Identification of novel immune correlates of protection against acute bovine babesiosis by superinfecting cattle with in vitro culture attenuated and virulent Babesia bovis strains

Authors: BASTOS, REGINALDO - Washington State University; LAUGHERY, JACOB - Washington State University; OZUBEK, SEZAYI - University Of Firat; ALZAN, HEBA - Egypt National Research Center; Taus, Naomi; Ueti, Massaro; Suarez, Carlos

Submitted to: Frontiers in Immunology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 11/2/2022
Publication Date: 11/18/2022
Citation: Bastos, R.G., Laughery, J.M., Ozubek, S., Alzan, H.F., Taus, N.S., Ueti, M.W., Suarez, C.E. 2022. Identification of novel immune correlates of protection against acute bovine babesiosis by superinfecting cattle with in vitro culture attenuated and virulent Babesia bovis strains. Frontiers in Immunology. 13. Article 1045608. https://doi.org/10.3389/fimmu.2022.1045608.
DOI: https://doi.org/10.3389/fimmu.2022.1045608

Interpretive Summary: Tickborne blood parasites of the genus Babesia are the causative agent of babesiosis, a disease that negatively impacts public and animal health around the world. Effective and sustainable vaccines against babesiosis are urgently needed. In this study attenuated and virulent homologous strains of Babesia bovis were used to investigate which immune responses correlated with protection of cattle against acute bovine babesiosis. Holstein calves were infected intravenously (IV) with an in vitro culture of attenuated B. bovis strain while three age-matched Holstein calves were inoculated IV with uninfected red blood cells (RBC) as controls. Calves were monitored for signs of disease and blood was collected to measure changes in immune responses. All calves infected with the attenuated strain of B. bovis developed mild signs of acute babesiosis characterized by increase in temperature as early as 6 days post-infection, but the animals recovered without treatment. The calves also showed changes in immune cells and cytokines compared to the control animals. At 46 days after infection with the attenuated strain, all experimental calves were challenged IV with the homologues virulent B. bovis strain. Interestingly, calves infected with the attenuated strain survived challenge with the virulent strain without displaying signs of acute babesiosis. In contrast, control animals presented marked signs of acute disease, starting at day 10 post-challenge, and two of them were humanely euthanized at days 13 and 14 after challenge due to the severity of the disease. Following challenge, control calves showed higher parasite load in peripheral blood than animals previously exposed to the attenuated strain. However, no significant alterations in the profile of immune cells and cytokines were observed in attenuated strain infected and control calves after challenge. In conclusion, by using attenuated and virulent homologous strains of B. bovis and cattle as a model of infection, we identified novel alterations in blood immune cells and profile cytokine expression in peripheral blood that are associated with protection against acute B. bovis infection. These immune correlates of protection may be useful in the design of effective and sustainable vaccines against human and animal babesiosis.

Technical Abstract: The apicomplexan tickborne parasites Babesia bovis and B. bigemina are the major causative agents of bovine babesiosis, a disease that negatively affects the cattle industry and food safety around the world. The absence of correlates of protection represents one major impediment for the development of effective and sustainable vaccines against bovine babesiosis. Herein we superinfected cattle with attenuated and virulent strains of B. bovis to investigate immune correlates of protection against acute bovine babesiosis. Three 6-month-old Holstein calves were infected intravenously (IV) with the in vitro culture attenuated Att-S74-T3Bo B. bovis strain (106 infected bovine red blood cells (iRBC)/calf) while three age-matched Holstein calves were inoculated IV with normal RBC as controls (106 RBC/calf). All Att-S74-T3Bo-infected calves showed a significant increase in temperature early after inoculation but recovered without treatment. Att-S74-T3Bo-infected calves also developed: (a) monocytosis, neutropenia, and CD4+ lymphopenia in peripheral blood on days 3 to 7 post-inoculation; (b) significant levels of TNFa, CXCL10, IFN', IL-4, and IL-10 in sera at day 6 after infection; and (c) IgM and IgG against B. bovis antigens, starting at days 10 and 30 post-inoculation, respectively. At 46 days post-Att-S74-T3Bo inoculation, all experimental calves were infected IV with the homologous virulent B. bovis strain Vir-S74-T3Bo (107 iRBC/calf). All Att-S74-T3Bo-infected calves survived superinfection with Vir-S74-T3Bo without displaying signs of acute babesiosis. In contrast, control animals showed signs of acute disease, starting at day 10 post-Vir-S74-T3Bo infection, and two of them were humanely euthanized at days 13 and 14 after inoculation due to the severity of their symptoms. Also, control calves showed higher (P<0.05) parasite load in peripheral blood compared to animals previously exposed to Att-S74-T3Bo. No significant alterations in the profile of leukocytes and cytokines were observed in Att-S74-T3Bo-inoculated after Vir-S74-T3Bo infection. In conclusion, data demonstrate novel changes in the profile of blood immune cells and cytokine expression in peripheral blood that are associated with protection against acute bovine babesiosis. These identified immune correlates of protection may be useful for designing effective and sustainable vaccines against babesiosis in cattle.