Distribution and hydrolytic enzyme characteristics of Candida albicans strains isolated from diabetic patients and their non-diabetic consorts

Introduction:  The aim of this study was to investigate the oral colonization profile of Candida albicans strains isolated from diabetic patients and their non-diabetic consorts. In addition hydrolytic enzyme activity of these isolates was analysed.
Methods:  The genetic diversity of C. albicans oral isolates from 52 couples was established using isoenzyme marker and cluster analysis. Hydrolytic enzyme characteristics, namely secreted aspartyl proteinases (SAPs) and phospholipases (PLs) were also analysed.
Results:  Simultaneous colonization by C. albicans was observed in the consorts of 12 couples (23.1%). Patterns of monoclonal and polyclonal oral colonization by C. albicans strains were identified and the coexistence of identical or highly related strains was observed in both members of eight couples. The genetic diversity observed in the total yeast population revealed four large, genetically distinct groups (A to D) and the coexistence of strains in couples or consorts conjugally unrelated. SAP and PL activity was observed in the majority of C. albicans isolates without any association to particular strain, strain clusters (highly related isolates), or clinical characteristics of the consorts (diabetic, non-diabetic, and gender).
Conclusion:  Possible sources of transmission and oral propagation of groups (clusters) of strains of C. albicans can occur between diabetic and non-diabetic consorts. A conjugal genotypic identity exists in most C. albicans -positive couples, that is, both consorts share identical or highly related strains; however, this identity is not couple-specific as seen by the coexistence of clusters in couples and unrelated consorts.     

Clonal identity of Candida albicans in the oral cavity and the gastrointestinal tract of pre‐school children

The clonal relationship between oral and fecal Candida albicans isolated from children of pre‐school age was examined using RAPD analysis. Significantly higher levels of C. albicans were found in saliva, dental plaque, carious specimens and stools of 56 patients with severe caries as compared to 52 healthy control subjects. The highest prevalence was found in carious specimens and a strong correlation was observed between its presence in saliva, dental plaque, carious specimen and feces. RAPD analysis of isolates from 23 patients with simultaneous oral and fecal C. albicans revealed clonal counterparts present in both oral and stool samples in 15 cases; five patients harbored closely related strains; and three patients harbored unrelated strains. Our results demonstrate a strong correlation between oral and gastrointestinal C. albicans colonization. We assume that carious teeth may constitute an ecologic niche for C. albicans potentially responsible for recurrent oral and non‐oral candidiasis.     

Analysis of the strain relatedness of oral Candida albicans in patients with diabetes mellitus using polymerase chain reaction‐fingerprinting

To increase our understanding of Candida pathogenicity, the identification of those strains most frequently associated with infections is of paramount importance. Polymerase chain reaction (PCR)‐based methods are extremely effective in differentiating and determining reproducibility, they require minimum starting material and are rapid and simple to perform. In this study, the genetic relatedness of Candida albicans was assessed for two geographically different patient groups (London, UK and Parma, Italy) affected by diabetes mellitus. C. albicans samples from the oral cavities of non‐diabetic healthy subjects were also examined by PCR fingerprinting to evaluate the possible genetic differences among endogenous strains in individuals with and without diabetes mellitus. PCR fingerprinting, with subsequent phylogenetic analysis of C. albicans isolates from the diabetic patients from London and Italy and from the non‐diabetic subjects, revealed that there were significant differences (P < 0.0001) between C. albicans isolates indicative of the distinct ecological niches that occur in the oral cavities of these patient cohorts. The most diverse group comprised the isolates from the diabetic patients in the UK, possibly reflecting the antifungal treatment that these patients had received. Further studies that include isolates from patient cohorts with systemic diseases other than diabetes mellitus, and from more diverse geographic localities are required to explain the relatedness of C. albicans isolates in the mouth.     

Epidemiology of oral yeast colonization and infection in patients with hematological malignancies,head neck and solid tumors

J Oral Pathol Med (2011) 40 : 83–89 Background: The aim of this study was to investigate the epidemiology of oral yeast colonization and infection amongst cancer patients. Methods: Patients with solid tumor, head‐neck cancer or hematological malignancy were recruited into the study. Demographic data on age, gender, type of cancer, preceding treatment with antibiotics, anti‐fungal agents, chemotherapy, radiation or surgery and presence of dentures were recorded on admission. Oral examination and microbial swabs were obtained and yeast culture, identification and antifungal susceptibility performed. Results: Oral yeast colonization was prevalent in 56.8% (227/400) of all cancer patients and 18.9% (43/227) of those had clinical and microbiological evidence of infection. The incidence of oral candidiasis in yeast colonized patients was highest in head neck cancer (29.2%) followed by hematological malignancies (20.5%) and solid tumor (17%) patients. Age and dentures were identified as independent risk factors associated with yeast carriage. Candida albicans was the dominant (74%) species (497.5 per 1000 cancer admissions) followed by C. glabrata (11.5%), C. tropicalis (2.6%), C. krusei (2.6%) and C. parapsilosis (1.9%). The overall resistance to azoles was 28.2% (75/266). Resistance to specific drugs was seen for fluconazole (4.5%), itraconazole (11.7%), ketoconazole (11.3%), voriconazole (0.75%) and caspofungin (41.1%) but none to amphotericin B or nystatin. Conclusions: The highest incidence of oral candidiasis amongst cancer patients was seen in head neck cancers. The majority of infections were caused by C. albicans but almost one third of patients harbored non‐C. albicans strains such as C. glabrata which were often more resistant to anti‐fungal agents.     

Correlation between enzyme production,germ tube formation and susceptibility to fluconazole in Candida species isolated from patients with denture‐related stomatitis and control individuals

Introduction: Phospholipase and proteinase secretion in yeasts of the genus Candida has been described as a relevant virulence factor. Also, germ tube formation by Candida albicans is associated with its invasive capacity and is considered an important pathogenic mechanism. Methods: To link the production of hydrolytic enzymes with the capacity to produce infection, 232 clinical isolates of yeasts from the oral cavity of 140 individuals wearing removable maxillary protheses were studied. The sample was composed of 70 patients with denture‐related stomatitis (DRS) and 70 individuals with normal palatal mucosa. For strains identified as C. albicans, the correlation between germ tube formation and their capacity to cause infection was studied and the presence of Candida dubliniensis was investigated. Susceptibility to fluconazole was evaluated. Results: Candida albicans was the only species producing phospholipase and germ tube. We observed a higher level of production of phospholipase in cases of infection compared with commensals. Significant differences between the two groups of C. albicans isolates were observed as to germ tube production. Only, Candida glabrata showed lower susceptibility to fluconazole. Conclusion: The results reinforced the idea that C. albicans is the most frequent and can be the most pathogenic yeast in oral candidosis. However, the strains isolated from DRS patients and healthy individuals showed the same virulence factors. It seems that several virulence attributes are involved in the infective process but no single factor contributes to Candida virulence. Candida dubliniensis was absent in the oral cavity of individuals with and without DRS.     

Colonization of Candida: prevalence among tongue‐pierced and non‐pierced immunocompetent adults

Oral Diseases (2010) 16 , 176–175 Objective: To evaluate the colonization of Candida at the tongue‐piercing site of immunocompetent individuals. Subjects and methods: Swabs samples were obtained from the anterior lingual mucosa of healthy young adults with tongue piercing (N = 115); 86 subjects with (non‐intra‐oral) facial piercing served as a comparison group. Candida colonization was examined by light microscopy after 5‐day incubation. Positive specimens were re‐cultured on Chromagar?Candida plates for species identifying. Results: Candida colonization was more prevalent among tongue‐pierced (20.0%) than facial‐pierced subjects (9.4%; P = 0.048). All colonies were of Candida albicans. No difference was found between current tongue ornament wearers (21.2%) and non‐wearers (19.5%; P = 0.803). In multivariate analysis, the only significantly positive influencing factors on colonization were tongue piercing (P = 0.034) and daily smoking of more than 10 cigarettes (P = 0.024). Conclusions: Piercing of the tongue was found to be a risk factor for colonization of Candida albicans, without an influence of whether or not an ornament is in place.     

Identification of Candida dubliniensis among oral yeast isolates from an Italian population of human immunodeficiency virus‐infected (HIV+) subjects

Candida dubliniensis, an emerging oral pathogen, phenotypically resembles Candida albicans so closely that it is easily misidentified as such. The aim of the present study was to evaluate the usefulness of two phenotypic methods, growth at 45°C and 2,3,5‐triphenyltetrazolium chloride (TTC) reduction, for confirming presumptive identification of C. dubliniensis and C. albicans by colony color on CHROMagar Candida (CAC) medium. A combination of these methods was used to establish the prevalence of oral C. dubliniensis in an Italian population of 45 human immunodeficiency virus (HIV)‐infected subjects. Twenty‐two samples (48.9%) were positive for yeasts on CAC medium producing a total of 37 fungal isolates. The colony color and 45°C growth ability test correctly identified all C. dubliniensis and C. albicans isolates (5/37, 13.5%, and 16/37, 43.2%, respectively), while assessment of TTC reduction misidentified one C. albicans isolate. The isolation rate of C. dubliniensis was 11.1% (5/45 patients). All of the C. dubliniensis isolates were highly susceptible to fluconazole (MIC = 0.5 µg/ml). The combination of CAC medium screening with growth at 45°C and TTC reduction tests may represent a simple, reliable and inexpensive identification protocol for C. dubliniensis.     

Oral epithelium–Candida glabrata interactions in vitro

Background: Oropharyngeal candidiasis is a common opportunistic infection and Candida glabrata is the second or third most frequently isolated species from oropharyngeal candidiasis lesions, after Candida albicans. The aim of this study was to study the cytokine‐inducing and cell‐damaging potential of C. glabrata in oral epithelial cells and compare this to C. albicans. Methods: Oral epithelial cell lines and primary gingival epithelial cells were cocultured with C. glabrata strains GDH2269 and 94‐11 or C. albicans strains SC5314 and ATCC28366. Supernatants were analysed for the presence of interleukin‐1α (IL‐1α), IL‐8 and granulocyte–macrophage colony‐stimulating factor (GM‐CSF) by enzyme‐linked immunosorbent assay. The cytotoxity of different strains was determined using the CytoTox‐96 assay. Results: Compared to C. albicans, C. glabrata induced different proinflammatory cytokine responses in oral epithelial cells; a high level of GM‐CSF induction was only detected in C. glabrata‐infected cells and not in C. albicans‐infected cells, regardless of the origin of these cells (cell lines or primary cells) or the strain used. Like C. albicans, C. glabrata induced an IL‐1α response by oral epithelial cells, but this response was both strain‐dependent and epithelial cell origin‐dependent. Unlike C. albicans, C. glabrata failed to induce a strong IL‐8 response in any of the cell systems studied. Finally, in these studies C. glabrata showed lower cytotoxicity than C. albicans. Conclusions: C. glabrata is less cytotoxic than C. albicans and induces different proinflammatory cytokine responses in oral epithelial cells.     

Genetic diversity and exoenzyme activities of Candida albicans and Candida dubliniensis isolated from the oral cavity of Brazilian periodontal patients

Objective

Mucosal surfaces are the primary oral reservoirs of Candida species, but these species can also be found in subgingival biofilm. The present study investigated the genetic diversity and production of exoenzymes of C. albicans and C. dubliniensis isolated from the oral cavity of systemically healthy patients with periodontitis.

Design

Fifty-three patients were analysed. Samples were collected from three oral cavity sites (periodontal pocket, gingival sulci and oral mucosa), plated and, after isolation, suspect strains of C. albicans and C. dubliniensis were identified by PCR. The genetic diversity of the isolates was evaluated by RAPD and the activities of the secreted aspartyl proteinases and phospholipases were evaluated by the agar plate method.

Results

Twenty-one patients showed positive results for Candida spp. There were no statistically significant differences between genders, or between sites. C. albicans was the most frequently found specie, while C. dubliniensis was isolated from the periodontal pocket of only one patient. Sixteen genotypes were detected among the C. albicans isolates, and one among the C. dubliniensis isolates. The similarity coefficient (S_SM) values among the C. albicans genotypes ranged from 0.684 to 1.0 with an average of 0.905 ± 0.074. All isolates produced high levels of Saps and most of them produced high levels of phospholipases. No relationship was found between the genotypes and the pattern of enzymatic production. There was no association between specific genotypes and their site of isolation.

Conclusions

The results of the present study suggest that genetically homogeneous strains of C. albicans are present in the oral cavity of patients with periodontitis and that these strains are capable of producing high levels of exoenzyme.     

Oral Candida colonization and its relation with predisposing factors in HIV‐infected children and their uninfected siblings in Brazil: the era of highly active antiretroviral therapy

J Oral Pathol Med (2010) 39 188–194 Objectives: To evaluate predisposing factors such as orofacial manifestations, immunosuppression status and antiretroviral therapy in relation to oral colonization by Candida spp. in Brazilian HIV‐infected children and their uninfected siblings in the era of highly active antiretroviral therapy (HAART). Methods: Whole stimulated saliva was collected from 65 HIV‐infected children (HIV+) and 40 uninfected siblings (HIV–), followed by assessment of orofacial manifestation, caries indexes and the number of cavitated dentinal carious teeth (CDT). The salivary samples were cultured and the colonies were counted. After which they were identified by sugar assimilation and fermentation (API 20C). Data was analyzed using chi‐square, Mann–Whitney, Spearman tests and logistic regression. Results: Regarding positive growth, HIV+ presented 80% (52/65) and HIV? 57.5% (23/40) (P = 0.013). Absence of antiretroviral therapy and HAART increased the probability of Candida isolation (P < 0.05). Mean CD4%, immune‐status and history of recurrent oral candidiasis (OC) had no influence on Candida isolation. Mixed Candida spp. cultures were observed in HIV+ (40%) and HIV? (52%): C. albicans was more frequently found in both groups, with a higher prevalence in HIV+ (P = 0.05); other non‐albicans species were isolated in HIV+ and HIV?. Low prevalence of orofacial manifestations was observed in HIV+ (10.7% of OC). There was an association between means of CDT and Candida growth (P < 0.05) and a positive correlation between number of CDT and Candida cfu‐counts in HIV+ and HIV?. Mean CD4% and immune‐status had no influence on Candida isolation. Absence of antiretroviral therapy and HAART increased the probability of Candida isolation (P < 0.05). Conclusions: The HIV infected children had a significantly higher prevalence of oral Candida spp. compared to their uninfected siblings. Absence of HAART and presence of dentinal carious teeth increased significantly Candida spp. colonization in these children.     

Adherence of Candida albicans to silicone is promoted by the human salivary protein SPLUNC2/PSP/BPIFA2

Interactions between Candida albicans, saliva and saliva‐coated oral surfaces are initial events in the colonization of the oral cavity by this commensal yeast, which can cause oral diseases such as candidiasis and denture stomatitis. Candida albicans also colonizes silicone voice prostheses, and the microbial biofilm formed can impair valve function, necessitating frequent prosthesis replacement. We have previously shown that saliva promoted binding of C. albicans cells to silicone in vitro, and that the selective binding of specific salivary proteins to voice prosthesis silicone mediated attachment of C. albicans cells. The C. albicans cells adhered to a polypeptide (or polypeptides) of ~36 kDa eluted from saliva‐treated silicone. We show here that a protein of similar size was identified in replicate blots of the eluate from saliva‐treated silicone when the blots were probed with antibodies to human SPLUNC2, a salivary protein with reported microbial agglutination properties. In addition, SPLUNC2 was depleted from saliva that had been incubated with silicone coupons. To determine whether SPLUNC2 is a yeast‐binding protein, SPLUNC2 cDNA was expressed in Escherichia coli. Purified recombinant His‐tagged protein (SPLUNC2r) bound to silicone as demonstrated by immunoblot analysis of an eluate from SPLUNC2r‐treated silicone coupons and ³⁵S‐radiolabelled C. albicans cells adhered in a dose‐dependent manner to SPLUNC2r‐coated silicone. We conclude that SPLUNC2 binds to silicone and acts as a receptor for C. albicans adherence to, and subsequent colonization of, voice prosthesis silicone.     

Reducing the Incidence of Denture Stomatitis: Are Denture Cleansers Sufficient?

Purpose: Candida albicans is the predominant oral yeast associated with denture stomatitis. With an increasing population of denture wearers, the incidence of denture stomatitis is increasing. Effective management of these patients will alleviate the morbidity associated with this disease. The aim of this study was to examine the capacity of four denture cleansers to efficiently decontaminate and sterilize surfaces covered by C. albicans biofilms. Materials and Methods: Sixteen C. albicans strains isolated from denture stomatitis patients and strain ATCC 90028 were grown as mature confluent biofilms on a 96‐well format and immersed in Dentural, Medical? Interporous^®, Steradent Active Plus, and Boots Smile denture cleansers according to the manufacturers’ instructions or overnight. The metabolic activity and biomass of the biofilms were then quantified, and scanning electron microscopy (SEM) used to examine treated biofilms. Results: Dentural was the most effective denture cleanser, reducing the biomass by greater than 90% after 20 minutes. Steradent Active plus was significantly more effective following 10‐minute immersion than overnight (p < 0.001). All cleansers reduced the metabolic activity by greater than 80% following overnight immersion; however, Boots Smile exhibited significantly reduced metabolic activity following only a 15‐minute immersion (p < 0.001). SEM revealed residual C. albicans material following Dentural treatment. Conclusions: This study showed that denture cleansers exhibit effective anti‐C. albicans biofilm activity, both in terms of removal and disinfection; however, residual biofilm retention that could lead to regrowth and denture colonization was observed. Therefore, alternative mechanical disruptive methods are required to enhance biofilm removal.     

The development and validation of a rapid genetic method for species identification and genotyping of medically important fungal pathogens using high‐resolution melting curve analysis

Accurate, rapid and economical fungal species identification has been a major aim in mycology. In this study, our goal was to examine the feasibility of a high‐resolution melting curve analysis (HRMA) of internal transcribed regions ITS1 and ITS2 in ribosomal DNA (rDNA) for a rapid, simple and inexpensive differentiation of eight clinically relevant Candida species (Candida albicans, Candida glabrata, Candida parapsilosis, Candida krusei, Candida tropicalis, Candida guilliermondii, Candida dubliniensis and Candida lusitaniae). In addition, for the first time, we tested the applicability of HRMA to classify C. albicans strains into four previously described genotypes (A, B, C and D) using a primer set that spans the transposable intron region of 25S of rDNA. Type and unknown clinical oral isolates were used in this study and the melting curve analysis was compared with both amplicons' sequencing and agarose gel electrophoresis analysis. Real‐time PCR and subsequent HRMA of the two described rDNA regions generated distinct melting curve profiles that were in accord with sequencing and gel electrophoresis analysis, highly reproducible, and characteristic of each of the eight Candida species and C. albicans genotypes. Moreover, results were obtained in 4 h and without the need for any post‐amplification handling, so reducing time and cost. Owing to its simplicity and speed, this technique is a good fit for genotypic analysis of hundreds of clinical strains in large epidemiological settings.     

The isolation,identification and molecular analysis of Candida spp. isolated from the oral cavities of patients with diabetes mellitus

Previous studies have shown a high incidence (77%) of isolation of Candida spp. from the oral cavities of patients with type 1 diabetes mellitus. The aim of the present study was to assess the prevalence of yeast in the oral cavities of patients suffering from type 1 and type 2 diabetes mellitus. The patients were classified according to the level of diabetic control (HbA1_c), and further stratified on the presence or absence of dental prosthesis. Oral rinse samples were assessed for the growth of yeast and the degree of colonization. Oral isolates were defined to the species level by both phenotypic and novel molecular methods. The overall proportion (60%) of diabetic patients who had Candida spp. isolated from the oral cavity was similar to that previously reported. Local oral factors, such as the presence of dentures, seemed to have a greater influence than diabetic status on the amount and species of Candida isolated from the oral cavities of diabetic patients. Diabetic patients with dentures had more non‐albicans Candida isolated from their mouths than dentate diabetic patients. Candida dubliniensis was isolated from diabetic patients and may have a predilection for dentate patients.     

Stable azole drug resistance associated with a substrain of Candida albicans from an HIV-infected patient

Oral candidiasis is one of the earliest and most frequent complications of a failing immune system in HIV-infected individuals. For several years, oral candidiasis has been treated effectively with azole drugs, the one most frequently used is fluconazole. Unfortunately, extensive use of the drug for treatment and prophylaxis has led to treatment failure in an increasing number of patients. In most of these cases, strains of C. albicans isolated from the infection are less susceptible to fluconazole. The development of azole resistance in strains of C. albicans has been studied biochemically and more recently with molecular techniques. One excellent example of the development of azole resistance in C. albicans has been documented in a series of 17 C. albicans isolates from a single patient over a 2-year period. During this time, the patient experienced 14 episodes of oral candidiasis and was treated with increasing doses of fluconazole. Molecular and biochemical analyses confirms that the isolates are the same strain of C. albicans and that the resistance in these isolates is stable over 600 generations, suggesting that the changes in this strain are genetic in nature. In addition, the development of resistance is correlated with the identification of a substrain or variant of the original strain, as identified by restriction fragment length polymorphism (RFLP) analysis with the moderately repetitive probe, Ca3. The analysis of this series of isolates demonstrates that azole drug resistance is associated with several small genetic changes, each of which contributes to the overall resistance of the strain. Clearly, continual use of azole drugs by a patient can select for genetic changes that render oral candidiasis refractory to treatment.     

Chitinase expression in parotid glands of non-obese diabetic mice

Oral Diseases (2012) 18 , 506–512 Objective: This investigation was a basal study that used a mouse model of xerostomia to identify protein biomarkers of xerostomia in saliva. We identified genes expressed differently in parotid glands from non‐obese diabetic mice with diabetes and those from control mice; subsequently, we investigated expression of the proteins encoded by these genes in parotid glands and saliva. Materials and Methods: DNA microarray and real‐time PCR analyses were performed to detect differences between NOD/ShiJcl and C57BL/6JJcl (control) female mice in gene expression from parotid glands or parotid acinar cells. Subsequently, protein expression was assessed using immunoblotting and immunohistochemistry. Similarly, enzyme activity in saliva was assessed using zymography. Results: Based on gene expression analyses, Chia expression was higher in diabetic mice than non‐diabetic mice and control mice; similarly, expression of chitinase, the protein encoded by Chia, was higher in diabetic mice. Saliva from NOD/ShiJcl mice had more chitinase than saliva from control mice. Conclusions: Chitinase was highly expressed in parotid acinar cells from diabetic mice compared with non‐diabetic and control mice. Increased chitinase expression and enzyme activity may characterize the autoimmune diabetes in mice; however, further investigation is required to assess its use as a biomarker of xerostomia in humans.     

Antigen I/II mediates interactions between Streptococcus mutans and Candida albicans

Streptococcus mutans and Candida albicans are frequently co‐isolated from dental plaque of children with early childhood caries (ECC) and are only rarely found in children without ECC, suggesting that these species interact in a manner that contributes to the pathogenesis of ECC. Previous studies have demonstrated that glucans produced by S. mutans are crucial for promoting the formation of biofilm and cariogenicity with C. albicans; however, it is unclear how non‐glucan S. mutans biofilm factors contribute to increased biofilm formation in the presence of C. albicans. In this study we examined the role of S. mutans antigen I/II in two‐species biofilms with C. albicans, and determined that antigen I/II is important for the incorporation of C. albicans into the two‐species biofilm and is also required for increased acid production. The interaction is independent of the proteins Als1 and Als3, which are known streptococcal receptors of C. albicans. Moreover, antigen I/II is required for the colonization of both S. mutans and C. albicans during co‐infection of Drosophila melanogaster in vivo. Taken together, these results demonstrate that antigen I/II mediates the increase of C. albicans numbers and acid production in the two‐species biofilm, representing new activities associated with this known S. mutans adhesin.     

Mutacin production in Streptococcus mutans genotypes isolated from caries‐affected and caries‐free individuals

Relationships between genetic diversity and mutacin production in Streptococcus mutans were evaluated in 319 clinical isolates from eight caries‐affected and eight caries‐free individuals. The isolates were submitted to mutacin typing and AP‐PCR (arbitrarily primed polymerase chain reaction) assay. The mutacin production was detected for 12 Streptococcus sp. indicator strains. Results showed significant variations in the mutacin production profiles and the inhibitory spectra of both groups. A possible association was seen between mutacin activity and the distinct patterns of Streptococcus sp. colonization in the two groups. Genotyping by AP‐PCR using the primers OPA‐02 and OPA‐13 revealed 101 distinct genotypes against 48 phenotypes identified by mutacin typing. No correlation was observed between the inhibitory spectra of mutacin and genotypic similarities based on AP‐PCR analyses. According to our results, strains of the same S. mutans genotype showed different mutacin profiles, suggesting a high degree of interstrain diversity. In conclusion, mutacin production seems to be of clinical importance in the colonization of S. mutans and is highly diversified in the S. mutans species.     

Antifungal drug susceptibility of oral Candida albicans isolates may be associated with apoptotic responses to Amphotericin B

J Oral Pathol Med (2010) 39 182–187 Background: Candida albicans is the important opportunistic fungal pathogens which can cause oral Candidiasis and even more seriously systemic infection. Apoptosis of C. albicans induced by environmental factor such as weak acid and antifungal drugs were studied recently. Illustrating the phenomenon of apoptosis in C. albicans may help us to discover new antifungal therapy by activating the fungal cells to suicide. Methods: Two oral C. albians clinical isolates which isolated respectively from healthy host [Strain 23C: minimal inhibition concentration (MIC) is 0.125 μg/ml for Amphotericin B (AmB)] and advanced cancer patient (Strain 28A: MIC is 2 μg/ml for AmB), were induced by 1 μg/ml AmB in vitro for 200 min, and then studied the apoptosis markers using terminal deoxynucletidyltransferase‐mediated dUTP nick end labeling (TUNEL) (shown by diaminobenzidine and fluorescent isothiocyanate), and the ultrastructure of cell nuclear using transmission electron microscope (TEM), quantitative analysis using flow cytometry for the rapid exposure of phosphatidylserine at the outer membrane and propodium iodide (PI) double staining. C. albicans conference strain YEM30 was used as the control strain. Results: With TUNEL assay and TEM, we detected the typical characteristics of apoptosis. Strain 23C (with low MIC) showed significantly higher percentage of apoptosis (19.92%) compared with Strain 28A (with high MIC) which was isolated from the cancer patient (7.29%) (P < 0.01). In addition, 7.3% of early apoptosis cells of Strain 23C can form colonies on the plates, while 15% for Strain 28A. None of the PI (+) cells can form colony. Conclusions: Apoptosis of oral C. albicans isolates can be induced by AmB. The feature of antifungal drug susceptibility of the oral C. albicans clinical isolates may associate with the response of apoptosis inducing.     

Human oral keratinocyte E‐cadherin degradation by Candida albicans and Candida glabrata

J Oral Pathol Med (2010) 39 : 275–278 Background: E‐cadherin (E‐Cad) is a 120‐kDa adhesive protein found in adherens junctions of the digestive tract epithelium. We tested the ability of two Candida strains to degrade human E‐Cad in the Candida virulence factor perspective. Materials and methods: We set out to study oral mucosal E‐Cad degradation by clinical and reference strains of Candida albicans and Candida glabrata. We also included hyphal and secreted aspartic proteinase (Sap) mutants of C. albicans to test the effect of yeast/hyphal transition on the ability to degrade E‐Cad. The tests were performed at pH 4 and pH 6 to determine the effect of local tissue acidity on the activation of Saps. The C. albicans strains used were: CCUG 32723; clinical strain SC5314 which is known to be strongly invasive; hyphal mutants of SC5314: HLC52 (efg1/efg1), HLC54 (cph1/cph1 efg1/efg1) and JKC19 (cph1/cph1); clinical strain B1134; Sap 1–3 and Sap 4–6 mutants of SC5314. The C. glabrata strains used were ATCC 90030, and the clinical strains 5WT and G212. Results: The sonicated yeast cells of C. albicans JKC19 and SC5314, both in hyphal form, degraded E‐Cad at pH 4. The 10× concentrated growth media of the strains HLC‐52, HLC‐54, 32723 and B1134; all in yeast form, caused degradation at pH 4, HLC‐52 and HLC‐54 also at pH 6. The C. glabrata strains did not degrade E‐Cad. Conclusions: pH is a strain dependent triggering factor in activating yeast or hyphal form related Candida Saps in degrading epithelial cell associated E‐Cads.