Escherichia coli EAST1 toxin toxicity of variants 17-2 and O 42

EAST1 (EnteroAggregative heat-Stable Toxin 1) is a 4.1 kDa toxin that was first detected in the enteroaggregative Escherichia coli (EAEC) strain 17-2 (O3:H2) isolated from the stools of a Chilean child with diarrhoea. Accordingly, EAST1 is thought to play a role in the pathogenicity of EAEC. The goal of this study was to obtain purified biologically active forms of two EAST1 variants (17-2 and O 42). Purified toxin samples were treated with protein disulfide isomerase (PDI) to ascertain the integrity of the disulfide bridges. Since EAST1 is often compared to STa (heat-Stable Toxin a), both purified EAST1 variants were tested for biological activity using the suckling mouse assay, the reference test for STa. A positive gut to body (G/B) weight ratio was not observed for any of the EAST1 preparations tested, although STa was active. Exposure of the purified toxins to T84 cell monolayers, an epithelial cell line derived from a human colon carcinoma, in modified Ussing flux chambers resulted in a rapidly attained and prolonged increase in short circuit current, a sensitive measure of net ion transport. Responses to 17-2 and O 42 variants were comparable in magnitude and inhibitable by bumetanide and DASU-02, indicating net anion secretion. The results demonstrate that EAST1 toxin stimulates anion secretion by T84 cell monolayers and it is sustained for the duration of toxin exposure.