A 37-KILODALTON GLYCOPROTEIN OF BABESIA DIVERGENS IS A MAJOR COMPONENT OF A PROTECTIVE FRACTION CONTAINING LOW-MOLECULAR-MASS CULTURE-DERIVED EXOANTIGENS

The supernatants of in vitro cultures of Babesia divergens Rouen 1987 in human erythrocytes, obtained by using a semidefined medium based on human high-density lipoproteins, were fractionated by get filtration chromatography into four fractions, F-1 to F-4. The crude supernatant as well as each fraction adjuvanted with Quil-A protected gerbils from mortality due to a homologous infectious challenge. Analysis of the h umoral response of the 10 protected gerbils with fraction F-4, contain ing major proteins with molecular masses lower than 50 kDa, showed tha t a few antigens (from 50 to 17 kDa) could be important candidates for an improved vaccine against B. divergens babesiosis. As an immunodomi nant response was directed against the 37-kDa antigen (Bd37) in two di fferent B. divergens strains tested, a polyclonal antibody directed ag ainst Bd37 was produced in a rabbit. In an immunofluorescence assay, t he anti-Bd37 antiserum strongly labelled small internal vesicles of th e merozoites and the cell surface was diffusely labelled after fixatio n, whereas on live merozoites, this labelling was not observed, [H-3]g lucosamine-radiolabelling experiments demonstrate that Bd37 is a glyco protein. The Bd37 protein can also be labelled with [C-14]palmitate bu t not with [H-3]myristic acid, In Triton X-114 temperature phase parti tioning of B. divergens-infected erythrocyte extracts, Bd37 was exclus ively found into the detergent phase, indicating that the palmitoylate d Bd37 protein was in the membrane fraction. In the in vitro supernata nt, the glycoprotein Bd37 was found in a nonpalmitoylated form, indica ting excretion and/or release of the glycoprotein from the merozoite.