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At least 19 records
All sourcesNASA NTRSDOE OSTI
DOE OSTI2023

De Novo Assembly and Annotation of 11 Diverse Shrub Willow (Salix) Genomes Reveals Novel Gene Organization in Sex-Linked Regions

Poplar and willow species in the Salicaceae are dioecious, yet have been shown to use different sex determination systems located on different chromosomes. Willows in the subgenus Vetrix are interesting for comparative studies of sex determination systems, yet genomic resources for these species are still quite limited. Only a few annotated reference genome assemblies are available, despite many species in use in breeding programs. Here we present de novo assemblies and annotations of 11 shrub willow genomes from six species. Copy number variation of candidate sex determination genes within each genome was characterized and revealed remarkable differences in putative master regulator gene duplication and deletion. We also analyzed copy number and expression of candidate genes involved in floral secondary metabolism, and identified substantial variation across genotypes, which can be used for parental selection in breeding programs. Lastly, we report on a genotype that produces only female descendants and identified gene presence/absence variation in the mitochondrial genome that may be responsible for this unusual inheritance.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

Survey of Thirteen Novel Pseudomonas putida Bacteriophages

Bacteriophages have been widely investigated as a promising treatment of food, medical equipment, and humans colonized by antibiotic-resistant bacteria. Phages pose particular interest in combating those bacteria which form biofilms, such as the medically important human pathogen Pseudomonas aeruginosa and several plant pathogens, including P. syringae . In an undergraduate lab course, P. putida was used as the host to isolate novel anti-pseudomonal bacteriophages. Environmental samples of soil and water were collected, and purified phage isolates were obtained. After Illumina sequencing, genomes of these phages were assembled de novo and annotated. Assembled genomes were compared with known genomes in the literature and GenBank to identify taxonomic relations and to refine their functional annotations. The thirteen phages described are sipho-, myo-, and podoviruses in several families of Caudoviricetes , spanning several novel genera, with genomes ranging from 40,000 to 96,000 bp. One phage (DDSR119) is unique and is the first reported P. putida siphovirus. The remaining 12 can be clustered into four distinct groups. Six are highly related to each other and to previously described Autotranscriptaviridae phages: Waldo5, PlaquesPlease, and Laces98 all belong to the Waldovirus genus, whereas Stalingrad, Bosely, and Stamos belong to the Troedvirus genus. Zuri was previously classified as the founding member of a new genus Zurivirus within the family Schitoviridae . Ebordelon and Holyagarpour each represent different species within Zurivirus , whereas Meara is a more distantly related member of the Schitoviridae . Dolphis and Jeremy are similar enough to form a genus but have only a few distant relatives among sequenced phages and are notable for being temperate. We identified the lysis cassettes in all 13 phages, compared tail spike structures, and found auxiliary metabolic genes in several. Studies like these, which isolate and characterize infectious virions, enable the identification of novel proteins and molecular systems and also provide the raw materials for further study, evaluation, and manipulation of phage proteins and their hosts.

Pseudomonas putida↗
DOE OSTI2023

Two major chromosome evolution events with unrivaled conserved gene content in pomegranate

Pomegranate has a unique evolutionary history given that different cultivars have eight or nine bivalent chromosomes with possible crossability between the two classes. Therefore, it is important to study chromosome evolution in pomegranate to understand the dynamics of its population. Here, we de novo assembled the Azerbaijani cultivar “Azerbaijan guloyshasi” (AG2017; 2n = 16) and re-sequenced six cultivars to track the evolution of pomegranate and to compare it with previously published de novo assembled and re-sequenced cultivars. High synteny was observed between AG2017, Bhagawa (2n = 16), Tunisia (2n = 16), and Dabenzi (2n = 18), but these four cultivars diverged from the cultivar Taishanhong (2n = 18) with several rearrangements indicating the presence of two major chromosome evolution events. Major presence/absence variations were not observed as >99% of the five genomes aligned across the cultivars, while >99% of the pan-genic content was represented by Tunisia and Taishanhong only. We also revisited the divergence between soft- and hard-seeded cultivars with less structured population genomic data, compared to previous studies, to refine the selected genomic regions and detect global migration routes for pomegranate. We reported a unique admixture between soft- and hard-seeded cultivars that can be exploited to improve the diversity, quality, and adaptability of local pomegranate varieties around the world. Our study adds body knowledge to understanding the evolution of the pomegranate genome and its implications for the population structure of global pomegranate diversity, as well as planning breeding programs aiming to develop improved cultivars.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2021

Accelerating large scale de novo metagenome assembly using GPUs

Metagenomic workflows involve studying uncultured microorganisms directly from the environment. These environmental samples when processed by modern sequencing machines yield large and complex datasets that exceed the capabilities of metagenomic software. The increasing sizes and complexities of datasets make a strong case for exascale-capable metagenome assemblers. However, the underlying algorithmic motifs are not well suited for GPUs. This poses a challenge since the majority of next-generation supercomputers will rely primarily on GPUs for computation. In this paper we present the first of its kind GPU-Accelerated implementation of the local assembly approach that is an integral part of a widely used large-scale metagenome assembler, MetaHipMer. Local assembly uses algorithms that induce random memory accesses and non-deterministic workloads, which make GPU offloading a challenging task. Our GPU implementation outperforms the CPU version by about 7x and boosts the performance of MetaHipMer by 42% when running on 64 Summit nodes.

Awan, Muaaz Gul↗
DOE OSTI2022

HiFiAdapterFilt, a memory efficient read processing pipeline, prevents occurrence of adapter sequence in PacBio HiFi reads and their negative impacts on genome assembly

Abstract Background Pacific Biosciences HiFi read technology is currently the industry standard for high accuracy long-read sequencing that has been widely adopted by large sequencing and assembly initiatives for generation of de novo assemblies in non-model organisms. Though adapter contamination filtering is routine in traditional short-read analysis pipelines, it has not been widely adopted for HiFi workflows. Results Analysis of 55 publicly available HiFi datasets revealed that a read-sanitation step to remove sequence artifacts derived from PacBio library preparation from read pools is necessary as adapter sequences can be erroneously integrated into assemblies. Conclusions Here we describe the nature of adapter contaminated reads, their consequences in assembly, and present HiFiAdapterFilt, a simple and memory efficient solution for removing adapter contaminated reads prior to assembly.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

Population Genomics of Pseudocercospora griseola Reveals New Groups in the Middle American Clade and the Presence of the Endophytic Bacterium Achromobacter xylosoxidans

Angular leaf spot (ALS), caused by Pseudocercospora griseola is an important disease of common beans. P. griseola, is highly variable and has co-evolved with its host. In this study, 48 isolates of P. griseola from Puerto Rico, Guatemala, Honduras and Tanzania were sequenced (3RADseq), resulting in the de novo assembly of 42,214 contigs. Phylogenomic, population genetic structure and principal component analyses using 1,260 SNPs divided these isolates into two populations, Andean and Middle American, while the Middle American population was further divided into three sub-populations. There were moderate to high levels of differentiation between P. griseola populations, with pairwise Fst values ranging from 0.11 to 0.95. The Andean population was composed of isolates from Tanzania, and was separated from the Middle American population (Fst = 0.95). The Middle American population was separated into 3 subpopulations including isolates from: 1. Guatemala and Honduras, 2. Tanzania, and 3. Puerto Rico. Pathogenicity testing of 27 isolates from Puerto Rico, using 12 common bean differential lines, identified ten races, but these races were not associated with SNPs found in virulence genes. DNA of an endophytic bacterium (Achromobacter xylosoxidans) was found in seven mildly virulent isolates suggesting a possible role of the bacterium in the observed virulence patterns. To understand the evolution and diversity of P. griseola, further study of the virulence genes and the interactions among the endophytic bacterium, the fungus, and the host plant is required. Such information is critical to inform breeding strategies for the development of resistant germplasm and cultivars.

Serrato-Diaz, Luz M. [U.S. Department of Agricultu↗
DOE OSTI2022

Protocol for single-cell isolation and genome amplification of environmental microbial eukaryotes for genomic analysis

We describe environmental microbial eukaryotes (EMEs) sample collection, single-cell isolation, lysis, and genome amplification, followed by the rDNA amplification and OTU screening for recovery of high-quality species-specific genomes for de novo assembly. These protocols are part of our pipeline that also includes bioinformatic methods. The pipeline and its application on a wide range of phyla of different sample complexity are described in our complementary paper. In addition, this protocol describes optimized lysis, genome amplification, and OTU screening steps of the pipeline. For complete details on the use and execution of this protocol, please refer to Ciobanu et al. (2021).

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2021

Genomic insights into the origin, domestication and diversification of Brassica juncea

Despite early domestication around 3000 BC, the evolutionary history of the ancient allotetraploid species Brassica juncea (L.) Czern & Coss remains uncertain. Here, we report a chromosome-scale de novo assembly of a yellow-seeded B. juncea genome by integrating long-read and short-read sequencing, optical mapping and Hi-C technologies. Nuclear and organelle phylogenies of 480 accessions worldwide supported that B. juncea is most likely a single origin in West Asia, 8,000–14,000 years ago, via natural interspecific hybridization. Subsequently, new crop types evolved through spontaneous gene mutations and introgressions along three independent routes of eastward expansion. Selective sweeps, genome-wide trait associations and tissue-specific RNA-sequencing analysis shed light on the domestication history of flowering time and seed weight, and on human selection for morphological diversification in this versatile species. Our data provide a comprehensive insight into the origin and domestication and a foundation for genomics-based breeding of B. juncea.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Full genome sequence for the African swine fever virus outbreak in the Dominican Republic in 1980

African swine fever is a lethal disease of domestic pigs, geographically expanding as a pandemic, that is affecting countries across Eurasia and severely damaging their swine production industry. After more than 40 years of being absent in the Western hemisphere, in 2020 ASF reappeared in the Dominican Republic and Haiti. The recent outbreak strain in the Dominican Republic has been identified as a genotype II ASFV a derivative of the ASF strain circulating in Asia and Europe. However, to date no full-length genome sequence from either the 1978–1980 Here we report the complete genome sequence of an African swine fever virus (ASFV) (DR-1980) that was previously isolated from blood collected in 1980 from the Dominican Republic at the end of the last outbreak, before culling of all swine on the island of Hispaniola and stored in the Plum Island Animal Disease Center ASFV repository. A contig representing the full-length genome (183,687 base pairs) was de novo assembled into a single contig using both Nanopore and Illumina sequences. DR-1980 was determined to belong to genotype I and, as determined by full genome comparison, a close relative to the sequenced Sardinia viruses that were causing outbreaks at this time.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2021

NLR diversity and candidate fusiform rust resistance genes in loblolly pine

Resistance to fusiform rust disease in loblolly pine (Pinus taeda) is a classic gene-for-gene system. Early resistance gene mapping in the P. taeda family 10-5 identified RAPD markers for a major fusiform rust resistance gene, Fr1. More recently, single nucleotide polymorphism (SNP) markers associated with resistance were mapped to a full-length gene model in the loblolly pine genome encoding for a nucleotide-binding site leucine-rich repeat (NLR) protein. NLR genes are one of the most abundant gene families in plant genomes and are involved in effector-triggered immunity. Inter- and intraspecies studies of NLR gene diversity and expression have resulted in improved disease resistance. To characterize NLR gene diversity and discover potential resistance genes, we assembled de novo transcriptomes from 92 loblolly genotypes from across the natural range of the species. In these transcriptomes, we identified novel NLR transcripts that are not present in the loblolly pine reference genome and found significant geographic diversity of NLR genes providing evidence of gene family evolution. We designed capture probes for these NLRs to identify and map SNPs that stably cosegregate with resistance to the SC20-21 isolate of Cronartium quercuum f.sp. fusiforme (Cqf) in half-sib progeny of the 10-5 family. We identified 10 SNPs and 2 quantitative trait loci associated with resistance to SC20-21 Cqf. The geographic diversity of NLR genes provides evidence of NLR gene family evolution in loblolly pine. The SNPs associated with rust resistance provide a resource to enhance breeding and deployment of resistant pine seedlings.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

Improving the efficiency of Rubisco by resurrecting its ancestors in the family Solanaceae

Plants and photosynthetic organisms have a remarkably inefficient enzyme named Rubisco that fixes atmospheric CO 2 into organic compounds. Understanding how Rubisco has evolved in response to past climate change is important for attempts to adjust plants to future conditions. In this study, we developed a computational workflow to assemble de novo both large and small subunits of Rubisco enzymes from transcriptomics data. Next, we predicted sequences for ancestral Rubiscos of the (nightshade) family Solanaceae and characterized their kinetics after coexpressing them inEscherichia coli. Predicted ancestors of C 3 Rubiscos were identified that have superior kinetics and excellent potential to help plants adapt to anthropogenic climate change. Our findings also advance understanding of the evolution of Rubisco’s catalytic traits.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

Phenotypic and genomic characterization of Methanothermobacter wolfeii strain BSEL, a CO 2 -capturing archaeon with minimal nutrient requirements

A new variant of Methanothermobacter wolfeii was isolated from an anaerobic digester using enrichment cultivation in anaerobic conditions. Here, the new isolate was taxonomically identified via 16S rRNA gene sequencing and tagged as M. wolfeii BSEL. The whole genome of the new variant was sequenced and de novo assembled. Genomic variations between the BSEL strain and the type strain were discovered, suggesting evolutionary adaptations of the BSEL strain that conferred advantages while growing under a low concentration of nutrients. M. wolfeii BSEL displayed the highest specific growth rate ever reported for the wolfeii species (0.27 ± 0.03 h –1 ) using carbon dioxide (CO 2 ) as unique carbon source and hydrogen (H 2 ) as electron donor. M. wolfeii BSEL grew at this rate in an environment with ammonium (NH 4 + ) as sole nitrogen source. The minerals content required to cultivate the BSEL strain was relatively low and resembled the ionic background of tap water without mineral supplements. Optimum growth rate for the new isolate was observed at 64°C and pH 8.3. In this work, it was shown that wastewater from a wastewater treatment facility can be used as a low-cost alternative medium to cultivate M. wolfeii BSEL. Continuous gas fermentation fed with a synthetic biogas mimic along with H 2 in a bubble column bioreactor using M. wolfeii BSEL as biocatalyst resulted in a CO 2 conversion efficiency of 97% and a final methane (CH 4 ) titer of 98.5%v, demonstrating the ability of the new strain for upgrading biogas to renewable natural gas.

09 BIOMASS FUELS↗
DOE OSTI2024

Horizontal gene transfer of the Mer operon is associated with large effects on the transcriptome and increased tolerance to mercury in nitrogen-fixing bacteria

Abstract Background Mercury (Hg) is highly toxic and has the potential to cause severe health problems for humans and foraging animals when transported into edible plant parts. Soil rhizobia that form symbiosis with legumes may possess mechanisms to prevent heavy metal translocation from roots to shoots in plants by exporting metals from nodules or compartmentalizing metal ions inside nodules. Horizontal gene transfer has potential to confer immediate de novo adaptations to stress. We used comparative genomics of high quality de novo assemblies to identify structural differences in the genomes of nitrogen-fixing rhizobia that were isolated from a mercury (Hg) mine site that show high variation in their tolerance to Hg. Results Our analyses identified multiple structurally conserved merA homologs in the genomes of Sinorhizobium medicae and Rhizobium leguminosarum but only the strains that possessed a Mer operon exhibited 10-fold increased tolerance to Hg. RNAseq analysis revealed nearly all genes in the Mer operon were significantly up-regulated in response to Hg stress in free-living conditions and in nodules. In both free-living and nodule environments, we found the Hg-tolerant strains with a Mer operon exhibited the fewest number of differentially expressed genes (DEGs) in the genome, indicating a rapid and efficient detoxification of Hg from the cells that reduced general stress responses to the Hg-treatment. Expression changes in S. medicae while in bacteroids showed that both rhizobia strain and host-plant tolerance affected the number of DEGs. Aside from Mer operon genes, nif genes which are involved in nitrogenase activity in S. medicae showed significant up-regulation in the most Hg-tolerant strain while inside the most Hg-accumulating host-plant. Transfer of a plasmid containing the Mer operon from the most tolerant strain to low-tolerant strains resulted in an immediate increase in Hg tolerance, indicating that the Mer operon is able to confer hyper tolerance to Hg. Conclusions Mer operons have not been previously reported in nitrogen-fixing rhizobia. This study demonstrates a pivotal role of the Mer operon in effective mercury detoxification and hypertolerance in nitrogen-fixing rhizobia. This finding has major implications not only for soil bioremediation, but also host plants growing in mercury contaminated soils.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2021

Genome, transcriptome and secretome analyses of the antagonistic, yeast-like fungus Aureobasidium pullulans to identify potential biocontrol genes

Aureobasidium pullulans is an extremotolerant, cosmopolitan yeast-like fungus that successfully colonises vastly different ecological niches. The species is widely used in biotechnology and successfully applied as a commercial biocontrol agent against postharvest diseases and fireblight. However, the exact mechanisms that are responsible for its antagonistic activity against diverse plant pathogens are not known at the molecular level. Thus, it is difficult to optimise and improve the biocontrol applications of this species. As a foundation for elucidating biocontrol mechanisms, we have de novo assembled a high-quality reference genome of a strongly antagonistic A. pullulans strain, performed dual RNA-seq experiments, and analysed proteins secreted during the interaction with the plant pathogen Fusarium oxysporum. Based on the genome annotation, potential biocontrol genes were predicted to encode secreted hydrolases or to be part of secondary metabolite clusters (e.g., NRPS-like, NRPS, T1PKS, terpene, and β-lactone clusters). Transcriptome and secretome analyses defined a subset of 79 A. pullulans genes (among the 10,925 annotated genes) that were transcriptionally upregulated or exclusively detected at the protein level during the competition with F. oxysporum. These potential biocontrol genes comprised predicted secreted hydrolases such as glycosylases, esterases, and proteases, as well as genes encoding enzymes, which are predicted to be involved in the synthesis of secondary metabolites. This study highlights the value of a sequential approach starting with genome mining and consecutive transcriptome and secretome analyses in order to identify a limited number of potential target genes for detailed, functional analyses.

transcriptome↗
DOE OSTI2023

HiC manuscript: bulk metagenomic data_MG_raw Upload

The raw bulk metagenomic data were trimmed and de-novo assembled. The assemblies were screened for viral contigs (by virsorter2, Deepvirfinder, VIBRANT, checkV) and binned to generate MAGs that were further refined by Hi-C read links.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

HiC manuscript: bulk metagenomic HiC metaG Raw

The raw Hi-C metagenomic data were trimmed and aligned to the de-novo assemblies obtained from the paired shotgun metagenomes following the Hi-C kit manufacturer's recommendations (https://phasegenomics.github.io/2019/09/19/hic-alignment-and-qc.html). The contig-contig links or Hi-C links were quality-filtered for the identification of virus-host pairs.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Genome-Resolved Metagenomics of Nitrogen Transformations in the Switchgrass Rhizosphere Microbiome on Marginal Lands

Switchgrass (Panicum virgatum L.) remains the preeminent American perennial (C4) bioenergy crop for cellulosic ethanol, that could help displace over a quarter of the US current petroleum consumption. Intriguingly, there is often little response to nitrogen fertilizer once stands are established. The rhizosphere microbiome plays a critical role in nitrogen cycling and overall plant nutrient uptake. We used high-throughput metagenomic sequencing to characterize the switchgrass rhizosphere microbial community before and after a nitrogen fertilization event for established stands on marginal land. We examined community structure and bulk metabolic potential, and resolved 29 individual bacteria genomes via metagenomic de novo assembly. Community structure and diversity were not significantly different before and after fertilization; however, the bulk metabolic potential of carbohydrate-active enzymes was depleted after fertilization. We resolved 29 metagenomic assembled genomes, including some from the ‘most wanted’ soil taxa such as Verrucomicrobia, Candidate phyla UBA10199, Acidobacteria (rare subgroup 23), Dormibacterota, and the very rare Candidatus Eisenbacteria. The Dormibacterota (formally candidate division AD3) we identified have the potential for autotrophic CO utilization, which may impact carbon partitioning and storage. Our study also suggests that the rhizosphere microbiome may be involved in providing associative nitrogen fixation (ANF) via the novel diazotroph Janthinobacterium to switchgrass.

60 APPLIED LIFE SCIENCES↗