Search for Antibiotic Substances Using High-Throughput Droplet Microfluidics

The rise of multidrug-resistant pathogens has intensified the need for novel antibiotics with unique mechanisms of action, as resistance to existing drugs continues to emerge rapidly after clinical introduction. In the race against time, traditional antibiotic discovery methods have proven insufficient to address this escalating challenge, hindered by the rediscovery of known cdmpounds and a lack of highly parallelized, culture-dependent techniques. Droplet microfluidics has emerged as a transformative technology, providing unmatched speed and efficiency in cultivating and screening microorganisms for secondary metabolite production. By creating millions of controlled microenvironments, this approach enables rapid access to microbial diversity while streamlining the discovery of urgently needed antibiotics. We have developed an ultra-high-throughput droplet microfluidic platform to cultivate and screen microbial resources, including environmental metagenomic libraries in defined expression hosts or microbial extracts from diverse habitats, for secondary metabolite potential. A novel feature of this platform is the use of multiplexed culturomics at the picoliter scale, enabling the evaluation of diverse growth media to target specific segments of microbial diversity in environmental samples, including low-abundance taxa. Furthermore, cultured droplets can be interrogated using reporter strains of clinically relevant pathogens to assess bioactivity. After a co-incubation step, the droplets are analyzed and sorted using an optical fiber-based detection setup that measures fluorescent readouts indicating reporter strain inhibition. Hits identified in the ultra-high-throughput droplet screen are subsequently transferred to high-throughput plate-based assays. To enable this transfer, droplets are upscaled from the picoliter to the nanoliter scale, allowing precise deposition into microtiter plates and facilitating automated, mode-of-action-based screening on the robotic platform. Promising strains are subsequently subjected to metabolic profiling and scaled up to liter­scale fermentations (1-300 L), yielding purified active compounds for LC-MS/MS and NMR analyses with the potential to reveal nature-inspired lead compounds for preclinical development. This multiscale pipeline for antibiotic discovery accelerates the identification of novel anti-infectives and drives innovation against one of the most urgent public health challenges of our time.

PhD start: 12/01/2024