Performances of Four Real-Time PCR Assays for Diagnosis of Pneumocystis jirovecii Pneumonia

Abstract

Pneumonia due to Pneumocystis jirovecii (PCP) is a frequent infection among HIVThe performances of these PCR assays were also evaluated according to the classification of the probability of PCP (proven, probable, possible, or no final diagnosis of PCP) based on clinical and radiological signs as well as on the direct examination of bronchoalveolar lavage samples. In the proven PCP category, Pneumocystis jirovecii DNA was detected with all four assays. In the probable PCP category, the in-house PCR, AmpliSens, and the MycAssay PCR were positive for all samples, while the Bio-Evolution PCR failed to detect Pneumocystis jirovecii DNA in two samples. In the possible PCP category, the percentage of positive samples according to PCR varied from 54.5% to 86.4%. Detection of colonized patients is discussed. Finally, among the four evaluated PCR assays, one was not suitable for colonization detection but showed good performance in the proven and probable PCP groups. For the three other assays, performances were excellent and allowed detection of a very low fungal burden.

A New Age in Molecular Diagnostics for Invasive Fungal Disease: Are We Ready?

“…Several commercial qPCR assays for PCP detection are available but there are few comparative studies. A performance evaluation of three kits (AmpliSens P. jirovecii-FRT, MycAssay Pneumocystis from Myconostica, and Bio-Evolution P. jirovecii PCR) together with an in-house assay targeting the major surface glycoprotein gene found excellent concordance between the in-house assay, Ampli-Sens, and MycAssay (kappa, > 0.8), with all three assays confirming the diagnosis in 100% of proven (n = 12) and probable (n = 25) PCP groups, compared with 100 and 92%, respectively, of cases confirmed by the Bio-Evolution assay (Sasso et al, 2016). The percentage of positive results was more variable for the "possible PCP" category, ranging from 54.5% (Bio-Evolution) to 86.4% (AmpliSens PCR), but all four assays were effective for PCP diagnosis.…”

Diagnostic workup for ARDS patients

“…The first step is a direct microscopic examination of BAL fluid, which allows one to look for bacteria using Gram staining, but also to use Giemsa-modified staining which offers a number of advantages over Gram staining, including better visualization of host cell morphology (including viral cytopathic inclusions [12]), improved detection of bacteria, particularly intracellular bacteria, and detection of some protozoan and fungal pathogens [19]. The second step is to culture for bacteria using quantitative culture of BAL fluid to test for respiratory viruses (Table 1), Pneumocystis or other fungi such as Aspergillus using PCR [20,21] (see Table 1). Moreover, specific staining (immunostaining or immunofluorescence) may be used to detect Legionella pneumophila, Pneumocystis jirovecii, Toxoplasma gondii and other pathogens responsible for pneumonia in immunocompetent or immunosuppressed patients.…”

A Molecular Window into the Biology and Epidemiology of Pneumocystis spp

“…In recent years, multiple PCR assays have been developed for detection of P. jirovecii, most commonly utilizing primers for the mitochondrial large-subunit rRNA gene (mtLSU) or the multicopy major surface glycoprotein (msg) gene family (39)(40)(41)(42)(43)(44)(45)(46), and these assays appear to be 10 to 100 times more sensitive than microscopic detection of stained organisms. Several commercially available diagnostic PCR kits are approved for clinical use in a number of countries, primarily in Europe (47)(48)(49)(50); none are currently FDA approved for use in the United States.…”