Polymerase Chain Reaction targeting insertion sequence IS6110 for the diagnosis of pulmonary tuberculosis among Sudanese children and young adults

Abstract

This study was carried out in Khartoum State during the period from January 2011 to December 2013 to improve the rate of detection of Mycobacterium tuberculosis (MTB) in children with symptoms of tuberculosis (TB) infection using different conventional and advanced diagnostic techniques. One hundred and ninety-seven specimens of gastric lavage and sputum were collected from different hospitals in Khartoum State, including Elbolok Hospital, Jafar Ibn Owf Hospital, Elasha'ab Teaching Hospital, Soba University Hospital and Academy Charity Hospital. All children participating in the study were subjected to the Mantoux test after obtaining appropriate consent injected by 5 tuberculin units of tuberculin purified protein derivative, and the results were recorded after three days. Specimens were decontaminated and inoculated on Lowenstein-Jensen media according to the modified Petroff's method. Two smears were prepared and stained by Ziehl-Neelsen stain and Auramine fluorescent dye; bacterial DNA was extracted from each specimen by using phenol chloroform method, and then the Polymerase Chain Reaction technique was adopted to detect Insertion Sequence IS6110 gene of MTB in these specimens. This study showed that the positive results for TST, ZN, Auramine, Culture and PCR were 86 (43.7%), 16 (8.1%), 22 (11.2%), 32 (16.2%) and 35(17.8%), respectively. The study concluded that the PCR technique is the most sensitive and specific technique for a quick identification of MTB in gastric lavage and sputum from children who are unable to expectorate a good quality sputum sample or who are diagnosed as negative using conventional diagnostic methods.

Journey towards National Institute of One Health in India

“…Standard tube agglutination test (STAT) and Rose Bengal plate test (RBPT) were performed according to the procedure described by the World Organization for Animal Health (OIE) 4 . The samples from animals were screened for tuberculosis and leptospirosis by targeting the IS1160 gene 5 and secY gene 6 , respectively.…”

Sensitive detection of the IS6110 sequence of Mycobacterium tuberculosis complex based on PCR-magnetic bead ELISA

“…8 The polymerase chain reaction (PCR) amplication of IS6110 sequence has been successfully applied for the detection of M. tuberculosis complex from the various types of clinical specimens. [9][10][11] However, the lower sensitivity of agarose gel electrophoresis, the commonly used method for the detection and visualization of PCR products, is the limitation of conventional PCR amplication. 12 Quantitative PCR amplication technique offers the real-time quantitation of target nucleic acids during PCR reaction cycles overcoming the laborious post-PCR detection methods.…”

Diagnóstico de tuberculosis pulmonar en lavado broncoalveolar: desempeño de la PCR en comparación con las pruebas microbiológicas de rutina

“…Las muestras obtenidas por lavados broncoalveolares se deben procesar precozmente, antes de cumplir las 12 horas de recolectadas, debido a que los anestésicos empleados en la broncoscopia disminuyen la viabilidad del bacilo y pueden producir un retardo en el desarrollo o un cultivo falso negativo [15]. Actualmente, en el mundo se cuenta con metodologías moleculares, rápidas y sensibles, que acortan el tiempo de diagnóstico de la tuberculosis, lo que indirectamente minimiza los riesgos de exposición de la población susceptible al contagio y la instauración de tratamientos innecesarios [16][17][18][19][20][21]. El objetivo de este trabajo fue describir la eficacia de la reacción en cadena de la polimerasa (PCR) para la detección de Mycobacterium tuberculosis, en comparación con el cultivo y la baciloscopia de muestras de lavado broncoalveolar, en casos sospechosos de tuberculosis pulmonar.…”