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Comparative Analysis of Chlorhexidine Derivatives and Alternative Agents on Streptococcus mutans Viability, Biofilm Formation, and Gene Expression.

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Title: Comparative Analysis of Chlorhexidine Derivatives and Alternative Agents on Streptococcus mutans Viability, Biofilm Formation, and Gene Expression.
Authors: Kamalloo H; Department of Microbiology, Science and Research Branch, Islamic Azad University, Tehran, Iran., Moeininejad M; School of Dentistry, Tehran University of Medical Sciences, Tehran, Iran., Golshoui MK; Infectious Diseases and Tropical Medicine Research Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran., Samadyar F; Department of Microbiology, Faculty of Basic Science, Karaj Branch, Islamic Azad University Karaj, Karaj, Iran., Shivaee A; Infectious Diseases and Tropical Medicine Research Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran., Kalani BS; Department of Medical Microbiology, School of Medicine, Ilam University of Medical Sciences, Ilam, Iran.; Student Research Committee, Ilam University of Medical Sciences, Ilam, Iran.
Source: MicrobiologyOpen [Microbiologyopen] 2026 Aug; Vol. 15 (4), pp. e70360.
Publication Type: Journal Article; Comparative Study
Language: English
Journal Info: Publisher: Wiley-Blackwell Country of Publication: England NLM ID: 101588314 Publication Model: Print Cited Medium: Internet ISSN: 2045-8827 (Electronic) Linking ISSN: 20458827 NLM ISO Abbreviation: Microbiologyopen Subsets: MEDLINE
Imprint Name(s): Original Publication: Oxford : Wiley-Blackwell
MeSH Terms: Biofilms*/drug effects , Biofilms*/growth & development , Chlorhexidine*/pharmacology , Chlorhexidine*/analogs & derivatives , Streptococcus mutans*/drug effects , Streptococcus mutans*/physiology , Streptococcus mutans*/genetics , Microbial Viability*/drug effects , Anti-Bacterial Agents*/pharmacology , Gene Expression Regulation, Bacterial*/drug effects, Fibroblasts/drug effects ; Humans ; Microbial Sensitivity Tests ; Gene Expression Profiling ; Biguanides
Abstract: This study evaluates the antimicrobial and antibiofilm effects of chlorhexidine and related compounds against S. mutans, focusing on bacterial viability, biofilm formation, and expression of key biofilm genes. Eight compounds were tested for their chemical properties, MICs, and interactions via FIC analyses. Time-kill assays assessed bactericidal activity over 24 h, while cytotoxicity was evaluated in human gingival fibroblasts. Gene expression of biofilm-related genes was analyzed to determine transcriptional modulation. Chlorhexidine gluconate and alexidine showed the strongest antimicrobial activity, with MICs of 2 μg/mL against the reference strain and 4 μg/mL for clinical isolates. Time-kill results demonstrated bacterial reductions from ~7.2 to < 1.0 log CFU/mL in the reference strain but less effect in clinical isolates. Chlorhexidine reduced biofilm biomass by nearly 19% at 48 h, whereas amoxicillin increased biofilm formation by over 12%. Cytotoxicity varied, with proguanil showing high cell viability (90%), while chlorhexidine and alexidine reduced viability to 50-70%. Gene expression analyzes revealed significant downregulation of gtfD and brpA by chlorhexidine and analogs, contrasting with mild upregulation by amoxicillin. These findings highlight chlorhexidine compounds as effective anti-biofilm agents, emphasizing the need for continued exploration of alternatives and combination therapies to enhance dental caries treatment.
(© 2026 The Author(s). MicrobiologyOpen published by John Wiley & Sons Ltd.)
References: Int J Oral Sci. 2021 Sep 30;13(1):30. (PMID: 34588414)
Odontology. 2020 Apr;108(2):260-270. (PMID: 31624978)
Can J Microbiol. 2011 Jan;57(1):1-20. (PMID: 21217792)
Clin Oral Investig. 2025 Nov 7;29(12):555. (PMID: 41198897)
Trends Microbiol. 2018 Mar;26(3):229-242. (PMID: 29097091)
Microbiol Res. 2023 Oct 14;278:127526. (PMID: 39491258)
Int Dent J. 2022 Jun;72(3):269-277. (PMID: 35287956)
J Dent. 2020 Dec;103:103497. (PMID: 33075450)
BMC Oral Health. 2025 Mar 13;25(1):379. (PMID: 40082866)
Int J Dent. 2023 Jan 3;2023:9365845. (PMID: 36636170)
J Clin Periodontol. 2017 Mar;44 Suppl 18:S23-S38. (PMID: 28266108)
Curr Oral Health Rep. 2016 Mar;3(1):36-44. (PMID: 27134811)
Molecules. 2025 Apr 14;30(8):. (PMID: 40333726)
Microb Pathog. 2021 Jan;150:104669. (PMID: 33278519)
Cell Host Microbe. 2023 Apr 12;31(4):528-538. (PMID: 36933557)
FEMS Microbiol Lett. 2025 Jan 10;372:. (PMID: 40985596)
J Dent Res. 2021 Jun;100(6):631-638. (PMID: 33530836)
Microorganisms. 2024 Aug 07;12(8):. (PMID: 39203455)
Arch Oral Biol. 2021 Jul;127:105137. (PMID: 33965851)
J Am Dent Assoc. 2008 Feb;139(2):178-83. (PMID: 18245686)
Front Microbiol. 2019 Mar 22;10:587. (PMID: 30967854)
Pathog Dis. 2022 Jun 22;80(1):. (PMID: 35481543)
Trends Microbiol. 2025 Apr;33(4):459-471. (PMID: 39828459)
Acta Microbiol Immunol Hung. 2018 Jun 1;65(2):229-240. (PMID: 29685049)
Antimicrob Agents Chemother. 1993 May;37(5):1158-9. (PMID: 8517706)
Grant Information: Ilam University of Medical Sciences
Contributed Indexing: Keywords: Streptococcus mutans; antimicrobial activity; biofilm inhibition; chlorhexidine; virulence gene expression
Substance Nomenclature: R4KO0DY52L (Chlorhexidine)
0 (Anti-Bacterial Agents)
GVN71CAL3G (alexidine)
MOR84MUD8E (chlorhexidine gluconate)
0 (Biguanides)
Entry Date(s): Date Created: 20260715 Date Completed: 20260715 Latest Revision: 20260813
Update Code: 20260814
PubMed Central ID: PMC13369289
DOI: 10.1002/mbo3.70360
PMID: 42449561
Database: MEDLINE
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  Data: Comparative Analysis of Chlorhexidine Derivatives and Alternative Agents on Streptococcus mutans Viability, Biofilm Formation, and Gene Expression.
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– Name: Abstract
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  Data: This study evaluates the antimicrobial and antibiofilm effects of chlorhexidine and related compounds against S. mutans, focusing on bacterial viability, biofilm formation, and expression of key biofilm genes. Eight compounds were tested for their chemical properties, MICs, and interactions via FIC analyses. Time-kill assays assessed bactericidal activity over 24 h, while cytotoxicity was evaluated in human gingival fibroblasts. Gene expression of biofilm-related genes was analyzed to determine transcriptional modulation. Chlorhexidine gluconate and alexidine showed the strongest antimicrobial activity, with MICs of 2 μg/mL against the reference strain and 4 μg/mL for clinical isolates. Time-kill results demonstrated bacterial reductions from ~7.2 to &amp;lt; 1.0 log CFU/mL in the reference strain but less effect in clinical isolates. Chlorhexidine reduced biofilm biomass by nearly 19% at 48 h, whereas amoxicillin increased biofilm formation by over 12%. Cytotoxicity varied, with proguanil showing high cell viability (90%), while chlorhexidine and alexidine reduced viability to 50-70%. Gene expression analyzes revealed significant downregulation of gtfD and brpA by chlorhexidine and analogs, contrasting with mild upregulation by amoxicillin. These findings highlight chlorhexidine compounds as effective anti-biofilm agents, emphasizing the need for continued exploration of alternatives and combination therapies to enhance dental caries treatment.&lt;br /&gt; (&#169; 2026 The Author(s). MicrobiologyOpen published by John Wiley &amp; Sons Ltd.)
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  Data: Int J Oral Sci. 2021 Sep 30;13(1):30. (PMID: &lt;searchLink fieldCode=&quot;PM&quot; term=&quot;%2234588414%22&quot;&gt;34588414)&lt;/searchLink&gt;&lt;br /&gt;Odontology. 2020 Apr;108(2):260-270. (PMID: &lt;searchLink fieldCode=&quot;PM&quot; term=&quot;%2231624978%22&quot;&gt;31624978)&lt;/searchLink&gt;&lt;br /&gt;Can J Microbiol. 2011 Jan;57(1):1-20. (PMID: &lt;searchLink fieldCode=&quot;PM&quot; term=&quot;%2221217792%22&quot;&gt;21217792)&lt;/searchLink&gt;&lt;br /&gt;Clin Oral Investig. 2025 Nov 7;29(12):555. (PMID: &lt;searchLink fieldCode=&quot;PM&quot; term=&quot;%2241198897%22&quot;&gt;41198897)&lt;/searchLink&gt;&lt;br /&gt;Trends Microbiol. 2018 Mar;26(3):229-242. (PMID: &lt;searchLink fieldCode=&quot;PM&quot; term=&quot;%2229097091%22&quot;&gt;29097091)&lt;/searchLink&gt;&lt;br /&gt;Microbiol Res. 2023 Oct 14;278:127526. (PMID: &lt;searchLink fieldCode=&quot;PM&quot; term=&quot;%2239491258%22&quot;&gt;39491258)&lt;/searchLink&gt;&lt;br /&gt;Int Dent J. 2022 Jun;72(3):269-277. (PMID: &lt;searchLink fieldCode=&quot;PM&quot; term=&quot;%2235287956%22&quot;&gt;35287956)&lt;/searchLink&gt;&lt;br /&gt;J Dent. 2020 Dec;103:103497. (PMID: &lt;searchLink fieldCode=&quot;PM&quot; term=&quot;%2233075450%22&quot;&gt;33075450)&lt;/searchLink&gt;&lt;br /&gt;BMC Oral Health. 2025 Mar 13;25(1):379. (PMID: &lt;searchLink fieldCode=&quot;PM&quot; term=&quot;%2240082866%22&quot;&gt;40082866)&lt;/searchLink&gt;&lt;br /&gt;Int J Dent. 2023 Jan 3;2023:9365845. (PMID: &lt;searchLink fieldCode=&quot;PM&quot; term=&quot;%2236636170%22&quot;&gt;36636170)&lt;/searchLink&gt;&lt;br /&gt;J Clin Periodontol. 2017 Mar;44 Suppl 18:S23-S38. (PMID: &lt;searchLink fieldCode=&quot;PM&quot; term=&quot;%2228266108%22&quot;&gt;28266108)&lt;/searchLink&gt;&lt;br /&gt;Curr Oral Health Rep. 2016 Mar;3(1):36-44. (PMID: &lt;searchLink fieldCode=&quot;PM&quot; term=&quot;%2227134811%22&quot;&gt;27134811)&lt;/searchLink&gt;&lt;br /&gt;Molecules. 2025 Apr 14;30(8):. (PMID: &lt;searchLink fieldCode=&quot;PM&quot; term=&quot;%2240333726%22&quot;&gt;40333726)&lt;/searchLink&gt;&lt;br /&gt;Microb Pathog. 2021 Jan;150:104669. (PMID: &lt;searchLink fieldCode=&quot;PM&quot; term=&quot;%2233278519%22&quot;&gt;33278519)&lt;/searchLink&gt;&lt;br /&gt;Cell Host Microbe. 2023 Apr 12;31(4):528-538. (PMID: &lt;searchLink fieldCode=&quot;PM&quot; term=&quot;%2236933557%22&quot;&gt;36933557)&lt;/searchLink&gt;&lt;br /&gt;FEMS Microbiol Lett. 2025 Jan 10;372:. (PMID: &lt;searchLink fieldCode=&quot;PM&quot; term=&quot;%2240985596%22&quot;&gt;40985596)&lt;/searchLink&gt;&lt;br /&gt;J Dent Res. 2021 Jun;100(6):631-638. (PMID: &lt;searchLink fieldCode=&quot;PM&quot; term=&quot;%2233530836%22&quot;&gt;33530836)&lt;/searchLink&gt;&lt;br /&gt;Microorganisms. 2024 Aug 07;12(8):. (PMID: &lt;searchLink fieldCode=&quot;PM&quot; term=&quot;%2239203455%22&quot;&gt;39203455)&lt;/searchLink&gt;&lt;br /&gt;Arch Oral Biol. 2021 Jul;127:105137. (PMID: &lt;searchLink fieldCode=&quot;PM&quot; term=&quot;%2233965851%22&quot;&gt;33965851)&lt;/searchLink&gt;&lt;br /&gt;J Am Dent Assoc. 2008 Feb;139(2):178-83. (PMID: &lt;searchLink fieldCode=&quot;PM&quot; term=&quot;%2218245686%22&quot;&gt;18245686)&lt;/searchLink&gt;&lt;br /&gt;Front Microbiol. 2019 Mar 22;10:587. (PMID: &lt;searchLink fieldCode=&quot;PM&quot; term=&quot;%2230967854%22&quot;&gt;30967854)&lt;/searchLink&gt;&lt;br /&gt;Pathog Dis. 2022 Jun 22;80(1):. (PMID: &lt;searchLink fieldCode=&quot;PM&quot; term=&quot;%2235481543%22&quot;&gt;35481543)&lt;/searchLink&gt;&lt;br /&gt;Trends Microbiol. 2025 Apr;33(4):459-471. (PMID: &lt;searchLink fieldCode=&quot;PM&quot; term=&quot;%2239828459%22&quot;&gt;39828459)&lt;/searchLink&gt;&lt;br /&gt;Acta Microbiol Immunol Hung. 2018 Jun 1;65(2):229-240. (PMID: &lt;searchLink fieldCode=&quot;PM&quot; term=&quot;%2229685049%22&quot;&gt;29685049)&lt;/searchLink&gt;&lt;br /&gt;Antimicrob Agents Chemother. 1993 May;37(5):1158-9. (PMID: &lt;searchLink fieldCode=&quot;PM&quot; term=&quot;%228517706%22&quot;&gt;8517706)&lt;/searchLink&gt;
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    Subjects:
      – SubjectFull: Fibroblasts drug effects
        Type: general
      – SubjectFull: Humans
        Type: general
      – SubjectFull: Microbial Sensitivity Tests
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      – SubjectFull: Gene Expression Profiling
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      – SubjectFull: Chlorhexidine analogs & derivatives
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      – SubjectFull: Streptococcus mutans drug effects
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      – SubjectFull: Anti-Bacterial Agents pharmacology
        Type: general
      – SubjectFull: Gene Expression Regulation, Bacterial drug effects
        Type: general
    Titles:
      – TitleFull: Comparative Analysis of Chlorhexidine Derivatives and Alternative Agents on Streptococcus mutans Viability, Biofilm Formation, and Gene Expression.
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              Text: 2026 Aug
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              Value: 4
          Titles:
            – TitleFull: MicrobiologyOpen
              Type: main
ResultId 1