Interação entre bactérias cariogênicas
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Data
2016-03-21
Autores
Oliveira, Rosa Virginia Dutra de [UNESP]
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Universidade Estadual Paulista (Unesp)
Resumo
A proximidade entre micro-organismos presentes no biofilme dentário determina a existência de interações entre eles, que podem beneficiar ou antagonizar as espécies envolvidas. Os objetivos deste estudo foram: 1) apresentar uma revisão de literatura sobre interações entre espécies bacterianas cariogênicas no biofilme oral; 2) validar o uso do reator ‘‘drip-flow’’ (DFR) para desenvolver biofilmes dentários e testar agentes antimicrobianos; 3) avaliar o crescimento e a suscetibilidade à clorexidina de biofilmes compostos por Streptococccus mutans ATCC 25175 e Lactobacillus acidophilus ATCC 4356, usando o DFR; 4) avaliar o crescimento e a suscetibilidade à clorexidina de biofilmes compostos por Streptococccus mutans e Actinomyces naeslundii ATCC 12104, usando o DFR. Biofilmes cresceram sobre lâminas de vidro cobertas com hidroxiapatita e caldo BHI suplementado com sacarose 0.2% ou 0.5%, dependendo das espécies e um fluxo de 10 mL/h foi usado. O DFR foi incubado por 24 h a 37 °C / 5% CO2. Biofilmes foram tratados com clorexidina 0.2% (CHX) ou NaCl 0.9% por 2 min. Crescimento e efeito dos tratamentos foram determinados por contagem de unidades formadoras de colônias (UFC). Biofilmes foram marcados com o kit de Viabilidade Live/Dead e analisados por microscopia confocal de varredura a laser para diferenciar células com membranas íntegras das lesadas pela ação da CHX. A validação do DFR foi analisada por teste t (α=0.05). Avaliação da interação foi analisada por ANOVA dois fatores e teste de Tukey (α=0.05). A revisão de literatura mostrou o papel das interações microbianas no balanço competição/ coexistência. Na validação do DFR, resultados mostraram que, apesar da diferença inicial das concentrações de mono-culturas de S. mutans, o tratamento com clorexidina afetou ambos os biofilmes na mesma proporção. Não foi observada interação entre solução de tratamento e condição de cultura em biofilmes de S. mutans e L. acidophilus. Entretanto, a viabilidade foi significativamente reduzida após o tratamento com CHX. Mono-culturas de L. acidophilus cresceram significativamente menos que ambas mono- culturas e culturas mistas de S. mutans. S. mutans e A. naeslundii cresceram similarmente em ambas condições de cultura, dentro do grupo NaCl 0.9%. A viabilidade bacteriana foi significativamente reduzida em todos os grupos tratados com clorexidina, exceto para culturas mistas de S. mutans. Mono-culturas de A. naeslundii foram as mais suscetíveis, enquanto culturas mistas de S. mutans forma as menos suscetíveis à CHX. Em conclusão, relações entre os micro-organismos podem influenciar a ocorrência de cárie dentária. O presente estudo mostrou a aplicabilidade do DFR para crescer biofilmes orais e testar o uso de agentes antimicrobianos. Foram encontradas interações significantes entre S. mutans e A. naeslundii, mas não entre S. mutans e L. acidophilus.
The proximity between microorganisms present in dental biofilm determines the existence of interaction between them, which can benefit or antagonize the involved species. The aims of this study were: 1) to present a review about interactions among cariogenic bacterial species within oral biofilm; 2) to show the applicability of the drip flow reactor (DFR) for developing oral biofilms and testing antimicrobial agents; 3) to evaluate the growth and chlorhexidine susceptibility of biofilms comprised by Streptococcus mutans ATCC 25175 and Lactobacillus acidophilus ATCC 4356, using DFR; 3) to assess the growth and chlorhexidine susceptibility of biofilms comprised by Streptococcus mutans ATCC 25175 and Actinomyces naeslundii ATCC 12104, using DFR. Biofilms grew on hydroxyapatite coated glass slides, with BHI broth at 10 mL/h flow rate supplemented with 0.2 or 0.5% sucrose depending on the species used. DFR was incubated for 24 h at 37 °C / 5% CO2. Biofilms were treated with 0.2% chlorhexidine (CHX) or 0.9% NaCl for 2 min. Growth and effect of treatments were determined by colony forming units (CFU) counts. Biofilms were stained using the Live/Dead Viability kit and analyzed by confocal laser scanning microscopy (CLSM) to differentiate bacterial cells without damage and damaged by the action of CHX. DFR validation was analyzed by unpaired t test (α=0.05). Interaction evaluation was analyzed by two-way ANOVA and Tukey test (α=0.05). Literature review showed the role of microbial interactions in balancing competition/coexistence. For DFR validation, results showed that despite distinct initial concentrations of S. mutans mono-cultures, chlorhexidine treatment affected both biofilms at the same proportion. No interaction between treatment solution and culture condition was found in S. mutans and L. acidophilus biofilms. However, viability was significantly reduced by CHX treatment. L. acidophilus in mono-culture grew significantly less than S. mutans in either mono or mixed-culture. S. mutans and A. naeslundii grew similarly in both culture conditions within NaCl group. Bacterial viability was significantly reduced in all groups treated with chlorhexidine, except for S. mutans in mixed-cultures. A. naeslundii in mono-culture was the most susceptible group, whereas S. mutans in mixed-cultures was the least susceptible. In conclusion, relationships among microorganisms may influence the occurrence of dental caries. The present study showed the applicability of the DFR for growing oral biofilms and testing antimicrobial agents. Significant interactions were found between S. mutans and A. naeslundii but not between S. mutans and L. acidophilus.
The proximity between microorganisms present in dental biofilm determines the existence of interaction between them, which can benefit or antagonize the involved species. The aims of this study were: 1) to present a review about interactions among cariogenic bacterial species within oral biofilm; 2) to show the applicability of the drip flow reactor (DFR) for developing oral biofilms and testing antimicrobial agents; 3) to evaluate the growth and chlorhexidine susceptibility of biofilms comprised by Streptococcus mutans ATCC 25175 and Lactobacillus acidophilus ATCC 4356, using DFR; 3) to assess the growth and chlorhexidine susceptibility of biofilms comprised by Streptococcus mutans ATCC 25175 and Actinomyces naeslundii ATCC 12104, using DFR. Biofilms grew on hydroxyapatite coated glass slides, with BHI broth at 10 mL/h flow rate supplemented with 0.2 or 0.5% sucrose depending on the species used. DFR was incubated for 24 h at 37 °C / 5% CO2. Biofilms were treated with 0.2% chlorhexidine (CHX) or 0.9% NaCl for 2 min. Growth and effect of treatments were determined by colony forming units (CFU) counts. Biofilms were stained using the Live/Dead Viability kit and analyzed by confocal laser scanning microscopy (CLSM) to differentiate bacterial cells without damage and damaged by the action of CHX. DFR validation was analyzed by unpaired t test (α=0.05). Interaction evaluation was analyzed by two-way ANOVA and Tukey test (α=0.05). Literature review showed the role of microbial interactions in balancing competition/coexistence. For DFR validation, results showed that despite distinct initial concentrations of S. mutans mono-cultures, chlorhexidine treatment affected both biofilms at the same proportion. No interaction between treatment solution and culture condition was found in S. mutans and L. acidophilus biofilms. However, viability was significantly reduced by CHX treatment. L. acidophilus in mono-culture grew significantly less than S. mutans in either mono or mixed-culture. S. mutans and A. naeslundii grew similarly in both culture conditions within NaCl group. Bacterial viability was significantly reduced in all groups treated with chlorhexidine, except for S. mutans in mixed-cultures. A. naeslundii in mono-culture was the most susceptible group, whereas S. mutans in mixed-cultures was the least susceptible. In conclusion, relationships among microorganisms may influence the occurrence of dental caries. The present study showed the applicability of the DFR for growing oral biofilms and testing antimicrobial agents. Significant interactions were found between S. mutans and A. naeslundii but not between S. mutans and L. acidophilus.
Descrição
Palavras-chave
Streptococcus mutans, Lactobacillus acidophilus, Actinomyces, Clorexidina, Cárie dentária, Biofilmes, Streptococcus mutans, Lactobacillus acidophilus, Actinomyces, Chlorhexidine, Dental caries, Biofilms