Highly efficient intranasal H5N1 vaccine based on a safe vesicular stomatitis virus (VSV) vector

Recombinant vesicular stomatitis virus (rVSV) is an increasingly popular vaccine vector, but it still faces challenges such as difficulties in construction, low in vitro amplification titers, safety concerns, and uncertain mucosal immune effects. Therefore, there is an urgent need to construct new VSV vectors for next-generation vaccines with high immunogenicity and excellent safety. In this study, we independently constructed a high-expression 6-9G recombinant replication-competent VSV vector. Based on the 6-9G vector, site-directed mutations were introduced into the M gene, including deletion of the 51st amino acid (Δ51), Δ51 + V221F, and Δ51 + S226R, to generate vectors designated as 6-9G-Δ51, 6-9G-Δ51 + V221F, and 6-9G-Δ51 + S226R. Through in vitro replication kinetics and in vivo immunogenicity studies, we identified 6-9G-Δ51 + S226R vector with a favorable safety profile and strong immunogenicity. The highly pathogenic avian influenza virus H5 subtype clade 2.3.4.4b poses a global pandemic threat, urgently necessitating new strategies for its prevention and control. Based on the 6-9G-Δ51 + S226R vector, we constructed the 6-9ΔG_Δ51+S226R-H5N1 influenza virus vectored vaccine by replacing the VSV-G gene with the HA and NA genes. This candidate demonstrated further improved safety while eliciting robust and durable humoral as well as mucosal immune responses after two doses, providing 100% protection in mice against lethal viral challenge. The successfully developed novel 6-9ΔG_Δ51+S226R-H5N1 vaccine candidate not only exhibits excellent safety and immunogenicity but also represents a powerful potential tool for the effective prevention and control of highly pathogenic avian influenza.