Detection of Clostridium tetani Neurotoxins Inhibited In Vivo by Botulinum Antitoxin B: Potential for Misleading Mouse Test Results in Food Controls

Abstract

The presence of botulinum neurotoxin-producing Clostridia (BPC) in food sources is a public health concern. In favorable environmental conditions, BPC can produce botulinum neurotoxins (BoNTs) outside or inside the vertebrate host, leading to intoxications or toxico-infectious forms of botulism, respectively. BPC in food are almost invariably detected either by PCR protocols targeted at the known neurotoxin-encoding genes, or by the mouse test to assay for the presence of BoNTs in the supernatants of enrichment broths inoculated with the tested food sample. The sample is considered positive for BPC when the supernatant contains toxic substances that are lethal to mice, heat-labile and neutralized in vivo by appropriate polyclonal antibodies raised against purified BoNTs of different serotypes. Here, we report the detection in a food sample of a Clostridium tetani strain that produces tetanus neurotoxins (TeNTs) with the above-mentioned characteristics: lethal for mice, heat-labile and neutralized by botulinum antitoxin type B. Notably, neutralization occurred with two different commercially available type B antitoxins, but not with type A, C, D, E and F antitoxins. Although TeNT and BoNT fold very similarly, evidence that antitoxin B antiserum can neutralize the neurotoxic effect of TeNT in vivo has not been documented before. The presence of C. tetani strains in food can produce misleading results in BPC detection using the mouse test.

Detection of Active BoNT/C and D by EndoPep-MS Using MALDI Biotyper Instrument and Comparison with the Mouse Test Bioassay

“…However, it does pose ethical issues because it requires the sacrifice of many animals, it is time consuming since four days are needed for confirmation of negative samples, and it requires a large volume of sample [ 11 ]. Furthermore, other substances in the sample, e.g., other toxins or irritating ingredients such as spices in foods, can occasionally lead to inconclusive results [ 12 , 13 ]. Many efforts have been made in recent years to develop alternative laboratory methods to the mouse lethality test and some are very promising.…”

Introduction to the Toxins Special Issue on Botulinum Neurotoxins in the Nervous System: Future Challenges for Novel Indications

“…The last paper was contributed by Bano et al [ 25 ]. This paper reports a relevant observation on the fact that a tetanus neurotoxin (TeNT), a relative of BoNT/B produced by a Clostridia tetani strain, is neutralized by antisera raised against BoNT/B.…”