Heterogeneous distribution of Candida albicans cell-surface antigens demonstrated with an Als1-specific monoclonal antibody

Despite an abundance of data describing expression of genes in the Candida albicans ALS (agglutinin-like sequence) gene family, little is known about the production of Als proteins on individual cells, their spatial localization or stability. Als proteins are most commonly discussed with respect to function in adhesion of C. albicans to host and abiotic surfaces. Development of a mAb specific for Als1, one of the eight large glycoproteins encoded by the ALS family, provided the opportunity to detect Als1 during growth of yeast and hyphae, both in vitro and in vivo, and to demonstrate the utility of the mAb in blocking C. albicans adhesion to host cells. Although most C. albicans yeast cells in a saturated culture are Als1-negative by indirect immunofluorescence, Als1 is detected on the surface of nearly all cells shortly after transfer into fresh growth medium. Als1 covers the yeast cell surface, with the exception of bud scars. Daughters of the inoculum cells, and sometimes granddaughters, also have detectable Als1, but Als1 is not detectable on cells from subsequent generations. On germ tubes and hyphae, most Als1 is localized proximal to the mother yeast. Once deposited on yeasts or hyphae, Als1 persists long after the culture has reached saturation. Growth stage-dependent production of Als1, coupled with its persistence on the cell surface, results in a heterogeneous population of cells within a C. albicans culture. Anti-Als1 immunolabelling patterns vary depending on the source of the C. albicans cells, with obvious differences between cells recovered from culture and those from a murine model of disseminated candidiasis. Results from this work highlight the temporal parallels for ALS1 expression and Als1 production in yeasts and germ tubes, the specialized spatial localization and persistence of Als1 on the C. albicans cell surface, and the differences in Als1 localization that occur in vitro and in vivo

Evaluation of the Role of Candida albicans Agglutinin-Like Sequence (Als) Proteins in Human Oral Epithelial Cell Interactions

The fungus C. albicans uses adhesins to interact with human epithelial surfaces in the processes of colonization and pathogenesis. The C. albicans ALS (agglutinin-like sequence) gene family encodes eight large cell-surface glycoproteins (Als1-Als7 and Als9) that have adhesive function. This study utilized C. albicans Δals mutant strains to investigate the role of the Als family in oral epithelial cell adhesion and damage, cytokine induction and activation of a MAPK-based (MKP1/c-Fos) signaling pathway that discriminates between yeast and hyphae. Of the eight Δals mutants tested, only the Δals3 strain showed significant reductions in oral epithelial cell adhesion and damage, and cytokine production. High fungal:epithelial cell multiplicities of infection were able to rescue the cell damage and cytokine production phenotypes, demonstrating the importance of fungal burden in mucosal infections. Despite its adhesion, damage and cytokine induction phenotypes, the Δals3 strain induced MKP1 phosphorylation and c-Fos production to a similar extent as control cells. Our data demonstrate that Als3 is involved directly in epithelial adhesion but indirectly in cell damage and cytokine induction, and is not the factor targeted by oral epithelial cells to discriminate between the yeast and hyphal form of C. albicans

A Human-Curated Annotation of the Candida albicans Genome

Recent sequencing and assembly of the genome for the fungal pathogen Candida albicans used simple automated procedures for the identification of putative genes. We have reviewed the entire assembly, both by hand and with additional bioinformatic resources, to accurately map and describe 6,354 genes and to identify 246 genes whose original database entries contained sequencing errors (or possibly mutations) that affect their reading frame. Comparison with other fungal genomes permitted the identification of numerous fungus-specific genes that might be targeted for antifungal therapy. We also observed that, compared to other fungi, the protein-coding sequences in the C. albicans genome are especially rich in short sequence repeats. Finally, our improved annotation permitted a detailed analysis of several multigene families, and comparative genomic studies showed that C. albicans has a far greater catabolic range, encoding respiratory Complex 1, several novel oxidoreductases and ketone body degrading enzymes, malonyl-CoA and enoyl-CoA carriers, several novel amino acid degrading enzymes, a variety of secreted catabolic lipases and proteases, and numerous transporters to assimilate the resulting nutrients. The results of these efforts will ensure that the Candida research community has uniform and comprehensive genomic information for medical research as well as for future diagnostic and therapeutic applications

Evolution of pathogenicity and sexual reproduction in eight Candida genomes

Candida species are the most common cause of opportunistic fungal infection worldwide. Here we report the genome sequences of six Candida species and compare these and related pathogens and non-pathogens. There are significant expansions of cell wall, secreted and transporter gene families in pathogenic species, suggesting adaptations associated with virulence. Large genomic tracts are homozygous in three diploid species, possibly resulting from recent recombination events. Surprisingly, key components of the mating and meiosis pathways are missing from several species. These include major differences at the mating-type loci (MTL); Lodderomyces elongisporus lacks MTL, and components of the a1/2 cell identity determinant were lost in other species, raising questions about how mating and cell types are controlled. Analysis of the CUG leucine-to-serine genetic-code change reveals that 99% of ancestral CUG codons were erased and new ones arose elsewhere. Lastly, we revise the Candida albicans gene catalogue, identifying many new genes.publishe

Cellular and Molecular Biology of Candida albicans Estrogen Response

Candida albicans is the most common etiological agent of vaginal candidiasis. Elevated host estrogen levels and the incidence of vaginal candidiasis are positively associated. Elevated estrogen levels may affect host and/or fungal cells. This study investigates the effect of 17-β-estradiol, 17-α-estradiol, ethynyl estradiol, and estriol on several C. albicans strains at concentrations ranging from 10(−5) to 10(−10) M. The addition of 17-β-estradiol or ethynyl estradiol to C. albicans cells caused an increase in the number of cells forming germ tubes and an increase in germ tube length in a dose- and strain-dependent manner. The addition of 17-α-estradiol or estriol did not have a significant effect on germ tube formation by the cultured cells. Exposure to exogenous estrogens did not significantly change the biomass of any C. albicans culture tested. The transcriptional profile of estrogen-treated C. albicans cells showed increased expression of CDR1 and CDR2 across several strain-estrogen concentration-time point combinations, suggesting that these genes are the most responsive to estrogen exposure. Analysis of strain DSY654, which lacks the CDR1 and CDR2 coding sequences, showed a significantly decreased number of germ tube-forming cells in the presence of 17-β-estradiol. PDR16 was the most highly up-regulated gene in strain DSY654 under these growth conditions. The cell biology and gene expression data from this study led to a model that proposes how components of the phospholipid and sterol metabolic pathways may interact to affect C. albicans germ tube formation and length

Comparison between Candida albicans Agglutinin-Like Sequence Gene Expression Patterns in Human Clinical Specimens and Models of Vaginal Candidiasis

Expression of the eight genes in the Candida albicans agglutinin-like sequence (ALS) family was studied by reverse transcription-PCR of RNA isolated from clinical vaginal fluid specimens and vaginal candidiasis model systems. Although expression of all ALS genes was detected across the set of clinical specimens, ALS1, ALS2, ALS3, and ALS9 transcripts were detected most frequently, and expression of ALS4 and ALS5 was detected least frequently. Laboratory strain 3153A and two C. albicans strains isolated from the clinical specimens were studied using two models of vaginal candidiasis to determine how closely these models mimicked the clinical specimens at the level of gene expression. ALS gene expression patterns in a murine vaginitis model were identical to those from the clinical specimens. Expression of more ALS genes was detected in specimens collected 7 days after infection compared to those collected at 4 days. Similar patterns of ALS gene expression were observed when the three C. albicans strains were tested in the reconstituted human vaginal epithelium model. In this model, expression of ALS4, ALS5, ALS6, and ALS7 was least frequently detected. Negative or weakened signals for ALS4 expression were observed at early time points, suggesting that ALS4 expression, which was strong in the inoculum cells, was down-regulated upon contact of C. albicans with vaginal epithelial cells in this model. The data presented here support the conclusion of host-site-specific influences on ALS gene expression and validate the use of the experimental models for evaluating the phenotype of als/als mutant strains