Applied Microbiology and Biotechnology

, Volume 99, Issue 10, pp 4343–4353 | Cite as

Relationships between the genome and some phenotypical properties of Lactobacillus fermentum CECT 5716, a probiotic strain isolated from human milk

  • Nivia Cárdenas
  • Jonathan E. Laiño
  • Susana Delgado
  • Esther Jiménez
  • Marianela Juárez del Valle
  • Graciela Savoy de Giori
  • Fernando Sesma
  • Baltasar Mayo
  • Leónides Fernández
  • Jean Guy LeBlanc
  • Juan M. RodríguezEmail author
Applied genetics and molecular biotechnology


Lactobacillus fermentum CECT 5716, isolated from human milk, has immunomodulatory, anti-inflammatory, and anti-infectious properties, as revealed by several in vitro and in vivo assays, which suggests a strong potential as a probiotic strain. In this work, some phenotypic properties of L. fermentum CECT 5716 were evaluated, and the genetic basis for the obtained results was searched for in the strain genome. L. fermentum CECT 5716 does not contain plasmids and showed neither bacteriocin nor biogenic amine biosynthesis ability but was able to produce organic acids, glutathione, riboflavin, and folates and to moderately stimulate the maturation of mouse dendritic cells. No prophages could be induced, and the strain was sensitive to all antibiotics proposed by European Food Safety Authority (EFSA) standards, while no transmissible genes potentially involved in antibiotic resistance were detected in its genome. Globally, there was an agreement between the phenotype properties of L. fermentum CECT 5716 and the genetic information contained in its genome.


Lactobacillus fermentum Genome Glutathione Riboflavin Folate Antibiotic resistance Dendritic cells Human milk 



This work was supported by CSD2007-00063 (FUN-C-FOOD, Consolider-Ingenio 2010), CYTED-IBEROFUN, and AGL2013-41980-P projects from the Ministerio de Ciencia e Innovación (Spain), a research contract funded by Biosearch Life (Granada, Spain) and by funding from the Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET) and Agencia Nacional de Promoción Científica y Tecnológica (ANPCyT), Argentina.

Supplementary material

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  1. Ardite E, Sans M, Panes J, Romero FJ, Pique JM, Fernandez-Checa JC (2000) Replenishment of glutathione levels improves mucosal function in experimental colitis. Lab Invest 80:735–744CrossRefPubMedGoogle Scholar
  2. Arribas B, Rodriguez-Cabezas ME, Comalada M, Bailon E, Camuesco D, Olivares M, Xaus J, Zarzuelo A, Gálvez J (2009) Evaluation of the preventative effects exerted by Lactobacillus fermentum in an experimental model of septic shock induced in mice. Br J Nutr 101:2–4CrossRefGoogle Scholar
  3. Asrar FM, O’Connor DL (2005) Bacterially synthesized folate and supplemental folic acid are absorbed across the large intestine of piglets. J Nutr Biochem 16:587–593CrossRefPubMedGoogle Scholar
  4. Bañuelos O, Fernández L, Corral JM, Valdivieso-Ugarte M, Adrio JL, Velasco J (2008) Metabolism of prebiotic products containing beta(2–1) fructan mixtures by two Lactobacillus strains. Anaerobe 14:184–189CrossRefPubMedGoogle Scholar
  5. Bover-Cid S, Holzapfel WH (1999) Improved screening procedure for biogenic amine production by lactic acid bacteria. Int J Food Microbiol 53:33–41CrossRefPubMedGoogle Scholar
  6. Camuesco D, Comalada M, Rodriguez-Cabezas ME, Nieto A, Lorente MD, Concha A, Zarzuelo A, Galvez J (2004) The intestinal anti-inflammatory effect of quercitrin is associated with an inhibition in iNOS expression. Br J Pharmacol 143:908–918CrossRefPubMedCentralPubMedGoogle Scholar
  7. Cárdenas N, Calzada J, Peirotén A, Jiménez E, Escudero R, Rodríguez JM, Medina M, Fernández L (2014) Development of a potential probiotic fresh cheese using two Lactobacillus salivarius strains isolated from human milk. Biomed Res Int. doi: 10.1155/2014/801918 PubMedCentralPubMedGoogle Scholar
  8. Church FC, Swaisgood HE, Porter DH, Catignain GL (1983) Spectrophotometric assay using o-phthaldialdehyde for determination of proteolysis in milk and isolated milk proteins. J Dairy Sci 66:1219–1227CrossRefGoogle Scholar
  9. Connell SR, Tracz DM, Nierhaus KH, Taylor DE (2003) Ribosomal protection proteins and their mechanism of tetracycline resistance. Antimicrob Agents Chemother 47:3675–3681CrossRefPubMedCentralPubMedGoogle Scholar
  10. Cueva C, Moreno-Arribas MV, Martín-Alvarez PJ, Bills G, Vicente MF, Basilio A, Rivas CL, Requena T, Rodríguez JM, Bartolomé B (2010) Antimicrobial activity of phenolic acids against commensal, probiotic and pathogenic bacteria. Res Microbiol 161:372–382CrossRefPubMedGoogle Scholar
  11. Danielsen M, Wind A (2003) Susceptibility of Lactobacillus spp. to antimicrobial agents. Int J Food Microbiol 82:1–11CrossRefPubMedGoogle Scholar
  12. Díaz-Ropero M, Martin R, Sierra S, Lara-Villoslada F, Rodriguez JM, Xaus J, Olivares M (2006) Two Lactobacillus strains, isolated from breast milk, differently modulate the immune response. J Appl Microbiol 102:337–343Google Scholar
  13. Dudeja PK, Kode A, Alnounou M, Tyagi S, Torania S, Subramanian VS, Said HM (2001) Mechanism of folate transport across the human colonic basolateral membrane. Am J Physiol Gastrointest Liver Physiol 281:G54–60PubMedGoogle Scholar
  14. EFSA (2012) Guidance on the assessment of bacterial susceptibility to antimicrobials of human and veterinary importance. EFSA J 10:2740Google Scholar
  15. Egervarn M, Danielsen M, Roos S, Lindmark H, Lindgren S (2007) Antibiotic susceptibility profiles of Lactobacillus reuteri and Lactobacillus fermentum. J Food Prot 70:412–418PubMedGoogle Scholar
  16. Fernández L, Langa S, Martín V, Maldonado A, Jiménez E, Martín R, Rodríguez JM (2013) The human milk microbiota: origin and potential roles in health and disease. Pharmacol Res 69:1–10CrossRefPubMedGoogle Scholar
  17. Gil-Campos M, López MÁ, Rodriguez-Benítez MV, Romero J, Roncero I, Linares MD, Maldonado J, López-Huertas E, Berwind R, Ritzenthaler KL, Navas V, Sierra C, Sempere L, Geerlings A, Maldonado-Lobón JA, Valero AD, Lara-Villoslada F, Olivares M (2012) Lactobacillus fermentum CECT 5716 is safe and well tolerated in infants of 1–6 months of age: a randomized controlled trial. Pharmacol Res 65:231–238CrossRefPubMedGoogle Scholar
  18. Gonzalez R, Sanchez de Medina F, Galvez J, Rodriguez-Cabezas ME, Duarte J, Zarzuelo A (2001) Dietary vitamin E supplementation protects the rat large intestine from experimental inflammation. Int J Vitam Nutr Res 71:243–250CrossRefPubMedGoogle Scholar
  19. Grover S, Sharma VK, Mallapa RH, Batish VK (2013) Draft genome sequence of Lactobacillus fermentum Lf1, an Indian isolate of human gut origin. Genome Announc 1:e00883–13PubMedCentralPubMedGoogle Scholar
  20. Horwitz W (2000) Official methods of analysis of AOAC international. AOAC International, GaithersburgGoogle Scholar
  21. Jayashree S, Rajendhran J, Jayaraman K, Kalaichelvan G, Gunasekaran P (2011) Improvement of riboflavin production by Lactobacillus fermentum isolated from yogurt. Food Biotechnol 25:240–251CrossRefGoogle Scholar
  22. Jeurink PV, van Bergenhenegouwen J, Jiménez E, Knippels LM, Fernández L, Garssen J, Knol J, Rodríguez JM, Martín R (2013) Human milk: a source of more life than we imagine. Ben Microbes 4:17–30CrossRefGoogle Scholar
  23. Jiménez E, Langa S, Martín V, Arroyo R, Martín R, Fernández L, Rodríguez JM (2010) Complete genome sequence of Lactobacillus fermentum CECT 5716, a probiotic strain isolated from human milk. J Bacteriol 192:4800CrossRefPubMedCentralPubMedGoogle Scholar
  24. Juarez del Valle M, Laiño J, Savoy de Giori G, LeBlanc JG (2014) Use of lactic acid bacteria as a biotechnological strategy to increase riboflavin levels in soymilk. Food Res Int 62:1015–1019CrossRefGoogle Scholar
  25. Katla A-K, Kruse H, Johnsen G, Herikstad H (2001) Antimicrobial susceptibility of starter culture bacteria used in Norwegian dairy products. Int J Food Microbiol 67:147e52CrossRefGoogle Scholar
  26. Krause LJ, Forsberg CW, O’Connor DL (1996) Feeding human milk to rats increases Bifidobacterium in the cecum and colon which correlates with enhanced folate status. J Nutr 126:1505–1511PubMedGoogle Scholar
  27. Kullisaar T, Zilmer M, Mikelsaar M, Vihalemm T, Annuk H, Kairane C, Kilk A (2002) Two antioxidative lactobacilli strains as promising probiotics. Int J Food Microbiol 72:215–224CrossRefPubMedGoogle Scholar
  28. Laiño JE, LeBlanc JG, Savoy de Giori GS (2012) Selection of folate producing starter cultures of yogurt isolated from Northwestern Argentina. Can J Microbiol 58:581–588CrossRefPubMedGoogle Scholar
  29. Langa S, Maldonado A, Delgado S, Martín R, Martín V, Jiménez E, Ruíz-Barba JL, Mayo B, Connor RI, Suárez JE, Rodríguez JM (2012) Characterization of Lactobacillus salivarius CECT 5713, a strain isolated from human milk: from genotype to phenotype. Appl Microbiol Biotechnol 94:1279–1287CrossRefPubMedGoogle Scholar
  30. Lara-Villoslada F, Sierra S, Díaz-Ropero MP, Rodríguez JM, Xaus J, Olivares M (2009) Safety assessment of Lactobacillus fermentum CECT5716, a probiotic strain isolated from human milk. J Dairy Res 76:216–221CrossRefPubMedGoogle Scholar
  31. LeBlanc JG, Piard JC, Sesma F, de Giori GS (2005) Lactobacillus fermentum CRL 722 is able to deliver active alpha-galactosidase activity in the small intestine of rats. FEMS Microbiol Lett 248:177–182CrossRefPubMedGoogle Scholar
  32. LeBlanc JG, Giori GSD, Smid EJ, Hugenholtz J, Sesma F (2007) Folate production by lactic acid bacteria and other food-grade microorganisms. Commun Curr Res Educ Topics Trends Appl Microbiol 1:329–339Google Scholar
  33. LeBlanc JG, Ledue-Clier F, Bensaada M, de Giori GS, Guerekobaya T, Sesma F, Juillard V, Rabot S, Piard JC (2008) Ability of Lactobacillus fermentum to overcome host alpha-galactosidase deficiency, as evidenced by reduction of hydrogen excretion in rats consuming soya alpha-galacto-oligosaccharides. BMC Microbiol 8:22CrossRefPubMedCentralPubMedGoogle Scholar
  34. LeBlanc JG, Milani C, Savoy de Giori GS, Sesma F, van Sinderen D, Ventura M (2013) Bacteria as vitamin suppliers to their host: a gut microbiota perspective. Curr Opin Biotechnol 24:160–168CrossRefPubMedGoogle Scholar
  35. Loguercio C, D’Argenio G, Delle Cave M, Cosenza V, Della Valle N, Mazzacca G, Del Vecchio BC (2003) Glutathione supplementation improves oxidative damage in experimental colitis. Dig Liver Dis 35:635–641CrossRefPubMedGoogle Scholar
  36. Lu L, Hsieh M, Oriss TB, Morelf PA, Starzl TE, Rao AS, Thomson AW (1995) Generation of DC from mouse spleen cell cultures in response to GM-CSF: immunophenotypic and functional analyses. Immunology 84:127–134PubMedCentralPubMedGoogle Scholar
  37. Maldonado J, Cañabate F, Sempere L, Vela F, Sánchez AR, Narbona E, López-Huertas E, Geerlings A, Valero AD, Olivares M, Lara-Villoslada F (2012) Human milk probiotic Lactobacillus fermentum CECT5716 reduces the incidence of gastrointestinal and upper respiratory tract infections in infants. J Pediatr Gastroenterol Nutr 54:55–61CrossRefPubMedGoogle Scholar
  38. Mañé J, Lorén V, Pedrosa E, Ojanguren I, Xaus J, Cabré E, Domènech E, Gassull MA (2009) Lactobacillus fermentum CECT5716 prevents and reverts intestinal damage on TNBS-induced colitis in mice. Inflamm Bowel Dis 15:1155–1163CrossRefPubMedGoogle Scholar
  39. Martín R, Langa S, Reviriego C, Jimenez E, Marin ML, Xaus J, Fernández L, Rodríguez JM (2003) Human milk is a source of lactic acid bacteria for the infant gut. J Pediatr 143:754–758CrossRefPubMedGoogle Scholar
  40. Martin R, Olivares M, Marin ML, Fernandez L, Xaus J, Rodríguez JM (2005) Probiotic potential of 3 lactobacilli strains isolated from breast milk. J Hum Lact 21:8–17CrossRefPubMedGoogle Scholar
  41. Martín R, Olivares M, Marín ML, Xaus J, Fernández L, Rodríguez JM (2005) Characterization of a reuterin-producing Lactobacillus coryniformis strain isolated from a goat’s milk cheese. Int J Food Microbiol 104:267–277CrossRefPubMedGoogle Scholar
  42. Martín R, Olivares M, Pérez M, Xaus J, Torre C, Fernández L, Rodríguez JM (2010) Identification and evaluation of the probiotic potential of lactobacilli isolated from canine milk. Vet J 185:193–198CrossRefPubMedGoogle Scholar
  43. Miralles-Barrachina O, Savoye G, Belmonte-Zalar L, Hochain P, Ducrotte P, Hecketsweiler B, Lerebours E, Dechelotte P (1999) Low levels of glutathione in endoscopic biopsies of patients with Crohn’s colitis: the role of malnutrition. Clin Nutr 18:313–317CrossRefPubMedGoogle Scholar
  44. Morita H, Toh H, Fukuda S, Horikawa H, Oshima K, Suzuki T, Murakami M, Hisamatsu S, Kato Y, Takizawa T, Fukuoka H, Yoshimura T, Itoh K, O’Sullivan DJ, McKay LL, Ohno H, Kikuchi J, Masaoka T, Hattori M (2008) Comparative genome analysis of Lactobacillus reuteri and Lactobacillus fermentum reveal a genomic island for reuterin and cobalamin production. DNA Res 15:151–161CrossRefPubMedCentralPubMedGoogle Scholar
  45. Olivares M, Díaz-Ropero MP, Martín R, Rodríguez JM, Xaus J (2006) Antimicrobial potential of four Lactobacillus strains isolated from breast milk. J Appl Microbiol 101:72–79CrossRefPubMedGoogle Scholar
  46. Olivares M, Díaz-Ropero MP, Sierra S, Lara-Villoslada F, Fonollá J, Navas M, Rodríguez JM, Xaus J (2007) Oral intake of Lactobacillus fermentum CECT5716 enhances the effects of influenza vaccination. Nutrition 23:254–260CrossRefPubMedGoogle Scholar
  47. Peran L, Camuesco D, Comalada M, Nieto A, Concha A, Adrio JL, Olivares M, Xaus J, Zarzuelo A, Galvez J (2006) Lactobacillus fermentum, a probiotic capable to release glutathione, prevents colonic inflammation in the TNBS model of rat colitis. Int J Colorectal Dis 21:737–746CrossRefPubMedGoogle Scholar
  48. Pérez-Cano FJ, Dong H, Yaqoob P (2010) In vitro immunomodulatory activity of Lactobacillus fermentum CECT5716 and Lactobacillus salivarius CECT5713: two probiotic strains isolated from human breast milk. Immunobiology 12:996–1004CrossRefGoogle Scholar
  49. Piddock LJ (2006) Multidrug-resistance efflux pumps—not just for resistance. Nat Rev Microbiol 4:629–636CrossRefPubMedGoogle Scholar
  50. Santos F, Vera JL, van der Heijden R, Valdez G, de Vos WM, Sesma F, Hugenholtz J (2008) The complete coenzyme B12 biosynthesis gene cluster of Lactobacillus reuteri CRL1098. Microbiology 154:81–93CrossRefPubMedGoogle Scholar
  51. Song Y-L, Kato N, Matsumiya Y, Liu C-X, Kato H, Watanabe K (1999) Identification of and hydrogen peroxide production by fecal and vaginal lactobacilli isolated from Japanese women and newborn infants. J Clin Microbiol 37:3062–3064PubMedCentralPubMedGoogle Scholar
  52. Sybesma W, Starrenburg M, Tijsseling L, Hoefnagel MH, Hugenholtz J (2003) Effects of cultivation conditions on folate production by lactic acid bacteria. Appl Environ Microbiol 69:4542–4548CrossRefPubMedCentralPubMedGoogle Scholar
  53. Taranto MP, Vera JL, Hugenholtz J, De Valdez GF, Sesma F (2003) Lactobacillus reuteri CRL1098 produces cobalamin. J Bacteriol 185:5643–5647CrossRefPubMedCentralPubMedGoogle Scholar
  54. Yap PS, Gilliland SE (2000) Comparison of newly isolated strains of Lactobacillus delbrueckii susp. lactis for hydrogen peroxide production at 5°C. J Dairy Sci 83:628–632CrossRefPubMedGoogle Scholar
  55. Zhang Z, Schwartz S, Wagner L, Miller W (2000) A greedy algorithm for aligning DNA sequences. J Comput Biol 7:203–214CrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2015

Authors and Affiliations

  • Nivia Cárdenas
    • 1
    • 2
  • Jonathan E. Laiño
    • 3
  • Susana Delgado
    • 4
  • Esther Jiménez
    • 1
    • 2
  • Marianela Juárez del Valle
    • 3
  • Graciela Savoy de Giori
    • 3
  • Fernando Sesma
    • 3
  • Baltasar Mayo
    • 4
  • Leónides Fernández
    • 1
    • 2
  • Jean Guy LeBlanc
    • 3
  • Juan M. Rodríguez
    • 1
    • 2
    Email author
  1. 1.Departamento de Nutrición, Bromatología y Tecnología de los AlimentosUniversidad Complutense de MadridMadridSpain
  2. 2.ProbisearchMadridSpain
  3. 3.CERELA-CONICET, Centro de Referencia para LactobacilosSan Miguel de TucumánArgentina
  4. 4.Instituto de Productos Lácteos de Asturias (IPLA-CSIC)VillaviciosaSpain

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