Bovine babesiosis: Partial purification and characterization of blood culture-derived Babesia bovis

Morphologic, biologic and immunologic properties of corpuscular and soluble fractions of Babesia bovis purified from in vitro blood cultures were studied. Supernatant fluids obtained during routine medium exchange were studied. Supernatant fluids obtained during routine medium exchange were submitted to differential centrifugation to separate soluble and corpuscular babesial antigens from erythrocyte stroma. Extracellular babesiae were sedimented with infected and uninfected erythrocyte ghosts. The majority of babesiae were found in erythrocyte ghosts. Clumps of extracellular parasites were sometimes formed in vitro and generally could not be separated from uninfected erythrocytes. Centrifugation over a discontinuous Ficoll density gradient did not improve separations. Parasites remained viable throughout the purification procedure but were killed by freezing and rapid thawing. Both corpuscular and soluble antigen fractions elicited the production of specific anti-babesial antibodies when injected into calves. Electron microscopy of corpuscular antigen revealed the presence of intra- and extraerythrocytic babesial merozoites. A surface coat was visible loosely adhering to the plasma membrane of the parasites. Parasite suspensions and cell-free supernatant fluids obtained from in vitro cultures of B. bovis should provide a variety of unique antigens for further in vitro and in vivo studies.