Emergent Biological Endurance Depends on Extracellular Matrix Composition of Three-Dimensionally Printed Escherichia coli Biofilms

Abstract

Biofilms are three-dimensional (3D) bacterial communities that exhibit a highly self-organized nature in terms of their composition and complex architecture. Bacteria in biofilms display emergent biological properties, such as resistance to antimicrobials and disinfectants that the individual planktonic cells lack. Bacterial biofilms possess specialized architectural features including unique extracellular matrix compositions and a distinct spatially patterned arrangement of cells and matrix components within the biofilm. It is unclear which of these architectural elements of bacterial biofilms lead to the development of their emergent biological properties. Here, we report a 3D printing-based technique for studying the emergent resistance behaviors of Escherichia coli biofilms as a function of their architecture. Cellulose and curli are the major extracellular-matrix components in E. coli biofilms. We show that 3D-printed biofilms expressing either curli alone or both curli and cellulose in their extracellular matrices show higher resistance to exposure against disinfectants than 3D prints expressing either cellulose alone or no biofilm-matrix components. The 3D-printed biofilms expressing cellulose and/or curli also show thicker anaerobic zones than nonbiofilm-forming E. coli 3D prints. Thus, the matrix composition plays a crucial role in the emergent spatial patterning and biological endurance of 3D-printed biofilms. In contrast, initial spatial distribution of bacterial density or curli-producing cells does not have an effect on biofilm resistance phenotypes. Further, these 3D-printed biofilms could be reversibly attached to different surfaces (bacterial cellulose, glass, and polystyrene) and display resistance to physical distortions by retaining their shape and structure. This physical robustness highlights their potential in applications including bioremediation, protective coatings against pathogens on medical devices, or wastewater treatment, among many others. This new understanding of the emergent behavior of bacterial biofilms could aid in the development of novel engineered living materials using synthetic biology and materials science approaches.

Biofilms: Formation, Research Models, Potential Targets, and Methods for Prevention and Treatment

“…For example, a better in vitro biofilm model could be developed by printing bacteria and hydrogels which will be capable of mimicking native biofilms. [ 355 , 356 , 357 , 358 , 359 ]…”

“…For example, a better in vitro biofilm model could be developed by printing bacteria and hydrogels which will be capable of mimicking native biofilms. [355][356][357][358][359] In order to develop more effective approaches for managing biofilms, future studies should aim to identify new targets for disrupting biofilms. Furthermore, the advances in genetic tools and resources (e.g., genome-wide transposon insertion libraries) may provide a high-throughput screening platform to find out new targets among the non-essential genes.…”

Biofabrication and characterization of multispecies electroactive biofilms in stratified paper-based scaffolds

“…24,25 Even the latest advances in 3-D printing require complicated and time-consuming processes for bioink preparation, and it is controver-sial whether the 3-D printed biofilm resembles the actual in vivo biofilm environments. 26,27 Above all, reported work with electrogenic bacterial cells for electroactive biofilm formation is unavailable or quite limited.…”

Design and biofabrication of bacterial living materials with robust and multiplexed biosensing capabilities

“…Extrusion-based bacterial printing using custom-built or commercial bioprinters was demonstrated on wild-type [ [10] , [11] , [12] , [13] , [14] ] and engineered [ [15] , [16] , [17] , [18] , [19] , [20] , [21] ] bacterial species. Stereolithographic and inkjet-based bioprinting of engineered strains were also reported in recent studies, the latter being used to fabricate the sole abiotic component [ 22 , 23 ].…”