Development of a Sensitive Aptamer-based PCR Method with Magnetic Immunoseparation for Detection of Salmonella Typhimurium in Ground Turkey

Introduction: Aptamers are single-stranded oligonucleotide ligands that can bind targets with high affinity and specificity. They have been widely studied in the field of diagnostics as alternatives to antibodies due to their favorable features such as easy labeling, temperature tolerance, and lower cost. PCR methods have been used broadly to detect and identify foodborne pathogens because of its simplicity, rapidity, and specificity.

Purpose: The aim of this research was to develop a sensitive aptamer-based PCR method coupled with magnetic immunoseparation for detection of Salmonella Typhimurium in ground turkey.

Methods: First, biotinylated polyclonal anti-Salmonella Typhimurium antibody was immobilized on streptavidin-coated magnetic nanobeads to capture Salmonella Typhimurium. Second, the selected aptamer B5 was added to bind to the surface of Salmonella Typhimurium captured by the magnetic nanobeads. Third, the aptamer B5 was released by heating and amplified by PCR. Finally, PCR products were separated with 2% agarose gel electrophoresis and visualized with a UV trans-illuminator.

Results: DNA aptamer specific for Salmonella Typhimurium had been selected using quartz crystal microbalance based SELEX. The developed assay was able to detect 10² CFU/ml of Salmonella Typhimurium in pure culture and 10³ CFU/ml of Salmonella Typhimurium in ground turkey.

Significance: This study demonstrated the feasibility and application of an aptamer PCR method coupled with magnetic immunoseparation for sensitive detection of Salmonella Typhimurium in ground turkey.