Exploring the Virulence Gene Expression of Arcobacter butzleri during Simulated Infection of Human Gut Models

Introduction: Arcobacter butzleri is an emerging foodborne pathogen often isolated from pork, chicken and beef meat, which causes different gastrointestinal diseases in human due to its invasive behaviour. However, the pathogenicity of A. butzleri is still underestimated due to a substantial lack of information on its virulence mechanisms, metabolic and genomic features. Purpose: This study aims to explore, in simulated host-pathogen interactions, the expression of nine genes that are currently correlated with virulence traits of A. butzleri. Methods: After the ex novo design of primers for the nine genes (cadF, ciaB, cj1349, irgA, hecA, hecB, mviN, pldA, tlyA) on an A. butzleri reference genome (type strain LMG 10828T), their relative expression was quantified by quantitative RT-PCR under simulated host-pathogen interaction conditions. Briefly, In Vitro gut models of mucus-producing (HT29–MTX) and non-producing (HT29, Caco-2) human cells were co-incubated with the pathogen and total bacterial RNA was recovered at different time points. At the same time, bacterial counts were performed to describe the colonization and translocation capabilities of A. butzleri, by using two dimensional and three dimensional gut models, respectively. Results: As the first outcome, an RT-qPCR protocol, suitable to quantify the relative expression of the nine virulence genes of A. butzleri in the presence of human cells, was optimized. Applying this protocol, an upregulation of part of those genes along the co-incubation time was observed. In addition, a favourable role of the mucus for the pathogen colonization was observed whereas the data from the three-dimensional models suggested an intracellular passage of A. butzleri through the epithelial barrier. Significance: The results of this study represent a first step in the understanding of A. butzleri pathogenicity and are important to explain its role in gastrointestinal diseases.