Application of immunoproteomics to leptospirosis: towards clinical diagnostics and vaccine discovery.

Each of the currently available methods for serodiagnosis of leptospirosis, including the microscopic agglutination test (MAT), has its own drawback(s) when used in clinical practice. A new diagnostic test is therefore required for an earlier and more accurate diagnosis of leptospirosis. We applied immunoproteomics to define potential immunogens from five serovars of Leptospira reference strains. A leptospiral whole cell lysate from each serovar was used as the antigen to react with IgG and IgM in the sera from four patients with a positive MAT. Sera from four non-leptospirosis patients with a negative MAT were pooled and used as the negative control. 2-D Western blot analysis showed that the degree of immunoreactivity corresponded with the MAT titers. No immunoreactive spots were detected when the pooled control sera were used. A total of 24 protein spots immunoreacted with IgM and/or IgG from patients with leptospirosis. These immunoreactive proteins were identified by MALDI-TOF MS and were classified into five groups, including flagellar proteins, chaperones/heat shock proteins, transport proteins, metabolic enzymes, and hypothetical proteins. More immunoreactive spots were detected with anti-human IgG in the sera of all patients and with all the serovars of leptospires used. Some of the identified proteins immunoreacted only with IgG, whereas the others were detectable with both IgM and IgG. Among the immunoreactive proteins identified, FlaB proteins (flagellin and flagellar core protein) have been shown to have a potential role in clinical diagnostics and vaccine development. These data underscore the significant impact of immunoproteomics in clinical applications.