RESUMO
Despite the great increase in the understanding of the biology and pathogenesis of Mycobacterium tuberculosis achieved by the scientific community in recent decades, tuberculosis (TB) still represents one of the major threats to global human health. The only available vaccine (Mycobacterium bovis BCG) protects children from disseminated forms of TB but does not effectively protect adults from the respiratory form of the disease, making the development of new and more-efficacious vaccines against the pulmonary forms of TB a major goal for the improvement of global health. Among the different strategies being developed to reach this goal is the construction of attenuated strains more efficacious and safer than BCG. We recently showed that a sigE mutant of M. tuberculosis was more attenuated and more efficacious than BCG in a mouse model of infection. In this paper, we describe the construction and characterization of an M. tuberculosissigE fadD26 unmarked double mutant fulfilling the criteria of the Geneva Consensus for entering human clinical trials. The data presented suggest that this mutant is even more attenuated and slightly more efficacious than the previous sigE mutant in different mouse models of infection and is equivalent to BCG in a guinea pig model of infection.
Assuntos
Ligases/deficiência , Mycobacterium tuberculosis/imunologia , Fator sigma/deficiência , Vacinas contra a Tuberculose/imunologia , Tuberculose Pulmonar/prevenção & controle , Animais , Proteínas de Bactérias , Modelos Animais de Doenças , Cobaias , Camundongos , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/patogenicidade , Vacinas contra a Tuberculose/efeitos adversos , Vacinas contra a Tuberculose/genética , Vacinas Atenuadas/efeitos adversos , Vacinas Atenuadas/genética , Vacinas Atenuadas/imunologia , VirulênciaRESUMO
The diagnosis of leprosy continues to be based on clinical symptoms and early diagnosis and treatment are critical to preventing disability and transmission. Sensitive and specific laboratory tests are not available for diagnosing leprosy. Despite the limited applicability of anti-phenolic glycolipid-I (PGL-I) serology for diagnosis, it has been suggested as an additional tool to classify leprosy patients (LPs) for treatment purposes. Two formats of rapid tests to detect anti-PGL-I antibodies [ML immunochromatography assay (ICA) and ML Flow] were compared in different groups, multibacillary patients, paucibacillary patients, household contacts and healthy controls in Brazil and Nepal. High ML Flow intra-test concordance was observed and low to moderate agreement between the results of ML ICA and ML Flow tests on the serum of LPs was observed. LPs were "seroclassified" according to the results of these tests and the seroclassification was compared to other currently used classification systems: the World Health Organization operational classification, the bacilloscopic index and the Ridley-Jopling classification. When analysing the usefulness of these tests in the operational classification of PB and MB leprosy for treatment and follow-up purposes, the ML Flow test was the best point-of-care test for subjects in Nepal and despite the need for sample dilution, the ML ICA test yielded better performance among Brazilian subjects. Our results identified possible ways to improve the performance of both tests.
Assuntos
Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Criança , Pré-Escolar , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Adulto Jovem , Antígenos de Bactérias/sangue , Glicolipídeos/sangue , Isotipos de Imunoglobulinas/sangue , Hanseníase/diagnóstico , Mycobacterium leprae/imunologia , Brasil , Estudos de Casos e Controles , Imunoensaio/métodos , Cromatografia de Afinidade/métodos , Hanseníase/imunologia , Nepal , Valor Preditivo dos Testes , Kit de Reagentes para Diagnóstico , Sensibilidade e EspecificidadeRESUMO
Leprosy is not eradicable with currently available diagnostics or interventions, as evidenced by its stable incidence. Early diagnosis of Mycobacterium leprae infection should therefore be emphasized in leprosy research. It remains challenging to develop tests based on immunological biomarkers that distinguish individuals controlling bacterial replication from those developing disease. To identify biomarkers for field-applicable diagnostics, we determined cytokines/chemokines induced by M. leprae proteins in blood of leprosy patients and endemic controls (EC) from high leprosy-prevalence areas (Bangladesh, Brazil, Ethiopia) and from South Korea, where leprosy is not endemic anymore. M. leprae-sonicate-induced IFN-γ was similar for all groups, excluding M. leprae/IFN-γ as a diagnostic readout. By contrast, ML2478 and ML0840 induced high IFN-γ concentrations in Bangladeshi EC, which were completely absent for South Korean controls. Importantly, ML2478/IFN-γ could indicate distinct degrees of M. leprae exposure, and thereby the risk of infection and transmission, in different parts of Brazilian and Ethiopian cities. Notwithstanding these discriminatory responses, M. leprae proteins did not distinguish patients from EC in one leprosy-endemic area based on IFN-γ. Analyses of additional cytokines/chemokines showed that M. leprae and ML2478 induced significantly higher concentrations of MCP-1, MIP-1ß, and IL-1ß in patients compared with EC, whereas IFN-inducible protein-10, like IFN-γ, differed between EC from areas with dissimilar leprosy prevalence. This study identifies M. leprae-unique Ags, particularly ML2478, as biomarker tools to measure M. leprae exposure using IFN-γ or IFN-inducible protein-10, and also shows that MCP-1, MIP-1ß, and IL-1ß can potentially distinguish pathogenic immune responses from those induced during asymptomatic exposure to M. leprae.
Assuntos
Citocinas/sangue , Hanseníase/epidemiologia , Hanseníase/imunologia , Mycobacterium leprae/imunologia , Adulto , Idoso , Antígenos de Bactérias/imunologia , Bangladesh/epidemiologia , Biomarcadores/sangue , Brasil/epidemiologia , Citocinas/biossíntese , Citocinas/genética , Etiópia/epidemiologia , Feminino , Humanos , Interferon gama/biossíntese , Interferon gama/sangue , Interferon gama/genética , Hanseníase/diagnóstico , Masculino , Pessoa de Meia-Idade , República da Coreia/epidemiologia , Células Th1/imunologia , Células Th1/microbiologia , Células Th2/imunologia , Células Th2/microbiologia , Adulto JovemRESUMO
The diagnosis of leprosy continues to be based on clinical symptoms and early diagnosis and treatment are critical to preventing disability and transmission. Sensitive and specific laboratory tests are not available for diagnosing leprosy. Despite the limited applicability of anti-phenolic glycolipid-I (PGL-I) serology for diagnosis, it has been suggested as an additional tool to classify leprosy patients (LPs) for treatment purposes. Two formats of rapid tests to detect anti-PGL-I antibodies [ML immunochromatography assay (ICA) and ML Flow] were compared in different groups, multibacillary patients, paucibacillary patients, household contacts and healthy controls in Brazil and Nepal. High ML Flow intra-test concordance was observed and low to moderate agreement between the results of ML ICA and ML Flow tests on the serum of LPs was observed. LPs were "seroclassified" according to the results of these tests and the seroclassification was compared to other currently used classification systems: the World Health Organization operational classification, the bacilloscopic index and the Ridley-Jopling classification. When analysing the usefulness of these tests in the operational classification of PB and MB leprosy for treatment and follow-up purposes, the ML Flow test was the best point-of-care test for subjects in Nepal and despite the need for sample dilution, the ML ICA test yielded better performance among Brazilian subjects. Our results identified possible ways to improve the performance of both tests.
Assuntos
Antígenos de Bactérias/sangue , Glicolipídeos/sangue , Isotipos de Imunoglobulinas/sangue , Hanseníase/diagnóstico , Mycobacterium leprae/imunologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Brasil , Estudos de Casos e Controles , Criança , Pré-Escolar , Cromatografia de Afinidade/métodos , Feminino , Humanos , Imunoensaio/métodos , Hanseníase/imunologia , Masculino , Pessoa de Meia-Idade , Nepal , Valor Preditivo dos Testes , Kit de Reagentes para Diagnóstico , Sensibilidade e Especificidade , Adulto JovemRESUMO
The detection of hundreds of thousands of new cases of leprosy every year suggests that transmission of Mycobacterium leprae infection still continues. Unfortunately, tools for identification of asymptomatic disease and/or early-stage M. leprae infection (likely sources of transmission) are lacking. The recent identification of M. leprae-unique genes has allowed the analysis of human T-cell responses to novel M. leprae antigens. Antigens with the most-promising diagnostic potential were tested for their ability to induce cytokine secretion by using peripheral blood mononuclear cells from leprosy patients and controls in five different areas where leprosy is endemic; 246 individuals from Brazil, Nepal, Bangladesh, Pakistan, and Ethiopia were analyzed for gamma interferon responses to five recombinant proteins (ML1989, ML1990, ML2283, ML2346, and ML2567) and 22 synthetic peptides. Of these, the M. leprae-unique protein ML1989 was the most frequently recognized and ML2283 the most specific for M. leprae infection/exposure, as only a limited number of tuberculosis patients responded to this antigen. However, all proteins were recognized by a significant number of controls in areas of endemicity. T-cell responses correlated with in vitro response to M. leprae, suggesting that healthy controls in areas where leprosy is endemic are exposed to M. leprae. Importantly, 50% of the healthy household contacts and 59% of the controls in areas of endemicity had no detectable immunoglobulin M antibodies to M. leprae-specific PGL-I but responded in T-cell assays to >or=1 M. leprae protein. T-cell responses specific for leprosy patients and healthy household contacts were observed for ML2283- and ML0126-derived peptides, indicating that M. leprae peptides hold potential as diagnostic tools. Future work should concentrate on the development of a sensitive and field-friendly assay and identification of additional peptides and proteins that can induce M. leprae-specific T-cell responses.
Assuntos
Interferon gama/biossíntese , Hanseníase/diagnóstico , Mycobacterium leprae/imunologia , Linfócitos T/imunologia , Adulto , Antígenos de Bactérias , Bangladesh , Brasil , Etiópia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Nepal , Paquistão , Proteínas Recombinantes , Sensibilidade e Especificidade , Adulto JovemRESUMO
Leprosy, a chronic human disease with potentially debilitating neurological consequences, results from infection with Mycobacterium leprae. This unculturable pathogen has undergone extensive reductive evolution, with half of its genome now occupied by pseudogenes. Using comparative genomics, we demonstrated that all extant cases of leprosy are attributable to a single clone whose dissemination worldwide can be retraced from analysis of very rare single-nucleotide polymorphisms. The disease seems to have originated in Eastern Africa or the Near East and spread with successive human migrations. Europeans or North Africans introduced leprosy into West Africa and the Americas within the past 500 years.
Assuntos
Humanos , História Antiga , História Medieval , História do Século XVIII , História do Século XIX , Ásia/epidemiologia , América/epidemiologia , Pseudogenes , Genoma Bacteriano , Análise de Sequência com Séries de Oligonucleotídeos , Polimorfismo de Nucleotídeo Único , África/epidemiologia , Emigração e Imigração , Europa (Continente)/epidemiologia , Genes Bacterianos , Hanseníase/história , Hanseníase/microbiologia , Hanseníase/transmissão , Hanseníase/epidemiologia , Mycobacterium leprae/classificação , Mycobacterium leprae/genéticaRESUMO
Only native products of Mycobacterium leprae, whether cell wall, cytosol, or membrane derived, can confer protective immunity against challenge in the mouse footpad. Previously, recombinant proteins were shown to be ineffective. The cell wall skeleton-the mycolyl-arabinogalactan-peptidoglycan complex-devoid of proteins is not protective.