Effect of infectious bursal disease virus infection on the expression of immune checkpoint molecules in SPF chickens

Infectious bursal disease (IBD) is an acute, highly contagious, and immunosuppressive avian disease caused by infectious bursal disease virus (IBDV). Immune checkpoint molecules are vital for regulating immune equilibrium, preventing autoimmune responses, and controlling the scale and duration of immune reactions. These molecules are implicated in the immunosuppressive mechanisms initiated by viral infections. Currently, the correlation between IBDV infection and the expression of immune checkpoint molecules is not clearly defined. In this study, White Leghorn SPF chickens were inoculated with very virulent (vvIBDV), attenuated (attIBDV), or novel variant (nvIBDV) IBDV strains. Using real-time PCR, we quantified immune checkpoint genes and cytokines in the bursa of fabricius (BF), thymus, spleen, and peripheral blood mononuclear cells (PBMCs). The findings revealed a significant upregulation of immune checkpoint genes (PD-1, PD-L1, PD-L2, LAG3, CTLA4) in the BF, thymus, spleen, and PBMCs following infection with various virulent strains of IBDV compared to control tissues. Specifically, PD-L1 in BF surged to 7.32-fold (vvIBDV, 3 dpi), 7.44-fold (attIBDV, 3 dpi), and 6.47-fold (nvIBDV, 3 dpi) compared to controls. PD-L2 in spleen surged to 7.66-fold (attIBDV, 21 dpi), and 7.82-fold (nvIBDV, 21 dpi) compared to controls. Moreover, the expression levels of inhibitory and pro-inflammatory cytokines (TGF-β2, IL-10, IL-6, IL-1β, TNF-α) were notably elevated in infected tissues compared to control tissues, with IL-1β upregulated by 7.47-fold in BF at 3 dpi. TGF-β2 in spleen surged to 8.16-fold (vvIBDV, 21 dpi), 6.33-fold (attIBDV, 21 dpi), and 7.61-fold (nvIBDV, 21 dpi). These results suggest that IBDV infection in SPF chickens induces robust upregulation of immune checkpoint molecules (PD-1, PD-L1, PD-L2, LAG3, CTLA4) and cytokines (TGF-β2, IL-10, IL-6, IL-1β, TNF-α), correlating with viral virulence and immunosuppression. Our findings suggest that immune checkpoint dysregulation may contribute to IBDV pathogenesis, particularly in mediating sustained immunosuppression in poultry.