Rapid, Quantitative PCR Monitoring of Growth of Clostridium botulinum Type E in Modified-Atmosphere-Packaged Fish
AUTOR(ES)
Kimura, B.
FONTE
American Society for Microbiology
RESUMO
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.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=92548Documentos Relacionados
- Growth of Listeria monocytogenes and Yersinia enterocolitica on Cooked Modified-Atmosphere-Packaged Poultry in the Presence and Absence of a Naturally Occurring Microbiota
- Thermal Inactivation of Nonproteolytic Clostridium botulinum Type E Spores in Model Fish Media and in Vacuum-Packaged Hot-Smoked Fish Products
- Growth potential of Clostridium botulinum in fresh mushrooms packaged in semipermeable plastic film.
- Isolation of Clostridium botulinum type E from Cayuga Lake fish.
- Relative Neurotoxin Gene Expression in Clostridium botulinum Type B, Determined Using Quantitative Reverse Transcription-PCR
