• Vaccine Name: F. tularensis vaccine LVS LPS
• Target Pathogen: Francisella tularensis
• Target Disease: Tularemia
• Vaccine Ontology ID: VO_0000308
• Type: Subunit vaccine
• Antigen: Francisella tularensis live vaccine strain (LVS) lipopolysaccharide (LPS) (Dreisbach et al., 2000). The chemical composition of the lipid A moiety of LVS LPS is quite different from that of enteric LPS. The LVS LPS lacks a functional lipid A. This apparently confers a survival advantage on the bacterium, since its LPS does not participate in induction of nitric oxide production that might limit its intracellular growth. Further, since other data indicate that LVS LPS is unable to block macrophage stimulation by functional LPS, the structure must be distinct enough to not permit recognition as an antagonist for traditional LPSs. The ability of LVS LPS to stimulate protection in C3H/HeJ mice, which are defective in the ability to recognize and respond to enteric LPS at least in part due to a point mutation in Toll-like receptor 4, suggests that LVS LPS is recognized through receptors other than Tlr4 (Dreisbach et al., 2000).
• Preparation: Francisella tularensis live vaccine strain (LVS) LPS is purified from whole F. tularensis LVS bacteria. A culture of F. tularensis LVS is grown in 4-L flasks at 37°C for 48 h in Trypticase soy broth with cysteine. The cultures are centrifuged for 15 min, washed 3 times in PBS, once in methanol, and once in acetone, and then lyophilized. The LPS is then extracted. After the crude LPS is treated with DNase, RNase, and proteinase K, the pellet is harvested by centrifugation 3 times for 12 h. LPS preparations are then reconstituted in endotoxin-free PBS and stored at 4°C (Dreisbach et al., 2000).
• Virulence: No traditional endotoxin has been associated with virulent F. tularensis. More recent reports indicated that purified LVS LPS was not endotoxic in d-galactosamine-sensitized mice and failed to activate Limulus amoebocyte lysate. LVS LPS also failed to stimulate human monocytes or peripheral blood lymphocytes to proliferate, produce TNF-α, or produce IL-1. Mouse peritoneal exudate macrophages treated with LVS LPS did not produce TNF-α or nitric oxide, and there was no increase in surface immunoglobulin expression by a mouse pre-B-cell line in response to LVS LPS. The only reported biological activity of LVS LPS is activation of complement (Dreisbach et al., 2000). LPS purified from F. tularensis LVS did not activate murine B cells for proliferation or polyclonal immunoglobulin secretion, nor did it activate murine splenocytes for secretion of interleukin-4 (IL-4), IL-6, IL-12, or IFN-gamma. However, in vivo experiments have suggested that LVS LPS contributes to the virulence of Francisella, in that LPS-defective Lpsd C3H/HeJ mice are reported to be more susceptible to LVS infection than Lpsn C3H/HeN (Dreisbach et al., 2000).

Mouse Response

• Host Strain: BALB/cByJ, C57BL/6J, BALB/c.scid
• Vaccination Protocol: Normal mouse serum (NMS) was obtained by bleeding normal BALB/cByJ mice from the lateral tail vein and pooling the resulting serum. Normal BALB/cByJ mice were then immunized with 10 µg of LPS intradermally (i.d.), and immune mouse serum (IMS) was obtained 3 or 30 days later. BALB/cByJ mice were given 0.5 ml of a 1:4 dilution of these sera i.p. 1 day before challenge with 10^3 LVS bacteria i.p. (Dreisbach et al., 2000).
• Persistence: T-cell-deficient BALB/c.nu/nu mice treated with LVS LPS survived lethal challenge for 2 to 3 weeks longer than control (PBS-treated) mice; these mice eventually succumbed to challenge after ~3-4 weeks (Dreisbach et al., 2000).
• Immune Response: LPS purified from F. tularensis LVS lacks the ability to nonspecifically activate murine B cells for proliferation or polyclonal immunoglobulin secretion. It is also unable to activate murine or human macrophages (Dreisbach et al., 2000). It does not stimulate murine splenocytes to secrete IL-12, IL-6, IL-4, or IFN-γ (Dreisbach et al., 2000). LVS LPS is unable to activate murine or human macrophages. LVS LPS does stimulate specific antibody production and in purified form is a weak immunogen that induces primarily an IgM response. When recognized by the murine immune system as a component of the bacterium, however, the resulting specific antibody response is vigorous and is characterized by the production of large amounts of IgG, particularly IgG2a. Despite the apparent absence of nonspecific immunostimulatory activity and minimal specific antibody production, however, treatment of mice with surprisingly small amounts of LVS LPS stimulates very strong B-cell-dependent protection against lethal LVS challenge within 2 to 3 days (Dreisbach et al., 2000).
• Side Effects: None were noted (Dreisbach et al., 2000).
• Challenge Protocol: BALB/cByJ mice were given various doses of LVS LPS i.d. on day 0 and challenged 3 days later with lethal doses of LVS, either 10^3 (1,000 LD50) or 10^4 (10,000 LD50) bacteria (Dreisbach et al., 2000).
• Efficacy: Very strong protection against lethal LVS infection was demonstrated. Mice given doses as low as 0.1 ng of LVS LPS i.d. survived lethal LVS challenge, depending on the strength of the challenge given. Mice given 100 ng of LVS LPS i.d. survived LVS challenge doses approaching 10^6 bacteria (1,000,000 LD50). Further, in three experiments, 15 of 15 mice challenged with 10^4 LVS bacteria i.p. 3, 10, or 35 days after treatment with 100 ng of LVS LPS i.d. survived; however, in the same experiments none of 15 mice treated with 100 ng of LVS LPS i.d. and challenged 3, 10, or 35 days with 20 LD50 of L. monocytogenes survived. Early protection also could not be demonstrated using other types of LPS, in that mice given 100 ng of E. coli O55 LPS, Salmonella serovar Typhimurium LPS, or E. coli K235 LPS i.d. on day 0 and challenged with 103 LVS bacteria i.p. on day 3 did not survive.