Enriching the unculturable microbial majority of the human gastrointestinal tract through microfluidic encapsulation

Abstract

The human gut microbiota is a consortium of microorganisms made up primarily of bacteria, fungi, and archaea. The complex community of bacteria, in particular, is highly investigated due to its link to human health and disease. Unfortunately, studies aimed at understanding this convoluted relationship are missing key information as a large portion of this commensal population has not been cultured in vitro. One of the dominating theories for the lack of cultured strains is due to the inadequacies of imitating native conditions in vitro that are required for bacterial growth. Due to the growing awareness of gut microbiota’s importance for human health, there is an established need for improved culture techniques. Notably, cell encapsulation is a promising approach to improve culture outputs due to the mechanostimulatory effects and spatial isolation of cells. Microfluidic droplet generation is a preferable method of cell encapsulation due to the high control of microbead formation and biomaterial compatibility. In this thesis, the microfluidic encapsulation of anaerobic gut bacteria is explored to improve in vitro culture. The microfluidic droplet generation was characterized and optimized for compatibility with a synthetic biomaterial, namely four-arm poly(ethylene glycol maleimide)(PEG4MAL). The suitability of PEG4MAL for anaerobic bacteria culture was characterized by assessing mechanical properties, microstructure, and biocompatibility. Lastly, the effect of encapsulation was evaluated by examining cell viability and colony formation. The application of PEG4MAL microencapsulation for the enrichment of difficult-to-grow anaerobic gut bacteria has led to high-density growth within the microbeads. This work provides the foundation for future studies exploring the possibility of in situ culture, sampling, and delivery within the human gastrointestinal tract.

Date	15 May 2025
Original language	American English
Awarding Institution	École de technologie supérieure
Supervisor	Ali Ahmadi (Supervisor), Sophie Lerouge (Co-supervisor) & Corinne F. Maurice (Co-supervisor)