Electrochemical detection of Legionella pneumophila using DNAzymes and under continuous flow in cooling tower water

Rapid detection of Legionella pneumophila in cooling tower water is crucial to mitigate the fatal consequences of Legionnaires disease. This study presents a microfluidic system that employs RNA-cleaving DNAzymes (RCDs) for continuous real time monitoring of this pathogen directly in a single sample of cooling tower water without the need for lengthy bacterial culture. The RCDs, coupled to microgel magnetic beads, are programmed to release an electroactive DNA barcode in the presence of L. pneumophila, which is detected by a downstream electrochemical sensor in real time. Our system identifies key parameters such as peak current, slope of signal increase, and lag time that correlate with L. pneumophila concentration, achieving a limit of detection of 1.4 × 10³ CFU/mL in buffer and 1.9 × 10³ CFU/mL in cooling tower water, meeting regulatory requirements. This system was further used to identify different serotypes of L. pneumophila amongst other waterborne bacterial species including non pneumophila species of Legionella, creating a highly specific tool for identifying this high-risk pathogen.