Umbilical Cord Blood IgA Levels and Bacterial Profile in Preterm Neonates Born with Maternal Risk Factors for Early-Onset Neonatal Sepsis.

OBJECTIVES: To investigate the IgA levels and bacterial profile in umbilical cord blood (UCB) samples of mothers with risk factors compared to those without risk factors; and to understand the link between UCB culture positivity and neonatal outcomes [early-onset sepsis (EOS) or death within 7 d of life]. METHODS: This is a pilot prospective case-control study. Mothers with preterm deliveries (gestational age <34 wk) were enrolled in two groups- Cases: Those with antenatal risk factors (prolonged duration of rupture of membranes of ≥24 h or chorioamnionitis) and controls: Those without these two risk factors. Serum IgA levels was assayed and microbiological culture was tested in UCB samples. 16S sequencing to determine the UCB microbiome was performed in a subset of samples (n = 15). Neonates were followed-up for the occurrence of EOS or death until 7 d of life. RESULTS: Forty-nine mothers as cases and 50 mothers as controls were consecutively enrolled. No significant difference was observed in the IgA levels (60.5 vs. 58.1 mg/L; p = 0.71), neonatal blood culture positivity (4.1% vs. 8.0%; p = 0.41) and UCB culture positivity (30.6% vs. 26.0%; p = 0.61) in the two groups. No difference was observed between the groups in occurrence of EOS or death within 7 d of life. Proteobacteria, Firmicutes and Actinobacteria were the most abundant phyla. Serratia, Bifidobacterium, Collinsella, Meganomas and Blautia being the most common genera. CONCLUSIONS: Cord blood IgA concentration could not differentiate the neonates at-risk of infection due to its presence in both the groups.

Comparison of Small Gut and Whole Gut Microbiota of First-Degree Relatives With Adult Celiac Disease Patients and Controls.

Recent studies on celiac disease (CeD) have reported alterations in the gut microbiome. Whether this alteration in the microbial community is the cause or effect of the disease is not well understood, especially in adult onset of disease. The first-degree relatives (FDRs) of CeD patients may provide an opportunity to study gut microbiome in pre-disease state as FDRs are genetically susceptible to CeD. By using 16S rRNA gene sequencing, we observed that ecosystem level diversity measures were not significantly different between the disease condition (CeD), pre-disease (FDR) and control subjects. However, differences were observed at the level of amplicon sequence variant (ASV), suggesting alterations in specific ASVs between pre-disease and diseased condition. Duodenal biopsies showed higher differences in ASVs compared to fecal samples indicating larger disruption of the microbiota at the disease site. The duodenal microbiota of FDR was characterized by significant abundance of ASVs belonging to Parvimonas, Granulicatella, Gemella, Bifidobacterium, Anaerostipes, and Actinomyces genera. The duodenal microbiota of CeD was characterized by higher abundance of ASVs from genera Megasphaera and Helicobacter compared to the FDR microbiota. The CeD and FDR fecal microbiota had reduced abundance of ASVs classified as Akkermansia and Dorea when compared to control group microbiota. In addition, predicted functional metagenome showed reduced ability of gluten degradation by CeD fecal microbiota in comparison to FDRs and controls. The findings of the present study demonstrate differences in ASVs and predicts reduced ability of CeD fecal microbiota to degrade gluten compared to the FDR fecal microbiota. Further research is required to investigate the strain level and active functional profiles of FDR and CeD microbiota to better understand the role of gut microbiome in pathophysiology of CeD.

First Degree Relatives of Patients with Celiac Disease Harbour an Intestinal Transcriptomic Signature that Might Protect them from Enterocyte Damage.

INTRODUCTION: Celiac disease (CeD) is an autoimmune enteropathy which affects approximately 0.7% of the global population. While first-degree relatives (FDR) of patients with CeD have a 7.5% risk of developing enteropathy, many remain protected. Therefore, intestinal mucosa of FDR might have protective compensatory mechanisms against immunological injury. We have explored the protective mechanisms that may be active in intestinal mucosa of FDR. METHODS: Intestinal mucosal biopsies (4-5 pieces) from treatment naïve patients with CeD (n = 12), FDR (n = 12) (anti-tTG negative) and controls (n = 12) (anti-tTG negative) were obtained from each individual and subjected to microarray analysis using HT-12-v4 Human Expression BeadChips (Illumina). Differential gene expression analysis was carried out among CeD, FDR and controls; and resulting gene lists were analyzed using gene ontology and pathway enrichment tools. RESULTS: Patients with CeD, FDR and control groups displayed significant differential gene expression. Thirty seven genes were upregulated and 372 were downregulated in the intestinal mucosa of FDR in comparison to CeD and controls. Pseudogenes constituted about 18% (315/1751) of FDR differentially expressed genes, and formed "clusters" that associated uniquely with individual study groups. The three study groups segregated into distinct clusters in unsupervised (PCA) and supervised (random forests) modelling approaches. Pathways analysis revealed an emphasis on crypt-villous maintenance and immune regulation in the intestinal mucosa of FDR. CONCLUSIONS: Our analysis suggests that the intestinal mucosa of celiac FDR consist of a unique molecular phenotype that is distinct from CeD and controls. The transcriptomic landscape of FDR promotes maintenance of crypt-villous axis and modulation of immune mechanisms. These differences clearly demonstrate the existence of compensatory protective mechanisms in the FDR intestinal mucosa.

Proteome analysis of the macroscopically affected colonic mucosa of Crohn's disease and intestinal tuberculosis.

Differentiation between intestinal tuberculosis (ITB) and Crohn's disease (CD) is challenging in geographical regions where both these diseases are prevalent. There is a need of biomarkers for differentiation between these two disorders. Colonic biopsies from inflamed mucosa of treatment-naive patients with ITB, CD and controls were used for analysis. Protein extracted from biopsies was digested with trypsin and resulting peptides were labeled with iTRAQ reagents. The peptides were subsequently analyzed using LC-MS/MS for identification and quantification. Gene ontology annotation for proteins was analyzed in PANTHER. Validation experiments were done for six differentially expressed proteins using immunohistochemistry. 533 proteins were identified and 241 proteins were quantified from 5 sets of iTRAQ experiments. While 63 were differentially expressed in colonic mucosa of patients with CD and ITB in at least one set of iTRAQ experiment, 11 proteins were differentially expressed in more than one set of experiments. Six proteins used for validation using immunohistochemistry in a larger cohort of patients; none of them however was differentially expressed in patients with ITB and CD. There are differentially expressed proteins in tissue proteome of CD and ITB. Further experiments are required using a larger cohort of homogeneous tissue samples.

Prevalence of celiac disease among first-degree relatives of Indian celiac disease patients.

BACKGROUND: Celiac disease, once thought to be uncommon in Asia, is now recognized in Asian nations as well. We investigated the prevalence of celiac disease in first-degree relatives of celiac disease patients followed in our centre. METHODS: First-degree relatives were screened prospectively for celiac disease using questionnaire-based interview and anti-tissue transglutaminase antibody. Serology positive first-degree relatives underwent duodenal biopsies. Diagnosis of celiac disease was made based on positive serology and villous abnormality Marsh grade 2 or higher. Human leucocyte antigen DQ2/-DQ8 was also assessed in 127 first-degree relatives. RESULTS: 434 first-degree relatives of 176 celiac disease patients were prospectively recruited; 282 were symptomatic (64.9%), 58 were positive for serology (13.3%). Seroprevalence was higher in female than in males (19% vs 8.5%; p=0.001) and highest in siblings (16.9%) than parents (13.6%) and children (5.9%) of celiac patients (p=0.055); 87.4% first-degree relatives were human leucocyte antigen-DQ2/-DQ8 positive. Overall prevalence of celiac disease was 10.9% amongst first-degree relatives. CONCLUSIONS: The prevalence of celiac disease in first-degree relatives of celiac disease patients was 10.9% in our cohort, and 87% had human leucocyte antigen-DQ2 or -DQ8 haplotype. All first-degree relatives of celiac disease patients should be screen for celiac disease even if asymptomatic or with atypical manifestations.

Structural and Functional Changes in the Tight Junctions of Asymptomatic and Serology-negative First-degree Relatives of Patients With Celiac Disease.

BACKGROUND: Ten to 15% of first-degree relatives (FDRs) of celiac disease (CeD) patients develop CeD. Although intestinal barrier functions (intestinal permeability) are abnormal in the subset of serology-negative FDRs, what leads to the abnormal barrier function is not known. GOALS: To study the ultrastructure and functions of tight junctions in serology-negative FDRs of CeD patients. STUDY: The intestinal permeability was measured in 97 asymptomatic and anti-tissue transglutaminase antibody (anti-tTG Ab)-negative FDRs (using the lactulose mannitol ratio) and in 75 controls. The ultrastructure of tight junctions using transmission electron microscopy, and the expression of key tight junction proteins (claudin-2, claudin-3, occludin, JAM-A, and ZO-1) and zonulin using real-time PCR and immunohistochemistry were assessed in anti-tTG Ab-negative, HLA-DQ2/-DQ8-positive FDRs having normal villi and in disease controls. In addition, the serum zonulin level was measured in 172 anti-tTG Ab-negative FDRs and 198 controls. RESULTS: The intestinal permeability was significantly increased in FDRs than in controls. Ultrastructural abnormalities such as dilatation of the tight junction (P=0.004) and loss of the pentalaminar structure (P=0.001) were more common in FDRs than in disease controls. There was significant underexpression of tight junction proteins ZO-1 (P=0.040) and occludin (P=0.041) in FDRs. There was no significant difference in the serum zonulin level between FDRs and controls (P=0.154). CONCLUSIONS: Even asymptomatic, anti-tTG-Ab-negative FDRs with a normal villous histology have both ultrastructural and functional abnormalities in tight junctions. These findings are indirect evidence of the presence of tight junction abnormalities before the onset of the disease and may have therapeutic implications.

Intermittent Directly Observed Therapy for Abdominal Tuberculosis: A Multicenter Randomized Controlled Trial Comparing 6 Months Versus 9 Months of Therapy.

BACKGROUND: The duration of treatment of gastrointestinal tuberculosis continues to be a matter of debate. The World Health Organization advocates intermittent directly observed short-course therapy (DOTs), but there is a lack of data of its efficacy in abdominal tuberculosis. We therefore conducted a multicenter randomized controlled trial to compare 6 months and 9 months of antituberculosis therapy using DOTs. METHODS: One hundred ninety-seven patients with abdominal tuberculosis (gastrointestinal, 154; peritoneal, 40; mixed, 3) were randomized to receive 6 months (n = 104) or 9 months (n = 93) of antituberculosis therapy using intermittent directly observed therapy. Patients were followed up 1 year after completion of treatment to assess recurrence. Patients were evaluated for primary endpoint (complete clinical response, partial response, and no response) and secondary endpoint (recurrence of the disease at the end of 1 year of follow-up). RESULTS: Baseline characteristics were similar between the 2 randomized groups. There was no difference between the 6-month group and 9-month group in the complete clinical response rate on per-protocol analysis (91.5% vs 90.8%; P = .88) or intent-to-treat analysis (75% vs 75.8%; P = .89). Only 1 patient in the 9-month group and no patients in the 6-month group had recurrence of disease. Side effects occurred in 21 (21.3%) and 16 (18.2%) patients in the 6-month and 9-month groups, respectively. CONCLUSIONS: There was no difference in efficacy of antituberculosis therapy delivered for either 6 months or 9 months in either gastrointestinal or peritoneal tuberculosis, confirming the efficacy of intermittent directly observed therapy. CLINICAL TRIALS REGISTRATION: NCT01124929.

Metabolic abnormalities of gastrointestinal mucosa in celiac disease: An in vitro proton nuclear magnetic resonance spectroscopy study.

BACKGROUND AND AIM: Celiac disease (CeD) is a common autoimmune disorder in which ingestion of gluten and related proteins leads to inflammation in the small intestine. Although the histological findings in CeD are characteristic, they are not specific. In this study, proton nuclear magnetic resonance (NMR) spectroscopy was used to investigate the differences in metabolic profile of duodenal mucosal biopsies of patients with CeD and controls to find out the biomarker/s of villous atrophy. METHODS: Duodenal mucosal biopsies were collected from 29 CeD patients (mean age 26.2 ± 10.8 years) and 17 controls (mean age 34.1 ± 11.1 years) and were subjected to proton NMR spectroscopy following perchloric acid extraction. Assignment of metabolite resonances was carried out and their concentrations were determined. For comparison between the groups unpaired t-test/Wilcoxon rank sum test was used. Partial least squares-discriminant analysis was performed to study the clustering behavior of the samples from CeD patients and controls using the Unscrambler 10.2 software. RESULTS: Partial least squares-discriminant analysis clearly differentiated CeD patients from controls. Significantly higher concentrations of isoleucine, leucine, aspartate, succinate, and pyruvate, and lower concentration of glycerophosphocholine, were observed in the duodenal mucosa of CeD patients compared with controls. The results suggest abnormalities in glycolysis, Krebs cycle (energy deficiency), and amino acid metabolism, which may affect the biosynthetic pathways and consequently contribute to villous atrophy. CONCLUSIONS: NMR spectroscopy with multivariate analysis of duodenal mucosal biopsies revealed a characteristic metabolic profile in CeD patients. The work provided an insight in determining biomarker/s for villous atrophy and diagnosis of CeD patients.

Celiac disease: a disease with varied manifestations in adults and adolescents.

OBJECTIVE: We aimed to determine the characteristics of patients with celiac disease and differences between those who presented during adolescence or adulthood. METHODS: We retrospectively reviewed the case records of 233 consecutive patients with celiac disease who were diagnosed at 12-18 years or >18 years of age. The diagnosis of celiac disease was made on the basis of the modified European Society of Pediatric Gastroenterology, Hepatology and Nutrition criteria. RESULTS: The diagnosis of celiac disease was made after 18 years of age in 153 (65.7%) patients. Median duration of symptoms at the diagnosis was 54 months (range 1 month to 29 years). In all, 103 (44.2%) patients with atypical manifestations were referred by other departments for evaluation. Chronic diarrhea (48.5%), short stature (27.0%) and chronic anemia (9.0%) were the common modes of presentation. Elevated level of aminotransaminase were present in 50 (24.3%) patients. Chronic diarrhea, hypocalcemia and hypoalbuminemia were present in significantly higher number of adult than adolescent patients. In all, 227 (97.4%) patients responded to a 6-month gluten-free diet and six non-responders were non-compliant. CONCLUSIONS: More than 40% of the patients with celiac disease present to clinicians other than gastroenterologists or internists with atypical manifestations. A high index of suspicion is required for diagnosing its variant forms.

Techniques of functional and motility test: how to perform and interpret intestinal permeability.

Transport of molecules across the intestinal epithelium takes place through 2 major routes, ie, trans-cellular and paracellular. Assessment of intestinal permeability is performed to assess the overall function of transport through the intestinal epithelial paracellular route. Urinary excretion of disaccharides and monosaccharides and ratio of their excretion is a basis for measurement of intestinal permeability. Lactulose and mannitol ratio is the most commonly used test for assessment of small intestinal permeability and the most reliable method for measurement of concentration of lactulose and mannitol in the urine is high performance liquid chromatography. After the measurement of concentration of probes in the urine; the results are expressed as the ratio of percentage excretion of the ingested dose of lactulose and mannitol in the urine. Testing of intestinal permeability is not required for routine patient care, however it is an important tool to understand the function of the paracellular transport in the research setting. Increase in intestinal permeability has been implicated in the pathogenesis of many autoimmune diseases including celiac disease, Crohn's disease, type I diabetes and food allergy.