Live Attenuated Tularemia Vaccines for Protection Against Respiratory Challenge With Virulent F. tularensis subsp. tularensis.

Francisella tularensis is the causative agent of tularemia and a Tier I bioterrorism agent. In the 1900s, several vaccines were developed against tularemia including the killed "Foshay" vaccine, subunit vaccines comprising F. tularensis protein(s) or lipoproteins(s) in an adjuvant formulation, and the F. tularensis Live Vaccine Strain (LVS); none were licensed in the U.S.A. or European Union. The LVS vaccine retains toxicity in humans and animals-especially mice-but has demonstrated efficacy in humans, and thus serves as the current gold standard for vaccine efficacy studies. The U.S.A. 2001 anthrax bioterrorism attack spawned renewed interest in vaccines against potential biowarfare agents including F. tularensis. Since live attenuated-but not killed or subunit-vaccines have shown promising efficacy and since vaccine efficacy against respiratory challenge with less virulent subspecies holarctica or F. novicida, or against non-respiratory challenge with virulent subsp. tularensis (Type A) does not reliably predict vaccine efficacy against respiratory challenge with virulent subsp. tularensis, the route of transmission and species of greatest concern in a bioterrorist attack, in this review, we focus on live attenuated tularemia vaccine candidates tested against respiratory challenge with virulent Type A strains, including homologous vaccines derived from mutants of subsp. holarctica, F. novicida, and subsp. tularensis, and heterologous vaccines developed using viral or bacterial vectors to express F. tularensis immunoprotective antigens. We compare the virulence and efficacy of these vaccine candidates with that of LVS and discuss factors that can significantly impact the development and evaluation of live attenuated tularemia vaccines. Several vaccines meet what we would consider the minimum criteria for vaccines to go forward into clinical development-safety greater than LVS and efficacy at least as great as LVS, and of these, several meet the higher standard of having efficacy ≥LVS in the demanding mouse model of tularemia. These latter include LVS with deletions in purMCD, sodBFt , capB or wzy; LVS ΔcapB that also overexpresses Type VI Secretion System (T6SS) proteins; FSC200 with a deletion in clpB; the single deletional purMCD mutant of F. tularensis SCHU S4, and a heterologous prime-boost vaccine comprising LVS ΔcapB and Listeria monocytogenes expressing T6SS proteins.

Humanized Mice for the Evaluation of Francisella tularensis Vaccine Candidates.

Francisella tularensis (FT), a highly infectious pathogen, is considered to be a potential biological weapon owing to the current lack of a human vaccine against it. Tul4 and FopA, both outer membrane proteins of FT, play an important role in the bacterium's immunogenicity. In the present study, we evaluated the immune response of mice-humanized with human CD34+ cells (hu-mice)-to a cocktail of recombinant Tul4 and FopA (rTul4 and rFopA), which were codon-optimized and expressed in Escherichia coli. Not only did the cocktail-immunized hu-mice produce a significant human immunoglobulin response, they also exhibited prolonged survival against an attenuated live vaccine strain as well as human T cells in the spleen. These results suggest that the cocktail of rTul4 and rFopA had successfully induced an immune response in the hu-mice, demonstrating the potential of this mouse model for use in the evaluation of FT vaccine candidates.

Nasal Acai polysaccharides potentiate innate immunity to protect against pulmonary Francisella tularensis and Burkholderia pseudomallei Infections.

Pulmonary Francisella tularensis and Burkholderia pseudomallei infections are highly lethal in untreated patients, and current antibiotic regimens are not always effective. Activating the innate immune system provides an alternative means of treating infection and can also complement antibiotic therapies. Several natural agonists were screened for their ability to enhance host resistance to infection, and polysaccharides derived from the Acai berry (Acai PS) were found to have potent abilities as an immunotherapeutic to treat F. tularensis and B. pseudomallei infections. In vitro, Acai PS impaired replication of Francisella in primary human macrophages co-cultured with autologous NK cells via augmentation of NK cell IFN-γ. Furthermore, Acai PS administered nasally before or after infection protected mice against type A F. tularensis aerosol challenge with survival rates up to 80%, and protection was still observed, albeit reduced, when mice were treated two days post-infection. Nasal Acai PS administration augmented intracellular expression of IFN-γ by NK cells in the lungs of F. tularensis-infected mice, and neutralization of IFN-γ ablated the protective effect of Acai PS. Likewise, nasal Acai PS treatment conferred protection against pulmonary infection with B. pseudomallei strain 1026b. Acai PS dramatically reduced the replication of B. pseudomallei in the lung and blocked bacterial dissemination to the spleen and liver. Nasal administration of Acai PS enhanced IFN-γ responses by NK and γδ T cells in the lungs, while neutralization of IFN-γ totally abrogated the protective effect of Acai PS against pulmonary B. pseudomallei infection. Collectively, these results demonstrate Acai PS is a potent innate immune agonist that can resolve F. tularensis and B. pseudomallei infections, suggesting this innate immune agonist has broad-spectrum activity against virulent intracellular pathogens.

[Evaluation of immunobiological activity of Francisella tularensis C-complex preparations as promising component of subunit vaccines].

Data on influence of Francisella tularensis C-complex preparations on formation of immunity against tularemia are presented. Study of cellular immunity characteristics as well as dynamics of antibody response was carried out on white mice and guinea pigs models. Absence of toxicity, pyrogenicity, and negative effects on immunocompetent cells in combination with protective activity points to possibility of use the C-complex as a component of a subunit vaccine.

Preparing for bioterrorism: category A agents.

September 11, 2001, brought the possibility of biologic acts of terrorism against the United States into the national consciousness. As the American people brace themselves for this new threat to the national well-being, clinicians must understand how to prevent, recognize, and treat the biologic agents that could be used in terrorist attacks. This article discusses the most likely biologic agents, including diagnostic laboratory procedures, treatment options, psychological effects, special populations, and reporting requirements.

Do homeopathic nosodes protect against infection? An experimental test.

CONTEXT: For centuries, homeopathic practitioners have claimed that serially agitated dilutions of infectious agents (called "nosodes") are effective in the prevention of infectious disease. However, no rigorous tests of this claim have been performed. OBJECTIVE: To test whether a nosode of Francisella tularensis-infected tissue could protect from subsequent challenge with this pathogen in vivo. DESIGN: Experimental laboratory test. SETTING: A P3 containment laboratory at an infectious disease research facility. PARTICIPANTS: 142 male C3H/HeN specific, pathogen-free mice. INTERVENTION: Six levels of a nosode prepared from tularemia-infected tissue were produced. All exposures were below the lowest level at which a classical vaccination response was expected. The nosode and dilutent control solutions were administered orally (.03 mL, 3 times per week) for 1 month before and after challenge. Animals were challenged with a potentially lethal dose (LD50 or LD75) of F tularensis, then evaluated for time of death and total mortality. MAIN OUTCOME MEASURES: Mortality and time to death. RESULTS: In a series of 15 trials (n = 142), the tularemia nosode consistently produced increased mean times to death. All but 2 of 15 trials showed reduced time to death in the nosode group and decreased mortality compared with controls. Protection rates averaged 22% over controls compared to 100% protection by standard vaccination. CONCLUSIONS: This study found partial protection from a nosode of tularemia in dilutions below those expected to have protective effects, but not as great as those produced by standard vaccination. If homeopathic nosodes can induce protection from infectious agents for which vaccination is currently unavailable, they may provide an interim method of reducing morbidity or mortality from such agents.

The efficacy of ciprofloxacin and doxycycline against experimental tularaemia.

The efficacy of doxycycline and ciprofloxacin against an experimental tularaemia infection was assessed by comparing the median lethal dose (MLD) of Francisella tularensis Schu4 biotype A strain given intraperitoneally to antibiotic-treated and untreated mice. In untreated Porton outbred mice this was <1 cfu. Ciprofloxacin and doxycycline given at 40 mg/kg bd, initiated 48 h before infection and continued for 5 days after infection, afforded protection against intraperitoneal challenges of 3.7 x 10(6) cfu and 6.0 x 10(6) cfu, respectively. Protection was reduced when both antibiotics were given over a similar period at a lower dose regimen (20 mg/kg bd) to 8.8 x 10(5) cfu and 3.5 x 10(2) cfu, respectively. The greater reduction in protection offered by doxycycline was a reflection of the higher in-vitro MIC. Protection also decreased when the antibiotics were initiated 24 h after challenge. The MLD was 3.2 x 10(5) cfu and 1.6 x 10(6) cfu for ciprofloxacin and doxycycline respectively given at 40 mg/kg bd and was reduced further using the lower dose regimen. Overall, 90% of the deaths occurred following the withdrawal of antibiotic, irrespective of the antibiotic dose or type. It was possible to prevent this relapse by extending the antibiotic administration to 10 days after challenge. Ciprofloxacin and doxycycline may be useful for treating tularaemia, although the possibility of relapse should be considered.

[The immunological efficacy of Francisella tularensis outer membranes for hamadryas baboons]. / Immunologicheskaia éffektivnost' vneshnikh membran Francisella tularensis dlia obez'ian-gamadril.

The protective properties of the preparation of F. tularensis outer membranes (OM), obtained from F. tularensis vaccine strain 15, were studied in experiments on hamadryas baboons challenged subcutaneously with F. tularensis virulent strain Schu (nonarctic subspecies). The subcutaneous immunization with the OM preparation prevented the development of clinically pronounced infection in more than 70% of the monkeys challenged with F. tularensis strain Schu in a dose of 787 live microbial cells 30 days after immunization. Antibody titers determined in the immunized monkeys with the use of the agglutination test (AT) and the passive hemagglutination test (PHAT) were usual in minimal diagnostic limits (1:80 for AT and 1:320 for PHAT) and did not significantly rise by day 20 after immunization. In all intact animals infected with F. tularensis strain Schu the development of the infectious process was registered, which was accompanied by a rise in temperature exceeding 39.5 degrees C and a rise in the titer of specific antibodies.

[The protective properties of the outer membranes of Francisella tularensis in an experimental infection in guinea pigs]. / Zashchitnye svoistva vneshnykh membran Francisella tularensis pri éksperimental'noi infektsii morskikh svinok.

Subcutaneous immunization, made in a single injection, with outer membrane preparations obtained from F.tularensis vaccine strain 15 and virulent strain A'Cole results in intensive immunity to tularemia in guinea pigs, ensuring the protection of 60-100% of the animals within a month after challenge with F.tularensis virulent strain 503 in a dose of 1,000 DCL. The development of protective effect induced by F.tularensis outer membranes can be observed during the first 24 hours and reaches its maximum by days 15-21 after immunization.

[The preventive activity of monoclonal antibodies specific to the lipopolysaccharide of Francisella tularensis]. / Preventivnaia aktivnost' monoklonal'nykh antitel, spetsifichnykh k lipopolisakharidu Francisella tularensis.

The preventive activity of five monoclonal antibodies (McAb) in experimental tularemia was evaluated. McAb produced by hybridoma FB11-k (IgG2a), specific to F. tularensis lipopolysaccharide, prevented the death of mice and guinea pigs infected with F. tularensis virulent strain 503 of the holarctic subspecies.