scholarly journals Outer membrane vesicles derived from Salmonella Typhimurium mutants with truncated LPS induce cross-protective immune responses against infection of Salmonella enterica serovars in the mouse model

2016 ◽  
Vol 306 (8) ◽  
pp. 697-706 ◽  
Author(s):  
Qiong Liu ◽  
Qing Liu ◽  
Jie Yi ◽  
Kang Liang ◽  
Tian Liu ◽  
...  
Keyword(s):  
Mouse Model ◽  
Immune Responses ◽  
Salmonella Typhimurium ◽  
Outer Membrane ◽  
Salmonella Enterica ◽  
Membrane Vesicles ◽  
Outer Membrane Vesicles

scholarly journals Display of Recombinant Proteins on Bacterial Outer Membrane Vesicles by Using Protein Ligation

2018 ◽  
Vol 84 (8) ◽  
pp. e02567-17 ◽  
Author(s):  
H. Bart van den Berg van Saparoea ◽  
Diane Houben ◽  
Marien I. de Jonge ◽  
Wouter S. P. Jong ◽  
Joen Luirink
Keyword(s):  
Escherichia Coli ◽  
Outer Membrane ◽  
Salmonella Enterica ◽  
Membrane Vesicles ◽  
Therapeutic Agents ◽  
Outer Membrane Vesicles ◽  
High Density ◽  
Content Type ◽  
Protein Ligation ◽  
Serovar Typhimurium

ABSTRACT 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.IMPORTANCE Outer membrane vesicles (OMVs) derived from Gram-negative bacteria attract increasing interest in the development of vaccines and therapeutic agents. We aim to construct a semisynthetic OMV platform for recombinant antigen presentation on OMVs derived from attenuated Salmonella enterica serovar Typhimurium cells displaying an adapted Escherichia coli autotransporter, Hbp, at the surface. Although this autotransporter accepts substantial modifications, its capacity with respect to the number, size, and structural complexity of the antigens genetically fused to the Hbp carrier is restricted. Here we describe the application of SpyCatcher/SpyTag protein ligation technology to enzymatically link antigens to Hbp present at high density in OMVs. Protein ligation was apparently unobstructed by the membrane environment and allowed a high surface density of coupled antigens, a property we have shown to be important for vaccine efficacy. The OMV coupling procedure appears versatile and robust, allowing fast production of experimental vaccines and therapeutic agents through a modular plug-and-display procedure.

​Optimization of Cultural Conditions for Maximum Production of Outer Membrane Vesicles from Salmonella Typhimurium

2021 ◽  
Author(s):  
Menguzotunuo Solo ◽  
Shantanu Tamuly ◽  
Luit Moni Barkalita ◽  
Girin Kumar Saikia ◽  
Dhruba Jyoti Kalita
Keyword(s):  
Oxidative Stress ◽  
Salmonella Typhimurium ◽  
Outer Membrane ◽  
Large Scale ◽  
Multiple Drug Resistance ◽  
Membrane Vesicles ◽  
Outer Membrane Vesicles ◽  
Multiple Drug ◽  
Scale Production ◽  
Maximum Production

Background: The non-typhoidal Salmonella causes gastroenteritis in humans that makes its way to the food chain mainly through the animal products. The multiple drug resistance imposes one of the major hurdle in the treatment of the disease. The vaccination appears to be the most important method for prevention of the disease. Unfortunately, there is no liscenced vaccine available against non-typhoidal Salmonellae. The use of outer membrane vesicles (OMVs) of Salmonella as a vaccine candidate has attained significant centre-stage in the recent years given to its protective immunogenicity. However, the large scale production of OMVs is difficult owing to low yield per liter of culture. Methods: In the present study, we have optimized the culture conditions viz. pH, phase of growth and presence of oxidative stress for maximum production of OMVs from Salmonella Typhimurium. The OMVs were characterized based on yield based on protein concentration, lipopolysaccharide concentration and zeta size. Result: In the present study, it was found that incubation of Salmonella Typhimurium up to peak of the growth phase at pH 7 in presence of oxidative stress was found to be the most suitable condition for maximum production of OMVs.

scholarly journals Decoration of Outer Membrane Vesicles with Multiple Antigens by Using an Autotransporter Approach

2014 ◽  
Vol 80 (18) ◽  
pp. 5854-5865 ◽  
Author(s):  
Maria H. Daleke-Schermerhorn ◽  
Tristan Felix ◽  
Zora Soprova ◽  
Corinne M. ten Hagen-Jongman ◽  
David Vikström ◽  
...  
Keyword(s):  
Immune Responses ◽  
Outer Membrane ◽  
Vaccine Development ◽  
Membrane Vesicles ◽  
Outer Membrane Vesicles ◽  
Heterologous Proteins ◽  
Content Type ◽  
Serovar Typhimurium ◽  
Heterologous Antigens

ABSTRACTOuter membrane vesicles (OMVs) are spherical nanoparticles that naturally shed from Gram-negative bacteria. They are rich in immunostimulatory proteins and lipopolysaccharide but do not replicate, which increases their safety profile and renders them attractive vaccine vectors. By packaging foreign polypeptides in OMVs, specific immune responses can be raised toward heterologous antigens in the context of an intrinsic adjuvant. Antigens exposed at the vesicle surface have been suggested to elicit protection superior to that from antigens concealed inside OMVs, but hitherto robust methods for targeting heterologous proteins to the OMV surface have been lacking. We have exploited our previously developed hemoglobin protease (Hbp) autotransporter platform for display of heterologous polypeptides at the OMV surface. One, two, or three of theMycobacterium tuberculosisantigens ESAT6, Ag85B, and Rv2660c were targeted to the surface ofEscherichia coliOMVs upon fusion to Hbp. Furthermore, a hypervesiculating ΔtolRΔtolAderivative of attenuatedSalmonella entericaserovar Typhimurium SL3261 was generated, enabling efficient release and purification of OMVs decorated with multiple heterologous antigens, exemplified by theM. tuberculosisantigens and epitopes fromChlamydia trachomatismajor outer membrane protein (MOMP). Also, we showed that delivery ofSalmonellaOMVs displaying Ag85B to antigen-presenting cellsin vitroresults in processing and presentation of an epitope that is functionally recognized by Ag85B-specific T cell hybridomas. In conclusion, the Hbp platform mediates efficient display of (multiple) heterologous antigens, individually or combined within one molecule, at the surface of OMVs. Detection of antigen-specific immune responses upon vesicle-mediated delivery demonstrated the potential of our system for vaccine development.

scholarly journals Immunization with Vibrio cholerae Outer Membrane Vesicles Induces Protective Immunity in Mice

2008 ◽  
Vol 76 (10) ◽  
pp. 4554-4563 ◽  
Author(s):  
Stefan Schild ◽  
Eric J. Nelson ◽  
Andrew Camilli
Keyword(s):  
Immune Responses ◽  
Outer Membrane ◽  
Vibrio Cholerae ◽  
High Titer ◽  
Immunoglobulin A ◽  
Membrane Vesicles ◽  
Oral Route ◽  
Outer Membrane Vesicles ◽  
Intranasal Route ◽  
Female Mice

ABSTRACT The gram-negative bacterium Vibrio cholerae releases outer membrane vesicles (OMVs) during growth. In this study, we immunized female mice by the intranasal, intragastric, or intraperitoneal route with purified OMVs derived from V. cholerae. Independent of the route of immunization, mice induced specific, high-titer immune responses of similar levels against a variety of antigens present in the OMVs. After the last immunization, the half-maximum total immunoglobulin titer was stable over a 3-month period, indicating that the immune response was long lasting. The induction of specific isotypes, however, was dependent on the immunization route. Immunoglobulin A, for example, was induced to a significant level only by mucosal immunization, with the intranasal route generating the highest titers. We challenged the offspring of immunized female mice with V. cholerae via the oral route in two consecutive periods, approximately 30 and 95 days after the last immunization. Regardless of the route of immunization, the offspring was protected against colonization with V. cholerae in both challenge periods. Our results show that mucosal immunizations via both routes with OMVs derived from V. cholerae induce long-term protective immune responses against this gastrointestinal pathogen. These findings may contribute to the development of “nonliving,” OMV-based vaccines against V. cholerae and other enteric pathogens, using the oral or intranasal route of immunization.

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