Alvin L. Rogers

Variation in adhesion and cell surface hydrophobicity in Candida albicans white and opaque phenotypes

A previous study had established that a select group of pathogenic isolates of Candida albicans was capable of switching heritably, reversibly and at a high frequency (10−2 to 10−3) between two phenotypes (‘white’ or ‘opaque’) readily distinguishable by the size, shape, and color of colonies formed on agar at 25°C. This paper describes experiments designed to determine the ability of these two phenotypes to attach to buccal epithelial cells (BECs) and plastic, and to compare the cell surface hydrophobicities of white and opaque phenotypes from three clinical isolates. ‘White cells’ were found to be significantly more adhesive to BECs, and a strong correlation was also found between phenotype adhesiveness and the percentage of BECs to which C. albicans had attached. The percentage of BECs with one or more attached C. albicans was approximately 90% for the white phenotype and approximately 50% for the opaque phenotype. ‘Opaque cells’, in contrast, were twice as hydrophobic as white cells, and the percentage of opaque cells bound to BECs by coadhesion was also double that of white cells. The differences in adhesion to plastic between the two phenotypes were not statistically significant and there was no distinct trend to suggest which phenotype might be more adhesive to plastic. These results indicate that several factors are involved in the adhesion of C. albicans to plastic, and confirm the hypothesis that cell surface hydrophobicity is of minor importance in direct adhesion to epithelial cells but that it may contribute to indirect attachment to epithelial cells by promoting yeast coadhesion. Moreover, the data presented in this paper also revealed that under identical growth conditions, adhesion of C. albicans was significantly altered depending on the phenotypic state of the organism tested. Therefore, because C. albicans can switch at a high frequency to various phenotypes in vitro, it may be that in future adhesion studies involving Candida the phenotypic state of the organism at the time of testing will have to be determined. Otherwise, the results, even within the same laboratory, may be difficult to interpret.

Dermatophytosis of the scalp: Incidence, immune response, and epidemiology

Tinea capitis remains a common infection among the pediatric population of North America. The ‘gray patch’ Microsporum audouinii infections of the 1950s have been supplanted by the ‘black dot’ ringworm of Trichophyton tonsurans. The clinical presentation of T. tonsurans infection is quite variable and may be related to specific host T-lymphocyte response. This dermatophytosis is most frequently incurred from contact with an infected child either directly or via a variety of fomites. Current studies indicate that an asymptomatic adult carrier state may also exist which could contribute to the morbidity of this mycosis.

Human pathogenic fungi recovered from Brasilian soil.

A total of 202 soil samples from areas around the cities of Belo Horizonte, Santos, São Paulo, Piracicaba and Itatiaia park were examined by various techniques for the presence of fungi that may be pathogenic to humans and animals. The fungi isolated were:Microsporon gypseum (40),M. cookei (2),Keratinomyces ajelloi (2),T. mentagrophytes (9),T. terrestre (49),T. verrucosum (2),Allescheria boydii (1),Aspergillus fumigatus (2),Candida albicans (21),Cryptococcus neoformans (4), andSporotrichum schenckii (4). Many of the pathogenic fungi were found in habitats where they might be expected to occur in relation to activities of humans and animals. The probable basis for the apparent absence ofParacoccidioides brasiliensis from soil samples was discussed. The following organisms have apparently been reported for the first time from soil samples in Brasil;T. mentagrophytes, T. terrestre, T. verrucosum, A. fumigatus, C. albicans, andS. schenckii.

Candida albicans resistance to 5-fluorocytosine: frequency of partially resistant strains among clinical isolates.

Resistance to 5-fluorocytosine was studied in 137 independent Candida albicans clinical isolates. Seventy-eight isolates (57%) were susceptible; 51 isolates (37%) were partially resistant; 8 isolates (6%) were highly resistant. All partially resistant isolates gave rise to variants which were highly resistant. Some susceptible isolates gave rise to variants which were highly resistant; two such isolates were shown to be heterozygous for resistance, and these isolates define a new type of heterozygote. A partially resistant isolate gave rise to resistant variants which were auxotrophic for lysine; this result was interpreted as preliminary evidence that the allele which determined resistance was linked to an allele which determined auxotrophy for lysine. It is suggested that heterozygotes constitute a source of preexisting mutant alleles which determine resistance, and that 5-fluorocytosine treatment of infections due to heterozygotes may result in significant selection for resistant variants. A simple screening procedure is described by which partially resistant strains may be recognized. Images

Influence of mucosal cell origin on the in vitro adherence of Candida albicans: Are mucosal cells from different sources equivalent?

The influence of collecting mucosal cells from various anatomical sites, and varying the date of collection and cell donor on adhesion of Candida albicans to human epithelial cells was examined by using an in vitro adherence assay. Examination of buccal mucosal cells from twenty-four donors showed statistically significant differences in the number of attached yeasts between individuals. Sex did not exert a significant influence on adhesion. Examination of buccal mucosal cells from ten donors collected on five different dates revealed that yeast attachment to mucosal epithelial cells varied significantly within subjects across time. Epithelial cells from some donors manifested greater date-to-date variations in yeast adhesion than others. Adherence of Candida to mucosal cells from three anatomical sites (mouth, vagina and urinary tract) collected from ten different donors was also tested. Yeast adherence to buccal cells was highest, lowest using urinary tract cells, while vaginal epithelium was intermediate. Adherence to mucosal cells from three sites was significantly different both within and between individuals although some subjects manifested larger variations than others. These data suggest that the in vitro adherence of Candida albicans is influenced by mucosal cell donor, date of collection and body site of origin. Mucosal cells from different sources do not appear to be equivalent in receptiveness to C. albicans and this might explain some of the discrepancies observed when adhesion studies performed by different investigators are compared. The existing need for a more uniform methodology with which to pursue studies on fungal attachment to mucosal surfaces is emphasized.

Segregation of 5-Fluorocytosine-Resistant Variants by Candida albicans

Spontaneous production of 5-fluorocytosine-resistant variants by three Candida albicans isolates is due to segregation from a preexisting heterozygous state. Images

Effect of attachment of anticandidal antibody to the surfaces of liposomes encapsulating amphotericin B in the treatment of murine candidiasis.

The effect produced by antibody specific to Candida albicans when attached to liposomes containing amphotericin B was studied in vivo. Liposomal amphotericin B bearing specific immunoglobulin (LAMB-Ab) was compared with the unencapsulated drug (fAMB) and other liposomal amphotericin B formulations in the short-term survival (21 days) of mice with disseminated candidiasis. Both the treatment and prophylaxis of the murine model of candidiasis were explored in these trials. LAMB-Ab increased survival rates in the model more than other liposomal preparations containing amphotericin B. Liposomal amphotericin B compounds as a group prolonged survival over fAMB. Liposomal preparations used for comparison included liposomes with attached nonspecific antibody (LAMB-Ab-), liposomes without antibody (LAMB), and liposomes with unattached specific antibody (LAMB+).

Inhibition of adherence ofCandida albicans to human epithelial cells

A mannose-specific lectin, Concanavalin A, was used to pretreatCandida albicans before using the yeats in anin vitro adherence assay. Adherence to buccal cells was inhibited but could be restored by preincubation of the lectin with a specific haptenic sugar, a-D-methylmannopyranoside, prior in the assay but not by using D-galactose, D-ribose and D-raffinose, sugars which the lectin does not recognize.

Development of amphotericin B liposomes bearing antibody specific to Candida albicans.

SummaryLiposomes expressing external antibody specific for Candida albicans and encapsulating amphotericin B were developed and characterized in this study. Antibody was first modified by the covalent attachment of palmitic acid residues. Liposomes were produced by reverse-phase evaporation and modified antibody was incorporated into these liposomes via the hydrophobic interaction between the palmitic acid and the phospholipids composing the liposomes. The liposomes were characterized as to the amount of amphotericin B by spectroscopy and for the presence of antibody by protein analysis and secondary immunolabeling by fluorescent and electron microscopic methods. Immunogold labeling showed that the antibody was being expressed externally on the liposomes in the electron microscopic studies and the specificity of these liposomes for C. albicans was observed by secondary immunofluorescence.

Aberrant Histoplasma capsulatum confirmation of identity by a chemiluminescence-labeled DNA probe

A cottony, light tan, filamentous fungus with pear-shaped microconidia and lacking tuberculated macroconidia was isolated from a bronchial lavage specimen. Subculture on several media at 37 degrees C failed to convert the fungus to a yeast form after several weeks; attempts at in vivo conversion in mice were also unsuccessful. Sera obtained several months apart showed M bands with Histoplasma capsulatum (HC) antigen by immunodiffusion and an increase in complement fixation titers with mycelial and yeast phase antigens of HC. Parallel identity was obtained on two occasions with exoantigen culture confirmation reagents for HC from Immuno-Mycologics as well as one of identity with Nolan reagents. Extracts from four Chrysosporium spp. strains had no identity reactions with HC with either kit. The fungus was identified as HC by the Accuprobe Histoplasma chemiluminescence-labeled DNA probe directed at ribosomal RNA, whereas all four Chrysosporium spp. isolates tested negative. DNA probes are a fast and accurate method to confirm the identity of aberrant fungal isolates.