Application of PCR to a clinical and environmental investigation of a case of equine botulism

• Published in: Journal of Clinical Microbiology 1994-08, Vol.32 (8), p.1986-1991
• Main Authors: SZABO, E. A, PEMBERTON, J. M, GIBSON, A. M, THOMAS, R. J, PASCOE, R. R, DESMARCHELIER, P. M
• Format: Article
• Language: English
• Subjects: amplification chaine polymerase; Animals; Archives; Bacteriology; Base Sequence; bioassays; Biological and medical sciences; Biological Assay; Botulinum Toxins - biosynthesis; Botulinum Toxins - genetics; botulism; Botulism - epidemiology; Botulism - microbiology; Botulism - veterinary; botulisme; botulismo; caballos; cheval; Clostridium - isolation & purification; Clostridium - metabolism; Clostridium botulinum; DNA Primers; dosage biologique; electroforesis; electrophorese; electrophoresis; ensayo biologico; Epidemiology; Fundamental and applied biological sciences. Psychology; gene; genes; Genes, Bacterial - genetics; Horse Diseases - epidemiology; Horse Diseases - microbiology; Horses; Microbiology; Molecular Sequence Data; nervous system; Neurotoxins - biosynthesis; Neurotoxins - genetics; polymerase chain reaction; Polymerase Chain Reaction - methods; queensland; reaccion de cadenas de polimerasa; Sensitivity and Specificity; sistema nervioso; soil; Soil Microbiology; sol; Specimen Handling; suelo; systeme nerveux; toxinas; toxine; toxins; United Kingdom - epidemiology
• ISSN: 0095-1137; 1098-660X
• DOI: 10.1128/jcm.32.8.1986-1991.1994

PCR for the detection of botulinum neurotoxin gene types A to E was used in the investigation of a case of equine botulism. Samples from a foal diagnosed with toxicoinfectious botulism in 1985 were reanalyzed by PCR and the mouse bioassay in conjunction with an environmental survey. Neurotoxin B was detected by mouse bioassay in culture enrichments of serum, spleen, feces, and intestinal contents. PCR results compared well with mouse bioassay results, detecting type B neurotoxin genes in these samples and also in a liver sample. Other neurotoxin types were not detected by either test. Clostridium botulinum type B was shown to be prevalent in soils collected from the area in which the foal was raised. Four methods were used to test for the presence of botulinum neurotoxin-producing organisms in 66 soil samples taken within a 5-km radius: PCR and agarose gel electrophoresis (types A to E), PCR and an enzyme-linked assay (type B), hybridization of crude alkaline cell lysates with a type B-specific probe, and the mouse bioassay (all types). Fewer soil samples were positive for C. botulinum type B by the mouse bioassay (15%) than by any of the DNA-based detection systems. Hybridization of a type B-specific probe to DNA dot blots (26% of the samples were positive) and PCR-enzyme-linked assay (77% of the samples were positive) were used for the rapid analysis of large numbers of samples, with sensitivity limits of 3 X 10(6) and 3,000 cells, respectively. Conventional detection of PCR products by gel electrophoresis was the most sensitive method (300-cell limit), and in the present environmental survey, neurotoxin B genes only were detected in 94% of the samples.