Whole community shotgun metagenomes of two biological soil crust types from the Mojave Desert.

We present six whole community shotgun metagenomic sequencing data sets of two types of biological soil crusts sampled at the ecotone of the Mojave Desert and Colorado Desert in California. These data will help us understand the diversity and function of biocrust microbial communities, which are essential for desert ecosystems.

Metatranscriptomes of two biological soil crust types from the Mojave desert in response to wetting.

We present eight metatranscriptomic datasets of light algal and cyanolichen biological soil crusts from the Mojave Desert in response to wetting. These data will help us understand gene expression patterns in desert biocrust microbial communities after they have been reactivated by the addition of water.

Metatranscriptomics sheds light on the links between the functional traits of fungal guilds and ecological processes in forest soil ecosystems.

Soil fungi belonging to different functional guilds, such as saprotrophs, pathogens, and mycorrhizal symbionts, play key roles in forest ecosystems. To date, no study has compared the actual gene expression of these guilds in different forest soils. We used metatranscriptomics to study the competition for organic resources by these fungal groups in boreal, temperate, and Mediterranean forest soils. Using a dedicated mRNA annotation pipeline combined with the JGI MycoCosm database, we compared the transcripts of these three fungal guilds, targeting enzymes involved in C- and N mobilization from plant and microbial cell walls. Genes encoding enzymes involved in the degradation of plant cell walls were expressed at a higher level in saprotrophic fungi than in ectomycorrhizal and pathogenic fungi. However, ectomycorrhizal and saprotrophic fungi showed similarly high expression levels of genes encoding enzymes involved in fungal cell wall degradation. Transcripts for N-related transporters were more highly expressed in ectomycorrhizal fungi than in other groups. We showed that ectomycorrhizal and saprotrophic fungi compete for N in soil organic matter, suggesting that their interactions could decelerate C cycling. Metatranscriptomics provides a unique tool to test controversial ecological hypotheses and to better understand the underlying ecological processes involved in soil functioning and carbon stabilization.

Multiomics in the central Arctic Ocean for benchmarking biodiversity change.

Multiomics approaches need to be applied in the central Arctic Ocean to benchmark biodiversity change and to identify novel species and their genes. As part of MOSAiC, EcoOmics will therefore be essential for conservation and sustainable bioprospecting in one of the least explored ecosystems on Earth.

Metagenome Sequencing to Explore Phylogenomics of Terrestrial Cyanobacteria.

Cyanobacteria are ubiquitous microorganisms with crucial ecosystem functions, yet most knowledge of their biology relates to aquatic taxa. We have constructed metagenomes for 50 taxonomically well-characterized terrestrial cyanobacterial cultures. These data will support phylogenomic studies of evolutionary relationships and gene content among these unique algae and their aquatic relatives.

DOE JGI Metagenome Workflow.

The DOE Joint Genome Institute (JGI) Metagenome Workflow performs metagenome data processing, including assembly; structural, functional, and taxonomic annotation; and binning of metagenomic data sets that are subsequently included into the Integrated Microbial Genomes and Microbiomes (IMG/M) (I.-M. A. Chen, K. Chu, K. Palaniappan, A. Ratner, et al., Nucleic Acids Res, 49:D751-D763, 2021, https://doi.org/10.1093/nar/gkaa939) comparative analysis system and provided for download via the JGI data portal (https://genome.jgi.doe.gov/portal/). This workflow scales to run on thousands of metagenome samples per year, which can vary by the complexity of microbial communities and sequencing depth. Here, we describe the different tools, databases, and parameters used at different steps of the workflow to help with the interpretation of metagenome data available in IMG and to enable researchers to apply this workflow to their own data. We use 20 publicly available sediment metagenomes to illustrate the computing requirements for the different steps and highlight the typical results of data processing. The workflow modules for read filtering and metagenome assembly are available as a workflow description language (WDL) file (https://code.jgi.doe.gov/BFoster/jgi_meta_wdl). The workflow modules for annotation and binning are provided as a service to the user community at https://img.jgi.doe.gov/submit and require filling out the project and associated metadata descriptions in the Genomes OnLine Database (GOLD) (S. Mukherjee, D. Stamatis, J. Bertsch, G. Ovchinnikova, et al., Nucleic Acids Res, 49:D723-D733, 2021, https://doi.org/10.1093/nar/gkaa983).IMPORTANCE The DOE JGI Metagenome Workflow is designed for processing metagenomic data sets starting from Illumina fastq files. It performs data preprocessing, error correction, assembly, structural and functional annotation, and binning. The results of processing are provided in several standard formats, such as fasta and gff, and can be used for subsequent integration into the Integrated Microbial Genomes and Microbiomes (IMG/M) system where they can be compared to a comprehensive set of publicly available metagenomes. As of 30 July 2020, 7,155 JGI metagenomes have been processed by the DOE JGI Metagenome Workflow. Here, we present a metagenome workflow developed at the JGI that generates rich data in standard formats and has been optimized for downstream analyses ranging from assessment of the functional and taxonomic composition of microbial communities to genome-resolved metagenomics and the identification and characterization of novel taxa. This workflow is currently being used to analyze thousands of metagenomic data sets in a consistent and standardized manner.

A single-cell genomics pipeline for environmental microbial eukaryotes.

Single-cell sequencing of environmental microorganisms is an essential component of the microbial ecology toolkit. However, large-scale targeted single-cell sequencing for the whole-genome recovery of uncultivated eukaryotes is lagging. The key challenges are low abundance in environmental communities, large complex genomes, and cell walls that are difficult to break. We describe a pipeline composed of state-of-the art single-cell genomics tools and protocols optimized for poorly studied and uncultivated eukaryotic microorganisms that are found at low abundance. This pipeline consists of seven distinct steps, beginning with sample collection and ending with genome annotation, each equipped with quality review steps to ensure high genome quality at low cost. We tested and evaluated each step on environmental samples and cultures of early-diverging lineages of fungi and Chromista/SAR. We show that genomes produced using this pipeline are almost as good as complete reference genomes for functional and comparative genomics for environmental microbial eukaryotes.

Proteome specialization of anaerobic fungi during ruminal degradation of recalcitrant plant fiber.

The rumen harbors a complex microbial mixture of archaea, bacteria, protozoa, and fungi that efficiently breakdown plant biomass and its complex dietary carbohydrates into soluble sugars that can be fermented and subsequently converted into metabolites and nutrients utilized by the host animal. While rumen bacterial populations have been well documented, only a fraction of the rumen eukarya are taxonomically and functionally characterized, despite the recognition that they contribute to the cellulolytic phenotype of the rumen microbiota. To investigate how anaerobic fungi actively engage in digestion of recalcitrant fiber that is resistant to degradation, we resolved genome-centric metaproteome and metatranscriptome datasets generated from switchgrass samples incubated for 48 h in nylon bags within the rumen of cannulated dairy cows. Across a gene catalog covering anaerobic rumen bacteria, fungi and viruses, a significant portion of the detected proteins originated from fungal populations. Intriguingly, the carbohydrate-active enzyme (CAZyme) profile suggested a domain-specific functional specialization, with bacterial populations primarily engaged in the degradation of hemicelluloses, whereas fungi were inferred to target recalcitrant cellulose structures via the detection of a number of endo- and exo-acting enzymes belonging to the glycoside hydrolase (GH) family 5, 6, 8, and 48. Notably, members of the GH48 family were amongst the highest abundant CAZymes and detected representatives from this family also included dockerin domains that are associated with fungal cellulosomes. A eukaryote-selected metatranscriptome further reinforced the contribution of uncultured fungi in the ruminal degradation of recalcitrant fibers. These findings elucidate the intricate networks of in situ recalcitrant fiber deconstruction, and importantly, suggest that the anaerobic rumen fungi contribute a specific set of CAZymes that complement the enzyme repertoire provided by the specialized plant cell wall degrading rumen bacteria.

Metagenomes and Metatranscriptomes of a Glucose-Amended Agricultural Soil.

The addition of glucose to soil has long been used to study the metabolic activity of microbes in soil; however, the response of the microbial ecophysiology remains poorly characterized. To address this, we sequenced the metagenomes and metatranscriptomes of glucose-amended soil microbial communities in a laboratory incubation.

Metagenomes from Experimental Hydrologic Manipulation of Restored Coastal Plain Wetland Soils (Tyrell County, North Carolina).

Hydrologic changes modify microbial community structure and ecosystem functions, especially in wetland systems. Here, we present 24 metagenomes from a coastal freshwater wetland experiment in which we manipulated hydrologic conditions and plant presence. These wetland soil metagenomes will deepen our understanding of how hydrology and vegetation influence microbial functional diversity.

Terabase-scale metagenome coassembly with MetaHipMer.

Metagenome sequence datasets can contain terabytes of reads, too many to be coassembled together on a single shared-memory computer; consequently, they have only been assembled sample by sample (multiassembly) and combining the results is challenging. We can now perform coassembly of the largest datasets using MetaHipMer, a metagenome assembler designed to run on supercomputers and large clusters of compute nodes. We have reported on the implementation of MetaHipMer previously; in this paper we focus on analyzing the impact of very large coassembly. In particular, we show that coassembly recovers a larger genome fraction than multiassembly and enables the discovery of more complete genomes, with lower error rates, whereas multiassembly recovers more dominant strain variation. Being able to coassemble a large dataset does not preclude one from multiassembly; rather, having a fast, scalable metagenome assembler enables a user to more easily perform coassembly and multiassembly, and assemble both abundant, high strain variation genomes, and low-abundance, rare genomes. We present several assemblies of terabyte datasets that could never be coassembled before, demonstrating MetaHipMer's scaling power. MetaHipMer is available for public use under an open source license and all datasets used in the paper are available for public download.

Metatranscriptomic Sequencing of a Cyanobacterial Soil-Surface Consortium with and without a Diverse Underlying Soil Microbiome.

Soil surface consortia are easily observed and sampled, allowing examination of their interactions with soil microbiomes. Here, we present metatranscriptomic sequences from Dark Green 1 (DG1), a cyanobacterium-based soil surface consortium, in the presence and absence of an underlying soil microbiome and/or urea.

Shotgun metagenomic analysis of microbial communities from the Loxahatchee nature preserve in the Florida Everglades.

BACKGROUND: Currently, much is unknown about the taxonomic diversity and the mechanisms of methane metabolism in the Florida Everglades ecosystem. The Loxahatchee National Wildlife Refuge is a section of the Florida Everglades that is almost entirely unstudied in regard to taxonomic profiling. This short report analyzes the metagenome of soil samples from this Refuge to investigate the predominant taxa, as well as the abundance of genes involved in environmentally significant metabolic pathways related to methane production (nitrogen fixation and dissimilatory sulfite reduction). METHODS: Shotgun metagenomic sequencing using the Illumina platform was performed on 17 soil samples from four different sites within the Loxahatchee National Wildlife Refuge, and underwent quality control, assembly, and annotation. The soil from each sample was tested for water content and concentrations of organic carbon and nitrogen. RESULTS: The three most common phyla of bacteria for every site were Actinobacteria, Acidobacteria, and Proteobacteria; however, there was variation in relative phylum composition. The most common phylum of Archaea was Euryarchaeota for all sites. Alpha and beta diversity analyses indicated significant congruity in taxonomic diversity in most samples from Sites 1, 3, and 4 and negligible congruity between Site 2 and the other sites. Shotgun metagenomic sequencing revealed the presence of biogeochemical biomarkers of particular interest (e.g., mrcA, nifH, and dsrB) within the samples. The normalized abundances of mcrA, nifH, and dsrB exhibited a positive correlation with nitrogen concentration and water content, and a negative correlation with organic carbon concentration. CONCLUSION: This Everglades soil metagenomic study allowed examination of wetlands biological processes and showed expected correlations between measured organic constituents and prokaryotic gene frequency. Additionally, the taxonomic profile generated gives a basis for the diversity of prokaryotic microbial life throughout the Everglades.

Shotgun metagenome data of a defined mock community using Oxford Nanopore, PacBio and Illumina technologies.

Metagenomic sequence data from defined mock communities is crucial for the assessment of sequencing platform performance and downstream analyses, including assembly, binning and taxonomic assignment. We report a comparison of shotgun metagenome sequencing and assembly metrics of a defined microbial mock community using the Oxford Nanopore Technologies (ONT) MinION, PacBio and Illumina sequencing platforms. Our synthetic microbial community BMock12 consists of 12 bacterial strains with genome sizes spanning 3.2-7.2 Mbp, 40-73% GC content, and 1.5-7.3% repeats. Size selection of both PacBio and ONT sequencing libraries prior to sequencing was essential to yield comparable relative abundances of organisms among all sequencing technologies. While the Illumina-based metagenome assembly yielded good coverage with few misassemblies, contiguity was greatly improved by both, Illumina + ONT and Illumina + PacBio hybrid assemblies but increased misassemblies, most notably in genomes with high sequence similarity to each other. Our resulting datasets allow evaluation and benchmarking of bioinformatics software on Illumina, PacBio and ONT platforms in parallel.

Metagenomes and metatranscriptomes from boreal potential and actual acid sulfate soil materials.

Natural sulfide rich deposits are common in coastal areas worldwide, including along the Baltic Sea coast. When artificial drainage exposes these deposits to atmospheric oxygen, iron sulfide minerals in the soils are rapidly oxidized. This process turns the potential acid sulfate soils into actual acid sulfate soils and mobilizes large quantities of acidity and leachable toxic metals that cause severe environmental problems. It is known that acidophilic microorganisms living in acid sulfate soils catalyze iron sulfide mineral oxidation. However, only a few studies regarding these communities have been published. In this study, we sampled the oxidized actual acid sulfate soil, the transition zone where oxidation is actively taking place, and the deepest un-oxidized potential acid sulfate soil. Nucleic acids were extracted and 16S rRNA gene amplicons, metagenomes, and metatranscriptomes generated to gain a detailed insight into the communities and their activities. The project will be of great use to microbiologists, environmental biologists, geochemists, and geologists as there is hydrological and geochemical monitoring from the site stretching back for many years.

Microbiomes of Velloziaceae from phosphorus-impoverished soils of the campos rupestres, a biodiversity hotspot.

The rocky, seasonally-dry and nutrient-impoverished soils of the Brazilian campos rupestres impose severe growth-limiting conditions on plants. Species of a dominant plant family, Velloziaceae, are highly specialized to low-nutrient conditions and seasonal water availability of this environment, where phosphorus (P) is the key limiting nutrient. Despite plant-microbe associations playing critical roles in stressful ecosystems, the contribution of these interactions in the campos rupestres remains poorly studied. Here we present the first microbiome data of Velloziaceae spp. thriving in contrasting substrates of campos rupestres. We assessed the microbiomes of Vellozia epidendroides, which occupies shallow patches of soil, and Barbacenia macrantha, growing on exposed rocks. The prokaryotic and fungal profiles were assessed by rRNA barcode sequencing of epiphytic and endophytic compartments of roots, stems, leaves and surrounding soil/rocks. We also generated root and substrate (rock/soil)-associated metagenomes of each plant species. We foresee that these data will contribute to decipher how the microbiome contributes to plant functioning in the campos rupestres, and to unravel new strategies for improved crop productivity in stressful environments.

Microbial metagenomes and metatranscriptomes during a coastal phytoplankton bloom.

Metagenomic and metatranscriptomic time-series data covering a 52-day period in the fall of 2016 provide an inventory of bacterial and archaeal community genes, transcripts, and taxonomy during an intense dinoflagellate bloom in Monterey Bay, CA, USA. The dataset comprises 84 metagenomes (0.8 terabases), 82 metatranscriptomes (1.1 terabases), and 88 16S rRNA amplicon libraries from samples collected on 41 dates. The dataset also includes 88 18S rRNA amplicon libraries, characterizing the taxonomy of the eukaryotic community during the bloom. Accompanying the sequence data are chemical and biological measurements associated with each sample. These datasets will facilitate studies of the structure and function of marine bacterial communities during episodic phytoplankton blooms.

Microbial Community Structure and Functional Potential in Cultivated and Native Tallgrass Prairie Soils of the Midwestern United States.

The North American prairie covered about 3.6 million-km2 of the continent prior to European contact. Only 1-2% of the original prairie remains, but the soils that developed under these prairies are some of the most productive and fertile in the world, containing over 35% of the soil carbon in the continental United States. Cultivation may alter microbial diversity and composition, influencing the metabolism of carbon, nitrogen, and other elements. Here, we explored the structure and functional potential of the soil microbiome in paired cultivated-corn (at the time of sampling) and never-cultivated native prairie soils across a three-states transect (Wisconsin, Iowa, and Kansas) using metagenomic and 16S rRNA gene sequencing and lipid analysis. At the Wisconsin site, we also sampled adjacent restored prairie and switchgrass plots. We found that agricultural practices drove differences in community composition and diversity across the transect. Microbial biomass in prairie samples was twice that of cultivated soils, but alpha diversity was higher with cultivation. Metagenome analyses revealed denitrification and starch degradation genes were abundant across all soils, as were core genes involved in response to osmotic stress, resource transport, and environmental sensing. Together, these data indicate that cultivation shifted the microbiome in consistent ways across different regions of the prairie, but also suggest that many functions are resilient to changes caused by land management practices - perhaps reflecting adaptations to conditions common to tallgrass prairie soils in the region (e.g., soil type, parent material, development under grasses, temperature and rainfall patterns, and annual freeze-thaw cycles). These findings are important for understanding the long-term consequences of land management practices to prairie soil microbial communities and their genetic potential to carry out key functions.