Wânia da Silva Carvalho

Spoligotyping of clinical Mycobacterium tuberculosis isolates from the state of Minas Gerais, Brazil

We performed spoligotyping on 114 strains of the Mycobacterium tuberculosis (Mtb) complex that had been isolated from patients in Minas Gerais Health Units during 2004. A total of 82/114 (72%) clinical isolates were clustered and 32/114 (28%) were unique. Seven shared types containing nine strains were newly created. A total of nine patterns corresponded to unreported orphan strains, as evaluated against all of the strains recorded in the SITVIT2 proprietary database in the Institut Pasteur de la Guadeloupe. The major clades were composed of isolates that belong to the following genotypes: Latin-America and Mediterranean (63/114, 55.3%) (the ill-defined T superfamily) (12/114, 10.5%), Haarlem (8/114, 7%), X clade (6/114, 5.3%), S clade (3/114, 2.6%) and the East-African Indian and Manu types, each with 1/114 (0.9%) isolates. A considerable number of strains (n = 20, 17.5%) showed patterns that did not fall within any of the previously described major clades. We conclude the bulk of tuberculosis (TB) (92/114, 80.7%) in our location is recent evolutionary strains that belong to the principal genetic groups 2/3. Further studies on epidemiology of TB are required to understand Mtb biodiversity and TB transmission in this region.

Evaluation of six different DNA extraction methods for detection of Mycobacterium tuberculosis by means of PCR-IS6110: preliminary study

BackgroundDevelopments in molecular detection and strain differentiation of members of Mycobacterium tuberculosis complex have proved to be useful. The DNA extraction method influences the amplification efficiency, causing interference on the sensitivity and respective inhibitors. The aim of this study was to standardize a simple and fast DNA extraction method, providing DNA amplification by IS6110-PCR effectively free from undue interferences.FindingsThe efficiency of the six different protocols tested in M. tuberculosis cultures has varied from 75% to 92.5%. This preliminary study evaluating the IS6110 PCR sensitivity and specificity was developed in DNA extracted from microscope slides, and achieved 100% of efficiency.ConclusionsDNA extraction by Chelex + NP-40 method from both, cultures of M. tuberculosis and smear slides, resulted in good quantity of interference free DNA, especially in samples with low concentrations of genetic material; therefore, such technique may be used for the molecular diagnosis of tuberculosis.

Diagnóstico de resistência do Mycobacterium tuberculosis à rifampicina utilizando-se da reação em cadeia da polimerase

A resistencia do Mycobacterium tuberculosis aos tuberculostaticos tem surgido como grande ameaca a Saude Publica. A resistencia a rifampicina pode ser considerada como um marcador para a multi-resistencia a farmacos e tem sido atribuida a mudancas estruturais da RNA polimerase, produto de expressao do gene rpoB. Os pacientes portadores dessas cepas tem baixa perspectiva frente ao tratamento. Os testes convencionais de sensibilidade aos farmacos realizados em cultura do Mtb requerem varias semanas para o crescimento. Por este motivo, a Reacao em Cadeia da Polimerase (PCR), metodo de baixo custo e que pode reduzir o tempo para o diagnostico, representa alternativa viavel e promissora. Neste artigo estao descritos os metodos mais comumente empregados na deteccao de mutantes resistentes a rifampicina baseados na PCR, como analise de Polimorfismo Conformacional de Fita Simples (Single-Strand Conformation Polymorphism, SSCP), PCR Heteroduplex e INNO-LIPA. Recentemente, padronizou-se a tecnica de PCR em baixa estringencia, usando um unico iniciador (Low Stringency Single Specific Primer, LSSP), que se mostrou um metodo rapido e sensivel na deteccao de mutacoes no gene rpoB.

Positive tuberculin test and risk of infection by Mycobacterium tuberculosis in a tuberculosis clinic settled in an upright building, in Minas Gerais, Brazil

Antecedentes: La tuberculosis ocupacional causa ausentismo laboral. Conocer la reactividad tuberculinica de los trabajadores de la salud ayuda a mejorar las medidas de control para evitar la infeccion laboral con M. tuberculosis. Objetivo: Estudiar la reactividad tuberculinica de un grupo de trabajadores de la salud de un centro de referencia de tuberculosis que esta ubicado en una edificacion vertical. Material y Metodos: Se realizo una prueba de tuberculina a 251 empleados de una clinica de tuberculosis, que no tenian antecedentes o sospecha de tuberculosis y quienes no habian sido sometidos previamente a la prueba. Aquellos sujetos que no respondieron a tuberculina fueron sometidos a una nueva prueba para evaluar la reaccion de refuerzo. Resultados: El 51% de los sujetos estudiados tuvieron una reaccion tuber-culinica positiva. El fenomeno de refuerzo se observo en 69 de 190 sujetos (35,7%). La tasa de seroconversion en aquellos sujetos con una tuberculina negativa en que se repitio la prueba la cabo de un ano, fue de 5,1%. Conclusiones: En este grupo de trabajadores de la salud expuestos a M. tuberculosis, hay una alta tasa de reactividad negativa a tuberculina.

Prevalence of latent Mycobacterium tuberculosis infection in prisoners

ABSTRACT Objective: To determine the prevalence of and the factors associated with latent Mycobacterium tuberculosis infection (LTBI) in prisoners in the state of Minas Gerais, Brazil. Methods: This was a cross-sectional cohort study conducted in two prisons in Minas Gerais. Tuberculin skin tests were performed in the individuals who agreed to participate in the study. Results: A total of 1,120 individuals were selected for inclusion in this study. The prevalence of LTBI was 25.2%. In the multivariate analysis, LTBI was associated with self-reported contact with active tuberculosis patients within prisons (adjusted OR = 1.51; 95% CI: 1.05-2.18) and use of inhaled drugs (adjusted OR = 1.48; 95% CI: 1.03-2.13). Respiratory symptoms were identified in 131 (11.7%) of the participants. Serological testing for HIV was performed in 940 (83.9%) of the participants, and the result was positive in 5 (0.5%). Two cases of active tuberculosis were identified during the study period. Conclusions: Within the prisons under study, the prevalence of LTBI was high. In addition, LTBI was associated with self-reported contact with active tuberculosis patients and with the use of inhaled drugs. Our findings demonstrate that it is necessary to improve the conditions in prisons, as well as to introduce strategies, such as chest X-ray screening, in order to detect tuberculosis cases and, consequently, reduce M. tuberculosis infection within the prison system.

Evaluation of the Mean Cost and Activity Based Cost in the Diagnosis of Pulmonary Tuberculosis in the Laboratory Routine of a High-Complexity Hospital in Brazil

At a global level, with the increase in healthcare costs, there is a need to assess the economic impact of the incorporation of new technologies in different health disorders in different countries. There is scarce information regarding costs incurred with the use of current or new diagnostic tests for tuberculosis or from the vantage point of their incorporation within the healthcare systems of high-burden countries. The present study aimed to assess the mean cost and the activity based cost of the laboratory diagnosis for tuberculosis by means of conventional techniques and from the Detect TB®LabTest molecular test kit in a general high-complexity hospital of the public health system in Brazil. Cost analysis was performed by means of primary data, collected in the Mycobacteria and Molecular Biology Laboratory in 2013. The mean cost and activity based cost were, respectively, U

In-house PCR with DNA extracted directly from positive slides to confirm or exclude the diagnosis of tuberculosis: focus on biosafety

10.06/U

Correlation between the BACTEC MGIT 960 culture system with Genotype MTBDRplus and TB-SPRINT in multidrug resistant Mycobacterium tuberculosis clinical isolates from Brazil

5.61 for centrifuged bacilloscopy by Ziehl Neelsen (ZN) and Auramine (AU); U

Prevalence of latent tuberculosis infection and risk of infection in patients with chronic kidney disease undergoing hemodialysis in a referral center in Brazil

7.42/U

Evaluation of the nitrate reductase assay for the rapid detection of resistance to first-line medications in Mycobacterium tuberculosis strains isolated from patients in a general hospital

4.15 for direct bacilloscopy by ZN; U