Novel approach for a user-friendly aptamer-based detection kit for Staphylococcus aureus in biological fluids

Staphylococcus aureus is a common Gram-positive pathogen, present in inadequately treated food, causing a wide spectrum of diseases and one of the leading causes of mortality and morbidity in health care associated infections. Nowadays, antibiotic-resistant strains of S. aureus are seriously challenging global monitoring platforms. Currently, the most important detection methods are classical cultures and biochemical tests. However, they are time-consuming and not highly sensitive. For this reason, there is currently a huge demand for rapid methods for S. aureus detection to be incorporated into point-of-care diagnostic systems. A novel and highly performing approach is offered by aptamers, tailored DNA or RNA sequences acting as artificial recognition elements for conserved epitopes on the bacterial surface. The technological challenges for the use of aptamers-based biosensors in the clinical practice is the lack of materials for their high-density immobilization. Since aptamers interacting with S. aureus are already known, we took advantage of this knowledge tackling two principal drawbacks of the aptamer-based biosensor available to date: the lack of materials for high-density surface immobilization with aptamers and the restriction in the user-friendly read-out of the detected molecules. We developed a device where the detection of S. aureus is based on the displacement by bacteria of a fluorescein-labeled peptide that selectively recognizes the aptamer. Nanostructured zirconium dioxide supports have been used for immobilizing the aptamers, allowing high-density immobilization while retaining their functionality. The displacement of the peptide is associated with a detectable change in the fluorescence of the labeled peptide. For the design of peptides specific for S. aureus aptamers we exploited the energy-based amino acid-base recognition code, previously obtained for several protein-DNA complexes. This approach can be expanded towards any biological agent for which selective aptamers have been identified, allowing immediate applications in clinical and environmental monitoring, particularly in those conditions where a rapid, and specific detection of one or multiple bacterial agents is needed.