Análise da virulência de biofilmes de Candida albicans desenvolvidos sob diferentes condições

Orientadores: Wander José da Silva, Altair Antoninha Del Bel CuryTese (doutorado) - Universidade Estadual de Campinas, Faculdade de Odontologia de PiracicabaResumo: A prevalência das infecções por Candida é elevada; logo, melhor compreensão dos mecanismos de desenvolvimento do biofilme é necessária para a redução da virulência e apropriado manejo clínico. Objetivou-se analisar a virulência de biofilmes de Candida albicans desenvolvidos sob diferentes condições. O papel das superfícies de biomateriais, da película salivar, e do estágio de desenvolvimento dos biofilmes foi avaliado no Capítulo 1. A influência da presença de outros microrganismos na virulência de C. albicans e na interação com o epitélio foi avaliada no Capítulo 2. O papel da atmosfera e da população bacteriana dos biofilmes foi investigado no Capítulo 3. No Capítulo 1, biofilmes de C. albicans foram desenvolvidos sobre discos de resina acrílica e titânio recobertos com película de saliva, ou de saliva com plasma. A superfície dos materiais foi analisada quanto a rugosidade e energia livre de superfície (ELS). Avaliou-se o número de microrganismos viáveis, a concentração de DNA, a atividade metabólica, a expressão de fatores de virulência e a estrutura dos biofilmes. Não houve diferenças quanto a rugosidade das superfícies. A película minimizou as diferenças entre a ELS dos materiais, sendo maior ELS observada para películas de saliva com plasma. O número de microrganismos viáveis, a concentração de DNA e a atividade metabólica aumentaram ao longo da maturação do biofilme. Maior atividade metabólica, maior expressão de ALS1, ALS3 e HWP1, e maior número de hifas foram verificados em biofilmes desenvolvidos na presença de película de saliva com plasma. Não houve diferenças entre os materiais. Concluiu-se que a presença de plasma na película salivar aumenta a virulência de C. albicans. No capítulo 2, biofilmes maduros (72 h) de C. albicans, e de C. albicans com bactérias, foram desenvolvidos sobre a superfície de resina acrílica. Esses biofilmes foram então analisados ou utilizados para infectar um modelo de epitélio oral humano reconstituído (RHOE). Avaliou-se o número de hifas e a expressão de fatores de virulência. A resposta epitelial foi avaliada por meio da expressão de IL-18 e Dectin1, da atividade da enzima lactato-desidrogenase (LDH), e a invasão epitelial. Maior número de hifas e maior expressão de HWP1, SAP4 e SAP6 foram verificados em biofilmes multi-espécie desenvolvidos sobre o acrílico. Biofilmes multi-espécies que infectaram o RHOE também apresentaram maior número de hifas e maior expressão de ALS3, EPA1, SAP6 e HWP1. Consequentemente, esses epitélios apresentaram maior invasão tecidual, maior atividade de LDH e maior expressão de IL-18. Concluiu-se que a presença de bactérias aumenta a virulência e patogenicidade de C. albicans. No capítulo 3, biofilmes de C. albicans e de C. albicans com estreptococos foram desenvolvidos sobre titânio em atmosfera de aerobiose e anaerobiose. Biofilmes de C. albicans (com ou sem estreptococos) também foram desenvolvidos na presença de Porphyromonas gingivalis, em anaerobiose. Avaliou-se a formação de hifas e a expressão de fatores de virulência. A atmosfera de anaerobiose e o co-cultivo com estreptococos geraram maior número de hifas e maior expressão de fatores de virulência. Embora P. gingivalis tenha inibido a virulência de C. albicans em biofilmes duo-espécies, esse efeito foi revertido na presença de estreptococos. Concluiu-se que a atmosfera de anaerobiose e a presença de estreptococos estimulam a virulência de C. albicansAbstract: Candida infections have high prevalence; therefore, better understanding of the development mechanisms of biofilms is required for the reduction of virulence and for more appropriated clinical management. The aim of this study was to analyse the virulence of Candida albicans biofilms developed under different conditions. The role of biomaterial surfaces, salivary pellicle, and stage of biofilm development has been reported in Chapter 1. The influence of the presence of other microorganisms in virulence of C. albicans and on the interaction with the epithelium was evaluated in Chapter 2. The role of the atmosphere and of the biofilm¿s bacterial population was investigated in Chapter 3. In Chapter 1, C. albicans biofilms were developed on acrylic resin and titanium discs coated with saliva pellicle, or saliva with blood plasma pellicle. The materials¿ surfaces were analysed with regards to surface roughness and surface free energy (SFE). We evaluated the number of viable organisms, the DNA concentration, the metabolic activity, the expression of virulence factors and the architecture of the biofilms. There were no differences in the surface roughness of materials. The pellicles eliminated the differences between SFE of materials. Higher SFE was observed for discs coated with pellicles of saliva with plasma. The number of viable organisms, the DNA concentration and the metabolic activity increased throughout the biofilm maturation. Higher metabolic activity, increased expression of ALS1, ALS3 and HWP1, and more hyphae were observed in biofilms grown in the presence of saliva with plasma pellicle. There were no differences between the materials. In conclusion, the presence of saliva with plasma pellicle increases the C. albicans virulence. In chapter 2, mature biofilms (72 h) of C. albicans, or C. albicans with bacteria, were developed on the surface of acrylic resin discs. These biofilms were then analysed or used to infect a reconstituted human oral epithelium model (RHOE). Biofilms were evaluated according to the number of hyphae and the expression of virulence factors. The epithelial response was evaluated by the expression of IL-18 and Dectin1, by the activity of lactate dehydrogenase enzyme (LDH), and with regards the epithelial invasion. Increased number of hyphae and increased expression of HWP1, SAP4 and SAP6 were observed in multi-species biofilms developed on acrylic. Multi-species biofilms that have infected RHOE also had a greater number of hyphae and increased expression of ALS3, EPA1, SAP6 and HWP1. Consequently, these epithelia showed greater tissue invasion, higher LDH activity and higher IL-18 expression. We concluded, therefore, that the presence of bacteria promotes the virulence and pathogenicity of C. albicans. In Chapter 3, biofilms of C. albicans and C. albicans with streptococci were developed on titanium, aerobically and anaerobically. C. albicans biofilms (mixed or not-mixed with streptococci) were also grown in the presence Porphyromonas gingivalis, anaerobically. Biofilms were evaluated according to the formation of hyphae and expression of virulence factors. Anaerobic conditions and co-cultivation of streptococci produced higher number of hyphae and increased expression of virulence factors. Although P. gingivalis has inhibited the virulence of C. albicans, this effect was reversed in the presence of streptococci. We concluded that the anaerobic atmosphere and the presence of streptococci stimulate the C. albicans virulenceDoutoradoProtese DentalDoutor em Clínica Odontológic

Maxillofacial trauma due to traffic accidents and falls:an exploratory study of associated factors

This study aimed to determine the pattern of Maxillofacial trauma (MFT) due to traffic accidents and falls in a reference hospital in a rural region of northeastern Brazil between December 2011 and December 2018 and to identify associated factors. This was a cross-sectional study using 585 medical records of patients with MFT. The data were subjected to a Poisson-Tweedie multiple regression analysis to estimate the Prevalence ratio (PR), with a 95% confidence interval (95% CI) and a significance level of p<0.05. MFT due to traffic accidents was more prevalent among patients 21 to 40 years old (PR=2.30; 95% CI=1.20-4.41; p<0.001) diagnosed with zygomatic-orbital complex fractures (PR=1.80; 95% CI=1.08-2.98; p=0.023). Falls were more frequent among older groups of 41 to 60 years (PR=1.83; 95% CI=1.09-3.06; p=0.022) and over 61 years (PR=2.23; 95% CI=1.09-3.06; p=0.022). In traffic accidents, alcohol consumption increased the length of stay (PR=2.081; 95% CI=1.553-2.787; p<0.001), and patients who did not use personal protective equipment (PPE) had higher hospital costs (PR=179.964; 95% CI=1.485-1.994; p<0.001) for this etiology. Traffic accidents and falls are two of the main etiologies of MFT, especially for males in the young adult age group (traffic accidents) and those above 41 years (falls). Alcohol consumption and the nonuse of PPE influenced the length of the hospital stay and hospital costs. Strategies to confront this problem, such as road and highway improvements, effective enforcement of laws and intersectoral coordination involving the entire community to implement policies and prevention programs targeted at these populations, can be implemented

Qualification of management and attention processes in the city of Caaporã, state of Paraíba, Brazil: mentoring reports on permanent education in health

The aim of this paper was to report the training on Permanent Education in Health (PEH) of Family's Health Teams (FHT) from the Muncipality of Caapor&#227;, State of Para&#237;ba. The problems identified involved the difficulties to access health services, fragility of the link with the community, and lack of educational and preventive actions. The proposed interventions encompass redirection of access strategies, integration between ESF and the community, and redistribution of tasks among FHT members. Results show the elimination of queue for care service, strengthening of the link with the community, and introduction of preventive measures in the FHT. The PEH in Caapor&#227; contributed to modify the management and health care practices.Objetivou-se relatar a forma&#231;&#227;o em Educa&#231;&#227;o Permanente em Sa&#250;de (EPS) de Equipes de Sa&#250;de da Fam&#237;lia (ESF) do munic&#237;pio de Caapor&#227; - PB. Os problemas identificados dizem respeito a dificuldades de acesso dos usu&#225;rios, fragilidade do v&#237;nculo com a comunidade e aus&#234;ncia de a&#231;&#245;es educativas e preventivas. As interven&#231;&#245;es propostas envolvem a reorienta&#231;&#227;o das estrat&#233;gias de acesso, integra&#231;&#227;o entre a ESF e comunidade e a redistribui&#231;&#227;o das tarefas entre os membros da ESF. Os resultados revelam a elimina&#231;&#227;o da fila de espera para o atendimento, o fortalecimento do v&#237;nculo com a comunidade e a introdu&#231;&#227;o de a&#231;&#245;es preventivas na ESF. A EPS em Caapor&#227; contribuiu para modificar as pr&#225;ticas de gest&#227;o e aten&#231;&#227;o em sa&#250;de.17018

Acesso em saúde bucal no Brasil: análise das iniquidades e não acesso na perspectiva do usuário, segundo o Programa de Melhoria do Acesso e da Qualidade da Atenção Básica, 2014 e 2018

Objective: To investigate factors associated with non-access to oral health in Brazil. Methods: Cross-sectional study that analyzed data from the external evaluation of the National Program to Improve Access and Quality (2014 and 2018), using hierarchical multivariate logistic regression. Non-access category was defined as the situation in which user was unable to make an appointment with a dentist. Results: We analyzed data from 37.262 individuals (2014 sample) and 117.570 people (2018 sample). A greater chance of non-access was found for those who: live in more unequal municipalities and with less oral health coverage, whose travel time to the unit is more than 11 minutes; female, aged between 25 and 39 years and whose income was up to 1 minimum wage. Conclusion: Non-access was associated with municipal factors such as grater inequality; organizational factors such as less coverage and travel time to the unit; and individual factors such as sex, age and income.Objetivo: Investigar os fatores associados ao não acesso em saúde bucal no Brasil. Métodos: Estudo transversal, sobre dados da avaliação externa do Programa Nacional de Melhoria do Acesso e da Qualidade da Atenção Básica, em 2014 e 2018, mediante regressão logística multivariada hierarquizada. Definiu-se como ‘não acesso’ quando o usuário não consegue marcar consulta com cirurgião-dentista. Resultados: Analisou-se dados de 37.262 indivíduos do segundo ciclo (2014) e 117.570 do terceiro ciclo (2018). Maior chance de não acesso ocorreu para residentes em municípios mais desiguais e com menor cobertura de saúde bucal, deslocamento para a unidade de saúde superior a 11 minutos, sexo feminino, idade entre 25 e 39 anos e renda de até 1 salário mínimo. Conclusão: O não acesso associou-se a fatores municipais, como maior desigualdade; fatores organizacionais, como menor cobertura e tempo de deslocamento até a unidade; e fatores individuais, como sexo, idade e rend

PERFIL DE URGÊNCIAS ODONTOLÓGICAS NO ESTADO DA PARAÍBA: UMA ANÁLISE DO PMAQ-AB

Objetivo: Avaliar os fatores associados à procura por atendimentos de urgência odontológica na Atenção Primária à Saúde. Metodologia: Foi realizado estudo transversal analítico, a partir dos microdados do 2° ciclo de avaliação externa do Programa de Melhoria do Acesso e da Qualidade da Atenção Básica (PMAQ-AB) no estado da Paraíba. Dados de relato de casos de urgência odontológica (n=1.531) e da procura por atendimento após episódio de urgência (n=824) foram avaliados segundo características sociodemográficas (sexo, idade, estado civil, cor da pele, analfabetismo, renda e cadastro no bolsa família). Os dados foram analisados por meio de regressão logística multivariada, pela qual foram obtidas as medidas de razão de chances (OR) e intervalo de confiança (IC95%), considerando-se p0,05). Entre os indivíduos que relataram urgência odontológica, 57,9% informaram procurar atendimento odontológico após o episódio de urgência. A procura por atendimento odontológico em casos de urgência se deu preferencialmente por indivíduos com maior idade (OR=1,014, IC95%= 1,003-1,026). Conclusões: As urgências odontológicas foram mais frequentes entre indivíduos que não sabem ler, o que reflete uma iniquidade em saúde. A maior procura dos idosos por serviços odontológicos nos casos de urgência revela o acesso preferencial dessa população via SUS.

Antibacterial and Solubility Analysis of Experimental Phytotherapeutic Paste for Endodontic Treatment of Primary Teeth

Objective: To evaluate the antibacterial effect and the solubility of experimental root canal filling pastes containing the phytoconstituents terpineol and cinnamaldehyde. Material and Methods: Minimum Inhibitory Concentration (MIC) of each phytoconstituent was determined against Enterococcus faecalis. Five groups of antibiotic pastes based on zinc oxide were obtained by mixing: only terpineol, only cinnamaldehyde, terpineol and cinnamaldehyde combined, chlorhexidine (antibiotic control), and CTZ paste (control paste). Antibacterial activity was analyzed through direct contact test within 24 and 72 hours. Solubility was evaluated by spectrophotometry within 48 and 144 hours. Antibacterial activity data were analyzed descriptively, and solubility data was analyzed using ANOVA and Tukey tests (p&lt;0.05). Results: The MIC obtained for terpineol and cinnamaldehyde were, respectively, 2000 μg/mL and 500 μg/mL. After 24h, only the terpineol paste did not inhibit E. faecalis growth. After 72h, all groups inhibited E. faecalis growth. After 48h, the highest solubility was verified in the terpineol paste (p&lt;0.05), and no differences were detected among other groups (p&gt;0.05). After and 144h, highest solubility was observed in the terpineol paste (p&lt;0.05), followed by the CTZ paste (p&lt;0.05). No differences were detected for cinnamaldehyde, terpineol+cinnamaldehyde and chlorhexidine pastes (p&gt;0.05). Conclusion: Pastes containing cinnamaldehyde or terpineol+cinnamaldehyde showed antibacterial activity against E. faecalis similar to CTZ paste, with lower solubility

Antibacterial and Solubility Analysis of Experimental Phytotherapeutic Paste for Endodontic Treatment of Primary Teeth

Objective: To evaluate the antibacterial effect and the solubility of experimental root canal filling pastes containing the phytoconstituents terpineol and cinnamaldehyde. Material and Methods: Minimum Inhibitory Concentration (MIC) of each phytoconstituent was determined against Enterococcus faecalis. Five groups of antibiotic pastes based on zinc oxide were obtained by mixing: only terpineol, only cinnamaldehyde, terpineol and cinnamaldehyde combined, chlorhexidine (antibiotic control), and CTZ paste (control paste). Antibacterial activity was analyzed through direct contact test within 24 and 72 hours. Solubility was evaluated by spectrophotometry within 48 and 144 hours. Antibacterial activity data were analyzed descriptively, and solubility data was analyzed using ANOVA and Tukey tests (p&lt;0.05). Results: The MIC obtained for terpineol and cinnamaldehyde were, respectively, 2000 μg/mL and 500 μg/mL. After 24h, only the terpineol paste did not inhibit E. faecalis growth. After 72h, all groups inhibited E. faecalis growth. After 48h, the highest solubility was verified in the terpineol paste (p&lt;0.05), and no differences were detected among other groups (p&gt;0.05). After and 144h, highest solubility was observed in the terpineol paste (p&lt;0.05), followed by the CTZ paste (p&lt;0.05). No differences were detected for cinnamaldehyde, terpineol+cinnamaldehyde and chlorhexidine pastes (p&gt;0.05). Conclusion: Pastes containing cinnamaldehyde or terpineol+cinnamaldehyde showed antibacterial activity against E. faecalis similar to CTZ paste, with lower solubility

Salivary pellicles equalise surfaces' charges and modulate the virulence of Candida albicans biofilm

Numerous environmental factors influence the pathogenesis of Candida biofilms and an uderstanding of these is necessary for appropriate clinical management. Aims: To investigate the role of material type, pellicle and stage of biofilm development on the viability, bioactivity, virulence and structure of C. albicans biofilms. Methods: The surface roughness (SR) and surface free energy (SFE) of acrylic and titanium discs was measured. Pellicles of saliva, or saliva supplemented with plasma, were formed on acrylic and titanium discs. Candida albicans biofilms were then generated for 1.5 h, 24 h, 48 h and 72 h. The cell viability in biofilms was analysed by culture, whilst DNA concentration and the expression of Candida virulence genes (ALS1, ALS3 and HWP1) were evaluated using qPCR. Biofilm metabolic activity was determined using XTT reduction assay, and biofilm structure analysed by Scanning Electron Microscopy (SEM). Results: Whilst the SR of acrylic and titanium did not significantly differ, the saliva with plasma pellicle increased significantly the total SFE of both surface. The number of viable microorganisms and DNA concentration increased with biofilm development, not differing within materials and pellicles. Biofilms developed on saliva with plasma pellicle surfaces had significantly higher activity after 24 h and this was accompanied with higher expression of virulence genes at all periods. Conclusion: Induction of C. albicans virulence occurs with the presence of plasma proteins in pellicles, throughout biofilm growth. To mitigate such effects, reduction of increased plasmatic exudate, related to chronic inflammatory response, could aid the management of candidal biofilm-related infections

Salivary pellicles equalise surfaces’ charges and modulate the virulence of candida albicans biofilm

Numerous environmental factors influence the pathogenesis of Candida biofilms and an understanding of these is necessary for appropriate clinical management. Aims To investigate the role of material type, pellicle and stage of biofilm development on the viability, bioactivity, virulence and structure of C. albicans biofilms. Methods The surface roughness (SR) and surface free energy (SFE) of acrylic and titanium discs was measured. Pellicles of saliva, or saliva supplemented with plasma, were formed on acrylic and titanium discs. Candida albicans biofilms were then generated for 1.5 h, 24 h, 48 h and 72 h. The cell viability in biofilms was analysed by culture, whilst DNA concentration and the expression of Candida virulence genes (ALS1, ALS3 and HWP1) were evaluated using qPCR. Biofilm metabolic activity was determined using XTT reduction assay, and biofilm structure analysed by Scanning Electron Microscopy (SEM). Results Whilst the SR of acrylic and titanium did not significantly differ, the saliva with plasma pellicle increased significantly the total SFE of both surface. The number of viable microorganisms and DNA concentration increased with biofilm development, not differing within materials and pellicles. Biofilms developed on saliva with plasma pellicle surfaces had significantly higher activity after 24 h and this was accompanied with higher expression of virulence genes at all periods. Conclusion Induction of C. albicans virulence occurs with the presence of plasma proteins in pellicles, throughout biofilm growth. To mitigate such effects, reduction of increased plasmatic exudate, related to chronic inflammatory response, could aid the management of candidal biofilm-related infections66129140COORDENAÇÃO DE APERFEIÇOAMENTO DE PESSOAL DE NÍVEL SUPERIOR - CAPESFUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DE SÃO PAULO - FAPESP007355/2013-002012/07436-