Outer Membrane Protein A (OmpA) of Shigella flexneri 2a, Induces Protective Immune Response in a Mouse Model

Abstract

BackgroundIn our earlier studies 34 kDa outer membrane protein (OMP) of Shigella flexneri 2a has been identified as an efficient immunostimulant.Key ResultsIn the present study MALDI-TOF MS analysis of the purified 34 kDa OMP of Shigella flexneri 2a shows considerable sequence homology (Identity 65%) with the OmpA of S. flexneri 2a. By using the specific primers, the gene of interest has been amplified from S. flexneri 2a (N.Y-962/92) genomic DNA, cloned in pET100/D-TOPO® vector and expressed using induction with isopropyl thiogalactoside (IPTG) for the first time. Immunogenicity and protective efficacy of the recombinant OmpA has been evaluated in an intranasally immunized murine pulmonary model. The recombinant protein induces significantly enhanced protein specific IgG and IgA Abs in both mucosal and systemic compartments and IgA secreting cells in the systemic compartment (spleen). The mice immunized with OmpA have been protected completely from systemic challenge with a lethal dose of virulent S. flexneri 2a. Immunization with the protein causes mild polymorphonuclear neutrophil infiltration in the lung, without inducing the release of large amounts of proinflammatory cytokines.ConclusionThese results suggest that the OmpA of S. flexneri 2a can be an efficacious mucosal immunogen inducing protective immune responses. Our findings also demonstrate that antibodies and Th1 immune response may be associated with the marked protective efficacy of immunized mice after intranasal shigellae infection.

Outer Membrane Protein A (OmpA) of Shigella flexneri 2a Links Innate and Adaptive Immunity in a TLR2-dependent Manner and Involvement of IL-12 and Nitric Oxide

“…Immunogen-OmpA of S. flexneri 2a was expressed in E. coli BL21(DE3) and purified as described previously (5). The lipopolysaccharide present in the recombinant OmpA was removed by passing the protein through Detoxi-Gel endotoxinremoving resin and S3⌬ peptide affinity gel columns, respectively.…”

“…Immunization of mice was performed as described previously (5). Briefly, 7-week-old BALB/c female mice weighing ϳ25 g were sedated by intramuscular injection of a mixture of 0.3 mg of xylazine hydrochloride and 1.0 mg of ketamine hydrochloride in 50 l of saline.…”

“…Furthermore, it has been observed that the protein commences innate immune response through up-regulation of TLR2, adaptor protein MyD88, p38 MAP kinase, NF-B, production of type-1 cytokines and chemokines, as well as T cell costimulatory molecules (MHC II, CD40, and CD80) in macrophages (4). Recently the 34-kDa OMP has been identified as OmpA of S. flexneri 2a (5). Moreover, it has been observed that OmpA of S. flexneri 2a elicits strong protective immunity in mice against shigellosis, which may involve a Th1-directed cell-mediated response (5).…”

“…Recently the 34-kDa OMP has been identified as OmpA of S. flexneri 2a (5). Moreover, it has been observed that OmpA of S. flexneri 2a elicits strong protective immunity in mice against shigellosis, which may involve a Th1-directed cell-mediated response (5). OmpA of Klebsiella pneumoniae has been shown to bind to and activate human macrophages and immature monocyte-derived dendritic cells in a TLR2-dependent manner (6,7).…”

Characterization of a Novel Fusion Protein from IpaB and IpaD of Shigella spp. and Its Potential as a Pan-Shigella Vaccine

“…Several groups have explored different approaches in the search of a Shigella vaccine. These include live attenuated strains of Shigella (16), LPS-protein conjugates (17), mixtures of subunit components (18), and recombinant proteins (19). Our hypothesis is that a vaccine comprised of highly conserved protein antigens would provide broad, serotype-independent protection, thus bypassing the need to consider multiple serotypes as are needed for vaccines that target LPS or O antigen.…”