Effect of α-Hemolysin Producing <i>E. coli</i> in Two Different Mouse Strains in a DSS Model of Inflammatory Bowel Disease

Background: Phylogroup B2 Escherichia coli have been associated with ulcerative colitis (UC). In this study, we aimed to compare colonization with the UC-associated E. coli p19A in different mice strains, to investigate the role of alpha hemolysin in a UC mouse model. Methods: In this study, Sigirr &minus;/&minus; and C57BL/6 mice were chosen, and UC was induced by adding dextran sulfate sodium (DSS) to the drinking water. The mice were pre-treated with ciprofloxacin. p19A expressing luminescence and GFP, alpha-hemolysin knock out p19A-&Delta;hlyI II, and non-pathogenic lab E. coli DH10B were cultured in LB broth, and orally gavaged into the mice. Colonization with p19A WT was visualized using an in vivo imaging system. Results: p19A WT colonized the colon, ileum, Peyer&rsquo;s patches, liver, and spleen of infected C57BL/6 and Sigirr &minus;/&minus; mice. A total of 99% of the p19A WT infected C57BL/6 mice and 29% of the p19A WT infected Sigirr &minus;/&minus; mice survived to the 4th post infection day. Conclusion: UC-associated E. coli p19A WT colonized the intestines of DSS-treated mice and caused extra-intestinal infection. Hemolysin is an important factor in this pathogenesis, since isogenic hemolysin mutants did not cause the same inflammation

Extraintestinal pathogenic <i>Escherichia coli</i> are associated with intestinal inflammation in patients with ulcerative colitis

E. coli of the phylogenetic group B2 harbouring Extra intestinal Pathogenic Escherichia coli (ExPEC) genes are frequently seen as colonizers of the intestine in patients with active ulcerative colitis (UC). In this study, we describe the influence of E. coli Nissle (EcN) B2 as add-on treatment to conventional therapies in patients with active UC. For this study one hundred active UC patients were randomized to ciprofloxacin or placebo for 1 week followed by EcN or placebo for 7 weeks. Stool samples were collected at weeks 0, 1, 8, 12, where E. coli were characterized and fecal calprotectin was measured. We showed that in the active UC patient group receiving Placebo/EcN, fewer patients reached remission, in comparison to the patient group receiving Placebo/placebo (p < 0.05). Active UC patients initially colonized with E. coli B2 had increased fecal calprotectin values and Colitis Activity Index scores in comparison to patients colonized with E. coli A and D (p < 0.05*). In conclusion, treatment of UC patients with E. coli Nissle (B2) does not promote clinical remission and active UC patients colonized with E. coli B2 have an increased intestinal inflammation

Predictors of response and disease course in patients with inflammatory bowel disease treated with biological therapy-the Danish IBD Biobank Project:protocol for a multicentre prospective cohort study

IntroductionInflammatory bowel diseases (IBDs) are chronic diseases of unknown cause characterised by a progressive and unpredictable disease course. In the last decade, biological treatment has become a cornerstone in the treatment of IBD. However, one-in-three-to-four patients do not respond to first-line biological agents and another third of patients see their response diminish over time. This highlights an unmet need for optimising the use of biologicals and the prediction of treatment response. Considering the multifaceted nature of IBD, we hypothesise that multiomics profiling of sequential samples from single patients could facilitate the discovery of predictive biomarkers of response to biological therapy and disease course.MethodsThis is a multicentre prospective cohort study which will enrol 840 biological-naïve patients with IBD who initiate biological therapy in a 3-year period. Primary outcomes are the occurrence of primary non-response (evaluated at weeks 14–16) and loss of response (evaluated during entire follow-up in patients who obtain partial or full response after induction period). Each patient will be followed up for their clinical data for at least 1 year or till the end of study period (up to 4 years). Blood and stool samples will be collected sequentially during the first year of biological treatment. Intestinal tissue will be sampled after 1 year of treatment and whenever an endoscopy is performed. Samples will undergo transcriptomic, proteomic and microbial DNA analyses. Omics data will be integrated with clinical data to identify a panel of predictive biomarkers of response to biological therapy and disease behaviour in patients with IBD.Ethics and disseminationEthical approval has been obtained from the Danish Ethics Committee (H-18064178). Inclusion is ongoing at three study centres and will be initiated in two additional centres. Both positive and negative study results will be disseminated through peer-reviewed journals according to Strengthening the Reporting of Observational Studies in Epidemiology guidelines, as well as presented at international conferences

Substantial Intestinal Microbiota Differences Between Patients With Ulcerative Colitis From Ghana and Denmark:Ulcerative Colitis in Denmark vs Ghana

BACKGROUND: Ulcerative colitis (UC) is a relapsing nontransmural inflammatory disease that is restricted to the colon and is characterized by flare-ups of bloody diarrhea. In this study, we aimed to investigate intestinal bacterial diversity in healthy controls and patients with UC with and without active disease, from Ghana and Denmark. METHODS: The study included 18 UC patients (9 with active and 9 with inactive disease) and 18 healthy controls from Ghana. In addition 16 UC patients from Denmark (8 UC with active and 8 UC with inactive disease) and 19 healthy controls from Denmark. Microbiota diversity analysis relied on sequencing of ribosomal small subunit genes. Purified genomic DNA was submitted to PCR using a primer set targeting prokaryotes and eukaryotes. The purified DNA was sequenced on the Illumina MiSeq system in a 2 × 250 bp set up (Illumina, San Diego, CA, USA). Blinded analysis of the taxonomy table was performed using BioNumerics-7.5 (Applied Maths NV, Sint-Martens-Latem, Belgium). RESULTS: When analyzing the taxonomy data for prokaryotes, cluster and principal component analysis shows Danish healthy controls clustered together, but separate from healthy controls from Ghana, which also clustered together. The Shannon diversity index (SDI) for prokaryotes shows significant differences between Danish healthy controls and patients in comparison with the corresponding groups from Ghana (p = 0.0056). Significant increased abundance of Escherichia coli was detected in healthy controls from Ghana in comparison with healthy controls from Denmark. The SDI of the prokaryotes ranges between 0 and 3.1 in the Ghana study groups, while in the Danish study groups it ranges between 1.4 and 3.2, the difference is however not significant (p = 0.138). Our data show a significant increased abundance of eukaryotes species in the healthy control group from Ghana and Denmark in comparison with patient groups from Ghana and Denmark. CONCLUSION: Overall, healthy controls and patients with UC from Denmark have increased diversity of prokaryotes. Healthy controls from Denmark and Ghana have increased abundance of eukaryotes in comparison with UC patient groups from Denmark and Ghana

Fecal microbiota transplantation in the treatment of chronic pouchitis:A systematic review

The objective was to evaluate available literature on treatment of chronic pouchitis with fecal microbiota transplantation (FMT) focusing on clinical outcomes, safety, and different approaches to FMT preparation and delivery. A systematic review of electronic databases was conducted using Medline, EMBASE, and the Cochrane Central Register of Controlled Trials Library from inception through April 2020. Human studies of all study types reporting results of FMT to treat chronic pouchitis were included. Nine studies, reporting FMT treatment of 69 patients with chronic pouchitis were found eligible for the review. Most studies were case series and cohort studies rated as having fair to poor quality due to high risk of bias and small sample size. Only one randomized controlled trial was included, finding no beneficial effect of FMT. In total clinical response after FMT was reported in 14 (31.8%) out of 44 evaluated patients at various timepoints after FMT, and clinical remission in ten (22.7%) patients. Only minor self-limiting adverse events were reported. FMT varied greatly regarding preparation, length of treatment, and route of delivery. The effects of FMT on symptoms of chronic pouchitis are not established, though some studies show promising results. Future controlled well-designed studies are warranted

Secretion of alpha-hemolysin by<i> Escherichia coil</i> disrupts tight junctions in ulcerative colitis patients

OBJECTIVES: The potential of Escherichia coli (E. coli) isolated from inflammatory bowel disease (IBD) patients to damage the integrity of the intestinal epithelium was investigated. METHODS: E. coli strains isolated from patients with ulcerative colitis (UC) and healthy controls were tested for virulence capacity by molecular techniques and cytotoxic assays and transepithelial electric resistance (TER). E. coliisolate p19A was selected, and deletion mutants were created for alpha-hemolysin (α-hemolysin) (hly) clusters and cytotoxic necrotizing factor type 1 (cnf1). ProbioticE. coliNissle and pathogenicE. coliLF82 were used as controls. RESULTS: E. colistrains from patients with active UC completely disrupted epithelial cell tight junctions shortly after inoculation. These strains belong to phylogenetic group B2 and are all α-hemolysin positive. In contrast, probioticE. coliNissle, pathogenicE. coliLF82, fourE. colifrom patients with inactive UC and threeE. colistrains from healthy controls did not disrupt tight junctions.E. colip19A WT as well ascnf1, and single loci ofhlymutants from cluster I and II were all able to damage Caco-2 (Heterogeneous human epithelial colorectal adenocarcinoma) cell tight junctions. However, this phenotype was lost in a mutant with knockout (Δ) of bothhlyloci (P<0.001). CONCLUSIONS: UC-associated E. coliproducing α-hemolysin can cause rapid loss of tight junction integrity in differentiated Caco-2 cell monolayers. This effect was abolished in a mutant unable to express α-hemolysin. These results suggest that high Hly expression may be a mechanism by which specific strains of E. colipathobionts can contribute to epithelial barrier dysfunction and pathophysiology of disease in IBD

Effects of probiotics (Vivomixx®) in obese pregnant women and their newborn:Study protocol for a randomized controlled trial

BACKGROUND: Maternal obesity is associated with increased risks of adverse pregnancy-related complications and outcomes for both mothers and infants. Overweight and obese women have an increased risk of pregnancy-induced hypertension, preeclampsia and gestational diabetes mellitus (GDM). Infant Body Mass index (BMI) and the risk of obesity in adulthood are related to maternal gestational weight gain (GWG). Preventive lifestyle and dietary interventions are time-consuming and do not always reduce GWG or the risk of maternal pregnancy complications. Recent research has indicated that the gut microbiota may play a significant role in the development of obesity. Some studies have indicated that the daily consumption of probiotics may reduce the risk of preeclampsia, maintain serum insulin levels and reduce the frequency of GDM in pregnant women. The aims of this study are to investigate whether daily probiotic supplements in obese women during pregnancy can limit gestational weight gain, improve glucose homeostasis and thereby improve maternal, fetal and infant health outcomes. METHODS: A pilot study including 50 obese pregnant nulliparous women with a prepregnancy BMI of between 30 and 35 kg/m(2) will be randomized to receive daily probiotics (four capsules of Vivomixx®; total of 450 billion CFU/day, including eight probiotic bacterial strains) or placebo from gestational age 14–20 weeks until delivery. The infants will be followed until 9 months of age. The women will be monitored by weight, blood, fecal, vaginal and urine samples, diet questionnaires and hospital record review. Primary outcomes are: maternal weight gain, glycated hemoglobin (HbA1c) level and changes in glucose concentration measured during an oral glucose tolerance test. Secondary outcomes are: microbiota and inflammatory markers in mother and child, pregnancy complications, pregnancy outcomes, physical activity and the body composition of the neonate. DISCUSSION: We expect to find alterations in the metabolic profiles, microbiota and possibly pregnancy outcomes. From a clinical point of view the effects of Vivomixx® could control weight gain and reduce complications during pregnancy by inducing changes in the gut microbiota. Furthermore, this intervention during pregnancy could influence the infant’s microbiota, which could have important implications for infant development and health. TRIAL REGISTRATION: ClincalTrials.gov Identifier: NCT02508844, registered on 11 May 2015. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s13063-016-1617-5) contains supplementary material, which is available to authorized users

Characterization of Diarrheagenic Enteroaggregative Escherichia coli in Danish Adults—Antibiotic Treatment Does Not Reduce Duration of Diarrhea

Enteroaggregative Escherichia coli (EAEC) is frequently isolated from sporadic cases of diarrhea and in outbreaks of gastroenteritis in several regions of the world. The pathophysiology of EAEC continues to be enigmatic, and the efficacy of antibiotic treatment in EAEC-associated diarrhea has been discussed. Since the level of antibiotic resistance is increasing, it is essential to restrict the use of antibiotics to prevent further resistance development. We aimed to investigate EAEC strains in adult Danish patients suffering from diarrhea and from healthy controls. We examined the antibiotic resistance in EAEC strains, the clinical response to antibiotic treatment in EAEC diarrheal cases, and the distribution of virulence genes in diarrheal cases. The EAEC strains were collected from patients suffering from diarrhea in a Danish multicenter study. A medical doctor interviewed the patients by using a questionnaire regarding gastrointestinal symptoms, exposures, and use of antibiotic and over-the-counter antidiarrheal drugs. Follow-up was performed after 3–5 months to inquire about differential diagnosis to gastrointestinal disease. A multiplex polymerase chain reaction characterized virulence genes in diarrheal cases. Finally, the level of antibiotic resistance was examined by using the disc diffusion method. Asymptomatic carriage of EAEC in the adult Danish population was rare, in contrast to findings in healthy Danish children. The duration of diarrhea was not shortened by antibiotic treatment, specifically ciprofloxacin treatment, or by over-the-counter antidiarrheal drugs. Follow-up revealed no pathology in diarrheal patients apart from irritable bowel syndrome in two patients. A high number of patients suffered from long-term diarrhea, which was associated with the enterotoxin EAST-1 and a high virulence factor score. A high level of antibiotic resistance was observed and 58% of the EAEC strains were multidrug resistant. Multidrug resistance was most pronounced in cases of travelers' diarrhea, and it was seen that antibiotic treatment did not reduce the duration of diarrhea