Motile and Chemotactic Minicells and Minicell-Driven Biohybrids Engineered for Active Cargo Delivery

Abstract

Bacterial minicells are submicrometer-sized spherical compartments produced by bacteria as a result of aberrant cell division. Minicells have a similar cellular composition to the parental bacteria but lack chromosomal DNA and are thus unable to proliferate. Due to that, minicells have attracted attention as potential means of effector delivery in bioengineering and biomedical applications. However, until now, the efficiency of delivery by minicells has been limited by passive collisions with their targets. To develop minicell-based active delivery, here we engineer Escherichia coli strains generating motile minicells with enhanced swimming properties by introducing genetic modifications specifically targeting flagella number, length, and rotation speed. The engineered minicells preserve motility over an extended period of time and, in contrast to parental E. coli cells, increase their swimming speed for the intermediate viscosity of the medium. Despite their small size, minicells show an efficient chemotactic response and utilize the same chemotactic strategy as parental E. coli cells. Moreover, we develop a procedure for conjugating minicells with cargo particles and demonstrate that such minicell-driven biohybrid swimmers are chemotactic and thus capable of actively accumulating at the source of an attractant. These engineered chemotactic minicells and minicell-based biohybrids can serve as cargo delivery platforms with active targeting, thus overcoming the challenges posed by nontargeted therapies.