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Brandt B.W.,VU University Amsterdam | Bonder M.J.,VU University Amsterdam | Huse S.M.,Josephine Bay Paul Center for Comparative Molecular Biology and Evolution | Zaura E.,VU University Amsterdam
Nucleic Acids Research | Year: 2012

Amplicon sequencing of the hypervariable regions of the small subunit ribosomal RNA gene is a widely accepted method for identifying the members of complex bacterial communities. Several rRNA gene sequence reference databases can be used to assign taxonomic names to the sequencing reads using BLAST, USEARCH, GAST or the RDP classifier. Next-generation sequencing methods produce ample reads, but they are short, currently ∼100-450nt (depending on the technology), as compared to the full rRNA gene of ∼1550nt. It is important, therefore, to select the right rRNA gene region for sequencing. The primers should amplify the species of interest and the hypervariable regions should differentiate their taxonomy. Here, we introduce TaxMan: a web-based tool that trims reference sequences based on user-selected primer pairs and returns an assessment of the primer specificity by taxa. It allows interactive plotting of taxa, both amplified and missed in silico by the primers used. Additionally, using the trimmed sequences improves the speed of sequence matching algorithms. The smaller database greatly improves run times (up to 98) and memory usage, not only of similarity searching (BLAST), but also of chimera checking (UCHIME) and of clustering the reads (UCLUST). TaxMan is available at http://www.ibi.vu.nl/ programs/taxmanwww/. © 2012 The Author(s).

Vick-Majors T.J.,Montana State University | Priscu J.C.,Josephine Bay Paul Center for Comparative Molecular Biology and Evolution | Amaral-Zettler L.A.,Brown University
ISME Journal | Year: 2014

High-latitude environments, such as the Antarctic McMurdo Dry Valley lakes, are subject to seasonally segregated light-dark cycles, which have important consequences for microbial diversity and function on an annual basis. Owing largely to the logistical difficulties of sampling polar environments during the darkness of winter, little is known about planktonic microbial community responses to the cessation of photosynthetic primary production during the austral sunset, which lingers from approximately February to April. Here, we hypothesized that changes in bacterial, archaeal and eukaryotic community structure, particularly shifts in favor of chemolithotrophs and mixotrophs, would manifest during the transition to polar night. Our work represents the first concurrent molecular characterization, using 454 pyrosequencing of hypervariable regions of the small-subunit ribosomal RNA gene, of bacterial, archaeal and eukaryotic communities in permanently ice-covered lakes Fryxell and Bonney, before and during the polar night transition. We found vertically stratified populations that varied at the community and/or operational taxonomic unit-level between lakes and seasons. Network analysis based on operational taxonomic unit level interactions revealed nonrandomly structured microbial communities organized into modules (groups of taxa) containing key metabolic potential capacities, including photoheterotrophy, mixotrophy and chemolithotrophy, which are likely to be differentially favored during the transition to polar night. © 2014 International Society for Microbial Ecology All rights reserved.

Liebeskind B.J.,University of Texas at Austin | Hillis D.M.,University of Texas at Austin | Zakon H.H.,University of Texas at Austin | Zakon H.H.,Josephine Bay Paul Center for Comparative Molecular Biology and Evolution
Proceedings of the National Academy of Sciences of the United States of America | Year: 2015

Multicellularity has evolved multiple times, but animals are the only multicellular lineage with nervous systems. This fact implies that the origin of nervous systems was an unlikely event, yet recent comparisons among extant taxa suggest that animal nervous systems may have evolved multiple times independently. Here, we use ancestral gene content reconstruction to track the timing of gene family expansions for the major families of ion-channel proteins that drive nervous system function. We find that animals with nervous systems have broadly similar complements of ionchannel types but that these complements likely evolved independently. We also find that ion-channel gene family evolution has included large loss events, two of which were immediately followed by rounds of duplication. Ctenophores, cnidarians, and bilaterians underwent independent bouts of gene expansion in channel families involved in synaptic transmission and action potential shaping. We suggest that expansions of these family types may represent a genomic signature of expanding nervous system complexity. Ancestral nodes in which nervous systems are currently hypothesized to have originated did not experience large expansions, making it difficult to distinguish among competing hypotheses of nervous system origins and suggesting that the origin of nerves was not attended by an immediate burst of complexity. Rather, the evolution of nervous system complexity appears to resemble a slow fuse in stem animals followed by many independent bouts of gene gain and loss.

Amaral-Zettler L.A.,Josephine Bay Paul Center for Comparative Molecular Biology and Evolution | Amaral-Zettler L.A.,Brown University
Frontiers in Microbiology | Year: 2012

Extremely acidic (pH < 3) and extremely alkaline (pH > 9) environments support a diversity of single-cell and to a lesser extent, multicellular eukaryotic life. This study compared alpha and beta diversity in eukaryotic communities from seven diverse aquatic environments with pH values ranging from 2 to 11 using massively-parallel pyrotag sequencing targeting the V9 hypervariable region of the 18S ribosomal RNA (rRNA) gene. A total of 946 operational taxonomic units (OTUs) were recovered at a 6% cut-off level (94% similarity) across the sampled environments. Hierarchical clustering of the samples segregated the communities into acidic and alkaline groups. Similarity percentage (SIMPER) analysis followed by indicator OTU analysis (IOA) and non-metric multidimensional scaling (NMDS) were used to determine which characteristic groups of eukaryotic taxa typify acidic or alkaline extremes and the extent to which pH explains eukaryotic community structure in these environments. Spain's Rio Tinto yielded the fewest observed OTUs while Nebraska Sandhills alkaline lakes yielded the most. Distinct OTUs, including metazoan OTUs, numerically dominated pH extreme sites. Indicator OTUs included the diatom Pinnularia and unidentified opisthokonts (Fungi and Filasterea) in the extremely acidic environments, and the ciliate Frontonia across the extremely alkaline sites. Inferred from NMDS, pH explained only a modest fraction of the variation across the datasets, indicating that other factors influence the underlying community structure in these environments. The findings from this study suggest that the ability for eukaryotes to adapt to pH extremes over a broad range of values may be rare, but further study of taxa that can broadly adapt across diverse acidic and alkaline environments, respectively present good models for understanding adaptation and should be targeted for future investigations. © 2013 Amaral-Zettler.

Akerman N.H.,Josephine Bay Paul Center for Comparative Molecular Biology and Evolution | Butterfield D.A.,University of Washington | Huber J.A.,Josephine Bay Paul Center for Comparative Molecular Biology and Evolution
Frontiers in Microbiology | Year: 2013

Microorganisms throughout the dark ocean use reduced sulfur compounds for chemolithoautotrophy. In many deep-sea hydrothermal vents, sulfide oxidation is quantitatively the most important chemical energy source for microbial metabolism both at and beneath the seafloor. In this study, the presence and activity of vent endemic Epsilonproteobacteria was examined in six low-temperature diffuse vents over a range of geochemical gradients from Axial Seamount, a deep-sea volcano in the Northeast Pacific. PCR primers were developed and applied to target the sulfur oxidation soxB gene of Epsilonproteobacteria. soxB genes belonging to the genera Sulfurimonas and Sulfurovum are both present and expressed at most diffuse vent sites, but not in background seawater. Although Sulfurovum-like soxB genes were detected in all fluid samples, the RNA profiles were nearly identical among the vents and suggest that Sulfurimonas-like species are the primary Epsilonproteobacteriaresponsible for actively oxidizing sulfur via the Sox pathway at each vent. Community patterns of subseafloor Epsilonproteobacteria 16S rRNA genes werebest matched to methane concentrations in vent fluids, as well as individual vent locations, indicating that both geochemistry and geographical isolation play a role in structuring subseafloor microbial populations. The data show that in the subseafloor at Axial Seamount, Epsilonproteobacteria are expressing the soxB gene and that microbial patterns in community distribution are linked to both vent location and chemistry. ©2013 Akerman, Butterfield and Huber.

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