Safety and Stability of Two Potentially Probiotic Lactobacillus Strains After In Vitro Gastrointestinal Transit

  • Wilson José Fernandes Lemos Junior
  • André Fioravante Guerra
  • Armin Tarrah
  • Vinícius da Silva Duarte
  • Alessio GiacominiEmail author
  • Rosa Helena Luchese
  • Viviana Corich


According to FAO and WHO, probiotics are defined as live microorganisms that, when administered in adequate amounts, confer a health benefit on the host. Most probiotic bacteria used today belong to the genera Lactobacillus and Bifidobacterium and are of animal or human origin. The fundamental characteristic routinely evaluated in potential probiotics strains is their limited viability loss during gastrointestinal transit (GIT), but to date, no studies reported whether probiotics, besides viability, still also maintain their beneficial properties intact. To study this aspect, we considered two strains, Lactobacillus rhamnosus DTA 79 and L. paracasei DTA 83, previously characterised for the presence of some probiotic properties, isolated from faeces of 7- to 21-day-old babies. Here, we examined some additional properties, namely antibiotic resistance, resistance to lysozyme, presence of haemolytic activity and inhibition of pathogen biofilm formation. We then tested the effect of in vitro GIT on all these features and our results show evidence that this procedure had in some cases limited and in others no significant effects on them. Additionally, we examined the gastrointestinal resistance of the strains after skim milk fermentation and successive storage of the product for 20 and 40 days at refrigeration temperature, to see whether prolonged storage could weaken cell resistance to GIT. Our results demonstrate that a protracted refrigeration period before in vitro GIT did not affect or influenced very weakly this essential probiotic property.


Lactobacillus rhamnosus Lactobacillus paracasei Probiotics Dairy Biofilm 


Funding Information

WJFLJ was financially supported by CAPES—Coordenação de Aperfeiçoamento de Pessoal de Nível Superior.

Compliance with Ethical Standards

Conflict of Interest

The authors declare that they have no conflicts of interest.


  1. 1.
    Caballero-Franco C, Keller K, De Simone C, Chadee K (2007) The VSL#3 probiotic formula induces mucin gene expression and secretion in colonic epithelial cells. Am J Physiol Liver Physiol 292:G315–G322. Google Scholar
  2. 2.
    Padmavathi T, Bhargavi R, Priyanka PR, Niranjan NR, Veerabhadrappa PP (2018) Screening of potential probiotic lactic acid bacteria and production of amylase and its partial purification. J Genet Eng Biotechnol 16:357–362. CrossRefGoogle Scholar
  3. 3.
    Tarrah A, de CJ, Rossi RC, da Silva Duarte V, Righetto Ziegler D, Corich V, Giacomini A (2018) In vitro probiotic potential and anti-cancer activity of newly isolated folate-producing Streptococcus thermophilus strains. Front Microbiol 9:1–11. CrossRefGoogle Scholar
  4. 4.
    Ouwehand AC, Salminen S, Isolauri E (2002) Probiotics: an overview of beneficial effects. Antonie Van Leeuwenhoek 82:279–289CrossRefGoogle Scholar
  5. 5.
    Rezac S, Kok CR, Heermann M, Hutkins R (2018) Fermented foods as a dietary source of live organisms. Front Microbiol 9:1785. CrossRefGoogle Scholar
  6. 6.
    Pogačić T, Samaržija D, Corich V, D’Andrea M, Kagkli DM, Giacomini A, Majhenič AC, Rogelj I (2010) Microbiota of Karakačanski skakutanac, an artisanal fresh sheep cheese studied by culture-independent PCR-ARDRA and PCR-DGGE. Dairy Sci Technol 90:461–468. CrossRefGoogle Scholar
  7. 7.
    Campanaro S, Treu L, Vendramin V, Bovo B, Giacomini A, Corich V (2014) Metagenomic analysis of the microbial community in fermented grape marc reveals that Lactobacillus fabifermentans is one of the dominant species: insights into its genome structure. Appl Microbiol Biotechnol 98:6015–6037. CrossRefGoogle Scholar
  8. 8.
    Kim H-J, Lee HJ, Lim B, Kim E, Kim HY, Suh M, Hur M (2018) Lactobacillus terrae sp. nov., a novel species isolated from soil samples in the Republic of Korea. Int J Syst Evol Microbiol 68:2906–2911. CrossRefGoogle Scholar
  9. 9.
    Grumezescu AM, Holban AM (eds) (2018) Diet, microbiome and health. Academic PressGoogle Scholar
  10. 10.
    Tarrah A, da Silva Duarte V, de Castilhos J, Pakroo S, Lemos Junior WJF, Luchese RH, Guerra AF, Rossi RC, Righetto Ziegler D, Corich V, Giacomini A (2019) Probiotic potential and biofilm inhibitory activity of Lactobacillus casei group strains isolated from infant feces. J Funct Foods 54:489–497. CrossRefGoogle Scholar
  11. 11.
    Alessandri G, Milani C, Duranti S, Mancabelli L, Ranjanoro T, Modica S, Carnevali L, Statello R, Bottacini F, Turroni F, Ossiprandi MC, Sgoifo A,- van Sinderen D, Ventura M (2019) Ability of bifidobacteria to metabolize chitin-glucan and its impact on the gut microbiota. Sci Rep 9:5755. CrossRefGoogle Scholar
  12. 12.
    Saarela M, Mogensen G, Fondén R, Mättö J, Sandholm TM (2000) Probiotic bacteria: safety, functional and technological properties. J Biotechnol 84:197–215. CrossRefGoogle Scholar
  13. 13.
    FAO/WHO (2006) Probiotics in food: health and nutritional properties and guidelines for evaluation. FAO Food and Nutrition Paper 85. Available at: Accessed 12 June 2019
  14. 14.
    Millette M, Nguyen A, Amine KM, Lacroix M (2013) Gastrointestinal survival of bacteria in commercial probiotic products. Int J Probiotics Prebiotics 8:149–156Google Scholar
  15. 15.
    EFSA (2016) Update of the list of QPS-recommended biological agents intentionally added to food or feed as notified to EFSA 4: suitability of taxonomic units notified to EFSA until March 2016. EFSA J 14(7):e04522. Google Scholar
  16. 16.
    Lemos Junior WJF, Fioravante Guerra A, da Silva Duarte V, Treu L, Tarrah A, Campanaro S, Luchese RH, Giacomini A, Corich V (2019) Draft genome sequence data of Lactobacillus paracasei strain DTA83 isolated from infant stools. Data Br 22:1064–1067. CrossRefGoogle Scholar
  17. 17.
    Guerra AF, Lemos Junior WJF, dos SGO, Andrighetto C, Giacomini A, Corich V, Luchese RH (2018) Lactobacillus paracasei probiotic properties and survivability under stress-induced by processing and storage of ice cream bar or ice-lolly. Ciência Rural 48(9):e20170601. CrossRefGoogle Scholar
  18. 18.
    Pogačić T, D’Andrea M, Kagkli DM, Corich V, Giacomini A, Baldan E, Majhenič AC, Obermajer T, Rogelj I, Samaržija D (2011) Biodiversity of microbial consortia isolated from traditional fresh sheep cheese Karakačanski skakutanac. Mljekarstvo 61:208–219Google Scholar
  19. 19.
    Favarin L, Laureano-Melo R, Luchese RH (2015) Survival of free and microencapsulated Bifidobacterium: effect of honey addition. J Microencapsul 32:329–335. CrossRefGoogle Scholar
  20. 20.
    Charteris WP, Kelly PM, Morelli L, Collins JK (1998) Antibiotic susceptibility of potentially probiotic Lactobacillus species. J Food Prot 61:1636–1643CrossRefGoogle Scholar
  21. 21.
    Baldassarri L, Creti R, Recchia S, Imperi M, Facinelli B, Giovanetti E, Pataracchia M, Alfarone G, Orefici G (2006) Therapeutic failures of antibiotics used to treat macrolide-susceptible Streptococcus pyogenes infections may be due to biofilm formation. J Clin Microbiol 44:2721–2727. CrossRefGoogle Scholar
  22. 22.
    Beck BR, Park G-S, Lee YH, Im S, Jeong DY, Kang J (2019) Whole genome analysis of Lactobacillus plantarum strains isolated from kimchi and determination of probiotic properties to treat mucosal infections by Candida albicans and Gardnerella vaginalis. Front Microbiol 10:433. CrossRefGoogle Scholar
  23. 23.
    Fernández MF, Boris S, Barbes C (2003) Probiotic properties of human lactobacilli strains to be used in the gastrointestinal tract. J Appl Microbiol 94:449–455CrossRefGoogle Scholar
  24. 24.
    Zárate G, Chaia AP, González S, Oliver G (2000) Viability and-galactosidase activity of dairy propionibacteria subjected to digestion by artificial gastric and intestinal fluids. J Food Prot 63(9):1214–1221CrossRefGoogle Scholar
  25. 25.
    Flach J, van der Waal MB, van den Nieuwboer M, Claassen E, Larsen OFA (2018) The underexposed role of food matrices in probiotic products: reviewing the relationship between carrier matrices and product parameters. Crit Rev Food Sci Nutr 58:2570–2584. CrossRefGoogle Scholar
  26. 26.
    Nawaz M, Wang J, Zhou A, Ma C, Wu X, Moore JE, Millar BC, Xu J (2011) Characterization and transfer of antibiotic resistance in lactic acid bacteria from fermented food products. Curr Microbiol 62:1081–1089. CrossRefGoogle Scholar
  27. 27.
    EFSA (2012) Guidance on the assessment of bacterial susceptibility to antimicrobials of human and veterinary importance. EFSA J 10(6):2740. Google Scholar
  28. 28.
    Wang J, Guo H, Cao C, Zhao W, Kwok LY, Zhang H, Zhang W (2018) Characterization of the adaptive amoxicillin resistance of Lactobacillus casei Zhang by proteomic analysis. Front Microbiol 9:292. CrossRefGoogle Scholar
  29. 29.
    Ruiz J (2003) Mechanisms of resistance to quinolones: target alterations, decreased accumulation and DNA gyrase protection. J Antimicrob Chemother 51:1109–1117. CrossRefGoogle Scholar
  30. 30.
    Campedelli I, Mathur H, Salvetti E, Clarke S, Rea MC, Torriani S, Ross RP, Hill C, O’Toole PW (2018) Genus-wide assessment of antibiotic resistance in Lactobacillus spp. Appl Environ Microbiol 85(1):e01738–e01718. CrossRefGoogle Scholar
  31. 31.
    Yamaguchi A, Ohmori H, Kaneko-Ohdera M, Nomura T, Sawai T (1991) Delta pH-dependent accumulation of tetracycline in Escherichia coli. Antimicrob Agents Chemother 35:53–56CrossRefGoogle Scholar
  32. 32.
    Berger LR, Weiser RS (1957) The β-glucosaminidase activity of egg-white lysozyme. Biochim Biophys Acta 26:517–521. CrossRefGoogle Scholar
  33. 33.
    Valenta C, Schwarz E, Bernkop-Schnürch A (1998) Lysozyme-caffeic acid conjugates: possible novel preservatives for dermal formulations. Int J Pharm 174:125–132. CrossRefGoogle Scholar
  34. 34.
    Zago M, Fornasari ME, Carminati D, Burns P, Suàrez V, Vinderola G, Reinheimer J, Giraffa G (2011) Characterization and probiotic potential of Lactobacillus plantarum strains isolated from cheeses. Food Microbiol 28:1033–1040. CrossRefGoogle Scholar
  35. 35.
    Institute of Medicine (U.S.). Subcommittee on Nutrition during Lactation. Health Resources and Services Administration. (1991) Nutrition during lactation. National Academy PressGoogle Scholar
  36. 36.
    Tong J, Wei H, Liu X, Hu W, Bi M, Wang Y, Li Q, Li N (2011) Production of recombinant human lysozyme in the milk of transgenic pigs. Transgenic Res 20:417–419. CrossRefGoogle Scholar
  37. 37.
    Kumar M, Dhaka P, Vijay D, Vergis J, Mohan V, Kumar A, Kurkure NV, Barbuddhe SB, Malik SVS, Rawool DB (2016) Antimicrobial effects of Lactobacillus plantarum and Lactobacillus acidophilus against multidrug-resistant enteroaggregative Escherichia coli. Int J Antimicrob Agents 48:265–270. CrossRefGoogle Scholar
  38. 38.
    Gopalsamy SN, Woodworth MH, Wang T, Carpentieri CT, Mehta N, Friedman-Moraco RJ, Mehta AK, Larsen CP, Kraft CS (2018) The use of microbiome restoration therapeutics to eliminate intestinal colonization with multidrug-resistant organisms. Am J Med Sci 356:433–440. CrossRefGoogle Scholar
  39. 39.
    Ouwehand AC, Saxelins M, Salminen S (2004) Phenotypic differences between commercial Lactobacillus rhamnosus GG and L. rhamnosus strains recovered from blood. Clin Infect Dis 39:1858–1860. CrossRefGoogle Scholar
  40. 40.
    García A, Navarro K, Sanhueza E, Pineda S, Pastene E, Quezada M, Henríquez K, Karlyshev A, Villena J, González C (2017) Characterization of Lactobacillus fermentum UCO-979C, a probiotic strain with a potent anti-Helicobacter pylori activity. Electron J Biotechnol 25:75–83. CrossRefGoogle Scholar
  41. 41.
    WHO/FAO (2006) Probiotics in food health and nutritional properties and guidelines for evaluation. FAO Food Nutr Pap 85Google Scholar
  42. 42.
    Ziarno M, Zaręba D (2015) Effects of milk components and food additives on survival of three bifidobacteria strains in fermented milk under simulated gastrointestinal tract conditions. Microb Ecol Health Dis 26:27812. Google Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  • Wilson José Fernandes Lemos Junior
    • 1
  • André Fioravante Guerra
    • 2
  • Armin Tarrah
    • 3
  • Vinícius da Silva Duarte
    • 3
  • Alessio Giacomini
    • 3
    Email author
  • Rosa Helena Luchese
    • 4
  • Viviana Corich
    • 3
  1. 1.Department of BiotechnologyUniversity of VeronaVeronaItaly
  2. 2.Federal Center of Technological Education Celso Suckow da FonsecaValençaBrazil
  3. 3.Department of Agronomy Food Natural resources Animals and EnvironmentUniversity of PadovaLegnaroItaly
  4. 4.Department of Food TechnologyFederal Rural University of Rio de JaneiroSeropédicaBrazil

Personalised recommendations