High-throughput anaerobic screening for identifying compounds acting against gut bacteria in monocultures or communities

The human gut microbiome is a key contributor to health, and its perturbations are linked to many diseases. Small-molecule xenobiotics such as drugs, chemical pollutants and food additives can alter the microbiota composition and are now recognized as one of the main factors underlying microbiome diversity. Mapping the effects of such compounds on the gut microbiome is challenging because of the complexity of the community, anaerobic growth requirements of individual species and the large number of interactions that need to be quantitatively assessed. High-throughput screening setups offer a promising solution for probing the direct inhibitory effects of hundreds of xenobiotics on tens of anaerobic gut bacteria. When automated, such assays enable the cost-effective investigation of a wide range of compound-microbe combinations. We have developed an experimental setup and protocol that enables testing of up to 5,000 compounds on a target gut species under strict anaerobic conditions within 5 d. In addition, with minor modifications to the protocol, drug effects can be tested on microbial communities either assembled from isolates or obtained from stool samples. Experience in working in an anaerobic chamber, especially in performing delicate work with thick chamber gloves, is required for implementing this protocol. We anticipate that this protocol will accelerate the study of interactions between small molecules and the gut microbiome and provide a deeper understanding of this microbial ecosystem, which is intimately intertwined with human health. This is a high-throughput screening protocol that allows users to test the growth effects of diverse drugs on bacterial monocultures, synthetic communities and communities derived from stool samples.Compared with other methods, it provides increased throughput and cost-effectiveness and can be performed under anaerobic conditions. The authors describe a high-throughput screening strategy that can be carried out anaerobically for studying the effects of drugs in vitro on individual gut microbes or microbial communities created synthetically or obtained from stool samples.