Background: The emergence of drug-resistant strain of M.tuberculosis is one of the most critical issues facing TB researchers and clinicians. Rapid diagnosis of drug-resistant tuberculosis is essential for the prompt initiation of effective second-line therapy to improve treatment outcome and limit transmission of this obstinate disease. The aim of this study is to develop a Real-time PCR assay for the detection of mutations in RRDR (rifampcin resistance determinant region) of rpoB which conferring rifampicin resistance in Mycobacterium tuberculosis.

 Materials and Methods: In this experimental study, the primer and probe set were designed for a RRDR region of rpoB gene using a specialized software. Clinical specimens that had previously been evaluated resistant or sensitive by using convential method, were used for assessing the clinical sensitivity and specificity of the assay.

Results: The clinical sensitivity of the assay was determined 100%. The primers and the probes were rpoB specific and no cross-reaction was observed with other microorganisms and human genome bioinformatically. The clinical specificity of developed Real-time PCR assay was examined experimentally using 25 negative samples and determined to be 100%.

Conclusion: The developed real-time PCR assay can be used as an appropriate and efficient tool for the rapid detection of rifampicin-resistant Mycobacterium tuberculosis.

Background: The Real-time PCR assay has been established as the standard method for Human Cytomegalovirus (HCMV) quantitation in immunocompromised patients. However, the question of which one of whole blood or plasma specimens is better for viral quantitation is still unresolved for many clinical laboratories. To answer this question, the current study compares HCMV DNA load in whole blood and plasma samples.

Materials and Methods: In this prospective study, the whole blood and plasma samples were obtained from 41 transplantated patients and the viral load was detected using a validated, in-house Real-time PCR assay.

Results: Of the total 193 examined specimens, 174 were negative and 19 samples, from 16 patients, were positive in at least one of whole blood or plasma samples. Based on the results of linear regression analysis, the cytomegalovirus viral load was correlated in whole blood and plasma samples (R2: 0.872). However, the regression equation shows that the HCMV load in whole blood samples is higher than load of this virus in plasma. The validity of the quantitative results was confirmed by repeating the tests and analyzing the results using the repeated measure analysis.

Conclusion: Based on the results of the present study, HCMV quantitation in whole blood samples has a higher analytical sensitivity than in plasma samples.