Molecular basis for improved anthrax vaccines.

Granules of mast cells (MCs) enhance adaptive immunity when, on activation, they are released as stable particles. Here we show that submicrometre particles modelled after MC granules augment immunity when used as adjuvants in vaccines. The synthetic particles, which consist of a carbohydrate backbone with encapsulated inflammatory mediators such as tumour necrosis factor, replicate attributes of MCs in vivo including the targeting of draining lymph nodes and the timed release of the encapsulated mediators. When used as an adjuvant during vaccination of mice with haemagglutinin from the influenza virus, the particles enhanced adaptive immune responses and increased survival of mice on lethal challenge. Furthermore, differential loading of the particles with the cytokine IL-12 directed the character of the response towards Th1 lymphocytes. The synthetic MC adjuvants replicate and enhance the functions of MCs during vaccination, and can be extended to polarize the resulting immunity.

Synthetic mast-cell granules as adjuvants to promote and polarize immunity in lymph nodes

Here we describe the characterization of a lipoprotein previously proposed as a potential Bacillus anthracis virulence determinant and vaccine candidate. This protein, designated MntA, is the solute-binding component of a manganese ion ATP-binding cassette transporter. Coupled proteomic-serological screen of a fully virulent wild-type B. anthracis Vollum strain, confirmed that MntA is expressed both in vitro and during infection. Expression of MntA is shown to be independent of the virulence plasmids pXO1 and pXO2. An mntA deletion, generated by allelic replacement, results in complete loss of MntA expression and its phenotypic analysis revealed: (i) impaired growth in rich media, alleviated by manganese supplementation; (ii) increased sensitivity to oxidative stress; and (iii) delayed release from cultured macrophages. The DeltamntA mutant expresses the anthrax-associated classical virulence factors, lethal toxin and capsule, in vitro as well as in vivo, and yet the mutation resulted in severe attenuation; a 10(4)-fold drop in LD(50) in a guinea pig model. MntA expressed in trans allowed to restore, almost completely, the virulence of the DeltamntA B. anthracis strain. We propose that MntA is a novel B. anthracis virulence determinant essential for the development of anthrax disease, and that B. anthracisDeltamntA strains have the potential to serve as platform for future live attenuated vaccines.

The solute-binding component of a putative Mn(II) ABC transporter (MntA) is a novel Bacillus anthracis virulence determinant.

Protective host immune responses to anthrax infection in humans and animal models are characterized by the development of neutralizing Abs against the receptor-binding anthrax protective Ag (PA), which, together with the lethal factor (LF) protease, composes anthrax lethal toxin (LT). We now report that B cells, in turn, are targets for LT. Anthrax PA directly binds primary B cells, resulting in the LF-dependent cleavage of the MAPK kinases (MAPKKs) and disrupted signaling to downstream MAPK targets. Although not directly lethal to B cells, anthrax LT treatment causes severe B cell dysfunction, greatly reducing proliferative responses to IL-4-, anti-IgM-, and/or anti-CD40 stimulation. Moreover, B cells treated with anthrax LT in vitro or isolated from mice treated with anthrax LT in vivo have a markedly diminished capacity to proliferate and produce IgM in response to TLR-2 and TLR-4 ligands. The suppressive effects of anthrax LT on B cell function occur at picomolar concentrations in vitro and at sublethal doses in vivo. These results indicate that anthrax LT directly inhibits the function of B cells in vitro and in vivo, revealing a potential mechanism through which the pathogen could bypass protective immune responses.

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Anthrax Lethal Toxin Has Direct and Potent Inhibitory Effects on B Cell Proliferation and Immunoglobulin Production

Bacterial infections are common in tropical parts of the world and can include those species also seen regularly in temperate climates. Many tropical bacterial infections, however, are rarely diagnosed in temperate parts of the world and include bartonellosis, tropical ulcer, tropical pyomyositis, granuloma inguinale, lymphogranuloma venereum, yaws, pinta, melioidosis, and glanders. Some tropical bacterial diseases, eg, plague and anthrax, are associated with high mortality rates and are of potential use in bioterrorism. Some tropical bacterial diseases are closely associated with specific activities such as hunting (ie, tularemia) or eating raw seafood ( Vibrio vulnificus infection). The bacterial diseases having the most severe medical impact in the tropics are those caused by members of the Mycobacterium genus. Millions of persons throughout the world suffer from tuberculosis and leprosy; Buruli ulcers are common causes of morbidity in many tropical countries. Because of the increasing frequency of travel to tropical parts of the world for tourism and work as well as the increasing number of immigrants and adoptees from these areas, it is imperative that physicians practicing in temperate climates be able to recognize the signs and symptoms of tropical bacterial diseases, carry out the proper diagnostic tests, and initiate appropriate therapy and prevention. Learning objective At the completion of this learning activity, participants should be familiar with the clinical presentations, epidemiologies, diagnoses, therapies, and preventions of bacterial tropical diseases.

Tropical dermatology: Bacterial tropical diseases

Protective antigen (PA)-based vaccination is an effective countermeasure to anthrax infection. While neutralizing anti-PA antibody titers elicited by this vaccine serve as good correlates for protection against anthrax (S. Reuveny, M. D. White, Y. Y. Adar, Y. Kafri, Z. Altboum, Y. Gozes, D. Kobiler, A. Shafferman, and B. Velan, Infect. Immun. 69:2888-2893, 2001), no data are available on the contribution of the immunological memory for PA itself to protection. We therefore developed a guinea pig model in which a primary immunization with threshold levels of PA can induce a long-term T-cell immunological memory response without inducing detectable anti-PA antibodies. A revaccination of primed animals with the same threshold PA levels was effective for memory activation, yielding a robust and rapid secondary response. A challenge with a lethal dose (40 50% lethal doses; 2,000 spores) of spores after the booster vaccinations indicated that animals were not protected at days 2, 4, and 6 postboosting. Protection was achieved only from the 8th day postboosting, concomitant with the detection of protective levels of neutralizing antibody titers in the circulation. The practical implications from the studies reported herein are that, as expected, the protective capacity of memory depends on the PA dose used for the primary immunization and that the effectiveness of booster immunizations for the postexposure treatment of anthrax may be very limited when no detectable antibodies are present in primed animals prior to Bacillus anthracis spore exposure. Therefore, to allow for the establishment of memory-dependent protection prior to the expected onset of disease, booster immunizations should not be used without concomitant antimicrobial treatment in postexposure scenarios.

Rachel Danieli

Papers

Contribution of Immunological Memory to Protective Immunity Conferred by a Bacillus anthracis Protective Antigen-Based Vaccine