Multiple Confirmation and RAPD-genotyping of Enterobacter sakazakii Isolated from Sunsik

선식에서 분리한 Enterobacter sakazakii의 복합동정 및 RAPD를 이용한 genotyping

  • Choi, Jae-Won (Food Safety Research Division, Korea Food Research Institute) ;
  • Kim, Yun-Ji (Food Safety Research Division, Korea Food Research Institute) ;
  • Lee, Jong-Kyung (Food Safety Research Division, Korea Food Research Institute) ;
  • Kim, Young-Ho (Food Safety Research Division, Korea Food Research Institute) ;
  • Kwon, Ki-Sung (Center for Food Evaluation, Korea Food and Drug Adminstration) ;
  • Hwang, In-Gyun (Center for Food Evaluation, Korea Food and Drug Adminstration) ;
  • Oh, Se-Wook (Food Safety Research Division, Korea Food Research Institute)
  • 최재원 (한국식품연구원 안전성연구단) ;
  • 김윤지 (한국식품연구원 안전성연구단) ;
  • 이종경 (한국식품연구원 안전성연구단) ;
  • 김영호 (한국식품연구원 안전성연구단) ;
  • 권기성 (식품의약품안전청 식품평가부) ;
  • 황인균 (식품의약품안전청 식품평가부) ;
  • 오세욱 (한국식품연구원 안전성연구단)
  • Published : 2008.02.28

Abstract

Enterobacter sakazakii is implicated in severe forms of neonatal infections such as meningitis and sepsis. This organism has been isolated from a wide range of foods, including cheese, vegetables, grains, herbs, and spices, but its primary environment is still unknown. Generally, dried infant milk formula has been epidemiologically identified as the source of E. sakazakii. Sunsik (a powdered mixture of roasted grains and other foodstuffs) is widely consumed in Korea as a side dish or energy supplement. Sunsik is consumed without heat treatment; thus, lacking an additional opportunity to inactivate foodborne pathogens. Therefore, its microbiological safety should be guaranteed. In this study, the prevalence of E. sakazakii was monitored in 23 different sunsik component flours, using FDA recommended methods; but E. sakazakii medium (Neogen) and Chromogenic E. sakazakii medium (Oxoid) were used as the selective media. In total, presumptive E. sakazakii strains were isolated from 8 different sunsik powders. Subsequently, an API 20E test was conducted, and 15 strains from 5 different sunsik flours (sea tangle, brown rice, non-glutinous rice, cheonggukjang, dried anchovy) were confirmed as E. sakazakii. Fifteen strains were again confirmed by PCR amplification, using three different primer sets (tDNA sequence, ITS sequence, 16S rRNA sequence), and compared to ATCC strains (12868, 29004, 29544, 51329). They were once again confirmed by their enzyme production profiles using an API ZYM kit. Finally, RAPD (random amplified polymorphic DNA)-genotyping was carried out as a monitoring tool to determine the contamination route of E. sakazakii during processing.

Keywords

Enterobacter sakazakii;isolation;confirmation;PCR;genotyping

References

  1. Emery CL, Weymouth LA. Detection and clinical significance of extended-spectrum beta-lactamases in a tertiary-care medical center. J. Clin. Microbiol. 35: 2061-2067 (1997)
  2. Pribyl C, Salzer R, Beskin J, Haddad RJ, Pollock B, Beville R, Holmes B, Mogabgab WJ. Aztreonam in the treatment of serious orthopedic infections. Am. J. Med. 78: 51-56 (1985) https://doi.org/10.1016/0002-9343(85)90205-0
  3. Lai KK. Enterobacter sakazakii infections among neonates, infants, children, and adults. Case reports and a review of the literature. Medicine 80: 113-122 (2001) https://doi.org/10.1097/00005792-200103000-00004
  4. Bar-Oz B, Preminger A, Peleg O, Block C, Arad I. Enterobacter sakazakii infection in the newborn. Acta Paediatr. 90: 356-358 (2001) https://doi.org/10.1111/j.1651-2227.2001.tb00319.x
  5. Biering G, Karlsson S, Clark NC, Jonsdottir KE, Ludvigsson P, Steingrimsson O. Three cases of neonatal meningitis caused by Enterobacter sakazakii in powdered milk. J. Clin. Microbiol. 27: 2054-2056 (1989)
  6. Muytjens HL, Zanen HC, Sonderkamp HJ, Kollee LA, Wachsmuth IK, Farmer JJ. Analysis of eight cases of neonatal meningitis and sepsis due to Enterobacter sakazakii. J. Clin. Microbiol. 18: 115-120 (1983)
  7. The Food Safety Authority of Ireland. Pregestimil infant formula recall. http://www.fsai.ie/alerts/archive/fa20050120.asp. Accessed Jan. 20, 2005
  8. Gurtler JB, Kornacki JL, Beuchat LR. Enterobacter sakazakii: A coliform of increased concern to infant health. Int. J. Food Microbiol. 104: 1-34 (2005) https://doi.org/10.1016/j.ijfoodmicro.2005.02.013
  9. FDA. http://www.cfsan.fda.gov/~ebam/bam-a2.html#excl. Accessed Dec. 29, 2006
  10. Lehner A, Tasara T, Stephan R. 16S rRNA genes based analysis of Enterobacter sakazakii strains from different sources and development of a PCR assay for identification. BMC Microbiol. 4: 43-50 (2004) https://doi.org/10.1186/1471-2180-4-43
  11. Chung SS, Han YS. Consumer's recognition, nutrient composition, and safety evaluation of commercial sunsik and saengsik. Korean J. Food Culture 18: 235-243 (2003)
  12. Dennison SK, Morris J. Multiresistant Enterobacter sakazakii wound infection in an adult. Infect. Med. 19: 533-535 (2002)
  13. Noriega FR, Kotloff KL, Martin MA, Schwalbe RS. Nosocomial bacteremia caused by Enterobacter Sakazakii and Leuconostoc mesenteroides resulting from extrinsic contamination of infant formula. Pediatr. Infect. Dis. J. 9: 447-449 (1990) https://doi.org/10.1097/00006454-199006000-00018
  14. Jimenez E, Gimenez C. Septic shock due to Enterobacter sakazakii. Clin. Microbiol. Newslett. 4: 30 (1982) https://doi.org/10.1016/S0196-4399(82)80129-3
  15. Oh SW, Kang DH. Fluorogenic selective and differential medium for isolation of Enterobacter sakazakii. Appl. Environ. Microb. 70: 5692-5694 (2004) https://doi.org/10.1128/AEM.70.9.5692-5694.2004
  16. Van Acker J, De Smet F, Muyldermans G, Bougatef A, Naessens A, Lauwers S. Outbreak of necrotizing enterocolitis associated with Enterobacter sakazakii in powdered milk formula. J. Clin. Microbiol. 39: 293-297 (2001) https://doi.org/10.1128/JCM.39.1.293-297.2001
  17. Enterobacter sakazakii and other microorganisms in powdered infant formula: meeting report, MRA Series 6. Microbiological Risk Assessment Series, No. 6 ISBN: 92 4 156262 5 (WHO) http://www.who.int/foodsafety/publications/micro/es.pdf. Accessed Dec. 30, 2006
  18. Kandhai MC, Reij MW, Gorris LG, Guillaume-Gentil O, van Schothorst M. Occurrence of Enterobacter sakazakii in food production environments and households. Lancet 363: 39-40 (2004) https://doi.org/10.1016/S0140-6736(03)15169-0
  19. Farmer JJ, Asbury MA, Hickman FW, Brenner DJ. The Enterobacteriaceae study group (USA). Enterobacter sakazakii: A new species of 'Enterobacteriaceae' isolated from clinical specimens. Int. J. Syst. Bacteriol. 30: 569-584 (1980) https://doi.org/10.1099/00207713-30-3-569
  20. Wyeth Nutrition. Wyeth Nutrition initiates voluntary recall of selected store brand infant formula from U.S. marketplace. http:// www.wyeth.com/news?nav=display&navTo=/wyeth_html/home/ news/pressreleases/2002/1146452500787.html. Accessed Nov. 1, 2002
  21. FDA. Food and Drug Administration, Isolation and enumeration of Enterobacter sakazakii from dehydrated powdered infant formula. http://www.cfsan.fda. gov/~comm/mmesakaz.html. Accessed Sep. 19, 2003
  22. Muytjens HL, Kollee LA. Enterobacter sakazakii meningitis in neonates: Causative role of formula? Pediatr. Infect. Dis. J. 9: 372-373 (1990) https://doi.org/10.1097/00006454-199005000-00016
  23. Keyser M, Withuhn RC, Ronquest LC, Britz T. Treatment of winery effluent with upflow anaerobic sludge blanket (UASB)- granular sludges enriched with Enterobacter sakazakii. Biotechnol. Lett. 25: 1893-1898 (2003) https://doi.org/10.1023/B:BILE.0000003978.72266.96
  24. Simmons BP, Gelfand MS, Haas M, Metts L, Ferguson F. Enterobacter sakazakii infections in neonates associated with intrinsic contamination of a powdered infant formula. Infect. Cont. Hosp. Ep. 10: 398-401 (1989) https://doi.org/10.1086/646060
  25. Iverson C, Forsythe SJ. Risk profiles of Enterobacter sakazakii, an emergent pathogen associated with infant milk formula. Trends Food Sci. Tech. 14: 443-454 (2003) https://doi.org/10.1016/S0924-2244(03)00155-9
  26. Hawkins RE, Lissner CR, Sanford JP. Enterobacter sakazakii bacteremia in an adult. South. Med. J. 84: 793-795 (1991) https://doi.org/10.1097/00007611-199106000-00033
  27. Clementino MM, Filippis ID, Nascimento CR, Branquinho R, Rocha CL, Martins OB. PCR analyses of tRNA intergenic spacer, 16S-23S internal transcribed spacer, and randomly amplified polymorphic DNA reveal inter- and intraspecific relationship of Enterobacter cloacae strains. J. Clin. Microbiol. 39: 3865-3870 (2001) https://doi.org/10.1128/JCM.39.11.3865-3870.2001
  28. Clark NC, Hill BC, O'Hara CM, Steingrimsson O, Cooksey RC. Epidemiologic typing of Enterobacter sakazakii in two neonatal nosocomial outbreaks. Diagn. Micr. Infec. Dis. 13: 467-72 (1990) https://doi.org/10.1016/0732-8893(90)90078-A
  29. Nazarowec-White M, Farber JM. Incidence, survival, and growth of Enterobacter sakazakii in infant formula. J. Food Protect. 60: 226-230 (1997) https://doi.org/10.4315/0362-028X-60.3.226
  30. Iverson C, Druggan P, Forsythe S. A selective differential medium for Enterobacter sakazakii, a preliminary study. Int. J. Food Microbiol. 96: 133-139 (2004) https://doi.org/10.1016/j.ijfoodmicro.2004.01.024
  31. Muytjens HL, Roelofs-Willemse H, Jaspar GH. Quality of powdered substitutes for breast milk with regard to members of the family Enterobacteriaceae. J. Clin. Microbiol. 26: 743-746 (1988)