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There is increasing evidence that induction of local immune responses is a key component of effective vaccines. For respiratory pathogens, for example tuberculosis and influenza, aerosol delivery is being actively explored as a method to administer vaccine antigens. Current animal models used to study respiratory pathogens suffer from anatomical disparity with humans. The pig is a natural and important host of influenza viruses and is physiologically more comparable to humans than other animal models in terms of size, respiratory tract biology and volume. It may also be an important vector in the birds to human infection cycle. A major drawback of the current pig model is the inability to analyze antigen-specific CD8+ T-cell responses, which are critical to respiratory immunity. Here we address this knowledge gap using an established in-bred pig model with a high degree of genetic identity between individuals, including the MHC (Swine Leukocyte Antigen (SLA)) locus. We developed a toolset that included long-term in vitro pig T-cell culture and cloning and identification of novel immunodominant influenza-derived T-cell epitopes. We also generated structures of the two SLA class I molecules found in these animals presenting the immunodominant epitopes. These structures allowed definition of the primary anchor points for epitopes in the SLA binding groove and established SLA binding motifs that were used to successfully predict other influenza-derived peptide sequences capable of stimulating T-cells. Peptide-SLA tetramers were constructed and used to track influenza-specific T-cells ex vivo in blood, the lungs and draining lymph nodes. Aerosol immunization with attenuated single cycle influenza viruses (S-FLU) induced large numbers of CD8+ T-cells specific for conserved NP peptides in the respiratory tract. Collectively, these data substantially increase the utility of pigs as an effective model for studying protective local cellular immunity against respiratory pathogens.

We identify amino acid variants within dominant SARS-CoV-2 T cell epitopes by interrogating global sequence data. Several variants within nucleocapsid and ORF3a epitopes have arisen independently in multiple lineages and result in loss of recognition by epitope-specific T cells assessed by IFN-g and cytotoxic killing assays. Complete loss of T cell responsiveness was seen due to Q213K in the A*01:01-restricted CD8+ ORF3a epitope FTSDYYQLY 207-215 ; due to P13L, P13S, and P13T in the B*27:05-restricted CD8+ nucleocapsid epitope QRNAP-RITF 9-17 ; and due to T362I and P365S in the A*03:01/A*11:01-restricted CD8+ nucleocapsid epitope KTFPPTEPK 361-369 . CD8+ T cell lines unable to recognize variant epitopes have diverse T cell receptor repertoires. These data demonstrate the potential for T cell evasion and highlight the need for ongoing surveillance for variants capable of escaping T cell as well as humoral immunity.

Professor Satu Mustjoki has received honoraria and research funding from Novartis, Pfizer and Bristol-Myers Squibb (not related to this study). Professor Vincenzo Cerullo is cofounder and shareholder of the Valo Therapeutics LTD (not related to this study). All other named authors have no conflict of interest, financial or otherwise. Synopsis:Molecular mimicry can induce autoimmunity. By developing and using a bioinformatic tool to analyze molecular mimicry between tumor and viral antigens, the authors show this phenomenon can also play a role in antitumor immune responses.