Pre-treatment of Chicken Epithelial Cells with Intracellular PRR Agonists Leads to Alterations in C. jejuni Strain Invasion and Cytokine Phenotype

Abstract P10

Presenter: Heather Chick (Swansea University)

Authors: Heather M. Chick (1), Lisa K. Williams (2), Bernd Kaspers (3) and Thomas S. Wilkinson (1)

This work is part of the BBSRC-funded CampAttack Consortium (2018-2021)

• 1 Swansea University, Swansea, Wales
• 2 Hartpury University, Hartpury, England
• 3 Ludwig-Maximilians-Universitaet, Muenchen, Germany

Extra-intestinal spread of C. jejuni in broiler chickens represents a significant public health threat. Some pathogenic C. jejuni strains possess the ability to invade intestinal epithelial cells, migrate across the gut wall and disseminate into edible tissues such as the muscle and the liver. There is good in vivo evidence in broiler chickens that C. jejuni-induced localised and systemic inflammation may be responsible for compromising epithelial cell barrier integrity and may be a pre-requisite for extra-intestinal spread. Understanding the specific inflammatory stimuli that link C. jejuni, the gut epithelium and inflammation is integral in designing intervention methods for Campylobacter control. Using cell culture, 8E11 avian intestinal epithelial cells were treated for 6-24 hours with various pattern recognition receptor (PRR) agonists or the cytokines IL-6, IFN- and IL-17 in order to induce defined local immune response. Epithelial cells were then either lysed and RNA extracted, or were treated with C. jejuni M1 (reference strain), C. jejuni LE55 (high invasion) or C. jejuni LE17 (low invasion) for 4 hours, before RNA was extracted or a gentamicin protection assay (GPA) performed. Quantitative RT-PCR was performed on RNA extracts to determine gene expression ratios of CXCLi1, CXCLi2, IL-10 and TNF-, and viable counts performed from GPA plates. Pre-treatment of 8E11 cells resulted in strain dependent alterations in the invasion patterns of C. jejuni; in particular, the low-invasion strain (LE17) showed significantly enhanced invasion in comparison with M1 control or invasive (LE55) strain when epithelial cells had been pre-treated with a NOD-2 agonist (MDP) or a TLR3 agonist (Poly I:C). Pre-treatment of chicken epithelial cells with Poly I:C, LPS, MDP and IFN- led to CXCLi2 production, an effect which was significantly enhanced up to 60-fold when IFN- and LPS where given in combination. Together, these results suggest that the type of T-helper lymphocyte derived inflammatory milieu generated promotes key differences in the invasive capabilities of certain strains of C. jejuni in vitro. These results are important because they suggest that pre-existing inflamed epithelium induced by intracellular PRRs and Th1 cytokines lead to enhanced C. jejuni invasion and CXCLi2 production. This suggests a strong link between inflammation and the ability of C. jejuni to invade gut cells and recruit blood derived phagocytes. Anti-inflammatory strategies must therefore focus on the type of inflammation generated in order to gain specific control of Campylobacter.

Presenting in Speaking session 2 - Pathogenesis