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Canadian Journal of Infectious Diseases and Medical Microbiology
Volume 26, Issue 5, Pages 263-267
http://dx.doi.org/10.1155/2015/138787
Original Article

Development and Validation of a Pneumocystis jirovecii Real-time Polymerase Chain Reaction Assay for Diagnosis of Pneumocystis Pneumonia

Deirdre L Church,1,2,3 Anshula Ambasta,3 Amanda Wilmer,4 Holly Williscroft,1 Gordon Ritchie,4,5 Dylan R Pillai,1,2,3 Sylvie Champagne,4,5 and Daniel G Gregson1,2,3

1Division of Microbiology, Calgary Laboratory Services, Departments of Pathology & Laboratory Medicine, Calgary, Alberta, Canada
2Division of Medical Microbiology, Alberta Health Services and the University of Calgary, Calgary, Alberta, Canada
3Division of Medical Microbiology and Medicine, Alberta Health Services and the University of Calgary, Calgary, Alberta, and Department of Pathology & Laboratory Medicine, the University of British Columbia, Vancouver, British Columbia, Canada
4Division of Medical Microbiology, the University of British Columbia, Vancouver, British Columbia, Canada
5St Paul’s Hospital and the University of British Columbia, Vancouver, British Columbia, Canada

Copyright © 2015 Hindawi Publishing Corporation. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

BACKGROUND: Pneumocystis jirovecii (PJ), a pathogenic fungus, causes severe interstitial Pneumocystis pneumonia (PCP) among immunocompromised patients. A laboratory-developed real-time polyermase chain reaction (PCR) assay was validated for PJ detection to improve diagnosis of PCP.

METHODS: Forty stored bronchoalveolar lavage (BAL) samples (20 known PJ positive [PJ+] and 20 known PJ negative [PJ−]) were initially tested using the molecular assay. Ninety-two sequentially collected BAL samples were then analyzed using an immunofluorescence assay (IFA) and secondarily tested using the PJ real-time PCR assay. Discrepant results were resolved by retesting BAL samples using another real-time PCR assay with a different target. PJ real-time PCR assay performance was compared with the existing gold standard (ie, IFA) and a modified gold standard, in which a true positive was defined as a sample that tested positive in two of three methods in a patient suspected to have PCP.

RESULTS: Ninety of 132 (68%) BAL fluid samples were collected from immunocompromised patients. Thirteen of 92 (14%) BALs collected were PJ+ when tested using IFA. A total of 40 BAL samples were PJ+ in the present study including: all IFA positive samples (n=13); all referred PJ+ BAL samples (n=20); and seven additional BAL samples that were IFA negative, but positive using the modified gold standard. Compared with IFA, the PJ real-time PCR had sensitivity, specificity, and positive and negative predictive values of 100%, 91%, 65% and 100%, respectively. Compared with the modified gold standard, PJ real-time PCR had a sensitivity, specificity, and positive and negative predictive values of 100%.

CONCLUSION: PJ real-time PCR improved detection of PJ in immunocompromised patients.