Investigation of ERG11 Gene Mutations in Fluconazole Resistant Candida albicans Isolated from a Number of Rasht Hospitals

Authors
A. D. Eftekhari 1
M. Anvari 2
N. Ranji 3
1 Department of Biology, Rasht Branch, Islamic Azad University, Rasht, Iran
2 Department of Microbiology, Faculty of Sciences, Rasht Branch of Islamic Azad University, Rasht, Iran
3 Department of Genetics, Faculty of Sciences, Rasht Branch of Islamic Azad University, Rasht, Iran
Abstract
Objective: Azole antifungal drugs have been a treatment option for Candida albicans infections. However, azole resistance can occur through different mechanisms such as alterations in ERG11 (lanosterol 14α-demethylase). This study assesses ERG11 gene mutations in Candida albicans strains isolated from patients with Candidia volvovaginitis in a number of Rasht hospitals between 2012-2014 by direct PCR and sequencing.
Methods: We identified the yeast strains by standard identification methods, such as germ tubes. Drug sensitivity was determined as MIC 90 values by the macrodilution broth method based on the CLSI protocol. We screened the resistant strains prior to DNA extraction and ERG11 gene mutations were confirmed by PCR sequencing.
Results: From 40 strains, 4 showed high levels of resistance to fluconazole. Of these, two species had a MIC 90 of 512μg/ml and the other two species had a MIC 90 of 1024 μg/ml. Three strains had D116E and V456G polymorphisms.
Conclusion: The most fluconazole resistant Candida albicans strains worldwide were reported. Our results suggested a correlation between the polymorphism and fluconazole resistance in the Candida albicans strains.

Keywords

[1]  Davies JM, Stacey AJ, Gilligan CA. Candida albicans hyphal invasion: thigmotropism or chemotropism? FEMS Microbiol Lett 1999; 171(2): 245-9.
[2]  Kavanaugh NL, Zhang AQ, Nobile CJ, Johnson AD, Ribbeck K. Mucins suppress virulence traits of Candida albicans. MBio 2014; 5(6): e01911.
[3]  Seneviratne CJ, Jin LJ, Samaranayake YH, Samaranayake LP. Cell density and cell aging as factors modulating antifungal resistance of Candida albicans biofilms. Antimicrob Agents Chemother 2008; 52(9): 3259-66.
[4]  Xiang MJ, Liu JY, Ni PH, Wang S, Shi C, Wei B, Ni YX, Ge HL. Erg11 mutations associated with azole resistance in clinical isolates of Candida albicans. FEMS Yeast Res 2013; 13(4): 386-93.
[5]  Perea S, López-Ribot JL, Kirkpatrick WR, McAtee RK, Santillán RA, Martínez M, Calabrese D, Sanglard D, Patterson TF. Prevalence of molecular mechanisms of resistance to azole antifungal agents in Candida albicans strains displaying high-level fluconazole resistance isolated from human immunodeficiency virus-infected patients. Antimicrob Agents Chemother 2001; 45(10): 2676-84.
[6]  Xu Y, Chen L, Li C. Susceptibility of clinical isolates of Candida species to fluconazole and detection of Candida albicans ERG11 mutations. J Antimicrob Chemother 2008; 61(4): 798-804.
[7]  Feng LJ, Wan Z, Wang XH, Li RY, Liu W. Relationship between antifungal resistance of fluconazole resistant Candida albicans and mutations in ERG11 gene. Chin Med J (Engl) 2010; 123(5): 544-8.
[8]  Yan L, Zhang J, Li M, Cao Y, Xu Z, Cao Y, Gao P, Wang Y, Jiang Y. DNA microarray analysis of fluconazole resistance in a laboratory Candida albicans strain. Acta Biochim Biophys Sin (Shanghai) 2008; 40(12): 1048-60.
[9]  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 14alpha-demethylase (Erg11p, Cyp51p) to azole resistance in Candida albicans. Microbiology 1999; 145(Pt 10): 2701-13.
[10]   Wang H, Kong F, Sorrell TC, Wang B, McNicholas P, Pantarat N, Ellis D, Xiao M, Widmer F, Chen SC. Rapid detection of ERG11 gene mutations in clinical Candida albicans isolates with reduced susceptibility to fluconazole by rolling circle amplification and DNA sequencing. BMC Microbiol 2009; 9: 167.
[11]   Gołąbek K, Strzelczyk JK, Owczarek A, Cuber P, Ślemp-Migiel A, Wiczkowski A. Selected mechanisms of molecular resistance of Candida albicans to azole drugs. Acta Biochim Pol 2015; 62(2): 247-51.
[12] Sambrook J, Russell DW. Rapid isolation of yeast DNA. CSH Protoc 2006; 2006(1). pii: pdb.prot4039.
[13]   Zarei Mahmoudabadi A, Rezaei-Matehkolaei A, Navid M, Torabizadeh M, Mazdarani S. Colonization and antifungals susceptibility patterns of Candida species isolated from hospitalized patients in ICUs and NICUs. J Nephropathol 2015; 4(3): 77-84.
[14]   Teymuri M, Mamishi S, Pourakbari B, Mahmoudi S, Ashtiani MT, Sadeghi RH, Yadegari MH. Investigation of ERG11 gene expression among fluconazole-resistant Candida albicans: first report from an Iranian referral paediatric hospital. Br J Biomed Sci 2015; 72(1): 28-31.
[15]   Mohamadi J, Motaghi M, Panahi J, Havasian MR, Delpisheh A, Azizian M, Pakzad I. Anti-fungal resistance in Candida isolated from oral and diaper rash candidiasis in neonates. Bioinformation 2014; 10(11): 667-70.
[16]   Sobel JD, Zervos M, Reed BD, Hooton T, Soper D, Nyirjesy P, Heine MW, Willems J, Panzer H. Fluconazole susceptibility of vaginal isolates obtained from women with complicated Candida vaginitis: clinical implications. Antimicrob Agents Chemother 2003; 47(1): 34-8.
[17]   Shokohi T, Bandalizadeh Z, Hedayati MT, Mayahi S. In vitro antifungal susceptibility of Candida species isolated from oropharyngeal lesions of patients with cancer to some antifungal agents. JJM 2011; 4(Supplement 1): S19-S26.
[18]   Vazquez JA, Peng G, Sobel JD, Steele-Moore L, Schuman P, Holloway W, Neaton JD. Evolution of antifungal susceptibility among Candida species isolates recovered from human immunodeficiency virus-infected women receiving fluconazole prophylaxis. Clin Infect Dis 2001; 33(7): 1069-75.
[19]   Sanglard D, Ischer F, Koymans L, Bille J. Amino acid substitutions in the cytochrome P-450 lanosterol 14alpha-demethylase (CYP51A1) from azole-resistant Candida albicans clinical isolates contribute to resistance to azole antifungal agents. Antimicrob Agents Chemother 1998; 42(2): 241-53.
[20]   Lee MK, Williams LE, Warnock DW, Arthington-Skaggs BA. Drug resistance genes and trailing growth in Candida albicans isolates. J Antimicrob Chemother 2004; 53(2): 217-24.
[21]   Strzelczyk JK, Slemp-Migiel A, Rother M, Gołąbek K, Wiczkowski A. Nucleotide substitutions in the Candida albicans ERG11 gene of azole-susceptible and azole-resistant clinical isolates. Acta Biochim Pol 2013; 60(4): 547-52.
[22]   Goldman GH, da Silva Ferreira ME, dos Reis Marques E, Savoldi M, Perlin D, Park S, Godoy Martinez PC, Goldman MH, Colombo AL. Evaluation of fluconazole resistance mechanisms in candida albicans clinical isolates from HIV-infected patients in Brazil. Diagn Microbiol Infect Dis 2004; 50(1): 25-32.

 
 
Pathobiology Reserach
Volume 18, Issue 3
October 2015
Pages 98-107