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Open Access Research article

Comparison of two laboratory-developed PCR methods for the diagnosis of Pulmonary Tuberculosis in Brazilian patients with and without HIV infection

Luciene C Scherer13*, Rosa D Sperhacke2, Carla Jarczewski5, Patrícia I Cafrune2, Candice T Michelon2, Rubia Rupenthal2, Marta Osorio Ribeiro6, Antonio Ruffino Netto4, Maria LR Rossetti23 and Afrânio L Kritski4

Author Affiliations

1 Post Graduation Program in Biological Science-Biochemistry Department, Federal University of Rio Grande do Sul-UFRGS, Porto Alegre/RS/Brazil

2 Technological and Scientific Development Center-CDCT, State Foundation in Production and Health Research - FEPPS/RS) Porto Alegre/RS/Brazil

3 Lutheran University of Brasil-ULBRA, Canoas/RS/Brazil

4 Academic Tuberculosis Program, Medical School, Clementino Fraga Filho Hospital, ATP-MS/HUCFF, Federal University of Rio de Janeiro-UFRJ, Rio de Janeiro, Brazil

5 Parthenon Hospital/Secretary of Health of Rio Grande do Sul/Porto Alegre/RS/Brazil

6 Public Laboratory of the State of Rio Grande do Sul (LACEN/RS), State Foundation in Production and Health Research - FEPPS/RS) Porto Alegre/RS/Brazil

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BMC Pulmonary Medicine 2011, 11:15  doi:10.1186/1471-2466-11-15

Published: 29 March 2011

Abstract

Background

Direct smear examination with Ziehl-Neelsen (ZN) staining for the diagnosis of pulmonary tuberculosis (PTB) is cheap and easy to use, but its low sensitivity is a major drawback, particularly in HIV seropositive patients. As such, new tools for laboratory diagnosis are urgently needed to improve the case detection rate, especially in regions with a high prevalence of TB and HIV.

Objective

To evaluate the performance of two in house PCR (Polymerase Chain Reaction): PCR dot-blot methodology (PCR dot-blot) and PCR agarose gel electrophoresis (PCR-AG) for the diagnosis of Pulmonary Tuberculosis (PTB) in HIV seropositive and HIV seronegative patients.

Methods

A prospective study was conducted (from May 2003 to May 2004) in a TB/HIV reference hospital. Sputum specimens from 277 PTB suspects were tested by Acid Fast Bacilli (AFB) smear, Culture and in house PCR assays (PCR dot-blot and PCR-AG) and their performances evaluated. Positive cultures combined with the definition of clinical pulmonary TB were employed as the gold standard.

Results

The overall prevalence of PTB was 46% (128/277); in HIV+, prevalence was 54.0% (40/74). The sensitivity and specificity of PCR dot-blot were 74% (CI 95%; 66.1%-81.2%) and 85% (CI 95%; 78.8%-90.3%); and of PCR-AG were 43% (CI 95%; 34.5%-51.6%) and 76% (CI 95%; 69.2%-82.8%), respectively. For HIV seropositive and HIV seronegative samples, sensitivities of PCR dot-blot (72% vs 75%; p = 0.46) and PCR-AG (42% vs 43%; p = 0.54) were similar. Among HIV seronegative patients and PTB suspects, ROC analysis presented the following values for the AFB smear (0.837), Culture (0.926), PCR dot-blot (0.801) and PCR-AG (0.599). In HIV seropositive patients, these area values were (0.713), (0.900), (0.789) and (0.595), respectively.

Conclusion

Results of this study demonstrate that the in house PCR dot blot may be an improvement for ruling out PTB diagnosis in PTB suspects assisted at hospitals with a high prevalence of TB/HIV.