Date Published: March 30, 2017
Publisher: Public Library of Science
Author(s): Cecilie Juul Hviid, Marianne Lund, Allan Sørensen, Svend Ellermann- Eriksen, Bente Jespersen, Mette Yde Dam, Jens Frederik Dahlerup, Thomas Benfield, Sanne Jespersen, Lars Jørgen Østergaard, Alex Lund Laursen, Roberto F. Speck.
Diagnosis of Pneumocystis jirovecii (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.
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.
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.
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.
The opportunistic pathogen Pneumocystis jirovecii (PJ) is a well-known cause of severe Pneumocystis pneumonia (PCP) among immunosuppressed patients, particularly those infected with HIV [1, 2]. PCP also occurs in non-HIV immunosuppressed patients, including those with hematologic malignancies and transplant recipients. Recently it has become a significant opportunistic infection in patients receiving immunosuppressive medication for autoimmune or inflammatory diseases [3–5].
In conclusion, we have developed a sensitive quantitative PCR assay with a large dynamic range and applied the method to characterize the level of carriage in different groups of asymptomatic immunosuppressed patients and the dynamics in patients with PCP using oral wash. Carriage was most frequently seen in renal transplanted patients within the first 6 months after operation and in Danish HIV infected patients with viremia combined with low CD4 count, whereas carriage was less frequent among untreated HIV infected patients from Guinea Bissau and patients on immunosuppressive therapy. A fungal load of >1000 copies was suggestive of PCP, whereas a copy number below 30 was seen in asymptomatic patients, who did not develop symptomatic disease on follow up. A grey zone was seen between these 2 extremes suggesting that the method should be used with caution until larger studies have been undertaken.