Abstract
ABSTRACT There is a need for development of an effective vaccine against Francisella tularensis, as this potential bioweapon has a high mortality rate and low infectious dose when delivered via the aerosol route. Moreover, this Tier 1 agent has a history of weaponization. We engineered targeted mutations in the Type A strain F. tularensis subspecies tularensis Schu S4 in aro genes encoding critical enzymes in aromatic amino acid biosynthesis. F. tularensis Schu S4ΔaroC, Schu S4ΔaroD, and Schu S4ΔaroCΔaroD mutant strains were attenuated for intracellular growth in vitro and for virulence in vivo and, conferred protection against pulmonary wild-type (WT) F. tularensis Schu S4 challenge in the C57BL/6 mouse model. F. tularensis Schu S4ΔaroD was identified as the most promising vaccine candidate, demonstrating protection against high-dose intranasal challenge; it protected against 1,000 CFU Schu S4, the highest level of protection tested to date. It also provided complete protection against challenge with 92 CFU of a F. tularensis subspecies holarctica strain (Type B). Mice responded to vaccination with Schu S4ΔaroD with systemic IgM and IgG2c, as well as the production of a functional T cell response as measured in the splenocyte-macrophage co-culture assay. This vaccine was further characterized for dissemination, histopathology, and cytokine/chemokine gene induction at defined time points following intranasal vaccination which confirmed its attenuation compared to WT Schu S4. Cytokine, chemokine, and antibody induction patterns compared to wild-type Schu S4 distinguish protective vs. pathogenic responses to F. tularensis and elucidate correlates of protection associated with vaccination against this agent.
Highlights
Francisella tularensis is a Gram-negative coccobacillus and the etiological agent of the human disease tularemia which manifests clinically in multiple forms depending on the route of infection
This study identifies a promising, live attenuated vaccine candidate that is protective against high-dose respiratory challenge in mice and which may be useful in elucidating vaccineassociated correlates of protection against tularemia in the mouse model
Unmarked deletions in the FTT0876 c and FTT0471 genes were created in F. tularensis strain Schu S4 using a two-step allelic exchange strategy as previously described [9]
Summary
Francisella tularensis is a Gram-negative coccobacillus and the etiological agent of the human disease tularemia which manifests clinically in multiple forms depending on the route of infection. F. tularensis subspecies tularensis (Type A) causes the most severe form of disease, and subspecies holarctica (Type B) causes disease with reduced severity [6]. Previous attempts to develop a tularemia vaccine included the Live Vaccine Strain (LVS), an attenuated derivative of F. tularensis subspecies holarctica (Type B strain), which was tested extensively in humans [1,2]. LVS is not currently licensed for human use, it serves as proof of principle that a live attenuated vaccine can provide at least partial protection against subsequent challenge with the highly virulent wild-type (WT) F. tularensis subspecies tularensis (Type A). We proposed that generating rationally designed live attenuated vaccines in the subspecies tularensis background
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