Mechanistic Insight into the Anti-Bacterial/Anti-Biofilm Effects of Low Chlorhexidine Concentrations on <i>Enterococcus faecalis</i>—In Vitro Study
Background: Endodontic treatment failures are often linked to the persistence of <i>Enterococcus faecalis</i> in the root canal system. This study aimed to investigate the antibacterial/antibiofilm mechanism of chlorhexidine (CHX), particularly at low concentrations, against <i>E....
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2024-11-01
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| author | Nathanyel Sebbane Itzhak Abramovitz Nurit Kot-Limon Doron Steinberg |
| author_facet | Nathanyel Sebbane Itzhak Abramovitz Nurit Kot-Limon Doron Steinberg |
| author_sort | Nathanyel Sebbane |
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| description | Background: Endodontic treatment failures are often linked to the persistence of <i>Enterococcus faecalis</i> in the root canal system. This study aimed to investigate the antibacterial/antibiofilm mechanism of chlorhexidine (CHX), particularly at low concentrations, against <i>E. faecalis</i>, to improve endodontic treatment protocols. Methods: The antibacterial activity of CHX (0.125–20 μg/mL) was evaluated against <i>E. faecalis</i> ATCC 29212 using various assays, including planktonic growth inhibition, colony-forming units (CFUs), membrane permeability and potential assays, high-resolution scanning electron microscopy (HR-SEM), confocal laser scanning microscopy of biofilms, biomass and metabolic activity assays on matured biofilm, and quantitative real-time PCR for gene expression. Statistical analysis was performed using Student’s <i>t</i>-test and ANOVA. Results: CHX demonstrated concentration-dependent inhibition of <i>E. faecalis</i>, significantly reducing planktonic growth and CFUs. Membrane assays showed increased permeability and depolarization, indicating damage. HR-SEM revealed morphological changes, such as pore formation, while confocal microscopy showed a reduction in biofilm mass and extracellular substances. Gene expression analysis indicated the downregulation of virulence genes and upregulation of stress response genes. Conclusions: CHX at low concentrations disrupts <i>E. faecalis</i> at multiple levels, from membrane disruption to gene expression modulation, affecting mature biofilm. These findings support the refinement of endodontic disinfection protocols to reduce microbial persistence. |
| format | Article |
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| institution | Kabale University |
| issn | 2076-2607 |
| language | English |
| publishDate | 2024-11-01 |
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| spelling | doaj-art-41e5f9a9578f486e8c1dc176bb961cf12024-11-26T18:14:51ZengMDPI AGMicroorganisms2076-26072024-11-011211229710.3390/microorganisms12112297Mechanistic Insight into the Anti-Bacterial/Anti-Biofilm Effects of Low Chlorhexidine Concentrations on <i>Enterococcus faecalis</i>—In Vitro StudyNathanyel Sebbane0Itzhak Abramovitz1Nurit Kot-Limon2Doron Steinberg3Biofilm Research Laboratory, The Institute of Biomedical and Oral Research (IBOR), The Faculty of Dental Medicine, The Hebrew University of Jerusalem, Jerusalem 9112102, IsraelDepartment of Endodontics, The Faculty of Dental Medicine, The Hebrew University-Hadassah, Jerusalem 9112102, IsraelDepartment of Endodontics, The Faculty of Dental Medicine, The Hebrew University-Hadassah, Jerusalem 9112102, IsraelBiofilm Research Laboratory, The Institute of Biomedical and Oral Research (IBOR), The Faculty of Dental Medicine, The Hebrew University of Jerusalem, Jerusalem 9112102, IsraelBackground: Endodontic treatment failures are often linked to the persistence of <i>Enterococcus faecalis</i> in the root canal system. This study aimed to investigate the antibacterial/antibiofilm mechanism of chlorhexidine (CHX), particularly at low concentrations, against <i>E. faecalis</i>, to improve endodontic treatment protocols. Methods: The antibacterial activity of CHX (0.125–20 μg/mL) was evaluated against <i>E. faecalis</i> ATCC 29212 using various assays, including planktonic growth inhibition, colony-forming units (CFUs), membrane permeability and potential assays, high-resolution scanning electron microscopy (HR-SEM), confocal laser scanning microscopy of biofilms, biomass and metabolic activity assays on matured biofilm, and quantitative real-time PCR for gene expression. Statistical analysis was performed using Student’s <i>t</i>-test and ANOVA. Results: CHX demonstrated concentration-dependent inhibition of <i>E. faecalis</i>, significantly reducing planktonic growth and CFUs. Membrane assays showed increased permeability and depolarization, indicating damage. HR-SEM revealed morphological changes, such as pore formation, while confocal microscopy showed a reduction in biofilm mass and extracellular substances. Gene expression analysis indicated the downregulation of virulence genes and upregulation of stress response genes. Conclusions: CHX at low concentrations disrupts <i>E. faecalis</i> at multiple levels, from membrane disruption to gene expression modulation, affecting mature biofilm. These findings support the refinement of endodontic disinfection protocols to reduce microbial persistence.https://www.mdpi.com/2076-2607/12/11/2297<i>E. faecalis</i>chlorhexidineintracanal medicationendodontic treatmentbiofilm |
| spellingShingle | Nathanyel Sebbane Itzhak Abramovitz Nurit Kot-Limon Doron Steinberg Mechanistic Insight into the Anti-Bacterial/Anti-Biofilm Effects of Low Chlorhexidine Concentrations on <i>Enterococcus faecalis</i>—In Vitro Study Microorganisms <i>E. faecalis</i> chlorhexidine intracanal medication endodontic treatment biofilm |
| title | Mechanistic Insight into the Anti-Bacterial/Anti-Biofilm Effects of Low Chlorhexidine Concentrations on <i>Enterococcus faecalis</i>—In Vitro Study |
| title_full | Mechanistic Insight into the Anti-Bacterial/Anti-Biofilm Effects of Low Chlorhexidine Concentrations on <i>Enterococcus faecalis</i>—In Vitro Study |
| title_fullStr | Mechanistic Insight into the Anti-Bacterial/Anti-Biofilm Effects of Low Chlorhexidine Concentrations on <i>Enterococcus faecalis</i>—In Vitro Study |
| title_full_unstemmed | Mechanistic Insight into the Anti-Bacterial/Anti-Biofilm Effects of Low Chlorhexidine Concentrations on <i>Enterococcus faecalis</i>—In Vitro Study |
| title_short | Mechanistic Insight into the Anti-Bacterial/Anti-Biofilm Effects of Low Chlorhexidine Concentrations on <i>Enterococcus faecalis</i>—In Vitro Study |
| title_sort | mechanistic insight into the anti bacterial anti biofilm effects of low chlorhexidine concentrations on i enterococcus faecalis i in vitro study |
| topic | <i>E. faecalis</i> chlorhexidine intracanal medication endodontic treatment biofilm |
| url | https://www.mdpi.com/2076-2607/12/11/2297 |
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