Detection of Pneumocystis jirovecii by quantitative real-time PCR in oral rinses from Pneumocystis pneumonia asymptomatic human immunodeficiency virus patients.

Pneumocystis pneumonia (PCP) is a potentially life-threatening fungal infection usually seen in immunocompromised patients. Pneumocystis jirovecii can be easily detected from oral rinse samples in HIV patients with suspected PCP. In this study, a quantitative real-time PCR assay was used to establish the frequency of detection of P. jirovecii in oral rinses from HIV patients without respiratory symptoms or suspicion of PCP. Two saline oral rinses were collected from 100 ambulant HIV patients and from 60 COPD patients (comparator group). Four HIV patients were positive for P. jirovecii. In three patients, the first sample was positive and in one the second one was positive. One of these patients was on PCP prophylaxis and had a CD4+ count of 76 cells/mm3. The mean CD4+ count for all patients was 527 cells/mm3. All qRT-PCR test results for the COPD patients were negative. No patient developed PCP at six months follow-up. The qRT-PCR assay can be used to detect P. jirovecii DNA in oral rinse samples from HIV patients without evident clinical symptoms, however the oral carriage of this fungus was rare in our cohort of patients. In conclusion, although rare, a positive oral rinse P. jirovecii result may reflect colonisation, in particular in patients with HIV. This needs to be kept in mind when using oral rinses and qRT-PCR in the diagnosis of P. jirovecii infection.

2-Hydroxyisocaproic acid is bactericidal in human dental root canals ex vivo.

AIM: To compare the activity of 2-hydroxyisocaproic acid (HICA), calcium hydroxide (Ca(OH)2 ) and chlorhexidine digluconate (CHG) against Enterococcus faecalis T-75359 (root canal isolate) in the root canals of extracted human teeth. METHODOLOGY: Bacterial suspensions (108  cfu mL-1 ) were incubated in root canals with 0.9 mm diameter root blocks (n = 73) for 21 days. Bacterial penetration into dentine was analysed by the Brown and Brenn method (n = 5). Canals (n = 17/group) were medicated with 40% of HICA paste, 40% of Ca(OH)2 paste, 2% of CHG solution or 0.9% of saline solution for 7 days. Samples taken from the inner (first 0.1 mm) and deeper (second 0.1 mm) dentine, and residual roots were cultured in broth for 24 h. Bacterial growth was detected by spectrophotometry (optical density, OD) and confirmed by culture on agar. The OD data were analysed with Kruskal-Wallis and Friedman with Wilcoxon signed-rank test between and within groups, respectively, and agar culture data with Pearson chi-square with Mann-Whitney and Cochran with McNemar tests, respectively (P < 0.05). RESULTS: Bacterial invasion into dentine tubules was confirmed. In deeper dentine, HICA inhibited >90% of bacterial growth in comparison with saline. No bacterial growth was observed in 82-100% of inner and deeper dentine samples. CHG prevented the growth in 88%, Ca(OH)2 in 59-76% and saline in 65-71%, respectively. HICA was significantly more active than Ca(OH)2 (P = 0.008) in the residual roots. The viability testing on agar showed essentially the same result. CONCLUSION: HICA paste exerted superior activity against E. faecalis and could have potential for root canal medication.

Antimicrobial 2-hydroxyisocaproic acid and chlorhexidine resist inactivation by dentine.

AIM: To compare the antibacterial activity of 2-hydroxyisocaproic acid (HICA) with currently used root canal medicaments and to examine their interactions with potential inhibitors in nutrient-deficient and nutrient-rich conditions. METHODOLOGY: First, the antibacterial activity of single concentrations of HICA, calcium hydroxide solution or slurry, chlorhexidine digluconate or acetate was tested against Enterococcus faecalis with and without potential inhibitors: dentine powder (DP), hydroxyapatite or bovine serum albumin, in a low concentration of peptone water. Relative viable counts were determined by culture at 1, 24 and 48 h. In the second set of experiments, the activity of three concentrations of HICA was evaluated against two isolates of E. faecalis with and without potential inhibitors in nutrient-rich thioglycollate broth using a modification of a standard microdilution method. The minimum bactericidal concentration was determined by culture at 1, 24 and 48 h. RESULTS: Concentrations of ≥33 mg mL(-1) of HICA were found to be bactericidal against E. faecalis in both nutrient-deficient and nutrient-rich environments at 24- to 48-h incubation, whereas the initial activity of Ca(OH)2 slurry was lost at 48-h incubation. HICA tolerated well all tested potential inhibitors up to 19 mg mL(-1) . DP concentrations higher than this inhibited its activity in a dose-dependent manner in both environments. DP demonstrated moderate antibacterial activity, and it enhanced the otherwise limited activity of Ca(OH)2 slurry and solution. DP did not impact on the activity of chlorhexidine. CONCLUSIONS: These results support the long-term antibacterial activity of HICA and indicate its tolerance to clinically relevant concentrations of dentine and other inhibitors commonly present in the root canal system. Therefore, HICA may have potential as an interappointment medication in the treatment of root canal infections.

DL-2-hydroxyisocaproic acid attenuates inflammatory responses in a murine Candida albicans biofilm model.

Chronic biofilm infections are often accompanied by a chronic inflammatory response, leading to impaired healing and increased, irreversible damage to host tissues. Biofilm formation is a major virulence factor for Candida albicans and a challenge for treatment. Most current antifungals have proved ineffective in eradicating infections attributed to biofilms. The biofilm structure protects Candida species against antifungals and provides a way for them to evade host immune systems. This leads to a very distinct inflammatory response compared to that seen in planktonic infections. Previously, we showed the superior efficacy of dl-2-hydroxyisocaproic acid (HICA) against various bacteria and fungi. However, the immunomodulatory properties of HICA have not been studied. Our aim was to investigate the potential anti-inflammatory response to HICA in vivo. We hypothesized that HICA reduces the levels of immune mediators and attenuates the inflammatory response. In a murine model, a robust biofilm was formed for 5 days in a diffusion chamber implanted underneath mouse skin. The biofilm was treated for 12 h with HICA, while caspofungin and phosphate-buffered saline (PBS) were used as controls. The pathophysiology and immunoexpression in the tissues surrounding the chamber were determined by immunohistochemistry. Histopathological examination showed an attenuated inflammatory response together with reduced expression of matrix metalloproteinase 9 (MMP-9) and myeloperoxidase (MPO) compared to those of chambers containing caspofungin and PBS. Interestingly, the expression of developmental endothelial locus 1 (Del-1), an antagonist of neutrophil extravasation, increased after treatment with HICA. Considering its anti-inflammatory and antimicrobial activity, HICA may have enormous therapeutic potential in the treatment of chronic biofilm infections and inflammation, such as those seen with chronic wounds.

2-hydroxyisocaproic acid is fungicidal for Candida and Aspergillus species.

The amino acid derivative 2-hydroxyisocaproic acid (HICA) is a nutritional additive used to increase muscle mass. Low levels can be detected in human plasma as a result of leucine metabolism. It has broad antibacterial activity but its efficacy against pathogenic fungi is not known. The aim was to test the efficacy of HICA against Candida and Aspergillus species. Efficacy of HICA against 19 clinical and reference isolates representing five Candida and three Aspergillus species with variable azole antifungal sensitivity profiles was tested using a microdilution method. The concentrations were 18, 36 and 72 mg ml(-1) . Growth was determined spectrophotometrically for Candida isolates and by visual inspection for Aspergillus isolates, viability was tested by culture and impact on morphology by microscopy. HICA of 72 mg ml(-1) was fungicidal against all Candida and Aspergillus fumigatus and Aspergillus terreus isolates. Lower concentrations were fungistatic. Aspergillus flavus was not inhibited by HICA. HICA inhibited hyphal formation in susceptible Candida albicans and A. fumigatus isolates and affected cell wall integrity. In conclusion, HICA has broad antifungal activity against Candida and Aspergillus at concentrations relevant for topical therapy. As a fungicidal agent with broad-spectrum bactericidal activity, it may be useful in the topical treatment of multispecies superficial infections.

Fermentative 2-carbon metabolism produces carcinogenic levels of acetaldehyde in Candida albicans.

UNLABELLED: Acetaldehyde is a carcinogenic product of alcohol fermentation and metabolism in microbes associated with cancers of the upper digestive tract. In yeast acetaldehyde is a by-product of the pyruvate bypass that converts pyruvate into acetyl-Coenzyme A (CoA) during fermentation. THE AIMS OF OUR STUDY WERE: (i) to determine the levels of acetaldehyde produced by Candida albicans in the presence of glucose in low oxygen tension in vitro; (ii) to analyse the expression levels of genes involved in the pyruvate-bypass and acetaldehyde production; and (iii) to analyse whether any correlations exist between acetaldehyde levels, alcohol dehydrogenase enzyme activity or expression of the genes involved in the pyruvate-bypass. Candida albicans strains were isolated from patients with oral squamous cell carcinoma (n = 5), autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy (APECED) patients with chronic oral candidosis (n = 5), and control patients (n = 5). The acetaldehyde and ethanol production by these isolates grown under low oxygen tension in the presence of glucose was determined, and the expression of alcohol dehydrogenase (ADH1 and ADH2), pyruvate decarboxylase (PDC11), aldehyde dehydrogenase (ALD6) and acetyl-CoA synthetase (ACS1 and ACS2) and Adh enzyme activity were analysed. The C. albicans isolates produced high levels of acetaldehyde from glucose under low oxygen tension. The acetaldehyde levels did not correlate with the expression of ADH1, ADH2 or PDC11 but correlated with the expression of down-stream genes ALD6 and ACS1. Significant differences in the gene expressions were measured between strains isolated from different patient groups. Under low oxygen tension ALD6 and ACS1, instead of ADH1 or ADH2, appear the most reliable indicators of candidal acetaldehyde production from glucose.

Chlorhexidine-impregnated PEM/THFM polymer exhibits superior activity to fluconazole-impregnated polymer against Candida albicans biofilm formation.

Biofilm-associated infections represent a major challenge for biomaterials. Methods to alter the chemical characteristics of biomaterials offer an attractive solution for enhanced microbial control. The aim of this study was to investigate the efficacy of a poly(ethyl methacrylate)/tetrahydrofurfuryl methacrylate (PEM/THFM) acrylic model impregnated with fluconazole (FLU) or chlorhexidine (CHX) in preventing Candida biofilm formation in vitro. PEM/THFM disks impregnated with CHX (n=50) or FLU (n=50) and drug-free control disks (n=50) were infected with Candida albicans ATCC 90028. Disks were incubated for 2, 7, 14, 21 or 28 days at 37 °C and the biofilm biomass and metabolic activity was quantified at each time point using crystal violet staining and XTT [2,3-bis(2-methoxy-4-nitro-5-sulfo-phenyl)-2H-tetrazolium-5-carboxanilide] reduction assay. FLU disks exhibited poor overall biofilm inhibitory characteristics, with mean metabolic and biomass inhibition of 12.6% and 8.8%, respectively. Conversely, CHX disks were highly effective, significantly inhibiting biofilm development by 75% (P ≤ 0.001) and its metabolism by 84% (P ≤ 0.001) for all time points tested. The notable efficacy of CHX against C. albicans biofilms is a promising outcome to overcome the side effects and poor relative activity of conventional antifungal agents against Candida biofilms. These findings indicate that impregnation of PEM/THFM with antimicrobials has potential as a treatment modality for denture stomatitis.

Candidacidal effect of fluconazole and chlorhexidine released from acrylic polymer.

OBJECTIVES: To investigate the efficacy and rate of killing of a fluconazole- or chlorhexidine-impregnated polymeric delivery system against fluconazole-susceptible and -resistant Candida albicans and fluconazole-resistant Candida glabrata. METHODS: Poly(ethyl methacrylate)/tetrahydrofurfuryl methacrylate (PEM/THFM) discs impregnated with chlorhexidine, pure fluconazole (FLCp) or fluconazole from capsules (FLCc) were prepared by substituting a portion of PEM powder with an equivalent amount of each drug. Discs were incubated in sterile water for 1, 3, 7, 14, 21 and 28 days. The amounts of drugs in the leachates were measured spectrophotometrically and their antifungal activity against fluconazole-susceptible (n=1) and fluconazole-resistant (n=2) candidal isolates was determined using a time-kill method and by comparing the released concentrations with the corresponding MICs. RESULTS: Fluconazole and chlorhexidine leached from PEM/THFM polymer for up to 28 days and the released concentrations were fungicidal against all three Candida isolates for at least the first 7 days. Chlorhexidine leachates killed all Candida isolates more rapidly than the two fluconazole formulation leachates throughout the study period. FLCc leachates required longer incubation for 100% killing than FLCp leachates. The proportion of viable C. glabrata dropped more slowly than that of C. albicans with the same MIC. CONCLUSIONS: The concentrations of chlorhexidine and fluconazole leached from the PEM/THFM polymer were fungicidal against all Candida isolates, including those resistant to fluconazole, for the first 7 days. Chlorhexidine leachates showed a rapid fungicidal activity for up to 4 weeks, which can be of use in cases with poor response to conventional antifungals.

Production of carcinogenic acetaldehyde by Candida albicans from patients with potentially malignant oral mucosal disorders.

OBJECTIVES: Production of carcinogenic acetaldehyde by Candida has been suggested to contribute to epithelial dysplasia and oral carcinogenesis. Oral lichen planus (OLP), oral lichenoid lesion (OLL) and oral leukoplakia (OL) are potentially carcinogenic oral diseases where colonisation by Candida is common, but acetaldehyde production by Candida has not been studied. STUDY DESIGN: Acetaldehyde production in ethanol (11 mM), glucose (100 mM), ethanol-glucose (11 mM and 100 mM) or red wine (1200 mM ethanol) incubation by Candida albicans from patients with OLL (n = 6), OLP (n = 16), OL (n = 6) and controls (n = 6) was measured by gas chromatography. Participants completed a questionnaire regarding their smoking habits and alcohol consumption. RESULTS: All Candida albicans isolates produced potentially carcinogenic levels of acetaldehyde (>100 µM) in all incubations containing ethanol. The control group isolates produced the highest acetaldehyde levels. Isolates from smokers produced more acetaldehyde in all incubations than those from non-smokers. The difference was significant in ethanol-glucose incubation. Isolates from patients who were both smokers and drinkers produced the highest amounts when incubated in ethanol, ethanol-glucose and wine. CONCLUSIONS: Candida albicans isolated from potentially carcinogenic oral diseases can produce mutagenic amounts of acetaldehyde. Cigarette smoking and alcohol consumption may favour adaptational changes resulting in the upregulation of candidal acetaldehyde metabolism.

Chlorhexidine is a highly effective topical broad-spectrum agent against Candida spp.

The objective of this study was to compare the in vitro antifungal activities of chlorhexidine (CHX) and fluconazole (FLZ) against Candida isolates comprising eight different species associated with oral candidosis. A broth microdilution method as described in Clinical and Laboratory Standards Institute (CLSI) protocol M27-A3 was used to determine susceptibility. A total of 79 clinical isolates and reference strains belonging to eight different Candida spp. was tested. The minimum inhibitory concentration (MIC) was the lowest drug concentration that reduced growth by 50% for FLZ at 48 h and by 80% for CHX at 24h and 48 h. The geometric mean MIC (and MIC range) at 48 h for CHX was 3.03 mg/L (0.78-6.25mg/L) and for FLZ was 19.12 mg/L (≤0.125-256 mg/L). Of the 79 isolates, 14 (18%) were resistant to FLZ (MIC≥64 mg/L). All isolates were effectively inhibited by ≤6.25mg/L CHX, and Candida CHX MICs are below the CHX levels found in saliva following normal dosing. No cross-resistance between CHX and FLZ was detected (r(s)=0.039, P=0.733). CLSI M27-A3 methodology proved to provide reproducible results with clear endpoints for CHX. In conclusion, the findings showed that CHX has excellent broad-spectrum antifungal activity in vitro. It was effective at concentrations detected in saliva when using standard dosing regimens. Moreover, no cross-resistance was detected between CHX and FLZ, even among Candida spp. highly resistant to FLZ.

Spectroscopy in the analysis of bacterial and eukaryotic cell footprints on implant surfaces.

We tested the suitability of two spectroscopic methods, x-ray photoelectron spectroscopy (XPS) and time of flight secondary ion mass spectrometry (ToF-SIMS), in the recognition of bacterial and eukaryotic cell footprints on implant surfaces. Human mesenchymal stem cells (MSCs) and Staphylococcus aureus were cultured on sample surfaces and detached using trypsin. Scanning electron microscopy confirmed that the processed surfaces did not contain any human or microbial cells. The footprints were then analysed using XPS and ToF-SIMS. XPS results showed no significant differences between the footprints, but principal component analysis of the ToF-SIMS data enabled clear separation of MSC-footprints from the S. aureus and co-culture footprints (p < 0.03). ToF-SIMS also demonstrated 'race for the surface' between proteins, which suggest surface charge (zeta-potential) dependent protein adsorption. ToF-SIMS differentiated eukaryotic and bacterial footprints and has potential for post-hoc detection of implant-related infections based on the typical ToF-SIMS spectra.

Clinical manifestations and management of patients with autoimmune polyendocrine syndrome type I.

Autoimmune polyendocrine syndrome type I (APS-I) is a monogenic model disease of autoimmunity. Its hallmarks are chronic mucocutaneous candidosis, hypoparathyroidism and adrenal insufficiency, but many other autoimmune disease components occur less frequently. The first components usually appear in childhood, but may be delayed to adolescence or early adult life. There is enormous variation in presentation and phenotype, which makes the diagnosis difficult. Antibodies against interferon-omega and -alpha have recently been shown to be sensitive and relatively specific markers for APS-I, and mutational analysis of the autoimmune regulator gene gives the diagnosis in >95% of cases. The treatment and follow-up of patients is demanding and requires the collaboration of specialists of several fields. However, the literature is especially sparse regarding information on treatment and follow-up; hence, we present here a comprehensive overview on clinical characteristics, treatment and follow-up based on personal experience and published studies.

Oral infections and systemic disease--an emerging problem in medicine.

The relationship between oral and general health has been increasingly recognised during the past two decades. Several epidemiological studies have linked poor oral health with cardiovascular disease, poor glycaemic control in diabetics, low birth-weight pre-term babies, and a number of other conditions, including rheumatoid arthritis and osteoporosis. Oral infections are also recognised as a problem for individuals suffering from a range of chronic conditions, including cancer and infection with human immunodeficiency virus, as well as patients with ventilator-associated pneumonia. This review considers the systemic consequences of odontogenic infections and the possible mechanisms by which oral infection and inflammation can contribute to cardiovascular disease, as well as the oral conditions associated with medically compromised patients. A large number of clinical studies have established the clinical efficacy of topical antimicrobial agents, e.g., chlorhexidine and triclosan, in the prevention and control of oral disease, especially gingivitis and dental plaque. The possible risks of antimicrobial resistance are a concern, and the benefits of long-term use of triclosan require further evaluation. Oral infections have become an increasingly common risk-factor for systemic disease, which clinicians should take into account. Clinicians should increase their knowledge of oral diseases, and dentists must strengthen their understanding of general medicine, in order to avoid unnecessary risks for infection that originate in the mouth.

Candida albicans does not invade carious human dentine.

AIM: Candida albicans has been proposed to be a caries pathogen, but the evidence for its specific role is lacking. To be considered significant in caries progression, a marked amount of yeasts should be present in a lesion. The aim of the study was to investigate the presence of C. albicans in dentinal caries lesions. MATERIALS AND METHODS: To demonstrate the extension of caries and to identify the bacteria in a lesion, sections of 10 carious human teeth were stained with Gram and Giemsa stains. C. albicans was detected with periodic acid-Schiff (PAS) staining and by immunohistochemistry using a C. albicans-specific antibody 3H8. Thirty sections were used for each staining (in total 120 sections). RESULTS: Extensive bacterial invasion and intensive staining by PAS occurred in all samples. However, with the C. albicans-specific antibody, only 30 (3.3%) sections stained weakly positive, with a few stained cells on the lesion surface. However, the positive identification of C. albicans, based on the morphology of the cells, was not possible. CONCLUSIONS: The results do not support the previous suggestion that C. albicans is important in the dentine caries pathology. In addition, because of its unspecific nature, PAS turned out to be an unsuitable method for detecting yeasts in carious tooth samples.

Expression of interleukin-8 and its receptor IL-8RA in chronic hyperplastic candidosis.

INTRODUCTION: Neutrophils are the main opponents of Candida albicans in chronic hyperplastic candidosis. They migrate from the circulation to the epithelium where they form microabscesses. We therefore hypothesized that the neutrophil chemokine interleukin-8 (IL-8) might play a role in the neutrophil-Candida interaction. METHODS: Biopsies from patients with chronic hyperplastic candidosis (n = 10) were stained using the avidin-biotin-peroxidase complex protocol for IL-8 and IL-8 receptor A and were compared to healthy control mucosa (n = 3). A set of C. albicans agar sections was similarly analysed. RESULTS: In chronic hyperplastic candidosis lesions IL-8 was strongly expressed in both vascular endothelium and mucosal epithelium. Many resident and immigrant inflammatory cells, including intraepithelial neutrophils, were IL-8 receptor A positive. In addition, IL-8 (or an analogue) was found in the candidal mother cell in chronic hyperplastic candidosis and in agar, whereas the tips of the hyphae expressed IL-8 receptor A (or an analogue). CONCLUSION: IL-8 may play a role in the recruitment of neutrophils from the vascular compartment to the epithelial microabscesses. C. albicans may have developed an ability to sense IL-8. The IL-8 ligand-receptor interaction may help to direct the growth of the IL-8-receptor-containing tips of the hyphae away from the IL-8-producing candidal cell body (a centrifugal growth pattern to facilitate host tissue penetration). Later, this ability might help to keep the vulnerable hyphal tips away from areas with high concentrations of host IL-8 and candidacidal neutrophils. We suggest that this phenomenon, in contrast to chemotropism, is named chemophobia.

Bacterial aerosols in dental practice - a potential hospital infection problem?

Aerosols containing microbes from the oral cavity of the patient are created when using modern high-speed rotating instruments in restorative dentistry. How far these aerosols spread and what level of contamination they cause in the dental surgery has become a growing concern as the number of patients with oro-nasal meticillin-resistant Staphylococcus aureus colonization has increased. The present study aimed to determine how far airborne bacteria spread during dental treatment, and the level of contamination. Fall out samples were collected on blood agar plates placed in six different sectors, 0.5-2m from the patient. Restorative dentistry fallout samples (N=72) were collected from rooms (N=6) where high-speed rotating instruments were used, and control samples (N=24) were collected from rooms (N=4) used for periodontal and orthodontic treatment where rotating and ultrasonic instruments were not used. The collection times were 1.5 and 3 h. In addition, samples were taken from facial masks of personnel and from surfaces in the rooms before and after disinfection. After 48 h of incubation at 37 degrees C, colonies were counted and classified by Gram stain. The results showed significant contamination of the room at all distances sampled when high-speed instruments were used (mean 970 colony-forming units/m2/h). The bacterial density was found to be higher in the more remote sampling points. Gram-positive cocci, namely viridans streptococci and staphylococci, were the most common findings. The area that becomes contaminated during dental procedures is far larger than previously thought and practically encompasses the whole room. These results emphasize the need for developing new means for preventing microbial aerosols in dentistry and protection of all items stored temporarily on work surfaces. This is especially important when treating generally ill or immunocompromised patients at dental surgeries in hospital environments.

Specificity of the monoclonal antibody 3H8 in the immunohistochemical identification of Candida species.

Candida albicans has been shown to be involved in the pathogenesis of adult periodontitis (AP). The diagnosis of Candida-associated AP depends largely on the identification of yeast and pseudomycelial forms in gingival tissue samples by using periodic acid-Schiff and Gomori methenamine silver stains. However, these stains are non-specific and also reveal confusing artifacts seemingly rather difficult to distinguish from yeasts. With the recent development and availability of monoclonal antibodies (Mabs) to various epitopes of C. albicans, for example Mab 3H8 which recognizes a mannoprotein, it is now possible to identify Candida in human tissue biopsies. To explore further the usefulness of this Mab in detecting Candida in periodontal disease the antibody was tested against a wide range of yeast species and strains and various morphological forms, grown in agar blocks at various temperatures and for various time periods. Furthermore, considering the location of the 3H8 epitope on the external cell wall of certain C. albicans strains, it seemed reasonable to determine whether the epitope could be expressed into the surrounding environment, further aiding the recognition of the organism in tissue. The 3H8 epitope appeared to be located at the external surface and on the septum between the mother cell and germ tube of some C. albicans strains but it was partially cryptic in the cell wall of other strains. Both yeast blastoconidia and pseudohyphae were labeled by the 3H8 antibody. Candida lusitaniae, C. glabrata, C. krusei, C. parapsilosis and C. tropicalis did not posses the epitope. The epitope was expressed extracellularly by both blastoconidia and pseudohyphae of C. albicans. This Mab appears to be suitable for the identification of C. albicans in periodontal tissue and may provide further insight into the role of C. albicans in the pathogenesis and diagnosis of periodontal diseases.

In vitro adhesion of Candida species to denture base materials.

Adhesion of Candida species to prosthetic acrylic resins is an essential first step in the pathogenesis of denture stomatitis. Data on the relative adhesion of pathogenic non-albicans Candida species to different denture base materials are sparse. The purpose of the present study was to investigate in vitro adhesion of C. albicans, C. glabrata, C. krusei and C. dubliniensis to four different denture base materials. Specimens of both heat-cured resins (Vertex(TM) Rapid Simplified and ProBase Hot) and cold-cured resins (Paladur A and Paladur B) were prepared using a novel method and the adhesion of four strains each of the foregoing Candida species evaluated microscopically using a soft imaging system. There was a significant difference in yeast adherence between Vertex and the other resins. Only C. glabrata attached to Vertex, while all the remainder of the tested species adhered to all other resins tested except ProBase, which resisted C. krusei adhesion. There was a significant difference in candidal adhesion between cold-cured and heat-cured resins for three Candida species (C. albicans, P = 0.039; C. glabrata, P = 0.002 and C. krusei, P = 0.000). The type of denture base material and whether they are heat-cured or cold-cured play an important role in modifying candidal adhesion.

Intracellular localization of Porphyromonas gingivalis thiol proteinase in periodontal tissues of chronic periodontitis patients.

OBJECTIVES: Porphyromonas gingivalis is a significant periodontal pathogen that has been shown in vitro to be able to invade gingival epithelial cells and grow intracellularly. The aim of the present study was to detect P. gingivalis in gingival tissues from chronic periodontitis (CP) patients. MATERIALS AND METHODS: Monoclonal antibodies specific to a cell membrane-bound thiol proteinase of P. gingivalis were used to detect the microbe in gingival tissues of CP patients (n = 13) by immunohistochemistry. The presence of P. gingivalis was also analysed by polymerase chain reaction (PCR). RESULTS: Immunohistochemical analysis of the periodontal tissues revealed positive staining for P. gingivalis thiol proteinase in 11 of the 13 patients. Positive staining was mainly located intracellularly in the perinuclear region of the cytoplasm in the periodontal epithelial cells and it could be detected throughout the whole depth of both pocket and oral epithelium. The sensitivity of immunohistochemistry was found to be comparable with that of PCR. CONCLUSIONS: Our results provide in vivo evidence of the ability of P. gingivalis to enter human gingival epithelial cells. Intracellular localization of P. gingivalis contributes to its evasion of the host immune surveillance and eventually increases its resistance to conventional treatments of periodontal diseases.