Oral Colonization and Virulence Factors of Candida spp. in Babies With Cleft Palate.

OBJECTIVE: To examine oral colonization and virulence factors of Candida spp. in patients aged from 0 to 18 months with cleft palate (CP). MATERIALS AND METHODS: Sixty babies were allocated into 3 groups: CP, CP with orthodontic plate (CPwP), and control group (Ctrl) without CP. Information on feeding habits, hygiene, and history of candidosis was collected. The presence of Candida spp. was investigated in samples of saliva. Fungal hydrophobicity, protease, esterase, phospholipase, and hemolysin were evaluated in a semiquantitative manner. RESULTS: Positive oral isolations of Candida spp. were detected in CP (89.5%), CPwP (100%), and Ctrl (44%) groups. Candidosis was more reported in the cleft groups than in the Ctrl group (P ≤ .023). There was a higher prevalence of Candida albicans, followed by Candida krusei, Candida tropicalis, and Candida parapsilosis in all groups. There was no uniformity of expression of virulence factors, either among different species or among different groups. CONCLUSION: Candida spp. colonization occurred in all groups, being superior in CPwP group. Candidosis episodes were more reported in patients from CPwP than in other groups, although candidosis was also registered in other groups. Candida albicans was the predominant species and virulence factors did not exhibit any pattern for species or groups of patients.

Randomized clinical trial of complete denture cleaning methods in residents of care institutions

Aim: To evaluate the efficacy of ultrasonic bath and traditional brushing in the hygiene of complete dentures of dependent residents in long-term care institutions (LTCIs). Methods: A randomized cross-over clinical study was conducted in 17 maxillary complete denture wearers living in LTCIs. Cleaning protocols were brushing or ultrasonic bath, both applied with neutral liquid soap. Biofilm biomass were estimated by MTT reduction assay and specific microbial load (CFU) of Streptococcus spp., Staphylococcus spp. and Candida spp. were quantified by selective and differential culture media. Results: Ultrasound method showed higher percentage reduction of biofilm biomass and specific microbial loads of Streptococcus spp. compared to brushing (p<0.01). Reduction of microbial loads of the other microorganisms were not different between cleaning methods (p>0.05). Conclusions: The ultrasonic bath proved to be a feasible alternative method for the mechanical cleaning of complete dentures in LTCIs

Growing Candida albicans Biofilms on Paper Support and Dynamic Conditions.

A stainless steel paper-embedded biofilm reactor (PEBR) was developed for Candida spp. growth, permitting confluent distribution of nutrients by capillary diffusion through ordinary laboratory filter paper. Antibiogram disks were distributed along the filter paper rim, and the PEBR received 0.1 or 0.01 % crystal violet (CV) at 200 µL min(-1) and at 37 °C, for 48 h. CV was recovered from the disks and measured at 540 nm. Candida albicans SC5314 cells were applied onto antibiogram disks. The bioreactor was assembled, and YEPD broth was admitted (200 µL min(-1)) at 37 °C, for 72 h. Biofilm growth was estimated via the MTT reduction test. Controls were disks that received the same treatments, except for the fungus. The PEBR was considered high-throughput table, low-cost, and feasible to grow C. albicans biofilms.

Influence of glyphosate in planktonic and biofilm growth of Pseudomonas aeruginosa.

This study evaluated the impact of different concentrations of glyphosate (Rondup(®)) on planktonic and biofilm growth of P. aeruginosa. Aerobic and anaerobic cultures of P. aeruginosa ATCC(®)15442 inoculated in MHB + glyphosate (0.845 ppm, 1.690 ppm, 8.45 ppm, 16.90 ppm, 84.50 ppm, 169 ppm, 845 ppm, and 1690 ppm) and cultured in normoxia and anoxia, following their OD(560nm) every hour for 24 h. Biofilms of adapted cells were formed in the presence of glyphosate (0.845 to 1690 ppm) in normoxia and anoxia for 36 h. Glyphosate at concentrations higher than 84.5 ppm reduces the cell density of planktonic aerobic cultures (p < 0.05). However, these same concentrations favor the planktonic anaerobic growth (p < 0.05). On the other hand, the herbicide favors a slight growth of biofilms in a concentration-dependent manner up to 84.5 ppm (p > 0.05), and more pronounced over 169 ppm. Anaerobic biofilms have their growth more readily favored (p < 0.05), regardless of concentration. In a concentration-dependent manner, glyphosate interferes with the growth ability of P. aeruginosa ATCC(®)15442.

Influence of glyphosate in planktonic and biofilm growth of Pseudomonas aeruginosa

This study evaluated the impact of different concentrations of glyphosate (Rondup®) on planktonic and biofilm growth of P. aeruginosa. Aerobic and anaerobic cultures of P. aeruginosa ATCC®15442 inoculated in MHB + glyphosate (0.845 ppm, 1.690 ppm, 8.45 ppm, 16.90 ppm, 84.50 ppm, 169 ppm, 845 ppm, and 1690 ppm) and cultured in normoxia and anoxia, following their OD560nm every hour for 24 h. Biofilms of adapted cells were formed in the presence of glyphosate (0.845 to 1690 ppm) in normoxia and anoxia for 36 h. Glyphosate at concentrations higher than 84.5 ppm reduces the cell density of planktonic aerobic cultures (p < 0.05). However, these same concentrations favor the planktonic anaerobic growth (p < 0.05). On the other hand, the herbicide favors a slight growth of biofilms in a concentration-dependent manner up to 84.5 ppm (p > 0.05), and more pronounced over 169 ppm. Anaerobic biofilms have their growth more readily favored (p < 0.05), regardless of concentration. In a concentration-dependent manner, glyphosate interferes with the growth ability of P. aeruginosa ATCC®15442.

Low virulent oral Candida albicans strains isolated from smokers.

It is widely accepted that tabagism is a predisposing factor to oral candidosis and cumulate data suggest that cigarette compounds may increase candidal virulence. To verify if enhanced virulence occurs in Candida albicans from chronic smokers, a cohort of 42 non-smokers and other of 58 smokers (all with excellent oral conditions and without signs of candidosis) were swabbed on tong dorsum and jugal mucosa. Results showed that oral candidal loads do not differ between smoker and non-smokers. Activities of secreted aspartyl-protease (Sap), phospholipase, chondroitinase, esterase-lipase, and haemolysin secretions were screened for thirty-two C. albicans isolates. There were detected significant increments in phospholipasic and chondroitinasic activities in isolates from non-smokers. For other virulence factors, no differences between both cohorts were achieved.

Enhancement of secretory aspartyl protease production in biofilms of Candida albicans exposed to sub-inhibitory concentrations of fluconazole.

The production of Secretory Aspartyl Proteases (Sap) is an important virulence factor of Candida albicans. Many studies have shown that a challenge with sub-inhibitory concentrations of antifungals lead species of Candida to the secretion of higher concentrations of Sap. Nevertheless, published studies only reported the secretion of such enzymes by cells growing in planktonic phase, with few mention of biofilms. The present study evaluated the alterations in the secretion of Sap by C. albicans grown in biofilms and exposed to sub-inhibitory concentrations of fluconazole. The MICs for fluconazole of seven clinical strains were determined for planktonic cells. Biofilm and planktonic cells were grown in the presence of ½ MIC, » MIC, and no medication (control). The relative metabolic activity, indirectly related to cell loads, were estimated by the absorbance of reduced XTT and the Sap activity was evaluated by bovine albumin test. It was observed that 72 h-old biofilms under the influence of ½ MIC had fewer cells than » MIC and control. The production of Sap was inversely proportional to the cell content, with higher secretion in ½ MIC, followed by » MIC and control. Biofilms of C. albicans challenged by sub-MICs of fluconazole tend to secrete higher quantities of Sap.

Time-related increase of staphylococci, Enterobacteriaceae and yeasts in the oral cavities of comatose patients.

BACKGROUND/PURPOSE: The composition of oral microbiota in comatose patients remains uncertain. Some pulmonary pathogens may be found in dental biofilms or as part of the saliva microbiota. It is supposed that some pneumopathogenic microorganisms may overgrow in the mouths of comatose patients and spread to their lungs. METHODS: The oral colonization dynamics of staphylococci, Enterobacteriaceae and yeasts in nine comatose patients (group 1), and in 12 conscious patients that brushed their teeth at least twice a day (group 2) was evaluated. Both groups were followed up for 7 days after hospitalization. Daily samples of saliva were obtained, dispersed and plated on selective culture media and colony forming units of each microbial group were obtained. RESULTS: For patients in group 1, the counts of total viable bacteria, staphylococci, Enterobacteriaceae and yeasts progressively increased in a time-dependant manner. For the conscious patients of group 2, there was no increase. CONCLUSION: It would appear that concomitant consciousness and brushing teeth are determinants in controlling the selected pneumopathogen counts in resting saliva. The increase in microbial counts in comatose patients is understandable because these microorganisms could spread to the lungs.

Cell surface hydrophobicity of Candida albicans isolated from elder patients undergoing denture-related candidosis.

BACKGROUND: The virulence potential of Candida albicans strains enrolled in denture-related candidosis still remains uncertain. Candida albicans cells with higher cell surface hydrophobicity (CSH) rates, so-called hydrophobic, present higher adhesion success in different host tissues than cells with lower rates, or even hydrophilic. OBJECTIVE: The proposition of this study was to evaluate the differences in the CSH of strains isolated from denture users with and without denture-related candidosis. MATERIAL AND METHODS: The strains were obtained from two paired groups of patients living a same retirement house. Fungal cells were submitted to CSH evaluation by the hydrocarbon partition test using xylene. RESULTS: The measures revealed that the yeasts from patients with candidosis had CSH values ranging from 4.52% to 12.24%, with an average of 8.22 +/- 2.92%. In the countergroup, the CSH ranged from 3.86% to 14.36%, with an average of 8.38 +/- 3.76%. The difference between the groups were considered not relevant (p = 0.997). CONCLUSION: The results let to the inference that natural populations of C. albicans from patients with and without clinical manifestation denture-related candidosis do not differ one from the other regarding to CSH.

Proposal of a low-cost protocol for colorimetric semi-quantification of secretory phospholipase by Candida albicans grown in planktonic and biofilm phases.

Biofilms are aggregates of microorganisms living in multilayered structures inside polymeric matrices onto surfaces. These biofilms may subvert the physiological properties of adjacent tissues causing morphofunctional failure. Many studies have shown that the expression of virulence attributes is maximized when microbes form such communities. This study evaluated the differential phospholipasic activity of Candida albicans SC5314 grown in planktonic phase and in biofilm. We propose two distinct protocols for the colorimetric evaluation of phosphatidylcholine hydrolysis in neutral and acidic conditions. The results showed that both protocols are suitable for the proposed intention and that 72 h-old planktonic cultures of C. albicans SC5314 secrete higher quantities of neutral (6.42-fold) and acidic (3.85-fold) phospholipases than biofilms.

Phenotypic evaluation of the effect of anaerobiosis on some virulence attributes of Candida albicans.

The current assumption that Candida albicans is a facultatively anaerobic organism has been widely accepted since its recovery from anoxic sites became common. However, the link between anaerobiosis and virulence remains uncertain. This study investigated the differential cell-surface hydrophobicity (CSH) using a hydrocarbon/water partition technique and analysed the differential secretion rates of secretory aspartyl proteases (Saps), esterase, chondroitinase and haemolysins of C. albicans strains recovered from periodontal pockets and non-periodontium-related intra-oral sites. For the enzymic tests, all strains from both sets were grown under aerobic and anaerobic conditions and the harvested cells were inoculated onto suitable normal or pre-reduced culture media in the presence or absence of molecular oxygen, respectively. The results showed that no variations were perceptible for CSH and chondroitinase (P>0.05). The secretion rates of esterase and haemolysins strongly decreased in an anoxic environment (P<0.0001). However, a consistent increment (P<0.0001) in Sap secretion was detected when cultures were grown under anaerobic conditions. Based on these results, it is suggested that the oxygen concentration in the atmosphere surrounding cells exerts a variable influence on the virulence attributes of C. albicans.

The role of candidal histolytic enzymes on denture-induced stomatitis in patients living in retirement homes.

MATERIAL AND METHODS: Fifty nine elders wearing complete dentures and living in retirement homes in Curitiba (southern Brazil), were divided into two groups: group #1, 26 patients with denture-induced stomatitis and group #2, 33 patients without denture-induced stomatitis. The two groups were evaluated in relation to the degree of denture-induced stomatitis, salivary fungal loads, and secretion of some histolytic enzymes. RESULTS: Patients from group #1 showed higher degrees of colonisation by Candida albicans (p = 0.031). Candida krusei, Candida tropicalis, and Candida parapsilosis were also isolated, but there were no differences between the groups (p > 0.05). Secretory aspartyl protease (Sap) and chondroitinase did not show significant differences among the isolated Candida spp. in the two groups. Phospholipase secretion rates were higher among the strains of C. albicans from group #2 (p = 0.036). The same behaviour was not detected for non-albicans Candida species. CONCLUSIONS: The results could infer that differences in the secretion rates of candidal histolytic enzymes should not be imputed as imperative for the progress of denture-induced stomatitis.

Candida albicans biofilms produce more secreted aspartyl protease than the planktonic cells.

By using a simple, low-cost system of polystyrene centrifuge tubes we compared the secreted aspartyl proteases (Saps) secretion during the biphasic growth modes of Candida albicans using twenty-one clinical isolates. Our results indicate that biofilms of C. albicans consistently secrete more Saps than their planktonic counterparts.

Concentrações inibitórias mínimas de amoxicilina em medicamento referência, genérico e similar / Minimum inhibitory concentration of different presentations of amoxicilin

Os estreptococos do grupo viridans são organismos comumente associados à endocardite após extrações dentais. Por isso, a profilaxia adotada antes desses procedimentos envolve o uso de penicilinas, em especial a amoxicilina. Foram determinadas as Concentrações Inibitórias Mínimas (MIC) de diferentes apresentações de amoxicilina (medicamento de referência, genérico e similar) frente cepas-padrão de estreptococos orais. Não foram detectadas diferenças estatísticas significativas na eficácia das diferentes categorias de amoxicilina e o antibiótico mostrou potência superior à preconizada pela legislação vigente.

Clonal characterization of Streptococcus mutans strains by multilocus enzyme electrophoresis

Vinte e uma cepas de Streptococcus mutans foram agrupadas pela eletroforese de enzimas codificadas por multilocus (MLEE). Seis isoenzimas apresentaram forte poder discriminatório (M1P, MPI, PLP, NSP, GOT e LAP). A MLEE é uma técnica robusta que pode ser empregada no estudo da diversidade clonal de cepas de S. mutans, em estudos epidemiológicos.

Genotypic diversity and virulence traits of Streptococcus mutans in caries-free and caries-active individuals.

The present study evaluated the relationship between clonal diversity and some virulence traits of Streptococcus mutans isolated from eight caries-free and eight caries-active subjects. A total of 155 S. mutans isolates from caries-free subjects and 144 isolates from caries-active subjects were obtained from samples of saliva, dental plaque and tongue surface and identified by PCR. The isolates were submitted to arbitrarily primed (AP)-PCR (OPA-2 and OPA-13) and multilocus enzyme electrophoresis (MLEE) to establish the genotypic diversity. Production of water-insoluble glucan (WIG) (monitored by SDS-PAGE), final pH of cultures and the ability of bacterial cells to adhere to smooth glass in the presence of sucrose were measured. High and comparable abilities of MLEE and AP-PCR were found to distinguish S. mutans genotypes, using Simpson's index of discrimination (0.971 and 0.968, respectively). The results showed a significant difference (P < 0.01) in the number of genotypes when caries-free and caries-active groups were compared by both fingerprinting methods used. Final pH (P = 0.32) and the percentage of adherence to a glass surface (P = 0.62) did not show differences between the two groups; however, the intensities of WIG bands from the caries-active group were greater than those from the caries-free group (P < 0.01). In addition, WIG was positively correlated with the ability of S. mutans to adhere to a glass surface (r = 0.34, P = 0.02) from caries-active subjects. These data showed that AP-PCR analysis and MLEE are both effective methods for assessing the genetic relatedness of S. mutans. Using these techniques, it was found that there is a larger number of genotypes of S. mutans with increased ability to synthesize WIG in caries-active individuals.

Açäo de amostras de Steptococcus mutans e Streptococcus sobrinus sobre diferentes carboidratos com ênfase na cárie dentária: estudo in vitro / Effect of Streptococcus mutans and Streptococcus sobrinus strains on different carbohydrates with emphasis on dental caries: in vitro study

A presente pesquisa tem como objetivo o estudo in vitro da produçäo de placa bacteriana e ácidos por amostras de S. mutans e S. sobrinus cultivadas isoladamente e em associaçäo, frente a diferentes substratos glicídicos. Para quantificaçäo da placa in vitro, foram utilizadas culturas recentes dessas espécies inoculadas, isoladas e em associaçäo, em tubos contendo meio BHI acrescido dos diferentes carboidrato (individualmente) - sacarose, glicose, frutose, glicose + frutose acompanhados de uma bastäo-capilar previamente pesado. Após incubação a 37ºC/48 horas (10 por cento CO²) os bastöes foram submetidos a nova passagem para obtençäo da produçäo de placa. A determinaçäo da produçäo de ácido pelas amostras de S. mutans e S. sobrinus - isoladas e em associaçäo - foi realizada através de mediçöes contínuas, em intervalos de tempo regulares, do pH do meio de cultura contendo os diferentes substratos. A sacarose apresentou valores estatisticamente superiores em relaçäo à produçäo de placa bacteriana quando comparada aos demais carboidratos analisados. A espécie S. mutans, isoladamente, e a associaçäo S. mutans / S. sobrinus produziram quantidades estatisticamente superiores de placa bacteriana in vitro quando cultivados em meio acrescido de 10 por cento de sacarose. As amostras de S. mutans e S. sobrinus, isoladas e em associaçäo, näo demonstraram um comportamento homogêneo para a produçäo de ácidos, apresentando momentos distintos de maior abaixamento do pH ao longo do período de incubaçäo