sybr-green-i and Pneumonia--Pneumocystis

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.

Topics: Animals; Benzothiazoles; CDC2 Protein Kinase; Diamines; DNA, Fungal; Fluorescent Dyes; Genes, cdc; Humans; Microscopy, Fluorescence; Organic Chemicals; Pneumocystis; Pneumonia, Pneumocystis; Quinolines; Rats; Reverse Transcriptase Polymerase Chain Reaction; Sensitivity and Specificity; Tetrahydrofolate Dehydrogenase