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Nontuberculous mycobacterial cutaneous infection confirmed by biochemical tests, polymerase chain reaction-restriction fragment length polymorphism analysis and sequencing of hsp65 gene.  

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We report a woman in whom a slow-growing scotochromogenic strain of Mycobacterium was cultured from skin lesions. According to its phenotypic and biochemical characteristics we could predict only that it might be M. szulgai, M. scrofulaceum or M. gordonae. Polymerase chain reaction amplification of the hsp65 gene and subsequent restriction fragment length polymorphism analysis on the isolated strain showed that its restriction pattern differed from both M. scrofulaceum and other scotochromogenic species. Ninety-nine per cent similarity was detected between the isolated strain and M. gordonae by sequencing of the hsp65 gene. This result suggests that the isolated strain may be either a slow-growing scotochromogenic Mycobacterium most resembling M. gordonae or a novel mycobacterial species. PMID:14511003

Li, X J; Wu, Q X; Zeng, X S

2003-09-01

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Assessment of partial sequencing of the 65-kilodalton heat shock protein gene (hsp65) for routine identification of Mycobacterium species isolated from clinical sources.  

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We assessed the ability of an in-house database, consisting of 111 hsp65 sequences from putative and valid Mycobacterium species or described groups, to identify 689 mycobacterial clinical isolates from 35 species or groups. A preliminary assessment indicated that hsp65 sequencing confirmed the identification of 79.4% of the isolates from the 32 species examined, including all Mycobacterium tuberculosis complex isolates, all isolates from 13 other species, and 95.6% of all M. avium-M. intracellulare complex isolates. Identification discrepancies were most frequently encountered with isolates submitted as M. chelonae, M. fortuitum, M. gordonae, M. scrofulaceum, and M. terrae. Reexamination of isolates with discrepant identifications confirmed that hsp65 identifications were correct in a further 40 isolates. This brought the overall agreement between hsp65 sequencing and the other identification methods to 85.2%. The remaining 102 isolates had sequence matches below our acceptance criterion, had nondifferential sequence matches between two or more species, were identified by 16S rRNA sequencing as a putative taxonomic group not contained in our database, or were identified by hsp65 and 16S rRNA gene sequencing as a species not in our biochemical test database or had conflicting identifications. Therefore, to incorporate the unconfirmed isolates it was necessary to create 29 additional entries in our hsp65 identification database: 18 associated with valid species, 7 indicating unique sequences not associated with valid or putative species or groups, and 4 associated with unique, but currently described taxonomic groups. Confidence in the hsp65 sequence identification of a clinical isolate is best when sequence matches of 100% occur, but our data indicate that correct identifications can be confidently made when unambiguous matches exceeding 97% occur, but are dependent on the completeness of the database. Our study indicates that for hsp65 sequencing to be an effective means for identifying mycobacteria a comprehensive database must be constructed. hsp65 sequencing has the advantage of being more rapid and less expensive than biochemical test panels, uses a single set of reagents to identify both rapid- and slow-growing mycobacteria, and can provide a more definitive identification. PMID:15243051

McNabb, Alan; Eisler, Diane; Adie, Kathy; Amos, Marie; Rodrigues, Mabel; Stephens, Gwen; Black, William A; Isaac-Renton, Judith

2004-07-01

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The genomic heterogeneity among Mycobacterium terrae complex displayed by sequencing of 16S rRNA and hsp 65 genes.  

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The species identification within Mycobacterium terrae complex has been known to be very difficult. In this study, the genomic diversity of M. terrae complex with eighteen clinical isolates, which were initially identified as M. terrae complex by phenotypic method, was investigated, including that of three type strains (M. terrae, M. nonchromogenicum, and M. triviale ). 16S rRNA and 65-kDa heat shock protein (hsp 65) gene sequences of mycobacteria were determined and aligned with eleven other references for the comparison using similarity search against the GenBank and Ribosomal Database Project II (RDP) databases. 16S rRNA and hsp 65 genes of M. terrae complex showed genomic heterogeneity. Amongst the eighteen clinical isolates, nine were identified as M. nonchromogenicum, eight as M. terrae, one as M. mucogenicum with the molecular characteristic of rapid growth. M. nonchromogenicum could be subdivided into three subgroups, while M. terrae could be subdivided into two subgroups using a 5 bp criterion (>1% difference). Seven isolates in two subgroups of M. nonchromogenicum were Mycobacterium sp. strain MCRO 6, which was closely related to M. nonchromogenicum. The hsp 65 gene could not differentiate one M. nonchromogenicum from M. avium or one M. terrae from M. intracellulare. The nucleotide sequence analysis of 16S rRNA and hsp 65 genes was shown to be useful in identifying the M. terrae complex, but hsp 65 was less discriminating than 16S rRNA. PMID:14978332

Lee, Chang Kyu; Gi, Hyun Mi; Cho, Yunjung; Kim, Young Kee; Lee, Kap No; Song, Ki-Joon; Song, Jin-Won; Park, Kwang Sook; Park, Eun Mi; Lee, Hyeyoung; Bai, Gill-Han

2004-01-01

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Assessment of Partial Sequencing of the 65-Kilodalton Heat Shock Protein Gene (hsp65) for Routine Identification of Mycobacterium Species Isolated from Clinical Sources  

OpenAIRE

We assessed the ability of an in-house database, consisting of 111 hsp65 sequences from putative and valid Mycobacterium species or described groups, to identify 689 mycobacterial clinical isolates from 35 species or groups. A preliminary assessment indicated that hsp65 sequencing confirmed the identification of 79.4% of the isolates from the 32 species examined, including all Mycobacterium tuberculosis complex isolates, all isolates from 13 other species, and 95.6% of all M. avium-M. intrace...

Mcnabb, Alan; Eisler, Diane; Adie, Kathy; Amos, Marie; Rodrigues, Mabel; Stephens, Gwen; Black, William A.; Isaac-renton, Judith

2004-01-01

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Análise de restrição enzimática do gene hsp65 de isolados clínicos de pacientes com suspeita de tuberculose pulmonar em Teresina, Piauí / Restriction enzyme analysis of the hsp65 gene in clinical isolates from patients suspected of having pulmonary tuberculosis in Teresina, Brazil  

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Full Text Available SciELO Brazil | Language: Portuguese Abstract in portuguese OBJETIVO: Identificar as espécies de micobactérias encontradas no escarro de pacientes com suspeita de tuberculose pulmonar e analisar o impacto dessas identificações na abordagem terapêutica. MÉTODOS: Foram avaliados 106 pacientes com suspeita de tuberculose pulmonar encaminhados para o serviço de [...] pneumologia de um hospital público em Teresina, Piauí. Espécimes de escarro matinal foram avaliados quanto à presença de micobactérias por baciloscopia e cultura. Foram utilizadas PCR e análise de restrição enzimática do gene hsp65 (PRA-hsp65) para a identificação das cepas de micobactérias isoladas em cultura. RESULTADOS: Foram analisadas 206 amostras de escarro. A idade dos pacientes variou de 15 a 87 anos, sendo 67% do gênero masculino. Tosse ocorreu em 100% dos casos. O padrão radiográfico predominante foi de lesão moderada, observada em 70%. A positividade no esfregaço foi de 76%, e isolamento em cultura ocorreu em 91% das culturas executadas. Testes tradicionais identificaram micobactérias não tuberculosas (MNT) em 9% dos isolados. O método PRA-hsp65 confirmou esses dados, mostrando sete padrões de bandas capazes de identificar as espécies de MNT isoladas: Mycobacterium kansasii; M. abscessus 1; M. abscessus 2; M. smegmatis; M. flavescens 1; M. gordonae 5 e M. gordonae 7. Todos os pacientes com MNT tinham mais de 60 anos, e observaram-se bronquiectasias em 88% das radiografias. Houve dois casos de reinfecção, identificados inicialmente como infecção por M. abscessus e M. kansasii. CONCLUSÕES: As MNT causam infecção pulmonar em pacientes imunocompetentes, e a identificação das MNT é importante para estabelecer o diagnóstico correto e a decisão terapêutica mais adequada. O método PRA-hsp65 é útil para identificar espécies de MNT e pode ser implantado em laboratórios de biologia molecular não especializados em micobactérias. Abstract in english OBJECTIVE: To identify mycobacterial species in the sputum of patients suspected of having pulmonary tuberculosis and to determine the impact that the acquisition of this knowledge has on the therapeutic approach. METHODS: We evaluated 106 patients suspected of having pulmonary tuberculosis and refe [...] rred to the pulmonology department of a public hospital in the city of Teresina, Brazil. Morning sputum specimens were evaluated for the presence of mycobacteria by sputum smear microscopy and culture. We used PCR and restriction enzyme analysis of the hsp65 gene (PRA-hsp65) to identify the strains of mycobacteria isolated in culture. RESULTS: A total of 206 sputum samples were analyzed. Patient ages ranged from 15 to 87 years, and 67% were male. There was cough in 100% of the cases. The predominant radiographic pattern was moderate disease, observed in 70%. Smear positivity was 76%, and isolation in culture occurred in 91% of the cultures. Traditional tests identified nontuberculous mycobacteria (NTM) in 9% of the isolates. The PRA-hsp65 method confirmed these data, showing seven band patterns that were able to identify the isolated species of NTM: Mycobacterium kansasii; M. abscessus 1; M. abscessus 2; M. smegmatis; M. flavescens 1; M. gordonae 5; and M. gordonae 7. All of the patients with NTM were over 60 years of age, and bronchiectasis was seen in 88% of the X-rays. There were two cases of reinfection, initially attributed to M. abscessus and M. kansasii. CONCLUSIONS: In immunocompetent patients, NTM can infect the lungs. It is important to identify the specific NTM in order to establish the correct diagnosis and choose the most appropriate therapeutic regimen. The PRA-hsp65 method is useful in identifying NTM species and can be implemented in molecular biology laboratories that do not specialize in the identification of mycobacteria.

Maria das Graças Motta e, Bona; Maria José Soares, Leal; Liline Maria Soares, Martins; Raimundo Nonato da, Silva; José Adail Fonseca de, Castro; Semiramis Jamil Hadad do, Monte.

2011-10-01

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Análise de restrição enzimática do gene hsp65 de isolados clínicos de pacientes com suspeita de tuberculose pulmonar em Teresina, Piauí Restriction enzyme analysis of the hsp65 gene in clinical isolates from patients suspected of having pulmonary tuberculosis in Teresina, Brazil  

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Full Text Available OBJETIVO: Identificar as espécies de micobactérias encontradas no escarro de pacientes com suspeita de tuberculose pulmonar e analisar o impacto dessas identificações na abordagem terapêutica. MÉTODOS: Foram avaliados 106 pacientes com suspeita de tuberculose pulmonar encaminhados para o serviço de pneumologia de um hospital público em Teresina, Piauí. Espécimes de escarro matinal foram avaliados quanto à presença de micobactérias por baciloscopia e cultura. Foram utilizadas PCR e análise de restrição enzimática do gene hsp65 (PRA-hsp65 para a identificação das cepas de micobactérias isoladas em cultura. RESULTADOS: Foram analisadas 206 amostras de escarro. A idade dos pacientes variou de 15 a 87 anos, sendo 67% do gênero masculino. Tosse ocorreu em 100% dos casos. O padrão radiográfico predominante foi de lesão moderada, observada em 70%. A positividade no esfregaço foi de 76%, e isolamento em cultura ocorreu em 91% das culturas executadas. Testes tradicionais identificaram micobactérias não tuberculosas (MNT em 9% dos isolados. O método PRA-hsp65 confirmou esses dados, mostrando sete padrões de bandas capazes de identificar as espécies de MNT isoladas: Mycobacterium kansasii; M. abscessus 1; M. abscessus 2; M. smegmatis; M. flavescens 1; M. gordonae 5 e M. gordonae 7. Todos os pacientes com MNT tinham mais de 60 anos, e observaram-se bronquiectasias em 88% das radiografias. Houve dois casos de reinfecção, identificados inicialmente como infecção por M. abscessus e M. kansasii. CONCLUSÕES: As MNT causam infecção pulmonar em pacientes imunocompetentes, e a identificação das MNT é importante para estabelecer o diagnóstico correto e a decisão terapêutica mais adequada. O método PRA-hsp65 é útil para identificar espécies de MNT e pode ser implantado em laboratórios de biologia molecular não especializados em micobactérias.OBJECTIVE: To identify mycobacterial species in the sputum of patients suspected of having pulmonary tuberculosis and to determine the impact that the acquisition of this knowledge has on the therapeutic approach. METHODS: We evaluated 106 patients suspected of having pulmonary tuberculosis and referred to the pulmonology department of a public hospital in the city of Teresina, Brazil. Morning sputum specimens were evaluated for the presence of mycobacteria by sputum smear microscopy and culture. We used PCR and restriction enzyme analysis of the hsp65 gene (PRA-hsp65 to identify the strains of mycobacteria isolated in culture. RESULTS: A total of 206 sputum samples were analyzed. Patient ages ranged from 15 to 87 years, and 67% were male. There was cough in 100% of the cases. The predominant radiographic pattern was moderate disease, observed in 70%. Smear positivity was 76%, and isolation in culture occurred in 91% of the cultures. Traditional tests identified nontuberculous mycobacteria (NTM in 9% of the isolates. The PRA-hsp65 method confirmed these data, showing seven band patterns that were able to identify the isolated species of NTM: Mycobacterium kansasii; M. abscessus 1; M. abscessus 2; M. smegmatis; M. flavescens 1; M. gordonae 5; and M. gordonae 7. All of the patients with NTM were over 60 years of age, and bronchiectasis was seen in 88% of the X-rays. There were two cases of reinfection, initially attributed to M. abscessus and M. kansasii. CONCLUSIONS: In immunocompetent patients, NTM can infect the lungs. It is important to identify the specific NTM in order to establish the correct diagnosis and choose the most appropriate therapeutic regimen. The PRA-hsp65 method is useful in identifying NTM species and can be implemented in molecular biology laboratories that do not specialize in the identification of mycobacteria.

Maria das Graças Motta e Bona

2011-10-01

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First Report of Disseminated Mycobacterium Skin Infections in Two Liver Transplant Recipients and Rapid Diagnosis by hsp65 Gene Sequencing ?  

OpenAIRE

We present here the first report of disseminated skin Mycobacterium infections in two liver transplant recipients, in which hsp65 gene sequencing was used for rapid species identification. Both patients had hepatitis B virus-related cirrhosis and diabetes mellitus and presented with progressive generalized, nodular skin lesions. In one patient, a 50-year-old woman who had frequent contact with marine fish, an acid-fast bacillus was isolated from skin biopsy tissue after 2 months of culture. W...

Lau, Susanna K. P.; Curreem, Shirly O. T.; Ngan, Antonio H. Y.; Yeung, Chi-keung; Yuen, Kwok-yung; Woo, Patrick C. Y.

2011-01-01

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hsp65 PCR-restriction enzyme analysis (PRA) for identification of mycobacteria in the clinical laboratory / PCR e análise de padrões de restrição do gene hsp65 (PRA) para identificação de micobactérias no laboratório clínico  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: English Abstract in portuguese Mais de 70 espécies de micobactérias já foram definidas e algumas delas podem causar enfermidade em humanos, especialmente em pacientes imunocomprometidos. A identificação de espécie, na maioria dos laboratórios clínicos, se baseia em características fenotípicas e testes bioquímicos e resultados def [...] initivos só são obtidos após duas a quatro semanas. Métodos rápidos de identificação reduzem o tempo necessário para o diagnóstico e podem antecipar a instituição do tratamento específico, aumentando as chances de sucesso. A análise de padrões de restrição do gene hsp65 amplificado por PCR (PRA) foi utilizada como método rápido de identificação em 103 isolamentos clínicos. Os padrões de bandas foram interpretados por comparação com tabelas publicadas e padrões disponíveis em um site de Internet (http://www.hospvd.ch:8005). Resultados concordantes de PRA e identificação bioquímica foram obtidos em 76 de 83 isolamentos (91,5%). Os resultados de 20 isolamentos não puderam ser comparados porque a identificação fenotípica ou por PRA foi inconclusiva. Os resultados deste trabalho mostram que PRA pode ser útil para identificação de rotina de micobactérias por ser um método acurado, rápido e mais econômico do que a identificação convencional. Abstract in english More than 70 species of mycobacteria have been defined, and some can cause disease in humans, especially in immunocompromised patients. Species identification in most clinical laboratories is based on phenotypic characteristics and biochemical tests and final results are obtained only after two to f [...] our weeks. Quick identification methods, by reducing time for diagnosis, could expedite institution of specific treatment, increasing chances of success. PCR restriction-enzyme analysis (PRA) of the hsp65 gene was used as a rapid method for identification of 103 clinical isolates. Band patterns were interpreted by comparison with published tables and patterns available at an Internet site (http://www.hospvd.ch:8005). Concordant results of PRA and biochemical identification were obtained in 76 out of 83 isolates (91.5%). Results from 20 isolates could not be compared due to inconclusive PRA or biochemical identification. The results of this work showed that PRA could improve identification of mycobacteria in a routine setting because it is accurate, fast, and cheaper than conventional phenotypic identification.

Carolina Feher da, SILVA; Suely Yoko Mizuka, UEKI; Débora de Cássia Pires, GEIGER; Sylvia Cardoso, LEÃO.

2001-02-01

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The hsp65 gene patterns of less common Mycobacterium and Nocardia spp. by polymerase chain reaction-restriction fragment length polymorphism analysis with capillary electrophoresis.  

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To rapidly identify Mycobacterium and Nocardia spp. without costly probes, we had implemented capillary electrophoresis (CE) in polymerase chain reaction (PCR)-restriction fragment length polymorphism (RFLP) analysis to analyze their 65-kDa heat shock protein (hsp65) gene. The PCR-RFLP analysis with CE (PRACE) involved only one restriction enzyme, HaeIII, and a single electrophoretic separation less than 10 min. Full-range (10-200 bp) RFLP patterns of 12 less common Mycobacterium and 7 Nocardia spp. were investigated. A good agreement was observed between the sizes of restriction fragments resolved by CE and the real sizes deduced from sequence analysis. Including hsp65 gene patterns of 12 Mycobacterium spp. published earlier, differentiation was distinct among 24 Mycobacterium and 7 Nocardia spp. Some closely related species exhibiting similar biochemical characteristics could be well discriminated by an extra HaeIII digestion site. Thus, PRACE offers a nonprobe alternative for rapid identification of various cultured Mycobacterium and Nocardia to the species level. PMID:17382507

Chang, Po-Ling; Hsieh, Wen-Shyang; Chiang, Chia-Lien; Tuohy, Marion J; Hall, Gerri S; Procop, Gary W; Chang, Huan-Tsung; Ho, Hsin-Tsung

2007-07-01

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DETECÇÃO DO COMPLEXO Mycobacterium tuberculosis NO LEITE PELA REAÇÃO EM CADEIA DA POLIMERASE SEGUIDA DE ANÁLISE DE RESTRIÇÃO DO FRAGMENTO AMPLIFICADO (PRA DETECTION OF Mycobacterium tuberculosis COMPLEX BY PCR-RESTRICTION FRAGMENT LENGTH POLYMORFISM ANALYSIS OF THE HSP65 GENE  

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Full Text Available Mycobacterium bovis é membro do complexo Mycobacterium tuberculosis (MTBC, grupo este composto por espécies com grande homologia genética. É o agente etiológico da tuberculose bovina, importante zoonose transmissível ao homem, principalmente através da inalação do bacilo e/ou pelo consumo de leite e derivados não-pasteurizados provenientes de vacas tuberculosas. O objetivo deste estudo foi padronizar a identificação de micobactérias do complexo M. tuberculosis presentes no leite, por metodologia molecular. Fez-se a extração de DNA diretamente do leite contaminado e realizou-se a identificação molecular pela reação em cadeia da polimerase seguida de análise de restrição do fragmento amplificado (PRA. Utilizaram-se inhagens de referência e leite cru artificialmente contaminado com M. bovis IP. Um fragmento de 441pb do gene hsp65 foi amplificado, tratado com BstEII e HaeIII e empregou-se o perfil de restrição enzimática obtido para identificar o complexo M. tuberculosis no leite. Com a PRA foi possível detectar com especificidade e sensibilidade a presença de M. bovis em até 10 UFC/mL de leite. A metodologia padronizada poderá auxiliar os métodos microbiológicos e bioquímicos tradicionalmente usados na identificação do bacilo em alimentos suspeitos de contaminação, como, por exemplo, o leite proveniente de animais suspeitos de infecção por M. bovis.

Palavras-chaves: Análise de perfil de restrição enzimática (PRA, complexo Mycobacterium tuberculosis, leite, Mycobacterium bovis, limite de detecção (PCR. Mycobacterium bovis is a member of the M. tuberculosis complex, a group composed by species with high genetic homology. The pathogen is the etiological agent of bovine tuberculosis, an important zoonosis that is mainly transmitted by inhalation of infectious droplet nuclei or by ingestion of milk and crude milk derivative products from tuberculosis cows. The definitive identification of M. bovis, up to species level, is time consuming and difficult. In this work, the objective was to standardize a polymerase chain reaction followed by an enzyme restriction analysis in order to identify the M. tuberculosis complex in milk, without a microbiological isolation step. Reference strains and raw milk seeded with M. Bovis, were used as the starting material.  A 441pb fragment of the hsp65 gene was amplified and digested by two restriction enzymes BstEII and HaeIII. The obtained profile was used to identify the M. tuberculosis complex in milk. The minimum limit of detection of M. bovis in milk was 10CFU/mL. PRA methodology proved to be a specific and sensible method. It can be used to assist the microbiological and biochemical methods commonly used to identifying the bacilli in clinical samples, as milk 

Key word: Detection limit (PRA, Mycobacterium tuberculosis complex, milk Mycobacterium bovis, Restriction Enzyme Analysis (PCR,

Joab Trajano Silva

2008-12-01

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Th1 polarized response induced by intramuscular DNA-HSP65 immunization is preserved in experimental atherosclerosis  

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Full Text Available We previously reported that a DNA vaccine constructed with the heat shock protein (HSP65 gene from Mycobacterium leprae (DNA-HSP65 was protective and also therapeutic in experimental tuberculosis. By the intramuscular route, this vaccine elicited a predominant Th1 response that was consistent with its protective efficacy against tuberculosis. It has been suggested that the immune response to Hsp60/65 may be the link between exposure to microorganisms and increased cardiovascular risk. Additionally, the high cholesterol levels found in atherosclerosis could modulate host immunity. In this context, we evaluated if an atherogenic diet could modulate the immune response induced by the DNA-HSP65 vaccine. C57BL/6 mice (4-6 animals per group were initially submitted to a protocol of atherosclerosis induction and then immunized by the intramuscular or intradermal route with 4 doses of 100 µg DNA-HSP65. On day 150 (15 days after the last immunization, the animals were sacrificed and antibodies and cytokines were determined. Vaccination by the intramuscular route induced high levels of anti-Hsp65 IgG2a antibodies, but not anti-Hsp65 IgG1 antibodies and a significant production of IL-6, IFN-g and IL-10, but not IL-5, indicating a Th1 profile. Immunization by the intradermal route triggered a mixed pattern (Th1/Th2 characterized by synthesis of anti-Hsp65 IgG2a and IgG1 antibodies and production of high levels of IL-5, IL-6, IL-10, and IFN-g. These results indicate that experimentally induced atherosclerosis did not affect the ability of DNA-HSP65 to induce a predominant Th1 response that is potentially protective against tuberculosis.

D.M. Fonseca

2007-11-01

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Th1 polarized response induced by intramuscular DNA-HSP65 immunization is preserved in experimental atherosclerosis  

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Full Text Available SciELO Brazil | Language: English Abstract in english We previously reported that a DNA vaccine constructed with the heat shock protein (HSP65) gene from Mycobacterium leprae (DNA-HSP65) was protective and also therapeutic in experimental tuberculosis. By the intramuscular route, this vaccine elicited a predominant Th1 response that was consistent with [...] its protective efficacy against tuberculosis. It has been suggested that the immune response to Hsp60/65 may be the link between exposure to microorganisms and increased cardiovascular risk. Additionally, the high cholesterol levels found in atherosclerosis could modulate host immunity. In this context, we evaluated if an atherogenic diet could modulate the immune response induced by the DNA-HSP65 vaccine. C57BL/6 mice (4-6 animals per group) were initially submitted to a protocol of atherosclerosis induction and then immunized by the intramuscular or intradermal route with 4 doses of 100 µg DNA-HSP65. On day 150 (15 days after the last immunization), the animals were sacrificed and antibodies and cytokines were determined. Vaccination by the intramuscular route induced high levels of anti-Hsp65 IgG2a antibodies, but not anti-Hsp65 IgG1 antibodies and a significant production of IL-6, IFN-g and IL-10, but not IL-5, indicating a Th1 profile. Immunization by the intradermal route triggered a mixed pattern (Th1/Th2) characterized by synthesis of anti-Hsp65 IgG2a and IgG1 antibodies and production of high levels of IL-5, IL-6, IL-10, and IFN-g. These results indicate that experimentally induced atherosclerosis did not affect the ability of DNA-HSP65 to induce a predominant Th1 response that is potentially protective against tuberculosis.

D.M., Fonseca; V.L.D., Bonato; C.L., Silva; A., Sartori.

1495-15-01

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Recurrent nontuberculous mycobacterial endophthalmitis: a diagnostic conundrum  

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Full Text Available Nandini Venkateswaran,1 Gabrielle Yeaney,2 Mina Chung,3,4 Holly B Hindman3,41University of Rochester School of Medicine and Dentistry, University of Rochester, 2Department of Pathology and Laboratory Medicine, 3Flaum Eye Institute, 4Center for Visual Science, University of Rochester School of Medicine and Dentistry, Rochester, NY, USAObjective: To report a case of recurrent nontuberculous mycobacterial endophthalmitis in the context of neurotrophic keratopathy secondary to herpes zoster ophthalmicus that had an atypical presentation and complex course, and highlights the challenges of causative organism identification and therapeutic interventions in this condition.Methods: A retrospective chart review was conducted to determine the visual outcomes of the patient.Results: A 68-year-old pseudophakic male with long-standing neurotrophic keratopathy and perforated descemetocele managed with cyanoacrylate glue and a contact bandage lens in the left eye, began experiencing recurrent episodes of endophthalmitis after undergoing a penetrating keratoplasty. Several therapeutic procedures including an anterior chamber washout, two pars plana vitrectomies, explantation of the posterior chamber intraocular lens and capsular bag, and multiple intravitreal antimicrobial injections, were performed to which he has ultimately responded favorably, with no signs of infection to date and stable visual acuity. The causative organism of his recurrent infections was initially identified as Mycobacterium abscessus through biochemical testing and 16S ribosomal ribonucleic acid gene sequencing; however, repeat polymerase chain reaction (PCR and sequencing of the 65 kDa heat shock protein (hsp65 gene for experimental purposes confirmed the accurate identification of the organism to be Mycobacterium chelonae. Given the greater reliability of PCR and sequencing of the hsp65 gene over traditional biochemical tests and culture techniques, M. chelonae was likely the infectious agent all along, and the organism was originally misidentified on the basis of less accurate tests.Conclusion: Recurrent atypical mycobacterial endophthalmitis requires expedient identification and management to prevent poor visual outcomes. Standard biochemical testing can identify the causative organism but is limited by the inability to distinguish between nontuberculous species reliably. We recommend the use of PCR in conjunction with sequencing of the hsp65 gene for reliable differentiation of M. chelonae and M. abscessus in atypical mycobacterial ocular infections. Minimum inhibitory concentration antibiotic susceptibility tests on cultured strains are the best guide to antibiotic selection, given the rapidly rising resistance to antimicrobials in atypical mycobacterial species.Keywords: atypical mycobacteria, herpes zoster ophthalmicus, hsp65, Mycobacterium chelonae, neurotrophic keratopathy, visual outcome

Venkateswaran N

2014-05-01

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Comparative evaluation of polymerase chain reaction and restriction enzyme analysis: two amplified targets, hsp65 and rpoB, for identification of cultured mycobacteria.  

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The increasing incidence of tuberculosis and other mycobacterial infections due to AIDS epidemic resulted in the need of rapid and accurate identification of isolated mycobacteria. The correct identification result leads to the selection of an appropriate therapeutic regimen. Polymerase chain reaction and restriction enzyme analysis (PCR-REA) has been developed since 1992 and used as the rapid method for identifying mycobacteria. Several genes or sequences have been used as an amplified target for PCR-REA. The present study aims to evaluate the potential use of PCR-REA of gene-encoding heat shock protein 65 kDa (hsp65) and beta-subunit RNA polymerase (rpoB) for the identification of mycobacteria compared with conventional biochemical identification. Two hundreds clinical isolates, consisting of 50 isolates of Mycobacterium tuberculosis and 150 isolates of nontuberculous mycobacteria (NTM), were submitted for identification using PCR-REA and biochemical method. The results demonstrated that PCR-REA identified 188 isolates of both M. tuberculosis and NTM concordantly with biochemical identification. Discordant identification results obtained from 12 isolates, comprised of 8 M. scrofulaceum, 1 M. avium complex, 1 M. malmoense, 1 M. terrae complex, and 1 M. chelonae/abscessus. Overall, the concordant percentage of results obtained from PCR-REA compared with biochemical method was 100%, 98.8%, and 83.3% for M. tuberculosis complex, rapidly growing, and slowly growing mycobacteria, respectively, and the results of hsp65 PCR-REA was in agreement with those obtained from rpoB PCR-REA. From this study, PCR-REA appears to be a simple, rapid, and reliable method for identifying mycobacteria in a routine microbiology laboratory. PMID:15766601

Cheunoy, Wattana; Prammananan, Therdsak; Chaiprasert, Angkana; Foongladda, Suporn

2005-03-01

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Ub Combination Enhanced Cellular Immune Response Elicited by HSP65 DNA Vaccine against Mycobacterium tuberculosis  

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 This study observed the immune response induced by a HSP65 DNA vaccine fused with UbGR against Mycobacterium tuberculosis. BALB/c mice were inoculated with HSP65 DNA vaccine, UbGR-fused HSP65 DNA vaccine (Ub-GR-HSP65) and blank vector respectively. HSP65 DNA vaccine elicited a Thl-polarized immune response. The Thl-type cytokine (IFN-?) and proliferative T cell responses from spleen were improved significantly in Ub...

Qingmin Wang; Chengxiang Lei; Qiuhong Liu

2013-01-01

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Influência do biofármaco DNA-hsp65 na lesão pulmonar induzida por bleomicina / Influence of a DNA-hsp65 vaccine on bleomycin-induced lung injury  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: Portuguese Abstract in portuguese OBJETIVO: Avaliar a influência do biofármaco DNA-hsp65 em um modelo de distúrbio fibrosante pulmonar experimental. MÉTODOS: Foram estudados 120 camundongos machos C57BL/6, divididos em quatro grupos: grupo SS, animais tratados com salina (placebo) e injetados com salina intratraqueal (IT); grupo SB, [...] tratados com salina (placebo) e injetados com bleomicina IT; grupo PB, tratados com plasmídeo, sem gene bacteriano, e injetados com bleomicina IT; e grupo BB, tratados com DNA-hsp65 e injetados com bleomicina IT. A bleomicina foi injetada 15 dias após a última imunização, e os animais sacrificados seis semanas após o uso da droga IT. O pulmão esquerdo retirado foi utilizado para análise morfológica, e o pulmão direito para dosagens de hidroxiprolina. RESULTADOS: A proporção de camundongos que apresentaram morte não-programada depois de 48 h da injeção IT foi maior no grupo SB em comparação ao grupo SS (57,7% vs. 11,1%). A área percentual média de interstício septal foi maior nos grupos SB e PB (53,1 ± 8,6% e 53,6 ± 9,3%, respectivamente) em comparação aos grupos SS e BB (32,9 ± 2,7% e 34,3 ± 6,1%, respectivamente). Os grupos SB, PB e BB mostraram aumentos nos valores médios da área de interstício septal corada por picrosirius em comparação ao grupo SS (SS: 2,0 ± 1,4%; SB: 8,2 ± 4,9%; PB: 7,2 ± 4,2%; e BB:6,6±4,1%).O conteúdo pulmonar de hidroxiprolina no grupo SS foi inferior ao dos demais grupos (SS: 104,9 ± 20,9 pg/pulmão; SB: 160,4 ±47,8 pg/pulmão; PB:170,0 ± 72,0 pg/pulmão; e BB: 162,5 ± 39,7 pg/pulmão). CONCLUSÕES: A imunização com o biofármaco DNA-hsp65 interferiu na deposição de matriz não-colágena em um modelo de lesão pulmonar induzida por bleomicina. Abstract in english OBJECTIVE: To evaluate the effects of immunization with a DNA-hsp65 vaccine in an experimental model of pulmonary fibrosis. METHODS: A total of 120 male C57BL/6 mice were distributed into four groups: SS, injected with saline (placebo) and then receiving intratracheal (IT) instillation of saline; SB [...] , injected with saline (placebo) and then receiving IT instillation of bleomycin; PB, treated with plasmid only, without bacterial genome, and then receiving IT instillation of bleomycin; and BB, treated with the vaccine and then receiving IT instillation of bleomycin. Bleomycin was instilled 15 days after the last immunization, and the animals were killed six weeks thereafter. The left and right lungs were removed, the former for morphological analysis and the latter for hydroxyproline measurements. RESULTS: The proportion of deaths within the first 48 h after the IT instillation (deaths attributed to the surgical procedure) was higher in the SB group than in the SS group (57.7% vs. 11.1%). The mean area of pulmonary interstitial septa was greater in the SB and PB groups (53.1 ± 8.6% and 53.6±9.3%, respectively) than in the SS and BB groups (32.9 ± 2.7% and 34.3 ± 6.1%, respectively). The mean area of interstitial septa stained by picrosirius was greater in the SB, PB and BB groups than in the SS group (8.2 ± 4.9%, 7.2 ± 4.2% and 6.6 ± 4.1%, respectively, vs. 2.0±1.4%). The total hydroxyproline content in the lung was significantly lower in the SS group (104.9 ± 20.9 pg/lung) than in the other groups (SB: 160.4 ± 47.8 pg/lung; PB: 170.0 ± 72.0 pg/lung; and BB: 162.5 ± 39.7 pg/lung). CONCLUSIONS: Immunization with the DNA-hsp65 vaccine reduced the deposition of noncollagen matrix in a model of bleomycin-induced lung lesion.

Adriana Ignacio de, Padua; Célio Lopes, Silva; Simone Gusmão, Ramos; Lúcia Helena, Faccioli; José Antônio Baddini, Martinez.

2008-11-01

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Novel Allelic Variants of Mycobacteria Isolated in Brazil as Determined by PCR-Restriction Enzyme Analysis of hsp65  

OpenAIRE

Human isolates of Mycobacterium collected in 16 different states of Brazil were submitted to PCR-restriction analysis (PRA) of a 439-bp fragment of the hsp65 gene with HaeIII and BstEII. Fourteen allelic variants not described in clinical isolates so far were observed among 36 (10%) of 356 Brazilian strains, including a new pattern for Mycobacterium scrofulaceum, M. intracellulare, and M. flavescens, two new patterns for M. fortuitum, three new patterns each for M. gordonae and M. terrae, and...

Da Silva Rocha, A.; Werneck Barreto, A. M.; Dias Campos, C. E.; Villas-bo?as Da Silva, M.; Fonseca, L.; Saad, M. H.; Degrave, W. M.; Suffys, P. N.

2002-01-01

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Mycobacterium tuberculosis Chaperonin 60.1 Is a More Potent Cytokine Stimulator than Chaperonin 60.2 (Hsp 65) and Contains a CD14-Binding Domain  

OpenAIRE

Much attention has focused on the Mycobacterium tuberculosis molecular chaperone chaperonin (Cpn) 60.2 (Hsp 65) in the pathology of tuberculosis because of its immunogenicity and ability to directly activate human monocytes and vascular endothelial cells. However, M. tuberculosis is one of a small group of bacteria that contain multiple genes encoding Cpn 60 proteins. We have now cloned and expressed both M. tuberculosis proteins and report that the novel chaperonin 60, Cpn 60.1, is a more po...

Lewthwaite, Jo C.; Coates, Anthony R. M.; Tormay, Peter; Singh, Mahavir; Mascagni, Paolo; Poole, Stephen; Roberts, Michael; Sharp, Lindsay; Henderson, Brian

2001-01-01

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Antigen-presenting cells transfected with Hsp65 messenger RNA fail to treat experimental tuberculosis  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: English Abstract in english In the last several years, the use of dendritic cells has been studied as a therapeutic strategy against tumors. Dendritic cells can be pulsed with peptides or full-length protein, or they can be transfected with DNA or RNA. However, comparative studies suggest that transfecting dendritic cells with [...] messenger RNA (mRNA) is superior to other antigen-loading techniques in generating immunocompetent dendritic cells. In the present study, we evaluated a new therapeutic strategy to fight tuberculosis using dendritic cells and macrophages transfected with Hsp65 mRNA. First, we demonstrated that antigen-presenting cells transfected with Hsp65 mRNA exhibit a higher level of expression of co-stimulatory molecules, suggesting that Hsp65 mRNA has immunostimulatory properties. We also demonstrated that spleen cells obtained from animals immunized with mock and Hsp65 mRNA-transfected dendritic cells were able to generate a mixed Th1/Th2 response with production not only of IFN-? but also of IL-5 and IL-10. In contrast, cells recovered from mice immunized with Hsp65 mRNA-transfected macrophages were able to produce only IL-5. When mice were infected with Mycobacterium tuberculosis and treated with antigen-presenting cells transfected with Hsp65 mRNA (therapeutic immunization), we did not detect any decrease in the lung bacterial load or any preservation of the lung parenchyma, indicating the inability of transfected cells to confer curative effects against tuberculosis. In spite of the lack of therapeutic efficacy, this study reports for the first time the use of antigen-presenting cells transfected with mRNA in experimental tuberculosis.

C.D., Rocha; A.P.F., Trombone; J.C.C., Lorenzi; L.P., Almeida; A.F., Gembre; E., Padilha; S.G., Ramos; C.L., Silva; A.A.M., Coelho-Castelo.

1183-11-01

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Novel allelic variants of Mycobacteria isolated in Brazil as determined by PCR-restriction enzyme analysis of hsp65.  

Science.gov (United States)

Human isolates of Mycobacterium collected in 16 different states of Brazil were submitted to PCR-restriction analysis (PRA) of a 439-bp fragment of the hsp65 gene with HaeIII and BstEII. Fourteen allelic variants not described in clinical isolates so far were observed among 36 (10%) of 356 Brazilian strains, including a new pattern for Mycobacterium scrofulaceum, M. intracellulare, and M. flavescens, two new patterns for M. fortuitum, three new patterns each for M. gordonae and M. terrae, and one new pattern for M. avium complex-like strains. Two unidentified strains each also presented a new pattern, strongly suggesting that Mycobacterium genotypes are distributed biogeographically. The PRA procedure was also performed with 43 reference isolates belonging to 34 species, adding a further six new patterns to the identification algorithm. A database containing the normalized restriction patterns of both enzymes was constructed. Patterns available on the Internet can be introduced into this database, which will make possible the comparison of genotypes from isolates from different parts of the world. PMID:12409396

da Silva Rocha, A; Werneck Barreto, A M; Dias Campos, C E; Villas-Bôas da Silva, M; Fonseca, L; Saad, M H; Degrave, W M; Suffys, P N

2002-11-01

21

Mycobacterial FurA is a negative regulator of catalase-peroxidase gene katG.  

Science.gov (United States)

In several bacteria, the catalase-peroxidase gene katG is under positive control by oxyR, a transcriptional regulator of the peroxide stress response. The Mycobacterium tuberculosis genome also contains sequences corresponding to oxyR, but this gene has been inactivated in the tubercle bacillus because of the presence of multiple mutations and deletions. Thus, M. tuberculosis katG and possibly other parts of the oxidative stress response in this organism are either not regulated or are controlled by a factor different from OxyR. The mycobacterial FurA is a homologue of the ferric uptake regulator Fur and is encoded by a gene located immediately upstream of katG. Here, we examine the possibility that FurA regulates katG expression. Inactivation of furA on the Mycobacterium smegmatis chromosome, a mycobacterial species that also lacks an oxyR homologue, resulted in derepression of katG, concomitant with increased resistance of the furA mutant to H2O2. In addition, M. smegmatis furA::Km(r) was more sensitive to the front-line antituberculosis agent isonicotinic acid hydrazide (INH) compared with the parental furA+ strain. The phenotypic manifestations were specific, as the mutant strain did not show altered sensitivity to organic peroxides, and both H2O2 and INH susceptibility profiles were complemented by the wild-type furA+ gene. We conclude that FurA is a second regulator of oxidative stress response in mycobacteria and that it negatively controls katG. In species lacking a functional oxyR, such as M. tuberculosis and M. smegmatis, FurA appears to be a dominant regulator affecting mycobacterial physiology and intracellular survival. PMID:11251835

Zahrt, T C; Song, J; Siple, J; Deretic, V

2001-03-01

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Distribution of hsp65 PCR-Restriction Enzyme Analysis Patterns among Mycobacterium avium Complex Isolates in Thailand?  

OpenAIRE

A total of 227 clinical Mycobacterium avium complex isolates from Thailand were differentiated into species and types by using PCR-restriction enzyme analysis of hsp65. The distribution of types showed the predominance of M. avium I (77%) in blood specimens, whereas M. intracellulare I was more commonly found in pulmonary specimens (44.2%). In addition, infections with M. avium were more likely to be found in younger adults (20 to 39 years old), while infections with M. intracellulare were mo...

Prammananan, Therdsak; Phunpruch, Saranya; Tingtoy, Nipa; Srimuang, Somboon; Chaiprasert, Angkana

2006-01-01

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The 'tubulin-like' S1 protein of Spirochaeta is a member of the hsp65 stress protein family  

Science.gov (United States)

A 65-kDa protein (called S1) from Spirochaeta bajacaliforniensis was identified as 'tubulin-like' because it cross-reacted with at least four different antisera raised against tubulin and was isolated, with a co-polymerizing 45-kDa protein, by warm-cold cycling procedures used to purify tubulin from mammalian brain. Furthermore, at least three genera of non-cultivable symbiotic spirochetes (Pillotina, Diplocalyx, and Hollandina) that contain conspicuous 24-nm cytoplasmic tubules displayed a strong fluorescence in situ when treated with polyclonal antisera raised against tubulin. Here we summarize results that lead to the conclusion that this 65-kDa protein has no homology to tubulin. S1 is an hsp65 stress protein homologue. Hsp65 is a highly immunogenic family of hsp60 proteins which includes the 65-kDa antigens of Mycobacterium tuberculosis (an active component of Freund's complete adjuvant), Borrelia, Treponema, Chlamydia, Legionella, and Salmonella. The hsp60s, also known as chaperonins, include E. coli GroEL, mitochondrial and chloroplast chaperonins, the pea aphid 'symbionin' and many other proteins involved in protein folding and the stress response.

Munson, D.; Obar, R.; Tzertzinis, G.; Margulis, L.

1993-01-01

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Comparison of polymerase chain reaction amplification of two mycobacterial DNA sequences, IS6110 and the 65kDa antigen gene, in the diagnosis of tuberculosis.  

OpenAIRE

BACKGROUND: Knowledge of the sequences of mycobacterial genes and the availability of DNA amplification techniques have raised the possibility that identification of mycobacterial DNA may offer a rapid and specific diagnostic test for tuberculosis. The correlation between the presence of Mycobacterium tuberculosis DNA and clinical tuberculosis, however, is not known. This study compared the results of polymerase chain reaction amplification of two M tuberculosis DNA sequences, IS6110 and the ...

Walker, D. A.; Taylor, I. K.; Mitchell, D. M.; Shaw, R. J.

1992-01-01

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Mycobacterial Diseases  

Science.gov (United States)

... Between Lab and Live Results . Related Links Tuberculosis Leprosy (Hansen's Disease) National Library of Medicine, MedlinePlus Javascript ... can be found throughout the world. Tuberculosis and leprosy (Hansen’s disease) are the best known mycobacterial diseases. ...

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Green fluorescent protein as a marker for gene expression and cell biology of mycobacterial interactions with macrophages.  

Science.gov (United States)

The green fluorescent protein (GFP) of the jellyfish Aequorea victoria offers certain advantages over other bioluminescence systems because no exogenously added substrate or co-factors are necessary, and fluorescence can be elicited by irradiation with blue light without exposing the cells producing GFP to invasive treatments. A mycobacterial shuttle-plasmid vector carrying gfp cDNA was constructed and used to generate transcriptional fusions with promoters of interest and to examine their expression in Mycobacterium smegmatis and Mycobacterium bovis BCG grown in macrophages or on laboratory media. The promoters studied were: (i) ahpC from Mycoosis and Mycobacterium leprae, a gene encoding alkyl hydroperoxide reductase which, along with the divergently transcribed regulator oxyR, are homologues of corresponding stress-response systems in enteric bacteria and play a role in isoniazid sensitivity; (ii) mtrA, an M. tuberculosis response regulator belonging to the superfamily of bacterial two-component signal-transduction systems; (iii) hsp60, a previously characterized heat-shock gene from M. bovis; and (iv) tbprc3, a newly isolated promoter from M. tuberculosis. Expression of these promoters in mycobacteria was analysed using epifluorescence microscopy, laser scanning confocal microscopy, fluorescence spectroscopy, and flow cytometry. These approaches permitted assessment of fluorescence prior to and after macrophage infection, and analyses of promoter expression in individual mycobacteria and its distribution within populations of bacterial cells. Bacteria expressing GFP from a strong promoter could be separated by fluorescence-activated cell sorting from cells harbouring the vector used to construct the fusion. In addition, the stable expression of mtrA-gfp fusion in M. bovis BCG facilitated localization and isolation of phagocytic vesicles containing mycobacteria. The experiments presented here suggest that GFP will be a useful tool for analysis of mycobacterial gene expression and a convenient cell biology marker to study mycobacterial interactions with macrophages. PMID:8596439

Dhandayuthapani, S; Via, L E; Thomas, C A; Horowitz, P M; Deretic, D; Deretic, V

1995-09-01

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Transcriptional Control of the Iron-Responsive fxbA Gene by the Mycobacterial Regulator IdeR†  

OpenAIRE

Exochelin is the primary extracellular siderophore of Mycobacterium smegmatis, and the iron-regulated fxbA gene encodes a putative formyltransferase, an essential enzyme in the exochelin biosynthetic pathway (E. H. Fiss, Y. Yu, and W. R. Jacobs, Jr., Mol. Microbiol. 14:557–569, 1994). We investigated the regulation of fxbA by the mycobacterial IdeR, a homolog of the Corynebacterium diphtheriae iron regulator DtxR (M. P. Schmitt, M. Predich, L. Doukhan, I. Smith, and R. K. Holmes, Infect. Im...

Dussurget, Olivier; Timm, Juliano; Gomez, Manuel; Gold, Benjamin; Yu, Shengwei; Sabol, Sue Z.; Holmes, Randall K.; Jacobs, William R.; Smith, Issar

1999-01-01

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Identification of Mycobacterium avium Genotypes with Distinctive Traits by Combination of IS1245-Based Restriction Fragment Length Polymorphism and Restriction Analysis of hsp65  

OpenAIRE

One-hundred eight Mycobacterium avium isolates from pigs, humans, birds, and bovines were typed by the IS1245-based restriction fragment length polymorphism (RFLP) method and PCR-restriction enzyme analysis (PRA) of hsp65. Nine clusters of isolates showing more than 80% similarity in their RFLP profiles were detected. The largest cluster (cluster B) included 32 of 79 pig isolates (40.5%), 3 of 25 human isolates (12%), and 1 of 2 bovine isolates, comprising 33% of all isolates. The second larg...

Oliveira, R. S.; Sircili, M. P.; Oliveira, E. M. D.; Balian, S. C.; Ferreira-neto, J. S.; Lea?o, S. C.

2003-01-01

29

Molecular Characterization of Heterologous HIV-1gp120 Gene Expression Disruption in Mycobacterium bovis BCG Host Strain: A Critical Issue for Engineering Mycobacterial Based-Vaccine Vectors  

Directory of Open Access Journals (Sweden)

Full Text Available Mycobacterium bovis Bacillus Calmette-Guérin (BCG as a live vector of recombinant bacterial vaccine is a promising system to be used. In this study, we evaluate the disrupted expression of heterologous HIV-1gp120 gene in BCG Pasteur host strain using replicative vectors pMV261 and pJH222. pJH222 carries a lysine complementing gene in BCG lysine auxotrophs. The HIV-1 gp120 gene expression was regulated by BCG hsp60 promoter (in plasmid pMV261 and Mycobacteria spp. ?-antigen promoter (in plasmid pJH222. Among 14 rBCG:HIV-1gp120 (pMV261 colonies screened, 12 showed a partial deletion and two showed a complete deletion. However, deletion was not observed in all 10 rBCG:HIV-1gp120 (pJH222 colonies screened. In this study, we demonstrated that E. coli/Mycobacterial expression vectors bearing a weak promoter and lysine complementing gene in a recombinant lysine auxotroph of BCG could prevent genetic rearrangements and disruption of HIV 1gp120 gene expression, a key issue for engineering Mycobacterial based vaccine vectors.

Joan Joseph

2010-01-01

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Identification of Mycobacterium avium genotypes with distinctive traits by combination of IS1245-based restriction fragment length polymorphism and restriction analysis of hsp65.  

Science.gov (United States)

One-hundred eight Mycobacterium avium isolates from pigs, humans, birds, and bovines were typed by the IS1245-based restriction fragment length polymorphism (RFLP) method and PCR-restriction enzyme analysis (PRA) of hsp65. Nine clusters of isolates showing more than 80% similarity in their RFLP profiles were detected. The largest cluster (cluster B) included 32 of 79 pig isolates (40.5%), 3 of 25 human isolates (12%), and 1 of 2 bovine isolates, comprising 33% of all isolates. The second largest cluster (cluster A) included 18 pig isolates (22.8%) and 6 human isolates (24%). Six smaller clusters included six pig isolates (clusters C and D), four and two human isolates (clusters E and F, respectively), two pig isolates (cluster I), and two pig isolates plus one bovine isolate and the avian purified protein derivative strain (cluster H). Cluster G represented the "bird-type" profile and included the bird isolate in this series, one pig isolate, plus reference strain R13. PRA revealed four allelic variants. Seventy-seven isolates were identified as M. avium PRA variant I, 24 were identified as M. avium PRA variant II, 6 were identified as M. avium PRA variant III, and 1 was identified as M. avium PRA variant IV. Except for three isolates from cluster B, each of the RFLP clusters was associated with a single PRA pattern. Isolates with unique (nonclustered) RFLP profiles were distributed between PRA variants I and II, and there was one unique isolate of PRA variant IV. These observations are consistent with divergent evolution within M. avium, resulting in the emergence of distinct lineages with particular competence to infect animals and humans. PMID:12517823

Oliveira, R S; Sircili, M P; Oliveira, E M D; Balian, S C; Ferreira-Neto, J S; Leão, S C

2003-01-01

31

Exposure to Antibiotics Induces Expression of the Mycobacterium tuberculosis sigF Gene: Implications for Chemotherapy against Mycobacterial Persistors  

OpenAIRE

The sigF gene encodes an alternate sigma factor found in Mycobacterium tuberculosis and related pathogenic mycobacteria. Determination of conditions of sigF expression is an important step in understanding the conditional gene regulation which may govern such processes as virulence and dormancy in mycobacteria. We constructed an in-frame translational lacZ-kan fusion within the sigF gene to determine the conditions of sigF expression. This reporter construct was expressed from a multicopy pla...

Michele, Theresa M.; Ko, Chiew; Bishai, William R.

1999-01-01

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The First Korean Case of Nontuberculous Mycobacterial Lung Disease Caused by Mycobacterium abscessus Subspecies bolletii in a Patient with Bronchiectasis  

OpenAIRE

We report the first Korean case of lung diseases caused by Mycobacterium abscessus subsp. bolletii in a previously healthy male, except for a previous history of pulmonary tuberculosis and bronchiectasis. All serial isolates are identified as M. abscessus subsp. bolletii by multi-locus sequence analysis based on the hsp65, rpoB, and 16S rRNA fragments. At the genetic level, the isolate has the erm(41) gene with a T28 sequevar, associated with clarithromycin resistance, and no rrl mutation. Th...

Jeong, Byeong-ho; Kim, Su-young; Jeon, Kyeongman; Huh, Hee Jae; Ki, Chang-seok; Lee, Nam Yong; Shin, Sung Jae; Koh, Won-jung

2014-01-01

33

Quinolones for mycobacterial infections.  

Science.gov (United States)

The fluoroquinolones (FQs) are important agents for the treatment of mycobacterial infections. In leprosy, the use of FQs has enabled a dramatic shortening of formerly long and complicated therapy. Both animal and human studies support the inclusion of certain FQs as a cornerstone of leprosy therapy. In tuberculosis (TB), particularly in multidrug-resistant (MDR) infections, the place of the major antimycobacterial FQs is less clear as there is widespread resistance to these agents in areas of the world in which MDR-TB and extensively drug-resistant (XDR)-TB are prevalent, particularly in Southeast Asia. The place of the newly developed FQ-related diarylquinoline compound known as bedaquiline in the treatment of drug-resistant TB is unclear; however, human studies suggest that it might be effective for this indication. PMID:23643393

Rubinstein, Ethan; Keynan, Yoav

2013-07-01

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Phosphorylation regulates mycobacterial proteasome.  

Science.gov (United States)

Mycobacterium tuberculosis possesses a proteasome system that is required for the microbe to resist elimination by the host immune system. Despite the importance of the proteasome in the pathogenesis of tuberculosis, the molecular mechanisms by which proteasome activity is controlled remain largely unknown. Here, we demonstrate that the ?-subunit (PrcA) of the M. tuberculosis proteasome is phosphorylated by the PknB kinase at three threonine residues (T84, T202, and T178) in a sequential manner. Furthermore, the proteasome with phosphorylated PrcA enhances the degradation of Ino1, a known proteasomal substrate, suggesting that PknB regulates the proteolytic activity of the proteasome. Previous studies showed that depletion of the proteasome and the proteasome-associated proteins decreases resistance to reactive nitrogen intermediates (RNIs) but increases resistance to hydrogen peroxide (H2O2). Here we show that PknA phosphorylation of unprocessed proteasome ?-subunit (pre-PrcB) and ?-subunit reduces the assembly of the proteasome complex and thereby enhances the mycobacterial resistance to H2O2 and that H2O2 stress diminishes the formation of the proteasome complex in a PknA-dependent manner. These findings indicate that phosphorylation of the M. tuberculosis proteasome not only modulates proteolytic activity of the proteasome, but also affects the proteasome complex formation contributing to the survival of M. tuberculosis under oxidative stress conditions. PMID:25224505

Anandan, Tripti; Han, Jaeil; Baun, Heather; Nyayapathy, Seeta; Brown, Jacob T; Dial, Rebekah L; Moltalvo, Juan A; Kim, Min-Seon; Yang, Seung Hwan; Ronning, Donald R; Husson, Robert N; Suh, Joowon; Kang, Choong-Min

2014-09-01

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Análise de restrição enzimática do gene hsp65 de isolados clínicos de pacientes com suspeita de tuberculose pulmonar em Teresina, Piauí Restriction enzyme analysis of the hsp65 gene in clinical isolates from patients suspected of having pulmonary tuberculosis in Teresina, Brazil  

OpenAIRE

OBJETIVO: Identificar as espécies de micobactérias encontradas no escarro de pacientes com suspeita de tuberculose pulmonar e analisar o impacto dessas identificações na abordagem terapêutica. MÉTODOS: Foram avaliados 106 pacientes com suspeita de tuberculose pulmonar encaminhados para o serviço de pneumologia de um hospital público em Teresina, Piauí. Espécimes de escarro matinal foram avaliados quanto à presença de micobactérias por baciloscopia e cultura. Foram utilizadas PCR ...

Bona, Maria Das Grac?as Motta E.; Maria José Soares Leal; Liline Maria Soares Martins; Raimundo Nonato da Silva; José Adail Fonseca De Castro; Semiramis Jamil Hadad do Monte

2011-01-01

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Mycobacterial diseases of deer.  

Science.gov (United States)

The most significant mycobacterial diseases of free-living, captive and farmed deer are bovine tuberculosis, caused by Mycobacterium bovis, Johne's disease (paratuberculosis), caused by Mycobacterium avium subsp paratuberculosis (basonym M. paratuberculosis), and avian tuberculosis, caused principally by M. avium subsp avium. The first case of M. bovis infection in farmed deer was identified in New Zealand in 1978. In 1983, a voluntary scheme was introduced in New Zealand to control tuberculosis in farmed deer, followed by a compulsory tuberculosis control scheme in 1990. The primary control measure is the slaughter of infected animals, detected by skin testing and blood testing, together with movement control and vector control. The number of infected deer herds peaked in the mid 1990s at over 160 herds, but by 30 June 2002 this had been reduced to 79 (1.45%), and to 67 (1.23%) by June 2003. Deer-to-deer transmission occurs, but the majority of herd breakdowns are believed to be from infected vectors. Factors likely to affect the susceptibility of deer include age, environment, population density, exposure and genetics. Avian tuberculosis occasionally causes clinical disease in wild, captive and farmed deer in New Zealand and overseas. Mycobacterium intracellulare, and subspecies of M. avium other than M. paratuberculosis, are widespread throughout New Zealand and are thought to be largely responsible for the high level of sensitisation to avian purified protein derivative (PPD), which is used for comparison purposes in tuberculosis skin testing of deer in this country. Infections with these organisms are usually subclinical in farmed deer, although M. avium subsp avium commonly causes lesions in retropharyngeal, mesenteric and ileocaecal lymph nodes. These lesions cause problems because of their gross and microscopic similarity to those due to M. bovis infection. Birds and domestic animals are most likely to become infected via environmental contamination of food, water, bedding litter or soil, while carnivores or scavengers may also become infected by ingesting infected carcasses. Johne's disease has been reported in deer in the wild and in zoos, especially in North America, the United Kingdom (UK) and Europe. Since first being confirmed in farmed deer in New Zealand in 1979, the incidence of Johne's disease has increased steadily. To date, M. paratuberculosis has been identified in >600 farmed deer on 300 properties. The majority of cases have been identified from suspected tuberculous lesions submitted from deer slaughter plants. Clinically, Johne's disease in deer is similar to the disease in sheep and cattle, with typical signs of loss of weight and condition, and diarrhoea. However, outbreaks of Johne's disease frequently occur in young red deer, 8-15 months of age, whereas the clinical disease in sheep and cattle is sporadic and usually affects adults 3-5 years of age. The disease is characterised by a chronic granulomatous enteritis and lymphadenitis, especially affecting the jejunum and ileum and the mesenteric lymph nodes. Deer affected subclinically may have lesions in these lymph nodes at slaughter, which are grossly indistinguishable from those due to bovine tuberculosis. Because of the antigenic similarity between M. intracellulare and all the subspecies of M. avium, including M. paratuberculosis, the diagnostic tests for Johne's disease lack sensitivity and specificity, making control difficult. PMID:15726126

Mackintosh, C G; de Lisle, G W; Collins, D M; Griffin, J F T

2004-08-01

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Phenotypic heterogeneity in mycobacterial stringent response  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background A common survival strategy of microorganisms subjected to stress involves the generation of phenotypic heterogeneity in the isogenic microbial population enabling a subset of the population to survive under stress. In a recent study, a mycobacterial population of M. smegmatis was shown to develop phenotypic heterogeneity under nutrient depletion. The observed heterogeneity is in the form of a bimodal distribution of the expression levels of the Green Fluorescent Protein (GFP as reporter with the gfp fused to the promoter of the rel gene. The stringent response pathway is initiated in the subpopulation with high rel activity. Results In the present study, we characterise quantitatively the single cell promoter activity of the three key genes, namely, mprA, sigE and rel, in the stringent response pathway with gfp as the reporter. The origin of bimodality in the GFP distribution lies in two stable expression states, i.e., bistability. We develop a theoretical model to study the dynamics of the stringent response pathway. The model incorporates a recently proposed mechanism of bistability based on positive feedback and cell growth retardation due to protein synthesis. Based on flow cytometry data, we establish that the distribution of GFP levels in the mycobacterial population at any point of time is a linear superposition of two invariant distributions, one Gaussian and the other lognormal, with only the coefficients in the linear combination depending on time. This allows us to use a binning algorithm and determine the time variation of the mean protein level, the fraction of cells in a subpopulation and also the coefficient of variation, a measure of gene expression noise. Conclusions The results of the theoretical model along with a comprehensive analysis of the flow cytometry data provide definitive evidence for the coexistence of two subpopulations with overlapping protein distributions.

Bose Indrani

2011-01-01

38

Mycobacterial DNA gyrase: enzyme purification and characterization of supercoiling activity.  

Science.gov (United States)

Putative structural genes encoding Mycobacterium bovis BCG gyrase A and gyrase B subunits were expressed in Escherichia coli under the control of a regulated promoter. Upon induction, high levels of proteins of M(r) 92,000 and 75,000 were generated. Purification and reconstitution of these proteins yielded an enzyme with bacterial DNA gyrase activity. DNA supercoiling activity of the mycobacterial enzyme required ATP, Mg2+, and spermidine. Like other bacterial DNA gyrases, the supercoiling activity of the mycobacterial enzyme was inhibited by low concentration of the classical gyrase B subunit inhibitors novobiocin and coumermycin. Older gyrase A subunit inhibitors, nalidixic and oxolinic acid, had no effect on the supercoiling activity at 400 to 800 micrograms/ml. However, in vitro assays to show the inhibition of supercoiling activity and stimulation of cleavable complex formation demonstrated that ciprofloxacin is a potent inhibitor of mycobacterial DNA gyrase. The availability of highly purified mycobacterial DNA gyrase could aid in future investigations of quinolone derivatives targeting Mycobacterium specifically. PMID:7503546

Wu, L C; Shahied, S I

1995-12-01

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The First Korean Case of Nontuberculous Mycobacterial Lung Disease Caused by Mycobacterium abscessus Subspecies bolletii in a Patient with Bronchiectasis.  

Science.gov (United States)

We report the first Korean case of lung diseases caused by Mycobacterium abscessus subsp. bolletii in a previously healthy male, except for a previous history of pulmonary tuberculosis and bronchiectasis. All serial isolates are identified as M. abscessus subsp. bolletii by multi-locus sequence analysis based on the hsp65, rpoB, and 16S rRNA fragments. At the genetic level, the isolate has the erm(41) gene with a T28 sequevar, associated with clarithromycin resistance, and no rrl mutation. The isolate is resistant to clarithromycin. Although the symptoms and radiographic findings have improved after combination of antibiotics, the follow-up sputum cultures are persistently positive. PMID:24523815

Jeong, Byeong-Ho; Kim, Su-Young; Jeon, Kyeongman; Huh, Hee Jae; Ki, Chang-Seok; Lee, Nam Yong; Shin, Sung Jae; Koh, Won-Jung

2014-01-01

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Molecular Characterization of Heterologous HIV-1gp120 Gene Expression Disruption in Mycobacterium bovis BCG Host Strain: A Critical Issue for Engineering Mycobacterial Based-Vaccine Vectors  

OpenAIRE

Mycobacterium bovis Bacillus Calmette-Guérin (BCG) as a live vector of recombinant bacterial vaccine is a promising system to be used. In this study, we evaluate the disrupted expression of heterologous HIV-1gp120 gene in BCG Pasteur host strain using replicative vectors pMV261 and pJH222. pJH222 carries a lysine complementing gene in BCG lysine auxotrophs. The HIV-1 gp120 gene expression was regulated by BCG hsp60 promoter (in plasmid pMV261) and Mycobacteria spp. ?-antigen promo...

Joan Joseph; Raquel Fernández-Lloris; Elías Pezzat; Saubi, Narc Amp S.; Pere-Joan Cardona; Beatriz Mothe; Josep Maria Gatell

2010-01-01

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Protein Turnover in Mycobacterial Proteomics  

Directory of Open Access Journals (Sweden)

Full Text Available Understanding the biology of Mycobacterium tuberculosis is one of the primary challenges in current tuberculosis research. Investigation of mycobacterial biology using the systems biology approach has deciphered much information with regard to the bacilli and tuberculosis pathogenesis. The modulation of its environment and the ability to enter a dormant phase are the hallmarks of M. tuberculosis. Until now, proteome studies have been able to understand much about the role of various proteins, mostly in growing M. tuberculosis cells. It has been difficult to study dormant M. tuberculosis by conventional proteomic techniques with very few proteins being found to be differentially expressed. Discrepancy between proteome and transcriptome studies lead to the conclusion that a certain aspect of the mycobacterial proteome is not being explored. Analysis of protein turnover may be the answer to this dilemma. This review, while giving a gist of the proteome response of mycobacteria to various stresses, analyzes the data obtained from abundance studies versus data from protein turnover studies in M. tuberculosis. This review brings forth the point that protein turnover analysis is capable of discerning more subtle changes in protein synthesis, degradation, and secretion activities. Thus, turnover studies could be incorporated to provide a more in-depth view into the proteome, especially in dormant or persistent cells. Turnover analysis might prove helpful in drug discovery and a better understanding of the dynamic nature of the proteome of mycobacteria.

Prahlad K. Rao

2009-08-01

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Surgical treatment of nontuberculous mycobacterial lung disease.  

Science.gov (United States)

While the prevalence of pulmonary tuberculosis has been decreasing, the prevalence of nontuberculous mycobacterial lung disease has been increasing. Unlike tuberculosis, nontuberculous mycobacterial disease is not communicable. However, their indolent nature may result in extensive parenchymal destruction, causing respiratory failure and vulnerability to airway infection. Nontuberculous mycobacterial lung disease, therefore, has been becoming a significant health problem. According to the 2007 American Thoracic Society/Infectious Diseases Society of America statement on nontuberculous mycobacterial diseases, the primary treatment is a multidrug treatment regimen. However, its efficacy is less than satisfactory for Mycobacterium avium complex lung disease, which is the most common type of nontuberculous mycobacterial lung diseases, and for Mycobacterium abscessus lung disease, which is notoriously resistant to chemotherapeutic drugs. The statement, therefore, has proposed a multidisciplinary treatment approach for these types of nontuberculous mycobacterial lung diseases: a combination of multidrug treatment regimen and adjuvant resectional surgery. This review covers the rationale, indication, procedure, and outcome of surgical treatment of nontuberculous mycobacterial lung disease. The rationale of surgery is to prevent disease progressing by removing the areas of lung most affected, harboring the largest amounts of mycobacteria. The indications for surgery include a poor response to drug therapy, the development of macrolide-resistant disease, or the presence of a significant disease-related complication such as hemoptysis. The surgical procedures of choice are various types of pulmonary resections, including wedge resection, segmentectomy, lobectomy, or pneumonectomy. The reported series have achieved favorable treatment outcomes in surgically treated patients with acceptable morbidity and mortality rates. PMID:24740640

Shiraishi, Yuji

2014-08-01

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Mycobacterial skin and soft tissue infection.  

Science.gov (United States)

Mycobacterial skin and soft tissue infection (SSTI) includes nontuberculous mycobacterial (NTM) infections, tuberculosis (TB), and leprosy. Diagnosis of mycobacterial SSTI can be challenging due to diverse clinical presentation, low yield from cultured specimens, and nonspecific histopathology on tissue biopsy. In addition, immunosuppressed patients may present with atypical or disseminated disease. Despite aggressive medical treatment and often with surgical intervention, results may be suboptimal with poor outcomes. Regimens typically require multiple antibiotics for extended periods of time and are often complicated by medication side effects and drug-drug interactions. Biopsy with culture is the gold standard for diagnosis, but newer molecular diagnostics and proteomics such as matrix-assisted laser desorption ionization-time of flight mass spectrometry have improved diagnosis with increased identification of clinically significant mycobacteria species in clinically relevant time frames. We will review updates in diagnostic tests along with clinical presentation and treatment of mycobacterial SSTI for NTM, TB, and leprosy. PMID:25339245

Wang, Shu-Hua; Pancholi, Preeti

2014-11-01

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Nontuberculous mycobacterial peritonitis in peritoneal dialysis patients.  

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While nontuberculous mycobacterial peritonitis is uncommon among peritoneal dialysis (PD) patients, these infections have serious consequences. They present a significant diagnostic and therapeutic challenge for clinicians. Diagnosis can be delayed due to the slow growth rate of some mycobacterial species. These organisms can also be overlooked when adequate culture media are not used in the microbiological evaluation process. The choice of antimicrobial therapy depends upon isolation and speciation of the infecting Mycobacterium species, and prompt catheter removal is essential. Because serious intra-abdominal complications may follow infection, identifying patient risk factors for nontuberculous mycobacterial peritonitis and initiating prompt diagnosis and treatment are essential. We report three cases of peritonitis associated with Mycobacterium chelonae and Mycobacterium gordonae, each with a unique presentation, and discuss the appropriate diagnosis and treatment strategies for the management of PD-associated mycobacterial infections. PMID:17555495

Rho, Mira; Bia, Frank; Brewster, Ursula C

2007-01-01

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Serine threonine protein kinases of mycobacterial genus: phylogeny to function.  

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Serine/threonine protein kinases (STPKs) are known to act as sensors of environmental signals that thereby regulate developmental changes and host pathogen interactions. In this study, we carried out comparative genome analysis of six completely sequenced pathogenic and nonpathogenic mycobacterial species to systematically characterize the STPK complement of mycobacterium. Our analysis revealed that while Mycobacterium tuberculosis strains have 11 conserved kinases, this number varies from 4 to 24 in other mycobacterial species. pknA, an essential STPK encoding gene, was found to be truncated in the initial analysis of M. avium subsp. paratuberculosis (Map) and M. tuberculosis C genomes. However, resequencing of pknA gene in Map confirmed that the truncation was due to a sequencing error. The conservation of division and cell wall gene cluster involved in cell envelope biosynthesis and cell division, in the vicinity of pknL locus, implicates a possible role of PknL in cell division and envelop biosynthesis. We identified a cyclophilin domain as part of a mycobacterial kinase in Map that suggests a plausible regulation of cyclophilins by phosphorylation. The co-inheritance of pknA, pknB, pknG, and pknL loci across genomes and some unique repertoire of pathogen-specific kinases such as pknI and pknJ of Mtb complex suggest similitude and divergence between pathogenic and nonpathogenic signaling. This study would add another dimension toward identification of physiological substrates and thereby function, while resolving the existing complexities in signaling network between the two domains of life, pathogen and nonpathogen. PMID:17148687

Narayan, Azeet; Sachdeva, Preeti; Sharma, Kirti; Saini, Adesh K; Tyagi, Anil K; Singh, Yogendra

2007-03-14

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Nontuberculous Mycobacterial Ocular Infections—Comparing the Clinical and Microbiological Characteristics between Mycobacterium abscessus and Mycobacterium massiliense  

Science.gov (United States)

Purpose To analyze the clinical characteristics of nontuberculous mycobacterial (NTM) ocular infections and the species-specific in vitro antimicrobial susceptibility. Material and Methods In 2000 to 2011 at the National Taiwan University Hospital, multilocus sequencing of rpoB, hsp65 and secA was used to identify NTM isolates from ocular infections. The clinical presentation and treatment outcomes were retrospectively compared between species. Broth microdilution method was used to determine the minimum inhibitory concentrations of amikacin (AMK), clarithromycin (CLA), ciprofloxacin (CPF), levofloxacin (LVF), moxifloxacin (MXF) and gatifloxacin (GAF) against all strains. The activities of antimicrobial combinations were assessed by the checkerboard titration method. Results A total of 24 NTM strains (13 Mycobacterium abscessus and 11 Mycobacterium massiliense) were isolated from 13 keratitis, 10 buckle infections, and 1 canaliculitis cases. Clinically, manifestations and outcomes caused by these two species were similar and surgical intervention was necessary for medically unresponsive NTM infection. Microbiologically, 100% of M. abscessus and 90.9% of M. massiliense ocular isolates were susceptible to amikacin but all were resistant to fluoroquinolones. Inducible clarithromycin resistance existed in 69.3% of M. abscessus but not in M. massiliense isolates. None of the AMK-CLA, AMK-MXF, AMK-GAF, CLA-MXF and CLA-GAF combinations showed synergistic or antagonistic effect against both species in vitro. Conclusions M. abscessus and M. massiliense are the most commonly identified species for NTM ocular infections in Taiwan. Both species were resistant to fluoroquinolones, susceptible to amikacin, and differ in clarithromycin resistance. Combined antimicrobial treatments showed no interaction in vitro but could be considered in combination with surgical interventions for eradication of this devastating ocular infection. PMID:25581038

Chu, Hsiao-Sang; Chang, Shan-Chwen; Shen, Elizabeth P.; Hu, Fung-Rong

2015-01-01

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Mycobacterial species as case-study of comparative genome analysis  

DEFF Research Database (Denmark)

The genus Mycobacterium represents more than 120 species including important pathogens of human and cause major public health problems and illnesses. Further, with more than 100 genome sequences from this genus, comparative genome analysis can provide new insights for better understanding the evolutionary events of these species and improving drugs, vaccines, and diagnostics tools for controlling Mycobacterial diseases. In this present study we aim to outline a comparative genome analysis of fourteen Mycobacterial genomes: M. avium subsp. paratuberculosis K—10, M. bovis AF2122/97, M. bovis BCG str. Pasteur 1173P2, M. leprae Br4923, M. marinum M, M. sp. KMS, M. sp. MCS, M. tuberculosis CDC1551, M. tuberculosis F11, M. tuberculosis H37Ra, M. tuberculosis H37Rv, M. tuberculosis KZN 1435 , M. ulcerans Agy99,and M. vanbaalenii PYR—1, For this purpose a comparison has been done based on their length of genomes, GC content, number of genes in different data bases (Genbank, Refseq, and Prodigal). The BLAST matrix of these genomes has been figured to give a lot of information about the similarity between species in a simple scheme. As a result of multiple genome analysis, the pan and core genome have been defined for twelve Mycobacterial species. We have also introduced the genome atlas of the reference strain M. tuberculosis H37Rv which can give a good overview of this genome. And for examining the phylogenetic relationships among these bacteria, a phylogenic tree has been constructed from 16S rRNA gene for tuberculosis and non tuberculosis Mycobacteria to understand the evolutionary events of these species.

Zakham, F.; Belayachi, L.

2011-01-01

48

MENDELIAN SUSCEPTIBILITY TO MYCOBACTERIAL DISEASE IN EGYPTIAN CHILDREN  

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Full Text Available

Background: Tuberculosis remains a major health problem in developing countries especially with the emergence of multidrug resistant strains. Mendelian Susceptibility to Mycobacterial Disease (MSMD is a rare disorder with impaired immunity against mycobacterial pathogens. Reported MSMD etiologies highlight the crucial role of the Interferon gamma /Interleukin 12 (IFN-g/ IL-12 axis and the phagocyte respiratory burst axis.

Purpose: Screen patients with possible presentations for MSMD.

Methods: Patients with disseminated BCG infection following vaccination, atypical mycobacterial infections or recurrent tuberculosis infections were recruited from the Primary Immune Deficiency Clinic at Cairo University Specialized Pediatric Hospital, Egypt and immune and genetic laboratory investigations were conducted at Human Genetic of Infectious Diseases laboratory in Necker Medical School, France from 2005-2009. IFN-g level in patient’s plasma as well as mutations in the eight previously identified MSMD-causing genes were explored.

Results: Nine cases from eight (unrelated kindreds were evaluated in detail. We detected a high level of IFN-g in plasma in one patient. Through Sanger sequencing, a homozygous mutation in the IFNGR1 gene at position 485 corresponding to an amino acid change from serine to phenylalanine (S485F, was detected in this patient.

Conclusion: We report the first identified cases of MSMD among Egyptian patients, including in particular a new IFNGR1 mutation underlying IFN-gR1 deficiency. The eight remaining patients need to be explored further. These findings have implications regarding the compulsory Bacillus Calmette Guerin vaccination policy in Egypt, especially given the high consanguinity rate.

Keywords: Interferon gamma axis, mycobacterium tuberculosis, BCG, consanguinity

Nermeen Galal

2012-01-01

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DETECÇÃO DO COMPLEXO Mycobacterium tuberculosis NO LEITE PELA REAÇÃO EM CADEIA DA POLIMERASE SEGUIDA DE ANÁLISE DE RESTRIÇÃO DO FRAGMENTO AMPLIFICADO (PRA) DETECTION OF Mycobacterium tuberculosis COMPLEX BY PCR-RESTRICTION FRAGMENT LENGTH POLYMORFISM ANALYSIS OF THE HSP65 GENE  

OpenAIRE

Mycobacterium bovis é membro do complexo Mycobacterium tuberculosis (MTBC), grupo este composto por espécies com grande homologia genética. É o agente etiológico da tuberculose bovina, importante zoonose transmissível ao homem, principalmente através da inalação do bacilo e/ou pelo consumo de leite e derivados não-pasteurizados provenientes de vacas tuberculosas. O objetivo deste estudo foi padronizar a identificação de micobactérias do complexo M. tuberculosis presentes no leite...

Joab Trajano Silva; Leila Souza Fonseca; Marlei Gomes da Silva; Eduardo Eustáquio de Souza Figueiredo; Vânia Margaret Flosi Paschoalin

2008-01-01

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Nontuberculous mycobacterial infections in children with inborn errors of the immune system.  

Science.gov (United States)

Severe mycobacterial disease is mostly confined to patients who are immunocompromized either by acquired or inherited causes. One such genetic disorder is Mendelian Susceptibility to Mycobacterial Disease (MSMD), a hot topic within the field of primary immunodeficiency. This single gene disorder is characterized by isolated infection with mycobacteria or Salmonella due to a defect in the type-1 cytokine response. In the last two decades, ten genes have been labeled as causing MSMD when they harbor germline mutations, namely IL12B, IL12RB1, IFNGR1, IFNGR2, STAT1, IKBKG, CYBB, TYK2, IRF8 and ISG15. The mutations lead to either insufficient production of IFN-?, or to an insufficient response to the cytokine. Current treatment options include recombinant IFN-? and hematologic stem cell transplantation (HSCT). In the future, gene therapy, antisense-mediated exon skipping and chemical intervention in glycosylation problems may become successful alternatives. Furthermore, it is likely that many new candidate genes and pathways crucial for mycobacterial immunity will be identified. PMID:24119826

Haverkamp, Margje H; van de Vosse, Esther; van Dissel, Jaap T

2014-01-01

51

Non-tuberculous mycobacterial tenosynovitis: a review.  

Science.gov (United States)

The clinical characteristics, outcome and treatment of non-tuberculous mycobacterial tenosynovitis are reviewed. From lesions localized in the hand, 10 different species of non-tuberculous mycobacteria have been reported. The most common are Mycobacterium marinum and Mycobacterium kansasii. Other less frequent organisms are Mycobacterium avium complex, Mycobacterium szulgai, Mycobacterium terrae, Mycobacterium fortuitum, Mycobacterium chelonae, Mycobacterium abscessus, Mycobacterium malmoense and Mycobacterium xenopi. The infections appear to be the result of previous trauma, surgical procedure, corticosteroid injection or non-apparent inoculation (water contamination). Immunosuppression is sometimes associated with the infections and can be considered as a risk factor. Surgical debridement and appropriate mycobacterial cultures are critical to enable diagnosis and appropriate management. Specimens should be inoculated on a range of media and incubated at a range of temperatures in order to isolate mycobacteria with different growth characteristics (with prolonged incubation). The optimal treatment of these infections is discussed. PMID:10482048

Zenone, T; Boibieux, A; Tigaud, S; Fredenucci, J F; Vincent, V; Chidiac, C; Peyramond, D

1999-01-01

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Computational genomics-proteomics and Phylogeny analysis of twenty one mycobacterial genomes (Tuberculosis & non Tuberculosis strains  

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Full Text Available Abstract Background The genus Mycobacterium comprises different species, among them the most contagious and infectious bacteria. The members of the complex Mycobacterium tuberculosis are the most virulent microorganisms that have killed human and other mammals since millennia. Additionally, with the many different mycobacterial sequences available, there is a crucial need for the visualization and the simplification of their data. In this present study, we aim to highlight a comparative genome, proteome and phylogeny analysis between twenty-one mycobacterial (Tuberculosis and non tuberculosis strains using a set of computational and bioinformatics tools (Pan and Core genome plotting, BLAST matrix and phylogeny analysis. Results Considerably the result of pan and core genome Plotting demonstrated that less than 1250 Mycobacterium gene families are conserved across all species, and a total set of about 20,000 gene families within the Mycobacterium pan-genome of twenty one mycobacterial genomes. Viewing the BLAST matrix a high similarity was found among the species of the complex Mycobacterium tuberculosis and less conservation is found with other slow growing pathogenic mycobacteria. Phylogeny analysis based on both protein conservation, as well as rRNA clearly resolve known relationships between slow growing mycobacteria. Conclusion Mycobacteria include important pathogenic species for human and animals and the Mycobacterium tuberculosis complex is the most cause of death of the humankind. The comparative genome analysis could provide a new insight for better controlling and preventing these diseases.

Zakham Fathiah

2012-08-01

53

Mycobacterial disease in renal allograft recipients  

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Full Text Available Implication for health policy/practice/research/medical education:Solid organ transplant recipients have impaired cell-mediated immunity, and are at increased risk of mycobacterial infection. Mycobacterium tuberculosis infection (TB has a high mortality rate among this population. The diagnosis of tuberculosis in solid organ transplant recipients is a big challenge and needs rapid and accurate modalities. These patients have 3.8 time greater risk of developing extra-pulmonary TB than general population. High index of suspicion and applying with invasive diagnostic procedure are needed for diagnosis of TB in renal transplanted patients.

Ardalan Mohammad-Reza

2013-06-01

54

Role of Iron in Nramp1-Mediated Inhibition of Mycobacterial Growth  

OpenAIRE

Innate resistance to mycobacterial growth is mediated by a gene, Nramp1. We have previously reported that Nramp1 mRNA from macrophages of Mycobacterium bovis BCG-resistant (Bcgr) mice is more stable than Nramp1 mRNA from macrophages of BCG-susceptible (Bcgs) mice. Based on these observations and on reports that show that the closely related Nramp2 gene is a metal ion transporter, we evaluated the effect of iron on the growth of Mycobacterium avium within macrophages as well as on the stabilit...

Zwilling, Bruce S.; Kuhn, Donald E.; Wikoff, Lisa; Brown, David; Lafuse, William

1999-01-01

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Recurrent pulmonary aspergillosis and mycobacterial infection in an unsplenectomized patient with type 1 Gaucher disease  

Science.gov (United States)

Background The clinical presentation of Gaucher disease (GD), an inherited lysosomal storage disorder caused by the deficient activity of the lysosomal enzyme glucocerebrosidase, is highly variable, and three clinical types are distinguished based upon the presence of neurologic symptoms. Thrombocytopenia, anemia, hepatosplenomegaly, and bone manifestations are the most typical signs of GD type 1 (GD1). Case presentation We present the case of an unsplenectomized man suffering from heterozygous GD1 with mutations of c.1226A>G (N370S) and RecNci I (L444P, A456P, and V460V) in the GBA1 gene, who developed recurrent pulmonary aspergillosis caused by Aspergillus fumigatus and a mycobacterial infection caused by Mycobacterium avium. Despite long-lasting therapy of both aspergillosis (including antifungal drugs and surgery), and the mycobacterial infection (triple therapy with rifampicin, ethambutol, and clarithromycin), recurrent positivity for M. avium and A. fumigatus was detected. Conclusions Symptomatic lung involvement and an increased susceptibility to pulmonary infections are uncommon in GD and, if present, are often associated with more severe disease manifestations. To our knowledge, this is the first published report on the association of GD and pulmonary aspergillosis and mycobacterial infection. It illustrates the increased susceptibility of untreated GD patients to opportunistic pulmonary infections and ineffective eradication of these infections despite adequate therapy. PMID:24195576

Lorenz, Fryderyk; Kleinotiene, Grazina; Bulanda, Agnieszka; Markuszewska-Kuczy?ska, Alicja; Raistenskis, Juozas; Klimkowska, Monika

2014-01-01

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Tetrahydrolipstatin Inhibition, Functional Analyses, and Three-dimensional Structure of a Lipase Essential for Mycobacterial Viability  

Energy Technology Data Exchange (ETDEWEB)

The highly complex and unique mycobacterial cell wall is critical to the survival of Mycobacteria in host cells. However, the biosynthetic pathways responsible for its synthesis are, in general, incompletely characterized. Rv3802c from Mycobacterium tuberculosis is a partially characterized phospholipase/thioesterase encoded within a genetic cluster dedicated to the synthesis of core structures of the mycobacterial cell wall, including mycolic acids and arabinogalactan. Enzymatic assays performed with purified recombinant proteins Rv3802c and its close homologs from Mycobacterium smegmatis (MSMEG{_}6394) and Corynebacterium glutamicum (NCgl2775) show that they all have significant lipase activities that are inhibited by tetrahydrolipstatin, an anti-obesity drug that coincidently inhibits mycobacterial cell wall biosynthesis. The crystal structure of MSMEG{_}6394, solved to 2.9 {angstrom} resolution, revealed an {alpha}/{beta} hydrolase fold and a catalytic triad typically present in esterases and lipases. Furthermore, we demonstrate direct evidence of gene essentiality in M. smegmatis and show the structural consequences of loss of MSMEG{_}6394 function on the cellular integrity of the organism. These findings, combined with the predicted essentiality of Rv3802c in M. tuberculosis, indicate that the Rv3802c family performs a fundamental and indispensable lipase-associated function in mycobacteria.

Crellin, Paul K.; Vivian, Julian P.; Scoble, Judith; Chow, Frances M.; West, Nicholas P.; Brammananth, Rajini; Proellocks, Nicholas I.; Shahine, Adam; Le Nours, Jerome; Wilce, Matthew C.J.; Britton, Warwick J.; Coppel, Ross L.; Rossjohn, Jamie; Beddoe, Travis (Monash); (Centenary)

2010-09-17

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Nontuberculous mycobacterial pulmonary diseases in immunocompetent patients  

Energy Technology Data Exchange (ETDEWEB)

Nontuberculous mycobacterial (NTM) infections are an increasingly recognized cause of chronic lung disease in immunocompetent adults, and the M. Avium complex, M. Kansasii, and a rapidly growing mycobacteria such as M. abscessus, M. Fortuitum, and M. Chelonae account for most of the pathogens involved. Because the clinical features of NTM disease are not distinguishable from those of tuberculosis, and NTM are ubiquitous in the environment, diagnosis requires that the bacilli are isolated and identified. NTM diseases have been difficult to treat, though since the introduction of new macrolides, the outcome for patients with some NTM diseases has improved significantly. For correct diagnosis and the successful treatment of NTM pulmonary disease, a knowledge of the full spectrum of clinical and radiological findings is important.

Koh, Won Jung; Kwon, O Jung; Lee, Kyung Soo [Sungkyunkwan University School of Medicine, Seoul (Korea, Republic of)

2002-09-01

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Nontuberculous mycobacterial pulmonary diseases in immunocompetent patients  

International Nuclear Information System (INIS)

Nontuberculous mycobacterial (NTM) infections are an increasingly recognized cause of chronic lung disease in immunocompetent adults, and the M. Avium complex, M. Kansasii, and a rapidly growing mycobacteria such as M. abscessus, M. Fortuitum, and M. Chelonae account for most of the pathogens involved. Because the clinical features of NTM disease are not distinguishable from those of tuberculosis, and NTM are ubiquitous in the environment, diagnosis requires that the bacilli are isolated and identified. NTM diseases have been difficult to treat, though since the introduction of new macrolides, the outcome for patients with some NTM diseases has improved significantly. For correct diagnosis and the successful treatment of NTM pulmonary disease, a knowledge of the full spectrum of clinical and radiological findings is important

59

Immune Response to Mycobacterial Infection: Lessons from Flow Cytometry  

OpenAIRE

Detecting and treating active and latent tuberculosis are pivotal elements for effective infection control; yet, due to their significant inherent limitations, the diagnostic means for these two stages of tuberculosis (TB) to date remain suboptimal. This paper reviews the current diagnostic tools for mycobacterial infection and focuses on the application of flow cytometry as a promising method for rapid and reliable diagnosis of mycobacterial infection as well as discrimination between active...

Nikoletta Rovina; Marios Panagiotou; Konstantinos Pontikis; Magdalini Kyriakopoulou; Koulouris, Nikolaos G.; Antonia Koutsoukou

2013-01-01

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Polyphenolic acetates : A newer anti-Mycobacterial therapeutic option  

OpenAIRE

The objective of our research project was screening of various highly specific substrates of Acetoxy Drug: Protein Transacytylase (M.TAase) for antimycobacterial activity. Mycobacterial culture was done in Middlebrook’s 7H9 media. Protein purification (Mycobacterial Tranacetylase, M.TAase) was done by ion exchange chromatography and its demonstration was done on SDS- polyacrylamide gel electrophoresis (SDS-PAGE) and western blot. Middlebrook’s 7H9 broth was procured from Becton Dickinson....

Santram; Petra, Surajeet K.; Raj, H. G.; Parmar, V. S.; Garima Gupta; Bhagel, A. S.; Bose, M.; Mala Chandra

2014-01-01

61

Clinical agreement in pulmonary nontuberculous mycobacterial disease  

Directory of Open Access Journals (Sweden)

Full Text Available Peter J Nolan Department of Internal Medicine, Toowoomba General Hospital, Toowoomba, Queensland, Australia Purpose: To consider the clinical agreement among respiratory and infectious disease physicians, working in a tertiary chest diseases center serving a population with a low incidence of pulmonary tuberculosis (<3/100,000/year, in the assessment of cases of pulmonary nontuberculous mycobacterial (NTM lung disease. Method: A series of previously notified cases of NTM disease was abstracted and anonymously presented to a cohort of seven respiratory and infectious disease physicians. Their individual decisions to notify, treat, and follow the cases was evaluated and compared using the intraclass correlation coefficient. Results: A wide range was demonstrated in the diagnostic and management decision triage of each case by the physicians participating in the study. Clinical agreement on the likelihood of disease was limited, with an intraclass correlation coefficient of 0.394. Indication to notify the case to the state registry was linked to the clinical intent to initiate a treatment program. Conclusion: There appears to be limited agreement on the clinical significance of NTM isolates from pulmonary specimens among this cohort of experienced clinicians. If this trend is generalizable to a wider population of respiratory and infectious disease physicians, the number of notified and treated cases of disease is likely to be an underestimate of the true burden of disease in the general population. Keywords: diagnostic certainty, Kappa index, intraclass correlation coefficient, lung disease

Nolan PJ

2013-11-01

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Anti-mycobacterial peptides: from human to phage.  

Science.gov (United States)

Mycobacterium tuberculosis is the major pathogen of tuberculosis (TB). With the growing problem of M. tuberculosis resistant to conventional antibiotics, especially multi-drug resistant tuberculosis (MDR-TB) and extensively-drug resistant tuberculosis (XDR-TB), the need for new TB drugs is now more prominent than ever. Among the promising candidates for anti-TB drugs, anti-mycobacterial peptides have a few advantages, such as low immunogenicity, selective affinity to prokaryotic negatively charged cell envelopes, and diverse modes of action. In this review, we summarize the recent progress in the anti-mycobacterial peptides, highlighting the sources, effectiveness and bactericidal mechanisms of these antimicrobial peptides. Most of the current anti-mycobacterial peptides are derived either from host immune cells, bacterial extraction, or mycobacteriophages. Besides trans-membrane pore formation, which is considered to be the common bactericidal mechanism, many of the anti-mycobacterial peptides have the second non-membrane targets within mycobacteria. Additionally, some antimicrobial peptides play critical roles in innate immunity. However, a few obstacles, such as short half-life in vivo and resistance to antimicrobial peptides, need overcoming before clinical applications. Nevertheless, the multiple functions of anti-mycobacterial peptides, especially direct killing of pathogens and immune-modulators in infectious and inflammatory conditions, indicate that they are promising candidates for future drug development. © 2015 S. Karger AG, Basel. PMID:25613372

Teng, Tieshan; Liu, Jiafa; Wei, Hongping

2015-01-01

63

Mycobacterial infections in a large Virginia hospital, 2001-2009  

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Full Text Available Abstract Background In areas where both tuberculosis (TB and nontuberculous mycobacteria (NTM are prevalent, descriptive studies of the clinical features of individual mycobacteria are needed to inform clinical triage. Methods We queried the University of Virginia Clinical Data Repository for all mycobacterial infections from 2001-2009. Results Of 494 mycobacterial infections in 467 patients there were 22 species. Patients with pulmonary Tb were more likely to be reported as immigrants (p M. kansasii, M. xenopi, and M. fortuitum were more likely than MAC to have cavities. There were at least 83 patients that met criteria for NTM lung disease and these were caused by 9 species. M. abscessus infection was associated with cystic fibrosis and M. xenopi infection was associated with male gender. Conclusions In our center mycobacterial infections were common and of diverse species. Immigrant status, cavities, and effusion were associated with TB vs. NTM.

Scully Kenneth W

2011-05-01

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Immune response to mycobacterial infection: lessons from flow cytometry.  

Science.gov (United States)

Detecting and treating active and latent tuberculosis are pivotal elements for effective infection control; yet, due to their significant inherent limitations, the diagnostic means for these two stages of tuberculosis (TB) to date remain suboptimal. This paper reviews the current diagnostic tools for mycobacterial infection and focuses on the application of flow cytometry as a promising method for rapid and reliable diagnosis of mycobacterial infection as well as discrimination between active and latent TB: it summarizes diagnostic biomarkers distinguishing the two states of infection and also features of the distinct immune response against Mycobacterium tuberculosis (Mtb) at certain stages of infection as revealed by flow cytometry to date. PMID:24376464

Rovina, Nikoletta; Panagiotou, Marios; Pontikis, Konstantinos; Kyriakopoulou, Magdalini; Koulouris, Nikolaos G; Koutsoukou, Antonia

2013-01-01

65

Nontuberculous mycobacterial infections in Indian AIDS patients detected by a novel set of ESAT-6 polymerase chain reaction primers.  

Science.gov (United States)

Nontuberculous mycobacteria are often underdiagnosed due to lack of proper diagnostic facilities. To overcome this, we created a rapid PCR method for the species-specific diagnosis of Mycobacterium tuberculosis and its differentiation from other mycobacteria. A set of PCR primers targeting the gene encoding for early-secreted antigen-6 (ESAT-6) of the M. tuberculosis complex was designed and standardized on mycobacterial standard strains and on 75 recent isolates from AIDS patients and 70 isolates from HIV-negative patients seen at the hospital of the All India Institute of Medical Sciences, New Delhi, India. All 145 fresh mycobacterial isolates were identified using phenotypic methods and 16S rRNA PCR followed by sequencing of hypervariable region A. The ESAT-6 PCR detected all of the M. tuberculosis strains correctly (100% sensitivity), but none of the nontuberculous Mycobacterium spp. gave positive results (100% specific). Most nontuberculous mycobacteria were identified in patients with AIDS (24%) followed by those with tuberculous lymphadenitis (12.5%) and those with pulmonary tuberculosis whose treatment had failed (4.3%). The most common nontuberculous mycobacterial species isolated from AIDS patients was M. avium (6.6%), followed by M. fortuitum (5.7%), M. intracellulare and M. terrae (2.6% each). M. celatum, M. duvalii, M. austroafricanum, M. phlei and M. flavescence were also isolated from one patient each. The combination of genus-specific PCR primers with the novel ESAT-6 primer set could provide accurate and rapid diagnosis of mycobacteriosis. PMID:17314419

Singh, Sarman; Gopinath, Krishnamoorthy; Shahdad, Saba; Kaur, Manjot; Singh, Balwan; Sharma, Pawan

2007-02-01

66

Mycobacterial chaperonins: the tail wags the dog.  

Science.gov (United States)

Molecular chaperones are defined as proteins that assist the noncovalent assembly of other protein-containing structures in vivo, but which are not components of these structures when they are carrying out their normal biological functions. There are numerous families of protein that fit this definition of molecular chaperones, the most ubiquitous of which are the chaperonins and the Hsp70 families, both of which are required for the correct folding of nascent polypeptide chains and thus essential genes for cell viability. The groE genes of Escherichia coli were the first chaperonin genes to be discovered, within an operon comprising two genes, groEL and groES, that function together in the correct folding of nascent polypeptide chains. The identification of multiple groEL genes in mycobacteria, only one of which is operon-encoded with a groES gene, has led to debate about the functions of their encoded proteins, especially as the essential copies are surprisingly often not the operon-encoded genes. Comparisons of these protein sequences reveals a consistent functional homology and identifies an actinomycete-specific chaperonin family, which may chaperone the folding of enzymes involved in mycolic acid synthesis and thus provide a unique target for the development of a new class of broad-spectrum antimycobacterial drugs. PMID:24102684

Colaco, Camilo A; MacDougall, Alistair

2014-01-01

67

Highlight on Advances in Nontuberculous Mycobacterial Disease in North America  

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Nontuberculous mycobacteria (NTM) are ubiquitous in the environment and exist as an important cause of pulmonary infections in humans. Pulmonary involvement is the most common disease manifestation of NTM and the incidence of NTM is growing in North America. Susceptibility to NTM infection is incompletely understood; therefore preventative tools are not well defined. Treatment of pulmonary nontuberculous mycobacterial (NTM) infection is difficult and entails multiple antibiotics and an extended treatment course. Also, there is a considerable variation in treatment management that should be considered before initiating treatment. We highlight the new findings in the epidemiology diagnosis and treatment of mycobacterial infections. We debate new advances regarding NTM infection in cystic fibrosis patients and solid organ transplant recipients. Finally, we introduce a new epidemiologic model for NTM disease based on virulence-exposure-host factors.

Mirsaeidi, Mehdi; Farshidpour, Maham; Allen, Mary Beth; Ebrahimi, Golnaz; Falkinham, Joseph O.

2014-01-01

68

DNA encoding individual mycobacterial antigens protects mice against tuberculosis  

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Full Text Available Over the last few years, some of our experiments in which mycobacterial antigens were presented to the immune system as if they were viral antigens have had a significant impact on our understanding of protective immunity against tuberculosis. They have also markedly enhanced the prospects for new vaccines. We now know that individual mycobacterial protein antigens can confer protection equal to that from live BCG vaccine in mice. A critical determinant of the outcome of immunization appears to be the degree to which antigen-specific cytotoxic T cells are generated by the immune response. Our most recent studies indicate that DNA vaccination is an effective way to establish long-lasting cytotoxic T cell memory and protection against tuberculosis.

C.L. Silva

1999-02-01

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DNA encoding individual mycobacterial antigens protects mice against tuberculosis  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: English Abstract in english Over the last few years, some of our experiments in which mycobacterial antigens were presented to the immune system as if they were viral antigens have had a significant impact on our understanding of protective immunity against tuberculosis. They have also markedly enhanced the prospects for new v [...] accines. We now know that individual mycobacterial protein antigens can confer protection equal to that from live BCG vaccine in mice. A critical determinant of the outcome of immunization appears to be the degree to which antigen-specific cytotoxic T cells are generated by the immune response. Our most recent studies indicate that DNA vaccination is an effective way to establish long-lasting cytotoxic T cell memory and protection against tuberculosis.

C.L., Silva; V.L.D., Bonato; V.M.F., Lima.

1999-02-01

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Correlation between Serum and Plasma Antibody Titers to Mycobacterial Antigens ?  

OpenAIRE

The ability to utilize serum or plasma samples interchangeably is useful for tuberculosis (TB) serology. We demonstrate a strong correlation between antibody titers to several mycobacterial antigens in serum versus plasma from HIV-infected and non-HIV-infected TB and non-TB patients (r = 0.99 to 0.89; P < 0.0001). Plasma and serum can be used interchangeably in the same antibody detection assays.

Siev, Michael; Yu, Xian; Prados-rosales, Rafael; Martiniuk, Frank T.; Casadevall, Arturo; Achkar, Jacqueline M.

2010-01-01

71

Correlation between Serum and Plasma Antibody Titers to Mycobacterial Antigens ?  

Science.gov (United States)

The ability to utilize serum or plasma samples interchangeably is useful for tuberculosis (TB) serology. We demonstrate a strong correlation between antibody titers to several mycobacterial antigens in serum versus plasma from HIV-infected and non-HIV-infected TB and non-TB patients (r = 0.99 to 0.89; P < 0.0001). Plasma and serum can be used interchangeably in the same antibody detection assays. PMID:21047999

Siev, Michael; Yu, Xian; Prados-Rosales, Rafael; Martiniuk, Frank T.; Casadevall, Arturo; Achkar, Jacqueline M.

2011-01-01

72

Correlation between serum and plasma antibody titers to mycobacterial antigens.  

Science.gov (United States)

The ability to utilize serum or plasma samples interchangeably is useful for tuberculosis (TB) serology. We demonstrate a strong correlation between antibody titers to several mycobacterial antigens in serum versus plasma from HIV-infected and non-HIV-infected TB and non-TB patients (r = 0.99 to 0.89; P < 0.0001). Plasma and serum can be used interchangeably in the same antibody detection assays. PMID:21047999

Siev, Michael; Yu, Xian; Prados-Rosales, Rafael; Martiniuk, Frank T; Casadevall, Arturo; Achkar, Jacqueline M

2011-01-01

73

Induction of Mycobacterial Resistance to Quinolone Class Antimicrobials  

OpenAIRE

An agar plate assay was developed for detecting the induction of drug-resistant mycobacterial mutants during exposure to inhibitors of DNA gyrase. When Mycobacterium smegmatis on drug-containing agar, resistant colonies arose over a period of 2 weeks. A recA deficiency reduced mutant recovery, consistent with involvement of the SOS response in mutant induction. The C-8-methoxy compounds gatifloxacin and moxifloxacin allowed the recovery of fewer resistant mutants than either ciprofloxacin or ...

Malik, Muhammad; Chavda, Kalyan; Zhao, Xilin; Shah, Nirali; Hussain, Syed; Kurepina, Natalia; Kreiswirth, Barry N.; Kerns, Robert J.; Drlica, Karl

2012-01-01

74

Prospective study of mycobacterial infections in patients with cystic fibrosis.  

OpenAIRE

Fifty four patients with cystic fibrosis, aged 3-67 years, were studied prospectively for pulmonary mycobacterial infection. Sputum smears and cultures were carried out and intradermal skin tests performed. Mycobacteria were cultured from six patients in association with clinical deterioration; four patients had positive direct smears. Mycobacterium tuberculosis, M aviumintracellulare, M kansasii, and M gordonae were isolated. There were no deaths and all improved with chemotherapy. A third o...

Hjelte, L.; Petrini, B.; Ka?llenius, G.; Strandvik, B.

1990-01-01

75

Nontuberculous Mycobacterial Infection after Fractionated CO2 Laser Resurfacing  

OpenAIRE

Nontuberculous mycobacteria are increasingly associated with cutaneous infections after cosmetic procedures. Fractionated CO2 resurfacing, a widely used technique for photorejuvenation, has been associated with a more favorable side effect profile than alternative procedures. We describe 2 cases of nontuberculous mycobacterial infection after treatment with a fractionated CO2 laser at a private clinic. Densely distributed erythematous papules and pustules developed within the treated area wit...

Culton, Donna A.; Lachiewicz, Anne M.; Miller, Becky A.; Miller, Melissa B.; Mackuen, Courteney; Groben, Pamela; White, Becky; Cox, Gary M.; Stout, Jason E.

2013-01-01

76

Multiplex PCR assay for immediate identification of the infecting species in patients with mycobacterial disease.  

OpenAIRE

Rapid identification of infecting mycobacterial species enables appropriate medical care decisions to be made. Our aim was to demonstrate the clinical usefulness of the multiplex PCR assay, a test based on PCR, which permits direct identification of 12 mycobacterial species in clinical specimens. A total of 259 specimens from 177 patients who had clinical symptoms of mycobacterial disease but for whom there were difficulties in diagnosis were tested. Specimens were analyzed within 48 h of rec...

Kox, L. F. F.; Jansen, H. M.; Kuijper, S.; Kolk, A. H. J.

1997-01-01

77

Polyphenolic acetates : A newer anti-Mycobacterial therapeutic option  

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Full Text Available The objective of our research project was screening of various highly specific substrates of Acetoxy Drug: Protein Transacytylase (M.TAase for antimycobacterial activity. Mycobacterial culture was done in Middlebrook’s 7H9 media. Protein purification (Mycobacterial Tranacetylase, M.TAase was done by ion exchange chromatography and its demonstration was done on SDS- polyacrylamide gel electrophoresis (SDS-PAGE and western blot. Middlebrook’s 7H9 broth was procured from Becton Dickinson. CM-Sepharose, DEAE-Sepharose and Q-Sephharose were purchased from Amersham Pharmacia. Anti acetyl lysine polyclonal antibody was purchased from Cell Signaling. The Middlebrook 7H9 medium was used for M. smegmatis culture. The media was prepared according to the manufacturer’s instructions. The various Polyphenol acetate compounds were tested for their antimycobacterial activities. Minimal inhibitory concentrations (MIC were calculated by Alamar blue dye assay method. The GST protein was used as a receptor protein and purified Mycobacterial Glutamine Synthetase (GS as TAase for acetylation by DAMC. To demonstrate the TAase catalyzed acetylation of GST by DAMC, purified M.TAase (GS was preincubated with GST and DAMC followed by western blot using anti acetyl lysine antibody, which avidly react with the acetylated proteins. The growth pattern of M. smegmatis was diminished under the influence of various polyphenolic acetates (PA tested for their anti-mycobacterial activity. DAMC and DAMC-5-carboxylic acid was found to have MIC of 40?g/ml whereas DAMC-6-carboxylic acid was reported to have MIC value of 35?g/ml and for ellagic acid tetra acetate (EATA it was 60?g/ml. Previous work in our lab has led to discovery of a novel enzyme acetoxy drug: protein transacetylase (TAase, catalyzing transfer of acetyl group from various polyphenolic peracetate (PA to certain receptor proteins such as cytochromes P-450, NADPH cytochrome reductase, nitric oxide synthase (NOS has been established in various eukaryotic as well as prokaryotic sources. PA(s irreversible inhibitors of mammalian CYP linked MFO, possibly due to modification of cytochrome p- 450 by acetylation in a reaction catalysed by M.TAase (GS utilizing PA(s as a donor of acetyl groups. Accordingly, it was hypothesized that the CYP51 of mycobacteria involved in the cell wall sterol synthesis could possibly be modified by our PA(s through the novel unknown action of GS as transacetylase leading to the death of mycobacterial cell by way of acetylation catalyzed by acetoxy drug: protein transacetylase (M.TAase or GS.

Santram

2014-01-01

78

Assembly and proteolytic processing of mycobacterial ClpP1 and ClpP2  

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Full Text Available Abstract Background Caseinolytic proteases (ClpPs are barrel-shaped self-compartmentalized peptidases involved in eliminating damaged or short-lived regulatory proteins. The Mycobacterium tuberculosis (MTB genome contains two genes coding for putative ClpPs, ClpP1 and ClpP2 respectively, that are likely to play a role in the virulence of the bacterium. Results We report the first biochemical characterization of ClpP1 and ClpP2 peptidases from MTB. Both proteins were produced and purified in Escherichia coli. Use of fluorogenic model peptides of diverse specificities failed to show peptidase activity with recombinant mycobacterial ClpP1 or ClpP2. However, we found that ClpP1 had a proteolytic activity responsible for its own cleavage after the Arg8 residue and cleavage of ClpP2 after the Ala12 residue. In addition, we showed that the absence of any peptidase activity toward model peptides was not due to an obstruction of the entry pore by the N-terminal flexible extremity of the proteins, nor to an absolute requirement for the ClpX or ClpC ATPase complex. Finally, we also found that removing the putative propeptides of ClpP1 and ClpP2 did not result in cleavage of model peptides. We have also shown that recombinant ClpP1 and ClpP2 do not assemble in the conventional functional tetradecameric form but in lower order oligomeric species ranging from monomers to heptamers. The concomitant presence of both ClpP1 and ClpP2 did not result in tetradecameric assembly. Deleting the amino-terminal extremity of ClpP1 and ClpP2 (the putative propeptide or entry gate promoted the assembly in higher order oligomeric species, suggesting that the flexible N-terminal extremity of mycobacterial ClpPs participated in the destabilization of interaction between heptamers. Conclusion Despite the conservation of a Ser protease catalytic triad in their primary sequences, mycobacterial ClpP1 and ClpP2 do not have conventional peptidase activity toward peptide models and display an unusual mechanism of self-assembly. Therefore, the mechanism underlying their peptidase and proteolytic activities might differ from that of other ClpP proteolytic complexes.

Benaroudj Nadia

2011-12-01

79

Anti-Mycobacterial Activity of Marine Fungus-Derived 4-Deoxybostrycin and Nigrosporin  

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Full Text Available 4-Deoxybostrycin is a natural anthraquinone compound isolated from the Mangrove endophytic fungus Nigrospora sp. collected from the South China Sea. Nigrosporin is the deoxy-derivative of 4-deoxybostrycin. They were tested against mycobacteria, especially Mycobacterium tuberculosis. In the Kirby-Bauer disk diffusion susceptibility test, they both had inhibition zone sizes of over 25 mm. The results of the absolute concentration susceptibility test suggested that they had inhibitory effects against mycobacteria. Moreover, 4-deoxybostrycin exhibited good inhibition which was even better than that of first line anti-tuberculosis (TB drugs against some clinical multidrug-resistant (MDR M. tuberculosis strains. The gene expression profile of M. tuberculosis H37Rv after treatment with 4-deoxybostrycin was compared with untreated bacteria. One hundred and nineteen out of 3,875 genes were significantly different in M. tuberculosis exposed to 4-deoxybostrycin from control. There were 46 functionally known genes which are involved in metabolism, information storage and processing and cellular processes. The differential expressions of six genes were further confirmed by quantitative real-time polymerase chain reaction (qRT-PCR. The present study provides a useful experiment basis for exploitation of correlative new drugs against TB and for finding out new targets of anti-mycobacterial therapy.

Xiaomin Lai

2013-01-01

80

Cooling towers--a potential environmental source of slow-growing mycobacterial species.  

Science.gov (United States)

Over the last decade a rise in the frequency of disease caused by nontuberculous mycobacteria (NTM) has occurred, especially among AIDS patients. The lack of evidence for person-to-person transmission indicates the environment is a source of infection. The ecology and environmental sources of NTMs are poorly understood, and many pathogenic strains have not been observed outside of clinical cases. Several species of NTMs have been reported from treated water distribution systems; however, one type of manmade environment that has not been examined for mycobacteria is that of cooling towers of air-conditioning systems. Such environments not only harbor a variety of microbial species, they also disseminate them in aerosols. The present investigation examined nine cooling towers from various locations in the United States. Cooling tower water was concentrated, treated with cetylpyridinium chloride, and plated onto Middlebrook 7H10 agar supplemented with OADC and cycloheximide. Colonies presumed to be mycobacterial species were isolated and acid-fast stained. Identification was made by amplifying and sequencing 1450 bp fragments of the 16S rRNA gene in both directions, and comparing resulting sequences with those in GenBank. Results showed that at least 75% of tower samples contained NTMs, and most of the isolates closely matched known mycobacterial pathogens. Isolates most closely matched the following GenBank sequences: Mycobacterium intracellulare, M. szulgai, M. bohemicum, M. gordonae, M. nonchromogenicum, and M. n. sp. "Fuerth 1999." This is the first report of specific NTMs in cooling tower water, and the first report of M. n. sp. "Fuerth 1999" from any environmental sample. Although cooling towers have a relatively high pH, they may favor the growth and dissemination of such potential pathogens, and future epidemiologic investigations should consider cooling towers as possible environmental sources of mycobacteria. PMID:12688848

Black, Walter C; Berk, Sharon G

2003-01-01

81

Synthesis and evaluation of small libraries of triazolylmethoxy chalcones, flavanones and 2-aminopyrimidines as inhibitors of mycobacterial FAS-II and PknG.  

Science.gov (United States)

A synthetic strategy to access small libraries of triazolylmethoxy chalcones 4{1-20}, triazolylmethoxy flavanones 5{1-10} and triazolylmethoxy aminopyrimidines 6{1-17} from a common substrate 4-propargyloxy-2-hydroxy acetophenone using a set of different reactions has been developed. The chalcones and flavanones were screened against mycobacterial FAS-II pathway using a recombinant mycobacterial strain, against which the most potent compound showed ?88% inhibition in bacterial growth and substantially induction of reporter gene activity at 100 ?M concentration. The triazolylmethoxy aminopyrimdines were screened against PknG of Mycobaceterium tuberculosis displaying moderate to good activity (23-53% inhibition at 100 ?M), comparable to the action of a standard inhibitor. PMID:22854194

Anand, Namrata; Singh, Priyanka; Sharma, Anindra; Tiwari, Sameer; Singh, Vandana; Singh, Diwakar K; Srivastava, Kishore K; Singh, B N; Tripathi, Rama Pati

2012-09-01

82

Mendelian susceptibility to mycobacterial disease: Genetic, immunological, and clinical features of inborn errors of IFN-? immunity.  

Science.gov (United States)

Mendelian susceptibility to mycobacterial disease (MSMD) is a rare condition characterized by predisposition to clinical disease caused by weakly virulent mycobacteria, such as BCG vaccines and environmental mycobacteria, in otherwise healthy individuals with no overt abnormalities in routine hematological and immunological tests. MSMD designation does not recapitulate all the clinical features, as patients are also prone to salmonellosis, candidiasis and tuberculosis, and more rarely to infections with other intramacrophagic bacteria, fungi, or parasites, and even, perhaps, a few viruses. Since 1996, nine MSMD-causing genes, including seven autosomal (IFNGR1, IFNGR2, STAT1, IL12B, IL12RB1, ISG15, and IRF8) and two X-linked (NEMO, and CYBB) genes have been discovered. The high level of allelic heterogeneity has already led to the definition of 18 different disorders. The nine gene products are physiologically related, as all are involved in IFN-?-dependent immunity. These disorders impair the production of (IL12B, IL12RB1, IRF8, ISG15, NEMO) or the response to (IFNGR1, IFNGR2, STAT1, IRF8, CYBB) IFN-?. These defects account for only about half the known MSMD cases. Patients with MSMD-causing genetic defects may display other infectious diseases, or even remain asymptomatic. Most of these inborn errors do not show complete clinical penetrance for the case-definition phenotype of MSMD. We review here the genetic, immunological, and clinical features of patients with inborn errors of IFN-?-dependent immunity. PMID:25453225

Bustamante, Jacinta; Boisson-Dupuis, Stéphanie; Abel, Laurent; Casanova, Jean-Laurent

2014-10-26

83

The mycobacterial DNA-binding protein 1 (MDP1 from Mycobacterium bovis BCG influences various growth characteristics  

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Full Text Available Abstract Background Pathogenic mycobacteria such as M. tuberculosis, M. bovis or M. leprae are characterised by their extremely slow growth rate which plays an important role in mycobacterial virulence and eradication of the bacteria. Various limiting factors influence the generation time of mycobacteria, and the mycobacterial DNA-binding protein 1 (MDP1 has also been implicated in growth regulation. Our strategy to investigate the role of MDP1 in mycobacterial growth consisted in the generation and characterisation of a M. bovis BCG derivative expressing a MDP1-antisense gene. Results The expression rate of the MDP1 protein in the recombinant M. bovis BCG containing the MDP1-antisense plasmid was reduced by about 50% compared to the reference strain M. bovis BCG containing the empty vector. In comparison to this reference strain, the recombinant M. bovis BCG grew faster in broth culture and reached higher cell masses in stationary phase. Likewise its intracellular growth in mouse and human macrophages was ameliorated. Bacterial clumping in broth culture was reduced by the antisense plasmid. The antisense plasmid increased the susceptibility of the bacteria towards Ampicillin. 2-D protein gels of bacteria maintained under oxygen-poor conditions demonstrated a reduction in the number and the intensity of many protein spots in the antisense strain compared to the reference strain. Conclusion The MDP1 protein has a major impact on various growth characteristics of M. bovis BCG. It plays an important role in virulence-related traits such as aggregate formation and intracellular multiplication. Its impact on the protein expression in a low-oxygen atmosphere indicates a role in the adaptation to the hypoxic conditions present in the granuloma.

Maurischat Sven

2008-06-01

84

Partial overlap of anti-mycobacterial, and anti-Saccharomyces cerevisiae mannan antibodies in Crohns disease  

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Full Text Available AIM: To test whether humoral immune reaction against mycobacteria may play a role in anti-Saccharomyces cerevisiae antibodies (ASCA generation in Crohn's disease (CD and/or whether it correlates with clinical subtypes.METHODS: The dominant ASCA epitope was detected by Galanthus nivalis lectin (GNL-binding assay. ASCA and IgG against mycobacterial lysates [M avium, M smegmatis, M chelonae, M bovis BCG, M avium ssp. paratuberculosis (MAP] or purified lipoarabinomannans (LAM were detected by ELISA. ASCA and anti-mycobacterial antibodies were affinity purified to assess cross-reactivities. Anti-mycobacterial IgG were induced by BCG-infection of mice.RESULTS: GNL bound to different extents to mycobacterial lysates, abundantly to purified mannose-capped (Man LAM from M tuberculosis, but not to uncapped LAM from M smegmatis. Fifteen to 45% of CD patients but only 0%-6% of controls were seropositive against different mycobacterial antigens. Anti-mycobacterial IgG correlated with ASCA (r = 0.37-0.64; P = 0.003-P < 0.001. ASCA-positivity and deficiency for mannan-binding lectin synergistically associated with anti-mycobacterial IgG. In some patients, anti-mycobacterial antibodies represent cross-reactive ASCA. Vice-versa, the predominant fraction of ASCA did not cross-react with mycobacteria. Finally, fistulizing disease associated with antibodies against M avium, M smegmatis and MAP (P = 0.024, 0.004 and 0.045, respectively.CONCLUSION: Similar to ASCA, seroreactivity against mycobacteria may define CD patients with complicated disease and a predisposition for immune responses against ubiquitous antigens. While in some patients anti-mycobacterial antibodies strongly cross-react with yeast mannan; these cross-reactive antibodies only represent a minor fraction of total ASCA. Thus, mycobacterial infection unlikely plays a role in ASCA induction.

Stefan Müller, Thomas Schaffer, Alain M Schoepfer, Annamarie Hilty, Thomas Bodmer, Frank Seibold

2008-06-01

85

Respiratory review of 2014: tuberculosis and nontuberculous mycobacterial pulmonary disease.  

Science.gov (United States)

Since tuberculosis (TB) remains a major global health concern and the incidence of multi-drug resistant (MDR)-TB is increasing globally, new modalities for the detection of TB and drug resistant TB are needed to improve TB control. The Xpert MTB/RIF test can be a valuable new tool for early detection of TB and rifampicin resistance, with a high sensitivity and specificity. Late-generation fluoroquinolones, levofloxacin, and moxifloxacin, which are the principal drugs for the treatment of MDR-TB, show equally high efficacy and safety. Systemic steroids may reduce the overall TB mortality attributable to all forms of TB across all organ systems, although inhaled corticosteroids can increase the risk of TB development. Although fixed dose combinations were expected to reduce the risk of drug resistance and increase drug compliance, a recent meta-analysis found that they might actually increase the risk of relapse and treatment failure. Regarding treatment duration, patients with cavitation and culture positivity at 2 months of TB treatment may require more than 6 months of standard treatment. New anti-TB drugs, such as linezolid, bedaquiline, and delamanid, could improve the outcomes in drug-resistant TB. Nontuberculous mycobacterial lung disease has typical clinical and immunological phenotypes. Mycobacterial genotyping may predict disease progression, and whole genome sequencing may reveal the transmission of Mycobacterium abscessus. In refractory Mycobacterium avium complex lung disease, a moxifloxacin-containing regimen was expected to improve the treatment outcome. PMID:25368661

Park, Cheol Kyu; Kwon, Yong Soo

2014-10-01

86

A mycobacterial coinfection in a dog suspected on blood smear.  

Science.gov (United States)

A 4-year-old neutered female crossbred Shepherd was referred for a history of 10 days of anorexia, polyuria, polydipsia, polyadenomegaly, and diarrhea. On physical examination, the dog appeared quiet, responsive, and apyretic, with generalized and severe lymphadenomegaly. Hematologic abnormalities included neutrophilic leukocytosis with left shift, and lymphopenia. Blood smears revealed intracytoplasmic bacilli negatively stained with May-Grünwald-Giemsa in neutrophils and monocytes. Lymph node smears revealed pyogranulomatous adenitis with calcified deposits and many negative-staining rod structures, both within the cytoplasm of neutrophils and macrophages, and free in the background. An acid-fast stain (Ziehl-Neelsen) confirmed the diagnosis of mycobacterial infection. The dog was euthanized for public health and ethical reasons, and the postmortem examination revealed severe and generalized granulomatous and necrotizing lymphadenitis, panniculitis, and hepatitis, and infiltration of epithelioid macrophages in the lungs, colon, and spleen. Numerous acid-fast bacilli, consistent with mycobacterial infection, were observed both in the cytoplasm of epithelioid macrophages and giant cells, and free in the background. Mycobacterium bovis was first confirmed by conventional PCR of organ extracts. Mycobacterium avium was detected in a culture of the same organs. Further PCR amplifications and sequencing revealed a coinfection with 2 different species of mycobacterium, one belonging to the Mycobacterium avium complex and the other to the Mycobacterium tuberculosis complex. PMID:24320783

Etienne, Claire-Lise; Granat, Fanny; Trumel, Catherine; Raymond-Letron, Isabelle; Lucas, Marie-Noëlle; Boucraut-Baralon, Corine; Pingret, Jean-Luc; Magne, Laurent; Delverdier, Maxence

2013-12-01

87

A Pseudokinase Debut at the Mycobacterial Cell Wall  

Science.gov (United States)

Mycobacterium tuberculosis, the causative agent of tuberculosis, has a complex cellular envelope that comprises both the cytoplasmic membrane and the outer cell wall. Despite advances in elucidating the structural and biochemical composition of these features, the processes that ensure cell wall homeostasis remain poorly understood. New findings implicate the essential mycobacterial serine-threonine protein kinase (STPK), PknB, in regulating the formation of a regulatory complex that includes the integral membrane protein MviN, which is required for peptidoglycan biosynthesis, and a forkhead-associated (FHA) domain protein, FhaA. A model has emerged in which a peptidoglycan-derived muropeptide signal triggers the PknB-mediated phosphorylation of the MviN pseudokinase domain, which in turn recruits the FHA-containing regulatory protein to inhibit peptidoglycan biosynthesis at the cell poles and septum. In establishing PknB as central regulator of this pathway, the model reinforces the major role of this STPK network in the orchestration of fundamental mycobacterial processes, and, with the identification of MviN as having a catalytically inactive and highly divergent kinase homology domain, the model establishes a pseudokinase as a key player in cell wall metabolism.

Digby F. Warner (University of Cape Town; Faculty of Health Sciences REV)

2012-01-24

88

Contribution of the multidrug efflux pump LfrA to innate mycobacterial drug resistance.  

Science.gov (United States)

Multidrug resistance (MDR) in bacteria has been associated with efflux pumps that export structurally unrelated compounds and decrease cytoplasmic drug accumulation. To investigate MDR in mycobacteria, we studied the Mycobacterium smegmatis mutant mc(2)11, which is resistant to doxorubicin, tetracycline, rhodamine, ethidium bromide and the hydrophilic fluoroquinolones. A genomic library constructed from this mutant was used to select clones conferring resistance to doxorubicin. Surprisingly, the clone selected encodes the efflux pump LfrA, which has been reported to confer resistance to hydrophilic fluoroquinolones, ethidium bromide, rhodamine, and acriflavine. To define the contribution of LfrA to the innate mycobacterial drug resistance and to the MDR phenotype in mc(2)11, the lfrA gene was disrupted in both the mc(2)11 mutant and the mc(2)155 wild-type parent. LfrA disruption of the wild-type strain decreased resistance to ethidium bromide and acriflavine, and increased accumulation of ethidium bromide. However, disruption of lfrA gene results only in a 2-fold decrease in minimal inhibitory concentrations (MICs) for ciprofloxacin, doxorubicin, rhodamine, and accumulation of [(14)C]ciprofloxacin was unchanged. LfrA disruption of the MDR strain mc(2)11 produced a similar phenotype. Thus, LfrA contributes significantly to the intrinsic MICs of M. smegmatis for ethidium bromide and acriflavine, but not for ciprofloxacin, doxorubicin or rhodamine. PMID:11094273

Sander, P; De Rossi, E; Böddinghaus, B; Cantoni, R; Branzoni, M; Böttger, E C; Takiff, H; Rodriquez, R; Lopez, G; Riccardi, G

2000-12-01

89

Effect of silver-carrying photocatalyst "Hikari-Gintech" on mycobacterial growth in vitro.  

Science.gov (United States)

The antimycobacterial activity of "Hikari-Gintech" powder, which has photocatalytic activity, was examined in vitro. Both powder dissolved in liquid and Hikari-Gintech-coated cloths showed strong antimycobacterial activity against Mycobacterium tuberculosis H37Rv, M. bovis BCG Pasteur, multi-drug-resistant M. tuberculosis (a clinical isolate) and M. avium. Hikari-Gintech powder appeared to affect mycobacterial cell wall metabolism rather than mycobacterial DNA because no damage to mycobacterial DNA was detected after spraying with Hikari-Gintech solution. PMID:15272193

Matsui, Yoshifumi; Otomo, Koji; Ishida, Shinichiro; Yanagihara, Kazuo; Kawanobe, Yasutaka; Kida, Shoji; Taruoka, Eiichi; Sugawara, Isamu

2004-01-01

90

Diagnosis and management of atypical mycobacterial infection after laparoscopic surgery.  

Science.gov (United States)

Atypical mycobacterial infections at the laparoscopic port site are a frequent problem encountered in patients undergoing laparoscopic surgery. In this study we concentrate on the clinical diagnosis, management and prevention of this problem. In this series we assess 19 patients presenting with port hole infections after laparoscopic surgery and were treated with a combination of oral clarithromycin and ciprofloxacin. Seven patients who had persistent nodules were given injections of amikacin directly into the infection foci along with standard oral therapy. Most of the patients treated with standard oral therapy for 28 days showed recovery. The patients with persistent nodules 4 weeks after completion of therapy were treated with injections of amikacin directly into the nodule which lead to resolution of symptoms. For prevention of infection, proper sterilization and storage of instruments is recommended. Laparoscopic port hole infections is a preventable problem and can also be treated by nonsurgical method. PMID:22131651

Chaudhuri, Sumit; Sarkar, Debojyoti; Mukerji, Reshmi

2010-12-01

91

Mycobacterium abscessus: a new player in the mycobacterial field.  

Science.gov (United States)

Mycobacterium abscessus, a relative of Koch's bacillus (the bacterium that causes tuberculosis), has recently emerged as the cause of an increasing number of both community- and hospital-acquired infections in humans; it also constitutes a serious threat for cystic fibrosis patients. This situation is worsened by its exceptionally high natural and acquired antibiotic resistance that complicates treatment. Although a rapid grower, it shares some traits with Koch's bacillus, including the ability to induce a persistent lung disease associated with caseous lesions, a landmark of Mycobacterium tuberculosis infection. Its genome sequence and microarrays are now available, and efficient genetic tools have recently been developed. Here we consider the various advantages of using this species as an experimental model to study tuberculosis and other related mycobacterial diseases. PMID:20060723

Medjahed, Halima; Gaillard, Jean-Louis; Reyrat, Jean-Marc

2010-03-01

92

Imaging of tuberculous and non-tuberculous mycobacterial infections  

International Nuclear Information System (INIS)

Full text: Both tuberculosis (TB) and non-tuberculous mycobacterial (NTM) infections are increasing in prevalence and radiologists must be aware of the increasing likelihood of encountering these infections. The NTM organisms are often overlooked as potential infecting organisms, diagnosis is often delayed as they tend not to be included in the radiologic differential diagnosis. Clinical and radiological manifestations of pulmonary TB depend on whether the host is naive to the infecting organism (primary TB) or whether there has been reactivation or reexposure (postprimary TB). In immunocompetent patients primary TB tends to be self-limiting, but resolution of the primary lesion is usually slow, and often results in permanent scarring. Postprimary TB lesions (reactivation of a dormant primary infection) tend to be focal, nodular, with a slow progressive course resulting in high morbidity and mortality if not adequately treated. The most common atypical mycobacterial infection is caused by Mycobacterium avium intracellulare complex. Two target groups are mainly encountered in NTM infections: 1) males more than 50 years old with preexisting lung disease presenting constitutional symptoms and 2) elderly women with no preexisting pulmonary disease with less pronounced constitutional symptoms. NTM infection is usually indolent, sometimes remaining stable for a long period; however, without treatment it may progress and ultimately be fatal. CT findings of primary tuberculose fatal. CT findings of primary tuberculosis consist of consolidation mainly in middle and lower lobes and lymphadenopathy with hypodense centre and rim enhancement on contrast-enhanced CT. Tree-in bud appearance adjacent to areas of consolidation is consistent with endobronchial spread. Cavitary lesions are less common than in primary.

93

Novel STAT1 Alleles in Otherwise Healthy Patients with Mycobacterial Disease  

OpenAIRE

The transcription factor signal transducer and activator of transcription-1 (STAT1) plays a key role in immunity against mycobacterial and viral infections. Here, we characterize three human STAT1 germline alleles from otherwise healthy patients with mycobacterial disease. The previously reported L706S, like the novel Q463H and E320Q alleles, are intrinsically deleterious for both interferon gamma (IFNG)-induced gamma-activating factor-mediated immunity and interferon alpha (IFNA)-induced int...

Chapgier, A.; Boisson-dupuis, S.; Jouanguy, E.; Vogt, G.; Feinberg, J.; Prochnicka-chalufour, A.; Casrouge, A.; Yang, K.; Soudais, C.; Fieschi, C.; Santos, O. F.; Bustamante, J.; Picard, C.; Beaucoudrey, L.; Emile, J. F.

2006-01-01

94

Mycobacterial Infection of the Gallbladder Masquerading as Gallbladder Cancer with a False Positive Pet Scan  

OpenAIRE

Isolated mycobacterial infection of gall bladder is an extremely rare entity. Only anecdotal reports are evident in the literature. A preoperative diagnosis of mycobacterial infection of gallbladder is therefore very difficult. The case of a 72-year-old male who underwent surgery for suspected gallbladder cancer is presented. The diagnosis of cancer was based on radiological findings and an abnormal uptake of fluorine-18-fluoro-2-deoxy-D-glucose (FDG) on positron emission tomography (PET) sca...

Adeeb Majid; Ravish Sanghi Raju; Markus Trochsler; Kanhere, Harsh A.; Maddern, Guy J.

2013-01-01

95

Imbalanced effector and regulatory cytokine responses may underlie mycobacterial immune restoration disease  

OpenAIRE

Abstract Background Immune restoration disease (IRD) is an adverse consequence of antiretroviral therapy, where the restored pathogen-specific response causes immunopathology. Mycobacteria are the pathogens that most frequently provoke IRD and mycobacterial IRD is a common cause of morbidity in HIV-infected patients co-infected with mycobacteria. We hypothesised that the excessive effector immune response in mycobacterial IRD reflects impaired regulation by IL-10. Resu...

Price Patricia; Orsogna Lloyd, D.; Lim Andrew; French Martyn A

2008-01-01

96

Molecular-based mycobacterial identification in a clinical laboratory setting: a comparison of two methods.  

LENUS (Irish Health Repository)

Many mycobacterial species are pathogenic to humans, with infection occurring worldwide. Infection with Mycobacterium tuberculosis is a well-described global phenomenon, but other mycobacterial species are increasingly shown to be the cause of both pulmonary and extrapulmonary infection and are managed differently from M. tuberculosis infection. Rapid and accurate differentiation of mycobacterial species is, therefore, critical to guide timely and appropriate therapeutic and public health management. This study evaluates two commercially available DNA strip assays, the Genotype Common Mycobacteria (CM) assay (Hain Lifescience, Nehren, Germany) and the Speed-oligo Mycobacteria assay (Vircell, Spain) for their usefulness in a clinical laboratory setting. Both assays were evaluated on 71 clinical mycobacterial isolates, previously identified using Gen-Probe AccuProbe and through a UK mycobacteriology reference laboratory, as well as 29 non-mycobacterial isolates. Concordant results were obtained for 98% of isolates using both assays. The sensitivity was 97% (95% confidence interval [CI]: 93.3-100%) for the CM assay and 98.6% (95% CI: 95.9-100%) for the Speed-oligo assay. Overall, both assays proved to be useful tools for rapid and sensitive mycobacterial species identification, although interpretation of results was easier with the CM assay. Finally, results were available within one day, compared to current identification times which range between seven days and four weeks.

O'Donnell, N

2012-01-01

97

Phagocyte NADPH oxidase, chronic granulomatous disease and mycobacterial infections.  

Science.gov (United States)

Infection of humans with Mycobacterium tuberculosis remains frequent and may still lead to death. After primary infection, the immune system is often able to control M. tuberculosis infection over a prolonged latency period, but a decrease in immune function (from HIV to immunosenescence) leads to active disease. Available vaccines against tuberculosis are restricted to BCG, a live vaccine with an attenuated strain of M. bovis. Immunodeficiency may not only be associated with an increased risk of tuberculosis, but also with local or disseminated BCG infection. Genetic deficiency in the reactive oxygen species (ROS)-producing phagocyte NADPH oxidase NOX2 is called chronic granulomatous disease (CGD). CGD is among the most common primary immune deficiencies. Here we review our knowledge on the importance of NOX2-derived ROS in mycobacterial infection. A literature review suggests that human CGD patient frequently have an increased susceptibility to BCG and to M. tuberculosis. In vitro studies and experiments with CGD mice are incomplete and yielded - at least in part - contradictory results. Thus, although observations in human CGD patients leave little doubt about the role of NOX2 in the control of mycobacteria, further studies will be necessary to unequivocally define and understand the role of ROS. PMID:24916152

Deffert, Christine; Cachat, Julien; Krause, Karl-Heinz

2014-08-01

98

Anti-mycobacterial activity of 1,3-diaryltriazenes.  

Science.gov (United States)

The rapid generation and spread of the drug resistant tuberculosis has led to an ongoing demand for novel compounds for therapeutic use. Identification and study of compounds with the ability to inhibit Mycobacterium tuberculosis is of paramount importance. For this reason, a library of substituted 1,3-diaryltriazenes based on the acting component of the anti-trypanosomal drug, diminazene aceturate was created and evaluated for its potential as anti-tubercular agent. Several compounds were identified with sub-micro molar inhibitory concentrations against M. tuberculosis and other clinically relevant mycobacterial species such as Mycobacterium bovis, Mycobacterium avium and Mycobacterium ulcerans. Although the library of the compounds showed a considerable acute cytotoxicity, a genotoxicity could not be observed. Finally, the triazene 14 was selected with the best biological properties (IC50 = 3.26 ?M, NI50 = 24.22 ?M, SI = 7.44). The compound 14 showed the ability to inhibit the growth of intracellular replicating and multi-drug resistant M. tuberculosis. The results suggest the molecule to be an interesting scaffold for further study and optimization. PMID:24631899

Cappoen, Davie; Vajs, Jure; Uythethofken, Cynthia; Virag, Andrej; Mathys, Vanessa; Ko?evar, Marijan; Verschaeve, Luc; Gazvoda, Martin; Polanc, Slovenko; Huygen, Kris; Košmrlj, Janez

2014-04-22

99

Ultrastructural morphologic changes in mycobacterial biofilm in different extreme condition.  

Science.gov (United States)

Abstract The aim of this study was to investigate the morphologic and ultrastructural features of biofilms of slow and fast-growing mycobacteria in different stress conditions, presence and absence of oleic acid albumin dextrose catalase (OADC) enrichment and at different temperatures: 30, 37 and 42?°C. Four hundred mycobacterial isolates were taken. The biomass of each biofilm was quantified using a modified microtiter plate assay method. Isolates were divided into those that formed fully established biofilms, moderately attached biofilms and weakly adherent biofilms by comparison with a known biofilm-forming strain. The large quantity of biofilm was produced by Mycobacterium smegmatis at temperature 37 and 42?°C as compared to 30?°C. Mycobacterium fortuitum and M. avium developed large amount of biofilm at 30?°C as compared to 37 and 42?°C. Mycobacterium tuberculosis developed strong biofilm at 37?°C and no biofilm at 30 and 42?°C in Sauton's media. The selected non-tuberculous mycobacteria and H37Rv developed strong biofilm in the presence of OADC enrichment in Sauton's medium. Microscopic examination of biofilms by scanning electron microscopy revealed that poorly adherent biofilm formers failed to colonize the entire surface of the microtiter well. While moderately adherent biofilm formers grew in uniform monolayers but failed to develop a mature three-dimensional structure. SEM analysis of an isolate representative of the group formed fully established biofilms with a textured, multi-layered, three-dimensional structure. PMID:25192360

Kumar, Virendra; Sachan, Tarun Kumar; Sharma, Pragya; Rawat, Krishna Dutta

2015-02-01

100

Inhibition of mycobacterial alanine racemase activity and growth by thiadiazolidinones.  

Science.gov (United States)

The genus Mycobacterium includes non-pathogenic species such as M. smegmatis, and pathogenic species such as M. tuberculosis, the causative agent of tuberculosis (TB). Treatment of TB requires a lengthy regimen of several antibiotics, whose effectiveness has been compromised by the emergence of resistant strains. New antibiotics that can shorten the treatment course and those that have not been compromised by bacterial resistance are needed. In this study, we report that thiadiazolidinones, a relatively little-studied heterocyclic class, inhibit the activity of mycobacterial alanine racemase, an essential enzyme that converts l-alanine to d-alanine for peptidoglycan synthesis. Twelve members of the thiadiazolidinone family were evaluated for inhibition of M. tuberculosis and M. smegmatis alanine racemase activity and bacterial growth. Thiadiazolidinones inhibited M. tuberculosis and M. smegmatis alanine racemases to different extents with 50% inhibitory concentrations (IC50) ranging from 150?M, respectively. The compounds also inhibited the growth of these bacteria, including multidrug resistant strains of M. tuberculosis. The minimal inhibitory concentrations (MIC) for drug-susceptible M. tuberculosis and M. smegmatis ranged from 6.25?g/ml to 100?g/ml, and from 1.56 to 6.25?g/ml for drug-resistant M. tuberculosis. The in vitro activities of thiadiazolidinones suggest that this family of compounds might represent starting points for medicinal chemistry efforts aimed at developing novel antimycobacterial agents. PMID:23680030

Lee, Yashang; Mootien, Sara; Shoen, Carolyn; Destefano, Michelle; Cirillo, Pier; Asojo, Oluwatoyin A; Yeung, Kacheong R; Ledizet, Michel; Cynamon, Michael H; Aristoff, Paul A; Koski, Raymond A; Kaplan, Paul A; Anthony, Karen G

2013-07-15

101

Non-major histocompatibility complex-restricted cytotoxic activity of blood mononuclear cells stimulated with secreted mycobacterial proteins and other mycobacterial antigens  

DEFF Research Database (Denmark)

Several observations indicate that non-major histocompatibility complex (MHC)-restricted cytotoxicity, mediated for example by natural killer cells and lymphokine-activated killer cells, may serve as an important antimicrobial defense mechanism. The purpose of the present study was to investigate the influences of different mycobacterial antigens on non-MHC-restricted cytotoxicity and further to investigate the ways by which various lymphocyte subpopulations contribute to the development of this cytotoxicity. Non-MHC-restricted cytotoxicity was induced following stimulation of mononuclear cells with tuberculin purified protein derivative, Mycobacterium bovis bacillus Calmette-Guérin (BCG), short- and long-term culture filtrates of virulent Mycobacterium tuberculosis H37Rv, and 30-31-kDa secreted mycobacterial protein. These antigens also induced proliferation and production of gamma interferon. The CD4+ cells proliferated and expressed interleukin-2 receptors following stimulation with mycobacterial antigens. Depletion studies after antigen stimulation showed that the cytotoxic effector cells were CD16+ CD56+ and CD4-; the CD4+ cells alone did not mediate non-MHC-restricted cytotoxicity. To evaluate the influence of CD4+ cells on the development of non-MHC-restricted cytotoxicity, blood mononuclear cells were depleted of CD4+ cells before antigen stimulation. When mononuclear cells were incubated with purified protein derivative or short-term culture filtrate in the absence of CD4+ cells, cytotoxic activity was reduced. This reduction was abolished by interleukin-2 but not by gamma interferon. We conclude that several mycobacterial antigens are able to induce non-MHC-restricted cytotoxicity. This study indicates that non-MHC-restricted cytotoxicity following stimulation with mycobacterial antigens is induced by cytokines released by antigen-specific activated CD4+ cells.

Ravn, P; Pedersen, B K

1994-01-01

102

Synthesis and selection of de novo proteins that bind and impede cellular functions of an essential mycobacterial protein.  

Science.gov (United States)

Recent advances in nonrational and part-rational approaches to de novo peptide/protein design have shown increasing potential for development of novel peptides and proteins of therapeutic use. We demonstrated earlier the usefulness of one such approach recently developed by us, called "codon shuffling," in creating stand-alone de novo protein libraries from which bioactive proteins could be isolated. Here, we report the synthesis and selection of codon-shuffled de novo proteins that bind to a selected Mycobacterium tuberculosis protein target, the histone-like protein HupB, believed to be essential for mycobacterial growth. Using a versatile bacterial two-hybrid system that entailed utilization of HupB and various codon-shuffled protein libraries as bait and prey, respectively, we were able to identify proteins that bound strongly to HupB. The observed interaction was also confirmed using an in vitro assay. One of the protein binders was expressed in Mycobacterium smegmatis and was shown to appreciably affect growth in the exponential phase, a period wherein HupB is selectively expressed. Furthermore, the transcription profile of hupB gene showed a significant reduction in the transcript quantity in mycobacterial strains expressing the protein binder. Electron microscopy of the affected mycobacteria elaborated on the extent of cell damage and hinted towards a cell division malfunction. It is our belief that a closer inspection of the obtained de novo proteins may bring about the generation of small-molecule analogs, peptidomimetics, or indeed the proteins themselves as realistic leads for drug candidates. Furthermore, our strategy is adaptable for large-scale targeting of the essential protein pool of Mycobacterium tuberculosis and other pathogens. PMID:17189438

Rao, Alka; Ram, Geeta; Saini, Adesh Kumar; Vohra, Reena; Kumar, Krishan; Singh, Yogendra; Ranganathan, Anand

2007-02-01

103

Isolation by genetic labeling of a new mycobacterial plasmid, pJAZ38, from Mycobacterium fortuitum.  

Science.gov (United States)

In a two-step mating experiment with recipient strains of Mycobacterium smegmatis, the Mycobacterium fortuitum cryptic plasmid pJAZ38 was isolated. Plasmid pJAZ38 was genetically labeled by cointegration formation mediated by the kanamycin-resistant mycobacterial transposon Tn611. The region responsible for replication of pJAZ38 was located and sequenced. This region showed homology with the Mycobacterium avium plasmid pLR7 and the Mycobacterium scrofulaceum plasmid pMSC262, a family of plasmids which have been found to be widespread throughout the mycobacteria. Further experiments showed pJAZ38 to be stably inherited in the absence of selection pressure and compatible with the most commonly used mycobacterial replicon, pAL5000. In contrast to pLR7 and pMSC262, pJAZ38 was able to replicate in M. smegmatis mc(2)155, making it a useful tool for mycobacterial genetics. PMID:9209023

Gavigan, J A; Aínsa, J A; Pérez, E; Otal, I; Martín, C

1997-07-01

104

Mycobacterial infection in a fairy bluebird (Irena ract puella): a diagnostic conundrum.  

Science.gov (United States)

An adult male, wild-caught fairy bluebird (Irena puella) was evaluated after diagnosis of hepatic mycobacterial disease in a bird sharing the same quarantine space. Initial results did not reveal leukocytosis or acid-fast organisms in a liver biopsy. However, Mycobacterium avium was found in the liver via polymerase chain reaction (PCR). After euthanasia, acid-fast stains remained negative in the liver, although PCR was positive and M. avium complex (identified by high-performance liquid chromatography) was isolated from the liver. PCR could offer a relatively sensitive and rapid diagnostic test in the investigation of mycobacterial disease in avian patients. PMID:19368260

Robbins, P K; Terrell, Scott P; Bradway, Dan; Wier, Fonda

2009-03-01

105

Activation of T cells recognizing self 60-kD heat shock protein can protect against experimental arthritis  

OpenAIRE

Lewis rats are susceptible to several forms of experimental arthritis- induced using heat-killed Mycobacterium tuberculosis (adjuvant arthritis, or AA), streptococcal cell walls, collagen type II, and the lipoidal amine CP20961. Prior immunization with the mycobacterial 65-kD heat shock protein (hsp65) was reported to protect against AA, and other athritis models not using M. tuberculosis, via a T cell-mediated mechanism. Hsp65 shares 48% amino acid identity with mammalian hsp60, which is exp...

1995-01-01

106

Immunology of atherosclerosis. Demonstration of heat shock protein 60 expression and T lymphocytes bearing alpha/beta or gamma/delta receptor in human atherosclerotic lesions.  

OpenAIRE

Our previous work revealed the presence of a great number of activated T lymphocytes in early human atherosclerotic lesions, and we were able to induce atherosclerosis in normocholesterolemic rabbits by immunization with Mycobacterium tuberculosis heat-shock protein (HSP) 65. We hypothesized this latter phenomenon to arise from cross-reactivity of mycobacterial HSP 65 with the endogenously expressed homologous 60-kd form of this stress protein. To study HSP 60 expression and the phenotype of ...

Kleindienst, R.; Xu, Q.; Willeit, J.; Waldenberger, F. R.; Weimann, S.; Wick, G.

1993-01-01

107

Lack of Adherence to Evidence-based Treatment Guidelines for Nontuberculous Mycobacterial Lung Disease  

OpenAIRE

Rationale: The 2007 American Thoracic Society (ATS) and Infectious Diseases Society of America (IDSA) recommend that patients with pulmonary nontuberculous mycobacterial (PNTM) disease caused by Mycobacterium avium complex (MAC) or M. abscessus be treated with a macrolide-based multidrug antibiotic regimen until sputum culture negative for 1 year. After 6 years, the degree of adherence to recommended guidelines among physicians remains unknown.

Adjemian, Jennifer; Prevots, D. Rebecca; Gallagher, Jack; Heap, Kylee; Gupta, Renu; Griffith, David

2014-01-01

108

Rapid assay for mycobacterial growth and antibiotic susceptibility using gel microdrop encapsulation.  

OpenAIRE

Effective control of tuberculosis transmission in vulnerable population groups is dependent on rapid identification of the infectious agent and its drug susceptibility. However, the slow growth rate of mycobacteria has undermined the ability to quickly identify antimicrobial resistance. These studies describe a mycobacterial growth assay based on microencapsulation technology used in conjunction with flow cytometric analysis. Mycobacteria were encapsulated in agarose gel microdrops approximat...

Ryan, C.; Nguyen, B. T.; Sullivan, S. J.

1995-01-01

109

Evaluation of Oral Antiseptic Rinsing before Sputum Collection To Reduce Contamination of Mycobacterial Cultures?  

Science.gov (United States)

To assess whether rinsing with oral antiseptics before sputum collection would reduce contamination of mycobacterial cultures, 120 patients with suspected tuberculosis were randomly assigned to rinse with chlorhexidine or cetylpyridinium mouthwash before collection. The culture contamination rate was significantly lower after rinsing with chlorhexidine before collection, especially for cultures grown in MGIT medium. PMID:21677070

Peres, Renata L.; Palaci, Moisés; Loureiro, Rafaela B.; Dietze, Reynaldo; Johnson, John L.; Golub, Jonathan E.; Ruffino-Netto, A.; Maciel, Ethel L.

2011-01-01

110

Molecular and Physiological Effects of Mycobacterial oxyR Inactivation  

OpenAIRE

The majority of slow-growing mycobacteria have a functional oxyR, the central regulator of the bacterial oxidative stress response. In contrast, this gene has been inactivated during the evolution of Mycobacterium tuberculosis. Here we inactivated the oxyR gene in Mycobacterium marinum, an organism used to model M. tuberculosis pathogenesis. Inactivation of oxyR abrogated induction of ahpC, a gene encoding alkylhydroperoxide reductase, normally activated upon peroxide challenge. The absence o...

Paga?n-ramos, Eileen; Master, Sharon S.; Pritchett, Christopher L.; Reimschuessel, Renate; Trucksis, Michele; Timmins, Graham S.; Deretic, Vojo

2006-01-01

111

Evaluation of the LiPA MYCOBACTERIA Assay for Identification of Mycobacterial Species from BACTEC 12B Bottles  

OpenAIRE

The LiPA MYCOBACTERIA (Innogenetics NV, Ghent, Belgium) assay was used to identify mycobacterial isolates using culture fluid from positive BACTEC 12B bottles. The LiPA method involves reverse hybridization of a biotinylated mycobacterial PCR fragment, a 16 to 23S rRNA spacer region, to oligonucleotide probes arranged in lines on a membrane strip, with detection via biotin-streptavidin coupling by a colorimetric system. This system identifies Mycobacterium species and differentiates M. tuberc...

Miller, Nancimae; Infante, Susanna; Cleary, Tim

2000-01-01

112

Biochemical and Structural Characterization of Mycobacterial Aspartyl-tRNA Synthetase AspS, a Promising TB Drug Target  

Science.gov (United States)

The human pathogen Mycobacterium tuberculosis is the causative agent of pulmonary tuberculosis (TB), a disease with high worldwide mortality rates. Current treatment programs are under significant threat from multi-drug and extensively-drug resistant strains of M. tuberculosis, and it is essential to identify new inhibitors and their targets. We generated spontaneous resistant mutants in Mycobacterium bovis BCG in the presence of 10× the minimum inhibitory concentration (MIC) of compound 1, a previously identified potent inhibitor of mycobacterial growth in culture. Whole genome sequencing of two resistant mutants revealed in one case a single nucleotide polymorphism in the gene aspS at 535GAC>535AAC (D179N), while in the second mutant a single nucleotide polymorphism was identified upstream of the aspS promoter region. We probed whole cell target engagement by overexpressing either M. bovis BCG aspS or Mycobacterium smegmatis aspS, which resulted in a ten-fold and greater than ten-fold increase, respectively, of the MIC against compound 1. To analyse the impact of inhibitor 1 on M. tuberculosis AspS (Mt-AspS) activity we over-expressed, purified and characterised the kinetics of this enzyme using a robust tRNA-independent assay adapted to a high-throughput screening format. Finally, to aid hit-to-lead optimization, the crystal structure of apo M. smegmatis AspS was determined to a resolution of 2.4 Å. PMID:25409504

Cox, Jonathan A. G.; Fütterer, Klaus; Abrahams, Katherine A.; Bhatt, Apoorva; Alderwick, Luke J.; Reynolds, Robert C.; Loman, Nicholas J.; Nataraj, VijayaShankar; Alemparte, Carlos; Barros, David; Lloyd, Adrian J.; Ballell, Lluis; Hobrath, Judith V.; Besra, Gurdyal S.

2014-01-01

113

Platelets Direct Monocyte Differentiation Into Epithelioid-Like Multinucleated Giant Foam Cells With Suppressive Capacity Upon Mycobacterial Stimulation  

Science.gov (United States)

Background.?Epithelioid, foam, and multinucleated giant cells (MNGCs) are characteristics of tuberculosis granulomas, yet the precise genesis and functions of these transformed macrophages are unclear. We evaluated the role of platelets as drivers of macrophage transformation in mycobacterial infection. Methods.?We employed flow cytometry and microscopy to assess cellular phenotype and phagocytosis. Immune assays allowed quantification of cytokines and chemokines, whereas gene microarray technology was applied to estimate global transcriptome alterations. Immunohistochemical investigations of tuberculosis granulomas substantiated our findings at the site of infection. Results.?Monocytes differentiated in presence of platelets (MP-Macs) acquired a foamy, epithelioid appearance and gave rise to MNGCs (MP-MNGCs). MP-Macs up-regulated activation markers, phagocytosed mycobacteria, and released abundant interleukin 10. Upon extended culture, MP-Macs shared transcriptional features with epithelioid cells and M2 macrophages and up-regulated CXCL5 transcripts. In line with this, CXCL5 concentrations were significantly increased in airways of active tuberculosis patients. The platelet-specific CD42b antigen was detected in MP-Macs, likewise in macrophages, MNGCs, and epithelioid cells within tuberculosis granulomas, along with the platelet aggregation-inducing factor PDPN. Conclusions.?Platelets drive macrophage differentiation into MNGCs with characteristics of epithelioid, foam, and giant cells observed in tuberculosis granulomas. Our data define platelets as novel participants in tuberculosis pathogenesis. PMID:24987031

Feng, Yonghong; Dorhoi, Anca; Mollenkopf, Hans-Joachim; Yin, Hongyun; Dong, Zhengwei; Mao, Ling; Zhou, Jun; Bi, Aixiao; Weber, Stephan; Maertzdorf, Jeroen; Chen, Gang; Chen, Yang; Kaufmann, Stefan H. E.

2014-01-01

114

Under-explored experimental topics related to integral mycobacterial vaccines for leprosy.  

Science.gov (United States)

Many leprosy vaccine studies have utilized live or killed whole mycobacteria, such as Bacille Calmette-Guérin, Indian Cancer Research Center (ICRC) bacilli and Mycobacterium w either alone or in combination with killed Mycobacterium leprae. For Bacille Calmette-Guérin, the vaccine dose is generally that which gives the largest delayed-type hypersensitivity response with minimal side effects. The doses of other integral mycobacterial vaccines appear to be arbitrarily chosen. Hypotheses governing immunologic responses to complex antigens predict that the doses used may be too high, resulting in protection of some individuals and increasing the susceptibility of other individuals to leprosy. The natural history of an individual's prior exposure to environmental mycobacteria will affect the outcome of protective vaccination using a given dose of mycobacterial vaccine in the individual. PMID:14711362

Gormus, Bobby J; Meyers, Wayne M

2003-12-01

115

Bacillus calmette-guerin infection in NADPH oxidase deficiency: defective mycobacterial sequestration and granuloma formation  

OpenAIRE

Patients with chronic granulomatous disease (CGD) lack generation of reactive oxygen species (ROS) through the phagocyte NADPH oxidase NOX2. CGD is an immune deficiency that leads to frequent infections with certain pathogens; this is well documented for S. aureus and A. fumigatus, but less clear for mycobacteria. We therefore performed an extensive literature search which yielded 297 cases of CGD patients with mycobacterial infections; M. bovis BCG was most commonly described (74%). The rela...

Deffert, Christine; Scha?ppi, Michela G.; Pache, Jean-claude; Cachat, Julien; Vesin, Dominique; Bisig, Ruth; Ma Mulone, Xiaojuan; Kelkka, Tiina; Holmdahl, Rikard; Garcia, Ire?ne; Olleros, Maria-luisa; Krause, Karl-heinz

2014-01-01

116

Reduction of Contamination of Mycobacterial Growth Indicator Tubes with a Modified Antimicrobial Combination  

OpenAIRE

Culture in the fluorimetric Mycobacteria Growth Indicator Tube (MGIT) treated with a combination of vancomycin, amphotericin B, and nalidixic acid (VAN) showed growth of most strains of 31 mycobacterial species with a less-than-1-day delay. The results were similar to those in the MGIT with polymyxin B, amphotericin B, nalidixic acid, trimethoprim, and azlocillin, but with respiratory specimens, the MGIT with VAN showed a lower contamination rate with no change in the detection rate or time.

Chang, Chulhun Ludgerus; Park, Tae Sung; Oh, Seung Hwan; Kim, Hyung Hoi; Lee, Eun Yup; Son, Han Chul; Kim, Cheol Min

2002-01-01

117

Biochip System for Rapid and Accurate Identification of Mycobacterial Species from Isolates and Sputum?  

OpenAIRE

The accurate detection of mycobacterial species from isolates and clinical samples is important for pathogenic diagnosis and treatment and for disease control. There is an urgent need for the development of a rapid, simple, and accurate detection method. We established a biochip assay system, including a biochip, sample preparation apparatus, hybridization instrument, chip washing machine, and laser confocal scanner equipped with interpretation software for automatic diagnosis. The biochip si...

Zhu, Lingxiang; Jiang, Guanglu; Wang, Shengfen; Wang, Can; Li, Qiang; Yu, Hao; Zhou, Yang; Zhao, Bing; Huang, Hairong; Xing, Wanli; Mitchelson, Keith; Cheng, Jing; Zhao, Yanlin; Guo, Yong

2010-01-01

118

In Vitro Synergy between Clofazimine and Amikacin in Treatment of Nontuberculous Mycobacterial Disease  

OpenAIRE

Disease caused by nontuberculous mycobacteria (NTM) is increasing in frequency. The outcome of treatment for NTM lung disease is poor, particularly lung disease caused by Mycobacterium simiae and M. abscessus. Exploring synergy between active available drugs is a sensible way forward given the lack of new active drugs. We tested for synergy between amikacin and clofazimine, using standardized methods, in 564 consecutive clinical isolates identified as 21 species of rapidly growing mycobacteri...

Ingen, Jakko; Totten, Sarah E.; Helstrom, Niels K.; Heifets, Leonid B.; Boeree, Martin J.; Daley, Charles L.

2012-01-01

119

T-Cell Recognition of Mycobacterial GroES Peptides in Thai Leprosy Patients and Contacts  

OpenAIRE

We report here the mapping of T-cell-stimulatory determinants of the GroES 10-kDa heat shock protein homologues from Mycobacterium leprae and Mycobacterium tuberculosis, which are known as major immunogens in mycobacterial infections. Peripheral blood mononuclear cells (PBMC) from treated tuberculoid leprosy or lepromatous leprosy patients and from healthy household or hospital staff contacts of the patients were cultured with 20 16-mer peptides covering the entire sequences of both M. leprae...

Chua-intra, Boosbun; Peerapakorn, Somchai; Davey, Nick; Jurcevic, Stipo; Busson, Marc; Vordermeier, H. Martin; Pirayavaraporn, Charoon; Ivanyi, Juraj

1998-01-01

120

Specific detection of unamplified mycobacterial DNA using fluorescent semiconductor quantum dots and magnetic beads  

OpenAIRE

Here we present the development of a specific DNA detection method using fluorescent semiconductor quantum dots (QDs) and magnetic beads (MBs) for fast detection of Mycobacterium spp. dispensing with the need for DNA amplification. Two biotinylated oligonucleotide probes were used to recognize and detect specific complementary mycobacterial target DNA through a sandwich hybridization reaction. Cadmim selenite QDs conjugated with streptavidin and species specific probes were used to pro...

Gazouli, Maria; Liandris, Emmanouil; Andreadou, Margarita; Sechi, Leonardo Antonio; Masala, Speranza; Paccagnini, Daniela; Ikonomopoulos, John

2010-01-01

121

Gamma Interferon Production by Bovine ?? T Cells following Stimulation with Mycobacterial Mycolylarabinogalactan Peptidoglycan  

OpenAIRE

A large percentage of lymphocytes in the blood of cattle express the ?? T-cell receptor, but specific functions for these cells have not yet been clearly defined. There is evidence, however, that human, murine, and bovine ?? T cells have a role in the immune response to mycobacteria. This study investigated the ability of bovine ?? T cells to expand and produce gamma interferon (IFN-?) in response to stimulation with mycobacterial products. Bovine ?? T cells, isolated from the periph...

Vesosky, B.; Turner, O. C.; Turner, J.; Orme, I. M.

2004-01-01

122

Alkaline decontamination of sputum specimens adversely affects stability of mycobacterial mRNA.  

OpenAIRE

Reverse transcriptase PCR (RT-PCR) is an important tool for Mycobacterium tuberculosis research and diagnostics. A standard procedure using N-acetyl-L-cysteine (NALC) and NaOH has been widely adopted for digestion and decontamination of sputum specimens for mycobacterial culture. The objective of this study was to determine the compatibility of this method with the recovery of RNA for RT-PCR assays. Nineteen sputum specimens were collected from smear-positive, pretreatment tuberculosis patien...

Desjardin, L. E.; Perkins, M. D.; Teixeira, L.; Cave, M. D.; Eisenach, K. D.

1996-01-01

123

Comparative Genomic and Phylogenetic Approaches to Characterize the Role of Genetic Recombination in Mycobacterial Evolution  

OpenAIRE

The genus Mycobacterium encompasses over one hundred named species of environmental and pathogenic organisms, including the causative agents of devastating human diseases such as tuberculosis and leprosy. The success of these human pathogens is due in part to their ability to rapidly adapt to their changing environment and host. Recombination is the fastest way for bacterial genomes to acquire genetic material, but conflicting results about the extent of recombination in the genus Mycobacteri...

Smith, Silvia E.; Showers-corneli, Patrice; Dardenne, Caitlin N.; Harpending, Henry H.; Martin, Darren P.; Beiko, Robert G.

2012-01-01

124

Pseudo-Gaucher cells in mycobacterial infection: a report of two cases  

OpenAIRE

This report describes two cases of mycobacterial infection with pseudo-Gaucher cells. Both patients had no clinical evidence of inherited Gaucher disease. The first case was a patient with AIDS and Mycobacterium avium intracellulare involving the lung, spleen, and bone marrow. The bone marrow aspirates showed many histiocytes with needle-like inclusions. Acid fast staining showed that these histiocytes contained acid fast bacilli. Bone marrow biopsies revealed granulomatous lesions with aggre...

Dunn, P.; Kuo, M-c; Sun, C-f

2005-01-01

125

Mycobacterial laminin-binding histone-like protein mediates collagen-dependent cytoadherence  

Directory of Open Access Journals (Sweden)

Full Text Available When grown in the presence of exogenous collagen I, Mycobacterium bovis BCG was shown to form clumps. Scanning electron microscopy examination of these clumps revealed the presence of collagen fibres cross-linking the bacilli. Since collagen is a major constituent of the eukaryotic extracellular matrices, we assayed BCG cytoadherence in the presence of exogenous collagen I. Collagen increased the interaction of the bacilli with A549 type II pneumocytes or U937 macrophages, suggesting that BCG is able to recruit collagen to facilitate its attachment to host cells. Using an affinity chromatography approach, we have isolated a BCG collagen-binding protein corresponding to the previously described mycobacterial laminin-binding histone-like protein (LBP/Hlp, a highly conserved protein associated with the mycobacterial cell wall. Moreover, Mycobacterium leprae LBP/Hlp, a well-characterized adhesin, was also able to bind collagen I. Finally, using recombinant fragments of M. leprae LBP/Hlp, we mapped the collagen-binding activity within the C-terminal domain of the adhesin. Since this protein was already shown to be involved in the recognition of laminin and heparan sulphate-containing proteoglycans, the present observations reinforce the adhesive activities of LBP/Hlp, which can be therefore considered as a multifaceted mycobacterial adhesin, playing an important role in both leprosy and tuberculosis pathogenesis.

André Alves Dias

2012-12-01

126

Mycobacterial laminin-binding histone-like protein mediates collagen-dependent cytoadherence.  

Science.gov (United States)

When grown in the presence of exogenous collagen I, Mycobacterium bovis BCG was shown to form clumps. Scanning electron microscopy examination of these clumps revealed the presence of collagen fibres cross-linking the bacilli. Since collagen is a major constituent of the eukaryotic extracellular matrices, we assayed BCG cytoadherence in the presence of exogenous collagen I. Collagen increased the interaction of the bacilli with A549 type II pneumocytes or U937 macrophages, suggesting that BCG is able to recruit collagen to facilitate its attachment to host cells. Using an affinity chromatography approach, we have isolated a BCG collagen-binding protein corresponding to the previously described mycobacterial laminin-binding histone-like protein (LBP/Hlp), a highly conserved protein associated with the mycobacterial cell wall. Moreover, Mycobacterium leprae LBP/Hlp, a well-characterized adhesin, was also able to bind collagen I. Finally, using recombinant fragments of M. leprae LBP/Hlp, we mapped the collagen-binding activity within the C-terminal domain of the adhesin. Since this protein was already shown to be involved in the recognition of laminin and heparan sulphate-containing proteoglycans, the present observations reinforce the adhesive activities of LBP/Hlp, which can be therefore considered as a multifaceted mycobacterial adhesin, playing an important role in both leprosy and tuberculosis pathogenesis. PMID:23283469

Dias, André Alves; Raze, Dominique; de Lima, Cristiana Soares; Marques, Maria Angela de Melo; Drobecq, Hervé; Debrie, Anne-Sophie; Ribeiro-Guimarães, Michelle Lopes; Biet, Franck; Pessolani, Maria Cristina Vidal

2012-12-01

127

Molecular basis for the differential quinolone susceptibility of mycobacterial DNA gyrase.  

Science.gov (United States)

DNA gyrase is a type II topoisomerase that catalyzes the introduction of negative supercoils in the genomes of eubacteria. Fluoroquinolones (FQs), successful as drugs clinically, target the enzyme to trap the gyrase-DNA complex, leading to the accumulation of double-strand breaks in the genome. Mycobacteria are less susceptible to commonly used FQs. However, an 8-methoxy-substituted FQ, moxifloxacin (MFX), is a potent antimycobacterial, and a higher susceptibility of mycobacterial gyrase to MFX has been demonstrated. Although several models explain the mechanism of FQ action and gyrase-DNA-FQ interaction, the basis for the differential susceptibility of mycobacterial gyrase to various FQs is not understood. We have addressed the basis of the differential susceptibility of the gyrase and revisited the mode of action of FQs. We demonstrate that FQs bind both Escherichia coli and Mycobacterium tuberculosis gyrases in the absence of DNA and that the addition of DNA enhances the drug binding. The FQs bind primarily to the GyrA subunit of mycobacterial gyrase, while in E. coli holoenzyme is the target. The binding of MFX to GyrA of M. tuberculosis correlates with its effectiveness as a better inhibitor of the enzyme and its efficacy in cell killing. PMID:24419347

Kumar, Rupesh; Madhumathi, Bhavani Shankar; Nagaraja, Valakunja

2014-01-01

128

Recombinant gamma interferon for the treatment of pulmonary and mycobacterial diseases  

International Nuclear Information System (INIS)

An increased antibiotic resistance is described for Mycobacterium tuberculosis and atypical mycobacterial species; therefore, new treatments are required. Immunocompromised patients have increased risk, as demonstrated by complications after BCG vaccination. On the other hand, idiopathic pulmonary fibrosis is a fatal disease, with no therapy available to modify course of the disease. Gamma interferon (IFN-?) plays an essential role as main activator of cytokine secretion in macrophages, also showing a potent anti-fibrotic effects. To evaluate the adjuvant effect of IFN-? on these three clinical scenarios, five clinical trials were carried out. Patients treated with IFN gamma had satisfactory response according to clinical, imaging and functional criteria since their first evaluations, significantly improving when compared to the control group receiving placebo in a study of pulmonary atypical mycobacteriosis. Fast sputum conversion was obtained in mycobacterial infections, including tuberculosis. In the idiopathic pulmonary fibrosis study, 75% of treated patients were considered as responders (improvement + stable). Here we report the cases of two nursing babies with suppurative regional lymphadenitis caused by BCG, who were successfully treated with recombinant human IFN-?. Treatment was well tolerated, with most of the adverse reactions corresponding to classical flu-like symptoms produced by the cytokine. We can conclude that IFN-? is useful and well tolerated at IFN-? is useful and well tolerated as adjuvant therapy in patients with pulmonary mycobacterial diseases or idiopathic pulmonary fibrosis. (author)

129

Mycobacterial laminin-binding histone-like protein mediates collagen-dependent cytoadherence  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: English Abstract in english When grown in the presence of exogenous collagen I, Mycobacterium bovis BCG was shown to form clumps. Scanning electron microscopy examination of these clumps revealed the presence of collagen fibres cross-linking the bacilli. Since collagen is a major constituent of the eukaryotic extracellular mat [...] rices, we assayed BCG cytoadherence in the presence of exogenous collagen I. Collagen increased the interaction of the bacilli with A549 type II pneumocytes or U937 macrophages, suggesting that BCG is able to recruit collagen to facilitate its attachment to host cells. Using an affinity chromatography approach, we have isolated a BCG collagen-binding protein corresponding to the previously described mycobacterial laminin-binding histone-like protein (LBP/Hlp), a highly conserved protein associated with the mycobacterial cell wall. Moreover, Mycobacterium leprae LBP/Hlp, a well-characterized adhesin, was also able to bind collagen I. Finally, using recombinant fragments of M. leprae LBP/Hlp, we mapped the collagen-binding activity within the C-terminal domain of the adhesin. Since this protein was already shown to be involved in the recognition of laminin and heparan sulphate-containing proteoglycans, the present observations reinforce the adhesive activities of LBP/Hlp, which can be therefore considered as a multifaceted mycobacterial adhesin, playing an important role in both leprosy and tuberculosis pathogenesis.

André Alves, Dias; Dominique, Raze; Cristiana Soares de, Lima; Maria Angela de Melo, Marques; Hervé, Drobecq; Anne-Sophie, Debrie; Michelle Lopes, Ribeiro-Guimarães; Franck, Biet; Maria Cristina Vidal, Pessolani.

2012-12-01

130

Molecular and physiological effects of mycobacterial oxyR inactivation.  

Science.gov (United States)

The majority of slow-growing mycobacteria have a functional oxyR, the central regulator of the bacterial oxidative stress response. In contrast, this gene has been inactivated during the evolution of Mycobacterium tuberculosis. Here we inactivated the oxyR gene in Mycobacterium marinum, an organism used to model M. tuberculosis pathogenesis. Inactivation of oxyR abrogated induction of ahpC, a gene encoding alkylhydroperoxide reductase, normally activated upon peroxide challenge. The absence of oxyR also resulted in increased sensitivity to the front-line antituberculosis drug isoniazid. Inactivation of oxyR in M. marinum did not affect either virulence in a fish infection model or survival in human macrophages. Our findings demonstrate, at the genetic and molecular levels, a direct role for OxyR in ahpC regulation in response to oxidative stress. Our study also indicates that oxyR is not critical for virulence in M. marinum. However, oxyR inactivation confers increased sensitivity to isonicotinic acid hydrazide, suggesting that the natural loss of oxyR in the tubercle bacillus contributes to the unusually high sensitivity of M. tuberculosis to isoniazid. PMID:16547055

Pagán-Ramos, Eileen; Master, Sharon S; Pritchett, Christopher L; Reimschuessel, Renate; Trucksis, Michele; Timmins, Graham S; Deretic, Vojo

2006-04-01

131

Mycobacterium arosiense sp. nov., a slowly growing, scotochromogenic species causing osteomyelitis in an immunocompromised child  

DEFF Research Database (Denmark)

A yellow-pigmented, scotochromogenic, slowly growing mycobacterial strain, designated T1921(T), was isolated from the disseminated osteomyelitic lesions of a 7-year-old child with an underlying partial gamma interferon receptor alpha-1 deficiency. Hybridization by the line probe assay indicated the presence of a Mycobacterium species. Sequencing of the 16S rRNA gene, the internally transcribed spacer (ITS) region and the hsp65 and rpoB genes revealed that strain T1921(T) could be differentiated from all recognized species of the genus Mycobacterium. Phylogenetic analysis based on the 16S rRNA gene indicated that strain T1921(T) was related most closely to Mycobacterium intracellulare, whereas analysis based on the ITS and hsp65 and rpoB genes indicated that it was most closely related to Mycobacterium avium. Phenotypic tests were not able to differentiate strain T1921(T) from similar slowly growing mycobacteria. Strain T1921(T) is considered to represent a novel species of the genus Mycobacterium, for which the name Mycobacterium arosiense sp. nov. is proposed. The type strain is T1921(T) (=DSM 45069(T) =ATCC BAA-1401(T)) Udgivelsesdato: 2008/10

Bang, D.; Herlin, T.

2008-01-01

132

Chronic mycobacterial meningitis due to Mycobacterium chelonae: a case report.  

Science.gov (United States)

We report a case of chronic meningitis due to Mycobacterium chelonae. This organism is a rapidly growing Mycobacterium (RGM) and can be found worldwide in environmental sources such as soil, dust, and water. M. chelonae is an uncommon cause of meningitis; the majority of infections caused by this organism are localized cutaneous or soft tissue infections, and rarely lung infections. The organism is indistinguishable phenotypically, so we applied PCR based on the rpoB gene sequence followed by restriction fragment length polymorphism (RFLP) for molecular identification. The subsequent sequencing of RFLP products revealed 99.7% similarity with M. chelonae. PMID:25195074

Salmanzadeh, Shokrallah; Honarvar, Negin; Goodarzi, Hamed; Khosravi, Azar Dokht; Nashibi, Roohangiz; Serajian, Amir Arsalan; Hashemzadeh, Mohammad

2014-10-01

133

Identification of drug susceptibility pattern and mycobacterial species in sputum smear positive pulmonary tuberculosis patients with and without HIV co-infection in north west Ethiopia  

DEFF Research Database (Denmark)

Ethiopia is among the high-burden countries of tuberculosis (TB) in the world Since mycobacterial culture and susceptibility testing are not routinely performed in Ethiopia, recent data on susceptibility patterns and the mycobacterial species cultured from sputum smear positive patients are limited.

Mekonen, Mekdem; Abate, Ebba

2010-01-01

134

Probing the interaction of the diarylquinoline TMC207 with its target mycobacterial ATP synthase.  

Science.gov (United States)

Infections with Mycobacterium tuberculosis are substantially increasing on a worldwide scale and new antibiotics are urgently needed to combat concomitantly emerging drug-resistant mycobacterial strains. The diarylquinoline TMC207 is a highly promising drug candidate for treatment of tuberculosis. This compound kills M. tuberculosis by binding to a new target, mycobacterial ATP synthase. In this study we used biochemical assays and binding studies to characterize the interaction between TMC207 and ATP synthase. We show that TMC207 acts independent of the proton motive force and does not compete with protons for a common binding site. The drug is active on mycobacterial ATP synthesis at neutral and acidic pH with no significant change in affinity between pH 5.25 and pH 7.5, indicating that the protonated form of TMC207 is the active drug entity. The interaction of TMC207 with ATP synthase can be explained by a one-site binding mechanism, the drug molecule thus binds to a defined binding site on ATP synthase. TMC207 affinity for its target decreases with increasing ionic strength, suggesting that electrostatic forces play a significant role in drug binding. Our results are consistent with previous docking studies and provide experimental support for a predicted function of TMC207 in mimicking key residues in the proton transfer chain and blocking rotary movement of subunit c during catalysis. Furthermore, the high affinity of TMC207 at low proton motive force and low pH values may in part explain the exceptional ability of this compound to efficiently kill mycobacteria in different microenvironments. PMID:21858172

Haagsma, Anna C; Podasca, Ioana; Koul, Anil; Andries, Koen; Guillemont, Jerome; Lill, Holger; Bald, Dirk

2011-01-01

135

Elucidation of the dual role of Mycobacterial MoeZR in molybdenum cofactor biosynthesis and cysteine biosynthesis.  

Science.gov (United States)

The pathway of molybdenum cofactor biosynthesis has been studied in detail by using proteins from Mycobacterium species, which contain several homologs associated with the first steps of Moco biosynthesis. While all Mycobacteria species contain a MoeZR, only some strains have acquired an additional homolog, MoeBR, by horizontal gene transfer. The role of MoeBR and MoeZR was studied in detail for the interaction with the two MoaD-homologs involved in Moco biosynthesis, MoaD1 and MoaD2, in addition to the CysO protein involved in cysteine biosynthesis. We show that both proteins have a role in Moco biosynthesis, while only MoeZR, but not MoeBR, has an additional role in cysteine biosynthesis. MoeZR and MoeBR were able to complement an E. coli moeB mutant strain, but only in conjunction with the Mycobacterial MoaD1 or MoaD2 proteins. Both proteins were able to sulfurate MoaD1 and MoaD2 in vivo, while only MoeZR additionally transferred the sulfur to CysO. Our in vivo studies show that Mycobacteria have acquired several homologs to maintain Moco biosynthesis. MoeZR has a dual role in Moco- and cysteine biosynthesis and is involved in the sulfuration of MoaD and CysO, whereas MoeBR only has a role in Moco biosynthesis, which is not an essential function for Mycobacteria. PMID:22140533

Voss, Martin; Nimtz, Manfred; Leimkühler, Silke

2011-01-01

136

Comparison of three methods of decontamination of sputum for Mycobacterial culture  

OpenAIRE

Three methods of decontamination of sputum for mycobacterial culture viz. Swah method of Nassau, Petroff’s method and NALC-NaOH method have been compared. Swab method was found to be best giving 92.3% positive cultures. Positive cultures in the other two methods were 80.7% and 78.8% respectively. The rate of contamination was equal (3.9%) in Swab and Petroff’s method, while NALC-NaOH method gave a higher contamination are (8.6%).

Damle, A. S.; Kaundinya, D. V.

1986-01-01

137

Alteraciones en el reclutamiento y activación de proteínas Rab durante la infección micobacteriana / Alterations in recruitment and activation of Rab proteins during mycobacterial infection  

Scientific Electronic Library Online (English)

Full Text Available SciELO Colombia | Language: Spanish Abstract in spanish En el fagosoma, Mycobacterium spp. altera la activación y reclutamiento de diferentes proteínas "del gen Ras de cerebro de rata", comúnmente conocidas como Rab. En este manuscrito se revisa una serie de reportes que han demostrado que los fagosomas que contienen micobacterias tienen una expresión ma [...] yor y sostenida de Rab5, Rab11, Rab14 y Rab22a, y menor o ninguna expresión de Rab7, Rab9 y Rab6. Esto se correlaciona con aumento de la fusión de estos fagosomas con endosomas tempranos y de reciclaje, lo que les permite mantener ciertas características de compartimentos tempranos, permite que las bacterias obtengan acceso a nutrientes y previene la activación de mecanismos contra la micobacteria. La expresión de mutantes constitutivamente activos de las Rab de endosomas tempranos impide la maduración de fagosomas que contienen esferas de látex o micobacterias inactivadas por calor. Mientras que su silenciamiento, mediante ARN de interferencia o mediante dominantes negativos, induce la maduración de fagosomas micobacterianos. Los mecanismos exactos por los que las micobacterias alteran la dinámica de expresión de estas GTPasas, afectando la maduración fagolisosómica, no se han establecido. El problema podría explicarse por defectos en el reclutamiento de las proteínas que interactúan con Rab, como la cinasa-3 del fosfatidilinositol y el antígeno endosómico temprano 1. La identificación de los mecanismos empleados por Mycobacterium spp. para interrumpir el ciclo de activación de las Rab, será esencial para comprender la fisiopatología de la infección micobacteriana y útil como posibles blancos farmacológicos. Abstract in english At the phagosome level, Mycobacterium spp. alters activation and recruitment of several "Ras gene from rat brain" proteins, commonly known as Rab. Mycobacterial phagosomes have a greater and sustained expression of Rab5, Rab11, Rab14 and Rab22a, and lowered or no expression of Rab7, Rab9 and Rab6. T [...] his correlates with increased fusion of the phagosomes with early and recycling endosomes acquiring some features of early phogosomes, allowing the bacteria to gain access to nutrients and preventing the activation of anti-mycobacterial mechanisms. The expression of constitutively active mutants of Rab from the early stage endosomes prevents the maturation of phagosomes containing latex beads or heat-inactivated mycobacteria. Silencing of these mutants by interference RNA or dominant negative forms induces the maturation of mycobacterial phagosomes. The mechanisms have not been established by which mycobacteria alter the expression of these GTPases and thereby shift the phagolysosomal maturation. The problem can be explained by alterations in the recruitment of proteins that interact with Rab, such as phosphoinositide 3-kinases and early endosomal antigen 1. Identifying the mechanisms used by Mycobacterium spp. to disrupt the cycle of Rab activation will be essential to understand the pathophysiology of mycobacterial infections and usefully to potential drug targets.

Diana, Castaño; Mauricio, Rojas.

2010-06-01

138

Genes  

Science.gov (United States)

Illustration of the placement of genes in a chromosome. A gene can be defined as a region of DNA that controls a hereditary characteristic. It usually corresponds to a sequence used in the production of a specific protein or RNA. A gene carries biological information in a form that must be copied and transmitted from each cell to all its progeny. This includes the entire functional unit: coding DNA sequences, non-coding regulatory DNA sequences, and introns. Genes can be as short as 1000 base pairs or as long as several hundred thousand base pairs. It can even be carried by more than one chromosome. The estimate for the number of genes in humans has decreased as our knowledge has increased. As of 2001, humans are thought to have between 30,000 and 40,000 genes.

Access Excellence

2005-03-12

139

Heterogeneity of RNA polymerase gene (rpoB) sequences of Mycobacterium gordonae clinical isolates identified with a DNA probe kit and by conventional methods.  

Science.gov (United States)

In a previous study, we have evaluated genetic identification by using the rpoB gene, which was recently introduced by Kim et al. (J. Clin. Microbiol. 39:2102-2109, 2001; J. Clin. Microbiol. 37:1714-1720, 1999). In this process, we examined the rpoB gene heterogeneity of clinical isolates identified as Mycobacterium gordonae with the conventional biological and biochemical tests and/or a commercially available DNA probe kit. Sequencing of the rpoB gene of 34 clinical isolates revealed that M. gordonae clinical isolates were classified into four major clusters (A, B, C, and D). Interestingly, organisms belonging to cluster D (15 isolates) did not hybridize with M. gordonae ATCC 14470 and specifically possessed urease activity. Therefore, it could be considered to be a novel mycobacterium. The identification of M. gordonae is known to have ambiguous results sometimes. On the other hand, identification of clinical isolates seems to be inconvenient and unsuitable because of a more than 99% 16S rRNA gene similarity value between clusters. These findings suggest that the existence of M. gordonae-like mycobacteria that share similar biochemical and biological characteristics with the 16S rRNA gene of an M. gordonae type strain but less similarity at the genomic DNA level may have complicated the identification of M. gordonae in many laboratories. Furthermore, compared with hsp65 PCR restriction analysis (PRA), rpoB PRA would have the advantage of producing no ambiguous results because of the intracluster homogeneity of the rpoB gene. In this case, rpoB would provide clearer results than hsp65, even if PRA analysis was used. We demonstrated that these M. gordonae-like mycobacteria were easily distinguished by PRA of the rpoB sequence. Additionally, the significance of this M. gordonae-like cluster may help to establish the comparison between the M. gordonae isolates from a clinical specimen and an infectious process in a given patient and to determine the true incidence of infection with this microorganism. PMID:12682157

Itoh, Saotomo; Kazumi, Yuko; Abe, Chiyoji; Takahashi, Mitsuyoshi

2003-04-01

140

Characterization of two heparan sulphate-binding sites in the mycobacterial adhesin Hlp  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background The histone-like Hlp protein is emerging as a key component in mycobacterial pathogenesis, being involved in the initial events of host colonization by interacting with laminin and glycosaminoglycans (GAGs. In the present study, nuclear magnetic resonance (NMR was used to map the binding site(s of Hlp to heparan sulfate and identify the nature of the amino acid residues directly involved in this interaction. Results The capacity of a panel of 30 mer synthetic peptides covering the full length of Hlp to bind to heparin/heparan sulfate was analyzed by solid phase assays, NMR, and affinity chromatography. An additional active region between the residues Gly46 and Ala60 was defined at the N-terminal domain of Hlp, expanding the previously defined heparin-binding site between Thr31 and Phe50. Additionally, the C-terminus, rich in Lys residues, was confirmed as another heparan sulfate binding region. The amino acids in Hlp identified as mediators in the interaction with heparan sulfate were Arg, Val, Ile, Lys, Phe, and Thr. Conclusion Our data indicate that Hlp interacts with heparan sulfate through two distinct regions of the protein. Both heparan sulfate-binding regions here defined are preserved in all mycobacterial Hlp homologues that have been sequenced, suggesting important but possibly divergent roles for this surface-exposed protein in both pathogenic and saprophic species.

Previato Jose O

2008-05-01

141

Husbandry stress exacerbates mycobacterial infections in adult zebrafish, Danio rerio (Hamilton)  

Science.gov (United States)

Mycobacteria are significant pathogens of laboratory zebrafish, Danio rerio (Hamilton). Stress is often implicated in clinical disease and morbidity associated with mycobacterial infections but has yet to be examined with zebrafish. The aim of this study was to examine the effects of husbandry stressors on zebrafish infected with mycobacteria. Adult zebrafish were exposed to Mycobacterium marinum or Mycobacterium chelonae, two species that have been associated with disease in zebrafish. Infected fish and controls were then subjected to chronic crowding and handling stressors and examined over an 8-week period. Whole-body cortisol was significantly elevated in stressed fish compared to non-stressed fish. Fish infected with M. marinum ATCC 927 and subjected to husbandry stressors had 14% cumulative mortality while no mortality occurred among infected fish not subjected to husbandry stressors. Stressed fish, infected with M. chelonae H1E2 from zebrafish, were 15-fold more likely to be infected than non-stressed fish at week 8 post-injection. Sub-acute, diffuse infections were more common among stressed fish infected with M. marinum or M. chelonae than non-stressed fish. This is the first study to demonstrate an effect of stress and elevated cortisol on the morbidity, prevalence, clinical disease and histological presentation associated with mycobacterial infections in zebrafish. Minimizing husbandry stress may be effective at reducing the severity of outbreaks of clinical mycobacteriosis in zebrafish facilities. ?? 2009 Blackwell Publishing Ltd.

Ramsay, J.M.; Watral, V.; Schreck, C.B.; Kent, M.L.

2009-01-01

142

Mycobacterial spindle cell pseudotumor of the appendix vermiformis in a patient with AIDS  

Directory of Open Access Journals (Sweden)

Full Text Available Mycobacterial pseudotumor (MP is a rare pathologic presentation of both Mycobacterium tuberculosis and non-tuberculous mycobacterial disease, hitherto reported to occur only in immunosuppressed patients with or without human immunodeficiency virus infection. This lesion shares close pathologic resemblance to certain mesenchymal neoplasms, particularly Kaposi's sarcoma (KS, from which it must be properly differentiated due to distinct prognosis and therapy. We report a case of MP obliterating the lumen of the appendix vermiformis in a 34-year-old patient who died of complications of AIDS at our hospital in Rio de Janeiro. A total of 24 cases of MP (including our patient have been described in the literature. MP has been found especially in lymph nodes, but extranodal lesions have been described in the skin, spleen, lung, bone marrow, brain and, in our patient, the appendix vermiformis. We offer a review of the other 23 published case reports of MP in both HIV-infected and uninfected patients and discuss the pathologic features that differentiate MP from KS.

Carlos Alberto Basílio-de-Oliveira

2001-04-01

143

Restoration of innate immune activation accelerates Th1-cell priming and protection following pulmonary mycobacterial infection.  

Science.gov (United States)

The immune mechanisms underlying delayed induction of Th1-type immunity in the lungs following pulmonary mycobacterial infection remain poorly understood. We have herein investigated the underlying immune mechanisms for such delayed responses and whether a selected innate immune-modulating strategy can accelerate Th1-type responses. We have found that, in the early stage of pulmonary infection with attenuated Mycobacterium tuberculosis (M.tb H37Ra), the levels of infection in the lung continue to increase logarithmically until days 14 and 21 postinfection in C57BL/6 mice. The activation of innate immune responses, particularly DCs, in the lung is delayed. This results in a delay in the subsequent downstream immune responses including the migration of antigen-bearing DCs to the draining lymph node (dLN), the Th1-cell priming in dLN, and the recruitment of Th1 cells to the lung. However, single lung mucosal exposure to the TLR agonist FimH postinfection is able to accelerate protective Th1-type immunity via facilitating DC migration to the lung and draining lymph nodes, enhancing DC antigen presentation and Th1-cell priming. These findings hold implications for the development of immunotherapeutic and vaccination strategies and suggest that enhancement of early innate immune activation is a viable option for improving Th1-type immunity against pulmonary mycobacterial diseases. PMID:24519467

Lai, Rocky; Jeyanathan, Mangalakumari; Shaler, Christopher R; Damjanovic, Daniela; Khera, Amandeep; Horvath, Carly; Ashkar, Ali A; Xing, Zhou

2014-05-01

144

[Evaluation of mycobacterial infections using 18F-fluorodeoxyglucose-positron emission tomography: results of nine cases].  

Science.gov (United States)

18F-fluorodeoxyglucose-positron emission tomography (FDG-PET/CT) is a useful technique for distinguishing malignant and benign lesions, although the occurrence of false-positive results in cases involving benign lesions is possible. We evaluated nine patients with mycobacterial infections who underwent FDG-PET/CT from April 2008 to July 2010. FDG-PET/CT was performed 1-2h (during the early and late phases) after administration of FDG at a dose of 185 MBq/individual after fasting for at least 5h. Out of the nine patients, four were diagnosed with pulmonary nonmycobacterium tuberculosis, two with pulmonary tuberculosis, two with tuberculous lymphadenopathy, and one with pleural tuberculoma. All patients had a maximum standardized uptake value (SUV(max)) of > 2.5, and the SUV(max) increased from the early to the late phase. One lesion that occurred due to tuberculous pleurisy after treatment demonstrated high FDG uptake, similar to the other cases. It is difficult to distinguish mycobacterial infections from malignant diseases using FGD-PET alone; hence, the use of high-resolution CT and bacteriological tests is required for diagnosis and distinction. PMID:24716357

Uruga, Hironori; Ishihara, Makiko; Hanada, Shigeo; Takaya, Hisashi; Miyamoto, Atsushi; Morokawa, Nasa; Fujii, Takeshi; Kurosaki, Atsuko; Kishi, Kazuma

2014-02-01

145

Rapid susceptibility testing of Mycobacterium tuberculosis by bioluminescence assay of mycobacterial ATP  

International Nuclear Information System (INIS)

Mycobacterial growth was monitored by bioluminescence assay of mycobacterial ATP. Cultures of Mycobacterium tuberculosis H37Rv and of 25 clinical isolates of the same species were exposed to serial dilutions of ethambutol, isoniazid, rifampin, and streptomycin. A suppression of ATP, indicating growth inhibition, occurred for susceptible but not resistant strains within 5 to 7 days of incubation. Breakpoint concentrations between susceptibility and resistance were determined by comparing these results with those obtained by reference techniques. Full agreement was found in 99% of the assays with the resistance ratio method on Lowenstein-Jensen medium, and 98% of the assays were in full agreement with the radiometric system (BACTEC). A main advantage of the bioluminescence method is its rapidity, with results available as fast as with the radiometric system but at a lower cost and without the need for radioactive culture medium. The method provides kinetic data concerning drug effects within available in vivo drug concentrations and has great potential for both rapid routine susceptibility testing and research applications in studies of drug effects on mycobacteria

146

Development of a Murine Mycobacterial Growth Inhibition Assay for Evaluating Vaccines against Mycobacterium tuberculosis? †  

Science.gov (United States)

The development and characterization of new tuberculosis (TB) vaccines has been impeded by the lack of reproducible and reliable in vitro assays for measuring vaccine activity. In this study, we developed a murine in vitro mycobacterial growth inhibition assay for evaluating TB vaccines that directly assesses the capacity of immune splenocytes to control the growth of Mycobacterium tuberculosis within infected macrophages. Using this in vitro assay, protective immune responses induced by immunization with five different types of TB vaccine preparations (Mycobacterium bovis BCG, an attenuated M. tuberculosis mutant strain, a DNA vaccine, a modified vaccinia virus strain Ankara [MVA] construct expressing four TB antigens, and a TB fusion protein formulated in adjuvant) can be detected. Importantly, the levels of vaccine-induced mycobacterial growth-inhibitory responses seen in vitro after 1 week of coculture correlated with the protective immune responses detected in vivo at 28 days postchallenge in a mouse model of pulmonary tuberculosis. In addition, similar patterns of cytokine expression were evoked at day 7 of the in vitro culture by immune splenocytes taken from animals immunized with the different TB vaccines. Among the consistently upregulated cytokines detected in the immune cocultures are gamma interferon, growth differentiation factor 15, interleukin-21 (IL-21), IL-27, and tumor necrosis factor alpha. Overall, we have developed an in vitro functional assay that may be useful for screening and comparing new TB vaccine preparations, investigating vaccine-induced protective mechanisms, and assessing manufacturing issues, including product potency and stability. PMID:19458207

Parra, Marcela; Yang, Amy L.; Lim, JaeHyun; Kolibab, Kristopher; Derrick, Steven; Cadieux, Nathalie; Perera, Liyanage P.; Jacobs, William R.; Brennan, Michael; Morris, Sheldon L.

2009-01-01

147

Development of a murine mycobacterial growth inhibition assay for evaluating vaccines against Mycobacterium tuberculosis.  

Science.gov (United States)

The development and characterization of new tuberculosis (TB) vaccines has been impeded by the lack of reproducible and reliable in vitro assays for measuring vaccine activity. In this study, we developed a murine in vitro mycobacterial growth inhibition assay for evaluating TB vaccines that directly assesses the capacity of immune splenocytes to control the growth of Mycobacterium tuberculosis within infected macrophages. Using this in vitro assay, protective immune responses induced by immunization with five different types of TB vaccine preparations (Mycobacterium bovis BCG, an attenuated M. tuberculosis mutant strain, a DNA vaccine, a modified vaccinia virus strain Ankara [MVA] construct expressing four TB antigens, and a TB fusion protein formulated in adjuvant) can be detected. Importantly, the levels of vaccine-induced mycobacterial growth-inhibitory responses seen in vitro after 1 week of coculture correlated with the protective immune responses detected in vivo at 28 days postchallenge in a mouse model of pulmonary tuberculosis. In addition, similar patterns of cytokine expression were evoked at day 7 of the in vitro culture by immune splenocytes taken from animals immunized with the different TB vaccines. Among the consistently upregulated cytokines detected in the immune cocultures are gamma interferon, growth differentiation factor 15, interleukin-21 (IL-21), IL-27, and tumor necrosis factor alpha. Overall, we have developed an in vitro functional assay that may be useful for screening and comparing new TB vaccine preparations, investigating vaccine-induced protective mechanisms, and assessing manufacturing issues, including product potency and stability. PMID:19458207

Parra, Marcela; Yang, Amy L; Lim, JaeHyun; Kolibab, Kristopher; Derrick, Steven; Cadieux, Nathalie; Perera, Liyanage P; Jacobs, William R; Brennan, Michael; Morris, Sheldon L

2009-07-01

148

Rapid identification of strains belonging to the Mycobacterium abscessus group through erm(41) gene pyrosequencing.  

Science.gov (United States)

Mycobacterium abscessus and Mycobacterium massiliense lung infections have different clarithromycin susceptibilities, making proper identification important; however, standard multi-gene sequencing in clinical laboratories is laborious and time consuming. We developed a pyrosequencing-based method for rapid identification of strains belonging to the M. abscessus group by targeting erm(41). We examined 55 isolates from new pulmonary M. abscessus infections and identified 28 M. abscessus, 25 M. massiliense, and 2 Mycobacterium bolletii isolates. Multi-gene sequencing of 16S rRNA, hsp65, rpoB, and the 16S-23S ITS region was concordant with the results of erm(41) pyrosequencing; thus, the M. abscessus group can be identified by single-nucleotide polymorphisms in erm(41). The method also enables rapid identification of polymorphic, inducible clarithromycin-resistant sequevars (T28 or C28). Pyrosequencing of erm(41) is a rapid, reliable, high-throughput alternative method for identifying and characterizing M. abscessus species. Further testing of a diverse collection of isolates is necessary to demonstrate the discriminatory power of erm(41) sequencing to differentiating species with this highly divergent group. PMID:24809859

Yoshida, Shiomi; Tsuyuguchi, Kazunari; Suzuki, Katsuhiro; Tomita, Motohisa; Okada, Masaji; Shimada, Ryoko; Hayashi, Seiji

2014-07-01

149

The DNA damage-regulated autophagy modulator DRAM1 links mycobacterial recognition via TLP-MYD88 to authophagic defense.  

Science.gov (United States)

Autophagy is an important defense mechanism against mycobacteria, the causative agents of tuberculosis. The molecular mechanisms that link mycobacterial recognition to autophagy remain unclear. Our analysis in zebrafish and human macrophage models of mycobacterial infection reveals that the DNA damage-regulated autophagy modulator DRAM1 functions downstream of pathogen recognition by the Toll-like receptor (TLR)/interleukin-1 receptor (IL1R)-MYD88-NF-?B innate immune sensing pathway to activate selective autophagy. Mycobacterial infection of human macrophages and zebrafish embryos induced DRAM1 expression in a MYD88 and NF-?B-dependent manner. DRAM1 knockdown increased mycobacterial infection, whereas overexpression lowered infection by hyperactivating autophagy. DRAM1-mediated selective autophagic defenses require the cytosolic DNA sensor STING and the selective autophagy receptor p62/SQSTM1. Contrary to its known role in autophagy-mediated cell death and cancer, this DRAM1 function is p53 independent. We propose that DRAM1 mediates autophagic defense against a broader range of intracellular pathogens, since DRAM1 expression was also induced by the common bacterial endotoxin lipopolysaccharide. PMID:24922577

van der Vaart, Michiel; Korbee, Cornelis J; Lamers, Gerda E M; Tengeler, Anouk C; Hosseini, Rohola; Haks, Mariëlle C; Ottenhoff, Tom H M; Spaink, Herman P; Meijer, Annemarie H

2014-06-11

150

Immune reconstitution inflammatory syndrome in HIV-infected patients with mycobacterial infections starting highly active anti-retroviral therapy  

International Nuclear Information System (INIS)

AIM: To describe the radiological appearances of immune reconstitution inflammatory syndrome (IRIS) in human immunodeficiency virus (HIV)-infected patients with mycobacterial infections starting highly active anti-retroviral therapy (HAART). MATERIALS AND METHODS: Five consecutive HIV infected patients with IRIS due to mycobacterial infection were studied. Intercurrent infection and poor drug compliance were excluded as causes of presentation. The chest radiological appearances at the time of starting HAART and at the time of diagnosis of IRIS were compared. RESULTS: In these five patients there was clinical and radiological deterioration, occurring between 10 days and 7 months after starting HAART, leading to unmasking of previously undiagnosed mycobacterial infection or to worsening of mycobacterial disease. All five patients had HAART-induced increases in CD4+ T lymphocyte counts and reductions in peripheral blood HIV 'viral load'. Chest radiographic abnormalities due to IRIS included marked mediastinal lymphadenopathy in three patients--severe enough to produce tracheal compression in two patients (one of whom had stridor)--and was associated with new pulmonary infiltrates in two patients. The other two patients had new infiltrates, which in one patient was associated with a pleural effusion. CONCLUSION: These cases illustrate the diverse chest radiographic appearances of IRIS occurring after HAART in patients with mycobacterial and HIV co-infection. Marked mediasterial and HIV co-infection. Marked mediastinal lymphadenopathy occurred in three of these five patients (with associated tracheal narrowing in two patients); four patients developed pulmonary infiltrates and one had an effusion. The cases further highlight that the onset of IRIS may be delayed for several months after HAART is started

151

Mycobacterial Infections  

Science.gov (United States)

... especially people with other problems that affect their immunity, such as AIDS. Sometimes you can have these infections with no symptoms at all. At other times, they can cause lung symptoms similar to tuberculosis: Cough Weight loss Coughing up blood or mucus ...

152

Detection of Mycobacterium bovis and Mycobacterium tuberculosis from Cattle: Possible Public Health Relevance  

DEFF Research Database (Denmark)

Mycobacterium bovis and Mycobacterium tuberculosis infect both animals and humans. The disease epidemiology by these agents differs in developed and developing countries due to the differences in the implementation of the prevention and control strategies. The present study describes the detection of M. bovis and M. tuberculosis from specimens of lungs and pulmonary lymph nodes of four cattle died in an organized herd of 183 cattle in the state of Himachal Pradesh, India, with inconclusive skin test results. Identification and distinction of these closely related mycobacterial species was done by PCR-RFLP targeting hsp65 gene followed by spacer oligonucleotide typing. Mixed infection of M. bovis and M. tuberculosis was detected in one cattle.

Thakur, Aneesh; Sharma, Mandeep

2012-01-01

153

Simple and rational approach to the identification of Mycobacterium tuberculosis, Mycobacterium avium complex species, and other commonly isolated mycobacteria.  

Science.gov (United States)

A novel PCR-restriction fragment length polymorphism analysis of the hsp65 gene was developed. The restriction patterns for Mycobacterium tuberculosis and Mycobacterium avium complex (MAC) species were designed to be highly distinct, and the overall number of restriction patterns was designed to be limited. Four hundred specimens (17 reference strains and 383 clinical isolates) were tested, of which 98 were M. tuberculosis and 132 were MAC species. The assay was virtually 100% sensitive and specific for M. tuberculosis and MAC species. Moreover, it gave highly concordant results for other mycobacterial species other than M. terrae complex species. This assay can be completed in one day and is user-friendly and robust. Therefore, it is highly suitable for large-scale use in a clinical laboratory. PMID:11574614

Wong, D A; Yip, P C; Cheung, D T; Kam, K M

2001-10-01

154

Evaluation of Biphasic Culture System for Mycobacterial Isolation from the Sputum of Patients with Pulmonary Tuberculosis  

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Full Text Available Mycobacterial diseases continue to cause high morbidity and mortality. Isolation, identification and sensitivity testing form the backbone of laboratory investigations. M. tuberculosis isolation needs 6-8 weeks on conventional egg containing media. For rapid isolation various methods have been evaluated. We evaluated biphasic system (Middlebrook 7H11 agar slant + Middlebrook 9H broth in comparison with Lowenstein - Jensen (LJ medium. In smear positive cases biphasic system showed the recovery rate of 97.05% as against 79.41% on LJ on incubation for 21± 4.44 and 28±3.76 days respectively. In smear negative and culture positive cases biphasic system and LJ showed isolation rates of 91.66% and 66.6% after 36±3.44 and 41± 4.09 days respectively. Biphasic system showed lower contamination rate (1.33%. Biphasic medium is superior to LJ medium in isolation of M. tuberculosis .

Ghatole M

2005-01-01

155

Mycobacterial Ser/Thr protein kinases and phosphatases: physiological roles and therapeutic potential.  

Science.gov (United States)

Reversible protein phosphorylation is a major regulation mechanism of fundamental biological processes, not only in eukaryotes but also in bacteria. A growing body of evidence suggests that Ser/Thr phosphorylation play important roles in the physiology and virulence of Mycobacterium tuberculosis, the etiological agent of tuberculosis. This pathogen uses 'eukaryotic-like' Ser/Thr protein kinases and phosphatases not only to regulate many intracellular metabolic processes, but also to interfere with signaling pathways of the infected host cell. Disrupting such processes by means of selective inhibitors may thus provide new pharmaceutical weapons to combat the disease. Here we review the current knowledge on Ser/Thr protein kinases and phosphatases in M. tuberculosis, their regulation mechanisms and putative substrates, and we explore their therapeutic potential as possible targets for the development of new anti-mycobacterial compounds. PMID:17869195

Wehenkel, Annemarie; Bellinzoni, Marco; Graña, Martin; Duran, Rosario; Villarino, Andrea; Fernandez, Pablo; Andre-Leroux, Gwénaëlle; England, Patrick; Takiff, Howard; Cerveñansky, Carlos; Cole, Stewart T; Alzari, Pedro M

2008-01-01

156

Detection and Differentiation of Mycobacterium tuberculosis and Nontuberculous Mycobacterial Isolates by Real-Time PCR  

Science.gov (United States)

Mycobacteria cause a variety of illnesses that differ in severity and public health implications. The differentiation of Mycobacterium tuberculosis from nontuberculous mycobacteria (NTM) is of primary importance for infection control and choice of antimicrobial therapy. Despite advances in molecular diagnostics, the ability to rapidly diagnose M. tuberculosis infections by PCR is still inadequate, largely because of the possibility of false-negative reactions. We designed and validated a real-time PCR for mycobacteria by using the LightCycler system with 18 reference strains and 168 clinical mycobacterial isolates. All clinically significant mycobacteria were detected; the mean melting temperatures (with 99.9% confidence intervals [99.9% CI] in parentheses) for the different mycobacteria were as follows: M. tuberculosis, 64.35°C (63.27 to 65.42°C); M. kansasii, 59.20°C (58.07 to 60.33°C); M. avium, 57.82°C (57.05 to 58.60°C); M. intracellulare, 54.46°C (53.69 to 55.23°C); M. marinum, 58.91°C (58.28 to 59.55°C); rapidly growing mycobacteria, 53.09°C (50.97 to 55.20°C) or 43.19°C (42.19 to 44.49°C). This real-time PCR assay with melting curve analysis consistently accurately detected and differentiated M. tuberculosis from NTM. Detection of an NTM helps ensure that the negative result for M. tuberculosis is a true negative. The specific melting temperature also provides a suggestion of the identity of the NTM present, when the most commonly encountered mycobacterial species are considered. In a parallel comparison, both the LightCycler assay and the COBAS Amplicor M. tuberculosis assay correctly categorized 48 of 50 specimens that were proven by culture to contain M. tuberculosis, and the LightCycler assay correctly characterized 3 of 3 specimens that contained NTM. PMID:14605148

Shrestha, Nabin K.; Tuohy, Marion J.; Hall, Gerri S.; Reischl, Udo; Gordon, Steven M.; Procop, Gary W.

2003-01-01

157

Rapid radiometric methods to detect and differentiate Mycobacterium tuberculosis/M. bovis from other mycobacterial species  

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Rapid methods for the differentiation of Mycobacterium tuberculosis/M. bovis (TB complex) from other mycobacteria (MOTT bacilli) were developed and evaluated in a three-phase study. In the first phase, techniques for identification of Mycobacterium species were developed by using radiometric technology and BACTEC Middlebrook 7H12 liquid medium. Based on /sup 14/CO/sub 2/ evolution, characteristic growth patterns were established for 13 commonly encountered mycobacterial species. Mycobacteria belonging to the TB complex were differentiated from other mycobacteria by cellular morphology and rate of /sup 14/CO/sub 2/ evolution. For further differentiation, radiometric tests for niacin production and inhibition by Q-nitro-alpha-acetyl amino-beta-hydroxy-propiophenone (NAP) were developed. In the second phase, 100 coded specimens on Lowenstein-Jensen medium were identified as members of the TB complex, MOTT bacilli, bacteria other than mycobacteria, or ''no viable organisms'' within 3 to 12 (average 6.4) days of receipt from the Centers for Disease Control. Isolation and identification of mycobacteria from 20 simulated sputum specimens were carried out in phase III. Out of 20 sputum specimens, 16 contained culturable mycobacteria, and all of the positives were detected by the BACTEC method in an average of 7.3 days. The positive mycobacterial cultures were isolated and identified as TB complex or MOTT bacilli in an average of 12.8 days. The radiometric NAP test was found to be highly sensitive and specific for a rapid identification of TB complex, whereas the radiometric niacin test was found to have some inherent problems. Radiometric BACTEC and conventional methodologies were in complete agreement in Phase II as well as in Phase III.

Siddiqi, S.H.; Hwangbo, C.C.; Silcox, V.; Good, R.C.; Snider, D.E. Jr.; Middlebrook, G.

1984-10-01

158

Rapid radiometric methods to detect and differentiate Mycobacterium tuberculosis/M. bovis from other mycobacterial species  

International Nuclear Information System (INIS)

Rapid methods for the differentiation of Mycobacterium tuberculosis/M. bovis (TB complex) from other mycobacteria (MOTT bacilli) were developed and evaluated in a three-phase study. In the first phase, techniques for identification of Mycobacterium species were developed by using radiometric technology and BACTEC Middlebrook 7H12 liquid medium. Based on 14CO2 evolution, characteristic growth patterns were established for 13 commonly encountered mycobacterial species. Mycobacteria belonging to the TB complex were differentiated from other mycobacteria by cellular morphology and rate of 14CO2 evolution. For further differentiation, radiometric tests for niacin production and inhibition by Q-nitro-alpha-acetyl amino-beta-hydroxy-propiophenone (NAP) were developed. In the second phase, 100 coded specimens on Lowenstein-Jensen medium were identified as members of the TB complex, MOTT bacilli, bacteria other than mycobacteria, or ''no viable organisms'' within 3 to 12 (average 6.4) days of receipt from the Centers for Disease Control. Isolation and identification of mycobacteria from 20 simulated sputum specimens were carried out in phase III. Out of 20 sputum specimens, 16 contained culturable mycobacteria, and all of the positives were detected by the BACTEC method in an average of 7.3 days. The positive mycobacterial cultures were isolated and identified as TB complex or MOTT bacilli in an average of 12.8 days. The radiometric an average of 12.8 days. The radiometric NAP test was found to be highly sensitive and specific for a rapid identification of TB complex, whereas the radiometric niacin test was found to have some inherent problems. Radiometric BACTEC and conventional methodologies were in complete agreement in Phase II as well as in Phase III

159

Characteristics of suppressor macrophages induced by mycobacterial and protozoal infections in relation to alternatively activated M2 macrophages.  

Science.gov (United States)

In the advanced stages of mycobacterial infections, host immune systems tend to change from a Th1-type to Th2-type immune response, resulting in the abrogation of Th1 cell- and macrophage-mediated antimicrobial host protective immunity. Notably, this type of immune conversion is occasionally associated with the generation of certain types of suppressor macrophage populations. During the course of Mycobacterium tuberculosis (MTB) and Mycobacterium avium-intracellulare complex (MAC) infections, the generation of macrophages which possess strong suppressor activity against host T- and B-cell functions is frequently encountered. This paper describes the immunological properties of M1- and M2-type macrophages generated in tumor-bearing animals and those generated in hosts with certain microbial infections. In addition, this paper highlights the immunological and molecular biological characteristics of suppressor macrophages generated in hosts with mycobacterial infections, especially MAC infection. PMID:22666284

Tomioka, Haruaki; Tatano, Yutaka; Maw, Win Win; Sano, Chiaki; Kanehiro, Yuichi; Shimizu, Toshiaki

2012-01-01

160

Comparison of Clinical and Radiographic Characteristics between Nodular Bronchiectatic Form of Nontuberculous Mycobacterial Lung Disease and Diffuse Panbronchiolitis  

OpenAIRE

The nodular bronchiectatic form of nontuberculous mycobacterial (NTM) lung disease and diffuse panbronchiolits (DPB) show similar clinical and radiographic findings. The present study was performed to clarify the clinicoradiographic similarities as well as the differences between NTM lung disease and DPB. The initial clinicoradiographic features of 78 patients with the nodular bronchiectatic form of NTM lung disease (41 patients with Mycobacterium avium complex infection and 37 patients with ...

Park, Hye Yun; Suh, Gee Young; Chung, Man Pyo; Kim, Hojoong; Kwon, O. Jung; Chung, Myung Jin; Kim, Tae Sung; Lee, Kyung Soo; Koh, Won-jung

2009-01-01

161

Structural reorganization of the antigen-binding groove of human CD1b for presentation of mycobacterial sulfoglycolipids  

OpenAIRE

The mechanisms permitting nonpolymorphic CD1 molecules to present lipid antigens that differ considerably in polar head and aliphatic tails remain elusive. It is also unclear why hydrophobic motifs in the aliphatic tails of some antigens, which presumably embed inside CD1 pockets, contribute to determinants for T-cell recognition. The 1.9-Å crystal structure of an active complex of CD1b and a mycobacterial diacylsulfoglycolipid presented here provides some clues. Upon antigen binding, endoge...

Garcia-alles, Luis F.; Collmann, Anthony; Versluis, Cees; Lindner, Buko; Guiard, Julie; Maveyraud, Laurent; Huc, Emilie; Im, Jin S.; Sansano, Sebastiano; Brando, The?re?se; Julien, Sylviane; Prandi, Jacques; Gilleron, Martine; Porcelli, Steven A.; La Salle, Henri

2011-01-01

162

Effect of rifampin and rifabutin on serum itraconazole levels in patients with chronic pulmonary aspergillosis and coexisting nontuberculous mycobacterial infection.  

Science.gov (United States)

We investigated the effects of rifampin and rifabutin on serum itraconazole levels in patients with chronic pulmonary aspergillosis. Serum itraconazole concentrations were significantly lower in patients who received itraconazole with rifampin (median, 0.1 ?g/ml; P itraconazole alone (median, 5.92 ?g/ml). Concomitant use of rifampin or rifabutin and itraconazole should be avoided in patients with chronic pulmonary aspergillosis and coexisting mycobacterial infections. PMID:25313207

Moon, Seong Mi; Park, Hye Yun; Jeong, Byeong-Ho; Jeon, Kyeongman; Lee, Soo-Youn; Koh, Won-Jung

2015-01-01

163

Atypical Mycobacterial Exit-Site Infection and Peritonitis in Peritoneal Dialysis Patients on Prophylactic Exit-Site Gentamicin Cream  

OpenAIRE

We report 9 cases of exit-site infection and continuous ambulatory peritoneal dialysis peritonitis associated with atypical mycobacteria. All patients had been using topical gentamicin cream as prophylaxis for exit-site infection before the onset of these infections. Gentamicin cream is postulated to be a potential risk factor for atypical mycobacterial infection because of selective pressure on other micro-organisms. The microbiology of atypical mycobacteria and the treatment for atypical my...

Lo, Man-wai; Mak, Siu-ka; Wong, Yuk-yi; Lo, Kwok-chi; Chan, Shuk-fan; Tong, Gensy M. W.; Lo, Kin-yee; Wong, Ping-nam; Tse, Cindy W. S.; Kam, Kai-man; Wong, Andrew K. M.

2013-01-01

164

Protection against Mycobacterium avium by DNA Vaccines Expressing Mycobacterial Antigens as Fusion Proteins with Green Fluorescent Protein  

OpenAIRE

Mycobacterium avium causes disseminated disease in humans with AIDS, paratuberculosis in ruminants, lymphadenopathy in swine, and tuberculosis in birds. We constructed DNA vaccines expressing mycobacterial antigens as fusion proteins with enhanced green fluorescent protein (EGFP). Plasmids p65K-EGFP, p85A-EGFP, and p85B-EGFP expressed the M. avium 65-kDa antigen, the Mycobacterium bovis BCG 85A antigen, and the M. avium 85B antigen, respectively, as EGFP fusion proteins. We visualized protein...

Velaz-faircloth, Maria; Cobb, Alison J.; Horstman, Amanda L.; Henry, Stanley C.; Frothingham, Richard

1999-01-01

165

The role of the mycobacterial DNA-binding protein 1 (MDP1) from Mycobacterium bovis BCG in host cell interaction  

OpenAIRE

Background: Mycobacterium tuberculosis differs from most pathogens in its ability to multiply inside monocytes and to persist during long periods of time within granuloma in a status of latency. A class of proteins called mycobacterial histone-like proteins has been associated with regulation of replication and latency, but their precise role in the infection process has yet to be uncovered. Our study aimed at defining the impact of the histone-like protein MDP1 from M. bovis BCG (m...

Kunisch, Ralph; Kamal, Elisabeth; Lewin, Astrid

2012-01-01

166

Diacylated Sulfoglycolipids Are Novel Mycobacterial Antigens Stimulating CD1-restricted T Cells during Infection with Mycobacterium tuberculosis  

OpenAIRE

Mycobacterial lipids comprise a heterogeneous group of molecules capable of inducing T cell responses in humans. To identify novel antigenic lipids and increase our understanding of lipid-mediated immune responses, we established a panel of T cell clones with different lipid specificities. Using this approach we characterized a novel lipid antigen belonging to the group of diacylated sulfoglycolipids purified from Mycobacterium tuberculosis. The structure of this sulfoglycolipid was identifie...

Gilleron, Martine; Stenger, Steffen; Mazorra, Zaima; Wittke, Frederick; Mariotti, Sabrina; Bo?hmer, Gabriele; Prandi, Jacques; Mori, Lucia; Puzo, Germain; Libero, Gennaro

2004-01-01

167

Rapid detection and identification of nontuberculous mycobacterial pathogens in fish by using high-resolution melting analysis  

OpenAIRE

Mycobacterial infections in fish are commonly referred to as piscine mycobacteriosis, irrespectively of the specific identity of the causal organism. They usually cause a chronic disease and sometimes may result in high mortalities and severe economic losses. Nearly 20 species of Mycobacterium have been reported to infect fish. Among them, Mycobacterium marinum, M. fortuitum, and M. chelonae are generally considered the major agents responsible for fish mycobacteriosis. As no quick and inexpe...

Phung, Thu Nguyet; Caruso, Domenico; Godreuil, S.; Keck, N.; Vallaeys, T.; Avarre, Jean-christophe

2013-01-01

168

Unraveling the transcriptional regulatory networks associated to mycobacterial cell wall defective form induction by glycine and lysozyme treatment  

OpenAIRE

It is known that a combined glycine/lysozyme treatment is able to induce in vitro the mycobacterial conversion from the bacillary to the cell wall defective forms. These forms also naturally occur in vivo as a response to various antimicrobial factors such as lysozyme released by phagocytic cells. Although they have been successfully isolated from patients with several chronic diseases, their role in pathogenesis is still unknown, mainly due to the difficulties in handling the in vi...

Rosu, Valentina; Bandino, Ennio; Cossu, Andrea

2013-01-01

169

Targeted delivery of mycobacterial antigens to human dendritic cells via Siglec-7 induces robust T cell activation.  

Science.gov (United States)

Lipids from mycobacteria can be presented to human T cells by group 1 CD1 Ag-presenting molecules (CD1a, CD1b, and CD1c). Group 1 CD1-restricted T cells are activated by lipid Ags presented by myeloid dendritic cells (DCs), after which they generate antibacterial effector functions, including IFN-? secretion and cytolysis. Thus, mycobacterial lipids are being investigated as components of novel vaccines for mycobacterial infections. In this study we show that the mycobacterial lipid Ag C80 glucose-6-monomycolate can be delivered to human CD1b(+) DCs via targeted liposomal nanoparticles, leading to robust group 1 CD1-restricted activation of T cells. Targeting was achieved by decorating the liposomes with a high-affinity glycan ligand of sialic acid-binding Ig-like lectin (Siglec)-7, a siglec receptor expressed on DCs that mediates rapid endocytosis and transport of its cargo to lysosomes. An Ab to Siglec-7 completely blocked the binding of targeted liposomes to human monocyte-derived DCs (Mo-DCs), demonstrating their targeting specificity. Mo-DCs pulsed with targeted liposomes containing C80 glucose-6-monomycolate more potently activated a CD1b-restricted T cell line relative to Mo-DCs pulsed with free lipid Ag or antigenic liposomes without Siglec-7 ligand. These data suggest that the endocytic function of Siglec-7 can be exploited to deliver glycolipid Ags to their target cell and increase the efficiency of display to T cells. PMID:25000981

Kawasaki, Norihito; Rillahan, Cory D; Cheng, Tan-Yun; Van Rhijn, Ildiko; Macauley, Matthew S; Moody, D Branch; Paulson, James C

2014-08-15

170

Molecular basis of mycobacterial lipid antigen presentation by CD1c and its recognition by ?? T cells.  

Science.gov (United States)

CD1c is a member of the group 1 CD1 family of proteins that are specialized for lipid antigen presentation. Despite high cell surface expression of CD1c on key antigen-presenting cells and the discovery of its mycobacterial lipid antigen presentation capability, the molecular basis of CD1c recognition by T cells is unknown. Here we present a comprehensive functional and molecular analysis of ?? T-cell receptor (TCR) recognition of CD1c presenting mycobacterial phosphomycoketide antigens. Our structure of CD1c with the mycobacterial phosphomycoketide (PM) shows similarities to that of CD1c-mannosyl-?1-phosphomycoketide in that the A' pocket accommodates the mycoketide alkyl chain; however, the phosphate head-group of PM is shifted ?6 Å in relation to that of mannosyl-?1-PM. We also demonstrate a bona fide interaction between six human TCRs and CD1c-mycoketide complexes, measuring high to moderate affinities. The crystal structure of the DN6 TCR and mutagenic studies reveal a requirement of five complementarity determining region (CDR) loops for CD1c recognition. Furthermore, mutagenesis of CD1c reveals residues in both the ?1 and ?2 helices involved in TCR recognition, yet not entirely overlapping among the examined TCRs. Unlike patterns for MHC I, no archetypical binding footprint is predicted to be shared by CD1c-reactive TCRs, even when recognizing the same or similar antigens. PMID:25298532

Roy, Sobhan; Ly, Dalam; Li, Nan-Sheng; Altman, John D; Piccirilli, Joseph A; Moody, D Branch; Adams, Erin J

2014-10-28

171

Anti-mycobacterial activity of root and leaf extracts of Anthocleista djalonensis (Loganiaceae and Diospyros mespiliformis (Ebenaceae  

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Full Text Available We screened the aqueous and methanol leaf and root extracts of Anthocleista djalonensis, Diospyros mespiliformis, and their combinations for possible anti-mycobacterial activities using Mycobacterium smegmatis as a surrogate screen. These plants are reputed among folk practices as potent remedy in the management of tuberculosis and leprosy cases. In the sensitivity screening study, only the methanol extracts of A. djalonensis and D. mespiliformis showed anti-mycobacterial activity, while the aqueous extracts exhibited no inhibitory activity on M. smegmatis. The minimum inhibitory concentration (MIC of the methanol leaf and root extract of A. djalonensis against M. smegmatis were 125 ?g/ml. The MIC of the methanol leaf and root extracts of D. mespiliformis is 167 and 250 ?g/ml, respectively. In the interaction studies, four out of nine decimal combinations of the two medicinal plant extracts exhibited synergism with fractional inhibitory concentration indices < 1 and a negative activity index values. The 8:2 ratio of D. mespiliformis and A. djalonensis exhibited the greatest degree of antimycobacterial synergy against M. smegmatis. The result of this study supports the claims of efficacy reported in the folk use of these plants in mycobacterial infection and the plants could therefore be investigated further and harnessed as potent antimycobacterial agents.

Esimone Charles

2009-01-01

172

Increased serum anti-mycobacterial antibody titers in rheumatoid arthritis patients: Is there any specific antigenic target?  

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Objective was to investigate the presence of immunoreactivity against mycobacterial antigens in the sera of patients with rheumatoid arthritis (Ra) and to detect the target of the immune reaction. This study was carried out on 60 patients with RA, and 25 patients with no joint diseases in the laboratory of Clinical Microbiology Department of Ankara University Medical Faculty, Ankara, Turkey between July 2003 to January 2004. Secreted and cellular antigens of Mycobacterium tuberculosis (M. tuberculosis) H37Rv and Mycobacterium bovis (M. bovis) were isolated and purified by high performance liquid chromatography to antigenic fractions. The immunoreactivity of patient and control sera against these antigens were determined by enzyme-linked immunosorbent assay (ELISA). Immunoreactivity against mycobacterial antigens in RA patients were significantly higher than controls. Significant difference between patients and controls has been determined with M. bovis Bacillus Calmette Guerin (BCG) culture fluid and sonicate antigens, but not with M. tuberculosis H37Rv. This suggests that the antigen triggering immune response in patients with RA may belong to or mainly expressed on M. bovis BCG. The ELISA results showed significant difference between RA patients and controls with all antigenic fractions. Presence of increased immunoreactivity against mycobacterial antigens in the sera of patients with RA was detected. When statistical analysis was considered, we cannot put forward ais was considered, we cannot put forward any antigenic fraction alone as the one responsible for the increased reactivity. (author)

173

Detection and differentiation of Mycobacterium tuberculosis and nontuberculous mycobacterial isolates by real-time PCR.  

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Mycobacteria cause a variety of illnesses that differ in severity and public health implications. The differentiation of Mycobacterium tuberculosis from nontuberculous mycobacteria (NTM) is of primary importance for infection control and choice of antimicrobial therapy. Despite advances in molecular diagnostics, the ability to rapidly diagnose M. tuberculosis infections by PCR is still inadequate, largely because of the possibility of false-negative reactions. We designed and validated a real-time PCR for mycobacteria by using the LightCycler system with 18 reference strains and 168 clinical mycobacterial isolates. All clinically significant mycobacteria were detected; the mean melting temperatures (with 99.9% confidence intervals [99.9% CI] in parentheses) for the different mycobacteria were as follows: M. tuberculosis, 64.35 degrees C (63.27 to 65.42 degrees C); M. kansasii, 59.20 degrees C (58.07 to 60.33 degrees C); M. avium, 57.82 degrees C (57.05 to 58.60 degrees C); M. intracellulare, 54.46 degrees C (53.69 to 55.23 degrees C); M. marinum, 58.91 degrees C (58.28 to 59.55 degrees C); rapidly growing mycobacteria, 53.09 degrees C (50.97 to 55.20 degrees C) or 43.19 degrees C (42.19 to 44.49 degrees C). This real-time PCR assay with melting curve analysis consistently accurately detected and differentiated M. tuberculosis from NTM. Detection of an NTM helps ensure that the negative result for M. tuberculosis is a true negative. The specific melting temperature also provides a suggestion of the identity of the NTM present, when the most commonly encountered mycobacterial species are considered. In a parallel comparison, both the LightCycler assay and the COBAS Amplicor M. tuberculosis assay correctly categorized 48 of 50 specimens that were proven by culture to contain M. tuberculosis, and the LightCycler assay correctly characterized 3 of 3 specimens that contained NTM. PMID:14605148

Shrestha, Nabin K; Tuohy, Marion J; Hall, Gerri S; Reischl, Udo; Gordon, Steven M; Procop, Gary W

2003-11-01

174

Drug treatment of pulmonary nontuberculous mycobacterial disease in HIV-negative patients: the evidence.  

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Pulmonary disease (PD) caused by nontuberculous mycobacteria is an emerging infection mainly in countries where the incidence of tuberculosis is in decline. It affects an elderly population, often with underlying chronic lung diseases, but its epidemiology shows significant regional variation. Guidelines and recommendations for treatment of these infections exist, but build strongly on expert opinion, as very few good quality clinical trials have been performed in this field. Only for the most frequent causative agents, the Mycobacterium avium complex, Mycobacterium kansasii and Mycobacterium abscessus, a reasonable number of trials and case series is now available. For the less frequent causative agents of pulmonary nontuberculous mycobacterial (NTM) disease (Mycobacterium xenopi, Mycobacterium malmoense, Mycobacterium fortuitum, Mycobacterium chelonae) data is mostly limited to a few very small case series. Within this review, we have collected and combined evidence from all available trials and case series. From the data of these trials and case series, we reconstruct a more evidence-based overview of possible drug treatment regimens and their outcomes. PMID:24124798

van Ingen, Jakko; Ferro, Beatriz E; Hoefsloot, Wouter; Boeree, Martin J; van Soolingen, Dick

2013-10-01

175

The cost of medical management of pulmonary nontuberculous mycobacterial disease in Ontario, Canada.  

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Treatment of pulmonary nontuberculous mycobacterial (NTM) infection is complex, requiring multiple antibiotics and a prolonged treatment course. We determined the monthly cost of treating patients with pulmonary NTM infections in our clinic, a tertiary care centre in Toronto, Ontario, Canada. We reviewed records of a single clinic at the University Health Network (Toronto) for all patients with pulmonary NTM isolates. Pharmacological and nonpharmacological treatment costs were calculated using a number of Canadian references. 172 patients were reviewed, 91 of whom were treated pharmacologically. The median total duration and cost per treated patient were 14 months (interquartile range (IQR) 9-23 months) and CAD 4,916 (IQR CAD 2,934-9,063), respectively. Median monthly drug treatment cost was CAD 321 (IQR CAD 254-458) for all patients, CAD 289 (IQR CAD 237-341) for patients receiving exclusively oral antibiotics and CAD 1,161 (IQR CAD 795-1,646) for patients whose treatment included i.v. antibiotics. The most costly oral regiment consisted of a fluroquinolone, macrolide and rifampin. In multivariable analysis, Mycobacterium abscessus infection, i.v. therapy and Mycobacterium xenopi infection were all associated with increased monthly treatment costs. The direct medical costs of NTM infections are substantial. Less expensive alternative therapies might be most helpful for M. abscessus infection and when i.v. antibiotics are deemed necessary. PMID:20817704

Leber, A; Marras, T K

2011-05-01

176

Exosomes carrying mycobacterial antigens can protect mice against Mycobacterium tuberculosis infection.  

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Approximately 2 billion people are infected with Mycobacterium tuberculosis, the etiological agent of tuberculosis (TB), and an estimated 1.5 million individuals die annually from TB. Presently, Mycobacterium bovis BCG remains the only licensed TB vaccine; however, previous studies suggest its protective efficacy wanes over time and fails in preventing pulmonary TB. Therefore, a safe and effective vaccine is urgently required to replace BCG or boost BCG immunizations. Our previous studies revealed that mycobacterial proteins are released via exosomes from macrophages infected with M. tuberculosis or pulsed with M. tuberculosis culture filtrate proteins (CFP). In the present study, exosomes purified from macrophages treated with M. tuberculosis CFP were found to induce antigen-specific IFN-? and IL-2-expressing CD4(+) and CD8(+) T cells. In exosome-vaccinated mice, there was a similar TH1 immune response but a more limited TH2 response compared to BCG-vaccinated mice. Using a low-dose M. tuberculosis mouse aerosol infection model, exosomes from CFP-treated macrophages were found to both prime a protective immune response as well as boost prior BCG immunization. The protection was equal to or superior to BCG. In conclusion, our findings suggest that exosomes might serve as a novel cell-free vaccine against an M. tuberculosis infection. PMID:23943377

Cheng, Yong; Schorey, Jeffery S

2013-12-01

177

The feasibility of sputum transportation system in china: effect of sputum storage on the mycobacterial detection.  

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Sputum transportation from county-level to prefecture-level is an ideal strategy to cover the shortage of the laboratory capability in the resource-poor setting. Here, we firstly evaluated the feasibility of sputum transportation system in China by analyzing the culture and molecular diagnosis results from 1982 smear-positive patients with different delay in processing for culture. In this study, the total contamination rate was 2.32% and the total smear positive/culture negative (S+/C-) rate was 7.57%. We found that sputum specimens refrigerated for no more than 7 d before mycobacterial detection did not affect culture significantly. In addition, the invalid result rates among 0-3 d, 3-7 d, and 7+ d group were 3.63%, 3.14%, and 12.48%, respectively. Statistic analysis revealed that molecular diagnostic results while the invalid result rate of genechip for the specimen with more than 7 d delay was significantly higher (P<0.001). The refrigerators equipped in county laboratories, transport at low temperature and frequent transport services once a week will ensure the feasibility of sputum transportation system in China. PMID:25484017

Pang, Yu; DU, Jian; Zhang, Zhi Ying; Ou, Xi Chao; Li, Qiang; Xia, Hui; Qu, Yan; Zhao, Yan Lin

2014-12-01

178

Pre-Columbian mycobacterial genomes reveal seals as a source of New World human tuberculosis.  

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Modern strains of Mycobacterium tuberculosis from the Americas are closely related to those from Europe, supporting the assumption that human tuberculosis was introduced post-contact. This notion, however, is incompatible with archaeological evidence of pre-contact tuberculosis in the New World. Comparative genomics of modern isolates suggests that M. tuberculosis attained its worldwide distribution following human dispersals out of Africa during the Pleistocene epoch, although this has yet to be confirmed with ancient calibration points. Here we present three 1,000-year-old mycobacterial genomes from Peruvian human skeletons, revealing that a member of the M. tuberculosis complex caused human disease before contact. The ancient strains are distinct from known human-adapted forms and are most closely related to those adapted to seals and sea lions. Two independent dating approaches suggest a most recent common ancestor for the M. tuberculosis complex less than 6,000 years ago, which supports a Holocene dispersal of the disease. Our results implicate sea mammals as having played a role in transmitting the disease to humans across the ocean. PMID:25141181

Bos, Kirsten I; Harkins, Kelly M; Herbig, Alexander; Coscolla, Mireia; Weber, Nico; Comas, Iñaki; Forrest, Stephen A; Bryant, Josephine M; Harris, Simon R; Schuenemann, Verena J; Campbell, Tessa J; Majander, Kerttu; Wilbur, Alicia K; Guichon, Ricardo A; Wolfe Steadman, Dawnie L; Cook, Della Collins; Niemann, Stefan; Behr, Marcel A; Zumarraga, Martin; Bastida, Ricardo; Huson, Daniel; Nieselt, Kay; Young, Douglas; Parkhill, Julian; Buikstra, Jane E; Gagneux, Sebastien; Stone, Anne C; Krause, Johannes

2014-10-23

179

[Uncommon mycobacterial infections in domestic and zoo animals: four cases with special emphasis on pathology].  

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Infections caused by classical tubercle bacilli are rare during the last years. Nevertheless, diseases caused by other mycobacteria have to be considered clinically and in diagnostic pathology especially in cases of immunosuppression and due to their potential zoonosis risk. An infection by mycobacteria was diagnosed in four animals (Mayotte Maki, Blue-headed Parrot, Patagonian sealion, Beagle) necropsied between 1995 and 2002 in the Institute of Veterinary-Pathology of the University of Leipzig. The Maki, the blue-headed parrot and the dog showed a disseminated character of the disease caused by Mycobacterium genavense (monkey and bird) resp. Mycobacterium avium (dog), while an open chronical tuberculosis of the lungs due to a pathogenic member of Mycobacterium tuberculosis complex was observed in the seal. All these bacteria are potential causes of zoonoses. So, if granulomatous or disseminated histiocytic alterations are detected in diagnostic pathology, mycobacterial infections should always be included in differential diagnoses and require careful aetiological investigations by histopathological and bacteriological methods. PMID:14560447

Steiger, K; Ellenberger, C; Schüppel, K F; Richter, E; Schmerbach, K; Krautwald-Junghanns, M E; Wünnemann, K; Eulenberger, K; Schoon, H A

2003-09-01

180

Mycobacterial envelope lipids fingerprint from direct MALDI-TOF MS analysis of intact bacilli.  

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Mycobacterium tuberculosis (Mtb) lipids including glycolipids and lipoglycans play a crucial role in the modulation of the host immune response by targeting the innate receptors C-type lectins, TLRs and the CD1 proteins of class 1. Glycolipids have been shown to be biomarkers of M. tuberculosis strains and also of opportunistic mycobacteria called non-tuberculous mycobacteria. Most of the structural and functional work of the Mtb lipids has been done using lipids arising from M. tuberculosis cell growth in vitro. However it is likely that lipid structures can change during infection or among the M. tuberculosis or opportunistic clinical strains. Here we describe a new, rapid and sensitive analysis of lipids directly on whole mycobacteria which can be done in few minutes and on less than 1000 mycobacteria by direct matrix-assisted laser desorption/ionization mass spectrometry using an unusual solvent matrix. By this new methodology, which does not require extraction or purification steps, we are able to discriminate mycobacteria belonging to the Mtb complex as well as opportunistic and non-pathogenic mycobacteria. This method was also found to be successful for identification of an envelope lipid mutant. This work opens a new analytical route for in vivo analysis of mycobacterial lipids. PMID:25488848

Larrouy-Maumus, Gérald; Puzo, Germain

2015-01-01

181

Detection and identification of non-tuberculous mycobacterial infections in 6,472 tuberculosis suspected patients.  

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Between March 1993 and March 1994, 82 patients with infection by non-tuberculous mycobacteria (NTM) and 443 patients with tuberculosis (TB) were registered among 6,472 patients with suspected tuberculosis. Skin-test reactivity to purified protein derivative (PPD) in patients demonstrated indurations of 10-14 mm or more for the majority of patients in both groups. Most patients with NTM infection had abnormal chest roentgenograms showing sporadic infiltrations, nodular abscesses, and cavities resembling TB radiological evidence. The similarity in age range, PPD skin reaction, and radiological evidence in patients infected with NTM or Mycobacterium tuberculosis (MTB) can mislead the physician. Some NTM species were recovered more often than others. Mycobacterium fortuitum from 22 clinical specimens (26.8%); Mycobacterium gastric 19 (23.1%); and Mycobacterium terrae complex 15 (18.3%). The antimicrobial drug susceptibility tests of the isolated organisms showed that 42 (9.5%) isolates of MTB were resistant to isoniazid and 31 (7.0%) to streptomycin. a few strains (1.3%) were identified as being resistant to a combination of 3 primary drugs. These findings suggest that drug-resistant mycobacterial infections are becoming an important problem in the region. PMID:8863361

Bahrmand, A R; Madani, H; Samar, G; Khalilzadeh, L; Bakayev, V V; Yaghli, M; babaei, M H

1996-01-01

182

Structure of mycobacterial maltokinase, the missing link in the essential GlgE-pathway  

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A novel four-step pathway identified recently in mycobacteria channels trehalose to glycogen synthesis and is also likely involved in the biosynthesis of two other crucial polymers: intracellular methylglucose lipopolysaccharides and exposed capsular glucan. The structures of three of the intervening enzymes - GlgB, GlgE, and TreS - were recently reported, providing the first templates for rational drug design. Here we describe the structural characterization of the fourth enzyme of the pathway, mycobacterial maltokinase (Mak), uncovering a eukaryotic-like kinase (ELK) fold, similar to methylthioribose kinases and aminoglycoside phosphotransferases. The 1.15?Å structure of Mak in complex with a non-hydrolysable ATP analog reveals subtle structural rearrangements upon nucleotide binding in the cleft between the N- and the C-terminal lobes. Remarkably, this new family of ELKs has a novel N-terminal domain topologically resembling the cystatin family of protease inhibitors. By interfacing with and restraining the mobility of the phosphate-binding region of the N-terminal lobe, Mak's unusual N-terminal domain might regulate its phosphotransfer activity and represents the most likely anchoring point for TreS, the upstream enzyme in the pathway. By completing the gallery of atomic-detail models of an essential pathway, this structure opens new avenues for the rational design of alternative anti-tubercular compounds. PMID:25619172

Fraga, Joana; Maranha, Ana; Mendes, Vitor; Pereira, Pedro José Barbosa; Empadinhas, Nuno; Macedo-Ribeiro, Sandra

2015-01-01

183

Occurrence of Nontuberculous Mycobacterial Pulmonary Infection in an Endemic Area of Tuberculosis  

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The majority of investigations of the epidemiology of nontuberculous mycobacteria (NTM) have focused on highly developed nations with a low prevalence of tuberculosis. In contrast, the Para state of north Brazil represents an area of high tuberculosis prevalence and increasing NTM incidence. Toward the goal of understanding the dynamics of infection by all Mycobacterium species, we report patient characteristics and the identification of NTM strains isolated from sputum samples from patients that were residents of Para, a state in the Amazon region, Northern of Brazil, over the period January 2010 through December 2011 (2 years). The 29 NTM patients comprised 13.5% of positive mycobacterial cultures over the 2-year period. A major risk factor for NTM pulmonary disease was previous tuberculosis (76%). Further, the average age of NTM patients (52 years) was significantly higher than that of tuberculosis patients (39 years) and more were female (72.4% vs. 37.4%). Unlike other Brazilian states, NTM pulmonary patients in Para were infected with a different spectrum of mycobacteria; primarily the rapidly growing Mycobacterium massiliense and Mycobacterium simiae complex. PMID:23875055

da Costa, Ana Roberta Fusco; Falkinham, Joseph O.; Lopes, Maria Luiza; Barretto, Adriana Rodrigues; Felicio, João Soares; Sales, Lúcia Helena Messias; Bahia, Jeann Ricardo da Costa; Conceição, Emilyn Costa; Lima, Karla Valéria Batista

2013-01-01

184

Autoimmunity, hypogammaglobulinemia, lymphoproliferation, and mycobacterial disease in patients with activating mutations in STAT3.  

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The signal transducer and activator of transcription (STAT) family of transcription factors orchestrate hematopoietic cell differentiation. Recently, mutations in STAT1, STAT5B, and STAT3 have been linked to development of immunodysregulation polyendocrinopathy enteropathy X-linked-like syndrome. Here, we immunologically characterized 3 patients with de novo activating mutations in the DNA binding or dimerization domains of STAT3 (p.K392R, p.M394T, and p.K658N, respectively). The patients displayed multiorgan autoimmunity, lymphoproliferation, and delayed-onset mycobacterial disease. Immunologically, we noted hypogammaglobulinemia with terminal B-cell maturation arrest, dendritic cell deficiency, peripheral eosinopenia, increased double-negative (CD4(-)CD8(-)) T cells, and decreased natural killer, T helper 17, and regulatory T-cell numbers. Notably, the patient harboring the K392R mutation developed T-cell large granular lymphocytic leukemia at age 14 years. Our results broaden the spectrum of phenotypes caused by activating STAT3 mutations, highlight the role of STAT3 in the development and differentiation of multiple immune cell lineages, and strengthen the link between the STAT family of transcription factors and autoimmunity. PMID:25349174

Haapaniemi, Emma M; Kaustio, Meri; Rajala, Hanna L M; van Adrichem, Arjan J; Kainulainen, Leena; Glumoff, Virpi; Doffinger, Rainer; Kuusanmäki, Heikki; Heiskanen-Kosma, Tarja; Trotta, Luca; Chiang, Samuel; Kulmala, Petri; Eldfors, Samuli; Katainen, Riku; Siitonen, Sanna; Karjalainen-Lindsberg, Marja-Liisa; Kovanen, Panu E; Otonkoski, Timo; Porkka, Kimmo; Heiskanen, Kaarina; Hänninen, Arno; Bryceson, Yenan T; Uusitalo-Seppälä, Raija; Saarela, Janna; Seppänen, Mikko; Mustjoki, Satu; Kere, Juha

2015-01-22

185

Autoimmunity, hypogammaglobulinemia, lymphoproliferation, and mycobacterial disease in patients with activating mutations in STAT3  

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The signal transducer and activator of transcription (STAT) family of transcription factors orchestrate hematopoietic cell differentiation. Recently, mutations in STAT1, STAT5B, and STAT3 have been linked to development of immunodysregulation polyendocrinopathy enteropathy X-linked–like syndrome. Here, we immunologically characterized 3 patients with de novo activating mutations in the DNA binding or dimerization domains of STAT3 (p.K392R, p.M394T, and p.K658N, respectively). The patients displayed multiorgan autoimmunity, lymphoproliferation, and delayed-onset mycobacterial disease. Immunologically, we noted hypogammaglobulinemia with terminal B-cell maturation arrest, dendritic cell deficiency, peripheral eosinopenia, increased double-negative (CD4?CD8?) T cells, and decreased natural killer, T helper 17, and regulatory T-cell numbers. Notably, the patient harboring the K392R mutation developed T-cell large granular lymphocytic leukemia at age 14 years. Our results broaden the spectrum of phenotypes caused by activating STAT3 mutations, highlight the role of STAT3 in the development and differentiation of multiple immune cell lineages, and strengthen the link between the STAT family of transcription factors and autoimmunity. PMID:25349174

Haapaniemi, Emma M.; Kaustio, Meri; Rajala, Hanna L. M.; van Adrichem, Arjan J.; Kainulainen, Leena; Glumoff, Virpi; Doffinger, Rainer; Kuusanmäki, Heikki; Heiskanen-Kosma, Tarja; Trotta, Luca; Chiang, Samuel; Kulmala, Petri; Eldfors, Samuli; Katainen, Riku; Siitonen, Sanna; Karjalainen-Lindsberg, Marja-Liisa; Kovanen, Panu E.; Otonkoski, Timo; Porkka, Kimmo; Heiskanen, Kaarina; Hänninen, Arno; Bryceson, Yenan T.; Uusitalo-Seppälä, Raija; Saarela, Janna; Seppänen, Mikko; Kere, Juha

2015-01-01

186

Expression and Immunogenicity of the Mycobacterial Ag85B/ESAT-6 Antigens Produced in Transgenic Plants by Elastin-Like Peptide Fusion Strategy  

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Full Text Available This study explored a novel system combining plant-based production and the elastin-like peptide (ELP fusion strategy to produce vaccinal antigens against tuberculosis. Transgenic tobacco plants expressing the mycobacterial antigens Ag85B and ESAT-6 fused to ELP (TBAg-ELP were generated. Purified TBAg-ELP was obtained by the highly efficient, cost-effective, inverse transition cycling (ICT method and tested in mice. Furthermore, safety and immunogenicity of the crude tobacco leaf extracts were assessed in piglets. Antibodies recognizing mycobacterial antigens were produced in mice and piglets. A T-cell immune response able to recognize the native mycobacterial antigens was detected in mice. These findings showed that the native Ag85B and ESAT-6 mycobacterial B- and T-cell epitopes were conserved in the plant-expressed TBAg-ELP. This study presents the first results of an efficient plant-expression system, relying on the elastin-like peptide fusion strategy, to produce a safe and immunogenic mycobacterial Ag85B-ESAT-6 fusion protein as a potential vaccine candidate against tuberculosis.

Udo Conrad

2010-01-01

187

CFP10 and ESAT6 aptamers as effective Mycobacterial antigen diagnostic reagents.  

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The development of effective Mycobacterial antigen diagnostic reagents remains a high priority. The 6-kDa early secreted antigenic target (ESAT6) and 10-kDa culture filtrate protein (CFP10) are secreted early by virulent Mycobacterium tuberculosis (M. tb) and are not present in the non-virulent Bacillus Calmette-Guerin (BCG). In this study, we used a Systematic Evolution of Ligands by Exponential Enrichment (SELEX) technique to screen for a functional ssDNA aptamer "antibody" that specifically bound to ESAT6-CFP10 (CE) protein. The selected single ssDNA aptamers (CE24 and CE15) demonstrated the highest specificity and binding affinity to CFP10 (CE24: Kd = 3.75 × 10(-7) M) and ESAT6 (CE15: Kd = 1.6 × 10(-7) M). We further detected CFP10 and ESAT6 proteins in serum samples from active pulmonary tuberculosis (TB) patients, extrapulmonary TB patients and healthy donors by using an enzyme-linked oligonucleotide assay (ELONA). The results showed that the sensitivity and specificity were 100% and 94.1% (using CE24 aptamer-based ELONA) and 89.6% and 94.1% (using CE15 aptamer-based ELONA), respectively. A good correlation was observed between aptamer-based ELONA and T-SPOT TB assay. Thus, our study suggests that CE24 and CE15 have potentially broad applications as early antigen diagnostic agents not only for active pulmonary TB, extrapulmonary TB, but also possibly for latent TB infection and TB with immune-deficiency. PMID:24968239

Tang, Xiao-Lei; Zhou, Ya-Xiong; Wu, Si-Min; Pan, Qin; Xia, Bing; Zhang, Xiao-Lian

2014-12-01

188

Bacillus calmette-guerin infection in NADPH oxidase deficiency: defective mycobacterial sequestration and granuloma formation.  

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Patients with chronic granulomatous disease (CGD) lack generation of reactive oxygen species (ROS) through the phagocyte NADPH oxidase NOX2. CGD is an immune deficiency that leads to frequent infections with certain pathogens; this is well documented for S. aureus and A. fumigatus, but less clear for mycobacteria. We therefore performed an extensive literature search which yielded 297 cases of CGD patients with mycobacterial infections; M. bovis BCG was most commonly described (74%). The relationship between NOX2 deficiency and BCG infection however has never been studied in a mouse model. We therefore investigated BCG infection in three different mouse models of CGD: Ncf1 mutants in two different genetic backgrounds and Cybb knock-out mice. In addition, we investigated a macrophage-specific rescue (transgenic expression of Ncf1 under the control of the CD68 promoter). Wild-type mice did not develop severe disease upon BCG injection. In contrast, all three types of CGD mice were highly susceptible to BCG, as witnessed by a severe weight loss, development of hemorrhagic pneumonia, and a high mortality (? 50%). Rescue of NOX2 activity in macrophages restored BCG resistance, similar as seen in wild-type mice. Granulomas from mycobacteria-infected wild-type mice generated ROS, while granulomas from CGD mice did not. Bacterial load in CGD mice was only moderately increased, suggesting that it was not crucial for the observed phenotype. CGD mice responded with massively enhanced cytokine release (TNF-?, IFN-?, IL-17 and IL-12) early after BCG infection, which might account for severity of the disease. Finally, in wild-type mice, macrophages formed clusters and restricted mycobacteria to granulomas, while macrophages and mycobacteria were diffusely distributed in lung tissue from CGD mice. Our results demonstrate that lack of the NADPH oxidase leads to a markedly increased severity of BCG infection through mechanisms including increased cytokine production and impaired granuloma formation. PMID:25188296

Deffert, Christine; Schäppi, Michela G; Pache, Jean-Claude; Cachat, Julien; Vesin, Dominique; Bisig, Ruth; Ma Mulone, Xiaojuan; Kelkka, Tiina; Holmdahl, Rikard; Garcia, Irene; Olleros, Maria L; Krause, Karl-Heinz

2014-09-01

189

CT findings of mycobacterial infection other than tuberculosis : comparison with tuberculosis  

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To compare the CT findings of mycobacterial infection other than tuberculosis (MOTT) with those of tuberculosis (TB). The chest CT scans of 30 immunocompetent patients with culture-proven pulmonary MOTT (M:F =3D 11:19; mean age, 51.2 yrs.) and of 24 patients with active tuberculosis (M:F 12:12; mean age, 42.5 yrs.) were analyzed by two radiologists; decisions were reached by consensus. Common findings for both MOTT and TB included brochogenically-spread bronchogenic spread nodular lesion (93.3% for MOTT, 100% for TB), bronchiectasis (90%, 83.3%), bronchial wall thickening (66.7%, 54.2%), granuloma (63.3%, 75%), parenchymal scarring (53.3%, 54.2%), and mediastinal lymphadenopathy (50%, 37.5%). Less commonly observed findings were emphysema (46.7%, 29.7%), atelectasis (36.7%, 29.2%), narrowing of a major airway (23.3%, 25%), consolidation (23.3%, 29.2%)and pleural disease (16.7%, 29.2%). Except for cavity (30%, 53.3%; P less than 0.05), the frequencies of each finding were not different between the two groups. A lobe-matched frequency comparison showed that only bronchiectasis in the right middle lobe (40%, 16.7%), right lower lobe (63.3%, 33.3%) and lingula division (53.3%, 25%) was significantly more common in MOTT than in TB (p less than 0.05). The number of lobes in which bronchiectasis and bronchial wall thickening were involved was greater in MOTT (3.20) than in TB (2.04) (p=3D 0.011). Although the CT findings of MOTT and TB overlap considerably, cavities are more common in TB, while in MOTT, bronchiectasis in the lower lung zone is more common and bronchiectasis tends to be more extensive. (author)

Yoon, Chang Jin; Goo, Jin Mo; Seo, Joon Beom; Kim, Se Hyung; Im, Jung Gi [College of Medicine and the Institute of Radiation Medicine, Seoul National University, Seoul (Korea, Republic of)

2000-03-01

190

Epidemiology of Nontuberculous Mycobacterial Infections and Associated Chronic Macrolide Use among Persons with Cystic Fibrosis  

Science.gov (United States)

Rationale: Persons with cystic fibrosis (CF) are at high risk of nontuberculous mycobacterial (NTM) infection, with treatment requiring prolonged multidrug regimens that include macrolides. Although macrolides, specifically azithromycin, are used in the management of patients with CF with chronic Pseudomonas, macrolide-resistant NTM infections are of growing concern. Objectives: To evaluate the relationship between chronic macrolide use and NTM infection among patients with CF included in the 2011 CF Patient Registry (CFPR). Methods: We performed a nested case-control study: incident NTM cases were persons aged more than 5 years with at least one positive culture for NTM in 2011. Controls were persons with negative cultures in 2010 and 2011. Measurements and Main Results: The 2011 CFPR included 27,112 patients; 5,403 (20%) were cultured for mycobacteria in 2010–2011 and met all inclusion criteria. Of these, 191 (4%) were NTM-positive in 2011 only (cases); 5,212 (96%) were NTM-negative in 2010 and 2011 (control subjects). Among the cases, 122 (64%) were culture-positive for Mycobacterium avium complex (MAC) and 69 (36%) for M. abscessus. Azithromycin use in 2010 was less frequently reported among MAC cases (57%; odds ratio = 0.7, P < 0.05) and M. abscessus cases (51%; odds ratio = 0.5, P < 0.01) than in control subjects (66%). Among adolescents and adults, patients with the greatest number of years on chronic macrolides were the least likely to develop incident NTM in 2011 (P < 0.01). Conclusions: Patients with incident NTM infections from either MAC or M. abscessus were less likely to have had chronic azithromycin treatment in the past year. However, because macrolide monotherapy may lead to macrolide resistance, routine screening for NTM should be considered for persons with CF. PMID:23927602

Binder, Alison M.; Olivier, Kenneth N.; Prevots, D. Rebecca

2013-01-01

191

Diffuse Pulmonary Uptake of Tc-99m Methylene Diphosphonate in a Patient with Non-tuberculosis Mycobacterial Infection  

International Nuclear Information System (INIS)

Extra-osseous uptake of bone-seeking radiopharmaceuticals has been reported at various sites and it is known to be induced by various causes. Diffuse pulmonary infection, such as tuberculosis, can be a cause of lung uptake of bone-scan agent. Here we report on a patient with non-tuberculosis mycobacterial infection (NTM) who demonstrated diffuse pulmonary uptake on Tc-99m MDP bone scan. After medical treatment for NTM, the patient's lung lesions improved. Estra skeletal lung Tc-99m MDP uptake on bone scan may suggest lung parenchymal damage associated with disease activity.

192

Characterization of the Mycobacterial AdnAB DNA Motor Provides Insights into the Evolution of Bacterial Motor-Nuclease Machines*  

OpenAIRE

Mycobacterial AdnAB exemplifies a family of heterodimeric motor-nucleases involved in processing DNA double strand breaks (DSBs). The AdnA and AdnB subunits are each composed of an N-terminal UvrD-like motor domain and a C-terminal RecB-like nuclease module. Here we conducted a biochemical characterization of the AdnAB motor, using a nuclease-inactivated heterodimer. AdnAB is a vigorous single strand DNA (ssDNA)-dependent ATPase (kcat 415 s?1), and the affinity of the motor for the ssDNA co...

Unciuleac, Mihaela-carmen; Shuman, Stewart

2010-01-01

193

The application of luciferase as a reporter of environmental regulation of gene expression in mycobacteria.  

Science.gov (United States)

This paper describes the construction of pSG10, the first mycobacterial promoter probe shuttle vector to use the structural gene of a bacterial luciferase as a reporter gene. To examine the utility of using bacterial luciferase to measure gene expression in mycobacteria, the authors have used this vector to monitor the induction of the acetamidase gene promoter of Mycobacterium smegmatis. Luciferase proved to be a rapid, sensitive and easily assayable reporter of changes in gene activity in response to environment in mycobacteria. PMID:7765445

Gordon, S; Parish, T; Roberts, I S; Andrew, P W

1994-11-01

194

Double Strand Break Unwinding and Resection by the Mycobacterial Helicase-Nuclease AdnAB in the Presence of Single Strand DNA-binding Protein (SSB)*  

OpenAIRE

Mycobacterial AdnAB is a heterodimeric DNA helicase-nuclease and 3? to 5? DNA translocase implicated in the repair of double strand breaks (DSBs). The AdnA and AdnB subunits are each composed of an N-terminal motor domain and a C-terminal nuclease domain. Inclusion of mycobacterial single strand DNA-binding protein (SSB) in reactions containing linear plasmid dsDNA allowed us to study the AdnAB helicase under conditions in which the unwound single strands are coated by SSB and thereby pre...

Unciuleac, Mihaela-carmen; Shuman, Stewart

2010-01-01

195

The role of multiple SOS boxes upstream of the Mycobacterium tuberculosis lexA gene--identification of a novel DNA-damage-inducible gene.  

Science.gov (United States)

Four potential binding sites for LexA were identified upstream of the Mycobacterium tuberculosis lexA gene. A mutational analysis of these sites in a lexA-lacZ reporter construct revealed that only one of these SOS boxes was required for DNA-damage-mediated regulation of lexA expression. A novel DNA-damage-inducible gene, Rv2719c, was identified that was divergently transcribed relative to lexA; the other three SOS boxes were found to be involved in regulating expression of this novel mycobacterial-specific gene. The SOS boxes lay in the respective promoter regions of the genes that they regulated. PMID:12427951

Dullaghan, Edith M; Brooks, Patricia C; Davis, Elaine O

2002-11-01

196

Correlation Between Microscopic Examination and Culture for Detection and Differentiation of Mycobacterial Isolates from Cattle in the Sudan  

Directory of Open Access Journals (Sweden)

Full Text Available One hundred and sixty seven caseated lymph nodes and tuberculous lungs were collected from cattle slaughtered in various locations in Khartoum State (Sudan and examined bacteriologically. Microscopic examination using Zeilh-Neelsen stain and culture on Lowenstein-Jensen (L-J medium were carried out to detect and differentiate between the mycobacterial species in the specimens. Thirty-five, 35 (20.96% samples were found to harbor acid-fast bacteria when examined microscopically. Out of the 35 acid-fast bacteria, 22 (62.86% showed branching filaments and were identified as Mycobacterium farcinogenes. The remaining 13(37.14% were bacilli and identified as Mycobacterium bovis. The 35 specimens that proved to harbor acid fast bacteria were cultured on L-J medium. None of the 22 specimens with branching filaments (Mycobacterium farcinogenes grew on L-J medium when incubated aerobically at 37 ° C between four and eight weeks. 12 (92.31% of the 13 bacilli (Mycobacterium bovis showed visible growth using the above growth conditions. In conclusion, microscopic examination can only detect acid fast organisms in the clinical samples whereas culture on L-J medium can differentiate between acid-fast mycobacterial species.

S.H. Manal

2005-01-01

197

Abnormal Nasal Nitric Oxide Production, Ciliary Beat Frequency, and Toll-like Receptor Response in Pulmonary Nontuberculous Mycobacterial Disease Epithelium  

Science.gov (United States)

Rationale: Pulmonary nontuberculous mycobacterial (PNTM) disease has increased over the past several decades, especially in older women. Despite extensive investigation, no consistent immunological abnormalities have been found. Using evidence from diseases such as cystic fibrosis and primary ciliary dyskinesia, in which mucociliary dysfunction predisposes subjects to high rates of nontuberculous mycobacterial disease that increase with age, we investigated correlates of mucociliary function in subjects with PNTM infections and healthy control subjects. Objectives: To define ex vivo characteristics of PNTM disease. Methods: From 2009 to 2012, 58 subjects with PNTM infections and 40 control subjects were recruited. Nasal nitric oxide (nNO) was determined at the time of respiratory epithelial collection. Ciliary beat frequency at rest and in response to Toll-like receptor (TLR) and other agonists was determined using high-speed video microscopy. Measurements and Main Results: We found decreased nNO production, abnormally low resting ciliary beat frequency, and abnormal responses to agonists of TLR2, -3, -5, -7/8, and -9 in subjects with PNTM compared with healthy control subjects. The low ciliary beat frequency in subjects with PNTM was normalized ex vivo by augmentation of the NO–cyclic guanosine monophosphate pathway without normalization of their TLR agonist responses. Conclusions: Impaired nNO, ciliary beat frequency, and TLR responses in PNTM disease epithelium identify possible underlying susceptibility mechanisms as well as possible avenues for directed investigation and therapy. PMID:23593951

Fowler, Cedar J.; Olivier, Kenneth N.; Leung, Janice M.; Smith, Caroline C.; Huth, Andrea G.; Root, Heather; Kuhns, Douglas B.; Logun, Carolea; Zelazny, Adrian; Frein, Cathleen A.; Daub, Janine; Haney, Carissa; Shelhamer, James H.; Bryant, Clare E.

2013-01-01

198

An outbreak of Mycobacterium genavense infection in a flock of captive diamond doves (Geopelia cuneata).  

Science.gov (United States)

Two diamond doves (Geopelia cuneata) in a flock of 23 birds housed in an aviary in a zoo in central Japan were found dead as a result of mycobacteriosis. Fecal samples of the remaining doves were positive for mycobacterial infection, and thus they were euthanatized. Clinical signs and gross pathology, including weight loss and sudden death and slight enlargement of the liver and intestine, were observed in a small number of birds (3/23). Disseminated histiocytic infiltration of either aggregates or sheets of epithelioid cells containing acid-fast bacilli, in the absence of caseous necrosis, were observed in different organs of the infected doves, especially lungs (23/23), intestines (9/23), livers (7/23), and hearts (6/23). Mycobacterium sp. was isolated from the livers of three birds (3/23). DNA extracted from frozen liver and formalin-fixed, paraffin-embedded tissues (5/23) were used for amplification of the gene encoding mycobacterial 65-kDa heat shock protein (hsp65). The causative Mycobacterium species was identified by PCR-restriction fragment length polymorphism analysis. Mycobacterium genavense infection was confirmed in three of the diamond doves. Moreover, partial 16S rDNA gene sequencing revealed 100% identity across the three samples tested, and 99.77% nucleotide homology of the isolate sequence to M. genavense. The main route of M. genavense infection in the diamond doves was most likely airborne, suggesting a potential zoonotic risk of airborne transmission between humans and birds. PMID:25518432

Haridy, Mohie; Fukuta, Mayumi; Mori, Yasuyuki; Ito, Hideyuki; Kubo, Masahito; Sakai, Hiroki; Yanai, Tokuma

2014-09-01

199

Predicting the subcellular localization of mycobacterial proteins by incorporating the optimal tripeptides into the general form of pseudo amino acid composition.  

Science.gov (United States)

Mycobacterium tuberculosis is a bacterium that causes tuberculosis, one of the most prevalent infectious diseases. Predicting the subcellular localization of mycobacterial proteins in this bacterium may provide vital clues for the prediction of protein function as well as for drug discovery and design. Therefore, a computational method that can predict the subcellular localization of mycobacterial proteins with high precision is highly desirable. We propose a computational method to predict the subcellular localization of mycobacterial proteins. An objective and strict benchmark dataset was constructed after collecting 272 non-redundant proteins from the universal protein resource (the UniProt database). Subsequently, a novel feature selection strategy based on binomial distribution was used to optimize the feature vector. Finally, a subset containing 219 chosen tripeptide features was imported into a support vector machine-based method to estimate the performance of the dataset in accurately and sensitively identifying these proteins. We found that the proposed method gave a maximum overall accuracy of 89.71% with an average accuracy of 81.12% in the jackknife cross-validation. The results indicate that our prediction method gave an efficient and powerful performance when compared with other published methods. We made the proposed method available on a purpose built Web server called MycoSub that is freely accessible at . We anticipate that MycoSub will become a useful tool for studying the functions of mycobacterial proteins and for designing and developing anti-mycobacterium drugs. PMID:25437899

Zhu, Pan-Pan; Li, Wen-Chao; Zhong, Zhe-Jin; Deng, En-Ze; Ding, Hui; Chen, Wei; Lin, Hao

2015-02-20

200

Combined Multilocus Short-Sequence-Repeat and Mycobacterial Interspersed Repetitive Unit-Variable-Number Tandem-Repeat Typing of Mycobacterium avium subsp. paratuberculosis Isolates? ‡  

OpenAIRE

Short-sequence-repeat (SSR) sequencing was applied to 127 Mycobacterium avium subsp. paratuberculosis isolates typed by mycobacterial interspersed repetitive unit-variable-number tandem repeats (MIRU-VNTR) and IS900 restriction fragment length polymorphism (RFLP). Combined MIRU-VNTR and SSR typing followed by secondary IS900 RFLP typing is an improved approach to high-resolution genotyping of this pathogen.

Thibault, Virginie C.; Grayon, Maggy; Boschiroli, Maria Laura; Willery, Eve; Allix-be?guec, Caroline; Stevenson, Karen; Biet, Franck; Supply, Philip

2008-01-01

201

Structural and functional characterization of an arylamine N-acetyltransferase from the pathogen Mycobacterium abscessus : differences from other mycobacterial isoforms and implications for selective inhibition  

DEFF Research Database (Denmark)

Mycobacterium abscessus is the most pathogenic rapid-growing mycobacterium and is one of the most resistant organisms to chemotherapeutic agents. However, structural and functional studies of M. abscessus proteins that could modify/inactivate antibiotics remain nonexistent. Here, the structural and functional characterization of an arylamine N-acetyltransferase (NAT) from M. abscessus [(MYCAB)NAT1] are reported. This novel prokaryotic NAT displays significant N-acetyltransferase activity towards aromatic substrates, including antibiotics such as isoniazid and p-aminosalicylate. The enzyme is endogenously expressed and functional in both the rough and smooth M. abscessus morphotypes. The crystal structure of (MYCAB)NAT1 at 1.8?Å resolution reveals that it is more closely related to Nocardia farcinica NAT than to mycobacterial isoforms. In particular, structural and physicochemical differences from other mycobacterial NATs were found in the active site. Peculiarities of (MYCAB)NAT1 were further supported by kinetic and docking studies showing that the enzyme was poorly inhibited by the piperidinol inhibitor of mycobacterial NATs. This study describes the first structure of an antibiotic-modifying enzyme from M. abscessus and provides bases to better understand the substrate/inhibitor-binding specificities among mycobacterial NATs and to identify/optimize specific inhibitors. These data should also contribute to the understanding of the mechanisms that are responsible for the pathogenicity and extensive chemotherapeutic resistance of M. abscessus.

Cocaign, Angélique; Kubiak, Xavier Jean Philippe

2014-01-01

202

Rapid construction of mycobacterial mutagenesis vectors using ligation-independent cloning  

OpenAIRE

Targeted mutagenesis is one of the major tools for determining the function of a given gene and its involvement in bacterial pathogenesis. In mycobacteria, gene deletion is often accomplished by using allelic exchange techniques that commonly utilise a suicide delivery vector. We have adapted a widely-used suicide delivery vector (p1NIL) for cloning two flanking regions of a gene using ligation independent cloning (LIC). The pNILRB plasmid series produced allow a faster, more efficient and le...

Balhana, Ricardo; Stoker, Neil G.; Sikder, Mahmudul Hasan; Chauviac, Francois-xavier; Kendall, Sharon L.

2010-01-01

203

The gene encoding mycobacterial DNA-binding protein I (MDPI) transformed rapidly growing bacteria to slowly growing bacteria.  

Science.gov (United States)

Pathogenic species of Mycobacterium are slowly growing intracellular bacteria. Slow growth is important for the parasitism of these organisms and chronicity of the disease, but its precise mechanism has not been elucidated. Recently, we found that a novel DNA-binding protein (MDPI) was expressed (7-10% in total protein) in mycobacteria, such as Mycobacterium bovis bacillus Calmette-Guérin, Mycobacterium tuberculosis, and Mycobacterium leprae. In this study, we observed that MDPI interfered with replication, transcription, and translation in the analysis in in vitro E. coli cell-free macromolecular biosynthesizing systems. Furthermore, MDPI inhibited the rapid growth of both Escherichia coli and Mycobacterium smegmatis, and NH(2)-terminal second amino acid, asparagine, was observed to be important in terms of this function. These data suggest an important role of MDPI for suppression of growth rates of mycobacteria. PMID:10620682

Matsumoto, S; Furugen, M; Yukitake, H; Yamada, T

2000-01-15

204

Association of IFNGR2 gene polymorphisms with pulmonary tuberculosis among the Vietnamese  

OpenAIRE

Interferon-? (IFN-?) is a key molecule of T helper 1 (Th1)-immune response against tuberculosis (TB), and rare genetic defects of IFN-? receptors cause disseminated mycobacterial infection. The aim of the present study was to investigate whether genetic polymorphisms found in the Th1-immune response genes play a role in TB. In our study, DNA samples were collected from two series of cases including 832 patients with new smear-positive TB and 506 unrelated individuals with no history of TB ...

Hijikata, Minako; Shojima, Junko; Matsushita, Ikumi; Tokunaga, Katsushi; Ohashi, Jun; Hang, Nguyen T. L.; Horie, Toru; Sakurada, Shinsaku; Hoang, Nguyen P.; Thuong, Pham H.; Lien, Luu T.; Keicho, Naoto

2012-01-01

205

Transcriptional profiling of mycobacterial antigen-induced responses in infants vaccinated with BCG at birth  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background Novel tuberculosis (TB vaccines recently tested in humans have been designed to boost immunity induced by the current vaccine, Mycobacterium bovis Bacille Calmette-Guérin (BCG. Because BCG vaccination is used extensively in infants, this population group is likely to be the first in which efficacy trials of new vaccines will be conducted. However, our understanding of the complexity of immunity to BCG in infants is inadequate, making interpretation of vaccine-induced immune responses difficult. Methods To better understand BCG-induced immunity, we performed gene expression profiling in five 10-week old infants routinely vaccinated with BCG at birth. RNA was extracted from 12 hour BCG-stimulated or purified protein derivative of tuberculin (PPD-stimulated PBMC, isolated from neonatal blood collected 10 weeks after vaccination. RNA was hybridised to the Sentrix® HumanRef-8 Expression BeadChip (Illumina to measure expression of >16,000 genes. Results We found that ex vivo stimulation of PBMC with PPD and BCG induced largely similar gene expression profiles, except that BCG induced greater macrophage activation. The peroxisome proliferator-activated receptor (PPAR signaling pathway, including PPAR-?, involved in activation of the alternative, anti-inflammatory macrophage response was down-regulated following stimulation with both antigens. In contrast, up-regulation of genes associated with the classic, pro-inflammatory macrophage response was noted. Further analysis revealed a decrease in the expression of cell adhesion molecules (CAMs, including integrin alpha M (ITGAM, which is known to be important for entry of mycobacteria into the macrophage. Interestingly, more leukocyte genes were down-regulated than up-regulated. Conclusion Our results suggest that a combination of suppressed and up-regulated genes may be key in determining development of protective immunity to TB induced by vaccination with BCG.

Hill Adrian VS

2009-02-01

206

AADNMR: A Simple Method for Rapid Identification of Bacterial/Mycobacterial Infections in Antibiotic Treated Peritoneal Dialysis Effluent Samples for Diagnosis of Infectious Peritonitis  

CERN Document Server

An efficient method is reported for rapid identification of bacterial or mycobacterial infection in a suspected clinical/biological sample. The method is based on the fact that the ring methylene protons of cyclic fatty acids (constituting the cell membrane of several species of bacteria and mycobacteria) resonate specifically between -0.40 and 0.68 ppm region of the 1H NMR spectrum. These cyclic fatty acids are rarely found in the eukaryotic cell membranes. Therefore, the signals from cyclic ring moiety of these fatty acids can be used as markers (a) for the identification of bacterial and mycobacterial infections and (b) for differential diagnosis of bacterial and fungal infections. However, these microbial fatty acids when present inside the membrane are not easily detectable by NMR owing to their fast T2 relaxation. Nonetheless, the problem can easily be circumvented if these fatty acids become suspended in solution. This has been achieved by abolishing the membrane integrity using broad spectrum antibiot...

Guleria, Anupam; Rawat, Atul; Khetrapal, C L; Prasad, Narayan; Kumar, Dinesh

2014-01-01

207

Thin-Section CT Findings of Nontuberculous Mycobacterial Pulmonary Diseases: Comparison Between Mycobacterium avium-intracellulare Complex and Mycobacterium abscessus Infection  

OpenAIRE

We aimed to compare the CT findings of nontuberculous mycobacterial pulmonary diseases caused by Mycobacterium avium-intracellulare complex (MAC) and Mycobacterium abscessus. Two chest radiologists analyzed retrospectively the thin-section CT findings of 51 patients with MAC and 36 with M. abscessus infection in terms of patterns and forms of lung lesions. No significant difference was found between MAC and M. abscessus infection in the presence of small nodules, tree-in-bud pattern, and bron...

Chung, Myung Jin; Lee, Kyung Soo; Koh, Won-jung; Lee, Ju Hyun; Kim, Tae Sung; Kwon, O. Jung; Kim, Seonwoo

2005-01-01

208

T Cell Reactivity against Mycolyl Transferase Antigen 85 of M. tuberculosis in HIV-TB Coinfected Subjects and in AIDS Patients Suffering from Tuberculosis and Nontuberculous Mycobacterial Infections  

OpenAIRE

The mycolyl transferase antigen 85 complex is a major secreted protein family from mycobacterial culture filtrate, demonstrating powerful T cell stimulatory properties in most HIV-negative, tuberculin-positive volunteers with latent M.tuberculosis infection and only weak responses in HIV-negative tuberculosis patients. Here, we have analyzed T cell reactivity against PPD and Ag85 in HIV-infected individuals, without or with clinical symptoms of tuberculosis, and in AIDS patients with disease ...

Kris Huygen; Jean-Paul Van Vooren; Claire-Michèle Farber; Pierre Couppie; Eliane Bourreau; Annie Drowart; Pascal Launois

2011-01-01

209

Combined Multilocus Short-Sequence-Repeat and Mycobacterial Interspersed Repetitive Unit-Variable-Number Tandem-Repeat Typing of Mycobacterium avium subsp. paratuberculosis Isolates? ‡  

Science.gov (United States)

Short-sequence-repeat (SSR) sequencing was applied to 127 Mycobacterium avium subsp. paratuberculosis isolates typed by mycobacterial interspersed repetitive unit-variable-number tandem repeats (MIRU-VNTR) and IS900 restriction fragment length polymorphism (RFLP). Combined MIRU-VNTR and SSR typing followed by secondary IS900 RFLP typing is an improved approach to high-resolution genotyping of this pathogen. PMID:18923016

Thibault, Virginie C.; Grayon, Maggy; Boschiroli, Maria Laura; Willery, Eve; Allix-Béguec, Caroline; Stevenson, Karen; Biet, Franck; Supply, Philip

2008-01-01

210

Combined multilocus short-sequence-repeat and mycobacterial interspersed repetitive unit-variable-number tandem-repeat typing of Mycobacterium avium subsp. paratuberculosis isolates.  

Science.gov (United States)

Short-sequence-repeat (SSR) sequencing was applied to 127 Mycobacterium avium subsp. paratuberculosis isolates typed by mycobacterial interspersed repetitive unit-variable-number tandem repeats (MIRU-VNTR) and IS900 restriction fragment length polymorphism (RFLP). Combined MIRU-VNTR and SSR typing followed by secondary IS900 RFLP typing is an improved approach to high-resolution genotyping of this pathogen. PMID:18923016

Thibault, Virginie C; Grayon, Maggy; Boschiroli, Maria Laura; Willery, Eve; Allix-Béguec, Caroline; Stevenson, Karen; Biet, Franck; Supply, Philip

2008-12-01

211

Calcium Signaling in Dendritic Cells by Human or Mycobacterial Hsp70 Is Caused by Contamination and Is Not Required for Hsp70-mediated Enhancement of Cross-presentation*  

OpenAIRE

Extracellular heat shock proteins (HSPs) can stimulate antigen-specific immune responses. Using recombinant human (rhu)Hsp70, we previously demonstrated that through complex formation with exogenous antigenic peptides, rhuHsp70 can enhance cross-presentation by antigen-presenting cells (APCs) resulting in stronger T cell stimulation. T cell stimulatory activity has also been described for mycobacterial (myc)Hsp70. MycHsp70-assisted T cell activation has been reported t...

Bendz, Henriette; Marincek, Boris-christian; Momburg, Frank; Ellwart, Joachim W.; Issels, Rolf D.; Nelson, Peter J.; Noessner, Elfriede

2008-01-01

212

Mycobacterial Interspersed Repetitive Unit Typing of Mycobacterium tuberculosis Compared to IS6110-Based Restriction Fragment Length Polymorphism Analysis for Investigation of Apparently Clustered Cases of Tuberculosis  

OpenAIRE

An evaluation of the utility of IS6110-based restriction fragment length polymorphism (RFLP) typing compared to a combination of variable number tandem repeat (VNTR) typing and mycobacterial interspersed repetitive unit (MIRU) typing was undertaken. A total of 53 patient isolates of Mycobacterium tuberculosis from four presumed episodes of cross-infection were examined. Genomic DNA was extracted from the isolates by a cetyl trimethylammonium bromide method. The number of copies of tandem repe...

Hawkey, Peter M.; Smith, E. Grace; Evans, Jason T.; Monk, Philip; Bryan, Gerry; Mohamed, Huda H.; Bardhan, Madhu; Pugh, R. Nicholas

2003-01-01

213

Last Step in the Conversion of Trehalose to Glycogen: A MYCOBACTERIAL ENZYME THAT TRANSFERS MALTOSE FROM MALTOSE 1-PHOSPHATE TO GLYCOGEN  

OpenAIRE

We show that Mycobacterium smegmatis has an enzyme catalyzing transfer of maltose from [14C]maltose 1-phosphate to glycogen. This enzyme was purified 90-fold from crude extracts and characterized. Maltose transfer required addition of an acceptor. Liver, oyster, or mycobacterial glycogens were the best acceptors, whereas amylopectin had good activity, but amylose was a poor acceptor. Maltosaccharides inhibited the transfer of maltose from [14C]maltose-1-P to glycogen because they were also ac...

Elbein, Alan D.; Pastuszak, Irena; Tackett, Alan J.; Wilson, Tyler; Pan, Yuan T.

2010-01-01

214

In Vitro Gene Expression Dissected: Chemostat Surgery for Mycobacterium Tuberculosis  

Directory of Open Access Journals (Sweden)

Full Text Available A unique approach, combining defined and reproducible in vitro models with DNA microarrays, has been developed to study environmental modulation of mycobacterial gene expression. The gene expression profiles of samples of Mycobacterium tuberculosis, from independent chemostat cultures grown under defined and reproducible conditions, were found to be highly correlated. This approach is now being used to study the effect of relevant stimuli, such as limited oxygen availability, on mycobacterial gene expression. A modification of the chemostat culture system, enabling largevolume controlled batch culture, has been developed to study starvation survival. Cultures of M. tuberculosis have been maintained under nutrient-starved conditions for extended periods, with 106 – 107 bacilli surviving in a culturable state after 100 days. The design of the culture system has made it possible to control the environment and collect multiple time-course samples to study patterns of gene expression. These studies demonstrate that it is possible to perform long-term studies and obtain reproducible expression data using controlled and defined in vitro models.

Philip D. Marsh

2002-01-01

215

atpE gene as a new useful specific molecular target to quantify Mycobacterium in environmental samples  

OpenAIRE

BackgroundThe environment is the likely source of many pathogenic mycobacterial species but detection of mycobacteria by bacteriological tools is generally difficult and time-consuming. Consequently, several molecular targets based on the sequences of housekeeping genes, non-functional RNA and structural ribosomal RNAs have been proposed for the detection and identification of mycobacteria in clinical or environmental samples. While certain of these targets were proposed as specific for...

Radomski, Nicolas; Roguet, Ade?lai?de; Lucas, Franc?oise; Veyrier, Fre?de?ric; Cambau, Emmanuelle; Accrombessi, He?berte; Moilleron, Re?gis; Behr, Marcel; Moulin, Laurent

2013-01-01

216

rpoB Gene Mutations and Molecular Characterization of Rifampin-Resistant Mycobacterium tuberculosis Isolates from Shandong Province, China  

OpenAIRE

Sixty rifampin (RIF)-resistant and 75 RIF-susceptible Mycobacterium tuberculosis isolates from Shandong Province, China, were analyzed for rpoB gene mutations and genotyped. Mycobacterial interspersed repetitive unit (MIRU) genotype 223325173533 was overrepresented among RIF-resistant isolates. MIRU combined with IS6110 restriction fragment length polymorphism analysis as the second-line genotyping method may reflect epidemiologic links more reliably than each method alone.

Ma, Xin; Wang, Haiying; Deng, Yunfeng; Liu, Zhimin; Xu, Yong; Pan, Xi; Musser, James M.; Graviss, Edward A.

2006-01-01

217

Assembly and proteolytic processing of mycobacterial ClpP1 and ClpP2.  

OpenAIRE

Abstract Background Caseinolytic proteases (ClpPs) are barrel-shaped self-compartmentalized peptidases involved in eliminating damaged or short-lived regulatory proteins. The Mycobacterium tuberculosis (MTB) genome contains two genes coding for putative ClpPs, ClpP1 and ClpP2 respectively, that are likely to play a role in the virulence of the bacterium. Results We report the first biochemical characterization of ClpP1 and ClpP2 peptidases from MTB. Bot...

Benaroudj Nadia; Raynal Bertrand; Miot Marika; Ortiz-Lombardia Miguel

2011-01-01

218

Effective inhibitors of the essential kinase PknB and their potential as anti-mycobacterial agents.  

Science.gov (United States)

PknB is an essential serine/threonine kinase of Mycobacterium tuberculosis with possible roles in a number of signalling pathways involved in cell division and metabolism. We screened a library of >50,000 compounds for inhibitors of the in vitro phosphorylation of GarA (Rv1827) by PknB and identified a number of inhibitors. A program of synthetic medicinal chemistry was subsequently conducted around one class of inhibitors and was successful in generating ATP competitive inhibitors with potency in the nanomolar range. Compounds in this class showed cross-reactivity with the related M. tuberculosis kinase, PknF, but not with PknG in an in vitro autophosphorylation assay. These synthesised inhibitors were able to prevent the growth of M. tuberculosis in an Alamar blue assay and in an intracellular model of infection, but only in the micromolar range. We attempted to determine if cell wall permeability was an explanation for the discrepancy between the potent in vitro compared with relatively poor in vivo activity, but found no evidence that the activity of the inhibitors could be improved by weakening the cell wall. Despite a number of drug discovery efforts attempting to develop inhibitors against PknB, it is yet to be reported that any such inhibitors prevent mycobacterial growth at submicromolar concentrations. PMID:21482481

Lougheed, Kathryn E A; Osborne, Simon A; Saxty, Barbara; Whalley, David; Chapman, Tim; Bouloc, Nathalie; Chugh, Jasveen; Nott, Timothy J; Patel, Dony; Spivey, Vicky L; Kettleborough, Catherine A; Bryans, Justin S; Taylor, Debra L; Smerdon, Stephen J; Buxton, Roger S

2011-07-01

219

Mycobacterial and nonbacterial pulmonary complications in hospitalized patients with human immunodeficiency virus infection: A prospective, cohort study  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background A prospective observational study was done to describe nonbacterial pulmonary complications in hospitalized patients with human immunodeficiency virus (HIV infection. Methods The study included 1,225 consecutive hospital admissions of 599 HIV-infected patients treated from April 1995 through March 1998. Data included demographics, risk factors for HIV infection, Acute Physiology and Chronic Health Evaluation (APACHE II score, pulmonary complications, CD4+ lymphocyte count, hospital stay and case-fatality rate. Results Patient age (mean ± SD was 38.2 ± 8.9 years, 62% were men, and 84% were African American. The median APACHE II score was 14, and median CD4+ lymphocyte count was 60/?L. Pulmonary complications were Pneumocystis carinii pneumonia (85 in 78 patients, Mycobacterium avium complex (51 in 38, Mycobacterium tuberculosis (40 in 35, Mycobacterium gordonae (11 in 11, Mycobacterium kansasii (10 in 9, Cytomegalovirus (10 in 10, Nocardia asteroides (3 in 3, fungus ball (2 in 2, respiratory syncytial virus (1, herpes simplex virus (1, Histoplasma capsulatum (1, lymphoma (3 in 3, bronchogenic carcinoma (2 in 2, and Kaposi sarcoma (1. The case-fatality rate of patients was 11% with Pneumocystis carinii pneumonia; 5%, Mycobacterium tuberculosis; 6%, Mycobacterium avium complex; and 7%, noninfectious pulmonary complications. Conclusion Most pulmonary complications in hospitalized patients with HIV are from Pneumocystis and mycobacterial infection.

Afessa Bekele

2001-09-01

220

Mycobacterial species diversity at a general hospital on the island of Crete: first detection of Mycobacterium lentiflavum in Greece.  

Science.gov (United States)

The objective of the present study was to investigate the diversity of mycobacterial isolates in a general hospital in Crete, Greece. 48 positive Lowenstein-Jensen cultures over a 3-y period were analysed by means of AccuProbe and GenoType assays. Non-tuberculous mycobacteria (NTM) comprised the majority of the isolates (56.3%, 27/48) vs 33.3% (16/48) of M. tuberculosis; 10.4% of the isolates could not be classified. Among NTM, M. lentiflavum was the predominant species isolated (9/27) followed by M. kansasii, M. gordonae and M. peregrinum, whereas no M. avium complex isolates were detected. This is the first detection of M. lentiflavum in Greece. The susceptibilities of the M. lentiflavum isolates to an extended panel of antibiotics were determined by the proportions method and the medical files of the 9 patients were reviewed. Three isolates were from urine, which is an unusual site. All strains exhibited multidrug resistance. The patients were adults with immunosuppression or predisposing conditions for NTM infection. Diagnosis of true infection was either not pursued or the patients died shortly after isolation. PMID:17852893

Neonakis, Ioannis K; Gitti, Zoe; Kourbeti, Irene S; Michelaki, Helen; Baritaki, Maria; Alevraki, Georgia; Papadomanolaki, Evangelia; Tsafaraki, Ekaterini; Tsouri, Anna; Baritaki, Stavroula; Krambovitis, Elias; Spandidos, Demetrios A

2007-01-01

221

Systemic BCG immunization induces persistent lung mucosal multifunctional CD4 T(EM) cells which expand following virulent mycobacterial challenge.  

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To more closely understand the mechanisms of how BCG vaccination confers immunity would help to rationally design improved tuberculosis vaccines that are urgently required. Given the established central role of CD4 T cells in BCG induced immunity, we sought to characterise the generation of memory CD4 T cell responses to BCG vaccination and M. bovis infection in a murine challenge model. We demonstrate that a single systemic BCG vaccination induces distinct systemic and mucosal populations of T effector memory (T(EM)) cells in vaccinated mice. These CD4+CD44(hi)CD62L(lo)CD27? T cells concomitantly produce IFN-? and TNF-?, or IFN-?, IL-2 and TNF-? and have a higher cytokine median fluorescence intensity MFI or 'quality of response' than single cytokine producing cells. These cells are maintained for long periods (>16 months) in BCG protected mice, maintaining a vaccine-specific functionality. Following virulent mycobacterial challenge, these cells underwent significant expansion in the lungs and are, therefore, strongly associated with protection against M. bovis challenge. Our data demonstrate that a persistent mucosal population of T(EM) cells can be induced by parenteral immunization, a feature only previously associated with mucosal immunization routes; and that these multifunctional T(EM) cells are strongly associated with protection. We propose that these cells mediate protective immunity, and that vaccines designed to increase the number of relevant antigen-specific T(EM) in the lung may represent a new generation of TB vaccines. PMID:21720558

Kaveh, Daryan A; Bachy, Véronique S; Hewinson, R Glyn; Hogarth, Philip J

2011-01-01

222

Systemic BCG Immunization Induces Persistent Lung Mucosal Multifunctional CD4 TEM Cells which Expand Following Virulent Mycobacterial Challenge  

Science.gov (United States)

To more closely understand the mechanisms of how BCG vaccination confers immunity would help to rationally design improved tuberculosis vaccines that are urgently required. Given the established central role of CD4 T cells in BCG induced immunity, we sought to characterise the generation of memory CD4 T cell responses to BCG vaccination and M. bovis infection in a murine challenge model. We demonstrate that a single systemic BCG vaccination induces distinct systemic and mucosal populations of T effector memory (TEM) cells in vaccinated mice. These CD4+CD44hiCD62LloCD27? T cells concomitantly produce IFN-? and TNF-?, or IFN-?, IL-2 and TNF-? and have a higher cytokine median fluorescence intensity MFI or ‘quality of response’ than single cytokine producing cells. These cells are maintained for long periods (>16 months) in BCG protected mice, maintaining a vaccine–specific functionality. Following virulent mycobacterial challenge, these cells underwent significant expansion in the lungs and are, therefore, strongly associated with protection against M. bovis challenge. Our data demonstrate that a persistent mucosal population of TEM cells can be induced by parenteral immunization, a feature only previously associated with mucosal immunization routes; and that these multifunctional TEM cells are strongly associated with protection. We propose that these cells mediate protective immunity, and that vaccines designed to increase the number of relevant antigen-specific TEM in the lung may represent a new generation of TB vaccines. PMID:21720558

Kaveh, Daryan A.; Bachy, Véronique S.; Hewinson, R. Glyn; Hogarth, Philip J.

2011-01-01

223

Interleukin-1? triggers the differentiation of macrophages with enhanced capacity to present mycobacterial antigen to T cells.  

Science.gov (United States)

The rapid differentiation of monocytes into macrophages (M?) and dendritic cells is a pivotal aspect of the innate immune response. Differentiation is triggered following recognition of microbial ligands that activate pattern recognition receptors or directly by pro-inflammatory cytokines. We demonstrate that interleukin-1? (IL-1?) induces the rapid differentiation of monocytes into CD209(+) M?, similar to activation via Toll-like receptor 2/1, but with distinct phenotypic and functional characteristics. The IL-1? induced M? express higher levels of key markers of phagocytosis, including the Fc-receptors CD16 and CD64, as well as CD36, CD163 and CD206. In addition, IL-1?-induced M? exert potent phagocytic activity towards inert particles, oxidized low-density lipoprotein and mycobacteria. Furthermore, IL-1?-induced M? express higher levels of HLA-DR and effectively present mycobacterial antigens to T cells. Therefore, the ability of IL-1? to induce monocyte differentiation into M? with both phagocytosis and antigen-presenting function is a distinct part of the innate immune response in host defence against microbial infection. PMID:24032597

Schenk, Mirjam; Fabri, Mario; Krutzik, Stephan R; Lee, Delphine J; Vu, David M; Sieling, Peter A; Montoya, Dennis; Liu, Philip T; Modlin, Robert L

2014-02-01

224

Infection of an equine placenta with a novel mycobacterial species leading to abortion.  

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A 25-year-old pregnant American Quarter Horse mare presented with a 1-week history of progressively worsening vaginal discharge. Transrectal ultrasound revealed increased thickness of the combined uterus and placenta with evidence of chorioallantoic edema but no placental separation. A thickened amnion was visible on transabdominal ultrasound. Abortion occurred 2 days after presentation despite medical treatment. At necropsy, the chorioallantois had variable but diffuse thickening with focally extensive browning of the chorionic surface in the right horn and adjacent body. There were fluid-filled sacculations on the allantoic surface of the umbilical cord, allantoamnion, and chorioallantois associated with diffuse perivascular fluid microscopically. A nonbranching acid-fast bacterium identified as belonging to the genus Mycobacterium Runyon group IV was isolated from the chorioallantois and uterine fluid. Ziehl-Neelsen stain confirmed the presence of intracellular acid-fast bacilli in trophoblasts of the gravid horn and the cervical star area. The current case is unique in that the mycobacteria did not initiate a significant granulomatous inflammatory response in the chorion unless villar necrosis occurred. Sequence analysis of the 16S ribosomal RNA gene and the rpo? gene, encoding the ? subunit of RNA polymerase, indicated that the strain of mycobacteria isolated in this case belonged to a novel species of rapidly growing mycobacteria and not to an established species. Mycobacteria are an uncommon and sporadic cause of placentitis and abortion, but should be suspected in cases of chronic placentitis that are not restricted to the cervical star area. PMID:22585955

Johnson, Aime K; Roberts, John F; Hagan, Alexander; Wilborn, Robyn R; Dujovne, Ghislaine; Sells, Stephen F; Donahue, J Michael

2012-07-01

225

The role of transcriptional regulation in maintaining the availability of mycobacterial adenylate cyclases  

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Full Text Available Mycobacterium species have a complex cAMP regulatory network indicated by the high number of adenylate cyclases annotated in their genomes. However the need for a high level of redundancy in adenylate cyclase genes remains unknown. We have used semiquantitiative RT-PCR to examine the expression of eight Mycobacterium smegmatis cyclases with orthologs in the human pathogen Mycobacterium tuberculosis, where cAMP has recently been shown to be important for virulence. All eight cyclases were transcribed in all environments tested, and only four demonstrated environmental-mediated changes in transcription. M. smegmatis genes MSMEG_0545 and MSMEG_4279 were upregulated during starvation conditions while MSMEG_0545 and MSMEG_4924 were downregulated in H2O2 and MSMEG_3780 was downregulated in low pH and starvation. Promoter fusion constructs containing M. tuberculosis H37Rv promoters showed consistent regulation compared to their M. smegmatis orthologs. Overall our findings indicate that while low levels of transcriptional regulation occur, regulation at the mRNA level does not play a major role in controlling cellular cyclase availability in a given environment.

Sarah J. Casey

2014-03-01

226

Interruption of the phosphoglucose isomerase gene results in glucose auxotrophy in Mycobacterium smegmatis.  

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Two glycerol utilization mutants of Mycobacterium smegmatis that were unable to utilize most carbon sources except glucose were isolated. Supplementation of these media with small amounts of glucose restored growth in the mutants; these strains are therefore glucose auxotrophs. The mutant phenotype is complemented by the gene encoding phosphoglucose isomerase (pgi), and direct measurement of enzyme activities in the mutants suggests that this gene product is absent in the auxotrophic strains. Mapping of the mutant allele by Southern analysis demonstrates the presence of a 1-kb deletion extending into the coding sequence of pgi. The possible roles of phosphoglucose isomerase in mycobacterial cell wall synthesis and metabolic regulation are discussed. PMID:9098072

Tuckman, D; Donnelly, R J; Zhao, F X; Jacobs, W R; Connell, N D

1997-04-01

227

Cloning of the Gene Encoding a 22-Kilodalton Cell Surface Antigen of Mycobacterium bovis BCG and Analysis of Its Potential for DNA Vaccination against Tuberculosis  

OpenAIRE

Using spleen cells from mice vaccinated with live Mycobacterium bovis BCG, we previously generated three monoclonal antibodies reactive against a 22-kDa protein present in mycobacterial culture filtrate (CF) (K. Huygen et al., Infect. Immun. 61:2687–2693, 1993). These monoclonal antibodies were used to screen an M. bovis BCG genomic library made in phage ?gt11. The gene encoding a 233-amino-acid (aa) protein, including a putative 26-aa signal sequence, was isolated, and sequence analysis i...

Lefe?vre, Philippe; Denis, Olivier; Wit, Lucas; Tanghe, Audrey; Vandenbussche, Paul; Content, Jean; Huygen, Kris

2000-01-01

228

High Throughput Phenotypic Selection of Mycobacterium tuberculosis Mutants with Impaired Resistance to Reactive Oxygen Species Identifies Genes Important for Intracellular Growth  

OpenAIRE

Mycobacterium tuberculosis has the remarkable capacity to survive within the hostile environment of the macrophage, and to resist potent antibacterial molecules such as reactive oxygen species (ROS). Thus, understanding mycobacterial resistance mechanisms against ROS may contribute to the development of new anti-tuberculosis therapies. Here we identified genes involved in such mechanisms by screening a high-density transposon mutant library, and we show that several of them are involved in th...

Mestre, Olga; Hurtado-ortiz, Raquel; Dos Vultos, Tiago; Namouchi, Amine; Cimino, Mena; Pimentel, Madalena; Neyrolles, Olivier; Gicquel, Brigitte

2013-01-01

229

Perspective on sequence evolution of microsatellite locus (CCG)n in Rv0050 gene from Mycobacterium tuberculosis  

OpenAIRE

Abstract Background The mycobacterial genome is inclined to polymerase slippage and a high mutation rate in microsatellite regions due to high GC content and absence of a mismatch repair system. However, the exact molecular mechanisms underlying microsatellite variation have not been fully elucidated. Here, we investigated mutation events in the hyper-variable trinucleotide microsatellite locus MML0050 located in the Rv0050 gene of W-Beijing and non-W-Beijing Mycobacteriu...

Jin Ruiliang; Cui Zhenling; Liu Zhonghua; Yang Hua; Lu Junmei; Zheng Ruijuan; Wang Jie; Qin Lianhua; Feng Yonghong; Hu Zhongyi

2011-01-01

230

Novel Diagnostic Algorithm for Identification of Mycobacteria Using Genus-Specific Amplification of the 16S-23S rRNA Gene Spacer and Restriction Endonucleases  

OpenAIRE

A novel genus-specific PCR for mycobacteria with simple identification to the species level by restriction fragment length polymorphism (RFLP) was established using the 16S-23S ribosomal RNA gene (rDNA) spacer as a target. Panspecificity of primers was demonstrated on the genus level by testing 811 bacterial strains (122 species in 37 genera from 286 reference strains and 525 clinical isolates). All mycobacterial isolates (678 strains among 48 defined species and 5 indeterminate taxons) were ...

Roth, Andreas; Reischl, Udo; Streubel, Anna; Naumann, Ludmila; Kroppenstedt, Reiner M.; Habicht, Marion; Fischer, Marga; Mauch, Harald

2000-01-01

231

Overexpression of proinflammatory TLR-2-signalling lipoproteins in hypervirulent mycobacterial variants.  

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Changes in the cell envelope composition of mycobacteria cause major changes in cytokine profiles of infected antigen presenting cells. We describe here the modulation of inflammatory responses by Mycobacterium abscessus, an emerging pathogen in cystic fibrosis. M. abscessus is able to switch from a smooth (S) to a rough (R) morphotype by the loss of a surface glycopeptidolipid. R variants are associated with severe clinical forms and a 'hyper-proinflammatory' response in ex vivo and in vivo models. Using partitioning of cell surface components we found that a complex fraction, more abundant in R variants than in S variants, made a major contribution to the TLR-2-dependent hyper-proinflammatory response induced by R variants. Lipoproteins were the main TLR-2 agonists in this fraction, consistent with the larger amounts of 16 lipoproteins in cell surface extracts from R variants; 15 out of 16 being more strongly induced in R variant than in S variant. Genetic interruption of glycopeptidolipid pathway in wild-type S variant resulted in R phenotype with similar induction of lipoprotein genes. In conclusion, R morphotype in M. abscessus is associated with increased synthesis/exposure at the cell surface of lipoproteins, these changes profoundly modifying the innate immune response through TLR-2-dependent mechanisms. PMID:21143571

Roux, Anne-Laure; Ray, Aurélie; Pawlik, Alexandre; Medjahed, Halima; Etienne, Gilles; Rottman, Martin; Catherinot, Emilie; Coppée, Jean-Yves; Chaoui, Karima; Monsarrat, Bernard; Toubert, Antoine; Daffé, Mamadou; Puzo, Germain; Gaillard, Jean-Louis; Brosch, Roland; Dulphy, Nicolas; Nigou, Jérôme; Herrmann, Jean-Louis

2011-05-01

232

The DosS-DosT/DosR Mycobacterial Sensor System  

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Full Text Available DosS/DosR is a two-component regulatory system in which DosS, a heme-containing sensor also known as DevS, under certain conditions undergoes autophosphorylation and then transfers the phosphate to DosR, a DNA-binding protein that controls the entry of Mycobacterium tuberculosis and other mycobacteria into a latent, dormant state. DosT, a second sensor closely related to DosS, is present in M. tuberculosis and participates in the control of the dormancy response mediated by DosR. The binding of phosphorylated DosR to DNA initiates the expression of approximately fifty dormancy-linked genes. DosT is accepted to be a gas sensor that is activated in the ferrous state by the absence of an oxygen ligand or by the binding of NO or CO. DosS functions in a similar fashion as a gas sensor, but contradictory evidence has led to the suggestion that it also functions as a redox state sensor. This review focuses on the structure, biophysical properties, and function of the DosS/DosT heme sensors.

Santhosh Sivaramakrishnan

2013-07-01

233

Tomography high Resolution CT findings of nontuberculous mycobacterial pulmonary disease: Comparison between the first treatment and the re treatment group  

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To analyze and compare the thin section CT findings of first and re treatment nontuberculous mycobacterial (NTM) pulmonary disease. Between January 2005 and April 2010, 121 patients with positive sputum culture for NTM were recruited. We included only 32 patients underwent high resolution chest CT and were confirmed by American Thoracic Society criteria NTM pulmonary infection (first treatment 15, re treatment 17 patients). CT images of 32 patients were reviewed retrospectively. We evaluated the frequency and laterality of the followings; nodule, increased density, bronchial change, parenchymal change. The significantly frequent CT findings of the re treatment NTM group were well defined nodules (retreatment 82.4%, first treatment 33.3%, p = 0.00), consolidations (retreatment 88.2%, first treatment 53.3%, p = 0.03), bronchial changes (bronchiectasis; retreatment 100%, first treatment 66.6%, p = 0.01, bronchial narrowing; retreatment 23.5%, first treatment 0%, p = 0.04 and mucoid impaction; retreatment-58.8%, first treatment-20.0%, p = 0.03) and atelectasis with bronchiectasis (retreatment-88.2%, first treatment 26.7%, p = 0.00). However, most of the evaluated thin section CT findings, such as centrilobular and ill defined nodules, lobular, segmental and subpleural consolidations, ground glass attenuation, bronchial wall thickening, cavities, pleural lesions, fibrotic band, emphysema and laterality of lesions, have not shown significant differences between first treatment and the re treatment group. Thin section CT findings of well defined nodules, consolidations, bronchial changes (bronchiectasis, bronchial narrowing and mucoid impaction) and atelectasis with bronchiectasis are highly suggestive of re treatment NTM pulmonary disease.

Gwak, Soon Hyuk; Cho, Bum Sang; Jeon, Min Hee; Kim, Eun Young; Kang, Min Ho; Yi, Kyung Sik; Lee, Seung Young; Kim, Sung Jin; Lee, Ki Man [Chungbuk National Univ., Cheongju, (Korea, Republic of)

2012-06-15

234

Tomography high Resolution CT findings of nontuberculous mycobacterial pulmonary disease: Comparison between the first treatment and the re treatment group  

International Nuclear Information System (INIS)

To analyze and compare the thin section CT findings of first and re treatment nontuberculous mycobacterial (NTM) pulmonary disease. Between January 2005 and April 2010, 121 patients with positive sputum culture for NTM were recruited. We included only 32 patients underwent high resolution chest CT and were confirmed by American Thoracic Society criteria NTM pulmonary infection (first treatment 15, re treatment 17 patients). CT images of 32 patients were reviewed retrospectively. We evaluated the frequency and laterality of the followings; nodule, increased density, bronchial change, parenchymal change. The significantly frequent CT findings of the re treatment NTM group were well defined nodules (retreatment 82.4%, first treatment 33.3%, p = 0.00), consolidations (retreatment 88.2%, first treatment 53.3%, p = 0.03), bronchial changes (bronchiectasis; retreatment 100%, first treatment 66.6%, p = 0.01, bronchial narrowing; retreatment 23.5%, first treatment 0%, p = 0.04 and mucoid impaction; retreatment-58.8%, first treatment-20.0%, p = 0.03) and atelectasis with bronchiectasis (retreatment-88.2%, first treatment 26.7%, p = 0.00). However, most of the evaluated thin section CT findings, such as centrilobular and ill defined nodules, lobular, segmental and subpleural consolidations, ground glass attenuation, bronchial wall thickening, cavities, pleural lesions, fibrotic band, emphysema and laterality of lesions, have not shown significant differences between first treatmgnificant differences between first treatment and the re treatment group. Thin section CT findings of well defined nodules, consolidations, bronchial changes (bronchiectasis, bronchial narrowing and mucoid impaction) and atelectasis with bronchiectasis are highly suggestive of re treatment NTM pulmonary disease

235

A comparison of gross pathology, histopathology, and mycobacterial culture for the diagnosis of tuberculosis in elk (Cervus elaphus).  

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Using the isolation of Mycobacterium bovis as the reference standard, this study evaluated the sensitivity, specificity and kappa statistic of gross pathology (abattoir postmortem inspection), histopathology, and parallel or series combinations of the two for the diagnosis of tuberculosis in 430 elk and red deer. Two histopathology interpretations were evaluated: histopathology I, where the presence of lesions compatible with tuberculosis was considered positive, and histopathology II, where lesions compatible with tuberculosis or a select group of additional possible diagnoses were considered positive. In the 73 animals from which M. bovis was isolated, gross lesions of tuberculosis were most often in the lung (48), the retropharyngeal lymph nodes (36), the mesenteric lymph node (35), and the mediastinal lymph nodes (16). Other mycobacterial isolates included: 11 M. paratuberculosis, 11 M. avium, and 28 rapidly growing species or M. terrae complex. The sensitivity estimates of gross pathology and histopathology I were 93% (95% confidence limits [CL] 84.97%) and 88% [CL 77.94%], respectively, and the specificity of both was 89% [CL 85.92%]). The sensitivity and specificity of histopathology II were 89% (CL 79.95%) and 77% (CL 72.81%), respectively. The highest sensitivity estimates (93-95% [CL 84.98%]) were obtained by interpreting gross pathology and histopathology in parallel (where an animal had to be positive on at least one of the two, to be classified as combination positive). The highest specificity estimates (94-95% [CL 91-97%] were generated when the two tests were interpreted in series (an animal had to be positive on both tests to be classified as combination positive). The presence of gross or microscopic lesions showed moderate to good agreement with the isolation of M. bovis (Kappa = 65-69%). The results showed that post-mortem inspection, histopathology and culture do not necessarily recognize the same infected animals and that the spectra of animals identified by the tests overlaps. PMID:8785715

Rohonczy, E B; Balachandran, A V; Dukes, T W; Payeur, J B; Rhyan, J C; Saari, D A; Whiting, T L; Wilson, S H; Jarnagin, J L

1996-04-01

236

High-Throughput Mycobacterial Interspersed Repetitive-Unit-Variable-Number Tandem-Repeat Genotyping for Mycobacterium tuberculosis Epidemiological Studies.  

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The emergence of drug-resistant forms of tuberculosis (TB) represents a major public health concern. Understanding the transmission routes of the disease is a key factor for its control and for the implementation of efficient interventions. Mycobacterial interspersed repetitive-unit-variable-number tandem-repeat (MIRU-VNTR) marker typing is a well-described method for lineage identification and transmission tracking. However, the conventional manual genotyping technique is cumbersome and time-consuming and entails many risks for errors, thus hindering its implementation and dissemination. We describe here a new approach using the QIAxcel system, an automated high-throughput capillary electrophoresis system that also carries out allele calling. This automated method was assessed on 1,824 amplicons from 82 TB isolates and tested with sets of markers of 15 or 24 loci. Overall allele-calling concordance between the methods from 140 to 1,317 bp was 98.9%. DNA concentrations and repeatability and reproducibility performances showed no biases in allele calling. Furthermore, turnaround time using this automated system was reduced by 81% compared to the conventional manual agarose gel method. In sum, this new automated method facilitates MIRU-VNTR genotyping and provides reliable results. Therefore, it is well suited for field genotyping. The implementation of this method will help to achieve accurate and cost-effective epidemiological studies, especially in countries with a high prevalence of TB, where the high number of strains complicates the surveillance of circulating lineages and requires efficient interventions to be carried out in an urgent manner. PMID:25428144

Gauthier, Marie; Bidault, Floriane; Mosnier, Amandine; Bablishvili, Nino; Tukvadze, Nestani; Somphavong, Silaphet; Paboriboune, Phimpha; Ocheretina, Oksana; Pape, Jean William; Paranhos-Baccala, Glaucia; Berland, Jean-Luc

2015-02-01

237

The mycobacterial glycolipid glucose monomycolate induces a memory T cell response comparable to a model protein antigen and no B cell response upon experimental vaccination of cattle  

Science.gov (United States)

Glycolipids are presented to T cells by human group 1 CD1 proteins, but are not used as subunit vaccines yet. Experimental immunizations with pure mycobacterial glucose monomycolate (GMM) and keyhole limpet haemocyanin (KLH) in cattle, a species which, unlike mice, expresses group 1 CD1, showed that GMM was equally efficient as KLH in generating T cell responses in blood, but not in the draining lymph node. Also, KLH induced strong antibody responses whereas GMM did not. These data suggest that non-overlapping T cell populations are targeted and demonstrate the potential of glycolipids as a special class of subunit vaccine candidates. PMID:19538998

Nguyen, Thi Kim Anh; Koets, Ad P.; Santema, Wiebren J.; van Eden, Willem; Rutten, Victor P.M.G.; Van Rhijn, Ildiko

2009-01-01

238

Real-Time PCR Assay Using Fine-Needle Aspirates and Tissue Biopsy Specimens for Rapid Diagnosis of Mycobacterial Lymphadenitis in Children  

OpenAIRE

A real-time PCR assay was developed to diagnose and identify the causative agents of suspected mycobacterial lymphadenitis. Primers and probes for the real-time PCR were designed on the basis of the internal transcribed spacer sequence, enabling the recognition of the genus Mycobacterium and the species Mycobacterium avium and M. tuberculosis. The detection limit for the assay was established at 1,100 CFU/ml of pus, and the specificity tests showed no false-positive reaction with other mycoba...

Coppenraet, E. S. Bruijnesteijn; Lindeboom, J. A.; Prins, J. M.; Peeters, M. F.; Claas, E. C. J.; Kuijper, E. J.

2004-01-01

239

Genes and Gene Therapy  

Science.gov (United States)

... correctly, a child can have a genetic disorder. Gene therapy is an experimental technique that uses genes to ... or prevent disease. The most common form of gene therapy involves inserting a normal gene to replace an ...

240

Drug-sensitive tuberculosis, multidrug-resistant tuberculosis, and nontuberculous mycobacterial pulmonary disease in nonAIDS adults: comparisons of thin-section CT findings  

Energy Technology Data Exchange (ETDEWEB)

The aim of this work was to compare thin-section CT (TSCT) findings of drug-sensitive (DS) tuberculosis (TB), multidrug-resistant (MDR) TB, and nontuberculous mycobacterial (NTM) pulmonary disease in nonAIDS adults. During 2003, 216 (113 DS TB, 35 MDR TB, and 68 NTM) patients with smear-positive sputum for acid-fast bacilli (AFB), and who were subsequently confirmed to have mycobacterial pulmonary disease, underwent thoracic TSCT. The frequency of lung lesion patterns on TSCT and patients' demographic data were compared. The commonest TSCT findings were tree-in-bud opacities and nodules. On a per-person basis, significant differences were found in the frequency of multiple cavities and bronchiectasis (P<0.001, chi-square test and multiple logistic regression analysis). Multiple cavities were more frequent in MDR TB than in the other two groups and extensive bronchiectasis in NTM disease (multiple logistic regression analysis). Patients with MDR TB were younger than those with DS TB or NTM disease (P<0.001, multiple logistic regression analysis). Previous tuberculosis treatment history was significantly more frequent in patients with MDR TB or NTM disease (P<0.001, chi-square test and multiple logistic regression analysis). In patients with positive sputum AFB, multiple cavities, young age, and previous tuberculosis treatment history imply MDR TB, whereas extensive bronchiectasis, old age, and previous tuberculosis treatment history NTM disease. (orig.)

Chung, Myung Jin; Lee, Kyung Soo; Kim, Tae Sung; Kim, Sung Mok [Sungkyunkwan University School of Medicine, Department of Radiology and Center for Imaging Science, Samsung Medical Center, Seoul (Korea); Koh, Won-Jung; Kwon, O Jung [Sungkyunkwan University School of Medicine, Division of Pulmonary and Critical Care Medicine, Department of Medicine, Samsung Medical Center, Seoul (Korea); Kang, Eun Young [Korea University Guro Hospital, Department of Diagnostic Radiology, Korea University College of Medicine, Seoul (Korea); Kim, Seonwoo [Sungkyunkwan University School of Medicine, Biostatistics Unit of the Samsung Biomedical Research Institute, Samsung Medical Center, Seoul (Korea)

2006-09-15

241

Drug-sensitive tuberculosis, multidrug-resistant tuberculosis, and nontuberculous mycobacterial pulmonary disease in nonAIDS adults: comparisons of thin-section CT findings  

International Nuclear Information System (INIS)

The aim of this work was to compare thin-section CT (TSCT) findings of drug-sensitive (DS) tuberculosis (TB), multidrug-resistant (MDR) TB, and nontuberculous mycobacterial (NTM) pulmonary disease in nonAIDS adults. During 2003, 216 (113 DS TB, 35 MDR TB, and 68 NTM) patients with smear-positive sputum for acid-fast bacilli (AFB), and who were subsequently confirmed to have mycobacterial pulmonary disease, underwent thoracic TSCT. The frequency of lung lesion patterns on TSCT and patients' demographic data were compared. The commonest TSCT findings were tree-in-bud opacities and nodules. On a per-person basis, significant differences were found in the frequency of multiple cavities and bronchiectasis (P<0.001, chi-square test and multiple logistic regression analysis). Multiple cavities were more frequent in MDR TB than in the other two groups and extensive bronchiectasis in NTM disease (multiple logistic regression analysis). Patients with MDR TB were younger than those with DS TB or NTM disease (P<0.001, multiple logistic regression analysis). Previous tuberculosis treatment history was significantly more frequent in patients with MDR TB or NTM disease (P<0.001, chi-square test and multiple logistic regression analysis). In patients with positive sputum AFB, multiple cavities, young age, and previous tuberculosis treatment history imply MDR TB, whereas extensive bronchiectasis, old age, and previous tuberculosis treatment history NTM disease. (orig.)story NTM disease. (orig.)

242

Both Corynebacterium diphtheriae DtxR(E175K) and Mycobacterium tuberculosis IdeR(D177K) Are Dominant Positive Repressors of IdeR-Regulated Genes in M. tuberculosis  

OpenAIRE

The diphtheria toxin repressor (DtxR) is an important iron-dependent transcriptional regulator of known virulence genes in Corynebacterium diphtheriae. The mycobacterial iron-dependent repressor (IdeR) is phylogenetically closely related to DtxR, with high amino acid similarity in the DNA binding and metal ion binding site domains. We have previously shown that an iron-insensitive, dominant-positive dtxR(E175K) mutant allele from Corynebacterium diphtheriae can be expressed in Mycobacterium t...

Manabe, Yukari C.; Hatem, Christine L.; Kesavan, Anup K.; Durack, Justin; Murphy, John R.

2005-01-01

243

The Differential Gene Expression Pattern of Mycobacterium tuberculosis in Response to Capreomycin and PA-824 versus First-Line TB Drugs Reveals Stress- and PE/PPE-Related Drug Targets  

OpenAIRE

Tuberculosis is a leading infectious disease causing millions of deaths each year. How to eradicate mycobacterial persistence has become a central research focus for developing next-generation TB drugs. Yet, the knowledge in this area is fundamentally limited and only a few drugs, notably capreomycin and PA-824, have been shown to be active against non-replicating persistent TB bacilli. In this study, we performed a new bioinformatics analysis on microarray-based gene expression data obtained...

Fu, Li M.; Tai, Shu C.

2009-01-01

244

ClpR Protein-like Regulator Specifically Recognizes RecA Protein-independent Promoter Motif and Broadly Regulates Expression of DNA Damage-inducible Genes in Mycobacteria*  

OpenAIRE

The RecA-dependent DNA damage response pathway (SOS response) appears to be the major DNA repair mechanism in most bacteria, but it has been suggested that a RecA-independent mechanism is responsible for controlling expression of most damage-inducible DNA repair genes in Mycobacterium tuberculosis. The specific reparative responses and molecular mediators involved in the DNA repair mechanism remain largely unclear in this pathogen and its related species. In this study, a mycobacterial ClpR-l...

Wang, Yi; Huang, Yuanxia; Xue, Chaolun; He, Yang; He, Zheng-guo

2011-01-01

245

Nontuberculous mycobacterial pulmonary infections  

OpenAIRE

Pulmonary infections due to nontuberculous mycobacteria (NTM) are increasingly recognized worldwide. Although over 150 different species of NTM have been described, pulmonary infections are most commonly due to Mycobacterium avium complex (MAC), Mycobacterium kansasii, and Mycobacterium abscessus. The identification of these organisms in pulmonary specimens does not always equate with active infection; supportive radiographic and clinical findings are needed to establish the diagnosis. It is ...

Johnson, Margaret M.; Odell, John A.

2014-01-01

246

Nontuberculous mycobacterial pulmonary infections.  

Science.gov (United States)

Pulmonary infections due to nontuberculous mycobacteria (NTM) are increasingly recognized worldwide. Although over 150 different species of NTM have been described, pulmonary infections are most commonly due to Mycobacterium avium complex (MAC), Mycobacterium kansasii, and Mycobacterium abscessus. The identification of these organisms in pulmonary specimens does not always equate with active infection; supportive radiographic and clinical findings are needed to establish the diagnosis. It is difficult to eradicate NTM infections. A prolonged course of therapy with a combination of drugs is required. Unfortunately, recurrent infection with new strains of mycobacteria or a relapse of infection caused by the original organism is not uncommon. Surgical resection is appropriate in selected cases of localized disease or in cases in which the infecting organism is resistant to medical therapy. Additionally, surgery may be required for infections complicated by hemoptysis or abscess formation. This review will summarize the practical aspects of the diagnosis and management of NTM thoracic infections, with emphasis on the indications for surgery and the results of surgical intervention. The management of NTM disease in patients with human immunodeficiency virus (HIV) infections is beyond the scope of this article and, unless otherwise noted, comments apply to hosts without HIV infection. PMID:24624285

Johnson, Margaret M; Odell, John A

2014-03-01

247

Mycobacterial infections in Finland.  

Science.gov (United States)

In Finland, the situation with regard to tuberculosis has been constantly improving during recent years, and the WHO criteria for a low incidence country (10/100,000) are presently satisfied. In contrast, the number of clinical specimens yielding atypical mycobacteria by isolation has rapidly increased. Organisms belonging in Mycobaterium avium-intracellulare complex dominate among the isolated species. PMID:8867168

Tala, E; Viljanen, M

1995-01-01

248

Disseminated infection due to Mycobacterium avium subsp. avium in an Asian elephant (Elephas maximus).  

Science.gov (United States)

A disseminated infection caused by Mycobacterium avium subspecies avium (MAA) was diagnosed in a 57-yr-old male Asian elephant (Elephas maximus) housed at the Seoul Zoo, Gyeonggi, Republic of Korea. An apparent granulomatous inflammation with central caseous necrosis was evident in the lung sections. To confirm mycobacterial infection, polymerase chain reaction-restriction enzyme polymorphism analysis (PCR-RFLP) of the rpoB and hsp65 genes was performed from multiple organs and cultured bacteria. The PCR-RFLP revealed a M. avium subspecies. MAA was identified by multiplex PCR for detection of IS901 and IS1311. Thus, it is believed that MAA caused the disseminated infection in this case. Although the source of infection was not determined, the elephant may have become infected through contamination of soil and feed by free-living birds infected with MAA. This is the first reported case of disseminated infection due to MAA in a captive elephant in the Republic of Korea. PMID:22204075

Yong, Hwanyul; Choi, Go-Eun; Lee, Byung Soo; Whang, Jake; Shin, Sung Jae

2011-12-01

249

Avian mycobacteriosis in psittacines: a retrospective study of 123 cases.  

Science.gov (United States)

One hundred and twenty-three cases of mycobacterioses were diagnosed in psittacine birds from a total of 9,241 submissions for necropsy examination or histopathology made to the California Animal Health and Food Safety Laboratory System between 1990 and 2007. The species affected most commonly were Amazon parrots (Amazona spp.)(n = 32; 26%) and grey-cheeked parakeets Brotogeris pyrrophterus (n = 23; 18.7%). The main gross findings on necropsy examination were enlarged and mottled pale livers and spleens and thickening of the small intestinal wall with numerous pale miliary nodules on the mucosa. Microscopical examination revealed infiltration of foamy macrophages and giant cells containing acid-fast bacteria in various organs. The gene encoding mycobacterial 65 kDa heat shock protein (hsp65) was amplified by nested polymerase chain reaction (PCR) from DNA extracted from 22 cases. The species of Mycobacterium involved was determined by analysis of restriction endonuclease patterns of the PCR products. Mycobacterium genavense was detected in 19 cases and Mycobacterium avium in two cases. One parrotlet (Touit spp.) had a mixed infection of both species of mycobacteria. It is concluded that M. genavense is the primary cause of mycobacteriosis in psittacine birds and the potential for zoonotic disease should be considered, especially for immunocompromised owners. PMID:22884283

Palmieri, C; Roy, P; Dhillon, A S; Shivaprasad, H L

2013-02-01

250

Mycobacterium salmoniphilum infection in burbot Lota lota.  

Science.gov (United States)

Burbot Lota lota sampled from lakes Mjosa and Losna in southeastern Norway between 2005 and 2008 were found to be infected with Mycobacterium salmoniphilum at a culture-positive prevalence of 18.6 and 3.3%, respectively. The condition factor (CF) of mycobacteria-affected fish sampled from Mjøsa in 2008 was lower than the average CF of total sampled fish the same year. Externally visible pathological changes included skin ulceration, petechiae, exopthalmia and cataract. Internally, the infections were associated with capsulated, centrally necrotic granulomas, containing large numbers of acid-fast bacilli, found mainly in the mesenteries, spleen, heart and swim bladder. Mycobacterial isolates recovered on Middlebrook 7H10 agar were confirmed as M. salmoniphilum by phenotypical investigation and by partial sequencing of the 16S rRNA, rpoB and Hsp65genes as well as the internal transcribed spacer (ITS1) locus. This study adds burbot to the list of fish species susceptible to piscine mycobacteriosis and describes M. salmoniphilum infection in a non-salmonid fish for the first time. PMID:21797036

Zerihun, Mulualem Adam; Berg, Vidar; Lyche, Jan L; Colquhoun, Duncan J; Poppe, Trygve T

2011-05-24

251

Evaluation of polymerase chain reaction and restriction enzyme analysis for routine identification of mycobacteria: accuracy, rapidity, and cost analysis.  

Science.gov (United States)

Polymerase chain reaction and restriction enzyme analysis (PCR-REA) of the hsp65 gene was evaluated for use as a routine identification method for identifying mycobacteria. The accuracy, rapidity, and cost were assessed compared with the conventional biochemical method. Five hundred and forty-one mycobacterial clinical isolates obtained from the Department of Microbiology, Faculty of Medicine at Siriraj Hospital, Mahidol University, were submitted for PCR-REA and biochemical identification. PCR-REA showed high concordant result with 100, 96.2, and 94.1% for identification of Mycobacterium tuberculosis, rapid- and slow-growing mycobacteria, respectively. Discordant results were obtained from 24 (4.4%) out of 541 isolates, consisting of 9 rapid growers (6 M. chelonae, 2 M. abscessus, and 1 M. fortuitum) and 15 slow growers (9 M. scrofulaceum, 2 M. gordonae, 1 M. avium, 1 M. kansasii, 1 M. malmoense, and 1 M. terrae complex). PCR-REA demonstrated not only accurate results but was also less expensive (2.1 US dollars/sample). This method was rapid with a turn-around time of 30 hours compared with 2-4 weeks for the conventional method. PMID:16438154

Prammananan, Therdsak; Cheunoy, Wattana; Na-Ubol, Preeyawit; Tingtoy, Nipa; Srimuang, Somboon; Chaiprasert, Angkana

2005-09-01

252

Evaluation of INNO-LiPA mycobacteria v2 assay for identification of rapidly growing mycobacteria  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: English Abstract in english A total of 54 rapidly growing mycobacteria (RGM) isolated from patients attended in the two hospitals of Cádiz Bay (Spain) were selected during a seven-year-period (2000-2006) in order to evaluate the INNO-LiPA Mycobacteria v2 assay for mycobacterial identification, based on the reverse hybridizatio [...] n principle. The strains were cultured in Löwenstein-Jensen and Middlebrook 7H9 media and identified to the species level by sequencing of the 16S rRNA, PCR-restriction enzyme analysis of the hsp65 gene, conventional tests and INNO-LiPA Mycobacteria v2 assay. By the molecular methods we identified a total of 12 different species: 23 Mycobacterium fortuitum, 11 M. chelonae, 10 M. abscessus, 2 M. senegalense, 1 M. alvei, 1 M. brumae, 1 M. mageritense, 1 M. mucogenicum, 1 M. neoaurum, 1 M. peregrinum, 1 M. septicum and 1 M. smegmatis. Fifty two strains (96.3%) were correctly identified by conventional techniques and 47 strains (87.0%) by INNO-LiPA Mycobacteria v2 assay. We find INNO-LiPA Mycobacteria v2 assay simple to perform but it provides few advantages in comparison with conventional methods and sometimes needs complementary tests to identify Mycobacterium fortuitum complex, M. chelonae complex and specific species due to the great heterogeneity in the RGM group.

Lidia, García-Agudo; Iría, Jesús; Manuel, Rodríguez-Iglesias; Pedro, García-Martos.

1220-12-01

253

Evaluation of the post-antibiotic effect of six anti-mycobacterial agents against Mycobacterium avium by the Bactec radiometric method.  

Science.gov (United States)

We compared and evaluated the duration of the post-antibiotic effect (PAE) of six anti-mycobacterial agents (ethambutol, isoniazid, rifampicin, amikacin, ciprofloxacin and clarithromycin), alone and in combination, against ten strains of Mycobacterium avium. PAE was determined by conventional viable counting and the Bactec growth system, analysed by the T100 value (the time to produce a cumulative growth index of 100). A 2 h exposure to the drug did not induce significant killing. Thus, comparison of regrowth could be evaluated without the potential need to adjust viable counts. The correlation of duration of PAE between the viable count method and the T100 Bactec method was excellent (r = 0.94). No significant PAE ( or = 8 h) which was observed in half the strains tested. It was concluded that the T100 Bactec method is highly suitable for evaluating PAE against M. avium, producing a definitive result in 5 days. PMID:9249202

Fuursted, K

1997-07-01

254

Comparison of the anti-inflammatory activity of Commiphora mukul (an indigenous drug) with those of phenylbutazone and ibuprofen in experimental arthritis induced by mycobacterial adjuvant.  

Science.gov (United States)

In the present investigation a method of induction of experimental arthritis in animals was modified to provide a better model replica of human arthritis. Inflammatory syndrome, resembling rheumatoid arthritis in man, was induced in the right hock joint of albino rabbits by intra-articular injection of the killed mycobacterial adjuvant in liquid paraffin. Development of this arthritic syndrome was studied from a period of five months with and without drugs. Anti-inflammatory agents such as phenylbutazone, ibuprofen and fraction "A" of gum-guggual from Commiphora mukkul were administered orally at a daily dose of 100, 100 and 500 mg/kg, respectively, for a period of five months. All three drugs decreased the thickness of the joint swelling during the course of drug treatment. These results indicate the beneficial role of phenylbutazone, ibuprofen and fraction "A" of gum-guggul in experimental arthritis. PMID:578471

Sharma, J N; Sharma, J N

1977-07-01

255

Rapid Rebound of the Treg Compartment in DEREG Mice Limits the Impact of Treg Depletion on Mycobacterial Burden, but Prevents Autoimmunity  

Science.gov (United States)

The development of an effective vaccine against tuberculosis (Tb) represents one of the major medical challenges of this century. Mycobacterium bovis Bacille Calmette-Guerin (BCG), the only vaccine available at present, is mostly effective at preventing disseminated Tb in children, but shows variable protection against pulmonary Tb, the most common form in adults. The reasons for this poor efficacy are not completely understood, but there is evidence that T regulatory cells (Tregs) might be involved. Similarly, Tregs have been associated with the immunosuppression observed in patients infected with Tb and are therefore believed to play a role in pathogen persistence. Thus, Treg depletion has been postulated as a novel strategy to potentiate M. bovis BCG vaccination on one side, while on the other, employed as a therapeutic approach during chronic Tb infection. Yet since Tregs are critically involved in controlling autoimmune inflammation, elimination of Tregs may therefore also incur the danger of an excessive inflammatory immune response. Thus, understanding the dynamics and function of Tregs during mycobacterial infection is crucial to evaluate the potential of Treg depletion as a medical option. To address this, we depleted Tregs after infection with M. bovis BCG or Mycobacterium tuberculosis (Mtb) using DEREG mice, which express the diphtheria toxin (DT) receptor under the control of the FoxP3 locus, thereby allowing the selective depletion of FoxP3+ Tregs. Our results show that after depletion, the Treg niche is rapidly refilled by a population of DT-insensitive Tregs (diTregs) and bacterial load remains unchanged. On the contrary, impaired rebound of Tregs in DEREG × FoxP3GFP mice improves pathogen burden, but is accompanied by detrimental autoimmune inflammation. Therefore, our study provides the proof-of-principle that, although a high degree of Treg depletion may contribute to the control of mycobacterial infection, it carries the risk of autoimmunity. PMID:25050936

Varela, Filipa; Behrends, Jochen; Swallow, Maxine; Kruse, Friederike; Krull, Freyja; Ghorbani, Peyman; Mayer, Christian T.

2014-01-01

256

Genes of Mycobacterium tuberculosis H37Rv downregulated in the attenuated strain H37Ra are restricted to M. tuberculosis complex species.  

Science.gov (United States)

By comparing gene expression of virulent Mycobacterium tuberculosis H37Rv and attenuated strain H37Ra, we previously detected six genes that appear to be markedly downregulated in the attenuated strain compared with the virulent one. Three of these genes, i.e. Rv1345, Rv2770c, and Rv0288, code for proteins that can be predictively associated to immunological or pathogenetic aspects of M. tuberculosis infection; the other genes, i.e. Rv2336, Rv1320c, and Rv2819c, code for proteins with unknown functions (Rindi et al., 1999). In this paper we searched for the above mentioned genes in Pvu II-digested genomic DNA of a number of mycobacterial species by southern blot analysis employing PCR-generated probes in high-stringency conditions. Hybridization signals were only found in species belonging to the M. tuberculosis complex, i.e., M. tuberculosis, M. bovis, including the BCG strain, and M. microti, but not in other mycobacterial species, including M. avium, M. intracellulare, M. malmoense, M. xenopi, M. kansasii, M. simiae, M. marinum, M. scrofulaceum, M. gordonae, M. fortuitum, and M. smegmantis. These results indicate that genes Rv1345, Rv2770c, Rv0288, Rv2336, Rv1320c, and Rv2819c are associated with the most virulent mycobacteria and further support their potential role in M. tuberculosis virulence. PMID:11497087

Rindi, L; Lari, N; Garzelli, C

2001-07-01

257

Estimation of the spread of pathogenic mycobacteria in organic broiler farms by the polymerase chain reaction.  

Science.gov (United States)

Organic poultry breeding allows for increased exposure of birds to soil, faeces, and wildlife, which have been associated with the transmission of mycobacterial infections. Therefore the aim of this study was to investigate the spread of the major pathogenic mycobacteria in organically reared broilers in Greece using a diagnostic algorithm that relied on a combination of the polymerase chain reaction (PCR) and the restriction fragment length polymorphism analysis (RFLP). Liver, spleen and gonads from 81 to 150 days old broilers were aseptically collected post-mortem. 500 broilers from a population of 35,370, reared in the 25 registered as organic farms in Greece for the 2005 were used. DNA was isolated and incorporated to PCR targeted to 16S-rRNA gene (for Mycobacterium spp.), IS6110 (for Mycobacterium tuberculosis complex-MTBc), IS1245 (for Mycobacterium avium complex-MAC), IS901 (for M. avium subsp. avium-MAA) and hsp65 (for Mycobacterium genavense, by PCR-RFLP). The mean prevalence of mycobacteria detected by PCR with a 95% confidence interval was estimated to 4.4-8.8%. The relevant percentage with regard to the mycobacterial species that were included in this study was 0.17-2.03% for MAC, 2.11-3.39% for MTBc and 0.66-3.08% for mycobacteria not belonging to any of the above groups. None of the mycobacteria detected were identified as MAA or M. genavense. Considering that avian tuberculosis has been eradicated from conventional farms, the level and the pattern of positivity recorded here, indicates that our results may be associated with the specific conditions that apply to organic breeding. PMID:18774661

Ikonomopoulos, J; Fragkiadaki, E; Liandris, E; Sotirakoglou, K; Xylouri, E; Gazouli, M

2009-01-13

258

Mutation in alkylhydroperoxidase D gene dramatically decreases persistence of Mycobacterium bovis bacillus calmette-guerin in infected macrophage  

Directory of Open Access Journals (Sweden)

Full Text Available Background and Objectives: Mycobacterium tuberculosis is the leading cause of death from a single bacterial species in the world and is subjected to a highly oxidative environment in its host macrophage and consequently has evolved protective mechanisms against reactive oxygen and nitrogen intermediates. Alkyl hydroperoxidase D (AhpD is a molecule from these mycobacterial defense systems that has a dual function. It not only works with Alkyl hydroperoxidase C (AhpC in mycobacterial defense system against oxidative stress but also has a role in oxidation/reduction of succinyltransferase B (SucB, dihydrolipoamide dehydrogenase (LPD and AhpC. The present study was undertaken to find out the effects of inactivation of ahpD gene in the intra-macrophage persistence of resulted BCG mutant. Materials and Methods: We did allelic exchange mutagenesis in Mycobacterium bovis BCG and evaluate the effects of this mutagenesis in intracellular persistence of wild type BCG strains and ahpD mutant ones by comparing colony forming units (CFU in infected macrophage. Results: Our findings showed that after producing allelic exchange mutagenesis in ahpD gene of M.bovis BCG a sever decrease in the CFU?s of ahpD mutant BCG strains has been observed and intracellular persistence of ahpD mutant BCG strains decreased significantly. Conclusion: Mutagenesis in ahpD gene will cause significant decrease in intracellular survival of ahpD mutant strains than wild type M.bovis BCG strains and could leads to an inefficiency in pyruvate dehydrogenase pathway and could also impair impairs mycobacterial defense system against oxidative and nitrosative stress.

Farivar Taghi

2008-07-01

259

Determination of Genotypic Diversity of Mycobacterium avium Subspecies from Human and Animal Origins by Mycobacterial Interspersed Repetitive-Unit-Variable-Number Tandem-Repeat and IS1311 Restriction Fragment Length Polymorphism Typing Methods ? †  

OpenAIRE

Members of the Mycobacterium avium complex (MAC) are ubiquitous bacteria that can be found in water, food, and other environmental samples and are considered opportunistic pathogens for numerous animal species, mainly birds and pigs, as well as for humans. We have recently demonstrated the usefulness of a PCR-based mycobacterial interspersed repetitive-unit-variable-number tandem-repeat (MIRU-VNTR) typing for the molecular characterization of M. avium subsp. paratuberculosis and M. avium stra...

Radomski, Nicolas; Thibault, Virginie C.; Karoui, Claudine; Cruz, Krystel; Cochard, Thierry; Gutie?rrez, Cristina; Supply, Philip; Biet, Frank; Boschiroli, Mari?a Laura

2010-01-01

260

Novel, potent, orally bioavailable and selective mycobacterial ATP synthase inhibitors that demonstrated activity against both replicating and non-replicating M. tuberculosis.  

Science.gov (United States)

The mycobacterial F0F1-ATP synthase (ATPase) is a validated target for the development of tuberculosis (TB) therapeutics. Therefore, a series of eighteen novel compounds has been designed, synthesized and evaluated against Mycobacterium smegmatis ATPase. The observed ATPase inhibitory activities (IC50) of these compounds range between 0.36 and 5.45?M. The lead compound 9d [N-(7-chloro-2-methylquinolin-4-yl)-N-(3-((diethylamino)methyl)-4-hydroxyphenyl)-2,3-dichlorobenzenesulfonamide] with null cytotoxicity (CC50>300?g/mL) and excellent anti-mycobacterial activity and selectivity (mycobacterium ATPase IC50=0.51?M, mammalian ATPase IC50>100?M, and selectivity >200) exhibited a complete growth inhibition of replicating Mycobacterium tuberculosis H37Rv at 3.12?g/mL. In addition, it also exhibited bactericidal effect (approximately 2.4log10 reductions in CFU) in the hypoxic culture of non-replicating M. tuberculosis at 100?g/mL (32-fold of its MIC) as compared to positive control isoniazid [approximately 0.2log10 reduction in CFU at 5?g/mL (50-fold of its MIC)]. The pharmacokinetics of 9d after p.o. and IV administration in male Sprague-Dawley rats indicated its quick absorption, distribution and slow elimination. It exhibited a high volume of distribution (Vss, 0.41L/kg), moderate clearance (0.06L/h/kg), long half-life (4.2h) and low absolute bioavailability (1.72%). In the murine model system of chronic TB, 9d showed 2.12log10 reductions in CFU in both lung and spleen at 173?mol/kg dose as compared to the growth of untreated control group of Balb/C male mice infected with replicating M. tuberculosis H37Rv. The in vivo efficacy of 9d is at least double of the control drug ethambutol. These results suggest 9d as a promising candidate molecule for further preclinical evaluation against resistant TB strains. PMID:25614114

Singh, Supriya; Roy, Kuldeep K; Khan, Shaheb R; Kashyap, Vivek Kr; Sharma, Abhisheak; Jaiswal, Swati; Sharma, Sandeep K; Krishnan, Manju Yasoda; Chaturvedi, Vineeta; Lal, Jawahar; Sinha, Sudhir; Gupta, Arnab D; Srivastava, Ranjana; Saxena, Anil K

2015-02-15

261

Capacity of murine T cells to retain long-term responsiveness to mycobacterial antigens is controlled by the H-2 complex.  

Science.gov (United States)

It is firmly established that the allelic composition of the H-2 complex has a prominent impact on the course of tuberculosis (TB) infection in mice, including granuloma formation, mycobacterial spread in the lungs, and the dynamics of mortality. Although intuitively obvious, the role of long-term specific T cell responses in the expression of corresponding phenotypes is poorly understood. In this study we have compared polyclonal lymph node cell response (cell yield, proliferation, surface markers, IL-4/interferon-gamma (IFN-gamma) production) to Mycobacterium tuberculosis H37Rv sonicate in repeated 10-day cycles of stimulation/rest between H-2 congenic IE-negative mouse strains, categorized on the basis of mortality following lethal challenge as TB-susceptible (C57B1/6), TB-resistant (4R) and BCG non-protected (B10.M). The capacity to retain specific responsiveness to repeated stimulation by mycobacterial antigens depended upon both the H-2 haplotype of the host and the immunizing dose of the antigen. 4R lymph node cells following either 50 microg/mouse or 100 microg/mouse immunization constantly responded to sonicate, increased in numbers, and after the third stimulation/rest cycle developed into a stable CD3+CD4+ cell line. B6 cells following either 50 microg/mouse or 100 microg/mouse immunization, and B10.M cells following 100 microg/mouse (but not 50 microg/mouse) immunization, lost the capacity to incorporate methyl-3H-thymidine during the second cycle, and died. Analogous results were obtained in the in vivo experiments, when the dynamics of the response over 12 weeks following a single immunization with the antigen was studied. In response to the antigen, cells from all three mouse strains produced significant amounts of IL-2 and IFN-gamma, but not IL-4, indicating that they belong predominantly to the Th1-like subset. Among noteworthy differences between the mouse strains was a clear deficiency of CD8+ T cells in B6 cultures, and an unusually high proportion of CD3+CD4-CD8- (double-negative) T cells in B10.M cultures following a high-dose immunization. PMID:9486398

Pichugin, A V; Khaidukov, S V; Moroz, A M; Apt, A S

1998-02-01

262

The Contraceptive Depot Medroxyprogesterone Acetate Impairs Mycobacterial Control and Inhibits Cytokine Secretion in Mice Infected with Mycobacterium tuberculosis  

OpenAIRE

The contraceptive depot medroxyprogesterone acetate (DMPA), with progestin as the single active compound, possesses selective glucocorticoid activity and can alter the expression of glucocorticoid receptor-regulated genes. We therefore propose that pharmacological doses of DMPA used for endocrine therapy could have significant immune modulatory effects and impact on susceptibility to, as well as clinical manifestation and outcome of, infectious diseases. We investigated the effect of contrace...

Kleynhans, Le?anie; Du Plessis, Nelita; Allie, Nasiema; Jacobs, Muazzam; Kidd, Martin; Helden, Paul D.; Walzl, Gerhard; Ronacher, Katharina

2013-01-01

263

Structural Plasticity and Distinct Drug-Binding Modes of LfrR, a Mycobacterial Efflux Pump Regulator?  

OpenAIRE

The TetR-like transcriptional repressor LfrR controls the expression of the gene encoding the Mycobacterium smegmatis efflux pump LfrA, which actively extrudes fluoroquinolones, cationic dyes, and anthracyclines from the cell and promotes intrinsic antibiotic resistance. The crystal structure of the apoprotein form of the repressor reveals a structurally asymmetric homodimer exhibiting local unfolding and a blocked drug-binding site, emphasizing the significant conformational plasticity of th...

Bellinzoni, Marco; Buroni, Silvia; Schaeffer, Francis; Riccardi, Giovanna; Rossi, Edda; Alzari, Pedro M.

2009-01-01

264

Use of the hupB Gene Encoding a Histone-Like Protein of Mycobacterium tuberculosis as a Target for Detection and Differentiation of M. tuberculosis and M. bovis  

OpenAIRE

The gene for histone-like protein (hupB [Rv2986c]) of Mycobacterium tuberculosis has been identified as a singular target which allows differentiation of two closely related mycobacterial species, namely, M. tuberculosis and M. bovis of the MTB complex, by a PCR assay. The N and S primer-generated PCR amplicons differed in M. tuberculosis and M. bovis; these amplicons were determined to be 645 and 618 bp, respectively. This difference was localized to the C-terminal part of the gene by using ...

Prabhakar, S.; Mishra, A.; Singhal, A.; Katoch, V. M.; Thakral, S. S.; Tyagi, J. S.; Prasad, H. K.

2004-01-01

265

T cell reactivity against mycolyl transferase antigen 85 of M. tuberculosis in HIV-TB coinfected subjects and in AIDS patients suffering from tuberculosis and nontuberculous mycobacterial infections.  

Science.gov (United States)

The mycolyl transferase antigen 85 complex is a major secreted protein family from mycobacterial culture filtrate, demonstrating powerful T cell stimulatory properties in most HIV-negative, tuberculin-positive volunteers with latent M.tuberculosis infection and only weak responses in HIV-negative tuberculosis patients. Here, we have analyzed T cell reactivity against PPD and Ag85 in HIV-infected individuals, without or with clinical symptoms of tuberculosis, and in AIDS patients with disease caused by nontuberculous mycobacteria. Whereas responses to PPD were not significantly different in HIV-negative and HIV-positive tuberculin-positive volunteers, responses to Ag85 were significantly decreased in the HIV-positive (CDC-A and CDC-B) group. Tuberculosis patients demonstrated low T cell reactivity against Ag85, irrespective of HIV infection, and finally AIDS patients suffering from NTM infections were completely nonreactive to Ag85. A one-year follow-up of twelve HIV-positive tuberculin-positive individuals indicated a decreased reactivity against Ag85 in patients developing clinical tuberculosis, highlighting the protective potential of this antigen. PMID:20936150

Launois, Pascal; Drowart, Annie; Bourreau, Eliane; Couppie, Pierre; Farber, Claire-Michèle; Van Vooren, Jean-Paul; Huygen, Kris

2011-01-01

266

Patients with non-tuberculous mycobacterial lung disease have elevated transforming growth factor-beta following ex vivo stimulation of blood with live Mycobacterium intracellulare.  

Science.gov (United States)

We previously found that a subset of patients with pulmonary non-tuberculous mycobacterial (pNTM) disease were taller, leaner, and had a higher prevalence of pectus excavatum and scoliosis than uninfected controls. Additionally, whole blood of pNTM patients stimulated ex vivo with live Mycobacterium intracellulare produced significantly less interferon-gamma (IFN?) compared to that of uninfected controls. Since IFN? production can be suppressed by transforming growth factor-beta (TGF?), an immunosuppressive cytokine, we measured basal and M. intracellulare-stimulated blood levels of TGF? in a group of 20 pNTM patients and 20 uninfected controls. In contrast to the IFN? findings, we found that stimulated blood from pNTM patients produced significantly higher levels of TGF? compared to controls. Since pNTM patients frequently possess body features that overlap with Marfan syndrome (MFS), and increased TGF? expression is important in the pathogenesis of MFS, we posit that a yet-to-be-identified syndrome related to MFS predisposes certain individuals to develop pNTM disease. PMID:23808720

Ovrutsky, Alida R; Merkel, Patricia A; Schonteich, Eric; Bai, Xiyuan; Kinney, William; Iseman, Michael D; Kartalija, Marinka; Knight, Vijaya; Chan, Edward D

2013-09-01

267

Comparison of spoligotyping, mycobacterial interspersed repetitive units typing and IS6110-RFLP in a study of genotypic diversity of Mycobacterium tuberculosis in Delhi, North India  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: English Abstract in english The aim of the present study was to compare polymerase chain reaction (PCR)-based methods - spoligotyping and mycobacterial interspersed repetitive units (MIRU) typing - with the gold-standard IS6110 restriction fragment length polymorphism (RFLP) analysis in 101 isolates of Mycobacterium tuberculos [...] is to determine the genetic diversity of M. tuberculosis clinical isolates from Delhi, North India. Spoligotyping resulted in 49 patterns (14 clusters); the largest cluster was composed of Spoligotype International Types (SITs)26 [Central-Asian (CAS)1-Delhi lineage], followed by SIT11 [East-African-Indian (EAI) 3-Indian lineage]. A large number of isolates (75%) belonged to genotypic lineages, such as CAS, EAI and Manu, with a high specificity for the Indian subcontinent, emphasising the complex diversity of the phylogenetically coherent M. tuberculosis in North India. MIRU typing, using 11 discriminatory loci, was able to distinguish between all but two strains based on individual patterns. IS6110-RFLP analysis (n = 80 strains) resulted in 67 unique isolates and four clusters containing 13 strains. MIRUs discriminated all 13 strains, whereas spoligotyping discriminated 11 strains. Our results validate the use of PCR-based molecular typing of M. tuberculosis using repetitive elements in Indian isolates and demonstrate the usefulness of MIRUs for discriminating low-IS6110-copy isolates, which accounted for more than one-fifth of the strains in the present study.

Mandira, Varma-Basil; Sujeet, Kumar; Jyoti, Arora; Archana, Angrup; Thierry, Zozio; Jayant Nagesh, Banavaliker; Urvashi Balbir, Singh; Nalin, Rastogi; Mridula, Bose.

2011-08-01

268

Comparison of spoligotyping, mycobacterial interspersed repetitive units typing and IS6110-RFLP in a study of genotypic diversity of Mycobacterium tuberculosis in Delhi, North India  

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Full Text Available The aim of the present study was to compare polymerase chain reaction (PCR-based methods - spoligotyping and mycobacterial interspersed repetitive units (MIRU typing - with the gold-standard IS6110 restriction fragment length polymorphism (RFLP analysis in 101 isolates of Mycobacterium tuberculosis to determine the genetic diversity of M. tuberculosis clinical isolates from Delhi, North India. Spoligotyping resulted in 49 patterns (14 clusters; the largest cluster was composed of Spoligotype International Types (SITs26 [Central-Asian (CAS1-Delhi lineage], followed by SIT11 [East-African-Indian (EAI 3-Indian lineage]. A large number of isolates (75% belonged to genotypic lineages, such as CAS, EAI and Manu, with a high specificity for the Indian subcontinent, emphasising the complex diversity of the phylogenetically coherent M. tuberculosis in North India. MIRU typing, using 11 discriminatory loci, was able to distinguish between all but two strains based on individual patterns. IS6110-RFLP analysis (n = 80 strains resulted in 67 unique isolates and four clusters containing 13 strains. MIRUs discriminated all 13 strains, whereas spoligotyping discriminated 11 strains. Our results validate the use of PCR-based molecular typing of M. tuberculosis using repetitive elements in Indian isolates and demonstrate the usefulness of MIRUs for discriminating low-IS6110-copy isolates, which accounted for more than one-fifth of the strains in the present study.

Mandira Varma-Basil

2011-08-01

269

Phenolic-glycolipid-1 and lipoarabinomannan preferentially modulate TCR- and CD28-triggered proximal biochemical events, leading to T-cell unresponsiveness in mycobacterial diseases  

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Full Text Available Background Advanced stages of leprosy show T cell unresponsiveness and lipids of mycobacterial origin are speculated to modulate immune responses in these patients. Present study elucidates the role of phenolicglycolipid (PGL-1 and Mannose-capped lipoarabinomannan (Man-LAM on TCR- and TCR/CD28- mediated signalling. Results We observed that lipid antigens significantly inhibit proximal early signalling events like Zap-70 phosphorylation and calcium mobilization. Interestingly, these antigens preferentially curtailed TCR-triggered early downstream signalling events like p38 phosphorylation whereas potentiated that of Erk1/2. Further, at later stages inhibition of NFAT binding, IL-2 message, CD25 expression and T-cell blastogenesis by PGL-1 and Man-LAM was noted. Conclusion Altogether, we report that Man-LAM and PGL-1 preferentially interfere with TCR/CD28-triggered upstream cell signalling events, leading to reduced IL-2 secretion and T-cell blastogenesis which potentially could lead to immunosupression and thus, disease exacerbation, as noted in disease spectrum.

Dagur Pradeep

2012-09-01

270

Reactividad serológica y celular frente a proteínas micobacterianas en la enfermedad de Hansen / Serological and cellular reactivity to mycobacterial proteins in Hansen´s disease  

Scientific Electronic Library Online (English)

Full Text Available SciELO Venezuela | Language: Spanish Abstract in spanish Se diseñó un estudio para evaluar la reactividad inmunológica frente a diferentes preparaciones proteicas micobacterianas utilizando pruebas serológicas y de inmunidad celular. Para el estudio fueron incluídos pacientes con manifestaciones clínicas de lepra predominantemente de la forma multibacilar [...] . Todos los pacientes fueron adultos con edad comprendida entre 20 y 39 años. El 58% correspondía a la forma clínica de Lepra Lepromatosa (LL) n= 81, el 29% a la forma Borderline Lepromatosa (BL) n=41 y 10% a Borderline Borderline (BB) n=14. Solo el 3% fueron pacientes Borderline Tuberculoide (BT): 74% masculino y 26% femenino. El fenómeno reaccional más frecuente fue del tipo eritema nodoso leproso (ENL). Las proteínas micobacterianas ensayadas fueron: antígenos proteicos crudos totales de Mycobacterium leprae (MlSA), Mycobacterium bovis (MbSA y MbSA de excreción), antígeno proteico de excreción parcialmente purificado con una movilidad relativa de 30 kDa (Ml 30) y proteínas recombinantes de Mycobacterium (Mt70, Mb 65, Ml 36, 28, 18 y 10 kDa) encontrandose que las proteínas recombinantes (Ml10 kDa, Ml 36 kDa) a mayor carga bacilar presentaban una mayor reactividad serológica estadísticamente significativa (p= 0,0051 y 0,050 respectivamente). La proteína de 30 kDa fue predominantemente reconocida por anticuerpos de los pacientes multibacilares. Los resultados demuestran que el promedio de los valores de anticuerpos en pacientes no reaccionales fueron superiores en presencia de proteínas completas (MbSA y MbSA de exc) en comparación con el grupo de pacientes que presentaron fenómenos reaccionales (p=0,000567 y 0,000061 respectivamente) Este mismo comportamiento se observó frente a las proteínas micobacterianas individuales (30 kDa, 10 kDa y 36 kDa). La respuesta proliferativa de los linfocitos T en los pacientes multibacilares reaccionales y no reaccionales frente a las proteínas micobacterianas (MlSA, Ml 10 kDa, MbSA, MbSA de excreción) fue negativa en ambos grupos. Abstract in english The study was designed for evaluating immunological reactivity to various mycobacterial protein preparations using serological and cell-mediated immunological tests in patients with clinical leprosy signs, predominantly, with the multibacillary forms. All patients were adults with ages between 20 an [...] d 30 years. Fifty eight (n= 81) percent corresponded to Lepromatous Leprosy (LL), 29% (n= 41) to Borderline Lepromatous Leprosy (BL) and 10% (n=41) to Borderline Borderline Leprosy (BB); only 3% were Borderline Tuberculoid (BT) patients: 74% males and 26% females. The most frequent reactional phenomenon was of the Erythema Nodosum (ENL) type. The mycobacterial proteins tested were: total crude Mycobacterium leprae antigens (MISA); Mycobacterium bovis (MbSA and excretion MbSA); partially purified excretion protein antigen, with a 30kDa relative movility (Ml30); and recombinant M. leprae proteins (Mt70, Mb 65, Ml 36, 28, 18 and 10 kDa). Two of the recombinant proteins (Ml10 and Ml 36 kDa) presented a statiscally significant higher serological reactivity, directly related with a larger bacillary load (p= 0.0051 and 0.050 respectively). The 30 kDa protein was predominantly recognized by antibodies from multibacillary patients. Results show that mean antibody values were higher in non reactional patients when tested against complete proteins (MbSA and ex MbSA) when compared with the group of patients who presented reactional phenomena (p= 0.000567 and 0.000061, respectively). Comparing reactional with non reactional patients, it was seen that mean antibody values against complete proteins (MbSA and ex MbSA) were higher in non reactional individuals (p= 0.000567 and 0.000061, respectively). This same behavior occurred towards individual mycobacterial proteins (30, 10 and 36 kDa). The T lymphocyte prolypherative response in reactional and non reactional patients towards mycobacterial proteins (MlSA, Ml 10 kDa, MbSA, ex MbSA) was negative.

Elsa, Rada; Nacarid, Aranzazu; Vestalia, Rodríguez; Rafael, Borges; Jacinto, Convit.

2010-09-01

271

Effect of study design and setting on tuberculosis clustering estimates using Mycobacterial Interspersed Repetitive Units-Variable Number Tandem Repeats (MIRU-VNTR): a systematic review  

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Objectives To systematically review the evidence for the impact of study design and setting on the interpretation of tuberculosis (TB) transmission using clustering derived from Mycobacterial Interspersed Repetitive Units-Variable Number Tandem Repeats (MIRU-VNTR) strain typing. Data sources MEDLINE, EMBASE, CINHAL, Web of Science and Scopus were searched for articles published before 21st October 2014. Review methods Studies in humans that reported the proportion of clustering of TB isolates by MIRU-VNTR were included in the analysis. Univariable meta-regression analyses were conducted to assess the influence of study design and setting on the proportion of clustering. Results The search identified 27 eligible articles reporting clustering between 0% and 63%. The number of MIRU-VNTR loci typed, requiring consent to type patient isolates (as a proxy for sampling fraction), the TB incidence and the maximum cluster size explained 14%, 14%, 27% and 48% of between-study variation, respectively, and had a significant association with the proportion of clustering. Conclusions Although MIRU-VNTR typing is being adopted worldwide there is a paucity of data on how study design and setting may influence estimates of clustering. We have highlighted study design variables for consideration in the design and interpretation of future studies. PMID:25609667

Mears, Jessica; Abubakar, Ibrahim; Cohen, Theodore; McHugh, Timothy D; Sonnenberg, Pam

2015-01-01

272

Novel Role of Phosphorylation-Dependent Interaction between FtsZ and FipA in Mycobacterial Cell Division  

OpenAIRE

The bacterial divisome is a multiprotein complex. Specific protein-protein interactions specify whether cell division occurs optimally, or whether division is arrested. Little is known about these protein-protein interactions and their regulation in mycobacteria. We have investigated the interrelationship between the products of the Mycobacterium tuberculosis gene cluster Rv0014c-Rv0019c, namely PknA (encoded by Rv0014c) and FtsZ-interacting protein A, FipA (encoded by Rv0019c) and the produc...

Sureka, Kamakshi; Hossain, Tofajjen; Mukherjee, Partha; Chatterjee, Paramita; Datta, Pratik; Kundu, Manikuntala; Basu, Joyoti

2010-01-01

273

Elucidation of the Dual Role of Mycobacterial MoeZR in Molybdenum Cofactor Biosynthesis and Cysteine Biosynthesis  

OpenAIRE

The pathway of molybdenum cofactor biosynthesis has been studied in detail by using proteins from Mycobacterium species, which contain several homologs associated with the first steps of Moco biosynthesis. While all Mycobacteria species contain a MoeZR, only some strains have acquired an additional homolog, MoeBR, by horizontal gene transfer. The role of MoeBR and MoeZR was studied in detail for the interaction with the two MoaD-homologs involved in Moco biosynthesis, MoaD1 and MoaD2, in addi...

Voss, Martin; Nimtz, Manfred; Leimku?hler, Silke

2011-01-01

274

Antigen Specificity in Experimental Bovine Tuberculosis  

OpenAIRE

This report describes the kinetics of T-cell responses to a panel of mycobacterial antigens (PPD-M, PPD-A, ESAT-6, Ag85, 38kD, MPB64, MPB70, MPB83, hsp16.1, hsp65, and hsp70) following experimental infection of cattle with Mycobacterium bovis. Increased antigen-specific lymphocyte proliferation, gamma interferon, and interleukin-2 responses were observed in all calves following infection. Positive lymphocyte proliferation and cytokine responses to PPD-M and ESAT-6 were observed throughout the...

Rhodes, S. G.; Gavier-widen, D.; Buddle, B. M.; Whelan, A. O.; Singh, M.; Hewinson, R. G.; Vordermeier, H. M.

2000-01-01

275

Inositol monophosphate phosphatase genes of Mycobacterium tuberculosis  

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Full Text Available Abstract Background Mycobacteria use inositol in phosphatidylinositol, for anchoring lipoarabinomannan (LAM, lipomannan (LM and phosphatidylinosotol mannosides (PIMs in the cell envelope, and for the production of mycothiol, which maintains the redox balance of the cell. Inositol is synthesized by conversion of glucose-6-phosphate to inositol-1-phosphate, followed by dephosphorylation by inositol monophosphate phosphatases (IMPases to form myo-inositol. To gain insight into how Mycobacterium tuberculosis synthesises inositol we carried out genetic analysis of the four IMPase homologues that are present in the Mycobacterium tuberculosis genome. Results Mutants lacking either impA (Rv1604 or suhB (Rv2701c were isolated in the absence of exogenous inositol, and no differences in levels of PIMs, LM, LAM or mycothiol were observed. Mutagenesis of cysQ (Rv2131c was initially unsuccessful, but was possible when a porin-like gene of Mycobacterium smegmatis was expressed, and also by gene switching in the merodiploid strain. In contrast, we could only obtain mutations in impC (Rv3137 when a second functional copy was provided in trans, even when exogenous inositol was provided. Experiments to obtain a mutant in the presence of a second copy of impC containing an active-site mutation, in the presence of porin-like gene of M. smegmatis, or in the absence of inositol 1-phosphate synthase activity, were also unsuccessful. We showed that all four genes are expressed, although at different levels, and levels of inositol phosphatase activity did not fall significantly in any of the mutants obtained. Conclusions We have shown that neither impA, suhB nor cysQ is solely responsible for inositol synthesis. In contrast, we show that impC is essential for mycobacterial growth under the conditions we used, and suggest it may be required in the early stages of mycothiol synthesis.

Parish Tanya

2010-02-01

276

Discovery of pyrazolopyridones as a novel class of noncovalent DprE1 inhibitor with potent anti-mycobacterial activity.  

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A novel pyrazolopyridone class of inhibitors was identified from whole cell screening against Mycobacterium tuberculosis (Mtb). The series exhibits excellent bactericidality in vitro, resulting in a 4 log reduction in colony forming units following compound exposure. The significant modulation of minimum inhibitory concentration (MIC) against a Mtb strain overexpressing the Rv3790 gene suggested the target of pyrazolopyridones to be decaprenylphosphoryl-?-D-ribose-2'-epimerase (DprE1). Genetic mapping of resistance mutation coupled with potent enzyme inhibition activity confirmed the molecular target. Detailed biochemical characterization revealed the series to be a noncovalent inhibitor of DprE1. Docking studies at the active site suggest that the series can be further diversified to improve the physicochemical properties without compromising the antimycobacterial activity. The pyrazolopyridone class of inhibitors offers an attractive non-nitro lead series targeting the essential and vulnerable DprE1 enzyme for the discovery of novel antimycobacterial agents to treat both drug susceptible and drug resistant strains of Mtb. PMID:24818517

Panda, Manoranjan; Ramachandran, Sreekanth; Ramachandran, Vasanthi; Shirude, Pravin S; Humnabadkar, Vaishali; Nagalapur, Kavitha; Sharma, Sreevalli; Kaur, Parvinder; Guptha, Supreeth; Narayan, Ashwini; Mahadevaswamy, Jyothi; Ambady, Anisha; Hegde, Naina; Rudrapatna, Suresh S; Hosagrahara, Vinayak P; Sambandamurthy, Vasan K; Raichurkar, Anandkumar

2014-06-12

277

Factors associated with tuberculosis infection, and with anti-mycobacterial immune responses, among five year olds BCG-immunised at birth in Entebbe, Uganda  

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Background BCG is used widely as the sole licensed vaccine against tuberculosis, but it has variable efficacy and the reasons for this are still unclear. No reliable biomarkers to predict future protection against, or acquisition of, TB infection following immunisation have been identified. Lessons from BCG could be valuable in the development of effective tuberculosis vaccines. Objectives Within the Entebbe Mother and Baby Study birth cohort in Uganda, infants received BCG at birth. We investigated factors associated with latent tuberculosis infection (LTBI) and with cytokine response to mycobacterial antigen at age five years. We also investigated whether cytokine responses at one year were associated with LTBI at five years of age. Methods Blood samples from age one and five years were stimulated using crude culture filtrates of Mycobacterium tuberculosis in a six-day whole blood assay. IFN-?, IL-5, IL-13 and IL-10 production was measured. LTBI at five years was determined using T-SPOT.TB® assay. Associations with LTBI at five years were assessed using multivariable logistic regression. Multiple linear regression with bootstrapping was used to determine factors associated with cytokine responses at age five years. Results LTBI prevalence was 9% at age five years. Only urban residence and history of TB contact/disease were positively associated with LTBI. BCG vaccine strain, LTBI, HIV infection, asymptomatic malaria, growth z-scores, childhood anthelminthic treatment and maternal BCG scar were associated with cytokine responses at age five. Cytokine responses at one year were not associated with acquisition of LTBI by five years of age. Conclusion Although multiple factors influenced anti-myocbacterial immune responses at age five, factors likely to be associated with exposure to infectious cases (history of household contact, and urban residence) dominated the risk of LTBI. PMID:25529292

Lule, Swaib Abubaker; Mawa, Patrice A.; Nkurunungi, Gyaviira; Nampijja, Margaret; Kizito, Dennison; Akello, Florence; Muhangi, Lawrence; Elliott, Alison M.; Webb, Emily L.

2015-01-01

278

The radiology of IRIS (immune reconstitution inflammatory syndrome) in patients with mycobacterial tuberculosis and HIV co-infection: appearances in 11 patients  

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Aim: To determine the radiological manifestations of IRIS (immune reconstitution inflammatory syndrome) in patients with HIV and mycobacterium tuberculosis co-infection, in the context of their demographic and clinical data. Materials and methods: The radiological imaging, demographic and clinical data of 11 patients diagnosed with IRIS associated with HIV and mycobacterial tuberculosis co-infection were studied retrospectively. Where available, follow-up imaging studies were also reviewed. Results: The most common radiological feature of IRIS was lymph node enlargement (73%), with central low attenuation centres, in keeping with necrosis, present in most of these cases (88%). Most commonly affected were intra-abdominal nodes (70%), followed by axillary (40%) and mediastinal lymph nodes (36%). Within the lung parenchyma, diffuse, bilateral pulmonary nodules were seen in 55% of cases. Unilateral small volume pleural effusions were seen in two cases with associated parenchymal changes seen in only one. Small volume ascites was seen in two cases. Thirty-six percent of cases presented with new or worsening abscesses despite treatment. In this context, image-guided radiological drainage proved a useful adjunct to the conventional medical therapy for IRIS. The most common clinical signs of IRIS included fever (64%), abdominal pain (36%) and cough (27%). Conclusion: We have described the radiological features that are characteristic in IRIS and the importance of putting these into context with the clinical and pathological findings as part of a multidisciplinary approach in making the diagnosis. The role of the radiologist is central in diagnosis, monitoring of disease progression and management of complications in patients with IRIS.

Rajeswaran, G. [Department of Radiology and Department of HIV/GU Medicine, Chelsea and Westminster Hospital, London (United Kingdom)]. E-mail: grajeswaran@hotmail.com; Becker, J.L. [Department of Radiology and Department of HIV/GU Medicine, Chelsea and Westminster Hospital, London (United Kingdom); Michailidis, C. [Department of Radiology and Department of HIV/GU Medicine, Chelsea and Westminster Hospital, London (United Kingdom); Pozniak, A.L. [Department of Radiology and Department of HIV/GU Medicine, Chelsea and Westminster Hospital, London (United Kingdom); Padley, S.P.G. [Department of Radiology and Department of HIV/GU Medicine, Chelsea and Westminster Hospital, London (United Kingdom)

2006-10-15

279

Mycobacterial interspersed repetitive unit typing and mutational profile for multidrug-resistant and extensively drug-resistant tuberculosis surveillance in Portugal: a 3-year period overview.  

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Multidrug tuberculosis (MDR-TB) and extensively drug-resistant tuberculosis (XDR-TB) cases constitute a serious health problem in Portugal, of which the majority of isolates belong to the Lisboa family and the Q1 cluster, highly related to the Lisboa family. Here we sought to investigate the molecular basis of resistant TB as well as to determine the prevalence of specific drug resistance mutations and their association with MDR-TB and/or XDR-TB. In total, 74 Mycobacterium tuberculosis clinical isolates collected in Lisbon Health Region were genotyped by 24-loci mycobacterial interspersed repetitive units-variable number of tandem repeats (MIRU-VNTR), and the mutational profile associated with first- and second-line drug resistance was studied. Seven new mutations were found, whilst the remaining 28 mutations had been previously associated with drug resistance. None of the mutations was specifically associated with MDR-TB. The mutational patterns observed among isolates belonging to Lisboa3 and Q1 clusters were also observed in isolates with unique MIRU-VNTR patterns but closely related to these strains. Such data suggest that the genotyping technique employed discriminates isolates with the same mutational profile. To establish the most adequate genotyping technique, the discriminatory power of three different MIRU-VNTR sets was analysed. The 15-loci MIRU-VNTR set showed adequate discriminatory power, comparable with the 24-loci set, allowing clustering of 60% and 86% of the MDR-TB and XDR-TB isolates, respectively, the majority of which belonged to the Lisboa3 and Q1 clusters. From an epidemiological standpoint, this study suggests combined mutational and genotyping analysis as a valuable tool for drug resistance surveillance. PMID:25270633

Silva, Carla; Perdigão, João; Jordão, Luísa; Portugal, Isabel

2014-12-01

280

The radiology of IRIS (immune reconstitution inflammatory syndrome) in patients with mycobacterial tuberculosis and HIV co-infection: appearances in 11 patients  

International Nuclear Information System (INIS)

Aim: To determine the radiological manifestations of IRIS (immune reconstitution inflammatory syndrome) in patients with HIV and mycobacterium tuberculosis co-infection, in the context of their demographic and clinical data. Materials and methods: The radiological imaging, demographic and clinical data of 11 patients diagnosed with IRIS associated with HIV and mycobacterial tuberculosis co-infection were studied retrospectively. Where available, follow-up imaging studies were also reviewed. Results: The most common radiological feature of IRIS was lymph node enlargement (73%), with central low attenuation centres, in keeping with necrosis, present in most of these cases (88%). Most commonly affected were intra-abdominal nodes (70%), followed by axillary (40%) and mediastinal lymph nodes (36%). Within the lung parenchyma, diffuse, bilateral pulmonary nodules were seen in 55% of cases. Unilateral small volume pleural effusions were seen in two cases with associated parenchymal changes seen in only one. Small volume ascites was seen in two cases. Thirty-six percent of cases presented with new or worsening abscesses despite treatment. In this context, image-guided radiological drainage proved a useful adjunct to the conventional medical therapy for IRIS. The most common clinical signs of IRIS included fever (64%), abdominal pain (36%) and cough (27%). Conclusion: We have described the radiological features that are characteristic in IRIS and the importance of putting thesin IRIS and the importance of putting these into context with the clinical and pathological findings as part of a multidisciplinary approach in making the diagnosis. The role of the radiologist is central in diagnosis, monitoring of disease progression and management of complications in patients with IRIS

281

Gene Cloning  

Science.gov (United States)

This lesson covers the utilization of gene cloning to isolate and copy a specific gene of interest. The transformation of bacteria with plasmids containing antibiotic resistance genes to make gene libraries and the selection of bacteria colonies that contain the specific gene of interest are described.

282

The contraceptive depot medroxyprogesterone acetate impairs mycobacterial control and inhibits cytokine secretion in mice infected with Mycobacterium tuberculosis.  

Science.gov (United States)

The contraceptive depot medroxyprogesterone acetate (DMPA), with progestin as the single active compound, possesses selective glucocorticoid activity and can alter the expression of glucocorticoid receptor-regulated genes. We therefore propose that pharmacological doses of DMPA used for endocrine therapy could have significant immune modulatory effects and impact on susceptibility to, as well as clinical manifestation and outcome of, infectious diseases. We investigated the effect of contraceptive doses of DMPA in two different murine Mycobacterium tuberculosis models. Multiplex bead array analysis revealed that DMPA altered serum cytokine levels of tumor necrosis factor alpha (TNF-?), granulocyte colony-stimulating factor (G-CSF), and interleukin 10 (IL-10) in C57BL/6 mice and gamma interferon (IFN-?) in BALB/c mice. DMPA also suppressed antigen-specific production of TNF-?, G-CSF, IL-10, and IL-6 and induced the production of IP-10 in C57BL/6 mice. In BALB/c mice, DMPA altered the antigen-specific secretion of IFN-?, IL-17, granulocyte-macrophage colony-stimulating factor (GM-CSF), IL-6, and monocyte chemotactic protein 1 (MCP-1). Furthermore, we show that C57BL/6 mice treated with doses of DMPA, which result in serum concentrations similar to those observed in contraceptive users, have a significantly higher bacterial load in their lungs. Our data show for the first time that DMPA impacts tuberculosis (TB) disease severity in a mouse model and that the effects of this contraceptive are not confined to infections of the genital tract. This could have major implications for the contraceptive policies not only in developing countries like South Africa but also worldwide. PMID:23381991

Kleynhans, Léanie; Du Plessis, Nelita; Allie, Nasiema; Jacobs, Muazzam; Kidd, Martin; van Helden, Paul D; Walzl, Gerhard; Ronacher, Katharina

2013-04-01

283

GeneXpert MTB/RIF Testing in the Management of Patients with Active Tuberculosis; A Real Life Experience from Saudi Arabia  

Science.gov (United States)

Background GeneXpert MTB/RIF is a real-time PCR assay with established diagnostic performance in pulmonary and extra-pulmonary forms of tuberculosis. The aim of this study was to assess the contribution of GeneXpert MTB/RIF assay to the management of patients with any form of active tuberculosis in a single large tertiary center in Saudi Arabia, with a special focus on the impact on time to start of antituberculous therapy compared with Ziehl-Neelsen (ZN) smears and mycobacterial cultures. Materials and Methods Clinical, radiological and laboratory records for all patients who were commenced on antituberculous therapy between March 2011 and February 2013 were retrospectively reviewed. Results A total of 140 patients were included, 38.6% of which had pulmonary tuberculosis. GeneXpert MTB/RIF was requested for only 39.2% of patients and was the only reason for starting antituberculous therapy for only 12.1%. The median time to a positive GeneXpert MTB/RIF result was 0 days (IQR 3) compared with 0 day (IQR 1) for smear microscopy (P > 0.999) and 22 days (IQR 21) for mycobacterial cultures (P < 0.001). No patients discontinued antituberculous therapy because of a negative GeneXpert MTB/RIF result. Conclusions In a setting wherein physicians are highly experienced in the diagnosis and treatment of tuberculosis, GeneXpert MTB/RIF was remarkably under-utilized and had only a limited impact on decisions related to starting or stopping antituberculous therapy. Cost-effectiveness and clinical utility of routine testing of all smear-negative clinical samples submitted for tuberculosis investigations by GeneXpert MTB/RIF warrant further study. PMID:24693467

Al-Otaibi, Mohammed F.; Al-Ateah, Souad M.; Al-Onazi, Fahad M.; Baig, Kamran; El-Khizzi, Noura A.; Albarrak, Ali M.

2014-01-01

284

Systems Biology-Based Identification of Mycobacterium tuberculosis Persistence Genes in Mouse Lungs  

Science.gov (United States)

ABSTRACT Identifying Mycobacterium tuberculosis persistence genes is important for developing novel drugs to shorten the duration of tuberculosis (TB) treatment. We developed computational algorithms that predict M. tuberculosis genes required for long-term survival in mouse lungs. As the input, we used high-throughput M. tuberculosis mutant library screen data, mycobacterial global transcriptional profiles in mice and macrophages, and functional interaction networks. We selected 57 unique, genetically defined mutants (18 previously tested and 39 untested) to assess the predictive power of this approach in the murine model of TB infection. We observed a 6-fold enrichment in the predicted set of M. tuberculosis genes required for persistence in mouse lungs relative to randomly selected mutant pools. Our results also allowed us to reclassify several genes as required for M. tuberculosis persistence in vivo. Finally, the new results implicated additional high-priority candidate genes for testing. Experimental validation of computational predictions demonstrates the power of this systems biology approach for elucidating M. tuberculosis persistence genes. PMID:24549847

Dutta, Noton K.; Bandyopadhyay, Nirmalya; Veeramani, Balaji; Lamichhane, Gyanu; Karakousis, Petros C.; Bader, Joel S.

2014-01-01

285

Identification of a repetitive sequence belonging to a PPE gene of Mycobacterium tuberculosis and its use in diagnosis of tuberculosis.  

Science.gov (United States)

A repetitive sequence specific to Mycobacterium tuberculosis was isolated from a lambda gt11 library of M. tuberculosis by DNA-DNA hybridization using genomic DNA of M. tuberculosis as probe followed by subtractive hybridization with a cocktail of other mycobacterial DNA. This led to identification of CD192, a 1291 bp fragment of M. tuberculosis containing repetitive sequences, which produced positive hybridization signals with M. tuberculosis DNA within 30 min. Nucleotide sequencing revealed the presence of several direct and inverted repeats within the 1291 bp fragment that belonged to a PPE family gene (Rv0355) of M. tuberculosis. The use of CD192 as a DNA probe for the identification of M. tuberculosis in culture and clinical samples was investigated. The 1291 bp sequence was present in M. tuberculosis, Mycobacterium bovis and M. bovis BCG, but was not present in many of the other mycobacterial strains tested, including M. tuberculosis H37Ra. More than 300 clinical isolates of M. tuberculosis were probed with CD192, and the presence of the 1291 bp sequence was observed in all the clinical strains, including those lacking IS6110. The sequence displayed RFLP among the clinical isolates. A PCR assay was developed which detected M. tuberculosis with 100% specificity from specimens of sputum, cerebrospinal fluid and pleural effusion from clinically diagnosed cases of tuberculosis. PMID:16849727

Srivastava, Ranjana; Kumar, D; Waskar, M N; Sharma, Meera; Katoch, V M; Srivastava, Brahm S

2006-08-01

286

Expression of the Mycobacterium bovis P36 gene in Mycobacterium smegmatis and the baculovirus/insect cell system  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: English Abstract in english In the present study we evaluated different systems for the expression of mycobacterial antigen P36 secreted by Mycobacterium bovis. P36 was detected by Western blot using a specific antiserum. The P36 gene was initially expressed in E. coli, under the control of the T7 promoter, but severe proteoly [...] sis prevented its purification. We then tried to express P36 in M. smegmatis and insect cells. For M. smegmatis, we used three different plasmid vectors differing in copy number and in the presence of a promoter for expression of heterologous proteins. P36 was detected in the cell extract and culture supernatant in both expression systems and was recognized by sera from M. bovis-infected cattle. To compare the expression level and compartmentalization, the MPB70 antigen was also expressed. The highest production was reached in insect cell supernatants. In conclusion, M. smegmatis and especially the baculovirus expression system are good choices for the production of proteins from pathogenic mycobacteria for the development of mycobacterial vaccines and diagnostic reagents.

F., Bigi; O., Taboga; M.I., Romano; A., Alito; J.C., Fisanotti; A.A., Cataldi.

1999-01-01

287

Expression of the Mycobacterium bovis P36 gene in Mycobacterium smegmatis and the baculovirus/insect cell system  

Directory of Open Access Journals (Sweden)

Full Text Available In the present study we evaluated different systems for the expression of mycobacterial antigen P36 secreted by Mycobacterium bovis. P36 was detected by Western blot using a specific antiserum. The P36 gene was initially expressed in E. coli, under the control of the T7 promoter, but severe proteolysis prevented its purification. We then tried to express P36 in M. smegmatis and insect cells. For M. smegmatis, we used three different plasmid vectors differing in copy number and in the presence of a promoter for expression of heterologous proteins. P36 was detected in the cell extract and culture supernatant in both expression systems and was recognized by sera from M. bovis-infected cattle. To compare the expression level and compartmentalization, the MPB70 antigen was also expressed. The highest production was reached in insect cell supernatants. In conclusion, M. smegmatis and especially the baculovirus expression system are good choices for the production of proteins from pathogenic mycobacteria for the development of mycobacterial vaccines and diagnostic reagents.

Bigi F.

1999-01-01

288

ClpR protein-like regulator specifically recognizes RecA protein-independent promoter motif and broadly regulates expression of DNA damage-inducible genes in mycobacteria.  

Science.gov (United States)

The RecA-dependent DNA damage response pathway (SOS response) appears to be the major DNA repair mechanism in most bacteria, but it has been suggested that a RecA-independent mechanism is responsible for controlling expression of most damage-inducible DNA repair genes in Mycobacterium tuberculosis. The specific reparative responses and molecular mediators involved in the DNA repair mechanism remain largely unclear in this pathogen and its related species. In this study, a mycobacterial ClpR-like regulator, corresponding to Rv2745c in M. tuberculosis and to Ms2694 in M. smegmatis mc(2)155, was found to interact with the promoter regions of multiple damage-inducible DNA repair genes. Specific binding of the ClpR-like factor to the conserved RecA-independent promoter RecA-NDp motif was then confirmed using in vitro electrophoretic mobility shift assays as well as in vivo chromatin immunoprecipitation experiments. The ClpR knock-out experiments, in combination with quantitative real time PCR assays, demonstrated that the expression of these RecA-independent genes were significantly down-regulated in the mutant strain of M. smegmatis in response to a DNA-damaging agent compared with the wild type strain. Furthermore, the ClpR-like factor was shown to contribute to mycobacterial genomic stability. These results enhance our understanding of the function of the ClpR regulator and the regulatory mechanism of RecA-independent DNA repair in mycobacteria. PMID:21771781

Wang, Yi; Huang, Yuanxia; Xue, Chaolun; He, Yang; He, Zheng-Guo

2011-09-01

289

ClpR Protein-like Regulator Specifically Recognizes RecA Protein-independent Promoter Motif and Broadly Regulates Expression of DNA Damage-inducible Genes in Mycobacteria*  

Science.gov (United States)

The RecA-dependent DNA damage response pathway (SOS response) appears to be the major DNA repair mechanism in most bacteria, but it has been suggested that a RecA-independent mechanism is responsible for controlling expression of most damage-inducible DNA repair genes in Mycobacterium tuberculosis. The specific reparative responses and molecular mediators involved in the DNA repair mechanism remain largely unclear in this pathogen and its related species. In this study, a mycobacterial ClpR-like regulator, corresponding to Rv2745c in M. tuberculosis and to Ms2694 in M. smegmatis mc2155, was found to interact with the promoter regions of multiple damage-inducible DNA repair genes. Specific binding of the ClpR-like factor to the conserved RecA-independent promoter RecA-NDp motif was then confirmed using in vitro electrophoretic mobility shift assays as well as in vivo chromatin immunoprecipitation experiments. The ClpR knock-out experiments, in combination with quantitative real time PCR assays, demonstrated that the expression of these RecA-independent genes were significantly down-regulated in the mutant strain of M. smegmatis in response to a DNA-damaging agent compared with the wild type strain. Furthermore, the ClpR-like factor was shown to contribute to mycobacterial genomic stability. These results enhance our understanding of the function of the ClpR regulator and the regulatory mechanism of RecA-independent DNA repair in mycobacteria. PMID:21771781

Wang, Yi; Huang, Yuanxia; Xue, Chaolun; He, Yang; He, Zheng-Guo

2011-01-01

290

Diagnosis of nontuberculous mycobacterial infections.  

Science.gov (United States)

The nontuberculous mycobacteria (NTM) are typically environmental organisms residing in soil and water. Although generally of low pathogenicity to humans, NTM can cause a wide array of clinical diseases; pulmonary disease is most frequent, followed by lymphadenitis in children, skin disease by M. marinum (particularly in fish tank fanciers), and other extrapulmonary or disseminated infections in severely immunocompromised patients. Of the >140 NTM species reported in the literature, 25 species have been strongly associated with NTM diseases; the remainder are environmental organisms rarely encountered in clinical samples. Correct species identification is very important because NTM species differ in their clinical relevance. Further, NTM differ strongly in their growth rate, temperature tolerance, and drug susceptibility. The diagnosis of NTM disease is complex and requires good communication between clinicians, radiologists, and microbiologists. Isolation of M. kansasii and (in northwestern Europe) M. malmoense from pulmonary specimens usually indicates disease, whereas Mycobacterium gordonae and, to a lesser extent, M. simiae or M. chelonae are typically contaminants rather than causative agents of true disease. Mycobacterium avium complex (MAC), M. xenopi, and M. abscessus form an intermediate category between these two extremes. This review covers the clinical and laboratory diagnosis of NTM diseases and particularities for the different disease types and patient populations. Because of limited sensitivity and specificity of symptoms, radiology, and direct microscopy of clinical samples, culture remains the gold standard. Yet culture is time consuming and demands the use of multiple media types and incubation temperatures to optimize the yield. Outside of reference centers, such elaborate culture algorithms are scarce. PMID:23460010

van Ingen, Jakko

2013-02-01

291

É possível uma vacina gênica auxiliar no controle da tuberculose? Could a DNA vaccine be useful in the control of tuberculosis?  

Directory of Open Access Journals (Sweden)

Full Text Available Vacinas de DNA, ainda em fase de experimentação e testes clínicos, podem se tornar uma importante ferramenta de combate a doenças infecciosas para as quais, até hoje, não existe prevenção segura e eficaz, como a tuberculose. Nos últimos anos vários estudos têm sido dedicados ao desenvolvimento de vacinas de DNA que codificam proteínas de micobactérias, entre as quais destacam-se as que codificam o antígeno 85 (Ag 85 e a proteína de choque térmico de 65 kDa (hsp65. Estes dois antígenos foram os mais estudados apresentando resultados bastante satisfatórios em ensaios pré-clínicos e com grande volume de dados registrados na literatura. Além de proteger contra infecção experimental por Mycobacterium tuberculosis virulenta, a vacina DNA-hsp65 também apresenta atividade terapêutica, ou seja, é capaz de curar os animais previamente infectados, inclusive aqueles com bacilos resistentes a múltiplas drogas. Esta vacina, hoje em avaliação clínica no Brasil também para o tratamento de câncer, é capaz de induzir a produção de citocinas de padrão Th1 tal como IFN- interferon-gama, associadas ao controle da doença. Além disso, a vacina de DNA-hsp65 é capaz de estimular clones de células CD8 citotóxicos e CD4 que podem ser caracterizados como células de memória sendo responsáveis por conferir imunidade duradoura contra a infecção. Quando utilizada na terapia da infecção, a vacina de DNA-hsp65 faz com que haja uma mudança no padrão de resposta imune, induzindo a secreção de citocinas de padrão Th1 criando um ambiente favorável à erradicação do bacilo. Os resultados demonstram ainda que a via de administração e a formulação na qual a vacina é administrada exerce fundamental influência no padrão e duração da resposta imune desencadeada. O conjunto de resultados hoje disponíveis mostra que uma vacina de DNA contra a tuberculose contribuirá de maneira significativa no controle desta doença.The DNA vaccines currently under pre-clinical and clinical development may prove to be important tools in combating infectious diseases, such as tuberculosis, for which no safe and effective form of prevention has yet been developed. In recent years, several studies have aimed to develop a DNA vaccine encoding mycobacterial proteins such as antigen 85 (Ag85 and the 65-kDa mycobacterial heat shock protein (hsp65. The latter is protective against virulent infection with Mycobacterium tuberculosis (including multidrug-resistant strains. The hsp65 DNA vaccine, currently under clinical evaluation in Brazil for cancer therapy, is able to induce the secretion of Th1 cytokines, such as gamma-interferon, associated with disease control. Furthermore, this vaccine stimulates cytotoxic CD8 and CD4 T-cell clones that can be characterized as memory cells, which are responsible for effective and long-lasting immunity against tuberculosis. When used as a therapeutic agent in inoculated mice, the hsp65 DNA vaccine promotes changes in the immunity profile, triggering the secretion of Th1 cytokines and establishing a favorable environment for the elimination of bacilli. The results also demonstrate that the route of administration, as well as the formulation in which the vaccine is administered, fundamentally influence the pattern and duration of the immune response induced. Taking all currently available data into account, we can conclude that a DNA vaccine against tuberculosis could contribute significantly to the control of the disease.

José Maciel Rodrigues Júnior

2004-08-01

292

É possível uma vacina gênica auxiliar no controle da tuberculose? / Could a DNA vaccine be useful in the control of tuberculosis?  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: Portuguese Abstract in portuguese Vacinas de DNA, ainda em fase de experimentação e testes clínicos, podem se tornar uma importante ferramenta de combate a doenças infecciosas para as quais, até hoje, não existe prevenção segura e eficaz, como a tuberculose. Nos últimos anos vários estudos têm sido dedicados ao desenvolvimento de va [...] cinas de DNA que codificam proteínas de micobactérias, entre as quais destacam-se as que codificam o antígeno 85 (Ag 85) e a proteína de choque térmico de 65 kDa (hsp65). Estes dois antígenos foram os mais estudados apresentando resultados bastante satisfatórios em ensaios pré-clínicos e com grande volume de dados registrados na literatura. Além de proteger contra infecção experimental por Mycobacterium tuberculosis virulenta, a vacina DNA-hsp65 também apresenta atividade terapêutica, ou seja, é capaz de curar os animais previamente infectados, inclusive aqueles com bacilos resistentes a múltiplas drogas. Esta vacina, hoje em avaliação clínica no Brasil também para o tratamento de câncer, é capaz de induzir a produção de citocinas de padrão Th1 tal como IFN- interferon-gama, associadas ao controle da doença. Além disso, a vacina de DNA-hsp65 é capaz de estimular clones de células CD8 citotóxicos e CD4 que podem ser caracterizados como células de memória sendo responsáveis por conferir imunidade duradoura contra a infecção. Quando utilizada na terapia da infecção, a vacina de DNA-hsp65 faz com que haja uma mudança no padrão de resposta imune, induzindo a secreção de citocinas de padrão Th1 criando um ambiente favorável à erradicação do bacilo. Os resultados demonstram ainda que a via de administração e a formulação na qual a vacina é administrada exerce fundamental influência no padrão e duração da resposta imune desencadeada. O conjunto de resultados hoje disponíveis mostra que uma vacina de DNA contra a tuberculose contribuirá de maneira significativa no controle desta doença. Abstract in english The DNA vaccines currently under pre-clinical and clinical development may prove to be important tools in combating infectious diseases, such as tuberculosis, for which no safe and effective form of prevention has yet been developed. In recent years, several studies have aimed to develop a DNA vacci [...] ne encoding mycobacterial proteins such as antigen 85 (Ag85) and the 65-kDa mycobacterial heat shock protein (hsp65). The latter is protective against virulent infection with Mycobacterium tuberculosis (including multidrug-resistant strains). The hsp65 DNA vaccine, currently under clinical evaluation in Brazil for cancer therapy, is able to induce the secretion of Th1 cytokines, such as gamma-interferon, associated with disease control. Furthermore, this vaccine stimulates cytotoxic CD8 and CD4 T-cell clones that can be characterized as memory cells, which are responsible for effective and long-lasting immunity against tuberculosis. When used as a therapeutic agent in inoculated mice, the hsp65 DNA vaccine promotes changes in the immunity profile, triggering the secretion of Th1 cytokines and establishing a favorable environment for the elimination of bacilli. The results also demonstrate that the route of administration, as well as the formulation in which the vaccine is administered, fundamentally influence the pattern and duration of the immune response induced. Taking all currently available data into account, we can conclude that a DNA vaccine against tuberculosis could contribute significantly to the control of the disease.

José Maciel, Rodrigues Júnior; Karla de Melo, Lima; Arlete Aparecida Martins Coelho, Castelo; Vânia Luiza Deperon Bonato, Martins; Sandra Aparecida dos, Santos; Lucia Helena, Faccioli; Célio Lopes, Silva.

2004-08-01

293

[Activation of immunoproteasome subunit genes transcription in mice monocytes infected with different strains of mycobacteria].  

Science.gov (United States)

Immunoproteasomal processing of mycobacterial antigens is necessary to control the infection and to protect the organism from development of active form of tuberculosis. Here we investigate the activation of immunoproteasome subunit genes transcription in peritoneal monocytes of C57Bl/6 mice infected with vaccine M. bovis BCG and virulent strain M. tuberculosis H37Rv. The level of transcription of LMP2, LMP7, MECL1 subunits didn't increase for one and two days after a single infection. Two rounds of infection with BCG strain M. bovis led to enhancement of the only LMP7 subunit gene transcription. However after subsequent infection of monocytes with vaccine followed by virulent strain infection the dramatic rise of all immunoproteasomal subunit genes transcription was observed. Activation of transcription of the gene coding the PA28alpha subunit of regulatory complex PA28 was observed only after a single infection of monocytes with strain M. bovis BCG. Thus, vaccination with strain M. bovis BCG promotes effective activation of immunoproteasomal genes in case of subsequent contact with virulent strain M. tuberculosis H37Rv. PMID:23808166

Timofeev, A V; Kuz'menko, Iu V; Zharkov, I I; Starodubova, E S; Karpov, V L

2013-01-01

294

Gene Regions  

Science.gov (United States)

This animation shows the three gene coding regions. This is the fourth of a series of seven animations that detail the process of crop genetic engineering. To begin at the beginning, see Overview of Crop Genetic Engineering. (To return to the animation previous to this, go to Gene Cloning. To go to the next animation, go to Gene Modification.)

295

Patients with inhibitory and neutralizing auto-antibodies to interferon-? resemble the sporadic adult-onset phenotype of Mendelian Susceptibility to Mycobacterial Disease (MSMD) lacking Bacille Calmette-Guerin (BCG)-induced diseases.  

Science.gov (United States)

To recognize patients with inhibitory and neutralizing auto-antibodies to interferon-? (AutoAbs-IFN-?) presenting with the sporadic phenotype of Mendelian Susceptibility to Mycobacterial Disease (MSMD) mainly characterized by recurrent intracellular mycobacterium or/and salmonella infections, we comprehensively investigated IL12/23-IFN-? signaling, candidate genetic sequencings or/and protein expressions of IL12RB1, IFNRG1, IL12p40, IFNRG2, STAT1, IKKA, NEMO, CYBB and IRF8 in four patients. Their serum was further titrated to detect AutoAbs-IFN-?, for which the biological activity was assessed in Jurkat T cells. The patients mainly presented with recurrent non-tuberculous mycobacterium osteomyelitis and lymphadenopathy (Mycobacterium abscessus, chelonae and avium intracellular complex), and salmonella sepsis (S. enterica serogroup B, C2 and D). Additionally, Penicillium marneffei, varicella-zoster virus, and herpes simplex virus infections occurred. Inhibitory and neutralizing IFN-? downstream signaling was elucidated in Jurkat cell lines as decreased MHC class I and phosphorylated STAT1 expression. Together with 24 patients from the PubMed search, the majority of the AutoAbs-IFN-? patients were Asian (25/28). The most common involvement was lymph nodes (in 22/28), lungs (19/28) and bones (12/28). Mycobacterium avium complex (in 14) and chelonae (7) were the most common pathogens from 40 isolations. In contrast to those with the mild form of MSMD phenotype, AutoAbs-IFN-? patients, in the absence of BCG-induced diseases, had a more persistent course and poor response to IFN-? treatment. In conclusion, AutoAbs-IFN-? patients may have a sporadic adult-onset MSMD phenotype in Asian regions endemic for mycobacterial infections. PMID:23083630

Lee, Wen-I; Huang, Jing-Long; Wu, Ting-Shu; Lee, Ming-Hsun; Chen, I-Jung; Yu, Kuang-Hiu; Liu, Chien-Ying; Yang, Chih-Hsun; Hsieh, Meng-Ying; Lin, Yi-Ling; Shih, Ying-Fan; Jaing, Tang-Her; Huang, Shih-Chiang; Kuo, Tseng-Tong; Ku, Cheng-Lung

2013-05-01

296

Gene Modifications  

Science.gov (United States)

This animation shows how a gene is constructed to eventually produce a protein in a Bt corn plant. This is the fifth of a series of seven animations that detail the process of crop genetic engineering. To begin at the beginning, see Overview of Crop Genetic Engineering. (To return to the animation previous to this, go to Gene Regions. To go to the next animation, go to Gene Gun.)

297

Cancer genes.  

OpenAIRE

Cancer is a genetic disease; tumor cells differ from their normal progenitors by genetic alterations that affect growth-regulatory genes. There exist 2 classes of such cancer genes: the oncogenes, which function as positive growth regulators, and the tumor suppressor genes, which function as negative growth regulators. Oncogenes are widely conserved among diverse forms of life and are active in transmitting growth signals from the cell periphery to the cell nucleus. These signaling functions ...

Vogt, P. K.

1993-01-01

298

Genetic polymorphisms in TNF genes and tuberculosis in North Indians  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background Pulmonary tuberculosis, the most common clinical form of mycobacterial diseases, is a granulomatous disease of the lungs caused by Mycobaterium tuberculosis. A number of genes have been identified in studies of diverse origins to be important in tuberculosis. Of these, both tumor necrosis factor ? (TNF-? and lymphotoxin ? (LT-? play important immunoregulatory roles. Methods To investigate the association of TNF polymorphisms with tuberculosis in the Asian Indians, we genotyped five potentially functional promoter polymorphisms in the TNFA gene and a LTA_NcoI polymorphism (+252 position of the LTA gene in a clinically well-defined cohort of North-Indian patients with tuberculosis (N = 185 and their regional controls (N = 155. Serum TNF-? (sTNF-? levels were measured and correlated with genotypes and haplotypes. Results The comparison of the allele frequencies for the various loci investigated revealed no significant differences between the tuberculosis patients and controls. Also, when the patients were sub-grouped into minimal, moderately advanced and far advanced disease on the basis of chest radiographs, TST and the presence/absence of cavitary lesions, none of the polymorphisms showed a significant association with any of the patient sub-groups. Although a significant difference was observed in the serum TNF-? levels in the patients and the controls, none of the investigated polymorphisms were found to affect the sTNF-? levels. Interestingly, it was observed that patients with minimal severity were associated with lower log sTNF-? levels when compared to the patients with moderately advanced and far advanced severity. However, none of these differences were found to be statistically significant. Furthermore, when haplotypes were analyzed, no significant difference was observed. Conclusions Thus, our findings exclude the TNF genes as major risk factor for tuberculosis in the North Indians.

Ghosh Balaram

2010-06-01

299

Trichoderma genes  

Energy Technology Data Exchange (ETDEWEB)

Described herein are novel gene sequences isolated from Trichoderma reesei. Two genes encoding proteins comprising a cellulose binding domain, one encoding an arabionfuranosidase and one encoding an acetylxylanesterase are described. The sequences, CIP1 and CIP2, contain a cellulose binding domain. These proteins are especially useful in the textile and detergent industry and in pulp and paper industry.

Foreman, Pamela (Los Altos, CA); Goedegebuur, Frits (Vlaardingen, NL); Van Solingen, Pieter (Naaldwijk, NL); Ward, Michael (San Francisco, CA)

2012-06-19

300

Gene Gun  

Science.gov (United States)

How the gene gun works to transform cells with new DNA. This is thesixth of a series of seven animations that detail the process of cropgenetic engineering. To begin at the beginning, see Overview of Crop Genetic Engineering. (To return to the animation previous to this, go to Gene Modification. To go to the next animation, go to Backcross Breeding.)

301

Perspective on sequence evolution of microsatellite locus (CCGn in Rv0050 gene from Mycobacterium tuberculosis  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background The mycobacterial genome is inclined to polymerase slippage and a high mutation rate in microsatellite regions due to high GC content and absence of a mismatch repair system. However, the exact molecular mechanisms underlying microsatellite variation have not been fully elucidated. Here, we investigated mutation events in the hyper-variable trinucleotide microsatellite locus MML0050 located in the Rv0050 gene of W-Beijing and non-W-Beijing Mycobacterium tuberculosis strains in order to gain insight into the genomic structure and activity of repeated regions. Results Size analysis indicated the presence of five alleles that differed in length by three base pairs. Moreover, nucleotide gains occurred more frequently than loses in this trinucleotide microsatellite. Mutation frequency was not completely related with the total length, though the relative frequency in the longest allele was remarkably higher than that in the shortest. Sequence analysis was able to detect seven alleles and revealed that point mutations enhanced the level of locus variation. Introduction of an interruptive motif correlated with the total allele length and genetic lineage, rather than the length of the longest stretch of perfect repeats. Finally, the level of locus variation was drastically different between the two genetic lineages. Conclusion The Rv0050 locus encodes the bifunctional penicillin-binding protein ponA1 and is essential to mycobacterial survival. Our investigations of this particularly dynamic genomic region provide insights into the overall mode of microsatellite evolution. Specifically, replication slippage was implicated in the mutational process of this microsatellite and a sequence-based genetic analysis was necessary to determine that point mutation events acted to maintain microsatellite size integrity while providing genomic diversity.

Jin Ruiliang

2011-08-01

302

Clinical and laboratory features of Mycobacterium porcinum.  

Science.gov (United States)

Recent molecular studies have shown Mycobacterium porcinum, recovered from cases of lymphadenitis in swine, to have complete 16S rDNA sequence identity and >70% DNA-DNA homology with human isolates within the M. fortuitum third biovariant complex. We identified 67 clinical and two environmental isolates of the M. fortuitum third biovariant sorbitol-negative group, of which 48 (70%) had the same PCR restriction enzyme analysis (PRA) profile as the hsp65 gene of M. porcinum (ATCC 33776(T)) and were studied in more detail. Most U.S. patient isolates were from Texas (44%), Florida (19%), or other southern coastal states (15%). Clinical infections included wound infections (62%), central catheter infections and/or bacteremia (16%), and possible pneumonitis (18%). Sequencing of the 16S rRNA gene (1,463 bp) showed 100% identity with M. porcinum ATCC 33776(T). Sequencing of 441 bp of the hsp65 gene showed four sequevars that differed by 2 to 3 bp from the porcine strains. Clinical isolates were positive for arylsulfatase activity at 3 days, nitrate, iron uptake, D-mannitol, i-myo-inositol, and catalase at 68 degrees C. They were negative for L-rhamnose and D-glucitol (sorbitol). Clinical isolates were susceptible to ciprofloxacin, sulfamethoxazole, and linezolid and susceptible or intermediate to cefoxitin, clarithromycin, imipenem, and amikacin. M. porcinum ATCC 33776(T) gave similar results except for being nitrate negative. These studies showed almost complete phenotypic and molecular identity between clinical isolates of the M. fortuitum third biovariant D-sorbitol-negative group and porcine strains of M. porcinum and confirmed that they belong to the same species. Identification of M. porcinum presently requires hsp65 gene PRA or 16S rRNA or hsp65 gene sequencing. PMID:15583300

Wallace, Richard J; Brown-Elliott, Barbara A; Wilson, Rebecca W; Mann, Linda; Hall, Leslie; Zhang, Yansheng; Jost, Kenneth C; Brown, June M; Kabani, Amin; Schinsky, Mark F; Steigerwalt, Arnold G; Crist, Christopher J; Roberts, Glenn D; Blacklock, Zeta; Tsukamura, Michio; Silcox, Vella; Turenne, Christine

2004-12-01

303

Clinical and Laboratory Features of Mycobacterium porcinum†  

Science.gov (United States)

Recent molecular studies have shown Mycobacterium porcinum, recovered from cases of lymphadenitis in swine, to have complete 16S rDNA sequence identity and >70% DNA-DNA homology with human isolates within the M. fortuitum third biovariant complex. We identified 67 clinical and two environmental isolates of the M. fortuitum third biovariant sorbitol-negative group, of which 48 (70%) had the same PCR restriction enzyme analysis (PRA) profile as the hsp65 gene of M. porcinum (ATCC 33776T) and were studied in more detail. Most U.S. patient isolates were from Texas (44%), Florida (19%), or other southern coastal states (15%). Clinical infections included wound infections (62%), central catheter infections and/or bacteremia (16%), and possible pneumonitis (18%). Sequencing of the 16S rRNA gene (1,463 bp) showed 100% identity with M. porcinum ATCC 33776T. Sequencing of 441 bp of the hsp65 gene showed four sequevars that differed by 2 to 3 bp from the porcine strains. Clinical isolates were positive for arylsulfatase activity at 3 days, nitrate, iron uptake, d-mannitol, i-myo-inositol, and catalase at 68°C. They were negative for l-rhamnose and d-glucitol (sorbitol). Clinical isolates were susceptible to ciprofloxacin, sulfamethoxazole, and linezolid and susceptible or intermediate to cefoxitin, clarithromycin, imipenem, and amikacin. M. porcinum ATCC 33776T gave similar results except for being nitrate negative. These studies showed almost complete phenotypic and molecular identity between clinical isolates of the M. fortuitum third biovariant d-sorbitol-negative group and porcine strains of M. porcinum and confirmed that they belong to the same species. Identification of M. porcinum presently requires hsp65 gene PRA or 16S rRNA or hsp65 gene sequencing. PMID:15583300

Wallace, Richard J.; Brown-Elliott, Barbara A.; Wilson, Rebecca W.; Mann, Linda; Hall, Leslie; Zhang, Yansheng; Jost, Kenneth C.; Brown, June M.; Kabani, Amin; Schinsky, Mark F.; Steigerwalt, Arnold G.; Crist, Christopher J.; Roberts, Glenn D.; Blacklock, Zeta; Tsukamura, Michio; Silcox, Vella; Turenne, Christine

2004-01-01

304

Characterization of the first report of Mycobacterium timonense infecting an HIV patient in an Ecuadorian hospital.  

Science.gov (United States)

Mycobacterium timonense is a non-tuberculous mycobacteria (NTM) described in southern France in 2009, and to our knowledge, not reported again as a human pathogen in indexed literature. The aim of this work was to characterize the first clinical isolate of M. timonense in Ecuador. Time of growth, biochemical tests, thin layer growth test, PCR-RFLP analysis of the hsp65 gene and MALDI-TOF spectra analysis were not able to identify the species. The species identification was achieved through sequencing of rrs, hsp65 and rpoB genes. The results highlight the necessity to set up a sequencing method to identify emerging NTM in Ecuadorian clinical facilities. PMID:24813256

Zurita, J; Ortega-Paredes, D; Mora, M; Espinel, N; Parra, H; Febres, L; Zurita-Salinas, C

2014-12-01

305

Enfermedades micobacterianas diseminadas en pacientes con VIH/SIDA. Evaluación de los hemocultivos por método rápido Disseminated mycobacterial infections in patients with HIV/AIDS. Evaluation of blood cultures  

Directory of Open Access Journals (Sweden)

Full Text Available Mil cuarenta hemocultivos correspondientes a 451 enfermos uruguayos con SIDA y diagnóstico clínico de micobacteriosis diseminada fueron evaluados entre 1999 y 2003. Las muestras fueron procesadas en el Centro de Referencia Nacional para Micobacterias (Montevideo, Uruguay, utilizando el sistema de hemocultivos automatizado para micobacterias MB - BacT (BioMérieux. Se detectaron 45 muestras positivas (4,3% correspondientes a 26 enfermos (promedio 2,3 muestras por paciente. En 10/26 casos se identificó M. avium complex (MAC y en 13/26 el germen aislado fue M. tuberculosis. El tiempo medio de incubación fue de 12,4 días (intervalo 6-19 días para MAC y de 22,6 días (intervalo 7-35 días para M. tuberculosis. El hemocultivo ha demostrado ser la mejor muestra para la confirmación bacteriológica de las enfermedades micobacterianas diseminadas cuando se estudian por lo menos 2 muestras por paciente. La frecuencia de aislamientos de M. tuberculosis y MAC aislados en pacientes con SIDA en Uruguay, corresponde a la de un país con una moderada prevalencia de tuberculosis.One thousand-forty blood cultures corresponding to 451 Uruguayan patients with AIDS and clinic diagnosis of disseminated mycobacterial infection were evaluated between 1999 and 2003. Samples were processed in the NationalReferenceCenter for Mycobacteria (Montevideo, Uruguay, using the automated blood culture system for mycobacteria MB -BacT (BioMérieux. Forty-five positive samples were detected (4.3% corresponding to 26 patients with AIDS (average 2.3 samples per patient. In 10/26 patients M. avium complex (MAC was identified and in 13/26 the isolated germ was M. tuberculosis. The average time of incubation was of 12.4 days (range 6-19 days for MAC and of 22.6 days (range 7-35 days for M. tuberculosis. Blood culture has demonstrated to be the best sample for the bacteriological confirmation of the disseminated mycobacterial infections when at least 2 samples by patient are studied. The frequency of isolates of M. tuberculosis and MAC in AIDS patients is according with a moderate prevalence of tuberculosis in Uruguay.

C. Coitinho

2005-12-01

306

Activity-based metabolomic profiling of enzymatic function: Identification of Rv1248c as a mycobacterial 2-hydroxy-3-oxoadipate synthase  

OpenAIRE

Activity based metabolomic profiling (ABMP) allows unbiased discovery of enzymatic activities encoded by genes of unknown function. ABMP applies liquid chromatography-mass spectrometry (LC-MS) to analyze the impact of a recombinant enzyme on the homologous cellular extract as a physiologic library of potential substrates and products. The Mycobacterium tuberculosis protein Rv1248c was incompletely characterized as a thiamine diphosphate-dependent ?-ketoglutarate decarboxylase. Here, recombin...

Carvalho, Luiz Pedro S.; Zhao, Hong; Dickinson, Caitlyn E.; Arango, Nancy M.; Lima, Christopher D.; Fischer, Steven M.; Ouerfelli, Ouathek; Nathan, Carl; Rhee, Kyu Y.

2010-01-01

307

Gene Cloning  

Science.gov (United States)

This interactive activity adapted from the University of Nebraska's Library of Crop Technologies details the steps involved in producing clones of genes that can then be used to transform the characteristics of an organism.

2009-09-08

308

Gene therapy.  

OpenAIRE

In recent years, there have been a number of technological breakthroughs that have allowed for clinical trials in gene therapy to be initiated. In combination with the genome initiative, the potential for new therapeutics is limitless. Although an enormous amount of information has been obtained in a relatively short period of time, gene therapy is not yet ready for wide-scale practice. Some of the successes and obstacles that remain are summarized in this report.

Mota Biosca, Anna

1992-01-01

309

Key Hub and Bottleneck Genes Differentiate the Macrophage Response to Virulent and Attenuated Mycobacterium bovis  

Science.gov (United States)

Mycobacterium bovis is an intracellular pathogen that causes tuberculosis in cattle. Following infection, the pathogen resides and persists inside host macrophages by subverting host immune responses via a diverse range of mechanisms. Here, a high-density bovine microarray platform was used to examine the bovine monocyte-derived macrophage transcriptome response to M. bovis infection relative to infection with the attenuated vaccine strain, M. bovis Bacille Calmette–Guérin. Differentially expressed genes were identified (adjusted P-value ?0.01) and interaction networks generated across an infection time course of 2, 6, and 24?h. The largest number of biological interactions was observed in the 24-h network, which exhibited scale-free network properties. The 24-h network featured a small number of key hub and bottleneck gene nodes, including IKBKE, MYC, NFKB1, and EGR1 that differentiated the macrophage response to virulent and attenuated M. bovis strains, possibly via the modulation of host cell death mechanisms. These hub and bottleneck genes represent possible targets for immuno-modulation of host macrophages by virulent mycobacterial species that enable their survival within a hostile environment. PMID:25324841

Killick, Kate E.; Magee, David A.; Park, Stephen D. E.; Taraktsoglou, Maria; Browne, John A.; Conlon, Kevin M.; Nalpas, Nicolas C.; Gormley, Eamonn; Gordon, Stephen V.; MacHugh, David E.; Hokamp, Karsten

2014-01-01

310

Simple and Rational Approach to the Identification of Mycobacterium tuberculosis, Mycobacterium avium Complex Species, and Other Commonly Isolated Mycobacteria  

OpenAIRE

A novel PCR-restriction fragment length polymorphism analysis of the hsp65 gene was developed. The restriction patterns for Mycobacterium tuberculosis and Mycobacterium avium complex (MAC) species were designed to be highly distinct, and the overall number of restriction patterns was designed to be limited. Four hundred specimens (17 reference strains and 383 clinical isolates) were tested, of which 98 were M. tuberculosis and 132 were MAC species. The assay was virtually 100% sensitive and s...

Wong, Derek A.; Yip, Peter C. W.; Cheung, Danny T. L.; Kam, Kai Man

2001-01-01

311

Molecular characterization of Mycobacterium avium subsp. paratuberculosis using multi-locus short sequence repeat (MLSSR) and mycobacterial interspersed repetitive units-variable number tandem repeat (MIRU-VNTR) typing methods.  

Science.gov (United States)

The aim of this study was to characterize 38 bovine strains of Mycobacterium avium subsp. paratuberculosis isolated in Ireland using 11 multi locus short sequence repeat (MLSSR) loci and 8 mycobacterial interspersed repetitive units-variable number of tandem repeat (MIRU-VNTR) loci. The discriminatory power of these two typing methods alone and combined was evaluated, as was the epidemiology of the isolates and the genotypes obtained. Using the MIRU-VNTR typing method (8 loci analysed), only 3 subtypes were detected with a discrimination index (DI) of 0.54, but one MIRU-VNTR type has not been identified in other studies. In contrast the MLSSR method (using 11 loci) differentiated the 38 MAP isolates into 18 types with DI of 0.92. Among these 18 types 6 have not been recorded previously. The combination of the 2 methods (MIRU-VNTR and MLSSR) produced 22 distinct genotypes giving a maximal DI of 0.94. According to the allelic diversity, some markers are more polymorphic than others and must be applied in priority for the differentiation of MAP bovine isolates. This is the first report of genotyping data for MAP on the island of Ireland and will be very useful for analysing its epidemiology, transmission and virulence. PMID:21269784

Douarre, P E; Cashman, W; Buckley, J; Coffey, A; O'Mahony, J

2011-05-01

312

Designer Genes.  

Science.gov (United States)

Genetic technologies may soon help fill some of the most important needs of humanity from food to energy to health care. The research of major designer genes companies and reasons why the initial mad rush for biotechnology has slowed are reviewed. (SR)

Miller, Judith; Miller, Mark

1983-01-01

313

Evaluating gene × gene and gene × smoking interaction in rheumatoid arthritis using candidate genes in GAW15  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract We examined the potential gene × gene interactions and gene × smoking interactions in rheumatoid arthritis (RA using the candidate gene data sets provided by Genetic Analysis Workshop 15 Problem 2. The multifactor dimensionality reduction (MDR method was used to test gene × gene interactions among candidate genes. The case-only sample was used to test gene × smoking interactions. The best predictive model was the single-locus model with single-nucleotide polymorphism (SNP rs2476601 in gene PTPN22. However, no clear gene × gene interaction was identified. Substantial departure from multiplicativity was observed between smoking and SNPs in genes CTLA4, PADI4, MIF, and SNPs on chromosome 5 and one haplotype of PTPN22. The strongest evidence of association was identified between the PTPN22 gene and RA status, which was consistently detected in single SNP association, gene × gene interaction and gene × smoking interaction analyses.

Mei Ling

2007-12-01

314

Characteristics of inpatients with nontuberculous mycobacterial infections in a highly complex hospital in Colombia / Caracterización de pacientes hospitalizados con infecciones causadas por micobacterias no tuberculosas, en un hospital de alta complejidad en Colombia  

Scientific Electronic Library Online (English)

Full Text Available SciELO Colombia | Language: English Abstract in spanish Antecedentes: Las infecciones por micobacterias no tuberculosas (MNT) se describen en los últimos años con mayor frecuencia, especialmente en pacientes con inmunosupresión y en pacientes tratados por procedimientos estéticos. Las MNT incluyen especies del género Mycobacterium , diferentes del comple [...] jo Mycobacterium tuberculosis y Mycobacterium leprae . Objetivo: Describir las características demográficas y clínicas de pacientes hospitalizados con infecciones por MNT. Metodología: Estudio descriptivo retrospectivo. Resultados: De 187 pacientes con infección por micobacterias documentadas por cultivo, 17 (9,1%) tuvieron infección por MNT. Edad promedio de 38,4 ± 19,2 años. El 58,82% fueron hombres. Las principales comorbilidades fueron VIH/sida (41,17%), diabetes mellitus (23,53%), enfermedad renal crónica (17,64%), terapia inmunosupresora (17,64%) y neoplasias (17,64%). En los coinfectados con VIH el recuento de CD4 fue Abstract in english Background: Nontuberculous mycobacteria (NTM) infections has been described more frequently in recent years, especially in immunosuppression conditions and after cosmetic surgical procedures. The NTM include species of the genus Mycobacterium , other than Mycobacterium tuberculosis complex and Mycob [...] acterium leprae. Objective: To describe the demographic and clinical characteristics of Colombian in-patientswith NTM infections. Methodology: A retrospective descriptive study. Results: In 187 patients with culture- confirmed mycobacterial infection, 17 (9,1%) had NTM.The mean age was 38,4 ± 19,2 and 58,82% were men. Major comorbidities were: HIV/AIDS(41,1%), diabetes mellitus (23,5%), chronic renal disease (17,6%), immunosuppressive therapy(17,6%) and neoplasms (17,6%). In patients co-infected with HIV, CD4 count was

Franco Eduardo, Montúfar; Camilo A., Madrid; María C., Montufar; Carolina, Aguilar; Carolina, Saldarriaga; Miguel A., Mesa; Alicia, Quiroga; Carlos E., Builes; John J., Zuleta; Olga L., Molina.

2014-12-30

315

High Genetic Diversity among Mycobacterium avium subsp. paratuberculosis Strains from German Cattle Herds Shown by Combination of IS900 Restriction Fragment Length Polymorphism Analysis and Mycobacterial Interspersed Repetitive Unit-Variable-Number Tandem-Repeat Typing?  

Science.gov (United States)

Mycobacterium avium subsp. paratuberculosis is the etiologic agent of Johne's disease and is endemic to the national cattle herds of many countries. Because of the very low level of genetic heterogeneity of this organism, it is difficult to select a workable procedure for strain differentiation at a resolution sufficient to investigate epidemiological links between herds or different ruminant species and the suggested zoonotic potential of M. avium subsp. paratuberculosis for Crohn's disease. Analysis of restriction fragment length polymorphisms (RFLPs) based on the insertion element IS900 (IS900 RFLP) with four restriction enzymes and 10 markers of specific mycobacterial interspersed repetitive units (MIRUs) and variable-number tandem repeats (VNTRs) was applied to 71 bovine M. avium subsp. paratuberculosis isolates originating from 14 herds from different regions in Germany. Among these isolates, all of which belonged to the M. avium subsp. paratuberculosis type II group, 17 genotypes were detected by IS900 RFLP and consisted of a combination of seven BstEII, eight PstI, nine PvuII, and four BamHI restriction patterns. Novel RFLP types were found. The diversity of the M. avium subsp. paratuberculosis isolates inside the herds was different depending on the frequency of animal purchase. The results of typing by IS900 RFLP and MIRU-VNTR analyses were not associated. Fifteen MIRU-VNTR patterns were identified with a discriminatory index of 0.905. The most common BstEII-based IS900 RFLP type, type C1 (72%), was subdivided into 14 types by MIRU-VNTR analysis. A combination of fingerprinting and PCR-based techniques resulted in 24 M. avium subsp. paratuberculosis genotypes and achieved a discriminatory index of 0.997. By using only BstEII and PstI digestion together with typing by MIRU-VNTR analysis, a discriminatory index of 0.993 was achieved. This is high enough to support epidemiological studies on a national as well as a global scale. PMID:18174306

Möbius, Petra; Luyven, Gabriele; Hotzel, Helmut; Köhler, Heike

2008-01-01

316

Determination of Genotypic Diversity of Mycobacterium avium Subspecies from Human and Animal Origins by Mycobacterial Interspersed Repetitive-Unit-Variable-Number Tandem-Repeat and IS1311 Restriction Fragment Length Polymorphism Typing Methods ? †  

Science.gov (United States)

Members of the Mycobacterium avium complex (MAC) are ubiquitous bacteria that can be found in water, food, and other environmental samples and are considered opportunistic pathogens for numerous animal species, mainly birds and pigs, as well as for humans. We have recently demonstrated the usefulness of a PCR-based mycobacterial interspersed repetitive-unit-variable-number tandem-repeat (MIRU-VNTR) typing for the molecular characterization of M. avium subsp. paratuberculosis and M. avium strains exclusively isolated from AIDS patients. In the present study we extended our analysis, based on eight MIRU-VNTR markers, to a strain collection comprehensively comprising the other M. avium subspecies, including M. avium subsp. avium, M. avium subsp. hominissuis, and M. avium subsp. silvaticum, isolated from numerous animal species, HIV-positive and HIV-negative humans, and environmental sources. All strains were fully typeable, with the discriminatory index being 0.885, which is almost equal to that obtained by IS1311 restriction fragment length polymorphism (RFLP) typing as a reference. In contrast to IS1311 RFLP typing, MIRU-VNTR typing was able to further discriminate M. avium subsp. avium strains. MIRU-VNTR alleles strongly associated with or specific for M. avium subspecies were detected in several markers. Moreover, the MIRU-VNTR typing-based results were consistent with a scenario of the independent evolution of M. avium subsp. avium/M. avium subsp. silvaticum and M. avium subsp. paratuberculosis from M. avium subsp. hominissuis, previously proposed on the basis of multilocus sequence analysis. MIRU-VNTR typing therefore appears to be a convenient typing method capable of distinguishing the three main subspecies and strains of the complex and providing new epidemiological knowledge on MAC. PMID:20107094

Radomski, Nicolas; Thibault, Virginie C.; Karoui, Claudine; de Cruz, Krystel; Cochard, Thierry; Gutiérrez, Cristina; Supply, Philip; Biet, Frank; Boschiroli, María Laura

2010-01-01

317

High genetic diversity among Mycobacterium avium subsp. paratuberculosis strains from German cattle herds shown by combination of IS900 restriction fragment length polymorphism analysis and mycobacterial interspersed repetitive unit-variable-number tandem-repeat typing.  

Science.gov (United States)

Mycobacterium avium subsp. paratuberculosis is the etiologic agent of Johne's disease and is endemic to the national cattle herds of many countries. Because of the very low level of genetic heterogeneity of this organism, it is difficult to select a workable procedure for strain differentiation at a resolution sufficient to investigate epidemiological links between herds or different ruminant species and the suggested zoonotic potential of M. avium subsp. paratuberculosis for Crohn's disease. Analysis of restriction fragment length polymorphisms (RFLPs) based on the insertion element IS900 (IS900 RFLP) with four restriction enzymes and 10 markers of specific mycobacterial interspersed repetitive units (MIRUs) and variable-number tandem repeats (VNTRs) was applied to 71 bovine M. avium subsp. paratuberculosis isolates originating from 14 herds from different regions in Germany. Among these isolates, all of which belonged to the M. avium subsp. paratuberculosis type II group, 17 genotypes were detected by IS900 RFLP and consisted of a combination of seven BstEII, eight PstI, nine PvuII, and four BamHI restriction patterns. Novel RFLP types were found. The diversity of the M. avium subsp. paratuberculosis isolates inside the herds was different depending on the frequency of animal purchase. The results of typing by IS900 RFLP and MIRU-VNTR analyses were not associated. Fifteen MIRU-VNTR patterns were identified with a discriminatory index of 0.905. The most common BstEII-based IS900 RFLP type, type C1 (72%), was subdivided into 14 types by MIRU-VNTR analysis. A combination of fingerprinting and PCR-based techniques resulted in 24 M. avium subsp. paratuberculosis genotypes and achieved a discriminatory index of 0.997. By using only BstEII and PstI digestion together with typing by MIRU-VNTR analysis, a discriminatory index of 0.993 was achieved. This is high enough to support epidemiological studies on a national as well as a global scale. PMID:18174306

Möbius, Petra; Luyven, Gabriele; Hotzel, Helmut; Köhler, Heike

2008-03-01

318

Determination of genotypic diversity of Mycobacterium avium subspecies from human and animal origins by mycobacterial interspersed repetitive-unit-variable-number tandem-repeat and IS1311 restriction fragment length polymorphism typing methods.  

Science.gov (United States)

Members of the Mycobacterium avium complex (MAC) are ubiquitous bacteria that can be found in water, food, and other environmental samples and are considered opportunistic pathogens for numerous animal species, mainly birds and pigs, as well as for humans. We have recently demonstrated the usefulness of a PCR-based mycobacterial interspersed repetitive-unit-variable-number tandem-repeat (MIRU-VNTR) typing for the molecular characterization of M. avium subsp. paratuberculosis and M. avium strains exclusively isolated from AIDS patients. In the present study we extended our analysis, based on eight MIRU-VNTR markers, to a strain collection comprehensively comprising the other M. avium subspecies, including M. avium subsp. avium, M. avium subsp. hominissuis, and M. avium subsp. silvaticum, isolated from numerous animal species, HIV-positive and HIV-negative humans, and environmental sources. All strains were fully typeable, with the discriminatory index being 0.885, which is almost equal to that obtained by IS1311 restriction fragment length polymorphism (RFLP) typing as a reference. In contrast to IS1311 RFLP typing, MIRU-VNTR typing was able to further discriminate M. avium subsp. avium strains. MIRU-VNTR alleles strongly associated with or specific for M. avium subspecies were detected in several markers. Moreover, the MIRU-VNTR typing-based results were consistent with a scenario of the independent evolution of M. avium subsp. avium/M. avium subsp. silvaticum and M. avium subsp. paratuberculosis from M. avium subsp. hominissuis, previously proposed on the basis of multilocus sequence analysis. MIRU-VNTR typing therefore appears to be a convenient typing method capable of distinguishing the three main subspecies and strains of the complex and providing new epidemiological knowledge on MAC. PMID:20107094

Radomski, Nicolas; Thibault, Virginie C; Karoui, Claudine; de Cruz, Krystel; Cochard, Thierry; Gutiérrez, Cristina; Supply, Philip; Biet, Frank; Boschiroli, María Laura

2010-04-01

319

Use of sequence microdivergence in mycobacterial ortholog to analyze contributions of the water-activating loop histidine of Escherichia coli uracil-DNA glycosylase in reactant binding and catalysis  

International Nuclear Information System (INIS)

Uracil-DNA glycosylase (Ung), a DNA repair enzyme, pioneers uracil excision repair pathway. Structural determinations and mutational analyses of the Ung class of proteins have greatly facilitated our understanding of the mechanism of uracil excision from DNA. More recently, a hybrid quantum-mechanical/molecular mechanical analysis revealed that while the histidine (H67 in EcoUng) of the GQDPYH motif (? loop) in the active site pocket is important in positioning the reactants, it makes an unfavorable energetic contribution (penalty) in achieving the transition state intermediate. Mutational analysis of this histidine is unavailable from any of the Ung class of proteins. A complication in demonstrating negative role of a residue, especially when located within the active site pocket, is that the mutants with enhanced activity are rarely obtained. Interestingly, unlike the most Ung proteins, the H67 equivalent in the ? loop in mycobacterial Ung is represented by P67. Exploiting this natural diversity to maintain structural integrity of the active site, we transplanted an H67P mutation in EcoUng. Uracil inhibition assays and binding of a proteinaceous inhibitor, Ugi (a transition state substrate mimic), with the mutant (H67P) revealed that its active site pocket was not perturbed. The catalytic efficiency (Vmax/Km) of the mutant was similar to that of the wild type Ung. However, the mutant showed increased Km and Vmax. Togethesub>m and Vmax. Together with the data from a double mutation H67P/G68T, these observations provide the first biochemical evidence for the proposed diverse roles of H67 in catalysis by Ung

320

Evaluating gene × gene and gene × smoking interaction in rheumatoid arthritis using candidate genes in GAW15  

OpenAIRE

Abstract We examined the potential gene × gene interactions and gene × smoking interactions in rheumatoid arthritis (RA) using the candidate gene data sets provided by Genetic Analysis Workshop 15 Problem 2. The multifactor dimensionality reduction (MDR) method was used to test gene × gene interactions among candidate genes. The case-only sample was used to test gene × smoking interactions. The best predictive model was the single-locus model with single-nucleotide polymorphism ...

Mei Ling; Li Xiaohui; Yang Kai; Cui Jinrui; Fang Belle; Guo Xiuqing; Rotter Jerome I

2007-01-01

321

Rapid detection of viable M. tuberculosis directly from sputum by reverse transcriptase PCR targeting 85B gene.  

Science.gov (United States)

This study was carried out to standardize reverse transcriptase PCR (RT-PCR) targeting 85B gene for the rapid detection of viable Mycobacterium tuberculosis (M tuberculosis) from sputum specimens. 100 sputum samples from clinically suspected tuberculosis patients were included in the study. The sputum samples were collected in sterile diethyl pyrocarbonate (DEPC) treated containers and transported in ice to the laboratory within 2 hours to prevent degradation of RNA. The following microbiological analysis was performed on the sputum specimens: Ziehl Neelsen staining, Mycobacterial culture by BACTEC TB 460 reader and RT-PCR targeting 85B gene. Out of the 100 sputum samples, 40 sputum samples were Ziehl Neelsen stain positive, 58 sputum samples were culture and RT-PCR positive and 42 were culture and RT-PCR were negative. Among direct smear positive specimens, 3 specimens did not grow in culture and was RT-PCR negative indicating the non-viability of the acid-fast bacilli seen in the direct smear. The results of RT-PCR and culture against control group and clinically diagnosed tuberculosis patients were statistically significant by Chi square test (P < 0.001). RT-PCR targeting 85B gene of M tuberculosis standardized in our laboratory for is a rapid, reliable and sensitive technique to detect the viable M. tuberculosis directly from sputum specimens. PMID:23785865

Gayathri, Ramasubban; Therese, Kulandai Lily; Sridhar, R; Kannan, Durai; Madhavan, Hajib; Meenakshi, N

2011-06-01

322

Overexpression of the Salmonella KdpF membrane peptide modulates expression of kdp genes and intramacrophage growth.  

Science.gov (United States)

Membrane peptides appear as an emerging class of regulatory molecules in bacteria, which can interact with membrane proteins, including transporters and sensor kinases. The KdpF peptide, which is cotranscribed with kdpABC genes and regulated by the KdpDE two-component system, is supposed to stabilize the KdpABC potassium transporter complex but may also exhibit unsuspected regulatory function(s). The mycobacterial KdpF can interact with the KdpD histidine kinase, and kdpF overexpression has been shown to reduce intramacrophage replication of Mycobacterium bovis BCG. In this study, we investigated whether KdpF displays similar behavior in another intracellular pathogen, Salmonella enterica serovar Typhimurium. We show that Salmonella KdpF can interact with KdpD in a bacterial two-hybrid assay. We have constructed a Salmonella strain overexpressing kdpF, and we have investigated expression of the kdp regulon, as well as intramacrophage survival. We show that kdpF overexpression reduces expression of kdpA and kdpD genes under potassium limitation. Moreover, kdpF overexpression increases intramacrophage multiplication of S. Typhimurium. Hence, our results indicate that KdpF can play a regulatory role in S. Typhimurium, modulating kdp gene expression and intramacrophage survival, but in a way that differs from the one reported for M. bovis BCG. PMID:25197761

Gannoun-Zaki, Laila; Belon, Claudine; Dupont, Christian; Hilbert, Friederike; Kremer, Laurent; Blanc-Potard, Anne-Béatrice

2014-10-01

323

Gene expression and gene therapy imaging  

International Nuclear Information System (INIS)

The fast growing field of molecular imaging has achieved major advances in imaging gene expression, an important element of gene therapy. Gene expression imaging is based on specific probes or contrast agents that allow either direct or indirect spatio-temporal evaluation of gene expression. Direct evaluation is possible with, for example, contrast agents that bind directly to a specific target (e.g., receptor). Indirect evaluation may be achieved by using specific substrate probes for a target enzyme. The use of marker genes, also called reporter genes, is an essential element of MI approaches for gene expression in gene therapy. The marker gene may not have a therapeutic role itself, but by coupling the marker gene to a therapeutic gene, expression of the marker gene reports on the expression of the therapeutic gene. Nuclear medicine and optical approaches are highly sensitive (detection of probes in the picomolar range), whereas MRI and ultrasound imaging are less sensitive and require amplification techniques and/or accumulation of contrast agents in enlarged contrast particles. Recently developed MI techniques are particularly relevant for gene therapy. Amongst these are the possibility to track gene therapy vectors such as stem cells, and the techniques that allow spatiotemporal control of gene expression by non-invasive heating (with MRI guided focused ultrasound) and the use of temperature sensitive promoters. (orig.)

324

Gene Switches  

Science.gov (United States)

In this activity, learners explore how genetic switches function and the role of genetic switches in the process of evolution. To make these concepts less abstract and more understandable, learners first view a series of video clips and animations from the HHMI DVD (or online) "Evolution: Constant Change and Common Threads." Then, learners construct a model of a gene switch using craft materials or FridgiGears (magnetic gears). This activity can be done as a demonstration, a student inquiry activity, or a combination of the two.

Colvard, Mary

2010-01-01

325

Novel diagnostic algorithm for identification of mycobacteria using genus-specific amplification of the 16S-23S rRNA gene spacer and restriction endonucleases.  

Science.gov (United States)

A novel genus-specific PCR for mycobacteria with simple identification to the species level by restriction fragment length polymorphism (RFLP) was established using the 16S-23S ribosomal RNA gene (rDNA) spacer as a target. Panspecificity of primers was demonstrated on the genus level by testing 811 bacterial strains (122 species in 37 genera from 286 reference strains and 525 clinical isolates). All mycobacterial isolates (678 strains among 48 defined species and 5 indeterminate taxons) were amplified by the new primers. Among nonmycobacterial isolates, only Gordonia terrae was amplified. The RFLP scheme devised involves estimation of variable PCR product sizes together with HaeIII and CfoI restriction analysis. It yielded 58 HaeIII patterns, of which 49 (84%) were unique on the species level. Hence, HaeIII digestion together with CfoI results was sufficient for correct identification of 39 of 54 mycobacterial taxons and one of three or four of seven RFLP genotypes found in Mycobacterium intracellulare and Mycobacterium kansasii, respectively. Following a clearly laid out diagnostic algorithm, the remaining unidentified organisms fell into five clusters of closely related species (i.e., the Mycobacterium avium complex or Mycobacterium chelonae-Mycobacterium abscessus) that were successfully separated using additional enzymes (TaqI, MspI, DdeI, or AvaII). Thus, next to slowly growing mycobacteria, all rapidly growing species studied, including M. abscessus, M. chelonae, Mycobacterium farcinogenes, Mycobacterium fortuitum, Mycobacterium peregrinum, and Mycobacterium senegalense (with a very high 16S rDNA sequence similarity) were correctly identified. A high intraspecies sequence stability and the good discriminative power of patterns indicate that this method is very suitable for rapid and cost-effective identification of a wide variety of mycobacterial species without the need for sequencing. Phylogenetically, spacer sequence data stand in good agreement with 16S rDNA sequencing results, as was shown by including strains with unsettled taxonomy. Since this approach recognized significant subspecific genotypes while identification of a broad spectrum of mycobacteria rested on identification of one specific RFLP pattern within a species, this method can be used by both reference (or research) and routine laboratories. PMID:10699003

Roth, A; Reischl, U; Streubel, A; Naumann, L; Kroppenstedt, R M; Habicht, M; Fischer, M; Mauch, H

2000-03-01

326

Amoebal coculture of "Mycobacterium massiliense" sp. nov. from the sputum of a patient with hemoptoic pneumonia.  

Science.gov (United States)

A nonphotochromogenic, rapidly growing Mycobacterium strain was isolated in pure culture from the sputum and the bronchoalveolar fluid of a patient with hemoptoic pneumonia by using axenic media and an amoebal coculture system. Both isolates grew in less than 7 days at 24 to 37 degrees C with an optimal growth temperature of 30 degrees C. The isolates exhibited biochemical and antimicrobial susceptibility profiles overlapping those of Mycobacterium abscessus, Mycobacterium chelonae, and Mycobacterium immunogenum, indicating that they belonged to M. chelonae-M. abscessus group. They differed from M. abscessus in beta-galactosidase, beta-N-acetyl-beta-glucosaminidase, and beta-glucuronidase activities and by the lack of nitrate reductase and indole production activities, as well as in their in vitro susceptibilities to minocycline and doxycycline. These isolates and M. abscessus differed from M. chelonae and M. immunogenum by exhibiting gelatinase and tryptophane desaminase activities. Their 16S rRNA genes had complete sequence identity with that of M. abscessus and >99.6% similarity with those of M. chelonae and M. immunogenum. Further molecular investigations showed that partial hsp65 and sodA gene sequences differed from that of M. abscessus by five and three positions over 441 bp, respectively. Partial rpoB and recA gene sequence analyses showed 96 and 98% similarities with M. abscessus, respectively. Similarly, 16S-23S rRNA internal transcribed spacer sequence of the isolates differed from that of M. abscessus by a A-->G substitution at position 60 and a C insertion at position 102. Phenotypic and genotypic features of these two isolates indicated that they were representative of a new mycobacterial species within the M. chelonae-M. abscessus group. Phylogenetic analysis suggested that these isolates were perhaps recently derived from M. abscessus. We propose the name of "Mycobacterium massiliense" for this new species. The type strain has been deposited in the Collection Institut Pasteur as CIP 108297(T) and in Culture Collection of the University of Goteborg, Goteborg, Sweden, as CCUG 48898(T). PMID:15583272

Adékambi, Toïdi; Reynaud-Gaubert, Martine; Greub, Gilbert; Gevaudan, Marie-José; La Scola, Bernard; Raoult, Didier; Drancourt, Michel

2004-12-01

327

STRESS AND ATHEROSCLEROSIS: MAY HSP60 BE THE MOLECULAR LINK?  

Directory of Open Access Journals (Sweden)

Full Text Available In last decades, incidence of cardiovascular diseases is increased. Among them, atherosclerosis is one of the most commons. It is a disorder of in- flammation and innate immunity following lipid accumulation. From a bio- logic perspective, the process of adhesion and transmigration of immune cells (monocytes and macrophages across the endothelium is a crucial step for atherogenesis and mature plaque rupture. Moreover, there is a relationship between inflammation, infection, autoimmunity and athero- sclerosis. Inflammation has received increasing attention in recent years as a cause of atherosclerosis and cardiovascular diseases. Autoimmune diseases are characterized by enhanced atherosclerosis. Humoral immune responses to mycobacterial Hsp65, as well as to human Hsp60 and oxLDL, have been established in a number of human autoimmune diseases and are considered to be significantly associated also with atherosclerosis.

Luana Lipari

2009-01-01

328

CD4+, CD8+, CD3+ cell counts and CD4+/CD8+ ratio among patients with mycobacterial diseases (leprosy, tuberculosis), HIV infections, and normal healthy adults: a comparative analysis of studies in different regions of India.  

Science.gov (United States)

In this study, we estimated the CD4+, CD8+, CD3+ cell counts and the CD4/CD8 ratio among normal healthy controls (adults and children), leprosy patients (without any complications and during reactional states), TB patients (with and without HIV), and HIV-positive patients (early infection and full-blown AIDS) and correlated the changes with disease progression. In our study, it was observed that among adults, CD4+ cell counts ranged from 518-1098, CD8+ from 312-952, whereas CD4/CD8 ratio from 0.75-2.30. Among children, both CD4+ and CD8+ cells were more and the CD4/CD8 ratio varied from 0.91-3.17. With regard to leprosy patients, we observed that CD4+ and CD8+ cell counts were lower among PB (pauci-bacillary) and MB (multi-bacillary) patients. CD4/CD8 ratio was 0.99 ± 0.28 among PB patients while the ratio was lower, 0.78 ± 0.20, among MB patients. CD4+ cell counts were raised during RR (reversal reactions) and ENL (erythema nodosum leprosum) among the PB and MB patients whereas the CD8+ cell counts were lower among PB and MB patients. CD4/CD8 ratio doubled during reactional episodes of RR and ENL. Among the HIV-negative tuberculosis (TB) patients, both the CD4+ and CD8+ cell counts were found to be less and the CD4/CD8 ratio varied between 0.53-1.75. Among the HIV-positive TB patients and HIV-positive patients, both the CD4+ and CD8+ cells were very less and ratio drops significantly. In the initial stages of infection, as CD4+ counts drop, an increase in the CD8+ cell counts was observed and the ratio declines. In full-blown cases, CD4+ cell counts were very low, 3-4 to 54 cells, CD8+ cells from 12-211 and the ratio drops too low. This study is the first of its kind in this region of the country and assumes importance since no other study has reported the values of CD4+ and CD8+ T-lymphocyte counts among patients with mycobacterial diseases (leprosy and TB), HIV infections along with normal healthy individuals of the region, and correlation with clinical presentations of patients. PMID:25350657

Hussain, Tahziba; Kulshreshtha, K K; Yadav, V S; Katoch, Kiran

2015-01-01

329

Deep stromal mycobacterial keratitis: viable bacteria after six months of treatment: case report and literature review / Ceratite estromal profunda por micobactéria: bactéria viável após seis meses de tratamento: relato de caso e revisão da literatura  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: English Abstract in portuguese O objetivo do caso é descrever a presença de micobactérias viáveis em pacientes com ceratite, 6 meses após tratamento intensivo. A identificação de espécies, foi efetuada usando método PRA (polymerase chain reaction seguida pela restriction endonuclease analysis). Clonalidade foi avaliada pelos méto [...] dos RAPD (randomly amplified polymorphic DNA) e ERIC-PCR (enterobacterial repetitive intergenic consensus - polymerase chain reaction). Paciente refere trauma com corpo estranho metálico há 3 semanas. A cultura da córnea revelou Mycobacterium abscessus. Após 6 meses de tratamento tópico e sistêmico, paciente apresentava-se sem inflamação, sendo considerado clinicamente curado. Realizou-se então, uma ceratoplastia penetrante com intuitos ópticos. A cultura da córnea transplantada revelou micobactérias de mesma origem clonal. O achado mais interessante neste relato, foi a positividade da cultura da córnea transplantada após 6 meses de intenso tratamento específico. Ao nosso conhecimento, esse é o primeiro caso relatado na literatura mostrando essa possibilidade em tratamento de ceratites por micobactérias. Assim, os pacientes com ceratite por Mycobacterium abscessus podem apresentar bactérias viáveis após longo tempo de tratamento específico e precisam ser seguidos cuidadosamente por um longo período de tempo. Abstract in english To report the presence of viable mycobacteria in a patient with keratitis treated for 6 months. Species identification was performed using the PRA method (polymerase chain reaction followed by restriction endonuclease analysis). Clonality was evaluated with RAPD (randomly amplified polymorphic DNA) [...] and ERIC-PCR (enterobacterial repetitive intergenic consensus - polymerase chain reaction) methods. The patient reported trauma due to a metallic foreign body 3 weeks prior to presentation. Initial corneal scraping cultures revealed Mycobacterium abscessus. After 6 months of topical and systemic treatment the patient presented with no active inflammation and was considered clinically cured. An optic penetrating keratoplasty was performed. Culture of the excised cornea revealed Mycobacterium abscessus. Both isolates had the same clonal origin. The most interesting finding of this case report was the positive culture of the excised cornea after 6 months of intensive specific topical therapy. To our knowledge, this is the first report in the literature showing this possibility in the treatment of Mycobacterial keratitis. Thus, Mycobacterium abscessus may present viable bacteria after long-term treatment and should be followed carefully for a long period of time after tapering the medication.

Filipe Accioly de, Gusmão; Lênio, Alvarenga; Luciene, Barbosa; Jorge, Sampaio; Sylvia Cardoso, Leão; Ana Luisa, Hofling-Lima; Denise de, Freitas.

2005-08-01

330

Deep stromal mycobacterial keratitis: viable bacteria after six months of treatment: case report and literature review Ceratite estromal profunda por micobactéria: bactéria viável após seis meses de tratamento: relato de caso e revisão da literatura  

Directory of Open Access Journals (Sweden)

Full Text Available To report the presence of viable mycobacteria in a patient with keratitis treated for 6 months. Species identification was performed using the PRA method (polymerase chain reaction followed by restriction endonuclease analysis. Clonality was evaluated with RAPD (randomly amplified polymorphic DNA and ERIC-PCR (enterobacterial repetitive intergenic consensus - polymerase chain reaction methods. The patient reported trauma due to a metallic foreign body 3 weeks prior to presentation. Initial corneal scraping cultures revealed Mycobacterium abscessus. After 6 months of topical and systemic treatment the patient presented with no active inflammation and was considered clinically cured. An optic penetrating keratoplasty was performed. Culture of the excised cornea revealed Mycobacterium abscessus. Both isolates had the same clonal origin. The most interesting finding of this case report was the positive culture of the excised cornea after 6 months of intensive specific topical therapy. To our knowledge, this is the first report in the literature showing this possibility in the treatment of Mycobacterial keratitis. Thus, Mycobacterium abscessus may present viable bacteria after long-term treatment and should be followed carefully for a long period of time after tapering the medication.O objetivo do caso é descrever a presença de micobactérias viáveis em pacientes com ceratite, 6 meses após tratamento intensivo. A identificação de espécies, foi efetuada usando método PRA (polymerase chain reaction seguida pela restriction endonuclease analysis. Clonalidade foi avaliada pelos métodos RAPD (randomly amplified polymorphic DNA e ERIC-PCR (enterobacterial repetitive intergenic consensus - polymerase chain reaction. Paciente refere trauma com corpo estranho metálico há 3 semanas. A cultura da córnea revelou Mycobacterium abscessus. Após 6 meses de tratamento tópico e sistêmico, paciente apresentava-se sem inflamação, sendo considerado clinicamente curado. Realizou-se então, uma ceratoplastia penetrante com intuitos ópticos. A cultura da córnea transplantada revelou micobactérias de mesma origem clonal. O achado mais interessante neste relato, foi a positividade da cultura da córnea transplantada após 6 meses de intenso tratamento específico. Ao nosso conhecimento, esse é o primeiro caso relatado na literatura mostrando essa possibilidade em tratamento de ceratites por micobactérias. Assim, os pacientes com ceratite por Mycobacterium abscessus podem apresentar bactérias viáveis após longo tempo de tratamento específico e precisam ser seguidos cuidadosamente por um longo período de tempo.

Filipe Accioly de Gusmão

2005-08-01

331

Biochemical and immunological characterization of a cpn60.1 knockout mutant of Mycobacterium bovis BCG.  

Science.gov (United States)

Pathogenic mycobacteria possess two homologous chaperones encoded by cpn60.1 and cpn60.2. Cpn60.2 is essential for survival, providing the basic chaperone function, while Cpn60.1 is not. In the present study, we show that inactivation of the Mycobacterium bovis BCG cpn60.1 (Mb3451c) gene does not significantly affect bacterial growth in 7H9 broth, but that this knockout mutant (?cpn60.1) forms smaller colonies on solid 7H11 medium than the parental and complemented strains. When growing on Sauton medium, the ?cpn60.1 mutant exhibits a thinner surface pellicle and is associated with higher culture filtrate protein content and, coincidentally, with less protein in its outermost cell envelope in comparison with the parental and complemented strains. Interestingly, in this culture condition, the ?cpn60.1 mutant is devoid of phthiocerol dimycocerosates, and its mycolates are two carbon atoms longer than those of the wild-type, a phenotype that is fully reversed by complementation. In addition, ?cpn60.1 bacteria are more sensitive to stress induced by H(2)O(2) but not by SDS, high temperature or acidic pH. Taken together, these data indicate that the cell wall of the ?cpn60.1 mutant is impaired. Analysis by 2D gel electrophoresis and MS reveals the upregulation of a few proteins such as FadA2 and isocitrate lyase in the cell extract of the mutant, whereas more profound differences are found in the composition of the mycobacterial culture filtrate, e.g. the well-known Hsp65 chaperonin Cpn60.2 is particularly abundant and increases about 200-fold in the filtrate of the ?cpn60.1 mutant. In mice, the ?cpn60.1 mutant is less persistent in lungs and, to a lesser extent, in spleen, but it induces a comparable mycobacteria-specific gamma interferon production and protection against Mycobacterium tuberculosis H37Rv challenge as do the parental and complemented BCG strains. Thus, by inactivating the cpn60.1 gene in M. bovis BCG we show that Cpn60.1 is necessary for the integrity of the bacterial cell wall, is involved in resistance to H(2)O(2)-induced stress but is not essential for its vaccine potential. PMID:21127129

Wang, Xiao-Ming; Lu, Changlong; Soetaert, Karine; S'Heeren, Catherine; Peirs, Priska; Lanéelle, Marie-Antoinette; Lefèvre, Philippe; Bifani, Pablo; Content, Jean; Daffé, Mamadou; Huygen, Kris; De Bruyn, Jacqueline; Wattiez, Ruddy

2011-04-01

332

Identifying Gene Interaction Enrichment for Gene Expression Data  

OpenAIRE

Gene set analysis allows the inclusion of knowledge from established gene sets, such as gene pathways, and potentially improves the power of detecting differentially expressed genes. However, conventional methods of gene set analysis focus on gene marginal effects in a gene set, and ignore gene interactions which may contribute to complex human diseases. In this study, we propose a method of gene interaction enrichment analysis, which incorporates knowledge of predefined gene sets (e.g. gene ...

Zhang, Jigang; Li, Jian; Deng, Hong-wen

2009-01-01

333

Imaging gene expression in gene therapy  

International Nuclear Information System (INIS)

Full text. Gene therapy can be used to introduce new genes, or to supplement the function of indigenous genes. At the present time, however, there is non-invasive test to demonstrate efficacy of the gene transfer and expression processes. It has been postulated that scintigraphic imaging can offer unique information on both the site at which the transferred gene is expressed, and the degree of expression, both of which are critical issue for safety and clinical efficacy. Many current studies are based on 'suicide gene therapy' of cancer. Cells modified to express these genes commit metabolic suicide in the presence of an enzyme encoded by the transferred gene and a specifically-convertible pro drug. Pro drug metabolism can lead to selective metabolic trapping, required for scintigraphy. Herpes simplex virus type-1 thymidine kinase (H S V-1 t k+) has been use for 'suicide' in vivo tumor gene therapy. It has been proposed that radiolabelled nucleosides can be used as radiopharmaceuticals to detect H S V-1 t k+ gene expression where the H S V-1 t k+ gene serves a reporter or therapeutic function. Animal gene therapy models have been studied using purine-([18 F]F H P G; [18 F]-A C V), and pyrimidine- ([123/131 I]I V R F U; [124/131I]) antiviral nucleosides. Principles of gene therapy and gene therapy imaging will be reviewed and experimental data for [123/131rimental data for [123/131I]I V R F U imaging with the H S V-1 t k+ reporter gene will be presented

334

Gene doping: gene delivery for olympic victory.  

Science.gov (United States)

With one recently recommended gene therapy in Europe and a number of other gene therapy treatments now proving effective in clinical trials it is feasible that the same technologies will soon be adopted in the world of sport by unscrupulous athletes and their trainers in so called 'gene doping'. In this article an overview of the successful gene therapy clinical trials is provided and the potential targets for gene doping are highlighted. Depending on whether a doping gene product is secreted from the engineered cells or is retained locally to, or inside engineered cells will, to some extent, determine the likelihood of detection. It is clear that effective gene delivery technologies now exist and it is important that detection and prevention plans are in place. PMID:23082866

Gould, David

2013-08-01

335

Gene conversion in steroid 21-hydroxylase genes.  

Science.gov (United States)

The steroid 21-hydroxylase gene, CYP21B, encodes cytochrome P450c21, which mediates 21-hydroxylation. The gene is located about 30 kb downstream from pseudogene CYP21A. The CYP21A gene is homologous to the CYP21B gene but contains some mutations, including a C----T change which leads a termination codon, TAG, in the eighth exon. We found the same change in a mutant CYP21B gene isolated from a patient with 21-hydroxylase deficiency. Furthermore, a reciprocal change--i.e., a T----C change in the eighth exon of the CYP21A gene--was observed in the Japanese population and was associated with the two HLA haplotypes, HLA-B44-DRw13 and HLA-Bw46-DRw8. These changes may be considered the result of gene conversion-like events. PMID:1971153

Urabe, K; Kimura, A; Harada, F; Iwanaga, T; Sasazuki, T

1990-06-01

336

Plasmid instability when the hsp60 gene promoter is used to express the protective non-toxic fragment B of the diphtheria toxin in recombinant BCG  

Directory of Open Access Journals (Sweden)

Full Text Available The genetic modification of the live attenuated Mycobacterium bovis BCG to deliver a protective Corynebacterium diphtheriae antigen in vivo could be a safer and less costly alternative to the new and more expensive DTP vaccines available today, in particular to third world-countries. The stability of expression of heterologous antigens in BCG, however, is a major challenge to the use of live recombinant bacteria in vaccine development and appears to be dependent to a certain extent, on a genetic compatibility between the expression cassette within the plasmid construct and the mycobacterium host. In the quest for the best recombinant BCG transformant to express the dtb gene of C. diphtheriae we generated two new rBCG strains by transforming the Moreau substrain of BCG with the mycobacterial expression vectors pUS973 and pUS977, each one carrying a different promoter to drive the expression of the target antigen. After transformation recombinant BCG clones were selected on Middlebrook 7H10 kanamycin Agar plates, expanded in Middlebrook 7H9 kanamycin Broth and analyzed by agarose gel electrophoresis and immunoblotting. rBCGs transformed with the construct carrying the weak PAN promoter from M. paratuberculosis stably expressed the dtb gene. Conversely, rBCGs transformed with the construct carrying the strong mycobacterium hsp60 promoter were unstable and consequently unfit for the expression of the C. diphtheriae gene.

Dilzamar V. Nascimento

2013-04-01

337

Nontuberculous Mycobacterial Lung Disease in Southern Taiwan  

OpenAIRE

Background: The aim of this study was to assess the prevalence of the major nontuberculousmycobacterium (NTM) species and the outcome of their treatment insouthern Taiwan (a high-prevalence area for mycobacterium tuberculosis[MTB]).Methods: The study was a retrospective review of patients with NTM pulmonary diseaseat the Kaohsiung Chang Gung Memorial Hospital from 2004 to 2005.The variables recorded and analyzed included demographics, particularly ageand gender; primary clinical presentations...

Chin-Chou Wang; Meng-Chih Lin; Jien-Wei Li; Yi-Hsi Wang

2009-01-01

338

The changing pattern of nontuberculous mycobacterial disease  

OpenAIRE

Nontuberculous mycobacteria are human opportunistic pathogens whose source of infection is the environment. These include both slow-growing (eg, Mycobacterium kansasii and Mycobacterium avium) and rapid-growing (eg, Mycobacterium abscessus and Mycobacterium fortuitum) species. Transmission is through ingestion or inhalation of water, particulate matter or aerosols, or through trauma. The historic presentation of pulmonary disease in older individuals with predisposing lung conditions and in c...

Falkinham, Joseph O.

2003-01-01

339

Nontuberculous Mycobacterial Lung Disease in Southern Taiwan  

Directory of Open Access Journals (Sweden)

Full Text Available Background: The aim of this study was to assess the prevalence of the major nontuberculousmycobacterium (NTM species and the outcome of their treatment insouthern Taiwan (a high-prevalence area for mycobacterium tuberculosis[MTB].Methods: The study was a retrospective review of patients with NTM pulmonary diseaseat the Kaohsiung Chang Gung Memorial Hospital from 2004 to 2005.The variables recorded and analyzed included demographics, particularly ageand gender; primary clinical presentations; chest radiographic findings; riskfactors; medication and outcome of treatment.Results: The study included 67 patients with NTM pulmonary disease. The averageage was 66.6 14.5 years and they were predominantly male (70.1%. Ofthese patients, 88.1% had pre-existing lung disease, with chronic obstructivepulmonary disease (61.2% and TB (58.2% as the main underlying lung diseases.Rapid-growth species (M. abscessus, 44.8% and M. fortuitum, 23.9%were the most commonly isolated species. Of the forty patients that weretreated and followed up for at least one year, 31 had a favorable outcome(mean duration of therapy, 8.46 2.96 months.Conclusions: The predominant species in southern Taiwan differ from those in other countriesas well as in northern Taiwan, with rapid-growth species predominatingin southern Taiwan.

Chin-Chou Wang

2009-10-01

340

Subcutaneous aspergillosis with coexisting atypical mycobacterial infection  

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Full Text Available A 60-year-old woman, a known diabetic and asthmatic, was admitted for acute exacerbation of chronic obstructive pulmonary disease. Physical examination revealed two soft nodules in the left infra axillary region. Fine needle aspiration cytology (FNAC showed fungal granulomatous reaction suggestive of fungal infection. Periodic acid Schiff stain (PAS stain revealed PAS positive, acutely branching, septate fungal hyphae. Wet mount of the aspirate revealed plenty of pus cells and branching septate hyphae. Ziehl-Neelsen (ZN stain showed moderate numbers of acid fast bacilli. Culture yielded Aspergillus flavus and Mycobacterium fortuitum.

Duraipandian Jeyakumari

2010-04-01

341

Detection of Mycobacterial DNA in Andean Mummies  

OpenAIRE

The identification of genetic material from pathogenic organisms in ancient tissues provides a powerful tool for the study of certain infectious diseases in historic populations. We have obtained tissue samples from the genital areas of 12 mummies in the American Museum of Natural History collection in New York, N.Y. The mummies were excavated in the Andes Mountain region of South America, and radiocarbon dating estimates that the mummies date from a.d. 140 to 1200. DNAs were successfully ext...

Konomi, Nami; Lebwohl, Eve; Mowbray, Ken; Tattersall, Ian; Zhang, David

2002-01-01

342

Mycobacterial subcutaneous arteritis / Arterite subcutânea por micobactéria  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: English Abstract in portuguese Os autores descrevem três casos de pacientes com nódulos subcutâneos em diferentes fases de evolução, com sintomas sistêmicos inespe-cíficos, PPD positivo e raio-X de tórax normal. O estudo anátomo-patológico dos nódulos antigos mostrou arterite granulomatosa e com raros bacilos álcool-ácido resiste [...] ntes na parede vascular; os nódulos recentes mostraram paniculite inespecífica com alguns bacilos álcool-ácido resistentes na parede de vasos cutâneos aparentemente normais. Esses achados sugerem que a micobactéria tem um tropismo vascular e pode causar arterite granulomatosa. Abstract in english The authors report three patients with subcutaneous erythematous nodules in different phases of development, inespecific systemic symptoms, positive PPD test, and normal chest X-rays. The histopathological study of the older nodules showed a granulomatous arteritis with a few acid-fast bacilli in th [...] e vascular wall. The nodules at an early phase showed an inespecific panniculitis with some acid-fast bacilli in apparently normal cutaneous vessels. These findings suggest that the mycobacterium has a vascular tropism and may cause a primary granulomatous arteritis.

Consuelo Junqueira, Rodrigues; Fernando Peixoto Ferraz de, Campos; Lais Lage, Furtado-Mendonça; Rosa Maria Rodrigues, Pereira; Berilo, Langer; Jayme, Diament; Ricardo Manoel de, Oliveira; Wilson, Cossermelli.

1990-10-01

343

Mycobacterial subcutaneous arteritis Arterite subcutânea por micobactéria  

Directory of Open Access Journals (Sweden)

Full Text Available The authors report three patients with subcutaneous erythematous nodules in different phases of development, inespecific systemic symptoms, positive PPD test, and normal chest X-rays. The histopathological study of the older nodules showed a granulomatous arteritis with a few acid-fast bacilli in the vascular wall. The nodules at an early phase showed an inespecific panniculitis with some acid-fast bacilli in apparently normal cutaneous vessels. These findings suggest that the mycobacterium has a vascular tropism and may cause a primary granulomatous arteritis.Os autores descrevem três casos de pacientes com nódulos subcutâneos em diferentes fases de evolução, com sintomas sistêmicos inespe-cíficos, PPD positivo e raio-X de tórax normal. O estudo anátomo-patológico dos nódulos antigos mostrou arterite granulomatosa e com raros bacilos álcool-ácido resistentes na parede vascular; os nódulos recentes mostraram paniculite inespecífica com alguns bacilos álcool-ácido resistentes na parede de vasos cutâneos aparentemente normais. Esses achados sugerem que a micobactéria tem um tropismo vascular e pode causar arterite granulomatosa.

Consuelo Junqueira Rodrigues

1990-10-01

344

Mycobacterial infection masquerading as cutaneous sarcoidosis.  

Science.gov (United States)

Granulomatous skin inflammation can be secondary to a number of infectious and noninfectious aetiologies. Although the presence of Mycobacterium gordonae is commonly thought to reflect environmental contamination, infection can occur even in immunocompetent hosts. Proper diagnostic evaluation of granulomatous inflammation of the skin is essential in order to optimize treatment recommendations. PMID:19040506

Moss, J; Zic, J; Drake, W

2009-07-01

345

Mycobacterial disease, immunosuppression, and acquired immunodeficiency syndrome.  

OpenAIRE

The mycobacteria are an important group of acid-fast pathogens ranging from obligate intracellular parasites such as Mycobacterium leprae to environmental species such as M. gordonae and M. fortuitum. The latter may behave as opportunistic human pathogens if the host defenses have been depleted in some manner. The number and severity of such infections have increased markedly with the emergence of the acquired immunodeficiency syndrome (AIDS) epidemic. These nontuberculous mycobacteria tend t...

Collins, F. M.

1989-01-01

346

Principles of gene therapy  

Directory of Open Access Journals (Sweden)

Full Text Available Genes are specific sequences of bases that encode instructions to make proteins. When genes are altered so that encoded proteins are unable to carry out their normal functions, genetic disorders can result. Gene therapy is designed to introduce genetic material into cells to compensate for abnormal genes or to make a beneficial protein. This article reviews the fundamentals in gene therapy and its various modes of administration with an insight into the role of gene therapy in Periodontics and future percepts and the technical and ethical issues of using gene therapy.

Mammen Biju

2007-01-01

347

Gene-Gene Cooperativity in Small Networks  

OpenAIRE

We show how to construct a reduced description of interacting genes in noisy, small regulatory networks using coupled binary spin variables. Treating both the protein number and gene expression state variables stochastically and on equal footing, we propose a mapping that connects the molecular level description of networks to the binary representation. We construct a phase diagram indicating when genes can be considered to be independent and when the coupling between them cannot be neglected...

Walczak, Aleksandra M.; Wolynes, Peter G.

2009-01-01

348

Exploiting gene × gene interaction in linkage analysis  

OpenAIRE

Abstract When two genes interact to cause a clinically important phenotype, it would seem reasonable to expect that we could leverage genotypic information at one of the loci in order to improve our ability to detect the other. We were therefore interested in extending the posterior probability of linkage (PPL), a class of linkage statistics we have been developing over the past decade, in order to explicitly allow for gene × gene interaction. In this report we utilize a new implem...

Huang Yungui; Bartlett Christopher W; Segre Alberto M; O'Connell Jeffrey R; Mangin LaVonne; Vieland Veronica J

2007-01-01

349

Gene finding by integrating gene finders  

Directory of Open Access Journals (Sweden)

Full Text Available Gene finding, the accurate annotation of genomic DNA, has become one of the central topics in biological research. Although various computational methods (gene finders have been proposed and developed, they all have their own limitations in gene findings. In this paper, we introduce an integrating gene finder, which combines the results of several existing gene finders together, to improve the accuracy of gene finding. Four integration schemes, based on majority voting, are developed for the analysis of two datasets – the basic dataset and the testing dataset. The basic dataset consists of 1500 DNA sequences and the testing dataset consists of 103 DNA sequences. It is demonstrated that a simple integration (a simple voting for each nucleotide can significantly improve the finding performance, and removing confusing gene finders, caused by poor performance or redundant results, is important for a further improvement of the integration. The best prediction results are obtained using weighted majority voting, aided by the mRMR (Minimum Redundancy Maximum Relevance (Peng, 2005 method for the gene finder selection. The prediction accuracies are 84.16% and 90.06% for the basic dataset and testing dataset respectively, which are better than any individual gene finding software in our research.

Yudong Cai

2010-11-01

350

Human Gene Therapy: Genes without Frontiers?  

Science.gov (United States)

Describes the latest advancements and setbacks in human gene therapy to provide reference material for biology teachers to use in their science classes. Focuses on basic concepts such as recombinant DNA technology, and provides examples of human gene therapy such as severe combined immunodeficiency syndrome, familial hypercholesterolemia, and…

Simon, Eric J.

2002-01-01

351

A Pilot Study of Gene/Gene and Gene/Environment Interactions in Alzheimer Disease  

OpenAIRE

Background: Although some genes associated with increased risk of Alzheimer Disease (AD) have been identified, few data exist related to gene/gene and gene/environment risk of AD. The purpose of this pilot study was to explore gene/gene and gene/environment associations in AD and to obtain data for sample size estimates for larger, more definitive studies of AD.

Ghebranious, Nader; Mukesh, Bickol; Giampietro, Philip F.; Glurich, Ingrid; Mickel, Susan F.; Waring, Stephen C.; Mccarty, Catherine A.

2011-01-01

352

Gene therapy in transplantation.  

OpenAIRE

In transplantation, gene therapy strategies to prolong graft survival involve gene transfer and expression of immunomodulatory or graft-protecting molecules. The local production of immunosuppressive molecules has the potential to reduce their systemic side effects, and to increase their bioavailability and hence their therapeutic efficiency. Ex vivo gene transfer enables manipulation prior to engraftment. Vectors have now been developed that can optimally transfer the relevant genes to vario...

Wood, Kj; Prior, Tg

2001-01-01

353

Reading and Generalist Genes  

Science.gov (United States)

Twin-study research suggests that many (but not all) of the same genes contribute to genetic influence on diverse learning abilities and disabilities, a hypothesis called "generalist genes". This generalist genes hypothesis was tested using a set of 10 DNA markers (single nucleotide polymorphisms [SNPs]) found to be associated with early reading…

Haworth, Claire M. A.; Meaburn, Emma L.; Harlaar, Nicole; Plomin, Robert

2007-01-01

354

Down-weighting overlapping genes improves gene set analysis  

OpenAIRE

Abstract Background The identification of gene sets that are significantly impacted in a given condition based on microarray data is a crucial step in current life science research. Most gene set analysis methods treat genes equally, regardless how specific they are to a given gene set. Results In this work we propose a new gene set analysis method that computes a gene set score as the mean of absolute values of weighted moderated gene t-scores. The gene weights...

Tarca Adi; Draghici Sorin; Bhatti Gaurav; Romero Roberto

2012-01-01

355

Community structure and PAH ring-hydroxylating dioxygenase genes of a marine pyrene-degrading microbial consortium.  

Science.gov (United States)

Marine microbial consortium UBF, enriched from a beach polluted by the Prestige oil spill and highly efficient in degrading this heavy fuel, was subcultured in pyrene minimal medium. The pyrene-degrading subpopulation (UBF-Py) mineralized 31 % of pyrene without accumulation of partially oxidized intermediates indicating the cooperation of different microbial components in substrate mineralization. The microbial community composition was characterized by culture dependent and PCR based methods (PCR-DGGE and clone libraries). Molecular analyses showed a highly stable community composed by Alphaproteobacteria (84 %, Breoghania, Thalassospira, Paracoccus, and Martelella) and Actinobacteria (16 %, Gordonia). The members of Thalasosspira and Gordonia were not recovered as pure cultures, but five additional strains, not detected in the molecular analysis, that classified within the genera Novosphingobium, Sphingopyxis, Aurantimonas (Alphaproteobacteria), Alcanivorax (Gammaproteobacteria) and Micrococcus (Actinobacteria), were isolated. None of the isolates degraded pyrene or other PAHs in pure culture. PCR amplification of Gram-positive and Gram-negative dioxygenase genes did not produce results with any of the cultured strains. However, sequences related to the NidA3 pyrene dioxygenase present in mycobacterial strains were detected in UBF-Py consortium, suggesting the representative of Gordonia as the key pyrene degrader, which is consistent with a preeminent role of actinobacteria in pyrene removal in coastal environments affected by marine oil spills. PMID:24356981

Gallego, Sara; Vila, Joaquim; Tauler, Margalida; Nieto, José María; Breugelmans, Philip; Springael, Dirk; Grifoll, Magdalena

2014-07-01

356

Gene Gateway: Exploring Genes and Genetic Disorders  

Science.gov (United States)

This collection of guides and tutorials is intended to help users take advantage of of online data sources from the Human Genome Project for learning about genetic disorders, genes, and proteins. Resources include the Gene Gateway Workbook, a downloadable tutorial consisting of activities with screenshots and instructions, that helps new users locate and use genetic-disorder and bioinformatics resources on the web. There are also resources for learning about genes and the proteins they encode; tips, tutorials, and terminology for using selected resources in the Genome Database Guide; a guide to nontechnical resources on genetic disorder descriptions and treatments; a human genome landmarks poster; and others.

357

Gene hunting in autoinflammation.  

Science.gov (United States)

Steady progress in our understanding of the genetic basis of autoinflammatory diseases has been made over the past 16 years. Since the discovery of the familial Mediterranean fever gene MEFV (also known as marenostrin) in 1997, 18 other genes responsible for monogenic autoinflammatory diseases have been identified to date. The discovery of these genes was made through the utilisation of many genetic mapping techniques, including next generation sequencing platforms. This review article clearly describes the gene hunting approaches, methods of data analysis and the technological platforms used, which has relevance to all those working within the field of gene discovery for Mendelian disorders. PMID:24070009

Standing, Ariane; Omoyinmi, Ebun; Brogan, Paul

2013-01-01

358

Gene expression in fungi  

Directory of Open Access Journals (Sweden)

Full Text Available This contribution is based on the four presentations made at the Special Interest Group (SIG meeting titled Gene Expression in Fungi held during IMC9 in Edinburgh. This overview is independent from other articles published or that will be published by each speaker. In the SIG meeting, basic principles of in vivo animal models for virulence studies were discussed. Infection associated genes of Candida albicans and fungal adaptation to the host was summarized. Azole susceptibility was evaluated as a combined result of several changes in expression of pertinent genes. Gene transfer in fungi, resulting in fungal evolution and gene adaptation to environmental factors, was reported.

A. Kalkanci

2011-06-01

359

UniGene  

Science.gov (United States)

Created by the National Center for Biotechnology Information, UniGene is "an experimental system for automatically partitioning GenBank sequences into a non-redundant set of gene-oriented clusters." In addition to gene sequences, this Web site also offers thousands of novel expressed sequence tag (EST) sequences, a useful gene discovery resource. Organisms currently cataloged include human, rat, mouse, cow, zebrafish, clawed frog, fruitfly, mosquito, wheat, rice, barley, maize, and cress. Users may also access the Digital Differential Display to compare gene expression fingerprints for cancer cells and their normal counterparts. Other Web site features include query tips, FAQs, and relevant external links.

360

Retrieval with gene queries  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background Accuracy of document retrieval from MEDLINE for gene queries is crucially important for many applications in bioinformatics. We explore five information retrieval-based methods to rank documents retrieved by PubMed gene queries for the human genome. The aim is to rank relevant documents higher in the retrieved list. We address the special challenges faced due to ambiguity in gene nomenclature: gene terms that refer to multiple genes, gene terms that are also English words, and gene terms that have other biological meanings. Results Our two baseline ranking strategies are quite similar in performance. Two of our three LocusLink-based strategies offer significant improvements. These methods work very well even when there is ambiguity in the gene terms. Our best ranking strategy offers significant improvements on three different kinds of ambiguities over our two baseline strategies (improvements range from 15.9% to 17.7% and 11.7% to 13.3% depending on the baseline. For most genes the best ranking query is one that is built from the LocusLink (now Entrez Gene summary and product information along with the gene names and aliases. For others, the gene names and aliases suffice. We also present an approach that successfully predicts, for a given gene, which of these two ranking queries is more appropriate. Conclusion We explore the effect of different post-retrieval strategies on the ranking of documents returned by PubMed for human gene queries. We have successfully applied some of these strategies to improve the ranking of relevant documents in the retrieved sets. This holds true even when various kinds of ambiguity are encountered. We feel that it would be very useful to apply strategies like ours on PubMed search results as these are not ordered by relevance in any way. This is especially so for queries that retrieve a large number of documents.

Srinivasan Padmini

2006-04-01

361

Essential Bacillus subtilis genes  

DEFF Research Database (Denmark)

To estimate the minimal gene set required to sustain bacterial life in nutritious conditions, we carried out a systematic inactivation of Bacillus subtilis genes. Among approximate to4,100 genes of the organism, only 192 were shown to be indispensable by this or previous work. Another 79 genes were predicted to be essential. The vast majority of essential genes were categorized in relatively few domains of cell metabolism, with about half involved in information processing, one-fifth involved in the synthesis of cell envelope and the determination of cell shape and division, and one-tenth related to cell energetics. Only 4% of essential genes encode unknown functions. Most essential genes are present throughout a wide range of Bacteria, and almost 70% can also be found in Archaea and Eucarya. However, essential genes related to cell envelope, shape, division, and respiration tend to be lost from bacteria with small genomes. Unexpectedly, most genes involved in the Embden-Meyerhof-Parnas pathway are essential.Identification of unknown and unexpected essential genes opens research avenues to better understanding of processes that sustain bacterial life.

Kobayashi, K.; Ehrlich, S.D.

2003-01-01

362

Cancer gene therapy  

Directory of Open Access Journals (Sweden)

Full Text Available Cancer gene therapy can be defined as transfer of nucleic acids into tumor or normal cells with aim to eradicate or reduce tumor mass by direct killing of cells, immunomodulation or correction of genetic errors, and reversion of malignant status. Initially started with lots of optimism and enthusiasm, cancer gene therapy has shown limited success in treatment of patients. This review highlights current limitations and almost endless possibilities of cancer gene therapy. The major difficulty in advancing gene therapy technology from the bench to the clinical practice is problem with gene delivery vehicles (so called vectors needed to ferry genetic material into a cell. Despite few reports of therapeutic responses in some patients, there is still no proof of clinical efficacy of most cancer gene therapy approaches, primarily due to very low transduction and expression efficacy in vivo of available vectors. An "ideal" gene therapy vector should be administrated through a noninvasive route and should be targeted not only to primary tumor mass but also to disseminated tumor cells and micrometastases; it should also carry therapeutic gene with tumor-restricted, time-regulated, and sustained expression. Current strategies for combating the cancer with gene therapy can be divided into four basic concepts: (1 replacement of missing tumor suppressor gene and/or blocking of oncogenes or pro-inflammatory genes, (2 suicide gene strategies, (3 induction of immune-mediated destruction, and (4 inhibition of tumor angiogenesis. The advance in the clinical benefit of gene therapy will probably be first achieved with combining it with standard cancer treatment: chemotherapy, radiotherapy, and immunotherapy.

Mitrovi? Tatjana

2005-01-01

363

A Fragment of 21 ORFs Around the Direct Repeat (DR Region of Mycobacterium tuberculosis is Absent From the Other Sequenced Mycobacterial Genomes: Implications for the Evolution of the DR Region  

Directory of Open Access Journals (Sweden)

Full Text Available The direct repeat (DR region is a singular locus of the Mycobacterium tuberculosis complex genome. This region consists of 36 bp repetitive sequences separated by non-repetitive unique spacer sequences. Around this region there are several genes coding for proteins of unknown function. To determine whether the M. smegmatis, M. avium, M. marinum and M. leprae genomes contain sequences and ORFs similar to those of the DR locus of the M. tuberculosis complex, we analysed the corresponding regions in these species. As a first step, some conserved genes that flank the DR genes [Rv2785c (rpsO, Rv2786c (ribF, Rv2790c (ltp1 , Rv2793c (truB, Rv2800, Rv2825, Rv2828, Rv2831 (echA16 , Rv2838 (rbfA and Rv2845 (proS ] were used as markers to locate the corresponding orthologues in M. smegmatis, M. avium, M. marinum and M. leprae in silico. Most of these M. tuberculosis marker genes have highly similar orthologues located in the same order and orientation in the other mycobacteria. In contrast, no orthologues were found for ORFs Rv2801–Rv2824, suggesting that these genes are unique to M. tuberculosis within the genus Mycobacterium.We observed that in M. smegmatis and M. avium, Rv2800 and Rv2825 are adjacent. This observation was experimentally confirmed by PCR. In conclusion, as the DR locus and the ORFs around it are absent in M. smegmatis and M. avium and, as it is possible that these species are older than M. tuberculosis, we postulated that the DR locus was acquired by the M. tuberculosis complex species or by an ancestor bacterium.

Angel Cataldi

2004-01-01

364

Gene Positioning and Expression  

OpenAIRE

Within the nucleus, the genome is spatially organized. Individual chromosomes are non-randomly positioned with respect to each other and with respect to nuclear landmarks [1,2]. Furthermore, the position of individual genes can reflect their expression. Here we discuss two well-characterized examples of gene relocalization associated with transcriptional activation: 1) developmentally regulated genes that move from the nuclear periphery to transcription factories in the nucleoplasm upon induc...

Egecioglu, Defne; Brickner, Jason H.

2011-01-01

365

DNA, Genes and Chromosomes  

Science.gov (United States)

Today you will learn about the parts of DNA and what DNA, genes and chromosomes are. Today you will learn what DNA, genes and chromosomes are and the parts of the DNA molecule. Look at all of the websites, take whatever notes you need to. At the end of the assignment, be able to describle DNA, the parts of DNA, genes and chromosomes. Covers Biology Core Curriculum, ...

Mrs. Fomby

2007-11-07

366

Genes and Disease  

Science.gov (United States)

The National Center for Biotechnology Information of the National Library of Medicine (part of the National Institutes of Health) has posted this webpage, Genes and Disease, which provides information "for some 60 diseases associated with specific genes, and has links to the 1998 Gene Map as well as to PubMed, protein sequences, Online Mendelian Inheritance in Man, and associations related to each disease."

1998-01-01

367

Functional analyses of mycobacterial lipoprotein diacylglyceryl transferase and comparative secretome analysis of a mycobacterial lgt mutant.  

Science.gov (United States)

Preprolipopoprotein diacylglyceryl transferase (Lgt) is the gating enzyme of lipoprotein biosynthesis, and it attaches a lipid structure to the N-terminal part of preprolipoproteins. Using Lgt from Escherichia coli in a BLASTp search, we identified the corresponding Lgt homologue in Mycobacterium tuberculosis and two homologous (MSMEG_3222 and MSMEG_5408) Lgt in Mycobacterium smegmatis. M. tuberculosis lgt was shown to be essential, but an M. smegmatis ?MSMEG_3222 mutant could be generated. Using Triton X-114 phase separation and [(14)C]palmitic acid incorporation, we demonstrate that MSMEG_3222 is the major Lgt in M. smegmatis. Recombinant M. tuberculosis lipoproteins Mpt83 and LppX are shown to be localized in the cell envelope of parental M. smegmatis but were absent from the cell membrane and cell wall in the M. smegmatis ?MSMEG_3222 strain. In a proteomic study, 106 proteins were identified and quantified in the secretome of wild-type M. smegmatis, including 20 lipoproteins. All lipoproteins were secreted at higher levels in the ?MSMEG_3222 mutant. We identify the major Lgt in M. smegmatis, show that lipoproteins lacking the lipid anchor are secreted into the culture filtrate, and demonstrate that M. tuberculosis lgt is essential and thus a validated drug target. PMID:22609911

Tschumi, Andreas; Grau, Thomas; Albrecht, Dirk; Rezwan, Mandana; Antelmann, Haike; Sander, Peter

2012-08-01

368

Gene positioning and expression.  

Science.gov (United States)

Within the nucleus, the genome is spatially organized. Individual chromosomes are non-randomly positioned with respect to each other and with respect to nuclear landmarks [1,2]. Furthermore, the position of individual genes can reflect their expression. Here we discuss two well-characterized examples of gene relocalization associated with transcriptional activation: 1) developmentally regulated genes that move from the nuclear periphery to transcription factories in the nucleoplasm upon induction and 2) genes that are targeted from the nucleoplasm to the nuclear periphery, through interactions with the nuclear pore complex (NPC), upon activation. Finally, we speculate as to the mechanistic and functional commonalities of these phenomena. PMID:21292462

Egecioglu, Defne; Brickner, Jason H

2011-06-01

369

Gene Conversion and Evolution of Gene Families: An Overview  

OpenAIRE

The importance of gene conversion for the evolution of gene families is reviewed. Four problems concerning gene conversion, i.e., concerted evolution, generation of useful variation, deleterious effects, and relation to neofunctionalization, are discussed by surveying reported examples of evolving gene families. Emphasis is given toward understanding interactive effects of gene conversion and natural selection.

Tomoko Ohta

2010-01-01

370

Gene Conversion and Evolution of Gene Families: An Overview  

Directory of Open Access Journals (Sweden)

Full Text Available The importance of gene conversion for the evolution of gene families is reviewed. Four problems concerning gene conversion, i.e., concerted evolution, generation of useful variation, deleterious effects, and relation to neofunctionalization, are discussed by surveying reported examples of evolving gene families. Emphasis is given toward understanding interactive effects of gene conversion and natural selection.

Tomoko Ohta

2010-09-01

371

Clinical and Laboratory Features of Mycobacterium porcinum†  

OpenAIRE

Recent molecular studies have shown Mycobacterium porcinum, recovered from cases of lymphadenitis in swine, to have complete 16S rDNA sequence identity and >70% DNA-DNA homology with human isolates within the M. fortuitum third biovariant complex. We identified 67 clinical and two environmental isolates of the M. fortuitum third biovariant sorbitol-negative group, of which 48 (70%) had the same PCR restriction enzyme analysis (PRA) profile as the hsp65 gene of M. porcinum (ATCC 33776T) and we...

Wallace, Richard J.; Brown-elliott, Barbara A.; Wilson, Rebecca W.; Mann, Linda; Hall, Leslie; Zhang, Yansheng; Jost, Kenneth C.; Brown, June M.; Kabani, Amin; Schinsky, Mark F.; Steigerwalt, Arnold G.; Crist, Christopher J.; Roberts, Glenn D.; Blacklock, Zeta; Tsukamura, Michio

2004-01-01

372

Mycobacterium paraterrae sp. nov. recovered from a clinical specimen: novel chromogenic slow growing mycobacteria related to Mycobacterium terrae complex.  

Science.gov (United States)

A previously unidentified, slowly growing scotochromogenic Mycobacterium was isolated from a Korean patient with symptomatic pulmonary infection. Phenotypically, this strain was generally similar to Mycobacterium terrae complex strains, however it uniquely produced orange pigmentation. Unique mycolic acid profiles and phylogenetic analyses based on three alternative chronometer molecules, 16S rRNA gene, hsp65 and rpoB, confirmed the taxonomic status of this strain as a novel species. These results support that this strain represents a novel Mycobacterium species. The name Mycobacterium paraterrae sp. nov. is proposed. The type strain is 05-2522 (= DSM 45127 = KCTC 19556). PMID:20055942

Lee, Hyungki; Lee, Seoung-Ae; Lee, In-Kyung; Yu, Hee-Kyung; Park, Young-Gil; Jeong, Joseph; Lee, Seon Ho; Kim, Sung-Ryul; Hyun, Jin-Won; Kim, Kijeong; Kook, Yoon-Hoh; Kim, Bum-Joon

2010-01-01

373

A review on microcephaly genes  

OpenAIRE

This review aims to summarize the recent findings regarding microcephaly genes. We have discussed the molecular genetics studies of microcephaly genes including a comprehensive appraisal of the seven mapped loci (MCPH1–MCPH7), their corresponding genes and protein products of the genes, their likely role in normal brain development and the details of the mutations reported in these genes.

Irshad S.; Shahid S.

2012-01-01

374

GENE EXPRESSION NETWORKS  

Science.gov (United States)

"Gene expression network" is the term used to describe the interplay, simple or complex, between two or more gene products in performing a specific cellular function. Although the delineation of such networks is complicated by the existence of multiple and subtle types of intera...

375

Making a Gene Library  

Science.gov (United States)

This animation shows how to make a colony of new DNA in order to locate a specific gene of interest. This is the third of four animations detailing the gene cloning process. To begin at the beginning, choose Making a Recombinant Plasmid. (The animation prior to this one is Bacteria Transformation. The animation just after this one is Screening a DNA Library.)

376

Development of a quantitative analysis method for mRNA from Mycobacterium leprae and slow-growing acid-fast bacteria  

International Nuclear Information System (INIS)

This study aimed to develop a specific method for detection and quantitative determination of mRNA that allows estimation of viable counts of M. leprae and other mycobacteria. Of heart-shock protein of 65 kDa (hsp65), mRNA was used as an indicator to discriminate the living cells and died ones. To compare mRNA detections by RNase protection assay (RPA) and Northern blot hybridization (NBH), labelled anti-sense RNA for hsp65 gene of M. leprae was synthesized using plasmid pUC8/N5. The anti-sense RNA synthesized from the template DNA containing about 580 bp (194 to 762) of hsp65 gene. When compared with NBH method, the amount of probe required for the detection by RPA method was 1/30 or less and the detection sensitivity of RPA was also 10 times higher. In addition, complicated procedures were needed to eliminate non-specific reactions in NBH method. These results indicated that RPA method is more convenient and superior for the mRNA detection. However, isotope degradation in the probe used for RPA method might affect the results. Therefore, 33P of 35P, of which degradation energy is less that 32P should be used for labelling. Total RNA was effectively extracted from M. chelonae, M. marinum by AGPC method, but not from M. leprae. In conclusion, RPA is a very effective detection method for these mRNA, but it seems necessary to further improve the sensitivity of detection for a small amount of test materials. (M.N.) materials. (M.N.)

377

Molecular Characterization of Mycobacterium massiliense and Mycobacterium bolletii in Isolates Collected from Outbreaks of Infections after Laparoscopic Surgeries and Cosmetic Procedures?  

Science.gov (United States)

An outbreak of infections affecting 311 patients who had undergone different invasive procedures occurred in 2004 and 2005 in the city of Belém, in the northern region of Brazil. Sixty-seven isolates were studied; 58 were from patients who had undergone laparoscopic surgeries, 1 was from a patient with a postinjection abscess, and 8 were from patients who had undergone mesotherapy. All isolates were rapidly growing nonpigmented mycobacteria and presented a pattern by PCR-restriction enzyme analysis of the hsp65 gene with BstEII of bands of 235 and 210 bp and with HaeIII of bands of 200, 70, 60, and 50 bp, which is common to Mycobacterium abscessus type 2, Mycobacterium bolletii, and Mycobacterium massiliense. hsp65 and rpoB gene sequencing of a subset of 20 isolates was used to discriminate between these three species. hsp65 and rpoB sequences chosen at random from 11 of the 58 isolates from surgical patients and the postinjection abscess isolate presented the highest degrees of similarity with the corresponding sequences of M. massiliense. In the same way, the eight mesotherapy isolates were identified as M. bolletii. Molecular typing by pulsed-field gel electrophoresis (PFGE) grouped all 58 surgical isolates, while the mesotherapy isolates presented three different PFGE patterns and the postinjection abscess isolate showed a unique PFGE pattern. In conclusion, molecular techniques for identification and typing were essential for the discrimination of two concomitant outbreaks and one case, the postinjection abscess, not related to either outbreak, all of which were originally attributed to a single strain of M. abscessus. PMID:18174307

Viana-Niero, Cristina; Lima, Karla Valéria Batista; Lopes, Maria Luiza; da Silva Rabello, Michelle Christiane; Marsola, Lourival Rodrigues; Brilhante, Vânia Cristina Ribeiro; Durham, Alan Mitchel; Leão, Sylvia Cardoso

2008-01-01

378

GeneCards Version 3: the human gene integrator  

OpenAIRE

GeneCards (www.genecards.org) is a comprehensive, authoritative compendium of annotative information about human genes, widely used for nearly 15 years. Its gene-centric content is automatically mined and integrated from over 80 digital sources, resulting in a web-based deep-linked card for each of >73 000 human gene entries, encompassing the following categories: protein coding, pseudogene, RNA gene, genetic locus, cluster and uncategorized. We now introduce GeneCards Version 3, featuring a ...

Safran, Marilyn; Dalah, Irina; Alexander, Justin; Rosen, Naomi; Iny Stein, Tsippi; Shmoish, Michael; Nativ, Noam; Bahir, Iris; Doniger, Tirza; Krug, Hagit; Sirota-madi, Alexandra; Olender, Tsviya; Golan, Yaron; Stelzer, Gil; Harel, Arye

2010-01-01

379

Gene-gene interaction filtering with ensemble of filters  

OpenAIRE

Abstract Background Complex diseases are commonly caused by multiple genes and their interactions with each other. Genome-wide association (GWA) studies provide us the opportunity to capture those disease associated genes and gene-gene interactions through panels of SNP markers. However, a proper filtering procedure is critical to reduce the search space prior to the computationally intensive gene-gene interaction identification step. In this study, we show that two commonly ...

Wk, Ho Joshua; Yang Pengyi; Yang Yee; Zhou Bing B

2011-01-01

380

Methods for detecting gene × gene interaction in multiplex extended pedigrees  

OpenAIRE

Abstract Complex diseases are multifactorial in nature and can involve multiple loci with gene × gene and gene × environment interactions. Research on methods to uncover the interactions between those genes that confer susceptibility to disease has been extensive, but many of these methods have only been developed for sibling pairs or sibships. In this report, we assess the performance of two methods for finding gene × gene interactions that are applicable to arbitrarily sized pe...

Cooper Margaret E; Goldstein Toby H; Maher Brion S; Brock Guy N; Marazita Mary L

2005-01-01

381

GeneMark.hmm: new solutions for gene finding.  

OpenAIRE

The number of completely sequenced bacterial genomes has been growing fast. There are computer methods available for finding genes but yet there is a need for more accurate algorithms. The GeneMark. hmm algorithm presented here was designed to improve the gene prediction quality in terms of finding exact gene boundaries. The idea was to embed the GeneMark models into naturally derived hidden Markov model framework with gene boundaries modeled as transitions between hidden states. We also used...

Lukashin, A. V.; Borodovsky, M.

1998-01-01

382

Gene expression networks.  

Science.gov (United States)

With the advent of microarrays and next-generation biotechnologies, the use of gene expression data has become ubiquitous in biological research. One potential drawback of these data is that they are very rich in features or genes though cost considerations allow for the use of only relatively small sample sizes. A useful way of getting at biologically meaningful interpretations of the environmental or toxicological condition of interest would be to make inferences at the level of a priori defined biochemical pathways or networks of interacting genes or proteins that are known to perform certain biological functions. This chapter describes approaches taken in the literature to make such inferences at the biochemical pathway level. In addition this chapter describes approaches to create hypotheses on genes playing important roles in response to a treatment, using organism level gene coexpression or protein-protein interaction networks. Also, approaches to reverse engineer gene networks or methods that seek to identify novel interactions between genes are described. Given the relatively small sample numbers typically available, these reverse engineering approaches are generally useful in inferring interactions only among a relatively small or an order 10 number of genes. Finally, given the vast amounts of publicly available gene expression data from different sources, this chapter summarizes the important sources of these data and characteristics of these sources or databases. In line with the overall aims of this book of providing practical knowledge to a researcher interested in analyzing gene expression data from a network perspective, the chapter provides convenient publicly accessible tools for performing analyses described, and in addition describe three motivating examples taken from the published literature that illustrate some of the relevant analyses. PMID:23086841

Thomas, Reuben; Portier, Christopher J

2013-01-01

383

Gene Testing for Hereditary Ataxia  

Science.gov (United States)

... onset. For which of the hereditary ataxias is gene testing available? Discovery of specific ataxia genes makes ... develop more effective treatments. FREQUENTLY ASKED QUESTIONS ABOUT... Gene Testing for Hereditary Ataxia This fact sheet provides ...

384

Gene amplification in carcinogenesis  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: English Abstract in english Gene amplification increases the number of genes in a genome and can give rise to karyotype abnormalities called double minutes (DM) and homogeneously staining regions (HSR), both of which have been widely observed in human tumors but are also known to play a major role during embryonic development [...] due to the fact that they are responsible for the programmed increase of gene expression. The etiology of gene amplification during carcinogenesis is not yet completely understood but can be considered a result of genetic instability. Gene amplification leads to an increase in protein expression and provides a selective advantage during cell growth. Oncogenes such as CCND1, c-MET, c-MYC, ERBB2, EGFR and MDM2 are amplified in human tumors and can be associated with increased expression of their respective proteins or not. In general, gene amplification is associated with more aggressive tumors, metastases, resistance to chemotherapy and a decrease in the period during which the patient stays free of the disease. This review discusses the major role of gene amplification in the progression of carcinomas, formation of genetic markers and as possible therapeutic targets for the development of drugs for the treatment of some types of tumors.

Lucimari, Bizari; Ana Elizabete, Silva; Eloiza H., Tajara.

385

SOS induction in mycobacteria: analysis of the DNA-binding activity of a LexA-like repressor and its role in DNA damage induction of the recA gene from Mycobacterium smegmatis.  

Science.gov (United States)

The protein encoded by the lexA gene from Mycobacterium leprae was overproduced in Escherichia coli. The recombinant protein bound to the promoter regions of the M. leprae lexA, M. leprae recA and M. smegmatis recA genes at sites with the sequences 5'-GAACACATGTTT and 5'-GAACAGGTGTTC, which belong to the 'Cheo box' family of binding sites recognized by the SOS repressor from Bacillus subtilis. Gel mobility shift assays were used to confirm that proteins with the same site specificity of DNA binding are also present in Mycobacterium tuberculosis and M. smegmatis. Complex formation was impaired by mutagenic disruption of the dyad symmetry of the M. smegmatis recA Cheo box. LexA binding was also inhibited by preincubation of the M. smegmatis and M. tuberculosis extracts with anti-M. leprae LexA antibodies, suggesting that the mycobacterial LexA proteins are functionally conserved at the level of DNA binding. Finally, exposure of M. smegmatis to DNA-damaging agents resulted in induction of the M. smegmatis recA promoter with concomitant loss of DNA binding of LexA to its Cheo box, confirming that this organism possesses the key regulatory elements of a functional SOS induction system. PMID:9427395

Durbach, S I; Andersen, S J; Mizrahi, V

1997-11-01

386

Terapia gênica Gene therapy  

Directory of Open Access Journals (Sweden)

Full Text Available Terapia gênica é um procedimento médico que envolve a modificação genética de células como forma de tratar doenças. Os genes influenciam praticamente todas as doenças humanas, seja pela codificação de proteínas anormais diretamente responsáveis pela doença, seja por determinar suscetibilidade a agentes ambientais que a induzem. A terapia gênica é ainda experimental, e está sendo estudada em protocolos clínicos para diferentes tipos de doenças. O desenvolvimento de métodos seguros e eficientes de transferência gênica para células humanas é um dos pontos mais importantes na terapia gênica. Apesar do grande esforço dirigido na última década para o aperfeiçoamento dos protocolos de terapia gênica humana, e dos avanços importantes na pesquisa básica, as aplicações terapêuticas da tecnologia de transferência gênica continuam ainda em grande parte teóricas. O potencial da terapia gênica é muito grande, devendo ainda causar grande impacto em todos os aspectos da medicina.Gene therapy is a medical intervention that involves modifying the genetic material of living cells to fight disease. Genes influence virtually every human disease, either by encoding for abnormal proteins, which are directly responsible for the disease, or by causing a susceptibility to environmental agents which induce it. Gene therapy is still experimental, and is being studied in clinical trials for many different types of diseases. The development of safe and effective methods of implanting normal genes into the human cell is one of the most important technical issues in gene therapy. Although much effort has been directed in the last decade toward improvement of protocols in human gene therapy, and in spite of many considerable achievements in basic research, the therapeutic applications of gene transfer technology still remain mostly theoretical. The potential for gene therapy is huge and likely to impact on all aspects of medicine.

Nance Beyer Nardi

2002-01-01

387

Methanogenesis and methane genes  

International Nuclear Information System (INIS)

An overview of the pathways leading to methane biosynthesis is presented. The steps investigated to date by gene cloning and DNA sequencing procedures are identified and discussed. The primary structures of component C of methyl coenzyme M reductase encoded by mcr operons in different methanogens are compared. Experiments to detect the primary structure of the genes encoding F420 reducing hydrogenase (frhABG) and methyl hydrogen reducing hydrogenase (mvhDGA) in methanobacterium thermoautotrophicum strain H are compared with each other and with eubacterial hydrogenase encoding genes. A biotechnological use for hydrogenases from hypermorphillic archaebacteria is suggested. (author)

388

Further understanding human disease genes by comparing with housekeeping genes and other genes  

OpenAIRE

Abstract Background Several studies have compared various features of heritable disease genes with other so called non-disease genes, but they have yielded some conflicting results. A potential problem in those studies is that the non-disease genes contained a large number of essential genesgenes which are indispensable for humans to survive and reproduce. Since a functional disruption of an essential gene has fatal consequences, it's more reasonable to regard essential ...

Chen Ting; Zhou Xianghong; Xu Min; Wang Li; Tu Zhidong; Sun Fengzhu

2006-01-01

389

GEIRA: gene-environment and gene–gene interaction research application  

OpenAIRE

Abstract The GEIRA (Gene-Environment and Gene–Gene Interaction Research Application) algorithm and subsequent program is dedicated to genome-wide gene-environment and gene–gene interaction analysis. It implements concepts of both additive and multiplicative interaction as well as calculations based on dominant, recessive and co-dominant genetic models, respectively. Estimates of interactions are incorporated in a single table to make the output easily read. The algorithm is cod...

2011-01-01

390

Detection of Mycobacterium tuberculosis in sputum, pleural and bronchoalveolar lavage fluid using DNA amplification of the MPB 64 protein coding gene and IS6110 insertion element.  

Science.gov (United States)

Two gene sequences specific for Mycobacterium tuberculosis were evaluated for the diagnosis of pulmonary tuberculous (PTB) in pleural fluid (PF), bronchoalveolar lavage fluid (BAL) and sputum (Sp). The 240 bp sequence (nts 460-700) coding for the MPB 64 protein coding gene and the 123 bp IS6110 insertion element present in multiple copies in the mycobacterial genome were amplified using the polymerase chain reaction. Fifty-nine clinical specimens were studied. The diagnosis of PTB was confirmed by positive M. tuberculosis cultures in 14 specimens, and by the presence of characteristic histological features of granuloma and Langerhan's giant cells on pleural biopsy in 3 PF specimens through cultures for M. tuberculosis were negative. The remaining 42 specimens were obtained from patient's with non-tuberculosis pulmonary infections or malignancy, and these served as negative controls. Our results showed that the IS6110 insertion element and MPB 64 gene sequence were detected in all 14 culture positive PTB cases, although detection of the latter sequence required both DNA amplification and oligonucleotide hybridization. There was however one false positive specimen with the MPB 64 detection protocol. More importantly, both the MPB 64 sequence and IS6110 insertion element protocols were unable to detect M. tuberculosis DNA in the 3 PF samples diagnosed by histological characteristics on pleural biopsy and culture negative. We conclude that DNA amplification for M. tuberculosis-specific sequences is a useful method for rapid diagnosis of PTB in culture positive specimens. However, the false negative results with TB culture negative cases of tuberculosis pleurisy, limits its usefulness for the diagnosis of tuberculous pleurisy. PMID:8629054

Tan, J; Lee, B W; Lim, T K; Chin, N K; Tan, C B; Xia, J R; Yap, H K; Kumarasinghe, G

1995-06-01

391

Hypospadias : Gene mapping and candidate gene studied  

OpenAIRE

Hypospadias is a common congenital malformation in boys, characterized by incomplete fusion of the urethral folds, abnormal opening of urethra and different degrees of curvature of the penis. In Sweden, the incidence of hypospadias is 1.14 per 300 male live-births according to the annual Swedish Malformation Registry. Hypospadias is considered to be a complex genetic disorder caused by the interplay between environmental factors and the additive effects of multiple genes. Se...

Trinh, Thi Thai Hanh

2009-01-01

392

Some Genes Are Dominant  

Science.gov (United States)

This interactive activity, adapted from the Dolan DNA Learning Center, illustrates how Gregor Mendel used pure-bred yellow and green peas to show that some genes are dominant and others are recessive.

Foundation, Wgbh E.

2007-04-19

393

Finding schizophrenia genes  

OpenAIRE

Genetic epidemiological studies suggest that individual variation in susceptibility to schizophrenia is largely genetic, reflecting alleles of moderate to small effect in multiple genes. Molecular genetic studies have identified a number of potential regions of linkage and 2 associated chromosomal abnormalities, and accumulating evidence favors several positional candidate genes. These findings are grounds for optimism that insight into genetic factors associated with schizophrenia will help ...

Kirov, George; O’donovan, Michael C.; Owen, Michael J.

2005-01-01

394

Genes, chromosomes, and rhabdomyosarcoma.  

Science.gov (United States)

Rhabdomyosarcomas are a heterogeneous group of malignant tumors and are the most common soft-tissue sarcoma of childhood. Rhabdomyosarcomas resemble developing skeletal muscle, notably in their expression of the MRF family of transcription factors and the PAX3 and PAX7 genes. These PAX genes are also involved through specific translocations, t(2;13)(q35;q14) and variant t(1;13)(p36;q14) in the alveolar subtype, which result in PAX3-FKHR and PAX7-FKHR fusion genes, respectively. The fusion genes are thought critically to affect downstream targets of PAX3 and PAX7 or possibly have novel targets. Similar downstream changes may also be involved in embryonal and fusion gene negative cases. Genomic amplification of such genes as MYCN, MDM2, CDK4, and PAX7-FKHR is a feature mainly of the alveolar subtype, while specific chromosomal gains, including chromosomes 2, 8, 12, and 13, are associated with the embryonal subtype. Loss of alleles and imprinting at 11p15.5 and disruption of genes such as IGF2, ATR, PTC, P16, and TP53 have also been implicated in rhabdomyosarcoma development. Whereas there is now a realistic possibility of cure in the majority of cases, there remains a subset that is resistant to multimodality therapy, including high-dose chemotherapy. Characterization of the defining molecular features of tumors that are likely to behave aggressively represents a particular challenge. Current research is leading toward a better understanding of rhabdomyosarcoma tumorigenesis, which may ultimately result in novel therapeutic strategies that increase the overall cure. Genes Chromosomes Cancer 26:275-285, 1999. PMID:10534762

Anderson, J; Gordon, A; Pritchard-Jones, K; Shipley, J

1999-12-01

395

Selection of reference genes for gene expression studies in astrocytomas.  

Science.gov (United States)

This study was aimed to test a panel of six housekeeping genes (GAPDH, HPRT1, POLR2A, RPLP0, ACTB, and H3F) so as to identify and validate the most suitable reference genes for expression studies in astrocytomas. GAPDH was the most stable and HPRT1 was the least stable reference gene. The effect of reference gene selection on quantitative real-time polymerase chain reaction data interpretation was demonstrated, normalizing the expression data of a selected gene of interest. Thus, GAPDH may be recommended for data normalization in gene expression studies in astrocytomas. Nevertheless, a preliminary validation of reference gene stability is required prior to every study. PMID:20849807

Gresner, Sylwia M; Golanska, Ewa; Kulczycka-Wojdala, Dominika; Jaskolski, Dariusz J; Papierz, Wielislaw; Liberski, Pawel P

2011-01-01

396

Stochastic Gene Expression Model Base Gene Regulatory Networks  

Science.gov (United States)

Gene regulatory networks consist of a number of genes and their interactions which regulate expressions of the genes. Along with the development of gene regulatory network studies, computer simulations have become a valuable tool to evaluate complex relationships between genes. Due to the stochastic nature of gene expressions, various stochastic approaches have attracted increasing interest. In this study, we build gene regulatory networks based on a stochastic gene expression model with delicate assumptions such as transcription, translation, DNA-protein, protein-protein associations and time delay for protein activation. Two simple in-silico gene regulatory network models are constructed and monitored their expression profiles reflecting the inhibition and activation of the gene regulations.

Kim, Haseong; Gelenbe, Erol

397

Evidence for homosexuality gene  

Energy Technology Data Exchange (ETDEWEB)

A genetic analysis of 40 pairs of homosexual brothers has uncovered a region on the X chromosome that appears to contain a gene or genes for homosexuality. When analyzing the pedigrees of homosexual males, the researcheres found evidence that the trait has a higher likelihood of being passed through maternal genes. This led them to search the X chromosome for genes predisposing to homosexuality. The researchers examined the X chromosomes of pairs of homosexual brothers for regions of DNA that most or all had in common. Of the 40 sets of brothers, 33 shared a set of five markers in the q28 region of the long arm of the X chromosome. The linkage has a LOD score of 4.0, which translates into a 99.5% certainty that there is a gene or genes in this area that predispose males to homosexuality. The chief researcher warns, however, that this one site cannot explain all instances of homosexuality, since there were some cases where the trait seemed to be passed paternally. And even among those brothers where there was no evidence that the trait was passed paternally, seven sets of brothers did not share the Xq28 markers. It seems likely that homosexuality arises from a variety of causes.

Pool, R.

1993-07-16

398

GeneCards Version 3: the human gene integrator.  

Science.gov (United States)

GeneCards (www.genecards.org) is a comprehensive, authoritative compendium of annotative information about human genes, widely used for nearly 15 years. Its gene-centric content is automatically mined and integrated from over 80 digital sources, resulting in a web-based deep-linked card for each of >73,000 human gene entries, encompassing the following categories: protein coding, pseudogene, RNA gene, genetic locus, cluster and uncategorized. We now introduce GeneCards Version 3, featuring a speedy and sophisticated search engine and a revamped, technologically enabling infrastructure, catering to the expanding needs of biomedical researchers. A key focus is on gene-set analyses, which leverage GeneCards' unique wealth of combinatorial annotations. These include the GeneALaCart batch query facility, which tabulates user-selected annotations for multiple genes and GeneDecks, which identifies similar genes with shared annotations, and finds set-shared annotations by descriptor enrichment analysis. Such set-centric features address a host of applications, including microarray data analysis, cross-database annotation mapping and gene-disorder associations for drug targeting. We highlight the new Version 3 database architecture, its multi-faceted search engine, and its semi-automated quality assurance system. Data enhancements include an expanded visualization of gene expression patterns in normal and cancer tissues, an integrated alternative splicing pattern display, and augmented multi-source SNPs and pathways sections. GeneCards now provides direct links to gene-related research reagents such as antibodies, recombinant proteins, DNA clones and inhibitory RNAs and features gene-related drugs and compounds lists. We also portray the GeneCards Inferred Functionality Score annotation landscape tool for scoring a gene's functional information status. Finally, we delineate examples of applications and collaborations that have benefited from the GeneCards suite. Database URL: www.genecards.org. PMID:20689021

Safran, Marilyn; Dalah, Irina; Alexander, Justin; Rosen, Naomi; Iny Stein, Tsippi; Shmoish, Michael; Nativ, Noam; Bahir, Iris; Doniger, Tirza; Krug, Hagit; Sirota-Madi, Alexandra; Olender, Tsviya; Golan, Yaron; Stelzer, Gil; Harel, Arye; Lancet, Doron

2010-01-01

399

Predicting Gene Ontology Biological Process From Temporal Gene Expression Patterns  

OpenAIRE

The aim of the present study was to generate hypotheses on the involvement of uncharacterized genes in biological processes. To this end, supervised learning was used to analyze microarray-derived time-series gene expression data. Our method was objectively evaluated on known genes using cross-validation and provided high-precision Gene Ontology biological process classifications for 211 of the 213 uncharacterized genes in the data set used. In addition, new roles in biological process were h...

Lægreid, Astrid; Hvidsten, Torgeir R.; Midelfart, Herman; Komorowski, Jan; Sandvik, Arne K.

2003-01-01

400

Gene Circuit Analysis of the Terminal Gap Gene huckebein  

OpenAIRE

The early embryo of Drosophila melanogaster provides a powerful model system to study the role of genes in pattern formation. The gap gene network constitutes the first zygotic regulatory tier in the hierarchy of the segmentation genes involved in specifying the position of body segments. Here, we use an integrative, systems-level approach to investigate the regulatory effect of the terminal gap gene huckebein (hkb) on gap gene expression. We present quantitative expression data for the Hkb p...

Ashyraliyev, Maksat; Siggens, Ken; Janssens, Hilde; Blom, Joke; Akam, Michael; Jaeger, Johannes

2009-01-01

401

Recombination in immunoglobulin gene loci  

OpenAIRE

Gene network of the lymphoid cell differentiation coordinates precisely the recombination process in immunoglobulin gene loci. In our opinion, cellular microRNAs can contribute to the allelic exclusion through microRNA-directed DNA methylation and participate in retargeting recombinases activity from the gene loci of heavy immunoglobulin chains to the gene loci of light chains

Komisarenko S. V.; Halytskiy V. A.

2009-01-01