Phenotypic switching and filamentation in Candida glabrata

Abstract

Candida glabrata switches spontaneously, reversibly and at high frequency among the following four phenotypes distinguishable by graded colony colouration on CuSO 4 -containing agar : white (Wh), light brown (LB), dark brown (DB) and very dark brown (vDB). These phenotypes also differ in a graded fashion in the level of expression of the metallothionein gene MTII (Wh T LB T DB a vDB), the frequency of switching (Wh S LB S DB S vDB) and colouration on phloxine B-containing agar (Wh S LB S DB S vDB). Switching among the four graded phenotypes is referred to as ' the core switching system '. An additional switch phenotype, ' irregular wrinkle ' (IWr), has been identified, which exhibits a highly wrinkled colony morphology. The characteristics of IWr suggest that switching to and from this phenotype represents a second high-frequency switching system. A microscopic analysis revealed that during the first 3 days of colony development, cells in the centres of Wh, LB, DB and vDB colonies expressed almost exclusively the budding yeast phenotype. After 3 days, however, pseudohyphae and cells extending tubes accumulated, so that by 7 days the proportions of these two cellular phenotypes reached 40-50 % and 10-20 %, respectively. In contrast, IWr colonies were composed almost exclusively of pseudohyphae through the first 6 days of colony development. After 6 days, IWr colonies began to accumulate both budding yeast cells and tubes. The tubes formed by C. glabrata reached lengths of up to six cell diameters, but the tubes did not represent traditional compartmentalized hyphae. Tube growth ended when the tube tip expanded to form a bud. Tubes then functioned as corridors for daughter nucleus migration to the apical bud, and were ultimately left uncompartmentalized and nucleus free. Core switching, pseudohypha formation and tube formation occurred in a majority of 62 tested clinical isolates, demonstrating that these developmental programmes are general characteristics of most strains of C. glabrata.

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

“…Metallothionein production by switch phenotypes of C. glabrata was detected by the method described by Lachke et al (2002). Cells from a primary colony (cultured on RPMI agar containing 120 mg FL ml 21 ) and visually homogeneous for a single switch phenotype were inoculated into liquid yeast-peptone-glucose medium containing 1 mM CuSO 4 (YPG-CuSO 4 ) and grown at 25 uC until they reached a concentration of~5610 6 c.f.u.…”

“…glabrata is known to switch to pseudohyphal growth when starved of nitrogen (Csank & Haynes, 2000), but the fungus does not form hyphae during host invasion (Lachke et al, 2002) and only budding growth has been observed in animal tissues (Fidel et al, 1999). In this study, highly FLresistant C. glabrata mutants formed a higher number of budding yeasts and also larger aggregates than the parental control when cultured in FCS, indicating that this attribute may be associated with high FL resistance.…”

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

“…We previously demonstrated in limited plating experiments that 25 of 62 clinical isolates of C. glabrata (40 %) underwent core switching (Lachke et al, 2002). Strains that had not been observed to switch in the initial experiments did so in larger-scale plating experiments.…”

“…The core phenotypes are white (Wh), light brown (LB), dark brown (DB) and very dark brown (vDB). Although core switching was first observed and defined on agar containing CuSO 4 , it has been shown to occur in the absence of CuSO 4 (Lachke et al, 2002) and can also be distinguished by graded pink coloration when grown on agar containing phloxine B (Lachke et al, 2002). Cells expressing any of the core phenotypes can also switch to the irregular wrinkle (IWr) phenotype, then back to the original core phenotype (Lachke et al, 2002).…”

“…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 .…”