Phenotypic and genotypic analysis of anti-tuberculosis drug resistance in Mycobacterium tuberculosis isolates in Myanmar.

Science.gov (United States)

Aung, Wah Wah; Ei, Phyu Win; Nyunt, Wint Wint; Swe, Thyn Lei; Lwin, Thandar; Htwe, Mi Mi; Kim, Kyung Jun; Lee, Jong Seok; Kim, Chang Ki; Cho, Sang Nae; Song, Sun Dae; Chang, Chulhun L

2015-09-01

Tuberculosis (TB) is one of the most serious health problems in Myanmar. Because TB drug resistance is associated with genetic mutation(s) relevant to responses to each drug, genotypic methods for detecting these mutations have been proposed to overcome the limitations of classic phenotypic drug susceptibility testing (DST). We explored the current estimates of drug-resistant TB and evaluated the usefulness of genotypic DST in Myanmar. We determined the drug susceptibility of Mycobacterium tuberculosis isolated from sputum smear-positive patients with newly diagnosed pulmonary TB at two main TB centers in Myanmar during 2013 by using conventional phenotypic DST and the GenoType MTBDRplus assay (Hain Lifescience, Germany). Discrepant results were confirmed by sequencing the genes relevant to each type of resistance (rpoB for rifampicin; katG and inhA for isoniazid). Of 191 isolates, phenotypic DST showed that 27.7% (n=53) were resistant to at least one first-line drug and 20.9% (n=40) were resistant to two or more, including 18.3% (n=35) multidrug-resistant TB (MDR-TB) strains. Monoresistant strains accounted for 6.8% (n=13) of the samples. Genotypic assay of 189 isolates showed 17.5% (n=33) MDR-TB and 5.3% (n=10) isoniazid-monoresistant strains. Genotypic susceptibility results were 99.5% (n=188) concordant and agreed almost perfectly with phenotypic DST (kappa=0.99; 95% confidence interval 0.96-1.01). The results highlight the burden of TB drug resistance and prove the usefulness of the genotypic DST in Myanmar.

Tuberculosis associated factors caused by Mycobacterium tuberculosis of the RDRio genotype

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Eloise Brasil Moraes

Full Text Available BACKGROUND Tuberculosis (TB continues to be a disease that affects many countries around the world, including Brazil. Recently, a subtype of Latin American-Mediterranean family strain was identified and characterised by RDRio. The strain has been associated with different characteristics of the disease. OBJECTIVES In the present study we investigated the association of epidemiological, clinical, radiological and bacteriological variables with pulmonary tuberculosis caused by RDRioMycobacterium tuberculosis strain in large regions of São Paulo. METHODS We conducted a cross-sectional study in 530 patients with pulmonary tuberculosis, diagnosed using sputum culture, from two regions of the São Paulo state in Brazil. The samples were brought to São Paulo reference laboratories for epidemiological, clinical, radiological and bacteriological analyses, and the data were obtained from a TB notification system. RDRio genotyping and Spoligotyping of the samples were performed. For the analysis of the categorical variables we used the chi-square test or the Fisher’s exact test, and for the continuous variables, the Mann-Whitney test. In addition, a logistic regression was used for multivariate analysis. Differences with p < 0.05 were considered significant. FINDINGS The RDRio deletion was identified in 152 (28.7% samples. In the univariate analysis, both the age groups above 25 years and alcohol consumption were associated with the RDRio deletion. The multivariate analysis confirmed the association of the RDRio deletion with the age groups: 25-35 years old [OR: 2.28 (1.02-5.07; p = 0.04] and 36-60 years old (OR: 2.36 (1.11-5.05; p = 0.03], and also with alcohol consumption [OR: 1.63 (1.05-2.54; p = 0,03]. MAIN CONCLUSIONS In this study, we identified new factors associated with the M. tuberculosis of the RDRio deletion strains infection.

A Mycobacterium tuberculosis cluster demonstrating the use of genotyping in urban tuberculosis control

NARCIS (Netherlands)

G. de Vries (Gerard); R.M. van Hest (Reinier); C.C.A. Burdo (Conny); D. van Soolingen (Dick); J.H. Richardus (Jan Hendrik)

2009-01-01

textabstractBackground: DNA fingerprinting of Mycobacterium tuberculosis isolates offers better opportunities to study links between tuberculosis (TB) cases and can highlight relevant issues in urban TB control in low-endemic countries. Methods: A medium-sized molecular cluster of TB cases with

Genotypes of Mycobacterium tuberculosis in patients at risk of drug resistance in Bolivia.

Science.gov (United States)

Monteserin, Johana; Camacho, Mirtha; Barrera, Lucía; Palomino, Juan Carlos; Ritacco, Viviana; Martin, Anandi

2013-07-01

Bolivia ranks among the 10 Latin American countries with the highest rates of tuberculosis (TB) and multidrug resistant (MDR) TB. In view of this, and of the lacking information on the population structure of Mycobacterium tuberculosis in the country, we explored genotype associations with drug resistance and clustering by analyzing isolates collected in 2010 from 100 consecutive TB patients at risk of drug resistance in seven of the nine departments in which Bolivia is divided. Fourteen isolates were MDR, 29 had other drug resistance profiles, and 57 were pansusceptible. Spoligotype family distribution was: Haarlem 39.4%, LAM 26.3%, T 22.2%, S 2.0%, X 1.0%, orphan 9.1%, with very low intra-family diversity and absence of Beijing genotypes. We found 66 different MIRU-VNTR patterns; the most frequent corresponded to Multiple Locus Variable Analysis (MLVA) MtbC15 patterns 860, 372 and 873. Twelve clusters, each with identical MIRU-VNTR and spoligotypes, gathered 35 patients. We found no association of genotype with drug resistant or MDR-TB. Clustering associated with SIT 50 and the H3 subfamily to which it belongs (pBolivia. However, results should be taken cautiously because the sample is small and includes a particular subset of M. tuberculosis population. Copyright © 2013 Elsevier B.V. All rights reserved.

First insight into the genotypic diversity of clinical Mycobacterium tuberculosis isolates from Gansu Province, China.

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Jie Liu

Full Text Available BACKGROUND: Investigations of Mycobacterium tuberculosis genetic diversity in China have indicated a significant regional distribution. The aim of this study was to characterize the genotypes of clinical M. tuberculosis isolates obtained from Gansu, which has a special geographic location in China. METHODOLOGY/PRINCIPAL FINDINGS: A total of 467 clinical M. tuberculosis strains isolated in Gansu Province were genotyped by 15-locus mycobacterial interspersed repetitive units-variable number tandem repeats (MIRU-VNTR and spoligotyping. The results showed that 445 isolates belonged to six known spoligotype lineages, whereas 22 isolates were unknown. The Beijing genotype was the most prevalent (87.58%, n = 409, while the shared type 1 was the dominant genotype (80.94%, n = 378. The second most common lineage was the T lineage, with 25 isolates (5.35%, followed by the H lineage with 5 isolates (1.07%, the MANU family (0.64%, 3 isolates, the U family (0.43%, 2 isolates and the CAS lineage with 1 isolate (0.21%. By using the VNTR15China method, we observed 15 groups and 228 genotypes among the 467 isolates. We found no association between the five larger groups (including the Beijing genotype and sex, age, or treatment status, and there was no noticeable difference in the group analysis in different areas. In the present study, seven of the 15 MIRU-VNTR loci were highly or moderately discriminative according to their Hunter-Gaston discriminatory index. CONCLUSIONS/SIGNIFICANCE: The Beijing genotype is the predominant genotype in Gansu province. We confirm that VNTR15China is suitable for typing Beijing strains in China and that it has a better discriminatory power than spoligotyping. Therefore, the use of both methods is the most suitable for genotyping analysis of M. tuberculosis.

Automated High-Throughput Genotyping for Study of Global Epidemiology of Mycobacterium tuberculosis Based on Mycobacterial Interspersed Repetitive Units

Science.gov (United States)

Supply, Philip; Lesjean, Sarah; Savine, Evgueni; Kremer, Kristin; van Soolingen, Dick; Locht, Camille

2001-01-01

Large-scale genotyping of Mycobacterium tuberculosis is especially challenging, as the current typing methods are labor-intensive and the results are difficult to compare among laboratories. Here, automated typing based on variable-number tandem repeats (VNTRs) of genetic elements named mycobacterial interspersed repetitive units (MIRUs) in 12 mammalian minisatellite-like loci of M. tuberculosis is presented. This system combines analysis of multiplex PCRs on a fluorescence-based DNA analyzer with computerized automation of the genotyping. Analysis of a blinded reference set of 90 strains from 38 countries (K. Kremer et al., J. Clin. Microbiol. 37:2607–2618, 1999) demonstrated that it is 100% reproducible, sensitive, and specific for M. tuberculosis complex isolates, a performance that has not been achieved by any other typing method tested in the same conditions. MIRU-VNTRs can be used for analysis of the global genetic diversity of M. tuberculosis complex strains at different levels of evolutionary divergence. To fully exploit the portability of this typing system, a website was set up for the analysis of M. tuberculosis MIRU-VNTR genotypes via the Internet. This opens the way for global epidemiological surveillance of tuberculosis and should lead to novel insights into the evolutionary and population genetics of this major pathogen. PMID:11574573

Occupational Tuberculosis in Denmark through 21 Years Analysed by Nationwide Genotyping

DEFF Research Database (Denmark)

Pedersen, Mathias Klok; Andersen, Aase Bengaard; Andersen, Peter Henrik

2016-01-01

Tuberculosis (TB) is a well-known occupational hazard. Based on more than two decades (1992-2012) of centralized nationwide genotyping of all Mycobacterium tuberculosis culture-positive TB patients in Denmark, we compared M. tuberculosis genotypes from all cases notified as presumed occupational (N...... = 130) with M. tuberculosis genotypes from all TB cases present in the country (N = 7,127). From 1992 through 2006, the IS6110 Restriction Fragment Length Polymorphism (RFLP) method was used for genotyping, whereas from 2005 to present, the 24-locus-based Mycobacterial Interspersed Repetitive Unit...

Tuberculosis genotyping information management system: enhancing tuberculosis surveillance in the United States.

Science.gov (United States)

Ghosh, Smita; Moonan, Patrick K; Cowan, Lauren; Grant, Juliana; Kammerer, Steve; Navin, Thomas R

2012-06-01

Molecular characterization of Mycobacterium tuberculosis complex isolates (genotyping) can be used by public health programs to more readily identify tuberculosis (TB) transmission. The Centers for Disease Control and Prevention's National Tuberculosis Genotyping Service has offered M. tuberculosis genotyping for every culture-confirmed case in the United States since 2004. The TB Genotyping Information Management System (TB GIMS), launched in March 2010, is a secure online database containing genotype results linked with case characteristics from the national TB registry for state and local TB programs to access, manage and analyze these data. As of September 2011, TB GIMS contains genotype results for 89% of all culture-positive TB cases for 2010. Over 400 users can generate local and national reports and maps using TB GIMS. Automated alerts on geospatially concentrated cases with matching genotypes that may represent outbreaks are also generated by TB GIMS. TB genotyping results are available to enhance national TB surveillance and apply genotyping results to conduct TB control activities in the United States. Published by Elsevier B.V.

Association between genotype and drug resistance profiles of Mycobacterium tuberculosis strains circulating in China in a national drug resistance survey

NARCIS (Netherlands)

Zhou, Yang; van den Hof, Susan; Wang, Shengfen; Pang, Yu; Zhao, Bing; Xia, Hui; Anthony, Richard; Ou, Xichao; Li, Qiang; Zheng, Yang; Song, Yuanyuan; Zhao, Yanlin; van Soolingen, Dick

2017-01-01

We describe the population structure of a representative collection of 3,133 Mycobacterium tuberculosis isolates, collected within the framework of a national resistance survey from 2007 in China. Genotyping data indicate that the epidemic strains in China can be divided into seven major complexes,

The Mycobacterium tuberculosis homologue of the Mycobacterium ...

African Journals Online (AJOL)

With the completion of genome sequencing of Mycobacterium tuberculosis and upsurge in the incidence of M. tuberculosis infection worldwide partly as a result of HIV pandemic, there is need for rationale approach to vaccine and chemotherapy discoveries for M. tuberculosis. The homologue of mig gene of. Mycobacterium ...

High genetic diversity among Mycobacterium tuberculosis strains in Tehran, Iran

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Taher Azimi

2018-05-01

Full Text Available Introduction: Tuberculosis (TB still remains an important public health problem in Iran. The genotyping of Mycobacterium tuberculosis isolates is expected to lead to a better understanding of M. tuberculosis transmission in Tehran, the most populated city of Iran. Materials and Methods: A total of 2300 clinical specimens were obtained from TB suspected patients who were referred to a TB center in Tehran from Jan 2014 to Dec 2016. Identification was performed using both conventional and molecular methods. The presence of resistance to rifampicin was examined by the GeneXpert MTB/RIF. The standard 15-locus mycobacterial interspersed repetitive units-variable number of tandem repeats (MIRU-VNTR typing method was applied to genotype of clinical isolates. Results: Of 2300 specimens, 80 isolates were identified as M. tuberculosis by using biochemical and molecular tests. Of 80 M. tuberculosis isolates, 76 (95% had unique genotypic profiles and 4 (5% shared a profile with one or more other strains. Based on single loci variation (SLV 4 clonal complexes were observed. NEW-1 was found to be the most predominant lineage (22.5% followed by West African (1.25%, Central Asian (CAS/Delhi (1.25%, Bovis (1.25%, H37Rv (1.25% and multiple matches (1.25%. Loci MIRU10, MIRU26, MTUB21 and QUB26 were found as highly discriminative. No mutation was detected in the hotspot region of rifampicin by using GeneXpert MTB/RIF. Conclusions: Our study findings show that there was considerable genotypic diversity among M. tuberculosis isolates in Tehran. The 15-locus MIRU-VNTR showed high HGDI and could be used as a first-line genotyping method for epidemiological studies. Keywords: Mycobacterium tuberculosis, Genotyping, MIRU-VNTR, Tehran, Iran

Modern lineages of Mycobacterium tuberculosis in Addis Ababa ...

African Journals Online (AJOL)

Background: The genotyping of Mycobacterium tuberculosis strains is important to have unique insights into the dissemination dynamics and evolutionary genetics of this pathogen and for TB control as it allows the detection of suspected outbreaks and the tracing of transmission chains. Objective: To characterize M.

Polymorphisms of twenty regulatory proteins between Mycobacterium tuberculosis and Mycobacterium bovis

Science.gov (United States)

Mycobacterium tuberculosis and Mycobacterium bovis are responsible for tuberculosis in humans or animals, respectively. Both species are closely related and belong to the Mycobacterium tuberculosis complex (MTC). M. tuberculosis is the most ancient species from which M. bovis and the other members o...

Tracing Mycobacterium tuberculosis transmission by whole genome sequencing in a high incidence setting

DEFF Research Database (Denmark)

Bjorn-Mortensen, K; Soborg, B; Koch, A

2016-01-01

In East Greenland, a dramatic increase of tuberculosis (TB) incidence has been observed in recent years. Classical genotyping suggests a genetically similar Mycobacterium tuberculosis (Mtb) strain population as cause, however, precise transmission patterns are unclear. We performed whole genome...

Genotyping and drug resistance patterns of Mycobacterium tuberculosis strains observed in a tuberculosis high-burden municipality in Northeast, Brazil

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Roberta dos Santos Silva Luiz

2013-06-01

Full Text Available OBJECTIVES: This study has used a combination of clinical information, spoligotyping, and georeferencing system to elucidate the genetic diversity of the Mycobacterium tuberculosis isolates circulating in a TB-prevalent municipality of Northeast Brazil. METHODS: A total of 115 M. tuberculosis strains were isolated from pulmonary tuberculosis patients from January 2007 to March 2008 in Fortaleza. Drug susceptibility and spoligotyping assays were performed and place of residence of the patients were georeferenced. RESULTS: Of the M. tuberculosis strains studied, 51 (44.3% isolates were resistant to at least one drug (R-TB and 64 (55.7% were sensitive to all the drugs tested (S-TB. A high frequency of resistance was found in previously treated cases (84% and among new cases (16%; p < 0.001. a total of 74 (64% isolates were grouped into 22 spoligotyped lineages, while 41 (36% isolates were identified as new. among the predominant genotypes, 33% were latim american mediterranean (lam, 12% haarlem (h, and 5% u. there was no association of geographic distribution of rt-tb patients as compared to the controls and also the geographic location to the spoligotype patterns. the geospatial analysis revealed that 24 (23% patients (hot spot zones either shared the same residence or lived in a close neighborhood of a case. among these concentration zones, the patients lived in the same residence and shared a common genotype pattern and resistance pattern. DISCUSSION: it was observed that the spoligopatterns family distribution was similar to that reported for south america, prevailing the lam and h lineages. a high rate-case among the resistant TB group occurs as a result of transmitted and acquired resistance. A more effective surveillance program is needed in order to succeed in reducing tuberculosis in Northeast Brazil.

Genotyping and drug resistance patterns of Mycobacterium tuberculosis strains observed in a tuberculosis high-burden municipality in Northeast, Brazil

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Roberta dos Santos Silva Luiz

Full Text Available OBJECTIVES: This study has used a combination of clinical information, spoligotyping, and georeferencing system to elucidate the genetic diversity of the Mycobacterium tuberculosis isolates circulating in a TB-prevalent municipality of Northeast Brazil. METHODS: A total of 115 M. tuberculosis strains were isolated from pulmonary tuberculosis patients from January 2007 to March 2008 in Fortaleza. Drug susceptibility and spoligotyping assays were performed and place of residence of the patients were georeferenced. RESULTS: Of the M. tuberculosis strains studied, 51 (44.3% isolates were resistant to at least one drug (R-TB and 64 (55.7% were sensitive to all the drugs tested (S-TB. A high frequency of resistance was found in previously treated cases (84% and among new cases (16%; p < 0.001. a total of 74 (64% isolates were grouped into 22 spoligotyped lineages, while 41 (36% isolates were identified as new. among the predominant genotypes, 33% were latim american mediterranean (lam, 12% haarlem (h, and 5% u. there was no association of geographic distribution of rt-tb patients as compared to the controls and also the geographic location to the spoligotype patterns. the geospatial analysis revealed that 24 (23% patients (hot spot zones either shared the same residence or lived in a close neighborhood of a case. among these concentration zones, the patients lived in the same residence and shared a common genotype pattern and resistance pattern. DISCUSSION: it was observed that the spoligopatterns family distribution was similar to that reported for south america, prevailing the lam and h lineages. a high rate-case among the resistant TB group occurs as a result of transmitted and acquired resistance. A more effective surveillance program is needed in order to succeed in reducing tuberculosis in Northeast Brazil.

Genotyping did not evidence any contribution of Mycobacterium bovis to human tuberculosis in Brazil.

Science.gov (United States)

Rocha, Adalgiza; Elias, Atina R; Sobral, Luciana F; Soares, Diego F; Santos, Alexandre C; Marsico, Ana-Grazia; Hacker, Mariana A; Caldas, Paulo C; Parente, Luiz C; Silva, Marcio R; Fonseca, Leila; Suffys, Philip; Boéchat, Neio

2011-01-01

The contribution of Mycobacterium bovis to the global burden of tuberculosis (TB) in man is likely to be underestimated due to its dysgonic growth characteristics and because of the absence of pyruvate in most used media is disadvantageous for its primary isolation. In Brazil Mycobacterium culture, identification and susceptibility tests are performed only in TB reference centers, usually for selected cases. Moreover, solid, egg-based, glycerol-containing (without pyruvate supplementation) Löwenstein-Jensen (L-J) or Ogawa media are routinely used, unfavouring M. bovis isolation. To determine the importance of M. bovis as a public health threat in Brazil we investigated 3046 suspected TB patients inoculating their clinical samples onto routine L-J and L-J pyruvate enriched media. A total of 1796 specimens were culture positive for Mycobacterium spp. and 702 TB cases were confirmed. Surprisingly we did not detect one single case of M. bovis in the resulting collection of 1674 isolates recovered from M. bovis favourable medium analyzed by conventional and molecular speciation methods. Also, bacillary DNA present on 454 sputum smears from 223 TB patients were OxyR genotyped and none was recognized as M. bovis. Our data indicate that M. bovis importance on the burden of human TB in Brazil is marginal. Copyright © 2010 Elsevier Ltd. All rights reserved.

Mycobacterium tuberculosis infection of domesticated Asian elephants, Thailand.

OpenAIRE

2011-01-01

Four Asian elephants were confirmed to be infected with Mycobacterium tuberculosis by bacterial culture, other diagnostic procedures, and sequencing of 16S–23S rDNA internal transcribed spacer region, 16S rRNA, and gyrase B gene sequences. Genotyping showed that the infectious agents originated from 4 sources in Thailand. To identify infections, a combination of diagnostic assays is essential.

First description of Mycobacterium tuberculosis Beijing genotype in Argentina Primera descripción del genotipo Beijing de Mycobacterium tuberculosis en la Argentina

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N. Morcillo

2005-06-01

Full Text Available During a population-based study to genotype isolates of Mycobacterium tuberculosis from Buenos Aires Northern suburbs, we found isolates with molecular patterns related to those of the Beijing genotype. Five out of 590 (0.85% patients had isolates with spoligopattern identical to that of the Beijing family. Since two of these isolates showed identical IS6110RFLP pattern, we found only four different patterns containing 11 to 19 bands. The isolates were obtained from young people (including a 7 years-old child who were born in Argentina, and were living in a small area of our region. However, conventional contact tracing did not prove epidemiological linkage among them. These isolates were fully drug-susceptible to the first-line drugs. The comparison of the IS6110RFLP patterns from our isolates against a set of 19 reference Beijing patterns from the RIVM (The Netherlands confirmed that the strains belonged to the Beijing lineage. These findings might be partially explained by the important migration phenomena occurred during the last decade. Further surveillance studies would help in the following of Beijing family strain dissemination in our community.Durante un estudio poblacional para genotipificar aislamientos de Mycobacterium tuberculosis de Gran Buenos Aires Norte, se encontraron aislamientos con patrones moleculares relacionados con el genotipo Beijing. Cinco de 590 (0,85% pacientes tuvieron aislamientos con un patrón de spoligotyping idéntico al de la familia Beijing. Dos de esos aislamientos mostraron un patrón de IS6110 RFLP idéntico, por lo que finalmente fueron encontrados cuatro patrones diferentes conteniendo entre 11 y 19 bandas. Las cepas fueron aisladas de personas jóvenes incluyendo un niño de siete años de edad, todos ellos nacidos en Argentina y residentes en un pequeño distrito de nuestra región. Sin embargo, no pudo hallarse ninguna conexión epidemiológica entre ellos. La comparación de los patrones de IS6110

Mycobacterium bovis and Other Uncommon Members of the Mycobacterium tuberculosis Complex.

Science.gov (United States)

Esteban, Jaime; Muñoz-Egea, Maria-Carmen

2016-12-01

Since its discovery by Theobald Smith, Mycobacterium bovis has been a human pathogen closely related to animal disease. At present, M. bovis tuberculosis is still a problem of importance in many countries and is considered the main cause of zoonotic tuberculosis throughout the world. Recent development of molecular epidemiological tools has helped us to improve our knowledge about transmission patterns of this organism, which causes a disease indistinguishable from that caused by Mycobacterium tuberculosis. Diagnosis and treatment of this mycobacterium are similar to those for conventional tuberculosis, with the important exceptions of constitutive resistance to pyrazinamide and the fact that multidrug-resistant and extremely drug-resistant M. bovis strains have been described. Among other members of this complex, Mycobacterium africanum is the cause of many cases of tuberculosis in West Africa and can be found in other areas mainly in association with immigration. M. bovis BCG is the currently available vaccine for tuberculosis, but it can cause disease in some patients. Other members of the M. tuberculosis complex are mainly animal pathogens with only exceptional cases of human disease, and there are even some strains, like "Mycobacterium canettii," which is a rare human pathogen that could have an important role in the knowledge of the evolution of tuberculosis in the history.

In vitro susceptibility of Mycobacterium tuberculosis, Mycobacterium africanum, Mycobacterium bovis, Mycobacterium avium, Mycobacterium fortuitum, and Mycobacterium chelonae to ticarcillin in combination with clavulanic acid.

OpenAIRE

Casal, M J; Rodriguez, F C; Luna, M D; Benavente, M C

1987-01-01

The in vitro susceptibility of Mycobacterium tuberculosis, Mycobacterium bovis, Mycobacterium africanum, Mycobacterium avium, Mycobacterium fortuitum, and Mycobacterium chelonae (M. chelonei) to ticarcillin in combination with calvulanic acid (CA) was studied by the agar dilution method. All the M. tuberculosis, M. bovis, and M. africanum strains were inhibited at a ticarcillin concentration of 32 micrograms/ml or lower in combination with 5 micrograms of CA. M. chelonae and M. avium strains ...

Line probe assay for differentiation within Mycobacterium tuberculosis complex. Evaluation on clinical specimens and isolates including Mycobacterium pinnipedii

DEFF Research Database (Denmark)

Kjeldsen, Marianne Kirstine; Bek, Dorte; Rasmussen, Erik Michael

2009-01-01

A line probe assay (GenoType MTBC) was evaluated for species differentiation within the Mycobacterium tuberculosis complex (MTBC). We included 387 MTBC isolates, 43 IS6110 low-copy MTBC isolates, 28 clinical specimens with varying microscopy grade, and 30 isolates of non-tuberculous mycobacteria...

Multidrug-Resistant Mycobacterium tuberculosis of the Latin American Mediterranean Lineage, Wrongly Identified as Mycobacterium pinnipedii (Spoligotype International Type 863 [SIT863]), Causing Active Tuberculosis in South Brazil

KAUST Repository

Dalla Costa, Elis R.; Vasconcelos, Sidra E. G.; Esteves, Leonardo S.; Gomes, Harrison M.; Gomes, Lia L.; Almeida da Silva, Pedro; Perdigã o, Joã o; Portugal, Isabel; Viveiros, Miguel; McNerney, Ruth; Pain, Arnab; Clark, Taane G.; Rastogi, Nalin; Unis, Gisela; Rossetti, Maria Lucia R.; Suffys, Philip Noel

2015-01-01

We recently detected the spoligotype patterns of strains of Mycobacterium pinnipedii, a species of the Mycobacterium tuberculosis complex, in sputum samples from nine cases with pulmonary tuberculosis residing in Porto Alegre, South Brazil. Because this species is rarely encountered in humans, we further characterized these nine isolates by additional genotyping techniques, including 24-locus mycobacterial interspersed repetitive-unit–variable-number tandem-repeat (MIRU-VNTR) typing, verification of the loci TbD1, RD9, pks15/1, RDRio, and fbpC, the insertion of IS6110 at a site specific to the M. tuberculosis Latin American Mediterranean (LAM) lineage, and whole-genome sequencing. The combined analysis of these markers revealed that the isolates are in fact M. tuberculosis and more specifically belong to the LAM genotype. Most of these isolates (n = 8) were shown to be multidrug resistant (MDR), which prompted us to perform partial sequencing of the rpoA, rpoB, rpoC, katG, and inhA genes. Seven isolates (77.8%) carried the S315T mutation in katG, and one of these (11%) also presented the C(−17)T single-nucleotide polymorphism (SNP) in inhA. Interestingly, six of the MDR isolates also presented an undescribed insertion of 12 nucleotides (CCA GAA CAA CCC) in codon 516 of rpoB. No putative compensatory mutation was found in either rpoA or rpoC. This is the first report of an M. tuberculosis LAM family strain with a convergent M. pinnipedii spoligotype. These spoligotypes are observed in genotype databases at a modest frequency, highlighting that care must be taken when identifying isolates in the M. tuberculosis complex on the basis of single genetic markers.

Multidrug-Resistant Mycobacterium tuberculosis of the Latin American Mediterranean Lineage, Wrongly Identified as Mycobacterium pinnipedii (Spoligotype International Type 863 [SIT863]), Causing Active Tuberculosis in South Brazil

KAUST Repository

Dalla Costa, Elis R.

2015-09-23

We recently detected the spoligotype patterns of strains of Mycobacterium pinnipedii, a species of the Mycobacterium tuberculosis complex, in sputum samples from nine cases with pulmonary tuberculosis residing in Porto Alegre, South Brazil. Because this species is rarely encountered in humans, we further characterized these nine isolates by additional genotyping techniques, including 24-locus mycobacterial interspersed repetitive-unit–variable-number tandem-repeat (MIRU-VNTR) typing, verification of the loci TbD1, RD9, pks15/1, RDRio, and fbpC, the insertion of IS6110 at a site specific to the M. tuberculosis Latin American Mediterranean (LAM) lineage, and whole-genome sequencing. The combined analysis of these markers revealed that the isolates are in fact M. tuberculosis and more specifically belong to the LAM genotype. Most of these isolates (n = 8) were shown to be multidrug resistant (MDR), which prompted us to perform partial sequencing of the rpoA, rpoB, rpoC, katG, and inhA genes. Seven isolates (77.8%) carried the S315T mutation in katG, and one of these (11%) also presented the C(−17)T single-nucleotide polymorphism (SNP) in inhA. Interestingly, six of the MDR isolates also presented an undescribed insertion of 12 nucleotides (CCA GAA CAA CCC) in codon 516 of rpoB. No putative compensatory mutation was found in either rpoA or rpoC. This is the first report of an M. tuberculosis LAM family strain with a convergent M. pinnipedii spoligotype. These spoligotypes are observed in genotype databases at a modest frequency, highlighting that care must be taken when identifying isolates in the M. tuberculosis complex on the basis of single genetic markers.

[Study on VNTR diversity of clinical Mycobacterium tuberculosis isolates from Qinghai].

Science.gov (United States)

Li, Bin; Liu, Haican; Wang, Zhaofen; Ma, Yongcheng; Su, Xiaodong; Jiang, Mingxia; Wan, Kanglin; Liu, Shou; Zhao, Xiuqin; Qu, Shugen

2015-10-01

To investigate the variable number tandem repeats (VNTR) genetic polymorphisms, genotyping and distribution pattern of clinical Mycobacterium (M.) tuberculosis isolates from Qinghai province. The clinical M. tuberculosis strains isolated from the patients with tuberculosis and related background data were collected from Qinghai Provincial Center for Disease Control and Prevention from 2009 to 2012. Genotyping was conducted by using multiple locus VNTR analysis (MLVA). Genomic DNA was extracted and 15 VNTR loci were amplified with PCR and the PCR products were detected with gel electrophoresis. The VNTR diversity and clusters of genotyping were analyzed with BioNumerics (Version 5.0). A total of 251 clinical M. tuberculosis isolates were analyzed with 15 VNTR loci showing that there were great genetic diversity in these isolates. Six of the 15 VNTR loci, showed that the Hunter-Gaston index (HGI) were higher than 0.6, in which the highest resolution was MIRU26. The clusters of genotyping showed that these isolates could be categorized into four gene clusters and 238 genotypes. The four gene clusters accounted for 4.9%, 91.9%, 1.6% and 1.6% of the clinical isolates, respectively. The results showed that there is great variety of VNTR genetic polymorphisms in clinical M. tuberculosis isolates in Qinghai province.

Genotyping of rifampin-resistant Mycobacterium tuberculosis isolates from Western Turkey

International Nuclear Information System (INIS)

Cavasoglu, Cengiz; Bilgic, Altinay; Durmaz, Riza; Gunal, Selami

2004-01-01

Although the rate of multiple drug resistance is high there is no published data on the transmission rate of drug-resistant strains of Mycobacterium tuberculosis in the Aegean region of Western Turkey that are based on molecular methods. IS6110 and pTBN12 restriction fragment lengthpolymorphism (RFLP) methods were used for typing Mycobacterium tuberculosis isolated from 26 sputum samples from 26 patients. 19 of rifampin-resistant isolates (73.1%) contained 6 to 11 copies of 156110. Eighteen different IS6110 DNA fingerprint patterns were observed in the 26 rifampin resistant isolates. 23 of the 26 rifampin-resistant isolates were also resistant to isoniazid. When evaluated together, both methods yielded 21 (80.9%) different banding patterns and the level of clustering was 34.6%. The average number per pattern was 1.23 (26/21). IS6110 fingerprinting suggests that the rifampin-resistant isolates obtained from the Aegean region had a relatively high clustering rate and were clonally related. These findings showed that the rifampin-resistant isolates are actively transmitted between patients. Urgent measures should be taken to prevent the spread of these resistant strains. (author)

Mycobacterium tuberculosis Isolates from Single Outpatient Clinic in Panama City Exhibit Wide Genetic Diversity

Science.gov (United States)

Sambrano, Dilcia; Correa, Ricardo; Almengor, Pedro; Domínguez, Amada; Vega, Silvio; Goodridge, Amador

2014-01-01

Understanding Mycobacterium tuberculosis biodiversity and transmission is significant for tuberculosis control. This short report aimed to determine the genetic diversity of M. tuberculosis isolates from an outpatient clinic in Panama City. A total of 62 M. tuberculosis isolates were genotyped by 12 loci mycobacterial interspersed repetitive units-variable number of tandem repeats (MIRU-VNTR) and Spoligotyping. Forty-five (72.6%) of the isolates showed unique MIRU-VNTR genotypes, and 13 (21%) of the isolates were grouped into four clusters. Four isolates showed polyclonal MIRU-VNTR genotypes. The MIRU-VNTR Hunter-Gaston discriminatory index reached 0.988. The Spoligotyping analysis revealed 16 M. tuberculosis families, including Latin American-Mediterranean, Harlem, and Beijing. These findings suggest a wide genetic diversity of M. tuberculosis isolates at one outpatient clinic. A detailed molecular epidemiology survey is now warranted, especially following second massive immigration for local Panama Canal expansion activities. PMID:24865686

Radiographic differentiation of atypical tuberculosis from mycobacterium tuberculosis

International Nuclear Information System (INIS)

Tarver, R.D.; Pearcy, E.A.; Conces, D.J. Jr.; Mathur, P.N.

1987-01-01

The chest radiographs of 95 patients with the new diagnosis of atypical turberculosis were reviewed to determine if any significant differences between atypical tuberculosis and that caused by Mycobacterium tuberculosis could be discerned. Findings included upper lobe involvement in B4 of the 95 patients and cavities in 76, with nearly equal groups having no, moderate, or extensive surrounding alveolar disease. Nodules were common; in six patients a nodule was the sole manifestation of disease. Adenopathy was seen in 12 of the 95 patients, atlectasis in 45, pleural thickening in 90, and effusions in three. These radiographic findings did not allow the radiographic differentiation of atypical tuberculosis from Mycobacterium tuberculosis infection

Drug Resistance of Mycobacterium tuberculosis Complex among ...

African Journals Online (AJOL)

BACKGROUND: In Burkina Faso, there is no recent data about the level of drug resistance in Mycobacterium tuberculosis strains among newly diagnosed tuberculosis cases. OBJECTIVE: To provide an update of the primary drug resistance of mycobacterium tuberculosis among patients in Burkina faso. METHODS: ...

First baseline of circulating genotypic lineages of Mycobacterium tuberculosis in patients from the brazilian borders with Argentina and Paraguay.

Science.gov (United States)

Machado, Luzia Neri C; Marcondes, Nadir R; Leite, Clarice Q Fijimura; Santos, Adolfo C Barreto; Pavan, Fernando Rogério; Baldin, Vanessa Pietrowski; Castilho, Aline Lemes; Siqueira, Vera Lúcia D; Baeza, Lilian Cristiane; Berghs, Henri; Cardoso, Rosilene Fressatti

2014-01-01

At the triple border Brazil/Paraguay/Argentina there is easy mobility from one city to another for economic and tourism activities. This constant and fast population mobility is mainly to visit Iguazu Falls, in the Iguazu River, on the border of the Brazilian state of Paraná and the Argentina. As the incidence of tuberculosis is high in this setting, our study aimed to establish a first baseline of circulating genotypic lineages of Mycobacterium tuberculosis. This study included 120 patients from 10 cities in southwestern Paraná, Brazil with pulmonary symptoms, from July 2009 to July 2011. Information about sex, age, clinical features and address was collected by reviewing the national tuberculosis notification database. Of these, 96 (80%) isolates were identified as M. tuberculosis and 22 (22.9%) were drug resistant (20, 20.8% INH mono-resistant and 2, 2.1% multidrug-resistant). All isolates were subjected to genotyping by Spoligotyping and MIRU-VNTR typing. The distribution of the isolates analyzed by spoligotyping revealed 30 distinct patterns. The four mainly detected clades were Latin American and Mediterranean (LAM), ill-defined T, Haarlem (H) and S. The MIRU-VNTR showed 85 distinct patterns. Spoligotyping combined to MIRU-VNTR allowed 90 distinct patterns. Our study demonstrated that there is significant molecular diversity in circulating M. tuberculosis, with predominance of the LAM and T clades in cities of southwestern Paraná, Brazil, bordering Argentina and Paraguay.

First baseline of circulating genotypic lineages of Mycobacterium tuberculosis in patients from the brazilian borders with Argentina and Paraguay.

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Luzia Neri C Machado

Full Text Available BACKGROUND: At the triple border Brazil/Paraguay/Argentina there is easy mobility from one city to another for economic and tourism activities. This constant and fast population mobility is mainly to visit Iguazu Falls, in the Iguazu River, on the border of the Brazilian state of Paraná and the Argentina. As the incidence of tuberculosis is high in this setting, our study aimed to establish a first baseline of circulating genotypic lineages of Mycobacterium tuberculosis. METHODOLOGY/PRINCIPAL FINDINGS: This study included 120 patients from 10 cities in southwestern Paraná, Brazil with pulmonary symptoms, from July 2009 to July 2011. Information about sex, age, clinical features and address was collected by reviewing the national tuberculosis notification database. Of these, 96 (80% isolates were identified as M. tuberculosis and 22 (22.9% were drug resistant (20, 20.8% INH mono-resistant and 2, 2.1% multidrug-resistant. All isolates were subjected to genotyping by Spoligotyping and MIRU-VNTR typing. The distribution of the isolates analyzed by spoligotyping revealed 30 distinct patterns. The four mainly detected clades were Latin American and Mediterranean (LAM, ill-defined T, Haarlem (H and S. The MIRU-VNTR showed 85 distinct patterns. Spoligotyping combined to MIRU-VNTR allowed 90 distinct patterns. CONCLUSIONS/SIGNIFICANCE: Our study demonstrated that there is significant molecular diversity in circulating M. tuberculosis, with predominance of the LAM and T clades in cities of southwestern Paraná, Brazil, bordering Argentina and Paraguay.

A case of Manila type Mycobacterium tuberculosis infection in Japan

Science.gov (United States)

Usami, Osamu; Nakajima, Chie; Endo, Shiro; Inomata, Shinya; Kanamori, Hajime; Hirakata, Yoichi; Uchiyama, Bine; Kaku, Mitsuo; Suzuki, Yasuhiko; Hattori, Toshio

2015-01-01

Key Clinical Message A 76-year-old Japanese woman contracted a Mycobacterium tuberculosis (TB, Manila type) infection in Japan, despite never having traveled. However, her son was treated for TB in the Philippines 3 years before he stayed at her house. Spoligotyping allows us to identify the TB genotype and identify the route of infection. PMID:26273455

Representativeness of Tuberculosis Genotyping Surveillance in the United States, 2009-2010.

Science.gov (United States)

Shak, Emma B; France, Anne Marie; Cowan, Lauren; Starks, Angela M; Grant, Juliana

2015-01-01

Genotyping of Mycobacterium tuberculosis isolates contributes to tuberculosis (TB) control through detection of possible outbreaks. However, 20% of U.S. cases do not have an isolate for testing, and 10% of cases with isolates do not have a genotype reported. TB outbreaks in populations with incomplete genotyping data might be missed by genotyping-based outbreak detection. Therefore, we assessed the representativeness of TB genotyping data by comparing characteristics of cases reported during January 1, 2009-December 31, 2010, that had a genotype result with those cases that did not. Of 22,476 cases, 14,922 (66%) had a genotype result. Cases without genotype results were more likely to be patients <19 years of age, with unknown HIV status, of female sex, U.S.-born, and with no recent history of homelessness or substance abuse. Although cases with a genotype result are largely representative of all reported U.S. TB cases, outbreak detection methods that rely solely on genotyping data may underestimate TB transmission among certain groups.

[Discriminatory power of variable number on tandem repeats loci for genotyping Mycobacterium tuberculosis strains in China].

Science.gov (United States)

Chen, H X; Cai, C; Liu, J Y; Zhang, Z G; Yuan, M; Jia, J N; Sun, Z G; Huang, H R; Gao, J M; Li, W M

2017-06-10

Objective: Using the standard genotype method, variable number of tandem repeats (VNTR), we constructed a VNTR database to cover all provinces and proposed a set of optimized VNTR loci combinations for each province, in order to improve the preventive and control programs on tuberculosis, in China. Methods: A total of 15 loci VNTR was used to analyze 4 116 Mycobacterium tuberculosis strains, isolated from national survey of Drug Resistant Tuberculosis, in 2007. Hunter-Gaston Index (HGI) was also used to analyze the discriminatory power of each VNTR site. A set combination of 12-VNTR, 10-VNTR, 8-VNTR and 5-VNTR was respectively constructed for each province, based on 1) epidemic characteristics of M. tuberculosis lineages in China, with high discriminatory power and genetic stability. Results: Through the completed 15 loci VNTR patterns of 3 966 strains under 96.36 % (3 966/4 116) coverage, we found seven high HGI loci (including QUB11b and MIRU26) as well as low stable loci (including QUB26, MIRU16, Mtub21 and QUB11b) in several areas. In all the 31 provinces, we found an optimization VNTR combination as 10-VNTR loci in Inner Mongolia, Chongqing and Heilongjiang, but with 8-VNTR combination shared in other provinces. Conclusions: It is necessary to not only use the VNTR database for tracing the source of infection and cluster of M. tuberculosis in the nation but also using the set of optimized VNTR combinations in monitoring those local epidemics and M. tuberculosis (genetics in local) population.

Molecular Strain Typing of Mycobacterium tuberculosis: a Review of Frequently Used Methods

Science.gov (United States)

2016-01-01

Tuberculosis, caused by the bacterium Mycobacterium tuberculosis, remains one of the most serious global health problems. Molecular typing of M. tuberculosis has been used for various epidemiologic purposes as well as for clinical management. Currently, many techniques are available to type M. tuberculosis. Choosing the most appropriate technique in accordance with the existing laboratory conditions and the specific features of the geographic region is important. Insertion sequence IS6110-based restriction fragment length polymorphism (RFLP) analysis is considered the gold standard for the molecular epidemiologic investigations of tuberculosis. However, other polymerase chain reaction-based methods such as spacer oligonucleotide typing (spoligotyping), which detects 43 spacer sequence-interspersing direct repeats (DRs) in the genomic DR region; mycobacterial interspersed repetitive units–variable number tandem repeats, (MIRU-VNTR), which determines the number and size of tandem repetitive DNA sequences; repetitive-sequence-based PCR (rep-PCR), which provides high-throughput genotypic fingerprinting of multiple Mycobacterium species; and the recently developed genome-based whole genome sequencing methods demonstrate similar discriminatory power and greater convenience. This review focuses on techniques frequently used for the molecular typing of M. tuberculosis and discusses their general aspects and applications. PMID:27709842

Association between Mycobacterium tuberculosis complex phylogenetic lineage and acquired drug resistance.

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Courtney M Yuen

Full Text Available BACKGROUND: Development of resistance to antituberculosis drugs during treatment (i.e., acquired resistance can lead to emergence of resistant strains and consequent poor clinical outcomes. However, it is unknown whether Mycobacterium tuberculosis complex species and lineage affects the likelihood of acquired resistance. METHODS: We analyzed data from the U.S. National Tuberculosis Surveillance System and National Tuberculosis Genotyping Service for tuberculosis cases during 2004-2011 with assigned species and lineage and both initial and final drug susceptibility test results. We determined univariate associations between species and lineage of Mycobacterium tuberculosis complex bacteria and acquired resistance to isoniazid, rifamycins, fluoroquinolones, and second-line injectables. We used Poisson regression with backward elimination to generate multivariable models for acquired resistance to isoniazid and rifamycins. RESULTS: M. bovis was independently associated with acquired resistance to isoniazid (adjusted prevalence ratio = 8.46, 95% CI 2.96-24.14 adjusting for HIV status, and with acquired resistance to rifamycins (adjusted prevalence ratio = 4.53, 95% CI 1.29-15.90 adjusting for homelessness, HIV status, initial resistance to isoniazid, site of disease, and administration of therapy. East Asian lineage was associated with acquired resistance to fluoroquinolones (prevalence ratio = 6.10, 95% CI 1.56-23.83. CONCLUSIONS: We found an association between mycobacterial species and lineage and acquired drug resistance using U.S. surveillance data. Prospective clinical studies are needed to determine the clinical significance of these findings, including whether rapid genotyping of isolates at the outset of treatment may benefit patient management.

[Rd7 genotyping of M. tuberculosis strains isolated from patients with lung tuberculosis in different areas of Kazakhstan].

Science.gov (United States)

Demkin, V V; Korneva, I N; Riazanova, Iu A; Muminov, T A; Beĭsembaeva, Sh A; Zhakipbaeva, B T; Shopaeva, G A; Dauletbakova, A M

2008-01-01

A three-primer PCR assay was designed for detection of possible deletions in the RD7 region of the Mycobacterium tuberculosis complex chromosome. The assay produced amplicons of different size depending on the presence or absence of the deletions. The PCR assay was applied to 176 isolates from patients with lung tuberculosis collected in different areas of Kazakhstan in summer 2004. The isolates were initially characterized by culture and biochemical tests. The RD7 genotyping results demonstrated no polymorphism and the absence of deletions in the RD7 genome region. Some strains were additionally characterized using PCR-RFLP analysis of gyrB and hsp64 genes. The RFLP-patterns obtained corresponded to the M. tuberculosis genotypes. The results of this work are consistent with certain previous studies, indicating population stability of the RD7 region in M. tuberculosis strains. Species characterization of the isolates shows that M. tuberculosis sensu stricto is the principal causative agent of human lung tuberculosis in Kazakhstan.

Transmission of Mycobacterium tuberculosis from patients who are nucleic acid amplification test- negative.

Science.gov (United States)

Xie, Yingda L; Cronin, Wendy A; Proschan, Michael; Oatis, Richard; Cohn, Silvia; Curry, Scott R; Golub, Jonathan E; Barry Iii, Clifton E; Dorman, Susan E

2018-04-24

Among adults with signs and symptoms of pulmonary tuberculosis (TB), recognition of transmissible TB has implications for airborne infection isolation and public health activities. Sputum smear-negative TB patients account for around one-fifth of tuberculosis transmission. The tuberculosis transmission risk of TB patients with negative results on nucleic acid amplification (NAA) testing of respiratory specimens has not been established. We sought to estimate the tuberculosis transmission risk of NAA test-negative TB patients. We retrospectively reviewed Maryland TB program data from 2004 to 2009 during which NAA testing by the Mycobacterium Tuberculosis Direct Test (MTD) was performed routinely. Patients with sputum Mycobacterium tuberculosis (M.tb) isolates having matching genotypes were assigned to clusters. Transmission sequence was approximated by collection order of individuals' first culture-positive specimens. Minimum transmission risks of NAA (MTD)-negative TB patients and of smear-negative TB patients were estimated based on individuals' positions within clusters. Among 809 patients with culture-confirmed TB, M.tb genotypes were available for 782 (96.7%). For NAA-negative TB patients the minimum transmission risk estimate was 5.1% (95% CI 0-11.4). For smear-negative TB patients the minimum transmission risk estimate was 11.2% (95% CI 7.2-15.3). Minimum transmission risk of NAA-negative TB patients was lower than that of smear-negative TB patients. However, transmission risk of NAA-negative TB patients appears to not be negligible.

Optimal Combination of VNTR Typing for Discrimination of Isolated Mycobacterium tuberculosis in Korea

OpenAIRE

Lee, Jihye; Kang, Heeyoon; Kim, Sarang; Yoo, Heekyung; Kim, Hee Jin; Park, Young Kil

2014-01-01

Background Variable-number tandem repeat (VNTR) typing is a promising method to discriminate the Mycobacterium tuberculosis isolates in molecular epidemiology. The purpose of this study is to determine the optimal VNTR combinations for discriminating isolated M. tuberculosis strains in Korea. Methods A total of 317 clinical isolates collected throughout Korea were genotyped by using the IS6110 restriction fragment length polymorphism (RFLP), and then analysed for the number of VNTR copies fro...

Progression to active tuberculosis, but not transmission, varies by Mycobacterium tuberculosis lineage in The Gambia

NARCIS (Netherlands)

de Jong, Bouke C.; Hill, Philip C.; Aiken, Alex; Awine, Timothy; Antonio, Martin; Adetifa, Ifedayo M.; Jackson-Sillah, Dolly J.; Fox, Annette; Deriemer, Kathryn; Gagneux, Sebastien; Borgdorff, Martien W.; McAdam, Keith P. W. J.; Corrah, Tumani; Small, Peter M.; Adegbola, Richard A.

2008-01-01

BACKGROUND: There is considerable variability in the outcome of Mycobacterium tuberculosis infection. We hypothesized that Mycobacterium africanum was less likely than M. tuberculosis to transmit and progress to tuberculosis disease. METHODS: In a cohort study of patients with tuberculosis and their

Drug Resistance and Population Structure of Mycobacterium tuberculosis Beijing Strains Isolated in Poland.

Science.gov (United States)

Kozińska, Monika; Augustynowicz-Kopeć, Ewa

2015-01-01

In total, 1095 Mycobacterium tuberculosis clinical isolates from 282 patients with drug-resistant and 813 with drug-sensitive tuberculosis (TB) in Poland during 2007-2011 were analysed. Seventy-one (6.5%) patients were found to have strains of Beijing genotype as defined by spoligotyping. The majority of patients were Polish-born; among foreign-born a large proportion came from Chechnya and Vietnam. Analysis showed strong associations between Beijing genotype infection and MDR, pre-XDR and XDR resistance, with a considerable relative risk among new patients, suggesting that this is due to increased spread of drug-resistant strains rather than acquisition of resistance during treatment.

Real-Time Measurement of Host Bioenergetics During Mycobacterium Tuberculosis Infection

Science.gov (United States)

2015-05-01

AWARD NUMBER: W81XWH-13-1-0149 TITLE: “Real-Time Measurement of Host Bioenergetics During Mycobacterium Tuberculosis Infection...successfully adapted metabolic flux analysis using a Seahorse XF96 metabolic flux analyzer to study Mycobacterium tuberculosis energy metabolism in an...Mycobacterium tuberculosis function. In: Systems Biology of Tuberculosis . Editors: J McFadden, D Beste and A Kierzek. 2013. Springer, New York, NY. 2

[Proposal of a five MIRU-VNTR panel to screen clinical isolates of Mycobacterium tuberculosis in Mexico].

Science.gov (United States)

Bolado-Martínez, Enrique; Candia-Plata, Maria Del Carmen; Zenteno-Cuevas, Roberto; Mendoza Damián, Fabiola; Avilés-Acosta, Magali; Álvarez-Hernández, Gerardo

2015-11-01

Tuberculosis is a public health problem across Mexico. This paper aims to select a panel, with a minimum number of repetitive elements (MIRU-VNTR) for genotypic characterization of Mycobacterium tuberculosis (M. tuberculosis) clinical isolates. In this study, a full panel of 24 MIRU-VNTR loci was used to discriminate 65 clinical isolates of M. tuberculosis from three different geographical regions of Mexico. Those loci with the highest discriminatory power were subsequently selected. The panel, including five loci, was obtained by selecting the highest values of allelic diversity among the genotypes obtained. The dendrogram, generated by the panel MIRU-VNTR 5, showed a high discriminatory power with 65 unique genotype profiles and formed clusters according to the geographical region of origin. The panel MIRU-VNTR 5 can be useful for characterizing clinical isolates of M. tuberculosis in Mexico. Copyright © 2014 Elsevier España, S.L.U. y Sociedad Española de Enfermedades Infecciosas y Microbiología Clínica. All rights reserved.

First Insight into a Nationwide Genotypic Diversity of Mycobacterium tuberculosis among Previously Treated Pulmonary Tuberculosis Cases in Benin, West Africa.

Science.gov (United States)

Affolabi, Dissou; Sanoussi, N'Dira; Codo, Sergio; Sogbo, Fréderic; Wachinou, Prudence; Massou, Faridath; Kehinde, Aderemi; Anagonou, Séverin

2017-01-01

Molecular studies on tuberculosis (TB) are rare in low-resource countries like Benin, where data on molecular study on previously treated TB cases is unavailable. From January to December 2014, all smear- and culture-positive previously treated pulmonary TB patients from all TB clinics were systematically recruited. Drug susceptibility testing and spoligotyping were performed on all isolates. Of the 100 patients recruited, 71 (71.0%) were relapse cases and 24 (24.0%) were failure cases, while 5 (5.0%) were default cases. Resistance rate to any first-line drug was 40.0%, while 12.0% of strains were multidrug-resistant (MDR) and no strain was extensively drug-resistant (XDR). A total of 40 distinct spoligotypes were found to be corresponding to a genotypic diversity of 40.0%. ST61 was the most predominant spoligotype with prevalence of 33.0%. In all, 31 single spoligotypes and nine clusters were observed with 2 to 33 strains per cluster giving a clustering rate of 69.0%. Euro-American (Lineage 4) was the most prevalent lineage (74.0%) and Lineage 2 was associated with resistance to streptomycin. This first insight into genetic diversity of previously treated pulmonary TB patients in Benin showed a relatively high genetic diversity of Mycobacterium tuberculosis .

First Insight into a Nationwide Genotypic Diversity of Mycobacterium tuberculosis among Previously Treated Pulmonary Tuberculosis Cases in Benin, West Africa

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Dissou Affolabi

2017-01-01

Full Text Available Background. Molecular studies on tuberculosis (TB are rare in low-resource countries like Benin, where data on molecular study on previously treated TB cases is unavailable. Materials and Methods. From January to December 2014, all smear- and culture-positive previously treated pulmonary TB patients from all TB clinics were systematically recruited. Drug susceptibility testing and spoligotyping were performed on all isolates. Results. Of the 100 patients recruited, 71 (71.0% were relapse cases and 24 (24.0% were failure cases, while 5 (5.0% were default cases. Resistance rate to any first-line drug was 40.0%, while 12.0% of strains were multidrug-resistant (MDR and no strain was extensively drug-resistant (XDR. A total of 40 distinct spoligotypes were found to be corresponding to a genotypic diversity of 40.0%. ST61 was the most predominant spoligotype with prevalence of 33.0%. In all, 31 single spoligotypes and nine clusters were observed with 2 to 33 strains per cluster giving a clustering rate of 69.0%. Euro-American (Lineage 4 was the most prevalent lineage (74.0% and Lineage 2 was associated with resistance to streptomycin. Conclusion. This first insight into genetic diversity of previously treated pulmonary TB patients in Benin showed a relatively high genetic diversity of Mycobacterium tuberculosis.

Correlations between major risk factors and closely related Mycobacterium tuberculosis isolates grouped by three current genotyping procedures: a population-based study in northeast Mexico.

Science.gov (United States)

Peñuelas-Urquides, Katia; Martínez-Rodríguez, Herminia Guadalupe; Enciso-Moreno, José Antonio; Molina-Salinas, Gloria María; Silva-Ramírez, Beatriz; Padilla-Rivas, Gerardo Raymundo; Vera-Cabrera, Lucio; Torres-de-la-Cruz, Víctor Manuel; Martínez-Martínez, Yazmin Berenice; Ortega-García, Jorge Luis; Garza-Treviño, Elsa Nancy; Enciso-Moreno, Leonor; Saucedo-Cárdenas, Odila; Becerril-Montes, Pola; Said-Fernández, Salvador

2014-09-01

The characteristics of tuberculosis (TB) patients related to a chain of recent TB transmissions were investigated. Mycobacterium tuberculosis (MTB) isolates (120) were genotyped using the restriction fragment length polymorphism-IS6110 (R), spacer oligotyping (S) and mycobacterial interspersed repetitive units-variable number of tandem repeats (M) methods. The MTB isolates were clustered and the clusters were grouped according to the similarities of their genotypes. Spearman's rank correlation coefficients between the groups of MTB isolates with similar genotypes and those patient characteristics indicating a risk for a pulmonary TB (PTB) chain transmission were ana- lysed. The isolates showing similar genotypes were distributed as follows: SMR (5%), SM (12.5%), SR (1.67%), MR (0%), S (46.67%), M (5%) and R (0%). The remaining 35 cases were orphans. SMR exhibited a significant correlation (p < 0.05) with visits to clinics, municipalities and comorbidities (primarily diabetes mellitus). S correlated with drug consumption and M with comorbidities. SMR is needed to identify a social network in metropolitan areas for PTB transmission and S and M are able to detect risk factors as secondary components of a transmission chain of TB.

Representativeness of Tuberculosis Genotyping Surveillance in the United States, 2009–2010

Science.gov (United States)

Shak, Emma B.; Cowan, Lauren; Starks, Angela M.; Grant, Juliana

2015-01-01

Genotyping of Mycobacterium tuberculosis isolates contributes to tuberculosis (TB) control through detection of possible outbreaks. However, 20% of U.S. cases do not have an isolate for testing, and 10% of cases with isolates do not have a genotype reported. TB outbreaks in populations with incomplete genotyping data might be missed by genotyping-based outbreak detection. Therefore, we assessed the representativeness of TB genotyping data by comparing characteristics of cases reported during January 1, 2009–December 31, 2010, that had a genotype result with those cases that did not. Of 22,476 cases, 14,922 (66%) had a genotype result. Cases without genotype results were more likely to be patients <19 years of age, with unknown HIV status, of female sex, U.S.-born, and with no recent history of homelessness or substance abuse. Although cases with a genotype result are largely representative of all reported U.S. TB cases, outbreak detection methods that rely solely on genotyping data may underestimate TB transmission among certain groups. PMID:26556930

Mycobacterium tuberculosis infection in cattle from the Eastern Cape Province of South Africa.

Science.gov (United States)

Hlokwe, Tiny Motlatso; Said, Halima; Gcebe, Nomakorinte

2017-10-10

Mycobacterium tuberculosis is the main causative agent of tuberculosis (TB) in human and Mycobacterium bovis commonly causes tuberculosis in animals. Transmission of tuberculosis caused by both pathogens can occur from human to animals and vice versa. In the current study, M. tuberculosis, as confirmed by polymerase chain reaction (PCR) using primers targeting 3 regions of difference (RD4, RD9 and RD12) on the genomes, was isolated from cattle originating from two epidemiologically unrelated farms in the Eastern Cape (E.C) Province of South Africa. Although the isolates were genotyped with variable number of tandem repeat (VNTR) typing, no detailed epidemiological investigation was carried out on the respective farms to unequivocally confirm or link humans as sources of TB transmission to cattle, a move that would have embraced the 'One Health' concept. In addition, strain comparison with human M. tuberculosis in the database from the E.C Province and other provinces in the country did not reveal any match. This is the first report of cases of M. tuberculosis infection in cattle in South Africa. The VNTR profiles of the M. tuberculosis strains identified in the current study will form the basis for creating M. tuberculosis VNTR database for animals including cattle for future epidemiological studies. Our findings however, call for urgent reinforcement of collaborative efforts between the veterinary and the public health services of the country.

Host immunity to Mycobacterium tuberculosis and risk of tuberculosis

DEFF Research Database (Denmark)

Michelsen, Sascha Wilk; Soborg, Bolette; Agger, Else-Marie

2016-01-01

BACKGROUND: Human immune responses to latent Mycobacterium tuberculosis (Mtb) infection (LTBI) may enable individuals to control Mtb infection and halt progression to tuberculosis (TB), a hypothesis applied in several novel TB vaccines. We aimed to evaluate whether immune responses to selected LTBI...

Utility and diagnostic performance of Mycobacterium tuberculosis complex by two immunochromatographic assays as compared with the molecular Genotype assay in Nigeria

Directory of Open Access Journals (Sweden)

Benjamin Thumamo Pokam

2013-01-01

Full Text Available Among the disadvantages of smear microscopy for detection of tuberculosis cases is its inability to differentiate between Mycobacterium tuberculosis (MTB and non-tuberculous mycobacteria (NTM. This study evaluated two, new immunochromatographic assays – Capilia TB-Neo and SD Bioline – on unheated and heated cultures at 80 °C for 30 min respectively for their ability to discriminate between MTB complex and NTM as compared with the molecular Genotype assay. Mycobacteria used in the study were obtained from smear-positive specimens collected from patients at four major hospitals in Cross River State, Nigeria. Capilia TB-Neo and SD Bioline showed sensitivities of 98.8% and 93.8% respectively and 100% specificity for both assays. Heating the isolates did not significantly impact the test performance. Both tests are recommended for use in rapid differentiation of strains isolated in Nigeria.

[Rapid, simple genotyping method by the variable numbers of tandem repeats (VNTR) for Mycobacterium tuberculosis isolates in Japan--analytical procedure of JATA (12)-VNTR].

Science.gov (United States)

Maeda, Shinji; Murase, Yoshiro; Mitarai, Satoshi; Sugawara, Isamu; Kato, Seiya

2008-10-01

The discriminatory power of each locus in variable numbers of tandem repeats (VNTR) analyses was evaluated for development of the genotyping method of Mycobacterium tuberculosis (TB) in Japan. By using 325 TB strains collected from whole Japan and 24 mass infection cases (74 isolates), IS6110 restriction fragment length polymorphism (RFLP), spoligotyping and VNTR (35 loci) were analyzed. We excluded 4 loci (VNTRs 2163a, 3232, 3820, and 4120) and selected in top 12 loci (VNTRs 0424, 0960, 1955, 2074, 2163b, 2372, 2996, 3155, 3192, 3336, 4052, and 4156). The cluster rate of IS6110 RFLP was higher than that of 12-locus [Japan Anti-Tuberculosis Association (JATA)] VNTR. And in comparison of the discriminatory power of 12-locus JATA VNTR and that of Supply (15)-VNTR, the JATA (12)-VNTR was superior, even though less loci analyses. Therefore, this JATA (12)-VNTR could be used for TB genotyping in areas where Beijing strains are prevalent.

Molecular diversity of Mycobacterium tuberculosis isolates from patients with tuberculosis in Honduras

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Ghebremichael Solomon

2010-08-01

Full Text Available Abstract Background Tuberculosis persists as a public health problem in Honduras. A better knowledge of the molecular characteristics of Mycobacterium tuberculosis strains will contribute to understand the transmission dynamics of the disease within the country. The aim of this study was to provide an insight of the genetic biodiversity of M. tuberculosis clinical isolates collected in Honduras between 1994 and 2002. Genotyping was performed using spoligotyping and RFLP. The spoligotypes obtained were compared with the SITVIT2 proprietary database of the Pasteur Institute of Guadeloupe. Results Spoligotyping grouped 84% of the isolates into 27 clusters (2 to 43 strains per cluster. Of the 44 shared international types (SITs identified among the Honduran stains, 8 SITs were newly identified either within the present study or after match with an orphan type previously identified in the SITVIT2 database. In addition, 16 patterns corresponded to orphan, previously unreported isolates. The Latin American Mediterranean (LAM lineage was the most common in this study; 55% of the strains belonged to this family. Other genotypes found were Haarlem (16%, T (16%, X-clade (6%, Unknown signature (5% and S (1%. Only one Beijing strain was identified (0.5%. We observed a high degree of diversity after characterizing the 43 isolates belonging to the main spoligotyping cluster (SIT 33, LAM3 with IS6110-RFLP. A total of 35 different RFLP-fingerprints were detected, of which 6 patterns corresponded to the same number of clusters comprising 14 strains. Conclusions The findings obtained in this study show that tuberculosis transmission in Honduras is due to modern M. tuberculosis lineages with high level of biodiversity.

Molecular diversity of Mycobacterium tuberculosis isolates from patients with tuberculosis in Honduras

Science.gov (United States)

2010-01-01

Background Tuberculosis persists as a public health problem in Honduras. A better knowledge of the molecular characteristics of Mycobacterium tuberculosis strains will contribute to understand the transmission dynamics of the disease within the country. The aim of this study was to provide an insight of the genetic biodiversity of M. tuberculosis clinical isolates collected in Honduras between 1994 and 2002. Genotyping was performed using spoligotyping and RFLP. The spoligotypes obtained were compared with the SITVIT2 proprietary database of the Pasteur Institute of Guadeloupe. Results Spoligotyping grouped 84% of the isolates into 27 clusters (2 to 43 strains per cluster). Of the 44 shared international types (SITs) identified among the Honduran stains, 8 SITs were newly identified either within the present study or after match with an orphan type previously identified in the SITVIT2 database. In addition, 16 patterns corresponded to orphan, previously unreported isolates. The Latin American Mediterranean (LAM) lineage was the most common in this study; 55% of the strains belonged to this family. Other genotypes found were Haarlem (16%), T (16%), X-clade (6%), Unknown signature (5%) and S (1%). Only one Beijing strain was identified (0.5%). We observed a high degree of diversity after characterizing the 43 isolates belonging to the main spoligotyping cluster (SIT 33, LAM3) with IS6110-RFLP. A total of 35 different RFLP-fingerprints were detected, of which 6 patterns corresponded to the same number of clusters comprising 14 strains. Conclusions The findings obtained in this study show that tuberculosis transmission in Honduras is due to modern M. tuberculosis lineages with high level of biodiversity. PMID:20678242

Tuberculosis transmission of predominant genotypes of Mycobacterium tuberculosis in northern suburbs of Buenos Aires city region.

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Morcillo, N; Zumarraga, M; Imperiale, B; Di Giulio, B; Chirico, C; Kuriger, A; Alito, A; Kremer, K; Cataldi, A

2007-01-01

In 2003, the incidence of tuberculosis in Argentina showed an increase compared to 2002. The severe national crisis at the end of the 90s has probably strongly contributed to this situation. The goal of this work was to estimate the extent of the spread of the most predominant Mycobacterium tuberculosis strains and to assess the spread of predominant M. tuberculosis clusters as determined by spoligotyping and IS6110 RFLP. The study involved 590 pulmonary, smear-positive TB cases receiving medical attention at health centers and hospitals in Northern Buenos Aires (NBA) suburbs, from October 2001 to December 2002. From a total of 208 clinical isolates belonging to 6 major clusters, 63 (30.2%) isolates had identical spoligotyping and IS6110 RFLP pattern. Only 22.2% were shown to have epidemiological connections with another member of their respective cluster. In these major clusters, 30.2% of the 208 TB cases studied by both molecular techniques and contact tracing could be convincingly attributable to a recently acquired infection. This knowledge may be useful to assess the clonal distribution of predominant M. tuberculosis clusters in Argentina, which may make an impact on TB control strategies.

Comparison between RFLP and MIRU-VNTR genotyping of Mycobacterium tuberculosis strains isolated in Stockholm 2009 to 2011.

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Jonsson, Jerker; Hoffner, Sven; Berggren, Ingela; Bruchfeld, Judith; Ghebremichael, Solomon; Pennhag, Alexandra; Groenheit, Ramona

2014-01-01

Our aim was to analyze the difference between methods for genotyping of Mycobacterium tuberculosis complex isolates. We collected genotyping results from Restriction Fragment Length Polymorphism (RFLP) and Mycobacterial Interspersed Repetitive Units-Variable Numbers of Tandem Repeat (MIRU-VNTR) in a geographically limited area (Stockholm) during a period of three years. The number and proportion of isolates belonging to clusters was reduced by 45 and 35% respectively when combining the two methods compared with using RFLP or MIRU-VNTR only. The mean size of the clusters was smaller when combining methods and smaller with RFLP compared to MIRU-VNTR. In clusters with confirmed epidemiological links RFLP coincided slightly better than MIRU-VNTR but where there was a difference, the variation in MIRU-VNTR pattern was only in a single locus. In isolates with few IS6110 bands in RFLP, MIRU-VNTR differentiated the isolates more, dividing the RFLP clusters. Since MIRU-VNTR is faster and less labour-intensive it is the method of choice for routine genotyping. In most cases it will be sufficient for epidemiological purposes but true clustering might still be considered if there are epidemiological links and the MIRU-VNTR results differ in only one of its 24 loci.

A Case of False-Positive Mycobacterium tuberculosis Caused by Mycobacterium celatum

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Edward Gildeh

2016-01-01

Full Text Available Mycobacterium celatum is a nontuberculous mycobacterium shown to cause symptoms similar to pulmonary M. tuberculosis. Certain strains have been shown to cross-react with the probes used to detect M. tuberculosis, making this a diagnostic challenge. We present a 56-year-old gentleman who developed signs and symptoms of lung infection with computed tomography scan of the chest showing right lung apex cavitation. Serial sputum samples were positive for acid-fast bacilli and nucleic acid amplification testing identified M. tuberculosis ribosomal RNA, resulting in treatment initiation. Further testing with high performance liquid chromatography showed a pattern consistent with M. celatum. This case illustrates the potential for M. celatum to mimic M. tuberculosis in both its clinical history and laboratory testing due to the identical oligonucleotide sequence contained in both. An increasing number of case reports suggest that early reliable differentiation could reduce unnecessary treatment and public health intervention associated with misdiagnosed tuberculosis.

Mycobacterium tuberculosis monoarthritis in a child

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Rosenberg Alan M

2008-09-01

Full Text Available Abstract A child with isolated Mycobacterium tuberculosis monoarthritis, with features initially suggesting oligoarthritis subtype of juvenile idiopathic arthritis, is presented. This patient illustrates the need to consider the possibility of tuberculosis as the cause of oligoarthritis in high-risk pediatric populations even in the absence of a tuberculosis contact history and without evidence of overt pulmonary disease.

First insights into circulating Mycobacterium tuberculosis complex lineages and drug resistance in Guinea

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Ejo, Mebrat; Gehre, Florian; Barry, Mamadou Dian; Sow, Oumou; Bah, Nene Mamata; Camara, Mory; Bah, Boubacar; Uwizeye, Cecile; Nduwamahoro, Elie; Fissette, Kristina; Rijk, Pim De; Merle, Corinne; Olliaro, Piero; Burgos, Marcos; Lienhardt, Christian; Rigouts, Leen; de Jong, Bouke C.

2015-01-01

In this study we assessed first-line anti-tuberculosis drug resistance and the genotypic distribution of Mycobacterium tuberculosis complex (MTBC) isolates that had been collected from consecutive new tuberculosis patients enrolled in two clinical trials conducted in Guinea between 2005 and 2010. Among the total 359 MTBC strains that were analyzed in this study, 22.8% were resistant to at least one of the first line anti-tuberculosis drugs, including 2.5% multidrug resistance and 17.5% isoniazid resistance, with or without other drugs. In addition, further characterization of isolates from a subset of the two trials (n = 184) revealed a total of 80 different spoligotype patterns, 29 “orphan” and 51 shared patterns. We identified the six major MTBC lineages of human relevance, with predominance of the Euro-American lineage. In total, 132 (71.7%) of the strains were genotypically clustered, and further analysis (using the DESTUS model) suggesting significantly faster spread of LAM10_CAM family (p = 0.00016). In conclusion, our findings provide a first insight into drug resistance and the population structure of the MTBC in Guinea, with relevance for public health scientists in tuberculosis control programs. PMID:26004194

Phenotypic and genomic comparison of Mycobacterium aurum and surrogate model species to Mycobacterium tuberculosis: implications for drug discovery.

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Namouchi, Amine; Cimino, Mena; Favre-Rochex, Sandrine; Charles, Patricia; Gicquel, Brigitte

2017-07-13

Tuberculosis (TB) is caused by Mycobacterium tuberculosis and represents one of the major challenges facing drug discovery initiatives worldwide. The considerable rise in bacterial drug resistance in recent years has led to the need of new drugs and drug regimens. Model systems are regularly used to speed-up the drug discovery process and circumvent biosafety issues associated with manipulating M. tuberculosis. These include the use of strains such as Mycobacterium smegmatis and Mycobacterium marinum that can be handled in biosafety level 2 facilities, making high-throughput screening feasible. However, each of these model species have their own limitations. We report and describe the first complete genome sequence of Mycobacterium aurum ATCC23366, an environmental mycobacterium that can also grow in the gut of humans and animals as part of the microbiota. This species shows a comparable resistance profile to that of M. tuberculosis for several anti-TB drugs. The aims of this study were to (i) determine the drug resistance profile of a recently proposed model species, Mycobacterium aurum, strain ATCC23366, for anti-TB drug discovery as well as Mycobacterium smegmatis and Mycobacterium marinum (ii) sequence and annotate the complete genome sequence of this species obtained using Pacific Bioscience technology (iii) perform comparative genomics analyses of the various surrogate strains with M. tuberculosis (iv) discuss how the choice of the surrogate model used for drug screening can affect the drug discovery process. We describe the complete genome sequence of M. aurum, a surrogate model for anti-tuberculosis drug discovery. Most of the genes already reported to be associated with drug resistance are shared between all the surrogate strains and M. tuberculosis. We consider that M. aurum might be used in high-throughput screening for tuberculosis drug discovery. We also highly recommend the use of different model species during the drug discovery screening process.

Mycobacterium tuberculosis Metabolism

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Warner, Digby F.

2015-01-01

Metabolism underpins the physiology and pathogenesis of Mycobacterium tuberculosis. However, although experimental mycobacteriology has provided key insights into the metabolic pathways that are essential for survival and pathogenesis, determining the metabolic status of bacilli during different stages of infection and in different cellular compartments remains challenging. Recent advances—in particular, the development of systems biology tools such as metabolomics—have enabled key insights into the biochemical state of M. tuberculosis in experimental models of infection. In addition, their use to elucidate mechanisms of action of new and existing antituberculosis drugs is critical for the development of improved interventions to counter tuberculosis. This review provides a broad summary of mycobacterial metabolism, highlighting the adaptation of M. tuberculosis as specialist human pathogen, and discusses recent insights into the strategies used by the host and infecting bacillus to influence the outcomes of the host–pathogen interaction through modulation of metabolic functions. PMID:25502746

The Genotype MTBDRplus ver. 2.0 test as a quick indicator of resistance to rifampicin and isoniazid in Mycobacterium tuberculosis strains

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Salvatore Nisticò

2013-08-01

Full Text Available Tuberculosis is still a global emergency and a major public health problem, in some cases related to the appearance of strains of multi drug resistance (MDR and extensive drug resistance (XDR Mycobacterium tuberculosis complex.The correct determination of antibiotic sensitivity profiles is therefore crucial to carry out appropriate treatment aimed to decrease the infectivity of each patient and to reduce mortality. The poor adherence to treatment by the patient or the use of therapies based on a single drug, as a result of incorrect requirements, promote the development of drug-resistance. Have some time on the market of molecular diagnostic tests that allow, quickly and directly from biological sample to search for resistance genes some key drugs of anti-TB therapy (Rifampicin and Isoniazid. One of the tests in question is the Genotype MTBDRplus ver 2.0 which can reveal the presence of genes for resistance to Isoniazid (INH and Rifampin (RMP.The loci analyzed are those corresponding to the rpoB gene for rifampicin, katG and inhA for isoniazid. Our study is based on the analysis of 83 strains of tubercular Mycobacteria identified and isolated from patients with tuberculosis disease and subjected to the tests sensitivity, searching for mutations and phenotypic susceptibility testing for Rifampicin and Isoniazid.The comparison of the results has shown that the results obtained using the Genotype MTBDRplus ver 2.0 test, were similar to the results obtained by the traditional susceptibility testing.

Genotypic characterization of multi-drug-resistant Mycobacterium tuberculosis isolates in Myanmar.

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Aye, Khin Saw; Nakajima, Chie; Yamaguchi, Tomoyuki; Win, Min Min; Shwe, Mu Mu; Win, Aye Aye; Lwin, Thandar; Nyunt, Wint Wint; Ti, Ti; Suzuki, Yasuhiko

2016-03-01

The number of multi-drug-resistant tuberculosis (MDR-TB) cases is rising worldwide. As a countermeasure against this situation, the implementation of rapid molecular tests to identify MDR-TB would be effective. To develop such tests, information on the frequency and distribution of mutations associating with phenotypic drug resistance in Mycobacterium tuberculosis is required in each country. During 2010, the common mutations in the rpoB, katG and inhA of 178 phenotypically MDR M. tuberculosis isolates collected by the National Tuberculosis Control Program (NTP) in Myanmar were investigated by DNA sequencing. Mutations affecting the 81-bp rifampicin (RIF) resistance-determining region (RRDR) of the rpoB were identified in 127 of 178 isolates (71.3%). Two of the most frequently affected codons were 531 and 526, with percentages of 48.3% and 14.0% respectively. For isoniazid (INH) resistance, 114 of 178 MDR-TB isolates (64.0%) had mutations in the katG in which a mutation-conferring amino acid substitution at codon 315 from Ser to Thr was the most common. Mutations in the inhA regulatory region were also detected in 20 (11.2%) isolates, with the majority at position -15. Distinct mutation rate and pattern from surrounding countries might suggest that MDR-TB has developed and spread domestically in Myanmar. Copyright © 2015 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

Characterization of drug susceptibility of Mycobacterium tuberculosis isolated from new cases of tuberculosis concurrent with HIV infection

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G. V. Panov

2015-01-01

Full Text Available The paper characterizes drug susceptibility in Mycobacterium tuberculosis isolated from new cases of tuberculosis concurrent with HIV infection. The investigators have studied the spectrum of drug resistance in Mycobacterium tuberculosis isolated from new cases of tuberculosis concurrent with and without HIV infection (172 and 309 clinical isolates, respectively. There are differences in the rate of primary drug resistance to antituberculosis drugs in patients with and without HIV infection (59 and 43.5% of the cases, respectively. The HIV-infected have also shown high rifampicin resistance rates in Mycobacterium tuberculosis (41.7%. The reasons for these differences are as yet unknown and call for further investigation.

[Clustering analysis of Mycobacterium tuberculosis using the JATA(12)-VNTR system for molecular epidemiological surveillance in broad areas of Japan].

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Wada, Takayuki; Tamaru, Aki; Iwamoto, Tomotada; Arikawa, Kentaro; Nakanishi, Noriko; Komukai, Jun; Matsumoto, Kenji; Hase, Atsushi

2013-04-01

Japan Anti-Tuberculosis Association (JATA) (12)-variable numbers of tandem repeats (VNTR) is a standard method for genotyping of clinical isolates of Mycobacterium tuberculosis in Japan. As a model study for nationwide surveillance, this study aimed to describe the tendency and frequency of genotypes of M. tuberculosis in a large number of clinical samples. Clinical isolates of M. tuberculosis (n = 1,778) were obtained from patients with tuberculosis in 3 areas, i.e., Osaka City, Osaka Prefecture, and Kobe City, during 2007 and 2008. The samples were analyzed using JATA (12)-VNTR. All genotypes were subjected to clustering analysis. In total, 1,086 (61.1%) isolates showed clustering. The most common clusters were composed of 3 members. Such clusters were considered to reflect either actual transmission or low discriminatory power of JATA (12)-VNTR. Several prevalent JATA(12)-VNTR genotypes formed large clusters and were discussed in relation with epidemiological findings of other studies. The findings of this study will aid in the construction of an effective genotyping-based surveillance system of M. tuberculosis, through improvement of interpretation of VNTR types, observation of certain particular strains in an area, and efficient detection of unidentified outbreaks.

Rapid Detection of Cell-Free Mycobacterium tuberculosis DNA in Tuberculous Pleural Effusion.

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Che, Nanying; Yang, Xinting; Liu, Zichen; Li, Kun; Chen, Xiaoyou

2017-05-01

Tuberculous pleurisy is one of the most common types of extrapulmonary tuberculosis, but its diagnosis remains difficult. In this study, we report for the first time on the detection of cell-free Mycobacterium tuberculosis DNA in pleural effusion and an evaluation of a newly developed molecular assay for the detection of cell-free Mycobacterium tuberculosis DNA. A total of 78 patients with pleural effusion, 60 patients with tuberculous pleurisy, and 18 patients with alternative diseases were included in this study. Mycobacterial culture, the Xpert MTB/RIF assay, the adenosine deaminase assay, the T-SPOT.TB assay, and the cell-free Mycobacterium tuberculosis DNA assay were performed on all the pleural effusion samples. The cell-free Mycobacterium tuberculosis DNA assay and adenosine deaminase assay showed significantly higher sensitivities of 75.0% and 68.3%, respectively, than mycobacterial culture and the Xpert MTB/RIF assay, which had sensitivities of 26.7% and 20.0%, respectively ( P pleural effusion showed the highest sensitivity of 95.0% but the lowest specificity of 38.9%. The cell-free Mycobacterium tuberculosis DNA assay detected as few as 1.25 copies of IS 6110 per ml of pleural effusion and showed good accordance of the results between repeated tests ( r = 0.978, P = 2.84 × 10 -10 ). These data suggest that the cell-free Mycobacterium tuberculosis DNA assay is a rapid and accurate molecular test which provides direct evidence of Mycobacterium tuberculosis etiology. Copyright © 2017 American Society for Microbiology.

The draft genome of Mycobacterium aurum, a potential model organism for investigating drugs against Mycobacterium tuberculosis and Mycobacterium leprae

KAUST Repository

Phelan, Jody; Maitra, Arundhati; McNerney, Ruth; Nair, Mridul; Gupta, Antima; Coll, Francesc; Pain, Arnab; Bhakta, Sanjib; Clark, Taane G.

2015-01-01

Mycobacterium aurum (M. aurum) is an environmental mycobacteria that has previously been used in studies of anti-mycobacterial drugs due to its fast growth rate and low pathogenicity. The M. aurum genome has been sequenced and assembled into 46 contigs, with a total length of 6.02 Mb containing 5684 annotated protein-coding genes. A phylogenetic analysis using whole genome alignments positioned M. aurum close to Mycobacterium vaccae and Mycobacterium vanbaalenii, within a clade related to fast-growing mycobacteria. Large-scale genomic rearrangements were identified by comparing the M. aurum genome to those of Mycobacterium tuberculosis and Mycobacterium leprae. M. aurum orthologous genes implicated in resistance to anti-tuberculosis drugs in M. tuberculosis were observed. The sequence identity at the DNA level varied from 68.6% for pncA (pyrazinamide drug-related) to 96.2% for rrs (streptomycin, capreomycin). We observed two homologous genes encoding the catalase-peroxidase enzyme (katG) that is associated with resistance to isoniazid. Similarly, two embB homologues were identified in the M. aurum genome. In addition to describing for the first time the genome of M. aurum, this work provides a resource to aid the use of M. aurum in studies to develop improved drugs for the pathogenic mycobacteria M. tuberculosis and M. leprae.

The draft genome of Mycobacterium aurum, a potential model organism for investigating drugs against Mycobacterium tuberculosis and Mycobacterium leprae

KAUST Repository

Phelan, Jody

2015-06-04

Mycobacterium aurum (M. aurum) is an environmental mycobacteria that has previously been used in studies of anti-mycobacterial drugs due to its fast growth rate and low pathogenicity. The M. aurum genome has been sequenced and assembled into 46 contigs, with a total length of 6.02 Mb containing 5684 annotated protein-coding genes. A phylogenetic analysis using whole genome alignments positioned M. aurum close to Mycobacterium vaccae and Mycobacterium vanbaalenii, within a clade related to fast-growing mycobacteria. Large-scale genomic rearrangements were identified by comparing the M. aurum genome to those of Mycobacterium tuberculosis and Mycobacterium leprae. M. aurum orthologous genes implicated in resistance to anti-tuberculosis drugs in M. tuberculosis were observed. The sequence identity at the DNA level varied from 68.6% for pncA (pyrazinamide drug-related) to 96.2% for rrs (streptomycin, capreomycin). We observed two homologous genes encoding the catalase-peroxidase enzyme (katG) that is associated with resistance to isoniazid. Similarly, two embB homologues were identified in the M. aurum genome. In addition to describing for the first time the genome of M. aurum, this work provides a resource to aid the use of M. aurum in studies to develop improved drugs for the pathogenic mycobacteria M. tuberculosis and M. leprae.

The draft genome of Mycobacterium aurum , a potential model organism for investigating drugs against Mycobacterium tuberculosis and Mycobacterium leprae

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Jody Phelan

2015-01-01

Full Text Available Mycobacterium aurum (M. aurum is an environmental mycobacteria that has previously been used in studies of anti-mycobacterial drugs due to its fast growth rate and low pathogenicity. The M. aurum genome has been sequenced and assembled into 46 contigs, with a total length of 6.02 Mb containing 5684 annotated protein-coding genes. A phylogenetic analysis using whole genome alignments positioned M. aurum close to Mycobacterium vaccae and Mycobacterium vanbaalenii, within a clade related to fast-growing mycobacteria. Large-scale genomic rearrangements were identified by comparing the M. aurum genome to those of Mycobacterium tuberculosis and Mycobacterium leprae. M. aurum orthologous genes implicated in resistance to anti-tuberculosis drugs in M. tuberculosis were observed. The sequence identity at the DNA level varied from 68.6% for pncA (pyrazinamide drug-related to 96.2% for rrs (streptomycin, capreomycin. We observed two homologous genes encoding the catalase-peroxidase enzyme (katG that is associated with resistance to isoniazid. Similarly, two emb B homologues were identified in the M. aurum genome. In addition to describing for the first time the genome of M. aurum , this work provides a resource to aid the use of M. aurum in studies to develop improved drugs for the pathogenic mycobacteria M. tuberculosis and M. leprae.

SUSCEPTIBILITY OF RIFAMPICIN-ISONIAZID RESISTANT MYCOBACTERIUM TUBERCULOSIS ISOLATES AGAINST LEVOFLOXACIN

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A. H. Kurniawan

2016-01-01

Full Text Available Background: Tuberculosis (TB is a high burden disease in Indonesia with multidrug-resistant (MDR TB incidence started to increase. Treatment success of MDR-TB globally was low in number than it was targeted which was especially caused by fluoroquinolone resistance. One of the fluoroquinolone is levofloxacin, an antibiotic that has been widely used irrationally as antimicrobial treatment. Therefore, this study investigated the sensitivity and MBC of MDR Mycobacterium tuberculosis isolates against Levofloxacin. Method: The susceptibility test for MDR-Mycobacterium tuberculosis on levofloxacin by standard method with levofloxacin were on concentrations 0,5 μg/ml, 1 μg/ml, and 2 μg/ml. Sample of 8 strains MDR-Mycobacterium tuberculosis were cultured with each concentrations on Middlebrook 7H9 for 1 week incubation. Next, each of the incubated concentration was subcultured on solid media Middlebrook 7H10 for 3 weeks incubation. Colonized agar plates after 3 weeks incubation were confirmed with acid-fast stain. Results: On MB 7H10 with levofloxacin concentration 2 μg/ml showed bactericidal effect 100% by no MDR Mycobacterium tuberculosis colony grew (0/8 while the MB 7H10 with levofloxacin concentration 1 μg/ml and 0,5 μg/ml showed the bactericidal effect 37,5% and 25% respectively. The colonized agar plate implied that the MDR Mycobacterium tuberculosis with levofloxacin concentration 1 μg/ml (5/8 and 0,5 μg/ml (6/8 grew well. Conclusion: Levofloxacin concentration 2 μg/ml was susceptible on MDR Mycobacterium tuberculosis. The concentration 2 μg/ml of levofloxacin could be considered as MBC.

Molecular characterization of Mycobacterium tuberculosis isolates from elephants of Nepal.

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Paudel, Sarad; Mikota, Susan K; Nakajima, Chie; Gairhe, Kamal P; Maharjan, Bhagwan; Thapa, Jeewan; Poudel, Ajay; Shimozuru, Michito; Suzuki, Yasuhiko; Tsubota, Toshio

2014-05-01

Mycobacterium tuberculosis was cultured from the lung tissues of 3 captive elephants in Nepal that died with extensive lung lesions. Spoligotyping, TbD1 detection and multi-locus variable number of tandem repeat analysis (MLVA) results suggested 3 isolates belonged to a specific lineage of Indo-Oceanic clade, EAI5 SIT 138. One of the elephant isolates had a new synonymous single nucleotide polymorphism (SNP) T231C in the gyrA sequence, and the same SNP was also found in human isolates in Nepal. MLVA results and transfer history of the elephants suggested that 2 of them might be infected with M. tuberculosis from the same source. These findings indicated the source of M. tuberculosis infection of those elephants were local residents, presumably their handlers. Further investigation including detailed genotyping of elephant and human isolates is needed to clarify the infection route and eventually prevent the transmission of tuberculosis to susceptible hosts. Copyright © 2014 Elsevier Ltd. All rights reserved.

Human Tuberculosis Caused by Mycobacterium bovis in the United States, 2006-2013.

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Scott, Colleen; Cavanaugh, Joseph S; Pratt, Robert; Silk, Benjamin J; LoBue, Philip; Moonan, Patrick K

2016-09-01

Using genotyping techniques that have differentiated Mycobacterium bovis from Mycobacterium tuberculosis since 2005, we review the epidemiology of human tuberculosis caused by M. bovis in the United States and validate previous findings nationally. All tuberculosis cases with a genotyped M. tuberculosis complex isolate reported during 2006-2013 in the United States were eligible for analysis. We used binomial regression to identify characteristics independently associated with M. bovis disease using adjusted prevalence ratios (aPRs) and corresponding 95% confidence intervals (CIs). During 2006-2013, the annual percentages of tuberculosis cases attributable to M. bovis remained consistent nationally (range, 1.3%-1.6%) among all tuberculosis cases (N = 59 273). Compared with adults 25-44 years of age, infants aged 0-4 years (aPR, 1.9 [95% CI, 1.4-2.8]) and children aged 5-14 years (aPR, 4.0 [95% CI, 3.1-5.3]) had higher prevalences of M. bovis disease. Patients who were foreign-born (aPR, 1.4 [95% CI, 1.2-1.7]), Hispanic (aPR, 3.9 [95% CI, 3.0-5.0]), female (aPR, 1.4 [95% CI, 1.3-1.6]), and resided in US-Mexico border counties (aPR, 2.0 [95% CI, 1.7-2.4]) also had higher M. bovis prevalences. Exclusively extrapulmonary disease (aPR, 3.7 [95% CI, 3.3-4.2]) or disease that was both pulmonary and extrapulmonary (aPR, 2.4 [95% CI, 2.1-2.9]) were associated with a higher prevalence of M. bovis disease. Children, foreign-born persons, Hispanics, and females are disproportionately affected by M. bovis, which was independently associated with extrapulmonary disease. Targeted prevention efforts aimed at Hispanic mothers and caregivers are warranted. Published by Oxford University Press for the Infectious Diseases Society of America 2016. This work is written by (a) US Government employee(s) and is in the public domain in the US.

A Mycobacterium tuberculosis cluster demonstrating the use of genotyping in urban tuberculosis control

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Burdo Conny CA

2009-09-01

Full Text Available Abstract Background DNA fingerprinting of Mycobacterium tuberculosis isolates offers better opportunities to study links between tuberculosis (TB cases and can highlight relevant issues in urban TB control in low-endemic countries. Methods A medium-sized molecular cluster of TB cases with identical DNA fingerprints was used for the development of a visual presentation of epidemiologic links between cases. Results Of 32 cases, 17 (53% were linked to the index case, and 11 (34% to a secondary case. The remaining four (13% could not be linked and were classified as possibly caused by the index patient. Of the 21 cases related to the index case, TB developed within one year of the index diagnosis in 11 patients (52%, within one to two years in four patients (19%, and within two to five years in six patients (29%. Conclusion Cluster analysis underscored several issues for TB control in an urban setting, such as the recognition of the outbreak, the importance of reinfections, the impact of delayed diagnosis, the contribution of pub-related transmissions and its value for decision-making to extend contact investigations. Visualising cases in a cluster diagram was particularly useful in finding transmission locations and the similarities and links between patients.

Genotyping and drug susceptibility testing of mycobacterial isolates from population-based tuberculosis prevalence survey in Ghana.

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Addo, Kennedy Kwasi; Addo, Samuel Ofori; Mensah, Gloria Ivy; Mosi, Lydia; Bonsu, Frank Adae

2017-12-02

Mycobacterium tuberculosis complex (MTBC) and Non-tuberculosis Mycobacterium (NTM) infections differ clinically, making rapid identification and drug susceptibility testing (DST) very critical for infection control and drug therapy. This study aims to use World Health Organization (WHO) approved line probe assay (LPA) to differentiate mycobacterial isolates obtained from tuberculosis (TB) prevalence survey in Ghana and to determine their drug resistance patterns. A retrospective study was conducted whereby a total of 361 mycobacterial isolates were differentiated and their drug resistance patterns determined using GenoType Mycobacterium Assays: MTBC and CM/AS for differentiating MTBC and NTM as well MTBDRplus and NTM-DR for DST of MTBC and NTM respectively. Out of 361 isolates, 165 (45.7%) MTBC and 120 (33.2%) NTM (made up of 14 different species) were identified to the species levels whiles 76 (21.1%) could not be completely identified. The MTBC comprised 161 (97.6%) Mycobacterium tuberculosis and 4 (2.4%) Mycobacterium africanum. Isoniazid and rifampicin monoresistant MTBC isolates were 18/165 (10.9%) and 2/165(1.2%) respectively whiles 11/165 (6.7%) were resistant to both drugs. Majority 42/120 (35%) of NTM were M. fortuitum. DST of 28 M. avium complex and 8 M. abscessus complex species revealed that all were susceptible to macrolides (clarithromycin, azithromycin) and aminoglycosides (kanamycin, amikacin, and gentamicin). Our research signifies an important contribution to TB control in terms of knowledge of the types of mycobacterium species circulating and their drug resistance patterns in Ghana.

Role of genotype® mycobacterium common mycobacteria/additional species assay for rapid differentiation between Mycobacterium tuberculosis complex and different species of non-tuberculous mycobacteria

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Amresh Kumar Singh

2013-01-01

Full Text Available Background: Mycobacterium tuberculosis complex (MTBC and non-tuberculous mycobacteria (NTM may or may not have same clinical presentations, but the treatment regimens are always different. Laboratory differentiation between MTBC and NTM by routine methods are time consuming and cumbersome to perform. We have evaluated the role of GenoType® Mycobacterium common mycobacteria/additional species (CM/AS assay for differentiation between MTBC and different species of NTM in clinical isolates from tuberculosis (TB cases. Materials and Methods: A total of 1080 clinical specimens were collected from January 2010 to June 2012. Diagnosis was performed by Ziehl-Neelsen staining followed by culture in BacT/ALERT 3D system (bioMerieux, France. A total of 219 culture positive clinical isolates (BacT/ALERT® MP cultures were selected for differentiation by p-nitrobenzoic acid (PNB sensitivity test as and BIO-LINE SD Ag MPT64 TB test considering as the gold standard test. Final identification and differentiation between MTBC and different species of NTM were further confirmed by GenoType® Mycobacterium CM/AS assay (Hain Lifescience, Nehren, Germany. Results: Out of 219 BacT/ALERT® MP culture positive isolates tested by PNB as 153 MTBC (69.9% and by GenoType® Mycobacterium CM/AS assay as 159 (72.6% MTBC and remaining 60 (27.4% were considered as NTM species. The GenoType® Mycobacterium CM/AS assay was proved 99.3% sensitive and 98.3% specific for rapid differentiation of MTBC and NTM. The most common NTM species were; Mycobacterium fortuitum 20 (33.3% among rapid growing mycobacteria and Mycobacterium intracellulare 11 (18.3% among slow growing mycobacteria. Conclusion: The GenoType® Mycobacterium assay makes rapid and accurate identification of NTM species as compared with different phenotypic and molecular diagnostic tool and helps in management of infections caused by different mycobacteria.

Evaluation of Genetic Pattern of Non-Tuberculosis Mycobacterium Using VNTR Method

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Noorozi J

2011-06-01

Full Text Available Background and Objectives: Epidemiological studies of Non-tuberculosis Mycobacterium is important because of the drug resistance pattern and worldwide dissemination of these organisms. One of genetic fingerprinting methods for epidemiological studies is VNTR (Variable Number Tandem Repeat. In this study genetic pattern of atypical Mycobacterium was evaluated by VNTR method for epidemiologic studies. Methods: 48 pulmonary and non pulmonary specimens separated from patients with the symptoms of pulmonary tuberculosis (PTB and identified as Non-tuberculosis Mycobacteriumby phenotypic and PCR-RFLP methods were selected for this study. Clinical samples and their standard strains were evaluated according to VNTR pattern using the 7 genetic loci including ETR-B. ETR-F. ETR-C. MPTR-A. ETR-A. ETR-E. ETR-D.Results: The results of VNTR method showed that none of the 7 loci had any polymorphism in the standard strains of atypical mycobacterium. Some of these variable number tandem repeat in 42 clinical samples of non-tuberculosis Mycobacterium were polymorphic while the PCR product (for any loci was not found in the remaining 6 specimens. Conclusion: Although the used genetic loci of this study were suitable for epidemiological studies of Mycobacterium tuberculosis, these loci were not able to determine the diversity of genetics of non-tuberculosis Mycobacterium Therefore, it seems necessary that other loci be studied using VNTR method.

Management of latent Mycobacterium tuberculosis infection: WHO guidelines for low tuberculosis burden countries

NARCIS (Netherlands)

Getahun, Haileyesus; Matteelli, Alberto; Abubakar, Ibrahim; Aziz, Mohamed Abdel; Baddeley, Annabel; Barreira, Draurio; den Boon, Saskia; Borroto Gutierrez, Susana Marta; Bruchfeld, Judith; Burhan, Erlina; Cavalcante, Solange; Cedillos, Rolando; Chaisson, Richard; Chee, Cynthia Bin-Eng; Chesire, Lucy; Corbett, Elizabeth; Dara, Masoud; Denholm, Justin; de Vries, Gerard; Falzon, Dennis; Ford, Nathan; Gale-Rowe, Margaret; Gilpin, Chris; Girardi, Enrico; Go, Un-Yeong; Govindasamy, Darshini; D Grant, Alison; Grzemska, Malgorzata; Harris, Ross; Horsburgh, C. Robert; Ismayilov, Asker; Jaramillo, Ernesto; Kik, Sandra; Kranzer, Katharina; Lienhardt, Christian; LoBue, Philip; Lönnroth, Knut; Marks, Guy; Menzies, Dick; Migliori, Giovanni Battista; Mosca, Davide; Mukadi, Ya Diul; Mwinga, Alwyn; Nelson, Lisa; Nishikiori, Nobuyuki; Oordt-Speets, Anouk; Rangaka, Molebogeng Xheedha; Reis, Andreas; Rotz, Lisa; Sandgren, Andreas; Sañé Schepisi, Monica; Schünemann, Holger J.; Sharma, Surender Kumar; Sotgiu, Giovanni; Stagg, Helen R.; Sterling, Timothy R.; Tayeb, Tamara; Uplekar, Mukund; van der Werf, Marieke J.; Vandevelde, Wim; van Kessel, Femke; van't Hoog, Anna; Varma, Jay K.; Vezhnina, Natalia; Voniatis, Constantia; Vonk Noordegraaf-Schouten, Marije; Weil, Diana; Weyer, Karin; Wilkinson, Robert John; Yoshiyama, Takashi; Zellweger, Jean Pierre; Raviglione, Mario

2015-01-01

Latent tuberculosis infection (LTBI) is characterised by the presence of immune responses to previously acquired Mycobacterium tuberculosis infection without clinical evidence of active tuberculosis (TB). Here we report evidence-based guidelines from the World Health Organization for a public health

Comparative genomics of archived pyrazinamide resistant Mycobacterium tuberculosis complex isolates from Uganda

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Bovine tuberculosis is a ‘neglected zoonosis’ and its contribution to the proportion of Mycobacterium tuberculosis complex infections in humans is unknown. A retrospective study on archived Mycobacterium tuberculosis complex (MTC) isolates from a reference laboratory in Uganda was undertaken to iden...

Tuberculosis patients co-infected with Mycobacterium bovis and Mycobacterium tuberculosis in an urban area of Brazil.

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Silva, Marcio Roberto; Rocha, Adalgiza da Silva; da Costa, Ronaldo Rodrigues; de Alencar, Andrea Padilha; de Oliveira, Vania Maria; Fonseca Júnior, Antônio Augusto; Sales, Mariana Lázaro; Issa, Marina de Azevedo; Filho, Paulo Martins Soares; Pereira, Omara Tereza Vianello; dos Santos, Eduardo Calazans; Mendes, Rejane Silva; Ferreira, Angela Maria de Jesus; Mota, Pedro Moacyr Pinto Coelho; Suffys, Philip Noel; Guimarães, Mark Drew Crosland

2013-05-01

In this cross-sectional study, mycobacteria specimens from 189 tuberculosis (TB) patients living in an urban area in Brazil were characterised from 2008-2010 using phenotypic and molecular speciation methods (pncA gene and oxyR pseudogene analysis). Of these samples, 174 isolates simultaneously grew on Löwenstein-Jensen (LJ) and Stonebrink (SB)-containing media and presented phenotypic and molecular profiles of Mycobacterium tuberculosis, whereas 12 had molecular profiles of M. tuberculosis based on the DNA analysis of formalin-fixed paraffin wax-embedded tissue samples (paraffin blocks). One patient produced two sputum isolates, the first of which simultaneously grew on LJ and SB media and presented phenotypic and molecular profiles of M. tuberculosis, and the second of which only grew on SB media and presented phenotypic profiles of Mycobacterium bovis. One patient provided a bronchial lavage isolate, which simultaneously grew on LJ and SB media and presented phenotypic and molecular profiles of M. tuberculosis, but had molecular profiles of M. bovis from paraffin block DNA analysis, and one sample had molecular profiles of M. tuberculosis and M. bovis identified from two distinct paraffin blocks. Moreover, we found a low prevalence (1.6%) of M. bovis among these isolates, which suggests that local health service procedures likely underestimate its real frequency and that it deserves more attention from public health officials.

Tuberculosis patients co-infected with Mycobacterium bovis and Mycobacterium tuberculosis in an urban area of Brazil

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Marcio Roberto Silva

2013-05-01

Full Text Available In this cross-sectional study, mycobacteria specimens from 189 tuberculosis (TB patients living in an urban area in Brazil were characterised from 2008-2010 using phenotypic and molecular speciation methods (pncA gene and oxyR pseudogene analysis. Of these samples, 174 isolates simultaneously grew on Löwenstein-Jensen (LJ and Stonebrink (SB-containing media and presented phenotypic and molecular profiles of Mycobacterium tuberculosis, whereas 12 had molecular profiles of M. tuberculosis based on the DNA analysis of formalin-fixed paraffin wax-embedded tissue samples (paraffin blocks. One patient produced two sputum isolates, the first of which simultaneously grew on LJ and SB media and presented phenotypic and molecular profiles of M. tuberculosis, and the second of which only grew on SB media and presented phenotypic profiles of Mycobacterium bovis. One patient provided a bronchial lavage isolate, which simultaneously grew on LJ and SB media and presented phenotypic and molecular profiles of M. tuberculosis, but had molecular profiles of M. bovis from paraffin block DNA analysis, and one sample had molecular profiles of M. tuberculosis and M. bovis identified from two distinct paraffin blocks. Moreover, we found a low prevalence (1.6% of M. bovis among these isolates, which suggests that local health service procedures likely underestimate its real frequency and that it deserves more attention from public health officials.

The draft genome of Mycobacterium aurum, a potential model organism for investigating drugs against Mycobacterium tuberculosis and Mycobacterium leprae.

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Phelan, Jody; Maitra, Arundhati; McNerney, Ruth; Nair, Mridul; Gupta, Antima; Coll, Francesc; Pain, Arnab; Bhakta, Sanjib; Clark, Taane G

2015-09-01

Mycobacterium aurum (M. aurum) is an environmental mycobacteria that has previously been used in studies of anti-mycobacterial drugs due to its fast growth rate and low pathogenicity. The M. aurum genome has been sequenced and assembled into 46 contigs, with a total length of 6.02Mb containing 5684 annotated protein-coding genes. A phylogenetic analysis using whole genome alignments positioned M. aurum close to Mycobacterium vaccae and Mycobacterium vanbaalenii, within a clade related to fast-growing mycobacteria. Large-scale genomic rearrangements were identified by comparing the M. aurum genome to those of Mycobacterium tuberculosis and Mycobacterium leprae. M. aurum orthologous genes implicated in resistance to anti-tuberculosis drugs in M. tuberculosis were observed. The sequence identity at the DNA level varied from 68.6% for pncA (pyrazinamide drug-related) to 96.2% for rrs (streptomycin, capreomycin). We observed two homologous genes encoding the catalase-peroxidase enzyme (katG) that is associated with resistance to isoniazid. Similarly, two embB homologues were identified in the M. aurum genome. In addition to describing for the first time the genome of M. aurum, this work provides a resource to aid the use of M. aurum in studies to develop improved drugs for the pathogenic mycobacteria M. tuberculosis and M. leprae. Copyright © 2015 Asian-African Society for Mycobacteriology. Published by Elsevier Ltd. All rights reserved.

Risk factors for Mycobacterium tuberculosis infection among children in Greenland

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Søborg, Bolette; Andersen, Aase Bengaard; Melbye, Mads

2011-01-01

To examine the risk factors for Mycobacterium tuberculosis infection (MTI) among Greenlandic children for the purpose of identifying those at highest risk of infection.......To examine the risk factors for Mycobacterium tuberculosis infection (MTI) among Greenlandic children for the purpose of identifying those at highest risk of infection....

Characterization of extensively drug-resistant Mycobacterium tuberculosis in Nepal.

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Poudel, Ajay; Maharjan, Bhagwan; Nakajima, Chie; Fukushima, Yukari; Pandey, Basu D; Beneke, Antje; Suzuki, Yasuhiko

2013-01-01

The emergence of extensively drug-resistant tuberculosis (XDR-TB) has raised public health concern for global control of TB. Although molecular characterization of drug resistance-associated mutations in multidrug-resistant isolates in Nepal has been made, mutations in XDR isolates and their genotypes have not been reported previously. In this study, we identified and characterized 13 XDR Mycobacterium tuberculosis isolates from clinical isolates in Nepal. The most prevalent mutations involved in rifampicin, isoniazid, ofloxacin, and kanamycin/capreomycin resistance were Ser531Leu in rpoB gene (92.3%), Ser315Thr in katG gene (92.3%), Asp94Gly in gyrA gene (53.9%) and A1400G in rrs gene (61.5%), respectively. Spoligotyping and multilocus sequence typing revealed that 69% belonged to Beijing family, especially modern types. Further typing with 26-loci variable number of tandem repeats suggested the current spread of XDR M. tuberculosis. Our result highlights the need to reinforce the TB policy in Nepal with regard to control and detection strategies. Copyright © 2012 Elsevier Ltd. All rights reserved.

Peritoneal tuberculosis due to Mycobacterium caprae

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T. Nebreda

2016-01-01

Full Text Available The incidence of tuberculosis in humans due to Mycobacterium caprae is very low and is almost confined to Europe. We report a case of a previously healthy 41-year-old Moroccan with a 6 month history of abdominal pain, weight loss, fatigue and diarrhea. A diagnosis of peritoneal tuberculosis due to M. caprae was made.

[Characterization of clinical isolates of Mycobacterium tuberculosis from HIV positive individuals in Colombia, 2012].

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Castro, Claudia; Ricardo, Alba; Zabaleta, Angie; Llerena, Claudia; Puerto, Gloria

2017-01-24

One third of the increase in tuberculosis cases is attributed to the spread of HIV. In 2012, 1,397 HIV-associated tuberculosis cases were reported in Colombia, i.e., 11.8% of the total cases. Molecular epidemiology tools help to understand the transmission of tuberculosis. To characterize clinical isolates of Mycobacterium tuberculosis derived from HIV-infected individuals, received at the Laboratorio Nacional de Referencia in the Instituto Nacional de Salud. This was a descriptive observational study. We analyzed 63 isolates of M. tuberculosis from HIV-infected individuals. Identification, drug susceptibility and genotyping assays were performed. Of the new cases evaluated, three (5.0%) were resistant to isoniazid combined with streptomycin; two (3.3%) to rifampicin, and one (1.6%) to isoniazid. Previously treated cases were sensitive. No multidrug resistance was evident. Among the predominant genotypes, 20 isolates were (31.7%) LAM9, eight (12.7%), H1, and seven (11.1%), T1. Nineteen isolates corresponded to orphan patterns. One single grouping was observed among tested isolates. We found no statistically significantdifference between the proportions of the antituberculous drug resistance and genotypes. We found resistant isolates to the most powerful drugs, rifampicin and isoniazid, among new cases, showing the transmission of resistant strains. Genetic families of M. tuberculosis LAM9, T1 and H1 correspond to those described in the general population. We detected no active transmission among studied isolates. More comprehensive studies are needed to assess the real situation of HIV associated tuberculosis in the country regarding sensitivity and transmission.

Detection of Mycobacterium Tuberculosis by using PCR

International Nuclear Information System (INIS)

Suhadi, F; Dadang-Sudrajat; Maria-Lina, R.

1996-01-01

Polymerase Chain Reaction (PCR) procedure using three primary set derived from repetitive DNA sequence specific to mycobacteria was used to diagnose pathogenic Mycobacterium tuberculosis. The assay was specific for M. tuberculosis and could be used to detect the amount DNA less than 10 -9 g

Modelling the Transitional Dynamics of Mycobacterium Tuberculosis ...

African Journals Online (AJOL)

The World Health Organization's targets of eliminating Tuberculosis (TB) by 2050 is challenged by the emergence and spread of drug resistance TB. However, the traditional mechanism of resistance is that of acquired resistance, whereby the mycobacterium Tuberculosis (MTB) strain develops mutations under selective ...

Extra pulmonary tuberculosis: Rapid identification of Mycobacterium tuberculosis grown in Mycobacterium growth indicator tube 960 and Lowenstein-Jensen media, employing Standard diagnostics Bioline Mycobacterium tuberculosis protein 64 antigen detection kit

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G Kandhakumari

2015-01-01

Full Text Available Background: Investigation of extra pulmonary tuberculosis (EPTB in and around Pondicherry is being carried out since August 2011 in our tertiary care super specialty hospital. Objectives: To compare the rapid Kit SD Bio-Line MPT 64 Ag with conventional and time consuming biochemical tests. Confirmation of Mycobacterium tuberculosis at a reasonable time frame is the main thrust. Materials and Methods: Thirty three Mycobacterium tuberculosis and four Non-Tuberculous Mycobacteria (NTM grown in MGIT960 system/Lowenstein-Jensen media (LJ were examined by the rapid MPT 64 antigen detection as well as a battery of conventional tests like niacin, nitrate reduction, paraminobenzoic acid susceptibility and cord formation. Results and Conclusion: . Both the rapid kit and conventional tests correctly identified 33 M.tuberculosis isolates. Keeping conventional identification as reference, sensitivity and specificity for rapid kit was 100%. Rapid kit which takes only 15 minutes is accurate, cost effective, and facilitates early treatment for these EPTB patients, whose clinical specimens are paucibacillary.

Mycobacterium tuberculosis Infection in a Domesticated Korean Wild Boar ( Sus scrofa coreanus).

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Seo, Min-Goo; Ouh, In-Ohk; Kim, Munki; Lee, Jienny; Kim, Young-Hoan; Do, Jae-Cheul; Kwak, Dongmi

2017-06-01

Tuberculosis, a chronic progressive disease, has been reported in bovine, swine, and primate species. Here, we report the first case of Mycobacterium tuberculosis infection in a Korean wild boar ( Sus scrofa coreanus). The owners this domesticated boar brought it to the Gyeongbuk Veterinary Service Laboratory in Korea after it was found dead and severely emaciated. Demarcated yellowish white nodules were found around the larynx and retropharyngeal lymph node during necropsy. The lungs had diffuse fibrinous pleuritis, severe congestion, and scattered nodules. More nodules were found in the spleen. Tuberculosis is characterized by massive macrophage infiltration and central caseous necrosis; both characteristics were found in the lungs. Histopathologic examination revealed that the alveolar lumen had marked fibrosis and exudates. Examination of the fluid revealed extensive macrophage permeation. To confirm a Mycobacterium infection, PCR was performed using two primer sets specific to the rpoB gene of Mycobacterium; Mycobacterium was detected in the lungs and spleen. To identify the species of Mycobacterium, immunohistochemical evaluation was performed using antibodies against Mycobacterium tuberculosis and Mycobacterium bovis . The results revealed immunoreactivity against M. tuberculosis but not against M. bovis . The consumption of undercooked or raw meat from game animals may expose humans and other animals to sylvatic infection. Consequently, Koreans who ingest wild boar may be at risk of a tuberculosis infection. To reduce the risk of foodborne infection and maintain public health, continuous monitoring and control strategies are required.

Comparative Mycobacteriology of the Mycobacterium tuberculosis complex

OpenAIRE

Gordon, Stephen V.; Behr, Marcel A.

2015-01-01

The Mycobacterium tuberculosis complex (MTBC) is a group of highly genetically related pathogens that cause tuberculosis (TB) in mammalian species. However, the very name of the complex underlines the fact that our knowledge of these pathogens is dominated by studies on the human pathogen, M. tuberculosis. Of course this is entirely justified; M. tuberculosis is a major global pathogen that exacts a horrendous burden in terms of mortality and morbidity so it is appropriate that it is...

Coinfección por Mycobacterium malmoense y Mycobacterium tuberculosis en paciente con el síndrome de inmunodeficiencia humana

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Lilian María Mederos Cuervo

Full Text Available Se presenta un caso de coinfección por Mycobacterium malmoense y Mycobacterium tuberculosis en un paciente cubano con síndrome de inmunodeficiencia adquirida (sida, que producía enfermedad respiratoria y hepática respectivamente. Los cultivos realizados a partir de las muestras de esputo demostraron la presencia de una cepa micobacteriana no pigmentada de crecimiento lento perteneciente al grupo III de Runyon e identificada como Mycobacterium malmoense. A partir de los cultivos del tejido hepático extraído laparoscópicamente se aisló una cepa posteriormente identificada como Mycobacterium tuberculosis. El estudio anatomopatológico confirmó el diagnóstico de tuberculosis, el paciente recibió tratamiento específico y evolucionó clínicamente bien. Se reporta un caso infrecuente de coinfección por Mycobacterium, el cual describe el primer reporte de tuberculosis hepática en una paciente con sida en Cuba.

Diffuse Type Primary Mycobacterium Tuberculosis of the Breast: A Case Report

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Kim, Hyun A; Kang, Bong Joo; Kim, Sung Hun; Kim, Hnana; Lee, Ah Won [Seoul St. Mary' s Hospital, The Catholic University of Korea, Seoul (Korea, Republic of)

2011-12-15

Tuberculous mastitis is a rare manifestation of mycobacterium tuberculosis infection. It mimics inflammatory breast cancer or other pyogenic inflammations. In most of the tuberculous mastitis reports, coexisting or prior tuberculosis infection and secondary infection of the breast by direct spread via axillary or cervical lymphadenopathy, or hematogenous spread have been noted. We describe the mammographic and ultrasonographic findings of a case of diffuse type mycobacterium tuberculosis of the breast showing diffuse edema which was confirmed as tuberculosis through biopsy and had no evidence of old or concurrent pulmonary tuberculosis on chest computed tomography

Safety assessment in primary Mycobacterium tuberculosis smear ...

African Journals Online (AJOL)

Introduction Tuberculosis (TB) is caused by Mycobacterium tuberculosis and is transmitted mainly through aerosolization of infected sputum which puts laboratory workers at risk in spite of the laboratory workersf risk of infection being at 3 to 9 times higher than the general public. Laboratory safety should therefore be ...

Immunological crossreactivity of the Mycobacterium leprae CFP-10 with its homologue in Mycobacterium tuberculosis

NARCIS (Netherlands)

Geluk, A.; van Meijgaarden, K. E.; Franken, K. L. M. C.; Wieles, B.; Arend, S. M.; Faber, W. R.; Naafs, B.; Ottenhoff, T. H. M.

2004-01-01

Mycobacterium tuberculosis culture filtrate protein-10 (CFP-10) (Rv3874) is considered a promising antigen for the immunodiagnosis of tuberculosis (TB) together with early secreted antigens of M. tuberculosis (ESAT-6). Both ESAT-6 and CFP-10 are encoded by the RD1 region that is deleted from all

Novos dados sobre os Spoligotypes de estirpes do complexo Mycobacterium tuberculosis isoladas no Hospital Fernando Fonseca (Amadora-Sintra, Portugal

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Suzana David

2005-11-01

Full Text Available Resumo: O presente estudo populacional, que decorreu entre 1999 e 2003, foi baseado na utilização do Spoligotyping na genotipagem de 452 isolados do complexo Mycobacterium tuberculosis de doentes com tuberculose internados no Hospital Fernando Fonseca. Spoligotypes foram identificados como shared types (ST recorrendo a uma base de dados internacional. Onze ST raros, não identificados na base de dados, acomodaram 8,4% dos isolados. Aliás, particular a Portugal poderá ser a predominância de ST identificados na base de dados mas não previamente classificados como famílias genotípicas, tais como o ST244, ST150 e ST389, representando 13,3 % do total. A identificação de isolados clínicos de M. africanum de genótipo Afri1 e de M. tuberculosis de genótipo CAS1 poderá confirmar a importação de isolados de origem africana e asiática. M. tuberculosis da família Beijing foi pela primeira vez por nós assinalado a partir de 1999. Desde então, o número de isolados provenientes do hospital passou de um para cinco, anualmente, representando actualmente 2,2%, o que a coloca em décimo lugar em prevalência. M. tuberculosis Beijing poderá corresponder a um problema emergente em Portugal devido à recente imigração proveniente da Europa Oriental e da Ásia. Outros genótipos, ST150 e ST389, mostraram um incremento, cujo significado não é claro. No entanto, as frequências relativas das famílias predominantes LAM, T1 e Haarlem mantiveram-se relativamente estáveis. O presente estudo confirma a variabilidade genética em Portugal dos isolados do complexo M. tuberculosis. Estes estudos poderão contribuir para a definição de prioridades nos programas nacionais de luta contra a tuberculose.Rev Port Pneumol 2005; XI (6: 513-531 Abstract: The present population study, from 1999 to 2003, has been based on the use of Spoligotyping in the genotyping of 452 isolates of

First molecular epidemiology study of Mycobacterium tuberculosis in Kiribati.

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Eman Aleksic

Full Text Available Tuberculosis incidence rates in Kiribati are among the highest in the Western Pacific Region, however the genetic diversity of circulating Mycobacterium tuberculosis complex strains (MTBC and transmission dynamics are unknown. Here, we analysed MTBC strains isolated from culture positive pulmonary tuberculosis (TB cases from the main TB referral centre between November 2007 and October 2009. Strain genotyping (IS6110 typing, spoligotyping, 24-loci MIRU-VNTR and SNP typing was performed and demographic information collected. Among 73 MTBC strains analysed, we identified seven phylogenetic lineages, dominated by Beijing strains (49%. Beijing strains were further differentiated in two main branches, Beijing-A (n = 8 and -B (n = 28, that show distinct genotyping patterns and are characterized by specific deletion profiles (Beijing A: only RD105, RD207 deleted; Beijing B: RD150 and RD181 additionally deleted. Many Kiribati strains (59% based on IS6110 typing of all strains occurred in clusters, suggesting ongoing local transmission. Beijing-B strains and over-crowded living conditions were associated with strain clustering (likely recent transmission, however little evidence of anti-tuberculous drug resistance was observed. We suggest enhanced case finding amongst close contacts and continued supervised treatment of all identified cases using standard first-line drugs to reduce TB burden in Kiribati. Beijing strains can be subdivided in different principle branches that might be associated with differential spreading patterns in the population.

Adaptation and evolution of drug-resistant Mycobacterium tuberculosis

NARCIS (Netherlands)

Bergval, I.L.

2013-01-01

Many studies have been conducted on drug resistance and the evolution of Mycobacterium tuberculosis. Notwithstanding, many molecular mechanisms facilitating the emergence, adaptation and spread of drug-resistant tuberculosis have yet to be discovered. This thesis reports studies of the adaptive

Conspicuous multidrug-resistant Mycobacterium tuberculosis cluster strains do not trespass country borders in Latin America and Spain.

Science.gov (United States)

Ritacco, Viviana; Iglesias, María-José; Ferrazoli, Lucilaine; Monteserin, Johana; Dalla Costa, Elis R; Cebollada, Alberto; Morcillo, Nora; Robledo, Jaime; de Waard, Jacobus H; Araya, Pamela; Aristimuño, Liselotte; Díaz, Raúl; Gavin, Patricia; Imperiale, Belen; Simonsen, Vera; Zapata, Elsa M; Jiménez, María S; Rossetti, Maria L; Martin, Carlos; Barrera, Lucía; Samper, Sofia

2012-06-01

Multidrug-resistant Mycobacterium tuberculosis strain diversity in Ibero-America was examined by comparing extant genotype collections in national or state tuberculosis networks. To this end, genotypes from over 1000 patients with multidrug-resistant tuberculosis diagnosed from 2004 through 2008 in Argentina, Brazil, Chile, Colombia, Venezuela and Spain were compared in a database constructed ad hoc. Most of the 116 clusters identified by IS6110 restriction fragment length polymorphism were small and restricted to individual countries. The three largest clusters, of 116, 49 and 25 patients, were found in Argentina and corresponded to previously documented locally-epidemic strains. Only 13 small clusters involved more than one country, altogether accounting for 41 patients, of whom 13 were, in turn, immigrants from Latin American countries different from those participating in the study (Peru, Ecuador and Bolivia). Most of these international clusters belonged either to the emerging RD(Rio) LAM lineage or to the Haarlem family of M. tuberculosis and four were further split by country when analyzed with spoligotyping and rifampin resistance-conferring mutations, suggesting that they did not represent ongoing transnational transmission events. The Beijing genotype accounted for 1.3% and 10.2% of patients with multidrug-resistant tuberculosis in Latin America and Spain, respectively, including one international cluster of two cases. In brief, Euro-American genotypes were widely predominant among multidrug-resistant M. tuberculosis strains in Ibero-America, reflecting closely their predominance in the general M. tuberculosis population in the region, and no evidence was found of acknowledged outbreak strains trespassing country borders. Copyright © 2011 Elsevier B.V. All rights reserved.

Molecular characterisation of Mycobacterium caprae strains isolated in Poland.

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Krajewska-Wędzina, Monika; Kozińska, Monika; Orłowska, Blanka; Weiner, Marcin; Szulowski, Krzysztof; Augustynowicz-Kopeć, Ewa; Anusz, Krzysztof; Smith, Noel H

2018-03-10

Bovine tuberculosis (bovine TB, bTB) is caused by bovine bacilli: Mycobacterium bovis and M caprae The studies conducted in Poland, in the National Bovine Tuberculosis Reference Laboratory in the Department of Microbiology of the National Veterinary Research Institute in Pulawy, show that animal tuberculosis in Poland is also caused by M caprae We here describe the identification and genotypic assessment of 52 isolates of M caprae obtained from Polish cattle and wild animals over the last five years. We show that strains isolated from bison have significant genotypic diversity and are distinct compared with the genotypes of strains isolated from cattle. Similarly, isolates from cattle herds can be highly genotypically variable. Formal designation of the members of the Mycobacterium tuberculosis complex is controversial in Poland; there is a gap in veterinary legislation with regard to bTB and no explicit mention of M caprae causing tuberculosis in animal. © British Veterinary Association (unless otherwise stated in the text of the article) 2018. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

Whole genome sequencing versus traditional genotyping for investigation of a Mycobacterium tuberculosis outbreak: a longitudinal molecular epidemiological study.

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Andreas Roetzer

Full Text Available BACKGROUND: Understanding Mycobacterium tuberculosis (Mtb transmission is essential to guide efficient tuberculosis control strategies. Traditional strain typing lacks sufficient discriminatory power to resolve large outbreaks. Here, we tested the potential of using next generation genome sequencing for identification of outbreak-related transmission chains. METHODS AND FINDINGS: During long-term (1997 to 2010 prospective population-based molecular epidemiological surveillance comprising a total of 2,301 patients, we identified a large outbreak caused by an Mtb strain of the Haarlem lineage. The main performance outcome measure of whole genome sequencing (WGS analyses was the degree of correlation of the WGS analyses with contact tracing data and the spatio-temporal distribution of the outbreak cases. WGS analyses of the 86 isolates revealed 85 single nucleotide polymorphisms (SNPs, subdividing the outbreak into seven genome clusters (two to 24 isolates each, plus 36 unique SNP profiles. WGS results showed that the first outbreak isolates detected in 1997 were falsely clustered by classical genotyping. In 1998, one clone (termed "Hamburg clone" started expanding, apparently independently from differences in the social environment of early cases. Genome-based clustering patterns were in better accordance with contact tracing data and the geographical distribution of the cases than clustering patterns based on classical genotyping. A maximum of three SNPs were identified in eight confirmed human-to-human transmission chains, involving 31 patients. We estimated the Mtb genome evolutionary rate at 0.4 mutations per genome per year. This rate suggests that Mtb grows in its natural host with a doubling time of approximately 22 h (400 generations per year. Based on the genome variation discovered, emergence of the Hamburg clone was dated back to a period between 1993 and 1997, hence shortly before the discovery of the outbreak through epidemiological

Revisiting host preference in the Mycobacterium tuberculosis complex: experimental infection shows M. tuberculosis H37Rv to be avirulent in cattle.

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Adam O Whelan

Full Text Available Experiments in the late 19th century sought to define the host specificity of the causative agents of tuberculosis in mammals. Mycobacterium tuberculosis, the human tubercle bacillus, was independently shown by Smith, Koch, and von Behring to be avirulent in cattle. This finding was erroneously used by Koch to argue the converse, namely that Mycobacterium bovis, the agent of bovine tuberculosis, was avirulent for man, a view that was subsequently discredited. However, reports in the literature of M. tuberculosis isolation from cattle with tuberculoid lesions suggests that the virulence of M. tuberculosis for cattle needs to be readdressed. We used an experimental bovine infection model to test the virulence of well-characterized strains of M. tuberculosis and M. bovis in cattle, choosing the genome-sequenced strains M. tuberculosis H37Rv and M. bovis 2122/97. Cattle were infected with approximately 10(6 CFU of M. tuberculosis H37Rv or M. bovis 2122/97, and sacrificed 17 weeks post-infection. IFN-gamma and tuberculin skin tests indicated that both M. bovis 2122 and M. tuberculosis H37Rv were equally infective and triggered strong cell-mediated immune responses, albeit with some indication of differential antigen-specific responses. Postmortem examination revealed that while M. bovis 2122/97-infected animals all showed clear pathology indicative of bovine tuberculosis, the M. tuberculosis-infected animals showed no pathology. Culturing of infected tissues revealed that M. tuberculosis was able to persist in the majority of animals, albeit at relatively low bacillary loads. In revisiting the early work on host preference across the M. tuberculosis complex, we have shown M. tuberculosis H37Rv is avirulent for cattle, and propose that the immune status of the animal, or genotype of the infecting bacillus, may have significant bearing on the virulence of a strain for cattle. This work will serve as a baseline for future studies into the genetic basis

Application of Pulsed Field Gel Electrophoresis for Study of Genetic Diversity in Mycobacterium tuberculosis Strains Isolated From Tuberculosis Patients.

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Khosravi, Azar Dokht; Vatani, Shideh; Feizabadi, Mohammad Mehdi; Abasi Montazeri, Effat; Jolodar, Abbas

2014-05-01

Mycobacterium tuberculosis genotyping can effectively improve tuberculosis (TB) control programs by controlling disease transmission. Pulsed field gel electrophoresis (PFGE) is a particularly powerful tool for determination of clonal identity of bacteria providing information for understanding and controlling the spread of disease. The aim of present study was to investigate the genetic diversity of M. tuberculosis strains in Khuzestan province by the PFGE technique. In total, 80 M. tuberculosis positive cultures were obtained from tuberculosis patients. PFGE was performed on 60 PCR-confirmed isolates by using DraI and XbaI restriction enzymes according to standard protocols. Plugs containing digested DNA were then loaded on agarose gels and run using contour-clamped homogenous electric fields. Fifty distinct DNA banding patterns were obtained by digestion of DNA with DraI and 38 DNA banding patterns by digestion with XbaI restriction enzymes. The patterns comprised of 17 different clusters in which cluster I was the major one, containing six strains. Three clusters contained three strains each and the 13 remaining clusters comprised of two strains each. Digestion with DraI yielded 15-20 DNA fragments with 50-485 kb size, while digestion by XbaI produced DNA fragments with a size smaller than 50-242 kb. Despite the ability of PFGE for study of genetic diversity of many mycobacterial species and it being considered as a robust and useful tool, in this study we only found a 15% epidemiological relationship amongst the isolates. Thus, for higher discrimination of genotypic clusters among M. tuberculosis clinical isolates, the application of more sophisticated complementary techniques is required.

Collectin CL-LK Is a Novel Soluble Pattern Recognition Receptor for Mycobacterium tuberculosis

DEFF Research Database (Denmark)

Troegeler, Anthony; Lugo-Villarino, Geanncarlo; Hansen, Søren

2015-01-01

Understanding the molecular components of immune recognition of the tuberculosis (TB) bacillus, Mycobacterium tuberculosis, can help designing novel strategies to combat TB. Here, we identify collectin CL-LK as a novel soluble C-type lectin able to bind M. tuberculosis, and characterize mycobacte......Understanding the molecular components of immune recognition of the tuberculosis (TB) bacillus, Mycobacterium tuberculosis, can help designing novel strategies to combat TB. Here, we identify collectin CL-LK as a novel soluble C-type lectin able to bind M. tuberculosis, and characterize...

Benzothiazinones kill Mycobacterium tuberculosis by blocking arabinan synthesis

DEFF Research Database (Denmark)

Makarov, Vadim; Manina, Giulia; Mikusova, Katarina

2009-01-01

New drugs are required to counter the tuberculosis (TB) pandemic. Here, we describe the synthesis and characterization of 1,3-benzothiazin-4-ones (BTZs), a new class of antimycobacterial agents that kill Mycobacterium tuberculosis in vitro, ex vivo, and in mouse models of TB. Using genetics...

Characterization of a Mycobacterium leprae antigen related to the secreted Mycobacterium tuberculosis protein MPT32

NARCIS (Netherlands)

Wieles, B.; van Agterveld, M.; Janson, A.; Clark-Curtiss, J.; Rinke de Wit, T.; Harboe, M.; Thole, J.

1994-01-01

Secreted proteins may serve as major targets in the immune response to mycobacteria. To identify potentially secreted Mycobacterium leprae antigens, antisera specific for culture filtrate proteins of Mycobacterium tuberculosis were used to screen a panel of recombinant antigens selected previously

Triple valve endocarditis by mycobacterium tuberculosis. A case report

Directory of Open Access Journals (Sweden)

Shaikh Quratulain

2012-09-01

Full Text Available Abstract Background Granulomas caused by Mycobacterium Tuberculosis have been observed at autopsy in the heart, pre-dominantly in the myocardium and endocardium, but rarely involving the coronary vessels and valvular structures. Mycobacterium tuberculosis valvular endocarditis is extremely rare, with most reports coming from autopsy series. Case presentation We report the case of a 17 year old immunocompetent girl who presented with history of fever, malaise, foot gangrene and a left sided hemiparesis. On investigation she was found to have infective endocarditis involving the aortic, mitral and tricuspid valves. She had developed a right middle cerebral artery stroke. She underwent dual valve replacement and tricuspid repair. The vegetations showed granulomatous inflammation but blood cultures and other biological specimen cultures were negative for any organisms. She was started on antituberculous treatment and anticoagulation. Conclusion This is the first reported case of triple valve endocarditis by Mycobacterium Tuberculosis in an immunocompetent host. Especially important is the fact that the right heart is involved which has been historically described in the setting of intravenous drug abuse. This implies that Tuberculosis should be considered in cases of culture negative endocarditis in endemic areas like Pakistan even in immunocompetent hosts.

Advances in the Laboratory Diagnosis of Mycobacterium Tuberculosis

African Journals Online (AJOL)

Mycobacterium tuberculosis (MTB), the agent of human tuberculosis remains a leading cause of mortality globally. Its resurgence during the last two decades is a reflection of its opportunistic relationship with HIV. The challenges associated with the disease are enormous and often debilitating. The role of clinical and ...

Comparative Genomics and Proteomic Analysis of Four Non-tuberculous Mycobacterium Species and Mycobacterium tuberculosis Complex : Occurrence of Shared Immunogenic Proteins

NARCIS (Netherlands)

Gcebe, Nomakorinte; Michel, Anita; Gey van Pittius, Nicolaas C; Rutten, Victor

2016-01-01

The Esx and PE/PPE families of proteins are among the most immunodominant mycobacterial antigens and have thus been the focus of research to develop vaccines and immunological tests for diagnosis of bovine and human tuberculosis, mainly caused by Mycobacterium bovis and Mycobacterium tuberculosis,

Buoyant density of Mycobacterium tuberculosis: implications for sputum processing

NARCIS (Netherlands)

den Hertog, A. L.; Klatser, P. R.; Anthony, R. M.

2009-01-01

A tuberculosis (TB) research laboratory in the Netherlands. The concentration of Mycobacterium tuberculosis cells from sputum is almost universally performed by centrifugation after chemical liquefaction. These methods are thus dependent on the effective sedimentation of mycobacterial cells, and the

Intraocular manifestations of mycobacterium tuberculosis: A review of the literature

Directory of Open Access Journals (Sweden)

Lauren A. Dalvin

2017-05-01

Full Text Available Mycobacterium tuberculosis: is most commonly associated with pulmonary infection. However, tuberculosis (TB can also affect the eye. TB can affect nearly any tissue in the eye, and a high index of suspicion is required for accurate diagnosis, as many of the intraocular manifestations of TB can mimic other, more common diseases. Correct diagnosis is critical because systemic anti-tuberculosis treatment may be required, and vision loss or even loss of the affected eye can occur without proper treatment. Thus, it is important for ophthalmologists and infectious disease specialists to work together to accurately diagnose and treat intraocular TB. This article reports the various known presentations of intraocular TB and reviews important elements of diagnosis and treatment. Keywords: Mycobacterium, Tuberculosis, Choroidal granuloma, Retinal vasculitis

Diversity and evolution of drug resistance mechanisms in Mycobacterium tuberculosis

Directory of Open Access Journals (Sweden)

Al-Saeedi M

2017-10-01

Full Text Available Mashael Al-Saeedi, Sahal Al-Hajoj Department of Infection and Immunity, Mycobacteriology Research Section, King Faisal Specialist Hospital and Research Center, Riyadh, Saudi Arabia Abstract: Despite the efficacy of antibiotics to protect humankind against many deadly pathogens, such as Mycobacterium tuberculosis, nothing can prevent the emergence of drug-resistant strains. Several mechanisms facilitate drug resistance in M. tuberculosis including compensatory evolution, epistasis, clonal interference, cell wall integrity, efflux pumps, and target mimicry. In this study, we present recent findings relevant to these mechanisms, which can enable the discovery of new drug targets and subsequent development of novel drugs for treatment of drug-resistant M. tuberculosis. Keywords: Mycobacterium tuberculosis, antibiotic resistance, compensatory evolution, epistasis, efflux pumps, fitness cost

Unraveling Mycobacterium tuberculosis genomic diversity and evolution in Lisbon, Portugal, a highly drug resistant setting

KAUST Repository

Perdigão, João

2014-11-18

Background Multidrug- (MDR) and extensively drug resistant (XDR) tuberculosis (TB) presents a challenge to disease control and elimination goals. In Lisbon, Portugal, specific and successful XDR-TB strains have been found in circulation for almost two decades. Results In the present study we have genotyped and sequenced the genomes of 56 Mycobacterium tuberculosis isolates recovered mostly from Lisbon. The genotyping data revealed three major clusters associated with MDR-TB, two of which are associated with XDR-TB. Whilst the genomic data contributed to elucidate the phylogenetic positioning of circulating MDR-TB strains, showing a high predominance of a single SNP cluster group 5. Furthermore, a genome-wide phylogeny analysis from these strains, together with 19 publicly available genomes of Mycobacterium tuberculosis clinical isolates, revealed two major clades responsible for M/XDR-TB in the region: Lisboa3 and Q1 (LAM). The data presented by this study yielded insights on microevolution and identification of novel compensatory mutations associated with rifampicin resistance in rpoB and rpoC. The screening for other structural variations revealed putative clade-defining variants. One deletion in PPE41, found among Lisboa3 isolates, is proposed to contribute to immune evasion and as a selective advantage. Insertion sequence (IS) mapping has also demonstrated the role of IS6110 as a major driver in mycobacterial evolution by affecting gene integrity and regulation. Conclusions Globally, this study contributes with novel genome-wide phylogenetic data and has led to the identification of new genomic variants that support the notion of a growing genomic diversity facing both setting and host adaptation.

Molecular detection methods of resistance to antituberculosis drugs in Mycobacterium tuberculosis.

Science.gov (United States)

Brossier, F; Sougakoff, W

2017-09-01

Molecular methods predict drug resistance several weeks before phenotypic methods and enable rapid implementation of appropriate therapeutic treatment. We aimed to detail the most representative molecular tools used in routine practice for the rapid detection of resistance to antituberculosis drugs among Mycobacterium tuberculosis strains. The molecular diagnosis of resistance to antituberculosis drugs in clinical samples or from in vitro cultures is based on the detection of the most common mutations in the genes involved in the development of resistance in M. tuberculosis strains (encoding either protein targets of antibiotics, or antibiotic activating enzymes) by commercial molecular kits or by sequencing. Three hypotheses could explain the discrepancies between the genotypic results and the phenotypic drug susceptibility testing results: a low percentage of resistant mutants precluding the detection by genotypic methods on the primary culture; a low level of resistance not detected by phenotypic testing; and other resistance mechanisms not yet characterized. Molecular methods have varying sensitivity with regards to detecting antituberculosis drug resistance; that is why phenotypic susceptibility testing methods are mandatory for detecting antituberculosis drug-resistant isolates that have not been detected by molecular methods. The questionable ability of existing phenotypic and genotypic drug susceptibility testing to properly classify strains as susceptible or resistant, and at what level of resistance, was raised for several antituberculosis agents. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

21 CFR 866.3370 - Mycobacterium tuberculosis immunofluorescent reagents.

Science.gov (United States)

2010-04-01

... HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents... used to identify Mycobacterium tuberculosis directly from clinical specimens. The identification aids...

Comparison of antigen-specific T-cell responses of tuberculosis patients using complex or single antigens of Mycobacterium tuberculosis

DEFF Research Database (Denmark)

Mustafa, A S; Amoudy, H A; Wiker, H G

1998-01-01

GroES, rPstS, rGroEL and rDnaK) antigens of Mycobacterium tuberculosis. The responses of PBMC to these defined antigens were compared with the corresponding results obtained with complex antigens, such as whole-cell M. tuberculosis, M. tuberculosis culture filtrate (MT-CF) and cell wall antigens, as well...... as the vaccine strain, Mycobacterium bovis bacillus Calmette-Guerin (BCG). In addition, M. tuberculosis and MT-CF-induced T-cell lines were tested in the same assays against the panel of purified and complex antigens. The compiled data from PBMC and T-cell lines tested for antigen-induced proliferation and IFN...

Tuberculosis transmission of predominant genotypes of Mycobacterium tuberculosis in Northern suburbs of Buenos Aires city region Transmisión de la tuberculosis por genotipos predominantes de Mycobacterium tuberculosis en la región Gran Buenos Aires Norte

Directory of Open Access Journals (Sweden)

N. Morcillo

2007-09-01

Full Text Available In 2003, the incidence of tuberculosis in Argentina showed an increase compared to 2002. The severe national crisis at the end of the 90s has probably strongly contributed to this situation. The goal of this work was to estimate the extent of the spread of the most predominant Mycobacterium tuberculosis strains and to assess the spread of predominant M. tuberculosis clusters as determined by spoligotyping and IS6110 RFLP. The study involved 590 pulmonary, smear-positive TB cases receiving medical attention at health centers and hospitals in Northern Buenos Aires (NBA suburbs, from October 2001 to December 2002. From a total of 208 clinical isolates belonging to 6 major clusters, 63 (30.2% isolates had identical spoligotyping and IS6110 RFLP pattern. Only 22.2% were shown to have epidemiological connections with another member of their respective cluster. In these major clusters, 30.2% of the 208 TB cases studied by both molecular techniques and contact tracing could be convincingly attributable to a recently acquired infection. This knowledge may be useful to assess the clonal distribution of predominant M. tuberculosis clusters in Argentina, which may make an impact on TB control strategies.La incidencia de la tuberculosis en Argentina mostró en 2003 un incremento en comparación con 2002. La grave crisis nacional a fines de los 90 ha probablemente contribuido en gran medida a esta situación. El objetivo del presente trabajo fue determinar la diversidad genética de aislamientos de Mycobacterium tuberculosis y el grado de dispersión de algunas cepas mayoritarias genéticamente relacionadas. El estudio involucró 590 aislamientos clínicos provenientes de muestras respiratorias con examen directo positivo, de pacientes atendidos en los hospitales y centros de salud que conforman la región Gran Buenos Aires Norte (NBA, de octubre de 2001 a diciembre de 2002. De 208 aislamientos que se encontraron en los 6 mayores clusters, 63 (30,2% ten

Integration of published information into a resistance-associated mutation database for Mycobacterium tuberculosis.

Science.gov (United States)

Salamon, Hugh; Yamaguchi, Ken D; Cirillo, Daniela M; Miotto, Paolo; Schito, Marco; Posey, James; Starks, Angela M; Niemann, Stefan; Alland, David; Hanna, Debra; Aviles, Enrique; Perkins, Mark D; Dolinger, David L

2015-04-01

Tuberculosis remains a major global public health challenge. Although incidence is decreasing, the proportion of drug-resistant cases is increasing. Technical and operational complexities prevent Mycobacterium tuberculosis drug susceptibility phenotyping in the vast majority of new and retreatment cases. The advent of molecular technologies provides an opportunity to obtain results rapidly as compared to phenotypic culture. However, correlations between genetic mutations and resistance to multiple drugs have not been systematically evaluated. Molecular testing of M. tuberculosis sampled from a typical patient continues to provide a partial picture of drug resistance. A database of phenotypic and genotypic testing results, especially where prospectively collected, could document statistically significant associations and may reveal new, predictive molecular patterns. We examine the feasibility of integrating existing molecular and phenotypic drug susceptibility data to identify associations observed across multiple studies and demonstrate potential for well-integrated M. tuberculosis mutation data to reveal actionable findings. © The Author 2014. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Immune responses to the Mycobacterium tuberculosis-specific antigen ESAT-6 signal subclinical infection among contacts of tuberculosis patients

DEFF Research Database (Denmark)

Doherty, T Mark; Demissie, Abebech; Olobo, Joseph

2002-01-01

Diagnosis of latent Mycobacterium tuberculosis infection is considered essential for tuberculosis control but is hampered by the lack of specific reagents. We report that strong recognition of tuberculosis complex-specific antigen ESAT-6 by healthy household contacts of tuberculosis patients...

Antibacterial Activity of Medicinal Aqueous Plant Extracts against Mycobacterium tuberculosis

Directory of Open Access Journals (Sweden)

Muna Mohammed Buzayan

2012-09-01

Full Text Available Tuberculosis (TB remains a serious health problem in many regions of the world, and the development of resistance to antibiotics by this microbe created the need for new drugs to replace those which have lost effectiveness. This study assesses the medicinal anti-Mycobacterium tuberculosis properties of natural products obtained from plants collected from Eastern Libya. In this study aqueous extracts of nine different plants were assayed for their Mycobacterium tuberculosis inhibitory activity using the BACTEC MGIT960 susceptibility test method. The aqueous extracts of Ceratonia siliqua L, Helichrysum stoechas (L. Moench and Thymus algeriensis did not show any activity against M. tuberculosis in different concentrations. The aqueous extract of Marrubium vulgare L. from Syria showed high activity against M. tuberculosis. Marrubium alysson L., Marrubium vulgare L., Pistacia lentiscus L, Quercus coccifera L, Thymus capitatus (L. Hoffm. & Link, showed varying degrees of activity against M. tuberculosis. The results of this study show that aqueous extracts from six different medicinal plants have different effects against M. tuberculosis in vitro.

Evaluation of customised lineage-specific sets of MIRU-VNTR loci for genotyping Mycobacterium tuberculosis complex isolates in Ghana.

Science.gov (United States)

Asante-Poku, Adwoa; Nyaho, Michael Selasi; Borrell, Sonia; Comas, Iñaki; Gagneux, Sebastien; Yeboah-Manu, Dorothy

2014-01-01

Different combinations of variable number of tandem repeat (VNTR) loci have been proposed for genotyping Mycobacterium tuberculosis complex (MTBC). Existing VNTR schemes show different discriminatory capacity among the six human MTBC lineages. Here, we evaluated the discriminatory power of a "customized MIRU12" loci format proposed previously by Comas et al. based on the standard 24 loci defined by Supply et al. for VNTR-typing of MTBC in Ghana. One hundred and fifty-eight MTBC isolates classified into Lineage 4 and Lineage 5 were used to compare a customized lineage-specific panel of 12 MIRU-VNTR loci ("customized MIRU-12") to the standard MIRU-15 genotyping scheme. The resolution power of each typing method was determined based on the Hunter-Gaston- Discriminatory Index (HGDI). A minimal set of customized MIRU-VNTR loci for typing Lineages 4 (Euro-American) and 5 (M. africanum West African 1) strains from Ghana was defined based on the cumulative HGDI. Among the 106 Lineage 4 strains, the customized MIRU-12 identified a total of 104 distinct genotypes consisting of 2 clusters of 2 isolates each (clustering rate 1.8%), and 102 unique strains while standard MIRU-15 yielded a total of 105 different genotypes, including 1 cluster of 2 isolates (clustering rate: 0.9%) and 104 singletons. Among, 52 Lineage 5 isolates, customized MIRU-12 genotyping defined 51 patterns with 1 cluster of 2 isolates (clustering rate: 0.9%) and 50 unique strains whereas MIRU-15 classified all 52 strains as unique. Cumulative HGDI values for customized MIRU-12 for Lineages 4 and 5 were 0.98 respectively whilst that of standard MIRU-15 was 0.99. A union of loci from the customised MIRU-12 and standard MIRU-15 revealed a set of customized eight highly discriminatory loci: 4052, 2163B, 40, 4165, 2165, 10,16 and 26 with a cumulative HGDI of 0.99 for genotyping Lineage 4 and 5 strains from Ghana.

Evaluation of customised lineage-specific sets of MIRU-VNTR loci for genotyping Mycobacterium tuberculosis complex isolates in Ghana.

Directory of Open Access Journals (Sweden)

Adwoa Asante-Poku

Full Text Available BACKGROUND: Different combinations of variable number of tandem repeat (VNTR loci have been proposed for genotyping Mycobacterium tuberculosis complex (MTBC. Existing VNTR schemes show different discriminatory capacity among the six human MTBC lineages. Here, we evaluated the discriminatory power of a "customized MIRU12" loci format proposed previously by Comas et al. based on the standard 24 loci defined by Supply et al. for VNTR-typing of MTBC in Ghana. METHOD: One hundred and fifty-eight MTBC isolates classified into Lineage 4 and Lineage 5 were used to compare a customized lineage-specific panel of 12 MIRU-VNTR loci ("customized MIRU-12" to the standard MIRU-15 genotyping scheme. The resolution power of each typing method was determined based on the Hunter-Gaston- Discriminatory Index (HGDI. A minimal set of customized MIRU-VNTR loci for typing Lineages 4 (Euro-American and 5 (M. africanum West African 1 strains from Ghana was defined based on the cumulative HGDI. RESULTS AND CONCLUSION: Among the 106 Lineage 4 strains, the customized MIRU-12 identified a total of 104 distinct genotypes consisting of 2 clusters of 2 isolates each (clustering rate 1.8%, and 102 unique strains while standard MIRU-15 yielded a total of 105 different genotypes, including 1 cluster of 2 isolates (clustering rate: 0.9% and 104 singletons. Among, 52 Lineage 5 isolates, customized MIRU-12 genotyping defined 51 patterns with 1 cluster of 2 isolates (clustering rate: 0.9% and 50 unique strains whereas MIRU-15 classified all 52 strains as unique. Cumulative HGDI values for customized MIRU-12 for Lineages 4 and 5 were 0.98 respectively whilst that of standard MIRU-15 was 0.99. A union of loci from the customised MIRU-12 and standard MIRU-15 revealed a set of customized eight highly discriminatory loci: 4052, 2163B, 40, 4165, 2165, 10,16 and 26 with a cumulative HGDI of 0.99 for genotyping Lineage 4 and 5 strains from Ghana.

Putative in vitro expressed gene fragments unique to Mycobacterium avium subspecies para tuberculosis

DEFF Research Database (Denmark)

Nielsen, Kirstine Klitgaard; Ahrens, Peter

2002-01-01

By a suppression subtractive hybridization based method, nine novel Mycobacterium avium subsp. para tuberculosis (M. para tuberculosis) fragments of between 318 and 596 bp have been identified and characterized. Database search revealed little or no similarity with other mycobacteria. The uniquen......By a suppression subtractive hybridization based method, nine novel Mycobacterium avium subsp. para tuberculosis (M. para tuberculosis) fragments of between 318 and 596 bp have been identified and characterized. Database search revealed little or no similarity with other mycobacteria....... The uniqueness and diagnostic potential of seven of these fragments in relation to M. paratuberculosis closest relative Mycobacterium avium subsp. avium (M. avium) was confirmed by species-specific PCR and Southern blot. Furthermore, RT-PCR indicated that eight of the nine fragments originate from areas...

Phylogenetic analysis of vitamin B12-related metabolism in Mycobacterium tuberculosis

OpenAIRE

Young, Douglas B.; Comas, I?aki; de Carvalho, Luiz P. S.

2015-01-01

Comparison of genome sequences from clinical isolates of Mycobacterium tuberculosis with phylogenetically-related pathogens Mycobacterium marinum, Mycobacterium kansasii, and Mycobacterium leprae reveals diversity amongst genes associated with vitamin B12-related metabolism. Diversity is generated by gene deletion events, differential acquisition of genes by horizontal transfer, and single nucleotide polymorphisms (SNPs) with predicted impact on protein function and transcriptional regulation...

Siderocalin inhibits the intracellular replication of Mycobacterium tuberculosis in macrophages

DEFF Research Database (Denmark)

Johnson, Erin E; Srikanth, Chittur V; Sandgren, Andreas

2010-01-01

Siderocalin is a secreted protein that binds to siderophores to prevent bacterial iron acquisition. While it has been shown to inhibit the growth of Mycobacterium tuberculosis (M.tb) in extracellular cultures, its effect on this pathogen within macrophages is not clear. Here, we show that sideroc......Siderocalin is a secreted protein that binds to siderophores to prevent bacterial iron acquisition. While it has been shown to inhibit the growth of Mycobacterium tuberculosis (M.tb) in extracellular cultures, its effect on this pathogen within macrophages is not clear. Here, we show...... findings are consistent with an important role for siderocalin in protection against M.tb infection and suggest that exogenously administered siderocalin may have therapeutic applications in tuberculosis....

The Genotypic Population Structure of Mycobacterium tuberculosis Complex from Moroccan Patients Reveals a Predominance of Euro-American Lineages

Science.gov (United States)

Lahlou, Ouafae; Millet, Julie; Chaoui, Imane; Sabouni, Radia; Filali-Maltouf, Abdelkarim; Akrim, Mohammed; El Mzibri, Mohammed; Rastogi, Nalin; El Aouad, Rajae

2012-01-01

Background Tuberculosis (TB) remains a major health problem in Morocco. Characterization of circulating Mycobacterium tuberculosis genotypic lineages, important to understand the dynamic of the disease, was hereby addressed for the first time at a national level. Methodology/Principal Findings Spoligotyping was performed on a panel of 592 M. tuberculosis complex strains covering a 2-year period (2004–2006). It identified 129 patterns: 105 (n = 568 strains) corresponded to a SIT number in the SITVIT2 database, while 24 patterns were labeled as orphan. A total of 523 (88.3%) strains were clustered vs. 69 or 11.7% unclustered. Classification of strains within 3 large phylogenetical groups was as follows: group 1– ancestral/TbD1+/PGG1 (EAI, Bovis, Africanum), group 2– modern/TbD1−/PGG1 group (Beijing, CAS), group 3– evolutionary recent/TbD1−/PGG2/3 (Haarlem, X, S, T, LAM; alternatively designated as the Euro-American lineage). As opposed to group 3 strains (namely LAM, Haarlem, and T) that predominated (86.5% of all isolates), 6 strains belonged to group 2 (Beijing n = 5, CAS n = 1), and 3 strains (BOV_1 n = 2, BOV_4-CAPRAE) belonged to ancestral group 1 (EAI and AFRI lineage strains were absent). 12-loci MIRU-VNTR typing of the Casablanca subgroup (n = 114 strains) identified 71 patterns: 48 MITs and 23 orphan patterns; it allowed to reduce the clustering rate from 72.8% to 29.8% and the recent transmission rate from 64% to 20.2%. Conclusion The M. tuberculosis population structure in Morocco is highly homogeneous, and is characterized by the predominance of the Euro-American lineages, namely LAM, Haarlem, and T, which belong to the “evolutionary recent” TbD1−/PGG2/3 phylogenetic group. The combination of spoligotyping and MIRUs decreased the clustering rate significantly, and should now be systematically applied in larger studies. The methods used in this study appear well suited to monitor the M. tuberculosis population

The Use Of Rap-PCR In Studying Mycobacterium tuberculosis ...

African Journals Online (AJOL)

Mycobacterium tuberculosis is the second leading cause of death from infectious agent. This study sought to detect M. tuberculosis genes, which were specifically expressed, or upregulated during intracellular infection of. J774 murine macrophages; as such genes may be potential targets for novel drug action. J774 murine ...

Genetic Mimetics of Mycobacterium tuberculosis and Methicillin-Resistant Staphylococcus aureus as Verification Standards for Molecular Diagnostics.

Science.gov (United States)

Machowski, Edith Erika; Kana, Bavesh Davandra

2017-12-01

Molecular diagnostics have revolutionized the management of health care through enhanced detection of disease or infection and effective enrollment into treatment. In recognition of this, the World Health Organization approved the rollout of nucleic acid amplification technologies for identification of Mycobacterium tuberculosis using platforms such as GeneXpert MTB/RIF, the GenoType MTBDR plus line probe assay, and, more recently, GeneXpert MTB/RIF Ultra. These assays can simultaneously detect tuberculosis infection and assess rifampin resistance. However, their widespread use in health systems requires verification and quality assurance programs. To enable development of these, we report the construction of genetically modified strains of Mycobacterium smegmatis that mimic the profile of Mycobacterium tuberculosis on both the GeneXpert MTB/RIF and the MTBDR plus line probe diagnostic tests. Using site-specific gene editing, we also created derivatives that faithfully mimic the diagnostic result of rifampin-resistant M. tuberculosis , with mutations at positions 513, 516, 526, 531, and 533 in the rifampin resistance-determining region of the rpoB gene. Next, we extended this approach to other diseases and demonstrated that a Staphylococcus aureus gene sequence can be introduced into M. smegmatis to generate a positive response for the SCC mec probe in the GeneXpert SA Nasal Complete molecular diagnostic cartridge, designed for identification of methicillin-resistant S. aureus These biomimetic strains are cost-effective, have low biohazard content, accurately mimic drug resistance, and can be produced with relative ease, thus illustrating their potential for widespread use as verification standards for diagnosis of a variety of diseases. Copyright © 2017 American Society for Microbiology.

Immune responses to the Mycobacterium tuberculosis-specific antigen ESAT-6 signal subclinical infection among contacts of tuberculosis patients

DEFF Research Database (Denmark)

Doherty, T Mark; Demissie, Abebech; Olobo, Joseph

2002-01-01

Diagnosis of latent Mycobacterium tuberculosis infection is considered essential for tuberculosis control but is hampered by the lack of specific reagents. We report that strong recognition of tuberculosis complex-specific antigen ESAT-6 by healthy household contacts of tuberculosis patients...... correlates with the subsequent development of active tuberculosis during a 2-year follow-up period....

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

DEFF Research Database (Denmark)

Thakur, Aneesh; Sharma, Mandeep; Katoch, Vipin C.

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 detectio...

Microbe Profile: Mycobacterium tuberculosis: Humanity's deadly microbial foe.

Science.gov (United States)

Gordon, Stephen V; Parish, Tanya

2018-04-01

Mycobacterium tuberculosis is an expert and deadly pathogen, causing the disease tuberculosis (TB) in humans. It has several notable features: the ability to enter non-replicating states for long periods and cause latent infection; metabolic remodelling during chronic infection; a thick, waxy cell wall; slow growth rate in culture; and intrinsic drug resistance and antibiotic tolerance. As a pathogen, M. tuberculosis has a complex relationship with its host, is able to replicate inside macrophages, and expresses diverse immunomodulatory molecules. M. tuberculosis currently causes over 1.8 million deaths a year, making it the world's most deadly human pathogen.

Phylogenetic analysis of vitamin B12-related metabolism in Mycobacterium tuberculosis

Directory of Open Access Journals (Sweden)

Douglas B. Young

2015-03-01

Full Text Available Comparison of genome sequences from clinical isolates of Mycobacterium tuberculosis with phylogenetically-related pathogens Mycobacterium marinum, Mycobacterium kansasii and Mycobacterium leprae reveals diversity amongst genes associated with vitamin B12-related metabolism. Diversity is generated by gene deletion events, differential acquisition of genes by horizontal transfer, and single nucleotide polymorphisms with predicted impact on protein function and transcriptional regulation. Differences in the B12 synthesis pathway, methionine biosynthesis, fatty acid catabolism, and DNA repair and replication are consistent with adaptations to different environmental niches and pathogenic lifestyles. While there is no evidence of further gene acquisition during expansion of the M. tuberculosis complex, the emergence of other forms of genetic diversity provides insights into continuing host-pathogen co-evolution and has the potential to identify novel targets for disease intervention.

DIGITAL DETECTION SYSTEM DESIGN OF MYCOBACTERIUM TUBERCULOSIS THROUGH EXTRACTION OF SPUTUM IMAGE USING NEURAL NETWORK METHOD

Directory of Open Access Journals (Sweden)

Franky Arisgraha

2012-01-01

Full Text Available Tuberculosis (TBC is an dangerous disease and many people has been infected. One of many important steps to control TBC effectively and efficiently is by increasing case finding using right method and accurate diagnostic. One of them is to detect Mycobacterium Tuberculosis inside sputum. Conventional detection of Mycobacterium Tuberculosis inside sputum can need a lot of time, so digitally detection method of Mycobacterium Tuberculosis was designed as an effort to get better result of detection. This method was designed by using combination between digital image processing method and Neural Network method. From testing report that was done, Mycobacterium can be detected with successful value reach 77.5% and training error less than 5%.

The Evolution of Strain Typing in the Mycobacterium tuberculosis Complex.

Science.gov (United States)

Merker, Matthias; Kohl, Thomas A; Niemann, Stefan; Supply, Philip

2017-01-01

Tuberculosis (TB) is a contagious disease with a complex epidemiology. Therefore, molecular typing (genotyping) of Mycobacterium tuberculosis complex (MTBC) strains is of primary importance to effectively guide outbreak investigations, define transmission dynamics and assist global epidemiological surveillance of the disease. Large-scale genotyping is also needed to get better insights into the biological diversity and the evolution of the pathogen. Thanks to its shorter turnaround and simple numerical nomenclature system, mycobacterial interspersed repetitive unit-variable-number tandem repeat (MIRU-VNTR) typing, based on 24 standardized plus 4 hypervariable loci, optionally combined with spoligotyping, has replaced IS6110 DNA fingerprinting over the last decade as a gold standard among classical strain typing methods for many applications. With the continuous progress and decreasing costs of next-generation sequencing (NGS) technologies, typing based on whole genome sequencing (WGS) is now increasingly performed for near complete exploitation of the available genetic information. However, some important challenges remain such as the lack of standardization of WGS analysis pipelines, the need of databases for sharing WGS data at a global level, and a better understanding of the relevant genomic distances for defining clusters of recent TB transmission in different epidemiological contexts. This chapter provides an overview of the evolution of genotyping methods over the last three decades, which culminated with the development of WGS-based methods. It addresses the relative advantages and limitations of these techniques, indicates current challenges and potential directions for facilitating standardization of WGS-based typing, and provides suggestions on what method to use depending on the specific research question.

Variable host-pathogen compatibility in Mycobacterium tuberculosis.

NARCIS (Netherlands)

Gagneux, Sebastien; DeRiemer, Kathryn; Van, Tran; Kato-Maeda, Midori; Jong, Bouke C de; Narayanan, Sujatha; Nicol, Mark; Niemann, Stefan; Kremer, Kristin; Gutierrez, M Cristina; Hilty, Markus; Hopewell, Philip C; Small, Peter M

2006-01-01

Mycobacterium tuberculosis remains a major cause of morbidity and mortality worldwide. Studies have reported human pathogens to have geographically structured population genetics, some of which have been linked to ancient human migrations. However, no study has addressed the potential evolutionary

Beta-lactamases of Mycobacterium tuberculosis and Mycobacterium kansasii.

Science.gov (United States)

Segura, C; Salvadó, M

1997-09-01

Re-emergence of infectious diseases caused by mycobacteria as well as the emergence of multiresistant strains of Mycobacterium has promoted the research on the use of beta-lactames in the treatment of such diseases. Mycobacteria produce beta-lactamases: M. tuberculosis produces a wide-spectrum beta-lactamase whose behaviour mimicks those of Gram-negative bacteria. M. kansasii produces also beta-lactamase which can be inhibited by clavulanic acid. An overview on beta-lactamases from both species is reported.

Specific T-cell epitopes for immunoassay-based diagnosis of Mycobacterium tuberculosis infection

DEFF Research Database (Denmark)

Brock, I; Weldingh, K; Leyten, EM

2004-01-01

Specific T-cell epitopes for immunoassay-based diagnosis of Mycobacterium tuberculosis infection.Brock I, Weldingh K, Leyten EM, Arend SM, Ravn P, Andersen P. Department of Infectious Disease Immunology, Statens Serum Institute, Artillerivej 5, DK-2300 Copenhagen S, Denmark. The currently used...... method for immunological detection of tuberculosis infection, the tuberculin skin test, has low specificity. Antigens specific for Mycobacterium tuberculosis to replace purified protein derivative are therefore urgently needed. We have performed a rigorous assessment of the diagnostic potential of four...... recently identified antigens (Rv2653, Rv2654, Rv3873, and Rv3878) from genomic regions that are lacking from the Mycobacterium bovis bacillus Calmette-Guerin (BCG) vaccine strains as well as from the most common nontuberculous mycobacteria. The fine specificity of potential epitopes in these molecules...

Population Genomics of Mycobacterium tuberculosis in Ethiopia Contradicts the Virgin Soil Hypothesis for Human Tuberculosis in Sub-Saharan Africa.

Science.gov (United States)

Comas, Iñaki; Hailu, Elena; Kiros, Teklu; Bekele, Shiferaw; Mekonnen, Wondale; Gumi, Balako; Tschopp, Rea; Ameni, Gobena; Hewinson, R Glyn; Robertson, Brian D; Goig, Galo A; Stucki, David; Gagneux, Sebastien; Aseffa, Abraham; Young, Douglas; Berg, Stefan

2015-12-21

Colonial medical reports claimed that tuberculosis (TB) was largely unknown in Africa prior to European contact, providing a "virgin soil" for spread of TB in highly susceptible populations previously unexposed to the disease [1, 2]. This is in direct contrast to recent phylogenetic models which support an African origin for TB [3-6]. To address this apparent contradiction, we performed a broad genomic sampling of Mycobacterium tuberculosis in Ethiopia. All members of the M. tuberculosis complex (MTBC) arose from clonal expansion of a single common ancestor [7] with a proposed origin in East Africa [3, 4, 8]. Consistent with this proposal, MTBC lineage 7 is almost exclusively found in that region [9-11]. Although a detailed medical history of Ethiopia supports the view that TB was rare until the 20(th) century [12], over the last century Ethiopia has become a high-burden TB country [13]. Our results provide further support for an African origin for TB, with some genotypes already present on the continent well before European contact. Phylogenetic analyses reveal a pattern of serial introductions of multiple genotypes into Ethiopia in association with human migration and trade. In place of a "virgin soil" fostering the spread of TB in a previously naive population, we propose that increased TB mortality in Africa was driven by the introduction of European strains of M. tuberculosis alongside expansion of selected indigenous strains having biological characteristics that carry a fitness benefit in the urbanized settings of post-colonial Africa. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

Identification of a novel 27-kDa protein from Mycobacterium tuberculosis culture fluid by a monoclonal antibody specific for the Mycobacterium tuberculosis complex

NARCIS (Netherlands)

Rambukkana, A.; Das, P. K.; Kolk, A. H.; Burggraaf, J. D.; Kuijper, S.; Harboe, M.

1993-01-01

Mycobacterium tuberculosis antigens inducing species-specific immune responses are likely to be particularly important for serodiagnosis or for skin testing of tuberculosis. In the present study, we describe the characterization of two novel monoclonal antibodies (MoAbs) A3h4 (IgG2a) and B5g1 (IgM)

Bioaktivitas Ekstrak Metanol Daun Pegagan (Centella Asiatica L. Terhadap Pertumbuhan Bakteri Mycobacterium Tuberculosis

Directory of Open Access Journals (Sweden)

Yusran Yusran

2016-01-01

Full Text Available Plants gotu kola (Centella Asiatica L .Urban is a wild plant that efficacious as remedies traditional cure disease tuberculosis (TB.TB is disease contagious infection caused by bacteria mycobacterium tuberculosis. Research aims to understand the ability extract methanol leaves gotu kola red and leaves gotu kola green and determines the concentration optimal extract methanol leaves gotu kola red and leaves gotu kola green and to know the comparison between extract methanol leaves gotu kola red with an extract methanol leaves gotu kola green in inhibits the activity of mycobacterium tuberculosis.Extraction done with the methods maceration use methanol and continued with evaporation until obtained extract viscous .Testing antibacterial activity done in a microscopic observation drug susceptibility ( mods use plate petri dish 24 hole with the variation of concentration ie 20%,40%, 60%, 80% and 100%.The results of testing show that extracts methanol leaves gotu kola red and leaves gotu kola green positive capable of inhibiting the growth of bacteria mycobacterium tuberculosis with inhibition optimal in concentration 80 % and 100 % characterized by the absence of growth bacteria colonies which are (- or 0 %.Extract methanol leaves gotu kola green capable of inhibiting the growth of bacteria mycobacterium tuberculosis better than extract methanol leaves gotu kola red seen in concentration 40% and 60%.

Proteasomal control of cytokinin synthesis protects Mycobacterium tuberculosis against nitric oxide

Science.gov (United States)

Samanovic, Marie I.; Tu, Shengjiang; Novák, Ondřej; Iyer, Lakshminarayan M.; McAllister, Fiona E.; Aravind, L.; Gygi, Steven P.; Hubbard, Stevan R.; Strnad, Miroslav; Darwin, K. Heran

2015-01-01

Summary One of several roles of the Mycobacterium tuberculosis proteasome is to defend against host-produced nitric oxide (NO), a free radical that can damage numerous biological macromolecules. Mutations that inactivate proteasomal degradation in Mycobacterium tuberculosis result in bacteria that are hypersensitive to NO and attenuated for growth in vivo, but it was not known why. To elucidate the link between proteasome function, NO-resistance, and pathogenesis, we screened for suppressors of NO hypersensitivity in a mycobacterial proteasome ATPase mutant and identified mutations in Rv1205. We determined that Rv1205 encodes a pupylated proteasome substrate. Rv1205 is a homologue of the plant enzyme LONELY GUY, which catalyzes the production of hormones called cytokinins. Remarkably, we report for the first time that an obligate human pathogen secretes several cytokinins. Finally, we determined that the Rv1205-dependent accumulation of cytokinin breakdown products is likely responsible for the sensitization of Mycobacterium tuberculosis proteasome-associated mutants to NO. PMID:25728768

Insights on the Emergence of Mycobacterium tuberculosis from the Analysis of Mycobacterium kansasii

Science.gov (United States)

Wang, Joyce; McIntosh, Fiona; Radomski, Nicolas; Dewar, Ken; Simeone, Roxane; Enninga, Jost; Brosch, Roland; Rocha, Eduardo P.; Veyrier, Frédéric J.; Behr, Marcel A.

2015-01-01

By phylogenetic analysis, Mycobacterium kansasii is closely related to Mycobacterium tuberculosis. Yet, although both organisms cause pulmonary disease, M. tuberculosis is a global health menace, whereas M. kansasii is an opportunistic pathogen. To illuminate the differences between these organisms, we have sequenced the genome of M. kansasii ATCC 12478 and its plasmid (pMK12478) and conducted side-by-side in vitro and in vivo investigations of these two organisms. The M. kansasii genome is 6,432,277 bp, more than 2 Mb longer than that of M. tuberculosis H37Rv, and the plasmid contains 144,951 bp. Pairwise comparisons reveal conserved and discordant genes and genomic regions. A notable example of genomic conservation is the virulence locus ESX-1, which is intact and functional in the low-virulence M. kansasii, potentially mediating phagosomal disruption. Differences between these organisms include a decreased predicted metabolic capacity, an increased proportion of toxin–antitoxin genes, and the acquisition of M. tuberculosis-specific genes in the pathogen since their common ancestor. Consistent with their distinct epidemiologic profiles, following infection of C57BL/6 mice, M. kansasii counts increased by less than 10-fold over 6 weeks, whereas M. tuberculosis counts increased by over 10,000-fold in just 3 weeks. Together, these data suggest that M. kansasii can serve as an image of the environmental ancestor of M. tuberculosis before its emergence as a professional pathogen, and can be used as a model organism to study the switch from an environmental opportunistic pathogen to a professional host-restricted pathogen. PMID:25716827

Pyrosequencing for Rapid Detection of Mycobacterium tuberculosis Resistance to Rifampin, Isoniazid, and Fluoroquinolones ▿

Science.gov (United States)

Bravo, Lulette Tricia C.; Tuohy, Marion J.; Ang, Concepcion; Destura, Raul V.; Mendoza, Myrna; Procop, Gary W.; Gordon, Steven M.; Hall, Geraldine S.; Shrestha, Nabin K.

2009-01-01

After isoniazid and rifampin (rifampicin), the next pivotal drug class in Mycobacterium tuberculosis treatment is the fluoroquinolone class. Mutations in resistance-determining regions (RDR) of the rpoB, katG, and gyrA genes occur with frequencies of 97%, 50%, and 85% among M. tuberculosis isolates resistant to rifampin, isoniazid, and fluoroquinolones, respectively. Sequences are highly conserved, and certain mutations correlate well with phenotypic resistance. We developed a pyrosequencing assay to determine M. tuberculosis genotypic resistance to rifampin, isoniazid, and fluoroquinolones. We characterized 102 M. tuberculosis clinical isolates from the Philippines for susceptibility to rifampin, isoniazid, and ofloxacin by using the conventional submerged-disk proportion method and validated our pyrosequencing assay using these isolates. DNA was extracted and amplified by using PCR primers directed toward the RDR of the rpoB, katG, and gyrA genes, and pyrosequencing was performed on the extracts. The M. tuberculosis H37Rv strain (ATCC 25618) was used as the reference strain. The sensitivities and specificities of pyrosequencing were 96.7% and 97.3%, 63.8% and 100%, and 70.0% and 100% for the detection of resistance to rifampin, isoniazid, and ofloxacin, respectively. Pyrosequencing is thus a rapid and accurate method for detecting M. tuberculosis resistance to these three drugs. PMID:19846642

Inactivation of Mycobacterium paratuberculosis and Mycobacterium tuberculosis in fresh soft cheese by gamma radiation

International Nuclear Information System (INIS)

Badr, Hesham M.

2011-01-01

The effectiveness of gamma irradiation on the inactivation of Mycobacterium paratuberculosis, Mycobacterium bovis and Mycobacterium tuberculosis in fresh soft cheese that prepared from artificially inoculated milk samples was studied. Irradiation at dose of 2 kGy was sufficient for the complete inactivation of these mycobacteria as they were not detected in the treated samples during storage at 4±1 o C for 15 days. Moreover, irradiation of cheese samples, that were prepared from un-inoculated milk, at this effective dose had no significant effects on their gross composition and contents from riboflavin, niacin and pantothenic acid, while significant decreases in vitamin A and thiamin were observed. In addition, irradiation of cheese samples had no significant effects on their pH and nitrogen fractions contents, except for the contents of ammonia, which showed a slight, but significant, increases due to irradiation. The analysis of cheese fats indicated that irradiation treatment induced significant increase in their oxidation parameters and contents from free fatty acids; however, the observed increases were relatively low. On the other hand, irradiation of cheese samples induced no significant alterations on their sensory properties. Thus, irradiation dose of 2 kGy can be effectively applied to ensure the safety of soft cheese with regards to these harmful mycobacteria. - Highlights: → We examined the effectiveness of gamma irradiation on inactivation of Mycobacterium paratuberculosis, Mycobacterium bovis and Mycobacterium tuberculosis in fresh soft cheese. → Irradiation at dose of 2 kGy was sufficient for complete inactivation of these mycobacteria. → Irradiation of cheese samples induced no significant alterations on their sensory properties.

Inactivation of Mycobacterium paratuberculosis and Mycobacterium tuberculosis in fresh soft cheese by gamma radiation

Energy Technology Data Exchange (ETDEWEB)

Badr, Hesham M., E-mail: heshambadr_aea@yahoo.co.uk [Atomic Energy Authority, Nuclear Research Center, Abou Zaabal, P.O. Box 13759 Cairo (Egypt)

2011-11-15

The effectiveness of gamma irradiation on the inactivation of Mycobacterium paratuberculosis, Mycobacterium bovis and Mycobacterium tuberculosis in fresh soft cheese that prepared from artificially inoculated milk samples was studied. Irradiation at dose of 2 kGy was sufficient for the complete inactivation of these mycobacteria as they were not detected in the treated samples during storage at 4{+-}1 {sup o}C for 15 days. Moreover, irradiation of cheese samples, that were prepared from un-inoculated milk, at this effective dose had no significant effects on their gross composition and contents from riboflavin, niacin and pantothenic acid, while significant decreases in vitamin A and thiamin were observed. In addition, irradiation of cheese samples had no significant effects on their pH and nitrogen fractions contents, except for the contents of ammonia, which showed a slight, but significant, increases due to irradiation. The analysis of cheese fats indicated that irradiation treatment induced significant increase in their oxidation parameters and contents from free fatty acids; however, the observed increases were relatively low. On the other hand, irradiation of cheese samples induced no significant alterations on their sensory properties. Thus, irradiation dose of 2 kGy can be effectively applied to ensure the safety of soft cheese with regards to these harmful mycobacteria. - Highlights: > We examined the effectiveness of gamma irradiation on inactivation of Mycobacterium paratuberculosis, Mycobacterium bovis and Mycobacterium tuberculosis in fresh soft cheese. > Irradiation at dose of 2 kGy was sufficient for complete inactivation of these mycobacteria. > Irradiation of cheese samples induced no significant alterations on their sensory properties.

Genomic diversity among drug sensitive and multidrug resistant isolates of Mycobacterium tuberculosis with identical DNA fingerprints.

Directory of Open Access Journals (Sweden)

Stefan Niemann

2009-10-01

Full Text Available Mycobacterium tuberculosis complex (MTBC, the causative agent of tuberculosis (TB, is characterized by low sequence diversity making this bacterium one of the classical examples of a genetically monomorphic pathogen. Because of this limited DNA sequence variation, routine genotyping of clinical MTBC isolates for epidemiological purposes relies on highly discriminatory DNA fingerprinting methods based on mobile and repetitive genetic elements. According to the standard view, isolates exhibiting the same fingerprinting pattern are considered direct progeny of the same bacterial clone, and most likely reflect ongoing transmission or disease relapse within individual patients.Here we further investigated this assumption and used massively parallel whole-genome sequencing to compare one drug-susceptible (K-1 and one multidrug resistant (MDR isolate (K-2 of a rapidly spreading M. tuberculosis Beijing genotype clone from a high incidence region (Karakalpakstan, Uzbekistan. Both isolates shared the same IS6110 RFLP pattern and the same allele at 23 out of 24 MIRU-VNTR loci. We generated 23.9 million (K-1 and 33.0 million (K-2 paired 50 bp purity filtered reads corresponding to a mean coverage of 483.5 fold and 656.1 fold respectively. Compared with the laboratory strain H37Rv both Beijing isolates shared 1,209 SNPs. The two Beijing isolates differed by 130 SNPs and one large deletion. The susceptible isolate had 55 specific SNPs, while the MDR variant had 75 specific SNPs, including the five known resistance-conferring mutations.Our results suggest that M. tuberculosis isolates exhibiting identical DNA fingerprinting patterns can harbour substantial genomic diversity. Because this heterogeneity is not captured by traditional genotyping of MTBC, some aspects of the transmission dynamics of tuberculosis could be missed or misinterpreted. Furthermore, a valid differentiation between disease relapse and exogenous reinfection might be impossible using

Genomic diversity among drug sensitive and multidrug resistant isolates of Mycobacterium tuberculosis with identical DNA fingerprints.

Science.gov (United States)

Niemann, Stefan; Köser, Claudio U; Gagneux, Sebastien; Plinke, Claudia; Homolka, Susanne; Bignell, Helen; Carter, Richard J; Cheetham, R Keira; Cox, Anthony; Gormley, Niall A; Kokko-Gonzales, Paula; Murray, Lisa J; Rigatti, Roberto; Smith, Vincent P; Arends, Felix P M; Cox, Helen S; Smith, Geoff; Archer, John A C

2009-10-12

Mycobacterium tuberculosis complex (MTBC), the causative agent of tuberculosis (TB), is characterized by low sequence diversity making this bacterium one of the classical examples of a genetically monomorphic pathogen. Because of this limited DNA sequence variation, routine genotyping of clinical MTBC isolates for epidemiological purposes relies on highly discriminatory DNA fingerprinting methods based on mobile and repetitive genetic elements. According to the standard view, isolates exhibiting the same fingerprinting pattern are considered direct progeny of the same bacterial clone, and most likely reflect ongoing transmission or disease relapse within individual patients. Here we further investigated this assumption and used massively parallel whole-genome sequencing to compare one drug-susceptible (K-1) and one multidrug resistant (MDR) isolate (K-2) of a rapidly spreading M. tuberculosis Beijing genotype clone from a high incidence region (Karakalpakstan, Uzbekistan). Both isolates shared the same IS6110 RFLP pattern and the same allele at 23 out of 24 MIRU-VNTR loci. We generated 23.9 million (K-1) and 33.0 million (K-2) paired 50 bp purity filtered reads corresponding to a mean coverage of 483.5 fold and 656.1 fold respectively. Compared with the laboratory strain H37Rv both Beijing isolates shared 1,209 SNPs. The two Beijing isolates differed by 130 SNPs and one large deletion. The susceptible isolate had 55 specific SNPs, while the MDR variant had 75 specific SNPs, including the five known resistance-conferring mutations. Our results suggest that M. tuberculosis isolates exhibiting identical DNA fingerprinting patterns can harbour substantial genomic diversity. Because this heterogeneity is not captured by traditional genotyping of MTBC, some aspects of the transmission dynamics of tuberculosis could be missed or misinterpreted. Furthermore, a valid differentiation between disease relapse and exogenous reinfection might be impossible using standard

Structural studies on Mycobacterium tuberculosis RecA

Indian Academy of Sciences (India)

Structures of crystals of Mycobacterium tuberculosis RecA, grown and analysed under different conditions, provide insights into hitherto underappreciated details of molecular structure and plasticity. In particular, they yield information on the invariant and variable features of the geometry of the P-loop, whose binding to ATP ...

DNA repair in Mycobacterium tuberculosis revisited.

Science.gov (United States)

Dos Vultos, Tiago; Mestre, Olga; Tonjum, Tone; Gicquel, Brigitte

2009-05-01

Our understanding of Mycobacterium tuberculosis DNA repair mechanisms is still poor compared with that of other bacterial organisms. However, the publication of the first complete M. tuberculosis genome sequence 10 years ago boosted the study of DNA repair systems in this organism. A first step in the elucidation of M. tuberculosis DNA repair mechanisms was taken by Mizrahi and Andersen, who identified homologs of genes involved in the reversal or repair of DNA damage in Escherichia coli and related organisms. Genes required for nucleotide excision repair, base excision repair, recombination, and SOS repair and mutagenesis were identified. Notably, no homologs of genes involved in mismatch repair were identified. Novel characteristics of the M. tuberculosis DNA repair machinery have been found over the last decade, such as nonhomologous end joining, the presence of Mpg, ERCC3 and Hlr - proteins previously presumed to be produced exclusively in mammalian cells - and the recently discovered bifunctional dCTP deaminase:dUTPase. The study of these systems is important to develop therapeutic agents that can counteract M. tuberculosis evolutionary changes and to prevent adaptive events resulting in antibiotic resistance. This review summarizes our current understanding of the M. tuberculosis DNA repair system.

Specific T-cell epitopes for immunoassay-based diagnosis of Mycobacterium tuberculosis infection

DEFF Research Database (Denmark)

Brock, I; Weldingh, K; Leyten, EM

2004-01-01

Specific T-cell epitopes for immunoassay-based diagnosis of Mycobacterium tuberculosis infection.Brock I, Weldingh K, Leyten EM, Arend SM, Ravn P, Andersen P. Department of Infectious Disease Immunology, Statens Serum Institute, Artillerivej 5, DK-2300 Copenhagen S, Denmark. The currently used...... method for immunological detection of tuberculosis infection, the tuberculin skin test, has low specificity. Antigens specific for Mycobacterium tuberculosis to replace purified protein derivative are therefore urgently needed. We have performed a rigorous assessment of the diagnostic potential of four...... selected and combined the specific peptide stretches from the four proteins not recognized by M. bovis BCG-vaccinated individuals. These peptide stretches were tested with peripheral blood mononuclear cells obtained from patients with microscopy- or culture-confirmed tuberculosis and from healthy M. bovis...

Molecular epidemiology and evolutionary genetics of Mycobacterium tuberculosis in Taipei.

Science.gov (United States)

Dou, Horng-Yunn; Tseng, Fan-Chen; Lin, Chih-Wei; Chang, Jia-Ru; Sun, Jun-Ren; Tsai, Wen-Shing; Lee, Shi-Yi; Su, Ih-Jen; Lu, Jang-Jih

2008-12-22

The control of tuberculosis in densely populated cities is complicated by close human-to-human contacts and potential transmission of pathogens from multiple sources. We conducted a molecular epidemiologic analysis of 356 Mycobacterium tuberculosis (MTB) isolates from patients presenting pulmonary tuberculosis in metropolitan Taipei. Classical antibiogram studies and genetic characterization, using mycobacterial interspersed repetitive-unit-variable-number tandem-repeat (MIRU-VNTR) typing and spoligotyping, were applied after culture. A total of 356 isolates were genotyped by standard spoligotyping and the strains were compared with in the international spoligotyping database (SpolDB4). All isolates were also categorized using the 15 loci MIRU-VNTR typing method and combin with NTF locus and RD deletion analyses. Of 356 isolates spoligotyped, 290 (81.4%) displayed known spoligotypes and 66 were not identified in the database. Major spoligotypes found were Beijing lineages (52.5%), followed by Haarlem lineages (13.5%) and EAI plus EAI-like lineages (11%). When MIRU-VNTR was employed, 140 patterns were identified, including 36 clusters by 252 isolates and 104 unique patterns, and the largest cluster comprised 95 isolates from the Beijing family. The combination of spoligotyping and MIRU-VNTR revealed that 236 (67%) of the 356 isolates were clustered in 43 genotypes. Strains of the Beijing family was more likely to be of modern strain and a higher percentage of multiple drug resistance than other families combined (P = 0.08). Patients infected with Beijing strains were younger than those with other strains (mean 58.7 vs. 64.2, p = 0.02). Moreover, 85.3% of infected persons younger than 25 years had Beijing modern strain, suggesting a possible recent spread in the young population by this family of TB strain in Taipei. Our data on MTB genotype in Taipei suggest that MTB infection has not been optimally controlled. Control efforts should be reinforced in view of the

Clustered tuberculosis in a low-burden country

DEFF Research Database (Denmark)

Kamper-Jørgensen, Z; Andersen, A B; Kok-Jensen, A

2012-01-01

Molecular genotyping of Mycobacterium tuberculosis has proved to be a powerful tool in tuberculosis surveillance, epidemiology, and control. Based on results obtained through 15 years of nationwide IS6110 restriction fragment length polymorphism (RFLP) genotyping of M. tuberculosis cases in Denmark......, a country on the way toward tuberculosis elimination, we discuss M. tuberculosis transmission dynamics and point to areas for control interventions. Cases with 100% identical genotypes (RFLP patterns) were defined as clustered, and a cluster was defined as cases with an identical genotype. Of 4,601 included...... cases, corresponding to 76% of reported and 97% of culture-verified tuberculosis cases in the country, 56% were clustered, of which 69% were Danes. Generally, Danes were more often in large clusters (= 50 persons), older (mean age, 45 years), and male (male/female ratio, 2.5). Also, Danes had a higher...

Efficacy of amikacin and ciprofloxacin against clinical isolates of mycobacterium tuberculosis

International Nuclear Information System (INIS)

Satti, M.; Faqir, F.; Sattar, A.; Abbasi, S.; Butt, T.; Karamat, K.A.; Abidi, M.

2010-01-01

Background: Tuberculosis was a leading cause of death at the turn of the 20 century and continues to be one of the medical scourges of mankind. Before the availability of antimicrobial drugs the cornerstone of treatment was rest in the open air in sanatoria. The major breakthrough in treatment of tuberculosis came with the discovery of Streptomycin. Later, INH, Ethambutol, Pyrazinamide, Rifampicin were added to the arsenal. Objective of this study was to determine the sensitivity of clinical isolates of Mycobacterium tuberculosis against two second-line anti-tuberculosis drugs, Amikacin and Ciprofloxacin. Methods: This cross-sectional study was conducted at Department of Microbiology, Armed Forces Institute of Pathology (AFIP) Rawalpindi. All routine clinical samples received for acid fast bacilli (AFB) in the Department of Microbiology, AFIP, Rawalpindi were processed by modified Petroff's technique and inoculated on Lowenstein Jensen (LJ) medium and Bactec 460 Mycobacterium tuberculosis culture system. After identification of M. tuberculosis sensitivity was performed against first-line anti-tuberculosis drugs. Then susceptibility of M. tuberculosis isolates against Amikacin and Ciprofloxacin was performed on LJ medium. H37Rv was used as control strain. Results: Results were interpreted using resistance ratio method. Out of 100 M. tuberculosis isolates, 98% were sensitive to Amikacin and 97% to Ciprofloxacin. Conclusion: Amikacin and Ciprofloxacin are very effective second line anti-tuberculosis drugs against tuberculosis isolates in our set-up. (author)

Drug susceptibility testing of Mycobacterium tuberculosis to fluoroquinolones

DEFF Research Database (Denmark)

Johansen, I S; Larsen, A R; Sandven, P

2003-01-01

In the first attempt to establish a quality assurance programme for susceptibility testing of Mycobacterium tuberculosis to fluoroquinolones, 20 strains with different fluoroquinolone susceptibility patterns were distributed by the Supranational Reference Laboratory in Stockholm to the other...

Bloodstream Infections with Mycobacterium tuberculosis among HIV patients

Centers for Disease Control (CDC) Podcasts

This podcast looks at bloodstream infections with Mycobacterium tuberculosis and other pathogens among outpatients infected with HIV in Southeast Asia. CDC health scientist Kimberly McCarthy discusses the study and why bloodstream infections occur in HIV-infected populations.

Mycobacterium tuberculosis: nepřekonatelný nepřítel

Czech Academy of Sciences Publication Activity Database

Machová, Iva; Pichová, Iva

2014-01-01

Roč. 24, č. 4 (2014), s. 98-100 ISSN 1210-1737 R&D Projects: GA MŠk LO1302; GA MŠk(CZ) 7E11070 EU Projects: European Commission(XE) 241587 - SYSTEMTB Institutional support: RVO:61388963 Keywords : Mycobacterium tuberculosis * tuberculosis * latent infection * resistance * metabolism * drugs Subject RIV: CE - Biochemistry

Molecular epidemiology of Mycobacterium tuberculosis clinical isolates in Southwest Ireland.

LENUS (Irish Health Repository)

Ojo, Olabisi O

2010-10-01

Tuberculosis has had significant effects on Ireland over the past two centuries, causing persistently higher morbidity and mortality than in neighbouring countries until the last decade. This study describes the results of genotyping and drug susceptibility testing of 171 strains of Mycobacterium tuberculosis complex isolated between January 2004 and December 2006 in a region of Ireland centred on the city of Cork. Spoligotype comparisons were made with the SpolDB4 database and clustered 130 strains in 23 groups, forty-one strains showed unique Spoligotyping patterns. The commonest spoligotypes detected were ST0137 (X2) (16.9%), and ST0351 (15.8%) (\\'U\\' clade). The major spoligotype clades were X (26.2%), U (19.3%), T (15.2%), Beijing (5.9%), Haarlem (4.7%), LAM (4.1%), BOVIS (1.75%), with 12.9% unassigned strains. A 24-locus VNTR genotyping produced 15 clusters containing 49 isolates, with high discrimination index (HGDI>0.99). A combination of Spoligotyping and VNTR reduced the number of clustered isolates to 47 in 15 clusters (27.5%). This study identified ST351 as common among Irish nationals, and found a low rate of drug resistance with little evidence of transmission of drug resistant strains. Strain clustering was significantly associated with age under 55 years and Irish nationality. Only strains of Euro-American lineage formed clusters. Molecular typing did not completely coincide with the results of contact investigations.

Clustering of Mycobacterium tuberculosis strains from foreign-born patients in Korea.

Science.gov (United States)

Jeon, Christie Y; Kang, Heeyoon; Kim, Mihye; Murray, Megan B; Kim, Heejin; Cho, Eun Hee; Park, Young Kil

2011-12-01

Information on drug resistance and transmission patterns of tuberculosis (TB) in foreign-born patients is lacking in Asia where immigration is increasing. We examined the drug-resistance profiles of 288 Mycobacterium tuberculosis isolates from foreign-born patients in South Korea, and assessed for potential transmission in the host country by analysing their IS6110 genotypes, as well as those of 4780 strains from native Korean TB patients. The prevalence of multidrug-resistant (MDR) TB was 9.7% and 42% among new and previously treated patients, respectively. Chinese nationality was associated with MDR TB (OR(China)=3.0, 95% CI 1.1-9.3). Of the 288 strains, 51 (17.7%) formed 31 clusters, of which 22 were identical to strains from native Koreans. A number of strains belonged to the K family, subtypes known to occur endemically in Korea. MDR TB was common, and clustering patterns showed potential cross-cultural transmission among foreign-born TB patients. Further molecular epidemiological studies of all isolates in the area are needed to determine the extent of international TB transmission in Asia. © 2011 SGM

Combating highly resistant emerging pathogen Mycobacterium abscessus and Mycobacterium tuberculosis with novel salicylanilide esters and carbamates.

Science.gov (United States)

Baranyai, Zsuzsa; Krátký, Martin; Vinšová, Jarmila; Szabó, Nóra; Senoner, Zsuzsanna; Horváti, Kata; Stolaříková, Jiřina; Dávid, Sándor; Bősze, Szilvia

2015-08-28

In the Mycobacterium genus over one hundred species are already described and new ones are periodically reported. Species that form colonies in a week are classified as rapid growers, those requiring longer periods (up to three months) are the mostly pathogenic slow growers. More recently, new emerging species have been identified to lengthen the list, all rapid growers. Of these, Mycobacterium abscessus is also an intracellular pathogen and it is the most chemotherapy-resistant rapid-growing mycobacterium. In addition, the cases of multidrug-resistant Mycobacterium tuberculosis infection are also increasing. Therefore there is an urgent need to find new active molecules against these threatening strains. Based on previous results, a series of salicylanilides, salicylanilide 5-chloropyrazinoates and carbamates was designed, synthesized and characterised. The compounds were evaluated for their in vitro activity on M. abscessus, susceptible M. tuberculosis H37Rv, multidrug-resistant (MDR) M. tuberculosis MDR A8, M. tuberculosis MDR 9449/2006 and on the extremely-resistant Praha 131 (XDR) strains. All derivatives exhibited a significant activity with minimum inhibitory concentrations (MICs) in the low micromolar range. Eight salicylanilide carbamates and two salicylanilide esters exhibited an excellent in vitro activity on M. abscessus with MICs from 0.2 to 2.1 μM, thus being more effective than ciprofloxacin and gentamicin. This finding is potentially promising, particularly, as M. abscessus is a threateningly chemotherapy-resistant species. M. tuberculosis H37Rv was inhibited with MICs from 0.2 μM, and eleven compounds have lower MICs than isoniazid. Salicylanilide esters and carbamates were found that they were effective also on MDR and XDR M. tuberculosis strains with MICs ≥1.0 μM. The in vitro cytotoxicity (IC50) was also determined on human MonoMac-6 cells, and selectivity index (SI) of the compounds was established. In general, salicylanilide

Evidence of presence of Mycobacterium tuberculosis in bovine tissue samples by multiplex PCR: possible relevance to reverse zoonosis.

Science.gov (United States)

Mittal, M; Chakravarti, S; Sharma, V; Sanjeeth, B S; Churamani, C P; Kanwar, N S

2014-04-01

Bovine tuberculosis, caused by Mycobacterium bovis, remains one of the most important zoonotic health concerns worldwide. The transmission of Mycobacterium tuberculosis from humans to animals also occurs especially in countries where there is close interaction of humans with the animals. In the present study, thirty bovine lung tissue autopsy samples from an organized dairy farm located in North India were screened for the presence of Mycobacterium tuberculosis complex by smear microscopy, histopathological findings and PCR. Differential diagnosis of M. tuberculosis and M. bovis was made based on the deletion of mce-3 operon in M. bovis. The present study found eight of these samples positive for M. tuberculosis by multiplex PCR. Sequencing was performed on two PCR-positive representative samples and on annotation, and BLAST analysis confirmed the presence of gene fragment specific to Mycobacterium tuberculosis. The presence of M. tuberculosis in all the positive samples raises the possibility of human-to-cattle transmission and possible adaptation of this organism in bovine tissues. This study accentuates the importance of screening and differential diagnosis of Mycobacterium tuberculosis complex in humans and livestock for adopting effective TB control and eradication programmes. © 2014 Blackwell Verlag GmbH.

Comparison of the UDP-N-Acetylmuramate:l-Alanine Ligase Enzymes from Mycobacterium tuberculosis and Mycobacterium leprae

OpenAIRE

Mahapatra, Sebabrata; Crick, Dean C.; Brennan, Patrick J.

2000-01-01

In the peptidoglycan of Mycobacterium leprae, l-alanine of the side chain is replaced by glycine. When expressed in Escherichia coli, MurC (UDP-N-acetyl-muramate:l-alanine ligase) of M. leprae showed Km and Vmax for l-alanine and glycine similar to those of Mycobacterium tuberculosis MurC, suggesting that another explanation should be sought for the presence of glycine.

Characterization of the genetic diversity of Mycobacterium tuberculosis in São Paulo city, Brazil.

Science.gov (United States)

Mendes, Natália H; Melo, Fernando Af; Santos, Adolfo Cb; Pandolfi, José Rc; Almeida, Elisabete A; Cardoso, Rosilene F; Berghs, Henri; David, Suzana; Johansen, Faber K; Espanha, Lívia G; Leite, Sergio Ra; Leite, Clarice Qf

2011-07-29

Tuberculosis is a major health problem in São Paulo, Brazil, which is the most populous and one of the most cosmopolitan cities in South America. To characterize the genetic diversity of Mycobacterium tuberculosis in the population of this city, the genotyping techniques of spoligotyping and MIRU were applied to 93 isolates collected in two consecutive years from 93 different tuberculosis patients residing in São Paulo city and attending the Clemente Ferreira Institute (the reference clinic for the treatment of tuberculosis). Spoligotyping generated 53 different spoligotype patterns. Fifty-one isolates (54.8%) were grouped into 13 spoligotyping clusters. Seventy- two strains (77.4%) showed spoligotypes described in the international databases (SpolDB4, SITVIT), and 21 (22.6%) showed unidentified patterns. The most frequent spoligotype families were Latin American Mediterranean (LAM) (26 isolates), followed by the T family (24 isolates) and Haarlem (H) (11 isolates), which together accounted for 65.4% of all the isolates. These three families represent the major genotypes found in Africa, Central America, South America and Europe. Six Spoligo-International-types (designated SITs by the database) comprised 51.8% (37/72) of all the identified spoligotypes (SIT53, SIT50, SIT42, SIT60, SIT17 and SIT1). Other SITs found in this study indicated the great genetic diversity of M. tuberculosis, reflecting the remarkable ethnic diversity of São Paulo city inhabitants. The MIRU technique was more discriminatory and did not identify any genetic clusters with 100% similarity among the 93 isolates. The allelic analysis showed that MIRU loci 26, 40, 23 and 10 were the most discriminatory. When MIRU and spoligotyping techniques were combined, all isolates grouped in the 13 spoligotyping clusters were separated. Our data indicated the genomic stability of over 50% of spoligotypes identified in São Paulo and the great genetic diversity of M. tuberculosis isolates in the remaining

Characterization of the genetic diversity of Mycobacterium tuberculosis in São Paulo city, Brazil

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Espanha Lívia G

2011-07-01

Full Text Available Abstract Background Tuberculosis is a major health problem in São Paulo, Brazil, which is the most populous and one of the most cosmopolitan cities in South America. To characterize the genetic diversity of Mycobacterium tuberculosis in the population of this city, the genotyping techniques of spoligotyping and MIRU were applied to 93 isolates collected in two consecutive years from 93 different tuberculosis patients residing in São Paulo city and attending the Clemente Ferreira Institute (the reference clinic for the treatment of tuberculosis. Findings Spoligotyping generated 53 different spoligotype patterns. Fifty-one isolates (54.8% were grouped into 13 spoligotyping clusters. Seventy- two strains (77.4% showed spoligotypes described in the international databases (SpolDB4, SITVIT, and 21 (22.6% showed unidentified patterns. The most frequent spoligotype families were Latin American Mediterranean (LAM (26 isolates, followed by the T family (24 isolates and Haarlem (H (11 isolates, which together accounted for 65.4% of all the isolates. These three families represent the major genotypes found in Africa, Central America, South America and Europe. Six Spoligo-International-types (designated SITs by the database comprised 51.8% (37/72 of all the identified spoligotypes (SIT53, SIT50, SIT42, SIT60, SIT17 and SIT1. Other SITs found in this study indicated the great genetic diversity of M. tuberculosis, reflecting the remarkable ethnic diversity of São Paulo city inhabitants. The MIRU technique was more discriminatory and did not identify any genetic clusters with 100% similarity among the 93 isolates. The allelic analysis showed that MIRU loci 26, 40, 23 and 10 were the most discriminatory. When MIRU and spoligotyping techniques were combined, all isolates grouped in the 13 spoligotyping clusters were separated. Conclusions Our data indicated the genomic stability of over 50% of spoligotypes identified in São Paulo and the great genetic

Comparison of antigen-specific T-cell responses of tuberculosis patients using complex or single antigens of Mycobacterium tuberculosis

DEFF Research Database (Denmark)

Mustafa, A S; Amoudy, H A; Wiker, H G

1998-01-01

We have screened peripheral blood mononuclear cells (PBMC) from tuberculosis (TB) patients for proliferative reactivity and interferon-gamma (IFN-gamma) secretion against a panel of purified recombinant (r) and natural (n) culture filtrate (rESAT-6, nMPT59, nMPT64 and nMPB70) and somatic-derived (r......GroES, rPstS, rGroEL and rDnaK) antigens of Mycobacterium tuberculosis. The responses of PBMC to these defined antigens were compared with the corresponding results obtained with complex antigens, such as whole-cell M. tuberculosis, M. tuberculosis culture filtrate (MT-CF) and cell wall antigens, as well...... as the vaccine strain, Mycobacterium bovis bacillus Calmette-Guerin (BCG). In addition, M. tuberculosis and MT-CF-induced T-cell lines were tested in the same assays against the panel of purified and complex antigens. The compiled data from PBMC and T-cell lines tested for antigen-induced proliferation and IFN...

JST Thesaurus Headwords and Synonyms: Mycobacterium tuberculosis [MeCab user dictionary for science technology term[Archive

Lifescience Database Archive (English)

Full Text Available MeCab user dictionary for science technology term Mycobacterium tuberculosis 名詞 一般 * * * * 結核...菌 ケッカクキン ケッカクキン Thesaurus2015 200906007893342100 C LS07 UNKNOWN_2 Mycobacterium tuberculosis

IS6110 restriction fragment length polymorphism of Mycobacterium tuberculosis isolated from patients with pulmonary tuberculosis in Campinas, Brazil: evidence of intercontinental distribution of strains

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Ana Lucia Roscani Calusni

2003-07-01

Full Text Available Tuberculosis (TB is a major concern in developing countries. In Brazil, few genotyping studies have been conducted to verify the number of IS6110 copies present in local prevalent strains of Mycobacterium tuberculosis, the distribution and clustering of strains. IS6110 DNA fingerprinting was performed on a sample of M. tuberculosis isolates from patients with AFB smear-positive pulmonary TB, at a hospital in Brazil. The IS6110 profiles were analyzed and compared to a M. tuberculosis database of the Houston Tuberculosis Initiative, Houston, US. Seventy-six fingerprints were obtained from 98 patients. All M. tuberculosis strains had an IS6110 copy number between 5-21 allowing for differentiation of the isolates. Human immunodeficiency virus infection was confirmed in nearly half the patients of whom data was available. Fifty-eight strains had unique patterns, while 17 strains were grouped in 7 clusters (2 to 6 strains. When compared to the HTI database, 6 strains matched isolates from El Paso, Ciudad de Juarez, Houston, and New York. Recently acquired infections were documented in 19% of cases. The community transmission of infection is intense, since some clustered strains were recovered during the four-year study period. The intercontinental dissemination of M. tuberculosis strains is suspected by demonstration of identical fingerprints in a distant country.

Molecular identification of Mycobacterium tuberculosis complex isolates from Kermanshah Province, Iran

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Roghieh Moghaddam

2016-01-01

Full Text Available Tuberculosis is one of the most important zoonotic diseases in the world. Rapid diagnosis of the disease and identification of species is extremely important for proper treatment of the disease as some species of the complex are resistant to the first-line of tuberculosis drugs. The aim of present study was molecular identification of Mycobacterium tuberculosis (MTB complex isolates from Kermanshah Province, Iran, which were submitted to the Tuberculosis Reference Laboratory at Razi Vaccine and Serum Research Institute (Tehran, Iran. To identify the genus Mycobacterium, all isolates were subjected to 16S rRNA polymerase chain reaction (PCR, and PCR-IS6110 was subsequently used to confirm that the isolates belonged to MTB complex. Finally, region of difference (RD typing was used to identify the species in the complex. The results of 16S rRNA and IS6110 PCR analysis showed the presence of 543-bp and 245-bp bands, respectively. Furthermore, 146bp, 172bp, 235bp, and 369bp at RD1, RD4, RD9, and RD12, respectively, were observed during RD typing. Thus, based on the results, all isolates were identified as MTB. It is worth mentioning that most tuberculosis cases are identified on the basis of acid-fast bacilli detection, and antibiotic therapy is immediately initiated subsequently. Moreover, it should be noted that some of these acid-fast positive cases might not be of genus Mycobacterium, and thus, the antibiotics prescribed might threaten the health of the patients. Additionally, if the identified bacilli are not within MTB complex, the drug therapy would differ. However, Mycobacterium bovis, which is a member of MTB complex and is resistant to pyrazinamide, requires exact strain identification. Based on the findings, individual isolates should be identified by RD typing methods, which could clearly discriminate the species from each other.

Distinct Spatiotemporal Dynamics of Peptidoglycan Synthesis between Mycobacterium smegmatis and Mycobacterium tuberculosis

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Helene Botella

2017-09-01

Full Text Available Peptidoglycan (PG, a polymer cross-linked by d-amino acid-containing peptides, is an essential component of the bacterial cell wall. We found that a fluorescent d-alanine analog (FDAA incorporates chiefly at one of the two poles in Mycobacterium smegmatis but that polar dominance varies as a function of the cell cycle in Mycobacterium tuberculosis: immediately after cytokinesis, FDAAs are incorporated chiefly at one of the two poles, but just before cytokinesis, FDAAs are incorporated comparably at both. These observations suggest that mycobacterial PG-synthesizing enzymes are localized in functional compartments at the poles and septum and that the capacity for PG synthesis matures at the new pole in M. tuberculosis. Deeper knowledge of the biology of mycobacterial PG synthesis may help in discovering drugs that disable previously unappreciated steps in the process.

Rapid drug susceptibility test of mycobacterium tuberculosis by bioluminescence sensor

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Lu, Bin; Xu, Shunqing; Chen, Zifei; Zhou, Yikai

2001-09-01

With the persisting increase of drug-resistant stains of M. Tuberculosis around the world, rapid and sensitive detection of antibiotic of M. Tuberculosis is becoming more and more important. In the present study, drug susceptibility of M. tuberculosis were detected by recombination mycobacteriophage combined with bioluminescence sensor. It is based on the use of recombination mycobacteriophage which can express firefly luciferase when it infects viable mycobacteria, and can effectively produce quantifiable photon. Meanwhile, in mycobacterium cells treated with active antibiotic, no light is observed. The emitted light is recorded by a bioluminscence sensor, so the result of drug-resistant test can be determined by the naked eye. 159 stains of M. tuberculosis were applied to this test on their resistant to rifampin, streptomycin and isoniazid. It is found that the agreement of this assay with Liewenstein- Jensen slat is: rifampin 95.60 percent, isoniazid 91.82 percent, streptomycin 88.68 percent, which showed that it is a fast and practical method to scene and detect drug resistant of mycobacterium stains.

Progenitor “Mycobacterium canettii” clone responsible for lymph node tuberculosis epidemic, Djibouti.

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Blouin, Yann; Cazajous, Géraldine; Dehan, Céline; Soler, Charles; Vong, Rithy; Hassan, Mohamed Osman; Hauck, Yolande; Boulais, Christian; Andriamanantena, Dina; Martinaud, Christophe; Martin, Émilie; Pourcel, Christine; Vergnaud, Gilles

2014-01-01

“Mycobacterium canettii,” an opportunistic human pathogen living in an unknown environmental reservoir, is the progenitor species from which Mycobacterium tuberculosis emerged. Since its discovery in 1969, most of the ≈70 known M. canettii strains were isolated in the Republic of Djibouti, frequently from expatriate children and adults. We show here, by whole-genome sequencing, that most strains collected from February 2010 through March 2013, and associated with 2 outbreaks of lymph node tuberculosis in children, belong to a unique epidemic clone within M. canettii. Evolution of this clone, which has been recovered regularly since 1983, may mimic the birth of M. tuberculosis. Thus, recognizing this organism and identifying its reservoir are clinically important.

Comparison of the UDP-N-Acetylmuramate:l-Alanine Ligase Enzymes from Mycobacterium tuberculosis and Mycobacterium leprae

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Mahapatra, Sebabrata; Crick, Dean C.; Brennan, Patrick J.

2000-01-01

In the peptidoglycan of Mycobacterium leprae, l-alanine of the side chain is replaced by glycine. When expressed in Escherichia coli, MurC (UDP-N-acetyl-muramate:l-alanine ligase) of M. leprae showed Km and Vmax for l-alanine and glycine similar to those of Mycobacterium tuberculosis MurC, suggesting that another explanation should be sought for the presence of glycine. PMID:11073931

First approach to molecular epidemiology of bovine tuberculosis in Colombia

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Jimena Jojoa-Jojoa

2015-12-01

Full Text Available Objective. To investigate the presence of Mycobacterium bovis and other Mycobacterium species in samples of cattle and buffalo in Colombia, to start the molecular characterization of M. bovis in the country. Material and methods. 492 samples were collected from herds identified with the presence of infected animals through the PPD, by the Group of Bovine Tuberculosis ICA Colombian Agricultural Institute in eight departments of Colombia. Lymph nodes of head, thorax and abdomen, gross lesions of tissues with tuberculosis, nasal swabs, milk, blood and fresh cheeses were included. Samples were subjected to detection of Mycobacterium bovis and other mycobacteria by conventional microbiological analysis and PCR-6110 and spoligotyping molecular assays. Results. In the samples analyzed especially in lymph nodes, Mycobacterium bovis was demonstrated with genotypes reported and not previously reported in the world, as well as M. tuberculosis in Antioquia, Cundinamarca, Boyacá and Magdalena departments. Conclusions. In Colombia there are at least 7 genotypes of M. bovis that are infected and sick cattle and buffalo from four different departments becoming serious threat to public health.

Cellular immune responses to ESAT-6 discriminate between patients with pulmonary disease due to Mycobacterium avium complex and those with pulmonary disease due to Mycobacterium tuberculosis

DEFF Research Database (Denmark)

Lein, A D; von Reyn, C F; Ravn, P

1999-01-01

ESAT-6 (for 6-kDa early secreted antigenic target) is a secreted antigen found almost exclusively in organisms of the Mycobacterium tuberculosis complex. We compared in vitro gamma interferon (IFN-gamma) responses by peripheral blood mononuclear cells to this antigen in patients with pulmonary...... disease due to either Mycobacterium avium complex (MAC) or Mycobacterium tuberculosis with those in healthy, skin test-negative, control subjects. Significant IFN-gamma responses to ESAT-6 were detected in 16 (59%) of 27 M. tuberculosis pulmonary disease patients, 0 (0%) of 8 MAC disease patients, and 0...... (0%) of 8 controls. Significant IFN-gamma responses to M. tuberculosis purified protein derivative were detected in 23 (85%) of 27 M. tuberculosis disease patients, 2 (25%) of 8 MAC disease patients, and 5 (63%) of 8 healthy controls. M. avium sensitin was recognized in 24 (89%) of 27 M. tuberculosis...

Spoligotyping of Mycobacterium tuberculosis isolates at a tertiary care hospital in India.

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Suzana, Shirly; Shanmugam, Sivakumar; Uma Devi, K R; Swarna Latha, P N; Michael, Joy S

2017-06-01

Spoligotyping is a valuable genotyping tool to study the genetic diversity and molecular epidemiology of Mycobacterium tuberculosis (M. tb). The aim of this study was to analyse different spoligotype patterns of M. tb strains isolated from patients with tuberculosis from different parts of India. A total of 163 M. tb isolates were spoligotyped between January 2014 and January 2015. About 47% (n = 77) were from patients with extrapulmonary tuberculosis; of these, 10 were MDR, and seven were Pre-XDR. Of the 86 M. tb isolates from patients with pulmonary tuberculosis, 25 were MDR, and 25 were Pre-XDR. We found 61 spoligo patterns, 128 clusters in the spoligotype data base (spoldb4 data base) with spoligo international type (SIT) number and 35 true unique isolates. The most pre-dominant spoligotype was EAI lineage (56), followed by Beijing (28), CAS (20), T(9), U(7), X(3), H(3), BOVIS_1 BCG(1) and LAM(1). Although our study identified EAI, CAS and Beijing strain lineages as pre-dominant, we also found a large number of orphan strains (20%) in our study. Beijing strains were more significantly associated with MDR TB than CAS and EAI lineages. Further studies on large sample sizes would help to clearly describe the epidemiology of M. tb in India. © 2017 John Wiley & Sons Ltd.

Mycobacterium tuberculosis bacteremia detected by the Isolator lysis-centrifugation blood culture system.

OpenAIRE

Kiehn, T E; Gold, J W; Brannon, P; Timberger, R J; Armstrong, D

1985-01-01

Mycobacterium tuberculosis was detected by the Isolator lysis-centrifugation blood culture system from the blood of a patient with tuberculosis of the breast. The organism also grew on conventional laboratory media inoculated with pleural fluid from the patient.

Targeting phenotypically tolerant Mycobacterium tuberculosis

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Gold, Ben; Nathan, Carl

2016-01-01

While the immune system is credited with averting tuberculosis in billions of individuals exposed to Mycobacterium tuberculosis, the immune system is also culpable for tempering the ability of antibiotics to deliver swift and durable cure of disease. In individuals afflicted with tuberculosis, host immunity produces diverse microenvironmental niches that support suboptimal growth, or complete growth arrest, of M. tuberculosis. The physiological state of nonreplication in bacteria is associated with phenotypic drug tolerance. Many of these host microenvironments, when modeled in vitro by carbon starvation, complete nutrient starvation, stationary phase, acidic pH, reactive nitrogen intermediates, hypoxia, biofilms, and withholding streptomycin from the streptomycin-addicted strain SS18b, render M. tuberculosis profoundly tolerant to many of the antibiotics that are given to tuberculosis patients in a clinical setting. Targeting nonreplicating persisters is anticipated to reduce the duration of antibiotic treatment and rate of post-treatment relapse. Some promising drugs to treat tuberculosis, such as rifampicin and bedaquiline, only kill nonreplicating M. tuberculosis in vitro at concentrations far greater than their minimal inhibitory concentrations against replicating bacilli. There is an urgent demand to identify which of the currently used antibiotics, and which of the molecules in academic and corporate screening collections, have potent bactericidal action on nonreplicating M. tuberculosis. With this goal, we review methods of high throughput screening to target nonreplicating M. tuberculosis and methods to progress candidate molecules. A classification based on structures and putative targets of molecules that have been reported to kill nonreplicating M. tuberculosis revealed a rich diversity in pharmacophores. However, few of these compounds were tested under conditions that would exclude the impact of adsorbed compound acting during the recovery phase of

Mycobacterium tuberculosis effectors interfering host apoptosis signaling.

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Liu, Minqiang; Li, Wu; Xiang, Xiaohong; Xie, Jianping

2015-07-01

Tuberculosis remains a serious human public health concern. The coevolution between its pathogen Mycobacterium tuberculosis and human host complicated the way to prevent and cure TB. Apoptosis plays subtle role in this interaction. The pathogen endeavors to manipulate the apoptosis via diverse effectors targeting key signaling nodes. In this paper, we summarized the effectors pathogen used to subvert the apoptosis, such as LpqH, ESAT-6/CFP-10, LAMs. The interplay between different forms of cell deaths, such as apoptosis, autophagy, necrosis, is also discussed with a focus on the modes of action of effectors, and implications for better TB control.

Molecular Epidemiology of Mycobacterium Tuberculosis Strains in ...

African Journals Online (AJOL)

Doroudchi M, Kremer K, Basiri EA, Kadivar MR,. Van Soolingen D, Ghaderi AA. IS6110‑RFLP and spoligotyping of Mycobacterium tuberculosis isolates in Iran. Scand J Infect. Dis 2000;32:663‑8. 13. Farnia P, Masjedi MR, Mirsaeidi M, Mohammadi F,. Jallaledin‑Ghanavi, Vincent V, et al. Prevalence of Haarlem I and Beijing ...

Horizontal acquisition of a hypoxia-responsive molybdenum cofactor biosynthesis pathway contributed to Mycobacterium tuberculosis pathoadaptation.

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Levillain, Florence; Poquet, Yannick; Mallet, Ludovic; Mazères, Serge; Marceau, Michael; Brosch, Roland; Bange, Franz-Christoph; Supply, Philip; Magalon, Axel; Neyrolles, Olivier

2017-11-01

The unique ability of the tuberculosis (TB) bacillus, Mycobacterium tuberculosis, to persist for long periods of time in lung hypoxic lesions chiefly contributes to the global burden of latent TB. We and others previously reported that the M. tuberculosis ancestor underwent massive episodes of horizontal gene transfer (HGT), mostly from environmental species. Here, we sought to explore whether such ancient HGT played a part in M. tuberculosis evolution towards pathogenicity. We were interested by a HGT-acquired M. tuberculosis-specific gene set, namely moaA1-D1, which is involved in the biosynthesis of the molybdenum cofactor. Horizontal acquisition of this gene set was striking because homologues of these moa genes are present all across the Mycobacterium genus, including in M. tuberculosis. Here, we discovered that, unlike their paralogues, the moaA1-D1 genes are strongly induced under hypoxia. In vitro, a M. tuberculosis moaA1-D1-null mutant has an impaired ability to respire nitrate, to enter dormancy and to survive in oxygen-limiting conditions. Conversely, heterologous expression of moaA1-D1 in the phylogenetically closest non-TB mycobacterium, Mycobacterium kansasii, which lacks these genes, improves its capacity to respire nitrate and grants it with a marked ability to survive oxygen depletion. In vivo, the M. tuberculosis moaA1-D1-null mutant shows impaired survival in hypoxic granulomas in C3HeB/FeJ mice, but not in normoxic lesions in C57BL/6 animals. Collectively, our results identify a novel pathway required for M. tuberculosis resistance to host-imposed stress, namely hypoxia, and provide evidence that ancient HGT bolstered M. tuberculosis evolution from an environmental species towards a pervasive human-adapted pathogen.

"Pseudo-Beijing": evidence for convergent evolution in the direct repeat region of Mycobacterium tuberculosis.

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Lukas Fenner

Full Text Available Mycobacterium tuberculosis has a global population structure consisting of six main phylogenetic lineages associated with specific geographic regions and human populations. One particular M. tuberculosis genotype known as "Beijing" has repeatedly been associated with drug resistance and has been emerging in some parts of the world. "Beijing" strains are traditionally defined based on a characteristic spoligotyping pattern. We used three alternative genotyping techniques to revisit the phylogenetic classification of M. tuberculosis complex (MTBC strains exhibiting the typical "Beijing" spoligotyping pattern.MTBC strains were obtained from an ongoing molecular epidemiological study in Switzerland and Nepal. MTBC genotyping was performed based on SNPs, genomic deletions, and 24-loci MIRU-VNTR. We identified three MTBC strains from patients originating from Tibet, Portugal and Nepal which exhibited a spoligotyping patterns identical to the classical Beijing signature. However, based on three alternative molecular markers, these strains were assigned to Lineage 3 (also known as Delhi/CAS rather than to Lineage 2 (also known as East-Asian lineage. Sequencing of the RD207 in one of these strains showed that the deletion responsible for this "Pseudo-Beijing" spoligotype was about 1,000 base pairs smaller than the usual deletion of RD207 in classical "Beijing" strains, which is consistent with an evolutionarily independent deletion event in the direct repeat (DR region of MTBC.We provide an example of convergent evolution in the DR locus of MTBC, and highlight the limitation of using spoligotypes for strain classification. Our results indicate that a proportion of "Beijing" strains may have been misclassified in the past. Markers that are more phylogenetically robust should be used when exploring strain-specific differences in experimental or clinical phenotypes.

Mutations in DNA repair genes are associated with the Haarlem lineage of Mycobacterium tuberculosis independently of their antibiotic resistance.

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Olano, Juanita; López, Beatriz; Reyes, Alejandro; Lemos, María del Pilar; Correa, Nidia; Del Portillo, Patricia; Barrera, Lucia; Robledo, Jaime; Ritacco, Viviana; Zambrano, María Mercedes

2007-11-01

The analysis of the DNA repair genes ogt and ung was carried out in 117 Mycobacterium tuberculosis clinical isolates from Argentina and Colombia in order to explore correlation between mutations in these genes and multi-drug resistance. With the exception of two Beijing family isolates, the rest of the strains harbored either two wild-type or two mutant alleles with identical single nucleotide polymorphisms (SNPs) in each gene (ogt44 and ung501). These ogt44 and ung501 mutations were not associated with multi-drug resistance and occurred simultaneously in circulating Haarlem genotype M. tuberculosis strains. We therefore propose the use of these markers as tools in phylogenetic and epidemiologic studies.

Molecular epidemiology of Mycobacterium tuberculosis in aboriginal peoples of Taiwan, 2006-2011.

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Chen, Yih-Yuan; Chang, Jia-Ru; Huang, Wei-Feng; Kuo, Shu-Chen; Yeh, Jun-Jun; Lee, Jen-Jyh; Jang, Chang-Sheng; Sun, Jun-Ren; Chiueh, Tzong-Shi; Su, Ih-Jen; Dou, Horng-Yunn

2014-04-01

Previous research revealed a 6-fold higher incidence of tuberculosis (TB) amongst aborigines compared to Han Chinese in Taiwan. To investigate the reasons for this disparity, we genotyped Mycobacterium tuberculosis (MTB) strains obtained from members of different aboriginal tribes in different geographical regions of Taiwan by using molecular methods. In total, 177 isolates of MTB collected from patients at four hospitals in Taiwan from January 2006 to December 2011 were analysed by spoligotyping, mycobacterial interspersed repetitive unit-variable number tandem-repeat (MIRU-VNTR) typing. The most prevalent strains in the eastern and central regions of Taiwan were Beijing (45.7% in eastern) and Haarlem (39.1% in eastern, 37.1% in central) lineages, whereas in southern regions the most prevalent strains were EAI (47.7%) and Haarlem (20.5%) lineages. The high prevalence of EAI in southern Taiwan aborigines may be closely associated with Austronesian culture. This study provides a first overview of the M. tuberculosis strains circulating in aboriginal populations in Taiwan. The high prevalences of certain MTB lineages within aboriginal sub-populations suggest that transmission of MTB may have been restricted to close contacts. Copyright © 2013 The British Infection Association. Published by Elsevier Ltd. All rights reserved.

Multiplexed Quantitation of Intraphagocyte Mycobacterium tuberculosis Secreted Protein Effectors

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Fadel Sayes

2018-04-01

Full Text Available Summary: The pathogenic potential of Mycobacterium tuberculosis largely depends on ESX secretion systems exporting members of the multigenic Esx, Esp, and PE/PPE protein families. To study the secretion and regulation patterns of these proteins while circumventing immune cross-reactions due to their extensive sequence homologies, we developed an approach that relies on the recognition of their MHC class II epitopes by highly discriminative T cell receptors (TCRs of a panel of T cell hybridomas. The latter were engineered so that each expresses a unique fluorescent reporter linked to specific antigen recognition. The resulting polychromatic and multiplexed imaging assay enabled us to measure the secretion of mycobacterial effectors inside infected host cells. We applied this novel technology to a large panel of mutants, clinical isolates, and host-cell types to explore the host-mycobacteria interplay and its impact on the intracellular bacterial secretome, which also revealed the unexpected capacity of phagocytes from lung granuloma to present mycobacterial antigens via MHC class II. : Sayes et al. develop an approach to express distinct fluorescent reporters that is based on the recognition of specific Mycobacterium tuberculosis MHC class II epitopes by highly discriminative T cell hybridomas. This multiplexed technology allows the study of secretion, subcellular location, and regulation patterns of these instrumental protein members. Keywords: mycobacterium tuberculosis, type VII secretion systems, intracellular bacteria, T-cell hybridomas, mycobacterial virulence factors, bacterial antigen presentation, lentiviral vectors, reporter T cells, in vivo antigen presentation, protein localization

Virulence factors of the Mycobacterium tuberculosis complex

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Forrellad, Marina A.; Klepp, Laura I.; Gioffré, Andrea; Sabio y García, Julia; Morbidoni, Hector R.; Santangelo, María de la Paz; Cataldi, Angel A.; Bigi, Fabiana

2013-01-01

The Mycobacterium tuberculosis complex (MTBC) consists of closely related species that cause tuberculosis in both humans and animals. This illness, still today, remains to be one of the leading causes of morbidity and mortality throughout the world. The mycobacteria enter the host by air, and, once in the lungs, are phagocytated by macrophages. This may lead to the rapid elimination of the bacillus or to the triggering of an active tuberculosis infection. A large number of different virulence factors have evolved in MTBC members as a response to the host immune reaction. The aim of this review is to describe the bacterial genes/proteins that are essential for the virulence of MTBC species, and that have been demonstrated in an in vivo model of infection. Knowledge of MTBC virulence factors is essential for the development of new vaccines and drugs to help manage the disease toward an increasingly more tuberculosis-free world. PMID:23076359

Phenotypic assays for Mycobacterium tuberculosis infection.

Science.gov (United States)

Song, Ok-Ryul; Deboosere, Nathalie; Delorme, Vincent; Queval, Christophe J; Deloison, Gaspard; Werkmeister, Elisabeth; Lafont, Frank; Baulard, Alain; Iantomasi, Raffaella; Brodin, Priscille

2017-10-01

Tuberculosis (TB) is still a major global threat, killing more than one million persons each year. With the constant increase of Mycobacterium tuberculosis strains resistant to first- and second-line drugs, there is an urgent need for the development of new drugs to control the propagation of TB. Although screenings of small molecules on axenic M. tuberculosis cultures were successful for the identification of novel putative anti-TB drugs, new drugs in the development pipeline remains scarce. Host-directed therapy may represent an alternative for drug development against TB. Indeed, M. tuberculosis has multiple specific interactions within host phagocytes, which may be targeted by small molecules. In order to enable drug discovery strategies against microbes residing within host macrophages, we developed multiple fluorescence-based HT/CS phenotypic assays monitoring the intracellular replication of M. tuberculosis as well as its intracellular trafficking. What we propose here is a population-based, multi-parametric analysis pipeline that can be used to monitor the intracellular fate of M. tuberculosis and the dynamics of cellular events such as phagosomal maturation (acidification and permeabilization), zinc poisoning system or lipid body accumulation. Such analysis allows the quantification of biological events considering the host-pathogen interplay and may thus be derived to other intracellular pathogens. © 2017 International Society for Advancement of Cytometry. © 2017 International Society for Advancement of Cytometry.

Genetic diversity of drug and multidrug-resistant Mycobacterium tuberculosis circulating in Veracruz, Mexico

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Munro-Rojas, Daniela; Fernandez-Morales, Esdras; Zarrabal-Meza, José; Martínez-Cazares, Ma. Teresa; Parissi-Crivelli, Aurora; Fuentes-Domínguez, Javier; Séraphin, Marie Nancy; Lauzardo, Michael; González-y-Merchand, Jorge Alberto; Rivera-Gutierrez, Sandra

2018-01-01

Background Mexico is one of the most important contributors of drug and multidrug-resistant tuberculosis in Latin America; however, knowledge of the genetic diversity of drug-resistant tuberculosis isolates is limited. Methods In this study, the genetic structure of 112 Mycobacterium tuberculosis strains from the southeastern Mexico was determined by spoligotyping and 24-loci MIRU-VNTRs. Findings The results show eight major lineages, the most of which was T1 (24%), followed by LAM (16%) and H (15%). A total of 29 (25%) isolates were identified as orphan. The most abundant SITs were SIT53/T1 and SIT42/LAM9 with 10 isolates each and SIT50/H3 with eight isolates. Fifty-two spoligotype patterns, twenty-seven clusters and ten clonal complexes were observed, demonstrating an important genetic diversity of drug and multidrug-resistant tuberculosis isolates in circulation and transmission level of these aggravated forms of tuberculosis. Being defined as orphan or as part of an orphan cluster, was a risk factor for multidrug resistant-tuberculosis (OR 2.5, IC 1.05–5.86 and OR 3.3, IC 1–11.03, respectively). Multiple correspondence analyses showed association of some clusters and SITs with specific geographical locations. Conclusions Our study provides one of the most detailed description of the genetic structure of drug and multidrug-resistant tuberculosis strains in southeast Mexico, establishing for the first time a baseline of the genotypes observed in resistant isolates circulating, however further studies are required to better elucidate the genetic structure of tuberculosis in region and the factors that could be participating in their dispersion. PMID:29543819

Genome sequencing and annotation of multidrug resistant Mycobacterium tuberculosis (MDR-TB PR10 strain

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Mohd Zakihalani A. Halim

2016-03-01

Full Text Available Here, we report the draft genome sequence and annotation of a multidrug resistant Mycobacterium tuberculosis strain PR10 (MDR-TB PR10 isolated from a patient diagnosed with tuberculosis. The size of the draft genome MDR-TB PR10 is 4.34 Mbp with 65.6% of G + C content and consists of 4637 predicted genes. The determinants were categorized by RAST into 400 subsystems with 4286 coding sequences and 50 RNAs. The whole genome shotgun project has been deposited at DDBJ/EMBL/GenBank under the accession number CP010968. Keywords: Mycobacterium tuberculosis, Genome, MDR, Extrapulmonary

Molecular epidemiology and evolutionary genetics of Mycobacterium tuberculosis in Taipei

OpenAIRE

Su Ih-Jen; Lee Shi-Yi; Tsai Wen-Shing; Sun Jun-Ren; Chang Jia-Ru; Lin Chih-Wei; Tseng Fan-Chen; Dou Horng-Yunn; Lu Jang-Jih

2008-01-01

Abstract Background The control of tuberculosis in densely populated cities is complicated by close human-to-human contacts and potential transmission of pathogens from multiple sources. We conducted a molecular epidemiologic analysis of 356 Mycobacterium tuberculosis (MTB) isolates from patients presenting pulmonary tuberculosis in metropolitan Taipei. Classical antibiogram studies and genetic characterization, using mycobacterial interspersed repetitive-unit-variable-number tandem-repeat (M...

Mycobacterium Tuberculosis Pyomyositis in an Infant

Science.gov (United States)

Malik, ZA; Shehab, M

2013-01-01

Mycobacterium tuberculosis is endemic to many parts of the world. It may have variable clinical presentations, especially in the pediatric age group. Presented here is the case of a 9-month old infant who was referred for infectious disease opinion when his thigh induration failed to improve after surgical drainage and a course of oral antibiotic therapy. Mycobacterial PCR on the operative sample fluid was found to be positive; and mycobacterial culture grew M. tuberculosis. He received 9 months of treatment with anti-TB medications, with excellent results and complete recovery. This is the first report of TB pyomyositis in an infant; and highlights the need to have a high index of suspicion for unusual organisms when conventional therapy fails to demonstrate expected results. PMID:23919207

Mechanish of dTTP Inhibition of the Bifunctional dCTP Deaminase:dUTPase Encoded by Mycobacterium tuberculosis

DEFF Research Database (Denmark)

Helt, Signe Smedegaard; Thymark, Majbritt; Harris, Pernille

2008-01-01

Recombinant deoxycytidine triphosphate (dCTP) deaminase from Mycobacterium tuberculosis was produced in Escherichia coli and purified. The enzyme proved to be a bifunctional dCTP deaminase:deoxyuridine triphosphatase. As such, the M. tuberculosis enzyme is the second bifunctional enzyme to be cha......Recombinant deoxycytidine triphosphate (dCTP) deaminase from Mycobacterium tuberculosis was produced in Escherichia coli and purified. The enzyme proved to be a bifunctional dCTP deaminase:deoxyuridine triphosphatase. As such, the M. tuberculosis enzyme is the second bifunctional enzyme...

Phylogenetic reconstruction of Mycobacterium tuberculosis within four settings of the Caribbean region: tree comparative analyse and first appraisal on their phylogeography.

Science.gov (United States)

Duchêne, Véronique; Ferdinand, Séverine; Filliol, Ingrid; Guégan, Jean François; Rastogi, Nalin; Sola, Christophe

2004-03-01

In order to compare phylogenetic methods and to reconstruct the evolutionary history of the tubercle bacilli, a set of macro-array-based genotyping data of Mycobacterium tuberculosis clinical isolates (called spoligotyping for spacer oligonucleotide typing, which assays the variability of the Direct Repeat -DR- locus), was analyzed in four settings of the Caribbean region (Guadeloupe, Martinique, Cuba and Haiti). A set of 47 alleles, split into 26 shared and 21 unique alleles) representative of 321 individual M. tuberculosis clinical isolates from patients residing in the above regions was studied. The following methods (and software in brackets) were investigated: numerical taxonomy distance methods (TAXOTRON), maximum parsimony procedure (PAUP), median-joining networks (NETWORK), and nested clade analysis (GEODIS). Results using these methods were analyzed, compared and discussed. The latter method (GEODIS) was investigated in detail by introducing geographical data together with genetic variability results to detect a link between population structure and population history, and to test the null hypothesis of no association between geography and genotypes. Irrespective of the methods used, our findings demonstrate that a core structure of four families (or clades) of M. tuberculosis strains is highly prevalent within the islands studied, indirectly reflecting passed colonization history of these different settings. Specificity of M. tuberculosis genotypes in each of the islands is discussed in the light of their respective colonial and contemporary histories.

Genetic characterization of Mycobacterium tuberculosis in the West Bank, Palestinian Territories

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Ereqat Suheir

2012-06-01

Full Text Available Abstract Background The World Health Organization (WHO declared human tuberculosis (TB a global health emergency and launched the “Global Plan to Stop Tuberculosis” which aims to save a million lives by 2015. Global control of TB is increasingly dependent on rapid and accurate genetic typing of species of the Mycobacterium tuberculosis (MTB complex including M. tuberculosis. The aim of this study was to identify and genetically characterize the MTB isolates circulating in the West Bank, Palestinian Territories. Genotyping of the MTB isolates from patients with pulmonary TB was carried out using two molecular genetic techniques, spoligotyping and mycobacterial interspersed repetitive units-variable number of tandem repeat (MIRU-VNTR supported by analysis of the MTB specific deletion 1 (TbD1. Findings A total of 17 MTB patterns were obtained from the 31 clinical isolates analyzed by spoligotyping; corresponding to 2 orphans and 15 shared-types (SITs. Fourteen SITs matched a preexisting shared-type in the SITVIT2 database, whereas a single shared-type SIT3348 was newly created. The most common spoligotyping profile was SIT53 (T1 variant, identified in 35.5 % of the TB cases studied. Genetic characterization of 22 clinical isolates via the 15 loci MIRU-VNTR typing distinguished 19 patterns. The 15-loci MIT144 and MIT145 were newly created within this study. Both methods determined the present of M. bovis strains among the isolates. Conclusions Significant diversity among the MTB isolates circulating in the West Bank was identified with SIT53-T1 genotype being the most frequent strain. Our results are used as reference database of the strains circulating in our region and may facilitate the implementation of an efficient TB control program.

Evaluation of rapid immuno chromatographic assay kit using monoclonal mpt64 antibodies for identification of mycobacterium tuberculosis complex

International Nuclear Information System (INIS)

Satti, L.; Ikram, A.; Malik, N.

2010-01-01

To evaluate the performance of rapid immuno chromatographic kit MPT64 Ag for the identification of Mycobacterium tuberculosis complex from various Mycobacterium tuberculosis culture positive specimens. Department of Microbiology, Armed Forces Institute of Pathology Rawalpindi, from August 2008 through March 2009. Eighty four Mycobacterium tuberculosis positive cultures on I BACTEC 460 and MGIT 960, one ATCC 25177 MTB strain, three institutional control MTB strains, two institutional control MOTT strains and 20 different bacterial isolates were tested. Tests were performed according to the instructional manual. Out of total 84 tested samples, MPT64 showed positive result in 80 cultures. Only four positive cultures did not display any band on MPT64 kit. These four strains were reconfirmed as Mycobacterium tuberculosis by PCR method. MOTT control strains and all the 20 bacterial isolates were negative for band. The sensitivity and specificity of ICT assay in our study was 95.2% and 100% respectively. Rapid MPT64 Kit is a good diagnostic tool to differentiate between Mycobacterium tuberculosis complex and MOTT with 100% specificity. The technique is simple and can provide prompt information to the clinicians to initiate early and appropriate antituberculosis therapy. (author)

In vitro efficacy of ethionamide and clarithromycin in mycobacterium tuberculosis isolates

International Nuclear Information System (INIS)

Satti, M.S.; Abbasi, S.; Rafi, S.; Butt, T.; Karamat, K.A.

2010-01-01

To determine the sensitivity of clinical isolates of Mycobacterium tuberculosis isolates against ethionamide, and clarithromycin. Department of Microbiology, Armed Forces institute of Pathology (AFIP) Rawalpindi from June 2003 to June 2004. Materials and Methods: All routine clinical samples received for acid fast bacilli (AFB) culture and yielding positive growth on Lowenstien Jensen medium and Bactec 460 were include in the study. The isolates were from sputum (n=70), bronchioalveolar lavage (n=10), fine needle aspiration (n=6), lymph nodes (n=7), pleural fluid (n=4), endometrium (n=3). After the identification of M. tuberculosis (MTB) sensitivity was performed against first-line antituberculosis drugs. Then susceptibility of M. tuberculosis isolates against ethionamide and clarithromycih was performed on LJ medium. Mycobacterium H37Rv was used as control strain. Results were interpreted using resistance ratio method. Out of 100 M. tuberculosis isolates, sensitivity to ethionamide was 93% and 9% to clarithromycih. Clarithromycin when used alone is ineffective as antituberculosis drug but its efficacy in combination needs to be tested. However ethionamide may be used as an alternative antituberculosis drug. (author)

Identification and characterization of the ESAT-6 homologue of Mycobacterium leprae and T-cell cross-reactivity with Mycobacterium tuberculosis

NARCIS (Netherlands)

Geluk, Annemieke; van Meijgaarden, Krista E.; Franken, Kees L. M. C.; Subronto, Yanri W.; Wieles, Brigitte; Arend, Sandra M.; Sampaio, Elizabeth P.; de Boer, Tjitske; Faber, William R.; Naafs, Ben; Ottenhoff, Tom H. M.

2002-01-01

In this paper we describe identification and characterization of Mycobacterium leprae ESAT-6 (L-ESAT-6), the homologue of M. tuberculosis ESAT-6 (T-ESAT-6). T-ESAT-6 is expressed by all pathogenic strains belonging to the M. tuberculosis complex but is absent from virtually all other mycobacterial

Marked microevolution of a unique Mycobacterium tuberculosis strain in 17 years of ongoing transmission in a high risk population.

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Carolina Mehaffy

Full Text Available The transmission and persistence of Mycobacterium tuberculosis within high risk populations is a threat to tuberculosis (TB control. In the current study, we used whole genome sequencing (WGS to decipher the transmission dynamics and microevolution of M. tuberculosis ON-A, an endemic strain responsible for an ongoing outbreak of TB in an urban homeless/under-housed population. Sixty-one M. tuberculosis isolates representing 57 TB cases from 1997 to 2013 were subjected to WGS. Sequencing data was integrated with available epidemiological information and analyzed to determine how the M. tuberculosis ON-A strain has evolved during almost two decades of active transmission. WGS offers higher discriminatory power than traditional genotyping techniques, dividing the M. tuberculosis ON-A strain into 6 sub-clusters, each defined by unique single nucleotide polymorphism profiles. One sub-cluster, designated ON-ANM (Natural Mutant; 26 isolates from 24 cases was also defined by a large, 15 kb genomic deletion. WGS analysis reveals the existence of multiple transmission chains within the same population/setting. Our results help validate the utility of WGS as a powerful tool for identifying genomic changes and adaptation of M. tuberculosis.

Characterisation of pks15/1 in clinical isolates of Mycobacterium tuberculosis from Mexico

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Roberto Zenteno-Cuevas

2013-09-01

Full Text Available Tuberculosis (TB is an infectocontagious respiratory disease caused by members of the Mycobacterium tuberculosis complex. A 7 base pair (bp deletion in the locus polyketide synthase (pks15/1 is described as polymorphic among members of the M. tuberculosis complex, enabling the identification of Euro-American, Indo-Oceanic and Asian lineages. The aim of this study was to characterise this locus in TB isolates from Mexico. One hundred twenty clinical isolates were recovered from the states of Veracruz and Estado de Mexico. We determined the nucleotide sequence of a ± 400 bp fragment of the locus pks15/1, while genotypic characterisation was performed by spoligotyping. One hundred and fifty isolates contained the 7 bp deletion, while five had the wild type locus. Lineages X (22%, LAM (18% and T (17% were the most frequent; only three (2% of the isolates were identified as Beijing and two (1% EAI-Manila. The wild type pks15/1 locus was observed in all Asian lineage isolates tested. Our results confirm the utility of locus pks15/1 as a molecular marker for identifying Asian lineages of the M. tuberculosis complex. This marker could be of great value in the epidemiological surveillance of TB, especially in countries like Mexico, where the prevalence of such lineages is unknown.

Associations between Mycobacterium tuberculosis Strains and Phenotypes

Science.gov (United States)

Brown, Timothy; Nikolayevskyy, Vladyslav; Velji, Preya

2010-01-01

To inform development of tuberculosis (TB) control strategies, we characterized a total of 2,261 Mycobacterium tuberculosis complex isolates by using multiple phenotypic and molecular markers, including polymorphisms in repetitive sequences (spoligotyping and variable-number tandem repeats [VNTRs]) and large sequence and single-nucleotide polymorphisms. The Beijing family was strongly associated with multidrug resistance (p = 0.0001), and VNTR allelic variants showed strong associations with spoligotyping families: >5 copies at exact tandem repeat (ETR) A, >2 at mycobacterial interspersed repetitive unit 24, and >3 at ETR-B associated with the East African–Indian and M. bovis strains. All M. tuberculosis isolates were differentiated into 4 major lineages, and a maximum parsimony tree was constructed suggesting a more complex phylogeny for M. africanum. These findings can be used as a model of pathogen global diversity. PMID:20113558

Study on drug resistance of mycobacterium tuberculosis in patients with pulmonary tuberculosis by drug resistance gene detecting

International Nuclear Information System (INIS)

Wang Wei; Li Hongmin; Wu Xueqiong; Wang Ansheng; Ye Yixiu; Wang Zhongyuan; Liu Jinwei; Chen Hongbing; Lin Minggui; Wang Jinhe; Li Sumei; Jiang Ping; Feng Bai; Chen Dongjing

2004-01-01

To investigate drug resistance of mycobacterium tuberculosis in different age group, compare detecting effect of two methods and evaluate their the clinical application value, all of the strains of mycobacterium tuberculosis were tested for resistance to RFP, INH SM PZA and EMB by the absolute concentration method on Lowenstein-Jensen medium and the mutation of the rpoB, katG, rpsL, pncA and embB resistance genes in M. tuberculosis was tested by PCR-SSCP. In youth, middle and old age group, the rate of acquired drug resistance was 89.2%, 85.3% and 67.6% respectively, the gene mutation rate was 76.2%, 81.3% and 63.2% respectively. The rate of acquired drug resistance and multiple drug resistance in youth group was much higher than those in other groups. The gene mutation was correlated with drug resistance level of mycobacterium tuberculosis. The gene mutation rate was higher in strains isolated from high concentration resistance than those in strains isolated from low concentration resistance. The more irregular treatment was longer, the rate of drug resistance was higher. Acquired drug resistance varies in different age group. It suggested that surveillance of drug resistence in different age group should be taken seriously, especially in youth group. PCR - SSCP is a sensitive and specific method for rapid detecting rpoB, katG, rpsL, pncA and embB genes mutations of MTB. (authors)

A robust SNP barcode for typing Mycobacterium tuberculosis complex strains

KAUST Repository

Coll, Francesc; McNerney, Ruth; Guerra-Assunç ã o, José Afonso; Glynn, Judith R.; Perdigã o, Joã o; Viveiros, Miguel; Portugal, Isabel; Pain, Arnab; Martin, Nigel; Clark, Taane G.

2014-01-01

Strain-specific genomic diversity in the Mycobacterium tuberculosis complex (MTBC) is an important factor in pathogenesis that may affect virulence, transmissibility, host response and emergence of drug resistance. Several systems have been proposed

T-cell recognition of Mycobacterium tuberculosis culture filtrate fractions in tuberculosis patients and their household contacts

DEFF Research Database (Denmark)

Demissie, A; Ravn, P; Olobo, J

1999-01-01

We examined the immune responses of patients with active pulmonary tuberculosis (TB) and their healthy household contacts to short-term culture filtrate (ST-CF) of Mycobacterium tuberculosis or molecular mass fractions derived from it. Our goal was to identify fractions strongly recognized......, to secreted mycobacterial antigens is suggestive of an early stage of infection by M. tuberculosis, which could in time result in overt disease or containment of the infection. This possibility is currently being investigated by follow-up studies of the household contacts....

Genotypic diversity of multidrug-, quinolone- and extensively drug-resistant Mycobacterium tuberculosis isolates in Thailand.

Science.gov (United States)

Disratthakit, Areeya; Meada, Shinji; Prammananan, Therdsak; Thaipisuttikul, Iyarit; Doi, Norio; Chaiprasert, Angkana

2015-06-01

Drug-resistant tuberculosis (TB), which includes multidrug-resistant (MDR-TB), quinolone-resistant (QR-TB) and extensively drug-resistant tuberculosis (XDR-TB), is a serious threat to TB control. We aimed to characterize the genotypic diversity of drug-resistant TB clinical isolates collected in Thailand to establish whether the emergence of drug-resistant TB is attributable to transmitted resistance or acquired resistance. We constructed the first molecular phylogeny of MDR-TB (n=95), QR-TB (n=69) and XDR-TB (n=28) in Thailand based on spoligotyping and proposed 24-locus multilocus variable-number of tandem repeat analysis (MLVA). Clustering analysis was performed using the unweighted pair group method with arithmetic mean. Spoligotyping identified the Beijing strain (SIT1) as the most predominant genotype (n=139; 72.4%). The discriminatory power of 0.9235 Hunter-Gaston Discriminatory Index (HGDI) with the 15-locus variable-number tandem repeats of mycobacterial interspersed repetitive units typing was improved to a 0.9574 HGDI with proposed 24-locus MLVA, thereby resulting in the subdivision of a large cluster of Beijing strains (SIT1) into 17 subclusters. We identified the spread of drug-resistant TB clones caused by three different MLVA types in the Beijing strain (SIT1) and a specific clone of XDR-TB caused by a rare genotype, the Manu-ancestor strain (SIT523). Overall, 49.5% of all isolates were clustered. These findings suggest that a remarkable transmission of drug-resistant TB occurred in Thailand. The remaining 50% of drug-resistant TB isolates were unique genotypes, which may have arisen from the individual acquisition of drug resistance. Our results suggest that transmitted and acquired resistance have played an equal role in the emergence of drug-resistant TB. Further characterization of whole genome sequences of clonal strains could help to elucidate the mycobacterial genetic factors relevant for drug resistance, transmissibility and virulence

Protective and therapeutic efficacy of Mycobacterium smegmatis expressing HBHA-hIL12 fusion protein against Mycobacterium tuberculosis in mice.

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Shanmin Zhao

Full Text Available Tuberculosis (TB remains a major worldwide health problem. The only vaccine against TB, Mycobacterium bovis Bacille Calmette-Guerin (BCG, has demonstrated relatively low efficacy and does not provide satisfactory protection against the disease. More efficient vaccines and improved therapies are urgently needed to decrease the worldwide spread and burden of TB, and use of a viable, metabolizing mycobacteria vaccine may be a promising strategy against the disease. Here, we constructed a recombinant Mycobacterium smegmatis (rMS strain expressing a fusion protein of heparin-binding hemagglutinin (HBHA and human interleukin 12 (hIL-12. Immune responses induced by the rMS in mice and protection against Mycobacterium tuberculosis (MTB were investigated. Administration of this novel rMS enhanced Th1-type cellular responses (IFN-γ and IL-2 in mice and reduced bacterial burden in lungs as well as that achieved by BCG vaccination. Meanwhile, the bacteria load in M. tuberculosis infected mice treated with the rMS vaccine also was significantly reduced. In conclusion, the rMS strain expressing the HBHA and human IL-12 fusion protein enhanced immunogencity by improving the Th1-type response against TB, and the protective effect was equivalent to that of the conventional BCG vaccine in mice. Furthermore, it could decrease bacterial load and alleviate histopathological damage in lungs of M. tuberculosis infected mice.

Deciphering the biology of Mycobacterium tuberculosis from thecomplete genome sequence

DEFF Research Database (Denmark)

Cole, S.T.; Krogh, Anders Stærmose

1998-01-01

Countless millions of people have died from tuberculosis, a chronic infectious disease caused by the tubercle bacillus. The complete genome sequence of the best-characterized strain of Mycobacterium tuberculosis, H37Rv, has been determined and analysed in order to improve our understanding....... tuberculosis differs radically from other bacteria in that a very large portion of its coding capacity is devoted to the production of enzymes involved in lipogenesis and lipolysis, and to two new families of glycine-rich proteins with a repetitive structure that may represent a source of antigenic variation....

The transmission of Mycobacterium tuberculosis in high burden settings

NARCIS (Netherlands)

Yates, Tom A.; Khan, Palwasha Y.; Knight, Gwenan M.; Taylor, Jonathon G.; McHugh, Timothy D.; Lipman, Marc; White, Richard G.; Cohen, Ted; Cobelens, Frank G.; Wood, Robin; Moore, David A. J.; Abubakar, Ibrahim

2016-01-01

Unacceptable levels of Mycobacterium tuberculosis transmission are noted in high burden settings and a renewed focus on reducing person-to-person transmission in these communities is needed. We review recent developments in the understanding of airborne transmission. We outline approaches to measure

Uracil excision repair in Mycobacterium tuberculosis cell-free extracts.

Science.gov (United States)

Kumar, Pradeep; Bharti, Sanjay Kumar; Varshney, Umesh

2011-05-01

Uracil excision repair is ubiquitous in all domains of life and initiated by uracil DNA glycosylases (UDGs) which excise the promutagenic base, uracil, from DNA to leave behind an abasic site (AP-site). Repair of the resulting AP-sites requires an AP-endonuclease, a DNA polymerase, and a DNA ligase whose combined activities result in either short-patch or long-patch repair. Mycobacterium tuberculosis, the causative agent of tuberculosis, has an increased risk of accumulating uracils because of its G + C-rich genome, and its niche inside host macrophages where it is exposed to reactive nitrogen and oxygen species, two major causes of cytosine deamination (to uracil) in DNA. In vitro assays to study DNA repair in this important human pathogen are limited. To study uracil excision repair in mycobacteria, we have established assay conditions using cell-free extracts of M. tuberculosis and M. smegmatis (a fast-growing mycobacterium) and oligomer or plasmid DNA substrates. We show that in mycobacteria, uracil excision repair is completed primarily via long-patch repair. In addition, we show that M. tuberculosis UdgB, a newly characterized family 5 UDG, substitutes for the highly conserved family 1 UDG, Ung, thereby suggesting that UdgB might function as backup enzyme for uracil excision repair in mycobacteria. Copyright © 2011 Elsevier Ltd. All rights reserved.

Mycobacterium tuberculosis population in northwestern Russia: an update from Russian-EU/Latvian border region.

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Igor Mokrousov

Full Text Available This study aimed to characterize the population structure of Mycobacterium tuberculosis in Pskov oblast in northwestern Russia, to view it in the geographical context, to compare drug resistance properties across major genetic families. Ninety M. tuberculosis strains from tuberculosis (TB patients, permanent residents in Pskov oblast were subjected to LAM-specific IS6110-PCR and spoligotyping, followed by comparison with SITVITWEB and MIRU-VNTRplus databases. The Beijing genotype (n = 40 was found the most prevalent followed by LAM (n = 18, T (n = 13, Haarlem (n = 10, Ural (n = 5, and Manu2 (n = 1; the family status remained unknown for 3 isolates. The high rate of Beijing genotype and prevalence of LAM family are similar to those in the other Russian settings. A feature specific for M. tuberculosis population in Pskov is a relatively higher rate of Haarlem and T types. Beijing strains were further typed with 12-MIRU (followed by comparison with proprietary global database and 3 hypervariable loci QUB-3232, VNTR-3820, VNTR-4120. The 12-MIRU typing differentiated 40 Beijing strains into 14 types (HGI = 0.82 while two largest types were M2 (223325153533 prevalent throughout former USSR and M11 (223325173533 prevalent in Russia and East Asia. The use of 3 hypervariable loci increased a discrimination of the Beijing strains (18 profiles, HGI = 0.89. Both major families Beijing and LAM had similar rate of MDR strains (62.5 and 55.6%, respectively that was significantly higher than in other strains (21.9%; P = 0.001 and 0.03, respectively. The rpoB531 mutations were more frequently found in Beijing strains while LAM drug resistant strains mainly harbored rpoB516 and inhA -15 mutations. Taken together with a high rate of multidrug resistance among Beijing strains from new TB cases (79.3% versus 44.4% in LAM, these findings suggest the critical impact of the Beijing genotype on the current situation with MDR-TB in the

Evaluation of GenoType NTM-DR Assay for Identification of Mycobacterium chimaera.

Science.gov (United States)

Mok, Simone; Rogers, Thomas R; Fitzgibbon, Margaret

2017-06-01

Identification of species within the Mycobacterium avium complex (MAC) is difficult, and most current diagnostic laboratory tests cannot distinguish between species included in the complex. Differentiation of species within the MAC is important, as Mycobacterium chimaera has recently emerged as a major cause of invasive cardiovascular infections following open heart surgery. A new commercial diagnostic assay, GenoType NTM-DR ver. 1.0, is intended to differentiate between three species within the MAC, namely, Mycobacterium avium , Mycobacterium intracellulare , and Mycobacterium chimaera In this study, we investigated an archival collection of 173 MAC isolates using 16S rRNA and 16S-23S internal transcribed spacer (ITS) gene sequencing, and GenoType NTM-DR was evaluated for identifying M. chimaera and other species belonging to the MAC. Species identification of 157/173 (91%) isolates with the GenoType NTM-DR assay was in agreement with 16S rRNA and 16S-23S ITS gene sequencing results. Misidentification occurred with 16 isolates which belonged to four species included in the MAC that are rarely encountered in clinical specimens. Despite some limitations of this assay, GenoType NTM-DR had 100% specificity for identifying M. chimaera This novel assay will enable diagnostic laboratories to differentiate species belonging to the Mycobacterium avium complex and to accurately identify M. chimaera It can produce rapid results and is also more cost efficient than gene sequencing methods. Copyright © 2017 American Society for Microbiology.

Anti-mycobacterium tuberculosis activity of polyherbal medicines used for the treatment of tuberculosis in Eastern Cape, South Africa.

Science.gov (United States)

Famewo, Elizabeth B; Clarke, Anna M; Wiid, Ian; Ngwane, Andile; van Helden, Paul; Afolayan, Anthony J

2017-09-01

The emergence of drug-resistant strains of Mycobacterium tuberculosis has become a global public health problem. Polyherbal medicines offer great hope for developing alternative drugs for the treatment of tuberculosis. To evaluate the anti-tubercular activity of polyherbal medicines used for the treatment of tuberculosis. The remedies were screened against Mycobacterium tuberculosis H37Rv using Middlebrook 7H9 media and MGIT BACTEC 960 system. They were liquid preparations from King Williams Town site A (KWTa), King Williams Town site B (KWTb), King Williams Town site C (KWTc), Hogsback first site (HBfs), Hogsback second site (HBss), Hogsback third site (HBts), East London (EL), Alice (AL) and Fort Beaufort (FB). The susceptibility testing revealed that all the remedies contain anti-tubercular activity with KWTa, KWTb, KWTc, HBfs, HBts, AL and FB exhibiting more activity at a concentration below 25 µl/ml. Furthermore, MIC values exhibited inhibitory activity with the most active remedies from KWTa, HBfs and HBts at 1.562 µg/ml. However, isoniazid showed more inhibitory activity against M. tuberculosis at 0.05 µg/ml when compare to the polyherbal remedies. This study has indicated that these remedies could be potential sources of new anti-mycobacterial agents against M. tuberculosis . However, the activity of these preparations and their active principles still require in vivo study in order to assess their future as new anti-tuberculosis agents.

Molecular Identification of Mycobacterium Tuberculosis and Analysis of Its Resistance to Rifampin in Sputa from Tuberculosis Suspected Patients

International Nuclear Information System (INIS)

Syaifudin, M.

2010-01-01

An accurate identification of different species of Mycobacterium provides to allow appropriate treatment for Mycobacterium tuberculosis infection. Beside that, drug resistance of M. tuberculosis strains to rifampin is not clearly understood in contributing to the spread of tuberculosis in Indonesia. To assess the molecular mechanism of rifampin resistance, a number of clinical specimens of M. tuberculosis were analyzed their molecular nature of a part of the rpoB gene using polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) methods. DNA's extracted from sputum samples were amplified and 32 P-labeled by PCR with the specific primers and the product was analyzed their mutation conferring resistance by MDE gel electrophoresis. Of the 70 specimens tested, 57 specimens were positive for M. tuberculosis organism only, three specimens contained a mixture of M. tuberculosis and non tuberculosis mycobacteria (NTM), and 10 specimens were negative approved by Duplex PCR. Of these sixty DNA positive samples (thus the sensitivity of PCR was 85.71%), 5 (8.3%) of them suspected to contain mutations in rpoB which were associated with rifampin resistance. Even though the frequency of mutation was low, the results from our study clearly indicate that the molecular mechanism of rifampin resistance in M. tuberculosis isolates from Indonesia involves alterations in the rpoB gene. Molecular diagnosis by PCR which is fast and easy to perform is useful for early and rapid detection of TB in sputum specimen. (author)

Prevalence of pulmonary TB and spoligotype pattern of Mycobacterium tuberculosis among TB suspects in a rural community in Southwest Ethiopia

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Deribew Amare

2012-03-01

in Ethiopia. A large scale study on the genotyping of Mycobacterium tuberculosis in Ethiopia is crucial to understand transmission dynamics, identification of drug resistant strains and design preventive strategies.

Sensitivity Pattern of Second Line Anti-Tuberculosis Drugs against Clinical Isolates of Multidrug Resistant Mycobacterium Tuberculosis

International Nuclear Information System (INIS)

Ghafoor, T.; Ikram, A.; Abbasi, S. A.; Zaman, G.; Ayyub, M.; Palomino, J. C.; Vandamme, P.; Martin, A.

2015-01-01

Objective:To determine the current sensitivity pattern of second line anti-tuberculosis drugs against clinical isolates of Multidrug Resistant Mycobacterium tuberculosis (MDR-TB). Study Design: A cross-sectional study. Place and Duration of Study: Department of Microbiology, Armed Forces Institute of Pathology (AFIP), Rawalpindi, from November 2011 to April 2013. Methodology: Samples received during the study period were processed on BACTEC MGIT 960 system for Mycobacterium tuberculosis (MTB) culture followed by first line drugs susceptibility testing of culture proven MTB isolates. On the basis of resistance to rifampicin and isoniazid, 100 clinical isolates of MDR-TB were further subjected to susceptibility testing against amikacin (AMK), capreomycin (CAP), ofloxacin (OFL) and ethionamide (ETH) as per standard BACTEC MGIT 960 instructions. Results: Out of 100 MDR-TB isolates, 62% were from male patients and 38% from female patients. 97% were sensitive to AMK, 53% to OFL, 87% to CAP; and 87% were sensitive to ETH. Conclusion: The majority of the MDR-TB isolates showed excellent sensitivity against AMK, CAP and ETH. However, sensitivity of MDR-TB isolates against fluoroquinolones like OFL was not encouraging. (author)

Epidemiological evaluation of spatiotemporal and genotypic clustering of Mycobacterium tuberculosis in Ontario, Canada.

Science.gov (United States)

Tuite, A R; Guthrie, J L; Alexander, D C; Whelan, M S; Lee, B; Lam, K; Ma, J; Fisman, D N; Jamieson, F B

2013-10-01

In Canada, tuberculosis (TB) rates are at a historic low, with the remaining risk concentrated in a few vulnerable population subgroups. To describe the epidemiology of TB in the Canadian province of Ontario and to characterise risk factors associated with transmission events, identified using genetic typing techniques. Retrospective analysis of 2186 culture-positive TB cases between August 2007 and December 2011. Temporal trends and risk of spatiotemporal and genotypic clustering were evaluated using Poisson and logistic regression models. Being in a spatiotemporal cluster was associated with Aboriginal status (odds ratio [OR] 3.63, 95% confidence interval [CI] 1.23-10.71). Cases in genotypic clusters were more likely to report homelessness as a risk factor (adjusted OR [aOR] 2.92, 95%CI 1.74-4.90) or be male (aOR 1.35, 95%CI 1.09-1.68), and were less likely to be aged ≥ 65 years (aOR 0.63, 95%CI 0.49-0.82), foreign-born (aOR 0.32, 95%CI 0.24-0.43) or Aboriginal (aOR 0.40, 95%CI 0.16-0.99). The Beijing lineage had an annual rate of increase of almost 10% (P = 0.047), and was associated with genotypic clustering (aOR 2.84, 95%CI 2.19-3.67). Genotypic data suggest that disease clusters are smaller, but far more common, than would be estimated using spatiotemporal clustering.

Interaction of Mycobacterium tuberculosis with human respiratory mucosa.

Science.gov (United States)

Middleton, A M; Chadwick, M V; Nicholson, A G; Dewar, A; Groger, R K; Brown, E J; Ratliff, T L; Wilson, R

2002-01-01

Endobronchial infection is associated with pulmonary tuberculosis in the majority of cases. We have investigated the adherence of Mycobacterium tuberculosis to the human respiratory mucosa. Organ cultures constructed with human tissue were infected with M. tuberculosis in the presence or absence of mycobacterial fibronectin attachment cell surface proteins and examined by scanning electron microscopy. M. tuberculosis adhered mainly to extracellular matrix (ECM) in areas of mucosal damage, but not to ciliated mucosa, intact extruded cells, basement membrane or collagen fibres. Bacteria also adhered to fibrous but not globular mucus and occasionally to healthy unciliated mucosa, open tight junctions and to extruded cells that had degenerated, exposing their contents. There was a significant reduction (pprotein (FAP) and M. bovis antigen 85B protein, in a concentration dependent manner. The combined effect of FAP and antigen 85B protein was significantly greater than either protein alone. Bacterial adherence to fibrous mucus was not influenced by fibronectin. We conclude that M. tuberculosis adheres to ECM in areas of mucosal damage at least in part via FAP and antigen 85B protein.

Whole genome sequencing of Mycobacterium tuberculosis SB24 isolated from Sabah, Malaysia

Directory of Open Access Journals (Sweden)

Noraini Philip

2016-09-01

Full Text Available Mycobacterium tuberculosis (M. tuberculosis is the causative agent of tuberculosis (TB that causes millions of death every year. We have sequenced the genome of M. tuberculosis isolated from cerebrospinal fluid (CSF of a patient diagnosed with tuberculous meningitis (TBM. The isolated strain was referred as M. tuberculosis SB24. Genomic DNA of the M. tuberculosis SB24 was extracted and subjected to whole genome sequencing using PacBio platform. The draft genome size of M. tuberculosis SB24 was determined to be 4,452,489 bp with a G + C content of 65.6%. The whole genome shotgun project has been deposited in NCBI SRA under the accession number SRP076503.

Mycobacterium tuberculosis whole genome sequencing and protein structure modelling provides insights into anti-tuberculosis drug resistance

KAUST Repository

Phelan, Jody; Coll, Francesc; McNerney, Ruth; Ascher, David B.; Pires, Douglas E. V.; Furnham, Nick; Coeck, Nele; Hill-Cawthorne, Grant A.; Nair, Mridul; Mallard, Kim; Ramsay, Andrew; Campino, Susana; Hibberd, Martin L.; Pain, Arnab; Rigouts, Leen; Clark, Taane G.

2016-01-01

of 144 Mycobacterium tuberculosis clinical isolates from The Special Programme for Research and Training in Tropical Diseases (TDR) collection sourced from 20 countries in four continents. A genome-wide approach was applied to 127 isolates to identify

Transmission of Mycobacterium tuberculosis Undetected by Tuberculin Skin Testing

Czech Academy of Sciences Publication Activity Database

Anderson, S. T.; Williams, A. J.; Brown, J. R.; Newton, S. M.; Šimšová, Marcela; Nicol, M. P.; Šebo, Peter; Levin, M.; Wilkinson, R. J.; Wilkinson, K. A.

2006-01-01

Roč. 173, - (2006), s. 1038-1042 ISSN 1073-449X R&D Projects: GA AV ČR IAA5020406 Institutional research plan: CEZ:AV0Z50200510 Keywords : adenylate cyclase * diagnostic tests and procedures * mycobacterium tuberculosis Subject RIV: EE - Microbiology, Virology Impact factor: 9.091, year: 2006

DNA fingerprinting of Mycobacterium tuberculosis: from phage typing to whole-genome sequencing.

Science.gov (United States)

Schürch, Anita C; van Soolingen, Dick

2012-06-01

Current typing methods for Mycobacterium tuberculosis complex evolved from simple phenotypic approaches like phage typing and drug susceptibility profiling to DNA-based strain typing methods, such as IS6110-restriction fragment length polymorphisms (RFLP) and variable number of tandem repeats (VNTR) typing. Examples of the usefulness of molecular typing are source case finding and epidemiological linkage of tuberculosis (TB) cases, international transmission of MDR/XDR-TB, the discrimination between endogenous reactivation and exogenous re-infection as a cause of relapses after curative treatment of tuberculosis, the evidence of multiple M. tuberculosis infections, and the disclosure of laboratory cross-contaminations. Simultaneously, phylogenetic analyses were developed based on single nucleotide polymorphisms (SNPs), genomic deletions usually referred to as regions of difference (RDs) and spoligotyping which served both strain typing and phylogenetic analysis. National and international initiatives that rely on the application of these typing methods have brought significant insight into the molecular epidemiology of tuberculosis. However, current DNA fingerprinting methods have important limitations. They can often not distinguish between genetically closely related strains and the turn-over of these markers is variable. Moreover, the suitability of most DNA typing methods for phylogenetic reconstruction is limited as they show a high propensity of convergent evolution or misinfer genetic distances. In order to fully explore the possibilities of genotyping in the molecular epidemiology of tuberculosis and to study the phylogeny of the causative bacteria reliably, the application of whole-genome sequencing (WGS) analysis for all M. tuberculosis isolates is the optimal, although currently still a costly solution. In the last years WGS for typing of pathogens has been explored and yielded important additional information on strain diversity in comparison to the

Lymphatic endothelial cells are a replicative niche for Mycobacterium tuberculosis

Science.gov (United States)

Lerner, Thomas R.; de Souza Carvalho-Wodarz, Cristiane; Repnik, Urska; Russell, Matthew R.G.; Borel, Sophie; Diedrich, Collin R.; Rohde, Manfred; Wainwright, Helen; Collinson, Lucy M.; Wilkinson, Robert J.; Griffiths, Gareth; Gutierrez, Maximiliano G.

2016-01-01

In extrapulmonary tuberculosis, the most common site of infection is within the lymphatic system, and there is growing recognition that lymphatic endothelial cells (LECs) are involved in immune function. Here, we identified LECs, which line the lymphatic vessels, as a niche for Mycobacterium tuberculosis in the lymph nodes of patients with tuberculosis. In cultured primary human LECs (hLECs), we determined that M. tuberculosis replicates both in the cytosol and within autophagosomes, but the bacteria failed to replicate when the virulence locus RD1 was deleted. Activation by IFN-γ induced a cell-autonomous response in hLECs via autophagy and NO production that restricted M. tuberculosis growth. Thus, depending on the activation status of LECs, autophagy can both promote and restrict replication. Together, these findings reveal a previously unrecognized role for hLECs and autophagy in tuberculosis pathogenesis and suggest that hLECs are a potential niche for M. tuberculosis that allows establishment of persistent infection in lymph nodes. PMID:26901813

High proportion of modern genotypes of M. tuberculosis and their affinity with drug resistance in northern region of India.

Science.gov (United States)

Dhatwalia, Sunil Kumar; Yadav, Rakesh; Behera, Digambar; Kaur, Harsimran; Kumar, Manoj; Sethi, Sunil

2017-09-01

Comparative genomics on the basis of TbD1 deletion has differentiated the members of Mycobacterium tuberculosis complex (MTC) in two major genogroups. They exhibit differential distribution and virulence potential. The present study was carried out to see the proportion of these genogroups and their association with drug resistance. The drug resistance pattern of 205 culture positive cases of M. tuberculosis and their relation with TbD1 deletion was analysed from the tertiary care centre. Overall proportion of genotypes (TbD1- and Tbd1+) and their association with drug resistance was also observed from the various studies from India. Our study reports that 85.4% of the isolates of M. tuberculosis were modern genotypes (TbD1-) and rest of 14.6% were ancient genotypes (TbD1+). 37 cases were of multiple drug resistant-TB (MDR-TB), 35 of them belongs to modern genogrop and rest of (2) were in ancient genogroup (p=0.12). Overall pooled estimate of proportion of modern genotype is 75.5% (CI 95%, 73.03-77.87) and 24.55% (CI 95%, 22.13-26.97) for ancient genotypes from the studies carried out in India. Modern genotypes were more rarely drug sensitive phenotypes with a relative risk (RR) of 0.89 (CI 95%, 0.74-1.07) while MDR cases were more in this group with an odds ratio (OR) of 2.27 (CI 95%, 0-1.07). This study demonstrates a higher proportion of modern genotypes in our region/India; which are more likely to be associated with drug resistance. Future, epidemiological/in vitro studies are required to ascertain the relationship between genotypes and their virulence potential. Copyright © 2017 International Society for Chemotherapy of Infection and Cancer. Published by Elsevier Ltd. All rights reserved.

MenA Is a Promising Drug Target for Developing Novel Lead Molecules to Combat Mycobacterium tuberculosis

OpenAIRE

Kurosu, Michio; Crick, Dean C.

2009-01-01

Potent inhibitors of MenA (1,4-dihydroxy-2-naphtoate prenyltrasferase) in Mycobacterium tuberculosis are identified, and are also effective in inhibiting growth of Mycobacterium tuberculosis at low concentrations. The MenA inhibitors possess common chemical structural features of ((alkylamino)alkoxyphenyl)(phenyl)methanones. Significantly, the MenA inhibitors can be synthesized in a few steps with high overall yields. The representative MenA inhibitors are highly effective in killing nonrepli...

Bloodstream Infections with Mycobacterium tuberculosis among HIV patients

Centers for Disease Control (CDC) Podcasts

2010-09-23

This podcast looks at bloodstream infections with Mycobacterium tuberculosis and other pathogens among outpatients infected with HIV in Southeast Asia. CDC health scientist Kimberly McCarthy discusses the study and why bloodstream infections occur in HIV-infected populations.  Created: 9/23/2010 by National Center for Emerging and Zoonotic Infectious Diseases (NCEZID).   Date Released: 9/23/2010.

Bacteriological diagnosis and molecular strain typing of Mycobacterium bovis and Mycobacterium caprae.

Science.gov (United States)

Gormley, E; Corner, L A L; Costello, E; Rodriguez-Campos, S

2014-10-01

The primary isolation of a Mycobacterium sp. of the Mycobacterium tuberculosis complex from an infected animal provides a definitive diagnosis of tuberculosis. However, as Mycobacterium bovis and Mycobacterium caprae are difficult to isolate, particularly for animals in the early stages of disease, success is dependent on the optimal performance of all aspects of the bacteriological process, from the initial choice of tissue samples at post-mortem examination or clinical samples, to the type of media and conditions used to cultivate the microorganism. Each step has its own performance characteristics, which can contribute to sensitivity and specificity of the procedure, and may need to be optimized in order to achieve the gold standard diagnosis. Having isolated the slow-growing mycobacteria, species identification and fine resolution strain typing are keys to understanding the epidemiology of the disease and to devise strategies to limit transmission of infection. New technologies have emerged that can now even discriminate different isolates from the same animal. In this review we highlight the key factors that contribute to the accuracy of bacteriological diagnosis of M. bovis and M. caprae, and describe the development of advanced genotyping techniques that are increasingly used in diagnostic laboratories for the purpose of supporting detailed epidemiological investigations. Copyright © 2014 Elsevier Ltd. All rights reserved.

Assessment of tuberculosis spatial hotspot areas in Antananarivo, Madagascar, by combining spatial analysis and genotyping.

Science.gov (United States)

Ratovonirina, Noël Harijaona; Rakotosamimanana, Niaina; Razafimahatratra, Solohery Lalaina; Raherison, Mamy Serge; Refrégier, Guislaine; Sola, Christophe; Rakotomanana, Fanjasoa; Rasolofo Razanamparany, Voahangy

2017-08-14

Tuberculosis (TB) remains a public health problem in Madagascar. A crucial element of TB control is the development of an easy and rapid method for the orientation of TB control strategies in the country. Our main objective was to develop a TB spatial hotspot identification method by combining spatial analysis and TB genotyping method in Antananarivo. Sputa of new pulmonary TB cases from 20 TB diagnosis and treatment centers (DTCs) in Antananarivo were collected from August 2013 to May 2014 for culture. Mycobacterium tuberculosis complex (MTBC) clinical isolates were typed by spoligotyping on a Luminex® 200 platform. All TB patients were respectively localized according to their neighborhood residence and the spatial distribution of all pulmonary TB patients and patients with genotypic clustered isolates were scanned respectively by the Kulldorff spatial scanning method for identification of significant spatial clustering. Areas exhibiting spatial clustering of patients with genotypic clustered isolates were considered as hotspot TB areas for transmission. Overall, 467 new cases were included in the study, and 394 spoligotypes were obtained (84.4%). New TB cases were distributed in 133 of the 192 Fokontany (administrative neighborhoods) of Antananarivo (1 to 15 clinical patients per Fokontany) and patients with genotypic clustered isolates were distributed in 127 of the 192 Fokontany (1 to 13 per Fokontany). A single spatial focal point of epidemics was detected when ignoring genotypic data (p = 0.039). One Fokontany of this focal point and three additional ones were detected to be spatially clustered when taking genotypes into account (p Madagascar and will allow better TB control strategies by public health authorities.

Mycobacterium tuberculosis and Mycobacterium marinum non-homologous end-joining proteins can function together to join DNA ends in Escherichia coli.

Science.gov (United States)

Wright, Douglas G; Castore, Reneau; Shi, Runhua; Mallick, Amrita; Ennis, Don G; Harrison, Lynn

2017-03-01

Mycobacterium tuberculosis and Mycobacterium smegmatis express a Ku protein and a DNA ligase D and are able to repair DNA double strand breaks (DSBs) by non-homologous end-joining (NHEJ). This pathway protects against DNA damage when bacteria are in stationary phase. Mycobacterium marinum is a member of this mycobacterium family and like M. tuberculosis is pathogenic. M. marinum lives in water, forms biofilms and infects fish and frogs. M. marinum is a biosafety level 2 (BSL2) organism as it can infect humans, although infections are limited to the skin. M. marinum is accepted as a model to study mycobacterial pathogenesis, as M. marinum and M. tuberculosis are genetically closely related and have similar mechanisms of survival and persistence inside macrophage. The aim of this study was to determine whether M. marinum could be used as a model to understand M. tuberculosis NHEJ repair. We identified and cloned the M. marinum genes encoding NHEJ proteins and generated E. coli strains that express the M. marinum Ku (Mm-Ku) and ligase D (Mm-Lig) individually or together (LHmKumLig strain) from expression vectors integrated at phage attachment sites in the genome. We demonstrated that Mm-Ku and Mm-Lig are both required to re-circularize Cla I-linearized plasmid DNA in E. coli. We compared repair of strain LHmKumLig with that of an E. coli strain (BWKuLig#2) expressing the M. tuberculosis Ku (Mt-Ku) and ligase D (Mt-Lig), and found that LHmKumLig performed 3.5 times more repair and repair was more accurate than BWKuLig#2. By expressing the Mm-Ku with the Mt-Lig, or the Mt-Ku with the Mm-Lig in E. coli, we have shown that the NHEJ proteins from M. marinum and M. tuberculosis can function together to join DNA DSBs. NHEJ repair is therefore conserved between the two species. Consequently, M. marinum is a good model to study NHEJ repair during mycobacterial pathogenesis. © The Author 2016. Published by Oxford University Press on behalf of the UK Environmental Mutagen

Evolutionary history and global spread of the Mycobacterium tuberculosis Beijing lineage.

OpenAIRE

Merker Matthias; Blin Camille; Mona Stefano; Duforet-Frebourg Nicolas; Lecher Sophie; Willery Eve; Blum Michael G B; Rüsch-Gerdes Sabine; Mokrousov Igor; Aleksic Eman; Allix-Béguec Caroline; Antierens Annick; Augustynowicz-Kopec Ewa; Ballif Marie; Barletta Francesca

2015-01-01

International audience; Mycobacterium tuberculosis strains of the Beijing lineage are globally distributed and are associated with the massive spread of multidrug-resistant (MDR) tuberculosis in Eurasia. Here we reconstructed the biogeographical structure and evolutionary history of this lineage by genetic analysis of 4,987 isolates from 99 countries and whole-genome sequencing of 110 representative isolates. We show that this lineage initially originated in the Far East, from where it radiat...

Contribution of spoligotyping and MIRU-VNTRs to characterize prevalent Mycobacterium tuberculosis genotypes infecting tuberculosis patients in Morocco.

Science.gov (United States)

Chaoui, Imane; Zozio, Thierry; Lahlou, Ouafae; Sabouni, Radia; Abid, Mohammed; El Aouad, Rajae; Akrim, Mohammed; Amzazi, Said; Rastogi, Nalin; El Mzibri, Mohammed

2014-01-01

In the present study, Mycobacterium tuberculosis complex (MTBC) clinical isolates from culture-positive TB patients in Morocco were studied by spoligotyping and 12-loci MIRU-VNTR typing methods to characterize prevalent genotypes (n = 219 isolates from 208 patients). Spoligotyping resulted in 39 unique patterns and 167 strains in 30 clusters (2-50 strains per cluster). Comparison with international database showed that 29 of 39 unique patterns matched existing shared spoligotype international types (SITs). Nine shared types containing 10 strains were newly created (SIT 2891 to SIT 2899); this led to the description of 69 SITs with 206 strains and two orphan patterns. The most prevalent spoligotype was SIT42 (LAM; n = 50 or 24% of isolates). The repartition of strains according to major MTBC clades was as follows LAM (46.1%)> Haarlem (26%) >ill-defined T superfamily (22.6%) and S clade (0.96%). On the other hand, Beijing, CAS (Central Asian) and EAI (East-African Indian) strains were absent in this setting. Subsequent 12-Loci MIRU typing resulted in a total of 25 SIT/MIT clusters (n = 66 isolates, 2-6 isolates per cluster), with a resulting recent transmission rate of 22.3%. The MIRU-VNTR patterns corresponded to 69 MITs for 138 strains and 46 orphan patterns. The most frequent patterns were MIT43 (n = 8), MIT9 (n = 7) and MIT42 (n = 7). HGDI analysis of the 12 MIRU loci showed that loci 10, 23 and 40 were highly discriminative in our setting. The results also underlined the usefulness of spoligotyping and MIRU-VNTR to detect mixed infections among certain of our TB patients. Globally, the results obtained showed that TB is almost exclusively transmitted in Morocco through evolutionary-modern MTBC lineages belonging to principal genetic groups 2/3 strains (Haarlem, LAM, T), with a high level of biodiversity seen by MIRU typing. This study provides with a 1st global snapshot of MTBC population structure in Morocco, and validates the potential use of spoligotyping in

Direct detection of Mycobacterium tuberculosis and drug resistance in respiratory specimen using Abbott Realtime MTB detection and RIF/INH resistance assay.

Science.gov (United States)

Tam, Kingsley King-Gee; Leung, Kenneth Siu-Sing; To, Sabrina Wai-Chi; Siu, Gilman Kit-Hang; Lau, Terrence Chi-Kong; Shek, Victor Chi-Man; Tse, Cindy Wing-Sze; Wong, Samson Sai-Yin; Ho, Pak-Leung; Yam, Wing-Cheong

2017-10-01

Abbott RealTime MTB (Abbott-RT) in conjunction with Abbott RealTime MTB RIF/INH Resistance (Abbott-RIF/INH) is a new, high-throughput automated nucleic acid amplification platform (Abbott-MDR) for detection of Mycobacterium tuberculosis complex (MTBC) and the genotypic markers for rifampicin (RIF) and isoniazid (INH) resistance directly from respiratory specimens. This prospective study evaluated the diagnostic performance of this new platform for MTBC and multidrug-resistant tuberculosis (MDR-TB) using 610 sputum specimens in a tuberculosis high-burden setting. Using conventional culture results and clinical background as reference standards, Abbott-RT exhibited an overall sensitivity and specificity of 95.2% and 99.8%, respectively. Genotypic RIF/INH resistance of 178 "MTB detected" specimens was subsequently analyzed by Abbott-RIF/INH. Compared to phenotypic drug susceptibility test results, Abbott-RIF/INH detected resistance genotypic markers in 84.6% MDR-TB, 80% mono-RIF-resistant and 66.7% mono-INH-resistant specimens. Two of the RIF-resistant specimens carried a novel single, nonsense mutation at rpoB Q513 and in silico simulation demonstrated that the truncated RpoB protein failed to bind with other subunits for transcription. Overall, Abbott-MDR platform provided high throughput and reliable diagnosis of MDR-TB within a TB high-burden region. Copyright © 2017 Elsevier Inc. All rights reserved.

Análise genotípica de isolados de Mycobacterium tuberculosis de um hospital em Lisboa, Portugal

Directory of Open Access Journals (Sweden)

João Perdigão

2009-09-01

Full Text Available Resumo: Portugal apresenta uma das taxas de incidência de tuberculose mais altas da União Europeia, tendo a Região de Saúde de Lisboa uma taxa de incidência muito acima da média nacional. O presente estudo analisa a transmissão, susceptibilidade aos antibacilares e características de uma população de estudo de um hospital central de Lisboa. Cento e trinta e dois isolados clínicos de Mycobacterium tuberculosis foram pre-viamente testados quanto à susceptibilidade aos antibacilares de primeira linha. A taxa de multirresistência encontrada foi de 3,0%, enquanto 13,6% dos isolados eram resistentes a um ou mais antibacilares de primeira linha. A serologia para o VIH estava disponível para 98 doentes, 26 (26,5% eram positivos. Os isolados foram genotipados por MIRU-VNTR e 53 (40,2% dos 132 isolados encontravam-se distribuídos por 17 clusters MIRU-VNTR diferentes de dois ou mais isolados. De todos os isolados analisados, 25,8% perten-ciam à família Lisboa. Concluímos que a transmissão de estirpes de Mycobacterium tuberculosis, resistentes e susceptíveis, está a ocorrer, com especial preocupação para as estirpes Lisboa.Rev Port Pneumol 2009; XV (5: 761-769 Abstract: Portugal has one of the highest tuberculosis notification rates of the European Union with Lisbon Health Region having an incidence rate well above the national average. The present study analyses the transmission, drug susceptibility and characteristics of a study population from a Central Lisbon’s Hospital. One hundred and thirty-two Mycobacterium tuberculosis clinical isolates were previously tested for drug susceptibility to first-line drugs. The multidrug (MDR resistance rate was found to be 3.0%, while 13.6% of the isolates were resistant to one or more first-line drugs. HIV serology was available for 98 patients, 26 (26.5% were positive. Genotyping was performed by MIRU-VNTR and 53 (40,2% out of the 132 isolates

Genome-wide analysis of multi- and extensively drug-resistant Mycobacterium tuberculosis

KAUST Repository

Coll, Francesc; Phelan, Jody; Hill-Cawthorne, Grant A.; Nair, Mridul; Mallard, Kim; Ali, Shahjahan; Abdallah, Abdallah; Alghamdi, Saad; Alsomali, Mona; Ahmed, Abdallah O.; Portelli, Stephanie; Oppong, Yaa; Alves, Adriana; Bessa, Theolis Barbosa; Campino, Susana; Caws, Maxine; Chatterjee, Anirvan; Crampin, Amelia C.; Dheda, Keertan; Furnham, Nicholas; Glynn, Judith R.; Grandjean, Louis; Minh Ha, Dang; Hasan, Rumina; Hasan, Zahra; Hibberd, Martin L.; Joloba, Moses; Jones-Ló pez, Edward C.; Matsumoto, Tomoshige; Miranda, Anabela; Moore, David J.; Mocillo, Nora; Panaiotov, Stefan; Parkhill, Julian; Penha, Carlos; Perdigã o, Joã o; Portugal, Isabel; Rchiad, ‍ Zineb; Robledo, Jaime; Sheen, Patricia; Shesha, Nashwa Talaat; Sirgel, Frik A.; Sola, Christophe; Oliveira Sousa, Erivelton; Streicher, Elizabeth M.; Helden, Paul Van; Viveiros, Miguel; Warren, Robert M.; McNerney, Ruth; Pain, Arnab; Clark, Taane G.

2018-01-01

To characterize the genetic determinants of resistance to antituberculosis drugs, we performed a genome-wide association study (GWAS) of 6,465 Mycobacterium tuberculosis clinical isolates from more than 30 countries. A GWAS approach within a mixed

Organization of the origins of replication of the chromosomes of Mycobacterium smegmatis, Mycobacterium leprae and Mycobacterium tuberculosis and isolation of a functional origin from M. smegmatis.

Science.gov (United States)

Salazar, L; Fsihi, H; de Rossi, E; Riccardi, G; Rios, C; Cole, S T; Takiff, H E

1996-04-01

The genus Mycobacterium is composed of species with widely differing growth rates ranging from approximately three hours in Mycobacterium smegmatis to two weeks in Mycobacterium leprae. As DNA replication is coupled to cell duplication, it may be regulated by common mechanisms. The chromosomal regions surrounding the origins of DNA replication from M. smegmatis, M. tuberculosis, and M. leprae have been sequenced, and show very few differences. The gene order, rnpA-rpmH-dnaA-dnaN-recF-orf-gyrB-gyrA, is the same as in other Gram-positive organisms. Although the general organization in M. smegmatis is very similar to that of Streptomyces spp., a closely related genus, M. tuberculosis and M. leprae differ as they lack an open reading frame, between dnaN and recF, which is similar to the gnd gene of Escherichia coli. Within the three mycobacterial species, there is extensive sequence conservation in the intergenic regions flanking dnaA, but more variation from the consensus DnaA box sequence was seen than in other bacteria. By means of subcloning experiments, the putative chromosomal origin of replication of M. smegmatis, containing the dnaA-dnaN region, was shown to promote autonomous replication in M. smegmatis, unlike the corresponding regions from M. tuberculosis or M. leprae.

SIMULATION OF EPIDEMIC TRANSMISSION OF МYCOBACTERIUM TUBERCULOSIS IN THE SAKHA REPUBLIC (YUKUTIA

Directory of Open Access Journals (Sweden)

S. N. Zhdаnovа

2017-01-01

Full Text Available Goal: reconstruction of epidemic transmission of Beijing genotype of Mycobacterium tuberculosis on the territory with limited migration based on the comparative study of 153 isolates isolated from patients of different generations in theSakhaRepublic (Yakutia.Materials and methods. The structure of Mycobacterium tuberculosis population was described and analyzed through MIRU-VNTR-genotyping, subtyping of the parts of RD105/RD207 genome, pattern classification as per clonal pattern of Merker M. et al (2015, and phylogenetic simulation.Results. It was found out that highly transmissive subtypes of Beijing genotype having a high potential of developing drug resistance was confidently more prevalent (χ2 = 8.3, p < 0.01 among younger people (born after 1990 compared to the older generation (born before 1959. It was also found out that during previous five decades a certain shift occurred in the structure of M. tuberculosis population.Conclusion. The obtained data confirm the hypothesis that epidemic sybtypes ofBeijing genotype were brought to the territory of theSakhaRepublic (Yakutia fairly recently (approximately fifty years ago compared to the other regions ofRussia.

Metabolic principles of persistence and pathogenicity in Mycobacterium tuberculosis.

Science.gov (United States)

Ehrt, Sabine; Schnappinger, Dirk; Rhee, Kyu Y

2018-04-24

Metabolism was once relegated to the supply of energy and biosynthetic precursors, but it has now become clear that it is a specific mediator of nearly all physiological processes. In the context of microbial pathogenesis, metabolism has expanded outside its canonical role in bacterial replication. Among human pathogens, this expansion has emerged perhaps nowhere more visibly than for Mycobacterium tuberculosis, the causative agent of tuberculosis. Unlike most pathogens, M. tuberculosis has evolved within humans, which are both host and reservoir. This makes unrestrained replication and perpetual quiescence equally incompatible strategies for survival as a species. In this Review, we summarize recent work that illustrates the diversity of metabolic functions that not only enable M. tuberculosis to establish and maintain a state of chronic infection within the host but also facilitate its survival in the face of drug pressure and, ultimately, completion of its life cycle.

Genetic diversity of the Mycobacterium tuberculosis complex in San Luis Potosí, México.

Science.gov (United States)

López-Rocha, Estela; Juárez-Álvarez, Julio; Riego-Ruiz, Lina; Enciso-Moreno, Leonor; Ortega-Aguilar, Francisco; Hernández-Nieto, Julián; Enciso-Moreno, José A; López-Revilla, Rubén

2013-05-01

Although epidemiologic and socioeconomic criteria and biomedical risk factors indicate high-priority for tuberculosis (TB) control in Mexico, molecular epidemiology studies of the disease in the country are scarce. Complete sociodemographic and clinical data were obtained from 248 of the 432 pulmonary TB (PTB) cases confirmed from 2006 to 2010 on the population under epidemiological surveillance in the state of San Luis Potosí, México. From most PTB cases with complete data Mycobacterium tuberculosis complex (MTC) isolates were recovered and their spoligotypes, lineages and families, geographic distribution and drug resistance determined. Pulmonary tuberculosis incidence ranged from 2.4 to 33.4 (cases per 100,000 inhabitants) in the six state sanitary jurisdictions that were grouped in regions of low (jurisdictions I-II-III), intermediate (jurisdictions IV-V) and high incidence (jurisdiction VI) with 6.2, 17.3 and 33.4 rates, respectively. Most patients were poor, 50-years-median-age males and housewives. Among the 237 MTC spoligotyped isolates, 232 corresponded to M. tuberculosis (104 spoligotypes in 24 clusters) and five to M. bovis. The predominant Euro-American lineage was distributed all over the state, the East-Asian lineage (Beijing family) in the capital city, the Indo-Oceanic (Manila family) in eastern localities, and M. bovis in rural localities. In San Luis Potosí TB affects mainly poor male adults and is caused by M. tuberculosis and to a minor extent by M. bovis. There is great genotypic diversity among M. tuberculosis strains, the Euro-American lineage being much more prevalent than the Indo-Oceanic and East-Asian lineages. The frequency of resistant strains is relatively low and not associated to any particular lineage.

Siderocalin inhibits the intracellular replication of Mycobacterium tuberculosis in macrophages

DEFF Research Database (Denmark)

Johnson, Erin E; Srikanth, Chittur V; Sandgren, Andreas

2010-01-01

Siderocalin is a secreted protein that binds to siderophores to prevent bacterial iron acquisition. While it has been shown to inhibit the growth of Mycobacterium tuberculosis (M.tb) in extracellular cultures, its effect on this pathogen within macrophages is not clear. Here, we show that sideroc...

Added value of IP-10 as a read-out of Mycobacterium tuberculosis

DEFF Research Database (Denmark)

Jenum, Synne; Dhanasekaran, Sivmakumaran; Ritz, Christian

2016-01-01

We have explored the added value of IFNγ-inducible protein 10 as a read-out of Mycobacterium tuberculosis specific immunity in young Indian children where the sensitivity of the IGRA for tuberculosis (TB) is poor. Reduced frequency of indeterminate results and an increased sensitivity for TB, sug...

Pulmonary Mycobacterium tuberculosis (Beijing strain infection in a stray dog : clinical communication

Directory of Open Access Journals (Sweden)

S.D.C. Parsons

2008-05-01

Full Text Available Mycobacterium tuberculosis infection in dogs is rarely reported and has not previously been documented in South Africa. A case of a stray Maltese crossbreed dog with extensive multifocal pulmonary tuberculosis due to M. tuberculosis is described. Pulmonary granulomas in this case were poorly encapsulated and contained large numbers of acid-fast bacteria, highlighting the potential for infected companion animals to excrete the pathogen. Treatment of canine tuberculosis is generally not advised, and for this reason, euthanasia of diseased animals must be advocated in most instances. Physicians and veterinarians must be aware that companion animals with active disease caused by M. tuberculosis could act as a potential source of infection.

In vitro and in vivo activities of the nitroimidazole TBA-354 against Mycobacterium tuberculosis.

Science.gov (United States)

Upton, A M; Cho, S; Yang, T J; Kim, Y; Wang, Y; Lu, Y; Wang, B; Xu, J; Mdluli, K; Ma, Z; Franzblau, S G

2015-01-01

Nitroimidazoles are a promising new class of antitubercular agents. The nitroimidazo-oxazole delamanid (OPC-67683, Deltyba) is in phase III trials for the treatment of multidrug-resistant tuberculosis, while the nitroimidazo-oxazine PA-824 is entering phase III for drug-sensitive and drug-resistant tuberculosis. TBA-354 (SN31354[(S)-2-nitro-6-((6-(4-trifluoromethoxy)phenyl)pyridine-3-yl)methoxy)-6,7-dihydro-5H-imidazo[2,1-b][1,3]oxazine]) is a pyridine-containing biaryl compound with exceptional efficacy against chronic murine tuberculosis and favorable bioavailability in preliminary rodent studies. It was selected as a potential next-generation antituberculosis nitroimidazole following an extensive medicinal chemistry effort. Here, we further evaluate the pharmacokinetic properties and activity of TBA-354 against Mycobacterium tuberculosis. TBA-354 is narrow spectrum and bactericidal in vitro against replicating and nonreplicating Mycobacterium tuberculosis, with potency similar to that of delamanid and greater than that of PA-824. The addition of serum protein or albumin does not significantly alter this activity. TBA-354 maintains activity against Mycobacterium tuberculosis H37Rv isogenic monoresistant strains and clinical drug-sensitive and drug-resistant isolates. Spontaneous resistant mutants appear at a frequency of 3 × 10(-7). In vitro studies and in vivo studies in mice confirm that TBA-354 has high bioavailability and a long elimination half-life. In vitro studies suggest a low risk of drug-drug interactions. Low-dose aerosol infection models of acute and chronic murine tuberculosis reveal time- and dose-dependent in vivo bactericidal activity that is at least as potent as that of delamanid and more potent than that of PA-824. Its superior potency and pharmacokinetic profile that predicts suitability for once-daily oral dosing suggest that TBA-354 be studied further for its potential as a next-generation nitroimidazole. Copyright © 2015, American

[Increased IL-4 production in response to virulent Mycobacterium tuberculosis in tuberculosis patients with advanced disease].

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Ordway, Diane J; Martins, Marta S; Costa, Leonor M; Freire, Mónica S; Arroz, Maria J; Dockrell, Hazel M; Ventura, Fernando A

2005-01-01

The study was designed to compare immune responses to Mycobacterium tuberculosis bacilli and antigens in healthy Portuguese subjects and pulmonary tuberculosis patients (TB), and to correlate immune status with clinical severity of tuberculosis disease. PBMC were cultured and stimulated with live and killed M. tuberculosis H37Rv and purified protein derivative (PPD) and lymphoproliferation and production of IFN-gamma and IL-5/IL-4 by these cultures were evaluated by the use of ELISA and multi-parameter flow cytometry. PBMC from 30 tuberculosis patients demonstrated significantly reduced amounts of proliferation and IFN-gamma when stimulated with live M. tuberculosis compared the control group. Of 15 tuberculosis patients tested for intracellular IL-4 following stimulation with M. tuberculosis, 7 showed greatly increased IL-4 production in CD8+ and gammadelta+ T cells. Tuberculosis patients demonstrated an increase of intracellular IL-4 after PBMC were stimulated with live M. tuberculosis in the CD4+ phenotype, but more notably in CD8+ and gammadelta TCR+ subsets. Increased production of IL-4 in tuberculosis patients was primarily in individuals with advanced involvement of lung parenchymal with high bacterial loads in sputum. These results suggest that an alteration in type 1 and type 2 cytokine balance can occur in patients with tuberculosis at an advanced clinical stage of disease.

Risk factors associated with cluster size of Mycobacterium tuberculosis (Mtb) of different RFLP lineages in Brazil.

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Peres, Renata Lyrio; Vinhas, Solange Alves; Ribeiro, Fabíola Karla Correa; Palaci, Moisés; do Prado, Thiago Nascimento; Reis-Santos, Bárbara; Zandonade, Eliana; Suffys, Philip Noel; Golub, Jonathan E; Riley, Lee W; Maciel, Ethel Leonor

2018-02-08

Tuberculosis (TB) transmission is influenced by patient-related risk, environment and bacteriological factors. We determined the risk factors associated with cluster size of IS6110 RFLP based genotypes of Mycobacterium tuberculosis (Mtb) isolates from Vitoria, Espirito Santo, Brazil. Cross-sectional study of new TB cases identified in the metropolitan area of Vitoria, Brazil between 2000 and 2010. Mtb isolates were genotyped by the IS6110 RFLP, spoligotyping and RD Rio . The isolates were classified according to genotype cluster sizes by three genotyping methods and associated patient epidemiologic characteristics. Regression Model was performed to identify factors associated with cluster size. Among 959 Mtb isolates, 461 (48%) cases had an isolate that belonged to an RFLP cluster, and six clusters with ten or more isolates were identified. Of the isolates spoligotyped, 448 (52%) were classified as LAM and 412 (48%) as non-LAM. Our regression model found that 6-9 isolates/RFLP cluster were more likely belong to the LAM family, having the RD Rio genotype and to be smear-positive (adjusted OR = 1.17, 95% CI 1.08-1.26; adjusted OR = 1.25, 95% CI 1.14-1.37; crude OR = 2.68, 95% IC 1.13-6.34; respectively) and living in a Serra city neighborhood decrease the risk of being in the 6-9 isolates/RFLP cluster (adjusted OR = 0.29, 95% CI, 0.10-0.84), than in the others groups. Individuals aged 21 to 30, 31 to 40 and > 50 years were less likely of belonging the 2-5 isolates/RFLP cluster than unique patterns compared to individuals cluster group (adjustment OR = 0.45, 95% CI 0.24-0.85) than unique patterns. We found that a large proportion of new TB infections in Vitoria is caused by prevalent Mtb genotypes belonging to the LAM family and RD Rio genotypes. Such information demonstrates that some genotypes are more likely to cause recent transmission. Targeting interventions such as screening in specific areas and social risk groups, should be a priority

Shared Mycobacterium avium genotypes observed among unlinked clinical and environmental isolates

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Our understanding of the sources of Mycobacterium avium infection is partially based on genotypic matching of pathogen isolates from cases and environmental sources. These approaches assume that genotypic identity is rare in isolates from unlinked cases or sources. To test this, ...

A quantitative and efficient approach to select MIRU-VNTR loci based on accumulation of the percentage differences of strains for discriminating divergent Mycobacterium tuberculosis sublineages.

Science.gov (United States)

Pan, Xin-Ling; Zhang, Chun-Lei; Nakajima, Chie; Fu, Jin; Shao, Chang-Xia; Zhao, Li-Na; Cui, Jia-Yi; Jiao, Na; Fan, Chang-Long; Suzuki, Yasuhiko; Hattori, Toshio; Li, Di; Ling, Hong

2017-07-26

Although several optimal mycobacterial interspersed repetitive units-variable number tandem repeat (MIRU-VNTR) loci have been suggested for genotyping homogenous Mycobacterium tuberculosis, including the Beijing genotype, a more efficient and convenient selection strategy for identifying optimal VNTR loci is needed. Here 281 M. tuberculosis isolates were analyzed. Beijing genotype and non-Beijing genotypes were identified, as well as Beijing sublineages, according to single nucleotide polymorphisms. A total of 22 MIRU-VNTR loci were used for genotyping. To efficiently select optimal MIRU-VNTR loci, we established accumulations of percentage differences (APDs) between the strains among the different genotypes. In addition, we constructed a minimum spanning tree for clustering analysis of the VNTR profiles. Our findings showed that eight MIRU-VNTR loci displayed disparities in h values of ≥0.2 between the Beijing genotype and non-Beijing genotype isolates. To efficiently discriminate Beijing and non-Beijing genotypes, an optimal VNTR set was established by adding loci with APDs ranging from 87.2% to 58.8%, resulting in the construction of a nine-locus set. We also found that QUB11a is a powerful locus for separating ST10s (including ST10, STF and STCH1) and ST22s (including ST22 and ST8) strains, whereas a combination of QUB11a, QUB4156, QUB18, Mtub21 and QUB26 could efficiently discriminate Beijing sublineages. Our findings suggested that two nine-locus sets were not only efficient for distinguishing the Beijing genotype from non-Beijing genotype strains, but were also suitable for sublineage genotyping with different discriminatory powers. These results indicate that APD represents a quantitative and efficient approach for selecting MIRU-VNTR loci to discriminate between divergent M. tuberculosis sublineages.

A quantitative and efficient approach to select MIRU–VNTR loci based on accumulation of the percentage differences of strains for discriminating divergent Mycobacterium tuberculosis sublineages

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Pan, Xin-Ling; Zhang, Chun-Lei; Nakajima, Chie; Fu, Jin; Shao, Chang-Xia; Zhao, Li-Na; Cui, Jia-Yi; Jiao, Na; Fan, Chang-Long; Suzuki, Yasuhiko; Hattori, Toshio; Li, Di; Ling, Hong

2017-01-01

Although several optimal mycobacterial interspersed repetitive units–variable number tandem repeat (MIRU–VNTR) loci have been suggested for genotyping homogenous Mycobacterium tuberculosis, including the Beijing genotype, a more efficient and convenient selection strategy for identifying optimal VNTR loci is needed. Here 281 M. tuberculosis isolates were analyzed. Beijing genotype and non-Beijing genotypes were identified, as well as Beijing sublineages, according to single nucleotide polymorphisms. A total of 22 MIRU–VNTR loci were used for genotyping. To efficiently select optimal MIRU–VNTR loci, we established accumulations of percentage differences (APDs) between the strains among the different genotypes. In addition, we constructed a minimum spanning tree for clustering analysis of the VNTR profiles. Our findings showed that eight MIRU–VNTR loci displayed disparities in h values of ≥0.2 between the Beijing genotype and non-Beijing genotype isolates. To efficiently discriminate Beijing and non-Beijing genotypes, an optimal VNTR set was established by adding loci with APDs ranging from 87.2% to 58.8%, resulting in the construction of a nine-locus set. We also found that QUB11a is a powerful locus for separating ST10s (including ST10, STF and STCH1) and ST22s (including ST22 and ST8) strains, whereas a combination of QUB11a, QUB4156, QUB18, Mtub21 and QUB26 could efficiently discriminate Beijing sublineages. Our findings suggested that two nine-locus sets were not only efficient for distinguishing the Beijing genotype from non-Beijing genotype strains, but were also suitable for sublineage genotyping with different discriminatory powers. These results indicate that APD represents a quantitative and efficient approach for selecting MIRU–VNTR loci to discriminate between divergent M. tuberculosis sublineages. PMID:28745309

Dehalogenation of Haloalkanes by Mycobacterium tuberculosis H37Rv and Other Mycobacteria

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Jesenská, Andrea; Sedlác̆ek, Ivo; Damborský, Jir̆í

2000-01-01

Haloalkane dehalogenases convert haloalkanes to their corresponding alcohols by a hydrolytic mechanism. To date, various haloalkane dehalogenases have been isolated from bacteria colonizing environments that are contaminated with halogenated compounds. A search of current databases with the sequences of these known haloalkane dehalogenases revealed the presence of three different genes encoding putative haloalkane dehalogenases in the genome of the human parasite Mycobacterium tuberculosis H37Rv. The ability of M. tuberculosis and several other mycobacterial strains to dehalogenate haloaliphatic compounds was therefore studied. Intact cells of M. tuberculosis H37Rv were found to dehalogenate 1-chlorobutane, 1-chlorodecane, 1-bromobutane, and 1,2-dibromoethane. Nine isolates of mycobacteria from clinical material and four strains from a collection of microorganisms were found to be capable of dehalogenating 1,2-dibromoethane. Crude extracts prepared from two of these strains, Mycobacterium avium MU1 and Mycobacterium smegmatis CCM 4622, showed broad substrate specificity toward a number of halogenated substrates. Dehalogenase activity in the absence of oxygen and the identification of primary alcohols as the products of the reaction suggest a hydrolytic dehalogenation mechanism. The presence of dehalogenases in bacterial isolates from clinical material, including the species colonizing both animal tissues and free environment, indicates a possible role of parasitic microorganisms in the distribution of degradation genes in the environment. PMID:10618227

Correlations between major risk factors and closely related Mycobacterium tuberculosis isolates grouped by three current enotyping procedures: a population-based study in northeast Mexico

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Katia Peñuelas-Urquides

2014-09-01

Full Text Available The characteristics of tuberculosis (TB patients related to a chain of recent TB transmissions were investigated. Mycobacterium tuberculosis (MTB isolates (120 were genotyped using the restriction fragment length polymorphism-IS6110 (R, spacer oligotyping (S and mycobacterial interspersed repetitive units-variable number of tandem repeats (M methods. The MTB isolates were clustered and the clusters were grouped according to the similarities of their genotypes. Spearman’s rank correlation coefficients between the groups of MTB isolates with similar genotypes and those patient characteristics indicating a risk for a pulmonary TB (PTB chain transmission were ana- lysed. The isolates showing similar genotypes were distributed as follows: SMR (5%, SM (12.5%, SR (1.67%, MR (0%, S (46.67%, M (5% and R (0%. The remaining 35 cases were orphans. SMR exhibited a significant correlation (p < 0.05 with visits to clinics, municipalities and comorbidities (primarily diabetes mellitus. S correlated with drug consumption and M with comorbidities. SMR is needed to identify a social network in metropolitan areas for PTB transmission and S and M are able to detect risk factors as secondary components of a transmission chain of TB.

Correlations between major risk factors and closely related Mycobacterium tuberculosis isolates grouped by three current enotyping procedures: a population-based study in northeast Mexico

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Peñuelas-Urquides, Katia; Martínez-Rodríguez, Herminia Guadalupe; Enciso-Moreno, José Antonio; Molina-Salinas, Gloria María; Silva-Ramírez, Beatriz; Padilla-Rivas, Gerardo Raymundo; Vera-Cabrera, Lucio; Torres-de-la-Cruz, Víctor Manuel; Martínez-Martínez, Yazmin Berenice; Ortega-García, Jorge Luis; Garza-Treviño, Elsa Nancy; Enciso-Moreno, Leonor; Saucedo-Cárdenas, Odila; Becerril-Montes, Pola; Said-Fernández/, Salvador

2014-01-01

The characteristics of tuberculosis (TB) patients related to a chain of recent TB transmissions were investigated. Mycobacterium tuberculosis (MTB) isolates (120) were genotyped using the restriction fragment length polymorphism-IS6110 (R), spacer oligotyping (S) and mycobacterial interspersed repetitive units-variable number of tandem repeats (M) methods. The MTB isolates were clustered and the clusters were grouped according to the similarities of their genotypes. Spearman’s rank correlation coefficients between the groups of MTB isolates with similar genotypes and those patient characteristics indicating a risk for a pulmonary TB (PTB) chain transmission were ana- lysed. The isolates showing similar genotypes were distributed as follows: SMR (5%), SM (12.5%), SR (1.67%), MR (0%), S (46.67%), M (5%) and R (0%). The remaining 35 cases were orphans. SMR exhibited a significant correlation (p < 0.05) with visits to clinics, municipalities and comorbidities (primarily diabetes mellitus). S correlated with drug consumption and M with comorbidities. SMR is needed to identify a social network in metropolitan areas for PTB transmission and S and M are able to detect risk factors as secondary components of a transmission chain of TB. PMID:25317710

Evaluation of the Mycobacterium tuberculosis SO2 vaccine using a natural tuberculosis infection model in goats.

Science.gov (United States)

Bezos, J; Casal, C; Álvarez, J; Roy, A; Romero, B; Rodríguez-Bertos, A; Bárcena, C; Díez, A; Juste, R; Gortázar, C; Puentes, E; Aguiló, N; Martín, C; de Juan, L; Domínguez, L

2017-05-01

The development of new vaccines against animal tuberculosis (TB) is a priority for improving the control and eradication of this disease, particularly in those species not subjected to compulsory eradication programmes. In this study, the protection conferred by the Mycobacterium tuberculosis SO 2 experimental vaccine was evaluated using a natural infection model in goats. Twenty-six goats were distributed in three groups: (1) 10 goats served as a control group; (2) six goats were subcutaneously vaccinated with BCG; and (3) 10 goats were subcutaneously vaccinated with SO 2 . Four months after vaccination, all groups were merged with goats infected with Mycobacterium bovis or Mycobacterium caprae, and tested over a 40 week period using a tuberculin intradermal test and an interferon-γ assay for mycobacterial reactivity. The severity of lesions was determined at post-mortem examination and the bacterial load in tissues were evaluated by culture. The two vaccinated groups had significantly lower lesion and bacterial culture scores than the control group (P<0.05); at the end of the study, the SO 2 vaccinated goats had the lowest lesion and culture scores. These results suggest that the SO 2 vaccine provides some protection against TB infection acquired from natural exposure. Copyright © 2017 Elsevier Ltd. All rights reserved.

rBCG30-induced immunity and cross-protection against Mycobacterium leprae challenge are enhanced by boosting with the Mycobacterium tuberculosis 30-kilodalton antigen 85B.

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Gillis, Thomas P; Tullius, Michael V; Horwitz, Marcus A

2014-09-01

Leprosy remains a major global health problem and typically occurs in regions in which tuberculosis is endemic. Vaccines are needed that protect against both infections and do so better than the suboptimal Mycobacterium bovis BCG vaccine. Here, we evaluated rBCG30, a vaccine previously demonstrated to induce protection superior to that of BCG against Mycobacterium tuberculosis and Mycobacterium bovis challenge in animal models, for efficacy against Mycobacterium leprae challenge in a murine model of leprosy. rBCG30 overexpresses the M. tuberculosis 30-kDa major secretory protein antigen 85B, which is 85% homologous with the M. leprae homolog (r30ML). Mice were sham immunized or immunized intradermally with BCG or rBCG30 and challenged 2.5 months later by injection of viable M. leprae into each hind footpad. After 7 months, vaccine efficacy was assessed by enumerating the M. leprae bacteria per footpad. Both BCG and rBCG30 induced significant protection against M. leprae challenge. In the one experiment in which a comparison between BCG and rBCG30 was feasible, rBCG30 induced significantly greater protection than did BCG. Immunization of mice with purified M. tuberculosis or M. leprae antigen 85B also induced protection against M. leprae challenge but less so than BCG or rBCG30. Notably, boosting rBCG30 with M. tuberculosis antigen 85B significantly enhanced r30ML-specific immune responses, substantially more so than boosting BCG, and significantly augmented protection against M. leprae challenge. Thus, rBCG30, a vaccine that induces improved protection against M. tuberculosis, induces cross-protection against M. leprae that is comparable or potentially superior to that induced by BCG, and boosting rBCG30 with antigen 85B further enhances immune responses and protective efficacy. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

MIRU-VNTR Genotyping of Mycobacterium tuberculosis Strains Using QIAxcel Technology: A Multicentre Evaluation Study.

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Nikolayevskyy, Vladyslav; Trovato, Alberto; Broda, Agnieszka; Borroni, Emanuele; Cirillo, Daniela; Drobniewski, Francis

2016-01-01

Molecular genotyping of M.tuberculosis is an important laboratory tool in the context of emerging drug resistant TB. The standard 24-loci MIRU-VNTR typing includes PCR amplification followed by the detection and sizing of PCR fragments using capillary electrophoresis on automated sequencers or using agarose gels. The QIAxcel Advanced system might offer a cost-effective medium-throughput alternative. Performance characteristics of the QIAxcel Advanced platform for the standard 24 VNTR loci panel was evaluated at two centres on a total of 140 DNA specimens using automated capillary electrophoresis as a reference method. Additionally 4 hypervariable MIRU-VNTR loci were evaluated on 53 crude DNA extracts. The sizing accuracy, interlaboratory reproducibility and overall instrument's performance were assessed during the study. An overall concordance with the reference method was high reaching 98.5% and 97.6% for diluted genomic and crude DNA extracts respectively. 91.4% of all discrepancies were observed in fragments longer than 700bp. The concordance for hypervariable loci was lower except for locus 4120 (96.2%). The interlaboratory reproducibility agreement rates were 98.9% and 91.3% for standard and hypervariable loci, respectively. Overall performance of the QIAxcel platform for M.tuberculosis genotyping using a panel of standard loci is comparable to that of established methods for PCR fragments up to 700bp. Inaccuracies in sizing of longer fragments could be resolved through using in-house size markers or introduction of offset values. To conclude, the QiaXcel system could be considered an effective alternative to existing methods in smaller reference and regional laboratories offering good performance and shorter turnaround times.

MIRU-VNTR Genotyping of Mycobacterium tuberculosis Strains Using QIAxcel Technology: A Multicentre Evaluation Study.

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Vladyslav Nikolayevskyy

Full Text Available Molecular genotyping of M.tuberculosis is an important laboratory tool in the context of emerging drug resistant TB. The standard 24-loci MIRU-VNTR typing includes PCR amplification followed by the detection and sizing of PCR fragments using capillary electrophoresis on automated sequencers or using agarose gels. The QIAxcel Advanced system might offer a cost-effective medium-throughput alternative.Performance characteristics of the QIAxcel Advanced platform for the standard 24 VNTR loci panel was evaluated at two centres on a total of 140 DNA specimens using automated capillary electrophoresis as a reference method. Additionally 4 hypervariable MIRU-VNTR loci were evaluated on 53 crude DNA extracts. The sizing accuracy, interlaboratory reproducibility and overall instrument's performance were assessed during the study.An overall concordance with the reference method was high reaching 98.5% and 97.6% for diluted genomic and crude DNA extracts respectively. 91.4% of all discrepancies were observed in fragments longer than 700bp. The concordance for hypervariable loci was lower except for locus 4120 (96.2%. The interlaboratory reproducibility agreement rates were 98.9% and 91.3% for standard and hypervariable loci, respectively. Overall performance of the QIAxcel platform for M.tuberculosis genotyping using a panel of standard loci is comparable to that of established methods for PCR fragments up to 700bp. Inaccuracies in sizing of longer fragments could be resolved through using in-house size markers or introduction of offset values. To conclude, the QiaXcel system could be considered an effective alternative to existing methods in smaller reference and regional laboratories offering good performance and shorter turnaround times.

Resistance to first-line anti-TB drugs is associated with reduced nitric oxide susceptibility in Mycobacterium tuberculosis

DEFF Research Database (Denmark)

Idh, Jonna; Mekonnen, Mekidim; Abate, Ebba

2012-01-01

The relative contribution of nitric oxide (NO) to the killing of Mycobacterium tuberculosis in human tuberculosis (TB) is controversial, although this has been firmly established in rodents. Studies have demonstrated that clinical strains of M. tuberculosis differ in susceptibility to NO, but how...

Impact of fgd1 and ddn Diversity in Mycobacterium tuberculosis Complex on In Vitro Susceptibility to PA-824

KAUST Repository

Feuerriegel, S.

2011-09-19

PA-824 is a promising drug candidate for the treatment of tuberculosis (TB). It is in phase II clinical trials as part of the first newly designed regimen containing multiple novel antituberculosis drugs (PA-824 in combination with moxifloxacin and pyrazinamide). However, given that the genes involved in resistance against PA-824 are not fully conserved in the Mycobacterium tuberculosis complex (MTBC), this regimen might not be equally effective against different MTBC genotypes. To investigate this question, we sequenced two PA-824 resistance genes (fgd1 [Rv0407] and ddn [Rv3547]) in 65 MTBC strains representing major phylogenetic lineages. The MICs of representative strains were determined using the modified proportion method in the Bactec MGIT 960 system. Our analysis revealed single-nucleotide polymorphisms in both genes that were specific either for several genotypes or for individual strains, yet none of these mutations significantly affected the PA-824 MICs (≤0.25 μg/ml). These results were supported by in silico modeling of the mutations identified in Fgd1. In contrast, “Mycobacterium canettii” strains displayed a higher MIC of 8 μg/ml. In conclusion, we found a large genetic diversity in PA-824 resistance genes that did not lead to elevated PA-824 MICs. In contrast, M. canettii strains had MICs that were above the plasma concentrations of PA-824 documented so far in clinical trials. As M. canettii is also intrinsically resistant against pyrazinamide, new regimens containing PA-824 and pyrazinamide might not be effective in treating M. canettii infections. This finding has implications for the design of multiple ongoing clinical trials.

Shared Mycobacterium avium genotypes observed among unlinked clinical and environmental isolates*

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Our understanding of the sources of Mycobacterium avium infection is partially based on genotypic matching of pathogen isolates from cases and environmental sources. These approaches assume that genotypic identity is rare in isolates from unlinked cases or sources. To test this a...

Immune Responses Involved in Mycobacterium Tuberculosis Infection

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Roghayeh Teimourpour

2016-09-01

Full Text Available Background and Objectives: Mycobacterium tuberculosis is the causative agent of tuberculosis (TB. Approximately one-third of the world's population is infected with M. tuberculosis. Despite the availability of drug and vaccine, it remains one of the leading causes of death in humans especially in developing countries. Epidemiological studies have indicated that only 10-30% of people exposed to tubercle bacillus are infected with M. tuberculosis, and at least 90% of the infected people finally do not acquire TB. The studies have indicated that the host efficient immune system has essential roles in the control of TB infection such that the highest rate of mortality and morbidity is seen in immunocompromised patients such as people infected with HIV. M. tuberculosis is an obligatory intracellular bacterium. It enters the body mainly through the respiratory tract and alveolar macrophages combat this pathogen most commonly. In addition to alveolar macrophages, various T-cell subpopulations need to be activated to overcome this bacterium's resistance to the host defense systems. CD4+ T cells, through production of several cytokines such as IFN-γ and TNF-α, and CD8+ T cells, through cytotoxic activities and induction of apoptosis in infected cells, play critical roles in inducing appropriate immune responses against M. tuberculosis. Although cell-mediated immunity is the cornerstone of host responses against TB and the recent studies have provided evidence for the importance of humoral and innate immune system in the control of TB, a profound understanding of the immune responses would provide a basis for development of new generations of vaccines and drugs. The present study addresses immune responses involved in M. tuberculosis infection.

Detection of mycobacterium tuberculosis in clinical samples by smear and culture

International Nuclear Information System (INIS)

Aftab, R.; Amjad, F.; Khurshid, R.

2009-01-01

A retrospective study was carried out in order to compare the smear stained by ZN and Lowenstein-Jensen (U) medium for the detection of Mycobacterium in clinical samples from different categories. Study Design: Laboratory based, Retrospective. Place and Duration: Sir Ganga Ram Hospital Fatima Jinnah Medical College, Lahore over a 5 year period between Jan 2001 and June 2006. Material and Methods: A total of 798 clinical samples were collected from patients of both sexes and all ages with a provisional diagnosis of tuberculosis. A Ziehl-Neelsen stain (ZN) and culture on U medium was performed for the detection of Mycobacterium. The specimen categories were sputum, pus, lymph node aspirate, urine and endometrial curetting. Results: Out of 5 types of 798 specimens received over a period of five years, only 46.3%) (n=369) were respiratory whereas the remaining 53.7% (n=429) were non respiratory tract category samples including sputum, pus, lymph node aspirate, urine and endometrial curetting. All were examined for the presence of acid-fast-bacilli (AFB) in ZN smear. Among these 3.578% gave a positive ZN stain while 11.65% were positive on culture. Out of a total of 369 respiratory tract category samples, 38 (10.3%) sputum samples were positive for AFB on both ZN and culture. Among the non respiratory tract category, 47 (28.2%) pus, 26 (31%) LN aspirate, 5 (15.6%) urine, 5 (3.42%) endometrial curetting were reported positive. Only 15.16% of clinical samples belonging to 5 different categories of specimens received from patients of both sexes with a provisional diagnosis of tuberculosis, tested positive for Mycobacterium by both ZN stain smear and culture on U medium. Among these, 3.57% were positive for AFB on ZN smear and 11.65% were positive on culture on U medium. Conclusion: These conventional techniques have proved to be reliable testing tools for detection of Mycobacterium tuberculosis in our settings but there is an urgent need to promote the use of Biotic and

Mycobacterial diversity causing multi- and extensively drug-resistant tuberculosis in Djibouti, Horn of Africa.

Science.gov (United States)

Millán-Lou, M I; Ollé-Goig, J E; Tortola, M T; Martin, C; Samper, S

2016-02-01

On detecting a high prevalence of multidrug-resistant tuberculosis (TB) in Djibouti, 32 Mycobacterium tuberculosis isolates of patients hospitalised in the TB referral centre of the capital were genotyped. A high variety of M. tuberculosis lineages, including lineage 1, Indo-Oceanic, lineage 2, East-Asian, lineage 3, East-African Indian and lineage 4, Euro-American, were detected.

Increasing incidence of fluoroquinolone-resistant Mycobacterium tuberculosis in Mumbai, India.

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Agrawal, D; Udwadia, Z F; Rodriguez, C; Mehta, A

2009-01-01

Tertiary referral centre, private hospital, Mumbai, India. To analyse the incidence of fluoroquinolone (FQ) resistant Mycobacterium tuberculosis (TB) in our laboratory from 1995 to 2004. Retrospective review and analysis of the drug susceptibility test records of all M. tuberculosis culture-positive samples from our Microbiology Department from 1995 to 2004. FQ resistance has increased exponentially in our laboratory, from 3% in 1996 to 35% in 2004. The incidence of multidrug-resistant tuberculosis has also increased during the same period, from 33% in 1995 to 56% in 2004. The incidence of FQ-resistant M. tuberculosis is gradually increasing to alarming levels. This may be due to widespread use of this vital group of drugs in the treatment of community-acquired infections. We urge that these broad spectrum antibiotics be used judiciously, and ideally be reserved for treatment of resistant TB in TB-endemic areas.

Role of Granulocyte-Macrophage Colony-Stimulating Factor Production by T Cells during Mycobacterium tuberculosis Infection.

Science.gov (United States)

Rothchild, Alissa C; Stowell, Britni; Goyal, Girija; Nunes-Alves, Cláudio; Yang, Qianting; Papavinasasundaram, Kadamba; Sassetti, Christopher M; Dranoff, Glenn; Chen, Xinchun; Lee, Jinhee; Behar, Samuel M

2017-10-24

Mice deficient for granulocyte-macrophage colony-stimulating factor (GM-CSF -/- ) are highly susceptible to infection with Mycobacterium tuberculosis , and clinical data have shown that anti-GM-CSF neutralizing antibodies can lead to increased susceptibility to tuberculosis in otherwise healthy people. GM-CSF activates human and murine macrophages to inhibit intracellular M. tuberculosis growth. We have previously shown that GM-CSF produced by iNKT cells inhibits growth of M. tuberculosis However, the more general role of T cell-derived GM-CSF during infection has not been defined and how GM-CSF activates macrophages to inhibit bacterial growth is unknown. Here we demonstrate that, in addition to nonconventional T cells, conventional T cells also produce GM-CSF during M. tuberculosis infection. Early during infection, nonconventional iNKT cells and γδ T cells are the main source of GM-CSF, a role subsequently assumed by conventional CD4 + T cells as the infection progresses. M. tuberculosis -specific T cells producing GM-CSF are also detected in the peripheral blood of infected people. Under conditions where nonhematopoietic production of GM-CSF is deficient, T cell production of GM-CSF is protective and required for control of M. tuberculosis infection. However, GM-CSF is not required for T cell-mediated protection in settings where GM-CSF is produced by other cell types. Finally, using an in vitro macrophage infection model, we demonstrate that GM-CSF inhibition of M. tuberculosis growth requires the expression of peroxisome proliferator-activated receptor gamma (PPARγ). Thus, we identified GM-CSF production as a novel T cell effector function. These findings suggest that a strategy augmenting T cell production of GM-CSF could enhance host resistance against M. tuberculosis IMPORTANCE Mycobacterium tuberculosis is the bacterium that causes tuberculosis, the leading cause of death by any infection worldwide. T cells are critical components of the immune

Highly structured genetic diversity of the Mycobacterium tuberculosis population in Djibouti.

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Godreuil, S; Renaud, F; Choisy, M; Depina, J J; Garnotel, E; Morillon, M; Van de Perre, P; Bañuls, A L

2010-07-01

Djibouti is an East African country with a high tuberculosis incidence. This study was conducted over a 2-month period in Djibouti, during which 62 consecutive patients with pulmonary tuberculosis (TB) were included. Genetic characterization of Mycobacterium tuberculosis, using mycobacterial interspersed repetitive-unit variable-number tandem-repeat typing and spoligotyping, was performed. The genetic and phylogenetic analysis revealed only three major families (Central Asian, East African Indian and T). The high diversity and linkage disequilibrium within each family suggest a long period of clonal evolution. A Bayesian approach shows that the phylogenetic structure observed in our sample of 62 isolates is very likely to be representative of the phylogenetic structure of the M. tuberculosis population in the total number of TB cases.

Characterization of Mycobacterium tuberculosis nicotinamidase/pyrazinamidase.

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Zhang, Hua; Deng, Jiao-Yu; Bi, Li-Jun; Zhou, Ya-Feng; Zhang, Zhi-Ping; Zhang, Cheng-Gang; Zhang, Ying; Zhang, Xian-En

2008-02-01

The nicotinamidase/pyrazinamidase (PncA) of Mycobacterium tuberculosis is involved in the activation of the important front-line antituberculosis drug pyrazinamide by converting it into the active form, pyrazinoic acid. Mutations in the pncA gene cause pyrazinamide resistance in M. tuberculosis. The properties of M. tuberculosis PncA were characterized in this study. The enzyme was found to be a 20.89 kDa monomeric protein. The optimal pH and temperature of enzymatic activity were pH 7.0 and 40 degrees C, respectively. Inductively coupled plasma-optical emission spectrometry revealed that the enzyme was an Mn(2+)/Fe(2+)-containing protein with a molar ratio of [Mn(2+)] to [Fe(2+)] of 1 : 1; furthermore, the external addition of either type of metal ion had no apparent effect on the wild-type enzymatic activity. The activity of the purified enzyme was determined by HPLC, and it was shown that it possessed similar pyrazinamidase and nicotinamidase activity, by contrast with previous reports. Nine PncA mutants were generated by site-directed mutagenesis. Determination of the enzymatic activity and metal ion content suggested that Asp8, Lys96 and Cys138 were key residues for catalysis, and Asp49, His51, His57 and His71 were essential for metal ion binding. Our data show that M. tuberculosis PncA may bind metal ions in a manner different from that observed in the case of Pyrococcus horikoshii PncA.

Aspectos relevantes del uso de Mycobacterium´habana´ como candidato vacunal contra la tuberculosis

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Iliana Valdés

2011-12-01

Full Text Available La eficacia protectora de la actual vacuna (BCG contra la tuberculosis, para contrarrestar las formas pulmonares de esta enfermedad y su reactivación, resulta variable o poco eficiente, lo cual impone la búsqueda urgente de nuevas alternativas profilácticas contra esta enfermedad. Basados en las ventajas inmunogénicas que ofrece el uso de vacunas vivas, se han encaminado diferentes estrategias de este tipo empleando mutantes auxotróficos de Mycobacterium tuberculosis , BCG recombinante o micobacterias no tuberculosas. Existen evidencias experimentales acerca de la protección conferida tras la vacunación con cepas vivas, inactivadas o fracciones proteicas de Mycobacterium'habana' TMC-5135 contra la infección por Mycobacterium tuberculosis. Esta respuesta protectora parece ir aparejada de escasos signos de virulencia en los modelos animales ensayados, lo cual coloca a M.´habana´ dentro de los posibles candidatos vacunales contra la tuberculosis al ajustarse a la condición que impone una vacuna clásica de reproducir la infección y los eventos inmunes que le suceden lo más fielmente posible a como ocurren de manera natural, sin causar extensos daños en el receptor.

Consequences of genomic diversity in Mycobacterium tuberculosis

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Coscolla, Mireia; Gagneux, Sebastien

2014-01-01

The causative agent of human tuberculosis, Mycobacterium tuberculosis complex (MTBC), comprises seven phylogenetically distinct lineages associated with different geographical regions. Here we review the latest findings on the nature and amount of genomic diversity within and between MTBC lineages. We then review recent evidence for the effect of this genomic diversity on mycobacterial phenotypes measured experimentally and in clinical settings. We conclude that overall, the most geographically widespread Lineage 2 (includes Beijing) and Lineage 4 (also known as Euro-American) are more virulent than other lineages that are more geographically restricted. This increased virulence is associated with delayed or reduced pro-inflammatory host immune responses, greater severity of disease, and enhanced transmission. Future work should focus on the interaction between MTBC and human genetic diversity, as well as on the environmental factors that modulate these interactions. PMID:25453224

Total hip replacement infected with Mycobacterium tuberculosis complicated by Addison disease and psoas muscle abscess: a case report

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De Nardo Pasquale

2012-01-01

Full Text Available Abstract Introduction Prosthetic joint infection due to Mycobacterium tuberculosis is occasionally encountered in clinical practice. To the best of our knowledge, this is the first report of a prosthetic joint infection due to Mycobacterium tuberculosis complicated by psoas abscesses and secondary Addison disease. Case presentation A 67-year-old immunocompetent Caucasian woman underwent total left hip arthroplasty because of osteoarthritis. After 18 months, she underwent arthroplasty revision for a possible prosthetic infection. Periprosthetic tissue specimens for bacteria were negative, and empirical antibiotic therapy was unsuccessful. She was then admitted to our department because of complications arising 22 months after arthroplasty. A physical examination revealed a sinus tract overlying her left hip and skin and mucosal pigmentation. Her levels of C-reactive protein, basal cortisol, adrenocorticotropic hormone, and sodium were out of normal range. Results of the tuberculin skin test and QuantiFERON-TB Gold test were positive. Computed tomography revealed a periprosthetic abscess and the inclusion of the left psoas muscle. Results of microbiological tests were negative, but polymerase chain reaction of a specimen taken from the hip fistula was positive for Mycobacterium tuberculosis. Our patient's condition was diagnosed as prosthetic joint infection and muscle psoas abscess due to Mycobacterium tuberculosis and secondary Addison disease. She underwent standard treatment with rifampicin, ethambutol, isoniazid, and pyrazinamide associated with hydrocortisone and fludrocortisone. At 15 months from the beginning of therapy, she was in good clinical condition and free of symptoms. Conclusions Prosthetic joint infection with Mycobacterium tuberculosis is uncommon. A differential diagnosis of tuberculosis should be considered when dealing with prosthetic joint infection, especially when repeated smears and histology examination from infected

Identification of Secretory Proteins in Mycobacterium tuberculosis Using Pseudo Amino Acid Composition

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Huan Yang

2016-01-01

Full Text Available Tuberculosis is killing millions of lives every year and on the blacklist of the most appalling public health problems. Recent findings suggest that secretory protein of Mycobacterium tuberculosis may serve the purpose of developing specific vaccines and drugs due to their antigenicity. Responding to global infectious disease, we focused on the identification of secretory proteins in Mycobacterium tuberculosis. A novel method called MycoSec was designed by incorporating g-gap dipeptide compositions into pseudo amino acid composition. Analysis of variance-based technique was applied in the process of feature selection and a total of 374 optimal features were obtained and used for constructing the final predicting model. In the jackknife test, MycoSec yielded a good performance with the area under the receiver operating characteristic curve of 0.93, demonstrating that the proposed system is powerful and robust. For user’s convenience, the web server MycoSec was established and an obliging manual on how to use it was provided for getting around any trouble unnecessary.

In-Silico Testing Of Nutraceutical Against The Murd Enzymes From Mycobacterium Tuberculosis

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Mohammad Teimouri

2015-08-01

Full Text Available In spite of availability of moderately protective vaccine and antibiotics new antibacterial agents are urgently needed to decrease the global incidence of Mycobacterium tuberculosis infections. Mur family is an important target for the development of new drugs as they are involved in the biosynthesis of bacterial cell wall. MurC-MurF ligases catalyze a series of irreversible steps in the biosynthesis of peptidoglycan precursor i.e. MurD catalyzes the ligation of D-glutamate to the nucleotide precursor UMA. Here we developed a homology model of MurD from M. Tuberculosis and was validated by using rampage Errat and ProSA online servers. Different nutraceuticals were tested and reported for their activity. Among the 14 nutraceuticals Diosgenin Xanthohumol Capsaicin 1-acetoxychavicol acetate and 6-Gingerol have best docking score. The best of all was Diosgenin with the docking score -14.22988 Xanthohumol with -13.923555 Capsaicin with -12.880404 1-acetoxychavicol acetate with -12.573502 and 6-Gingerol -12.349156 which will play a guiding role in the experimental design and development of mycobacterium tuberculosis MurD

[Identification of the Mycobacterium tuberculosis Beijing lineage in Ecuador].

Science.gov (United States)

Jiménez, Patricia; Calvopiña, Karina; Herrera, Diana; Rojas, Carlos; Pérez-Lago, Laura; Grijalva, Marcelo; Guna, Remedios; García-de Viedma, Darío

2017-06-01

Mycobacterium tuberculosis Beijing lineage isolates are considered to be especially virulent, transmissible and prone to acquire resistances. Beijing strains have been reported worldwide, but studies in Latin America are still scarce. The only multinational study performed in the region indicated a heterogeneous distribution for this lineage, which was absent in Chile, Colombia and Ecuador, although further studies found the lineage in Chile and Colombia. To search for the presence of the Beijing lineage in Ecuador, the only country in the region where it remains unreported. We obtained a convenience sample (2006-2012) from two hospitals covering different populations. The isolates were genotyped using 24-MIRU-VNTR. Lineages were assigned by comparing their patterns to those in the MIRU-VNTRplus platform. Isolates belonging to the Beijing lineage were confirmed by allele-specific PCR. We identified the first Beijing isolate in Ecuador in an unexpected epidemiological scenario: A patient was infected in the Andean region, in a population with low mobility and far from the borders of the neighboring countries where Beijing strains had been previously reported. This is the first report of the presence of the Beijing lineage in Ecuador in an unusual epidemiological context that deserves special attention.

Human B cells produce chemokine CXCL10 in the presence of Mycobacterium tuberculosis specific T cells

DEFF Research Database (Denmark)

Hoff, Soren T; Salman, Ahmed M; Ruhwald, Morten

2015-01-01

BACKGROUND: The role of B cells in human host response to Mycobacterium tuberculosis (Mtb) infection is still controversial, but recent evidence suggest that B cell follicle like structures within the lung may influence host responses through regulation of the local cytokine environment. A candid......BACKGROUND: The role of B cells in human host response to Mycobacterium tuberculosis (Mtb) infection is still controversial, but recent evidence suggest that B cell follicle like structures within the lung may influence host responses through regulation of the local cytokine environment...

Interplay between Mutations and Efflux in Drug Resistant Clinical Isolates of Mycobacterium tuberculosis

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Miguel Viveiros

2017-04-01

Full Text Available Numerous studies show efflux as a universal bacterial mechanism contributing to antibiotic resistance and also that the activity of the antibiotics subject to efflux can be enhanced by the combined use of efflux inhibitors. Nevertheless, the contribution of efflux to the overall drug resistance levels of clinical isolates of Mycobacterium tuberculosis is poorly understood and still is ignored by many. Here, we evaluated the contribution of drug efflux plus target-gene mutations to the drug resistance levels in clinical isolates of M. tuberculosis. A panel of 17 M. tuberculosis clinical strains were characterized for drug resistance associated mutations and antibiotic profiles in the presence and absence of efflux inhibitors. The correlation between the effect of the efflux inhibitors and the resistance levels was assessed by quantitative drug susceptibility testing. The bacterial growth/survival vs. growth inhibition was analyzed through the comparison between the time of growth in the presence and absence of an inhibitor. For the same mutation conferring antibiotic resistance, different MICs were observed and the different resistance levels found could be reduced by efflux inhibitors. Although susceptibility was not restored, the results demonstrate the existence of a broad-spectrum synergistic interaction between antibiotics and efflux inhibitors. The existence of efflux activity was confirmed by real-time fluorometry. Moreover, the efflux pump genes mmr, mmpL7, Rv1258c, p55, and efpA were shown to be overexpressed in the presence of antibiotics, demonstrating the contribution of these efflux pumps to the overall resistance phenotype of the M. tuberculosis clinical isolates studied, independently of the genotype of the strains. These results showed that the drug resistance levels of multi- and extensively-drug resistant M. tuberculosis clinical strains are a combination between drug efflux and the presence of target-gene mutations, a reality

Expression of Mycobacterium smegmatis pyrazinamidase in Mycobacterium tuberculosis confers hypersensitivity to pyrazinamide and related amides.

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Boshoff, H I; Mizrahi, V

2000-10-01

A pyrazinamidase (PZase)-deficient pncA mutant of Mycobacterium tuberculosis, constructed by allelic exchange, was used to investigate the effects of heterologous amidase gene expression on the susceptibility of this organism to pyrazinamide (PZA) and related amides. The mutant was highly resistant to PZA (MIC, >2,000 microg/ml), in accordance with the well-established role of pncA in the PZA susceptibility of M. tuberculosis (A. Scorpio and Y. Zhang, Nat. Med. 2:662-667, 1996). Integration of the pzaA gene encoding the major PZase/nicotinamidase from Mycobacterium smegmatis (H. I. M. Boshoff and V. Mizrahi, J. Bacteriol. 180:5809-5814, 1998) or the M. tuberculosis pncA gene into the pncA mutant complemented its PZase/nicotinamidase defect. In both pzaA- and pncA-complemented mutant strains, the PZase activity was detected exclusively in the cytoplasm, suggesting an intracellular localization for PzaA and PncA. The pzaA-complemented strain was hypersensitive to PZA (MIC, /=20 microg/ml) and was also sensitive to benzamide (MIC, 20 microg/ml), unlike the wild-type and pncA-complemented mutant strains, which were highly resistant to this amide (MIC, >500 microg/ml). This finding was consistent with the observation that benzamide is hydrolyzed by PzaA but not by PncA. Overexpression of PzaA also conferred sensitivity to PZA, nicotinamide, and benzamide on M. smegmatis (MIC, 150 microg/ml in all cases) and rendered Escherichia coli hypersensitive for growth at low pH.

Whole-genome-based Mycobacterium tuberculosis surveillance: a standardized, portable, and expandable approach.

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Kohl, Thomas A; Diel, Roland; Harmsen, Dag; Rothgänger, Jörg; Walter, Karen Meywald; Merker, Matthias; Weniger, Thomas; Niemann, Stefan

2014-07-01

Whole-genome sequencing (WGS) allows for effective tracing of Mycobacterium tuberculosis complex (MTBC) (tuberculosis pathogens) transmission. However, it is difficult to standardize and, therefore, is not yet employed for interlaboratory prospective surveillance. To allow its widespread application, solutions for data standardization and storage in an easily expandable database are urgently needed. To address this question, we developed a core genome multilocus sequence typing (cgMLST) scheme for clinical MTBC isolates using the Ridom SeqSphere(+) software, which transfers the genome-wide single nucleotide polymorphism (SNP) diversity into an allele numbering system that is standardized, portable, and not computationally intensive. To test its performance, we performed WGS analysis of 26 isolates with identical IS6110 DNA fingerprints and spoligotyping patterns from a longitudinal outbreak in the federal state of Hamburg, Germany (notified between 2001 and 2010). The cgMLST approach (3,041 genes) discriminated the 26 strains with a resolution comparable to that of SNP-based WGS typing (one major cluster of 22 identical or closely related and four outlier isolates with at least 97 distinct SNPs or 63 allelic variants). Resulting tree topologies are highly congruent and grouped the isolates in both cases analogously. Our data show that SNP- and cgMLST-based WGS analyses facilitate high-resolution discrimination of longitudinal MTBC outbreaks. cgMLST allows for a meaningful epidemiological interpretation of the WGS genotyping data. It enables standardized WGS genotyping for epidemiological investigations, e.g., on the regional public health office level, and the creation of web-accessible databases for global TB surveillance with an integrated early warning system. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Genetic diversity of the Mycobacterium tuberculosis Complex in San Luis Potosí, México

Science.gov (United States)

2013-01-01

Background Although epidemiologic and socioeconomic criteria and biomedical risk factors indicate high-priority for tuberculosis (TB) control in Mexico, molecular epidemiology studies of the disease in the country are scarce. Methods Complete sociodemographic and clinical data were obtained from 248 of the 432 pulmonary TB (PTB) cases confirmed from 2006 to 2010 on the population under epidemiological surveillance in the state of San Luis Potosí, México. From most PTB cases with complete data Mycobacterium tuberculosis complex (MTC) isolates were recovered and their spoligotypes, lineages and families, geographic distribution and drug resistance determined. Results Pulmonary tuberculosis incidence ranged from 2.4 to 33.4 (cases per 100,000 inhabitants) in the six state sanitary jurisdictions that were grouped in regions of low (jurisdictions I-II-III), intermediate (jurisdictions IV-V) and high incidence (jurisdiction VI) with 6.2, 17.3 and 33.4 rates, respectively. Most patients were poor, 50-years-median-age males and housewives. Among the 237 MTC spoligotyped isolates, 232 corresponded to M. tuberculosis (104 spoligotypes in 24 clusters) and five to M. bovis. The predominant Euro-American lineage was distributed all over the state, the East-Asian lineage (Beijing family) in the capital city, the Indo-Oceanic (Manila family) in eastern localities, and M. bovis in rural localities. Conclusions In San Luis Potosí TB affects mainly poor male adults and is caused by M. tuberculosis and to a minor extent by M. bovis. There is great genotypic diversity among M. tuberculosis strains, the Euro-American lineage being much more prevalent than the Indo-Oceanic and East-Asian lineages. The frequency of resistant strains is relatively low and not associated to any particular lineage. PMID:23635381

Updates on antibody functions in Mycobacterium tuberculosis infection and their relevance for developing a vaccine against tuberculosis.

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Achkar, Jacqueline M; Prados-Rosales, Rafael

2018-04-12

A more effective vaccine to control tuberculosis (TB), a major global public health problem, is urgently needed. Current vaccine candidates focus predominantly on eliciting cell-mediated immunity but other arms of the immune system also contribute to protection against TB. We review here recent studies that enhance our current knowledge of antibody-mediated functions against Mycobacterium tuberculosis. These findings, which contribute to the increasing evidence that antibodies have a protective role against TB, include demonstrations that firstly distinct human antibody Fc glycosylation patterns, found in latent M. tuberculosis infection but not in active TB, influence the efficacy of the host to control M. tuberculosis infection, secondly antibody isotype influences human antibody functions, and thirdly that antibodies targeting M. tuberculosis surface antigens are protective. We discuss these findings in the context of TB vaccine development and highlight the need for further research on antibody-mediated immunity in M. tuberculosis infection. Copyright © 2018 Elsevier Ltd. All rights reserved.

Distribution of Insertion- and Deletion-Associated Genetic Polymorphisms among Four Mycobacterium tuberculosis Phospholipase C Genes and Associations with Extrathoracic Tuberculosis: a Population-Based Study

OpenAIRE

Kong, Y.; Cave, M. D.; Yang, D.; Zhang, L.; Marrs, C. F.; Foxman, B.; Bates, J. H.; Wilson, F.; Mukasa, L. N.; Yang, Z. H.

2005-01-01

The Mycobacterium tuberculosis genome contains four phospholipase C (PLC)-encoding genes, designated plcA, plcB, plcC, and plcD, respectively. Each of the four genes contributes to the overall PLC activity of M. tuberculosis. PLC is hypothesized to contribute to M. tuberculosis virulence. Infection of M. tuberculosis strains carrying a truncated plcD gene is associated with the occurrence of extrathoracic tuberculosis. However, whether the other three plc genes are also associated with extrat...

Disinfectant-susceptibility of multi-drug-resistant Mycobacterium tuberculosis isolated in Japan

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Noriko Shinoda

2016-02-01

Full Text Available Abstract Background Multi-drug-resistant Mycobacterium tuberculosis has been an important problem in public health around the world. However, limited information about disinfectant-susceptibility of multi-drug-resistant strain of M. tuberculosis was available. Findings We studied susceptibility of several Japanese isolates of multi-drug-resistant M. tuberculosis against disinfectants, which are commonly used in clinical and research laboratories. We selected a laboratory reference strain (H37Rv and eight Japanese isolates, containing five drug-susceptible strains and three multi-drug-resistant strains, and determined profiles of susceptibility against eight disinfectants. The M. tuberculosis strains were distinguished into two groups by the susceptibility profile. There was no relationship between multi-drug-resistance and disinfectant-susceptibility in the M. tuberculosis strains. Cresol soap and oxydol were effective against all strains we tested, regardless of drug resistance. Conclusions Disinfectant-resistance is independent from multi-drug-resistance in M. tuberculosis. Cresol soap and oxydol were effective against all strains we tested, regardless of drug resistance.

Orchestration of pulmonary T cell immunity during Mycobacterium tuberculosis infection: immunity interruptus

Science.gov (United States)

Behar, Samuel M.; Carpenter, Stephen M.; Booty, Matthew G.; Barber, Daniel L.; Jayaraman, Pushpa

2014-01-01

Despite the introduction almost a century ago of Mycobacterium bovis BCG (BCG), an attenuated form of M. bovis that is used as a vaccine against Mycobacterium tuberculosis, tuberculosis remains a global health threat and kills more than 1.5 million people each year. This is mostly because BCG fails to prevent pulmonary disease – the contagious form of tuberculosis. Although there have been significant advances in understanding how the immune system responds to infection, the qualities that define protective immunity against M. tuberculosis remain poorly characterized. The ability to predict who will maintain control over the infection and who will succumb to clinical disease would revolutionize our approach to surveillance, control, and treatment. Here we review the current understanding of pulmonary T cell responses following M. tuberculosis infection. While infection elicits a strong immune response that contains infection, M. tuberculosis evades eradication. Traditionally, its intracellular lifestyle and alteration of macrophage function are viewed as the dominant mechanisms of evasion. Now we appreciate that chronic inflammation leads to T cell dysfunction. While this may arise as the host balances the goals of bacterial sterilization and avoidance of tissue damage, it is becoming clear that T cell dysfunction impairs host resistance. Defining the mechanisms that lead to T cell dysfunction is crucial as memory T cell responses are likely to be subject to the same subject to the same pressures. Thus, success of T cell based vaccines is predicated on memory T cells avoiding exhaustion while at the same time not promoting overt tissue damage. PMID:25311810

Pulmonary tuberculosis

Science.gov (United States)

TB; Tuberculosis - pulmonary; Mycobacterium - pulmonary ... Pulmonary TB is caused by the bacterium Mycobacterium tuberculosis (M tuberculosis) . TB is contagious. This means the bacteria is easily spread from an infected person ...

bioA mutant of Mycobacterium tuberculosis shows severe growth defect and imparts protection against tuberculosis in guinea pigs

Science.gov (United States)

Kar, Ritika; Nangpal, Prachi; Mathur, Shubhita; Singh, Swati

2017-01-01

Owing to the devastation caused by tuberculosis along with the unsatisfactory performance of the Bacillus Calmette–Guérin (BCG) vaccine, a more efficient vaccine than BCG is required for the global control of tuberculosis. A number of studies have demonstrated an essential role of biotin biosynthesis in the growth and survival of several microorganisms, including mycobacteria, through deletion of the genes involved in de novo biotin biosynthesis. In this study, we demonstrate that a bioA mutant of Mycobacterium tuberculosis (MtbΔbioA) is highly attenuated in the guinea pig model of tuberculosis when administered aerogenically as well as intradermally. Immunization with MtbΔbioA conferred significant protection in guinea pigs against an aerosol challenge with virulent M. tuberculosis, when compared with the unvaccinated animals. Booster immunization with MtbΔbioA offered no advantage over a single immunization. These experiments demonstrate the vaccinogenic potential of the attenuated M. tuberculosis bioA mutant against tuberculosis. PMID:28658275

bioA mutant of Mycobacterium tuberculosis shows severe growth defect and imparts protection against tuberculosis in guinea pigs.

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Ritika Kar

Full Text Available Owing to the devastation caused by tuberculosis along with the unsatisfactory performance of the Bacillus Calmette-Guérin (BCG vaccine, a more efficient vaccine than BCG is required for the global control of tuberculosis. A number of studies have demonstrated an essential role of biotin biosynthesis in the growth and survival of several microorganisms, including mycobacteria, through deletion of the genes involved in de novo biotin biosynthesis. In this study, we demonstrate that a bioA mutant of Mycobacterium tuberculosis (MtbΔbioA is highly attenuated in the guinea pig model of tuberculosis when administered aerogenically as well as intradermally. Immunization with MtbΔbioA conferred significant protection in guinea pigs against an aerosol challenge with virulent M. tuberculosis, when compared with the unvaccinated animals. Booster immunization with MtbΔbioA offered no advantage over a single immunization. These experiments demonstrate the vaccinogenic potential of the attenuated M. tuberculosis bioA mutant against tuberculosis.

Immunogenic membrane-associated proteins of Mycobacterium tuberculosis revealed by proteomics.

Science.gov (United States)

Sinha, Sudhir; Kosalai, K; Arora, Shalini; Namane, Abdelkader; Sharma, Pawan; Gaikwad, Anil N; Brodin, Priscille; Cole, Stewart T

2005-07-01

Membrane-associated proteins of Mycobacterium tuberculosis offer a challenge, as well as an opportunity, in the quest for better therapeutic and prophylactic interventions against tuberculosis. The authors have previously reported that extraction with the detergent Triton X-114 (TX-114) is a useful step in proteomic analysis of mycobacterial cell membranes, and detergent-soluble membrane proteins of mycobacteria are potent stimulators of human T cells. In this study 1-D and 2-D gel electrophoresis-based protocols were used for the analysis of proteins in the TX-114 extract of M. tuberculosis membranes. Peptide mass mapping (using MALDI-TOF-MS, matrix assisted laser desorption/ionization time of flight mass spectrometry) of 116 samples led to the identification of 105 proteins, 9 of which were new to the M. tuberculosis proteome. Functional orthologues of 73 of these proteins were also present in Mycobacterium leprae, suggesting their relative importance. Bioinformatics predicted that as many as 73% of the proteins had a hydrophobic disposition. 1-D gel electrophoresis revealed more hydrophobic/transmembrane and basic proteins than 2-D gel electrophoresis. Identified proteins fell into the following major categories: protein synthesis, cell wall biogenesis/architecture and conserved hypotheticals/unknowns. To identify immunodominant proteins of the detergent phase (DP), 14 low-molecular-mass fractions prepared by continuous-elution gel electrophoresis were subjected to T cell activation assays using blood samples from BCG-vaccinated healthy donors from a tuberculosis endemic area. Analysis of the responses (cell proliferation and IFN-gamma production) showed that the immunodominance of certain DP fractions was most probably due to ribosomal proteins, which is consistent with both their specificity for mycobacteria and their abundance. Other membrane-associated proteins, including transmembrane proteins/lipoproteins and ESAT-6, did not appear to contribute

Inhibition of Glutamine Synthetase: A Potential Drug Target in Mycobacterium tuberculosis

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Sherry L. Mowbray

2014-08-01

Full Text Available Tuberculosis is an infectious disease caused by Mycobacterium tuberculosis. Globally, tuberculosis is second only to AIDS in mortality and the disease is responsible for over 1.3 million deaths each year. The impractically long treatment schedules (generally 6–9 months and unpleasant side effects of the current drugs often lead to poor patient compliance, which in turn has resulted in the emergence of multi-, extensively- and totally-drug resistant strains. The development of new classes of anti-tuberculosis drugs and new drug targets is of global importance, since attacking the bacterium using multiple strategies provides the best means to prevent resistance. This review presents an overview of the various strategies and compounds utilized to inhibit glutamine synthetase, a promising target for the development of drugs for TB therapy.

Infections with Mycobacterium tuberculosis and Mycobacterium avium among HIV-infected patients after the introduction of highly active antiretroviral therapy. EuroSIDA Study Group JD

DEFF Research Database (Denmark)

Kirk, O; Gatell, J M; Mocroft, A

2000-01-01

the introduction of HAART, using data from the EuroSIDA study, a European, multicenter observational cohort of more than 7,000 patients. Overall incidences of Mycobacterium tuberculosis (TB) and Mycobacterium avium complex (MAC) were 0.8 and 1.4 cases/100 person-years of follow-up (PYF), decreasing from 1.8 (TB...

Proteogenomic Investigation of Strain Variation in Clinical Mycobacterium tuberculosis Isolates

KAUST Repository

Heunis, Tiaan

2017-08-18

Mycobacterium tuberculosis consists of a large number of different strains that display unique virulence characteristics. Whole-genome sequencing has revealed substantial genetic diversity among clinical M. tuberculosis isolates, and elucidating the phenotypic variation encoded by this genetic diversity will be of utmost importance to fully understand M. tuberculosis biology and pathogenicity. In this study we integrated whole-genome sequencing and mass spectrometry (GeLC-MS/MS) to reveal strain-specific characteristics in the proteomes of two clinical M. tuberculosis Latin American-Mediterranean isolates. Using this approach we identified 59 peptides containing single amino acid variants, which covered ~9% of all total coding nonsynonymous single nucleotide variants detected by whole-genome sequencing. Furthermore, we identified 29 distinct peptides that mapped to a hypothetical protein not present in the M. tuberculosis H37Rv reference proteome. Here we provide evidence for the expression of this protein in the clinical M. tuberculosis SAWC3651 isolate. The strain-specific databases enabled confirmation of genomic differences (i.e. large genomic regions of difference and nonsynonymous single nucleotide variants) in these two clinical M. tuberculosis isolates and allowed strain differentiation at the proteome level. Our results contribute to the growing field of clinical microbial proteogenomics and can improve our understanding of phenotypic variation in clinical M. tuberculosis isolates.

Proteogenomic Investigation of Strain Variation in Clinical Mycobacterium tuberculosis Isolates

KAUST Repository

Heunis, Tiaan; Dippenaar, Anzaan; Warren, Robin M.; van Helden, Paul D.; van der Merwe, Ruben G.; Gey van Pittius, Nicolaas C.; Pain, Arnab; Sampson, Samantha L.; Tabb, David L.

2017-01-01

Mycobacterium tuberculosis consists of a large number of different strains that display unique virulence characteristics. Whole-genome sequencing has revealed substantial genetic diversity among clinical M. tuberculosis isolates, and elucidating the phenotypic variation encoded by this genetic diversity will be of utmost importance to fully understand M. tuberculosis biology and pathogenicity. In this study we integrated whole-genome sequencing and mass spectrometry (GeLC-MS/MS) to reveal strain-specific characteristics in the proteomes of two clinical M. tuberculosis Latin American-Mediterranean isolates. Using this approach we identified 59 peptides containing single amino acid variants, which covered ~9% of all total coding nonsynonymous single nucleotide variants detected by whole-genome sequencing. Furthermore, we identified 29 distinct peptides that mapped to a hypothetical protein not present in the M. tuberculosis H37Rv reference proteome. Here we provide evidence for the expression of this protein in the clinical M. tuberculosis SAWC3651 isolate. The strain-specific databases enabled confirmation of genomic differences (i.e. large genomic regions of difference and nonsynonymous single nucleotide variants) in these two clinical M. tuberculosis isolates and allowed strain differentiation at the proteome level. Our results contribute to the growing field of clinical microbial proteogenomics and can improve our understanding of phenotypic variation in clinical M. tuberculosis isolates.

Proteogenomic Investigation of Strain Variation in Clinical Mycobacterium tuberculosis Isolates.

Science.gov (United States)

Heunis, Tiaan; Dippenaar, Anzaan; Warren, Robin M; van Helden, Paul D; van der Merwe, Ruben G; Gey van Pittius, Nicolaas C; Pain, Arnab; Sampson, Samantha L; Tabb, David L

2017-10-06

Mycobacterium tuberculosis consists of a large number of different strains that display unique virulence characteristics. Whole-genome sequencing has revealed substantial genetic diversity among clinical M. tuberculosis isolates, and elucidating the phenotypic variation encoded by this genetic diversity will be of the utmost importance to fully understand M. tuberculosis biology and pathogenicity. In this study, we integrated whole-genome sequencing and mass spectrometry (GeLC-MS/MS) to reveal strain-specific characteristics in the proteomes of two clinical M. tuberculosis Latin American-Mediterranean isolates. Using this approach, we identified 59 peptides containing single amino acid variants, which covered ∼9% of all coding nonsynonymous single nucleotide variants detected by whole-genome sequencing. Furthermore, we identified 29 distinct peptides that mapped to a hypothetical protein not present in the M. tuberculosis H37Rv reference proteome. Here, we provide evidence for the expression of this protein in the clinical M. tuberculosis SAWC3651 isolate. The strain-specific databases enabled confirmation of genomic differences (i.e., large genomic regions of difference and nonsynonymous single nucleotide variants) in these two clinical M. tuberculosis isolates and allowed strain differentiation at the proteome level. Our results contribute to the growing field of clinical microbial proteogenomics and can improve our understanding of phenotypic variation in clinical M. tuberculosis isolates.

Genetic Determinants of Drug Resistance in Mycobacterium tuberculosis and Their Diagnostic Value

NARCIS (Netherlands)

Farhat, M.R.; Sultana, R.; Iartchouk, O.; Bozeman, S.; Galagan, J.; Sisk, P.; Stolte, C.; Nebenzahl-Guimaraes, H.; Jacobson, K.; Sloutsky, A.; Kaur, D.; Posey, J.; Kreiswirth, B.N.; Kurepina, N.; Rigouts, L.; Streicher, E.M.; Victor, T.C.; Warren, R.M.; Soolingen, D. van; Murray, M.

2016-01-01

RATIONALE: The development of molecular diagnostics that detect both the presence of Mycobacterium tuberculosis in clinical samples and drug resistance-conferring mutations promises to revolutionize patient care and interrupt transmission by ensuring early diagnosis. However, these tools require the

Dominant incidence of multidrug and extensively drug-resistant specific Mycobacterium tuberculosis clones in Osaka Prefecture, Japan.

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Aki Tamaru

Full Text Available Infection and transmission of multidrug-resistant Mycobacterium tuberculosis (MDR-Mtb and extensively drug-resistant M. tuberculosis (XDR-Mtb is a serious health problem. We analyzed a total of 1,110 Mtb isolates in Osaka Prefecture and neighboring areas from April 2000 to March 2009. A total of 89 MDR-Mtb were identified, 36 (48.5% of which were determined to be XDR-Mtb. Among the 89 MDR-Mtb isolates, 24 (27.0% phylogenetically distributed into six clusters based on mycobacterial interspersed repetitive units-various number of tandem repeats (MIRU-VNTR typing. Among these six clusters, the MIRU-VNTR patterns of four (OM-V02, OM-V03, OM-V04, and OM-V06 were only found for MDR-Mtb. Further analysis revealed that all isolates belonging to OM-V02 and OM-V03, and two isolates from OM-V04 were clonal. Importantly such genotypes were not observed for drug-sensitive isolates. These suggest that few but transmissible clones can transmit after acquiring multidrug resistance and colonize even in a country with a developed, well-organized healthcare system.

Clinical implications of molecular drug resistance testing for Mycobacterium tuberculosis: a TBNET/RESIST-TB consensus statement

NARCIS (Netherlands)

Domínguez, J.; Boettger, E. C.; Cirillo, D.; Cobelens, F.; Eisenach, K. D.; Gagneux, S.; Hillemann, D.; Horsburgh, R.; Molina-Moya, B.; Niemann, S.; Tortoli, E.; Whitelaw, A.; Lange, C.

2016-01-01

The emergence of drug-resistant strains of Mycobacterium tuberculosis is a challenge to global tuberculosis (TB) control. Although culture-based methods have been regarded as the gold standard for drug susceptibility testing (DST), molecular methods provide rapid information on mutations in the M.

Studium role fosfofruktokinasy A a B v metabolismu Mycobacterium tuberculosis

Czech Academy of Sciences Publication Activity Database

Machová, Iva; Snášel, Jan; Pichová, Iva

2014-01-01

Roč. 108, č. 5 (2014), s. 542 ISSN 0009-2770. [Mezioborové setkání mladých biologů, biochemiků a chemiků /14./. 13.05.2014-16.05.2014, Milovy] Grant - others:European Research Council(XE) FP7-245187 Institutional support: RVO:61388963 Keywords : Mycobacterium tuberculosis * phosphofructokinase Subject RIV: CE - Biochemistry

In vitro Inhibition of Mycobacterium smegmatis and Mycobacterium ...

African Journals Online (AJOL)

Some Nigerian plants used in traditional medicine to treat tuberculosis and/or some of its symptoms were screened for in vitro activity against Mycobacterium smegmatis and a clinical isolate of Mycobacterium tuberculosis. Only 3 of the 6 crude methanolic extracts of the 6 plant species exhibited inhibitory activities against ...

Immunogenic Properties of Lactobacillus plantarum Producing Surface-Displayed Mycobacterium tuberculosis Antigens.

Science.gov (United States)

Kuczkowska, Katarzyna; Kleiveland, Charlotte R; Minic, Rajna; Moen, Lars F; Øverland, Lise; Tjåland, Rannei; Carlsen, Harald; Lea, Tor; Mathiesen, Geir; Eijsink, Vincent G H

2017-01-15

Tuberculosis (TB) remains among the most deadly diseases in the world. The only available vaccine against tuberculosis is the bacille Calmette-Guérin (BCG) vaccine, which does not ensure full protection in adults. There is a global urgency for the development of an effective vaccine for preventing disease transmission, and it requires novel approaches. We are exploring the use of lactic acid bacteria (LAB) as a vector for antigen delivery to mucosal sites. Here, we demonstrate the successful expression and surface display of a Mycobacterium tuberculosis fusion antigen (comprising Ag85B and ESAT-6, referred to as AgE6) on Lactobacillus plantarum The AgE6 fusion antigen was targeted to the bacterial surface using two different anchors, a lipoprotein anchor directing the protein to the cell membrane and a covalent cell wall anchor. AgE6-producing L. plantarum strains using each of the two anchors induced antigen-specific proliferative responses in lymphocytes purified from TB-positive donors. Similarly, both strains induced immune responses in mice after nasal or oral immunization. The impact of the anchoring strategies was reflected in dissimilarities in the immune responses generated by the two L. plantarum strains in vivo The present study comprises an initial step toward the development of L. plantarum as a vector for M. tuberculosis antigen delivery. This work presents the development of Lactobacillus plantarum as a candidate mucosal vaccine against tuberculosis. Tuberculosis remains one of the top infectious diseases worldwide, and the only available vaccine, bacille Calmette-Guérin (BCG), fails to protect adults and adolescents. Direct antigen delivery to mucosal sites is a promising strategy in tuberculosis vaccine development, and lactic acid bacteria potentially provide easy, safe, and low-cost delivery vehicles for mucosal immunization. We have engineered L. plantarum strains to produce a Mycobacterium tuberculosis fusion antigen and to anchor this

Mycobacterium tuberculosis Controls Phagosomal Acidification by Targeting CISH-Mediated Signaling

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Christophe J. Queval

2017-09-01

Full Text Available Pathogens have evolved a range of mechanisms to counteract host defenses, notably to survive harsh acidic conditions in phagosomes. In the case of Mycobacterium tuberculosis, it has been shown that regulation of phagosome acidification could be achieved by interfering with the retention of the V-ATPase complexes at the vacuole. Here, we present evidence that M. tuberculosis resorts to yet another strategy to control phagosomal acidification, interfering with host suppressor of cytokine signaling (SOCS protein functions. More precisely, we show that infection of macrophages with M. tuberculosis leads to granulocyte-macrophage colony-stimulating factor (GM-CSF secretion, inducing STAT5-mediated expression of cytokine-inducible SH2-containing protein (CISH, which selectively targets the V-ATPase catalytic subunit A for ubiquitination and degradation by the proteasome. Consistently, we show that inhibition of CISH expression leads to reduced replication of M. tuberculosis in macrophages. Our findings further broaden the molecular understanding of mechanisms deployed by bacteria to survive.

Evaluation of highly conserved hsp65-specific nested PCR primers for diagnosing Mycobacterium tuberculosis.

Science.gov (United States)

Priyadarshini, P; Tiwari, K; Das, A; Kumar, D; Mishra, M N; Desikan, P; Nath, G

2017-02-01

To evaluate the sensitivity and specificity of a new nested set of primers designed for the detection of Mycobacterium tuberculosis complex targeting a highly conserved heat shock protein gene (hsp65). The nested primers were designed using multiple sequence alignment assuming the nucleotide sequence of the M. tuberculosis H37Rv hsp65 genome as base. Multidrug-resistant Mycobacterium species along with other non-mycobacterial and fungal species were included to evaluate the specificity of M. tuberculosis hsp65 gene-specific primers. The sensitivity of the primers was determined using serial 10-fold dilutions, and was 100% as shown by the bands in the case of M. tuberculosis complex. None of the other non M. tuberculosis complex bacterial and fungal species yielded any band on nested polymerase chain reaction (PCR). The first round of amplification could amplify 0.3 ng of the template DNA, while nested PCR could detect 0.3 pg. The present hsp65-specific primers have been observed to be sensitive, specific and cost-effective, without requiring interpretation of biochemical tests, real-time PCR, sequencing or high-performance liquid chromatography. These primer sets do not have the drawbacks associated with those protocols that target insertion sequence 6110, 16S rDNA, rpoB, recA and MPT 64.

Current Methods in the Molecular Typing of Mycobacterium tuberculosis and Other Mycobacteria

Science.gov (United States)

van Ingen, Jakko; Dziadek, Jarosław; Mazur, Paweł K.; Bielecki, Jacek

2014-01-01

In the epidemiology of tuberculosis (TB) and nontuberculous mycobacterial (NTM) diseases, as in all infectious diseases, the key issue is to define the source of infection and to disclose its routes of transmission and dissemination in the environment. For this to be accomplished, the ability of discerning and tracking individual Mycobacterium strains is of critical importance. Molecular typing methods have greatly improved our understanding of the biology of mycobacteria and provide powerful tools to combat the diseases caused by these pathogens. The utility of various typing methods depends on the Mycobacterium species under investigation as well as on the research question. For tuberculosis, different methods have different roles in phylogenetic analyses and person-to-person transmission studies. In NTM diseases, most investigations involve the search for environmental sources or phylogenetic relationships. Here, too, the type of setting determines which methodology is most suitable. Within this review, we summarize currently available molecular methods for strain typing of M. tuberculosis and some NTM species, most commonly associated with human disease. For the various methods, technical practicalities as well as discriminatory power and accomplishments are reviewed. PMID:24527454

Differential T-cell recognition of native and recombinant Mycobacterium tuberculosis GroES

DEFF Research Database (Denmark)

Rosenkrands, I; Weldingh, K; Ravn, P

1999-01-01

Mycobacterium tuberculosis GroES was purified from culture filtrate, and its identity was confirmed by immunoblot analysis and N-terminal sequencing. Comparing the immunological recognition of native and recombinant GroES, we found that whereas native GroES elicited a strong proliferative response...

The curative activity of thioridazine on mice infected with Mycobacterium tuberculosis

DEFF Research Database (Denmark)

Martins, Marta; Viveiros, Miguel; Kristiansen, Jette E

2007-01-01

BACKGROUND: The aim of the study was to evaluate the effectiveness of thioridazine (TZ) at different dose levels on mice that had been infected intraperitoneally (i.p.) with a high dose of the Mycobacterium tuberculosis ATCC H37Rv strain. SUBJECTS AND METHODS: Groups of five female BALB/C mice were...

Total Hip Arthroplasty Loosening Due to Mycobacterium Tuberculosis: A Case Report and Review of the Literature

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Anis Tebourbi

2017-04-01

Full Text Available Context: Prosthetic joint infection due to Mycobacterium tuberculosis with no previous history of pulmonary or extra pulmonary tuberculosis is an extremely rare complication. Aims To report the case of a patient with tuberculous mycobacterial prosthetic hip infection, 14 years after surgery for post traumatic osteoarthritis, with no previous history of tuberculosis. Methods A 46-year-old male presented an acetabular loosening of a cemented total hip arthroplasty with subnormal biologic parameters. A one stage revision surgery was planned. Intraoperative findings suggested mycobacterial tuberculous infection with presence of periacetabular yellowish rice-shaped granules. Results A one-stage prosthesis exchange was performed; Culture on Löwenstein-Jensen medium grew MTB days after inoculation and histological examination confirmed tuberculous infection. Patient was treated by antituberculous agents for 12 months with optimal clinical and biological response and no prosthetic loosening signs at eighteen months follow up. Conclusions Total hip arthroplasty loosening due to mycobacterium tuberculosis is a rare entity, which should be evoked even when no inflammatory signs are shown. Discovery of yellowish rice-shaped granules is an indicator to investigate for tuberculosis. Management of prosthetic joint infection due to M.tuberculosis must involve both medical and surgical approach.

Extracellular Sphingomyelinase Rv0888 of Mycobacterium tuberculosis Contributes to Pathological Lung Injury of Mycobacterium smegmatis in Mice via Inducing Formation of Neutrophil Extracellular Traps.

Science.gov (United States)

Dang, Guanghui; Cui, Yingying; Wang, Lei; Li, Tiantian; Cui, Ziyin; Song, Ningning; Chen, Liping; Pang, Hai; Liu, Siguo

2018-01-01

Mycobacterium tuberculosis is the causative agent of tuberculosis (TB), which mainly causes pulmonary injury and tubercles. Although macrophages are generally considered to harbor the main cells of M. tuberculosis , new evidence suggests that neutrophils are rapidly recruited to the infected lung. M. tuberculosis itself, or its early secreted antigenic target protein 6 (ESAT-6), can induce formation of neutrophil extracellular traps (NETs). However, NETs trap mycobacteria but are unable to kill them. The role of NETs' formation in the pathogenesis of mycobacteria remains unclear. Here, we report a new M. tuberculosis extracellular factor, bifunctional enzyme Rv0888, with both nuclease and sphingomyelinase activities. Rv0888 sphingomyelinase activity can induce NETs' formation in vitro and in the lung of the mice and enhance the colonization ability of Mycobacterium smegmatis in the lungs of mice. Mice infected by M. smegmatis harboring Rv0888 sphingomyelinase induced pathological injury and inflammation of the lung, which was mainly mediated by NETs, induced by Rv0888 sphingomyelinase, associated protein (myeloperoxidase) triggered caspase-3. In summary, the study sheds new light on the pathogenesis of mycobacteria and reveals a novel target for TB treatment.

Mycobacterium tuberculosis TlyA Protein Negatively Regulates T Helper (Th) 1 and Th17 Differentiation and Promotes Tuberculosis Pathogenesis*

Science.gov (United States)

Rahman, Md. Aejazur; Sobia, Parveen; Dwivedi, Ved Prakash; Bhawsar, Aakansha; Singh, Dhiraj Kumar; Sharma, Pawan; Moodley, Prashini; Van Kaer, Luc; Bishai, William R; Das, Gobardhan

2015-01-01

Mycobacterium tuberculosis, the causative agent of tuberculosis, is an ancient pathogen and a major cause of death worldwide. Although various virulence factors of M. tuberculosis have been identified, its pathogenesis remains incompletely understood. TlyA is a virulence factor in several bacterial infections and is evolutionarily conserved in many Gram-positive bacteria, but its function in M. tuberculosis pathogenesis has not been elucidated. Here, we report that TlyA significantly contributes to the pathogenesis of M. tuberculosis. We show that a TlyA mutant M. tuberculosis strain induces increased IL-12 and reduced IL-1β and IL-10 cytokine responses, which sharply contrasts with the immune responses induced by wild type M. tuberculosis. Furthermore, compared with wild type M. tuberculosis, TlyA-deficient M. tuberculosis bacteria are more susceptible to autophagy in macrophages. Consequently, animals infected with the TlyA mutant M. tuberculosis organisms exhibited increased host-protective immune responses, reduced bacillary load, and increased survival compared with animals infected with wild type M. tuberculosis. Thus, M. tuberculosis employs TlyA as a host evasion factor, thereby contributing to its virulence. PMID:25847237

DETECTION OF MYCOBACTERIUM TUBERCULOSIS IN BLOOD FOR DIAGNOSIS OF GENERALISED TUBERCULOSIS IN HIV-POSITIVE PATIENTS

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V. N. Zimina

2017-01-01

Full Text Available Objective: To study the informative value of the detection of mycobacteria in blood with the cultural method in patients with suspected tuberculous sepsis and to determine the most significant clinical and laboratory criteria for testing. Materials and methods: The investigation to detect M.tuberculosis was fulfilled in 159 HIV-positive patients with suspected tuberculosis sepsis. Blood culture was completed with culture medium Myco/F Lytic Culture Vials and analyzer BACTEC 9050. Results: Mycobacteria were detected in blood of 19 patients (11,9% of all patients: in 18 patients the growth of М. tuberculosis complex was detected (25,3% of all patients with diagnosed tuberculosis and in 1 patient it was Mycobacterium avium complex (0,6% of all patients. It was shown, that the probability of M.tuberculosis detection was especially associated with the severity of the disease, immunosupression (less than 100 cells/mkl, hemoglobin quantity less than 90 g/l (levels were determined through the seeking for the most significant cutoffs. It was not proofed, that meningoencephalitis develops more often in patients with proven bacteremia. There were no evident differences in detection frequency of mycobacteria in sputum between patients with tuberculous sepsis and without it.

MOLECULAR CHARACTERIZATION OF MYCOBACTERIUM TUBERCULOSIS STRAINS CIRCULATING IN THE URAL REGION, RUSSIA

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T. V. Umpeleva

2013-01-01

Full Text Available Abstract. Overall 178 Mycobacterium tuberculosis isolates recovered in 2009–2011 from newly diagnosed epidemiologically unlinked to TB patients in the Ural region of Russia have been studied. The absolute concentration method was used for drug susceptibility testing. Mutations katG, inhA and rpoB associated with resistance to isoniazid and rifampicin were detected by microchip technology («TB-Biochip». The isolates were genotyped by real-time PCR for the detection of Beijing/non-Beijing genotypes and 15-locus MIRU-VNTR typing according to «MIRU-VNTRplus» (http://www.miru-vntrplus.org. More than half (55.1% of 178 isolates belonged to the Beijing family, 58.7% of them were multiple drug resistant (MDR mostly due to rpoBSer531→Leu and katGSer315→Thr1 substitutions. Fifty VNTR profiles were found in 98 Beijing isolates; 57 of them grouped into 9 clusters. The largest VNTR cluster included 23 (23.5% Beijing isolates and 21 of them were MDR. The 80 non-Beijing isolates showed 64 distinct VNTR patterns which belonged to 6 genetic families: LAM, Ural, Haarlem, etc. Among LAM and Ural isolates 30.4% and 28.6% were MDR, respectively. The 5 of 7 MDR LAM isolates had specific mutation profile:  rpoBAsp516→Val substitution and mutations katGSer315→Thr1 and inhA_T15. The MDR Ural isolates showed the heterogeneity of mutations in rpoB gene compared to other genotypes. Taken together, these findings suggest the emergence and spread of MDR-TB in the Ural region which is characterized by circulation of MDR strains of different genotypes with the Beijing family genotype to be predominant.

Two-Component Systems of Mycobacterium tuberculosis—Structure-Based Approaches

DEFF Research Database (Denmark)

Tucker, Paul; Nowak, Elzbieta; Morth, Jens Preben

2007-01-01

Mycobacterium tuberculosis contains few two‐component systems compared to many other bacteria, possibly because it has more serine/threonine signaling pathways. Even so, these two‐component systems appear to play an important role in early intracellular survival of the pathogen as well as in aspe...

Genomic diversity of drug-resistant Mycobacterium tuberculosis isolates in Lisbon Portugal: Towards tuberculosis genomic epidemiology

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João Perdigão

2015-01-01

Full Text Available Multidrug- (MDR and extensively drug-resistant (XDR tuberculosis (TB present a challenge to disease control and elimination goals. Lisbon, Portugal, has a high TB incidencerate and unusual and successful XDR-TB strains that have been found in circulation foralmost two decades. For the last 20 years, a continued circulation of two phylogenetic clades, Lisboa3 and Q1, which are highly associated with MDR and XDR, have been observed. In recent years, these strains have been well characterized regarding the molecular basis of drug resistance and have been inclusively subjected to whole genome sequencing (WGS. Researchers have been studying the genomic diversity of strains circulating in Lisbon and its genomic determinants through cutting-edge next generation sequencing. An enormous amount of whole genome sequence data are now available for the most prevalent and clinically relevant strains circulating in Lisbon. It is the persistence, prevalence and rapid evolution towards drug resistance that has prompted researchers to investigate the properties of these strains at the genomic level and in the future at a global transcriptomic level. Seventy Mycobacterium tuberculosis (MTB isolates, mostly recovered in Lisbon, were genotyped by 24-loci Mycobacterial Interspersed Repetitive Unit – Variable Number of Tandem Repeats (MIRU-VNTR and the genomes sequenced using a next generation sequencing platform – Illumina HiSeq 2000. The genotyping data revealed three major clusters associated with MDR-TB (Lisboa3-A, Lisboa3-B and Q1, two of which are associated with XDR-TB (Lisboa3-B and Q1, whilst the genomic data contributed to elucidating the phylogenetic positioning of circulating MDR-TB strains, showing a high predominance of a single SNP cluster group 5. Furthermore, a genome-wide phylogeny analysis from these strains, together with 19 publicly available genomes of MTB clinical isolates, revealed two major clades responsible for MDR/XDR-TB in the region

Genomic diversity of drug-resistant Mycobacterium tuberculosis isolates in Lisbon Portugal: Towards tuberculosis genomic epidemiology

KAUST Repository

Perdigã o, Joã o; Silva, Hugo; Machado, Diana; Macedo, Rita; Maltez, Fernando; Silva, Carla; Jordao, Luisa; Couto, Isabel; Mallard, Kim; Coll, Francesc; Hill-Cawthorne, Grant A.; McNerney, Ruth; Pain, Arnab; Clark, Taane G.; Viveiros, Miguel; Portugal, Isabel

2015-01-01

Multidrug- (MDR) and extensively drug-resistant (XDR) tuberculosis (TB) present a challenge to disease control and elimination goals. Lisbon, Portugal, has a high TB incidence rate and unusual and successful XDR-TB strains that have been found in circulation for almost two decades. For the last 20. years, a continued circulation of two phylogenetic clades, Lisboa3 and Q1, which are highly associated with MDR and XDR, have been observed. In recent years, these strains have been well characterized regarding the molecular basis of drug resistance and have been inclusively subjected to whole genome sequencing (WGS). Researchers have been studying the genomic diversity of strains circulating in Lisbon and its genomic determinants through cutting-edge next generation sequencing. An enormous amount of whole genome sequence data are now available for the most prevalent and clinically relevant strains circulating in Lisbon.It is the persistence, prevalence and rapid evolution towards drug resistance that has prompted researchers to investigate the properties of these strains at the genomic level and in the future at a global transcriptomic level. Seventy Mycobacterium tuberculosis (MTB) isolates, mostly recovered in Lisbon, were genotyped by 24-. loci Mycobacterial Interspersed Repetitive Unit - Variable Number of Tandem Repeats (MIRU-VNTR) and the genomes sequenced using a next generation sequencing platform - Illumina HiSeq 2000.The genotyping data revealed three major clusters associated with MDR-TB (Lisboa3-A, Lisboa3-B and Q1), two of which are associated with XDR-TB (Lisboa3-B and Q1), whilst the genomic data contributed to elucidating the phylogenetic positioning of circulating MDR-TB strains, showing a high predominance of a single SNP cluster group 5. Furthermore, a genome-wide phylogeny analysis from these strains, together with 19 publicly available genomes of MTB clinical isolates, revealed two major clades responsible for MDR/XDR-TB in the region: Lisboa3 and Q

Genomic diversity of drug-resistant Mycobacterium tuberculosis isolates in Lisbon Portugal: Towards tuberculosis genomic epidemiology

KAUST Repository

Perdigão, João

2015-03-01

Multidrug- (MDR) and extensively drug-resistant (XDR) tuberculosis (TB) present a challenge to disease control and elimination goals. Lisbon, Portugal, has a high TB incidence rate and unusual and successful XDR-TB strains that have been found in circulation for almost two decades. For the last 20. years, a continued circulation of two phylogenetic clades, Lisboa3 and Q1, which are highly associated with MDR and XDR, have been observed. In recent years, these strains have been well characterized regarding the molecular basis of drug resistance and have been inclusively subjected to whole genome sequencing (WGS). Researchers have been studying the genomic diversity of strains circulating in Lisbon and its genomic determinants through cutting-edge next generation sequencing. An enormous amount of whole genome sequence data are now available for the most prevalent and clinically relevant strains circulating in Lisbon.It is the persistence, prevalence and rapid evolution towards drug resistance that has prompted researchers to investigate the properties of these strains at the genomic level and in the future at a global transcriptomic level. Seventy Mycobacterium tuberculosis (MTB) isolates, mostly recovered in Lisbon, were genotyped by 24-. loci Mycobacterial Interspersed Repetitive Unit - Variable Number of Tandem Repeats (MIRU-VNTR) and the genomes sequenced using a next generation sequencing platform - Illumina HiSeq 2000.The genotyping data revealed three major clusters associated with MDR-TB (Lisboa3-A, Lisboa3-B and Q1), two of which are associated with XDR-TB (Lisboa3-B and Q1), whilst the genomic data contributed to elucidating the phylogenetic positioning of circulating MDR-TB strains, showing a high predominance of a single SNP cluster group 5. Furthermore, a genome-wide phylogeny analysis from these strains, together with 19 publicly available genomes of MTB clinical isolates, revealed two major clades responsible for MDR/XDR-TB in the region: Lisboa3 and Q

Cyclic GMP-AMP Synthase Is an Innate Immune DNA Sensor for Mycobacterium tuberculosis.

Science.gov (United States)

Collins, Angela C; Cai, Haocheng; Li, Tuo; Franco, Luis H; Li, Xiao-Dong; Nair, Vidhya R; Scharn, Caitlyn R; Stamm, Chelsea E; Levine, Beth; Chen, Zhijian J; Shiloh, Michael U

2015-06-10

Activation of the DNA-dependent cytosolic surveillance pathway in response to Mycobacterium tuberculosis infection stimulates ubiquitin-dependent autophagy and inflammatory cytokine production, and plays an important role in host defense against M. tuberculosis. However, the identity of the host sensor for M. tuberculosis DNA is unknown. Here we show that M. tuberculosis activated cyclic guanosine monophosphate-adenosine monophosphate (cGAMP) synthase (cGAS) in macrophages to produce cGAMP, a second messenger that activates the adaptor protein stimulator of interferon genes (STING) to induce type I interferons and other cytokines. cGAS localized with M. tuberculosis in mouse and human cells and in human tuberculosis lesions. Knockdown or knockout of cGAS in human or mouse macrophages blocked cytokine production and induction of autophagy. Mice deficient in cGAS were more susceptible to lethality caused by infection with M. tuberculosis. These results demonstrate that cGAS is a vital innate immune sensor of M. tuberculosis infection. Copyright © 2015 Elsevier Inc. All rights reserved.

Origins and properties of Mycobacterium tuberculosis isolates in London.

Science.gov (United States)

Dale, Jeremy W; Bothamley, Graham H; Drobniewski, Francis; Gillespie, Stephen H; McHugh, Timothy D; Pitman, Richard

2005-06-01

Using similarities of IS6110 banding patterns, isolates of Mycobacterium tuberculosis from a population-based study in London were assigned to 12 large groups termed 'superfamilies' (sfams). Analysis of patient data showed a marked geographical association in the distribution of these sfams. In particular, isolates from patients born in Europe were from different sfams than those born elsewhere, indicating that there had been relatively little transmission of tuberculosis in London from immigrant communities into the endogenous population. Multivariate analysis showed that certain sfams were significantly associated with pulmonary rather than extrapulmonary disease, or with sputum smear negativity, independently of country of birth or ethnicity, suggesting that the properties of the infecting organism play a role in the nature of the disease process.

Substituted aminopyrimidine protein kinase B (PknB) inhibitors show activity against Mycobacterium tuberculosis

Science.gov (United States)

Chapman, Timothy M.; Bouloc, Nathalie; Buxton, Roger S.; Chugh, Jasveen; Lougheed, Kathryn E.A.; Osborne, Simon A.; Saxty, Barbara; Smerdon, Stephen J.; Taylor, Debra L.; Whalley, David

2012-01-01

A high-throughput screen against PknB, an essential serine–threonine protein kinase present in Mycobacterium tuberculosis (M. tuberculosis), allowed the identification of an aminoquinazoline inhibitor which was used as a starting point for SAR investigations. Although a significant improvement in enzyme affinity was achieved, the aminoquinazolines showed little or no cellular activity against M. tuberculosis. However, switching to an aminopyrimidine core scaffold and the introduction of a basic amine side chain afforded compounds with nanomolar enzyme binding affinity and micromolar minimum inhibitory concentrations against M. tuberculosis. Replacement of the pyrazole head group with pyridine then allowed equipotent compounds with improved selectivity against a human kinase panel to be obtained. PMID:22469702

Urease Activity Represents an Alternative Pathway for Mycobacterium tuberculosis Nitrogen Metabolism

Science.gov (United States)

Lin, Wenwei; Mathys, Vanessa; Ang, Emily Lei Yin; Koh, Vanessa Hui Qi; Martínez Gómez, Julia María; Ang, Michelle Lay Teng; Zainul Rahim, Siti Zarina; Tan, Mai Ping; Pethe, Kevin

2012-01-01

Urease represents a critical virulence factor for some bacterial species through its alkalizing effect, which helps neutralize the acidic microenvironment of the pathogen. In addition, urease serves as a nitrogen source provider for bacterial growth. Pathogenic mycobacteria express a functional urease, but its role during infection has yet to be characterized. In this study, we constructed a urease-deficient Mycobacterium tuberculosis strain and confirmed the alkalizing effect of the urease activity within the mycobacterium-containing vacuole in resting macrophages but not in the more acidic phagolysosomal compartment of activated macrophages. However, the urease-mediated alkalizing effect did not confer any growth advantage on M. tuberculosis in macrophages, as evidenced by comparable growth profiles for the mutant, wild-type (WT), and complemented strains. In contrast, the urease-deficient mutant exhibited impaired in vitro growth compared to the WT and complemented strains when urea was the sole source of nitrogen. Substantial amounts of ammonia were produced by the WT and complemented strains, but not with the urease-deficient mutant, which represents the actual nitrogen source for mycobacterial growth. However, the urease-deficient mutant displayed parental colonization profiles in the lungs, spleen, and liver in mice. Together, our data demonstrate a role for the urease activity in M. tuberculosis nitrogen metabolism that could be crucial for the pathogen's survival in nutrient-limited microenvironments where urea is the sole nitrogen source. Our work supports the notion that M. tuberculosis virulence correlates with its unique metabolic versatility and ability to utilize virtually any carbon and nitrogen sources available in its environment. PMID:22645285

Prevalence of latent Mycobacterium tuberculosis infection in prisoners.

Science.gov (United States)

Navarro, Pedro Daibert de; Almeida, Isabela Neves de; Kritski, Afrânio Lineu; Ceccato, Maria das Graças; Maciel, Mônica Maria Delgado; Carvalho, Wânia da Silva; Miranda, Silvana Spindola de

2016-01-01

To determine the prevalence of and the factors associated with latent Mycobacterium tuberculosis infection (LTBI) in prisoners in the state of Minas Gerais, Brazil. This was a cross-sectional cohort study conducted in two prisons in Minas Gerais. Tuberculin skin tests were performed in the individuals who agreed to participate in the study. A total of 1,120 individuals were selected for inclusion in this study. The prevalence of LTBI was 25.2%. In the multivariate analysis, LTBI was associated with self-reported contact with active tuberculosis patients within prisons (adjusted OR = 1.51; 95% CI: 1.05-2.18) and use of inhaled drugs (adjusted OR = 1.48; 95% CI: 1.03-2.13). Respiratory symptoms were identified in 131 (11.7%) of the participants. Serological testing for HIV was performed in 940 (83.9%) of the participants, and the result was positive in 5 (0.5%). Two cases of active tuberculosis were identified during the study period. Within the prisons under study, the prevalence of LTBI was high. In addition, LTBI was associated with self-reported contact with active tuberculosis patients and with the use of inhaled drugs. Our findings demonstrate that it is necessary to improve the conditions in prisons, as well as to introduce strategies, such as chest X-ray screening, in order to detect tuberculosis cases and, consequently, reduce M. tuberculosis infection within the prison system. Determinar a prevalência e os fatores associados à infecção latente por Mycobacterium tuberculosis (ILTB) em pessoas privadas de liberdade no Estado de Minas Gerais. Estudo de coorte transversal realizado em duas penitenciárias em Minas Gerais. Foi realizada a prova tuberculínica nos indivíduos que aceitaram participar do estudo. Foram selecionados 1.120 indivíduos para a pesquisa. A prevalência da ILTB foi de 25,2%. Na análise multivariada, a ILTB esteve associada com relato de contato com caso de tuberculose ativa dentro da penitenciária (OR ajustada = 1,51; IC95%: 1

CCL2 responses to Mycobacterium tuberculosis are associated with disease severity in tuberculosis.

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Zahra Hasan

Full Text Available BACKGROUND: Leucocyte activating chemokines such as CCL2, CCL3, and CXCL8 together with proinflammatory IFNgamma, TNFalpha and downmodulatory IL10 play a central role in the restriction of M. tuberculosis infections, but is unclear whether these markers are indicative of tuberculosis disease severity. METHODOLOGY: We investigated live M. tuberculosis- and M. bovis BCG-induced peripheral blood mononuclear cell responses in patients with tuberculosis (TB and healthy endemic controls (ECs, n = 36. TB patients comprised pulmonary (PTB, n = 34 and extrapulmonary groups, subdivided into those with less severe localized extrapulmonary TB (L-ETB, n = 16 or severe disseminated ETB (D-ETB, n = 16. Secretion of CCL2, IFNgamma, IL10 and CCL3, and mRNA expression of CCL2, TNFalpha, CCL3 and CXCL8 were determined. RESULTS: M. tuberculosis- and BCG-induced CCL2 secretion was significantly increased in both PTB and D-ETB (p<0.05, p<0.01 as compared with L-ETB patients. CCL2 secretion in response to M. tuberculosis was significantly greater than to BCG in the PTB and D-ETB groups. M. tuberculosis-induced CCL2 mRNA transcription was greater in PTB than L-ETB (p = 0.023, while CCL2 was reduced in L-ETB as compared with D-ETB (p = 0.005 patients. M. tuberculosis-induced IFNgamma was greater in L-ETB than PTB (p = 0.04, while BCG-induced IFNgamma was greater in L-ETB as compared with D-ETB patients (p = 0.036. TNFalpha mRNA expression was raised in PTB as compared with L-ETB group in response to M. tuberculosis (p = 0.02 and BCG (p = 0.03. Mycobacterium-induced CCL3 and CXCL8 was comparable between TB groups. CONCLUSIONS: The increased CCL2 and TNFalpha in PTB patients may support effective leucocyte recruitment and M. tuberculosis localization. CCL2 alone is associated with severity of TB, possibly due to increased systemic inflammation found in severe disseminated TB or due to increased monocyte infiltration to lung parenchyma in pulmonary disease.

Impact of Laboratory Cross-Contamination on Molecular Epidemiology Studies of Tuberculosis

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Martínez, Miguel; de Viedma, Darío García; Alonso, María; Andrés, Sandra; Bouza, Emilio; Cabezas, Teresa; Cabeza, Isabel; Reyes, Armando; Sánchez-Yebra, Waldo; Rodríguez, Manuel; Sánchez, M. Isabel; Rogado, M. Cruz; Fernández, Rosa; Peñafiel, Teresa; Martínez, Juan; Barroso, Pilar; Lucerna, M. Ángeles; Diez, L. Felipe; Gutiérrez, Carmelo

2006-01-01

Laboratory cross-contamination by Mycobacterium tuberculosis is known to be responsible for the misdiagnosis of tuberculosis, but its impact on other contexts has not been analyzed. We present the findings of a molecular epidemiology analysis in which the recent transmission events identified by a genotyping reference center were overestimated as a result of unnoticed laboratory cross-contamination in the original diagnostic laboratories. PMID:16891518

Bovine tuberculosis

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Tuberculosis (TB) in animals and humans may result from exposure to bacilli within the Mycobacterium tuberculosis complex (i.e., M. tuberculosis, M. bovis, M. africanum, M. pinnipedii, M. microti, M. caprae, or M. canetti) . Mycobacterium bovis is the species most often isolated from tuberculous cat...

Biochemická charakterizace fosfoenolpyruvát karboxykinasy z Mycobacterium tuberculosis

Czech Academy of Sciences Publication Activity Database

Machová, Iva; Snášel, Jan; Pichová, Iva

2013-01-01

Roč. 107, č. 5 (2013), s. 425-425 ISSN 0009-2770. [Mezioborové setkání mladých biologů, biochemiků a chemiků /13./. 14.5.2013-17.5.2013, Žďár nad Sázavou] Grant - others:European Research Council(XE) FP7-245187 Institutional support: RVO:61388963 Keywords : Mycobacterium tuberculosis * Pepck * GDP/GTP Subject RIV: CE - Biochemistry

Structural and Functional Studies of Phosphoenolpyruvate Carboxykinase from Mycobacterium tuberculosis

Czech Academy of Sciences Publication Activity Database

Machová, Iva; Snášel, Jan; Dostál, Jiří; Brynda, Jiří; Fanfrlík, Jindřich; Singh, M.; Tarábek, Ján; Vaněk, O.; Bednárová, Lucie; Pichová, Iva

2015-01-01

Roč. 10, č. 3 (2015), e0120682/1-e0120682/21 E-ISSN 1932-6203 R&D Projects: GA MŠk LO1302 EU Projects: European Commission(XE) 241587 - SYSTEMTB Institutional support: RVO:61388963 Keywords : crystal structure * noncovalent complexes * Mycobacterium tuberculosis * mechanism Subject RIV: CE - Biochemistry Impact factor: 3.057, year: 2015 http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0120682

Mycobacterium tuberculosis Controls Phagosomal Acidification by Targeting CISH-Mediated Signaling.

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Queval, Christophe J; Song, Ok-Ryul; Carralot, Jean-Philippe; Saliou, Jean-Michel; Bongiovanni, Antonino; Deloison, Gaspard; Deboosère, Nathalie; Jouny, Samuel; Iantomasi, Raffaella; Delorme, Vincent; Debrie, Anne-Sophie; Park, Sei-Jin; Gouveia, Joana Costa; Tomavo, Stanislas; Brosch, Roland; Yoshimura, Akihiko; Yeramian, Edouard; Brodin, Priscille

2017-09-26

Pathogens have evolved a range of mechanisms to counteract host defenses, notably to survive harsh acidic conditions in phagosomes. In the case of Mycobacterium tuberculosis, it has been shown that regulation of phagosome acidification could be achieved by interfering with the retention of the V-ATPase complexes at the vacuole. Here, we present evidence that M. tuberculosis resorts to yet another strategy to control phagosomal acidification, interfering with host suppressor of cytokine signaling (SOCS) protein functions. More precisely, we show that infection of macrophages with M. tuberculosis leads to granulocyte-macrophage colony-stimulating factor (GM-CSF) secretion, inducing STAT5-mediated expression of cytokine-inducible SH2-containing protein (CISH), which selectively targets the V-ATPase catalytic subunit A for ubiquitination and degradation by the proteasome. Consistently, we show that inhibition of CISH expression leads to reduced replication of M. tuberculosis in macrophages. Our findings further broaden the molecular understanding of mechanisms deployed by bacteria to survive. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

In vivo induction of neutrophil extracellular traps by Mycobacterium tuberculosis in a guinea pig model.

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Filio-Rodríguez, Georgina; Estrada-García, Iris; Arce-Paredes, Patricia; Moreno-Altamirano, María M; Islas-Trujillo, Sergio; Ponce-Regalado, M Dolores; Rojas-Espinosa, Oscar

2017-10-01

In 2004, a novel mechanism of cellular death, called 'NETosis', was described in neutrophils. This mechanism, different from necrosis and apoptosis, is characterized by the release of chromatin webs admixed with microbicidal granular proteins and peptides (NETs). NETs trap and kill a variety of microorganisms. Diverse microorganisms, including Mycobacterium tuberculosis, are NET inducers in vitro. The aim of this study was to examine whether M. tuberculosis can also induce NETs in vivo and if the NETs are bactericidal to the microorganism. Guinea pigs were intradermally inoculated with M. tuberculosis H37Rv, and the production of NETs was investigated at several time points thereafter. NETs were detected as early as 30 min post-inoculation and were clearly evident by 4 h post-inoculation. NETs produced in vivo contained DNA, myeloperoxidase, elastase, histones, ROS and acid-fast bacilli. Viable and heat-killed M. tuberculosis, as well as Mycobacterium bovis BCG were efficient NET inducers, as were unilamellar liposomes prepared with lipids from M. tuberculosis. In vitro, guinea pig neutrophils also produced NETs in response to M. tuberculosis. However, neither the in vivo nor the in vitro-produced NETs were able to kill M. tuberculosis. Nevertheless, in vivo, neutrophils might propitiate recruitment and activation of more efficient microbicidal cells.

MicroRNA-365 in macrophages regulates Mycobacterium tuberculosis-induced active pulmonary tuberculosis via interleukin-6.

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Song, Qingzhang; Li, Hui; Shao, Hua; Li, Chunling; Lu, Xiao

2015-01-01

The present study is to investigate the relationship between microRNA (miR)-365 expression and the levels of interleukin (IL)-6 mRNA and protein in patients with active tuberculosis. From June 2011 to June 2014, 48 patients with active pulmonary tuberculosis induced by Mycobacterium tuberculosis were included in the study. In addition, 23 healthy subjects were enrolled as control. Macrophages were collected by pulmonary alveolus lavage. In addition, serum and mononuclear cells were isolated from peripheral blood. The levels of miR-365 and IL-6 in macrophages, mononuclear cells and serum were determined using quantitative real-time polymerase chain reaction. The protein expression of IL-6 in macrophages and mononuclear cells was measured using Western blotting, while that in serum was detected by enzyme-linked immunoabsorbent assay. Expression of IL-6 mRNA and protein was significantly enhanced in patients with active pulmonary tuberculosis. Increase of IL-6 protein concentration in serum was probably due to the release of IL-6 protein from mononuclear cells in the blood. In addition, miR-365 levels were significantly lowered in patients with active pulmonary tuberculosis. Up-regulated IL-6 expression in macrophages, mononuclear cells and serum in patients with active pulmonary tuberculosis is related to the down-regulation of miR-365, suggesting that miR-365 may regulate the occurrence and immune responses of active pulmonary tuberculosis via IL-6.

Spoligotyping of Mycobacterium tuberculosis isolates from patients with pulmonary tuberculosis in Mumbai, India.

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Kulkarni, Savita; Sola, Christophe; Filliol, Ingrid; Rastogi, Nalin; Kadival, Gururaj

2005-05-01

Tuberculosis remains a major health problem in India, with 2 million new cases and 421,000 deaths each year. In this paper, we describe the spoligotyping results of 216 Mycobacterium tuberculosis culture isolates from patients with pulmonary tuberculosis in Mumbai, India. As spoligotyping data from India have rarely been described until now, and as there is limited information on the major circulating clades of M. tuberculosis, the data obtained were also compared to an international spoligotype database (SpolDB4) that contained patterns from 22,546 isolates from more than 100 countries. Eighty-four (39%) of the isolates were definitively marked as orphan strains, indicating the paucity of such data from India. The remaining 132 isolates clustered among 59 shared types; among these, 42 shared types were already present in the database, 17 were newly created, and 5 of them were specifically reported from Mumbai. A total of 9 major types in this study clustered 32% of the isolates. At the phylogenetic level, 30% of the isolates belonged to the Central Asian families CAS1 and CAS2, of the major genetic group (MGG) 1, 29% to MGG 2 and 3 families (spacers 33-36 missing) and 17% to the ancestral East African Indian (EAI) family. Finally, nearly 10% of the isolates belonged to the W-Beijing family in a broad sense, also in the MGG 1 group. In conclusion, historic clones of the MGG 1 group of M. tuberculosis are responsible for roughly 60% of all tuberculosis cases in Mumbai. Together with the fact that organisms presumably of European descent (such as the Haarlem family) were only rarely found, our observations suggest that tuberculosis in Mumbai, India is essentially caused by historical clones of tubercle bacilli undergoing active circulation due to uncontrolled demography, high prevalence of the disease, and a paucity of resources.

Discovery of dormancy associated antigens of Mycobacterium tuberculosis : novel targets for the development of post-exposure or therapeutic tuberculosis vaccines

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Lin, May Young

2009-01-01

The growing number of tuberculosis (TB) casualties urges development of not only more effective drugs and preventive vaccines but also development of post-exposure/therapeutic TB vaccines. Post-exposure/therapeutic TB vaccines are needed since 2 billion people worldwide harbor a latent Mycobacterium

Evaluation of the efficacy of valproic acid and suberoylanilide hydroxamic acid (vorinostat in enhancing the effects of first-line tuberculosis drugs against intracellular Mycobacterium tuberculosis

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Martin Rao

2018-04-01

Full Text Available Background: New tuberculosis (TB drug treatment regimens are urgently needed. This study evaluated the potential of the histone deacetylase inhibitors (HDIs valproic acid (VPA and suberoylanilide hydroxamic acid (SAHA to enhance the effects of first-line anti-TB drugs against intracellular Mycobacterium tuberculosis. Methods: M. tuberculosis H37Rv cultures were exposed to VPA or SAHA over 6 days, in the presence or absence of isoniazid (INH and rifampicin (RIF. The efficacy of VPA and SAHA against intracellular M. tuberculosis with and without INH or RIF was tested by treating infected macrophages. Bactericidal activity was assessed by counting mycobacterial colony-forming units (CFU. Results: VPA treatment exhibited superior bactericidal activity to SAHA (2-log CFU reduction, while both HDIs moderately improved the activity of RIF against extracellular M. tuberculosis. The bactericidal effect of VPA against intracellular M. tuberculosis was greater than that of SAHA (1-log CFU reduction and equalled that of INH (1.5-log CFU reduction. INH/RIF and VPA/SAHA combination treatment inhibited intracellular M. tuberculosis survival in a shorter time span than monotherapy (3 days vs. 6 days. Conclusions: VPA and SAHA have adjunctive potential to World Health Organization-recommended TB treatment regimens. Clinical evaluation of the two drugs with regard to reducing the treatment duration and improving treatment outcomes in TB is warranted. Keywords: Mycobacterium tuberculosis, Adjunct host-directed therapy, Tuberculosis, Histone deacetylase inhibitors, Repurposed drugs

Ancient origin and gene mosaicism of the progenitor of Mycobacterium tuberculosis.

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M Cristina Gutierrez

2005-09-01

Full Text Available The highly successful human pathogen Mycobacterium tuberculosis has an extremely low level of genetic variation, which suggests that the entire population resulted from clonal expansion following an evolutionary bottleneck around 35,000 y ago. Here, we show that this population constitutes just the visible tip of a much broader progenitor species, whose extant representatives are human isolates of tubercle bacilli from East Africa. In these isolates, we detected incongruence among gene phylogenies as well as mosaic gene sequences, whose individual elements are retrieved in classical M. tuberculosis. Therefore, despite its apparent homogeneity, the M. tuberculosis genome appears to be a composite assembly resulting from horizontal gene transfer events predating clonal expansion. The amount of synonymous nucleotide variation in housekeeping genes suggests that tubercle bacilli were contemporaneous with early hominids in East Africa, and have thus been coevolving with their human host much longer than previously thought. These results open novel perspectives for unraveling the molecular bases of M. tuberculosis evolutionary success.

New Approaches and Therapeutic Options for Mycobacterium tuberculosis in a Dormant State.

NARCIS (Netherlands)

Caño-Muñiz, Santiago; Anthony, Richard; Niemann, Stefan; Alffenaar, Jan-Willem C

We are far away from the days when tuberculosis (TB) accounted for 1 in 4 deaths during the 19th century. However,Mycobacterium tuberculosiscomplex (MTBC) strains are still the leading cause of morbidity and mortality by a single infectious disease, with 9.6 million cases and 1.5 million deaths

Comparative Analyses of Nonpathogenic, Opportunistic, and Totally Pathogenic Mycobacteria Reveal Genomic and Biochemical Variabilities and Highlight the Survival Attributes of Mycobacterium tuberculosis

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Singh, Yadvir; Kohli, Sakshi; Ahmad, Javeed; Ehtesham, Nasreen Z.; Tyagi, Anil K.

2014-01-01

ABSTRACT Mycobacterial evolution involves various processes, such as genome reduction, gene cooption, and critical gene acquisition. Our comparative genome size analysis of 44 mycobacterial genomes revealed that the nonpathogenic (NP) genomes were bigger than those of opportunistic (OP) or totally pathogenic (TP) mycobacteria, with the TP genomes being smaller yet variable in size—their genomic plasticity reflected their ability to evolve and survive under various environmental conditions. From the 44 mycobacterial species, 13 species, representing TP, OP, and NP, were selected for genomic-relatedness analyses. Analysis of homologous protein-coding genes shared between Mycobacterium indicus pranii (NP), Mycobacterium intracellulare ATCC 13950 (OP), and Mycobacterium tuberculosis H37Rv (TP) revealed that 4,995 (i.e., ~95%) M. indicaus pranii proteins have homology with M. intracellulare, whereas the homologies among M. indicus pranii, M. intracellulare ATCC 13950, and M. tuberculosis H37Rv were significantly lower. A total of 4,153 (~79%) M. indicus pranii proteins and 4,093 (~79%) M. intracellulare ATCC 13950 proteins exhibited homology with the M. tuberculosis H37Rv proteome, while 3,301 (~82%) and 3,295 (~82%) M. tuberculosis H37Rv proteins showed homology with M. indicus pranii and M. intracellulare ATCC 13950 proteomes, respectively. Comparative metabolic pathway analyses of TP/OP/NP mycobacteria showed enzymatic plasticity between M. indicus pranii (NP) and M. intracellulare ATCC 13950 (OP), Mycobacterium avium 104 (OP), and M. tuberculosis H37Rv (TP). Mycobacterium tuberculosis seems to have acquired novel alternate pathways with possible roles in metabolism, host-pathogen interactions, virulence, and intracellular survival, and by implication some of these could be potential drug targets. PMID:25370496

Tumor necrosis factor blockers influence macrophage responses to Mycobacterium tuberculosis

OpenAIRE

HARRIS, JAMES; HARRIS, JAMES

2008-01-01

PUBLISHED umor necrosis factor (TNF)?? is a proinflammatory cytokine that mediates inflammation in response to various pathogens, including Mycobacterium tuberculosis, but is also a key factor in the pathogenesis of rheumatoid arthritis and other autoimmune diseases. Three TNF???suppressing drugs have been approved to treat selected autoimmune diseases; 2 are monoclonal antibodies against TNF?? (adalimumab and infliximab), and the other is a soluble TNF receptor/Fc fusion protein (etanerce...

Performance of a Highly Sensitive Mycobacterium tuberculosis Complex Real-Time PCR Assay for Diagnosis of Pulmonary Tuberculosis in a Low-Prevalence Setting: a Prospective Intervention Study.

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Vinuesa, Víctor; Borrás, Rafael; Briones, María Luisa; Clari, María Ángeles; Cresencio, Vicenta; Giménez, Estela; Muñoz, Carmen; Oltra, Rosa; Servera, Emilio; Scheelje, Talia; Tornero, Carlos; Navarro, David

2018-05-01

The potential impact of routine real-time PCR testing of respiratory specimens from patients with presumptive tuberculosis in terms of diagnostic accuracy and time to tuberculosis treatment inception in low-prevalence settings remains largely unexplored. We conducted a prospective intervention cohort study. Respiratory specimens from 1,020 patients were examined by acid-fast bacillus smear microscopy, tested by a real-time Mycobacterium tuberculosis complex PCR assay (Abbott RealTi me MTB PCR), and cultured in mycobacterial media. Seventeen patients tested positive by PCR (5 were acid-fast bacillus smear positive and 12 acid-fast bacillus smear negative), and Mycobacterium tuberculosis was recovered from cultures for 12 of them. Patients testing positive by PCR and negative by culture ( n = 5) were treated and deemed to have responded to antituberculosis therapy. There were no PCR-negative/culture-positive cases, and none of the patients testing positive for nontuberculous mycobacteria ( n = 20) yielded a positive PCR result. The data indicated that routine testing of respiratory specimens from patients with presumptive tuberculosis by the RealTi me MTB PCR assay improves the tuberculosis diagnostic yield and may reduce the time to antituberculosis treatment initiation. On the basis of our data, we propose a novel mycobacterial laboratory algorithm for tuberculosis diagnosis. Copyright © 2018 American Society for Microbiology.

Clustering of Beijing genotype Mycobacterium tuberculosis isolates from the Mekong delta in Vietnam on the basis of variable number of tandem repeat versus restriction fragment length polymorphism typing

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Huyen Mai NT

2013-02-01

Full Text Available Abstract Background In comparison to restriction fragment length polymorphism (RFLP typing, variable number of tandem repeat (VNTR typing is easier to perform, faster and yields results in a simple, numerical format. Therefore, this technique has gained recognition as the new international gold standard in typing of Mycobacterium tuberculosis. However, some reports indicated that VNTR typing may be less suitable for Beijing genotype isolates. We therefore compared the performance of internationally standardized RFLP and 24 loci VNTR typing to discriminate among 100 Beijing genotype isolates from the Southern Vietnam. Methods Hundred Beijing genotype strains defined by spoligotyping were randomly selected and typed by RFLP and VNTR typing. The discriminatory power of VNTR and RFLP typing was compared using the Bionumerics software. Results Among 95 Beijing strains available for analysis, 14 clusters were identified comprising 34 strains and 61 unique profiles in 24 loci VNTR typing ((Hunter Gaston Discrimination Index (HGDI = 0.994. 13 clusters containing 31 strains and 64 unique patterns in RFLP typing (HGDI = 0.994 were found. Nine RFLP clusters were subdivided by VNTR typing and 12 VNTR clusters were split by RFLP. Five isolates (5% revealing double alleles or no signal in two or more loci in VNTR typing could not be analyzed. Conclusions Overall, 24 loci VNTR typing and RFLP typing had similar high-level of discrimination among 95 Beijing strains from Southern Vietnam. However, loci VNTR 154, VNTR 2461 and VNTR 3171 had hardly added any value to the level of discrimination.

Identification of MHC class II restricted T‐cell‐mediated reactivity against MHC class I binding Mycobacterium tuberculosis peptides

DEFF Research Database (Denmark)

Wang, Mingjun; Tang, Sheila Tuyet; Stryhn, Anette

2011-01-01

Major histocompatibility complex (MHC) class I restricted cytotoxic T lymphocytes (CTL) are known to play an important role in the control of Mycobacterium tuberculosis infection so identification of CTL epitopes from M. tuberculosis is of importance for the development of effective peptide...

[Resistance to first-line drugs and major genotypic lineages of Mycobacterium tuberculosis in the 3 French Department of the Americas: Profiles, evolution, and trends (1995-2011)].

Science.gov (United States)

Millet, J; Berchel, M; Prudenté, F; Streit, E; Bomer, A-G; Schuster, F; Vanhomwegen, J; Paasch, D; Galbert, I; Valery, E; Aga, R; Rastogi, N

2014-05-01

This is the first overview on resistant and multidrug resistant isolates of Mycobacterium tuberculosis circulating in the French Department of the Americas (Guadeloupe, Martinique, and French Guiana) over 17 years (January 1995-December 2011). A total of 1,239 cases were studied: 1,199 new cases (primary and multidrug resistance of 11.8 and 1.6% respectively), and 40 persistent (defined as cases with a previous history of positive culture over 6 months interval and whose spoligotypes remain unchanged), in which significantly higher proportions of resistance to at least isoniazid (22.5%, P = 0.002), rifampicin (20.0%, P < 0.001), and multidrug resistance (17.5%, P < 0.001) were observed as compared to new cases. The 281 spoligotypes obtained showed the presence of five major lineages, T (29.9%), LAM (23.9%), Haarlem (22.1%), EAI (7.1%), and X (6.7%). Two of these lineages, X and LAM, predominate among resistant and multidrug resistant isolates respectively (X: 10.5% of resistant isolates, P = 0.04; LAM: 42.3% of multidrug resistant isolates, P = 0.02). Four of the 19 major spoligo-profiles, corresponding to SIT 20, 64, 45, and 46, were significantly associated with drug resistance. Among them, genotype SIT 20, associated with monoresistance to isoniazid and multidrug resistance, would be actively and persistently in circulation, since 1999, in French Guiana, department in which one may also observe the presence of strains of M. tuberculosis phylogeographically associated to Guiana and Suriname (SIT 131 and SIT 1340).

The cell envelope glycoconjugates of Mycobacterium tuberculosis

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Angala, Shiva Kumar; Belardinelli, Juan Manuel; Huc-Claustre, Emilie; Wheat, William H.; Jackson, Mary

2015-01-01

Tuberculosis (TB) remains the second most common cause of death due to a single infectious agent. The cell envelope of Mycobacterium tuberculosis (Mtb), the causative agent of the disease in humans, is a source of unique glycoconjugates and the most distinctive feature of the biology of this organism. It is the basis of much of Mtb pathogenesis and one of the major causes of its intrinsic resistance to chemotherapeutic agents. At the same time, the unique structures of Mtb cell envelope glycoconjugates, their antigenicity and essentiality for mycobacterial growth provide opportunities for drug, vaccine, diagnostic and biomarker development, as clearly illustrated by recent advances in all of these translational aspects. This review focuses on our current understanding of the structure and biogenesis of Mtb glycoconjugates with particular emphasis on one of most intriguing and least understood aspect of the physiology of mycobacteria: the translocation of these complex macromolecules across the different layers of the cell envelope. It further reviews the rather impressive progress made in the last ten years in the discovery and development of novel inhibitors targeting their biogenesis. PMID:24915502

In vitro activities of DA-7157 and DA-7218 against Mycobacterium tuberculosis and Nocardia brasiliensis.

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Vera-Cabrera, Lucio; Gonzalez, Eva; Rendon, Adrian; Ocampo-Candiani, Jorge; Welsh, Oliverio; Velazquez-Moreno, Victor M; Choi, Sung Hak; Molina-Torres, Carmen

2006-09-01

The in vitro activities of DA-7157, a novel oxazolidinone, against clinical isolates of Nocardia brasiliensis and Mycobacterium tuberculosis were determined. Equal MIC(50)s and MIC(90)s (0.25 and 0.5 microg/ml, respectively) were found for susceptible and multidrug-resistant isolates of M. tuberculosis. The N. brasiliensis isolates showed an MIC(90) of 1 microg/ml and an MIC(50) of 1 microg/ml. The DA-7157 prodrug, DA-7218, exhibited similar MICs for M. tuberculosis but fivefold-higher MICs for N. brasiliensis.

Duplex detection of the Mycobacterium tuberculosis complex and medically important non-tuberculosis mycobacteria by real-time PCR based on the rnpB gene.

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Abdeldaim, Guma; Svensson, Erik; Blomberg, Jonas; Herrmann, Björn

2016-11-01

A duplex real-time PCR based on the rnpB gene was developed for Mycobacterium spp. The assay was specific for the Mycobacterium tuberculosis complex (MTB) and also detected all 19 tested species of non-tuberculous mycobacteria (NTM). The assay was evaluated on 404 clinical samples: 290 respiratory samples and 114 from tissue and other non-respiratory body sites. M. tuberculosis was detected by culture in 40 samples and in 30 samples by the assay. The MTB assay showed a sensitivity similar to Roche Cobas Amplicor MTB-PCR (Roche Molecular Systems, Pleasanton, CA, USA). There were only nine samples with non-tuberculous mycobacteria detected by culture. Six of them were detected by the PCR assay. © 2016 APMIS. Published by John Wiley & Sons Ltd.

Diagnosis of intestinal tuberculosis using a monoclonal antibody to Mycobacterium tuberculosis

Institute of Scientific and Technical Information of China (English)

Yasushi Ihama; Akira Hokama; Kenji Hibiya; Kazuto Kishimoto; Manabu Nakamoto; Tetsuo Hirata; Nagisa Kinjo

2012-01-01

AIM:To investigate the utility of immunohistochemical (IHC) staining with an antibody to Mycobacterium tuberculosis (M.tuberculosis) for the diagnosis of intestinal tuberculosis (TB).METHODS:We retrospectively identified 10 patients (4 males and 6 females; mean age =65.1 ± 13.6 years)with intestinal TB.Clinical characteristics,including age,gender,underlying disease,and symptoms were obtained.Chest radiograph and laboratory tests,including sputum Ziehl-Neelsen (ZN) staining,M.tuberculosis culture,and sputum polymerase chain reaction (PCR)for tubercle bacilli DNA,as well as Tuberculin skin test (TST) and QuantiFERON-TB gold test (QFT),were examined.Colonoscopic records recorded on the basis of Sato's classification were also reviewed,in addition to data from intestinal biopsies examined for histopathological findings,including hematoxylin and eosin staining,and ZN staining,as well as M.tuberculosis culture,and PCR for tubercle bacilli DNA.For the present study,archived formalin-fixed paraffin-embedded (FFPE) intestinal tissue samples were immunohistochemically stained using a commercially available species-specific monoclonal antibody to the 38-kDa antigen of the M.tuberculosis complex.These sections were also stained with the pan-macrophage marker CD68 antibody.RESULTS:From the clinical data,we found that no patients were immunocompromised,and that the main symptoms were diarrhea and weight loss.Three patients displayed active pulmonary TB,six patients (60％) had a positive TST,and 4 patients (40％) had a positive QFT.Colonoscopic findings revealed that all patients had type 1 findings (linear ulcers in a circumferential arrangement or linear ulcers arranged circumferentially with mucosa showing multiple nodules),all of which were located in the right hemicolon and/or terminal ileum.Seven patients (70％) had concomitant healed lesions in the ileocecal area.No acid-fast bacilli were detected with ZN staining of the intestinal tissue samples,and both M.tuberculosis

Laboratory Diagnosis and Susceptibility Testing for Mycobacterium tuberculosis.

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Procop, Gary W

2016-12-01

The laboratory, which utilizes some of the most sophisticated and rapidly changing technologies, plays a critical role in the diagnosis of tuberculosis. Some of these tools are being employed in resource-challenged countries for the rapid detection and characterization of Mycobacterium tuberculosis. Foremost, the laboratory defines appropriate specimen criteria for optimal test performance. The direct detection of mycobacteria in the clinical specimen, predominantly done by acid-fast staining, may eventually be replaced by rapid-cycle PCR. The widespread use of the Xpert MTB/RIF (Cepheid) assay, which detects both M. tuberculosis and key genetic determinants of rifampin resistance, is important for the early detection of multidrug-resistant strains. Culture, using both broth and solid media, remains the standard for establishing the laboratory-based diagnosis of tuberculosis. Cultured isolates are identified far less commonly by traditional biochemical profiling and more commonly by molecular methods, such as DNA probes and broad-range PCR with DNA sequencing. Non-nucleic acid-based methods of identification, such as high-performance liquid chromatography and, more recently, matrix-assisted laser desorption/ionization-time of flight mass spectrometry, may also be used for identification. Cultured isolates of M. tuberculosis should be submitted for susceptibility testing according to standard guidelines. The use of broth-based susceptibility testing is recommended to significantly decrease the time to result. Cultured isolates may also be submitted for strain typing for epidemiologic purposes. The use of massive parallel sequencing, also known as next-generation sequencing, promises to continue to this molecular revolution in mycobacteriology, as whole-genome sequencing provides identification, susceptibility, and typing information simultaneously.

Production of antibodies against glycolipids from the Mycobacterium tuberculosis cell wall in aerosol murine models of tuberculosis.

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Cardona, P J; Julián, E; Vallès, X; Gordillo, S; Muñoz, M; Luquin, M; Ausina, V

2002-06-01

Evolution of antibodies against glycolipids from the Mycobacterium tuberculosis cell wall has been studied for the first time in experimental murine models of tuberculosis induced by aerosol, in which infection, reinfection, reactivation, prophylaxis and treatment with antibiotics have been assayed. Results show a significant humoral response against these antigens, where diacyltrehaloses (DAT) and sulpholipid I (SL-I) elicited higher antibody levels than protein antigens like antigen 85 protein complex (Ag85), culture filtrate proteins (CFP) and purified protein derivative (PPD). Only immunoglobulin M (IgM) antibodies have been detected against DAT and SL-I. Their evolution has a positive correlation with bacillary concentration in tissues.

Immunological response to Mycobacterium tuberculosis infection in blood from type 2 diabetes patients.

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Raposo-García, Sara; Guerra-Laso, José Manuel; García-García, Silvia; Juan-García, Javier; López-Fidalgo, Eduardo; Diez-Tascón, Cristina; Nebreda-Mayoral, Teresa; López-Medrano, Ramiro; Rivero-Lezcano, Octavio Miguel

2017-06-01

The convergence of tuberculosis and diabetes represents a co-epidemic that threatens progress against tuberculosis. We have investigated type 2 diabetes as a risk factor for tuberculosis susceptibility, and have used as experimental model whole blood infected in vitro with Mycobacterium tuberculosis. Blood samples from diabetic patients were found to have a higher absolute neutrophil count that non-diabetic controls, but their immune functionality seemed impaired because they displayed a lower capacity to phagocytose M. tuberculosis, a finding that had been previously reported only for monocytes. In contrast, an increased production of TNFα was detected in infected blood from diabetic patients. Despite the altered phagocytic capacity showed by cells from these patients, the antimicrobial activity measured in both whole blood and monocyte derived macrophages was similar to that of controls. This unexpected result prompts further improvements in the whole blood model to analyze the immune response of diabetes patients to tuberculosis. Copyright © 2017 European Federation of Immunological Societies. Published by Elsevier B.V. All rights reserved.

High prevalence of clustered tuberculosis cases in Peruvian migrants in Florence, Italy

Directory of Open Access Journals (Sweden)

Lorenzo Zammarchi

2014-12-01

Full Text Available Tuberculosis is a leading cause of morbidity for Peruvian migrants in Florence, Italy, where they account for about 20% of yearly diagnosed cases. A retrospective study on cases notified in Peruvian residents in Florence in the period 2001-2010 was carried out and available Mycobacterium tuberculosis strains were genotyped (MIRU-VNTR-24 and Spoligotyping. One hundred thirty eight cases were retrieved. Genotyping performed in 87 strains revealed that 39 (44.8% belonged to 12 clusters. Assuming that in each cluster the transmission of tuberculosis from the index case took place in Florence, a large proportion of cases could be preventable by improving early diagnosis of contagious cases and contact tracing.

Clinical Concentrations of Thioridazine Kill Intracellular Multidrug-Resistant Mycobacterium tuberculosis

Science.gov (United States)

Ordway, Diane; Viveiros, Miguel; Leandro, Clara; Bettencourt, Rosário; Almeida, Josefina; Martins, Marta; Kristiansen, Jette E.; Molnar, Joseph; Amaral, Leonard

2003-01-01

The phenothiazines chlorpromazine (CPZ) and thioridazine (TZ) have equal in vitro activities against antibiotic-sensitive and -resistant Mycobacterium tuberculosis. These compounds have not been used as anti-M. tuberculosis agents because their in vitro activities take place at concentrations which are beyond those that are clinically achievable. In addition, chronic administration of CPZ produces frequent severe side effects. Because CPZ has been shown to enhance the killing of intracellular M. tuberculosis at concentrations in the medium that are clinically relevant, we have investigated whether TZ, a phenothiazine whose negative side effects are less frequent and serious than those associated with CPZ, kills M. tuberculosis organisms that have been phagocytosed by human macrophages, which have nominal killing activities against these bacteria. Both CPZ and TZ killed intracellular antibiotic-sensitive and -resistant M. tuberculosis organisms when they were used at concentrations in the medium well below those present in the plasma of patients treated with these agents. These concentrations in vitro were not toxic to the macrophage, nor did they affect in vitro cellular immune processes. TZ thus appears to be a serious candidate for the management of a freshly diagnosed infection of pulmonary tuberculosis or as an adjunct to conventional antituberculosis therapy if the patient originates from an area known to have a high prevalence of multidrug-resistant M. tuberculosis isolates. Nevertheless, we must await the outcomes of clinical trials to determine whether TZ itself may be safely and effectively used as an antituberculosis agent. PMID:12604522

Meropenem-Clavulanate is Effective Against Extensive Drug-Resistant Mycobacterium Tuberculosis

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Hugonnet, J.; Tremblay, L; Boshoff, H; Barry, C; Blanchard, J

2009-01-01

e-lactam antibiotics are ineffective against Mycobacterium tuberculosis, being rapidly hydrolyzed by the chromosomally encoded blaC gene product. The carbapenem class of e-lactams are very poor substrates for BlaC, allowing us to determine the three-dimensional structure of the covalent BlaC-meropenem covalent complex at 1.8 angstrom resolution. When meropenem was combined with the e-lactamase inhibitor clavulanate, potent activity against laboratory strains of M. tuberculosis was observed [minimum inhibitory concentration (MICmeropenem) less than 1 microgram per milliliter], and sterilization of aerobically grown cultures was observed within 14 days. In addition, this combination exhibited inhibitory activity against anaerobically grown cultures that mimic the 'persistent' state and inhibited the growth of 13 extensively drug-resistant strains of M. tuberculosis at the same levels seen for drug-susceptible strains. Meropenem and clavulanate are Food and Drug Administration-approved drugs and could potentially be used to treat patients with currently untreatable disease.

Human tuberculosis caused by Mycobacterium bovis: a retrospective comparison with Mycobacterium tuberculosis in a Mexican tertiary care centre, 2000-2015.

Science.gov (United States)

Torres-Gonzalez, Pedro; Cervera-Hernandez, Miguel E; Martinez-Gamboa, Areli; Garcia-Garcia, Lourdes; Cruz-Hervert, Luis P; Bobadilla-Del Valle, Miriam; Ponce-de Leon, Alfredo; Sifuentes-Osornio, Jose

2016-11-08

Human tuberculosis caused by Mycobacterium bovis is believed to be frequent in developing countries. Transmission is usually through ingestion of unpasteurized dairy products, although airborne contagion is possible. Disease caused by M. tuberculosis or M. bovis is clinically indistinguishable from each other. The aim of this study was to determine the factors associated with M. bovis disease. Retrospective analysis of all culture-positive cases of M. bovis and M. tuberculosis from 2000 to 2015, in a Mexican tertiary-care centre. Sociodemographic, clinical, and radiographic data from medical records were compared. Disease site was classified as pulmonary, extrapulmonary, or pulmonary and extrapulmonary, based on cultures. We evaluated 533 cases, 372 (69.7 %) of which were caused by M. tuberculosis and 161 (30.2 %) by M. bovis. Characteristics associated with M. bovis disease were: younger age (aOR 0.97, 95 % CI 0.95-0.98), glucocorticoid use (aOR 2.27, 95 % CI 1.42-3.63), and extrapulmonary disease (aOR 1.80, 95 % CI 1.21-2.69). M. tuberculosis was associated with lower socioeconomic status (aOR 0.52, 95 % CI 0.28-0.97). When we analysed only pulmonary cases, younger age (aOR 0.97, 95 % CI 0.96-0.99), glucocorticoid use (aOR 2.41, 95 % CI 1.30-4.46), and smoking (aOR 1.94, CI 95 % 1.15-3.27) were associated with M. bovis. Both groups showed similar proportions of direct microscopy smear results (respiratory samples) and chest X-ray cavitations. Younger age, glucocorticoid use, and extrapulmonary disease were associated with M. bovis as the causative agent of tuberculosis in a group of patients from a tertiary care centre in a country where bovine tuberculosis is endemic. Further studies must be conducted in the general population to determine pathogen-specific associated factors and outcomes.

Activity of Scottish plant, lichen and fungal endophyte extracts against Mycobacterium aurum and Mycobacterium tuberculosis.

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Gordien, Andréa Y; Gray, Alexander I; Ingleby, Kevin; Franzblau, Scott G; Seidel, Véronique

2010-05-01

With tuberculosis the leading bacterial killer worldwide and other mycobacterial diseases on the increase, the search for new antimycobacterial agents is timely. In this study, extracts from plants, lichens and fungal endophytes of Scottish provenance were screened for activity against Mycobacterium aurum and M. tuberculosis H(37)Rv. The best activity against M. aurum was observed for extracts of Juniperus communis roots and Cladonia arbuscula (MIC = 4 microg/mL), and a fungal endophyte isolated from Vaccinium myrtillus (MIC = 8 microg/mL). The best activity against M. tuberculosis was observed for extracts of C. arbuscula, Empetrum nigrum, J. communis roots, Calluna vulgaris aerial parts, Myrica gale roots and stems (93 to 99% inhibition at 100 microg/mL). Potent antitubercular activity (90 to 96% inhibition at 100 microg/mL) was also observed for the ethanol extracts of Xerocomus badius, Chalciporus piperatus, Suillus luteus and of endophytes isolated from C. vulgaris, E. nigrum, Vaccinium vitis-idaea and V. myrtillus. The results obtained this study provide, in part, some scientific basis for the traditional use of some of the selected plants in the treatment of tuberculosis. They also indicate that fungal endophytes recovered from Scottish plants are a source of antimycobacterial agents worthy of further investigation. Copyright (c) 2009 John Wiley & Sons, Ltd.

Microbial sensor for drug susceptibility testing of Mycobacterium tuberculosis.

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Zhang, Z-T; Wang, D-B; Li, C-Y; Deng, J-Y; Zhang, J-B; Bi, L-J; Zhang, X-E

2018-01-01

Drug susceptibility testing (DST) of clinical isolates of Mycobacterium tuberculosis is critical in treating tuberculosis. We demonstrate the possibility of using a microbial sensor to perform DST of M. tuberculosis and shorten the time required for DST. The sensor is made of an oxygen electrode with M. tuberculosis cells attached to its surface. This sensor monitors the residual oxygen consumption of M. tuberculosis cells after treatment with anti-TB drugs with glycerine as a carbon source. In principle, after drug pretreatment for 4-5 days, the response differences between the sensors made of drug-sensitive isolates are distinguishable from the sensors made of drug-resistant isolates. The susceptibility of the M. tuberculosis H37Ra strain, its mutants and 35 clinical isolates to six common anti-TB drugs: rifampicin, isoniazid, streptomycin, ethambutol, levofloxacin and para-aminosalicylic acid were tested using the proposed method. The results agreed well with the gold standard method (LJ) and were determined in significantly less time. The whole procedure takes approximately 11 days and therefore has the potential to inform clinical decisions. To our knowledge, this is the first study that demonstrates the possible application of a dissolved oxygen electrode-based microbial sensor in M. tuberculosis drug resistance testing. This study used the microbial sensor to perform DST of M. tuberculosis and shorten the time required for DST. The overall detection result of the microbial sensor agreed well with that of the conventional LJ proportion method and takes less time than the existing phenotypic methods. In future studies, we will build an O 2 electrode array microbial sensor reactor to enable a high-throughput drug resistance analysis. © 2017 The Authors. Journal of Applied Microbiology published by John Wiley & Sons Ltd on behalf of The Society for Applied Microbiology.

Mycobacterium tuberculosis strains exhibit differential and strain-specific molecular signatures in pulmonary epithelial cells.

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Mvubu, Nontobeko Eunice; Pillay, Balakrishna; Gamieldien, Junaid; Bishai, William; Pillay, Manormoney

2016-12-01

Although pulmonary epithelial cells are integral to innate and adaptive immune responses during Mycobacterium tuberculosis infection, global transcriptomic changes in these cells remain largely unknown. Changes in gene expression induced in pulmonary epithelial cells infected with M. tuberculosis F15/LAM4/KZN, F11, F28, Beijing and Unique genotypes were investigated by RNA sequencing (RNA-Seq). The Illumina HiSeq 2000 platform generated 50 bp reads that were mapped to the human genome (Hg19) using Tophat (2.0.10). Differential gene expression induced by the different strains in infected relative to the uninfected cells was quantified and compared using Cufflinks (2.1.0) and MeV (4.0.9), respectively. Gene expression varied among the strains with the total number of genes as follows: F15/LAM4/KZN (1187), Beijing (1252), F11 (1639), F28 (870), Unique (886) and H37Rv (1179). A subset of 292 genes was commonly induced by all strains, where 52 genes were down-regulated while 240 genes were up-regulated. Differentially expressed genes were compared among the strains and the number of induced strain-specific gene signatures were as follows: F15/LAM4/KZN (138), Beijing (52), F11 (255), F28 (55), Unique (186) and H37Rv (125). Strain-specific molecular gene signatures associated with functional pathways were observed only for the Unique and H37Rv strains while certain biological functions may be associated with other strain signatures. This study demonstrated that strains of M. tuberculosis induce differential gene expression and strain-specific molecular signatures in pulmonary epithelial cells. Specific signatures induced by clinical strains of M. tuberculosis can be further explored for novel host-associated biomarkers and adjunctive immunotherapies. Copyright © 2016 Elsevier Ltd. All rights reserved.

Human ULK1 Variation and Susceptibility to Mycobacterium tuberculosis Infection.

Science.gov (United States)

Horne, David J; Graustein, Andrew D; Shah, Javeed A; Peterson, Glenna; Savlov, Meg; Steele, Sergio; Narita, Masahiro; Hawn, Thomas R

2016-10-15

Unlike tuberculosis, few studies have evaluated a host genetic basis for variability in susceptibility to latent Mycobacterium tuberculosis infection (LTBI). We performed a candidate gene association study of autophagy-related genes and LTBI. We enrolled close contacts of individuals with pulmonary tuberculosis, assessed LTBI status, and determined clinical and sociodemographic risk factors for LTBI. In participants who self-identified as Asian or black, we compared haplotype-tagging single-nucleotide polymorphisms (SNPs) in ULK1 and GABARAP between cases (n = 143) and controls (n = 106). Using CRISPR/Cas9 in U937 monocytes, we investigated the effect of ULK1 deficiency on cytokine expression, autophagy, and M. tuberculosis replication. In Asian participants, we identified 2 ULK1 SNPs (rs12297124 and rs7300908) associated with LTBI. After adjustment for population admixture and clinical risk for LTBI, each rs12297124 minor allele conferred 80% reduction in LTBI risk (odds ratio, 0.18; 95% confidence interval, .07-.46). Compared with controls, ULK1-deficient cells exhibited decreased tumor necrosis factor secretion after stimulation with Toll-like receptor ligands and M. tuberculosis whole-cell lysate, increased M. tuberculosis replication, and decreased selective autophagy. These results demonstrate a strong association of rs12297124, a noncoding ULK1 SNP, with LTBI and a role for ULK1 regulation of TNF secretion, nonspecific and M. tuberculosis-induced autophagy, and M. tuberculosis replication in monocytes. © The Author 2016. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail journals.permissions@oup.com.

Comparative 'omics analyses differentiate Mycobacterium tuberculosis and Mycobacterium bovis and reveal distinct macrophage responses to infection with the human and bovine tubercle bacilli

Science.gov (United States)

Malone, Kerri M.; Rue-Albrecht, Kévin; Magee, David A.; Conlon, Kevin; Schubert, Olga T.; Nalpas, Nicolas C.; Browne, John A.; Smyth, Alicia; Gormley, Eamonn; Aebersold, Ruedi; MacHugh, David E.; Gordon, Stephen V.

2018-01-01

Members of the Mycobacterium tuberculosis complex (MTBC) are the causative agents of tuberculosis in a range of mammals, including humans. A key feature of MTBC pathogens is their high degree of genetic identity yet distinct host tropism. Notably, while Mycobacterium bovis is highly virulent and pathogenic for cattle, the human pathogen M. tuberculosis is attenuated in cattle. Previous research also suggests that host preference amongst MTBC members has a basis in host innate immune responses. To explore MTBC host tropism, we present in-depth profiling of the MTBC reference strains M. bovis AF2122/97 and M. tuberculosis H37Rv at both the global transcriptional and the translational level via RNA-sequencing and SWATH MS. Furthermore, a bovine alveolar macrophage infection time course model was used to investigate the shared and divergent host transcriptomic response to infection with M. tuberculosis H37Rv or M. bovis AF2122/97. Significant differential expression of virulence-associated pathways between the two bacilli was revealed, including the ESX-1 secretion system. A divergent transcriptional response was observed between M. tuberculosis H37Rv and M. bovis AF2122/97 infection of bovine alveolar macrophages, in particular cytosolic DNA-sensing pathways at 48 h post-infection, and highlights a distinct engagement of M. bovis with the bovine innate immune system. The work presented here therefore provides a basis for the identification of host innate immune mechanisms subverted by virulent host-adapted mycobacteria to promote their survival during the early stages of infection. PMID:29557774

Genetic Diversity and Transmission Characteristics of Beijing Family Strains of Mycobacterium tuberculosis in Peru

Science.gov (United States)

Iwamoto, Tomotada; Grandjean, Louis; Arikawa, Kentaro; Nakanishi, Noriko; Caviedes, Luz; Coronel, Jorge; Sheen, Patricia; Wada, Takayuki; Taype, Carmen A.; Shaw, Marie-Anne; Moore, David A. J.; Gilman, Robert H.

2012-01-01

Beijing family strains of Mycobacterium tuberculosis have attracted worldwide attention because of their wide geographical distribution and global emergence. Peru, which has a historical relationship with East Asia, is considered to be a hotspot for Beijing family strains in South America. We aimed to unveil the genetic diversity and transmission characteristics of the Beijing strains in Peru. A total of 200 Beijing family strains were identified from 2140 M. tuberculosis isolates obtained in Lima, Peru, between December 2008 and January 2010. Of them, 198 strains were classified into sublineages, on the basis of 10 sets of single nucleotide polymorphisms (SNPs). They were also subjected to variable number tandem-repeat (VNTR) typing using an international standard set of 15 loci (15-MIRU-VNTR) plus 9 additional loci optimized for Beijing strains. An additional 70 Beijing family strains, isolated between 1999 and 2006 in Lima, were also analyzed in order to make a longitudinal comparison. The Beijing family was the third largest spoligotyping clade in Peru. Its population structure, by SNP typing, was characterized by a high frequency of Sequence Type 10 (ST10), which belongs to a modern subfamily of Beijing strains (178/198, 89.9%). Twelve strains belonged to the ancient subfamily (ST3 [n = 3], ST25 [n = 1], ST19 [n = 8]). Overall, the polymorphic information content for each of the 24 loci values was low. The 24 loci VNTR showed a high clustering rate (80.3%) and a high recent transmission index (RTIn−1 = 0.707). These strongly suggest the active and on-going transmission of Beijing family strains in the survey area. Notably, 1 VNTR genotype was found to account for 43.9% of the strains. Comparisons with data from East Asia suggested the genotype emerged as a uniquely endemic clone in Peru. A longitudinal comparison revealed the genotype was present in Lima by 1999. PMID:23185395

Cytokine responses in relation to age, gender, body mass index, Mycobacterium tuberculosis infection, and otitis media among inuit in greenland

DEFF Research Database (Denmark)

Nielsen, Nina Odgaard; Soborg, Bolette; Børresen, Malene

2013-01-01

To evaluate the cytokine response pattern in Inuit in Greenland in relation to age, gender, body mass index (BMI), Mycobacterium tuberculosis infection (MTI), and otitis media (OM) to assess whether Inuit may have signs of impaired immune responsiveness to infection.......To evaluate the cytokine response pattern in Inuit in Greenland in relation to age, gender, body mass index (BMI), Mycobacterium tuberculosis infection (MTI), and otitis media (OM) to assess whether Inuit may have signs of impaired immune responsiveness to infection....

Description of polymerase chain reaction and sequencing DNA Mycobacterium tuberculosis from specimen sputum of tuberculosis patients in Medan

Science.gov (United States)

Lily; Siregar, Y.; Ilyas, S.

2018-03-01

This study purposed to describe the product Polymerase Chain Reaction (PCR) and sequencing of DNA Mycobacterium (M.) tuberculosis from sputum of tuberculosis (TB) patients in Medan. Sputum was collected from patients that diagnosed with pulmonary TB by a physician. Specimen processed by PCR method of Li et al. and sequencing at Macrogen Laboratory. All of 12 product PCR were showed brightness bands at 126 base pair (bp). These results indicated similarity to the study of Li et al. Sequencing analysis showed the presence of a mutation and non-mutation groups of M. tuberculosis. The reference and outcome berange of the mutation and non-mutation of M. tuberculosis were 56-107, 59-85, 60-120 and 63-94, respectively. The percentage bp difference between the outcome and references for mutation and non-mutation were 3.448-6.569and 3.278-7.428%, respectively. In conclusion, the successful amplification of PCR products in a 1.5% agarose gel electrophoresis where all 12 sputa contained rpoB-positive M. tuberculosis and 0.644% difference was found between the outcome with reference bp of the mutation and non-mutation M. tuberculosis groups.

Sulfonamide-Based Inhibitors of Aminoglycoside Acetyltransferase Eis Abolish Resistance to Kanamycin in Mycobacterium tuberculosis

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Garzan, Atefeh; Willby, Melisa J.; Green, Keith D.; Gajadeera, Chathurada S.; Hou, Caixia; Tsodikov, Oleg V.; Posey, James E.; Garneau-Tsodikova, Sylvie

2016-12-08

A two-drug combination therapy where one drug targets an offending cell and the other targets a resistance mechanism to the first drug is a time-tested, yet underexploited approach to combat or prevent drug resistance. By high-throughput screening, we identified a sulfonamide scaffold that served as a pharmacophore to generate inhibitors of Mycobacterium tuberculosis acetyltransferase Eis, whose upregulation causes resistance to the aminoglycoside (AG) antibiotic kanamycin A (KAN) in Mycobacterium tuberculosis. Rational systematic derivatization of this scaffold to maximize Eis inhibition and abolish the Eis-mediated KAN resistance of M. tuberculosis yielded several highly potent agents. A crystal structure of Eis in complex with one of the most potent inhibitors revealed that the inhibitor bound Eis in the AG-binding pocket held by a conformationally malleable region of Eis (residues 28–37) bearing key hydrophobic residues. These Eis inhibitors are promising leads for preclinical development of innovative AG combination therapies against resistant TB.

Mapping of Mycobacterium tuberculosis Complex Genetic Diversity Profiles in Tanzania and Other African Countries

Science.gov (United States)

Mbugi, Erasto V.; Katale, Bugwesa Z.; Streicher, Elizabeth M.; Keyyu, Julius D.; Kendall, Sharon L.; Dockrell, Hazel M.; Michel, Anita L.; Rweyemamu, Mark M.; Warren, Robin M.; Matee, Mecky I.; van Helden, Paul D.; Couvin, David; Rastogi, Nalin

2016-01-01

The aim of this study was to assess and characterize Mycobacterium tuberculosis complex (MTBC) genotypic diversity in Tanzania, as well as in neighbouring East and other several African countries. We used spoligotyping to identify a total of 293 M. tuberculosis clinical isolates (one isolate per patient) collected in the Bunda, Dar es Salaam, Ngorongoro and Serengeti areas in Tanzania. The results were compared with results in the SITVIT2 international database of the Pasteur Institute of Guadeloupe. Genotyping and phylogeographical analyses highlighted the predominance of the CAS, T, EAI, and LAM MTBC lineages in Tanzania. The three most frequent Spoligotype International Types (SITs) were: SIT21/CAS1-Kili (n = 76; 25.94%), SIT59/LAM11-ZWE (n = 22; 7.51%), and SIT126/EAI5 tentatively reclassified as EAI3-TZA (n = 18; 6.14%). Furthermore, three SITs were newly created in this study (SIT4056/EAI5 n = 2, SIT4057/T1 n = 1, and SIT4058/EAI5 n = 1). We noted that the East-African-Indian (EAI) lineage was more predominant in Bunda, the Manu lineage was more common among strains isolated in Ngorongoro, and the Central-Asian (CAS) lineage was more predominant in Dar es Salaam (p-value<0.0001). No statistically significant differences were noted when comparing HIV status of patients vs. major lineages (p-value = 0.103). However, when grouping lineages as Principal Genetic Groups (PGG), we noticed that PGG2/3 group (Haarlem, LAM, S, T, and X) was more associated with HIV-positive patients as compared to PGG1 group (Beijing, CAS, EAI, and Manu) (p-value = 0.03). This study provided mapping of MTBC genetic diversity in Tanzania (containing information on isolates from different cities) and neighbouring East African and other several African countries highlighting differences as regards to MTBC genotypic distribution between Tanzania and other African countries. This work also allowed underlining of spoligotyping patterns tentatively grouped within the newly designated EAI3-TZA

Mapping of Mycobacterium tuberculosis Complex Genetic Diversity Profiles in Tanzania and Other African Countries.

Directory of Open Access Journals (Sweden)

Erasto V Mbugi

Full Text Available The aim of this study was to assess and characterize Mycobacterium tuberculosis complex (MTBC genotypic diversity in Tanzania, as well as in neighbouring East and other several African countries. We used spoligotyping to identify a total of 293 M. tuberculosis clinical isolates (one isolate per patient collected in the Bunda, Dar es Salaam, Ngorongoro and Serengeti areas in Tanzania. The results were compared with results in the SITVIT2 international database of the Pasteur Institute of Guadeloupe. Genotyping and phylogeographical analyses highlighted the predominance of the CAS, T, EAI, and LAM MTBC lineages in Tanzania. The three most frequent Spoligotype International Types (SITs were: SIT21/CAS1-Kili (n = 76; 25.94%, SIT59/LAM11-ZWE (n = 22; 7.51%, and SIT126/EAI5 tentatively reclassified as EAI3-TZA (n = 18; 6.14%. Furthermore, three SITs were newly created in this study (SIT4056/EAI5 n = 2, SIT4057/T1 n = 1, and SIT4058/EAI5 n = 1. We noted that the East-African-Indian (EAI lineage was more predominant in Bunda, the Manu lineage was more common among strains isolated in Ngorongoro, and the Central-Asian (CAS lineage was more predominant in Dar es Salaam (p-value<0.0001. No statistically significant differences were noted when comparing HIV status of patients vs. major lineages (p-value = 0.103. However, when grouping lineages as Principal Genetic Groups (PGG, we noticed that PGG2/3 group (Haarlem, LAM, S, T, and X was more associated with HIV-positive patients as compared to PGG1 group (Beijing, CAS, EAI, and Manu (p-value = 0.03. This study provided mapping of MTBC genetic diversity in Tanzania (containing information on isolates from different cities and neighbouring East African and other several African countries highlighting differences as regards to MTBC genotypic distribution between Tanzania and other African countries. This work also allowed underlining of spoligotyping patterns tentatively grouped within the newly designated EAI3

Mapping of Mycobacterium tuberculosis Complex Genetic Diversity Profiles in Tanzania and Other African Countries.

Science.gov (United States)

Mbugi, Erasto V; Katale, Bugwesa Z; Streicher, Elizabeth M; Keyyu, Julius D; Kendall, Sharon L; Dockrell, Hazel M; Michel, Anita L; Rweyemamu, Mark M; Warren, Robin M; Matee, Mecky I; van Helden, Paul D; Couvin, David; Rastogi, Nalin

2016-01-01

The aim of this study was to assess and characterize Mycobacterium tuberculosis complex (MTBC) genotypic diversity in Tanzania, as well as in neighbouring East and other several African countries. We used spoligotyping to identify a total of 293 M. tuberculosis clinical isolates (one isolate per patient) collected in the Bunda, Dar es Salaam, Ngorongoro and Serengeti areas in Tanzania. The results were compared with results in the SITVIT2 international database of the Pasteur Institute of Guadeloupe. Genotyping and phylogeographical analyses highlighted the predominance of the CAS, T, EAI, and LAM MTBC lineages in Tanzania. The three most frequent Spoligotype International Types (SITs) were: SIT21/CAS1-Kili (n = 76; 25.94%), SIT59/LAM11-ZWE (n = 22; 7.51%), and SIT126/EAI5 tentatively reclassified as EAI3-TZA (n = 18; 6.14%). Furthermore, three SITs were newly created in this study (SIT4056/EAI5 n = 2, SIT4057/T1 n = 1, and SIT4058/EAI5 n = 1). We noted that the East-African-Indian (EAI) lineage was more predominant in Bunda, the Manu lineage was more common among strains isolated in Ngorongoro, and the Central-Asian (CAS) lineage was more predominant in Dar es Salaam (p-value<0.0001). No statistically significant differences were noted when comparing HIV status of patients vs. major lineages (p-value = 0.103). However, when grouping lineages as Principal Genetic Groups (PGG), we noticed that PGG2/3 group (Haarlem, LAM, S, T, and X) was more associated with HIV-positive patients as compared to PGG1 group (Beijing, CAS, EAI, and Manu) (p-value = 0.03). This study provided mapping of MTBC genetic diversity in Tanzania (containing information on isolates from different cities) and neighbouring East African and other several African countries highlighting differences as regards to MTBC genotypic distribution between Tanzania and other African countries. This work also allowed underlining of spoligotyping patterns tentatively grouped within the newly designated EAI3-TZA

Crystallization and preliminary X-ray characterization of phosphoglucose isomerase from Mycobacterium tuberculosis H37Rv

International Nuclear Information System (INIS)

Mathur, Divya; Anand, Kanchan; Mathur, Deepika; Jagadish, Nirmala; Suri, Anil; Garg, Lalit C.

2007-01-01

The phosphoglucose isomerase from Mycobacterium tuberculosis H37Rv was crystallized and diffraction data were collected to 2.8 Å resolution. Phosphoglucose isomerase is a ubiquitous enzyme that catalyzes the isomerization of d-glucopyranose-6-phosphate to d-fructofuranose-6-phosphate. The present investigation reports the expression, purification, crystallization and preliminary crystallographic studies of the phosphoglucose isomerase from Mycobacterium tuberculosis H37Rv, which shares 46% sequence identity with that of its human host. The recombinant protein, which was prepared using an Escherichia coli expression system, was crystallized by the hanging-drop vapour-diffusion method. The crystals diffracted to a resolution of 2.8 Å and belonged to the orthorhombic space group I2 1 2 1 2 1 , with unit-cell parameters a = 109.0, b = 119.8, c = 138.9 Å

Novel genetic polymorphisms that further delineate the phylogeny of the Mycobacterium tuberculosis complex.

NARCIS (Netherlands)

Huard, Richard C; Fabre, Michel; Haas, Petra de; Lazzarini, Luiz Claudio Oliveira; Soolingen, Dick van; Cousins, Debby; Ho, John L

2006-01-01

In a previous report, we described a PCR protocol for the differentiation of the various species of the Mycobacterium tuberculosis complex (MTC) on the basis of genomic deletions (R. C. Huard, L. C. de Oliveira Lazzarini, W. R. Butler, D. van Soolingen, and J. L. Ho, J. Clin. Microbiol.

Gamma Interferon Release Assays for Detection of Mycobacterium tuberculosis Infection

Science.gov (United States)

Denkinger, Claudia M.; Kik, Sandra V.; Rangaka, Molebogeng X.; Zwerling, Alice; Oxlade, Olivia; Metcalfe, John Z.; Cattamanchi, Adithya; Dowdy, David W.; Dheda, Keertan; Banaei, Niaz

2014-01-01

SUMMARY Identification and treatment of latent tuberculosis infection (LTBI) can substantially reduce the risk of developing active disease. However, there is no diagnostic gold standard for LTBI. Two tests are available for identification of LTBI: the tuberculin skin test (TST) and the gamma interferon (IFN-γ) release assay (IGRA). Evidence suggests that both TST and IGRA are acceptable but imperfect tests. They represent indirect markers of Mycobacterium tuberculosis exposure and indicate a cellular immune response to M. tuberculosis. Neither test can accurately differentiate between LTBI and active TB, distinguish reactivation from reinfection, or resolve the various stages within the spectrum of M. tuberculosis infection. Both TST and IGRA have reduced sensitivity in immunocompromised patients and have low predictive value for progression to active TB. To maximize the positive predictive value of existing tests, LTBI screening should be reserved for those who are at sufficiently high risk of progressing to disease. Such high-risk individuals may be identifiable by using multivariable risk prediction models that incorporate test results with risk factors and using serial testing to resolve underlying phenotypes. In the longer term, basic research is necessary to identify highly predictive biomarkers. PMID:24396134

Phosphorylation of InhA inhibits mycolic acid biosynthesis and growth of Mycobacterium tuberculosis

Energy Technology Data Exchange (ETDEWEB)

Molle, Virginie; Gulten, Gulcin; Vilchèze, Catherine; Veyron-Churlet, Romain; Zanella-Cléon, Isabelle; Sacchettini, James C.; Jacobs, Jr, William R.; Kremer, Laurent (CNRS-UMR); (Einstein); (TAM)

2011-08-24

The remarkable survival ability of Mycobacterium tuberculosis in infected hosts is related to the presence of cell wall-associated mycolic acids. Despite their importance, the mechanisms that modulate expression of these lipids in response to environmental changes are unknown. Here we demonstrate that the enoyl-ACP reductase activity of InhA, an essential enzyme of the mycolic acid biosynthetic pathway and the primary target of the anti-tubercular drug isoniazid, is controlled via phosphorylation. Thr-266 is the unique kinase phosphoacceptor, both in vitro and in vivo. The physiological relevance of Thr-266 phosphorylation was demonstrated using inhA phosphoablative (T266A) or phosphomimetic (T266D/E) mutants. Enoyl reductase activity was severely impaired in the mimetic mutants in vitro, as a consequence of a reduced binding affinity to NADH. Importantly, introduction of inhA{_}T266D/E failed to complement growth and mycolic acid defects of an inhA-thermosensitive Mycobacterium smegmatis strain, in a similar manner to what is observed following isoniazid treatment. This study suggests that phosphorylation of InhA may represent an unusual mechanism that allows M. tuberculosis to regulate its mycolic acid content, thus offering a new approach to future anti-tuberculosis drug development.

Phosphorylation of InhA inhibits mycolic acid biosynthesis and growth of Mycobacterium tuberculosis.

Science.gov (United States)

Molle, Virginie; Gulten, Gulcin; Vilchèze, Catherine; Veyron-Churlet, Romain; Zanella-Cléon, Isabelle; Sacchettini, James C; Jacobs, William R; Kremer, Laurent

2010-12-01

The remarkable survival ability of Mycobacterium tuberculosis in infected hosts is related to the presence of cell wall-associated mycolic acids. Despite their importance, the mechanisms that modulate expression of these lipids in response to environmental changes are unknown. Here we demonstrate that the enoyl-ACP reductase activity of InhA, an essential enzyme of the mycolic acid biosynthetic pathway and the primary target of the anti-tubercular drug isoniazid, is controlled via phosphorylation. Thr-266 is the unique kinase phosphoacceptor, both in vitro and in vivo. The physiological relevance of Thr-266 phosphorylation was demonstrated using inhA phosphoablative (T266A) or phosphomimetic (T266D/E) mutants. Enoyl reductase activity was severely impaired in the mimetic mutants in vitro, as a consequence of a reduced binding affinity to NADH. Importantly, introduction of inhA_T266D/E failed to complement growth and mycolic acid defects of an inhA-thermosensitive Mycobacterium smegmatis strain, in a similar manner to what is observed following isoniazid treatment. This study suggests that phosphorylation of InhA may represent an unusual mechanism that allows M. tuberculosis to regulate its mycolic acid content, thus offering a new approach to future anti-tuberculosis drug development. © 2010 Blackwell Publishing Ltd.

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

Bim is a crucial regulator of apoptosis induced by Mycobacterium tuberculosis

Science.gov (United States)

Aguiló, N; Uranga, S; Marinova, D; Martín, C; Pardo, J

2014-01-01

Mycobacterium tuberculosis, the causative agent of tuberculosis, induces apoptosis in infected macrophages in vitro and in vivo. However, the molecular mechanism controlling this process is not known. In order to study the involvement of the mitochondrial apoptotic pathway in M. tuberculosis-induced apoptosis, we analysed cell death in M. tuberculosis-infected embryonic fibroblasts (MEFs) derived from different knockout mice for genes involved in this route. We found that apoptosis induced by M. tuberculosis is abrogated in the absence of Bak and Bax, caspase 9 or the executioner caspases 3 and 7. Notably, we show that MEF deficient in the BH3-only BCL-2-interacting mediator of cell death (Bim) protein were also resistant to this process. The relevance of these results has been confirmed in the mouse macrophage cell line J774, where cell transfection with siRNA targeting Bim impaired apoptosis induced by virulent mycobacteria. Notably, only infection with a virulent strain, but not with attenuated ESX-1-defective strains, such as Bacillus Calmette-Guerin and live-attenuated M. tuberculosis vaccine strain MTBVAC, induced Bim upregulation and apoptosis, probably implicating virulence factor early secreted antigenic target 6-kDa protein in this process. Our results suggest that Bim upregulation and apoptosis is mediated by the p38MAPK-dependent pathway. Our findings show that Bim is a master regulator of apoptosis induced by M. tuberculosis. PMID:25032866

A broad set of different llama antibodies specific for a 16 kDa heat shock protein of Mycobacterium tuberculosis

NARCIS (Netherlands)

Trilling, Anke K.; de Ronde, Hans; Noteboom, Linda; van Houwelingen, Adèle; Roelse, Margriet; Srivastava, Saurabh K.; Haasnoot, Willem; Jongsma, Maarten A.; Kolk, Arend; Zuilhof, Han; Beekwilder, Jules

2011-01-01

Recombinant antibodies are powerful tools in engineering of novel diagnostics. Due to the small size and stable nature of llama antibody domains selected antibodies can serve as a detection reagent in multiplexed and sensitive assays for M. tuberculosis. Antibodies for Mycobacterium tuberculosis (M.

Supplementary Material for: Whole genome sequencing reveals genomic heterogeneity and antibiotic purification in Mycobacterium tuberculosis isolates

KAUST Repository

Black, PA; Vos, M. de; Louw, GE; Merwe, RG van der; Dippenaar, A.; Streicher, EM; Abdallah, AM; Sampson, SL; Victor, TC; Dolby, T.; Simpson, JA; Helden, PD van; Warren, RM; Pain, Arnab

2015-01-01

Abstract Background Whole genome sequencing has revolutionised the interrogation of mycobacterial genomes. Recent studies have reported conflicting findings on the genomic stability of Mycobacterium tuberculosis during the evolution of drug

A Rational Approach to Identify Inhibitors of Mycobacterium tuberculosis Enoyl Acyl Carrier Protein Reductase

Czech Academy of Sciences Publication Activity Database

Chhabria, M. T.; Parmar, K. B.; Brahmkshatriya, Pathik

2013-01-01

Roč. 19, č. 21 (2013), s. 3878-3883 ISSN 1381-6128 Institutional support: RVO:61388963 Keywords : mycobacterium tuberculosis * enoyl acyl carrier protein reductase * pharmacophore modeling * molecular docking * binding interactions Subject RIV: FR - Pharmacology ; Medidal Chemistry Impact factor: 3.288, year: 2013

Proteomic profiling of Mycobacterium tuberculosis identifies nutrient-starvation-responsive toxin-antitoxin systems

DEFF Research Database (Denmark)

Albrethsen, Jakob; Agner, Jeppe; Piersma, Sander R

2013-01-01

In order to successfully enter the latent stage, Mycobacterium tuberculosis must adapt to conditions such as nutrient limitation and hypoxia. In vitro models that mimic latent infection are valuable tools for describing the changes in metabolism that occur when the bacterium exists in a non......-growing form. We used two complementary proteomic approaches, label-free LC-MS/MS analysis and two-dimensional difference gel electrophoresis, to determine the proteome profile of extracellular proteins from M. tuberculosis cultured under nutrient starvation. Through the label-free LC-MS/MS analysis......, significant differences in the overall metabolism during nutrient starvation were detected. Notably, members of the toxin-antitoxin systems were present in larger quantities in nutrient-starved cultures, supporting a role for these global modules as M. tuberculosis switches its metabolism into dormancy...

Capsular glucan and intracellular glycogen of Mycobacterium tuberculosis: biosynthesis and impact on the persistence in mice

DEFF Research Database (Denmark)

Sambou, Tounkang; Dinadayala, Premkumar; Stadthagen, Gustavo

2008-01-01

Mycobacterium tuberculosis and other pathogenic mycobacterial species produce large amounts of a glycogen-like alpha-glucan that represents the major polysaccharide of their outermost capsular layer. To determine the role of the surface-exposed glucan in the physiology and virulence of these bact......Mycobacterium tuberculosis and other pathogenic mycobacterial species produce large amounts of a glycogen-like alpha-glucan that represents the major polysaccharide of their outermost capsular layer. To determine the role of the surface-exposed glucan in the physiology and virulence...... of these bacteria, orthologues of the glg genes involved in the biosynthesis of glycogen in Escherichia coli were identified in M. tuberculosis H37Rv and inactivated by allelic replacement. Biochemical analyses of the mutants and complemented strains indicated that the synthesis of glucan and glycogen involves...... the alpha-1,4-glucosyltransferases Rv3032 and GlgA (Rv1212c), the ADP-glucose pyrophosphorylase GlgC (Rv1213) and the branching enzyme GlgB (Rv1326c). Disruption of glgC reduced by half the glucan and glycogen contents of M. tuberculosis, whereas the inactivation of glgA and Rv3032 affected the production...

Clustering of Mycobacterium tuberculosis Cases in Acapulco: Spoligotyping and Risk Factors

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Elizabeth Nava-Aguilera

2011-01-01

Full Text Available Recurrence and reinfection of tuberculosis have quite different implications for prevention. We identified 267 spoligotypes of Mycobacterium tuberculosis from consecutive tuberculosis patients in Acapulco, Mexico, to assess the level of clustering and risk factors for clustered strains. Point cluster analysis examined spatial clustering. Risk analysis relied on the Mantel Haenszel procedure to examine bivariate associations, then to develop risk profiles of combinations of risk factors. Supplementary analysis of the spoligotyping data used SpolTools. Spoligotyping identified 85 types, 50 of them previously unreported. The five most common spoligotypes accounted for 55% of tuberculosis cases. One cluster of 70 patients (26% of the series produced a single spoligotype from the Manila Family (Clade EAI2. The high proportion (78% of patients infected with cluster strains is compatible with recent transmission of TB in Acapulco. Geomatic analysis showed no spatial clustering; clustering was associated with a risk profile of uneducated cases who lived in single-room dwellings. The Manila emerging strain accounted for one in every four cases, confirming that one strain can predominate in a hyperendemic area.

[Tuberculosis caused by Mycobacterium bovis in workers of bovine tuberculosis sanitation farms in Antioquia, Boyacá and Cundinamarca].

Science.gov (United States)

Leal-Bohórquez, Andrés F; Castro-Osorio, Claudia M; Wintaco-Martínez, Luz M; Villalobos, Rafael; Puerto-Castro, Gloria M

2016-01-01

To perform classic and molecular epidemiological surveillance of human tuberculosis caused by Mycobacterium bovis in bovine supply chains at farms with PPD positive bovines in the departments of Antioquia, Boyacá and Cundinamarca during a one-year period. Livestock farms with PPD positive bovines or buffalos were visited in the study departments according to information obtained in the "Programa Nacional de Tuberculosis bovina" (National program on bovine Tuberculosis) released by ICA (Colombian Agriculture and Livestock Institute). Data on socio-demographic information and tuberculosis risk factors associated to the occupation were collected through a survey applied to all workers at the visited farms. Sputum samples were obtained after informed consent. The sputa underwent microbiological and molecular testing to identify members of the M. tuberculosis complex. Thirty-three livestock farms were visited and information of 164 workers from the bovine supply chain was collected. Staying in a PPD positive farm for more than a year, ignorance about the disease and the presence of possible vectors, like dogs and cats, were identified as possible risk factors for developing tuberculosis. No cases of tuberculosis caused by M. bovis or M. tuberculosis in workers of the visited farms were found. No cases of the disease caused by this zoonotic agent were documented in the departments of Antioquia, Boyacá and Cundinamarca.

Management of latent Mycobacterium tuberculosis infection: WHO guidelines for low tuberculosis burden countries.

Science.gov (United States)

Getahun, Haileyesus; Matteelli, Alberto; Abubakar, Ibrahim; Aziz, Mohamed Abdel; Baddeley, Annabel; Barreira, Draurio; Den Boon, Saskia; Borroto Gutierrez, Susana Marta; Bruchfeld, Judith; Burhan, Erlina; Cavalcante, Solange; Cedillos, Rolando; Chaisson, Richard; Chee, Cynthia Bin-Eng; Chesire, Lucy; Corbett, Elizabeth; Dara, Masoud; Denholm, Justin; de Vries, Gerard; Falzon, Dennis; Ford, Nathan; Gale-Rowe, Margaret; Gilpin, Chris; Girardi, Enrico; Go, Un-Yeong; Govindasamy, Darshini; D Grant, Alison; Grzemska, Malgorzata; Harris, Ross; Horsburgh, C Robert; Ismayilov, Asker; Jaramillo, Ernesto; Kik, Sandra; Kranzer, Katharina; Lienhardt, Christian; LoBue, Philip; Lönnroth, Knut; Marks, Guy; Menzies, Dick; Migliori, Giovanni Battista; Mosca, Davide; Mukadi, Ya Diul; Mwinga, Alwyn; Nelson, Lisa; Nishikiori, Nobuyuki; Oordt-Speets, Anouk; Rangaka, Molebogeng Xheedha; Reis, Andreas; Rotz, Lisa; Sandgren, Andreas; Sañé Schepisi, Monica; Schünemann, Holger J; Sharma, Surender Kumar; Sotgiu, Giovanni; Stagg, Helen R; Sterling, Timothy R; Tayeb, Tamara; Uplekar, Mukund; van der Werf, Marieke J; Vandevelde, Wim; van Kessel, Femke; van't Hoog, Anna; Varma, Jay K; Vezhnina, Natalia; Voniatis, Constantia; Vonk Noordegraaf-Schouten, Marije; Weil, Diana; Weyer, Karin; Wilkinson, Robert John; Yoshiyama, Takashi; Zellweger, Jean Pierre; Raviglione, Mario

2015-12-01

Latent tuberculosis infection (LTBI) is characterised by the presence of immune responses to previously acquired Mycobacterium tuberculosis infection without clinical evidence of active tuberculosis (TB). Here we report evidence-based guidelines from the World Health Organization for a public health approach to the management of LTBI in high risk individuals in countries with high or middle upper income and TB incidence of <100 per 100 000 per year. The guidelines strongly recommend systematic testing and treatment of LTBI in people living with HIV, adult and child contacts of pulmonary TB cases, patients initiating anti-tumour necrosis factor treatment, patients receiving dialysis, patients preparing for organ or haematological transplantation, and patients with silicosis. In prisoners, healthcare workers, immigrants from high TB burden countries, homeless persons and illicit drug users, systematic testing and treatment of LTBI is conditionally recommended, according to TB epidemiology and resource availability. Either commercial interferon-gamma release assays or Mantoux tuberculin skin testing could be used to test for LTBI. Chest radiography should be performed before LTBI treatment to rule out active TB disease. Recommended treatment regimens for LTBI include: 6 or 9 month isoniazid; 12 week rifapentine plus isoniazid; 3-4 month isoniazid plus rifampicin; or 3-4 month rifampicin alone. Copyright ©ERS 2015.

Mycobacterium tuberculosis exploits asparagine to assimilate nitrogen and resist acid stress during infection.

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Alexandre Gouzy

2014-02-01

Full Text Available Mycobacterium tuberculosis is an intracellular pathogen. Within macrophages, M. tuberculosis thrives in a specialized membrane-bound vacuole, the phagosome, whose pH is slightly acidic, and where access to nutrients is limited. Understanding how the bacillus extracts and incorporates nutrients from its host may help develop novel strategies to combat tuberculosis. Here we show that M. tuberculosis employs the asparagine transporter AnsP2 and the secreted asparaginase AnsA to assimilate nitrogen and resist acid stress through asparagine hydrolysis and ammonia release. While the role of AnsP2 is partially spared by yet to be identified transporter(s, that of AnsA is crucial in both phagosome acidification arrest and intracellular replication, as an M. tuberculosis mutant lacking this asparaginase is ultimately attenuated in macrophages and in mice. Our study provides yet another example of the intimate link between physiology and virulence in the tubercle bacillus, and identifies a novel pathway to be targeted for therapeutic purposes.

Usefulness of the secondary probe pTBN12 in DNA fingerprinting of Mycobacterium tuberculosis.

OpenAIRE

Chaves, F; Yang, Z; el Hajj, H; Alonso, M; Burman, W J; Eisenach, K D; Dronda, F; Bates, J H; Cave, M D

1996-01-01

A comparison was made between DNA fingerprints of Mycobacterium tuberculosis produced with the insertion sequence IS6110 and those produced with the polymorphic GC-rich repetitive sequence contained in the plasmid pTBN12. A total of 302 M. tuberculosis isolates from the prison system in Madrid, Spain, and the Denver Public Health Department (Denver, Colo.) were analyzed with the two probes. Both probes identified the same isolates in the same clusters when the fingerprints had six or more cop...

Mycobacterium tuberculosis, but not vaccine BCG, specifically upregulates matrix metalloproteinase-1.

Science.gov (United States)

Elkington, Paul T G; Nuttall, Robert K; Boyle, Joseph J; O'Kane, Cecilia M; Horncastle, Donna E; Edwards, Dylan R; Friedland, Jon S

2005-12-15

Pulmonary cavitation is fundamental to the global success of Mycobacterium tuberculosis. However, the mechanisms of this lung destruction are poorly understood. The biochemistry of lung matrix predicts matrix metalloproteinase (MMP) involvement in immunopathology. We investigated gene expression of all MMPs, proteins with a disintegrin and metalloproteinase domain, and tissue inhibitors of metalloproteinases in M. tuberculosis-infected human macrophages by real-time polymerase chain reaction. MMP secretion was measured by zymography and Western analysis, and expression in patients with pulmonary tuberculosis was localized by immunohistochemistry. MMP-1 and MMP-7 gene expression and secretion are potently upregulated by M. tuberculosis, and no increase in tissue inhibitor of metalloproteinase expression occurs to oppose their activity. Dexamethasone completely suppresses MMP-1 but not MMP-7 gene expression and secretion. In patients with active tuberculosis, macrophages express MMP-1 and MMP-7 adjacent to areas of tissue destruction. MMP-1 but not MMP-7 expression and secretion are relatively M. tuberculosis specific, are not upregulated by tuberculosis-associated cytokines, and are prostaglandin dependent. In contrast, the vaccine M. bovis bacillus Calmette-Guérin (BCG) does not stimulate MMP-1 secretion from human macrophages, although M. tuberculosis and BCG do upregulate MMP-7 equally. BCG-infected macrophages secrete reduced prostaglandin E2 concentrations compared with M. tuberculosis-infected macrophages, and prostaglandin pathway supplementation augments MMP-1 secretion from BCG-infected cells. M. tuberculosis specifically upregulates MMP-1 in a cellular model of human infection and in patients with tuberculosis. In contrast, vaccine BCG, which does not cause lung cavitation, does not upregulate prostaglandin E2-dependent MMP-1 secretion.

PREVALÊNCIA DA Mycobacterium tuberculosis NO COMPLEXO PRISIONAL DO MUNICÍPIO DE ITAPERUNA, RJ

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Francislene FERREIRA SILVA

2015-12-01

Full Text Available A presença da tuberculose em sistemas prisionais é descrito como uma ameaça a saúde pública. O presente estudo objetivou a determinar a prevalência da Mycobacterium tuberculosis no sistema prisional Diomedes Vinhosa Muniz em Itaperuna, Rio de Janeiro. O estudo foi submetido e aprovado pelo comitê de ética em pesquisa da Faculdade Redentor em Itaperuna e autorizado pela Secretaria de Estado de Administração Penitenciária (SEAP do estado do Rio de Janeiro. Participaram da pesquisa 10% do total de detentos condenados, estes assinaram um termo de consentimento livre e esclarecido, preencheram um questionário sócio econômico e epidemiológico em seguida foi coletado o material biológico para realização da baciloscopia. Os resultados indicaram que 69% dos detentos analisados não completaram o ensino fundamental, 30% é originário de comunidades faveladas, 55% possuem uma renda mensal de até 2 salários mínimos, 79% consumiam tabaco, 64% relataram que tiveram tosse por mais de 3 semanas, 65% declaram a maconha como a droga ilícita preferencial e 51% usavam cocaína, 3% declararam se portador do HIV, 42% tiveram contato com indivíduos com a tuberculose e 3% dos detentos haviam a presença da Mycobacterium tuberculosis. Logo, conclui-se que no sistema prisional Diomedes Vinhosa Muniz houve uma baixa prevalência da M.ycobacterium tuberculosis.

Human tuberculosis caused by Mycobacterium bovis: a retrospective comparison with Mycobacterium tuberculosis in a Mexican tertiary care centre, 2000–2015

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Pedro Torres-Gonzalez

2016-11-01

Full Text Available Abstract Background Human tuberculosis caused by Mycobacterium bovis is believed to be frequent in developing countries. Transmission is usually through ingestion of unpasteurized dairy products, although airborne contagion is possible. Disease caused by M. tuberculosis or M. bovis is clinically indistinguishable from each other. The aim of this study was to determine the factors associated with M. bovis disease. Methods Retrospective analysis of all culture-positive cases of M. bovis and M. tuberculosis from 2000 to 2015, in a Mexican tertiary-care centre. Sociodemographic, clinical, and radiographic data from medical records were compared. Disease site was classified as pulmonary, extrapulmonary, or pulmonary and extrapulmonary, based on cultures. Results We evaluated 533 cases, 372 (69.7 % of which were caused by M. tuberculosis and 161 (30.2 % by M. bovis. Characteristics associated with M. bovis disease were: younger age (aOR 0.97, 95 % CI 0.95–0.98, glucocorticoid use (aOR 2.27, 95 % CI 1.42–3.63, and extrapulmonary disease (aOR 1.80, 95 % CI 1.21–2.69. M. tuberculosis was associated with lower socioeconomic status (aOR 0.52, 95 % CI 0.28–0.97. When we analysed only pulmonary cases, younger age (aOR 0.97, 95 % CI 0.96–0.99, glucocorticoid use (aOR 2.41, 95 % CI 1.30–4.46, and smoking (aOR 1.94, CI 95 % 1.15–3.27 were associated with M. bovis. Both groups showed similar proportions of direct microscopy smear results (respiratory samples and chest X-ray cavitations. Conclusions Younger age, glucocorticoid use, and extrapulmonary disease were associated with M. bovis as the causative agent of tuberculosis in a group of patients from a tertiary care centre in a country where bovine tuberculosis is endemic. Further studies must be conducted in the general population to determine pathogen-specific associated factors and outcomes.

Dendritic Cells Activate and Mature after Infection with Mycobacterium tuberculosis

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Mamo Gezahagne

2011-07-01

Full Text Available Abstract Background Dendritic cells (DCs can take up an array of different antigens, including microorganisms which they can process and present more effectively than any other antigen presenting cell. However, whether the interaction between the human DC and Mycobacterium tuberculosis represents a defense mechanism by the invaded host, or helping the invader to evade the defense mechanism of the host is still not clearly understood. Findings To analyze the interactions between M. tuberculosis and immune cells, human peripheral blood monocyte-derived immature DCs were infected with M. tuberculosis H37Rv wild type strain and flow cytometry was used to analyse cell surface expression markers. The ability of the M. tuberculosis infected DC to induce T cell proliferation using 5 and 6-carboxyfluorescein diacetate succinimidyl ester (CFSE dilution technique was also investigated. DCs were found to internalize the mycobacteria and show dose dependent infection and necrosis with different multiplicity of infection. Flow cytometry analysis of cell surface expression markers CD40, CD54, CD80, CD83, CD86 and HLA DR in infected DC revealed significant (p M. tuberculosis in comparison to immature DC with no stimulation. Lipopolysaccharide (LPS from Salmonella abortus equi, a known DC maturation agent, was used as a positive control and showed a comparable up regulation of cell surface markers as observed with M. tuberculosis infected DC. It was revealed that the M. tuberculosis infected DC induced T cell proliferation. Conclusion These data clearly demonstrate that M. tuberculosis induces activation and maturation of human monocyte-derived immature DC as well as induces T cell proliferation in vitro.

Studies on mycobacterium tuberculosis sensitivity test by using the method of rapid radiometry with appendixes of clinical results

International Nuclear Information System (INIS)

Yang Yongqing; Jiang Yimin; Lu Wendong; Zhu Rongen

1987-01-01

Three standard strains of mycobacterium tuberculosis (H 37 RV-fully sensitive, SM-R1000 μg/ml, RFP-R 100 μg/ml) were tested with 10 concentration of 5 antitubercular agent, INH, SM, PAS, RFP and EB. 114 isolates of mycobacterium tuberculosis taken from patients were tested with INH, PAS, SM and RFP. They were agreed with the results of standard Lowenstein-Jensen method in 81.7%. 82% of the isolate test were completed within 5 days. The method may be used in routine clinical work. The liquid media prepared by authors do not require human serum albumin and it is less expensive and readily available

The endothelin system has a significant role in the pathogenesis and progression of Mycobacterium tuberculosis infection.

Science.gov (United States)

Correa, Andre F; Bailão, Alexandre M; Bastos, Izabela M D; Orme, Ian M; Soares, Célia M A; Kipnis, Andre; Santana, Jaime M; Junqueira-Kipnis, Ana Paula

2014-12-01

Tuberculosis (TB) remains a major global health problem, and although multiple studies have addressed the relationship between Mycobacterium tuberculosis and the host on an immunological level, few studies have addressed the impact of host physiological responses. Proteases produced by bacteria have been associated with important alterations in the host tissues, and a limited number of these enzymes have been characterized in mycobacterial species. M. tuberculosis produces a protease called Zmp1, which appears to be associated with virulence and has a putative action as an endothelin-converting enzyme. Endothelins are a family of vasoactive peptides, of which 3 distinct isoforms exist, and endothelin 1 (ET-1) is the most abundant and the best-characterized isoform. The aim of this work was to characterize the Zmp1 protease and evaluate its role in pathogenicity. Here, we have shown that M. tuberculosis produces and secretes an enzyme with ET-1 cleavage activity. These data demonstrate a possible role of Zmp1 for mycobacterium-host interactions and highlights its potential as a drug target. Moreover, the results suggest that endothelin pathways have a role in the pathogenesis of M. tuberculosis infections, and ETA or ETB receptor signaling can modulate the host response to the infection. We hypothesize that a balance between Zmp1 control of ET-1 levels and ETA/ETB signaling can allow M. tuberculosis adaptation and survival in the lung tissues. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Development of a Hierarchical Variable-Number Tandem Repeat Typing Scheme for Mycobacterium tuberculosis in China

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Luo, Tao; Yang, Chongguang; Pang, Yu; Zhao, Yanlin; Mei, Jian; Gao, Qian

2014-01-01

Molecular typing based on variable-number tandem repeats (VNTR) analysis is a promising tool for identifying transmission of Mycobacterium tuberculosis. However, the currently proposed 15- and 24-locus VNTR sets (VNTR-15/24) only have limited resolution and contain too many loci for large-scale typing in high burden countries. To develop an optimal typing scheme in China, we evaluated the resolution and robustness of 25 VNTR loci, using population-based collections of 1362 clinical isolates from six provinces across the country. The resolution of most loci showed considerable variations among regions. By calculating the average resolution of all possible combinations of 20 robust loci, we identified an optimal locus set with a minimum of 9 loci (VNTR-9) that could achieve comparable resolution of the standard VNTR-15. The VNTR-9 had consistently high resolutions in all six regions, and it was highly concordant with VNTR-15 for defining both clustered and unique genotypes. Furthermore, VNTR-9 was phylogenetically informative for classifying lineages/sublineages of M. tuberculosis. Three hypervariable loci (HV-3), VNTR 3232, VNTR 3820 and VNTR 4120, were proved important for further differentiating unrelated clustered strains based on VNTR-9. We propose the optimized VNTR-9 as first-line method and the HV-3 as second-line method for molecular typing of M. tuberculosis in China and surrounding countries. The development of hierarchical VNTR typing methods that can achieve high resolution with a small number of loci could be suitable for molecular epidemiology study in other high burden countries. PMID:24586989

A first insight on the population structure of Mycobacterium tuberculosis complex as studied by spoligotyping and MIRU-VNTRs in Santiago, Chile.

Science.gov (United States)

Balcells, María Elvira; García, Patricia; Meza, Paulina; Peña, Carlos; Cifuentes, Marcela; Couvin, David; Rastogi, Nalin

2015-01-01

Tuberculosis (TB) remains a significant public health problem worldwide, but the ecology of the prevalent mycobacterial strains, and their transmission, can vary depending on country and region. Chile is a country with low incidence of TB, that has a geographically isolated location in relation to the rest of South American countries due to the Andes Mountains, but recent migration from neighboring countries has changed this situation. We aimed to assess the genotypic diversity of Mycobacterium tuberculosis complex (MTBC) strains in Santiago, Chile, and compare with reports from other Latin-American countries. We analyzed MTBC isolates from pulmonary tuberculosis cases collected between years 2008 and 2013 in Central Santiago, using two genotyping methods: spoligotyping and 12-loci mycobacterial interspersed repetitive unit-variable number of tandem repeats (MIRU-VNTRs). Data obtained were analyzed and compared to the SITVIT2 database. Mean age of the patients was 47.5 years and 61% were male; 11.6% were migrants. Of 103 strains (1 isolate/patient) included, there were 56 distinct spoligotype patterns. Of these, 16 strains (15.5%) corresponded to orphan strains in the SITVIT2 database, not previously reported. Latin American and Mediterranean (LAM) (34%) and T (33%) lineages were the most prevalent strains, followed by Haarlem lineage (16.5%). Beijing family was scarcely represented with only two cases (1.9%), one of them isolated from a Peruvian migrant. The most frequent clustered spoligotypes were SIT33/LAM3 (10.7%), SIT53/T1 (8.7%), SIT50/H3 (7.8%), and SIT37/T3 (6.8%). We conclude that LAM and T genotypes are the most prevalent genotypes of MTBC in Santiago, Chile, and together correspond to almost two thirds of analyzed strains, which is similar to strain distribution reported from other countries of Latin America. Nevertheless, the high proportion of SIT37/T3, which was rarely found in other Latin American countries, may underline a specific history or

Diversity of DNA fingerprints of Mycobacterium tuberculosis isolates in the United States.

Science.gov (United States)

Yang, Z; Barnes, P F; Chaves, F; Eisenach, K D; Weis, S E; Bates, J H; Cave, M D

1998-04-01

To investigate the diversity of IS6110 fingerprints of Mycobacterium tuberculosis isolates in the United States and to determine if matching IS6110 fingerprints represent recent interstate tuberculosis transmission, we performed restriction fragment length polymorphism analysis of M. tuberculosis isolates from 1,326 patients in three geographically separated states. Seven hundred ninety-five different IS6110 fingerprint patterns were generated, and pattern diversity was similar in each state. Ninety-six percent of the fingerprint patterns were observed in only one state, demonstrating that most IS6110 fingerprint patterns are confined to a single geographic location. Of the IS6110 fingerprint patterns that were shared by isolates from more than one state, most isolates with 1 to 5 IS6110 copies were separable by pTBN12 fingerprinting whereas those with > 15 copies were not. One high-copy-number M. tuberculosis strain had identical IS6110 and pTBN12 fingerprints and included 57 isolates from three states. Epidemiological data demonstrated significant recent transmission of tuberculosis within each city but not among the states. This suggests that identical fingerprints of isolates from geographically separate locations most likely reflect interstate tuberculosis transmission in the past, with subsequent intrastate spread of disease. Further evaluation of M. tuberculosis strains that cause outbreaks in different geographic locations will provide insight into the epidemiological and bacteriological factors that facilitate the spread of tuberculosis.

Cutaneous squamous cell carcinoma in lupus vulgaris caused by drug resistant Mycobacterium tuberculosis

Directory of Open Access Journals (Sweden)

Muthu S Kumaran

2017-01-01

Full Text Available Tuberculosis (TB is still a major public health problem in the world, with many factors contributing to this burden, including poor living conditions, overcrowding, poverty, malnutrition, illiteracy, and rapid spread of human immunodeficiency virus infection. Cutaneous tuberculosis is a less common form of extrapulmonary tuberculosis, and in this paucibacillary form the diagnosis depends on histopathology, tuberculin positivity, and response to treatment. The diagnosis is even more difficult in cases with drug resistant Mycobacterium tuberculosis due to lack of awareness and lack of facilities to diagnose drug resistant tuberculosis. In this article, we describe an unusual case of multidrug resistant lupus vulgaris (LV, in a 34-year-old male who responded to anti-tubercular treatment (ATT initially, but developed recurrent disease which failed to respond to standard four-drug ATT; subsequently, tissue culture showed growth of multidrug resistant M. tuberculosis. Subsequently, he also developed cutaneous squamous cell carcinoma. This article aims to exemplify a grave complication that can occur in long-standing case of LV, the limitations faced by clinicians in developing countries where tuberculosis is endemic, and classical methods of proving drug resistance are generally unavailable or fail.

Anti-Mycobacterium tuberculosis activity of fungus Phomopsis stipata

Directory of Open Access Journals (Sweden)

Karina Andrade de Prince

2012-03-01

Full Text Available Our purpose was to determine the anti-Mycobacterium tuberculosis activity of the metabolites produced by the endophitic fungus Phomopsis stipata (Lib. B. Sutton, (Diaporthaceae, cultivated in different media. The antimycobacterial activity was assessed through the Resazurin Microtiter Assay (REMA and the cytotoxicity test performed on macrophage cell line. The extracts derived from fungi grown on Corn Medium and Potato Dextrose Broth presented the smallest values of Minimum Inhibitory Concentration (MIC and low cytotoxicity, which implies a high selectivity index. This is the first report on the chemical composition and antitubercular activity of metabolites of P. stipata, as well as the influence of culture medium on these properties.

Development of a One-Step Multiplex PCR Assay for Differential Detection of Major Mycobacterium Species.

Science.gov (United States)

Chae, Hansong; Han, Seung Jung; Kim, Su-Young; Ki, Chang-Seok; Huh, Hee Jae; Yong, Dongeun; Koh, Won-Jung; Shin, Sung Jae

2017-09-01

The prevalence of tuberculosis continues to be high, and nontuberculous mycobacterial (NTM) infection has also emerged worldwide. Moreover, differential and accurate identification of mycobacteria to the species or subspecies level is an unmet clinical need. Here, we developed a one-step multiplex PCR assay using whole-genome analysis and bioinformatics to identify novel molecular targets. The aims of this assay were to (i) discriminate between the Mycobacterium tuberculosis complex (MTBC) and NTM using rv0577 or RD750, (ii) differentiate M. tuberculosis ( M. tuberculosis ) from MTBC using RD9, (iii) selectively identify the widespread M. tuberculosis Beijing genotype by targeting mtbk_20680 , and (iv) simultaneously detect five clinically important NTM ( M. avium , M. intracellulare , M. abscessus , M. massiliense , and M. kansasii ) by targeting IS 1311 , DT1, mass_3210 , and mkan_rs12360 An initial evaluation of the multiplex PCR assay using reference strains demonstrated 100% specificity for the targeted Mycobacterium species. Analytical sensitivity ranged from 1 to 10 pg for extracted DNA and was 10 3 and 10 4 CFU for pure cultures and nonhomogenized artificial sputum cultures, respectively, of the targeted species. The accuracy of the multiplex PCR assay was further evaluated using 55 reference strains and 94 mycobacterial clinical isolates. Spoligotyping, multilocus sequence analysis, and a commercial real-time PCR assay were employed as standard assays to evaluate the multiplex PCR assay with clinical M. tuberculosis and NTM isolates. The PCR assay displayed 100% identification agreement with the standard assays. Our multiplex PCR assay is a simple, convenient, and reliable technique for differential identification of MTBC, M. tuberculosis , M. tuberculosis Beijing genotype, and major NTM species. Copyright © 2017 American Society for Microbiology.

Structural and Functional Aspects of the Sensor Histidine Kinase PrrB from Mycobacterium tuberculosis

DEFF Research Database (Denmark)

Nowak, E.; Panjikar, S.; Morth, J.P.

2006-01-01

We describe the solution structures of two- and three-domain constructs of the sensor histidine kinase PrrB from Mycobacterium tuberculosis, which allow us to locate the HAMP linker relative to the ATP binding and dimerization domains. We show that the three-domain construct is active both...

T-cell recognition of Mycobacterium tuberculosis culture filtrate fractions in tuberculosis patients and their household contacts

DEFF Research Database (Denmark)

Demissie, A; Ravn, P; Olobo, J

1999-01-01

We examined the immune responses of patients with active pulmonary tuberculosis (TB) and their healthy household contacts to short-term culture filtrate (ST-CF) of Mycobacterium tuberculosis or molecular mass fractions derived from it. Our goal was to identify fractions strongly recognized...... antigens and immune responses were determined. Household contacts produced significantly higher levels of gamma interferon (IFN-gamma) than the TB patients in response to antigens present in ST-CF and the 10 narrow-molecular-mass fractions. A similar difference in leukocyte proliferative responses...... to the antigens between the two groups was also found. In general, while all fractions stimulated immune responses, the highest activity was seen with the low-molecular-mass fractions, which include well-defined TB antigens such as ESAT-6. Leukocytes from contacts of TB patients with severe disease produced...