Phononic Crystal-Based Acoustofluidic Device for Sorting Gas Vesicle-Containing Bacteria

Biosynthetic gas vesicles (GVs) have garnered substantial attention owing to their potential as novel ultrasound contrast agents. Compared to traditional microbubble contrast agents, GVs have numerous evident advantages, including nanoscale size, good stability, low cost, and biocompatibility. More importantly, GVs can function as acoustic reporter genes for cell imaging owing to their gene-encoding features. Numerous new versions of GVs have been developed by heterologously expressing GV-encoding gene variants in recombinant Escherichia coli . However, only a few genetically engineered bacteria can successfully produce GVs, which limits their biomedical applications. Therefore, a precise, simple, and rapid method for sorting GV-containing bacteria must be developed. Here, we propose a phononic crystal-based sorting method and fabricate a fully packaged acoustofluidic device that can separate GV-containing E. coli ( GVs-E. coli ) from non-GV-containing E. coli based on the differences in their acoustic contrast factor and in the radiation forces produced by the localized sound field of an integrated resonant phononic crystal plate. The isolated GVs-E. coli demonstrated a purity exceeding 90%, with 89% purification efficiency. This study provides a powerful method for screening GV-containing cells and a perspective for the biomedical applications of acoustic metamaterials.