Abstract
BackgroundCampylobacter jejuni (C. jejuni) has been assigned as an important food-borne pathogen for human health but many pathogenicity factors of C. jejuni and human host cell responses related to the infection have not yet been adequately clarified. This study aimed to determine further C. jejuni pathogenicity factors and virulence genes based on a random mutagenesis approach. A transposon mutant library of C. jejuni NCTC 11168 was constructed and the ability of individual mutants to adhere to and invade human intestinal epithelial cells was evaluated compared to the wild type. We identified two mutants of C. jejuni possessing altered phenotypes with transposon insertions in the genes Cj1492c and Cj1507c. Cj1492c is annotated as a two-component sensor and Cj1507c is described as a regulatory protein. However, functions of both mutated genes are not clarified so far.ResultsIn comparison to the wild type, Cj::1492c and Cj::1507c showed around 70–80% relative motility and Cj::1492c had around 3-times enhanced adhesion and invasion rates whereas Cj::1507c had significantly impaired adhesive and invasive capability. Moreover, Cj::1492c had a longer lag phase and slower growth rate while Cj::1507c showed similar growth compared to the wild type. Between 5 and 24 h post infection, more than 60% of the intracellular wild type C. jejuni were eliminated in HT-29/B6 cells, however, significantly fewer mutants were able to survive intracellularly. Nevertheless, no difference in host cell viability and induction of the pro-inflammatory chemokine IL-8 were determined between both mutants and the wild type.ConclusionWe conclude that genes regulated by Cj1507c have an impact on efficient adhesion, invasion and intracellular survival of C. jejuni in HT-29/B6 cells. Furthermore, potential signal sensing by Cj1492c seems to lead to limiting attachment and hence internalisation of C. jejuni. However, as the intracellular survival capacities are reduced, we suggest that signal sensing by Cj1492c impacts several processes related to pathogenicity of C. jejuni.
Highlights
Campylobacter jejuni (C. jejuni) has been assigned as an important food-borne pathogen for human health but many pathogenicity factors of C. jejuni and human host cell responses related to the infection have not yet been adequately clarified
Transposon mutagenesis of C. jejuni NCTC11168 In this study, the HyperMu Transposon was randomly inserted into the DNA of C. jejuni NCTC 11168 using a 2 h in vitro reaction catalysed by HyperMu MuA Transposase and subsequently transformed into C. jejuni NCTC 11168 by natural competence
To identify genes involved in C. jejuni pathogenicity, the obtained mutants were subsequently screened using invasion assays with HT-29/B6 cells
Summary
Campylobacter jejuni (C. jejuni) has been assigned as an important food-borne pathogen for human health but many pathogenicity factors of C. jejuni and human host cell responses related to the infection have not yet been adequately clarified. Cj1492c is annotated as a two-component sensor and Cj1507c is described as a regulatory protein Functions of both mutated genes are not clarified so far. Extensive studies have investigated that C. jejuni possesses numerous pathogenicity-associated factors involved in bacterial colonisation, transmigration and intracellular survival in intestinal epithelial cells [7, 8]. The paracellular pathway is characterised by breaking the tight junction and adherens junction complexes and crossing epithelial barriers by passage between neighbouring cells [7]. It has been demonstrated recently, that the C. jejuni protease HtrA is involved in breaching the epithelial barrier and enables invasion of epithelial cells via the paracellular pathway [9]
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