Interindividual Variation in Dietary Carbohydrate Metabolism by Gut Bacteria Revealed with Droplet Microfluidic Culture

Abstract

Culture and screening of gut bacteria enable testing of microbial function and therapeutic potential. However, the diversity of human gut microbial communities (microbiota) impedes comprehensive experimental studies of individual bacterial taxa. Here, we combine advances in droplet microfluidics and high-throughput DNA sequencing to develop a platform for separating and assaying growth of microbiota members in picoliter droplets (MicDrop). MicDrop enabled us to cultivate 2.8 times more bacterial taxa than typical batch culture methods. We then used MicDrop to test whether individuals possess similar abundances of carbohydrate-degrading gut bacteria, using an approach which had previously not been possible due to throughput limitations of traditional bacterial culture techniques. Single MicDrop experiments allowed us to characterize carbohydrate utilization among dozens of gut bacterial taxa from distinct human stool samples. Our aggregate data across nine healthy stool donors revealed that all of the individuals harbored gut bacterial species capable of degrading common dietary polysaccharides. However, the levels of richness and abundance of polysaccharide-degrading species relative to monosaccharide-consuming taxa differed by up to 2.6-fold and 24.7-fold, respectively. Additionally, our unique dataset suggested that gut bacterial taxa may be broadly categorized by whether they can grow on single or multiple polysaccharides, and we found that this lifestyle trait is correlated with how broadly bacterial taxa can be found across individuals. This demonstration shows that it is feasible to measure the function of hundreds of bacterial taxa across multiple fecal samples from different people, which should in turn enable future efforts to design microbiota-directed therapies and yield new insights into microbiota ecology and evolution. IMPORTANCE Bacterial culture and assay are components of basic microbiological research, drug development, and diagnostic screening. However, community diversity can make it challenging to comprehensively perform experiments involving individual microbiota members. Here, we present a new microfluidic culture platform that makes it feasible to measure the growth and function of microbiota constituents in a single set of experiments. As a proof of concept, we demonstrate how the platform can be used to measure how hundreds of gut bacterial taxa drawn from different people metabolize dietary carbohydrates. Going forward, we expect this microfluidic technique to be adaptable to a range of other microbial assay needs.

Droplet-based high-throughput cultivation for accurate screening of antibiotic resistant gut microbes

“…However, some bacteria require minimal medium with specific carbohydrates, vitamins, or trace elements ( Tramontano et al, 2018 ; Oberhardt et al, 2015 ) while others utilize surface features such as hydrophobicity, roughness, and surface chemistry to form biofilms and proliferate ( Tuson and Weibel, 2013 ). Further enrichment of anaerobic organisms within our droplet platform could be achieved by incorporating droplet generation with defined medium ( Villa et al, 2020 ), combinatorially generated gradients of medium ( Churski et al, 2012 ), or varying the droplet surfactant chemistries to improve biofilm formation ( Chang et al, 2015 ).…”

“…One of the essential steps to address such shortcomings is microbial cultivation, which enables the recovery of complete reference genomes ( Mukherjee et al, 2017 ), accurate identification of taxonomy and functional potential of new strains ( Forster et al, 2019 ; Zou et al, 2019 ), and validation of causality through perturbation experiments ( Schmidt et al, 2018 ). Microbial cultivation is currently experiencing a pronounced revival ( Forster et al, 2019 ; Zou et al, 2019 ; Browne et al, 2016 ; Lagier et al, 2016 ; Villa et al, 2020 ), yet the majority of cultivation efforts that rely on traditional cultivation strategies require arduous manual picking of thousands of colonies, impeding the efforts to harmonize discoveries that emerge from ‘omics strategies with downstream mechanistic investigations in the rapidly advancing field of microbiome research.…”

“…For instance, Jiang et al, 2016 isolated environmental soil-associated bacteria in droplets and found an increase in the diversity of taxa with an increased representation of rare organisms ( Jiang et al, 2016 ). Villa et al, 2020 recently cultivated human gut microbes in thousands of nanoliter droplets to characterize metabolic variation in polysaccharide-degrading gut bacteria and analyzed their growth kinetics ( Villa et al, 2020 ). Due to small droplet volume, bacteria confined within droplets can reach a critical threshold concentration of quorum sensing molecules faster than would occur in bulk culture, which can lead to improved growth in certain culture medium ( Boedicker et al, 2009 ).…”

Microbiota responses to different prebiotics are conserved within individuals and associated with habitual fiber intake

“…From a microbial ecological perspective, our findings support a model where microbial metabolic repertoires within individuals' microbiota have the capacity to utilize a diverse array of chemically distinct fiber types. Evidence in favor of this model include in vitro prebiotic screens suggesting that individuals uniformly harbor bacteria capable of degrading a wide variety of carbohydrate species (81). Such metabolic plasticity may benefit gut microbiota by enhancing community stability and invasion resistance (82)(83)(84), and could be enabled by substrateswitching at the level of strains (84,85) or cross-feeding interactions (86).…”

Towards culturing the microbe of your choice

“…These microdroplets ensure effective cell separation and can keep them among closely associated organisms on which they may depend. Microdroplet encapsulation has been successful in a variety of environments from aquatic to human gut (Zengler et al ., 2002; Boedicker et al ., 2009; Jiang et al ., 2016; Villa et al ., 2020; Watterson et al ., 2020). Another less conventional approach is the application of electrochemical methods whereby microbes grow on or near electrode surfaces.…”