Journal of Antimicrobial Chemotherapy (2002) 49, 973-980
© 2002 The British Society for Antimicrobial Chemotherapy
Investigation of multidrug efflux pumps in relation to fluconazole resistance in Candida albicans biofilms
Departments of 1Microbiology and 2Medicine, Division of Infectious Diseases, The University of Texas Health Science Center at San Antonio, San Antonio, TX, USA
Received 6 July 2001; returned 23 November 2001; revised 2 January 2002; accepted 4 March 2002.
A main characteristic associated with microbial biofilms is their increased resistance to antimicrobial chemotherapies. However, at present very little is known about the phenotypic changes that occur during the transition from the planktonic to the biofilm mode of growth. Candida albicans biofilms displayed an organized three-dimensional structure, and consisted of a dense network of yeasts and filamentous cells deeply embedded in exopolymeric matrix. These biofilms were intrinsically resistant to fluconazole. Moreover, the resistance phenotype was maintained by sessile cells when resuspended as free-floating cells, thus demonstrating that biofilm integrity and the presence of exopolymeric material are not the sole determinants of biofilm resistance. Under planktonic conditions, one of the main mechanisms of azole resistance in C. albicans is through active efflux of these drugs mediated by ATP-binding cassette (ABC) transporters and major facilitators. In this study we used northern hybridization to monitor expression of genes belonging to two different types of efflux pump, the ABC transporters and major facilitators (encoded by CDR and MDR genes, respectively), in C. albicans populations under both planktonic and biofilm growth. It was demonstrated that expression of genes encoding both types of efflux pump were up-regulated during the course of biofilm formation and development. Moreover, antifungal susceptibilities of biofilms formed by a set of C. albicans mutant strains deficient in efflux pumps were investigated to determine their contribution to biofilm resistance. Remarkably, mutants carrying single and double deletion mutations in 
cdr1, cdr2, mdr1, cdr1/cdr2 and mdr1/cdr1 were hypersusceptible to fluconazole when planktonic, but still retained the resistant phenotype during biofilm growth. These analyses demonstrate that C. albicans biofilm resistance is a complex phenomenon that cannot be explained by one mechanism alone, instead it is multifactorial and may involve different molecular mechanisms of resistance compared with those displayed by planktonic cells.
Keywords: Candida albicans, biofilm, efflux pumps, resistance
* Correspondence address. Department of Medicine, Division of Infectious Diseases, The University of Texas Health Science Center at San Antonio, South Texas Centers for Biology in Medicine, Texas Research Park, 15355 Lambda Drive, San Antonio, TX 78245, USA. Tel: +1-210-562-5017; Fax: +1-210-562-5016; E-mail: RIBOT{at}UTHSCSA.EDU
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