International Journal of Antimicrobial Agents 27 (2006) 403–408 Resistance mechanisms in fluconazole-resistant Candida albicans isolates from vaginal candidiasis Jana Cernicka, Julius Subik ∗ Comenius University in Bratislava, Faculty of Natural Sciences, Department of Microbiology and Virology, Mlynska dolina B-2, 842 15 Bratislava 4, Slovak Republic Received 25 October 2005; accepted 8 December 2005 Abstract Candida albicans is the most frequently identified yeast species causing mycotic vaginitis. A significant number of vaginal yeast isolates are resistant to azole antifungal agents in vitro. Here we investigated the molecular mechanisms of resistance in 22 randomly selected fluconazole- resistant vaginal C. albicans isolates. Twelve isolates in this collection were found to be cross-resistant to itraconazole and 15 to voriconazole. Most of them also displayed decreased susceptibility to terbinafine. Northern blot analyses revealed overexpression of the MDR1 gene in all isolates, which in some isolates was accompanied by elevated levels of CDR1/CDR2 and ERG11 expression. Sequence analysis of the polymerase chain reaction-amplified ERG11 gene of selected azole-resistant isolates identified D116E and V488I amino acid alterations in Erg11p that are known to be conserved in fluconazole-resistant strains. The results demonstrate that decreased susceptibilities of vaginal yeast isolates to clinically used azole derivatives are the result of a combination of several molecular mechanisms involving drug efflux and alterations in the structure or cellular amount of 14--lanosterol demethylase. © 2006 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved. Keywords: Antifungal agents; Candida albicans; Drug resistance; Fluconazole; Gene expression; Vaginal isolates 1. Introduction Fungal infections caused by opportunistic pathogens have become more frequent, partially as a result of prolonged antibiotic therapy and the increasing number of immuno- compromised patients [1–3]. Mycotic vaginitis is a com- mon mucosal infection caused mainly by the yeast Candida albicans. It occurs with the highest prevalence in women aged 20–30 years [4–6]. Therapy of vulvovaginal candidi- asis frequently fails due to resistance of yeast pathogens to the drugs used. Resistance of Candida species to azole antifungals is the most prevalent type of resistance to antimy- cotics [7,8]. In recent reports, 3.6% of C. albicans vaginal isolates were found to be resistant to fluconazole [9,10], whilst resistance to itraconazole was considerably higher (16.2%) [10]. On the other hand, a significantly higher fre- quency of fluconazole resistance has been reported in an ∗ Corresponding author. Tel.: +421 2 6029 6631; fax: +421 2 6542 9064. E-mail address: subik@fns.uniba.sk (J. Subik). Argentinian study [11] and by us [12] (13.4% and 12.8%, respectively). Several mechanisms by which yeasts become resistant to azole antifungals have been elucidated [13,14]. Resistance can result from: (i) overexpression of the ERG11 gene encod- ing an enzyme involved in demethylation of 14--lanosterol; (ii) a decreased affinity of mutant Erg11p for azoles; (iii) fail- ure of cells to accumulate antifungals owing to overexpres- sion of the MDR1, CDR1 and CDR2 genes encoding major drug efflux pumps using the protonmotive force and ATP to transport drugs across the plasma membrane; and (iv) an altered ergosterol biosynthesis pathway stemming from inac- tivation of the sterol C5,6-desaturase encoded by the ERG3 gene. Analysis of drug resistance mechanisms in matched sets of susceptible and resistant isolates of the same strain from human immunodeficiency virus (HIV)-infected patients confirmed the multifactorial nature of azole resistance, with a predominance of overexpression of genes encoding drug efflux pumps [15]. A similar study using vaginal yeast iso- lates has not yet been reported. 0924-8579/$ – see front matter © 2006 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved. doi:10.1016/j.ijantimicag.2005.12.005