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The gut microbiota plays an important role in human health and disease. Stool, rectal swab and rectal mucosal tissue samples have been used in individual studies to survey the microbial community but the consequences of using these different sample types are not completely understood. In this study, we report differences in stool, rectal swab and rectal mucosal tissue microbial communities with shotgun metagenome sequencing of 1397 stool, swab and mucosal tissue samples from 240 participants. The taxonomic composition of stool and swab samples was distinct, but less different to each other than mucosal tissue samples. Functional profile differences between stool and swab samples are smaller, but mucosal tissue samples remained distinct from the other two types. When the taxonomic and functional profiles were used for inference in association with host phenotypes of age, sex, body mass index (BMI), antibiotics or non-steroidal anti-inflammatory drugs (NSAIDs) use, hypothesis testing using either stool or rectal swab gave broadly significantly correlated results, but inference performed on mucosal tissue samples gave results that were generally less consistent with either stool or swab. Our study represents an important resource for determination of how inference can change for taxa and pathways depending on the choice of where to sample within the human gut.

BACKGROUND: Bacterial exposure from house dust has been associated with asthma and atopy in children but whether these relationships are present in adults remains unclear. OBJECTIVE: To examine associations of house dust microbiota with adult asthma, atopy, and hay fever. METHODS: Vacuumed bedroom dust samples from the homes of 879 participants (average age 62) in the Agricultural Lung Health Study, a case-control study of asthma nested within a farming cohort, were subjected to 16S rRNA amplicon sequencing to characterize bacterial communities. We defined current asthma and hay fever using questionnaires and current atopy by blood specific immunoglobulin E > 0.70 IU/ml to ≥ 1 of ten common allergens. We used linear regression to examine whether overall within-sample bacterial diversity differed by outcome, microbiome regression-based kernel association test (MiRKAT) to evaluate whether between-sample bacterial community compositions differed by outcome, and analysis of composition of microbiomes (ANCOM) to identify differentially abundant bacterial taxa. RESULTS: Overall diversity of bacterial communities in house dust was similar by asthma status but was lower (p-value < 0.05) with atopy or hay fever. Many individual bacterial taxa were differentially abundant (false discovery rate < 0.05) by asthma, atopy, or hay fever. Several taxa from Cyanobacteria, Bacteroidetes, and Fusobacteria were more abundant with asthma, atopy, or hay fever. In contrast, several taxa from Firmicutes were more abundant in homes of individuals with adequately controlled asthma (vs inadequately controlled asthma), non-atopics, or individuals without hay fever. CONCLUSION: Microbial composition of house dust may influence allergic outcomes in adults.

Significance Cancer chemotherapy often causes side effects that require modulations in dosing, which then reduce anticancer efficacy. Here, we show that targeted inhibition of gut bacterial enzymes alleviates key stages of gut epithelial damage caused by the cancer drug irinotecan, blunts stark gut microbial compositional shifts caused by irinotecan, and enhances irinotecan’s antitumor effectiveness by reducing its gastrointestinal toxicity., Irinotecan treats a range of solid tumors, but its effectiveness is severely limited by gastrointestinal (GI) tract toxicity caused by gut bacterial β-glucuronidase (GUS) enzymes. Targeted bacterial GUS inhibitors have been shown to partially alleviate irinotecan-induced GI tract damage and resultant diarrhea in mice. Here, we unravel the mechanistic basis for GI protection by gut microbial GUS inhibitors using in vivo models. We use in vitro, in fimo, and in vivo models to determine whether GUS inhibition alters the anticancer efficacy of irinotecan. We demonstrate that a single dose of irinotecan increases GI bacterial GUS activity in 1 d and reduces intestinal epithelial cell proliferation in 5 d, both blocked by a single dose of a GUS inhibitor. In a tumor xenograft model, GUS inhibition prevents intestinal toxicity and maintains the antitumor efficacy of irinotecan. Remarkably, GUS inhibitor also effectively blocks the striking irinotecan-induced bloom of Enterobacteriaceae in immune-deficient mice. In a genetically engineered mouse model of cancer, GUS inhibition alleviates gut damage, improves survival, and does not alter gut microbial composition; however, by allowing dose intensification, it dramatically improves irinotecan’s effectiveness, reducing tumors to a fraction of that achieved by irinotecan alone, while simultaneously promoting epithelial regeneration. These results indicate that targeted gut microbial enzyme inhibitors can improve cancer chemotherapeutic outcomes by protecting the gut epithelium from microbial dysbiosis and proliferative crypt damage.

Objective Gut microorganisms contribute to the metabolism of environmental toxicants, including methylmercury (MeHg). Our main objective was to investigate whether associations between biomarkers for prenatal MeHg exposure and maternal gut microbiota differed between early and late gestation. Methods Maternal blood and stool samples were collected during early (8.3–17 weeks, n=28) and late (27–36 weeks, n=24) gestation. Total mercury and MeHg concentrations were quantified in biomarkers, and inorganic mercury was estimated by subtraction. The diversity and structure of the gut microbiota were investigated using 16S rRNA gene profiling (n = 52). Biomarkers were dichotomized, and diversity patterns were compared between high/low mercury concentrations. Spearman's correlation was used to assess bivariate associations between MeHg biomarkers (stool, blood, and meconium), and 23 gut microbial taxa (genus or family level, >1% average relative abundance). Results Within-person and between-person diversity patterns in gut microbiota differed between early/late gestation. The overall composition of the microbiome differed between high/low MeHg concentrations (in blood and stool) during early gestation, but not late gestation. Ten (of 23) taxa were significantly correlated with MeHg biomarkers (increasing or decreasing); however, associations differed, depending on whether the sample was collected during early or late gestation. A total of 43% of associations (69/161) reversed the direction of correlation between early/late gestation. Conclusions The time point at which a maternal fecal sample is collected may yield different associations between gut microorganisms and MeHg biomarkers, which may be due in part to remodeling of maternal microbiota during pregnancy. Our results suggest the effectiveness of dietary interventions to reduce prenatal MeHg exposure may differ between early and late gestation.

Background: Measuring and understanding the function of the human microbiome is key for several aspects of health; however, the development of statistical methods specifically for the analysis of microbial gene expression (i.e., metatranscriptomics) is in its infancy. Many currently employed differential expression analysis methods have been designed for different data types and have not been evaluated in metatranscriptomics settings. To address this knowledge gap, we undertook a comprehensive evaluation and benchmarking of eight differential analysis methods for metatranscriptomics data. Results: We used a combination of real and simulated metatranscriptomics data to evaluate the performance (i.e., model fit, Type-I error, and statistical power) of eights methods: log-normal (LN), logistic-beta (LB), MAST, Kruskal-Wallis, two-part Kruskal-Wallis, DESeq2, and ANCOM-BC and metagenomeSeq. The simulation was informed by supragingival biofilm microbiome data from about 300 preschool-age children enrolled in a study of early childhood caries (ECC), whereas validations were sought in two additional datasets, including an ECC and an inflammatory bowel disease one. The LB test showed the highest power in both small and large sample sizes and reasonably controlled Type-I error. Contrarily, MAST was hampered by inflated Type-I error. Using LN and LB tests, we found that genes C8PHV7 and C8PEV7, harbored by the lactate-producing Campylobacter gracilis, had the strongest association with ECC. Conclusion: This comprehensive model evaluation findings offer practical guidance for the selection of appropriate methods for rigorous analyses of differential expression in metatranscriptomics data. Selection of an optimal method is likely to increase the possibility of detecting true signals while minimizing the chance of claiming false ones.

OBJECTIVES: The objective of this study was to determine if perturbations in gut microbial composition and the gut metabolome could be linked to individuals with obesity and osteoarthritis (OA). METHODS: Fecal samples were collected from 92 participants with obesity recruited from the Johnston County Osteoarthritis Project. OA cases (n = 59) had radiographic hand plus knee OA, defined as involvement of at least 3 joints across both hands, and a Kellgren-Lawrence (KL) grade 2–4 in at least one knee. Controls (n = 33) were without hand OA and with KL grade 0–1 knees. Fecal metabolomes were analyzed by a UHPLC/Q Exactive HFx mass spectrometer. Microbiome composition was determined in fecal samples by 16S ribosomal RNA amplicon sequencing (rRNA-seq). Stepwise logistic regression models were built to determine predictors of OA status. Spearman correlations were performed to determine associations between metabolites and microbiota in OA or healthy individuals. RESULTS: Untargeted metabolomics analysis indicated that OA cases had significantly higher levels of di- and tri-peptides (P

Experimental manipulation of gut microbes in animal models alters fear behavior and relevant neurocircuitry. In humans, the first year of life is a key period for brain development, the emergence of fearfulness, and the establishment of the gut microbiome. Variation in the infant gut microbiome has previously been linked to cognitive development, but its relationship with fear behavior and neurocircuitry is unknown. In this pilot study of 34 infants, we find that 1-year gut microbiome composition (Weighted Unifrac; lower abundance of Bacteroides, increased abundance of Veillonella, Dialister, and Clostridiales) is significantly associated with increased fear behavior during a non-social fear paradigm. Infants with increased richness and reduced evenness of the 1-month microbiome also display increased non-social fear. This study indicates associations of the human infant gut microbiome with fear behavior and possible relationships with fear-related brain structures on the basis of a small cohort. As such, it represents an important step in understanding the role of the gut microbiome in the development of human fear behaviors, but requires further validation with a larger number of participants., Experimental manipulation of the gut microbiome in animal models impacts fear behaviours. Here, the authors show in a pilot study that features of the human infant gut microbiome are associated with non-social fear behaviours during a laboratory based assessment.

Complex dietary carbohydrate structures including β(1–4) galacto-oligosaccharides (GOS) are resistant to digestion in the upper gastrointestinal (GI) tract and arrive intact to the colon where they benefit the host by selectively stimulating microbial growth. Studies have reported the beneficial impact of GOS (alone or in combination with other prebiotics) by serving as metabolic substrates for modulating the assembly of the infant gut microbiome while reducing GI infections. N-Acetyl-D-lactosamine (LacNAc, Galβ1,4GlcNAc) is found in breast milk as a free disaccharide. This compound is also found as a component of human milk oligosaccharides (HMOs), which have repeating and variably branched lactose and/or LacNAc units, often attached to sialic acid and fucose monosaccharides. Human glycosyl-hydrolases do not degrade most HMOs, indicating that these structures have evolved as natural prebiotics to drive the proper assembly of the infant healthy gut microbiota. Here, we sought to develop a novel enzymatic method for generating LacNAc-enriched GOS, which we refer to as humanized GOS (hGOS). We showed that the membrane-bound β-hexosyl transferase (rBHT) from Hamamotoa (Sporobolomyces) singularis was able to generate GOS and hGOS from lactose and N-Acetyl-glucosamine (GlcNAc). The enzyme catalyzed the regio-selective, repeated addition of galactose from lactose to GlcNAc forming the β-galactosyl linkage at the 4-position of the GlcNAc and at the 1-position of D-galactose generating, in addition to GOS, LacNAc, and Galactosyl-LacNAc trisaccharides which were produced by two sequential transgalactosylations. Humanized GOS is chemically distinct from HMOs, and its effects in vivo have yet to be determined. Thus, we evaluated its safety and demonstrated the prebiotic's ability to modulate the gut microbiome in 6-week-old C57BL/6J mice. Longitudinal analysis of gut microbiome composition of stool samples collected from mice fed a diet containing hGOS for 5 weeks showed a transient reduction in alpha diversity. Differences in microbiome community composition mostly within the Firmicutes phylum were observed between hGOS and GOS, compared to control-fed animals. In sum, our study demonstrated the biological synthesis of hGOS, and signaled its safety and ability to modulate the gut microbiome in vivo, promoting the growth of beneficial microorganisms, including Bifidobacterium and Akkermansia.

The protozoal pathogen Tritrichomonas foetus infects the colon of domestic cats and is a major cause of chronic colitis and diarrhea. Treatment failure is common, but antibiotics may improve clinical signs in a subset of cats, leading researchers to question involvement of the colonic microbiota in disease pathogenesis. Studies performed in women with venereal Trichomonas vaginalis infections have revealed that dysbiosis of host microbiota contributes to pathogenicity with similar findings also found in mice with intestinal Tritrichomonas musculis The aim of this study was to characterize differences in the fecal microbiota of cats with and without naturally occurring T. foetus infection and in a group of kittens prior to and after experimentally induced infection. Archived fecal DNA from cats undergoing testing for T. foetus infection (n = 89) and experimentally infected kittens (n = 4; at pre-, 2 weeks, and 9 weeks post-infection) were analyzed by sequencing of 16S rRNA genes. Amongst the naturally infected population, the genera Megamonas and Helicobacter were significantly increased in prevalence and abundance in cats testing positive for T. foetus infection. In the group of four experimentally infected kittens, fecal samples post-infection had significantly lower abundance of genus Dialister and Megamonas and greater abundance of the class Betaproteobacteria and family Succinivibrionaceae. We hypothesize that T. foetus promotes dysbiosis by competition for fermentable substrates used by these bacteria and that metabolic byproducts may contribute to the pathogenesis of colonic inflammation and diarrhea. Future studies are warranted for the measurement of fecal concentrations of microbial and protozoal metabolites in cats with T. foetus infection for the identification of potential therapeutic targets.

Background Prebiotic galacto-oligosaccharides (GOS) have an extensively demonstrated beneficial impact on intestinal health. In this study, we determined the impact of GOS diets on hallmarks of gut aging: microbiome dysbiosis, inflammation, and intestinal barrier defects (“leaky gut”). We also evaluated if short-term GOS feeding influenced how the aging gut responded to antibiotic challenges in a mouse model of Clostridioides difficile infection. Finally, we assessed if colonic organoids could reproduce the GOS responder—non-responder phenotypes observed in vivo. Results Old animals had a distinct microbiome characterized by increased ratios of non-saccharolytic versus saccharolytic bacteria and, correspondingly, a lower abundance of β-galactosidases compared to young animals. GOS reduced the overall diversity, increased the abundance of specific saccharolytic bacteria (species of Bacteroides and Lactobacillus), increased the abundance of β-galactosidases in young and old animals, and increased the non-saccharolytic organisms; however, a robust, homogeneous bifidogenic effect was not observed. GOS reduced age-associated increased intestinal permeability and increased MUC2 expression and mucus thickness in old mice. Clyndamicin reduced the abundance Bifidobacterium while increasing Akkermansia, Clostridium, Coprococcus, Bacillus, Bacteroides, and Ruminococcus in old mice. The antibiotics were more impactful than GOS on modulating serum markers of inflammation. Higher serum levels of IL-17 and IL-6 were observed in control and GOS diets in the antibiotic groups, and within those groups, levels of IL-6 were higher in the GOS groups, regardless of age, and higher in the old compared to young animals in the control diet groups. RTqPCR revealed significantly increased gene expression of TNFα in distal colon tissue of old mice, which was decreased by the GOS diet. Colon transcriptomics analysis of mice fed GOS showed increased expression of genes involved in small-molecule metabolic processes and specifically the respirasome in old animals, which could indicate an increased oxidative metabolism and energetic efficiency. In young mice, GOS induced the expression of binding-related genes. The galectin gene Lgals1, a β-galactosyl-binding lectin that bridges molecules by their sugar moieties and is an important modulator of the immune response, and the PI3K-Akt and ECM-receptor interaction pathways were also induced in young mice. Stools from mice exhibiting variable bifidogenic response to GOS injected into colon organoids in the presence of prebiotics reproduced the response and non-response phenotypes observed in vivo suggesting that the composition and functionality of the microbiota are the main contributors to the phenotype. Conclusions Dietary GOS modulated homeostasis of the aging gut by promoting changes in microbiome composition and host gene expression, which was translated into decreased intestinal permeability and increased mucus production. Age was a determining factor on how prebiotics impacted the microbiome and expression of intestinal epithelial cells, especially apparent from the induction of galectin-1 in young but not old mice.

Aim: This in vivo experimental study investigated bacterial microbiome and metabolome longitudinal changes associated with enamel caries lesion progression and arrest. Methods: We induced natural caries activity in three caries-free volunteers prior to four premolar extractions for orthodontic reasons. The experimental model included placement of a modified orthodontic band on smooth surfaces and a mesh on occlusal surfaces. We applied the caries-inducing protocol for 4- and 6-weeks, and subsequently promoted caries lesion arrest via a 2-week toothbrushing period. Lesions were verified clinically and quantitated via micro-CT enamel density measurements. The biofilm microbial composition was determined via 16S rRNA gene Illumina sequencing and NMR spectrometry was used for metabolomics. Results: Biofilm maturation and caries lesion progression were characterized by an increase in Gram-negative anaerobes, including Veillonella and Prevotella. Streptococcus was associated caries lesion progression, while a more equal distribution of Streptococcus, Bifidobacterium, Atopobium, Prevotella, Veillonella, and Saccharibacteria (TM7) characterized arrest. Lactate, acetate, pyruvate, alanine, valine, and sugars were more abundant in mature biofilms compared to newly formed biofilms. Conclusions: These longitudinal bacterial microbiome and metabolome results provide novel mechanistic insights into the role of the biofilm in caries progression and arrest and offer promising candidate biomarkers for validation in future studies.

Background Early life exposure to adverse environments, and maternal stress in particular, has been shown to increase risk for metabolic diseases and neurobehavioral disorders. While many studies have examined the hypothalamic-pituitary-adrenal axis (HPA axis) as the primary mechanism behind these relationships, emerging research on the brain-gut axis suggests that the microbiome may play a role. In this study, we tested the relationships among maternal precarity and HPA axis dysregulation during the peripartum period, infant gut microbiome composition, and infant HPA axis functioning. Methods Data come from 25 mother-infant dyads in the Galápagos, Ecuador. Women completed surveys on precarity measures (food insecurity, low social support, depression, and stress) and gave salivary cortisol samples during and after pregnancy. Infant salivary cortisol and stool were collected in the postpartum. Statistical significance of differences in microbial diversity and relative abundance were assessed with respect to adjusted linear regression models. Results Maternal precarity was associated with lower diversity and higher relative abundance of Enterobacteriaceae and Streptococcaceae and a lower relative abundance of Bifidobacterium and Lachnospiraceae. These patterns of colonization for Enterobacteriaceae and Bifidobacterium mirrored those found in infants with HPA axis dysregulation. Maternal HPA axis dysregulation during pregnancy was also associated with a greater relative abundance of Veillonella. Conclusions Overall, exposures to precarity and HPA axis dysregulation were associated with an increase in groups that include potentially pathogenic bacteria, including Enterobacteriaceae, Streptococcaceae, and Veillonella, and a decrease in potentially protective bacteria, including Bifidobacterium and Lachnospiraceae, as well as a decrease in overall diversity.

Recent and ongoing developments in microbiome science are enabling new frontiers of research for probiotics and prebiotics. Novel types, mechanisms, and applications currently under study have the potential to change scientific understanding as well as nutritional and healthcare applications of these interventions. The expansion of related fields of microbiome-targeted interventions, and an evolving landscape for implementation across regulatory, policy, prescriber, and consumer spheres, portends an era of significant change. In this review we examine recent, emerging, and anticipated trends in probiotic and prebiotic science, and create a vision for broad areas of developing influence in the field.

Recently, there has been a surge of interest in the possibility that microbial communities inhabiting the human gut could affect cognitive development and increase risk for mental illness via the “microbiome-gut-brain axis”. Infancy likely represents a critical period for the establishment of these relationships, as it is the most dynamic stage of postnatal brain development and a key period in the maturation of the microbiome. Indeed, recent reports indicate that characteristics of the infant gut microbiome are associated with both temperament and cognitive performance. The neural circuits underlying these relationships have not yet been delineated. To address this gap, resting-state fMRI scans were acquired from 39 one-year-old human infants who had provided fecal samples for identification and relative quantification of bacterial taxa. Measures of alpha diversity were generated and tested for associations with measures of functional connectivity. Primary analyses focused on the amygdala as manipulation of the gut microbiota in animal models alters the structure and neurochemistry of this brain region. Secondary analyses explored functional connectivity of nine canonical resting state functional networks. Alpha diversity was significantly associated with functional connectivity between the amygdala and thalamus and between the anterior cingulate cortex and anterior insula. These regions play an important role in processing/responding to threat. Alpha diversity was also associated with functional connectivity between the supplementary motor area (SMA, representing the sensorimotor network) and the inferior parietal lobule (IPL). Importantly, SMA-IPL connectivity also related to cognitive outcomes at 2 years of age, suggesting a potential pathway linking gut microbiome diversity and cognitive outcomes during infancy. These results provide exciting new insights into the gut-brain axis during early human development and should stimulate further studies into whether microbiome–associated changes in brain circuitry influence later risk for psychopathology.

Although colorectal cancer is the third leading cause of death in Morocco, there are no studies of the microbiome changes associated with the disease in the Moroccan population. The aim of our study was to compare the stool microbiome of Moroccan cancer patients with healthy individuals. We analyzed the microbiome composition of samples from 11 CRC patients and 12 healthy individuals by 16S rRNA amplicon sequencing. Principal coordinate analysis of samples revealed defined cancer versus healthy clusters. Our findings showed that cancer samples had higher proportions of Firmicutes (T = 50.5%; N = 28.4%; p = 0.04), specifically of Clostridia (T = 48.3%; N = 19.0%; p = 0.002), and Fusobacteria (T = 0.1%; N = 0.0%; p = 0.02), especially of Fusobacteriia (T = 0.1%; N = 0.0%; p = 0.02), while Bacteroidetes were enriched in healthy samples (T = 35.1%; N = 62.8%; p = 0.06), particularly the class Bacteroidia (T = 35.1%; N = 62.6%; p = 0.06). Porphyromonas, Clostridium, Ruminococcus, Selenomonas, and Fusobacterium were significantly overrepresented in diseased patients, similarly to other studies. Predicted functional information showed that bacterial motility proteins, flagellar assembly, and fatty acid biosynthesis metabolism were significantly overrepresented in cancer patients, while amino acid metabolism and glycan biosynthesis were overrepresented in controls. This suggests that involvement of these functional metagenomes is similar and relevant in the carcinogenesis process, independent of the origin of the samples. Results from this study allowed identification of bacterial taxa relevant to the Moroccan population and encourages larger studies to facilitate population-directed therapeutic approaches. Electronic supplementary material The online version of this article (10.1007/s00430-018-0542-5) contains supplementary material, which is available to authorized users.

Objectives Inadequate sleep increases the risk for age-related cognitive decline and recent work suggests a possible role of the gut microbiota in this phenomenon. Partial sleep deprivation alters the human gut microbiome, and its composition is associated with cognitive flexibility in animal models. Given these findings, we examined the possible relationship among the gut microbiome, sleep quality, and cognitive flexibility in a sample of healthy older adults. Methods Thirty-seven participants (age 64.59 ± 7.54 years) provided a stool sample for gut microbial sequencing and completed the Pittsburgh Sleep Quality Index and Stroop Color Word Test as part of a larger project. Results Better sleep quality was associated with better Stroop performance and higher proportions of the gut microbial phyla Verrucomicrobia and Lentisphaerae. Stroop Word and Color-Word performance correlated with higher proportions of Verrucomicrobia and Lentisphaerae. Partial correlations suggested that the relationship between Lentisphaerae and Stroop Color-Word performance was better accounted for by sleep quality; sleep quality remained a significant predictor of Color-Word performance, independent of the Lentisphaerae proportion, while the relationship between Lentisphaerae and Stroop performance was non-significant. Verrucomicrobia and sleep quality were not associated with Stroop Word performance independent of one another. Conclusions The current findings suggest a possible relationship among sleep quality, composition of the gut microbiome, and cognitive flexibility in healthy older adults. Prospective and experimental studies are needed to confirm these findings and determine whether improving microbiome health may buffer against sleep-related cognitive decline in older adults.

Early childhood caries (ECC) is a biofilm-mediated disease. Social, environmental, and behavioral determinants as well as innate susceptibility are major influences on its incidence; however, from a pathogenetic standpoint, the disease is defined and driven by oral dysbiosis. In other words, the disease occurs when the natural equilibrium between the host and its oral microbiome shifts toward states that promote demineralization at the biofilm-tooth surface interface. Thus, a comprehensive understanding of dental caries as a disease requires the characterization of both the composition and the function or metabolic activity of the supragingival biofilm according to well-defined clinical statuses. However, taxonomic and functional information of the supragingival biofilm is rarely available in clinical cohorts, and its collection presents unique challenges among very young children. This paper presents a protocol and pipelines available for the conduct of supragingival biofilm microbiome studies among children in the primary dentition, that has been designed in the context of a large-scale population-based genetic epidemiologic study of ECC. The protocol is being developed for the collection of two supragingival biofilm samples from the maxillary primary dentition, enabling downstream taxonomic (e.g., metagenomics) and functional (e.g., transcriptomics and metabolomics) analyses. The protocol is being implemented in the assembly of a pediatric precision medicine cohort comprising over 6000 participants to date, contributing social, environmental, behavioral, clinical, and biological data informing ECC and other oral health outcomes.

The gut microbiota plays a critical role in the overall health of its host. Benefits derived from bacterial members of the gut microbiota can influence host growth, immune response, pathogen colonization, and intestinal physiology. Use of probiotics, prebiotics, and synbiotics are emerging as effective mechanisms to selectively modulate composition and function of the gut microbiota. This chapter introduces the concept of probiotics and prebiotics from a historic perspective, and attempts to answer the fundamental questions of the impact of probiotics and prebiotics on microbiome composition in health versus disease states, colonization of the human gut by probiotics (is it necessary?), and how the food or product matrix impact probiotic delivery and effect. The conclusion of this chapter focuses on the next generation of probiotics: novel species and bacterial consortia.

Our gut microbiota is composed of an assortment of (semi)permanent intestinal inhabitants that co-exist with temporary microbes. Although it is hard to keep up with the fast advancing field of microbiome research, this book bid to provide, first, a look into key players of the gut microbiota found in food from breast milk to dairy, meats and vegetables, to a revision of methods to generate efficacious probiotics. Then, we approached the oral microbiota as the host-microbe ecosystem that selects which bacteria will progress into the gastrointestinal tract, highlighting the importance of oral health for the overall health of the host. Finally, the third part of the book focused on age and the gut microbiome, how the consumption of foods with high microbial loads may lead to temporary changes in microbial diversity, to end on what is commonly known now as “the diseases of the Western civilization” and the potential to modulate the gut microbiota for a more resilient microbial population. Hence, what do we know now that we did not know 10 years ago?

Previous studies on the differences in gut microbiota between exclusively breastfed (EBF) and non-EBF infants have provided highly variable results. Here we perform a meta-analysis of seven microbiome studies (1825 stool samples from 684 infants) to compare the gut microbiota of non-EBF and EBF infants across populations. In the first 6 months of life, gut bacterial diversity, microbiota age, relative abundances of Bacteroidetes and Firmicutes, and predicted microbial pathways related to carbohydrate metabolism are consistently higher in non-EBF than in EBF infants, whereas relative abundances of pathways related to lipid metabolism, vitamin metabolism, and detoxification are lower. Variation in predicted microbial pathways associated with non-EBF infants is larger among infants born by Caesarian section than among those vaginally delivered. Longer duration of exclusive breastfeeding is associated with reduced diarrhea-related gut microbiota dysbiosis. Furthermore, differences in gut microbiota between EBF and non-EBF infants persist after 6 months of age. Our findings elucidate some mechanisms of short and long-term benefits of exclusive breastfeeding across different populations., Studies on the effects of breastfeeding on the infant gut microbiota have provided inconsistent results. Here, Ho et al. perform a meta-analysis of seven studies across different populations, supporting that exclusive breastfeeding is associated with short-term and long-term alterations in the infant gut microbiota.

Salmonella is estimated to cause one million foodborne illnesses in the United States every year. Salmonella -contaminated poultry products are one of the major sources of salmonellosis. Given the critical role of the gut microbiota in Salmonella transmission, a manipulation of the chicken intestinal microenvironment could prevent animal colonization by the pathogen. In Salmonella , the global regulator gene fnr ( f umarate n itrate r eduction) regulates anaerobic metabolism and is essential for adapting to the gut environment. This study tested the hypothesis that an attenuated Fnr mutant of Salmonella enterica serovar Typhimurium (attST) or prebiotic galacto-oligosaccharides (GOS) could improve resistance to wild-type Salmonella via modifications to the structure of the chicken gut microbiome. Intestinal samples from a total of 273 animals were collected weekly for 9 weeks to evaluate the impact of attST or prebiotic supplementation on microbial species of the cecum, duodenum, jejunum, and ileum. We next analyzed changes to the gut microbiome induced by challenging the animals with a wild-type Salmonella serovar 4,[5],12:r:− (Nal r ) strain and determined the clearance rate of the virulent strain in the treated and control groups. Both GOS and the attenuated Salmonella strain modified the gut microbiome but elicited alterations of different taxonomic groups. The attST produced significant increases of Alistipes and undefined Lactobacillus , while GOS increased Christensenellaceae and Lactobacillus reuteri . The microbiome structural changes induced by both treatments resulted in a faster clearance after a Salmonella challenge. IMPORTANCE With an average annual incidence of 13.1 cases/100,000 individuals, salmonellosis has been deemed a nationally notifiable condition in the United States by the Centers for Disease Control and Prevention (CDC). Earlier studies demonstrated that Salmonella is transmitted by a subset of animals (supershedders). The supershedder phenotype can be induced by antibiotics, ascertaining an essential role for the gut microbiota in Salmonella transmission. Consequently, modulation of the gut microbiota and modification of the intestinal microenvironment could assist in preventing animal colonization by the pathogen. Our study demonstrated that a manipulation of the chicken gut microbiota by the administration of an attenuated Salmonella strain or prebiotic galacto-oligosaccharides (GOS) can promote resistance to Salmonella colonization via increases of beneficial microorganisms that translate into a less hospitable gut microenvironment.

Large-scale microbiome studies have established that most of the diversity contained in the gastrointestinal tract is represented at the strain level; however, exhaustive genomic and physiological characterization of human isolates is still lacking. With increased use of probiotics as interventions for gastrointestinal disorders, genomic and functional characterization of novel microorganisms becomes essential. In this study, we explored the impact of strain-level genomic variability on bacterial physiology of two novel human Lactobacillus rhamnosus strains (AMC143 and AMC010) of probiotic potential in relation to stress resistance. The strains showed differences with known probiotic strains (L. rhamnosus GG, Lc705, and HN001) at the genomic level, including nucleotide polymorphisms, mutations in non-coding regulatory regions, and rearrangements of genomic architecture. Transcriptomics analysis revealed that gene expression profiles differed between strains when exposed to simulated gastrointestinal stresses, suggesting the presence of unique regulatory systems in each strain. In vitro physiological assays to test resistance to conditions mimicking the gut environment (acid, alkali, and bile stress) showed that growth of L. rhamnosus AMC143 was inhibited upon exposure to alkaline pH, while AMC010 and control strain LGG were unaffected. AMC143 also showed a significant survival advantage compared to the other strains upon bile exposure. Reverse transcription qPCR targeting the bile salt hydrolase gene (bsh) revealed that AMC143 expressed bsh poorly (a consequence of a deletion in the bsh promoter and truncation of bsh gene in AMC143), while AMC010 had significantly higher expression levels than AMC143 or LGG. Insertional inactivation of the bsh gene in AMC010 suggested that bsh could be detrimental to bacterial survival during bile stress. Together, these findings show that coupling of classical microbiology with functional genomics methods for the characterization of bacterial strains is critical for the development of novel probiotics, as variability between strains can dramatically alter bacterial physiology and functionality.

Colon cancer is a multifactorial disease associated with a variety of lifestyle factors. Alterations in the gut microbiota and the intestinal metabolome are noted during colon carcinogenesis, implicating them as critical contributors or results of the disease process. Diet is a known determinant of health, and as a modifier of the gut microbiota and its metabolism, a critical element in maintenance of intestinal health. This review summarizes recent evidence demonstrating the role and responses of the intestinal microbiota during colon tumorigenesis and the ability of dietary bioactive compounds and probiotics to impact colon health from the intestinal lumen to the epithelium and systemically. We first describe changes to the intestinal microbiome, metabolome, and epithelium associated with colon carcinogenesis. This is followed by a discussion of recent evidence indicating how specific classes of dietary bioactives, prebiotics, or probiotics affect colon carcinogenesis. Lastly, we briefly address the prospects of using multiple 'omics' techniques to integrate the effects of diet, host, and microbiota on colon tumorigenesis with the goal of more fully appreciating the interconnectedness of these systems and thus, how these approaches can be used to advance personalized nutrition strategies and nutrition research.

Directed modulation of the colonic bacteria to metabolize lactose effectively is a potentially useful approach to improve lactose digestion and tolerance. A randomized, double-blind, multisite placebo-controlled trial conducted in human subjects demonstrated that administration of a highly purified (>95%) short-chain galactooligosaccharide (GOS), designated “RP-G28,” significantly improved clinical outcomes for lactose digestion and tolerance. In these individuals, stool samples were collected pretreatment (day 0), after GOS treatment (day 36), and 30 d after GOS feeding stopped and consumption of dairy products was encouraged (day 66). In this study, changes in the fecal microbiome were investigated using 16S rRNA amplicon pyrosequencing and high-throughput quantitative PCR. At day 36, bifidobacterial populations were increased in 27 of 30 of GOS subjects (90%), demonstrating a bifidogenic response in vivo. Relative abundance of lactose-fermenting Bifidobacterium, Faecalibacterium, and Lactobacillus were significantly increased in response to GOS. When dairy was introduced into the diet, lactose-fermenting Roseburia species increased from day 36 to day 66. The results indicated a definitive change in the fecal microbiome of lactose-intolerant individuals, increasing the abundance of lactose-metabolizing bacteria that were responsive to dietary adaptation to GOS. This change correlated with clinical outcomes of improved lactose tolerance.

Objectives: Dysbiosis of the gut microbiome is implicated in numerous human health conditions. Animal studies have linked microbiome disruption to changes in cognitive functioning, although no study has examined this possibility in neurologically healthy older adults. Methods: Participants were 43 community-dwelling older adults (50–85 years) that completed a brief cognitive test battery and provided stool samples for gut microbiome sequencing. Participants performing≥1 SD below normative performance on two or more tests were compared to persons with one or fewer impaired scores. Results: Mann Whitney U tests revealed different distributions of Bacteroidetes (p=.01), Firmicutes (p=.02), Proteobacteria (p=.04), and Verrucomicrobia (p=.003) between Intact and Impaired groups. These phyla were significantly correlated with cognitive test performances, particularly Verrucomicrobia and attention/executive function measures. Conclusions: The current findings suggest that composition of the gut microbiome is associated with cognitive test performance in neurologically healthy older adults. Future studies are needed to confirm these findings and explore possible mechanisms. (JINS, 2017, 23, 700–705)

Investigation of bacteria involved in pathogenesis of necrotizing enterocolitis (NEC) is limited by infant fragility, analysis restricted to feces, use of culture-based methods, and lack of clinically-relevant animal models. This study used a unique preterm piglet model to characterize spontaneous differences in microbiome composition of NEC-predisposed regions of gut. Preterm piglets (n=23) were cesarean-delivered and nurtured for 30 hours over which time 52% developed NEC. Bacterial DNA from ileal content, ileal mucosa, and colonic mucosa were PCR amplified, subjected to terminal restriction fragment length polymorphism (TRFLP) analysis and targeted 16S rDNA qPCR. Preterm ileal mucosa was specifically bereft in diversity of bacteria compared to ileal content and colonic mucosa. Preterm ileum was restricted to representation by only Proteobacteria, Firmicutes, Cyanobacteria and Chloroflexi. In piglets with NEC, ileal mucosa was uniquely characterized by increases in number of Firmicutes and diversity of phyla to include Actinobacteria and uncultured bacteria. Five specific TRFLP profiles, corresponding in closest identity to Clostridium butyricum, C. neonatale, C. proteolyticum, Streptomyces spp., and Leptolyngbya spp., were significantly more prevalent or observed only among samples from piglets with NEC. Total numbers of Clostridium spp. and C. butyricum were significantly greater in samples of NEC ileal mucosa but not ileal content or colonic mucosa. These results provide strong support for ileal mucosa as a focus for investigation of specific dysbiosis associated with NEC and suggest a significant role for Clostridium spp., and members of the Actinobacteria and Cyanobacteria in the pathogenesis of NEC in preterm piglets.

Understanding the importance of the gut microbiome on modulation of host health has become a subject of great interest for researchers across disciplines. As an intrinsically multidisciplinary field, microbiome research has been able to reap the benefits of technological advancements in systems and synthetic biology, biomaterials engineering, and traditional microbiology. Gut microbiome research has been revolutionized by high-throughput sequencing technology, permitting compositional and functional analyses that were previously an unrealistic undertaking. Emerging technologies including engineered organoids derived from human stem cells, high-throughput culturing, and microfluidics assays allowing for the introduction of novel approaches will improve the efficiency and quality of microbiome research. Here, we will discuss emerging technologies and their potential impact on gut microbiome studies.

Ileocecal resection (ICR) is a commonly required surgical intervention in unmanageable Crohn's disease and necrotizing enterocolitis. However, the impact of ICR, and the concomitant doses of antibiotic routinely given with ICR, on the intestinal commensal microbiota has not been determined. In this study, wild-type C57BL6 mice were subjected to ICR and concomitant single intraperitoneal antibiotic injection. Intestinal lumen contents were collected from jejunum and colon at 7, 14, and 28 days after resection and compared to non-ICR controls. Samples were analyzed by16S rRNA gene pyrosequencing and quantitative PCR. The intestinal microbiota was altered by 7 days after ICR and accompanying antibiotic treatment, with decreased diversity in the colon. Phylogenetic diversity (PD) decreased from 11.8 ± 1.8 in non-ICR controls to 5.9 ± 0.5 in 7-day post-ICR samples. There were also minor effects in the jejunum where PD values decreased from 8.3 ± 0.4 to 7.5 ± 1.4. PCoA analysis indicated that bacterial populations 28 days post-ICR differed significantly from non-ICR controls. Moreover, colon and jejunum bacterial populations were remarkably similar 28 days after resection, whereas the initial communities differed markedly. Firmicutes and Bacteroidetes were the predominant phyla in jejunum and colon before ICR; however, Firmicutes became the vastly predominant phylum in jejunum and colon 28 days after ICR. Although the microbiota returned towards a homeostatic state, with re-establishment of Firmicutes as the predominant phylum, we did not detect Bacteroidetes in the colon 28 days after ICR. In the jejunum Bacteroidetes was detected at a 0.01% abundance after this time period. The changes in jejunal and colonic microbiota induced by ICR and concomitant antibiotic injection may therefore be considered as potential regulators of post-surgical adaptive growth or function, and in a setting of active IBD, potential contributors to post-surgical pathophysiology of disease recurrence. © 2013 Devine et al.

Astronaut intestinal health may be impacted by microgravity, radiation, and diet. The aim of this study was to characterize how high and low linear energy transfer (LET) radiation, microgravity, and elevated dietary iron affect colon microbiota (determined by 16S rDNA pyrosequencing) and colon function. Three independent experiments were conducted to achieve these goals: 1) fractionated low LET γ radiation (137Cs, 3 Gy, RAD), high Fe diet (IRON) (650 mg/kg diet), and a combination of low LET γ radiation and high Fe diet (IRON+RAD) in male Sprague-Dawley rats; 2) high LET 38Si particle exposure (0.050 Gy), 1/6 G partial weight bearing (PWB), and a combination of high LET38Si particle exposure and PWB in female BalbC/ByJ mice; and 3) 13 d spaceflight in female C57BL/6 mice. Low LET radiation, IRON and spaceflight increased Bacteroidetes and decreased Firmicutes. RAD and IRON+RAD increased Lactobacillales and lowered Clostridiales compared to the control (CON) and IRON treatments. Low LET radiation, IRON, and spaceflight did not significantly affect diversity or richness, or elevate pathogenic genera. Spaceflight increased Clostridiales and decreased Lactobacillales, and similar trends were observed in the experiment using a ground-based model of microgravity, suggesting altered gravity may affect colonic microbiota. Although we noted no differences in colon epithelial injury or inflammation, spaceflight elevated TGFβ gene expression. Microbiota and mucosal characterization in these models is a first step in understanding the impact of the space environment on intestinal health.