Evaluación del método de Petroff modificado con solución salina para el diagnóstico de micobacterias en el sistema Bact/Alert 3D / Assessment of Petroff method modified with saline solution for the diagnosis of mycobacteria in Bact/Alert 3D system

Introducción: Existen diferentes métodos de descontaminación de muestras pulmonares para el diagnóstico de micobacterias. El Programa Nacional de Control de Tuberculosis recomienda el método de Petroff modificado con solución salina, pero no existen evidencias documentadas que avalen este método. Objetivo: Evaluar el método de Petroff modificado con solución salina para el diagnóstico de micobacterias en el sistema Bact/Alert 3D. Métodos: Se realizó un estudio observacional analítico de pruebas diagnósticas utilizando 100 muestras pulmonares recibidas en el Laboratorio Nacional de Referencia e Investigaciones de Tuberculosis, Lepra y Micobacterias del Instituto Pedro Kourí, abril 2016 enero 2017. La muestra se dividió en 3 alícuotas y se descontaminaron mediante 3 métodos; luego se inocularon en los medios de cultivo sólido y líquido. Se compararon los resultados del cultivo en cuanto: tiempo de detección de crecimiento, tasa de contaminación, por ciento de positividad, además se calcularon indicadores de desempeño. Resultados: Al comparar el método Petroff modificado con solución salina con el Petroff modificado con solución fosfato en Löwenstein Jensen, el tiempo de detección de crecimiento, por ciento de positividad y la tasa de contaminación se comportaron de forma similar y la sensibilidad (93,75 por ciento), concordancia (96,47 por ciento) e índice de Youden (0,91) fueron elevadas. Al compararlo el Petroff modificado con solución salina con el N-Acetil-L-Cisteína, las variables no mostraron diferencias significativas y los Indicadores de Desempeño se comportaron por encima del 93 por ciento, para el medio sólido y líquido. Conclusiones: Los resultados avalan la continuidad del uso del Petroff modificado con solución salina como método de descontaminación de las muestras pulmonares en la red de laboratorios de Cuba y como alternativa en el pretratamiento de las muestras para el medio líquido (Bact/Alert 3D), además constituye un soporte para el Programa Nacional de Control de Tuberculosis(AU)

Introduction: There are different decontamination methods of pulmonary samples for the diagnosis of mycobacteria. The National Program for the Control of Tuberculosis recommends Petroff method modified with saline solution; but there are not documented evidences that endorse it. Objective: Assess Petroff method modified with saline solution for the diagnosis of mycobacteria in Bact / Alert 3D system. Methods: An observational analytic study of diagnostic tests was conducted; there were used 100 pulmonary samples received in the National Laboratory of References and Researches of Tuberculosis, Leprosy and Mycobacteria of Pedro Kourí Institute, from April 2016 to January 2017. The sample was divided in 3 aliquots and those were decontaminated using 3 methods; then, they were inoculated in the solid and liquid culture means. Cultures´ results were compared according to: growth's detection time, contamination rate, percent of positivity; in addition, performance indicators were calculated. Results: When comparing Petroff method modified with saline solution with Petroff method modified with phosphate solution in Löwenstein Jensen, the growth's detection time, the percent of positivity and the rate of contamination behaved similarly, and sensitivity (93,75percent), concordance (96,47percent) and Youden´s index (0,91) were high. When the Petroff method modified with saline solution was compared with N-Acetil-L- Cisteina, the variables did not show significative differences and the behaviour indicators were over 93percent for the solid and liquid mean. Conclusions: The results endorse the continuity of the use of Petroff method modified with saline solution as a decontamination method of pulmonary samples in the network of Cuban laboratories and as alternative to the pre-treatment of the samples for the liquid mean (Bact/Alert 3D); it also constitutes a support for the National Program for the Control of Tuberculosis(AU)

Evaluation of modified petroff’s method and manual MGIT culture system to facilitate detection of acid-fast bacilli in smear-negative presumptive cases of pulmonary tuberculosis

Background: Tuberculosis is a major public health problem in India. According to RNTCP guidelines, all efforts are to be made to diagnose tuberculosis on the basis of detection of AFB/ MTB in the clinical specimen; hence all those cases where AFB/ MTB have not been detected are presumptive cases of tuberculosis. In this background, the present study aims to detect AFB/ MTB in the sputum of clinico-radiologically presumptive cases of pulmonary tuberculosis using direct ZN microscopy, Modified Petroff’s method followed by ZN microscopy & MGIT liquid culture method. Materials and Methods: A total of 94 sputum samples which were negative for AFB by direct ZN microscopy at the DMC of the institute, were subjected to Modified Petroff’s method followed by microscopy & MGIT liquid culture test to find out any additional yield of bacteriologically confirmed disease. Result: Out of 94 specimens, 20 (21.28%) were positive for AFB on ZN microscopy post Modified Petroff’s method. Among all the specimens, 30 (31.91%) were found to be positive by MGIT liquid culture method with average time to detection growth (TTD) around 16.73 days (2-38 days). Conclusion: Modified Petroff’s method followed by ZN microscopy & MGIT test for mycobacterial culture can improve case detection.

Comparison between Ogawa-Kudoh and modified Petroff techniques for mycobacteria cultivation in the diagnosis of pulmonary tuberculosis / Comparação entre os métodos de Ogawa-Kudoh e Petroff modificado para o cultivo de micobactérias no diagnóstico da tuberculose pulmonar

ABSTRACT Objective To compare the performance of the Ogawa-Kudoh method with the modified Petroff technique in diagnosis of pulmonary tuberculosis. Methods A total of 205 sputum samples from 166 patients with clinical suspicion or under pulmonary tuberculosis follow-up, seen at a public tertiary care hospital, from July 2014 to July 2016 were used. All samples were simultaneously processed using the Ogawa-Kudoh and modified Petroff decontamination methods, according to the recommendations of the Ministry of Health. In the statistical analysis, the McNemar test and the Kappa index were used, respectively, to compare proportions and verify agreement between data. Results The Ogawa-Kudoh and modified Petroff methods were efficient in mycobacteria detection, with no significant differences in results (p=0.549) and contamination rate of the cultures (p=0.065). The agreement between techniques was considered excellent (Kappa index of 0.877) and Ogawa-Kudoh, as compared to the modified Petroff technique, showed sensitivity of 90.4%, specificity of 96.6%, positive predictive value of 94.3% and negative predictive value of 94.2%. Conclusion The Ogawa-Kudoh technique proved to be sufficiently sensitive and specific for diagnosis of pulmonary tuberculosis, and, therefore, suitable for routine laboratory application. Since it is simple, low-cost and has less technical requirements for biosafety and professional training, Ogawa-Kudoh is an alternative for managers and healthcare professionals to promote the expansion of bacteriological diagnostic coverage of pulmonary tuberculosis.

RESUMO Objetivo Comparar o desempenho do método de Ogawa-Kudoh ao de Petroff modificado no diagnóstico da tuberculose pulmonar. Métodos Utilizaram-se 205 amostras de escarro de 166 pacientes com suspeita clínica ou controle de tuberculose pulmonar atendidos em um hospital público terciário, entre os meses de julho de 2014 a julho de 2016. Todas as amostras foram processadas simultaneamente pelos métodos de descontaminação Ogawa-Kudoh e Petroff modificado, seguindo as recomendações do Ministério da Saúde. Na análise estatística, foi empregado o teste de McNemar, para comparação de proporções, e o índice Kappa, para verificar o grau de concordância entre os dados. Resultados Os métodos Ogawa-Kudoh e Petroff modificado mostraram-se eficientes na detecção de micobactérias, não sendo verificadas discordâncias significativas tanto nas comparações de pares de resultados (p=0,549), como na taxa de contaminação das culturas (p=0,065). O grau de concordância das técnicas foi considerado excelente (índice Kappa de 0,877), e o Ogawa-Kudoh, em relação ao Petroff modificado, apresentou 90,4% de sensibilidade, 96,6% de especificidade, 94,3% de valor preditivo positivo e 94,2% de valor preditivo negativo. Conclusão O método de Ogawa-Kudoh revelou-se suficientemente sensível e específico para o diagnóstico da tuberculose pulmonar e, portanto, adequado para a aplicação na rotina laboratorial. Por ser mais simples, de baixo custo e com menores exigências técnicas de biossegurança e capacitação profissional, o Ogawa-Kudoh apresenta-se como alternativa para gestores e profissionais da área promoverem a ampliação da cobertura diagnóstica bacteriológica da tuberculose pulmonar.

Is universal sample processing methodology better than conventional techniques for detection of tuberculosis.

Background: The identification of infectious cases is a crucial first step for tuberculosis control programmes worldwide. It relies exclusively on the detection of acid‑fast bacilli in sputum by smear microscopy. Therefore, there is an urgent and definite need to improve the sensitivity of smear microscopy. Objective: The USP method was compared with the two most commonly used conventional methods of smear microscopy namely; direct smear microscopy and the microscopy by modified Petroff’s method. Materials and Methods: Two samples from each patient were taken from 197 patients of presumptive tuberculosis. One smear was made for direct Ziehl‑Neelsen staining and two smears were made after processing by two concentration methods i.e., modified Petroff’s and USP solution. LJ media were inoculated for culture after processing by both concentration methods. Results: Among 197 cases 93 were culture positive by either method. Out of 93 culture‑positive sample, 78.5% were direct smear positive, 89% were 4%NaOH smear positive and 96% were USP smear‑positive samples but difference in diagnostic accuracy of USP (96%) and modified Petroff method (93%) is not statistically significant (P > 0.01). Conclusion: The present study evaluated the smear microscopy by USP method with two conventional methods, direct microscopy and microscopy by modified Petroff’s method. The study concludes that although USP method is more sensitive than conventional methods, it is not feasible to include it in diagnosis of early tuberculosis within RNTCP.

Comparison of three decontamination methods for Mycobacterium bovis isolation / Comparação de três métodos de descontaminação para isolamento de Mycobacterium bovis

Sixty samples of tissue fragments with lesions suggestive of tuberculosis from bovine abattoirs, kept in saturated solution of sodium borate, were subjected to four treatments: 4 percent NaOH (Petroff Method), 12 percent H2SO4 and 1.5 percent HPC (1-Hexadecylpyridinium Chloride) decontamination, and physiological saline solution (control). The HPC method showed the lowest contamination rate (3 percent) when compared to control (88 percent, p<0.001), NaOH (33 percent, p<0.001) and H2SO4 (21.7 percent, p<0.002). Regarding the isolation success, the HPC method was better (40 percent) than the control (3 percent, p<0.001), NaOH (13 percent, p=0.001) and H2SO4 (1.7 percent, p<0.001) methods. These results indicate that HPC is an alternative to the Petroff method.

Sessenta amostras de fragmentos de tecidos com lesões sugestivas de tuberculose provenientes de abatedouros bovinos, conservadas em solução saturada de borato de sódio, foram submetidas a quatro tratamentos: descontaminação através dos métodos NaOH 4 por cento (Método Petroff), H2SO4 12 por cento e HPC (Cloreto de hexadecilpiridínio) 1,5 por cento, e solução salina (controle). O método HPC apresentou a menor proporção de contaminação (3 por cento), em relação ao controle (88 por cento, p<0,001), NaOH (33 por cento, p<0,001) e H2SO4 (21,7 por cento, p=0,002). Em relação ao sucesso no isolamento, o método HPC apresentou o melhor resultado (40 por cento), em relação ao controle (3 por cento, p<0,001), NaOH (13 por cento, p=0,001) e H2SO4 (1,7 por cento, p<0,001). Os resultados indicam que o HPC é uma alternativa à utilização do método Petroff.