IADR Abstract Archives
Reuterin Derived Lactobacillus-reuteri Antibiofilm Activitiy towards Clinical Isolate Porphyromonas-gingivalis
Objectives: To investigate the effect of reuterin derived Lactobacillus-reuteri Indonesian strain towards clinically isolated Porphyromonas-gingivalis biofilm.
Methods: P.gingivalis was isolated from subgingival plaque samples of fixed orthodontic patients. P.gingivalis was cultured in blood agar, at 37°C for 24-h, anaerob condition (CO2,H,N) and confirmed with PCR with 16s rDNA primers specific gene. Primers were used in this study: 5'-AGGCAGCTTGCCATACTGCG-3' (forward) and 5'-ACTGTTAGCAACTACCGATGT-3' (revese). L.reuteri BEA 230424 (Indonesian strain) was cultured in de Mann Rogosa Sharpe (MRS) agar, 37°C, 48-h, anaerob condition. To produce reuterin, L.reuteri (1.5×1010CFU/ml) were suspended in 5ml glycerol (300mM) in MRS broth and incubated for 3-h, anaerobic conditions at 37°C. Subsequently, the supernatant was collected by centrifugation and then filtered (0.22μm). Reuterin effect on biofilm was evaluated by biofilm assay. A dose-dependent study using five reuterin concentrations (100%,50%,25%,12.5% and 6.25%) were performed against biofilms in a timescale of 15min,1h, 3h, 6h, and 24h. Chlorhexidine (0.2%) and biofilm wells without reuterin was used as positive and negative control respectively. Biofilm was assessed using crystal-violet dye with microplate-spectrophotometric (600nm). Data were statistically analyzed using one-way analysis of variance (ANOVA), with p<0.05 set as the level of significance.
Results: Study showed a 405-bp PCR result for P.gingivalis which confirmed the bacterial strain. Result showed a significant reduction of the biofilm formation of P.gingivalis after treatment with reuterin at all concentration and incubation times (p<0.05). The effective concentrations in reducing biofilm formation were found to be reuterin 50% (OD600 nm=0.268±0.091) in 6h incubation time compared to negative control (OD600 nm=2.537±0.559) with 89.4% biofilm reduction.
Conclusions: Reuterin proved to have inhibitory ability against P.gingivalis biofilm as periodontitis-associated bacteria. Therefore, reuterin isolated probiotic L.reuteri may have the capability to reduce the risk of periodontitis. However, further studies are needed to confirm the effect of reuterin isolated probiotic L.reuteri against periodontal pathogen in-vivo.
Methods: P.gingivalis was isolated from subgingival plaque samples of fixed orthodontic patients. P.gingivalis was cultured in blood agar, at 37°C for 24-h, anaerob condition (CO2,H,N) and confirmed with PCR with 16s rDNA primers specific gene. Primers were used in this study: 5'-AGGCAGCTTGCCATACTGCG-3' (forward) and 5'-ACTGTTAGCAACTACCGATGT-3' (revese). L.reuteri BEA 230424 (Indonesian strain) was cultured in de Mann Rogosa Sharpe (MRS) agar, 37°C, 48-h, anaerob condition. To produce reuterin, L.reuteri (1.5×1010CFU/ml) were suspended in 5ml glycerol (300mM) in MRS broth and incubated for 3-h, anaerobic conditions at 37°C. Subsequently, the supernatant was collected by centrifugation and then filtered (0.22μm). Reuterin effect on biofilm was evaluated by biofilm assay. A dose-dependent study using five reuterin concentrations (100%,50%,25%,12.5% and 6.25%) were performed against biofilms in a timescale of 15min,1h, 3h, 6h, and 24h. Chlorhexidine (0.2%) and biofilm wells without reuterin was used as positive and negative control respectively. Biofilm was assessed using crystal-violet dye with microplate-spectrophotometric (600nm). Data were statistically analyzed using one-way analysis of variance (ANOVA), with p<0.05 set as the level of significance.
Results: Study showed a 405-bp PCR result for P.gingivalis which confirmed the bacterial strain. Result showed a significant reduction of the biofilm formation of P.gingivalis after treatment with reuterin at all concentration and incubation times (p<0.05). The effective concentrations in reducing biofilm formation were found to be reuterin 50% (OD600 nm=0.268±0.091) in 6h incubation time compared to negative control (OD600 nm=2.537±0.559) with 89.4% biofilm reduction.
Conclusions: Reuterin proved to have inhibitory ability against P.gingivalis biofilm as periodontitis-associated bacteria. Therefore, reuterin isolated probiotic L.reuteri may have the capability to reduce the risk of periodontitis. However, further studies are needed to confirm the effect of reuterin isolated probiotic L.reuteri against periodontal pathogen in-vivo.
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