Artificial Intelligence-Assisted Automatic Raman-Activated Cell Sorting (AI-RACS) System for Mining Specific Functional Microorganisms in the Microbiome

The microbiome represents the natural presence of microorganisms, and exploring, understanding, and leveraging its functions will bring about significant breakthroughs in life sciences and applications. Raman-activated cell sorting (RACS) enables the correlation of phenotype and genotype at the single-cell level, offering a solution to the bottleneck in microbial community functional analysis caused by challenges in cultivating diverse microorganisms. However, current labor-intensive manual procedures fall short in catering to the demands of single-cell functional analysis in microbial communities. To address this issue, we developed an artificial intelligence-assisted Raman-activated cell sorting system (AI-RACS) that integrates precise single-cell positioning, automated data collection, optical tweezers capture, and single-cell printing to elevate microbial single-cell RACS from manual to automated, validating the efficacy of the system by isolating aluminum-tolerant microbes from acidic soil microbiota. Leveraging the AI-RACS framework, we sorted 13 strains from red soil samples under near-in situ conditions, with all demonstrating strong aluminum tolerance. AI-RACS efficiently segregates microbial cells from intricate environmental samples, investigating their functional attributes and presenting a novel tool for microbial research and applications.