Milène Sasso

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 HIV-positive or other immunocompromised patients. In the past several years, PCR on pulmonary samples has become an essential element for the laboratory diagnosis of PCP. Nevertheless, very few comparative studies of available PCR assays have been published. In this work, we evaluated the concordance between four real-time PCR assays, including three commercial kits, AmpliSens, MycAssay, and Bio-Evolution PCR, and an in-house PCR (J. Fillaux et al. 2008, J Microbiol Methods 75:258–261, doi:http://dx.doi.org/10.1016/j.mimet.2008.06.009), on 148 pulmonary samples. The results showed concordance rates ranging from 81.6% to 96.6% (kappa, 0.64 to 0.93). Concordance was excellent between three assays: the in-house assay, AmpliSens, and the MycAssay PCR (kappa, >0.8). The 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.

Changes in the distribution of colonising and infecting Candida spp. isolates, antifungal drug consumption and susceptibility in a French intensive care unit: A 10‐year study

Monitoring fungal ecology and resistance to antifungal agents within intensive care units (ICU) is essential for the management of invasive fungal infections. Therefore, a retrospective descriptive study was carried in the ICU of Nimes University Hospital, France, from 2007 to 2016. As the majority of invasive fungal infections in ICU are caused by Candida species, the study objectives were to describe Candida species distribution, to assess candidaemia incidence and to monitor the antifungal drug susceptibility of Candida isolates and the consumption of antifungal agents. Among the recorded invasive Candida infections (n=244), 43% were intra‐abdominal and 22% bloodstream infections. Candida albicans was the most frequent species (55.8%), followed by Candida glabrata (14.1%), Candida tropicalis (10%), Candida parapsilosis (8%) and Candida krusei (5.3%). Candidaemia incidence was 4.49 per 1000 admissions. The mean consumption of antifungal agents was of 170.5 defined daily doses (DDD) for 1000 hospital days (HD) per year. Changes in antifungal drug consumption were observed, with an increased use of echinocandins (from 17.96 DDD/1000 HD in 2007 to 48.76 DDD/1000 HD in 2016), and the total treatment cost tripled during the study period. No significant change in fungal ecology or in the emergence of resistant species was observed; indeed, only 1.1% of isolates presented an unusual resistance to antifungal agents.

Rapid emergence of FKS mutations in Candida glabrata isolates in a peritoneal candidiasis

We report a rapid acquisition of echinocandin resistance after 12 days of micafungin treatment, without prior exposure, in a patient with peritoneal candidiasis due to C. glabrata. Isolates recovered before and after treatment were compared by multilocus sequence typing. Results of antifungal susceptibility testing and FKS mutations were reported. The interest of repeating antifungal susceptibility testing for echinocandin molecules during the treatment is discussed and a strategy to research FKS mutations proposed.

Direct identification of molds by sequence analysis in fungal chronic rhinosinusitis

Fungi are widely implicated in chronic rhinosinusitis. Direct microscopic examination (DME) is used to confirm the biological diagnosis of fungal rhinosinusitis (FRS). Diagnostic sensitivity of DME is better than culture, however DME does not allow fungal species identification. In this study, we included 54 sinus samples demonstrating hyphae on DME. Direct sequencing was compared to culture for the identification of the fungal species. Sequence analysis identified fungi in 81.5% of cases while culture was positive in only 31.5%. The most common genus was Aspergillus and the identified species belonged to section Fumigati or to section Flavi. Among other fungi identified by sequence analysis, Schizophyllum commune was present in three samples attesting to the importance of this Basidiomycetes in FRS. Our results clearly demonstrate the superiority of sequencing compared to culture when performed on specimens with hyphal elements at DME, and contributes to the epidemiological knowledge of fungi involved in FRS.

Pityriasis capitis et dermatite séborrhéique du cuir chevelu : rôle du laboratoire dans l’évaluation d’une implication fongique

Resume Le pityriasis capitis gras et la dermatite seborrheique sont les deux principaux contextes cliniques au cours desquels le biologiste est appele a confirmer un diagnostic de «levurose» du cuir chevelu. La prevalence mondiale de la dermatite seborrheique, plus frequente dans la population âgee, serait de 2 a 3 %, avec une predominance masculine. Celle du pityriasis capitis gras depasserait 50 % de la population generale post-pubertaire et d’âge moyen. L’examen du cuir chevelu sous rayonnement ultra-violet (lampe de Wood) met en evidence l’activite metabolique du champignon, revelee par des placards de tailles diverses, fluorescents jaune verdâtre (teinte «bronze»), mais sans fluorescence des cheveux eux-memes. Deux especes, Malassezia globosa et M. restricta, semblent aujourd’hui plus etroitement associees a ces formes cliniques. L’examen biologique permettra d’evaluer le niveau d’implication et la densite relative du champignon sur le cuir chevelu. Les modalites de prelevement sont essentielles a connaitre et conditionnent la qualite de l’examen biologique qui est base sur un examen direct bien conduit.