Single-cell approaches in human microbiome research.

Microbial culturing and meta-omic profiling technologies have significantly advanced our understanding of the taxonomic and functional variation of the human microbiome and its impact on host processes. The next increase in resolution will come by understanding the role of low-abundant and less-prevalent bacteria and the study of individual cell behaviors that underlie the complexity of microbial ecosystems. To this aim, single-cell techniques are being rapidly developed to isolate, culture, and characterize the genomes and transcriptomes of individual microbes in complex communities. Here, we discuss how these single-cell technologies are providing unique insights into the biology and behavior of human microbiomes.

High Acetate Concentration Protects Intestinal Barrier and Exerts Anti-Inflammatory Effects in Organoid-Derived Epithelial Monolayer Cultures from Patients with Ulcerative Colitis.

Short-chain fatty acids as well as their bacterial producers are of increasing interest in inflammatory bowel diseases. Although less studied compared to butyrate, acetate might also be of interest as it may be less toxic to epithelial cells, stimulate butyrate-producing bacteria by cross-feeding, and have anti-inflammatory and barrier-protective properties. Moreover, one of the causative factors of the probiotic potency of Saccharomyces cerevisae var. boulardii is thought to be its high acetate production. Therefore, the objective was to preclinically assess the effects of high acetate concentrations on inflammation and barrier integrity in organoid-based monolayer cultures from ulcerative colitis patients. Confluent organoid-derived colonic epithelial monolayers (n = 10) were exposed to basolateral inflammatory stimulation or control medium. After 24 h, high acetate or control medium was administered apically for an additional 48 h. Changes in TEER were measured after 48 h. Expression levels of barrier genes and inflammatory markers were determined by qPCR. Pro-inflammatory proteins in the supernatant were quantified using the MSD platform. Increased epithelial resistance was observed with high acetate administration in both inflamed and non-inflamed conditions, together with decreased expression levels of IL8 and TNFα and CLDN1. Upon high acetate administration to inflamed monolayers, upregulation of HIF1α, MUC2, and MKI67, and a decrease of the majority of pro-inflammatory cytokines was observed. In our patient-derived human epithelial cell culture model, a protective effect of high acetate administration on epithelial resistance, barrier gene expression, and inflammatory protein production was observed. These findings open up new possibilities for acetate-mediated management of barrier defects and inflammation in IBD.

Treponema peruense sp. nov., a commensal spirochaete isolated from human faeces.

A Gram-stain-negative, obligatory anaerobic spirochaete (RCC2812T) was isolated from a faecal sample obtained from an individual residing in a remote Amazonian community in Peru. The bacterium showed highest 16S rRNA gene sequence similarity to the pig intestinal spirochete Treponema succinifaciens (89.48 %). Average nucleotide identity values between strain RCC2812T and all available Treponema genomes from validated type strains were all <73 %, thus clearly lower than the species delineation threshold. The DNA G+C content of RCC2812T was 41.24 mol%. Phenotypic characterization using the API-ZYM and API 20A systems confirmed the divergent position of this bacterium within the genus Treponema. Strain RCC2812T could be differentiated from the phylogenetically most closely related T. succinifaciens by the presence of alkaline phosphatase and α -glucosidase activities. Unlike T. succinifaciens, strain RCC2812T grew equally well with or without serum. Strain RCC2812T is the first commensal Treponema isolated from the human faecal microbiota of remote populations, and based on the collected data represents a novel Treponema species for which the name Treponema peruense sp. nov. is proposed. The type strain is RCC2812T (=LMG 31794T=CIP 111910T).

Isolation and Quantification of Uremic Toxin Precursor-Generating Gut Bacteria in Chronic Kidney Disease Patients.

In chronic kidney disease (CKD), impaired kidney function results in accumulation of uremic toxins, which exert deleterious biological effects and contribute to inflammation and cardiovascular morbidity and mortality. Protein-bound uremic toxins (PBUTs), such as p-cresyl sulfate, indoxyl sulfate and indole-3-acetic acid, originate from phenolic and indolic compounds, which are end products of gut bacterial metabolization of aromatic amino acids (AAA). This study investigates gut microbial composition at different CKD stages by isolating, identifying and quantifying PBUT precursor-generating bacteria. Fecal DNA extracts from 14 controls and 138 CKD patients were used to quantify total bacterial number and 11 bacterial taxa with qPCR. Moreover, isolated bacteria from CKD 1 and CKD 5 fecal samples were cultured in broth medium supplemented with AAA under aerobic and anaerobic conditions, and classified as PBUT precursor-generators based on their generation capacity of phenolic and indolic compounds, measured with U(H)PLC. In total, 148 different fecal bacterial species were isolated, of which 92 were PBUT precursor-generators. These bacterial species can be a potential target for reducing PBUT plasma levels in CKD. qPCR indicated lower abundance of short chain fatty acid-generating bacteria, Bifidobacterium spp. and Streptococcus spp., and higher Enterobacteriaceae and E. coli with impaired kidney function, confirming an altered gut microbial composition in CKD.

Specific members of the predominant gut microbiota predict pouchitis following colectomy and IPAA in UC.

OBJECTIVE: Pouchitis is the most common complication after colectomy with ileal pouch-anal anastomosis (IPAA) for UC and the risk is the highest within the 1st year after surgery. The pathogenesis is not completely understood but clinical response to antibiotics suggests a role for gut microbiota. We hypothesised that the risk for pouchitis can be predicted based on the faecal microbial composition before colectomy. DESIGN: Faecal samples from 21 patients with UC undergoing IPAA were prospectively collected before colectomy and at predefined clinical visits at 1 month, 3 months, 6 months and 12 months after IPAA. The predominant microbiota was analysed using community profiling with denaturing gradient gel electrophoresis followed by quantitative real-time PCR validation. RESULTS: Cluster analysis before colectomy distinguished patients with pouchitis from those with normal pouch during the 1st year of follow-up. In patients developing pouchitis, an increase of Ruminococcus gnavus (p<0.001), Bacteroides vulgatus (p=0.043), Clostridium perfringens (p=0.011) and a reduction of two Lachnospiraceae genera (Blautia (p=0.04), Roseburia (p=0.008)) was observed. A score combining these five bacterial risk factors was calculated and presence of at least two risk factors showed a sensitivity and specificity of 100% and 63.6%, respectively. CONCLUSIONS: Presence of R. gnavus, B. vulgatus and C. perfringens and absence of Blautia and Roseburia in faecal samples of patients with UC before surgery is associated with a higher risk of pouchitis after IPAA. Our findings suggest new predictive and therapeutic strategies in patients undergoing colectomy with IPAA.

Sphingobacterium cellulitidis sp. nov., isolated from clinical and environmental sources.

The taxonomic position of two isolates belonging to the genus Sphingobacterium was determined. The first isolate, R-53603T, was obtained from purulent discharge from the toe of a cellulitis patient in Kuwait. Comparative 16S rRNA gene sequence analysis revealed 99.87 % similarity of R-53603T with environmental isolate P031 (=R-53745) originating from activated sludge in Singapore. The two isolates were phylogenetically positioned on the same sub-branch. Highest 16S rRNA gene sequence similarity was found with the type strains of Sphingobacterium mizutaii (98.23 %), Sphingobacterium lactis (97.78 %) and Sphingobacterium daejeonense (97.14 %). DNA-DNA hybridizations revealed <70 % relatedness between the two isolates and the type strains of the close phylogenetic neighbours S. mizutaii(18.0-24.5 %), S. lactis(20.3-25.9 %) and S. daejeonense(13.2-20.0 %). The high relative contribution of iso-C15 : 0, iso-C17 : 0 3-OH and summed feature 3 (iso-C15 : 0 2-OH and/or C16 : 1ω7c) in the cellular fatty acid profiles of R-53603T and R-53745, the presence of sphingophospholipids, MK-7 as the dominant menaquinone and phosphatidylethanolamine as the major polar lipid in strain R-53603T are typical chemotaxonomic characteristics for members of the genus Sphingobacterium. Phenotypic features most useful for differentiation of the two novel strains from the most closely related species S. mizutaii include growth on MacConkey agar, and utilization of stachyose, guanidine HCl and lithium chloride in Biolog GEN III tests. Strains R-53603T and R-53745 thus represent a novel species, for which the name Sphingobacterium cellulitidis sp. nov. is proposed. The type strain is R-53603T (=LMG 28764T=DSM 102028T).

Acinetobacter dijkshoorniae sp. nov., a member of the Acinetobacter calcoaceticus-Acinetobacter baumannii complex mainly recovered from clinical samples in different countries.

The recent advances in bacterial species identification methods have led to the rapid taxonomic diversification of the genus Acinetobacter. In the present study, phenotypic and molecular methods have been used to determine the taxonomic position of a group of 12 genotypically distinct strains belonging to the Acinetobacter calcoaceticus-Acinetobacter baumannii (ACB) complex, initially described by Gerner-Smidt and Tjernberg in 1993, that are closely related to Acinetobacter pittii. Strains characterized in this study originated mostly from human samples obtained in different countries over a period of 15 years. rpoB gene sequences and multilocus sequence typing were used for comparisons against 94 strains representing all species included in the ACB complex. Cluster analysis based on such sequences showed that all 12 strains grouped together in a distinct clade closest to Acinetobacter pittiithat was supported by bootstrap values of 99 %. Values of average nucleotide identity based on blast between the genome sequence of strain JVAP01T (NCBI accession no. LJPG00000000) and those of other species from the ACB complex were always <91.2 %, supporting the species status of the group. In addition, the metabolic characteristics of the group matched those of the ACB complex and the analysis of their protein signatures by matrix-assisted laser desorption ionization time-of-flight MS identified some specific peaks. Our results support the designation of these strains as representing a novel species, for which the name Acinetobacter dijkshoorniae sp. nov. is proposed. The type strain is JVAP01T (=CECT 9134T=LMG 29605T).

Wheat bran extract alters colonic fermentation and microbial composition, but does not affect faecal water toxicity: a randomised controlled trial in healthy subjects.

Wheat bran extract (WBE), containing arabinoxylan-oligosaccharides that are potential prebiotic substrates, has been shown to modify bacterial colonic fermentation in human subjects and to beneficially affect the development of colorectal cancer (CRC) in rats. However, it is unclear whether these changes in fermentation are able to reduce the risk of developing CRC in humans. The aim of the present study was to evaluate the effects of WBE on the markers of CRC risk in healthy volunteers, and to correlate these effects with colonic fermentation. A total of twenty healthy subjects were enrolled in a double-blind, cross-over, randomised, controlled trial in which the subjects ingested WBE (10 g/d) or placebo (maltodextrin, 10 g/d) for 3 weeks, separated by a 3-week washout period. At the end of each study period, colonic handling of NH3 was evaluated using the biomarker lactose[15N, 15N']ureide, colonic fermentation was characterised through a metabolomics approach, and the predominant microbial composition was analysed using denaturing gradient gel electrophoresis. As markers of CRC risk, faecal water genotoxicity was determined using the comet assay and faecal water cytotoxicity using a colorimetric cell viability assay. Intake of WBE induced a shift from urinary to faecal 15N excretion, indicating a stimulation of colonic bacterial activity and/or growth. Microbial analysis revealed a selective stimulation of Bifidobacterium adolescentis. In addition, WBE altered the colonic fermentation pattern and significantly reduced colonic protein fermentation compared with the run-in period. However, faecal water cytotoxicity and genotoxicity were not affected. Although intake of WBE clearly affected colonic fermentation and changed the composition of the microbiota, these changes were not associated with the changes in the markers of CRC risk.

Phylogenetic analysis of faecal microbiota from captive cheetahs reveals underrepresentation of Bacteroidetes and Bifidobacteriaceae.

BACKGROUND: Imbalanced feeding regimes may initiate gastrointestinal and metabolic diseases in endangered felids kept in captivity such as cheetahs. Given the crucial role of the host's intestinal microbiota in feed fermentation and health maintenance, a better understanding of the cheetah's intestinal ecosystem is essential for improvement of current feeding strategies. We determined the phylogenetic diversity of the faecal microbiota of the only two cheetahs housed in an EAZA associated zoo in Flanders, Belgium, to gain first insights in the relative distribution, identity and potential role of the major community members. RESULTS: Taxonomic analysis of 16S rRNA gene clone libraries (702 clones) revealed a microbiota dominated by Firmicutes (94.7%), followed by a minority of Actinobacteria (4.3%), Proteobacteria (0.4%) and Fusobacteria (0.6%). In the Firmicutes, the majority of the phylotypes within the Clostridiales were assigned to Clostridium clusters XIVa (43%), XI (38%) and I (13%). Members of the Bacteroidetes phylum and Bifidobacteriaceae, two groups that can positively contribute in maintaining intestinal homeostasis, were absent in the clone libraries and detected in only marginal to low levels in real-time PCR analyses. CONCLUSIONS: This marked underrepresentation is in contrast to data previously reported in domestic cats where Bacteroidetes and Bifidobacteriaceae are common residents of the faecal microbiota. Next to methodological differences, these findings may also reflect the apparent differences in dietary habits of both felid species. Thus, our results question the role of the domestic cat as the best available model for nutritional intervention studies in endangered exotic felids.

Psychrotrophic members of Leuconostoc gasicomitatum, Leuconostoc gelidum and Lactococcus piscium dominate at the end of shelf-life in packaged and chilled-stored food products in Belgium.

Previously, a considerable underestimation (+0.5-3.2 log CFU/g) on the contamination levels of psychrotrophic lactic acid bacteria (LAB) was observed for 33 retail, packaged food products stored at chilling temperature when the mesophilic enumeration technique was implemented as reference shelf-life parameter. In the present study, the microbial diversity of the dominant psychrotrophic LAB recovered after incubation of plates at 22 °C for 5 days was determined using a polyphasic taxonomic approach. A total of 212 LAB isolates were identified using a combination of rep-PCR fingerprinting, amplified fragment length polymorphism (AFLP) analysis and pheS gene sequencing. Leuconostoc gasicomitatum, Leuconostoc gelidum, Leuconostoc spp., Lactococcus piscium and Lactobacillus algidus proved to be the most competent and predominant species that may go undetected by the widely applied mesophilic enumeration protocols (ISO 4833:2003 and ISO 15214:1998). This study has assessed the interspecific variation among potential spoilage LAB, and highlights the significance of implementing a reference shelf-life parameter based on the enumeration of the total psychrotrophic bacterial load for industrial microbiological routine analyses.

Acinetobacter kookii sp. nov., isolated from soil.

Two Gram-stain-negative, non-fermentative bacterial strains, designated 11-0202(T) and 11-0607, were isolated from soil in South Korea, and four others, LUH 13522, LUH 8638, LUH 10268 and LUH 10288, were isolated from a beet field in Germany, soil in the Netherlands, and sediment of integrated fish farms in Malaysia and Thailand, respectively. Based on 16S rRNA, rpoB and gyrB gene sequences, they are considered to represent a novel species of the genus Acinetobacter. Their 16S rRNA gene sequences showed greatest pairwise similarity to Acinetobacter beijerinckii NIPH 838(T) (97.9-98.4 %). They shared highest rpoB and gyrB gene sequence similarity with Acinetobacter johnsonii DSM 6963(T) and Acinetobacter bouvetii 4B02(T) (85.4-87.6 and 78.1-82.7 %, respectively). Strain 11-0202(T) displayed low DNA-DNA reassociation values (<40 %) with the most closely related species of the genus Acinetobacter. The six strains utilized azelate, 2,3-butanediol, ethanol and dl-lactate as sole carbon sources. Cellular fatty acid analyses showed similarities to profiles of related species of the genus Acinetobacter: summed feature 3 (C16 : 1ω7c, C16 : 1ω6c; 24.3-27.2 %), C18 : 1ω9c (19.9-22.1 %), C16 : 0 (15.2-22.0 %) and C12 : 0 (9.2-14.2 %). On the basis of the current findings, it is concluded that the six strains represent a novel species, for which the name Acinetobacter kookii sp. nov. is proposed. The type strain is 11-0202(T) ( = KCTC 32033(T) = JCM 18512(T)).

Application of culture-dependent and culture-independent methods for the identification of Lactobacillus kefiranofaciens in microbial consortia present in kefir grains.

The biological and technological characteristics of kefiran as well as its importance in grain integrity led us to analyze the microbial kefir grain consortium with focus on Lactobacillus kefiranofaciens. The presence of L. kefiranofaciens in the nine kefir grains studied was demonstrated by denaturing gradient gel electrophoresis. By culture dependent methods applying a methodology focused on the search of this species, 22 isolates with typical morphology were obtained and identified applying a combination of SDS-PAGE of whole cell proteins, (GTG)5-PCR and sequence analysis of the housekeeping gene encoding the α-subunit of bacterial phenylalanyl-tRNA synthase (pheS). This polyphasic approach allowed the reliable identification of 11 L. kefiranofaciens, 5 Lactobacillus paracasei, 4 Lactobacillus kefiri and 2 Lactobacillus parakefiri isolates. Isolated L. kefiranofaciens strains produced polysaccharide in strain-dependent concentrations and EPS produced by them also differed in the degree of polymerization. The isolation and accurate identification of L. kefiranofaciens is relevant taking into account the important role of this microorganism in the grain ecosystem as well as its potential application as starter in food fermentations.

Gut microbiome studies in CKD: opportunities, pitfalls and therapeutic potential.

Interest in gut microbiome dysbiosis and its potential association with the development and progression of chronic kidney disease (CKD) has increased substantially in the past 6 years. In parallel, the microbiome field has matured considerably as the importance of host-related and environmental factors is increasingly recognized. Past research output in the context of CKD insufficiently considered the myriad confounding factors that are characteristic of the disease. Gut microbiota-derived metabolites remain an interesting therapeutic target to decrease uraemic (cardio)toxicity. However, future studies on the effect of dietary and biotic interventions will require harmonization of relevant readouts to enable an in-depth understanding of the underlying beneficial mechanisms. High-quality standards throughout the entire microbiome analysis workflow are also of utmost importance to obtain reliable and reproducible results. Importantly, investigating the relative composition and abundance of gut bacteria, and their potential association with plasma uraemic toxins levels is not sufficient. As in other fields, the time has come to move towards in-depth quantitative and functional exploration of the patient's gut microbiome by relying on confounder-controlled quantitative microbial profiling, shotgun metagenomics and in vitro simulations of microorganism-microorganism and host-microorganism interactions. This step is crucial to enable the rational selection and monitoring of dietary and biotic intervention strategies that can be deployed as a personalized intervention in CKD.

Dysbiosis of the faecal microbiota in patients with Crohn's disease and their unaffected relatives.

BACKGROUND AND AIMS: A general dysbiosis of the intestinal microbiota has been established in patients with Crohn's disease (CD), but a systematic characterisation of this dysbiosis is lacking. Therefore the composition of the predominant faecal microbiota of patients with CD was studied in comparison with the predominant composition in unaffected controls. Whether dysbiosis is present in relatives of patients CD was also examined. METHODS: Focusing on families with at least three members affected with CD, faecal samples of 68 patients with CD, 84 of their unaffected relatives and 55 matched controls were subjected to community fingerprinting of the predominant microbiota using denaturing gradient gel electrophoresis (DGGE). To analyse the DGGE profiles, BioNumerics software and non-parametric statistical analyses (SPSS V.17.0) were used. Observed differences in the predominant microbiota were subsequently confirmed and quantified with real-time PCR. RESULTS: Five bacterial species characterised dysbiosis in CD, namely a decrease in Dialister invisus (p=0.04), an uncharacterised species of Clostridium cluster XIVa (p = 0.03), Faecalibacterium prausnitzii (p < 1.3 × 10⁻5) and Bifidobacterium adolescentis (p = 5.4 × 10⁻6), and an increase in Ruminococcus gnavus (p = 2.1 × 10⁻7). Unaffected relatives of patients with CD had less Collinsella aerofaciens (p = 0.004) and a member of the Escherichia coli-Shigella group (p = 0.01) and more Ruminococcus torques (p = 0.02) in their predominant microbiota as compared with healthy subjects. CONCLUSION: Unaffected relatives of patients with CD have a different composition of their microbiota compared with healthy controls. This dysbiosis is not characterised by lack of butyrate producing-bacteria as observed in CD but suggests a role for microorganisms with mucin degradation capacity.

Effect of cryopreservation medium conditions on growth and isolation of gut anaerobes from human faecal samples.

BACKGROUND: Novel strategies for anaerobic bacterial isolations from human faecal samples and various initiatives to generate culture collections of gut-derived bacteria have instigated considerable interest for the development of novel microbiota-based treatments. Early in the process of building a culture collection, optimal faecal sample preservation is essential to safeguard the viability of the broadest taxonomic diversity range possible. In contrast to the much more established faecal storage conditions for meta-omics applications, the impact of stool sample preservation conditions on bacterial growth recovery and isolation remains largely unexplored. In this study, aliquoted faecal samples from eleven healthy human volunteers selected based on a range of physicochemical and microbiological gradients were cryopreserved at - 80 °C either without the addition of any medium (dry condition) or in different Cary-Blair medium conditions with or without a cryoprotectant, i.e. 20% (v/v) glycerol or 5% (v/v) DMSO. Faecal aliquots were subjected to bulk 16S rRNA gene sequencing as well as dilution plating on modified Gifu Anaerobic Medium after preservation for culturable fraction profiling and generation of bacterial culture collections. RESULTS: Analyses of compositional variation showed that cryopreservation medium conditions affected quantitative recovery but not the overall community composition of cultured fractions. Post-preservation sample dilution and richness of the uncultured source samples were the major drivers of the cultured fraction richness at genus level. However, preservation conditions differentially affected recovery of specific genera. Presence-absence analysis indicated that twenty-two of the 45 most abundant common genera (>0.01% abundance, dilution 10-4) were recovered in cultured fractions from all preservation conditions, while nine genera were only detected in fractions from a single preservation condition. Overall, the highest number of common genera (i.e. 35/45) in cultured fractions were recovered from sample aliquots preserved without medium and in the presence of Cary-Blair medium containing 5% (v/v) DMSO. Also, in the culture collection generated from the cultured fractions, these two preservation conditions yielded the highest species richness (72 and 66, respectively). CONCLUSION: Our results demonstrate that preservation methods partly determine richness and taxonomic diversity of gut anaerobes recovered from faecal samples. Complementing the current standard practice of cryopreserving stool samples in dry conditions with other preservation conditions, such as Cary-Blair medium with DMSO, could increase the species diversity of gut-associated culture collections. Video abstract.

Fast quantification of gut bacterial species in cocultures using flow cytometry and supervised classification.

A bottleneck for microbial community experiments with many samples and/or replicates is the fast quantification of individual taxon abundances, which is commonly achieved through sequencing marker genes such as the 16S rRNA gene. Here, we propose a new approach for high-throughput and high-quality enumeration of human gut bacteria in a defined community, combining flow cytometry and supervised classification to identify and quantify species mixed in silico and in defined communities in vitro. We identified species in a 5-species in silico community with an F1 score of 71%. In addition, we demonstrate in vitro that our method performs equally well or better than 16S rRNA gene sequencing in two-species cocultures and agrees with 16S rRNA gene sequencing data on the most abundant species in a four-species community. We found that shape and size differences alone are insufficient to distinguish species, and that it is thus necessary to exploit the multivariate nature of flow cytometry data. Finally, we observed that variability of flow cytometry data across replicates differs between gut bacterial species. In conclusion, the performance of supervised classification of gut species in flow cytometry data is species-dependent, but is for some combinations accurate enough to serve as a faster alternative to 16S rRNA gene sequencing.

Cross-sectional and longitudinal comparisons of the predominant fecal microbiota compositions of a group of pediatric patients with cystic fibrosis and their healthy siblings.

Although only poorly documented, it can be assumed that intensive antibiotic treatments of chronic lung infections in patients with cystic fibrosis (CF) also affect the diversity and metabolic functioning of the gastrointestinal microbiota and potentially lead to a state of dysbiosis. A better knowledge of the differences in gut microbiota composition and stability between patients with CF and healthy subjects could lead to optimization of current antibiotic therapies and/or development of add-on therapies. Using conventional culturing and population fingerprinting by denaturing gradient gel electrophoresis (DGGE) of 16S rRNA amplicons, we compared the predominant fecal microbiota of 21 patients with CF and 24 healthy siblings in a cross-sectional study. General medium counts, as well as counts on media specific for lactic acid bacteria, clostridia, Bifidobacterium spp., Veillonella spp., and Bacteroides-Prevotella spp., were consistently higher in sibling samples than in CF samples, whereas the reverse was found for enterobacterial counts. DGGE fingerprinting uncovered large intersubject variations in both study groups. On the other hand, the cross-sectional data indicated that the predominant fecal microbiota of patients and siblings had comparable species richness. In addition, a longitudinal study was performed on 7 or 8 consecutive samples collected over a 2-year period from two patients and their respective siblings. For these samples, DGGE profiling indicated an overall trend toward lower temporal stability and lower species richness in the predominant fecal CF microbiota. The observed compositional and dynamic perturbations provide the first evidence of a general dysbiosis in children with CF compared to their siblings.

Metatranscriptome analysis for insight into whole-ecosystem gene expression during spontaneous wheat and spelt sourdough fermentations.

Lactic acid bacteria (LAB) are of industrial importance in the production of fermented foods, including sourdough-derived products. Despite their limited metabolic capacity, LAB contribute considerably to important characteristics of fermented foods, such as extended shelf-life, microbial safety, improved texture, and enhanced organoleptic properties. Triggered by the considerable amount of LAB genomic information that became available during the last decade, transcriptome and, by extension, metatranscriptome studies have become one of the most appropriate research approaches to study whole-ecosystem gene expression in more detail. In this study, microarray analyses were performed using RNA sampled during four 10-day spontaneous sourdough fermentations carried out in the laboratory with an in-house-developed LAB functional gene microarray. For data analysis, a new algorithm was developed to calculate a net expression profile for each of the represented genes, allowing use of the microarray analysis beyond the species level. In addition, metabolite target analyses were performed on the sourdough samples to relate gene expression with metabolite production. The results revealed the activation of different key metabolic pathways, the ability to use carbohydrates other than glucose (e.g., starch and maltose), and the conversion of amino acids as a contribution to redox equilibrium and flavor compound generation in LAB during sourdough fermentation.

Gut Microbiome Profiling Uncovers a Lower Abundance of Butyricicoccus in Advanced Stages of Chronic Kidney Disease.

Chronic kidney disease (CKD) is characterized by the accumulation of uremic toxins which exert deleterious effects on various organ systems. Several of these uremic toxins originate from the bacterial metabolization of aromatic amino acids in the colon. This study assessed whether the gut microbial composition varies among patients in different stages of CKD. Uremic metabolites were quantified by UPLC/fluorescence detection and microbial profiling by 16S rRNA amplicon sequencing. Gut microbial profiles of CKD patients were compared among stages 1-2, stage 3 and stages 4-5. Although a substantial inter-individual difference in abundance of the top 15 genera was observed, no significant difference was observed between groups. Bristol stool scale (BSS) correlated negatively with p-cresyl sulfate and hippuric acid levels, irrespective of the intake of laxatives. Butyricicoccus, a genus with butyrate-generating properties, was decreased in abundance in advanced stages of CKD compared to the earlier stages (p = 0.043). In conclusion, in this cross-sectional study no gradual differences in the gut microbial profile over the different stages of CKD were observed. However, the decrease in the abundance of Butyricicoccus genus with loss of kidney function stresses the need for more in-depth functional exploration of the gut microbiome in CKD patients not on dialysis.