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.

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

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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

Diversity of DNA fingerprints of Mycobacterium tuberculosis isolates in the United States.

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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.

Microevolution of Mycobacterium tuberculosis in a tuberculosis patient.

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Al-Hajoj, S.A.; Akkerman, O.; Parwati, I.; Al-Gamdi, S.; Rahim, Z.; Soolingen, D. van; Ingen, J. van; Supply, P.; Zanden, A.G. van der

2010-01-01

Five Mycobacterium tuberculosis isolates were obtained from three body sites from a Dutch patient. The isolates displayed a single genotype by 24-locus MIRU-VNTR typing (except for a single locus not amplified from one isolate) but were differentiated by small variations in IS6110 fingerprints,

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 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.

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.

Genomic diversity among Beijing and non-Beijing Mycobacterium tuberculosis isolates from Myanmar.

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Ruth Stavrum

2008-04-01

Full Text Available The Beijing family of Mycobacterium tuberculosis is dominant in countries in East Asia. Genomic polymorphisms are a source of diversity within the M. tuberculosis genome and may account for the variation of virulence among M. tuberculosis isolates. Till date there are no studies that have examined the genomic composition of M. tuberculosis isolates from the high TB-burden country, Myanmar.Twenty-two M. tuberculosis isolates from Myanmar were screened on whole-genome arrays containing genes from M. tuberculosis H37Rv, M. tuberculosis CDC1551 and M. bovis AF22197. Screening identified 198 deletions or extra regions in the clinical isolates compared to H37Rv. Twenty-two regions differentiated between Beijing and non-Beijing isolates and were verified by PCR on an additional 40 isolates. Six regions (Rv0071-0074 [RD105], Rv1572-1576c [RD149], Rv1585c-1587c [RD149], MT1798-Rv1755c [RD152], Rv1761c [RD152] and Rv0279c were deleted in Beijing isolates, of which 4 (Rv1572-1576c, Rv1585c-1587c, MT1798-Rv1755c and Rv1761c were variably deleted among ST42 isolates, indicating a closer relationship between the Beijing and ST42 lineages. The TbD1 region, Mb1582-Mb1583 was deleted in Beijing and ST42 isolates. One M. bovis gene of unknown function, Mb3184c was present in all isolates, except 11 of 13 ST42 isolates. The CDC1551 gene, MT1360 coding for a putative adenylate cyclase, was present in all Beijing and ST42 isolates (except 1. The pks15/1 gene, coding for a putative virulence factor, was intact in all Beijing and non-Beijing isolates, except in ST42 and ST53 isolates.This study describes previously unreported deletions/extra regions in Beijing and non-Beijing M. tuberculosis isolates. The modern and highly frequent ST42 lineage showed a closer relationship to the hypervirulent Beijing lineage than to the ancient non-Beijing lineages. The pks15/1 gene was disrupted only in modern non-Beijing isolates. This is the first report of an in-depth analysis on

Molecular characterization of multidrug-resistant Mycobacterium tuberculosis isolated in Nepal.

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Poudel, Ajay; Nakajima, Chie; Fukushima, Yukari; Suzuki, Haruka; Pandey, Basu Dev; Maharjan, Bhagwan; Suzuki, Yasuhiko

2012-06-01

Despite the fact that Nepal is one of the first countries globally to introduce multidrug-resistant tuberculosis (MDR-TB) case management, the number of MDR-TB cases is continuing to rise in Nepal. Rapid molecular tests applicable in this setting to identify resistant organisms would be an effective tool in reversing this trend. To develop such tools, information about the frequency and distribution of mutations that are associated with phenotypic drug resistance in Mycobacterium tuberculosis is required. In the present study, we investigated the prevalence of mutations in rpoB and katG genes and the inhA promoter region in 158 M. tuberculosis isolates (109 phenotypically MDR and 49 non-MDR isolates collected in Nepal) by DNA sequencing. Mutations affecting the 81-bp rifampin (RIF) resistance-determining region (RRDR) of rpoB were identified in 106 of 109 (97.3%) RIF-resistant isolates. Codons 531, 526, and 516 were the most commonly affected, at percentages of 58.7, 15.6, and 15.6%, respectively. Of 113 isoniazid (INH)-resistant isolates, 99 (87.6%) had mutations in the katG gene, with Ser315Thr being the most prevalent (81.4%) substitution. Mutations in the inhA promoter region were detected in 14 (12.4%) INH-resistant isolates. The results from this study provide an overview of the current situation of RIF and INH resistance in M. tuberculosis in Nepal and can serve as a basis for developing or improving rapid molecular tests to monitor drug-resistant strains in this country.

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 ...

Molecular Genetic Analysis of Multi-drug Resistance in Indian Isolates of Mycobacterium tuberculosis

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Noman Siddiqi

1998-09-01

Full Text Available A total of 116 isolates from patients attending the out-patient department at the All India Institute of Medical Sciences, New Delhi and the New Delhi Tuberculosis Centre, New Delhi, India were collected. They were analyzed for resistance to drugs prescribed in the treatment for tuberculosis. The drug resistance was initially determined by microbiological techniques. The Bactec 460TB system was employed to determine the type and level of resistance in each isolate. The isolates were further characterized at molecular level. The multi-drug loci corresponding to rpo b, gyr A, kat G were studied for mutation(s by the polymerase chain reaction-single strand conformational polymorphism (PCR-SSCP technique. The SSCP positive samples were sequenced to characterize the mutations in rpo b, and gyr A loci. While previously reported mutations in the gyr A and rpo b loci were found to be present, several novel mutations were also scored in the rpo b locus. Interestingly, analysis of the gyr A locus showed the presence of point mutation(s that could not be detected by PCR-SSCP. Furthermore, rifampicin resistance was found to be an important marker for checking multi-drug resistance (MDR in clinical isolates of Mycobacterium tuberculosis. This is the first report on molecular genetic analysis of MDR tuberculosis one from India, highlights the increasing incidence of MDR in the Indian isolates of M. tuberculosis.

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.

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

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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

[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.

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.

Tuberculosis relapse in Vietnam is significantly associated with Mycobacterium tuberculosis Beijing genotype infections

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Huyen, Mai N. T.; Buu, Tran N.; Tiemersma, Edine; Lan, Nguyen T. N.; Dung, Nguyen H.; Kremer, Kristin; Soolingen, Dick V.; Cobelens, Frank G. J.

2013-01-01

In Vietnam, the Mycobacterium tuberculosis Beijing genotype is associated with multi-drug resistance and is emerging. A possible explanation for this genotype's success is an increased rate of relapse. In a prospective cohort study, isolates from patients with smear-positive tuberculosis were

Polymorphisms of twenty regulatory proteins between Mycobacterium tuberculosis and Mycobacterium bovis

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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...

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

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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.

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.

Spoligotyping of Mycobacterium tuberculosis isolates from tuberculosis diagnosed patients at Dilla University Referral Hospital and other private clinics, Southern Ethiopia

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Gebremedhin Gebrezgabiher

2015-04-01

Full Text Available Objective: To assess Mycobacterium tuberculosis (M. tuberculosis strains exsisting in Gedeo zone and the surrounding areas of the Southern Ethiopia using spoligotyping. Methods: A cross sectional study was carried out from February, 2012 to June, 2013 and 97 (76 sputum and 21 fine needle aspirate samples were taken from tuberculosis diagnosed patients at Dilla University Referral Hospital and other private clinics. Culturing, region of difference (RD9 deletion typing and spoligotyping techniques were employed to isolate M. tuberculosis strains. Results: Growth of mycobacteria was observed in 35.1% (34/97. Speciation of isolates showed that 91.2% (31/34 of the isolates were M. tuberculosis. Further characterization led to the identification of 23 different spoligotype patterns of M. tuberculosis of which 61% and 39% displayed unique and cluster patterns, respectively. The most dominant shared type was spoligotype international type 53. Of the 23 strains, 12 have not been registered in the international spoligotyping database (SpolDB4. Seventy one percent of the strains belonged to the Euro-American lineage. Conclusions: This study revealed the existence of both genetically diverse and clustered M. tuberculosis strains from tuberculosis patients in the area, suggesting reactivation of infection and recent transmission, respectively. Molecular epidemiology of M. tuberculosis should be done nationwide in order to set appropriate control measures.

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

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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.

Detection of mutation in isoniazid-resistant Mycobacterium tuberculosis isolates from tuberculosis patients in Belarus

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Bostanabad S

2008-01-01

Full Text Available The aim of this study was to investigate the frequency, location and type of katG mutations in Mycobacterium tuberculosis strains isolated from patients in Belarus. Forty two isoniazid-resistant isolates were identified from sputum of 163 patients with active pulmonary tuberculosis. Drug susceptibility testing was determined by using CDC standard conventional proportional method and BACTEC system. Standard PCR method for detection of isoniazid resistance associated mutations was performed by katG gene amplification and DNA sequencing. Most mutations were found in katG gene codons 315, 316 and 309. Four types of mutations were identified in codon 315: AGC→ACC ( n = 36 85%, AGC→AGG ( n = 1 2.3%, AGC→AAC ( n = 2 4.7%, AGC→GGC ( n = 1 2.3%. One type of mutation was found in codon 316: GGC→AGC ( n = 1841.4%, four types of mutations were detected in codon 309: GGT→GGT ( n = 716.1%, GGT→GCT ( n = 49.2%, GGT→GTC ( n = 36.9%, GGT→GGG ( n = 12.7%. The highest frequency of mutations sharing between primary and secondary infections was found in codon 315.

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.

Predominance of Central Asian and European families among Mycobacterium tuberculosis isolates in Kashmir Valley, India.

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Bashir, Gulnaz; Wani, Tehmeena; Sharma, Pragya; Katoch, V M; Lone, Rubina; Shah, Azra; Katoch, Kiran; Kakru, D K; Chauhan, Devendra Singh

2017-10-01

As there are no data available regarding the strains of Mycobacterium tuberculosis circulating in Kashmir Valley, India, the current study aimed at describing the genetic diversity of M. tuberculosis strains in this region, by spoligotyping and 12-locus-based MIRU-VNTR typing (Mycobacterial Interspersed Repetitive Unit-Variable Number Tandem Repeat). Sputa from 207 smear positive cases with newly diagnosed pulmonary tuberculosis were subjected to culture for M. tuberculosis. Eighty-five isolates confirmed as M. tuberculosis were subjected to drug susceptibility testing and molecular typing by spoligotyping and MIRU-VNTRs. Drug susceptibility results of 72 isolates revealed 76.3% as fully sensitive while 5.5% as multidrug resistant (MDR). Spoligotyping of 85 isolates detected 42 spoligotypes with 50 isolates (58.8%) clustered into seven spoligotypes. SIT26/CAS1_Del was the major spoligotype (23, 27%) followed by SIT127/H4 (12, 14.1%); CAS lineage (37.6%) was predominant, followed by Haarlem (25.8%) and ill-defined T clade (23.5%). MIRU-VNTR analysis displayed 82 MIRU patterns from 85 strains, including 3 small clusters and 79 unique. MIRU 26 was found to be the most discriminatory locus. Kashmir Valley has CAS as the predominant lineage of M. tuberculosis similar to the rest of the Indian sub-continent, while it is peculiar in having Euro American lineages such as Haarlem and ill-defined T clade. Copyright © 2017 Tuberculosis Association of India. Published by Elsevier B.V. All rights reserved.

Beijing Lineage of MDR Mycobacterium tuberculosis in Bulgaria, 2007-2011

NARCIS (Netherlands)

Panaiotov, Stefan; Bachiyska, Elizabeta; Yordanova, Stanislava; Atanasova, Yuliana; Brankova, Nadia; Levterova, Viktoria; Sengstake, Sarah; Anthony, Richard; Bergval, Indra; Sola, Christophe; Kantardjiev, Todor

2014-01-01

To assess the spread of the Mycobacterium tuberculosis Beijing genotype among patients with multidrug-resistant and extensively resistant tuberculosis in Bulgaria, we genotyped 188 (72%) of 261 microbiologically confirmed resistant isolates obtained during 2007-2011. The estimated prevalence of the

Characterisation of Mycobacterium tuberculosis isolates lacking IS6110 in Viet Nam.

Science.gov (United States)

Huyen, M N T; Tiemersma, E W; Kremer, K; de Haas, P; Lan, N T N; Buu, T N; Sola, C; Cobelens, F G J; van Soolingen, D

2013-11-01

The molecular diagnosis of tuberculosis (TB) in Viet Nam is often based on the detection of insertion sequence (IS) 6110 in Mycobacterium tuberculosis. However, 8-11% of M. tuberculosis strains in South-East Asia do not contain this target and this undermines the validity of these molecular tests. We quantified the frequency of M. tuberculosis strains lacking IS6110 in rural Viet Nam and studied their epidemiological and clinical characteristics. Consecutively diagnosed adult TB patients in rural Southern Viet Nam submitted two sputum samples for culture, IS6110 restriction fragment length polymorphism (RFLP) spoligotyping and 15-loci variable number tandem repeat typing. Polymerase chain reaction (PCR) was performed to confirm the absence of IS6110 elements in strains lacking IS6110 hybridisation in RFLP. Among 2664 TB patient isolates examined, 109 (4.1%) had no IS6110 element. Compared to other strains, these no-copy strains were less often resistant to anti-tuberculosis drugs, particularly to streptomycin (adjusted OR 0.2, 95%CI 0.1-0.5), and showed significant geographic variation. No associations with TB history or demographic factors were found. Strains without the IS6110 target pose a problem in Viet Nam as regards false-negative molecular TB diagnosis in PCR. Compared to other strains circulating in Viet Nam, no-copy strains are more susceptible to anti-tuberculosis drugs.

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...

Molecular characterisation of Mycobacterium caprae strains isolated in Poland.

Science.gov (United States)

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.

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.

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

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

Science.gov (United States)

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.

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 ...

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.

Clinical data and molecular analysis of Mycobacterium tuberculosi isolates from drug-resistant tuberculosis patients in Goiás, Brazil

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Sueli Lemes de Ávila Alves

2011-09-01

Full Text Available Drug resistance is one of the major concerns regarding tuberculosis (TB infection worldwide because it hampers control of the disease. Understanding the underlying mechanisms responsible for drug resistance development is of the highest importance. To investigate clinical data from drug-resistant TB patients at the Tropical Diseases Hospital, Goiás (GO, Brazil and to evaluate the molecular basis of rifampin (R and isoniazid (H resistance in Mycobacterium tuberculosis. Drug susceptibility testing was performed on 124 isolates from 100 patients and 24 isolates displayed resistance to R and/or H. Molecular analysis of drug resistance was performed by partial sequencing of the rpoB and katGgenes and analysis of the inhA promoter region. Similarity analysis of isolates was performed by 15 loci mycobacterial interspersed repetitive unit-variable number tandem repeat (MIRU-VNTR typing. The molecular basis of drug resistance among the 24 isolates from 16 patients was confirmed in 18 isolates. Different susceptibility profiles among the isolates from the same individual were observed in five patients; using MIRU-VNTR, we have shown that those isolates were not genetically identical, with differences in one to three loci within the 15 analysed loci. Drug-resistant TB in GO is caused by M. tuberculosis strains with mutations in previously described sites of known genes and some patients harbour a mixed phenotype infection as a consequence of a single infective event; however, further and broader investigations are needed to support our findings.

rpoB gene mutations among Mycobacterium tuberculosis isolates from extrapulmonary sites.

Science.gov (United States)

Khosravi, Azar Dokht; Meghdadi, Hossein; Ghadiri, Ata A; Alami, Ameneh; Sina, Amir Hossein; Mirsaeidi, Mehdi

2018-03-01

The aim of this study was to analyze mutations occurring in the rpoB gene of Mycobacterium tuberculosis (MTB) isolates from clinical samples of extrapulmonary tuberculosis (EPTB). Seventy formalin-fixed, paraffin-embedded samples and fresh tissue samples from confirmed EPTB cases were analyzed. Nested PCR based on the rpoB gene was performed on the extracted DNAs, combined with cloning and subsequent sequencing. Sixty-seven (95.7%) samples were positive for nester PCR. Sequence analysis of the 81 bp region of the rpoB gene demonstrated mutations in 41 (61.2%) of 67 sequenced samples. Several point mutations including deletion mutations at codons 510, 512, 513 and 515, with 45% and 51% of the mutations in codons 512 and 513 respectively were seen, along with 26% replacement mutations at codons 509, 513, 514, 518, 520, 524 and 531. The most common alteration was Gln → His, at codon 513, presented in 30 (75.6%) isolates. This study demonstrated sequence alterations in codon 513 of the 81 bp region of the rpoB gene as the most common mutation occurred in 75.6% of molecularly confirmed rifampin-resistant strains. In addition, simultaneous mutation at codons 512 and 513 was demonstrated in 34.3% of the isolates. © 2018 APMIS. Published by John Wiley & Sons Ltd.

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

Whole genome sequencing reveals genomic heterogeneity and antibiotic purification in Mycobacterium tuberculosis isolates

KAUST Repository

Black, PA

2015-10-24

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 resistance. In an age where whole genome sequencing is increasingly relied upon for defining the structure of bacterial genomes, it is important to investigate the reliability of next generation sequencing to identify clonal variants present in a minor percentage of the population. This study aimed to define a reliable cut-off for identification of low frequency sequence variants and to subsequently investigate genetic heterogeneity and the evolution of drug resistance in M. tuberculosis. Methods Genomic DNA was isolated from single colonies from 14 rifampicin mono-resistant M. tuberculosis isolates, as well as the primary cultures and follow up MDR cultures from two of these patients. The whole genomes of the M. tuberculosis isolates were sequenced using either the Illumina MiSeq or Illumina HiSeq platforms. Sequences were analysed with an in-house pipeline. Results Using next-generation sequencing in combination with Sanger sequencing and statistical analysis we defined a read frequency cut-off of 30 % to identify low frequency M. tuberculosis variants with high confidence. Using this cut-off we demonstrated a high rate of genetic diversity between single colonies isolated from one population, showing that by using the current sequencing technology, single colonies are not a true reflection of the genetic diversity within a whole population and vice versa. We further showed that numerous heterogeneous variants emerge and then disappear during the evolution of isoniazid resistance within individual patients. Our findings allowed us to formulate a model for the selective bottleneck which occurs during the course of infection, acting as a genomic purification event. Conclusions Our study demonstrated true levels of genetic diversity

Use of the VNTR typing technique to determine the origin of Mycobacterium tuberculosis strains isolated from Filipino patients in Korea.

Science.gov (United States)

Lee, Jihye; Tupasi, Thelma E; Park, Young Kil

2014-05-01

With increasing international interchange of personnel, international monitoring is necessary to decrease tuberculosis incidence in the world. This study aims to develop a new tool to determine origin of Mycobacterium tuberculosis strains isolated from Filipino patients living in Korea. Thirty-two variable number tandem repeat (VNTR) loci were used for discrimination of 50 Filipino M. tuberculosis strains isolated in the Philippines, 317 Korean strains isolated in Korea, and 8 Filipino strains isolated in Korea. We found that the VNTR loci 0580, 0960, 2531, 2687, 2996, 0802, 2461, 2163a, 4052, 0424, 1955, 2074, 2347, 2401, 3171, 3690, 2372, 3232, and 4156 had different mode among copy numbers or exclusively distinct copy number in VNTR typing between Filipino and Korean M. tuberculosis strains. When these differences of the VNTR loci were applied to 8 Filipino M. tuberculosis strains isolated in Korea, 6 of them revealed Filipino type while 2 of them had Korean type. Using the differences of mode or repeated number of VNTR loci were very useful in distinguishing the Filipino strain from Korean strain.

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.

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: ...

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

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...

Phylogenetic analysis of vitamin B12-related metabolism in Mycobacterium tuberculosis

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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.

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

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.

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 ...

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

Differential Expression of Immunogenic Proteins on Virulent Mycobacterium tuberculosis Clinical Isolates

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Pablo Schierloh

2014-01-01

Full Text Available Molecular epidemiology has revealed that Mycobacterium tuberculosis (Mtb, formerly regarded as highly conserved species, displays a considerable degree of genetic variability that can influence the outcome of the disease as well as the innate and adaptive immune response. Recent studies have demonstrated that Mtb families found worldwide today differ in pathology, transmissibility, virulence, and development of immune response. By proteomic approaches seven proteins that were differentially expressed between a local clinical isolate from Latin-American-Mediterranean (LAM and from Haarlem (H lineages were identified. In order to analyze the immunogenic ability, recombinant Rv2241, Rv0009, Rv0407, and Rv2624c proteins were produced for testing specific antibody responses. We found that these proteins induced humoral immune responses in patients with drug-sensitive and drug-resistant tuberculosis with substantial cross-reactivity among the four proteins. Moreover, such reactivity was also correlated with anti-Mtb-cell surface IgM, but not with anti-ManLAM, anti-PPD, or anti-Mtb-surface IgG antibodies. Therefore, the present results describe new Mtb antigens with potential application as biomarkers of TB.

DNA restriction fragment length polymorphism analysis of Mycobacterium tuberculosis isolates from HIV-seropositive and HIV-seronegative patients in Kampala, Uganda

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Katabazi Fred A

2009-02-01

Full Text Available Abstract Background The identification and differentiation of strains of Mycobacterium tuberculosis by DNA fingerprinting has provided a better understanding of the epidemiology and tracing the transmission of tuberculosis. We set out to determine if there was a relationship between the risk of belonging to a group of tuberculosis patients with identical mycobacterial DNA fingerprint patterns and the HIV sero-status of the individuals in a high TB incidence peri-urban setting of Kampala, Uganda. Methods One hundred eighty three isolates of Mycobacterium tuberculosis from 80 HIV seropositive and 103 HIV seronegative patients were fingerprinted by standard IS6110-RFLP. Using the BioNumerics software, strains were considered to be clustered if at least one other patient had an isolate with identical RFLP pattern. Results One hundred and eighteen different fingerprint patterns were obtained from the 183 isolates. There were 34 clusters containing 54% (99/183 of the patients (average cluster size of 2.9, and a majority (96.2% of the strains possessed a high copy number (≥ 5 copies of the IS6110 element. When strains with P = 0.615, patients aged P = 0.100, and sex (aOR 1.12, 95%CI 0.60–2.06, P = 0.715. Conclusion The sample showed evidence of a high prevalence of recent transmission with a high average cluster size, but infection with an isolate with a fingerprint found to be part of a cluster was not associated with any demographic or clinical characteristics, including HIV status.

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.

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...

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

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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

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)

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.

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

Science.gov (United States)

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.

Molecular analysis of Rv0679c and Rv0180c genes of Mycobacterium tuberculosis from clinical isolates of pulmonary tuberculosis

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L Rupa

2016-01-01

Full Text Available Context: Two novel proteins/genes Rv0679c and Rv0180c of Mycobacterium tuberculosis (MTB H37Rv were classified as a hypothetical membrane and transmembrane proteins which might have a role in the invasion. Molecular analysis of these genes in human clinical isolates of pulmonary tuberculosis (PTB patients was not well characterised. Aims: To assess the molecular diversity of Rv0679c and Rv0180c genes of MTB from clinical isolates of PTB patients. Settings and Design: DNA from 97 clinical isolates was extracted and subjected to amplification using selective primers by polymerase chain reaction (PCR. The PCR product obtained was sequenced commercially. Patients and Methods: Clinical isolates obtained from tuberculosis patients were investigated for polymorphisms in the Rv0679c and Rv0180c genes by PCR and DNA sequencing. Genomic DNA isolated by cetyltrimethylammonium bromide method was used for amplification of genes. Results: Rv0679c gene was highly conserved in 61 out of 65 clinical isolates assessed for sequence homology with wild-type H37Rv gene and was identical using ClustalW. Fifty-five out of 78 (70.5% clinical isolates assessed for Rv0180c were positive for single nucleotide polymorphism (SNP at 258th position where the nucleotide G was replaced with T (G to T. In clinical isolates of untreated cases, the frequency was 54.5% for SNP at 258th position which is low compared to cases undergoing treatment where the frequency was 73.1%. Conclusions: Molecular analysis of Rv0180c in clinical isolates of PTB assessed in this study was the first report, where an SNP at 258th position G to T was identified within the gene. Rv0679c gene was highly conserved (94%, within Indian clinical isolates as compared to reports from other nations.

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

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

RAPD Analysis and Antibiotic Susceptibility for Mycobacterium tuberculosis Strains Isolated from Different Locations in Egypt

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Ali, A. M.

2011-01-01

Full Text Available The routine identification of mycobacterial strains isolated from patients in different locations in Egypt was confirmed by specific DNA fragment amplification. The susceptibilities of 72 Mycobacterium tuberculosis strains against the four antibiotics used in tuberculosis treatment (Isoniazid, INH; Rifampicin, Rif; Streptomycin, St and Ethambutol, E were examined. Our results indicated that, multi drug resistant tuberculosis (MDR-TB represents about 19.5% of the tested strains, whereas sensitive strains represented 26.4%. The genetic polymorphism of the tested strains was examined using RAPD analysis. Six selected strains represent the different antibiotic susceptibility groups were examined using RAPD fingerprinting. No difference between the strains was recorded using the RFLP analysis of amplified specific fragment. The discrimination power of RAPD analysis was inadequate to clarify the genetic correlation between the tested strains. MDR-TB was approximately double time in 2008 compared with the value in 2007. Most of the new MDRTB was correlated with resident dense population regions.

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

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)

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

Dehalogenation of Haloalkanes by Mycobacterium tuberculosis H37Rv and Other Mycobacteria

Science.gov (United States)

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

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.

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

Science.gov (United States)

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.

Determination of in vitro synergy between linezolid and other antimicrobial agents against Mycobacterium tuberculosis isolates.

Science.gov (United States)

Zou, Lin; Liu, Min; Wang, Yufeng; Lu, Jie; Pang, Yu

2015-12-01

In this study, our objective was to explore the potential in vitro synergy between linezolid (LZD) and six other anti-TB drugs in Mycobacterium tuberculosis strains, especially multidrug-resistant tuberculosis (MDR-TB) strains. Among the different combinations, the LZD-clarithromycin (CLA) combination showed the best synergism, which was observed in 85% (34/40) of 40 isolates. In addition, one (2.5%) and twenty-one (52.5%) of 40 isolates showed synergism for the LZD-levofloxcin (LEV) and LZD-moxifloxacin (MOX) combinations, respectively, and the difference in the proportion of synergy between these two combinations was significantly different (P synergy against non-MDR group seemed higher than that against MDR group in each combination, while the significant difference was only observed in the LZD-EMB combination (P = 0.046). In conclusion, our findings demonstrate that LZD shows the synergistic activity against both non-MDR and MDR M. tuberculosis strains when in combination with CLA, EMB, MOX, amikacin and clofazimine, indicating that LZD may be considered as a promising component involving the regimen for the treatment of MDR-TB. Copyright © 2015 Elsevier Ltd. All rights reserved.

Comparative study of IS6110 restriction fragment length polymorphism and variable-number tandem-repeat typing of Mycobacterium tuberculosis isolates in the Netherlands, based on a 5-year nationwide survey

NARCIS (Netherlands)

de Beer, Jessica L.; van Ingen, Jakko; de Vries, Gerard; Erkens, Connie; Sebek, Maruschka; Mulder, Arnout; Sloot, Rosa; van den Brandt, Anne-Marie; Enaimi, Mimount; Kremer, Kristin; Supply, Philip; van Soolingen, Dick

2013-01-01

In order to switch from IS6110 and polymorphic GC-rich repetitive sequence (PGRS) restriction fragment length polymorphism (RFLP) to 24-locus variable-number tandem-repeat (VNTR) typing of Mycobacterium tuberculosis complex isolates in the national tuberculosis control program in The Netherlands, a

Comparative Study of IS6110 Restriction Fragment Length Polymorphism and Variable-Number Tandem-Repeat Typing of Mycobacterium tuberculosis Isolates in the Netherlands, Based on a 5-Year Nationwide Survey

NARCIS (Netherlands)

Beer, J.L. de; Ingen, J. van; Vries, G. de; Erkens, C.; Sebek, M.; Mulder, A.; Sloot, R.; Brandt, A.M. van den; Enaimi, M.; Kremer, K.; Supply, P.; Soolingen, D. van

2013-01-01

In order to switch from IS6110 and polymorphic GC-rich repetitive sequence (PGRS) restriction fragment length polymorphism (RFLP) to 24-locus variable-number tandem-repeat (VNTR) typing of Mycobacterium tuberculosis complex isolates in the national tuberculosis control program in The Netherlands, a

Trends of Mycobacterium bovis Isolation and First-Line Anti-tuberculosis Drug Susceptibility Profile: A Fifteen-Year Laboratory-Based Surveillance.

Directory of Open Access Journals (Sweden)

Miriam Bobadilla-del Valle

2015-09-01

Full Text Available Mycobacterium tuberculosis causes the majority of tuberculosis (TB cases in humans; however, in developing countries, human TB caused by M. bovis may be frequent but undetected. Human TB caused by M. bovis is considered a zoonosis; transmission is mainly through consumption of unpasteurized dairy products, and it is less frequently attributed to animal-to-human or human-to-human contact. We describe the trends of M. bovis isolation from human samples and first-line drug susceptibility during a 15-year period in a referral laboratory located in a tertiary care hospital in Mexico City.Data on mycobacterial isolates from human clinical samples were retrieved from the laboratory's database for the 2000-2014 period. Susceptibility to first-line drugs: rifampin, isoniazid, streptomycin (STR and ethambutol was determined. We identified 1,165 isolates, 73.7% were M. tuberculosis and 26.2%, M. bovis. Among pulmonary samples, 16.6% were M. bovis. The proportion of M. bovis isolates significantly increased from 7.8% in 2000 to 28.4% in 2014 (X(2trend, p<0.001. Primary STR resistance was higher among M. bovis compared with M. tuberculosis isolates (10.9% vs.3.4%, p<0.001. Secondary multidrug resistance (MDR rates were 38.5% and 34.4% for M. bovis and M. tuberculosis, respectively (p = 0.637. A rising trend of primary STR monoresistance was observed for both species (3.4% in 2000-2004 vs. 7.6% in 2010-2014; p = 0.02.There is a high prevalence and a rising trend of M. bovis isolates in our region. The proportion of pulmonary M. bovis isolates is higher than in previous reports. Additionally, we report high rates of primary anti-tuberculosis resistance and secondary MDR in both M. tuberculosis and M. bovis. This is one of the largest reports on drug susceptibility of M. bovis from human samples and shows a significant proportion of first-line anti-tuberculosis drug resistance.

Trends of Mycobacterium bovis Isolation and First-Line Anti-tuberculosis Drug Susceptibility Profile: A Fifteen-Year Laboratory-Based Surveillance.

Science.gov (United States)

Bobadilla-del Valle, Miriam; Torres-González, Pedro; Cervera-Hernández, Miguel Enrique; Martínez-Gamboa, Areli; Crabtree-Ramirez, Brenda; Chávez-Mazari, Bárbara; Ortiz-Conchi, Narciso; Rodríguez-Cruz, Luis; Cervantes-Sánchez, Axel; Gudiño-Enríquez, Tomasa; Cinta-Severo, Carmen; Sifuentes-Osornio, José; Ponce de León, Alfredo

2015-09-01

Mycobacterium tuberculosis causes the majority of tuberculosis (TB) cases in humans; however, in developing countries, human TB caused by M. bovis may be frequent but undetected. Human TB caused by M. bovis is considered a zoonosis; transmission is mainly through consumption of unpasteurized dairy products, and it is less frequently attributed to animal-to-human or human-to-human contact. We describe the trends of M. bovis isolation from human samples and first-line drug susceptibility during a 15-year period in a referral laboratory located in a tertiary care hospital in Mexico City. Data on mycobacterial isolates from human clinical samples were retrieved from the laboratory's database for the 2000-2014 period. Susceptibility to first-line drugs: rifampin, isoniazid, streptomycin (STR) and ethambutol was determined. We identified 1,165 isolates, 73.7% were M. tuberculosis and 26.2%, M. bovis. Among pulmonary samples, 16.6% were M. bovis. The proportion of M. bovis isolates significantly increased from 7.8% in 2000 to 28.4% in 2014 (X(2)trend, ptuberculosis isolates (10.9% vs.3.4%, ptuberculosis, respectively (p = 0.637). A rising trend of primary STR monoresistance was observed for both species (3.4% in 2000-2004 vs. 7.6% in 2010-2014; p = 0.02). There is a high prevalence and a rising trend of M. bovis isolates in our region. The proportion of pulmonary M. bovis isolates is higher than in previous reports. Additionally, we report high rates of primary anti-tuberculosis resistance and secondary MDR in both M. tuberculosis and M. bovis. This is one of the largest reports on drug susceptibility of M. bovis from human samples and shows a significant proportion of first-line anti-tuberculosis drug resistance.

Trends of Mycobacterium bovis Isolation and First-Line Anti-tuberculosis Drug Susceptibility Profile: A Fifteen-Year Laboratory-Based Surveillance

Science.gov (United States)

Bobadilla-del Valle, Miriam; Torres-González, Pedro; Cervera-Hernández, Miguel Enrique; Martínez-Gamboa, Areli; Crabtree-Ramirez, Brenda; Chávez-Mazari, Bárbara; Ortiz-Conchi, Narciso; Rodríguez-Cruz, Luis; Cervantes-Sánchez, Axel; Gudiño-Enríquez, Tomasa; Cinta-Severo, Carmen; Sifuentes-Osornio, José; Ponce de León, Alfredo

2015-01-01

Background Mycobacterium tuberculosis causes the majority of tuberculosis (TB) cases in humans; however, in developing countries, human TB caused by M. bovis may be frequent but undetected. Human TB caused by M. bovis is considered a zoonosis; transmission is mainly through consumption of unpasteurized dairy products, and it is less frequently attributed to animal-to-human or human-to-human contact. We describe the trends of M. bovis isolation from human samples and first-line drug susceptibility during a 15-year period in a referral laboratory located in a tertiary care hospital in Mexico City. Methodology/Principal Findings Data on mycobacterial isolates from human clinical samples were retrieved from the laboratory’s database for the 2000–2014 period. Susceptibility to first-line drugs: rifampin, isoniazid, streptomycin (STR) and ethambutol was determined. We identified 1,165 isolates, 73.7% were M. tuberculosis and 26.2%, M. bovis. Among pulmonary samples, 16.6% were M. bovis. The proportion of M. bovis isolates significantly increased from 7.8% in 2000 to 28.4% in 2014 (X 2 trend, ptuberculosis isolates (10.9% vs.3.4%, ptuberculosis, respectively (p = 0.637). A rising trend of primary STR monoresistance was observed for both species (3.4% in 2000–2004 vs. 7.6% in 2010–2014; p = 0.02). Conclusions/Significance There is a high prevalence and a rising trend of M. bovis isolates in our region. The proportion of pulmonary M. bovis isolates is higher than in previous reports. Additionally, we report high rates of primary anti-tuberculosis resistance and secondary MDR in both M. tuberculosis and M. bovis. This is one of the largest reports on drug susceptibility of M. bovis from human samples and shows a significant proportion of first-line anti-tuberculosis drug resistance. PMID:26421930

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.

Supplementary Material for: Whole genome sequencing reveals genomic heterogeneity and antibiotic purification in Mycobacterium tuberculosis isolates

KAUST Repository

Black, PA

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 resistance. In an age where whole genome sequencing is increasingly relied upon for defining the structure of bacterial genomes, it is important to investigate the reliability of next generation sequencing to identify clonal variants present in a minor percentage of the population. This study aimed to define a reliable cut-off for identification of low frequency sequence variants and to subsequently investigate genetic heterogeneity and the evolution of drug resistance in M. tuberculosis. Methods Genomic DNA was isolated from single colonies from 14 rifampicin mono-resistant M. tuberculosis isolates, as well as the primary cultures and follow up MDR cultures from two of these patients. The whole genomes of the M. tuberculosis isolates were sequenced using either the Illumina MiSeq or Illumina HiSeq platforms. Sequences were analysed with an in-house pipeline. Results Using next-generation sequencing in combination with Sanger sequencing and statistical analysis we defined a read frequency cut-off of 30 % to identify low frequency M. tuberculosis variants with high confidence. Using this cut-off we demonstrated a high rate of genetic diversity between single colonies isolated from one population, showing that by using the current sequencing technology, single colonies are not a true reflection of the genetic diversity within a whole population and vice versa. We further showed that numerous heterogeneous variants emerge and then disappear during the evolution of isoniazid resistance within individual patients. Our findings allowed us to formulate a model for the selective bottleneck which occurs during the course of infection, acting as a genomic purification event. Conclusions Our study demonstrated true levels of genetic

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...

[Identification and drug susceptibility testing of Mycobacterium thermoresistibile and Mycobacterium elephantis isolated from a cow with mastitis].

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Li, W B; Ji, L Y; Xu, D L; Liu, H C; Zhao, X Q; Wu, Y M; Wan, K L

2018-05-10

Objective: To understand the etiological characteristics and drug susceptibility of Mycobacterium thermoresistibile and Mycobacterium elephantis isolated from a cow with mastitis and provide evidence for the prevention and control of infectious mastitis in cows. Methods: The milk sample was collected from a cow with mastitis, which was pretreated with 4 % NaOH and inoculated with L-J medium for Mycobacterium isolation. The positive cultures were initially identified by acid-fast staining and multi-loci PCR, then Mycobacterium species was identified by the multiple loci sequence analysis (MLSA) with 16S rRNA , hsp65 , ITS and SodA genes. The drug sensitivity of the isolates to 27 antibiotics was tested by alamar blue assay. Results: Two anti-acid stain positive strains were isolated from the milk of a cow with mastitis, which were identified as non- tuberculosis mycobacterium by multi-loci PCR, and multi-loci nucleic acid sequence analysis indicated that one strain was Mycobacterium thermoresistibile and another one was Mycobacterium elephantis . The results of the drug susceptibility test showed that the two strains were resistant to most antibiotics, including rifampicin and isoniazid, but they were sensitive to amikacin, moxifloxacin, levofloxacin, ethambutol, streptomycin, tobramycin, ciprofloxacin and linezolid. Conclusions: Mycobacterium thermoresistibile and Mycobacterium elephantis were isolated in a cow with mastitis and the drug susceptibility spectrum of the pathogens were unique. The results of the study can be used as reference for the prevention and control the infection in cows.

Overview of errors in the reference sequence and annotation of Mycobacterium tuberculosis H37Rv, and variation amongst its isolates

KAUST Repository

Köser, Claudio U.

2012-06-01

Since its publication in 1998, the genome sequence of the Mycobacterium tuberculosis H37Rv laboratory strain has acted as the cornerstone for the study of tuberculosis. In this review we address some of the practical aspects that have come to light relating to the use of H37Rv throughout the past decade which are of relevance for the ongoing genomic and laboratory studies of this pathogen. These include errors in the genome reference sequence and its annotation, as well as the recently detected variation amongst isolates of H37Rv from different laboratories. © 2011 Elsevier B.V..

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 ...

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.

Population Structure among Mycobacterium tuberculosis Isolates from Pulmonary Tuberculosis Patients in Colombia

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Realpe, Teresa; Correa, Nidia; Rozo, Juan Carlos; Ferro, Beatriz Elena; Gomez, Verónica; Zapata, Elsa; Ribon, Wellman; Puerto, Gloria; Castro, Claudia; Nieto, Luisa María; Diaz, Maria Lilia; Rivera, Oriana; Couvin, David; Rastogi, Nalin; Arbelaez, Maria Patricia; Robledo, Jaime

2014-01-01

Background Phylogeographic composition of M. tuberculosis populations reveals associations between lineages and human populations that might have implications for the development of strategies to control the disease. In Latin America, lineage 4 or the Euro-American, is predominant with considerable variations among and within countries. In Colombia, although few studies from specific localities have revealed differences in M. tuberculosis populations, there are still areas of the country where this information is lacking, as is a comparison of Colombian isolates with those from the rest of the world. Principal Findings A total of 414 M. tuberculosis isolates from adult pulmonary tuberculosis cases from three Colombian states were studied. Isolates were genotyped using IS6110-restriction fragment length polymorphism (RFLP), spoligotyping, and 24-locus Mycobacterial interspersed repetitive units variable number tandem repeats (MIRU-VNTRs). SIT42 (LAM9) and SIT62 (H1) represented 53.3% of isolates, followed by 8.21% SIT50 (H3), 5.07% SIT53 (T1), and 3.14% SIT727 (H1). Composite spoligotyping and 24-locus MIRU- VNTR minimum spanning tree analysis suggest a recent expansion of SIT42 and SIT62 evolved originally from SIT53 (T1). The proportion of Haarlem sublineage (44.3%) was significantly higher than that in neighboring countries. Associations were found between M. tuberculosis MDR and SIT45 (H1), as well as HIV-positive serology with SIT727 (H1) and SIT53 (T1). Conclusions This study showed the population structure of M. tuberculosis in several regions from Colombia with a dominance of the LAM and Haarlem sublineages, particularly in two major urban settings (Medellín and Cali). Dominant spoligotypes were LAM9 (SIT 42) and Haarlem (SIT62). The proportion of the Haarlem sublineage was higher in Colombia compared to that in neighboring countries, suggesting particular conditions of co-evolution with the corresponding human population that favor the success of this

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

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

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

2003-11-01

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

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...

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

Science.gov (United States)

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.

Bovine tuberculosis

Science.gov (United States)

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...

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.

Mycobacterium tuberculosis Metabolism

Science.gov (United States)

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

Microbial sensor for drug susceptibility testing of Mycobacterium tuberculosis.

Science.gov (United States)

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.

Molecular characterization of Mycobacterium tuberculosis isolates from Tehran, Iran by restriction fragment length polymorphism analysis and spoligotyping

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Seifu Gizaw Feyisa

2016-04-01

Full Text Available Abstract: INTRODUCTION Characterization of Mycobacterium tuberculosis (MTB isolates by DNA fingerprinting has contributed to tuberculosis (TB control. The aim of this study was to determine the genetic diversity of MTB isolates from Tehran province in Iran. METHODS MTB isolates from 60 Iranian and 10 Afghan TB patients were fingerprinted by standard IS6110-restriction fragment length polymorphism (RFLP analysis and spoligotyping. RESULTS The copy number of IS6110 ranged from 10-24 per isolate. The isolates were classified into 22 clusters showing ≥ 80% similarity by RFLP analysis. Fourteen multidrug-resistant (MDR isolates were grouped into 4 IS6110-RFLP clusters, with 10 isolates [71% (95% CI: 45-89%] in 1 cluster, suggesting a possible epidemiological linkage. Eighteen Iranian isolates showed ≥ 80% similarity with Afghan isolates. There were no strains with identical fingerprints. Spoligotyping of 70 isolates produced 23 distinct patterns. Sixty (85.7% isolates were grouped into 13 clusters, while the remaining 10 isolates (14.2% were not clustered. Ural (formerly Haarlem4 (n = 22, 31.4% was the most common family followed by Central Asian strain (CAS (n = 18, 25.7% and T (n = 9, 12.8% families. Only 1strain was characterized as having the Beijing genotype. Among 60 Iranian and 10 Afghan MTB isolates, 25% (95% CI: 16-37 and 70% (95% CI: 39-89 were categorized as Ural lineage, respectively. CONCLUSIONS A higher prevalence of Ural family MTB isolates among Afghan patients than among Iranian patients suggests the possible transmission of this lineage following the immigration of Afghans to Iran.

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

Directory of Open Access Journals (Sweden)

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.

Limited variation of DNA fingerprints (IS6110 and IS1081) in Korean strains of Mycobacterium tuberculosis.

Science.gov (United States)

Huh, Y J; Ahn, D I; Kim, S J

1995-08-01

To establish the usefulness of DNA fingerprinting for the epidemiology of Mycobacterium tuberculosis isolated from Korean tuberculosis patients. Comparison of restriction fragment length polymorphism (RFLP) patterns produced by southern hybridization of PvuII-digested chromosomal DNA. IS6110-associated banding patterns of 41 isolates varied considerably, containing 1-13 copies. The RFLP pattern of the epidemiologically related M. tuberculosis isolates was identical in 8 of 10 groups of close contact patients. No noticeable differences in RFLP were observed between drug-sensitive and drug-resistant isolates. IS1081-containing restriction fragment analysis of 52 isolates showed 6 different banding patterns, and the C type was found dominant in Korea. Identification of G type M. tuberculosis, which has a 8.0 kb IS1081-containing PvuII fragment, is unusual because it has been observed only in M. bovis BCG so far. IS6110 was a very useful tool for tracing the transmission route of tuberculosis; IS1081 was also useful for subdividing M. tuberculosis into several groups.

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.

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.

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.

Genome sequence of Mycobacterium yongonense RT 955-2015 isolate from a patient misdiagnosed with multi-drug resistant tuberculosis: first clinical isolate in Tanzania.

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Mnyambwa, Nicholaus Peter; Kim, Dong-Jin; Ngadaya, Esther; Chun, Jongsik; Ha, Sung-Min; Petrucka, Pammla; Addo, Kennedy Kwasi; Kazwala, Rudovick R; Mfinanga, Sayoki G

2018-04-24

Mycobacterium yongonense is a recently described novel species belonging to Mycobacterium avium complex which is the most prevalent etiology of non-tuberculous mycobacteria associated with pulmonary infections, and posing tuberculosis diagnostic challenges in high-burden, resource-constrained settings. We used whole genome shotgun sequencing and comparative microbial genomic analyses to characterize the isolate from a patient diagnosed with multi-drug resistant tuberculosis (MDR-TB) after relapse. We present a genome sequence of the first case of M. yongonense (M. yongonense RT 955-2015) in Tanzania. Sequence analysis revealed that the RT 955-2015 strain had a high similarity to M. yongonense 05-1390(T) (98.74%) and M. chimaera DSM 44623(T) (98%). Its 16S rRNA showed similarity to M. paraintracellulare KCTC 290849(T) (100%); M. intracellulare ATCC 13950(T) (100%); M. chimaera DSM 44623(T) (99.9%); and M. yongonense 05-1390(T) (98%). The strain had a substantially different rpoB sequence from that of M. yongonense 05-1390 (95.16%) but exhibited a sequence closely related to M. chimaera DSM 44623(T) (99.86%), M. intracellulare ATCC 13950(T) (99.53%), and M. paraintracellulare KCTC 290849(T) (99.53%). In light of the OrthoANI algorithm, and phylogenetic analysis, we conclude that the isolate was M. yongonense Type II genotype, which is an indication that the patient was misdiagnosed with TB/MDR-TB and received inappropriate treatment. Copyright © 2018. Published by Elsevier Ltd.

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.

Molecular typing of mycobacterium tuberculosis isolates circulating in Jiangsu Province, China

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Dong Haiyan

2011-10-01

Full Text Available Abstract Background Globally, China is the second place with high burden of tuberculosis (TB. To explore the characteristics of the pathogens of Mycobacterium tuberculosis (MTB circulating in this area is helpful for understanding and controlling the spread of the strains. Recent developments in molecular biology have allowed prompt identification and tracking specific strains of MTB spreading through the population. Methods Spacer-oligonucleotide typing (spoligotyping and mycobacterial interspersed repetitive units variable number tandem repeat (MIRU-VNTR were performed in combination to yield specific genetic profiles of 260 MTB strains isolated from 30 counties of Jiangsu province in China between June and July 2010. The spoligotyping results were in comparison to the world Spoligotyping Database of Institute Pasteur de Guadeloupe (SpolDB4. Drug susceptibility test (DST was performed on all strains by proportion method on Lowenstein-Jensen (LJ culture media. Results Based on the spoligotyping method, 246 strains displayed known patterns and 14 were absent in the database. Predominant spoligotypes belonged to the Beijing family (80.4%. By using the 24-loci VNTR typing scheme, 224 different patterns were identified, including 20 clusters and 204 unique patterns. The largest clade comprised 195 strains belonging to the Beijing family. The combination of spoligotyping and 24-loci MIRU-VNTR demonstrated maximal discriminatory power. Furthermore, we observed a significant association between Beijing family strains and drug-resistant phenotypes. The Beijing family strains presented increased risks for developing multi-drug resistant TB, with the OR (95% CI of 11.07(1.45-84.50. Conclusions The present study demonstrated that Beijing family isolates were the most prevalent strains circulating in Jiangsu province of China. The utility of spoligotyping in combination with 24-loci MIRU-VNTR might be a useful tool for epidemiological analysis of MTB

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.

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.

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

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

Assessment of trends of ofloxacin resistance in Mycobacterium tuberculosis

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J S Verma

2011-01-01

Full Text Available Purpose: Ofloxacin (OFX is one of the potent fluoroquinolone (FQ recommended to treat MDR-TB. Over a decade, the preexposure of this drug for the treatment of other bacterial infections has resulted in acquisition of FQ resistance among Mycobacterium tuberculosis strains. Considering this possibility, a study was undertaken in a tertiary care center in the capital city (India to assess the drug resistance trends of OFX among susceptible and multidrug resistant (MDR strains of M. tuberculosis. Materials and Methods: A total of 102 M. tuberculosis isolates (47 susceptible to first-line drugs and 55 MDR isolates were screened for susceptibility testing of OFX with a critical concentration of 2 μg/ml by Lowenstein Jensen (LJ proportion method. Results: The results showed 40 (85.1% isolates among 47 susceptible isolates and 34 (61.8% isolates among 55 MDR isolates, were found to be susceptible to OFX. Fisher′s exact test showed significant P-value (0.0136 demonstrating 1.377 fold (95% confidence interval increased risk to become resistant to OFX than susceptible isolates. These finding shows decreased OFX susceptibility is not only limited to MDR isolates but also increasingly seen in susceptible strains as a result of drug abuse. Conclusions: Our finding were not alarming, but highlights the general risk of acquiring resistance to OFX, jeopardizing the potential for these drugs to be used as second-line anti-TB agents in the management of drug-resistant TB and creating incurable TB strains .

Rapid screening of rpoB and katG mutations in Mycobacterium tuberculosis isolates by high-resolution melting curve analysis

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M Haeili

2014-01-01

Full Text Available Background: Early detection of multidrug-resistant tuberculosis (MDR-TB is essential to prevent its transmission in the community and initiate effective anti-TB treatment regimen. Materials and Methods: High-resolution melting curve (HRM analysis was evaluated for rapid detection of resistance conferring mutations in rpoB and katG genes. We screened 95 Mycobacterium tuberculosis clinical isolates including 20 rifampin resistant (RIF-R, 21 isoniazid resistant (INH-R and 54 fully susceptible (S isolates determined by proportion method of drug susceptibility testing. Nineteen M. tuberculosis isolates with known drug susceptibility genotypes were used as references for the assay validation. The nucleotide sequences of the target regions rpoB and katG genes were determined to investigate the frequency and type of mutations and to confirm HRM results. Results: HRM analysis of a 129-bp fragment of rpoB allowed correct identification of 19 of the 20 phenotypically RIF-R and all RIF-S isolates. All INH-S isolates generated wild-type HRM curves and 18 out of 21 INH-R isolates harboured any mutation in 109-bp fragment of katG exhibited mutant type HRM curves. However, 1 RIF-R and 3 INH-R isolates were falsely identified as susceptible which were confirmed for having no mutation in their target regions by sequencing. The main mutations involved in RIF and INH resistance were found at codons rpoB531 (60% of RIF-R isolates and katG315 (85.7% of INH-R isolates, respectively. Conclusion: HRM was found to be a reliable, rapid and low cost method to characterise drug susceptibility of clinical TB isolates in resource-limited settings.

Detection of 123 bp fragment of insertion element IS6110 Mycobacterium tuberculosis for diagnosis of extrapulmonary tuberculosis

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A K Maurya

2012-01-01

Full Text Available Purpose: Extrapulmonary tuberculosis (EPTB is emerging problem in developing and developed countries. The diagnosis of EPTB in its different clinical presentations remains a true challenge. IS6110-based polymerase chain reaction (PCR is used for rapid identification and positivity rate of the Mycobacterium tuberculosis complex in clinical isolates of different sites of EPTB. The present study was carried out to study the prevalence of M. tuberculosis complex in clinical isolates of EPTB at tertiary care centres in Lucknow. Materials and Methods: Seven hundred fifty-six specimens were collected from the suspected cases of EPTB which were processed for Mycobacteria by Ziehl Neelson (ZN staining and BACTEC culture. All the specimens were also processed for IS6110-based PCR amplification with primers targeting 123 bp fragment of insertion element IS6110 of the M. tuberculosis complex. Results: Of these 756 specimens, 71(9.3% were positive for acid fast bacilli (AFB by ZN staining, 227(30.1% were positive for mycobacteria by BACTEC culture and IS6110 PCR were positive for M. tuberculosis complex in 165 (20.7% isolates. We found a significant difference in sensitivities of different tests (P<0.05. Conclusions: This study reveals the positivity of M. tuberculosis complex in clinical isolates of EPTB case in tertiary care hospitals in Northern India. 72.7% of M. tuberculosis complex was confirmed by IS6110-PCR in culture isolates from different sites of EPTB. The high prevalence of the M. tuberculosis complex was seen in lymph node aspirate and synovial fluid. However, utility of PCR may play a potentially significant role in strengthening the diagnosis of EPTB especially targeting IS6110.

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.

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)

Evaluation of the Sensititre MycoTB plate for susceptibility testing of the Mycobacterium tuberculosis complex against first- and second-line agents.

Science.gov (United States)

Hall, Leslie; Jude, Kurt P; Clark, Shirley L; Dionne, Kim; Merson, Ryan; Boyer, Ana; Parrish, Nicole M; Wengenack, Nancy L

2012-11-01

The Sensititre MycoTB plate (TREK Diagnostic Systems, Cleveland, OH) uses a microtiter plate MIC format for susceptibility testing of Mycobacterium tuberculosis complex isolates against first- and second-line antituberculosis agents. Categorical agreement versus the agar proportion method for 122 M. tuberculosis complex isolates was 94% to 100%.

Extended spectrum of antibiotic susceptibility for tuberculosis, Djibouti.

Science.gov (United States)

Bouzid, Fériel; Astier, Hélène; Osman, Djaltou Aboubaker; Javelle, Emilie; Hassan, Mohamed Osman; Simon, Fabrice; Garnotel, Eric; Drancourt, Michel

2018-02-01

In the Horn of Africa, there is a high prevalence of tuberculosis that is reported to be partly driven by multidrug-resistant (MDR) Mycobacterium tuberculosis strictu sensu strains. We conducted a prospective study to investigate M. tuberculosis complex species causing tuberculosis in Djibouti, and their in vitro susceptibility to standard anti-tuberculous antibiotics in addition to clofazimine, minocycline, chloramphenicol and sulfadiazine. Among the 118 mycobacteria isolates from 118 successive patients with suspected pulmonary tuberculosis, 111 strains of M. tuberculosis, five Mycobacterium canettii, one 'Mycobacterium simulans' and one Mycobacterium kansasii were identified. Drug-susceptibility tests performed on the first 78 isolates yielded nine MDR M. tuberculosis isolates. All isolates were fully susceptible to clofazimine, minocycline and chloramphenicol, and 75 of 78 isolates were susceptible to sulfadiazine. In the Horn of Africa, patients with confirmed pulmonary tuberculosis caused by an in vitro susceptible strain may benefit from anti-leprosy drugs, sulfamides and phenicol antibiotics. Copyright © 2017 Elsevier B.V. and International Society of Chemotherapy. All rights reserved.

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.

Tuberculosis Caused by Mycobacterium orygis in Dairy Cattle and Captured Monkeys in Bangladesh: a New Scenario of Tuberculosis in South Asia.

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Rahim, Z; Thapa, J; Fukushima, Y; van der Zanden, A G M; Gordon, S V; Suzuki, Y; Nakajima, C

2017-12-01

Mycobacterium orygis, commonly known as the oryx bacillus and a newly proposed Mycobacterium tuberculosis complex subspecies, was isolated from 18 cattle in a dairy farm and two captured rhesus monkeys in a zoo in Bangladesh. All the infected animals had tuberculosis lesions in their lungs, suggesting transmission and infection with M. orygis by an airborne route. The 20 isolates were analysed using a range of conventional and molecular typing methods, and RD-deletion typing and sequencing of selected genes confirmed the isolates as M. orygis. Multiple-locus variable-number tandem repeat analysis (MLVA) allowed the isolates to be divided into three clusters based on the relatedness of their MLVA profiles. The two monkey isolates shared the same MLVA pattern with 15 of the cattle isolates, whereas the remaining three cattle isolates had different patterns, even though the latter animals had been kept in the same dairy farm. The diversity observed among isolates may suggest the bacteria have been established in this area for a long period. This study along with other recent findings that report the detection of M. orygis from animals as well as humans originating from South Asia potentially indicate endemic distribution of M. orygis in South Asia. © 2016 Blackwell Verlag GmbH.

Risk factors for Mycobacterium tuberculosis infection among children in Greenland

DEFF Research Database (Denmark)

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....

Investigation of Susceptibility of Mycobacterium tuberculosis Complex Strains Isolated from Clinical Samples Against the First and Second-Line Anti-tuberculosis Drugs by the Sensititre MycoTB Plate Method

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Figen KAYSERİLİ ORHAN

2018-03-01

Full Text Available Introduction: Phenotypic methods for drug susceptibility testing of Mycobacterium tuberculosis complex (MTC to second-line drugs are not yet standardized. The Sensititre MycoTB Plate is a microtiter plate containing lyophilized antibiotics and configured for determination of MIC to first and second-line anti-tuberculosis drugs. The purpose of this study is to detect the susceptibility rates of MTC strains isolated from patients’ specimens for first and second-line anti-tuberculosis drugs. Materials and Methods: This study included 50 MTC strains isolated from various clinical specimens. Out of the 50 strains, 38 were isolated from sputum, three from cerebrospinal fluid, three from bronchoalveolar lavage, and six from other samples in this study. The susceptibility of strains to anti-tuberculosis drugs were determined by the Sensititre MycoTB Plate Method. Thawed isolates were subcultured, and dilutions were inoculated into MycoTB wells. The results were read at days 7, 14 and 21. Results: At the end of study, out of 50 MTC isolates, 7 (14% showed resistance to Isoniazid (INH, 5 (10% to streptomycin (SM, 4 (8% to ethambutol (EMB, 4 (8% to ethionamide (ETH, 3 (6% to rifampicin (RIF, 3 (6% to rifabutin (RFB, 2 (4% to kanamycin (KAN, 2 (4% to ofloxacin (OFL, 2 (4% to P-aminosalicyclic acid (PAS, 1 (2% to moxiflocacin (MOX, and 1 (2% to cycloserine (CYC. All strains were found sensitive to amikacin while 2 strains (4% were identified as multidrug-resistant tuberculosis (MDR-TB. Thirty-five strains (70% were sensitive to all drugs. Extensively drug resistant tuberculosis (XDR-TB was not determined in this study. Conclusion: This is the first study that tests second line anti-tuberculosis drugs in our location and provides us valuable data regarding MDR-TB and XDR-TB rates. The Sensititre MycoTB Plate Method is a fast, reliable and practical method and can be used to determine the susceptibility of first and second-line anti-tuberculosis drugs.

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.

Analysis of expression profile of mce operon genes (mce1, mce2, mce3 operon) in different Mycobacterium tuberculosis isolates at different growth phases.

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Singh, Pratibha; Katoch, V M; Mohanty, K K; Chauhan, Devendra Singh

2016-04-01

Mycobacterium tuberculosis (M. tuberculosis) has four homologous mammalian cell entry (mce) operons (mce1-4) that encode exported proteins and have a possible role in the virulence mechanism of this pathogen. The expression of mce operon is considered to be complex and not completely understood. Although expression of mce operon at different in vitro growth phases has been studied earlier, its expression in different M. tuberculosis isolates under different growth phases is not yet studied. The present preliminary study was conducted on a limited number of isolates to know the trend of expression pattern of mce operon genes in different M. tuberculosis isolates under different growth stages. In this study, we monitored the transcriptional profile of selected mce operon genes (mce1A, mce1D, mce2A, mce2D, mce3A, mce3C) in different M.tuberculosis isolates (MDR1, MDR2, and sensitive isolate) at early exponential and stationary phases using real-time quantitative PCR. The expression ratio of all selected mce operon genes in all M. tuberculosis isolates was reduced at the initial phase and increased substantially at a later phase of growth. Higher expression of mce1 operon genes was found in all M. tuberculosis isolates as compared to other mce operon genes (mce2 and mce3 operons) at stationary growth phase. the higher expression of mce operon genes at stationary phase (as compared to early exponential phase) suggested growth phase dependent expression of mce operon genes. This indicated that the mce operon genes might have a role in M. tuberculosis survival and adaptation on the onset of adverse condition like stationary phase. Identification of differentially expressed genes will add to our understanding of the bacilli involved in adaptation to different growth conditions.

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 ...

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

Science.gov (United States)

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.

Variations in the occurrence of specific rpoB mutations in rifampicin-resistant Mycobacterium tuberculosis isolates from patients of different ethnic groups in Kuwait.

Science.gov (United States)

Ahmad, Suhail; Al-Mutairi, Noura M; Mokaddas, Eiman

2012-05-01

Frequency of resistance-conferring mutations vary among isoniazid- and ethambutol-resistant Mycobacterium tuberculosis isolates obtained from patients of various ethnic groups. This study was aimed to determine the occurrence of specific rpoB mutations in rifampicin-resistant M. tuberculosis isolates from tuberculosis patients of various ethnic groups in Kuwait. Rifampicin-resistant M. tuberculosis isolates (n=119) from South Asian (n=55), Southeast Asian (n=23), Middle Eastern (n=39) and other (n=2) patients and 107 rifampicin-susceptible isolates were tested. Mutations in rpoB were detected by DNA sequencing. Polymorphisms at katG463 and gyrA95 were detected by PCR-RFLP for genetic group assignment. None of rifampicin-susceptible but 116 of 119 rifampicin-resistant isolates showed rpoB mutation(s). Mutations among isolates from South Asian patients were distributed at rpoB516 (20%), rpoB526 (24%) and rpoB531 (27%) while 78 and 51 per cent of isolates from Southeast Asian and Middle Eastern patients, respectively, contained a mutated rpoB531. All isolates with rpoB N-terminal and cluster II mutations were obtained from Middle Eastern and South Asian patients. Most isolates from South Asian (84%) and Southeast Asian (70%) patients belonged to genetic group I while nearly all remaining isolates belonged to genetic group II. Isolates from Middle Eastern patients were distributed among genetic group I (46%), genetic group II (33%) and genetic group III (21%). The occurrence of specific rpoB mutations varied considerably in rifampicin-resistant M. tuberculosis isolates obtained from patients of different ethnic groups within the same country. The present data have important implications for designing region-specific rapid methods for detecting majority of rifampicin-resistant strains.

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.

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

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

Energy Technology Data Exchange (ETDEWEB)

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

First worldwide proficiency study on variable-number tandem-repeat typing of Mycobacterium tuberculosis complex strains.

NARCIS (Netherlands)

Beer, J.L. de; Kremer, K.; Kodmon, C.; Supply, P.; Soolingen, D. van

2012-01-01

Although variable-number tandem-repeat (VNTR) typing has gained recognition as the new standard for the DNA fingerprinting of Mycobacterium tuberculosis complex (MTBC) isolates, external quality control programs have not yet been developed. Therefore, we organized the first multicenter proficiency

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

Science.gov (United States)

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.

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 ...

Cytosolic proteome profiling of aminoglycosides resistant Mycobacterium tuberculosis clinical isolates using MALDI-TOF/MS

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Divakar Sharma

2016-11-01

Full Text Available Emergence of extremely drug resistant tuberculosis (XDR-TB is the consequence of the failure of second line TB treatment. Aminoglycosides are the important second line anti-TB drugs used to treat the multi drug resistant tuberculosis (MDR-TB. Main known mechanism of action of aminoglycosides is to inhibit the protein synthesis by inhibiting the normal functioning of ribosome. Primary target of aminoglycosides are the ribosomal RNA and its associated proteins. Various mechanisms have been proposed for aminoglycosides resistance but still some are unsolved. As proteins are involved in most of the biological processes, these act as a potential diagnostic markers and drug targets. In the present study we analyzed the purely cytosolic proteome of amikacin (AK and kanamycin (KM resistant Mycobacterium tuberculosis isolates by proteomic and bioinformatic approaches. Twenty protein spots were found to have over expressed in resistant isolates and were identified. Among these Rv3208A, Rv2623, Rv1360, Rv2140c, Rv1636 and Rv2185c are six proteins with unknown functions or undefined role. Docking results showed that AK and KM binds to the conserved domain (DUF, USP-A, Luciferase, PEBP and Polyketidecyclase/dehydrase domain of these hypothetical proteins and over expression of these proteins might neutralize/modulate the effect of drug molecules. TBPred and GPS-PUP predicted cytoplasmic nature and potential pupylation sites within these identified proteins respectively. String analysis also suggested that over expressed proteins along with their interactive partners might be involved in aminoglycosides resistance. Cumulative effect of these over expressed proteins could be involved in AK and KM resistance by mitigating the toxicity, repression of drug target and neutralizing affect. These findings need further exploitation for the expansion of newer therapeutics or diagnostic markers against AK and KM resistance so that an extreme condition like XDR-TB can

Cytosolic Proteome Profiling of Aminoglycosides Resistant Mycobacterium tuberculosis Clinical Isolates Using MALDI-TOF/MS.

Science.gov (United States)

Sharma, Divakar; Lata, Manju; Singh, Rananjay; Deo, Nirmala; Venkatesan, Krishnamurthy; Bisht, Deepa

2016-01-01

Emergence of extensively drug resistant tuberculosis (XDR-TB) is the consequence of the failure of second line TB treatment. Aminoglycosides are the important second line anti-TB drugs used to treat the multi drug resistant tuberculosis (MDR-TB). Main known mechanism of action of aminoglycosides is to inhibit the protein synthesis by inhibiting the normal functioning of ribosome. Primary target of aminoglycosides are the ribosomal RNA and its associated proteins. Various mechanisms have been proposed for aminoglycosides resistance but still some are unsolved. As proteins are involved in most of the biological processes, these act as a potential diagnostic markers and drug targets. In the present study we analyzed the purely cytosolic proteome of amikacin (AK) and kanamycin (KM) resistant Mycobacterium tuberculosis isolates by proteomic and bioinformatic approaches. Twenty protein spots were found to have over expressed in resistant isolates and were identified. Among these Rv3208A, Rv2623, Rv1360, Rv2140c, Rv1636, and Rv2185c are six proteins with unknown functions or undefined role. Docking results showed that AK and KM binds to the conserved domain (DUF, USP-A, Luciferase, PEBP and Polyketidecyclase/dehydrase domain) of these hypothetical proteins and over expression of these proteins might neutralize/modulate the effect of drug molecules. TBPred and GPS-PUP predicted cytoplasmic nature and potential pupylation sites within these identified proteins, respectively. String analysis also suggested that over expressed proteins along with their interactive partners might be involved in aminoglycosides resistance. Cumulative effect of these over expressed proteins could be involved in AK and KM resistance by mitigating the toxicity, repression of drug target and neutralizing affect. These findings need further exploitation for the expansion of newer therapeutics or diagnostic markers against AK and KM resistance so that an extreme condition like XDR-TB can be prevented.

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

[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.

Genetic diversity of Mycobacterium tuberculosis isolates from central India

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Prabha Desikan

2016-01-01

Interpretation & conclusions: Well established SITs were found to be predominant in our study. SIT26/CAS1_DEL was the most predominant type. However, the occurrence of a substantial number of orphan isolates may indicate the presence of active spatial and temporal evolutionary dynamics within the isolates of M. tuberculosis.

Shortening Isolation of Patients With Suspected Tuberculosis by Using Polymerase Chain Reaction Analysis

DEFF Research Database (Denmark)

Fløe, Andreas; Hilberg, Ole; Thomsen, Vibeke Østergaard

2015-01-01

Background. Isolation of patients suspected for pulmonary tuberculosis is guided by serial sputum smears. This can result in isolation for days for patients with noncontagious tuberculosis. To determine whether a single sample negative for Mycobacterium tuberculosis complex at polymerase chain...... reaction (PCR) can guide isolation. Methods. We retrospectively evaluated sputum samples analyzed for M. tuberculosis complex at the International Reference Laboratory of Mycobacteriology, Copenhagen, Denmark in 2002–2011. We selected culture-confirmed tuberculosis cases with ≥3 samples within 14 days...... before or after the initial culture-positive sample. We repeated the process for those with ≥2 samples within 28 days. The primary outcome was PCR-negative, smear-positive patients. Results. We included 53 533 sputum samples from 20 928 individuals; 1636 had culture-confirmed tuberculosis. Of these, 856...

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

Science.gov (United States)

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.

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

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

Science.gov (United States)

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

Proteome analysis of ofloxacin and moxifloxacin induced mycobacterium tuberculosis isolates by proteomic approach.

Science.gov (United States)

Lata, Manju; Sharma, Divakar; Kumar, Bhavnesh; Deo, Nirmala; Tiwari, Pramod Kumar; Bisht, Deepa; Venkatesan, Krishnamurthy

2015-01-01

Ofloxacin (OFX) and moxifloxacin (MOX) are the most promising second line drugs for tuberculosis treatment. Although the primary mechanism of action of OFX and MOX is gyrase inhibition, other possible mechanisms cannot be ruled out. Being the functional moiety of cell, the proteins act as primary targets for developing drugs, diagnostics and therapeutics. In this study we have investigated the proteomic changes of Mycobacterium tuberculosis isolates induced by OFX and MOX by applying comparative proteomic approaches based on two-dinensional gel electrophoresis (2DE) along with matrix assisted laser desorption ionisation time of flight mass spectrometry (MALDI TOF/TOF-MS) and bioinformatic tools. The findings are likely to provide new understanding of OFX and MOX mechanisms that might be helpful in exploring new diagnostics and drug targets. Our study explored eleven proteins (Rv2889c, Rv2623, Rv0952, Rv1827, Rv1932, Rv0054, Rv1080c, Rv3418c, Rv3914, Rv1636 and Rv0009) that were overexpressed in the presence of drugs. Among them, Rv2623, Rv1827 and Rv1636 were identified as proteins with unknown function. InterProScan and molecular docking revealed that the conserved domain of hypothetical proteins interact with OFX and MOX which indicate a probable inhibition/modulation of the functioning of these proteins by both drugs, which might be overexpressed to overcome this effect.

The Association between Mycobacterium Tuberculosis Genotype and Drug Resistance in Peru.

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Louis Grandjean

Full Text Available The comparison of Mycobacterium tuberculosis bacterial genotypes with phenotypic, demographic, geospatial and clinical data improves our understanding of how strain lineage influences the development of drug-resistance and the spread of tuberculosis.To investigate the association of Mycobacterium tuberculosis bacterial genotype with drug-resistance. Drug susceptibility testing together with genotyping using both 15-loci MIRU-typing and spoligotyping, was performed on 2,139 culture positive isolates, each from a different patient in Lima, Peru. Demographic, geospatial and socio-economic data were collected using questionnaires, global positioning equipment and the latest national census.The Latin American Mediterranean (LAM clade (OR 2.4, p<0.001 was significantly associated with drug-resistance and alone accounted for more than half of all drug resistance in the region. Previously treated patients, prisoners and genetically clustered cases were also significantly associated with drug-resistance (OR's 2.5, 2.4 and 1.8, p<0.001, p<0.05, p<0.001 respectively.Tuberculosis disease caused by the LAM clade was more likely to be drug resistant independent of important clinical, genetic and socio-economic confounding factors. Explanations for this include; the preferential co-evolution of LAM strains in a Latin American population, a LAM strain bacterial genetic background that favors drug-resistance or the "founder effect" from pre-existing LAM strains disproportionately exposed to drugs.

Proteomic analysis of drug-resistant Mycobacterium tuberculosis by one-dimensional gel electrophoresis and charge chromatography.

Science.gov (United States)

Yari, Shamsi; Hadizadeh Tasbiti, Alireza; Ghanei, Mostafa; Shokrgozar, Mohammad Ali; Fateh, Abolfazl; Mahdian, Reza; Yari, Fatemeh; Bahrmand, Ahmadreza

2017-01-01

Multidrug-resistant tuberculosis (MDR-TB) is a form of TB caused by Mycobacterium tuberculosis (M. tuberculosis) that do not respond to, at least, isoniazid and rifampicin, the two most powerful, first-line (or standard) anti-TB drugs. Novel intervention strategies for eliminating this disease were based on finding proteins that can be used for designing new drugs or new and reliable kits for diagnosis. The aim of this study was to compare the protein profiles of MDR-TB with sensitive isolates. Proteomic analysis of M. tuberculosis MDR-TB and sensitive isolates was obtained with ion exchange chromatography coupled with MALDI-TOF-TOF (matrix-assisted laser desorption/ionization) in order to identify individual proteins that have different expression in MDR-TB to be used as a drug target or diagnostic marker for designing valuable TB vaccines or TB rapid tests. We identified eight proteins in MDR-TB isolates, and analyses showed that these proteins are absent in M. tuberculosis-sensitive isolates: (Rv2140c, Rv0009, Rv1932, Rv0251c, Rv2558, Rv1284, Rv3699 and MMP major membrane proteins). These data will provide valuable clues in further investigation for suitable TB rapid tests or drug targets against drug-resistant and sensitive M. tuberculosis isolates.

Mycobacterium tuberculosis, Beijing genotype strains not associated with radiological presentation of pulmonary tuberculosis

NARCIS (Netherlands)

Borgdorff, Martien W.; van Deutekom, Henk; de Haas, Petra E. W.; Kremer, Kristin; van Soolingen, Dick

2004-01-01

Mycobacterium tuberculosis strains of the Beijing genotype have been involved in various outbreaks of multidrug-resistant tuberculosis. Some studies suggest that the infection with the Beijing genotype is associated with a different host immune response. Since this might also lead to a different

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...

PolyTB: A genomic variation map for Mycobacterium tuberculosis

KAUST Repository

Coll, Francesc

2014-02-15

Tuberculosis (TB) caused by Mycobacterium tuberculosis (Mtb) is the second major cause of death from an infectious disease worldwide. Recent advances in DNA sequencing are leading to the ability to generate whole genome information in clinical isolates of M. tuberculosis complex (MTBC). The identification of informative genetic variants such as phylogenetic markers and those associated with drug resistance or virulence will help barcode Mtb in the context of epidemiological, diagnostic and clinical studies. Mtb genomic datasets are increasingly available as raw sequences, which are potentially difficult and computer intensive to process, and compare across studies. Here we have processed the raw sequence data (>1500 isolates, eight studies) to compile a catalogue of SNPs (n = 74,039, 63% non-synonymous, 51.1% in more than one isolate, i.e. non-private), small indels (n = 4810) and larger structural variants (n = 800). We have developed the PolyTB web-based tool (http://pathogenseq.lshtm.ac.uk/polytb) to visualise the resulting variation and important meta-data (e.g. in silico inferred strain-types, location) within geographical map and phylogenetic views. This resource will allow researchers to identify polymorphisms within candidate genes of interest, as well as examine the genomic diversity and distribution of strains. PolyTB source code is freely available to researchers wishing to develop similar tools for their pathogen of interest. 2014 Elsevier Ltd. All rights reserved.

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

Molecular characterization of Mycobacterium orygis isolates from wild animals of Nepal.

Science.gov (United States)

Thapa, Jeewan; Nakajima, Chie; Maharjan, Bhagwan; Poudell, Ajay; Suzuki, Yasuhiko

2015-08-01

Mycobacterium orygis, a new member of the Mycobacterium tuberculosis complex, was isolated from a captive spotted deer (Axis axis) and a blue bull (Boselaphus tragocamelus) in Nepal. Analyses by spoligotyping, mycobacterial interspersed repetitive units-variable number of tandem repeats (MIRU-VNTR) typing, region of difference and single nucleotide polymorphism of genes gyrB, mmpL6, TbD1, PPE55 and Rv2042c confirmed the isolates as M. orygis. Moreover, analyses by spoligotyping (SIT587) as well as MIRU-VNTR showed that the isolates shared a similar pattern with many reported isolates. From previous and the present studies, it can be inferred that South Asia is one of the endemic regions for M. orygis. Further investigation including a larger sample size and different host interaction will help to understand the ecology and epidemiology of M. orygis in Nepal.

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

Triple valve endocarditis by mycobacterium tuberculosis. A case report

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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,

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.

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

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

Overview of errors in the reference sequence and annotation of Mycobacterium tuberculosis H37Rv, and variation amongst its isolates

KAUST Repository

Kö ser, Claudio U.; Niemann, Stefan; Summers, David K.; Archer, John A.C.

2012-01-01

Since its publication in 1998, the genome sequence of the Mycobacterium tuberculosis H37Rv laboratory strain has acted as the cornerstone for the study of tuberculosis. In this review we address some of the practical aspects that have come to light

Intraocular manifestations of mycobacterium tuberculosis: A review of the literature

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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

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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

Molecular Characterization of the Resistance of Mycobacterium ...

African Journals Online (AJOL)

Purpose: To characterize the resistance of Mycobacterium tuberculosis to second line drugs using a line probe assay. Methods: Multi-drug resistant strains of Mycobacterium tuberculosis isolated between December 2008 and December 2009 were tested for resistance to fluoroquinolones and second-line injectable drugs ...

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

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...

Secondary typing of Mycobacterium tuberculosis isolates with matching IS6110 fingerprints from different geographic regions of the United States.

Science.gov (United States)

Yang, Z H; Bates, J H; Eisenach, K D; Cave, M D

2001-05-01

Fifty-nine isolates of Mycobacterium tuberculosis obtained from different states in the United States and representing 25 interstate clusters were investigated. These clusters were identified by computer-assisted analysis of DNA fingerprints submitted during 1996 and 1997 by different laboratories participating in the CDC National Genotyping and Surveillance Network. Isolates were fingerprinted with the IS6110 right-hand probe (IS6110-3'), the IS6110 left-hand probe (IS6110-5'), and the probe pTBN12, containing the polymorphic GC-rich sequence (PGRS). Spoligotyping based on the polymorphism in the 36-bp direct-repeat locus was also performed. As a control, 43 M. tuberculosis isolates in 17 clusters obtained from patients in Arkansas during the study period were analyzed. Of the 25 interstate clusters, 19 were confirmed as correctly clustered when all the isolates were analyzed on the same gel using the IS6110-3' probe. Of the 19 true IS6110-3' clusters, 10 (53%) were subdivided by one or more secondary typing methods. Clustering of the control group was virtually identical by all methods. Of the three different secondary typing methods, spoligotyping was the least discriminating. IS6110-5' fingerprinting was as discriminating as PGRS fingerprinting. The data indicate that the IS6110-5' probe not only is a useful secondary typing method but also probably would prove to be a more useful primary typing method for a genotyping network which involves isolates from different geographic regions.

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...

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

Science.gov (United States)

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

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...

Clonality and Micro-Diversity of a Nationwide Spreading Genotype of Mycobacterium tuberculosis in Japan

Science.gov (United States)

Wada, Takayuki; Iwamoto, Tomotada; Tamaru, Aki; Seto, Junji; Ahiko, Tadayuki; Yamamoto, Kaori; Hase, Atushi; Maeda, Shinji; Yamamoto, Taro

2015-01-01

Mycobacterium tuberculosis transmission routes can be estimated from genotypic analysis of clinical isolates from patients. In Japan, still a middle-incidence country of TB, a unique genotype strain designated as ‘M-strain’ has been isolated nationwide recently. To ascertain the history of the wide spread of the strain, 10 clinical isolates from different areas were subjected to genome-wide analysis based on deep sequencers. Results show that all isolates possessed common mutations to those of referential strains. The greatest number of accumulated single nucleotide variants (SNVs) from the oldest coalescence was 13 nucleotides, indicating high clonality of these isolates. When an SNV common to the isolates was used as a surrogate marker of the clone, authentic clonal isolates with variation in a reliable subset of variable number of tandem repeat (VNTR) genotyping method can be selected successfully from clinical isolates populations of M. tuberculosis. When the authentic clones can also be assigned to sub-clonal groups by SNVs derived from the genomic comparison, they are classifiable into three sub-clonal groups with a bias of geographical origins. Feedback from genomic analysis of clinical isolates of M. tuberculosis to genotypic markers will be an efficient strategy for the big data in various settings for public health actions against TB. PMID:25734518

Antibacterial Activity of Medicinal Aqueous Plant Extracts against Mycobacterium tuberculosis

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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.

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...

Mycobacterium tuberculosis Genotype and Case Notification Rates, Rural Vietnam, 2003-2006

NARCIS (Netherlands)

Buu, T.N.; Huyen, M.N.T.; Lan, N.N.T.; Quy, H.T.; Hen, N.V.; Zignol, M.; Borgdorff, M.W.; van Soolingen, D.; Cobelens, F.G.J.

2009-01-01

Tuberculosis case notification rates (CNRs) for young adults in Vietnam are increasing. To determine whether this finding could reflect emergence of Mycobacterium tuberculosis Beijing genotype, we studied all new sputum smear-positive pulmonary tuberculosis patients registered for treatment in 3

Increased level of acute phase reactants in patients infected with modern Mycobacterium tuberculosis genotypes in Mwanza, Tanzania

DEFF Research Database (Denmark)

Stavrum, Ruth; PrayGod, George; Range, Nyagosya

2014-01-01

a distinct genetic ancestry. This study describes the genetic biodiversity of M. tuberculosis genotypes in Mwanza city, Tanzania and the clinical presentation of the disease caused by isolates of different lineages. METHODS: Two-hundred-fifty-two isolates from pulmonary TB patients in Mwanza, Tanzania were......BACKGROUND: There is increasing evidence to suggest that different Mycobacterium tuberculosis lineages cause variations in the clinical presentation of tuberculosis (TB). Certain M. tuberculosis genotypes/lineages have been shown to be more likely to cause active TB in human populations from....... tuberculosis lineage of the infectious agent for each patient. RESULTS: The most frequent genotype was ST59 (48 out of 248 [19.4%]), belonging to the Euro-American lineage LAM11_ZWE, followed by ST21 (CAS_KILI lineage [44 out of 248 [17.7%]). A low degree of diversity (15.7% [39 different ST's out of 248...

Mycobacterium tuberculosis strains of the Beijing genotype are rarely observed in tuberculosis patients in South America.

Science.gov (United States)

Ritacco, Viviana; López, Beatriz; Cafrune, Patricia I; Ferrazoli, Lucilaine; Suffys, Philip N; Candia, Norma; Vásquez, Lucy; Realpe, Teresa; Fernández, Jorge; Lima, Karla V; Zurita, Jeannete; Robledo, Jaime; Rossetti, Maria L; Kritski, Afranio L; Telles, Maria A; Palomino, Juan C; Heersma, Herre; van Soolingen, Dick; Kremer, Kristin; Barrera, Lucía

2008-08-01

The frequency of the Beijing genotype of Mycobacterium tuberculosis as a cause of tuberculosis (TB) in South America was determined by analyzing genotypes of strains isolated from patients that had been diagnosed with the disease between 1997 and 2003 in seven countries of the subcontinent. In total, 19 of the 1,202 (1.6%) TB cases carried Beijing isolates, including 11 of the 185 patients from Peru (5.9%), five of the 512 patients from Argentina (1.0%), two of the 252 Brazilian cases (0.8%), one of the 166 patients from Paraguay (0.6%) and none of the samples obtained from Chile (35), Colombia (36) and Ecuador (16). Except for two patients that were East Asian immigrants, all cases with Beijing strains were native South Americans. No association was found between carrying a strain with the Beijing genotype and having drug or multi-drug resistant disease. Our data show that presently transmission of M. tuberculosis strains of the Beijing genotype is not frequent in Latin America. In addition, the lack of association of drug resistant TB and infection with M. tuberculosis of the Beijing genotype observed presently demands efforts to define better the contribution of the virulence and lack of response to treatment to the growing spread of Beijing strains observed in other parts of the world.

Mycobacterium tuberculosis strains of the Beijing genotype are rarely observed in tuberculosis patients in South America

Directory of Open Access Journals (Sweden)

Viviana Ritacco

2008-08-01

Full Text Available The frequency of the Beijing genotype of Mycobacterium tuberculosis as a cause of tuberculosis (TB in South America was determined by analyzing genotypes of strains isolated from patients that had been diagnosed with the disease between 1997 and 2003 in seven countries of the subcontinent. In total, 19 of the 1,202 (1.6% TB cases carried Beijing isolates, including 11 of the 185 patients from Peru (5.9%, five of the 512 patients from Argentina (1.0%, two of the 252 Brazilian cases (0.8%, one of the 166 patients from Paraguay (0.6% and none of the samples obtained from Chile (35, Colombia (36 and Ecuador (16. Except for two patients that were East Asian immigrants, all cases with Beijing strains were native South Americans. No association was found between carrying a strain with the Beijing genotype and having drug or multi-drug resistant disease. Our data show that presently transmission of M. tuberculosis strains of the Beijing genotype is not frequent in Latin America. In addition, the lack of association of drug resistant TB and infection with M. tuberculosis of the Beijing genotype observed presently demands efforts to define better the contribution of the virulence and lack of response to treatment to the growing spread of Beijing strains observed in other parts of the world.

Correlates between Models of Virulence for Mycobacterium tuberculosis among Isolates of the Central Asian Lineage: a Case for Lysozyme Resistance Testing?

Science.gov (United States)

Casali, Nicola; Clark, Simon O.; Hooper, Richard; Williams, Ann; Velji, Preya; Gonzalo, Ximena

2015-01-01

Virulence factors (VFs) contribute to the emergence of new human Mycobacterium tuberculosis strains, are lineage dependent, and are relevant to the development of M. tuberculosis drugs/vaccines. VFs were sought within M. tuberculosis lineage 3, which has the Central Asian (CAS) spoligotype. Three isolates were selected from clusters previously identified as dominant in London, United Kingdom. Strain-associated virulence was studied in guinea pig, monocyte-derived macrophage, and lysozyme resistance assays. Whole-genome sequencing, single nucleotide polymorphism (SNP) analysis, and a literature review contributed to the identification of SNPs of interest. The animal model revealed borderline differences in strain-associated pathogenicity. Ex vivo, isolate C72 exhibited statistically significant differences in intracellular growth relative to C6 and C14. SNP candidates inducing lower fitness levels included 123 unique nonsynonymous SNPs, including three located in genes (lysX, caeA, and ponA2) previously identified as VFs in the laboratory-adapted reference strain H37Rv and shown to confer lysozyme resistance. C72 growth was most affected by lysozyme in vitro. A BLAST search revealed that all three SNPs of interest (C35F, P76Q, and P780R) also occurred in Tiruvallur, India, and in Uganda. Unlike C72, however, no single isolate identified through BLAST carried all three SNPs simultaneously. CAS isolates representative of three medium-sized human clusters demonstrated differential outcomes in models commonly used to estimate strain-associated virulence, supporting the idea that virulence varies within, not just across, M. tuberculosis lineages. Three VF SNPs of interest were identified in two additional locations worldwide, which suggested independent selection and supported a role for these SNPs in virulence. The relevance of lysozyme resistance to strain virulence remains to be established. PMID:25776753

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....

Mycobacterium tuberculosis Lineage 4 comprises globally distributed and geographically restricted sublineages

Science.gov (United States)

Coscolla, Mireia; Liu, Qingyun; Trauner, Andrej; Fenner, Lukas; Rutaihwa, Liliana; Borrell, Sonia; Luo, Tao; Gao, Qian; Kato-Maeda, Midori; Ballif, Marie; Egger, Matthias; Macedo, Rita; Mardassi, Helmi; Moreno, Milagros; Tudo Vilanova, Griselda; Fyfe, Janet; Globan, Maria; Thomas, Jackson; Jamieson, Frances; Guthrie, Jennifer L.; Asante-Poku, Adwoa; Yeboah-Manu, Dorothy; Wampande, Eddie; Ssengooba, Willy; Joloba, Moses; Henry Boom, W.; Basu, Indira; Bower, James; Saraiva, Margarida; Vaconcellos, Sidra E. G.; Suffys, Philip; Koch, Anastasia; Wilkinson, Robert; Gail-Bekker, Linda; Malla, Bijaya; Ley, Serej D.; Beck, Hans-Peter; de Jong, Bouke C.; Toit, Kadri; Sanchez-Padilla, Elisabeth; Bonnet, Maryline; Gil-Brusola, Ana; Frank, Matthias; Penlap Beng, Veronique N.; Eisenach, Kathleen; Alani, Issam; Wangui Ndung’u, Perpetual; Revathi, Gunturu; Gehre, Florian; Akter, Suriya; Ntoumi, Francine; Stewart-Isherwood, Lynsey; Ntinginya, Nyanda E.; Rachow, Andrea; Hoelscher, Michael; Cirillo, Daniela Maria; Skenders, Girts; Hoffner, Sven; Bakonyte, Daiva; Stakenas, Petras; Diel, Roland; Crudu, Valeriu; Moldovan, Olga; Al-Hajoj, Sahal; Otero, Larissa; Barletta, Francesca; Jane Carter, E.; Diero, Lameck; Supply, Philip; Comas, Iñaki; Niemann, Stefan; Gagneux, Sebastien

2016-01-01

Generalist and specialist species differ in the breadth of their ecological niche. Little is known about the niche width of obligate human pathogens. Here we analyzed a global collection of Mycobacterium tuberculosis Lineage 4 clinical isolates, the most geographically widespread cause of human tuberculosis. We show that Lineage 4 comprises globally distributed and geographically restricted sublineages, suggesting a distinction between generalists and specialists. Population genomic analyses showed that while the majority of human T cell epitopes were conserved in all sublineages, the proportion of variable epitopes was higher in generalists. Our data further support a European origin for the most common generalist sublineage. Hence, the global success of Lineage 4 reflects distinct strategies adopted by different sublineages and the influence of human migration. PMID:27798628

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 ...

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

Science.gov (United States)

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-Kopeć, Ewa; Ballif, Marie; Barletta, Francesca; Beck, Hans Peter; Barry, Clifton E; Bonnet, Maryline; Borroni, Emanuele; Campos-Herrero, Isolina; Cirillo, Daniela; Cox, Helen; Crowe, Suzanne; Crudu, Valeriu; Diel, Roland; Drobniewski, Francis; Fauville-Dufaux, Maryse; Gagneux, Sébastien; Ghebremichael, Solomon; Hanekom, Madeleine; Hoffner, Sven; Jiao, Wei-wei; Kalon, Stobdan; Kohl, Thomas A; Kontsevaya, Irina; Lillebæk, Troels; Maeda, Shinji; Nikolayevskyy, Vladyslav; Rasmussen, Michael; Rastogi, Nalin; Samper, Sofia; Sanchez-Padilla, Elisabeth; Savic, Branislava; Shamputa, Isdore Chola; Shen, Adong; Sng, Li-Hwei; Stakenas, Petras; Toit, Kadri; Varaine, Francis; Vukovic, Dragana; Wahl, Céline; Warren, Robin; Supply, Philip; Niemann, Stefan; Wirth, Thierry

2015-03-01

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 radiated worldwide in several waves. We detected successive increases in population size for this pathogen over the last 200 years, practically coinciding with the Industrial Revolution, the First World War and HIV epidemics. Two MDR clones of this lineage started to spread throughout central Asia and Russia concomitantly with the collapse of the public health system in the former Soviet Union. Mutations identified in genes putatively under positive selection and associated with virulence might have favored the expansion of the most successful branches of the lineage.

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

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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

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.

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

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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%.

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

Science.gov (United States)

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.

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

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

Conserved hypothetical protein Rv1977 in Mycobacterium tuberculosis strains contains sequence polymorphisms and might be involved in ongoing immune evasion.

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Jiang, Yi; Liu, Haican; Wang, Xuezhi; Li, Guilian; Qiu, Yan; Dou, Xiangfeng; Wan, Kanglin

2015-01-01

Host immune pressure and associated parasite immune evasion are key features of host-pathogen co-evolution. A previous study showed that human T cell epitopes of Mycobacterium tuberculosis are evolutionarily hyperconserved and thus it was deduced that M. tuberculosis lacks antigenic variation and immune evasion. Here, we selected 151 clinical Mycobacterium tuberculosis isolates from China, amplified gene encoding Rv1977 and compared the sequences. The results showed that Rv1977, a conserved hypothetical protein, is not conserved in M. tuberculosis strains and there are polymorphisms existed in the protein. Some mutations, especially one frameshift mutation, occurred in the antigen Rv1977, which is uncommon in M.tb strains and may lead to the protein function altering. Mutations and deletion in the gene all affect one of three T cell epitopes and the changed T cell epitope contained more than one variable position, which may suggest ongoing immune evasion.

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...

Bacteriological and virulence study of a Mycobacterium chimaera isolate from a patient in China.

Science.gov (United States)

Liu, Guan; Chen, Su-Ting; Yu, Xia; Li, Yu-Xun; Ling, Ying; Dong, Ling-Ling; Zheng, Su-Hua; Huang, Hai-Rong

2015-04-01

A clinical isolate from a patient was identified as Mycobacterium chimaera, a recently identified species of nontuberculous Mycobacteria. The biochemical and molecular identity, drug sensitivity and virulence of this isolated strain were investigated. 16S rRNA, the 16S-23S ITS, hsp65 and rpoB were amplified, and their sequence similarities with other mycobacteria were analyzed. The minimum inhibitory concentrations of 22 anti-microbial agents against this isolate were established, and the virulence of the isolate was evaluated by intravenous injection into C57BL/6 mice using Mycobacterium tuberculosis H37Rv as a control strain. Growth and morphological characteristics and mycolic acid profile analysis revealed that this isolated strain was a member of the Mycobacterium avium complex. BLAST analysis of the amplified sequences showed that the isolated strain was closely related to M. chimaera. Susceptibility testing showed that the isolate was sensitive to rifabutin, rifapentine, clarithromycin, azithromycin, imipenem and cefoxitin. Bacterial load determination and tissue histopathology of the infected mice indicated that the isolate was highly virulent. The first case of M. chimaera infection in China was evaluated. The information derived from this case may offer valuable guidance for clinical diagnosis and treatment.

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

Global transcriptional profiling of longitudinal clinical isolates of Mycobacterium tuberculosis exhibiting rapid accumulation of drug resistance.

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Anirvan Chatterjee

Full Text Available The identification of multidrug resistant (MDR, extensively and totally drug resistant Mycobacterium tuberculosis (Mtb, in vulnerable sites such as Mumbai, is a grave threat to the control of tuberculosis. The current study aimed at explaining the rapid expression of MDR in Directly Observed Treatment Short Course (DOTS compliant patients, represents the first study comparing global transcriptional profiles of 3 pairs of clinical Mtb isolates, collected longitudinally at initiation and completion of DOTS. While the isolates were drug susceptible (DS at onset and MDR at completion of DOTS, they exhibited identical DNA fingerprints at both points of collection. The whole genome transcriptional analysis was performed using total RNA from H37Rv and 3 locally predominant spoligotypes viz. MANU1, CAS and Beijing, hybridized on MTBv3 (BuG@S microarray, and yielded 36, 98 and 45 differentially expressed genes respectively. Genes encoding transcription factors (sig, rpoB, cell wall biosynthesis (emb genes, protein synthesis (rpl and additional central metabolic pathways (ppdK, pknH, pfkB were found to be down regulated in the MDR isolates as compared to the DS isolate of the same genotype. Up regulation of drug efflux pumps, ABC transporters, trans-membrane proteins and stress response transcriptional factors (whiB in the MDR isolates was observed. The data indicated that Mtb, without specific mutations in drug target genes may persist in the host due to additional mechanisms like drug efflux pumps and lowered rate of metabolism. Furthermore this population of Mtb, which also showed reduced DNA repair activity, would result in selection and stabilization of spontaneous mutations in drug target genes, causing selection of a MDR strain in the presence of drug pressures. Efflux pump such as drrA may play a significant role in increasing fitness of low level drug resistant cells and assist in survival of Mtb till acquisition of drug resistant mutations with

Direct DNA Extraction from Mycobacterium tuberculosis Frozen Stocks as a Reculture-Independent Approach to Whole-Genome Sequencing

DEFF Research Database (Denmark)

Bjorn-Mortensen, K; Zallet, J; Lillebaek, T

2015-01-01

Culturing before DNA extraction represents a major time-consuming step in whole-genome sequencing of slow-growing bacteria, such as Mycobacterium tuberculosis. We report a workflow to extract DNA from frozen isolates without reculturing. Prepared libraries and sequence data were comparable...... with results from recultured aliquots of the same stocks....

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)

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

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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

Chromosomal rearrangements and protein globularity changes in Mycobacterium tuberculosis isolates from cerebrospinal fluid

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Seow Hoon Saw

2016-09-01

Full Text Available Background Meningitis is a major cause of mortality in tuberculosis (TB. It is not clear what factors promote central nervous system invasion and pathology but it has been reported that certain strains of Mycobacterium tuberculosis (Mtb might have genetic traits associated with neurotropism. Methods In this study, we generated whole genome sequences of eight clinical strains of Mtb that were isolated from the cerebrospinal fluid (CSF of patients presenting with tuberculous meningitis (TBM in Malaysia, and compared them to the genomes of H37Rv and other respiratory Mtb genomes either downloaded from public databases or extracted from local sputum isolates. We aimed to find genomic features that might be distinctly different between CSF-derived and respiratory Mtb. Results Genome-wide comparisons revealed rearrangements (translocations, inversions, insertions and deletions and non-synonymous SNPs in our CSF-derived strains that were not observed in the respiratory Mtb genomes used for comparison. These rearranged segments were rich in genes for PE (proline-glutamate/PPE (proline-proline-glutamate, transcriptional and membrane proteins. Similarly, most of the ns SNPs common in CSF strains were noted in genes encoding PE/PPE proteins. Protein globularity differences were observed among mycobacteria from CSF and respiratory sources and in proteins previously reported to be associated with TB meningitis. Transcription factors and other transcription regulators featured prominently in these proteins. Homologs of proteins associated with Streptococcus pneumoniae meningitis and Neisseria meningitidis virulence were identified in neuropathogenic as well as respiratory mycobacterial spp. examined in this study. Discussion The occurrence of in silico genetic differences in CSF-derived but not respiratory Mtb suggests their possible involvement in the pathogenesis of TBM. However, overall findings in this comparative analysis support the postulation that TB

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

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

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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%.

Isolation, Identification, and Characterization of a New Highly Pathogenic Field Isolate of Mycobacterium avium spp. avium

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Liangquan Zhu

2018-01-01

Full Text Available Avian tuberculosis is a chronic, contagious zoonotic disease affecting birds, mammals, and humans. The disease is most often caused by Mycobacterium avium spp. avium (MAA. Strain resources are important for research on avian tuberculosis and vaccine development. However, there has been little reported about the newly identified MAA strain in recent years in China. In this study, a new strain was isolated from a fowl with symptoms of avian tuberculosis by bacterial culture. The isolated strain was identified to be MAA by culture, staining, and biochemical and genetic analysis, except for different colony morphology. The isolated strain was Ziehl-Zeelsen staining positive, resistant to p-nitrobenzoic acid, and negative for niacin production, Tween-80 hydrolysis, heat stable catalase and nitrate production. The strain had the DnaJ gene, IS1245, and IS901, as well. Serum agglutination indicated that the MAA strain was of serotype 1. The MAA strain showed strong virulence via mortality in rabbits and chickens. The prepared tuberculin of the MAA strain had similar potency compared to the MAA reference strain and standard tuberculin via a tuberculin skin test. Our studies suggested that this MAA strain tends to be a novel subtype, which might enrich the strain resource of avian tuberculosis.

Isolation and molecular characterization of Mycobacterium bovis causing pulmonary tuberculosis and epistaxis in a Thoroughbred horse.

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Hlokwe, Tiny Motlatso; Sutton, David; Page, Patrick; Michel, Anita Luise

2016-09-02

Tuberculosis caused by Mycobacterium bovis (M. bovis) is very uncommon in horses worldwide. In the current study, an eight-year-old male Thoroughbred in good body condition was admitted to the Equine Clinic at the Onderstepoort Veterinary Academic Hospital in 2005 due to bilateral epistaxis accompanied by coughing. Routine examinations were conducted to determine the cause of the condition. Endoscopic examination revealed the major source of the epistaxis as the trachea, whereas thoracic radiography indicated the presence of a primary pulmonary mass. M. bovis was isolated from a broncho-alveolar lavage (BAL) sample collected. The pulmonary mass reduced in size three months later following an oral administration of enrofloxacin (7.5 mg/kg PO SID). Genetic fingerprinting by spoligotyping identified the M. bovis isolate as spoligotype SB0868 strain. This M. bovis strain type was never described previously in South Africa (SA). This is the first case of M. bovis infection in a horse in SA which has been fully documented including clinical findings, isolation and genetic characterisation of the causative pathogen. This report indicates that horses may contract and harbour M. bovis despite their lower susceptibility compared to other domestic animals. It also suggests that the infection may be more easily contained and eliminated from the host.

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

Science.gov (United States)

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.

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

Antimicrobial susceptibility determined by the E test, Löwenstein-Jensen proportion, and DNA sequencing methods among Mycobacterium tuberculosis isolates discrepancies, preliminary results

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Maria Inês Moura Freixo

2004-02-01

Full Text Available Mycobacterium tuberculosis strains resistant to streptomycin (SM, isoniazid (INH, and/or rifampin (RIF as determined by the conventional Löwenstein-Jensen proportion method (LJPM were compared with the E test, a minimum inhibitory concentration susceptibility method. Discrepant isolates were further evaluated by BACTEC and by DNA sequence analyses for mutations in genes most often associated with resistance to these drugs (rpsL, katG, inhA, and rpoB. Preliminary discordant E test results were seen in 75% of isolates resistant to SM and in 11% to INH. Discordance improved for these two drugs (63% for SM and none for INH when isolates were re-tested but worsened for RIF (30%. Despite good agreement between phenotypic results and sequencing analyses, wild type profiles were detected on resistant strains mainly for SM and INH. It should be aware that susceptible isolates according to molecular methods might contain other mechanisms of resistance. Although reproducibility of the LJPM susceptibility method has been established, variable E test results for some M. tuberculosis isolates poses questions regarding its reproducibility particularly the impact of E test performance which may vary among laboratories despite adherence to recommended protocols. Further studies must be done to enlarge the evaluated samples and looked possible mutations outside of the hot spot sequenced gene among discrepant strains.

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

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 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.

Correlations between major risk factors and closely related Mycobacterium tuberculosis isolates grouped by three current enotyping 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

2014-01-01

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

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

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

Directory of Open Access Journals (Sweden)

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.

Rapid antibiotic susceptibility testing of Mycobacterium tuberculosis : Its utility in resource poor settings

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Poojary A

2006-01-01

Full Text Available Purpose: To compare the rapid colorimetric nitrate reductase based antibiotic susceptibility (CONRAS test performed on Mycobacterium tuberculosis isolates with the conventional method i.e., the proportion method. Methods: One hundred clinical isolates of M. tuberculosis were tested for susceptibility to isoniazid (INH and rifampicin (RIF by the conventional proportion method and CONRAS in Middlebrook 7H9 liquid medium enriched with growth supplements (MB7H9S. Results: The performance of the CONRAS test was evaluated using proportion method as the gold standard. The sensitivity (ability to detect true drug resistance and specificity (ability to detect true drug susceptibility of the CONRAS test to INH was 93.75 and 98.52% and for RIF it was 96.10 and 100% respectively. The mean time for reporting was 6.3 days and the test showed excellent reproducibility. The kappa (k value for INH was 0.92 and for RIF was 0.99, indicating excellent agreement between the two methods. Conclusions: CONRAS test is a rapid and reliable method of drug susceptibility for M. tuberculosis.

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.

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.

Rapid detection of multidrug-resistant Mycobacterium tuberculosis using the malachite green decolourisation assay

Science.gov (United States)

Coban, Ahmet Yilmaz; Uzun, Meltem

2013-01-01

Early detection of drug resistance in Mycobacterium tuberculosis isolates allows for earlier and more effective treatment of patients. The aim of this study was to investigate the performance of the malachite green decolourisation assay (MGDA) in detecting isoniazid (INH) and rifampicin (RIF) resistance in M. tuberculosis clinical isolates. Fifty M. tuberculosis isolates, including 19 multidrug-resistant, eight INH-resistant and 23 INH and RIF-susceptible samples, were tested. The sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV) and agreement of the assay for INH were 92.5%, 91.3%, 92.5%, 91.3% and 92%, respectively. Similarly, the sensitivity, specificity, PPV, NPV and agreement of the assay for RIF were 94.7%, 100%, 100%, 96.8% and 98%, respectively. There was a major discrepancy in the tests of two isolates, as they were sensitive to INH by the MGDA test, but resistant by the reference method. There was a minor discrepancy in the tests of two additional isolates, as they were sensitive to INH by the reference method, but resistant by the MGDA test. The drug susceptibility test results were obtained within eight-nine days. In conclusion, the MGDA test is a reliable and accurate method for the rapid detection of INH and RIF resistance compared with the reference method and the MGDA test additionally requires less time to obtain results. PMID:24402143

Rapid detection of multidrug-resistant Mycobacterium tuberculosis using the malachite green decolourisation assay

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Ahmet Yilmaz Coban

2013-12-01

Full Text Available Early detection of drug resistance in Mycobacterium tuberculosis isolates allows for earlier and more effective treatment of patients. The aim of this study was to investigate the performance of the malachite green decolourisation assay (MGDA in detecting isoniazid (INH and rifampicin (RIF resistance in M. tuberculosis clinical isolates. Fifty M. tuberculosis isolates, including 19 multidrug-resistant, eight INH-resistant and 23 INH and RIF-susceptible samples, were tested. The sensitivity, specificity, positive predictive value (PPV, negative predictive value (NPV and agreement of the assay for INH were 92.5%, 91.3%, 92.5%, 91.3% and 92%, respectively. Similarly, the sensitivity, specificity, PPV, NPV and agreement of the assay for RIF were 94.7%, 100%, 100%, 96.8% and 98%, respectively. There was a major discrepancy in the tests of two isolates, as they were sensitive to INH by the MGDA test, but resistant by the reference method. There was a minor discrepancy in the tests of two additional isolates, as they were sensitive to INH by the reference method, but resistant by the MGDA test. The drug susceptibility test results were obtained within eight-nine days. In conclusion, the MGDA test is a reliable and accurate method for the rapid detection of INH and RIF resistance compared with the reference method and the MGDA test additionally requires less time to obtain results.

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

Full Text Available Abstract Background In Ethiopia where there is no strong surveillance system and state of the art diagnostic facilities are limited, the real burden of tuberculosis (TB is not well known. We conducted a community based survey to estimate the prevalence of pulmonary TB and spoligotype pattern of the Mycobacterium tuberculosis isolates in Southwest Ethiopia. Methods A total of 30040 adults in 10882 households were screened for pulmonary TB in Gilgel Gibe field research centre in Southwest Ethiopia. A total of 482 TB suspects were identified and smear microscopy and culture was done for 428 TB suspects. Counseling and testing for HIV/AIDS was done for all TB suspects. Spoligotyping was done to characterize the Mycobacterium tuberculosis isolates. Results Majority of the TB suspects were females (60.7% and non-literates (83.6%. Using smear microscopy, a total of 5 new and 4 old cases of pulmonary TB cases were identified making the prevalence of TB 30 per 100,000. However, using the culture method, we identified 17 new cases with a prevalence of 76.1 per 100,000. There were 4.3 undiagnosed pulmonary TB cases for every TB case who was diagnosed through the passive case detection mechanism in the health facility. Eleven isolates (64.7% belonged to the six previously known spoligotypes: T, Haarlem and Central-Asian (CAS. Six new spoligotype patterns of Mycobacterium tuberculosis, not present in the international database (SpolDB4 were identified. None of the rural residents was HIV infected and only 5 (5.5% of the urban TB suspects were positive for HIV. Conclusion The prevalence of TB in the rural community of Southwest Ethiopia is low. There are large numbers of undiagnosed TB cases in the community. However, the number of sputum smear-positive cases was very low and therefore the risk of transmitting the infection to others may be limited. Active case finding through health extension workers in the community can improve the low case detection rate

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.

Design and Construction of a Cloning Vector Containing the hspX Gene of Mycobacterium tuberculosis.

Science.gov (United States)

Yaghoubi, Atieh; Aryan, Ehsan; Zare, Hosna; Alami, Shadi; Teimourpour, Roghayeh; Meshkat, Zahra

2016-10-01

Tuberculosis (TB) is a major cause of death worldwide. Finding an effective vaccine against TB is the best way to control it. Several vaccines against this disease have been developed but none are completely protective. The aim of this study was to design and construct a cloning vector containing the Mycobacterium tuberculosis (M. tuberculosis) heat shock protein X (hspX) . First, an hspX fragment was amplified by PCR and cloned into plasmid pcDNA3.1(+) and recombinant vector was confirmed. A 435 bp hspX fragment was isolated. The fragment was 100% homologous with hspX of M. tuberculosis strain H37Rv in GenBank. In this study, the cloning vector pcDNA3.1(+), containing a 435-bp hspX fragment of M. tuberculosis , was constructed. This could be used as a DNA vaccine to induce immune responses in animal models in future studies.

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...

MIRU-VNTR typing of drug-resistant tuberculosis isolates in Greece.

Science.gov (United States)

Rovina, Nikoletta; Karabela, Simona; Constantoulakis, Pantelis; Michou, Vassiliki; Konstantinou, Konstantinos; Sgountzos, Vassileios; Roussos, Charis; Poulakis, Nikolaos

2011-08-01

The increasing immigration rate in Greece from countries with a high prevalence of Mycobacterium tuberculosis (MTB) and multidrug-resistant tuberculosis (MDR-TB) may have an impact οn the number of MDR-TB cases in Greece. The aim of this study was to genotypically characterize the MTB isolates from patients with pulmonary drug-resistant tuberculosis (DR-TB) in Greece, and to determine whether there is any association between the prevalent genotypes and drug resistance. Fifty-three drug-resistant MTB strains isolated from culture specimens of clinical material from native Greeks and immigrant patients with pulmonary tuberculosis were genotyped using the mycobacterial interspersed repetitive units-variable number of tandem repeats (MIRU-VNTR) method. The phylogenetically distinct groups of isolates identified were: the Beijing (34%), the LAM (11%), the Haarlem (24.5%), the Uganda I (9.4%), the Ural (3.8%), the Delhi/CAS (9.4%) and the Cameroon (3.8%) families. Greek patients were more likely to have monoresistant and polyresistant TB with the most prevalent isolates belonging to the Haarlem family. Among foreign-born patients with MDR-TB, the most prevalent genotypes belonged to the Beijing family. MIRU-VNTR rapidly obtained clinically useful genotyping data, by characterizing clonal MTB heterogeneity in the isolated strains. Our results underline the need for more effective antituberculosis control programs in order to control the expansion of DR-TB in Greece.

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...

Public health impact of isoniazid-resistant Mycobacterium tuberculosis strains with a mutation at amino-acid position 315 of katG: a decade of experience in The Netherlands

NARCIS (Netherlands)

van Doorn, H. R.; de Haas, P. E. W.; Kremer, K.; Vandenbroucke-Grauls, C. M. J. E.; Borgdorff, M. W.; van Soolingen, D.

2006-01-01

A previous limited study demonstrated that Mycobacterium tuberculosis isolates with a mutation at amino-acid position 315 of katG (Delta315) exhibited high-level resistance to isoniazid and were more frequently resistant to streptomycin. In the present study, isoniazid-resistant M. tuberculosis

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

Directory of Open Access Journals (Sweden)

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

Mycobacterium tuberculosis Complex Members Adapted to Wild and Domestic Animals.

Science.gov (United States)

Malone, Kerri M; Gordon, Stephen V

2017-01-01

The Mycobacterium tuberculosis complex (MTBC) is composed of several highly genetically related species that can be broadly classified into those that are human-host adapted and those that possess the ability to propagate and transmit in a variety of wild and domesticated animals. Since the initial description of the bovine tubercle bacillus, now known as Mycobacterium bovis, by Theobald Smith in the late 1800's, isolates originating from a wide range of animal hosts have been identified and characterized as M. microti, M. pinnipedii, the Dassie bacillus, M. mungi, M. caprae, M. orygis and M. suricattae. This chapter outlines the events resulting in the identification of each of these animal-adapted species, their close genetic relationships, and how genome-based phylogenetic analyses of species-specific variation amongst MTBC members is beginning to unravel the events that resulted in the evolution of the MTBC and the observed host tropism between the human- and animal-adapted member species.

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.

Different Transcriptional Profiles of Human Monocyte-Derived Dendritic Cells Infected with Distinct Strains of Mycobacterium tuberculosis and Mycobacterium bovis Bacillus Calmette-Guérin

Directory of Open Access Journals (Sweden)

Nunzia Sanarico

2011-01-01

Full Text Available In order to analyze dendritic cells (DCs activation following infection with different mycobacterial strains, we studied the expression profiles of 165 genes of human monocyte-derived DCs infected with H37Rv, a virulent Mycobacterium tuberculosis (MTB laboratory strain, CMT97, a clinical MTB isolate, Mycobacterium bovis bacillus Calmette-Guérin (BCG, Aventis Pasteur, and BCG Japan, both employed as vaccine against tuberculosis. The analysis of the gene expression reveals that, despite a set of genes similarly modulated, DCs response resulted strain dependent. In particular, H37Rv significantly upregulated EBI3 expression compared with BCG Japan, while it was the only strain that failed to release a significant IL-10 amount. Of note, BCG Japan showed a marked increase in CCR7 and TNF-α expression regarding both MTB strains and it resulted the only strain failing in exponential intracellular growth. Our results suggest that DCs display the ability to elicit a tailored strain-specific immune response.

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

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

Susceptibility of Mycobacterium tuberculosis to sulfamethoxazole, trimethoprim and their combination over a 12 year period in Taiwan.

Science.gov (United States)

Huang, Tsi-Shu; Kunin, Calvin M; Yan, Bo-Shiun; Chen, Yao-Shen; Lee, Susan Shin-Jung; Syu, Wan

2012-03-01

This study was designed to determine the susceptibility of clinical isolates of multidrug-resistant (MDR) and non-MDR Mycobacterium tuberculosis to sulfamethoxazole, trimethoprim and trimethoprim/sulfamethoxazole over a 12 year period in Taiwan. We examined a total of 117 clinical isolates of M. tuberculosis collected from Southern Taiwan, 116 from 1995 to 2006 and an extensively drug-resistant (XDR) isolate in 2009. These included 28 isolates susceptible to all four first-line agents, 52 MDR isolates and 36 isolates with a mixed combination of drug resistance patterns other than MDR and 1 XDR isolate. Sulfamethoxazole inhibited 80% growth of all 117 isolates regardless of their susceptibility to the first-line agents at an MIC(90) of 9.5 mg/L. The concentration required to inhibit 99% growth was 38 mg/L. There were no significant changes in the MIC(50) or MIC(90) of sulfamethoxazole over a 12 year period. All 117 isolates were resistant to trimethoprim at >8 mg/L. The combination of trimethoprim/sulfamethoxazole at a ratio of 1:19 had no additive or synergistic effects. Sulfamethoxazole inhibited the growth of clinical isolates of M. tuberculosis at achievable concentrations in plasma after oral administration. Susceptibility to sulfamethoxazole remained constant over a 12 year period. Trimethoprim was inactive against M. tuberculosis and trimethoprim/sulfamethoxazole provided no additional activity. Although the current and prior studies demonstrate that sulfamethoxazole is active against M. tuberculosis the search needs to continue for more active, lipid-soluble sulphonamides that are better absorbed into tissues and have improved therapeutic efficacy.

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.

Genetic diversity and distribution of Mycobacterium tuberculosis genotypes in Limpopo, South Africa.

Science.gov (United States)

Maguga-Phasha, N T C; Munyai, N S; Mashinya, F; Makgatho, M E; Mbajiorgu, E F

2017-12-12

Tuberculosis remains a major health problem and knowledge of the diversity of Mycobacterium tuberculosis strains in specific geographical regions can contribute to the control of the disease. This study describes the genetic profile of M. tuberculosis in five districts of Limpopo Province. A total 487 isolates were collected from the National Health Laboratory Services from all regions/districts of Limpopo Province. Only 215 isolates were confirmed to be M. tuberculosis by Bactec Mycobacterium Growth Indicator Tube 960® and Rhodamine-Auramine staining. Isolates were subcultured on Löwenstein-Jensen medium agar slants to validate purity. They were spoligotyped and data analysed using the international spoligotyping database 4 (SpolDB4). Of the 215 isolates, 134 (62.3%) were genotyped into 21 genotype families while 81 (37.7%) were orphans. The 81 orphans were further subjected to resolution employing SpolDB3/RIM. Overall, the study revealed a high diversity of strains of 32 predominantly the non-Beijing lineages: the LAM- LAM3 (9.8%), LAM9 (4.7%) and LAM11- ZWE (3.3%), the T-T1(15.0%), T2 (0.9%), T2-T3 (1.4%), the CAS-CAS1-Delhi 5 (1.9%) and CAS1-KILI (1.4%) the MANU2 (1.4%), U (0.5%), X-X1(1.4%), X3 (1.9%), S (9.8%), CAS (1.4%), LAM7(0.9%), T3(0.5%), LAM8(4.7%), T4(1.4%), X2(0.4%), AI5(1.9%), LAM1(0.5%), FAMILY33 (1.9%), EAI4(1.4%), M. microti (1.9%). The Beijing and Beijing-like families were (14.9%) and (0.9%), respectively. A total of 28(13%) clusters and 77(36%) unique cases were identified. Beijing strain (SIT 1) formed the biggest cluster constituting 14%, followed by LAM3 (SIT 33), T1 (SIT 53) and LAM4 (SIT 811) with 7%, 5.1% and 2.8%, respectively. The Beijing family was the only genotype found in all the five districts and was predominant in Mopani (18.8%), Sekhukhune (23.7%) and Vhembe (23.3%). Dominant genotypes in Capricorn and Waterberg were LAM3 (11.9%) and T1 (13.3%), respectively. A wide diversity of lineages was demonstrated at district level. A

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,

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.

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

Preclinical Testing of the Nitroimidazopyran PA-824 for Activity against Mycobacterium tuberculosis in a Series of In Vitro and In Vivo Models

OpenAIRE

Lenaerts, Anne J.; Gruppo, Veronica; Marietta, Karen S.; Johnson, Christine M.; Driscoll, Diane K.; Tompkins, Nicholas M.; Rose, Jerry D.; Reynolds, Robert C.; Orme, Ian M.

2005-01-01

This study extends earlier reports regarding the in vitro and in vivo efficacies of the nitroimidazopyran PA-824 against Mycobacterium tuberculosis. PA-824 was tested in vitro against a broad panel of multidrug-resistant clinical isolates and was found to be highly active against all isolates (MIC < 1 μg/ml). The activity of PA-824 against M. tuberculosis was also assessed grown under conditions of oxygen depletion. PA-824 showed significant activity at 2, 10, and 50 μg/ml, similar to that of...

Interleukin-1 or tumor necrosis factor-alpha augmented the cytotoxic effect of mycobacteria on human fibroblasts: application to evaluation of pathogenesis of clinical isolates of Mycobacterium tuberculosis and M. avium complex.

Science.gov (United States)

Takii, T; Abe, C; Tamura, A; Ramayah, S; Belisle, J T; Brennan, P J; Onozaki, K

2001-03-01

Mycobacteria-induced in vitro events reflecting human tuberculosis can contribute to the evaluation of the pathogenesis of Mycobacterium tuberculosis (MTB). In this study, we propose such an in vitro method based on live mycobacteria-induced cytotoxicity to human cell lines. When human lung-derived normal fibroblast cell line MRC-5 was infected with various strains of mycobacteria (M. tuberculosis H(37)Rv and H(37) Ra, Mycobacterium avium 427S and 2151SmO, and Mycobacterium bovis BCG Pasteur and Tokyo), the fibroblasts were killed by mycobacteria according to the degree of virulence. Other human originated macrophage (U-937, THP-1), myeloid (HL-60), and epithelial carcinoma (A549) cell lines exhibited a similar cytotoxic response to virulent mycobacteria. MRC-5 was most susceptible to virulent mycobacteria among various human cell lines examined. The cytotoxicity was enhanced by the proinflammatory cytokines, interleukin-1 (IL-1) and tumor necrosis factor-a (TNF-alpha), which in the absence of mycobacteria stimulate the growth of normal human fibroblasts. This in vitro evaluation system was applied to clinical isolates of drug-sensitive MTB (DS-MTB), drug-resistant MTB (DR-MTB) including multidrug-resistant (MDR-MTB), and M. avium complex (MAC). MTB strains (n = 24) exhibited strong cytotoxic activity, but MAC strains (n = 5) had only weak activity. Furthermore, there was no significant difference in cytotoxicity between DS-MTB (n = 11) and DR-MTB (n = 13). Collectively, these results suggest that this new in vitro system is useful for evaluating the pathogenesis of mycobacteria and that there was no difference in the pathogenesis between drug-susceptible and drug-resistant clinical isolates.

Design and Construction of a Cloning Vector Containing the hspX Gene of Mycobacterium tuberculosis

Directory of Open Access Journals (Sweden)

Atieh Yaghoubi

2016-10-01

Full Text Available Background: Tuberculosis (TB is a major cause of death worldwide. Finding an effective vaccine against TB is the best way to control it. Several vaccines against this disease have been developed but none are completely protective. The aim of this study was to design and construct a cloning vector containing the Mycobacterium tuberculosis (M. tuberculosis heat shock protein X (hspX. Methods: First, an hspX fragment was amplified by PCR and cloned into plasmid pcDNA3.1(+ and recombinant vector was confirmed. Results: A 435 bp hspX fragment was isolated. The fragment was 100% homologous with hspX of M. tuberculosis strain H37Rv in GenBank. Conclusions: In this study, the cloning vector pcDNA3.1(+, containing a 435-bp hspX fragment of M. tuberculosis, was constructed. This could be used as a DNA vaccine to induce immune responses in animal models in future studies.

Snapshot of the genetic diversity of Mycobacterium tuberculosis isolates in Iraq

Directory of Open Access Journals (Sweden)

Mohanad Mohsin Ahmed

2014-01-01

Conclusion: Iraq is specific in having its own most predominant lineage (SIT1144/T1 which is not found among neighboring countries. The 15-locus MIRU-VNTR can be useful in discriminating M. tuberculosis isolates in Iraq.

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

Directory of Open Access Journals (Sweden)

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

Science.gov (United States)

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.

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

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

Science.gov (United States)

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

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

Directory of Open Access Journals (Sweden)

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.

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...

Targeting phenotypically tolerant Mycobacterium tuberculosis

Science.gov (United States)

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

Beijing/W genotype Mycobacterium tuberculosis and drug resistance.

NARCIS (Netherlands)

Glynn, Judith R; Kremer, Kristin; Borgdorff, Martien W; Rodriguez, Mar Pujades; Soolingen, Dick van

2006-01-01

Beijing/W genotype Mycobacterium tuberculosis is widespread, may be increasing, and may have a predilection for drug resistance. Individual-level data on >29,000 patients from 49 studies in 35 countries were combined to assess the Beijing genotype's prevalence worldwide, trends over time and with

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.

First insights into the genetic diversity of Mycobacterium tuberculosis strains in Salvador, Bahia State, Brazil Primeiro ensaio sobre diversidade genética das cepas de Mycobacterium tuberculosis em Salvador, Bahia, Brasil

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Aída Cristina do Nascimento Silva

2011-09-01

Full Text Available This study constitutes a first attempt to describe the genetic population structure of Mycobacterium tuberculosis circulating in Salvador, Bahia State, Brazil. A total of 56 confirmed cases of pulmonary tuberculosis, identified between March and June 2008, were analyzed using restriction fragment length polymorphism (IS6110-RFLP. The study population was characterized by a predominance of males (71.43% over 30 years of age (68.75%. Forty-one isolates were found to belong to a single pattern (73.2%, while 15 (26.7% were found in group patterns, forming six clusters. The higher level of diversity observed is much more suggestive of endogenous reactivation than recent transmission.Este é o primeiro estudo realizado na Bahia, Brasil, visando à descrição da estrutura da população genética circulante do Mycobacterium tuberculosis na cidade de Salvador. Um total de 56 casos confirmados de tuberculose pulmonar, identificados entre março e junho de 2008, foi analisado pelo método Restriction Fragment Lenght Polymorphism (IS6110-RFLP. A população de estudo foi caracterizada como a maioria do sexo masculino (71,43 %, idade acima de 30 anos (68,75%. Quarenta e um isolados (73,21% com padrão único, enquanto 15 (26,75% apresentaram padrões agrupáveis, formando seis clusters. A alta taxa de diversidade das cepas de M. tuberculosis observada é mais sugestiva de reativação endógena do que transmissão recente.

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.

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.

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

KAUST Repository

Phelan, Jody

2016-03-23

Background Combating the spread of drug resistant tuberculosis is a global health priority. Whole genome association studies are being applied to identify genetic determinants of resistance to anti-tuberculosis drugs. Protein structure and interaction modelling are used to understand the functional effects of putative mutations and provide insight into the molecular mechanisms leading to resistance. Methods To investigate the potential utility of these approaches, we analysed the genomes 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 polymorphisms associated with minimum inhibitory concentrations for first-line anti-tuberculosis drugs. In addition, the effect of identified candidate mutations on protein stability and interactions was assessed quantitatively with well-established computational methods. Results The analysis revealed that mutations in the genes rpoB (rifampicin), katG (isoniazid), inhA-promoter (isoniazid), rpsL (streptomycin) and embB (ethambutol) were responsible for the majority of resistance observed. A subset of the mutations identified in rpoB and katG were predicted to affect protein stability. Further, a strong direct correlation was observed between the minimum inhibitory concentration values and the distance of the mutated residues in the three-dimensional structures of rpoB and katG to their respective drugs binding sites. Conclusions Using the TDR resource, we demonstrate the usefulness of whole genome association and convergent evolution approaches to detect known and potentially novel mutations associated with drug resistance. Further, protein structural modelling could provide a means of predicting the impact of polymorphisms on drug efficacy in the absence of phenotypic data. These approaches could ultimately lead to novel resistance

Performance Assessment of the CapitalBio Mycobacterium Identification Array System for Identification of Mycobacteria

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Liu, Jingbo; Yan, Zihe; Han, Min; Han, Zhijun; Jin, Lingjie; Zhao, Yanlin

2012-01-01

The CapitalBio Mycobacterium identification microarray system is a rapid system for the detection of Mycobacterium tuberculosis. The performance of this system was assessed with 24 reference strains, 486 Mycobacterium tuberculosis clinical isolates, and 40 clinical samples and then compared to the “gold standard” of DNA sequencing. The CapitalBio Mycobacterium identification microarray system showed highly concordant identification results of 100% and 98.4% for Mycobacterium tuberculosis complex (MTC) and nontuberculous mycobacteria (NTM), respectively. The sensitivity and specificity of the CapitalBio Mycobacterium identification array for identification of Mycobacterium tuberculosis isolates were 99.6% and 100%, respectively, for direct detection and identification of clinical samples, and the overall sensitivity was 52.5%. It was 100% for sputum, 16.7% for pleural fluid, and 10% for bronchoalveolar lavage fluid, respectively. The total assay was completed in 6 h, including DNA extraction, PCR, and hybridization. The results of this study confirm the utility of this system for the rapid identification of mycobacteria and suggest that the CapitalBio Mycobacterium identification array is a molecular diagnostic technique with high sensitivity and specificity that has the capacity to quickly identify most mycobacteria. PMID:22090408

Mycobacterium tuberculosis strains potentially involved in the TB epidemic in Sweden a century ago.

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Ramona Groenheit

Full Text Available UNLABELLED: A hundred years ago the prevalence of tuberculosis (TB in Sweden was one of the highest in the world. In this study we conducted a population-based search for distinct strains of Mycobacterium tuberculosis complex isolated from patients born in Sweden before 1945. Many of these isolates represent the M. tuberculosis complex population that fueled the TB epidemic in Sweden during the first half of the 20(th century. METHODS: Genetic relationships between strains that caused the epidemic and present day strains were studied by spoligotyping and restriction fragment length polymorphism. RESULTS: The majority of the isolates from the elderly population were evolutionary recent Principal Genetic Group (PGG2/3 strains (363/409 or 88.8%, and only a low proportion were ancient PGG1 strains (24/409 or 5.9%. Twenty-two were undefined. The isolates demonstrated a population where the Euro-American superlineage dominated; in particular with Haarlem (41.1% and T (37.7% spoligotypes and only 21.2% belonged to other spoligotype families. Isolates from the elderly population clustered much less frequently than did isolates from a young control group population. CONCLUSIONS: A closely knit pool of PGG2/3 strains restricted to Sweden and its immediate neighbours appears to have played a role in the epidemic, while PGG1 strains are usually linked to migrants in todaýs Sweden. Further studies of these outbreak strains may give indications of why the epidemic waned.

19-VNTR loci used in genotyping Chinese clinical Mycobacterium tuberculosis complex strains and in association with spoligotyping.

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Jiang, Yi; Liu, Hai-can; Zheng, Huajun; Dou, Xiangfeng; Tang, Biao; Zhao, Xiu-qin; Zhu, Yongqiang; Lu, Bing; Wang, Shengyue; Dong, Hai-yan; Zhang, Yuan-yuan; Zhao, Guoping; Wan, Kanglin

2013-07-01

Recently, tandem repeat typing has emerged as a rapid and easy method for the molecular epidemiology of the Mycobacterium tuberculosis (M. tuberculosis) complex. In this study, a collection of 19 VNTRs incorporating 15 previously described loci and 4 newly evaluated markers were used to genotype 206 Chinese M. tuberculosis isolates and 9 BCG strains. The discriminatory power was evaluated and compared with that obtained by Spoligotyping. It turned out that 15-locus VNTR could be very useful in M. tuberculosis complex strains genotyping in China. The 4 newly evaluated loci were proved informative and could be useful for future epidemiology studies, especially in Beijing family strains. In addition, a unique pattern of the latter 4 loci were found in Chinese BCG strains. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

One-tube loop-mediated isothermal amplification combined with restriction endonuclease digestion and ELISA for colorimetric detection of resistance to isoniazid, ethambutol and streptomycin in Mycobacterium tuberculosis isolates.

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Lee, Mei-Feng; Chen, Yen-Hsu; Hsu, Hui-Jine; Peng, Chien-Fang

2010-10-01

In this study, we designed a simple and rapid colorimetric detection method, a one-tube loop-mediated isothermal amplification (LAMP)-PCR-hybridization-restriction endonuclease-ELISA [one-tube LAMP-PCR-HY-RE-ELISA] system, to detect resistance to isoniazid, ethambutol and streptomycin in strains of Mycobacterium tuberculosis isolated from clinical specimens. The clinical performance of this method for detecting isoniazid-resistant, ethambutol-resistant and streptomycin-resistant isolates of M. tuberculosis showed 98.9%, 94.3% and 93.8%, respectively. This assay is rapid and convenient that can be performed within one working day. One-tube LAMP-PCR-HY-RE-ELISA system was designed based on hot spot point mutations in target drug-resistant genes, using LAMP-PCR, hybridization, digestion with restriction endonuclease and colorimetric method of ELISA. In this study, LAMP assay was used to amplify DNA from drug-resistant M. tuberculosis, and ELISA was used for colorimetrical determination. This assay will be a useful tool for rapid diagnosis of mutant codons in strains of M. tuberculosis for isoniazid at katG 315 and katG 463, ethambutol at embB 306 and embB 497, and streptomycin at rpsL 43. Crown Copyright © 2010. Published by Elsevier B.V. All rights reserved.

Drug-resistance patterns of Mycobacterium tuberculosis strains and associated risk factors among multi drug-resistant tuberculosis suspected patients from Ethiopia.

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Mesfin, Eyob Abera; Beyene, Dereje; Tesfaye, Abreham; Admasu, Addisu; Addise, Desalegn; Amare, Miskir; Dagne, Biniyam; Yaregal, Zelalem; Tesfaye, Ephrem; Tessema, Belay

2018-01-01

Multidrug drug-resistant tuberculosis (MDR-TB) is a major health problem and seriously threatens TB control and prevention efforts globally. Ethiopia is among the 30th highest TB burden countries for MDR-TB with 14% prevalence among previously treated cases. The focus of this study was on determining drug resistance patterns of Mycobacterium tuberculosis among MDR-TB suspected cases and associated risk factors. A cross-sectional study was conducted in Addis Ababa from June 2015 to December 2016. Sputum samples and socio-demographic data were collected from 358 MDR-TB suspected cases. Samples were analyzed using Ziehl-Neelsen technique, GeneXpert MTB/RIF assay, and culture using Lowenstein-Jensen and Mycobacterial growth indicator tube. Data were analyzed using SPSS version 23. A total of 226 the study participants were culture positive for Mycobacterium tuberculosis, among them, 133 (58.8%) participants were males. Moreover, 162 (71.7%) had been previously treated for tuberculosis, while 128 (56.6%) were TB/HIV co-infected. A majority [122 (54%)] of the isolates were resistant to any first-line anti-TB drugs. Among the resistant isolates, 110 (48.7%) were determined to be resistant to isoniazid, 94 (41.6%) to streptomycin, 89 (39.4%) to rifampicin, 72 (31.9%) to ethambutol, and 70 (30.9%) to pyrazinamide. The prevalence of MDR-TB was 89 (39.4%), of which 52/89 (58.4%) isolates were resistance to all five first-line drugs. Risk factors such as TB/HIV co-infection (AOR = 5.59, p = 0.00), cigarette smoking (AOR = 3.52, p = 0.045), alcohol drinking (AOR = 5.14, p = 0.001) hospital admission (AOR = 3.49, p = 0.005) and visiting (AOR = 3.34, p = 0.044) were significantly associated with MDR-TB. The prevalence of MDR-TB in the study population was of a significantly high level among previously treated patients and age group of 25-34. TB/HIV coinfection, smoking of cigarette, alcohol drinking, hospital admission and health facility visiting were identified as risk factors

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.

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

Science.gov (United States)

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.

Wild-type MIC distributions for aminoglycoside and cyclic polypeptide antibiotics used for treatment of Mycobacterium tuberculosis infections.

Science.gov (United States)

Juréen, P; Angeby, K; Sturegård, E; Chryssanthou, E; Giske, C G; Werngren, J; Nordvall, M; Johansson, A; Kahlmeter, G; Hoffner, S; Schön, T

2010-05-01

The aminoglycosides and cyclic polypeptides are essential drugs in the treatment of multidrug-resistant tuberculosis, underscoring the need for accurate and reproducible drug susceptibility testing (DST). The epidemiological cutoff value (ECOFF) separating wild-type susceptible strains from non-wild-type strains is an important but rarely used tool for indicating susceptibility breakpoints against Mycobacterium tuberculosis. In this study, we established wild-type MIC distributions on Middlebrook 7H10 medium for amikacin, kanamycin, streptomycin, capreomycin, and viomycin using 90 consecutive clinical isolates and 21 resistant strains. Overall, the MIC variation between and within runs did not exceed +/-1 MIC dilution step, and validation of MIC values in Bactec 960 MGIT demonstrated good agreement. Tentative ECOFFs defining the wild type were established for all investigated drugs, including amikacin and viomycin, which currently lack susceptibility breakpoints for 7H10. Five out of seven amikacin- and kanamycin-resistant isolates were classified as susceptible to capreomycin according to the current critical concentration (10 mg/liter) but were non-wild type according to the ECOFF (4 mg/liter), suggesting that the critical concentration may be too high. All amikacin- and kanamycin-resistant isolates were clearly below the ECOFF for viomycin, and two of them were below the ECOFF for streptomycin, indicating that these two drugs may be considered for treatment of amikacin-resistant strains. Pharmacodynamic indices (peak serum concentration [Cmax]/MIC) were more favorable for amikacin and viomycin compared to kanamycin and capreomycin. In conclusion, our data emphasize the importance of establishing wild-type MIC distributions for improving the quality of drug susceptibility testing against Mycobacterium tuberculosis.

Comparison of manual mycobacteria growth indicator tube and epsilometer test with agar proportion method for susceptibility testing of Mycobacterium tuberculosis

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N Karabulut

2014-01-01

Full Text Available Background and Objectives: Antimycobacterial susceptibility tests take weeks, and delayed therapy can lead to spread of Mycobacterium tuberculosis. Therefore, rapid, accurate and cost-effective methods are required for proper therapy selection. In this study, the Mycobacteria growth indicator tube (MGIT and epsilometer test (Etest methods were compared to the agar proportion method for susceptibility testing of Mycobacterium tuberculosis. Materials and Methods: The susceptibility tests against isoniazid (INH, rifampin (RIF, streptomycin (STM and ethambutol (ETM of 51 M. tuberculosis complex isolates were analyzed by the MGIT, Etest and agar proportion methods. Results: The concordance between MGIT/Etest and agar proportion methods was 98% for INH and 100% for RIF, STM, ETM. There were not statistically significant differences in results of the susceptibility tests between MGIT/Etest and the reference agar proportion method. Conclusion: The results have shown that MGIT and Etest methods can be used instead of the agar proportion method, because these two methods are more rapid and easier than the agar proportion method.

Virulence factors of the Mycobacterium tuberculosis complex

Science.gov (United States)

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

Decreasing prevalence of multi-drugs resistant Mycobacterium tuberculosis in Nashik City, India

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Arun P. More

2013-03-01

Full Text Available Objective: In India, increasing prevalence of multi-drug resistant tuberculosis (MDR has aggravated the control oftuberculosis problem. In many urban and semi-urban regions of India, no surveillance data of multidrug resistance inMycobacterium tuberculosisis available.Methods: A surveillance study on multidrug resistance was carried out in semi-urban and rural regions in and aroundNashik City of Maharashtra, India. The surveillance study was conducted in this region found that the prevalence ofcombined resistance to first and second-line anti-tuberculosis drugs is remarkably high. The isolates of M. tuberculosiswas identified and subjected to drug susceptibility testing. The patterns of drug susceptibility of isolates of M. tuberculosisduring the periods 2000 and 2004 were compared with drug susceptibility patterns of the organisms during theperiod 2008 to 2011.Results: The 260 isolates identified as M. tuberculosis show mean drug resistance prevalence of 45.6% for more than anytwo drugs and the MDR rate as 37% in the years 2000 to 2004 whereas 305 isolates of the organism show mean drugresistance prevalence of 30.2% and the MDR rate as 25% in the years 2008 to 2011.Conclusion: The researcher found that, though the prevalence of multidrug resistance to the drugs tested is remarkablyhigh, it has come down noticeably during the past seven years due to efforts of State Government and strict implementationof treatment guidelines of WHO by the physicians. J Microbiol Infect Dis 2013; 3(1: 12-17Key words: MDR-TB, XDR-TB, DOTS, drug-resistance prevalence rate.

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.

Thr202Ala in thyA Is a Marker for the Latin American Mediterranean Lineage of the Mycobacterium tuberculosis Complex Rather than Para-Aminosalicylic Acid Resistance

KAUST Repository

Feuerriegel, S.; Koser, C.; Trube, L.; Archer, John A.C.; Rusch Gerdes, S.; Richter, E.; Niemann, S.

2010-01-01

Single nucleotide polymorphisms (SNPs) involved in the development of resistance represent powerful markers for the rapid detection of first- and second-line resistance in clinical Mycobacterium tuberculosis complex (MTBC) isolates. However

Drug Susceptibility of Mycobacterium tuberculosis Beijing Genotype and Association with MDR TB

Science.gov (United States)

ten Kate, Marian T.; de Knegt, Gerjo J.; Kremer, Kristin; Aarnoutse, Rob E.; Boeree, Martin J.; Verbrugh, Henri A.; van Soolingen, Dick; Bakker-Woudenberg, Irma A.J.M.

2012-01-01

To determine differences in the ability of Mycobacterium tuberculosis strains to withstand antituberculosis drug treatment, we compared the activity of antituberculosis drugs against susceptible Beijing and East-African/Indian genotype M. tuberculosis strains. Beijing genotype strains showed high rates of mutation within a wide range of drug concentrations, possibly explaining this genotype’s association with multidrug-resistant tuberculosis. PMID:22469099

Mycobacterium Tuberculosis Pyomyositis in an Infant

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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

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.

[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.

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...

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.

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

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

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

Science.gov (United States)

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.

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....

Rapid presumptive identification of the Mycobacterium tuberculosis-bovis complex by radiometric determination of heat stable urease

International Nuclear Information System (INIS)

Gandy, J.H.; Pruden, E.L.; Cox, F.R.

1983-01-01

Simple and rapid Bactec methodologies for the determination of neat (unaltered) and heat stable urease activity of mycobacteria are presented. Clinical isolates (63) and stock cultures (32)--consisting of: M. tuberculosis (19), M. bovis (5), M. kansasii (15), M. marinum (4), M. simiae (3), M. scrofulaceum (16), M. gordonae (6), M. szulgai (6), M. flavescens (1), M. gastri (1), M. intracellulare (6), M. fortuitum-chelonei complex (12), and M. smegmatis (1)--were tested for neat urease activity by Bactec radiometry. Mycobacterial isolates (50-100 mg wet weight) were incubated at 35 degrees C for 30 minutes with microCi14C-urea. Urease-positive mycobacteria gave Bactec growth index (GI) values greater than 100 units, whereas urease-negative species gave values less than 10 GI units. Eighty-three isolates possessing neat urease activity were heated at 80 degrees C for 30 minutes followed by incubation at 35 degrees C for 30 minutes with 1 microCi14C-urea. Mycobacterium tuberculosis-bovis complex demonstrated heat-stable urease activity (GI more than 130 units) and could be distinguished from mycobacteria other than tuberculosis (MOTT), which gave GI values equal to or less than 40 units

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.

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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.

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...

Optimization of Standard In-House 24-Locus Variable-Number Tandem-Repeat Typing for Mycobacterium tuberculosis and Its Direct Application to Clinical Material

NARCIS (Netherlands)

de Beer, Jessica L.; Akkerman, Onno W.; Schurch, Anita C.; Mulder, Arnout; van der Werf, Tjip S.; van der Zanden, Adri G. M.; van Ingen, Jakko; van Soolingen, Dick

Variable-number tandem-repeat (VNTR) typing with a panel of 24 loci is the current gold standard in the molecular typing of Mycobacterium tuberculosis complex isolates. However, because of technical problems, a part of the loci often cannot be amplified by multiplex PCRs. Therefore, a considerable

High-resolution minisatellite-based typing as a portable approach to global analysis of Mycobacterium tuberculosis molecular epidemiology

Science.gov (United States)

Mazars, Edith; Lesjean, Sarah; Banuls, Anne-Laure; Gilbert, Michèle; Vincent, Véronique; Gicquel, Brigitte; Tibayrenc, Michel; Locht, Camille; Supply, Philip

2001-01-01

The worldwide threat of tuberculosis to human health emphasizes the need to develop novel approaches to a global epidemiological surveillance. The current standard for Mycobacterium tuberculosis typing based on IS6110 restriction fragment length polymorphism (RFLP) suffers from the difficulty of comparing data between independent laboratories. Here, we propose a high-resolution typing method based on variable number tandem repeats (VNTRs) of genetic elements named mycobacterial interspersed repetitive units (MIRUs) in 12 human minisatellite-like regions of the M. tuberculosis genome. MIRU-VNTR profiles of 72 different M. tuberculosis isolates were established by PCR analysis of all 12 loci. From 2 to 8 MIRU-VNTR alleles were identified in the 12 regions in these strains, which corresponds to a potential of over 16 million different combinations, yielding a resolution power close to that of IS6110-RFLP. All epidemiologically related isolates tested were perfectly clustered by MIRU-VNTR typing, indicating that the stability of these MIRU-VNTRs is adequate to track outbreak episodes. The correlation between genetic relationships inferred from MIRU-VNTR and IS6110-RFLP typing was highly significant. Compared with IS6110-RFLP, high-resolution MIRU-VNTR typing has the considerable advantages of being fast, appropriate for all M. tuberculosis isolates, including strains that have a few IS6110 copies, and permitting easy and rapid comparison of results from independent laboratories. This typing method opens the way to the construction of digital global databases for molecular epidemiology studies of M. tuberculosis. PMID:11172048

Whole genome sequence analysis of Mycobacterium suricattae

KAUST Repository

Dippenaar, Anzaan; Parsons, Sven David Charles; Sampson, Samantha Leigh; Van Der Merwe, Ruben Gerhard; Drewe, Julian Ashley; Abdallah, Abdallah; Siame, Kabengele Keith; Gey Van Pittius, Nicolaas Claudius; Van Helden, Paul David; Pain, Arnab; Warren, Robin Mark

2015-01-01

Tuberculosis occurs in various mammalian hosts and is caused by a range of different lineages of the Mycobacterium tuberculosis complex (MTBC). A recently described member, Mycobacterium suricattae, causes tuberculosis in meerkats (Suricata suricatta) in Southern Africa and preliminary genetic analysis showed this organism to be closely related to an MTBC pathogen of rock hyraxes (Procavia capensis), the dassie bacillus. Here we make use of whole genome sequencing to describe the evolution of the genome of M. suricattae, including known and novel regions of difference, SNPs and IS6110 insertion sites. We used genome-wide phylogenetic analysis to show that M. suricattae clusters with the chimpanzee bacillus, previously isolated from a chimpanzee (Pan troglodytes) in West Africa. We propose an evolutionary scenario for the Mycobacterium africanum lineage 6 complex, showing the evolutionary relationship of M. africanum and chimpanzee bacillus, and the closely related members M. suricattae, dassie bacillus and Mycobacterium mungi.

Whole genome sequence analysis of Mycobacterium suricattae

KAUST Repository

Dippenaar, Anzaan

2015-10-21

Tuberculosis occurs in various mammalian hosts and is caused by a range of different lineages of the Mycobacterium tuberculosis complex (MTBC). A recently described member, Mycobacterium suricattae, causes tuberculosis in meerkats (Suricata suricatta) in Southern Africa and preliminary genetic analysis showed this organism to be closely related to an MTBC pathogen of rock hyraxes (Procavia capensis), the dassie bacillus. Here we make use of whole genome sequencing to describe the evolution of the genome of M. suricattae, including known and novel regions of difference, SNPs and IS6110 insertion sites. We used genome-wide phylogenetic analysis to show that M. suricattae clusters with the chimpanzee bacillus, previously isolated from a chimpanzee (Pan troglodytes) in West Africa. We propose an evolutionary scenario for the Mycobacterium africanum lineage 6 complex, showing the evolutionary relationship of M. africanum and chimpanzee bacillus, and the closely related members M. suricattae, dassie bacillus and Mycobacterium mungi.

Spoligotyping based genetic diversity of Mycobacterium tuberculosis in Ethiopia: a systematic review.

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Tulu, Begna; Ameni, Gobena

2018-03-27

Understanding the types of strains and lineages of Mycobacterium tuberculosis (M. tuberculosis) circulating in a country is of paramount importance for tuberculosis (TB) control program of that country. The main aim of this study was to review and compile the results of studies conducted on strains and lineages of M. tuberculosis in Ethiopia. A systematic search and review of articles published on M. tuberculosis strains and lineages in Ethiopia were made. PubMed and Google Scholar databases were considered for the search while the keywords used were M. tuberculosis, molecular epidemiology, molecular typing spoligotyping and Ethiopia. Twenty-one studies were considered in this review and a total of 3071 M. tuberculosis isolates and 3067 strains were included. These studies used spoligotyping and identified five lineages including Indo-Ocean, East Asian/Beijing, East African-Indian, Euro-American and Ethiopian in a proportion of 7.1%, 0.2%, 23.0%, 64.8%, and 4.1%, respectively. Thus, Euro-American was the most frequently (64.8%) occurring Lineage while East Asian was the least (0.2%) frequently occurring Lineage in the country. Surprisingly, the Ethiopian Lineage seemed to be localized to northeastern Ethiopia. In addition, the top five clades identified by this review were T, CAS, H, Manu and Ethiopian comprising of 48.0%, 23.0%, 11.0%, 6.0% and 4.1% of the strains, respectively. Furthermore, predominant shared types (spoligotype patterns) identified were SIT149, SIT53, SIT25, SIT37, and SIT21, each consisting of 420, 343, 266, 162 and 102 isolates, respectively, while, on the other hand, 15% of the strains were orphan. According to the summary of the results of this review, diversified strains and lineages of M. tuberculosis were found in Ethiopia, and the frequencies of occurrence of these strains and lineages were variable in different regions of the country. This systematic review is registered in the PRISMA with the registration number of 42017059263.

Predicting Mycobacterium tuberculosis Complex Clades Using Knowledge-Based Bayesian Networks

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Minoo Aminian

2014-01-01

Full Text Available We develop a novel approach for incorporating expert rules into Bayesian networks for classification of Mycobacterium tuberculosis complex (MTBC clades. The proposed knowledge-based Bayesian network (KBBN treats sets of expert rules as prior distributions on the classes. Unlike prior knowledge-based support vector machine approaches which require rules expressed as polyhedral sets, KBBN directly incorporates the rules without any modification. KBBN uses data to refine rule-based classifiers when the rule set is incomplete or ambiguous. We develop a predictive KBBN model for 69 MTBC clades found in the SITVIT international collection. We validate the approach using two testbeds that model knowledge of the MTBC obtained from two different experts and large DNA fingerprint databases to predict MTBC genetic clades and sublineages. These models represent strains of MTBC using high-throughput biomarkers called spacer oligonucleotide types (spoligotypes, since these are routinely gathered from MTBC isolates of tuberculosis (TB patients. Results show that incorporating rules into problems can drastically increase classification accuracy if data alone are insufficient. The SITVIT KBBN is publicly available for use on the World Wide Web.

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.

Wild-Type MIC Distributions for Aminoglycoside and Cyclic Polypeptide Antibiotics Used for Treatment of Mycobacterium tuberculosis Infections▿

Science.gov (United States)

Juréen, P.; Ängeby, K.; Sturegård, E.; Chryssanthou, E.; Giske, C. G.; Werngren, J.; Nordvall, M.; Johansson, A.; Kahlmeter, G.; Hoffner, S.; Schön, T.

2010-01-01

The aminoglycosides and cyclic polypeptides are essential drugs in the treatment of multidrug-resistant tuberculosis, underscoring the need for accurate and reproducible drug susceptibility testing (DST). The epidemiological cutoff value (ECOFF) separating wild-type susceptible strains from non-wild-type strains is an important but rarely used tool for indicating susceptibility breakpoints against Mycobacterium tuberculosis. In this study, we established wild-type MIC distributions on Middlebrook 7H10 medium for amikacin, kanamycin, streptomycin, capreomycin, and viomycin using 90 consecutive clinical isolates and 21 resistant strains. Overall, the MIC variation between and within runs did not exceed ±1 MIC dilution step, and validation of MIC values in Bactec 960 MGIT demonstrated good agreement. Tentative ECOFFs defining the wild type were established for all investigated drugs, including amikacin and viomycin, which currently lack susceptibility breakpoints for 7H10. Five out of seven amikacin- and kanamycin-resistant isolates were classified as susceptible to capreomycin according to the current critical concentration (10 mg/liter) but were non-wild type according to the ECOFF (4 mg/liter), suggesting that the critical concentration may be too high. All amikacin- and kanamycin-resistant isolates were clearly below the ECOFF for viomycin, and two of them were below the ECOFF for streptomycin, indicating that these two drugs may be considered for treatment of amikacin-resistant strains. Pharmacodynamic indices (peak serum concentration [Cmax]/MIC) were more favorable for amikacin and viomycin compared to kanamycin and capreomycin. In conclusion, our data emphasize the importance of establishing wild-type MIC distributions for improving the quality of drug susceptibility testing against Mycobacterium tuberculosis. PMID:20237102

Molecular characterization of Mycobacterium tuberculosis isolates from Tanga, Tanzania: First insight of MIRU-VNTR and microarray-based spoligotyping in a high burden country.

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Hoza, Abubakar S; Mfinanga, Sayoki G; Moser, Irmgard; König, Brigitte

2016-05-01

Molecular typing of Mycobacterium tuberculosis(MTB) has greatly enhanced the understanding of the population structure of MTB isolates and epidemiology of tuberculosis (TB). To characterize prevalent genotypes of MTB, microarrays‑based spoligotyping and mycobacterial interspersed repetitive unit‑variable number of tandem repeats (MIRU‑VNTR) were applied on 80 isolates collected from primary health care facilities in Tanga, North‑eastern Tanzania. A total of 18 distinct spoligotypes were identified. The lineages by order of their predominance were EAI and CAS families (26.25%, 21 isolates each), LAM family and T super‑family (10%, 8 isolates each), MANU family (3.75%, 3 isolates), Beijing family (2.5%, 2 isolates) and S family (1.25%, 1 isolate). Overall, sixteen (20%) strains could not be allocated to any lineage according to the SITVIT_WEB database. The allelic diversity (h) for specific MIRU‑VNTR loci showed a considerable variation ranging from 0.826 of VNTR locus 3192 to 0.141 of VNTR locus 2059. The allelic diversity for 11 loci (VNTR 3192, 2996, 2165, 960, 4052, 424, 4156, 2531, 1644, 802 and 3690) exceeded 0.6, indicating highly discriminatory power. Seven loci (VNTR 2163b, 2401, 1955, 577, 4348, 2687 and 580) showed moderate discrimination (0.3 ≤ h ≥ 0.6), and three loci (VNTR3007, 154 and 2059) were less polymorphic. The present study suggests that the TB cases in Tanga might be caused by a diverse array of MTB strain families that may be indicative of a cosmopolitan population with frequent migration and travel. Microarray‑based spoligotyping and MIRU‑VNTR could be reliable tools in detecting different MTB genotypes in high burden settings. Copyright © 2016 Elsevier Ltd. All rights reserved.

Draft genome sequence of Mycobacterium tuberculosis strain B9741 of Beijing B0/W lineage from HIV positive patient from Siberia

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K.V. Shur

2016-12-01

Full Text Available We report a draft genome sequence of Mycobacterium tuberculosis strain B9741 belonging to Beijing B0/W lineage isolated from a HIV patient from Siberia, Russia. This clinical isolate showed MDR phenotype and resistance to isoniazid, rifampin, streptomycin and pyrazinamide. We analyzed SNPs associated with virulence and resistance. The draft genome sequence and annotation have been deposited at GenBank under the accession NZ_LVJJ00000000.

Development of a highly sensitive one-tube nested real-time PCR for detecting Mycobacterium tuberculosis.

Science.gov (United States)

Choi, Yeonim; Jeon, Bo-Young; Shim, Tae Sun; Jin, Hyunwoo; Cho, Sang-Nae; Lee, Hyeyoung

2014-12-01

Rapid, accurate detection of Mycobacterium tuberculosis is crucial in the diagnosis of tuberculosis (TB), but conventional diagnostic methods have limited sensitivity and specificity or are time consuming. A new highly sensitive nucleic acid amplification test, combined nested and real-time polymerase chain reaction (PCR) in a single tube (one-tube nested real-time PCR), was developed for detecting M. tuberculosis, which takes advantage of two PCR techniques, i.e., nested PCR and real-time PCR. One-tube nested real-time PCR was designed to have two sequential reactions with two sets of primers and dual probes for the insertion sequence (IS) 6110 sequence of M. tuberculosis in a single closed tube. The minimum limits of detection of IS6110 real-time PCR and IS6110 one-tube nested real-time PCR were 100 fg/μL and 1 fg/μL of M. tuberculosis DNA, respectively. AdvanSure TB/non-tuberculous mycobacteria (NTM) real-time PCR, IS6110 real-time PCR, and two-tube nested real-time PCR showed 100% sensitivity and 100% specificity for clinical M. tuberculosis isolates and NTM isolates. In comparison, the sensitivities of AdvanSure TB/NTM real-time PCR, single IS6110 real-time PCR, and one-tube nested real-time PCR were 91% (152/167), 94.6% (158/167), and 100% (167/167) for sputum specimens, respectively. In conclusion, IS6110 one-tube nested real-time PCR is useful for detecting M. tuberculosis due to its high sensitivity and simple manipulation. Copyright © 2014 Elsevier Inc. All rights reserved.

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.

Mycobacterium malmesburyense sp. nov., a non-tuberculous species of the genus Mycobacterium revealed by multiple gene sequence characterization.

Science.gov (United States)

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

2017-04-01

Non-tuberculous mycobacteria (NTM) are ubiquitous in the environment, and an increasing number of NTM species have been isolated and characterized from both humans and animals, highlighting the zoonotic potential of these bacteria. Host exposure to NTM may impact on cross-reactive immune responsiveness, which may affect diagnosis of bovine tuberculosis and may also play a role in the variability of the efficacy of Mycobacterium bovis BCG vaccination against tuberculosis. In this study we characterized 10 NTM isolates originating from water, soil, nasal swabs of cattle and African buffalo as well as bovine tissue samples. These isolates were previously identified during an NTM survey and were all found, using 16S rRNA gene sequence analysis to be closely related to Mycobacterium moriokaense. A polyphasic approach that included phenotypic characterization, antibiotic susceptibility profiling, mycolic acid profiling and phylogenetic analysis of four gene loci, 16S rRNA, hsp65, sodA and rpoB, was employed to characterize these isolates. Sequence data analysis of the four gene loci revealed that these isolates belong to a unique species of the genus Mycobacterium. This evidence was further supported by several differences in phenotypic characteristics between the isolates and the closely related species. We propose the name Mycobacterium malmesburyense sp. nov. for this novel species. The type strain is WCM 7299T (=ATCC BAA-2759T=CIP 110822T).

[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.

Insertion sequence typing of Mycobacterium tuberculosis: characterization of a widespread subtype with a single copy of IS6110.

Science.gov (United States)

Fomukong, N G; Tang, T H; al-Maamary, S; Ibrahim, W A; Ramayah, S; Yates, M; Zainuddin, Z F; Dale, J W

1994-12-01

DNA fingerprinting with the insertion sequence IS6110 (also known as IS986) has become established as a major tool for investigating the spread of tuberculosis. Most strains of Mycobacterium tuberculosis have multiple copies of IS6110, but a small minority carry a single copy only. We have examined selected strains from Malaysia, Tanzania and Oman, in comparison with M. bovis isolates and BCG strains carrying one or two copies of IS6110. The insertion sequence appears to be present in the same position in all these strains, which suggests that in these organisms the element is defective in transposition and that the loss of transposability may have occurred at an early stage in the evolution of the M. tuberculosis complex.

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

Directory of Open Access Journals (Sweden)

Joab Trajano Silva

2008-12-01

, up to species level, is time consuming and difficult. In this work, the objective was to standardize a polymerase chain reaction followed by an enzyme restriction analysis in order to identify the M. tuberculosis complex in milk, without a microbiological isolation step. Reference strains and raw milk seeded with M. Bovis, were used as the starting material.  A 441pb fragment of the hsp65 gene was amplified and digested by two restriction enzymes BstEII and HaeIII. The obtained profile was used to identify the M. tuberculosis complex in milk. The minimum limit of detection of M. bovis in milk was 10CFU/mL. PRA methodology proved to be a specific and sensible method. It can be used to assist the microbiological and biochemical methods commonly used to identifying the bacilli in clinical samples, as milk 

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

Efficient Differentiation of Mycobacterium tuberculosis Strains of the W-Beijing Family from Russia using Highly Polymorphic VNTR Loci

International Nuclear Information System (INIS)

Surikova, O. V.; Voitech, D. S.; Kuzmicheva, G.; Tatkov, S. I.; Mokrousov, I. V.; Narvskaya, O. V.; Rot, M. A.; Soolingen, D. van; Filipenko, M. L.

2005-01-01

The W-Beijing family is a widespread Mycobacterium tuberculosis clonal lineage that frequently causes epidemic outbreaks. This family is genetically homogeneous and conserved, so ETR-VNTR (exact tandem repeat-variable number of tandem repeats) typing is insufficient for strain differentiation, due to a common ETR-A to E profile (42435). This leads to the false clustering in molecular epidemiological studies, especially in the regions of predominance of the W-Beijing family. In this study, we searched for VNTR loci with a high evolutionary rate of polymorphism in the W-Beijing genome. Here we further evaluated VNTR typing on a set of 99 Mycobacterium tuberculosis clinical isolates and reference strains. These isolates were characterized and classified into several genotype families based on three ETR loci (A, C, E) and eight additional loci [previously described as QUB (Queen's University Belfast) or MIRU (Mycobacterial Interspersed Repetitive Units) or Mtubs]. Ninety-nine strains were divided into 74 VNTR-types, 51 isolates of the W-Beijing family identified by IS6110 RFLP-typing (the restriction fragment length polymorphism-typing) and/or spoligotyping were subdivided into 30 VNTR-types. HGDI (the Hunter-Gaston discriminatory index) for all studied loci was close to that of IS6110 RFLP typing, a 'gold standard' method for subtyping M. tuberculosis complex strains. The QUB 26 and QUB 18 loci located in the PPE genes were highly polymorphic and more discriminative than other loci (HGDI is 0.8). Statistically significant increase of tandem repeats number in loci ETR-A, -E, QUB 26, QUB 18, QUB 11B, Mtub21 was revealed in the W-Beijing group compared to genetically divergent non-W-Beijing strains. Thirty-six isolates were subjected to IS6110 RFLP typing. The congruence between results of the IS6110 RFLP typing and 11-loci VNTR typing was estimated on 23 isolates of the W-Beijing family. These isolates were subdivided into 9 IS6110-RFLP types and 13 VNTR types. The poor

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.

Genetic diversity of the Mycobacterium tuberculosis East African–Indian family in three tropical Asian countries

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Yih-Yuan Chen

2017-12-01

Full Text Available Background: The Beijing lineage of Mycobacterium tuberculosis (MTB is the most predominant MTB strain in Asian countries and is spreading worldwide, however, the East African–Indian (EAI lineage is also particularly prevalent in many tropical Asian countries. The evolutionary relationships among MTB EAI isolates from Taiwan and those of tropical Asian countries remain unknown. Methods: The EAI strains collected from patients in Taiwan were analyzed using spacer oligonucleotide typing and mycobacterial interspersed repetitive unit–variable number of tandem repeats (MIRU-VNTR typing, and compared with published profiles from Cambodia and Singapore to investigate potential epidemiological linkages. Results: Among the three countries, the EAI lineage was most prevalent in Cambodia (60%; Singapore, 25.62%; and Taiwan, 21.85%, having also the highest rates of multidrug resistance and lowest rates of clustering of MTB isolates. We describe a convenient method using seven selected MIRU-VNTR loci for first-line typing to discriminate Beijing and EAI lineages. A potential epidemiological linkage in these tropical Asian countries is also discussed based on a minimum-spanning tree constructed using 24 MIRU-VNTR loci of MTB EAI strains. Conclusion: This study identified evolutionary relationships among MTB EAI isolates from Taiwan and those of two other tropical Asian countries, Cambodia and Singapore. Keywords: East African–Indian family, Mycobacterium tuberculosis, tropical Asian countries

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

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.

[DNA mutations associated to rifampicin or isoniazid resistance in M. tuberculosis clinical isolates from Sonora, Mexico].

Science.gov (United States)

Bolado-Martínez, Enrique; Pérez-Mendoza, Ansix; Alegría-Morquecho, Francisco Monserrat; Candia-Plata, María del Carmen; Aguayo-Verdugo, María del Rosario; Alvarez-Hernández, Gerardo

2012-01-01

To perform the analysis of specific regions of the major genes associated with resistance to isoniazid or rifampin. Twenty two M. tuberculosis strains, isolated from human samples obtained in Sonora, Mexico. Specific primers for hotspots of the rpoB, katG, inhA genes and the ahpC-oxyR intergenic region were used. The purified PCR products were sequenced. Mutations in the promoter of inhA, the ahpC-oxyR region, and codon 315 of katG and in 451 or 456 codons of rpoB, were identified. Detection of mutations not previously reported requires further genotypic analysis of Mycobacterium tuberculosis isolates in Sonora.

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

[Description of Mycobacterium tuberculosis mutations conferring resistance to rifampicin and isoniazid detected by GenoType® MTBDRplus V.2 in Colombia].

Science.gov (United States)

Llerena, Claudia; Medina, Raquel

2017-01-24

The GenoType®MTBDRplusV.2 assay is a molecular technique endorsed by the World Health Organization and the Pan American Health Organization that allows for the identification of the Mycobacterium tuberculosis complex and the detection of mutations in the rpoβ gene for rifampicin resistance, and katG and inhA genes for isoniazid resistance. Due to the genetic variability in the circulating strains around the world, the national tuberculosis control programs should assess the performance of these new diagnostic technologies and their use under program conditions as rapid tests. To describe the mutations identified by the GenoType®MTBDRplusV.2 assay in pulmonary samples and Mycobacterium tuberculosis isolates in the Laboratorio Nacional de Referencia of the Instituto Nacional de Salud in 2014. We conducted a retrospective, descriptive study to detect the expression of inhA, KatG and rpoβ genes, responsible for resistence against isoniazid and rifampicin using the GenoType® MTBDRplus V.2 assay in 837 samples and isolates from tuberculosis cases. Several mutations in the rpoβ gene were identified. Ser531Leu was the most frequent (36.6%) followed by Asp516Val (21.6%), while Ser315Thr1 was the most frequent mutation in the katG gene (91.9%). We were able to identify different mutations present in MDR-TB strains in the country, with frequencies similar to those reported in other countries in the South American region.

Mycobacterium grossiae sp. nov., a rapidly growing, scotochromogenic species isolated from human clinical respiratory and blood culture specimens.

Science.gov (United States)

Paniz-Mondolfi, Alberto Enrique; Greninger, Alexander L; Ladutko, Lynn; Brown-Elliott, Barbara A; Vasireddy, Ravikiran; Jakubiec, Wesley; Vasireddy, Sruthi; Wallace, Richard J; Simmon, Keith E; Dunn, Bruce E; Jackoway, Gary; Vora, Surabhi B; Quinn, Kevin K; Qin, Xuan; Campbell, Sheldon

2017-11-01

A previously undescribed, rapidly growing, scotochromogenic species of the genus Mycobacterium (represented by strains PB739 T and GK) was isolated from two clinical sources - the sputum of a 76-year-old patient with severe chronic obstructive pulmonary disease, history of tuberculosis exposure and Mycobacterium avium complex isolated years prior; and the blood of a 15-year-old male with B-cell acute lymphoblastic leukaemia status post bone marrow transplant. The isolates grew as dark orange colonies at 25-37 °C after 5 days, sharing features in common with other closely related species. Analysis of the complete 16S rRNA gene sequence (1492 bp) of strain PB739 T demonstrated that the isolate shared 98.8 % relatedness with Mycobacterium wolinskyi. Partial 429 bp hsp65 and 744 bp rpoB region V sequence analyses revealed that the sequences of the novel isolate shared 94.8 and 92.1 % similarity with those of Mycobacterium neoaurum and Mycobacterium aurum, respectively. Biochemical profiling, antimicrobial susceptibility testing, HPLC/gas-liquid chromatography analyses and multilocus sequence typing support the taxonomic status of these isolates (PB739 T and GK) as representatives of a novel species. Both isolates were susceptible to the Clinical and Laboratory Standards Institute recommended antimicrobials for susceptibility testing of rapidly growing mycobacteria including amikacin, ciprofloxacin, moxifloxacin, doxycycline/minocycline, imipenem, linezolid, clarithromycin and trimethropin/sulfamethoxazole. Both isolates PB739 T and GK showed intermediate susceptibility to cefoxitin. We propose the name Mycobacterium grossiae sp. nov. for this novel species and have deposited the type strain in the DSMZ and CIP culture collections. The type strain is PB739 T (=DSM 104744 T =CIP 111318 T ).

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...

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.

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.

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.

Attempts to revive Mycobacterium tuberculosis from 300-year-old human mummies.

Science.gov (United States)

Lemma, Eshetu; Zimhony, Oren; Greenblatt, Charles L; Koltunov, Viktoria; Zylber, María Inés; Vernon, Kim; Spigelman, Mark

2008-06-01

Environmental persistence of Mycobacterium tuberculosis is subject to speculation. However, the reality that infected postmortem tissues can be a danger to pathologists and embalmers has worrisome implications. A few experimental studies have demonstrated the organism's ability to withstand exposure to embalming fluid and formalin. Recently, a failure was reported in an attempt to resuscitate an original isolate of Robert Koch to determine the lifetime of the tubercle bacillus. The present study also considers a historical approach to determine persistence under favorable environmental conditions. It asks whether acid-fast forms observed in tissues of 300-year-old Hungarian mummies can be resuscitated. Finding organisms before the advent of antibiotics and pasteurization may yield valuable genetic information. Using various media modifications, as well as guinea pig inoculation, an attempt was made to culture these tissues for M. tuberculosis. In addition, a resuscitation-promoting factor, known to increase colony counts in high G+C bacteria, was applied to the cultures. Although an occasional PCR-positive sample was detected, no colonies of M. tuberculosis were obtained. Our results may indicate that the life span of the tubercle bacillus is less than a few hundred years, even though in the short run it can survive harsh chemical treatment.

Mycobacterium tuberculosis population structure and molecular epidemiological analysis in Sucre municipality, Miranda state, Venezuela.

Science.gov (United States)

Patiño, Margareth A; Abadía, Edgar; Solalba Gómez; Maes, Mailis; Muñoz, Mariana; Gómez, Daniela; Guzmán, Patricia; Méndez, María Victoria; Ramirez, Carmen; Mercedes, España; de Waard, Jacobus; Takiff, Howard

2014-12-01

Sucre municipality is a large, densely populated marginal area in the eastern part of Caracas, Venezuela that consistently has more cases of tuberculosis than other municipalities in the country. To identify the neighborhoods in the municipality with the highest prevalence of tuberculosis, and determine whether the Mycobacterium tuberculosis strain distribution in this municipality is different from that previously found in the western part of Caracas and the rest of Venezuela, we collected data on all tuberculosis cases in the municipality diagnosed in 2005-6. We performed two separate molecular epidemiological studies, spoligotyping 44 strains in a first study, and spoligotyping 131 strains, followed by MIRU-VNTR 15 on 21 clustered isolates in the second. With spoligotyping, the most common patterns were Shared International Type SIT17 (21%); SIT42 (15%); SIT93 (11%); SIT20 (7%); SIT53 (6%), a distribution similar to other parts of Venezuela, except that SIT42 and SIT20 were more common. MIRU-VNTR 15 showed that six of seven SIT17 strains examined belonged to a large cluster previously found circulating in Venezuela, but all of the SIT42 strains were related to a cluster centered in the neighborhoods of Unión and Maca, with a MIRU-VNTR pattern not previously seen in Venezuela. It appears that a large percentage of the tuberculosis in the Sucre municipality is caused by the active transmission of two strain families centered within distinct neighborhoods, one reflecting communication with the rest of the country, and the other suggesting the insular, isolated nature of some sectors.

Cloning and expression of pab gene of M. tuberculosis isolated from pulmonary TB patient in E.coli DH5α

Directory of Open Access Journals (Sweden)

Tri Y. M. Raras

2011-11-01

Full Text Available Background: Mycobacterium tuberculosis antigen38 is a potent serodiagnostic agent containing two M. tuberculosisspecific B-cell epitopes. The high price of imported diagnostic agents hinders realization of fast clinical TB diagnosis in developing countries. Therefore, we produced recombinant antigen38 (recAg38M from M. tuberculosis local strain, which might be used to produce economical tuberculosis serodiagnostic kit.Methods: Pab gene that was isolated from pulmonary TB patient in Malang was cloned into a plasmid vector (pGEMTeasy to construct pMB38. The E.coli DH5α clone carrying pMb38 was selected on X-gal medium. The expression of pab was mediated using pPRoExHTc under the control of Trc promoter and E.coli DH5α as host.Results: Alignment of the pab sequence from the white E.coli DH5α clones with that of M. tuberculosis H37Rv showed 98% homology. The recombinant protein in which the signal peptide has been deleted to prevent the protein being secreted into medium was found in the cytoplasm.Conclusion: pab gene of M. tuberculosis isolated from a TB patient could be expressed in heterologous system in E.coliDH5α. (Med J Indones 2011; 20:247-54Keywords: Mycobacterium tuberculosis, Pab gene expression, recombinant antigen38

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

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.

Identification and characterization of a 29-kilodalton protein from Mycobacterium tuberculosis culture filtrate recognized by mouse memory effector cells

DEFF Research Database (Denmark)

Rosenkrands, I; Rasmussen, P.B.; Carnio, M

1998-01-01

Culture filtrate proteins from Mycobacterium tuberculosis induce protective immunity in various animal models of tuberculosis. Two molecular mass regions (6 to 10 kDa and 24 to 36 kDa) of short-term culture filtrate are preferentially recognized by Th1 cells in animal models as well as by patients...... the antigen 85 complex was selected. The 29-kDa antigen (CFP29) was purified from M. tuberculosis short-term culture filtrate by thiophilic adsorption chromatography, anion-exchange chromatography, and gel filtration, In its native form, CFP29 forms a polymer with a high molecular mass. CFP29 was mapped......, and they both elicited the release of high levels of gamma interferon from mouse memory effector cells isolated during the recall of protective immunity to tuberculosis. Interspecies analysis by immunoblotting and PCR demonstrated that CFP29 is widely distributed in mycobacterial species....

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

KAUST Repository

Phelan, Jody

2016-01-01

Abstract Background Combating the spread of drug resistant tuberculosis is a global health priority. Whole genome association studies are being applied to identify genetic determinants of resistance to anti-tuberculosis drugs. Protein structure and interaction modelling are used to understand the functional effects of putative mutations and provide insight into the molecular mechanisms leading to resistance. Methods To investigate the potential utility of these approaches, we analysed the genomes 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 polymorphisms associated with minimum inhibitory concentrations for first-line anti-tuberculosis drugs. In addition, the effect of identified candidate mutations on protein stability and interactions was assessed quantitatively with well-established computational methods. Results The analysis revealed that mutations in the genes rpoB (rifampicin), katG (isoniazid), inhA-promoter (isoniazid), rpsL (streptomycin) and embB (ethambutol) were responsible for the majority of resistance observed. A subset of the mutations identified in rpoB and katG were predicted to affect protein stability. Further, a strong direct correlation was observed between the minimum inhibitory concentration values and the distance of the mutated residues in the three-dimensional structures of rpoB and katG to their respective drugs binding sites. Conclusions Using the TDR resource, we demonstrate the usefulness of whole genome association and convergent evolution approaches to detect known and potentially novel mutations associated with drug resistance. Further, protein structural modelling could provide a means of predicting the impact of polymorphisms on drug efficacy in the absence of phenotypic data. These approaches could ultimately lead to novel

Mutation of katG in a clinical isolate of Mycobacterium tuberculosis: effects on catalase-peroxidase for isoniazid activation.

Science.gov (United States)

Purkan; Ihsanawati; Natalia, D; Syah, Y M; Retnoningrum, D S; Kusuma, H S

2016-01-01

Mutations in katG gene are often associated with isoniazid (INH) resistance in Mycobacterium tuberculosis strain. This research was perfomed to identify the katG mutation in clinical isolate (L8) that is resistant to INH at 1 μg/ml. In addition to characterize the catalase-peroxidase of KatG L8 and perform the ab initio structural study of the protein to get a more complete understanding in drug activation and the resistan­ce mechanism. The katG gene was cloned and expressed in Escherichia coli, then followed by characterization of catalase-peroxidase of KatG. The structure modelling was performed to know a basis of alterations in enzyme activity. A substitution of A713G that correspond to Asn238Ser replacement was found in the L8 katG. The Asn238Ser modification leads to a decline in the activity of catalase-peroxidase and INH oxidation of the L8 KatG protein. The catalytic efficiency (Kcat/KM) of mutant KatGAsn238Ser respectively decreases to 41 and 52% for catalase and peroxidase. The mutant KatGAsn238Ser also shows a decrease of 62% in INH oxidation if compared to a wild type KatG (KatGwt). The mutant Asn238Ser might cause instability in the substrate binding­ site of KatG, because of removal of a salt bridge connecting the amine group of Asn238 to the carbo­xyl group of Glu233, which presents in KatGwt. The lost of the salt bridge in the substrate binding site in mutant KatGAsn238Ser created changes unfavorable for enzyme activities, which in turn emerge as INH resistan­ce in the L8 isolate of M. tuberculosis.

Whole genome sequencing of the monomorphic pathogen Mycobacterium bovis reveals local differentiation of cattle clinical isolates.

Science.gov (United States)

Lasserre, Moira; Fresia, Pablo; Greif, Gonzalo; Iraola, Gregorio; Castro-Ramos, Miguel; Juambeltz, Arturo; Nuñez, Álvaro; Naya, Hugo; Robello, Carlos; Berná, Luisa

2018-01-02

Bovine tuberculosis (bTB) poses serious risks to animal welfare and economy, as well as to public health as a zoonosis. Its etiological agent, Mycobacterium bovis, belongs to the Mycobacterium tuberculosis complex (MTBC), a group of genetically monomorphic organisms featured by a remarkably high overall nucleotide identity (99.9%). Indeed, this characteristic is of major concern for correct typing and determination of strain-specific traits based on sequence diversity. Due to its historical economic dependence on cattle production, Uruguay is deeply affected by the prevailing incidence of Mycobacterium bovis. With the world's highest number of cattle per human, and its intensive cattle production, Uruguay represents a particularly suited setting to evaluate genomic variability among isolates, and the diversity traits associated to this pathogen. We compared 186 genomes from MTBC strains isolated worldwide, and found a highly structured population in M. bovis. The analysis of 23 new M. bovis genomes, belonging to strains isolated in Uruguay evidenced three groups present in the country. Despite presenting an expected highly conserved genomic structure and sequence, these strains segregate into a clustered manner within the worldwide phylogeny. Analysis of the non-pe/ppe differential areas against a reference genome defined four main sources of variability, namely: regions of difference (RD), variable genes, duplications and novel genes. RDs and variant analysis segregated the strains into clusters that are concordant with their spoligotype identities. Due to its high homoplasy rate, spoligotyping failed to reflect the true genomic diversity among worldwide representative strains, however, it remains a good indicator for closely related populations. This study introduces a comprehensive population structure analysis of worldwide M. bovis isolates. The incorporation and analysis of 23 novel Uruguayan M. bovis genomes, sheds light onto the genomic diversity of this

Infection caused by Mycobacterium tuberculosis.

Science.gov (United States)

Peloquin, C A; Berning, S E

1994-01-01

To update readers on the clinical management of infections caused by Mycobacterium tuberculosis, to provide a general description of the organism, culture and susceptibility testing, and clinical manifestations of the disease, and to provide several aspects of the treatment of the disease, including historical perspective, current approaches, and research opportunities for the future. The current medical literature, including abstracts presented at recent international meetings, is reviewed. References were identified through MEDLINE, MEDLARS II, Current Contents, and published meeting abstracts. Data regarding the epidemiology, clinical manifestations, culture and susceptibility testing, and treatment of tuberculosis are cited. Specific attention has been focused on the clinical management of patients with noncontagious infection and potentially contagious active disease (TB) caused by M. tuberculosis. Information contributing to the discussion of the topics selected by the authors is reviewed. Data supporting and disputing specific conclusions are presented. The incidence of TB is increasing in the US, despite the fact that available technologies are capable of controlling the vast majority of existing cases. Fueling the fire is the problem of coinfection with HIV and M. tuberculosis. Very few drugs are available for the treatment of TB, and few of these approach the potency of isoniazid and rifampin. Preventive therapy of patients exposed to multiple-drug-resistant M. tuberculosis (MDR-TB) is controversial and of unknown efficacy. Treatment of active disease caused by MDR-TB requires up to four times longer, is associated with increased toxicity, and is far less successful than the treatment of drug-susceptible TB. Strategies for the management of such cases are presented. The rising incidence of TB in the US reflects a breakdown in the healthcare systems responsible for controlling the disease, which reflects the past budgetary reductions. Although TB control

Metabolic principles of persistence and pathogenicity in Mycobacterium tuberculosis.

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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.

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

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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.

Whole Genome Sequencing Based Characterization of Extensively Drug-Resistant Mycobacterium tuberculosis Isolates from Pakistan

KAUST Repository

Ali, Asho; Hasan, Zahra; McNerney, Ruth; Mallard, Kim; Hill-Cawthorne, Grant A.; Coll, Francesc; Nair, Mridul; Pain, Arnab; Clark, Taane G.; Hasan, Rumina

2015-01-01

Improved molecular diagnostic methods for detection drug resistance in Mycobacterium tuberculosis (MTB) strains are required. Resistance to first- and second- line anti-tuberculous drugs has been associated with single nucleotide polymorphisms (SNPs) in particular genes. However, these SNPs can vary between MTB lineages therefore local data is required to describe different strain populations. We used whole genome sequencing (WGS) to characterize 37 extensively drug-resistant (XDR) MTB isolates from Pakistan and investigated 40 genes associated with drug resistance. Rifampicin resistance was attributable to SNPs in the rpoB hot-spot region. Isoniazid resistance was most commonly associated with the katG codon 315 (92%) mutation followed by inhA S94A (8%) however, one strain did not have SNPs in katG, inhA or oxyR-ahpC. All strains were pyrazimamide resistant but only 43% had pncA SNPs. Ethambutol resistant strains predominantly had embB codon 306 (62%) mutations, but additional SNPs at embB codons 406, 378 and 328 were also present. Fluoroquinolone resistance was associated with gyrA 91-94 codons in 81% of strains; four strains had only gyr B mutations, while others did not have SNPs in either gyrA or gyrB. Streptomycin resistant strains had mutations in ribosomal RNA genes; rpsL codon 43 (42%); rrs 500 region (16%), and gidB (34%) while six strains did not have mutations in any of these genes. Amikacin/kanamycin/capreomycin resistance was associated with SNPs in rrs at nt1401 (78%) and nt1484 (3%), except in seven (19%) strains. We estimate that if only the common hot-spot region targets of current commercial assays were used, the concordance between phenotypic and genotypic testing for these XDR strains would vary between rifampicin (100%), isoniazid (92%), flouroquinolones (81%), aminoglycoside (78%) and ethambutol (62%); while pncA sequencing would provide genotypic resistance in less than half the isolates. This work highlights the importance of expanded

Whole Genome Sequencing Based Characterization of Extensively Drug-Resistant Mycobacterium tuberculosis Isolates from Pakistan

KAUST Repository

Ali, Asho

2015-02-26

Improved molecular diagnostic methods for detection drug resistance in Mycobacterium tuberculosis (MTB) strains are required. Resistance to first- and second- line anti-tuberculous drugs has been associated with single nucleotide polymorphisms (SNPs) in particular genes. However, these SNPs can vary between MTB lineages therefore local data is required to describe different strain populations. We used whole genome sequencing (WGS) to characterize 37 extensively drug-resistant (XDR) MTB isolates from Pakistan and investigated 40 genes associated with drug resistance. Rifampicin resistance was attributable to SNPs in the rpoB hot-spot region. Isoniazid resistance was most commonly associated with the katG codon 315 (92%) mutation followed by inhA S94A (8%) however, one strain did not have SNPs in katG, inhA or oxyR-ahpC. All strains were pyrazimamide resistant but only 43% had pncA SNPs. Ethambutol resistant strains predominantly had embB codon 306 (62%) mutations, but additional SNPs at embB codons 406, 378 and 328 were also present. Fluoroquinolone resistance was associated with gyrA 91-94 codons in 81% of strains; four strains had only gyr B mutations, while others did not have SNPs in either gyrA or gyrB. Streptomycin resistant strains had mutations in ribosomal RNA genes; rpsL codon 43 (42%); rrs 500 region (16%), and gidB (34%) while six strains did not have mutations in any of these genes. Amikacin/kanamycin/capreomycin resistance was associated with SNPs in rrs at nt1401 (78%) and nt1484 (3%), except in seven (19%) strains. We estimate that if only the common hot-spot region targets of current commercial assays were used, the concordance between phenotypic and genotypic testing for these XDR strains would vary between rifampicin (100%), isoniazid (92%), flouroquinolones (81%), aminoglycoside (78%) and ethambutol (62%); while pncA sequencing would provide genotypic resistance in less than half the isolates. This work highlights the importance of expanded

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

[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.

Protective Vaccine Efficacy of the Complete Form of PPE39 Protein from Mycobacterium tuberculosis Beijing/K Strain in Mice.

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Kim, Ahreum; Hur, Yun-Gyoung; Gu, Sunwha; Cho, Sang-Nae

2017-11-01

The aim of this study was to evaluate the protective efficacy of MTBK_24820, a complete form of PPE39 protein derived from a predominant Beijing/K strain of Mycobacterium tuberculosis in South Korea. Mice were immunized with MTKB_24820, M. bovis Bacilli Calmette-Guérin (BCG), or adjuvant prior to a high-dosed Beijing/K strain aerosol infection. After 4 and 9 weeks, bacterial loads were determined and histopathologic and immunologic features in the lungs and spleens of the M. tuberculosis -infected mice were analyzed. Putative immunogenic T-cell epitopes were examined using synthetic overlapping peptides. Successful immunization of MTBK_24820 in mice was confirmed by increased IgG responses ( P tuberculosis Beijing/K-strain is frequently isolated from TB patients. Copyright © 2017 American Society for Microbiology.

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.

Detection of rifampin resistance patterns in Mycobacterium tuberculosis strains isolated in Iran by polymerase chain reaction-single-strand conformation polymorphism and direct sequencing methods

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Bahram Nasr Isfahani

2006-09-01

Full Text Available Mutations in the rpoB locus confer conformational changes leading to defective binding of rifampin (RIF to rpoB and consequently resistance in Mycobacterium tuberculosis. Polymerase chain reaction-single-strand conformation polymorphism (PCR-SSCP was established as a rapid screening test for the detection of mutations in the rpoB gene, and direct sequencing has been unambiguously applied to characterize mutations. A total of 37 of Iranian isolates of M. tuberculosis, 16 sensitive and 21 resistant to RIF, were used in this study. A 193-bp region of the rpoB gene was amplified and PCR-SSCP patterns were determined by electrophoresis in 10% acrylamide gel and silver staining. Also, 21 samples of 193-bp rpoB amplicons with different PCR-SSCP patterns from RIFr and 10 from RIFs were sequenced. Seven distinguishable PCR-SSCP patterns were recognized in the 21 Iranian RIFr strains, while 15 out of 16 RIFs isolates demonstrated PCR-SSCP banding patterns similar to that of sensitive standard strain H37Rv. However one of the sensitive isolates demonstrated a different pattern. There were seen six different mutations in the amplified region of rpoB gene: codon 516(GAC/GTC, 523(GGG/GGT, 526(CAC/TAC, 531(TCG/TTG, 511(CTG/TTG, and 512(AGC/TCG. This study demonstrated the high specificity (93.8% and sensitivity (95.2% of PCR-SSCP method for detection of mutation in rpoB gene; 85.7% of RIFr strains showed a single mutation and 14.3% had no mutations. Three strains showed mutations caused polymorphism. Our data support the common notion that rifampin resistance genotypes are generally present mutations in codons 531 and 526, most frequently found in M. tuberculosis populations regardless of geographic origin.

Genomic analysis of globally diverse Mycobacterium tuberculosis strains provides insights into emergence and spread of multidrug resistance

Science.gov (United States)

Manson, Abigail L.; Cohen, Keira A.; Abeel, Thomas; Desjardins, Christopher A.; Armstrong, Derek T.; Barry, Clifton E.; Brand, Jeannette; Chapman, Sinéad B.; Cho, Sang-Nae; Gabrielian, Andrei; Gomez, James; Jodals, Andreea M.; Joloba, Moses; Jureen, Pontus; Lee, Jong Seok; Malinga, Lesibana; Maiga, Mamoudou; Nordenberg, Dale; Noroc, Ecaterina; Romancenco, Elena; Salazar, Alex; Ssengooba, Willy; Velayati, A. A.; Winglee, Kathryn; Zalutskaya, Aksana; Via, Laura E.; Cassell, Gail H.; Dorman, Susan E.; Ellner, Jerrold; Farnia, Parissa; Galagan, James E.; Rosenthal, Alex; Crudu, Valeriu; Homorodean, Daniela; Hsueh, Po-Ren; Narayanan, Sujatha; Pym, Alexander S.; Skrahina, Alena; Swaminathan, Soumya; Van der Walt, Martie; Alland, David; Bishai, William R.; Cohen, Ted; Hoffner, Sven; Birren, Bruce W.; Earl, Ashlee M.

2017-01-01

Multidrug-resistant tuberculosis (MDR-TB), caused by drug resistant strains of Mycobacterium tuberculosis, is an increasingly serious problem worldwide. In this study, we examined a dataset of 5,310 M. tuberculosis whole genome sequences from five continents. Despite great diversity with respect to geographic point of isolation, genetic background and drug resistance, patterns of drug resistance emergence were conserved globally. We have identified harbinger mutations that often precede MDR. In particular, the katG S315T mutation, conferring resistance to isoniazid, overwhelmingly arose before rifampicin resistance across all lineages, geographic regions, and time periods. Molecular diagnostics that include markers for rifampicin resistance alone will be insufficient to identify pre-MDR strains. Incorporating knowledge of pre-MDR polymorphisms, particularly katG S315, into molecular diagnostics will enable targeted treatment of patients with pre-MDR-TB to prevent further development of MDR-TB. PMID:28092681

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.

TLR2-Modulating Lipoproteins of the Mycobacterium tuberculosis Complex Enhance the HIV Infectivity of CD4+ T Cells.

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Ciaran Skerry

Full Text Available Co-infection with Mycobacterium tuberculosis accelerates progression from HIV to AIDS. Our previous studies showed that M. tuberculosis complex, unlike M. smegmatis, enhances TLR2-dependent susceptibility of CD4+ T cells to HIV. The M. tuberculosis complex produces multiple TLR2-stimulating lipoproteins, which are absent in M. smegmatis. M. tuberculosis production of mature lipoproteins and TLR2 stimulation is dependent on cleavage by lipoprotein signal peptidase A (LspA. In order to determine the role of potential TLR2-stimulating lipoproteins on mycobacterial-mediated HIV infectivity of CD4+ T cells, we generated M. smegmatis recombinant strains overexpressing genes encoding various M. bovis BCG lipoproteins, as well as a Mycobacterium bovis BCG strain deficient in LspA (ΔlspA. Exposure of human peripheral blood mononuclear cells (PBMC to M. smegmatis strains overexpressing the BCG lipoproteins, LprF (p<0.01, LprH (p<0.05, LprI (p<0.05, LprP (p<0.001, LprQ (p<0.005, MPT83 (p<0.005, or PhoS1 (p<0.05, resulted in increased HIV infectivity of CD4+ T cells isolated from these PBMC. Conversely, infection of PBMC with ΔlspA reduced HIV infectivity of CD4+ T cells by 40% relative to BCG-infected cells (p<0.05. These results may have important implications for TB vaccination programs in areas with high mother-to-child HIV transmission.

TLR2-Modulating Lipoproteins of the Mycobacterium tuberculosis Complex Enhance the HIV Infectivity of CD4+ T Cells.

Science.gov (United States)

Skerry, Ciaran; Klinkenberg, Lee G; Page, Kathleen R; Karakousis, Petros C

2016-01-01

Co-infection with Mycobacterium tuberculosis accelerates progression from HIV to AIDS. Our previous studies showed that M. tuberculosis complex, unlike M. smegmatis, enhances TLR2-dependent susceptibility of CD4+ T cells to HIV. The M. tuberculosis complex produces multiple TLR2-stimulating lipoproteins, which are absent in M. smegmatis. M. tuberculosis production of mature lipoproteins and TLR2 stimulation is dependent on cleavage by lipoprotein signal peptidase A (LspA). In order to determine the role of potential TLR2-stimulating lipoproteins on mycobacterial-mediated HIV infectivity of CD4+ T cells, we generated M. smegmatis recombinant strains overexpressing genes encoding various M. bovis BCG lipoproteins, as well as a Mycobacterium bovis BCG strain deficient in LspA (ΔlspA). Exposure of human peripheral blood mononuclear cells (PBMC) to M. smegmatis strains overexpressing the BCG lipoproteins, LprF (p<0.01), LprH (p<0.05), LprI (p<0.05), LprP (p<0.001), LprQ (p<0.005), MPT83 (p<0.005), or PhoS1 (p<0.05), resulted in increased HIV infectivity of CD4+ T cells isolated from these PBMC. Conversely, infection of PBMC with ΔlspA reduced HIV infectivity of CD4+ T cells by 40% relative to BCG-infected cells (p<0.05). These results may have important implications for TB vaccination programs in areas with high mother-to-child HIV transmission.

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...

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

Science.gov (United States)

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.

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

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

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

Effect of egg yolk on growth of Mycobacterium tuberculosis in 7H12 liquid medium

International Nuclear Information System (INIS)

Kononov, Y.; Ta, K.D.; Heifets, L.

1988-01-01

Of 92 drug-resistant Mycobacterium tuberculosis strains isolated from sputum specimens, 86 showed growth in two types of 7H12 broth, one with egg yolk and the other without egg yolk. In addition, two strains grew only in plain 7H12 broth without yolk, and four others were recovered only in the medium supplemented with egg yolk. The radiometrically detected growth was higher in the presence of egg yolk, corresponding to a higher number of CFU per milliliter in these cultures. The improvement of growth in 7H12 broth supplemented with egg yolk was most noticeable in cultures isolated from sputum specimens having a low number of acid-fast bacilli in the smear and producing only a few colonies on solid media

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

Mycobacterium bovis in a European bison (Bison bonasus) raises concerns about tuberculosis in Brazilian captive wildlife populations: a case report.

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Zimpel, Cristina Kraemer; Brum, Juliana Sperotto; de Souza Filho, Antônio Francisco; Biondo, Alexander Welker; Perotta, João Henrique; Dib, Cristina Corsi; Bonat, Marcelo; Neto, José Soares Ferreira; Brandão, Paulo Eduardo; Heinemann, Marcos Bryan; Guimaraes, Ana Marcia Sa

2017-02-10