Leptospiral proteins recognized during the humoral immune response to leptospirosis in humans

Leptospirosis is an emerging zoonosis caused by pathogenic spirochetes belo nging to the genus Leptospira. An understanding of leptospiral protein expr ession regulation is needed to develop new immunoprotective and serodiagnos tic strategies. We used the Immoral immune response during human leptospiro sis as a reporter of protein antigens expressed during infection. Qualitati ve and quantitative immunoblot analysis was performed using sera from 105 p atients from Brazil and Barbados. Sera from patients with other diseases an d healthy individuals were evaluated as controls. Seven proteins, p76, p62, p48, p45, p41, p37, and p32, were identified as targets of the humoral res ponse during natural infection. In both acute and convalescent phases of il lness, antibodies to lipopolysaccharide were predominantly immunoglobulin M (IgM) while antibodies to proteins were exclusively IgG. Anti-p32 reactivi ty had the greatest sensitivity and specificity: positive reactions were ob served in 37 and 84% of acute- and convalescent-phase sera, respectively, w hile only 5% of community control individuals demonstrated positive reactio ns. Six immunodominant antigens were expressed by all pathogenic leptospira l strains tested; only p37 was inconsistently expressed. Two-dimensional im munoblots identified four of the seven infection-associated antigens as bei ng previously characterized proteins: LipL32 (the major outer membrane lipo protein), LipL41 (a surface-exposed outer membrane lipoprotein), and heat s hock proteins GroEL and DnaK. Fractionation studies demonstrated LipL32 and LipL41 reactivity in the outer membrane fraction and GroEL and DnaK in the cytoplasmic fraction, while p37 appeared to be a soluble periplasmic prote in. Most of the other immunodominant proteins, including p48 and p45, were localized to the inner membrane. These findings indicate that leptospiral p roteins recognized during natural infection are potentially useful for sero diagnosis and may serve as targets for vaccine design.