Highly sensitive and fast Legionella spp. in situ detection based on a loop mediated isothermal amplification technique combined to an electrochemical transduction system

Abstract

A rapid highly sensitive genosensor has been developed for monitoring the presence of Legionella spp. in different water systems (domestic hot water, heating/cooling systems or cooling towers) in order to avoid its spreading from the source of contamination. The genosensor integrates a loop mediated isothermal amplification (LAMP) reaction with an electrochemical transduction signal, producing a very simple, rapid to perform and cost effective method, suitable for in situ analyses. This approach detects as low as 10 fg of Legionella nucleic acid, corresponding to only 2 number copies of the bacteria. The use of an electrochemical redox-active double stranded DNA (dsDNA) intercalating molecule, known as methylen blue (MB), allows the immediate electrochemical reading during the DNA polymerization. The sensor can obtain quantitative results in 20 min with a correlation between the electrochemical data and Legionella spp. copy number (at a logarithmic scale) of r = -0.97. In conclusion, a fast, easy to use, and accurate electrochemical genosensor, with high precision, sensitivity, and specificity has been developed for in situ detection of Legionella spp. enabling real time decision making and improving significantly the current detection methods for the prevention and screening of Legionella.

Keywords: Electrochemical dsDNA detection; Genosensor; LAMP; Legionella spp..