On-site detection of methicillin-resistant Staphylococcus aureus (MRSA) utilizing G-quadruplex based isothermal exponential amplification reaction (GQ-EXPAR)

Abstract

Methicillin-resistant Staphylococcus aureus (MRSA) has a high incidence in infectious hospitals and communities, highlighting the need for early on-site detection due to its resistance to methicillin antibiotics. The present study introduces a highly sensitive detection system for mecA, a crucial methicillin marker, utilizing an RCA-based isothermal exponential amplification reaction. The G-quadruplex-based isothermal exponential amplification reaction (GQ-EXPAR) method designs probes to establish G-quadruplex secondary structures incorporating thioflavin T for fluorescence. The system, unlike conventional genetic detection methods, works with portable isothermal PCR devices (Isoquark), facilitating on-site detection. A detection limit of 0.1 fmol was demonstrated using synthetic DNA, and effective detection was proven using thermal lysis. The study also validated the detection of targets swabbed from surfaces within bacterial 3D nanostructures using the GQ-EXPAR method. After applying complementary sequences to the padlock probe for the target, the GQ-EXPAR method can be used on various targets. The developed method could facilitate rapid and accurate diagnostics within MRSA strains.