Phenotype and cytokine production of Histomonas meleagridis-specific T cells isolated from chickens and turkeys infected with the parasite or vaccinated against histomonosis
Gisela F. Erf
Dissertation - University of Veterinary Medicine Vienna - 2020
The aim of this PhD-project was to establish assays for detection of type-1 (IFN-γ) and type-2 (IL-13) cytokines to study the T-cell response towards H. meleagridis of chickens and turkeys. During the pilot experiments, intracellular cytokine staining (IFN-γ) and PrimeFlowTM RNA (IL-13) assays could be successfully developed and further scrutinized in controlled infection/vaccination studies within the chicken and turkey, respectively. Clonal monoxenic histomonads cultures were applied for either infection using the virulent strain (low passage) or vaccination using the attenuated strain (high passage). Findings obtained from the pilot experiments in the chicken showed significant increases of H. meleagridis-specific IFN-γ- producing splenocytes within the CD4+ and CD4-CD8β- T-cell subset of infected chickens compared to naïve controls. For vaccinated turkeys, very slight increments of IFN-γ+ liver cells were found within CD4+ and non-CD4 T cells in comparison to non-treated control birds. On the contrary to IFN-γ, extremely low frequencies of IL-13 mRNA+ cells were detected within spleens and livers of both species. As these observations suggested a dominating type-1 immune response towards H. meleagridis, the subsequent main experiments focused on IFN-γ detection employing an optimized staining panel for T-cell subsets. In addition, the T-cell immune responses of chickens and turkeys vaccinated and subsequent challenge infected were studied next to infected or vaccinated birds compared to non-inoculated control animals. While inoculation with the attenuated parasite strain seems to induce a rather weak systemic immune response in both species, the virulent strain leads to a strong systemic T-cell response in the spleen and in addition in the liver of vaccinated/infected turkeys. Significant increases of IFN- γ-producing splenocytes were detected within the CD4+ T-cell subset isolated from infected chickens while no such differences were found in other T-cell subsets, inoculation groups or the liver. These findings suggest a strong involvement of responsive T cells at the local mucosal site of infection resulting in a retention of histomonads in the cecum. In contrast, vaccinated and infected turkeys showed an induction of IFN-γ-producing lymphocytes within all major T- cell subsets (CD4+, CD8α+ and CD3+CD4-CD8α-) from spleen and liver, which indicates that immune responses at the mucosal site do not seem to be potent enough for restricting the infection to the ceca. To fully understand H. meleagridis-specific responses and potentially 89 identify correlates of protection, investigation of INF-γ-producing T cells at the mucosal site in addition to screening a broader panel of cytokines would be desirable as soon as suitabletools for detection are available.
Dissertation - Veterinärmedizinische Universität Wien - 2020
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