Multiple Cross Displacement Amplification Coupled With Gold Nanoparticles-Based Lateral Flow Biosensor for Detection...

Area of Science:

• Microbiology
• Molecular Biology
• Biosensing

Background:

• The rapid spread of the mobilized colistin resistance (mcr-1) gene in Enterobacteriaceae poses a significant threat to treating severe infections.
• Colistin resistance necessitates rapid and accurate detection methods for effective control.

Purpose of the Study:

• To establish a simple, rapid, specific, and sensitive method for identifying the mcr-1 gene.
• To develop a diagnostic tool suitable for surveillance in clinical and livestock environments.

Main Methods:

• Developed a multiple cross displacement amplification (MCDA) assay combined with a gold nanoparticles-based lateral flow biosensor (LFB) for mcr-1 gene detection.
• The MCDA-LFB assay operates at a constant isothermal temperature (63°C) for 30 minutes, with results available from the LFB within 2 minutes.
• The entire process, from DNA extraction to result interpretation, is completed in under 60 minutes.

Main Results:

• The assay demonstrated high analytical specificity, correctly identifying all 16 mcr-1-positive strains and yielding negative results for non-mcr-1 isolates.
• The sensitivity reached as low as 600 fg of plasmid DNA per reaction in pure cultures.
• In spiked fecal samples, the sensitivity was approximately 4.5 × 10^3 CFU/mL, equivalent to about 4.5 CFU per reaction.

Conclusions:

• The developed MCDA-LFB assay is a highly efficient tool for the rapid and reliable detection of the mcr-1 gene.
• This technique is suitable for widespread surveillance of mcr-1 gene dissemination in both clinical settings and the livestock industry.
• The assay's simplicity, speed, and sensitivity contribute to combating the threat of colistin resistance.