Terapia fotodinâmica em esporos de bacillus atrophaeus e bacillus subtilis: estudos com LASER, LED, azul de metileno, rosa bengala e verde malaquita
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2012-07-30
Autores
Silva, Michelle Peneluppi [UNESP]
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Universidade Estadual Paulista (Unesp)
Resumo
Os esporos de Bacillus spp. são encontrados amplamente distribuídos na natureza, podendo ocasionar contaminação do meio ambiente e, eventualmente, doenças ao homem e animais. Em resposta a crescente resistência microbiana, a terapia fotodinâmica (PDT) surge para atuar como um tratamento alternativo e eficaz. O presente estudo teve como objetivo comparar e avaliar a ação exercida pelo LASER de baixa intensidade (vermelho visível) e pelo diodo emissor de luz verde (LED) em esporos de Bacillus atrophaeus e Bacillus subtilis na PDT, com o uso dos fotossensibilizadores azul de metileno (37,5 M), rosa bengala (12,5 M) e verde malaquita (300 M). Utilizou-se cepa padrão de Bacillus atrophaeus (ATCC 9372) e Bacillus subtilis (ATCC 19659). As cepas foram cultivadas, durante 7 dias, em Ágar Nutriente acrescidas de 0,003% de sulfato de manganês e analisadas quanto a formação de esporos (coloração de Wirtz-Conklin). Os esporos foram suspensos em água destilada esterilizada e centrifugados por 10 min a 653 Xg. As suspensões receberam choque térmico de 70 °C por 30 min. As suspensões foram padronizadas com 106 células/mL. Em placas de 96 poços adicionou-se 0,1 mL de suspensão dos esporos de Bacillus atrophaeus ou de Bacillus subtilis e 0,1 mL do fotossensibilizador ou de solução de NaCl a 0,9%, sendo agitadas durante 5, 10 e 30 minutos e irradiadas. Realizaram-se diluições seriadas. Alíquotas de 0,1 mL das diluições foram semeadas em placas com Ágar Infusão Cérebro-Coração e incubadas a 37 °C por 48 horas. Os resultados foram analisados estatisticamente (ANOVA, teste de Tukey, p<0,05). As maiores reduções observadas em UFC/mL (Log10) para os esporos de Bacillus atrophaeus foram 0,71 Log10 para azul de metileno (10 min); 2,49 Log10 para rosa bengala (30 min) e 0,42 Log10 para verde...
The spores of Bacillus spp. are found widely distributed in nature, which may cause contamination of the environment and eventually diseases to humans and animals. In response to increasing microbial resistance, photodynamic therapy (PDT) appears to act as an alternative treatment and effective. The present study aimed to compare and evaluate the action exerted by low intensity laser and green light emitting diode (LED) in spores of Bacillus atrophaeus and Bacillus subtilis in PDT, with the use of photosensitizers blue methylene (37.5 M), rose bengal (12.5 M) and malachite green (300 M). It was used a standard strain of Bacillus atrophaeus (ATCC 9372) and Bacillus subtilis (ATCC 19659). The strains were cultivated for 7 days in Nutrient Agar added to 0.003% of manganese sulphate and analyzed for the formation of spores (Wirtz-Conklin staining). The spores were suspended in sterile distilled water and centrifuged for 10 min at 653 Xg. The suspensions were heat shock at 70 °C for 30 min. The suspensions were standardized to 106 cells / mL. In 96-well plates were added 0.1 ml of Bacillus subtilis or Bacillus atrophaeus and 0.1 ml of the photosensitizer or NaCl 0.9%, stirred for 5, 10 and 30 minutes and irradiated. Serial dilutions were performed. Aliquots of 0.1 mL of the dilutions were plated on Brain Heart Infusion Agar and incubated at 37 °C for 48 hours. The results were analyzed statistically (ANOVA, Tukey test, p<0.05). The highest observed reductions in CFU/ml (Log10) for Bacillus atrophaeus were 0.71 Log10 for methylene blue (10 minutes), 2.49 Log10 for rose bengal (30 min) and 0.42 Log10 for malachite green (10 min). In relation to Bacillus subtilis the largest reductions were 0.82 Log10 for methylene blue (10 minutes), 3.86 Log10 for rose bengal (30 min) and 0.63 Log10 for malachite green (10 min). The conclusion... (Complete abstract click electronic access below)
The spores of Bacillus spp. are found widely distributed in nature, which may cause contamination of the environment and eventually diseases to humans and animals. In response to increasing microbial resistance, photodynamic therapy (PDT) appears to act as an alternative treatment and effective. The present study aimed to compare and evaluate the action exerted by low intensity laser and green light emitting diode (LED) in spores of Bacillus atrophaeus and Bacillus subtilis in PDT, with the use of photosensitizers blue methylene (37.5 M), rose bengal (12.5 M) and malachite green (300 M). It was used a standard strain of Bacillus atrophaeus (ATCC 9372) and Bacillus subtilis (ATCC 19659). The strains were cultivated for 7 days in Nutrient Agar added to 0.003% of manganese sulphate and analyzed for the formation of spores (Wirtz-Conklin staining). The spores were suspended in sterile distilled water and centrifuged for 10 min at 653 Xg. The suspensions were heat shock at 70 °C for 30 min. The suspensions were standardized to 106 cells / mL. In 96-well plates were added 0.1 ml of Bacillus subtilis or Bacillus atrophaeus and 0.1 ml of the photosensitizer or NaCl 0.9%, stirred for 5, 10 and 30 minutes and irradiated. Serial dilutions were performed. Aliquots of 0.1 mL of the dilutions were plated on Brain Heart Infusion Agar and incubated at 37 °C for 48 hours. The results were analyzed statistically (ANOVA, Tukey test, p<0.05). The highest observed reductions in CFU/ml (Log10) for Bacillus atrophaeus were 0.71 Log10 for methylene blue (10 minutes), 2.49 Log10 for rose bengal (30 min) and 0.42 Log10 for malachite green (10 min). In relation to Bacillus subtilis the largest reductions were 0.82 Log10 for methylene blue (10 minutes), 3.86 Log10 for rose bengal (30 min) and 0.63 Log10 for malachite green (10 min). The conclusion... (Complete abstract click electronic access below)
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Fotossintese, Esporos bacterianos, Bacillus subtilis
Como citar
SILVA, Michelle Peneluppi. Terapia fotodinâmica em esporos de bacillus atrophaeus e bacillus subtilis: estudos com LASER, LED, azul de metileno, rosa bengala e verde malaquita. 2012. 104 f. Dissertação (mestrado) – Universidade Estadual Paulista, Faculdade de Odontologia de São José dos Campos, 2012.