Qualitative and Quantitative DNA- and RNA-Based Analysis of the Bacterial Stomach Microbiota in Humans, Mice, and Gerbils.

Clinical interventions in the stomach have been linked to fecal microbiota alterations, suggesting a function of the stomach in gastrointestinal (GI) homeostasis. We sought to determine the taxonomic bacterial biogeography of the upper GI tract, including different sites within the human stomach (cardia, corpus, and antrum), adjacent upstream (esophagus) and downstream (duodenum) locations, and luminal contents (aspirate), as well as whole-stomach samples from mice and gerbils. Qualitative and quantitative DNA- and RNA-based taxonomic microbiota analyses were combined to study the relationship of relative and absolute bacterial abundances and transcriptionally active bacterial microbiota components in the stomach of humans and mice. Stomach microbiota compositions resembled those of esophagus and duodenum. However, along the descending GI tract, the relative abundances of specific oropharyngeal commensals decreased (Streptococcus) or increased (Rothia mucilaginosa, Porphyromonas, and Lachnospiraceae). Furthermore, the compositional similarity (weighted UniFrac) between stomach aspirates and esophageal biopsy samples increased with gastric Streptococcus relative abundance. In both human aspirate and mouse stomach samples, Firmicutes were more abundant among transcriptionally active bacteria than Bacteroidetes. The relative abundance of Firmicutes in the stomach was negatively correlated and that of Bacteroidetes was positively correlated with absolute bacterial abundance, suggesting a disproportionate increase of Bacteroidetes over Firmicutes at higher bacterial densities. Human, mouse, and gerbil stomach samples showed similarities at higher taxonomic levels but differences at lower taxonomic levels. Our findings suggest selective enrichment and depletion of specific bacterial taxa in the stomach and Firmicutes being transcriptionally more active than Bacteroidetes that increase in relative abundance with total bacterial load. IMPORTANCE Clinical stomach interventions, such as acid inhibition or bypass surgery, have been linked to fecal microbiota alterations. We demonstrate that the stomach microbiota largely overlaps those of adjacent gastrointestinal locations and identify gradual decreases and increases in the relative abundances of specific bacteria within the stomach, suggesting selective enrichment and depletion. Moreover, similarities between stomach and esophagus samples are proportional to the concentrations of Streptococcus (Firmicutes) in the stomach. The relative abundance of Firmicutes in the stomach, compared to that of Bacteroidetes, is increased in RNA relative to DNA, indicating higher transcriptional activity. Moreover, increased absolute bacterial loads are associated with decreased relative abundance of Firmicutes and higher relative abundance of Bacteroidetes. Our findings characterize the stomach microbiota as influenced by Bacteroidetes influx against a background of transcriptionally more active Firmicutes. Human, mouse, and gerbil stomach microbiotas differ at lower taxonomic levels, which might affect the utility of these model organisms.

Gut microbiota-dependent modulation of innate immunity and lymph node remodeling affects cardiac allograft outcomes.

We hypothesized that the gut microbiota influences survival of murine cardiac allografts through modulation of immunity. Antibiotic pretreated mice received vascularized cardiac allografts and fecal microbiota transfer (FMT), along with tacrolimus immunosuppression. FMT source samples were from normal, pregnant (immune suppressed), or spontaneously colitic (inflammation) mice. Bifidobacterium pseudolongum (B. pseudolongum) in pregnant FMT recipients was associated with prolonged allograft survival and lower inflammation and fibrosis, while normal or colitic FMT resulted in inferior survival and worse histology. Transfer of B. pseudolongum alone resulted in reduced inflammation and fibrosis. Stimulation of DC and macrophage lines with B. pseudolongum induced the antiinflammatory cytokine IL-10 and homeostatic chemokine CCL19 but induced lesser amounts of the proinflammatory cytokines TNFα and IL-6. In contrast, LPS and Desulfovibrio desulfuricans (D. desulfuricans), more abundant in colitic FMT, induced a more inflammatory cytokine response. Analysis of mesenteric and peripheral lymph node structure showed that B. pseudolongum gavage resulted in a higher laminin α4/α5 ratio in the lymph node cortical ridge, indicative of a suppressive environment, while D. desulfuricans resulted in a lower laminin α4/α5 ratio, supportive of inflammation. Discrete gut bacterial species alter immunity and may predict graft outcomes through stimulation of myeloid cells and shifts in lymph node structure and permissiveness.

Microbiota­implications for immunity and transplantation.

Each individual harbours a unique set of commensal microorganisms, collectively referred to as the microbiota. Notably, these microorganisms exceed the number of cells in the human body by 10-fold. This finding has accelerated a shift in our understanding of human physiology, with the realization that traits necessary for health are both encoded and influenced by the human genome and the microbiota. Our understanding of the aetiology of complex diseases has, therefore, evolved with increasing awareness that the human microbiota has an active and critical role in maintaining health and inducing disease. Indeed, findings from bioinformatic studies indicate that the microbiota and microbiome have multiple effects on the innate and adaptive immune systems, with effects on infection, autoimmune disease and cancer. In this Review, we first address the important statistical and informatics aspects that should be considered when characterizing the composition of microbiota. We next highlight the effects of the microbiota on the immune system and the implications of these effects on organ failure and transplantation. Finally, we reflect on the future perspectives for studies of the microbiota, including novel diagnostic tests and therapeutics.

A physicians' wish list for the clinical application of intestinal metagenomics.

Christoph Steininger and colleagues explore how multiple infectious, autoimmune, metabolic, and neoplastic diseases have been associated with changes in the intestinal microbiome, although a cause-effect relationship is often difficult to establish. Integration of metagenomics into clinical medicine is a challenge, and the authors highlight clinical approaches that are of high priority for the useful medical application of metagenomics. Please see later in the article for the Editors' Summary.

Efficacy of combined jejunal and colonic fecal microbiota transplantation for recurrent Clostridium difficile Infection.

The prevalence of recurrent Clostridium difficile infection (RCDI) is increasing; fecal microbiota transplantation (FMT) is an effective therapy. However, there have been no studies of the efficacy of a single session of combined enteral and colonic FMT or characterizations of changes in the microbiota between donors and recipients. We performed a study of 27 patients with RCDI who were given a fixed volume of processed fecal filtrate via enteroscopy and colonoscopy in a single session. Patients were closely monitored, and fecal samples were collected from 2 patient-donor pairs for 16S rRNA analysis. All patients had reduced stool frequency, abdominal pain, white blood cell counts, and elimination of fecal C difficile toxin (P < .05). FMT increased microbial diversity, increasing proportions of Lachnospiraceae (phylum Firmicutes) and reducing proportions of Enterobacteriaceae. FMT was associated with marked changes in the composition of fecal microbiota in 2 patients with RCDI.

Characterization of the bacterial community of the chemically defended Hawaiian sacoglossan Elysia rufescens.

Sacoglossans are characterized by the ability to sequester functional chloroplasts from their algal diet through a process called kleptoplasty, enabling them to photosynthesize. The bacterial diversity associated with sacoglossans is not well understood. In this study, we coupled traditional cultivation-based methods with 454 pyrosequencing to examine the bacterial communities of the chemically defended Hawaiian sacoglossan Elysia rufescens and its secreted mucus. E. rufescens contains a defense molecule, kahalalide F, that is possibly of bacterial origin and is of interest because of its antifungal and anticancer properties. Our results showed that there is a diverse bacterial assemblage associated with E. rufescens and its mucus, with secreted mucus harboring higher bacterial richness than entire-E. rufescens samples. The most-abundant bacterial groups affiliated with E. rufescens and its mucus are Mycoplasma spp. and Vibrio spp., respectively. Our analyses revealed that the Vibrio spp. that were highly represented in the cultivable assemblage were also abundant in the culture-independent community. Epifluorescence microscopy and matrix-assisted laser desorption-ionization mass spectrometry (MALDI-MS) were utilized to detect the chemical defense molecule kahalalide F on a longitudinal section of the sacoglossan.

Genome sequences of 65 Helicobacter pylori strains isolated from asymptomatic individuals and patients with gastric cancer, peptic ulcer disease, or gastritis.

Helicobacter pylori, inhabitant of the gastric mucosa of over half of the world population, with decreasing prevalence in the U.S., has been associated with a variety of gastric pathologies. However, the majority of H. pylori-infected individuals remain asymptomatic, and negative correlations between H. pylori and allergic diseases have been reported. Comprehensive genome characterization of H. pylori populations from different human host backgrounds including healthy individuals provides the exciting potential to generate new insights into the open question whether human health outcome is associated with specific H. pylori genotypes or dependent on other environmental factors. We report the genome sequences of 65 H. pylori isolates from individuals with gastric cancer, preneoplastic lesions, peptic ulcer disease, gastritis, and from asymptomatic adults. Isolates were collected from multiple locations in North America (USA and Canada) as well as from Columbia and Japan. The availability of these H. pylori genome sequences from individuals with distinct clinical presentations provides the research community with a resource for detailed investigations into genetic elements that correlate either positively or negatively with the epidemiology, human host adaptation, and gastric pathogenesis and will aid in the characterization of strains that may favor the development of specific pathology, including gastric cancer.

Complex microbiome underlying secondary and primary metabolism in the tunicate-Prochloron symbiosis.

The relationship between tunicates and the uncultivated cyanobacterium Prochloron didemni has long provided a model symbiosis. P. didemni is required for survival of animals such as Lissoclinum patella and also makes secondary metabolites of pharmaceutical interest. Here, we present the metagenomes, chemistry, and microbiomes of four related L. patella tunicate samples from a wide geographical range of the tropical Pacific. The remarkably similar P. didemni genomes are the most complex so far assembled from uncultivated organisms. Although P. didemni has not been stably cultivated and comprises a single strain in each sample, a complete set of metabolic genes indicates that the bacteria are likely capable of reproducing outside the host. The sequences reveal notable peculiarities of the photosynthetic apparatus and explain the basis of nutrient exchange underlying the symbiosis. P. didemni likely profoundly influences the lipid composition of the animals by synthesizing sterols and an unusual lipid with biofuel potential. In addition, L. patella also harbors a great variety of other bacterial groups that contribute nutritional and secondary metabolic products to the symbiosis. These bacteria possess an enormous genetic potential to synthesize new secondary metabolites. For example, an antitumor candidate molecule, patellazole, is not encoded in the genome of Prochloron and was linked to other bacteria from the microbiome. This study unveils the complex L. patella microbiome and its impact on primary and secondary metabolism, revealing a remarkable versatility in creating and exchanging small molecules.