Microbial phylogenetic diversity preserved in facies-specific modern, recent, Holocene and Pleistocene hot-spring travertine deposits of Yellowstone and Turkey

A systematic evaluation has been undertaken of the mechanisms and products of microbial community preservation within modern-to-ancient terrestrial hot-spring calcium carbonate (CaCO3) limestone deposits called travertine. Microbial 16S rRNA gene sequences preserved within Modern travertine deposited within the Proximal Slope Facies (PSF) at Mammoth Hot Springs (MHS), Yellowstone National Park, has been directly compared with analogous Holocene-Late Pleistocene PSF travertine at Gardiner, Montana, and Middle Pleistocene PSF travertine in Denizli, Turkey. Analyses have included: (1) Modern microbial communities inhabiting the PSF of an actively depositing travertine hot-spring system (water, microbial mats, travertine) at MHS (0 YBP Modern travertine, Angel Terrace, Spring AT-1); (2) 9 YBP Modern PSF travertine (Angel Terrace, Spring AT-2; MHS); (2) ~100 YBP Recent travertine (New Highland Terrace, MHS); (3) ~4,000 YBP Holocene travertine (USGS Y-10 Core); (3) ~30,000 YBP Late Pleistocene travertine (Gardiner Quarry); and (4) ~1.1 Ma YBP Middle Pleistocene travertine (all of the Mammoth Hot Springs (YNP) and (Cakmak Quarry, Turkey). Genomic DNA entombed during rapid (up to 5 mm/day) travertine deposition and preserved within CaCO3 fluid inclusions and between crystals, was extracted via bulk rock drilling under sterile clean room conditions. Pooled 16S rRNA gene sequence libraries were constructed via polymerase chain reactions (PCRs), terminal-restriction fragment length polymorphisms (T-RFLP), and MiSeq amplicon sequencing. Blast searches using multiple web-based bioinformatics tools identified over 400 operational taxonomic units (OTUs) affiliated with a total of 19 phyla (16 phyla and 3 candidate phyla) within the Domain Bacteria. Previous analyses of the living microbial communities inhabiting modern active PSF depositional environments at MHS have provided a baseline with which to compare the ancient travertine analyses. Combined results from both MHS and Denizli indicate that only 3 of 19 phyla were detected in PSF travertine samples of all ages. Increasing depositional age of the travertine deposits was associated with increasing extents of post-depositional water-rock geochemical alteration (diagenesis). Microbial community structure shifted across this spatial and temporal transect from being dominated by Cyanobacteria and Proteobacteria in the Modern and Recent, to dominance by Firmicutes in the Holocene and Pleistocene. This is not unexpected, as endospores of Firmicutes are known to be resistant and also persist under harsh environmental conditions and therefore show higher relative abundance in ancient samples in comparison to modern samples. Preliminary blast search results imply that libraries from all PSF travertine samples of all ages contain photoautotrophic, chemoautotrophic and heterotrophic metabolic activities, which is likely the result of both original hot-spring depositional processes and secondary diagenetic alteration

Global phylogeography and ancient evolution of the widespread human gut virus crAssphage

Microbiomes are vast communities of microorganisms and viruses that populate all natural ecosystems. Viruses have been considered to be the most variable component of microbiomes, as supported by virome surveys and examples of high genomic mosaicism. However, recent evidence suggests that the human gut virome is remarkably stable compared with that of other environments. Here, we investigate the origin, evolution and epidemiology of crAssphage, a widespread human gut virus. Through a global collaboration, we obtained DNA sequences of crAssphage from more than one-third of the world's countries and showed that the phylogeography of crAssphage is locally clustered within countries, cities and individuals. We also found fully colinear crAssphage-like genomes in both Old-World and New-World primates, suggesting that the association of crAssphage with primates may be millions of years old. Finally, by exploiting a large cohort of more than 1,000 individuals, we tested whether crAssphage is associated with bacterial taxonomic groups of the gut microbiome, diverse human health parameters and a wide range of dietary factors. We identified strong correlations with different clades of bacteria that are related to Bacteroidetes and weak associations with several diet categories, but no significant association with health or disease. We conclude that crAssphage is a benign cosmopolitan virus that may have coevolved with the human lineage and is an integral part of the normal human gut virome

Global phylogeography and ancient evolution of the widespread human gut virus crAssphage

Microbiomes are vast communities of microorganisms and viruses that populate all natural ecosystems. Viruses have been considered to be the most variable component of microbiomes, as supported by virome surveys and examples of high genomic mosaicism. However, recent evidence suggests that the human gut virome is remarkably stable compared with that of other environments. Here, we investigate the origin, evolution and epidemiology of crAssphage, a widespread human gut virus. Through a global collaboration, we obtained DNA sequences of crAssphage from more than one-third of the world’s countries and showed that the phylogeography of crAssphage is locally clustered within countries, cities and individuals. We also found fully colinear crAssphage-like genomes in both Old-World and New-World primates, suggesting that the association of crAssphage with primates may be millions of years old. Finally, by exploiting a large cohort of more than 1,000 individuals, we tested whether crAssphage is associated with bacterial taxonomic groups of the gut microbiome, diverse human health parameters and a wide range of dietary factors. We identified strong correlations with different clades of bacteria that are related to Bacteroidetes and weak associations with several diet categories, but no significant association with health or disease. We conclude that crAssphage is a benign cosmopolitan virus that may have coevolved with the human lineage and is an integral part of the normal human gut virome

Insights into the primate microbiome

The study of the primate microbiome (genetic and environmental content of the microbiota) has served as a critical tool for understanding the impact of microbial communities on host health and disease. An in-depth understanding of the healthy primate microbial community composition and function is needed in order to identify changes in the microbiome that cause and/or result in unhealthy states in the host. In this dissertation project, I focused my efforts on better understanding the factors that influence or shape a healthy non-human primate (NHP) microbiome using high throughput sequencing (16S rRNA and metagenomics), bioinformatics and various statistical tools. In the first study, investigating the confounding effects of sample storage preservatives in the form of technical variations in the microbiome, I was able to determine whether technical variations due to sample storage preservatives (no preservative, 96% ethanol and RNAlater) outweighed alterations due to biological factors of interest. The results showed that while these technical variations in the microbiome are not large enough to obscure the differences due to host species, body site and habitat differences, they may promote misleading interpretations if they have large enough effect sizes compared to the biological factors such as host population. While most microbiome studies focus on one body site from one host species at a time, I leveraged the advantages of multi-body site and multi host species dataset to determine the extent to which the various factors influence the different types of microbiome in different host species. In the second study, I analyzed the microbial community composition of eight distinct sites (rectal, fecal, oral, nasal, otic/ear, vaginal, penile and skin) from 17 non-human primate species, the majority of which were wild. The results revealed a possibly conserved NHP oral microbiome with relatively low alpha and beta diversities as seen by the distinct clustering of the oral samples away from samples from all other body sites. Without the oral samples however, host species differences shaped the microbiome with samples clustered by body site within each host species due to niche specialization. The results also showed a stronger captivity effect on the gut microbiome in comparison to the vaginal microbiome possibly due to dietary differences between wild and captive individuals which in turn affects the microbiome more than the vaginal microbiome. These results help to elucidate the wide variation present in the microbiomes from various body sites both within and between NHPs. To better understand the possibly conserved NHP oral microbiome observed in the second study, I analyzed the oral samples of 12 predominantly wild non-human primates (NHPs) to better understand the extent to which factors including host species, host phylogeny, diet and habitat influence the NHP oral microbiome in this study. While both alpha and beta diversity results showed statistically significant differences in the NHP oral microbiome due to all four factors, LEfSe and MaAsLin2 results revealed significant associations between the microbiome and host species. These results are also indicative of a distinct, possibly conserved NHP oral microbiome with certain host specific differences. Finally, I also analyzed and compared the gut microbiomes of five closely related wild lemurs (Propithecus verreauxi, Propithecus diadema, Propithecus edwardsi, Varecia variegata, and Lemur catta) occupying different dietary niches. The results revealed host-specific gut microbiota clustered within broad clusters of host phylogenetic and dietary groups. While the lowest diversity was observed in the sifakas (folivores), the omnivorous L. catta showed the highest alpha diversity possibly due to their generalist feeding strategy with wide range of food sources. With the clear confoundment between host phylogeny and diet, I included three old world monkeys (Procolobus rufomitratus tephrosceles, Cercopithecus ascanius schmidti and Papio Anubis) in the analysis and obtained 22 discriminative biomarkers due host phylogeny compared to 5 due diet suggestive of a stronger host phylogenetic group effect on the NHP gut microbiome compared to diet.LimitedAuthor requested closed access (OA after 2yrs) in Vireo ETD syste

Plant cell wall degrading enzymes and their potential application to biofuel production

The soaring high price of petroleum price as well as the impact of the fossil fuel combustion process on atmospheric carbon dioxide (CO2) has necessitated the development of alternative fuel energy. Even with the most recent technology, oil, natural gas, and coal will emit not only climate-threatening greenhouse gases and other pollutants, but also the quantity of undiscovered stocks will always be a matter of great concern Plants have evolved photosynthetic mechanisms in which solar energy is used to fix CO2 into carbohydrates. Thus, combustion of biofuels, derived from plant biomass, can be considered a potentially carbon neutral process. Plants cell walls which are composed mostly of lignocellulosic materials (lignin, cellulose, and hemicellulose) are recalcitrant and are difficult to breakdown. This therefore represents a major limitation in the conversion of plant biomass to biofuels. Microbes have evolved a plethora of enzymatic strategies for hydrolyzing plant cell wall into its constituent sugars for subsequent fermentation to biofuels. Therefore, microorganisms are considered an important source of biocatalysts in the emerging biofuel industry. Caldicellulosiruptor bescii and Themoanaerobacterium bryantii are both thermophilic anaerobes capable of secreting enzymes for degrading plant cell walls. To gain insight into the cellular machinery that these organisms elaborate to degrade cellulose and the hemicellulosic polymer xylan, genes with various putative functions from both organisms were identified, cloned and expressed the recombinant proteins in Escherichia coli. A hemicellulase gene cluster from T. bryantii encoding genes predicted to have endoxylanase activity, -xylosidase activity, -glucoronidase activity, acetyl xylan esterase activity as well as oxido-reductase activity was isolated for further studies. We hypothesized that the endoxylanase should work synergistically with the other accessory enzymes in the cluster to break down xylan releasing mainly xylose for the utilization of the organism. As expected the endoxylanase was able to hydrolyze a number of xylan containing polysaccharides releasing undecorated xylo-oligosaccharides with the -xylosidases converting the xylobiose released to xylose. A putative β-glucosidase Cb486 from C. bescii was screened for enzymatic properties, and it was shown to exhibit activity against pNP-α-L-arabinopyranoside, pNP-β-D-fucopyranoside, pNP-β-D-galactopyranoside, pNP-β-D-glucopyranoside, pNP-β-D-xylopyranoside and pNP-β-D-cellobioside suggesting that Cb486 is a multi-functional enzyme. We therefore hypothesized that Cb486 incubated with endoglucanases or endoxylanases should be able to enhance the release of glucose or xylose from cellulose and xylans respectively. As expected, Cb486 was also able to work synergistically with a number of endo-glucanases from the genome of C. bescii to release glucose from cellulosic substrates as well as xylose from xylan containing substrates

How Workflow Documentation Facilitates Curation Planning

The description of the specific processes and artifacts that led to the creation of a data product provide a detailed picture of data provenance in the form of a workflow. The Site-Based Data Curation project, hosted by the Center for Informatics Research in Science and Scholarship at the University of Illinois, has been investigating how workflows can be used in developing curation processes and policies that move curation "upstream" in the research process. The team has documented an individual workflow for geobiology data collected during a single field trip to Yellowstone National Park. This specific workflow suggests a generalized three-part process for field data collection that comprises three distinct elements: a Planning Stage, a Fieldwork Stage, and a Processing and Analysis Stage. Beyond supplying an account of data provenance, the workflow has allowed the team to identify 1) points of intervention for curation processes and 2) data products that are likely candidates for sharing or deposit. Although these objects may be viewed by individual researchers as 'intermediate' data products, discussions with geobiology researchers have suggested that with appropriate packaging and description they may serve as valuable observational data for other researchers. Curation interventions may include the introduction of regularized data formats during the planning process, data description procedures, the identification and use of established controlled vocabularies, and data quality and validation procedures. We propose a poster that shows the individual workflow and our generalization into a three-stage process. We plan to discuss with attendees how well the three-stage view applies to other types of field-based research, likely points of intervention, and what kinds of interventions are appropriate and feasible in the example workflow.Institute of Museum and Library Service National Leadership Grant number LG-06-12-0706-12Ope

Xylan Deconstruction by Thermophilic <i>Thermoanaerobacterium bryantii</i> Hemicellulases Is Stimulated by Two Oxidoreductases

Thermoanaerobacterium bryantii strain mel9T is a thermophilic bacterium isolated from a waste pile of a corn-canning factory. The genome of T. bryantii mel9T was sequenced and a hemicellulase gene cluster was identified. The cluster encodes seven putative enzymes, which are likely an endoxylanase, an α-glucuronidase, two oxidoreductases, two β-xylosidases, and one acetyl xylan esterase. These genes were designated tbxyn10A, tbagu67A, tbheoA, tbheoB, tbxyl52A, tbxyl39A, and tbaxe1A, respectively. Only TbXyn10A released reducing sugars from birchwood xylan, as shown by thin-layer chromatography analysis. The five components of the hemicellulase cluster (TbXyn10A, TbXyl39A, TbXyl52A, TbAgu67A, and TbAxe1A) functioned in synergy to hydrolyze birchwood xylan. Surprisingly, the two putative oxidoreductases increased the enzymatic activities of the gene products from the xylanolytic gene cluster in the presence of NADH and manganese ions. The two oxidoreductases were therefore named Hemicellulase-Enhancing Oxidoreductases (HEOs). All seven enzymes were thermophilic and acted in synergy to degrade xylans at 60 °C. Except for TbXyn10A, the other enzymes encoded by the gene cluster were conserved with high amino acid identities (85–100%) in three other Thermoanaerobacterium species. The conservation of the gene cluster is, therefore, suggestive of an important role of these enzymes in xylan degradation by these bacteria. The mechanism for enhancement of hemicellulose degradation by the HEOs is under investigation. It is anticipated, however, that the discovery of these new actors in hemicellulose deconstruction will have a significant impact on plant cell wall deconstruction in the biofuel industry

Site-Based Data Curation at Yellowstone National Park

The Site-Based Data Curation (SBDC) project at the University of Illinois at Urbana-Champaign is developing a framework of policies and processes for the curation of research data generated at scientifically significant sites. The model will be built around the case of systems geobiology at Yellowstone National Park.IMLS / LG-06-12-0706-12unpublishedis peer reviewe

Diagnostic and prognostic potential of the microbiome in ovarian cancer treatment response

Abstract Ovarian cancer (OC) is the second most common gynecological malignancy and the fifth leading cause of death due to cancer in women in the United States mainly due to the late-stage diagnosis of this cancer. It is, therefore, critical to identify potential indicators to aid in early detection and diagnosis of this disease. We investigated the microbiome associated with OC and its potential role in detection, progression as well as prognosis of the disease. We identified a distinct OC microbiome with general enrichment of several microbial taxa, including Dialister, Corynebacterium, Prevotella, and Peptoniphilus in the OC cohort in all body sites excluding stool and omentum which were not sampled from the benign cohort. These taxa were, however, depleted in the advanced-stage and high-grade OC patients compared to early-stage and low-grade OC patients suggestive of decrease accumulation in advanced disease and could serve as potential indicators for early detection of OC. Similarly, we also observed the accumulation of these mainly pathogenic taxa in OC patients with adverse treatment outcomes compared to those without events and could also serve as potential indicators for predicting patients’ responses to treatment. These findings provide important insights into the potential use of the microbiome as indicators in (1) early detection of and screening for OC and (2) predicting patients’ response to treatment. Given the limited number of patients enrolled in the study, these results would need to be further investigated and confirmed in a larger study