The role of yeasts in oral cancer by means of endogenous nitrosation

Oral leukoplakias, particularly non-homogeneous types, are often invaded by yeasts, with Candida albicans being the dominant species. The more advanced precancerous leukoplakia lesions yield more rarely occurring biotypes of C. albicans, suggesting a causal role for these biotypes in the malignant transformation. N-nitroso-benzylmethylamine (NBMA) is a compound able to induce carcinoma of the esophagus and the oral cavity in the rat. The catalytic potential of yeasts, isolated from leukoplakia lesions and from normal mucosa, to produce NBMA from the precursors N-benzyl-methylamine and nitrite was assessed at pH 6.8. The yeast strains differed in nitrosation potential, ranking from 0 to 1.2 micrograms NBMA/10(6) cells. C. albicans strains of the more rarely occurring biotypes showed the highest nitrosation potential, whereas C. tropicalis, C. parapsilosis, and Torulopsis glabrata were ranked lower. Strains with high nitrosation potential were generally isolated from lesions with more advanced precancerous changes. Thus, further evidence is provided supporting the hypothesis that yeasts play a causal role in oral cancer by means of endogenous nitrosamine production.     

Yeast species and biotypes associated with oral leukoplakia and lichen planus

Of 36 patients, 17 had oral leukoplakia, including homogeneous and nonhomogeneous types, and 19 had reticular lesions of oral lichen planus. A sample of yeast flora in each patient was taken from the pathologic lesion as well as from normal-appearing mucosa. The isolated yeasts were identified according to species level, and identification was extended beyond the species level for one species, Candida albicans, to reveal the biotype by means of the Odds and Abbott procedure comprising tests for acid and salt tolerance, proteinase production, resistance to 5-fluorocytosine and safranine, and assimilation of urea, sorbose, and citrate. Yeasts were present in the lesions of 82% of leukoplakia patients, compared to 37% of lichen planus patients, a frequency of yeasts corresponding to that in healthy adults. C. albicans was the dominating species in lesions of both diseases, constituting 82% of all yeasts in the leukoplakia lesions. In addition, the following species were identified: Candida tropicalis, Candida pintolopesii, Torulopsis glabrata, and Saccharomyces cerevisiae. Eighteen biotypes of C. albicans were encountered, the most frequently occurring biotypes being 355 and 177. Differences between C. albicans biotypes isolated from pathologic and normal mucosa were encountered in five of eleven leukoplakia patients and in one of three lichen planus patients. This indicates that the oral cavity comprises several ecologic niches for yeasts. As nonhomogeneous leukoplakias are more likely to develop into carcinoma than are homogeneous leukoplakias, it is interesting to note that the C. albicans biotypes isolated from nodular lesions (one type of nonhomogeneous leukoplakia)--biotypes 145, 175, and 575--rarely occur.(ABSTRACT TRUNCATED AT 250 WORDS)     

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.     

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.     

Candida infection in oral leukoplakia: an unperceived public health problem

Objectives: The study aimed to determine the proportion, known risk factors and etiology for Candida infection in leukoplakia lesions among patients with oral leukoplakia attending the Oral and Maxillofacial Clinic at a Tertiary Care Hospital in Sri Lanka.

Materials and methods: Eighty clinically suspected oral leukoplakia patients were included. Two oral swabs each, from leukoplakia patients: one swab from the lesion and the other one from the contralateral unaffected corresponding area (as a control) were collected. Direct microscopy and culture followed by colony count and phenotypic identification were performed to identify pathogenic Candida species.

Results: Candida infection was seen in 47% of patients with oral leukoplakia. Candida albicans (94.7%) was the most common Candida species followed by Candida tropicalis (5.3%). Majority of Candida-infected lesions were seen in the buccal mucosa region. Alteration of taste (p?=?0.021), having other oral lesions (p?=?0.008), angular cheilitis (p?=?0.024) and periodontitis (p?=?0.041) showed a significant association with Candida-associated leukoplakia. Increasing age showed a significant tendency for Candida infection (p?=?0.020). Smoking (p?=?0.026) and betel-quid chewing (p?=?0.006) were also found to be significantly associated, although alcohol consumption alone did not show a significant association. Oral leukoplakia patients who had all three habits: alcohol consumption, smoking and betel-quid chewing had a significant association with Candida infection (p?=?0.004).

Conclusions: Patients who had a combination of risk factors: smoking, betel-quid chewing and alcohol consumption were seen to have a significant association with Candida infection. Further betel-quid chewing alone and smoking singly was also significantly associated with Candida infection in oral leukoplakia.     

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.     

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.     

Podoplanin expression in oral leukoplakia: prognostic value and clinicopathological implications

Oral Diseases (2012) 18 , 692–699 Objectives: Current clinicopathological parameters cannot predict the risk of malignant transformation in oral leukoplakia sufficiently. Recent studies have shown that podoplanin is expressed in oral cancer and precancerous lesions. The aim of our study was to assess whether podoplanin expression in pretreatment biopsies could serve as a biomarker to predict the risk of malignant transformation in patients with oral leukoplakia. Materials and Methods: In this retrospective study, podoplanin expression was analysed in 60 patients with previously untreated oral leukoplakia by immunohistochemistry. We investigated the associations between podoplanin expression and various clinicopathological variables including oral cancer‐free survival (OCFS) and the SIN‐classification. Results: The chi‐square‐test revealed that high expression of podoplanin in pretreatment biopsies was associated with malignant transformation (P = 0.003) and increasing SIN‐classification (P = 0.009). In univariate analysis, podoplanin expression in oral leukoplakia had a significant impact on OCFS (P = 0.009). The 5‐year OCFS rate decreased from 100% for patients with no podoplanin expression to 41.7% for patients with the highest level of podoplanin expression. Conclusion: Although podoplanin expression and the SIN‐classification served as factors to predict malignant transformation in patients with oral leukoplakia in univariate analysis, no significant impact was found for both factors in multivariate analysis.     

Oral precancerous disorders associated with areca quid chewing, smoking, and alcohol drinking in southern Taiwan

OBJECTIVE: To investigate the prevalence and the associated risk factors of oral precancerous disorders in southern Taiwan. METHODS: We conducted a cross-sectional community survey interviewing 1075 adult subjects, 15 years of age and over, gathered from randomly selected 591 households, and spanning five villages in southern Taiwan. The study protocol included a visual oral soft tissue examination and a questionnaire-based interview. The chi-square test was used to test the differences in prevalence of oral precancerous lesions and conditions by different "life styles" relating to current risk habits of current areca quid chewing, smoking, and alcohol drinking. To control for possible confounding, a logistic regression model was used to estimate the Odds Ratios (OR) for leukoplakia and oral submucous fibrosis (OSF). RESULTS: 136 precancerous lesions and conditions were detected among 1075 subjects (12.7%). The analysis of the spectrum of oral precancerous disorders detected, leukoplakia (n = 80), OSF (n = 17) and verrucous lesions (n = 9), demonstrated an association with gender (P < 0.001). There were statistically significant associations among leukoplakia (P < 0.01), OSF (P < 0.0001), and verrucous lesions (P < 0.0001) and the life style of current areca quid chewing, smoking, and alcohol drinking. The synergistic effect of smoking and areca quid chewing habit on leukoplakia and OSF was demonstrated. CONCLUSION: This study reinforces the association of current areca quid chewing without tobacco, cigarette smoking, and alcohol drinking to leukoplakia, OSF, and verrucous lesions in Taiwan.     

Fluconazole-resistant Candida species in the oral flora of fluconazole-exposed HIV-positive patients

The purpose of this study was to examine the effect of preceding fluconazole treatment on the oral mycologic flora and on the sensitivity of oral Candida albicans isolates to fluconazole. Saline oral rinses were collected from 89 HIV-positive patients, of whom 48 had been exposed to fluconazole and 41 were fluconazole-naive. The rinses were cultured on Sabouraud's and Pagano Levin agars, and yeasts were identified by standard methods. Fluconazole sensitivity of C. albicans isolates was measured by disk diffusion assay. C. albicans was isolated from 69% of patients who had received fluconazole and from 93% of the patients who were fluconazole-naive (p < 0.05). Nine other species of yeasts were also isolated, most commonly C. glabrata. Five patients previously exposed to fluconazole harbored fluconazole-resistant C. albicans, whereas no resistance was detected among the patients who were fluconazole-naive (p < 0.01). Sixteen of the patients who were fluconazole-exposed carried yeasts other than C. albicans, compared with only five patients in the fluconazole-naive group (p < 0.01). All of the fluconazole-resistant strains were isolated from patients with low CD4 counts (less than 100 cells/ml) and after lengthy fluconazole exposures. Nevertheless, patients in Charlotte, N.C., who had a greater mean fluconazole exposure time (10.25 ± 1.41 months) than patients in Glasgow, UK, (0.65 ± 0.18 months; p < 0.005), did not develop significantly more in vitro resistance or species diversity. This study indicates that long-term fluconazole treatment can have significant effects on the yeast flora of the mouth, particularly in a patient with a CD4 count of less than 100 cells/ml.     

Short telomeres in an oral precancerous lesion: Q-FISH analysis of leukoplakia

J Oral Pathol Med (2012) 41 : 372–378 Objectives: A precancerous condition is a lesion that, if left untreated, leads to cancer or can be induced to become malignant. In the oral region, leukoplakia is a lesion that has been regarded as precancerous. In cases of oral carcinoma, we have frequently noticed that a type of leukoplakia histologically demonstrating hyper‐orthokeratosis and mild atypia (ortho‐keratotic dysplasia; OKD) is often associated with carcinoma, either synchronously or metachronously. Therefore, we consider OKD‐type leukoplakia to be a true precancerous lesion. Materials and Methods: In an attempt to clarify the relationship between OKD as a precancerous condition in the oral mucosa and telomere length, we estimated telomere lengths in this type of leukoplakia using quantitative fluorescence in situ hybridization, and also quantified the frequency of anaphase–telophase bridges (ATBs) in comparison with squamous cell carcinoma in situ (CIS) and the background tissues of CIS and OKD. Results: Ortho‐keratotic dysplasia was frequently associated with squamous cell carcinoma (45.0%) and showed significantly shorter telomeres than normal control epithelium, CIS, or the background of CIS or OKD. The frequency of ATBs was much higher in OKD than in control epithelium or CIS. Conclusion: Ortho‐keratotic dysplasia appears to be frequently associated with carcinoma, chromosomal instability, and excessively shortened telomeres, not only in the lesion itself but also in the surrounding background. Therefore, when this type of leukoplakia is recognized in the oral region, strict follow‐up for oral squamous cell carcinoma is necessary, focusing not only on the areas of leukoplakia, but also the surrounding background.     

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.     

Clinical aspects related to endodontic yeast infections

Yeasts can be detected in 7–18% of infected root canals. They are commonly associated with persistent cases of apical periodontitis, but yeasts can also be isolated in primary apical periodontitis. Their relative proportion of the root canal flora is low, usually below 1%. Therefore, yeast detection with conventional microbiological methods may be difficult. Selective culture media such as Sabouraud dextrose agar combined with cultivation from undiluted sample should be used for primary isolation of yeasts in endodontic infections. Preliminary identification of yeasts is based on typical colony and cellular morphology and a positive catalase test. Detection by molecular biological methods is sensitive, but may occasionally also give false‐positive results. Candida albicans is the most commonly detected yeast species from infected root canals. It is typically found together with Gram‐positive bacteria such as streptococci, but can also be isolated in pure culture, which is an indication of its pathogenicity. A variety of virulence factors enable C. albicans to adhere to and penetrate into dentine. Furthermore, C. albicans tolerates harsh ecological conditions including high alkalinity. Calcium hydroxide is generally not efficient against oral yeasts in vitro; the in vivo effectiveness of calcium hydroxide is not known. However, sodium hypochlorite, iodine compounds, and chlorhexidine have proven effective against yeasts both in experimental conditions as well as in vivo and may offer effective treatment possibilities against endodontic yeast infections.     

The genotypes and virulence attributes of C. albicans isolates from oral leukoplakia

Background There is a debate as to whether some types of oral leucoplakias (OL) are caused by Candida species, and whether they contribute to the malignant transformation, associated with a minority of such lesions. As no detailed population analysis of yeast isolates from OL is available, we evaluated the virulence attributes, and genotypes of 35 C. albicans from OL, and compared their genotypes with 18 oral isolates from healthy individuals.Material and Methods The virulence traits evaluated were esterase, phospholipase, proteinase, haemolysin and coagulase production, and phenotypic switching activity, and yeast adherence and biofilm formation. DNA from OL and control yeasts were evaluated for A, B or C genotype status.Results Phospholipase, proteinase, and coagulase activity and biofilm formation was observed in 80%, 66%, 97 % and 77 % of the isolates, respectively. Phenotypic switching was detected in 8.6%, while heamolytic, and esterase activity and adherence were noted in all isolates.Conclusions The genotype A was predominant amongst both the OL and control groups. Due to the small sample size of our study a larger investigation to define the role of candidal virulent attributes in the pathogenicity of OL is warranted, and the current data should serve as a basis until then. Key words:C. albicans, oral cavity, leukoplakia, virulence factors, genotyping.     

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.     

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.     

Immunohistochemical expression of DNA methyltransferases 1, 3a and 3b in oral leukoplakias and squamous cell carcinomas

Over-expression of DNA methyltransferases DNMT1, DNMT3a and DNMT3b has been reported in various cancers and precancerous lesions.

Objective

To investigate DNMT1, DNMT3a and DNMT3b enzymes in oral squamous cell carcinoma (SCC) and leukoplakia, and their relationship with histopathologic/clinical parameters.

Study design

Immunohistochemistry was carried out to evaluate the three DNMTs in 60 samples of oral SCC and 37 samples of oral leukoplakia.

Results

DNMT3a immunoreactivity in the three groups of oral SCC (39.8%) was significantly higher than in control (22.6%) (ANOVA, Student–Newman–Keuls test, P < 0.05), but not when compared to oral leukoplakia groups (28.2%). For DNMT1 and DNMT3b, there were no statistically significant differences between oral SCC groups (65% and 74.7%), oral leukoplakia groups (68.3% and 70.9%) and control (65.4% and 76.5%). There was a significantly higher mean percentage of DNMT1 immunoreactivity in non-smokers (ANOVA, P = 0.048), and a higher DNMT3a immunoreactivity in alcohol users (ANOVA, P = 0.01).

Conclusions

Higher DNMT3a immunopositivity may be associated with oral SCC and alcohol use, whilst lower levels of DNMT1 may be related with smoking habit. However, there was a significantly higher mean percentage of DNMT1 immunoreactivity in non-smokers (ANOVA, P = 0.048), and a higher DNMT3a immunoreactivity in alcohol users (ANOVA, P = 0.010).     

Candida glabrata and Candida albicans co‐infection of an in vitro oral epithelium

J Oral Pathol Med (2010) 40 : 421–427 Background: Candida albicans is regarded as the leading of candidosis. However, Candida glabrata has emerged as an important pathogen of oral mucosa, occurring both singly or in mixed species infections, often with C. albicans. Compared with C. albicans, little is known about the role of C. glabrata in oral infection. The aim of this study was to examine single and mixed species infection of oral epithelium involving C. glabrata and establish its ability to invade and damage tissue. Methods: A reconstituted human oral epithelium (RHOE) was infected only with C. glabrata, or simultaneously with C. glabrata and C. albicans. The ability of both species to invade the tissue was examined using species specific peptide nucleic acid (PNA) probe hybridization and confocal laser scanning microscopy. Epithelial damage was assessed by measuring lactate dehydrogenase (LDH) activity. Results: Candida glabrata strains were able to colonize the RHOE, in a strain dependent manner. Candida glabrata single infection after 12 h, generally revealed no invasion of the RHOE, which contrasted with extensive tissue invasion demonstrated by C. albicans. Mixed infection showed that C. albicans enhanced the invasiveness of C. glabrata, and led to increased LDH release by the RHOE, which paralleled the observed histological damage. Conclusions: The results obtained demonstrating enhanced invasion and increased tissue damage caused by mixed C. glabrata and C. albicans infections have important clinical significance and highlight the need to identify Candida species involved in oral candidosis.     

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.     

Natural history of Candida species and yeasts in the oral cavities of infants

A longitudinal study was made of the presence of Candida species and yeasts in infants from the day of birth until 1-yr-old, diagnosed by the culture of oral swab samples. The isolation rate of Candida rose from 5.7 per cent neonatally to 14.2 per cent on discharge from hospital at 7 days. The prevalence increased sharply when the infants returned home, reaching 82 per cent at 4 weeks of age, but then slowly declined to 50 per cent at 1 yr. At 4 weeks, yeasts were isolated from an additional 14 per cent of infants. The predominant species was C. albicans with relatively few isolates of C. tropicalis, C. parapsilosis and C. pseudotropicalis. Estimates of neonatal serum antibody to C. albicans showed no correlation with initial or subsequent oral colonization by this organism.