Protection against lethal leptospirosis after vaccination with LipL32 coupled or coadministered with the B subunit of Escherichia coli heat-labile enterotoxin

Abstract

Leptospirosis, a worldwide zoonosis, lacks an effective, safe, and cross-protective vaccine. LipL32, the most abundant, immunogenic, and conserved surface lipoprotein present in all pathogenic species of Leptospira, is a promising antigen candidate for a recombinant vaccine. However, several studies have reported a lack of protection when this protein is used as a subunit vaccine. In an attempt to enhance the immune response, we used LipL32 coupled to or coadministered with the B subunit of the Escherichia coli heat-labile enterotoxin (LTB) in a hamster model of leptospirosis. After homologous challenge with 5× the 50% lethal dose (LD(50)) of Leptospira interrogans, animals vaccinated with LipL32 coadministered with LTB and LTB::LipL32 had significantly higher survival rates (P < 0.05) than animals from the control group. This is the first report of a protective immune response afforded by a subunit vaccine using LipL32 and represents an important contribution toward the development of improved leptospirosis vaccines.

Fig 1

Characterization of purified recombinant proteins. (A) SDS-PAGE analysis; (B) anti-LTB Western blot analysis; (C) anti-LipL32 Western blot analysis; (D) Western blot analysis of human convalescent-phase sera. Lanes: 1, E. coli whole-cell extract; 2, rLTB; 3, rLipL32; 4, rLTB::LipL32 (A to C). Lanes: 1, L. interrogans strain L1-130 whole-cell extract; 2, rLTB; 3, rLipL32; 4, rLTB::LipL32 (D).

Fig 2

Recombinant protein GM1-binding ELISA. Different letters represent statistical differences (P < 0.05). Uppercase letters relate to significance for the anti-CT antibody, and lowercase letters relate to significance for the anti-LipL32 antibody. OD, optical density.

Fig 3

Humoral immune response against rLipL32 examined by ELISA. Error bars represent standard deviations. Both postimmune treatment groups were different from the preimmune group and control serum at all dilutions.

Fig 4

Hamster survival timeline (grouped results of three independent experiments). Different letters represent statistically different results (P < 0.05). Timelines represent the rLTB::LipL32 (■), rLTB-plus-rLipL32 (△), rLTB (▽), bacterin (----), and negative-control (□) groups. Statistical analyses and graph generation were carried out with GraphPad Prism 4 software systems (GraphPad Software).