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Many microbial pathogens produce a β-(1→6)-linked poly- N -acetyl-d-glucosamine (PNAG) surface capsule, including bacterial, fungal, and protozoan cells. Broadly protective immune responses to this single conserved polysaccharide antigen in animals are possible but only when a deacetylated poly- N -acetyl-d-glucosamine (dPNAG; <30% acetate) glycoform is administered as a conjugate to a carrier protein. Unfortunately, conventional methods for natural extraction or chemical synthesis of dPNAG and its subsequent conjugation to protein carriers can be technically demanding and expensive. Here, we describe an alternative strategy for creating broadly protective vaccine candidates that involved coordinating recombinant poly- N -acetyl-d-glucosamine (rPNAG) biosynthesis with outer membrane vesicle (OMV) formation in laboratory strains of Escherichia coli The glycosylated outer membrane vesicles (glycOMVs) released by these engineered bacteria were decorated with the PNAG glycopolymer and induced high titers of PNAG-specific IgG antibodies after immunization in mice. When a Staphylococcus aureus enzyme responsible for PNAG deacetylation was additionally expressed in these cells, glycOMVs were generated that elicited antibodies to both highly acetylated PNAG (∼95-100% acetate) and a chemically deacetylated dPNAG derivative (∼15% acetate). These antibodies mediated efficient in vitro killing of two distinct PNAG-positive bacterial species, namely S. aureus and Francisella tularensis subsp. holarctica , and mice immunized with PNAG-containing glycOMVs developed protective immunity against these unrelated pathogens. Collectively, our results reveal the potential of glycOMVs for targeting this conserved polysaccharide antigen and engendering protective immunity against the broad range of pathogens that produce surface PNAG., Competing Interests: Conflict of interest statement: C.C.-B. is an inventor of intellectual properties (use of human mAb to PNAG and use of PNAG vaccines) that are licensed by Brigham and Women’s Hospital to Alopexx Vaccine, LLC, and Alopexx Pharmaceuticals, LLC. As an inventor of intellectual properties, C.C.-B. also has the right to receive a share of licensing-related income (royalties, fees) through Brigham and Women’s Hospital from Alopexx Pharmaceuticals, LLC, and Alopexx Vaccine, LLC. D.P. and M.P.D. have a financial interest in Versatope, Inc., and M.P.D. also has a financial interest in Glycobia, Inc. The interests of D.P. and M.P.D. are reviewed and managed by Cornell University in accordance with their conflict of interest policies. G.B.P. is an inventor of intellectual properties (human mAb to PNAG and PNAG vaccines) that are licensed by Brigham and Women’s Hospital to Alopexx Vaccine, LLC, and Alopexx Pharmaceuticals, LLC, entities, in which G.B.P. also holds equity. As an inventor of intellectual properties, G.B.P. also has the right to receive a share of licensing-related income (royalties, fees) through Brigham and Women’s Hospital from Alopexx Pharmaceuticals, LLC, and Alopexx Vaccine, LLC. The interests of G.B.P. are reviewed and managed by Brigham and Women’s Hospital and Partners Healthcare in accordance with their conflict of interest policies.

Vaccines often require adjuvants to be effective. Traditional adjuvants, like alum, activate the immune response but in an uncontrolled way. Newer adjuvants help to direct the immune response in a more coordinated fashion. Here, we review the opportunity to use the outer membrane vesicles (OMVs) of bacteria as a way to modulate the immune response toward making more effective vaccines. This review outlines the different types of OMVs that have been investigated for vaccine delivery and how they are produced. Because OMVs are derived from bacteria, they have compositions that may not be compatible with parenteral delivery in humans; therefore, we also review the strategies brought to bear to detoxify OMVs while maintaining an adjuvant profile. OMV-based vaccines can be derived from the pathogens themselves, or can be used as surrogate constructs to mimic a pathogen through the heterologous expression of specific antigens in a desired host source strain, and approaches to doing so are reviewed. Additionally, the emerging area of engineered pathogen-specific carbohydrate sequences, or glycosylated OMVs is reviewed and contrasted with protein antigen delivery. Existing OMV-based vaccines as well as their routes of administration round out the text. Overall, this is an exciting time in the OMV field as it matures and leads to more effective and targeted ways to induce desired pathogen-specific immune responses., (Copyright © 2017 Elsevier B.V. All rights reserved.)