Abstract
The polymerase chain reaction used for amplifying an internal sequence of a previously amplified fragment (nested-PCR) was investigated as a complementary alternative for searching for alcohol-acid resistant bacilli and Mycobacterium tuberculosis cultures in Lowenstein-Jensen medium. 144 sputum samples were investigated from patients with suspected tuberculosis that were sent to the Tuberculosis Laboratory of the Evandro Chagas Institute in Belém, between June 2002 and December 2003. From the 144 samples, 121 were characterized as tuberculosis: 119 were positive in cultures, 95 under bacilloscopy and 128 using nested-PCR. The sensibility of the nested-PCR was 96% (116/121), while the specificity was 48% (11/23). Nested-PCR may be a complementary tool for diagnosing tuberculosis, since it presents sensitivity equivalent to that of cultures. However, further evaluations are needed with the aim of minimizing the number of false-positive results.
Highlights
The polymerase chain reaction used for amplifying an internal sequence of a previously amplified fragment was investigated as a complementary alternative for searching for alcohol-acid resistant bacilli and Mycobacterium tuberculosis cultures in Lowenstein-Jensen medium. 144 sputum samples were investigated from patients with suspected tuberculosis that were sent to the Tuberculosis Laboratory of the Evandro Chagas Institute in Belém, between June 2002 and December 2003
É importante relatar que todos os procedimentos de extração realizados para este trabalho foram acompanhados por controle negativo que não amplificaram nas reações de PCR, no entanto, estes controles, eram normalmente manipulados antes das amostras testes ou controles positivos (o que impossibilitava a detecção de falha técnica por contaminação do controle negativo após a pipetagem de amplicons)
Em nenhum dos casos de cultura negativa e nested-PCR positivo foi relatado tratamento anterior ou história de contato com paciente infectado pelo M. tuberculosis, o que poderia sugerir presença de DNA de M. tuberculosis nos indivíduos investigados
Summary
A reação em cadeia da polimerase (PCR) vem sendo investigada como alternativa de alta sensibilidade e especificidade para o diagnóstico rápido de doenças infecciosas como a tuberculose[11 12]. A nested-PCR, constituída por duas amplificações sucessivas do marcador molecular, é defendida por conciliar ainda maior sensibilidade e especificidade[7 9]. Este trabalho propõe avaliar a aplicabilidade da nested-PCR do gene que codifica o antígeno b para o diagnóstico da tuberculose
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