Real Time Quantitative PCR and RT--PCR for Analysis of Pneumocystis Carinii Hominis

J Microbiol Methods. 2001 Jun;45(2):113-8. doi: 10.1016/s0167-7012(01)00241-x.

Abstract

Pneumocystis carinii hominis is a common cause of pneumonia in immunocompromised patients and particularly in those infected by HIV. Giemsa- and Gomori--Grocott-stained smears are widely used for detection and quantification of this opportunistic fungus obtained from biological samples or from in vitro culture. But these methods are fastidious and time-consuming. Thus, instead of performing a count of organisms, we focused our attention on the level of specific DNA by a quantitative PCR technique. This procedure has the advantage of greater precision and more objectivity. To verify the presence of organisms, quantitative RT--PCR based on DHFR and a cell cycle mRNA have been developed. In this current study, we present a detailed description of these methods and their applications for analysis of P. carinii hominis.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • CDC2 Protein Kinase / genetics
  • DNA, Fungal / chemistry*
  • DNA, Fungal / genetics
  • DNA, Fungal / isolation & purification
  • Fluorescent Dyes / chemistry
  • Genes, cdc
  • Humans
  • Microscopy, Fluorescence
  • Organic Chemicals*
  • Pneumocystis / genetics*
  • Pneumocystis / isolation & purification
  • Pneumonia, Pneumocystis / diagnosis
  • Pneumonia, Pneumocystis / microbiology*
  • Rats
  • Reverse Transcriptase Polymerase Chain Reaction*
  • Sensitivity and Specificity
  • Tetrahydrofolate Dehydrogenase / chemistry
  • Tetrahydrofolate Dehydrogenase / genetics

Substances

  • DNA, Fungal
  • Fluorescent Dyes
  • Organic Chemicals
  • SYBR Green I
  • Tetrahydrofolate Dehydrogenase
  • CDC2 Protein Kinase