Noriyuki Takabatake

Inhibitory Effect of Antiserum to Surface Antigen P50 of Babesia gibsoni on Growth of Parasites in Severe Combined Immunodeficiency Mice Given Canine Red Blood Cells

ABSTRACT The inhibitory effect of an antiserum to surface protein P50 of Babesia gibsoni on the growth of the parasite was determined with severe combined immunodeficiency mice given canine red blood cells. The antiserum to the recombinant P50 protein significantly inhibited the parasite growth, indicating that P50 might be a useful vaccine candidate.

Modification of Host Erythrocyte Membranes by Trypsin and Chymotrypsin Treatments and Effects on the In Vitro Growth of Bovine and Equine Babesia Parasites

In the present study, we investigated the effects of protease pretreatments of host erythrocytes (RBC) on the in vitro growth of bovine Babesia parasites (Babesia bovis and B. bigemina) and equine Babesia parasites (B. equi and B. caballi). The selected proteases, trypsin and chymotrypsin, clearly modified several membrane proteins of both bovine and equine RBC, as demonstrated by SDS-PAGE analysis; however, the protease treatments also modified the sialic acid content exclusively in bovine RBC, as demonstrated by lectin blot analysis. An in vitro growth assay using the protease-treated RBC showed that the trypsin-treated bovine RBC, but not the chymotrypsin-treated ones, significantly reduced the growth of B. bovis and B. bigemina as compared to the control. In contrast, the growth of B. equi and B. caballi was not affected by any of these proteases. Thus, the bovine, but not the equine, Babesia parasites require the trypsin-sensitive membrane (sialoglyco) proteins to infect the RBC.

Amount of cholesterol in host membrane affects erythrocyte invasion and replication by Babesia bovis

Cholesterol is a major component of the erythrocyte membrane. In the present study, we investigated the effects of cholesterol reduction in host bovine erythrocytes (RBC) on the growth of Babesia bovis, a major bovine haemoprotozoon. An in vitro growth assay with bovine RBC that had been prepared by pre-treatment with a cholesterol depletion agent (methyl-beta-cyclodextrin, MCD) showed that the culture with 5 mM MCD-treated RBC inhibited the growth of B. bovis significantly as compared with that with the control RBC. In further experiments, the treatment with 5 mM MCD was proved to suppress both activities of the parasite, erythrocyte invasion and replication within the infected RBC. In contrast, a slight reduction in the membrane cholesterol by 1 mM MCD treatment promoted both their growth and erythrocyte invasion activity. These results indicate that erythrocyte invasion and replication by B. bovis are affected by the amount of cholesterol in the host erythrocyte membrane.

ASEXUAL GROWTH OF BABESIA BOVIS IS INHIBITED BY SPECIFIC SULFATED GLYCOCONJUGATES

In the present study, inhibitory effects of several sulfated and nonsulfated glycoconjugates were evaluated on the in vitro asexual growth of Babesia bovis. Among the selected sulfated glycoconjugates, dextran sulfate, heparin, heparan sulfate, fucoidan, and chondroitin sulfate B strongly inhibited the parasitic growth, and all but chondroitin sulfate B induced a significant accumulation of extracellular merozoites in culture. In contrast, chondroitin sulfate A, keratan sulfate, and protamine sulfate, as well as nonsulfated dextran and hyaluronic acid, did not influence the growth. These findings indicate that the asexual growth of B. bovis merozoites is inhibited by specific sulfated glycoconjugates, possibly providing us with an important insight into the molecular interaction(or interactions) during the process of the erythrocyte invasion by B. bovis merozoites.

Molecular Cloning of a Babesia caballi Gene Encoding the 134-Kilodalton Protein and Evaluation of Its Diagnostic Potential in an Enzyme-Linked Immunosorbent Assay

ABSTRACT A Babesia caballi gene encoding the 134-kDa (BC134) protein was immunoscreened with B. caballi-infected horse serum. An enzyme-linked immunosorbent assay (ELISA) using recombinant BC134 protein could effectively differentiate B. caballi-infected horse sera from Babesia equi-infected or noninfected control horse sera. These results suggest that the recombinant BC134 protein is a potential diagnostic antigen in the detection of B. caballi infection.

Isolation and pathogenic characterization of an OB1 variant of Babesia rodhaini which has a glycophorin A-independent pathway to murine red blood cells.

Recent studies using several Babesia spp. have demonstrated that these species commonly recognize host sialic acids of red blood cells (RBCs) for their invasion. Glycophorin A (GPA), which is a major carrier of the sialic acids on RBCs, is a possible invasive receptor for Babesia parasites. In the present study, a variant of Babesia rodhaini was successfully isolated from a GPA homozygous knockout (GPA(-/-)) mouse infected with an Australian strain of B. rodhaini which had originally been unable to replicate in GPA(-/-) mice. The isolated parasite (designated as an OB1 variant) caused lethal infection to wild-type mice, as in the case of the parent Australian strain. However, although the growth of the OB1 variant in GPA(-/-) mice was comparable with that in wild-type mice at 1-4 days after infection, the growth was significantly inhibited from day 5 onward, leading to the eventual survival of the GPA(-/-) mice. Resistance of GPA(-/-) mice against OB1 infection was lost by splenectomy, although the cytokine responses to the infection in the sera of GPA(-/-) mice were similar to those of wild-type mice. The autoantibody levels to GPA-defective RBCs in the sera of GPA(-/-) mice were depressed at a lower level at 0-2 days after infection than those of wild-type mice, while the levels of GPA(-/-) mice progressively increased and reached comparable levels to those of wild-type mice at day 3 or later. These results indicate that the isolated OB1 variant has a GPA-independent invasion pathway into murine RBCs and suggest that the resistance of GPA(-/-) mice against infection with the OB1 variant may be attributed to the effective clearance of the parasitized RBCs lacking GPA in the spleen, possibly mediated by preferential autoantibody binding to the RBC membrane.