Designing Bioprinted Agar-Based Scaffolds for Investigating Bacterial Migration 

• Miles Meadows, Chemical Engineering, University of Delaware

• Victoria Muir, Chemical and Biomolecular Engineering, University of Delaware

Designing Bioprinted Agar-Based Scaffolds for Investigating Bacterial Migration 

Miles Meadows, Lawrence Azzariti, Hannah Zucco, Ruth Pereira, Victoria Muir, PhD 

Every day all around us bacteria are constantly on the move. Many bacteria migrate through collective chemotaxis, coordinating movement as a group towards favorable nutrients, signals, and even other organisms. In environments such as our gut and soil, bacteria collectives encounter obstacles and navigate through pores. These biophysical complexities are not replicated in typical laboratory liquid culture experiments. Herein, we develop microporous agar-based hydrogels to study collective bacterial migration in porous environments. To fabricate microporous hydrogels, we use mechanical fragmentation by extrusion to generate agar-based microgels with an average size of ~100 µm for low (1.5 wt.%) and ~200 µm for high (3.0 wt.%) concentration agar bulk hydrogels containing 2 wt.% Luria-Bertani (LB) media. We then assemble these microgels into a jammed state using either low (centrifugation) or high (vacuum filtration) degrees of packing, which results in an agar-based granular hydrogel. This simple, low-cost, and accessible approach allows for the formation of modular microporous environments that are compatible with common microbial culture techniques. We characterize the rheological properties of these agar-based granular hydrogels, which have a storage modulus (G’) of 4.3 ± 0.3 kPa and 16.4 ± 0.4 kPa for low and high agar-based granular hydrogels, respectively, when jammed via vacuum filtration. Within the agar-based granular hydrogels, the microgel phase acts as a physical barrier, much like a grain of soil. Bacterial collectives can move within the microporous void space between the microgels, much like the porous pathways encountered in real-world environments like soil and tissue. Using extrusion printing, we create 1 cm long pathways of agar-based granular hydrogel, and, utilizing time-lapse fluorescence microscopy, we demonstrate that Escherichia coli (E. coli) will collectively perform chemotaxis through the medium. In our ongoing work, we continue exploring E. coli chemotaxis by designing printed pathways of larger scale and complexity. Overall, we present agar-based granular hydrogels as a simple and effective model to replicate the porous environments bacteria encounter every day in an effort to better our understanding of bacterial collective migration.