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Labelling of Candida auris Cell Walls to Examine Levels of PAMP Exposure by Flow Cytometry and Fluorescence Microscopy

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Candida auris

Abstract

Pathogen-associated molecular patterns (PAMPs) of the fungal cell wall are primary targets for the innate immune system of animals. Therefore, characterizing PAMP exposure of fungal pathogens helps to elucidate how they interact with their hosts at a molecular level. Fluorescent labelling can be used to monitor exposure of multiple fungal cell wall PAMPs in a single experiment. Here, we describe a protocol to simultaneously label chitin, mannan, and β-1,3-glucan in Candida auris to study these PAMPs by fluorescence microscopy and allow high-throughput examination of their exposure by flow cytometry.

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References

  1. Satoh K, Makimura K, Hasumi Y et al (2009) Candida auris sp. nov., a novel ascomycetous yeast isolated from the external ear canal of an inpatient in a Japanese hospital. Microbiol Immunol 53:41–44. https://doi.org/10.1111/j.1348-0421.2008.00083.x

    Article  PubMed  CAS  Google Scholar 

  2. Forsberg K, Woodworth K, Walters M et al (2019) Candida auris: the recent emergence of a multidrug-resistant fungal pathogen. Med Mycol 57:1–12. https://doi.org/10.1093/mmy/myy054

    Article  PubMed  Google Scholar 

  3. Hopke A, Brown AJP, Hall RA, Wheeler RT (2018) Dynamic fungal cell wall architecture in stress adaptation and immune evasion. Trends Microbiol 26:284–295. https://doi.org/10.1016/j.tim.2018.01.007

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  4. Gow NAR, Latge J-P, Munro CA (2017) The fungal cell wall: structure, biosynthesis, and function. Microbiol Spectr 5(3):FUNK-0035-2016. https://doi.org/10.1128/microbiolspec.FUNK-0035-2016

    Article  Google Scholar 

  5. Navarro-Arias MJ, Hernández-Chávez MJ, García-Carnero LC et al (2019) Differential recognition of Candida tropicalis, Candida guilliermondii, Candida krusei, and Candida auris by human innate immune cells. Infect Drug Resist 12:783–794. https://doi.org/10.2147/IDR.S197531

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  6. Bruno M, Kersten S, Bain JM et al (2020) Transcriptional and functional insights into the host immune response against the emerging fungal pathogen Candida auris. Nat Microbiol 5:1516–1531. https://doi.org/10.1038/s41564-020-0780-3

    Article  PubMed  CAS  Google Scholar 

  7. McKinnon KM (2018) Flow cytometry: an overview. Curr Protoc Immunol 120:5.1.1–5.1.11. https://doi.org/10.1002/cpim.40

    Article  Google Scholar 

  8. Macey MG (2007) Flow cytometry: principles and applications. Humana Press, Totowa

    Book  Google Scholar 

  9. Pawley JB (2006) Handbook of biological confocal microscopy, 3rd edn. Springer, Boston

    Book  Google Scholar 

  10. Walker LA, Munro CA, de Bruijn I et al (2008) Stimulation of chitin synthesis rescues Candida albicans from echinocandins. PLoS Pathog 4:e1000040. https://doi.org/10.1371/journal.ppat.1000040

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  11. Sherrington SL, Sorsby E, Mahtey N et al (2017) Adaptation of Candida albicans to environmental pH induces cell wall remodelling and enhances innate immune recognition. PLoS Pathog 13:e1006403. https://doi.org/10.1371/journal.ppat.1006403

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  12. Ballou ER, Avelar GM, Childers DS et al (2016) Lactate signalling regulates fungal β-glucan masking and immune evasion. Nat Microbiol 2:16238. https://doi.org/10.1038/nmicrobiol.2016.238

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  13. Pradhan A, Mol Avelar G, Bain JM et al (2019) Non-canonical signalling mediates changes in fungal cell wall PAMPs that drive immune evasion. Nat Commun 10:5315. https://doi.org/10.1038/s41467-019-13298-9

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  14. Mora-Montes HM, Netea MG, Ferwerda G et al (2011) Recognition and blocking of innate immunity cells by Candida albicans chitin. Infect Immun 79:1961–1970. https://doi.org/10.1128/IAI.01282-10

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  15. Ene IV, Adya AK, Wehmeier S et al (2012) Host carbon sources modulate cell wall architecture, drug resistance and virulence in a fungal pathogen. Cell Microbiol 14:1319–1335. https://doi.org/10.1111/j.1462-5822.2012.01813.x

    Article  PubMed  PubMed Central  CAS  Google Scholar 

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Acknowledgments

This work is supported by the Medical Research Council (MRC) Centre for Medical Mycology at the University of Exeter, UK [grant number MR/P501955/1]. The authors are grateful for the support and technical assistance from the Microscopy and Histology Core Facility (Kevin MacKenzie, Lucy Wight) and the Iain Fraser Cytometry Centre (Andrea Holme, Kate Burgoyne) of the University of Aberdeen.

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Correspondence to Alexander Lorenz .

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Pelletier, C., Lorenz, A. (2022). Labelling of Candida auris Cell Walls to Examine Levels of PAMP Exposure by Flow Cytometry and Fluorescence Microscopy. In: Lorenz, A. (eds) Candida auris. Methods in Molecular Biology, vol 2517. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2417-3_14

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  • DOI: https://doi.org/10.1007/978-1-0716-2417-3_14

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-2416-6

  • Online ISBN: 978-1-0716-2417-3

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