Skip to main content

Advertisement

SpringerLink
Log in

Candida species causing fungal keratitis: molecular identification, antifungal susceptibility, biofilm formation, and clinical aspects

  • Bacterial and Fungal Pathogenesis - Research Paper
  • Published:
Brazilian Journal of Microbiology Aims and scope Submit manuscript

Abstract

The study aimed to evaluate the clinical aspects, molecular identification, biofilm formation, and antifungal susceptibility profile of Candida species isolated from fungal keratitis. Thirteen Candida isolates from 13 patients diagnosed with Candida keratitis were retrieved and grown in pure culture. Species identification was performed by micromorphology analysis and ITS-rDNA sequencing. The broth microdilution method tested the minimum inhibitory concentration (MIC) of four antifungal drugs (fluconazole, amphotericin B, voriconazole, and anidulafungin). The biofilms were cultured and incubated with antifungal drugs for 24 h. The XTT reduction assay measured the biofilm activity. Biofilm MICs were calculated based on a 50% reduction in metabolic activity compared with the activity of the drug-free control. Among isolates, two were C. albicans, 10 were C. parapsilosis (sensu stricto), and one was C. orthopsilosis. All isolates were classified as susceptible or intermediate to all four antifungal drugs. Four isolates were very low biofilm producers (30%). Nine isolates were biofilm producers, and all biofilm samples were unsusceptible to all drugs tested. Previous ocular surgery was the most common underlying condition for fungal keratitis (84.6%), and C. parapsilosis was the most frequent Candida species (76.9%). Four patients (30.7%) needed keratoplasty, whereas two (15.3%) required evisceration. The biofilm formation ability of Candida isolates decreased antifungal susceptibility compared with planktonic cells. Despite in vitro antifungal susceptibility, almost half of the patients were unresponsive to clinical treatment and needed surgery.

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

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1

Similar content being viewed by others

Data availability

The data corresponding to this article are available in the article as well as in its online supplementary material.

References

  1. Bezerra FM, Höfling-Lima AL, de Oliveira LA (2020) Fungal keratitis management in a referral cornea center in Brazil. Rev Bras Oftalmol 79:315–319. https://doi.org/10.5935/0034-7280.20200067

    Article  Google Scholar 

  2. Höfling-Lima AL, Forseto A, Duprat JP, Andrade A, Souza LB, Godoy P et al (2005) Laboratory study of the mycotic infectious eye diseases and factors associated with keratitis. Arq Bras Oftalmol 68:21–27. https://doi.org/10.1590/s0004-27492005000100005

    Article  PubMed  Google Scholar 

  3. Keay LJ, Gower EW, Iovieno A, Oechsler RA, Alfonso EC, Matoba A et al (2011) Clinical and microbiological characteristics of fungal keratitis in the United States, 2001–2007: a multicenter study. Ophthalmology 118:920–926. https://doi.org/10.1016/j.ophtha.2010.09.011

    Article  PubMed  Google Scholar 

  4. Agrawal V, Biswas J, Madhavan HN, Mangat G, Reddy MK, Saini JS et al (1994) Current perspectives in infectious keratitis. Indian J Ophthalmol 42:171–192

    CAS  PubMed  Google Scholar 

  5. Tanure MAG, Cohen EJ, Sudesh S, Rapuano CJ, Laibson PR (2000) Spectrum of fungal keratitis at Wills Eye Hospital, Philadelphia Pennsylvania. Cornea 19:307–312. https://doi.org/10.1097/00003226-200005000-00010

    Article  CAS  PubMed  Google Scholar 

  6. Jastaneiah SS, Al-Rajhi AA, Abbott D (2011) Ocular mycosis at a referral center in Saudi Arabia: a 20-year study. Saudi J Ophthalmol 25:231–238. https://doi.org/10.1016/j.sjopt.2011.04.004

    Article  PubMed  PubMed Central  Google Scholar 

  7. Rizvi Y, Agarwal PM, Mishra PP, Dokania A (2016) Microbiological pattern and epidemiologic trends of fungal keratitis in North India. Jebmh 3:2445–2450. https://doi.org/10.18410/jebmh/2016/537

    Article  Google Scholar 

  8. Ansari Z, Miller D, Galor A (2013) Current thoughts in fungal keratitis: diagnosis and treatment. Curr Fungal Infect Rep 7:209–218. https://doi.org/10.1007/s12281-013-0150-110.1007/s12281-013-0150-1

    Article  PubMed  PubMed Central  Google Scholar 

  9. Qiao GL, Ling J, Wong T, Yeung SN, Iovieno A (2020) Candida keratitis: epidemiology, management, and clinical outcomes. Cornea. 39:801–805. https://doi.org/10.1097/ICO.0000000000002306

    Article  PubMed  Google Scholar 

  10. Merseguel KB, Nishikaku AS, Rodrigues AM, Padovan AC, e Ferreira RC, de Azevedo Melo AS et al (2015) Genetic diversity of medically important and emerging Candida species causing invasive infection. BMC Infect Dis 15:57. https://doi.org/10.1186/s12879-015-0793-3

    Article  PubMed  PubMed Central  Google Scholar 

  11. Desai JV, Mitchell AP, Andes DR (2014) Fungal biofilms, drug resistance, and recurrent infection. Cold Spring Harb Perspect Med. 4:a019729. https://doi.org/10.1101/cshperspect.a019729

    Article  PubMed  PubMed Central  Google Scholar 

  12. Melo AS, Bizerra FC, Freymüller E, Arthington-Skaggs BA, Colombo AL (2011) Biofilm production and evaluation of antifungal susceptibility amongst clinical Candida spp. isolates, including strains of the Candida parapsilosis complex. Med Mycol 49:253–262. https://doi.org/10.3109/13693786.2010.530032

    Article  CAS  PubMed  Google Scholar 

  13. CLSI. Performance standards for antifungal susceptibility testing of yeasts. 1st ed. CLSI supplement M60. Wayne PA: CLSI; 2017.

  14. Jin Y, Yip HK, Samaranayake YH, Yau JY, Samaranayake LP (2003) Biofilm-forming ability of Candida albicans is unlikely to contribute to high levels of oral yeast carriage in cases of human immunodeficiency virus infection. J Clin Microbiol 41:2961–2967. https://doi.org/10.1128/JCM.41.7.2961-2967.2003

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Iturrieta-González IA, Padovan ACB, Bizerra FC, Hahn RC, Colombo AL (2014) Multiple species of Trichosporon produce biofilms highly resistant to triazoles and amphotericin B. PLOS ONE 9:e109553. https://doi.org/10.1371/journal.pone.0109553

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. CLSI. Reference method for broth dilution antifungal susceptibility testing of yeasts. 4th ed. CLSI M27. Wayne, PA: CLSI; 2017.

  17. Cariello AJ, Passos RM, Yu MC, Hofling-Lima AL (2011) Microbial keratitis at a referral center in Brazil. Int Ophthalmol 31:197–204. https://doi.org/10.1007/s10792-011-9441-0

    Article  PubMed  Google Scholar 

  18. Weems JJ Jr (1992) Candida parapsilosis: epidemiology, pathogenicity, clinical manifestations, and antimicrobial susceptibility. Clin Infect Dis 14:756–766. https://doi.org/10.1093/clinids/14.3.756

    Article  PubMed  Google Scholar 

  19. Strausbaugh LJ, Sewell DL, Ward TT, Pfaller MA, Heitzman T, Tjoelker R (1994) High frequency of yeast carriage on hands of hospital personnel. J Clin Microbiol 32:2299–2300. https://doi.org/10.1128/jcm.32.9.2299-2300.1994

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Sun RL, Jones DB, Wilhelmus KR (2007) Clinical characteristics and outcome of Candida keratitis. Am J Ophthalmol 143:1043–1045. https://doi.org/10.1016/j.ajo.2007.02.016

    Article  PubMed  PubMed Central  Google Scholar 

  21. Bourcier T, Touzeau O, Thomas F, Chaumeil C, Baudrimont M, Borderie V et al (2003) Candida parapsilosis keratitis. Cornea 22:51–55. https://doi.org/10.1097/00003226-200301000-00012

    Article  PubMed  Google Scholar 

  22. Pereira R, dos Santos Fontenelle RO, de Brito EHS, de Morais SM (2021) Biofilm of Candida albicans: formation, regulation and resistance. J Appl Microbiol 131:11–22. https://doi.org/10.1111/jam.14949

    Article  CAS  PubMed  Google Scholar 

  23. Tits J, Cammue BPA, Thevissen K (2020) Combination therapy to treat fungal biofilm-based infections. Int J Mol Sci 21:8873. https://doi.org/10.3390/ijms21228873

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Kuhn DM, George T, Chandra J, Mukherjee PK, Ghannoum MA (2002) Antifungal susceptibility of Candida biofilms: unique efficacy of amphotericin B lipid formulations and echinocandins. Antimicrob Agents Chemother 46:1773–1780. https://doi.org/10.1128/AAC.46.6.1773-1780.2002

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Chandra J, Mukherjee PK, Leidich SD, Faddoul FF, Hoyer LL, Douglas LJ et al (2001) Antifungal resistance of Candidal biofilms formed on denture acrylic in vitro. J Dent Res 80:903–908. https://doi.org/10.1177/00220345010800031101

    Article  CAS  PubMed  Google Scholar 

  26. Muzny CA, Schwebke JR (2015) Biofilms: an underappreciated mechanism of treatment failure and recurrence in vaginal infections. Clin Infect Dis 61:601–606. https://doi.org/10.1093/cid/civ353

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Kumamoto CA (2002) Candida biofilms. Curr Opin Microbiol 5:608–611. https://doi.org/10.1016/s1369-5274(02)00371-5

    Article  CAS  PubMed  Google Scholar 

  28. Ramage G, Saville SP, Thomas DP, López-Ribot JL (2005) Candida biofilms: an update. Eukaryot Cell 4:633–638. https://doi.org/10.1128/EC.4.4.633-638.2005

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Chamilos G, Lewis RE, Albert N, Kontoyiannis DP (2007) Paradoxical effect of echinocandins across Candida species in vitro: evidence for echinocandin-specific and Candida species-related differences. Antimicrob Agents Chemother 51:2257–2259. https://doi.org/10.1128/AAC.00095-07

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Bizerra FC, Melo ASA, Katchburian E, Freymüller E, Straus AH, Takahashi HK et al (2011) Changes in cell wall synthesis and ultrastructure during paradoxical growth effect of caspofungin on four different Candida species. Antimicrob Agents Chemother 55:302–310. https://doi.org/10.1128/AAC.00633-10

    Article  CAS  PubMed  Google Scholar 

  31. Pate JC, Jones DB, Wilhelmus KR (2006) Prevalence and spectrum of bacterial co-infection during fungal keratitis. Br J Ophthalmol 90:289–292. https://doi.org/10.1136/bjo.2005.081869

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

We acknowledge Larissa Molina Favarello, MSc., for collaborating in performing the susceptibility tests.

Author information

Authors and Affiliations

  1. Department of Ophthalmology and Visual Sciences, Federal University of São Paulo, São Paulo, Brazil

    Fernanda M. Bezerra, Talita T. Rocchetti, Maria Cecília Z. Yu, Ana Luisa Höfling-Lima & Lauro A. de Oliveira

  2. Special Laboratory of Mycology, Federal University of São Paulo, São Paulo, Federal University of São Paulo, São Paulo, Brazil

    Soraia L. Lima, Daniel A. da Matta & Analy S. A. Melo

Authors
  1. Fernanda M. Bezerra
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Talita T. Rocchetti
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Soraia L. Lima
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Maria Cecília Z. Yu
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Daniel A. da Matta
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Ana Luisa Höfling-Lima
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Analy S. A. Melo
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Lauro A. de Oliveira
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

The study conception and design was performed by Analy Salles de Azevedo Melo, Daniel Archimedes da Matta, and Lauro Augusto de Oliveira. All authors contributed to material preparation, data collection, and analysis. The first draft of the manuscript was written by Lauro Augusto de Oliveira, Fernanda Machado Bezerra Linhares, and Talita Trevizani Rocchetti, and all authors commented on previous versions of the manuscript. All authors have read and approved the final manuscript.

Corresponding author

Correspondence to Lauro A. de Oliveira.

Ethics declarations

Ethics approval

This study was performed in accordance with the principles of the Declaration of Helsinki. Approval was granted by the Ethics Committee of the Federal University of São Paulo.

Consent to participate

Informed consent was obtained from all individual participants included in this study.

Consent for publication

This manuscript does not contain any photos or individual person’s data.

Conflict of interest

The authors declare no competing interests.

Additional information

Responsible Editor: Carlos Pelleschi Taborda

Publisher’s note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Lay summary

Molecular identification, biofilm formation, and antifungal susceptibility of Candida species isolated from fungal keratitis were evaluated. Biofilm formation, which was frequently seen in Candida sp., decreased antifungal susceptibility and could thus justify the unresponsiveness to treatment.

Rights and permissions

Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Bezerra, F.M., Rocchetti, T.T., Lima, S.L. et al. Candida species causing fungal keratitis: molecular identification, antifungal susceptibility, biofilm formation, and clinical aspects. Braz J Microbiol 54, 629–636 (2023). https://doi.org/10.1007/s42770-023-00964-w

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s42770-023-00964-w

Keywords

Search

Navigation