Strain-level Identification of Beneficial Lactobacilli of Dairy Origin using 16S rRNA Sequencing: A Biotechnology Approach
Microbiology Research Journal International,
Aim: This study investigated the cultural method and 16S rRNA gene analysis to reveal the composition and diversity of lactic acid bacteria (LAB) from dairy origin (spontaneous fermented cow milk) in Nigeria.
Methods: Six dairy samples which includes two raw cow milk, two raw goat milk and two fermented cow milk (nono) were collected and subjected to standard microbiological investigation using both cultural and molecular methods. The dairy samples were cultured on MRS media, and the isolates were identified using physiological and biochemical parameters. DNA of four selected probiotic LAB isolates from nono were amplified using PCR while the amplicons were electrophoresed in agarose gel, pre-stained with ethidium bromide and characterized by 16S rRNA gene analysis. The result of the DNA sequencing were analyzed using NCBI BLAST.
Results: A total of 55 presumptive LAB were isolated. Twenty nine (29) Lactobacillus plantarum representing 52.7%, Pediococcus acidilactici 15(27.2%), Lactobacillus casei and Lactobacillus brevis 4(7.3%) while Lactobacillus fermentum 3(5.5%) were recorded respectively. Result of the gel electrophoresis revealed DNA size of approximately 1500bp. The selected probiotic LAB from nono used in this study were confirmed as Lactobacillus plantarum N17, Lactobacillus plantarum N24, Lactobacillus brevis N10 and Lactobacillus casei N1 based on 16S rRNA gene sequence analysis while the phylogenetic analysis revealed a 98-100% similarity with a high homology level which affirms the strain of the organism.
Conclusion: This study has demonstrated the diversity of LAB existing in dairy samples known as fermented cow milk (nono) which could be harnessed as valuable sources for LAB isolation and potential probiotic organisms.
- Lactobacillus species
- gene sequence
- fermented cow milk
How to Cite
Fooks LJ, Fuller R, Gibson GR. Prebiotics, probiotics and human gut microbiology. International Dairy Journal. 1999;9(1):53-61.
Holzapfel EH, Haberer P, Geisen R, Björkroth J, Schillinger U. Taxonomy and important features of probiotic microorganisms in food and nutrition. American Journal of Clinical Nutrition. 2001;73(2):365S-73S
Galia Amirbozorgi, Hamidreza Samadlouie, Seyed, Ahmad Shahidi. Identification and characterization of lactic acid bacteria isolated from Iranian traditional dairy products. International Biological and Biomedical Journal. 2016; 2(1):47-52.
Irma Caro, Germán Bécares, Lucía Fuentes, María R. Garcia-Armesto, Javier Rúa, José M. Castro, Emiliano J. Quinto and Javier Mateo. Evaluation of three PCR primers based on the 16S rRNA gene for the identification of lactic acid bacteria from dairy origin. CyTA - Journal of Food. 2015;13(2):181-187. DOI: 10.1080/19476337
Sharpe ME. The genus Lactobacillus. In: The Prokaryotes. A handbook on habitat, isolation and identification of bacteria. Starr, M.P., H. Stolp, H.G., A. Balows and H.G. Schelegel (Eds.). Springer-Verlag, Berlin. 1981;1653-1679.
Oliveira RBP, Oliveira, A. de L. and Glória MBA. Screening of lactic acid bacteria from vacuum packaged beef for antimicrobial activity. Brazillian Journal of Microbiology. 2008;39(2):368-74.
Rachel Oluwayemisi Ishola, Bukola Christianah Adebayo-Tayo. Screening of Lactic Acid Bacteria Isolated from Fermented Food for Bio-molecules Production. AU J.T. 2012;15(4):205-217.
Amarela Terzic-Vidojevic, Maja Tolinacki, Milica Nikolic, Jelena Lozo, Jelena Begovic, Sahib Gurban Oglu Gulahmadov, Akif Alekperovich Kuliev, Michèle Dalgalarrondo, Jean-Marc Chobert, Thomas Haertlé, Ljubisa Topisirovic. Phenotypic and genotypic characterization of lactic acid bacteria isolated from Azerbaijani traditional dairy products. African Journal of Biotechnology. 2009; 8(11):2576-2588.
Ray B, Bhunia A. Fundamental food microbiology (5th ed.). Boca Raton, FL: Taylor & Francis Group LLC, CRC Press; 2014.
Nero LA, De Mattos MR, De Aguiar Ferreira Barros M, Ortolani MBT, Beloti V, De Melo Franco BDG. Listeria monocytogenes and Salmonella spp. in raw milk produced in Brazil: Occurrence and interference of indigenous microbiota in their isolation and development. Zoonoses and Public Health. 2008; 5(6):299–305.
Henr´ıquez-Aedo K, Dur´an D, Garcia A, Hengst MB, Aranda M. Identification of biogenic amines-producing lactic acid bacteria isolated from spontaneous malolactic fermentation of chilean red wines. LWT- Food Science and Technology. 2016;68:183–189.
Federici S, Ciarrocchi F, Campana R, Ciandrini E, Blasi G, Baffone W. Identification and functional traits of lactic acid bacteria isolated from Ciauscolo salami produced in Central Italy. Meat Science. 2014;98(4):575–584.
Fortune Akabanda, James Owusu-Kwarteng, Kwaku Tano-Debrah, Charles Parkouda, and Lene Jespersen. The Use of Lactic Acid Bacteria Starter Culture in the Production of Nunu, a Spontaneously Fermented Milk Product in Ghana. International Journal of Food Science; 2014. Article ID 721067, 11 pages. Available:http://dx.doi.org/10.1155/2014/721067
Leite AMO, Miguel MAL, Peixoto RS, et al. Probiotic potential of selected lactic acid bacteria strains isolated from Brazilian kefir grains. Journal of Dairy Science. 2015; 98(6):3622–3632.
Tormo A Djamila, Ali H, Lekhal C. Roques Phenotypic and genotypic characterization of lactic acid bacteria isolated from raw goat milk and effect of farming practices on the dominant species of lactic acid bacteria. International Journal of Food Microbiology. 2015;210:9-15. ISSN: 0168-1605.
Aforijiku S, Onilude AA, Wakil SM. Antagonistic effect of developed probiotic yoghurt against some selected food-borne pathogens during cold storage. Greener Journal of Biological Sciences. 2019; 9(2):22-28.
Wawrik B, Kerkho FL, Zylstra GJ, Kukor JJ. Identification of Unique Type II Polyketide Synthase Genes in Soil. Applied Environmental Microbiology. 2005;71(5):2232–2238. DOI: 10.1128/AEM.71.5.2232-2238.2005
Tamura K, Stecher G, Peterson D, Filipski A, Kumar S. MEGA6: Molecular Evolutionary Genetics Analysis version 6.0,” Molecular Biology and Evolution. 2013;30(12):2725-2729.
Bukola C. Adebayo-Tayo, Abiodun A. Onilude. Screening of Lactic Acid Bacteria Strains Isolated from Some Nigerian Fermented Foods for EPS Production. World Applied Science Journal. 2008;4(5): 741-747.
Willey JM, Sherwood LM, Woolverton CJ. Prescott’s principles of microbiology. Boston: McGraw-Hill Higher Education; 2009. ISBN: 9780073375236
Saitou N, Nei M. The neighbor-joining method: a new method for reconstructing phylogenetic trees. Molecular Biology and Evolution. 1987;4(4):406–425.
Felsenstein J. Confidence limits on phylogenies: An approach using the bootstrap. Evolution. 1985;39(4):783–791.
Lili Anggraini, Yetti Marlida, Wizna Wizna, Jamsari Jamsari, Mirzah Mirzah, Frederick Adzitey, Nurul Huda. Molecular identification and phylogenetic analysis of GABA-producing lactic acid bacteria isolated from indigenous dadih of West Sumatera, Indonesia [version awaiting peer review F1000 Research. 2018; 7:1663. Available:https://doi.org/10.12688/f1000research.16224.1
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