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 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 Diverse Myeloid and Lymphoid Cell Subpopulations Produce Gamma Interferon during Early Innate Immune Responses to Francisella tularensis Live Vaccine Strain

2008 ◽  
Vol 76 (9) ◽  
pp. 4311-4321 ◽  
Author(s):  
Roberto De Pascalis ◽  
Betsy C. Taylor ◽  
Karen L. Elkins
Keyword(s):  
T Cells ◽  
Immune Responses ◽  
Nk Cells ◽  
Francisella Tularensis ◽  
Vaccine Strain ◽  
Live Vaccine ◽  
Innate Immune ◽  
Live Vaccine Strain ◽  
Ifn Γ ◽  
Cell Subpopulations

ABSTRACT Francisella tularensis, a small gram-negative intracellular bacterium responsible for causing tularemia, is highly pathogenic and classified as a category A agent of bioterrorism. As for other intracellular pathogens, successful protective immune responses to Francisella tularensis require rapid and efficient induction of gamma interferon (IFN-γ) production. Studies using intracellular bacteria such as Listeria monocytogenes as well as Francisella suggest that natural killer (NK) and T cells are important sources of IFN-γ. However, comprehensive characterization of specific sources of IFN-γ produced during Francisella infection in vivo remains incomplete, and depletion of NK cells before infection of mice with the F. tularensis live vaccine strain (LVS) has little impact on the course or outcome of infection. In this study, we determined the cell subpopulations that respond quickly to intradermal F. tularensis LVS infection of mice by producing IFN-γ within hours to a few days. Splenic and liver lymphocytes were obtained from LVS-infected mice and analyzed for IFN-γ mRNA by reverse transcription-PCR, for intracellular cytokine expression by multiparameter flow cytometry, and for ex vivo production of IFN-γ protein by enzyme-linked immunosorbent assay. Cells producing IFN-γ were readily detectable by day 3 after infection, and numbers progressively increased through days 5 to 7. Importantly, the cell types responsible for IFN-γ production were much more varied than expected: these included not only NK cells and T cells, which might be predicted, but also other cells, including dendritic cells (DCs), “NK DCs,” NK T cells, and neutrophils. Most importantly, since RAG-1 knockout mice appeared to exhibit a frequency of IFN-γ-producing cells comparable to that of intact wild-type mice, early IFN-γ production by innate immune cells does not depend on the presence of T or B cells.

scholarly journals T-bet Regulates Immunity to Francisella tularensis Live Vaccine Strain Infection, Particularly in Lungs

2014 ◽  
Vol 82 (4) ◽  
pp. 1477-1490 ◽  
Author(s):  
Amanda A. Melillo ◽  
Oded Foreman ◽  
Catharine M. Bosio ◽  
Karen L. Elkins
Keyword(s):  
Francisella Tularensis ◽  
Vaccine Strain ◽  
Live Vaccine ◽  
Interleukin 17 ◽  
Live Vaccine Strain ◽  
Respiratory Protection ◽  
Necrosis Factor Alpha ◽  
Secondary Challenge ◽  
Ifn Γ

ABSTRACTUpregulation of the transcription factor T-bet is correlated with the strength of protection against secondary challenge with the live vaccine strain (LVS) ofFrancisella tularensis. Thus, to determine if this mediator had direct consequences in immunity to LVS, we examined its role in infection. Despite substantialin vivogamma interferon (IFN-γ) levels, T-bet-knockout (KO) mice infected intradermally (i.d.) or intranasally (i.n.) with LVS succumbed to infection with doses 2 log units less than those required for their wild-type (WT) counterparts, and exhibited significantly increased bacterial burdens in the lung and spleen. Lungs of LVS-infected T-bet-KO mice contained fewer lymphocytes and more neutrophils and interleukin-17 than WT mice. LVS-vaccinated T-bet-KO mice survived lethal LVS intraperitoneal secondary challenge but not high doses of LVS i.n. challenge, independently of the route of vaccination. Immune T lymphocytes from the spleens of i.d. LVS-vaccinated WT or KO mice controlled intracellular bacterial replication in anin vitrococulture system, but cultures with T-bet-KO splenocyte supernatants contained less IFN-γ and increased amounts of tumor necrosis factor alpha. In contrast, immune T-bet-KO lung lymphocytes were greatly impaired in controlling intramacrophage growth of LVS; this functional defect is the likely mechanism underpinning the lack of respiratory protection. Taken together, T-bet is important in host resistance to primary LVS infection and i.n. secondary challenge. Thus, T-bet represents a true, useful correlate for immunity to LVS.

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 Susceptibility to Secondary Francisella tularensis Live Vaccine Strain Infection in B-Cell-Deficient Mice Is Associated with Neutrophilia but Not with Defects in Specific T-Cell-Mediated Immunity

2001 ◽  
Vol 69 (1) ◽  
pp. 194-203 ◽  
Author(s):  
Catharine M. Bosio ◽  
Karen L. Elkins
Keyword(s):  
T Cells ◽  
B Cells ◽  
B Cell ◽  
Francisella Tularensis ◽  
Vaccine Strain ◽  
Culture System ◽  
Live Vaccine ◽  
Cell Mediated Immunity ◽  
Live Vaccine Strain ◽  
Wild Type

ABSTRACT Previous studies have demonstrated a role for B cells, not associated with antibody production, in protection against lethal secondary infection of mice with Francisella tularensislive vaccine strain (LVS). However, the mechanism by which B cells contribute to this protection is not known. To study the specific role of B cells during secondary LVS infection, we developed an in vitro culture system that mimics many of the same characteristics of in vivo infection. Using this culture system, we showed that B cells do not directly control LVS infection but that control of LVS growth is mediated primarily by LVS-primed T cells. Importantly, B cells were not required for the generation of effective memory T cells since LVS-primed, B-cell-deficient (BKO) mice generated CD4+ and CD8+ T cells that controlled LVS infection similarly to LVS-primed CD4+ and CD8+ T cells from wild-type mice. The control of LVS growth appeared to depend primarily on gamma interferon and nitric oxide and was similar in wild-type and BKO mice. Rather, the inability of BKO mice to survive secondary LVS infection was associated with marked neutrophil influx into the spleen very early after challenge. The neutrophilia was directly associated with B cells, since BKO mice reconstituted with naive B cells prior to a secondary challenge with LVS had decreased bacterial loads and neutrophils in the spleen and survived.

scholarly journals Respiratory Francisella tularensis Live Vaccine Strain Infection Induces Th17 Cells and Prostaglandin E2, Which Inhibits Generation of Gamma Interferon-Positive T Cells

2008 ◽  
Vol 76 (6) ◽  
pp. 2651-2659 ◽  
Author(s):  
Matthew D. Woolard ◽  
Lucinda L. Hensley ◽  
Thomas H. Kawula ◽  
Jeffrey A. Frelinger
Keyword(s):  
T Cells ◽  
Francisella Tularensis ◽  
Vaccine Strain ◽  
Th17 Cells ◽  
Live Vaccine ◽  
Prostaglandin E ◽  
Live Vaccine Strain ◽  
Intranasal Inoculation ◽  
Adaptive Immune ◽  
Ifn Γ

ABSTRACT Two key routes of Francisella tularensis infection are through the skin and airway. We wished to understand how the route of inoculation influenced the primary acute adaptive immune response. We show that an intranasal inoculation of the F. tularensis live vaccine strain (LVS) with a 1,000-fold-smaller dose than an intradermal dose results in similar growth kinetics and peak bacterial burdens. In spite of similar bacterial burdens, we demonstrate a difference in the quality, magnitude, and kinetics of the primary acute T-cell response depending on the route of inoculation. Further, we show that prostaglandin E2 secretion in the lung is responsible for the difference in the gamma interferon (IFN-γ) response. Intradermal inoculation led to a large number of IFN-γ+ T cells 7 days after infection in both the spleen and the lung. In contrast, intranasal inoculation induced a lower number of IFN-γ+ T cells in the spleen and lung but an increased number of Th17 cells in the lung. Intranasal infection also led to a significant increase of prostaglandin E2 (PGE2) in the bronchoalveolar lavage fluid. Inhibition of PGE2 production with indomethacin treatment resulted in increased numbers of IFN-γ+ T cells and decreased bacteremia in the lungs of intranasally inoculated mice. This research illuminates critical differences in acute adaptive immune responses between inhalational and dermal infection with F. tularensis LVS mediated by the innate immune system and PGE2.

scholarly journals Increased Susceptibility of IgA-Deficient Mice to Pulmonary Francisella tularensis Live Vaccine Strain Infection

2013 ◽  
Vol 81 (9) ◽  
pp. 3434-3441 ◽  
Author(s):  
Yoichi Furuya ◽  
Girish S. Kirimanjeswara ◽  
Sean Roberts ◽  
Dennis W. Metzger
Keyword(s):  
Francisella Tularensis ◽  
Vaccine Strain ◽  
Early Time ◽  
Iga Deficiency ◽  
Live Vaccine ◽  
Interleukin 12 ◽  
Live Vaccine Strain ◽  
Content Type ◽  
Ifn Γ ◽  
Increased Susceptibility

ABSTRACTFrancisella tularensis, the causative agent of tularemia, is most deadly in the pneumonic form; therefore, mucosal immunity is an important first line of defense against this pathogen. We have now evaluated the lethality of primaryF. tularensislive vaccine strain (LVS) pulmonary infection in mice that are defective in IgA (IgA−/−mice), the predominant mucosal Ig isotype. The results showed that IgA−/−mice were more susceptible than IgA+/+mice to intranasalF. tularensisLVS infection, despite developing higher levels of LVS-specific total, IgG, and IgM antibodies in the bronchoalveolar lavage specimens following infection. In addition, the absence of IgA resulted in a significant increase in bacterial loads and reduced survival. Interestingly, IgA−/−mice had lower pulmonary gamma interferon (IFN-γ) levels and decreased numbers of IFN-γ-secreting CD4+and CD8+T cells in the lung on day 9 postinfection compared to IgA+/+mice. Furthermore, IgA−/−mice displayed reduced interleukin 12 (IL-12) levels at early time points, and supplementing IgA−/−mice with IL-12 prior to LVS challenge induced IFN-γ production by NK cells and rescued them from mortality. Thus, IgA−/−mice are highly susceptible to primary pulmonary LVS infections not only because of IgA deficiency but also because of reduced IFN-γ responses.

scholarly journals Importance of B cells, but Not Specific Antibodies, in Primary and Secondary Protective Immunity to the Intracellular BacteriumFrancisella tularensis Live Vaccine Strain

1999 ◽  
Vol 67 (11) ◽  
pp. 6002-6007 ◽  
Author(s):  
Karen L. Elkins ◽  
Catharine M. Bosio ◽  
Tonya R. Rhinehart-Jones
Keyword(s):  
B Cells ◽  
Vaccine Strain ◽  
Protective Immunity ◽  
Secondary Infection ◽  
Live Vaccine ◽  
Intracellular Bacteria ◽  
Live Vaccine Strain ◽  
Lethal Challenge ◽  
Specific Antibodies

ABSTRACT Although there appears to be little if any role for specific antibodies in protection against intracellular bacteria, such as the model pathogen F. tularensis live vaccine strain (LVS), the role of B cells themselves in primary and secondary infection with such bacteria has not been examined directly. We show here that mice deficient in mature B cells and antibodies (B-cell knockout mice) are marginally compromised in controlling primary sublethal infection but are 100-fold less well protected against secondary lethal challenge than are their normal counterparts. This defect in optimal specific protective immunity was readily reconstituted by the transfer of primed, and to a lesser degree, unprimed B cells, but not by the transfer of specific antibodies. The results indicate a previously unappreciated role for B cells in secondary immunity to intracellular pathogens through a function other than antibody production.

scholarly journals Inactivated Francisella tularensis Live Vaccine Strain Protects against Respiratory Tularemia by Intranasal Vaccination in an Immunoglobulin A-Dependent Fashion

2007 ◽  
Vol 75 (5) ◽  
pp. 2152-2162 ◽  
Author(s):  
Shawn D. Baron ◽  
Rajendra Singh ◽  
Dennis W. Metzger
Keyword(s):  
Francisella Tularensis ◽  
Vaccine Strain ◽  
Serum Antibody ◽  
Immunoglobulin A ◽  
Live Vaccine ◽  
Interleukin 12 ◽  
Live Vaccine Strain ◽  
Lethal Challenge ◽  
Mucosal Vaccination ◽  
Ifn Γ

ABSTRACT Francisella tularensis is a gram-negative intracellular bacterium that is considered to be a potential category A biological weapon due to its extreme virulence. Although vaccination with the attenuated live vaccine strain (LVS) of F. tularensis can protect against lethal challenge, use of inactivated or subunit forms as vaccine candidates for induction of protective antibody responses has not been fully evaluated. In the present study, we examined whether immune protection in the lung could be stimulated by intranasal administration of inactivated LVS together with interleukin-12 (IL-12) as an adjuvant. LVS was inactivated by heat, paraformaldehyde treatment, or exposure to UV, and inactivation of the preparations was confirmed by assessing bacterial growth and the survival of mice after direct inoculation. We found that mucosal vaccination with inactivated LVS provided 90 to 100% protection in mice after lethal intranasal challenge with 104 CFU of LVS, and this protection was dependent on inclusion of exogenous IL-12 during vaccine administration. Survival of vaccinated mice after live bacterial challenge was correlated with reduced bacterial burden, decreased pulmonary inflammation, increased serum antibody titers, and lower levels of gamma interferon (IFN-γ), tumor necrosis factor alpha, and IL-6 in the lungs, livers, and spleens. Whereas NK cells were primarily responsible for the production of IFN-γ in unvaccinated, challenged animals, vaccinated mice had increased levels of lung IFN-γ+ CD4+ T cells after challenge. Significantly, mice genetically deficient in immunoglobulin A (IgA) expression were unable to survive lethal challenge after vaccination. These results are the first results to demonstrate that IgA-mediated protection against lethal respiratory tularemia occurs after mucosal vaccination with inactivated F. tularensis LVS.

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