Matthew C. Ross

Evaluation of 16s rDNA-based community profiling for human microbiome research

The Human Microbiome Project will establish a reference data set for analysis of the microbiome of healthy adults by surveying multiple body sites from 300 people and generating data from over 12,000 samples. To characterize these samples, the participating sequencing centers evaluated and adopted 16S rDNA community profiling protocols for ABI 3730 and 454 FLX Titanium sequencing. In the course of establishing protocols, we examined the performance and error characteristics of each technology, and the relationship of sequence error to the utility of 16S rDNA regions for classification- and OTU-based analysis of community structure. The data production protocols used for this work are those used by the participating centers to produce 16S rDNA sequence for the Human Microbiome Project. Thus, these results can be informative for interpreting the large body of clinical 16S rDNA data produced for this project.

The gut mycobiome of the Human Microbiome Project healthy cohort

BackgroundMost studies describing the human gut microbiome in healthy and diseased states have emphasized the bacterial component, but the fungal microbiome (i.e., the mycobiome) is beginning to gain recognition as a fundamental part of our microbiome. To date, human gut mycobiome studies have primarily been disease centric or in small cohorts of healthy individuals. To contribute to existing knowledge of the human mycobiome, we investigated the gut mycobiome of the Human Microbiome Project (HMP) cohort by sequencing the Internal Transcribed Spacer 2 (ITS2) region as well as the 18S rRNA gene.ResultsThree hundred seventeen HMP stool samples were analyzed by ITS2 sequencing. Fecal fungal diversity was significantly lower in comparison to bacterial diversity. Yeast dominated the samples, comprising eight of the top 15 most abundant genera. Specifically, fungal communities were characterized by a high prevalence of Saccharomyces, Malassezia, and Candida, with S. cerevisiae, M. restricta, and C. albicans operational taxonomic units (OTUs) present in 96.8, 88.3, and 80.8% of samples, respectively. There was a high degree of inter- and intra-volunteer variability in fungal communities. However, S. cerevisiae, M. restricta, and C. albicans OTUs were found in 92.2, 78.3, and 63.6% of volunteers, respectively, in all samples donated over an approximately 1-year period. Metagenomic and 18S rRNA gene sequencing data agreed with ITS2 results; however, ITS2 sequencing provided greater resolution of the relatively low abundance mycobiome constituents.ConclusionsCompared to bacterial communities, the human gut mycobiome is low in diversity and dominated by yeast including Saccharomyces, Malassezia, and Candida. Both inter- and intra-volunteer variability in the HMP cohort were high, revealing that unlike bacterial communities, an individual’s mycobiome is no more similar to itself over time than to another person’s. Nonetheless, several fungal species persisted across a majority of samples, evidence that a core gut mycobiome may exist. ITS2 sequencing data provided greater resolution of the mycobiome membership compared to metagenomic and 18S rRNA gene sequencing data, suggesting that it is a more sensitive method for studying the mycobiome of stool samples.

16S gut community of the Cameron County Hispanic Cohort

BackgroundObesity and type 2 diabetes (T2D) are major public health concerns worldwide, and their prevalence has only increased in recent years. Mexican Americans are disproportionately afflicted by obesity and T2D, and rates are even higher in the United States-Mexico border region. To determine the factors associated with the increased risk of T2D, obesity, and other diseases in this population, the Cameron County Hispanic Cohort was established in 2004.ResultsIn this study, we characterized the 16S gut community of a subset of 63 subjects from this unique cohort. We found that these communities, when compared to Human Microbiome Project subjects, exhibit community shifts often observed in obese and T2D individuals in published studies. We also examined microbial network relationships between operational taxonomic units (OTUs) in the Cameron County Hispanic Cohort (CCHC) and three additional datasets. We identified a group of seven genera that form a tightly interconnected network present in all four tested datasets, dominated by butyrate producers, which are often increased in obese individuals while being depleted in T2D patients.ConclusionsThrough a combination of increased disease prevalence and relatively high gut microbial homogeneity in the subset of CCHC members we examined, we believe that the CCHC may represent an ideal community to dissect mechanisms underlying the role of the gut microbiome in human health and disease. The lack of CCHC subject gut community segregation based on all tested metadata suggests that the community structure we observe in the CCHC likely occurs early in life, and endures. This persistent ‘disease’-related gut microbial community in CCHC subjects may enhance existing genetic or lifestyle predispositions to the prevalent diseases of the CCHC, leading to increased attack rates of obesity, T2D, non-alcoholic fatty liver disease, and others.

Maternal micronutrients can modify colonic mucosal microbiota maturation in murine offspring

Epidemiologic data suggest that early nutritional exposures may inflict persistent changes in the developing mammalian “super-organism” (i.e., the host and its residing microbiota). Such persistent modifications could predispose young adults to inflammatory bowel diseases (IBD). We recently observed that the dietary supplementation of four micronutrients to dams augmented colitis susceptibility in murine offspring in association with mucosal microbiota composition changes. In this study the effects of the four micronutrients on the microbiota of dams and female mice was examined. Additionally, age dependent microbiota composition shifts during pediatric development were delineated from the previous offspring data sets. Maternal and adult female microbiota did not separate secondary to the nutritional intervention. Significant microbiota composition changes occurred from postnatal day 30 (P30) to P90 at the level of 1 phylum and 15 genera. Most of these changes were absent or opposite in the maternally supplemented offspring. Nutritionally induced alterations in mucosal microbiota maturation may be contributors to colitis susceptibility in mammals.

Monotonous diets protect against acute colitis in mice: Epidemiologic and therapeutic implications

Objectives: Multiple characteristics of industrialization have been proposed to contribute to the global emergence of inflammatory bowel diseases (IBDs: Crohn disease and ulcerative colitis). Major changes in eating habits during the last decades and the effectiveness of exclusive enteral nutrition in the treatment of Crohn disease indicate the etiologic importance of dietary intake in IBDs. A uniform characteristic of nutrition in developing countries (where the incidence of IBD is low) and exclusive enteral nutrition is their consistent nature for prolonged periods; however, the potentially beneficial effect of dietary monotony in respect to mammalian intestinal inflammation has not been examined. Methods: The association between alternating (2 different complete chows) and persistent regular diets, and dextran sulfate sodium colitis susceptibility in C57BL/6J mice was studied. Colonic mucosal microbiota changes were investigated by high-throughput pyrosequencing of the 16S rRNA gene. Results: The severity of colitis increased upon dietary alternation compared with consistent control feeding. The microbiota of the alternating nutritional group clustered discretely from both control groups. Conclusions: Our findings highlight that monotonous dietary intake may decrease mammalian vulnerability against colitis in association with microbiota separation. The epidemiologic and therapeutic implications of our results are also discussed.

Maximal viral information recovery from sequence data using VirMAP

Accurate classification of the human virome is critical to a full understanding of the role viruses play in health and disease. This implies the need for sensitive, specific, and practical pipelines that return precise outputs while still enabling case-specific post hoc analysis. Viral taxonomic characterization from metagenomic data suffers from high background noise and signal crosstalk that confounds current methods. Here we develop VirMAP that overcomes these limitations using techniques that merge nucleotide and protein information to taxonomically classify viral reconstructions independent of genome coverage or read overlap. We validate VirMAP using published data sets and viral mock communities containing RNA and DNA viruses and bacteriophages. VirMAP offers opportunities to enhance metagenomic studies seeking to define virome-host interactions, improve biosurveillance capabilities, and strengthen molecular epidemiology reporting.Viral taxonomic characterization from metagenomic data suffers from high background noise and signal crosstalk. Here, the authors develop VirMAP, a novel pipeline for analyses of metagenomic data that classifies viral reconstructions independent of genome coverage or read overlap.

Giardia Infection of the Small Intestine Induces Chronic Colitis in Genetically Susceptible Hosts

The lumen-dwelling protozoan Giardia is an important parasitic cause of diarrheal disease worldwide. Infection can persist over extended periods with minimal intestinal inflammation, suggesting that Giardia may attenuate host responses to ensure its survival, although clearance eventually occurs in most cases. IL-10 is an anti-inflammatory regulator critical for intestinal homeostasis and controlling host responses to bacterial exposure, yet its potential role in coordinating antiprotozoal host defense in the intestine is not known. In this study, we found that murine infection with the natural enteric pathogen Giardia muris induced a transient IL-10 response after 2–4 wk at the primary site of infection in the upper small intestine, but parasite colonization and eradication were not affected by the absence of the cytokine in gene-targeted mice. However, IL-10 was critical for controlling infection-associated immunological sequelae in the colon because severe and persistent diarrhea and colitis were observed in IL-10–deficient mice within 1–2 wk postinfection but not in uninfected littermate controls. Inflammation was characterized by epithelial hyperplasia, neutrophil and macrophage expansion, and Th1 induction and could be prevented by blockade of IL-12/IL-23 p40 but not depletion of CD11c+ dendritic cells. Furthermore, the intestinal microbiota underwent characteristic shifts in composition and was required for disease because antibiotics and loss of TLR signaling in MyD88-deficient mice protected against colitis. Together, our data suggest that transient infection by a luminal and seemingly noninflammatory pathogen can trigger sustained colitis in genetically susceptible hosts, which has broader implications for understanding postinfectious syndromes and other chronic intestinal inflammatory conditions.