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Stimuli-responsive hydrogels that provide controlled degradation can be used as bacteria delivery systems for advanced therapeutic applications. Here, we report the first use of photodegradable hydrogels as materials that can direct bacterial movement, tune mean bacteria speed, and control bacteria delivery through spatiotemporal control of degradation. Hydrogels were formed using base-catalyzed Michael addition reactions between photodegradable poly(ethylene glycol) (PEG) o-nitrobenzyl diacrylate macromers and PEG tetra-thiol cross-linkers within microfluidic channels. Nutrient gradients were generated across the channel, and micron-scale regions of the hydrogel were partially degraded by exposure to controlled doses (2.1–168 mJ/mm^2) of patterned 365 nm light. Hydrogel degradation was then characterized in situ using fluorescence visualization of fluorescein-labeled hydrogels. Following characterization, Bacillus subtilis expressing green fluorescent protein was introduced into the device, and its movement up the nutrient gradient was monitored using time-lapse fluorescence microscopy to enable a systematic study of bacteria chemotaxis through the hydrogels at varied levels of degradation. B. subtilis showed minimal adhesion to partially degraded PEG hydrogels, and bacteria mean speed and mean directional change were tunable according to the level of hydrogel photodegradation, with a 2.6-fold difference in mean cell speed measured across the partially degraded hydrogel regions. Finally, the ability to alter bacteria speed and directionality through tunable degradation and without significant adhesion was used to achieve controlled release profiles of bacteria to delivery sites. These findings advance the use of PEG-based hydrogel materials as delivery vehicles for bacterial therapeutic applications and other living material applications that require controlled bacteria transport. 

Abstract Hydrogel materials can be used to integrate bacteria cells into biohybrid systems. Here, we investigate the use of polyethylene glycol-based hydrogels that employ different Michael-type addition crosslinking chemistries, including thiol-acrylate, thiol-vinyl sulfone, and thiol-maleimide click reactions, for covalent hydrogel network formation and bacteria encapsulation. All crosslinking chemistries generated hydrogels that provided stable encapsulation and culture ofBacillus subtilis; however, significant differences in cell viability and cell morphology after encapsulation were identified. Thiol-acrylate hydrogels provided the highest cell viability and favored encapsulation of single cells, while thiol-maleimide hydrogels had the lowest cell viability and favored encapsulation of larger aggregates. These findings demonstrate the impact of crosslinking strategies for encapsulation of microorganisms into hydrogel networks and suggest that thiol-acrylate chemistries are favorable for many applications. Graphical abstract