Scanning Electron Microscopy of Candida albicans Chlamydospores

Summary:  Candida albicans chlamydospores were studied using scanning electron microscopy method. The various relationships of chlamydospore and pseudohyphae were documented as follow: terminal chlamydospore with or without suspensor cell; lateral chlamydospore with or without suspensor cell; terminal chlamydospore pairs with or without suspensor cell; lateral chlamydospore pairs without suspensor cell.
Zusammenfassung:  Die Chlamydosporen von Candida albicans wurden rasterelektronenmikroskopisch untersucht. Die unterschiedlichen Beziehungen zwischen Chlamydosporen und Pseudohyphen werden wie folgt beschrieben: Terminate Chlamydospore mit oder ohne Suspensorzelle; laterale Chlamydospore mit oder ohne Suspensorzelle; terminale Chlamydosporen-Paare mit oder ohne Suspensorzelle; laterale Chlamydosporen-Paare ohne Suspensorzelle.     

Chlamydospores of Dermatophytes

Summary: Chlamydospores of dermatophytes were differentiated from gemmae, vesicules, protoplasm reservoirs and aleuriospores. Chlamydospores were not found to be resting structures, but possess multiple budding. So-called terminal and lateral chlamydospores become generally intercalary as they get older. Thick, sausage-like hyphae which we found in the area of chlamydospore-chains are supposedly rows of pre-chlamyoospores. Heads of some racket hyphae transform into chlamydospores.
Zusammenfassung: Die Chlamydosporen der Dermatophyten werden abgegrenzt gegen Gemmen, Vesiculae, Protoplasma-Reservoire und Aleuriosporen. Diese Chlamydosporen stellen sich nicht als ruhende Strukturen dar, sondern sie zeigen multiple Sprossungsvorgänge. Sogenannte terminale und laterale Chlamydosporen werden im Verlaufe der Zeit im allgemeinen zu interkalaren Chlamydosporen. Dicke, wurstförmige Hyphen, die wir im Bereich von Chlamydosporenketten fanden, sind vermutlich Reihen von Prä-Chlamydosporen. Die Auftreibungen einiger Raquette-Hyphen wandeln sich in Chlamydosporen um.     

Chlamydospore formation on Staib agar. Observations made before Candida dubliniensis was described

When routinely using Staib agar to detect Cryptococcus neoformans in AIDS patients by the brown colour effect of its colonies, rough-looking colonies of a questionable variety of Candida albicans were also found. Microscopically, these colonies consisted of pseudohyphae with abundant masses of chlamydospores. However, the colonies of C. albicans were smooth-edged and formed by round-oval blastospores only. Such observations were made during the mycological supervision of 36 cryptococcosis cases during the 1987-94 period. All these questionable cultures of Candida spp. were discarded. However, because the corresponding photographs of and records on such strains were found to be identical with those recently published by molecular biologists under the title 'Chlamydospore formation on Staib agar as a species-specific characteristic of Candida dubliniensis' [Staib, P. & Morschh?user, J. (1999) Mycoses 42, 521-524], the present communication presents a report on such observations in a representative and exemplary case of an AIDS patient.     

Adherence of Candida Albicans to Vaginal Epithelia: Significance of Morphological Form and Effect of Ketoconazole:Adhärenz von Candida albicans an Viginalepithelien: Einfluß verschiedener Wuchsformen und von Ketokonazol.

Summary: Candida albicans yeast cells, germ tubes and hyphae of equivalent ages were tested for their ability to adhere to vaginal epithelia when the viable biomasses of the different morphological forms were standardized by ATP measurment. No significant differences were found between mean numbers of yeasts and germ tubes adhering per epithelial cell, but numbers of 6-hour-old hyphae per epithelial cell were significantly lower. However, in terms of fungal biomass attached per epithelial cell, the 6h hyphae attached significantly better than the other cell forms, and in terms of surfaces areas of fungi per epitehlial cell, yeast cells were more poorly attached than either germ tubes or hyphae. No differences were seen in adherence of C. albicans to epithelia from pregnant and nonpregnant females, but previous observations that ketoconazole can reduce attachment of C. albicans to vaginal epithelia were confirmed. Scanning electron microscopy of adherent yeast and hyphal forms showed that only the tips of germ tubes and hyphae were involved in penetration of epithelial surfaces, and in some instances the hyphal tip appeared to be the only point truly attached to the epithelial cells. Zusammenfassung: Sproßzellen, Keimschläuche und Hyphen gleichen Alters von Candida albicans wurden auf ihre Fähigkeit untersucht, an Vaginalzellen zu haften. Die Lebendmasse der verschiedenen Wuchsformen wurde dabei durcb ATP-Messungen standardisiert. Es bestand kein signifikanter Unterschied in der Adhärenz, wenn die Zahlen von Sproßzellen und Keimschläuchen miteinander verglichen wurden. Dagegen war die Zahl der pro Epithelzelle anhaftenden Hyphen deutlich geringer. Der Vedeich der Lebendmasse der Pilze zeigte, daß bezogen auf die einzehen Epithelzeilen 6 Stunden alte Hyphen signifikant besser hafteten als die anderen Zellformen. Bezog man den Vergleich auf das Maß der Zelloberfläche der Pilzelemente pro Epithelzelle, so war die Haftfäihigkeit der Sproßzellen geringer als die der beiden anderen Form-elemente. Kein Unterschied in der Adhärenz fand sich beim Vergleich von Vaginalepithelien schwangerer und nicht-whwangerer Frauen. Die Früen Die mere Beobachtung, daß Ketokonazol die Haftfägkeit von C. albicans an Vaginalepithelien verringert, konnte bestätigt werden. Rasterelektronenmikroskopie Aufnahmen ließen erkennen, daß nur die Spitzen von Keimschäiuchen und Hyphen epitheliale Oberflächen penetrieren. Bei einigen Befnnden schien es, daß ausscbließlich die Hyphenspitze emdentig an der Epitheloberfläche haftet.     

The Effect of Antimycetics in Vitro on the Growth of Candida albicans and its Developmental Forms Die Wirkung der Antimykotika in vitro auf das Wachstum von Candida albicans und auf die Morphogenese ihrer Entwicklungsformen

Summary: The inpeding effect of three antifungal substances - Clotrimazole, Nystatin and Pimaricin - on the growth of Candida albicans and the morphogenesis of its developmental forms has been studied in vitro. The antimycetics were employed in indentical concentrations, Pimaricin exerting the strongest impediment, followed by Nystatin, and this by Clotrimazole. The formation of chlamydospores and pseudomycelium on rice agar is impeded in the presence of the antifungal substances in question. Swelling and vacuolation present themselves in the various developmental forms, and these changes depend on the concentration of the antimycetics. With reference to data in the literature, the impediment of growth is considered by us to be the primary effect of the antimycetics. In all probability, it enforces itself through the impediment of the synthesis of RNA and protein, whereas swelling and vacuolation are probably secondary processes, which are the result of the autolysis taking place in the cells. Zusammenfassung: Wir haben die inhibierende Wirkung von drei antimykotischen Präparaten - Clotrimazol, Nystatin und Pimaricin - auf das Wachstum von Candida albicans und auf die Morphogenese ihrer Entwicklungsformen in vitro untersucht. Bei gleicher Konzentration hat von den drei Antimykotika Pimaricin die stärkste Hemmwirkung gezeigt, dann folgte Nystatin und schließlich Clotrimazol. Die genannten antimykotischen Mittel hemmen die Bildung von Chlamydosporen und von Pseudomycelium auf Reis-Agar. Bei den verschiedenen Entwicklungsformen treten Schwellung und Vakuolisation auf, deren Ausmaß von der Konzentration der Antibiotika abhängt. Aufgrund von Angaben der einschlägigen Literatur betrachten wir die Wachstumsinhibition als Primärwirkung der Antimykotika, die aller Wahrscheinlichkeit nach infolge der Hemmung der RNS- und der Eiweiß-Synthese zustandekommen, während die Schwellung und die Vakuolisation ein Sekundärprozeß sind, welcher infolge der sich in den Zellen abspielenden Autolyse auftritt.     

The protein secretory pathway of Candida albicans

Virulence of the opportunistic pathogen, Candida albicans , relies on an assemblage of attributes. These include the secretion of hydrolytic enzymes, cell surface adhesins, morphological transition between yeast and hyphae, phenotypic switching and biofilm formation. These diverse features are united by their dependence on the protein secretory apparatus for expression. Although the secretory apparatus of C. albicans has been studied limitedly, it appears to conform to the well-conserved eukaryotic system of vesicle-mediated transport between intracellular compartments and the cell surface. Genome comparison with Saccharomyces cerevisiae , however, shows multiple differences whose functional significance is yet unstudied. A unique aspect of the secretory pathway of C. albicans is its structural, and perhaps functional, rearrangement in hyphal vs. yeast cells. This, and evidence of non-conserved secretion mechanism(s), suggest that there is much fundamental knowledge to be derived from the analysis of secretion in C. albican s, which will be relevant to its ability to cause disease.     

Changes in Candida albicans colonization and morphology under influence of voriconazole

The aim was the investigation of fungal colonization and morphological alterations under the influence of voriconazole in an in vitro system. Voriconazole stopped growth and colonization of Candida albicans (wild type SC5314) on cover slips in microtiter plates dependent on drug concentration, the time of Candida growth before the input of voriconazole and oxygen concentration. The direct microscopy by fluorescence staining with the optical brightener Blancophor showed short bizarrely deformed mycelia looking swollen. The colonization on cover glass was diminished. Microcolonies or starting of biofilm formation as in the control was not observed. The metabolic activity was demonstrated by vital staining with FUN 1 resulting in red fluorescent structures in the yeast forms and mycelia in the controls. Under voriconazole influence the remaining cells only showed a green or pale yellow fluorescence. Most of the cells lost their metabolic activity.     

Diagnostische Bedeutung des Candida albicans abtotenden Faktors*

Zusammenfassung: In der vorliegenden Arbeit wird die Methodik des Nachweises des Candida albicans abtötenden Faktors und dessen Bewertung beschrieben. Die Untersuchung wurde an Seren von Patienten, die durch eine invasive Kandidose gefährdet sind und von gesunden Menschen durchgeführt. Der candidocide Faktor wurde bei gesunden Menschen, aber auch bei Asthmatikern, die mit Kortikosteroiden behandelt wurden, weiter bei Kranken mit Niereninsuffizienz und bei Leukämikera festgestellt Zur wesentlichen Reduktion dieses Faktors kam es bei Kranken mit Tumoren, Leberzirrhose und besonders bei Diabetikern. Diese Patienten sind deshalb gegen die invasive Form der Kandidose anfällig, denn Candida-Zellen können sich in ihrem Blut ungestört vermehren. Bei den untersuchten Kranken wurden auch komplementfixierende und präzipitierende Anti-körper vorgefunden, die nut dem erhöhten Vorkonunen von Candida im Infektionsmaterial korrelierten. Diese immunologische Reaktion spricht auch für die parasitische Wirkung der Candida-Pilze. Summary: Proof and evaluation of the candidacidal factor ist described in the sera of patients compromised of invasive candidiasis as well as in the sera of healthy persons. The candidacidal factor was found in the sera of healthy persons, of patients treated with corticosteroids, of patients with leukaemia and kidney insufficiency. The factor was substantially reduced in the sera of patients with cancer, cirrhosis and especially with diabetes mellitus. These patients are therefore sensitive to invasive candidiasis as the Candida cells can multiply uninhibited in the serum. Complement fixing antibodies as well as precipitins were found in the investigated patients. The antibodies were in correlation with the high Candida colonisation of these persons. This immunological reaction agrees with the parasitic effect of Candida in these cases.     

Virulence of Candida albicans mutants

Mutant strains of the fungal pathogen Candida albicans blocked in pyrimidine transport and salvage metabolism were tested for virulence in various animal models. The growth rate, germination and proteolytic enzyme production did not correlate with the virulence of the strains. However, a defect in the uridine transport system significantly decreased virulence in murine candidosis, although it had no effect in vaginal candidosis or in a Candida cyst model. It remains unclear whether this is due to the differing host defence mechanisms involved in systemic and superficial mycoses, or to the different requirements of the fungal systems for adherence and tissue invasion in the two types of infection.     

Candida albicans and the principle of opportunism. An essay

Zusammenfassung.   Die Begriffe Opportunismus und Opportunist werden seit langem in vielerlei Hinsicht in Politik, Sozialwissenschaften, Wirtschaftswissenschaften und Naturwissenschaften benutzt. Trotzdem sind diese Begriffe nur unzureichend definiert. Es werden die verschiedenen Bedeutungen dieser Begriffe in den unterschiedlichen Zusammenhängen mit besonderem Augenmerk auf dem mikrobiologischen Bereich diskutiert, und hier im Speziellen in Zusammenhang mit Candida albicans , die neben Pseudomonas aeruginosa am häufigsten als Opportunist bezeichnet oder des Opportunismus verdächtigt wird.     

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.     

Zur Korrelation von Biotyp und Antimykotika-sensibilität bei Candida albicans. On the Correlation of Biotype and Antimicrobial Susceptibility in Candida albicans

Zusammenfassung: 54 rezente Candida albicans-isolate von Haut und angrenzenden Schleimhäuten wurden zunächst auf Grand der von Odds und Abbott 1983 (23) angegebenen Eigenschaften biotypisiert. Anschließend wurde ihre Antimykotikaempfindlichkeit nüt Hilfe von Agardilutions- und Mikrodilutionstest untersucht. Insgesamt fanden sich 42 verschiedene Biotypen, die im häufigsten Falle viermal vertreten waren. Die minimalen Hemmkonzentrationen (MHK) von 5-Fluorcytosin (5FC) schwankten zwischen 0,016 und mehr als 128 mg/l. Die IC₃₀-Werte nach Johnson et al. 1984 (17) reichten von 0,25 bis mehr als 64 mg/l. Bei Ketoconazol lagen die MHK-Werte zwischen 32 und 128 mg/l. Wie angesichts dieser geringen Bandbreite nicht anders zu erwarten, fand sich bei diesem Antimykotikum keine klare Korrelation zwischen Biotyp und minimalen Hemmkonzentrationen. Bei 5FC ließ sich demgegenüber eine derartige Korrelation innerhalb der Biotypengruppen 1 und 3 nachweisen: Innerhalb der Gruppe 1 erwiesen sich die mit den Kodenummern 4 und 5 als besonders resistent. Innerhalb der Gruppe 3 bestand ein entsprechender Zusammenhang mit der Kodenummer 7. Summary: 54 recent clinical mucocutaneous isolates of Candida albicans were differentiated using several different characteristics as described 1983 by Odds and Abbott (23). In addition antimicrobial susceptibility testing was performed using both the agardilution and microdilution method. 42 different biotypes were found, the most frequent one turning up four times. The minimum inhibitory concentrations (MIC) of 5-fluorocytosine (5FC) ranged from 0.016 to more than 128 mg/l. The IC₃₀ values according to Johnson et al. 1984 (17) lay between 0.25 and more than 64 mg/l. With ketoconazole the MICs ranged from 32 to 128 mg/l. As could be exspected from this small range no clear correlation could be demonstrated between biotype and MIC with this antimycotic. With 5FC, however, there was a correlation within the biotype groups 1 and 3: within group 1 the strains carrying the code numbers 4 and 5 were especially resistent to 5FC. Within group 3 this especially was the case with code number 7.     

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.     

Stimulierung der Filamentbildung von Candida albicans durch Kammerwasser

Zusammenfassung:  Kammerwasser von Kaninchen stimuliert das filamentöse Wachstum von Candida albicans in stammabhängiger Weise. Die Zugabe von Kammerwasser zu Blastosporen des Stammes DSM 70010 in Nährbouillon und Inkubation bei 37°C für 18–24 h führt ebenso zur Filamentbildung wie die Inkubation der Blastosporen mit Blutse-rum. Bei Stamm CBS 2730 ergibt nur Serum Filamentwachstum, nicht aber Kammerwasser. Falls im Verlauf einer Candida -Keratitis die Descement'sche Membran durchwachsen wird, könnte die Filamentstimulation durch Kammerwasser klinische Relevanz erlangen.
Summary:  Aqueous humor of rabbit eyes stimulates the filamentous growth of Candida albicans in a strain-dependent manner. Addition of aqueous humor to blastospores of strain DSM 70010 in nutrient broth and incubation at 37°C for 18–24 h yields filaments which are comparable to filaments after incubation of blastospores with blood serum. In strain CBS 2730 serum promotes the formation of filaments but not aqueous humor. If in the course of Candida keratitis the fungus penetrates Descement's membrane, the filamentation in aqueous humor may influence the outcome of the mycosis.     

Ultrastructural characters defining the terms 'yeast cell', 'pseudomycelium' and 'true mycelium' in Candida albicans

The discrimination of the morphological elements, 'yeast cell', 'pseudomycelium' and 'true mycelium' is indispensable in the mycological and histopathological diagnostics of fungal infections caused by Candida albicans. However, precisely defined distinctive criteria are lacking in the medical mycological literature. Such criteria based upon electron microscopical studies of the cell wall, are described.     

The involvement of physico-chemical interactions in the adhesion of Candida albicans and Candida dubliniensis to epithelial cells

Candida albicans and Candida dubliniensis are two pathogenic yeasts particularly hazardous to immunocompromised patients. Adhesion of yeast cells to epithelium is considered one of the virulence factors and its study is of major importance. The main aim of this study was the comparison of the influence of physico-chemical properties on the adhesion of C. albicans and C. dubliniensis to epithelium. Two strains of each Candida species were used in the adhesion assays to HeLa cells. Adhered cells were enumerated by direct microscopic images observation. Yeast cell surface tension parameters and degree of hydrophobicity were determined by contact angle measurement. Pseudohyphae and hyphae formation was analysed by scanning electron microscopy. Yeast cells presented no statistical differences concerning their physico-chemical surface properties. However, the extent of adhesion to epithelium was different among the four strains. As general conclusion, yeast adhesion to epithelium seems to be strain-dependent and not directly correlated with pseudohyphae formation.     

Differences in patient risk factors and source of candidaemia caused by Candida albicans and Candida glabrata

There is an increasing frequency of candidaemia caused by Candida glabrata which has decreased in vitro susceptibility to fluconazole. Differences in risk factors for candidaemia caused by C. glabrata and C. albicans have not been formally evaluated in a diverse patient group. We performed a retrospective study of adult inpatients from January 1, 2003 to April 30, 2008 with C. glabrata and C. albicans candidaemia at a single tertiary care centre in Detroit, Michigan to evaluate for differences in risk factors and presumed source of infection in these groups. Patients’ underlying conditions, risk factors and source of infection (probable or definite) were compared. Among 119 patients, 80 (67.2%) were C. albicans and 39 (32.8%) C. glabrata. Using logistic regression analysis, patients with C. glabrata infection were more likely to have diabetes mellitus (OR 2.43; 95% CI, 1.06–5.54) and abdominal source of infection (OR 4.53, 95% CI, 1.72–11.92). Mortality rates in the two groups were similar. Patients with C. glabrata candidaemia are more likely to be diabetic and have an abdominal source of infection compared with patients with C. albicans.     

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.     

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.