Prevotella copri and microbiota members mediate the beneficial effects of a therapeutic food for malnutrition.

Microbiota-directed complementary food (MDCF) formulations have been designed to repair the gut communities of malnourished children. A randomized controlled trial demonstrated that one formulation, MDCF-2, improved weight gain in malnourished Bangladeshi children compared to a more calorically dense standard nutritional intervention. Metagenome-assembled genomes from study participants revealed a correlation between ponderal growth and expression of MDCF-2 glycan utilization pathways by Prevotella copri strains. To test this correlation, here we use gnotobiotic mice colonized with defined consortia of age- and ponderal growth-associated gut bacterial strains, with or without P. copri isolates closely matching the metagenome-assembled genomes. Combining gut metagenomics and metatranscriptomics with host single-nucleus RNA sequencing and gut metabolomic analyses, we identify a key role of P. copri in metabolizing MDCF-2 glycans and uncover its interactions with other microbes including Bifidobacterium infantis. P. copri-containing consortia mediated weight gain and modulated energy metabolism within intestinal epithelial cells. Our results reveal structure-function relationships between MDCF-2 and members of the gut microbiota of malnourished children with potential implications for future therapies.

Bioactive glycans in a microbiome-directed food for children with malnutrition.

Evidence is accumulating that perturbed postnatal development of the gut microbiome contributes to childhood malnutrition1-4. Here we analyse biospecimens from a randomized, controlled trial of a microbiome-directed complementary food (MDCF-2) that produced superior rates of weight gain compared with a calorically more dense conventional ready-to-use supplementary food in 12-18-month-old Bangladeshi children with moderate acute malnutrition4. We reconstructed 1,000 bacterial genomes (metagenome-assembled genomes (MAGs)) from the faecal microbiomes of trial participants, identified 75 MAGs of which the abundances were positively associated with ponderal growth (change in weight-for-length Z score (WLZ)), characterized changes in MAG gene expression as a function of treatment type and WLZ response, and quantified carbohydrate structures in MDCF-2 and faeces. The results reveal that two Prevotella copri MAGs that are positively associated with WLZ are the principal contributors to MDCF-2-induced expression of metabolic pathways involved in utilizing the component glycans of MDCF-2. The predicted specificities of carbohydrate-active enzymes expressed by their polysaccharide-utilization loci are correlated with (1) the in vitro growth of Bangladeshi P. copri strains, possessing varying degrees of polysaccharide-utilization loci and genomic conservation with these MAGs, in defined medium containing different purified glycans representative of those in MDCF-2, and (2) the levels of faecal carbohydrate structures in the trial participants. These associations suggest that identifying bioactive glycan structures in MDCFs metabolized by growth-associated bacterial taxa will help to guide recommendations about their use in children with acute malnutrition and enable the development of additional formulations.

Prevotella copri-related effects of a therapeutic food for malnutrition.

Preclinical and clinical studies are providing evidence that the healthy growth of infants and children reflects, in part, healthy development of their gut microbiomes1-5. This process of microbial community assembly and functional maturation is perturbed in children with acute malnutrition. Gnotobiotic animals, colonized with microbial communities from children with severe and moderate acute malnutrition, have been used to develop microbiome-directed complementary food (MDCF) formulations for repairing the microbiomes of these children during the weaning period5. Bangladeshi children with moderate acute malnutrition (MAM) participating in a previously reported 3-month-long randomized controlled clinical study of one such formulation, MDCF-2, exhibited significantly improved weight gain compared to a commonly used nutritional intervention despite the lower caloric density of the MDCF6. Characterizing the 'metagenome assembled genomes' (MAGs) of bacterial strains present in the microbiomes of study participants revealed a significant correlation between accelerated ponderal growth and the expression by two Prevotella copri MAGs of metabolic pathways involved in processing of MDCF-2 glycans1. To provide a direct test of these relationships, we have now performed 'reverse translation' experiments using a gnotobiotic mouse model of mother-to-offspring microbiome transmission. Mice were colonized with defined consortia of age- and ponderal growth-associated gut bacterial strains cultured from Bangladeshi infants/children in the study population, with or without P. copri isolates resembling the MAGs. By combining analyses of microbial community assembly, gene expression and processing of glycan constituents of MDCF-2 with single nucleus RNA-Seq and mass spectrometric analyses of the intestine, we establish a principal role for P. copri in mediating metabolism of MDCF-2 glycans, characterize its interactions with other consortium members including Bifidobacterium longum subsp. infantis, and demonstrate the effects of P. copri-containing consortia in mediating weight gain and modulating the activities of metabolic pathways involved in lipid, amino acid, carbohydrate plus other facets of energy metabolism within epithelial cells positioned at different locations in intestinal crypts and villi. Together, the results provide insights into structure/function relationships between MDCF-2 and members of the gut communities of malnourished children; they also have implications for developing future prebiotic, probiotic and/or synbiotic therapeutics for microbiome restoration in children with already manifest malnutrition, or who are at risk for this pervasive health challenge.

Bioactive glycans in a microbiome-directed food for malnourished children.

Evidence is accumulating that perturbed postnatal development of the gut microbiome contributes to childhood malnutrition1-4. Designing effective microbiome-directed therapeutic foods to repair these perturbations requires knowledge about how food components interact with the microbiome to alter its expressed functions. Here we use biospecimens from a randomized, controlled trial of a microbiome-directed complementary food prototype (MDCF-2) that produced superior rates of weight gain compared to a conventional ready-to-use supplementary food (RUSF) in 12-18-month-old Bangladeshi children with moderate acute malnutrition (MAM)4. We reconstructed 1000 bacterial genomes (metagenome-assembled genomes, MAGs) present in their fecal microbiomes, identified 75 whose abundances were positively associated with weight gain (change in weight-for-length Z score, WLZ), characterized gene expression changes in these MAGs as a function of treatment type and WLZ response, and used mass spectrometry to quantify carbohydrate structures in MDCF-2 and feces. The results reveal treatment-induced changes in expression of carbohydrate metabolic pathways in WLZ-associated MAGs. Comparing participants consuming MDCF-2 versus RUSF, and MDCF-2-treated children in the upper versus lower quartiles of WLZ responses revealed that two Prevotella copri MAGs positively associated with WLZ were principal contributors to MDCF-2-induced expression of metabolic pathways involved in utilization of its component glycans. Moreover, the predicted specificities of carbohydrate active enzymes expressed by polysaccharide utilization loci (PULs) in these two MAGs correlate with the (i) in vitro growth of Bangladeshi P. copri strains, possessing differing degrees of PUL and overall genomic content similarity to these MAGs, cultured in defined medium containing different purified glycans representative of those in MDCF-2, and (ii) levels of carbohydrate structures identified in feces from clinical trial participants. In the accompanying paper5, we use a gnotobiotic mouse model colonized with age- and WLZ-associated bacterial taxa cultured from this study population, and fed diets resembling those consumed by study participants, to directly test the relationship between P. copri, MDCF-2 glycan metabolism, host ponderal growth responses, and intestinal gene expression and metabolism. The ability to identify bioactive glycan structures in MDCFs that are metabolized by growth-associated bacterial taxa will help guide recommendations about use of this MDCF for children with acute malnutrition representing different geographic locales and ages, as well as enable development of bioequivalent, or more efficacious, formulations composed of culturally acceptable and affordable ingredients.

Genotype and clinical characteristics of patients with Wolfram syndrome and WFS1-related disorders.

Objective: Wolfram syndrome (WFS) is an autosomal recessive disorder associated with juvenile-onset diabetes mellitus, optic atrophy, diabetes insipidus, and sensorineural hearing loss. We sought to elucidate the relationship between genotypic and phenotypic presentations of Wolfram syndrome which would assist clinicians in classifying the severity and prognosis of Wolfram syndrome more accurately. Approach: Patient data from the Washington University International Registry and Clinical Study for Wolfram Syndrome and patient case reports were analyzed to select for patients with two recessive mutations in the WFS1 gene. Mutations were classified as being either nonsense/frameshift variants or missense/in-frame insertion/deletion variants. Missense/in-frame variants were further classified as transmembrane or non-transmembrane based on whether they affected amino acid residues predicted to be in transmembrane domains of WFS1. Statistical analysis was performed using Wilcoxon rank-sum tests with multiple test adjustment applied via the Bonferonni correction. Results: A greater number of genotype variants correlated with earlier onset and a more severe presentation of Wolfram syndrome. Secondly, non-sense and frameshift variants had more severe phenotypic presentations than missense variants, as evidenced by diabetes mellitus and optic atrophy emerging significantly earlier in patients with two nonsense/frameshift variants compared with zero or one nonsense/frameshift variants. In addition, the number of transmembrane in-frame variants demonstrated a statistically significant dose-effect on age of onset of diabetes mellitus and optic atrophy among patients with either one or two in-frame variants. Summary/Conclusion: The results contribute to our current understanding of the genotype-phenotype relationship of Wolfram syndrome, suggesting that alterations in coding sequences result in significant changes in the presentation and severity of Wolfram. The impact of these findings is significant, as the results will aid clinicians in predicting more accurate prognoses and pave the way for personalized treatments for Wolfram syndrome.

Genotype and Clinical Characteristics of Patients with Wolfram Syndrome and WFS1-related Disorders.

Objective: Wolfram syndrome (WFS) is an autosomal recessive disorder associated with juvenile-onset diabetes mellitus, optic atrophy, diabetes insipidus, and sensorineural hearing loss. We sought to elucidate the relationship between genotypic and phenotypic presentations of Wolfram syndrome which would assist clinicians in classifying the severity and prognosis of Wolfram syndrome more accurately. Approach: Patient data from the Washington University International Registry and Clinical Study for Wolfram Syndrome and patient case reports were analyzed to select for patients with two recessive mutations in the WFS1 gene. Mutations were classified as being either nonsense/frameshift variants or missense/in-frame insertion/deletion variants and statistical analysis was performed using unpaired and paired t-tests and one- and two-way ANOVA with Tukey's or Dunnett's tests. Results: A greater number of genotype variants correlated with earlier onset and a more severe presentation of Wolfram syndrome. Secondly, non-sense and frameshift variants had more severe phenotypic presentations than missense variants, as evidenced by optic atrophy emerging significantly earlier in patients with 2 nonsense/frameshift alleles compared with 0 missense transmembrane variants. In addition, the number of transmembrane in-frame variants demonstrated a statistically significant dose-effect on age of onset of diabetes mellitus and optic atrophy. Summary / Conclusions: The results contribute to our current understanding of the genotype-phenotype relationship of Wolfram syndrome, suggesting that alterations in coding sequences result in significant changes in the presentation and severity of Wolfram. The impact of these findings is significant, as the results will aid clinicians in predicting more accurate prognoses and pave the way for personalized treatments for Wolfram syndrome.

Cytoprotection by a naturally occurring variant of ATP5G1 in Arctic ground squirrel neural progenitor cells.

Many organisms in nature have evolved mechanisms to tolerate severe hypoxia or ischemia, including the hibernation-capable Arctic ground squirrel (AGS). Although hypoxic or ischemia tolerance in AGS involves physiological adaptations, little is known about the critical cellular mechanisms underlying intrinsic AGS cell resilience to metabolic stress. Through cell survival-based cDNA expression screens in neural progenitor cells, we identify a genetic variant of AGS Atp5g1 that confers cell resilience to metabolic stress. Atp5g1 encodes a subunit of the mitochondrial ATP synthase. Ectopic expression in mouse cells and CRISPR/Cas9 base editing of endogenous AGS loci revealed causal roles of one AGS-specific amino acid substitution in mediating cytoprotection by AGS ATP5G1. AGS ATP5G1 promotes metabolic stress resilience by modulating mitochondrial morphological change and metabolic functions. Our results identify a naturally occurring variant of ATP5G1 from a mammalian hibernator that critically contributes to intrinsic cytoprotection against metabolic stress.

When animals hibernate, they lower their body temperature and metabolism to conserve the energy they need to withstand cold harsh winters. One such animal is the Arctic ground squirrel, an extreme hibernator that can drop its body temperatures to below 0°C. This hibernation ability means the cells of Arctic ground squirrels can survive severe shortages of blood and oxygen. But, it is unclear how their cells are able to endure this metabolic stress. To answer this question, Singhal, Bai et al. studied the cells of Arctic ground squirrels for unique features that might make them more durable to stress. Examining the genetic code of these resilient cells revealed that Arctic ground squirrels may have a variant form of a protein called ATP5G1. This protein is found in a cellular compartment called the mitochondria, which is responsible for supplying energy to the rest of the cell and therefore plays an important role in metabolic processes. Singhal, Bai et al. found that when this variant form of ATP5G1 was introduced into the cells of mice, their mitochondria was better at coping with stress conditions, such as low oxygen, low temperature and poisoning. Using a gene editing tool to selectively substitute some of the building blocks, also known as amino acids, that make up the ATP5G1 protein revealed that improvements to the mitochondria were caused by switching specific amino acids. However, swapping these amino acids, which presumably affects the role of ATP5G1, did not completely remove the cells' resilience to stress. This suggests that variants of other genes and proteins may also be involved in providing protection. These findings provide the first evidence of a protein variant that is responsible for protecting cells during the metabolic stress conditions caused by hibernation. The approach taken by Singhal, Bai et al. could be used to identify and study other proteins that increase resilience to metabolic stress. These findings could help develop new treatments for diseases caused by a limited blood supply to human organs, such as a stroke or heart attack.

Resilience to Injury: A New Approach to Neuroprotection?

Despite thousands of neuroprotectants demonstrating promise in preclinical trials, a neuroprotective therapeutic has yet to be approved for the treatment of acute brain injuries such as stroke or traumatic brain injury. Developing a more detailed understanding of models and populations demonstrating "neurological resilience" in spite of brain injury can give us important insights into new translational therapies. Resilience is the process of active adaptation to a stressor. In the context of neuroprotection, models of preconditioning and unique animal models of extreme physiology (such as hibernating species) reliably demonstrate resilience in the laboratory setting. In the clinical setting, resilience is observed in young patients and can be found in those with specific genetic polymorphisms. These important examples of resilience can help transform and extend the current neuroprotective framework from simply countering the injurious cascade into one that anticipates, monitors, and optimizes patients' physiological responses from the time of injury throughout the process of recovery. This review summarizes the underpinnings of key adaptations common to models of resilience and how this understanding can be applied to new neuroprotective approaches.

Neuronal GDPGP1 and glycogen metabolism: friend or foe?

The adult brain consumes glucose for energy needs and stores glucose as glycogen mainly in astrocytes. Schulz et al. (2020. J. Cell Biol.https://doi.org/10.1083/jcb.201807127) identify the stress-regulated metabolic enzyme GDPGP1 that promotes neuronal survival likely through glycogen reserves in mouse and C. elegans neurons.

Online continuing medical education as a key link for successful noncommunicable disease self-management: the CASALUD™ Model.

PURPOSE: The purpose of this study is to evaluate how the benefits of online continuing medical education (CME) provided to health care professionals traveled along a patient "educational chain". In this study, the educational chain begins with the influence that CME can have on the quality of health care, with subsequent influence on patient knowledge, disease self-management, and disease biomarkers. METHODS: A total of 422 patients with at least one noncommunicable disease (NCD) treated in eight different Mexican public health clinics were followed over 3 years. All clinics were participants in the CASALUD Model, an NCD care model for primary care, where all clinic staff were offered CME. Data were collected through a questionnaire on health care, patient disease knowledge, and self-management behaviors; blood samples and anthropometric measurements were collected to measure patient disease biomarkers. RESULTS: Between 2013 and 2015, the indexes measuring quality of health care, patient health knowledge, and diabetes self-management activities rose moderately but significantly (from 0.54 to 0.64, 0.80 to 0.84, and 0.62 to 0.67, respectively). Performing self-care activities - including owning and using a glucometer and belonging to a disease support group - saw the highest increase (from 0.65 to 0.75). A1C levels increased between 2013 and 2015 from 7.95 to 8.41% (63-68 mmol/mol) (P<0.001), and blood pressure decreased between 2014 and 2015 from 143.7/76.8 to 137.5/74.4 (systolic/diastolic reported in mmHg) (P<0.001). The mean levels of other disease biomarkers remained statistically unchanged, despite the improvements seen in the previous "links" of the educational chain. CONCLUSION: Online CME can effect certain changes in the educational chain linking quality of health care, patient knowledge, and self-management behaviors. However, in order to assure adequate NCD control, the entire health care system must be improved in tandem. Online CME programs, such as CASALUD's, are feasible strategies for impacting changes in disease self-management at a clinic level throughout a country.