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Vaccination has become an increasingly attractive strategy for protecting against antibiotic-resistant infections. Nanovaccines based on the outer membrane from Gram-negative bacteria are appealing due to their multiantigenic nature and inherent immunogenicity. Here, we develop cellular nanodiscs made of bacterial outer membrane (OM-NDs), as a platform for antibacterial vaccination. Using Pseudomonas aeruginosa as a model pathogen, the resulting OM-NDs can effectively interact with antigen-presenting cells, exhibiting accelerated uptake and an improved capacity for immune stimulation. With their small size, the OM-NDs are also capable of efficiently transporting to the lymph nodes after in vivo administration. As a result, the nanovaccine is effective at eliciting potent humoral and cellular immune responses against P. aeruginosa. In a murine model of pneumonia, immunization with OM-NDs confers strong protection against subsequent lung infection, resulting in improved survival, reduced bacterial loads, and alleviation of immune overactivation. Overall, this report illustrates the advantages of cellular nanodiscs, which can be readily generalized to other pathogens and may be applied toward other biomedical applications.

Objective: Porphyromonas gingivalis (Pg) is thought to be involved in the progression of Alzheimer's disease (AD). Whether Pg or its contents can reach the brain and directly affect neuropathology is, however, unknown. Here, we investigated whether outer membrane vesicles (OMVs) of Pg translocate to the brain and induce the pathogenic features of AD. Material and Methods: Pg OMVs were injected into the abdominal cavity of mice for 12 weeks. Pg OMV translocation to the brain was detected by immunohistochemistry using an anti-gingipain antibody. Tau protein and microglial activation in the mouse brain were examined by western blotting and immunohistochemistry. The effect of gingipains on inflammation was assessed by real-time polymerase chain reaction using human microglial HMC3 cells. Results: Gingipains were detected in the region around cerebral ventricles, choroid plexus, and ventricular ependymal cells in Pg OMV-administered mice. Tau and phosphorylated Tau protein increased and microglia were activated. Pg OMVs also increased the gene expression of proinflammatory cytokines in HMC3 cells in a gingipain-dependent manner. Conclusion: Pg OMVs, including gingipains, can reach the cerebral ventricle and induce neuroinflammation by activating microglia. Pg OMVs may provide a better understanding of the implications of periodontal diseases in neurodegenerative conditions such as AD.

Flocculation is an aggregation phenomenon of microbial cells in which they form flocs or flakes. In this study, it was found that addition of glycerol to a complex glucose medium promoted spontaneous floc formation by an Escherichia coli degP-deficient mutant strain …

Flocculation is an aggregation phenomenon of microbial cells in which they form flocs or flakes. In this study, it was found that addition of glycerol to a complex glucose medium promoted spontaneous floc formation by an Escherichia coli degP-deficient mutant strain …

Outer membrane vesicles (OMVs) are extracellular vesicles released from the surface of Gram-negative bacteria, including Escherichia coli. Several gene-deficient mutants relating to envelope stress (nlpI and degP) and phospholipid accumulation in the outer leaflet of the outer membrane (mlaA and mlaE) increase OMV production. This study examined the combinatorial deletion of these genes in E. coli and its effect on OMV production....

Outer membrane vesicles (OMVs) are extracellular vesicles released from the surface of Gram-negative bacteria, including Escherichia coli. Several gene-deficient mutants relating to envelope stress (nlpI and degP) and phospholipid accumulation in the outer leaflet of the outer membrane (mlaA and mlaE) increase OMV production. This study examined the combinatorial deletion of these genes in E. coli and its effect on OMV production....

The Escherichia coli virulence factor hemoglobin protease (Hbp) has been engineered into a surface display system that can be expressed to high density on live E. coli and Salmonella enterica serovar Typhimurium cells or derived outer membrane vesicles (OMVs). Multiple antigenic sequences can be genetically fused into the Hbp core structure for optimal exposure to the immune system. Although the Hbp display platform is relatively tolerant, increasing the number, size, and complexity of integrated sequences generally lowers the expression of the fused constructs and limits the density of display. This is due to the intricate mechanism of Hbp secretion across the outer membrane and the efficient quality control of translocation-incompetent chimeric Hbp molecules in the periplasm. To address this shortcoming, we explored the coupling of purified proteins to the Hbp carrier after its translocation across the outer membrane using the recently developed SpyTag/ SpyCatcher protein ligation system. As expected, fusion of the small SpyTag to Hbp did not hamper display on OMVs. Subsequent addition of purified proteins fused to the SpyCatcher domain resulted in efficient covalent coupling to Hbp-SpyTag. Using in addition the orthogonal SnoopTag/SnoopCatcher system, multiple antigen modules could be coupled to Hbp in a sequential ligation strategy. Not only antigens proved suitable for Spy-mediated ligation but also nanobodies. Addition of this functionality to the platform might allow the targeting of live bacterial or OMV vaccines to certain tissues or immune cells to tailor immune responses.

The Gram-negative bacterium Aggregatibacter actinomycetemcomitans is an oral and systemic pathogen, which is linked to aggressive forms of periodontitis and can be associated with endocarditis. The outer membrane vesicles (OMVs) of this species contain effector proteins such as cytolethal distending toxin (CDT) and leukotoxin (LtxA), which they can deliver into human host cells. The OMVs can also activate innate immunity through NOD1- and NOD2-active pathogen-associated molecular patterns. This dataset provides a proteome of highly purified OMVs from A. actinomycetemcomitans serotype e strain 173. The experimental data do not only include the raw data of the LC-MS/MS analysis of four independent preparations of purified OMVs but also the mass lists of the processed data and the Mascot.dat files from the database searches. In total 501 proteins are identified, of which 151 are detected in at least three of four independent preparations. In addition, this dataset contains the COG definitions and the predicted subcellular locations (PSORTb 3.0) for the entire genome of A. actinomycetemcomitans serotype e strain SC1083, which is used for the evaluation of the LC-MS/MS data. These data are deposited in ProteomeXchange in the public dataset PXD002509. In addition, a scientific interpretation of this dataset by Kieselbach et al. (2015) [2] is available at http://dx.doi.org/10.1371/journal.pone.0138591.

The Gram-negative bacterium Aggregatibacter actinomycetemcomitans is an oral and systemic pathogen, which is linked to aggressive forms of periodontitis and can be associated with endocarditis. The outer membrane vesicles (OMVs) of this species contain effector proteins such as cytolethal distending toxin (CDT) and leukotoxin (LtxA), which they can deliver into human host cells. The OMVs can also activate innate immunity through NOD1- and NOD2-active pathogen-associated molecular patterns. This dataset provides a proteome of highly purified OMVs from A. actinomycetemcomitans serotype e strain 173. The experimental data do not only include the raw data of the LC-MS/MS analysis of four independent preparations of purified OMVs but also the mass lists of the processed data and the Mascot.dat files from the database searches. In total 501 proteins are identified, of which 151 are detected in at least three of four independent preparations. In addition, this dataset contains the COG definitions and the predicted subcellular locations (PSORTb 3.0) for the entire genome of A. actinomycetemcomitans serotype e strain SC1083, which is used for the evaluation of the LC-MS/MS data. These data are deposited in ProteomeXchange in the public dataset PXD002509. In addition, a scientific interpretation of this dataset by Kieselbach et al. (2015) [2] is available at http://dx.doi.org/10.1371/journal.pone.0138591.

The Gram-negative bacterium Aggregatibacter actinomycetemcomitans is an oral and systemic pathogen, which is linked to aggressive forms of periodontitis and can be associated with endocarditis. The outer membrane vesicles (OMVs) of this species contain effector proteins such as cytolethal distending toxin (CDT) and leukotoxin (LtxA), which they can deliver into human host cells. The OMVs can also activate innate immunity through NOD1- and NOD2-active pathogen-associated molecular patterns. This dataset provides a proteome of highly purified OMVs from A. actinomycetemcomitans serotype e strain 173. The experimental data do not only include the raw data of the LC-MS/MS analysis of four independent preparations of purified OMVs but also the mass lists of the processed data and the Mascot.dat files from the database searches. In total 501 proteins are identified, of which 151 are detected in at least three of four independent preparations. In addition, this dataset contains the COG definitions and the predicted subcellular locations (PSORTb 3.0) for the entire genome of A. actinomycetemcomitans serotype e strain SC1083, which is used for the evaluation of the LC-MS/MS data. These data are deposited in ProteomeXchange in the public dataset PXD002509. In addition, a scientific interpretation of this dataset by Kieselbach et al. (2015) [2] is available at http://dx.doi.org/10.1371/journal.pone.0138591.

Shewanella oneidensis is a Gram-negative facultative anaerobe that can use a wide variety of terminal electron acceptors for anaerobic respiration. In this study, S. oneidensis degQ gene, encoding a putative periplasmic serine protease, was cloned and expressed. The activity of purified DegQ was inhibited by diisopropyl fluorophosphate, a typical serine protease-specific inhibitor, indicating that DegQ is a serine protease....

Shewanella oneidensis is a Gram-negative facultative anaerobe that can use a wide variety of terminal electron acceptors for anaerobic respiration. In this study, S. oneidensis degQ gene, encoding a putative periplasmic serine protease, was cloned and expressed. The activity of purified DegQ was inhibited by diisopropyl fluorophosphate, a typical serine protease-specific inhibitor, indicating that DegQ is a serine protease....

The Gram-negative bacterium Aggregatibacter actinomycetemcomitans is an oral and systemic pathogen, which is linked to aggressive forms of periodontitis and can be associated with endocarditis. The outer membrane vesicles (OMVs) of this species contain effector proteins such as cytolethal distending toxin (CDT) and leukotoxin (LtxA), which they can deliver into human host cells. The OMVs can also activate innate immunity through NOD1- and NOD2-active pathogen-associated molecular patterns. This dataset provides a proteome of highly purified OMVs from A. actinomycetemcomitans serotype e strain 173. The experimental data do not only include the raw data of the LC-MS/MS analysis of four independent preparations of purified OMVs but also the mass lists of the processed data and the Mascot.dat files from the database searches. In total 501 proteins are identified, of which 151 are detected in at least three of four independent preparations. In addition, this dataset contains the COG definitions and the predicted subcellular locations (PSORTb 3.0) for the entire genome of A. actinomycetemcomitans serotype e strain SC1083, which is used for the evaluation of the LC-MS/MS data. These data are deposited in ProteomeXchange in the public dataset PXD002509. In addition, a scientific interpretation of this dataset by Kieselbach et al. (2015) [2] is available at http://dx.doi.org/10.1371/journal.pone.0138591.

The Gram-negative bacterium Aggregatibacter actinomycetemcomitans is an oral and systemic pathogen, which is linked to aggressive forms of periodontitis and can be associated with endocarditis. The outer membrane vesicles (OMVs) of this species contain effector proteins such as cytolethal distending toxin (CDT) and leukotoxin (LtxA), which they can deliver into human host cells. The OMVs can also activate innate immunity through NOD1- and NOD2-active pathogen-associated molecular patterns. This dataset provides a proteome of highly purified OMVs from A. actinomycetemcomitans serotype e strain 173. The experimental data do not only include the raw data of the LC-MS/MS analysis of four independent preparations of purified OMVs but also the mass lists of the processed data and the Mascot.dat files from the database searches. In total 501 proteins are identified, of which 151 are detected in at least three of four independent preparations. In addition, this dataset contains the COG definitions and the predicted subcellular locations (PSORTb 3.0) for the entire genome of A. actinomycetemcomitans serotype e strain SC1083, which is used for the evaluation of the LC-MS/MS data. These data are deposited in ProteomeXchange in the public dataset PXD002509. In addition, a scientific interpretation of this dataset by Kieselbach et al. (2015) [2] is available at http://dx.doi.org/10.1371/journal.pone.0138591.

Vibrios are associated with a broad diversity of hosts that produce antimicrobial peptides (AMPs) as part of their defense against microbial infections. In particular, vibrios colonize epithelia, which function as protective barriers and express AMPs as a first line of chemical defense against pathogens. Recent studies have shown they can also colonize phagocytes, key components of the animal immune system. Phagocytes infiltrate infected tissues and use AMPs to kill the phagocytosed microorganisms intracellularly, or deliver their antimicrobial content extracellularly to circumvent tissue infection. We review here the mechanisms by which vibrios have evolved the capacity to evade or resist the potent antimicrobial defenses of the immune cells or tissues they colonize. Among their strategies to resist killing by AMPs, primarily vibrios use membrane remodeling mechanisms. In particular, some highly resistant strains substitute hexaacylated Lipid A with a diglycine residue to reduce their negative surface charge, thereby lowering their electrostatic interactions with cationic AMPs. As a response to envelope stress, which can be induced by membrane-active agents including AMPs, vibrios also release outer membrane vesicles to create a protective membranous shield that traps extracellular AMPs and prevents interaction of the peptides with their own membranes. Finally, once AMPs have breached the bacterial membrane barriers, vibrios use RND efflux pumps, similar to those of other species, to transport AMPs out of their cytoplasmic space.

Vibrios are associated with a broad diversity of hosts that produce antimicrobial peptides (AMPs) as part of their defense against microbial infections. In particular, vibrios colonize epithelia, which function as protective barriers and express AMPs as a first line of chemical defense against pathogens. Recent studies have shown they can also colonize phagocytes, key components of the animal immune system. Phagocytes infiltrate infected tissues and use AMPs to kill the phagocytosed microorganisms intracellularly, or deliver their antimicrobial content extracellularly to circumvent tissue infection. We review here the mechanisms by which vibrios have evolved the capacity to evade or resist the potent antimicrobial defenses of the immune cells or tissues they colonize. Among their strategies to resist killing by AMPs, primarily vibrios use membrane remodeling mechanisms. In particular, some highly resistant strains substitute hexaacylated Lipid A with a diglycine residue to reduce their negative surface charge, thereby lowering their electrostatic interactions with cationic AMPs. As a response to envelope stress, which can be induced by membrane-active agents including AMPs, vibrios also release outer membrane vesicles to create a protective membranous shield that traps extracellular AMPs and prevents interaction of the peptides with their own membranes. Finally, once AMPs have breached the bacterial membrane barriers, vibrios use RND efflux pumps, similar to those of other species, to transport AMPs out of their cytoplasmic space.

Vibrios are associated with a broad diversity of hosts that produce antimicrobial peptides (AMPs) as part of their defense against microbial infections. In particular, vibrios colonize epithelia, which function as protective barriers and express AMPs as a first line of chemical defense against pathogens. Recent studies have shown they can also colonize phagocytes, key components of the animal immune system. Phagocytes infiltrate infected tissues and use AMPs to kill the phagocytosed microorganisms intracellularly, or deliver their antimicrobial content extracellularly to circumvent tissue infection. We review here the mechanisms by which vibrios have evolved the capacity to evade or resist the potent antimicrobial defenses of the immune cells or tissues they colonize. Among their strategies to resist killing by AMPs, primarily vibrios use membrane remodeling mechanisms. In particular, some highly resistant strains substitute hexaacylated Lipid A with a diglycine residue to reduce their negative surface charge, thereby lowering their electrostatic interactions with cationic AMPs. As a response to envelope stress, which can be induced by membrane-active agents including AMPs, vibrios also release outer membrane vesicles to create a protective membranous shield that traps extracellular AMPs and prevents interaction of the peptides with their own membranes. Finally, once AMPs have breached the bacterial membrane barriers, vibrios use RND efflux pumps, similar to those of other species, to transport AMPs out of their cytoplasmic space.

Vibrios are associated with a broad diversity of hosts that produce antimicrobial peptides (AMPs) as part of their defense against microbial infections. In particular, vibrios colonize epithelia, which function as protective barriers and express AMPs as a first line of chemical defense against pathogens. Recent studies have shown they can also colonize phagocytes, key components of the animal immune system. Phagocytes infiltrate infected tissues and use AMPs to kill the phagocytosed microorganisms intracellularly, or deliver their antimicrobial content extracellularly to circumvent tissue infection. We review here the mechanisms by which vibrios have evolved the capacity to evade or resist the potent antimicrobial defenses of the immune cells or tissues they colonize. Among their strategies to resist killing by AMPs, primarily vibrios use membrane remodeling mechanisms. In particular, some highly resistant strains substitute hexaacylated Lipid A with a diglycine residue to reduce their negative surface charge, thereby lowering their electrostatic interactions with cationic AMPs. As a response to envelope stress, which can be induced by membrane-active agents including AMPs, vibrios also release outer membrane vesicles to create a protective membranous shield that traps extracellular AMPs and prevents interaction of the peptides with their own membranes. Finally, once AMPs have breached the bacterial membrane barriers, vibrios use RND efflux pumps, similar to those of other species, to transport AMPs out of their cytoplasmic space.

Vibrios are associated with a broad diversity of hosts that produce antimicrobial peptides (AMPs) as part of their defense against microbial infections. In particular, vibrios colonize epithelia, which function as protective barriers and express AMPs as a first line of chemical defense against pathogens. Recent studies have shown they can also colonize phagocytes, key components of the animal immune system. Phagocytes infiltrate infected tissues and use AMPs to kill the phagocytosed microorganisms intracellularly, or deliver their antimicrobial content extracellularly to circumvent tissue infection. We review here the mechanisms by which vibrios have evolved the capacity to evade or resist the potent antimicrobial defenses of the immune cells or tissues they colonize. Among their strategies to resist killing by AMPs, primarily vibrios use membrane remodeling mechanisms. In particular, some highly resistant strains substitute hexaacylated Lipid A with a diglycine residue to reduce their negative surface charge, thereby lowering their electrostatic interactions with cationic AMPs. As a response to envelope stress, which can be induced by membrane-active agents including AMPs, vibrios also release outer membrane vesicles to create a protective membranous shield that traps extracellular AMPs and prevents interaction of the peptides with their own membranes. Finally, once AMPs have breached the bacterial membrane barriers, vibrios use RND efflux pumps, similar to those of other species, to transport AMPs out of their cytoplasmic space.

Vibrios are associated with a broad diversity of hosts that produce antimicrobial peptides (AMPs) as part of their defense against microbial infections. In particular, vibrios colonize epithelia, which function as protective barriers and express AMPs as a first line of chemical defense against pathogens. Recent studies have shown they can also colonize phagocytes, key components of the animal immune system. Phagocytes infiltrate infected tissues and use AMPs to kill the phagocytosed microorganisms intracellularly, or deliver their antimicrobial content extracellularly to circumvent tissue infection. We review here the mechanisms by which vibrios have evolved the capacity to evade or resist the potent antimicrobial defenses of the immune cells or tissues they colonize. Among their strategies to resist killing by AMPs, primarily vibrios use membrane remodeling mechanisms. In particular, some highly resistant strains substitute hexaacylated Lipid A with a diglycine residue to reduce their negative surface charge, thereby lowering their electrostatic interactions with cationic AMPs. As a response to envelope stress, which can be induced by membrane-active agents including AMPs, vibrios also release outer membrane vesicles to create a protective membranous shield that traps extracellular AMPs and prevents interaction of the peptides with their own membranes. Finally, once AMPs have breached the bacterial membrane barriers, vibrios use RND efflux pumps, similar to those of other species, to transport AMPs out of their cytoplasmic space.

Vibrios are associated with a broad diversity of hosts that produce antimicrobial peptides (AMPs) as part of their defense against microbial infections. In particular, vibrios colonize epithelia, which function as protective barriers and express AMPs as a first line of chemical defense against pathogens. Recent studies have shown they can also colonize phagocytes, key components of the animal immune system. Phagocytes infiltrate infected tissues and use AMPs to kill the phagocytosed microorganisms intracellularly, or deliver their antimicrobial content extracellularly to circumvent tissue infection. We review here the mechanisms by which vibrios have evolved the capacity to evade or resist the potent antimicrobial defenses of the immune cells or tissues they colonize. Among their strategies to resist killing by AMPs, primarily vibrios use membrane remodeling mechanisms. In particular, some highly resistant strains substitute hexaacylated Lipid A with a diglycine residue to reduce their negative surface charge, thereby lowering their electrostatic interactions with cationic AMPs. As a response to envelope stress, which can be induced by membrane-active agents including AMPs, vibrios also release outer membrane vesicles to create a protective membranous shield that traps extracellular AMPs and prevents interaction of the peptides with their own membranes. Finally, once AMPs have breached the bacterial membrane barriers, vibrios use RND efflux pumps, similar to those of other species, to transport AMPs out of their cytoplasmic space.

Vibrios are associated with a broad diversity of hosts that produce antimicrobial peptides (AMPs) as part of their defense against microbial infections. In particular, vibrios colonize epithelia, which function as protective barriers and express AMPs as a first line of chemical defense against pathogens. Recent studies have shown they can also colonize phagocytes, key components of the animal immune system. Phagocytes infiltrate infected tissues and use AMPs to kill the phagocytosed microorganisms intracellularly, or deliver their antimicrobial content extracellularly to circumvent tissue infection. We review here the mechanisms by which vibrios have evolved the capacity to evade or resist the potent antimicrobial defenses of the immune cells or tissues they colonize. Among their strategies to resist killing by AMPs, primarily vibrios use membrane remodeling mechanisms. In particular, some highly resistant strains substitute hexaacylated Lipid A with a diglycine residue to reduce their negative surface charge, thereby lowering their electrostatic interactions with cationic AMPs. As a response to envelope stress, which can be induced by membrane-active agents including AMPs, vibrios also release outer membrane vesicles to create a protective membranous shield that traps extracellular AMPs and prevents interaction of the peptides with their own membranes. Finally, once AMPs have breached the bacterial membrane barriers, vibrios use RND efflux pumps, similar to those of other species, to transport AMPs out of their cytoplasmic space.