Polychip‐A High‐Throughput Droplet Microfluidics Platform for Interrogating Microbial Interactions

ABSTRACT Microbial interactions are fundamental to ecology, health, and disease, yet high‐throughput tools to study these complex relationships remain scarce. We introduce Polychip, a fully integrated, high‐throughput droplet microfluidics platform that revolutionizes microbial interaction screening. By seamlessly combining six microfluidic operations onto a single chip, Polychip achieves 99.7% efficiency and accelerates screening by 11–14 times compared to traditional high‐throughput liquid handling robotic methods, with minimal human input. Using the Polychip, we screened 2.24 × 10 6 soil‐extracted microorganisms against the multidrug‐resistant (MDR) pathogen Pseudomonas aeruginosa . This process recovered 1.96 × 10 4 hit droplets with an antimicrobial activity confirmation accuracy of 48%. Three environmental isolates ( Stenotrophomonas sp.) exhibiting high potency and broad‐spectrum antimicrobial activity were identified through this screen. Supernatants from these three environmental isolates suppressed growth of both gram‐negative MDR pathogen Acinetobacter baumannii and gram‐positive methicillin‐resistant Staphylococcus aureus (MRSA). Through whole‐genome sequencing, metabolic pathway analysis, and mass spectrometry, we identified the secreted compound as enterochelin. This demonstrates Polychip's unprecedented single‐cell resolution and high throughput screening capability in rapidly discovering antimicrobial activities, opening new frontiers in combating resistant microbial pathogens, as well as more broadly advancing microbial ecology research.