Intercomparison of Two Fluorescent Dyes to Visualize Parasitic Fungi (Chytridiomycota) on Phytoplankton

• Isabell Klawonn ^{[6] [2]} ; Susanne Dunker ^{[7] [3]} ; Maiko Kagami ^{[1] [4]} ; Hans-Peter Grossart ^{[6] [8]} ; Silke Van den Wyngaert ^{[6] [9] [5]}
  1. [1] Toho University

     Toho University

     Japón

     
  2. [2] Leibniz Institute for Baltic Sea Research

     Leibniz Institute for Baltic Sea Research

     Kreisfreie Stadt Rostock, Alemania

     
  3. [3] German Center for Integrative Biodiversity Research

     German Center for Integrative Biodiversity Research

     Kreisfreie Stadt Leipzig, Alemania

     
  4. [4] Yokohama National University

     Yokohama National University

     Naka Ku, Japón

     
  5. [5] University of Turku

     University of Turku

     Turku, Finlandia

     
  6. [6] Department of Experimental Limnology, Leibniz-Institute of Freshwater Ecology and Inland Fisheries (IGB), 12587, Berlin, Germany
  7. [7] Department for Physiological Diversity, Helmholtz Centre for Environmental Research (UFZ), 04318, Leipzig, Germany
  8. [8] Institute of Biochemistry and Biology, Potsdam University, 14476, Potsdam, Germany
  9. [9] WasserCluster Lunz, Biologische Station, Dr. Carl Kupelwieser Promenade 5, 3293, Lunz am See, Austria

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• Localización: Microbial ecology, ISSN-e 1432-184X, ISSN 0095-3628, Vol. 85, Nº. 1, 2023, págs. 9-23
• Idioma: inglés
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  • Fungal microparasites (here chytrids) are widely distributed and yet, they are often overlooked in aquatic environments. To facilitate the detection of microparasites, we revisited the applicability of two fungal cell wall markers, Calcofluor White (CFW) and wheat germ agglutinin (WGA), for the direct visualization of chytrid infections on phytoplankton in laboratory-maintained isolates and field-sampled communities. Using a comprehensive set of chytrid–phytoplankton model pathosystems, we verified the staining pattern on diverse morphological structures of chytrids via fluorescence microscopy. Empty sporangia were stained most effectively, followed by encysted zoospores and im-/mature sporangia, while the staining success was more variable for rhizoids, stalks, and resting spores. In a few instances, the staining was unsuccessful (mostly with WGA), presumably due to insufficient cell fixation, gelatinous cell coatings, and multilayered cell walls. CFW and WGA staining could be done in Utermöhl chambers or on polycarbonate filters, but CFW staining on filters seemed less advisable due to high background fluorescence. To visualize chytrids, 1 µg dye mL−1 was sufficient (but 5 µg mL−1 are recommended). Using a dual CFW–WGA staining protocol, we detected multiple, mostly undescribed chytrids in two natural systems (freshwater and coastal), while falsely positive or negative stained cells were well detectable. As a proof-of-concept, we moreover conducted imaging flow cytometry, as a potential high-throughput technology for quantifying chytrid infections. Our guidelines and recommendations are expected to facilitate the detection of chytrid epidemics and to unveil their ecological and economical imprint in natural and engineered aquatic systems.