PCR in diagnosis of pneumocystosis of rats

Polymerase chain reaction (PCR) is a powerful technique to detect scanty amount of DNA from living organisms. The present study intended to develop specific primers for PCR diagnosis of pneumocystosis and to evaluate diagnostic efficacy by preparation of template DNAs from invasive BAL fluid and also to screen serum or blood as a non-invasive specimen. Albino rats of Wistar or Fischer strains were experimentally infected by Pneumocystis carinii. Extracted DNAs or cell lysates of their blood, bronchoalveolar lavage fluid, and lung homogenate were used as the template DNA. Primers were synthetic oligonucleotides among 16s rDNA sequences. All of the primer combinations gave PCR products, but the primer pair of #24 and #27 gave best quality product of 666 bp. The sensitivity of PCR with lysates of BAL fluid was 57.7% but it increased to 84.6% with extracted DNAs. None of BAL lysate or DNA was positive among 13 microscopically negatives. The serum DNAs were positive only in 2 cases out of 20 morphologically positive rats. DNAs of human, rat, other parasites, yeast, and microorganisms were negative. The findings suggest that the present primers are specific but simple lysate of BAL fluid is not sensitive. PCR may be used as a routine diagnostic method of pneumocystosis if simple and rapid preparation of non-invasive clinical specimens are available.