Changes in gut microbiota and lactose intolerance symptoms before and after daily lactose supplementation in individuals with the lactase nonpersistent genotype.

BACKGROUND: Approximately 70%-100% of the Asian adult population is lactase nonpersistent (LNP). The literature shows that many individuals with the LNP-genotype can consume ≤12 g of lactose without experiencing gastrointestinal discomfort. Repetitive consumption of lactose may reduce intolerance symptoms via adaptation of the gut microbiota. OBJECTIVE: This study aimed to assess the effects of daily consumption of incremental lactose doses on microbiota composition and function, and intolerance symptoms. METHODS: Twenty-five healthy adults of Asian origin, carrying the LNP-genotype were included in this 12-wk before and after intervention trial. Participants consumed gradually increasing lactose doses from 3 to 6 g to 12 g twice daily, each daily dose of 6 g, 12 g, or 24 g being provided for 4 consecutive weeks. Participants handed-in repeated stool samples and underwent a 25 g lactose challenge hydrogen breath test (HBT) before and after the 12-wk intervention. Daily gastrointestinal symptoms and total symptom scores (TSSs) during the lactose challenge were recorded. RESULTS: A significant increase from 5.5% ± 7.6% to 10.4% ± 9.6% was observed in Bifidobacterium relative abundance after the intervention (P = 0.009), accompanied by a 2-fold increase (570 ± 269 U/g; P < 0.001) in fecal ß-galactosidase activity compared with baseline (272 ± 158 U/g). A 1.5-fold decrease (incremental area under the curve; P = 0.01) in expired hydrogen was observed during the second HBT (38 ± 35 ppm·min), compared with the baseline HBT (57 ± 38 ppm·min). There was a nonsignificant decrease in TSS (10.6 ± 8.3 before compared with 8.1 ± 7.2 after intervention; P = 0.09). Daily consumption of lactose was well tolerated, with mild to no gastrointestinal complaints reported during the intervention. CONCLUSIONS: Increased levels of Bifidobacterium indicate an adaptation of the gut microbiota upon repetitive consumption of incremental doses of lactose, which was well tolerated as demonstrated by reduced expired hydrogen concentrations during the second 25-g lactose HBT. Bifidobacteria metabolize lactose without gas production thereby potentially reducing intestinal gas formation in the gut of individuals with the LNP-genotype. This increased lactose tolerance possibly lifts the necessity to remove nutrient-rich dairy foods completely from the diet. The trial is registered at the International Clinical Trials Registry Platform: NL9516. The effect of dietary lactose in lactase nonpersistent individuals on gut microbiota.

Exploring the Link between Leaky-Gut-Related Markers and Metabolic Health in a Large Dutch Adult Population.

A leaky gut can trigger chronic inflammation and poses a primary risk for metabolic diseases. This study established a relationship between intestinal integrity (leaky gut) and metabolic health in a general population. Leaky-gut markers (LGMs) were studied in a large population of Dutch adults with a broad spectrum of metabolic health. This study enrolled 500 individuals selected within the NQplus cohort study (n = 2048) by stratified randomization, based on waist circumference, fasting glucose, and high-density lipoprotein (HDL) cholesterol to obtain a representative and balanced population in terms of metabolic health parameters, sex (male/female), and age (<54/≥54 years). LGMs-zonulin, lipopolysaccharide-binding protein (LBP), and soluble CD14 (sCD14)-were measured in EDTA plasma or serum. Zonulin was most strongly associated with metabolic health. Zonulin and LBP were most strongly associated with the inflammatory marker C-reactive protein (CRP). The quartile analysis for zonulin and LBP showed that most metabolic health parameters and CRP levels increased from Q1 to Q4, with significant differences between quartiles, except for markers related to glucose homeostasis (glucose and glycated hemoglobin A1c (HbA1c)). Associations between LGMs and metabolic health parameters in this large Dutch adult population indicate that LGMs are valuable markers for identifying people at risk of a leaky gut and subsequent chronic inflammation linked to metabolic disorders.

An Observational Study to Evaluate the Association between Intestinal Permeability, Leaky Gut Related Markers, and Metabolic Health in Healthy Adults.

We explored whether metabolic health is linked to intestinal permeability, using a multi-sugar (MS) permeability test, and whether intestinal permeability is correlated with the leaky gut-related markers (LGM) zonulin, LBP, and sCD14. Metabolically healthy (n = 15) and unhealthy subjects (n = 15) were recruited based on waist circumference, fasting glucose, and high-density lipoprotein cholesterol levels. Participants underwent an MS permeability test that assessed site-specific permeabilities of the gastroduodenum and small and large intestines. The test was performed with/without an acetylsalicylic acid challenge to measure and correlate the gut permeability, LGM, and metabolic health. At baseline, metabolic health showed no correlation with gut permeability. Significant correlations were found between the metabolic health parameters and LGM. In the acetylsalicylic acid challenged MS permeability test, low-density lipoprotein cholesterol was correlated with the sucralose/erythritol ratio, reflecting the whole intestinal permeability. Correlations between most metabolic health parameters and LGM during the acetylsalicylic acid challenge were less pronounced than at baseline. In both MS permeability tests, no significant correlations were found between LGM (plasma and serum) and gut permeability. Thus, correlations between LGM and metabolic health might not be linked with paracellular gut permeability. Transcellular translocation and/or lipoprotein-related transportation is a more likely mechanism underlying the association between LGM and metabolic health.

Clostridioides difficile toxin A-mediated Caco-2 cell barrier damage was attenuated by insect-derived fractions and corresponded to increased gene transcription of cell junctional and proliferation proteins.

Pathogenesis of C. difficile in the intestine is associated with the secretion of toxins which can damage the intestinal epithelial layer and result in diseases such as diarrhoea. Treatment for C. difficile infections consists of antibiotics which, however, have non-specific microbiocidal effects and may cause intestinal dysbiosis which results in subsequent health issues. Therefore, alternative treatments to C. difficile infections are required. We investigated whether different black soldier fly- and mealworm-derived fractions, after applying the INFOGEST digestion protocol, could counteract C. difficile toxin A-mediated barrier damage of small intestinal Caco-2 cells. Treatment and pre-treatment with insect-derived fractions significantly (p < 0.05) mitigated the decrease of the transepithelial electrical resistance (TEER) of Caco-2 cells induced by C. difficile toxin A. In relation to these effects, RNA sequencing data showed an increased transcription of cell junctional and proliferation protein genes in Caco-2 cells. Furthermore, the transcription of genes regulating immune signalling was also increased. To identify whether this resulted in immune activation we used a Caco-2/THP-1 co-culture model where the cells were only separated by a permeable membrane. However, the insect-derived fractions did not change the basolateral secreted IL-8 levels in this model. To conclude, our findings suggest that black soldier fly- and mealworm-derived fractions can attenuate C. difficile induced intestinal barrier disruption and they might be promising tools to reduce the symptoms of C. difficile infections.

Identification of leaky gut-related markers as indicators of metabolic health in Dutch adults: The Nutrition Questionnaires plus (NQplus) study.

BACKGROUND AND AIM: Chronic inflammation is a primary risk factor for chronic metabolic disease and may be triggered by a "leaky gut." Several biomarkers have been recognized to indicate intestinal permeability (i.e., leaky gut) and bacterial translocation. Nonetheless, which of these biomarkers exhibit the highest correlation with metabolic health parameters remains unclear. Hence, this study aimed to explore the correlation between leaky gut-related markers and metabolic health. METHODS: Based on waist circumference, plasma fasting glucose, plasma gamma-glutamyl transpeptidase (GGT), and plasma LDL cholesterol, two groups of 40 subjects with the most extreme metabolic health profiles were selected from the NQplus cohort study (n = 2048), which was previously conducted by the Wageningen University's Division of Human Nutrition. Eight potential leaky gut-related markers were selected from the literature and measured in serum or EDTA plasma samples of these selected individuals. These samples were also obtained from the NQplus cohort study. RESULTS: From the leaky gut markers, levels of zonulin, lipopolysaccharide-binding protein, soluble CD14, bactericidal/permeability-increasing protein, and peptidoglycan were significantly higher in individuals with unhealthy metabolic profiles (p<0.05). No differences in EndoCAb IgM, EndoCAb IgA, and EndoCAb IgG were observed between healthy and unhealthy individuals. Stepwise regression analysis revealed that zonulin was substantially associated with metabolic health parameters such as BMI, blood glucose, triglyceride, GGT, and C-reactive protein levels. C-reactive protein, an inflammation marker, showed the most pronounced association with zonulin. CONCLUSIONS: Biomarkers that link a leaky gut and subsequent bacterial translocation to metabolic health were identified in this study. Especially zonulin may aid in monitoring a leaky gut and detecting individuals at risk for developing chronic metabolic diseases.

A combined microphysiological-computational omics approach in dietary protein evaluation.

Food security is under increased pressure due to the ever-growing world population. To tackle this, alternative protein sources need to be evaluated for nutritional value, which requires information on digesta peptide composition in comparison to established protein sources and coupling to biological parameters. Here, a combined experimental and computational approach is presented, which compared seventeen protein sources with cow's whey protein concentrate (WPC) as the benchmark. In vitro digestion of proteins was followed by proteomics analysis and statistical model-based clustering. Information on digesta peptide composition resulted in 3 cluster groups, primarily driven by the peptide overlap with the benchmark protein WPC. Functional protein data was then incorporated in the computational model after evaluating the effects of eighteen protein digests on intestinal barrier integrity, viability, brush border enzyme activity, and immune parameters using a bioengineered intestine as microphysiological gut system. This resulted in 6 cluster groups. Biological clustering was driven by viability, brush border enzyme activity, and significant differences in immune parameters. Finally, a combination of proteomic and biological efficacy data resulted in 5 clusters groups, driven by a combination of digesta peptide composition and biological effects. The key finding of our holistic approach is that protein source (animal, plant or alternative derived) is not a driving force behind the delivery of bioactive peptides and their biological efficacy.

In Vitro Studies Toward the Use of Chitin as Nutraceutical: Impact on the Intestinal Epithelium, Macrophages, and Microbiota.

SCOPE: Chitin, the most abundant polysaccharide found in nature after cellulose, is known for its ability to support wound healing and to lower plasma-oxidized low-density lipoprotein (LDL) levels. Studies have also revealed immunomodulatory potential but contradicting results are often impossible to coalesce through usage of chitin of different or unknown physicochemical consistency. In addition, only a limited set of cellular models have been used to test the bioactivity of chitin. METHODS AND RESULTS: Chitin is investigated with well-defined physicochemical consistency for its immunomodulatory potency using THP-1 macrophages, impact on intestinal epithelial barrier using Caco-2 cells, and fermentation by fecal-derived microbiota. Results show that chitin with a degree of acetylation (DA) of ≈83%, regardless of size, does not affect the intestinal epithelial barrier integrity. Large-sized chitin significantly increases acetic acid production by gut microbiota without altering the composition. Exposure of small-sized chitin to THP-1 macrophages lead to significantly increased secretion of IL-1ß, IL-8, IL-10, and CXCL10 in a multi-receptor and clathrin-mediated endocytosis dependent manner. CONCLUSIONS: These findings indicate that small-sized chitin does not harm the intestinal barrier nor affects SCFA secretion and microbiota composition, but does impact immune activity which could be beneficial to subjects in need of immune support or activation.

Identification and in silico bioinformatics analysis of PR10 proteins in cashew nut.

Proteins from cashew nut can elicit mild to severe allergic reactions. Three allergenic proteins have already been identified, and it is expected that additional allergens are present in cashew nut. pathogenesis-related protein 10 (PR10) allergens from pollen have been found to elicit similar allergic reactions as those from nuts and seeds. Therefore, we investigated the presence of PR10 genes in cashew nut. Using RNA-seq analysis, we were able to identify several PR10-like transcripts in cashew nut and clone six putative PR10 genes. In addition, PR10 protein expression in raw cashew nuts was confirmed by immunoblotting and liquid chromatography-mass spectrometry (LC-MS/MS) analyses. An in silico allergenicity assessment suggested that all identified cashew PR10 proteins are potentially allergenic and may represent three different isoallergens.

Validation of three rapid screening methods for detection of verotoxin-producing Escherichia coli in foods: interlaboratory study.

An interlaboratory study was conducted for the validation of 3 methods for the detection of all verotoxin-producing Escherichia coli (VTEC) in foods. The methods were a multi-analyte 1-step lateral flow immunoassay (LFIA) for detection of E. coli O157 and verotoxin (VT); an enzyme-linked immunosorbent assay targeted against VT1, VT2, and VT2c (VT-ELISA); and a polymerase chain reaction (PCR) method for detection of VT genes (VT-PCR). Aliquots (25 g or 25 mL) of 4 food types (raw minced [ground] beef, unpasteurized milk, unpasteurized apple juice [cider], and salami) were individually inoculated with low numbers (<9 to 375 cells/25 g) of 6 test strains of E. coli (serogroups O26, O103, O111, O145, and O157) with differing VT-producing capabilities. Five replicates for each test strain and 5 uninoculated samples were prepared for each food type. Fourteen participating laboratories analyzed samples using the LFIA, 9 analyzed the samples by ELISA, and 9 by PCR. The LFIA for O157 and VT had a specificity (correct identification of negative samples) of 92 and 94%, respectively, and a sensitivity (correct identification of positive samples) of 94 and 55%, respectively. The VT-ELISA and VT-PCR had a specificity of 98 and 99%, respectively, and a sensitivity of 89 and 72%, respectively.