Visual and Rapid Detection of Leptospira interrogans Using Multiple Cross-Displacement Amplification Coupled with Nanoparticle-Based Lateral Flow Biosensor

摘要

Leptospirosis is a worldwide zoonosis caused by spirochetes from the genus Leptospira. In the present study, a visual and rapid method for detecting Leptospira interrogans was developed based on multiple cross-displacement amplification (MCDA) and nanoparticle-based lateral flow biosensor (LFB). A set of 10 primers was specifically designed to recognize 10 regions of the lipL 41 gene of L. interrogans. The MCDA reaction was optimized at 64 degrees C for only 40 min, and the amplification products were directly applied to the biosensor. The entire process, including DNA extraction (25 min), MCDA reaction (40 min), and result interpretation (similar to 2 min), could be completed within 70 min. Amplification products were detectable from as few as 10 genomic equivalents per reaction of pure L. interrogans DNA. No cross-reaction with nonpathogenic Leptospira or other bacteria was observed. The MCDA-LFB method established in the current report is suitable for the rapid screening of L. interrogans in clinical, animal, and environmental samples.