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ARS Home » Pacific West Area » Pullman, Washington » Animal Disease Research » Research » Publications at this Location » Publication #373289

Research Project: Development of Detection and Control Strategies for Bovine Babesiosis and Equine Piroplasmosis

Location: Animal Disease Research

Title: Flow cytometric analysis of the cytotoxic T-cell recall response to Theileria parva in cattle following vaccination by the infection and treatment method

item ELNAGGAR, MAHMOUD - Washington State University
item KNOWLES, DONALD - Washington State University
item DAVIS, WILLIAM - Washington State University
item Fry, Lindsay

Submitted to: Veterinary Sciences
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 6/11/2021
Publication Date: 6/18/2021
Citation: Elnaggar, M.M., Knowles, D.P., Davis, W., Fry, L.M. 2021. Flow cytometric analysis of the cytotoxic T-cell recall response to Theileria parva in cattle following vaccination by the infection and treatment method. Veterinary Sciences. 8(6). Article 114.

Interpretive Summary: Theileria parva, an intracellular protozoan parasite, is the leading infectious cause of death of cattle in sub-Saharan Africa. Current preventive strategies are incredibly limited, and improved vaccination and treatment strategies are urgently needed to decrease the socioeconomic impact of this parasite. One significant hindrance to the development of a new vaccine is an incomplete understanding of protective immunity to T. parva. In large part, this lack of understanding can be attributed to a lack of methods to fully characterize the T cells that comprise the protective response. In this manuscript, we describe the development of a flow-cytometry-based assay that can be used to simultaneously assess not only the killing ability of bovine T cells, but also the phenotypic signature of those cells. This information can be used to expedite the development and laboratory assessment of next-generation vaccine candidates for T. parva.

Technical Abstract: The intra-lymphocytic, apicomplexan parasite, Theileria parva, is the causative agent of East Coast Fever (ECF), the leading cause of death in cattle of sub-Saharan Africa. Although the development of a sustainable, next-generation vaccine to protect cattle from T. parva is a major focus of international aid organizations, progress has been impeded by the lack of a complete understanding of protective immunity that develops following natural, tick-borne exposure or via the infection treatment method (ITM) of immunization. This is in part attributable to a paucity of methods to fully characterize the phenotype and function of the memory T-cell repertoire that develops following exposure to T. parva. As reported here, ex vivo methods developed to study the immune response against mycobacterial pathogens were adapted for use in T. parva to overcome this impediment. Consistent with previous observations, a single stimulation of peripheral blood mononuclear cells (PBMC) from ITM-immunized steers using irradiated, autologous, T. parva-infected cells (TpC) elicited a proliferative recall response detectable 7 days post stimulation. Flow cytometric analysis showed CD45R0+/CCR7- CD4 and CD8 T cells comprised the proliferating cell populations after antigenic stimulation. A second round of stimulation with TpC boosted the proliferative response of both CD4 and CD8 T cell populations, yielding sufficient cell numbers for functional analysis by 14 days post stimulation. Co-incubation of stimulated PBMC cultures with autologous TpC demonstrated the presence of cytotoxic T cells (CTL) with ability to kill TpC. Comparison of CTL activity of cultures depleted of either CD4 or CD8 T cells demonstrated CTL activity was attributed primarily to CD8 T cells. An important observation made during the development of the assay was that stimulation of PBMC from steers vaccinated by the ITM always elicited proliferation of CD4 and CD8 T cells, suggesting CD4 T cell is likely essential for development of a CTL response. The assay described herein can be used to further define the immune phenotype of memory CTL and CD4 T cells in T. parva-immune cattle, facilitating subsequent development and evaluation of candidate, next-generation vaccines to prevent ECF.