Objectives To compare three methods of intratubular contamination that simulate endodontic infections using confocal laser scanning microscopy (CLSM).Material and Methods Two pre-existing models of dentinal contamination were used to induce intratubular infection (groups A and B). These methods were modified in an attempt to improve the model (group C). Among the modifications it may be included: specimen contamination for five days, ultrasonic bath with BHI broth after specimen sterilization, use of E. faecalis during the exponential growth phase, greater concentration of inoculum, and two cycles of centrifugation on alternate days with changes of culture media. All specimens were longitudinally sectioned and stained with of LIVE/DEAD® for 20 min. Specimens were assessed using CLSM, which provided images of the depth of viable bacterial proliferation inside the dentinal tubules. Additionally, three examiners used scores to classify the CLSM images according to the following parameters: homogeneity, density, and depth of the bacterial contamination inside the dentinal tubules. Kruskal-Wallis and Dunn’s tests were used to evaluate the live and dead cells rates, and the scores obtained.Results The contamination scores revealed higher contamination levels in group C when compared with groups A and B (p<0.05). No differences were observed between group A and B (p>0.05). The volume of live cells in group C was higher than in groups A and B (p<0.05).Conclusion The new protocol for intratubular infection resulted in high and uniform patterns of bacterial contamination and higher cell viability in all specimens when compared with the current methods.

Objective The antimicrobial effect of ultrasonic agitation of calcium hydroxide (CH) pastes in infected bovine dentin and their penetrability were evaluated using confocal laser scanning microscopy (CLSM) and microbiological culture.Material and Methods Fifty-two bovine teeth were infected with Enterococcus faecalis using a new contamination protocol; then they received CH paste and were divided into groups with or without ultrasound. Ultrasonic agitation was conducted for 1 min with a plain point insert. After 15 d, the CLSM analyzed the viable and dead bacteria with Live and Dead assay. The dentinal wall debris was collected by burs, and the colony forming units (CFU/mL) were counted. The penetrability of the paste inside dentinal tubules was tested using the B-rodamine dye.Results The calcium hydroxide paste showed better results with the use of ultrasonic agitation (p<0.05).Conclusion The ultrasonic agitation of CH paste increased its antimicrobial action and was responsible for intradentinal penetration with the fulfilment of the tubules.

A Faculdade de Odontologia de Bauru, por proporcionar a estrutura para formar e transformar alunos em profissionais da melhor qualidade.Ao CNPQ, pelo apoio financeiro fundamental para a execução das pesquisas.A professora Flaviana Bombarda de Andrade, orientadora e amiga, que no exercício do seu dom me ensinou a pesquisar e me apaixonar pela microbiologia.Obrigada por todos os ensinamentos repassados, por sua paciência, atenção, vontade de ensinar e principalmente pela convivência, carinho e compreensão. Você como professora e sua história de vida em busca dos seus objetivos serão um exemplo para minha vida e carreira.Ao professor Marco Antonio Hungaro Duarte, por sua ajuda, atenção, boas ideias e, principalmente, pelo conhecimento compartilhado desde as conversas mais informais até as discussões durante as aulas. Obrigada por me permitir esta vivência.Aos professores Ivaldo Gomes de Moraes, Roberto Brandão, Clóvis Bramante e Norberti Bernadineli, Marco Antonio e Flaviana Andrade, pela hospitalidade e bom relacionamento com os alunos, por todo o conhecimento transmitido e pelas correções, sugestões, críticas sempre construtivas e, principalmente, pelos incentivos ao longo desses anos, objetivando o nosso crescimento profissional e pessoal. Agradeço imensamente pela oportunidade da experiência vivida e pelo privilégio de ter absorvido um pouco do conhecimento de todos vocês.Aos funcionários do departamento de Endodontia, Suelly, Andressa e Edi Mauro, por sua ajuda, atenção, gentileza e convivência.A Márcia Graeff, por fazer seu trabalho com amor e paciência. Obrigada por sua companhia, bom humor e por suportar a bagunça que fazíamos (eu e Amanda) na sua sala com a euforia pelas belas imagens durante os experimentos e inúmeros pilotos.Aos colegas da turma de mestrado, Amanda Maliza, Bruno Guimarães,

Objective: To assess the antimicrobial activity, solubility, setting time, flowability, pH, calcium release and surface characteristic of mineral trioxide aggregate (MTA) associated with different substances: 1% sodium hypochlorite gel, 2% chlorhexidine gel, K-Y gel, distilled water associated with 10% propylene glycol (CCPG), saline solution, and distilled water alone. Methods: The antimicrobial test included Streptococcus mutans, Lactobacillus casei, Enterococcus faecalis and Candida albicans, and the direct contact method was used. To assess setting time and flowability, ASTM 266/08 and ADA 57/2007 specifications were used. To assess pH and calcium release, the different types of material were inserted into retrograde cavities of acrylic resin teeth and immersed in ultrapure water for reading at different periods. The latter was performed with a pHmeter and an atomic absorption spectrophotometer. To assess the surface characteristics, acrylic teeth were analyzed under scanning electron micrograph (SEM). Results: The results yielded by the antimicrobial activity test revealed that additives associated with MTA were more inhibitory than bactericidal, with chlorhexidine achieving the best results. The K-Y vehicle made the additive more soluble than others. There were statistical differences amongst groups (p > 0.05) for flowability. The greatest setting time was observed in the CCPG group. The K-Y group presented the lowest pH and calcium release values in the 3-hour period. In the other periods, there was greater uniformity amongst groups. As regards the surface characteristics analysis, the CCPG group presented the greatest porosity (p < 0.05). Conclusions:Only the chlorhexidine gel brought some improvement to the antimicrobial effect, K-Y gel interfered in the physical-chemical properties of MTA and the addition of CCPG provided the greatest porosity.Keywords: Antimicrobial sensitivity test. Calcium release. Solubility. pH level. Physical-chemical properties. MTA.