A droplet microfluidic strategy for cultivation, investigation, and high-throughput isolation of mouse gut microbiome...

Understanding the gut microbiome's intricate dynamics and its impact on host health necessitates the cultivation and isolation of its constituent microorganisms. Traditional culturing techniques often fall short in capturing the diversity of the gut microbiota, particularly for rare and slow-growing species. In this study, we present a droplet microfluidic platform as a high-throughput and efficient method for the cultivation and isolation of mouse gut microorganisms. Droplets, each encapsulating a single cell, were incubated under both aerobic and anaerobic conditions, thereby providing individual microenvironments without nutrient competition and facilitating the growth of a wide range of microorganisms. We validated the platform by successfully cultivating and isolating a diverse array of gut microorganisms, including strains with probiotic potential. A comparative analysis with traditional agar plating techniques revealed a higher number of unique isolates from the droplet cultivation method, demonstrating its enhanced capability to capture the cultivable fraction of the gut microbiome. Beyond isolation, 16S rDNA amplicon sequencing of the diverse microbial cultures in droplets demonstrated that our system reflects changes in microbial diversity induced by dietary interventions in mice. Droplet microfluidics offers a powerful and scalable tool for the high-throughput cultivation, evaluation, and isolation of gut microorganisms, paving the way for deeper insights into the gut microbiome's role in health and disease.IMPORTANCEThe gut microbiome plays a crucial role in health and disease, yet many of its microbial members remain difficult to cultivate using traditional methods. In this study, we present a droplet microfluidic platform that advances our ability to cultivate, isolate, and analyze mouse gut microorganisms. By providing individual microenvironments for single cells, this high-throughput method overcomes limitations of traditional culturing techniques, enhancing microbial diversity recovery compared to standard techniques. Furthermore, this platform can reflect changes in microbial diversity in response to dietary changes in mice, highlighting its potential for studying gut microbial dynamics.