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  • 1
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    Exploring microbial diversity and taxonomy using SSU rRNA hypervariable tag sequencing (2009)
    Public Library of Science
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    Publication Date: 2022-05-25
    Description: © 2008 Huse et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License. The definitive version was published in PLoS Genetics 4 (2008): e1000255, doi:10.1371/journal.pgen.1000255.
    Description: Massively parallel pyrosequencing of hypervariable regions from small subunit ribosomal RNA (SSU rRNA) genes can sample a microbial community two or three orders of magnitude more deeply per dollar and per hour than capillary sequencing of full-length SSU rRNA. As with full-length rRNA surveys, each sequence read is a tag surrogate for a single microbe. However, rather than assigning taxonomy by creating gene trees de novo that include all experimental sequences and certain reference taxa, we compare the hypervariable region tags to an extensive database of rRNA sequences and assign taxonomy based on the best match in a Global Alignment for Sequence Taxonomy (GAST) process. The resulting taxonomic census provides information on both composition and diversity of the microbial community. To determine the effectiveness of using only hypervariable region tags for assessing microbial community membership, we compared the taxonomy assigned to the V3 and V6 hypervariable regions with the taxonomy assigned to full-length SSU rRNA sequences isolated from both the human gut and a deep-sea hydrothermal vent. The hypervariable region tags and full-length rRNA sequences provided equivalent taxonomy and measures of relative abundance of microbial communities, even for tags up to 15% divergent from their nearest reference match. The greater sampling depth per dollar afforded by massively parallel pyrosequencing reveals many more members of the “rare biosphere” than does capillary sequencing of the full-length gene. In addition, tag sequencing eliminates cloning bias and the sequences are short enough to be completely sequenced in a single read, maximizing the number of organisms sampled in a run while minimizing chimera formation. This technique allows the cost-effective exploration of changes in microbial community structure, including the rare biosphere, over space and time and can be applied immediately to initiatives, such as the Human Microbiome Project.
    Description: Woods Hole Center for Oceans and Human Health from the National Institutes of Health and National Science Foundation (NIH/NIEHS 1 P50 ES012742-01 and NSF/OCE 0430724-J Stegeman PI to MLS). NIH Director's Pioneer Award and Doris Duke Distinguished Clinical Scientist Award to DAR.
    Repository Name: Woods Hole Open Access Server
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  • 2
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    Unique prokaryotic consortia in geochemically distinct sediments from Red Sea Atlantis II and Discovery Deep brine pools (2012)
    Public Library of Science
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    Publication Date: 2022-05-25
    Description: © The Author(s), 2012. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in PLoS ONE 7 (2012): e42872, doi:10.1371/journal.pone.0042872.
    Description: The seafloor is a unique environment, which allows insights into how geochemical processes affect the diversity of biological life. Among its diverse ecosystems are deep-sea brine pools - water bodies characterized by a unique combination of extreme conditions. The ‘polyextremophiles’ that constitute the microbial assemblage of these deep hot brines have not been comprehensively studied. We report a comparative taxonomic analysis of the prokaryotic communities of the sediments directly below the Red Sea brine pools, namely, Atlantis II, Discovery, Chain Deep, and an adjacent brine-influenced site. Analyses of sediment samples and high-throughput pyrosequencing of PCR-amplified environmental 16S ribosomal RNA genes (16S rDNA) revealed that one sulfur (S)-rich Atlantis II and one nitrogen (N)-rich Discovery Deep section contained distinct microbial populations that differed from those found in the other sediment samples examined. Proteobacteria, Actinobacteria, Cyanobacteria, Deferribacteres, and Euryarchaeota were the most abundant bacterial and archaeal phyla in both the S- and N-rich sections. Relative abundance-based hierarchical clustering of the 16S rDNA pyrotags assigned to major taxonomic groups allowed us to categorize the archaeal and bacterial communities into three major and distinct groups; group I was unique to the S-rich Atlantis II section (ATII-1), group II was characteristic for the N-rich Discovery sample (DD-1), and group III reflected the composition of the remaining sediments. Many of the groups detected in the S-rich Atlantis II section are likely to play a dominant role in the cycling of methane and sulfur due to their phylogenetic affiliations with bacteria and archaea involved in anaerobic methane oxidation and sulfate reduction.
    Description: This work was supported by King Abdullah University for Science and Technology Global Collaborative Partners (GCR) program.
    Repository Name: Woods Hole Open Access Server
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  • 3
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    Shifts in the microbial community composition of Gulf Coast beaches following beach oiling (2014)
    Public Library of Science
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    Publication Date: 2022-05-25
    Description: © The Author(s), 2013. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in PLoS One 8 (2013): e74265, doi:10.1371/journal.pone.0074265.
    Description: Microorganisms associated with coastal sands serve as a natural biofilter, providing essential nutrient recycling in nearshore environments and acting to maintain coastal ecosystem health. Anthropogenic stressors often impact these ecosystems, but little is known about whether these disturbances can be identified through microbial community change. The blowout of the Macondo Prospect reservoir on April 20, 2010, which released oil hydrocarbons into the Gulf of Mexico, presented an opportunity to examine whether microbial community composition might provide a sensitive measure of ecosystem disturbance. Samples were collected on four occasions, beginning in mid-June, during initial beach oiling, until mid-November from surface sand and surf zone waters at seven beaches stretching from Bay St. Louis, MS to St. George Island, FL USA. Oil hydrocarbon measurements and NOAA shoreline assessments indicated little to no impact on the two most eastern beaches (controls). Sequence comparisons of bacterial ribosomal RNA gene hypervariable regions isolated from beach sands located to the east and west of Mobile Bay in Alabama demonstrated that regional drivers account for markedly different bacterial communities. Individual beaches had unique community signatures that persisted over time and exhibited spatial relationships, where community similarity decreased as horizontal distance between samples increased from one to hundreds of meters. In contrast, sequence analyses detected larger temporal and less spatial variation among the water samples. Superimposed upon these beach community distance and time relationships, was increased variability in bacterial community composition from oil hydrocarbon contaminated sands. The increased variability was observed among the core, resident, and transient community members, indicating the occurrence of community-wide impacts rather than solely an overprinting of oil hydrocarbon-degrading bacteria onto otherwise relatively stable sand population structures. Among sequences classified to genus, Alcanivorax, Alteromonas, Marinobacter, Winogradskyella, and Zeaxanthinibacter exhibited the largest relative abundance increases in oiled sands.
    Description: Financial support for this work was provided by the University of Wisconsin-Milwaukee National Institute of Environmental Health Sciences program, grant ES-004184 to SLM and the Alfred P. Sloan Foundation's grant for the Rare Biosphere in the Built Environment MLS.
    Repository Name: Woods Hole Open Access Server
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  • 4
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    Core microbial functional activities in ocean environments revealed by global metagenomic profiling analyses (2014)
    Public Library of Science
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    Publication Date: 2022-05-25
    Description: © The Author(s), 2014. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in PLoS One 9 (2014): e97338, doi:10.1371/journal.pone.0097338.
    Description: Metagenomics-based functional profiling analysis is an effective means of gaining deeper insight into the composition of marine microbial populations and developing a better understanding of the interplay between the functional genome content of microbial communities and abiotic factors. Here we present a comprehensive analysis of 24 datasets covering surface and depth-related environments at 11 sites around the world's oceans. The complete datasets comprises approximately 12 million sequences, totaling 5,358 Mb. Based on profiling patterns of Clusters of Orthologous Groups (COGs) of proteins, a core set of reference photic and aphotic depth-related COGs, and a collection of COGs that are associated with extreme oxygen limitation were defined. Their inferred functions were utilized as indicators to characterize the distribution of light- and oxygen-related biological activities in marine environments. The results reveal that, while light level in the water column is a major determinant of phenotypic adaptation in marine microorganisms, oxygen concentration in the aphotic zone has a significant impact only in extremely hypoxic waters. Phylogenetic profiling of the reference photic/aphotic gene sets revealed a greater variety of source organisms in the aphotic zone, although the majority of individual photic and aphotic depth-related COGs are assigned to the same taxa across the different sites. This increase in phylogenetic and functional diversity of the core aphotic related COGs most probably reflects selection for the utilization of a broad range of alternate energy sources in the absence of light.
    Description: This work was supported by King Abdullah University for Science and Technology Global Collaborative Partners (GCR) program.
    Repository Name: Woods Hole Open Access Server
    Type: Article
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  • 5
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    Comparison of bacterial communities in sands and water at beaches with bacterial water quality violations (2014)
    Public Library of Science
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    Publication Date: 2022-05-26
    Description: © The Author(s), 2014. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in PLoS One 9 (2014): e90815, doi:10.1371/journal.pone.0090815.
    Description: Recreational water quality, as measured by culturable fecal indicator bacteria (FIB), may be influenced by persistent populations of these bacteria in local sands or wrack, in addition to varied fecal inputs from human and/or animal sources. In this study, pyrosequencing was used to generate short sequence tags of the 16S hypervariable region ribosomal DNA from shallow water samples and from sand samples collected at the high tide line and at the intertidal water line at sites with and without FIB exceedance events. These data were used to examine the sand and water bacterial communities to assess the similarity between samples, and to determine the impact of water quality exceedance events on the community composition. Sequences belonging to a group of bacteria previously identified as alternative fecal indicators were also analyzed in relationship to water quality violation events. We found that sand and water samples hosted distinctly different overall bacterial communities, and there was greater similarity in the community composition between coastal water samples from two distant sites. The dissimilarity between high tide and intertidal sand bacterial communities, although more similar to each other than to water, corresponded to greater tidal range between the samples. Within the group of alternative fecal indicators greater similarity was observed within sand and water from the same site, likely reflecting the anthropogenic contribution at each beach. This study supports the growing evidence that community-based molecular tools can be leveraged to identify the sources and potential impact of fecal pollution in the environment, and furthermore suggests that a more diverse bacterial community in beach sand and water may reflect a less contaminated site and better water quality.
    Description: This work was supported by the National Science Foundation grant OCE-0430724, and the National Institute of Environmental Health Sciences grant P0ES012742 to the Woods Hole Center for Ocean and Human Health. E. Halliday was partially supported by WHOI Academic Programs and grants from the WHOI Ocean Ventures Fund and the WHOI Coastal Ocean Institute.
    Repository Name: Woods Hole Open Access Server
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  • 6
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    Analysis of lung microbiota in bronchoalveolar lavage, protected brush and sputum samples from subjects with mild-to-moderate cystic fibrosis Lung Disease (2016)
    Public Library of Science
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    Publication Date: 2022-05-26
    Description: © The Author(s), 2016. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in PLoS 11 (2016): e0149998, doi:10.1371/journal.pone.0149998.
    Description: Individuals with cystic fibrosis (CF) often acquire chronic lung infections that lead to irreversible damage. We sought to examine regional variation in the microbial communities in the lungs of individuals with mild-to-moderate CF lung disease, to examine the relationship between the local microbiota and local damage, and to determine the relationships between microbiota in samples taken directly from the lung and the microbiota in spontaneously expectorated sputum. In this initial study, nine stable, adult CF patients with an FEV1〉50% underwent regional sampling of different lobes of the right lung by bronchoalveolar lavage (BAL) and protected brush (PB) sampling of mucus plugs. Sputum samples were obtained from six of the nine subjects immediately prior to the procedure. Microbial community analysis was performed on DNA extracted from these samples and the extent of damage in each lobe was quantified from a recent CT scan. The extent of damage observed in regions of the right lung did not correlate with specific microbial genera, levels of community diversity or composition, or bacterial genome copies per ml of BAL fluid. In all subjects, BAL fluid from different regions of the lung contained similar microbial communities. In eight out of nine subjects, PB samples from different regions of the lung were also similar in microbial community composition, and were similar to microbial communities in BAL fluid from the same lobe. Microbial communities in PB samples were more diverse than those in BAL samples, suggesting enrichment of some taxa in mucus plugs. To our knowledge, this study is the first to examine the microbiota in different regions of the CF lung in clinically stable individuals with mild-to-moderate CF-related lung disease.
    Description: Support from the Cystic Fibrosis Foundation Research Development Program (STANTO07R0) as a pilot grant to DAH and AA. Research reported in this publication was also supported by grants from the National Institutes of Health to DAH (R01 AI091702 to DAH) and the American Asthma Foundation Scholars Award and CFFT-ASHARE15A0 and R01HL122372 to AA and R01 HL074175 (BAS). The Dartmouth Lung Biology Center and CF Translational Research Core was supported by an Institutional Development Award (IDeA) from the National Institute of General Medical Sciences of the National Institutes of Health under grant number P30GM106394 and by the CFF RDP (CFRDP STANTO11R0).
    Repository Name: Woods Hole Open Access Server
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  • 7
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    The pervasive effects of an antibiotic on the human gut microbiota, as revealed by deep 16S rRNA sequencing (2008)
    Public Library of Science
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    Publication Date: 2022-05-26
    Description: © 2008 Author et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. The definitive version was published in PLoS Biology 6 (2008): e280, doi:10.1371/journal.pbio.0060280.
    Description: The intestinal microbiota is essential to human health, with effects on nutrition, metabolism, pathogen resistance, and other processes. Antibiotics may disrupt these interactions and cause acute disease, as well as contribute to chronic health problems, although technical challenges have hampered research on this front. Several recent studies have characterized uncultured and complex microbial communities by applying a new, massively parallel technology to obtain hundreds of thousands of sequences of a specific variable region within the small subunit rRNA gene. These shorter sequences provide an indication of diversity. We used this technique to track changes in the intestinal microbiota of three healthy humans before and after treatment with the antibiotic ciprofloxacin, with high sensitivity and resolution, and without sacrificing breadth of coverage. Consistent with previous results, we found that the microbiota of these individuals was similar at the genus level, but interindividual differences were evident at finer scales. Ciprofloxacin reduced the diversity of the intestinal microbiota, with significant effects on about one-third of the bacterial taxa. Despite this pervasive disturbance, the membership of the communities had largely returned to the pretreatment state within 4 weeks.
    Repository Name: Woods Hole Open Access Server
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  • 8
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    Evolutionary strategies of viruses, bacteria and archaea in hydrothermal vent ecosystems revealed through metagenomics (2014)
    Public Library of Science
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    Publication Date: 2022-05-26
    Description: © The Author(s), 2014. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in PLoS One 9 (2014): e109696, doi:10.1371/journal.pone.0109696.
    Description: The deep-sea hydrothermal vent habitat hosts a diverse community of archaea and bacteria that withstand extreme fluctuations in environmental conditions. Abundant viruses in these systems, a high proportion of which are lysogenic, must also withstand these environmental extremes. Here, we explore the evolutionary strategies of both microorganisms and viruses in hydrothermal systems through comparative analysis of a cellular and viral metagenome, collected by size fractionation of high temperature fluids from a diffuse flow hydrothermal vent. We detected a high enrichment of mobile elements and proviruses in the cellular fraction relative to microorganisms in other environments. We observed a relatively high abundance of genes related to energy metabolism as well as cofactors and vitamins in the viral fraction compared to the cellular fraction, which suggest encoding of auxiliary metabolic genes on viral genomes. Moreover, the observation of stronger purifying selection in the viral versus cellular gene pool suggests viral strategies that promote prolonged host integration. Our results demonstrate that there is great potential for hydrothermal vent viruses to integrate into hosts, facilitate horizontal gene transfer, and express or transfer genes that manipulate the hosts’ functional capabilities.
    Description: Funding for sequencing of the viral metagenome was provided by the Gordon and Betty Moore Foundation. All other funding was provided by a NASA Astrobiology Institute grant through Cooperative Agreement NNA04CC09A to the Geophysical Laboratory at the Carnegie Institution for Science. R.A. was funded by a NSF Graduate Research Fellowship through NSF grant number DGE-0718124, an NSF IGERT grant to the University of Washington Astrobiology Program, and the ARCS Foundation.
    Repository Name: Woods Hole Open Access Server
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