Author(s):
Soumya Gulab Katre, Jaykishor A. Chhangani, Payal S. Kathale
Email(s):
saumyakatre48@gmail.com
DOI:
10.52711/0975-4377.2026.00017 
Address:
Soumya Gulab Katre*, Jaykishor A. Chhangani, Payal S. Kathale
Bio-processing and Herbal Division, Mahatma Gandhi Institute for Rural Industrialization, Wardha – 442001, Maharashtra, India.
*Corresponding Author
Published In:
Volume - 18,
Issue - 2,
Year - 2026
ABSTRACT:
Reliable revival of lyophilized bacterial cultures is essential for routine microbiological applications; however, commonly recommended media such as Brain Heart Infusion (BHI) broth are often costly and not readily available. This article describes a simple, reproducible, and cost-effective method for the revival, cultivation, and short-term maintenance of lyophilized Streptococcus mutans obtained from the Microbial Type Culture Collection (MTCC) using tryptic soy and nutrient media as alternatives. Lyophilized cultures were rehydrated and incubated under microaerophilic conditions, followed by isolation on nutrient agar and tryptic soy agar. Method validation was performed using visual turbidity assessment, optical density measurements at 600 nm (OD600), colony-forming unit (CFU) enumeration, colony morphology, and subculture viability. The method enabled rapid revival in tryptic soy media within 12–14h, yielding higher biomass and CFU counts than nutrient media, which exhibited delayed growth. Cultures revived in tryptic soy media and maintained on tryptic soy agar remained viable for up to 10–12 days at 4°C. This validated protocol provides an economical and practical alternative to BHI for routine revival and short-term maintenance of lyophilized S. mutans.
• Validated, low-cost method using tryptic soy and nutrient media
• Reliable revival of lyophilized Streptococcus mutans
• Suitable for routine microbiological and teaching laboratories
Cite this article:
Soumya Gulab Katre, Jaykishor A. Chhangani, Payal S. Kathale. A Validated, Cost-effective Method for Revival and Maintenance of Lyophilized Streptococcus mutans using Nutrient and Tryptic Soy Media. Research Journal of Pharmaceutical Dosage Forms and Technology. 2026; 18(2):108-4. doi: 10.52711/0975-4377.2026.00017
Cite(Electronic):
Soumya Gulab Katre, Jaykishor A. Chhangani, Payal S. Kathale. A Validated, Cost-effective Method for Revival and Maintenance of Lyophilized Streptococcus mutans using Nutrient and Tryptic Soy Media. Research Journal of Pharmaceutical Dosage Forms and Technology. 2026; 18(2):108-4. doi: 10.52711/0975-4377.2026.00017 Available on: https://www.rjpdft.com/AbstractView.aspx?PID=2026-18-2-3
8. REFERENCES:
1. Baker JL, Mark Welch JL, Kauffman KM, McLean JS, He X. The oral microbiome: diversity, biogeography and human health. Nat Rev Microbiol. 2024; 22(2): 89-104. doi:10.1038/s41579-023-00963-6
2. Kaushita Banerjee, Gourav Saha, Pitambar Sahoo, Sourav Chattaraj, Dhamodhar P PT. Synthesis and Characterization of a Lemongrass Oil Emulsion Formulation incorporating Alumina Nanoparticles for activity against Streptococcus mutans Isolated from Dental Caries. Res J Pharm Technol. 2020; 13(5): 2291-2296. doi:10.5958/0974-360X.2020.00413.8
3. Saumya G Nair, Rony T Kondody SB. Evaluation of Effect of Vitamin C on Streptococcus Mutans Induced Secondary Caries: An In vitro Study. Res J Pharm Technol. 2023; 16(7): 3245-3250. doi:10.52711/0974-360X.2023.00533
4. Syed Junaid, Dileep N, Rakesh K.N PKT. Anticaries Activity of Selected Plants against Clinical Isolates of Streptococcus mutans. Asian J Pharm Technol. 2013; 3(3): 107-108. https://ajptonline.com/HTMLPaper.aspx?Journal=Asian Journal of Pharmacy and Technology; PID=2013-3-3-4
5. Zahra M. Ali Mohammed, Jebor Baydaa AH. The effect of Glucoamylase and Glucose oxidase from the Aspergillus niger and Penicillium notatum respectively on inhibition of Streptococcus mutans causes the Dental caries. Res J Pharm Technol. 2018; 11(8): 3419-3422. doi:10.5958/0974-360X.2018.00630.3
6. Taufiqi Hidayatullah, Vera Yulina, Citra Feriana Putri BAG. In-vitro and In-silico Analyses of Ziziphus mauritiana Lam effect on the Virulence properties of Streptococcus mutans. Res J Pharm Technol. 2025; 18(5). doi:10.52711/0974-360X.2025.00314
7. Ida Bagus Narmada, Muhammad Fulan Ardhani SLR, Inggit Dwi Virgianti, Putri Pramita Larasati ERW, Alexander Patera Nugraha TNE binti TAN. Hibiscus sabdariffa L. Flower Distilled Water Extract Antibacterial Activity towards Streptococcus mutans: An in vitro Study. Res J Pharm Technol. 2024; 17(1): 25-30. doi:10.52711/0974-360X.2024.00005
8. Christy. S NMS. Antimicrobial Efficacy of Azadirachta indica against Streptococcus mutans– An In vitro Study. Asian J Pharm Technol. 2019; 9(3): 149-153. doi:10.5958/2231-5713.2019.00025.4
9. Lemos JA, Quivey RG, Koo H, Abranches J. Streptococcus mutans: a new Gram-positive paradigm? Microbiology. 2013; 159(Pt 3): 436-445. doi:10.1099/mic.0.066134-0
10. Lemos JA, Palmer SR, Zeng L, et al. The Biology of Streptococcus mutans. Microbiol Spectr. 2019; 7(1). doi: 10.1128/microbiolspec.GPP3-0051-2018
11. Utamaningyas A, Pramesti H, Balafif F. The Streptococcus mutans ability to survive in biofilms and during dental caries formation: scoping review. J Syiah Kuala Dent Soc. 2023; 7: 150-158. doi:10.24815/jds. v7i2.30295
12. Gao Z, Chen X, Wang C, et al. new strategies and mechanisms for targeting Streptococcus mutans biofilm formation to prevent dental caries: A review. Microbiol Res. 2024; 278: 127526. doi: https://doi.org/10.1016/j.micres.2023.127526
13. Samar Ali, AL Salameh, Mustafa Alammory OH. Comparative Effect of laser treatment on Streptococcus mutans Biofilm adhered to Dental implant surface. Res J Pharm Technol. 2020; 13(7): 3311-3316. doi:10.5958/0974-360X.2020.00587.9
14. Prawati Nuraini, Mega Moeharyono Puteri EP. Anti-biofilm Activity of Epigallocatechin gallate (EGCG) against Streptococcus mutans bacteria. Res J Pharm Technol. 2021; 14(9): 5019-5023. doi:10.52711/0974-360X.2021.00875
15. G.А. Atazhanova, K.Zh. Badekova, S.А. Ivasenko1, Т. Kacergius, Ya.К. Levaya, G.К. Kurmantaeva, M.Yu. Ishmuratova МKS. Influence of Essential Oils on the Formation of Streptococcus mutans Biofilms. Res J Pharm Technol. 2022; 15(11): 4959-4966. doi:10.52711/0974-360X.2022.00834
16. Matsumoto-Nakano M. Role of Streptococcus mutans surface proteins for biofilm formation. Jpn Dent Sci Rev. 2018; 54(1): 22-29. doi: https://doi.org/10.1016/j.jdsr.2017.08.002
17. Rushita Shah, Amit Kumar, Ashutosh Agarwal, Neha TA and PB. Molecular Biology of Streptococcus Mutans: A Review. Acad J Med. 8(1): 25-28. doi:10.48165/ajm.2025.8.01.5
18. Bedoya-Correa CM, Rincón Rodríguez RJ, Parada-Sanchez MT. Genomic and phenotypic diversity of Streptococcus mutans. J Oral Biosci. 2019; 61(1): 22-31. doi:https://doi.org/10.1016/j.job.2018.11.001
19. J. Carlsson. Nutritional Requirements of Streptococcus mutans. Caries Res. 1970; 4(4): 305-320. doi: https://doi.org/10.1159/000259653
20. Zhou X, Li Y, eds. Chapter 2 - Techniques for Oral Microbiology. In: Atlas of Oral Microbiology. Academic Press; 2015:15-40. doi: https://doi.org/10.1016/B978-0-12-802234-4.00002-1
21. Hashimoto T, Koga M, Masaoka Y. Advantages of a diluted nutrient broth medium for isolating N2-producing denitrifying bacteria of α-Proteobacteria in surface and subsurface upland soils. Soil Sci Plant Nutr. 2009; 55(5): 647-659. doi:10.1111/j.1747-0765.2009. 00404.x
22. Okada A, Gotoh Y, Watanabe T, Furuta E, Yamamoto K, Utsumi R. Targeting Two‐Component Signal Transduction: A Novel Drug Discovery System. In: Simon MI, Crane BR, Crane A, eds. Two‐Component Signaling Systems, Part A. Vol 422. Methods in Enzymology. Academic Press; 2007: 386-395. doi: https://doi.org/10.1016/S0076-6879(06)22019-6
23. Agurokpon D. General Components of Culture Media.; 2021. doi:10.13140/RG.2.2.19467.00801
24. Heckly RJ. Preservation of Bacteria by Lyophilization. In: Umbreit WW, ed. Vol 3. Advances in Applied Microbiology. Academic Press; 1961:1-76. doi: https://doi.org/10.1016/S0065-2164(08)70506-9
25. Arguelles E. Maintenance and Preservation of Microalgal Cultures. In: 2020:53-60.
26. Liang X, Gong T, Chen JJ, Chen TJ, Yang JL, Zhu P. Influence of Long-Term Agar-Slant Preservation at 4 °C on the Recombinant Enzyme Activity of Engineered Yeast. Fermentation. 2023; 9(2). doi:10.3390/fermentation9020104
27. Sousa AM, Machado I, Nicolau A, Pereira MO. Improvements on colony morphology identification towards bacterial profiling. J Microbiol Methods. 2013; 95(3): 327-335. doi: https://doi.org/10.1016/j.mimet.2013.09.020
28. Laflamme C, Simard JR, Buteau S, et al. Effect of growth media and washing on the spectral signatures of aerosolized biological simulants. Appl Opt. 2011; 50: 788-796. doi:10.1364/AO.50.000788
29. Hatiboruah D, Yengkhom D, Namsa N, Nath P. Turbidimetric analysis of growth kinetics of bacteria in the laboratory environment using smartphone. J Biophotonics. 2020; 13. doi:10.1002/jbio.201960159
30. Kumar SB, Narayana IH, Murthy SB V, Mathew S, Damodhar P, Kumar SSS. Isolation and Genetic Characterization of Mutans Streptococci Associated with Dental Caries in Rural Field Practice of a Dental Institution: In Vivo: Study. Indian J Dent Res. 2019; 30(6). https://journals.lww.com/ijdr/fulltext/2019/30060/isolation_and_genetic_characterization_of_mutans.15.aspx
31. Thiruvengadam V, Chitharanjan AB, Kumar K, Singaram V. Comparison of Streptococcus mutans Adhesion on New and Recycled Metal Brackets: An In Vitro Study. Cureus. 2022; 14(3): e23574. doi:10.7759/cureus.23574
32. Khan AU, Islam B, Khan SN, Akram M. A proteomic approach for exploring biofilm in Streptococcus mutans. Bioinformation. 2011; 5(10): 440-445. doi:10.6026/97320630005440
33. Manohar P, Ramesh N. Improved lyophilization conditions for long-term storage of bacteriophages. Sci Rep. 2019; 9(1): 15242. doi:10.1038/s41598-019-51742-4
34. Ohta H, Hattori T. Bacteria sensitive to nutrient broth medium in terrestrial environments. Soil Sci Plant Nutr. 1980; 26: 99-107. doi:10.1080/00380768.1980.10433216