ABSTRACT
AIMS: To determine whether Clostridium botulinum neurotoxin (BoNT) production in anaerobic culture was affected by temperature and could influence the sandwich ELISA (sELISA) detection of group III toxins in pre-enriched gastrointestinal (GI) contents from clinically suspect cattle botulism cases. METHODS AND RESULTS: Bovine post-mortem GI samples taken from 124 and 96 animals with suspect and nonsuspect botulism, respectively, were pre-enriched anaerobically at 30 and 37°C prior to testing by sELISA. After enrichment at 37°C, BoNT was demonstrated in all clinically suspect bovine botulism cases that had been identified by the mouse bioassay, and enrichment by both temperatures enabled BoNT detection in a number of mouse bioassay-negative suspect cases. CONCLUSIONS: Culture temperature does influence the production of group III BoNT, and incubation at both 30 and 37°C is required for optimum detection. SIGNIFICANCE AND IMPACT OF THE STUDY: The in vitro assay defined in this study has the potential of improving the confirmation rate of clinically suspect cattle botulism cases whilst reducing the use of the costly and ethically sensitive mouse bioassay, the current diagnostic gold standard for BoNT testing.
Subject(s)
Botulinum Toxins/metabolism , Botulism/veterinary , Clostridium botulinum type C/metabolism , Clostridium botulinum type D/metabolism , Gastrointestinal Contents/microbiology , Anaerobiosis , Animals , Biological Assay , Body Temperature , Botulism/diagnosis , Cattle , Cattle Diseases/diagnosis , Enzyme-Linked Immunosorbent Assay/methods , Enzyme-Linked Immunosorbent Assay/veterinaryABSTRACT
In human subjects, the acute tryptophan (TRP) depletion (ATD) paradigm has been shown to have effects on mood and cognition. It is assumed that these effects are mediated through the serotonin system. In this study, we have examined the effects of ATD on the central concentrations of the monoamine transmitters, noradrenaline (NA) and dopamine (DA) as well as on serotonin (5-HT). Effects on NA and DA could also affect mood and cognition. Following oral administration of TRP-containing (TRP+) and TRP-free (TRP-) amino acid mixtures, neurotransmitter concentrations and free plasma TRP concentrations were determined by High Performance Liquid Chromatography (HPLC) with electrochemical detection. Free plasma TRP was significantly and substantially reduced (79%) in rats given a TRP- amino acid mixture when compared with those given a TRP+ mixture. ATD also significantly decreased 5-HT and 5-hydroxyindolacetic acid in the frontal cortex, remaining cortex and hippocampus, but did not significantly reduce these in the striatum. Furthermore, ATD did not significantly alter the concentration of NA and DA in any brain region examined. This study demonstrates that the administration of a TRP- amino acid mixture in rats can reduce free plasma TRP to levels comparable to those reported in human studies. These results indicate that behavioural and cognitive changes produced by ATD in preclinical or clinical studies are likely to be due to specific effects on the serotonergic system.