Microbiología Médica y Diagnóstico

The Effect of Three Decontamination Methods on COBAS Amplicor Mycobacterium tuberculosis PCR Assay

Abstract

Gülnur Tarhan, Mehmet Bakır Saygan, Salih Cesur, Fatih Ocak and Ä°smail Ceyhan

In this study, we evaluated the effect of three decontamination-homogenization-concentration (DHC) methods on COBAS Amplicor Mycobacterium tuberculosis (MTB) PCR system (Roche Diagnostics, Inc., Branchburg, USA) in three different periods. A total of 1210 clinical specimens (602 pulmonary, 608 extrapulmonary specimens) were investigated. Specimens were decontaminated periodically using three DHC methods (Method A: 3% NaOHtrisodium citrate-N-acetyl-L-cysteine (NALC), Method B: 4% NaOH-Bromothymol Blue (BTB) method-Direct, Method C: 4% NaOH-BTB method-irrigation before DNA extraction method). Definitive results were obtained from 1011 (83.6%) of 1210 samples. The inhibition rates according to DHC methods (Method A, B, C) were respectively 3.3% (10/302), 4.3% (7/162) and 17.3% (24/138) for pulmonary samples; 10.3% (30/291), 19.5% (33/169) and 35.1% (52/148) for extrapulmonary samples; 6.7% (40/593), 12.1% (40/331) and 26.5% (76/286) for all samples respectively When inhibition rates were compared in terms of samples types and numbers, high inhibitor rates were found in urine 24.4%, cerebrospinal fluid (CSF) 9.5% and gastric lavage fluid (GLF) 7.6%, respectively. Using culture results as standard, the sensitivity, specificity, positive (PPV) and negative (NPV) predictive values of COBAS Amplicor MTB PCR assay were, respectively, 68.2%, 99.1%, 75% and 98.8% for the method A, 66.7%, 98%, 75% and 96.9% for method B, 75.0%, 98.9%, 75% and 98.8% for method C. We conclude that laboratories planning to use nucleic acid amplification (NAA) methods as supplement to conventional methods, should be prefer 3% NaOH-trisodium citrate-NALC method.