Campylobacter jejuni genes Cj1492c and Cj1507c are involved in host cell adhesion and invasion

Background 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. 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. Results In 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. Conclusion We 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

The transcriptional response of Arcobacter butzleri to cold shock

Arcobacter (A.) butzleri is an emerging zoonotic pathogen associated with gastrointestinal diseases, such as abdominal cramps and diarrhea, and is widely detected in animals, showing a high prevalence in poultry and seafood. The survival and adaptation of A. butzleri to cold temperatures remains poorly studied, although it might be of interest for food safety considerations. To address this, growth patterns of eight A. butzleri isolates were determined at 8 °C for 28 days. A. butzleri isolates showed strain‐dependent behavior: six isolates were unculturable after day 18, one exhibited declining but detectable cell counts until day 28 and one grew to the stationary phase level. Out of 13 A. butzleri cold shock‐related genes homologous to Escherichia coli, 10 were up‐regulated in response to a temperature downshift to 8 °C, as demonstrated by reverse transcription‐quantitative PCR. Additionally, we compared these data with the cold‐shock response in E. coli. Overall, we provide a deeper insight into the environmental adaptation capacities of A. butzleri, which we find shares similarities with the E. coli cold‐shock response

Editorial: Campylobacter-associated food safety

Introduction: Campylobacteriosis is an enteric bacterial zoonotic infection caused by members of the Campylobacter genus (Kirkpatrick and Tribble, 2011). C. jejuni (> 85%) and C. coli (5–10%) are the most common species associated with the disease (Patrick et al., 2018). Ingestion of as few as 500 bacteria can cause campylobacteriosis (Robinson, 1981). Although Campylobacter typically causes self-limiting human gastroenteritis, it can lead to prolonged post-infectious complications, such as Guillain-Barré syndrome, reactive arthritis, and/or post infectious-irritable bowel syndrome (Rees et al., 1995). The treatment of campylobacteriosis poses significant economic burdens worldwide, resulting in $1.56 billion in healthcare costs in the USA,$80 million in Canada, and €2.4 billion in the European Union per year (Devleesschauwer et al., 2017). The high prevalence of Campylobacter in the agri-food system is likely a major contributing factor to the incidence of campylobacteriosis. Due to its microaerobic nature, Campylobacter can colonize the intestinal tract of food-producing animals such as poultry, cattle, sheep, and swine (Hansson et al., 2018). However, it can also survive under aerobic conditions and infect humans through the food supply chain by forming biofilms or entering the viable but non-culturable state (Lv et al., 2019; Ma et al., 2022). The main route of infection has been identified as the consumption of contaminated food commodities, such as unpasteurized dairy products, undercooked poultry meat and/or contaminated water (Silva et al., 2011). Therefore, detection, characterization, and reduction of Campylobacter in the agroecosystem are of great importance. This mini-review provides an overview of the current trends in understanding Campylobacter and its interaction with the agroecosystem. We first introduce the improved methods to detect Campylobacter in various agri-food settings. Then, the prevalence of this microbe in the agri-food system as well as its characteristics are summarized. Finally, novel control strategies of Campylobacter are summarized and discussed

Lessons from a Meta-Analysis of Murine Infection Studies

Background: Only limited information is available about the immunopathogenic properties of Arcobacter infection in vivo. Therefore, we performed a meta- analysis of published data in murine infection models to compare the pathogenic potential of Arcobacter butzleri with Campylobacter jejuni and commensal Escherichia coli as pathogenic and harmless reference bacteria, respectively. Methodology / Principal Findings: Gnotobiotic IL-10-/- mice generated by broad-spectrum antibiotic compounds were perorally infected with A. butzleri (strains CCUG 30485 or C1), C. jejuni (strain 81-176) or a commensal intestinal E. coli strain. Either strain stably colonized the murine intestines upon infection. At day 6 postinfection (p.i.), C. jejuni infected mice only displayed severe clinical sequelae such as wasting bloody diarrhea. Gross disease was accompanied by increased numbers of colonic apoptotic cells and distinct immune cell populations including macrophages and monocytes, T and B cells as well as regulatory T cells upon pathogenic infection. Whereas A. butzleri and E. coli infected mice were clinically unaffected, respective colonic immune cell numbers increased in the former, but not in the latter, and more distinctly upon A. butzleri strain CCUG 30485 as compared to C1 strain infection. Both, A. butzleri and C. jejuni induced increased secretion of pro-inflammatory cytokines such as IFN-γ, TNF, IL-6 and MCP-1 in large, but also small intestines. Remarkably, even though viable bacteria did not translocate from the intestines to extra-intestinal compartments, systemic immune responses were induced in C. jejuni, but also A. butzleri infected mice as indicated by increased respective pro-inflammatory cytokine concentrations in serum samples at day 6 p.i. Conclusion / Significance: A. butzleri induce less distinct pro-inflammatory sequelae as compared to C. jejuni, but more pronounced local and systemic immune responses than commensal E. coli in a strain-dependent manner. Hence, data point towards that A. butzleri is more than a commensal in vertebrate hosts

Prevalence, antimicrobial susceptibility and virulence gene profiles of Arcobacter species isolated from human stool samples, foods of animal origin, ready-to-eat salad mixes and environmental water

Background Members of the genus Arcobacter are considered as emerging zoonotic food and waterborne pathogens that cause gastroenteritis and bacteremia in humans. However, the potential risk that Arcobacter species pose to public health remains unassessed in various countries, including Baltic states. Therefore, the aim of this study was to determine the prevalence, antimicrobial susceptibility and presence of putative virulence genes of Arcobacter isolates recovered from humans, food products and environmental water in Lithuania. Results A total of 1862 samples were collected and examined from 2018 to 2020 in the city of Kaunas. Overall, 11.2% (n = 208) of the samples were positive for the presence of Arcobacter spp. The highest prevalence was detected in chicken meat (36%), followed by environmental water (28.1%), raw cow milk (25%), ready-to-eat salad mixes (7.1%) and human stool (1.7%). A. butzleri was the most frequently isolated species (n = 192; 92.3%), followed by A. cryaerophilus (n = 16; 7.7%). Arcobacter spp. antimicrobial susceptibility testing revealed unimodally distributed aggregated minimal inhibitory concentrations (MICs) for gentamicin, tetracycline, ciprofloxacin, ampicillin and erythromycin. However, a bimodal distribution for azithromycin was found with 96.2% of determined MICs above the epidemiological cut-off value (ECOFF) defined for Campylobacter jejuni (0.25 µg/ml). Majority of the Arcobacter isolates (n = 187; 89.9%) showed high susceptibility to ciprofloxacin with MICs below or equal to the ECOFF value of 0.5 µg/ml. The putative virulence genes cadF (100%), ciaB (100%), cj1349 (99%), tlyA (99%), mviN (97.9%) and pldA (95.8%) were the predominant genes detected among A. butzleri isolates. In contrast, the mviN and ciaB genes were present in all, whereas cj1349 (12.5%), tlyA (25%) and hecA (12.5%) were only detected in few A. cryaerophilus isolates. Conclusions Our results demonstrate that food products and environmental water in Lithuania are frequently contaminated with Arcobacter spp. that carry multiple putative virulence genes. Furthermore, A. butzleri were isolated from 1.7% of inpatients. Fluoroquinolones and aminoglycosides were found to be more effective against Arcobacter in comparison to other antimicrobial agents. However, further studies are needed to determine the pathogenic mechanisms and factors that facilitate the spread of Arcobacter infections

Intestinal Barrier in Post-Campylobacter jejuni Irritable Bowel Syndrome

Background: Campylobacter jejuni (C. jejuni) is one of the most common causes of bacterial gastroenteritis worldwide. One sequela of this infection is the development of post-infectious irritable bowel syndrome (PI-IBS). It has been suggested that a dysfunctional intestinal barrier may promote IBS development. We aimed to test this hypothesis against the background of the leaky gut concept for low-grade inflammation in PI-IBS. Methods: We identified patients with persistent PI-IBS symptoms after C. jejuni infection. During sigmoidoscopy, forceps biopsies were obtained for electrophysiological measurements of epithelial transport and barrier function in miniaturized Ussing devices. C. jejuni absence was checked by PCR and cytokine production with immunohistochemistry. Results: In PI-IBS, the epithelial resistance of the colon epithelium was unaltered, reflecting an intact paracellular pathway. In contrast, temperature-dependent horseradish peroxidase (HRP, 44 kDa) permeation increased. Short-circuit current (Isc) reflecting active anion secretion and ENaC-dependent electrogenic sodium absorption was unaffected. Early endosome antigen-1 (EEA1) and IL-4 levels increased. C. jejuni is not incorporated into the resident microbiota of the colon mucosa in PI-IBS. Conclusions: In PI-IBS after C. jejuni infection, macromolecule uptake via endocytosis was enhanced, leading to low-grade inflammation with pro-inflammatory cytokine release. The findings will allow C. jejuni-induced pathomechanisms to be targeted during infection and, thereafter to reduce sequelae such as PI-IBS

The glycosyltransferase ST3GAL2 is regulated by miR-615-3p in the intestinal tract of Campylobacter jejuni infected mice

Background Campylobacter jejuni (C. jejuni) infections are of increasing importance worldwide. As a typical mucosal pathogen, the interaction of C. jejuni with mucins is a prominent step in the colonisation of mucosal surfaces. Despite recent advances in understanding the interaction between bacterial pathogens and host mucins, the mechanisms of mucin glycosylation during intestinal C. jejuni infection remain largely unclear. This prompted us to identify relevant regulatory networks that are concerted by miRNAs and could play a role in the mucin modification and interaction. Results We firstly used a human intestinal in vitro model, in which we observed altered transcription of MUC2 and TFF3 upon C. jejuni NCTC 11168 infection. Using a combined approach consisting of in silico analysis together with in vitro expression analysis, we identified the conserved miRNAs miR-125a-5p and miR-615-3p associated with MUC2 and TFF3. Further pathway analyses showed that both miRNAs appear to regulate glycosyltransferases, which are related to the KEGG pathway ‘Mucin type O-glycan biosynthesis’. To validate the proposed interactions, we applied an in vivo approach utilising a well-established secondary abiotic IL-10−/− mouse model for infection with C. jejuni 81-176. In colonic tissue samples, we confirmed infection-dependent aberrant transcription of MUC2 and TFF3. Moreover, two predicted glycosyltransferases, the sialyltransferases ST3GAL1 and ST3GAL2, exhibited inversely correlated transcriptional levels compared to the expression of the identified miRNAs miR-125a-5p and miR-615-3p, respectively. In this study, we mainly focused on the interaction between miR-615-3p and ST3GAL2 and were able to demonstrate their molecular interaction using luciferase reporter assays and RNAi. Detection of ST3GAL2 in murine colonic tissue by immunofluorescence demonstrated reduced intensity after C. jejuni 81-176 infection and was thus consistent with the observations made above. Conclusions We report here for the first time the regulation of glycosyltransferases by miRNAs during murine infection with C. jejuni 81-176. Our data suggest that mucin type O-glycan biosynthesis is concerted by the interplay of miRNAs and glycosyltransferases, which could determine the shape of intestinal glycosylated proteins during infection