Ultrahigh-throughput screening of heterogeneous vancomycin-intermediate Staphylococcus aureus based on...

Area of Science:

• Microfluidics
• Bacteriology
• Antimicrobial Resistance

Background:

• Heterogeneous vancomycin-intermediate Staphylococcus aureus (hVISA) complicates treatment due to limitations in standard susceptibility testing.
• Current gold-standard methods like PAP-AUC are time-consuming and labor-intensive.
• There is a critical need for rapid and accurate hVISA detection methods.

Purpose of the Study:

• To develop and validate a rapid microfluidic platform for detecting and isolating hVISA.
• To optimize parameters for efficient hVISA screening using fluorescence-activated droplet sorting (FADS).
• To reduce the time required for hVISA identification compared to conventional methods.

Main Methods:

• A microfluidic system integrated with fluorescence-activated droplet sorting (FADS) was developed.
• Key parameters including droplet generation, bacterial concentration, and fluorescence probes were optimized.
• The system was validated using the hVISA reference strain Mu3 and 15 clinical isolates.

Main Results:

• A high-throughput platform achieved ~30% single-bacterium droplet encapsulation.
• Fluorescence signals were detectable within 6 hours and stable for over a week.
• The platform successfully isolated hVISA subpopulations from Mu3 within 12 hours, confirmed by clinical isolates.

Conclusions:

• The integrated microfluidic and FADS platform enables rapid hVISA detection in 12 hours.
• This technology significantly enhances the enrichment efficiency of drug-resistant bacterial populations.
• The developed system offers a promising alternative to conventional methods for timely hVISA diagnosis.