Phenotypic Switching in Candida glabrata Involves Phase-Specific Regulation of the Metallothionein Gene MT-II and the Newly Discovered Hemolysin Gene HLP

Abstract

Although Candida glabrata has emerged in recent years as a major fungal pathogen, there have been no reports demonstrating that it undergoes either the bud-hypha transition or high-frequency phenotypic switching, two developmental programs believed to contribute to the pathogenic success of other Candida species. Here it is demonstrated that C. glabrata undergoes reversible, high-frequency phenotypic switching between a white (Wh), light brown (LB), and dark brown (DB) colony phenotype discriminated on an indicator agar containing 1 mM Candida glabrata has emerged as one of the three most common Candida species colonizing humans (8, 12). C. glabrata now represents the second-most-common Candida species causing bloodstream infections (36) and, at least in the Detroit area, one of the prevalent species responsible for yeast vaginitis (42,52). A dramatic increase in the carriage of C. glabrata has also been demonstrated in dentate individuals over 80 years of age, and the proportion of elderly individuals with dentures carrying C. glabrata in one study was found to be greater than 50% (23). What is most worrisome about the recent emergence of C. glabrata as a major Candida pathogen and commensal is that it is naturally resistant to azole drug therapy (3,9,14,27).The success of the most prevalent Candida pathogen, C. albicans, depends in part on its phenotypic plasticity. C. albicans exhibits two developmental programs that provide a portion of its phenotypic plasticity, the bud-hypha transition (11, 44) and high-frequency phenotypic switching (45-47). Transition to a hyphal growth form provides C. albicans with the capacity to penetrate tissue and disseminate (35), and mutants of C. albicans that do not form hyphae exhibit a reduction in virulence in animal models (20,37,43). High-frequency phenotypic switching involves the combinatorial regulation of phase-specific genes (45-47), several of which appear to facilitate pathogenesis, including secreted aspartyl proteinases (15,32,33,55) and drug resistance genes (1). Misexpression of phase-specific genes in the wrong phase alters the specificity of virulence in different animal models (18,19). Surprisingly, C. glabrata has never been reported to undergo either the budhypha transition or high-frequency phenotypic switching. How, then, has C. glabrata achieved its recent success both as a commensal and as a pathogen? One possible answer is that the developmental plasticity afforded by the bud-hypha transition and high-frequency phenotypic switching is really not important in the overall pathogenesis of an infectious yeast. An alternative answer is that although these two developmental programs are important to C. albicans (45) and other highly related species (50; S. O. Soll, S. R. Lockhart, and D. R. Soll, unpublished observations), they may not be important for the pathogenesis of C. glabrata. C. glabrata may have developed alternative mechanisms that generate the plasticity that these developmental programs provide for rapid responses to environmental challenges. He...

Dark brown is the more virulent of the switch phenotypes of Candida glabrata

“…The fluorescent GFP-expressing strain Ca2-Gf2 and the non-fluorescent CFP-expressing strain Ca2-CF1 were selected for experimental use. Both strains were tested and found to undergo core switching normally (Lachke et al, 2000(Lachke et al, , 2002.…”

“…For monitoring phenotypic switching, cells were plated at a density of 40 c.f.u. per plate on YPD agar containing 1 mM CuSO 4 (Lachke et al, 2000).…”

“…In Candida glabrata, there are two high-frequency switching systems, core and irregular wrinkle (Lachke et al, 2000(Lachke et al, , 2002. In the core system, cells switch among four phenotypes distinguishable by differences in the brown coloration of colonies grown on agar containing 1 mM CuSO 4 .…”

Fluconazole resistance in Candida glabrata is associated with increased bud formation and metallothionein production

“…Each experimental condition was performed in duplicate (Pabinger et al, 2009) and repeated at least once on different days for reproducibility. A meltingcurve assay was carried out for each experiment to confirm the specificity of the primers (Lachke et al, 2000). …”

“…Confirming the latter, the mRNA levels of MTI and MTIIa of the two FL-resistant mutants grown on medium containing copper were far greater than that of the control parental strain (P,0.05) without copper supplements. It has also been demonstrated that genes other than those involved in copper assimilation, such as HLP1, may be induced in response to rapid environmental challenge (Lachke et al, 2000). Indeed, in C. albicans, high-frequency phenotypic switching involves the combinatorial regulation of phase-specific genes (Soll, 1997), several of which appear to facilitate pathogenesis, including secreted proteinases and drug resistance genes.…”