Dietary intake of inulin-type fructans in active and inactive Crohn's disease and healthy controls: a case-control study.

BACKGROUND AND AIMS: Prebiotic inulin-type fructans are widely consumed in the diet and may have contrasting effects in Crohn's disease by stimulating gut microbiota and/or by generating functional gastrointestinal symptoms. The aim of this study was to measure fructan and oligofructose intakes in patients with active and inactive Crohn's disease compared with healthy controls. METHODS: Patients with active Crohn's disease (n = 98), inactive Crohn's (n = 99) and healthy controls (n = 106) were recruited to a case-control study. Dietary intake of inulin-type fructans was measured using a specific food frequency questionnaire and was compared between the three groups and between patients with different disease phenotypes (Montreal classification). Associations between intakes and disease activity (Harvey-Bradshaw Index, HBI) were also undertaken. RESULTS: Patients with active Crohn's disease had lower fructan intakes (median 2.9 g/d, interquartile range [IQR] 1.8) than those with inactive Crohn's (3.6 g/d, 2.1, p = 0.036) or controls (3.9 g/d, 2.1, p = 0.003) and lower oligofructose intakes (2.8 g/d, 1.8) than those with inactive Crohn's (3.5 g/d, 2.2, p = 0.048) or controls (3.8 g/d, 2.1, p = 0.003). There were no differences in intakes related to disease site or behaviour. There were negative correlations between HBI well-being score and fructan intake (ρ = -0.154, p = 0.03) and oligofructose intake (ρ = -0.156, p = 0.028) and for the HBI abdominal pain score and fructan (ρ = -0.164, p = 0.021) and oligofructose intake (ρ = -0.157, p = 0.027). CONCLUSIONS: Patients with active Crohn's disease consume lower quantities of fructans and oligofructose than their inactive counterparts and healthy controls. The impact of lower intakes of prebiotic fructans on gut microbiota is unknown and warrants further research.

Smokers with active Crohn's disease have a clinically relevant dysbiosis of the gastrointestinal microbiota.

BACKGROUND: Patients with Crohn's disease (CD) have an intestinal dysbiosis with components of the microbiota exerting differential immune effects. Smoking is associated with an increased incidence of CD, more frequent relapse, and greater burden of surgery. This study aimed to investigate the association between smoking and the intestinal microbiota in patients with active CD. METHODS: Patients with active CD (n = 103) and healthy controls (n = 66) were recruited and demographic and clinical data recorded including current smoking behavior. Fecal samples were collected and analyzed by fluorescent in situ hybridization using probes targeting 16S rRNA of bacteria previously shown to be altered in active CD (bifidobacteria, bacteroides, Clostridium coccoides-Eubacterium rectale, Escherichia coli, and Faecalibacterium prausnitzii). RESULTS: In total, 29/101 (29%) patients with CD and 8/58 (14%) controls were current smokers (P = 0.032). Following multivariate analysis, smoking was found to have a significant and independent effect on the microbiota of patients with CD, with higher Bacteroides-Prevotella in smokers (38.4%) compared with nonsmokers (28.1%) (F((1,93)) = 12.6, P = 0.001). Healthy controls who smoked also had higher Bacteroides-Prevotella (34.8%) than nonsmokers (24.1%) (F((1,55)) = 4.5, P = 0.038). In the pooled multivariate analysis, patients with CD had higher bifidobacteria (F((1,156)) = 30.5, P < 0.001), higher Bacteroides-Prevotella (F((1,156)) = 6.5, P = 0.012), and lower F. prausnitzii (F((1,156)) = 3.8, P = 0.052) compared with healthy controls. CONCLUSIONS: Smokers have luminal microbiota that consist of significantly higher bacteroides. Investigation of whether this is one mechanism through which the negative effects of smoking on CD are mediated is warranted.

Randomised, double-blind, placebo-controlled trial of fructo-oligosaccharides in active Crohn's disease.

INTRODUCTION: The commensal intestinal microbiota drive the inflammation associated with Crohn's disease. However, bacteria such as bifidobacteria and Faecalibacterium prausnitzii appear to be immunoregulatory. In healthy subjects the intestinal microbiota are influenced by prebiotic carbohydrates such as fructo-oligosaccharides (FOS). Preliminary data suggest that FOS increase faecal bifidobacteria, induce immunoregulatory dendritic cell (DC) responses and reduce disease activity in patients with Crohn's disease. AIMS AND METHODS: To assess the impact of FOS in patients with active Crohn's disease using an adequately powered randomised double-blind placebo-controlled trial with predefined clinical, microbiological and immunological end points. Patients with active Crohn's disease were randomised to 15 g/day FOS or non-prebiotic placebo for 4 weeks. The primary end point was clinical response at week 4 (fall in Crohn's Disease Activity Index of ≥ 70 points) in the intention-to-treat (ITT) population. RESULTS: 103 patients were randomised to receive FOS (n = 54) or placebo (n = 49). More patients receiving FOS (14 (26%) vs 4 (8%); p = 0.018) withdrew before the 4-week end point. There was no significant difference in the number of patients achieving a clinical response between the FOS and placebo groups in the ITT analysis (12 (22%) vs 19 (39%), p = 0.067). Patients receiving FOS had reduced proportions of interleukin (IL)-6-positive lamina propria DC and increased DC staining of IL-10 (p < 0.05) but no change in IL-12p40 production. There were no significant differences in the faecal concentration of bifidobacteria and F prausnitzii between the groups at baseline or after the 4-week intervention. CONCLUSION: An adequately powered placebo-controlled trial of FOS showed no clinical benefit in patients with active Crohn's disease, despite impacting on DC function. ISRCTN50422530.

Development and field evaluation of a new serological test for Taenia saginata cysticercosis.

Cattle infected with the tapeworm cyst, Taenia saginata metacestode (synonym: Cysticercus bovis) are a source of human infection if affected beef is eaten raw or undercooked. Control measures targeted at individual cattle rather than all animals in a T. saginata-exposed herd should help reduce costs and alleviate current constraints associated with managing an outbreak. To that end, we have developed a reliable diagnostic test for use in live animals that would enable veterinary regulators to focus disease control strategies. The test detects bovine anti-T. saginata immunoglobulin G1 antibodies using an enzyme-linked immunosorbent assay (ELISA) which relies on the excretory-secretory antigens of T. saginata. Animals were inoculated with 10, 100 or 1000 viable T. saginata eggs in order to simulate the parasite burden of field-infected animals (parasite load=1-86; n=28). By testing sera obtained from the inoculated animals 84 days post-inoculation, test sensitivity was estimated to be 92.9% (95% confidence interval or CI=83.4-100.0%). Another 17 animals inoculated with 5000 or 10,000 viable eggs of T. saginata and shown to harbour metacestodes at post-mortem, all tested positive in the ELISA. Test specificity estimated from a herd of field animals with no historical, epidemiological, or post-mortem evidence of infection was 90.6% (95% CI=87.0-94.2%; n=256 field cattle). Using the test on samples (n=347) from a T. saginata-infected feedlot, the Bayesian approach estimate of seroprevalence was 4.6% (95% probability intervals=0.5-10.3%). The test performance characteristics of the ELISA suggest that it will be adequate for field application in bovine cysticercosis outbreaks.

Construction of a complementary DNA library for Parelaphostrongylus tenuis and identification of a potentially sero-diagnostic recombinant antigen.

Newly developed serological tests for diagnosing parelaphostrongylosis in cervids, using the excretory-secretory products (ES) of the infective larvae of Parelaphostrongylus tenuis in enzyme-linked immunosorbent assays (ELISAs), have demonstrable superiority over the traditional method of larval recovery and microscopic identification. To generate a source of ELISA antigen by genetic engineering, we created a complementary DNA (cDNA) expression library by the reverse transcription of mRNA of P. tenuis adult worms, and ligation with the vector lambda-ZAP II. The library was screened using antisera produced in mice by immunization with a somatic antigen preparation of adult worms. Seventeen clones were isolated, sequenced, and checked for similarity to other DNA sequences in GenBank. A previously identified parasite gene encoding an aspartyl protease inhibitor (API) was isolated from the cDNA library, subcloned and expressed using the pET expression vector to produce a glutathione S transferase (GST)-His-S.Tag-P. tenuis API fusion protein (molecular weight = 63 kDa). An enzyme-linked immunosorbent assay utilizing the API fusion protein as the coating antigen was used to serologically diagnose all white-tailed deer (WTD, 10 out of 10) that had been inoculated with 6 - 150 L3 P. tenuis, indicating that the antigen may be a useful serodiagnostic antigen for P. tenuis infection in this cervid species.

Accuracy of an indirect fluorescent-antibody test and of a complement-fixation test for the diagnosis of Babesia caballi in field samples from horses.

We evaluated the indirect fluorescent-antibody (IFA) test and complement-fixation (CF) test for diagnosis of equine piroplasmosis in the absence of a gold standard. Using Evan's blue, we estimated the specificity of the IFA test on a parasite-free, field horse population to be 98% (95% confidence interval=97, 99). We observed an excellent test agreement (kappa=0.83) between two collaborating laboratories when the IFA test was performed on identical samples from an endemic area. Using Bayesian analysis with informative prior probability distributions, we estimated the sensitivity of the IFA test to be 92% (95% probability interval, PI=81, 98), and specificity to be 95% (95% PI=88, 99). The CF test sensitivity and specificity estimates were 28% (95% PI=15, 47) and 99% (95% PI=96, 100), respectively. We found the IFA to be superior to the CF test, and the inclusion of Evan's blue in test protocol improved the performance of the IFA test. We conclude that the IFA test for Babesia caballi is a sensitive and specific test for the diagnosis of equine piroplasmosis.

Validation of the indirect fluorescent antibody and the complement fixation tests for the diagnosis of Theileria equi.

The indirect fluorescent antibody (IFA) test for Theileria equi was evaluated to assess test's suitability for the serological diagnosis of equine piroplasmosis, to provide performance parameters for the purpose of test validation, and to compare it with the complement fixation (CF) test. Using a protocol that included Evan's blue, the specificity of the IFA test was estimated at 99.0% for T. equi by the classical method of analysis, and 96.6% by the Bayesian method. The use of Evan's blue in the test protocol increased test specificity and contributed to an excellent test agreement between two collaborating laboratories (kappa = 0.96). Using Bayesian analysis, the sensitivity estimate for the IFA test was 89.2%. The CF test sensitivity and specificity estimates for T. equi were 63.1 and 96.4%, respectively, as determined by Bayesian analysis. The IFA test was more sensitive than the CF test but the specificity estimates were similar.