Skip to main content
SpringerLink
Log in

Cell Viability Assays for Candida auris

  • Protocol
  • First Online:
Candida auris

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2517))

Abstract

Cell viability assays are useful for assessing the efficacy of antifungal therapeutics and disinfection strategies in vitro. In recent years these assays have been fundamental for the testing of conventional and novel therapies against the nosocomial fungal pathogen Candida auris. Here we provide detailed descriptions of methods for assessing cellular viability of Candida auris in vitro, such as metabolic assays (XTT and resazurin), colony-forming unit counting, live/dead quantitative PCR, and fluorescent staining for microscopic analyses.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Log in via an institution

Subscribe and save

Springer+ Basic
$34.99 /Month
  • Get 10 units per month
  • Download Article/Chapter or eBook
  • 1 Unit = 1 Article or 1 Chapter
  • Cancel anytime
Subscribe now

Buy Now

Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 129.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 169.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 249.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

Similar content being viewed by others

References

  1. Ramage G, Vande Walle K, Wickes BL, Lopez-Ribot JL (2001) Standardized method for in vitro antifungal susceptibility testing of Candida albicans biofilms. Antimicrob Agents Chemother 45(9):2475–2479

    Article  CAS  Google Scholar 

  2. Pierce CG, Uppuluri P, Tristan AR, Wormley FL Jr, Mowat E, Ramage G, Lopez-Ribot JL (2008) A simple and reproducible 96-well plate-based method for the formation of fungal biofilms and its application to antifungal susceptibility testing. Nat Protoc 3(9):1494–1500

    Article  CAS  Google Scholar 

  3. Pierce CG, Uppuluri P, Tummala S, Lopez-Ribot JL (2010) A 96 well microtiter plate-based method for monitoring formation and antifungal susceptibility testing of Candida albicans biofilms. J Vis Exp 44:2287

    Google Scholar 

  4. Dalecki AG, Crawford CL, Wolschendorf F (2016) Targeting biofilm associated staphylococcus aureus using resazurin based drug-susceptibility assay. J Vis Exp 111:53925

    Google Scholar 

  5. Van den Driessche F, Rigole P, Brackman G, Coenye T (2014) Optimization of resazurin-based viability staining for quantification of microbial biofilms. J Microbiol Methods 98:31–34

    Article  CAS  Google Scholar 

  6. Loozen G, Boon N, Pauwels M, Quirynen M, Teughels W (2011) Live/dead real-time polymerase chain reaction to assess new therapies against dental plaque-related pathologies. Mol Oral Microbiol 26(4):253–261

    Article  CAS  Google Scholar 

  7. Agusti G, Fittipaldi M, Morato J, Codony F (2013) Viable quantitative PCR for assessing the response of Candida albicans to antifungal treatment. Appl Microbiol Biotechnol 97(1):341–349

    Article  CAS  Google Scholar 

  8. Boulos L, Prevost M, Barbeau B, Coallier J, Desjardins R (1999) LIVE/DEAD BacLight : application of a new rapid staining method for direct enumeration of viable and total bacteria in drinking water. J Microbiol Methods 37(1):77–86

    Article  CAS  Google Scholar 

  9. Millard PJ, Roth BL, Thi HP, Yue ST, Haugland RP (1997) Development of the FUN-1 family of fluorescent probes for vacuole labeling and viability testing of yeasts. Appl Environ Microbiol 63(7):2897–2905

    Article  CAS  Google Scholar 

  10. Satoh K, Makimura K, Hasumi Y, Nishiyama Y, Uchida K, Yamaguchi H (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(1):41–44

    Article  CAS  Google Scholar 

  11. Lockhart SR, Etienne KA, Vallabhaneni S, Farooqi J, Chowdhary A, Govender NP, Colombo AL, Calvo B, Cuomo CA, Desjardins CA, Berkow EL, Castanheira M, Magobo RE, Jabeen K, Asghar RJ, Meis JF, Jackson B, Chiller T, Litvintseva AP (2017) Simultaneous emergence of multidrug-resistant Candida auris on 3 continents confirmed by whole-genome sequencing and epidemiological analyses. Clin Infect Dis 64(2):134–140

    Article  CAS  Google Scholar 

  12. Kean R, Ramage G (2019) Combined antifungal resistance and biofilm tolerance: the global threat of Candida auris. mSphere 4(4):e00458–e00419

    Article  CAS  Google Scholar 

  13. Hager CL, Larkin EL, Long L, Zohra Abidi F, Shaw KJ, Ghannoum MA (2018) In vitro and in vivo evaluation of the antifungal activity of APX001A/APX001 against Candida auris. Antimicrob Agents Chemother 62(3):e02319–e02317

    Article  Google Scholar 

  14. de Oliveira HC, Monteiro MC, Rossi SA, Peman J, Ruiz-Gaitan A, Mendes-Giannini MJS, Mellado E, Zaragoza O (2019) Identification of off-patent compounds that present antifungal activity against the emerging fungal pathogen Candida auris. Front Cell Infect Microbiol 9:83

    Article  CAS  Google Scholar 

  15. Rudramurthy SM, Colley T, Abdolrasouli A, Ashman J, Dhaliwal M, Kaur H, Armstrong-James D, Strong P, Rapeport G, Schelenz S, Ito K, Chakrabarti A (2019) In vitro antifungal activity of a novel topical triazole PC945 against emerging yeast Candida auris. J Antimicrob Chemother 74(10):2943–2949

    Article  CAS  Google Scholar 

  16. Arias LS, Butcher MC, Short B, McKloud E, Delaney C, Kean R, Monteiro DR, Williams C, Ramage G, Brown JL (2020) Chitosan Ameliorates Candida auris Virulence in a Galleria mellonella Infection Model. Antimicrob Agents Chemother 64(8):e00476–e00420

    Article  CAS  Google Scholar 

  17. CLSI (2008) Reference method for broth dilution antifungal susceptibility testing of yeasts, 3rd edn. Clinical and Laboratory Standards Institute (CLSI). Document M27-A3.

    Google Scholar 

  18. Allkja J, Bjarnsholt T, Coenye T, Cos P, Fallarero A, Harrison J, Lopes S, Oliver A, Pereira M, Ramage G, Shirtliff M, Stoodley P, Webb J, Zaat S, Goeres D (2020) Minimum information guideline for spectrophotometric and fluorometric methods to assess biofilm formation in microplates. Biofilms 2:100010

    Article  Google Scholar 

  19. Kean R, Sherry L, Townsend E, McKloud E, Short B, Akinbobola A, Mackay WG, Williams C, Jones BL, Ramage G (2017) Surface disinfection challenges for Candida auris: an in vitro study. J Hosp Infect 98(4):433–436

    Article  Google Scholar 

  20. Azeredo J, Azevedo NF, Briandet R, Cerca N, Coenye T, Costa AR, Desvaux M, Di Bonaventura G, Hebraud M, Jaglic Z, Kacaniova M, Knochel S, Lourenco A, Mergulhao F, Meyer RL, Nychas G, Simoes M, Tresse O, Sternberg C (2017) Critical review on biofilm methods. Crit Rev Microbiol 43(3):313–351

    Article  CAS  Google Scholar 

  21. Jose A, Coco BJ, Milligan S, Young B, Lappin DF, Bagg J, Murray C, Ramage G (2010) Reducing the incidence of denture stomatitis: are denture cleansers sufficient? J Prosthodont 19(4):252–257

    Article  Google Scholar 

  22. Mowat E, Butcher J, Lang S, Williams C, Ramage G (2007) Development of a simple model for studying the effects of antifungal agents on multicellular communities of Aspergillus fumigatus. J Med Microbiol 56(9):1205–1212

    Article  CAS  Google Scholar 

  23. Miles AA, Misra SS, Irwin JO (1938) The estimation of the bactericidal power of the blood. J Hyg (Lond) 38(6):732–749

    CAS  Google Scholar 

  24. Kean R, Rajendran R, Haggarty J, Townsend EM, Short B, Burgess KE, Lang S, Millington O, Mackay WG, Williams C, Ramage G (2017) Candida albicans mycofilms support staphylococcus aureus colonization and enhances miconazole resistance in dual-species interactions. Front Microbiol 8:258

    Article  Google Scholar 

  25. Sherry L, Lappin G, O'Donnell LE, Millhouse E, Millington OR, Bradshaw DJ, Axe AS, Williams C, Nile CJ, Ramage G (2016) Viable compositional analysis of an eleven species oral polymicrobial biofilm. Front Microbiol 7:912

    Article  Google Scholar 

  26. Yue H, Bing J, Zheng Q, Zhang Y, Hu T, Du H, Wang H, Huang G (2018) Filamentation in Candida auris, an emerging fungal pathogen of humans: passage through the mammalian body induces a heritable phenotypic switch. Emerg Microbes Infect 7(1):188

    Article  CAS  Google Scholar 

  27. Wang X, Bing J, Zheng Q, Zhang F, Liu J, Yue H, Tao L, Du H, Wang Y, Wang H, Huang G (2018) The first isolate of Candida auris in China: clinical and biological aspects. Emerg Microbes Infect 7(1):93

    PubMed  PubMed Central  CAS  Google Scholar 

  28. Short B, Brown J, Delaney C, Sherry L, Williams C, Ramage G, Kean R (2019) Candida auris exhibits resilient biofilm characteristics in vitro: implications for environmental persistence. J Hosp Infect 103(1):92–96

    Article  CAS  Google Scholar 

  29. Cleare LG, Li KL, Abuzeid WM, Nacharaju P, Friedman JM, Nosanchuk JD (2020) NO Candida auris: nitric oxide in nanotherapeutics to combat emerging fungal pathogen Candida auris. J Fungi (Basel) 6(2):85

    Article  CAS  Google Scholar 

  30. Larkin E, Hager C, Chandra J, Mukherjee PK, Retuerto M, Salem I, Long L, Isham N, Kovanda L, Borroto-Esoda K, Wring S, Angulo D, Ghannoum M (2017) The emerging pathogen Candida auris: growth phenotype, virulence factors, activity of antifungals, and effect of SCY-078, a novel glucan synthesis inhibitor, on growth morphology and biofilm formation. Antimicrob Agents Chemother 61(5):e02396–e02316

    Article  CAS  Google Scholar 

  31. Essary BD, Marshall PA (2009) Assessment of FUN-1 vital dye staining: yeast with a block in the vacuolar sorting pathway have impaired ability to form CIVS when stained with FUN-1 fluorescent dye. J Microbiol Methods 78(2):208–212

    Article  CAS  Google Scholar 

  32. Ramage G (2016) Comparing apples and oranges: considerations for quantifying candidal biofilms with XTT [2,3-bis(2-methoxy-4-nitro-5-sulfo-phenyl)-2H-tetrazolium-5-carboxanilide] and the need for standardized testing. J Med Microbiol 65(4):259–260

    Article  CAS  Google Scholar 

  33. Nocker A, Cheung CY, Camper AK (2006) Comparison of propidium monoazide with ethidium monoazide for differentiation of live vs. dead bacteria by selective removal of DNA from dead cells. J Microbiol Methods 67(2):310–320

    Article  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Oral Sciences Research Group, College of Medical, Veterinary and Life Sciences, Glasgow University, Glasgow, UK

    Jason L. Brown, Bryn Short, Leighann Sherry & Gordon Ramage

  2. Glasgow Biofilm Research Network, Glasgow, UK

    Jason L. Brown, Bryn Short, Alicia Ware, Leighann Sherry, Ryan Kean & Gordon Ramage

  3. Department of Biological and Biomedical, School of Health and Life Sciences, Glasgow Caledonian University, Glasgow, UK

    Alicia Ware & Ryan Kean

Authors
  1. Jason L. Brown
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Bryn Short
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Alicia Ware
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Leighann Sherry
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Ryan Kean
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Gordon Ramage
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding authors

Correspondence to Jason L. Brown or Gordon Ramage .

Editor information

Editors and Affiliations

  1. Institute of Medical Sciences (IMS), University of Aberdeen, Aberdeen, UK

    Alexander Lorenz

Rights and permissions

Reprints and permissions

Copyright information

© 2022 The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature

About this protocol

Check for updates. Verify currency and authenticity via CrossMark

Cite this protocol

Brown, J.L., Short, B., Ware, A., Sherry, L., Kean, R., Ramage, G. (2022). Cell Viability Assays for Candida auris. 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_10

Download citation

  • DOI: https://doi.org/10.1007/978-1-0716-2417-3_10

  • Published:

  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-2416-6

  • Online ISBN: 978-1-0716-2417-3

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation