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Enterocin LD3 from Enterococcus hirae LD3 Inhibits the Growth of Salmonella enterica subsp. enterica serovar Typhimurium ATCC 13311 in Fruit Juice

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Abstract

In order to prevent the growth of pathogens in food, bacteriocins produced by various probiotic lactic acid bacteria have been recognized as potential substitutes of chemical preservatives. In this study, enterocin LD3 was purified from the cell-free supernatant of a food isolate, Enterococcus hirae LD3 using multistep chromatography. In the fruit juice, lethal concentration (LC50) of enterocin LD3 was found to be 260 µg/mL against Salmonella enterica subsp. enterica serovar Typhimurium ATCC 13311. The cells treated with enterocin LD3 were red colour indicating dead cells after propidium iodide staining, while untreated cells were found blue after staining with 4', 6-diamidino-2-phenylindole. The mechanism of cell killing was analyzed using infrared spectrum of cells treated with enterocin LD3 which was found altered in the range of 1,094.30 and 1,451.82 cm−1 corresponding to nucleic acids and phospholipids, respectively. The morphology of target cells were severely ruptured and lysed as observed under electron microscopy. Thus, the present study suggested that enterocin LD3 showed bactericidal activity against Salm. enterica subsp. enterica serovar Typhimurium ATCC 13311 and may be applied as a bio-preservative for the safety of fruit juices.

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References

  1. Hoffmann S, Scallan E (2017) Epidemiology, cost, and risk analysis of foodborne disease. In: Dodd CER, Aldsworth T, Stein RA, Cliver DO, Riemann HP (ed) Foodborne Diseases, 3rd edn. Academic Press pp 31–63. https://doi.org/10.1016/B978-0-12-385007-2.00002-4

  2. Law JW, Ab Mutalib NS, Chan KG, Lee LH (2015) Rapid methods for the detection of foodborne bacterial pathogens: Principles, applications, advantages and limitations. Front Microbiol 5:770. https://doi.org/10.3389/fmicb.2014.00770

    Article  PubMed  PubMed Central  Google Scholar 

  3. Liu H, Whitehouse CA, Li B (2018) Presence and persistence of Salmonella in water: The impact on microbial quality of water and food safety. Front Public Health 6:159. https://doi.org/10.3389/fpubh.2018.00159

    Article  PubMed  PubMed Central  Google Scholar 

  4. Kumar S, Kumar Y, Kumar G, Kumar G, Tahlan AK (2022) Non-typhoidal Salmonella infections across India: Emergence of a neglected group of enteric pathogens. J Taibah Univ Medical Sci 17:747–754. https://doi.org/10.1016/j.jtumed.2022.02.011

    Article  Google Scholar 

  5. Bisht A, Kamble MP, Choudhary P, Chaturvedi K, Kohli G, Juneja VK, Sehgal S, Taneja NK (2021) A surveillance of food borne disease outbreaks in India: 2009–2018. Food Control 121:107630. https://doi.org/10.1016/j.foodcont.2020.107630

    Article  Google Scholar 

  6. Kumar Y, Gupta N, Vaish VB, Gupta S (2016) Distribution trends & antibiogram pattern of Salmonella enterica serovar Newport in India. Indian J Med Res 144:82–86. https://doi.org/10.4103/0971-5916.193293,144(1),82

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Xiang YZ, Zhang YM, Liu YY, Zhang M, Lin LB, Zhang QL (2021) Purification, characterization, and antibacterial and antibiofilm activity of a novel bacteriocin against Salmonella Enteritidis. Food Control 127:108110. https://doi.org/10.1016/j.foodcont.2021.108110

    Article  CAS  Google Scholar 

  8. Meng F, Zhu X, Zhao H, Nie T, Lu F, Lu Z, Lu Y (2021) A class III bacteriocin with broad-spectrum antibacterial activity from Lactobacillus acidophilus NX2–6 and its preservation in milk and cheese. Food Control 121:107597. https://doi.org/10.1016/j.foodcont.2020.107597

    Article  CAS  Google Scholar 

  9. Aneja KR, Dhiman R, Aggarwal NK, Kumar V, Kaur M (2014) Microbes associated with freshly prepared juices of citrus and carrots. Int J Food Sci 2014:1–7. https://doi.org/10.1155/2014/408085

    Article  Google Scholar 

  10. Ahmed T, Das KK, Uddin MA (2018) The microbiological quality of commercial fruit juices-current perspectives. Bangladesh J Microbiol 35:128–133. https://doi.org/10.3329/bjm.v35i2.42643

    Article  Google Scholar 

  11. To HTA, Chhetri V, Settachaimongkon S, Prakitchaiwattana C (2022) Stress tolerance-Bacillus with a wide spectrum bacteriocin as an alternative approach for food bio-protective culture production. Food Control 133:108598. https://doi.org/10.1016/j.foodcont.2021.108598

    Article  CAS  Google Scholar 

  12. Sourri P, Tassou CC, Nychas GJE, Panagou EZ (2022) Fruit juice spoilage by Alicyclobacillus: Detection and control methods- A comprehensive review. Foods 11:747. https://doi.org/10.3390/foods11050747

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Avirvarei AC, Salanta LC, Pop CR, Mudura E, Pasqualone A, Anjos O, Barboza N, Usaga J, Darab CP, Burja-Udrea C, Zhao H (2023) Fruit-based fermented beverages: Contamination sources and emerging technologies applied to assure their safety. Foods 12:838. https://doi.org/10.3390/foods12040838

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Allen HK, Trachsel J, Looft T, Casey TA (2014) Finding alternatives to antibiotics. Ann N Y Acad Sci 1323:91–100. https://doi.org/10.1111/nyas.12468

    Article  PubMed  Google Scholar 

  15. Cotter PD, Ross RP, Hill C (2013) Bacteriocins - A viable alternative to antibiotics? Nat Rev Microbiol 11:95–105. https://doi.org/10.1038/nrmicro2937

    Article  CAS  PubMed  Google Scholar 

  16. Leon Madrazo A, Segura Campos MR (2020) Review of antimicrobial peptides as promoters of food safety: Limitations and possibilities within the food industry. J Food Saf 40:e12854. https://doi.org/10.1111/jfs.12854

    Article  CAS  Google Scholar 

  17. Gok Charyyev M, Ozden Tuncer B, Akpınar Kankaya D, Tuncer Y (2019) Bacteriocinogenic properties and safety evaluation of Enterococcus faecium YT52 isolated from Boza, a traditional cereal based fermented beverage. J Consum Prot Food Saf 14:41–53. https://doi.org/10.1007/s00003-019-01213-9

    Article  CAS  Google Scholar 

  18. Wang H, Zhang H, Zhang H, Jin J, Xie Y (2021) Outer membrane channel protein TolC regulates Escherichia coli K12 sensitivity to plantaricin BM-1 via the CpxR/CpxA two-component regulatory system. Probiotics Antimicrob Proteins 13:238–248. https://doi.org/10.1007/s12602-020-09671-6

    Article  CAS  PubMed  Google Scholar 

  19. Liang ZR, Hsiao HI, Jhang DJ (2020) Synergistic antibacterial effect of nisin, ethylenediaminetetraacetic acid, and sulfite on native microflora of fresh white shrimp during ice storage. J Food Saf 40:e12794. https://doi.org/10.1111/jfs.12794

    Article  CAS  Google Scholar 

  20. Silva CCG, Silva SPM, Ribeiro SC (2018) Application of bacteriocins and protective cultures in dairy food preservation. Front Microbiol 9:594. https://doi.org/10.3389/fmicb.2018.00594

    Article  PubMed  PubMed Central  Google Scholar 

  21. Cotter PD, Hill C, Ross RP (2005) Bacteriocins: Developing innate immunity for food. Nat Rev Microbiol 3:777–788. https://doi.org/10.1038/nrmicro1273

    Article  CAS  PubMed  Google Scholar 

  22. Egan K, Field D, Rea MC, Ross RP, Hill C, Cotter PD (2016) Bacteriocins: Novel solutions to age-old spore-related problems? Front Microbiol 7:461. https://doi.org/10.3389/fmicb.2016.00461

    Article  PubMed  PubMed Central  Google Scholar 

  23. Gupta A, Tiwari SK (2015) Probiotic potential of bacteriocin-producing Enterococcus hirae strain LD3 isolated from Dosa batter. Ann Microbiol 65:2333–2342. https://doi.org/10.1007/s13213-015-1075-4

    Article  CAS  Google Scholar 

  24. Gupta A, Tiwari SK, Netrebov V, Chikindas ML (2016) Biochemical properties and mechanism of action of enterocin LD3 purified from Enterococcus hirae LD3. Probiotics Antimicrob Proteins 8:161–169. https://doi.org/10.1007/s12602-016-9217-y

    Article  CAS  PubMed  Google Scholar 

  25. Sheoran P, Tiwari SK (2021) Synergistically-acting enterocin LD3 and plantaricin LD4 against Gram-positive and Gram-negative pathogenic bacteria. Probiotics Antimicrob Proteins 13:542–554. https://doi.org/10.1007/s12602-020-09708-w

    Article  CAS  PubMed  Google Scholar 

  26. Sheoran P, Tiwari SK (2019) Enterocin LD3 from Enterococcus hirae LD3 causing efflux of intracellular ions and UV-absorbing materials in Gram-negative bacteria. J Appl Microbiol 126:1059–1069. https://doi.org/10.1111/jam.14203

    Article  CAS  PubMed  Google Scholar 

  27. Podolak R, Enache E, Stone W, Black DG, Elliott PH (2010) Sources and risk factors for contamination, survival, persistence, and heat resistance of Salmonella in low-moisture foods. J Food Prot 73:1919–1936. https://doi.org/10.4315/0362-028X-73.10.1919

    Article  PubMed  Google Scholar 

  28. Zhao X, Shi C, Meng R, Liu Z, Huang Y, Zhao Z, Guo N (2016) Effect of nisin and perilla oil combination against Listeria monocytogenes and Staphylococcus aureus in milk. J Food Sci Technol 53:2644–2653. https://doi.org/10.1007/s13197-016-2236-6

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Starr T, Bauler TJ, Malik-Kale P, Steele-Mortimer O (2018) The phorbol 12-myristate-13-acetate differentiation protocol is critical to the interaction of THP-1 macrophages with Salmonella Typhimurium. PLoS ONE 13:e0193601. https://doi.org/10.1371/journal.pone.0193601

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Zoumpopoulou G, Pepelassi E, Papaioannou W, Georgalaki M, Maragkoudakis PA, Tarantilis PA, Polissiou M, Tsakalidou E, Papadimitriou K (2013) Incidence of bacteriocins produced by food-related lactic acid bacteria active towards oral pathogens. Int J Mol Sci 14:4640–4654. https://doi.org/10.3390/ijms14034640

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Amit SK, Uddin MM, Rahman R, Islam SMR, Khan MS (2017) A review on mechanisms and commercial aspects of food preservation and processing. Agric Food Secur 6:51. https://doi.org/10.1186/s40066-017-0130-8

    Article  Google Scholar 

  32. Haddad-Kashani H, Nikzad H, Mobaseri S, Hoseini ES (2012) Synergism effect of nisin peptide in reducing chemical preservatives in food industry. Life Sci J 9:496–501

    Google Scholar 

  33. Ghosh T, Purkait A, Garai B, Banerjee A (2021) Microbial production of bacteriocins and its applications in food preservation: A review. J Food Process Preserv 5:1–2

    CAS  Google Scholar 

  34. Pei J, Yue T, Yuan Y, Dai L (2017) Activity of paracin C from lactic acid bacteria against Alicyclobacillus in apple juice: Application of a novelty bacteriocin. J Food Saf 37:e12350. https://doi.org/10.1111/ijsa.12350

    Article  Google Scholar 

  35. Terabayashi Y, Juan A, Tamotsu H, Ashimine N, Nakano K, Shimoji M, Shiroma A, Teruya K, Satou K, Hirano T (2014) First complete genome sequence of Salmonella enterica subsp. enterica serovar Typhimurium strain ATCC 13311 (NCTC 74), a reference strain of multidrug resistance, as achieved by use of PacBio single-molecule real-time technology. Genome Announc 2:e00986-e1014. https://doi.org/10.1128/genomeA.00986-14

    Article  PubMed  PubMed Central  Google Scholar 

  36. Ferreira AE, Canal N, Morales D, Fuentefria DB, Corção G (2007) Characterization of enterocins produced by Enterococcus mundtii isolated from human feces. Braz Arch Biol Technol 50:249–258. https://doi.org/10.1590/S1516-89132007000200010

    Article  Google Scholar 

  37. Viedma PM, López AS, Omar NB, Abriouel H, López RL, Valdivia E, Belloso OM, Gálvez A (2008) Enhanced bactericidal effect of enterocin AS-48 in combination with high-intensity pulsed-electric field treatment against Salmonella enterica in apple juice. Int J Food Microbiol 128:244–249. https://doi.org/10.1016/j.ijfoodmicro.2008.08.014

    Article  CAS  Google Scholar 

  38. Liang Z, Mittal GS, Griffith MW (2002) Inactivation of Salmonella Typhimurium in orange juice containing antimicrobial agents by pulsed electric field. J Food Prot 65:1081–1087. https://doi.org/10.4315/0362-028X-65.7.1081

    Article  CAS  PubMed  Google Scholar 

  39. Rosenberg M, Azevedo NF, Ivask A (2019) Propidium iodide staining underestimates viability of adherent bacterial cells. Sci Rep 9:1–12. https://doi.org/10.1038/s41598-019-42906-3

    Article  CAS  Google Scholar 

  40. Sheoran P, Tiwari SK (2019) Anti-staphylococcal activity of bacteriocins of food isolates Enterococcus hirae LD3 and Lactobacillus plantarum LD4 in pasteurized milk. 3 Biotech 9:8. https://doi.org/10.1007/s13205-018-1546-y

    Article  PubMed  PubMed Central  Google Scholar 

  41. Klubthawee N, Aunpad R (2021) A thermostable, modified cathelicidin-derived peptide with enhanced membrane-active activity against Salmonella enterica serovar Typhimurium. Front Microbiol 11:592220. https://doi.org/10.3389/fmicb.2020.592220

    Article  PubMed  PubMed Central  Google Scholar 

  42. Gong HS, Meng XC, Wang H (2010) Mode of action of plantaricin MG, a bacteriocin active against Salmonella typhimurium. J Basic Microbiol 50:S37–S45. https://doi.org/10.1002/jobm.201000130

    Article  PubMed  Google Scholar 

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Acknowledgements

The Department of Genetics at M. D. University Rohtak provided support for MKY and IK through its University Research Scholarship.

Funding

The Department of Biotechnology (BT/PR8911/NDB/39/423/2013 and BT/PR8306/PID/6/738/2013), Indian Council of Medical Research (5/9/1117/2013-NUT), New Delhi, India and University Grants Commission-Scheme for Trans-disciplinary Research for India’s Developing Economy Component-1 (UGC-STRIDE-1), New Delhi, India [F.2–16/2019 (STRIDE 1)] provided financial support for this work.

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The experiments, data collection and analysis and designed the manuscript were performed by PS and MKY. The manuscript was formatted by IK. SKT, as corresponding authors, provided all laboratory facilities and chemicals and reviewed the manuscript. All authors read and approved the final version of the manuscript.

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Correspondence to Santosh Kumar Tiwari.

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Sheoran, P., Yadav, M.K., Kumari, I. et al. Enterocin LD3 from Enterococcus hirae LD3 Inhibits the Growth of Salmonella enterica subsp. enterica serovar Typhimurium ATCC 13311 in Fruit Juice. Probiotics & Antimicro. Prot. (2023). https://doi.org/10.1007/s12602-023-10108-z

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