Rapid identification of Mycobacterium tuberculosis in cultures by molecular and immunological methods / Identificação rápida de Mycobacterium tuberculosis em culturas por método molecular e método imunológico

Objective: The study aimed to evaluate molecular and immunological methods and to propose a workflow using them for tuberculosis (TB) diagnosis routine. Methods: A cross-sectional retrospective study was performed, including 121 liquid cultures from a TB laboratory located in the extreme south of Brazil. All cultures were positive for Mycobacterium tuberculosis complex (MTBC) by in-house Polymerase Chain Reaction (PCR) using DNA extracted by the CTAB method (PCR-CTAB) for IS6110 detection. These cultures were subjected to faster tests than this one, the immunological MPT64 assay and the PCR using DNA extracted by thermal lysis method (PCR-TL), and these were evaluated for MTBC identification using PCR-CTAB as a reference method. Results: The sensitivity of MPT64 assay and PCR-TL to identify MTBC in positive cultures by PCR-CTAB were 73.6% (89/121) and 98.3% (119/121), respectively. We proposed a workflow based on the use of MPT64 assay in liquid cultures suggestive of MTBC, and in case of a negative result, we suggest the performance of PCR-TL. The PCR-CTAB is suggested only if faster tests are negative. Conclusions: Methods capable of confirming MTBC in cultures should continue to be standardized, tested, and optimized to meet the ideal requirements of simplicity, quickness, and effectiveness. The molecular and immunological methods evaluated have differences in the execution and detection of MTBC in cultures, but they are rapid tools for laboratory TB diagnosi

Objetivos: O estudo objetivou avaliar métodos molecular e imunológico e propor um fluxo de trabalho utilizando-os para a rotina de diagnóstico da tuberculose (TB). Métodos: Foi realizado um estudo transversal retrospectivo, incluindo 121 culturas líquidas de um laboratório de TB localizado no extremo sul do Brasil. Todas as culturas foram positivas para o complexo Mycobacterium tuberculosis (CMTB) por Reação em Cadeia da Polimerase (PCR) in-house para detecção do IS6110, usando DNA extraído pelo método CTAB (PCR-CTAB). Essas culturas foram submetidas a testes mais rápidos que este, o ensaio imunológico MPT64 e a PCR com DNA extraído pelo método de lise térmica (PCR-LT), e estas foram avaliadas para identificação de CMTB usando PCR-CTAB como método de referência. Resultados: A sensibilidade do ensaio MPT64 e da PCR-LT para identificar o CMTB em culturas positivas pela PCRCTAB foi de 73,6% (89/121) e 98,3% (119/121), respectivamente. Propusemos um fluxo de trabalho baseado no uso do ensaio MPT64 em culturas líquidas sugestivas de CMTB e, em caso de resultado negativo, sugerimos a realização de PCR-LT. Sugere-se a PCR-CTAB apenas se os testes mais rápidos forem negativos. Conclusões: Os métodos capazes de confirmar o CMTB em culturas devem continuar sendo padronizados, testados e otimizados para atender aos requisitos ideais de simplicidade, rapidez e eficácia. Os métodos molecular e imunológico avaliados apresentam diferenças na execução e detecção do CMTB em culturas, mas são ferramentas rápidas para o diagnóstico laboratorial da TB.

Impact of the COVID-19 pandemic on laboratory diagnosis of tuberculosis in southern Brazil / Impacto da pandemia de COVID-19 no diagnóstico laboratorial de tuberculose no sul do Brasil / Impacto de la pandemia COVID-19 en el diagnóstico de laboratorio de la tuberculosis en el sur de Brasil

Background and objectives: to understand the impact of the COVID-19 pandemic on tuberculosis (TB) diagnosis in different settings is essential to guide the establishment of appropriate TB control strategies. This study aimed to assess the influence of COVID-19 pandemic in laboratory diagnosis of TB in patients tested and diagnosed for TB. Methods: a data survey was carried out in the database of laboratories that perform TB diagnosis for the public health system in Rio Grande city (Rio Grande do Sul, Brazil). Results: there was a decrease of 1,368 to 735 (reduction of 46.3%) in the number of patients tested for TB in public diagnostic services in 2019 and 2020, respectively, and a decrease of 197 to 119 (reduction of 39.6%) in the number of new TB cases diagnosed. In contrast, the positivity rate was 14.4% in 2019 and 16.2% in 2020. Moreover, it was observed that the laboratory that performs the diagnostic service for Primary Health Care was the most affected, when compared to Tertiary Health Care. Conclusion: as a consequence of measures to control the spread of SARS-CoV-2, there was a reduction in TB testing and in the detection of new cases, especially in Primary Health Care, where patients with less need for hospitalization are received.(AU)

Justificativa e objetivos: compreender o impacto da pandemia COVID-19 no diagnóstico da tuberculose (TB) em diferentes locais é essencial para orientar o estabelecimento de estratégias adequadas de controle da TB. O objetivo deste estudo foi avaliar a influência da pandemia de COVID-19 no diagnóstico laboratorial de TB, em pacientes testados e diagnosticados com TB. Métodos: foi realizado um levantamento de dados no banco de dados de laboratórios que realizam diagnóstico de TB para o sistema público de saúde na cidade de Rio Grande (Rio Grande do Sul, Brasil). Resultados: houve redução de 1.368 para 735 (redução de 46,3%) no número de pacientes testados para TB nos serviços públicos de diagnóstico em 2019 e 2020, respectivamente, e redução de 197 para 119 (redução de 39,6%) no número de novos casos de TB diagnosticados. Em contrapartida, a taxa de positividade foi de 14,4% em 2019 e 16,2% em 2020. Além disso, observou-se que o laboratório que realiza o serviço de diagnóstico para a Atenção Primária à Saúde foi o mais afetado, quando comparado com a Atenção Terciária à Saúde. Conclusão: como consequência das medidas de controle da disseminação do SARS-CoV-2, houve redução na testagem de TB e na detecção de novos casos, principalmente na atenção primária à saúde, onde são recebidos pacientes com menor necessidade de internação.(AU)

Justificación y objetivos: comprender el impacto de la pandemia Covid-19 en el diagnóstico de tuberculosis (TB) en diferentes lugares es fundamental para orientar el establecimiento de estrategias adecuadas de control de la TB. El objetivo de este estudio fue evaluar la influencia de la pandemia de COVID-19 en el diagnóstico de laboratorio de TB, en términos de pacientes examinados y diagnosticados de TB. Métodos: los datos fueron recolectados de la base de datos de los laboratorios que realizan el diagnóstico de TB para el sistema público de salud en la ciudad de Rio Grande (Rio Grande do Sul, Brasil). Resultados: hubo una reducción de 1.368 a 735 (reducción del 46,3%) en el número de pacientes sometidos a pruebas de TB en los servicios públicos de diagnóstico en 2019 y 2020, respectivamente, y una reducción de 197 a 119 (reducción del 39,6%) en el número de nuevos casos de TB diagnosticados. Por otro lado, la tasa de positividad fue de 14,4% en 2019 y 16,2% en 2020. Además, se observó que el laboratorio que realiza el servicio de diagnóstico para la Atención Primaria de Salud fue el más afectado, en comparación con la Atención Terciaria de Salud. Conclusiones: como consecuencia de las medidas para el control de la propagación del SARS-CoV-2, hubo una reducción en las pruebas de TB y en la detección de nuevos casos, especialmente en la Atención Primaria de Salud, donde se encuentran los pacientes con menor necesidad de hospitalización.(AU)

Tuberculosis cases in a prison in the extreme south of Brazil.

Introduction. Tuberculosis (TB) control is a challenge, especially in vulnerable populations, such as prisoners.Hypothesis. In prison houses, the transmission of micro-organisms that cause infectious diseases can occur due to the susceptibility and immune compromise of prisoners, and due to the precarious physical conditions of the prison houses. However, strategies such as monitoring by health professionals, can mitigate the transmission of these micro-organisms, as well as, reduce the number of coinfections and antimicrobials resistance.Aim. This study attempted to analyse the dynamics of transmission and the antimicrobial resistance profile of Mycobacterium tuberculosis strains obtained from prisoners and to characterize the epidemiological, clinical and laboratory profiles of prisoners diagnosed with TB.Methodology. A cross-sectional and retrospective study was conducted with sputum samples collected from 228 distinct prisoners who were treated at the Health Unit located in the Regional Penitentiary of Rio Grande, Rio Grande do Sul, Brazil. The antimicrobial resistance profile of the strains was evaluated using the Resazurin Microtiter Assay and the transmission dynamics was investigated using 15-loci MIRU-VNTR.Results. Thirty-five patients (15.4 %) were diagnosed with TB, and when a TB/HIV coinfection was assessed, 8.6 % (3/35) of the patients were positive. In addition, all patients with results available for HBV, HCV, syphilis and diabetes mellitus were negative. Based on the genotypic profile, 55.9 % of the clinical isolates were grouped into five groups. One isolate with mono-resistance to isoniazid and two with mono-resistance to streptomycin were found.Conclusion. The presence of a Health Unit may have influenced the low numbers of TB/HIV, TB/HBV, TB/HCV, TB/syphilis coinfections and TB cases resistant to antimicrobials. Recent M. tuberculosis transmission can be inferred based on the high percentage of formatting of clusters. This situation stresses the need to improve active and passive detection, the screening of individuals for TB upon entrance into prison for early detection, and the implementation of prophylactic measures to reduce M. tuberculosis transmission.

Nontuberculous mycobacteria in patients with suspected tuberculosis and the genetic diversity of Mycobacterium avium in the extreme south of Brazil.

OBJECTIVE: Nontuberculous mycobacteria (NTM) are a heterogeneous group of bacteria that are widely distributed in nature and associated with opportunistic infections in humans. The aims of this study were to identify NTM in patients with suspected tuberculosis who presented positive cultures and to evaluate the genetic diversity of strains identified as Mycobacterium avium. METHODS: We studied pulmonary and extrapulmonary samples obtained from 1,248 patients. The samples that tested positive on culture and negative for the M. tuberculosis complex by molecular identification techniques were evaluated by detection of the hsp65 and rpoB genes and sequencing of conserved fragments of these genes. All strains identified as M. avium were genotyped using the eight-locus mycobacterial interspersed repetitive unit-variable-number tandem-repeat method. RESULTS: We found that NTM accounted for 25 (7.5%) of the 332 mycobacteria isolated. Of those 25, 18 (72%) were M. avium, 5 (20%) were M. abscessus, 1 (4%) was M. gastri, and 1 (4%) was M. kansasii. The 18 M. avium strains showed high diversity, only two strains being genetically related. CONCLUSIONS: These results highlight the need to consider the investigation of NTM in patients with suspected active tuberculosis who present with positive cultures, as well as to evaluate the genetic diversity of M. avium strains.

Extrapulmonary TB or sarcoidosis associated with TB? Case report / Tuberculose extrapulmonar ou sarcoidose associada a tuberculose? Relato de caso / Tuberculosis extrapulmonar o sarcoidosis asociada a la tuberculosis? Relato de caso

Objetivos: O presente relato tem como objetivo discutir o diagnostico diferencial entre tuberculose e sarcoidose. Descrição do caso: Caso de um paciente do sexo masculino, 25 anos, que desenvolveu artrite granulomatosa e osteomielite com inflamação granulomatosa não caseosa sugestiva de sarcoidose. Vinte e quatro meses antes desse relato, o paciente teve resultados laboratoriais positivos para tuberculose pulmonar, no entanto, não foi tratado. Conclusão: Demonstramos como é necessário o acompanhamento do paciente após a realização de um teste diagnóstico para tuberculose e o desafio de confirmar um diagnóstico de sarcoidose em um paciente com tuberculose.(AU)

Objectives: The present report aims to discuss the differential diagnosis between tuberculosis and sarcoidosis. Case description: The case of a 25-year-old male patient who developed granulomatous arthritis and osteomyelitis with nonnecrotizing granulomatous inflammation suggesting a sarcoidosis. Twenty-four months before this report, he had positive laboratory results to pulmonary tuberculosis, however, he was not treated. Conclusion: This case report demonstrates how necessary is the follow-up of patient after the performance of a diagnostic testing to tuberculosis, and the challenge to confirm a diagnosis of sarcoidosis in a patient with tuberculosis.(AU)

Objetivos: El presente relato tiene como objetivo discutir el diagnóstico diferencial entre tuberculosis y sarcoidosis. Descripción del caso: Caso de un paciente del sexo masculino, 25 años, que desarrolló artritis granulomatosa y osteomielitis con inflamación granulomatosa no caseosa sugestiva de sarcoidosis. Veinticuatro meses antes de ese relato, el paciente tuvo resultados de laboratorio positivos para la tuberculosis pulmonar, por lo que no fue tratado. Conclusión: Demostramos cómo es necesario el acompañamiento del paciente después de la realización de una prueba diagnóstica para tuberculosis y el desafío de confirmar un diagnóstico de sarcoidosis en un paciente con tuberculosis.(Au)

The BACTEC MGIT(tm) 320 system as a laboratory tool to diagnose tuberculosis in a Brazilian hospital with a high prevalence of HIV infection.

INTRODUCTION: The World Health Organization endorses the BACTEC Mycobacterial Growth Indicator Tube (MGIT)(tm) system as a rapid, sensitive, and specific method to diagnostic of tuberculosis. Here, we compared the performance of this system against Ogawa-Kudoh cultures and microscopy. METHODS: A total of 927 samples were obtained between December 2011 and December 2013 from 652 cases of suspected tuberculosis at the School Hospital of the Federal University of Rio Grande in Brazil. RESULTS: The MGIT system confirmed tuberculosis in more cases in less time. CONCLUSIONS: The MGIT system is an effective tool for early diagnosis of tuberculosis, especially in patients with HIV/AIDS.

Molecular typing of Mycobacterium bovis isolated in the south of Brazil.

Bovine tuberculosis is a major infectious disease of the cattle. In this study, 85 M. bovis isolates from 162 lymph nodes, obtained from a herd of cattle on a farm in southern Brazil, were evaluated using spoligotyping and VNTR. The strains were grouped into five clusters and five orphans, showing a heterogenic genetic profile, what could represent diverse geographic origins of the introduced cows and/or the frequent movement of cattle between different properties.

Molecular typing of Mycobacterium bovis isolated in the south of Brazil

Bovine tuberculosis is a major infectious disease of the cattle. In this study, 85 M. bovis isolates from 162 lymph nodes, obtained from a herd of cattle on a farm in southern Brazil, were evaluated using spoligotyping and VNTR. The strains were grouped into five clusters and five orphans, showing a heterogenic genetic profile, what could represent diverse geographic origins of the introduced cows and/or the frequent movement of cattle between different properties.