Novel CRISPR-Cas13 Biosensor for Real-Time Detection of Antimicrobial Resistance Genes in Environmental Microbiomes

Background: The emergence and spread of antimicrobial resistance (AMR) genes in environmental microbiomes pose a critical threat to global health security. Current detection methods are time-consuming and often lack the sensitivity required for early environmental surveillance. Methods We developed a novel CRISPR-Cas13a-based biosensor system coupled with isothermal amplification for rapid, field-deployable detection of clinically relevant AMR genes (blaNDM-1, mcr-1, and vanA) in environmental samples. The system integrates microfluidic sample processing with fluorescent readout and smartphone-based detection. Results Our biosensor demonstrated exceptional sensitivity with detection limits of 10 copies/μL for target AMR genes, achieving 100% specificity against non-target sequences. Field testing across 150 environmental samples from wastewater treatment plants, agricultural runoff, and hospital effluents revealed previously undetected AMR hotspots. The system provided results within 45 minutes compared to 72 hours for conventional PCR-based methods. Longitudinal monitoring revealed seasonal fluctuations in AMR gene prevalence, with peak concentrations coinciding with agricultural antibiotic usage patterns. Conclusions This innovative biosensor platform enables rapid, sensitive detection of AMR genes in environmental settings, providing a powerful tool for real-time surveillance and early warning systems. The technology addresses critical gaps in current AMR monitoring capabilities and offers significant potential for global implementation in resource-limited settings.