Detection of <i>Pneumocystis jirovecii</i> in oral wash from immunosuppressed patients as a diagnostic tool

<div><p>Background</p><p>Diagnosis of <i>Pneumocystis jirovecii</i> (PJ) pneumonia ordinarily requires invasive procedures that could be avoided by PCR methodologies, if these could be designed with adequate cut-off values for confounding background carriage.</p><p>Methods</p><p>We designed a novel quantitative real-time PCR assay to detect the mitochondrial large subunit rRNA gene of PJ in oral washes. To benchmark levels of PJ carriage versus infection, we tested asymptomatic immunosuppressed patients including Danish (n = 88) and West African HIV-infected (n = 142) patients, renal transplant recipients (n = 51), rheumatologic patients (n = 102), patients with inflammatory bowel diseases (n = 98), and healthy blood donors (controls, n = 50). The fungal burden in patients with PJ pneumonia (PCP, n = 7) was also investigated.</p><p>Results</p><p>Danish HIV-infected patients (with viremia/low CD4) and recent transplant recipients were at most risk of being carriers (prevalence of 23% and 16.7% respectively), whereas PJ was rarely detected among rheumatologic patients, patients with inflammatory bowel diseases, and untreated West African HIV patients. PJ was not detected among healthy controls. The fungal burden in patients with PCP fell rapidly on treatment.</p><p>Conclusions</p><p>The quantitative PCR method described could conceivably discriminate between carriage and disease, given suitable threshold values for the former, and predict treatment efficacy by measures of the fungal burden in daily oral washes.</p></div

Renal transplant recipients.

<p>Demographics.</p

The standard curve shows linearity over a large dynamic detection range (from 3 to 30.000.000 copies) of the PCR assay.

<p>Mean and errorbars of one standard deviation are shown All negative (water) and positive controls (the standard in dilution 10⁻⁸) performed as expected. Inhibition of the PCR reaction occurred with only one patient, suspected to have PCP. The RNase P assay showed only minor variations between samples and standardization was not necessary. Two samples from a blood donor and a renal transplant patient were RNase P negative and were excluded.</p

HIV patients with PCP with follow-up.

<p>HIV patients with PCP with follow-up.</p

HIV patients.

<p>Demographics.</p