Intervención ergonómica evaluada por Ocra Check List a digitadores, Lima-2015 / Ergonomic intervention evaluated by Ocra Check List on data entry operators, Lima-2015

Los trastornos osteomusculares en digitadores están relacionados con los movimientos repetitivos. Objetivo:medir el efecto de una intervención ergonómica realizada a digitadores mediante Ocra Check List. Material y Métodos: investigación no experimental, longitudinal, cuantitativa en la que se realizó una medición del riesgo mediante OCRA Check List antes y después de una intervención ergonómica. Resultados: del total de 6 digitadores, 5 tenían entre 30-50 años y 4 eran mujeres. Los 6 digitadores se encontraban sintomáticos antes de la intervención y presentaban trastornos osteomusculares. Se midió con OCRA Check List antes de la intervención resultando 48.75 en a extremidad derecha y 52.50 en la extremidad izquierda. Luego de la intervención ergonómica, el riesgo resultante fue de 17.25 en la extremidad derecha y 18 en la extremidad izquierda. Además, el 80% de los digitadores se volvieron asintomáticos. Conclusión: la intervención ergonómica realizada en digitadores disminuyó el riesgo de presentar trastornos osteomusculares

Objective: to measure the effect of an ergonomic intervention using OcraCheckList risk assesment on data entry operators. Material and Methods: this is a longitudinal and quantitative study on data entry operators who underwent the Ocrachecklist risk assesment before and after the implementation of three 8 min pauses with reduction in the number of typed pages per hour. Results: there was a total of 6 data entry operators. Their ages ranged between 30 and 50 years old, four of the subjects were female. Work schedule involved five shifts of 9.5 hours a week with the number of pages typed per hour of 12. Risk assesment was performed usin the OcraCheckList Method and the number of hours with no apropriate recovery was estimated. In regards to symptomatology, six subjects presented manifestations of musculoskeletal disorders as follow : back pain, carpal tunnel syndrome and De Quervain tenosynovitis. An ergonomic intervention was implemented that included three effective pauses with a reduction in the number of typed pages per hour to 8.5 . Subjects underwent Ocrachecklist risk assesment before and after the intervention showing a score of 48.75 in RUE and 52.5 in LUE. Such scores represent a "significant" risk level (critical condition). Risk assessment was repeated two months after the intervention with resulting scores of 17.25 in RUE and 18 in LUE. In addition, 80% of the digitators became asymptomatic. Conclusion:this ergonomic intervention, on data entry operators, decreased the score of risk assesment using the OCRA CheckList Method

Imaging the Human Immunological Synapse.

The purpose of the method is to generate an immunological synapse (IS), an example of cell-to-cell conjugation formed by an antigen-presenting cell (APC) and an effector helper T lymphocyte (Th) cell, and to record the images corresponding to the first stages of the IS formation and the subsequent trafficking events (occurring both in the APC and in the Th cell). These events will eventually lead to polarized secretion at the IS. In this protocol, Jurkat cells challenged with Staphylococcus enterotoxin E (SEE)-pulsed Raji cells as a cell synapse model was used, because of the closeness of this experimental system to the biological reality (Th cell-APC synaptic conjugates). The approach presented here involves cell-to-cell conjugation, time-lapse acquisition, wide-field fluorescence microscopy (WFFM) followed by image processing (post-acquisition deconvolution). This improves the signal-to-noise ratio (SNR) of the images, enhances the temporal resolution, allows the synchronized acquisition of several fluorochromes in emerging synaptic conjugates and decreases fluorescence bleaching. In addition, the protocol is well matched with the end point cell fixation protocols (paraformaldehyde, acetone or methanol), which would allow further immunofluorescence staining and analyses. This protocol is also compatible with laser scanning confocal microscopy (LSCM) and other state-of-the-art microscopy techniques. As a main caveat, only those T cell-APC boundaries (called IS interfaces) that were at the right 90° angle to the focus plane along the Z-axis could be properly imaged and analyzed. Other experimental models exist that simplify imaging in the Z dimension and the following image analyses, but these approaches do not emulate the complex, irregular surface of an APC, and may promote non-physiological interactions in the IS. Thus, the experimental approach used here is suitable to reproduce and to confront some biological complexities occurring at the IS.

Bedaquiline and linezolid MIC distributions and epidemiological cut-off values for Mycobacterium tuberculosis in the Latin American region.

Objectives: To describe the distributions of bedaquiline and linezolid MIC values for the Mycobacterium tuberculosis WT population and to define the corresponding epidemiological cut-offs (ECOFFs) in three Latin American countries. Methods: MICs of bedaquiline and linezolid were determined by the resazurin microtitre assay (REMA). In phase 1, interlaboratory reproducibility was assessed using a panel of 10 fully susceptible M. tuberculosis strains. Phase 2 involved MIC determination for 248 clinical isolates from Argentina (n = 58), Brazil (n = 100) and Peru (n = 90) from patients who were treatment-naive for bedaquiline and linezolid. We then determined the ECOFFs for bedaquiline and linezolid by the eyeball method and the ECOFFinder statistical calculator. Results: Phase 1: REMA MIC values in the three sites were either identical to each other or differed by one 2-fold dilution from the consensus value with the exception of a single value. Phase 2: the bedaquiline MIC range was 0.0039-0.25 mg/L for pan-susceptible and drug-resistant isolates combined. The linezolid MIC range was 0.062-0.5 mg/L for pan-susceptible isolates and 0.031-4 mg/L for drug-resistant isolates. ECOFFs were 0.125 mg/L for bedaquiline and 0.50 mg/L for linezolid. Conclusions: REMA is reproducible and robust for the determination of bedaquiline and linezolid MIC distributions and ECOFF values when applied in laboratories of medium/low-resource countries. We suggest that WT MIC distributions for both drugs should be used as a monitoring tool to control the possible rapid emergence of resistance.

String test: A new tool for tuberculosis diagnosis and drug-resistance detection in children.

Background: There is a critical need to improve the diagnostic accuracy of tuberculosis (TB) in children. Several techniques have been developed to improve the quality of sputum samples; however, these procedures are very unpleasant and invasive and require hospitalization and trained personnel. This study aims to explore the potential use of a new and noninvasive tool, "string test," for TB diagnosis in children and in adults not able to render sputum samples and at risk of developing multidrug-resistant TB (MDR-TB). Methods: Children with clinical suspicion of TB attending the pediatric consultation at the Cetrangolo or Cordero Hospitals and adults suspected of MDR-TB and unable to produce sputum attending the Infectious Disease Unit of Cetrangolo Hospital were included in this study. Subjects and Methods: The "string test" is a string that is swallowed by the patients and exposed to gastrointestinal secretions that were late analyzed for TB diagnosis and drug-resistance detection by GenoType MTBDRplus. MedCalc software was used to perform statistical analysis. Results: This technique could be applied on 62.1% of selected children. About 11 (30.6%) children were diagnosed as TB cases, 8 (22.2%) from gastric aspirate and using the "string test." Six out of 19 adults were also diagnosed. Genotype directly on the string specimen detected two MDR-TB in adults and two isoniazid-resistant cases before obtaining the isolate. Conclusion: This test was safe, cheap, and easily implemented without requiring hospitalization. This research could represent a significant step forward to diagnose and rapidly detect drug-resistant TB in children.

Antimicrobial activity against Mycobacterium tuberculosis under in vitro lipid-rich dormancy conditions.

Although tuberculosis treatment is dependent on drug-susceptibility testing (DST) and molecular drug-resistance detection, treatment failure and relapse remain a challenge. This could be partially due to the emergence of antibiotic-tolerant dormant mycobacteria, where host lipids have been shown to play an important role. This study evaluated the susceptibility of Mycobacterium tuberculosis to two antibiotic combinations - rifampicin, moxifloxacin, amikacin and metronidazole (RIF-MXF-AMK-MTZ), and rifampicin, moxifloxacin, amikacin and pretomanid (RIF-MXF-AMK-PA) - in a lipid-rich dormancy model. Although their effectiveness in in vitro cultures with dextrose as a carbon source has been proved, we observed that none of the antibiotic mixtures were bactericidal in the presence of lipids. The presence of lipids may confer tolerance to M. tuberculosis against the mixture of antibiotics tested and such tolerance could be even higher during the dormant stages. The implementation of lipids in DST on clinical isolates could potentially lead to a better treatment strategy.

Performance of a highly successful outbreak strain of Mycobacterium tuberculosis in a multifaceted approach to bacterial fitness assessment.

Determining bacterial fitness represents a major challenge and no single parameter can accurately predict the ability of a certain pathogen to succeed. The M strain of Mycobacterium tuberculosis managed to spread and establish in the community and caused the largest multidrug-resistant tuberculosis outbreak in Latin America. We have previously shown that the M strain can manipulate the host immune response, but we still have no direct evidence, other than epidemiology, that can account for the enhanced fitness of the M strain. Our objective was to further characterize the performance of the outbreak strain M in different fitness assays. Two main aspects were evaluated: (1) molecular characterization of selected isolates from the M outbreak and related strains and (2) comparative fitness and in vivo performance of representative M strain isolates vs. the non-prosperous M strain variant 410. Our approach confirmed the multifaceted nature of fitness. Altogether, we conclude that the epidemiologically abortive strain 410 was vulnerable to drug-driven pressure, a weak competitor, and a stronger inductor of protective response in vivo. Conversely, the isolate 6548, representative of the M outbreak peak, had a growth disadvantage but performed very well in competition and induced lung damage at advanced stages in spite of reaching relatively low CFU counts. Integration of these observations supports the idea that the M strain managed to find a unique path to success.

Validación de la batería de instrumentos para la evaluación de factores de riesgo psicosocial / Validation of the battery of instruments for the evaluation of psychosocial risk factors

Objetivo: El presente estudio tuvo el objetivo de adaptar y validar los resultados de la Batería de Instrumentos para la Evaluación de Factores de Riesgo Psicosocial desarrollado por el Ministerio de la Protección Social de Colombia en el 2010. Materiales y métodos: Se realizó un estudio instrumental con los trabajadores que pasaron examen médico ocupacional de marzo a setiembre 2014, con un total de 478 personas. Se procedió al análisis de propiedades psicométricas mediante criterio de jueces, análisis de validez mediante el coeficiente V de Aiken, análisis de validez de constructo mediante análisis factorial de tipo exploratorios y el análisis de confiabilidad mediante la consistencia interna con el coeficiente Alpha de Cronbach. Resultados: Como resultados del análisis de la validez de contenido, al aplicar el criterio de jueces, se consideraron solo aquellos ítems con V = 1 (p = 0,032), quedando en la Forma A, 108 reactivos, en la Forma B, 96 y en Extralaborales, 29 ítems. Se evidenciaron índices de homogeneidad de los ítems mayor al criterio (rit > 0,20) e índices adecuados de confiabilidad por el método de consistencia interna con el estadístico Alfa de Cronbach en las escalas Intralaboral Forma A, Forma B, Extralaboral y Estrés. Para el grado de validez convergente, se obtuvieron correlaciones estadísticamente significativas en la mayoría de dimensiones entre las escalas Extralaboral y Estrés, y entre la Forma A y Estrés. Conclusiones: Se determinó la validez de criterio y validez convergente en las escalas Intralaboral A, Intralaboral B y Extralaboral, encontrándose evidencias empíricas de mediciones coherentes al constructo de Riesgos Psicosociales; asimismo, se calcularon los coeficientes de consistencia interna, obteniéndose en las dimensiones evaluadas adecuados índices de acuerdo al criterio mínimo sugerido para la investigación

Objective: This study aimed to adapt and validate the results of the Batería de Instrumentos para la Evaluación de Factores de Riesgo Psicosocial (Battery of Instruments for the Evaluation of Psychosocial Risk Factors) developed by the Ministry of Health and Social Protection of Colombia in 2010. Materials and methods: An instrumental study was conducted with a total of 478 workers who underwent an occupational medical examination from March to September 2014. The following analyses were performed: psychometric properties by judges' criteria, validity by Aiken's V coefficient, construct validity by exploratory factor analysis, and reliability and internal consistency by Cronbach's Alpha. Results: In the content validity analysis using judges' criteria, only items with V = 1 (p = 0.032) were considered, resulting in 108 items in the Inside-Work scale form A, 96 items in the Inside-Work scale form B, and 29 items in the Outside-Work scale. A higher-than-standard item homogeneity level (rit > 0.20), and adequate reliability and internal consistency levels were demonstrated by Cronbach's Alpha in the Inside-Work scale form A, Inside-Work scale form B, Outside-Work scale and Stress scale. In the convergent validity analysis, statistical significant correlations were obtained in most of the dimensions between the Outside-Work scale and Stress scale, and the Inside-Work scale form A and Stress scale. Conclusions: Criteria validity and convergent validity were determined in the Inside-Work scale form A, Inside-Work scale form B and Outside-Work scale, with empiric evidence of measurements consistent with the psychosocial risk construct being found. Moreover, internal consistency coefficients were estimated, with adequate levels being obtained in the assessed dimensions, according to the minimum level suggested for the research

The transcriptome of Mycobacterium tuberculosis in a lipid-rich dormancy model through RNAseq analysis.

Tuberculosis (TB) is currently the number one killer among infectious diseases worldwide. Lipids are abundant molecules during the infectious cycle of Mycobacterium tuberculosis (Mtb) and studies better mimicking its actual metabolic state during pathogenesis are needed. Though most studies have focused on the mycobacterial lipid metabolism under standard culture conditions, little is known about the transcriptome of Mtb in a lipid environment. Here we determined the transcriptome of Mtb H37Rv in a lipid-rich environment (cholesterol and fatty acid) under aerobic and hypoxic conditions, using RNAseq. Lipids significantly induced the expression of 368 genes. A main core lipid response was observed involving efflux systems, iron caption and sulfur reduction. In co-expression with ncRNAs and other genes discussed below, may act coordinately to prepare the machinery conferring drug tolerance and increasing a persistent population. Our findings could be useful to tag relevant pathways for the development of new drugs, vaccines and new strategies to control TB.

In vitro activity of bedaquiline against rapidly growing nontuberculous mycobacteria.

Bedaquiline (BDQ) has been proven to be effective in the treatment of multidrug-resistant tuberculosis. We hypothesized that BDQ could be a potential agent to treat nontuberculous mycobacterial (NTM) infection. The objective of this study was to evaluate the in vitro activity of BDQ against rapidly growing mycobacteria by assessing the minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) against 18 NTM strains. For MIC determination we performed the resazurin microtitre assay broth dilution, and for the MBC the c.f.u. was determined. BDQ exhibited a strong inhibitory effect against most NTM tested; however, for some NTM strains the MBC was significantly higher than the MIC. A new finding is that Mycobacterium flavescens has a mutation in the gene atpE associated with natural resistance to BDQ. These preliminary promising results demonstrate that BDQ could be potentially useful for the treatment of NTM.

Exploring tuberculosis by molecular tests on DNA isolated from smear microscopy slides.

Tuberculosis (TB) is an infectious disease of global public health importance caused by Mycobacterium tuberculosis complex. The disease has worsened with the emergence of multidrug-resistant (MDR)-TB strains. The timely diagnosis and treatment of TB remains a key public health priority, and laboratories have a critical role in the rapid and accurate detection of TB and drug resistance. Molecular assays based on nucleic acid amplification techniques have been developed for the rapid, sensitive, and specific diagnosis of TB, with the ability to determine the drug sensitivity status. These molecular techniques are now available or are being implemented in developing countries. However, traditional microscopy and culture methods cannot yet be replaced; the molecular assays can be applied in parallel with these tests for the diagnosis of TB or for drug susceptibility testing. Performing such molecular tests is often restricted by constraints with regard to sputum sample storage and safe transportation from remote health centres to central laboratories. Since smear slides are performed routinely for the diagnosis of TB in most TB diagnostic laboratories, they are readily available and could be the ideal tool to transport sputum for further molecular tests. The aim of this review was to provide a comprehensive survey on the use of smear slides for both TB diagnosis and the molecular test approach. Based on the literature, stained smear microscopy slides can be a safe system for the transportation of sputum specimens from remote health centres to reference TB laboratories for further molecular TB or MDR-TB detection, and could help in the rapid diagnosis and therefore timely management of TB patients.

"Genetic regulation of Mycobacterium tuberculosis in a lipid-rich environment".

Tuberculosis (TB) remains as one of the leading causes of morbidity and mortality among infectious diseases worldwide. Although lipids (mainly fatty acids and cholesterol) have been reported to play an important role during active and latent infection of M. tuberculosis, there are other molecular aspects of bacterial response to those substrates that are not fully understood, involving gene regulation background. This review highlights recent insights on pathogen gene expression: regulation during its active growth, during survival in presence of lipids and under variable hostile host microenvironments. We also propose several application options of this knowledge that may contribute for improved TB control.

Optimizing of a protein extraction method for Mycobacterium tuberculosis proteome analysis using mass spectrometry.

A critical step in proteomic analyses comprises the implementation of a reliable cell lysis method with high yields of qualitative proteins. In Mycobacteria, the protein extraction step is often hampered by the thick waxy cell wall which is rich in mycolic acids. Harsh disruption techniques to release proteins from the cells are thus required. Here, we demonstrate an optimized protein extraction procedure for Mycobacterium tuberculosis (Mbt) that results in protein extracts that are useful for all currently used proteomics platforms, including gel and LC-MS based strategies. We compared the effectiveness of using both thiourea and urea and/or SDS and DTT in the solubilization buffer, in combination or not with sonication and/or bead beating. After some preliminary optimization steps on fast-growing Mbt-like organisms, namely Mycobacterium smegmatis and Mycobacterium fortuitum, the final protein extraction protocol was tested on M. tuberculosis. Based on the concentrations of the proteins recovered from each of the tested methods and on the quality of the extracted proteins as evaluated by SDS PAGE, we propose a lysis buffer that contains both thiourea and urea, in combination with two mechanical cell disruption methods: sonication and bead beating. The optimized protocol results in protein extracts that are useful in M. tuberculosis proteomics studies based on any proteomics strategy or platform.

The potential role of trimethoprim-sulfamethoxazole in the treatment of drug-resistant tuberculosis.

Tuberculosis (TB) remains a serious public health threat worsened by emerging drug resistance. Mycobacterium tuberculosis has become resistant not only to front-line drugs but also to second-line antimicrobials directed at drug-resistant TB. Renewed efforts are devoted for the development of new antibiotics active against TB. Also, repurposing of other antibiotics is being explored to shorten the time to develop new drugs against M. tuberculosis. As a result, trimethoprim-sulfamethoxazole (SXT) has emerged as a potential new option to treat drug-resistant TB. SXT has been found to be surprisingly active against drug-resistant M. tuberculosis, not only in vitro but also in vivo. The potential role of SXT for the treatment of multidrug resistant/extensively drug resistant TB might be explored in further clinical evaluations.

Performance of Four Transport and Storage Systems for Molecular Detection of Multidrug-Resistant Tuberculosis.

BACKGROUND: Detection of drug-resistant tuberculosis is essential for the control of the disease but it is often hampered by the limitation of transport and storage of samples from remote locations to the reference laboratory. We performed a retrospective field study to evaluate the performance of four supports enabling the transport and storage of samples to be used for molecular detection of drug resistance using the GenoType MTBDRplus. METHODS: Two hundred Mycobacterium tuberculosis strains were selected and spotted on slides, FTA cards, GenoCards, and in ethanol. GenoType MTBDRplus was subsequently performed with the DNA extracted from these supports. Sensitivity and specificity were calculated and compared to the results obtained by drug susceptibility testing. RESULTS: For all supports, the overall sensitivity and specificity for detection of resistance to RIF was between 95% and 100%, and for INH between 95% and 98%. CONCLUSION: The four transport and storage supports showed a good sensitivity and specificity for the detection of resistance to RIF and INH in M. tuberculosis strains using the GenoType MTBDRplus. These supports can be maintained at room temperature and could represent an important alternative cost-effective method useful for rapid molecular detection of drug-resistant TB in low-resource settings.

Mycobacterium saopaulense sp. nov., a rapidly growing mycobacterium closely related to members of the Mycobacterium chelonae--Mycobacterium abscessus group.

Five isolates of non-pigmented, rapidly growing mycobacteria were isolated from three patients and,in an earlier study, from zebrafish. Phenotypic and molecular tests confirmed that these isolates belong to the Mycobacterium chelonae-Mycobacterium abscessus group, but they could not be confidently assigned to any known species of this group. Phenotypic analysis and biochemical tests were not helpful for distinguishing these isolates from other members of the M. chelonae­M.abscessus group. The isolates presented higher drug resistance in comparison with other members of the group, showing susceptibility only to clarithromycin. The five isolates showed a unique PCR restriction analysis pattern of the hsp65 gene, 100 % similarity in 16S rRNA gene and hsp65 sequences and 1-2 nt differences in rpoB and internal transcribed spacer (ITS) sequences.Phylogenetic analysis of a concatenated dataset including 16S rRNA gene, hsp65, and rpoB sequences from type strains of more closely related species placed the five isolates together, as a distinct lineage from previously described species, suggesting a sister relationship to a group consisting of M. chelonae, Mycobacterium salmoniphilum, Mycobacterium franklinii and Mycobacterium immunogenum. DNA­DNA hybridization values .70 % confirmed that the five isolates belong to the same species, while values ,70 % between one of the isolates and the type strains of M. chelonae and M. abscessus confirmed that the isolates belong to a distinct species. The polyphasic characterization of these isolates, supported by DNA­DNA hybridization results,demonstrated that they share characteristics with M. chelonae­M. abscessus members, butconstitute a different species, for which the name Mycobacterium saopaulense sp. nov. is proposed. The type strain is EPM10906T (5CCUG 66554T5LMG 28586T5INCQS 0733T).

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

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

Utilidad del método de la nitrato reductasa para la detección de resistencia a drogas antituberculosas de segunda línea / Usefulness of the nitrate reductase method to detect resistance to second-line anti-tuberculosis drugs

Introducción: el creciente hallazgo de cepas de Mycobacterium tuberculosis multidrogorresistentes extremadamente resistentes ratifica la importancia de ofrecer, de forma rápida, los resultados de susceptibilidad de M. tuberculosis a drogas de primera y segunda línea como única alternativa para evitar la transmisión. Objetivo: comparar el método de la nitrato reductasa y el de las proporciones para la detección de susceptibilidad a drogas antituberculosas de segunda línea en aislamientos clínicos de M. tuberculosis, recuperados de pacientes cubanos con tuberculosis multidrogorresistente. Métodos: se investigó, mediante el método de las proporciones en Löwenstein-Jensen y el de la nitrato reductasa, la susceptibilidad a la ofloxacina, la kanamicina y a la capreomicina en 34 aislamientos de M. tuberculosis multidrogorresistentes. Resultados: en tres aislamientos se evidenció un comportamiento extremadamente resistente por ambos métodos. Mediante el método de la nitrato reductasa los resultados estuvieron disponibles entre 7 y 14 días. La sensibilidad fue de 100 por ciento, 90,0 por ciento y 77,8 por ciento para la ofloxacina, la kanamicina y la capreomicina, respectivamente, mientras que la especificidad fue superior al 95,0 por ciento y el valor de kappa fue superior a 0,85 para las tres drogas. Conclusión: de acuerdo con los resultados alcanzados, consideramos que el método de la nitrato reductasa constituye una valiosa alternativa para la detección oportuna de tuberculosis extremadamente resistente en países con limitados recursos económicos(AU)

Introduction: The increase of multidrug resistant and extensively drug resistant tuberculosis underlines the urgent need to obtain early results of Mycobacterium tuberculosis susceptibility both to first and second line antituberculosis drugs in order to avoid dissemination of resistant isolates. Objective: The aim of this research was to compare the performance of the nitrate reductase assay and the proportion method for to detect the susceptibility to second line antituberculosis drugs in multidrug resistant clinical isolates of M. tuberculosis. Methods: The susceptibility to ofloxacin, kamamycin and capreomycin of 34 M. tuberculosis multidrug resistant isolates was investigated using the proportion method in Löwenstein-Jensen and the nitrate reductase assay. Results: Three isolates were identified as extensively drug resistant by both methods. The results of the nitrate reductase assay were obtained between 7-14 days achieving 100 percent, 90.0 percent and 77.8 percent of sensitivity for ofloxacin, kamamycin and capreomycin, respectively while specificity was higher than 95.0 percent and kappa value was higher to 0,85 for all drugs. Conclusion: The nitrate reductase assay represents a useful tool for the rapid identification of extensively drug resistant tuberculosis in low resources setting(AU)

Utilidad del método de la nitrato reductasa para la detección de resistencia a drogas antituberculosas de segunda línea / Usefulness of the nitrate reductase method to detect resistance to second-line anti-tuberculosis drugs

Introducción: el creciente hallazgo de cepas de Mycobacterium tuberculosis multidrogorresistentes extremadamente resistentes ratifica la importancia de ofrecer, de forma rápida, los resultados de susceptibilidad de M. tuberculosis a drogas de primera y segunda línea como única alternativa para evitar la transmisión. Objetivo: comparar el método de la nitrato reductasa y el de las proporciones para la detección de susceptibilidad a drogas antituberculosas de segunda línea en aislamientos clínicos de M. tuberculosis, recuperados de pacientes cubanos con tuberculosis multidrogorresistente. Métodos: se investigó, mediante el método de las proporciones en Löwenstein-Jensen y el de la nitrato reductasa, la susceptibilidad a la ofloxacina, la kanamicina y a la capreomicina en 34 aislamientos de M. tuberculosis multidrogorresistentes. Resultados: en tres aislamientos se evidenció un comportamiento extremadamente resistente por ambos métodos. Mediante el método de la nitrato reductasa los resultados estuvieron disponibles entre 7 y 14 días. La sensibilidad fue de 100 por ciento, 90,0 por ciento y 77,8 por ciento para la ofloxacina, la kanamicina y la capreomicina, respectivamente, mientras que la especificidad fue superior al 95,0 por ciento y el valor de kappa fue superior a 0,85 para las tres drogas. Conclusión: de acuerdo con los resultados alcanzados, consideramos que el método de la nitrato reductasa constituye una valiosa alternativa para la detección oportuna de tuberculosis extremadamente resistente en países con limitados recursos económicos(AU)

Introduction: the increase of multidrug resistant and extensively drug resistant tuberculosis underlines the urgent need to obtain early results of Mycobacterium tuberculosis susceptibility both to first and second line antituberculosis drugs in order to avoid dissemination of resistant isolates. Objective: the aim of this research was to compare the performance of the nitrate reductase assay and the proportion method for to detect the susceptibility to second line antituberculosis drugs in multidrug resistant clinical isolates of M. tuberculosis. Methods: the susceptibility to ofloxacin, kamamycin and capreomycin of 34 M. tuberculosis multidrug resistant isolates was investigated using the proportion method in Löwenstein-Jensen and the nitrate reductase assay. Results: three isolates were identified as extensively drug resistant by both methods. The results of the nitrate reductase assay were obtained between 7-14 days achieving 100 percent, 90.0 percent and 77.8 percent of sensitivity for ofloxacin, kamamycin and capreomycin, respectively while specificity was higher than 95.0 percent and kappa value was higher to 0,85 for all drugs. Conclusion: the nitrate reductase assay represents a useful tool for the rapid identification of extensively drug resistant tuberculosis in low resources setting(AU)

Evaluation of the Speed-Oligo Mycobacteria assay for the identification of nontuberculous mycobacteria.

Nontuberculous mycobacteria (NTM) causing human infectious disease have become increasingly common. Rapid and accurate identification to the species level is, therefore, critical. The Speed-Oligo Mycobacteria assay is an oligochromatographic method that was made available recently for the identification and differentiation of mycobacteria. The present study aimed to evaluate the performance of the Speed-Oligo Mycobacteria assay for the identification of NTM. We examined a total of 62 strains (9 type strains, 19 reference strains and 34 clinical isolates) belonging to 13 different species (Mycobacterium intracellulare, M. fortuitum, M. gordonae, M. kansasii, M. marinum, M. peregrinum, M. scrofulaceum, M. abscessus, M. bovis BCG, M. chelonae, M. avium, M. malmoense and M. xenopi). The Speed-Oligo Mycobacteria assay was performed according to the manufacturer's instructions. Discrepant results between Speed-Oligo Mycobacteria and the original identification were reassessed by the Speed-Oligo Mycobacteria assay and resolved by the GenoType Mycobacterium CM assay and by sequencing of 16S rRNA and protein-encoding genes. We found 93.5 % (58/62) concordance for the identification of NTM as compared with the original identification. Three strains were erroneously identified by Speed-Oligo Mycobacteria: one M. kansasii strain was identified as Mycobacterium tuberculosis complex, and one M. chelonae strain and one M. peregrinum strain were both identified as Mycobacterium abscessus. Moreover, one M. chelonae strain was not identified by Speed-Oligo Mycobacteria since it did not react with any species-specific probe. For these strains, sequencing of the genes hsp65, 16S rRNA and rpoB and the GenoType Mycobacterium CM assay were performed. The Speed-Oligo Mycobacteria assay can be a useful tool for the rapid and easy identification of the most common NTM. If applied in clinical practice it could reduce diagnostic delays and contribute to correct clinical and better management of infections caused by NTM.