Journal of Food Hygiene and Safety (한국식품위생안전성학회지)

The Korean Society of Food Hygiene and Safety (한국식품위생안전성학회)
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Development of a Rapid Enrichment Broth for Vibrio parahaemolyticus Using a Predictive Model of Microbial Growth with Response Surface Analysis

미생물 생장 예측모델과 반응표면분석법을 이용한 Vibrio parahaemolyticus의 신속 증균배지 개발

  • Yeon-Hee Seo (Department of Food and Nutrition, Kookmin University) ;
  • So-Young Lee (Department of Food and Nutrition, Kookmin University) ;
  • Unji Kim (Department of Food and Nutrition, Kookmin University) ;
  • Se-Wook Oh (Department of Food and Nutrition, Kookmin University)
  • Received : 2023.09.08
  • Accepted : 2023.10.29
  • Published : 2023.12.30
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Abstract

In this study, we developed Rapid Enrichment Broth for Vibrio parahaemolyticus (REB-V), a broth capable enriching V. parahaemolyticus from 100 CFU/mL to 106 CFU/mL within 6 hours, which greatly facilitates the rapid detection of V. parahaemolyticus. Using a modified Gompertz model and response surface methodology, we optimized supplement sources to rapidly enrich V. parahaemolyticus. The addition of 0.003 g/10 mL of D-(+)-mannose, 0.002 g/10 mL of L-valine, and 0.002 g/10 mL of magnesium sulfate to 2% (w/v) NaCl BPW was the most effective combination of V. parahaemolyticus enrichment. Optimal V. parahaemolyticus culture conditions using REB-V were at pH 7.84 and 37℃. To confirm REB-V culture efficiency compared to 2% (w/v) NaCl BPW, we assessed the amount of enrichment achieved in 7 hours in each medium and extracted DNA samples from each culture every hour. Real-time PCR was performed using the extracted DNA to verify the applicability of this REB-V culture method to molecular diagnosis. V. parahaemolyticus was enriched to 5.452±0.151 Log CFU/mL in 2% (w/v) NaCl BPW in 7 hours, while in REB-V, it reached 7.831±0.323 Log CFU/mL. This confirmed that REB-V enriched V. parahaemolyticus to more than 106 CFU/mL within 6 hours. The enrichment rate of REB-V was faster than that of 2% (w/v) NaCl BPW, and the amount of enrichment within the same time was greater than that of 2% (w/v) NaCl BPW, indicating that REB-V exhibits excellent enrichment efficiency.

본 논문은 적은 농도로 존재하는 Vibrio parahaemolyticus를 6시간이내에 106 CFU/mL까지 신속하게 배양가능한 배지인 Rapid Enrichment Broth for V. parahaemolyticus(REB-V)를 개발하여 V. parahaemolyticus를 신속하게 검출할 수 있도록 하였다. Modified Gompertz model과 RSM를 활용하여 V. parahaemolyticus를 신속하게 증균할 수 있도록 첨가 성분을 최적화하였다. 그 결과 2%(w/v) NaCl BPW에 D-(+)-mannose 0.3 g/L, L-valine 0.2 g/L, magnesium sulfate 0.2 g/L를 첨가하였을 때 V. parahaemolyticus의 증균량이 최대였다. REB-V의 배양조건을 pH 7.84와 37℃으로 최적화하였으며, 2%(w/v) NaCl BPW와 비교하여 REB-V의 증균 효율을 확인하기 위해 7시간 증균을 통해 증균량을 평가하였다. 또한 매시간마다 배양액에서 DNA를 추출하였고, 추출한 DNA로 real-time PCR을 수행하여 REB-V의 분자진단법 적용가능성을 확인하였다. 그 결과 7시간 동안 2%(w/v) NaCl BPW에서 V. parahaemolyticus는 5.452±0.151 Log CFU/mL까지 증균되었고, REB-V에서는 7.831±0.323 Log CFU/mL의 V. parahaemolyticus가 증균되었다. 최종적으로 REB-V에서 6시간 이내에 106 CFU/mL 이상으로 증균된 것을 확인하였고, REB-V의 증균속도가 2%(w/v) NaCl BPW보다 빠르고 같은 시간 내의 증균량이 2%(w/v) NaCl BPW보다 많았음을 통해 REB-V의 증균효율이 우수한 것으로 확인되었다.

Keywords

Acknowledgement

본 연구는 농림축산식품부 재원으로 농림식품기술기획평가원의 "식중독균의 신속배양 배지 및 신속 배양기의 개발"의 연구개발비 지원에 의해 수행되었음(821056032SB010).

References

  1. Bisha, B., Simonson, J., Janes, M., Bauman, K., Goodridge, L. D., A review of the current status of cultural and rapid detection of Vibrio parahaemolyticus. Int. J. Food Sci. Technol., 47, 885-899 (2012). https://doi.org/10.1111/j.1365-2621.2012.02950.x
  2. Igbinosa, E.O., Okoh, A.I., Emerging Vibrio species: an unending threat to public health in developing countries. Res. Microbiol., 159, 495-506 (2008). https://doi.org/10.1016/j.resmic.2008.07.001
  3. Ward, L.N., Bej, A.K., Detection of Vibrio parahaemolyticus in shellfish by use of multiplexed real-time PCR with TaqMan fluorescent probes. Appl. Environ. Microb., 72, 2031-2042 (2006). https://doi.org/10.1128/AEM.72.3.2031-2042.2006
  4. Ren, Y., Cao, L., Zhang, X., Jiao, R., Ou, D., Wang, Y., Zhang, D., Ling, N., Ye, Y., A novel fluorescence resonance energy transfer (FRET)-based paper sensor with smartphone for quantitative detection of Vibrio parahaemolyticus. Food Control., 145, 109412 (2023).
  5. Beshiru, A., Igbinosa, E.O., Surveillance of Vibrio parahaemolyticus pathogens recovered from ready-to-eat foods. Sci. Rep-UK., 13, 4186 (2023).
  6. Ministry of Food and Drug Safety (MFDS), (2023, August 7). Statistic system. Retrieved from http://www.foodsafetykorea.go.kr/portal/healthyfoodlife/foodPoisoningStat.do?menu_no=519&menu_grp=MENU_GRP02.
  7. Seung, H.J., Kim, L.L., Park, J.E., Ryu, S.H., Jin, Y.H., Kim, O.H., Yun, E.S., Virulence genes and antimicrobial resistance of Vibrio parahaemolyticus isolated from commercial marine products and aquarium waters. Korean J. Food Sci. Technol., 55, 159-164 (2023). https://doi.org/10.9721/KJFST.2023.55.2.159
  8. Ministry of Food and Drug Safety (MFDS), (2023, August 16), Korean food standards codex. Retrieved from https://www.foodsafetykorea.go.kr/foodcode/01_03.jsp?idx=382
  9. Lee, J., Seo, Y., Yoon, Y., Improvement of the detection technique of sisteria monocytogenes through modification of the enrichment medium and DNA extraction buffer J. Food Hyg. Saf., 35, 334-340 (2020). https://doi.org/10.13103/JFHS.2020.35.4.334
  10. Whitaker, W.B., Richards, G.P., Boyd, E.F., Loss of sigma factor RpoN increases intestinal colonization of Vibrio parahaemolyticus in an adult mouse model. Infect. Immun., 82, 544-556 (2014). https://doi.org/10.1128/IAI.01210-13
  11. Song, Y.J., Cho, K.J., Son, E.I., Jo, D.M., Kim, Y.M., Park, S.K., Detection of Salmonella spp. in seafood via desalinized DNA extraction method and preculture. J. Food Hyg. Saf., 38, 123-130 (2023). https://doi.org/10.13103/JFHS.2023.38.3.123
  12. Yamazaki, W., Ishibashi, M., Kawahara, R., Inoue, K., Development of a loop-mediated isothermal amplification assay for sensitive and rapid detection of Vibrio parahaemolyticus. BMC Microbiol., 8, 163 (2008).
  13. Otomo, Y., Hossain, F., Rabbi, F., Yakuwa, Y., Ahsan, C.R., Pre-enrichment of estuarine and fresh water environmental samples with sodium chloride yields in better recovery of Vibrio parahaemolyticus. Adv. Microb., 3, 21-25 (2013). https://doi.org/10.4236/aim.2013.31003
  14. Eom, T.K., Study on substrate removal and mechanism of Ruminococcus flavefaciens FD-1 under carbon- and nitrogen-limiting conditions. J. Korean Soc. Environ. Eng., 17, 295-302 (1995).
  15. Yang, Z.Q., Jiao, X.A., Li, P., Pan, Z.M., Huang, J.L., Gu, R.X., Fang, W.M., Chao, G.X., Predictive model of Vibrio parahaemolyticus growth and survival on salmon meat as a function of temperature. Food Microbiol., 26, 606-614 (2009). https://doi.org/10.1016/j.fm.2009.04.004
  16. Kim, S.Y., Oh, D.H., Predictive modeling of Bacillus cereus on carrot treated with slightly acidic electrolyzed water and ultrasonication at various storage temperatures, JKSFSN., 43, 1296-1303, (2014). https://doi.org/10.3746/jkfn.2014.43.8.1296
  17. Kim, H.D., Han, S.J., Choi, J.I., Im, Y.K., Optimization of physical factor for amylase production by Arthrobacter sp. by response surface methodology. Korean Chem. Eng. Res., 54, 140-144 (2016). https://doi.org/10.9713/kcer.2016.54.1.140
  18. Williams, S.L., Jensen, R.V., Kuhn, D.D., Stevens, A.M., Analyzing the metabolic capabilities of a Vibrio parahaemolyticus strain that causes early mortality syndrome in shrimp. Aquaculture., 476, 44-48 (2017). https://doi.org/10.1016/j.aquaculture.2017.03.030
  19. Regmi, A., Boyd, E.F., Carbohydrate metabolic systems present on genomic islands are lost and gained in Vibrio parahaemolyticus. BMC Microbiol., 19, 1-20 (2019). https://doi.org/10.1186/s12866-018-1372-8
  20. Ju, C.H., Yeung, P.M., Oesterling, J., Seigerman, D.A., Boor, K.J., Vibrio parahaemolyticus growth under low-iron conditions and survival under high-magnesium conditions. J. Food Prot., 69, 1040-1045 (2006). https://doi.org/10.4315/0362-028X-69.5.1040
  21. Tiruvayipati, S., Bhassu, S., Host, pathogen and the environment: the case of Macrobrachium rosenbergii, Vibrio parahaemolyticus and magnesium. GUT Pathog., 8, 1-8 (2016). https://doi.org/10.1186/s13099-015-0083-z
  22. Soto-Rodriguez, S.A., Lozano-Olvera, R., Palacios-Gonzalez, D.A., Bolan-Mejia, C., Rendon-Aguilar, K.G., Characterization and growth conditions of Vibrio parahaemolyticus strains with different virulence degrees that cause acute hepatopancreatic necrosis disease in Litopenaeus vannamei. J. World Aquacult. Soc., 50, 1002-1015 (2019). https://doi.org/10.1111/jwas.12617