Korean Journal of Food Science and Technology (한국식품과학회지)

Korean Society of Food Science and Technology (한국식품과학회)
권호 표지

Toxin Gene Analysis of Bacillus cereus and Bacillus thuringiensis Isolated from Cooked Rice

쌀밥에서 분리한 Bacillus cereus와 Bacillus thuringiensis의 독소유전자 분석

  • Jeon, Jong-Hyuk (Department of Food Science and Biotechnology, Kyungwon University) ;
  • Park, Jong-Hyun (Department of Food Science and Biotechnology, Kyungwon University)
  • 전종혁 (경원대학교 식품생물공학과) ;
  • 박종현 (경원대학교 식품생물공학과)
  • Received : 2010.04.12
  • Accepted : 2010.05.14
  • Published : 2010.06.30
Download PDF
⟨ Previous Next ⟩

Abstract

Bacterial contamination of cooked rice was analyzed to evaluate the microbial safety. Thirty raw rice samples were collected in Korea and cooked in an electric rice cooker. Mesophilic aerobe, food-poisoning Bacillus cereus group, and their toxin genes were determined on cooked rice. The percentage of total mesophilic aerobe based on 1-3 log CFU/g was 27% among the samples. Bacillus spp. in MYP selective medium was similar to the number of mesophilic aerobe, whileas Bacillus spp. was detected in most samples after enrichment. Thirty-seven isolates from 30 cooked rices were identified as B. thuringiensis, B. cereus, B. valismortis, B. pumilus, B. coagulans, B. licheniformis, Geobacillus stearothermophilus, and Brevibacillus laterosporus. Twenty isolates (54%), more than half of the isolates, were B. thuringiensis while nine (27%) were identified as B. cereus. All B. thuringiensis isolates possessed non-hemolytic toxin genes and interestingly, seven B. cereus among nine isolates possessed emetic toxin genes. More B. thuringiensis was present on the cooked rice than B. cereus and most B. cereus possessed emetic toxin genes rather than diarrheal toxin genes. Therefore, food-borne outbreak due to B.cereus on the cooked rice kept at room temperature might be examples of emetic food-poisoning.

쌀밥의 미생물 안전성을 평가하기 위하여 쌀을 취반한 후 세균을 분석하였다. 전국에서 생산되는 생쌀 30개를 수집하여 취반 후에 중온성 호기성균과 MYP선택배지에서 Bacillus cereus group를 분리 동정하여 그의 분포도와 독소유전자를 분석하였다. 취반 직후의 쌀밥 27%에서 1-3 log CFU/g정도의 총 중온성 호기균과 거의 같은 정도로의 Bacillus spp.가 존재하는 것으로 나타났다. 그러나 균이 검출되지 않은 시료들도 증균한 후에는 B. cereus group균들이 검출되어 사실상 대부분의 시료에서 Bacillus spp.가 분포하고 있는 것을 알 수 있었다. 이들 시료로부터 37개의 분리하여 동정한 균주는 B. thuringiensis, B. cereus, B. valismortis, B. pumilus, B. coagulans, B. licheniformis, Geobacillus stearo-thermophilus, Brevibacillus laterosporus 등으로 나타났다. 분리 균주중 20개(54%)의 분리주가 B. thuringiensis로 나타났고 그 다음으로 9개(27%)의 B. cereus이였다. 그리고 3개(8%)의 B. valismortis와 각각 1개(3%)의 B. pumilus, B. coagulans, B. licheniformis, Geobacillus stearothermophilus, Brevibacillus laterosporus이였다. B. thuringiensis는 모두에서 non-hemolytic toxin gene(nhe)을 가지고 있었고 9개의 B. cereus중 7균주가 emetic toxin 유전자를 함유하고 있었다. 따라서 쌀밥에는 B. thuringiensis가 B. cereus보다 더 높은 빈도로 분포되어 있고 B. cereus는 설사형 독소유전자 보다는 구토형 독소를 더 많이 가지고 있었다. 취반 후 쌀밥을 상온에서 보관하여 발생되는 B. cereus 식중독은 설사형보다 구토형일 가능성이 더 많을 것으로 보인다.

Keywords

References

  1. Kim SH, Kim JS, Choi JP, Park JH. Prevalence and frequency of food-borne pathogens on unprocessed agricultural and marine products. Korean J. Food Sci. Technol. 38: 594-598 (2006)
  2. Chang TE. Moon SY, Lee KW, Park JM, Han JS, Song OJ, Shin IS. Microflora of manufacturing process and final products of saengshik. Korean J. Food Sci. Technol. 36: 501-506 (2004)
  3. Korea Food and Drug Administration. http://www.kfda.go.kr. accessed on Dec. 18, 2009.
  4. Lechner S. Mayr R. Francis KP. Bacillus weihenstephanensis sp. nov. is a new psychrotolerant species of the Bacillus cereus group. Int. J. Syst. Bacteriol. 48: 1373-1382 (1998) https://doi.org/10.1099/00207713-48-4-1373
  5. Nakamura LK. Bacillus pseudomycoides sp. nov. Int. J. Syst. Bacteriol. 48: 1031-1035 (1998) https://doi.org/10.1099/00207713-48-3-1031
  6. Granum PE. Bacillus cereus and its toxins. J. Appl. Bacteriol. 23(Suppl.): 61S-66S (1994)
  7. Hansen BM, Hoiby PE, Jensen GB, Hendriksen NB. The Bacillus cereus bceT enterotoxin sequence reappraised. FEMS Microbiol. Lett. 223: 21-24 (2003) https://doi.org/10.1016/S0378-1097(03)00249-0
  8. Beecher DJ, Wong ACL. Tripartite hemolysin BL from Bacillus cereus. Hemolytic analysis of component interaction and a model for its characteristic paradoxical zone phenomenon. J. Biol. Chem. 272: 233-239 (1997) https://doi.org/10.1074/jbc.272.1.233
  9. Schoeni JL, Wong AC. Heterogeneity observed in the components of hemolysin BL, an enterotoxin produced by Bacillus cereus. Int. J. Food Microbiol. 53: 159-167 (1999) https://doi.org/10.1016/S0168-1605(99)00158-0
  10. Guinebretiere MH, Broussolle V, Nguyen-The C. Enterotoxigenic profiles of food-poisoning and food-born Bacillus cereus strains. J. Clin. Microbiol. 40: 3053-3056 (2002) https://doi.org/10.1128/JCM.40.8.3053-3056.2002
  11. Agata N, Mori M, Ohta M, Suwan S, Ohtani I, Isobe M. A novel dodecadepsipeptide, cereulide, isolated from Bacillus cereus causes vacuole formation in HEp-2 cells. FEMS Microbiol. Lett. 121: 31-34 (1994)
  12. Agata N, Ohta M, Yokoyama K. Production of Bacillus cereus emetic toxin (cereulide) in various foods. Int. J. Food Microbiol. 73: 23-27 (2002) https://doi.org/10.1016/S0168-1605(01)00692-4
  13. Mikkola R, Saris NE, Grigoriev PA, Andersson MA, Salkinoja-Salonen MS. Ionophoretic properties and mitochondrial effects of cereulide: The emetic toxin of B. cereus. Eur. J. Biochem. 263: 112-117 (1999) https://doi.org/10.1046/j.1432-1327.1999.00476.x
  14. Pirhonen T, Andersson MA, Jskelinen EL, Salkinoja-Salonen MS, Honkanen-Buzalski T, Johansson TM. Biochemical and toxic diversity of Bacillus cereus in a pasta and meat dish associated with a food-poisoning case. Food Microbiol. 22: 87-91 (2005) https://doi.org/10.1016/j.fm.2004.04.002
  15. Agata N, Ohta M, Mori M. Production of an emetic toxin, cereulide, is associated with a specific class of Bacillus cereus. Curr. Microb. 33: 67-69 (1996) https://doi.org/10.1007/s002849900076
  16. Glatz BA, Goepfert JM. Defined conditions for synthesis of Bacillus cereus enterotoxin by fermentor-grown cultures. Appl. Environ. Microbiol. 32: 400-404 (1976)
  17. Turnbull PCB. Bacillus cereus toxins. pp. 397-448. In: Pharmacology of Bacterial Toxins. Dorner F, Drews J (eds). Pergamon Press, Oxford, England (1986)
  18. Jackson SG, Goodbrand RB, Ahmed R, Kasatiya S. Bacillus cereus and Bacillus thuringiensis isolated in a gastroenteritis outbreak investigation. Lett. Appl. Microbiol. 21: 103-105 (1995) https://doi.org/10.1111/j.1472-765X.1995.tb01017.x
  19. Noguchi H. Development of Bacillus thuringiensis in Japen. pp. 283-291. In: Advanced engineered pesticides. Kim L. (eds). Marcel Dekker, New York, NY (1993)
  20. Perani M, Bishop AH, Vaid A. Prevalence of $\beta$-exotoxin, diarrhoeal toxin and specific $\delta$-endotoxin in natural isolates of Bacillus thuringiensis. FEMS Microbiol. Lett. 160: 55-60 (1998)
  21. Damgaard PH, Larsen HD, Hansen BM, Bresciani J, Jorgensen K. Enterotoxin-producing strains of Bacillus thuringiensis isolated from food. Lett. Appl. Microbiol. 23: 146-150 (1996)
  22. Prss BM, Dietrich R, Nibler B, Mrtlbauer E, Scherer S. The hemolytic enterotoxin HBL is broadly distributed among species of the Bacillus cereus group. Appl. Environ. Microb. 65: 5436-5442 (1999)
  23. Hwang JH. Biochemical characteristics and enterotoxin gene distribution of food-borne Bacillus cereus. MS thesis, Kyungwon University, Gyeonggi, Korea (2009)
  24. Yamada S, Ohashi E, Agata N, Venkateswaran K. Cloning and nucleotide sequence analysis of gyr B of Bacillus cereus, B. thuringiensis, B. mycoides, and B. anthracis and their application to the detection of B. cereus in rice. Appl. Environ. Microb. 65: 1483-1490 (1999)
  25. Kuo W-S, Chak K-F. Identification of novel cry-type genes from Bacillus thuringiensis strains on the basis of restriction fragment length polymorphism of the PCR-amplified DNA. Appl. Environ. Microbiol. 62: 1369-1377 (1996)
  26. Rowan NJ, Caldow G, Gemmell CG, Hunter IS. Production of diarrheal enterotoxins and other potential virulence factors by veterinary isolates of Bacillus species associated with nongastrointestinal infections. Appl. Environ. Microb. 69: 2372-2376 (2003) https://doi.org/10.1128/AEM.69.4.2372-2376.2003
  27. Celandroni EG, Salvetti SF, Barsotti C, Baggiani A, Senesi S. Identification and characterization of toxigenic Bacillus cereus isolates responsible for two food-poisoning outbreaks. FEMS Microbiol. Lett. 208: 129-134 (2002) https://doi.org/10.1111/j.1574-6968.2002.tb11072.x
  28. Lund T, De Buyser ML, Granum PE. A new cytotoxin from Bacillus cereus that may cause necrotic enteritis. Mol. Microbiol. 38: 254-261 (2000) https://doi.org/10.1046/j.1365-2958.2000.02147.x
  29. Ehling-Schulz M, Vukov N, Schulz A, Shaheen R, Andersson M, Mrtlbauer E, Scherer S. Identification and partial characterization of the nonribosomal peptide synthetase gene responsible for cereulide production in emetic Bacillus cereus. Appl. Environ. Microb. 71: 105-113 (2005) https://doi.org/10.1128/AEM.71.1.105-113.2005
  30. Kim YS, Oh BC, Shin DH. The extension of the shelf life of cooked rice by the treament with the plant extract and their volatile constituents. Food Sci. Biotechnol. 13: 519-522 (2004)
  31. Park SK, Cho YS, Shon MY, Seo KJ. Occurence and repression of off-odor in cooked rice during storage under low temperature warming condition of electric rice cooker. Korean J. Food Sci. Technol. 29: 919-924 (1997)
  32. Roh HJ, Shin YS, Lee KS, Shin MK. Antimicrobial activity of water extract of green tea against cooked rice putrefactive microorganism. Korean J. Food Sci. Technol. 28: 66-71 (1996)
  33. Oh MH, Cox JM. Development and application of a centrifuagation-plate method to study the biodiversity of Bacillus species in rice products. Food Control 21: 7-12 (2010) https://doi.org/10.1016/j.foodcont.2009.03.008
  34. Lee MS, Chang DS. Distribution and physiological characteristic of Bacillus cereus in rice and rice products. Bull. Korean Fish. Soc. 13: 163-172 (1980)
  35. From C, Hormazabal V, Granum PE. Food poisoning associated with pumilacidin-producing Bacillus pumilus in rice. Int. J. Food Microbiol. 115: 319-324 (2007) https://doi.org/10.1016/j.ijfoodmicro.2006.11.005
  36. Ankolekar C, Rahmati T, Labbe' RG. Detection of toxigenic Bacillus cereus and Bacillus thuringiensis spores in U.S. rice. Int. J. Food Microbiol. 128: 460-466 (2009) https://doi.org/10.1016/j.ijfoodmicro.2008.10.006
  37. Rosenquist H, Smidt L, Anderson SR, Jensen GB, Wilcks A. Occurrence and significance of Bacillus cereus and Bacillus thuringiensis in ready-to-eat food. FEMS Microbiol. Lett. 250: 129-136 (2005) https://doi.org/10.1016/j.femsle.2005.06.054
  38. Cronin UP, Wilkinson MG. The growth, physiology and potential of Bacillus cereus in cooked rice during storage temperature abuse. Food Control 20: 822-282 (2009) https://doi.org/10.1016/j.foodcont.2008.10.018
  39. Svensson B, Monthan A, Shaheen R, Ansesson MA, Salkinoja-Salonen M, Christiansson A. Occurence of emetic toxin producing Bacillus cereus in the dairy production chain. Iut. Dairy J. 16: 740-749 (2006) https://doi.org/10.1016/j.idairyj.2005.07.002
  40. Carlin F, Fricker M, Pielaat A, Heisterkamp S, Shaheen R, Salonen MS, Svensson B, Nguyen-the C, Ehling-Schulz. Emetic toxin-producing strains of B.cereus show distinct characteristics within the Bacillus cereus group. Int. J. Food Microbiol. 109: 132-138 (2006) https://doi.org/10.1016/j.ijfoodmicro.2006.01.022
  41. Apetroaie-Constantin C, Shaheen R, Andrup L, Smidt L, Rita H, Salkinja-Salonen. Environment driven cereulide production by emetic strains of Bacillus cereus. Int. J. Food Microbiol. 127: 60-67 (2008) https://doi.org/10.1016/j.ijfoodmicro.2008.06.006
  42. Rivera AMG, Granum PE, Priest FG. Common occurrence of enterotoxin genes and enterotoxicity in Bacillus thuringiensis. FEMS Microbiol. Lett. 190: 151-155 (2000) https://doi.org/10.1111/j.1574-6968.2000.tb09278.x