Variable efficiency of three primer pairs for the diagnosis of Pneumocystis carinii pneumonia by the polymerase chain reaction

Andrea De Luca, Enrica Tamburrini, Elena Ortona, Paola Mencarini, Paola Margutti, Andrea Antinori, Elena Visconti, Alessandra Siracusano

Research output: Contribution to journalArticlepeer-review

Abstract

The efficiency of three different primer pairs, complementary to different Pneumocystis carinii DNA regions, was compared in the polymerase chain reaction (PCR) for the diagnosis of Pneumocystis carinii pneumonia (PCP) on bronchoalveolar fluid (BALF) from patients with AIDS. PCR coupled with dot-blot hybridization (BLOT) using primers and probe from the mitochondrial 23SrDNA region showed the highest sensitivity, with a lower detection limit of 0·5-1 organisms μl-1. When testing 47 BALF, PCR plus BLOT if the mitochondrial 23SrDNA region showed also the best diagnostic efficiency (97% sensitivity, 100% specificity). Sensitivity was significantly higher than with PCR and BLOT of the 5SrDNA region (81·5% sensitivity; P = 0.025, McNemar test); and of the dehydrofolate reductase (DHFR) gene region (75·6% sensitivity; P = 0·019). Sensitivity was also significantly higher than indirect immunofluorescence (75·8% sensitivity; P = 0·008). Using DHFR primers and probe, specificity was also reduced. The diagnostic sensitivity in clinical specimens paralleled the detection limit in the standard dilutions. The use of repeated DNA sequences of proven specificity as target of PCR amplification favourably influences sensitivity and specificity. This comparative study demonstrates that primer selection plays a significant role in the diagnosis of PCP by PCR.

Original languageEnglish
Pages (from-to)333-340
Number of pages8
JournalMolecular and Cellular Probes
Volume9
Issue number5
DOIs
Publication statusPublished - 1995

Keywords

  • 5SrDNA
  • bronchoalveolar lavage fluid
  • dehydrofolate reductase gene
  • mitochondrial DNA
  • PCR
  • Pneumocystis carinii pneumonia

ASJC Scopus subject areas

  • Cell Biology
  • Molecular Biology

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