Amphotericin B-induced in vitro postantifungal effect on Candida species of oral origin.

OBJECTIVE: The aim of this investigation was to measure the postantifungal effect (PAFE) of 6 different oral Candida species following exposure to amphotericin B. MATERIALS AND METHODS: Five oral isolates each of Candida albicans, Candida tropicalis, Candida krusei, Candida parapsilosis, Candida glabrata and Candida guilliermondii (total of 30 isolates) were examined for the presence of PAFE after 1 h of exposure to the minimum inhibitory concentration of amphotericin B. The PAFE was determined as the difference in time (hours) required for the growth of the drug-free control and the drug-exposed test cultures to increase to 0.05 absorbance level following removal of amphotericin B. RESULTS: The mean duration of amphotericin B-induced PAFE was lowest for C. albicans (5.91 ± 0.31 h) and greatest for C. parapsilosis (12.72 ± 0.11 h), while C. guilliermondii (8.32 ± 0.33 h), C. glabrata (8.43 ± 0.21 h), C. krusei (9.68 ± 0.23 h) and C. tropicalis (10.98 ± 0.18 h) elicited intermediate values. CONCLUSION: Even a limited exposure to sublethal concentrations of amphotericin B suppressed growth of Candida species of oral origin. The significant variation in amphotericin B-induced PAFE amongst different Candida species may have clinical implications in terms of amphotericin B regimens used in the management of oral candidiasis.

Rapid differentiation of Candida dubliniensis from Candida albicans by early D-xylose assimilation.

OBJECTIVE: To determine if D-xylose (XYL) and/or α-methyl-D-glucoside (MDG) assimilation can be used reliably as a rapid test to differentiate Candida dubliniensis from Candida albicans at an earlier time point such as 2 h after inoculation. MATERIALS AND METHODS: Thirty isolates of C. albicans and C. dubliniensis recovered from anatomical sites and clinical specimens were used. Isolates were inoculated into the API 20C AUX yeast identification system, and incubated at 30°C. XYL and MDG assimilations were read at 2-hour intervals beginning 2 h after the initial inoculation and up to 24 h of incubation; thereafter, results were read after 48 and 72 h. RESULTS: Twenty-nine (97%) C. albicans isolates had assimilated XYL at 16 h and, by 24 h, all isolates were positive for XYL assimilation. None of the C. dubliniensis isolates assimilated XYL. The MDG assimilation revealed that 24, 40, 92 and 100% of C. albicans isolates became positive after 16, 24, 48 and 72 h of incubation, respectively, whereas only 3% of C. dubliniensis isolates assimilated MDG after 72 h. CONCLUSIONS: The findings showed that it is possible to rapidly differentiate C. albicans from C. dubliniensis isolates using the API 20C AUX carbohydrate assimilation kits after 16 h of incubation at 30°C based on the XYL assimilation.

Harassment of newly admitted undergraduates by senior students in a Faculty of Dentistry in Sri Lanka.

BACKGROUND: Harassment of new students by senior colleagues appears to be widespread in the industrialised countries. Although 'ragging' of new entrants to universities in Sri Lanka gets frequently publicised, its prevalence, severity and the consequences have not been documented. AIMS: This study aims to ascertain the extent of mistreatment of new dental students, the measures they take when harassed and any resulting negative effects. METHODS: We surveyed the year 2008 Dental students using 80 statements dealing with verbal/emotional, sexual and physical harassment. Sixty five students (91.5%) responded anonymously indicating whether a specific action occurred, the degree to which it affected them and any action taken to deal with it. RESULTS: Fifty percent of students had experienced mistreatment. Verbal and emotional abuse was more frequent than sexual or physical. Eighteen percent experienced sexual harassment, with a significantly higher proportion of males than females reporting it. A fifth of the students had upsetting memories of the event. Eighty five percent of the respondents stated that they did not suffer any ethnic or racial discrimination. CONCLUSIONS: Emotional harassment of new students by the seniors is a pervasive, yet under-reported problem. Definitive interventions need to be implemented to prevent untoward consequences that can undermine the educational goals of training.

Knowledge and attitudes towards HIV/AIDS amongst Kuwait University dental students.

The HIV and AIDS have emerged as complex health threats to the world population. As future dentists, it is pertinent that the dental students have sufficient knowledge and a positive approach towards the disease. Accordingly, the aim of this study was to assess the HIV/AIDS related knowledge and attitudes amongst clinical dental students at Kuwait University. A cross-sectional survey was conducted amongst the clinical dental students using a structured questionnaire with 60 questions to examine their knowledge under various categories and 13 questions to examine their attitudes towards the disease. The survey revealed that almost 58% of the respondents demonstrated a high level of knowledge (mean score: 45.23 ± 4.35 SD). Majority of the students (63.6%) expressed negative attitude (mean score: 5.36 ± 2.56 SD). The mean knowledge score of the fifth year dental students was significantly higher (P = 0.022) than that of the final year dental students regarding the knowledge of virus and disease process. However, no significant difference was observed with respect to other knowledge categories. Despite their high level of knowledge, the majority displayed a negative attitude towards HIV/AIDS. Hence, the findings imply that there is a need to address, more clearly, the students' misconceptions and attitudes towards the disease.

Antimicrobials as a contributory factor in oral candidosis--a brief overview.

The advent of the human immunodeficiency virus infection and the increasing prevalence of compromised individuals in the community due to modern therapeutic advances have resulted in a resurgence of opportunistic infections, including oral candidosis, which is by far the most common oral fungal infection in man. Broad-spectrum antibiotics used in the treatment of a wide range of disease conditions have also been attributed as a predisposing factor of oral candidosis. In this mini review we discuss the research findings on the relationship between antibiotics and oral candidosis and possible mechanisms of pathogenicity following such therapy.

Oral submucous fibrosis and oral yeast carriage - a case control study in Sri Lankan patients.

Oral submucous fibrosis (OSMF) is a well-known precancerous condition. Epithelial atrophy is one of the key features in OSMF. Presence of Candida in the mouth together with epithelial changes may predispose to candidal infection. Candidal infection together with other co-factors may also induce epithelial atypia and dysplasia leading to malignant change. The purpose of this study was to determine the prevalence of oral yeast carriage in patients with OSMF and to compare the carriage with the normal individuals. Thirty patients with histologically proven OSMF and healthy subjects were used as the test and control respectively. Oral rinse samples were collected from all the subjects and cultured on Sabouraud's agar. Species were identified using API 32C AUX identification kits. Nineteen (63.6%) of the test group and 15 (50%) of the control group had yeast isolated from their mouth. The carriage of yeast in the OSMF group was not statistically significant compared with the control group. We isolated C. dubliniensis in Sri Lanka for the first time and interestingly from the oral cavities of both OSMF patients and healthy individuals.

Diabetes mellitus as a contributory factor in oral candidosis.

It has been reported that poor glycaemic control predisposes to oral candidal infection in diabetic patients. For instance, the carriage of Candida species and the density of candidal growth in the oral cavity is frequently claimed to be increased in patients with diabetes mellitus. However, the validity of these observations remains controversial. Hence, we review and discuss here the clinical data in the literature on the relationship between diabetes and oral candidal carriage and infection, and possible mechanisms associated with its pathogenicity.

The impact of cigarette/tobacco smoking on oral candidosis: an overview.

Smoking is associated with a variety of changes in the oral cavity. Cigarette smoke has effects on saliva, oral commensal bacteria and fungi, mainly Candida, which causes oral candidosis, the most common opportunistic fungal infection in man. How cigarette smoke affects oral Candida is still controversial. This brief overview is an attempt to address the clinical findings on the relationship between smoking and oral candidosis and possible mechanisms of pathogenicity.

Rapid and unequivocal differentiation of Candida dubliniensis from other Candida species using species-specific DNA probes: comparison with phenotypic identification methods.

Candida dubliniensis is a recently described opportunistic pathogen which shares many phenotypic characteristics with Candida albicans but which has been reported to rapidly acquire resistance to azole antifungal drugs. Therefore, differentiation of C. dubliniensis from C. albicans becomes important to better understand the clinical significance and epidemiologic role of C. dubliniensis in candidiasis. We compared phenotypic methods for the differentiation of C. dubliniensis from C. albicans (i.e. the ability to grow at elevated temperatures, colony color on CHROMagar Candida medium, and carbohydrate assimilation patterns) to amplify the results of a polymerase chain reaction (PCR) assay using universal fungal primers to the internal transcribed spacer 2 (ITS2) region of rDNA and species-specific DNA probes in an enzyme immunoassay format (PCR-EIA). DNA sequencing of the ITS1 rDNA region was also conducted. The C. dubliniensis ITS2 probe correctly identified all C. dubliniensis isolates without cross-reaction with any other Candida species tested (mean A(650 nm) +/- SE, C. dubliniensis probe with C. dubliniensis DNA, 0.372 +/- 0.01, n = 22; C. dubliniensis probe with other Candida species DNA, 0.001 +/- 0.02 n = 16, P < 0.001). All other Candida species tested (C. albicans, Candida glabrata, Candida krusei, Candida parapsilosis, and Candida tropicalis) were also correctly identified by the PCR-EIA without any detectable cross-reactions among species. Phenotypically, C. dubliniensis isolates demonstrated an increased sensitivity to heat compared to C. albicans isolates. At 42 degrees C, only 50% of C. dubliniensis isolates grew compared to 73% of C. albicans isolates and, at 45 degrees C, 91% of C. dubliniensis isolates failed to grow compared to 64% of C. albicans isolates. C. albicans was more likely to demonstrate a dark green or blue green colony color on CHROMagar Candida medium obtained from Becton Dickinson (i.e. 100% of C. albicans isolates were dark green or blue green versus 64% of C. dubliniensis isolates) whereas no difference in the percentage of C. albicans or C. dubliniensis isolates producing dark green or blue green colony color was detected using CHROMagar Candida medium from Hardy Diagnostics (82% for both species). The API 20C AUX carbohydrate assimilation system incorrectly identified C. dubliniensis as C. albicans in all but three cases: remaining isolates were misidentified as C. albicans/C. tropicalis, C. tropicalis/C. albicans, and Candida lusitaniae/C. albicans. In all, 82% of C. albicans isolates and 100% of C. dubliniensis isolates assimilated trehalose; the latter finding was opposite to that reported for C. dubliniensis in the API 20C AUX profile index. Xylose and alpha-methyl-D-glucoside assimilation, respectively, were negative for 100 and 95% of C. dubliniensis isolates and positive for 100 and 91% of C. albicans isolates, confirming earlier reports that assimilation results for xylose and alpha-methyl-D-glucoside may be helpful in the discrimination of these two species. However, conventional phenotypic species identification tests required days for completion, whereas the PCR-EIA could be completed in a matter of hours. In addition, identification of Candida species by ITS1 rDNA sequencing gave 100% correspondence to the results obtained by the PCR-EIA, confirming the specificity of the PCR-EIA method. These data indicate that although a combination of phenotypic methods may help differentiate C. dubliniensis from C. albicans to some extent, the PCR-EIA can provide a simple, rapid, and unequivocal identification of the most medically important Candida species in a single test.

Adhesion and cell-surface-hydrophobicity of sequentially isolated genetic isotypes of Candida albicans in an HIV-infected Southern Chinese cohort.

Objectives of the study were to investigate the variability in yeast adhesion and cell-surface-hydrophobicity (CSH) during human immunodeficiency virus (HIV) disease progression, using a total of 60 sequential Candida albicans isolated from oral rinse samples of seven HIV-infected individuals with (4) and without (3) clinical symptoms of oropharyngeal candidosis. Significant differences in the adhesion to buccal epithelial cells (BECs) during sequential visits were observed for all genetic isotypes in five of the seven individuals and three isotypes belonging to the sixth individual. A single isotype of patient HK1 and another of HK4 (genotype I) demonstrated significant variations in their CSH during sequential visits whereas no such differences were noted for the remaining genotypes. On Spearman correlation analysis an isotype from HK1 demonstrated a significant increased adherence to BECs and CSH during HIV disease progression whereas no such correlation was noted for the remaining isotypes studied. No significant differences in adherence to BECs or CSH values were observed between the symptomatic oral candidosis and the asymptomatic carrier group. Further, on regression analysis only the single isotype of HK1 demonstrated a significant positive correlation between adherence to BECs and CSH whereas no such correlation was observed when all tested Candida isolates were pooled and evaluated as a single, large group.

Molecular heterogeneity of fluconazole-resistant and -susceptible oral Candida albicans isolates within a single geographic locale.

The emergence of drug-resistant Candida albicans in immunocompromised patients is common. A disconcerting aspect of this phenomenon is the rapid emergence of C. albicans strains that are resistant to a widely used azole drug, fluconazole (FLZ). To understand the origin of FLZ-resistant yeast isolates, we investigated molecular profiles of 20 geographically related oral C. albicans isolates using three genotyping methods: randomly amplified polymorphic DNA-PCR, with six different primers (OBU1, OBU2, OBU3 RSD6, RSD11 and RSD12); electrophoretic karyotyping by pulsed-field gel electrophoresis; and HinfI restriction fragment analysis. Of the 20 isolates studied, 10 were FLZ- resistant and originated from patients with oral candidosis with a history of FLZ therapy, and the remainder were FLZ susceptible from individuals with oral candidosis, but without a history of FLZ therapy. A composite genotype was generated for each strain by combining molecular types derived from the three independent molecular methods. The composite profiles indicated genetic diversity amongst both the FLZ-resistant as well as -sensitive isolates, and no specific features emerged distinguishing the drug-resistant and -sensitive groups. These observations cast doubt on the theory of a clonal origin of FLZ-resistant C. albicans isolates.

The impact of polyene, azole, and DNA analogue antimycotics on the cell surface hydrophobicity of Candida albicans and Candida tropicalis in HIV infection.

Oral candidiasis is the most common opportunistic infection in individuals infected with the human immunodeficiency virus. Though Candida albicans is the major aetiological agent, non-albicans species such Candida tropicalis are now emerging as important agents of such infection. The Candida cell surface hydrophobicity (CSH) is considered a critical factor contributing to its colonization potential and virulence. It is also known that brief exposure to sub-cidal concentrations of antifungal agents is a likely scenario in the oral environment where the administered drugs are diluted continuously due to the flushing action of saliva. Hence the objective of the present study was to compare the CSH of 10 isolates each of C. albicans and C. tropicalis from HIV-infected individuals following brief exposure (1 hour) of isolates to sub-therapeutic concentrations of nystatin, amphotericin B, ketoconazole, fluconazole and 5-flurocytosine. The CSH was assessed by a previously described biphasic aqueous-hydrocarbon assay. The mean percentage reduction of CSH of C. albicans following brief exposure to nystatin, amphotericin B, ketoconazole, fluconazole and 5-flurocytosine was 27.33 (p < 0.001), 21.34 (p < 0.05), 11.74 (p > 0.05), 18.4 (p > 0.05) and 14.64 (p > 0.05) respectively. The mean percentage reduction of CSH of C. tropicalis following brief exposure to nystatin, amphotericin B, ketoconazole, fluconazole and 5-flurocytosine was 33.81 (p < 0.01), 28.88 (p < 0.01), 12.6 (p > 0.05), 21.53 (p > 0.05) and 17.68 (p > 0.05) respectively. A significant interspecies variation in CSH was observed for nystatin and amphoterecin B. Overall the results reveal that the CSH of C. albicans is affected to a significantly lesser degree compared with C. tropicalis when exposed to the antifungals. These data further illustrate another mode of action of antifungals on Candida leading to a reduction in the CSH and thereby the yeast adherence to host tissues.

Post-antifungal effect of polyene, azole and DNA-analogue agents against oral Candida albicans and Candida tropicalis isolates in HIV disease.

Oropharyngeal candidiasis (OPC) is the most frequent AIDS-associated opportunistic infection, as up to 90% of HIV-infected individuals suffer at least one episode during the course of their disease. Various in vivo and in vitro procedures have been used to assess the effectiveness of antifungal agents used in HIV infection. In the present study, we evaluated in vitro the minimum inhibitory concentration (MIC) and the post-antifungal effect (PAFE) of two polyenes, two azoles and one DNA-analogue against 10 oral isolates of Candida albicans and 10 of Candida tropicalis, all from HIV-infected individuals, in order to obtain basic data on the pharmacodynamics of these drugs. One-hour exposure to twice the MIC of all the drugs, except fluconazole, elicited a consistently high PAFE in both Candida species. Furthermore, the PAFE elicited by the antifungals (except fluconazole) was significantly prolonged for C. tropicalis compared with C. albicans. This speedy recovery of C. albicans isolates exposed to transient low concentrations of antifungals appeared to reflect its virulence compared with lesser potent species, such as C. tropicalis. Taken together, the current data, while confirming the existence of PAFE in a non-albicans species of Candida, also provide further clues for the recalcitrance of C. albicans species in the face of antifungal therapy for oropharyngeal candidiasis.

Inhalational and topical steroids, and oral candidosis: a mini review.

Candidosis is by far the commonest oral fungal infection in man and could manifest as an adverse effect of drug therapy such as inhaled or topically applied corticosteroids. Due to the anti-inflammatory and immunosuppressive effect steroids are used in the management of bronchial asthma and oral mucosal diseases. In this mini review we discuss the clinical and laboratory findings on the relationship between steroid inhalers, other topical steroids and oral candidosis, possible mechanisms of pathogenicity following such therapy as well as the precautions that could be taken to minimize this adverse side effect.

Effects of two different growth media on the postantifungal effect induced by polyenes on Candida species.

There are no data on the effects of different growth media on polyene-induced postantifungal effect (PAFE) in Candida species. Hence, the nystatin- and amphotericin B-induced PAFEs in six Candida species (26 isolates) grown in Sabouraud's dextrose broth (SAB) and RPMI broth were evaluated, following limited exposure to the MICs of the two polyenes, using an automated turbidometric method. For nystatin, PAFE varied between 1.88 and 4.87 h in SAB and 0.66 and 6.89 h in RPMI, and for amphotericin B, the equivalent values were 3.13 to 10.98 h in SAB and 0.97 to 7.01 h in RPMI. These highly significant (P < 0.001) variations in the PAFE with both drugs, noted with most Candida strains grown in different media, call for standardization of intralaboratory methodology in measuring this parameter in order to obtain universally comparable data.

Adhesion of Candida parapsilosis to epithelial and acrylic surfaces correlates with cell surface hydrophobicity.

We investigated in vitro adherence of 24 isolates of Candida parapsilosis and 12 isolates of Candida albicans with regard to their relative cell-surface hydrophobicity (CSH), adherence to human buccal epithelial cells (BEC) and acrylic surfaces. There was no significant interspecies difference in the relative adherence of C. parapsilosis and C. albicans isolates to BEC, although the former demonstrated a tendency for increased adherence. However, a significant intra-species variation in adherence among isolates of C. parapsilosis (P < 0.0001) to BEC, but not of C. albicans was noted. The superficial isolates of C. parapsilosis demonstrated a higher avidity (33%) to BEC than the systemic isolates. On regression analysis a significant positive correlation between C. parapsilosis adherence to BEC and denture acrylic surfaces was noted (r = 0.45, P = 0.02). Similarly, buccal cell adherence correlated strongly with CSH of C. parapsilosis (r = 0.63, P = 0.0008). These results shed further light on the intimate relationship between adherence and CSH in candidal colonization and imply that both C. parapsilosis and C. albicans are equally potent in colonizing mucosal surfaces with respect to the attributes investigated.

Adjunctive use of chlorhexidine in oral candidoses: a review.

Oral candidosis is by far the commonest human fungal infection and manifests in a variety of clinical guises. The main reason for its high incidence appears to be the multiplicity of predisposing factors, which facilitate the conversion of oral commensal Candida to a parasitic existence. Despite the availability of a number of effective antimycotics for the treatment of oral candidoses, failure of therapy is not uncommon owing to the unique environment of the oral cavity where the flushing effect of saliva and the cleansing action of the oral musculature tend to reduce the drug concentration to sub-therapeutic levels. For these and other reasons chlorhexidine is widely prescribed in dentistry both as an antiseptic mouthwash and a denture disinfectant in order to supplement other antifungals. Chlorhexidine has a broad spectrum of antimicrobial activity including Candida albicans and other common non-albicans yeast species. In this review we outline the utility of chlorhexidine as an adjunct to conventional antimycotic therapy in the management of oral Candida infections.

The impact of chlorhexidine gluconate on the relative cell surface hydrophobicity of oral Candida albicans.

OBJECTIVES: Adherence of Candida albicans has been implicated as the first step in the pathogenesis of oral candidosis, and its relative cell surface hydrophobicity (CSH) a contributory physical force. Chlorhexidine gluconate is by far the commonest antiseptic mouthwash prescribed in dentistry. The intra-oral concentrations of the retained chlorhexidine mouthwash fluctuate considerably due to the dilution effect of saliva and the cleansing action of the oral musculature. Hence the objective of the present study was to investigate the effect of brief exposure to sub-therapeutic concentrations of chlorhexidine gluconate on the relative CSH of C. albicans. DESIGN: The CSH of the isolates was assessed by a biphasic aqueous-hydrocarbon assay. RESULTS: A statistically significant reduction in CSH was observed following the exposure of Candida isolates to 0.005 and 0.0025% chlorhexidine gluconate. CONCLUSIONS: These results elucidate additional mechanisms by which chlorhexidine gluconate suppress candidal pathogenicity despite a brief period of transient exposure within the oral environment.

Exposure to subtherapeutic concentrations of polyene antifungals suppresses the adherence of Candida species to denture acrylic.

BACKGROUND: The adherence of Candida species to denture acrylic is the initial event leading to Candida-associated denture stomatitis, with Candida albicans being the main aetiological agent. However, the increased incidence of immunocompromised patients in the community has resulted in the emergence of a number of non-albicans Candida species as causative agents of this disease, which is commonly managed by topically delivered polyene antifungals. Hence, we investigated the effect of the exposure of denture acrylic surfaces to nystatin and amphotericin B on the subsequent adhesion of six different Candida species. METHODS: Acrylic strips were exposed to subtherapeutic concentrations of the two polyenes for 30 min, and the adhesion of 4 isolates each of C. albicans, Candida glabrata, Candida guilliermondii, Candida krusei, Candida parapsilosis and Candida tropicalis was assessed using a previously described in vitro method with slight modifications. RESULTS: Overall, the results indicated a 35.9% (p < 0.01) and 63.1% (p < 0. 01) reduction, respectively, in yeast adhesion to denture acrylic following exposure to nystatin and amphotericin B, although this effect was not uniform for all the tested isolates. Thus, all C. glabrata, 3 C. guilliermondii and a single isolate each of C. krusei, C. parapsilosis and C. tropicalis were not significantly affected by nystatin exposure, and a single isolate each of C. glabrata and C. guilliermondii were not significantly affected by amphotericin B. CONCLUSIONS: The present data, the first on the effect of polyenes on a wide range of Candida species, indicate that the in vitro exposure of denture acrylic to subtherapeutic concentrations of nystatin and amphotericin B suppresses the adherence of pathogenic Candida species in general.

Sub-therapeutic exposure to polyene antimycotics elicits a post-antifungal effect (PAFE) and depresses the cell surface hydrophobicity of oral Candida albicans isolates.

Post-antifungal effect (PAFE) is defined as the suppression of growth that persists following limited exposure of fungi to antimycotics and subsequent removal of the drug. The fungal pathogen Candida albicans is the major aetiologic agent of oral candidosis, and the cell surface hydrophobicity (CSH) of this yeast is considered a critical factor contributing to its colonisation potential. As the concentration of topically prescribed antifungals reach sub-therapeutic levels at dosage intervals, the study of the polyene-induced PAFE and its impact on the CSH of oral C. albicans should be of clinical relevance. Hence the aims of this investigation were to measure the PAFE and CSH of 12 isolates of C. albicans following limited exposure (1 h) to nystatin and amphotericin B and also to investigate the ultrastructural features of yeast cells following such antifungal exposure. The yeasts were exposed to sub-lethal concentrations of nystatin (x2 MIC) and amphotericin B (x2 MIC) for a period of 1 h. Following subsequent removal of the drug, the PAFE and the CSH of the isolates were assessed by a turbidometric measurement of growth and a biphasic aqueous-hydrocarbon assay, respectively. The mean duration of PAFE of nystatin and amphotericin B were 5.99 (+/-0.49) h and 8.73 (+/-0.93) h, respectively, while the reduction in CSH following exposure to these drugs were 17.32% (P<0.05 for 83% of the isolates) and 14.26% (P<0.05 for 66% of the isolates), respectively. On scanning electron microscopy the exposed cells were seen to undergo collapse of the internal cell membrane, leaving an intact cell wall, while a proportion of cells were deflated. Some cells showed intense puckering of the cell wall, resulting in a mulberry appearance. Taken together, these data elucidate additional mechanisms by which polyene antimycotics may operate in vivo to suppress candidal pathogenicity.