Jejuni Infected Research Articles

Disease alleviating effects following prophylactic lemon and coriander essential oil treatment in mice with acute campylobacteriosis.

Given the worldwide increasing prevalence of human Campylobacter jejuni infections and the emergence of multi-drug resistant enteropathogenic strains, antibiotic-independent approaches applying non-toxic natural compounds for the treatment and prophylaxis of campylobacteriosis appear utmost desirable. In our placebo-controlled intervention study, we surveyed potential disease-alleviating including anti-pathogenic and immune-modulatory effects upon prophylactic oral application of lemon-essential oil (LEM-EO) and coriander-essential oil (COR-EO) in acute experimental campylobacteriosis. Therefore, secondary abiotic IL-10-/- mice were orally challenged with either LEM-EO or COR-EO starting seven days prior to peroral C. jejuni infection. Six days post-infection, slightly lower pathogen loads were assessed in the colon of mice from the LEM-EO as opposed to the COR-EO cohort if compared to placebo counterparts. Prophylactic application of both EOs improved the clinical outcome of acute campylobacteriosis which was paralleled by less distinct pathogen-induced colonic epithelial cell apoptosis. Moreover, mice subjected to LEM-EO and COR-EO prophylaxis displayed lower colonic numbers of macrophages/monocytes and of T lymphocytes, respectively, whereas in both verum groups, basal IL-6 and IFN-γ concentrations were measured in mesenteric lymph nodes on day 6 post-infection. The oral challenge with either EOs resulted in diminished secretion of distinct pro-inflammatory mediators in the kidney as well as serum samples derived from the infected mice. In conclusion, the results from our preclinical in vivo study provide evidence that LEM-EO and COR-EO constitute promising prophylactic measures to prevent severe campylobacteriosis which may help to reduce the risk for development of post-infectious sequelae in C. jejuni infected individuals.

Combination of organic acids benzoate, butyrate, caprylate, and sorbate provides a novel antibiotics-independent treatment option in the combat of acute campylobacteriosis.

The food-borne Gram-negative bacterial pathogen Campylobacter jejuni may cause the acute enterocolitis syndrome campylobacteriosis in infected humans. Given that human C. jejuni infections are rising globally which hold also true for resistance rates against antibiotic compounds such as macrolides and fluoroquinolones frequently prescribed for the treatment of severe infectious enteritis, novel antibiotics-independent therapeutic strategies are needed. Distinct organic acids are well known for their health-beneficial including anti-microbial and immunomodulatory properties. In our present study, we investigated potential pathogen-lowering and anti-inflammatory effects of benzoic acid, butyric acid, caprylic acid, and sorbic acid either alone or in combination during acute murine campylobacteriosis. Therefore, secondary abiotic IL-10-/- mice were perorally infected with C. jejuni strain 81-176 and subjected to a 4-day-course of respective organic acid treatment. On day 6 post-infection, mice from the combination cohort displayed slightly lower pathogen loads in the duodenum, but neither in the stomach, ileum nor large intestine. Remarkably, the clinical outcome of C. jejuni induced acute enterocolitis was significantly improved after combined organic acid treatment when compared to the placebo control group. In support, the combinatory organic acid treatment dampened both, macroscopic and microscopic inflammatory sequelae of C. jejuni infection as indicated by less colonic shrinkage and less pronounced histopathological including apoptotic epithelial cell changes in the colon on day 6 post-infection. Furthermore, mice from the combination as compared to placebo cohort exhibited lower numbers of innate and adaptive immune cells such as neutrophilic granulocytes, macrophages, monocytes, and T lymphocytes in their colonic mucosa and lamina propria, respectively, which also held true for pro-inflammatory cytokine secretion in the large intestines and mesenteric lymph nodes. Notably, the anti-inflammatory effects were not restricted to the intestinal tract, but could also be observed systemically given pro-inflammatory mediator concentrations in C. jejuni infected mice from the combination organic acid treatment that were comparable to basal values. In conclusion, our in vivo study provides first evidence that an oral application of distinct organic acids in combination exhibits pronounced anti-inflammatory effects and hence, constitutes a promising novel antibiotics-independent therapeutic strategy in the combat of acute campylobacteriosis.

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

BackgroundCampylobacter 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.ResultsWe 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.ConclusionsWe 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.

IL-23 Contributes to Campylobacter jejuni-Induced Intestinal Pathology via Promoting IL-17 and IFNγ Responses by Innate Lymphoid Cells.

Human pathogen Campylobacter jejuni is a significant risk factor for the development of long-term intestinal dysfunction although the cellular and molecular mechanisms remain scantily defined. IL-23 is an emerging therapeutic target for the treatment of inflammatory intestinal diseases, however its role in C. jejuni-driven intestinal pathology is not fully understood. IL-10 deficient mice represent a robust model to study the pathogenesis of C. jejuni infection because C. jejuni infection of mice lacking IL-10 results in symptoms and pathology that resemble human campylobacteriosis. To determine the role of IL-23 in C. jejuni-driven intestinal inflammation, we studied the disease pathogenesis in IL-23-/- mice with inhibited IL-10Rα signaling. These mice exhibited reduced intestinal pathology independent from bacterial clearance. Further, levels of IFNγ, IL-17, IL-22, TNF, and IL-6 were reduced and associated with reduced accumulation of neutrophils, monocytes and macrophages in the colon. Flow cytometry analysis revealed reduced production of IL-17 and IFNγ by group 1 and 3 innate lymphoid cells. Thus, our data suggest that IL-23 contributes to intestinal inflammation in C. jejuni infected mice by promoting IL-17 and IFNγ production by innate lymphoid cells.

Resveratrol Alleviates Acute Campylobacter jejuni Induced Enterocolitis in a Preclinical Murine Intervention Study.

The polyphenolic compound resveratrol has been shown to exert health-beneficial properties. Given globally emerging Campylobacter infections in humans, we addressed potential anti-pathogenic, immuno-modulatory and intestinal epithelial barrier preserving properties of synthetic resveratrol in the present preclinical intervention study applying a murine acute campylobacteriosis model. Two days following peroral C. jejuni infection, secondary abiotic IL-10−/− mice were either subjected to resveratrol or placebo via the drinking water. Whereas placebo mice suffered from acute enterocolitis at day 6 post-infection, resveratrol treatment did not only lead to improved clinical conditions, but also to less pronounced colonic epithelial apoptosis as compared to placebo application. Furthermore, C. jejuni induced innate and adaptive immune cell responses were dampened in the large intestines upon resveratrol challenge and accompanied by less colonic nitric oxide secretion in the resveratrol versus the placebo cohort. Functional analyses revealed that resveratrol treatment could effectively rescue colonic epithelial barrier function in C. jejuni infected mice. Strikingly, the disease-alleviating effects of resveratrol could additionally be found in extra-intestinal and also systemic compartments at day 6 post-infection. For the first time, our current preclinical intervention study provides evidence that peroral resveratrol treatment exerts potent disease-alleviating effects during acute experimental campylobacteriosis.

Murine Fecal Microbiota Transplantation Alleviates Intestinal and Systemic Immune Responses in Campylobacter jejuni Infected Mice Harboring a Human Gut Microbiota.

Human campylobacteriosis constitutes a zoonotic food-borne disease and a progressively rising health burden of significant socioeconomic impact. We have recently shown that conventional mice are protected from Campylobacter jejuni infection, which was not the case for human microbiota associated (hma) mice indicating that the host-specific gut microbiota composition primarily determines susceptibility to or resistance against C. jejuni infection. In our present preclinical intervention study we addressed whether gut microbiota changes in stably C. jejuni infected hma mice following murine fecal microbiota transplantation (mFMT) could alleviate pathogen-induced immune responses. To accomplish this, secondary abiotic C57BL/6 mice were generated by broad-spectrum antibiotic treatment, perorally reassociated with a complex human gut microbiota and challenged with C. jejuni by gavage. Seven days later C. jejuni infected hma mice were subjected to peroral mFMT on 3 consecutive days. Within a week post-mFMT fecal pathogenic burdens had decreased by two orders of magnitude, whereas distinct changes in the gut microbiota composition with elevated numbers of lactobacilli and bifidobacteria could be assessed. In addition, mFMT resulted in less C. jejuni induced apoptotic responses in colonic epithelia, reduced numbers of macrophages and monocytes as well as of T lymphocytes in the large intestinal mucosa and lamina propria and in less distinct intestinal pro-inflammatory cytokine secretion as compared to mock challenge. Strikingly, inflammation dampening effects of mFMT were not restricted to the intestinal tract, but could also be observed systemically as indicated by elevated serum concentrations of pro-inflammatory cytokines such as TNF-α, IL-12p70, and IL-6 in C. jejuni infected hma mice of the mock, but not the mFMT cohort. In conclusion, our preclinical mFMT intervention study provides evidence that changes in the gut microbiota composition which might be achieved by pre- or probiotic formulations may effectively lower intestinal C. jejuni loads, dampen both, pathogen-induced intestinal and systemic inflammatory sequelae and may represent a useful tool to treat continuous shedding of C. jejuni by asymptomatic carriers which is critical in the context of food production, hospitalization and immunosuppression.

Vitamin D in Acute Campylobacteriosis-Results From an Intervention Study Applying a Clinical Campylobacter jejuni Induced Enterocolitis Model.

Human Campylobacter infections are progressively rising and of high socioeconomic impact. In the present preclinical intervention study we investigated anti-pathogenic, immuno-modulatory, and intestinal epithelial barrier preserving properties of vitamin D applying an acute campylobacteriosis model. Therefore, secondary abiotic IL-10−/− mice were perorally treated with synthetic 25-OH-cholecalciferol starting 4 days before peroral Campylobacter jejuni infection. Whereas, 25-OH-cholecalciferol application did not affect gastrointestinal pathogen loads, 25-OH-cholecalciferol treated mice suffered less frequently from diarrhea in the midst of infection as compared to placebo control mice. Moreover, 25-OH-cholecalciferol application dampened C. jejuni induced apoptotic cell responses in colonic epithelia and promoted cell-regenerative measures. At day 6 post-infection, 25-OH-cholecalciferol treated mice displayed lower numbers of colonic innate and adaptive immune cell populations as compared to placebo controls that were accompanied by lower intestinal concentrations of pro-inflammatory mediators including IL-6, MCP1, and IFN-γ. Remarkably, as compared to placebo application synthetic 25-OH-cholecalciferol treatment of C. jejuni infected mice resulted in lower cumulative translocation rates of viable pathogens from the inflamed intestines to extra-intestinal including systemic compartments such as the kidneys and spleen, respectively, which was accompanied by less compromised colonic epithelial barrier function in the 25-OH-cholecalciferol as compared to the placebo cohort. In conclusion, our preclinical intervention study provides evidence that peroral synthetic 25-OH-cholecalciferol application exerts inflammation-dampening effects during acute campylobacteriosis.

Immune responses upon Campylobacter jejuni infection of secondary abiotic mice lacking nucleotide-oligomerization-domain-2

BackgroundCampylobacter jejuni infections are of rising importance worldwide. Given that innate immune receptors including nucleotide-oligomerization-domain-2 (Nod2) are essentially involved in combating enteropathogenic infections, we here surveyed the impact of Nod2 in murine campylobacteriosis.Methods and resultsIn order to overcome physiological colonization resistance preventing from C. jejuni infection, we generated secondary abiotic Nod2−/− and wildtype (WT) mice by broad-spectrum antibiotic treatment. Mice were then perorally infected with C. jejuni strain 81-176 on 2 consecutive days and could be stably colonized by the pathogen at high loads. Notably, Nod2 deficiency did not affect gastrointestinal colonization properties of C. jejuni. Despite high intestinal pathogenic burdens mice were virtually uncompromised and exhibited fecal blood in single cases only. At day 7 postinfection (p.i.) similar increases in numbers of colonic epithelial apoptotic cells could be observed in mice of either genotype, whereas C. jejuni infected Nod2−/− mice displayed more distinct regenerative properties in the colon than WT controls. C. jejuni infection was accompanied by increases in distinct immune cell populations such as T lymphocytes and regulatory T cells in mice of either genotype. Increases in T lymphocytes, however, were less pronounced in large intestines of Nod2−/− mice at day 7 p.i. when compared to WT mice, whereas colonic numbers of B lymphocytes were elevated in WT controls only upon C. jejuni infection. At day 7 p.i., colonic pro-inflammatory mediators including nitric oxide, TNF, IFN-γ and IL-22 increased more distinctly in Nod2−/− as compared to WT mice, whereas C. jejuni induced IL-23p19 and IL-18 levels were lower in the large intestines of the former. Converse to the colon, however, ileal concentrations of nitric oxide, TNF, IFN-γ, IL-6 and IL-10 were lower in Nod2−/− as compared to WT mice at day 7 p.i. Even though MUC2 was down-regulated in C. jejuni infected Nod2−/− mice, this did not result in increased pathogenic translocation from the intestinal tract to extra-intestinal compartments.ConclusionIn secondary abiotic mice, Nod2 signaling is involved in the orchestrated host immune responses upon C. jejuni infection, but does not control pathogen loads in the gastrointestinal tract.

Amelioration of intestinal and systemic sequelae of murine Campylobacter jejuni infection by probiotic VSL#3 treatment

BackgroundThe incidence of human Campylobacter jejuni infections is progressively increasing worldwide. Probiotic compounds might open up valuable tools to decrease pathogen burden and subsequent pro-inflammatory immune responses, but in vivo data are scarce.Methods and resultsSecondary abiotic mice generated by broad-spectrum antibiotic treatment were perorally challenged with the commercial probiotic compound VSL#3 consisting of Streptococcus thermophilus, Bifidobacterium breve, Bifidobacterium longum, Bifidobacterium infantis, Lactobacillus acidophilus, Lactobacillus plantarum, Lactobacillus paracasei, and Lactobacillus delbrueckii ssp. bulgaricus) either 5 days before (i.e. prophylactic regimen) or after (i.e. therapeutic regimen) peroral C. jejuni strain 81–176 infection, and analyzed 3 weeks following the initial bacterial re-association. Upon challenge, mice were colonized with the probiotic bacteria and/or C. jejuni at comparable intestinal loads, but co-colonization did not result in reduction of the pathogen burden. Remarkably, prophylactic as well as therapeutic VSL#3 treatment of C. jejuni infected mice ameliorated intestinal apoptosis and pro-inflammatory immune responses as indicated by lower numbers of innate and adaptive immune cell populations in the murine colon upon probiotic prophylaxis or treatment and reduced colonic concentrations of pro-inflammatory mediators including IL-6 and MCP-1. Importantly, concentrations of anti-inflammatory mediators such as IL-10 were significantly elevated in the colon of probiotics treated mice as compared to untreated controls. Strikingly, prophylactic VSL#3 treatment attenuated C. jejuni induced systemic pro-inflammatory responses as indicated by less TNF and IL-12p70 secretion in the spleen of VSL#3 pre-treated as compared to non-treated mice.ConclusionAdministration of probiotic formulations such as VSL#3 might open up valuable strategies for prophylaxis and/or treatment of C. jejuni induced intestinal and systemic sequelae in vivo by the suppression of pro-inflammatory and induction of anti-inflammatory responses.

Campylobacter jejuni infection of conventionally colonized mice lacking nucleotide-oligomerization-domain-2

BackgroundThe nucleotide-binding oligomerisaton protein 2 (NOD2) constitutes a pivotal sensor of bacterial muramyl dipeptide and assures expression of distinct antimicrobial peptides and mediators produced by enterocytes and immune cells directed against pathogens including Campylobacter jejuni. We here elucidated the role of NOD2 during murine C. jejuni infection in more detail.ResultsConventionally colonized NOD2 deficient (NOD2−/−) mice and corresponding wildtype (WT) counterparts were perorally infected with C. jejuni strain 81–176 on three consecutive days. The pathogen colonized both WT and NOD2−/− mice only sporadically until day 14 post infection (p.i.). However, the slightly higher prevalence of C. jejuni in NOD2−/− mice was accompanied by higher intestinal Escherichia coli loads known to facilitate C. jejuni colonization. Neither overt macroscopic (clinical) nor microscopic sequelae (such as colonic epithelial apoptosis) could be observed upon murine C. jejuni infection of either genotype. Innate immune responses were less distinctly induced in C. jejuni infected NOD2−/− versus WT mice as indicated by lower colonic numbers of neutrophils in the former. Conversely, adaptive immune cell counts including T lymphocytes were higher in large intestines of NOD2−/− as compared to WT mice that were paralleled by increased colonic IL-6 secretion and higher TNF and IL-18 mRNA expression levels in large intestines of the former. Only in NOD2−/− mice, however, colonic IL-22 mRNA expression was down-regulated at day 14 p.i. Whereas viable commensal intestinal bacteria could exclusively be detected in mesenteric lymph nodes and livers of NOD2−/− mice, bacterial translocation rates to kidneys and spleen were NOD2 independent. Notably, large intestinal mRNA expression levels of mucin-2, constituting a pivotal factor involved in epithelial barrier integrity, were comparable in naive and C. jejuni infected mice of either genotype.ConclusionNOD2 is involved in the well-balanced regulation of innate and adaptive pro-inflammatory immune responses of conventional mice upon C. jejuni infection.

Immunogens of interest for the diagnosis of Campylobacter jejuni infections

In order to identify the C. jejuni immunogens of interest for the diagnosis of Campylobacter infections, we analyzed the humoral response of 153 patients by using complement fixation (CF) and western blot assays. A first group of 79 sera was from C. jejuni infected patients suffering from enteritis ( n=16), Guillain-Barré syndrome (GBS) ( n=40) and arthritis ( n=23). A second group of 49 sera was from healthy blood donors and a third group consisted of 25 sera from children under 4 years old. Using the CF test, 88.6% of the C. jejuni infected patients were seropositive versus 28.5% of the healthy blood donors and none of the children. The Western blot assay allowed detection of antibodies directed against seven selected antigens ranging from 14 to 67 kDa. Three of these antigens with a molecular size of 29, 37 and 43 kDa were detected by 86.0%, 84.8% and 91.1% of the C. jejuni infected patients, respectively. These three antigens seem to be good candidates for the development of assays suitable for direct and indirect diagnosis of Campylobacter infections.

Calcium and protein kinase C play an important role in Campylobacter jejuni-induced changes in Na + and Cl − transport in rat ileum in vitro

The pathophysiological mechanism of Campylobacter jejuni (enterotoxigenic) induced secretory diarrhoea remains least understood. To investigate the mechanism(s) involved, the unidirectional fluxes of Na + and Cl − were measured across the C. jejuni live culture infected and control (non infected) rat ileum (unstriped), in vitro by Ussing technique under short circuit conditions, in the presence or absence of: Ca 2+ ionophore A23187 (5 μM), 1-verapamil (100 μM), calmodulin (CaM) antagonist W-7 (100 μM), dantrolene (25 μM), protein kinase C (PKC) activator PMA (100 ng/ml) and H-7 (60 μM), selective inhibitor of PKC. There was net absorption of Na + and enhanced Cl − secretion in infected animals while in control animals there was net absorption of Na + and marginal secretion Cl −. Ca 2+ ionophore A23187 mimicked the effects of C. jejuni infection whereas 1-verapamil had significant antisecretory effect on Na + and Cl − secretion in infected animals. In vitro measurement of unidirectional 45Ca fluxes in Ussing chamber experiments revealed net absorption of Ca 2+ in infected rat ileum as compared to net secretion of Ca 2+ in control rat ileum. These observations clearly indicate that there is increased stimulation of Ca 2+ uptake from extracellular milieu to the enterocytes during C. jejuni-induced diarrhoea. The intracellular calcium levels (Ca 2+] i (as measured by fluorescent probe Fura-2AM) were found to be raised significantly ( P < 0.001) in enterocytes isolated from C. jejuni infected ileum as compared to the enterocytes from control ileum. The observed increase in [Ca 2+] i in enterocytes isolated from C. jejuni live culture supernatant treated rat ileum further shows the involvement of enterotoxin in diarrhoeal process. Dantrolene decreased significantly C. jejuni-induced net Na + and Cl − secretion but it could not reverse it to absorption suggesting the partial involvement of Ca 2+ mobilised from intracellular stores in mediating secretion. W-7 failed to inhibit the C. jejuni-induced net Na + and Cl − secretion. In addition the CaM activity estimated in intestinal microvillar core remained same in both the control and C. jejuni infected animals. This indicates that C. jejuni-induced diarrhoea is not mediated through the activation of Ca 2+-CaM complex pathway of the Ca 2+ messenger system. The PKC activator PMA, induced net secretion of Na + and Cl − in the control animals but it could not enhance further the C. jejuni-induced Na + and Cl − secretion, suggesting that there is overlapping effect of PMA and C. jejuni live culture infection. H-7 significantly inhibited ( P < 0.001) net Na + and Cl − secretion in the infected animals but had no effect on basal rat ileal Na + and Cl − transport in control animals. This again suggest the activation of PKC signal in C. jejuni-induced ileal Na + and Cl − secretion. Although intracellular cAMP levels were found to be significantly raised in the enterocytes isolated from C. jejuni live culture infected and C. jejuni culture supernatant treated rat ileum, the mechanism of interaction between cAMP and Ca 2+ messenger system in infection remains undefined. Taken together, the present results therefore, indicate that enterotoxigenic C. jejuni live culture-induced Na + and Cl − secretion is a calcium-dependent process where both increased Ca 2+ entry from extracellular milieu and Ca 2+ mobilization from intracellular stores result in increased enterocyte (Ca 2+] i. The activation of PKC further appears to be the important intracellular mediator pathway involved in Ca 2+-dependent stimulation of C. jejuni-induced diarrhoea.