Abstract
BackgroundAcquired resistance to antifungal agents now supports the introduction of susceptibility testing for species-drug combinations for which this was previously thought unnecessary. For pathogenic yeasts, conventional phenotypic testing needs at least 24 h. Culture on a porous aluminum oxide (PAO) support combined with microscopy offers a route to more rapid results.MethodsMicrocolonies of Candida species grown on PAO were stained with the fluorogenic dyes Fun-1 and Calcofluor White and then imaged by fluorescence microscopy. Images were captured by a charge-coupled device camera and processed by publicly available software. By this method, the growth of yeasts could be detected and quantified within 2 h. Microcolony imaging was then used to assess the susceptibility of the yeasts to amphotericin B, anidulafungin and caspofungin (3.5 h culture), and voriconazole and itraconazole (7 h culture).SignificanceOverall, the results showed good agreement with EUCAST (86.5% agreement; n = 170) and E-test (85.9% agreement; n = 170). The closest agreement to standard tests was found when testing susceptibility to amphotericin B and echinocandins (88.2 to 91.2%) and the least good for the triazoles (79.4 to 82.4%). Furthermore, large datasets on population variation could be rapidly obtained. An analysis of microcolonies revealed subtle effects of antimycotics on resistant strains and below the MIC of sensitive strains, particularly an increase in population heterogeneity and cell density-dependent effects of triazoles. Additionally, the method could be adapted to strain identification via germ tube extension. We suggest PAO culture is a rapid and versatile method that may be usefully adapted to clinical mycology and has research applications.
Highlights
Pathogenic fungi are a major source of human infections and are a serious risk factor for mortality
Microcolony imaging on porous aluminum oxide (PAO) allowed the creation of rapid culture-based assays and the minimal inhibitory concentrations (MICs) testing on Candida on surfaces may be more relevant to how these yeasts exist in the human body than planktonic growth, as used in broth dilution methods
Poor nutrient environments with oxygen limitation and the potential for biofilm development can be created on PAO, which may allow further development to increase the relevance of MIC testing in vitro to clinical outcomes
Summary
Pathogenic fungi are a major source of human infections and are a serious risk factor for mortality. Limitations for molecular testing include multiple mutations conferring resistance and the lack of suitable genetic markers for some drugs [6]. These considerations suggest a need for further development of rapid, culture-based susceptibility testing in fungi. For yeasts and filamentous fungi, a number of rapid phenotypic methods have been developed. These include culture in microwells containing broth with antimycotics and colorimetric redox indicators [7,8,9]. Acquired resistance to antifungal agents supports the introduction of susceptibility testing for speciesdrug combinations for which this was previously thought unnecessary. Culture on a porous aluminum oxide (PAO) support combined with microscopy offers a route to more rapid results
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
