DOI QR코드

DOI QR Code

Application of MALDI-TOF mass spectrometry-based identification of foodborne pathogen tests to the Korea Food Standard Codex

MALDI-TOF 질량분석기를 이용한 식품중독균 확인시험 적용

  • 하미영 (농협중앙회 식품연구원) ;
  • 손은정 (농협중앙회 식품연구원) ;
  • 최은정 (농협중앙회 식품연구원)
  • Received : 2016.05.19
  • Accepted : 2016.07.05
  • Published : 2016.10.31

Abstract

Rapid and reliable identification of microorganisms is important to maintain food quality and to control safety. MALDI-TOF MS-based identification methods are relatively fast and simple compared to other conventional methods including gram staining and biochemical characterization. A colony on subcultured media can be directly prepared on the analysis plate without further complex treatments. In this study, we confirmed the applicability of MALDI-TOF MS-based identification of foodborne pathogens such as Salmonella Enteritidis/Typhimurium, Staphylococcus aureus, Vibrio parahaemolyticus, Clostridium perfringens, Listeria monocytogenes, Yersinia enterocolitica, Bacillus cereus, Campylobacter jejuni, Campylobacter coli, and Cronobacter sakazakii on the Korea Food Standard Codex. MALDI-TOF MS data of the pathogenic reference strains were incorporated into a commercial MicroID (ASTA Inc.) database. Other pathogenic reference strains and seven isolates from various food samples were correctly identified to the species level by using the MicroID database. In conclusion, MALDI-TOF MS is comparable with commercial biochemical identification.

최근 건강과 위생에 대한 소비자의 의식 향상으로 인해 농수축산 분야를 비롯한 식품의 가공 유통 분야에서도 식품의 안전성 확보를 위한 시험 검사가 실시되고 있고, HACCP과 같은 식품안전관리 프로그램의 조기 도입을 유도하고 있어 식품중독균에 대한 검사량이 증가하고 있다. 이에 따른 신속, 정확하고 대량의 시료를 처리할 수 있는 식품중독균 검사의 필요성이 증가되고 있다. 국내 식품 미생물의 확인시험방법은 전통적인 미생물 동정법인 그램 염색 등과 같은 형태학적 특성과 생화학적 분석에 의해서 주로 확인되는데, 확인 과정이 복잡하고 장시간이 소요된다. 이를 극복하기 위한 새로운 미생물 동정법인 MALDI-TOF 질량분석기반 미생물 동정법을 식품의 식품중독균 검사에 적용하기 위해 식품공전에서 주로 검사하는 식품중독균 10종에 대한 질량 패턴 데이터를 국내 질량분석 데이터베이스인 MicroID에 적용하였다. 표준 균주와 식품중독균이 검출된 시료에서 분리한 균으로 비교했을 때 질량분석기반 미생물 동정은 현재 사용되고 있는 생화학적 분석결과와 일치한 결과를 보여주었다. 또한, 식품중독균을 포함한 국내 미생물 균주를 이용해서 구축한 데이터베이스, MicroID는 기존의 상용화된 해외 MALDI-TOF 질량분석 데이터베이스 Biotyper와 동등 이상의 정확도를 나타내었다. 국내 식품관련 미생물에 대한 질량스펙트럼을 추가하여 데이터베이스를 지속적으로 확장시키면 신속 정확한 미생물 동정법으로 자리매김할 수 있을 것이다.

Keywords

References

  1. Kim HW, Ham JS, Seol KH, Han SH, Park BY, Oh MH. MALDI-TOF MS system for the identification of microorganisms in milk and dairy products. J. Milk Sci. Biotechnol. 30: 131-137 (2012)
  2. Wieser A, Schneider L, Jung J, Schubert S. MALDI-TOF MS in microbiological diagnostics-identification of microorganisms and beyond (mini review). Appl. Microbiol. Biotechnol. 93: 965-974 (2012) https://doi.org/10.1007/s00253-011-3783-4
  3. Murray PR. What is new in clinical microbiology-microbial identification by MALDI-TOF Mass Spectrometry. J. Mol. Diagn. 14: 419-423 (2012) https://doi.org/10.1016/j.jmoldx.2012.03.007
  4. Dingle TC, Butler Wu SM. MALDI-TOF mass spectrometry for microorganism identification. Clin. Lab. Med. 33: 589-609 (2013) https://doi.org/10.1016/j.cll.2013.03.001
  5. Bourassa L, Butler-Wu SM. MALDI-TOF Mass Spectrometry for Microorganism Identification. Vol. 42. pp. 37-85. In: Methods in Microbiology. Norris JR, Ribbons DW (eds). Academic Press, Cambridge, MA, USA (2015)
  6. Anhalt JP, Fenselau C. Identification of bacteria using mass spectrometry. Anal. Chem. 47: 219-225 (1975) https://doi.org/10.1021/ac60352a007
  7. Claydon MA, Davey SN, Edwards-Jones V, Gordon DB. The rapid identification of intact microorganisms using mass spectrometry. Nat. Biotechnol. 14: 1584-1586 (1996) https://doi.org/10.1038/nbt1196-1584
  8. Holland RD, Wilkes JG, Ralli F, Sutherland JB, Persons CC, Voorhees KJ, Lay Jr. JO. Rapid identification of intact whole bacteria based on spectral patterns using matrix-assisted laser desorption/ ionization with time-of-flight mass spectrometry. Rapid Commun. Mass Sp. 10: 1227-1232 (1996) https://doi.org/10.1002/(SICI)1097-0231(19960731)10:10<1227::AID-RCM659>3.0.CO;2-6
  9. Krishnamurthy T, Ross PL, Rajamani U. Detection of pathogenic and non-pathogenic bacteria by matrix-assisted laser desorption ionization time-of-flight mass spectrometry. Rapid Commun. Mass Sp. 10: 883-888 (1996) https://doi.org/10.1002/(SICI)1097-0231(19960610)10:8<883::AID-RCM594>3.0.CO;2-V
  10. Weisburg WG, Bams SM, Pelletier DA, Lane DJ. 16S Ribosomal DNA amplification for phylogenetic study. J. Bacteriol. 173: 697- 703 (1991) https://doi.org/10.1128/jb.173.2.697-703.1991
  11. Hansen BM, Leser TD, Hendriksen NB. Polymerase chain reaction assay for the detection of Bacillus cereus group cells. FEMS Microbiol. Lett. 202: 209-213 (2001) https://doi.org/10.1111/j.1574-6968.2001.tb10805.x
  12. Chen ML, Tsen HY. Discrimination of Bacillus cereus and Bacillus thuringiensis with 16S rRNA and gyrB gene based PCR primers and sequencing of their annealing sites. J. Appl. Microbiol. 92: 912-919 (2002) https://doi.org/10.1046/j.1365-2672.2002.01606.x
  13. Bakke I, Schryver PD, Boon N, Vadstein O. PCR-based community structure studies of Bacteria associated with eukaryotic organisms: A simple PCR strategy to avoid co-amplification of eukaryotic DNA. J. Microbiol. Meth. 84: 349-351 (2011) https://doi.org/10.1016/j.mimet.2010.12.015
  14. Smole SC, King LA, Leopold PE, Arbeit RD. Sample preparation of gram-positive bacteria for identification by matrix assisted laser desorption/ionization time-of-flight. J. Microbiol. Meth. 48: 107-115 (2002) https://doi.org/10.1016/S0167-7012(01)00315-3
  15. Bizzini A, Durussel C, Bille J, Greub G, Prodhom G. Performance of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry for identification of bacterial strains routinely isolated in a clinical microbiology laboratory. J. Clin. Microbiol. 48: 1549-1554 (2010) https://doi.org/10.1128/JCM.01794-09
  16. Alatoom AA, Cunningham SA, Ihde SM, Manrekar J, Patel R. Comparison of ditrect colony method versus extraction method for identfication gram-positive cocci by use of bruker biotyper matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. J. Clin. Microbiol. 49: 2868-2873 (2011) https://doi.org/10.1128/JCM.00506-11
  17. Haigh J, Degun A, Eydmann M, Millar M, Wilks M. Improved performance of bacterium and yeast identification by commercial in the clinical microbiology laboratory. J. Clin. Microbiol. 49: 3441 (2011) https://doi.org/10.1128/JCM.00576-11
  18. Evason DJ, Claydon MA, Gordon DB. Effects of ion mode and matrix additives in the identification of bacteria by intact cell mass spectrometry. Rapid Commun. Mass Sp. 14: 669-672 (2000) https://doi.org/10.1002/(SICI)1097-0231(20000430)14:8<669::AID-RCM932>3.0.CO;2-7
  19. Fenselau C, Demire PA. Characterization of intact microorganisms by MALDI mass spectrometry. Mass Spectrom. Rev. 20: 157-171 (2001) https://doi.org/10.1002/mas.10004
  20. Ford BA, Burnham CD. Optimization of routine identification of clinically relevant gram-negative bacteria by use of matrixassisted laser desorption ionization time-of-flight mass spectrometry and the Bruker Biotyper. J. Clin. Microbiol. 51: 1412-1420 (2013) https://doi.org/10.1128/JCM.01803-12
  21. TeKippe EM, Shuey S, Winkler DW, Butler MA, Burnham CD. Optimizing identification of clinically relevant gram-positive organisms by use of the Bruker Biotyper matrix-assisted laser desorption ionization time-of-flight mass spectrometry system, J. Clin. Microbiol. 51: 1421-1427 (2013) https://doi.org/10.1128/JCM.02680-12
  22. Seng P, Drancourt M, Gouriet F. Ongoing revolution in bacteriology: Routine identification of bacteria by matrix-assisted laser desorption ionization time-of-flight mass spectrometry. Clin. Infect. Dis. 49: 543-551 (2009) https://doi.org/10.1086/600885
  23. Mellmann A, Bimet F, Bizet C. High interlaboratory reproducibility of matrix-assisted laser desorption ionization-time of flight mass spectrometry-based species identification of nonfermenting bacteria. J. Clin. Microbiol. 47: 3732-3734 (2009) https://doi.org/10.1128/JCM.00921-09
  24. Croxatto A, Prod'hom G, Greub G. Application of MALDI-TOF mass spectrometry in clinical diagnostic microbiology. FEMS Microbiol. Rev. 36: 380-407 (2012) https://doi.org/10.1111/j.1574-6976.2011.00298.x
  25. Drobniewski FA. Bacillus cereus and related species. Clin. Microbiol. Rev. 6: 324-338 (1993) https://doi.org/10.1128/CMR.6.4.324
  26. Bremer H, Dennis PP. Modulation of chemical composition and other parameters of the cell by growth rate. pp. 167-182. In: Escherichia coli and Salmonella: Cellular and Molecular Biology. Neidhardt FC (ed). ASM Press, Washington, DC, USA (1996)
  27. Ryzhov V, Fenselau C. Characterization of the protein subset desorbed by MALDI from whole bacterial cells. Anal. Chem. 73:746-750 (2001) https://doi.org/10.1021/ac0008791
  28. Ash C, Farrow John AE, Dorsch M, Stackebrandt E, Collins MD. Comparative analysis of Bacillus anthracis, Bacillus cereus, and related species on the basis of reverse transcriptase gene sequencing of 16S rRNA. Int. J. Syst. Bacteriol. 41: 343-346 (1991) https://doi.org/10.1099/00207713-41-3-343
  29. Bessede E, Solecki O, Sifre E, Labadi L, Megraud F. Identification of Campylobacter species and related organisms by matrix assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry. Clin. Microbiol. Infect. 17: 1735-1739 (2011) https://doi.org/10.1111/j.1469-0691.2011.03468.x