Quantitative Pneumocystis jirovecii real-time PCR to differentiate disease from colonisation

We studied a Pneumocystis jirovecii quantitative polymerase chain reaction (qPCR) for distinguishing P. jirovecii disease from colonisation. Eighty-two respiratory samples from 65 patients with qPCR results were analysed against a gold standard clinical diagnosis of Pneumocystis pneumonia. High inter-assay reproducibility using recombinant and clinical material was observed. Contemporaneous samples from the same patient displayed high variability (median difference 2.6 log₁₀ copies/mL, IQR 2.1-3.1 log₁₀ copies/mL). Despite this, area under the receiver operator characteristic curve was 0.8. An optimum cut-off of 2.8 log₁₀ copies/mL (equivalent to C_T of 34.0 cycles) had 59% sensitivity and 92% specificity. The median P. jirovecii load was 7.3 log₁₀ copies/mL in HIV patients compared to 2.6 log₁₀ copies/mL in non-HIV patients. Specificity was 100% in non-HIV patients with qPCR of >3.8 log₁₀ copies/mL. qPCR was useful for distinguishing P. jirovecii disease from colonisation. A quantitative standard, standardisation of definitions and methods are required to improve the generalisability of results.