Biology subjects

Tully, B. J.

Publications and source records attributed to Tully, B. J..

11 recordsLinked to original sources

bioRxiv2023

Benchmarking A Novel Quantitative PCR-based Microbiome Profiling Platform Against Sequencing-based Methods

BackgroundPCR-based diagnostics, predominantly utilized for pathogen detection, have faced challenges in broader microbial profiling due to disparities in genomic data availability. This study addresses this limitation by exploiting the surge in the number of microbial genomes, facilitated by advancements in next-generation sequencing (NGS) and metagenomic-assembled genomes. The primary aim was to develop and validate quantitative PCR (qPCR) assays for a wide range of gut commensals, traditionally overlooked due to inadequate genomic information. We sought to compare the efficacy of these qPCR assays against established NGS microbiome profiling methodologies - 16S amplicon and metagenomic sequencing. MethodsWe designed 110 species-specific qPCR assays for gut commensals using a novel proprietary in silico pipeline and validated the assays against stool samples from three healthy donors. The quantitative microbiome profiles were compared to taxonomic profiles generated by standard bioinformatic approaches for 16S amplicon and metagenomic sequencing. 16S amplicons were analyzed as amplicon sequence variants produced by DADA2 and metagenomic sequences were analyzed by multiple iterations of MetaPhlAn (versions 2, 3, and 4) and Kraken2/Bracken paired with two different genomic databases. The qPCR assays were assessed for their ability to detect low abundance microbes and their correlation with NGS results, focusing on taxonomic resolution and limits of quantification. ResultsThe qPCR assays demonstrated high concordance with advanced metagenomic and the ineffectiveness of 16S amplicon methods to achieve species-level assignments. qPCR microbiome profiles were more highly correlated with the most current bioinformatic methods than the bioinformatics methods were to each other. The profile comparisons also highlight how the continued use of older bioinformatics protocols can limit results and lead to misinterpretation of data. Notably, qPCR identified taxa undetected or underestimated by metagenomic approaches, revealing limitations in current bioinformatics tools for differentiating closely related species and quantifying low abundance taxa. ConclusionsThis study establishes qPCR as a robust tool for large-scale microbiome profiling, offering enhanced accuracy, sensitivity, and quantitative capabilities compared to standard NGS methods. Our findings advocate for the integration of qPCR in standardizing microbiome detection, providing a pathway towards developing human microbiome profiling platforms capable of accurate species quantification. The adoption of qPCR assays could lead to more consistent, reliable, and cost-effective microbiome research and diagnostics.

microbiology↗

bioRxiv2023

Microbial abundance and diversity in 64-74 Ma subseafloor igneous basement from the Louisville Seamount Chain

The aquifer in subseafloor igneous basement is a massive, continuous microbial substrate, yet sparingly little is known about life in this habitat. The work to date has focused largely on describing microbial diversity in young basement (<10 Ma) at oceanic spreading regions and ridge flanks, where the basaltic crust is still porous and fluid flow through it is active. While the prevailing belief used to be that fluid flow through older parts of the seafloor was non-existent, recent heat flow models predict that fluid moves through subseafloor basement >65 Ma, and that seamounts can act as mid-plate conduits for fluids into and out of the subsurface aquifer in older crustal settings. Here we test the hypothesis that microbial life exists in subseafloor basement >65Ma using samples collected from the Louisville Seamount Chain via seafloor drilling. Cell biomass was heterogeneous in nature and ranged from below detection to [~]104 cells cm-3. Bacterial 16S rRNA genes from core samples and enrichment incubations are dominated by lineages putatively carrying out hydrocarbon oxidation and nitrogen, sulfur and metal redox processes. Samples from two different seamounts were statistically different, indicating some degree of biogeography. Archaea were not detected via quantitative polymerase chain reaction, indicating they are rare in the Louisville subsurface. Taken together, the data indicate that microbial life is indeed present in subseafloor igneous basement >65 Ma, which significantly expands the range of the subseafloor biosphere where microbial life is known to exist. Impact StatementThe aquifer in subseafloor igneous basement is the largest continuous microbial substrate on Earth, but it is difficult to access and therefore understudied. We here collected samples from the Louisville Seamount Chain using seafloor drilling to determine if microbial life exists in the >65 Ma subseafloor basement made at these seamounts. A low biomass environment dominated by Bacteria potentially capable of using the Fe and S inherent in subseafloor basalt was detected, including Bacteria that were revived in enrichment experiments. This discovery expands the range of seafloor where confirmed microbial life exists and indicates the interior of seamounts is habitable.

ecology↗

bioRxiv2022

High transposase abundance in the deep ocean is linked to a particle-associated lifestyle

Transposases are mobile genetic elements (MGEs) that move within and between genomes, promoting genomic plasticity in microorganisms. In marine microbial communities, the abundance of transposases increases with depth, but the reasons behind this trend remain unclear. Our analysis of metagenomes from the Tara Oceans and Malaspina Expeditions suggests that a particle-associated lifestyle is the main covariate for the high occurrence of transposases in the deep ocean. We observed a strong and depth-independent correlation between transposase abundance and the presence of biofilm-associated genes, as well as the prevalence of secretory enzymes. This suggests that mobile genetic elements readily propagate among microbial communities within crowded biofilms. Furthermore, we show that particle association positively correlates with larger genome size, which is in turn is associated with higher transposase abundance. Cassette sequences associated with transposons are enriched with genes related to defense mechanisms, which are more highly expressed in the deep sea. Thus, while transposons spread at the expense of their microbial hosts, they also introduce novel genes and potentially benefit the hosts in helping to compete for limited resources. Overall, our results suggest a new understanding of deep ocean particles as highways for gene sharing among defensively oriented microbial genomes. ImportanceGenes can move within and between microbial genomes via mobile genetic elements, which include transposases and transposons. In the oceans, there is a puzzling increase in transposase abundance in microbial genomes as depth increases. To gain insight into this trend, we conducted an extensive analysis of marine microbial metagenomes and metatranscriptomes. We found a significant correlation between transposase abundance and a particle-associated lifestyle among marine microbes. We also observed a link between transposase abundance and genes related to defense mechanisms. These results suggest that as microbes become densely packed into crowded particles, mobile genes are more likely to spread and carry genetic material that provides a competitive advantage in crowded habitats. This may enable deep sea microbes to effectively compete in such environments.

bioinformatics↗

bioRxiv2021

The high-throughput gene prediction of more than 1,700 eukaryote genomes using the software package EukMetaSanity

Gene prediction and annotation for eukaryotic genomes is challenging with large data demands and complex computational requirements. For most eukaryotes, genomes are recovered from specific target taxa. However, it is now feasible to reconstruct or sequence hundreds of metagenome-assembled genomes (MAGs) or single-amplified genomes directly from the environment. To meet this forth-coming wave of eukaryotic genome generation, we introduce EukMetaSanity, which combines state-of-the-art tools into three pipelines that have been specifically designed for extensive parallelization on high-performance computing infrastructure. EukMetaSanity performs an automated taxonomy search against a protein database of 1,482 species to identify phylogenetically compatible proteins to be used in downstream gene prediction. We present the results for intron, exon, and gene locus prediction for 112 genomes collected from NCBI, including fungi, plants, and animals, along with 1,669 MAGs and demonstrate that EukMetaSanity can provide reliable preliminary gene predictions for a single target taxon or at scale for hundreds of MAGs. EukMetaSanity is freely available at https://github.com/cjneely10/EukMetaSanity.

bioinformatics↗

bioRxiv2021

Eukaryotic genomes from a global metagenomic dataset illuminate trophic modes and biogeography of ocean plankton

Metagenomics is a powerful method for interpreting the ecological roles and physiological capabilities of mixed microbial communities. Yet, many tools for processing metagenomic data are not designed to consider eukaryotes, nor are they built for an increasing amount of sequence data. EukHeist is an automated pipeline to retrieve eukaryotic and prokaryotic metagenome assembled genomes (MAGs) from large-scale metagenomic datasets. We developed the EukHeist workflow to specifically process large amounts of both metagenomic and/or metatranscriptomic sequence data in an automated and reproducible fashion. Here, we applied EukHeist to the large-size fraction data (0.8-2000{micro}m) from Tara Oceans to recover both eukaryotic and prokaryotic MAGs, which we refer to as TOPAZ (Tara Oceans Particle-Associated MAGs). The TOPAZ MAGs consisted of >900 environmentally-relevant eukaryotic MAGs and >4,000 bacterial and archaeal MAGs. The bacterial and archaeal TOPAZ MAGs expand the known marine phylogenetic diversity through the increase in coverage of likely particle- and host-associated taxa. We also demonstrate an approach to infer the putative functional mode of the recovered eukaryotic MAGs. A global survey of the TOPAZ MAGs enabled the identification of ecological cohorts, driven by specific environmental factors, and putative host-microbe associations. ImportanceDespite the ecological importance of single-celled eukaryotic organisms in marine environments, the majority are difficult to cultivate in the lab. Sequencing genetic material extracted from environmental samples enables researchers to document naturally-occurring protistan communities. However, conventional sequencing methodologies cannot separate out the genomes of individual organisms. To more completely capture the entire genomic content of mixed protistan community, we can create bins of sequences that represent the same organism. We developed a pipeline that enables scientists to bin individual organisms out of metagenomic reads, and show results that provide exciting insights into what protistan communities are present in the ocean and what roles they play in the ecosystem. Here, a global survey of both eukaryotic and prokaryotic MAGs enabled the identification of ecological cohorts, driven by specific environmental factors, and putative host-microbe associations. Accessible and scalable computational tools, such as EukHeist, are likely to accelerate the identification of meaningful genetic signatures from large datasets, ultimately expanding the eukaryotic tree of life.

microbiology↗

bioRxiv2021

Microbial population dynamics are dominated by stochastic forces in a low biomass subseafloor habitat

The subseafloor is a vast habitat that supports microorganisms that have a global scale impact on geochemical cycles. Many of the endemic microbial communities inhabiting the subseafloor consist of small populations under growth-limited conditions. For small populations, stochastic evolutionary events can have large impacts on intraspecific population dynamics and allele frequencies. These conditions are fundamentally different from those experienced by most microorganisms in surface environments, and it is unknown how small population sizes and growth-limiting conditions influence evolution and population structure in the subsurface. Using a two-year, high-resolution environmental time-series, we examine the dynamics of microbial populations from cold, oxic crustal fluids collected from the subseafloor site North Pond, located near the mid-Atlantic ridge. Our results reveal rapid shifts in overall abundance, allele frequency, and strain abundance across the time points observed, with evidence for homologous recombination between coexisting lineages. We show that the subseafloor aquifer is a dynamic habitat that hosts microbial metapopulations that disperse frequently through the crustal fluids, enabling gene flow and recombination between microbial populations. The dynamism and stochasticity of microbial population dynamics in North Pond suggests that these forces are important drivers in the evolution of microbial populations in the vast subseafloor habitat. Significance StatementThe cold, oxic subseafloor is an understudied habitat that is difficult to access, yet important to global biogeochemical cycles and starkly different compared to microbial habitats on the surface of the Earth. Our understanding of microbial evolution and population dynamics is largely molded by studies of microbes living in surface habitats that can host 10-1,000 times more microbial biomass than is frequently observed in the subsurface. This study provides an opportunity to observe population dynamics within a low biomass, growth-limited environment and reveals that microbial populations in the subseafloor are influenced by changes in selection pressure and gene sweeps. In addition, recombination between strains that have dispersed from elsewhere within the aquifer has an important impact on the evolution of microbial populations. Much of the microbial life on the planet exists under growth-limited conditions and the subseafloor provides a natural laboratory to explore how life evolves in such environments.

microbiology↗

bioRxiv2020

Time-series transcriptomics from cold, oxic subseafloor crustal fluids reveals a motile, mixotrophic microbial community

The rock-hosted oceanic crustal aquifer is one of the largest habitable volumes on Earth, and it harbors a reservoir of microbial life that influences global-scale biogeochemical cycles. Here, we use time series metagenomic and metatranscriptomic data from a low-temperature, ridge flank environment that is representative of the majority of global hydrothermal fluid circulation in the ocean to reconstruct microbial metabolic potential, transcript abundance, and community dynamics. The data suggest that the microbial community in this subseafloor habitat is motile, chiefly heterotrophic or mixotrophic, and capable of using alternative electron acceptors such as nitrate and thiosulfate, in addition to oxygen. Anaerobic processes are most abundant in subseafloor horizons deepest in the aquifer, furthest from connectivity with the deep ocean, and there was little overlap in the active microbial populations between sampling horizons. Together, our results indicate the microbial community in the North Pond aquifer plays an important role in the oxidation of organic carbon within the crust, and is also metabolically flexible, with the ability to switch from autotrophy to heterotrophy, as well as function under low oxygen conditions. This work highlights the heterogeneity of microbial life in the subseafloor aquifer and provides new insights into biogeochemical cycling in ocean crust.

microbiology↗

bioRxiv2020

Marine Dadabacteria exhibit genome streamlining and phototrophy-driven niche partitioning

The remineralization of organic material via heterotrophy in the marine environment is performed by a diverse and varied group of microorganisms that can specialize in the type of organic material degraded and the niche they occupy. The marine Dadabacteria are cosmopolitan in the marine environment and belong to a candidate phylum for which there has not been a comprehensive assessment of the available genomic data to date. Here in, we assess the functional potential of the oligotrophic, marine Dadabacteria in comparison to terrestrial, coastal, and subsurface members of the phylum. Our analysis reveals that the marine Dadabacteria have undergone a genome streamlining event, reducing their genome size and the nitrogen content of their DNA and predicted proteome, relative to their terrestrial counterparts. Collectively, the Dadabacteria have the potential to degrade microbial particulate organic matter, specifically peptidoglycan and phospholipids. The marine Dadabacteria belong to two clades with distinct ecological niches in global metagenomic data: a shallow clade with the potential for photoheterotrophy through the use of proteorhodopsin, present predominantly in surface waters up to 100m depth; and a deep clade lacking the potential for photoheterotrophy that is more abundant in the deep photic zone.

microbiology↗

bioRxiv2019

MetaSanity: An integrated, customizable microbial genome evaluation and annotation pipeline

SummaryAs the importance of microbiome research continues to become more prevalent and essential to understanding a wide variety of ecosystems (e.g., marine, built, host-associated, etc.), there is a need for researchers to be able to perform highly reproducible and quality analysis of microbial genomes. MetaSanity incorporates analyses from eleven existing and widely used genome evaluation and annotation suites into a single, distributable workflow, thereby decreasing the workload of microbiologists by allowing for a flexible, expansive data analysis pipeline. MetaSanity has been designed to provide separate, reproducible workflows, that (1) can determine the overall quality of a microbial genome, while providing a putative phylogenetic assignment, and (2) can assign structural and functional gene annotations with varying degrees of specificity to suit the needs of the researcher. The software suite combines the results from several tools to provide broad insights into overall metabolic function and putative extracellular localization of peptidases and carbohydrate-active enzymes. Importantly, this software provides built-in optimization for \"big data\" analysis by storing all relevant outputs in an SQL database, allowing users to query all the results for the elements that will most impact their research.\n\nAvailability and implementationMetaSanity is provided under the GNU General Public License v.3.0 and is available for download at https://github.com/cjneely10/MetaSanity. This application is distributed as a Docker image. MetaSanity is implemented in Python3/Cython and C++.\n\nSupplementary informationSupplementary data are available below.

bioinformatics↗

bioRxiv2018

Metabolic Diversity within the Globally Abundant Marine Group II Euryarchaea Drives Ecological Patterns

Despite their discovery over 25 years ago, the Marine Group II Euryarchaea (MGII) have remained a difficult group of organisms to study, lacking cultured isolates and genome references. The MGII have been identified in marine samples from around the world and evidence supports a photoheterotrophic lifestyle combining phototrophy via proteorhodopsins with the remineralization of high molecular weight organic matter. Divided between two clades, the MGII have distinct ecological patterns that are not understood based on the limited number of available genomes. Here, I present the comparative genomic analysis of 250 MGII genomes, providing the most detailed view of these mesophilic archaea to-date. This analysis identified 17 distinct subclades including nine subclades that previously lacked reference genomes. The metabolic potential and distribution of the MGII genera revealed distinct roles in the environment, identifying algal-saccharide-degrading coastal subclades, protein-degrading oligotrophic surface ocean subclades, and mesopelagic subclades lacking proteorhodopsins common in all other subclades. This study redefines the MGII and provides an avenue for understanding the role these organisms play in the cycling of organic matter throughout the water column.

microbiology↗

bioRxiv2017

The Reconstruction of 2,631 Draft Metagenome-Assembled Genomes from the Global Oceans

Microorganisms play a crucial role in mediating global biogeochemical cycles in the marine environment. By reconstructing the genomes of environmental organisms through metagenomics, researchers are able to study the metabolic potential of Bacteria and Archaea that are resistant to isolation in the laboratory. Utilizing the large metagenomic dataset generated from 234 samples collected during the Tara Oceans circumnavigation expedition, we were able to assemble 102 billion paired-end reads into 562 million contigs, which in turn were co-assembled and consolidated in to 7.2 million contigs [≥]2kb in length. Approximately 1 million of these contigs were binned to reconstruct draft genomes. In total, 2,631 draft genomes with an estimated completion of [≥]50% were generated (1,491 draft genomes >70% complete; 603 high-quality genomes >90% complete). A majority of the draft genomes were manually assigned phylogeny based on sets of concatenated phylogenetic marker genes and/or 16S rRNA gene sequences. The draft genomes are now publically available for the research community at-large.

microbiology↗