Grey variants of the live vaccine strain of Francisella tularensis lack lipopolysaccharide O-antigen, show reduced ability to survive in macrophages and do not induce protective immunity in mice

Vaccine ◽  
2006 ◽  
Vol 24 (7) ◽  
pp. 989-996 ◽  
Author(s):  
Gill Hartley ◽  
Rosa Taylor ◽  
Jo Prior ◽  
Sarah Newstead ◽  
Paul G. Hitchen ◽  
...  
Keyword(s):  
Francisella Tularensis ◽  
Vaccine Strain ◽  
Protective Immunity ◽  
Live Vaccine ◽  
Live Vaccine Strain ◽  
O Antigen

scholarly journals A Francisella tularensis Live Vaccine Strain (LVS) Mutant with a Deletion in capB, Encoding a Putative Capsular Biosynthesis Protein, Is Significantly More Attenuated than LVS yet Induces Potent Protective Immunity in Mice against F. tularensis Challenge

2010 ◽  
Vol 78 (10) ◽  
pp. 4341-4355 ◽  
Author(s):  
Qingmei Jia ◽  
Bai-Yu Lee ◽  
Richard Bowen ◽  
Barbara Jane Dillon ◽  
Susan M. Som ◽  
...  
Keyword(s):  
Francisella Tularensis ◽  
Vaccine Strain ◽  
Protective Immunity ◽  
Lethal Dose ◽  
Live Vaccine ◽  
Human Macrophage ◽  
Live Vaccine Strain ◽  
Virulent Type ◽  
Marker Free ◽  
Cellular Immune

ABSTRACT Francisella tularensis, the causative agent of tularemia, is in the top category (category A) of potential agents of bioterrorism. The F. tularensis live vaccine strain (LVS) is the only vaccine currently available to protect against tularemia; however, this unlicensed vaccine is relatively toxic and provides incomplete protection against aerosolized F. tularensis, the most dangerous mode of transmission. Hence, a safer and more potent vaccine is needed. As a first step toward addressing this need, we have constructed and characterized an attenuated version of LVS, LVS ΔcapB, both as a safer vaccine and as a vector for the expression of recombinant F. tularensis proteins. LVS ΔcapB, with a targeted deletion in a putative capsule synthesis gene (capB), is antibiotic resistance marker free. LVS ΔcapB retains the immunoprotective O antigen, is serum resistant, and is outgrown by parental LVS in human macrophage-like THP-1 cells in a competition assay. LVS ΔcapB is significantly attenuated in mice; the 50% lethal dose (LD50) intranasally (i.n.) is >10,000-fold that of LVS. Providing CapB in trans to LVS ΔcapB partially restores its virulence in mice. Mice immunized with LVS ΔcapB i.n. or intradermally (i.d.) developed humoral and cellular immune responses comparable to those of mice immunized with LVS, and when challenged 4 or 8 weeks later with a lethal dose of LVS i.n., they were 100% protected from illness and death and had significantly lower levels (3 to 5 logs) of LVS in the lung, liver, and spleen than sham-immunized mice. Most importantly, mice immunized with LVS ΔcapB i.n. or i.d. and then challenged 6 weeks later by aerosol with 10× the LD50 of the highly virulent type A F. tularensis strain SchuS4 were significantly protected (100% survival after i.n. immunization). These results show that LVS ΔcapB is significantly safer than LVS and yet provides potent protective immunity against virulent F. tularensis SchuS4 challenge.

scholarly journals A Defined O-Antigen Polysaccharide Mutant of Francisella tularensis Live Vaccine Strain Has Attenuated Virulence while Retaining Its Protective Capacity

2007 ◽  
Vol 75 (5) ◽  
pp. 2591-2602 ◽  
Author(s):  
Shite Sebastian ◽  
Simon T. Dillon ◽  
Jillian G. Lynch ◽  
LeeAnn T. Blalock ◽  
Emmy Balon ◽  
...  
Keyword(s):  
Francisella Tularensis ◽  
Vaccine Strain ◽  
Lethal Dose ◽  
Live Vaccine ◽  
Effective Vaccine ◽  
Live Vaccine Strain ◽  
Virulence Determinants ◽  
Protective Capacity ◽  
O Antigen

ABSTRACT Francisella tularensis, the causative agent of tularemia, has been designated a CDC category A select agent because of its low infective dose (<10 CFU), its ready transmission by aerosol, and its ability to produce severe morbidity and high mortality. The identification and characterization of this organism's virulence determinants will facilitate the development of a safe and effective vaccine. We report that inactivation of the wbtA-encoded dehydratase of the O-antigen polysaccharide (O-PS) locus of the still-unlicensed live vaccine strain of F. tularensis (LVS) results in a mutant (the LVS wbtA mutant) with remarkably attenuated virulence. Western blot analysis and immune electron microscopy studies associate this loss of virulence with a complete lack of surface O-PS expression. A likely mechanism for attenuation is shown to be the transformation from serum resistance in the wild-type strain to serum sensitivity in the mutant. Despite this significant attenuation in virulence, the LVS wbtA mutant remains immunogenic and confers protective immunity on mice against challenge with an otherwise lethal dose of either F. tularensis LVS or a fully virulent clinical isolate of F. tularensis type B. Recognition and characterization of the pivotal role of O-PS in the virulence of this intracellular bacterial pathogen may have broad implications for the creation of a safe and efficacious vaccine.

scholarly journals Purified Lipopolysaccharide from Francisella tularensis Live Vaccine Strain (LVS) Induces Protective Immunity against LVS Infection That Requires B Cells and Gamma Interferon

2000 ◽  
Vol 68 (4) ◽  
pp. 1988-1996 ◽  
Author(s):  
Valley C. Dreisbach ◽  
Siobhan Cowley ◽  
Karen L. Elkins
Keyword(s):  
B Cells ◽  
Francisella Tularensis ◽  
Vaccine Strain ◽  
Protective Immunity ◽  
Gamma Interferon ◽  
Live Vaccine ◽  
Immunoglobulin M ◽  
Live Vaccine Strain ◽  
Murine Splenocytes ◽  
Ifn Γ

ABSTRACT Previous results have demonstrated that nonspecific protective immunity against lethal Francisella tularensis live vaccine strain (LVS) or Listeria monocytogenes infection can be stimulated either by sublethal infection with bacteria or by treatment with bacterial DNA given 3 days before lethal challenge. Here we characterize the ability of purified lipopolysaccharide (LPS) fromF. tularensis LVS to stimulate similar early protective immunity. Treatment of mice with surprisingly small amounts of LVS LPS resulted in very strong and long-lived protection against lethal LVS challenge within 2 to 3 days. Despite this strong protective response, 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 gamma interferon (IFN-γ). Immunization of mice with purified LVS LPS induced a weak specific anti-LPS immunoglobulin M (IgM) response and very little IgG; however, infection of mice with LVS bacteria resulted in vigorous IgM and IgG, particularly IgG2a, anti-LPS antibody responses. Studies using various immunodeficient mouse strains, including LPS-hyporesponsive C3H/HeJ mice, μMT− (B-cell-deficient) knockout mice, and IFN-γ-deficient mice, demonstrated that the mechanism of protection does not involve recognition through the Lpsn gene product; nonetheless, protection was dependent on B cells as well as IFN-γ.

scholarly journals Oral Live Vaccine Strain-Induced Protective Immunity against Pulmonary Francisella tularensis Challenge Is Mediated by CD4+ T Cells and Antibodies, Including Immunoglobulin A

2009 ◽  
Vol 16 (4) ◽  
pp. 444-452 ◽  
Author(s):  
Heather J. Ray ◽  
Yu Cong ◽  
Ashlesh K. Murthy ◽  
Dale M. Selby ◽  
Karl E. Klose ◽  
...  
Keyword(s):  
T Cells ◽  
Francisella Tularensis ◽  
Vaccine Strain ◽  
Protective Immunity ◽  
Immunoglobulin A ◽  
Live Vaccine ◽  
Live Vaccine Strain ◽  
Oral Vaccination ◽  
Enhanced Production ◽  
Schu S4

ABSTRACT Francisella tularensis is an intracellular gram-negative bacterium and the etiological agent of pulmonary tularemia. Given the high degrees of infectivity in the host and of dissemination of bacteria following respiratory infection, immunization strategies that target mucosal surfaces are critical for the development of effective vaccines against this organism. In this study, we have characterized the efficacy of protective immunity against pneumonic tularemia following oral vaccination with F. tularensis LVS (live vaccine strain). Mice vaccinated orally with LVS displayed colocalization of LVS with intestinal M cells, with subsequent enhanced production of splenic antigen-specific gamma interferon and of systemic and mucosal antibodies, including immunoglobulin A (IgA). LVS-vaccinated BALB/c mice were highly protected against intranasal (i.n.) SCHU S4 challenge and exhibited significantly less bacterial replication in the lungs, liver, and spleen than mock-immunized animals. Depletion of CD4+ T cells significantly abrogated the protective immunity, and mice deficient in B cells or IgA displayed partial protection against SCHU S4 challenge. These results suggest that oral vaccination with LVS induces protective immunity against i.n. challenge with F. tularensis SCHU S4 by a process mediated cooperatively by CD4+ T cells and antibodies, including IgA.

The Bla2 β-lactamase from the live-vaccine strain of Francisella tularensis encodes a functional protein that is only active against penicillin-class β-lactam antibiotics

2006 ◽  
Vol 186 (3) ◽  
pp. 219-228 ◽  
Author(s):  
Xiaowen R. Bina ◽  
Chunmei Wang ◽  
Mark A. Miller ◽  
James E. Bina
Keyword(s):  
Francisella Tularensis ◽  
Vaccine Strain ◽  
Live Vaccine ◽  
Live Vaccine Strain ◽  
Functional Protein

scholarly journals TolC-Dependent Modulation of Host Cell Death by the Francisella tularensis Live Vaccine Strain

2014 ◽  
Vol 82 (5) ◽  
pp. 2068-2078 ◽  
Author(s):  
Christopher R. Doyle ◽  
Ji-An Pan ◽  
Patricio Mena ◽  
Wei-Xing Zong ◽  
David G. Thanassi
Keyword(s):  
Cell Death ◽  
Immune Responses ◽  
Host Cell ◽  
Francisella Tularensis ◽  
Vaccine Strain ◽  
Live Vaccine ◽  
Live Vaccine Strain ◽  
Type I ◽  
Content Type ◽  
Host Cell Death

ABSTRACTFrancisella tularensisis a facultative intracellular, Gram-negative pathogen and the causative agent of tularemia. We previously identified TolC as a virulence factor of theF. tularensislive vaccine strain (LVS) and demonstrated that a ΔtolCmutant exhibits increased cytotoxicity toward host cells and elicits increased proinflammatory responses compared to those of the wild-type (WT) strain. TolC is the outer membrane channel component used by the type I secretion pathway to export toxins and other bacterial virulence factors. Here, we show that the LVS delays activation of the intrinsic apoptotic pathway in a TolC-dependent manner, both during infection of primary macrophages and during organ colonization in mice. The TolC-dependent delay in host cell death is required forF. tularensisto preserve its intracellular replicative niche. We demonstrate that TolC-mediated inhibition of apoptosis is an active process and not due to defects in the structural integrity of the ΔtolCmutant. These findings support a model wherein the immunomodulatory capacity ofF. tularensisrelies, at least in part, on TolC-secreted effectors. Finally, mice vaccinated with the ΔtolCLVS are protected from lethal challenge and clear challenge doses faster than WT-vaccinated mice, demonstrating that the altered host responses to primary infection with the ΔtolCmutant led to altered adaptive immune responses. Taken together, our data demonstrate that TolC is required for temporal modulation of host cell death during infection byF. tularensisand highlight how shifts in the magnitude and timing of host innate immune responses may lead to dramatic changes in the outcome of infection.

scholarly journals Intranasal Interleukin-12 Treatment for Protection against Respiratory Infection with the Francisella tularensis Live Vaccine Strain

2005 ◽  
Vol 73 (4) ◽  
pp. 2306-2311 ◽  
Author(s):  
Nathalie S. Duckett ◽  
Sofia Olmos ◽  
Douglas M. Durrant ◽  
Dennis W. Metzger
Keyword(s):  
Respiratory Infection ◽  
Francisella Tularensis ◽  
Vaccine Strain ◽  
Live Vaccine ◽  
Interleukin 12 ◽  
Live Vaccine Strain ◽  
Wild Type ◽  
Intranasal Infection ◽  
Ifn Γ

ABSTRACT Francisella tularensis is a gram-negative intracellular bacterium that can induce lethal respiratory infection in humans and rodents. However, little is known about the role of innate or adaptive immunity in protection from respiratory tularemia. In the present study, the role of interleukin-12 (IL-12) in inducing protective immunity in the lungs against intranasal infection of mice with the live vaccine strain (LVS) of F. tularensis was investigated. It was found that gamma interferon (IFN-γ) and IL-12 were strictly required for protection, since mice deficient in IFN-γ, IL-12 p35, or IL-12 p40 all succumbed to LVS doses that were sublethal for wild-type mice. Furthermore, exogenous IL-12 treatment 24 h before intranasal infection with a lethal dose of LVS (10,000 CFU) significantly decreased bacterial loads in the lungs, livers, and spleens of wild-type BALB/c and C57BL/6 mice and allowed the animals to survive infection; such protection was not observed in IFN-γ-deficient mice. The resistance induced by IL-12 to LVS infection was still observed in NK cell-deficient beige mice but not in CD8−/− mice. These results demonstrate that exogenous IL-12 delivered intranasally can prevent respiratory tularemia through a mechanism that is at least partially dependent upon the expression of IFN-γ and CD8 T cells.

scholarly journals O-Antigen-Deficient Francisella tularensis Live Vaccine Strain Mutants Are Ingested via an Aberrant Form of Looping Phagocytosis and Show Altered Kinetics of Intracellular Trafficking in Human Macrophages

2011 ◽  
Vol 80 (3) ◽  
pp. 952-967 ◽  
Author(s):  
Daniel L. Clemens ◽  
Bai-Yu Lee ◽  
Marcus A. Horwitz
Keyword(s):  
Vaccine Strain ◽  
Intracellular Trafficking ◽  
Live Vaccine ◽  
Live Vaccine Strain ◽  
Complement Components ◽  
Content Type ◽  
Human Macrophages ◽  
O Antigen ◽  
Kinetics Of ◽  
Terminal Complement

We examined the uptake and intracellular trafficking ofF. tularensisLive Vaccine Strain (LVS) and LVS with disruptions ofwbtDEFandwbtIgenes essential for synthesis of the O antigen of lipopolysaccharide. Unlike parental bacteria, O-antigen-deficient LVS is efficiently killed by serum with intact complement but not by serum lacking terminal complement components. Opsonization of O-antigen-deficient LVS in serum lacking terminal complement components allows efficient uptake of these live bacteria by macrophages. In the presence of complement, whereas parentalF. tularensisLVS is internalized within spacious pseudopod loops, mutant LVS is internalized within tightly juxtaposed multiple onion-like layers of pseudopodia. Without complement, both parental and mutant LVSs are internalized within spacious pseudopod loops. Thus, molecules other than O antigen are important in triggering dramatic pseudopod extensions and uptake by spacious pseudopod loops. Following uptake, both parental and mutant LVSs enter compartments that show limited staining for the lysosomal membrane glycoprotein CD63 and little fusion with secondary lysosomes. Subsequently, both parental and mutant LVSs lose their CD63 staining. Whereas the majority of parental LVS escapes into the cytosol by 6 h after uptake, mutant LVS shows a marked lag but does escape by 1 day after uptake. Despite the altered kinetics of phagosome escape, both mutant and parental strains grow to high levels within human macrophages. Thus, the O antigen plays a role in the morphology of uptake in the presence of complement and the kinetics of intracellular growth but is not essential for escape, survival, altered membrane trafficking, or intramacrophage growth.

iTRAQ quantitative analysis of Francisella tularensis ssp. holarctica live vaccine strain and Francisella tularensis ssp. tularensis SCHU S4 response to different temperatures and stationary phases of growth

PROTEOMICS ◽  
2009 ◽  
Vol 9 (10) ◽  
pp. 2875-2882 ◽  
Author(s):  
Juraj Lenco ◽  
Marek Link ◽  
Vojtech Tambor ◽  
Jitka Zaková ◽  
Lukas Cerveny ◽  
...  
Keyword(s):  
Quantitative Analysis ◽  
Francisella Tularensis ◽  
Vaccine Strain ◽  
Stationary Phases ◽  
Live Vaccine ◽  
Live Vaccine Strain ◽  
Different Temperatures ◽  
Schu S4
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