Bifidogenic properties of cell-free extracts derived from probiotic strains of Bifidobacterium bifidum and Lactobacillus reuteri

Keywords: probiotic bacteria; biofilm formation; biotechnological potential; proliferative activity; cell-free extract.


Comprehensive study of the biological activity of structural components and metabolites of “beneficial” microorganisms opens the prospects of efficient and rational use of their biotechnological potential in the correction of microecological and related disorders. The study tested proliferative activity and biofilm formation by Bifidobacterium bifidum probiotic strain under the influence of cell-free extracts containing structural components and metabolites of the probiotic strains of B. bifidum and Lactobacillus reuteri. Cell-free extracts were obtained by disintegrating suspensions of probiotic cells by cyclic freezing-thawing, cultivating probiotic microorganisms in their own disintegrates and subsequent filtration of the obtained disintegrates and cultures. The proliferative activity and biofilm formation of the probiotic test culture were studied by spectrophotometric microtiter plate method with 10%vol, 30%vol and 50%vol content of cell-free extracts in the cultivation medium. All investigated extracts showed a significant concentration-dependent stimulatory effect on the proliferative activity of B. bifidum. According to the degree of stimulatory effect on the B. bifidum proliferation, cell-free extracts arranged in ascending order: MLG (filtrate of L. reuteri culture, grown in L. reuteri disintegrate supplemented with 0.8 M glycerol and 0.4 M glucose) < MB (filtrate of В. bifidum culture, grown in В. bifidum disintegrate) < B (filtrate of В. bifidum disintegrate) < ML (filtrate of L. reuteri culture, grown in L. reuteri disintegrate) < L (filtrate of L. reuteri disintegrate). With the same content in the culture medium, filtrates of disintegrates had a more pronounced stimulatory effect than filtrates of cultures grown in their own disintegrates. Cell-free extracts from L. reuteri (L and ML) exerted a more pronounced stimulatory effect than cell-free extracts from B. bifidum. Not all studied cell-free extracts stimulated the biofilm formation by B. bifidum. The effect of cell-free extracts on this process depended on their type and concentration. Extract L had a predominantly inhibitory effect on biofilm formation by B. bifidum. The most pronounced stimulatory effect on biofilm formation by B. bifidum came from extract MLG. ML, B and MB extracts stimulated this process approximately equally. The detection of significant bifidogenic effect of the studied cell-free extracts may contribute to their pharmaceutical applications. Cell-free extracts can be used as metabiotics or prebiotics for increasing the survival of the injected probiotic, facilitating its inoculation in the gastrointestinal tract when used together. The obtained data encourage further careful study of the biochemical composition of cell-free extracts and efforts to clarify the mechanism of their action.


Cicvarek, J., Čurda, L., Elich, O., Dvořáková, E., & Dvořák, M. (2010). Effect of caseinomacropeptide concentrate addition on the growth of bifidobacteria. Czech Journal of Food Sciences, 28(6), 485–494.

Feng, Q., Chen, W. D., & Wang, Y. D. (2018). Gut microbiota: An integral mode rator in health and disease. Frontiers in Microbiology, 9, 151.

Gagliardi, A., Totino, V., Cacciotti, F., Iebba, V., Neroni, B., Bonfiglio, G., Tran cassini, M., Passariello, C., Pantanella, F, & Schippa, S. (2018). Rebuilding the gut microbiota ecosystem. International Journal of Environmental Research and Public Health, 15(8), 1679.

Hamet, M. F., Medrano, M., Perez, P. F., & Abraham, A. G. (2016). Oral administra tion of kefiran exerts a bifidogenic effect on BALB/c mice intestinal micro biota. Beneficial Microbes, 7(2), 237–246.

In Lee, S. H., Barancelli, G. V., de Camargo, T. M., Corassin, C. H., Rosim, R. E., da Cruz, A. G., Cappato, L. P., & de Oliveira, C. A. (2017). Biofilm-produ cing ability of Listeria monocytogenes isolates from Brazilian cheese proces sing plants. Food Research International, 91, 88–91.

Keith, J. W., & Pamer, E. G. (2018). Enlisting commensal microbes to resist anti biotic-resistant pathogens. Journal of Experimental Medicine, 2018, jem-20180399.

Kho, Z. Y., & Lal, S. K. (2018). The human gut microbiome – A potential controller of wellness and disease. Frontiers in Microbiology, 9, 1835.

Knysh, O. V., Isajenko, O. J., Babych, J. M., Poljans'ka, V. P., Zachepylo, S. V., Kompanijec', A. M., & Gorbach, T. V. (2018). Sposib oderzhannja biolo gichno aktyvnyh deryvativ bakterij probiotychnyh shtamiv [Method for obtai ning biologically active derivatives of bacteria of probiotic strains]. Patent of Ukraine for useful model No 122859. Derzhavne Patentne Vidomstvo Ukrainy, Kyiv (in Ukrainian).

Ku, S., Park, M., Ji, G., & You, H. (2016). Review on Bifidobacterium bifidum bgn4: Functionality and nutraceutical applications as a probiotic microorga nism. International Journal of Molecular Sciences, 17(9), 1544.

Landman, C., & Quevrain, E. (2016). Gut microbiota: Description, role and patho physiologic implications. La Revue de MedecineInterne, 37(6), 418–423.

Lindquist, J. A., & Mertens, P. R. (2018). Cold shock proteins: From cellular mec hanisms to pathophysiology and disease. Cell Communication and Signaling, 16(1), 63.

Litvak, Y., Byndloss, M. X., Tsolis, R. M., & Bäumler, A. J. (2017). Dysbiotic Proteobacteria expansion: A microbial signature of epithelial dysfunction. Current Opinion in Microbiology, 39, 1–6.

O’Callaghan, A., & van Sinderen, D. (2016). Bifidobacteria and their role as members of the human gut microbiota. Frontiers in Microbiology, 7, 925.

O'Neill, I., Schofield, Z., & Hall, L. J. (2017). Exploring the role of the microbiota member Bifidobacterium in modulating immune-linked diseases. Emerging Topics in Life Sciences, 1(4), 333–349.

Patel, A. K., Michaud, P., Singhania, R. R., Soccol, C. R., & Pandey, A. (2010). Polysaccharides from probiotics: New developments as food additives. Food Technology and Biotechnology, 48(4), 451–463.

Patten, D. A., & Laws, A. P. (2015). Lactobacillus-produced exopolysaccharides and their potential health benefits: A review. Beneficial Microbes, 6(4), 457–471.

Polak-Berecka, M., Wasko, A., Szwajgier, D., & Choma, A. (2013). Bifidogenic and antioxidant activity of exopolysaccharides produced by Lactobacillus rhamnosus E/N cultivated on different carbon sources. Polish Journal of Microbiology, 62(2), 81–189.

Quigley, E. M. M. (2016). Bifidobacterium bifidum. In: Floch, M. H., Ringel, Y., & Walker, W. A. (Eds.). The microbiota in gastrointestinal pathophysiology: Implications for human health, prebiotics, probiotics, and dysbiosis. Elsevier Inc. Part B, Chaptter 14. Pp. 131–133.

Ranieri, M. R., Whitchurch, C. B., & Burrows, L. L. (2018). Mechanisms of biofilm stimulation by subinhibitory concentrations of antimicrobials. Current Opinion in Microbiology, 45, 164–169.

Richards, J. L., Yap, Y. A., McLeod, K. H., Mackay, C. R., & Mariño, E. (2016). Dietary metabolites and the gut microbiota: An alternative approach to control inflammatory and autoimmune diseases. Clinical and Translational Immuno logy, 5(5), e82.

Rivière, A., Selak, M., Lantin, D., Leroy, F., & de Vuyst, L. (2016). Bifidobacteria and butyrate-producing colon bacteria: Importance and strategies for their stimulation in the human gut. Frontiers in Microbiology, 28(7), 979.

Ryan, P. M., Ross, R. P., Fitzgerald, G. F., Caplice, N. M., & Stanton, C. (2015). Sugar-coated: Exopolysaccharide producing lactic acid bacteria for food and human health applications. Food and Function, 6(3), 679–693.

Sarikaya, H., Aslim, B., & Yuksekdag, Z. (2017). Assessment of anti-biofilm activity and bifidogenic growth stimulator (BGS) effect of lyophilized exopolysac charides (l-EPSs) from Lactobacilli strains. International Journal of Food Properties, 20(2), 362–371.

Sarkar, A., & Mandal, S. (2016). Bifidobacteria ‒ insight into clinical outcomes and mechanisms of its probiotic action. Microbiological Research, 192, 159–171.

Serafini, F., Turroni, F., Ruas-Madiedo, P., Lugli, G. A., Milani, C., Duranti, S., Zam boni, N., Bottacini, F., Sinderen, D., Margolles, A., & Ventura, M. (2014). Kefir fermented milk and kefiran promote growth of Bifidobacterium bifidum PRL2010 and modulate its gene expression. International Journal of Food Microbiology,178, 50–59.

Shenderov, B. A. (2013). Metabiotics: Novel idea or natural development of pro biotic conception. Microbial Ecology in Health and Disease, 24(1), 20399.

Singh, A., Vishwakarma, V., & Singhal, B. (2018). Metabiotics: The functional metabolic signatures of probiotics: Current state-of-art and future research priorities. Advances in Bioscience and Biotechnology, 9, 147–189.

Spinler, J., Auchtung, J., Brown, A., Boonma, P., Oezguen, N., & Ross, C., Luna, R. A., Runge, J., Versalovic, J., Peniche, A., Dann, S. M., Britton, R. A., Haag, A., & Savidge, T. C. (2017). Next-generation probiotics targeting Clostridium difficile through precursor-directed antimicrobial biosynthesis. Infection and Immunity, 85(10), e00303-17.

Turroni, S., Brigidi, P., Cavalli, A., & Candela, M. (2017). Microbiota – host transgenomic metabolism, bioactive molecules from the inside: Miniperspec tive. Journal of Medicinal Chemistry, 61(1), 47–61.

Valdes, A. M., Walter, J., Segal, E., & Spector, T. D. (2018). Role of the gut mic robiota in nutrition and health. British Medical Journal, 361, k2179.

Wang, B., Yao, M., Lv, L., Ling, Z., & Li, L. (2017). The human microbiota in health and disease. Engineering, 3(1), 71–82.

Warminska-Radyko, I., Laniewska-Moroz, L., & Babuchowski, A. (2002). Possi bilities for stimulation of Bifidobacterium growth by propionibacteria. Le Lait, 82(1), 113–121.

How to Cite
Knysh , O. V. (2019). Bifidogenic properties of cell-free extracts derived from probiotic strains of Bifidobacterium bifidum and Lactobacillus reuteri . Regulatory Mechanisms in Biosystems, 10(1), 124-128.

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