SUMMARY
Real-time PCR has revolutionized the way clinical microbiology laboratories diagnose many human microbial infections. This testing method combines PCR chemistry with fluorescent probe detection of
amplified product in the same reaction vessel. In general, both PCR and
amplified product detection are completed in an hour or less, which is
considerably faster than conventional PCR detection methods. Real-time
PCR assays provide sensitivity and specificity equivalent to that of
conventional PCR combined with Southern blot analysis, and since
amplification and detection steps are performed in the same closed
vessel, the risk of releasing amplified nucleic acids into the
environment is negligible. The combination of excellent sensitivity and
specificity, low contamination risk, and speed has made real-time PCR
technology an appealing alternative to culture- or immunoassay-based
testing methods for diagnosing many infectious diseases. This review
focuses on the application of real-time PCR in the clinical
microbiology laboratory.
Pneumocystis carinii pneumonia has emerged as a significant cause of morbidity and mortality in immunocompromised patients with and without AIDS. To determine differences in P. carinii pneumonia in patients with and without AIDS, the P. carinii parasite numbers, lung inflammatory cell populations, gas exchange, and survival were assessed in a series of 75 consecutive patients with P. carinii pneumonia. Bronchoalveolar lavage was used to quantify the parasite and inflammatory cell numbers in these patients. The data from this study indicate: (1) patients with P. carinii pneumonia and AIDS have significantly greater numbers of P. carinii per ml of lavage compared to other immunocompromised patients with P. carinii pneumonia (p less than 0.0001); (2) patients with P. carinii pneumonia and AIDS also have significantly fewer neutrophils recovered in the lavage compared to other immunocompromised patients with P. carinii pneumonia (p = 0.0001); (3) patients with AIDS and P. carinii pneumonia have higher arterial oxygen tensions than those patients with P. carinii pneumonia in conditions other than AIDS (p = 0.008); and (4) increased lavage neutrophils (rather than parasite number) correlate with poorer oxygenation and poorer patient survival (p = 0.01). This investigation demonstrates substantial differences in lung inflammation and parasite number during P. carinii pneumonia in patients with and without AIDS. The data further suggest that lung inflammation contributes substantially to respiratory impairment in patients with P. carinii pneumonia.
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