Validation of Endopep-MS for qualitativedetection of BoNT/A, /C, /CD, /D, and /DC in animalspecimens and feed.
Project Number1U18FD008342-01
Contact PI/Project LeaderPOPPENGA, ROBERT HOWARD
Awardee OrganizationUNIVERSITY OF CALIFORNIA AT DAVIS
Description
Abstract Text
PROJECT SUMMARY/ABSTRACT
Botulism is a serious paralytic disease that affects both humans and animals which is caused by botulinum
toxins (BoNTs). Animal botulism outbreaks in farm animals, in particular, not only affects animals but also
result in financial losses for the industry and increases the risk of human exposure to BoNTs through animal
products such as milk. To prepare for a response to the future outbreaks, a fast, sensitive, and reliable
detection method for BoNTs is needed. The Endopep-MS method is one of the most promising methods that
has been used for confirmation of human botulism by the Centers for Disease Control and Prevention (CDC)
for years. However, a comprehensive validation study of the Endopep-MS for animalspecimens and feed is
lacking, and none of the veterinary diagnostic laboratories in the US have standard procedures for performing
this method. To fill the knowleage gap, our team at the California Animal Health and Food Safety Laboratory
(CAHFS), a founding member of VET-LIRN, is proposing a comprehensive validation study of the Endopep-MS
for qualitativedetection of BoNT/A, /C, /CD, /D, and /DC in animalspecimens and feed. Our study has three
specific aims as follows.
Specific Aim #1: To validate the Endopep-MS for qualitativedetection of BoNT/A, /C, /CD, /D, and /DC
in animalspecimens and feed. We will validate and compare the developed method to the standard mouse
bioassay (MBA) using toxin-spiked samples and archived samples collected during natural botulism cases.
The sample matrices include but are not limited to animal sera, liver, gastrointestinal contents, and feed.
Specific Aim #2: To develop monoclonal antibodies for the toxin extraction step in the Endopep-MS
method. Monoclonal antibodies are key for the Endopep-MS to achieve selectivity toward BoNT serotypes in
samples. Since there is currently only one source of the monoclonal antibodies in the US, we will collaborate
with Dr. Christina Tam at United States Department of Agriculture (USDA) and develop new monoclonal
antibodies for the method.
Specific Aim #3: To assess the effectiveness of polyclonal antibodies for the toxin extraction step in
the Endopep-MS method. The limited source of monoclonal antibodies can delay research development and
adoption of this method, an issue we have experienced ourselves. Alternately, polyclonal antibodies are
commercially available but the ability of polyclonal antibodies for BoNTs extraction has not been completely
explored. We will complete this knowledge gap by investigating capability of polyclonal antibodies for
extraction of BoNT/C, /CD, /D, and /DC.
If this project is funded and achieved, the lethal and time-consuming MBA that has been performed at CAHFS
for close to 30 years can be avoided. Our laboratory, as part of the Vet-LIRN, will have the Endopep-MS ready
to support botulism testing during outbreaks and large-scale animalfeed emergency events.
Public Health Relevance Statement
PROJECT NARRATIVE
If successful, this project will allow for the rapid detection of multiple botulinum toxin serotypes in feed and
animal samples, leading to an early diagnosis of botulism. Botulinum toxins are the most powerful bacterial
toxins and are also among the most toxic substances yet identified; they pose a serious threat to animals and
humans. Botulism in food animals can expose humans to the toxins via consumption of contaminated
products such as milk.
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