Enterococcus faecalis: fatores de virulência relacionados aos processos de natureza endodôntica

O objetivo principal deste estudo foi investigar a interação de 24 cepas de E. faecalis isoladas de infecções endodônticas primárias às proteínas de matriz dentinária, como também a moléculas de matriz presentes em lesões de endocardites. A análise desta interação foi feita através de técnica enzimática, com confirmação pela técnica de fluorescência. Além disto, foi realizada a confirmação do isolamento da espécie E. faecalis, através da técnica de PCR para o gene 16SrRNA e a análise da presença de genes de virulência da referida espécie microbiana para aderência às supostas proteínas de matriz incluindo às de ligação ao colágeno (ace, gelE, esp, agg e efaA). O maior padrão de interação das cepas ocorreu com a fibronectina (83,4%), seguido pelo fibrinogênio (62,5%) e colágeno humano tipo I (52%). Curiosamente, a aderência observada para o colágeno do tipo I, foi de pequena magnitude, quando comparado com a amostra padrão da ATCC 29212. As cepas ATCC 29212, A1, A43 e A68 interagiram com todas as proteínas de matriz utilizadas neste estudo. Um percentual expressivo das cepas testadas apresentou amplificação para efaA (86,9%) e para ace (73,9%). Paralelamente, todas as cepas apresentaram amplificação para gelE e foram negativas para os genes agg e esp. Adicionalmente, não houve correlação entre a detecção dos genes de virulência e a interação às proteínas de matriz, evidenciando que, mesmo com a detecção dos genes nas amostras, se faz necessário avaliar a expressão gênica por qPCR.

Effectiveness of 2% peracetic acid for the disinfection of gutta-percha cones

The aim of this study was to evaluate the effectiveness of 2% peracetic acid for the disinfection of gutta-percha cones contaminated in vitro with Escherichia coli, Staphylococcus aureus, Streptococcus mutans, Candida albicans and Bacillus subtilus (in spore form). Two hundred and twenty-five gutta-percha cones were contaminated with standardized suspensions of each microorganism and incubated at 37 degrees C for 24 h. The cones were divided into 10 experimental groups (n = 15), according to the microorganism tested and disinfection testing times. The disinfection procedure consisted of immersing each cone in a plastic tube containing the substance. The specimens remained in contact with the substance for 1 or 2.5 minutes. Afterwards, each cone was transferred to a 10% sodium thiosulphate solution (Na2S2O3) to neutralize the disinfectant. Microbial biofilms adhering to the cones were dispersed by agitation. Aliquots of 0.1 ml of the suspensions obtained were plated on Sabouraud dextrose agar, or brain and heart infusion agar, and incubated at 37 degrees C for 24 h. The results were expressed in colony forming units (CFU/ml) and the data were submitted to the Wilcoxon Signed Rank Test (level of significance at 0.05). A significant reduction was observed, after 1 minute of exposure, in the test solution for C. albicans (p = 0.0190), S. aureus (p = 0.0001), S. mutans (p = 0.0001), B. subtilis (p = 0.0001), and E. coli (p = 0.0001). After 2.5 minutes of exposure, 100% of the microbial inocula were eliminated. It was concluded that the 2% peracetic acid solution was effective against the biofilms of the tested microorganisms on gutta-percha cones at 1 minute of exposure.

Effect of rotary instrumentation and of the association of calcium hydroxide and chlorhexidine on the antisepsis of the root canal system in dogs

This study aimed at evaluating the antisepsis of the root canal system (RCS) and periapical region (PR) provided by rotary instrumentation associated with chlorhexidine + calcium hydroxide as intracanal medicament. Chronic periapical lesions were induced in 26 pre-molar roots in two dogs. After microbiological sampling, automatic instrumentation using the Profile system and irrigation with 5.25% sodium hypochlorite solution, with a final rinse of 14.3% EDTA followed by profuse irrigation with physiological saline were carried out in 18 root canals. After drying the canals, a paste based on calcium hydroxide associated with a 2% chlorhexidine digluconate solution was placed inside them. After 21 days, the medication was removed, leaving the root canals empty and coronally sealed. After 96 hours, a final microbiological sample was obtained, followed by histomicrobiological processing by the Brown & Brenn method. Eight untreated root canals represented the control group (C-G). Based on the Mann-Whitney test at a confidence level of 5% (p < 0.05), the procedures of antisepsis used offered significant efficacy (p < 0.05) resulting in 100.0% of the canals free of microorganisms. In the C-G, an elevated incidence of various microbial morphotypes was confirmed in all sites of the RCS, with the presence of microbial colonies in the periapical region. In contrast, the experimental group showed a similar pattern of infection in the RCS, although less intense and a reduced level of periapical infection (p < 0.05). It was concluded that adequate instrumentation followed by the application of calcium hydroxide + chlorhexidine offered significant elimination of microorganisms.

In vivo microbiological evaluation of the effect of biomechanical preparation of root canals using different irrigating solutions

The aim of this study was to evaluate the antimicrobial effect of biomechanical preparation using different irrigating solutions. Seventy-eight root canals from premolars of four dogs were used. After experimental induction of periapical lesions, the root canals were prepared using the following solutions for irrigation: Group 1) 2.5% sodium hypochlorite (NaOCl); Group 2) 2% chlorhexidine (CHX); Group 3) saline solution and Group 4) control group with no biomechanical preparation. The microbiological evaluation of the root canals was performed by counting the colony forming units (CFUs) using different culture mediums. Two absorbent paper cones were used in each root canal in order to collect the microbiological samples before, and thirty days after the biomechanical preparation. The culture plates were incubated in aerobic, anaerobic and microaerophilic environment. Statistical evaluation was carried out using analysis of variance, Tukey and Student tests. The results demonstrated that there was reduction in the number of microorganisms in the NaOCl and CHX groups (p<0.05). There was greater effectiveness in the chlorhexidine group. The group that used saline solution and the control group presented an increased number of microorganisms. It can be concluded that the use of antimicrobial irrigating solutions during biomechanical preparation promotes the reduction of endodontic microbiota. However, a considerable number of microorganisms were still observed.

Desinfeção aplicada aos cones de guta-percha

Introdução: Ao longo do tempo o Tratamento Endodôntico Não Cirúrgico tem sido das áreas da Medicina Dentária que mais tem evoluído. Todos os passos do tratamento têm sido revistos de forma a aumentar a taxa de sucesso. O controlo microbiológico é crucial para que o tratamento seja um sucesso a curto, médio e longo prazo. A assepsia deve ser mantida em todas as fases deste tratamento para que este seja um sucesso. Objetivo: Ao longo do meu percurso académico pude concluir que a fase da descontaminação dos cones, aquando a obturação (fase final do Tratamento Endodôntico Não Cirúrgico) era desvalorizada, o que me levou a efetuar uma revisão bibliográfica de modo a poder melhorar os meus conhecimentos e técnica. Material e Métodos: Para a elaboração deste trabalho foi realizada uma pesquisa bibliográfica recorrendo aos seguintes motores de busca: B-on, PubMed, Scielo e ScienceDirect, com as seguintes palavras-chave: “decontamination in endodontics”;” disinfection in endodontics”; “root canal irrigants”; “endodontics microbiology”; “Candida albicans“; “Enterococcus faecalis”; “sodium hypochlorite ”; “alcohol”; “contamination during Obturation”; “clorohexidine”; “filling materials endodontics”; “termoplastic gutta-percha”; “obturation material”; “Mineral Trioxide Aggregate”; “resilon”; “resin cement”; “resin material for root canal obturation”; “resin sealer”; “root canal”; “root canal sealing”; “root canal filling materials”; “condensation in endodontics”; “lateral condensation”; “gutta-percha”; “microlekeage”; “system B”; “fluid filtration model”;“dye penetration”. Como critério de inclusão estabeleceu-se que os artigos deveriam ser em Português, Inglês ou Espanhol e publicados entre 1995 e 2015. Dos resultados apresentados foram utilizados 110 artigos, pesquisados entre Maio de 2015 e 20 de Outubro de 2015. Foram ainda consultados livros de referência nestes mesmos locais. Conclusão: a presença de bactérias e os seus subprodutos no sistema tridimensional de canais está diretamente implicado com o insucesso do Tratamento Endodôntico. A descontaminação dos cones de guta-percha, é, portanto, um processo importante no Tratamento Endodôntico pois impede que os cones sejam colocados nos canais radiculares, estando contaminados por microorganismos que inviabilizam o tratamento efetuado. A submersão dos cones durante um minuto em clorohexidina a 2% ou hipoclorito a 5,25% está indicado e comprovado como um processo eficiente de desinfeção dos cones.

Endodontics pastes formulated with copaiba oil: action on oral microbiota and dentin bridge formation in dogs

This study analysed the effect of pastes formulated with calcium hydroxide P.A. and different vehicles (saline solution - paste A and Copaifera langsdorffii Desfon oil - paste B) on oral microorganisms and dentin bridge formation in dogs. The antimicrobial action of the pastes and their components was analysed by the minimum inhibitory concentration in agar gel technique. The components were diluted and tested on fifteen standard strains of microorganisms associated with endodontic diseases. The microorganisms were cultivated and after incubation data was analysed using One-Way ANOVA and Turkey's test (P≤0.05). Four superior incisors of ten animals were used to evaluate dentin bridge formation. Two incisors were capped with paste A (GA) and two with paste B (GB). After 90 days, the teeth were extracted for histological analysis and the degree of dentin bridge formation evaluated. Data was analysed by the Kruskal-Wallis test (P<0.05). The pastes and their components were classified in the following decreasing order of antimicrobial action: calcium hydroxide P.A., paste A, paste B and Copaifera langsdorffii Desfon oil. Calcium hydroxide P.A. showed significantly higher antimicrobial action than the pastes or their vehicles. No significant difference was observed between the two pastes in dentin bridge formation. Based on the microorganisms studied, it can be concluded that the pastes analysed showed similar antimicrobial potential but differed significantly from their individual components. No significant difference was observed in dentin bridge formation between the different pastes tested.

Water microbiology

Microbiology is the science devoted lo the study of organisms that are too small to be seen by the naked eye. These microorganisms are a large and diverse group of free-living forms that exist as single cells or cell clusters. Being free-living, microbial cells are distinct from the cells of animals and plants as the latter are not able to live alone in nature but only in characteristic groups. A single microbial cell, generally, is able to carry out its life processes of growth, respiration and reproduction independently of other cells, either of the same kind or of different kinds. There are five subdisciplines of microbiology: (a) the study of bacteria (bacteriology); (b) the study of viruses (virology); (c) the study of algae (phycology); (d) the study of fungi (mycology); and (e) the study of protozoa (protozoology). In the examination of the environment, all five areas of microbiology are studied. This becomes obvious when discussing the significance of each of these groups of organisms in relation to human health.

Airway cellularity, lipid laden macrophages and microbiology of gastric juice and airways in children with reflux oesophagitis

Background: Gastroesophageal reflux disease (GORD) can cause respiratory disease in children from recurrent aspiration of gastric contents. GORD can be defined in several ways and one of the most common method is presence of reflux oesophagitis. In children with GORD and respiratory disease, airway neutrophilia has been described. However, there are no prospective studies that have examined airway cellularity in children with GORD but without respiratory disease. The aims of the study were to compare (1) BAL cellularity and lipid laden macrophage index (LLMI) and, (2) microbiology of BAL and gastric juices of children with GORD (G+) to those without (G-). Methods: In 150 children aged <14-years, gastric aspirates and bronchoscopic airway lavage (BAL) were obtained during elective flexible upper endoscopy. GORD was defined as presence of reflux oesophagitis on distal oesophageal biopsies. Results: BAL neutrophil% in G- group (n = 63) was marginally but significantly higher than that in the G+ group (n = 77), (median of 7.5 and 5 respectively, p = 0.002). Lipid laden macrophage index (LLMI), BAL percentages of lymphocyte, eosinophil and macrophage were similar between groups. Viral studies were negative in all, bacterial cultures positive in 20.7% of BALs and in 5.3% of gastric aspirates. BAL cultures did not reflect gastric aspirate cultures in all but one child. Conclusion: In children without respiratory disease, GORD defined by presence of reflux oesophagitis, is not associated with BAL cellular profile or LLMI abnormality. Abnormal microbiology of the airways, when present, is not related to reflux oesophagitis and does not reflect that of gastric juices. © 2005 Chang et al; licensee BioMed Central Ltd.

Characterisation of the efficacy of endodontic medications using a three-dimensional fluorescent tooth model: An ex vivo study

The purpose of this study was to establish a three-dimensional fluorescent tooth model to investigate bacterial viability against intra-canal medicaments across the thickness and surface of root dentine. Dental microbial biofilms (Enterococcus faecalis and Streptococcus mutans) were established on the external root surface and bacterial kill was monitored over time against intra-canal medicament (Ca(OH)2 ) using fluorescent microscopy in conjunction with BacLight SYTO9 and propidium iodide stains. An Olympus digital camera fitted to SZX16 fluorescent microscope captured images of bacterial cells in biofilms on the external root surface. Viability of biofilm was measured by calculating the total pixel area of green (viable bacteria) and red (non-viable bacteria) for each image using ImageJ® software. All data generated were assessed for normality and then analysed using a Mann-Whitney t-test. The viability of S. mutans biofilm following Ca(OH)2 treatment showed a significant decline compared with the untreated group (P = 0.0418). No significant difference was seen for E. faecalis biofilm between the Ca(OH)2 and untreated groups indicating Ca(OH)2 medicament is ineffective against E. faecalis biofilm. This novel three-dimensional fluorescent biofilm model provides a new clinically relevant tool for testing of medicaments against dental biofilms.

20th annual meeting of the WEFTA Working Group on Analytical Methods in Fish and Fishery Products and 3rd annual meeting of the WEFTA Working Group on Microbiology. A retrospect on 20 years

After 20 annual meetings it is worth to have a look back and to see how it has started. There has been very little collaboration on research projects between member institutes under the auspices of WEFTA, co-operation in more neutral areas of common interest was developed at an early stage. The area which has proved very fruitful is methodology. It was agreed that probably the best way to make progress was to arrange meetings at each laboratory in turn where experienced, practising scientists could describe in detail how they carried out analyses. In this way, difficulties could be demonstrated or uncovered, and the accuracy, precision, efficiency and cost of the methods used in different laboratories could be compared.

Shining light on food microbiology; applications of luciferase-tagged microorganisms in the food industry

Bioluminescence is the production of light by living organisms as a result of a number of enzyme catalysed reactions caused by enzymes termed luciferases. The lux genes responsible for the emission of light can be cloned from one bioluminescent microorganism into one that is not bioluminescent. The light emitted can be monitored and quantified and will provide information on the metabolic activity, quantity and location of cells in a particular environment, in real-time. The primary aim of this thesis was to investigate and identify several food industry related applications of lux-tagged microorganisms. The first aim was to monitor a lux-tagged Cronobacter sakazakii in reconstituted infant milk formula, in realtime. The second aim was to investigate a bioluminescent-based early warning system for starter culture disruption by bacteriophages and antibiotic residues. The third of this thesis was to examine the use of a bioluminescent-based assay to test the activity of bioengineered Nisin derivatives M21V and S29A against foodborne pathogens in laboratory media and selected foods.