Discovery of Human HLA T Cell Epitopes in Human Metapneumovirus HMPV is a leading cause of lower respiratory infection in children and adults worldwide. Although nearly all people are infected with HMPV by age 5 years, immunity to HMPV is incomplete and re-infections occur throughout life. More severe disease occurs in persons with underlying conditions such as asthma, chronic obstructive pulmonary disease, HIV, or prematurity. Data from our group and others shows that T cells are important to clear infection in mice and humans, yet T cell response can also contribute to disease severity. Thus, characterizing the human T cell response to HMPV is important to guide safe and effective vaccine development. Our lab has established methods to discover human MHC-I and MHC-II epitopes using HLA-transgenic (huHLA-tg) mice and human PBMCs. The huHLA-tg mice express different HLA supertypes, which collectively capture a large proportion of different HLA alleles among diverse human populations. We propose to apply these methods to discover HLA-restricted HMPV epitopes recognized by CD8+ and CD4+ T cells. We will screen both overlapping peptide pools and predictope peptides by ELISPOT against T cells derived from huHLA-tg mice infected with HMPV. Individual epitope peptides that are confirmed by ELISpot will be ordered as tetramers from the NIH Tetramer Core. Tetramers will be used to thoroughly analyze and characterize HMPV epitope-specific CD8+ T cells, including: effector molecule production such as IFN-γ, TNF, IL-2, perforin, and granzyme; degranulation using CD107a; transcription factor profile; and inhibitory receptor expression. Dominant epitopes confirmed by tetramer staining will be tested for protective efficacy using peptide vaccination and live virus challenge of mice. HLA-typed human PBMCs will be used to confirm bone fide recognition of the epitopes by human CD8+ T cells. The results of this project will identify widely shared HMPV epitopes that can be used to design candidate vaccines and evaluate the response to any HMPV vaccine. These tools will be needed to understand human immune responses to both natural infection and vaccines, which is important given the immune-mediated pathology associated with HMPV.