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Open AccessArticle

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

1
Special Bacteriology Laboratory, SCT2—Istituto Zooprofilattico Sperimentale delle Venezie, Vicolo Mazzini 4, 31020 Fontane di Villorba, Italy
2
National Reference Centre for Botulism, Microbiological Foodborne Hazard Unit., Department of Food Safety, Nutrition and Veterinary Public Health, Istituto Superiore di Sanità, viale Regina Elena 299, 00161 Rome, Italy
3
Centers for Disease Control and Prevention, National Center for Environmental Health, Division of Laboratory Sciences, 4770 Buford Hwy NE, Atlanta, GA 30341, USA
*
Author to whom correspondence should be addressed.
Toxins 2021, 13(1), 10; https://doi.org/10.3390/toxins13010010
Received: 30 October 2020 / Revised: 18 December 2020 / Accepted: 21 December 2020 / Published: 24 December 2020
(This article belongs to the Special Issue Clostridium Neurotoxins)
Botulinum neurotoxins (BoNTs) are among the most poisonous known biological substances, and therefore the availability of reliable, easy-to use tools for BoNT detection are important goals for food safety and human and animal health. The reference method for toxin detection and identification is the mouse bioassay (MBA). An EndoPep-MS method for BoNT differentiation has been developed based on mass spectrometry. We have validated and implemented the EndoPep-MS method on a Bruker MALDI Biotyper for the detection of BoNT/C and D serotypes. The method was extensively validated using experimentally and naturally contaminated samples comparing the results with those obtained with the MBA. Overall, the limit of detection (LoD) for both C and D toxins were less than or equal to two mouse lethal dose 50 (mLD50) per 500 µL for all tested matrices with the exception of feces spiked with BoNT/C which showed signals not-related to specific peptide fragments. Diagnostic sensitivity, specificity and positive predictive value were 100% (95% CI: 87.66–100%), 96.08% (95% CI: 86.54–99.52%), and 93.33% (95% CI: 78.25–98.20%), respectively, and accuracy was 97.47% (95% CI: 91.15–99.69%). In conclusion, the tests carried out showed that the EndoPep-MS method, initially developed using more powerful mass spectrometers, can be applied to the Bruker MALDI Biotyper instrument with excellent results including for detection of the proteolytic activity of BoNT/C, BoNT/D, BoNT/CD, and BoNT/DC toxins. View Full-Text
Keywords: Clostridium botulinum; BoNT/C; BoNT/D; BoNT/CD; BoNT/DC; MALDI Biotyper Clostridium botulinum; BoNT/C; BoNT/D; BoNT/CD; BoNT/DC; MALDI Biotyper
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MDPI and ACS Style

Drigo, I.; Tonon, E.; Pascoletti, S.; Anniballi, F.; Kalb, S.R.; Bano, L. Detection of Active BoNT/C and D by EndoPep-MS Using MALDI Biotyper Instrument and Comparison with the Mouse Test Bioassay. Toxins 2021, 13, 10. https://doi.org/10.3390/toxins13010010

AMA Style

Drigo I, Tonon E, Pascoletti S, Anniballi F, Kalb SR, Bano L. Detection of Active BoNT/C and D by EndoPep-MS Using MALDI Biotyper Instrument and Comparison with the Mouse Test Bioassay. Toxins. 2021; 13(1):10. https://doi.org/10.3390/toxins13010010

Chicago/Turabian Style

Drigo, Ilenia; Tonon, Elena; Pascoletti, Simone; Anniballi, Fabrizio; Kalb, Suzanne R.; Bano, Luca. 2021. "Detection of Active BoNT/C and D by EndoPep-MS Using MALDI Biotyper Instrument and Comparison with the Mouse Test Bioassay" Toxins 13, no. 1: 10. https://doi.org/10.3390/toxins13010010

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