Abstract / Summary
Abstract Parenteral vaccines against respiratory viruses poorly block infection and transmission due to suboptimal induction of mucosal immunity. In contrast, hybrid immunity and heterologous-route immunization regimens generate protective mucosal and systemic immunity. Herein, we examine how chronological order in heterologous prime/boost immunization regimens impacts immune responses using combinations of intramuscular mRNA and intranasal adjuvanted subunit vaccines against SARS-CoV-2. Enhanced long-lived mucosal and systemic immunity depended strongly on vaccine delivery sequence, with heterologous regimens administered in alternate order of routes yielding markedly distinct immune profiles with different Th-biases and divergent localizations. Immunity imparted through heterologous immunization was more durable and responsive to delayed boosting than parenteral-only regimens. Furthermore, intramuscular mRNA immunization uniquely shaped gene expression locally and systemically through IFN-I induction, allowing for enhanced responses to subsequent mucosal immunizations. These findings demonstrate that the context of initial antigen exposure—including the route of exposure and associated inflammatory signals—profoundly shapes responses to subsequent encounters. Thus, vaccine responses in antigen-experienced hosts differ fundamentally from those induced on a naive baseline, reflecting the influence of pre-existing innate and adaptive immune memory.