Characterization and quantitative assessment of lactic acid bacteria in fermented finger millet (Eleusine coracana) Variety-ML365
Article Main
Abstract
Finger millet (Eleusine coracana), a nutrient-rich cereal, is traditionally consumed in India and Africa, particularly among low-income populations. It is a great source of proteins, fibres, carbohydrates, and minerals, including calcium, making it a viable substrate for the development of functional meals. The aim was to separate and molecularly characterize lactic acid bacteria (LAB) from the fermented flour of the Indian finger millet variety ML365. Twelve bacterial isolates were isolated on MRS (de Man Rogosa and Sharpe) agar after fermentation. Based on morphological, biochemical, and preliminary probiotic screening tests, five isolates (EC01–EC05) were selected for detailed study. Biochemical characterization revealed traits typical of LAB, including positive lactose utilization, starch hydrolysis, and negative catalase and oxidase activities. For molecular identification, the 16S rRNA gene was amplified and sequenced. Sequence analysis using BLAST revealed that the isolates were similar to species belonging to the Bacillus and Brevibacterium genera. In particular, isolates closely resembled Brevibacterium sp., Bacillus cereus, Bacillus subtilis, Bacillus amyloliquefaciens, and Bacillus pumilus. Their genetic ties were further validated by phylogenetic analysis. The accession numbers for the nucleotide sequences that were put into the GenBank database were PQ606366.1, PQ606365.1, PQ606371.1, PQ606367.1, and PQ606372.1. The study found that fermented finger millet flour is a good source of LAB and may possess probiotic qualities. Isolates like these may be utilized in the development of innovative probiotic formulations and functional foods. It is recommended that further in vivo research be conducted to investigate their safety profiles, potential technical applications in the food industry, and health-promoting properties.
Article Details
Article Details
Keywords
Bacillus species, Fermentation, Finger millet, Lactic acid bacteria (LAB), Probiotic isolates
References
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Adisa, A.M., Badejo, A.A., Ifesan, B.O.T. & Enujiugha, V.N. (2024). Phenotypic and molecular differentiation of lactic acid bacteria in fonio millet ogi fermentation and their potential as starter cultures. Food and Humanity 2, 100230. https://doi.org/10.1016/j.foohum.2024.100230
Akbulut, S., Baltaci, M.O., Adiguzel, G. & Adiguzel, A. (2022). Identification and potential biotechnological characterization of Lactic Acid Bacteria isolated from white cheese samples. Journal of Pure and Applied Microbiology, 16(4), 1-12. https://doi.org/10.21203/rs.3.rs-630567/v1.
Bindu, A. & Lakshmidevi, N. (2021). Identification and in vitro evaluation of probiotic attributes of lactic acid bacteria isolated from fermented food sources. Archives of Microbiology, 203, 579–595. https://doi.org/10.1007/s00203-020-02037-0
Chaudhary, J.K. & Mudgal, S. (2020). Effect of incorporation of Finger millet (Eleusine coracana) on the antimicrobial, ACE inhibitory, antioxidant and antidiabetic potential of a milk millet composite probiotic fermented product. Indian Journal of Dairy Science, 73(3), 222-230. https://doi.org/10.33785/IJDS.2020.v73i03.005
Divisekera, D.M.W.D., Samarasekera, J.K.R.R., Hettiarachchi, C., Gooneratne, J., Choudhary, M.I., Gopalakrishnan, S. & Wahab, A.T. (2019). Lactic acid bacteria isolated from fermented flour of finger millet, its probiotic attributes and bioactive properties. Annals of Microbiology, 69, 71-92. https://doi.org/10.1007/s13213-018-1399-y
Dorn-In, S., Bassitta, R., Schwaiger, K., Bauer, J. & Holzel., C.S. (2015). Specific amplification of bacterial DNA by optimized so-called universal bacterial primers in samples rich of plant DNA. Journal of Microbiological Methods, 113, 50-56. https://doi.org/10.1016/j.mimet.2015.04.001
Endo, A., Tanizawa, Y. & Arita, M. (2024). Isolation and identification of lactic Acid bacteria from environmental samples. Lactic Acid Bacteria, 2851, 3-13. doi: 10.1007/978-1-4939-8907-2_1
Frank, J.A., Reich, C.I., Sharma, S., Weisbaum, J.S., Wilson, B.A. & Olsen, G.J. (2008). Critical evaluation of two primers commonly used for amplification of bacterial 16S rRNA genes. Applied and Environmental Biology, 74 (8), 2461 – 2470. https://doi.org/10.1128/AEM.02272-07
Gouthami, B., Ramalakshmi, A., Balakrishnan, M., Karthikeyan, S., Muniraj, I. & Packialakshmi, J.S. (2024). Functional and molecular characterization of millet associated probiotic bacteria. BMC Microbiology, 24(485), 1-15. https://doi.org/10.1186/s12866-024-03606-9
Gupta, S., Mohanty, U. & Majumdar, R.K. (2021). Isolation and characterization of lactic acid bacteria from traditional fermented fish product Shidal of India with reference to their probiotic potential. LWT, 146, 11164. https://doi.org/10.1016/j.lwt.2021.111641
Gupta, R. & Gaur, S. (2025). Millet processing to fermented drinks: Evaluating changes in nutritional attributes, mineral accessibility, amino acid profile, and sensorial acceptability. International Journal of Gastronomy and Food Science, 101297, https://doi.org/10.1016/j.ijgfs.2025.101297
Huligere, S.S., Kumari, V.B.C., Alqadi, T., Kumar, S., Cull, C.A., Amachawadi, R.G. & Ramu, R. (2022). Isolation and characterization of lactic acid bacteria with potential probiotic activity and further investigation of their activity by α-amylase and α-glucosidase inhibitions of fermented batters. Frontiers in Microbiology, 13, 01-18. https://doi.org/10.3389/fmicb.2022.1042263
Keklik, G. (2023). Understanding Evolutionary Relationships and analysis methods through MEGA software. International Journal of New Horizons in the Sciences (JIHSCI), 1(2), 83-90. https://doi.org/10.5281/zenodo.10446714
Khushboo., Karnwal, A. & Mal, T. (2023). Characterization and selection of probiotic lactic acid bacteria from different dietary sources for development of functional foods. Frontiers of Microbiology, 14, 1-14. DOI 10.3389/fmicb.2023.1170725.
Lorn, D., Nguyen, T.K.C., HO, P.H., Tan, R., Licandro, H. & Wache, Y. (2021). Screening of lactic acid bacteria for their potential use as aromatic starters in fermented vegetables. International Journal of Food Microbiology, 350, 109242. https://doi.org/10.1016/j.ijfoodmicro.2021.109242
Lungani, A.N. (2021). Nutritional, antioxidant and microbiological properties of finger millet-based beverages as influenced by starch source during lactic acid fermentation. Doctoral dissertation, University of Pretoria, 1-159. https://repository.up.ac.za/handle/2263/85161
Mohanan, M.M., Shetty, R., Bhat, P.S., Deepashree, V.S., Thimulappa, R.K., Bang-Berthelsen, C.H. & Mudnakudu-Nagaraju, K.K. (2025). Isolation and characterization of biological traits of millet-derived lactic acid bacteria. International Journal of Food Science and Technology, 60(1), 1-28. vvaf074 https://doi.org/10.1093/ijfood/vvaf074
Obioha, P.I., Ouoba, L.I.I., Anyogu, A., Awamaria, B., Atchia, S., Ojimelukwe, P.C., Sutherland , J.P. & Ghoddusi, H.B. (2021). Identification and characterization of the lactic acid bacteria associated with the traditional fermentation of dairy fermented product. Brazilian Journal of Microbiology, 52(2), 869–881. doi: 10.1007/s42770-021-00461-y
Petkova, M., Stefanova, P., Gotcheya, V. & Angelov, A. (2021). Isolation and Characterization of Lactic Acid Bacteria and Yeast from typical Bulgarian sourdoughs. Microorganisms, 9(7), 1-18. https://doi.org/10.3390/microorganisms9071346
Saboori, K.B., Shahidi, F., Hedayati, S. & Javamanesh, A. (2022). Investigating the Probiotic Properties and Antimicrobial Activity of Lactic Acid Bacteria Isolated from an Iranian Fermented Dairy Product. Foods, 11,1-13. https:// doi.org/10.3390/foods11233904
Shivakumar, D., Gunduraj, A. & Sreenivasa, M.Y. (2023). Isolation, Evaluation, and In-Vitro Characterization of Lactobacillus Isolated from Traditional Fermented Food Finger Millet (Eleusine coracana) Batter for Their Probiotic Attributes. Food Biotechnology, 37(4), 338-363. https://doi.org/10.1080/08905436.2023.2267121
Singh, P., Singh, R., Bhadauria, V.& Singh, H. (2024). The functionality and extraction of protein from sorghum, finger millet, and Kodo millet: a review. International Journal of Food Science and Technology, 59(1), 512-521. https://doi.org/10.1111/ijfs.16747
Srinivash, M., Krishnamoorthi, R., Mahalingam, P.U., Malaikozhundan, B. & Keerthivasan, M. (2023). Probiotic potential of exopolysaccharide producing lactic acid bacteria isolated from homemade fermented food products. Journal of Agriculture and Food Research, 11, 1-15. https://doi.org/10.1016/j.jafr.2023.100517
Vila-Real, C., Pimenta-Martins, A., Mbugua, S., Hagretou, S.L., Katina K., Maina, N.H., Pinto, E. & Gomes, A.M.P. (2022). Novel synbiotic fermented finger millet-based yoghurt-like beverage: Nutritional, physicochemical, and sensory characterization. Journal of Functional Foods, 99, 1-10. https://doi.org/10.1016/j.jff.2022.105324
Yu, J., Wang, H.M., Zha, M.S., Qing, Y.T., Bai, N., Ren, Y., Xi, X.X., Liu, W.J., Menghe, B.L.G. & Zhang, H.P. (2015). Molecular identification and quantification of lactic acid bacteria in traditional fermented dairy foods of Russia. Journal of Dairy Science, 98(8), 5143–5154. http://dx.doi.org/10.3168/jds.2015-9460.
Zhao, Q., Zhang, Y., Xing, X., Li, S., Sun, R., Zhang, W.& Zhang, J. (2025). Genome-wide identification and abiotic stress response analysis of C2H2 zinc finger protein genes in foxtail millet (Setaria italica). Agronomy, 15(7), 1618. https://doi.org/10.3390/agronomy15071618
Aboutalebian, S., Ahmadikia, K., Fakhim, H., Chabavizadeh, J., Okhovat, A., Nikaeen, M. & Mirhendi, H. (2021). Direct detection and identification of the most common bacteria and fungi causing otitis externa by a stepwise multiplex PCR. Frontiers in Cellular and Infection Microbiology, 11, 1-11. doi: 10.3389/fcimb.2021.644060.1-11
Adisa, A.M., Badejo, A.A., Ifesan, B.O.T. & Enujiugha, V.N. (2024). Phenotypic and molecular differentiation of lactic acid bacteria in fonio millet ogi fermentation and their potential as starter cultures. Food and Humanity 2, 100230. https://doi.org/10.1016/j.foohum.2024.100230
Akbulut, S., Baltaci, M.O., Adiguzel, G. & Adiguzel, A. (2022). Identification and potential biotechnological characterization of Lactic Acid Bacteria isolated from white cheese samples. Journal of Pure and Applied Microbiology, 16(4), 1-12. https://doi.org/10.21203/rs.3.rs-630567/v1.
Bindu, A. & Lakshmidevi, N. (2021). Identification and in vitro evaluation of probiotic attributes of lactic acid bacteria isolated from fermented food sources. Archives of Microbiology, 203, 579–595. https://doi.org/10.1007/s00203-020-02037-0
Chaudhary, J.K. & Mudgal, S. (2020). Effect of incorporation of Finger millet (Eleusine coracana) on the antimicrobial, ACE inhibitory, antioxidant and antidiabetic potential of a milk millet composite probiotic fermented product. Indian Journal of Dairy Science, 73(3), 222-230. https://doi.org/10.33785/IJDS.2020.v73i03.005
Divisekera, D.M.W.D., Samarasekera, J.K.R.R., Hettiarachchi, C., Gooneratne, J., Choudhary, M.I., Gopalakrishnan, S. & Wahab, A.T. (2019). Lactic acid bacteria isolated from fermented flour of finger millet, its probiotic attributes and bioactive properties. Annals of Microbiology, 69, 71-92. https://doi.org/10.1007/s13213-018-1399-y
Dorn-In, S., Bassitta, R., Schwaiger, K., Bauer, J. & Holzel., C.S. (2015). Specific amplification of bacterial DNA by optimized so-called universal bacterial primers in samples rich of plant DNA. Journal of Microbiological Methods, 113, 50-56. https://doi.org/10.1016/j.mimet.2015.04.001
Endo, A., Tanizawa, Y. & Arita, M. (2024). Isolation and identification of lactic Acid bacteria from environmental samples. Lactic Acid Bacteria, 2851, 3-13. doi: 10.1007/978-1-4939-8907-2_1
Frank, J.A., Reich, C.I., Sharma, S., Weisbaum, J.S., Wilson, B.A. & Olsen, G.J. (2008). Critical evaluation of two primers commonly used for amplification of bacterial 16S rRNA genes. Applied and Environmental Biology, 74 (8), 2461 – 2470. https://doi.org/10.1128/AEM.02272-07
Gouthami, B., Ramalakshmi, A., Balakrishnan, M., Karthikeyan, S., Muniraj, I. & Packialakshmi, J.S. (2024). Functional and molecular characterization of millet associated probiotic bacteria. BMC Microbiology, 24(485), 1-15. https://doi.org/10.1186/s12866-024-03606-9
Gupta, S., Mohanty, U. & Majumdar, R.K. (2021). Isolation and characterization of lactic acid bacteria from traditional fermented fish product Shidal of India with reference to their probiotic potential. LWT, 146, 11164. https://doi.org/10.1016/j.lwt.2021.111641
Gupta, R. & Gaur, S. (2025). Millet processing to fermented drinks: Evaluating changes in nutritional attributes, mineral accessibility, amino acid profile, and sensorial acceptability. International Journal of Gastronomy and Food Science, 101297, https://doi.org/10.1016/j.ijgfs.2025.101297
Huligere, S.S., Kumari, V.B.C., Alqadi, T., Kumar, S., Cull, C.A., Amachawadi, R.G. & Ramu, R. (2022). Isolation and characterization of lactic acid bacteria with potential probiotic activity and further investigation of their activity by α-amylase and α-glucosidase inhibitions of fermented batters. Frontiers in Microbiology, 13, 01-18. https://doi.org/10.3389/fmicb.2022.1042263
Keklik, G. (2023). Understanding Evolutionary Relationships and analysis methods through MEGA software. International Journal of New Horizons in the Sciences (JIHSCI), 1(2), 83-90. https://doi.org/10.5281/zenodo.10446714
Khushboo., Karnwal, A. & Mal, T. (2023). Characterization and selection of probiotic lactic acid bacteria from different dietary sources for development of functional foods. Frontiers of Microbiology, 14, 1-14. DOI 10.3389/fmicb.2023.1170725.
Lorn, D., Nguyen, T.K.C., HO, P.H., Tan, R., Licandro, H. & Wache, Y. (2021). Screening of lactic acid bacteria for their potential use as aromatic starters in fermented vegetables. International Journal of Food Microbiology, 350, 109242. https://doi.org/10.1016/j.ijfoodmicro.2021.109242
Lungani, A.N. (2021). Nutritional, antioxidant and microbiological properties of finger millet-based beverages as influenced by starch source during lactic acid fermentation. Doctoral dissertation, University of Pretoria, 1-159. https://repository.up.ac.za/handle/2263/85161
Mohanan, M.M., Shetty, R., Bhat, P.S., Deepashree, V.S., Thimulappa, R.K., Bang-Berthelsen, C.H. & Mudnakudu-Nagaraju, K.K. (2025). Isolation and characterization of biological traits of millet-derived lactic acid bacteria. International Journal of Food Science and Technology, 60(1), 1-28. vvaf074 https://doi.org/10.1093/ijfood/vvaf074
Obioha, P.I., Ouoba, L.I.I., Anyogu, A., Awamaria, B., Atchia, S., Ojimelukwe, P.C., Sutherland , J.P. & Ghoddusi, H.B. (2021). Identification and characterization of the lactic acid bacteria associated with the traditional fermentation of dairy fermented product. Brazilian Journal of Microbiology, 52(2), 869–881. doi: 10.1007/s42770-021-00461-y
Petkova, M., Stefanova, P., Gotcheya, V. & Angelov, A. (2021). Isolation and Characterization of Lactic Acid Bacteria and Yeast from typical Bulgarian sourdoughs. Microorganisms, 9(7), 1-18. https://doi.org/10.3390/microorganisms9071346
Saboori, K.B., Shahidi, F., Hedayati, S. & Javamanesh, A. (2022). Investigating the Probiotic Properties and Antimicrobial Activity of Lactic Acid Bacteria Isolated from an Iranian Fermented Dairy Product. Foods, 11,1-13. https:// doi.org/10.3390/foods11233904
Shivakumar, D., Gunduraj, A. & Sreenivasa, M.Y. (2023). Isolation, Evaluation, and In-Vitro Characterization of Lactobacillus Isolated from Traditional Fermented Food Finger Millet (Eleusine coracana) Batter for Their Probiotic Attributes. Food Biotechnology, 37(4), 338-363. https://doi.org/10.1080/08905436.2023.2267121
Singh, P., Singh, R., Bhadauria, V.& Singh, H. (2024). The functionality and extraction of protein from sorghum, finger millet, and Kodo millet: a review. International Journal of Food Science and Technology, 59(1), 512-521. https://doi.org/10.1111/ijfs.16747
Srinivash, M., Krishnamoorthi, R., Mahalingam, P.U., Malaikozhundan, B. & Keerthivasan, M. (2023). Probiotic potential of exopolysaccharide producing lactic acid bacteria isolated from homemade fermented food products. Journal of Agriculture and Food Research, 11, 1-15. https://doi.org/10.1016/j.jafr.2023.100517
Vila-Real, C., Pimenta-Martins, A., Mbugua, S., Hagretou, S.L., Katina K., Maina, N.H., Pinto, E. & Gomes, A.M.P. (2022). Novel synbiotic fermented finger millet-based yoghurt-like beverage: Nutritional, physicochemical, and sensory characterization. Journal of Functional Foods, 99, 1-10. https://doi.org/10.1016/j.jff.2022.105324
Yu, J., Wang, H.M., Zha, M.S., Qing, Y.T., Bai, N., Ren, Y., Xi, X.X., Liu, W.J., Menghe, B.L.G. & Zhang, H.P. (2015). Molecular identification and quantification of lactic acid bacteria in traditional fermented dairy foods of Russia. Journal of Dairy Science, 98(8), 5143–5154. http://dx.doi.org/10.3168/jds.2015-9460.
Zhao, Q., Zhang, Y., Xing, X., Li, S., Sun, R., Zhang, W.& Zhang, J. (2025). Genome-wide identification and abiotic stress response analysis of C2H2 zinc finger protein genes in foxtail millet (Setaria italica). Agronomy, 15(7), 1618. https://doi.org/10.3390/agronomy15071618
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Characterization and quantitative assessment of lactic acid bacteria in fermented finger millet (Eleusine coracana) Variety-ML365. (2025). Journal of Applied and Natural Science, 17(4), 1561-1568. https://doi.org/10.31018/jans.v17i4.6875
