Development of a glycoconjugate vaccine to prevent invasive Salmonella Typhimurium infections in sub-Saharan Africa

Abstract

Invasive infections associated with non-typhoidal Salmonella (NTS) serovars Enteritidis (SE), Typhimurium (STm) and monophasic variant 1,4,[5],12:i:- are a major health problem in infants and young children in sub-Saharan Africa, and currently, there are no approved human NTS vaccines. NTS O-polysaccharides and flagellin proteins are protective antigens in animal models of invasive NTS infection. Conjugates of SE core and O-polysaccharide (COPS) chemically linked to SE flagellin have enhanced the anti-COPS immune response and protected mice against fatal challenge with a Malian SE blood isolate. We report herein the development of a STm glycoconjugate vaccine comprised of STm COPS conjugated to the homologous serovar phase 1 flagellin protein (FliC) with assessment of the role of COPS O-acetyls for functional immunity. Sun-type COPS conjugates linked through the polysaccharide reducing end to FliC were more immunogenic and protective in mice challenged with a Malian STm blood isolate than multipoint lattice conjugates (>95% vaccine efficacy [VE] versus 30–43% VE). Immunization with de-O-acetylated STm-COPS conjugated to CRM197 provided significant but reduced protection against STm challenge compared to mice immunized with native STm-COPS:CRM197 (63–74% VE versus 100% VE). Although OPS O-acetyls were highly immunogenic, post-vaccination sera that contained various O-acetyl epitope-specific antibody profiles displayed similar in vitro bactericidal activity when equivalent titers of anti-COPS IgG were assayed. In-silico molecular modeling further indicated that STm OPS forms a single dominant conformation, irrespective of O-acetylation, in which O-acetyls extend outward and are highly solvent exposed. These preclinical results establish important quality attributes for an STm vaccine that could be co-formulated with an SE-COPS:FliC glycoconjugate as a bivalent NTS vaccine for use in sub-Saharan Africa.

Impact of branching on the conformational heterogeneity of the lipopolysaccharide from Klebsiella pneumoniae: Implications for vaccine design

“…[23] In addition, it has been shown that antibody binding can alter the conformations sampled by an oligosaccharide, which may further facilitate binding. [24,46] For an O1 OPS vaccine, despite the reduced accessibility of the β-D-Galf and α-D-Galp linear backbone monosaccharides in the RUs of D-Gal-III OPS, antibodies induced against the terminal Gal-II region should be similarly efficacious in the context of D-Dal-I or D-Gal-III given the relative insensitivity of D-Gal-II to the presence of the branches. In addition, as the Gal-II region is membrane distal, it is expected that it would be more exposed compared to the membrane proximal D-Gal-I/D-Gal-III in O1.…”

“…Results are averaged over the entire HREST2-bpCMAP trajectories.Glycosidic linkage-based clustering analysis (GL): Additional analysis was undertaken to obtain a more quantitative estimate of the conformational states being sampled by the two OPS. Conformations of the saccharides were identified based on clustering of glycosidic linkages (GL) and their sampled populations determined, as previously presented [18,23,24,46]. To perform GL clustering it is necessary to assign the different glycosidic conformations to integers on which the clustering is performed.…”

Overexpression of O‐polysaccharide chain length regulators in Gram‐negative bacteria using the Wzx‐/Wzy‐dependent pathway enhances production of defined modal length O‐polysaccharide polymers for use as haptens in glycoconjugate vaccines

“…Typhimurium glycoconjugate vaccine using wzzB ‐controlled OPS antigen haptens, proof that this strategy is effective (Baliban et al . ). Salmonella Typhimurium overexpressing wzzB produced a single high molecular weight species of O‐polysaccharide as measured by HPLC‐SEC.…”

Development of a broad spectrum glycoconjugate vaccine to prevent wound and disseminated infections with Klebsiella pneumoniae and Pseudomonas aeruginosa

“…1.5% bacteriological agar (AmericanBio, MA) was added to HS for solid media preparations. Fully chemically defined medium (CDM) used for growth of KP in fermentation cultures and to assess guanine auxotrophy has been described previously [ 19 ]. 0.004%-0.025% guanine (Sigma Aldrich, MO) was added for the growth of CVD 3001 to supplement the guaBA mutation.…”

“…KP O1, O2, O3 and O5 reagent strains were grown in 4 L fermentation cultures in a BioStat A+ fermenter (0.6–5 L working volume, 6.6 L total volume) (Sartorius, Germany) essentially as described [ 19 ]. Briefly, 5–10 colonies were selected from an HS agar plate grown overnight at 37°C and used to inoculate a 250 mL shake flask containing 50 mL of CDM (supplemented with 0.025% guanine for CVD 3001).…”