Japanese encephalitis virus NS1' subunit vaccine confers protection against viral challenge and mosquito-mediated transmission.

Japanese encephalitis virus (JEV) is a mosquito-borne pathogen that threatens human health and the livestock industry. Current JEV vaccines, including inactivated and live attenuated types, still present several limitations, underscoring the urgent need for new, safe, and broad-spectrum JEV vaccines. Antibodies targeting the orthoflavivirus NS1 protein, which is highly conserved among orthoflaviviruses, have been demonstrated to provide protection against viral infection, with NS1' being known as an extended form of NS1 in the Japanese encephalitis serogroup. In this study, we successfully constructed and purified recombinant maltose-binding protein (MBP)-fused JEV NS1 and NS1' proteins. Systematic evaluation of the immunoreactive characteristics of these two subunit vaccines in mouse and piglet models demonstrated that both vaccines effectively induced high titers of specific antibodies against NS1 or NS1'. Moreover, antibodies elicited by MBP-NS1' immunization showed a stronger effect on inhibiting JEV infection in vitro. Notably, immunization with the MBP-NS1' formulation exhibited superior immunoprotective efficacy against JEV challenge compared to MBP-NS1. More importantly, mosquito artificial feeding experiments confirmed that immune sera from piglets, acquired by mosquitoes during blood feeding, potently inhibited JEV infection in mosquitoes, with NS1'-elicited antibodies exhibiting a stronger inhibitory effect. These findings suggest that JEV NS1 and NS1' can serve as candidate antigens for JEV vaccine design, with NS1' exhibiting greater potential than NS1.