Effects of repeated low-dose UVB irradiation on the hyphal growth of Candida albicans

Ultraviolet B light (UVB) can have negative phototropic effects on fungi. Candida albicans is often found on human skin exposed to UVB. Therefore, it is of medical interest to know whether a negative phototropic response to UVB irradiation can support an invasive growth of this potentially dangerous agent. In our study we investigated how repeated irradiation with low doses of UVB can influence the hyphal growth of C. albicans. Six randomly chosen strains of C. albicans were tested. Formation of hyphae was induced and maintained within transparent agar plates. The fungi were exposed to UVB three times daily for 7 days from either the observe or the reverse side during incubation. The wavelength spectrum was in the range of 310-315 nm, single doses were between 0.0018 and 0.432 J cm(-2). After 7 days the morphology and growth direction of C. albicans cells were determined microscopically. All six strains showed a common and dose-dependent response to UVB irradiation: the progression of hyphal growth was inhibited, no phototropic effects were seen and as a new finding an increased formation of blastospores was observed. We conclude that an irradiation of human skin colonized by C. albicans with doses of UVB that can occur under natural or artificial conditions is unlikely to trigger skin invasion by C. albicans.     

Spiral hyphae of Candida albicans formed in anaerobic culture

Candida albicans from a patient with dental caries grew on minimal medium consisting of agar supplemented with magnesium chloride and sodium phosphate. Hyphal growth was observed when the yeast was cultured between 26 °C and 28 °C under aerobic conditions, and typical chlamydospores were formed. However, when the yeast was cultured at the same temperature under anaerobic conditions, curly hyphae developed on the surface of the medium, but no chlamydospores were formed. This phenomenon was also observed if the culture was started under aerobic conditions but was continued under anaerobic conditions.     

Adherence ability of Candida africana: a comparative study with Candida albicans and Candida dubliniensis

In this study, we compared the adherence ability to human Hela cells and biofilm formation of three closely related Candida yeast. In our experiments, Candida africana showed poor adhesion ability to human Hela cells and the absence of biofilm formation on polyvinyl chloride strips. Conversely, Candida albicans and Candida dubliniensis formed mature biofilms and stable attachment to Hela cells. To our knowledge, this is the first comparative study reporting data on biofilm formation and adherence to human Hela cells by C. africana.     

Role of pH response in Candida albicans virulence

Over 25 years ago it was noted that the pH of the culture medium influenced germ tube formation of Candida albicans, an opportunistic fungal pathogen. This simple observation has been the stimulus for a number of investigations to discern the mechanisms controlling this response and the significance of this response to the biology of C. albicans. Recent studies have demonstrated that a signaling pathway conserved in several fungal species regulates this morphological response to ambient pH and controls the pH-conditional expression of multiple genes. Significantly, C. albicans responds to the pH of the host niche and this response is critical for virulence.     

Hydrolytic enzymes as virulence factors of Candida albicans

Candida albicans is a facultative pathogenic micro-organism that has developed several virulence traits enabling invasion of host tissues and avoidance of host defence mechanisms. Virulence factors that contribute to this process are the hydrolytic enzymes. Most of them are extracellularly secreted by the fungus. The most discussed hydrolytic enzymes produced by C. albicans are secreted aspartic proteinases (Saps). The role of these Saps for C. albicans infections was carefully evaluated in numerous studies, whereas only little is known about the physiological role of the secreted phospholipases (PL) and almost nothing about the involvement of lipases (Lip) in virulence. They may play an important role in the pathogenicity of candidosis and their hydrolytic activity probably has a number of possible functions in addition to the simple role of digesting molecules for nutrition. Saps as the best-studied member of this group of hydrolytic enzymes contribute to host tissue invasion by digesting or destroying cell membranes and by degrading host surface molecules. There is also some evidence that hydrolytic enzymes are able to attack cells and molecules of the host immune system to avoid or resist antimicrobial activity. High hydrolytic activity with broad substrate specificity has been found in several Candida species, most notably in C. albicans. This activity is attributed to multigene families with at least 10 members for Saps and Lips and several members for PL B. Distinct members of these gene families are differentially regulated in various Candida infections. In future, prevention and control of Candida infections might be achieved by pharmacological or immunological tools specifically modulated to inhibit virulence factors, e.g. the family of Saps.     

Inhibitory activity of thymol against the formation and viability of Candida albicans hyphae

As the capacity of Candida albicans to produce hyphae is considered an important virulence factor in the pathogenesis of candiasis, the aim of this study was to investigate whether thymol, the major component of thyme oil, can interfere with the filamentous forms of Candida albicans and their viability. The morphological transition from yeasts to filamentous forms was investigated by analysing the morphological index (MI), which classifies the differentiated forms and blastoconidia; viability was investigated by means of fluorescence microscopy using a new SYTO-9 and propidium iodide method previously used to stain only blastoconidia. Without thymol, there was an average of 94.00 +/- 3.06% hyphal forms. After 6 h of incubation with 1x MIC (125 microg ml(-1)), 1/2x MIC and 1/4x MIC of thymol, filamentation was, respectively, 14.33 +/- 8.25%, 28.33 +/- 7.17% and 45.67 +/- 8.09% in comparison with control (all statistically significant). In the absence of thymol, viable cells accounted for an average of 93.00 +/- 4.00% whereas, after 6 h of incubation with 1x MIC, 1/2x MIC and 1/4x MIC of thymol, the presence of 54.33 +/- 1.86%, 29.00 +/- 3.61% and 23.00 +/- 2.52% of yellow-orange coloured forms indicated damaged membranes and reduced viability. Our findings show that thymol interferes with the formation and viability of hyphae. This can be attributed to the characteristics of thymol disturbing Candida cell membranes and metabolism, probably by affecting fungal cell-wall synthesising enzymes.     

Antifungal activity of amphotericin B,caspofungin and posaconazole on Candida albicans biofilms in intermediate and mature development phases

The aim of this study was to examine the antifungal activity of amphotericin B, caspofungin and posaconazole on Candida albicans biofilms in the intermediate and mature development phases. Candida albicans biofilms, previously grown for either 24, 48 or 72 h in 96‐well microtitre plates, were treated for 48 h with amphotericin B, caspofungin or posaconazole in increasing concentrations according to the respective minimal inhibitory concentration (MIC) determined for planktonic cells (1–128 × MIC). The biofilms were quantified using the mean optical density (OD) determined by XTT assay. Antifungal activities were expressed as percentage of reduction in OD of drug‐treated biofilms compared to untreated biofilms. To test the fungicidal activity of antifungal agents, the unfixed biofilms were scraped off and seeded to Sabouraud agar. Caspofungin and amphotericin B showed higher activity against C. albicans biofilm grown for 24 h and 72 h (≥50% reduction of OD) than biofilms grown for 48 h, whereas posaconazole showed similar, but reduced activity against all phases of C. albicans biofilm (≤50% reduction of OD). Caspofungin at 1–4 × MIC achieved the greatest decrease in the biofilm OD grown for 24, 48 and 72 h, whereas amphotericin B showed dose‐dependent activity. However, all tested antifungals failed to reach fungicidal activity in all biofilm development phases.     

Effect of surface treatments of titanium on amphotericin B‐treated Candida albicans persister cells

Although persister cells in Candida albicans biofilm may contribute to its increased resistance to antifungal drugs, little information is available on the formation of Candida persister cells on titanium surfaces. The effect of different surface treatments of Ti on persister cells was determined in the present study. Titanium discs were surface‐treated by three different methods (Group A – polishing, Group B – sandblasting followed by acid‐etching, and Group C – sandblasting alone). Persister cells of two C. albicans strains, namely ATCC 90028 and HK30Aa, in biofilm and planktonic states, were measured as the percentage of colony forming units remaining after 24 h incubation with various concentrations of amphotericin B. No persister cells were detected in the planktonic cultures. However, 1.5%, 0.1% and 2.4%C. albicans ATCC 90028 persister cells were detected at an AmB concentration of 64 μg ml^?1 in groups A, B and C, respectively; and 0.3%, 0.2% and 0.6% for groups A, B and C, respectively, for HK30Aa. Group C of C. albicans ATCC 90028 appeared to provide a surface relatively unfavourable for the development of persister cells (P < 0.01). Whether these results may have implications on the clinical performance of titanium implants warrants further investigation.     

The Effect of Antimycotics on Secretory Acid Proteinase of Candida albicans

Summary

The effect of antimycotics on secretory aspartate (acid) proteinase, a virulence enzyme of Candida albicans, was investigated.

The conditions of the study were such as to induce proteinase production in the stationary phase of growth (25-40 hours), when no antifungal tested, except the polyene derivative methyl partricin, significantly reduced the viability of the culture.

Among azole derivatives, fenticonazole (FZ) but not miconazole, fluconazole or ketoconazole, exerted strong inhibition on proteinase, in typical dose-diphasic pattern, (0.01 μg/ml; 1-10 μg/ml). 5-fluorocytosine (5-FC) was also inhibitory at a dose interval 1-10 μg/ml. In all cases, the inhibition concerned the synthesis of the enzyme rather that its activity as suggested by the results of comparative ELISA, SDS-PAGE and spectrophotometric methods of proteinase detection.

Finally, the inhibition of proteinase production by FZ and 5-FC mainly reflected the effect of these antimycotics on general protein synthesis.     

Four cases of Candida albicans infections with isolates developing pink colonies on CHROMagar Candida plates

Candida albicans, the most commonly isolated yeast species, is typically identified by its green colony-colour on CHROMagar Candida plates. We here report four cases of Candida albicans infections, in which the initial identification was non-albicans isolates due to a clear pink colour of the colonies on CHROMagar Candida plates. However, classical phenotypic criteria, biochemical assimilation pattern and molecular characterisation identified all four isolates as C. albicans isolates.     

The Effect of Dexamethason and Oxytetracyclin on the Growth of Candida albicans

Summary: Candida albicans was grown in Sabouraud's dextrose liquid medium alone and with decadron®*, terramycin®** and both. Decadron® was incorporated in the strengths of 0.01, 0.05 and 0.1 % and terramycin® in 0.1, 0.2 and 0.5% strengths. Dry weight and pH values of the growths obtained were determined. The maximum increase in values occured when C. albicans was grown with decadron and terramycin together. The increase with decadron® or terramycin® individually or in combination was proportional to the increase in the concentration of the additives and was statistically significant when compared to the respective controls. The lesions in rabbit cornea were more severe with treated than untreated Candida.
Zusammenfassung: Candida albicans wurde in flüssigem Sabouraud-Dextrose-Me-dium allein, unter Zusatz von Dexamethason, von Oxytetracyclin und von Dexamethason + Oxytetracyclin gezüchtet. Dexamethason wurde in einer Konzentration von 0,01, 0,05 und 0,1 % angewendet, bei Oxytetracyclin betrugen die Konzentrationen 0,1, 0,2 und 0,5 %. Das Trockengewicht der gewachsenen Hefen und die pH-Werte im Nährboden wurden bestimmt. Der höchste Anstieg von Trockengewicht und pH-Wert trat auf, wenn Candida albicans unter gleichzeitigem Zusatz von Dexamethason und Oxytetracyclin gezüchtet wurde. Der Anstieg bei Züchtung mit Dexamethason oder Oxytetracyclin einzeln oder in Kombination war proportional zum Anstieg der Konzentrationen dieser Zusätze und war statistisch gegenüber den zugehörigen Kontrollen signifikant. An der Cornea des Kaninchenauges hervorgerufene Läsionen waren aus-geprägter, wenn Candida albicans-Zellen verwendet wurden, die im Nährmedium mit den genannten Zusätzen angezüchtet worden waren.     

The Effect of Dexamethason and Oxytetracyclin on the Growth of Candida albicans

Summary: Candida albicans was grown in Sabouraud's dextrose liquid medium alone and with decadron®*, terramycin®** and both. Decadron® was incorporated in the strengths of 0.01, 0.05 and 0.1 % and terramycin® in 0.1, 0.2 and 0.5% strengths. Dry weight and pH values of the growths obtained were determined. The maximum increase in values occured when C. albicans was grown with decadron and terramycin together. The increase with decadron® or terramycin® individually or in combination was proportional to the increase in the concentration of the additives and was statistically significant when compared to the respective controls. The lesions in rabbit cornea were more severe with treated than untreated Candida.
Zusammenfassung: Candida albicans wurde in flüssigem Sabouraud-Dextrose-Me-dium allein, unter Zusatz von Dexamethason, von Oxytetracyclin und von Dexamethason + Oxytetracyclin gezüchtet. Dexamethason wurde in einer Konzentration von 0,01, 0,05 und 0,1 % angewendet, bei Oxytetracyclin betrugen die Konzentrationen 0,1, 0,2 und 0,5 %. Das Trockengewicht der gewachsenen Hefen und die pH-Werte im Nährboden wurden bestimmt. Der höchste Anstieg von Trockengewicht und pH-Wert trat auf, wenn Candida albicans unter gleichzeitigem Zusatz von Dexamethason und Oxytetracyclin gezüchtet wurde. Der Anstieg bei Züchtung mit Dexamethason oder Oxytetracyclin einzeln oder in Kombination war proportional zum Anstieg der Konzentrationen dieser Zusätze und war statistisch gegenüber den zugehörigen Kontrollen signifikant. An der Cornea des Kaninchenauges hervorgerufene Läsionen waren aus-geprägter, wenn Candida albicans-Zellen verwendet wurden, die im Nährmedium mit den genannten Zusätzen angezüchtet worden waren.     

Comparative virulence of Candida albicans strains in CFW1 mice and Sprague-Dawley rats

Summary. To verify host-species specificities of virulence of Candida albicans in experimental systemic mycoses, 10 ATCC strains of Candida albicans were compared for their virulence in CFW1 mice and Sprague-Dawley rats. Virulence was parallel in mice and rats, four strains were avirulent (ATCC 10231, 18804, 38245, 44831), one strain had an intermediate virulence (ATCC 32354), and five strains (ATCC 10261, 44373, 44505, 62342, 90028) were highly virulent in both host species. Infection doses of 2 times 10⁶CFU per mouse and 5 times 10⁶ CFU per rat were comparable with respect to mortality of animals within 10 days; this represents a 4:1 ratio on the basis of body weight. In Sprague-Dawley rats haemorrhage occurred in infections with all virulent strains which was not observed in CFW1 mice.
Zusammenfassung. Zur Bewertung Wirtsspezies-spezifischer Virulenzunterschiede von Candida albicans bei tierexperimentellen systemischen Mykosen, führten wir für 10 ATCC-Stämme von Candida albicans Virulenzvergleiche für CFW1-Mäuse und Sprague-Dawley-Ratten durch. Die Virulenz der Stämme war parallel in Maus und Ratte. Vier Stämme waren apathogen (ATCC 10231, 18804, 38245, 44831), ein Stamm zeigte eine mitlere Virulenz (ATCC 32354), fünf Stämme (ATCC 10261, 44373, 44505, 62342, 90028) waren hochgradig virulent für beide Wirtsspezies. Infektionsdosen von 2 times 10⁶ KBE pro Maus und 5 times 10⁶ KBE pro Ratte waren für die Letalität über 10 Tage vergleichbar, auf das Körpergewicht bezogen wird bei CFW1-Mäusen eine vierfach höhere Infektionsdosis benötigt als bei Sprague-Dawley-Ratten. Bei Sprague-Dawley-Ratten wurden Blutungen bei der Infektion mit allen virulenten Stämmen beobachtet, was bei CFW1-Mäusen nicht auftrat.     

Growth of Candida albicans on the stratum corneum of diabetic and non-diabetic patients

We have used a novel approach to in vitro culture of Candida albicans on cyanoacrylate skin surface strippings. It appears that with this model yeasts and hyphae grew on large surfaces of stratum corneum. The area of extension of the fungal growth was larger on stratum corneum taken from diabetic than from non-diabetic volunteers.     

Effect of serum and surface characteristics on Candida albicans biofilm formation

Candida spp. biofilms can be established on a wide range of materials, including implanted medical devices, and can display a resistant phenotype to antifungal drugs. Several factors, including host and surface properties, may influence the establishment and the development of Candida albicans biofilms on biotic and abiotic surfaces. We therefore selected a collection of C. albicans clinical isolates to evaluate the effect of surface and serum on biofilm attachment and development. Disc coupons from the CDC biofilm reactor were used in a well plate assay to study biofilm production on six different surfaces with or without the addition of serum: polycarbonate, polystyrene, stainless steel, Teflon, polyvinyl chloride or hydroxyapatite. Our results showed that serum increases in vitro C. albicans biofilm formation on a wide range of distinct surfaces including metallic and non‐metallic materials, and that roughness and hydrophobicity can modulate C. albicans biofilm formation. These findings were also confirmed by scanning electron microscopy and it revealed the deposition of extracellular material on hyphae attached to a solid surface. Interestingly, adhesion can be significantly increased in the early stages of colonisation when serum is provided as a conditioning film in a surface‐dependent manner.     

Action of the Ketoconazole-Alkylaminoethylglycine Combination on Candida albicans

Summary

High concentrations of ketoconazole (Ktc) exhibit poor antimicrobial activity in microbicidal test.

Alkylaminoethylglycine (AAEG), an amphoteric surfaceactive agent, employed in subinhibitory concentrations, can enhance the action of ketoconazole. The Ktc-AAEG combination causes a higher rate of potassium ion release compared to both drugs separately.

The increased ketoconazole activity may be ascribed to the membrane damage indicated by increased permeability to potassium ions.     

Fluorometric determination of acid proteinase activity in Candida albicans strains from diabetic patients with vulvovaginal candidiasis

Vulvovaginal candidiasis is one of the most frequent disorders in obstetrics and gynaecology. Approximately three-quarters of all adult women experience at least one episode of vulvovaginal candidiasis during their life span. Diabetes mellitus (DM) increases the rate of vaginal colonisation and infection with Candida species. The secreted acid proteinase might be especially relevant in the pathogenesis of vulvovaginal candidiasis. The aim of this study was to determine the acid proteinase activity in the samples of Candida albicans from diabetic patients with vulvovaginal candidiasis by a fluorometric method. Vaginal swabs were taken from 33 women (aged between 22 and 57 years) having symptoms of vaginitis. Patients were divided into three groups: control group, controlled diabetic group and uncontrolled diabetic group. The proteinase activity in the culture supernatants was determined by a modified fluorometric method. Acid proteinase activities were significantly increased in the uncontrolled diabetic group in comparison with both the control group and the controlled diabetic group (P < 0.05). Acid proteinase may play an important role in C. albicans pathogenesis in diabetic patients. Improving glucose control may reduce the risk of Candida colonisation and potentially symptomatic infection, among women with diabetes and hence may be useful even for weaker enzyme activity measurements.     

The effects of rose bengal- and erythrosine-mediated photodynamic therapy on Candida albicans

The aim of this study was to evaluate the effects of photodynamic therapy (PDT) using rose bengal or erythrosine with light emitting diode (LED) on Candida albicans planktonic cultures and biofilms. Seven C. albicans clinical strains and one standard strain (ATCC 18804) were used. Planktonic cultures and biofilms of each C. albicans strain were submitted to the following experimental conditions: (a) treatment with rose bengal and LED (RB+L+); (b) treatment with erythrosine and LED (E+L+); and (c) control group, without LED irradiation or photosensitiser treatment (P-L-). After irradiation of the planktonic cultures and biofilms, the cultures were seeded onto Sabouraud dextrose agar (37 °C at 48 h) for counting of colony-forming units (CFU ml(-1) ) followed by posterior anova and Tukey's test analyses (P < 0.05). The biofilms were analysed using scanning electron microscopy (SEM). The results revealed a significant reduction of planktonic cultures (3.45 log(10) and 1.97 log(10) ) and of biofilms (<1 log(10) ) for cultures that were subjected to PDT mediated using either erythrosine or rose bengal, respectively. The SEM data revealed that the PDT was effective in reducing and destroying of C. albicans blastoconidia and hyphae. The results show that erythrosine- and rose bengal-mediated PDT with LED irradiation is effective in treating C. albicans.