Decreased survival of guinea pigs infected with Mycobacterium tuberculosis after multiple BCG vaccinations.

The BCG vaccine for tuberculosis has an outstanding safety record, with only occasional reports of adverse reactions. In some countries multiple BCG vaccinations have been given to children, but with no apparent benefit. We show here that in the highly susceptible guinea pig model of Mycobacterium tuberculosis infection animals receiving three sequential injections of BCG began to die unexpectedly at times after aerosol challenge much earlier than animals receiving the vaccine only once. Animals receiving three injections with BCG had extensive, disseminated lesions in the lungs, spleen and liver that were similar to non-vaccinated animals. In these animals there was effacement of the pulmonary, hepatic and splenic architecture by extensive areas of fibrosis with residual granulomatous inflammation and necrosis. This contrasted to the saline control group in which active inflammation was accompanied by extensive areas of necrosis but with less fibrosis in the lungs and spleen. Granulomatous inflammation was less severe and was not complicated by necrosis or extensive fibrosis in animals receiving a single injection with BCG. These data indicate that multiple BCG vaccination, which is akin to super-infection with the living vaccine, leads to major organ failure pathology.

NIH pre-clinical screening program: overview and current status.

The increase in the number of cases of Mycobacterium tuberculosis around the world has lead to a greater need for a more efficacious vaccine than the currently used M. bovis BCG. Despite the relative success of this attenuated vaccine there are multiple examples where alternative strategies are desperately needed. In 1996, the National Institutes of Health published a request calling for applications to test newly developed vaccines against tuberculosis. The current screening program at Colorado State University has tested a wide range of novel vaccine candidates.

Mice fed lipid-encapsulated Mycobacterium bovis BCG are protected against aerosol challenge with Mycobacterium tuberculosis.

Mice that consumed a single dose of 10(7) lipid-encapsulated Mycobacterium bovis BCG bacilli showed significant pulmonary and systemic protection against aerosol challenge with M. tuberculosis H37Rv. As an extension of previous challenge studies with virulent strains of M. bovis, this report describes a reduction in M. tuberculosis infection in mice vaccinated orally with lipid-encapsulated BCG comparable to that observed in mice vaccinated subcutaneously with BCG. These results are consistent with the induction of tuberculin-specific cell-mediated immune responses.

The protective effect of the Mycobacterium bovis BCG vaccine is increased by coadministration with the Mycobacterium tuberculosis 72-kilodalton fusion polyprotein Mtb72F in M. tuberculosis-infected guinea pigs.

A tuberculosis vaccine candidate consisting of a 72-kDa polyprotein or fusion protein based upon the Mtb32 and Mtb39 antigens of Mycobacterium tuberculosis and designated Mtb72F was tested for its protective capacity as a potential adjunct to the Mycobacterium bovis BCG vaccine in the mouse and guinea pig models of this disease. Formulation of recombinant Mtb72F (rMtb72F) in an AS02A adjuvant enhanced the Th1 response to BCG in mice but did not further reduce the bacterial load in the lungs after aerosol challenge infection. In the more stringent guinea pig disease model, rMtb72F delivered by coadministration with BCG vaccination significantly improved the survival of these animals compared to BCG alone, with some animals still alive and healthy in their appearance at >100 weeks post-aerosol challenge. A similar trend was observed with guinea pigs in which BCG vaccination was boosted by DNA vaccination, although this increase was not statistically significant due to excellent protection conferred by BCG alone. Histological examination of the lungs of test animals indicated that while BCG controls eventually died from overwhelming lung consolidation, the majority of guinea pigs receiving BCG mixed with rMtb72F or boosted twice with Mtb72F DNA had mostly clear lungs with minimal granulomatous lesions. Lesions were still prominent in guinea pigs receiving BCG and the Mtb72F DNA boost, but there was considerable evidence of lesion healing and airway remodeling and reestablishment. These data support the hypothesis that the coadministration or boosting of BCG vaccination with Mtb72F may limit the lung consolidation seen with BCG alone and may promote lesion resolution and healing. Collectively, these data suggest that enhancing BCG is a valid vaccination strategy for tuberculosis that is worthy of clinical evaluation.

Differential immune responses and protective efficacy induced by components of a tuberculosis polyprotein vaccine, Mtb72F, delivered as naked DNA or recombinant protein.

Key Ags of Mycobacterium tuberculosis initially identified in the context of host responses in healthy purified protein derivative-positive donors and infected C57BL/6 mice were prioritized for the development of a subunit vaccine against tuberculosis. Our lead construct, Mtb72F, codes for a 72-kDa polyprotein genetically linked in tandem in the linear order Mtb32(C)-Mtb39-Mtb32(N). Immunization of C57BL/6 mice with Mtb72F DNA resulted in the generation of IFN-gamma responses directed against the first two components of the polyprotein and a strong CD8(+) T cell response directed exclusively against Mtb32(C). In contrast, immunization of mice with Mtb72F protein formulated in the adjuvant AS02A resulted in the elicitation of a moderate IFN-gamma response and a weak CD8(+) T cell response to Mtb32c. However, immunization with a formulation of Mtb72F protein in AS01B adjuvant generated a comprehensive and robust immune response, resulting in the elicitation of strong IFN-gamma and Ab responses encompassing all three components of the polyprotein vaccine and a strong CD8(+) response directed against the same Mtb32(C) epitope identified by DNA immunization. All three forms of Mtb72F immunization resulted in the protection of C57BL/6 mice against aerosol challenge with a virulent strain of M. tuberculosis. Most importantly, immunization of guinea pigs with Mtb72F, delivered either as DNA or as a rAg-based vaccine, resulted in prolonged survival (>1 year) after aerosol challenge with virulent M. tuberculosis comparable to bacillus Calmette-Guérin immunization. Mtb72F in AS02A formulation is currently in phase I clinical trial, making it the first recombinant tuberculosis vaccine to be tested in humans.

Failure of the Mycobacterium bovis BCG vaccine: some species of environmental mycobacteria block multiplication of BCG and induction of protective immunity to tuberculosis.

The efficacy of Mycobacterium bovis bacillus Calmette-Guérin (BCG) vaccine against pulmonary tuberculosis (TB) varies enormously in different populations. The prevailing hypothesis attributes this variation to interactions between the vaccine and mycobacteria common in the environment, but the precise mechanism has so far not been clarified. Our study demonstrates that prior exposure to live environmental mycobacteria can result in a broad immune response that is recalled rapidly after BCG vaccination and controls the multiplication of the vaccine. In these sensitized mice, BCG elicits only a transient immune response with a low frequency of mycobacterium-specific cells and no protective immunity against TB. In contrast, the efficacy of TB subunit vaccines was unaffected by prior exposure to environmental mycobacteria. Six different isolates from soil and sputum samples from Karonga district in Northern Malawi (a region in which BCG vaccination has no effect against pulmonary TB) were investigated in the mouse model, and two strains of the Mycobacterium avium complex were found to block BCG activity completely.