Rapid detection of fungal pathogens in bronchoalveolar lavage samples using panfungal PCR combined with high resolution melting analysis
Language English Country England, Great Britain Media print-electronic
Document type Journal Article
PubMed
27161789
DOI
10.1093/mmy/myw032
PII: myw032
Knihovny.cz E-resources
- Keywords
- Bronchoalveolar lavage fluid, High resolution melting analysis, Immunocompromised patients, Panfungal PCR,
- MeSH
- Bronchoalveolar Lavage Fluid microbiology MeSH
- Time Factors MeSH
- Molecular Diagnostic Techniques methods MeSH
- DNA, Fungal chemistry genetics MeSH
- Fungi classification genetics isolation & purification MeSH
- Humans MeSH
- DNA, Ribosomal Spacer chemistry genetics MeSH
- Lung Diseases, Fungal diagnosis microbiology MeSH
- Polymerase Chain Reaction methods MeSH
- Predictive Value of Tests MeSH
- Sensitivity and Specificity MeSH
- Transition Temperature * MeSH
- Check Tag
- Humans MeSH
- Male MeSH
- Female MeSH
- Publication type
- Journal Article MeSH
- Names of Substances
- DNA, Fungal MeSH
- DNA, Ribosomal Spacer MeSH
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
References provided by Crossref.org
First case of invasive Magnusiomyces capitatus infection in Slovakia
