Augmented innate and humoral immune responses enhance early and durable rabies protection by a CD40L-overexpressing rabies vaccine

Rabies is the only acute zoonotic infectious disease with a mortality rate approaching 100%. Large-scale immunization of animals represents the most effective strategy for preventing human rabies. Consequently, there is a pressing need for a vaccine that is economical, safe, and efficient for controlling rabies transmitted by animals. A recombinant rabies virus (rLBNSE-CD40L) overexpressing murine CD40L was constructed using the RABV reverse genetics system. The possible adverse effects of rLBNSE-CD40L were evaluated using growth curves, body weight changes, survival rates, and inflammatory cell counts. The immunogenicity of rLBNSE-CD40L was assessed by measuring virus-neutralizing antibodies (VNA) levels, antiviral protection rates, and antibody isotype levels in mice. Flow cytometry, RNA sequencing, and lymph node immunofluorescence were employed to investigate the innate and humoral immune responses induced by rLBNSE-CD40L. Notably, rLBNSE-CD40L was found to rapidly generate significantly higher VNA levels as early as three days post-immunization, which relies on the activation of dendritic cells (DCs) and the engagement of innate immune-related pathways and genes. This vaccine candidate enhances innate immunity and elicits persistent primary and secondary antibody responses by promoting T helper 1 (Th1)-biased and T follicular helper (Tfh)-dependent germinal center (GC) humoral immunity. In conclusion, our study suggests that rLBNSE-CD40L has the potential to be developed into a non-pathogenic and effective animal rabies vaccine.