Development of real-time PCR tests for detecting botulinum neurotoxins A, B, E, F producing Clostridium botulinum, Clostridium baratii and Clostridium butyricum. 2009

P Fach, and P Micheau, and C Mazuet, and S Perelle, and M Popoff
Agence Française de Sécurité Sanitaire des Aliments, Laboratoire d'Etudes et de Recherches sur la Qualité des Aliments et les Procédés Agro-alimentaires, Maisons-Alfort, France. p.fach@afssa.fr

OBJECTIVE To develop real-time PCR assays for tracking and tracing clostridia responsible for human botulism. RESULTS Real-time PCR assays based on the detection of the genes ntnh encoding the nontoxin-nonhaemagglutinin (NTNH) proteins or the most homologous regions of the botulinum neurotoxin (bont) genes have been developed together with four real-time PCR assays, each being specific of the genes bont/A, bont/B, bont/E, bont/F and enables a toxin type-specific identification. The specificity of the assays was demonstrated using a panel of botulinum toxin producing clostridia (29 strains), nonbotulinum toxin producing clostridia (21 strains) and various other bacterial strains. The toxin type-specific assays had a sensitivity of 100 fg-1000 fg of total DNA in the PCR tube (25-250 genome equivalents) which correspond to 10(3) to 10(4) cells ml(-1). After a 48 h enrichment in anaerobic conditions, these PCR assays allowed the detection of Clostridium botulinum type A in a naturally contaminated sample of 'foie gras' suspected in a C. botulinum outbreak. CONCLUSIONS These PCR tests are specific and reliable for detection of heterogeneous BoNT producing clostridia responsible for human botulism. CONCLUSIONS Adoption of these PCR assays is a step forward a reliable and rapid detection of these clostridia in food samples.

UI MeSH Term Description Entries
D008969 Molecular Sequence Data Descriptions of specific amino acid, carbohydrate, or nucleotide sequences which have appeared in the published literature and/or are deposited in and maintained by databanks such as GENBANK, European Molecular Biology Laboratory (EMBL), National Biomedical Research Foundation (NBRF), or other sequence repositories. Sequence Data, Molecular,Molecular Sequencing Data,Data, Molecular Sequence,Data, Molecular Sequencing,Sequencing Data, Molecular
D001905 Botulinum Toxins Toxic proteins produced from the species CLOSTRIDIUM BOTULINUM. The toxins are synthesized as a single peptide chain which is processed into a mature protein consisting of a heavy chain and light chain joined via a disulfide bond. The botulinum toxin light chain is a zinc-dependent protease which is released from the heavy chain upon ENDOCYTOSIS into PRESYNAPTIC NERVE ENDINGS. Once inside the cell the botulinum toxin light chain cleaves specific SNARE proteins which are essential for secretion of ACETYLCHOLINE by SYNAPTIC VESICLES. This inhibition of acetylcholine release results in muscular PARALYSIS. Botulin,Botulinum Neurotoxin,Botulinum Neurotoxins,Clostridium botulinum Toxins,Botulinum Toxin,Neurotoxin, Botulinum,Neurotoxins, Botulinum,Toxin, Botulinum,Toxins, Botulinum,Toxins, Clostridium botulinum
D003013 Clostridium A genus of motile or nonmotile gram-positive bacteria of the family Clostridiaceae. Many species have been identified with some being pathogenic. They occur in water, soil, and in the intestinal tract of humans and lower animals.
D003014 Clostridium botulinum A species of anaerobic, gram-positive, rod-shaped bacteria in the family Clostridiaceae that produces proteins with characteristic neurotoxicity. It is the etiologic agent of BOTULISM in humans, wild fowl, HORSES; and CATTLE. Seven subtypes (sometimes called antigenic types, or strains) exist, each producing a different botulinum toxin (BOTULINUM TOXINS). The organism and its spores are widely distributed in nature.
D004269 DNA, Bacterial Deoxyribonucleic acid that makes up the genetic material of bacteria. Bacterial DNA
D005234 Fatty Liver Lipid infiltration of the hepatic parenchymal cells resulting in a yellow-colored liver. The abnormal lipid accumulation is usually in the form of TRIGLYCERIDES, either as a single large droplet or multiple small droplets. Fatty liver is caused by an imbalance in the metabolism of FATTY ACIDS. Liver Steatosis,Steatohepatitis,Steatosis of Liver,Visceral Steatosis,Liver Steatoses,Liver, Fatty,Steatohepatitides,Steatoses, Liver,Steatoses, Visceral,Steatosis, Liver,Steatosis, Visceral,Visceral Steatoses
D005777 Geese Any of various large waterfowl in the order Anseriformes, especially those of the genera Anser (gray geese) and Branta (black geese). They are larger than ducks but smaller than swans, prefer FRESH WATER, and occur primarily in the northern hemisphere. Goose,Gooses
D006388 Hemagglutinins Agents that cause agglutination of red blood cells. They include antibodies, blood group antigens, lectins, autoimmune factors, bacterial, viral, or parasitic blood agglutinins, etc. Isohemagglutinins,Exohemagglutinins,Hemagglutinin
D006801 Humans Members of the species Homo sapiens. Homo sapiens,Man (Taxonomy),Human,Man, Modern,Modern Man
D000818 Animals Unicellular or multicellular, heterotrophic organisms, that have sensation and the power of voluntary movement. Under the older five kingdom paradigm, Animalia was one of the kingdoms. Under the modern three domain model, Animalia represents one of the many groups in the domain EUKARYOTA. Animal,Metazoa,Animalia

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