Selection of G-rich ssDNA aptamers for the detection of enterotoxins of the cholera toxin family

Nerissa A. Molejon, Catherine M. Lapada, Vasso Skouridou and Ciara K. O'Sullivan published in collaboration with the University of the Philippinesand the King Abdulaziz University the following article:

Analytical Biochemistry, Volume 669, 15 May 2023, 115118

Abstract

Cholera and milder diarrheal disease are caused by Vibrio cholerae and enterotoxigenic Escherichia coli and are still a prominent public health concern. Evaluation of suspicious isolates is essential for the rapid containment of acute diarrhea outbreaks or prevention of epidemic cholera. Existing detection techniques require expensive equipment, trained personnel and are time-consuming. Antibody-based methods are also available, but cost and stability issues can limit their applications for point-of-care testing. This study focused on the selection of single stranded DNA aptamers as simpler, more stable and more cost-effective alternatives to antibodies for the co-detection of AB₅ toxins secreted by enterobacteria causing acute diarrheal infections. Cholera toxin and Escherichia coli heat-labile enterotoxin, the key toxigenicity biomarkers of these bacteria, were immobilized on magnetic beads and were used in a SELEX-based selection strategy. This led to the enrichment of sequences with a high % GC content and a dominant G-rich motif as revealed by Next Generation Sequencing. Enriched sequences were confirmed to fold into G-quadruplex structures and the binding of one of the most abundant candidates to the two enterotoxins was confirmed. Ongoing work is focused on the development of monitoring tools for potential environmental surveillance of epidemic cholera and milder diarrheal disease.

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