|Kong, Qiulian - Shanghai Academy Of Agricultural Sciences|
|Rubio, Fernando - Abraxis, Llc|
|Qi, Wenyuan - Shanghai Academy Of Agricultural Sciences|
Submitted to: Austin Immunology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 8/26/2016
Publication Date: 8/30/2016
Publication URL: http://handle.nal.usda.gov/10113/63165
Citation: Kong, Q., Patfield, S.A., Skinner, C.B., Stanker, L.H., Gehring, A.G., Fratamico, P.M., Rubio, F., Qi, W., He, X. 2016. Validation of two new immunoassays for sensative detection of a broad range of shiga toxins. Austin Immunology. 1(2):1007.
Interpretive Summary: Shiga toxin –producing E. coli (STEC) are important causes of foodborne illness. Shiga toxin (Stx) production is the common trait and major virulence determinant of all STEC strains. Individual STEC strains may produce one or more antigenically distinct Stxs, which include 3 subtypes of Stx1 and seven subtypes of Stx2. Reliable methods for detection of Stxs are crucial for identification of STEC. In this study, we evaluated two new commercial assays developed by Abraxis and compared them to the widely used Premier EHEC commercial assay for their ability to detect Stxs using the same set of standards. The new Abraxis Stx1 and Stx2 ELISAs were demonstrated to be more sensitive for most subtypes of Stxs than the Premier EHEC and detect all subtypes of Stxs in phosphate buffer. In contrast, the Premier EHEC test failed to detect Stx2g at 10 ng/mL. When these commercial kits were used to identify STEC strains based on the production of Stx as a marker, the Abraxis kits missed one out of 49 strains, while the Premier EHEC kit missed three out of 49 strains. These results indicate that the new Abraxis kits are a great addition to the existing commercial Stx detection tools and have a great potential for various applications by regulatory agencies, the food industry, and for diagnostic purposes.
Technical Abstract: The objective of this study was to evaluate two newly developed commercial assays, Abraxis Stx1 and Stx2 enzyme-linked immunosorbent assays (ELISAs), for their ability to detect Shiga toxin (Stx) produced by Escherichia coli (E. coli). The performance of these two assays were compared to a widely used commercial assay, the Premier EHEC (by Meridian Bioscience), using the same set of Stx standards developed in our laboratory. Significant improvements in assay sensitivity and specificity were observed using the new test kits. The limit of detection of the new kits for the Stx1a and Stx2a was 25 pg/mL, a 20-fold improvement over the Premier EHEC kit. In addition, the Abraxis kits were able to detect all subtypes of Stx1 and Stx2, but the Premier EHEC ELISA failed to detect Stx2g. For most subtypes of Stxs, the Abraxis kits were more sensitive but for the Stx1d, 2b, and 2d, the Premier EHEC kit showed higher sensitivity. When these commercial kits were used to analyze forty-nine bacterial solates, which represented 13 serotypes and 4 un-typed Shiga toxin-producing E. coli (STEC) strains, the Abraxis kits were capable of identifying all STEC strains based on Stx as the marker, except for one strain that produces Stx2b, and the Premier EHEC ELISA missed two Stx2e- and one Stx2g-producing stains. Furthermore, the Abraxis Stx ELISAs were also able to identify STEC strains using single colonies on agar plates without lengthy enrichment in liquid medium to allow toxin production. The results indicate that the new Abraxis Stx kits are useful for sensitive detection of Stxs, especially for the less common subtypes and they should improve the screening system for STEC.