Combinatorial Viral Vector-Based and Live Attenuated Vaccines without an Adjuvant to Generate Broader Immune Responses to Effectively Combat Pneumonic Plague

Mice immunized with a combination of an adenovirus vector (Ad5-YFV) and live-attenuated (LMA)-based vaccines were evaluated for protective efficacy against pneumonic plague. While the Ad5-YFV vaccine harbors a fusion cassette of three genes encoding YscF, F1, and LcrV, LMA represents a mutant of parental Yersinia pestis CO92 deleted for genes encoding Lpp, MsbB, and Ail. Ad5-YFV and LMA were either administered simultaneously (1-dose regimen) or 21 days apart in various orders and route of administration combinations (2-dose regimen). The 2-dose regimen induced robust immune responses to provide full protection to animals against parental CO92 and its isogenic F1 deletion mutant (CAF ) challenges during both short- and long-term studies. Mice intranasally (i.n.) immunized with Ad5-YFV first followed by LMA (i.n. or intramuscularly [i.m.]) had higher T- and B-cell proliferative responses and LcrV antibody titers than those in mice vaccinated with LMA (i.n. or i.m.) first ahead of Ad5-YFV (i.n.) during the long-term study. Specifically, the needle- and adjuvant-free vaccine combination (i.n.) is ideal for use in plague regions of endemicity. Conversely, with a 1-dose regimen, mice vaccinated with Ad5-YFV i.n. and LMA by the i.m. route provided complete protection to animals against CO92 and its CAF mutant challenges and elicited Th1/Th2, as well as Th17 responses, making it suitable for emergency vaccination during a plague outbreak or bioterrorist attack. This is a first study in which a viral vector-based and live-attenuated vaccines were effectively used in combination, representing adjuvant- and/or needle-free immunization, with each vaccine triggering a distinct cellular immune response. Yersinia pestis, the causative agent of plague, is a Tier-1 select agent and a reemerging human pathogen. A 2017 outbreak in Madagascar with >75% of cases being pneumonic and 8.6% causalities emphasized the importance of the disease. The World Health Organization has indicated an urgent need to develop new-generation subunit and live-attenuated plague vaccines. We have developed a subunit vaccine, including three components (YscF, F1, and LcrV) using an adenovirus platform (Ad5-YFV). In addition, we have deleted virulence genes of Y. pestis (e.g., , , and ) to develop a live-attenuated vaccine (LMA). Both of these vaccines generated robust humoral and cellular immunity and were highly efficacious in several animal models. We hypothesized the use of a heterologous prime-