Campylobacter jejuni ST353 and ST464 cause localised gut inflammation, crypt damage and 1 extraintestinal spread during large- and small-scale infection in broiler chickens.

Campylobacter infections in humans and chickens are a significant burden to health services 48 and the poultry industry. In the UK over 75% of chicken products are Campylobacter-positive 49 at retail but the knowledge of the mechanisms responsible for extraintestinal spread into 50 edible tissues remains incomplete. This work aimed to establish if two chicken-associated 51 lineages of C. jejuni, ST353 and ST464, have the potential for extraintestinal spread. Large- 52 and small-scale chicken colonisation trials investigated the infection biology of C. jejuni ST353 53 (3 strains) and ST464 (4 strains). Both lineages strongly colonised the ileum and caeca and 54 were detected in liver and spleen. C. jejuni ST353 and ST464 spleen load were significantly 55 increased compared to C. jejuni M1 controls. Immune responses in caecal tonsils exhibited 56 early induction of IFN-γ and suppressed TGFβ at 7dpi with C. jejuni ST464. Histochemistry of 57 gut tissue demonstrated significant decreases in intestinal crypt depth in ileal tissue with 58 increasing severity relative to Campylobacter lineage, M1<ST353<ST464. Pairwise correlation 59 analysis confirmed strong interdependencies between ‘caecal Campylobacter load’, ‘CXCLi1’, 60 ‘CXCLi2’ and ‘splenic Campylobacter load’. Furthermore, linear discriminant analysis (LDA) 61 confirmed that caecal tonsil derived IFNγ, TGFβ and CXCLi1 could predict splenic invasion at 62 71% accuracy. This work demonstrates that two chicken specialist C. jejuni lineages, ST353 63 and ST464, cause extraintestinal spread to liver and spleen and modelling suggests distinct 64 routes from ileum and caecum respectively. Recognition of these two routes of 65 Campylobacter extraintestinal spread (ileal/liver and caecal/spleen) provides better 66 understanding of this food derived pathogen for academia and the industry.