Influence of 2′-fucosyllactose on the microbiota composition and metabolic activity of fecal cultures from breastfed and formula-fed infants at two months of age

Although breast milk is considered the gold standard of nutrition for infant feeding, some circumstances may make breastfeeding difficult. Several commercial milk preparations include syn-thetic human milk oligosaccharides (HMOs) in their composition. However, the effect of HMOs on the establishment of the intestinal microbiota remains incompletely understood. Independent batch fermentations were performed with feces from six full-term infant donors of two months of age (three breastfed and three formula-fed, exclusively) in the presence of 2′fucosyllactose (2′FL), one of the most abundant HMOs in human milk. Microbiota composition was analyzed by 16S rRNA gene sequencing at baseline and at 24 h of incubation. The 2′FL consumption, gas accumulation, and levels of different metabolites were determined by chromatography. Microbiota profiles at baseline were clearly influenced by the mode of feeding and by the intrinsic ability of microbiotas to degrade 2′FL. The 2′FL degradation rate clustered fecal cultures into slow and fast degraders, regardless of feeding type, this being a determinant factor influencing the evolution of the microbiota during incubation, although the low number of donors precludes drawing sound conclusions. More studies are needed to decipher the extent to which the early intervention with HMOs could influence the microbiota as a function of its ability to utilize 2′FL.This research was funded by a contract of Lallemand Health Solutions Inc. with IPLA-CSIC and by the Intramural CSIC Research project PIE201970E061. S.A. was the recipient of a postdoctoral Juan de la Cierva Contract (Ministry of Science, Innovation and Universities, Ref. IJCI-2017-32156) and NS has a postdoctoral contract awarded by the Biosanitary Research Foundation in Asturias (FINBA, Spain)

In Vitro Probiotic Modulation of the Intestinal Microbiota and 2′Fucosyllactose Consumption in Fecal Cultures from Infants at Two Months of Age

2-fucosyllactose (2 FL) is one of the most abundant oligosaccharides in human milk, with benefits on neonatal health. Previous results point to the inability of the fecal microbiota from some infants to ferment 2 FL. We evaluated a probiotic formulation, including the strains Lactobacillus helveticus Rosell®-52 (R0052), Bifidobacterium longum subsp. infantis Rosell®-33 (R0033), and Bifidobacterium bifidum Rosell®-71 (R0071), individually or in an 80:10:10 combination on the microbiota and 2 FL degradation. Independent batch fermentations were performed with feces from six full-term infant donors of two months of age (three breastfed and three formula-fed) with added probiotic formulation or the constituent strains in the presence of 2 FL. Microbiota composition was analyzed by 16S rRNA gene sequencing. Gas accumulation, pH decrease and 2 FL consumption, and levels of different metabolites were determined by chromatography. B. bifidum R0071 was the sole microorganism promoting a partial increase of 2 FL degradation during fermentation in fecal cultures of 2 FL slow-degrading donors. However, major changes in microbiota composition and metabolic activity occurred with L. helveticus R0052 or the probiotic formulation in cultures of slow degraders. Further studies are needed to decipher the role of the host intestinal microbiota in the efficacy of these strains.This research was funded by a contract of Lallemand Health Solutions Inc. with IPLA-CSIC (Spanish National Research Council contract number 201266) and by the Intramural Spanish National Research Council project PIE201970E061. S.A. was the recipient of a postdoctoral Juan de la Cierva contract (Ministry of Science and Innovation, Ref. IJCI-2017-32156), and N.S. has a postdoctoral contract awarded by the Biosanitary Research Foundation in Asturias (FINBA, Spain)

The effects of antimicrobials and lipopolysaccharide on acute immune responsivity in pubertal male and female CD1 mice

Exposure to stress during critical periods of development—such as puberty—is associated with long-term disruptions in brain function and neuro-immune responsivity. However, the mechanisms underlying the effect of stress on the pubertal neuro-immune response has yet to be elucidated. Therefore, the objective of the current study was to investigate the effect antimicrobial and lipopolysaccharide (LPS) treatments on acute immune responsivity in pubertal male and female mice. Moreover, the potential for probiotic supplementation to mitigate these effects was also examined. 240 male and female CD1 mice were treated with one week of antimicrobial treatment (mixed antimicrobials or water) and probiotic treatment (L. rhamnosis R0011 and L. helveticus R0052 or L. helveticus R0052 and B. longum R0175) or placebo at five weeks of age. At six weeks of age (pubertal stress-sensitive period), the mice received a single injection of LPS or saline. Sickness behaviours were assessed, and mice were euthanized eight hours post-injection. Brain, blood, and intestinal samples were collected. The results indicated that the antimicrobial treatment reduced sickness behaviours, and potentiated LPS-induced plasma cytokine concentrations and pro-inflammatory markers in the pre-frontal cortex (PFC) and hippocampus, in a sex-dependent manner. However, probiotics reduced LPS-induced plasma cytokine concentrations along with hippocampal and PFC pro-inflammatory markers in a sex-dependent manner. L. rhamnosis R0011 and L. helveticus R0052 treatment also mitigated antimicrobial-induced plasma cytokine concentrations and sickness behaviours. These findings suggest that the microbiome is an important modulator of the pro-inflammatory immune response during puberty

Modulación in vitro de la microbiota intestinal de niños lactantes y del consumo de 2’-fucosillactosa con cepas de bacterias lácticas probióticas

Resumen del trabajo presentado a la 15ª Reunión de la Red Española de Bacterias Lácticas: Bacterias Lácticas en Alimentación y Salud, celebrada en Valencia (España), los días 26 y 27 de mayo de 2022.El establecimiento de la microbiota neonatal en las primeras etapas de la vida está modulado por varios tipos de factores entre los que destaca especialmente el tipo de alimentación (leche maternaLM o leche de fórmula-LF). La 2´-fucosillactosa (2´FL) es uno de los oligosacáridos más abundantes de la leche materna (“human milk oligosaccharides”-HMO), con actividad prebiótica. Los niños que no son amamantados con LM, lo ingieren en LF suplementadas, en combinación con cepas probióticas en muchas ocasiones. Sin embargo, el efecto tanto de este HMO como de la combinación con cepas probióticas a nivel de la microbiota intestinal y sus metabolitos no ha sido estudiado en detalle. Por tanto, es de gran interés la evaluación del efecto de la 2’FL y cepas probióticas destinadas a la alimentación de lactantes sobre la microbiota intestinal infantil (MI)

Roles of long non-coding RNAs in skeletal muscle development and disease

Skeletal muscle plays a central role in whole body metabolism and serves as the major site for insulin-stimulated glucose disposal. Insulin resistance in skeletal muscle is considered an initiating factor in the development of Type 2 diabetes (T2D). Skeletal muscle is also known for its ability to regenerate after injury in the process called myogenesis. Aberrant myogenesis will lead to many muscle disorders including various types of muscular dystrophies. The comprehensive delineation of genes involved in muscle regeneration and metabolism is an essential step towards understanding muscle biology and pathophysiology. While many studies have identified and dissected the role of protein-coding genes in muscle differentiation and function, recent discoveries demonstrate that the human genome also contains thousands of non-protein-coding genes termed long non-coding RNAs (lncRNAs). However, the identity and functions of many of these new genes are still unknown. Here, we describe the systematic discovery and characterization of lncRNAs in skeletal muscle. We focused on two main aspects of muscle biology which are muscle regeneration and metabolism.We used total RNA transcriptome sequencing in a panel of primary human myoblasts and myotubes to identify 191 known and novel lncRNAs with differential expression during muscle differentiation in vitro. We focused on one lncRNA, RMD1, whose expression is increased during myoblast differentiation in both mouse and humans. RMD1 expression also increases in the acute and chronic in vivo models of myogenesis; and RMD1 knockdown in vitro leads to decreased expression of myogenesis markers, which highlights its potential role in regulating skeletal muscle regeneration.We also present for the first time a comprehensive profile of known and novel lncRNAs whose expression changes in skeletal muscle obtained from diabetic subjects. These include a lncRNA we named TDNC1 (T2D down-regulated non-coding RNA 1) that is located near a previously identified T2D-associated variant (rs2943641). TDNC1 expression is correlated with rs2943641 genotype and is reduced in T2D as well as in young normoglycemic individuals with a family history of T2D. Compounds known to alter insulin sensitivity regulate TDNC1 expression. Moreover, ectopic expression of this lncRNA results in specific up-regulation of genes involved in insulin signaling as well as increased activity of this pathway.Taken together, this thesis presents a comprehensive assessment of lncRNAs as novel regulators in muscle differentiation and metabolism. Broadly, this work suggests that uncharacterized lncRNAs may play important roles not just in muscle biology and disease, but also in other biological processes and pathogenesis of various diseases.Le muscle squelettique contribue de manière significative à plusieurs fonctions corporelles. Il joue un rôle central dans le métabolisme du corps entier et il est le site principal pour l'élimination du glucose stimulée par l'insuline. La résistance à l'insuline dans le muscle squelettique est considérée comme un facteur déterminant qui mène à l'apparition du diabète de type 2 (DT2). Le muscle squelettique est également connu pour sa capacité à se régénérer après une blessure suivant un processus appelé myogenèse. La myogenèse aberrante peut entraîner de nombreux troubles musculaires, y compris différents types de dystrophies. L'identification et la caractérisation complète des gènes impliqués dans la régénération et le métabolisme du tissu musculaire sont des étapes essentielles vers la compréhension de la biologie musculaire et des pathologies qui lui sont associées. Bien que de nombreuses recherches aient identifié et disséqué le rôle des gènes codant pour les protéines, les découvertes récentes démontrent que le génome humain contient des milliers de gènes non codant appelés des longs ARN non codants (lncRNAs). Cependant, l'identité et la fonction de la majorité de ces nouveaux gènes restent à l'heure actuelle inconnues. Nous décrivons içi la découverte systématique et la caractérisation des lncRNAs dans le muscle squelettique. Nous nous sommes concentrés sur deux aspects principaux de la biologie musculaire qui sont la régénération musculaire et le métabolisme.Nous avons utilisé des données de séquençage de l'ARN total d'un large panel de myoblastes primaires et de myotubes pour identifier 191 lncRNAs, incluant des lnRNAs déjà connus, mais aussi d'autres nouveaux avec une expression différentielle lors de la différenciation in vitro. Nous nous sommes concentrés sur RMD1, un lncRNA, dont l'expression est augmentée lors de la différenciation de myoblaste à la fois chez la souris et l'humain. L'expression de ce gène augmente également dans les modèles in vivo de myogenèse aigüe et chronique. L'inactivation du gène RMD1 (knockdown) conduit à une diminution de l'expression des marqueurs de myogenèse ce qui souligne son rôle potentiel dans la régénération des muscles squelettiques.Nous présentons également pour la première fois un profil complet de lncRNAs, à la fois connus et nouveaux dont l'expression change dans le muscle squelettique de sujet diabétique de type 2. Ceux-ci incluent un lncRNA spécifique aux muscles, que nous avons appelé TDNC1 (T2D down-regulated non-coding RNA 1), situé à proximité d'une variante associée au DT2 précédemment identifiée (rs2943641). L'expression de TDNC1 est corrélée au génotype de rs2943641 et nous avons démontré qu'elle est réduite dans les cas de DT2 ainsi que chez les jeunes normoglycémiques avec des antécédents familiaux de DT2. Les composés connus pour modifier la sensibilité à l'insuline peuvent réguler l'expression de TDNC1. De plus, l'expression ectopique de ce lncRNA entraîne une régulation spécifique des gènes impliqués dans la signalisation de l'insuline ainsi qu'une augmentation de l'activité de cette voie.Dans l'ensemble, nos données représentent une évaluation complète des lncRNAs comme nouveaux régulateurs de la différenciation musculaire et du métabolisme. D'une manière générale, ce travail suggère que les lncRNAs non caractérisés peuvent jouer un rôle important non seulement dans la biologie musculaire et les maladies associées, mais aussi dans d'autres processus biologiques et pathologiques impliquant les muscles de diverses maladies

Complete Genome Sequence of Phascolarctobacterium faecium G 104, Isolated from the Stools of a Healthy Lean Donor

Phascolarctobacterium faecium is a strict anaerobe belonging to the Firmicutes phylum that is found abundantly in the human gastrointestinal tract. Here, we report the complete genome sequence of P. faecium G 104, a strain isolated from a fresh stool sample from a healthy lean donor.This work and the contract of E.M.G.D.P. were supported by the European Commission 7th Framework Program through the MyNewGut project (grant 613979). The contract of A.G. was supported by the European Commission H2020 Program through the EarlyCause project (grant 848158).Peer reviewe

Association of Levels of Fasting Glucose and Insulin With Rare Variants at the Chromosome 11p11.2- MADD

BACKGROUND: Common variation at the 11p11.2 locus, encompassing MADD, ACP2, NR1H3, MYBPC3 and SPI1, has been associated in genome-wide association studies with fasting glucose (FG) and insulin (FI). In the Cohorts for Heart and Aging Research in Genomic Epidemiology Targeted Sequencing Study, we sequenced five gene regions at 11p11.2 to identify rare, potentially functional variants influencing FG or FI levels. METHOD & RESULTS: Sequencing (mean depth 38×) across 16.1kb in 3,566 non-diabetic individuals identified 653 variants, 79.9% of which were rare (MAF <1%) and novel. We analyzed rare variants in five gene regions with FI or FG using the Sequence Kernel Association Test (SKAT). At NR1H3, 53 rare variants were jointly associated with FI (p=2.73 × 10(−3)); of these, seven were predicted to have regulatory function and showed association with FI (p=1.28 × 10(−3)). Conditioning on two previously associated variants at MADD (rs7944584, rs10838687) did not attenuate this association, suggesting that there are more than two independent signals at 11p11.2. One predicted regulatory variant, chr11:47227430 (hg18; MAF 0.00068), contributed 20.6% to the overall SKAT score at NR1H3, lies in intron 2 of NR1H3 and is a predicted binding site for FOXA1, a transcription factor associated with insulin regulation. In human HepG2 hepatoma cells, the rare chr11:47227430 A allele disrupted FOXA1 binding and reduced FOXA1-dependent transcriptional activity. CONCLUSION: Sequencing at 11p11.2- NR1H3 identified rare variation associated with FI. One variant, chr11:47227430, appears to be functional, with the rare A allele reducing transcription factor FOXA1 binding and FOXA1-dependent transcriptional activity

Association of Levels of Fasting Glucose and Insulin With Rare Variants at the Chromosome 11p11.2-MADD Locus

BackgroundCommon variation at the 11p11.2 locus, encompassing MADD, ACP2, NR1H3, MYBPC3, and SPI1, has been associated in genome-wide association studies with fasting glucose and insulin (FI). In the Cohorts for Heart and Aging Research in Genomic Epidemiology Targeted Sequencing Study, we sequenced 5 gene regions at 11p11.2 to identify rare, potentially functional variants influencing fasting glucose or FI levels.Methods and resultsSequencing (mean depth, 38×) across 16.1 kb in 3566 individuals without diabetes mellitus identified 653 variants, 79.9% of which were rare (minor allele frequency &lt;1%) and novel. We analyzed rare variants in 5 gene regions with FI or fasting glucose using the sequence kernel association test. At NR1H3, 53 rare variants were jointly associated with FI (P=2.73×10(-3)); of these, 7 were predicted to have regulatory function and showed association with FI (P=1.28×10(-3)). Conditioning on 2 previously associated variants at MADD (rs7944584, rs10838687) did not attenuate this association, suggesting that there are &gt;2 independent signals at 11p11.2. One predicted regulatory variant, chr11:47227430 (hg18; minor allele frequency=0.00068), contributed 20.6% to the overall sequence kernel association test score at NR1H3, lies in intron 2 of NR1H3, and is a predicted binding site for forkhead box A1 (FOXA1), a transcription factor associated with insulin regulation. In human HepG2 hepatoma cells, the rare chr11:47227430 A allele disrupted FOXA1 binding and reduced FOXA1-dependent transcriptional activity.ConclusionsSequencing at 11p11.2-NR1H3 identified rare variation associated with FI. One variant, chr11:47227430, seems to be functional, with the rare A allele reducing transcription factor FOXA1 binding and FOXA1-dependent transcriptional activity