Volume 3, Issue 1 (March 2017)                   Curr Med Mycol 2017, 3(1): 13-19 | Back to browse issues page


XML Print


Abstract:   (1068 Views)

Background and Purpose: The frequency of opportunistic fungal infections in immunocompromised patients, especially by Candida species, has sharply increased in the last few decades. The objective of this study was to analyse the ergosterol content and gene expression profiling of clinical isolates of fluconazole resistant Candida albicans.
Materials and Methods: Sixty clinical samples were identified and collected from immunocompromised patients, namely recurrent oral, vaginal, and cutaneous candidiasis, during 2015-16. Antifungal susceptibility testing of fluconazole against clinical Candida species was performed according to Clinical and Laboratory Standards Institute guidelines. Ergosterol content and gene expression profiling of sterol 14α-demethylase (ERG11) gene in fluconazole-susceptible and –resistant C.albicans were investigated.
Results: The specimens consisted of C. albicans (46.67%), Candida krusei (41.67%) (and Candida tropicalis (11.67%). All the isolates were resistant to fluconazole. No significant reduction was noted in total cellular ergosterol content in comparison with untreated controls in terms of fluconazole-resistant C. albicans. The expression level of ERG11 gene was down-regulated in fluconazole-susceptible C. albicans.Eventually, the expression pattern of ERG11 gene revealed no significant changes in fluconazole-resistant isolates compared to untreated controls. The results revealed no significant differences between fluconazole-susceptible and –resistant C. albicans sequences by comparison with ERG11 reference sequence.
Conclusion: Our findings provide an insight into the mechanism of fluconazole resistance in C. albicans. The mechanisms proposed for clinical isolates of fluconazole-resistant C. albicans are alteration in sterol biosynthesis, analysis of expression level of ERG11 gene, and analysis of gene sequences. Nonetheless, further studies are imperative to find molecular mechanisms that could be targeted to control fluconazole resistance.
 

Full-Text [PDF 854 kb]   (335 Downloads)    
Type of Study: Original Articles | Subject: Antifungal Susceptibility,
Received: 2017/06/21 | Accepted: 2017/08/20 | Published: 2017/12/12

References
1. Yang YL, Cheng HH, Ho YA, Hsiao CF, Lo HJ. Fluconazole resistance rate of Candida species from different regions and hospital types in Taiwan. J Microbiol Immunol Infect. 2003; 36(3): 187-91.
2. Marichal P, Koymans L, Willemsens S, Bellens D, Verhasselt P, Luyten W, Borgers M, Ramaekers FC, Odds FC, Bossche HV. Contribution of mutations in the cytochrome P450 14α-demethylase (Erg11p, Cyp51p) to azole resistance in Candida albicans. Microbiology. 1999; 145 (Pt 10): 2701-13. [DOI:10.1099/00221287-145-10-2701]
3. Sanglard D, Odds FC. Resistance of Candida species to antifungal agents: molecular mechanisms and clinical consequences. Lancet Infect Dis 2002; 2:73–85. [DOI:10.1016/S1473-3099(02)00181-0]
4. Loeffler J, Stevens DA. Antifungal drug resistance. Clin Infect Dis. 2003; 36: 31-41. [DOI:10.1086/344658]
5. Casalinuovo IA, Di Francesco P, Garaci E. Fluconazole resistance in Candida albicans: a review of mechanisms. Eur Rev Med Pharmacol Sci. 2004; 8(2): 69-77.
6. Sanglard D. Emerging threats in antifungal-resistant fungal pathogens. Front Med (Lausanne). 2016; 3: 11. [DOI:10.3389/fmed.2016.00011]
7. Franz R, Kelly SL, Lamb DC, Kelly DE, Ruhnke M, Morschhäuser J. Multiple molecular mechanisms contribute to a stepwise development of fluconazole resistance in clinical Candida albicansstrains. Antimicrob Agents Chemother. 1998; 42(12): 3065-72.
8. Xu Y, Sheng F, Zhao J, Chen L, Li C. ERG11 mutations and expression of resistance genes in fluconazole-resistant Candida albicans isolates. Arch Microbiol. 2015; 197(9): 1087-93. [DOI:10.1007/s00203-015-1146-8]
9. Alvarez-Rueda N, Fleury A, Logé C, Pagniez F, Robert E, Morio F, Le Pape P. The amino acid substitution N136Y in Candida albicans sterol 14alpha-demethylase is involved in fluconazole resistance. Med Mycol 2016; 54(7):764-75. [DOI:10.1093/mmy/myw023]
10. Song JL, Jo Beth Harry,‡ Richard T. Eastman, Brian G. Oliver, and Theodore C. White The Candida albicans lanosterol 14-α-demethylase (ERG11) gene promoter is maximally induced after prolonged growth with antifungal drugs. Antimicrob Agents Chemother. 2004; 48(4): 1136–44. [DOI:10.1128/AAC.48.4.1136-1144.2004]
11. Pfaller MA, Diekema DJ, Sheehan DJ. Interpretive breakpoints for fluconazole and Candida revisited: a blueprint for the future of antifungal susceptibility testing. Clin Microbiol Rev. 2006; 19(2): 435–447. [DOI:10.1128/CMR.19.2.435-447.2006]
12. Lamb DC, Kelly DE, Kelly SL. Molecular diversity of sterol 14α-demethylase substrates in plants, fungi and humans. FEBS Lett. 1998; 425(2): 263-5. [DOI:10.1016/S0014-5793(98)00247-6]
13. Dunkel N, Liu TT, Barker KS, Homayouni R, Morschhäuser J, Rogers PD. A gain-of-function mutation in the transcription factor Upc2p causes upregulation of ergosterol biosynthesis genes and increased fluconazole resistance in a clinical Candida albicans isolate. Eukaryot Cell. 2008; 7(7): 1180-90. [DOI:10.1128/EC.00103-08]
14. Evans EGV, Richardson MD. Medical Mycology: A Practical Approach (The Practical Approach Series). Oxford University Press: Newyork; 1989.
15. Harmal NS, Khodavandi A, Alshawsh MA, Farida J, Sekawi Z, Ng KP, Chong PP. Simplex and triplex polymerase chain reaction (PCR) for identification of three medically important Candida species. Afr J Biotechnol. 2012; 11(65): 12895–902.
16. Khodavandi A, Harmal NS, Alizadeh F, Scully OJ, Sidik SHM, Othman F, Sekawi Z, Ng KP, Chong PP. Comparison between allicin and fluconazole in Candida albicans biofilm inhibition and in suppression of HWP1 gene expression. Phytomedicine. 2011; 19(1): 56–63. [DOI:10.1016/j.phymed.2011.08.060]
17. Khodavandi A, Alizadeh Aghai Vanda N, Karimi G, Chong PP. Possible mechanisms of the antifungal activity of fluconazole in combination with terbinafine against Candida albicans. Pharm Biol. 2014; 52(12): 1505–9. [DOI:10.3109/13880209.2014.900808]
18. Pfaller MA, Diekema DJ. Progress in antifungal susceptibility testing of Candida spp. by use of Clinical and Laboratory Standards Institute broth microdilution methods, 2010 to 2012. J Clin Microbiol. 2012;50(9): 2846-56. [DOI:10.1128/JCM.00937-12]
19. Shokohi T, Badali H, Amirrajab N, Ataollahi MR, Kouhpayeh SA, Afsarian MH. In vitro activity of five antifungal agents against Candida albicans isolates, Sari, Iran. Curr Med Mycol. 2016; 2(2): 34-9.
20. Santos JRA, Gouveia LF, Taylor ELS, Resende-Stoianoff MA, Pianetti GA, César IC, Santos DA. Dynamic interaction between fluconazole and amphotericin b against Cryptococcus gattii. Antimicrob Agents Chemother. 2012; 56(5): 2553-8. [DOI:10.1128/AAC.06098-11]
21. Wilheim AB, Miranda-Filho Dde B, Nogueira RA, Rêgo RS, Lima Kde M, Pereira LM. The resistance to fluconazole in patients with esophageal candidiasis. Arq Gastroenterol. 2009; 46(1): 32-7. [DOI:10.1590/S0004-28032009000100011]
22. Ghannoum M, Roilides E, Katragkou A, Petraitis V, Walsh TJ. The role of echinocandins in Candida biofilm-related vascular catheter infections: in vitro and in vivo model systems. Clin Infect Dis. 2015; 61(S6): S618-21. [DOI:10.1093/cid/civ815]
23. Peron IH, Reichert-Lima F, Busso-Lopes AF, Nagasako CK, Lyra L, Moretti ML, Schreiber AZ. Resistance surveillance in Candida albicans: a five-year antifungal susceptibility evaluation in a Brazilian university hospital. PLoS One. 2016; 11(7): e0158126. [DOI:10.1371/journal.pone.0158126]
24. Morschhäuser J. The genetic basis of fluconazole resistance development in Candida albicans. Biochim Biophys Acta. 2002; 1587(2-3): 240-8. [DOI:10.1016/S0925-4439(02)00087-X]
25. Pappas PG, Kauffman CA, Andes DR, Clancy CJ, Marr KA, Ostrosky-Zeichner L, Reboli AC, Schuster MG, Vazquez JA, Walsh TJ, Zaoutis TE, Sobel JD. Clinical practice guideline for the management of candidiasis: 2016 update by the infectious diseases society of America. Clin Infect Dis. 2016; 62(4): e1-50. [DOI:10.1093/cid/civ1194]
26. Siikala E, Rautemaa R, Richardson M, Saxen H, Bowyer P, Sanglard D. Persistent Candida albicans colonization and molecular mechanisms of azole resistance in autoimmune polyendocrinopathy–candidiasis–ectodermal dystrophy (APECED) patients. J Antimicrob Chemother. 2010; 65(12): 2505-13. [DOI:10.1093/jac/dkq354]
27. Ahmad A, Khan A, Manzoor N, Khan LA. Evolution of ergosterol biosynthesis inhibitors as fungicidal against Candida. Microb Pathog. 2010; 48(1): 35-41. [DOI:10.1016/j.micpath.2009.10.001]
28. Spampinato C, Leonardi D. Candida infections, causes, targets, and resistance mechanisms: traditional and alternative antifungal agents. Biomed Res Int. 2013; 2013: 204237. [DOI:10.1155/2013/204237]
29. Arthington-Skaggs BA, Jradi H, Desai T, Morrison CJ. Quantitation of ergosterol content: novel method for determination of fluconazole susceptibility of Candida albicans. J Clin Microbiol. 1999; 37(10): 3332-7.
30. Salari S, Khosravi AR, Mousavi SA, Nikbakht-Brojeni GH. Mechanisms of resistance to fluconazole in Candida albicans clinical isolates from Iranian HIV-infected patients with oropharyngeal candidiasis. J Mycol Med. 2016; 26(1): 35 41. [DOI:10.1016/j.mycmed.2015.10.007]
31. Young LY, Hull CM, Heitman J. Disruption of ergosterol biosynthesis confers resistance to amphotericin B in Candida lusitaniae. Antimicrob Agents Chemother. 2003; 47(9): 2717-24. [DOI:10.1128/AAC.47.9.2717-2724.2003]
32. Borecká-Melkusová S, Moran GP, Sullivan DJ, Kucharíková S, Chorvát D Jr, Bujdáková H. The expression of genes involved in the ergosterol biosynthesis pathway in Candida albicans and Candida dubliniensis biofilms exposed to fluconazole. Mycoses. 2009; 52(2): 118-28. [DOI:10.1111/j.1439-0507.2008.01550.x]