Candida Albicans Blastospores Research Articles

Candida albicans blastospores and hyphae respond differentially to fluconazole: additional virulence factor of germination process

Background: The morphologic transition from yeasts (blastospores) to hyphae, through the germination process, is a crucial virulence factor of Candida albicans ( C. albicans), as it enhances adherence, tissue invasion and biofilm formation. The study aimed at comparing fluconazole susceptibility patterns of C. albicans yeast and germinated forms. Methods: Six C. albicans strains, including C. albicans ATCC 10231 and 5 clinical isolates were tested in yeast forms and germinated forms for fluconazole susceptibility using CLSI M27-A3 broth microdilution and CLSI M44-A2 disk diffusion reference methods. Results: Fluconazole minimal inhibitory concentrations (MICs) were 0.125-1 µg/ml and 0.25-4 µg/ml for yeasts and germinated forms, respectively. The sensitivities of yeasts were higher than those of their corresponding germinated forms with 1 to 4-fold dilutions differences. The MIC50s of yeasts and germinated forms were 0.125 and 1 µg/ml, respectively, with 3-fold dilution difference, while MIC90s were 1 and 4 µg/ml for yeasts and germinated forms, respectively, with 2-fold dilution difference. Means of inhibition zones were 31-46.7 mm and 29.3-43.7 mm for yeasts and germinated forms, respectively. The germinated forms of all tested strains had inhibition zones smaller than those of their corresponding yeast forms with 0.2-4.8 mm differences. There was significant statistical difference in sizes of inhibition zones between yeasts and germinated forms (p = 0.043). Conclusions: Yeast and hyphae of C. albicans, both in planktonic state, respond differentially to fluconazole, as the hyphae are less sensitive than their yeasts counterparts, which can be considered as a virulence factor for hyphae, which should be considered in treatment plans of candidal infections.

Treg dysfunction in aging oral mucosa

Abstract Aging leads to an increased susceptibility to infections, a phenomenon called immunosenescence. An imbalance between proinflammatory and anti-inflammatory mechanisms is also observed during aging. However, mechanisms of mucosal tissue specific infections and inflammation, and the underlying immune cell alterations in these tissues are incompletely understood. Here our objective was to understand oral mucosal immune cells, especially regulatory T cells, a population of CD4+ T cells that are central to immunomodulation. We infected young (2–3 months) and aged (>18–24months) C57BL/6 mice with a sublingual inoculation of 2 × 107 Candida albicans (SC5314) blastospores under anesthesia. We reinfected the mice on day 7 and day 16 after primary infection to mimic recurrent/secondary infections, monitored their weight loss and tongue immunopathology. Control mice were sham infected with PBS. We isolated oral draining cervical lymph nodes (CLN) cells and mouse oral intraepithelial and lamina propria cells (MOIL), and determined the frequency, absolute cell numbers and functions of T and inflammatory CD4 T cells ex vivo before and after infection. We found that aged mice continuously lost weight during the course of the recurrent infections, whereas young mice regained their weight after every infection. Immunosenescent phenotype in aged mice was associated with heightened tongue immunopathology, compared to young mice. Increased oral inflammation phenotype was associated with Treg dysfunctions in local oral tissue. Taken together, our study begins to define T cell dysfunctions and inflammatory processes underlying decline of the immune responses in oral mucosa.

Sporicidal effect of amorolfine and other antimycotics used in the therapy of fungal nail infections.

Although topical antifungal therapies for treating onychomycosis are available, the cure rate is unsatisfactorily low with a simultaneously high risk of recurrence. One reason might be the formation of dormant fungal cells by the pathogen, known as spores, which can survive in the affected nail keratin, thereby evading the effect of antifungal drugs. In this in vitro study, the ability of amorolfine and four other antimycotics (ciclopirox, bifonazole, terbinafine and fluconazole) to kill microconidia of the dermatophyte Trichophyton rubrum, chlamydospores of the dermatophyte Epidermophyton floccosum and blastospores of the yeast Candida albicans was extensively studied as these fungi occur predominantly in onychomycosis. The effectiveness of all five antimycotics depended on the drug concentration and the incubation time: a concentration of 10-1000 times the minimum inhibitory concentration against growing hyphae cells is needed to exert a sporicidal action. Amorolfine and ciclopirox showed the same sporicidal efficacy and kinetics for all three varieties of spores. Both were more effective than fluconazole and bifonazole against microconidia and chlamydospores as well as slightly more potent against chlamydospores and blastospores than terbinafine after 4 days of incubation and at concentrations of ≥10 μg ml(-1). Finally, sporicidal activity on the tested strains was demonstrated for all five different antimycotics used for onychomycosis treatment.

Morphologic and morphometric analysis of alternations in the oral cavity caused by Candida albicans: Experimental work

Candidiasis has become a human disease of increasing importance in the last decades. The aim of the study is to establish pathomorphological alterations caused by the blastospores of the Candida albicans as well as morphometric alterations. The experiment was carried out on 2.5-month-old rats, weighting 110-130 g. The study sample was divided into the animals infected by a submucous inoculation in the periodontal region and the controls. The gingival specimens were taken, preparations were done and stained by the hematoxylin-eosin and Periodic acid Schiff methods. The following alterations were found out by the stereological analysis: an average volume of nuclei of the gingival epithelial cells was 111.82 microm3 (SD = 25.34) on the first day. A statistically significant increase in the volume of nuclei in the experimental group began to occur from the fourth day (202.97 microm3; SD = 31.16, p < 0.05) and the highest value of the nuclei volume was found out on the eight day of the experiment (316.83 microm3; SD = 40.15). Blastospores of Candida albicans are pathogenic for the gingival tissue where they cause degenerative necrotic alterations of the granulomatous character and after the fourth day from the inoculation, the development of the pseudohyphae was observed. The obtained values of stereologic measurement show the acute increase in the volume of nuclei.

Frequency distribution of Candida albicans blastospores adhered to mucosal epithelial cells in-vitro.

Although several methods are available for examination of microbial adherence to epithelial cells, these do not distinguish between adherence of viable and non-viable micro-organisms. This study reports the use of acridine orange-stained blastospores of Candida albicans in conjunction with direct epifluorescence microscopy to determine viable (orange-fluorescing) and non-viable (green-fluorescing) blastospore adherence to buccal epithelial cells. The method was also employed to examine the effects of chlorhexidine treatment at subminimum inhibitory concentrations on the adherence of viable and non-viable blastospores. There was good correlation in the assessment of blastospore viability between the direct epifluorescence microscopy technique and the standard serial dilution and plating method for viable counting, confirming the reliability of direct epifluorescence microscopy. Chlorhexidine treatment before acridine orange staining did not alter this assessment of viability. Blastospore adherence to buccal epithelial cells resulted in a similarly skewed distribution whether examined using a crystal violet stain in conjunction with light microscopy or using direct epifluorescence microscopy, therefore validating the direct epifluorescence microscopy technique for the enumeration of blastospore adherence. Chlorhexidine treatment (0.0005% v/v, 30 min) of either blastospores or buccal epithelial cells altered the distribution of adherent blastospores per epithelial cell by increasing the number of epithelial cells having no adherent blastospores. No differences in adherence were, however, observed between blastospore or epithelial cells after treatment with this agent. Examination of the adherence of viable and non-viable blastospores to buccal epithelial cells using direct epifluorescence microscopy revealed a greater adherence capacity of non-viable than viable blastospores for buccal epithelial cells. Treatment of blastospores with chlorhexidine altered the frequency distributions of viable and non-viable blastospores with lower numbers of blastospores adherent per epithelial cell. The larger reduction in adherent viable blastospores in comparison with their non-viable counterparts is, however, an important observation which might have clinical relevance. Microbial cells adhere to epithelial cells resulting in a skewed distribution; study of this distribution gives useful information about the adherence process. Viable and non-viable components of a microbial population have different adherence capabilities and treatment of such populations with an antimicrobial agent exerting anti-adherent activity at sub-minimum inhibitory concentrations reduces the amount of adherence of these viable/non-viable components to different extents.

Interaction and intracellular killing of Candida albicans blastospores by human polymorphonuclear leucocytes, monocytes and monocyte-derived macrophages in aerobic and anaerobic conditions.

Polymorphonuclear leucocytes (PMN), monocytes and monocyte-derived macrophages were capable of interacting with opsonized C. albicans in both aerobic and anaerobic conditions. Superoxide anion release by these cells was inhibited in anaerobic conditions while lysozyme release and phagocytosis were equally efficient in both aerobic and anaerobic conditions. All cell types tested were capable of intracellular killing of C. albicans and this appeared to be maximum at 6 h for monocytes and macrophages and 24 h for PMN. Monocytes killed the lowest number of organisms, 1 x 10(6), and the killing was similar for aerobic and anaerobic conditions. In contrast, PMN and macrophages demonstrated greater killing of C. albicans in aerobic conditions compared with anaerobic conditions; PMN killed 1.9 x 10(6) organisms and macrophages 3 x 10(6) when incubated anaerobically. Inhibitors of oxygen metabolism decreased intracellular killing of C. albicans by macrophages and PMN in aerobic but not anaerobic conditions. The oxygen reaction products involved in the killing of C. albicans appeared to be different however: macrophage killing was decreased by superoxide anion and hydrogen peroxide inhibitors. PMN killing was decreased by superoxide anion, hydrogen peroxide, hypochlorous acid and hydroxyl radical inhibitors. The present study shows that although monocytes, macrophages and PMN function similarly in their interaction with C. albicans, they appear to use different oxygen reactive products for the intracellular killing of C. albicans.

Retention of microbial cells in substratum surface features of micrometer and sub-micrometer dimensions

Surfaces were produced with defined topographical features and surface chemistry. Silicon wafers, and wafers with attached nucleopore filters and quantifoils were coated with titanium using ion beam sputtering technology. Irregularly spaced, but regularly featured surface pits, sizes 0.2 and 0.5 μm, and regularly spaced pits with regular features (1 and 2 μm) diameter were produced. The smallest surface feature that could be successfully produced using this system was of diameter 0.2 μm. Ra, the average absolute deviation of the roughness irregularities from the mean line over one sampling length, Rz, the difference in height between the average of the five highest peaks, and the five lowest valleys along the assessment length of the profile and surface area values increased with surface feature size, with Ra values of 0.04–0.217 μm. There was no significant difference between the contact angles observed for smooth titanium surfaces with 0.2 and 0.5 μm features. However, a significant difference in contact angle was observed between the 1 and 2 μm featured surfaces ( p < 0.005). Substrata were used in microbial retention assays, using a range of unrelated, differently sized microorganisms. Staphylococcus aureus (cells 0.5–1 μm diameter) were retained in the highest numbers. S. aureus was well retained in the 0.5 μm sized pits and began to accumulate within larger surface features. Rod shaped Pseudomonas aeruginosa (1 μm × 3 μm) were preferentially retained, often end on, within the 1 μm surface features. Some daughter cells of Candida albicans blastospores were retained in 2 μm pits. For S. aureus and P. aeruginosa, the greatest numbers of cells were retained in the largest (2 μm) surface features. The number of C. albicans was similar across all the surfaces. The use of defined surfaces in microbial retention assays may lead to a better understanding of the interaction occurring between cells and surface features.

Detection of anti-Candida antibodies by the indirect immunofluorescence assay in patients with cancer in the orofacial region.

An indirect immunofluorescence assay was performed to detect antibodies to Candida albicans blastospores and germ tubes. Serum specimens were obtained from 82 patients with neoplastic diseases in the orofacial region and thrush of the oral mucosa. C. albicans was identified in the oral cavity of 63 patients investigated but serum anti-Candida antibodies were detected in only 23 of them. Serological examination showed that titers of antibodies to C. albicans blastospores ranged from 1:20 to 1:1280. High titers from 1:640 to 1:1280 were detected in patients without antibiotic, cytostatic, or radiotherapeutic treatment. The titers of antibodies to C. albicans germ tubes ranged from 1:20 to 1:640. Our results indicate that titers of antibodies to the C. albicans germ tubes were lower and were detected in a smaller number of patients.

Diagnosis and Therapeutic Management of Candidiasis in Falcons in Saudi Arabia

From 1998 to 2001, a total of 3760 falcons were presented to the Falcon Medical Research Hospital of the Fahad bin Sultan Falcon Center. From this total, 11 (0.3%) adult (>1-year-old) falcons, including 8 saker falcons (Falco cherrug), 2 peregrine falcons (Falco peregrinus), and 1 lanner falcon (Falco biarmicus), were admitted for treatment of candidiasis. Affected falcons showed general clinical signs including reduced to total absence of appetite, shredding and flicking of food, regurgitation, and progressive weight loss. The diagnosis of clinical candidiasis was made from clinical signs, endoscopic examination of the upper digestive tract, histologic observation of typical Candida albicans blastospores on samples obtained from the crop, and positive fungal cultures. Therapy consisted of applying a miconazole gel directly to the thoracic esophagus and crop twice daily for 5 consecutive days. Miconazole proved to be an effective, fast, and inexpensive antifungal agent suitable for the treatment of candidiasis in captive falcons.

In vitro Effect of Amikacin, Imipenem, Cefodizime, IFNα-2a Alone and Combinations of Antibiotics with IFNα-2a on Polymorphonuclear Leukocyte Function in Chronic Hepatitis Patients

The in vitro effect of amikacin (8 µg/ml), imipenem (30 µg/ml), cefodizime (10 µg/ml), interferon α-2a (IFNα-2a) (10 IU/ml) and antibiotic combinations with IFNα-2a on polymorphonuclear leukocyte (PMN) functions (phagocytosis and intracellular killing of Candida albicans blastospores) was investigated in chronic hepatitis B patients. Phagocytosis and candidacidal activity was not affected after pretreatment of PMNs with amikacin and imipenem (p > 0.05). Phagocytic activity was enhanced after pretreatment of PMNs with cefodizime and IFNα-2a compared with that of control PMNs (p < 0.05), but candidacidal activity was not affected by the same drugs (p > 0.05). Phagocytic and intracellular activity of PMNs were not affected by combinations of IFNα-2a and antibiotics (p > 0.05).

Experimental murine mycotic mastitis: a sensitive and lenient model for studies of antifungal chemotherapy.

The murine model of mycotic mastitis was used to study the efficacy of amphotericin B (AmB). Twenty-four BALB/cJ mice at the fifth day of lactation were anesthetized and inoculated through the teat canal (two glands) with 50 microl suspension containing 5.0 x 10(7) cfu ml(-1) Candida albicans blastospores. Mice were randomly divided into two groups: untreated controls and AmB treated. Animals were euthanized 3 and 6 days after infection and treatment (4 mg kg(-1) per day intraperitoneally). The fungal burden of the mammary gland was determined by quantitative cultures. The number of C. albicans cells recovered from mammary gland homogenates were significantly lower in the AmB treated animals (both 3 and 6 days post-infection) than in the untreated controls (P<0.007 and P<0.003, respectively). The mammary glands of all untreated control animals showed marked neutrophilic infiltration, severe necrosis, and presence of blastospores, hyphae and pseudohyphae. In contrast, 10 of 12 animals treated with AmB showed only a mild neutrophilic infiltration which was restricted to alveoli and excretory ducts. All extra-mammary organs were free of infection in both groups. The results demonstrate that the murine mycotic mastitis model is suitable for investigations of new antifungal compounds. In addition, this model is more lenient than the systemic candidiasis models.

Light and Electron Microscopic Findings in a Model of Human Cutaneous Candidosis Based on Reconstructed Human Epidermis Following the Topical Application of Different Econazole Formulations

The effects of two commercially available econazole formulations (econazole nitrate cream, econazole liposome gel) on uninfected reconstructed human epidermis and on a model of human cutaneous candidosis were investigated. The morphological alterations of the reconstructed epidermis after infection and treatment were analysed with light and electron microscopy. The most important Candida albicans-specific alterations of the recently established in vitro model of human cutaneous candidosis were scaling, hyperkeratosis, parakeratosis, dyskeratosis and spongiosis. A single application of the cream to the uninfected reconstructed epidermis caused more epidermal barrier damage and irritative toxic effects than the liposome gel. Treatment of the modelled human cutaneous candidosis with the cream also resulted in increased toxic effects, e.g., enhancement of scaling with invasion of Candida albicans blastospores into the stratum corneum and intracellular vacuoles. After application of the liposomal preparation invasion of Candida albicans in the stratum corneum could not be detected and toxic effects were reduced. Some of the Candida albicans-specific alterations such as hyperkeratosis, focal thickening of the stratum corneum, dyskeratosis and parakeratosis were completely eliminated. The liposomal formulation increased slightly the morphological alterations of the blastospores. Remnants of the cream formulation could be detected only very rarely in the stratum corneum or the blastospores. The liposomal preparation showed a strong affinity for the Candida albicans cells and the stratum corneum. Intact liposomes could even be observed in the intercellular spaces of the upper stratum corneum. As successful treatment depends on the ability to target the liposomal agent to the wanted site of action, this might be useful for more effective treatment of cutaneous candidosis.

A model of human cutaneous candidosis based on reconstructed human epidermis for the light and electron microscopic study of pathogenesis and treatment

The use of reconstructed human epidermis provided the basis for an in vitro model of human cutaneous candidosis. Candida albicans blastospores on the surface of reconstructed human epidermis provoked the following changes within 72 h: superficial keratin degradation, scaling, hyperkeratosis, parakeratosis, dyskeratosis representing hyperproliferative stress, spongiosis, and vesiculation. Great differences in the intensity of these reactions of intact reconstructed human epidermis or chemically or mechanically damaged reconstructed human epidermis illustrate the importance of the stratum corneum as a barrier. Uninfected reconstructed human epidermis showed prominent cell proliferation representing wound healing 72 h after mechanical or chemical pretreatment. These signs of repair were blocked in the presence of C. albicans and the blastospores were able to invade the stratum corneum. When desmosomes were accessible, a high affinity of C. albicans blastospores to these structures was observed. A single application of an econazole liposome dispersion decreased scaling, hyperkeratosis and dyskeratosis. Morphological alterations of C. albicans blastospores after treatment with the econazole liposome dispersion in the proposed ill vitro model were identical, as described in established animal models. This reconstructed human epidermis model of cutaneous disease may provide insight into the pathogenesis and treatment of cutaneous candidosis and may provide a substitute for animal models and investigations on humans.

A rapid evaluation of phagocytosis and killing of Candida albicans by CD13+ leukocytes

Flow cytometry can be adopted for routine monitoring of the immune functions of human polymorphonuclear leukocytes (PMNs) in several disease states. We recently developed a rapid and reproducible assay for the evaluation of the phagocytosis and killing of Candida albicans blastospores by human PMNs. Whole blood leukocytes were incubated with opsonized fluorescein isothiocyanate-labeled (FITC-labeled) blastospores for phagocytosis and killing assays. To discriminate between ingested, membrane-bound and free C. albicans blastospores, ethidium bromide (EtBr) was added to the samples prior to the flow cytometric analysis. EtBr induces a loss of green fluorescence in non-phagocytized C. albicans blastospores. Phagocytosis is determined by gating the phagocytes and calculating the percentage of phagocyte-associated green fluorescent cells. Intracellular killing is determined by first lysing phagocytes by hypotonic shock and then adding propidium iodide (PI) in order to identify red dead blastospores. Killing is measured in terms of the percentage of double-marked blastospore cells. We suggest that this method is a reliable and inexpensive technique to evaluate the immune reactivity of PMNs and peripheral blood monocytes (PBMs) in cases of immunosuppression.

Characterisation of microbial adherence to epithelial cells following biocide treatment by analysis of area under the frequency distribution curve

The aim of this study was to further characterise the anti-adherence effects of certain antimicrobial agents by evaluating their effects on the frequency distribution of Candida albicans blastospores (MEN strain) to buccal epithelial cells (BEC). A light microscopic assay was employed to evaluate the effects of super- and sub-minimal inhibitory concentrations of chlorhexidine gluconate, dequalinium chloride, cetrimide and cetylpyridinium chloride on candidal adherence in vitro. Frequency distributions were plotted to describe the adherence profile of each treatment and mathematical descriptions of these distributions were performed by calculating the areas under the frequency distribution curves (AUC). Comparison of the AUC of each treatment and control (sterile water treatment) was performed using a one-way analysis of variance. Each concentration of all antimicrobial agents significantly reduced the AUC. These reductions were due to greater percentages of BEC showing lower numbers (zero, one, two) of adherent blastospores and also lower percentages of BEC showing higher numbers of adherent blastospores in comparison to control. Generally, the anti-adherence effects of these antimicrobial agents (as measured using AUC determinations) were greater following treatment of blastospores and were observed with both super- and sub-minimal inhibitory concentrations. These findings suggest a greater affinity of the antimicrobial agents for the blastospore surface and also a greater ability to block, either directly or indirectly via steric hinderance, adhesins on the blastospore surface than receptors on the surface of BEC. It is suggested that to fully appreciate the anti-adherence potential of antimicrobial agents, the frequency distribution of microbial adherence data should be considered, as this will provide information on specific adherence population shifts, in conjunction with their effects on the mean number of adherent microorganisms per epithelial cell. In addition, the anti-adherence activity observed for the antimicrobial agents examined indicates their potential use in the prophylaxis of superficial candidal infections in the oral cavity.

Effects of amphotericin B on glucose metabolism in Candida albicans blastospores evidenced by 13C NMR.

Candida albicans blastospores harvested from 8- (exponential phase) or 48-h (stationary phase) cultures were incubated with 60 x 10(-3)M [1-(13)C]glucose with or without 10(-4)M amphotericin B (AmB). The utilization of [1-(13)C]glucose was monitored by in vivo 13C NMR under anaerobiosis. With exponential phase cells, in the presence of AmB, the consumption of glucose and the production of ethanol, trehalose, and glycerol continuously decreased with time, and after 25 min, the metabolism was blocked. On stationary phase cells AmB had almost no effect on glucose metabolism. Comparison with previous experiments evidenced that AmB induced first K+ leakage, then acidification, and finally a stop of the metabolism. In parallel, in vitro 13C NMR spectra were performed on supernatants and cell-free extracts of yeast suspension incubated under the same conditions. For both exponential and stationary phase cells, AmB induced an increase in the membrane permeability to glycerol; no change was observed for the other metabolites.

The Effects of Cetylpyridinium Chloride on the Cell Surface Charge (Zeta Potential) of Candida albicans: Implications for Anti‐adherence Effects

This study examined the effects of time of treatment with, and treatment concentration of cetylpyridinium chloride (CPC) on the zeta potential of blastospores of Candida albicans (MEN strain, exponential and stationary phases). The ranges of treatment times and concentrations of CPC employed were selected according to their reported abilities to reduce the adherence of this strain of C. albicans to human buccal epithelial cells in-vitro. The zeta potentials of both exponential and stationary phase blastospores were negative. However, these were significantly decreased following treatment with CPC (50 μg mL−1) over the range of treatment times examined (0·5–30 min), and also following treatment with CPC (0·5–50 μg mL−1) for a fixed period (15 min). It is suggested that the overall effects of CPC on blastospore adherence are not directly attributable to the observed effects on zeta potential, as these would be expected to enhance adherence. The anti-adherence effects of CPC are most likely due to its effects on other blastospore surface properties, e.g. surface morphology, surface hydrophobicity.

Modifications of pH and K + gradients in Candida albicans blastospores induced by Amphotericin B. A 31P NMR and K + atomic absorption study

Candida albicans blastospores harvested from 8 h (exponential) or 48 h (stationary) cultures were incubated with increasing doses of amphotericin B (AmB). The time course of H + influx and K + efflux was monitored by in vivo 31P NMR and K + atomic absorption respectively. AmB was shown to be more active on exponential phase cells than on stationary phase cells. For both growth phases, K + leakage ocurred before pH acidification. In light of these results, together with iodoacetate experiments, it seems difficult to assert that K + leakage is a secondary effect resulting from an increase in the permeability to protons, as formerly proposed. In addition, no H + over K + selectivity of pores formed by AmB could be detected. Finally, some unexpected results were afforded by 31P NMR experiments: a broadening of P i signals was detected on exponential phase cell spectra when the blastospores were incubated with 10 −3 and 10 −4 M AmB reflecting a transient heterogeneity of the intracellular pH within the cell population. For stationary phase blastospores, two subpopulations (IIa and IIb) were detected; population IIb, with a more acidic pH i, was much more sensitive to AmB action.

Intracellular pH of Candida albicans blastospores as measured by laser microspectrofluorimetry and 31P-NMR

The intracellular pH (pH i) of Candida albicans blastospores harvested from 8 h or 48 h cultures was determined under identical experimental conditions by two different techniques: 31P-NMR and laser microspectrofluorimetry. Time dependence of pH i was monitored by 31P-NMR on the whole cell population. Microspectrofluorimetry, after loading of the cells with SNARF-1, enabled the determination of pH i in isolated cells and its distribution among the cell population. By this method, the vacuolar pH could not be distinguished from the cytoplasmic pH in C. albicans blastospores, but alkalization of pH i was observed at the beginning of germ tubes. The absolute values of pH i determined by 31P-NMR were slightly different from those obtained by laser microspectrofluorimetry. However, the pH distributions in the cell population were converging. For blastospores in exponential phase a gaussian distribution of pH i was observed with both methods, the cells maintained a steady pH i value when the external pH was varied from 5.5 to 8.5. For cells in stationary phase two pools were identified: the combination of the two techniques demonstrated the presence of two different subpopulations. One of these population (with lower pH) was able to commute to the other one with time as shown by 31P-NMR kinetics. This information is reported here for the first time in C. albicans.

Primary interactions of three quaternary ammonium compounds with blastospores of Candida albicans (MEN strain).

The absorption of three quaternary ammonium compounds (QAC), cetylpyridinium chloride, cetrimide and benzalkonium chloride, onto the surface of blastospores of Candida albicans (MEN strain) was examined at room temperature. Equilibrium uptake occurred in less than 30 seconds for cetylpyridinium chloride and cetrimide whereas 5 min contact time was required for benzalkonium chloride. The adsorption of all three agents may be mathematically described as Langmuirian and hence a concentration-dependent formation of drug-monolayer on the surface of the blastospore occurred. From this the number of molecules adsorbed onto the surface of a single blastospore was calculated to be 1.33 x 10(12), 3.17 x 10(12) and 2.32 x 10(12) for cetylpyridinium chloride, cetrimide and benzalkonium chloride, respectively. These dissimilarities are most likely due to differences in the orientations of both the cationic nitrogen atom and the accompanying lipophilic portions of each QAC at the blastospore surface. Relating these observations to the known antiadherence effects of cetylpyridinium chloride and cetrimide, it can be concluded that monolayer coverage of the blastospore surface with QAC does not account for the observed reduced adherence. This suggests that the anti-adherence effects are due to either direct interaction with, or steric blockade of, adhesions on the blastospore surface.