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Accordingly, we could finally provide a definitive interpretation of qPCR results for 96% (267/278) of samples (Table?3). A fungal load >1900?TFEq/mL allowed us to conclude that there was ongoing PCP in 18 samples (6.5%). A fungal load Ceftiofur Since the 1990s, PCR has markedly improved the sensitivity of detection, revealing a new population of individuals in which detectable Pneumocystis DNA is present, but few or no organisms are visualized [13]. Indeed, the risk of false-positivity has been suggested, but it has been dramatically reduced with one-step processing techniques such as real-time PCR, which greatly limit specimen processing and avoid the risk of DNA carryover. The terms colonization, asymptomatic infection 3-Methyladenine in vitro and subclinical infection have been used to describe the presence of P.?jirovecii DNA in a host without the occurrence of clinical pneumonia [13], but, up to now, there has been no clear biological discrimination between colonization and disease. Here, we show in a large cohort of immunocompromised patients that this discrimination can be based on the estimation of fungal burdens in BAL fluids or IS with the use of real-time qPCR. This assumption is based on a prospective study performed in HIV-infected and non-HIV-infected patients in whom the high or low probability of PCP was estimated independently of qPCR tests Sunitinib mouse performed on BAL fluids and IS. Marked differences were found between the two groups of patients. The high-probability PCP group was characterized by a positive qPCR result, with high fungal burdens, and by the presence of P.?jirovecii on IIF smears in 81.3% of patients. In contrast, no low-probability PCP patients had P.?jirovecii by microscopy, and only 13.5% of them were qPCR-positive. Interestingly, there was no significant difference in fungal DNA load between IS and BAL fluid samples in the two groups of PCP patients, underlining the marked potential of qPCR on IS for the diagnosis of PCP [20]; however, as both IS and BAL fluids could be sampled in only five patients, this assertion has to be confirmed. The sensitivity and specificity of PCR on BAL fluids or IS for diagnosis and for discrimination between PCP and colonization were estimated with different qPCR cut-off values. As there was no significant difference in fungal DNA load between IS and BAL fluid samples, similar cut-offs were applicable for BAL fluid and IS samples.