Abdelrahman, M. K. | bioRxiv Science

bioRxiv2026

Engineered living materials suppress uropathogenic E. coli growth and invasion of urothelial cells through sustained probiotic release

Recurrent urinary tract infection (rUTI) is a significant public health problem. The most common cause of rUTI is uropathogenic Escherichia coli (UPEC). Antimicrobial prophylactic and therapeutic regimens for rUTI disrupts the microbiome and leads to infection with antimicrobial resistant organisms. Therefore, there is an urgent need to develop microbiome-sparing alternative approaches for preventing rUTI. Asymptomatic bacteriuria (ABU) E. coli strain 83972 (ABEC) outcompetes UPEC in the urinary tract without causing UTI symptoms, but its limited persistence in the bladder restricts its efficacy. Here, we investigate a device made from an engineered living material (ELM) that releases ABEC in a sustained manner as an antibiotic-free platform against UTIs and rUTIs. Using physiologically relevant in vitro models incorporating human urothelial cells, human urine, and periodic urine exchange, we show that ABEC-releasing ELMs suppress UPEC proliferation and inhibit UPEC attachment to and invasion into urothelial cells, particularly when ABEC is maintained at equal or higher levels than UPEC. Because ELMs continuously release ABEC, they sustain competitive pressure even as planktonic bacteria are cleared during voiding, outperforming a single dose of free-floating ABEC. In a rUTI model, sustained ABEC release from ELMs reduces the proliferation and re-invasion of UPEC expelled from infected urothelial cells, while UPEC infiltration into fractured ELMs remains negligible. Finally, we design a first-generation ELM device that can be transurethrally delivered and retained within the mouse urinary bladder, achieving sustained ABEC release in vivo, with ABEC persisting in the bladder, kidneys, and urine for at least 4 days. In summary, we report the development of an ELM device that continuously releases ABEC to suppress UPEC proliferation and urothelial cell invasion in in vitro models of rUTI, and demonstrate sustained ABEC release in vivo in a mouse bladder.

bioengineering↗

bioRxiv2024

Controlled release of microorganisms from engineered living materials

Probiotics offer therapeutic benefits by modulating the local microbiome, the host immune response, and the proliferation of pathogens. Probiotics have the potential to treat complex diseases, but their persistence or colonization is required at the target site for effective treatment. Although probiotic persistence can be achieved by repeated delivery, no biomaterial that releases clinically relevant doses of metabolically active probiotics in a sustained manner has been previously described. Here, we encapsulate stiff probiotic microorganisms within relatively less stiff hydrogels and show a generic mechanism where these microorganisms proliferate and induce hydrogel fracture, resulting in microbial release. Importantly, this fracture-based mechanism leads to microorganism release with zero-order release kinetics. Using this mechanism, small ([~]1 L) engineered living materials (ELMs) release >108 colony-forming-units (CFUs) of E. coli in 2 h. This release is sustained for at least 10 days. Cell release can be varied by more than three orders of magnitude by varying initial cell loading and modulating the mechanical properties of encapsulating matrix. As the governing mechanism of microbial release is entirely mechanical, we demonstrate controlled release of model Gram-negative, Gram-positive, and fungal probiotics from multiple hydrogel matrices. SIGNIFICANCEProbiotics offer therapeutic benefits and have the potential to treat complex diseases, but their persistence at the target site is often required for effective treatment. Although probiotic persistence can be achieved by repeated delivery, no biomaterial that releases metabolically active probiotics in a sustained manner has been developed yet. This work demonstrates a generic mechanism where stiff probiotics encapsulated within relatively less stiff hydrogels proliferate and induce hydrogel fracture. This allows a zero-order release of probiotics which can be easily controlled by adjusting the properties of the encapsulating matrices. This generic mechanism is applicable for a wide range of probiotics with different synthetic matrices and has the potential to be used in the treatment of a broad range of diseases.

bioengineering↗