Intestinal energy harvest is associated with post-bariatric surgery weight loss.

Background/Objectives: Metabolic and bariatric surgery (MBS) is the most effective treatment for severe obesity; however, a significant subset of patients does not achieve expected weight loss or have substantial weight recurrence over time. The intestinal energy harvest is a potential determinant of varying weight loss outcomes, but with limited exploration. We assess the relationships between diet, intestinal energy harvest, and weight outcomes over 24 months in individuals who have undergone MBS. Subjects/Methods: Calorie absorption was assessed with bomb calorimetry and dietary questionnaires before and after MBS. Within a total of 67 patients, fecal energy density was measured in 67, 56, 60, 67, 44, 47 samples at 0, 1, 6, 12, 18, and 24 months, respectively. Multivariate regression was developed to identify potential weight loss predictors, and random forest algorithms were employed to forecast weight results based on intestinal energy harvest. Results: Intestinal energy harvest enhanced the predictability of patient weight loss outcomes with random forest models. A notable difference in relative fecal energy content was observed between patients experiencing optimal and sub-optimal weight loss (p<0.01). Prior to MBS, an increased energy content in feces (indicating less energy absorption) is associated with greater weight loss after the operation. Associations between diet and energy harvest were insignificant. Conclusion: MBS changes energy harvest capacity post-surgery. A higher relative fecal energy content (lower energy absorption) at one month correlates with better weight loss outcomes at 6M, 12M, 18M and 24M post-MBS. Findings may guide the development of diagnostic tools and treatment guidelines for patients at risk of suboptimal weight loss outcomes. CLINICAL TRIAL REGISTRATION: The trial is registered at clinicaltrials.gov (NCT03065426).

Proportion-based normalizations outperform compositional data transformations in machine learning applications.

BACKGROUND: Normalization, as a pre-processing step, can significantly affect the resolution of machine learning analysis for microbiome studies. There are countless options for normalization scheme selection. In this study, we examined compositionally aware algorithms including the additive log ratio (alr), the centered log ratio (clr), and a recent evolution of the isometric log ratio (ilr) in the form of balance trees made with the PhILR R package. We also looked at compositionally naïve transformations such as raw counts tables and several transformations that are based on relative abundance, such as proportions, the Hellinger transformation, and a transformation based on the logarithm of proportions (which we call "lognorm"). RESULTS: In our evaluation, we used 65 metadata variables culled from four publicly available datasets at the amplicon sequence variant (ASV) level with a random forest machine learning algorithm. We found that different common pre-processing steps in the creation of the balance trees made very little difference in overall performance. Overall, we found that the compositionally aware data transformations such as alr, clr, and ilr (PhILR) performed generally slightly worse or only as well as compositionally naïve transformations. However, relative abundance-based transformations outperformed most other transformations by a small but reliably statistically significant margin. CONCLUSIONS: Our results suggest that minimizing the complexity of transformations while correcting for read depth may be a generally preferable strategy in preparing data for machine learning compared to more sophisticated, but more complex, transformations that attempt to better correct for compositionality. Video Abstract.

Western diet consumption impairs memory function via dysregulated hippocampus acetylcholine signaling.

Western diet (WD) consumption during early life developmental periods is associated with impaired memory function, particularly for hippocampus (HPC)-dependent processes. We developed an early life WD rodent model associated with long-lasting HPC dysfunction to investigate the neurobiological mechanisms mediating these effects. Rats received either a cafeteria-style WD (ad libitum access to various high-fat/high-sugar foods; CAF) or standard healthy chow (CTL) during the juvenile and adolescent stages (postnatal days 26-56). Behavioral and metabolic assessments were performed both before and after a healthy diet intervention period beginning at early adulthood. Results revealed HPC-dependent contextual episodic memory impairments in CAF rats that persisted despite the healthy diet intervention. Given that dysregulated HPC acetylcholine (ACh) signaling is associated with memory impairments in humans and animal models, we examined protein markers of ACh tone in the dorsal HPC (HPCd) in CAF and CTL rats. Results revealed significantly lower protein levels of vesicular ACh transporter in the HPCd of CAF vs. CTL rats, indicating chronically reduced ACh tone. Using intensity-based ACh sensing fluorescent reporter (iAChSnFr) in vivo fiber photometry targeting the HPCd, we next revealed that ACh release during object-contextual novelty recognition was highly predictive of memory performance and was disrupted in CAF vs. CTL rats. Neuropharmacological results showed that alpha 7 nicotinic ACh receptor agonist infusion in the HPCd during training rescued memory deficits in CAF rats. Overall, these findings reveal a functional connection linking early life WD intake with long-lasting dysregulation of HPC ACh signaling, thereby identifying an underlying mechanism for WD-associated memory impairments.

Mouse Adaptation of Human Inflammatory Bowel Diseases Microbiota Enhances Colonization Efficiency and Alters Microbiome Aggressiveness Depending on Recipient Colonic Inflammatory Environment.

Understanding the cause vs consequence relationship of gut inflammation and microbial dysbiosis in inflammatory bowel diseases (IBD) requires a reproducible mouse model of human-microbiota-driven experimental colitis. Our study demonstrated that human fecal microbiota transplant (FMT) transfer efficiency is an underappreciated source of experimental variability in human microbiota associated (HMA) mice. Pooled human IBD patient fecal microbiota engrafted germ-free (GF) mice with low amplicon sequence variant (ASV)-level transfer efficiency, resulting in high recipient-to-recipient variation of microbiota composition and colitis severity in HMA Il-10-/- mice. In contrast, mouse-to-mouse transfer of mouse-adapted human IBD patient microbiota transferred with high efficiency and low compositional variability resulting in highly consistent and reproducible colitis phenotypes in recipient Il-10-/- mice. Human-to-mouse FMT caused a population bottleneck with reassembly of microbiota composition that was host inflammatory environment specific. Mouse-adaptation in the inflamed Il-10-/- host reassembled a more aggressive microbiota that induced more severe colitis in serial transplant to Il-10-/- mice than the distinct microbiota reassembled in non-inflamed WT hosts. Our findings support a model of IBD pathogenesis in which host inflammation promotes aggressive resident bacteria, which further drives a feed-forward process of dysbiosis exacerbated gut inflammation. This model implies that effective management of IBD requires treating both the dysregulated host immune response and aggressive inflammation-driven microbiota. We propose that our mouse-adapted human microbiota model is an optimized, reproducible, and rigorous system to study human microbiome-driven disease phenotypes, which may be generalized to mouse models of other human microbiota-modulated diseases, including metabolic syndrome/obesity, diabetes, autoimmune diseases, and cancer.

Giardia Antagonizes Beneficial Functions of Indigenous and Therapeutic Intestinal Bacteria during Malnutrition.

Undernutrition in children commonly disrupts the structure and function of the small intestinal microbial community, leading to enteropathies, compromised metabolic health, and impaired growth and development. The mechanisms by which diet and microbes mediate the balance between commensal and pathogenic intestinal flora remain elusive. In a murine model of undernutrition, we investigated the direct interactions Giardia lamblia, a prevalent small intestinal pathogen, on indigenous microbiota and specifically on Lactobacillus strains known for their mucosal and growth homeostatic properties. Our research reveals that Giardia colonization shifts the balance of lactic acid bacteria, causing a relative decrease in Lactobacillus spp . and an increase in Bifidobacterium spp . This alteration corresponds with a decrease in multiple indicators of mucosal and nutritional homeostasis. Additionally, protein-deficient conditions coupled with Giardia infection exacerbate the rise of primary bile acids and susceptibility to bile acid-induced intestinal barrier damage. In epithelial cell monolayers, Lactobacillus spp . mitigated bile acid-induced permeability, showing strain-dependent protective effects. In vivo, L. plantarum, either alone or within a Lactobacillus spp consortium, facilitated growth in protein-deficient mice, an effect attenuated by Giardia , despite not inhibiting Lactobacillus colonization. These results highlight Giardia's potential role as a disruptor of probiotic functional activity, underscoring the imperative for further research into the complex interactions between parasites and bacteria under conditions of nutritional deficiency.

Acute Steroid-Refractory Gastrointestinal Graft-Versus-Host Disease Is Not Associated With Significant Differences in Gut Taxonomic Composition Compared to Steroid-Sensitive Gastrointestinal Graft-Versus-Host Disease Immediately Before Onset of Disease.

Taxonomic composition of the gut microbiota at the time of neutrophil engraftment is associated with the development of acute gastrointestinal graft-versus-host disease (GI GVHD) in patients undergoing allogeneic hematopoietic stem cell transplantation. However, less is known about the relationship between the gut microbiota and development of steroid-refractory GI GVHD immediately before the onset of disease. Markers of steroid-refractory GI GVHD are needed to identify patients who may benefit from the early initiation of non-corticosteroid-based GVHD treatment. Our aim was to identify differences in taxonomic composition in stool samples from patients without GVHD, with steroid-responsive GVHD and with steroid-refractory GI GVHD to identify predictive microbiome biomarkers of steroid-refractory GI GVHD. We conducted a retrospective case-control, single institution study, performing shotgun metagenomic sequencing on stool samples from patients with (n = 36) and without GVHD (n = 34) matched for time since transplantation. We compared the taxonomic composition of the gut microbiome in those with steroid-sensitive GI GVHD (n = 17) and steroid-refractory GI GVHD (n = 19) to each other and to those without GVHD. We also performed associations between steroid-refractory GI GVHD, gut taxonomic composition, and fecal calprotectin, a marker of GI GVHD to develop composite fecal markers of steroid-refractory GVHD before the onset of GI disease. We found that fecal samples within 30 days of GVHD onset from patients with and without GVHD or with and without steroid-refractory GI GVHD did not differ significantly in Shannon diversity (alpha-diversity) or in overall taxonomic composition (beta-diversity). Although those patients without GVHD had higher relative abundance of Clostridium spp., those with and without steroid-refractory GI GVHD did not significantly differ in taxonomic composition between one another. In our study, fecal calprotectin before disease onset was significantly higher in patients with GVHD compared to those without GVHD and higher in patients with steroid-refractory GI GVHD compared to steroid-sensitive GI GVHD. No taxa were significantly associated with higher levels of calprotectin.

A consortia of clinical E. coli strains with distinct in vitro adherent/invasive properties establish their own co-colonization niche and shape the intestinal microbiota in inflammation-susceptible mice.

BACKGROUND: Inflammatory bowel disease (IBD) patients experience recurrent episodes of intestinal inflammation and often follow an unpredictable disease course. Mucosal colonization with adherent-invasive Escherichia coli (AIEC) are believed to perpetuate intestinal inflammation. However, it remains unclear if the 24-year-old AIEC in vitro definition fully predicts mucosal colonization in vivo. To fill this gap, we have developed a novel molecular barcoding approach to distinguish strain variants in the gut and have integrated this approach to explore mucosal colonization of distinct patient-derived E. coli isolates in gnotobiotic mouse models of colitis. RESULTS: Germ-free inflammation-susceptible interleukin-10-deficient (Il10-/-) and inflammation-resistant WT mice were colonized with a consortium of AIEC and non-AIEC strains, then given a murine fecal transplant to provide niche competition. E. coli strains isolated from human intestinal tissue were each marked with a unique molecular barcode that permits identification and quantification by barcode-targeted sequencing. 16S rRNA sequencing was used to evaluate the microbiome response to E. coli colonization. Our data reveal that specific AIEC and non-AIEC strains reproducibly colonize the intestinal mucosa of WT and Il10-/- mice. These E. coli expand in Il10-/- mice during inflammation and induce compositional dysbiosis to the microbiome in an inflammation-dependent manner. In turn, specific microbes co-evolve in inflamed mice, potentially diversifying E. coli colonization patterns. We observed no selectivity in E. coli colonization patterns in the fecal contents, indicating minimal selective pressure in this niche from host-microbe and interbacterial interactions. Because select AIEC and non-AIEC strains colonize the mucosa, this suggests the in vitro AIEC definition may not fully predict in vivo colonization potential. Further comparison of seven E. coli genomes pinpointed unique genomic features contained only in highly colonizing strains (two AIEC and two non-AIEC). Those colonization-associated features may convey metabolic advantages (e.g., iron acquisition and carbohydrate consumption) to promote efficient mucosal colonization. CONCLUSIONS: Our findings establish the in vivo mucosal colonizer, not necessarily AIEC, as a principal dysbiosis driver through crosstalk with host and associated microbes. Furthermore, we highlight the utility of high-throughput screens to decode the in vivo colonization dynamics of patient-derived bacteria in murine models. Video Abstract.

Calorie restriction outperforms bariatric surgery in a murine model of obesity and triple-negative breast cancer.

Obesity promotes triple-negative breast cancer (TNBC), and effective interventions are urgently needed to break the obesity-TNBC link. Epidemiologic studies indicate that bariatric surgery reduces TNBC risk, while evidence is limited or conflicted for weight loss via low-fat diet (LFD) or calorie restriction (CR). Using a murine model of obesity-driven TNBC, we compared the antitumor effects of vertical sleeve gastrectomy (VSG) with LFD, chronic CR, and intermittent CR. Each intervention generated weight and fat loss and suppressed tumor growth relative to obese mice (greatest suppression with CR). VSG and CR regimens exerted both similar and unique effects, as assessed using multiomics approaches, in reversing obesity-associated transcript, epigenetics, secretome, and microbiota changes and restoring antitumor immunity. Thus, in a murine model of TNBC, bariatric surgery and CR each reverse obesity-driven tumor growth via shared and distinct antitumor mechanisms, and CR is superior to VSG in reversing obesity's procancer effects.

Metabolic and Immune Consequences of Antibiotic Related Microbiome Alterations during first-line Tuberculosis Treatment in Bamako, Mali.

Background: Tuberculosis (TB) infection is known to lead to the unbalance of the gut microbiota and act synergistically on the decline of the host immune response, when untreated. Moreover, previous work has found a correlation between dysbiosis in the gut microbiota composition and the use of antibiotics. However, there is a need for an in-depth understanding of the metabolic and immune consequences of antibiotic-related microbiome alterations during first-line TB treatment. Methods: In a longitudinal cohort study, which included TB-infected cohorts and healthy individuals (control group), we studied the anti-TB-related changes in the gut microbiota composition and related functional consequences. Sputum, whole blood and stool samples were collected from participants at four time-points including before (Month-0), during (Month-2), at the end of drug treatment (Month-6) and 9 months after treatment (Month-15). Controls were sampled at inclusion and Month-6. We analyzed the microbiota composition and microbial functional pathways with shotgun metagenomics, analyzed the blood metabolomics using high-performance liquid chromatography (HPLC), and measured the levels of metabolites and cytokines with cytometric bead array. Results: We found that the gut microbiota of patients infected with TB was different from that of the healthy controls. The gut microbiota became similar to healthy controls after treatment but was still significantly different after 6 months treatment and at the follow up 9 months after treatment. Our data also showed disturbance in the plasma metabolites such as tryptophan and tricarboxylic acids components of patients during TB treatment. Levels of IL-4, IL-6, IL-10, and IFN-γ decreased during treatment and levels were maintained after treatment completion, while IL-17A known to have a strong link with the gut microbiota was highly expressed during treatment period and longer than the 9-month post treatment completion. We found that some fatty acids were negatively correlated with the abundance of taxa. For example, Roseburia, Megasphaera, and alpha proteobacterium HIMB5 species were negatively correlated (rho = -0.6) with the quinolinate production. Conclusion: Changes in the composition and function of gut microbiota was observed in TB patients before and after treatment compared to healthy controls. The differences persisted at nine months after treatment completion. Alterations in some bacterial taxa were correlated to the changes in metabolite levels in peripheral blood, thus the altered microbial community might lead to changes in immune status that influence the disease outcome and future resistance to infections.

Gut Microbial Changes Following Fecal Microbiota Transplantation for D-Lactic Acidosis in Two Children.

D-lactic acidosis (D-LA) is an uncommon complication of short bowel syndrome characterized by elevated plasma D-lactate and encephalopathy. Treatments include rehydration, dietary carbohydrate restriction, and antibiotics to alter the gut microbiota. Fecal microbiota transplantation (FMT) has recently been used in children to successfully treat D-LA. We compared the clinical course and then utilized metagenomic shotgun sequencing to describe changes in the composition and function of the intestinal microbiome following FMT in 2 patients with recurrent D-LA. FMT altered the composition of the fecal microbiota in these 2 patients with recurrent D-LA, though not necessarily in a consistent manner. Importantly, microbial metabolic pathways were also impacted by FMT, which may be critical for achieving desired clinical outcomes. While sample size limits the generalizability of our results, these findings set the stage for further understanding of the role of microbes in the pathogenesis of recurrent D-LA.

Longer-Term Weight Loss Outcomes Are Not Primarily Driven by Diet Following Roux-en-Y Gastric Bypass and Sleeve Gastrectomy.

Metabolic and bariatric surgery (MBS) is the most effective long-term treatment for Class III obesity. Reduced dietary intake is considered a behavioral driver of post-surgical weight loss, but limited data have examined this association. Therefore, this study examined prospective, longitudinal relationships between dietary intake and weight loss over 24 months following Roux-en-Y Gastric Bypass and Sleeve Gastrectomy. Relationships between weight loss and dietary intake were examined using a validated 24-h dietary recall method. Associations between total energy/macronutrient intake and weight loss outcomes were assessed at 12-, 18-, and 24-months following MBS, defining patients as "responders" and "suboptimal responders". Consistent with previous literature, 12-month responders and suboptimal responders showed significant associations between weight loss and energy (p = 0.018), protein (p = 0.002), and total fat intake (p = 0.005). However, this study also revealed that many of these associations are no longer significant 24 months post-MBS (p > 0.05), despite consistent weight loss trends. This study suggests a short-term signal between these dietary factors and weight loss outcomes 12 months post-MBS; however, this signal does not persist beyond 12 months. These results are essential for interpreting and designing clinical studies measuring long-term post-surgical weight loss outcomes.

Western diet consumption impairs memory function via dysregulated hippocampus acetylcholine signaling.

Western diet (WD) consumption during development yields long-lasting memory impairments, yet the underlying neurobiological mechanisms remain elusive. Here we developed an early life WD rodent model to evaluate whether dysregulated hippocampus (HPC) acetylcholine (ACh) signaling, a pathology associated with memory impairment in human dementia, is causally-related to WD-induced cognitive impairment. Rats received a cafeteria-style WD (access to various high-fat/high-sugar foods; CAF) or healthy chow (CTL) during the juvenile and adolescent periods (postnatal days 26-56). Behavioral, metabolic, and microbiome assessments were performed both before and after a 30-day healthy diet intervention beginning at early adulthood. Results revealed CAF-induced HPC-dependent contextual episodic memory impairments that persisted despite healthy diet intervention, whereas CAF was not associated with long-term changes in body weight, body composition, glucose tolerance, anxiety-like behavior, or gut microbiome. HPC immunoblot analyses after the healthy diet intervention identified reduced levels of vesicular ACh transporter in CAF vs. CTL rats, indicative of chronically reduced HPC ACh tone. To determine whether these changes were functionally related to memory impairments, we evaluated temporal HPC ACh binding via ACh-sensing fluorescent reporter in vivo fiber photometry during memory testing, as well as whether the memory impairments could be rescued pharmacologically. Results revealed dynamic HPC ACh binding during object-contextual novelty recognition was highly predictive of memory performance and was disrupted in CAF vs. CTL rats. Further, HPC alpha-7 nicotinic receptor agonist infusion during consolidation rescued memory deficits in CAF rats. Overall, these findings identify dysregulated HPC ACh signaling as a mechanism underlying early life WD-associated memory impairments.

Dietary Calcium and Risk of Microscopic Colitis.

BACKGROUND: Microscopic colitis (MC) is an increasingly common cause of watery diarrhea particularly in older individuals. The role of diet in MC has received little study. METHODS: We conducted a case-control study at a single institution enrolling patients referred for elective outpatient colonoscopy for diarrhea. Patients were classified as cases with MC or non-MC controls after a review of colon biopsies by 1 research pathologist. Study subjects were interviewed by a trained telephone interviewer using a validated food frequency questionnaire. Adherent microbes were evaluated from colonic biopsies using 16s rRNA sequencing. RESULTS: The study population included 106 cases with MC and 215 controls. Compared with controls, the cases were older, better educated, and more likely to be female. Cases with MC had lower body mass index and were more likely to have lost weight. Subjects in the highest quartile of dietary calcium intake had a lower risk of MC compared with those in the lowest quartile (adjusted odds ratio 0.22, 95% confidence interval 0.07-0.76). The findings were not explained by dairy intake, body mass index, or weight loss. We found that dietary calcium intake had significant associations with the abundance of Actinobacteria and Coriobacteriales in the microbial community of colonic biopsies. DISCUSSION: Compared with patients with diarrhea, cases with MC had a lower intake of dietary calcium. Diet can be associated with alterations in the gut microbiota and with luminal factors that could affect the risk of MC.

Giardia hinders growth by disrupting nutrient metabolism independent of inflammatory enteropathy.

Giardia lamblia (Giardia) is among the most common intestinal pathogens in children in low- and middle-income countries (LMICs). Although Giardia associates with early-life linear growth restriction, mechanistic explanations for Giardia-associated growth impairments remain elusive. Unlike other intestinal pathogens associated with constrained linear growth that cause intestinal or systemic inflammation or both, Giardia seldom associates with chronic inflammation in these children. Here we leverage the MAL-ED longitudinal birth cohort and a model of Giardia mono-association in gnotobiotic and immunodeficient mice to propose an alternative pathogenesis of this parasite. In children, Giardia results in linear growth deficits and gut permeability that are dose-dependent and independent of intestinal markers of inflammation. The estimates of these findings vary between children in different MAL-ED sites. In a representative site, where Giardia associates with growth restriction, infected children demonstrate broad amino acid deficiencies, and overproduction of specific phenolic acids, byproducts of intestinal bacterial amino acid metabolism. Gnotobiotic mice require specific nutritional and environmental conditions to recapitulate these findings, and immunodeficient mice confirm a pathway independent of chronic T/B cell inflammation. Taken together, we propose a new paradigm that Giardia-mediated growth faltering is contingent upon a convergence of this intestinal protozoa with nutritional and intestinal bacterial factors.

A consortia of clinical E. coli strains with distinct in-vitro adherent/invasive properties establish their own co-colonization niche and shape the intestinal microbiota in inflammation-susceptible mice.

Background: Inflammatory bowel disease (IBD) patients experience recurrent episodes of intestinal inflammation and often follow an unpredictable disease course. Mucosal colonization with adherent-invasive Escherichia coli (AIEC) are believed to perpetuate intestinal inflammation. However, it remains unclear if the 24-year-old AIEC in-vitro definition fully predicts mucosal colonization in-vivo. To fill this gap, we have developed a novel molecular barcoding approach to distinguish strain variants in the gut and have integrated this approach to explore mucosal colonization of distinct patient-derived E. coli isolates in gnotobiotic mouse models of colitis. Results: Germ-free inflammation-susceptible interleukin-10-deficient (Il10-/-) and inflammation-resistant WT mice were colonized with a consortia of AIEC and non-AIEC strains, then given a murine fecal transplant to provide niche competition. E. coli strains isolated from human intestinal tissue were each marked with a unique molecular barcode that permits identification and quantification by barcode-targeted sequencing. 16S rRNA sequencing was used to evaluate the microbiome response to E. coli colonization. Our data reveal that specific AIEC and non-AIEC strains reproducibly colonize the intestinal mucosa of WT and Il10-/- mice. These E. coli expand in Il10-/- mice during inflammation and induce compositional dysbiosis to the microbiome in an inflammation-dependent manner. In turn, specific microbes co-evolve in inflamed mice, potentially diversifying E. coli colonization patterns. We observed no selectivity in E. coli colonization patterns in the fecal contents, indicating minimal selective pressure in this niche from host-microbe and interbacterial interactions. Because select AIEC and non-AIEC strains colonize the mucosa, this suggests the in vitro AIEC definition may not fully predict in vivo colonization potential. Further comparison of seven E. coli genomes pinpointed unique genomic features contained only in highly colonizing strains (two AIEC and two non-AIEC). Those colonization-associated features may convey metabolic advantages (e.g., iron acquisition and carbohydrate consumption) to promote efficient mucosal colonization. Conclusions: Our findings establish the in-vivo mucosal colonizer, not necessarily AIEC, as a principal dysbiosis driver through crosstalk with host and associated microbes. Furthermore, we highlight the utility of high-throughput screens to decode the in-vivo colonization dynamics of patient-derived bacteria in murine models.

Maternal low-calorie sweetener consumption rewires hypothalamic melanocortin circuits via a gut microbial co-metabolite pathway.

The prevalence of obesity and type 2 diabetes is growing at an alarming rate, including among pregnant women. Low-calorie sweeteners (LCSs) have increasingly been used as an alternative to sugar to deliver a sweet taste without the excessive caloric load. However, there is little evidence regarding their biological effects, particularly during development. Here, we used a mouse model of maternal LCS consumption to explore the impact of perinatal LCS exposure on the development of neural systems involved in metabolic regulation. We report that adult male, but not female, offspring from both aspartame- and rebaudioside A-exposed dams displayed increased adiposity and developed glucose intolerance. Moreover, maternal LCS consumption reorganized hypothalamic melanocortin circuits and disrupted parasympathetic innervation of pancreatic islets in male offspring. We then identified phenylacetylglycine (PAG) as a unique metabolite that was upregulated in the milk of LCS-fed dams and the serum of their pups. Furthermore, maternal PAG treatment recapitulated some of the key metabolic and neurodevelopmental abnormalities associated with maternal LCS consumption. Together, our data indicate that maternal LCS consumption has enduring consequences on the offspring's metabolism and neural development and that these effects are likely to be mediated through the gut microbial co-metabolite PAG.

Vaginal Microbiome Metagenome Inference Accuracy: Differential Measurement Error according to Community Composition.

Several studies have compared metagenome inference performance in different human body sites; however, none specifically reported on the vaginal microbiome. Findings from other body sites cannot easily be generalized to the vaginal microbiome due to unique features of vaginal microbial ecology, and investigators seeking to use metagenome inference in vaginal microbiome research are "flying blind" with respect to potential bias these methods may introduce into analyses. We compared the performance of PICRUSt2 and Tax4Fun2 using paired 16S rRNA gene amplicon sequencing and whole-metagenome sequencing data from vaginal samples from 72 pregnant individuals enrolled in the Pregnancy, Infection, and Nutrition (PIN) cohort. Participants were selected from those with known birth outcomes and adequate 16S rRNA gene amplicon sequencing data in a case-control design. Cases experienced early preterm birth (<32 weeks of gestation), and controls experienced term birth (37 to 41 weeks of gestation). PICRUSt2 and Tax4Fun2 performed modestly overall (median Spearman correlation coefficients between observed and predicted KEGG ortholog [KO] relative abundances of 0.20 and 0.22, respectively). Both methods performed best among Lactobacillus crispatus-dominated vaginal microbiotas (median Spearman correlation coefficients of 0.24 and 0.25, respectively) and worst among Lactobacillus iners-dominated microbiotas (median Spearman correlation coefficients of 0.06 and 0.11, respectively). The same pattern was observed when evaluating correlations between univariable hypothesis test P values generated with observed and predicted metagenome data. Differential metagenome inference performance across vaginal microbiota community types can be considered differential measurement error, which often causes differential misclassification. As such, metagenome inference will introduce hard-to-predict bias (toward or away from the null) in vaginal microbiome research. IMPORTANCE Compared to taxonomic composition, the functional potential within a bacterial community is more relevant to establishing mechanistic understandings and causal relationships between the microbiome and health outcomes. Metagenome inference attempts to bridge the gap between 16S rRNA gene amplicon sequencing and whole-metagenome sequencing by predicting a microbiome's gene content based on its taxonomic composition and annotated genome sequences of its members. Metagenome inference methods have been evaluated primarily among gut samples, where they appear to perform fairly well. Here, we show that metagenome inference performance is markedly worse for the vaginal microbiome and that performance varies across common vaginal microbiome community types. Because these community types are associated with sexual and reproductive outcomes, differential metagenome inference performance will bias vaginal microbiome studies, obscuring relationships of interest. Results from such studies should be interpreted with substantial caution and the understanding that they may over- or underestimate associations with metagenome content.

Reproducible changes in the anorexia nervosa gut microbiota following inpatient therapy remain distinct from non-eating disorder controls.

The composition of the gut microbiota in patients with anorexia nervosa (AN), and the ability of this microbial community to influence the host, remains uncertain. To achieve a broader understanding of the role of the intestinal microbiota in patients with AN, we collected fecal samples before and following clinical treatment at two geographically distinct eating disorder units (Center of Excellence for Eating Disorders [UNC-CH] and ACUTE Center for Eating Disorders [Denver Health]). Gut microbiotas were characterized in patients with AN, before and after inpatient treatment, and in non-eating disorder (non-ED) controls using shotgun metagenomic sequencing. The impact of inpatient treatment on the AN gut microbiota was remarkably consistent between eating disorder units. Although weight in patients with AN showed improvements, AN microbiotas post-treatment remained distinct from non-ED controls. Additionally, AN gut microbiotas prior to treatment exhibited more fermentation pathways and a lower ability to degrade carbohydrates than non-ED controls. As the intestinal microbiota can influence nutrient metabolism, our data highlight the complex microbial communities in patients with AN as an element needing further attention post inpatient treatment. Additionally, this study defines the effects of renourishment on the AN gut microbiota and serves as a platform to develop precision nutrition approaches to potentially mitigate impediments to recovery.

Ca:Mg ratio, medium-chain fatty acids, and the gut microbiome.

BACKGROUND & AIMS: Ketogenic medium-chain fatty acids (MCFAs) with profound health benefits are commonly found in dairy products, palm kernel oil and coconut oil. We hypothesize that magnesium (Mg) supplementation leads to enhanced gut microbial production of MCFAs and, in turn, increased circulating MCFAs levels. METHODS: We tested this hypothesis in the Personalized Prevention of Colorectal Cancer Trial (PPCCT) (NCT01105169), a double-blind 2 × 2 factorial randomized controlled trial enrolling 240 participants. Six 24-h dietary recalls were performed for all participants at the baseline and during the intervention period. Based on the baseline 24-h dietary recalls, the Mg treatment used a personalized dose of Mg supplementation that would reduce the calcium (Ca): Mg intake ratio to around 2.3. We measured plasma MCFAs, sugars, ketone bodies and tricarboxylic acid cycle (TCA cycle) metabolites using the Metabolon's global Precision Metabolomics™ LC-MS platform. Whole-genome shotgun metagenomics (WGS) sequencing was performed to assess microbiota in stool samples, rectal swabs, and rectal biopsies. RESULTS: Personalized Mg treatment (mean dose 205.58 mg/day with a range from 77.25 to 389.55 mg/day) significantly increased the plasma levels of C7:0, C8:0, and combined C7:0 and C8:0 by 18.45%, 25.28%, and 24.20%, respectively, compared to 14.15%, 10.12%, and 12.62% decreases in the placebo arm. The effects remain significant after adjusting for age, sex, race and baseline level (P = 0.0126, P = 0.0162, and P = 0.0031, respectively) and FDR correction at 0.05 (q = 0.0324 for both C7:0 and C8:0). Mg treatment significantly reduced the plasma level of sucrose compared to the placebo arm (P = 0.0036 for multivariable-adjusted and P = 0.0216 for additional FDR correction model) whereas alterations in daily intakes of sucrose, fructose, glucose, maltose and C8:0 from baseline to the end of trial did not differ between two arms. Mediation analysis showed that combined C7:0 and C8:0 partially mediated the effects of Mg treatment on total and individual ketone bodies (P for indirect effect = 0.0045, 0.0043, and 0.03, respectively). The changes in plasma levels of C7:0 and C8:0 were significantly and positively correlated with the alterations in stool microbiome α diversity (r = 0.51, p = 0.0023 and r = 0.34, p = 0.0497, respectively) as well as in stool abundance for the signatures of MCFAs-related microbiota with acyl-ACP thioesterase gene producing C7:0 (r = 0.46, p = 0.0067) and C8:0 (r = 0.49, p = 0.003), respectively, following Mg treatment. CONCLUSIONS: Optimizing Ca:Mg intake ratios to around 2.3 through 12-week personalized Mg supplementation leads to increased circulating levels of MCFAs (i.e. C7:0 and C8:0), which is attributed to enhanced production from gut microbial fermentation and, maybe, sucrose consumption.