This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowReprints and Permissions
Right arrow Copyright Information
Right arrow Books from ASM Press
Right arrow MicrobeWorld
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Kimura, B.
Right arrow Articles by Fujii, T.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Kimura, B.
Right arrow Articles by Fujii, T.
Agricola
Right arrow Articles by Kimura, B.
Right arrow Articles by Fujii, T.

 Previous Article  |  Next Article 

Applied and Environmental Microbiology, January 2001, p. 206-216, Vol. 67, No. 1
0099-2240/01/$04.00+0   DOI: 10.1128/AEM.67.1.206-216.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

Rapid, Quantitative PCR Monitoring of Growth of Clostridium botulinum Type E in Modified-Atmosphere-Packaged Fish

B. Kimura,1,* S. Kawasaki,1 H. Nakano,2 and T. Fujii1

Tokyo University of Fisheries, Department of Food Science and Technology, Tokyo 108-8477,1 and Hiroshima University, Department of Food Microbiology and Hygiene, Faculty of Applied Biological Science, Hiroshima 739-8528,2 Japan

Received 13 July 2000/Accepted 2 November 2000

A rapid, quantitative PCR assay (TaqMan assay) which quantifies Clostridium botulinum type E by amplifying a 280-bp sequence from the botulinum neurotoxin type E (BoNT/E) gene is described. With this method, which uses the hydrolysis of an internal fluoregenic probe and monitors in real time the increase in the intensity of fluorescence during PCR by using the ABI Prism 7700 sequence detection system, it was possible to perform accurate and reproducible quantification of the C. botulinum type E toxin gene. The sensitivity and specificity of the assay were verified by using 6 strains of C. botulinum type E and 18 genera of 42 non-C. botulinum type E strains, including strains of C. botulinum types A, B, C, D, F, and G. In both pure cultures and modified-atmosphere-packaged fish samples (jack mackerel), the increase in amounts of C. botulinum DNA could be monitored (the quantifiable range was 102 to 108 CFU/ml or g) much earlier than toxin could be detected by mouse assay. The method was applied to a variety of seafood samples with a DNA extraction protocol using guanidine isothiocyanate. Overall, an efficient recovery of C. botulinum cells was obtained from all of the samples tested. These results suggested that quantification of BoNT/E DNA by the rapid, quantitative PCR method was a good method for the sensitive assessment of botulinal risk in the seafood samples tested.

* Corresponding author. Mailing address: Tokyo University of Fisheries, Department of Food Science and Technology, Kounan 4-5-7, Minato-ku, Tokyo 108-8477, Japan. Phone: 81-3-5463-0603. Fax: 81-3-5463-0602. E-mail: kimubo{at}tokyo-u-fish.ac.jp.

Applied and Environmental Microbiology, January 2001, p. 206-216, Vol. 67, No. 1
0099-2240/01/$04.00+0   DOI: 10.1128/AEM.67.1.206-216.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


This article has been cited by other articles:

  • Artin, I., Bjorkman, P., Cronqvist, J., Radstrom, P., Holst, E. (2007). First Case of Type E Wound Botulism Diagnosed Using Real-Time PCR. J. Clin. Microbiol. 45: 3589-3594 [Abstract] [Full Text]  
  • Couesnon, A., Raffestin, S., Popoff, M. R. (2006). Expression of botulinum neurotoxins A and E, and associated non-toxin genes, during the transition phase and stability at high temperature: analysis by quantitative reverse transcription-PCR.. Microbiology 152: 759-770 [Abstract] [Full Text]  
  • Song, Y., Liu, C., Finegold, S. M. (2004). Real-Time PCR Quantitation of Clostridia in Feces of Autistic Children. Appl. Environ. Microbiol. 70: 6459-6465 [Abstract] [Full Text]  
  • Nol, P., Williamson, J. L., Rocke, T. E., Yuill, T. M. (2004). Detection of Clostridium botulinum Type C Cells in the Gastrointestinal Tracts of Mozambique Tilapia (Oreochromis mossambicus) by Polymerase Chain Reaction. J Wildl Dis 40: 749-753 [Abstract] [Full Text]  
  • O'Mahony, J., Hill, C. (2004). Rapid Real-Time PCR Assay for Detection and Quantitation of Mycobacterium avium subsp. paratuberculosis DNA in Artificially Contaminated Milk. Appl. Environ. Microbiol. 70: 4561-4568 [Abstract] [Full Text]  
  • Takahashi, H., Kimura, B., Yoshikawa, M., Fujii, T. (2003). Cloning and Sequencing of the Histidine Decarboxylase Genes of Gram-Negative, Histamine-Producing Bacteria and Their Application in Detection and Identification of These Organisms in Fish. Appl. Environ. Microbiol. 69: 2568-2579 [Abstract] [Full Text]  
  • Fach, P., Perelle, S., Dilasser, F., Grout, J., Dargaignaratz, C., Botella, L., Gourreau, J.-M., Carlin, F., Popoff, M. R., Broussolle, V. (2002). Detection by PCR-Enzyme-Linked Immunosorbent Assay of Clostridium botulinum in Fish and Environmental Samples from a Coastal Area in Northern France. Appl. Environ. Microbiol. 68: 5870-5876 [Abstract] [Full Text]  


Copyright © 2009 by the American Society for Microbiology.   For an alternate route to Journals.ASM.org, visit: http://intl-journals.asm.org | More Info»