Project description:We isolated Candida dubliniensis from a nonhuman source, namely, tick samples from an Irish seabird colony. The species was unambiguously identifi ed by phenotypic and genotypic means. Analysis of the 5.8S rRNA gene showed that the environmental isolates belong to C. dubliniensis genotype 1.
Project description:Candida dubliniensis is a recently described opportunistic fungal pathogen that is closely related to Candida albicans. Candida dubliniensis readily develops resistance to the azole antifungal agent fluconazole, both in vitro and in infected patients, and this resistance is usually associated with upregulation of the CdMDR1 gene, encoding a multidrug efflux pump of the major facilitator superfamily. To determine the role of CdMDR1 in drug resistance in C. dubliniensis, we constructed an mdr1 null mutant from the fluconazole-resistant clinical isolate CM2, which overexpressed the CdMDR1 gene. Sequential deletion of both CdMDR1 alleles was performed by the MPA(R)-flipping method, which is based on the repeated use of a dominant mycophenolic acid resistance marker for selection of integrative transformants and its subsequent deletion from the genome by FLP-mediated, site-specific recombination. In comparison with its parental strain, the mdr1 mutant showed decreased resistance to fluconazole but not to the related drug ketoconazole. In addition, we found that CdMDR1 confers resistance to the structurally unrelated drugs 4-nitroquinoline-N-oxide, cerulenin, and brefeldin A, since the enhanced resistance to these compounds of the parent strain CM2 compared with the matched susceptible isolate CM1 was abolished in the mdr1 mutant. In contrast, CdMDR1 inactivation did not cause increased susceptibility to amorolfine, terbinafine, fluphenazine, and benomyl, although overexpression of CdMDR1 in a hypersusceptible Saccharomyces cerevisiae strain had previously been shown to confer resistance to these compounds. The effect of CdMDR1 inactivation was identical to that seen in two similarly constructed C. albicans mdr1 mutants. Therefore, despite species-specific differences in the amino acid sequences of the Mdr1 proteins, overexpression of CaMDR1 and CdMDR1 in clinical C. albicans and C. dubliniensis strains seems to confer the same drug resistance profile in both species.
Project description:Telomeric ORFs (TLOs) in pathogenic Candida spp. encode Mediator subunits that regulate the transcription of distinct subsets of genes
Project description:Because Candida dubliniensis is closely related to Candida albicans, we tested whether it underwent white-opaque switching and mating and whether white-opaque switching depended on MTL homozygosity and mating depended on switching, as they do in C. albicans. We also tested whether C. dubliniensis could mate with C. albicans. Sequencing revealed that the MTLalpha locus of C. dubliniensis was highly similar to that of C. albicans. Hybridization with the MTLa1, MTLa2, MTLalpha1, and MTLalpha2 open reading frames of C. albicans further revealed that, as in C. albicans, natural strains of C. dubliniensis exist as a/alpha, a/a, and alpha/alpha, but the proportion of MTL homozygotes is 33%, 10 times the frequency of natural C. albicans strains. C. dubliniensis underwent white-opaque switching, and, as in C. albicans, the switching was dependent on MTL homozygosis. C. dubliniensis a/a and alpha/alpha cells also mated, and, as in C. albicans, mating was dependent on a switch from white to opaque. However, white-opaque switching occurred at unusually high frequencies, opaque cell growth was frequently aberrant, and white-opaque switching in many strains was camouflaged by an additional switching system. Mating of C. dubliniensis was far less frequent in suspension cultures, due to the absence of mating-dependent clumping. Mating did occur, however, at higher frequencies on agar or on the skin of newborn mice. The increases in MTL homozygosity, the increase in switching frequencies, the decrease in the quality of switching, and the decrease in mating efficiency all reflected a general deterioration in the regulation of developmental processes, very probably due to the very high frequency of recombination and genomic reorganization characteristic of C. dubliniensis. Finally, interspecies mating readily occurred between opaque C. dubliniensis and C. albicans strains of opposite mating type in suspension, on agar, and on mouse skin. Remarkably, the efficiency of interspecies mating was higher than intraspecies C. dubliniensis mating, and interspecies karyogamy occurred readily with apparently the same sequence of nuclear migration, fusion, and division steps observed during intraspecies C. albicans and C. dubliniensis mating and Saccharomyces cerevisiae mating.
Project description:Candida dubliniensis is the closest known relative of Candida albicans, the most pathogenic yeast species in humans. However, despite both species sharing many phenotypic characteristics, including the ability to form true hyphae, C. dubliniensis is a significantly less virulent and less versatile pathogen. Therefore, to identify C. albicans-specific genes that may be responsible for an increased capacity to cause disease, we have sequenced the C. dubliniensis genome and compared it with the known C. albicans genome sequence. Although the two genome sequences are highly similar and synteny is conserved throughout, 168 species-specific genes are identified, including some encoding known hyphal-specific virulence factors, such as the aspartyl proteinases Sap4 and Sap5 and the proposed invasin Als3. Among the 115 pseudogenes confirmed in C. dubliniensis are orthologs of several filamentous growth regulator (FGR) genes that also have suspected roles in pathogenesis. However, the principal differences in genomic repertoire concern expansion of the TLO gene family of putative transcription factors and the IFA family of putative transmembrane proteins in C. albicans, which represent novel candidate virulence-associated factors. The results suggest that the recent evolutionary histories of C. albicans and C. dubliniensis are quite different. While gene families instrumental in pathogenesis have been elaborated in C. albicans, C. dubliniensis has lost genomic capacity and key pathogenic functions. This could explain why C. albicans is a more potent pathogen in humans than C. dubliniensis.