Infection with Francisella tularensis LVSclpBLeads to an Altered yet Protective Immune Response

Abstract

Bacterial attenuation is typically thought of as reduced bacterial growth in the presence of constant immune pressure. Infection with Francisella tularensis elicits innate and adaptive immune responses. Several in vivo screens have identified F. tularensis genes necessary for virulence. Many of these mutations render F. tularensis defective for intracellular growth. However, some mutations have no impact on intracellular growth, leading us to hypothesize that these F. tularensis mutants are attenuated because they induce an altered host immune response. We were particularly interested in the F. tularensis LVS (live vaccine strain) clpB (FTL_0094) mutant because this strain was attenuated in pneumonic tularemia yet induced a protective immune response. The attenuation of LVS clpB was not due to an intracellular growth defect, as LVS clpB grew similarly to LVS in primary bone marrow-derived macrophages and a variety of cell lines. We therefore determined whether LVS clpB induced an altered immune response compared to that induced by LVS in vivo. We found that LVS clpB induced proinflammatory cytokine production in the lung early after infection, a process not observed during LVS infection. LVS clpB provoked a robust adaptive immune response similar in magnitude to that provoked by LVS but with increased gamma interferon (IFN-␥) and interleukin-17A (IL-17A) production, as measured by mean fluorescence intensity. Altogether, our results indicate that LVS clpB is attenuated due to altered host immunity and not an intrinsic growth defect. These results also indicate that disruption of a nonessential gene(s) that is involved in bacterial immune evasion, like F. tularensis clpB, can serve as a model for the rational design of attenuated vaccines.

Novel Catanionic Surfactant Vesicle Vaccines Protect against Francisella tularensis LVS and Confer Significant Partial Protection against F. tularensis Schu S4 Strain

“…In IL-6 Ϫ/Ϫ mice, susceptibility to F. tularensis LVS infection is enhanced (68). Moreover, live attenuated F. tularensis strains, such as the ⌬clpB strain, elicit early inflammatory responses and induce immunity to F. tularensis LVS (35)(36)(37). Immunization with attenuated Schu S4 mutants capable of protecting mice against subsequent virulent Schu S4 challenge was shown to correlate with higher blood serum levels of TNF-␣, IFN-␥, and MCP-1 within the first week postinfection than immunization with nonprotective mutants (60).…”

“…Although immunization with certain attenuated mutant strains of F. tularensis protects against subsequent challenge (35)(36)(37), they raise safety concerns due to the potential for reversion to full virulence. Therefore, subunit vaccines are generally considered to be safer than live attenuated vaccines (38).…”

Identification of Early Interactions between Francisella and the Host

“…Following humane euthanasia, lungs were aseptically removed after perfusion with PBS and digested into a single-cell suspension as previously described (15). For intranasally inoculated mice, 50 g/ml of gentamicin (Sigma) was added to the digestion mix to kill extracellular bacteria.…”

Janus Kinase 3 Activity Is Necessary for Phosphorylation of Cytosolic Phospholipase A2 and Prostaglandin E2 Synthesis by Macrophages Infected with Francisella tularensis Live Vaccine Strain

“…The ability of F. tularensis to invade and replicate within macrophages is paramount to its survival within the host, as F. tularensis mutants that fail to replicate in macrophages are avirulent (3). However, modulation of the host immune response by F. tularensis is also important to bacterial survival within the host (4). Numerous mechanisms of F. tularensis-mediated immune evasion have been identified (reviewed in reference 5).…”