Objective/background: The present investigation was undertaken to synthesize zinc oxide nanoparticles using Limonia acidissima L. and to test their efficacy against the growth of Mycobacterium tuberculosis. Methods: The formation of zinc oxide nanoparticles was confirmed with UV–visible spectrophotometry. Fourier transform infrared spectroscopy shows the presence of bio-molecules involved in the stabilization of zinc oxide nanoparticles. The shape and size was confirmed with atomic force microscope, X-ray diffraction, and high resolution transmission electron microscope. These nanoparticles were tested for their effect on the growth of M. tuberculosis through the microplate alamar blue assay technique. Results: The UV–visible data reveal that an absorbance peak at 374nm confirms formation of zinc oxide nanoparticles and they are spherical in shape with sizes between 12nm and 53nm. These nanoparticles control the growth of M. tuberculosis at 12.5μg/mL. Conclusion: Phytosynthesis of zinc oxide nanoparticles is a green, eco-friendly technology because it is inexpensive and pollution free. In the present investigation, based on our results we conclude that the aqueous extract of leaves of L. acidissima can be used for the synthesis of zinc oxide nanoparticles. These nanoparticles control the growth of M. tuberculosis and this was confirmed with the microplate alamar blue method. The potential of biogenic zinc oxide nanoparticles may be harnessed as a novel medicine ingredient to combat tuberculosis disease.

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Currently, one third of the world's population is latently infected with Mycobacterium tuberculosis (MTB), while 8.9–9.9 million new and relapse cases of tuberculosis (TB) are reported yearly. The renewed research interests in natural products in the hope of discovering new and novel antitubercular leads have been driven partly by the increased incidence of multidrug-resistant strains of MTB and the adverse effects associated with the first- and second-line antitubercular drugs. Natural products have been, and will continue to be a rich source of new drugs against many diseases. The depth and breadth of therapeutic agents that have their origins in the secondary metabolites produced by living organisms cannot be compared with any other source of therapeutic agents. Discovery of new chemical molecules against active and latent TB from natural products requires an interdisciplinary approach, which is a major challenge facing scientists in this field. In order to overcome this challenge, cutting edge techniques in mycobacteriology and innovative natural product chemistry tools need to be developed and used in tandem. The present review provides a cross-linkage to the most recent literature in both fields and their potential to impact the early phase of drug discovery against TB if seamlessly combined.

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Current World Health Organisation targets calling for an end to the global tuberculosis (TB) epidemic by 2035 require a dramatic improvement in current case-detection strategies. A reliance on passive case finding (PCF) has resulted consistently, in over three million infectious TB cases per year, being missed by the health system, leading to ongoing transmission of infection within families and communities. Active case finding (ACF) for TB has been recognized as an important complementary strategy to PCF, in order to diagnose and treat patients earlier, reducing the period of infectiousness and therefore transmission. ACF may also achieve substantial population-level TB control. Local TB epidemiology and the resources available in each setting will influence which populations should be screened, and the types of ACF interventions to use for maximal impact. TB control programs should begin with the highest risk groups and broaden their activities as resources allow. Mathematical models can help to predict the population-level effects and the cost-effectiveness of a variety of ACF strategies on different risk populations.

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Pulmonary infections with nontuberculous mycobacteria (NTM) are recognized as a problem in immunodeficient individuals and are increasingly common in older people with no known immune defects. NTM are found in soil and water, but factors influencing transmission from the environment to humans are mostly unknown. Studies of the epidemiology of NTM disease have matched some clinical isolates of NTM with isolates from the patient's local environment. Definitive matching requires strain level differentiation based on molecular analyses, including partial sequencing, PCR-restriction fragment length polymorphism (RFLP) analysis, random amplified polymorphic DNA (RAPD) PCR, repetitive element (rep-) PCR and pulsed field gel electrophoresis (PFGE) of large restriction fragments. These approaches have identified hospital and residential showers and faucets, hot-tubs and garden soil as sources of transmissible pathogenic NTM. However, gaps exist in the literature, with many clinical isolates remaining unidentified within environments that have been tested, and few studies investigating NTM transmission in developing countries. To understand the environmental reservoirs and transmission routes of pathogenic NTM, different environments, countries and climates must be investigated.

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Aims: The current study was designed to document medicinal plant species that are traditionally used to treat tuberculosis (TB) by Ghanaian communities. Methods: The medicinal plants used against TB or its signs and symptoms were selected using library and online published data searches. A guided questionnaire interview was also conducted with a botanist involved in plant collection at the Centre for Scientific Research into Plant Medicine (CSRPM) at Mampong. Data obtained were entered in Excel and summarized into means and frequencies using SPSS 12.0.1 for windows, and expressed as tables and bar graphs. Results: A total of 15 medicinal plant species distributed between 13 genera and 13 families were documented. The following medicinal plant species were found to be used against TB in Greater Accra and Eastern parts of Ghana: Azadirachta indica A. Juss. Stem bark (Meliaceae), Hygrophila auriculata Heine, whole plant (Acanthaceae), Chenopodium ambrosioides L. leaves (Amaranthaceae), Coix lacryma-jobi L. glumes (Poaceae), Solanum torvum Sw. unripe fruits (Solanaceae), Solanum torvum Sw. leaves (Solanaceae), Bidens pilosa L. whole plant (Asteraceae), Phyllanthus fraternus G.L. Webster leaves (Phyllanthaceae), Dissotis rotundifolia (Sm.) Triana, leaves (Melastomataceae), Cymbopogon giganteus Chiov. Leaves (Poaceae), Cyperus articulatus L. roots (Cyperaceae), Allium sativum L. bulb (Amaryllidaceae), Zingiber officinale Roscoe, rhizomes (Zingiberaceae), Allium cepa L. bulbs (Amaryllidaceae), Allium cepa L. leaves (Amaryllidaceae), Aloe vera var. barbadensis aqueous extract from leaves (Xanthorrhoeaceae), Aloe vera var. barbadensis organic extract from leaves (Xanthorrhoeaceae), Cocos nucifera Linn, water (Arecaceae) and Cocos nucifera Linn. Husk (Arecaceae). Conclusions: The collected plant species could be a source of a new class of drugs against TB. Bioactivity guided fractionation is recommended to identify lead compounds for antimycobacterial activity. The current paper documents for the first time medicinal plant species used by Ghanaian communities to treat TB. These results are a basis for selection of plants for further pharmacological, toxicological and phytochemical studies in developing new plant-based antimycobacterial drugs.

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Tuberculosis continues to cast a huge impact on humanity with its high incidence and mortality, especially in developing countries. For tuberculosis case detection, microscopy continues to be indispensible, given its low cost, rapidity, simplicity of procedure and high specificity. Modifications have attempted to improve the sensitivity of microscopy which include: concentration methods such as centrifugation, N-acetyl cysteine–sodium hydroxide, bleach, ammonium sulfate or chitin. Furthermore, classical Ziehl–Neelsen (ZN) staining has been subjected to varying carbol fuchsin concentrations or replaced by Kinyoun staining, fluorescent microscopy or immune-fluorescence. Currently, light emitting diode fluorescence is recognizably the most plausible method as an alternative to ZN staining.

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Background: Mycobacterium tuberculosis (Mtb) has evolved multiple mechanisms to manipulate its cellular niche for its own advantage. Many efforts have been made to understand basal mechanisms of mycobacterial infections. However, the underlying molecular regulation is not fully understood. Recently, a new class of non-coding, small RNAs, called microRNAs (miRNAs), has emerged as important regulators in biological processes, and their involvement in mycobacterial infection has been identified, thus opening a new field of research. Methods: This study aimed to determine by TaqMan Low Density Array the host genome-wide miRNA expression profile of primary human monocyte-derived macrophages (MDM) infected with two members of the Mtb complex: virulent Mtb H37Rv and the non-virulent vaccine strain Mycobacterium bovis Bacillus Calmette-Guerin (BCG) in comparison with chemically-inactivated Mtb bacilli. Results: The findings of this study showed that infection of MDM with H37Rv or BCG results in a signature of miRNA expression mostly overlapping between the two mycobacteria. A substantially different signature emerged from infection with killed virulent bacilli, suggesting an active influence of live intracellular bacteria on cellular miRNA metabolism. Specifically, Mtb induced miRNA signature is composed of miRNAs well established in immune regulation, miR-155 and miR-146a, as well as a set of miRNAs newly associated with Mtb infection: miR-145, miR-222*, miR-27a and miR-27b. All of these miRNAs are predicted to target important immune-related genes. Conclusions: This study signifies the miRNA host response upon intracellular mycobacterial infection in macrophages, providing new aspects of regulation in host-pathogen interactions, at post-transcriptional levels.

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Nontuberculous mycobacteria (NTM) are emergent pathogens whose importance in human health has been gaining relevance after being recognized as etiological agents of opportunist infections in HIV patients. Currently, NTM are recognized as etiological agents of several respiratory and extra-respiratory infections of immune-competent individuals. The environmental nature of NTM together with the ability to assemble biofilms on different surfaces plays a key role on their pathogenesis. In the present work the ability of three fast-growing NTM (Mycobacterium smegmatis, Mycobacterium fortuitum and Mycobacterium chelonae) to persist within a model of human alveolar macrophages was evaluated. Most often human infections with NTM occur by contact with the environment. Biofilms can work as environmental reservoirs. For this reason, it was decided to evaluate the ability of NTM to assemble biofilms on different surfaces. Scanning electron microscopy was used to elucidate the biofilm structure. The ability to assemble biofilms was connected with the ability to spread on solid media known as sliding. Biofilm assembly and intracellular persistence seems to be ruled by different mechanisms.

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To evaluate the performance of TrueNAT (RT Micro PCR device) assay in comparison with GeneXpert on sputum samples from pulmonary cases of tuberculosis. 274 samples were processed to detect MTB by ZN smear examination, MGIT culture and molecular methods that included RT-PCR (ABI 7500 & TrueNAT) and GeneXpert for case detection of TB. The overall performance of the test with MGIT(Mycobacterium Growth Indicator Tube) culture as gold standard, sensitivity of smear, RT PCR/TrueNAT and Genexpert was 61.5% (CI:53.3–69.3%), 94.7% (CI:89.8–97.6%) & 96.0% (CI: 91.5–98.5%), respectively. Amongst the S+ (108) samples, RT-PCR/TrueNAT and GeneXpert showed a sensitivity of 99% (CI:94.9%–99.8%) and 100% (98.6%–100.0%), respectively. High concordance was observed between GeneXpert and TrueNAT for case detection of TB. The GeneXpert MTB/RIF test was independent on the user's skills. It has a short turn-around time and simultaneously detects RIF resistance with M. tuberculosis in less than 3 h. The TrueNAT MTB has good sensitivity and specificity in case detection with hands on time of less than 3 h as well as fits the requirements in resourcelimited health care settings. Larger, multi-site studies are required to obtain better estimates of the performance of TrueNAT MTB.

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Objective/background: Microbial infections such as tuberculosis is a major cause of mortality worldwide. Plant-derived phytochemicals have a long history of providing much-needed novel therapeutics. Triterpenoids are among the prominent phytochemicals that possess numerous biological activities. Among them is maslinic acid (MA), a biologically active olean-type pentacyclic triterpenoid. In search of a novel antimicrobial agent, we aimed to evaluate the antimicrobial potential of MA. Methods: Antibacterial and antifungal activity was evaluated through the agar well diffusion method. Antitubercular activity was analysed through the agar well diffusion and disc diffusion methods, respectively. Antioxidant capacity was determined through assays for total antioxidant capacity, 2,2-diphenyl-1-picrylhydrazyl radical scavenging, hydrogen peroxide radical scavenging, and Fe³⁺ reducing power. The program Prediction of Activity Spectra for Substances was used to calculate the possible biological activity of MA. Results: MA showed dose-dependent antioxidant activity similar to that of ascorbic acid. It had no inhibitory effect on bacterial strains, but it had moderate activity against the fungi Aspergillus flavus and Ustilago maydis, with Aspergillus niger being the most sensitive to MA. MA also exhibited strong antimycobacterial activity. Probable antioxidant, antibacterial, and antifungal activity of MA based on software calculations are 0.479, 0.363 and 0.589 respectively. Conclusion: This work provides scientific evidence of the antioxidant, antifungal, and antimycobacterial activities of MA, showing its potential application in the development of natural antioxidants and antimicrobial agents for the agro-food and pharmaceutical industries.

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Purpose: PCR assay is a highly sensitive, specific and reliable diagnostic tool for the identification of pathogens in many infectious diseases. Genome sequencing Mycobacterium leprae revealed several gene targets that could be used for the detection of DNA from clinical and environmental samples. The PCR sensitivity of particular gene targets for specific clinical and environmental isolates has not yet been established. The present study was conducted to compare the sensitivity of RLEP, rpoT, Sod A and 16S rRNA gene targets in the detection of M. leprae in slit skin smear (SSS), blood, soil samples of leprosy patients and their surroundings. Method: Leprosy patients were classified into Paucibacillary (PB) and Multibacillary (MB) types. Ziehl–Neelsen (ZN) staining method for all the SSS samples and Bacteriological Index (BI) was calculated for all patients. Standard laboratory protocol was used for DNA extraction from SSS, blood and soil samples. PCR technique was performed for the detection of M. leprae DNA from all the above-mentioned samples. Results: RLEP gene target was able to detect the presence of M. leprae in 83% of SSS, 100% of blood samples and in 36% of soil samples and was noted to be the best out of all other gene targets (rpoT, Sod A and 16S rRNA). It was noted that the RLEP gene target was able to detect the highest number (53%) of BI-negative leprosy patients amongst all the gene targets used in this study. Conclusion: Amongst all the gene targets used in this study, PCR positivity using RLEP gene target was the highest in all the clinical and environmental samples. Further, the RLEP gene target was able to detect 53% of blood samples as positive in BI-negative leprosy cases indicating its future standardization and use for diagnostic purposes.

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Background: Antituberculosis drugs cause hepatotoxicity in some individuals leading to acute liver failure, which results in death. Such phenomena limit the clinical use of drugs, contributing to treatment failure that possibly causes drug resistance. Furthermore, associated risk factors for the development of antituberculosis-drug-induced hepatotoxicity (anti-TB-DIH) are found to be controversial among different study findings. Methods: A prospective cohort study was conducted from May 2014 to October 2014 in Dawro Zone, Tercha District Hospital Laboratory, South Ethiopia. One hundred and twenty-four new tuberculosis-positive individuals available from Tercha Hospital and five health centers during data collection were consecutively included. The sociodemographic data and anthropometric measurement were obtained. Then, 5 mL of venous blood was drawn from each individual, and the alanine transaminase, aspartate transaminase, and total bilirubin were measured photometrically at baseline, and then continuously monitored by measuring these liver enzymes every 2 weeks for 2 months. Data were analyzed with SPSS version 20 for Windows (SPSS Inc., Chicago, IL, USA). Results: The incidence of anti-TB-DIH was found to be 8% (10 patients out of 124). Raised serum transaminase and bilirubin level, as well as signs and symptoms of hepatotoxicity (nausea, anorexia, vomiting, malaise, and jaundice), were observed in the cases. The onset of hepatotoxicity ranged from 13 days to 58 days (median, 26 days) after treatment was initiated. Of the various risk factors analyzed, only high alcohol intake was associated with the incidence of anti-TB-DIH (odds ratio=9.3, 95% confidence interval 1.8–47, p <.007). Age, gender, extent of tuberculosis disease, and malnutrition were not significantly associated with anti-TB-DIH. Conclusion: The incidence of anti-TB-DIH in Dawro Zone was high. The drug responsible for the hepatotoxicity was not known. However, chronic high alcohol intake was associated with the development of anti-TB-DIH.

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Tuberculosis (TB) is an ancient disease caused by Mycobacterium tuberculosis (MTB), which remains a major cause for morbidity and mortality in several developing countries. Most drug-resistant MTB clinical strains are resistant to isoniazid (INH), a first-line anti-TB drug. Mutation in KatG, a catalase-peroxidase, of MTB is reported to be a major cause of INH resistance. Normally upon activation by KatG, INH is converted to an active intermediate which has antimycobacterial action in MTB. This INH intermediate in the presence of NADH forms INH-NAD adduct which inhibits inhA (2-trans-enoyl-acyl carrier protein reductase) of MTB, thus blocking the synthesis of mycolic acid, a major lipid of the mycobacterial cell wall. In this docking study, the high binding affinity of INH-NAD adduct towards InhA was observed in comparison with INH alone. In this study, two resistant mutants of KatG (S315T and S315N) were modeled using Modeller9v10 and docking analysis with INH was performed using AutoDock4.2 and the docking results of these mutants were compared with the wild type KatG. Docking results revealed the formation of a single hydrogen (H) bond between the secondary amine nitrogen (–NH) of INH with Thr or Asn residues in place of Serine at 315 position of KatG mutant strains respectively, whereas in the case of the wild type, there was no H-bond formation observed between INH and Ser315. The H-bond formation may prevent free radical formation by KatG in mutant strains thus the development of resistance to the drug. This in silico evidence may implicate the basis of INH resistance in KatG mutant strains.

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Objective/background: The in vitro drug-susceptibility testing of Mycobacterium tuberculosis reports isolates as resistant or susceptible on the basis of single critical concentrations. It is evident that drug resistance in M. tuberculosis is quite heterogeneous, and involves low level, moderate level, and high level of drug-resistant phenotypes. Thus, the aim of our study was to correlate rrs (X52917) and eis (AF144099) promoter mutations, found in M. tuberculosis isolates, with corresponding minimum inhibitory concentrations of amikacin, kanamycin, and capreomycin. Methods: Ninety M. tuberculosis clinical isolates were analyzed in this study. The minimum inhibitory concentrations were determined by MGIT 960 for 59 isolates with resistance-associated mutations in the rrs and eis promoter gene regions, and 31 isolates with wild-type sequences, as determined by the GenoType MTBDRsl (version 1) assay. Results: The rrs A1401G mutation was identified in 48 isolates resistant to the second-line injectables. The eis promoter mutations C-14T (n=3), G-10C (n=3), G-10A (n=3), and C-12T (n=2) were found within 11 isolates with various resistance profiles to the second-line injectables. Thirty-one isolates had wild-type sequences for the rrs and eis promoter gene regions of interest, one of which was amikacin, kanamycin, and capreomycin resistant. The isolates with the rrs A1401G mutation had amikacin, kanamycin, and capreomycin minimum inhibitory concentrations of >40 mg/L, >20 mg/L, and 5–15 mg/L, respectively. The isolates with eis promoter mutations had amikacin, kanamycin, and capreomycin minimum inhibitory concentrations of 0.25–1.0 mg/L, 0.625–10 mg/L, and 0.625–2.5 mg/L, respectively. Conclusion: This study provides a preliminary basis for the prediction of phenotypic-resistance levels to the second-line injectables based upon the presence of genetic mutations associated with amikacin, kanamycin, and capreomycin resistance. The results suggest that isolates with eis promoter mutations have consistently lower resistance levels to amikacin, kanamycin, and capreomycin than isolates with the rrs A1401G mutation.

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Objective/background: Nontuberculous mycobacteria (NTM) infection associated with pulmonary and extrapulmonary disease has been increasing globally. Despite an increase in incidence rate of NTM infection, its prevalence, species diversity, and circulation pattern in India is largely unknown. This study sought to investigate the overall burden and diversity of NTM among both pulmonary and extrapulmonary clinical isolates from a Northern Indian population. Methods: The study was conducted in the Department of Microbiology, from January 2013 to December 2015. A total of 4620 clinical samples were collected from patients suspected to have pulmonary and extrapulmonary tuberculosis. Preliminary diagnosis was performed using Ziehl–Neelsen staining followed by liquid culture in BacT/ALERT three-dimensional system. A total of 906 positive cultures obtained were differentiated as either NTM or Mycobacterium tuberculosis complex using a biochemical and MPT64 antigen test. Further identification of NTM species was confirmed with a line probe assay. Results: Out of 906 cultures isolates, 263 (29.0%) were confirmed as NTM and 643 (71.0%) were identified as Mycobacterium tuberculosis complex. A total of 79.4% of the NTM were recovered from pulmonary and 18.2% from extrapulmonary specimens. The diversity of NTM species was high (13 species) and predominated by Mycobacterium abscessus (31.3%) followed by Mycobacterium fortuitum (22%), Mycobacterium intracellulare (13.6%), Mycobacterium chelonae (9.1%), however, M. abscessus and M. fortuitum were the predominant species in both types of clinical isolates. Men (60.4%) and older patients aged greater than 55 years were the predominated risk group for NTM infection. Conclusion: The high prevalence and species diversity of NTM suggests the need for immediate and accurate characterization of NTM for proper treatment and management of patients.

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Background: Mycobacterium tuberculosis (Mtb) strains H37Ra and H37Rv are commonly used to study new and re-evaluate old antituberculous agents with respect to their pharmacodynamic effects in vitro. The differences in membrane proteins and, in particular, differences in carrier proteins between Mtb H37Ra and Mtb H37Rv may have an impact on antibiotic potency. The question of whether H37Ra can be used as a reliable surrogate for H37Rv and clinical strains has not been addressed sufficiently. The purpose of this study is to provide a full comparison of susceptibility data of the most common antituberculosis (TB) agents against both Mtb strains. Methods: In addition to a literature review, in vitro checkerboard susceptibility study was conducted comparing the in vitro minimum inhibitory concentration (MIC) of 16 common antituberculous drugs against H37Ra and H37Rv. Heifets–Sanchez TB agar drug susceptibility plates were utilized. Results: Half of the antibiotics demonstrated similar growth inhibition against both strains, while slightly differing MIC values were found for 7 of 16 drugs. With the exception of rifampicin, no marked difference in MIC against H37Ra and H37Rv was observed. Conclusion: While neither the attenuated (H37Ra) nor the virulent strain (H37Rv) is a clinical strain, both strains predicted MICs of clinical isolates equally well, when comparing the current in vitro results to clinical susceptibility data in the literature. H37Ra comes with the benefits of lower experimental costs and less administrative barriers including the requirement of a biosafety Level III environment.

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Epidemiological data show a worldwide increase in nontuberculous mycobacteriosis. Although it has been partially attributed to the improvement of microbiological methodologies that has allowed a better recovery and identification of nontuberculous mycobacteria (NTM), it is generally accepted that there is a genuine incidence augmentation. The reasons of the increase are likely multifactorial, depending on the nature of the pathogen, the host, and their interaction. Mycobacteria from the Mycobacterium tuberculosis complex has been regarded as pathogenic and NTM as opportunistic and nontransmissible. Nevertheless, few differences have been found in either their phenotypic or genotypic characteristics. The phenomenon of M. tuberculosis adaptation to the human host may be taking place again in NTM as a consequence of human environmental alterations that facilitate the interaction with the pathogen. The current worsening of the immunological status of increasing numbers of individuals, a result of factors such as malnutrition (obesity and diabetes), population aging or the widespread use of immunosuppressive medication, may be allowing the rapid evolution and person-to-person transmission of NTM. It is likely that mycobacteriosis incidence will keep escalating. New measures should be taken to deal with these diseases, including their reportability and the implementation of strain genotyping that would shed light on the NTM dissemination routes from the environment or human hosts.

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Introduction: GeneXpert MTB/RIF is a fully-automated diagnostic molecular test which simultaneously detects tuberculosis (TB) and rifampicin (RIF) drug resistance. The purpose of this study is to evaluate the performance of the GeneXpert MTB/RIF test for the detection of Mycobacterium tuberculosis complex (MTBC) in lymph node specimens and to show the place of Mycobacterium bovis as a major cause of TB lymphadenitis. Material and methods: This study was conducted simultaneously in the National Reference Laboratory for Mycobacteria of Ariana and the Central Laboratory of Sfax, from January to December 2013. In total, 174 lymph node specimens were processed simultaneously for Ziehl–Neelsen, auramine and immuno-histochemical staining. Conventional culture on both Lowenstein–Jensen and liquid medium (Bactec MGIT 960 BD system) and the new molecular-based GeneXpert MTB/RIF assay system were performed. Positive cultures were confirmed using molecular identification (Genotype MTBC Hain Lifescience). Results: Among the 174 samples tested, the GeneXpert detected the DNA of MTBC in 134 samples (77%). Standard bacteriological assays, including AFB microscopy and culture, were positive, respectively, in 41 (23.6%) and 79 (45.4%) specimens. M. bovis was isolated in 76% of positive cultures. GeneXpert sensitivity and specificity results were assessed according to smear and culture results, clinical and histological findings. The sensitivity and specificity of the Xpert assay were 87.5% (126/144) and 73.3%, respectively. Conclusion: The implementation of the GeneXpert MTB/RIF assay may dramatically improve the rapid diagnosis of lymph node TB.

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Objective: To evaluate risk factors for multidrug-resistant tuberculosis (MDR-TB) in an urban setting of Pakistan. Design and methods: In this multicenter case–control study, patients aged 15 years old or older with sputum culture and sensitivity (C/S) diagnosed with pulmonary MDR-TB were defined as cases, whereas patients aged 15 years old or older with sputum C/S diagnosed and susceptible to pulmonary TB were regarded as controls. Fifty cases and 75 controls were enrolled from three tertiary-care hospitals in Karachi. Results: Multivariable logistic regression models showed that cases were more likely to have had a TB patient in the house prior to the diagnosis of MDR-TB (adjusted odds ratio [OR_adj]=3.1, 95% confidence interval [CI]: 1.2, 8.3) or had a history of prior TB treatment (OR_adj = 4.2, 95% CI: 1.1, 15.4). Furthermore, cases compared with controls tended to be male (OR_adj = 3.6, 95% CI: 1.4, 9.7), 15–25 years of age (OR_adj = 3.7, 95% CI: 1.2, 11.3), of Sindhi ethnicity (adjusted OR=9.1, 95% CI: 1.9, 43.4) or with low educational attainment (OR_adj OR=5.5, 95% CI: 1.7–17.6, for no formal schooling; OR_adj = 3.8, 95% CI: 1.1–14.1, 1 for 1–5 school years). Conclusions: A TB patient in the house or a history of prior TB treatment was strongly associated with MDR-TB in this study. Furthermore, younger age, male gender, Sindhi ethnicity and poor educational attainment entailed a high risk for MDR-TB. Targeted educational intervention for patients and their contacts may minimize the noncompliance with prescribed TB treatment and lessen MDR-TB magnitude in settings like Karachi.

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Representative strains of Mycobacterium avium, Mycobacterium intracellulare and Mycobacterium scrofulaceum (MAIS) grew at equal rates in laboratory medium at 21% (air) and 12% oxygen. Growth in 6% oxygen proceeded at a 1.4–1.8-fold lower rate. Colony formation was the same at 21% (air) and 6% oxygen. The MAIS strains survived rapid shifts from aerobic to anaerobic conditions as measured by two experimental approaches (Falkinham (1996) ^[1]). MAIS cells grown aerobically to log phase in broth were diluted, spread on agar medium, and incubated anaerobically for up to 20 days at 37 °C. Although no colonies formed anaerobically, upon transfer to aerobic conditions, greater than 25% of the colony forming units (CFU) survived after 20 days of anaerobic incubation (Prince et al. (1989) ^[2]). MAIS cells grown in broth aerobically to log phase were sealed and vigorous agitation led to oxygen depletion (Wayne model). After 12 days anaerobic incubation, M. avium and M. scrofulaceum survival were high (>50%), while M. intracellulare survival was lower (22%). M. avium cells shifted to anaerobiosis in broth had increased levels of glycine dehydrogenase and isocitrate lyase. Growth of MAIS strains at low oxygen levels and their survival following a rapid shift to anaerobiosis is consistent with their presence in environments with fluctuating oxygen levels.

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Non-tuberculous mycobacteria (NTM) are environmental microbes that cause a variety of diseases both in immunocompromised and immunocompetent patients. Epidemiologic data indicate that there has been a global rise in the incidence of NTM infections. It has also been noted that NTM infections have a predilection to occur in postmenopausal women. In a recent study, it was demonstrated that in patients with non-CF bronchiectasis the probability of NTM isolation was significantly higher in elderly female patients and in those with a low body mass index. However, the mechanisms of causality of these gender differences and morpho-phenotypes remain enigmatic. The present study reviews the data and plausible mechanisms which might provide clues to this gender susceptibility and morpho-phenotypes of patients with bronchiectasis and NTM.

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Mycobacterium tuberculosis remains a major global health problem and is currently killing 1.5 million people every year. One of the most important steps in tuberculosis control is the rapid and accurate laboratory diagnosis. The Xpert MTB/RIF assay is a novel molecular, easy-to-use assay, which can lead to tuberculosis identification in less than 2 h. In this study, the Xpert MTB/RIF assay performance for rapid diagnosis of tuberculosis was evaluated in comparison with conventional culture methods; 361 pulmonary and extrapulmonary patient samples were collected between October 2010 and October 2012 and were analyzed at the National Reference laboratory for Mycobacteria, Zagreb, Croatia. For pulmonary samples the sensitivity and specificity were 86% and 100%, while for extrapulmonary samples the sensitivity and specificity were 75% and 99%, respectively. It was concluded that Xpert MTB/RIF assay has high sensitivity and specificity for both pulmonary and extrapulmonary specimens.

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Recurrent tuberculosis (TB) continues to be a significant problem and is an important indicator of the effectiveness of TB control. Recurrence can occur by relapse or exogenous reinfection. Recurrence of TB is still a major problem in high-burden countries, where there is lack of resources and no special attention is being given to this issue. The rate of recurrence is highly variable and has been estimated to range from 4.9% to 47%. This variability is related to differences in regional epidemiology of recurrence and differences in the definitions used by the TB control programs. In addition to treatment failure from noncompliance, there are several key host factors that are associated with high rates of recurrence. The widely recognized host factors independent of treatment program that predispose to TB recurrence include gender differences, malnutrition; comorbidities such as diabetes, renal failure, and systemic diseases, especially immunosuppressive states such as human immunodeficiency virus; substance abuse; and environmental exposures such as silicosis. With improved understanding of the human genome, proteome, and metabolome, additional host-specific factors that predispose to recurrence are being identified. Information on temporal and geographical trends of TB cases as well as studies with whole-genome sequencing might provide further information to enable us to fully understand TB recurrence and discriminate between reactivation and new infection. The recently launched World Health Organization End TB Strategy emphasizes the importance of integrated, patient-centered TB care. Continued improvement in diagnosis, treatment approaches, and an understanding of host-specific factors are needed to fully understand the clinical epidemiological and social determinants of TB recurrence.

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Objective/background: The nature and frequency of mutations in rifampicin (RIF) and isoniazid (INH) resistant Mycobacterium tuberculosis isolates vary considerably according to geographic locations. However, information regarding specific mutational patterns in Ethiopia remains limited. Methods: A cross-sectional prospective study was carried out among confirmed pulmonary tuberculosis cases in Southwest Ethiopia. Mutations associated with RIF and INH resistances were studied using GenoType MTBDRplus line probe assay in 112 M. tuberculosis isolates. Culture (MGIT960) and identification tests were performed at the Mycobacteriology Research Center of Jimma University, Jimma, Ethiopia. Results: Mutations conferring resistance to INH, RIF, and multidrug resistance were detected in 36.6% (41/112), 30.4% (34/112), and 27.7% (31/112) of M. tuberculosis isolates respectively. Among 34 RIF-resistant isolates, 82.4% (28/34) had rpoB gene mutations at S531L, 2.9% (1/34) at H526D, and 14.7% (5/34) had mutations only at wild type probes. Of 41 INH-resistant strains, 87.8% (36/41) had mutations in the katG gene at Ser315Thr1 and 9.8% (4/41) had mutations in the inhA gene at C15T. Mutations in inhA promoter region were strongly associated with INH monoresistance. Conclusion: A high rate of drug resistance was commonly observed among failure cases. The most frequent gene mutations associated with the resistance to INH and RIF were observed in the codon 315 of the katG gene and codon 531 of the rpoB gene, respectively. Further studies on mutations in different geographic regions using DNA sequencing techniques are warranted to improve the kit by including more specific mutation probes in the kit.

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Nontuberculous mycobacteria (NTM) are a diverse group of bacterial species that are distributed in the environment. Many of these environmental bacteria can cause disease in humans. The identification of NTM in environmental sources is important for both clinical and epidemiological purposes. In this study, the distribution of NTM species from environmental and clinical samples in the Middle East was reviewed. In order to provide an overview of NTM, as well as recent epidemiological trends, all studies addressing NTM in the Middle East from 1984 to 2014 were reviewed. A total of 96 articles were found, in which 1751 NTM strains were isolated and 1084 of which were obtained from clinical samples, 619 from environmental samples and 48 were cited by case reports. Mycobacterium fortuitum was the most common rapid growing mycobacteria (RGM) isolated from both clinical (269 out of 447 RGM; 60.1%) and environmental (135 out of 289 RGM; 46.7%) samples. Mycobacterium avium complex (MAC) was the most common slow growing mycobacteria (SGM) isolated from clinical samples (140 out of 637 SGM; 21.9%). An increasing trend in NTM isolation from the Middle East was noted over the last 5 years. This review demonstrates the increasing concern regarding NTM disease in the Middle East, emphasizing the need for regional collaboration and coordination in order to respond appropriately.

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