Quantification of Clostridium botulinum toxin gene expression by competitive reverse transcription-PCR

S McGrath, James Dooley, RW Haylock

Research output: Contribution to journalArticle

33 Citations (Scopus)

Abstract

Clostridium botulinum produces a characteristic botulinum neurotoxin which fan cause an often fatal neuroparalytic condition known as botulism. Although food-borne botulism is rare, critical screening by food companies is necessary to ensure that food products are safe. At present, the food industry assesses the risks of botulinum neurotoxin production by challenge testing to check any nerv food products and to check the efficacy of new storage regimes. Challenge testing involves artificial introduction of defined strains of microorganisms into food, and microbial growth and possible toxin production are then monitored. Botulinum toxin is normally analyzed by using the mouse bioassay. However, the mouse bioassay is expensive, slow, and politically sensitive because of animal rights issues, In this paper we describe adaptation of a new assay, competitive reverse transcription-PCR (RT-PCR), to monitor botulinum neurotoxin production. This method accurately measures the level of toxin-encoding mRNA in C, botulinum cells. Measurement of mRNA should provide a good indication of gene expression as mRNA is turned over rapidly in bacterial cells. In addition, the method is rapid, specific, and sensitive. The competitive RT-PCR method was developed to examine C. botulinum E VH toxin gene expression and was used to investigate the level of toxin production by C, botulinum E VH when the organism was grown in two different types of broth. The results which we obtained with the competitive RT-PCR method demonstrated that this method is more rapid and more sensitive than the mouse bioassay.
Original languageEnglish
Pages (from-to)1423-1428
JournalApplied and Environmental Microbiology
Volume66
Issue number4
DOIs
Publication statusPublished - Apr 2000

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