ARS | Publication request: Evaluation of a quali embryo model for the detection of botulism toxin type A activity

Submitted to: Interagency Botulism Research Committee
Publication Type: Proceedings
Publication Acceptance Date: August 13, 2008
Publication Date: September 14, 2008
Citation: Buhr, R.J., Bourassa, D.V., Cox Jr, N.A., Richardson, L.J., Phillips, R.W., Kelley, L.C. 2008. Evaluation of a quali embryo model for the detection of botulism toxin type A activity. Interagency Botulism Research Committee. P.225.

Technical Abstract: The Japanese quail embryo (Coturnix japonica) was evaluated for use as a bioassay to detect biologically active botulinum toxin serotype A (BoNT/A). Day 15 of incubation embryos were injected with decreasing dosages of BoNT/A from 250 to 0.5 ng of toxin. At 1 day post-injection, embryos receiving 20 ng of BoNT or higher had more than 74% of the embryos determined to be nonviable; embryos injected with 10, 5, 1, or 0.5 ng of BoNT had 70, 65, 33, and 45% of the embryos determined as nonviable, respectively. The control group (0 ng BoNT) maintained 100% viability 1 day post-injection. Premixing of the BoNT/A with serotype A specific antibody clearly demonstrated that the depression in the ability of the quail embryos to pip and hatch was attributable to biologically active botulinum toxin. When the antibody was preincubated with BoNT dosages from 50 to 5 ng, embryos maintained viability of greater than 64% through 3 days post-injection compared to viability of 24% for those embryos injected at the same dosages without antibody preincubation. The quail embryo bioassay detected biologically active BoNT/A from 0.5 to 250 ng at 1 day post-injection. Based on these data the LD50 for quail embryos was approximately 0.5 ng BoNT / embryo (71 µg BoNT / kg of body weight). Injection of day 15 quail embryos in the neck with BoNT did not interfere with the progression of yolk sac retraction into the abdominal cavity, but BoNT restricted aircell and eggshell pipping initiating morbidity and nonviable status enabling preemptive euthanasia. Results from current experiments to determine the minimal detectable dose in quail embryos will be presented.