Purification, characterization of O-acetylated sialoglycoconjugates-specific IgM, and development of an enzyme-linked immunosorbent assay for diagnosis and follow-up of indian visceral leishmaniasis patients

Abstract

The surface expression of 9-O-acetylated sialic acid (9-OAcSA) is elevated on hematopoietic cells and erythrocytes of visceral leishmaniasis (VL) patients. In this study, we show that VL patients contain elevated levels of IgM antibodies directed against 9-O-acetylated sialoglycoconjugates (9-OAcSG). These antibodies were affinity purified with bovine submaxillary protein as the affinity matrix containing the terminal epitope, 9-OAcSAα2-6GalNAc. They also bound to 9-OAcSGs on hematopoietic cells of patients with VL and to epitopes in the cytosol of Leishmania donovani promastigotes. A novel enzyme-linked immunosorbent assay was employed that showed 4-fold higher anti-OAcSG titers in VL patients (n = 38), mean ± S.E.M. being 0.83 ± 0.09 vs. 0.21 ± 0.04 detected in normal donors (n = 20) and patients with cross-reactive diseases such as malaria (n = 4) or tuberculosis (n = 4). Assay specificity and sensitivity was 100% and 92%, respectively, whereas positive and negative predictive values were 100% and 90%, respectively. Significantly, anti-OAcSG titers declined 30 days after completion of anti-leishmanial treatment, indicating that monitoring of anti-9-OAcSGs may be a valuable alternative toward increasing the efficiency of diagnosis and follow-up of VL.