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Sample records for mycobacterial hsp65 gene

  1. Ubiquitin-fusion degradation pathway: A new strategy for inducing CD8 cells specific for mycobacterial HSP65

    International Nuclear Information System (INIS)

    The ubiquitin-proteasome system (UPS) plays an indispensable role in inducing MHC class I-restricted CD8+ T cells. In this study, we exploited UPS to induce CD8+ T cells specific for mycobacterial HSP65 (mHSP65), one of the leading vaccine candidates against infection with Mycobacterium tuberculosis. A chimeric DNA termed pU-HSP65 encoding a fusion protein between murine ubiquitin and mHSP65 was constructed, and C57BL/6 (B6) mice were immunized with the DNA using gene gun bombardment. Mice immunized with the chimeric DNA acquired potent resistance against challenge with the syngeneic B16F1 melanoma cells transfected with the mHSP65 gene (HSP65/B16F1), compared with those immunized with DNA encoding only mHSP65. Splenocytes from the former group of mice showed a higher grade of cytotoxic activity against HSP65/B16F1 cells and contained a larger number of granzyme B- or IFN-?-producing CD8+ T cells compared with those from the latter group of mice

  2. 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

    Scientific Electronic Library Online (English)

    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

    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. 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.

  3. 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

    Directory of Open Access Journals (Sweden)

    Maria das Graças Motta e Bona

    2011-10-01

    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.

  4. Species-Specific Identification of Mycobacterium leprae by PCR-Restriction Fragment Length Polymorphism Analysis of the hsp65 Gene

    OpenAIRE

    Rastogi, Nalin; Goh, Khye Seng; Berchel, Mylene

    1999-01-01

    PCR-restriction fragment length polymorphism analysis (PRA) of the hsp65 gene present in all mycobacteria was used in the present investigation to characterize Mycobacterium leprae. Bacilli were extracted and purified from different organs from experimentally infected armadillos and nude mice (Swiss mice of nu/nu origin). A total of 15 samples were assayed in duplicate, and the results were compared with those obtained for a total of 147 cultivable mycobacteria representing 34 species. Irresp...

  5. Rapid differentiation of "Mycobacterium canettii" from other Mycobacterium tuberculosis complex organisms by PCR-restriction analysis of the hsp65 gene.

    Science.gov (United States)

    Goh, K S; Legrand, E; Sola, C; Rastogi, N

    2001-10-01

    A total of 102 isolates of the Mycobacterium tuberculosis complex, including available "M. canettii" isolates, were studied by PCR-restriction analysis of a 441-bp fragment of the hsp65 gene. PRA upon HhaI enzyme digestion (GCGC) allowed easy differentiation of "M. canettii" from other members of the M. tuberculosis complex (three bands of 260, 105, and 60 bp for "M. canetti," compared to four bands of 185, 105, 75, and 60 bp for other members of the M. tuberculosis complex). Sequencing of the 441-bp hsp65 fragment of "M. canettii" isolates showed the disappearance of an HhaI site at position 235 due to a C-to-T transition that corresponded to position 631 of the homologous hsp65 gene of M. tuberculosis H37Rv. Considering that "M. canettii" may also exist as a stable rough morphotype, we suggest that the true number of "M. canettii" isolates may be underestimated in clinical microbiology laboratories. PMID:11574597

  6. Cloning and Sequencing of a Part of the Heat Shock Protein 65 Gene (hsp65) of “Tropheryma whippelii” and Its Use for Detection of “T. whippelii” in Clinical Specimens by PCR

    OpenAIRE

    Morgenegg, Silvia; Dutly, Fabrizio; Altwegg, Martin

    2000-01-01

    Using broad-spectrum primers, we have amplified, cloned, and sequenced a 620-bp fragment of the “Tropheryma whippelii” heat shock protein 65 gene (hsp65) from the heart valve of a patient with Whipple's endocarditis. The deduced amino acid sequence shows high similarity to those from actinobacteria, confirming that “T. whippelii” is indeed a member of this phylum. Based on the nucleotide sequence, we have developed a “T. whippelii”-specific seminested PCR. Seventeen patients shown to be posit...

  7. Diffuse meningo-encefalitys due to Nocardia farcinica in a young kidney transplant recipient: identification of the strain using sequencing of hsp65 gene

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    Danila Costa

    2010-09-01

    Full Text Available Genus Nocardia belongs to aerobic Actinomycetes, a wide group of polymorphous Gram+, fixed and acapsusalated, branching, partially acid-fast bacilli. Nocardia organism are ubiquitous, soil-borne actinomycetes that usually infect humans as a result of the inhalation of airborne bacilli or traumatic inoculation. Nocardiosis is a rare opportunistic disease that affects mainly patients with impaired cellmediated immunity, such as those with acquired immunodeficiency syndrome (AIDS or transplant recipients, patients with pulmonary disease, haematological malignancies etc. Pulmonary disease is the most common presentation in immounosuppressed patients. Nocardial organism have a tendency to disseminate hematogenously from the primary site of infection.The central nervous system (CNS is one of the most frequent sites of dissemination. Herein we describe a rare and fatal case of diffuse meningo-encephalytis due to Nocardia farcinica in a young patient kidney transplant recipient. Nocardia has been isolated from Cerebrospinal fluid in a micobacteriology laboratory with identification of the strain using sequencing of hsp65 gene. The diagnosis can be challenging, as signs and symptoms are not specific and a high degree of clinical suspicion is required. Identification of Nocardia farcinica is important because of its aggressiveness, its tendency to disseminate, and its resistance to antibiotics. Susceptibilities to 10 antimicrobial agents were determined by E-test.The isolate was resistant to Gentamicin, Clarithromycin, Doxycicline, Cefotaxime and susceptible to Amikacin,Amoxicillin clavulanate, Imipenem, Ciprofloxacin, Linezolid and Trimethoprim–Sulfamethoxazole. The susceptibility profile was favourable since, in North Italy, the strains are generally resistant to Trimethoprim-Sulfamethoxazole.

  8. High genetic diversity revealed by variable-number tandem repeat genotyping and analysis of hsp65 gene polymorphism in a large collection of "Mycobacterium canettii" strains indicates that the M. tuberculosis complex is a recently emerged clone of "M. canettii".

    Science.gov (United States)

    Fabre, Michel; Koeck, Jean-Louis; Le Flèche, Philippe; Simon, Fabrice; Hervé, Vincent; Vergnaud, Gilles; Pourcel, Christine

    2004-07-01

    We have analyzed, using complementary molecular methods, the diversity of 43 strains of "Mycobacterium canettii" originating from the Republic of Djibouti, on the Horn of Africa, from 1998 to 2003. Genotyping by multiple-locus variable-number tandem repeat analysis shows that all the strains belong to a single but very distant group when compared to strains of the Mycobacterium tuberculosis complex (MTBC). Thirty-one strains cluster into one large group with little variability and five strains form another group, whereas the other seven are more diverged. In total, 14 genotypes are observed. The DR locus analysis reveals additional variability, some strains being devoid of a direct repeat locus and others having unique spacers. The hsp65 gene polymorphism was investigated by restriction enzyme analysis and sequencing of PCR amplicons. Four new single nucleotide polymorphisms were discovered. One strain was characterized by three nucleotide changes in 441 bp, creating new restriction enzyme polymorphisms. As no sequence variability was found for hsp65 in the whole MTBC, and as a single point mutation separates M. tuberculosis from the closest "M. canettii" strains, this diversity within "M. canettii" subspecies strongly suggests that it is the most probable source species of the MTBC rather than just another branch of the MTBC. PMID:15243089

  9. 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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    Joab Trajano Silva

    2008-12-01

    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,

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

    Scientific Electronic Library Online (English)

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

    2007-11-01

    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.

  11. Recurrent nontuberculous mycobacterial endophthalmitis: a diagnostic conundrum

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    Venkateswaran N

    2014-05-01

    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

  12. B cells expressing IL-10 mRNA modulate memory T cells after DNA-Hsp65 immunization.

    Science.gov (United States)

    Fontoura, I C; Trombone, A P F; Almeida, L P; Lorenzi, J C C; Rossetti, R A M; Malardo, T; Padilha, E; Schluchting, W; Silva, R L L; Gembre, A F; Fiuza, J E C; Silva, C L; Panunto-Castelo, A; Coelho-Castelo, A A M

    2015-12-01

    In DNA vaccines, the gene of interest is cloned into a bacterial plasmid that is engineered to induce protein production for long periods in eukaryotic cells. Previous research has shown that the intramuscular immunization of BALB/c mice with a naked plasmid DNA fragment encoding the Mycobacterium leprae 65-kDa heat-shock protein (pcDNA3-Hsp65) induces protection against M. tuberculosis challenge. A key stage in the protective immune response after immunization is the generation of memory T cells. Previously, we have shown that B cells capture plasmid DNA-Hsp65 and thereby modulate the formation of CD8+ memory T cells after M. tuberculosis challenge in mice. Therefore, clarifying how B cells act as part of the protective immune response after DNA immunization is important for the development of more-effective vaccines. The aim of this study was to investigate the mechanisms by which B cells modulate memory T cells after DNA-Hsp65 immunization. C57BL/6 and BKO mice were injected three times, at 15-day intervals, with 100 µg naked pcDNA-Hsp65 per mouse. Thirty days after immunization, the percentages of effector memory T (TEM) cells (CD4+ and CD8+/CD44high/CD62Llow) and memory CD8+ T cells (CD8+/CD44high/CD62Llow/CD127+) were measured with flow cytometry. Interferon ?, interleukin 12 (IL-12), and IL-10 mRNAs were also quantified in whole spleen cells and purified B cells (CD43-) with real-time qPCR. Our data suggest that a B-cell subpopulation expressing IL-10 downregulated proinflammatory cytokine expression in the spleen, increasing the survival of CD4+ TEM cells and CD8+ TEM/CD127+ cells. PMID:26397973

  13. 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)

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

    2008-11-01

    Full Text Available 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.

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

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    Adriana Ignacio de Padua

    2008-11-01

    Full Text Available 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.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.

  15. How M. leprae Hsp65 influences the immune response in genetically selected aged mice?

    Directory of Open Access Journals (Sweden)

    Estevam José Baldon

    2013-03-01

    Full Text Available Heat shock proteins may trigger innate immune responses and are involved in immunosenescence and autoimmunity. We characterized some cellular and humoral alterations after intraperitoneal administration of 2.5µg M. leprae Hsp65 in genetically selected mice for High (HIII or Low (LIII antibody production (9-months-old and in its F1 hybrids. Aged HIII female injected with Hsp65 presented a survival decrease of 42% when compared to untreated group (control; no changes in IgG1 or IgG2a anti-Hsp production were observed in HIII and LIII mice. Regarding the cellular changes, aged HIII female Hsp65-group presented amplified frequency in CD4+CD154+CD28+ cells (p<0.01 and reduced percentage of B and activated CD11c+ cells (p<0.01 in the spleen, and increased percentage of CD11c+ and NKG1A/C/E+ cells (p<0.01 in the blood compared to control. Hsp65 acts like an imbalance trigger: post-injection, the aged F1H female Hsp65-group died 2 months after the first death, as observed in aged HIII females; however there was no statistically significance compared with F1H control group. Furthermore, aged F1H and F1L female showed amplified frequency of naïve T cells and CD11c cells in spleen (p<0.001. In conclusion, our results confirm the sex dichotomy (sex effect of the Hsp65 interference in the immunity of aged mice, becoming evident in females. Next, we will characterize innate immune cells in peritoneal cavity after Hsp65 inoculation; in addition, the role of myeloid-derived suppressor cells will be investigated as these cells are increased during ageing process and have been associated with attenuation of experimental autoimmune diseases. Support: CNPq, FAPESP, INCT-TOX.

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

    International Nuclear Information System (INIS)

    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

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

    Energy Technology Data Exchange (ETDEWEB)

    Rocha, C.D.; Trombone, A.P.F.; Lorenzi, J.C.C.; Almeida, L.P.; Gembre, A.F.; Padilha, E. [Departamento de Bioquímica e Imunologia, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP (Brazil); Ramos, S.G. [Departamento de Patologia, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP (Brazil); Silva, C.L.; Coelho-Castelo, A.A.M. [Departamento de Bioquímica e Imunologia, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP (Brazil)

    2012-09-21

    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.

  18. Epitope specificity and MHC restriction of rheumatoid arthritis synovial T cell clones which recognize a mycobacterial 65 kDa heat shock protein.

    Science.gov (United States)

    Gaston, J S; Life, P F; van der Zee, R; Jenner, P J; Colston, M J; Tonks, S; Bacon, P A

    1991-10-01

    CD4+ T cell clones specific for the mycobacterial hsp 65 were obtained from synovial fluid of a DR4 homozygous rheumatoid arthritis (RA) patient. A stimulatory epitope was defined using both deletion mutants of the mycobacterial hsp 65 and synthetic peptides and proved to be in a highly conserved region of the molecule. Despite this, however, there was no recognition by these clones of either the recombinant human homologue of mycobacterial hsp 65, P60, nor of a synthetic peptide containing an amino acid sequence from P60 corresponding to the epitope defined in the mycobacterial hsp 65. When the pattern of HLA restriction shown by the hsp-65-specific T cell clones was investigated, all clones tested proved to be restricted by HLA-DP rather than the more usual HLA-DR. Inhibition experiments suggested that this restriction also applied to the polyclonal synovial T cell response to hsp 65, but not to other antigens. Exclusive restriction of T cell recognition of an antigen by HLA-DP has not been reported previously, and strongly suggests that in this case the T cell repertoire for recognizing hsp 65 in the context of DR4 is deficient. Such an association between DR4 and the inability to respond to an immunodominant bacterial antigen may have implications for the pathogenesis of RA. PMID:1721835

  19. Diversity of the 32-kilodalton protein gene may form a basis for species determination of potentially pathogenic mycobacterial species.

    OpenAIRE

    Soini, H.; Viljanen, M. K.

    1997-01-01

    In this study, partial gene sequences of the mycobacterial 32-kDa protein gene were determined by PCR-based sequencing. A total of 50 strains representing 18 potentially pathogenic mycobacterial species were studied. In 10 cases, all three strains of the species studied were identical, and intraspecies variability was found in 6 cases. Thus, the 32-kDa protein gene may be a good target for identification of mycobacteria by PCR-based sequencing.

  20. Presence of hsp65 in bacterial extracts (OM-89): a possible mediator of orally-induced tolerance?

    Science.gov (United States)

    Polla, B S; Baladi, S; Fuller, K; Rook, G

    1995-08-16

    Heat shock proteins (HSP) have been implicated in rodent models of autoimmunity, particularly arthritis, and there is suggestive though inconclusive evidence that they may also play a role in human autoimmune disease. The simplest hypothesis is based on molecular mimicry due to the amino-acid sequence homology between mammalian and microbial HSP. Recently OM-89, an extract of several strains of Escherichia coli, has shown some efficacy in the treatment of rheumatoid arthritis (RA) when taken orally. Using species-specific antibodies, we show here that OM-89 contains the 65 kDa HSP (hsp65), while hsp65 was not detected in another bacterial extract containing other microorganisms, including Staphylococcus aureus (OM-85). We suggest that if the human homologue of hsp65 is a relevant target antigen in the human disease, the efficacy of the preparation could be due to induction of oral tolerance or to switching the Th1 response towards Th2. Alternatively, even if the human hsp65 is not a target molecule in RA joints, OM-89 may evoke bystander suppression of joint inflammation via induction of TGF beta-secreting effector cells. These hypotheses should be tested in further studies. PMID:7649235

  1. Hsp65-producing Lactococcus lactis prevents experimental autoimmune encephalomyelitis in mice by inducing CD4+LAP+ regulatory T cells

    OpenAIRE

    Rezende, Rafael M.; Oliveira, Rafael P.; Medeiros, Samara R.; Gomes-Santos, Ana C.; Alves, Andrea C.; Loli, Flávia G.; Guimarães, Mauro A.F.; Amaral, Sylvia S; da Cunha, André P.; Weiner, Howard L.; Azevedo, Vasco; Miyoshi, Anderson; Ana M.C. Faria

    2012-01-01

    Heat shock proteins (Hsps) participate in the cellular response to stress and they are hiperexpressed in inflammatory conditions. They are also known to play a major role in immune modulation, controlling, for instance, autoimmune responses. In this study, we showed that oral administration of a recombinant Lactococcus lactis strain that produces and releases LPS-free Hsp65 prevented the development of experimental autoimmune encephalomyelitis (EAE) in C57BL/6 mice. This was confirmed by the ...

  2. Identification and functional annotation of mycobacterial septum formation genes using cell division mutants of Escherichia coli.

    Science.gov (United States)

    Gaiwala Sharma, Sujata S; Kishore, Vimal; Raghunand, Tirumalai R

    2016-01-01

    The major virulence trait of Mycobacterium tuberculosis is its ability to enter a latent state in the face of robust host immunity. Clues to the molecular basis of latency can emerge from understanding the mechanism of cell division, beginning with identification of proteins involved in this process. Using complementation of Escherichia coli mutants, we functionally annotated M. tuberculosis and Mycobacterium smegmatis homologs of divisome proteins FtsW and AmiC. Our results demonstrate that E. coli can be used as a surrogate model to discover mycobacterial cell division genes, and should prove invaluable in delineating the mechanisms of this fundamental process in mycobacteria. PMID:26577656

  3. Administration of Mycobacterium leprae rHsp65 Aggravates Experimental Autoimmune Uveitis in Mice

    OpenAIRE

    Marengo, Eliana B.; Commodaro, Alessandra Gonçalves; Jean Pierre S. Peron; de Moraes, Luciana V.; Portaro, Fernanda C. V.; Belfort, Rubens; Rizzo, Luiz Vicente; Sant'Anna, Osvaldo Augusto

    2009-01-01

    The 60kDa heat shock protein family, Hsp60, constitutes an abundant and highly conserved class of molecules that are highly expressed in chronic-inflammatory and autoimmune processes. Experimental autoimmune uveitis [EAU] is a T cell mediated intraocular inflammatory disease that resembles human uveitis. Mycobacterial and homologous Hsp60 peptides induces uveitis in rats, however their participation in aggravating the disease is poorly known. We here evaluate the effects of the Mycobacterium ...

  4. Identifying novel mycobacterial stress associated genes using a random mutagenesis screen in Mycobacterium smegmatis.

    Science.gov (United States)

    Viswanathan, Gopinath; Joshi, Shrilaxmi V; Sridhar, Aditi; Dutta, Sayantanee; Raghunand, Tirumalai R

    2015-12-10

    Cell envelope associated components of Mycobacterium tuberculosis (M.tb) have been implicated in stress response, immune modulation and in vivo survival of the pathogen. Although many such factors have been identified, there is a large disparity between the number of genes predicted to be involved in functions linked to the envelope and those described in the literature. To identify and characterise novel stress related factors associated with the mycobacterial cell envelope, we isolated colony morphotype mutants of Mycobacterium smegmatis (M. smegmatis), based on the hypothesis that mutants with unusual colony morphology may have defects in the biosynthesis of cell envelope components. On testing their susceptibility to stress conditions relevant to M.tb physiology, multiple mutants were found to be sensitive to Isoniazid, Diamide and H2O2, indicative of altered permeability due to changes in cell envelope composition. Two mutants showed defects in biofilm formation implying possible roles for the target genes in antibiotic tolerance and/or virulence. These assays identified novel stress associated roles for several mycobacterial genes including sahH, tatB and aceE. Complementation analysis of selected mutants with the M. smegmatis genes and their M.tb homologues showed phenotypic restoration, validating their link to the observed phenotypes. A mutant carrying an insertion in fhaA encoding a forkhead associated domain containing protein, showed reduced survival in THP-1 macrophages, providing in vivo validation to this screen. Taken together, these results suggest that the M.tb homologues of a majority of the identified genes may play significant roles in the pathogenesis of tuberculosis. PMID:26211627

  5. Mycobacterial Diseases

    Science.gov (United States)

    ... countries, non-tuberculous mycobacterial infections occur worldwide. NIAID Role in Research NIAID research on mycobacteria is leading ... Accessibility Privacy Policy Disclaimer Website Links & Policies FOIA Employee Emergency Information RELATED GOVERNMENT SITES U.S. Department of ...

  6. Dimerization of an Immunoactivating Peptide Derived from Mycobacterial hsp65 Using N-Hydroxysuccinimide Based Bifunctional Reagents Is Critical for Its Antitumor Properties.

    Czech Academy of Sciences Publication Activity Database

    Bezouška, Karel; Kubínková, Z.; St?íbrný, J.; Volfová, B.; Pompach, Petr; Kuzma, Marek; Šírová, Milada; ?íhová, Blanka

    2012-01-01

    Ro?. 23, ?. 10 (2012), s. 2032-2041. ISSN 1043-1802 R&D Projects: GA MŠk 1M0505; GA ?R GA303/09/0477; GA ?R GD305/09/H008 Institutional research plan: CEZ:AV0Z50200510 Keywords : HUMAN EOSINOPHILS * KILLER-CELLS * CD69 Subject RIV: CE - Biochemistry Impact factor: 4.580, year: 2012

  7. Enhanced immune response of a bicistronic DNA vaccine expressing fusion antigen Hsp65-Esat-6 of Mycobacterium Tuberculosis with GM-CSF as a molecular adjuvant

    Scientific Electronic Library Online (English)

    Yan, Dong; Jun-Yuan, Gong; Xin, Liu; Jun-Wu, Li.

    2013-10-01

    Full Text Available This study aimed to construct a bicistronic DNA vaccine expressing fusion antigen Hsp65-Esat-6 of Mycobacterium tuberculosis with cytokine GM-CSF as a molecular adjuvant (pIRES-Hsp65-ESAT-6-GM-CSF, pIRHEG), and the immune response in mice. C57BL/6 mice were immunized with the recombinant plasmid to [...] detect the titer of antibodies, lymphocyte proliferation, the ratio of CD4+, CD8+T cell and IFN ~ ?,IL-2 secretion. The titer of antibody, lymphocyte proliferation, the ratio of CD4+T and CD8+T cells and IFN ~ ?, IL-2 secretion of pIRHEG group was significant higher than other recombinant plasmid groups, which significant differed by statistical mean. The bicistronic DNA vaccine could induce an effective immune response in mice and could be used as vital ingredient of a new tuberculosis vaccine candidate.

  8. Macro-array and bioinformatic analyses reveal mycobacterial 'core' genes, variation in the ESAT-6 gene family and new phylogenetic markers for the Mycobacterium tuberculosis complex.

    Science.gov (United States)

    Marmiesse, Magali; Brodin, Priscille; Buchrieser, Carmen; Gutierrez, Christina; Simoes, Nathalie; Vincent, Veronique; Glaser, Philippe; Cole, Stewart T; Brosch, Roland

    2004-02-01

    To better understand the biology and the virulence determinants of the two major mycobacterial human pathogens Mycobacterium tuberculosis and Mycobacterium leprae, their genome sequences have been determined recently. In silico comparisons revealed that among the 1439 genes common to both M. tuberculosis and M. leprae, 219 genes code for proteins that show no similarity with proteins from other organisms. Therefore, the latter 'core' genes could be specific for mycobacteria or even for the intracellular mycobacterial pathogens. To obtain more information as to whether these genes really were mycobacteria-specific, they were included in a focused macro-array, which also contained genes from previously defined regions of difference (RD) known to be absent from Mycobacterium bovis BCG relative to M. tuberculosis. Hybridization of DNA from 40 strains of the M. tuberculosis complex and in silico comparison of these genes with the near-complete genome sequences from Mycobacterium avium, Mycobacterium marinum and Mycobacterium smegmatis were undertaken to answer this question. The results showed that among the 219 conserved genes, very few were not present in all the strains tested. Some of these missing genes code for proteins of the ESAT-6 family, a group of highly immunogenic small proteins whose presence and number is variable among the genomically highly conserved members of the M. tuberculosis complex. Indeed, the results suggest that, with few exceptions, the 'core' genes conserved among M. tuberculosis H37Rv and M. leprae are also highly conserved among other mycobacterial strains, which makes them interesting potential targets for developing new specific anti-mycobacterial drugs. In contrast, the genes from RD regions showed great variability among certain members of the M. tuberculosis complex, and some new specific deletions in Mycobacterium canettii, Mycobacterium microti and seal isolates were identified and further characterized during this study. Together with the distribution of a particular 6 or 7 bp micro-deletion in the gene encoding the polyketide synthase pks15/1, these results confirm and further extend the revised phylogenetic model for the M. tuberculosis complex recently presented. PMID:14766927

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

    OpenAIRE

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

    2003-01-01

    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...

  10. High-Resolution Phenotypic Profiling Defines Genes Essential for Mycobacterial Growth and Cholesterol Catabolism

    OpenAIRE

    Griffin, Jennifer E.; Gawronski, Jeffrey D.; DeJesus, Michael A.; Ioerger, Thomas R; Akerley, Brian J.; Sassetti, Christopher M.

    2011-01-01

    The pathways that comprise cellular metabolism are highly interconnected, and alterations in individual enzymes can have far-reaching effects. As a result, global profiling methods that measure gene expression are of limited value in predicting how the loss of an individual function will affect the cell. In this work, we employed a new method of global phenotypic profiling to directly define the genes required for the growth of Mycobacterium tuberculosis. A combination of high-density mutagen...

  11. 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)

    Joan Joseph

    2010-01-01

    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. α-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.

  12. Mycobacterial Pathogenomics and Evolution.

    Science.gov (United States)

    Bottai, Daria; Stinear, Timothy P; Supply, Philip; Brosch, Roland

    2014-02-01

    Most mycobacterial species are harmless saprophytes, often found in aquatic environments. A few species seem to have evolved from this pool of environmental mycobacteria into major human pathogens, such as Mycobacterium tuberculosis, the agent of tuberculosis, Mycobacterium leprae, the leprosy bacillus, and Mycobacterium ulcerans, the agent of Buruli ulcer. While the pathogenicity of M. ulcerans relates to the acquisition of a large plasmid encoding a polyketide-derived toxin, the molecular mechanisms by which M. leprae or M. tuberculosis have evolved to cause disease are complex and involve the interaction between the pathogen and the host. Here we focus on M. tuberculosis and closely related mycobacteria and discuss insights gained from recent genomic and functional studies. Comparison of M. tuberculosis genome data with sequences from nontuberculous mycobacteria, such as Mycobacterium marinum or Mycobacterium kansasii, provides a perception of the more distant evolution of M. tuberculosis, while the recently accomplished genome sequences of multiple tubercle bacilli with smooth colony morphology, named Mycobacterium canettii, have allowed the ancestral gene pool of tubercle bacilli to be estimated. The resulting findings are instrumental for our understanding of the pathogenomic evolution of tuberculosis-causing mycobacteria. Comparison of virulent and attenuated members of the M. tuberculosis complex has further contributed to identification of a specific secretion pathway, named ESX or Type VII secretion. The molecular machines involved are key elements for mycobacterial pathogenicity, strongly influencing the ability of M. tuberculosis to cope with the immune defense mounted by the host. PMID:26082120

  13. Genetic susceptibility to common mycobacterial diseases

    OpenAIRE

    Wong, Hei Sunny; Hill, Adrian Vivian Sinton

    2010-01-01

    Common mycobacterial diseases, including tuberculosis and leprosy, contribute to major mortality and morbidity worldwide. Despite evidence of an important role of host genetic factors in susceptibility to these infections, few compelling genetic associations have been identified with previous candidate gene and linkage approaches.This thesis investigates the genetic factors of human immunity to these mycobacterial diseases using a high-throughput approach of association testing. To assess gen...

  14. Release of mycobacterial antigens.

    Science.gov (United States)

    Majlessi, Laleh; Prados-Rosales, Rafael; Casadevall, Arturo; Brosch, Roland

    2015-03-01

    Mycobacterium tuberculosis has evolved from a Mycobacterium canettii-like progenitor pool into one of the most successful and widespread human pathogens. The pathogenicity of M. tuberculosis is linked to its ability to secrete/export/release selected mycobacterial proteins, and it is also established that active release of mycobacterial antigens is a prerequisite for strong immune recognition. Recent research has enabled mycobacterial secretion systems and vesicle-based release of mycobacterial antigens to be elucidated, which together with host-related specificities constitute key variables that determine the outcome of infection. Here, we discuss recently discovered, novel aspects on the nature and the regulation of antigen release of the tuberculosis agent with particular emphasis on the biological characterization of mycobacteria-specific ESX/type VII secretion systems and their secreted proteins, belonging to the Esx, PE, and PPE categories. The importance of specific mycobacterial antigen release is probably best exemplified by the striking differences observed between the cellular events during infection with the ESX-1-deficient, attenuated Mycobacterium bovis BCG compared to the virulent M. tuberculosis, which are clearly important for design of more specific diagnostics and more efficient vaccines. PMID:25703550

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

    OpenAIRE

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

    1999-01-01

    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...

  16. [Biologics and mycobacterial diseases].

    Science.gov (United States)

    Tsuyuguchi, Kazunari; Matsumoto, Tomoshige

    2013-03-01

    Various biologics such as TNF-alpha inhibitor or IL-6 inhibitor are now widely used for treatment of rheumatoid arthritis. Many reports suggested that one of the major issues is high risk of developing tuberculosis (TB) associated with using these agents, which is especially important in Japan where tuberculosis still remains endemic. Another concern is the risk of development of nontuberculous mycobacterial (NTM) diseases and we have only scanty information about it. The purpose of this symposium is to elucidate the role of biologics in the development of mycobacterial diseases and to establish the strategy to control them. First, Dr. Tohma showed the epidemiologic data of TB risks associated with using biologics calculated from the clinical database on National Database of Rheumatic Diseases by iR-net in Japan. He estimated TB risks in rheumatoid arthritis (RA) patients to be about four times higher compared with general populations and to become even higher by using biologics. He also pointed out a low rate of implementation of QuantiFERON test (QFT) as screening test for TB infection. Next, Dr. Tokuda discussed the issue of NTM disease associated with using biologics. He suggested the airway disease in RA patients might play some role in the development of NTM disease, which may conversely lead to overdiagnosis of NTM disease in RA patients. He suggested that NTM disease should not be uniformly considered a contraindication to treatment with biologics, considering from the results of recent multicenter study showing relatively favorable outcome of NTM patients receiving biologics. Patients with latent tuberculosis infection (LTBI) should receive LTBI treatment before starting biologics. Dr. Kato, a chairperson of the Prevention Committee of the Japanese Society for Tuberculosis, proposed a new LTBI guideline including active implementation of LTBI treatment, introducing interferon gamma release assay, and appropriate selection of persons at high risk for developing TB. Lastly, Dr. Matsumoto stressed the risk of discontinuing TNF-alpha inhibitor during treatment for tuberculosis. He showed from his clinical experience that TNF-alpha inhibitor can be safely used in active TB patient receiving effective antituberculosis chemotherapy and it is even more effective for prevention of paradoxical response. Active discussion was done about the four topics, including the matter beyond present guidelines. We hope these discussions will form the basis for the establishment of new guideline for the management of mycobacterial disease when using immunosuppressive agents including biologics. 1. The risk of developing tuberculosis (TB) and situations of screening for TB risk at administration of biologics-the case of rheumatoid arthritis: Shigeto TOHMA (Clinical Research Center for Allergy and Rheumatology, National Hospital Organization Sagamihara National Hospital) We calculated the standardized incidence ratio (SIR) of TB from the clinical data on National Database of Rheumatic Diseases by iR-net in Japan (NinJa) and compared with the SIR of TB from the data of the post-marketing surveillances of five biologics. Among 43584 patient-years, forty patients developed TB. The SIR of TB in NinJa was 4.34 (95%CI: 3.00-5.69). According to the post-marketing surveillances of 5 biologics, the SIR of TB were 3.62-34.4. The incidence of TB in patients with RA was higher than general population in Japan, and was increased more by some biologics. We have to recognize the risk of TB when we start biologics therapy to patients with RA. Although the frequency of implementation of QuantiFERON test (QFT) had gradually increased, it was still limited to 41%. In order to predict the risk of developing TB and to prevent TB, it might be better to check all RA patients by QFT at time time of biologics administration. 2. Biologics and nontuberculous mycobacterial diseases: Hitoshi TOKUDA (Social Insurance Central General Hospital) Several topics about the relationship between RA and nontuberculous mycobacterial (NTM) diseases were discussed, which is sti

  17. Mycobacterial codon optimization of the gene encoding the Sm14 antigen of Schistosoma mansoni in recombinant Mycobacterium bovis Bacille Calmette-Guérin enhances protein expression but not protection against cercarial challenge in mice.

    Science.gov (United States)

    Varaldo, Paula B; Miyaji, Eliane N; Vilar, Monica M; Campos, Adriano S; Dias, Waldely O; Armôa, Geraldo R G; Tendler, Miriam; Leite, Luciana C C; McIntosh, Douglas

    2006-10-01

    A mycobacterial codon-optimized gene encoding the Sm14 antigen of Schistosoma mansoni was generated using oligonucleotide assembly. This synthetic gene enhanced approximately fourfold the protein expression level in recombinant Mycobacterium bovis Bacille Calmette-Guérin (rBCG) when compared to that obtained using the native gene in the same expression vector. Immunization of mice with rBCG expressing Sm14 via the synthetic gene induced specific cellular Th1-predominant immune responses, as determined by interferon-gamma production of Sm14-stimulated splenocytes, which were comparable to those recorded in animals immunized with an rBCG strain expressing the native gene. Administration of a single dose of the rBCG-Sm14 construct carrying the synthetic gene conferred protection against cercarial challenge in outbred Swiss mice, at a level equivalent to those provided by either a single dose of rBCG expressing the native gene or three doses of Escherichia coli-derived recombinant Sm14. Our data demonstrated that despite improving the level of antigen expression, the codon optimization strategy did not result in enhanced immunity or protection against cercarial S. mansoni challenge. PMID:16965361

  18. Breast mycobacterial infection in a haemodialysis patient.

    OpenAIRE

    Modai, D.; Weissgarten, J.; Reiff, R.; Siegal, B.; Gabizon, D.

    1984-01-01

    Mycobacterial infection is relatively common among patients maintained on haemodialysis and may present in uncommon locations and acquire an unusual course. We present a patient in whom a breast mass was found to be caused by primary mycobacterial infection. This is to our knowledge the first report on breast mycobacterial infection in a haemodialysis patient.

  19. The cytopathology of mycobacterial infection.

    Science.gov (United States)

    Michelow, Pamela; Omar, Tanvier; Field, Andrew; Wright, Colleen

    2016-03-01

    Mycobacterial infection, tuberculosis (TB) in particular, remains one of the world's deadliest communicable diseases in adults and particularly in children, in low and middle income countries. The combination of human immunodeficiency virus (HIV) and TB is often lethal with TB accounting for 25% of deaths in the HIV population. One of the cornerstones for reducing the TB epidemic is early case detection using high quality diagnostic techniques. Cytology, especially fine needle aspiration biopsy (FNAB) is able to diagnose mycobacterial infection in a rapid and cost-effective manner without requiring surgery, thus allowing appropriate management to be quickly instituted. Confirmatory ancillary tests can effectively be performed on cytologic material. In this review, the pertinent cytomorphology of mycobacterial infection in various exfoliative and FNAB specimens is presented, in both immunocompetent and immunosuppressed patients. In the immunosuppressed, the typical cytomorphology of caseating granulomatous inflammation may not be seen but suppurative necrotic inflammation, mycobacterial spindle pseudotumour or a specimen comprised entirely of necrosis may be seen instead. This review includes discussion of currently available ancillary tests that can be performed on cytologic specimens. Diagn. Cytopathol. 2016;44:255-262. © 2016 Wiley Periodicals, Inc. PMID:26800030

  20. Murine T-lymphocyte activation by mycobacterial antigens

    International Nuclear Information System (INIS)

    There has been renewed interest in the diagnosis of tuberculosis and other mycobacterial infections in the United States. Effective immunity to mycobacterial infections, as well as diagnosis by the skin test, involves T-cells rather than antibodies. Studies currently underway use the new technologies of monoclonal antibodies and recombinant DNA to define better mycobacterial antigens for T-cell activation, in the hope of identifying species specific antigens. Lymph node cells from mice sensitized to Mycobacterium intracellulare and Mycobacterium avium were assayed for activation by mycobacterial fractions, and cell lines and clones were generated. Comparing BALB/c and B10 mice indicated better responses to M. avium sonicate by B10 mice. A recombinant gene product containing a M. intracellulare peptide was assayed with lymph node cells and indicated excellent T-cell stimulation in BALB/c lymph node cells and cell lines. However, assays using B10 T-cell clones have yet to detect responders to the recombinant protein. Future studies using synthetic epitopes produced by recombinant DNA techniques and defined by monoclonal antibodies are necessary for the identification of reactive T-cell epitopes that are potentially species specific. 4 refs, 7 figs, 1 tab

  1. Phenotypic heterogeneity in mycobacterial stringent response

    Directory of Open Access Journals (Sweden)

    Bose Indrani

    2011-01-01

    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.

  2. Cutaneous mycobacterial spindle cell pseudotumour

    OpenAIRE

    Tan, Geok Chin; Yap, Yen Piow; Shiran, Mohd Sidik; Sabariah, Abdul Rahman; PATHMANATHAN, RAJADURAI

    2009-01-01

    Mycobacterial spindle cell pseudotumour (MSCP) has been reported in various sites, including skin, lymph nodes, bone marrow, lung and spleen. Cutaneous lesions are extremely rare and the differential diagnoses include various spindle cell lesions. Literature review shows that this lesion has preponderance for upper limb involvement and occurs largely in immunosuppressed individuals. We report a case of MSCP of the skin due to atypical mycobacterium and discuss the risk of misdiagnosis as a sa...

  3. TNF-? modulates TLR2-dependent responses during mycobacterial infection.

    Science.gov (United States)

    Holla, Sahana; Trinath, Jamma; Balaji, Kithiganahalli Narayanaswamy

    2014-01-01

    Multifunctional roles of tumor necrosis factor-alpha (TNF-?) during the mycobacterial pathogenesis make it an important molecule to understand and to examine the course of infection. Identification and analysis of TNF-? response can largely contribute to determine the potential host mediators for therapeutic intervention against tuberculosis. The current chapter describes several methods to assess the ability of TNF-? signaling to modulate toll-like receptor (TLR)2 signaling, another key player in mycobacterial infection and its responses. Experiments involving neutralizing antibodies, antagonists, pharmacological inhibitors, and siRNA-mediated gene silencing are discussed in this chapter to establish the role of TNF-? signaling. The widely used protein and mRNA analysis readouts like enzyme-linked immunosorbent assay (ELISA), immunoblotting, fluorescence-activated cell sorting (FACS), and quantitative real-time RT-PCR are useful to estimate and confirm the mediators involved in TNF-? and TLR2 signaling. PMID:24788179

  4. Mycobacterial mutants with defective control of phagosomal acidification.

    Directory of Open Access Journals (Sweden)

    2005-11-01

    Full Text Available The pathogenesis of mycobacterial infection is associated with an ability to interfere with maturation of the phagosomal compartment after ingestion by macrophages. Identification of the mycobacterial components that contribute to this phenomenon will allow rational design of novel approaches to the treatment and prevention of tuberculosis. Microarray-based screening of a transposon library was used to identify mutations that influence the fate of Mycobacterium bovis bacille Calmette-Guérin (BCG following uptake by macrophages. A screen based on bacterial survival during a 3-d infection highlighted genes previously implicated in growth of Mycobacterium tuberculosis in macrophages and in mice, together with a number of other virulence genes including a locus encoding virulence-associated membrane proteins and a series of transporter molecules. A second screen based on separation of acidified and non-acidified phagosomes by flow cytometry identified genes involved in mycobacterial control of early acidification. This included the KefB potassium/proton antiport. Mutants unable to control early acidification were significantly attenuated for growth during 6-d infections of macrophages. Early acidification of the phagosome is associated with reduced survival of BCG in macrophages. A strong correlation exists between genes required for intracellular survival of BCG and those required for growth of M. tuberculosis in mice. In contrast, very little correlation exists between genes required for intracellular survival of BCG and those that are up-regulated during intracellular adaptation of M. tuberculosis. This study has identified targets for interventions to promote immune clearance of tuberculosis infection. The screening technologies demonstrated in this study will be useful to the study of pathogenesis in many other intracellular microorganisms.

  5. Leaderless Transcripts and Small Proteins Are Common Features of the Mycobacterial Translational Landscape

    Science.gov (United States)

    Lapierre, Pascal; Mir, Mushtaq; Chase, Michael R.; Pyle, Margaret M.; Gawande, Richa; Ahmad, Rushdy; Sarracino, David A.; Ioerger, Thomas R.; Fortune, Sarah M.; Derbyshire, Keith M.; Wade, Joseph T.; Gray, Todd A.

    2015-01-01

    RNA-seq technologies have provided significant insight into the transcription networks of mycobacteria. However, such studies provide no definitive information on the translational landscape. Here, we use a combination of high-throughput transcriptome and proteome-profiling approaches to more rigorously understand protein expression in two mycobacterial species. RNA-seq and ribosome profiling in Mycobacterium smegmatis, and transcription start site (TSS) mapping and N-terminal peptide mass spectrometry in Mycobacterium tuberculosis, provide complementary, empirical datasets to examine the congruence of transcription and translation in the Mycobacterium genus. We find that nearly one-quarter of mycobacterial transcripts are leaderless, lacking a 5’ untranslated region (UTR) and Shine-Dalgarno ribosome-binding site. Our data indicate that leaderless translation is a major feature of mycobacterial genomes and is comparably robust to leadered initiation. Using translational reporters to systematically probe the cis-sequence requirements of leaderless translation initiation in mycobacteria, we find that an ATG or GTG at the mRNA 5’ end is both necessary and sufficient. This criterion, together with our ribosome occupancy data, suggests that mycobacteria encode hundreds of small, unannotated proteins at the 5’ ends of transcripts. The conservation of small proteins in both mycobacterial species tested suggests that some play important roles in mycobacterial physiology. Our translational-reporter system further indicates that mycobacterial leadered translation initiation requires a Shine Dalgarno site in the 5’ UTR and that ATG, GTG, TTG, and ATT codons can robustly initiate translation. Our combined approaches provide the first comprehensive view of mycobacterial gene structures and their non-canonical mechanisms of protein expression. PMID:26536359

  6. Leaderless Transcripts and Small Proteins Are Common Features of the Mycobacterial Translational Landscape.

    Science.gov (United States)

    Shell, Scarlet S; Wang, Jing; Lapierre, Pascal; Mir, Mushtaq; Chase, Michael R; Pyle, Margaret M; Gawande, Richa; Ahmad, Rushdy; Sarracino, David A; Ioerger, Thomas R; Fortune, Sarah M; Derbyshire, Keith M; Wade, Joseph T; Gray, Todd A

    2015-11-01

    RNA-seq technologies have provided significant insight into the transcription networks of mycobacteria. However, such studies provide no definitive information on the translational landscape. Here, we use a combination of high-throughput transcriptome and proteome-profiling approaches to more rigorously understand protein expression in two mycobacterial species. RNA-seq and ribosome profiling in Mycobacterium smegmatis, and transcription start site (TSS) mapping and N-terminal peptide mass spectrometry in Mycobacterium tuberculosis, provide complementary, empirical datasets to examine the congruence of transcription and translation in the Mycobacterium genus. We find that nearly one-quarter of mycobacterial transcripts are leaderless, lacking a 5' untranslated region (UTR) and Shine-Dalgarno ribosome-binding site. Our data indicate that leaderless translation is a major feature of mycobacterial genomes and is comparably robust to leadered initiation. Using translational reporters to systematically probe the cis-sequence requirements of leaderless translation initiation in mycobacteria, we find that an ATG or GTG at the mRNA 5' end is both necessary and sufficient. This criterion, together with our ribosome occupancy data, suggests that mycobacteria encode hundreds of small, unannotated proteins at the 5' ends of transcripts. The conservation of small proteins in both mycobacterial species tested suggests that some play important roles in mycobacterial physiology. Our translational-reporter system further indicates that mycobacterial leadered translation initiation requires a Shine Dalgarno site in the 5' UTR and that ATG, GTG, TTG, and ATT codons can robustly initiate translation. Our combined approaches provide the first comprehensive view of mycobacterial gene structures and their non-canonical mechanisms of protein expression. PMID:26536359

  7. Targeting Mycobacterial Enzymes with Natural Products.

    Science.gov (United States)

    Sieniawska, Elwira

    2015-10-22

    Tuberculosis (TB) is a recurring threat to contemporary civilization. It affects not only those within developing countries, but has also appeared again in places where it was once considered eradicated. TB co-infection in patients infected by HIV is, at the time of writing, the most common cause of death. In the field of searching for new antimycobacterial drug leads, compounds of natural origin still remain a promising source. The review is intended to gather information about natural products (metabolites of plants, fungi, bacteria, and marine sponges) that show activity against mycobacterial enzymes. Here, natural metabolites are presented as being inhibitors/activators of the mycobacterial enzymes involved in mycobacterial growth in vitro (ClpC1, ClpP, MurE ligase, mycothiol S-conjugate amidase, ?-ketoacyl-ACP synthase, InhA) and in vivo, as regards the host cell (PtpB). Each enzyme is briefly described so as to generate an understanding of its role in mycobacterial growth and engender a perception of the mechanism of action of the studied natural compounds. Furthermore, after the introduction of the enzyme, its inhibitors are listed and exactly characterized. PMID:26441042

  8. The essential role of SepF in mycobacterial division.

    Science.gov (United States)

    Gola, Susanne; Munder, Thomas; Casonato, Stefano; Manganelli, Riccardo; Vicente, Miguel

    2015-08-01

    Mycobacteria lack several of the components that are essential in model systems as Escherichia coli or Bacillus subtilis for the formation of the divisome, a ring-like structure assembling at the division site to initiate bacterial cytokinesis. Divisome assembly depends on the correct placement of the FtsZ protein into a structure called the Z ring. Notably, early division proteins that assist in the localisation of the Z ring to the cytoplasmic membrane and modulate its structure are missing in the so far known mycobacterial cell division machinery. To find mycobacterium-relevant components of the divisome that might act at the level of FtsZ, a yeast two-hybrid screening was performed with FtsZ from Mycobacterium tuberculosis. We identified the SepF homolog as a new interaction partner of mycobacterial FtsZ. Depending on the presence of FtsZ, SepF-GFP fusions localised in ring-like structures at potential division sites. Alteration of SepF levels in Mycobacterium smegmatis led to filamentous cells, indicating a division defect. Depletion of SepF resulted in a complete block of division. The sepF gene is highly conserved in the M.?tuberculosis complex members. We therefore propose that SepF is an essential part of the core division machinery in the genus Mycobacterium. PMID:25943244

  9. Simple sequence repeats in mycobacterial genomes

    Indian Academy of Sciences (India)

    Vattipally B Sreenu; Pankaj Kumar; Javaregowda Nagaraju; Hampapathalu A Nagarajaram

    2007-01-01

    Simple sequence repeats (SSRs) or microsatellites are the repetitive nucleotide sequences of motifs of length 1–6 bp. They are scattered throughout the genomes of all the known organisms ranging from viruses to eukaryotes. Microsatellites undergo mutations in the form of insertions and deletions (INDELS) of their repeat units with some bias towards insertions that lead to microsatellite tract expansion. Although prokaryotic genomes derive some plasticity due to microsatellite mutations they have in-built mechanisms to arrest undue expansions of microsatellites and one such mechanism is constituted by post-replicative DNA repair enzymes MutL, MutH and MutS. The mycobacterial genomes lack these enzymes and as a null hypothesis one could expect these genomes to harbour many long tracts. It is therefore interesting to analyse the mycobacterial genomes for distribution and abundance of microsatellites tracts and to look for potentially polymorphic microsatellites. Available mycobacterial genomes, Mycobacterium avium, M. leprae, M. bovis and the two strains of M. tuberculosis (CDC1551 and H37Rv) were analysed for frequencies and abundance of SSRs. Our analysis revealed that the SSRs are distributed throughout the mycobacterial genomes at an average of 220–230 SSR tracts per kb. All the mycobacterial genomes contain few regions that are conspicuously denser or poorer in microsatellites compared to their expected genome averages. The genomes distinctly show scarcity of long microsatellites despite the absence of a post-replicative DNA repair system. Such severe scarcity of long microsatellites could arise as a result of strong selection pressures operating against long and unstable sequences although influence of GC-content and role of point mutations in arresting microsatellite expansions can not be ruled out. Nonetheless, the long tracts occasionally found in coding as well as non-coding regions may account for limited genome plasticity in these genomes.

  10. Octanoylation of early intermediates of mycobacterial methylglucose lipopolysaccharides

    Science.gov (United States)

    Maranha, Ana; Moynihan, Patrick J.; Miranda, Vanessa; Correia Lourenço, Eva; Nunes-Costa, Daniela; Fraga, Joana S.; José Barbosa Pereira, Pedro; Macedo-Ribeiro, Sandra; Ventura, M. Rita; Clarke, Anthony J.; Empadinhas, Nuno

    2015-01-01

    Mycobacteria synthesize unique intracellular methylglucose lipopolysaccharides (MGLP) proposed to modulate fatty acid metabolism. In addition to the partial esterification of glucose or methylglucose units with short-chain fatty acids, octanoate was invariably detected on the MGLP reducing end. We have identified a novel sugar octanoyltransferase (OctT) that efficiently transfers octanoate to glucosylglycerate (GG) and diglucosylglycerate (DGG), the earliest intermediates in MGLP biosynthesis. Enzymatic studies, synthetic chemistry, NMR spectroscopy and mass spectrometry approaches suggest that, in contrast to the prevailing consensus, octanoate is not esterified to the primary hydroxyl group of glycerate but instead to the C6 OH of the second glucose in DGG. These observations raise important new questions about the MGLP reducing end architecture and about subsequent biosynthetic steps. Functional characterization of this unique octanoyltransferase, whose gene has been proposed to be essential for M. tuberculosis growth, adds new insights into a vital mycobacterial pathway, which may inspire new drug discovery strategies. PMID:26324178

  11. Intranasal vaccination with messenger RNA as a new approach in gene therapy: Use against tuberculosis

    Directory of Open Access Journals (Sweden)

    Silva Aristóbolo M

    2010-10-01

    Full Text Available Abstract Background mRNAs are highly versatile, non-toxic molecules that are easy to produce and store, which can allow transient protein expression in all cell types. The safety aspects of mRNA-based treatments in gene therapy make this molecule one of the most promising active components of therapeutic or prophylactic methods. The use of mRNA as strategy for the stimulation of the immune system has been used mainly in current strategies for the cancer treatment but until now no one tested this molecule as vaccine for infectious disease. Results We produce messenger RNA of Hsp65 protein from Mycobacterium leprae and show that vaccination of mice with a single dose of 10 ?g of naked mRNA-Hsp65 through intranasal route was able to induce protection against subsequent challenge with virulent strain of Mycobacterium tuberculosis. Moreover it was shown that this immunization was associated with specific production of IL-10 and TNF-alpha in spleen. In order to determine if antigen presenting cells (APCs present in the lung are capable of capture the mRNA, labeled mRNA-Hsp65 was administered by intranasal route and lung APCs were analyzed by flow cytometry. These experiments showed that after 30 minutes until 8 hours the populations of CD11c+, CD11b+ and CD19+ cells were able to capture the mRNA. We also demonstrated in vitro that mRNA-Hsp65 leads nitric oxide (NO production through Toll-like receptor 7 (TLR7. Conclusions Taken together, our results showed a novel and efficient strategy to control experimental tuberculosis, besides opening novel perspectives for the use of mRNA in vaccines against infectious diseases and clarifying the mechanisms involved in the disease protection we noticed as well.

  12. Mycobacterial Pan-Genome Analysis Suggests Important Role of Plasmids in the Radiation of Type VII Secretion Systems

    Science.gov (United States)

    Dumas, Emilie; Christina Boritsch, Eva; Vandenbogaert, Mathias; Rodríguez de la Vega, Ricardo C.; Thiberge, Jean-Michel; Caro, Valerie; Gaillard, Jean-Louis; Heym, Beate; Girard-Misguich, Fabienne; Brosch, Roland; Sapriel, Guillaume

    2016-01-01

    In mycobacteria, various type VII secretion systems corresponding to different ESX (ESAT-6 secretory) types, are contributing to pathogenicity, iron acquisition, and/or conjugation. In addition to the known chromosomal ESX loci, the existence of plasmid-encoded ESX systems was recently reported. To investigate the potential role of ESX-encoding plasmids on mycobacterial evolution, we analyzed a large representative collection of mycobacterial genomes, including both chromosomal and plasmid-borne sequences. Data obtained for chromosomal ESX loci confirmed the previous five classical ESX types and identified a novel mycobacterial ESX-4-like type, termed ESX-4-bis. Moreover, analysis of the plasmid-encoded ESX loci showed extensive diversification, with at least seven new ESX profiles, identified. Three of them (ESX-P clusters 1–3) were found in multiple plasmids, while four corresponded to singletons. Our phylogenetic and gene-order-analyses revealed two main groups of ESX types: 1) ancestral types, including ESX-4 and ESX-4-like systems from mycobacterial and non-mycobacterial actinobacteria and 2) mycobacteria-specific ESX systems, including ESX-1-2-3-5 systems and the plasmid-encoded ESX types. Synteny analysis revealed that ESX-P systems are part of phylogenetic groups that derived from a common ancestor, which diversified and resulted in the different ESX types through extensive gene rearrangements. A converging body of evidence, derived from composition bias-, phylogenetic-, and synteny analyses points to a scenario in which ESX-encoding plasmids have been a major driving force for acquisition and diversification of type VII systems in mycobacteria, which likely played (and possibly still play) important roles in the adaptation to new environments and hosts during evolution of mycobacterial pathogenesis. PMID:26748339

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

    Directory of Open Access Journals (Sweden)

    Zakham Fathiah

    2012-08-01

    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.

  14. Evaluation of a low-density hydrogel microarray technique for mycobacterial species identification.

    Science.gov (United States)

    Zimenkov, Danila V; Kulagina, Elena V; Antonova, Olga V; Krasnova, Maria A; Chernyaeva, Ekaterina N; Zhuravlev, Vyacheslav Y; Kuz'min, Alexey V; Popov, Sergey A; Zasedatelev, Alexander S; Gryadunov, Dmitry A

    2015-04-01

    In addition to the obligatory pathogenic species of the Mycobacterium tuberculosis complex and Mycobacterium leprae, the genus Mycobacterium also includes conditionally pathogenic species that in rare cases can lead to the development of nontuberculous mycobacterial diseases. Because tuberculosis and mycobacteriosis have similar clinical signs, the accurate identification of the causative agent in a clinical microbiology laboratory is important for diagnostic verification and appropriate treatment. This report describes a low-density hydrogel-based microarray containing oligonucleotide probes based on the species-specific sequences of the gyrB gene fragment for mycobacterial species identification. The procedure included the amplification of a 352-nucleotide fragment of the gene and its hybridization on a microarray. The triple-species-specific probe design and the algorithm for hybridization profile recognition based on the calculation of Pearson correlation coefficients, followed by the construction of a profile database, allowed for the reliable and accurate identification of mycobacterial species, including mixed-DNA samples. The assay was used to evaluate 543 clinical isolates from two regions of Russia, demonstrating its ability to detect 35 mycobacterial species, with 99.8% sensitivity and 100% specificity when using gyrB, 16S, and internal transcribed spacer (ITS) fragment sequencing as the standard. The testing of clinical samples showed that the sensitivity of the assay was 89% to 95% for smear-positive samples and 36% for smear-negative samples. The large number of identified species, the high level of sensitivity, the ability to detect mycobacteria in clinical samples, and the up-to-date profile database make the assay suitable for use in routine laboratory practice. PMID:25609722

  15. Induction of antigen-specific CD4+ HLA-DR-restricted cytotoxic T lymphocytes as well as nonspecific nonrestricted killer cells by the recombinant mycobacterial 65-kDa heat-shock protein.

    Science.gov (United States)

    Ab, B K; Kiessling, R; Van Embden, J D; Thole, J E; Kumararatne, D S; Pisa, P; Wondimu, A; Ottenhoff, T H

    1990-02-01

    Acquired cell-mediated immunity to intracellular parasites like mycobacteria is dependent on antigen-specific T lymphocytes. We have recently found that mycobacteria not only induce helper T cells but also cytotoxic CD4+ and/or CD8+ T cells as well as nonspecific killer cells that lyse human macrophages in vitro. In addition, we have described that the recombinant heat-shock protein (hsp) 65 of Mycobacterium bovis BCG/M, tuberculosis is an important target antigen for CD4+CD8- cytotoxic T cells. We have now further investigated the cytotoxic effector cells that are induced by the hsp65 of BCG. Purified protein derivative of tuberculin (PPD)- or hsp65-specific cytotoxic T cells specifically lysed PPD, hsp65 of BCG and hsp65 of M. leprae-pulsed macrophages in an HLA-DR-restricted manner. Nonpulsed macrophages were lysed to a much lower but still significant extent. hsp65-induced effector cells expressed CD3, CD5, CD4, CD8 and CD56 markers. Depletion experiments showed that the antigen-specific HLA-DR-restricted killer cell was of the CD5+CD4+CD8-CD56- phenotype. Experiments using N-terminal truncated hsp65 fusion (cro-lacZ) proteins suggested that the N-terminal 65 amino acid residues of the 540 amino acid molecule are critical for the expression of the cytotoxic target epitope(s) in two individuals tested. In addition to inducing antigen-specific cytotoxic effector cells, the hsp65 also triggered nonspecific nonrestricted effector cells with lytic activity against nonpulsed autologous or allogeneic macrophages as well as K-562 and Daudi tumor cells. hsp65-stimulated effector cells produced both interferon and tumor necrosis factor-alpha. An important finding was that hsp65-stimulated effector cells strongly inhibited colony-forming unit formation from live BCG-infected autologous macrophages. PMID:1690136

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

    Energy Technology Data Exchange (ETDEWEB)

    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

    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.

  17. Imaging of non-tuberculous (atypical) mycobacterial pulmonary infection

    Energy Technology Data Exchange (ETDEWEB)

    Ellis, S.M.; Hansell, D.M

    2002-08-01

    Pulmonary infections due to mycobacterial organisms are increasing in incidence. Non-tuberculous (atypical) mycobacteria (NTM) represent a significant proportion of mycobacterial infections and may prove difficult to diagnose due to their non-specific clinical and radiographic presentations. An increasing volume of radiological data is now available for the more common non-tuberculous mycobacterial infections, and we have summarized the imaging features found in such cases, identifying radiographic features that would favour the diagnosis of a non-tuberculous mycobacterium and that, in some cases, suggest a specific organism. Ellis, S.M. and Hansell, D.M. (2002)

  18. 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

  19. Assembly of the Mycobacterial Cell Wall.

    Science.gov (United States)

    Jankute, Monika; Cox, Jonathan A G; Harrison, James; Besra, Gurdyal S

    2015-10-15

    Mycobacterium tuberculosis remains one of the most successful bacterial pathogens, claiming over 1.3 million lives worldwide in 2013. The emergence of multidrug-resistant and extensively drug-resistant isolates has prompted the need for new drugs and drug targets. M. tuberculosis possesses an unusual cell wall dominated by lipids and carbohydrates that provides a permeability barrier against hydrophilic drugs and is crucial for its survival and virulence. This large macromolecular structure, termed the mycolyl-arabinogalactan-peptidoglycan complex, and the phosphatidyl-myo-inositol-based lipoglycans are key features of the mycobacterial cell wall. Assembly of these cell wall components is an attractive target for the development of chemotherapeutics against tuberculosis. Herein, we focus on recent biochemical and molecular insights into these complex molecules of M. tuberculosis cell wall. PMID:26488279

  20. Nontuberculous mycobacterial pulmonary diseases in immunocompetent patients

    Energy Technology Data Exchange (ETDEWEB)

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

    2002-09-01

    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.

  1. Molecular Characterization of Environmental Non-Tuberculous Mycobacteria Using PCR- RFLP Analysis of 441 Bp Heat Shock Protein 65 Fragments

    Directory of Open Access Journals (Sweden)

    H Rezaei-Yazdi

    2012-04-01

    Full Text Available Background: Non- Tuberculous Mycobacteria are environmental opportunistic pathogens that can be found in various terrestrial and aquatic habitats. There are an epidemiological links between species isolated in tap water and those isolated from patients. hsp65 gene has more variability in its sequences, compared to the some more conserved genes in NTM, for identification of mycobacteria to species level. In this study, the prevalence of NTM in Isfahan City water samples was determined using culture, biochemical tests and PCR-RFLP analyses of hsp65 gene.Methods: Eighty-five water samples were collected and cultured. The mycobacterial isolates were identified by conventional biochemical tests. A 441 bp fragment of hsp65 genes was amplified and digested by two restriction enzymes, BstEII and HaeII. Digested products were analyzed using polyacrilamid gel electrophoresis (PAGE.Results: 25.9% of the water samples contained different species of NTM. Dominant isolates were M. fortuitum (26.7%, M. chelonae like organism (13.3% and M. mucogenicum (13.3%. Nineteen isolates of Mycobacteria were differentiated using hsp65 genes PCR-RFLP. Three isolates could not be identified at the species level because their RFLP patterns were different from other known PCR-RFLP profiles. There were different hsp65 gene PCR-RFLP profiles produced by digestion with BstEII and HaeIII. Conclusion: This study showed that PCR-RFLP of hsp65 gene in mycobacteria is more reliable method for identification of NTM at the specie level than conventional phenotypic methods (P<0.05. In comparing of RFLP patterns of this study to other investigation, some minor differences were negligible.

  2. Mycobacterial antigen detection by immunohistochemistry in pulmonary tuberculosis.

    Science.gov (United States)

    Humphrey, D M; Weiner, M H

    1987-07-01

    A preliminary diagnosis of tuberculosis can be established by the detection of acid-fast bacilli (AFB) and confirmed by culture of the microorganism. To evaluate an alternative method of diagnosis, the distribution of mycobacterial antigens in lung tissue specimens was characterized by an indirect peroxidase-antiperoxidase method and was compared to the detection of AFB by Ziehl-Neelsen stain. Histologic specimens were obtained from 59 hospital patients. Of nine patients with mycobacterial disease, seven had antigen detected in tissue. In two patients with tuberculous pneumonia, the distribution of mycobacterial antigens was approximately the same as that of AFB. In contrast, in four patients with caseating pulmonary granulomas, clumps of mycobacterial antigens were demonstrated in necrotic areas of the granulomas where there were few or no AFB. In one patient with Mycobacterium intracellulare infection, cross-reactive antigens stained weakly. Antigen was not found in tissue from two patients; one had miliary lung granulomas, and the second had mediastinal lymph node granulomas. Mycobacterial antigens were not detected in specimens from 50 control patients with nonmycobacterial diseases. On the basis of this study of 59 cases, immunohistochemical detection of microbial antigens appears to be useful for establishing the mycobacterial etiology of caseating pulmonary granulomas. PMID:3297995

  3. Mechanistic insight into mycobacterial MmpL protein function.

    Science.gov (United States)

    Székely, R; Cole, S T

    2016-03-01

    Mycobacterial cell walls are complex structures containing a broad range of unusual lipids, glycolipids and other polymers, some of which act as immunomodulators or virulence determinants. Better understanding of the enzymes involved in export processes would enlighten cell wall biogenesis. Bernut et al. () present the findings of a structural and functional investigation of one of the most important transporter families, the MmpL proteins, members of the resistance-nodulation-cell division (RND) superfamily. A Tyr842His missense mutation in the mmpL4a gene was shown to be responsible for the smooth-to-rough morphotype change of the near untreatable opportunistic pathogen Mycobacterium bolletii due to its failure to export a glycopeptidolipid (GPL). This mutation was pleiotropic and markedly increased virulence in infection models. Tyr842 is well conserved in all actinobacterial MmpL proteins suggesting that it is functionally important and this was confirmed by several approaches including replacing the corresponding residue in MmpL3 of Mycobacterium tuberculosis. Structural modelling combined with experimental results showed Tyr842 to be a critical residue for mediating the proton motive force required for GPL export. This mechanistic insight applies to all MmpL proteins and probably to all RND transporters. PMID:26710752

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

    DEFF Research Database (Denmark)

    Thakur, Aneesh; Sharma, Mandeep; Katoch, Vipin C.; Dhar, Prasenjit; Katoch, R. C.

    2012-01-01

    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....

  5. Inhibitors Selective for Mycobacterial Versus Human Proteasomes

    Energy Technology Data Exchange (ETDEWEB)

    Lin, G.; Li, D; Sorio de Carvalho, L; Deng, H; Tao, H; Vogt, G; Wu, K; Schneider, J; Chidawanyika, T; et. al.

    2009-01-01

    Many anti-infectives inhibit the synthesis of bacterial proteins, but none selectively inhibits their degradation. Most anti-infectives kill replicating pathogens, but few preferentially kill pathogens that have been forced into a non-replicating state by conditions in the host. To explore these alternative approaches we sought selective inhibitors of the proteasome of Mycobacterium tuberculosis. Given that the proteasome structure is extensively conserved, it is not surprising that inhibitors of all chemical classes tested have blocked both eukaryotic and prokaryotic proteasomes, and no inhibitor has proved substantially more potent on proteasomes of pathogens than of their hosts. Here we show that certain oxathiazol-2-one compounds kill non-replicating M.?tuberculosis and act as selective suicide-substrate inhibitors of the M.?tuberculosis proteasome by cyclocarbonylating its active site threonine. Major conformational changes protect the inhibitor-enzyme intermediate from hydrolysis, allowing formation of an oxazolidin-2-one and preventing regeneration of active protease. Residues outside the active site whose hydrogen bonds stabilize the critical loop before and after it moves are extensively non-conserved. This may account for the ability of oxathiazol-2-one compounds to inhibit the mycobacterial proteasome potently and irreversibly while largely sparing the human homologue.

  6. Mycobacterial sensitins: where are we now?

    Science.gov (United States)

    Magnusson, M

    1981-01-01

    A limited review of the results of using comparative reciprocal intradermal mycobacterial sensitin (CRIS) testing with guinea pigs for the classification and identification of mycobacteria is presented. The technical procedures and the materials used in CRIS testing are referred to only briefly. Strains of mycobacteria that grow well on a synthetic nonimmunogenic medium can be identified or classified at the species level with this method. At present some 50 species of mycobacteria can be identified by CRIS testing. With this method alone, delineation between the genera Mycobacterium and Nocardia (or between subgenera of Mycobacterium proposed earlier) and differentiation between individual strains of Mycobacterium tuberculosis, Mycobacterium bovis, and Mycobacterium africanum are not possible. In a blind study that utilized CRIS testing, a total of 10 strains of Mycobacterium avium complex serovars 1, 2, 4, and 6 and one strain of serovar 9 were identified as M. avium; a total of five strains of serovars 14, 16, 17, and 20 and one strain of serovar 12 were identified as Mycobacterium intracellulare; a total of seven strains of serovars 41, 42, and 43 and two other strains of serovar 9 were identified as Mycobacterium scrofulaceum; and two other strains of serovar 9 appeared to be distinct from M. avium and from the other two species just mentioned (author's unpublished data). CRIS tests cannot be used at present for the identification of mycobacteria that cannot be cultivated. PMID:7339825

  7. Mycobacterial infections in a large Virginia hospital, 2001-2009

    Directory of Open Access Journals (Sweden)

    Scully Kenneth W

    2011-05-01

    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.

  8. Identification of the sugars involved in mycobacterial cell aggregation.

    Science.gov (United States)

    Anton, V; Rougé, P; Daffé, M

    1996-11-01

    Incubation of Mycobacterium tuberculosis and M smegmatis cells with the sugar components of their surface-exposed glycans demonstrated that D-arabinose but not alpha-D-glucose or D-mannose, led to the dispersion of the large clumps formed by the bacilli in stationary liquid cultures. These results confirm the presence of arabinose-containing glycans on the mycobacterial cell surface and demonstrate the implication of selective sugars in cell aggregation, suggesting that the clumping of mycobacterial cells is probably mediated by lectin-carbohydrate interactions. PMID:8900060

  9. DNA vaccination boosts Bacillus Calmette-Guérin protection against mycobacterial infection in zebrafish.

    Science.gov (United States)

    Oksanen, Kaisa E; Myllymäki, Henna; Ahava, Maarit J; Mäkinen, Leena; Parikka, Mataleena; Rämet, Mika

    2016-01-01

    Despite the widespread use of the current Bacillus Calmette-Guérin (BCG) vaccine, tuberculosis is still a major cause of morbidity and mortality worldwide. Vaccination with BCG does not prevent a Mycobacterium tuberculosis infection, nor does it inhibit the reactivation of latent tuberculosis. Here, we show that adult zebrafish are modestly and variably protected from a mycobacterial infection by BCG vaccination. An intraperitoneal (i.p.) BCG vaccination was associated with enhanced survival upon a high-dose (20,000 bacteria) Mycobacterium marinum infection. In addition, BCG-vaccinated fish were more able to restrict a low-dose (30 bacteria) intraperitoneal infection with M. marinum, as indicated by lower bacterial loads at six weeks post infection (wpi). However, the vaccination could not completely prevent an infection. A qRT-PCR analysis comparing BCG-vaccinated and unvaccinated fish upon a mycobacterial infection indicated that the induction of Tumor necrosis factor (TNF) was more modest in vaccinated fish. The partial protection gained by BCG could be boosted by a DNA vaccine combining Ag85B, ESAT6 and a resuscitation-related gene RpfE, suggesting that this combination of antigens could be useful for a future BCG booster vaccine. We conclude that zebrafish is a useful early-phase preclinical model for studying subunit vaccines designed for boosting the effects of BCG. PMID:26363085

  10. HIV Disrupts Human T Cells That Target Mycobacterial Glycolipids.

    Science.gov (United States)

    Kasprowicz, Victoria O; Cheng, Tan-Yun; Ndung'u, Thumbi; Sunpath, Henry; Moody, D Branch; Kasmar, Anne G

    2016-02-15

    Single-cell analysis captures the heterogeneity of T-cell populations that target defined antigens. Human immunodeficiency virus (HIV) infection results in defects of antimycobacterial immunity, which remain poorly defined. We therefore recruited a small number of subjects, including those with latent and active M. tuberculosis infection, with or without concomitant HIV infection, and tracked the mycobacterial glycolipid-reactive T-cell repertoire by using CD1b tetramers. Glycolipid-reactive T cells expressed memory markers and the HIV coreceptors CD4 and CCR5; they were not detected in subjects with HIV-associated active M. tuberculosis infection. HIV infection may affect T cells that recognize mycobacterial glycolipids and influence immunity. PMID:26374910

  11. Polyphenolic acetates : A newer anti-Mycobacterial therapeutic option

    Directory of Open Access Journals (Sweden)

    Santram

    2014-01-01

    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.

  12. Assembly and proteolytic processing of mycobacterial ClpP1 and ClpP2

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    Benaroudj Nadia

    2011-12-01

    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.

  13. Sulfate Reducing Bacteria and Mycobacteria Dominate the Biofilm Communities in a Chloraminated Drinking Water Distribution System.

    Science.gov (United States)

    Gomez-Smith, C Kimloi; LaPara, Timothy M; Hozalski, Raymond M

    2015-07-21

    The quantity and composition of bacterial biofilms growing on 10 water mains from a full-scale chloraminated water distribution system were analyzed using real-time PCR targeting the 16S rRNA gene and next-generation, high-throughput Illumina sequencing. Water mains with corrosion tubercles supported the greatest amount of bacterial biomass (n = 25; geometric mean = 2.5 × 10(7) copies cm(-2)), which was significantly higher (P = 0.04) than cement-lined cast-iron mains (n = 6; geometric mean = 2.0 × 10(6) copies cm(-2)). Despite spatial variation of community composition and bacterial abundance in water main biofilms, the communities on the interior main surfaces were surprisingly similar, containing a core group of operational taxonomic units (OTUs) assigned to only 17 different genera. Bacteria from the genus Mycobacterium dominated all communities at the main wall-bulk water interface (25-78% of the community), regardless of main age, estimated water age, main material, and the presence of corrosion products. Further sequencing of the mycobacterial heat shock protein gene (hsp65) provided species-level taxonomic resolution of mycobacteria. The two dominant Mycobacteria present, M. frederiksbergense (arithmetic mean = 85.7% of hsp65 sequences) and M. aurum (arithmetic mean = 6.5% of hsp65 sequences), are generally considered to be nonpathogenic. Two opportunistic pathogens, however, were detected at low numbers: M. hemophilum (arithmetic mean = 1.5% of hsp65 sequences) and M. abscessus (arithmetic mean = 0.006% of hsp65 sequences). Sulfate-reducing bacteria from the genus Desulfovibrio, which have been implicated in microbially influenced corrosion, dominated all communities located underneath corrosion tubercules (arithmetic mean = 67.5% of the community). This research provides novel insights into the quantity and composition of biofilms in full-scale drinking water distribution systems, which is critical for assessing the risks to public health and to the water supply infrastructure. PMID:26098899

  14. Mycobacterial endocarditis: a comprehensive review / Endocardite micobacteriana: uma revisão abrangente

    Scientific Electronic Library Online (English)

    Shi-Min, Yuan.

    2015-02-01

    Full Text Available Objetivo: Uma análise sistemática foi feita considerando epidemiologia, quadro clínico, diagnóstico, tratamento e principais resultados da endocardite micobacteriana. Métodos: Foi realizada uma pesquisa bibliográfica abrangente no MEDLINE, Highwire Press e no Google para publicações sobre endocardi [...] te micobacteriana, publicados entre 2000 e 2013. Resultados: As micobactérias de crescimento rápido tornam-se os patógenos predominantes, com Mycobacterium chelonae sendo a mais comum. Essa condição se alterou significativamente em termos de epidemiologia, desde o início do século 21, abrangendo faixa etária mais ampla, maior latência, prevalecendo infecções da valva mitral e melhor prognóstico. Conclusão: Endocardite micobacteriana é rara e os patógenos causadores são predominantemente as micobactérias de crescimento rápido. Amicacina, ciprofloxacina e claritromicina são os agentes antimicrobianos mais frequentemente utilizados, mas muitas vezes apresentam respostas pobres. Pacientes com infecções profundas podem justificar um procedimento cirúrgico ou retirada de linha. Com a poliquimioterapia periódica guiada por testes de sensibilidade às drogas, e abordagens cirúrgicas, os pacientes podem obter bons resultados terapêuticos. Abstract in english Objective: A systematic analysis was made in view of the epidemiology, clinical features, diagnosis, treatment and main outcomes of mycobacterial endocarditis. Methods: The data source of the present study was based on a comprehensive literature search in MEDLINE, Highwire Press and Google search e [...] ngine for publications on mycobacterial endocarditis published between 2000 and 2013. Results: The rapidly growing mycobacteria become the predominant pathogens with Mycobacterium chelonae being the most common. This condition has changed significantly in terms of epidemiology since the 21st century, with more broad patient age range, longer latency, prevailed mitral valve infections and better prognosis. Conclusion: Mycobacterial endocarditis is rare and the causative pathogens are predominantly the rapidly growing mycobacteria. Amikacin, ciprofloxacin and clarithromycin are the most frequently used targeted antimicrobial agents but often show poor responses. Patients with deep infections may warrant a surgical operation or line withdrawal. With periodic multidrug therapy guided by drug susceptibility testing, and surgical managements, patients may achieve good therapeutic results.

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

    Directory of Open Access Journals (Sweden)

    Maurischat Sven

    2008-06-01

    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.

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

    Science.gov (United States)

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

    2014-01-01

    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, 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

  17. CD1 and mycobacterial lipids activate human T cells.

    Science.gov (United States)

    Van Rhijn, Ildiko; Moody, D Branch

    2015-03-01

    For decades, proteins were thought to be the sole or at least the dominant source of antigens for T cells. Studies in the 1990s demonstrated that CD1 proteins and mycobacterial lipids form specific targets of human ?? T cells. The molecular basis by which T-cell receptors (TCRs) recognize CD1-lipid complexes is now well understood. Many types of mycobacterial lipids function as antigens in the CD1 system, and new studies done with CD1 tetramers identify T-cell populations in the blood of tuberculosis patients. In human populations, a fundamental difference between the CD1 and major histocompatibility complex systems is that all humans express nearly identical CD1 proteins. Correspondingly, human CD1 responsive T cells show evidence of conserved TCRs. In addition to natural killer T cells and mucosal-associated invariant T (MAIT cells), conserved TCRs define other subsets of human T cells, including germline-encoded mycolyl-reactive (GEM) T cells. The simple immunogenetics of the CD1 system and new investigative tools to measure T-cell responses in humans now creates a situation in which known lipid antigens can be developed as immunodiagnostic and immunotherapeutic reagents for tuberculosis disease. PMID:25703557

  18. Flow cytometry sorting of recombinant mycobacterial species yields bacterial clones with enhanced insert expression.

    Science.gov (United States)

    Yu, Jae-Sung; Whitesides, John; Lee, Sun-Hee; Taylor, Natalie; Jacobs, William R; Letvin, Norman L; Haynes, Barton F

    2011-01-01

    Recombinant mycobacteria hold promise as vectors for delivery of HIV-1 and other pathogen antigen inserts for inducing systemic and mucosal immune responses. In general, the immunogenicity of the recombinant mycobacterial insert is proportional to the level of insert expression. In this study, a novel flow cytometry-based assay has been developed to sort live recombinant mycobacterial mutants with high expression of foreign inserts and to enrich those sorted bacterial populations. Sorted recombinant mycobacterial clones expressed higher levels of the ovalbumin SIINFEKL epitope, and select sorted clones showed better immunogenicity than unsorted recombinant mycobacteria. Thus, flow cytometry-based sorting can isolate recombinant mycobacteria enriched for higher insert expression. PMID:21068210

  19. Specific detection of the cleavage activity of mycobacterial enzymes using a quantum dot based DNA nanosensor

    DEFF Research Database (Denmark)

    Jepsen, Morten Leth; Harmsen, Charlotte; Godbole, Adwait Anand; Nagaraja, Valakunja; Knudsen, Birgitta R; Ho, Yi-Ping

    2016-01-01

    We present a quantum dot based DNA nanosensor specifically targeting the cleavage step in the reaction cycle of the essential DNA-modifying enzyme, mycobacterial topoisomerase I. The design takes advantages of the unique photophysical properties of quantum dots to generate visible fluorescence...... recovery upon specific cleavage by mycobacterial topoisomerase I. This report, for the first time, demonstrates the possibility to quantify the cleavage activity of the mycobacterial enzyme without the pre-processing sample purification or post-processing signal amplification. The cleavage induced signal...

  20. Immunoregulation of CNS autoimmunity by helminth and mycobacterial infections.

    Science.gov (United States)

    Sewell, Diane L; Reinke, Emily K; Hogan, Laura H; Sandor, Matyas; Fabry, Zsuzsa

    2002-06-01

    The 'hygiene hypothesis' has been proposed to explain apparent increases in autoimmune disease and allergy in areas of the world with improved health care and sanitation. This hypothesis proposes that the lack of serious childhood infections impairs development of an appropriately educated immune response. Imbalance of Th1 and Th2 responses and lack of regulatory T-cell populations are two of many proposed potential mechanisms for immune failures such as autoimmunity and allergy. We summarize the literature evidence for the influence of infectious organisms on autoimmunity with focus on helminth and mycobacterial infections. We also demonstrate that Schistosoma mansoni ova pretreatment, Mycobacterium bovis (BCG) infection, and lyophilized Mycobacterium tuberculosis all modify the course of clinical disease in mice induced for experimental autoimmune encephalomyelitis (a mouse model for human multiple sclerosis (MS)). Our data supports the applicability of the hygiene hypothesis to CNS autoimmune disease. PMID:12008041

  1. Color selection with a hygromycin-resistance-based Escherichia coli-mycobacterial shuttle vector.

    Science.gov (United States)

    Howard, N S; Gomez, J E; Ko, C; Bishai, W R

    1995-12-01

    Hygromycin-resistance (HyR)-based Escherichia coli-mycobacterial shuttle plasmids have high efficiencies of transformation and a broad mycobacterial host range. We have introduced a lacZ alpha (encoding the alpha-polypeptide fragment of beta-galactosidase (beta Gal))-multiple cloning site cassette into a HyR-based shuttle vector to generate a plasmid with nine unique cloning sites and the added feature of beta Gal color selection in appropriate E. coli host strains. PMID:8529888

  2. Comparison of Quantitative Techniques including Xpert MTB/RIF to Evaluate Mycobacterial Burden

    OpenAIRE

    van Zyl-Smit, R. N.; Binder, A; Meldau, R.; Mishra, H.; Semple, P. L.; Theron, G.; Peter, J.; Whitelaw, A.; Sharma, S. K.; R. Warren; Bateman, E D; Dheda, K.

    2011-01-01

    Introduction: Accurate quantification of mycobacterial load is important for the evaluation of patient infectiousness, disease severity and monitoring treatment response in human and in-vitro laboratory models of disease. We hypothesized that newer techniques would perform as well as solid media culture to quantify mycobacterial burden in laboratory specimens. Methods: We compared the turn-around-time, detection-threshold, dynamic range, reproducibility, relative discriminative ability, of 4 ...

  3. Mucosal cell-mediated immunity to mycobacterial, enterobacterial and other microbial antigens in inflammatory bowel disease.

    Science.gov (United States)

    Ibbotson, J P; Lowes, J R; Chahal, H; Gaston, J S; Life, P; Kumararatne, D S; Sharif, H; Alexander-Williams, J; Allan, R N

    1992-02-01

    Culture studies have suggested that Mycobacterium paratuberculosis may play a role in the aetiology of Crohn's disease. However, evidence of sensitization to mycobacterial antigens amongst patients with Crohn's disease has not yet been adequately demonstrated. Previous studies of cell-mediated immunity (CMI) in Crohn's disease were restricted to responses of peripheral blood mononuclear cells (PBMC) to mycobacterial antigens. In this study we have investigated the proliferative responses of both PBMC and mesenteric lymph node mononuclear cells (MLNMC) to a range of mycobacterial and non-mycobacterial antigens. There was no evidence of specific sensitization in the responses of MLNMC and PBMC from patients with inflammatory bowel disease (IBD) to the mycobacterial antigens. However, anergy to M. paratuberculosis could not be excluded. IBD MLNMC responses to most antigens were generally greater than those of PBMC, which were often undetectable. When compared with controls, there was evidence of increased CMI to a range of non-mycobacterial antigens, especially Yersinia enterocolitica, amongst both MLNMC and PBMC from patients with Crohn's disease and ulcerative colitis (UC). These results do not provide support to the proposed role of mycobacteria in the pathogenesis of Crohn's disease, but indicate that further investigation may determine a role for bacterial-specific T cell-mediated responses in the pathogenesis of IBD. PMID:1735186

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

    Directory of Open Access Journals (Sweden)

    Price Patricia

    2008-04-01

    Full Text Available 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. Results We studied two patients who experienced mycobacterial IRD during ART. One patient developed a second episode of IRD with distinct clinical characteristics. Findings were compared with patients 'at risk' of developing IRD who had uneventful immune recovery. Peripheral blood mononuclear cells (PBMC from all subjects were stimulated with mycobacterial antigens in the form of purified protein derivative (PPD. Supernatants were assayed for IFN? and IL-10. In response to PPD, PBMC from IRD patients generated IFN? during the first IRD episode, whilst cells from non-IRD controls produced more IL-10. Conclusion We present preliminary data from two HIV-infected patients showing an imbalance between IFN? and IL-10 responses to mycobacterial antigens during mycobacterial IRD. Our findings suggest that imbalanced effector and regulatory cytokine responses should be investigated as a cause of IRD.

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

    LENUS (Irish Health Repository)

    O'Donnell, N

    2012-01-01

    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.

  6. Adequate Th2-type response associates with restricted bacterial growth in latent mycobacterial infection of zebrafish.

    Science.gov (United States)

    Hammarén, Milka Marjut; Oksanen, Kaisa Ester; Nisula, Hanna Maria; Luukinen, Bruno Vincent; Pesu, Marko; Rämet, Mika; Parikka, Mataleena

    2014-06-01

    Tuberculosis is still a major health problem worldwide. Currently it is not known what kind of immune responses lead to successful control and clearance of Mycobacterium tuberculosis. This gap in knowledge is reflected by the inability to develop sufficient diagnostic and therapeutic tools to fight tuberculosis. We have used the Mycobacterium marinum infection model in the adult zebrafish and taken advantage of heterogeneity of zebrafish population to dissect the characteristics of adaptive immune responses, some of which are associated with well-controlled latency or bacterial clearance while others with progressive infection. Differences in T cell responses between subpopulations were measured at the transcriptional level. It was discovered that a high total T cell level was usually associated with lower bacterial loads alongside with a T helper 2 (Th2)-type gene expression signature. At late time points, spontaneous reactivation with apparent symptoms was characterized by a low Th2/Th1 marker ratio and a substantial induction of foxp3 reflecting the level of regulatory T cells. Characteristic gata3/tbx21 has potential as a biomarker for the status of mycobacterial disease. PMID:24968056

  7. Doença pulmonar por Mycobacterium tuberculosis e micobactérias não-tuberculosas entre pacientes recém-diagnosticados como HIV positivos em Moçambique, África Mycobacterium tuberculosis and nontuberculous mycobacterial isolates among patients with recent HIV infection in Mozambique

    Directory of Open Access Journals (Sweden)

    Elizabete Abrantes Nunes

    2008-10-01

    Full Text Available OBJETIVO: A micobacteriose é frequentemente diagnosticada entre pacientes infectados pelo HIV. Em Moçambique, onde apenas um pequeno número de pacientes encontra-se sob tratamento anti-retroviral, e a tuberculose tem alta prevalência, existe a necessidade de melhor caracterização destes agentes bacterianos, em nível de espécie, bem como de se caracterizar os padrões de resistência às drogas antituberculosas. MÉTODOS: Em uma coorte de 503 indivíduos HIV positivos suspeitos de tuberculose pulmonar, 320 apresentaram positividade para baciloscopia ou cultura no escarro e no lavado brônquico. RESULTADOS: Bacilos álcool-ácido resistentes foram detectados no escarro em 73% dos casos com cultura positiva. De 277 isolados em cultura, apenas 3 mostraram-se tratar de micobactérias não-tuberculosas: 2 Mycobacterium avium e uma M. simiae. Todos os isolados de M. tuberculosis inicialmente caracterizados através de reação em cadeia de polimerase (RCP do gene hsp65 foram posteriormente caracterizados como tal através de RCP do gene gyrB. Resistência à isoniazida foi encontrada em 14% dos casos; à rifampicina em 6%; e multirresistência em 5%. Pacientes previamente tratados para tuberculose mostraram tendência a taxas maiores de resistência às drogas de primeira linha. O padrão radiológico mais freqüente encontrado foi o infiltrado intersticial (67%, seguido da presença de linfonodos mediastinais (30%, bronquiectasias (28%, padrão miliar (18% e cavidades (12%. Os pacientes infectados por micobactérias não-tuberculosas não apresentaram manifestações clínicas distintas das apresentadas pelos outros pacientes. A mediana de linfócitos CD4 entre todos os pacientes foi de 134 células/mm³. CONCLUSÕES: Tuberculose e AIDS em Moçambique estão fortemente associadas, como é de se esperar em países com alta prevalência de tuberculose. Embora as taxas de resistência a drogas sejam altas, o esquema isoniazida-rifampicina continua sendo a escolha apropriada para o início do tratamento.OBJECTIVE: Mycobacteriosis is frequently diagnosed among HIV-infected patients. In Mozambique, where few patients are under antiretroviral therapy and the prevalence of tuberculosis is high, there is need for better characterization of mycobacteria at the species level, as well as for the identification of patterns of resistance to antituberculous drugs. METHODS: We studied a sample of 503 HIV-infected individuals suspected of having pulmonary tuberculosis. Of those 503, 320 tested positive for mycobacteria through sputum smear microscopy or culture of bronchoalveolar lavage fluid. RESULTS: Acid-fast bacilli were observed in the sputum of 73% of the individuals presenting positive cultures. Of 277 isolates tested, only 3 were nontuberculous mycobacteria: 2 were identified as Mycobacterium avium and one was identified as M. simiae. Strains initially characterized as M. tuberculosis complex through polymerase chain reaction restriction analysis (PRA of the hsp65 gene were later confirmed as such through PRA of the gyrB gene. Among the M. tuberculosis isolates, resistance patterns were as follows: to isoniazid, 14%; to rifampin, 6%; and multidrug resistance, 5%. Previously treated cases showed significantly higher rates of resistance to first-line antituberculous drugs. The most common radiological pattern was interstitial infiltrate (in 67%, followed by mediastinal lymph node enlargement (in 30%, bronchiectasis (in 28%, miliary nodules (in 18% and cavitation (in 12%. Patients infected with nontuberculous mycobacteria presented clinical profiles indistinguishable from those of other patients. The median CD4 lymphocyte count in this group was 134 cells/mm³. CONCLUSIONS: There is a strong association between tuberculosis and AIDS in Mozambique, as expected in a country with a high prevalence of tuberculosis. Although drug resistance rates are high, the isoniazid-rifampin regimen continues to be the appropriate choice for initial therapy.

  8. Molecular immunity to mycobacteria: knowledge from the mutation and phenotype spectrum analysis of Mendelian susceptibility to mycobacterial diseases

    OpenAIRE

    Qu, Hui-Qi; Susan P. Fisher-Hoch; McCormick, Joseph B

    2011-01-01

    Understanding molecular immunity against mycobacterial infection is critical for the development of effective strategies to control tuberculosis (TB), which is a major health issue in the developing world. Host immunogenetic studies represent an indispensable approach to understand the molecular mechanisms against mycobacterial infection. A superb paradigm is the identification of rare mutations causing Mendelian susceptibility to mycobacterial diseases (MSMD). Mutations in the interferon-gam...

  9. Cytokine gene expression in the BB rat pancreas: natural course and impact of bacterial vaccines.

    Science.gov (United States)

    Kolb, H; Wörz-Pagenstert, U; Kleemann, R; Rothe, H; Rowsell, P; Scott, F W

    1996-12-01

    In diabetes prone BB rat pancreas the Th1/ Th2 cytokine balance and the expression of inducible nitric oxide synthase (iNOS) was determined by mRNA analysis before and after the onset of insulitis. Specific mRNA was amplified by reverse transcriptase polymerase chain reaction, quantitated with radiolabelled probes by phosphoimaging and calibrated with the amount of co-amplified beta-actin mRNA. At 50 days of age, prior to recognizable insulitis, there was already significantly enhanced expression of both, Th1 and Th2 cytokines, and of iNOS mRNA, when compared to Wistar rat pancreas (p OM-89, an endotoxin free extract containing immunostimulatory glycolipopeptides and heat shock protein (hsp) 65, both downregulated IFN gamma mRNA while only OM-89 in addition suppressed iNOS mRNA and enhanced Th2 cytokine gene expression (p OM-89) from E. coli. PMID:8960825

  10. Atypical mycobacterial cutaneous infections in Egyptians: a clinicopathological study.

    Science.gov (United States)

    El-Khalawany, Mohamed A

    2014-04-01

    Atypical mycobacteria comprise an uncommon heterogenous non-tuberculous group of acid-fast bacteria that rarely involve skin. The pattern of atypical mycobacterial cutaneous infections (AMCI) has not been previously studied in Egypt. The aim of this study was to describe the clinical characteristics, pathological features and species profile of AMCI among Egyptian patients. A retrospective study included 46 cases, diagnosed with AMCI during the period 2002 to 2012. The study included 34 males (73.9%) and 12 females (26.9%). The average age of patients was 39 years while the average duration of lesions was 15 months. The lesions were mostly located on the extremities (91.3%) and there was predominance of single (65.2%) and nodular (41.4%) lesions. History of trauma was confirmed in 91.3%. Histologically, the granulomas were mostly superficial (67.4%) with predominance of nodular suppurative pattern (84.8%). Other significant histological findings included epidermal hypertrophy (100%), presence of large-sized multinucleated giant cells (87%) and intrafollicular neutrophilic abscesses (84.8%). The diagnosis was proved by direct smear in 6.5%, skin biopsy in 10.9%, tissue culture in 47.8% and polymerase chain reaction (PCR) in 34.8%. Isolated species included Mycobacterium marinum (84.8%), Mycobacterium fortuitum (10.9%) and Mycobacterium kansasii (4.3%). Although the results of this study recommend that the diagnosis of AMCI is based mainly on culture and PCR, other clinicopathological features such as history of trauma, acral location of the lesion and suppurative granulomatous reaction with intrafollicular abscesses could be helpful clues in suspecting AMCI. PMID:24533920

  11. Myeloid Growth Factors Promote Resistance to Mycobacterial Infection by Curtailing Granuloma Necrosis through Macrophage Replenishment.

    Science.gov (United States)

    Pagán, Antonio J; Yang, Chao-Tsung; Cameron, James; Swaim, Laura E; Ellett, Felix; Lieschke, Graham J; Ramakrishnan, Lalita

    2015-07-01

    The mycobacterial ESX-1 virulence locus accelerates macrophage recruitment to the forming tuberculous granuloma. Newly recruited macrophages phagocytose previously infected apoptotic macrophages to become new bacterial growth niches. Granuloma macrophages can then necrose, releasing mycobacteria into the extracellular milieu, which potentiates their growth even further. Using zebrafish with genetic or pharmacologically induced macrophage deficiencies, we find that global macrophage deficits increase susceptibility to mycobacterial infection by accelerating granuloma necrosis. This is because reduction in the macrophage supply below a critical threshold decreases granuloma macrophage replenishment to the point where apoptotic infected macrophages, failing to get engulfed, necrose. Reducing macrophage demand by removing bacterial ESX-1 offsets the susceptibility of macrophage deficits. Conversely, increasing macrophage supply in wild-type fish by overexpressing myeloid growth factors induces resistance by curtailing necrosis. These findings may explain the susceptibility of humans with mononuclear cytopenias to mycobacterial infections and highlight the therapeutic potential of myeloid growth factors in tuberculosis. PMID:26159717

  12. Specific detection of the cleavage activity of mycobacterial enzymes using a quantum dot based DNA nanosensor

    Science.gov (United States)

    Jepsen, Morten Leth; Harmsen, Charlotte; Godbole, Adwait Anand; Nagaraja, Valakunja; Knudsen, Birgitta R.; Ho, Yi-Ping

    2015-12-01

    We present a quantum dot based DNA nanosensor specifically targeting the cleavage step in the reaction cycle of the essential DNA-modifying enzyme, mycobacterial topoisomerase I. The design takes advantages of the unique photophysical properties of quantum dots to generate visible fluorescence recovery upon specific cleavage by mycobacterial topoisomerase I. This report, for the first time, demonstrates the possibility to quantify the cleavage activity of the mycobacterial enzyme without the pre-processing sample purification or post-processing signal amplification. The cleavage induced signal response has also proven reliable in biological matrices, such as whole cell extracts prepared from Escherichia coli and human Caco-2 cells. It is expected that the assay may contribute to the clinical diagnostics of bacterial diseases, as well as the evaluation of treatment outcomes.We present a quantum dot based DNA nanosensor specifically targeting the cleavage step in the reaction cycle of the essential DNA-modifying enzyme, mycobacterial topoisomerase I. The design takes advantages of the unique photophysical properties of quantum dots to generate visible fluorescence recovery upon specific cleavage by mycobacterial topoisomerase I. This report, for the first time, demonstrates the possibility to quantify the cleavage activity of the mycobacterial enzyme without the pre-processing sample purification or post-processing signal amplification. The cleavage induced signal response has also proven reliable in biological matrices, such as whole cell extracts prepared from Escherichia coli and human Caco-2 cells. It is expected that the assay may contribute to the clinical diagnostics of bacterial diseases, as well as the evaluation of treatment outcomes. Electronic supplementary information (ESI) available: Characterization of the QD-based DNA Nanosensor. See DOI: 10.1039/c5nr06326d

  13. Mycobacterial spindle cell pseudotumour of the brain in a patient with sarcoidosis.

    Science.gov (United States)

    Ismail, Iyad; Carey, Martyn; Trotter, Simon; Kunst, Heinke

    2015-01-01

    Mycobacterial spindle cell pseudotumours (MSP) are benign lesions characterised by local proliferation of spindle-shaped histiocytes caused by mycobacterial infections. Cerebral MSP due to Mycobacterium avium intracellulare (MAI) infection is rare, and is often misdiagnosed clinically and radiologically as a brain tumour. We present a case with underlying sarcoidosis and known pulmonary MAI infection presenting with partial seizures and headaches. Imaging of the brain revealed a solitary extra axial tumour within the right temporal area. Biopsy of the tumour showed evidence of MPS due to MAI infection. Prolonged treatment with antituberculous therapy showed complete resolution of the cerebral lesion. PMID:26153278

  14. Influence of polymerase brand on microarray-based spoligotyping in low concentrations of mycobacterial DNA.

    Science.gov (United States)

    Monecke, Stefan; Engelmann, Ines; Ehricht, Ralf

    2015-04-01

    Spoligotyping is a widely used typing method for the Mycobacterium tuberculosis complex. Protocols and platforms can be adapted for direct use on patient samples. Serial dilutions of genomic DNA from Mycobacterium bovis BCG strain DSM45071 were spoligotyped by array hybridization using 32 different commercial PCR polymerase preparations. In samples with very low concentrations of mycobacterial DNA, commercially available PCR polymerases differed in their performance, and some yielded no, or false, identification. Direct spoligotyping from samples with very low concentrations of mycobacterial DNA thus requires careful selection of polymerase and strict standardization. PMID:25656919

  15. No association between interferon-? receptor-1 gene polymorphism and pulmonary tuberculosis in a Gambian population sample

    OpenAIRE

    Awomoyi, A; Nejentsev, S.; Richardson, A.; Hull, J; Koch, O; Podinovskaia, M; Todd, J.; McAdam, K.; Blackwell, J.; Kwiatkowski, D.; Newport, M

    2004-01-01

    Background: Tuberculosis (TB) is a major global cause of mortality and morbidity, and host genetic factors influence disease susceptibility. Interferon-? mediates immunity to mycobacteria and rare mutations in the interferon-? receptor-1 gene (IFNGR1) result in increased susceptibility to mycobacterial infection, including TB, in affected families. The role of genetic variation in IFNGR1 in susceptibility to common mycobacterial diseases such as pulmonary TB in outbred populations has not pre...

  16. The hydroxy-naphthoquinone lapachol arrests mycobacterial growth and immunomodulates host macrophages.

    Science.gov (United States)

    Oliveira, Renato A S; Azevedo-Ximenes, Eulalia; Luzzati, Roberto; Garcia, Rodolfo C

    2010-11-01

    The present study reports the anti-mycobacterial activity of 2-hydroxy-3-(3-methyl-2-butenyl)-1,4-naphthoquinone (lapachol) as well as its influence on macrophage functions. Lapachol (L) did not induce apoptosis/necrosis of THP-1 macrophages at ?32 ?g/mL. Mycobacterium avium liquid growth was arrested by ?32 ?g/mL and intra-macrophage proliferation by ?16 ?g/mL lapachol. The main immuno-modulatory effects of lapachol observed were an up-regulation of interferon-?-receptor 1 (IFN-?R1) and major histocompatibility complex class II (MHCII) surface expression, and a marked inhibition of IL-10 secretion. Lapachol did not affect resting, IFN-?- or toll-like receptor 2 (TLR2)-induced levels of oxygen and nitrogen metabolism key proteins nor the TLR2-mediated secretion of TNF-?, nor induced either oxidative or endoplasmic reticulum (ER) stress. Lapachol inhibited the surface expression of the co-stimulatory molecule CD86 but not that of CD80 and CD83. The results obtained indicate that the substituted naphthoquinone lapachol exhibits an anti-mycobacterial activity that is more efficient intra- than extra-cellularly, and exerts immuno-modulatory effects some of which may enhance the capacity of the host cell to control mycobacterial growth. The immune-modulatory action of lapachol could contribute to its more efficient intra-macrophage anti-mycobacterial activity. PMID:20837170

  17. An essential nonredundant role for mycobacterial DnaK in native protein folding.

    Science.gov (United States)

    Fay, Allison; Glickman, Michael S

    2014-07-01

    Protein chaperones are essential in all domains of life to prevent and resolve protein misfolding during translation and proteotoxic stress. HSP70 family chaperones, including E. coli DnaK, function in stress induced protein refolding and degradation, but are dispensable for cellular viability due to redundant chaperone systems that prevent global nascent peptide insolubility. However, the function of HSP70 chaperones in mycobacteria, a genus that includes multiple human pathogens, has not been examined. We find that mycobacterial DnaK is essential for cell growth and required for native protein folding in Mycobacterium smegmatis. Loss of DnaK is accompanied by proteotoxic collapse characterized by the accumulation of insoluble newly synthesized proteins. DnaK is required for solubility of large multimodular lipid synthases, including the essential lipid synthase FASI, and DnaK loss is accompanied by disruption of membrane structure and increased cell permeability. Trigger Factor is nonessential and has a minor role in native protein folding that is only evident in the absence of DnaK. In unstressed cells, DnaK localizes to multiple, dynamic foci, but relocalizes to focal protein aggregates during stationary phase or upon expression of aggregating peptides. Mycobacterial cells restart cell growth after proteotoxic stress by isolating persistent DnaK containing protein aggregates away from daughter cells. These results reveal unanticipated essential nonredunant roles for mycobacterial DnaK in mycobacteria and indicate that DnaK defines a unique susceptibility point in the mycobacterial proteostasis network. PMID:25058675

  18. Nontuberculous Mycobacterial Disease Is Not a Contraindication to Lung Transplantation in Patients With Cystic Fibrosis

    DEFF Research Database (Denmark)

    Qvist, Tavs; Pressler, Tanja; Thomsen, V O; Skov, M; Iversen, M; Katzenstein, T L

    2013-01-01

    Whether nontuberculous mycobacterial (NTM) disease is a contraindication to lung transplantation remains controversial. We conducted a nationwide study to evaluate the clinical importance of NTM infection among lung transplant patients with cystic fibrosis (CF) in Denmark and to determine if NTM...

  19. A SHORT INTERFERING RNA MOLECULAR BEACON FOR THE ATTENUATION OF MYCOBACTERIAL INFECTION

    Directory of Open Access Journals (Sweden)

    Remo George

    2014-01-01

    Full Text Available The ability of the pathogen Mycobacterium Tuberculosis (MTB to invade and survive within macrophages of granulomas is attributed to the product of the Mammalian Cell Entry (MCE operon whose gene, mce4A, encodes a cholesterol transporter that transports host lipids into the bacterium that allows the bacterium to survive during chronic infection. Here, we proposed and tested the hypothesis that a mce4A siRNA molecular beacon can be used to attenuate mycobacterial infection in macrophages. Mce4A gene was cloned and expressed in E. coli (E. coli-4A and differentiated U937 cells were transduced with piLenti-siRNA-GFP phage expressing the mce4A siRNA for 24 h. This was followed by infection with either E. coli-4A or M. smegmatis for 3 h followed by incubation for 0, 3, 6, 24 and 48 h. The cells were lysed and the lysates were plated on LB agar plates containing ampicillin (100 µg mL-1 or on 7H11 media and incubated at 37°C overnight. Our results showed that the siRNA treatment attenuated E.coli-4A infection in macrophages at 3, 6, 24 and 48 h by 0, 77, 59.6 and 99.7%, respectively. Our results also showed that the siRNA treatment attenuated M. smegmatis infection in macrophages at 3, 6, 24 and 48 h. by 94.8, 70.3, 98.9 and 93.4%, respectively. In conclusion, a mce4A siRNA molecular beacon was successfully delivered and stably expressed in macrophages which attenuated E. coli expressing mce4A (E. coli-4A and M. smegmatis infection in macrophages.

  20. Immunohistologic analysis of mycobacterial antigens by monoclonal antibodies in tuberculosis and mycobacteriosis.

    Science.gov (United States)

    Barbolini, G; Bisetti, A; Colizzi, V; Damiani, G; Migaldi, M; Vismara, D

    1989-11-01

    Four monoclonal antibodies (MoAbs), 60.15, 61.3, 105.10, and 2.16, directed to different proteins of Mycobacterium tuberculosis were used by an indirect avidin-biotin complex peroxidase-antiperoxidase method to detect mycobacterial antigens in lung, lymph node, and joint tissue specimens of tuberculous patients. Using MoAb 60.15, which recognizes a broad range of cross-reactive mycobacterial proteins with a molecular mass of 28 kilodaltons (kD), scattered materials (mycobacterial in origin) were observed, many of which were located within phagocyte cytoplasm. With MoAb 61.3, which reacts with a 35 kD protein present in M tuberculosis, Mycobacterium africanum, and Mycobacterium bovis, many clumped particles similar in size and shape to acid-fast bacilli were observed within the phagocyte cytoplasm (lung tissue) and positive macrophages with lysosomes were distributed throughout the cytoplasm (bronchoalveolar lavage). The specificity of this MoAb (61.3) was confirmed by the negative staining of positive lymph node specimens obtained from a patient infected with Mycobacterium kansasii. MoAbs 105.10 and 2.16 bind to the cross-reactive 65 kD heat shock protein that is present in mycobacteria and stain scattered particles and dark clumps of bacilli within the phagocyte cytoplasm. On the basis of this study, immunohistochemical detection of mycobacterial antigens appears to be useful in establishing the mycobacterial etiology of caseating granulomas and in avoiding the false-negative results obtained by traditional staining methods. PMID:2807270

  1. Study of the gyrB gene polymorphism as a tool to differentiate among Mycobacterium tuberculosis complex subspecies further underlines the older evolutionary age of 'Mycobacterium canettii'.

    Science.gov (United States)

    Goh, Khye Seng; Fabre, Michel; Huard, Richard C; Schmid, Solveig; Sola, Christophe; Rastogi, Nalin

    2006-01-01

    The present investigation evaluated the PCR-restriction fragment length polymorphism (RFLP) analysis of hsp65 and gyrB targets for differentiation of the species within the Mycobacterium tuberculosis complex (MTC) both by including new restriction enzymes and previously unstudied species. The hsp65 restriction analysis using HhaI resulted in a characteristic 'Mycobacterium canettii' pattern. A study of the gyrB gene polymorphism using TaqIalpha and HinfI allowed the initial division of MTC into two major groups, one consisting of M. tuberculosis and 'M. canettii' as opposed to another single group with other species. Three different patterns were observed with RsaI, the first characteristic of Mycobacterium microti, the second with Mycobacterium bovis, M. bovis BCG and Mycobacterium caprae (M. caprae was easily separated from M. bovis, and M. bovis BCG by SacII digestion), and the third with M. tuberculosis, 'M. canettii', Mycobacterium africanum, Mycobacterium pinnipedii, and the dassie bacillus. Although further discrimination within the last group was not obtained using additional restriction enzymes, the HaeIII and RsaI digestions highlighted an important gyrB polymorphism among 'M. canettii' strains. A study of the single nucleotide polymorphisms (SNP) within the gyrB by sequence analysis not only confirmed the results of the restriction analysis, but showed further differences among 'M. canettii' isolates that were not picked up using the existing battery of restriction enzymes. As many as 11 different SNPs were identified in the collection of eight 'M. canettii' isolates studied. Considering that gyrB variability among MTC member species other than 'M. canettii' is as restricted as hsp65 variability among MTC, our data corroborate a recent proposition that the 'M. canettii' group is evolutionary much older than the other MTC members. In conclusion, gyrB PCR-RFLP is a simple and rapid low-cost method that combined with phenotypic characteristics, may be helpful to differentiate most of the subspecies within the MTC. PMID:16517119

  2. Synthesis and anti-mycobacterial activity of 2-chloronicotinaldehydes based novel 1H-1,2,3-triazolylbenzohydrazides.

    Science.gov (United States)

    Suman, Pathi; Dayakar, Cherupally; Rajkumar, Kommera; Yashwanth, Bomma; Yogeeswari, Perumal; Sriram, Dharmarajan; Rao, Janapala Venkateswara; Raju, Bhimapaka China

    2015-06-01

    1H-1,2,3-Triazolylbenzohydrazides (6a-h and 11a-l) were synthesized from 2-chloronicotinaldehydes and evaluated for anti-mycobacterial activity against Mycobacterium tuberculosis H37Rv strain (ATCC-27294). Seven compounds 6b, 6e,f, 11d, 11h, 11j and 11l displayed potent anti-mycobacterial activity (MIC 2.8-6.2 ?M). Potent anti-mycobacterial compounds were chosen for cytotoxicity studies by MTT protein assay against normal cell lines (PBMC and Raw 264.7) and shown low cytotoxicity. This is the first Letter assigning anti-mycobacterial activity, cytotoxicity and structure activity relationship for 1H-1,2,3-triazolylbenzohydrazides. PMID:25908515

  3. Engineering new mycobacterial vaccine design for HIV–TB pediatric vaccine vectored by lysine auxotroph of BCG

    OpenAIRE

    Saubi, Narcís; Gea-Mallorquí, Ester; Ferrer, Pau; Hurtado, Carmen; Sánchez-Úbeda, Sara; Eto, Yoshiki; Gatell, Josep M.; Hanke, Tomáš; Joseph, Joan

    2014-01-01

    In this study, we have engineered a new mycobacterial vaccine design by using an antibiotic-free plasmid selection system. We assembled a novel Escherichia coli (E. coli)–mycobacterial shuttle plasmid p2auxo.HIVA, expressing the HIV-1 clade A immunogen HIVA. This shuttle vector employs an antibiotic resistance-free mechanism for plasmid selection and maintenance based on glycine complementation in E. coli and lysine complementation in mycobacteria. This plasmid was first transformed into glyc...

  4. Suppression of lymphocyte responses by tuberculous plasma and mycobacterial arabinogalactan. Monocyte dependence and indomethacin reversibility.

    OpenAIRE

    Kleinhenz, M E; Ellner, J. J.; Spagnuolo, P J; Daniel, T M

    1981-01-01

    During tuberculosis, exposure of monocytes to circulating factors may induce the suppressor activity observed in some anergic patients. To explore this possibility, we examined the effects of plasma pooled from 28 untreated tuberculosis (TB) patients and the mycobacterial cell wall polysaccharide D-arabino-D-galactan (AG) on the in vitro function of peripheral blood mononuclear cells (PBMC) from healthy donors. In the [3H] thymidine incorporation assay, stimulated responses of PBMC incubated ...

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

    OpenAIRE

    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

    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...

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

    Scientific Electronic Library Online (English)

    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

    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 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.

  7. 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 as adjuvant therapy in patients with pulmonary mycobacterial diseases or idiopathic pulmonary fibrosis. (author)

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

    Directory of Open Access Journals (Sweden)

    André Alves Dias

    2012-12-01

    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.

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

    Science.gov (United States)

    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

    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

  10. The molecular biology of mycobacterial trehalose in the quest for advanced tuberculosis therapies.

    Science.gov (United States)

    Nobre, Ana; Alarico, Susana; Maranha, Ana; Mendes, Vitor; Empadinhas, Nuno

    2014-08-01

    Trehalose is a natural glucose disaccharide identified in the 19th century in fungi and insect cocoons, and later across the three domains of life. In members of the genus Mycobacterium, which includes the tuberculosis (TB) pathogen and over 160 species of nontuberculous mycobacteria (NTM), many of which are opportunistic pathogens, trehalose has been an important focus of research over the last 60 years. It is a crucial player in the assembly and architecture of the remarkable mycobacterial cell envelope as an element of unique highly antigenic glycolipids, namely trehalose dimycolate ('cord factor'). Free trehalose has been detected in the mycobacterial cytoplasm and occasionally in oligosaccharides with unknown function. TB and NTM infection statistics and death toll, the decline in immune responses in the aging population, human immunodeficiency virus/AIDS or other debilitating conditions, and the proliferation of strains with different levels of resistance to the dated drugs in use, all merge into a serious public-health threat urging more effective vaccines, efficient diagnostic tools and new drugs. This review deals with the latest findings on mycobacterial trehalose biosynthesis, catabolism, processing and recycling, as well with the ongoing quest for novel trehalose-related mechanisms to be targeted by novel TB therapeutics. In this context, the drug-discovery pipeline has recently included new lead compounds directed toward trehalose-related targets highlighting the potential of these pathways to stem the tide of rising drug resistance. PMID:24858083

  11. 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)

    Mekonen, Mekdem; Abate, Ebba; Aseffa, Abraham; Anagaw, Belay; Elias, Daniel; Hailu, Elena; Idh, Jonna; Moges, Feleke; Wolde-Amanuel, Yimtubezinash; Asrat, Daniel; Yamuah, Lawrence; Britton, Sven; Stendahl, Olle; Schön, Thomas

    2010-01-01

    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....

  12. Human TYK2 deficiency: Mycobacterial and viral infections without hyper-IgE syndrome.

    Science.gov (United States)

    Kreins, Alexandra Y; Ciancanelli, Michael J; Okada, Satoshi; Kong, Xiao-Fei; Ramírez-Alejo, Noé; Kilic, Sara Sebnem; El Baghdadi, Jamila; Nonoyama, Shigeaki; Mahdaviani, Seyed Alireza; Ailal, Fatima; Bousfiha, Aziz; Mansouri, Davood; Nievas, Elma; Ma, Cindy S; Rao, Geetha; Bernasconi, Andrea; Sun Kuehn, Hye; Niemela, Julie; Stoddard, Jennifer; Deveau, Paul; Cobat, Aurelie; El Azbaoui, Safa; Sabri, Ayoub; Lim, Che Kang; Sundin, Mikael; Avery, Danielle T; Halwani, Rabih; Grant, Audrey V; Boisson, Bertrand; Bogunovic, Dusan; Itan, Yuval; Moncada-Velez, Marcela; Martinez-Barricarte, Ruben; Migaud, Melanie; Deswarte, Caroline; Alsina, Laia; Kotlarz, Daniel; Klein, Christoph; Muller-Fleckenstein, Ingrid; Fleckenstein, Bernhard; Cormier-Daire, Valerie; Rose-John, Stefan; Picard, Capucine; Hammarstrom, Lennart; Puel, Anne; Al-Muhsen, Saleh; Abel, Laurent; Chaussabel, Damien; Rosenzweig, Sergio D; Minegishi, Yoshiyuki; Tangye, Stuart G; Bustamante, Jacinta; Casanova, Jean-Laurent; Boisson-Dupuis, Stéphanie

    2015-09-21

    Autosomal recessive, complete TYK2 deficiency was previously described in a patient (P1) with intracellular bacterial and viral infections and features of hyper-IgE syndrome (HIES), including atopic dermatitis, high serum IgE levels, and staphylococcal abscesses. We identified seven other TYK2-deficient patients from five families and four different ethnic groups. These patients were homozygous for one of five null mutations, different from that seen in P1. They displayed mycobacterial and/or viral infections, but no HIES. All eight TYK2-deficient patients displayed impaired but not abolished cellular responses to (a) IL-12 and IFN-?/?, accounting for mycobacterial and viral infections, respectively; (b) IL-23, with normal proportions of circulating IL-17(+) T cells, accounting for their apparent lack of mucocutaneous candidiasis; and (c) IL-10, with no overt clinical consequences, including a lack of inflammatory bowel disease. Cellular responses to IL-21, IL-27, IFN-?, IL-28/29 (IFN-?), and leukemia inhibitory factor (LIF) were normal. The leukocytes and fibroblasts of all seven newly identified TYK2-deficient patients, unlike those of P1, responded normally to IL-6, possibly accounting for the lack of HIES in these patients. The expression of exogenous wild-type TYK2 or the silencing of endogenous TYK2 did not rescue IL-6 hyporesponsiveness, suggesting that this phenotype was not a consequence of the TYK2 genotype. The core clinical phenotype of TYK2 deficiency is mycobacterial and/or viral infections, caused by impaired responses to IL-12 and IFN-?/?. Moreover, impaired IL-6 responses and HIES do not appear to be intrinsic features of TYK2 deficiency in humans. PMID:26304966

  13. A chemically synthesized peptide which elicits humoral and cellular immune responses to mycobacterial antigens.

    OpenAIRE

    Minden, P; Houghten, R A; Spear, J R; Shinnick, T M

    1986-01-01

    Monoclonal antibodies directed to Mycobacterium bovis BCG (BCG) and to M. tuberculosis H37Rv (H37Rv) were used in conjunction with affinity chromatography to prepare a mycobacterial component which was designated BCG-a. A synthetic peptide antigen was prepared based on the amino acid sequence of BCG-a and was designated BCG-a-P. Significant immunological similarities were found between BCG-a-P and antigens in extracts of BCG and H37Rv but not between BCG-a-P and antigens of nontuberculous myc...

  14. 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)

    Diana, Castaño; Mauricio, Rojas.

    2010-06-01

    Full Text Available 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.

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

    Directory of Open Access Journals (Sweden)

    Diana Castaño

    2010-08-01

    Full Text Available 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 mayor 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.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. This 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.

  16. A murine monoclonal antibody to glycogen: characterization of epitope-fine specificity by saturation transfer difference (STD) NMR spectroscopy and its use in mycobacterial capsular α-glucan research.

    Science.gov (United States)

    van de Weerd, Robert; Berbís, M Alvaro; Sparrius, Marrion; Maaskant, Janneke J; Boot, Maikel; Paauw, Nanne J; de Vries, Nadine; Boon, Louis; Baba, Otto; Cañada, F Javier; Geurtsen, Jeroen; Jiménez-Barbero, Jesús; Appelmelk, Ben J

    2015-04-13

    Mycobacterium tuberculosis, the causative agent of tuberculosis (TB), is a major pathogen responsible for 1.5 million deaths annually. This bacterium is characterized by a highly unusual and impermeable cell envelope, which plays a key role in mycobacterial survival and virulence. Although many studies have focused on the composition and functioning of the mycobacterial cell envelope, the capsular α-glucan has received relatively minor attention. Here we show that a murine monoclonal antibody (Mab) directed against glycogen cross-reacts with mycobacterial α-glucans, polymers of α(1-4)-linked glucose residues with α(1-6)-branch points. We identified the Mab epitope specificity by saturation transfer difference NMR and show that the α(1-4)-linked glucose residues are important in glucan-Mab interaction. The minimal epitope is formed by (linear) maltotriose. Notably, a Mycobacterium mutant lacking the branching enzyme GlgB does not react with the Mab; this suggests that the α(1-6)-branches form part of the epitope. These seemingly conflicting data can be explained by the fact that in the mutant the linear form of the α-glucan (amylose) is insoluble. This Mab was subsequently used to develop several techniques helpful in capsular α-glucan research. By using a capsular glucan-screening methodology based on this Mab we were able to identify several unknown genes involved in capsular α-glucan biogenesis. Additionally, we developed two methods for the detection of capsular α-glucan levels. This study therefore opens new ways to study capsular α-glucan and to identify possible targets for further research. PMID:25766777

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

    Directory of Open Access Journals (Sweden)

    Previato Jose O

    2008-05-01

    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.

  18. 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

  19. Pyridoxal-phosphate dependent mycobacterial cysteine synthases: Structure, mechanism and potential as drug targets.

    Science.gov (United States)

    Schnell, Robert; Sriram, Dharmarajan; Schneider, Gunter

    2015-09-01

    The alarming increase of drug resistance in Mycobacterium tuberculosis strains poses a severe threat to human health. Chemotherapy is particularly challenging because M. tuberculosis can persist in the lungs of infected individuals; estimates of the WHO indicate that about 1/3 of the world population is infected with latent tuberculosis providing a large reservoir for relapse and subsequent spread of the disease. Persistent M. tuberculosis shows considerable tolerance towards conventional antibiotics making treatment particularly difficult. In this phase the bacilli are exposed to oxygen and nitrogen radicals generated as part of the host response and redox-defense mechanisms are thus vital for the survival of the pathogen. Sulfur metabolism and de novo cysteine biosynthesis have been shown to be important for the redox homeostasis in persistent M. tuberculosis and these pathways could provide promising targets for novel antibiotics for the treatment of the latent form of the disease. Recent research has provided evidence for three de novo metabolic routes of cysteine biosynthesis in M. tuberculosis, each with a specific PLP dependent cysteine synthase with distinct substrate specificities. In this review we summarize our present understanding of these pathways, with a focus on the advances on functional and mechanistic characterization of mycobacterial PLP dependent cysteine synthases, their role in the various pathways to cysteine, and first attempts to develop specific inhibitors of mycobacterial cysteine biosynthesis. This article is part of a Special Issue entitled: Cofactor-dependent proteins: evolution, chemical diversity and bio-applications. PMID:25484279

  20. Sensitivity and Specificity of Immunocytochemical Staining of Mycobacterial Antigens in the Cytoplasm of Cerebrospinal Fluid Macrophages for Diagnosing Tuberculous Meningitis ?

    OpenAIRE

    Shao, YuQuan; Xia, Ping; Zhu, Tao; Zhou, Jiong; Yuan, Yuan; Zhang, Hao; Chen, Jianjun; Hu, Xingyue

    2011-01-01

    The sensitivity and specificity of immunocytochemical staining of mycobacterial antigens in the cytoplasm of cerebrospinal fluid (CSF) macrophages for diagnosis of tuberculous meningitis (TBM) was prospectively compared with Ahuja criteria from 393 consecutive CSF specimens. The assay can play an important role for the diagnosis of TBM, with sensitivity of 73.5% and specificity of 90.7%.

  1. Detection of mycobacterial DNA by a specific and simple lateral flow assay incorporating cadmium selenide quantum dots.

    Science.gov (United States)

    Cimaglia, Fabio; Liandris, Emmanouil; Gazouli, Maria; Sechi, Leonardo; Chiesa, Maurizio; De Lorenzis, Enrico; Andreadou, Margarita; Taka, Styliani; Mataragka, Antonia; Ikonomopoulos, John

    2015-12-01

    Cadmium selenide quantum dots have been incorporated to a lateral flow assay for the specific and very simple detection of different mycobacterial DNA targets within only a few minutes, bypassing the complexity of conventional DNA hybridization assays. The method extends our previous work on protein detection using an identical procedure. PMID:26070989

  2. 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 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

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

    Energy Technology Data Exchange (ETDEWEB)

    Buckingham, S.J.; Haddow, L.J.; Shaw, P.J.; Miller, R.F. E-mail: rmiller@gum.ucl.ac.uk

    2004-06-01

    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 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.

  4. Leveraging Advances in Tuberculosis Diagnosis and Treatment to Address Nontuberculous Mycobacterial Disease.

    Science.gov (United States)

    Raju, Ravikiran M; Raju, Sagar M; Zhao, Yanlin; Rubin, Eric J

    2016-03-01

    The nontuberculous mycobacteria (NTM), defined as any mycobacterial pathogen other than Mycobacterium tuberculosis or Mycobacterium leprae, are a diverse group of pathogens that collectively cause a substantive but often unappreciated worldwide burden of illness. Although NTMs may cause illness similar to M. tuberculosis, these pathogens generally do not respond to classic tuberculosis (TB) drug regimens, resulting in misdiagnosis and poor treatment, particularly in resource-poor settings. Although a few high-quality epidemiologic surveys have been made on the topic, existing evidence suggests that NTM-associated disease is much more common than previously thought: more common than TB in the industrialized world and likely increasing in prevalence globally. Despite this evidence, these organisms remain markedly understudied, and few international grants support basic science and clinical research. Here we suggest that the considerable efforts in developing new treatments and diagnostics for TB can be harnessed in the fight against NTM-associated illnesses. PMID:26886068

  5. Severe mycobacterial and Salmonella infections in interleukin-12 receptor-deficient patients.

    Science.gov (United States)

    de Jong, R; Altare, F; Haagen, I A; Elferink, D G; Boer, T; van Breda Vriesman, P J; Kabel, P J; Draaisma, J M; van Dissel, J T; Kroon, F P; Casanova, J L; Ottenhoff, T H

    1998-05-29

    Interleukin-12 (IL-12) is a cytokine that promotes cell-mediated immunity to intracellular pathogens by inducing type 1 helper T cell (TH1) responses and interferon-gamma (IFN-gamma) production. IL-12 binds to high-affinity beta1/beta2 heterodimeric IL-12 receptor (IL-12R) complexes on T cell and natural killer cells. Three unrelated individuals with severe, idiopathic mycobacterial and Salmonella infections were found to lack IL-12Rbeta1 chain expression. Their cells were deficient in IL-12R signaling and IFN-gamma production, and their remaining T cell responses were independent of endogenous IL-12. IL-12Rbeta1 sequence analysis revealed genetic mutations that resulted in premature stop codons in the extracellular domain. The lack of IL-12Rbeta1 expression results in a human immunodeficiency and shows the essential role of IL-12 in resistance to infections due to intracellular bacteria. PMID:9603733

  6. Mycobacterial bone marrow infections at a medical centre in Taiwan, 2001-2009.

    Science.gov (United States)

    Lin, S-H; Lai, C-C; Huang, S-H; Hung, C-C; Hsueh, P-R

    2014-07-01

    Mycobacterial bone marrow (BM) infection is the most common diagnosis established by BM examinations for fever of unknown origin. In this study, clinical features and outcomes of patients who fulfilled the criteria for BM infection due to Mycobacterium tuberculosis (MTB) and non-tuberculous mycobacteria (NTM) at a medical centre in Taiwan from 2001 to 2009 were investigated. The BM histopathological findings were also analysed. A total of 24 patients (16 men, eight women) with mycobacterial BM infections were found. Of these, nine (38%) were positive for human immunodeficiency virus (HIV) and six (25%) had no pre-existing immunocompromised conditions. MTB isolates were obtained from 11 (46%) patients and NTM species were isolated from 10 (42%) patients, including M. avium complex (MAC, n = 7) and M. kansasii (n = 3). Patients with MTB infections were significantly older than those with NTM infections (60·5 vs. 47·7 years, P = 0·043) and were less likely to have a positive BM culture (45% vs. 100%, P = 0·012). The 90-day survival rates for MTB and NTM BM infections were 68% and 60%, respectively (P = 0·61). In addition, the presence of BM granulomas was significantly more common in patients with MTB BM infections than in those with NTM infections (82% vs. 30%, P = 0·030). In Taiwan, the importance of NTM was not inferior to MTB and besides MAC, M. kansasii might be an important pathogen in non-HIV-infected patients. The presence of BM granulomas and caseation provides valuable information regarding early treatment pending culture results. PMID:24168831

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

    Energy Technology Data Exchange (ETDEWEB)

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

    1984-10-01

    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.

  8. 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 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

  9. Presence of mycobacterial L-forms in human blood: Challenge of BCG vaccination.

    Science.gov (United States)

    Markova, Nadya; Slavchev, Georgi; Michailova, Lilia

    2015-01-01

    Possible persistence of bacteria in human blood as cell wall deficient forms (L-forms) represents a top research priority for microbiologists. Application of live BCG vaccine and L-form transformation of vaccine strain may display a new intriguing aspect concerning the opportunity for occurrence of unpredictable colonization inside the human body by unusual microbial life forms. L-form cultures were isolated from 141 blood samples of people previously vaccinated with BCG, none with a history of exposure to tuberculosis. Innovative methodology to access the unusual L-form elements derived from human blood was developed. The methodology outlines the path of transformation of non- cultivable L-form element to cultivable bacteria and their adaptation for growth in vitro. All isolates showed typical L-forms growth features ("fried eggs" colonies and biofilm). Electron microscopy revealed morphology evidencing peculiar characteristics of bacterial L-form population (cell wall deficient polymorphic elements of variable shape and size). Regular detection of acid fast bacteria in smears of isolated blood L-form cultures, led us to start their identification by using specific Mycobactrium spp. genetic tests. Forty five of 97 genetically tested blood cultures provided specific positive signals for mycobacteria, confirmed by at least one of the 3 specific assays (16S rRNA PCR; IS6110 Real Time PCR and spoligotyping). In conclusion, the obtained genetic evidence suggests that these L-forms are of mycobacterial origin. As the investigated people had been vaccinated with BCG, we can assume that the identified mycobacterial L-forms may be produced by persisting live BCG vaccine. PMID:25874947

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

    DEFF Research Database (Denmark)

    Thakur, Aneesh; Sharma, Mandeep

    2012-01-01

    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.

  11. Reduced in vivo cytotoxicity and increased Mycobacterial burden are associated with virulent Mycobacterium tuberculosis strains during lung infection.

    Science.gov (United States)

    Quintero-Macías, Liz; Silva-Sánchez, Aarón; Valderrabano-Ortíz, Estela; Munguía-Fuentes, Rosario; Aguilar-León, Diana; Hernández-Pando, Rogelio; Flores-Romo, Leopoldo

    2012-01-01

    Cytotoxic cellular responses are crucial for clearing intracellular pathogens and generating host resistance. Experimental pulmonary tuberculosis is associated with an early delay in T cell responses and with elevated lung bacterial burden during chronic infection. In this study we quantified the in vivo cytotoxicity and the mycobacterial burden from two pertinent tissues in groups of mice infected each with a mycobacterial strain of different virulence. None of the strains induced cytotoxic responses during early (day 14) infection. Interestingly, at 21 and 60 days post-infection, Mycobacterium canettii (lowest virulence) triggered the strongest in vivo cytotoxicity both in lungs and mediastinal lymph nodes. In contrast, Mycobacterium tuberculosis H37Rv (intermediate virulence) and Beijing strains (highest virulence) induced lower cytotoxic responses, and exhibited high bacterial growth, especially in lungs. These in vivo data suggest that virulence of Mycobacterium strains are somehow associated with subverting cytotoxic responses, thus contributing to early bacterial replication and subsequent persistence in the lungs. PMID:21635179

  12. Exploring the Structure and Function of the Mycobacterial KatG Protein Using trans-Dominant Mutants

    OpenAIRE

    DeVito, Joseph A.; Morris, Sheldon

    2003-01-01

    In order to probe the structure and function of the mycobacterial catalase-peroxidase enzyme (KatG), we employed a genetic approach using dominant-negative analysis of katG merodiploids. Transformation of Mycobacterium bovis BCG with various katG point mutants (expressed from low-copy-number plasmids) resulted in reductions in peroxidase and catalase activities as measured in cell extracts. These reductions in enzymatic activity usually correlated with increased resistance to the antitubercul...

  13. ESX-1-induced apoptosis during mycobacterial infection: to be or not to be, that is the question

    OpenAIRE

    Aguiló, Nacho; Marinova, Dessislava; Martín, Carlos; Pardo, Julián

    2013-01-01

    The major Mycobacterium tuberculosis virulence factor ESAT-6 exported by the ESX-1 secretion system has been described as a pro-apoptotic factor by several independent groups in recent years, sustaining a role for apoptosis in M. tuberculosis pathogenesis. This role has been supported by independent studies in which apoptosis has been shown as a hallmark feature in human and mouse lungs infected with virulent strains. Nevertheless, the role of apoptosis during mycobacterial infection is subje...

  14. Detection of 2-eicosanol by gas chromatography-mass spectrometry in sputa from patients with pulmonary mycobacterial infections.

    OpenAIRE

    Alugupalli, S.; Olsson, B; Larsson, L.

    1993-01-01

    A total of 96 sputum specimens from patients with suspected or known mycobacterial and nonmycobacterial pulmonary infections were analyzed by gas chromatography-mass spectrometry for the presence of 2-eicosanol. This secondary alcohol was detected in all of the 25 sputum specimens culture positive for Mycobacterium tuberculosis, in 7 of the 9 sputum specimens culture positive for M. avium complex, and in all 3 of the studied sputum specimens associated with M. malmoense. The alcohol was not d...

  15. Two-dimensional gas chromatography with electron capture detection for the sensitive determination of specific mycobacterial lipid constituents.

    OpenAIRE

    Larsson, L; Jimenez, J.; Sonesson, A; F. Portaels

    1989-01-01

    A method was developed for determining two characteristic mycobacterial lipid constituents, tuberculostearic acid (as its pentafluorobenzyl ester) and 2-eicosanol (as its pentafluorobenzoyl ester), by using gas chromatography with electron capture detection. A microprocessor-controlled column-switching system (two-dimensional gas chromatography) facilitated sample preparation and increased specificity. The usefulness of the technique was illustrated by its ability to reveal picogram amounts o...

  16. Rapid diagnosis of mycobacterial infections and quantitation of Mycobacterium tuberculosis load by two real-time calibrated PCR assays.

    Science.gov (United States)

    Broccolo, Francesco; Scarpellini, Paolo; Locatelli, Giuseppe; Zingale, Anna; Brambilla, Anna M; Cichero, Paola; Sechi, Leonardo A; Lazzarin, Adriano; Lusso, Paolo; Malnati, Mauro S

    2003-10-01

    Sensitive and specific techniques to detect and identify Mycobacterium tuberculosis directly in clinical specimens are important for the diagnosis and management of patients with tuberculosis (TB). We developed two real-time PCR assays, based on the IS6110 multicopy element and on the senX3-regX3 intergenic region, which provide a rapid method for the diagnosis of mycobacterial infections. The sensitivity and specificity of both assays were established by using purified DNA from 71 clinical isolates and 121 clinical samples collected from 83 patients, 20 of whom were affected by TB. Both assays are accurate, sensitive, and specific, showing a complementary pattern of Mycobacterium recognition: broader for the IS6110-based assay and restricted to the M. tuberculosis complex for the senX3-regX3-based assay. Moreover, the addition of a synthetic DNA calibrator prior to DNA extraction allowed us to measure the efficiency of DNA recovery and to control for the presence of PCR inhibitors. The mycobacterial burden of the clinical samples, as assessed by direct microscopy, correlates with the M. tuberculosis DNA load measured by the senX3-regX3-based assay. In addition, reduced levels of M. tuberculosis DNA load are present in those patients subjected to successful therapy, suggesting a potential use of this assay for monitoring treatment efficacy. Therefore, these assays represent a fully controlled high-throughput system for the evaluation of mycobacterial burden in clinical specimens. PMID:14532183

  17. Structure of the mycobacterial ATP synthase Fo rotor ring in complex with the anti-TB drug bedaquiline

    Science.gov (United States)

    Preiss, Laura; Langer, Julian D.; Yildiz, Özkan; Eckhardt-Strelau, Luise; Guillemont, Jérôme E. G.; Koul, Anil; Meier, Thomas

    2015-01-01

    Multidrug-resistant tuberculosis (MDR-TB) is more prevalent today than at any other time in human history. Bedaquiline (BDQ), a novel Mycobacterium-specific adenosine triphosphate (ATP) synthase inhibitor, is the first drug in the last 40 years to be approved for the treatment of MDR-TB. This bactericidal compound targets the membrane-embedded rotor (c-ring) of the mycobacterial ATP synthase, a key metabolic enzyme required for ATP generation. We report the x-ray crystal structures of a mycobacterial c9 ring without and with BDQ bound at 1.55- and 1.7-Å resolution, respectively. The structures and supporting functional assays reveal how BDQ specifically interacts with the rotor ring via numerous interactions and thereby completely covers the c-ring’s ion-binding sites. This prevents the rotor ring from acting as an ion shuttle and stalls ATP synthase operation. The structures explain how diarylquinoline chemicals specifically inhibit the mycobacterial ATP synthase and thus enable structure-based drug design of next-generation ATP synthase inhibitors against Mycobacterium tuberculosis and other bacterial pathogens.

  18. Clinical Usefulness of PCR for Differential Diagnosis of Tuberculosis and Nontuberculous Mycobacterial Infection in Paraffin-Embedded Lung Tissues.

    Science.gov (United States)

    Kim, Yo Na; Kim, Kyoung Min; Choi, Ha Na; Lee, Ju Hyung; Park, Ho Sung; Jang, Kyu Yun; Moon, Woo Sung; Kang, Myoung Jae; Lee, Dong Geun; Chung, Myoung Ja

    2015-09-01

    The need for isolation of nontuberculous mycobacteria (NTM) from clinical specimens has increased in recent years. Our aim was to determine the clinical usefulness of PCR for differential diagnosis of tuberculosis and nontuberculous mycobacterial infection in lung tissue that show chronic granulomatous inflammation. A total of 199 formalin-fixed, paraffin-embedded specimens, including 137 Mycobacterium tuberculosis (MTB), 17 NTM cases, and 45 other than mycobacterial cases were collected. We performed acid-fast staining, MTB and NTM nested PCRs, and MTB and NTM real-time PCRs. No histologic difference between MTB and NTM infections was observed. Sensitivity and specificity for detecting MTB were 70.1% and 95.1% by nested PCR, respectively, and 70.8% and 100.0% by real-time PCR, respectively. Sensitivity and specificity for detecting NTM were 52.9% and 96.15% by nested PCR, respectively, and 35.3% and 100.0% by real-time PCR, respectively. Mycobacteria were identified by acid-fast staining in 50 of 154 cases (32.5%). All 50 acid-fast staining-positive cases showed positive nested and real-time PCR results (n = 47 MTB PCR positive; n = 3 NTM PCR positive), and results agreed with final diagnosis. PCR will be useful for the rapid diagnosis of mycobacterial infection and differentiation of MTB from NTM in formalin-fixed, paraffin-embedded specimens, especially in acid-fast staining-positive specimens. PMID:26163897

  19. A lacZ Reporter-Based Strategy for Rapid Expression Analysis and Target Validation of Mycobacterium tuberculosis Latent Infection Genes.

    Science.gov (United States)

    Sood, Shivani; Kaur, Satinder; Shrivastava, Rahul

    2016-02-01

    We report a novel lacZ fusion vector and demonstrate its utility for expression analysis of genes associated with Mycobacterium tuberculosis latent infection. The vector contains E. coli (oriE) and mycobacterial (oriM) origins of replication, a kanamycin resistance gene (Km(r)) as selection marker, and a lacZ reporter gene in fusion with MCS for cloning of upstream regulatory sequence of the desired genes. ?-galactosidase activity of the vector was standardized for expression analysis under latent mycobacterial conditions using Phsp60, a constitutive mycobacterial promoter, utilizing Mycobacterium smegmatis as model organism. Validation of the vector was done by cloning and expression analysis of PhspX (alpha crystalline) and Picl (isocitrate lyase), promoters from two of the genes shown to be involved in M. tuberculosis persistence. Both genes showed appreciable levels of ?-galactosidase expression under hypoxia-induced persistent conditions in comparison to their actively replicating state. Expression analysis of a set of hypothetical genes was also done, of which Rv0628c showed increased expression under persistent conditions. The reported fusion vector and the strategy can be effectively used for short listing and validation of drug targets deduced from various non-conclusive approaches such as bioinformatics and microarray analysis against latent/persistent form of mycobacterial infection. PMID:26597215

  20. Nontuberculous Mycobacterial (NTM) Disease in Immunocompetent Patients: Expanding Image Findings on Chest CT

    Energy Technology Data Exchange (ETDEWEB)

    Shin, Hyo Hyun; Seon, Hyun Ju; Kim, Mok Hee; Choi, Song; Song, Sang Gook; Shin, Sang Soo; Kim, Yun Hyeon; Park, Jin Gyoon [Chonnam National University Hospital, Gwangju (Korea, Republic of)

    2010-04-15

    The aim of this study was to evaluate the chest CT features of nontuberculous mycobacterial (NTM) disease regardless of the specific organisms. This study included 74 consecutive patients (35 men, 39 women; mean age, 63 years; age range, 25-89 years) who were diagnosed with NTM disease according to the American Thoracic Society Guidelines (1997 and 2007) between January 2005 and July 2007. Chest CT images were randomly reviewed by two radiologists with consensus. The most common organism associated with NTM disease is M. avium-intracellulare complex (87.8%), followed by M. abscesses, M. kansasii, and M. chelonae. The most common chest CT finding was a nodular bronchiectatic lesion (n = 35, 46.7%), followed by a cavitary lesion of the upper lobe (n = 21, 28.0%), combined lesions of two prior subtypes (n = 6, 8.0%), consolidative lesion (s) (n = 5, 6.7%), a bronchogenic spreading pulmonary tuberculosis-like lesion (n = 5, 6.7%), a cavitary mass lesion with small satellite nodules (n = 2, 2.7%), and a miliary nodular lesion (n = 1, 1.3%). More than 5 segments were involved in 60 cases (81.1%). The nodular bronchiectatic lesion or cavitary lesion of upper lobe presents with multi-segmental involvement and the occurrence of combined consolidation is indicative of NTM disease

  1. Nontuberculous Mycobacterial (NTM) Disease in Immunocompetent Patients: Expanding Image Findings on Chest CT

    International Nuclear Information System (INIS)

    The aim of this study was to evaluate the chest CT features of nontuberculous mycobacterial (NTM) disease regardless of the specific organisms. This study included 74 consecutive patients (35 men, 39 women; mean age, 63 years; age range, 25-89 years) who were diagnosed with NTM disease according to the American Thoracic Society Guidelines (1997 and 2007) between January 2005 and July 2007. Chest CT images were randomly reviewed by two radiologists with consensus. The most common organism associated with NTM disease is M. avium-intracellulare complex (87.8%), followed by M. abscesses, M. kansasii, and M. chelonae. The most common chest CT finding was a nodular bronchiectatic lesion (n = 35, 46.7%), followed by a cavitary lesion of the upper lobe (n = 21, 28.0%), combined lesions of two prior subtypes (n = 6, 8.0%), consolidative lesion (s) (n = 5, 6.7%), a bronchogenic spreading pulmonary tuberculosis-like lesion (n = 5, 6.7%), a cavitary mass lesion with small satellite nodules (n = 2, 2.7%), and a miliary nodular lesion (n = 1, 1.3%). More than 5 segments were involved in 60 cases (81.1%). The nodular bronchiectatic lesion or cavitary lesion of upper lobe presents with multi-segmental involvement and the occurrence of combined consolidation is indicative of NTM disease

  2. Evaluation of the humoral response against mycobacterial peptides, homologous to MOG?????, in multiple sclerosis patients.

    Science.gov (United States)

    Cossu, Davide; Mameli, Giuseppe; Masala, Speranza; Cocco, Eleonora; Frau, Jessica; Marrosu, Maria Giovanna; Sechi, Leonardo Antonio

    2014-12-15

    Bacillus Calmette-Guérin (BCG) and Mycobacterium avium subspecies paratuberculosis (MAP) have been associated with multiple sclerosis (MS). Clinical data indicates that BCG vaccination exerts anti-inflammatory effects in MS; conversely, MAP is thought to be one of the possible infectious factors responsible of MS through a molecular mimicry mechanism. A peptide-based indirect ELISA was used to detect antibodies against the encephalitogenic myelin oligodendrocyte glycoprotein (MOG)35-55 epitope, and two mycobacterial peptides sharing sequence homology with the latter: MAP_2619c352-361/BCG_1224355-364 and BCG_3329c64-74. Among 40 MS patients and 39 healthy volunteers included in the study, only MOG35-55 was capable of inducing a significantly higher humoral response in MS subjects compared to controls. Indeed, 11 out of 40 MS subjects (27.5%) and only 2 out of 39 controls (5%) were antibody-positive for MOG35-55 (p=0.01, AUC=0.65). These findings strengthen the importance of MOG35-55 in MS pathogenesis. The MAP and BCG MOG-homologues epitopes investigated were not recognized in MS patients. Overall, the results allow us concluding that sharing homology of linear epitopes is necessary but not sufficient to induce antibody-mediated cross-reactivity. PMID:25271190

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

    Science.gov (United States)

    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

    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

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

    Science.gov (United States)

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

    2015-01-01

    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

  5. Nosocomial rapidly growing mycobacterial infections following laparoscopic surgery: CT imaging findings

    International Nuclear Information System (INIS)

    To identify the distribution and frequency of computed tomography (CT) findings in patients with nosocomial rapidly growing mycobacterial (RGM) infection after laparoscopic surgery. A descriptive retrospective study in patients with RGM infection after laparoscopic surgery who underwent CT imaging prior to initiation of therapy. The images were analyzed by two radiologists in consensus, who evaluated the skin/subcutaneous tissues, the abdominal wall, and intraperitoneal region separately. The patterns of involvement were tabulated as: densification, collections, nodules (?1.0 cm), small nodules (<1.0 cm), pseudocavitated nodules, and small pseudocavitated nodules. Twenty-six patients met the established criteria. The subcutaneous findings were: densification (88.5 %), small nodules (61.5 %), small pseudocavitated nodules (23.1 %), nodules (38.5 %), pseudocavitated nodules (15.4 %), and collections (26.9 %). The findings in the abdominal wall were: densification (61.5 %), pseudocavitated nodules (3.8 %), and collections (15.4 %). The intraperitoneal findings were: densification (46.1 %), small nodules (42.3 %), nodules (15.4 %), and collections (11.5 %). Subcutaneous CT findings in descending order of frequency were: densification, small nodules, nodules, small pseudocavitated nodules, pseudocavitated nodules, and collections. The musculo-fascial plane CT findings were: densification, collections, and pseudocavitated nodules. The intraperitoneal CT findings were: densification, small nodules, nodules, and collections. (orig.)

  6. Nosocomial rapidly growing mycobacterial infections following laparoscopic surgery: CT imaging findings

    Energy Technology Data Exchange (ETDEWEB)

    Volpato, Richard [Cassiano Antonio de Moraes University Hospital, Department of Diagnostic Radiology, Vitoria, ES (Brazil); Campi de Castro, Claudio [University of Sao Paulo Medical School, Department of Radiology, Cerqueira Cesar, Sao Paulo (Brazil); Hadad, David Jamil [Cassiano Antonio de Moraes University Hospital, Nucleo de Doencas Infecciosas, Department of Internal Medicine, Vitoria, ES (Brazil); Silva Souza Ribeiro, Flavya da [Laboratorio de Patologia PAT, Department of Diagnostic Radiology, Unit 1473, Vitoria, ES (Brazil); Filho, Ezequiel Leal [UNIMED Diagnostico, Department of Diagnostic Radiology, Unit 1473, Vitoria, ES (Brazil); Marcal, Leonardo P. [The University of Texas M D Anderson Cancer Center, Department of Diagnostic Radiology, Unit 1473, Houston, TX (United States)

    2015-09-15

    To identify the distribution and frequency of computed tomography (CT) findings in patients with nosocomial rapidly growing mycobacterial (RGM) infection after laparoscopic surgery. A descriptive retrospective study in patients with RGM infection after laparoscopic surgery who underwent CT imaging prior to initiation of therapy. The images were analyzed by two radiologists in consensus, who evaluated the skin/subcutaneous tissues, the abdominal wall, and intraperitoneal region separately. The patterns of involvement were tabulated as: densification, collections, nodules (≥1.0 cm), small nodules (<1.0 cm), pseudocavitated nodules, and small pseudocavitated nodules. Twenty-six patients met the established criteria. The subcutaneous findings were: densification (88.5 %), small nodules (61.5 %), small pseudocavitated nodules (23.1 %), nodules (38.5 %), pseudocavitated nodules (15.4 %), and collections (26.9 %). The findings in the abdominal wall were: densification (61.5 %), pseudocavitated nodules (3.8 %), and collections (15.4 %). The intraperitoneal findings were: densification (46.1 %), small nodules (42.3 %), nodules (15.4 %), and collections (11.5 %). Subcutaneous CT findings in descending order of frequency were: densification, small nodules, nodules, small pseudocavitated nodules, pseudocavitated nodules, and collections. The musculo-fascial plane CT findings were: densification, collections, and pseudocavitated nodules. The intraperitoneal CT findings were: densification, small nodules, nodules, and collections. (orig.)

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

    Directory of Open Access Journals (Sweden)

    Udo Conrad

    2010-01-01

    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.

  8. Therapeutic DNA Vaccine Reduces Schistosoma mansoni–Induced Tissue Damage through Cytokine Balance and Decreased Migration of Myofibroblasts

    OpenAIRE

    Frantz, Fabiani Gai; Ito, Toshihiro; Cavassani, Karen Angélica; Hogaboam, Cory M.; Lopes Silva, Célio; Kunkel, Steven L; Faccioli, Lúcia H

    2011-01-01

    Helminths are known to elicit a wide range of immunomodulation characterized by dominant Th2-type immune responses. Our group previously showed that a DNA vaccine encoding the mycobacterial 65-kDa heat shock protein (DNA-hsp65) showed immunomodulatory properties. We also showed, using a helminth-tuberculosis (TB) co-infection model, that the DNA-hsp65 vaccine protected mice against TB. We next investigated the mechanistic role of the vaccine during helminth-TB co-infection. Clinically, helmin...

  9. Towards new antituberculotic targets: biochemical characterisation of mycobacterial RNase E/G

    Directory of Open Access Journals (Sweden)

    Agnes Csanadi

    2008-06-01

    Full Text Available The World Health Organization estimates that each year 3 million people die from tuberculosis (TB and 8 million people become infected. No new anti-TB drugs have been introduced in the past 30 years, even though their development becomes increasingly important to face new challenges posed by multidrug-resistant and extensively drug-resistant strains and by acute infection with M. tuberculosis of HIV positive patients. Owing to its apparently important role in RNA metabolism, the RNase E/G family of endoribonucleases can be considered as a promising target for antimicrobial drugs. This consideration promted us to characterise biochemical properties of the M. tuberculosis RNase E/G homologue. To learn more about specific properties of RNase E/G homologues a M. tuberculosis RNase E/G (MycRne was overexpressed in E. coli and purified as a 6His-tagged polypeptide. To characterise MycRne, we used in vitro cleavage assays and primer extension analysis of total RNA extracted from mycobacteria. We show that affinity purified MycRne has an endoribonucleolytic activity, which is dependent on the 5'-phosphorylation status of RNA. We could also show that RNase E/G has Mg2+ dependent activity and similar to E. coli RNase E, MycRne was able to cleave in an intercistronic region of the putative 9S precursor of 5S rRNA. Although, similar to E. coli RNase E, the mycobacterial RNase E/G homologue plays a role in rRNA processing, the substrate specificities of these enzymes show differences. This suggests that RNase E/G can be used as a promising target for antimicrobial drugs that can be optimized to specifically target pathogenic species.

  10. Optimizing Mycobacterial Culture in Smear-Negative, Human Immunodeficiency Virus-Infected Tuberculosis Cases

    Science.gov (United States)

    Ismail, N. A.; Said, H. M.; Pinini, Z.; Omar, S. V.; Beyers, N.; Naidoo, P.

    2015-01-01

    Introduction Tuberculosis (TB) is a significant public health problem and the diagnosis in human immunodeficiency virus (HIV)—infected individuals is challenging. The use of mycobacterial culture remains an important complementary tool and optimizing it has important benefits. We sought to determine the effect of an increase in the number of specimens evaluated, addition of nutritional supplementation to the culture medium, sputum appearance and volume on diagnostic yield and time to detection of pulmonary TB among smear-negative, HIV-infected adults. Methods In this prospective study conducted at the Tshwane District Hospital and Academic TB Laboratory, Pretoria, South Africa we collected three sputum specimens an hour apart from presumptive TB cases at an antiretroviral treatment site. We analysed specimens from 236 patients. Specimen appearance and volume were recorded. All specimens were processed for culture using both standard and supplemented media. Results A single specimen identified 79% of PTB cases using standard media; the second and third specimens added 12.5% and 8.3% respectively. Media supplementation, sputum appearance and specimen volume had no effect on culture yield or contamination rates. The mean time to detection was reduced from 19.8 days in standard cultures to 11.8 days in nutrient supplemented cultures (p = 0.002). For every 1 ml increase in sputum volume, time to detection was decreased by a factor of 0.797 (p = 0.011). Conclusion Use of an inexpensive culture supplement substantially reduced time to detection and could contribute to reducing treatment delay among HIV-infected cases. PMID:26544183

  11. Nontuberculous Mycobacterial Infection in the Uterine Cervix Mimics Invasive Cervical Cancer in Immunocompetent Woman.

    Science.gov (United States)

    Ukita, Masayo; Aoki, Masato; Murakami, Kosuke; Takaya, Hisamitsu; Kotani, Yasushi; Shimaoka, Masao; Tobiume, Takako; Nakai, Hidekatsu; Tsuji, Isao; Suzuki, Ayako; Mandai, Masaki

    2016-03-01

    Nontuberculous mycobacterial (NTM) infection is increasing across the world. Although the most common clinical manifestation of NTM disease is lung disease, a rare form of disseminated NTM disease has also been documented. Disseminated NTM usually develops in severely immunocompromised individuals, especially those with advanced AIDS. This manifestation is rare in non-HIV-infected hosts and is associated with immunosuppressed conditions. However, recent reports have suggested that disseminated NTM disease in immunocompetent patients without HIV infection has been increasing. Dissemination may involve any organ system, but a case in the female genital tract has never been reported. We report a case in a 67-yr-old previously healthy woman who presented with a disseminated NTM infection in the uterine cervix. The primary presentation was general fatigue and body weight loss. The patient also presented with a mass formation that mimicked cervical cancer on magnetic resonance imaging. In addition to the cervical mass, the patient presented with a mass formation in the omentum; wall thickening of the vagina, bladder, and ureter; and retention of pleural/peritoneal fluid. Vaginal cytology was negative. A diagnosis was made only after detecting acid-fast bacilli in a biopsy specimen of cervical mass, which was conducted under suspicion of cervical malignancy. Then, Mycobacterium aviumwas confirmed in a polymerase chain reaction test of cervical tissue. After administration of antimycobacterial therapy, the mass and other findings on magnetic resonance imaging disappeared. Infection in multiple organs leads to the diagnosis of disseminated NTM. This case indicates that, for prompt and accurate diagnosis, efforts to detect specific lesions by an imaging study and to confirm diagnosis pathologically are equally important, especially when local cytology is not convincing. The clinical course of this case may serve as a useful reference in the diagnosis and treatment of NTM. PMID:26535986

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

    International Nuclear Information System (INIS)

    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)

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

    OpenAIRE

    Unciuleac, Mihaela-Carmen; Shuman, Stewart

    2009-01-01

    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 cofa...

  14. Limited Contribution of IL-36 versus IL-1 and TNF Pathways in Host Response to Mycobacterial Infection

    OpenAIRE

    Segueni, Noria; Vigne, Solenne; Palmer, Gaby; Bourigault, Marie-Laure; Olleros, Maria L.; Vesin, Dominique; Garcia, Irene; Ryffel, Bernhard; Quesniaux, Valérie F. J.; Gabay, Cem

    2015-01-01

    IL-36 cytokines are members of the IL-1 family of cytokines that stimulate dendritic cells and T cells leading to enhanced T helper 1 responses in vitro and in vivo; however, their role in host defense has not been fully addressed thus far. The objective of this study was to examine the role of IL-36R signaling in the control of mycobacterial infection, using models of systemic attenuated M. bovis BCG infection and virulent aerogenic M. tuberculosis infection. IL-36? expression was increased ...

  15. Nitric oxide inhibits the accumulation of CD4+CD44hiTbet+CD69lo T cells in mycobacterial infection

    Science.gov (United States)

    Pearl, John E.; Torrado, Egidio; Tighe, Michael; Fountain, Jeffrey J.; Solache, Alejandra; Strutt, Tara; Swain, Susan; Appelberg, Rui; Cooper, Andrea M

    2013-01-01

    Summary Animals lacking the inducible nitric oxide synthase gene (nos2?/?) are less susceptible to M. avium strain 25291 and lack nitric oxide-mediated immunomodulation of CD4+ T cells. Here we show that the absence of nos2 results in increased accumulation of neutrophils and both CD4+ and CD8+ T cells within the M. avium-containing granuloma. Examination of the T-cell phenotype in M. avium-infected mice demonstrated that CD4+CD44hi effector T cells expressing the Th1 transcriptional regulator T-bet (T-bet+) were specifically reduced by the presence of nitric oxide. Importantly, the T-bet+ effector population could be separated into CD69hi and CD69lo populations, with the CD69lo population only able to accumulate during chronic infection within infected nos2?/? mice. Transcriptomic comparison between CD4+CD44hiCD69hi and CD4+CD44hiCD69lo populations revealed that CD4+CD44hiCD69lo cells had higher expression of the integrin itgb1/itga4 (VLA-4, CD49d/CD29). Inhibition of Nos2 activity allowed increased accumulation of the CD4+CD44hiT-bet+CD69lo population in WT mice as well as increased expression of VLA-4. These data support the hypothesis that effector T cells in mycobacterial granulomata are not a uniform effector population but exist in distinct subsets with differential susceptibility to the regulatory effects of nitric oxide. PMID:22890814

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

    OpenAIRE

    Unciuleac, Mihaela-Carmen; Shuman, Stewart

    2010-01-01

    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 prevent...

  17. Mycobacterial secretion systems ESX-1 and ESX-5 play distinct roles in host cell death and inflammasome activation

    KAUST Repository

    Abdallah, Abdallah

    2011-09-28

    During infection of humans and animals, pathogenic mycobacteria manipulate the host cell causing severe diseases such as tuberculosis and leprosy. To understand the basis of mycobacterial pathogenicity, it is crucial to identify the molecular virulence mechanisms. In this study, we address the contribution of ESX-1 and ESX-5 - two homologous type VII secretion systems of mycobacteria that secrete distinct sets of immune modulators - during the macrophage infection cycle. Using wild-type, ESX-1- and ESX-5-deficient mycobacterial strains, we demonstrate that these secretion systems differentially affect subcellular localization and macrophage cell responses. We show that in contrast to ESX-1, the effector proteins secreted by ESX-5 are not required for the translocation of Mycobacterium tuberculosis or Mycobacterium marinum to the cytosol of host cells. However, the M. marinum ESX-5 mutant does not induce inflammasome activation and IL-1b activation. The ESX-5 system also induces a caspase-independent cell death after translocation has taken place. Importantly, by means of inhibitory agents and small interfering RNA experiments, we reveal that cathepsin B is involved in both the induction of cell death and inflammasome activation upon infection with wild-type mycobacteria. These results reveal distinct roles for two different type VII secretion systems during infection and shed light on how virulent mycobacteria manipulate the host cell in various ways to replicate and spread. Copyright © 2011 by The American Association of Immunologists, Inc.

  18. Effects of epigallocatechin gallate on the cell-wall structure of Mycobacterial smegmatis mc(2)155.

    Science.gov (United States)

    Sun, Tieying; Qin, Biaojie; Gao, Mingchuan; Yin, Yuling; Wang, Changyuan; Zang, Shizhu; Li, Xinli; Zhang, Cuili; Xin, Yi; Jiang, Tao

    2015-11-01

    Epigallocatechin gallate (EGCG) is the main component of green tea extracts that inhibits the growth of Mycobacterial smegmatis mc(2)155, and the mechanism is not clear. This study showed the effects of EGCG on the growth of mc(2)155. The content and the structure of EGCG in LB medium with mc(2)155 were identified by HPLC and LC/MS. Transmission electron microscopy was utilised to identify the cell envelope structure. As a result, the optional inhibition concentration was determined to be 20 ?g mL(- 1). Most of EGCG was transferred into its isomeride in LB medium, but the inhibition effects against mc(2)155 had yet been maintained. The changes of cell envelope structure were showed after EGCG treatment for 18 h. The cell wall appeared to have a less electron-translucent zone, turn rougher and thicker. The results show that EGCG impacts the integrity of mycobacterial cell wall and is likely be a better prophylactic agent against tuberculosis. PMID:25495515

  19. Limited Contribution of IL-36 versus IL-1 and TNF Pathways in Host Response to Mycobacterial Infection.

    Science.gov (United States)

    Segueni, Noria; Vigne, Solenne; Palmer, Gaby; Bourigault, Marie-Laure; Olleros, Maria L; Vesin, Dominique; Garcia, Irene; Ryffel, Bernhard; Quesniaux, Valérie F J; Gabay, Cem

    2015-01-01

    IL-36 cytokines are members of the IL-1 family of cytokines that stimulate dendritic cells and T cells leading to enhanced T helper 1 responses in vitro and in vivo; however, their role in host defense has not been fully addressed thus far. The objective of this study was to examine the role of IL-36R signaling in the control of mycobacterial infection, using models of systemic attenuated M. bovis BCG infection and virulent aerogenic M. tuberculosis infection. IL-36? expression was increased in the lung of M. bovis BCG infected mice. However, IL-36R deficient mice infected with M. bovis BCG showed similar survival and control of the infection as compared to wild-type mice, although their lung pathology and CXCL1 response were transiently different. While highly susceptible TNF-? deficient mice succumbed with overwhelming M. tuberculosis infection, and IL-1RI deficient mice showed intermediate susceptibility, IL-36R-deficient mice controlled the infection, with bacterial burden, lung inflammation and pathology, similar to wild-type controls. Therefore, IL-36R signaling has only limited influence in the control of mycobacterial infection. PMID:25950182

  20. In Vivo and In Vitro Effects of Antituberculosis Treatment on Mycobacterial Interferon-? T Cell Response

    Science.gov (United States)

    Sauzullo, Ilaria; Mengoni, Fabio; Lichtner, Miriam; Massetti, Anna Paola; Rossi, Raffaella; Iannetta, Marco; Marocco, Raffaella; Borgo, Cosmo Del; Soscia, Fabrizio; Vullo, Vincenzo; Mastroianni, Claudio Maria

    2009-01-01

    Background In recent years, the impact of antituberculous treatment on interferon (IFN)-? response to Mycobacterium tuberculosis antigens has been widely investigated, but the results have been controversial. The objective of the present study was: i) to evaluate longitudinal changes of IFN-? response to M. tuberculosis-specific antigens in TB patients during antituberculous treatment by using the QuantiFERON-TB Gold (QFT-G) assay; ii) to compare the differences in T-cell response after a short or prolonged period of stimulation with mycobacterial antigens; iii) to assess the CD4+ and CD8+ T cells with effector/memory and central/memory phenotype; iv) to investigate the direct in vitro effects of antituberculous drugs on the secretion of IFN-?. Principal Findings 38 TB patients was evaluated at baseline and at month 2 and 4 of treatment and at month 6 (treatment completion). 27 (71%) patients had a QFT-G reversion (positive to negative) at the end of therapy, while 11 (29%) TB patients remained QFT-G positive at the end of therapy. Among the 11 patients with persistent positive QFT-G results, six had a complete response to the treatment, while the remaining 5 patients did not have a resolution of the disease. All 27 patients who became QFT-G negative had a complete clinical and microbiological recovery of the TB disease. In these patients the release of IFN-? is absent even after a prolonged 6-day incubation with both ESAT-6 and CFP-10 antigens and the percentage of effector/memory T-cells phenotype was markedly lower than subjects with persistent positive QFT-G results. The in vitro study showed that antituberculous drugs did not exert any inhibitory effect on IFN-? production within the range of therapeutically achievable concentrations. Conclusions The present study suggests that the decrease in the M. tuberculosis-specific T cells responses following successful anti-TB therapy may have a clinical value as a supplemental tool for the monitoring of the efficacy of pharmacologic intervention for active TB. In addition, the antituberculous drugs do not have any direct down-regulatory effect on the specific IFN-? response. PMID:19365543

  1. 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)

    Cocaign, Angélique; Kubiak, Xavier Jean Philippe

    2014-01-01

    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.

  2. Antigen stimulation of peripheral blood mononuclear cells from Mycobacterium bovis infected cattle yields evidence for a novel gene expression program

    OpenAIRE

    Zhao Yingdong; Keane Joseph; Costello Eamon; Park Stephen D; O'Farrelly Cliona; Gormley Eamonn; Meade Kieran G; MacHugh David E

    2008-01-01

    Abstract Background Bovine tuberculosis (BTB) caused by Mycobacterium bovis continues to cause substantial losses to global agriculture and has significant repercussions for human health. The advent of high throughput genomics has facilitated large scale gene expression analyses that present a novel opportunity for revealing the molecular mechanisms underlying mycobacterial infection. Using this approach, we have previously shown that innate immune genes in peripheral blood mononuclear cells ...

  3. Cutaneous manifestations of Nocardia brasiliensis infection in Taiwan during 2002-2012-clinical studies and molecular typing of pathogen by gyrB and 16S gene sequencing.

    Science.gov (United States)

    Chen, Kuo-Wei; Lu, Chun-Wei; Huang, Ting-Chi; Lu, Chin-Fang; Liau, Yea-Ling; Lin, Jeng-Fong; Li, Shu-Ying

    2013-09-01

    To observe the clinicopathologic and resistance profiles of the Nocardia brasiliensis causing cutaneous nocardiosis in Taiwan, 12 N. brasiliensis isolates were prospectively collected from patients with cutaneous nocardiosis in a hospital during 2002-2012. Clinicopathologic data were obtained, and isolates were identified by biochemical methods and 16S rRNA sequencing. Susceptibilities to 14 antimicrobial compounds were tested. Isolates were further genotyped by sequencing of 16S rRNA, secA1, hsp65, and gyrB genes. The nodulopustular pyoderma associated with sporotrichoid spreading was the most common skin presentations caused by N. brasiliensis. All of the isolates were susceptible to amikacin, gentamicin, tobramycin, piperacillin/tazobactam, and trimethoprim/sulfamethoxazole and resistant to kanamycin, erythromycin, and oxacillin, while susceptibilities to imipenem, vancomycin, penicillin-G, tetracycline, clindamycin, and ciprofloxacin varied among the 12 isolates. GyrB genotyping delineated the 12 isolates into 2 major groups, which was coincident with different single nucleotide substitutions at position 160 (G versus T) of 16S rRNA, different levels of imipenem minimum inhibition concentration (4-32 versus 0.25-0.75 mg/L), and prevalence of lymphadenitis (66.7 versus 16.7%). We have noted that tiny pustular lesions can be the first sign of cutaneous nocardiosis, which we believe has not been previously emphasized. No resistance to trimethoprim and sulfamethoxazole was found; therefore, sulphonamide drugs remain effective for treatment of cutaneous nocardiosis in Taiwan. PMID:23791388

  4. CMRegNet-An interspecies reference database for corynebacterial and mycobacterial regulatory networks

    DEFF Research Database (Denmark)

    Abreu, Vinicius A C; Almeida, Sintia; Tiwari, Sandeep; Hassan, Syed Shah; Mariano, Diego; Silva, Artur; Baumbach, Jan; Azevedo, Vasco; Röttger, Richard

    2015-01-01

    gene regulatory network can lead to various practical applications, creating a greater understanding of how organisms control their cellular behavior. DESCRIPTION: In this work, we present a new database, CMRegNet for the gene regulatory networks of Corynebacterium glutamicum ATCC 13032 and...

  5. [Implementation of the technical requirements of the UNE-EN-ISO 15189 quality standard in a mycobacterial laboratory].

    Science.gov (United States)

    Guna Serrano, M del Remedio; Ocete Mochón, M Dolores; Lahiguera, M José; Bresó, M Carmen; Gimeno Cardona, Concepción

    2013-02-01

    The UNE-EN-ISO 15189:2007 standard defines the requirements for quality and competence that must be met by medical laboratories. These laboratories should use this international standard to develop their own quality management systems and to evaluate their own competencies; in turn, this standard will be used by accreditation bodies to confirm or recognize the laboratories' competence. In clinical microbiology laboratories, application of the standard implies the implementation of the technical and specific management requirements that must be met to achieve optimal quality when carrying out microbiological tests. In Spain, accreditation is granted by the Spanish Accreditation Body (Entidad Nacional de Acreditación). This review aims to discuss the practical application of the standard's technical requirements in mycobacterial laboratory. Firstly, we define the scope of accreditation. Secondly, we specify how the items of the standard on personnel management, control of equipment, environmental facilities, method validation, internal controls and customer satisfaction surveys were developed and implemented in our laboratory. PMID:23453231

  6. Techniques of DNA hybridization detect small numbers of mycobacteria with no cross-hybridization with non-mycobacterial respiratory organisms

    International Nuclear Information System (INIS)

    The traditional methods used in identifying mycobacteria, such as acid-fast bacillus stains and culture, are often time-consuming, insensitive, and nonspecific. As part of an ongoing program to improve diagnosis and characterization of mycobacteria, the authors have found that deoxyribonucleic acid (DNA) hybridization techniques using isotopically labeled, single-stranded, total DNA can be used to detect as little as 10(-4) micrograms of Mycobacterium tuberculosis (MTb) DNA. This amount of DNA represents approximately 2 X 10(4) genomes. They have also shown the MTb DNA is sufficiently different from the DNA of non-mycobacterial microorganisms such that cross-hybridization with MTb DNA does not occur under the hybridization conditions employed. The authors speculate that DNA hybridization techniques may allow the rapid, sensitive, and specific identification of mycobacteria

  7. The external PASTA domain of the essential serine/threonine protein kinase PknB regulates mycobacterial growth

    Science.gov (United States)

    Turapov, Obolbek; Loraine, Jessica; Jenkins, Christopher H.; Barthe, Philippe; McFeely, Daniel; Forti, Francesca; Ghisotti, Daniela; Hesek, Dusan; Lee, Mijoon; Bottrill, Andrew R.; Vollmer, Waldemar; Mobashery, Shahriar; Cohen-Gonsaud, Martin; Mukamolova, Galina V.

    2015-01-01

    PknB is an essential serine/threonine protein kinase required for mycobacterial cell division and cell-wall biosynthesis. Here we demonstrate that overexpression of the external PknB_PASTA domain in mycobacteria results in delayed regrowth, accumulation of elongated bacteria and increased sensitivity to ?-lactam antibiotics. These changes are accompanied by altered production of certain enzymes involved in cell-wall biosynthesis as revealed by proteomics studies. The growth inhibition caused by overexpression of the PknB_PASTA domain is completely abolished by enhanced concentration of magnesium ions, but not muropeptides. Finally, we show that the addition of recombinant PASTA domain could prevent regrowth of Mycobacterium tuberculosis, and therefore offers an alternative opportunity to control replication of this pathogen. These results suggest that the PknB_PASTA domain is involved in regulation of peptidoglycan biosynthesis and maintenance of cell-wall architecture. PMID:26136255

  8. Implementation of MALDI-TOF MS technology for the identification of clinical isolates of Mycobacterium spp. in mycobacterial diagnosis.

    Science.gov (United States)

    Tudó, G; Monté, M R; Vergara, A; López, A; Hurtado, J C; Ferrer-Navarro, M; Vila, J; Gonzalez-Martin, J

    2015-08-01

    A total of 243 clinical isolates of the Mycobacterium genus were studied, 143 and 100 using two protocols (Protocol v2 and Protocol v3, respectively) provided by the manufacturer. The overall correlation of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) with the standard identification methods was 63.8 %. The rate of misidentification was 3.2 %, mainly affecting very close species. In Protocol v2, the correlation was 57.3 %, being greater in solid than in liquid media (71.7 % vs. 44.7 %, p?mycobacteria (NTM) (63.6 % vs. 55.5 %) was observed. In Protocol v3, the correlation was 73 %, with no significant differences between solid and liquid media (70.8 % vs. 75 %). In conclusion, MALDI-TOF MS may play a role in identifying mycobacterial species isolated from clinical samples, being faster than sequencing and hybridization-based techniques. PMID:25912272

  9. A novel inhibitor of gyrase B is a potent drug candidate for treatment of tuberculosis and nontuberculosis mycobacterial infections.

    Science.gov (United States)

    Locher, Christopher P; Jones, Steven M; Hanzelka, Brian L; Perola, Emanuele; Shoen, Carolyn M; Cynamon, Michael H; Ngwane, Andile H; Wiid, Ian J; van Helden, Paul D; Betoudji, Fabrice; Nuermberger, Eric L; Thomson, John A

    2015-03-01

    New drugs to treat drug-resistant tuberculosis are urgently needed. Extensively drug-resistant and probably the totally drug-resistant tuberculosis strains are resistant to fluoroquinolones like moxifloxacin, which target gyrase A, and most people infected with these strains die within a year. In this study, we found that a novel aminobenzimidazole, VXc-486, which targets gyrase B, potently inhibits multiple drug-sensitive isolates and drug-resistant isolates of Mycobacterium tuberculosis in vitro (MICs of 0.03 to 0.30 ?g/ml and 0.08 to 5.48 ?g/ml, respectively) and reduces mycobacterial burdens in lungs of infected mice in vivo. VXc-486 is active against drug-resistant isolates, has bactericidal activity, and kills intracellular and dormant M. tuberculosis bacteria in a low-oxygen environment. Furthermore, we found that VXc-486 inhibits the growth of multiple strains of Mycobacterium abscessus, Mycobacterium avium complex, and Mycobacterium kansasii (MICs of 0.1 to 2.0 ?g/ml), as well as that of several strains of Nocardia spp. (MICs of 0.1 to 1.0 ?g/ml). We made a direct comparison of the parent compound VXc-486 and a phosphate prodrug of VXc-486 and showed that the prodrug of VXc-486 had more potent killing of M. tuberculosis than did VXc-486 in vivo. In combination with other antimycobacterial drugs, the prodrug of VXc-486 sterilized M. tuberculosis infection when combined with rifapentine-pyrazinamide and bedaquiline-pyrazinamide in a relapse infection study in mice. Furthermore, the prodrug of VXc-486 appeared to perform at least as well as the gyrase A inhibitor moxifloxacin. These findings warrant further development of the prodrug of VXc-486 for the treatment of tuberculosis and nontuberculosis mycobacterial infections. PMID:25534737

  10. High Mortality of Disseminated Non-Tuberculous Mycobacterial Infection in HIV-Infected Patients in the Antiretroviral Therapy Era

    Science.gov (United States)

    Kobayashi, Tetsuro; Nishijima, Takeshi; Teruya, Katsuji; Aoki, Takahiro; Kikuchi, Yoshimi; Oka, Shinichi; Gatanaga, Hiroyuki

    2016-01-01

    Background Little information is available on the mortality and risk factors associated with death in disseminated non-tuberculous mycobacterial infection (dNTM) in HIV-infected patients in the ART-era. Methods In a single-center study, HIV-infected dNTM with positive NTM culture from sterile sites between 2000 and 2013 were analysed. The clinical characteristics at commencement of anti-mycobacterial treatment (baseline) were compared between those who survived and died. Results Twenty-four patients were analyzed. [The median CD4 27/μL (range 2–185)]. Mycobacterium avium and M. intracellulare accounted for 20 (83%) and 3 (13%) of isolated NTM. NTM bacteremia was diagnosed in 15 (63%) patients. Seven (29%) patients died, and NTM bacteremia was significantly associated with mortality (p = 0.022). The baseline CD4 count was significantly lower in the non-survivors than the survivors (median 7/μL versus 49, p = 0.034). Concomitant AIDS-defining diseases or malignancies were not associated with mortality. Immune-reconstitution syndrome (IRS) occurred to 19 (79%) patients (8 paradoxical and 11 unmasking), and prognosis tended to be better in unmasking-IRS than the other patients (n = 13) (p = 0.078). Patients with paradoxical-IRS had marginally lower CD4 count and higher frequency of bacteremia than those with unmasking-IRS (p = 0.051, and 0.059). Treatment with systemic corticosteroids was applied in 63% and 55% of patients with paradoxical and unmasking-IRS, respectively. Conclusion dNTM in HIV-infected patients resulted in high mortality even in the ART-era. NTM bacteremia and low CD4 count were risk factors for death, whereas patients presented with unmasking-IRS had marginally better prognosis. IRS occurred in 79% of the patients, suggesting difficulty in the management of dNTM. PMID:26985832

  11. CMRegNet-An interspecies reference database for corynebacterial and mycobacterial regulatory networks

    DEFF Research Database (Denmark)

    Abreu, Vinicius A C; Almeida, Sintia; Tiwari, Sandeep; Hassan, Syed Shah; Mariano, Diego; Silva, Artur; Baumbach, Jan; Azevedo, Vasco; Röttger, Richard

    2015-01-01

    BACKGROUND: Organisms utilize a multitude of mechanisms for responding to changing environmental conditions, maintaining their functional homeostasis and to overcome stress situations. One of the most important mechanisms is transcriptional gene regulation. In-depth study of the transcriptional gene regulatory network can lead to various practical applications, creating a greater understanding of how organisms control their cellular behavior. DESCRIPTION: In this work, we present a new database,...

  12. Molecular characterisation of clinical and environmental isolates of Mycobacterium kansasii isolates from South African gold mines.

    Science.gov (United States)

    Kwenda, Geoffrey; Churchyard, Gavin J; Thorrold, Catherine; Heron, Ian; Stevenson, Karen; Duse, Adriano G; Marais, Elsé

    2015-03-01

    Mycobacterium kansasii (M. kansasii) is a major cause of non-tuberculous mycobacterial pulmonary disease in the South African gold-mining workforce, but the source of infection and molecular epidemiology are unknown. This study investigated the presence of M. kansasii in gold and coal mine and associated hostel water supplies and compared the genetic diversity of clinical and environmental isolates of M. kansasii. Five M. kansasii and ten other potentially pathogenic mycobacteria were cultured mainly from showerhead biofilms. Polymerase chain reaction-restriction analysis of the hsp65 gene on 196 clinical and environmental M. kansasii isolates revealed 160 subtype I, eight subtype II and six subtype IV strains. Twenty-two isolates did not show the typical M. kansasii restriction patterns, suggesting that these isolates may represent new subtypes of M. kansasii. In contrast to the clonal population structure found amongst the subtype I isolates from studies in other countries, DNA fingerprinting of 114 clinical and three environmental subtype I isolates demonstrated genetic diversity amongst the isolates. This study demonstrated that showerheads are possible sources of M. kansasii and other pathogenic non-tuberculous mycobacterial infection in a gold-mining region, that subtype I is the major clinical isolate of M. kansasii strain and that this subtype exhibits genetic diversity. PMID:25719478

  13. 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

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

    2014-01-01

    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...

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

    Directory of Open Access Journals (Sweden)

    Hill Adrian VS

    2009-02-01

    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.

  15. Rapid Mycobacterial Liquid Culture-Screening Method for Mycobacterium avium Complex Based on Secreted Antigen-Capture Enzyme-Linked Immunosorbent Assay?

    OpenAIRE

    Shin, Sung Jae; Anklam, Kelly; Manning, Elizabeth J.B.; Collins, Michael T.

    2009-01-01

    Sensors in automated liquid culture systems for mycobacteria, such as MGIT, BacT/Alert 3D, and Trek ESP II, flag growth of any type of bacteria; a positive signal does not mean that the target mycobacteria are present. All signal-positive cultures thus require additional and often laborious testing. An immunoassay was developed to screen liquid mycobacterial cultures for evidence of Mycobacterium avium complex (MAC). The method, called the MAC-enzyme-linked immunosorbent assay (ELISA), relies...

  16. Implementation of a Consensus Set of Hypervariable Mycobacterial Interspersed Repetitive-Unit-Variable-Number Tandem-Repeat Loci in Mycobacterium tuberculosis Molecular Epidemiology.

    Science.gov (United States)

    Trovato, Alberto; Tafaj, Silva; Battaglia, Simone; Alagna, Riccardo; Bardhi, Donika; Kapisyzi, Perlat; Bala, Silvana; Haldeda, Migena; Borroni, Emanuele; Hafizi, Hasan; Cirillo, Daniela Maria

    2016-02-01

    This study shows that the addition of a consensus 4-locus set of hypervariable mycobacterial interspersed repetitive-unit-variable-number tandem repeat (MIRU-VNTR) loci to the spoligotyping-24-locus MIRU-VNTR typing strategy is a well-standardized approach that can contribute to an improvement of the true cluster definition while retaining high typeability in non-Beijing strains. PMID:26659207

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

    OpenAIRE

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

    2011-01-01

    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 Spoligot...

  18. Sensitivities and Specificities of Spoligotyping and Mycobacterial Interspersed Repetitive Unit-Variable-Number Tandem Repeat Typing Methods for Studying Molecular Epidemiology of Tuberculosis

    OpenAIRE

    Scott, Allison N; Menzies, Dick; Tannenbaum, Terry-Nan; Thibert, Louise; Kozak, Robert; Joseph, Lawrence; Schwartzman, Kevin; Behr, Marcel A

    2005-01-01

    The development of PCR-based genotyping modalities (spoligotyping and mycobacterial interspersed repetitive unit-variable-number tandem repeat [MIRU-VNTR] typing) offers promise for real-time molecular epidemiological studies of tuberculosis (TB). However, the utility of these methods depends on their capacity to appropriately classify isolates. To determine the operating parameters of spoligotyping and MIRU-VNTR typing, we have compared results generated by these newer tests to the standard ...

  19. Assessment of an Optimized Mycobacterial Interspersed Repetitive- Unit-Variable-Number Tandem-Repeat Typing System Combined with Spoligotyping for Population-Based Molecular Epidemiology Studies of Tuberculosis? ‡

    OpenAIRE

    Oelemann, Mara Cardoso; Diel, Roland; Vatin, Vincent; Haas, Walter; Rüsch-Gerdes, Sabine; Locht, Camille; Niemann, Stefan; Supply, Philip

    2006-01-01

    An optimized set of 24 mycobacterial interspersed repetitive-unit-variable-number tandem-repeat (MIRU-VNTR) loci, including a discriminatory subset of 15 loci, has recently been defined for the typing of Mycobacterium tuberculosis. Here, we evaluated the performances of this MIRU-VNTR typing system in combination with spoligotyping for the detection of transmission chains in a population-based study comprising 91% of culture-confirmed tuberculosis patients reported in 2003 in Hamburg, Germany...

  20. Identification of a Novel Galactosyl Transferase Involved in Biosynthesis of the Mycobacterial Cell Wall

    OpenAIRE

    Mikušová, Katarína; Belá?ová, Martina; Korduláková, Jana; Honda, Kristine; McNeil, Michael R.; Mahapatra, Sebabrata; Crick, Dean C.; Brennan, Patrick J.

    2006-01-01

    The possibility of the Rv3782 protein of Mycobacterium tuberculosis being a putative galactosyl transferase (GalTr) implicated in galactan synthesis arose from its similarity to the known GalTr Rv3808c, its classification as a nucleotide sugar-requiring inverting glycosyltransferase (GT-2 family), and its location within the “possible arabinogalactan biosynthetic gene cluster” of M. tuberculosis. In order to study the function of the enzyme, active membrane and cell wall fractions from Mycoba...

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

    OpenAIRE

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

    2006-01-01

    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.

  2. CMRegNet-An interspecies reference database for corynebacterial and mycobacterial regulatory networks

    DEFF Research Database (Denmark)

    Abreu, Vinicius A C; Almeida, Sintia

    2015-01-01

    BACKGROUND: Organisms utilize a multitude of mechanisms for responding to changing environmental conditions, maintaining their functional homeostasis and to overcome stress situations. One of the most important mechanisms is transcriptional gene regulation. In-depth study of the transcriptional gene regulatory network can lead to various practical applications, creating a greater understanding of how organisms control their cellular behavior. DESCRIPTION: In this work, we present a new database, CMRegNet for the gene regulatory networks of Corynebacterium glutamicum ATCC 13032 and Mycobacterium tuberculosis H37Rv. We furthermore transferred the known networks of these model organisms to 18 other non-model but phylogenetically close species (target organisms) of the CMNR group. In comparison to other network transfers, for the first time we utilized two model organisms resulting into a more diverse and complete network of the target organisms. CONCLUSION: CMRegNet provides easy access to a total of 3,103 known regulations in C. glutamicum ATCC 13032 and M. tuberculosis H37Rv and to 38,940 evolutionary conserved interactions for 18 non-model species of the CMNR group. This makes CMRegNet to date the most comprehensive database of regulatory interactions of CMNR bacteria. The content of CMRegNet is publicly available online via a web interface found at http://lgcm.icb.ufmg.br/cmregnet .

  3. In Vitro Gene Expression Dissected: Chemostat Surgery for Mycobacterium Tuberculosis

    Directory of Open Access Journals (Sweden)

    Philip D. Marsh

    2002-01-01

    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.

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

    Directory of Open Access Journals (Sweden)

    Afessa Bekele

    2001-09-01

    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.

  5. In vitro responses to a 65-kilodalton mycobacterial protein by synovial T cells from inflammatory arthritis patients.

    Science.gov (United States)

    Gaston, J S; Life, P F; Bailey, L C; Bacon, P A

    1989-10-15

    Bacterial Ag, especially those of mycobacteria, have been implicated in the pathogenesis of experimental inflammatory arthritis in rodents, while in man, reactive arthritis has a clear temporal relationship to infection with particular bacteria. To investigate the role of immune responses to bacterial Ag in inflammatory arthritis, we have examined the proliferative responses of paired synovial fluid and PBMC when stimulated with 1) suspensions of irradiated or heat-killed bacteria associated with reactive arthritis (ReA), 2) purified protein derivative, 3) a recombinant 65-kDa heat shock protein of Mycobacterium leprae. The 65-kDa Ag was stimulatory to synovial fluid mononuclear cells, but not PBMC, from patients with different arthropathies, including most of those with ReA, but also some with rheumatoid arthritis. Furthermore, the magnitude of these responses correlated more closely with responses to ReA-associated bacteria (such as Salmonella), than with responses to the mycobacterial Ag represented in purified protein derivative. These results suggest that the 65-kDa molecule, which is common to a wide range of bacteria, may be an important immunogen for the T cell-mediated immune responses within the joint in different clinically defined inflammatory arthropathies. PMID:2677142

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

    Science.gov (United States)

    Phung, Thu Nguyet; Caruso, Domenico; Godreuil, Sylvain; Keck, Nicolas; Vallaeys, Tatiana; Avarre, Jean-Christophe

    2013-12-01

    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 inexpensive diagnostic test exists, we tested the potential of high-resolution melting analysis (HRMA) to rapidly identify and differentiate several Mycobacterium species involved in fish infections. By analyzing both the melting temperature and melting profile of the 16S-23S rRNA internal transcribed spacer (ITS), we were able to discriminate 12 different species simultaneously. Sensitivity tests conducted on purified M. marinum and M. fortuitum DNA revealed a limit of detection of 10 genome equivalents per reaction. The primers used in this procedure did not lead to any amplification signal with 16 control non-Mycobacterium species, thereby demonstrating their specificity for the genus Mycobacterium. PMID:24123734

  7. Diagnosis of pulmonary and extrapulmonary tuberculosis based on detection of mycobacterial antigen 85B by immuno-PCR.

    Science.gov (United States)

    Singh, Netrapal; Sreenivas, Vishnubhatla; Gupta, Krishna B; Chaudhary, Anil; Mittal, Anshu; Varma-Basil, Mandira; Prasad, Rajendra; Gakhar, Surender K; Khuller, Gopal K; Mehta, Promod K

    2015-12-01

    We developed a novel indirect sandwich immuno-polymerase chain reaction (I-PCR) assay for the detection of mycobacterial antigen 85B (Ag85B, 30kDa, Rv1886c) in pulmonary tuberculosis (PTB) and extrapulmonary tuberculosis (EPTB) patients. The amino-modified reporter DNA was covalently attached with the antidetection antibody through a heterobifunctional cross-linking agent succinimidyl 4-[N-maleimidomethyl]-cyclohexane-1-carboxylate. The detection limit of Ag85B by I-PCR was found to be 1 femtogram (fg)/mL, which was 10(6)-fold lower than an analogous enzyme-linked immunosorbent assay (ELISA). The sensitivities of 85% and 77% with I-PCR and 77.6% and 62.5% with ELISA were observed in smear-positive and smear-negative PTB patients, respectively, with high specificity. On the other hand, sensitivities of 84% and 63.7% with I-PCR and 68% and 47.5% with ELISA were observed in confirmed and clinically suspected EPTB cases, respectively, with high specificity. PMID:26422085

  8. An In Silico Approach for Identification of Potential Anti-Mycobacterial Targets of Vasicine and Related Chemical Compounds.

    Science.gov (United States)

    Chaliha, Amrita Kashyap; Gogoi, Dhrubajyoti; Chetia, Pankaj; Sarma, Diganta; Buragohain, Alak Kumar

    2016-01-01

    Tuberculosis (TB) is known to mankind as one of the most pervasive and persistent of diseases since the early days of civilization. The growing resistance of the causative pathogen Mycobacterium tuberculosis to the standard drug regimen for TB poses further difficulty in its treatment and control. Screening of novel plant-derived compounds with promising anti-tubercular activity has been cited as a prospective route for new anti-tubercular drug discovery and design. Justicia adhatoda L. is a perennial evergreen shrub which is widely mentioned in scientific literature on account of its potent anti-mycobacterial properties. In the present study, we have employed a series of computational methodologies to reveal the probable molecular interactions of vasicine, the principal alkaloid of Justicia adhatoda L., and two of its close natural derivatives- vasicinone and deoxyvasicine, with certain biological targets in M. tuberculosis. Targets were identified from literature and through a reverse Pharmacophore-based approach. Subsequent comparative molecular docking to identify the best ligand-target interactions revealed Antigen 85C of M. tuberculosis as the most potent biological target of vasicine on the basis of optimum molecular docking values. A chemogenomics approach was also employed to validate the molecular interactions between the same class of chemical compounds as vasicine and Antigen 85C. Further, a library of structural analogs of vasicine was created by bioiosterism-based drug design to identify structural analogs with better inhibitory potential against Antigen 85C. PMID:26632438

  9. The chest radiographic appearances of non-tuberculous mycobacterial pulmonary infection in patients with acquired immunodeficiency syndrome

    International Nuclear Information System (INIS)

    Objective: To study the chest radiographic appearances of the non-tuberculous mycobacterial (NTM) pulmonary infection in patients with acquired immune deficiency syndrome (AIDS). Methods: Ten patients with AIDS and NTM underwent chest X-ray radiography and 7 patients performed high-resolution CT (HRCT) scan. Chest radiographic features of' NTM in patients with AIDS were retrospectively analyzed. Results: The chest radiograph showed bilateral pulmonary involvement in 6 cases and single lung involvement in 4 cases (3 cases in the right, 1 case in the left). Patchy air space consolidation (6 cases), large consolidation (5 cases), cavitation (5 cases), small nodules (3 cases), military nodules (2 cases), linear opacity (1 cases) were demonstrated on radiography. On HRCT, air space consolidation (7 cases), small nodules (6 cases), large consolidation (5 cases) with cavitation and cylindric bronchiectasis after the absorption of consolidation, enlarged hilar and mediastinal lymph nodes (4 cases), ground-glass opacities (3 cases), military nodules and 'tree-in-bud' sign (2 case), pleural effusion (1 case), pericardial effusion (1 case) and fibrotic band (1 case) were found. Conclusion: The most common radiographic appearances of NTM in patients with AIDS are bilateral small nodules, large consolidation with cavitation and cylindric bronchiectasis, enlarged hilar and mediastinal lymph nodes. (authors)

  10. The Warburg effect in mycobacterial granulomas is dependent on the recruitment and activation of macrophages by interferon-?.

    Science.gov (United States)

    Appelberg, Rui; Moreira, Diana; Barreira-Silva, Palmira; Borges, Margarida; Silva, Letícia; Dinis-Oliveira, Ricardo Jorge; Resende, Mariana; Correia-Neves, Margarida; Jordan, Michael B; Ferreira, Nuno C; Abrunhosa, Antero J; Silvestre, Ricardo

    2015-08-01

    Granulomas are the hallmark of mycobacterial disease. Here, we demonstrate that both the cell recruitment and the increased glucose consumption in granulomatous infiltrates during Mycobacterium avium infection are highly dependent on interferon-? (IFN-?). Mycobacterium avium-infected mice lacking IFN-? signalling failed to developed significant inflammatory infiltrations and lacked the characteristic uptake of the glucose analogue fluorine-18-fluorodeoxyglucose (FDG). To assess the role of macrophages in glucose uptake we infected mice with a selective impairment of IFN-? signalling in the macrophage lineage (MIIG mice). Although only a partial reduction of the granulomatous areas was observed in infected MIIG mice, the insensitivity of macrophages to IFN-? reduced the accumulation of FDG. In vivo, ex vivo and in vitro assays showed that macrophage activated by IFN-? displayed increased rates of glucose uptake and in vitro studies showed also that they had increased lactate production and increased expression of key glycolytic enzymes. Overall, our results show that the activation of macrophages by IFN-? is responsible for the Warburg effect observed in organs infected with M. avium. PMID:25807843

  11. Mucosal-associated invariant T cells are numerically and functionally deficient in patients with mycobacterial infection and reflect disease activity.

    Science.gov (United States)

    Kwon, Yong-Soo; Cho, Young-Nan; Kim, Moon-Ju; Jin, Hye-Mi; Jung, Hyun-Ju; Kang, Jeong-Hwa; Park, Ki-Jeong; Kim, Tae-Jong; Kee, Hae Jin; Kim, Nacksung; Kee, Seung-Jung; Park, Yong-Wook

    2015-05-01

    Mucosal-associated invariant T (MAIT) cells contribute to protection against certain microorganism infections. The aims of this study were to examine the levels of MAIT cells in pulmonary tuberculosis (TB) and nontuberculous mycobacteria (NTM) lung disease patients, to evaluate the clinical relevance of MAIT cell levels, and to investigate the functions of MAIT cells. Patients with pulmonary TB (n = 35), NTM (n = 29), and healthy controls (n = 75) were enrolled in the study. MAIT cell levels and functions were measured by flow cytometry. Circluating MAIT cell levels were found to be reduced in TB and NTM patients. MAIT cell deficiency reflects a variety of clinical conditions. In particular, MAIT cell numbers were significantly correlated with sputum AFB positivity, extent of disease, hemoglobin levels, lymphocyte counts, CRP and ESR levels. MAIT cells in TB patients failed to produce interferon-? irrespective of the mode of stimulation, whereas NTM patients displayed a defect in MR1-dependent signaling pathway. Notably, an elevated expression of programmed death-1 was also associated with MAIT cell deficiency in TB. This study shows that MAIT cells are numerically and functionally deficient in TB and NTM patients and these deficiencies could contribute to immune system dysreguation in mycobacterial infection. PMID:25837440

  12. Specific interaction between Mycobacterium tuberculosis lipoprotein-derived peptides and target cells inhibits mycobacterial entry in vitro

    Science.gov (United States)

    Ocampo, Marisol; Curtidor, Hernando; Vanegas, Magnolia; Patarroyo, Manuel Alfonso; Patarroyo, Manuel Elkin

    2014-01-01

    Summary Tuberculosis (TB) continues being one of the diseases having the greatest mortality rates around the world, 8.7 million cases having been reported in 2011. An efficient vaccine against TB having a great impact on public health is an urgent need. Usually, selecting antigens for vaccines has been based on proteins having immunogenic properties for patients suffering TB and having had promising results in mice and non-human primates. Our approach has been based on a functional approach involving the pathogen–host interaction in the search for antigens to be included in designing an efficient, minimal, subunit-based anti-tuberculosis vaccine. This means that Mycobacterium tuberculosis has mainly been involved in studies and that lipoproteins represent an important kind of protein on the cell envelope which can also contribute towards this pathogen's virulence. This study has assessed the expression of four lipoproteins from M. tuberculosis H37Rv, i.e. Rv1411c (LprG), Rv1911c (LppC), Rv2270 (LppN) and Rv3763 (LpqH), and the possible biological activity of peptides derived from these. Five peptides were found for these proteins which had high specific binding to both alveolar A549 epithelial cells and U937 monocyte-derived macrophages which were able to significantly inhibit mycobacterial entry to these cells in vitro. PMID:25041568

  13. Phylogenetic detection of horizontal gene transfer during the step-wise genesis of Mycobacterium tuberculosis

    Directory of Open Access Journals (Sweden)

    Turenne Christine

    2009-08-01

    Full Text Available Abstract Background In the past decade, the availability of complete genome sequence data has greatly facilitated comparative genomic research aimed at addressing genetic variability within species. More recently, analysis across species has become feasible, especially in genera where genome sequencing projects of multiple species have been initiated. To understand the genesis of the pathogen Mycobacterium tuberculosis within a genus where the majority of species are harmless environmental organisms, we have used genome sequence data from 16 mycobacteria to look for evidence of horizontal gene transfer (HGT associated with the emergence of pathogenesis. First, using multi-locus sequence analysis (MLSA of 20 housekeeping genes across these species, we derived a phylogeny that serves as the basis for HGT assignments. Next, we performed alignment searches for the 3989 proteins of M. tuberculosis H37Rv against 15 other mycobacterial genomes, generating a matrix of 59835 comparisons, to look for genetic elements that were uniquely found in M. tuberculosis and closely-related pathogenic mycobacteria. To assign when foreign genes were likely acquired, we designed a bioinformatic program called mycoHIT (mycobacterial homologue investigation tool to analyze these data in conjunction with the MLSA-based phylogeny. Results The bioinformatic screen predicted that 137 genes had been acquired by HGT at different phylogenetic strata; these included genes coding for metabolic functions and modification of mycobacterial lipids. For the majority of these genes, corroborating evidence of HGT was obtained, such as presence of phage or plasmid, and an aberrant GC%. Conclusion M. tuberculosis emerged through vertical inheritance along with the step-wise addition of genes acquired via HGT events, a process that may more generally describe the evolution of other pathogens.

  14. Characterization of the MSMEG_2631 Gene (mmp) Encoding a Multidrug and Toxic Compound Extrusion (MATE) Family Protein in Mycobacterium smegmatis and Exploration of Its Polyspecific Nature Using Biolog Phenotype MicroArray

    OpenAIRE

    Mishra, Mukti Nath; Daniels, Lacy

    2013-01-01

    In Mycobacterium, multidrug efflux pumps can be associated with intrinsic drug resistance. Comparison of putative mycobacterial transport genes revealed a single annotated open reading frame (ORF) for a multidrug and toxic compound extrusion (MATE) family efflux pump in all sequenced mycobacteria except Mycobacterium leprae. Since MATE efflux pumps function as multidrug efflux pumps by conferring resistance to structurally diverse antibiotics and DNA-damaging chemicals, we studied this gene (...

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

    Directory of Open Access Journals (Sweden)

    Sarah J. Casey

    2014-03-01

    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.

  16. Expression and participation of eotaxin during mycobacterial (type 1) and schistosomal (type 2) antigen-elicited granuloma formation.

    Science.gov (United States)

    Ruth, J H; Lukacs, N W; Warmington, K S; Polak, T J; Burdick, M; Kunkel, S L; Strieter, R M; Chensue, S W

    1998-10-15

    Eotaxin participation was analyzed during types 1 and 2 lung granuloma formation induced by embolizing Sepharose beads coupled to purified protein derivative (PPD) of Mycobacterium bovis or soluble Ags derived from Schistosoma mansoni eggs. Eotaxin was monitored by protein ELISA and semiquantitative reverse-transcriptase PCR mRNA analysis. Both types 1 and 2 granulomas released eotaxin, but levels were sixfold greater (on day 4) in the type 2 than for the type 1 or foreign body granulomas. Transcripts for eotaxin, IL-4, and CCR3 (eotaxin receptor) were also enhanced during type 2 granuloma formation. Anti-IL-4 treatment impaired eotaxin mRNA in lungs with type 2 granulomas, indicating that IL-4 promoted local eotaxin expression. In vivo, anti-eotaxin treatment caused modest reductions in the size of both types 1 and 2 lesions, with negligible effect on eosinophil recruitment. Surprisingly, anti-eotaxin treatment abrogated IFN-gamma-producing cells in regional lymph nodes during the type 1 PPD response. Lymph nodes draining both types 1 and 2 lesions showed enhanced CCR3 mRNA, but this followed the time of maximum eotaxin protein and mRNA expression. Correlative, in vitro studies revealed that graded doses of eotaxin increased IFN-gamma production from PPD-sensitive regional lymph node cultures, while monocyte-chemotactic protein-1, an important macrophage chemoattractant, had the opposite effect. These findings indicate that eotaxin expression is not limited to type 2 hypersensitivity granulomas, but also promotes IFN-gamma production during mycobacterial responses. PMID:9780203

  17. The DosS-DosT/DosR Mycobacterial Sensor System

    Directory of Open Access Journals (Sweden)

    Santhosh Sivaramakrishnan

    2013-07-01

    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.

  18. Molecular Identification and Conventional Susceptibility Testing of Iranian Clinical Mycobacterium fortuitum Isolates

    Directory of Open Access Journals (Sweden)

    Parvin Heidarieh

    2010-01-01

    Full Text Available Rapidly growing mycobacteria (RGM are capable of producing diseases in humans. Since mycobacteria vary in their susceptibility, precise identification is critical for adoption of correct drug therapy. The main aim of this study was molecular identification and evaluation of antimicrobial susceptibility pattern of Iranian clinically isolated Myocbacterium fortuitum.Materials and MethodsA total of 72 presumptively identified isolates of clinical atypical mycobacteria collected by Isfahan Research Center for Infectious Diseases & Tropical Medicine during 2006-2008 were included in the current study. A combination of conventional and molecular tests was applied to identify the isolates. Molecular methods including genus and group specific PCR and PCR-Restriction Algorithm (PRA based on hsp65 gene were applied to achieve exact identification of mycobacterial strains. Antimicrobial susceptibility testing on M. fortuitum isolates was performed by in-house prepared broth microdilution test..ResultsOut of 72 collected atypical mycobacteria isolates, we identified 25 strains of M. fortuitum. All strains had the specific molecular markers of mycobacterial identity and similar species specific PRA pattern of the international type strain of M. fortuitum. Drug susceptibility testing showed that the M. fortuitum isolates are sensitive to amikacin, sulfamethoxazole and ciprofloxacin (100%, imipenem (92%, clarithromycin (76%, cefoxitin (56% and doxycycline (16%.ConclusionMolecular identification of atypical mycobacteria based on PRA is a reliable and rapid approach which can identify mycobacterial strains to the species level. Our study showed that M. fortuitum plays a significant role in pulmonary and extrapulmonary infection in patients and should be given proper considerations when clinical samples are processed.

  19. A 475 years-old founder effect involving IL12RB1: a highly prevalent mutation conferring Mendelian Susceptibility to Mycobacterial Diseases in European descendants

    OpenAIRE

    Yancoski, J; Rocco, C; Bernasconi, A.; Oleastro, M; Bezrodnik, L; Vrátnica, C; Haerynck, F; Rosenzweig, SD

    2009-01-01

    Mutations in IFNGR1, IFNGR2, IL12RB1, IL12B, STAT1 and NEMO result in a common clinical phenotype known as Mendelian Susceptibility to Mycobacterial Diseases (MSMD). Interleukin-12 receptor ?1 (IL-12R?1) deficiency is the most common genetic etiology for MSMD. Known mutations affecting IL12RB1 are recessively inherited and are associated with null response to both IL-12 and IL-23. Mutation IL12RB1 1623_1624delinsTT was originally described in 5 families from European origin (2 from Germany; 1...

  20. Síndromes micobacterianos felinos y su importancia en la salud pública - Feline mycobacterial syndromes and its importance to public health

    Directory of Open Access Journals (Sweden)

    Jorge, María Cristina

    2012-01-01

    Full Text Available ResumenEn felinos domésticos las especies del género Mycobacterium causan tres síndromes: tuberculosis ocasionada por el complejo M. tuberculosis, lepra felina por M. lepraemurium y micobacteriosis atípica causada por varias especies de micobacterias no tuberculosas y no lepromatosas.AbstractIn domestic felines three mycobacterial syndromes are described: tuberculoses caused by M. tuberculosis complex, feline leprosy caused by M. lepraemurium and atypical mycobacteriosis caused by non tuberculous and non lepromatous mycobacteria.

  1. Cytotoxic human HLA class II restricted purified protein derivative-reactive T-lymphocyte clones. IV. Analysis of HLA restriction pattern and mycobacterial antigen specificity

    DEFF Research Database (Denmark)

    Hansen, P W; Petersen, C M; Povlsen, J V; Kristensen, T

    1987-01-01

    against autologous PPD-pulsed monocyte targets, was examined against a panel of allogeneic PPD pulsed targets. In agreement with our findings with bulk-expanded PPD-reactive cytotoxic T lymphocytes, all clones were restricted by HLA class II antigens: seven by HLA-DR 2 and one by HLA-DRw10--the other HLA...... monoclonal antibodies against monomorphic HLA-DR determinants. The antigen specificity of the clones was studied by using autologous monocyte targets pulsed with antigens prepared from a range of different mycobacterial species. All seven HLA-DR2-restricted clones reacted with the majority of antigens tested...

  2. Purification, crystallization and preliminary X-ray analysis of SGR6054, a Streptomyces homologue of the mycobacterial integration host factor mIHF

    International Nuclear Information System (INIS)

    A Streptomyces homologue of the mycobacterial integration host factor mIHF was heterologously produced, purified and crystallized in the presence of a 16-mer duplex DNA by the sitting-drop vapour-diffusion method. The best crystal diffracted X-rays to 2.22 Å resolution and belonged to space group C2. The mycobacterial integration host factor (mIHF) is a small nonspecific DNA-binding protein that is essential for the growth of Mycobacterium smegmatis. mIHF homologues are widely distributed among Actinobacteria, and a Streptomyces homologue of mIHF is involved in control of sporulation and antibiotic production in S. coelicolor A3(2). Despite their important biological functions, a structure of mIHF or its homologues has not been elucidated to date. Here, the S. griseus mIHF homologue (SGR6054) was expressed and purified from Escherichia coli and crystallized in the presence of a 16-mer duplex DNA by the sitting-drop vapour-diffusion method. The plate-shaped crystal belonged to space group C2, with unit-cell parameters a = 88.53, b = 69.35, c = 77.71 Å, ? = 96.63°, and diffracted X-rays to 2.22 Å resolution

  3. 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.)

  4. Direct Comparison of Xpert MTB/RIF Assay with Liquid and Solid Mycobacterial Culture for Quantification of Early Bactericidal Activity

    Science.gov (United States)

    Kayigire, Xavier A.; Friedrich, Sven O.; Venter, Amour; Dawson, Rodney; Gillespie, Stephen H.; Boeree, Martin J.; Heinrich, Norbert; Hoelscher, Michael

    2013-01-01

    The early bactericidal activity of antituberculosis agents is usually determined by measuring the reduction of the sputum mycobacterial load over time on solid agar medium or in liquid culture. This study investigated the value of a quantitative PCR assay for early bactericidal activity determination. Groups of 15 patients were treated with 6 different antituberculosis agents or regimens. Patients collected sputum for 16 h overnight at baseline and at days 7 and 14 after treatment initiation. We determined the sputum bacterial load by CFU counting (log CFU/ml sputum, reported as mean ± standard deviation [SD]), time to culture positivity (TTP, in hours [mean ± SD]) in liquid culture, and Xpert MTB/RIF cycle thresholds (CT, n [mean ± SD]). The ability to discriminate treatment effects between groups was analyzed with one-way analysis of variance (ANOVA). All measurements showed a decrease in bacterial load from mean baseline (log CFU, 5.72 ± 1.00; TTP, 116.0 ± 47.6; CT, 19.3 ± 3.88) to day 7 (log CFU, ?0.26 ± 1.23, P = 0.2112; TTP, 35.5 ± 59.3, P = 0.0002; CT, 0.55 ± 3.07, P = 0.6030) and day 14 (log CFU, ?0.55 ± 1.24, P = 0.0006; TTP, 54.8 ± 86.8, P < 0.0001; CT, 2.06 ± 4.37, P = 0.0020). The best discrimination between group effects was found with TTP at day 7 and day 14 (F = 9.012, P < 0.0001, and F = 11.580, P < 0.0001), followed by log CFU (F = 4.135, P = 0.0024, and F = 7.277, P < 0.0001). CT was not significantly discriminative (F = 1.995, P = 0.091, and F = 1.203, P = 0.316, respectively). Culture-based methods are superior to PCR for the quantification of early antituberculosis treatment effects in sputum. PMID:23596237

  5. 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 ...

  6. The Mycobacterium tuberculosis relBE toxin:antitoxin genes are stress-responsive modules that regulate growth through translation inhibition.

    Science.gov (United States)

    Korch, Shaleen B; Malhotra, Vandana; Contreras, Heidi; Clark-Curtiss, Josephine E

    2015-11-01

    Toxin-antitoxin (TA) genes are ubiquitous among bacteria and are associated with persistence and dormancy. Following exposure to unfavorable environmental stimuli, several species (Escherichia coli, Staphylococcus aureus, Myxococcus xanthus) employ toxin proteins such as RelE and MazF to downregulate growth or initiate cell death. Mycobacterium tuberculosis possesses three Rel TA modules (Rel Mtb ): RelBE Mtb , RelFG Mtb and RelJK Mtb (Rv1246c-Rv1247c, Rv2865-Rv2866, and Rv3357-Rv3358, respectively), which inhibit mycobacterial growth when the toxin gene (relE, relG, relK) is expressed independently of the antitoxin gene (relB, relF, relJ). In the present study, we examined the in vivo mechanism of the RelE Mtb toxin protein, the impact of RelE Mtb on M. tuberculosis physiology and the environmental conditions that regulate all three rel Mtb modules. RelE Mtb negatively impacts growth and the structural integrity of the mycobacterial envelope, generating cells with aberrant forms that are prone to extensive aggregation. At a time coincident with growth defects, RelE Mtb mediates mRNA degradation in vivo resulting in significant changes to the proteome. We establish that rel Mtb modules are stress responsive, as all three operons are transcriptionally activated following mycobacterial exposure to oxidative stress or nitrogen-limiting growth environments. Here we present evidence that the rel Mtb toxin:antitoxin family is stress-responsive and, through the degradation of mRNA, the RelE Mtb toxin influences the growth, proteome and morphology of mycobacterial cells. PMID:26502963

  7. Factors associated with pastoral community knowledge and occurrence of mycobacterial infections in Human-Animal Interface areas of Nakasongola and Mubende districts, Uganda

    Directory of Open Access Journals (Sweden)

    Biffa Demelash

    2010-08-01

    Full Text Available Abstract Background Nontuberculous mycobacteria (NTM are emerging opportunistic pathogens whose role in human and animal disease is increasingly being recognized. Major concerns are their role as opportunistic pathogens in HIV/AIDS infections. The role of open natural water sources as source and livestock/wildlife as reservoirs of infections to man are well documented. This presents a health challenge to the pastoral systems in Africa that rely mostly on open natural water sources to meet livestock and human needs. Recent study in the pastoral areas of Uganda showed infections with same genotypes of NTM in pastoralists and their livestock. The aim of this study was to determine the environmental, animal husbandry and socio-demographic factors associated with occurrence and the pastoral community knowledge of mycobacterial infections at the human-environment-livestock/wildlife interface (HELI areas in pastoral ecosystems of Uganda. Methods Two hundred and fifty three (253 individuals were subjected to a questionnaire survey across the study districts of Nakasongola and Mubende. Data were analyzed using descriptive statistics and multivariable logistic regression analysis. Results Humans sharing of the water sources with wild animals from the forest compared to savannah ecosystem (OR = 3.3, the tribe of herding pastoral community (OR = 7.9, number of rooms present in household (3-5 vs. 1-2 rooms (OR = 3.3 were the socio-demographic factors that influenced the level of knowledge on mycobacterial infections among the pastoral communities. Tribe (OR = 6.4, use of spring vs. stream water for domestic use (OR = 4.5, presence of sediments in household water receptacle (OR = 2.32, non separation of water containers for drinking and domestic use (OR = 2.46, sharing of drinking water sources with wild animals (OR = 2.1, duration of involvement of >5 yrs in cattle keeping (OR = 3.7 and distance of household to animal night shelters (>20 meters (OR = 3.8 were significant socio-demographic factors associated with the risk of occurrence of mycobacterioses among the pastoral communities in Uganda. Conclusion The socio-demographic, environmental and household related factors influence the risk of occurrence as well as pastoralists' knowledge of mycobacterial infections in the pastoral households at the human-environment-livestock/wildlife pastoral interface areas of Uganda.

  8. Micobacterias atípicas en cinco pacientes adultos sin evidencias de inmunosupresión: Construyendo una experiencia / Atypical mycobacterial infections in five adult patients without evidence of immunosuppression: Making an experience

    Scientific Electronic Library Online (English)

    Alberto, Fica; Andrés, Soto; Jeannette, Dabanch; Lorena, Porte; Marcelo, Castro; Luis, Thompson; M. Elvira, Balcells.

    2015-02-01

    Full Text Available El objetivo de este trabajo es reportar la experiencia acumulada sobre infecciones por micobacterias atípicas en pacientes sin inmunosupresión. Entre el año 2008 y 2013 se observaron cinco pacientes con infección por micobacterias atípicas: dos con infección cutánea y tres con infección pulmonar. Ni [...] nguno de estos pacientes tenía evidencias de inmunosupresión. Un paciente con bursitis de codo por M. chelonae tuvo un estudio citoquímico con aumento de celularidad de predominio mononuclear y desarrollo de bacterias al quinto día; respondió favorablemente a claritromicina. Un caso con infección cutánea por M. fortuitum evolucionó en forma prolongada con supuración ganglionar antes del diagnóstico y el cultivo solicitado a los 13 días fue positivo. Los tres pacientes con aislados pulmonares presentaron tos y expectoración y tenían en común ser mujeres en edad post-menopáusica y presentar pequeños infiltrados nodulares asociados a bronquiectasias en el estudio de imágenes pulmonares, un patrón descrito en la literatura científica. En estos tres casos, la latencia entre la toma de muestra y el informe definitivo tuvo un rango de 40 a 89 días. El aislamiento de micobacterias atípicas en muestras de expectoración en pacientes sin inmunosupresión se da en un contexto típico pero plantea dificultades diagnósticas y terapéuticas. El lento crecimiento de estos microorganismos en el laboratorio contribuye a este problema. Abstract in english We aim to communicate the experience gathered during the management of infections by atypical mycobacteria in immunocompetent patients in a general practice. Between 2008 and 2013, 5 patients with non-tuberculous mycobacterial infections were identified: 2 with cutaneous involvement and 3 with lung [...] infection. None of them had evidence of immunosuppression. A patient with elbow bursitis by M. chelonae presented with a high mononuclear count in fluid analysis with mycobacterial growth at the fifth day of culture. He evolved satisfactorily with clarithromycin. A case with M. fortuitum skin infection had a delayed initial diagnosis with progression to local draining lymph nodes; the culture when requested was positive after 13 days of incubation. Patients with pulmonary infection presented with prolonged cough and sputum and had in common to be postmenopausal women displaying small nodules and bronchiectases at lung images, a classical pattern. Time elapsed between respiratory sampling and a definitive inform ranged from 40 to 89 days. Non-tuberculous mycobacterial infections in non-immunosuppresed patients can generate diagnostic and therapeutic challenges. Delay in identification contributes to this problem.

  9. Nontuberculous mycobacterial pulmonary infections.

    Science.gov (United States)

    Johnson, Margaret M; Odell, John A

    2014-03-01

    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

  10. 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)

    Berod, Luciana; Stüve, Philipp; Varela, Filipa; Behrends, Jochen; Swallow, Maxine; Kruse, Friederike; Krull, Freyja; Ghorbani, Peyman; Mayer, Christian T; Hölscher, Christoph; Sparwasser, Tim

    2014-01-01

    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

  11. Diagnostic Yield of Bronchoalveolar Lavage Gene Xpert in Smear-Negative and Sputum-Scarce Pulmonary Tuberculosis

    International Nuclear Information System (INIS)

    Objective: To measure the diagnostic yield of Bronchoalveolar Lavage (BAL) gene Xpert (Xpert MTB/RIF assay), to detect Mycobacterium tuberculosis (MTB) and rifampicin resistance and compare it with that of mycobacterial cultures in a suspected case of pulmonary tuberculosis. Study Design: An analytical study. Place and Duration of Study: Department of Pulmonology, Fauji Foundation Hospital (FFH), Rawalpindi, from December 2012 to August 2013. Methodology: BAL specimens of 93 patients with suspected pulmonary tuberculosis with smear-negative or sputumscarce disease, who presented to the Department of Pulmonology, FFH, Rawalpindi were inducted. A smear-negative case was one in whom three consecutive early morning sputum samples did not reveal acid fast bacilli when examined by microscopy with Zeihl Nelson (ZN) stain. Patients who had sputum amount less than 1 ml were defined to have sputumscarce disease. The same was evaluated with ZN stain, gene Xpert and mycobacterial cultures. Sensitivity analysis was carried out using culture as the gold standard. Results: The frequency of positive mycobacterial cultures was 85 (91.4%). The sensitivity, specificity, positive predictive value and negative predictive values of BAL gene Xpert to detect Mycobacterium tuberculosis were 91.86%, 71.42%, 97.53% and 41.66% respectively. Xpert MTB/RIF assay had a sensitivity and specificity of 83.33% and 100% to detect rifampicin resistance. Conclusion: Bronchoalveolar lavage gene Xpert had a superior diagnostic yield in patients with either smear-negative or sputum-scarce pulmonary tuberculosis. Hence a positive Xpert MTB/RIF assay may be a useful adjunct to diagnosis and detection of MDR-TB in bronchoalveolar lavage specimens. (author)

  12. Characterization of the Mycobacterial Acyl-CoA Carboxylase Holo Complexes Reveals Their Functional Expansion into Amino Acid Catabolism

    OpenAIRE

    Ehebauer, Matthias T.; Zimmermann, Michael; Jakobi, Arjen J.; Noens, Elke E.; Laubitz, Daniel; Cichocki, Bogdan; Marrakchi, Hedia; Lanéelle, Marie-Antoinette; Daffé, Mamadou; Sachse, Carsten; Dziembowski, Andrzej; Sauer, Uwe; Wilmanns, Matthias

    2015-01-01

    Biotin-mediated carboxylation of short-chain fatty acid coenzyme A esters is a key step in lipid biosynthesis that is carried out by multienzyme complexes to extend fatty acids by one methylene group. Pathogenic mycobacteria have an unusually high redundancy of carboxyltransferase genes and biotin carboxylase genes, creating multiple combinations of protein/protein complexes of unknown overall composition and functional readout. By combining pull-down assays with mass spectrome...

  13. Mycobacteria mobility shift assay: a method for the rapid identification of Mycobacterium tuberculosis and nontuberculous mycobacteria

    Directory of Open Access Journals (Sweden)

    Letícia Muraro Wildner

    2014-06-01

    Full Text Available The identification of mycobacteria is essential because tuberculosis (TB and mycobacteriosis are clinically indistinguishable and require different therapeutic regimens. The traditional phenotypic method is time consuming and may last up to 60 days. Indeed, rapid, affordable, specific and easy-to-perform identification methods are needed. We have previously described a polymerase chain reaction-based method called a mycobacteria mobility shift assay (MMSA that was designed for Mycobacterium tuberculosis complex (MTC and nontuberculous mycobacteria (NTM species identification. The aim of this study was to assess the MMSA for the identification of MTC and NTM clinical isolates and to compare its performance with that of the PRA-hsp65 method. A total of 204 clinical isolates (102 NTM and 102 MTC were identified by the MMSA and PRA-hsp65. For isolates for which these methods gave discordant results, definitive species identification was obtained by sequencing fragments of the 16S rRNA and hsp65 genes. Both methods correctly identified all MTC isolates. Among the NTM isolates, the MMSA alone assigned 94 (92.2% to a complex or species, whereas the PRA-hsp65 method assigned 100% to a species. A 91.5% agreement was observed for the 94 NTM isolates identified by both methods. The MMSA provided correct identification for 96.8% of the NTM isolates compared with 94.7% for PRA-hsp65. The MMSA is a suitable auxiliary method for routine use for the rapid identification of mycobacteria.

  14. Evaluation of Mycobacterial Interspersed Repetitive-Unit-Variable-Number Tandem-Repeat Analysis and Spoligotyping for Genotyping of Mycobacterium bovis Isolates and a Comparison with Restriction Fragment Length Polymorphism Typing ?

    OpenAIRE

    McLernon, Joanne; Costello, Eamon; Flynn, Orla; Madigan, Gillian; Ryan, Fergus (Fergus W.)

    2010-01-01

    Common strain typing methods for differentiation of Mycobacterium bovis isolates include restriction endonuclease analysis (REA), restriction fragment length polymorphism (RFLP) analysis, spoligotyping, and, more recently, mycobacterial interspersed repetitive-unit-variable-number tandem-repeat (MIRU-VNTR) typing. MIRU-VNTR typing and spoligotyping were evaluated in this study, and these typing methods were compared with RFLP typing. A total of 386 M. bovis isolates from cattle, badgers, and ...

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

    Directory of Open Access Journals (Sweden)

    Farivar Taghi

    2008-07-01

    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.

  16. Rapid mycobacterial liquid culture-screening method for Mycobacterium avium complex based on secreted antigen-capture enzyme-linked immunosorbent assay.

    Science.gov (United States)

    Shin, Sung Jae; Anklam, Kelly; Manning, Elizabeth J B; Collins, Michael T

    2009-05-01

    Sensors in automated liquid culture systems for mycobacteria, such as MGIT, BacT/Alert 3D, and Trek ESP II, flag growth of any type of bacteria; a positive signal does not mean that the target mycobacteria are present. All signal-positive cultures thus require additional and often laborious testing. An immunoassay was developed to screen liquid mycobacterial cultures for evidence of Mycobacterium avium complex (MAC). The method, called the MAC-enzyme-linked immunosorbent assay (ELISA), relies on detection of MAC-specific secreted antigens in liquid culture. Secreted MAC antigens were captured by the MAC-ELISA with polyclonal anti- Mycobacterium avium subsp. paratuberculosis chicken immunoglobulin Y (IgY), detected using rabbit anti-MAC IgG, and then revealed using horseradish peroxidase-conjugated goat anti-rabbit IgG. When the MAC-ELISA was evaluated using pure cultures of known mycobacterial (n = 75) and nonmycobacterial (n = 17) organisms, no false-positive or false-negative MAC-ELISA results were found. By receiver operator characteristic (ROC) analysis of 1,275 previously identified clinical isolates, at the assay optimal cutoff the diagnostic sensitivity and specificity of the MAC-ELISA were 92.6% (95% confidence interval [95% CI], 90.3 to 94.5) and 99.9% (95% CI, 99.2 to 100), respectively, with an area under the ROC curve of 0.992. Prospective evaluation of the MAC-ELISA with an additional 652 clinical samples inoculated into MGIT ParaTB medium and signaling positive per the manufacturer's instructions found that the MAC-ELISA was effective in determining those cultures that actually contained MAC species and warranting the resources required to identify the organism by PCR. Of these 652 MGIT-positive cultures, the MAC-ELISA correctly identified 96.8% (of 219 MAC-ELISA-positive cultures) as truly containing MAC mycobacteria, based on PCR or high-performance liquid chromatography (HPLC) as reference tests. Only 6 of 433 MGIT signal-positive cultures (1.4%) were MAC-ELISA false negative, and only 7 of 219 MGIT signal-negative cultures (3.2%) were false positive. The MAC-ELISA is a low-cost, rapid, sensitive, and specific test for MAC in liquid cultures. It could be used in conjunction with or independent of automated culture reading instrumentation. For maximal accuracy and subspecies-specific identification, use of a confirmatory multiplex MAC PCR is recommended. PMID:19261776

  17. Gene Transfer in Mycobacterium tuberculosis: Shuttle Phasmids to Enlightenment

    Science.gov (United States)

    JACOBS, WILLIAM R.

    2016-01-01

    Infectious diseases have plagued humankind throughout history and have posed serious public health problems. Yet vaccines have eradicated smallpox and antibiotics have drastically decreased the mortality rate of many infectious agents. These remarkable successes in the control of infections came from knowing the causative agents of the diseases, followed by serendipitous discoveries of attenuated viruses and antibiotics. The discovery of DNA as genetic material and the understanding of how this information translates into specific phenotypes have changed the paradigm for developing new vaccines, drugs, and diagnostic tests. Knowledge of the mechanisms of immunity and mechanisms of action of drugs has led to new vaccines and new antimicrobial agents. The key to the acquisition of the knowledge of these mechanisms has been identifying the elemental causes (i.e., genes and their products) that mediate immunity and drug resistance. The identification of these genes is made possible by being able to transfer the genes or mutated forms of the genes into causative agents or surrogate hosts. Such an approach was limited in Mycobacterium tuberculosis by the difficulty of transferring genes or alleles into M. tuberculosis or a suitable surrogate mycobacterial host. The construction of shuttle phasmids—chimeric molecules that replicate in Escherichia coli as plasmids and in mycobacteria as mycobacteriophages—was instrumental in developing gene transfer systems for M. tuberculosis. This review will discuss M. tuberculosis genetic systems and their impact on tuberculosis research. “I had to know my enemy in order to prevail against him.”Nelson Mandela PMID:26105819

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

    OpenAIRE

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

    2009-01-01

    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...

  19. Identification of non-tuberculous mycobacteria from the Central Public Health Laboratory from Mato Grosso do Sul and analysis of clinical relevance

    OpenAIRE

    de Souza Moraes, Paulo Ricardo; Chimara, Erica; Telles, Maria Alice da Silva; Ueki, Suely Yoko Misuka; Cunha, Eunice Atsuko Totumi; Honer, Michael Robin; Sylvia Cardoso LEÃO

    2008-01-01

    Non-tuberculous mycobacteria isolated at the Central Public Health Laboratory from Mato Grosso do Sul in 2003 and 2004 were identified by conventional phenotypic methods (TI) and by PCR-Restriction Enzyme Analysis (PRA) using the hsp65 gene as target (PRA-hsp65). With 15 of the 32 analysed isolates, results of both methods were concordant, being 8 Mycobacterium avium, 3 M. fortutium, 1 M. kansasii, 1 M. flavescens, 1 M. peregrinum and 1 Nocardia brasiliensis. TI of 12 isolates was inconclusiv...

  20. Identification of non-tuberculous mycobacteria from the Central Public Health Laboratory from Mato Grosso do Sul and analysis of clinical relevance Identificação de micobactérias não-tuberculosas do Laboratório Central de Saúde Pública de Mato Grosso de Sul e análise de dados clínicos dos pacientes

    OpenAIRE

    Paulo Ricardo de Souza Moraes; Erica Chimara; Maria Alice da Silva Telles; Suely Yoko Misuka Ueki; Eunice Atsuko Totumi Cunha; Michael Robin Honer; Sylvia Cardoso Leão

    2008-01-01

    Non-tuberculous mycobacteria isolated at the Central Public Health Laboratory from Mato Grosso do Sul in 2003 and 2004 were identified by conventional phenotypic methods (TI) and by PCR-Restriction Enzyme Analysis (PRA) using the hsp65 gene as target (PRA-hsp65). With 15 of the 32 analysed isolates, results of both methods were concordant, being 8 Mycobacterium avium, 3 M. fortutium, 1 M. kansasii, 1 M. flavescens, 1 M. peregrinum and 1 Nocardia brasiliensis. TI of 12 isolates was inconclusiv...

  1. Identification of non-tuberculous mycobacteria from the Central Public Health Laboratory from Mato Grosso do Sul and analysis of clinical relevance Identificação de micobactérias não-tuberculosas do Laboratório Central de Saúde Pública de Mato Grosso de Sul e análise de dados clínicos dos pacientes

    Directory of Open Access Journals (Sweden)

    Paulo Ricardo de Souza Moraes

    2008-06-01

    Full Text Available Non-tuberculous mycobacteria isolated at the Central Public Health Laboratory from Mato Grosso do Sul in 2003 and 2004 were identified by conventional phenotypic methods (TI and by PCR-Restriction Enzyme Analysis (PRA using the hsp65 gene as target (PRA-hsp65. With 15 of the 32 analysed isolates, results of both methods were concordant, being 8 Mycobacterium avium, 3 M. fortutium, 1 M. kansasii, 1 M. flavescens, 1 M. peregrinum and 1 Nocardia brasiliensis. TI of 12 isolates was inconclusive. Novel PRA-hsp65 patterns were observed with 11 isolates. Medical data were evaluated for inference of clinical relevance of these isolates.Micobactérias não-tuberculosas isoladas no Laboratório Central de Saúde Pública de Mato Grosso do Sul em 2003 e 2004 foram identificadas usando métodos fenotípicos convencionais (TI e PCR-Restriction Enzyme Analysis (PRA tendo o gene hsp65 como alvo (PRA-hsp65. Em 15 dos 32 isolados analisados os resultados obtidos com ambos métodos foram concordantes, sendo 8 Mycobacterium avium, 3 M. fortutium, 1 M. kansasii, 1 M. flavescens, 1 M. peregrinum e 1 Nocardia brasiliensis. TI de 12 isolados não foi conclusiva. Perfis não descritos de PRA-hsp65 foram observados com 11 isolados. Dados dos prontuários médicos foram avaliados para inferir a relevância clínica dos isolados.

  2. The mycobacterial antibiotic resistance determinant WhiB7 acts as a transcriptional activator by binding the primary sigma factor SigA (RpoV).

    Science.gov (United States)

    Burian, Ján; Yim, Grace; Hsing, Michael; Axerio-Cilies, Peter; Cherkasov, Artem; Spiegelman, George B; Thompson, Charles J

    2013-12-01

    Tuberculosis therapeutic options are limited by the high intrinsic antibiotic resistance of Mycobacterium tuberculosis. The putative transcriptional regulator WhiB7 is crucial for the activation of systems that provide resistance to diverse antibiotic classes. Here, we used in vitro run-off, two-hybrid assays, as well as mutagenic, complementation and protein pull-down experiments, to characterize WhiB7 as an auto-regulatory, redox-sensitive transcriptional activator in Mycobacterium smegmatis. We provide the first direct biochemical proof that a WhiB protein promotes transcription and also demonstrate that this activity is sensitive to oxidation (diamide). Its partner protein for transcriptional activation was identified as SigA, the primary sigma factor subunit of RNA polymerase. Residues required for the interaction mapped to region 4 of SigA (including R515H) or adjacent domains of WhiB7 (including E63D). WhiB7's ability to provide a specific spectrum of antibiotic-resistance was dependent on these residues as well as its C-terminal AT-hook module that binds to an AT-rich motif immediately upstream of the -35 hexamer recognized by SigA. These experimentally established constrains, combined with protein structure predictions, were used to generate a working model of the WhiB7-SigA-promoter complex. Inhibitors preventing WhiB7 interactions could allow the use of previously ineffective antibiotics for treatment of mycobacterial diseases. PMID:23990327

  3. Incidence of active mycobacterial infections in Brazilian patients with chronic inflammatory arthritis and negative evaluation for latent tuberculosis infection at baseline - A longitudinal analysis after using TNF? blockers

    Science.gov (United States)

    Gomes, Carina Mori Frade; Terreri, Maria Teresa; de Moraes-Pinto, Maria Isabel; Barbosa, Cássia; Machado, Natália Pereira; Melo, Maria Roberta; Pinheiro, Marcelo Medeiros

    2015-01-01

    Several studies point to the increased risk of reactivation of latent tuberculosis infection (LTBI) in patients with chronic inflammatory arthritis (CIAs) after using tumour necrosis factor (TNF)? blockers. To study the incidence of active mycobacterial infections (aMI) in patients starting TNF ? blockers, 262 patients were included in this study: 109 with rheumatoid arthritis (RA), 93 with ankylosing spondylitis (AS), 44 with juvenile idiopathic arthritis (JIA) and 16 with psoriatic arthritis (PsA). All patients had indication for anti-TNF ? therapy. Epidemiologic and clinical data were evaluated and a simple X-ray and tuberculin skin test (TST) were performed. The control group included 215 healthy individuals. The follow-up was 48 months to identify cases of aMI. TST positivity was higher in patients with AS (37.6%) than in RA (12.8%), PsA (18.8%) and JIA (6.8%) (p aMI. The overall incidence rate of aMI was 86.93/100,000 person-years [95% confidence interval (CI) 23.6-217.9] for patients and 35.79/100,000 person-years (95% CI 12.4-69.6) for control group (p aMI had no evidence of LTBI at the baseline evaluation. Patients with CIA starting TNF ? blockers and no evidence of LTBI at baseline, particularly with nonreactive TST, may have higher risk of aMI. PMID:26560983

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

    Directory of Open Access Journals (Sweden)

    Dagur Pradeep

    2012-09-01

    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.

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

    Directory of Open Access Journals (Sweden)

    Mandira Varma-Basil

    2011-08-01

    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.

  6. 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)

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

    2011-08-01

    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 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.

  7. Serologic follow-up of IgG responses against recombinant mycobacterial proteins ML0405, ML2331 and LID-1 in a leprosy hyperendemic area in Venezuela

    Directory of Open Access Journals (Sweden)

    Elsa Rada

    2012-12-01

    Full Text Available Leprosy is a slowly evolving disease that occurs mainly in adults. In this study, the Mamaría Village, state of Portuguesa was selected because it had one of the highest prevalence rates (13.25% of leprosy cases in 1997. Between 1998-2004, 20.2% of the 89 cases registered in this village were less than 15 years old and 61.8% were males. Pau-cibacillary (PB lesions were the predominant clinical forms identified, although also multibacillary (MB forms were found. Additionally, 76% of the patients were bacteriologically negative. At the time of diagnosis, 75% of the patients presented with grade 0 disabilities, 23% with grade 1 and 2% with grade 2. Serum samples were collected from 18 PB and 15 MB patients, in addition to 14 family contacts, at the beginning and end of treatment. All the groups were re-evaluated during a three-year period (2008-2011. The proteins used for evaluation were ML0405, ML2331 and LID-1. These mycobacterial proteins were highly specific for Mycobacterium leprae and the IgG responses decreased in both MB and PB patients during multidrug treatment. Our results suggest that these antigens could be used as markers for successful treatment of non-reactional lepromatous patients.

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

    Energy Technology Data Exchange (ETDEWEB)

    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

    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.

  9. 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 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

  10. The twin-arginine translocation pathway of Mycobacterium smegmatis is functional and required for the export of mycobacterial beta-lactamases.

    Science.gov (United States)

    McDonough, Justin A; Hacker, Kari E; Flores, Anthony R; Pavelka, Martin S; Braunstein, Miriam

    2005-11-01

    The twin-arginine translocation (Tat) pathway exports folded proteins across the bacterial cytoplasmic membrane and is responsible for the proper extracytoplasmic localization of proteins involved in a variety of cellular functions, including pathogenesis. The Mycobacterium tuberculosis and Mycobacterium smegmatis genomes contain open reading frames with homology to components of the Tat export system (TatABC) as well as potential Tat-exported proteins possessing N-terminal signal sequences with the characteristic twin-arginine motif. Due to the importance of exported virulence factors in the pathogenesis of M. tuberculosis and the limited understanding of mycobacterial protein export systems, we sought to determine the functional nature of the Tat export pathway in mycobacteria. Here we describe phenotypic analyses of DeltatatA and DeltatatC deletion mutants of M. smegmatis, which demonstrated that tatA and tatC encode components of a functional Tat system capable of exporting characteristic Tat substrates. Both mutants displayed a growth defect on agar medium and hypersensitivity to sodium dodecyl sulfate. The mutants were also defective in the export of active beta-lactamases of M. smegmatis (BlaS) and M. tuberculosis (BlaC), both of which possess twin-arginine signal sequences. The Tat-dependent nature of BlaC was further revealed by mutation of the twin-arginine motif. Finally, we demonstrated that replacement of the native signal sequence of BlaC with the predicted Tat signal sequences of M. tuberculosis phospholipase C proteins (PlcA and PlcB) resulted in the Tat-dependent export of an enzymatically active 'BlaC. Thus, 'BlaC can be used as a genetic reporter for Tat-dependent export in mycobacteria. PMID:16267291

  11. Incidence of active mycobacterial infections in Brazilian patients with chronic inflammatory arthritis and negative evaluation for latent tuberculosis infection at baseline - A longitudinal analysis after using TNFa blockers.

    Science.gov (United States)

    Gomes, Carina Mori Frade; Terreri, Maria Teresa; Moraes-Pinto, Maria Isabel de; Barbosa, Cássia; Machado, Natália Pereira; Melo, Maria Roberta; Pinheiro, Marcelo Medeiros

    2015-11-01

    Several studies point to the increased risk of reactivation of latent tuberculosis infection (LTBI) in patients with chronic inflammatory arthritis (CIAs) after using tumour necrosis factor (TNF)a blockers. To study the incidence of active mycobacterial infections (aMI) in patients starting TNFa blockers, 262 patients were included in this study: 109 with rheumatoid arthritis (RA), 93 with ankylosing spondylitis (AS), 44 with juvenile idiopathic arthritis (JIA) and 16 with psoriatic arthritis (PsA). All patients had indication for anti-TNFatherapy. Epidemiologic and clinical data were evaluated and a simple X-ray and tuberculin skin test (TST) were performed. The control group included 215 healthy individuals. The follow-up was 48 months to identify cases of aMI. TST positivity was higher in patients with AS (37.6%) than in RA (12.8%), PsA (18.8%) and JIA (6.8%) (p < 0.001). In the control group, TST positivity was 32.7%. Nine (3.43%) patients were diagnosed with aMI. The overall incidence rate of aMI was 86.93/100,000 person-years [95% confidence interval (CI) 23.6-217.9] for patients and 35.79/100,000 person-years (95% CI 12.4-69.6) for control group (p < 0.001). All patients who developed aMI had no evidence of LTBI at the baseline evaluation. Patients with CIA starting TNFa blockers and no evidence of LTBI at baseline, particularly with nonreactive TST, may have higher risk of aMI. PMID:26560983

  12. Molecular identification of clinical Nocardia isolates from India.

    Science.gov (United States)

    Rudramurthy, Shivaprakash M; Honnavar, Prasanna; Kaur, Harsimran; Samanta, Palash; Ray, Pallab; Ghosh, Anup; Chakrabarti, Arunaloke

    2015-10-01

    The epidemiology of nocardiosis is evolving with increasing number of Nocardia spp. causing human infection. In recent years, molecular techniques have been used to identify Nocardia spp. There are limited data available on the spectrum of Nocardia spp. isolated from clinical samples in India. Here, a molecular study was carried on 30 clinical isolates maintained in our National Culture Collection to evaluate the techniques used for identifying the agents. The isolates were identified by sequencing two promising genes: the 16S rRNA gene and hsp65. Both hsp65 and the 16S rRNA gene could reliably identify 90 % of Nocardia isolates, i.e. N. farcinica, N. cyriacigeorgica, N. brasiliensis, N. otitidiscaviarum, N. amamiensis and N. pneumoniae. The mean percentage dissimilarity of sequence identification was higher using the hsp65 gene (4 %, range 0-7.9 %) compared with the 16S rRNA gene (2.3 %, range 0-8.9 %). Two isolates that showed ambiguous results in both the short segment of the 16S rRNA gene and hsp65 sequences could be resolved by sequencing a larger fragment (∼1000 bp) of the 16S rRNA gene. Both of these isolates were identified as N. beijingensis with similarities of 99.8 and 100 % compared with the standard strain. Genotyping of N. cyriacigeorgica strains was performed using hsp65 gene sequences and compared with previously described genotypes. Our N. cyriacigeorgica isolates belonged to genotype 1 (n = 4) and genotype 2 (n = 2). The present study highlights a wide spectrum of Nocardia spp. in India and emphasizes the need for molecular techniques for identification to the species level. PMID:26202321

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

    Science.gov (United States)

    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

    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

  14. Inositol monophosphate phosphatase genes of Mycobacterium tuberculosis

    Directory of Open Access Journals (Sweden)

    Parish Tanya

    2010-02-01

    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.

  15. Comparison of epitope specificity of anti-heat shock protein 60/65 IgG type antibodies in the sera of healthy subjects, patients with coronary heart disease and inflammatory bowel disease

    OpenAIRE

    Füst, George; Uray, Katalin; Bene, László; Hudecz, Ferenc; Karádi, István; Prohászka, Zoltán

    2011-01-01

    Previously, we reported on the presence of antibodies to linear epitopes of human and mycobacterial 60 kD heat shock proteins (HSP) in the sera of healthy blood donors. Since many recent findings indicate that the levels of these antibodies may be altered in coronary heart disease (CHD) and also inflammatory bowel diseases (IBD), it seemed worthwhile to compare the epitope specificity of the anti-HSP60 and anti-HSP65 antibodies in the sera of patients with these diseases to those in healthy s...

  16. A randomised controlled trial of the effects of albendazole in pregnancy on maternal responses to mycobacterial antigens and infant responses to bacille Calmette-Guérin (BCG immunisation [ISRCTN32849447

    Directory of Open Access Journals (Sweden)

    Nampijja Margaret

    2005-12-01

    Full Text Available Abstract Background Maternal schistosomiasis and filariasis have been shown to influence infant responses to neonatal bacille Calmette-Guérin (BCG immunisation but the effects of maternal hookworm, and of de-worming in pregnancy, are unknown. Methods In Entebbe, Uganda, we conducted a randomised, double-blind, placebo-controlled trial of a single dose of 400 mg of albendazole in the second trimester of pregnancy. Neonates received BCG. Interferon-gamma (IFN-? and interleukin (IL-5 responses to a mycobacterial antigen (crude culture filtrate proteins (CFP of Mycobacterium tuberculosis were measured in a whole blood assay. We analysed results for binary variables using ?2 tests and logistic regression. We analysed continuous variables using Wilcoxon's tests. Results Maternal hookworm was associated with reduced maternal IFN-? responses to CFP (adjusted odds ratio for IFN-? > median response: 0.14 (95% confidence interval 0.02–0.83, p = 0.021. Conversely, maternal hookworm was associated with subsequent increased IFN-? responses in their one-year-old infants (adjusted OR 17.65 (1.20–258.66; p = 0.013. Maternal albendazole tended to reduce these effects. Conclusion Untreated hookworm infection in pregnancy was associated with reduced maternal IFN-? responses to mycobacterial antigens, but increased responses in their infants one year after BCG immunisation. The mechanisms of these effects, and their implications for protective immunity remain, to be determined.

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

    Scientific Electronic Library Online (English)

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

    1999-01-01

    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 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.

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

    Directory of Open Access Journals (Sweden)

    F. Bigi

    1999-01-01

    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.

  19. 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)

    José Maciel Rodrigues Júnior

    2004-08-01

    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.

  20. 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)

    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

    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 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.

  1. Gene expression

    International Nuclear Information System (INIS)

    We prepared probes for isolating functional pieces of the metallothionein locus. The probes enabled a variety of experiments, eventually revealing two mechanisms for metallothionein gene expression, the order of the DNA coding units at the locus, and the location of the gene site in its chromosome. Once the switch regulating metallothionein synthesis was located, it could be joined by recombinant DNA methods to other, unrelated genes, then reintroduced into cells by gene-transfer techniques. The expression of these recombinant genes could then be induced by exposing the cells to Zn2+ or Cd2+. We would thus take advantage of the clearly defined switching properties of the metallothionein gene to manipulate the expression of other, perhaps normally constitutive, genes. Already, despite an incomplete understanding of how the regulatory switch of the metallothionein locus operates, such experiments have been performed successfully

  2. Structure of Mycobacterium tuberculosis Rv2714, a representative of a duplicated gene family in Actinobacteria

    International Nuclear Information System (INIS)

    The crystal structure of Rv2714, a protein of unknown function from M. tuberculosis, has been determined at 2.6 Å resolution using single-wavelength anomalous diffraction methods. The gene Rv2714 from Mycobacterium tuberculosis, which codes for a hypothetical protein of unknown function, is a representative member of a gene family that is largely confined to the order Actinomycetales of Actinobacteria. Sequence analysis indicates the presence of two paralogous genes in most mycobacterial genomes and suggests that gene duplication was an ancient event in bacterial evolution. The crystal structure of Rv2714 has been determined at 2.6 Å resolution, revealing a trimer in which the topology of the protomer core is similar to that observed in a functionally diverse set of enzymes, including purine nucleoside phosphorylases, some carboxypeptidases, bacterial peptidyl-tRNA hydrolases and even the plastidic form of an intron splicing factor. However, some structural elements, such as a ?-hairpin insertion involved in protein oligomerization and a C-terminal ?-helical domain that serves as a lid to the putative substrate-binding (or ligand-binding) site, are only found in Rv2714 bacterial homologues and represent specific signatures of this protein family

  3. Comparison of a Semiautomated Commercial Repetitive-Sequence-Based PCR Method with Spoligotyping, 24-Locus Mycobacterial Interspersed Repetitive-Unit–Variable-Number Tandem-Repeat Typing, and Restriction Fragment Length Polymorphism-Based Analysis of IS6110 for Mycobacterium tuberculosis Typing

    OpenAIRE

    Brossier, F; C. Sola; Millot, G.; Jarlier, V; Veziris, N.; Sougakoff, W.

    2014-01-01

    Fifty-two multidrug-resistant isolates of Mycobacterium tuberculosis representative of the currently predominant lineages in France were analyzed using repetitive-sequence-based PCR (rep-PCR) DiversiLab (DL), spoligotyping, 24-locus mycobacterial interspersed repetitive-unit–variable-number tandem-repeat typing (MIRU-VNTR), and restriction fragment length polymorphism of IS6110 (IS6110-RFLP). DL, as opposed to MIRU-VNTR and IS6110-RFLP analysis, did not allow discrimination among half of the ...

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

    Directory of Open Access Journals (Sweden)

    Jin Ruiliang

    2011-08-01

    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.

  5. Gene therapy.

    OpenAIRE

    Mota Biosca, Anna

    1992-01-01

    Applications of gene therapy have been evaluated in virtually every oral tissue, and many of these have proved successful at least in animal models. While gene therapy will not be used routinely in the next decade, practitioners of oral medicine should be aware of the potential of this novel type of treatment that doubtless will benefit many patients with oral diseases.

  6. Trichoderma genes

    Science.gov (United States)

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

    2012-06-19

    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.

  7. Ulcera lingual como signo único de infección recurrente por micobacteria en un paciente con VIH/SIDA / Lingual ulcer as the only sign of recurrent mycobacterial infection in an HIV/AIDS-infected patient

    Scientific Electronic Library Online (English)

    Velia, Ramírez Amador; Gabriela, Anaya Saavedra; Imelda, González Ramírez; Juan Luis, Mosqueda Gómez; Lilly, Esquivel Pedraza; Edgardo, Reyes Gutiérrez; Juan, Sierra Madero.

    2005-04-01

    Full Text Available Se describe un paciente con VIH/SIDA en el que se identificó una infección por micobacteria en la mucosa bucal, probablemente tuberculosis, en un centro de referencia para VIH/SIDA de la Ciudad de México. El propósito del presente informe es describir los hallazgos clínicos e histológicos en un paci [...] ente con VIH/SIDA, quien después de haber sido tratado exitosamente para tuberculosis ganglionar 4 años antes, presentó una úlcera lingual como único signo que sugirió recurrencia de infección por micobacteria, probablemente tuberculosis. Hombre de 39 años de edad, atendido desde 1991 en el Instituto Nacional de Ciencias Médicas y Nutrición "Salvador Zubirán", por el diagnóstico de infección con VIH. En 1999, el paciente presentó tuberculosis ganglionar, recibiendo tratamiento antifímico con involución de las adenopatías y desaparición de los síntomas sistémicos. En mayo del 2003 acudió a consulta por presentar una úlcera superficial en lengua, dolorosa, de 4 meses de evolución, de 0.7 cm. de diámetro, bien circunscrita, crateriforme, con bordes ligeramente elevados, irregulares e indurados. El estudio histopatológico mostró inflamación granulomatosa crónica con células gigantes multinucleadas sugestivas de infección por micobacteria, lo cual hizo pensar en recurrencia de tuberculosis, por lo que se indicó rifampicina, pirazinamida, etambutol y estreptomicina. En junio del 2003 el paciente inició TARAA, que incluyó dos ITRAN y un ITRNN. La lesión lingual evolucionó favorablemente, con cicatrización parcial a la primera semana y remisión total a los 45 días del inicio del tratamiento antifímico; a los 7 meses de seguimiento permanece sin lesión. El presente caso tiene la particularidad de que la úlcera lingual fue la única manifestación de infección por micobacteria, sugestiva de tuberculosis, en un paciente con VIH/SIDA, que pudo ocurrir como resultado de la recurrencia del episodio previo de TB ganglionar. Abstract in english The report describes an HIV/AIDS patient seen at a referral center in Mexico City, in whom a mycobacterial infection in the oral mucosa, probably tuberculosis (TB) was identified. The purpose is to describe the clinical and histological findings in an HIV-infected patient, who after being treated su [...] ccessfully for tuberculous lymphangitis 4 years ago, presented with a lingual ulcer as the only suggestive sign of recurrence of mycobacterial infection, probably M. tuberculosis. A 39-year-old man seen inthe HIV clinic of the Instituto Nacional de Ciencias Médicas y Nutrición "Salvador Zubirán" in Mexico City since 1991 for HIV infection. In 1999 the patient developed tuberculous lymphangitis; he was managed with a 4-drug regimen for 12 months, with improvement of local and systemic symptoms. In May of 2003, the patient presented a painful superficial lingual ulcer, 0.7 cm in diameter, well circumscribed, crateriform with slightly elevated, irregular and indurated borders, of 4 months duration. The histopathological examination showed chronic granulomatous inflammation with giant multinucleated cells, suggestive of mycobacterial infection, and recurrence of TB was considered. Rifampin, isoniazide, pyrazinamide, ethambutol and streptomycin were administered. The lingual lesion improved with partial healing at the first week and total remission at 45 days after the beginning of the antituberculous treatment. In June, 2003, the patient began highly active antiretroviral therapy (HAART) that included two NRTIs and one NNRTI. At 7 months of follow-up, the patient remains free of lingual lesions. The particularity of the present case is that the lingual ulcer was the only sign of infection by mycobacteria, suggestive of TB, in an HIV/AIDS patient that probably represented a recurrence of a previous episode.

  8. 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)

    C. Coitinho

    2005-12-01

    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.

  9. Gene Silencing.

    Czech Academy of Sciences Publication Activity Database

    Kertbundit, Sunee; Ju?í?ek, Miloslav; Hall, T.C.

    Dordrecht : Springer, 2010 - (Jain, S.; Brar, D.), s. 631-652 ISBN 978-90-481-2966-9 Institutional research plan: CEZ:AV0Z50380511 Keywords : Gene Silencing * RISC complex Subject RIV: EB - Genetics ; Molecular Biology

  10. Gene therapy

    OpenAIRE

    KAY, MARK A.; Liu, Dexi; Hoogerbrugge, Peter M.

    1997-01-01

    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.

  11. Studying Genes

    Science.gov (United States)

    ... the body's functions. Genes are passed from one generation to the next. What is a genome? A genome is all ... Highlight Header Highlight Body Related Links Up to top This page last reviewed on August ... U.S. Department of Health and Human Services National Institutes of Health: NIH...Turning Discovery Into ...

  12. 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)

    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

    Full Text Available 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

  13. Comparison of a semiautomated commercial repetitive-sequence-based PCR method with spoligotyping, 24-locus mycobacterial interspersed repetitive-unit-variable-number tandem-repeat typing, and restriction fragment length polymorphism-based analysis of IS6110 for Mycobacterium tuberculosis typing.

    Science.gov (United States)

    Brossier, F; Sola, C; Millot, G; Jarlier, V; Veziris, N; Sougakoff, W

    2014-11-01

    Fifty-two multidrug-resistant isolates of Mycobacterium tuberculosis representative of the currently predominant lineages in France were analyzed using repetitive-sequence-based PCR (rep-PCR) DiversiLab (DL), spoligotyping, 24-locus mycobacterial interspersed repetitive-unit-variable-number tandem-repeat typing (MIRU-VNTR), and restriction fragment length polymorphism of IS6110 (IS6110-RFLP). DL, as opposed to MIRU-VNTR and IS6110-RFLP analysis, did not allow discrimination among half of the isolates, an indication of comparatively lower resolving power. PMID:25210067

  14. Identificação e genotipagem de Mycobacterium bovis em bovinos positivos no teste intradérmico para tuberculose em Mato Grosso do Sul / Identification and genotyping of Mycobacterium bovis from positive cattle in skin test for tuberculosis in the State of Mato Grosso do Sul, Brazil

    Scientific Electronic Library Online (English)

    Daniela de O., Cazola; Klaudia dos S.G., Jorge; Martín J., Zumárraga; Antônio F., Souza-Filho; Flábio R., Araújo; Ana Luiza A.R., Osório.

    2015-02-01

    Full Text Available Neste estudo, realizou-se genotipagem de isolados de Mycobacterium bovis, provenientes de amostras de tecidos de bovinos positivos no teste cervical comparativo (TCC) para tuberculose em Mato Grosso do Sul, por meio da técnica de spoligotyping. Tecidos de 13 bovinos positivos, oriundos de diferentes [...] municípios do estado, foram cultivados em meio de Stonebrink. As colônias resultantes foram submetidas à coloração de Ziehl-Neelsen e todos os isolados apresentaram características tintoriais de BAAR. Os 13 isolados de BAAR foram identificados por PCR multiplex (mPCR). O gene hsp65 foi alvo para identificação de Mycobacterium spp, a sequência de inserção IS6110 foi alvo para identificação de complexo Mycobacterium tuberculosis (CMT) e a região rvd1rv2031c foi explorada para detecção de M. bovis. Os isolados micobacterianos foram genotipados pela técnica de spoligotyping. Dos 13 bovinos, sete tinham pelo menos uma lesão sugestiva de tuberculose em linfonodos retrofaríngeos, parotídeos e pulmonares ou no pulmão, e em seis não foram encontradas lesões visíveis sugestivas da doença. Na mPCR, 11/13 (84,6%) isolados foram positivos para Mycobacterium spp; 8/13 (61,5%) positivos para CMT e 7/13 (53,8%) positivos para M. bovis. Com base no spoligotyping, oito isolados de BAAR foram agrupados dentro de três diferentes agrupamentos de genótipos e uma amostra remanescente apresentou perfil único, sendo quatro isolados com padrão de espoligotipo SB0121, dois SB1145, dois SB0881 e um SB0140. A técnica de spoligotyping demonstrou que há diversidade genética entre os espoligotipos presentes no estado de Mato Grosso do Sul, embora predomine o perfil SB0121 Abstract in english Spoligotyping was performed in the present study to genotype Mycobacterium bovis isolates obtained from tissues of cattle that were positive in the comparative intradermal tuberculin test (CITT) in the state of Mato Grosso do Sul (Brazil). Tissue samples from 13 positive cattle from different munici [...] palities of the state were cultured using a Stonebrink medium. The resulting colonies were subjected to Ziehl-Neelsen staining and all isolates exhibited the staining characteristics of AFB. The 13 isolates of AFB were identified by means of a multiplex PCR (mPCR) assay. The hsp65 gene was targeted for the identification of Mycobacterium spp., whereas the IS6110 insertion sequence was targeted for the identification of the Mycobacterium tuberculosis complex (MTC) and the rvd1rv2031c region was explored for the detection of Mycobacterium bovis. The spoligotyping assay was performed to genotype mycobacterial isolates. Of the 13 cattle, seven had at least one lesion suggestive of tuberculosis in the retropharyngeal, parotid and lung lymph nodes or lung. The remaining six exhibited no lesions suggestive of the disease. In the mPCR, 11 of the 13 isolates (84.6%) were positive for Mycobacterium spp., 8/13 (61.5%) were positive for the MTC and 7/13 (53.8%) were positive for M. bovis. Based on the spoligotyping, eight isolates were grouped into three different groups of genotypes and one isolate exhibited an orphan type. Four isolates exhibited spoligotype pattern SB0121, while two isolates were associated with the pattern SB1145, another two were associated with pattern SB0881 and one was associated with pattern SB0140. Spoligotyping confirmed the genetic diversity present among isolates found in the state of Mato Grosso do Sul. In addition, SB0121 was confirmed as the predominant profile.

  15. Vaccination with a potent DNA vaccine targeting B-cell epitopes of hGRP induces prophylactic and therapeutic antitumor activity in vivo.

    Science.gov (United States)

    Yong, Lu; Huiyong, Zhang; Jing, Hou; Huaqian, Wang; Xiangbing, Hu; Yanjun, Ma; Xiaoyu, Ge; Li, Huang; Yanan, Yang; Rongyue, Cao; Hao, Fan; Jingjing, Liu; Jie, Wu

    2010-04-01

    Gastrin-releasing peptide (GRP), a bombesin-like peptide, is an autocrine or paracrine growth factor that can stimulate the growth of various cancer cells, making it an ideal target antigen to develop vaccines against cancer. In this study, we developed a novel DNA vaccine that encodes six tandem repeats of B-cell epitope GRP(18-27) (GRP6) flanked by HSP65 as carrier and four tandem repeats of mycobacterial HSP70(407-426) (M4) as helper T-cell epitopes for enhancement of immunogenicity. When intramuscularly immunized to mice, this anti-GRP DNA vaccine-induced GRP-specific antibody (Ab) responses that were at least 10-fold higher in magnitude compared with HSP65-GRP6 protein vaccine. Both prophylactic and therapeutic antitumor immunities induced by vaccination significantly suppressed the growth of GRP-dependent prostate carcinoma RM-1 in vivo and prolonged the survival of tumor-inoculated mice. Out results also showed that the immune sera with high titer of GRP-specific Abs effectively inhibited the growth of tumor in mice and dose dependently inhibited proliferation of cultured RM-1 cells in vitro, suggesting that the GRP neutralizing Ab is responsible for the protective and therapeutic antitumor activity of vaccination. These findings may be of great importance in the further exploration of the applications of growth factors identified in human in cancer therapy. PMID:20130655

  16. 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.)

  17. Imaging reporter gene for monitoring gene therapy

    International Nuclear Information System (INIS)

    Scintigraphic images can be obtained to document gene function at cellular level. This approach is presented here and the use of a reporter gene to monitor gene therapy is described. Two main ways are presented: either the use of a reporter gene coding for an enzyme the action of which will be monitored by radiolabeled pro-drug, or a cellular receptor gene, the action of which is documented by a radio labeled cognate receptor ligand. (author)

  18. 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)

    Filipe Accioly de, Gusmão; Lênio, Alvarenga; Luciene, Barbosa; Jorge, Sampaio; Sylvia Cardoso, Leão; Ana Luisa, Hofling-Lima; Denise de, Freitas.

    2005-08-01

    Full Text Available 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.

  19. Identificação de micobactérias não tuberculosas isoladas de sítios estéreis em pacientes em um hospital universitário na cidade do Rio de Janeiro Identification of nontuberculous mycobacteria isolated from clinical sterile sites in patients at a university hospital in the city of Rio de Janeiro, Brazil

    Directory of Open Access Journals (Sweden)

    Simone Gonçalves Senna

    2011-08-01

    Full Text Available OBJETIVO: Identificar micobactérias não tuberculosas (MNT isoladas de sítios estéreis em pacientes internados no Hospital Universitário Clementino Fraga Filho, Rio de Janeiro (RJ entre 2001 e 2006. MÉTODOS: Durante o período do estudo, 34 isolados de MNT de sítios estéreis de 14 pacientes, a maioria HIV positivos, foram submetidos a identificação fenotípica e hsp65 PCR-restriction enzyme analysis (PRA, análise por enzimas de restrição por PCR do gene hsp65. RESULTADOS: A maioria dos isolados foi identificada como Mycobacterium avium, seguida por M. monacense, M. kansasii e M. abscessus em menores proporções. CONCLUSÕES: A combinação de PRA, um método relativamente simples e de baixo custo, com algumas características fenotípicas pode fornecer a identificação correta de MNT na rotina de laboratórios clínicos.OBJECTIVE: To identify nontuberculous mycobacteria (NTM isolated from sterile sites in patients hospitalized between 2001 and 2006 at the Clementino Fraga Filho University Hospital, located in the city of Rio de Janeiro, Brazil. METHODS: During the study period, 34 NTM isolates from sterile sites of 14 patients, most of whom were HIV-positive, were submitted to phenotypic identification and hsp65 PCR-restriction enzyme analysis (PRA. RESULTS: Most isolates were identified as Mycobacterium avium, followed by M. monacense, M. kansasii, and M. abscessus. CONCLUSIONS: The combination of PRA, a relatively simple and inexpensive method, with the evaluation of a few phenotypic characteristics can allow NTM to be accurately identified in the routine of clinical laboratories.

  20. Identificação de micobactérias não tuberculosas isoladas de sítios estéreis em pacientes em um hospital universitário na cidade do Rio de Janeiro / Identification of nontuberculous mycobacteria isolated from clinical sterile sites in patients at a university hospital in the city of Rio de Janeiro, Brazil

    Scientific Electronic Library Online (English)

    Simone Gonçalves, Senna; Ana Grazia, Marsico; Gisele Betzler de Oliveira, Vieira; Luciana Fonseca, Sobral; Philip Noel, Suffys; Leila de Souza, Fonseca.

    2011-08-01

    Full Text Available OBJETIVO: Identificar micobactérias não tuberculosas (MNT) isoladas de sítios estéreis em pacientes internados no Hospital Universitário Clementino Fraga Filho, Rio de Janeiro (RJ) entre 2001 e 2006. MÉTODOS: Durante o período do estudo, 34 isolados de MNT de sítios estéreis de 14 pacientes, a maior [...] ia HIV positivos, foram submetidos a identificação fenotípica e hsp65 PCR-restriction enzyme analysis (PRA, análise por enzimas de restrição por PCR do gene hsp65). RESULTADOS: A maioria dos isolados foi identificada como Mycobacterium avium, seguida por M. monacense, M. kansasii e M. abscessus em menores proporções. CONCLUSÕES: A combinação de PRA, um método relativamente simples e de baixo custo, com algumas características fenotípicas pode fornecer a identificação correta de MNT na rotina de laboratórios clínicos. Abstract in english OBJECTIVE: To identify nontuberculous mycobacteria (NTM) isolated from sterile sites in patients hospitalized between 2001 and 2006 at the Clementino Fraga Filho University Hospital, located in the city of Rio de Janeiro, Brazil. METHODS: During the study period, 34 NTM isolates from sterile sites o [...] f 14 patients, most of whom were HIV-positive, were submitted to phenotypic identification and hsp65 PCR-restriction enzyme analysis (PRA). RESULTS: Most isolates were identified as Mycobacterium avium, followed by M. monacense, M. kansasii, and M. abscessus. CONCLUSIONS: The combination of PRA, a relatively simple and inexpensive method, with the evaluation of a few phenotypic characteristics can allow NTM to be accurately identified in the routine of clinical laboratories.

  1. STRESS AND ATHEROSCLEROSIS: MAY HSP60 BE THE MOLECULAR LINK?

    Directory of Open Access Journals (Sweden)

    Luana Lipari

    2009-01-01

    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.

  2. Identification of contaminating bacteria when attempting to isolate Mycobacterium avium subsp. paratuberculosis (MAP from bovine faecal and tissue samples using the BACTEC MGIT 960 system

    Directory of Open Access Journals (Sweden)

    P Steuer

    2015-01-01

    Full Text Available Diagnosis of Mycobacterium avium subsp. paratuberculosis infection by liquid culture is sensitive, faster than conventional solid culture and automated. However, a disadvantage of these culture systems is the potential for high frequency of culture contamination. Contaminant bacteria were identified as a step toward better contaminant control. No mycobacteria were detected by mycobacterial Polymerase Chain Reaction-Restriction Enzyme Analysis (PRA-hsp65. Ribosomal Intergenic Spacer Analysis (RISA followed by sequence analysis identified Paenibacillus sp., Enterobacteriaceae and Pseudomonas aeruginosa as common contaminants. The present study aimed to identify a representative sample of contaminants encountered when culturing clinical faecal samples from Chilean cattle. Further studies involving a larger and more representative sample of animals are required to extrapolate the results to a broader population.

  3. Gene gymnastics

    Science.gov (United States)

    Vijayachandran, Lakshmi S; Thimiri Govinda Raj, Deepak B; Edelweiss, Evelina; Gupta, Kapil; Maier, Josef; Gordeliy, Valentin; Fitzgerald, Daniel J; Berger, Imre

    2013-01-01

    Most essential activities in eukaryotic cells are catalyzed by large multiprotein assemblies containing up to ten or more interlocking subunits. The vast majority of these protein complexes are not easily accessible for high resolution studies aimed at unlocking their mechanisms, due to their low cellular abundance and high heterogeneity. Recombinant overproduction can resolve this bottleneck and baculovirus expression vector systems (BEVS) have emerged as particularly powerful tools for the provision of eukaryotic multiprotein complexes in high quality and quantity. Recently, synthetic biology approaches have begun to make their mark in improving existing BEVS reagents by de novo design of streamlined transfer plasmids and by engineering the baculovirus genome. Here we present OmniBac, comprising new custom designed reagents that further facilitate the integration of heterologous genes into the baculovirus genome for multiprotein expression. Based on comparative genome analysis and data mining, we herein present a blueprint to custom design and engineer the entire baculovirus genome for optimized production properties using a bottom-up synthetic biology approach. PMID:23328086

  4. Antigen stimulation of peripheral blood mononuclear cells from Mycobacterium bovis infected cattle yields evidence for a novel gene expression program

    Directory of Open Access Journals (Sweden)

    Zhao Yingdong

    2008-09-01

    Full Text Available Abstract Background Bovine tuberculosis (BTB caused by Mycobacterium bovis continues to cause substantial losses to global agriculture and has significant repercussions for human health. The advent of high throughput genomics has facilitated large scale gene expression analyses that present a novel opportunity for revealing the molecular mechanisms underlying mycobacterial infection. Using this approach, we have previously shown that innate immune genes in peripheral blood mononuclear cells (PBMC from BTB-infected animals are repressed in vivo in the absence of exogenous antigen stimulation. In the present study, we hypothesized that the PBMC from BTB-infected cattle would display a distinct gene expression program resulting from exposure to M. bovis. A functional genomics approach was used to examine the immune response of BTB-infected (n = 6 and healthy control (n = 6 cattle to stimulation with bovine tuberculin (purified protein derivative – PPD-b in vitro. PBMC were harvested before, and at 3 h and 12 h post in vitro stimulation with bovine tuberculin. Gene expression changes were catalogued within each group using a reference hybridization design and a targeted immunospecific cDNA microarray platform (BOTL-5 with 4,800 spot features representing 1,391 genes. Results 250 gene spot features were significantly differentially expressed in BTB-infected animals at 3 h post-stimulation contrasting with only 88 gene spot features in the non-infected control animals (P ? 0.05. At 12 h post-stimulation, 56 and 80 gene spot features were differentially expressed in both groups respectively. The results provided evidence of a proinflammatory gene expression profile in PBMC from BTB-infected animals in response to antigen stimulation. Furthermore, a common panel of eighteen genes, including transcription factors were significantly expressed in opposite directions in both groups. Real-time quantitative reverse transcription PCR (qRT-PCR demonstrated that many innate immune genes, including components of the TLR pathway and cytokines were differentially expressed in BTB-infected (n = 8 versus control animals (n = 8 after stimulation with bovine tuberculin. Conclusion The PBMC from BTB-infected animals exhibit different transcriptional profiles compared with PBMC from healthy control animals in response to M. bovis antigen stimulation, providing evidence of a novel gene expression program due to M. bovis exposure.

  5. Genes and Psoriasis

    Science.gov (United States)

    ... to find out more! Email * Zipcode Genes and Psoriasis Genes hold the key to understanding how the ... Are some genes linked to specific kinds of psoriasis? At the University of Utah, Drs. Gerald Krueger ...

  6. An atypical mycobacterial infection of the shoulder

    OpenAIRE

    Talbot, Christopher L.; Rhodes, Bryan

    2012-01-01

    Mycobacterium malmoense is an acid-fast non-tuberculous organism that most commonly causes pulmonary infection. Extrapulmonary infection has also been reported. With an increased emphasis being placed on the clinical importance of this organism, especially within Europe, we report the first case of septic arthritis of the shoulder caused by this organism. We also highlight the importance of considering atypical mycobacterium infection in the differential diagnosis of shoulder infection and is...

  7. Nontuberculous Mycobacterial Infections in Cystic Fibrosis.

    Science.gov (United States)

    Martiniano, Stacey L; Nick, Jerry A; Daley, Charles L

    2016-03-01

    Nontuberculous mycobacteria (NTM) are important emerging cystic fibrosis (CF) pathogens, with estimates of prevalence ranging from 6% to 13%. Diagnosis of NTM disease in patients with CF is challenging, as the infection may remain indolent in some, without evidence of clinical consequence, whereas other patients suffer significant morbidity and mortality. Treatment requires prolonged periods of multiple drugs and varies depending on NTM species, resistance pattern, and extent of disease. The development of a disease-specific approach to the diagnosis and treatment of NTM infection in CF patients is a research priority, as a lifelong strategy is needed for this high-risk population. PMID:26857770

  8. 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 [123/131I]I V R F U imaging with the H S V-1 t k+ reporter gene will be presented

  9. Identifying Gene Interaction Enrichment for Gene Expression Data

    OpenAIRE

    Zhang, Jigang; Li, Jian; Deng, Hong-Wen

    2009-01-01

    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 ...

  10. Essential Bacillus subtilis genes

    DEFF Research Database (Denmark)

    Kobayashi, K.; Ehrlich, S.D.; Albertini, A.; Amati, G.; Andersen, K.K.; Arnaud, M.; Asai, K.; Ashikaga, S.; Aymerich, S.; Bessieres, P.; Boland, F.; Brignell, S.C.; Bron, S.; Bunai, K.; Chapuis, J.; Christiansen, L.C.; Danchin, A.; Debarbouille, M.; Dervyn, E.; Deuerling, E.; Devine, K.; Devine, S.K.; Dreesen, O.; Errington, J.; Fillinger, S.; Foster, S.J.; Fujita, Y.; Galizzi, A.; Gardan, R.; Eschevins, C.; Fukushima, T.; Haga, K.; Harwood, C.R.; Hecker, M.; Hosoya, D.; Hullo, M.F.; Kakeshita, H.; Karamata, D.; Kasahara, Y.; Kawamura, F.; Koga, K.; Koski, P.; Kuwana, R.; Imamura, D.; Ishimaru, M.; Ishikawa, S.; Ishio, I.; Le Coq, D.; Masson, A.; Mauel, C.; Meima, R.; Mellado, R.P.; Moir, A.; Moriya, S.; Nagakawa, E.; Nanamiya, H.; Nakai, S.; Nygaard, P.; Ogura, M.; Ohanan, T.; O'Reilly, M.; O'Rourke, M.; Pragai, Z.; Pooley, H.M.; Rapoport, G.; Rawlins, J.P.; Rivas, L.A.; Rivolta, C.; Sadaie, A.; Sadaie, Y.; Sarvas, M.; Sato, T.; Saxild, Hans Henrik; Scanlan, E.; Schumann, W.; Seegers, J.F.M.L.; Sekiguchi, J.; Sekowska, A.; Seror, S.J.; Simon, M.; Stragier, P.; Studer, R.; Takamatsu, H.; Tanaka, T.; Takeuchi, M.; Thomaides, H.B.; Vagner, V.; van Dijl, J.M.; Watabe, K.; Wipat, A.; Yamamoto, H.; Yamamoto, M.; Yamamoto, Y.; Yamane, K.; Yata, K.; Yoshida, K.; Yoshikawa, H.; Zuber, U.; Ogasawara, N.

    2003-01-01

    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...

  11. Autism and Genes

    Science.gov (United States)

    National Institutes of Health, 2005

    2005-01-01

    This document defines and discusses autism and how genes play a role in the condition. Answers to the following questions are covered: (1) What are genes? (2) What is autism? (3) What causes autism? (4) Why study genes to learn about autism? (5) How do researchers look for the genes involved in autism? (screen the whole genome; conduct cytogenetic…

  12. Modelling Gene Regulatory Networks

    OpenAIRE

    Ingram, Piers J

    2008-01-01

    This thesis presents the results of mathematical modeling of both individual genes and small networks of genes. The regulation of gene activity is essential for the proper functioning of cells, which employ a variety of molecular mechanisms to control gene expression. Despite this, there is considerable variation in the precise number and timing of protein molecules that are produced. This is because gene expression is fundamentally a noisy process, subject to a number of sourc...

  13. Essential Bacillus subtilis genes

    DEFF Research Database (Denmark)

    Kobayashi, K.; Ehrlich, S.D.; Albertini, A.; Amati, G.; Andersen, K.K.; Arnaud, M.; Asai, K.; Ashikaga, S.; Aymerich, S.; Bessieres, P.; Boland, F.; Brignell, S.C.; Bron, S.; Bunai, K.; Chapuis, J.; Christiansen, L.C.; Danchin, A.; Debarbouille, M.; Dervyn, E.; Deuerling, E.; Devine, K.; Devine, S.K.; Dreesen, O.; Errington, J.; Fillinger, S.; Foster, S.J.; Fujita, Y.; Galizzi, A.; Gardan, R.; Eschevins, C.; Fukushima, T.; Haga, K.; Harwood, C.R.; Hecker, M.; Hosoya, D.; Hullo, M.F.; Kakeshita, H.; Karamata, D.; Kasahara, Y.; Kawamura, F.; Koga, K.; Koski, P.; Kuwana, R.; Imamura, D.; Ishimaru, M.; Ishikawa, S.; Ishio, I.; Le Coq, D.; Masson, A.; Mauel, C.; Meima, R.; Mellado, R.P.; Moir, A.; Moriya, S.; Nagakawa, E.; Nanamiya, H.; Nakai, S.; Nygaard, P.; Ogura, M.; Ohanan, T.; O'Reilly, M.; O'Rourke, M.; Pragai, Z.; Pooley, H.M.; Rapoport, G.; Rawlins, J.P.; Rivas, L.A.; Rivolta, C.; Sadaie, A.; Sadaie, Y.; Sarvas, M.; Sato, T.; Saxild, Hans Henrik; Scanlan, E.; Schumann, W.; Seegers, J.F.M.L.; Sekiguchi, J.; Sekowska, A.; Seror, S.J.; Simon, M.; Stragier, P.; Studer, R.; Takamatsu, H.; Tanaka, T.; Takeuchi, M.; Thomaides, H.B.; Vagner, V.; van Dijl, J.M.; Watabe, K.; Wipat, A.; Yamamoto, H.; Yamamoto, M.; Yamamoto, Y.; Yamane, K.; Yata, K.; Yoshida, K.; Yoshikawa, H.; Zuber, U.; Ogasawara, N.

    2003-01-01

    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...

  14. Gene symbol precision.

    Science.gov (United States)

    Bennani-Baiti, Barbara; Bennani-Baiti, Idriss M

    2012-01-10

    Several gene databases, including heavily used ones such as the National Center for Biotechnology Information (NCBI) database, erroneously assign, on occasion, literature references to genes or proteins. These mistakes are mostly due to an overlap in gene aliases, whereby two distinct genes share a pseudonym. This is particularly confusing when the gene products have also biological properties in common, are part of signaling pathways that cross-talk to one another, or are regulated by the same effectors. We present examples spanning several research fields including apoptosis, ubiquitin-dependent degradation, signaling by Notch, Wnt, and small G proteins, transporters of glutathione conjugates of electrophiles, and mitochondrial and ribosomal RNA genes. To solve the problem, we argue in favor of including Entrez gene numbers in papers submitted for publication as unique gene identifiers to allow precise identification of genes and species studied. PMID:22019431

  15. Interferon-? Gene Polymorphism in Pulmonary Tuberculosis: Performance in TB-Endemic Warao Amerindians population

    Directory of Open Access Journals (Sweden)

    Zaida Alicia Araujo

    2015-09-01

    Full Text Available Polymorphisms in the cytokine genes are known to influence cytokine levels and may be associated with outcome of infections. Interferon-? is the most important cytokine in resistance to mycobacterial diseases and common variants of IFN-gamma (IFN-? gene could be related to tuberculosis (TB susceptibility. The present study determined whether a pattern of a functional single-nucleotide polymorphism (SNP was present and could predispose Warao indigenous to infection by Mycobacterium tuberculosis. We determined the distribution of the IFN-?+874T/A polymorphism in Warao indigenous population of 24 patients with pulmonary TB and 111 healthy controls. As compared to Warao indigenous and Caucasian populations, Warao indigenous TB cases and controls showed a higher frequency of the IFN-? allele SNP (+874A; 23(95.8% and 108(97.3%, respectively, whose phenotypic expression is associated with decreased production of this cytokine. Indigenous homozygous for IFN-? (+874 A allele had 3.59-fold increased risk of developing tuberculosis (95% confidence interval, 2.60-4.96, p =0.0001. A decrease in the frequency of AT genotype was observed in TB cases (4.16% and controls (0.90%, moreover, the frequency of TT genotype was also decreased in controls (1.80%, while that this genotype was not observed among patients. The findings suggest that the presence of an AA genotype was frequent including cases and control indigenous, so given the role played by IFN-? in facilitating macrophage containment of M. tuberculosis; a low production of interferon-? in individuals homozygous for the (+874 A allele could contribute to increased risk of developing tuberculosis among Warao indigenous.

  16. Human Gene Therapy: Genes without Frontiers?

    Science.gov (United States)

    Simon, Eric J.

    2002-01-01

    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…

  17. Genes and Hearing Loss

    Science.gov (United States)

    ... Find an ENT Doctor Near You Genes and Hearing Loss Genes and Hearing Loss Patient Health Information News media interested in ... One of the most common birth defects is hearing loss or deafness (congenital), which can affect as ...

  18. Tumor targeted gene therapy

    International Nuclear Information System (INIS)

    Knowledge of molecular mechanisms governing malignant transformation brings new opportunities for therapeutic intervention against cancer using novel approaches. One of them is gene therapy based on the transfer of genetic material to an organism with the aim of correcting a disease. The application of gene therapy to the cancer treatment had led to the development of new experimental approaches such as suicidal gene therapy, inhibition of oncogenes and restoration of tumor-suppressor genes. Suicidal gene therapy is based on the expression in tumor cells of a gene encoding an enzyme that converts a prodrug into a toxic product. Representative suicidal genes are Herpes simplex virus type 1 thymidine kinase (HSV1-tk) and cytosine deaminase (CD). Especially, physicians and scientists of nuclear medicine field take an interest in suicidal gene therapy because they can monitor the location and magnitude, and duration of expression of HSV1-tk and CD by PET scanner

  19. What Is a Gene?

    Science.gov (United States)

    ... think about all the many different breeds of dogs. They all have the genes that make them dogs instead of cats, fish, or people. But those same genes that make a dog a dog also make different dog traits. So ...

  20. Tumor targeted gene therapy

    Energy Technology Data Exchange (ETDEWEB)

    Kang, Joo Hyun [Korea Institute of Radiological and Medical Science, Seoul (Korea, Republic of)

    2006-10-15

    Knowledge of molecular mechanisms governing malignant transformation brings new opportunities for therapeutic intervention against cancer using novel approaches. One of them is gene therapy based on the transfer of genetic material to an organism with the aim of correcting a disease. The application of gene therapy to the cancer treatment had led to the development of new experimental approaches such as suicidal gene therapy, inhibition of oncogenes and restoration of tumor-suppressor genes. Suicidal gene therapy is based on the expression in tumor cells of a gene encoding an enzyme that converts a prodrug into a toxic product. Representative suicidal genes are Herpes simplex virus type 1 thymidine kinase (HSV1-tk) and cytosine deaminase (CD). Especially, physicians and scientists of nuclear medicine field take an interest in suicidal gene therapy because they can monitor the location and magnitude, and duration of expression of HSV1-tk and CD by PET scanner.

  1. Community structure and PAH ring-hydroxylating dioxygenase genes of a marine pyrene-degrading microbial consortium.

    Science.gov (United States)

    Gallego, Sara; Vila, Joaquim; Tauler, Margalida; Nieto, José María; Breugelmans, Philip; Springael, Dirk; Grifoll, Magdalena

    2014-07-01

    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

  2. Evolutionary Fingerprinting of Genes

    OpenAIRE

    KOSAKOVSKY POND, Sergei L.; Scheffler, Konrad; Gravenor, Michael B.; Poon, Art F.Y.; Simon D.W. Frost

    2009-01-01

    Over time, natural selection molds every gene into a unique mosaic of sites evolving rapidly or resisting change—an “evolutionary fingerprint” of the gene. Aspects of this evolutionary fingerprint, such as the site-specific ratio of nonsynonymous to synonymous substitution rates (dN/dS), are commonly used to identify genetic features of potential biological interest; however, no framework exists for comparing evolutionary fingerprints between genes. We hypothesize that protein-coding genes wi...

  3. Genes underlying altruism

    OpenAIRE

    Thompson, Graham J; Hurd, Peter L; Crespi, Bernard J

    2013-01-01

    William D. Hamilton postulated the existence of ‘genes underlying altruism’, under the rubric of inclusive fitness theory, a half-century ago. Such genes are now poised for discovery. In this article, we develop a set of intuitive criteria for the recognition and analysis of genes for altruism and describe the first candidate genes affecting altruism from social insects and humans. We also provide evidence from a human population for genetically based trade-offs, underlain by oxytocin-system ...

  4. Cochlear Gene Therapy

    OpenAIRE

    Lustig, Lawrence R.; Akil, Omar

    2012-01-01

    The purpose of this review is to highlight recent advances in cochlear gene therapy over the past several years. Cochlear gene therapy has undergone tremendous advances over the past decade. Beginning with some groundbreaking work in 2005 documenting hair cell regeneration using virallymediated delivery of the mouse atonal 1 gene, gene therapy is now being explored as a possible treatment for a variety of causes of hearing loss.

  5. Discovering genes underlying QTL

    International Nuclear Information System (INIS)

    A map-based approach has allowed scientists to discover few genes at a time. In addition, the reproductive barrier between cultivated rice and wild relatives has prevented us from utilizing the germ plasm by a map-based approach. Most genetic traits important to agriculture or human diseases are manifested as observable, quantitative phenotypes called Quantitative Trait Loci (QTL). In many instances, the complexity of the phenotype/genotype interaction and the general lack of clearly identifiable gene products render the direct molecular cloning approach ineffective, thus additional strategies like genome mapping are required to identify the QTL in question. Genome mapping requires no prior knowledge of the gene function, but utilizes statistical methods to identify the most likely gene location. To completely characterize genes of interest, the initially mapped region of a gene location will have to be narrowed down to a size that is suitable for cloning and sequencing. Strategies for gene identification within the critical region have to be applied after the sequencing of a potentially large clone or set of clones that contains this gene(s). Tremendous success of positional cloning has been shown for cloning many genes responsible for human diseases, including cystic fibrosis and muscular dystrophy as well as plant disease resistance genes. Genome and QTL mapping, positional cloning: the pre-genomics era, comparative approaches to gene identification, and positional cloning: the genomics era are discussed in the report. (M. Suetake)

  6. Discovering genes underlying QTL

    Energy Technology Data Exchange (ETDEWEB)

    Vanavichit, Apichart [Kasetsart University, Kamphaengsaen, Nakorn Pathom (Thailand)

    2002-02-01

    A map-based approach has allowed scientists to discover few genes at a time. In addition, the reproductive barrier between cultivated rice and wild relatives has prevented us from utilizing the germ plasm by a map-based approach. Most genetic traits important to agriculture or human diseases are manifested as observable, quantitative phenotypes called Quantitative Trait Loci (QTL). In many instances, the complexity of the phenotype/genotype interaction and the general lack of clearly identifiable gene products render the direct molecular cloning approach ineffective, thus additional strategies like genome mapping are required to identify the QTL in question. Genome mapping requires no prior knowledge of the gene function, but utilizes statistical methods to identify the most likely gene location. To completely characterize genes of interest, the initially mapped region of a gene location will have to be narrowed down to a size that is suitable for cloning and sequencing. Strategies for gene identification within the critical region have to be applied after the sequencing of a potentially large clone or set of clones that contains this gene(s). Tremendous success of positional cloning has been shown for cloning many genes responsible for human diseases, including cystic fibrosis and muscular dystrophy as well as plant disease resistance genes. Genome and QTL mapping, positional cloning: the pre-genomics era, comparative approaches to gene identification, and positional cloning: the genomics era are discussed in the report. (M. Suetake)

  7. A Pilot Study of Gene/Gene and Gene/Environment Interactions in Alzheimer Disease

    OpenAIRE

    Ghebranious, Nader; Mukesh, Bickol; Giampietro, Philip F.; Glurich, Ingrid; Mickel, Susan F.; Waring, Stephen C.; McCarty, Catherine A.

    2011-01-01

    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.

  8. Gene expression in fungi

    Directory of Open Access Journals (Sweden)

    A. Kalkanci

    2011-06-01

    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.

  9. Interactions of OxyR with the promoter region of the oxyR and ahpC genes from Mycobacterium leprae and Mycobacterium tuberculosis.

    Science.gov (United States)

    Dhandayuthapani, S; Mudd, M; Deretic, V

    1997-01-01

    In contrast to the intact oxyR gene (a homolog of the central regulator of peroxide stress response in enteric bacteria) in Mycobacterium leprae, this gene is inactive in all strains of M. tuberculosis. In both species, oxyR is divergently transcribed from ahpC, which encodes a homolog of alkyl hydroperoxide reductase. To initiate investigations of the regulation of oxidative stress in mycobacteria and consequences of the elimination of oxyR in M. tuberculosis, in this work we tested the hypothesis that mycobacterial OxyR acts as a DNA binding protein and analyzed its interactions with the oxyR and ahpC promoters. M. leprae OxyR was overproduced and purified, and its binding to the oxyR-ahpC intergenic region of M. leprae was demonstrated. By using a sequential series of overlapping DNA fragments, the minimal OxyR binding site was delimited to a 30-bp DNA segment which included a palindromic sequence conforming with the established rules for the LysR family of regulators. A consensus sequence for the mycobacterial OxyR recognition site (cTTATCggc-N3-gccGATAAg) was deduced based on its conservation in different mycobacteria. A variance in two potentially critical nucleotides within this site was observed in M. tuberculosis, in keeping with its reduced affinity for OxyR. Transcription of plasmid-borne M. leprae oxyR and ahpC was investigated in M. smegmatis and M. bovis BCG by S1 nuclease protection and transcriptional fusion analyses. Two mRNA 5' ends were detected in each direction: (i) P1oxyR and P2oxyR and (ii) P1ahpC and P2ahpC. The binding site for OxyR overlapped P1oxyR, reminiscent of the autoregulatory loops controlling expression of oxyR in enteric bacteria and characteristic of the LysR superfamily in general. This site was also centered 65 bp upstream of P1ahpC, matching the usual position of LysR-type recognition sequences in relationship to positively controlled promoters. Superimposed on these features was the less orthodox presence of multiple transcripts and their unique arrangement, including a region of complementarity at the 5' ends of the P2ahpC and P2oxyR mRNAs, suggesting the existence of complex regulatory relationships controlling oxyR and ahpC expression in mycobacteria. PMID:9079928

  10. Gene therapy: An overview

    Directory of Open Access Journals (Sweden)

    Sudip Indu

    2013-01-01

    Full Text Available Gene therapy "the use of genes as medicine" involves the transfer of a therapeutic or working copy of a gene into specific cells of an individual in order to repair a faulty gene copy. The technique may be used to replace a faulty gene, or to introduce a new gene whose function is to cure or to favorably modify the clinical course of a condition. The objective of gene therapy is to introduce new genetic material into target cells while causing no damage to the surrounding healthy cells and tissues, hence the treatment related morbidity is decreased. The delivery system includes a vector that delivers a therapeutic gene into the patient′s target cell. Functional proteins are created from the therapeutic gene causing the cell to return to a normal stage. The vectors used in gene therapy can be viral and non-viral. Gene therapy, an emerging field of biomedicine, is still at infancy and much research remains to be done before this approach to the treatment of condition will realize its full potential.

  11. Gene therapy in periodontics

    Directory of Open Access Journals (Sweden)

    Anirban Chatterjee

    2013-01-01

    Full Text Available GENES are made of DNA - the code of life. They are made up of two types of base pair from different number of hydrogen bonds AT, GC which can be turned into instruction. Everyone inherits genes from their parents and passes them on in turn to their children. Every person?s genes are different, and the changes in sequence determine the inherited differences between each of us. Some changes, usually in a single gene, may cause serious diseases. Gene therapy is ?the use of genes as medicine?. It involves the transfer of a therapeutic or working gene copy into specific cells of an individual in order to repair a faulty gene copy. Thus it may be used to replace a faulty gene, or to introduce a new gene whose function is to cure or to favorably modify the clinical course of a condition. It has a promising era in the field of periodontics. Gene therapy has been used as a mode of tissue engineering in periodontics. The tissue engineering approach reconstructs the natural target tissue by combining four elements namely: Scaffold, signaling molecules, cells and blood supply and thus can help in the reconstruction of damaged periodontium including cementum, gingival, periodontal ligament and bone.

  12. [Imprinted genes in plants].

    Science.gov (United States)

    Zhang, Li-Geng; Yang, Ruo-Fei; Fu, Feng-Ling; Li, Wan-Chen

    2010-12-01

    The expression of imprinted genes is regulated by epigenetic mechanism. In plant endosperm, the allele of imprinted genes is expressed in a pattern of parent-of-origin-dependent. The expression of imprinted genes plays essential roles in the development of embryos and their annexe structures, as well as seed size, reproductive barriers and apomixis. Along with the progress of plant epigenetic research, the exploration of imprinted genes is becoming hotspot in epigenetic research. This review focused on the parental conflict theory about the origin of imprinted genes, and the latest research advances in expression regulation mechanism of plant imprinted genes, using the examples of the important imprinted genes MEA, FIS2, FWA, MPC, and PHE1 in Arabidopsis, and FIEI and FIE2 in maize. PMID:21513148

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

    OpenAIRE

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

    2007-01-01

    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) rs247660...

  14. Essential Bacillus subtilis genes

    DEFF Research Database (Denmark)

    Kobayashi, K.; Ehrlich, S.D.

    2003-01-01

    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.

  15. Cancer gene therapy

    Directory of Open Access Journals (Sweden)

    Mitrovi? Tatjana

    2005-01-01

    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.

  16. Further understanding human disease genes by comparing with housekeeping genes and other genes

    Directory of Open Access Journals (Sweden)

    Chen Ting

    2006-02-01

    Full Text Available 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 genes as extremely severe "disease" genes. Here we perform a comparative study on the features of human essential, disease, and other genes. Results In the absence of a set of well defined human essential genes, we consider a set of 1,789 ubiquitously expressed human genes (UEHGs, also known as housekeeping genes, as an approximation. We demonstrate that UEHGs are very likely to contain a large proportion of essential genes. We show that the UEHGs, disease genes and other genes are different in their evolutionary conservation rates, DNA coding lengths, gene functions, etc. Our findings systematically confirm that disease genes have an intermediate essentiality which is less than housekeeping genes but greater than other human genes. Conclusion The human genome may contain thousands of essential genes having features which differ significantly from disease and other genes. We propose to classify them as a unique group for comparisons of disease genes with non-disease genes. This new way of classification and comparison enables us to have a clearer understanding of disease genes.

  17. Antisense gene silencing

    DEFF Research Database (Denmark)

    Nielsen, Troels T; Nielsen, Jørgen E

    2013-01-01

    Since the first reports that double-stranded RNAs can efficiently silence gene expression in C. elegans, the technology of RNA interference (RNAi) has been intensively exploited as an experimental tool to study gene function. With the subsequent discovery that RNAi could also be applied to mammalian cells, the technology of RNAi expanded from being a valuable experimental tool to being an applicable method for gene-specific therapeutic regulation, and much effort has been put into further refine...

  18. Green genes gleaned.

    Science.gov (United States)

    Beale, Samuel I

    2005-07-01

    A recent paper by Ayumi Tanaka and colleagues identifying an Arabidopsis thaliana gene for 3,8-divinyl(proto)chlorophyllide 8-vinyl reductase brings a satisfying conclusion to the hunt for genes encoding enzymes for the steps in the chlorophyll biosynthetic pathway. Now, at least in angiosperm plants represented by Arabidopsis, genes for all 15 steps in the pathway from glutamyl-tRNA to chlorophylls a and b have been identified. PMID:15951223

  19. Gene therapy for haemophilia

    OpenAIRE

    Murphy, Samuel L.; HIGH, KATHERINE A.

    2008-01-01

    The ultimate goal of gene therapy is the replacement of a defective gene sequence with a corrected version to eliminate disease for the lifetime of the patient. This challenging task is not yet accomplished, however significant progress is evident. An initial spate of clinical trials attempting the treatment of haemophilia with gene transfer primarily resulted in the demonstration of good safety profiles, but without efficacy. Subsequent reengineering of vector plasmids and delivery systems r...

  20. Genes, economics, and happiness

    OpenAIRE

    De Neve, Jan-Emmanuel; Fowler, James H; Frey, Bruno S.

    2010-01-01

    Research on happiness has produced valuable insights into the sources of subjective well-being. A major finding from this literature is that people exhibit a baseline happiness that shows persistent strength over time, and twin studies have shown that genes play a significant role in explaining the variance of baseline happiness between individuals. However, these studies have not identified which genes might be involved. This article presents evidence of a specific gene that predicts subject...

  1. Cancer gene therapy

    OpenAIRE

    Mitrovi? Tatjana; Radulovi? Siniša

    2005-01-01

    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 i...

  2. Genes and Social Behavior

    OpenAIRE

    Robinson, Gene E; Fernald, Russell D.; Clayton, David F.

    2008-01-01

    What specific genes and regulatory sequences contribute to the organization and functioning of brain circuits that support social behavior? How does social experience interact with information in the genome to modulate these brain circuits? Here we address these questions by highlighting progress that has been made in identifying and understanding two key “vectors of influence” that link genes, brain, and social behavior: 1) social information alters gene readout in the brain to influence beh...

  3. History of gene therapy.

    Science.gov (United States)

    Wirth, Thomas; Parker, Nigel; Ylä-Herttuala, Seppo

    2013-08-10

    Two decades after the initial gene therapy trials and more than 1700 approved clinical trials worldwide we not only have gained much new information and knowledge regarding gene therapy in general, but also learned to understand the concern that has persisted in society. Despite the setbacks gene therapy has faced, success stories have increasingly emerged. Examples for these are the positive recommendation for a gene therapy product (Glybera) by the EMA for approval in the European Union and the positive trials for the treatment of ADA deficiency, SCID-X1 and adrenoleukodystrophy. Nevertheless, our knowledge continues to grow and during the course of time more safety data has become available that helps us to develop better gene therapy approaches. Also, with the increased understanding of molecular medicine, we have been able to develop more specific and efficient gene transfer vectors which are now producing clinical results. In this review, we will take a historical view and highlight some of the milestones that had an important impact on the development of gene therapy. We will also discuss briefly the safety and ethical aspects of gene therapy and address some concerns that have been connected with gene therapy as an important therapeutic modality. PMID:23618815

  4. Gene : CBRC-MMUR-01-1271 [SEVENS

    Lifescience Database Archive (English)

    Full Text Available CBRC-MMUR-01-1271 Novel UN B UNKNOWN SRGEF_MOUSE 1.1 34% ref|YP_639653.1| precorrin-3B synthase ... [Mycobacterium sp. MCS ] ref|YP_938521.1| precorrin-3B synthase [Mycobacte ... G08597.1| Precorrin-3B synthase [Mycobacterium sp. MCS ] gb|ABL91731.1| precorrin-3B synthase [Mycobacteri ...

  5. Gene Conversion and Evolution of Gene Families: An Overview

    OpenAIRE

    Tomoko Ohta

    2010-01-01

    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.

  6. A Second Endolysin Gene Is Fully Embedded In-Frame with the lysA Gene of Mycobacteriophage Ms6

    OpenAIRE

    Catalão, Maria João; Milho, Catarina; Gil, Filipa; Moniz-Pereira, José; Pimentel, Madalena

    2011-01-01

    Mycobacteriophages are dsDNA viruses that infect mycobacterial hosts. The mycobacteriophage Ms6 accomplishes lysis by producing two cell wall hydrolytic enzymes, Lysin A (LysA) that possesses a central peptidoglycan recognition protein (PGRP) super-family conserved domain with the amidase catalytic site, that cleaves the amide bond between the N-acetylmuramic acid and L-alanine residues in the oligopeptide crosslinking chains of the peptidoglycan and Lysin B (LysB) a mycolylarabinogalactan es...

  7. Gene promoters dictate histone occupancy within genes.

    Science.gov (United States)

    Perales, Roberto; Erickson, Benjamin; Zhang, Lian; Kim, Hyunmin; Valiquett, Elan; Bentley, David

    2013-10-01

    Spt6 is a transcriptional elongation factor and histone chaperone that reassembles transcribed chromatin. Genome-wide H3 mapping showed that Spt6 preferentially maintains nucleosomes within the first 500 bases of genes and helps define nucleosome-depleted regions in 5' and 3' flanking sequences. In Spt6-depleted cells, H3 loss at 5' ends correlates with reduced pol II density suggesting enhanced transcription elongation. Consistent with its 'Suppressor of Ty' (Spt) phenotype, Spt6 inactivation caused localized H3 eviction over 1-2 nucleosomes at 5' ends of Ty elements. H3 displacement differed between genes driven by promoters with 'open'/DPN and 'closed'/OPN chromatin conformations with similar pol II densities. More eviction occurred on genes with 'closed' promoters, associated with 'noisy' transcription. Moreover, swapping of 'open' and 'closed' promoters showed that they can specify distinct downstream patterns of histone eviction/deposition. These observations suggest a novel function for promoters in dictating histone dynamics within genes possibly through effects on transcriptional bursting or elongation rate. PMID:24013117

  8. What Is a Gene?

    Science.gov (United States)

    ... kitchen to greet the guests. As he helps everyone get settled, Emma wonders, "Aunt Rita's grandmother? It runs in the family? What are they talking about?" Genes (say: jeenz ), that's what they're talking about. Genes play an important role in determining physical traits — how we look —and ...

  9. Smart Genes, Stupid Science.

    Science.gov (United States)

    Randerson, Sherman; Mahadeva, Madhu N.

    1983-01-01

    Because many people still believe that specific, identifiable genes dictate the level of human intelligence and that the number/quality of these genes can be evaluated, presents evidence from human genetics (related to nervous system development) to counter this view. Also disputes erroneous assumptions made in "heritability studies" of human…

  10. DNA repair genes

    International Nuclear Information System (INIS)

    Fission yeast S. pombe is assumed to be a good model for cloning of human DNA repair genes, because human gene is normally expressed in S. pombe and has a very similar protein sequence to yeast protein. We have tried to elucidate the DNA repair mechanisms of S. pombe as a model system for those of mammals. (J.P.N.)

  11. Ocular Gene Therapy.

    Science.gov (United States)

    Campbell, J Peter; McFarland, Trevor J; Stout, J Timothy

    2016-01-01

    Ocular gene therapy involves the introduction of an exogenous gene product to a host's cellular and genetic machinery for endogenous production of a desired gene product. The eye represents an ideal target organ due to its easy visibility and accessibility, and several trials have demonstrated proof-of-principle safety and efficacy in a subtype of Leber's congenital amaurosis. There are numerous ongoing clinical trials exploring gene therapy in other retinal diseases. In autosomal recessively inherited retinal degenerations, the introduced gene product replaces a known genetically deficient gene product and provides restoration of function. In other disease states, such as neovascular age-related macular degeneration, the delivered gene product modulates existing proteins within a cell, such as vascular endothelial growth factor, for a desired therapeutic effect. This latter approach may have broader applications in other diseases such as diabetes and other retinal vascular diseases that are as yet unrealized. This review summarizes the current state of clinical research in ocular gene therapy focusing on those diseases in which the technology has reached clinical trials. PMID:26502313

  12. Gene therapy for immunodeficiency.

    Science.gov (United States)

    Candotti, F

    2001-09-01

    Since the early 1990s, primary immunodeficiency (ID) disorders have played a major role in the development of human gene therapy. Adenosine deaminase (ADA) deficiency was the first disease to be treated with a gene therapy approach in humans, and was also the first condition for which therapeutic gene transfer into the hematopoietic stem cell has been attempted in the clinical arena. A series of encouraging results obtained in chronic granulomatous disease (CGD) patients have followed these pioneer experiments and preceded the very recent and exciting reports of successful genetic correction procedures performed in patients affected with the X-linked form of severe combined immunodeficiency (XSCID). The technical progress made in the field of gene transfer in recent years is mostly responsible for these clinical advances, and will be critical for future development of gene therapy approaches for other forms of IDs. PMID:11892066

  13. Library Generation by Gene Shuffling

    Science.gov (United States)

    Meyer, Adam J.; Ellefson, JaredW.; Ellington, Andrew D.

    2014-01-01

    This unit describes the process of gene shuffling (also known as sexual PCR). Gene shuffling is a facile method for the generation of sequence libraries containing the information from a family of related genes. Essentially, related genes are fragmented by DNase I digestion and reassembled by primerless PCR. The resulting chimeric genes can then be screened or selected for a desired function. PMID:24510437

  14. A review on microcephaly genes

    OpenAIRE

    Irshad S.; Shahid S.

    2012-01-01

    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.

  15. Adenovirus Vectors for Gene Therapy, Vaccination and Cancer Gene Therapy

    OpenAIRE

    Wold, William S.M.; Toth, Karoly

    2013-01-01

    Adenovirus vectors are the most commonly employed vector for cancer gene therapy. They are also used for gene therapy and as vaccines to express foreign antigens. Adenovirus vectors can be replication-defective; certain essential viral genes are deleted and replaced by a cassette that expresses a foreign therapeutic gene. Such vectors are used for gene therapy, as vaccines, and for cancer therapy. Replication-competent (oncolytic) vectors are employed for cancer gene therapy. Oncolytic vector...

  16. GeneCards Version 3: the human gene integrator

    OpenAIRE

    2010-01-01

    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 ...

  17. Gene amplification in carcinogenesis

    Scientific Electronic Library Online (English)

    Lucimari, Bizari; Ana Elizabete, Silva; Eloiza H., Tajara.

    Full Text Available 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.

  18. Gene Variants Reduce Opioid Risks

    Science.gov (United States)

    ... Notes » Basic Science » Gene Variants Reduce Opioid Risks Gene Variants Reduce Opioid Risks Email Facebook Twitter June ... One study associated a rare variant of the gene for the μ-opioid receptor (OPRM1) with a ...

  19. 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.)

  20. Genome-wide transcriptional profiling of peripheral blood leukocytes from cattle infected with Mycobacterium bovis reveals suppression of host immune genes

    Directory of Open Access Journals (Sweden)

    Killick Kate E

    2011-12-01

    Full Text Available Abstract Background Mycobacterium bovis is the causative agent of bovine tuberculosis (BTB, a pathological infection with significant economic impact. Recent studies have highlighted the role of functional genomics to better understand the molecular mechanisms governing the host immune response to M. bovis infection. Furthermore, these studies may enable the identification of novel transcriptional markers of BTB that can augment current diagnostic tests and surveillance programmes. In the present study, we have analysed the transcriptome of peripheral blood leukocytes (PBL from eight M. bovis-infected and eight control non-infected age-matched and sex-matched Holstein-Friesian cattle using the Affymetrix® GeneChip® Bovine Genome Array with 24,072 gene probe sets representing more than 23,000 gene transcripts. Results Control and infected animals had similar mean white blood cell counts. However, the mean number of lymphocytes was significantly increased in the infected group relative to the control group (P = 0.001, while the mean number of monocytes was significantly decreased in the BTB group (P = 0.002. Hierarchical clustering analysis using gene expression data from all 5,388 detectable mRNA transcripts unambiguously partitioned the animals according to their disease status. In total, 2,960 gene transcripts were differentially expressed (DE between the infected and control animal groups (adjusted P-value threshold ? 0.05; with the number of gene transcripts showing decreased relative expression (1,563 exceeding those displaying increased relative expression (1,397. Systems analysis using the Ingenuity® Systems Pathway Analysis (IPA Knowledge Base revealed an over-representation of DE genes involved in the immune response functional category. More specifically, 64.5% of genes in the affects immune response subcategory displayed decreased relative expression levels in the infected animals compared to the control group. Conclusions This study demonstrates that genome-wide transcriptional profiling of PBL can distinguish active M. bovis-infected animals from control non-infected animals. Furthermore, the results obtained support previous investigations demonstrating that mycobacterial infection is associated with host transcriptional suppression. These data support the use of transcriptomic technologies to enable the identification of robust, reliable transcriptional markers of active M. bovis infection.

  1. 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)

  2. Antisense gene silencing

    DEFF Research Database (Denmark)

    Nielsen, Troels T; Nielsen, Jørgen E

    2013-01-01

    Since the first reports that double-stranded RNAs can efficiently silence gene expression in C. elegans, the technology of RNA interference (RNAi) has been intensively exploited as an experimental tool to study gene function. With the subsequent discovery that RNAi could also be applied to...... mammalian cells, the technology of RNAi expanded from being a valuable experimental tool to being an applicable method for gene-specific therapeutic regulation, and much effort has been put into further refinement of the technique. This review will focus on how RNAi has developed over the years and how the...

  3. Gene promoters dictate histone occupancy within genes

    OpenAIRE

    Perales, Roberto; Erickson, Benjamin; Zhang, Lian; Kim, Hyunmin; Valiquett, Elan; Bentley, David

    2013-01-01

    Spt6 is a transcriptional elongation factor and histone chaperone that reassembles transcribed chromatin. Genome-wide H3 mapping showed that Spt6 preferentially maintains nucleosomes within the first 500 bases of genes and helps define nucleosome-depleted regions in 5? and 3? flanking sequences. In Spt6-depleted cells, H3 loss at 5? ends correlates with reduced pol II density suggesting enhanced transcription elongation. Consistent with its ‘Suppressor of Ty' (Spt) phenotype, Spt6 inactivatio...

  4. The gene tree delusion.

    Science.gov (United States)

    Springer, Mark S; Gatesy, John

    2016-01-01

    Higher-level relationships among placental mammals are mostly resolved, but several polytomies remain contentious. Song et al. (2012) claimed to have resolved three of these using shortcut coalescence methods (MP-EST, STAR) and further concluded that these methods, which assume no within-locus recombination, are required to unravel deep-level phylogenetic problems that have stymied concatenation. Here, we reanalyze Song et al.'s (2012) data and leverage these re-analyses to explore key issues in systematics including the recombination ratchet, gene tree stoichiometry, the proportion of gene tree incongruence that results from deep coalescence versus other factors, and simulations that compare the performance of coalescence and concatenation methods in species tree estimation. Song et al. (2012) reported an average locus length of 3.1kb for the 447 protein-coding genes in their phylogenomic dataset, but the true mean length of these loci (start codon to stop codon) is 139.6kb. Empirical estimates of recombination breakpoints in primates, coupled with consideration of the recombination ratchet, suggest that individual coalescence genes (c-genes) approach ?12bp or less for Song et al.'s (2012) dataset, three to four orders of magnitude shorter than the c-genes reported by these authors. This result has general implications for the application of coalescence methods in species tree estimation. We contend that it is illogical to apply coalescence methods to complete protein-coding sequences. Such analyses amalgamate c-genes with different evolutionary histories (i.e., exons separated by >100,000bp), distort true gene tree stoichiometry that is required for accurate species tree inference, and contradict the central rationale for applying coalescence methods to difficult phylogenetic problems. In addition, Song et al.'s (2012) dataset of 447 genes includes 21 loci with switched taxonomic names, eight duplicated loci, 26 loci with non-homologous sequences that are grossly misaligned, and numerous loci with >50% missing data for taxa that are misplaced in their gene trees. These problems were compounded by inadequate tree searches with nearest neighbor interchange branch swapping and inadvertent application of substitution models that did not account for among-site rate heterogeneity. Sixty-six gene trees imply unrealistic deep coalescences that exceed 100 million years (MY). Gene trees that were obtained with better justified models and search parameters show large increases in both likelihood scores and congruence. Coalescence analyses based on a curated set of 413 improved gene trees and a superior coalescence method (ASTRAL) support a Scandentia (treeshrews)+Glires (rabbits, rodents) clade, contradicting one of the three primary systematic conclusions of Song et al. (2012). Robust support for a Perissodactyla+Carnivora clade within Laurasiatheria is also lost, contradicting a second major conclusion of this study. Song et al.'s (2012) MP-EST species tree provided the basis for circular simulations that led these authors to conclude that the multispecies coalescent accounts for 77% of the gene tree conflicts in their dataset, but many internal branches of their MP-EST tree are stunted by an order of magnitude or more due to wholesale gene tree reconstruction errors. An independent assessment of branch lengths suggests the multispecies coalescent accounts for ?15% of the conflicts among Song et al.'s (2012) 447 gene trees. Unfortunately, Song et al.'s (2012) flawed phylogenomic dataset has been used as a model for additional simulation work that suggests the superiority of shortcut coalescence methods relative to concatenation. Investigator error was passed on to the subsequent simulation studies, which also incorporated further logical errors that should be avoided in future simulation studies. Illegitimate branch length switches in the simulation routines unfairly protected coalescence methods from their Achilles' heel, high gene tree reconstruction error at short internodes. These simulations therefore provide no

  5. Gene Therapy of Cancerous Diseases

    OpenAIRE

    Valen?áková, A.; Dziaková, A.; Hatalová, E.

    2015-01-01

    Gene therapy of cancerous diseases provides new means of curing patients with oncologic illnesses. There are several approaches in treating cancer by gene therapy. Most commonly used methods are: cancer immunogene therapy, suicide gene therapy, application of tumor-suppressor genes, antiangiogenic therapy, mesenchymal stem cells used as vectors, gene directed enzyme/prodrug therapy and bacteria used as anti-cancer agents. Cancer gene immunotherapy uses several immunologic agents for the purp...

  6. "Bad genes" & criminal responsibility.

    Science.gov (United States)

    González-Tapia, María Isabel; Obsuth, Ingrid

    2015-01-01

    The genetics of the accused is trying to break into the courts. To date several candidate genes have been put forward and their links to antisocial behavior have been examined and documented with some consistency. In this paper, we focus on the so called "warrior gene", or the low-activity allele of the MAOA gene, which has been most consistently related to human behavior and specifically to violence and antisocial behavior. In preparing this paper we had two objectives. First, to summarize and analyze the current scientific evidence, in order to gain an in depth understanding of the state of the issue and determine whether a dominant line of generally accepted scientific knowledge in this field can be asserted. Second, to derive conclusions and put forward recommendations related to the use of genetic information, specifically the presence of the low-activity genotype of the MAOA gene, in modulation of criminal responsibility in European and US courts. PMID:25708001

  7. Imaging gene expression

    International Nuclear Information System (INIS)

    The rapid progress of molecular genetic methods over the past two decades has necessitated the development of methods to detect and quantify genetic activity within living bodies. Reporter genes provide a rapid and convenient tool to monitor gene expression by yielding a readily measurable phenotype upon expression when introduced into a biological system. Conventional reporter systems, however, are limited in their usefulness for in vivo experimjents or human gene therapy because of its invasive nature which requires cell damage before assays can be performed. This offers an unique opportunity for nuclear imaging techniques to develope a novel method for imaging both the location and amount of gene expression noninvasively. Current developments to achieve this goal rely on utilizing either reporter enzymes that accumulate radiolabeled substrates or reporter receptors that bind specific radioligands. This overview includes a brief introduction to the background for such research, a summary of published fresults, and an outlook for future directions

  8. Gene expression in fungi

    OpenAIRE

    Kalkanci, Ayse; Kadioglu, Aras; Wilson, Duncan; Jacobsen, Mette D.

    2011-01-01

    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 evaluate...

  9. Gene therapy in ophthalmology

    OpenAIRE

    Uthra Satagopan; Kumaramanickavel Govindasamy

    2009-01-01

    It has been more than a year since ophthalmologists and scientists under Dr. Robin Ali?s team at the Moorsfield Eye Hospital and the Institute of Ophthalmology, University College London, successfully treated patients with a severely blinding disease, Leber?s congenital amaurosis (LCA) using gene therapy. This success does not look to be transient, and this achievement in gene replacement therapy clinical trial for LCA has instilled hope in numerous families with patients suffer...

  10. Finding Genes for Schizophrenia

    OpenAIRE

    Åberg, Karolina

    2005-01-01

    Schizophrenia is one of our most common psychiatric diseases. It severely affects all aspects of psychological functions and results in loss of contact with reality. No cure exists and the treatments available today produce only partial relief for disease symptoms. The aim of this work is to better understand the etiology of schizophrenia by identification of candidate genes and gene pathways involved in the development of the disease. In a preliminarily study, the effects of medication and g...

  11. GEIRA: gene-environment and gene-gene interaction research application

    OpenAIRE

    Ding, Bo; Källberg, Henrik; Klareskog, Lars; Padyukov, Leonid; Alfredsson, Lars

    2011-01-01

    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 coded i...

  12. Radiosensitivity and genes

    International Nuclear Information System (INIS)

    Reported effects of some oncogenes, tumour suppressor genes and DNA repair genes on sensitivity of cells to ionizing radiation are reviewed. The role of oncogenes in cellular response to irradiation is discussed, especially the extensively studied oncogenes such as the ras gene family. For tumour suppressor genes, mainly the p53, which is increasingly implicated as a gene affecting radiosensitivity, is reviewed. It is considered that there is a cell cycle checkpoint determinant which is postulated to be able to arrest the irradiated cells in G1 phase to allow them to repair damage before they undergo DNA synthesis. So far there are six DNA repair genes which have been cloned in mammalian cells, but only one, XRCC1, appears to be involved in repair of human X-ray damage. XRCC1 can correct high sisterchromatid exchange levels when transferred into EM9 cells, but its expression seems to have no correlation with radiosensitivity of human neck and head tumour cells. Radiosensitivity is an intricate issue which may involve many factors. A scheme of cellular reactions after exposure to irradiation is proposed to indicate a possible sequence of events initiated by ionizing radiation

  13. Evidence for homosexuality gene

    Energy Technology Data Exchange (ETDEWEB)

    Pool, R.

    1993-07-16

    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.

  14. The Mycoplasma hominis vaa gene displays a mosaic gene structure

    DEFF Research Database (Denmark)

    Boesen, Thomas; Emmersen, Jeppe M. G.; Jensen, Lise T.; Ladefoged, Søren; Thorsen, Poul; Birkelund, Svend; Christiansen, Gunna; Jensen, Lise Torp

    1998-01-01

    Mycoplasma hominis contains a variable adherence-associated (vaa) gene. To classify variants of the vaa genes, we examined 42 M. hominis isolated by PCR, DNA sequencing and immunoblotting. This uncovered the existence of five gene categories. Comparison of the gene types revealed a modular...

  15. Identification of four soybean reference genes for gene expression normalization

    Science.gov (United States)

    Gene expression analysis requires the use of reference genes stably expressed independently of specific tissues or environmental conditions. Housekeeping genes (e.g., actin, tubulin, ribosomal, polyubiquitin and elongation factor 1-alpha) are commonly used as reference genes with the assumption tha...

  16. Mycobacterium anyangense sp. nov., a rapidly growing species isolated from blood of Korean native cattle, Hanwoo (Bos taurus coreanae).

    Science.gov (United States)

    Kim, Byoung-Jun; Kim, Jae-Myung; Kim, Bo-Ram; Lee, So-Young; Kim, GaNa; Jang, Yun-Ho; Ryoo, Soyoon; Jeon, Che-Ok; Jin, Hyun-Mi; Jeong, Joseph; Lee, Seon Ho; Lim, Ji-Hun; Kook, Yoon-Hoh; Kim, Bum-Joon

    2015-07-01

    From the whole blood of Korean native cattle, Hanwoo (Bos taurus coreanae), a previously undescribed, rapidly growing, scotochromogenic isolate of the genus Mycobacterium is reported. Its 16S rRNA gene sequence, and the sequences of three other genes (hsp65, recA and rpoB) were unique and phylogenetic analysis based on 16S rRNA gene sequence (1420 bp) placed the organism into the rapidly growing Mycobacterium group close to Mycobacterium smegmatis (98.5% sequence similarity). However, phylogenetic analyses based on three different gene sequences (hsp65, recA and rpoB) revealed its location to be distinct from the branch of rapidly growing species. Culture and biochemical characteristics were generally similar to those of Mycobacterium fortuitum. Unique matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) MS profiles of lipids, unique fatty acid profile, unique mycolic acids profiles and a low DNA-DNA relatedness to M. fortuitum (23.6%) and M. smegmatis (39.7%) strongly supported the taxonomic status of this strain as a representative of a novel species of rapidly growing mycobacteria named Mycobacterium anyangense. The type strain is strain QIA-38(T) (?= JCM 30275(T) = KCTC 29443(T)). PMID:25870258

  17. Applications of nanoparticle systems in gene delivery and gene therapy.

    Science.gov (United States)

    Kafshdooz, Taiebeh; Kafshdooz, Leila; Akbarzadeh, Abolfazl; Hanifehpour, Younes; Joo, Sang Woo

    2016-03-01

    For successful gene therapy, expansion of appropriate gene delivery systems could be one of the factors of major significance. Gene therapy provides large opportunities for treating diseases, including genetic disorders, infections, and cancer. Polymeric carriers have relatively low cytotoxicity and immunogenicity. Polymeric gene carriers are a potential substitute to using viral vectors. Overall, polymeric carriers can contain large-sized DNA, be conjugated with suitable functionalities, and be administered frequently. However, polymeric gene carriers have some restrictions, such as low gene transfection efficiencies and a moderately short period of gene expression. This study explores the current status of development of polymeric gene carriers, and presents guidelines for the prospective use of the polymer-based gene delivery systems in gene therapy. PMID:25365242

  18. Classification of genes based on gene expression analysis

    International Nuclear Information System (INIS)

    Systems biology and bioinformatics are now major fields for productive research. DNA microarrays and other array technologies and genome sequencing have advanced to the point that it is now possible to monitor gene expression on a genomic scale. Gene expression analysis is discussed and some important clustering techniques are considered. The patterns identified in the data suggest similarities in the gene behavior, which provides useful information for the gene functionalities. We discuss measures for investigating the homogeneity of gene expression data in order to optimize the clustering process. We contribute to the knowledge of functional roles and regulation of E. coli genes by proposing a classification of these genes based on consistently correlated genes in expression data and similarities of gene expression patterns. A new visualization tool for targeted projection pursuit and dimensionality reduction of gene expression data is demonstrated.

  19. Características clínicas, factores de riesgo y perfil de susceptibilidad de las infecciones por micobacterias documentadas por cultivo, en un hospital universitario de alta complejidad en Medellín (Colombia) / Clinical features, risk factors and susceptibility profile of mycobacterial infections documented by culture in a university hospital of high complexity in Medellin (Colombia)

    Scientific Electronic Library Online (English)

    Franco E, Montufar Andrade; Carolina, Aguilar Londoño; Carolina, Saldarriaga Acevedo; Alicia, Quiroga Echeverri; Carlos E, Builes Montaño; Miguel A, Mesa Navas; Olga L, Molina Upegüi; John J, Zuleta Tobón.

    2014-12-01

    Full Text Available Introducción: Tuberculosis (TBC) es aún una entidad de alta prevalencia y mortalidad en el mundo. La resistencia ascendente a fármacos es un problema de salud pública. Además se describen con mayor frecuencia infecciones por micobacterias no tuberculosas (MNT) en áreas de alta prevalencia de TBC. Ob [...] jetivos: Determinar características epidemiológicas, clínicas y microbiológicas de las infecciones por micobacterias documentadas por cultivo. Materiales y Métodos: Estudio observacional, descriptivo, en pacientes hospitalizados. Resultados: De 187 pacientes, en 90,9% se identificó complejo M. tuberculosis y en 9,1% MNT; 64% fueron hombres. Edad promedio 40 años (rango 1-88 años). Las principales co-morbilidades fueron infección por VIH/SIDA (23,5%), uso de corticoesteroides (13,3%) y enfermedad renal crónica (9,6%). Las formas clínicas fueron pulmonares (56,6%), extra-pulmonares (23,9%) y diseminadas (19,2%). El compromiso extra-pulmonar más frecuente fue ganglionar (7,4%) y gastrointestinal (7%). En M. tuberculosis 10,6% fueron multidrogoresistentes (MDR) y 2,12% con resistencia extendida (XDR). Mycobacterium avium y M. abscessus fueron las MNT más frecuentes. La mortalidad general fue 10%. Conclusiones: Inmuno-supresión es el principal factor de riesgo para enfermedad extrapulmonar y/o diseminada y la resistencia a fármacos en pacientes hospitalizados con TBC es llamativa, con mayor incidencia de MDR y XDR. Las infecciones por MNT no son infrecuentes en nuestro medio. Abstract in english Introduction: Tuberculosis (TB) remains an entity of high prevalence and mortality worldwide. The rising drug resistance is a public health problem. Besides, non-tuberculosis mycobacterial (NTM) infections are described with increasing frequency in areas of high prevalence of TB. Objectives: To dete [...] rmine epidemiological, clinical and microbiological characteristics of mycobacterial infections documented by culture. Materials and Methods: An observational, descriptive study in hospitalized patients. Results: M. tuberculosis complex was identified in 90,9% of 187 patients; 9,1% had NTM, 64% were male and the mean age was 40 years (range 1-88 years). The main co-morbidities were HIV / AIDS (23.5%), use of corticosteroids (13.3%) and chronic kidney disease (9.6%). Clinical forms were pulmonary (56.6%), extra-pulmonary (23.9%) and disseminated (19.2 The most common extra-pulmonary compromise was nodal (7.4%) and gastrointestinal (7%). 10.6% of M. tuberculosis were multi-drugresistant (MDR) and 2.12% had extended drug resistance (XDR). Mycobacterium avium andM. abscessus were the most frequent NTM. Overall mortality was 10%. Conclusions: In our study immune suppression is the main risk factor for extrapulmonary and disseminated disease. Resistance, MDR and XDR is higher in inpatients with TB. MNT infections are not uncommon in our country.

  20. Gene Circuit Analysis of the Terminal Gap Gene huckebein

    OpenAIRE

    Ashyraliyev, Maksat; Siggens, Ken; Janssens, Hilde; Blom, Joke; Akam, Michael; Jaeger, Johannes

    2009-01-01

    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...

  1. GeneCards Version 3: the human gene integrator.

    Science.gov (United States)

    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

    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

  2. [Gene therapy of hereditary diseases].

    Science.gov (United States)

    Ginter, E K

    2000-01-01

    In the review the main advantages in development of the approaches to gene therapy of hereditary diseases are presented. Now more than 1000 genes of hereditary diseases are mapped and some hundreds are cloned which is prerequisite for gene therapy. The transfer of the recombinant gene into the cell and the subsequent expression of the transgene product are the rate-limiting steps for successful gene therapy. A variety of methods, including the use of physical methods, modified viruses and synthetic vectors, are currently being used in experiments and clinical trials. Since the approval and initiation of the first human gene therapy trial to treat ADA deficiency, there have been several dozen approved gene therapy trials but clear clinical result was stated for ADA deficiency only. Cystic Fibrosis, CF was among several hereditary diseases which were considered as a target for gene therapy. Experiments on development of recombinant gene constructions, gene delivery by adenovirus vectors and liposomes as well as by other constructions into epithelial lung cells, gene expression and on the safety of gene therapy procedures were relatively successful. Phase 1 gene therapy clinical trials of CF showed that some unaccounted physiological peculiarities of lung tissue of the patients diminished effectiveness of gene transfer, longevity of CFTR gene expression and in some cases unexpected immunological complications arises during clinical trials. Now an intensive attempt to overcome these problems in gene therapy of CF are undertaken. PMID:11033886

  3. FunGene: the Functional Gene Pipeline and Repository

    Directory of Open Access Journals (Sweden)

    JamesR.Cole

    2013-10-01

    Full Text Available Ribosomal RNA genes have become the standard molecular markers for microbial community analysis for good reasons, including universal occurrence in cellular organisms, availability of large databases, and ease of rRNA gene region amplification and analysis. As markers, however, rRNA genes have some significant limitations. The rRNA genes are often present in multiple copies, unlike most protein-coding genes. The slow rate of change in rRNA genes means that multiple species sometimes share identical 16S rRNA gene sequences, while many more species share identical sequences in the short 16S rRNA regions commonly analyzed. In addition, the genes involved in many important processes are not distributed in a phylogenetically coherent manner, potentially due to gene loss or horizontal gene transfer. While rRNA genes remain the most commonly used markers, key genes in ecologically important pathways, e.g., those involved in carbon and nitrogen cycling, can provide important insights into community composition and function not obtainable through rRNA analysis. However, working with ecofunctional gene data requires some tools beyond those required for rRNA analysis. To address this, our Functional Gene Pipeline and Repository (FunGene; http://fungene.cme.msu.edu/ offers databases of many common ecofunctional genes and proteins, as well as integrated tools that allow researchers to browse these collections and choose subsets for further analysis, build phylogenetic trees, test primers and probes for coverage, and download aligned sequences. Additional FunGene tools are specialized to process coding gene amplicon data. For example, FrameBot produces frameshift-corrected protein and DNA sequences from raw reads while finding the most closely related protein reference sequence. These tools can help provide better insight into microbial communities by directly studying key genes involved in important ecological processes.

  4. Radionuclide reporter gene imaging for cardiac gene therapy

    International Nuclear Information System (INIS)

    In the field of cardiac gene therapy, angiogenic gene therapy has been most extensively investigated. The first clinical trial of cardiac angiogenic gene therapy was reported in 1998, and at the peak, more than 20 clinical trial protocols were under evaluation. However, most trials have ceased owing to the lack of decisive proof of therapeutic effects and the potential risks of viral vectors. In order to further advance cardiac angiogenic gene therapy, remaining open issues need to be resolved: there needs to be improvement of gene transfer methods, regulation of gene expression, development of much safer vectors and optimisation of therapeutic genes. For these purposes, imaging of gene expression in living organisms is of great importance. In radionuclide reporter gene imaging, ''reporter genes'' transferred into cell nuclei encode for a protein that retains a complementary ''reporter probe'' of a positron or single-photon emitter; thus expression of the reporter genes can be imaged with positron emission tomography or single-photon emission computed tomography. Accordingly, in the setting of gene therapy, the location, magnitude and duration of the therapeutic gene co-expression with the reporter genes can be monitored non-invasively. In the near future, gene therapy may evolve into combination therapy with stem/progenitor cell transplantation, so-called cell-based gene therapy or gene-modified cell therapy. Radionuclide reporter gene imaging is now expected to contribute in providing evidence on the usefulness of this novel therapeutic approach, as well as in investigating the molecular mechanisms underlying neovascularisation and safety issues relevant to further progress in conventional gene therapy. (orig.)

  5. Genes: an Open Access Journal

    OpenAIRE

    Young, J Peter W

    2009-01-01

    Genes have been in the scientific vocabulary for a hundred years. The term "gene" was proposed by the Danish plant scientist Wilhelm Johannsen in the first decade of the 20th century. For Johannsen, the gene remained an abstract concept, "free of any hypothesis" [1], but others were already pointing to chromosomes as the likely location of genes. The science of genetics was born at that time, and genes were rapidly connected with mutations, with patterns of inheritance, with development, with...

  6. Vertebrate gene predictions and the problem of large genes

    DEFF Research Database (Denmark)

    Wang, Jun; Li, ShengTing; Zhang, Yong; Zheng, HongKun; Xu, Zhao; Ye, Jia; Yu, Jun; Wong, Gane Ka-Shu

    2003-01-01

    To find unknown protein-coding genes, annotation pipelines use a combination of ab initio gene prediction and similarity to experimentally confirmed genes or proteins. Here, we show that although the ab initio predictions have an intrinsically high false-positive rate, they also have a consistently...... low false-negative rate. The incorporation of similarity information is meant to reduce the false-positive rate, but in doing so it increases the false-negative rate. The crucial variable is gene size (including introns)--genes of the most extreme sizes, especially very large genes, are most likely to...

  7. Gene therapy: progress and predictions

    Science.gov (United States)

    Collins, Mary; Thrasher, Adrian

    2015-01-01

    The first clinical gene delivery, which involved insertion of a marker gene into lymphocytes from cancer patients, was published 25 years ago. In this review, we describe progress since then in gene therapy. Patients with some inherited single-gene defects can now be treated with their own bone marrow stem cells that have been engineered with a viral vector carrying the missing gene. Patients with inherited retinopathies and haemophilia B can also be treated by local or systemic injection of viral vectors. There are also a number of promising gene therapy approaches for cancer and infectious disease. We predict that the next 25 years will see improvements in safety, efficacy and manufacture of gene delivery vectors and introduction of gene-editing technologies to the clinic. Gene delivery may also prove a cost-effective method for the delivery of biological medicines. PMID:26702034

  8. Gene therapy: progress and predictions.

    Science.gov (United States)

    Collins, Mary; Thrasher, Adrian

    2015-12-22

    The first clinical gene delivery, which involved insertion of a marker gene into lymphocytes from cancer patients, was published 25 years ago. In this review, we describe progress since then in gene therapy. Patients with some inherited single-gene defects can now be treated with their own bone marrow stem cells that have been engineered with a viral vector carrying the missing gene. Patients with inherited retinopathies and haemophilia B can also be treated by local or systemic injection of viral vectors. There are also a number of promising gene therapy approaches for cancer and infectious disease. We predict that the next 25 years will see improvements in safety, efficacy and manufacture of gene delivery vectors and introduction of gene-editing technologies to the clinic. Gene delivery may also prove a cost-effective method for the delivery of biological medicines. PMID:26702034

  9. Suicide genes or p53 gene and p53 target genes as targets for cancer gene therapy by ionizing radiation

    International Nuclear Information System (INIS)

    Radiotherapy has some disadvantages due to the severe side-effect on the normal tissues at a curative dose of ionizing radiation (IR). Similarly, as a new developing approach, gene therapy also has some disadvantages, such as lack of specificity for tumors, limited expression of therapeutic gene, potential biological risk. To certain extent, above problems would be solved by the suicide genes or p53 gene and its target genes therapies targeted by ionizing radiation. This strategy not only makes up the disadvantage from radiotherapy or gene therapy alone, but also promotes success rate on the base of lower dose. By present, there have been several vectors measuring up to be reaching clinical trials. This review focused on the development of the cancer gene therapy through suicide genes or p53 and its target genes mediated by IR. (authors)

  10. Multidimensional gene search with Genehopper.

    Science.gov (United States)

    Munz, Matthias; Tönnies, Sascha; Balke, Wolf-Tilo; Simon, Eric

    2015-07-01

    The high abundance of genetic information enables researchers to gain new insights from the comparison of human genes according to their similarities. However, existing tools that allow the exploration of such gene-to-gene relationships, apply each similarity independently. To make use of multidimensional scoring, we developed a new search engine named Genehopper. It can handle two query types: (i) the typical use case starts with a term-to-gene search, i.e. an optimized full-text search for an anchor gene of interest. The web-interface can handle one or more terms including gene symbols and identifiers of Ensembl, UniProt, EntrezGene and RefSeq. (ii) When the anchor gene is defined, the user can explore its neighborhood by a gene-to-gene search as the weighted sum of nine normalized gene similarities based on sequence homology, protein domains, mRNA expression profiles, Gene Ontology Annotation, gene symbols and other features. Each weight can be adjusted by the user, allowing flexible customization of the gene search. All implemented similarities have a low pairwise correlation (max r(2) = 0.4) implying a low linear dependency, i.e. any change in a single weight has an effect on the ranking. Thus, we treated them as separate dimensions in the search space. Genehopper is freely available at http://genehopper.ifis.cs.tu-bs.de. PMID:25990726

  11. Genes2FANs: connecting genes through functional association networks

    Directory of Open Access Journals (Sweden)

    Dannenfelser Ruth

    2012-07-01

    Full Text Available Abstract Background Protein-protein, cell signaling, metabolic, and transcriptional interaction networks are useful for identifying connections between lists of experimentally identified genes/proteins. However, besides physical or co-expression interactions there are many ways in which pairs of genes, or their protein products, can be associated. By systematically incorporating knowledge on shared properties of genes from diverse sources to build functional association networks (FANs, researchers may be able to identify additional functional interactions between groups of genes that are not readily apparent. Results Genes2FANs is a web based tool and a database that utilizes 14 carefully constructed FANs and a large-scale protein-protein interaction (PPI network to build subnetworks that connect lists of human and mouse genes. The FANs are created from mammalian gene set libraries where mouse genes are converted to their human orthologs. The tool takes as input a list of human or mouse Entrez gene symbols to produce a subnetwork and a ranked list of intermediate genes that are used to connect the query input list. In addition, users can enter any PubMed search term and then the system automatically converts the returned results to gene lists using GeneRIF. This gene list is then used as input to generate a subnetwork from the user’s PubMed query. As a case study, we applied Genes2FANs to connect disease genes from 90 well-studied disorders. We find an inverse correlation between the counts of links connecting disease genes through PPI and links connecting diseases genes through FANs, separating diseases into two categories. Conclusions Genes2FANs is a useful tool for interpreting the relationships between gene/protein lists in the context of their various functions and networks. Combining functional association interactions with physical PPIs can be useful for revealing new biology and help form hypotheses for further experimentation. Our finding that disease genes in many cancers are mostly connected through PPIs whereas other complex diseases, such as autism and type-2 diabetes, are mostly connected through FANs without PPIs, can guide better strategies for disease gene discovery. Genes2FANs is available at: http://actin.pharm.mssm.edu/genes2FANs.

  12. Radio-induced genes

    International Nuclear Information System (INIS)

    The monitoring system of the DNA integrity of an irradiated cell does not satisfy oneself to recruit the enzymes allowing the repair of detected damages. It sends an alarm signal whom transmission leads to the activation of specific genes in charge of stopping the cell cycle, the time to make the repair works, or to lead to the elimination of a too much damaged cell. Among the numerous genes participating to the monitoring of cell response to irradiation, the target genes of the mammalian P53 protein are particularly studied. Caretaker of the genome, this protein play a central part in the cell response to ionizing radiations. this response is less studied among plants. A way to tackle it is to be interested in the radioinduced genes identification in the vegetal cell, while taking advantage of knowledge got in the animal field. The knowledge of the complete genome of the arabette (arabidopsis thaliana), the model plant and the arising of new techniques allow to lead this research at a previously unknown rhythm in vegetal biology. (N.C.)

  13. Genes in mammalian reproduction

    Energy Technology Data Exchange (ETDEWEB)

    Gwatkin, R.B.L. [ed.

    1996-11-01

    This is an informative book which deals mainly with genomic imprinting, the role of steroid hormones in development, the expression of a variety of genes during development and the link to hereditary diseases. It is an up-to-date review in a field that is quickly changing and provides valuable basic information and current research trends.

  14. Genes and Vocal Learning

    Science.gov (United States)

    White, Stephanie A.

    2010-01-01

    Could a mutation in a single gene be the evolutionary lynchpin supporting the development of human language? A rare mutation in the molecule known as FOXP2 discovered in a human family seemed to suggest so, and its sequence phylogeny reinforced a Chomskian view that language emerged wholesale in humans. Spurred by this discovery, research in…

  15. Searching for speciation genes

    DEFF Research Database (Denmark)

    Holt, Benjamin George; Côté, Isabelle M; Emerson, Brent C

    2011-01-01

    Closely related species that show clear phenotypic divergence, but without obvious geographic barriers, can provide opportunities to study how diversification can occur when opportunities for allopatric speciation are limited. We examined genetic divergence in the coral reef fish genus Hypoplectr...... evidence for genes that may be associated with colour morphotype in the genus Hypoplectrus....

  16. IBMFS - gene mutations

    Science.gov (United States)

    A "mutation" is a change in a gene that prevents it from working properly. A "germline" mutation is a change that occurs in the egg or the sperm, or both, and is passed from one parent or both parents to the child.

  17. Gene therapy in ophthalmology

    Directory of Open Access Journals (Sweden)

    Uthra Satagopan

    2009-01-01

    Full Text Available It has been more than a year since ophthalmologists and scientists under Dr. Robin Ali?s team at the Moorsfield Eye Hospital and the Institute of Ophthalmology, University College London, successfully treated patients with a severely blinding disease, Leber?s congenital amaurosis (LCA using gene therapy. This success does not look to be transient, and this achievement in gene replacement therapy clinical trial for LCA has instilled hope in numerous families with patients suffering from this and similar retinal degenerative diseases, for whom restoration of lost vision has remained a distant dream so far. The encouragement that this success has given is expected to also lead to start of clinical trials for other blinding ocular diseases for which gene therapy experiments at the laboratory and animal levels have been successful. This article reviews the various studies that have led to the understanding of gene therapy outcomes in human ocular diseases and attempts to provide a brief sketch of successful clinical trials.

  18. Naming genes beyond Caenorhabditis

    Science.gov (United States)

    The nomenclature of genes in Caenorhabditis elegans is based on long-standing, successful guidelines established in the late 1970s. Over time these guidelines have matured into a comprehensive, systematic nomenclature that is easy to apply, descriptive and therefore highly informative. Recently, a f...

  19. Ultrasound mediated gene transfection

    Science.gov (United States)

    Williamson, Rene G.; Apfel, Robert E.; Brandsma, Janet L.

    2002-05-01

    Gene therapy is a promising modality for the treatment of a variety of human diseases both inherited and acquired, such as cystic fibrosis and cancer. The lack of an effective, safe method for the delivery of foreign genes into the cells, a process known as transfection, limits this effort. Ultrasound mediated gene transfection is an attractive method for gene delivery since it is a noninvasive technique, does not introduce any viral particles into the host and can offer very good temporal and spatial control. Previous investigators have shown that sonication increases transfection efficiency with and without ultrasound contrast agents. The mechanism is believed to be via a cavitation process where collapsing bubble nuclei permeabilize the cell membrane leading to increased DNA transfer. The research is focused on the use of pulsed wave high frequency focused ultrasound to transfect DNA into mammalian cells in vitro and in vivo. A better understanding of the mechanism behind the transfection process is also sought. A summary of some in vitro results to date will be presented, which includes the design of a sonication chamber that allows us to model the in vivo case more accurately.

  20. Novel targeting of PEGylated liposomes for codelivery of TGF-?1 siRNA and four antitubercular drugs to human macrophages for the treatment of mycobacterial infection: a quantitative proteomic study.

    Science.gov (United States)

    Niu, Ning-Kui; Yin, Juan-Juan; Yang, Yin-Xue; Wang, Zi-Li; Zhou, Zhi-Wei; He, Zhi-Xu; Chen, Xiao-Wu; Zhang, Xueji; Duan, Wei; Yang, Tianxin; Zhou, Shu-Feng

    2015-01-01

    Tuberculosis (TB) is still a major public health issue in developing countries, and its chemotherapy is compromised by poor drug compliance and severe side effects. This study aimed to synthesize and characterize new multimodal PEGylated liposomes encapsulated with clinically commonly used anti-TB drugs with linkage to small interfering RNA (siRNA) against transforming growth factor-?1 (TGF-?1). The novel NP-siRNA liposomes could target THP-1-derived human macrophages that were the host cells of mycobacterium infection. The biological effects of the NP-siRNA liposomes were evaluated on cell cycle distribution, apoptosis, autophagy, and the gene silencing efficiency of TGF-?1 siRNA in human macrophages. We also explored the proteomic responses to the newly synthesized NP-siRNA liposomes using the stable isotope labeling with amino acids in cell culture approach. The results showed that the multifunctional PEGylated liposomes were successfully synthesized and chemically characterized with a mean size of 265.1 nm. The novel NP-siRNA liposomes functionalized with the anti-TB drugs and TGF-?1 siRNA were endocytosed efficiently by human macrophages as visualized by transmission electron microscopy and scanning electron microscopy. Furthermore, the liposomes showed a low cytotoxicity toward human macrophages. There was no significant effect on cell cycle distribution and apoptosis in THP-1-derived macrophages after drug exposure at concentrations ranging from 2.5 to 62.5 ?g/mL. Notably, there was a 6.4-fold increase in the autophagy of human macrophages when treated with the NP-siRNA liposomes at 62.5 ?g/mL. In addition, the TGF-?1 and nuclear factor-?B expression levels were downregulated by the NP-siRNA liposomes in THP-1-derived macrophages. The Ingenuity Pathway Analysis data showed that there were over 40 signaling pathways involved in the proteomic responses to NP-siRNA liposome exposure in human macrophages, with 160 proteins mapped. The top five canonical signaling pathways were eukaryotic initiation factor 2 signaling, actin cytoskeleton signaling, remodeling of epithelial adherens junctions, epithelial adherens junction signaling, and Rho GDP-dissociation inhibitor signaling pathways. Collectively, the novel synthetic targeting liposomes represent a promising delivery system for anti-TB drugs to human macrophages with good selectivity and minimal cytotoxicity. PMID:26300629

  1. Genes contributing to prion pathogenesis

    DEFF Research Database (Denmark)

    Tamgüney, Gültekin; Giles, Kurt; Glidden, David V; Lessard, Pierre; Wille, Holger; Tremblay, Patrick; Groth, Darlene F; Yehiely, Fruma; Korth, Carsten; Moore, Richard C; Tatzelt, Jörg; Rubinstein, Eric; Boucheix, Claude; Yang, Xiaoping; Stanley, Pamela; Lisanti, Michael P; Dwek, Raymond A; Rudd, Pauline M; Moskovitz, Jackob; Epstein, Charles J; Cruz, Tracey Dawson; Kuziel, William A; Maeda, Nobuyo; Sap, Jan; Ashe, Karen Hsiao; Carlson, George A; Tesseur, Ina; Wyss-Coray, Tony; Mucke, Lennart; Weisgraber, Karl H; Mahley, Robert W; Cohen, Fred E; Prusiner, Stanley B

    incubation times, indicating that the conversion reaction may be influenced by other gene products. To identify genes that contribute to prion pathogenesis, we analysed incubation times of prions in mice in which the gene product was inactivated, knocked out or overexpressed. We tested 20 candidate genes...... show that many genes previously implicated in prion replication have no discernible effect on the pathogenesis of prion disease. While most genes tested did not significantly affect survival times, ablation of the amyloid beta (A4) precursor protein (App) or interleukin-1 receptor, type I (Il1r1), and...

  2. BRAF_(gene) - Wikipedia, the free encyclopedia [Gene Wiki

    Lifescience Database Archive (English)

    Full Text Available BRAF (gene) - Wikipedia, the free encyclopediaa:lang(ar),a:lang(kk-arab),a:lang(mzn),a:lang(ps), ... TD, Dolan CR; et al., eds. (1993–). GeneReviews™ [Internet ]. Seattle WA: University of Washington, Seattle. C ...

  3. Vertebrate gene predictions and the problem of large genes

    DEFF Research Database (Denmark)

    Wang, Jun; Li, ShengTing; Zhang, Yong; Zheng, HongKun; Xu, Zhao; Ye, Jia; Yu, Jun; Wong, Gane Ka-Shu

    2003-01-01

    To find unknown protein-coding genes, annotation pipelines use a combination of ab initio gene prediction and similarity to experimentally confirmed genes or proteins. Here, we show that although the ab initio predictions have an intrinsically high false-positive rate, they also have a consistently low false-negative rate. The incorporation of similarity information is meant to reduce the false-positive rate, but in doing so it increases the false-negative rate. The crucial variable is gene size...

  4. The Gene Wiki: community intelligence applied to human gene annotation

    OpenAIRE

    Huss, Jon W; Lindenbaum, Pierre; Martone, Michael; Roberts, Donabel; Pizarro, Angel; Valafar, Faramarz; Hogenesch, John B.; Su, Andrew I.

    2009-01-01

    Annotating the function of all human genes is a critical, yet formidable, challenge. Current gene annotation efforts focus on centralized curation resources, but it is increasingly clear that this approach does not scale with the rapid growth of the biomedical literature. The Gene Wiki utilizes an alternative and complementary model based on the principle of community intelligence. Directly integrated within the online encyclopedia, Wikipedia, the goal of this effort is to build a gene-specif...

  5. Using the Gene Ontology Hierarchy when Predicting Gene Function

    OpenAIRE

    Mostafavi, Sara; Morris, Quaid

    2012-01-01

    The problem of multilabel classification when the labels are related through a hierarchical categorization scheme occurs in many application domains such as computational biology. For example, this problem arises naturally when trying to automatically assign gene function using a controlled vocabularies like Gene Ontology. However, most existing approaches for predicting gene functions solve independent classification problems to predict genes that are involved in a given fu...

  6. Identification of significant periodic genes in microarray gene expression data

    OpenAIRE

    Chen Jie

    2005-01-01

    Abstract Background One frequent application of microarray experiments is in the study of monitoring gene activities in a cell during cell cycle or cell division. A new challenge for analyzing the microarray experiments is to identify genes that are statistically significantly periodically expressed during the cell cycle. Such a challenge occurs due to the large number of genes that are simultaneously measured, a moderate to small number of measurements per gene taken at different time points...

  7. Gene discovery and gene function assignment in filamentous fungi

    OpenAIRE

    Hamer, Lisbeth; Adachi, Kiichi; Montenegro-Chamorro, Maria V.; Tanzer, Matthew M.; Mahanty, Sanjoy K.; Lo, Clive; Tarpey, Rex W.; Skalchunes, Amy R.; Heiniger, Ryan W.; Frank, Sheryl A.; Darveaux, Blaise A.; Lampe, David J.; Slater, Ted M.; Ramamurthy, Lakshman; DeZwaan, Todd M.

    2001-01-01

    Filamentous fungi are a large group of diverse and economically important microorganisms. Large-scale gene disruption strategies developed in budding yeast are not applicable to these organisms because of their larger genomes and lower rate of targeted integration (TI) during transformation. We developed transposon-arrayed gene knockouts (TAGKO) to discover genes and simultaneously create gene disruption cassettes for subsequent transformation and mutant analysis. ...

  8. Gene doping in modern sport.

    Directory of Open Access Journals (Sweden)

    MAREK SAWCZUK

    2009-01-01

    Full Text Available Background: The subject of this paper is gene doping, which should be understood as "he non-therapeutic use of cells, genes, genetic elements, or of the modulation of gene expression, having the capacity to improve athletic performance". The authors of this work, based on the review of literature and previous research, make an attempt at wider characterization of gene doping and the discussion of related potential threats.Methods: This is a comprehensive survey of literature on the latest applications of molecular biology in medicine. The analysis involves a dozen scientific databases examined in order to find genes used in gene therapy and potentially useful in gene doping. Results: The obtained results enable better recognition of gene doping and indicate genes used in medicine that could be used in gene doping. This paper describes potential effects of their use and associated risk, and predicts the possible developments of gene doping in the future. Conclusion: Gene doping is undoubtedly a part of modern sport. Although WADA included gene doping on the list of banned methods as early as 2004, as previously stated above, it has not managed to develop efficient methods of detection.

  9. Avirulence Genes in Cereal Powdery Mildews: The Gene-for-Gene Hypothesis 2.0

    Science.gov (United States)

    Bourras, Salim; McNally, Kaitlin E.; Müller, Marion C.; Wicker, Thomas; Keller, Beat

    2016-01-01

    The gene-for-gene hypothesis states that for each gene controlling resistance in the host, there is a corresponding, specific gene controlling avirulence in the pathogen. Allelic series of the cereal mildew resistance genes Pm3 and Mla provide an excellent system for genetic and molecular analysis of resistance specificity. Despite this opportunity for molecular research, avirulence genes in mildews remain underexplored. Earlier work in barley powdery mildew (B.g. hordei) has shown that the reaction to some Mla resistance alleles is controlled by multiple genes. Similarly, several genes are involved in the specific interaction of wheat mildew (B.g. tritici) with the Pm3 allelic series. We found that two mildew genes control avirulence on Pm3f: one gene is involved in recognition by the resistance protein as demonstrated by functional studies in wheat and the heterologous host Nicotiana benthamiana. A second gene is a suppressor, and resistance is only observed in mildew genotypes combining the inactive suppressor and the recognized Avr. We propose that such suppressor/avirulence gene combinations provide the basis of specificity in mildews. Depending on the particular gene combinations in a mildew race, different genes will be genetically identified as the “avirulence” gene. Additionally, the observation of two LINE retrotransposon-encoded avirulence genes in B.g. hordei further suggests that the control of avirulence in mildew is more complex than a canonical gene-for-gene interaction. To fully understand the mildew–cereal interactions, more knowledge on avirulence determinants is needed and we propose ways how this can be achieved based on recent advances in the field.

  10. Novel targeting of PEGylated liposomes for codelivery of TGF-?1 siRNA and four antitubercular drugs to human macrophages for the treatment of mycobacterial infection: a quantitative proteomic study

    Directory of Open Access Journals (Sweden)

    Niu NK

    2015-08-01

    Full Text Available Ning-Kui Niu,1–3 Juan-Juan Yin,3 Yin-Xue Yang,4 Zi-Li Wang,1 Zhi-Wei Zhou,3 Zhi-Xu He,5 Xiao-Wu Chen,6 Xueji Zhang,7 Wei Duan,8 Tianxin Yang,9 Shu-Feng Zhou3 1Department of Orthopedics, General Hospital of Tianjin Medical University, Tianjin, 2Department of Spinal Surgery, General Hospital of Ningxia Medical University, Yinchuan, Ningxia, People’s Republic of China; 3Department of Pharmaceutical Sciences, College of Pharmacy, University of South Florida, Tampa, FL, USA; 4Department of Colorectal Surgery, General Hospital of Ningxia Medical University, Yinchuan, Ningxia, 5Guizhou Provincial Key Laboratory for Regenerative Medicine, Stem Cell and Tissue Engineering Research Center and Sino-US Joint Laboratory for Medical Sciences, Guizhou Medical University, Guiyang, Guizhou, 6Department of General Surgery, The First People’s Hospital of Shunde Affiliated to Southern Medical University, Shunde, Foshan, Guangdong, 7Research Center for Bioengineering and Sensing Technology, University of Science and Technology Beijing, Beijing, People’s Republic of China; 8School of Medicine, Deakin University, Waurn Ponds, VIC, Australia; 9Department of Internal Medicine, University of Utah and Salt Lake Veterans Affairs Medical Center, Salt Lake City, UT, USA Abstract: Tuberculosis (TB is still a major public health issue in developing countries, and its chemotherapy is compromised by poor drug compliance and severe side effects. This study aimed to synthesize and characterize new multimodal PEGylated liposomes encapsulated with clinically commonly used anti-TB drugs with linkage to small interfering RNA (siRNA against transforming growth factor-?1 (TGF-?1. The novel NP-siRNA liposomes could target THP-1-derived human macrophages that were the host cells of mycobacterium infection. The biological effects of the NP-siRNA liposomes were evaluated on cell cycle distribution, apoptosis, autophagy, and the gene silencing efficiency of TGF-?1 siRNA in human macrophages. We also explored the proteomic responses to the newly synthesized NP-siRNA liposomes using the stable isotope labeling with amino acids in cell culture approach. The results showed that the multifunctional PEGylated liposomes were successfully synthesized and chemically characterized with a mean size of 265.1 nm. The novel NP-siRNA liposomes functionalized with the anti-TB drugs and TGF-?1 siRNA were endocytosed efficiently by human macrophages as visualized by transmission electron microscopy and scanning electron microscopy. Furthermore, the liposomes showed a low cytotoxicity toward human macrophages. There was no significant effect on cell cycle distribution and apoptosis in THP-1-derived macrophages after drug exposure at concentrations ranging from 2.5 to 62.5 µg/mL. Notably, there was a 6.4-fold increase in the autophagy of human macrophages when treated with the NP-siRNA liposomes at 62.5 µg/mL. In addition, the TGF-?1 and nuclear factor-?B expression levels were downregulated by the NP-siRNA liposomes in THP-1-derived macrophages. The Ingenuity Pathway Analysis data showed that there were over 40 signaling pathways involved in the proteomic responses to NP-siRNA liposome exposure in human macrophages, with 160 proteins mapped. The top five canonical signaling pathways were eukaryotic initiation factor 2 signaling, actin cytoskeleton signaling, remodeling of epithelial adherens junctions, epithelial adherens junction signaling, and Rho GDP-dissociation inhibitor signaling pathways. Collectively, the novel synthetic targeting liposomes represent a promising delivery system for anti-TB drugs to human macrophages with good selectivity and minimal cytotoxicity. Keywords: tuberculosis, cytokine, liposome, apoptosis, autophagy, cell cycle, proteomics, SILAC, NF-?B, interleukin

  11. Eukaryotic Gene Prediction Using GeneMark.hmm-E and GeneMark-ES

    OpenAIRE

    Borodovsky, Mark; Lomsadze, Alex

    2011-01-01

    This unit describes how to use gene finding programs GeneMark.hmm-E and GeneMark-ES for finding protein-coding genes in genomic DNA of eukaryotic genomes. These bioinformatics tools were demonstrated to have state-of-the-art accuracy for many fungal, plant and animal genomes and have been frequently used for gene annotation in novel genomic sequences. Additional advantage of GeneMark-ES is that the problem of algorithm parameterization is solved automatically, with parameters estimated by ite...

  12. Genes, stress, and depression.

    Science.gov (United States)

    Wurtman, Richard J

    2005-05-01

    A relationship between genetic makeup and susceptibility to major depressive disorder (MDD) has long been suspected on the basis of family and twin studies. A metaanalysis of reports on the basis of twin studies has estimated MDD's degree of heritability to be 0.33 (confidence interval, 0.26-0.39). Among families exhibiting an increased prevalence of MDD, risk of developing the illness was enhanced in members exposed to a highly stressful environment. Aberrant genes can predispose to depression in a number of ways, for example, by diminishing production of growth factors that act during brain development. An aberrant gene could also increase or decrease a neurotransmitter's release into synapses, its actions, or its duration of activity. The gene products of greatest interest at present are those involved in the synthesis and actions of serotonin; among them, the serotonin-uptake protein localized within the terminals and dendrites of serotonin-releasing neurons. It has been found that the Vmax of platelet serotonin uptake is low in some patients with MDD; also, Vmax is highly correlated in twins. Antidepressant drugs such as the selective serotonin reuptake inhibitors act on this uptake protein. The specific genetic locus causing serotonin uptake to be lower in some patients with major depression involves a polymorphic region (5-HTTLPR) in the promoter region of the gene for the uptake protein. The gene itself exists as several alleles, the short "S" allele and the long "L" allele. The S variant is associated with less, and the L variant with more, of the uptake protein. The effect of stressful life events on depressive symptoms in young adults was found to be significantly stronger among SS or SL subjects than among LL subjects. Neuroimaging studies showed that people with the SS or SL alleles exhibited a greater activation of the amygdala in response to fearful stimuli than those with LL. It has been reported recently that mutations in the gene that controls serotonin synthesis in the human brain (tryptophan hydroxylase) also predispose to mood disturbances. It may be asked whether people who lack a psychiatric history should be advised to avoid stressful environments if they are found to carry the SS or SL alleles. PMID:15877307

  13. Gene Testing for Hereditary Ataxia

    Science.gov (United States)

    FAQ NATIONAL ATAXIA FOUNDATION FREQUENTLY ASKED QUESTIONS ABOUT... Gene Testing for Hereditary Ataxia This fact sheet provides an overview of gene testing for ataxia. It also addresses commonly asked ...

  14. Genes and human brain evolution

    OpenAIRE

    Geschwind, Daniel H.; Konopka, Genevieve

    2012-01-01

    Several genes were duplicated during human evolution. It seems that one such duplication gave rise to a gene that may have helped to make human brains bigger and more adaptable than those of our ancestors.

  15. Gene therapy of liver cancer

    OpenAIRE

    Hernandez-Alcoceba, R. (Rubén); B. Sangro; Prieto, J. (Jesús)

    2006-01-01

    The application of gene transfer technologies to the treatment of cancer has led to the development of new experimental approaches like gene directed enzyme/pro-drug therapy (GDEPT), inhibition of oncogenes and restoration of tumor-suppressor genes. In addition, gene therapy has a big impact on other fields like cancer immunotherapy, anti-angiogenic therapy and virotherapy. These strategies are being evaluated for the treatment of primary and metastatic liver cancer and some of them have reac...

  16. Human housekeeping genes are compact

    OpenAIRE

    Eisenberg, Eli; Levanon, Erez Y.

    2003-01-01

    We identify a set of 575 human genes that are expressed in all conditions tested in a publicly available database of microarray results. Based on this common occurrence, the set is expected to be rich in "housekeeping" genes, showing constitutive expression in all tissues. We compare selected aspects of their genomic structure with a set of background genes. We find that the introns, untranslated regions and coding sequences of the housekeeping genes are shorter, indicating ...

  17. Nontuberculous mycobacteria in respiratory samples from patients with pulmonary tuberculosis in the state of Rondonia, Brazil

    Directory of Open Access Journals (Sweden)

    Cleoni Alves Mendes de Lima

    2013-06-01

    Full Text Available The main cause of pulmonary tuberculosis (TB is infection with Mycobacterium tuberculosis (MTB. We aimed to evaluate the contribution of nontuberculous mycobacteria (NTM to pulmonary disease in patients from the state of Rondônia using respiratory samples and epidemiological data from TB cases. Mycobacterium isolates were identified using a combination of conventional tests, polymerase chain reaction-based restriction enzyme analysis of hsp65 gene and hsp65 gene sequencing. Among the 1,812 cases suspected of having pulmonary TB, 444 yielded bacterial cultures, including 369 cases positive for MTB and 75 cases positive for NTM. Within the latter group, 14 species were identified as Mycobacterium abscessus, Mycobacterium avium, Mycobacterium fortuitum, Mycobacterium intracellulare, Mycobacterium gilvum, Mycobacterium gordonae, Mycobacterium asiaticum, Mycobacterium tusciae, Mycobacterium porcinum, Mycobacterium novocastrense, Mycobacterium simiae, Mycobacterium szulgai, Mycobacterium phlei and Mycobacterium holsaticum and 13 isolates could not be identified at the species level. The majority of NTM cases were observed in Porto Velho and the relative frequency of NTM compared with MTB was highest in Ji-Paraná. In approximately half of the TB subjects with NTM, a second sample containing NTM was obtained, confirming this as the disease-causing agent. The most frequently observed NTM species were M. abscessus and M. avium and because the former species is resistant to many antibiotics and displays unsatisfactory cure rates, the implementation of rapid identification of mycobacterium species is of considerable importance.

  18. Genes and addiction.

    Science.gov (United States)

    Nestler, E J

    2000-11-01

    Drug addiction, like all psychiatric disorders, is defined solely in behavioural terms. For example, addiction can be considered a loss of control over drug-taking, or compulsive drug-seeking and -taking despite horrendous consequences. Abnormal behaviours are a consequence of aberrant brain function, which means that it is a tangible goal to identify the biological underpinnings of addiction. The genetic basis of addiction encompasses two broad areas of enquiry. One of these is the identification of genetic variation in humans that partly determines susceptibility to addiction. The other is the use of animal models to investigate the role of specific genes in mediating the development of addiction. Whereas recent advances in this latter effort are heartening, a major challenge remains: to understand how the many genes implicated in rodent models interact to yield as complex a phenotype as addiction. PMID:11062465

  19. Graphene based gene transfection

    Science.gov (United States)

    Feng, Liangzhu; Zhang, Shuai; Liu, Zhuang

    2011-03-01

    Graphene as a star in materials research has been attracting tremendous attentions in the past few years in various fields including biomedicine. In this work, for the first time we successfully use graphene as a non-toxic nano-vehicle for efficient gene transfection. Graphene oxide (GO) is bound with cationic polymers, polyethyleneimine (PEI) with two different molecular weights at 1.2 kDa and 10 kDa, forming GO-PEI-1.2k and GO-PEG-10k complexes, respectively, both of which are stable in physiological solutions. Cellular toxicity tests reveal that our GO-PEI-10k complex exhibits significantly reduced toxicity to the treated cells compared to the bare PEI-10k polymer. The positively charged GO-PEI complexes are able to further bind with plasmid DNA (pDNA) for intracellular transfection of the enhanced green fluorescence protein (EGFP) gene in HeLa cells. While EGFP transfection with PEI-1.2k appears to be ineffective, high EGFP expression is observed using the corresponding GO-PEI-1.2k as the transfection agent. On the other hand, GO-PEI-10k shows similar EGFP transfection efficiency but lower toxicity compared with PEI-10k. Our results suggest graphene to be a novel gene delivery nano-vector with low cytotoxicity and high transfection efficiency, promising for future applications in non-viral based gene therapy.Graphene as a star in materials research has been attracting tremendous attentions in the past few years in various fields including biomedicine. In this work, for the first time we successfully use graphene as a non-toxic nano-vehicle for efficient gene transfection. Graphene oxide (GO) is bound with cationic polymers, polyethyleneimine (PEI) with two different molecular weights at 1.2 kDa and 10 kDa, forming GO-PEI-1.2k and GO-PEG-10k complexes, respectively, both of which are stable in physiological solutions. Cellular toxicity tests reveal that our GO-PEI-10k complex exhibits significantly reduced toxicity to the treated cells compared to the bare PEI-10k polymer. The positively charged GO-PEI complexes are able to further bind with plasmid DNA (pDNA) for intracellular transfection of the enhanced green fluorescence protein (EGFP) gene in HeLa cells. While EGFP transfection with PEI-1.2k appears to be ineffective, high EGFP expression is observed using the corresponding GO-PEI-1.2k as the transfection agent. On the other hand, GO-PEI-10k shows similar EGFP transfection efficiency but lower toxicity compared with PEI-10k. Our results suggest graphene to be a novel gene delivery nano-vector with low cytotoxicity and high transfection efficiency, promising for future applications in non-viral based gene therapy. Electronic supplementary information (ESI) available: Thickness distribution of GO and GO-PEI; IR and TGA data; and confocal images of HeLa cells treated with bare EGFP pDNA and GO + pDNA. See DOI: 10.1039/c0nr00680g

  20. Gene and Aging

    Directory of Open Access Journals (Sweden)

    DD Farhud

    2008-09-01

    Full Text Available "nCollection of multiple processes that increase the chronological age of an organism leading to death is defined as aging, and even though important, it is poorly understood. Recent research has shown that aging is due to biochemical and genetic changes, in interaction with environmental effects, including diet and nutrition. Most knowledge on aging is based on ge­netic model system, but its molecular mechanisms are still not very clear. Discoveries in molecular biology have made way to look for candidate genes influencing lifespan. Furthermore, new investigations have stressed on the roles of mitochondria as the major generators and direct targets of reactive oxygen species. This paper reviews some recent literature on genes and ag­ing in model system, then discusses the role of mitochondria and nutrients in human aging.

  1. Alphaviruses in Gene Therapy

    Directory of Open Access Journals (Sweden)

    Kenneth Lundstrom

    2009-04-01

    Full Text Available Alphaviruses are enveloped single stranded RNA viruses, which as gene therapy vectors provide high-level transient gene expression. Semliki Forest virus (SFV, Sindbis virus (SIN and Venezuelan Equine Encephalitis (VEE virus have been engineered as efficient replication-deficient and -competent expression vectors. Alphavirus vectors have frequently been used as vehicles for tumor vaccine generation. Moreover, SFV and SIN vectors have been applied for intratumoral injections in animals implanted with tumor xenografts. SIN vectors have demonstrated natural tumor targeting, which might permit systemic vector administration. Another approach for systemic delivery of SFV has been to encapsulate replication-deficient viral particles in liposomes, which can provide passive targeting to tumors and allow repeated administration without host immune responses. This approach has demonstrated safe delivery of encapsulated SFV particles to melanoma and kidney carcinoma patients in a phase I trial. Finally, the prominent neurotropism of alphaviruses make them attractive for the treatment of CNS-related diseases.

  2. Genes y especies

    Directory of Open Access Journals (Sweden)

    A. G. Sáez

    2009-01-01

    Full Text Available La posibilidad de secuenciar y manipular los genes sigue abriendo fronteras en el estudio de las especies y la especiación. En tiempos recientes particularmente en dos direcciones. Por un lado, la secuenciación del gen COI (y otros en miles de muestras de forma sistemática y masiva, el llamado "DNA barcoding", está revelando una gran cantidad de biodiversidad, en buena medida previamente no sospechada y correspondiente a especies crípticas (morfológicamente irreconocibles. En segundo lugar, el estudio de los genes responsables de los cambios morfológicos nos está haciendo volver a dar una creciente importancia a la selección como motor de la especiación y de la evolución, incluso en presencia de dosis importantes de flujo génico.

  3. Endovascular Gene Delivery from a Stent Platform: Gene- Eluting Stents

    OpenAIRE

    Fishbein, Ilia; Chorny, Michael; Adamo, Richard F; Forbes, Scott P; Corrales, Ricardo A; Alferiev, Ivan S.; Levy, Robert J

    2013-01-01

    A synergistic impact of research in the fields of post-angioplasty restenosis, drug-eluting stents and vascular gene therapy over the past 15 years has shaped the concept of gene-eluting stents. Gene-eluting stents hold promise of overcoming some biological and technical problems inherent to drug-eluting stent technology. As the field of gene-eluting stents matures it becomes evident that all three main design modules of a gene-eluting stent: a therapeutic transgene, a vector and a delivery s...

  4. Gene therapy of cancer and development of therapeutic target gene

    International Nuclear Information System (INIS)

    We applied HSV-tk/GCV strategy to orthotopic rat hepatoma model and showed anticancer effects of hepatoma. The increased expression of Lac Z gene after adenovirus-mediated gene delivery throughout hepatic artery was thought that is increased the possibility of gene therapy for curing hepatoma. With the construction of kGLP-laboratory, it is possible to produce a good quantity and quality of adenovirus in lage-scale production and purification of adenovirus vector. Also, the analysis of hepatoma related genes by PCR-LOH could be used for the diagnosis of patients and the development of therapeutic gene

  5. GoGene: gene annotation in the fast lane

    OpenAIRE

    Plake, Conrad; Royer, Loic; Winnenburg, Rainer; Hakenberg, Jörg; Schroeder, Michael

    2009-01-01

    High-throughput screens such as microarrays and RNAi screens produce huge amounts of data. They typically result in hundreds of genes, which are often further explored and clustered via enriched GeneOntology terms. The strength of such analyses is that they build on high-quality manual annotations provided with the GeneOntology. However, the weakness is that annotations are restricted to process, function and location and that they do not cover all known genes in model organisms. GoGene addre...

  6. Gene therapy of cancer and development of therapeutic target gene

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Chang Min; Kwon, Hee Chung

    1998-04-01

    We applied HSV-tk/GCV strategy to orthotopic rat hepatoma model and showed anticancer effects of hepatoma. The increased expression of Lac Z gene after adenovirus-mediated gene delivery throughout hepatic artery was thought that is increased the possibility of gene therapy for curing hepatoma. With the construction of kGLP-laboratory, it is possible to produce a good quantity and quality of adenovirus in lage-scale production and purification of adenovirus vector. Also, the analysis of hepatoma related genes by PCR-LOH could be used for the diagnosis of patients and the development of therapeutic gene.

  7. Gene electrotransfer in clinical trials

    DEFF Research Database (Denmark)

    Gehl, Julie

    2014-01-01

    Electroporation is increasingly being used for delivery of chemotherapy to tumors. Likewise, gene delivery by electroporation is rapidly gaining momentum for both vaccination purposes and for delivery of genes coding for other therapeutic molecules, such as chronic diseases or cancer. This chapter describes how gene therapy may be performed using electric pulses to enhance uptake and expression.

  8. Vascular complications and gene therapy.

    Science.gov (United States)

    Roy, Sayon; Rothschild, Jennifer G; Chen, Amy

    2003-02-01

    For gene therapy, the last few years have been an exciting period. Encouraging results from several successful gene therapy trials were reported. Children born with a life-threatening immune system disorder, severe combined immune deficiency (SCID), were cured after receiving gene therapy for replacement of their defective adenosine deaminase (ADA) gene. Gene therapy successes related to vascular complications were also reported. The first human gene therapy trial for a blood-vessel disorder was performed successfully, in which copies of an angiogenic gene, the vascular endothelial growth factor (VEGF) gene, were directly delivered to the area surrounding the diseased artery of the leg of a patient with peripheral artery disease. Within a few days, this stimulated the growth of new blood vessels around the blockage in the ailing blood vessel and helped avoid amputation. In 1998, a patient with genetically small arteries became the first to receive VEGF gene therapy in the heart. Multiple copies of a plasmid with the VEGF gene were delivered into the damaged area of the heart, and a few days later angiogenesis ensued that helped bypass the blocked vessel, with markedly reduced chest pain in the patient. Gene therapy is becoming a reality and, more importantly, it appears to be safe and does not require supplementary immuno-suppressing drugs. Gene therapy seems to have begun delivering on its promises. PMID:12718732

  9. Gene therapy for thyroid cancer

    International Nuclear Information System (INIS)

    Gene therapy for thyroid cancer include immunotherapy, suicide gene therapy, tumor suppressor replacement, 131I therapy by sodium/iodide symporter and antisense therapy and so on. Gene therapy has wide perspectives, but there are many problems need to be solved for clinical application

  10. Compositional gradients in Gramineae genes

    DEFF Research Database (Denmark)

    Wong, Gane Ka-Shu; Wang, Jun; Tao, Lin; Tan, Jun; Zhang, JianGuo; Passey, Douglas A; Yu, Jun

    2002-01-01

    In this study, we describe a property of Gramineae genes, and perhaps all monocot genes, that is not observed in eudicot genes. Along the direction of transcription, beginning at the junction of the 5'-UTR and the coding region, there are gradients in GC content, codon usage, and amino-acid usage...

  11. Independent Gene Discovery and Testing

    Science.gov (United States)

    Palsule, Vrushalee; Coric, Dijana; Delancy, Russell; Dunham, Heather; Melancon, Caleb; Thompson, Dennis; Toms, Jamie; White, Ashley; Shultz, Jeffry

    2010-01-01

    A clear understanding of basic gene structure is critical when teaching molecular genetics, the central dogma and the biological sciences. We sought to create a gene-based teaching project to improve students' understanding of gene structure and to integrate this into a research project that can be implemented by instructors at the secondary level…

  12. Gene electrotransfer in clinical trials

    DEFF Research Database (Denmark)

    Gehl, Julie

    2014-01-01

    Electroporation is increasingly being used for delivery of chemotherapy to tumors. Likewise, gene delivery by electroporation is rapidly gaining momentum for both vaccination purposes and for delivery of genes coding for other therapeutic molecules, such as chronic diseases or cancer. This chapter...... describes how gene therapy may be performed using electric pulses to enhance uptake and expression....

  13. The Perils of Gene Patents

    OpenAIRE

    Salzberg, SL

    2012-01-01

    I argue here that gene patents, and patented genetic tests based on them, are a very bad idea. First, I discuss whether genes can reasonably be the subject of patents in the first place; I maintain that the answer is no. Second, I explain how gene patents interfere with scientific progress, slowing down the development of new cures and treatments for genetic diseases.

  14. Organellar gene expression

    OpenAIRE

    Preuten, Tobias

    2010-01-01

    Zusätzlich zu der eubakteriellen RNA-Polymerase (RNAP) der Plastiden sind im Zellkern von Arabidopsis thaliana drei weitere, phagentypische RNAP kodiert, die jeweils aus nur einer Einheit aufgebaut sind. Die Enzyme RpoTp und RpoTm werden in die Plastiden, bzw. die Mitochondrien transportiert, während RpoTmp in beiden Organellen zu finden ist. Um die Lichtabhängigkeit der RpoT-Gene zu untersuchen, wurde die lichtinduzierte Akkumulation ihrer Transkripte in 7-Tage alten Keimlingen, sowie 3- bzw...

  15. Genes, crianças e pediatras

    Scientific Electronic Library Online (English)

    Esmeralda, Martins; Teresa, Oliveira; Anabela, Bandeira.

    Full Text Available [...] Abstract in english Homocystinuria is an autosomal recessive disease due to cystathionine-synthase deficiency, with the gene CBS being located in chromosome 21. In its typical presentation the eye, skeleton, central nervous system, and vascular system are all involved. The patient is normal at birth and in non-treated [...] patients tall stature and ectopia lentis may be the first symptoms, as in the case we present.

  16. Gene therapy for arthritis

    OpenAIRE

    Traister, Russell S; Hirsch, Raphael

    2008-01-01

    Arthritis is among the leading causes of disability in the developed world. There remains no cure for this disease and the current treatments are only modestly effective at slowing the disease's progression and providing symptomatic relief. The clinical effectiveness of current treatment regimens has been limited by short half-lives of the drugs and the requirement for repeated systemic administration. Utilizing gene transfer approaches for the treatment of arthritis may overcome some of the ...

  17. Gene Expression in Bone

    Science.gov (United States)

    D'Ambrogio, A.

    Skeletal system has two main functions, to provide mechanical integrity for both locomotion and protection and to play an important role in mineral homeostasis. There is extensive evidence showing loss of bone mass during long-term Space-Flights. The loss is due to a break in the equilibrium between the activity of osteoblasts (the cells that forms bone) and the activity of osteoclasts (the cells that resorbs bone). Surprisingly, there is scanty information about the possible altered gene expression occurring in cells that form bone in microgravity.(Just 69 articles result from a "gene expression in microgravity" MedLine query.) Gene-chip or microarray technology allows to screen thousands of genes at the same time: the use of this technology on samples coming from cells exposed to microgravity could provide us with many important informations. For example, the identification of the molecules or structures which are the first sensors of the mechanical stress derived from lack of gravity, could help in understanding which is the first event leading to bone loss due to long-term exposure to microgravity. Consequently, this structure could become a target for a custom-designed drug. It is evident that bone mass loss, observed during long-time stay in Space, represents an accelerated model of what happens in aging osteoporosis. Therefore, the discovery and design of drugs able to interfere with the bone-loss process, could help also in preventing negative physiological processes normally observed on Earth. Considering the aims stated above, my research is designed to:

  18. Are there anxious genes?

    OpenAIRE

    Morris-Rosendahl, Deborah J.

    2002-01-01

    Anxiety comprises many clinical descriptions and phenotypes. A genetic predisposition to anxiety is undoubted; however, the nature and extent of that contribution is still unclear. Methods for the genetic analysis of such complex disorders is briefly reviewed, followed by a discussion of the comorbidity of anxiety with other psychiatric disorders and their possible common genetic etiology. Extensive genetic studies of the serotonin (5-hydroxytryptamine, 5-HT) transporter (5-HTT) gene have rev...

  19. Gene therapy in diabetes

    OpenAIRE

    Wong, Mary S; Hawthorne, Wayne J; Manolios, Nicholas

    2010-01-01

    Type 1 diabetes (T1D) is a chronic autoimmune disease, whereby auto-reactive cytotoxic T cells target and destroy insulin-secreting ?-cells in pancreatic islets leading to insulin deficiency and subsequent hyperglycemia. These individuals require multiple daily insulin injections every day of their life without which they will develop life-threatening diabetic ketoacidosis (DKA) and die. Gene therapy by viral vector and non-viral transduction may be useful techniques to treat T1D as it can be...

  20. nanosheets for gene therapy

    Science.gov (United States)

    Kou, Zhongyang; Wang, Xin; Yuan, Renshun; Chen, Huabin; Zhi, Qiaoming; Gao, Ling; Wang, Bin; Guo, Zhaoji; Xue, Xiaofeng; Cao, Wei; Guo, Liang

    2014-10-01

    A new class of two-dimensional (2D) nanomaterial, transition metal dichalcogenides (TMDCs) such as MoS2, MoSe2, WS2, and WSe2 which have fantastic physical and chemical properties, has drawn tremendous attention in different fields recently. Herein, we for the first time take advantage of the great potential of MoS2 with well-engineered surface as a novel type of 2D nanocarriers for gene delivery and therapy of cancer. In our system, positively charged MoS2-PEG-PEI is synthesized with lipoic acid-modified polyethylene glycol (LA-PEG) and branched polyethylenimine (PEI). The amino end of positively charged nanomaterials can bind to the negatively charged small interfering RNA (siRNA). After detection of physical and chemical characteristics of the nanomaterial, cell toxicity was evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Polo-like kinase 1 (PLK1) was investigated as a well-known oncogene, which was a critical regulator of cell cycle transmission at multiple levels. Through knockdown of PLK1 with siRNA carried by novel nanovector, qPCR and Western blot were used to measure the interfering efficiency; apoptosis assay was used to detect the transfection effect of PLK1. All results showed that the novel nanocarrier revealed good biocompatibility, reduced cytotoxicity, as well as high gene-carrying ability without serum interference, thus would have great potential for gene delivery and therapy.

  1. Gene Network Biological Validity Based on Gene-Gene Interaction Relevance

    OpenAIRE

    Francisco Gómez-Vela; Norberto Díaz-Díaz

    2014-01-01

    In recent years, gene networks have become one of the most useful tools for modeling biological processes. Many inference gene network algorithms have been developed as techniques for extracting knowledge from gene expression data. Ensuring the reliability of the inferred gene relationships is a crucial task in any study in order to prove that the algorithms used are precise. Usually, this validation process can be carried out using prior biological knowledge. The metabolic pathways stored in...

  2. Identification of Significant Association and Gene-Gene Interaction of GABA Receptor Subunit Genes in Autism

    OpenAIRE

    Ma, D. Q.; Whitehead, P.L.; Menold, M. M.; Martin, E R; Ashley-Koch, A. E.; Mei, H (Hailiang); Ritchie, M D; Delong, G R; Abramson, R K; Wright, H.H.; Cuccaro, M. L.; Hussman, J. P.; Gilbert, J. R.; Pericak-Vance, M A

    2005-01-01

    Autism is a common neurodevelopmental disorder with a significant genetic component. Existing research suggests that multiple genes contribute to autism and that epigenetic effects or gene-gene interactions are likely contributors to autism risk. However, these effects have not yet been identified. Gamma-aminobutyric acid (GABA), the primary inhibitory neurotransmitter in the adult brain, has been implicated in autism etiology. Fourteen known autosomal GABA receptor subunit genes were studied...

  3. DIFFERENTIATION BETWEEN Nocardia spp. AND Mycobacterium spp.: CRITICAL ASPECTS FOR BACTERIOLOGICAL DIAGNOSIS / Diferenciação de Nocardia spp. e Mycobacterium spp.: aspectos críticos para o diagnóstico bacteriológico

    Scientific Electronic Library Online (English)

    Edna Cleide Mendes, Muricy; Romilda Aparecida, Lemes; Sidney, Bombarda; Lucilaine, Ferrazoli; Erica, Chimara.

    2014-09-01

    Full Text Available Novas metodologias têm sido desenvolvidas para a identificação de Nocardia spp. mas o diagnóstico inicial ainda necessita de método rápido e preciso, principalmente devido à similaridade com o gênero Mycobacterium, clínica e bacteriologicamente. O crescimento em meio de Löwenstein Jensen (LJ), a pre [...] sença de bacilos corados pela coloração de Ziehl Neelsen e colônias com características diferentes podem ser fatores de confusão entre nocardias e micobactérias. Este estudo descreve a ocorrência de Nocardia spp. em laboratório de referência em micobacteriologia, observando-se as principais dificuldades em diferenciar Nocardia spp. e Mycobacterium spp., correlacionando isolados com casos de nocardiose. Os registros laboratoriais dos anos 2008 a 2012 foram analisados e os isolados identificados como Nocardia sp. ou como bacilos não álcool - ácido resistentes (NBAAR) foram selecionados. Os dados epidemiológicos e bacteriológicos foram analisados. Trinta e três isolados identificados como Nocardia sp. e 22 como NBAAR foram selecionados para este estudo, perfazendo 0,12% do total de isolados identificados no período estudado. A identificação presuntiva foi baseada na morfologia macroscópica e microscópica, resistência à lisozima e perfis de restrição pelo método PRA-hsp65. Nocardia spp. pode crescer em meios de isolamento para micobactérias (LJ e BBL MGIT™) e microscopia de morfologia e as colônias são muito semelhantes a algumas espécies de micobactérias. Dezessete pacientes (54,8%) foram notificados e tratados para tuberculose, mas apresentaram sinais e sintomas para nocardiose. Concluimos que a ocorrência de Nocardia sp. no período estudado foi de 0,12%. Os isolados com características de bacilos filamentosos, formadores de hifas aéreas, com colônias que podem ter pigmento, rugosas e que não possuem padrão de digestão para BstEII no método PRA-hsp65 são sugestivos de Nocardia spp. Para um laboratório de rotina de Micobactérias, um fluxo de identificação presuntiva para Nocardia spp. é essencial para permitir que esses isolados sejam identificados com técnicas mais precisas, para que seja oferecido o tratamento adequado e qualidade de vida aos pacientes. Abstract in english New methodologies were developed for the identification of Nocardia but the initial diagnosis still requires a fast and accurate method, mainly due to the similarity to Mycobacterium, both clinical and bacteriologically. Growth on Löwenstein-Jensen (LJ) medium, presence of acid-fast bacilli through [...] Ziehl-Neelsen staining, and colony morphology can be confusing aspects between Nocardia and Mycobacterium. This study describes the occurrence of Nocardia spp. in a mycobacterial-reference laboratory, observing the main difficulties in differentiating Nocardia spp. from Mycobacterium spp., and correlating isolates with nocardiosis cases. Laboratory records for the period between 2008 and 2012 were analyzed, and the isolates identified as Nocardia sp. or as non-acid-fast filamentous bacilli were selected. Epidemiological and bacteriological data were analyzed as well. Thirty-three isolates identified as Nocardia sp. and 22 as non-acid-fast bacilli were selected for this study, and represented 0.12% of isolates during the study period. The presumptive identification was based on macroscopic and microscopic morphology, resistance to lysozyme and restriction profiles using the PRA-hsp65 method. Nocardia spp. can grow on media for mycobacteria isolation (LJ and BBL MGIT™) and microscopy and colony morphology are very similar to some mycobacteria species. Seventeen patients (54.8%) were reported and treated for tuberculosis, but presented signs and symptoms of nocardiosis. It was concluded that the occurrence of Nocardia sp. during the study period was 0.12%. Isolates with characteristics of filamentous bacilli, forming aerial hyphae, with colonies that may be pigmented, rough and without the BstEII digestion pattern in PRA-hsp65 method are suggestive of Nocardia spp. For a myc

  4. Gene Therapy and Children (For Parents)

    Science.gov (United States)

    ... Kids Deal With Bullies Pregnant? What to Expect Gene Therapy and Children KidsHealth > Parents > Doctors & Hospitals > Medical Tests & ... by a "bad" gene. Continue Two Types of Gene Therapy The two forms of gene therapy are: Somatic ...

  5. Genetics Home Reference: What is gene therapy?

    Science.gov (United States)

    ... Precision Medicine Next Handbook > Gene Therapy > What is gene therapy? Gene therapy is an experimental technique that uses ... have no other cures. For general information about gene therapy: MedlinePlus from the National Library of Medicine offers ...

  6. Gene Therapy and Children (For Parents)

    Science.gov (United States)

    ... Caring for Your Child All About Food Allergies Gene Therapy and Children KidsHealth > For Parents > Gene Therapy ... that don't respond to conventional therapies. About Genes Our genes help make us unique. Inherited from ...

  7. Progress in gene targeting and gene therapy for retinitis pigmentosa

    Energy Technology Data Exchange (ETDEWEB)

    Farrar, G.J.; Humphries, M.M.; Erven, A. [Trinity College, Dublin (Ireland)] [and others

    1994-09-01

    Previously, we localized disease genes involved in retinitis pigmentosa (RP), an inherited retinal degeneration, close to the rhodopsin and peripherin genes on 3q and 6p. Subsequently, we and others identified mutations in these genes in RP patients. Currently animal models for human retinopathies are being generated using gene targeting by homologous recombination in embryonic stem (ES) cells. Genomic clones for retinal genes including rhodopsin and peripherin have been obtained from a phage library carrying mouse DNA isogenic with the ES cell line (CC1.2). The peripherin clone has been sequenced to establish the genomic structure of the mouse gene. Targeting vectors for rhodopsin and peripherin including a neomycin cassette for positive selection and thymidine kinase genes enabling selection against random intergrants are under construction. Progress in vector construction will be presented. Simultaneously we are developing systems for delivery of gene therapies to retinal tissues utilizing replication-deficient adenovirus (Ad5). Efficacy of infection subsequent to various methods of intraocular injection and with varying viral titers is being assayed using an adenovirus construct containing a CMV promoter LacZ fusion as reporter and the range of tissues infected and the level of duration of LacZ expression monitored. Viral constructs with the LacZ reporter gene under the control of retinal specific promoters such as rhodopsin and IRBP cloned into pXCJL.1 are under construction. An update on developments in photoreceptor cell-directed expression of virally delivered genes will be presented.

  8. Maximum Gene-Support Tree

    Directory of Open Access Journals (Sweden)

    Yunfeng Shan

    2008-01-01

    Full Text Available Genomes and genes diversify during evolution; however, it is unclear to what extent genes still retain the relationship among species. Model species for molecular phylogenetic studies include yeasts and viruses whose genomes were sequenced as well as plants that have the fossil-supported true phylogenetic trees available. In this study, we generated single gene trees of seven yeast species as well as single gene trees of nine baculovirus species using all the orthologous genes among the species compared. Homologous genes among seven known plants were used for validation of the ?nding. Four algorithms—maximum parsimony (MP, minimum evolution (ME, maximum likelihood (ML, and neighbor-joining (NJ—were used. Trees were reconstructed before and after weighting the DNA and protein sequence lengths among genes. Rarely a gene can always generate the “true tree” by all the four algorithms. However, the most frequent gene tree, termed “maximum gene-support tree” (MGS tree, or WMGS tree for the weighted one, in yeasts, baculoviruses, or plants was consistently found to be the “true tree” among the species. The results provide insights into the overall degree of divergence of orthologous genes of the genomes analyzed and suggest the following: 1 The true tree relationship among the species studied is still maintained by the largest group of orthologous genes; 2 There are usually more orthologous genes with higher similarities between genetically closer species than between genetically more distant ones; and 3 The maximum gene-support tree re?ects the phylogenetic relationship among species in comparison.

  9. Nonallelic Gene Conversion in the Genus Drosophila

    OpenAIRE

    Casola, Claudio; Ganote, Carrie L.; Hahn, Matthew W.

    2010-01-01

    Nonallelic gene conversion has been proposed as a major force in homogenizing the sequences of paralogous genes. In this work, we investigate the extent and characteristics of gene conversion among gene families in nine species of the genus Drosophila. We carried out a genome-wide study of 2855 gene families (including 17,742 genes) and determined that conversion events involved 2628 genes. The proportion of converted genes ranged across species from 1 to 9% when paralogs of all ages were inc...

  10. Genes, crianças e pediatras

    Scientific Electronic Library Online (English)

    Joana, Correia; Marta, Rios; Paula, Ferreira; Esmeralda, Martins; Anabela, Bandeira.

    2013-09-01

    Full Text Available [...] Abstract in english A male newborn, presenting hipotonia and posterior parietal bossing, developed, in the first 12 hours of life, refusal to feed and hypoglycaemia. A cranial ultrasound, skull X-ray and CT scan revealed an occipital and parietal fracture with an underlying haematoma and extensive extracranial soft-tis [...] sue swelling. He was submitted to surgical drainage. After 24 hours: new intracerebral bleeding. At the age of two-months he presented abnormal skin and sparse kinky hair. Serum copper and caeruloplasmin levels were below the normal range. Molecular diagnosis of Menkes disease was made by the identification of a new mutation in ATP7A gene.

  11. MUTATIONS IN CALMODULIN GENES

    DEFF Research Database (Denmark)

    The present invention relates to an isolated polynucleotide encoding at least a part of calmodulin and an isolated polypeptide comprising at least a part of a calmodulin protein, wherein the polynucleotide and the polypeptide comprise at least one mutation associated with a cardiac disorder. The ...... binding of calmodulin to ryanodine receptor 2 and use of such compound in a treatment of an individual having a cardiac disorder. The invention further provides a kit that can be used to detect specific mutations in calmodulin encoding genes....

  12. Alternative Gene Form Discovery and Candidate Gene Selection from Gene Indexing?Projects

    Science.gov (United States)

    Burke, John; Wang, Hui; Hide, Winston; Davison, Daniel B.

    1998-01-01

    Several efforts are under way to partition single-read expressed sequence tag (EST), as well as full-length transcript data, into large-scale gene indices, where transcripts are in common index classes if and only if they share a common progenitor gene. Accurate gene indexing facilitates gene expression studies, as well as inexpensive and early gene sequence discovery through assembly of ESTs that are derived from genes that have not been sequenced by classical methods. We extend, correct, and enhance the information obtained from index groups by splitting index classes into subclasses based on sequence dissimilarity (diversity). Two applications of this are highlighted in this report. First it is shown that our method can ameliorate the damage that artifacts, such as chimerism, inflict on index integrity. Additionally, we demonstrate how the organization imposed by an effective subpartition can greatly increase the sensitivity of gene expression studies by accounting for the existence and tissue- or pathology-specific regulation of novel gene isoforms and polymorphisms. We apply our subpartitioning treatment to the UniGene gene indexing project to measure a marked increase in information quality and abundance (in terms of assembly length and insertion/deletion error) after treatment and demonstrate cases where new levels of information concerning differential expression of alternate gene forms, such as regulated alternative splicing, are discovered. [Tables 2 and 3 can be viewed in their entirety as Online Supplements at http://www.genome.org.] PMID:9521931

  13. Many Ribosomal Protein Genes Are Cancer Genes in Zebrafish

    Directory of Open Access Journals (Sweden)

    Amsterdam Adam

    2004-01-01

    Full Text Available We have generated several hundred lines of zebrafish (Danio rerio, each heterozygous for a recessive embryonic lethal mutation. Since many tumor suppressor genes are recessive lethals, we screened our colony for lines that display early mortality and/or gross evidence of tumors. We identified 12 lines with elevated cancer incidence. Fish from these lines develop malignant peripheral nerve sheath tumors, and in some cases also other tumor types, with moderate to very high frequencies. Surprisingly, 11 of the 12 lines were each heterozygous for a mutation in a different ribosomal protein (RP gene, while one line was heterozygous for a mutation in a zebrafish paralog of the human and mouse tumor suppressor gene, neurofibromatosis type 2. Our findings suggest that many RP genes may act as haploinsufficient tumor suppressors in fish. Many RP genes might also be cancer genes in humans, where their role in tumorigenesis could easily have escaped detection up to now.

  14. Using the Gene Ontology Hierarchy when Predicting Gene Function

    CERN Document Server

    Mostafavi, Sara

    2012-01-01

    The problem of multilabel classification when the labels are related through a hierarchical categorization scheme occurs in many application domains such as computational biology. For example, this problem arises naturally when trying to automatically assign gene function using a controlled vocabularies like Gene Ontology. However, most existing approaches for predicting gene functions solve independent classification problems to predict genes that are involved in a given function category, independently of the rest. Here, we propose two simple methods for incorporating information about the hierarchical nature of the categorization scheme. In the first method, we use information about a gene's previous annotation to set an initial prior on its label. In a second approach, we extend a graph-based semi-supervised learning algorithm for predicting gene function in a hierarchy. We show that we can efficiently solve this problem by solving a linear system of equations. We compare these approaches with a previous ...

  15. Human AZU-1 gene, variants thereof and expressed gene products

    Science.gov (United States)

    Chen, Huei-Mei; Bissell, Mina

    2004-06-22

    A human AZU-1 gene, mutants, variants and fragments thereof. Protein products encoded by the AZU-1 gene and homologs encoded by the variants of AZU-1 gene acting as tumor suppressors or markers of malignancy progression and tumorigenicity reversion. Identification, isolation and characterization of AZU-1 and AZU-2 genes localized to a tumor suppressive locus at chromosome 10q26, highly expressed in nonmalignant and premalignant cells derived from a human breast tumor progression model. A recombinant full length protein sequences encoded by the AZU-1 gene and nucleotide sequences of AZU-1 and AZU-2 genes and variant and fragments thereof. Monoclonal or polyclonal antibodies specific to AZU-1, AZU-2 encoded protein and to AZU-1, or AZU-2 encoded protein homologs.

  16. Gene expression analysis identifies global gene dosage sensitivity in cancer

    DEFF Research Database (Denmark)

    Fehrmann, Rudolf S. N.; Karjalainen, Juha M.; Krajewska, Malgorzata; Westra, Harm-Jan; Maloney, David; Simeonov, Anton; Pers, Tune Hannes; Hirschhorn, Joel N.; Jansen, Ritsert C.; Schultes, Erik A.; van Haagenl, Herman H. H. B. M.; de Vries, Elisabeth G. E.; Meerman, Gerard J. te; Wijmenga, Cisca; van Vugt, Marcel A. T. M.; Franke, Lude

    2015-01-01

    Many cancer-associated somatic copy number alterations (SCNAs) are known. Currently, one of the challenges is to identify the molecular downstream effects of these variants. Although several SCNAs are known to change gene expression levels, it is not clear whether each individual SCNA affects gene expression. We reanalyzed 77,840 expression profiles and observed a limited set of 'transcriptional components' that describe well-known biology, explain the vast majority of variation in gene expressi...

  17. Gene Function Prediction Based on the Gene Ontology Hierarchical Structure

    OpenAIRE

    Cheng, Liangxi; Lin, Hongfei; Hu, Yuncui; Wang, Jian; Yang, Zhihao

    2014-01-01

    The information of the Gene Ontology annotation is helpful in the explanation of life science phenomena, and can provide great support for the research of the biomedical field. The use of the Gene Ontology is gradually affecting the way people store and understand bioinformatic data. To facilitate the prediction of gene functions with the aid of text mining methods and existing resources, we transform it into a multi-label top-down classification problem and develop a method that uses the hie...

  18. Gene therapy for Primary Immunodeficiencies: looking ahead, towards gene correction

    OpenAIRE

    Pessach, Itai M.; Luigi D. Notarangelo

    2011-01-01

    Allogeneic hematopoietic stem cell transplantation is the treatment of choice for severe primary immunodeficiencies (PIDs). For patients lacking an HLA-identical donor, gene therapy is an attractive therapeutic option. Approaches based on insertion of a functional gene using viral vectors have provided proof of concept for the ability of gene therapy to cure PIDs. However, leukemic transformation due to insertional mutagenesis has been observed, prompting development of novel approaches based...

  19. Identifying Driver Genes in Cancer by Triangulating Gene Expression, Gene Location, and Survival Data

    Science.gov (United States)

    Rouam, Sigrid; Miller, Lance D; Karuturi, R Krishna Murthy

    2014-01-01

    Driver genes are directly responsible for oncogenesis and identifying them is essential in order to fully understand the mechanisms of cancer. However, it is difficult to delineate them from the larger pool of genes that are deregulated in cancer (ie, passenger genes). In order to address this problem, we developed an approach called TRIAngulating Gene Expression (TRIAGE through clinico-genomic intersects). Here, we present a refinement of this approach incorporating a new scoring methodology to identify putative driver genes that are deregulated in cancer. TRIAGE triangulates – or integrates – three levels of information: gene expression, gene location, and patient survival. First, TRIAGE identifies regions of deregulated expression (ie, expression footprints) by deriving a newly established measure called the Local Singular Value Decomposition (LSVD) score for each locus. Driver genes are then distinguished from passenger genes using dual survival analyses. Incorporating measurements of gene expression and weighting them according to the LSVD weight of each tumor, these analyses are performed using the genes located in significant expression footprints. Here, we first use simulated data to characterize the newly established LSVD score. We then present the results of our application of this refined version of TRIAGE to gene expression data from five cancer types. This refined version of TRIAGE not only allowed us to identify known prominent driver genes, such as MMP1, IL8, and COL1A2, but it also led us to identify several novel ones. These results illustrate that TRIAGE complements existing tools, allows for the identification of genes that drive cancer and could perhaps elucidate potential future targets of novel anticancer therapeutics. PMID:25949096

  20. Mapping of repair genes

    International Nuclear Information System (INIS)

    Chromosome mapping of repair genes involved in U.V. sensitivity is reported. Twenty-three of 25 hybrid cells were resistant to U.V. light. Survival curves of 2 U.V.-resistant cell strains, which possessed mouse chromosomes and human chromosome No.7 - 16, were similar to those of wild strain (L5178Y). On the other hand, survival curves of U.V.-sensitive hybrid cells was analogous to those of Q31. There was a definitive difference in the frequency of inducible chromosome aberrations between U.V. resistant and sensitive mouse-human hybrid cells. U.V.-resistant cell strains possessed the ability of excision repair. Analysis of karyotype in hybrid cells showed that the difference in U.V. sensitivity is dependent upon whether or not human chromosome No.13 is present. Synteny test on esterase D-determining locus confirmed that there is an agreement between the presence of chromosome No.13 and the presence of human esterase D activity. These results led to a conclusion that human genes which compensate recessive character of U.V.-sensitive mutant strain, Q31, with mouse-human hybrid cells are located on the locus of chromosome No.13. (Namekawa, K.)

  1. Identification of significant periodic genes in microarray gene expression data

    Directory of Open Access Journals (Sweden)

    Chen Jie

    2005-11-01

    Full Text Available Abstract Background One frequent application of microarray experiments is in the study of monitoring gene activities in a cell during cell cycle or cell division. A new challenge for analyzing the microarray experiments is to identify genes that are statistically significantly periodically expressed during the cell cycle. Such a challenge occurs due to the large number of genes that are simultaneously measured, a moderate to small number of measurements per gene taken at different time points, and high levels of non-normal random noises inherited in the data. Results Based on two statistical hypothesis testing methods for identifying periodic time series, a novel statistical inference approach, the C&G procedure, is proposed to effectively screen out statistically significantly periodically expressed genes. The approach is then applied to yeast and bacterial cell cycle gene expression data sets, as well as to human fibroblasts and human cancer cell line data sets, and significantly periodically expressed genes are successfully identified. Conclusion The C&G procedure proposed is an effective method for identifying statistically significant periodic genes in microarray time series gene expression data.

  2. Molecular characterization of Mycobacterium kansasii isolates in the State of São Paulo between 1995-1998

    Directory of Open Access Journals (Sweden)

    Erica Chimara

    2004-11-01

    Full Text Available Mycobacterium kansasii is the most common cause of pulmonary nontuberculous mycobacteria infection and classical identification of this pathogen needs a time consuming phenotypic tests. Polymerase chain reaction-restriction fragment lenght polymorphism analysis (PRA of the gene enconding for the 65kDa heat shock (hsp65 protein offers an easy, rapid, and inexpensive procedure to identify and subtype M. kansasii isolates. In the present study, we performed a retrospective analysis of patients who had mycobacteria identified on the basis of phenotypic tests by means of a review of database at Mycobacteria Laboratory of the Instituto Adolfo Lutz in the period 1995-1998. A total of 9381 clinical isolates were analyzed of which 7777 (82.9% were identified as M. tuberculosis complex and 1604 (17.1% as nontuberculous mycobacteria. Of the 296 M. kansasii isolates, 189 (63.8% isolates obtained from 119 patients were viable and were analyzed by PRA-hsp65. Hundred eight two (98.9% were classified as M. kansasii type I. Two isolates were classified as type II and III and five isolates were characterized as other Mycobacterium species. Clinical isolates of M. kansasii in the state of São Paulo was almost exclusively subtype I regardless of HIV status.

  3. DNA-damage-inducible genes

    International Nuclear Information System (INIS)

    Ultraviolet (UV) lights, ionizing radiations and some chemical agents give rise to various kinds of DNA damages, such as pyrimidine dimers, DNA-strand scissions and base modification with bulky adducts. In response to the genotoxic stress caused by these DNA damages, a lot of mammalian genes are transcriptionally induced. Some of the induced genes have been identified to play important roles in cellular protection in association with DNA repair, G1/G2 checkpoint regulations or apoptosis. Ubiquitin, which has been revealed to be UV-inducible in cultured human cells (HeLa), has potential roles in cell cycle checkpoint activation and regulation of signal transduction pathway. In this article, we present the complete structure of a polyubiquitin gene CHUB2 isolated from the V79 Chinese hamster genome. The CHUB2 gene is characterized as a Chinese hamster equivalent to the human polyubiquitin gene UbC, which has been shown to be UV-inducible, because the nucleotide sequences in the 3' untranslated regions of both genes are highly homologous. Although the CHUB2 gene is not obviously induced by UV light, the structural characteristics in the 5' control region of the CHUB2 gene offers some hints concerning the human UbU gene regulation. In addition, we present a polymorphism which is attributable to the altered repeat number of the ubiquitin coding unit as has been similarly observed in the human UbC gene. The biological significance of this common feature to the CHUB2 gene and the human UbC gene will be discussed. (author)

  4. Hormones, genes, and?behavior

    OpenAIRE

    Pfaff, Donald W

    1997-01-01

    With assays of hormone-sensitive behaviors, it is possible to demonstrate both direct and indirect actions of genes on mammalian social behaviors. Direct effects of estrogen receptor gene expression and progesterone receptor gene expression figure prominently in well analyzed neuroendocrine mechanisms for sex behavior, operating through a neural circuit that has been delineated. Indirect effects, notably the consequences of sexual differentiation, display complex d...

  5. Gene Ontology Consortium: going forward

    OpenAIRE

    2014-01-01

    The Gene Ontology (GO; http://www.geneontology.org) is a community-based bioinformatics resource that supplies information about gene product function using ontologies to represent biological knowledge. Here we describe improvements and expansions to several branches of the ontology, as well as updates that have allowed us to more efficiently disseminate the GO and capture feedback from the research community. The Gene Ontology Consortium (GOC) has expanded areas of the ontology such as cilia...

  6. Why genes overlap in viruses

    OpenAIRE

    Chirico, Nicola; Vianelli, Alberto; Belshaw, Robert

    2010-01-01

    The genomes of most virus species have overlapping genes—two or more proteins coded for by the same nucleotide sequence. Several explanations have been proposed for the evolution of this phenomenon, and we test these by comparing the amount of gene overlap in all known virus species. We conclude that gene overlap is unlikely to have evolved as a way of compressing the genome in response to the harmful effect of mutation because RNA viruses, despite having generally higher mutation rates, have...

  7. Homeobox gene expression in Brachiopoda

    DEFF Research Database (Denmark)

    Altenburger, Andreas; Martinez, Pedro; Wanninger, Andreas

    2011-01-01

    The molecular control that underlies brachiopod ontogeny is largely unknown. In order to contribute to this issue we analyzed the expression pattern of two homeobox containing genes, Not and Cdx, during development of the rhynchonelliform (i.e., articulate) brachiopod Terebratalia transversa. Not is a homeobox containing gene that regulates the formation of the notochord in chordates, while Cdx (caudal) is a ParaHox gene involved in the formation of posterior tissues of various animal phyla. The...

  8. Gene therapy in pancreatic cancer

    OpenAIRE

    Liu, Si-Xue; Xia, Zhong-Sheng; Zhong, Ying-Qiang

    2014-01-01

    Pancreatic cancer (PC) is a highly lethal disease and notoriously difficult to treat. Only a small proportion of PC patients are eligible for surgical resection, whilst conventional chemoradiotherapy only has a modest effect with substantial toxicity. Gene therapy has become a new widely investigated therapeutic approach for PC. This article reviews the basic rationale, gene delivery methods, therapeutic targets and developments of laboratory research and clinical trials in gene therapy of PC...

  9. Gene set analysis for GWAS

    DEFF Research Database (Denmark)

    Debrabant, Birgit; Soerensen, Mette

    2014-01-01

    Abstract We discuss the use of modified Kolmogorov-Smirnov (KS) statistics in the context of gene set analysis and review corresponding null and alternative hypotheses. Especially, we show that, when enhancing the impact of highly significant genes in the calculation of the test statistic, the...... parameter and the genesis and distribution of the gene-level statistics, and illustrate the effects of differential weighting in a real-life example....

  10. Gene therapy: here to stay.

    OpenAIRE

    Dubé, I D; Cournoyer, D

    1995-01-01

    Advances in biotechnology have brought gene therapy to the forefront of medical research. The feasibility of gene transfer was first demonstrated in experiments using tumour viruses. This led to the development of a variety of viral and nonviral methods for the genetic modification of somatic cells. Two main approaches emerged: in-vivo modification, in which gene transfer vehicles are delivered directly into patients, and ex-vivo manipulation, in which cells from the patient are grown in cult...

  11. Detecting multivariate differentially expressed genes

    OpenAIRE

    Tegnér Jesper; Björkegren Johan; Peña José M; Nilsson Roland

    2007-01-01

    Abstract Background Gene expression is governed by complex networks, and differences in expression patterns between distinct biological conditions may therefore be complex and multivariate in nature. Yet, current statistical methods for detecting differential expression merely consider the univariate difference in expression level of each gene in isolation, thus potentially neglecting many genes of biological importance. Results We have developed a novel algorithm for detecting multivariate e...

  12. Discovering modulators of gene expression

    OpenAIRE

    Babur, Özgün; Demir, Emek; Gönen, Mithat; Sander, Chris; Dogrusoz, Ugur

    2010-01-01

    Proteins that modulate the activity of transcription factors, often called modulators, play a critical role in creating tissue- and context-specific gene expression responses to the signals cells receive. GEM (Gene Expression Modulation) is a probabilistic framework that predicts modulators, their affected targets and mode of action by combining gene expression profiles, protein–protein interactions and transcription factor–target relationships. Using GEM, we correctly predicted a significant...

  13. Delivery systems for gene therapy

    OpenAIRE

    Mali, Shrikant

    2013-01-01

    The structure of DNA was unraveled by Watson and Crick in 1953, and two decades later Arber, Nathans and Smith discovered DNA restriction enzymes, which led to the rapid growth in the field of recombinant DNA technology. From expressing cloned genes in bacteria to expressing foreign DNA in transgenic animals, DNA is now slated to be used as a therapeutic agent to replace defective genes in patients suffering from genetic disorders or to kill tumor cells in cancer patients. Gene therapy provid...

  14. Gene function diversification upon duplication

    OpenAIRE

    Gonzalez, Luís Manuel Baudouin, 1988-

    2013-01-01

    A duplicação de genes é um processo molecular que facilita a evolução de novos genes com novas funções. Isto acontece porque a presença de uma segunda cópia relaxa a pressão selectiva sobre estes genes, permintindo que se acumulem mutações nestes. A pseudogenização de uma das cópias é o destino mais provável após um evento de duplicação, assumindo que a fixação dá-se através de selecção neutral (o que é normalmente assumido), mas outros resultados podem ocorrer: ambas as cópias podem manter a...

  15. Discovering modulators of gene expression.

    Science.gov (United States)

    Babur, Ozgün; Demir, Emek; Gönen, Mithat; Sander, Chris; Dogrusoz, Ugur

    2010-09-01

    Proteins that modulate the activity of transcription factors, often called modulators, play a critical role in creating tissue- and context-specific gene expression responses to the signals cells receive. GEM (Gene Expression Modulation) is a probabilistic framework that predicts modulators, their affected targets and mode of action by combining gene expression profiles, protein-protein interactions and transcription factor-target relationships. Using GEM, we correctly predicted a significant number of androgen receptor modulators and observed that most modulators can both act as co-activators and co-repressors for different target genes. PMID:20466809

  16. Cytokines in mycobacterial infections: `in vitro` and `ex vivo` studies

    Energy Technology Data Exchange (ETDEWEB)

    Flad, H.D.; Gercken, J.; Huebner, L.; Schlueter, C.; Ernst, M. [Forschungsinstitut Borstel (Germany). Inst. fuer Experimentelle Biologie und Medizin; Pryjma, J. [Uniwersytet Jagiellonski, Cracow (Poland)

    1995-12-31

    Different species of mycobacteria differ in their capacity to induce the production of tumor necrosis factor-{alpha} (TNF-{alpha}) by human monocytes `in vitro`. Whereas `M. tuberculosis` is a potent inducer of TNF-{alpha}, `M. leprae` is much less potent. TNF-{alpha} production is found to be associated with the availability of H{sub 2}O{sub 2} generated by activated monocytes, as superoxide enhancing H{sub 2}O{sub 2} concentration increases and catalase degrading H{sub 2}O{sub 2} decreases TNF-{alpha} production. Furthermore, `M. kansasii` with high intrinsic catalase induce less TNF-{alpha} than mycobacteria with low intrinsic catalase. `In vitro` infection of monocytes with `M. tuberculosis` leads to an impairment of the antigen-presenting capacity, as determined by a reduction of antigen-induced T cell proliferation and interferon {gamma} (IFN-{gamma}) production. Of crucial importance in this impairment is the `M. tuberculosis`-induced down-modulation of MHC class II antigens. The role of TNF-{alpha} `in vivo` is reflected in patients with various forms of leprosy. In skin lesions of lepromatous leprosy patients TNF-{alpha}, interleukin 1{beta} (IL-1{beta}), and IFN-{gamma} production are found to be rare, whereas these cytokines are well expressed in skin lesions of patients with tuberculoid leprosy. After multidrug chemotherapy an increase of local cytokine production is found. Taken together, these findings suggest that components of mycobacteria may interfere with local cell-mediated immune reactions `in vivo`. The molecular mechanisms involved in these local responses need to be defined. (author). 10 refs, 3 figs, 5 tabs.

  17. Mycobacterial species as case-study of comparative genome analysis

    DEFF Research Database (Denmark)

    Zakham, F.; Belayachi, L.; Ussery, David; Akrim, M.; Benjouad, A.; El Aouad, R.; Ennaji, M. M.

    2011-01-01

    . 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...

  18. Cytokines in mycobacterial infections: 'in vitro' and 'ex vivo' studies

    International Nuclear Information System (INIS)

    Different species of mycobacteria differ in their capacity to induce the production of tumor necrosis factor-? (TNF-?) by human monocytes 'in vitro'. Whereas 'M. tuberculosis' is a potent inducer of TNF-?, 'M. leprae' is much less potent. TNF-? production is found to be associated with the availability of H2O2 generated by activated monocytes, as superoxide enhancing H2O2 concentration increases and catalase degrading H2O2 decreases TNF-? production. Furthermore, 'M. kansasii' with high intrinsic catalase induce less TNF-? than mycobacteria with low intrinsic catalase. 'In vitro' infection of monocytes with 'M. tuberculosis' leads to an impairment of the antigen-presenting capacity, as determined by a reduction of antigen-induced T cell proliferation and interferon ? (IFN-?) production. Of crucial importance in this impairment is the 'M. tuberculosis'-induced down-modulation of MHC class II antigens. The role of TNF-? 'in vivo' is reflected in patients with various forms of leprosy. In skin lesions of lepromatous leprosy patients TNF-?, interleukin 1? (IL-1?), and IFN-? production are found to be rare, whereas these cytokines are well expressed in skin lesions of patients with tuberculoid leprosy. After multidrug chemotherapy an increase of local cytokine production is found. Taken together, these findings suggest that components of mycobacteria may interfere with local cell-mediated immune reactions 'in vivo'. The molecular mechanisms involved in these local responses need to be defined. (author). 10 refs, 3 figs, 5 tabs

  19. Mycobacterial Pulmonary Infections in Patients with Idiopathic Pulmonary Fibrosis

    OpenAIRE

    Park, Sung-Woo; Song, Jin Woo; Shim, Tae Sun; Park, Moo-Suk; Lee, Hong-Lyeol; Uh, Soo-taek; Park, Choon-Sik; Kim, Dong Soon

    2012-01-01

    Patients with idiopathic pulmonary fibrosis (IPF) have an increased risk for developing tuberculosis (TB). However, no studies have been reported regarding the development of nontuberculous mycobacterium (NTM) lung disease (NTMLD). We reviewed 795 patients with IPF from five university hospitals who were diagnosed by histological or radio-clinical criteria. In the 795 patients with IPF, pulmonary infections with mycobacterium tuberculosis (MTB) and NTM were found in 35 (4.4%) and 16 patients ...

  20. An experimental model of mycobacterial infection under corneal flaps

    Directory of Open Access Journals (Sweden)

    C.B.D. Adan

    2004-07-01

    Full Text Available In order to develop a new experimental animal model of infection with Mycobacterium chelonae in keratomileusis, we conducted a double-blind prospective study on 24 adult male New Zealand rabbits. One eye of each rabbit was submitted to automatic lamellar keratotomy with the automatic corneal shaper under general anesthesia. Eyes were immunosuppressed by a single local injection of methyl prednisolone. Twelve animals were inoculated into the keratomileusis interface with 1 µl of 10(6 heat-inactivated bacteria (heat-inactivated inoculum controls and 12 with 1 µl of 10(6 live bacteria. Trimethoprim drops (0.1%, w/v were used as prophylaxis for the surgical procedure every 4 h (50 µl, qid. Animals were examined by 2 observers under a slit lamp on the 1st, 3rd, 5th, 7th, 11th, 16th, and 23rd postoperative days. Slit lamp photographs were taken to document clinical signs. Animals were sacrificed when corneal disease was detected and corneal samples were taken for microbiological analysis. Eleven of 12 experimental rabbits developed corneal disease, and M. chelonae could be isolated from nine rabbits. Eleven of the 12 controls receiving a heat-inactivated inoculum did not develop corneal disease. M. chelonae was not isolated from any of the control rabbits receiving a heat-inactivated inoculum, or from the healthy cornea of control rabbits. Corneal infection by M. chelonae was successfully induced in rabbits submitted to keratomileusis. To our knowledge, this is the first animal model of M. chelonae infection following corneal flaps for refractive surgery to be described in the literature and can be used for the analysis of therapeutic responses.

  1. An experimental model of mycobacterial infection under corneal flaps

    Scientific Electronic Library Online (English)

    C.B.D., Adan; E.H., Sato; L.B., Sousa; R.S., Oliveira; S.C., Leão; D., Freitas.

    2004-07-01

    Full Text Available In order to develop a new experimental animal model of infection with Mycobacterium chelonae in keratomileusis, we conducted a double-blind prospective study on 24 adult male New Zealand rabbits. One eye of each rabbit was submitted to automatic lamellar keratotomy with the automatic corneal shaper [...] under general anesthesia. Eyes were immunosuppressed by a single local injection of methyl prednisolone. Twelve animals were inoculated into the keratomileusis interface with 1 µl of 10(6) heat-inactivated bacteria (heat-inactivated inoculum controls) and 12 with 1 µl of 10(6) live bacteria. Trimethoprim drops (0.1%, w/v) were used as prophylaxis for the surgical procedure every 4 h (50 µl, qid). Animals were examined by 2 observers under a slit lamp on the 1st, 3rd, 5th, 7th, 11th, 16th, and 23rd postoperative days. Slit lamp photographs were taken to document clinical signs. Animals were sacrificed when corneal disease was detected and corneal samples were taken for microbiological analysis. Eleven of 12 experimental rabbits developed corneal disease, and M. chelonae could be isolated from nine rabbits. Eleven of the 12 controls receiving a heat-inactivated inoculum did not develop corneal disease. M. chelonae was not isolated from any of the control rabbits receiving a heat-inactivated inoculum, or from the healthy cornea of control rabbits. Corneal infection by M. chelonae was successfully induced in rabbits submitted to keratomileusis. To our knowledge, this is the first animal model of M. chelonae infection following corneal flaps for refractive surgery to be described in the literature and can be used for the analysis of therapeutic responses.

  2. Synthesis of glycoconjugate fragments of mycobacterial phosphatidylinositol mannosides and lipomannan

    Directory of Open Access Journals (Sweden)

    Spencer J. Williams

    2011-03-01

    Full Text Available Mycobacterium tuberculosis, the causitive agent of tuberculosis (TB, possesses a complex cell wall containing mannose-rich glycophospholids termed phosphatidylinositol mannosides (PIMs, lipomannan (LM, and lipoarabinomannan (LAM. These glycophospholipids play important roles in cell wall function and host–pathogen interactions. Synthetic PIM/LM/LAM substructures are useful biochemical tools to delineate and dissect the fine details of mannose glycophospholipid biosynthesis and their interactions with host cells. We report the efficient synthesis of a series of azidooctyl di- and trimannosides possessing the following glycan structures: ?-Man-1,6-?-Man, ?-Man-1,6-?-Man-1,6-?-Man, ?-Man-1,2-?-Man-1,6-?-Man and 2,6-di-(?-Man-?-Man. The synthesis includes the use of non-benzyl protecting groups compatible with the azido group and preparation of the branched trisaccharide structure 2,6-di-(?-Man-?-Man through a double glycosylation of a 3,4-butanediacetal-protected mannoside. The azidooctyl groups of these synthetic mannans were elaborated to fluorescent glycoconjugates and squaric ester derivatives useful for further conjugation studies.

  3. Triazaspirodimethoxybenzoyls as Selective Inhibitors of Mycobacterial Lipoamide Dehydrogenase

    Energy Technology Data Exchange (ETDEWEB)

    Bryk, Ruslana; Arango, Nancy; Venugopal, Aditya; Warren, J. David; Park, Yun-Hee; Patel, Mulchand S.; Lima, Christopher D.; Nathan, Carl (Weill-Med); (SKI); (SUNYB)

    2010-06-25

    Mycobacterium tuberculosis (Mtb) remains the leading single cause of death from bacterial infection. Here we explored the possibility of species-selective inhibition of lipoamide dehydrogenase (Lpd), an enzyme central to Mtb's intermediary metabolism and antioxidant defense. High-throughput screening of combinatorial chemical libraries identified triazaspirodimethoxybenzoyls as high-nanomolar inhibitors of Mtb's Lpd that were noncompetitive versus NADH, NAD{sup +}, and lipoamide and >100-fold selective compared to human Lpd. Efficacy required the dimethoxy and dichlorophenyl groups. The structure of an Lpd-inhibitor complex was resolved to 2.42 {angstrom} by X-ray crystallography, revealing that the inhibitor occupied a pocket adjacent to the Lpd NADH/NAD{sup +} binding site. The inhibitor did not overlap with the adenosine moiety of NADH/NAD{sup +} but did overlap with positions predicted to bind the nicotinamide rings in NADH and NAD{sup +} complexes. The dimethoxy ring occupied a deep pocket adjacent to the FAD flavin ring where it would block coordination of the NADH nicotinamide ring, while the dichlorophenyl group occupied a more exposed pocket predicted to coordinate the NAD{sup +} nicotinamide. Several residues that are not conserved between the bacterial enzyme and its human homologue were predicted to contribute both to inhibitor binding and to species selectivity, as confirmed for three residues by analysis of the corresponding mutant Mtb Lpd proteins. Thus, nonconservation of residues lining the electron-transfer tunnel in Mtb Lpd can be exploited for development of species-selective Lpd inhibitors.

  4. Mycobacterial Lineages Causing Pulmonary and Extrapulmonary Tuberculosis, Ethiopia

    OpenAIRE

    Firdessa, Rebuma; Berg, Stefan; Hailu, Elena; Schelling, Esther; Gumi, Balako; Erenso, Girume; Gadisa, Endalamaw; Kiros, Teklu; Habtamu, Meseret; Hussein, Jemal; Zinsstag, Jakob; Robertson, Brian D; Ameni, Gobena; Lohan, Amanda J; Loftus, Brendan

    2013-01-01

    Molecular typing of 964 specimens from patients in Ethiopia with lymph node or pulmonary tuberculosis showed a similar distribution of Mycobacterium tuberculosis strains between the 2 disease manifestations and a minimal role for M. bovis. We report a novel phylogenetic lineage of M. tuberculosis strongly associated with the Horn of Africa.

  5. Triazaspirodimethoxybenzoyls as selective inhibitors of mycobacterial lipoamide dehydrogenase†#

    OpenAIRE

    Bryk, Ruslana; Arango, Nancy; Venugopal, Aditya; Warren, J. David; Park, Yun-Hee; PATEL, MULCHAND S; Lima, Christopher D.; Nathan, Carl

    2010-01-01

    Mycobacterium tuberculosis (Mtb) remains the leading single cause of death from bacterial infection. Here we explored the possibility of species-selective inhibition of lipoamide dehydrogenase (Lpd), an enzyme central to Mtb’s intermediary metabolism and antioxidant defense. High-throughput screening of combinatorial chemical libraries identified triazaspirodimethoxybenzoyls as high-nanomolar inhibitors of Mtb’s Lpd that were noncompetitive versus NADH, NAD+, and lipoamide and >100-fold selec...

  6. Benzimidazoles: novel mycobacterial gyrase inhibitors from scaffold morphing.

    Science.gov (United States)

    Hameed P, Shahul; Raichurkar, Anandkumar; Madhavapeddi, Prashanti; Menasinakai, Sreenivasaiah; Sharma, Sreevalli; Kaur, Parvinder; Nandishaiah, Radha; Panduga, Vijender; Reddy, Jitendar; Sambandamurthy, Vasan K; Sriram, D

    2014-07-10

    Type II topoisomerases are well conserved across the bacterial species, and inhibition of DNA gyrase by fluoroquinolones has provided an attractive option for treatment of tuberculosis (TB). However, the emergence of fluoroquinolone-resistant strains of Mycobacterium tuberculosis (Mtb) poses a threat for its sustainability. A scaffold hopping approach using the binding mode of novel bacterial topoisomerase inhibitors (NBTIs) led to the identification of a novel class of benzimidazoles as DNA gyrase inhibitors with potent anti-TB activity. Docking of benzimidazoles to a NBTI bound crystal structure suggested that this class of compound makes key contacts in the enzyme active site similar to the reported NBTIs. This observation was further confirmed through the measurement of DNA gyrase inhibition, and activity against Mtb strains harboring mutations that confer resistance to aminopiperidines based NBTIs and Mtb strains resistant to moxifloxacin. Structure-activity relationship modification at the C-7 position of the left-hand side ring provided further avenue to improve hERG selectivity for this chemical series that has been the major challenges for NBTIs. PMID:25050172

  7. Macrophage Activation by Ursolic and Oleanolic Acids during Mycobacterial Infection

    Directory of Open Access Journals (Sweden)

    Sonia López-García

    2015-08-01

    Full Text Available Oleanolic (OA and ursolic acids (UA are triterpenes that are abundant in vegetables, fruits and medicinal plants. They have been described as active moieties in medicinal plants used for the treatment of tuberculosis. In this study, we analyzed the effects of these triterpenes on macrophages infected in vitro with Mycobacterium tuberculosis (MTB. We evaluated production of nitric oxide (NO, reactive oxygen species (ROS, and cytokines (TNF-? and TGF-? as well as expression of cell membrane receptors (TGR5 and CD36 in MTB-infected macrophages following treatment with OA and UA. Triterpenes caused reduced MTB growth in macrophages, stimulated production of NO and ROS in the early phase, stimulated TNF-?, suppressed TGF-? and caused over-expression of CD36and TGR5 receptors. Thus, our data suggest immunomodulatory properties of OA and UA on MTB infected macrophages. In conclusion, antimycobacterial effects induced by these triterpenes may be attributable to the conversion of macrophages from stage M2 (alternatively activated to M1 (classically activated.

  8. [Japanese new guidelines for nontuberculous mycobacterial pulmonary disease].

    Science.gov (United States)

    Kurashima, Atsuyuki

    2010-02-01

    Three important statements for Japanese pulmonary nontuberculous mycobacteriosis (NTM) were published in 2008. The first one is a new diagnostic criteria for pulmonary NTM, which was organized in association with the task force for nontuberculous mycobacteriois of the Japanese Society for Tuberculosis and the section for infectious disease and tuberculosis of the Japanese Respiratory Society. The second is a treatment guideline for pulmonary nontuberculous mycobacteriosis also which was made by the same joint working. The third is a sugical treatment guideline for pulmonary nontuberculous mycobacteriosis. The reason for the task of immediate importance is the number of pulmonary Mycobacterium avium complex (MAC) disease keeps increasing in our country and the disease cannot be disregarded widely in municipal hospitals or clinics. The morbidity rate of pulmonary MAC disease is assumed to be about 3.5 in the north American area. A lot of European nations are presumed that do not reach 1.0. Most of Asian researchers reply to our E-mail questions with the recent increasing of pulmonary MAC disease. Japanese estimated morbidity rate of this disease seems to be over 6.0 in 2007. It has been not clarified why a lot of this disease cases are in particular in Japan. In this situation, a concise diagnostic criteria is required from even a doctor who is not respiratory medicine specialists. The diagnosis can be confirmed by twice culture from sputa or one culture in case of bronchoscopic examination regardless of the bacterial strain. Moreover, it is possible to correspond to wider varieties of radiographic findings than 2007 diagnostic criteria of the United States. This disease became possible to diagnose before the consciousness syndrome appeared by the advancement of today's excellent imaging technology and nuclear acid amplification method. Therefore, the diagnosis confirmation and the beginning of chemotherapy time has become separated. In 2008, on Japanese medical insurance, the prescription of two drugs has become possible officially for pulmonary NTM due to the efforts of many stakeholders. However, pulmonary NTM is a disease to obtain a constant improvement at last continuing combination chemotherapy for a long term. Three drugs regiment of CAM as a main axis, adding EB, RFP or RBT is now a de facto international standard. New Japanese guideline for treatment describes the adverse events by a long-term administering more in detail than the previous one. However, it is difficult to control only by an internal therapy. In case of a localized lesion, we have recommended an appropriate surgical treatment. But a surgery treatment without combination of chemotherapy could not achieve an enough result. A multidisciplinary approach is important. The guideline of surgical treatment that reflected these content was also published in 2008. PMID:20229821

  9. STATE-OF-THE-ART HUMAN GENE THERAPY: PART I. GENE DELIVERY TECHNOLOGIES

    OpenAIRE

    Wang, Dan; Gao, Guangping

    2014-01-01

    Safe and effective gene delivery is a prerequisite for successful gene therapy. In the early age of human gene therapy, setbacks due to problematic gene delivery vehicles plagued the exciting therapeutic outcome. However, gene delivery technologies rapidly evolved ever since. With the advancement of gene delivery techniques, gene therapy clinical trials surged during the past decade. As the first gene therapy product has obtained regulatory approval and reached clinic, human gene therapy fina...

  10. Troyer Syndrome - GeneReviews - NCBI Bookshelf [GeneReviews

    Lifescience Database Archive (English)

    Full Text Available p a.figpopup{display:inline !important} .bk_tt {font-family: monospace} .body-content h2 {border ... m MP, Ardinger HH, et al., editors. GeneReviews? [Internet ]. Seattle (WA): University of Washington, Seattle; ... 1993-2014. GeneReviews ? [Internet ]. Show details Pagon RA, Adam MP, Ardinger HH, et ...

  11. Cherubism - GeneReviews - NCBI Bookshelf [GeneReviews

    Lifescience Database Archive (English)

    Full Text Available p a.figpopup{display:inline !important} .bk_tt {font-family: monospace} .body-content h2 {border ... m MP, Ardinger HH, et al., editors. GeneReviews? [Internet ]. Seattle (WA): University of Washington, Seattle; ... 1993-2014. GeneReviews ? [Internet ]. Show details Pagon RA, Adam MP, Ardinger HH, et ...

  12. CARASIL - GeneReviews - NCBI Bookshelf [GeneReviews

    Lifescience Database Archive (English)

    Full Text Available p a.figpopup{display:inline !important} .bk_tt {font-family: monospace} .body-content h2 {border ... m MP, Ardinger HH, et al., editors. GeneReviews? [Internet ]. Seattle (WA): University of Washington, Seattle; ... 1993-2014. GeneReviews ? [Internet ]. Show details Pagon RA, Adam MP, Ardinger HH, et ...

  13. Char Syndrome - GeneReviews - NCBI Bookshelf [GeneReviews

    Lifescience Database Archive (English)

    Full Text Available p a.figpopup{display:inline !important} .bk_tt {font-family: monospace} .body-content h2 {border ... m MP, Ardinger HH, et al., editors. GeneReviews? [Internet ]. Seattle (WA): University of Washington, Seattle; ... 1993-2014. GeneReviews ? [Internet ]. Show details Pagon RA, Adam MP, Ardinger HH, et ...

  14. Are TMEM genes potential candidate genes for panic disorder?

    DEFF Research Database (Denmark)

    NO, Gregersen; Buttenschøn, Henriette Nørmølle; Hedemand, Anne; Dahl, Hans Atli; Kristensen, Ann Suhl; B, Clementsen; Woldbye, David Paul Drucker; Koefoed, Pernille; Erhardt, A; Kruse, T A; Wang, August Gabriel; Børglum, Anders; Mors, Ole

    2014-01-01

    We analysed single nucleotide polymorphisms in two transmembrane genes (TMEM98 and TMEM132E) in panic disorder (PD) patients and control individuals from the Faroe Islands, Denmark and Germany. The genes encode single-pass membrane proteins and are located within chromosome 17q11.2-q12, a...

  15. Gene therapy of cancer by vaccines carrying inserted immunostimulatory genes.

    Czech Academy of Sciences Publication Activity Database

    Bubeník, Jan

    2007-01-01

    Ro?. 53, ?. 3 (2007), s. 71-73. ISSN 0015-5500 Grant ostatní: EU-FP6 NoE Clinigene(XE) 018933; Liga proti rakovin?, Praha(CZ) XX Institutional research plan: CEZ:AV0Z50520514 Keywords : gene therapy * immunostimulatory genes * vaccine Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 0.596, year: 2007

  16. GENE MUTATIONS, GENETIC DISEASE AND PHARMACOGENETIC GENES DISORDER

    Directory of Open Access Journals (Sweden)

    Ishak

    2010-12-01

    Full Text Available Somatic cell mutation is able to create genetic variance in a cell population and can induce cancer and tumor when gene mutations took place at repressor gene in controlling cell cycles such as p53 gene. Whereas germline cell mutation can cause genetic disease such as sickle cell anemia, breast cancer, thalassemia, parkinson’s as well as defect of biochemical pathway that influence drug-receptor interaction, which has negative effect and lead to hospitalized of patient. Most of reports mentioned that point mutation such as a single base of nucleotide substitution (purine replaced by purine or transversion (purine replaced by pyrimidine or vice versa that affected genetic disease as well as adverse drug reaction that involved genetic factors. Mutation that occurred in germline cell would be inherited to the progeny, and these mutated genes can spread in a population through fertilization process. Mutation that occur in coding frame of DNA region which of their expression are responsible for synthesis of specific products could be rise of genetic disease, because the lost of gene function. Similarly, mutation that take place for CYP450 gene family which related to drug metabolism included pharmacokinetic and pharmacodynamic gene function could affect drug biosynthesis and degradation. Abnormality of drug metabolism that results in pharmacogenetic effect which is indicated by adverse drug reaction to individual that severe metabolite defect. On the future, therapy of genetic disease as well as abnormal of drug metabolism can be directed into gene therapy techniques with using stem cell engineering.

  17. Gene Express Inc.

    Science.gov (United States)

    Saccomanno, Colette F

    2006-07-01

    Gene Express, Inc. is a technology-licensing company and provider of Standardized Reverse Transcription Polymerase Chain Reaction (StaRT-PCR) services. Designed by and for clinical researchers involved in pharmaceutical, biomarker and molecular diagnostic product development, StaRT-PCR is a unique quantitative and standardized multigene expression measurement platform. StaRT-PCR meets all of the performance characteristics defined by the US FDA as required to support regulatory submissions [101,102] , and by the Clinical Laboratory Improvement Act of 1988 (CLIA) as necessary to support diagnostic testing [1] . A standardized mixture of internal standards (SMIS), manufactured in bulk, provides integrated quality control wherein each native template target gene is measured relative to a competitive template internal standard. Bulk production enables the compilation of a comprehensive standardized database from across multiple experiments, across collaborating laboratories and across the entire clinical development lifecycle of a given compound or diagnostic product. For the first time, all these data are able to be directly compared. Access to such a database can dramatically shorten the time from investigational new drug (IND) to new drug application (NDA), or save time and money by hastening a substantiated 'no-go' decision. High-throughput StaRT-PCR is conducted at the company's automated Standardized Expression Measurement (SEM) Center. Currently optimized for detection on a microcapillary electrophoretic platform, StaRT-PCR products also may be analyzed on microarray, high-performance liquid chromatography (HPLC), or matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) platforms. SEM Center services deliver standardized genomic data--data that will accelerate the application of pharmacogenomic technology to new drug and diagnostic test development and facilitate personalized medicine. PMID:16886903

  18. Classifying genes to the correct Gene Ontology Slim term in Saccharomyces cerevisiae using neighbouring genes with classification learning

    Directory of Open Access Journals (Sweden)

    Tsatsoulis Costas

    2010-05-01

    Full Text Available Abstract Background There is increasing evidence that gene location and surrounding genes influence the functionality of genes in the eukaryotic genome. Knowing the Gene Ontology Slim terms associated with a gene gives us insight into a gene's functionality by informing us how its gene product behaves in a cellular context using three different ontologies: molecular function, biological process, and cellular component. In this study, we analyzed if we could classify a gene in Saccharomyces cerevisiae to its correct Gene Ontology Slim term using information about its location in the genome and information from its nearest-neighbouring genes using classification learning. Results We performed experiments to establish that the MultiBoostAB algorithm using the J48 classifier could correctly classify Gene Ontology Slim terms of a gene given information regarding the gene's location and information from its nearest-neighbouring genes for training. Different neighbourhood sizes were examined to determine how many nearest neighbours should be included around each gene to provide better classification rules. Our results show that by just incorporating neighbour information from each gene's two-nearest neighbours, the percentage of correctly classified genes to their correct Gene Ontology Slim term for each ontology reaches over 80% with high accuracy (reflected in F-measures over 0.80 of the classification rules produced. Conclusions We confirmed that in classifying genes to their correct Gene Ontology Slim term, the inclusion of neighbour information from those genes is beneficial. Knowing the location of a gene and the Gene Ontology Slim information from neighbouring genes gives us insight into that gene's functionality. This benefit is seen by just including information from a gene's two-nearest neighbouring genes.

  19. Variants in toll-like receptor 9 gene influence susceptibility to tuberculosis in a Mexican population

    OpenAIRE

    Torres-García, Diana; Cruz-Lagunas, Alfredo; García-Sancho Figueroa, Ma Cecilia; Fernández-Plata, Rosario; Baez-Saldaña, Renata; Mendoza-Milla, Criselda; Barquera, Rodrigo; Carrera-Eusebio, Aida; Ramírez-Bravo, Salomón; Campos, Lizeth; Angeles, Javier; Vargas-Alarcón, Gilberto; Granados, Julio; Gopal, Radha; Khader, Shabaana A

    2013-01-01

    Background: The control of Mycobacterium tuberculosis (Mtb) infection begins with the recognition of mycobacterial structural components by toll like receptors (TLRs) and other pattern recognition receptors. Our objective was to determine the influence of TLRs polymorphisms in the susceptibility to develop tuberculosis (TB) in Amerindian individuals from a rural area of Oaxaca, Mexico with high TB incidence. Methods: We carried out a case–control association community based study, genotyping ...

  20. Appearance frequency modulated gene set enrichment testing

    OpenAIRE

    Sartor Maureen A; Ma Jun; Jagadish HV

    2011-01-01

    Abstract Background Gene set enrichment testing has helped bridge the gap from an individual gene to a systems biology interpretation of microarray data. Although gene sets are defined a priori based on biological knowledge, current methods for gene set enrichment testing treat all genes equal. It is well-known that some genes, such as those responsible for housekeeping functions, appear in many pathways, whereas other genes are more specialized and play a unique role in a single pathway. Dra...

  1. Advancement and prospects of tumor gene therapy

    OpenAIRE

    Zhang, Chao; Wang, Qing-Tao; Liu, He; Zhang, Zhen-Zhu; Huang, Wen-Lin

    2011-01-01

    Gene therapy is one of the most attractive fields in tumor therapy. In past decades, significant progress has been achieved. Various approaches, such as viral and non-viral vectors and physical methods, have been developed to make gene delivery safer and more efficient. Several therapeutic strategies have evolved, including gene-based (tumor suppressor genes, suicide genes, antiangiogenic genes, cytokine and oxidative stress-based genes) and RNA-based (antisense oligonucleotides and RNA inter...

  2. The multiple gene duplication problem revisited

    OpenAIRE

    Bansal, Mukul S.; Eulenstein, Oliver

    2008-01-01

    Motivation: Deciphering the location of gene duplications and multiple gene duplication episodes on the Tree of Life is fundamental to understanding the way gene families and genomes evolve. The multiple gene duplication problem provides a framework for placing gene duplication events onto nodes of a given species tree, and detecting episodes of multiple gene duplication. One version of the multiple gene duplication problem was defined by Guigó et al. in 1996. Several heuristic solutions have...

  3. Evidence Based Selection of Housekeeping Genes

    OpenAIRE

    de Jonge, Hendrik J.M.; Fehrmann, Rudolf S. N.; de Bont, Eveline S. J. M.; Hofstra, Robert M. W.; Gerbens, Frans; Kamps, Willem A.; de Vries, Elisabeth G. E; VAN DER ZEE, ATE G.J.; te Meerman, Gerard J; ter Elst, Arja

    2007-01-01

    For accurate and reliable gene expression analysis, normalization of gene expression data against housekeeping genes (reference or internal control genes) is required. It is known that commonly used housekeeping genes (e.g. ACTB, GAPDH, HPRT1, and B2M) vary considerably under different experimental conditions and therefore their use for normalization is limited. We performed a meta-analysis of 13,629 human gene array samples in order to identify the most stable expressed genes. Here we show n...

  4. Integrating Gene Regulatory Networks to identify cancer-specific genes.

    Science.gov (United States)

    Bo, Valeria; Tucker, Allan

    2015-01-01

    Consensus approaches have been widely used to identify Gene Regulatory Networks (GRNs) that are common to multiple studies. However, in this research we develop an application that semi-automatically identifies key mechanisms that are specific to a particular set of conditions. We analyse four different types of cancer to identify gene pathways unique to each of them. To support the results reliability we calculate the prediction accuracy of each gene for the specified conditions and compare to predictions on other conditions. The most predictive are validated using the GeneCards encyclopaedia1 coupled with a statistical test for validating clusters. Finally, we implement an interface that allows the user to identify unique subnetworks of any selected combination of studies using AND & NOT logic operators. Results show that unique genes and sub-networks can be reliably identified and that they reflect key mechanisms that are fundamental to the cancer types under study. PMID:26306224

  5. A Combinatorial Approach to Detecting Gene-Gene and Gene-Environment Interactions in Family Studies

    OpenAIRE

    Lou, Xiang-Yang; Chen, Guo-Bo; YAN, LEI; MA, JENNIE Z.; MANGOLD, JAMIE E.; Zhu, Jun; Elston, Robert C.; Li, Ming D

    2008-01-01

    Widespread multifactor interactions present a significant challenge in determining risk factors of complex diseases. Several combinatorial approaches, such as the multifactor dimensionality reduction (MDR) method, have emerged as a promising tool for better detecting gene-gene (G × G) and gene-environment (G × E) interactions. We recently developed a general combinatorial approach, namely the generalized multifactor dimensionality reduction (GMDR) method, which can entertain both qualitative ...

  6. Principal Interactions Analysis for Repeated Measures Data: Application to Gene-Gene, Gene-Environment Interactions

    OpenAIRE

    Mukherjee, Bhramar; Ko, Yi-An; VanderWeele, Tyler; Roy, Anindya; Park, Sung Kyun; Chen, Jinbo

    2012-01-01

    Many existing cohorts with longitudinal data on environmental exposures, occupational history, lifestyle/behavioral characteristics and health outcomes have collected genetic data in recent years. In this paper, we consider the problem of modeling gene-gene, gene-environment interactions with repeated measures data on a quantitative trait. We review possibilities of using classical models proposed by Tukey (1949) and Mandel (1961) using the cell means of a two-way classification array for suc...

  7. On meme--gene coevolution.

    Science.gov (United States)

    Bull, L; Holland, O; Blackmore, S

    2000-01-01

    In this article we examine the effects of the emergence of a new replicator, memes, on the evolution of a pre-existing replicator, genes. Using a version of the NKCS model we examine the effects of increasing the rate of meme evolution in relation to the rate of gene evolution, for various degrees of interdependence between the two replicators. That is, the effects of memes' (suggested) more rapid rate of evolution in comparison to that of genes is investigated using a tunable model of coevolution. It is found that, for almost any degree of interdependence between the two replicators, as the rate of meme evolution increases, a phase transition-like dynamic occurs under which memes have a significantly detrimental effect on the evolution of genes, quickly resulting in the cessation of effective gene evolution. Conversely, the memes experience a sharp increase in benefit from increasing their rate of evolution. We then examine the effects of enabling genes to reduce the percentage of gene-detrimental evolutionary steps taken by memes. Here a critical region emerges as the comparative rate of meme evolution increases, such that if genes cannot effectively select memes a high percentage of the time, they suffer from meme evolution as if they had almost no selective capability. PMID:11224917

  8. Translational selection on SHH genes

    Scientific Electronic Library Online (English)

    Mohammadreza, Hajjari; Behnaz, Saffar; Atefeh, Khoshnevisan.

    Full Text Available Codon usage bias has been observed in various organisms. In this study, the correlation between SHH genes expression in some tissues and codon usage features was analyzed by bioinformatics. We found that translational selection may act on compositional features of this set of genes. [...

  9. Gene expression in colorectal cancer

    DEFF Research Database (Denmark)

    Birkenkamp-Demtroder, Karin; Christensen, Lise Lotte; Olesen, Sanne Harder; Frederiksen, Casper M; Laiho, Päivi; Aaltonen, Lauri A; Laurberg, Søren; Sørensen, Flemming Brandt; Hagemann, Rikke; ØRntoft, Torben F

    2002-01-01

    Understanding molecular alterations in colorectal cancer (CRC) is needed to define new biomarkers and treatment targets. We used oligonucleotide microarrays to monitor gene expression of about 6,800 known genes and 35,000 expressed sequence tags (ESTs) on five pools (four to six samples in each...

  10. REVIEW ARTICLE ON GENE THERAPY

    Directory of Open Access Journals (Sweden)

    Patil P.M., Chaudhari P.D., Megha Sahu and Duragkar N.J.

    2012-03-01

    Full Text Available Gene therapy can be broadly defined as the transfer of genetic material to cure a disease or at least to improve the clinical status of a patient. One of the basic concepts of gene therapy is to transform viruses into genetic shuttles, which will deliver the gene of interest into the target cells. Based on the nature of the viral genome, these gene therapy vectors can be divided into RNA and DNA viral vectors. The majority of RNA virus-based vectors have been derived from simple retroviruses like murine leukemia virus. A major shortcoming of these vectors is that they are not able to transducer nondividing cells. This problem may be overcome by the use of novel retroviral vectors derived from lentiviruses, such as human immunodeficiency virus (HIV. The most commonly used DNA virus vectors are based on adenoviruses and adeno-associated viruses. An example of gene-knockout mediated gene therapy is the knockout of the human CCR5 gene in T-cells in order to control HIV infection[1] Although the available vector systems are able to deliver genes in vivo into cells, the ideal delivery vehicle has not been found. Thus, the present viral vectors should be used only with great caution in human beings and further progress in vector development is necessary.

  11. On the development of mycobacterial infections II. Changing mycobacterial infection spectrum in the Cologne area 1983-1993.

    Science.gov (United States)

    Schütt-Gerowitt, H

    1995-12-01

    The overall detection rate of mycobacteria in the Cologne area did not change between 1983 and 1993. But a significant shift of the rate of Mycobacterium tuberculosis to nontuberculous mycobacteria, especially Mycobacterium avium-intracellulare complex, could be observed. Whereas the portion of Mycobacterium tuberculosis fell during these years from about 90% to 50-70% the portion of Mycobacterium avium-intracellulare increased steadily from 0 to 23% of all mycobacteria positive patients. The portion of tuberculosis patients positive for HIV remained constantly at 5-7% until 1992. Whether the 10% in 1993 signalizes a definitive increase remains to be proven. HIV positive patients with the diagnosis mycobacteria show another development. Whereas Mycobacterium tuberculosis is isolated constantly from about 2-3% of all HIV positive patients, the Mycobacterium avium-intracellulare complex increased from 0 to 10%. As methodical factors play a special role in the diagnostic procedure of mycobacteria, we evaluated some of the newer techniques for our routine laboratory: The well-known high sensitivity and specificity of the DNA probes (Gen-probe system) was confirmed, and by the radiometric culture system, in fact, the ubiquitous mycobacteria were detected much better. Otherwise from one third of special materials Mycobacterium tuberculosis and Mycobacterium bovis grew only on the conventional solid media. The susceptibility testing of 925 Mycobacterium tuberculosis strains revealed a rate of 10% (95 strains) with resistance to one antituberculotic drug only and 3% (29 strains) with resistance to two or more drugs. Only 0.9% (8 strains) were resistant to isoniacid and rifampicin (multidrug-resistant strains) indicating that drug resistance of Mycobacterium tuberculosis is still no serious problem in our region. PMID:8825114

  12. Imaging after vascular gene therapy

    International Nuclear Information System (INIS)

    Targets for cardiovascular gene therapy currently include limiting restenosis after balloon angioplasty and stent placement, inhibiting vein bypass graft intimal hyperplasia/stenosis, therapeutic angiogenesis for cardiac and lower-limb ischemia, and prevention of thrombus formation. While catheter angiography is still standard method to follow-up vascular gene transfer, other modern imaging techniques, especially intravascular ultrasound (IVUS), magnetic resonance (MR), and positron emission tomography (PET) imaging provide complementary information about the therapeutic effect of vascular gene transfer in humans. Although molecular imaging of therapeutic gene expression in the vasculatures is still in its technical development phase, it has already offered basic medical science an extremely useful in vivo evaluation tool for non- or minimally invasive imaging of vascular gene therapy

  13. Delivery systems for gene therapy

    Directory of Open Access Journals (Sweden)

    Shrikant Mali

    2013-01-01

    Full Text Available The structure of DNA was unraveled by Watson and Crick in 1953, and two decades later Arber, Nathans and Smith discovered DNA restriction enzymes, which led to the rapid growth in the field of recombinant DNA technology. From expressing cloned genes in bacteria to expressing foreign DNA in transgenic animals, DNA is now slated to be used as a therapeutic agent to replace defective genes in patients suffering from genetic disorders or to kill tumor cells in cancer patients. Gene therapy provides modern medicine with new perspectives that were unthinkable two decades ago. Progress in molecular biology and especially, molecular medicine is now changing the basics of clinical medicine. A variety of viral and non-viral possibilities are available for basic and clinical research. This review summarizes the delivery routes and methods for gene transfer used in gene therapy.

  14. Function analysis of unknown genes

    DEFF Research Database (Denmark)

    Rogowska-Wrzesinska, A.

    2002-01-01

      This thesis entitled "Function analysis of unknown genes" presents the use of proteome analysis for the characterisation of yeast (Saccharomyces cerevisiae) genes and their products (proteins especially those of unknown function). This study illustrates that proteome analysis can be used to...... describe different aspects of molecular biology of the cell, to study changes that occur in the cell due to overexpression or deletion of a gene and to identify various protein modifications. The biological questions and the results of the described studies show the diversity of the information that can be...... characterising yeast genes and proteins. It reports the first global proteome database collecting 36 yeast single gene deletion mutants and selecting over 650 differences between analysed mutants and the wild type strain. The obtained results show that two-dimensional gel electrophoresis and mass spectrometry...

  15. Imaging after vascular gene therapy

    Energy Technology Data Exchange (ETDEWEB)

    Manninen, Hannu I. [Department of Clinical Radiology, Kuopio University Hospital, Puijonlaaksontie 2, FIN-70210 Kuopio (Finland)]. E-mail: Hannu.manninen@kuh.fi; Yang, Xiaoming [Department of Radiology, Johns Hopkins University School of Medicine, Baltimore (United States)

    2005-11-01

    Targets for cardiovascular gene therapy currently include limiting restenosis after balloon angioplasty and stent placement, inhibiting vein bypass graft intimal hyperplasia/stenosis, therapeutic angiogenesis for cardiac and lower-limb ischemia, and prevention of thrombus formation. While catheter angiography is still standard method to follow-up vascular gene transfer, other modern imaging techniques, especially intravascular ultrasound (IVUS), magnetic resonance (MR), and positron emission tomography (PET) imaging provide complementary information about the therapeutic effect of vascular gene transfer in humans. Although molecular imaging of therapeutic gene expression in the vasculatures is still in its technical development phase, it has already offered basic medical science an extremely useful in vivo evaluation tool for non- or minimally invasive imaging of vascular gene therapy.

  16. Gene expression profiling: can we identify the right target genes?

    Directory of Open Access Journals (Sweden)

    J. E. Loyd

    2008-12-01

    Full Text Available Gene expression profiling allows the simultaneous monitoring of the transcriptional behaviour of thousands of genes, which may potentially be involved in disease development. Several studies have been performed in idiopathic pulmonary fibrosis (IPF, which aim to define genetic links to the disease in an attempt to improve the current understanding of the underlying pathogenesis of the disease and target pathways for intervention. Expression profiling has shown a clear difference in gene expression between IPF and normal lung tissue, and has identified a wide range of candidate genes, including those known to encode for proteins involved in extracellular matrix formation and degradation, growth factors and chemokines. Recently, familial pulmonary fibrosis cohorts have been examined in an attempt to detect specific genetic mutations associated with IPF. To date, these studies have identified families in which IPF is associated with mutations in the gene encoding surfactant protein C, or with mutations in genes encoding components of telomerase. Although rare and clearly not responsible for the disease in all individuals, the nature of these mutations highlight the importance of the alveolar epithelium in disease pathogenesis and demonstrate the potential for gene expression profiling in helping to advance the current understanding of idiopathic pulmonary fibrosis.

  17. Gene expression analysis identifies global gene dosage sensitivity in cancer

    DEFF Research Database (Denmark)

    Fehrmann, Rudolf S. N.; Karjalainen, Juha M.

    2015-01-01

    Many cancer-associated somatic copy number alterations (SCNAs) are known. Currently, one of the challenges is to identify the molecular downstream effects of these variants. Although several SCNAs are known to change gene expression levels, it is not clear whether each individual SCNA affects gene expression. We reanalyzed 77,840 expression profiles and observed a limited set of 'transcriptional components' that describe well-known biology, explain the vast majority of variation in gene expression and enable us to predict the biological function of genes. On correcting expression profiles for these components, we observed that the residual expression levels (in 'functional genomic mRNA' profiling) correlated strongly with copy number. DNA copy number correlated positively with expression levels for 99% of all abundantly expressed human genes, indicating global gene dosage sensitivity. By applying this method to 16,172 patient-derived tumor samples, we replicated many loci with aberrant copy numbers and identified recurrently disrupted genes in genomically unstable cancers.

  18. Gene expression profiling in rheumatology.

    Science.gov (United States)

    van der Pouw Kraan, Tineke C T M; van Baarsen, Lisa G M; Rustenburg, François; Baltus, Belinda; Fero, Mike; Verweij, Cornelis L

    2007-01-01

    In the last decade, the analysis of gene expression in tissues and cells has evolved from the analysis of a selected set of genes to an efficient high throughput whole-genome screening approach of potentially all genes expressed. Development of sophisticated methodologies such as microarray technology allows an open-ended survey to identify comprehensively the fraction of genes that are differentially expressed between samples and that define the samples' unique biology. By a global analysis of the genes that are expressed in cells and tissues of an individual under different conditions and during disease, we can build up "gene expression profiles (signatures)" which characterize the dynamic functioning of the genome under pathophysiological conditions. This strategy also provides the means to subdivide patients that suffer from a complex heterogeneous disease into more homogeneous subgroups. Such discovery-based research identifies biological processes that may include new genes with unknown function or genes not previously known to be involved in this process. The latter category may hold surprises that sometimes urge us to redirect our thinking. We have used microarrays to disclose the heterogeneity of rheumatoid arthritis (RA) patients at the level of gene expression of the affected synovial tissues. Analysis of the expression profiles of synovial tissues from different patients with RA revealed considerable variability, resulting in the identification of at least two molecularly distinct forms of RA tissues. One is characterized by genes that indicate an active inflammatory infiltrate with high immunoglobulin production, whereas the other type shows little immune activation and instead shows a higher stromal cell activity. These results confirm the heterogeneous nature of RA and suggest the existence of distinct pathogenic mechanisms that contribute to RA. The differences in expression profiles provide opportunities to stratify patients for intervention therapies based on molecular criteria. PMID:17983157

  19. REGULATORY GENES IN CREATING FLOWER COLOR PATTERNS

    Science.gov (United States)

    Differences in structural gene expression are responsible for a wide range of responses from human cancer to patterned flowers. Gene silencing is one of the ways in which gene expression is controlled. We have developed a model system to study gene silencing using a gene silencing mutation in Petun...

  20. How to Link to GeneReviews - GeneReviews - NCBI Bookshelf [GeneReviews

    Lifescience Database Archive (English)

    Full Text Available p a.figpopup{display:inline !important} .bk_tt {font-family: monospace} .body-content h2 {border ... m MP, Ardinger HH, et al., editors. GeneReviews? [Internet ]. Seattle (WA): University of Washington, Seattle; ... 1993-2014. GeneReviews ? [Internet ]. Show details Pagon RA, Adam MP, Ardinger HH, et ...

  1. GeneReviews Personnel - GeneReviews?? - NCBI Bookshelf [GeneReviews

    Lifescience Database Archive (English)

    Full Text Available p a.figpopup{display:inline !important} .bk_tt {font-family: monospace} .body-content h2 {border ... MP, Ardinger HH, et al., editors. GeneReviews?? [Internet ]. Seattle (WA): University of Washington, Seattle; ... 1993-2014. GeneReviews ?? [Internet ]. Show details Pagon RA, Adam MP, Ardinger HH, et ...

  2. GeneReviews — Review Processes - GeneReviews® - NCBI Bookshelf [GeneReviews

    Lifescience Database Archive (English)

    Full Text Available p a.figpopup{display:inline !important} .bk_tt {font-family: monospace} .body-content h2 {border ... am MP, Ardinger HH, et al., editors. GeneReviews® [Internet ]. Seattle (WA): University of Washington, Seattle; ... 1993-2014. GeneReviews ® [Internet ]. Show details Pagon RA, Adam MP, Ardinger HH, et ...

  3. GeneReviews Copyright Notice and Usage Disclaimer - GeneReviews - NCBI Bookshelf [GeneReviews

    Lifescience Database Archive (English)

    Full Text Available p a.figpopup{display:inline !important} .bk_tt {font-family: monospace} .body-content h2 {border ... m MP, Ardinger HH, et al., editors. GeneReviews? [Internet ]. Seattle (WA): University of Washington, Seattle; ... 1993-2014. GeneReviews ? [Internet ]. Show details Pagon RA, Adam MP, Ardinger HH, et ...

  4. What's New in GeneReviews - GeneReviews - NCBI Bookshelf [GeneReviews

    Lifescience Database Archive (English)

    Full Text Available p a.figpopup{display:inline !important} .bk_tt {font-family: monospace} .body-content h2 {border ... m MP, Ardinger HH, et al., editors. GeneReviews? [Internet ]. Seattle (WA): University of Washington, Seattle; ... 1993-2014. GeneReviews ? [Internet ]. Show details Pagon RA, Adam MP, Ardinger HH, et ...

  5. For Prospective GeneReviews Authors - GeneReviews - NCBI Bookshelf [GeneReviews

    Lifescience Database Archive (English)

    Full Text Available p a.figpopup{display:inline !important} .bk_tt {font-family: monospace} .body-content h2 {border ... m MP, Ardinger HH, et al., editors. GeneReviews? [Internet ]. Seattle (WA): University of Washington, Seattle; ... 1993-2014. GeneReviews ? [Internet ]. Show details Pagon RA, Adam MP, Ardinger HH, et ...

  6. GeneReviews Advanced Search Help - GeneReviews - NCBI Bookshelf [GeneReviews

    Lifescience Database Archive (English)

    Full Text Available p a.figpopup{display:inline !important} .bk_tt {font-family: monospace} .body-content h2 {border ... m MP, Ardinger HH, et al., editors. GeneReviews? [Internet ]. Seattle (WA): University of Washington, Seattle; ... 1993-2014. GeneReviews ? [Internet ]. Show details Pagon RA, Adam MP, Ardinger HH, et ...

  7. Gene-gene and gene-environment interactions: new insights into the prevention, detection and management of coronary artery disease

    OpenAIRE

    Lanktree, Matthew B.; Hegele, Robert A.

    2009-01-01

    Despite the recent success of genome-wide association studies (GWASs) in identifying loci consistently associated with coronary artery disease (CAD), a large proportion of the genetic components of CAD and its metabolic risk factors, including plasma lipids, type 2 diabetes and body mass index, remain unattributed. Gene-gene and gene-environment interactions might produce a meaningful improvement in quantification of the genetic determinants of CAD. Testing for gene-gene and gene-environment ...

  8. Validation of qPCR Methods for the Detection of Mycobacterium in New World Animal Reservoirs.

    Science.gov (United States)

    Housman, Genevieve; Malukiewicz, Joanna; Boere, Vanner; Grativol, Adriana D; Pereira, Luiz Cezar M; Silva, Ita de Oliveira e; Ruiz-Miranda, Carlos R; Truman, Richard; Stone, Anne C

    2015-11-01

    Zoonotic pathogens that cause leprosy (Mycobacterium leprae) and tuberculosis (Mycobacterium tuberculosis complex, MTBC) continue to impact modern human populations. Therefore, methods able to survey mycobacterial infection in potential animal hosts are necessary for proper evaluation of human exposure threats. Here we tested for mycobacterial-specific single- and multi-copy loci using qPCR. In a trial study in which armadillos were artificially infected with M. leprae, these techniques were specific and sensitive to pathogen detection, while more traditional ELISAs were only specific. These assays were then employed in a case study to detect M. leprae as well as MTBC in wild marmosets. All marmosets were negative for M. leprae DNA, but 14 were positive for the mycobacterial rpoB gene assay. Targeted capture and sequencing of rpoB and other MTBC genes validated the presence of mycobacterial DNA in these samples and revealed that qPCR is useful for identifying mycobacterial-infected animal hosts. PMID:26571269

  9. Validation of qPCR Methods for the Detection of Mycobacterium in New World Animal Reservoirs

    Science.gov (United States)

    Housman, Genevieve; Malukiewicz, Joanna; Boere, Vanner; Grativol, Adriana D.; Pereira, Luiz Cezar M.; Silva, Ita de Oliveira e; Ruiz-Miranda, Carlos R.; Truman, Richard; Stone, Anne C.

    2015-01-01

    Zoonotic pathogens that cause leprosy (Mycobacterium leprae) and tuberculosis (Mycobacterium tuberculosis complex, MTBC) continue to impact modern human populations. Therefore, methods able to survey mycobacterial infection in potential animal hosts are necessary for proper evaluation of human exposure threats. Here we tested for mycobacterial-specific single- and multi-copy loci using qPCR. In a trial study in which armadillos were artificially infected with M. leprae, these techniques were specific and sensitive to pathogen detection, while more traditional ELISAs were only specific. These assays were then employed in a case study to detect M. leprae as well as MTBC in wild marmosets. All marmosets were negative for M. leprae DNA, but 14 were positive for the mycobacterial rpoB gene assay. Targeted capture and sequencing of rpoB and other MTBC genes validated the presence of mycobacterial DNA in these samples and revealed that qPCR is useful for identifying mycobacterial-infected animal hosts. PMID:26571269

  10. GeneNet: a database on structure and functional organisation of gene networks

    OpenAIRE

    Ananko, E. A.; Podkolodny, N L; Stepanenko, I L; Ignatieva, E. V.; Podkolodnaya, O. A.; Kolchanov, N.A.

    2002-01-01

    The GeneNet database is designed for accumulation of information on gene networks. Original technology applied in GeneNet enables description of not only a gene network structure and functional relationships between components, but also metabolic and signal transduction pathways. Specialised software, GeneNet Viewer, automatically displays the graphical diagram of gene networks described in the database. Current release 3.0 of GeneNet database contains descriptions of 25 gene networks, 945 pr...

  11. Genes: an Open Access Journal

    Directory of Open Access Journals (Sweden)

    J. Peter W. Young

    2009-11-01

    Full Text Available Genes have been in the scientific vocabulary for a hundred years. The term "gene" was proposed by the Danish plant scientist Wilhelm Johannsen in the first decade of the 20th century. For Johannsen, the gene remained an abstract concept, "free of any hypothesis" [1], but others were already pointing to chromosomes as the likely location of genes. The science of genetics was born at that time, and genes were rapidly connected with mutations, with patterns of inheritance, with development, with quantitative traits, with evolution and with biochemical pathways. All this was achieved without knowledge of the physical nature of genes, but this changed in mid-century with the discoveries of molecular biology. DNA was revealed as the genetic material, and the mechanisms were elucidated by which the information was encoded, and propagated, and linked to the phenotype. However, the concept of a "gene" did not become clearer. Quite the reverse, as the units of mutation, of recombination, of inheritance, of expression, of regulation, etc. did not necessarily coincide. [...

  12. Gene expression profiles in skeletal muscle after gene electrotransfer

    DEFF Research Database (Denmark)

    Hojman, Pernille; Zibert, John R; Gissel, Hanne; Eriksen, Jens; Gehl, Julie

    2007-01-01

    BACKGROUND: Gene transfer by electroporation (DNA electrotransfer) to muscle results in high level long term transgenic expression, showing great promise for treatment of e.g. protein deficiency syndromes. However little is known about the effects of DNA electrotransfer on muscle fibres. We have...... therefore investigated transcriptional changes through gene expression profile analyses, morphological changes by histological analysis, and physiological changes by force generation measurements. DNA electrotransfer was obtained using a combination of a short high voltage pulse (HV, 1000 V/cm, 100 mus......) followed by a long low voltage pulse (LV, 100 V/cm, 400 ms); a pulse combination optimised for efficient and safe gene transfer. Muscles were transfected with green fluorescent protein (GFP) and excised at 4 hours, 48 hours or 3 weeks after treatment. RESULTS: Differentially expressed genes were...

  13. Detail Information [CGED - Cancer Gene Expression Database

    Lifescience Database Archive (English)

    Full Text Available Gene data inhouse ID GS16236 Symbol ECGF1 Gene name endothelial cell growth factor 1 (platelet-d ... Gene Ontology keywords GO 0000002 : mitochondrial genome ... maintenance GO 0001525 : angiogenesis GO 0005161 : ...

  14. BRCA1 and BRCA2 gene testing

    Science.gov (United States)

    The BRCA1 and BRCA2 gene test is a blood test that can tell you if you have a higher ... BRCA1 and BRCA2 are genes that suppress malignant tumors (cancer) in humans. When these genes change (become ...

  15. Gene Therapy for Diseases and Genetic Disorders

    Science.gov (United States)

    ... Mentor Submit Your Press Release Donate Home ASGCT Gene Therapy for Diseases Gene Therapy has made important ... Among the most notable advancements are the following: Gene Therapy for Genetic Disorders Severe Combined Immune Deficiency ( ...

  16. [Globin gene induction therapy for ?-thalassemia].

    Science.gov (United States)

    Lu, Zhou-Ming

    2014-02-01

    Globin gene induction therapy is a new treatment under study for ?-thalassemia. This review summarizes the research progress on the mechanisms of globin gene induction therapy for ?-thalassemia and current ?-globin gene induction medicines. PMID:24598686

  17. Compositional gradients in Gramineae genes

    DEFF Research Database (Denmark)

    Wong, Gane Ka-Shu; Wang, Jun; Tao, Lin; Tan, Jun; Zhang, JianGuo; Passey, Douglas A; Yu, Jun

    2002-01-01

    In this study, we describe a property of Gramineae genes, and perhaps all monocot genes, that is not observed in eudicot genes. Along the direction of transcription, beginning at the junction of the 5'-UTR and the coding region, there are gradients in GC content, codon usage, and amino-acid usage. The magnitudes of these gradients are large enough to hinder the annotation of the rice genome and to confound the detection of protein homologies across the monocot-eudicot divide. Udgivelsesdato: 200...

  18. Are TMEM genes potential candidate genes for panic disorder?

    DEFF Research Database (Denmark)

    Gregersen, Noomi O; Buttenschøn, Henriette Nørmølle; Hedemand, Anne; Dahl, Hans Atli; Kristensen, Ann S; Clementsen, Birita; Woldbye, David P D; Koefoed, Pernille; Erhardt, Angelika; Kruse, Torben A; Wang, August Gabriel; Børglum, Anders D; Mors, Ole

    2014-01-01

    We analysed single nucleotide polymorphisms in two transmembrane genes (TMEM98 and TMEM132E) in panic disorder (PD) patients and control individuals from the Faroe Islands, Denmark and Germany. The genes encode single-pass membrane proteins and are located within chromosome 17q11.2-q12, a previously reported candidate region for PD. Three single nucleotide polymorphisms (rs887231, rs887230 and rs4795942) located upstream and within TMEM132E showed a nominal significant association with PD primar...

  19. Gene-Environment Dependence Creates Spurious Gene-Environment Interaction

    OpenAIRE

    Dudbridge, Frank; Fletcher, Olivia

    2014-01-01

    Gene-environment interactions have the potential to shed light on biological processes leading to disease and to improve the accuracy of epidemiological risk models. However, relatively few such interactions have yet been confirmed. In part this is because genetic markers such as tag SNPs are usually studied, rather than the causal variants themselves. Previous work has shown that this leads to substantial loss of power and increased sample size when gene and environment are independent. Howe...

  20. Gene Duplication and Ectopic Gene Conversion in Drosophila

    OpenAIRE

    Tim Connallon; J. Roman Arguello

    2011-01-01

    The evolutionary impact of gene duplication events has been a theme of Drosophila genetics dating back to the Morgan School. While considerable attention has been placed on the genetic novelties that duplicates are capable of introducing, and the role that positive selection plays in their early stages of duplicate evolution, much less attention has been given to the potential consequences of ectopic (non-allelic) gene conversion on these evolutionary processes. In this paper we consider the ...