Comparison of PCR protocols for detecting Histoplasma capsulatum DNA through a multicenter study

Abstract

Background: A multicenter study was conducted. A panel containing DNA from Histoplasma capsulatum, as well as negative and cross-reaction controls, was sent to five different laboratories, members of the MICOMOL network from CYTED Program.

Aims: The objective was to assess the accuracy of different PCR protocols to detect H. capsulatum DNA.

Methods: Seven different PCR protocols were tested. They were based on PCR techniques and used unicopy and multicopy targets.

Results: Most of these protocols (4/7) were able to detect the smallest amounts of fungal DNA (10(2)fg/μl). Overall sensitivity was 86% and specificity was 100%. The protocol based on a unicopy target (SCAR220) presented lower sensitivity (43%) but 100% specificity. The real-time protocols tested were highly reproducible, sensitive, and specific. Neither false positives nor cross-reactions were detected in any protocol.

Conclusions: All laboratories were able to amplify H. capsulatum DNA, and real-time PCR seems to be a promising tool to efficiently detect this pathogen in clinical samples.

Keywords: Diagnosis; Diagnóstico; Histoplasmosis; Multicenter; Multicéntrico; PCR.