DOI QR코드

DOI QR Code

Antimicrobial effect of Enterococcus faecalis BMSE-HMP005 isolated from human breast milk against multidrug-resistant bacteria

모유 유래 유산균 Enterococcus faecalis BMSE-HMP005의 다제내성 균에 대한 항균효과

  • Lee, Jeong-Eun (Department of Alternative Medicine, Kyonggi University) ;
  • Kim, Soo-bin (Department of Food and Nutrition, Chung-Ang University) ;
  • Yu, Du-na (Department of Biotechnology, Kyung Hee University) ;
  • Jo, So-Yeon (Department of Food & Nutrition, Baewha Women's University) ;
  • Kim, Ae-Jung (The Graduate School of Alternative Medicine, Kyonggi University) ;
  • Kook, Moochang (Department of Food & Nutrition, Baewha Women's University)
  • 이정은 (경기대학교 일반대학원 대체의학과) ;
  • 김수빈 (중앙대학교 식품영양학과) ;
  • 유두나 (경희대학교 생명공학원) ;
  • 조소연 (배화여자대학교 식품영양학과) ;
  • 김애정 (경기대학교 대체의학대학원) ;
  • 국무창 (배화여자대학교 식품영양학과)
  • Received : 2021.12.23
  • Accepted : 2022.02.21
  • Published : 2022.04.30

Abstract

In this study, Enterococcus faecalis BMSE-HMP005 isolated from human breast milk demonstrated antimicrobial effects against multidrug-resistant (MDR) bacterial strains. The bacteriocin produced by E. faecalis BMSE-HMP005 was fractionated using reverse-phase high-performance liquid chromatography. This fraction showed antimicrobial effects against both gram-positive and gram-negative MDR bacteria. No hemolytic reactions were observed. E. faecalis BMSEHMP005 was resistant to vancomycin; however, kanamycin, ampicillin, and erythromycin showed minimum inhibitory concentrations that were lower than the acceptable range provided by the European Food Safety Authority. For artificial gastric juice and bile acid, the survival rates were 98.67% and 95.70%, respectively. These results show the potential utility of E. faecalis BMSE-HMP005 as a probiotic with remarkable antimicrobial effects against MDR bacteria.

본 연구에서는 모유 유래 유산균 E. faecalis BMSE-HMP005의 다제내성 균주에 대한 항균효과 및 probiotics로써 잠재적인 가능성을 확인하였다. E. faecalis BMSE-HMP005는 다제내성 균주 20주(Enterococcus spp., Staphylococcus spp., Escherichia spp., Pseudomonas spp., Salmonella spp., Klebsiella spp., Enterobacter spp.)에서 모두 MBC가 확인되어 다제내성 균주에 대한 우수한 항균효과를 입증하였다. 또한 RP-HPLC를 이용하여 배양액 내 bacteriocin을 확인하였으며, 이에 대한 분획은 gram 양성 및 gram 음성균주에서 모두 항균력이 나타나, E. faecalis BMSE-HMP005가 생산하는 bacteriocin의 광범위한 항균 스펙트럼을 입증하였다. E. faecalis BMSE-HMP005는 발암 화합물을 유발하는 β-glucuronidase에 대한 활성과 용혈성은 나타나지 않아 안전한 것으로 판단된다. E. faecalis BMSE-HMP005는 vancomycin에 대해서는 내성을 보이나, kanamycin (>0.058), ampicillin (>0.002), erythromycin (>0.002)은 EFSA 기준의 허용범위보다 낮은 MIC가 확인되었다. 또한 인공위액(pH 2.0) 및 인공 담즙산(0.3% bile acid) 조건에서 각각 98.67%, 95.70%의 생존율을 보였다. 이와 같은 결과는 본 연구에서 분리한 모유 유래 유산균 E. faecalis BMSE-HMP005가 다제내성 균주에 대한 우수한 항균활성을 갖는 probiotics로써 잠재적인 가능성을 보여준다. 따라서 건강기능식품의 기준 및 규격에 제시된 바와 같이 Enterococcus spp.는 항생제 내성 및 독성 유전자가 없는 경우에 한하여 probiotics로 사용이 가능한 고시하고 있어 E. faecalis BMSE-HMP005의 안전성에 대한 추가적인 검증이 필요할 것으로 판단된다.

Keywords

Acknowledgement

본 연구는 2019년도 과학기술정보통신부의 재원으로 한국연구재단 이공분야 기초연구사업의 지원으로 수행되었다(과제번호 NRF-2019R1FA105839).

References

  1. Almeida-Santos AC, Novais C, Peixe L, Freitas AR. Enterococcus spp. as a producer and target of bacteriocins: A double-edged sword in the antimicrobial resistance crisis context. Antibiotics 10: 1215 (2021) https://doi.org/10.3390/antibiotics10101215
  2. Argyri AA, Zoumpopoulou G, Karatzas KAG, Tsakalidou E, Nychas GJE, Panagou EZ, Tassou CC. Selection of potential probiotic lactic acid bacteria from fermented olives by in vitro tests. Food Microbiol. 33: 282-29 (2013) https://doi.org/10.1016/j.fm.2012.10.005
  3. Baccouri O, Boukerb AM, Farhat LB, Zebre A, Zimmermann K, Domann E, Cambronel M, Barreau M, Maillot O, Rince I, Muller C, Marzouki MN, Feuilloley M, Abidi F, Connil N. Probiotic potential and safety evaluation of Enterococcus faecalis OB14 and OB15, isolated from traditional Tunisian testouri cheese and rigouta, using physiological and genomic analysis. Front. Microbiol. 10: 881 (2019) https://doi.org/10.3389/fmicb.2019.00881
  4. Bagci U, Togay SO, Temiz A, Ay M. Probiotic characteristics of bacteriocin-producing Enterococcus faecium strains isolated from human milk and colostrum. Folia Microbiologica. 64: 735-750 (2019) https://doi.org/10.1007/s12223-019-00687-2
  5. Blessing EN, Chukwuemeka IS, David UC, Onuawuchi UG. Antibacterial properties of probiotics bacterial isolated from human breast milk. WNOFNS. 29: 290-297 (2020)
  6. Braiek OB, Smaoui S. Enterococci: Between emerging pathogens and potential probiotics. Biomed. Res. Int. 2019: 1-13 (2019) https://doi.org/10.1155/2019/5938210
  7. Brunel AS, Guery B. Multidrug resistant (or antimicrobial-resistant) pathogens-alternatives to new antibiotics? Swiss Med. Wkly. 147: w14553 (2017)
  8. Byakika S, Mukisa IM, Mugabi R, Muyanja C. Antimicrobial activity of lactic acid bacteria starters against acid tolerant, antibiotic resistant, and potentially virulent E. coli isolated from a fermented sorghum-millet beverage. Int. J. Microbiol. 2019: 2013539 (2019)
  9. Cleveland J, Montville TJ, Nes IF, Chikindas M. Bacteriocins: safe, natural antimicrobials for food preservation. Int. J. Food Microbiol. 71: 1-20 (2001) https://doi.org/10.1016/S0168-1605(01)00560-8
  10. Clinical and Laboratory Standards Institute. Methods for dilution antimicrobial susceptibility tests for bacteria that grow aerobically. 11th ed. Replaces M07-A10 (2018)
  11. Dabek M, McCrae SI, Stevens VJ, Duncan SH, Louis P. Distribution of β-glucosidase and β-glucuronidase activity and of β-glucuronidase gene gus in human colonic bacteria. FEMS Microbiology Ecology. 66: 487-495 (2008) https://doi.org/10.1111/j.1574-6941.2008.00520.x
  12. De Kwaadsteniet M, Todorov SD, Knoetze H, Dicks LMT. Characterization of a 3944 Da bacteriocin, produced by Enterococcus mundtii ST15, with activity against Gram-positive and Gram-negative bacteria. Int. J. Food Microbiol. 105: 433-44 (2005) https://doi.org/10.1016/j.ijfoodmicro.2005.03.021
  13. Ferchichi M, Sebei K, Boukerb AM, Karray-Bouraoui N, Chevalier S, Feuilloley MGJ, Connil N, Zommiti M. Enterococcus spp.: Is it a bad choice for a good use-a conundrum to solve? Microorganisms. 9: 2222 (2021) https://doi.org/10.3390/microorganisms9112222
  14. Hanchi H, Mottawea W, Sebei K, Hammami R. The genus Enterococcus: Between probiotic potential and safety concerns-An update. Front. Microbiol. 9: 1791 (2018) https://doi.org/10.3389/fmicb.2018.01791
  15. Henning C, Gautam D, Muriana P. Identification of multiple bacteriocins in Enterococcus spp. using an Enterococcus-specific bacteriocin PCR array. Microorganisms 3: 1-16 (2015) https://doi.org/10.3390/microorganisms3010001
  16. Huang MS, Cheng CC, Tseng SY, Lin YL, Lo HM, Chen PW. Most commensally bacterial strains in human milk of healthy mothers display multiple antibiotic resistance. Microbiologyopen. 8: e00618 (2019) https://doi.org/10.1002/mbo3.618
  17. Hunt KM, Foster JA, Forney LJ, Schutte UME, Beck DL, Abdo Z, Fox LK, Williams JE, McGuire MK, McGuire MA. Characterization of the diversity and temporal stability of bacterial communities in human milk. PLoS One. 6: e21313 (2011) https://doi.org/10.1371/journal.pone.0021313
  18. Izquierdo E, Bednarczyk A, Schaeffer C, Cai Y, Marchioni E, Dorsselaer AV, Ennahar S. Production of enterocins L50A, L50B, and IT, a new enterocin, by Enterococcus faecium IT62, a strain isolated from Italian ryegrass in Japan. Antimicrob. Agents Chemother. 52: 1917-23 (2008) https://doi.org/10.1128/AAC.01409-07
  19. Jimenez E, Delgado S, Fernandez L, Garcia N, Albujar M, Gomez A, Rodriguez JM. Assessment of the bacterial diversity of human colostrum and screening of Staphylococcal and Enterococcal populations for potential virulence factors. Research in Microbiology. 159: 595-601 (2008) https://doi.org/10.1016/j.resmic.2008.09.001
  20. Jose NM, Bunt CR, Hussain MA. Comparison of microbiological and probiotic characteristics of Lactobacilli isolates from dairy food products and animal rumen contents. Microorganisms. 3: 198-212 (2015) https://doi.org/10.3390/microorganisms3020198
  21. Kumar M, Srivastava S. Antilisterial activity of a broad-spectrum bacteriocin, enterocin LR/6 from Enterococcus faecium LR/6. Appl. Biochem. Biotechnol. 162: 698-706 (2009) https://doi.org/10.1007/s12010-009-8851-1
  22. Line JE, Svetoch EA, Eruslanov BV, Perelygin VV, Mitsevich EV, Mitsevich IP, Levchuk VP, Svetoch OE, Seal BS, Siragusa GR, Stern NJ. lation and purification of enterocin E-760 with broad antimicrobial activity against gram-positive and gram-negative bacteria. Antimicrob. Agents Chemother. 52: 1094-1100 (2008) https://doi.org/10.1128/AAC.01569-06
  23. Liu G, Ren L, Song Z, Wang C, Sun B. Purification and characteristics of bifidocin A, a novel bacteriocin produced by Bifidobacterium animals BB04 from centenarians' intestine. Food Control. 50: 889-895 (2015) https://doi.org/10.1016/j.foodcont.2014.10.049
  24. Martin R, Langa S, Reviriego C, Jimenez E, Marin M L, Olivares, Julio B, Jesus J, Leonides, F, Jordi X, Juan M. The commensal microflora of human milk: new perspectives for food bacteriotherapy and probiotics. Trends Food Sci. Tech. 15.3-4: 121-127 (2004) https://doi.org/10.1016/j.tifs.2003.09.010
  25. Mezaini A, Chihib NE, Bouras AD, Nedjar-arroume N, Hornez JP. Antibacterial activity of some lactic acid bacteria isolated from an algerian dairy product. J. Environ. Public Health. 2009: 678495 (2009) https://doi.org/10.1155/2009/678495
  26. Miller WR, Munita JM, Arias CA. Mechanisms of antibiotic resistance in enterococci. Expert Rev. Anti. Infect. Ther. 12: 1221-1236 (2015) https://doi.org/10.1586/14787210.2014.956092
  27. Ouwehand AC, Salminen S, Isolauri E. Probiotics: an overview of beneficial effects. Antonie van Leeuwenhoek. 82: 279-289 (2002) https://doi.org/10.1023/A:1020620607611
  28. Phumisantiphong U, Siripanichgon K, Reamtong O, Diraphat P. A novel bacteriocin from Enterococcus faecalis 478 exhibits a potent activity against vancomycin-resistant enterococci. PLos One. 12: e0186415 (2017) https://doi.org/10.1371/journal.pone.0186415
  29. Rahmani M, Saffari F, Aboubakri O, Mansouri S. Enterococci from breast-fed infants exert higher antibacterial effects than those from adults: A comparative study. Hum. Microbiome J. 17: 100072 (2020) https://doi.org/10.1016/j.humic.2020.100072
  30. Salminen S, Ouwehand ACO, Isolauri E. Clinical applications of probiotic bacteria. Int. Dairy Journal. 8: 563-572 (1998) https://doi.org/10.1016/S0958-6946(98)00077-6
  31. Vivas R, Teixeira-Barbosa AA, Dolabela SS, Jain S. Multidrug-resistant bacteria and alternative methods to control them: An overview. Microb. Drug Resist. 25: 1-19 (2019) https://doi.org/10.1089/mdr.2018.0104
  32. Wiegand I, Hilpert K, Hancock RE. Agar and broth dilution methods to determine the minimal inhibitory concentration (MIC) of antimicrobial substances. Nature protocols. 3.2: 163-175 (2008) https://doi.org/10.1038/nprot.2007.521
  33. Yasir M, Dutta D, Wilcox MDP. Mode of action of the antimicrobial peptide Mel4 is independent of Staphylococcus aureus cell membrane permeability. PLoS One. 14: e0215703 (2019) https://doi.org/10.1371/journal.pone.0215703
  34. Yoon SH, Ha SM, Kwon S, Lim J, Kim Y, Seo H, Chun J. Introducing EzBioCloud: A taxonomically united database of 16S rRNA and whole genome assemblies. Int. J. Syst. Evol. Microbiol. 67: 1613-1617 (2017) https://doi.org/10.1099/ijsem.0.001755
  35. Zhang F, Jiang M, Wan C, Chen X, Chen X, Tao X, Shah NP, Wei H. Screening probiotic strains for safety: Evaluation of virulence and antimicrobial susceptibility of enterococci from healthy Chinese infants. J. Dairy Sci. 99: 1-9 (2016) https://doi.org/10.3168/jds.2015-9445