Engineering topics

Rinke, Christian

Publications and source records attributed to Rinke, Christian.

14 records

DOE OSTI2018

Corrigendum: Minimum information about a single amplified genome (MISAG) and a metagenome-assembled genome (MIMAG) of bacteria and archaea.

© 2018 Nature Publishing Group. All rights reserved. In the version of this article initially published, the author A. Murat Eren was listed as A.M. Eren. The corresponding affiliation was given as the Knapp Center for Biomedical Discovery, rather than Department of Medicine, University of Chicago, Chicago, Illinois, USA, and Marine Biological Laboratory, Woods Hole, Massachusetts, USA. The errors have been corrected in the HTML and PDF versions of the article as of 29 November 2017. In the version of this article initially published, the following acknowledgment was omitted: A.L. was supported by the Russian Science Foundation (grant number 14-50-00069). The error has been corrected in the HTML and PDF versions of the article as of 7 December 2017.

Bowers, Robert M↗

DOE OSTI2017

Diverse Marinimicrobia bacteria may mediate coupled biogeochemical cycles along eco-thermodynamic gradients

Microbial communities drive biogeochemical cycles through networks of metabolite exchange that are structured along energetic gradients. As energy yields become limiting, these networks favor co-metabolic interactions to maximize energy disequilibria. Here we apply single-cell genomics, metagenomics, and metatranscriptomics to study bacterial populations of the abundant “microbial dark matter” phylum Marinimicrobia along defined energy gradients. We show that evolutionary diversification of major Marinimicrobia clades appears to be closely related to energy yields, with increased co-metabolic interactions in more deeply branching clades. Several of these clades appear to participate in the biogeochemical cycling of sulfur and nitrogen, filling previously unassigned niches in the ocean. Notably, two Marinimicrobia clades, occupying different energetic niches, express nitrous oxide reductase, potentially acting as a global sink for the greenhouse gas nitrous oxide.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2017

Recovery of nearly 8,000 metagenome-assembled genomes substantially expands the tree of life

Challenges in cultivating microorganisms have limited the phylogenetic diversity of currently available microbial genomes. This is being addressed by advances in sequencing throughput and computational techniques that allow for the cultivation-independent recovery of genomes from metagenomes. Here, we report the reconstruction of 7,903 bacterial and archaeal genomes from >1,500 public metagenomes. All genomes are estimated to be ≥50% complete and nearly half are ≥90% complete with ≤5% contamination. These genomes increase the phylogenetic diversity of bacterial and archaeal genome trees by >30% and provide the first representatives of 17 bacterial and three archaeal candidate phyla. We also recovered 245 genomes from the Patescibacteria superphylum (also known as the Candidate Phyla Radiation) and find that the relative diversity of this group varies substantially with different protein marker sets. The scale and quality of this data set demonstrate that recovering genomes from metagenomes provides an expedient path forward to exploring microbial dark matter.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2017

Minimum information about a single amplified genome (MISAG) and a metagenome-assembled genome (MIMAG) of bacteria and archaea

Here, we present two standards developed by the Genomic Standards Consortium (GSC) for reporting bacterial and archaeal genome sequences. Both are extensions of the Minimum Information about Any (x) Sequence (MIxS). The standards are the Minimum Information about a Single Amplified Genome (MISAG) and the Minimum Information about a MetagenomeAssembled Genome (MIMAG), including, but not limited to, assembly quality, and estimates of genome completeness and contamination. These standards can be used in combination with other GSC checklists, including the Minimum Information about a Genome Sequence (MIGS), Minimum Information about a Metagenomic Sequence (MIMS), and Minimum Information about a Marker Gene Sequence (MIMARKS). Community-wide adoption of MISAG and MIMAG will facilitate more robust comparative genomic analyses of bacterial and archaeal diversity.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2017

Corrigendum: Minimum information about a single amplified genome (MISAG) and a metagenome-assembled genome (MIMAG) of bacteria and archaea

Here, we present two standards developed by the Genomic Standards Consortium (GSC) for reporting bacterial and archaeal genome sequences. Both are extensions of the Minimum Information about Any (x) Sequence (MIxS). The standards are the Minimum Information about a Single Amplified Genome (MISAG) and the Minimum Information about a Metagenome- Assembled Genome (MIMAG), including, but not limited to, assembly quality, and estimates of genome completeness and contamination. These standards can be used in combination with other GSC checklists, including the Minimum Information about a Genome Sequence (MIGS), Minimum Information about a Metagenomic Sequence (MIMS), and Minimum Information about a Marker Gene Sequence (MIMARKS). Community-wide adoption of MISAG and MIMAG will facilitate more robust comparative genomic analyses of bacterial and archaeal diversity.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2017

Comparative Genomic Analysis of the Class Epsilonproteobacteria and Proposed Reclassification to Epsilonbacteraeota (phyl. nov.)

The Epsilonproteobacteria is the fifth validly described class of the phylum Proteobacteria, known primarily for clinical relevance and for chemolithotrophy in various terrestrial and marine environments, including deep-sea hydrothermal vents. As 16S rRNA gene repositories have expanded and protein marker analysis become more common, the phylogenetic placement of this class has become less certain. A number of recent analyses of the bacterial tree of life using both 16S rRNA and concatenated marker gene analyses have failed to recover the Epsilonproteobacteria as monophyletic with all other classes of Proteobacteria. In order to address this issue, we investigated the phylogenetic placement of this class in the bacterial domain using 16S and 23S rRNA genes, as well as 120 single-copy marker proteins. Singleand concatenated-marker trees were created using a data set of 4,170 bacterial representatives, including 98 Epsilonproteobacteria. Phylogenies were inferred under a variety of tree building methods, with sequential jackknifing of outgroup phyla to ensure robustness of phylogenetic affiliations under differing combinations of bacterial genomes. Based on the assessment of nearly 300 phylogenetic tree topologies, we conclude that the continued inclusion of Epsilonproteobacteria within the Proteobacteria is not warranted, and that this group should be reassigned to a novel phylum for which we propose the name Epsilonbacteraeota (phyl. nov.). We further recommend the reclassification of the order Desulfurellales (Deltaproteobacteria) to a novel class within this phylum and a number of subordinate changes to ensure consistency with the genome-based phylogeny. Phylogenomic analysis of 658 genomes belonging to the newly proposed Epsilonbacteraeota suggests that the ancestor of this phylum was an autotrophic, motile, thermophilic chemolithotroph that likely assimilated nitrogen from ammonium taken up from the environment or generated from environmental nitrate and nitrite by employing a variety of functional redox modules. The emergence of chemoorganoheterotrophic lifestyles in several Epsilonbacteraeota families is the result of multiple independent losses of various ancestral chemolithoautotrophic pathways. Our proposed reclassification of this group resolves an important anomaly in bacterial systematics and ensures that the taxonomy of Proteobacteria remains robust, specifically as genome-based taxonomies become more common.

evolution↗

DOE OSTI2017

Genomic Analysis of Caldithrix abyssi , the Thermophilic Anaerobic Bacterium of the Novel Bacterial Phylum Calditrichaeota

The genome of Caldithrix abyssi , the first cultivated representative of a phylum-level bacterial lineage, was sequenced within the framework of Genomic Encyclopedia of Bacteria and Archaea (GEBA) project. The genomic analysis revealed mechanisms allowing this anaerobic bacterium to ferment peptides or to implement nitrate reduction with acetate or molecular hydrogen as electron donors. The genome encoded five different [NiFe]- and [FeFe]-hydrogenases, one of which, group 1 [NiFe]-hydrogenase, is presumably involved in lithoheterotrophic growth, three other produce H 2 during fermentation, and one is apparently bidirectional. The ability to reduce nitrate is determined by a nitrate reductase of the Nap family, while nitrite reduction to ammonia is presumably catalyzed by an octaheme cytochrome c nitrite reductase εHao. The genome contained genes of respiratory polysulfide/thiosulfate reductase, however, elemental sulfur and thiosulfate were not used as the electron acceptors for anaerobic respiration with acetate or H 2 , probably due to the lack of the gene of the maturation protein. Nevertheless, elemental sulfur and thiosulfate stimulated growth on fermentable substrates (peptides), being reduced to sulfide, most probably through the action of the cytoplasmic sulfide dehydrogenase and/or NAD(P)-dependent [NiFe]-hydrogenase (sulfhydrogenase) encoded by the genome. Surprisingly, the genome of this anaerobic microorganism encoded all genes for cytochrome c oxidase, however, its maturation machinery seems to be non-operational due to genomic rearrangements of supplementary genes. Despite the fact that sugars were not among the substrates reported when C. abyssi was first described, our genomic analysis revealed multiple genes of glycoside hydrolases, and some of them were predicted to be secreted. This finding aided in bringing out four carbohydrates that supported the growth of C. abyssi : starch, cellobiose, glucomannan and xyloglucan. The genomic analysis demonstrated the ability of C. abyssi to synthesize nucleotides and most amino acids and vitamins. Finally, the genomic sequence allowed us to perform a phylogenomic analysis, based on 38 protein sequences, which confirmed the deep branching of this lineage and justified the proposal of a novel phylum Calditrichaeota .

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2016

acdc – Automated Contamination Detection and Confidence estimation for single-cell genome data

Abstract Background A major obstacle in single-cell sequencing is sample contamination with foreign DNA. To guarantee clean genome assemblies and to prevent the introduction of contamination into public databases, considerable quality control efforts are put into post-sequencing analysis. Contamination screening generally relies on reference-based methods such as database alignment or marker gene search, which limits the set of detectable contaminants to organisms with closely related reference species. As genomic coverage in the tree of life is highly fragmented, there is an urgent need for a reference-free methodology for contaminant identification in sequence data. Results We present acdc, a tool specifically developed to aid the quality control process of genomic sequence data. By combining supervised and unsupervised methods, it reliably detects both known and de novo contaminants. First, 16S rRNA gene prediction and the inclusion of ultrafast exact alignment techniques allow sequence classification using existing knowledge from databases. Second, reference-free inspection is enabled by the use of state-of-the-art machine learning techniques that include fast, non-linear dimensionality reduction of oligonucleotide signatures and subsequent clustering algorithms that automatically estimate the number of clusters. The latter also enables the removal of any contaminant, yielding a clean sample. Furthermore, given the data complexity and the ill-posedness of clustering, acdc employs bootstrapping techniques to provide statistically profound confidence values. Tested on a large number of samples from diverse sequencing projects, our software is able to quickly and accurately identify contamination. Results are displayed in an interactive user interface. Acdc can be run from the web as well as a dedicated command line application, which allows easy integration into large sequencing project analysis workflows. Conclusions Acdc can reliably detect contamination in single-cell genome data. In addition to database-driven detection, it complements existing tools by its unsupervised techniques, which allow for the detection of de novo contaminants. Our contribution has the potential to drastically reduce the amount of resources put into these processes, particularly in the context of limited availability of reference species. As single-cell genome data continues to grow rapidly, acdc adds to the toolkit of crucial quality assurance tools.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2016

Validation of picogram- and femtogram-input DNA libraries for microscale metagenomics

High-throughput sequencing libraries are typically limited by the requirement for nanograms to micrograms of input DNA. This bottleneck impedes the microscale analysis of ecosystems and the exploration of low biomass samples. Current methods for amplifying environmental DNA to bypass this bottleneck introduce considerable bias into metagenomic profiles. For this study, we describe and validate a simple modification of the Illumina Nextera XT DNA library preparation kit which allows creation of shotgun libraries from sub-nanogram amounts of input DNA. Community composition was reproducible down to 100 fg of input DNA based on analysis of a mock community comprising 54 phylogenetically diverse Bacteria and Archaea. The main technical issues with the low input libraries were a greater potential for contamination, limited DNA complexity which has a direct effect on assembly and binning, and an associated higher percentage of read duplicates. We recommend a lower limit of 1 pg (~100–1,000 microbial cells) to ensure community composition fidelity, and the inclusion of negative controls to identify reagent-specific contaminants. Applying the approach to marine surface water, pronounced differences were observed between bacterial community profiles of microliter volume samples, which we attribute to biological variation. This result is consistent with expected microscale patchiness in marine communities. We thus envision that our benchmarked, slightly modified low input DNA protocol will be beneficial for microscale and low biomass metagenomics.

100 fg↗

DOE OSTI2015

ProDeGe: A computational protocol for fully automated decontamination of genomes

Single amplified genomes and genomes assembled from metagenomes have enabled the exploration of uncultured microorganisms at an unprecedented scale. However, both these types of products are plagued by contamination. Since these genomes are now being generated in a high-throughput manner and sequences from them are propagating into public databases to drive novel scientific discoveries, rigorous quality controls and decontamination protocols are urgently needed. Here, we present ProDeGe (Protocol for fully automated Decontamination of Genomes), the first computational protocol for fully automated decontamination of draft genomes. ProDeGe classifies sequences into two classes—clean and contaminant—using a combination of homology and feature-based methodologies. On average, 84% of sequence from the non-target organism is removed from the data set (specificity) and 84% of the sequence from the target organism is retained (sensitivity). Lastly, the procedure operates successfully at a rate of ~0.30 CPU core hours per megabase of sequence and can be applied to any type of genome sequence.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2015

In Silico Analysis of the Metabolic Potential and Niche Specialization of Candidate Phylum " Latescibacteria " (WS3)

The “Latescibacteria” (formerly WS3), member of the Fibrobacteres–Chlorobi–Bacteroidetes (FCB) superphylum, represents a ubiquitous candidate phylum found in terrestrial, aquatic, and marine ecosystems. Recently, single-cell amplified genomes (SAGs) representing the “Latescibacteria” were obtained from the anoxic monimolimnion layers of Sakinaw Lake (British Columbia, Canada), and anoxic sediments of a coastal lagoon (Etoliko lagoon, Western Greece). Here, we present a detailed in-silico analysis of the four SAGs to gain some insights on their metabolic potential and apparent ecological roles. Metabolic reconstruction suggests an anaerobic fermentative mode of metabolism, as well as the capability to degrade multiple polysaccharides and glycoproteins that represent integral components of green (Charophyta and Chlorophyta) and brown (Phaeophycaea) algae cell walls (pectin, alginate, ulvan, fucan, hydroxyproline-rich glycoproteins), storage molecules (starch and trehalose), and extracellular polymeric substances (EPSs). The analyzed SAGs also encode dedicated transporters for the uptake of produced sugars and amino acids/oligopeptides, as well as an extensive machinery for the catabolism of all transported sugars, including the production of a bacterial microcompartment (BMC) to sequester propionaldehyde, a toxic intermediate produced during fucose and rhamnose metabolism. Finally, genes for the formation of gas vesicles, flagella, type IV pili, and oxidative stress response were found, features that could aid in cellular association with algal detritus. Collectively, these results indicate that the analyzed “Latescibacteria” mediate the turnover of multiple complex organic polymers of algal origin that reach deeper anoxic/microoxic habitats in lakes and lagoons. The implications of such process on our understanding of niche specialization in microbial communities mediating organic carbon turnover in stratified water bodies are discussed.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2015

Reconstructing each cell's genome within complex microbial communitiesâ€”dream or reality?

As the vast majority of microorganisms have yet to be cultivated in a laboratory setting, access to their genetic makeup has largely been limited to cultivation-independent methods. These methods, namely metagenomics and more recently single-cell genomics, have become cornerstones for microbial ecology and environmental microbiology. One ultimate goal is the recovery of genome sequences from each cell within an environment to move toward a better understanding of community metabolic potential and to provide substrate for experimental work. As single-cell sequencing has the ability to decipher all sequence information contained in an individual cell, this method holds great promise in tackling such challenge. Methodological limitations and inherent biases however do exist, which will be discussed here based on environmental and benchmark data, to assess how far we are from reaching this goal.

microbial ecology↗

DOE OSTI2014

Stop Codon Reassignment in the Wild

Since the discovery of the genetic code and protein translation mechanisms (1), a limited number of variations of the standard assignment between unique base triplets (codons) and their encoded amino acids and translational stop signals have been found in bacteria and phages (2-3). Given the apparent ubiquity of the canonical genetic code, the design of genomically recoded organisms with non-canonical codes has been suggested as a means to prevent horizontal gene transfer between laboratory and environmental organisms (4). It is also predicted that genomically recoded organisms are immune to infection by viruses, under the assumption that phages and their hosts must share a common genetic code (5). This paradigm is supported by the observation of increased resistance of genomically recoded bacteria to phages with a canonical code (4). Despite these assumptions and accompanying lines of evidence, it remains unclear whether differential and non-canonical codon usage represents an absolute barrier to phage infection and genetic exchange between organisms. Our knowledge of the diversity of genetic codes and their use by viruses and their hosts is primarily derived from the analysis of cultivated organisms. Advances in single-cell sequencing and metagenome assembly technologies have enabled the reconstruction of genomes of uncultivated bacterial and archaeal lineages (6). These initial findings suggest that large scale systematic studies of uncultivated microorganisms and viruses may reveal the extent and modes of divergence from the canonical genetic code operating in nature. To explore alternative genetic codes, we carried out a systematic analysis of stop codon reassignments from the canonical TAG amber, TGA opal, and TAA ochre codons in assembled metagenomes from environmental and host-associated samples, single-cell genomes of uncultivated bacteria and archaea, and a collection of phage sequences

09 BIOMASS FUELS↗

DOE OSTI2014

The Candidate Phylum Poribacteria by Single-Cell Genomics: New Insights into Phylogeny, Cell-Compartmentation, Eukaryote-Like Repeat Proteins, and Other Genomic Features

The candidate phylum Poribacteria is one of the most dominant and widespread members of the microbial communities residing within marine sponges. Cell compartmentalization had been postulated along with their discovery about a decade ago and their phylogenetic association to the Planctomycetes, Verrucomicrobia, Chlamydiae superphylum was proposed soon thereafter. In the present study we revised these features based on genomic data obtained from six poribacterial single cells. We propose that Poribacteria form a distinct monophyletic phylum contiguous to the PVC superphylum together with other candidate phyla. Our genomic analyses supported the possibility of cell compartmentalization in form of bacterial microcompartments. Further analyses of eukaryote-like protein domains stressed the importance of such proteins with features including tetratricopeptide repeats, leucin rich repeats as well as low density lipoproteins receptor repeats, the latter of which are reported here for the first time from a sponge symbiont. Finally, examining the most abundant protein domain family on poribacterial genomes revealed diverse phyH family proteins, some of which may be related to dissolved organic posphorus uptake.

59 BASIC BIOLOGICAL SCIENCES↗