Cell-based SELEX aptamer selection for electrochemical detection of Fluoribacter bozemanae bacteria

Abstract

Fluoriobacter bozemanae (F. bozemanae) can cause Legionnaires' disease. The conventional methods for the detection of F. bozemanae are laborious, costly and has to be performed in centralized labs by skilled personnel. Therefore, there is an urgent need for rapid detection methods for F. bozemanae. The present work reports the first electrochemical aptasensor for the rapid and specific detection of the most virulent type of Legionellosis. Firstly, high-affinity aptamer was selected against the F. bozemanae using cell-based systematic evolution of ligands by exponential enrichment (Cell SELEX). The aptamers selections were based on 10 rounds of cell-SELEX with live F. bozemanae cells, and the counter-selection was performed against five other closely related Legionella sub-species. The selected oligonucleotide sequences were characterized using the dissociation constant (Kd). Apt-FB6 sequence gave a Kd value as low as 9.516 nM. This selected aptamer was integrated with electrochemical impedance spectroscopy (EIS). The impedimetric aptasensor for F. bozemanae exhibited a detection limit down to 10 CFU/mL. Finally, the aptasensor was tested against Legionella anisa, Legionella micdadei, Fluoriobacter dumoffii, and Legionella pneumophila serogroup 1 and 3. The biosensor didn't show significant cross-reactivity with the closely-related sub-species of Legionella. The selected aptasensor can be further exploited for further biosensing applications for clinical, environmental, and diagnostic purposes.