Document Detail

Multiplex PCR for detection of botulinum neurotoxin-producing clostridia in clinical, food, and environmental samples.
MedLine Citation:
PMID:  19684163     Owner:  NLM     Status:  MEDLINE    
Botulinum neurotoxin (BoNT), the most toxic substance known, is produced by the spore-forming bacterium Clostridium botulinum and, in rare cases, also by some strains of Clostridium butyricum and Clostridium baratii. The standard procedure for definitive detection of BoNT-producing clostridia is a culture method combined with neurotoxin detection using a standard mouse bioassay (SMB). The SMB is highly sensitive and specific, but it is expensive and time-consuming and there are ethical concerns due to use of laboratory animals. PCR provides a rapid alternative for initial screening for BoNT-producing clostridia. In this study, a previously described multiplex PCR assay was modified to detect all type A, B, E, and F neurotoxin genes in isolated strains and in clinical, food, environmental samples. This assay includes an internal amplification control. The effectiveness of the multiplex PCR method for detecting clostridia possessing type A, B, E, and F neurotoxin genes was evaluated by direct comparison with the SMB. This method showed 100% inclusivity and 100% exclusivity when 182 BoNT-producing clostridia and 21 other bacterial strains were used. The relative accuracy of the multiplex PCR and SMB was evaluated using 532 clinical, food, and environmental samples and was estimated to be 99.2%. The multiplex PCR was also used to investigate 110 freshly collected food and environmental samples, and 4 of the 110 samples (3.6%) were positive for BoNT-encoding genes.
Dario De Medici; Fabrizio Anniballi; Gary M Wyatt; Miia Lindström; Ute Messelhäusser; Clare F Aldus; Elisabetta Delibato; Hannu Korkeala; Michael W Peck; Lucia Fenicia
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Publication Detail:
Type:  Comparative Study; Evaluation Studies; Journal Article; Research Support, Non-U.S. Gov't     Date:  2009-08-14
Journal Detail:
Title:  Applied and environmental microbiology     Volume:  75     ISSN:  1098-5336     ISO Abbreviation:  Appl. Environ. Microbiol.     Publication Date:  2009 Oct 
Date Detail:
Created Date:  2009-10-08     Completed Date:  2009-12-01     Revised Date:  2010-09-24    
Medline Journal Info:
Nlm Unique ID:  7605801     Medline TA:  Appl Environ Microbiol     Country:  United States    
Other Details:
Languages:  eng     Pagination:  6457-61     Citation Subset:  IM    
Department of Veterinary Public Health and Food Safety, Istituto Superiore di Sanità, Viale Regina Elena 299, 00161 Rome, Italy.
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MeSH Terms
Base Sequence
Biological Assay / statistics & numerical data
Botulinum Toxins / biosynthesis*,  genetics*
Clostridium / genetics*,  isolation & purification*,  metabolism
Clostridium botulinum / genetics,  isolation & purification,  metabolism
DNA Primers / genetics
DNA, Bacterial / genetics
Environmental Microbiology*
Food Microbiology*
Genes, Bacterial
Neurotoxins / biosynthesis*,  genetics*
Polymerase Chain Reaction / methods*,  statistics & numerical data
Sensitivity and Specificity
Grant Support
//Biotechnology and Biological Sciences Research Council
Reg. No./Substance:
0/Botulinum Toxins; 0/DNA Primers; 0/DNA, Bacterial; 0/Neurotoxins

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