Staff Publications

Staff Publications

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    'Staff publications' is the digital repository of Wageningen University & Research

    'Staff publications' contains references to publications authored by Wageningen University staff from 1976 onward.

    Publications authored by the staff of the Research Institutes are available from 1995 onwards.

    Full text documents are added when available. The database is updated daily and currently holds about 240,000 items, of which 72,000 in open access.

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    Does entry to center-based childcare affect gut microbial colonization in young infants?
    Hermes, Gerben D.A. ; Eckermann, Henrik A. ; Vos, Willem M. de; Weerth, Carolina de - \ 2020
    Scientific Reports 10 (2020)1. - ISSN 2045-2322

    Entry to center-based childcare (CC) at three months of life can be an important challenge for infants as it includes major stressors such as long maternal separations and frequently changing caregivers. Stress and the new environment may in turn alter the composition of the gut microbiota with possible implications for future health outcomes. As part of an ongoing longitudinal study, we investigated whether CC, as compared to being cared for by the parents at home, alters the composition of the gut microbiota, while accounting for known covariates of the infant gut microbiota. Stool samples of infants who entered CC (n = 49) and control infants (n = 49) were obtained before and four weeks after CC entrance. Using Redundancy analysis, Random Forests and Bayesian linear models we found that infant gut microbiota was not affected in a uniform way by entry to CC. In line with the literature, breastfeeding, birth mode, age, and the presence of siblings were shown to significantly impact the microbial composition.

    Fermentation of Chicory Fructo-Oligosaccharides and Native Inulin by Infant Fecal Microbiota Attenuates Pro-Inflammatory Responses in Immature Dendritic Cells in an Infant-Age-Dependent and Fructan-Specific Way
    Logtenberg, Madelon J. ; Akkerman, Renate ; An, Ran ; Hermes, Gerben D.A. ; Haan, Bart J. de; Faas, Marijke M. ; Zoetendal, Erwin G. ; Schols, Henk A. ; Vos, Paul de - \ 2020
    Molecular Nutrition & Food Research 64 (2020)13. - ISSN 1613-4125
    dendritic cells - in vitro fermentation - infant formula - inulin-type fructans - microbiota

    Scope: Inulin-type fructans are commonly applied in infant formula to support development of gut microbiota and immunity. These inulin-type fructans are considered to be fermented by gut microbiota, but it is unknown how fermentation impacts immune modulating capacity and whether the process of fermentation is dependent on the infant's age. Methods and results: The in vitro fermentation of chicory fructo-oligosaccharides (FOS) and native inulin are investigated using pooled fecal inocula of two- and eight-week-old infants. Both inocula primarily utilize the trisaccharides in FOS, while they almost completely utilize native inulin with degree of polymerization (DP) 3–8. Fecal microbiota of eight-week-old infants degrades longer chains of native inulin up to DP 16. This correlates with a higher abundance of Bifidobacterium and higher production of acetate and lactate after 26 h of fermentation. Fermented FOS and native inulin attenuate pro-inflammatory cytokines produced by immature dendritic cells (DCs), but profiles and magnitude of attenuation are stronger with native inulin than with FOS. Conclusion: The findings demonstrate that fermentation of FOS and native inulin is dependent on the infant's age and fructan structure. Fermentation enhances attenuating effects of pro-inflammatory responses in DCs, which depend mainly on microbial metabolites formed during fermentation.

    Individual and cohort-specific gut microbiota patterns associated with tissue-specific insulin sensitivity in overweight and obese males
    Hermes, Gerben D.A. ; Reijnders, Dorien ; Kootte, Ruud S. ; Goossens, Gijs H. ; Smidt, Hauke ; Nieuwdorp, Max ; Blaak, Ellen E. ; Zoetendal, Erwin G. - \ 2020
    Scientific Reports 10 (2020)1. - ISSN 2045-2322

    A growing body of evidence suggests that the human gut microbiota plays a role in the development of obesity and related metabolic diseases. However, there is little consensus between studies, which could be due to biological as well as technical variation. In addition, little human data are available to investigate whether tissue-specific insulin sensitivity is related to specific microbial patterns. We examined this relation in two independent cohorts of overweight and obese pre-diabetic men, using phylogenetic microarray data and hepatic, peripheral and adipose tissue insulin sensitivity that were determined by a two-step hyperinsulinemic-euglycemic clamp with [6,6-2H2]-glucose tracer infusion. Despite a prominent subject-specific microbiota, we found significant associations of microbial taxa with tissue-specific insulin sensitivity using regression analysis. Using random forests we found moderate associations with other measures of glucose homeostasis in only one of the cohorts (fasting glucose concentrations AUC = 0.66 and HbA1c AUC = 0.65). However, all findings were cohort-specific due to pronounced variation in microbiota between cohorts, suggesting the existence of alternative states for dysbiosis in metabolic syndrome patients. Our findings suggest individual or group related dynamics, instead of universal microbiota signals, related to the host when the overweight or obese state has already developed and argue that care should be taken with extrapolating significant correlations from single cohorts, into generalized biological relevance.

    Mediterranean diet intervention alters the gut microbiome in older people reducing frailty and improving health status : The NU-AGE 1-year dietary intervention across five European countries
    Ghosh, Tarini Shankar ; Rampelli, Simone ; Jeffery, Ian B. ; Santoro, Aurelia ; Neto, Marta ; Capri, Miriam ; Giampieri, Enrico ; Jennings, Amy ; Candela, Marco ; Turroni, Silvia ; Zoetendal, Erwin G. ; Hermes, Gerben D.A. ; Elodie, Caumon ; Brugere, Corinne Malpuech ; Pujos-Guillot, Estelle ; Berendsen, Agnes M. ; Groot, Lisette C.P.G.M. De; Feskins, Edith J.M. ; Kaluza, Joanna ; Pietruszka, Barbara ; Bielak, Marta Jeruszka ; Comte, Blandine ; Maijo-Ferre, Monica ; Nicoletti, Claudio ; Vos, Willem M. de; Fairweather-Tait, Susan ; Cassidy, Aedin ; Brigidi, Patrizia ; Franceschi, Claudio ; O'Toole, Paul W. - \ 2020
    Gut 69 (2020)7. - ISSN 0017-5749
    ageing - diet - enteric bacterial microflora - inflammation - intestinal bacteria

    Objective: Ageing is accompanied by deterioration of multiple bodily functions and inflammation, which collectively contribute to frailty. We and others have shown that frailty co-varies with alterations in the gut microbiota in a manner accelerated by consumption of a restricted diversity diet. The Mediterranean diet (MedDiet) is associated with health. In the NU-AGE project, we investigated if a 1-year MedDiet intervention could alter the gut microbiota and reduce frailty. Design: We profiled the gut microbiota in 612 non-frail or pre-frail subjects across five European countries (UK, France, Netherlands, Italy and Poland) before and after the administration of a 12-month long MedDiet intervention tailored to elderly subjects (NU-AGE diet). Results: Adherence to the diet was associated with specific microbiome alterations. Taxa enriched by adherence to the diet were positively associated with several markers of lower frailty and improved cognitive function, and negatively associated with inflammatory markers including C-reactive protein and interleukin-17. Analysis of the inferred microbial metabolite profiles indicated that the diet-modulated microbiome change was associated with an increase in short/branch chained fatty acid production and lower production of secondary bile acids, p-cresols, ethanol and carbon dioxide. Microbiome ecosystem network analysis showed that the bacterial taxa that responded positively to the MedDiet intervention occupy keystone interaction positions, whereas frailty-associated taxa are peripheral in the networks. Conclusion: Collectively, our findings support the feasibility of improving the habitual diet to modulate the gut microbiota which in turn has the potential to promote healthier ageing.

    Effect of wheat bran derived prebiotic supplementation on gastrointestinal transit, gut microbiota, and metabolic health: a randomized controlled trial in healthy adults with a slow gut transit
    Müller, Mattea ; Hermes, Gerben D.A. ; Canfora, Emanuel E. ; Holst, Jens J. ; Zoetendal, Erwin G. ; Smidt, Hauke ; Troost, Freddy ; Schaap, Frank G. ; Damink, Steven Olde ; Jocken, Johan W.E. ; Lenaerts, Kaatje ; Masclee, Ad A.M. ; Blaak, Ellen E. - \ 2020
    Gut Microbes (2020). - ISSN 1949-0976
    Arabinoxylan-Oligosaccharides - Energy metabolism - Gastrointestinal transit - Gut Hormones - Gut microbiota - Prebiotic - Stool consistency

    Acute intake of the wheat bran extract Arabinoxylan-Oligosaccharide (AXOS) modulates the gut microbiota, improves stool characteristics and postprandial glycemia in healthy humans. Yet, little is known on how long-term AXOS intake influences gastrointestinal (GI) functioning, gut microbiota, and metabolic health. In this randomized, placebo-controlled, double-blind study, we evaluated the effects of AXOS intake on GI function and metabolic health in adults with slow GI transit without constipation. Forty-eight normoglycemic adults were included with whole-gut transit time (WGTT) of >35 h receiving either 15 g/day AXOS or placebo (maltodextrin) for 12-wks. The primary outcome was WGTT, and secondary outcomes included stool parameters, gut permeability, short-chain fatty acids (SCFA), microbiota composition, energy expenditure, substrate oxidation, glucose, insulin, lipids, gut hormones, and adipose tissue (AT) function. WGTT was unchanged, but stool consistency softened after AXOS. 12-wks of AXOS intake significantly changed the microbiota by increasing Bifidobacterium and decreasing microbial alpha-diversity. With a good classification accuracy, overall microbiota composition classified responders with decreased WGTT after AXOS. The incretin hormone Glucagon-like protein 1 was reduced after AXOS compared to placebo. Energy expenditure, plasma metabolites, AT parameters, SCFA, and gut permeability were unchanged. In conclusion, intake of wheat bran extract increases fecal Bifidobacterium and softens stool consistency without major effects on energy metabolism in healthy humans with a slow GI transit. We show that overall gut microbiota classified responders with decreased WGTT after AXOS highlighting that GI transit and change thereof were associated with gut microbiota independent of Bifidobacterium. NCT02491125.

    Distal colonic transit is linked to gut microbiota diversity and microbial fermentation in humans with slow colonic transit
    Müller, Mattea ; Hermes, Gerben D.A. ; Canfora, Emanuel E. ; Smidt, Hauke ; Masclee, Ad A.M. ; Zoetendal, Erwin G. ; Blaak, Ellen E. - \ 2020
    American Journal of Physiology. Gastrointestinal and Liver Physiology 318 (2020)2. - ISSN 0193-1857 - p. G361 - G369.
    gastrointestinal transit - gut microbiota - short-chain fatty acids - stool consistency

    Longer colonic transit time and hard stools are associated with increased gut microbiota diversity. Here, we investigate to what extent quantitative measures of (segmental) colonic transit time were related to gut microbiota composition, microbial metabolites, and gut-related parameters in a human cross-sectional study. Using radiopaque markers, (segmental) colonic transit time (CTT) was measured in 48 lean/overweight participants with long colonic transit but without constipation. Fecal microbiota composition was determined using 16S rRNA gene amplicon sequencing. Associations between gastrointestinal transit (segmental CTT and stool frequency and consistency), microbiota diversity and composition, microbial metabolites [short-chain fatty acids (SCFA), branched-chain fatty acids, and breath hydrogen], habitual diet, and gut-related host parameters [lipopolysaccharide-binding protein (LBP) and fecal calprotectin] were investigated using univariate and multivariate approaches. Long descending (i.e., distal) colonic transit was associated with increased microbial α-diversity but not with stool consistency. Using unweighted and weighted UniFrac distance, microbiota variation was not related to (segmental) CTT but to demographics, diet, plasma LBP, and fecal calprotectin. Bray-Curtis dissimilarity related only to stool consistency. Rectosigmoid and descending colonic transit were negatively associated with fecal SCFA and plasma acetate, respectively. This study suggests that the distal colon transit may affect not only microbiota diversity but also microbial metabolism.NEW & NOTEWORTHY We extend previous findings showing that long distal colonic transit time influences microbial diversification and fermentation, whereas stool consistency is related to microbiota composition in humans with a long colonic transit. This study puts the importance of the (distal) colonic site in microbiota ecology forward, which should be considered in future therapeutic studies targeting, for instance, short-chain fatty acid production to improve metabolic health.

    NG-Tax 2.0: A Semantic Framework for High-Throughput Amplicon Analysis
    Poncheewin, Wasin ; Hermes, Gerben D.A. ; Dam, Jesse C.J. Van; Koehorst, Jasper J. ; Smidt, Hauke ; Schaap, Peter J. - \ 2020
    Frontiers in Genetics Livestock Genomics 10 (2020). - ISSN 1664-8021
    NG-Tax 2.0 is a semantic framework for FAIR high-throughput analysis and classification of marker gene amplicon sequences including bacterial and archaeal 16S ribosomal RNA (rRNA), eukaryotic 18S rRNA and ribosomal intergenic transcribed spacer sequences. It can directly use single or merged reads, paired-end reads and unmerged paired-end reads from long range fragments as input to generate de novo amplicon sequence variants (ASV). Using the RDF data model, ASV’s can be automatically stored in a graph database as objects that link ASV sequences with the full data-wise and element-wise provenance, thereby achieving the level of interoperability required to utilize such data to its full potential. The graph database can be directly queried, allowing for comparative analyses of over thousands of samples and is connected with an interactive Rshiny toolbox for analysis and visualization of (meta) data. Additionally, NG-Tax 2.0 exports an extended BIOM 1.0 (JSON) file as starting point for further analyses by other means. The extended BIOM file contains new attribute types to include information about the command arguments used, the sequences of the ASVs formed, classification confidence scores and is backwards compatible. The performance of NG-Tax 2.0 was compared with DADA2, using the plugin in the QIIME 2 analysis pipeline. Fourteen 16S rRNA gene amplicon mock community samples were obtained from the literature and evaluated. Precision of NG-Tax 2.0 was significantly higher with an average of 0.95 vs 0.58 for QIIME2-DADA2 while recall was comparable with an average of 0.85 and 0.77, respectively. NG-Tax 2.0 is written in Java. The code, the ontology, a Galaxy platform implementation, the analysis toolbox, tutorials and example SPARQL queries are freely available at under the MIT License.
    Arabinoxylan-Oligosaccharide Intake changes the microbiota and softens stool consistency without changes in gut transit and metabolic health in healthy adults
    Müller, Mattea ; Hermes, Gerben ; Canfora, Emanuel E. ; Holst, Jens J. ; Zoetendal, Erwin ; Smidt, Hauke ; Troost, Freddy ; Schaap, Frank G. ; Damink, Steven Olde ; Jocken, Johan W.E. ; Lenaerts, Kaatje ; Masclee, Ad A.M. ; Blaak, Ellen E. - \ 2019
    Wageningen University & Research
    PRJEB32919 - ERP115659 - human gut metagenome
    Prebiotic fibers may alter gastrointestinal (GI) transit time, microbiota composition and short chain fatty acid (SCFA) production, contributing to improved gut functionality and metabolic health. We investigated long-term effects of Arabinoxylan-Oligosaccharide (AXOS), a prebiotic dietary fiber on GI transit time, gut microbiota composition, and metabolic profile in adult participants.Methods: This randomized, placebo-controlled double-blind parallel study included 48 normoglycemic men and women (ages 20-55 y, body mass index (BMI) 19.8-30.5 kg/m2) with a slow whole-gut transit time (>35h) recruited during August 2015 to December 2016 in Maastricht, the Netherlands. Participants were randomly allocated to 12 weeks 15g/day AXOS or placebo (maltodextrin) intake. GI transit time, stool parameters, gut permeability, SCFA and microbiota composition were assessed before and after. Energy expenditure, substrate oxidation, glucose, insulin, lipids and incretin hormones were measured during a breakfast meal test before and after.Results: AXOS significantly changed the microbiota (p=0.05) mainly by increasing Bifidobacterium and decreased microbial alpha-diversity (P<.001) as compared to placebo. Whole-gut and upper intestinal transit were not affected, but stool consistency softened after AXOS (Bristol stool chart score 2.7 ± 0.19 to 3.3 ± 0.19, P<.01). Postprandial fat oxidation tended to increase (iAUC, P=.073) and early GLP-1 response (AUC0-90min, P=.005) was reduced after AXOS. Energy expenditure, plasma metabolites, SCFA concentrations and gut permeability were unchanged. Microbiota could classify responders in improved whole-gut transit after AXOS with an ([ROC] AUC 0.80%).Conclusion: AXOS intake, changed the microbiota, mainly increased fecal Bifidobacterium, tended to increase postprandial fat oxidation and decreased the early GLP-1 response. Whilst we did not observe changes in whole-gut transit time, overall microbiota could accurately classify responders with improved GI transit after AXOS intake.
    Exploring groundwater microbial communities for natural attenuation potential of micropollutants
    Aldas Vargas, A.B. ; Hauptfeld, Tina ; Hermes, G.D.A. ; Atashgahi, S. ; Smidt, H. ; Rijnaarts, H.H.M. ; Sutton, N.B. - \ 2019
    BioRxiv - 29 p.
    Groundwater is a key water resource, with 45.7% of all drinking water globally beingextracted from 15groundwater. Maintaining good groundwater quality is thus crucial to secure drinking water. 16Micropollutants, such as pesticides, threaten groundwater qualitywhich can be mitigated by 17biodegradation.Hence, exploringmicrobial communities in aquifers used for drinking water 18productionis essential for understanding micropollutantsbiodegradation capacity.This study aimed 19at understanding the interaction between groundwater geochemistry, pesticide presence, and 20microbial communities in aquifers used for drinking water production. Two groundwater monitoring 21wellslocated in the northeast of The Netherlands and at 500 m distance from each other were sampled 22in 2014, 2015, 2016 and 2018. In both wells, water was extracted from five discrete depths ranging 23from 13 to 54 m and used to analyze geochemical parameters, pesticide concentrations and microbial 24community dynamics using 16S rRNA gene sequencing and qPCR. Groundwater geochemistry was 25stable throughout the study period and pesticides were heterogeneously distributedat low 26concentrations (μg/L range). Integration of the groundwater chemical and microbial data showed that 27geochemical parameters and pesticides exerted selective pressure on microbial communities. 28Furthermore, microbial communities in both wells showed a more similar composition in the deeper 29part of the aquiferas compared to shallow sections, suggesting vertical differences in hydrological 30connection. This study provides initial insights into microbial communitycomposition and distribution 31in groundwater systems in relation to geochemical parameters. This information can contribute for 32the implementation of bioremediation technologies that guarantee safe drinking water production 33from clean aquifers.
    Assessment of the Accuracy of High-Throughput Sequencing of the ITS1 Region of Neocallimastigomycota for Community Composition Analysis
    Edwards, Joan E. ; Hermes, Gerben D.A. ; Kittelmann, Sandra ; Nijsse, Bart ; Smidt, Hauke - \ 2019
    Frontiers in Microbiology 10 (2019). - ISSN 1664-302X
    anaerobic fungi - clone library - high-throughput sequencing - internal transcribed spacer 1 region - Neocallimastigomycota - size polymorphism

    Anaerobic fungi (Neocallimastigomycota) are common inhabitants of the digestive tract of large mammalian herbivores, where they make an important contribution to plant biomass degradation. The internal transcribed spacer 1 (ITS1) region is currently the molecular marker of choice for anaerobic fungal community analysis, despite its known size polymorphism and heterogeneity. The aim of this study was to assess the accuracy of high-throughput sequencing of the ITS1 region of anaerobic fungi for community composition analysis. To this end, full-length ITS1 clone libraries from five pure cultures, representing the ITS1 region size range, were Sanger sequenced to generate a reference dataset. Barcoded amplicons of the same five pure cultures, and four different mock communities derived from them, were then sequenced using Illumina HiSeq. The resulting sequences were then assessed in relation to either the reference dataset (for the pure cultures) or the corresponding theoretical mock communities. Annotation of sequences obtained from individual pure cultures was not always consistent at the clade or genus level, irrespective of whether data from clone libraries or high-throughput sequencing were analyzed. The detection limit of the high-throughput sequencing method appeared to be influenced by factors other than the parameters used during data processing, as some taxa with theoretical values >0.6% were not detected in the mock communities. The high number of PCR cycles used was considered to be a potential explanation for this observation. Accuracy of two of the four mock communities was limited, and this was speculated to be due to preferential amplification of smaller sized ITS1 regions. If this is true, then this is predicted to be an issue with only six of the 32 named anaerobic fungal clades. Whilst high-throughput sequencing of the ITS1 region from anaerobic fungi can be used for environmental sample analysis, we conclude that the accuracy of the method is influenced by sample community composition. Furthermore, ambiguity in the annotation of sequences within pure cultures due to ITS1 heterogeneity reinforces the limitations of the ITS1 region for the taxonomic assignment of anaerobic fungi. In order to overcome these issues, there is a need to develop an alternative taxonomic marker for anaerobic fungi.

    Sugar Beet Pectin Supplementation Did Not Alter Profiles of Fecal Microbiota and Exhaled Breath in Healthy Young Adults and Healthy Elderly
    An, Ran ; Wilms, Ellen ; Smolinska, Agnieszka ; Hermes, Gerben D.A. ; Masclee, Ad A.M. ; Vos, Paul de; Schols, Henk A. ; Schooten, Frederik J. van; Smidt, Hauke ; Jonkers, Daisy M.A.E. ; Zoetendal, Erwin G. ; Troost, Freddy J. - \ 2019
    Nutrients 11 (2019)9. - ISSN 2072-6643
    aging - dietary fiber - elderly - exhaled air - microbiota - pectin - young adults

    Aging is accompanied with increased frailty and comorbidities, which is potentially associated with microbiome perturbations. Dietary fibers could contribute to healthy aging by beneficially impacting gut microbiota and metabolite profiles. We aimed to compare young adults with elderly and investigate the effect of pectin supplementation on fecal microbiota composition, short chain fatty acids (SCFAs), and exhaled volatile organic compounds (VOCs) while using a randomized, double-blind, placebo-controlled parallel design. Fifty-two young adults and 48 elderly consumed 15 g/day sugar beet pectin or maltodextrin for four weeks. Fecal and exhaled breath samples were collected before and after the intervention period. Fecal samples were used for microbiota profiling by 16S rRNA gene amplicon sequencing, and for analysis of SCFAs by gas chromatography (GC). Breath was used for VOC analysis by GC-tof-MS. Young adults and elderly showed similar fecal SCFA and exhaled VOC profiles. Additionally, fecal microbiota profiles were similar, with five genera significantly different in relative abundance. Pectin supplementation did not significantly alter fecal microbiota, SCFA or exhaled VOC profiles in elderly or young adults. In conclusion, aside from some minor differences in microbial composition, healthy elderly and young adults showed comparable fecal microbiota composition and activity, which were not altered by pectin supplementation.

    Financial development and the efficiency of microfinance institutions
    Hermes, Niels ; Lensink, Robert ; Meesters, Aljar - \ 2018
    In: Research Handbook on Small Business Social Responsibility / Spence, L.J., Frynas, J.G., Muthuri, J.N., Navare, J., Edward Elgar Publishing Ltd. - ISBN 9781784711818 - p. 177 - 205.
    Microfinance is an instrument for achieving responsible finance as its aim is to contribute to the financial inclusion of poor households and small businesses. We investigate whether the country-level financial environment in which microfinance institutions (MFIs) work affects their operations. In particular, we argue that the efficiency of MFIs and their capacity to contribute to increasing financial inclusion of poor households and small businesses is determined by the extent to which financial markets of countries are developed. On the one hand, well-developed financial markets provide an environment in which MFIs are able to flourish and increase their efficiency, which increases their capacity to contribute to financial inclusion. On the other hand, well-developed financial markets may also substitute for MFIs and/or may lead to MFI clients taking up multiple loans, reducing efficiency. We find strong evidence that MFI efficiency is positively associated with the level of financial market development.
    Assessment of the Accuracy of Anaerobic Fungal ITS1 Based Barcoded Amplicon Sequencing for Community Composition Analysis
    Edwards, Joan ; Hermes, Gerben ; Kittelmann, Sandra ; Nijsse, Bart ; Smidt, Hauke - \ 2018
    Wageningen University
    PRJEB29131 - ERP111407 - metagenome
    Background: Anaerobic fungi (Neocallimastigomycetes) are common inhabitants of the digestive tract of mammalian herbivores, where they play an important role in fibre degradation. The internal transcribed spacer 1 (ITS1) region is currently the molecular marker of choice for anaerobic fungal community analysis, despite its known size polymorphism and heterogeneity within pure cultures. The aim of this study was to assess the accurateness of anaerobic fungal ITS1 based barcoded amplicon sequencing for community composition analysis. The assessment was conducted using the following approach. Full-length ITS1 clone libraries from five pure cultures, representing the ITS1 region size range, were Sanger sequenced to generate a reference dataset. Barcoded amplicons of the same five pure cultures, and four different mock communities derived from them, were then sequenced using Illumina HiSeq. The resulting barcoded amplicon sequences were then assessed in relation to either the reference dataset (for the pure cultures) or the corresponding theoretical mock communities (derived from the reference dataset).Results: Annotation of sequences obtained from individual pure cultures was not always consistent at the clade or genus level, irrespective of whether clone libraries or barcoded amplicons were analysed. The minimum detection threshold of the barcoded amplicon method was above 1.5%, presumably due to the high number of PCR cycles used. Accuracy of two of the four mock communities was limited due to preferential amplification of smaller sized ITS1 regions. Of the 32 named anaerobic fungal clades, for which full ITS1 region sequence data is available, this preferential amplification is predicted to be an issue with only 6 of them (Orpinomyces 1a, Orpinomyces 1b, Orpinomyces 2, Cyllamyces 1, Cyllamyces 2 and Buwchfawromyces/SK2).Conclusions: Whilst anaerobic fungal ITS1 based barcoded amplicon sequencing is of value for environmental sample analysis, we conclude that the accuracy of the results is highly dependent on community composition due to preferential amplification of smaller sized ITS1 regions. Studies to date indicate that the D1/D2 region of the 28S rRNA gene is a promising alternative molecular marker. As such, there is an urgent need to obtain more reference sequences and develop an associated working taxonomic scheme in order to enable the suitability of the D1/D2 region as taxonomic marker for anaerobic fungal barcoded amplicon sequencing to be verified.
    NG-Tax, a highly accurate and validated pipeline for analysis of 16S rRNA amplicons from complex biomes
    Ramiro-Garcia, Javier ; Hermes, Gerben D.A. ; Giatsis, Christos ; Sipkema, Detmer ; Zoetendal, Erwin G. ; Schaap, Peter J. ; Smidt, Hauke - \ 2018
    F1000 Research 5 (2018). - ISSN 2046-1402

    Background Massive high-throughput sequencing of short, hypervariable segments of the 16S ribosomal RNA (rRNA) gene has transformed the methodological landscape describing microbial diversity within and across complex biomes. However, several studies have shown that the methodology rather than the biological variation is responsible for the observed sample composition and distribution. This compromises true meta-analyses, although this fact is often disregarded. Results To facilitate true meta-analysis of microbiome studies, we developed NG-Tax, a pipeline for 16S rRNA gene amplicon sequence analysis that was validated with different mock communities and benchmarked against QIIME as the currently most frequently used pipeline. The microbial composition of 49 independently amplified mock samples was characterized by sequencing two variable 16S rRNA gene regions, V4 and V5-V6, in three separate sequencing runs on Illumina's HiSeq2000 platform. This allowed evaluating important factors of technical bias in taxonomic classification: 1) run-to-run sequencing variation, 2) PCR-error, and 3) region/primer specific amplification bias. Despite the short read length (~140 nt) and all technical biases, the average specificity of the taxonomic assignment for the phylotypes included in the mock communities was 96%. On average 99.94% and 92.02% of the reads could be assigned to at least family or genus level, respectively, while assignment to 'spurious genera' represented on average only 0.02% of the reads per sample. Analysis of α- and β-diversity confirmed conclusions guided by biology rather than the aforementioned methodological aspects, which was not the case when samples were analysed using QIIME. Conclusions Different biological outcomes are commonly observed due to 16S rRNA region-specific performance. NG-Tax demonstrated high robustness against choice of region and other technical biases associated with 16S rRNA gene amplicon sequencing studies, diminishing their impact and providing accurate qualitative and quantitative representation of the true sample composition. This will improve comparability between studies and facilitate efforts towards standardization.

    Gut Microbiota and Body Weight in School-Aged Children : The KOALA Birth Cohort Study
    Mbakwa, Catherine A. ; Hermes, Gerben D.A. ; Penders, John ; Savelkoul, Paul H.M. ; Thijs, Carel ; Dagnelie, Pieter C. ; Mommers, Monique ; Zoetendal, Erwin G. ; Smidt, Hauke ; Arts, Ilja C.W. - \ 2018
    Obesity 26 (2018)11. - ISSN 1930-7381 - p. 1767 - 1776.

    Objective: This study aimed to examine the intestinal microbiota composition of school-aged children in association with (over)weight. Methods: The fecal microbiota composition of 295 children was analyzed using the Human Intestinal Tract Chip. Anthropometric outcomes (overweight [BMI ≥ 85th percentile], age- and sex-standardized BMI and weight z scores) were measured at 6 to 7 years of age, and elastic net was used to select genus-like bacterial groups related to all anthropometric outcomes. Subsequently, multiple linear and logistic regression models were used to model associations between selected bacterial groups and anthropometric measures while controlling for confounders. Results: Prevotella melaninogenica, Prevotella oralis, Dialister, and uncultured Clostridiales II (UCII) accounted for 26.1% of the variation in microbiota composition. Several bacterial groups were inversely associated with the anthropometric outcomes: Sutterella wadsworthensis, Marvinbryantia formatexigens, Prevotella melanogenica, P oralis, Burkholderia, uncultured Clostridiales II, and Akkermansia, while Streptococcus bovis was positively associated with overweight. Microbial diversity and richness, and Bacteroidetes to Firmicutes ratio, were not significantly associated with any of the outcomes. Conclusions: In the largest population-based study on childhood gut microbiota and body weight so far, both new and previously identified bacterial groups were found to be associated with overweight. Further research should elucidate their role in energy metabolism.

    Short-Term Microbiota Manipulation and Forearm Substrate Metabolism in Obese Men : A Randomized, Double-Blind, Placebo-Controlled Trial
    Reijnders, Dorien ; Goossens, Gijs H. ; Hermes, Gerben D.A. ; Smidt, Hauke ; Zoetendal, Erwin G. ; Blaak, Ellen E. - \ 2018
    Obesity Facts 11 (2018). - ISSN 1662-4025 - p. 318 - 326.

    Objective: To investigate the impact of gut microbiota manipulation on fasting and postprandial skeletal muscle metabolism in humans. Methods: 40 obese, insulin-resistant males were randomized to amoxicillin (broad-spectrum antibiotic), vancomycin (narrow-spectrum antibiotic), or placebo (7 days, 1,500 mg/day). Before and after treatment, forearm blood flow and metabolite fluxes across forearm muscle were measured under fasting and postprandial (high-fat mixed-meal) conditions. Results: Vancomycin decreased bacterial diversity, reduced the abundance of Gram-positive Firmicutes, and increased the abundance of Gram-negative Proteobacteria, whereas amoxicillin did not affect microbial composition. Neither vancomycin nor amoxicillin treatment affected fasting and postprandial plasma glucose, free fatty acid (FFA), triacylglycerol (TAG), glycerol, lactate, and insulin concentrations or forearm blood flow. Fasting and postprandial net forearm muscle glucose uptake and the release of lactate were not significantly altered by antibiotic treatment as compared to placebo. Finally, antibiotic treatment did not change fasting and postprandial glycerol, FFA, and TAG fluxes across forearm muscle. Conclusion: The present study demonstrates that short-term antibiotic treatment has no effects on fasting and postprandial forearm substrate metabolism and blood flow in obese men with impaired glucose metabolism. These data suggest that short-term strategies targeting the gut microbiota to improve metabolic health may not be effective in obese humans.

    Host and environmental factors affecting the intestinal microbiota in chickens
    Kers, Jannigje G. ; Velkers, Francisca C. ; Fischer, Egil A.J. ; Hermes, Gerben D.A. ; Stegeman, J.A. ; Smidt, Hauke - \ 2018
    Frontiers in Microbiology 9 (2018)FEB. - ISSN 1664-302X
    16S rRNA - Confounding factors - Gut health - Gut microbiota - Microbiome - Poultry
    The initial development of intestinal microbiota in poultry plays an important role in production performance, overall health and resistance against microbial infections. Multiplexed sequencing of 16S ribosomal RNA gene amplicons is often used in studies, such as feed intervention or antimicrobial drug trials, to determine corresponding effects on the composition of intestinal microbiota. However, considerable variation of intestinal microbiota composition has been observed both within and across studies. Such variation may in part be attributed to technical factors, such as sampling procedures, sample storage, DNA extraction, the choice of PCR primers and corresponding region to be sequenced, and the sequencing platforms used. Furthermore, part of this variation in microbiota composition may also be explained by different host characteristics and environmental factors. To facilitate the improvement of design, reproducibility and interpretation of poultry microbiota studies, we have reviewed the literature on confounding factors influencing the observed intestinal microbiota in chickens. First, it has been identified that host-related factors, such as age, sex, and breed, have a large effect on intestinal microbiota. The diversity of chicken intestinal microbiota tends to increase most during the first weeks of life, and corresponding colonization patterns seem to differ between layer- and meat-type chickens. Second, it has been found that environmental factors, such as biosecurity level, housing, litter, feed access and climate also have an effect on the composition of the intestinal microbiota. As microbiota studies have to deal with many of these unknown or hidden host and environmental variables, the choice of study designs can have a great impact on study outcomes and interpretation of the data. Providing details on a broad range of host and environmental factors in articles and sequence data repositories is highly recommended. This creates opportunities to combine data from different studies for meta-analysis, which will facilitate scientific breakthroughs toward nutritional and husbandry associated strategies to improve animal health and performance.
    Improvement of Insulin Sensitivity after Lean Donor Feces in Metabolic Syndrome Is Driven by Baseline Intestinal Microbiota Composition
    Kootte, Ruud S. ; Levin, Evgeni ; Salojärvi, Jarkko ; Smits, Loek P. ; Hartstra, Annick V. ; Udayappan, Shanti D. ; Hermes, Gerben ; Bouter, Kristien E. ; Koopen, Annefleur M. ; Holst, Jens J. ; Knop, Filip K. ; Blaak, Ellen E. ; Zhao, Jing Hua ; Smidt, Hauke ; Harms, Amy C. ; Hankemeijer, Thomas ; Bergman, Jacques J.G.H.M. ; Romijn, Hans A. ; Schaap, Frank G. ; Olde Damink, Steven W.M. ; Ackermans, Mariette T. ; Dallinga-Thie, Geesje M. ; Zoetendal, Erwin ; Vos, Willem M. de; Serlie, Mireille J. ; Stroes, Erik S.G. ; Groen, Albert K. ; Nieuwdorp, Max - \ 2017
    Cell Metabolism 26 (2017)4. - ISSN 1550-4131 - p. 611 - 619.e6.
    fecal microbiota transplantation - insulin sensitivity - intestinal microbiota composition - plasma metabolites

    The intestinal microbiota has been implicated in insulin resistance, although evidence regarding causality in humans is scarce. We therefore studied the effect of lean donor (allogenic) versus own (autologous) fecal microbiota transplantation (FMT) to male recipients with the metabolic syndrome. Whereas we did not observe metabolic changes at 18 weeks after FMT, insulin sensitivity at 6 weeks after allogenic FMT was significantly improved, accompanied by altered microbiota composition. We also observed changes in plasma metabolites such as γ-aminobutyric acid and show that metabolic response upon allogenic FMT (defined as improved insulin sensitivity 6 weeks after FMT) is dependent on decreased fecal microbial diversity at baseline. In conclusion, the beneficial effects of lean donor FMT on glucose metabolism are associated with changes in intestinal microbiota and plasma metabolites and can be predicted based on baseline fecal microbiota composition. Kootte et al. show that fecal microbiota transplantation from lean donors to obese patients with metabolic syndrome improves insulin sensitivity, a transient effect associated with changes in microbiota composition and fasting plasma metabolites. Baseline fecal microbiota composition in recipients predicts the response to lean donor fecal microbiota transplantation.

    Supplementation of Diet With Galacto-oligosaccharides Increases Bifidobacteria, but Not Insulin Sensitivity, in Obese Prediabetic Individuals
    Canfora, Emanuel E. ; Beek, Christina M. van der; Hermes, Gerben D.A. ; Goossens, Gijs H. ; Jocken, Johan W.E. ; Holst, Jens J. ; Eijk, Hans M. van; Venema, Koen ; Smidt, Hauke ; Zoetendal, Erwin G. ; Dejong, Cornelis H.C. ; Lenaerts, Kaatje ; Blaak, Ellen E. - \ 2017
    Gastroenterology 153 (2017)1. - ISSN 0016-5085 - p. 87 - 97.
    Metabolic Control - Microbial Obesity - Prebiotics - Short-Chain Fatty Acids

    Background & Aims: The gut microbiota affects host lipid and glucose metabolism, satiety, and chronic low-grade inflammation to contribute to obesity and type 2 diabetes. Fermentation end products, in particular the short-chain fatty acid (SCFA) acetate, are believed to be involved in these processes. We investigated the long-term effects of supplementation with galacto-oligosaccharides (GOS), an acetogenic fiber, on the composition of the human gut microbiota and human metabolism. Methods: We performed a double-blinded, placebo-controlled, parallel intervention study of 44 overweight or obese (body mass index, 28-40 kg/m2) prediabetic men and women (ages, 45-70 y) from October 2014 through October 2015 in Maastricht, The Netherlands. The participants were assigned randomly to groups who ingested 15 g GOS or isocaloric placebo (maltodextrin) daily with their regular meals for 12 weeks. Before and after this period, we collected data on peripheral and adipose tissue insulin sensitivity, fecal microbiota composition, plasma and fecal SCFA, energy expenditure and substrate oxidation, body composition, and hormonal and inflammatory responses. The primary outcome was the effect of GOS on peripheral insulin sensitivity, measured by the hyperinsulinemic-euglycemic clamp method. Results: Supplementation of diets with GOS, but not placebo, increased the abundance of Bifidobacterium species in feces by 5-fold (P = .009; q = 0.144). Microbial richness or diversity in fecal samples were not affected. We did not observe any differences in fecal or fasting plasma SCFA concentrations or in systemic concentrations of gut-derived hormones, incretins, lipopolysaccharide-binding protein, or other markers of inflammation. In addition, no significant alterations in peripheral and adipose tissue insulin sensitivity, body composition, and energy and substrate metabolism were found. Conclusions: Twelve-week supplementation of GOS selectively increased fecal Bifidobacterium species abundance, but this did not produce significant changes in insulin sensitivity or related substrate and energy metabolism in overweight or obese prediabetic men and women. number, NCT02271776.

    Effects of Gut Microbiota Manipulation by Antibiotics on Host Metabolism in Obese Humans: a Randomized Double-blind Placebo-controlled Trial
    Reijnders, Dorien ; Goossens, Gijs H. ; Neis, Evelien P.J.G. ; Beek, Christina M. van der; Most, Jasper ; Holst, Jens J. ; Lenaerts, Kaatje ; Kootte, Ruud S. ; Nieuwdorp, Max ; Groen, Albert K. ; Boekschoten, Mark ; Hermes, Gerben ; Smidt, Hauke ; Zoetendal, Erwin ; Dejong, Cornelis H.C. ; Blaak, Ellen E. - \ 2016
    Wageningen University
    Homo sapiens - GSE76003 - Homo sapiens - GSE76003 - PRJNA305937
    The gut microbiota has been implicated in obesity and cardiometabolic diseases, although evidence in humans is scarce. We investigated how gut microbiota manipulation by antibiotics (7-day administration of amoxicillin, vancomycin, or placebo) affects host metabolism in 57 obese, prediabetic men. Vancomycin, but not amoxicillin, decreased bacterial diversity and reduced Firmicutes involved in short-chain fatty acid and bile acid metabolism, concomitant with altered plasma and/or fecal metabolite concentrations. Adipose tissue gene expression of oxidative pathways was upregulated by antibiotics, whereas immune-related pathways were downregulated by vancomycin. Antibiotics did not affect tissue-specific insulin sensitivity, energy/substrate metabolism, postprandial hormones and metabolites, systemic inflammation, gut permeability, and adipocyte size. Importantly, energy harvest, adipocyte size, and whole-body insulin sensitivity were not altered at 8-week follow-up, despite a still considerably altered microbial composition, indicating that interference with adult microbiota by 7-day antibiotic treatment has no clinically relevant impact on metabolic health in obese humans.
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