Objective: To analyze the spatial distribution of tuberculosis in individuals under 15 years old and the socioeconomic factors in Paraíba, Brazil, 2007-2016. Methods: Ecological study, with Sinan’s data, considering the municipality as the unit of analysis. The spatial distribution of the incidence was performed, the local empirical Bayesian method and Moran's statistics were applied. Socioeconomic data were crossed to identify areas of social prosperity. Results: 426 cases were reported, with an average incidence of 4.5/100 thousand inhabitants. Moran's index = 0.59 (p=0.010). The Moran Map revealed a concentration of cases in children under 15 in 38 municipalities with a high priority of care, in conglomerates in the high-high and low-low patterns, in the east and northwest of the state, coincided with areas of low social prosperity. Conclusion: There are clusters with greater transmission of tuberculosis, indicating priority areas for approaching tuberculosis.

Objetivo: Analisar a distribuição espacial da tuberculose em indivíduos menores de 15 anos de idade e fatores socioeconômicos na Paraíba, Brasil, 2007-2016. Métodos: Estudo ecológico, com dados do Sinan, tendo-se o município como unidade de análise. Realizou-se distribuição espacial da incidência, aplicou-se o método bayesiano empírico local e a estatística de Moran. Dados socioeconômicos foram cruzados, para identificação das áreas de prosperidade social. Resultados: Foram notificados 426 casos, com incidência média de 4,5/100 mil habitantes. O índice de Moran foi de 0,59 (p-valor=0,010). O Moran Map revelou concentração de casos em menores de 15 anos em 38 municípios com alta prioridade da atenção, em conglomerados nos padrões alto-alto e baixo-baixo, nas regiões leste e noroeste do estado, coincidindo com áreas de baixa prosperidade social. Conclusão: Há clusters com maior transmissão da tuberculose, apontando áreas prioritárias para abordagem da tuberculose.

Resumo Objetivo: Analisar a distribuição espacial da tuberculose em indivíduos menores de 15 anos de idade e fatores socioeconômicos na Paraíba, Brasil, 2007-2016. Métodos: Estudo ecológico, com dados do Sistema de Informação de Agravos de Notificação, sendo o município a unidade de análise. Realizou-se distribuição espacial da incidência, aplicou-se o método bayesiano empírico local e a estatística de Moran. Dados socioeconômicos foram cruzados, para identificação das áreas de prosperidade social. Resultados: Foram notificados 426 casos, com incidência média de 4,5/100 mil habitantes. O índice de Moran foi de 0,59 (p-valor=0,010). O Moran Map revelou concentração de casos em menores de 15 anos em 38 municípios com alta prioridade da atenção, em conglomerados nos padrões alto-alto e baixo-baixo, nas regiões leste e noroeste do estado, coincidindo com áreas de baixa prosperidade social. Conclusão: Houve clusters com maior transmissão da tuberculose, apontando áreas prioritárias para abordagem da tuberculose.

Abstract Objective: To analyze spatial distribution of tuberculosis in individuals under 15 years old and socioeconomic factors in Paraíba, Brazil, 2007-2016. Methods: This was an ecological study based on data from the Notifiable Health Conditions Information System (SINAN), taking each municipality to be a unit of analysis. Spatial distribution of incidence was performed, the local empirical Bayesian method and Moran's I were applied. Socioeconomic data were crossed-checked to identify areas of social prosperity. Results: 426 cases were notified, with average incidence of 4.5/100,000 inhabitants. Moran's I was 0.59 (p=0.010). The Moran Map revealed concentration of cases in children under 15 in 38 high priority municipalities, in clusters with high-high and low-low patterns, in the east and northwest of the state, coinciding with areas of low social prosperity. Conclusion: There were clusters with greater tuberculosis transmission, indicating priority areas for addressing tuberculosis.

Resumen Objetivo: Analizar la distribución espacial de la tuberculosis en menores de 15 años y los factores socioeconómicos en Paraíba, Brasil, 2007-2016. Métodos: Estudio ecológico, con datos de Sistema de Información de Agravamientos de Notificación, considerando el municipio como unidad de análisis. Se realizó la distribución espacial de la incidencia, se aplicó el método empírico local Bayesiano y la estadística de Moran. Se cruzaron datos socioeconómicos para identificar áreas de vulnerabilidad social. Resultados: Se notificaron 426 casos, con una incidencia promedio de 4,5/100 mil habitantes. El índice de Moran = 0,59 (p=0,010). El Mapa de Moran reveló una concentración de casos en menores de 15 años en 38 municipios con alta prioridad de atención, en conglomerados en los patrones alto-alto y bajo-bajo, en las regiones este y noroeste del estado, coincidiendo con áreas de baja prosperidad social. Conclusión: Existían clusters con mayor transmisión de tuberculosis, lo que indica áreas prioritarias para abordar la tuberculosis.

Resumo Objetivo: Analisar a distribuição espacial da tuberculose em indivíduos menores de 15 anos de idade e fatores socioeconômicos na Paraíba, Brasil, 2007-2016. Métodos: Estudo ecológico, com dados do Sistema de Informação de Agravos de Notificação, sendo o município a unidade de análise. Realizou-se distribuição espacial da incidência, aplicou-se o método bayesiano empírico local e a estatística de Moran. Dados socioeconômicos foram cruzados, para identificação das áreas de prosperidade social. Resultados: Foram notificados 426 casos, com incidência média de 4,5/100 mil habitantes. O índice de Moran foi de 0,59 (p-valor=0,010). O Moran Map revelou concentração de casos em menores de 15 anos em 38 municípios com alta prioridade da atenção, em conglomerados nos padrões alto-alto e baixo-baixo, nas regiões leste e noroeste do estado, coincidindo com áreas de baixa prosperidade social. Conclusão: Houve clusters com maior transmissão da tuberculose, apontando áreas prioritárias para abordagem da tuberculose.

Abstract Objective: To analyze spatial distribution of tuberculosis in individuals under 15 years old and socioeconomic factors in Paraíba, Brazil, 2007-2016. Methods: This was an ecological study based on data from the Notifiable Health Conditions Information System (SINAN), taking each municipality to be a unit of analysis. Spatial distribution of incidence was performed, the local empirical Bayesian method and Moran's I were applied. Socioeconomic data were crossed-checked to identify areas of social prosperity. Results: 426 cases were notified, with average incidence of 4.5/100,000 inhabitants. Moran's I was 0.59 (p=0.010). The Moran Map revealed concentration of cases in children under 15 in 38 high priority municipalities, in clusters with high-high and low-low patterns, in the east and northwest of the state, coinciding with areas of low social prosperity. Conclusion: There were clusters with greater tuberculosis transmission, indicating priority areas for addressing tuberculosis.

Resumen Objetivo: Analizar la distribución espacial de la tuberculosis en menores de 15 años y los factores socioeconómicos en Paraíba, Brasil, 2007-2016. Métodos: Estudio ecológico, con datos de Sistema de Información de Agravamientos de Notificación, considerando el municipio como unidad de análisis. Se realizó la distribución espacial de la incidencia, se aplicó el método empírico local Bayesiano y la estadística de Moran. Se cruzaron datos socioeconómicos para identificar áreas de vulnerabilidad social. Resultados: Se notificaron 426 casos, con una incidencia promedio de 4,5/100 mil habitantes. El índice de Moran = 0,59 (p=0,010). El Mapa de Moran reveló una concentración de casos en menores de 15 años en 38 municipios con alta prioridad de atención, en conglomerados en los patrones alto-alto y bajo-bajo, en las regiones este y noroeste del estado, coincidiendo con áreas de baja prosperidad social. Conclusión: Existían clusters con mayor transmisión de tuberculosis, lo que indica áreas prioritarias para abordar la tuberculosis.

Abstract INTRODUCTION: Nontuberculous mycobacteria (NTM) species, as human pathogens, are increasing in the world, as is the difficulty of accurately identifying them. Differential diagnosis, especially between the M. tuberculosis complex and NTM species, and the characterization of NTM species is important. This study aimed to evaluate the performance of a molecular system based on multiplex real-time PCR with high-resolution melting (HRM) for the identification and differentiation of NTM species of clinical importance of an endemic area for tuberculosis in northeastern Brazil. METHODS: The technical protocol of the molecular system was based on multiplex real-time PCR-HRM, and evaluated the sensitivity and specificity of the detection of NTM species in mycobacterial clinical isolates from the studied region. The gold standard method was specific gene sequencing. RESULTS: The sensitivity and specificity of multiplex real-time PCR-HRM modified for differentiation between NTM and M. tuberculosis were 90% and 100%, respectively. The PCR-HRM sensitivities for the characterization of NTM species (M. kansasii, M. abscesses, M. avium, and M. fortuitum) were 94.59%, 80%, 57.14%, and 54%, respectively. CONCLUSIONS The multiplex real-time PCR-HRM modified assay has the potential to rapidly and efficiently identify nontuberculous mycobacteria of clinical importance, which is crucial for immediate implementation of the appropriate therapy and thus avoiding complications and sequelae in patients.

Abstract INTRODUCTION: Laboratory and clinical features of childhood tuberculosis (TB) are non-specific and establishing an accurate diagnosis remains a challenge. This study evaluated a Single tube nested-PCR (STNPCR) to detect genomic DNA of Mycobacterium tuberculosis complex in blood and urine. METHODS: Biological samples were obtained from children (<15 years old) with clinical suspicion of pulmonary and extrapulmonary TB at public hospitals in Recife-Pernambuco, Brazil. Cultures yielded negative results in a majority of childhood TB cases, which are generally paucibacillary. A set of clinical, epidemiological, radiological, and laboratory criteria with evident clinical improvement after anti-TB treatment were frequently used to define childhood TB cases. RESULTS: Ninety children with clinical suspicion were enrolled in this study (44 with TB and 46 without TB). The pulmonary TB group had 20 confirmed cases and 46 negative controls, while the extrapulmonary TB group had 24 confirmed cases. The STNPCR showed sensitivities to pulmonary and extrapulmonary TB of 47.4% and 52.2% (blood) and 38.8% and 20% (urine), respectively. Considering the low performance of STNPCR on separate samples, we decided to perform a combined analysis (parallel sensitivity analysis) of the results from blood and urine samples. The parallel sensitivity increased to 65% in blood and 62.5% in urine. The specificity in both samples ranged from 93.5-97.8%. CONCLUSIONS: Although STNPCR showed moderate sensitivity, the specificity is high; therefore, the test can be used as an auxiliary tool to diagnose TB in children. It is a rapid test that demonstrated better performance than other diagnostic tests in paucibacillary samples as it does in childhood tuberculosis.

Abstract INTRODUCTION: This study evaluated the performance of the IS6110-TaqMan® assay in different types of biological samples and tissues for laboratory diagnosis of extrapulmonary tuberculosis. METHODS: 143 biological samples and tissues from patients with suspected extrapulmonary tuberculosis from the health services of Recife/Pernambuco/Brazil were evaluated with the IS6110-TaqMan® assay. RESULTS: The sensitivities of the IS6110-TaqMan® assay calculated for blood, urine, both blood and urine samples, tissue biopsies, extrapulmonary body fluid samples, and all samples from patients calculated together were 55.9%, 33.3%, 68.8%, 43.8%, 29.6%, and 73.7%, respectively, and the specificities were 80%, 100%, 78.6%, 100%, 100%, and 84.2%, respectively. CONCLUSIONS The accuracy of qPCR was high in various clinical sample types. The analysis of more than one type of clinical sample collected from the same patient with extrapulmonary tuberculosis enhances the diagnostic power of the IS6110-TaqMan® assay when compared with the use of only one clinical sample.

Abstract β-Defensin-1, an antimicrobial peptide encoded by the DEFB1 gene, is known to play an important role in lung mucosal immunity. In our association study we analyzed three DEFB1 functional polymorphisms -52G>A (rs1799946), -44C>G (rs1800972) and -20G>A (rs11362) in 92 tuberculosis patients and 286 healthy controls, both from Northeast Brazil: no association was found between the studied DEFB1 polymorphisms and the disease. However we cannot exclude that this lack of association could be due to the low number of subjects analyzed, as suggested by the low statistical power achieved for the three analyzed SNPs (values between 0.16 and 0.50).

Abstract: INTRODUCTION : Molecular analyses are auxiliary tools for detecting Koch's bacilli in clinical specimens from patients with suspected tuberculosis (TB). However, there are still no efficient diagnostic tests that combine high sensitivity and specificity and yield rapid results in the detection of TB. This study evaluated single-tube nested polymerase chain reaction (STNPCR) as a molecular diagnostic test with low risk of cross contamination for detecting Mycobacterium tuberculosis in clinical samples. METHODS: Mycobacterium tuberculosis deoxyribonucleic acid (DNA) was detected in blood and urine samples by STNPCR followed by agarose gel electrophoresis. In this system, reaction tubes were not opened between the two stages of PCR (simple and nested). RESULTS: STNPCR demonstrated good accuracy in clinical samples with no cross contamination between microtubes. Sensitivity in blood and urine, analyzed in parallel, was 35%-62% for pulmonary and 41%-72% for extrapulmonary TB. The specificity of STNPCR was 100% in most analyses, depending on the type of clinical sample (blood or urine) and clinical form of disease (pulmonary or extrapulmonary). CONCLUSIONS: STNPCR was effective in detecting TB, especially the extrapulmonary form for which sensitivity was higher, and had the advantage of less invasive sample collection from patients for whom a spontaneous sputum sample was unavailable. With low risk of cross contamination, the STNPCR can be used as an adjunct to conventional methods for diagnosing TB.

The authors report a case of a 38-year-old HIV-positive woman, with subcutaneous nodules on the thoracic region with 3 months of evolution. Clinical, laboratory, and epidemiological features were evaluated and associated with apparent damage to the T11-T12 vertebrae, identification by imaging tests, positivity in a polymerase chain reaction-based test, and reactivity to the Mantoux tuberculin skin test (PPD-RT 23). The patient was diagnosed with osteoarticular tuberculosis and received treatment for a year, and clinical cure was achieved.

OBJETIVO: Comparar o desempenho da técnica nested polymerase chain reaction (NPCR) com aquele de culturas na detecção do complexo Mycobacterium tuberculosis em espécimes pulmonares e extrapulmonares.MÉTODOS: Analisamos 20 e 78 espécimes pulmonares e extrapulmonares, respectivamente, de 67 pacientes hospitalizados com suspeita de tuberculose. Um sistema automatizado foi utilizado na identificação de culturas de Mycobacterium spp., e M. tuberculosis IS6110 foi utilizada como sequência alvo na NPCR. A estatística kappa foi utilizada para verificar a concordância entre os resultados.RESULTADOS: Entre os 67 pacientes, 6 e 5, respectivamente foram diagnosticados com tuberculose pulmonar e extrapulmonar, e a NPCR foi positiva em todos os casos. Entre os 98 espécimes clínicos, a baciloscopia, cultura e NPCR foram positivas em 6,00%, 8,16% e 13,26%, respectivamente. Comparando-se os resultados da NPCR com aqueles da cultura (padrão ouro) nos espécimes pulmonares, a sensibilidade e a especificidade foram 100% e 83%, respectivamente, enquanto essas nos espécimes extrapulmonares foram 83% e 96% respectivamente, com boa concordância entre os testes (kappa, 0,50 e 0,6867, respectivamente).CONCLUSÕES: Embora a NPCR tenha se mostrado uma ferramenta muito útil na detecção do complexo M. tuberculosis, No entanto, os resultados positivos da NPCR devem ser associados à clínica, dados clínicos, epidemiológicos e outros dados laboratoriais devem também ser considerados no diagnóstico e tratamento da tuberculose pulmonar e extrapulmonar.

OBJECTIVE: To compare the performance of nested polymerase chain reaction (NPCR) with that of cultures in the detection of the Mycobacterium tuberculosis complex in pulmonary and extrapulmonary specimens.METHODS: We analyzed 20 and 78 pulmonary and extrapulmonary specimens, respectively, of 67 hospitalized patients suspected of having tuberculosis. An automated microbial system was used for the identification of Mycobacterium spp. cultures, and M. tuberculosis IS6110 was used as the target sequence in the NPCR. The kappa statistic was used in order to assess the level of agreement among the results.RESULTS: Among the 67 patients, 6 and 5, respectively, were diagnosed with pulmonary and extrapulmonary tuberculosis, and the NPCR was positive in all of the cases. Among the 98 clinical specimens, smear microscopy, culture, and NPCR were positive in 6.00%, 8.16%, and 13.26%, respectively. Comparing the results of NPCR with those of cultures (the gold standard), we found that NPCR had a sensitivity and specificity of 100% and 83%, respectively, in pulmonary specimens, compared with 83% and 96%, respectively, in extrapulmonary specimens, with good concordance between the tests (kappa, 0.50 and 0.6867, respectively).CONCLUSIONS: Although NPCR proved to be a very useful tool for the detection of M. tuberculosis complex, clinical, epidemiological, and other laboratory data should also be considered in the diagnosis and treatment of pulmonary and extrapulmonary tuberculosis.

Introduction This study evaluated the performance of an in-house nested-PCR system for the detection of the Mycobacterium tuberculosis complex in pleural fluid, blood and urine samples from pleural effusion tuberculosis patients by health services physicians in Pernambuco, Brazil. Methods A prospective double-blind study with 37 hospitalized patients of both sexes, aged over 15, was used to investigate the diagnosis of pleural effusion. The criteria used to define the cases included the demonstration of bacillus in biological samples by smear or culture or by a granulomatous finding in the histopathological examination, associated with an evident response to specific treatments to each clinical situation. Pleural fluid, blood and urine samples were collected and subjected to routine tests and the nested PCR technique to assess for M. tuberculosis amplification. Results In total, 37 pleural effusion patients took part in the study, of whom 19 (51.3%) had tubercular etiologies and 18 (48.7%) had etiologies from other causes. When the pleural fluid, blood and/or urine sample in-house nested-PCR sensitivities were evaluated simultaneously, the results were positive regardless of the biological specimen (the sensitivity was 84.2%); however, when the blood and/or urine samples were analyzed together, the sensitivity was 72.2%. When the pleural fluid samples were evaluated alone, the sensitivity was only 33.3%. Conclusions The performance of the diagnostic pleural tuberculosis nested-PCR was directly related to the diversity of the samples collected from the same patient. Additionally, this study may identify a need to prioritize non-invasive blood and urine collection for this diagnosis.

Introduction The early diagnosis of mycobacterial infections is a critical step for initiating treatment and curing the patient. Molecular analytical methods have led to considerable improvements in the speed and accuracy of mycobacteria detection. Methods The purpose of this study was to evaluate a multiplex polymerase chain reaction system using mycobacterial strains as an auxiliary tool in the differential diagnosis of tuberculosis and diseases caused by nontuberculous mycobacteria (NTM) Results Forty mycobacterial strains isolated from pulmonary and extrapulmonary origin specimens from 37 patients diagnosed with tuberculosis were processed. Using phenotypic and biochemical characteristics of the 40 mycobacteria isolated in LJ medium, 57.5% (n=23) were characterized as the Mycobacterium tuberculosis complex (MTBC) and 20% (n=8) as nontuberculous mycobacteria (NTM), with 22.5% (n=9) of the results being inconclusive. When the results of the phenotypic and biochemical tests in 30 strains of mycobacteria were compared with the results of the multiplex PCR, there was 100% concordance in the identification of the MTBC and NTM species, respectively. A total of 32.5% (n=13) of the samples in multiplex PCR exhibited a molecular pattern consistent with NTM, thus disagreeing with the final diagnosis from the attending physician. Conclusions Multiplex PCR can be used as a differential method for determining TB infections caused by NTM a valuable tool in reducing the time necessary to make clinical diagnoses and begin treatment. It is also useful for identifying species that were previously not identifiable using conventional biochemical and phenotypic techniques.

The authors report a case of a 12-year-old child with a complaint of pain and deformity in the lower thoracic region that had lasted for two years. Clinical, epidemiological and laboratory characteristics associated with images of apparent damage in the T9-T10 and T11-T12 vertebrae taken by radiography of the thoracic spine and nuclear magnetic resonance in addition to the positivity of the molecular test based on the polymerase chain reaction, led to tuberculous spondylitis being diagnosed and specific therapy was started. Culture of vertebral biopsy was positive for Mycobacterium tuberculosis after thirty days.

The polymerase chain reaction (PCR) and its variations, such as the nested-PCR, have been described as promising techniques for rapid diagnosis of tuberculosis (TB). With the aim of evaluating the usefulness of a nested-PCR method on samples of blood and urine of patients suspected of tuberculosis we analyzed 192 clinical samples, using as a molecular target the insertion element IS6110 specific of M. tuberculosis genome. Nested-PCR method showed higher sensitivity in patients with extrapulmonary tuberculosis (47.8% and 52% in blood and urine) when compared to patients with the pulmonary form of the disease (sensitivity of 29% and 26.9% in blood and urine), regardless of the type of biological sample used. The nested-PCR is a rapid technique that, even if not showing a good sensitivity, should be considered as a helpful tool especially in the extrapulmonary cases or in cases where confirmatory diagnosis is quite difficult to be achieved by routine methods. The performance of PCR-based techniques should be considered and tested in future works on other types of biological specimens besides sputum, like blood and urine, readily obtainable in most cases. The improving of M. tuberculosis nested-PCR detection in TB affected patients will give the possibility of an earlier detection of bacilli thus interrupting the transmission chain of the disease.

O trabalho visou à otimização de um método baseado na reação em cadeia da polimerase multiplex - para diferenciação de micobactérias de interesse para a saúde pública. A PCR Multiplex baseou-se na amplificação simultânea do genehsp65, presente em todo gênero Mycobacterium, do gene dnaJ, presente apenas em Mycobacterium tuberculosis e Mycobacterium avium e da sequência de inserção IS6110 presente no complexo Mycobacterium tuberculosis, gerando amplicons de 165pb, 365pb e 541pb, respectivamente. O limite de detecção foi de 1fg para o alvo hsp65, 100pg para o dnaJ e 0,1fg para o IS6110. A PCR multiplex detectou até 100pg de DNA de Mycobacterium tuberculosis. O sistema demonstrou ser específico e sensível na detecção de Mycobacterium tuberculosis, Mycobacterium bovis, Mycobacterium avium e Mycobacterium smegmatis. Os resultados obtidos utilizando cepas de referência demonstraram que a PCR multiplex pode ser uma ferramenta rápida, sensível e específica na diferenciação de micobactérias.

This study aimed to optimize a method based on the polymerase chain reaction - multiplex PCR - for differentiation of mycobacteria species of interest for public health. The multiplex PCR was based on simultaneous amplification of the hsp65 gene, which is present in all species of the Mycobacterium genus, the dnaJ gene, which is present only in Mycobacterium tuberculosis and Mycobacterium avium and the IS6110 insertion sequence, which is present in the Mycobacterium tuberculosis complex, generating amplicons of 165 bp, 365 bp and 541 bp, respectively. The detection limit was 1 fg for the hsp65 target, 100 pg for dnaJ and 0.1 fg for IS6110. The multiplex PCR detected down to 100 pg of DNA of Mycobacterium tuberculosis. The system was shown to be specific and sensitive for detection of Mycobacterium tuberculosis, Mycobacterium bovis, Mycobacterium avium and Mycobacterium smegmatis. The results obtained using reference strains of mycobacteria showed that multiplex PCR may be a fast, sensitive and specific tool for differentiation of mycobacteria.