Loss of neurovirulence is associated with reduction of cerebral capillary sequestration during acute Babesia bovis infection

Abstract

Background: Severe neurological signs that develop during acute infection by virulent strains of Babesia bovis are associated with sequestration of infected erythrocytes in cerebral capillaries. Serial passage of virulent strains in cattle results in attenuated derivatives that do not cause neurologic disease. We evaluated whether serial passage also results in a loss of cerebral capillary sequestration by examining brain biopsies during acute disease and at necropsy.

Findings: Cerebral biopsies of spleen intact calves inoculated intravenously with a virulent or attenuated strain pair of B. bovis were evaluated for capillary sequestration at the onset of babesiosis and during severe disease. In calves infected with the virulent strain, there was a significant increase in sequestration between the first and second biopsy timepoint. The attenuated strain was still capable of sequestration, but at a reduced level, and did not change significantly between the first and second biopsy. Necropsy examination confirmed the second biopsy results and demonstrated that sequestration identified at necropsy reflects pathologic changes occurring in live animals.

Conclusions: Loss of neurovirulence after serial in vivo passage of the highly virulent T2Bo strain of B. bovis in splenectomized animals is associated with a significant reduction of cerebral capillary sequestration. Previous genomic analysis of this and two other strain pairs suggests that this observation could be related to genomic complexity, particularly of the ves gene family, rather than consistent gene specific differences. Additional experiments will examine whether differential gene expression of ves genes is also associated with reduced cerebral sequestration and neurovirulence in attenuated strains.

Figure 1

Sequential evaluation of cerebral capillary sequestration during acute babesiosis. (A) The timing of biopsy correlated with changes in rectal temperature during infection. Animals infected with T2Bo_vir are in red, and animals infected with T2Bo_att are in blue. The red arrows at days 6 and 8 indicate the first and second biopsy for T2Bo_vir infected animals. The blue arrows at days 7 and 9–10 indicate the first and second biopsy, respectively, for T2Bo_att infected animals. (B) Representative photomicrographs of brain biopsies of either the T2Bo_vir or T2Bo_att infected group. First biopsy (panels 1 and 4) was taken one day following the onset of fever. Second biopsy (panels 2 and 5) was taken 2–3 days following the first biopsy. Panels 3 and 6 represent samples taken within one hour of death. (C) Levels of sequestration observed in brain biopsies. There is a significant difference in sequestration between first and second biopsies (p < 0.0001), as well as first biopsy and necropsy (p < 0.0001) for the T2Bo_vir group. A significant difference between the T2Bo_vir and T2Bo_att groups for second biopsy and necropsy (p < 0.05) was also observed.