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ARS Home » Pacific West Area » Albany, California » Western Regional Research Center » Foodborne Toxin Detection and Prevention Research » Research » Publications at this Location » Publication #410172

Research Project: Technologies for the Detection of Bacterial and Plant Toxins and Allergens that Impact Food Safety and Food Defense

Location: Foodborne Toxin Detection and Prevention Research

Title: Development of a rapid and sensitive CANARY biosensor assay for the detection of Shiga toxin 2 from Escherichia coli

item Tam, Christina
item WANG, YANGYANG - Smiths Detection
item Du, Wen-Xian
item FLANNERY, ANDREW - Smiths Detection
item He, Xiaohua

Submitted to: Toxins
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 3/12/2024
Publication Date: 3/14/2024
Citation: Tam, C.C., Wang, Y., Du, W.N., Flannery, A.R., He, X. 2024. Development of a rapid and sensitive CANARY biosensor assay for the detection of Shiga toxin 2 from Escherichia coli. Toxins. 16(3). Article 148.

Interpretive Summary: From 2009 to 2021, there were 1,019 reported Shiga-toxin producing Escherichia coli (STEC) outbreaks resulting in 14,010 illnesses, 2,218 hospitalizations, and 43 deaths in US alone. STEC has many different virulence factors, but Shiga toxins (Stxs) are the primary factor responsible for the development of severe complications like hemorrhagic colitis and hemolytic uremia syndrome. Early, rapid, and sensitive detection of STEC/Stx will greatly reduce morbidity and mortality since there are no therapeutics to treat STEC infections. In this study, we developed a B-cell based biosensor assay that was able to detect toxoid spiked in buffer or from toxin produced by various STEC strains. The assay was very sensitive in the pg/mL levels and required minimal sample preparation, small volumes, and rapid with output within 3 minutes after assay initiation. This assay may be a useful tool for environmental and food safety surveillance programs.

Technical Abstract: Shiga-toxin producing Escherichia coli (STEC) causes a wide spectrum of diseases including hemorrhagic colitis and hemolytic uremic syndrome (HUS). Current FSIS testing methods for STEC use the FDA BAM protocol which includes enrichment, cell plating, and genomic sequencing which takes time to complete thus delaying diagnosis and treatment. We wanted to develop a rapid, sensitive, and potentially portable assay that can identify STEC by detecting Shiga toxin (Stx) using the CANARY (Cellular Analysis and Notification of Antigen Risks and Yields) B-cell based biosensor technology. Five potential biosensor cell lines were evaluated for their ability to detect Stx2. Results using the best biosensor cell line (T5) indicated that this biosensor was stable after reconstitution with assay buffer covered in foil at 4 degrees or up to 10 days with an estimated limit of detection (LOD) of approximately 100-200 pg/mL and it detected a broad range of Stx2 subtypes, including Stx2a, Stx2b, Stx2c, Stx2d, and Stx2g) but not cross-react with any closely-related Stx1, abrin, and ricin. Additionally, this assay was able to detect Stx2 in culture supernatants of STEC grown in media with mitomycin C at 8- and 24-hours post-inoculation. These results indicate that the STEC CANARY biosensor developed in this study is sensitive, reproducible, specific, rapid (3 min) and can be a highly useful qualitative tool in environmental and food safety surveillance programs.