g Francisella tularensis is a Gram-negative immune-evasive coccobacillus that causes tularemia in humans and animals. A safe and efficacious vaccine that is protective against multiple F. tularensis strains has yet to be developed. In this study, we tested a novel vaccine approach using artificial pathogens, synthetic nanoparticles made from catanionic surfactant vesicles that are functionalized by the incorporation of either
Identification of antigens is important for vaccine production. We tested extraction protocols using cetyltrimethylammonium tosylate (CTAT) and sodium dodecylbenzenesulfonate (SDBS) to formulate surfactant vesicles (SVs) containing components from Neisseria gonorrhoeae. Carbohydrate and protein assays demonstrated that protein and carbohydrates were incorporated into the vesicle leaflet. Depending on the extraction protocol utilized, 100–400 µg of protein/mL of SVs solution was obtained. Gel electrophoresis followed by silver staining demonstrated that SV extracts contained lipooligosaccharide and a subset of bacterial proteins and lipoproteins. Western blotting and mass spectral analysis indicated that the majority of the proteins were derived from the outer membrane. Mass spectrometric and bioinformatics analysis of SVs identified 29 membrane proteins, including porin and opacity-associated protein. Proteins embedded in the SVs leaflet could be degraded by the addition of trypsin or proteinase K. Our data showed that the incorporation of CTAT and SDBS into vesicles eliminated their toxicity as measured by a THP-1 killing assay. Incorporation of gonococcal cell surface components into SVs reduced toxicity as compared to the whole cell extracts, as measured by cytokine induction, while retaining the immunogenicity. This process constitutes a general method for extracting bacterial surface components and identification of antigens that might be included in vaccines.
Francisella tularensis (Ft) is a Gram-negative, immune-evasive coccobacillus that causes tularemia for which there is no FDA-approved vaccine. We have utilized a novel vaccine approach using synthetic nanoparticles made from catanionic surfactant vesicles (V), functionalized by incorporation of either Ft type B Live Vaccine Strain (Ft LVS) or Ft type A Schu S4 strain (Ft Schu S4) components (i.e., LVS-V and Schu S4-V, respectively). Immunization of C57BL/6 mice with bare V partially protected against Ft LVS, presumably through activation of an innate immune response, yet failed to protect against Ft Schu S4. In contrast, immunization with LVS-V fully protected mice against intraperitoneal (i.p.) Ft LVS challenge, while immunization of mice with either LVS-V or Schu S4-V partially protected C57BL/6 mice against an intranasal (i.n.) Ft Schu S4 challenge. LVS-V-immunization, but not immunization with V, elicited high levels of IgG against non-LPS epitopes and these antisera conferred passive protection against challenge with Ft LVS. Our recently published and ongoing studies aim to identify the protein targets of mouse antisera, study the mechanism of non-specific protection gained by immunization with this nanoparticle vaccine platform (adjuvant effect) in Ft LVS challenge, and enhance protection in Ft Schu S4 challenge. Our data extend the utility of functionalized catanionic surfactant vesicles as a vaccine platform for pathogens.
ΔlgtD (a strain that produces lacto-N-neotetraose LOS and a peptide (PADRE) that possesses the ability to bind to a large number of HLA class II molecules, inserted into a surfactant vesicle. TRIAD is a catanionic surfactant vesicle formulation and the resulting vesicle is stable at room temperature for years, unlike a typical liposome. TRIAD is so robust that it can be autoclaved without any appreciable loss of structural integrity. Results Using TRIAD that contained LOS and PADRE at a ratio of 10:1, and immunising with 2 μg of LOS equivalent, we were able to demonstrate that our vaccine induces a high titer anti-LOS antibody response, with the majority of the elicited antibody being IgG. Intraperitoneal immunisation of mice with our vaccine construct produced no observable adverse effects in mice, while intraperitoneal immunisation with equivalent amounts of purified LOS induced significant adverse effects. Conclusions Our surfactant vesicle platform possesses all of the advantages seen with traditional liposome formulations, without any of the inherent problems associated with liposome-mediated vaccines. This vaccine platform readily lends itself to further modifications in that it is possible to include additional neisserial proteins into the vaccine via a novel whole cell extraction protocol. We believe that this will allow us to generate a universal vaccine able to protect against all serotypes of N. meningitidis. Background The HIV epidemic has shown a considerable amount of heterogeneity, complicating the design and placement of prevention, intervention and treatment programmes. Place-based analyses providing specific information on pathogen prevalence, risk behaviours and other micro-level behaviours can help to target public health responses. Methods Data were from a cross-sectional survey of most-at-risk populations (MARPs) from Winnipeg, Canada. Respondents were recruited through respondent-driven sampling, and biological, behavioural and egocentric network data were collected. Respondents named locations where they frequented and where they engaged in risk behaviours, including the use of crack cocaine, injectable and non-injectable drug use, and solvents; and either seeking sex work clients or sex workers. Locations were geo-coded up to Statistics Canada dissemination areas. Two-mode network visualisation and centrality, degree, and betweenness measures were generated using UciNet (V.6). O.20 -Sexual partnerships and networksResults From a sample size of 600, nine locations were named by 10 or more respondents. The following results pertain only to these nine locations (N: 231). Locations corresponded to three "hot spots" in Winnipeg's inner and outer core areas. Across the sample, HIV and HCV prevalence was 9.8% and 51.5%, respectively. Prevalence varied considerably by location, ranging from 0% to 15% for HIV and 20% to 70% for HCV. Degree ranged from 0.054 to 0.330, closeness from 0.178 to 0.411 and betweenness from 0.054 to 0.521. No association between prevalence of HIV and HCV and network met...
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