The association of Human Leukocyte Antigen (HLA) B27 with ankylosing spondylitis (AS) is one of the strongest links between possession of a number of closely related HLA alleles and an autoimmune disease. A prominent hypothesis for this association is the arthritogenic peptide theory, which invokes an argument that molecular mimicry between a foreign- and a self-peptide leads to the breakdown of immune tolerance and ensuing autoimmunity. A potential source of arthritogenic peptides comes from gastrointestinal bacteria, such as during Salmonella infection, which commonly precedes development of disease in a subgroup of AS patients. Another potential source of arthritogenic peptides is the newly described class of peptides, termed spliced peptides. These peptides form by fusion of two non-contiguous regions of the same or different proteins. The aim of this study is to investigate how the linear and spliced peptide pools are modulated post infection with Salmonella across the 8 most common HLA-B27 allotypes (HLA-B*27:02 – HLA-B*27:09). Methods: High resolution mass spectrometry combined with de novo sequencing were used to identify linear and spliced peptides across the 8 most common HLA-B27 in mock and Salmonella infected antigen presenting cells. Results: Our results showed that very low number of linear Salmonella-derived peptides are presented by the HLA-B27 allotypes studied. Interestingly, the number of spliced peptides derived from Salmonella antigens were higher than linear peptides, with some of the peptides forming human-Salmonella hybrids. We also observed that the consensus-binding motifs of the spliced peptides were significantly different to linear peptides, in particular the strong preference for arginine at the P2 anchor position was drastically reduced. Conclusion: The increased number and non-canonical motifs of spliced peptides could be potential sources of arthritogenic peptides and therefore could play a role in disease pathogenesis. However, further analyses such peptide binding studies, immunological assays and structural investigations are required to confirm immunogenicity of these peptides.