Optimizing the production, purification, characterization of E. coli OMVs

Outer membrane vesicles (OMVs) are 20-250 nm particles released from Gram-negative bacteria, including commensal and pathogenic Escherichia coli(E. coli). OMVs contain outer membrane and periplasmic proteins, toxins, nucleic acids, and pieces of peptidoglycan. OMV production is enhanced by environmental stressors, and OMVs are thought to facilitate bacterial cell growth and division, to contribute to bacterial communication (quorum sensing), and can also act as decoys during antibiotic or host immune system attacks. We aim to optimize E. coli OMV production and purification using ultracentrifugation and alternative techniques, to characterize OMVs using immunoblotting and nanoparticle tracking analysis (NTA), and to ultimately determine how OMVs can be used as molecular biomarkers for E. coli infections and Gram-negative sepsis. Preliminary studies have shown that antibodies to peptidoglycan associated lipoprotein (Pal) can be employed to successfully identify E. coli OMVs. © FASEB.