A Rapid, Sensitive, and Portable Biosensor Assay for the Detection of Botulinum Neurotoxin Serotype A in Complex Food Matrices

Botulinum neurotoxin (BoNT) intoxication can lead to the disease botulism, characterized by flaccid muscle paralysis that can cause respiratory failure and death.  Due to the significant morbidity and mortality costs associated with BoNTs high toxicity, developing highly sensitive, rapid, and field-deployable assays are critically important to protect the nation’s food supply against either accidental or intentional contamination. We report here that the B-cell based biosensor assay (CANARY® Zephyr) detects BoNT/A in buffer and various food matrices rapidly in ≤ 40 min, in small volumes ≈ 50 μL, with minimal processing of samples, and is extremely portable (suitcase-sized equipment). BoNT/A was detected at limits of detection (LOD) < 0.075 ng ± 0.02 in assay buffer while milk matrices (non-fat, 2 %, whole milk) increased the LOD to < 0.175 – 0.314 ng. Limits of detection for the assay in complex foods were < 1 ng ± 0.0 (neutralized acidic juices-carrot, orange and apple); < 16.7 ng ± 7.7 (liquid egg); and varied from < 0. 39 – 3.125 ng for solid complex foods (ground beef, green bean baby puree, smoked salmon). These results show that the CANARY® Zephyr assay can be a highly useful tool in clinical, environmental, and food safety surveillance programs.

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A Rapid, Sensitive, and Portable Biosensor Assay for the Detection of Botulinum Neurotoxin Serotype A in Complex Food Matrices

Toxins ◽

10.3390/toxins10110476 ◽

2018 ◽

Vol 10 (11) ◽

pp. 476 ◽

Cited By ~ 2

Author(s):

Christina Tam ◽

Andrew Flannery ◽

Luisa Cheng

Keyword(s):

Botulinum Neurotoxin ◽

Solid Complex ◽

Muscle Paralysis ◽

Cellular Analysis ◽

Smoked Salmon ◽

Whole Milk ◽

Highly Sensitive ◽

Surveillance Programs ◽

Botulinum Neurotoxin Serotype A ◽

Ml Detection

Botulinum neurotoxin (BoNT) intoxication can lead to the disease botulism, characterized by flaccid muscle paralysis that can cause respiratory failure and death. Due to the significant morbidity and mortality costs associated with BoNTs high toxicity, developing highly sensitive, rapid, and field-deployable assays are critically important to protect the nation’s food supply against either accidental or intentional contamination. We report here that the B-cell based biosensor assay CANARY® (Cellular Analysis and Notification of Antigen Risks and Yields) Zephyr detects BoNT/A holotoxin at limits of detection (LOD) of 10.0 ± 2.5 ng/mL in assay buffer. Milk matrices (whole milk, 2% milk and non-fat milk) with BoNT/A holotoxin were detected at similar levels (7.4–7.9 ng/mL). BoNT/A complex was positive in carrot, orange, and apple juices at LODs of 32.5–75.0 ng/mL. The detection of BoNT/A complex in solid complex foods (ground beef, smoked salmon, green bean baby puree) ranged from 14.8 ng/mL to 62.5 ng/mL. Detection of BoNT/A complex in the viscous liquid egg matrix required dilution in assay buffer and gave a LOD of 171.9 ± 64.7 ng/mL. These results show that the CANARY® Zephyr assay can be a highly useful qualitative tool in environmental and food safety surveillance programs.

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Fabrication of a Novel Highly Sensitive and Selective Immunosensor for Botulinum Neurotoxin Serotype A Based on an Effective Platform of Electrosynthesized Gold Nanodendrites/Chitosan Nanoparticles

Sensors ◽

10.3390/s17051074 ◽

2017 ◽

Vol 17 (5) ◽

pp. 1074 ◽

Cited By ~ 14

Author(s):

Rahim Sorouri ◽

Hasan Bagheri ◽

Abbas Afkhami ◽

Jafar Salimian

Keyword(s):

Botulinum Neurotoxin ◽

Chitosan Nanoparticles ◽

Serotype A ◽

Highly Sensitive ◽

Botulinum Neurotoxin Serotype A

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Analysis of Neurotoxin Cluster Genes in Clostridium botulinum Strains Producing Botulinum Neurotoxin Serotype A Subtypes

Applied and Environmental Microbiology ◽

10.1128/aem.02828-07 ◽

2008 ◽

Vol 74 (9) ◽

pp. 2778-2786 ◽

Cited By ~ 57

Author(s):

Mark J. Jacobson ◽

Guangyun Lin ◽

Brian Raphael ◽

Joanne Andreadis ◽

Eric A. Johnson

Keyword(s):

Amino Acid ◽

Clostridium Botulinum ◽

Botulinum Neurotoxin ◽

Amino Acid Sequences ◽

Gene Sequences ◽

Amino Acid Levels ◽

Cluster Type ◽

Botulinum Neurotoxin Serotype A ◽

Cluster Gene ◽

Degree Of Similarity

ABSTRACT Neurotoxin cluster gene sequences and arrangements were elucidated for strains of Clostridium botulinum encoding botulinum neurotoxin (BoNT) subtypes A3, A4, and a unique A1-producing strain (HA− Orfx+ A1). These sequences were compared to the known neurotoxin cluster sequences of C. botulinum strains that produce BoNT/A1 and BoNT/A2 and possess either a hemagglutinin (HA) or an Orfx cluster, respectively. The A3 and HA− Orfx+ A1 strains demonstrated a neurotoxin cluster arrangement similar to that found in A2. The A4 strain analyzed possessed two sets of neurotoxin clusters that were similar to what has been found in the A(B) strains: an HA cluster associated with the BoNT/B gene and an Orfx cluster associated with the BoNT/A4 gene. The nucleotide and amino acid sequences of the neurotoxin cluster-specific genes were determined for each neurotoxin cluster and compared among strains. Additionally, the ntnh gene of each strain was compared on both the nucleotide and amino acid levels. The degree of similarity of the sequences of the ntnh genes and corresponding amino acid sequences correlated with the neurotoxin cluster type to which the ntnh gene was assigned.

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