Investigating Responses of Bovine Gamma Delta T cell Subsets During Early MAP Infection

It has been suggested that gamma delta (γδ) T lymphocytes play a role in pathogenesis of mycobacterial infections. Unlike other species, ruminants, have many γδT cells, making them an excellent model to study these cells. Bovine γδ T cells are subdivided into two main subsets based on their expression of the surface molecule Workshop cluster 1 (WC1). WC1+ cells are located primarily in peripheral blood and because they are readily accessible, these are widely studied. The WC1- subset is preferentially localized along mucosal surfaces, suggesting their involvement in innate immunity and surveillance. Data have recently shown differential recruitment of γδ T cell subsets to Mycobacterium avium subspecies paratuberculosis (MAP) infection sites of calves and differential responses of subsets to MAP antigens; however the mechanisms driving these phenomena remain unknown and unexplored. Our first objective was to compare responses of subsets of naïve γδ T cells to various mycobacterial antigens in vitro. We measured cell proliferation and interferon gamma (IFNγ) secretion to characterize responses of subsets. The second objective was to evaluate recruitment and activation of γδ T cell subsets into the ileal mucosa during early experimental enteric MAP infection in calves by fluorescence microscopy and quantitative reverse transcription polymerase chain reaction (RT-qPCR) for cytokine gene expression. Our hypotheses were that naïve WC1- γδ T cells have an enhanced response to stimulation compared to WC1+ γδ T cells in vitro, and that WC1- cells are recruited to intestinal MAP infection sites where they are activated for pro-inflammatory cytokine secretion. Our findings support the hypotheses with in vitro assays showing distinctly different proliferative and cytokine responses of WC1- γδ T cell and WC1+ γδ T cells in various culture conditions, and in vivo experiments demonstrating distinct patterns of γδ T cell recruitment and cytokine gene expression within the distal ileum of MAP-infected compared to uninfected calves. This research provides a platform for the beginnings of understanding the differences in the immunological roles of bovine γδ T cell subsets and their participation in host immunity during MAP infection.