Biofilm-Forming oral Microorganisms in Dental Caries: Isolation, Identification, and Nanoemulsion Intervention
DOI:
https://doi.org/10.63682/jns.v14i24S.3996Abstract
Introduction: Oral biofilm, or dental plaque, is a complex microbial community composed of bacteria, fungi, and other microbes that form extracellular polymeric substances (EPS), leading to dental caries, periodontal disorders, and other oral health problems. Biofilms are resistant to antimicrobial agents and host immune responses, necessitating alternative treatment strategies. Herbal extracts, probiotics, quorum-sensing inhibitors, and biomaterials are being explored as biofilm inhibitors.
Objectives: This study aims to isolate and morphologically characterize microbial species from oral swabs and assess their biofilm-forming potential through a multidisciplinary approach involving colony morphology, VITEK® 2-based taxonomic identification, and biofilm evaluation using crystal violet and Congo red agar assays.
Methods: Swabs from eight patients with dental caries and plaque were collected and used for laboratory analysis. Bacteria were isolated through serial dilution and cultured on nutrient and Mitis salivarius agar (MSA) media. Anaerobic and aerobic bacteria were identified using Gram’s staining and the VITEK® 2 automated system. The next step will involve nanoemulsion preparation to assess its antimicrobial and biofilm inhibition effects.
Results: The study identified ten bacterial isolates from oral swabs, including Alloiococcus otitis, Enterococcus faecium, Klebsiella pneumoniae, Kocuria kristinae, Gemella bergeri, and Candida albicans, with distinct morphological features and confirmed taxonomy via the VITEK® 2 system. Among these, Enterococcus faecium (OD 1.32) and Alloiococcus otitis (OD 0.91) demonstrated significant biofilm-forming ability as confirmed by both crystal violet assay and Congo red agar assay, highlighting their potential role in oral biofilm development. Identified microorganisms included Alloiococcus otitis, Enterococcus faecium, Klebsiella pneumoniae, Kocuria kristinae, Gemella bergeri, and Candida albicans. Biofilm formation was assessed using Congo red and crystal violet assays, revealing biofilm production in Enterococcus faecalis and Alloiococcus otitis.
Conclusions: This study concludes that among the six microbial species identified from oral swabs of eight patients with dental plaques, Alloiococcus otitis and Enterococcus faecalis demonstrated significant biofilm-forming ability as confirmed by crystal violet and Congo red agar assays, suggesting the need for future exploration into antibiofilm strategies
Downloads
Metrics
References
Alghamdi, S. (2022). Isolation and identification of the oral bacteria and their characterization for bacteriocin production in the oral cavity. Saudi journal of biological sciences, 29(1), 318-323. doi: 10.1016/j.sjbs.2021.08.096. Epub 2021
Ali Hameed AL-Dabbagh, A., Ali Ajah, H., & Abdul Sattar Salman, J. (2023). Detection of virulence factors from Candida spp. Isolated from oral and vaginal candidiasis in Iraqi patients. Archives of Razi Institute, 78(1), 465-474. doi: 10.22092/ARI.2022.359464.2420
Aneja, K. R. (2009). Expriments in Microbiology, plant pathology and Biotechnology fourth ed. New Age International Publishers, Daryaganj, New Delhi.
Asghar, F., Bano, A., Waheed, F., Anjum, A. A., Ejaz, H., & Javed, N. (2023). Association of exogenous factors with molecular epidemiology of Staphylococcus aureus in human oral cavity. Saudi Journal of Biological Sciences, 30(4), 103613. Doi: 10.1016/j.sjbs.2023.103613.
Cappuccino, J.G. and Sherman, N. 2005. Microbiology: A Laboratory Manual. 7th Ed. Pearson Education, Inc., San Francisco,
Chen, X., Daliri, E. B. M., Chelliah, R., & Oh, D. H. (2020). Isolation and identification of potentially pathogenic microorganisms associated with dental caries in human teeth biofilms. Microorganisms, 8(10), 1596. doi: 10.3390/microorganisms8101596. PMID: 33081291; PMCID: PMC7603000.
Costa, R. C., Bertolini, M., Costa Oliveira, B. E., Nagay, B. E., Dini, C., Benso, B., ... & Souza, J. G. S. (2023). Polymicrobial biofilms related to dental implant diseases: unravelling the critical role of extracellular biofilm matrix. Critical reviews in microbiology, 49(3), 370-390. doi: 10.1080/1040841X.2022.2062219. Epub 2022 May 18. PMID: 35584310.
Garcia, C. R., Malik, M. H., Biswas, S., Tam, V. H., Rumbaugh, K. P., Li, W., & Liu, X. (2022). Nanoemulsion delivery systems for enhanced efficacy of antimicrobials and essential oils. Biomaterials Science, 10(3), 633-653. doi: 10.1039/d1bm01537k. PMID: 34994371.
Jakubovics, N. S., Goodman, S. D., Mashburn‐Warren, L., Stafford, G. P., & Cieplik, F. (2021). The dental plaque biofilm matrix. Periodontology 2000, 86(1), 32-56. doi: 10.1111/prd.12361. Epub 2021 Mar 10. PMID: 33690911; PMCID:
Khalili, H., Sisakhtpour, R., & Abdollahzadeh, S. (2018). Identification of biofilm-producing bacterial species causing dental caries and their sensitivity to zoocin A and clove hydroalcoholic extract. Immunopathologia Persa, 4(2), e27-e27. DOI:10.15171/ipp.2018.27
Khan, R. A., & Sarmah, R. (2023). Factors influencing microbial growth in the human oral cavity. Reviews and Research in Medical Microbiology, 34(3), 123-129. Doi: 10.1097/MRM.0000000000000338
Kolpen, M., Kragh, K. N., Enciso, J. B., Faurholt-Jepsen, D., Lindegaard, B., Egelund, G. B., ... & Bjarnsholt, T. (2022). Bacterial biofilms predominate in both acute and chronic human lung infections. Thorax, 77(10), 1015-1022. DOI: 10.1136/thoraxjnl-2021-217576
Mohammed, H. A., Sahi, N. M., Ahmed, R. T., & Fauzi Al-Rubaye, A. (2022). Antimicrobial activity of some nanoparticles synthesized by laser ablation technique against some bacteria isolated from oral cavity. Medical Journal of Babylon, 19(4), 601-608. DOI:10.4103/MJBL.MJBL_147_22
Nabawy, A., Makabenta, J. M., Schmidt-Malan, S., Park, J., Li, C. H., Huang, R., ... & Rotello, V. M. (2022). Dual antimicrobial-loaded biodegradable nanoemulsions for synergistic treatment of wound biofilms. Journal of Controlled Release, 347, 379-388. DOI: 10.1016/j.jconrel.2022.05.013
Ngwu, J. N., Uzoeto, H. O., Emaimo, J., Okorie, C., Mohammed, I. D., Edemekong, C. I., ... & Iroha, I. R. (2022). Antibiogram of biofilm forming oral streptococci species isolated from dental caries patients visiting federal college of dental technology and therapy, Enugu Nigeria. Int J Res Rep Dent, 5(1), 12-25.
Omori, M., Kato-Kogoe, N., Sakaguchi, S., Fukui, N., Yamamoto, K., Nakajima, Y., ... & Ueno, T. (2021). Comparative evaluation of microbial profiles of oral samples obtained at different collection time points and using different methods. Clinical Oral Investigations, 25, 2779-2789. DOI: 10.1007/s00784-020-03592-y
Prescott, L.M., Harley, J.P. and Klein, D.A. (2005) Microbiology. Sixth International Edition. Mcgraw-Hill Publishing Company, UK, 652-668CA, 73
Radunovic, M., Barac, M., Kuzmanovic Pficer, J., Pavlica, D., Jovanovic, A., Pucar, A., & Petrovic, S. (2022). Antifungal Susceptibility of Candida albicans Isolated from Tongue and Subgingival Biofilm of Periodontitis Patients. Antibiotics, 11(6), 802. DOI: 10.3390/antibiotics11060802
Ramírez-Larrota, J. S., & Eckhard, U. (2022). An introduction to bacterial biofilms and their proteases, and their roles in host infection and immune evasion. Biomolecules, 12(2), 306. DOI: 10.3390/biom12020306
Ray, R. R. (2022). Dental biofilm: Risks, diagnostics and management. Biocatalysis and Agricultural Biotechnology, 43, 102381.
Viksne, R., Racenis, K., Broks, R., Balode, A. O., Kise, L., & Kroica, J. (2023). In vitro assessment of biofilm production, antibacterial resistance of staphylococcus aureus, Klebsiella pneumoniae, Pseudomonas aeruginosa, and Acinetobacter spp. Obtained from tonsillar crypts of healthy adults. Microorganisms, 11(2), 258. DOI: 10.3390/microorganisms11020258
Wu, X., Al Farraj, D. A., Rajaselvam, J., Alkufeidy, R. M., Vijayaraghavan, P., Alkubaisi, N. A., ... & Alshammari, M. K. (2020). Characterization of biofilm formed by multidrug resistant Pseudomonas aeruginosa DC-17 isolated from dental caries. Saudi Journal of Biological Sciences, 27(11), 2955-2960. DOI: 10.1016/j.sjbs.2020.07.020
Downloads
Published
How to Cite
Issue
Section
License
Copyright (c) 2025 Sofia. A, Rimal Issac. R. S, Aneesh Nair, Sujina. M. G
This work is licensed under a Creative Commons Attribution 4.0 International License.
You are free to:
- Share — copy and redistribute the material in any medium or format
- Adapt — remix, transform, and build upon the material for any purpose, even commercially.
Terms:
- Attribution — You must give appropriate credit, provide a link to the license, and indicate if changes were made. You may do so in any reasonable manner, but not in any way that suggests the licensor endorses you or your use.
- No additional restrictions — You may not apply legal terms or technological measures that legally restrict others from doing anything the license permits.
