Despite advances in the treatment of invasive fungal diseases (IFD), mortality rates remain high. Moreover, due to the expanding spectrum of causative agents, fast and accurate pathogen identification is necessary. We designed a panfungal polymerase chain reaction (PCR), which targets the highly variable ITS2 region of rDNA genes and uses high resolution melting analysis (HRM) for subsequent species identification. The sensitivity and specificity of this method was tested on a broad spectrum of the most clinically important fungal pathogens including Aspergillus spp., Candida spp. and mucormycetes. Despite the fact that fluid from bronchoalveolar lavage (BAL) is one of the most frequently tested materials there is a lack of literature sources aimed at panfungal PCR as an IFD diagnostic tool from BAL samples. The applicability of this method in routine practice was evaluated on 104 BAL samples from immunocompromised patients. Due to high ITS region variability, we obtained divergent melting peaks for different fungal species. Thirteen out of 18 patients with proven or probable IFD were positive. Therefore, the sensitivity, specificity, positive predictive value and negative predictive value of our method were 67%, 100%, 100%, and 94%, respectively. In our assay, fungal pathogens identification is based on HRM, therefore omitting the expensive and time consuming sequencing step. With the high specificity, positive and negative predictive values, short time needed to obtain a result, and low price, the presented assay is intended to be used as a quick screening method for patients at risk of IFD.

Department of Clinical Microbiology University Hospital Brno Brno Czech Republic

Department of Internal Medicine Hematology and Oncology University Hospital Brno Brno Czech Republic

Department of Internal Medicine Hematology and Oncology University Hospital Brno Brno Czech Republic Department of Internal Medicine Hematology and Oncology Faculty of Medicine Masaryk University Brno Czech Republic

Department of Internal Medicine Hematology and Oncology University Hospital Brno Brno Czech Republic Department of Internal Medicine Hematology and Oncology Faculty of Medicine Masaryk University Brno Czech Republic CEITEC Central European Institute of Technology Masaryk University Brno Czech Republic

Department of Mycology HPL Ltd Bratislava Slovakia

Department of Oncohematology Comenius University in Bratislava and National Cancer Institute Bratislava Slovakia

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Detection and identification of fungi in bronchoalveolar lavage fluid from immunocompromised patients using panfungal PCR

First case of invasive Magnusiomyces capitatus infection in Slovakia