Impact of Xpert MTB/RIF implementation in tuberculosis case detection and control in Brazil: a nationwide intervention time-series analysis (2011-2022).

Background: Since 2014, Brazil has gradually implemented the Xpert MTB/RIF (Xpert) test to enhance early tuberculosis (TB) and drug-resistant (DR-TB) detection and control, yet its nationwide impact remains underexplored. Our study conducts an intervention time-series analysis (ITSA) to evaluate how the Xpert's implementation has improved TB and DR-TB detection nationwide. Methods: 1,061,776 cases from Brazil's National TB Registry (2011-2022) were reviewed and ITSA (2011-2019) was used to gauge the impact of the Xpert's adoption on TB and DR-TB notification. Granger Causality and dynamic regression modelling determined if incorporating Xpert testing as an external regressor enhanced forecasting accuracy for Brazil's future TB trends. Findings: Xpert implementation resulted in a 9.7% increase in TB notification and substantial improvements in DR-TB (63.6%) and drug-susceptible TB (92.1%) detection compared to expected notifications if it had not been implemented. Xpert testing counts also presented a time-dependent relationship with DR-TB detection post-implementation, and improved predictions in forecasting models, which depicted a potential increase in TB and DR-TB detection in the next six years. Interpretation: This study underscores the critical role of Xpert's adoption in boosting TB and DR-TB detection in Brazil, reinforcing the case for its widespread use in disease control. Improvements in prediction accuracy resulting from integrating Xpert data are crucial for allocating resources and reducing the incidence of TB. By acknowledging Xpert's role in both disease control and improving predictions, we advocate for its expanded use and further research into advanced molecular diagnostics for effective TB and DR-TB control. Funding: FIOCRUZ.

How has the municipal availability of the GeneXpert®MTB/RIF system affected the detection of drug-resistant tuberculosis in Brazil?

OBJECTIVE: To evaluate the association between the availability of GeneXpert®MTB/RIF in municipalities and the proportion of people who have access to this diagnostic technology for tuberculosis (TB), as well as the resistance detected by the surveillance system in Brazil. METHODS: We analysed 4998 Brazilian municipalities that reported 432,937 new TB cases between 2015 and 2020. We compared municipalities with and without the availability of GeneXpert®MTB/RIF regarding the effective access to GeneXpert®MTB/RIF diagnosis and the prevalence of detected resistance. RESULTS: Municipalities with at least one GeneXpert®MTB/RIF system had three times (95% CI 2.9-3.0) the access to diagnostic tests and 80.4% (95% CI 70.6%-90.2%) higher detection of resistance, compared with municipalities without this technology. We estimated that there have been 1890 cases of undetected resistance during this period in the country. CONCLUSIONS: The availability of GeneXpert®MTB/RIF system in the municipality increased the sensitivity of the surveillance for detecting TB resistance. PUBLIC HEALTH IMPLICATIONS: It is a priority to strengthen laboratory networks and narrow the gap in access to rapid diagnosis in remote areas to improve the detection and control of drug-resistant tuberculosis.

Assessing Polymorphic Purity of Rifampicin in Double and Triple-Drug Fixed-Dose Combination Products.

First-line tuberculostatic agents, Rifampicin (RIF), Isoniazid (ISH), Ethambutol (ETB), and Pyrazinamide (PZA) are generally administered as a fixed-dose combination (FDC) for improving patient adherence. The major quality challenge of these FDC products is their variable bioavailability, where RIF and its solid state are key factors. In this work, the analysis of the impact of the polymorphism in the performance of RIF in RIF-ISH and PZA-RIF-ISH combined products was carried out by an overall approach that included the development and validation of two methodologies combining near-infrared (NIR) spectroscopy and partial least squares (PLS) to the further evaluation of commercial products. For NIR-PLS methods, training and validation sets were prepared with mixtures of Form I/Form II of RIF, and the appropriate amount of ISH (for double associations) or ISH-PZA (for triple associations). The corresponding matrix of the excipients was added to the mixture of APIs to simulate the environment of each FDC product. Four PLS factors, reduced spectral range, and the combination of standard normal variate and Savitzky-Golay 1st derivative (SNV-D') were selected as optimum data pre-treatment for both methods, yielding satisfactory recoveries during the analysis of validation sets (98.5±2.0%, and 98.7±1.8% for double- and triple-FDC products, respectively). The NIR-PLS model for RIF-ISH successfully estimated the polymorphic purity of Form II in double-FDC capsules (1.02 ± 0.02w/w). On the other hand, the NIR-PLS model for RIF-ISH-PZA detected a low purity of Form II in triple FDC tablets (0.800 ± 0.021w/w), these results were confirmed by X-ray powder diffraction. Nevertheless, the triple-FDC tablets showed good performance in the dissolution test (Q=99-102%), implying a Form II purity about of 80% is not low enough to affect the safety and efficacy of the product.

The Role of GeneXpert® for Tuberculosis Diagnostics in Brazil: An Examination from a Historical and Epidemiological Perspective.

The rapid molecular test (RMT) performed on the GeneXpert® system is widely used as a control strategy and surveillance technique for tuberculosis (TB). In the region of the Americas, TB incidence is slowly increasing owing to an upward trend in Brazil, which is among the high TB-burden countries (HBCs), ranking in the 19th position. In this context, we aimed to (i) describe the implementation and history of RMT-TB (Xpert® MTB/RIF and Xpert® MTB/RIF Ultra) in Brazil; (ii) to evaluate the national RMT laboratory distribution, TB, and resistance to RIF detection by RMT; and (iii) to correlate these data with Brazilian TB incidence. The quantitative data of Xpert® MTB/RIF and Xpert® MTB/RIF Ultra assays performed in the pulmonary TB investigation from 2014 to 2020 were provided by the Brazilian Ministry of Health. A spatial visualization using ArcGIS software was performed. The Southeast region constituted about half of the RMT laboratories-from 39.4% to 45.9% of the total value over the five regions. Regarding the federal units, the São Paulo state alone represented from 20.2% to 34.1% (5.0 to 8.5 times the value) of RMT laboratories over the years observed. There were significant differences (p < 0.0001) in the frequency of RMT laboratories between all years of the historical series. There was an unequal distribution of RMT laboratories between Brazilian regions and federal units. This alerts us for the surveillance of rapid molecular detection of TB in different parts of the country, with the possibility of improving the distribution of tests in areas of higher incidence in order to achieve the level of disease control recommended by national and worldwide authorities.

Investigação da presença do bacilo Mycobacterium tuberculosis em amostras sugestivas de tuberculose oral / Investigation of the presence of Mycobacterium tuberculosis bacillus in samples suggestive of oral tuberculosis

A Tuberculose (TB) é um considerável problema de saúde pública mundial. Em 2021, de acordo com a Organização Mundial de Saúde (OMS) estimou-se que, no mundo, cerca de 10,6 milhões de pessoas desenvolveram TB e 1,4 milhão morreu devido à doença. Com isso, tornou-se a principal causa de morte por infecção em todo o mundo e uma das dez principais causas de morte em geral. A TB tem o pulmão como o principal sítio de acometimento, sendo denominada de TB Pulmonar (TBP). Porém pode ser diagnosticada em muitos órgãos do corpo de maneira Extrapulmonar (TBEP), sendo o linfonodo o local mais comum. Porém, o envolvimento pleural, neurológico, sinovial, pericárdico, abdominal, geniturinário e oral tem sido descrito, o que mostra a potencial capacidade de disseminação do Mycobacterium tuberculosis (MTB). A detecção do Bacilos Álcool-Ácido Resistentes (BAAR), geralmente ocorre pela observação das características microscópicas da morfologia dos tecidos, presença de granulomas com necrose caseosa, histiócitos epitelióides e células gigantes do tipo Langhans, associada à coloração para BAAR, pela técnica de Ziehl Neelsen (ZN). Ademais, investigação por imuno-histoquímica (IHQ), testes de amplificação de ácido nucleico pela Reação em Cadeia da Polimerase Hemianinhada (nested-PCR) e pelo sistema de detecção automatizado GeneXpert® MTB/RIF são métodos aplicados para o diagnóstico da infecção. Com isso, este estudo teve como objetivo investigar a presença do bacilo Mycobacterium tuberculosis em amostras orais em parafina que continham granulomas com necrose caseosa. Ao todo, como critério de inclusão, foram selecionadas biópsias que apresentaram granulomas com necrose caseosa, sugerindo o diagnóstico de TB. Foram excluídas aquelas que após a revisão das fichas e histológicas, não apresentavam os granulomas exibindo necrose caseosa e aquelas que foram de biópsias intraósseas. O M. tuberculosis foi procurado por meio da coloração de ZN, IHC, nested-PCR e ensaios GeneXpert® MTB/RIF. Foram então selecionadas nove amostras com granulomas com necrose caseosa. Houve predominância de indivíduos do sexo masculino (2,5:1), com idade média de 50 anos (±23,08; 19-89), sendo a língua o local anatômico mais afetado (n=4). O bacilo não foi identificado pela técnica de ZN em nenhuma amostra, e a coloração por IHC mostrou um padrão granular grosseiro, sugerindo M. tuberculosis, em três delas. Nested-PCR e os ensaios GeneXpert® MTB/RIF foram positivos em duas e três das amostras, respectivamente. Conclui-se que testes moleculares e IHC podem ser métodos auxiliares úteis para casos suspeitos de tuberculose.

Tuberculosis (TB) is a significant global public health issue. In 2021, according to the World Health Organization (WHO), it was estimated that approximately 10.6 million people developed TB worldwide, and 1.4 million died from the disease. Consequently, it became the leading cause of death due to infection worldwide and one of the top ten overall causes of death. TB primarily affects the lungs and is referred to as Pulmonary TB (PTB). However, it can be diagnosed in various organs of the body as Extrapulmonary TB (EPTB), with lymph nodes being the most common site of involvement. Moreover, pleural, neurological, synovial, pericardial, abdominal, genitourinary, and oral involvement have been described, demonstrating the potential for Mycobacterium tuberculosis (MTB) dissemination. The detection of Acid-Fast Bacilli (AFB) typically involves the observation of microscopic tissue characteristics, the presence of granulomas with caseous necrosis, epithelioid histiocytes, and Langhans giant cells, along with AFB staining using the Ziehl-Neelsen (ZN) technique. Furthermore, immunohistochemistry (IHC), nucleic acid amplification tests by Nested Polymerase Chain Reaction (nested-PCR), and the automated detection system GeneXpert® MTB/RIF are methods employed for diagnosing the infection. Therefore, the aim of this study was to investigate the presence of Mycobacterium tuberculosis in paraffin-embedded oral samples containing granulomas with caseous necrosis. Inclusion criteria were based on the selection of biopsies displaying granulomas with caseous necrosis, suggesting a diagnosis of TB. Biopsies without these features upon review of records and histological findings, as well as intraosseous biopsies, were excluded. M. tuberculosis was sought using ZN staining, IHC, nested-PCR, and GeneXpert® MTB/RIF assays. Nine samples with granulomas and caseous necrosis were selected. The majority of individuals were male (2.5:1 ratio), with an average age of 50 years (±23.08; range 19-89), and the tongue was the most affected anatomical site (n=4). AFB was not identified by the ZN technique in any of the samples, and IHC staining exhibited a coarse granular pattern, suggestive of M. tuberculosis, in three of them. Nested-PCR and GeneXpert® MTB/RIF assays yielded positive results in two and three of the samples, respectively. In conclusion, molecular tests and IHC can be valuable auxiliary methods for suspected cases of tuberculosis.

Linkage to care and treatment outcomes for patients diagnosed with drug-susceptible tuberculosis using Xpert MTB/RIF assay in Thaba-Tseka district in Lesotho.

Objectives: To evaluate linkage to care and treatment outcomes of patients with tuberculosis (TB) confirmed by Xpert MTB/RIF assay in Thaba-Tseka district, Lesotho. Design: This was a retrospective cohort study of adult patients diagnosed with drug-susceptible TB using the Xpert MTB/RIF assay at two laboratories in Thaba-Tseka district from January 2016 to December 2020. Results: Six hundred and fifty-five eligible participants were identified for inclusion in this study. Their median age was 40 [interquartile range (IQR) 32-54] years, and 468 (71.45%) were male. Evidence of linkage to care was found for 459 (70.08%) participants, but there was no documentation on treatment initiation for 196 (29.92%) participants. The median time to treatment initiation was 0 days (same-day initiation) (IQR 0-4) and the treatment success rate was 86%. Treatment success was associated with negative sputum smear results after 2, 5 and 6 months (χ2, P<0.001). The overall mortality rate was 10%, with no trend of mortality reduction. Conclusion: There is a need to address the issue of linkage to care of patients diagnosed with TB in Thaba-Tseka district. Efforts should be made to reduce TB mortality in line with the World Health Organization's 'End TB strategy' target.

Contribución del uso de XPERT MTB/RIF y su costo-efectividad en el diagnóstico de tuberculosis pulmonar y la resistencia a rifampicina: una comparación con métodos diagnósticos no moleculares / Contribution of the use of xpert MTB/RIF and its cost-effectiveness in the diagnosis of pulmonary tuberculosis and resistance to rifampicin: a comparison with non molecular diagnostic methods

Resumen Introducción: La tuberculosis es un problema de salud pública; su control requiere diagnóstico temprano y tratamiento oportuno. Xpert MTB/RIF® es una tecno logía diagnóstica basada en PCR en tiempo real, detecta el Complejo Mycobacterium tuberculosis y la susceptibilidad a rifampicina. Objetivo: Determinar la contribución del Xpert MTB/RIF y su costo-efectividad en la detección de tuberculosis y la resistencia a rifampicina en muestras respirato rias al compararlo con métodos de diagnóstico no moleculares Materiales y Métodos: Se analizaron 1.574 muestras de pacientes con sospecha de tuberculosis pulmonar que fueron procesadas para microscopía con coloración fluorescente de auramina-rodamina, Xpert MTB/RIF y cultivo en BACTEC MGIT 960. Los resultados obtenidos se compararon entre los métodos no moleculares y los moleculares para la detección de M. tuberculosis y susceptibilidad a rifampicina y se realizó un análisis comparativo de costos y costo efectividad. Resultados: 19,2% de las muestras fueron positivas por alguna de las técnicas usadas. Xpert MTB/RIF detectó M. tuberculosis en 90,4% del total de muestras positivas con un índice Kappa de 0,77 (IC95%: 0,74-0,82) comparado con el cultivo. La resistencia a rifampicina por Xpert fue 8,1%, sensibilidad 94,1% (IC95%: 73,0-99,0%), especificidad 98,4% (IC95%: 95,5-99,5%) y Kappa de 0,88 (IC95%: 0,76-1,00). La razón incremental de costo efectividad (RICE) fue menor en Xpert MTB/RIF comparada con el cultivo. Conclusión: Xpert MTB/RIF es una prueba eficiente y costo efectiva en la detección de casos de M. tuberculosis en muestras pulmonares comparado con los mé todos de diagnóstico basados en cultivo, sin embargo y a diferencia del Xpert MTB/RIF, estos pueden aportar en el diagnóstico con el aislamiento de especies de micobacterias no tuberculosas y la susceptibilidad a isoniazida y otros medicamentos.

Abstract Introduction: Tuberculosis is a public health problem its control requires early diagnosis and timely treatment. Xpert MTB/RIF is a real-time PCR based diagnostic technology, detects the Mycobacterium tuberculosis complex and rifampicin resistance. Objective: To determine the contribution of Xpert MTB/RIF and its cost-effectiveness in the detection of potential positive cases for tuberculosis and resistance to rifampicin in respiratory samples comparatively with diagnostic non molecular methods Materials and Methods: From 2013 to 2015, 1.574 clinical samples of patients with suspected pulmonary tuberculosis were evaluated by smear microscopy using auramina-rodamina stain, Xpert and culture in liquid medium BACTEC MGIT 960®. Results: 19,2% of the samples were positive for any of the methods used, Xpert detected M. tuberculosis in 90,4% of the positive samples and the concordance between Xpert and cultures had a Kappa index of 0,71 (IC95%: 0,62-0,72). Xpert identified resistance to rifampicin in 8,1% of the clinical samples studied with a sensitivity 94.1% (IC95%: 73,0-99,0%), specificity 98,4% (IC95%: 95,5-99,5%) and Kappa index 0,88 (IC95%: 0,76-1,00). Xpert had an incremental cost effectiveness ratio lower than culture (RICE). Conclusion: Xpert MTB/Rif is efficient diagnostic technique and comparable with culture in cost effectiveness for pulmonary tuberculosis diagnosis. However, culture based methods, in contrast to Xpert, may allow the isolation and identification of non tuberculosis mycobacterial species and the possibility to perform susceptibility for other antituberculous drugs.

Prueba Xpert® MTB/RIF para el diagnóstico de Tuberculosis en el Instituto Médico La Floresta

La necesidad de un diagnóstico rápido, sensible y específico para tuberculosis es apremiante; se estiman 4 000 muertes cada día y aproximadamente 28 000 personas se contagian de esta enfermedad. La prueba Xpert®MTB/RIF consiste en un ensayo de diagnóstico in vitro de reacción en cadena de la polimerasa (PCR) en tiempo real automatizada, semicuantitativa, que en tan solo dos horas, permite detectar el ácido desoxirribonucleico (ADN) del complejo Mycobacterium tuberculosis (MTB) y mutaciones asociadas a la resistencia a rifampicina. Objetivo: Describir la experiencia del ensayo Xpert®MTB/ RIF y su valor diagnóstico en Dpto. de Microbiología del Instituto Médico La Floresta, desde abril 2012 hasta abril 2021. Métodos: Estudio descriptivo de muestras respiratorias y de otras procedencias, de pacientes con sospecha de tuberculosis, ensayadas por PCR a través del Xpert®MTB/RIF. Adicionalmente se realizó Ziehl-Neelsen y cultivo por el método de Ogawa Kudoh modificado, dependiendo del volumen de muestra recibida. Resultados: De 618 PCR realizadas, 58 muestras fueron positivas (9,4 %), 56 correspondieron a adultos y 2 a niños, 81 % se clasificaron como tuberculosis pulmonar y 19 % como extrapulmonar. A 37 se les realizó cultivo, de las cuales 10 no desarrollaron crecimiento, y a 33 adicionalmente Ziehl-Neelsen, de las cuales 12 presentaron baciloscopias negativas. En cinco de las muestras se detectó resistencia a rifampicina. Conclusiones: A través de la prueba Xpert®MTB/RIF fue posible detectar MTB en aquellas con baciloscopias negativas, en las que no desarrollaron crecimiento en el cultivo y permitió la rápida instauración de tratamiento en la tuberculosis multirresistente.

The rapid, sensitive, and specific diagnosis for tuberculosis is urgent; 4 000 deaths are estimated every day and approximately 28 000 people are infected with this disease. The Xpert®MTB/RIF test consists of an automated, semi-quantitative real-time polymerase chain reaction (PCR) in vitro diagnostic assay that, in just two hours, allows the detection of Mycobacterium tuberculosis complex (MTB) DNA and mutations associated with resistance to rifampicin. Objective: To describe the experience of the Xpert®MTB/RIF assay and its diagnostic value in the Department of Microbiology of the La Floresta Medical Institute, from April 2012 to April 2021. Methods: Descriptive study of respiratory samples and other sources, from patients with suspected of tuberculosis, assayed by PCR through Xpert®MTB/RIF. Additionally, Ziehl-Neelsen and culture were performed by the modified Ogawa Kudoh method, depending on the volume of sample received. Results: Of 618 PCR performed, 58 samples were positive (9.4 %), 56 corresponded to adults and 2 to children, 81 % were classified as pulmonary tuberculosis and 19 % as extrapulmonary. Culture was performed on 37, of which 10 did not develop growth, and on 33 additionally Ziehl-Neelsen, of which 12 presented negative bacilloscopy. Rifampicin resistance was detected in five of the samples. Conclusions: Through the Xpert®MTB/RIF test, it was possible to detect MTB in those with negative bacilloscopy, in which they did not develop growth in the culture and allowed the rapid establishment of treatment in multidrug-resistant tuberculosis.

Transcriptome Profile Reveals Genetic and Metabolic Mechanisms Related to Essential Fatty Acid Content of Intramuscular Longissimus thoracis in Nellore Cattle.

Beef is a source of essential fatty acids (EFA), linoleic (LA) and alpha-linolenic (ALA) acids, which protect against inflammatory and cardiovascular diseases in humans. However, the intramuscular EFA profile in cattle is a complex and polygenic trait. Thus, this study aimed to identify potential regulatory genes of the essential fatty acid profile in Longissimus thoracis of Nellore cattle finished in feedlot. Forty-four young bulls clustered in four groups of fifteen animals with extreme values for each FA were evaluated through differentially expressed genes (DEG) analysis and two co-expression methodologies (WGCNA and PCIT). We highlight the ECHS1, IVD, ASB5, and ERLIN1 genes and the TF NFIA, indicated in both FA. Moreover, we associate the NFYA, NFYB, PPARG, FASN, and FADS2 genes with LA, and the RORA and ELOVL5 genes with ALA. Furthermore, the functional enrichment analysis points out several terms related to FA metabolism. These findings contribute to our understanding of the genetic mechanisms underlying the beef EFA profile in Nellore cattle finished in feedlot.

Caracterización clínica y microbiológica de pacientes con trazas en Xpert MTB/RIF Ultra / Clinical and microbiological characterization of patients with Xpert MTB/RIF Ultra traces

INTRODUCCIÓN: El Xpert MTB/RIF Ultra (Ultra) ha mejorado dramáticamente el diagnóstico de la tuberculosis (TBC). Con él ha nacido la categoría de trazas, que es la menor carga bacilar detectable por este examen. OBJETIVO: Describir las características clínicas de los pacientes con presencia de trazas en el Ultra y evaluar la confirmación de la TBC como diagnóstico clínico. MATERIALES Y MÉTODOS: Estudio descriptivo de serie de casos. Se extrajo la información de fichas clínicas de pacientes con positividad a trazas. Se confrontaron datos clínicos, microbiológicos e histopatológicos. RESULTADOS: Se analizaron 21 pacientes. La edad promedio fue de 52 años. Todos los casos presentaron baciloscopias negativas. Cuatro cultivos en medio líquido MGIT fueron positivos, dos en pleura parietal, uno en líquido pleural y otro en expectoración. En pleura parietal, tres casos presentaron granulomas con necrosis caseosa y un granuloma esbozos de necrosis. En tejido pulmonar se observaron dos casos con granulomas con esbozos de necrosis y dos con granulomas no necrotizantes. Tres pacientes tenían el antecedente de TBC previa, se interpretó la positividad de trazas en ellos como falsos positivos. Finalmente se diagnosticaron 13 casos como TBC activa, donde cinco de ellos fueron TBC pleurales. La mayor concordancia clínica, microbiológica e histopatológica fue en muestras de líquido y tejido pleural. DISCUSIÓN: Se debe interpretar con cautela los hallazgos de esta prueba en muestras de vía aérea; el análisis multidisciplinario (clínica, imágenes, microbiología, histología) es crucial en las decisiones de nuestras conductas clínicas futuras. El hallazgo de trazas en pleura tiene, a nuestro parecer, un alto valor diagnóstico en el estudio de la tuberculosis en esta localización.

INTRODUCTION: Xpert MTB/RIF Ultra has dramatically changed the diagnosis of tuberculosis. A new category called traces appeared, which is the smallest amount of bacillar load detectable. OBJECTIVE: Describe the clinical characteristics of patients that present traces in Xpert MTB/RIF Ultra test, and to evaluate the confirmation of tuberculosis as clinical diagnosis. METHODS: We perform a descriptive case series study. Information was recovered from clinical records of patients with positive test for traces. Clinical, histopathological and microbiological results were confronted. RESULTS: Twenty one patients were analyzed. The mean age was 52 years-old. All cases had negative smear microscopy and four MGIT cultures were positive, two in pleural fluid and another in sputum. In parietal pleura, three cases presented granulomas with caseous necrosis, and one showed granuloma with very little necrosis. In pleural tissue we observed two cases of granulomas with traces of necrosis and two with non-necrotizing granulomas. Three patients had history of previous tuberculosis and positive traces, the test was interpreted as a false positive result. Finally, active tuberculosis was diagnosed in 13 cases, and five of them were pleural tuberculosis. The highest clinical, microbiological and histopathological agreement was in fluid and pleural tissue samples. DISCUSSION: The findings of Xpert MTB/RIF Ultra in airway samples must be interpreted carefully. Multi-disciplinary analysis is crucial in future clinical decisions. The finding of traces in pleura has, in our opinion, a high diagnostic value in the study of tuberculosis in this location.

Viability assessment of Mycobacterium tuberculosis complex in OMNIgene • SPUTUM reagent using the BACTEC MGIT 960 System and Xpert MTB/RIF assay.

The World Health Organization advocates that sputum specimens submitted to tuberculosis (TB) diagnostic should be processed within 48 h after collection and be stored under cooling. We aimed to assess the performance of OMNIgene • SPUTUM reagent in maintaining viable specimens of Mycobacterium tuberculosis complex (MTBC) during transportation of sputum samples without refrigeration, in comparison to the standard protocol of the National TB Control Program. Sputum samples obtained in southeastern Brazil (June 2017 to July 2018) from 100 sequential patients with positive acid-fast bacillus smear microscopy were divided into two portions. Portion 1 continued to be cooled (standard protocol, STA), but portion 2 was added to OMNIgene • SPUTUM reagent (alternative protocol, OMS) until concomitant further processing. Both portions of all samples were cultured using MGIT and tested by Xpert MTB/RIF assay. Growth of MTBC in the first 42 days was detected in 96% of the cultures under the STA and 88% under the OMS. Intervals between processing and detecting MTBC growth in the two portions significantly differed (p = 0.0001). Portions under the two protocols showed similar results in the MTBC detection by Xpert assay and culture contamination by non-MTBC. The OMNIgene reagent liquefies and decontaminates sputum leading to a decrease in processing time. Although there was a small delay in mycobacterial growth, the OMNIgene reagent can be useful in specimens transported from collection sites over a long distance to centralized testing centers, maintaining viable MTBC for at least 8 days at room temperature.

De la Hogra al Hirak: neocolonialismo, memoria y disidencia política juvenil en el Rif / From Hogra to Hirak: Neocolonialism, memory and youth political dissidence in the Rif / De Hogra para Hirak: Neocolonialismo, memória e dissidência política juvenis no Rif

Resumen (analítico) Las teorías sobre movimientos sociales dominantes basadas en experiencias occidentales, pero ¿en qué medida pueden ayudarnos a comprender el proceso de construcción solidaria de prácticas políticas en contextos no occidentales políticamente cerrados? Según Bayat parece imprescindible una aproximación teórica que tenga en cuenta tanto la excepcionalidad árabe como la aplicación crítica de los modelos de las ciencias sociales en otros contextos que, muchas veces, olvidan los rasgos específicos que la protesta política ha adoptado en el mundo árabe. Este artículo, en el marco del proyecto Transgang, analizará un movimiento social marroquí aparecido en el Rif en 2016, protagonizado por poblaciones en edad juvenil, atendiendo a sus especificidades culturales usando las perspectivas decoloniales. Con este análisis, se presentarán algunas conclusiones para la investigación sobre movimientos sociales juveniles en sociedades árabes.

Abstract (analytical) Dominant social movement theories are based on Western experiences, but to what extent can they help us to understand the process of solidarity construction of political practices in closed political non-Western contexts? As Bayat points out, a theoretical approach that takes into account both the Arab exceptionality and the critical application of social science models in other contexts -that often forget the specific features that political protest has adopted in the Arab World- seems essential. This article, within the framework of the Transgang project, will analyse a Moroccan social movement that appeared in the Rif in 2016, led by youth populations, attending to their cultural specificities, and using decolonial perspectives. Through this analysis, some conclusions will be presented for research on youth social movements in Arab societies.

Resumo (analítico) As teorias sobre os movimentos sociais dominantes são baseadas em experiências ocidentais. No entanto, ¿em que medida elas podem nos ajudar a compreender o processo de construção solidária de práticas políticas em contextos politicamente autoritário não-ocidentais? Bayat e a sua excepcionalidade árabe nos advertem que, nas ciências sociais, o uso de modelos epistemológicos ocidentais em contextos não-ocidentais tem frequentemente esquecido as múltiplas particularidades desses territórios, sociedades, dos processos que nelas surgem e se desenvolvem. Este artigo, no âmbito do projeto Transgang, tomará como caso de estudo um movimento social que surgiu na região do Rif em 2016 o qual foi liderado por jovens, visando analisar as suas particularidades culturais a partir de perspectivas decoloniais. Finalmente, apresentar-se-ão algumas conclusões abertas para futuras pesquisas sobre movimentos sociais juvenis nas sociedades árabes.

Should treatment of low-level rifampicin mono-resistant tuberculosis be different?

BACKGROUND: Rifampicin resistant tuberculosis (RR-TB) was frequently detected in Suriname after the introduction of Xpert MTB/RIF in 2012. Subsequent phenotypic drug-susceptibility testing (DST) was not conclusive at that moment, while RR-TB patients treated with first-line tuberculostatics had good treatment outcome. In our study, we analysed this interesting observation. METHODS: We collected demographic and clinical characteristics and treatment outcome of TB patients from May 2012-December 2018 and performed a univariate and multivariate analysis to assess possible associations with resistance to rifampicin. Secondly, we conducted whole genome sequencing on all available Mycobacterium tuberculosis isolates that had a rifampicin resistance in the Xpert MTB/RIF test and performed phenotypic DST on selected isolates. FINDINGS: RR-TB was detected in 59 (9.6%) patients confirmed by Xpert. These patients were treated with rifampicin-containing regimens in most (88%) of the cases. In all 32 samples examined, a D435Y mutation in the rpoB gene was identified; only one isolate revealed an additional isoniazid mutation. Phenotypic DST indicated low-level rifampicin resistance. In multivariate analysis, the Creole ethnicity was a factor associated with rifampicin resistance (aOR 3.5; 95%CI 1.9-6.4). The treatment success rate for patients with RR-TB (78.0%) was comparable to the treatment outcome in non-RR-TB patients 77.8%. INTERPRETATION: This study confirms a low-level rifampicin mono-resistance in TB patients of Suriname. These patients could benefit from a first-line regimen with high dose rifampicin (or rifabutin), rather than from the lengthy treatment regimens for rifampicin-resistant and multi-drug resistant TB, a concept of stratified medicine also advocated for the treatment of TB. FUNDING: None.

Diagnosis of Tuberculosis Using Gastric Aspirates in Pediatric Patients in Haiti.

BACKGROUND: We aimed to determine whether the Xpert MTB/RIF (Xpert) assay is a useful adjunct to culture for the rapid diagnosis of tuberculosis (TB) using gastric lavage aspirates (GLAs) in children aged < 5 years. METHODS: We reviewed the yield from diagnostic modalities in children suspected of having TB followed at an infectious disease research and treatment center in Port-au-Prince, Haiti, from 2011 to 2016. RESULTS: In 187 children clinically diagnosed with TB, a microbiologic diagnosis could be established in 40 (21%). Cultures, Xpert, and smears were positive in 30 (19%), 28 (17%), and 3 (1.6%) children, respectively. Ten cases that would not have been diagnosed by culture alone were found by the use of the Xpert assay. Collecting 2 GLA samples optimized microbiologic yield. CONCLUSIONS: In GLAs, Xpert increased the yield of microbiologically documented cases by 33%. Additionally, the rapidity of diagnosis potentially makes Xpert a valuable adjunct in initiating treatment for TB in children. Smear microscopy has low sensitivity in GLA and did not add to the documented cases. Our findings also highlight the low rate of microbiologic confirmation of clinically diagnosed TB.

Normalization of endometrial histopathology and endometrial NK cells concentration predict successful pregnancy in repeated implantation failure.

OBJECTIVE: The primary objective was to establish the endometrial predictors of clinical pregnancy in a population of repeated implantation failure with oocyte donation after specific endometrial treatment. The secondary one was to evaluate reproduction outcomes in terms of Implantation rate (IR), Clinical pregnancy (CP), Live birth delivery rate (LBDR) and Prematurity, in relation to normalization or no-normalization of the predictors. METHODS: 66 patients were assigned to the study. We ran a Pipelle endometrial biopsy to investigate the endometrium lymphocyte population by Flow Cytometry and abnormal/normal patterns by histopathology in pre/post-treatment. We employed the binary logistic regression model to identify the predictors for CP. For the secondary objective, we assessed the clinical outcomes in function to the normalization or no normalization in post-treatment. RESULTS: Endometrial histopathology and endometrial NK cell counts resulted in CP predictors (Wald chi2 test (p=0.044 and 0.001)), respectively. We had a higher IR, CP and LBDR when both predictors were normalized in comparison with no normalization (p<0.001). There was a high percentage of prematurity in both normalized vs. non-normalized groups (34.4% (11/32) and 71.43% (5/7), respectively) without significant differences. CONCLUSION: Endometrial histopathology and endometrial NK cell counts showed that they are valid predictors of pregnancy outcome in repeated implantation failure after treatment. In post-treatment, the pregnancy outcomes were significantly higher in the presence of both normalized predictors. Pregnancy rates were zero in the no-normalization of both predictors. There was a high percentage of prematurity in both groups.

Cost-effectiveness of Xpert®MTB/RIF in the diagnosis of tuberculosis: pragmatic study

Abstract INTRODUCTION: The intensification of research and innovation with the creation of networks of rapid and effective molecular tests as strategies for the end of tuberculosis are essential to avoid late diagnosis and for the eradication of the disease. We aimed to evaluate the cost-effectiveness of Xpert®MTB/RIF (Xpert) in the diagnosis of drug-resistant tuberculosis in reference units, in scenarios with and without subsidies, and the respective cost adjustment for today. METHODS: The analyses were performed considering as criterion of effectiveness, negative culture or clinical improvement in the sixth month of follow-up. The comparison was performed using two diagnostic strategies for the drug susceptibility test (DST), BactecTMMGITTM960 System, versus Xpert. The cost effectiveness and incremental cost-effectiveness ratio (ICER) were calculated and dollar-corrected for American inflation (US$ 1.00 = R$ 5,29). RESULTS: Subsidized Xpert had the lowest cost of US$ 33.48 (R$67,52) and the highest incremental average efficiency (13.57), thus being a dominated analysis. After the inflation was calculated, the mean cost was DST-MGIT=US$ 74.85 (R$ 396,73) and Xpert = US$ 37.33 (R$197,86) with subsidies. CONCLUSIONS: The Xpert in the diagnosis of TB-DR in these reference units was cost-effective with subsidies. In the absence of a subsidy, Xpert in TB-DR is not characterized as cost effective. This factor reveals the vulnerability of countries dependent on international organizations' subsidy policies.

Rendimiento del GeneXpert en el diagnóstico de tuberculosis pulmonar y extrapulmonar en la edad pediátrica / GeneXpert for the diagnosis of pulmonary and extrapulmonary tuberculosis in pediatric patients / GeneXpert para o diagnóstico da tuberculose pulmonar e extrapulmonar em pacientes pediátricos

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Summary: Introduction: the World Health Organization (WHO) recommends molecular biology techniques, such as Xpert MTB/RIF, for the diagnosis of tuberculosis (TB) and for the detection of Rifampicin resistance. In Uruguay, the Xpert MTB/RIF has been used since 2014, and no research papers have yet assessed its performance. A Cochrane review recommends the assessment of the Xpert diagnostic accuracy in difficult to diagnose groups, such as, children, people living with HIV and with extrapulmonary tuberculosis. Objectives: describe cases of TB in children of under 15 years of age in Uruguay during 2018 and 2019 and describe the influence of the various diagnostic tests on the bacteriological confirmation of the disease. Evaluate the performance of the Xpert MTB/RIF for the diagnosis of TB in respiratory and non- respiratory samples using the culture as a reference standard. Compare the performance of GeneXpert with smear microscopy for TB diagnosis. Material and methods: analytical, retrospective study of children of under 15 years of age in Uruguay between January 2018 and June 2019, based on data obtained from the PNC-TB information system. Clinical-epidemiological characteristics of the TB cases were described. Definitions were taken from CHLA-EP, as per WHO recommendations. All respiratory and non-respiratory samples received by the National Reference Laboratory in Tuberculosis of the CHLA-EP from 1/1/2018 to 6/30/2019, entered in the IT system ("TB soft") were analyzed; they belonged to patients with clinical suspicion of TB, studied as contacts, or to TB risk groups (patients with immunodeficiency or at risk of immunosuppression, mainly). All samples underwent smear microscopy and/or Xpert MTB/RIF (according to the CHLA-EP protocol) and culture. The detection of Rifampicin resistance in the Xpert was compared with first- line drug sensitivity tests using molecular methods made from the cultures. The sensitivity, specificity, PPV and NPV of GeneXpert and ZN microscopy were calculated using Mycobacterium tuberculosis culture as gold standard. We calculated the Xpert positive and negative likelihood ratio (LR). Results: 67 patients under 15 years of age were diagnosed with TB, and 46% cases were bacteriologically confirmed. A total of 1670 samples were analyzed; 82% respiratory and 17% non-respiratory. A total of 32 samples showed a positive culture for M. tuberculosis (14 respiratory and 18 non- respiratory). One rifampicin resistance sample was detected in the Xpert that was not confirmed in the culture. The sensitivity of Xpert for all samples was 80%; the specificity 99,5%; PPV 80%; NPV 99,5%. In the case of smear microscopy for all samples: S 44,4%, specificity 99,4%, PPV 70,6%; NPV 98,2%. Respiratory samples: Xpert S 100%; E 99,4%; PPV 66,7%; NPV 100%. Bacilloscopy: S 72,7%; E 99,6%; PPV 72,7%; NPV 99,6%. Non-respiratory samples: Xpert S: 66,7%; E 100%; PPV 100%; NPV 97,9%. Bacilloscopy: S 25%; E 98,8%; PPV 66,7%; NPV 93,2%. The LR + of the Xpert for all samples was 160 and the LR - 0,2. Conclusions: TB in children under 15 remains difficult to diagnose. Bacteriological confirmation was attempted in 88% of TB cases, and almost 50% showed positive results by some bacteriological technique. The Xpert showed a good sensitivity and specificity profile in both respiratory and non-respiratory samples, similar to those reported in international papers. The main contribution in relation to smear microscopy is the greater sensitivity for the diagnosis of TB in children under 15 years of age. The Xpert is very useful for TB diagnosis when it is positive, although it does not ensure we can rule out the disease in case of negative results.

Resumo: Introdução: a Organização Mundial da Saúde (OMS) recomenda técnicas de biologia molecular, como o Xpert MTB / RIF para o diagnóstico de tuberculose (TB) e para a detecção de resistência à Rifampicina. No Uruguai, o Xpert MTB / RIF é usado desde 2014, e o seu desempenho ainda não tem sido avaliado. Uma revisão recente da Cochrane promove que pesquisas futuras devem avaliar a precisão diagnóstica do Xpert, em grupos difíceis de diagnosticar, como crianças, pessoas vivendo com HIV e pessoas com tuberculose extrapulmonar. Objetivos: descrever os casos de tuberculose em crianças menores de 15 anos no Uruguai nos anos 2018-2019 e a contribuição dos diferentes testes de diagnóstico na confirmação bacteriológica da doença. Avaliar o desempenho do Xpert MTB / RIF para o diagnóstico de TB em amostras respiratórias e não respiratórias de pacientes menores de 15 anos, utilizando a cultura como padrão de referência. Comparar o desempenho do GeneXpert com a baciloscopia para o diagnóstico da TB. Material e métodos: estudo analítico e retrospectivo de crianças menores de 15 anos estudadas para TB no Uruguai entre janeiro de 2018 e junho de 2019, utilizando a base em dados do sistema informático PNC-TB. Descrevemos as características clínico-epidemiológicas dos casos de TB. As definições foram retiradas do CHLA-EP de acordo com as recomendações da OMS. Todas as amostras respiratórias e não respiratórias recebidas pelo Laboratório Nacional de Referência (LNR) em Tuberculose do CHLA-EP de 01/01/2018 a 30/06/2019, inseridas no sistema computacional (TB Soft), corresponderam a pacientes com suspeita clínica de TB, estudados como contatos ou na detecção de TB em grupos de risco (pacientes com imunodeficiências ou com risco de imunossupressão, principalmente). As amostras foram realizadas por esfregaço (baciloscopia) e/ou Xpert MTB/RIF (de acordo com o protocolo CHLA-EP) e por cultura. A detecção da resistência à Rifampicina no Xpert foi comparada com os testes de sensibilidade a drogas de primeira linha (PSD), utilizando os métodos moleculares das culturas. A sensibilidade, especificidade, PPV e NPV do Xpert e esfregaço foram calculados usando a cultura como padrão de referência. Calculamos a razão de verossimilhança positiva e negativa (LR) do Xpert. Resultados: 67 crianças menores de 15 anos foram diagnosticadas com TB, e 46% dos casos foram confirmados bacteriologicamente. 1670 amostras foram analisadas; 82% respiratórias e 17% não respiratórias. 32 amostras tiveram uma cultura positiva para M. tuberculosis (14 respiratórias e 18 não respiratórias). A sensibilidade (S) do Xpert para todas as amostras foi de 80% (IC95% 37,5-96,3), especificidade (E) 99,5% (IC95% 97,3-99,9), PPV 80% (37,5-96,3), NPV 99,5% (97,3-99,9). Baciloscopia: S de 46,1% (28,7-64,5), E 99,5% (98,7-99,8), PPV 75% (50,5-89,8), VPL 98,3% (97,2-99). Amostras respiratórias: Xpert S 100%; E 99,3% VPP 66% e VPN 100%. Baciloscopia: S 66,6%, E 99,8%, PPV 80%, NPV 99,7%. Amostras não respiratórias: Xpert S: 66,6%, E 100%, PPV 100%, NPV 97,9%; Esfregaço S: 25%, E 99,3%, PPV 80%, NPV 93%. O LR + do Xpert para todas as amostras foi de 160 e o LR - 0,2. Conclusões: a TB em crianças menores de 15 anos é ainda difícil de diagnosticar. Tentamos a confirmação bacteriológica em 88% dos casos de TB, e quase 50% deles tiveram resultados positivos utilizando alguma técnica bacteriológica. O Xpert mostrou um bom perfil de sensibilidade e especificidade em amostras respiratórias e não respiratórias, semelhante ao relatado em papers internacionais. A principal contribuição em relação à baciloscopia é a maior sensibilidade para o diagnóstico de TB em menores de 15 anos. O Xpert é muito útil para o diagnóstico de TB em caso de ser positivo, embora não permita descartar a doença em casos negativos.

Association of Xpert MTB/RIF Cycle Threshold Values with Tuberculosis Treatment Outcomes.

PURPOSE: Considering the current recommendation of the World Health Organization to replace sputum smear microscopy with Xpert MTB/RIF as an initial diagnostic test for tuberculosis (TB), and that culture takes time to provide results, the cycle threshold (CT) of the Xpert test may be the only way to assess bacillary load. The objective of this study is to evaluate the association of bacillary load, measured by the Xpert CT, with the TB treatment outcomes. METHODS: In cohort study, Xpert CT values were evaluated in cured and non-cured (failure and death) patients. Multivariate analysis was performed to evaluate if CT is independently associated with TB treatment outcomes. RESULTS: During this study period, 155 patients (84 cured and 71 non-cured) met the inclusion and were included in the analysis. The mean CT value for Xpert MTB/RIF test was 20.7 ± 5.6 in cured patients and 17.1 ± 5.6 in non-cured patients (p < 0.0001). Previous TB was more frequent in non-cured (28.2%) than in cured patients (7.1%) (p < 0.0001). Non-cured patients were younger than cured ones (37.1 ± 13.3 vs 43.6 ± 16.2; p = 0.006). HIV was more frequent in non-cured (28.2%) than in cured patients (15.5%), although this difference was not statistically significant (p = 0.054). In multivariate analysis, CT values, age, previous TB, and HIV were independently associated with non-cure. CONCLUSIONS: Lower Xpert MTB/RIF CT values were independently associated with worse treatment outcomes. The information from even a single test performed before starting treatment proved to be a relatively good predictor of TB treatment outcome.

Transmission of Mycobacterium tuberculosis presenting unusually high discordance between genotypic and phenotypic resistance to rifampicin in an endemic tuberculosis setting.

BACKGROUND: Since the implementation of the Xpert MTB/RIF in Sao Paulo, Brazil, numerous Mycobacterium tuberculosis isolates presenting "rifampicin-resistant genotype with rifampicin-susceptible phenotype" were observed. OBJECTIVE: To evaluate the prevalence, rpoB mutations and transmission of M. tuberculosis resistant to rifampicin on Xpert MTB/RIF but susceptible on BACTEC MGIT system, in Sao Paulo state. METHODS: Patients' isolates with this pattern of rifampicin discordance, collected from 2014 to 2017, had their rpoB predominant rifampicin-resistance-determining region sequenced and were genotyped by IS6110 restriction fragment-length polymorphism. FINDINGS: The prevalence of rifampicin-discordant M. tuberculosis with genotypic resistance was 55.1% (156/283). Among the sequenced and genotyped isolates, 75.5% (111/147) were in clusters, largely associated with the type of rpoB mutation. Most isolates (98.6%; 72/73) harbouring the predominant mutation, His445Asn, were pooled into the two largest clusters, SP2ga (42/72; 58.3%) and SP5o (12/72; 16.7%). Ranking second, isolates carrying the silent mutation Phe433Phe were mostly (92.3%; 24/26) gathered into four groups of the family SP25. CONCLUSION: These findings suggest that this unusual high rifampicin discrepancy proportion was greatly influenced by few actively circulating clusters. Further studies on many of the rpoB mutations identified in our setting are needed to elucidate their association with phenotypic rifampicin resistance.

GeneXpert® MTB/RIF assay with transbronchial lung cryobiopsy for Mycobacterium tuberculosis diagnosis.

BACKGROUND: Early tuberculosis (TB) diagnostic is one of the critical steps to TB control. GeneXpert MTB/RIF has been widely proven for a prompt TB diagnosis. The use of GeneXpert MTB/RIF assay with transbronchial lung cryobiopsy samples may increase diagnostic accuracy. We aim to assess the diagnostic of TB with GeneXpert MTB/RIF assay with transbronchial lung cryobiopsy. METHODS: Patients with suspected diagnosis of TB and negative smear microscopies, with TB culture and GeneXpert MTB/RIF assay with transbronchial lung cryobiopsy were included in this cross-sectional study. Participants were enrolled from 2016 to 2018 at National Institute of Respiratory Diseases, Mexico. RESULTS: We included 54 patients (77.8% males) aged 30 to 65 years. The sensitivity of the GeneXpert MTB/RIF assay with transbronchial lung cryobiopsy was 81.3% (95% CI, 62.1-100%), with a specificity of 100% (95% CI, 100-100%) and a negative predictive value of 92.7% (95% CI, 84.7-100%). Twenty-two patients of the total population have HIV, the sensitivity of the test in these patients was 87.5% (95% CI, 64.6-100%). Also, 28 patients with a peripheral localized lesion which had a solid pattern were identified (51.9%). The sensitivity in patients with peripheral localized lesions was 88.9% (68.4-100%). CONCLUSIONS: The GeneXpert MTB/RIF assay with transbronchial lung cryobiopsy test is as efficient as broncho alveolar lavage for TB diagnosis. Transbronchial lung cryobiopsy increases a major diagnostic opportunity when the nature of illness is malignant. Transbronchial lung cryobiopsy is efficient in HIV patients, especially in patients with peripheral localized lesion.