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The gold standard for quantifying bacteria both in routine diagnostics and in research is plating followed by count of colony-forming units (CFU). But, manual CFU counting on plates is time-consuming and subjective. We evaluated fractal dimension as a new methodology for evaluating CFU. Twenty fragments of expanded polytetrafluoroethylene (ePTFE) synthetic vascular prosthesis and 20 silicone prostheses were embedded in bacterial suspensions and incubated. The prostheses were then sown in solid culture medium and incubated for 48 h. Petri dishes were photographed and analyzed by fractal dimension. There was correlation between the number of CFU in manual counting and the fractal dimension analysis (p = 0.0001). We demonstrated that fractal dimension is a useful method for microbiological analyses in researches. It makes CFU analysis easier and faster and can be used regardless of the culture medium.•Petri dishes with different bacterial colonies were photographed with a digital camera under natural light.•The images were binarized and analyzed with ImageJⓇ's "fractal dimension" tool.•Fractal dimension analysis showed to be a good tool for evaluating the amount of colony-forming unit.

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Antibacterial coatings have currently gained great importance in biomedical technology investigations. Because of the spatial arrangement of the film coatings, evaluation of antibacterial activity presents a new challenge regarding traditional bacterial counting methods. In this protocol, four clinically relevant pathogens, Salmonella typhimurium, Escherichia coli, Pseudomonas aeruginosa, and Staphylococcus aureus were incubated on titania mesostructured thin film coatings for 24 h. Then, cell viability was studied considering three methods: counting of the number of colony forming units (CFU), live/dead staining, and quantification of extracellular DNA in suspension. Firstly, bacterial count was determined by the standard plate-count technique. Secondly, bacteria membrane integrity was evaluated by utilization of two fluorescent dyes, which allow distinction between live (membrane intact) and dead (membrane disrupted) bacteria. Lastly, extracellular DNA was quantified by spectrophotometry. In this manner, the three aforementioned techniques enabled the study of bacterial viability by qualitative and quantitative analyses.

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Aim: To evaluate the effect of three natural antifungal agents combined with routine denture care on the treatment of DS, using a quantitative mycological culture analysis. Methods: Thirty denture wearers with denture stomatitis DS were treated using five substances: sterile distilled water (G1), nystatin oral suspension (G2), 20% alcoholic extract propolis (G3), Punica granatumLinné gel (G4), and Uncaria tomentosa gel (G5). The substances were used 3 times a day for 14 days. Quantitative mycological culture analysis of samples collected from the palatal mucosa was performed at three stages: before treatment (T0), after 14 days of treatment (T1), and 30 days after treatment completion (T2). Data were evaluated using Kruskal-Wallis and Friedman tests (p < 0.05). Results: Palatal mucosa intragroup analysis showed a significant reduction of Candida CFU/mL values for all groups at T1 compared to T0 (p < 0.05). However, they did not present statistical differences when comparing T1 and T2 (p > 0.05). The intergroup analysis demonstrated that there are no statistical differences, regardless of the evaluation time (p > 0.05). Conclusion:The natural products tested showed a satisfactory result on DS treatment, which proved to be equivalent to conventional topical therapy with nystatin and to treatment using only regular oral hygiene procedures.

Objetivo: Avaliar o efeito de três antifúngicos naturais combinados com o cuidado rotineiro com próteses dentárias no tratamento da EP, por meio de uma análise quantitativa de cultura micológica. Métodos: Trinta usuários de próteses dentárias com EP foram tratados com cinco substâncias: água destilada estéril (G1), suspensão oral de nistatina (G2), extrato alcoólico de própolis 20% (G3), gel Punica granatum L. (G4) e gel Uncaria tomentosa (G5). As substâncias foram utilizadas 3 vezes ao dia durante 14 dias. A análise micológica quantitativa das amostras coletadas da mucosa palatina foi realizada em três etapas: antes do tratamento (T0), após 14 dias do tratamento (T1) e 30 dias após o término do tratamento (T2). Os dados foram avaliados pelos testes de Kruskal-Wallis e Friedman (p < 0,05). Resultados: A análise intragrupo da mucosa palatina mostrou uma redução significativa dos valores de Candida UFC/mL para todos os grupos em T1 em comparação com T0 (p < 0,05). No entanto, não apresentaram diferenças estatísticas na comparação de T1 e T2 (p > 0,05). A análise intergrupos demonstrou que não há diferenças estatísticas, independentemente do tempo de avaliação (p > 0,05). Conclusão: Os produtos naturais testados apresentaram resultado satisfatório no tratamento da EP, sendo equivalente à terapia tópica convencional com nistatina e ao tratamento apenas com procedimentos rotineiros de higiene bucal.

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Objetivo: Determinar las propiedades antimicrobianas de la incorporación de nanopartículas de óxido de zinc y cobre en un adhesivo de grabado y lavado total sobre Streptococcus mutans en pacientes con restauraciones de resina compuesta confeccionadas con adhesivo cargado. Métodos: Estudio experimental, randomizado, la muestra estuvo conformada por 25 pacientes, de ambos sexos, pertenecientes al posgrado de Ortodoncia de la Facultad de Odontología de la Universidad de Chile, en los cuales se confirmó presencia de Streptococcus mutans en saliva. Se confeccionaron restauraciones de resina compuesta oclusales, en premolares superiores con indicación de exodoncia por el tratamiento de ortodoncia, con adhesivo cargado (cuya composición fue 5/0,2 por ciento ZnO y Cu, respectivamente) y control (sin presencia de nanopartículas en su composición), según el listado de aleatorización. Se tomaron muestras microbiológicas en tres tiempos con la técnica de la cubeta (antes, 1 semana y 4 semanas posterior a la confección de las restauraciones). Se obtuvieron, aislaron e identificaron colonias de Streptococcus mutans a partir de las muestras obtenidas. Se usó el test de Mann-Whitney mediante el paquete estadístico SPSS v.21 Resultados: El promedio del recuento de UFC de Streptococcus mutans en el grupo experimental fue mayor posterior a la confección de las restauraciones de resina compuesta. Los resultados de la identificación molecular por PCR demuestran la presencia de Streptococcus mutans en 20 de 25 muestras. Conclusiones: No existen diferencias en el recuento de Streptococcus mutans antes y después de la aplicación del adhesivo sobre las restauraciones de resina compuesta(AU)

Objective: To determine the antimicrobial properties of the incorporation of zinc and copper oxide nanoparticles in an etching and total wash adhesive on Streptococcus mutans in patients with composite resin restorations made with loaded adhesive. Methods: Experimental and randomized trial, the sample were 25 patients, of both sexes, belonging to the FOUCH Orthodontic postgraduate program, in whom the presence of Streptococcus mutans in saliva was confirmed. Occlusal composite resin restorations were made in upper premolars with indication of extraction by orthodontic treatment, with loaded adhesive (whose composition is 5 / 0.2% ZnO and Cu respectively) and control (without the presence of nanoparticles in their composition), according to the scrambling listing. Microbiological samples were taken in three stages with the cuvette technique (before, 1 week and 4 weeks after the restoration was made). Colonies of Streptococcus mutans were obtained, isolated and identified from the samples obtained. The statistical analysis used the SPSS v.21 software, the data was analyzed by Mann Whitney test Results: The average CFU count of Streptococcus mutans in the experimental group (adhesive modified with zinc oxide and copper nanoparticles) was higher after the fabrication of composite resin restorations. The results of molecular identification by PCR demonstrate the presence of Streptococcus mutans in 20 of 25 samples. Conclusions: There are no differences in the count of Streptococcus mutans before and after the application of the adhesive on the composite resin restorations(AU)

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A incerteza de medição representa o nível de confiança no resultado. Para a estimativa da incerteza de medição foi empregado o cálculo do desvio padrão da reprodutibilidade intralaboratorial de 48 ensaios de contagem de bactérias heterotróficas pela técnica da membrana filtrante com detecção por fluorescência pelo uso de substrato fluorogênico em amostras de água purificada contaminadas artificialmente entre 10 e 100 UFC/mL. O valor obtido, 1,3 x 10-3 (log10), indica que a técnica utilizada pode ser uma alternativa para a estimativa da incerteza de medição em ensaios microbiológicos quantitativos de contagem de bactérias heterotróficas em amostras de água purificada.(AU)

Measurement uncertainty represents the confidence level in the result. To estimate the expanded measurement uncertainty, the standard deviation of intra-laboratory reproducibility of 48 heterotrophic bacterial count assays by fluorescence detection by the use of fluorogenic substrate on artificially contaminated purified water samples between 10 and 100 CFU/mL was used. The value obtained, 1.3 x 10-3 (log10), indicates that the technique used can be an alternative to estimate measurement uncertainty in quantitative microbiological heterotrophic bacterial count assays in purified water samples using fluorogenic substrate.(AU)

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Objetivo: Avaliar a redução microbiana após antissepsia cirúrgica das mãos dos cirurgiões, realizada com preparação alcoólica, em diferentes tempos. Método: Estudo de prevalência, pragmático, de campo, realizado em hospital terciário do Brasil. Coletaram-se amostras microbiológicas das mãos de 54 cirurgiões após lavagem simples, para determinar a flora microbiana basal e, após a antissepsia cirúrgica alcoólica, para avaliar a redução microbiana imediata. Categorizaram-se os resultados da redução microbiana em redução leve (até 50% de redução da flora bacteriana), moderada (de 51 a 80%) e alta (acima de 80%). A pesquisa foi submetida e aprovada pelo Comitê de Ética e Pesquisa da instituição hospitalar privada, sede do estudo, e da instituição de ensino superior federal. Resultados: Nas técnicas realizadas em menos de 90 segundos, houve 80% de redução severa, 6,7% de redução moderada e 13,3% de redução leve. Nas técnicas desempenhadas em mais de 180 segundos, todas as amostras apresentaram redução de contagem bacteriana, o que não ocorreu em tempos menores de antissepsia. Conclusão: Quando a técnica e o tempo recomendados são seguidos, maior é a redução bacteriana, em comparação aos tempos menores.

Objective: To evaluate the microbial reduction after surgical hand antisepsis performed with alcohol solution at different application times among surgeons. Method: This is a pragmatic prevalence field study carried out in a Brazilian tertiary hospital. Microbiological samples were collected from the hands of 54 surgeons after simple washing to determine the baseline microbial flora and after surgical antisepsis with an alcohol solution to evaluate the immediate microbial reduction. We categorized the microbial reduction results as mild (up to 50% bacterial flora reduction), moderate (51 to 80%), and high (more than 80%). The research was submitted to and approved by the Research Ethics Committee of the private hospital (study site) and the federal institution of higher education. Results: Techniques performed in less than 90 seconds showed an 80% high reduction, 6.7% moderate reduction, and 13.3% mild reduction. In applications that lasted more than 180 seconds, all samples presented bacterial count reduction, which did not occur in shorter antisepsis times. Conclusion: When the recommended technique and time are followed, the bacterial reduction is greater compared to lower durations.

Objetivo: evaluar la reducción microbiana después de la antisepsia quirúrgica de las manos de los cirujanos, realizada con preparación alcohólica, en diferentes momentos. Método: Estudio pragmático de prevalencia de campo realizado en un hospital terciario de Brasil. Muestras microbiológicas recogidas de las manos de 54 cirujanos después de un simple lavado, para determinar la flora microbiana basal y después de la antisepsia quirúrgica alcohólica, para evaluar la reducción microbiana inmediata. Los resultados de la reducción microbiana se clasificaron como leves (hasta un 50% de reducción en la flora bacteriana), moderados (del 51 al 80%) y altos (más del 80%). La investigación fue presentada y aprobada por el Comité de Ética e Investigación de la institución del hospital privado, sede del estudio y de la institución federal de educación superior. Resultados: en las técnicas realizadas en menos de 90 segundos hubo una reducción severa del 80%; 6,7% de reducción moderada; 13,3% de ligera reducción. En las técnicas realizadas durante 180 segundos, todas las muestras presentaron una reducción en el recuento bacteriano, lo que no ocurrió en tiempos de antisepsia más cortos. Conclusión: Cuando se siguen la técnica y el tiempo recomendados, mayor es la reducción bacteriana, en comparación con los tiempos más cortos.

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A incerteza de medição representa o nível de confiança no resultado. Para a estimativa da incerteza de medição foi empregado o cálculo do desvio padrão da reprodutibilidade intralaboratorial de 48 ensaios de contagem de bactérias heterotróficas pela técnica da membrana filtrante com detecção por fluorescência pelo uso de substrato fluorogênico em amostras de água purificada contaminadas artificialmente entre 10 e 100 UFC/mL. O valor obtido, 1,3 x 10-3 (log10), indica que a técnica utilizada pode ser uma alternativa para a estimativa da incerteza de medição em ensaios microbiológicos quantitativos de contagem de bactérias heterotróficas em amostras de água purificada. (AU)

Measurement uncertainty represents the confidence level in the result. To estimate the expanded measurement uncertainty, the standard deviation of intra-laboratory reproducibility of 48 heterotrophic bacterial count assays by fluorescence detection by the use of fluorogenic substrate on artificially contaminated purified water samples between 10 and 100 CFU/mL was used. The value obtained, 1.3 x 10-3 (log10), indicates that the technique used can be an alternative to estimate measurement uncertainty in quantitative microbiological heterotrophic bacterial count assays in purified water samples using fluorogenic substrate. (AU)

Assuntos

RESUMO

A incerteza de medição representa o nível de confiança no resultado. Para a estimativa da incerteza de medição foi empregado o cálculo do desvio padrão da reprodutibilidade intralaboratorial de 48 ensaios de contagem de bactérias heterotróficas pela técnica da membrana filtrante com detecção por fluorescência pelo uso de substrato fluorogênico em amostras de água purificada contaminadas artificialmente entre 10 e 100 UFC/mL. O valor obtido, 1,3 x 10-3 (log10), indica que a técnica utilizada pode ser uma alternativa para a estimativa da incerteza de medição em ensaios microbiológicos quantitativos de contagem de bactérias heterotróficas em amostras de água purificada.

Measurement uncertainty represents the confidence level in the result. To estimate the expanded measurement uncertainty, the standard deviation of intra-laboratory reproducibility of 48 heterotrophic bacterial count assays by fluorescence detection by the use of fluorogenic substrate on artificially contaminated purified water samples between 10 and 100 CFU/mL was used. The value obtained, 1.3 x 10-3 (log10), indicates that the technique used can be an alternative to estimate measurement uncertainty in quantitative microbiological heterotrophic bacterial count assays in purified water samples using fluorogenic substrate.

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Pseudomonas aeruginosa is a human pathogen capable to form robust biofilms. P. aeruginosa biofilms represent a serious problem because of the adverse effects on human health and industry, from sanitary and economic points of view. Typical strategies to break down biofilms have been long used, such as the use of disinfectants or antibiotics, but also, according to their high resistance to standard antimicrobial approaches, alternative strategies employing photocatalysis or control of biofilm formation by modifying surfaces, have been proposed. Colony forming units (cfu) counting and live/dead staining, two classic techniques used for biofilm quantification, are detailed in this work. Both methods assess cell viability, a key factor to analyze the microbial susceptibility to given treatment, then, they represent a good approach for evaluation of an antibiofilm strategy.

RESUMO

Denture use may aggravate the occurrence of oral infections, considering it enhances microbial adherence. Aim: This study assessed the reduction of microbial loads of Candida albicans, Staphylococcus aureus, and Klebsiella oxytoca by disinfecting the polymethylmethacrylate (PMMA) of complete dentures with hydroalcoholic extract of Salvia officinalis. Additionally, the effect of such extract on the properties of PMMA was examined. Methods: Microorganisms were isolated from saliva samples collected from complete denture wearers. The hydroalcoholic extract of S. officinalis was produced according to the Brazilian Pharmacopoeia 5. The PMMA specimens (n=188) were immersed in microbial inoculum and incubated at 37°C for 16 hours per day. Then, they were subjected to a disinfection protocol for 30 days. The specimens were divided into five treatment groups: sterile saline solution (0.85%; control), 0.2% chlorhexidine digluconate, and hydroalcoholic extract of S. officinalis (0.2%, 0.8%, and 1.16%). Microorganism adherence to the PMMA surface was also assessed, as well as surface roughness (Ra in µm) and color stability of the PMMA (mean ΔE). Changes in microbial load and surface roughness after the disinfection protocol were verified with paired t-test. Substances at day 10, adherence, and color stability were compared by the Kruskall-Wallis and Mann-Whitney tests, and one-way ANOVA was used to compare substances at the beginning and end of the experiment (α=0.05). Results: The 1.16% S. officinalis extract significantly reduced the microbial load of all the microorganisms after 30 days of disinfection (p<0.05). The microbial load of K. oxytoca was also reduced at lower concentrations of the S. officinalis extract (0.2% and 0.8%) (p<0.02). Antimicrobial and anti-adherent effects against microorganisms isolated from the oral cavity were observed. There was no significant change in surface roughness (p>0.05) and color stability was significantly higher in the control group (p<0.0001). Conclusions: The hydroalcoholic extract of S. officinalis may be used as a disinfectant solution for dentures

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