from neonatal piglets are also susceptible to PRRSV, suggesting that non-macrophage cells can be infected by the virus. However, despite this susceptibility, PRRSV-infected cells were not found in the seminiferous tubules of prepubertal male pigs inoculated with a virulent PRRSV strain. This contrasts with sexually mature boars, where PRRSV-infected cells were prominently observed within the seminiferous tubules. The discrepancy is likely due to anatomical differences between the seminiferous tubules of sexually mature boars and prepubertal pigs. These findings provide new insights into PRRSV pathogenesis. Additionally, the ex vivo SSC culture provides a valuable model for identifying new viral receptors necessary for PRRSV infection and for investigating the virus's impact on spermatogenesis.