Journal of Food Hygiene and Safety (한국식품위생안전성학회지)

The Korean Society of Food Hygiene and Safety (한국식품위생안전성학회)
권호 표지
DOI QR코드

DOI QR Code

Improvement of the Detection Technique of Listeria monocytogenes through Modification of the Enrichment Medium and DNA Extraction Buffer

증균배지 및 DNA 추출법 개량을 통한 Listeria monocytogenes의 검출기법 개선 연구

  • Lee, Jeeyeon (Department of Food & Nutrition, Dong-eui University) ;
  • Seo, Yeongeun (Department of Food and Nutrition, Sookmyung Women's University) ;
  • Yoon, Yohan (Department of Food and Nutrition, Sookmyung Women's University)
  • 이지연 (동의대학교 식품영양학과) ;
  • 서영은 (숙명여자대학교 식품영양학과) ;
  • 윤요한 (숙명여자대학교 식품영양학과)
  • Received : 2020.08.11
  • Accepted : 2020.08.18
  • Published : 2020.08.30
Download PDF
⟨ Previous Next ⟩

Abstract

In this study we developed an enrichment medium and lysis buffers to detect Listeria monocytogenes in meat and processed meat products under various lysis conditions. The enrichment efficiency of L. monocytogenes medium listed in the Food Standards was compared, and thus, Listeria Enrichment Broth (LEB) was modified by adding supplements such as carbon source and minerals. The lysis buffers were developed to extract L. monocytogenes DNA quickly and efficiently under various lysis conditions. L. monocytogenes was most rapidly grown in LEB containing 0.1% pyruvate and 0.1% ferric citrate. A lysis buffer mixed with 0.5% or 1% N-lauroylasrcosine sodium salt, 0.5 N NaOH and 0.5 M EDTA for 30 min at room temperature was found to be the best in terms of DNA purity and yield. These results indicate that developed enrichment medium and lysis buffer can be used to detect L. monocytogenes in meat and processed meat products rapidly and efficiently.

기존의 L. monocytogenes 증균배지를 개량함으로써 증균배지를 개발하여 L. monocytogenes의 증균 효율을 높이며, DNA 추출에 사용되는 용해버퍼 및 용해조건을 개발함으로써 식육 및 식육가공품에서 L. monocytogenes를 효율적이고 신속하게 검출하고자 하였다. 식품공전에 등재되어 있는 L. monocytogenes 증균배지의 증균 효율을 비교하였으며, Listeria Enrichment Broth (LEB)가 가장 우수한 증균 효율을 보였다. LEB에 탄소원, 질소원, 미네랄 등 다양한 성분을 첨가하여 증균배지를 개발하였으며, 그 결과 LEB에 0.1% pyruvate, 0.1% ferric citrate를 첨가한 개발 증균배지에서 L. monocytogenes가 가장 빠르게 증균되었다. L. monocytogenes의 DNA를 신속하고 효율적으로 추출하기 위해 용해버퍼와 용해조건을 개발하였으며, 그 결과 0.5% 또는 1% N-lauroylsarcosine sodium salt에 0.5 N NaOH와 0.5 M EDTA가 혼합된 용해버퍼를 이용하여 실온에서 30분 간 L. monocytogenes 세포를 용해시키는 것이 DNA의 순도와 수율, 신규성과 경제적 측면에서 가장 우수한 것으로 확인되었다. 결론적으로 본 연구에서 개발된 증균배지 및 DNA 추출법을 활용한다면 식육 및 식육 가공품에서 L. monocytogenes를 보다 신속하게 검출할 수 있을 것이다.

Keywords

References

  1. Chen, Y., Chen, M., Wang, J., Wu, Q., Cheng, J., Zhang, J., Sun, Q., Xue, L., Zeng, H., Lei, T., Pang, R., Ye, Q., Wu, S., Zhang, S., Wu, H., Li, W., Kou, X., Heterogeneity, characteristics, and public health implications of Listeria monocytogenes in ready-to-eat foods and pasteurized milk in China. Front. Microbiol., 11, 642 (2020). https://doi.org/10.3389/fmicb.2020.00642
  2. Demaitre, N., Van Damme, I., De Zutter, L., Geeraerd, A.H., Rasschaert, G., De Reu, K., Occurrence, distribution and diversity of Listeria monocytogenes contamination on beef and pig carcasses after slaughter. Meat Sci., 108177 (2020).
  3. Lee, J., Gwak, E., Lee, H., Ha, J., Lee, S., Kim, S., Oh, M.H., Park, B.Y., Choi, K.H., Yoon, Y., Effects of low $NaNO_2$ and NaCl concentrations on Listeria monocytogenes growth in emulsion-type sausage. Asian-Australasian J. Anim. Sci., 30, 432-438 (2017). https://doi.org/10.5713/ajas.16.0391
  4. Liu, Y., Sun, W., Sun, T., Gorris, L.G., Wang, X., Liu, B., Dong, Q., The prevalence of Listeria monocytogenes in meat products in China: A systematic literature review and novel meta-analysis approach. Int. J. Food Microbiol., 312, 108358 (2020). https://doi.org/10.1016/j.ijfoodmicro.2019.108358
  5. Rietberg, K., Lloyd, J., Melius, B., Wyman, P., Treadwell, R., Olson, G., Kang, M.G., Duchin, J.S., Outbreak of Listeria monocytogenes infections linked to a pasteurized ice cream product served to hospitalized patients. Epidemiol. Infect., 144, 2728-2731 (2016). https://doi.org/10.1017/S0950268815003039
  6. Karyotis, D., Skandamis, P.N., Juneja, V.K., Thermal inactivation of Listeria monocytogenes and Salmonella spp. in sous-vide processed marinated chicken breast. Food Res. Int., 100, 894-898 (2017). https://doi.org/10.1016/j.foodres.2017.07.078
  7. Tangwatcharin, P., Sorapukdee, S., Kongsrirat, K., Sousvided restructured goat steaks: process optimized by thermal inactivation of Listeria monocytogenes and their quality characteristics. Food Sci. Anim. Resour., 39, 863-876 (2019). https://doi.org/10.5851/kosfa.2019.e64
  8. Thangavel, G., Subramaniyam, T., Antimicrobial efficacy of Leuconostoc spp. isolated from Indian meat against Escherichia coli and Listeria monocytogenes in Spinach leaves. Food Sci. Anim. Resour., 39, 677-685 (2019). https://doi.org/10.5851/kosfa.2019.e60
  9. Ministry of Food and Drug Safety, (2020, July 14). Food standards. Retrieved from http://www.foodsafetykorea.go.kr/foodcode/index.jsp
  10. Lee, Y., Yoon, Y., Seo, Y., Kim, S., Ha, J., Lee, J., Choi, Y., Oh, H., Kim, Y., Kang, J., Park, E., Kim, W.I., Lee, S., Combined enrichment and quantitative polymerase chain reaction to improve sensitivity and reduce time of detection of Listeria monocytogenes in mushrooms. Foodborne Pathog. Dis., 17, 276-283 (2020). https://doi.org/10.1089/fpd.2019.2688
  11. Delibato, E., Gattuso, A., Minucci, A., Auricchio, B., De Medici, D., Toti, L., Castangnola, M., Cpoluongo, E., Gianfranceschi, M.V., PCR experion automated electrophoresis system to detect Listeria monocytogenes in foods. J. Sep. Sci., 32, 3817-3821 (2009). https://doi.org/10.1002/jssc.200900166
  12. Furrer, B., Candrian, U., Hoefelein, C., Luethy, J., Detection and identification of Listeria monocytogenes in cooked sausage products and in milk by in vitro amplification of haemolysin gene fragments. J. Appl. Bacteriol., 70, 372-379 (1991). https://doi.org/10.1111/j.1365-2672.1991.tb02951.x
  13. Silk, T.M., Roth, T.M., Donnelly, C.W., Comparison of growth kinetics for healthy and heat-injured Listeria monocytogenes in eight enrichment broths. J. Food Prot., 65, 1333-1337 (2002). https://doi.org/10.4315/0362-028X-65.8.1333
  14. McLaughlin, H.P., Hill, C., Gahan, C.G., The impact of iron on Listeria monocytogenes; inside and outside the host. Curr. Opin. Biotechnol., 22, 194-199 (2011). https://doi.org/10.1016/j.copbio.2010.10.005
  15. Clark, M.S., 1997. Plant Molecular Biology - A Laboratory Manual. Springer, Berlin/ Heidelberg, Germany, pp.305-328.
  16. Popa, O.P., Murariu, D.U.M.I.T.R.U., Popa, L.O., Comparison of four DNA extraction methods from invasive freshwater bivalve species (Mollusca: Bivalvia) in Romanian fauna. Trav. Mus. Natl. Hist. Nat. Grigore Antipa, 6, 527-536 (2007).
  17. Poms, R., Glössl, J., Foissy, H., Increased sensitivity for detection of specific target DNA in milk by concentration in milk fat. Eur. Food Res. Technol., 213, 361-365 (2001). https://doi.org/10.1007/s002170100383
  18. Thermo Fisher Scientific, (2020, August 11). PCR setup-Six Critical Components to Consider. Retrieved from https://www.thermofisher.com/kr/ko/home/life-science/cloning/cloninglearning-center/invitrogen-school-of-molecular-biology/pcr-education/pcr-reagents-enzymes/pcr-component-considerations.html