intI1 gene abundance from septic tanks in Thailand using validated intI1 primers.

IMPORTANCE: Antimicrobial resistance is a global crisis, and wastewater treatment, including septic tanks, remains an important source of antimicrobial resistance (AMR) genes. The role of septic tanks in disseminating class 1 integron, and by extension AMR genes, in Thailand, where antibiotic use is unregulated remains understudied. We aimed to monitor gene abundance as a proxy to infer potential AMR from septic tanks in Thailand. We evaluated published intI1 primers due to the lack of consensus on optimal Q-PCR primers and the absence of standardization. Our findings confirmed septic tanks are a source of class 1 integron to the environment. We highlighted the significance of intI1 primer choice, in the context of interpretation of risk associated with AMR spread from septic tanks. We recommend the validated set (F3-R3) for optimal intI1 quantification toward the goal of achieving standardization across studies.

Microbial electrosynthesis of acetate from CO2 in three-chamber cells with gas diffusion biocathode under moderate saline conditions.

The industrial adoption of microbial electrosynthesis (MES) is hindered by high overpotentials deriving from low electrolyte conductivity and inefficient cell designs. In this study, a mixed microbial consortium originating from an anaerobic digester operated under saline conditions (∼13 g L-1 NaCl) was adapted for acetate production from bicarbonate in galvanostatic (0.25 mA cm-2) H-type cells at 5, 10, 15, or 20 g L-1 NaCl concentration. The acetogenic communities were successfully enriched only at 5 and 10 g L-1 NaCl, revealing an inhibitory threshold of about 6 g L-1 Na+. The enriched planktonic communities were then used as inoculum for 3D printed, three-chamber cells equipped with a gas diffusion biocathode. The cells were fed with CO2 gas and operated galvanostatically (0.25 or 1.00 mA cm-2). The highest production rate of 55.4 g m-2 d-1 (0.89 g L-1 d-1), with 82.4% Coulombic efficiency, was obtained at 5 g L-1 NaCl concentration and 1 mA cm-2 applied current, achieving an average acetate production of 44.7 kg MWh-1. Scanning electron microscopy and 16S rRNA sequencing analysis confirmed the formation of a cathodic biofilm dominated by Acetobacterium sp. Finally, three 3D printed cells were hydraulically connected in series to simulate an MES stack, achieving three-fold production rates than with the single cell at 0.25 mA cm-2. This confirms that three-chamber MES cells are an efficient and scalable technology for CO2 bio-electro recycling to acetate and that moderate saline conditions (5 g L-1 NaCl) can help reduce their power demand while preserving the activity of acetogens.

The Effects of Smoking on Human Pharynx Microbiota Composition and Stability.

The oral microbiota is essential to the health of the host, yet little is known about how it responds to disturbances. We examined the oropharyngeal microbiota of 30 individuals over 40 weeks. As the oropharynx is an important gateway to pathogens, and as smoking is associated with increased incidence and severity of respiratory infections, we compared the microbiota of smokers and nonsmokers to shed light on its potential for facilitating infections. We hypothesized that decreased species diversity, decreased community stability, or increased differences in community structure could facilitate invading pathogens. We found that smoking is associated with reduced alpha diversity, greater differences in community structure, and increased environmental filtering. The effects of short-term perturbations (antibiotic use and participants exhibiting cold symptoms) were also investigated. Antibiotic use had a negative effect on alpha diversity, irrespective of smoking status, and both antibiotic use and cold symptoms were associated with highly unique bacterial communities. A stability analysis of models built from the data indicated that there were no differences in local or global stability in the microbial communities of smokers, compared to nonsmokers, and that their microbiota are equally resistant to species invasions. Results from these models suggest that smoker microbiota are perturbed but characterized by alternative stable states that are as stable and invasion-resistant as are the microbiota of nonsmokers. Smoking is unlikely to increase the risk of infectious disease through the altered composition and ecological function of the microbiota; this is more likely due to the effects of smoking on the local and systemic immune system. IMPORTANCE Smoking is associated with an increased risk of respiratory infections. Hypothetically, the altered community diversity of smokers' pharyngeal microbiota, together with changes in their ecological stability properties, could facilitate their invasion by pathogens. To address this question, we analyzed longitudinal microbiota data of baseline healthy individuals who were either smokers or nonsmokers. While the results indicate reduced biodiversity and increased species turnover in the smokers' pharyngeal microbiota, their ecological stability properties were not different from those of the microbiota of nonsmokers, implying, in ecological terms, that the smokers' microbial communities are not less resistant to invasions. Therefore, the study suggests that the increased propensity of respiratory infections that is seen in smokers is more likely associated with changes in the local and systemic immune system than with ecological changes in the microbial communities.

Dynamic gill and mucus microbiomes during a gill disease episode in farmed Atlantic salmon.

Amoebic gill disease (AGD) and complex gill disease (CGD) are recurrent gill disorders in Atlantic salmon, resulting in significant aquaculture losses. The role of gill microbiomes in gill disease development is unclear. We undertook a longitudinal study to characterise the gill tissue and gill mucus microbiomes of farmed Atlantic salmon before, and during, a gill disease episode. Using a newly optimised DNA extraction protocol, we sequenced rRNA genes from microbiomes of gill samples taken from 105 individual salmon on a farm, over a summer season. The AGD aetiological agent, Neoparamoeba perurans, was PCR-quantified targeting 18S rRNA genes. Similar analyses were carried out on mucus samples. Mucus scrapings were suitable, non-lethal substitutes for characterisation of the gill prokaryotic community in this study. Gill tissue and gill mucus microbiomes changed during the campaign, correlating with N. perurans concentrations. Time explained 35% of the gill tissue and gill mucus microbiome variance, while N. perurans concentrations explained 5%. Genera including Dyadobacter, Shewanella and Pedobacter were maximally abundant in gill and mucus samples at the timepoint prior to the the detection of gill disorder signs, at T3. Shewanella was significantly more abundant before than during the gill disease episode, and we suggest this genus could be considered in future studies addressing relationships between gill disease and the gill microbiome.

Ecological Observations Based on Functional Gene Sequencing Are Sensitive to the Amplicon Processing Method.

Until recently, the de facto method for short-read-based amplicon reconstruction was a sequence similarity threshold approach (operational taxonomic units [OTUs]). This has changed with the amplicon sequence variant (ASV) method where distributions are fitted to abundance profiles of individual genes using a noise-error model. While OTU-based approaches are still useful for 16S rRNA/18S rRNA genes, where thresholds of 97% to 99% are used, their use for functional genes is still debatable as there is no consensus on clustering thresholds. Here, we compare OTU- and ASV-based reconstruction approaches and taxonomy assignment methods, the naive Bayesian classifier (NBC) and Bayesian lowest common ancestor (BLCA) algorithm, using a functional gene data set from the microbial nitrogen-cycling community in the Brouage mudflat (France). A range of OTU similarity thresholds and ASVs were used to compare amoA (ammonia-oxidizing archaea [AOA] and ammonia-oxidizing bacteria [AOB]), nxrB, nirS, nirK, and nrfA communities between differing sedimentary structures. Significant effects of the sedimentary structure on weighted UniFrac (WUniFrac) distances were observed for AOA amoA when using ASVs, an OTU at a threshold of 97% sequence identity (OTU-97%), and OTU-85%; AOB amoA when using OTU-85%; and nirS when using ASV, OTU-90%, and OTU-85%. For AOB amoA, significant effects of the sedimentary structures on UniFrac distances were observed when using OTU-97% but not ASVs, and the inverse was found for nrfA. Interestingly, conclusions drawn for nirK and nxrB were consistent between amplicon reconstruction methods. We also show that when the sequences in the reference database are related to the environment in question, the BLCA algorithm leads to more phylogenetically relevant classifications. However, when the reference database contains sequences more dissimilar to the ones retrieved, the NBC obtains more information. IMPORTANCE Several analysis pipelines are available to microbial ecologists to process amplicon sequencing data, yet to date, there is no consensus as to the most appropriate method, and it becomes more difficult for genes that encode a specific function (functional genes). Standardized approaches need to be adopted to increase the reliability and reproducibility of environmental amplicon-sequencing-based data sets. In this paper, we argue that the recently developed ASV approach offers a better opportunity to achieve such standardization than OTUs for functional genes. We also propose a comprehensive framework for quality filtering of the sequencing reads based on protein sequence verification.

Microbiomes in drinking water treatment and distribution: A meta-analysis from source to tap.

A meta-analysis of existing and available Illumina 16S rRNA datasets from drinking water source, treatment and drinking water distribution systems (DWDS) were collated to compare changes in abundance and diversity throughout. Samples from bulk water and biofilm were used to assess principles governing microbial community assembly and the value of amplicon sequencing to water utilities. Individual phyla relationships were explored to identify competitive or synergistic factors governing DWDS microbiomes. The relative importance of stochasticity in the assembly of the DWDS microbiome was considered to identify the significance of source and treatment in determining communities in DWDS. Treatment of water significantly reduces overall species abundance and richness, with chlorination of water providing the most impact to individual taxa relationships. The assembly of microbial communities in the bulk water of the source, primary treatment process and DWDS is governed by more stochastic processes, as is the DWDS biofilm. DWDS biofilm is significantly different from bulk water in terms of local contribution to beta diversity, type and abundance of taxa present. Water immediately post chlorination has a more deterministic microbial assembly, highlighting the significance of this process in changing the microbiome, although elevated levels of stochasticity in DWDS samples suggest that this may not be the case at customer taps. 16S rRNA sequencing is becoming more routine, and may have several uses for water utilities, including: detection and risk assessment of potential pathogens such as those within the genera of Legionella and Mycobacterium; assessing the risk of nitrification in DWDS; providing improved indicators of process performance and monitoring for significant changes in the microbial community to detect contamination. Combining this with quantitative methods like flow cytometry will allow a greater depth of understanding of the DWDS microbiome.

MdaB and NfrA, Two Novel Reductases Important in the Survival and Persistence of the Major Enteropathogen Campylobacter jejuni.

The paralogues RrpA and RrpB, which are members of the MarR family of DNA binding proteins, are important for the survival of the global bacterial foodborne pathogen Campylobacter jejuni under redox stress. We report that RrpA is a positive regulator of mdaB, encoding a flavin-dependent quinone reductase that contributes to the protection from redox stress mediated by structurally diverse quinones, while RrpB negatively regulates the expression of cj1555c (renamed nfrA for NADPH-flavin reductase A), encoding a flavin reductase. NfrA reduces riboflavin at a greater rate than its derivatives, suggesting that exogenous free flavins are the natural substrate. MdaB and NfrA both prefer NADPH as an electron donor. Cysteine substitution and posttranslational modification analyses indicated that RrpA and RrpB employ a cysteine-based redox switch. Complete genome sequence analyses revealed that mdaB is frequently found in Campylobacter and related Helicobacter spp., while nfrA is predominant in C. jejuni strains. Quinones and flavins are redox cycling agents secreted by a wide range of cell types that can form damaging superoxide by one-electron reactions. We propose a model for stress adaptation where MdaB and NfrA facilitate a two-electron reduction mechanism to the less toxic hydroquinones, thus aiding survival and persistence of this major pathogen. IMPORTANCE Changes in cellular redox potential result in alteration in the oxidation state of intracellular metabolites and enzymes; consequently, cells make adjustments that favor growth and survival. The work we present here answers some of the many questions that have remained elusive over the years of investigation into the enigmatic microaerophile bacterium Campylobacter jejuni. We employed molecular approaches to understand the regulation mechanisms and functional analyses to reveal the roles of two novel quinone and flavin reductases; both serve as major pools of cellular redox-active molecules. This work extends our knowledge on bacterial redox sensing mechanisms and the significance of hemostasis.

Putative Antimicrobial Peptides Within Bacterial Proteomes Affect Bacterial Predominance: A Network Analysis Perspective.

The predominance of bacterial taxa in the gut, was examined in view of the putative antimicrobial peptide sequences (AMPs) within their proteomes. The working assumption was that compatible bacteria would share homology and thus immunity to their putative AMPs, while competing taxa would have dissimilarities in their proteome-hidden AMPs. A network-based method ("Bacterial Wars") was developed to handle sequence similarities of predicted AMPs among UniProt-derived protein sequences from different bacterial taxa, while a resulting parameter ("Die" score) suggested which taxa would prevail in a defined microbiome. T he working hypothesis was examined by correlating the calculated Die scores, to the abundance of bacterial taxa from gut microbiomes from different states of health and disease. Eleven publicly available 16S rRNA datasets and a dataset from a full shotgun metagenomics served for the analysis. The overall conclusion was that AMPs encrypted within bacterial proteomes affected the predominance of bacterial taxa in chemospheres.

Dietary triggers of gut inflammation following exclusive enteral nutrition in children with Crohn's disease: a pilot study.

BACKGROUND: The anti-inflammatory effect of exclusive enteral nutrition on the gut of children with Crohn's disease is rapidly lost after food reintroduction. This study assessed disease dietary triggers following successful treatment with exclusive enteral nutrition. METHODS: Nutrient intake, dietary patterns and dietary biomarkers in faeces (gluten immunogenic peptides, undigestible starch, short chain fatty acids) were assessed in 14 children with Crohn's disease during early food reintroduction, following exclusive enteral nutrition. Groups above (Group A) and below (Group B) the median levels of faecal calprotectin after food reintroduction were assigned for comparative analysis. RESULTS: Intakes of fibre, gluten-containing cereals and red and processed meat were significantly higher in Group A than Group B; (median [Q1, Q3], g/day; Fibre: 12.1 [11.2, 19.9] vs. 9.9 [7.6, 12.1], p = 0.03; Red and processed meat: 151 [66.7, 190] vs. 63.3 [21.7, 67], p = 0.02; gluten-containing cereals: 289 [207, 402] vs. 203 [61, 232], p = 0.035). A diet consisting of cereals and meat products was predictive (92% accuracy) of higher faecal calprotectin levels after food reintroduction. In faeces, butyrate levels, expressed as absolute concentration and relative abundance, were higher in Group A than Group B by 28.4 µmol/g (p = 0.015) and 6.4% (p = 0.008), respectively. Levels of gluten immunogenic peptide and starch in faeces did not differ between the two groups. CONCLUSIONS: This pilot study identified potential dietary triggers of gut inflammation in children with Crohn's disease after food reintroduction following treatment with exclusive enteral nutrition. TRIAL REGISTRATION: Clinical trials.gov registration number: NCT02341248; Clinical trials.gov URL: https://clinicaltrials.gov/ct2/show/NCT02341248 (retrospectively registered).

The mannose receptor (CD206) identifies a population of colonic macrophages in health and inflammatory bowel disease.

To understand the contribution of mononuclear phagocytes (MNP), which include monocyte-derived intestinal macrophages, to the pathogenesis of inflammatory bowel disease (IBD), it is necessary to identify functionally-different MNP populations. We aimed to characterise intestinal macrophage populations in patients with IBD. We developed 12-parameter flow cytometry protocols to identify and human intestinal MNPs. We used these protocols to purify and characterize colonic macrophages from colonic tissue from patients with Crohn's disease (CD), ulcerative colitis (UC), or non-inflamed controls, in a cross-sectional study. We identify macrophage populations (CD45+CD64+ HLA-DR+) and describe two distinct subsets, differentiated by their expression of the mannose receptor, CD206. CD206+ macrophages expressed markers consistent with a mature phenotype: high levels of CD68 and CD163, higher transcription of IL-10 and lower expression of TREM1. CD206- macrophages appear to be less mature, with features more similar to their monocytic precursors. We identified and purified macrophage populations from human colon. These appear to be derived from a monocytic precursor with high CCR2 and low CD206 expression. As these cells mature, they acquire expression of IL-10, CD206, CD63, and CD168. Targeting the newly recruited monocyte-derived cells may represent a fruitful avenue to ameliorate chronic inflammation in IBD.

Inter-kingdom relationships in Crohn's disease explored using a multi-omics approach.

The etiology of Crohn's disease (CD) is multifactorial. Bacterial and fungal microbiota are involved in the onset and/or progression of the disease. A bacterial dysbiosis in CD patients is accepted; however, less is known about the mycobiome and the relationships between the two communities. We investigated the interkingdom relationships, their metabolic consequences, and the changes in the fungal community during relapse and remission in CD.Two cohorts were evaluated: a British cohort (n = 63) comprising CD and ulcerative colitis patients, and controls. The fungal and bacterial communities of biopsy and fecal samples were analyzed, with the fecal volatiles; datasets were also integrated; and a Dutch cohort (n = 41) comprising CD patients and healthy controls was analyzed for stability of the gut mycobiome.A dysbiosis of the bacterial community was observed in biopsies and stool. Results suggest Bacteroides is likely key in CD and may modulate Candida colonization. A dysbiosis of the fungal community was observed only in the Dutch cohort; Malassezia and Candida were increased in patients taking immunosuppressants. Longitudinal analysis showed an increase in Cyberlindnera in relapse. Saccharomyces was dominant in all fecal samples, but not in biopsies, some of which did not yield fungal reads; amino acid degradation was the main metabolic change associated with CD and both bacteria and fungi might be implicated.We have shown that Bacteroides and yeasts may play a role in CD; understanding their role and relationship in the disease would shed new light on the development and treatment of CD.

Oral Ferric Maltol Does Not Adversely Affect the Intestinal Microbiome of Patients or Mice, But Ferrous Sulphate Does.

BACKGROUND AND AIMS: Altering dietary ferrous sulphate (FS) consumption exacerbates a murine model of colitis and alters the intestinal microbiome. We investigated the impact of oral ferric maltol (FM) and FS on mice with dextran sodium sulphate (DSS) induced colitis, and the microbiome of patients with iron deficiency. METHODS: Mice had acute colitis induced, with 2% DSS for 5 days, followed by water. During this period, groups of mice were fed standard chow (200 ppm iron, SC, n = 8), or SC with 200ppm FS supplementation (n = 16, FSS), or SC with 200 ppm FM supplementation (n = 16, FMS). Clinical, pathological and microbiome assessments were compared at days 1 and 10. Fecal bacterial gDNA was extracted and the microbiome assessed by sequencing. Statistical inferences were made using MacQIIME. Principal Coordinates Analysis were used to visualize beta-diversity cluster analysis. Ten patients with IDA were treated with FS, and six with inactive inflammatory bowel disease received FM, supplements for four weeks: pre- and mid-treatment fecal samples were collected: the microbiome was assessed (see above). RESULTS: In mice, after DSS treatment, there was a decrease in many genera in the SC and FSS groups: Lactobacillales increased in mice that received FMS. In humans, FS treatment led to an increase in five genera, but FM was not associated with any measurable change. The severity of DSS-induced colitis was greater with FSS than FMS. CONCLUSIONS: This study demonstrates differential and unique influences of ferric maltol and ferrous sulphate supplements on intestinal microbiota. These differences might contribute to the different side effects associated with these preparations.

Socioeconomic position links circulatory microbiota differences with biological age.

Imbalanced nutrition is associated with accelerated ageing, possibly mediated by microbiota. An analysis of the circulatory microbiota obtained from the leukocytes of participants in the MRC Twenty-07 general population cohort was performed. We now report that in this cohort, the most biologically aged exhibit a significantly higher abundance of circulatory pathogenic bacteria, including Neisseria, Rothia and Porphyromonas, while those less biologically aged possess more circulatory salutogenic (defined as being supportive of human health and wellbeing) bacteria, including Lactobacillus, Lachnospiraceae UCG-004 and Kocuria. The presence of these salutogenic bactreria is consistent with a capacity to metabolise and produce Nrf2 agonists. We also demonstrate that associated one carbon metabolism, notably betaine levels, did not vary with chronological age, but displayed a difference with socioeconomic position (SEP). Those at lower SEP possessed significantly lower betaine levels indicative of a poorer diet and poorer health span and consistent with reduced global DNA methylation levels in this group. Our data suggest a clear route to improving age related health and resilience based on dietary modulation of the microbiota.

Nonlinear machine learning pattern recognition and bacteria-metabolite multilayer network analysis of perturbed gastric microbiome.

The stomach is inhabited by diverse microbial communities, co-existing in a dynamic balance. Long-term use of drugs such as proton pump inhibitors (PPIs), or bacterial infection such as Helicobacter pylori, cause significant microbial alterations. Yet, studies revealing how the commensal bacteria re-organize, due to these perturbations of the gastric environment, are in early phase and rely principally on linear techniques for multivariate analysis. Here we disclose the importance of complementing linear dimensionality reduction techniques with nonlinear ones to unveil hidden patterns that remain unseen by linear embedding. Then, we prove the advantages to complete multivariate pattern analysis with differential network analysis, to reveal mechanisms of bacterial network re-organizations which emerge from perturbations induced by a medical treatment (PPIs) or an infectious state (H. pylori). Finally, we show how to build bacteria-metabolite multilayer networks that can deepen our understanding of the metabolite pathways significantly associated to the perturbed microbial communities.

Carboxylic acids production and electrosynthetic microbial community evolution under different CO2 feeding regimens.

Microbial electrosynthesis (MES) is a potential technology for CO2 recycling, but insufficient information is available on the microbial interactions underpinning electrochemically-assisted reactions. In this study, a MES reactor was operated for 225 days alternately with bicarbonate or CO2 as carbon source, under batch or continuous feeding regimens, to evaluate the response of the microbial communities, and their productivity, to dynamic operating conditions. A stable acetic acid production rate of 9.68 g m-2 d-1, and coulombic efficiency up to 40%, was achieved with continuous CO2 sparging, higher than the rates obtained with bicarbonate (0.94 g m-2 d-1) and CO2 under fed-batch conditions (2.54 g m-2 d-1). However, the highest butyric acid production rate (0.39 g m-2 d-1) was achieved with intermittent CO2 sparging. The microbial community analyses focused on differential amplicon sequence variants (ASVs), allowing detection of ASVs significantly different across consecutive samples. This analysis, combined with co-occurence network analysis, and cyclic voltammetry, indicated that hydrogen-mediated acetogenesis was carried out by Clostridium, Eubacterium and Acetobacterium, whereas Oscillibacter and Caproiciproducens were involved in butyric acid production. The cathodic community was spatially inhomogeneous, with potential electrotrophs, such as Sulfurospirillum and Desulfovibrio, most prevalent near the current collector. The abundance of Sulfurospirillum positively correlated with that of Acetobacterium, supporting the syntrophic metabolism of both organisms.

Alterations in Intestinal Microbiota of Children With Celiac Disease at the Time of Diagnosis and on a Gluten-free Diet.

BACKGROUND AND AIMS: It is not clear whether alterations in the intestinal microbiota of children with celiac disease (CD) cause the disease or are a result of disease and/or its treatment with a gluten-free diet (GFD). METHODS: We obtained 167 fecal samples from 141 children (20 with new-onset CD, 45 treated with a GFD, 57 healthy children, and 19 unaffected siblings of children with CD) in Glasgow, Scotland. Samples were analyzed by 16S ribosomal RNA sequencing, and diet-related metabolites were measured by gas chromatography. We obtained fecal samples from 13 children with new-onset CD after 6 and 12 months on a GFD. Relationships between microbiota with diet composition, gastrointestinal function, and biomarkers of GFD compliance were explored. RESULTS: Microbiota α diversity did not differ among groups. Microbial dysbiosis was not observed in children with new-onset CD. In contrast, 2.8% (Bray-Curtis dissimilarity index, P = .025) and 2.5% (UniFrac distances, P = .027) of the variation in microbiota composition could be explained by the GFD. Between 3% and 5% of all taxa differed among all group comparisons. Eleven distinctive operational taxonomic units composed a microbe signature specific to CD with high diagnostic probability. Most operational taxonomic units that differed between patients on a GFD with new-onset CD vs healthy children were associated with nutrient and food group intake (from 75% to 94%) and with biomarkers of gluten ingestion. Fecal levels of butyrate and ammonia decreased during the GFD. CONCLUSIONS: Although several alterations in the intestinal microbiota of children with established CD appear to be effects of a GFD, specific bacteria were found to be distinct biomarkers of CD. Studies are needed to determine whether these bacteria contribute to pathogenesis of CD.

Differential prevalence and host-association of antimicrobial resistance traits in disinfected and non-disinfected drinking water systems.

Antimicrobial resistance (AMR) in drinking water has received less attention than its counterparts in the urban water cycle. While culture-based techniques or gene-centric PCR have been used to probe the impact of treatment approaches (e.g., disinfection) on AMR in drinking water, to our knowledge there is no systematic comparison of AMR trait distribution and prevalence between disinfected and disinfectant residual-free drinking water systems. We used metagenomics to assess the associations between disinfectant residuals and AMR prevalence and its host association in full-scale drinking water distribution systems (DWDSs) with and without disinfectant residuals. While the differences in AMR profiles between DWDSs were associated with the presence or absence of disinfectant, they were also associated with overall water chemistry and more importantly with microbial community structure. AMR genes and mechanisms differentially abundant in disinfected systems were primarily associated with nontuberculous mycobacteria (NTM). Finally, evaluation of metagenome assembled genomes (MAGs) also suggests that NTM possessing AMR genes conferring intrinsic resistance to key antibiotics were prevalent in disinfected systems, whereas such NTM genomes were not detected in disinfectant residual free DWDSs. Altogether, our findings provide insights into the drinking water resistome and its association with potential opportunistic pathogens, particularly in systems with disinfectant residual.

Dietary Strategies for Maintenance of Clinical Remission in Inflammatory Bowel Diseases: Are We There Yet?

The etiopathogenesis of Inflammatory bowel disease (IBD) is a result of a complex interaction between host immune response, the gut microbiome and environmental factors, such as diet. Although scientific advances, with the use of biological medications, have revolutionized IBD treatment, the challenge for maintaining clinical remission and delaying clinical relapse is still present. As exclusive enteral nutrition has become a well-established treatment for the induction of remission in pediatric Crohn's disease, the scientific interest regarding diet in IBD is now focused on the development of follow-on dietary strategies, which aim to suppress colonic inflammation and delay a disease flare. The objective of this review is to present an extensive overview of the dietary strategies, which have been used in the literature to maintain clinical remission in both Crohn's disease and Ulcerative colitis, and the evidence surrounding the association of dietary components with clinical relapse. We also aim to provide study-related recommendations to be encompassed in future research studies aiming to investigate the role of diet during remission periods in IBD.

Reverse transcriptase enzyme and priming strategy affect quantification and diversity of environmental transcripts.

Reverse-transcriptase-quantitative PCR (RT-Q-PCR) and RT-PCR amplicon sequencing, provide a convenient, target-specific, high-sensitivity approach for gene expression studies and are widely used in environmental microbiology. Yet, the effectiveness and reproducibility of the reverse transcription step has not been evaluated. Therefore, we tested a combination of four commercial reverse transcriptases with two priming techniques to faithfully transcribe 16S rRNA and amoA transcripts from marine sediments. Both enzyme and priming strategy greatly affected quantification of the exact same target with differences of up to 600-fold. Furthermore, the choice of RT system significantly changed the communities recovered. For 16S rRNA, both enzyme and priming had a significant effect with enzyme having a stronger impact than priming. Inversely, for amoA only the change in priming strategy resulted in significant differences between the same samples. Specifically, more OTUs and better coverage of amoA transcripts diversity were obtained with GS priming indicating this approach was better at recovering the diversity of amoA transcripts. Moreover, sequencing of RNA mock communities revealed that, even though transcript α diversities (i.e., OTU counts within a sample) can be biased by the RT, the comparison of ß diversities (i.e., differences in OTU counts between samples) is reliable as those biases are reproducible between environments.

Analysis of 61 exclusive enteral nutrition formulas used in the management of active Crohn's disease-new insights into dietary disease triggers.

BACKGROUND: Exclusive enteral nutrition (EEN) is an effective treatment for Crohn's disease. AIMS: To investigate the hypothesis that ingredients of EEN formulas are unlikely to initiate a disease flare and that their dietary elimination is not essential for disease amelioration. METHODS: We performed compositional analysis of EEN formulas with evidence of efficacy in management of active Crohn's disease. Macronutrient content was compared against the dietary reference values (DRV), the UK National Diet and Nutrition Survey (NDNS) and intake of Crohn's disease children. Food additives were cross-referenced against the FAO/WHO database. RESULTS: Sixty-one formulas were identified with variable composition (carbohydrates [22.8%-89.3%], protein [7.8%-30.1%], fat [0%-52.5%]). Maltodextrin, milk protein and vegetable/plant oils were the commonest macronutrient sources. Their n-6:n-3 fatty acid ratio varied from 0.25 to 46.5. 56 food additives were identified (median per formula: 11). All formulas were lactose-free, gluten-free, and 82% lacked fibre. The commonest food additives were emulsifiers, stabilisers, antioxidants, acidity regulators and thickeners. Food additives, implicated in Crohn's disease aetiology, were present in formulas (modified starches [100%], carrageenan [22%], carboxymethyl cellulose [13%] and polysorbate 80 [5%]). Remission rates did not differ between EEN formulas with and without those food additives. Analysis including only formulas from randomised controlled trials (RCTs) retained in the latest Cochrane meta-analysis produced similar findings. EEN formulas contained less energy from saturated fat than NDNS intake. CONCLUSION: We have identified food ingredients which are present in EEN formulas that are effective in Crohn's disease and challenge perceptions that these ingredients might be harmful.