Synthetic Oligomers Mimicking Capsular Polysaccharide Diheteroglycan are Potential Vaccine Candidates against Encapsulated Enterococcal Infections.

Infections caused by Enterococcus spp. are a major concern in the clinical setting. In Enterococcus faecalis, the capsular polysaccharide diheteroglycan (DHG), composed of ß-d-galactofuranose-(1 → 3)-ß-d-glucopyranose repeats, has been described as an important virulence factor and as a potential vaccine candidate against encapsulated strains. Synthetic structures emulating immunogenic polysaccharides present many advantages over native polysaccharides for vaccine development. In this work, we described the synthesis of a library of DHG oligomers, differing in length and order of the monosaccharide constituents. Using suitably protected thioglycoside building blocks, oligosaccharides up to 8-mer in length built up from either Galf-Glcp or Glcp-Galf dimers were generated, and we evaluated their immunoreactivity with antibodies raised against DHG. After the screening, we selected two octasaccharides, having either a galactofuranose or glucopyranose terminus, which were conjugated to a carrier protein for the production of polyclonal antibodies. The resulting antibodies were specific toward the synthetic structures and mediated in vitro opsonophagocytic killing of different encapsulated E. feacalis strains. The evaluated oligosaccharides are the first synthetic structures described to elicit antibodies that target encapsulated E. faecalis strains and are, therefore, promising candidates for the development of a well-defined enterococcal glycoconjugate vaccine.

Conjugation of Different Immunogenic Enterococcal Vaccine Target Antigens Leads to Extended Strain Coverage.

Enterococci have emerged as important nosocomial pathogens due to their resistance to the most commonly used antibiotics. Alternative treatments or prevention options are aimed at polysaccharides and surface-related proteins that play important roles in pathogenesis. Previously, we have shown that 2 Enterococcus faecium proteins, the secreted antigen A and the peptidyl-prolyl cis-trans isomerase, as well as the Enterococcus faecalis polysaccharide diheteroglycan, are able to induce opsonic and cross-protective antibodies. Here, we evaluate the use of glycoconjugates consisting of these proteins and an enterococcal polysaccharide to develop a vaccine with broader strain coverage. Diheteroglycan was conjugated to these 2 enterococcal proteins. Rabbit sera raised against these glycoconjugates showed Immunoglobulin G titers against the corresponding conjugate, as well as against the respective protein and carbohydrate antigens. Effective opsonophagocytic killing for the 2 sera was observed against different E. faecalis and E. faecium strains. Enzyme-linked immunosorbent assays against whole bacterial cells showed immune recognition of 22 enterococcal strains by the sera. Moreover, the sera conferred protection against E. faecalis and E. faecium strains in a mouse infection model. Our results suggest that these glycoconjugates are promising candidates for vaccine formulations with a broader coverage against these nosocomial pathogens and that the evaluated proteins are potential carrier proteins.