Impact of diet on human nutrition, immune response, gut microbiome, and cognition in an isolated and confined mission environment.

Long-duration spaceflight impacts human physiology, including well documented immune system dysregulation. The space food system has the potential to serve as a countermeasure to maladaptive physiological changes during spaceflight. However, the relationship between dietary requirements, the food system, and spaceflight adaptation requires further investigation to adequately define countermeasures and prioritize resources on future spaceflight missions. We evaluated the impact of an enhanced spaceflight diet, with increased quantity and variety of fruits, vegetables, fish, and other foods rich in flavonoids and omega-3 fatty acids, compared to a standard spaceflight diet on multiple health and performance outcomes in 16 subjects over four 45-day closed chamber missions in the NASA Human Exploration Research Analog (HERA). Subjects consuming the enhanced spaceflight diet had lower cholesterol levels, lower stress (i.e. cortisol levels), better cognitive speed, accuracy, and attention, and a more stable microbiome and metatranscriptome than subjects consuming the standard diet. Although no substantial changes were observed in the immune response, there were also no immune challenges, such as illness or infection, so the full benefits of the diet may not have been apparent in these analog missions. These results indicate that a spaceflight diet rich in fruits, vegetables, and omega-3 fatty acids produces significant health and performance benefits even over short durations. Further investigation is required to fully develop dietary countermeasures to physiological decrements observed during spaceflight. These results will have implications for food resource prioritization on spaceflight missions.

Sampling from four geographically divergent young female populations demonstrates forensic geolocation potential in microbiomes.

Studies of human microbiomes using new sequencing techniques have increasingly demonstrated that their ecologies are partly determined by the lifestyle and habits of individuals. As such, significant forensic information could be obtained from high throughput sequencing of the human microbiome. This approach, combined with multiple analytical techniques demonstrates that bacterial DNA can be used to uniquely identify an individual and to provide information about their life and behavioral patterns. However, the transformation of these findings into actionable forensic information, including the geolocation of the samples, remains limited by incomplete understanding of the effects of confounding factors and the paucity of diverse sequences. We obtained 16S rRNA sequences of stool and oral microbiomes collected from 206 young and healthy females from four globally diverse populations, in addition to supporting metadata, including dietary and medical information. Analysis of these microbiomes revealed detectable geolocation signals between the populations, even for populations living within the same city. Accounting for other lifestyle variables, such as diet and smoking, lessened but does not remove the geolocation signal.

Progressive dysbiosis of human orodigestive microbiota along the sequence of gastroesophageal reflux, Barrett's esophagus and esophageal adenocarcinoma.

The incidence of esophageal adenocarcinoma (EA) has drastically increased in the United States since 1970s for unclear reasons. We hypothesized that the widespread usage of antibiotics has increased the procarcinogenic potential of the orodigestive microbiota along the sequence of gastroesophageal reflux (GR), Barrett's esophagus (BE) and EA phenotypes. This case control study included normal controls (NC) and three disease phenotypes GR, BE and EA. Microbiota in the mouth, esophagus, and stomach, and rectum were analyzed using 16S rRNA gene sequencing. Overall, we discovered 44 significant pairwise differences in abundance of microbial taxa between the four phenotypes, with 12 differences in the mouth, 21 in the esophagus, two in the stomach, and nine in the rectum. Along the GR→BE→EA sequence, oral and esophageal microbiota were more diversified, the dominant genus Streptococcus was progressively depleted while six other genera Atopobium, Actinomyces, Veillonella, Ralstonia, Burkholderia and Lautropia progressively enriched. In NC, Streptococcus appeared to control populations of other genera in the foregut via numerous negative and positive connections, while in disease states, the rich network was markedly simplified. Inferred gene functional content showed a progressive enrichment through the stages of EA development in genes encoding antibiotic resistance, ligands of Toll-like and NOD-like receptors, nitrate-nitrite-nitric oxide pathway and acetaldehyde metabolism. The orodigestive microbiota is in a progressive dysbiotic state along the GR-BE-EA sequence. The increasing dysbiosis and antibiotic and procarcinogenic genes in the disease states warrants further study to define their roles in EA pathogenesis.

Gut microbiome differences between amyotrophic lateral sclerosis patients and spouse controls.

Objective: Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disease that is incurable and ultimately fatal. Few therapeutic options are available to patients. In this study, we explored differences in microbiome composition associated with ALS. Methods: We compared the gut microbiome and inflammatory marker profiles of ALS patients (n = 10) to those of their spouses (n = 10). Gut microbiome profiles were determined by 16S rRNA gene sequencing. Results: The gut microbial communities of the ALS patients were more diverse and were deficient in Prevotella spp. compared with those of their spouses. In contrast, healthy couples (n = 10 couples of the opposite sex) recruited from the same geographic region as the patient population did not exhibit these differences. Stool and plasma inflammatory markers were similar between ALS patients and their spouses. Predictive analysis of microbial enzymes revealed that ALS patients had decreased activity in several metabolic pathways, including carbon metabolism, butyrate metabolism, and systems involving histidine kinase and response regulators. Conclusions: ALS patients exhibit differences in their gut microbial communities compared with spouse controls. Our findings suggest that modifying the gut microbiome, such as via amelioration of Prevotella spp. deficiency, and/or altering butyrate metabolism may have translational value for ALS treatment.

Differential network analysis of oral microbiome metatranscriptomes identifies community scale metabolic restructuring in dental caries.

Dental caries is a microbial disease and the most common chronic health condition, affecting nearly 3.5 billion people worldwide. In this study, we used a multiomics approach to characterize the supragingival plaque microbiome of 91 Australian children, generating 658 bacterial and 189 viral metagenome-assembled genomes with transcriptional profiling and gene-expression network analysis. We developed a reproducible pipeline for clustering sample-specific genomes to integrate metagenomics and metatranscriptomics analyses regardless of biosample overlap. We introduce novel feature engineering and compositionally-aware ensemble network frameworks while demonstrating their utility for investigating regime shifts associated with caries dysbiosis. These methods can be applied when differential abundance modeling does not capture statistical enrichments or the results from such analysis are not adequate for providing deeper insight into disease. We identified which organisms and metabolic pathways were central in a coexpression network as well as how these networks were rewired between caries and caries-free phenotypes. Our findings provide evidence of a core bacterial microbiome that was transcriptionally active in the supragingival plaque of all participants regardless of phenotype, but also show highly diagnostic changes in the ways that organisms interact. Specifically, many organisms exhibit high connectedness with central carbon metabolism to Cardiobacterium and this shift serves a bridge between phenotypes. Our evidence supports the hypothesis that caries is a multifactorial ecological disease.

Urethral Catheter Biofilms Reveal Plasticity in Bacterial Composition and Metabolism and Withstand Host Immune Defenses in Hypoxic Environment.

Biofilms composed of multiple microorganisms colonize the surfaces of indwelling urethral catheters that are used serially by neurogenic bladder patients and cause chronic infections. Well-adapted pathogens in this niche are Escherichia coli, Proteus, and Enterococcus spp., species that cycle through adhesion and multilayered cell growth, trigger host immune responses, are starved off nutrients, and then disperse. Viable microbial foci retained in the urinary tract recolonize catheter surfaces. The molecular adaptations of bacteria in catheter biofilms (CBs) are not well-understood, promising new insights into this pathology based on host and microbial meta-omics analyses from clinical specimens. We examined catheters from nine neurogenic bladder patients longitudinally over up to 6 months. Taxonomic analyses from 16S rRNA gene sequencing and liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based proteomics revealed that 95% of all catheter and corresponding urinary pellet (UP) samples contained bacteria. CB biomasses were dominated by Enterobacteriaceae spp. and often accompanied by lactic acid and anaerobic bacteria. Systemic antibiotic drug treatments of patients resulted in either transient or lasting microbial community perturbations. Neutrophil effector proteins were abundant not only in UP but also CB samples, indicating their penetration of biofilm surfaces. In the context of one patient who advanced to a kidney infection, Proteus mirabilis proteomic data suggested a combination of factors associated with this disease complication: CB biomasses were high; the bacteria produced urease alkalinizing the pH and triggering urinary salt deposition on luminal catheter surfaces; P. mirabilis utilized energy-producing respiratory systems more than in CBs from other patients. The NADH:quinone oxidoreductase II (Nqr), a Na+ translocating enzyme not operating as a proton pump, and the nitrate reductase A (Nar) equipped the pathogen with electron transport chains promoting growth under hypoxic conditions. Both P. mirabilis and E. coli featured repertoires of transition metal ion acquisition systems in response to human host-mediated iron and zinc sequestration. We discovered a new drug target, the Nqr respiratory system, whose deactivation may compromise P. mirabilis growth in a basic pH milieu. Animal models would not allow such molecular-level insights into polymicrobial biofilm metabolism and interactions because the complexity cannot be replicated.

Characterizing Microbial Signatures on Sculptures and Paintings of Similar Provenance.

The preservation of artwork challenges museums, collectors, and art enthusiasts. Currently, reducing moisture, adjusting the type of lighting, and preventing the formation of mold are primary methods to preserving and preventing deterioration. Other methods such as ones based in detailed knowledge of molecular biology such as microbial community characterization using polymerase chain reaction (PCR) and sequencing have yet to be explored. Such molecular biology approaches are essential to explore as some environmental bacteria are capable of oxidizing nonpolar chemical substances rich in hydrocarbons such as oil-based paints. Using 16S rDNA Illumina Sequencing, we demonstrate a novel finding that there are differing bacterial communities for artwork from roughly the same era when comparing paintings on wood, paintings on canvases, and sculptures made of stone and marble. We also demonstrate that there are specific genera such as Aeromonas known for having oxidase positive strains, present on paintings on wood and paintings on canvas that could potentially be responsible for deterioration and fading as such organisms produce water or hydrogen peroxide as a byproduct of cytochrome c oxidase activity. The advantages of these genomics-based approaches to characterizing the microbial population on deteriorating artwork provides immense potential by identifying potentially damaging species that may not be detected using conventional methods in addition to addressing challenges to identification, restoration, and preservation efforts.

Oral Microbial Species and Virulence Factors Associated with Oral Squamous Cell Carcinoma.

The human microbiome has been the focus of numerous research efforts to elucidate the pathogenesis of human diseases including cancer. Oral cancer mortality is high when compared with other cancers, as diagnosis often occurs during late stages. Its prevalence has increased in the USA over the past decade and accounts for over 40,000 new cancer patients each year. Additionally, oral cancer pathogenesis is not fully understood and is likely multifactorial. To unravel the relationships that are associated with the oral microbiome and their virulence factors, we used 16S rDNA and metagenomic sequencing to characterize the microbial composition and functional content in oral squamous cell carcinoma (OSCC) tumor tissue, non-tumor tissue, and saliva from 18 OSCC patients. Results indicate a higher number of bacteria belonging to the Fusobacteria, Bacteroidetes, and Firmicutes phyla associated with tumor tissue when compared with all other sample types. Additionally, saliva metaproteomics revealed a significant increase of Prevotella in five OSCC subjects, while Corynebacterium was mostly associated with ten healthy subjects. Lastly, we determined that there are adhesion and virulence factors associated with Streptococcus gordonii as well as from known oral pathogens belonging to the Fusobacterium genera found mostly in OSCC tissues. From these results, we propose that not only will the methods utilized in this study drastically improve OSCC diagnostics, but the organisms and specific virulence factors from the phyla detected in tumor tissue may be excellent biomarkers for characterizing disease progression.

Traditional Human Populations and Nonhuman Primates Show Parallel Gut Microbiome Adaptations to Analogous Ecological Conditions.

Compared with urban-industrial populations, small-scale human communities worldwide share a significant number of gut microbiome traits with nonhuman primates. This overlap is thought to be driven by analogous dietary triggers; however, the ecological and functional bases of this similarity are not fully understood. To start addressing this issue, fecal metagenomes of BaAka hunter-gatherers and traditional Bantu agriculturalists from the Central African Republic were profiled and compared with those of a sympatric western lowland gorilla group (Gorilla gorilla gorilla) across two seasons of variable dietary intake. Results show that gorilla gut microbiomes shared similar functional traits with each human group, depending on seasonal dietary behavior. Specifically, parallel microbiome traits were observed between hunter-gatherers and gorillas when the latter consumed more structural polysaccharides during dry seasons, while small-scale agriculturalist and gorilla microbiomes showed significant functional overlap when gorillas consumed more seasonal ripe fruit during wet seasons. Notably, dominance of microbial transporters, transduction systems, and gut xenobiotic metabolism was observed in association with traditional agriculture and energy-dense diets in gorillas at the expense of a functional microbiome repertoire capable of metabolizing more complex polysaccharides. Differential abundance of bacterial taxa that typically distinguish traditional from industrialized human populations (e.g., Prevotella spp.) was also recapitulated in the human and gorilla groups studied, possibly reflecting the degree of polysaccharide complexity included in each group's dietary niche. These results show conserved functional gut microbiome adaptations to analogous diets in small-scale human populations and nonhuman primates, highlighting the role of plant dietary polysaccharides and diverse environmental exposures in this convergence.IMPORTANCE The results of this study highlight parallel gut microbiome traits in human and nonhuman primates, depending on subsistence strategy. Although these similarities have been reported before, the functional and ecological bases of this convergence are not fully understood. Here, we show that this parallelism is, in part, likely modulated by the complexity of plant carbohydrates consumed and by exposures to diverse xenobiotics of natural and artificial origin. Furthermore, we discuss how divergence from these parallel microbiome traits is typically associated with adverse health outcomes in human populations living under culturally westernized subsistence patterns. This is important information as we trace the specific dietary and environmental triggers associated with the loss and gain of microbial functions as humans adapt to various dietary niches.

Gut Microbial Changes in Diabetic db/db Mice and Recovery of Microbial Diversity upon Pirfenidone Treatment.

The leptin receptor-deficient db/db mouse model is an accepted in vivo model to study obesity, type 2 diabetes, and diabetic kidney disease. Healthy gastrointestinal (GI) microbiota has been linked to weight loss, improved glycemic control, and physiological benefits. We investigated the effect of various drugs on the GI microbiota of db/db mice as compared to control db/m mice. Treatment with long-acting pirfenidone (PFD) increased gut microbial diversity in diabetic db/db mice. Firmicutes, the most abundant phylum in db/m mice, decreased significantly in abundance in db/db mice but showed increased abundance with long-acting PFD treatment. Several bacterial taxa, including Lactobacillus and some Bacteroides, were less abundant in db/db mice and more abundant in long-acting-PFD-treated db/db mice. Long-acting PFD treatment reduced the abundance of Akkermansia muciniphila (5%) as compared to db/db mice (~15%). We conclude that gut microbial dysbiosis observed in db/db mice was partially reversed by long-acting PFD treatment and hypothesize that PFD has beneficial effects, in part, via its influence on the gut microbial metabolite profile. In quantitatively assessing urine metabolites, we observed a high abundance of diabetic ketoacidosis biomarkers, including 3-hydroxybutyric acid and acetoacetic acid in db/db mice, which were less abundant in the long-acting-PFD-treated db/db mice.

Longitudinal Study of Oral Microbiome Variation in Twins.

Humans are host to a multitude of microorganisms that rapidly populate the body at birth, subject to a complex interplay that is dependent on host genetics, lifestyle, and environment. The host-associated microbiome, including the oral microbiome, presents itself in a complex ecosystem important to health and disease. As the most common chronic disease globally, dental caries is induced by host-microbial dysbiosis in children and adults. Multiple biological and environmental factors are likely to impact disease predisposition, onset, progression, and severity, yet longitudinal studies able to capture these influences are missing. To investigate how host genetics and environment influenced the oral microbial communities over time, we profiled supragingival plaque microbiomes of dizygotic and monozygotic twins during 3 visits over 12-months. Dental plaque DNA samples were amplified by targeting the 16S rRNA gene V4 region, and microbial findings were correlated with clinical, diet and genetic metadata. We observed that the oral microbiome variances were shaped primarily by the environment when compared to host genetics. Among the environmental factors shaping microbial changes of our subjects, significant metadata included age of the subject, and the age by which subjects initiated brushing habits, and the types of actions post-brushing. Relevant heritability of the microbiome included Actinomyces and Capnocytophaga in monozygotic twins and Kingella in dizygotic twins. Corynebacterium and Veillonella abundances were associated with age, whereas Aggregatibacter was associated with younger subjects. Streptococcus abundance showed an inverse association over time, and Selenomonas abundances increased with brushing frequency per day. Unraveling the exact biological mechanisms in caries has the potential to reveal novel host-microbial biomarkers, pathways, and targets important to effective preventive measures, and early disease control in children.

Influence of Substrate Concentration on the Culturability of Heterotrophic Soil Microbes Isolated by High-Throughput Dilution-to-Extinction Cultivation.

The vast majority of microbes inhabiting oligotrophic shallow subsurface soil environments have not been isolated or studied under controlled laboratory conditions. In part, the challenges associated with isolating shallow subsurface microbes may persist because microbes in deeper soils are adapted to low nutrient availability or quality. Here, we use high-throughput dilution-to-extinction culturing to isolate shallow subsurface microbes from a conifer forest in Arizona, USA. We hypothesized that the concentration of heterotrophic substrates in microbiological growth medium would affect which microbial taxa were culturable from these soils. To test this, we diluted cells extracted from soil into one of two custom-designed defined growth media that differed by 100-fold in the concentration of amino acids and organic carbon. Across the two media, we isolated a total of 133 pure cultures, all of which were classified as Actinobacteria or Alphaproteobacteria The substrate availability dictated which actinobacterial phylotypes were culturable but had no significant effect on the culturability of Alphaproteobacteria We isolated cultures that were representative of the most abundant phylotype in the soil microbial community (Bradyrhizobium spp.) and representatives of five of the top 10 most abundant Actinobacteria phylotypes, including Nocardioides spp., Mycobacterium spp., and several other phylogenetically divergent lineages. Flow cytometry of nucleic acid-stained cells showed that cultures isolated on low-substrate medium had significantly lower nucleic acid fluorescence than those isolated on high-substrate medium. These results show that dilution-to-extinction is an effective method to isolate abundant soil microbes and that the concentration of substrates in culture medium influences the culturability of specific microbial lineages.IMPORTANCE Isolating environmental microbes and studying their physiology under controlled conditions are essential aspects of understanding their ecology. Subsurface ecosystems are typically nutrient-poor environments that harbor diverse microbial communities-the majority of which are thus far uncultured. In this study, we use modified high-throughput cultivation methods to isolate subsurface soil microbes. We show that a component of whether a microbe is culturable from subsurface soils is the concentration of growth substrates in the culture medium. Our results offer new insight into technical approaches and growth medium design that can be used to access the uncultured diversity of soil microbes.

Intestinal and hepatic microbiota changes associated with chronic ethanol administration in mice.

Alcohol-induced liver disease is closely related to translocation of bacterial products and bacteria from the intestine to the liver. However, it is not known whether bacterial translocation to the liver depends on certain intestinal microbiota changes that would predispose bacteria to translocate to the liver. In this study, we investigated the microbiota in the jejunum, ileum, cecum, feces and liver of mice subjected to chronic ethanol feeding using a Lieber DeCarli diet model of chronic ethanol feeding for 8 weeks. We demonstrate that chronic ethanol administration changes alpha diversity in the ileum and the liver and leads to compositional changes especially in the ileum. This is largely driven by an increase in gram-negative phyla - the source of endotoxins. Moreover, gram-negative Prevotella not only increased in the mucus layer of the ileum but also in liver samples. These results suggest that bacterial translocation to the liver might be associated with microbiota changes in the distal gastrointestinal tract.

Intestinal Fungal Dysbiosis and Systemic Immune Response to Fungi in Patients With Alcoholic Hepatitis.

Chronic alcohol consumption causes increased intestinal permeability and changes in the intestinal microbiota composition, which contribute to the development and progression of alcohol-related liver disease. In this setting, little is known about commensal fungi in the gut. We studied the intestinal mycobiota in a cohort of patients with alcoholic hepatitis, patients with alcohol use disorder, and nonalcoholic controls using fungal-specific internal transcribed spacer amplicon sequencing of fecal samples. We further measured serum anti-Saccharomyces cerevisiae antibodies (ASCA) as a systemic immune response to fungal products or fungi. Candida was the most abundant genus in the fecal mycobiota of the two alcohol groups, whereas genus Penicillium dominated the mycobiome of nonalcoholic controls. We observed a lower diversity in the alcohol groups compared with controls. Antibiotic or steroid treatment was not associated with a lower diversity. Patients with alcoholic hepatitis had significantly higher ASCA levels compared to patients with alcohol use disorder and to nonalcoholic controls. Within the alcoholic hepatitis cohort, patients with levels of at least 34 IU/mL had a significantly lower 90-day survival (59%) compared with those with ASCA levels less than 34 IU/mL (80%) with an adjusted hazard ratio of 3.13 (95% CI, 1.11-8.82; P = 0.031). Conclusion: Patients with alcohol-associated liver disease have a lower fungal diversity with an overgrowth of Candida compared with controls. Higher serum ASCA was associated with increased mortality in patients with alcoholic hepatitis. Intestinal fungi may serve as a therapeutic target to improve survival, and ASCA may be useful to predict the outcome in patients with alcoholic hepatitis.

Persistent Gut Microbial Dysbiosis in Children with Acute Lymphoblastic Leukemia (ALL) During Chemotherapy.

Prophylactic or therapeutic antibiotic use along with chemotherapy treatment potentially has a long-standing adverse effect on the resident gut microbiota. We have established a case-control cohort of 32 pediatric and adolescent acute lymphoblastic leukemia (ALL) patients and 25 healthy siblings (sibling controls) to assess the effect of chemotherapy as well as antibiotic prophylaxis on the gut microbiota. We observe that the microbiota diversity and richness of the ALL group is significantly lower than that of the control group at diagnosis and during chemotherapy. The microbiota diversity is even lower in antibiotics-exposed ALL patients. Although the gut microbial diversity tends to stabilize after 1-year post-chemotherapy, their abundances were altered because of chemotherapy and prophylactic antibiotic treatments. Specifically, the abundances of mucolytic gram-positive anaerobic bacteria, including Ruminococcus gnavus and Ruminococcus torques, tended to increase during the chemotherapy regimen and continued to be elevated 1 year beyond the initiation of chemotherapy. This dysbiosis may contribute to the development of gastrointestinal complications in ALL children following chemotherapy. These findings set the stage to further understand the role of the gut microbiome dynamics in ALL patients and their potential role in alleviating some of the adverse side effects of chemotherapy and antibiotics use in immunocompromised children.

Bacteriophage targeting of gut bacterium attenuates alcoholic liver disease.

Chronic liver disease due to alcohol-use disorder contributes markedly to the global burden of disease and mortality1-3. Alcoholic hepatitis is a severe and life-threatening form of alcohol-associated liver disease. The gut microbiota promotes ethanol-induced liver disease in mice4, but little is known about the microbial factors that are responsible for this process. Here we identify cytolysin-a two-subunit exotoxin that is secreted by Enterococcus faecalis5,6-as a cause of hepatocyte death and liver injury. Compared with non-alcoholic individuals or patients with alcohol-use disorder, patients with alcoholic hepatitis have increased faecal numbers of E. faecalis. The presence of cytolysin-positive (cytolytic) E. faecalis correlated with the severity of liver disease and with mortality in patients with alcoholic hepatitis. Using humanized mice that were colonized with bacteria from the faeces of patients with alcoholic hepatitis, we investigated the therapeutic effects of bacteriophages that target cytolytic E. faecalis. We found that these bacteriophages decrease cytolysin in the liver and abolish ethanol-induced liver disease in humanized mice. Our findings link cytolytic E. faecalis with more severe clinical outcomes and increased mortality in patients with alcoholic hepatitis. We show that bacteriophages can specifically target cytolytic E. faecalis, which provides a method for precisely editing the intestinal microbiota. A clinical trial with a larger cohort is required to validate the relevance of our findings in humans, and to test whether this therapeutic approach is effective for patients with alcoholic hepatitis.

Gastro-intestinal and oral microbiome signatures associated with healthy aging.

The human oral and gut microbiomes influence health via competition for a distinct niche in the body with pathogens, via metabolic capabilities that increase host digestive capacity and generate compounds engaged in signaling pathways and modulation of immune system functions. Old age alters our metabolic and regenerative capacity. Following recruitment of 65 human subjects in the age range of 70 to 82, we discerned healthy aging (HA) and non-healthy aging (NHA) cohorts discordant in the occurrence of one or more major diseases: (1) cancer, (2) acute or chronic cardiovascular diseases, (3) acute or chronic pulmonary diseases, (4) diabetes, and (5) stroke or neurodegenerative disorders. We analyzed these cohorts' oral microbiomes (saliva) and gut microbiomes (stool) to assess diversity and identify microbial biomarkers for HA. In contrast to the gut microbiome where no change was observed, we found that the saliva microbiome had higher α-diversity in the HA compared with the NHA group. We observed the genus Akkermansia to be significantly more abundant in the gut microbiota of the HA group. Akkermansia muciniphila is a colonic mucin-degrading bacterium believed to have beneficial effects on gastrointestinal health, particularly in the context of diabetes and obesity. Erysipelotrichaceae UCG-003 was a taxon increased in abundance in the HA cohort. Streptococcus was the only genus observed to be significantly decreased in abundance in both the gut and oral microbiomes of the HA cohort compared with the NHA cohort. Our data support the notion that these microbes are potential probiotics to decrease the risks of non-healthy aging.