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All sourcesNASA NTRSDOE OSTI
DOE OSTI2024

The Exoproteome and Surfaceome of Toxigenic Corynebacterium diphtheriae 1737 and Its Response to Iron Restriction and Growth on Human Hemoglobin

Toxin-producing Corynebacterium diphtheriae strains are the etiological agents of the severe upper respiratory disease, diphtheria. A global phylogenetic analysis revealed that biotype gravis is particularly lethal as it produces diphtheria toxin and a range of other virulence factors, particularly when it encounters low levels of iron at sites of infection. Here, to gain insight into how it colonizes its host, we have identified iron-dependent changes in the exoproteome and surfaceome of C. diphtheriae strain 1737 using a combination of whole-cell fractionation, intact cell surface proteolysis, and quantitative proteomics. In total, we identified 1414 of the predicted 2265 proteins (62%) encoded by its reference genome. For each protein, we quantified its degree of secretion and surface exposure, revealing that exoproteases and hydrolases predominate in the exoproteome, while the surfaceome is enriched with adhesins, particularly DIP2093. Our analysis provides insight into how components in the heme-acquisition system are positioned, showing pronounced surface exposure of the strain-specific ChtA/ChtC paralogues and high secretion of the species-conserved heme-binding HtaA protein, suggesting it functions as a hemophore. Profiling the response of the exoproteome and surfaceome after microbial exposure to human hemoglobin and iron limitation reveals potential virulence factors that may be expressed at sites of infection. Data are available via ProteomeXchange with identifier PXD051674.

cell envelope↗
DOE OSTI2026

The landscape of regulatory element evolution in a C4 perennial grass

Gene regulatory evolution is a well-known source of phenotypic diversity and adaptive evolution. Although cis-regulatory elements (CREs) play a vital role in gene expression evolution, the molecular evolution of CREs remains mostly unknown due to the difficulty in identifying and characterizing these functional elements. Comparative genomic analyses of noncoding DNA can be leveraged to identify conserved noncoding sequences (CNS), many of which may harbor functional CREs conserved by purifying selection. However, purely computational inference of CREs from putative CNS can be erroneous due to the complex genomic architecture in plants. One promising experimental approach to identify CREs is by profiling accessible chromatin regions (ACRs) that are often associated with the location of CREs. In this study, we use comparative genomics along with the profiling of ACRs to study the molecular evolution of putative functional noncoding regulatory regions in Panicoid grasses. We identified sets of CNS that varied in relationship to the degree of evolutionary divergence among the studied taxa, including identifying core-Panicoid-CNS. We augmented this analysis by profiling ACRs in Panicum hallii ecotypes using ATAC-seq. ACRs had low SNP density at the summit, harbored a high frequency of core-Panicoid-CNS, and were enriched with expression QTL. These data help to annotate the P. hallii genome for putative functional elements and suggest that a large proportion of these ACRs are evolving under purifying selection. Turnover in CNS and ACR between ecotypes of P. hallii identifies a small set of putatively divergent CREs that may underlie differences in gene regulation between genotypes from inland and coastal habitats. In summary, we profiled ACRs in Panicoid grasses and integrated this data with our putative CNS prediction framework, which provides unique insight into patterns of polymorphism and divergence in CREs in C4 perennial grasses.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

Testing for the Genomic Footprint of Conflict Between Life Stages in an Angiosperm and Moss Species

Abstract The maintenance of genetic variation by balancing selection is of considerable interest to evolutionary biologists. An important but understudied potential driver of balancing selection is antagonistic pleiotropy between diploid and haploid stages of the plant life cycle. Despite sharing a common genome, sporophytes (2n) and gametophytes (n) may undergo differential or even opposing selection. Theoretical work suggests antagonistic pleiotropy between life stages can generate balancing selection and maintain genetic variation. Despite the potential for far-reaching consequences of gametophytic selection, empirical tests of its pleiotropic effects (neutral, synergistic, or antagonistic) on sporophytes are generally lacking. Here, we examined the population genomic signals of selection across life stages in the angiosperm Rumex hastatulus and the moss Ceratodon purpureus. We compared gene expression between life stages and sexes, combined with neutral diversity statistics and the analysis of the distribution of fitness effects. In contrast to what would be predicted under balancing selection due to antagonistic pleiotropy, we found that unbiased genes between life stages were under stronger purifying selection, likely explained by a predominance of synergistic pleiotropy between life stages and strong purifying selection on broadly expressed genes. In addition, we found that 30% of candidate genes under balancing selection in R. hastatulus were located within inversion polymorphisms. Our findings provide novel insights into the genome-wide characteristics and consequences of plant gametophytic selection.

Evolutionary Biology↗
NASA NTRS2003

Phylogenetic analysis of anaerobic psychrophilic enrichment cultures obtained from a greenland glacier ice core

The examination of microorganisms in glacial ice cores allows the phylogenetic relationships of organisms frozen for thousands of years to be compared with those of current isolates. We developed a method for aseptically sampling a sediment-containing portion of a Greenland ice core that had remained at -9 degrees C for over 100,000 years. Epifluorescence microscopy and flow cytometry results showed that the ice sample contained over 6 x 10(7) cells/ml. Anaerobic enrichment cultures inoculated with melted ice were grown and maintained at -2 degrees C. Genomic DNA extracted from these enrichments was used for the PCR amplification of 16S rRNA genes with bacterial and archaeal primers and the preparation of clone libraries. Approximately 60 bacterial inserts were screened by restriction endonuclease analysis and grouped into 27 unique restriction fragment length polymorphism types, and 24 representative sequences were compared phylogenetically. Diverse sequences representing major phylogenetic groups including alpha, beta, and gamma Proteobacteria as well as relatives of the Thermus, Bacteroides, Eubacterium, and Clostridium groups were found. Sixteen clone sequences were closely related to those from known organisms, with four possibly representing new species. Seven sequences may reflect new genera and were most closely related to sequences obtained only by PCR amplification. One sequence was over 12% distant from its closest relative and may represent a novel order or family. These results show that phylogenetically diverse microorganisms have remained viable within the Greenland ice core for at least 100,000 years.

Evaluation Studies↗
NASA NTRS2006

Genetic mutation underlying orthostatic intolerance and diagnostic and therapeutic methods relating thereto

Isolated polynucleotide molecules and peptides encoded by these molecules are used in the analysis of human norepinephrine (NE) transporter variants, as well as in diagnostic and therapeutic applications, relating to a human NE transporter polymorphism. By analyzing genomic DNA or amplified genomic DNA, or amplified cDNA derived from mRNA, it is possible to type a human NE transporter with regard to the human NE transporter polymorphism, for example, in the context of diagnosing and treating NE transport impairments, and disorders associated with NE transport impairments, such as orthostatic intolerance.

Robertson, David↗
DOE OSTI2026

Compilation and utilization of a sorghum transcriptome compendium for gene regulatory network analysis and crop trait engineering

Sorghum bicolor (Sorghum) is a drought and heat tolerant C4 grass crop used to produce grain, forage, biofuels, and other bioproducts. Genetic improvement of sorghum hybrid crops is aided by a large and diverse germplasm, sorghum's diploid inbreeding genetics, and a relatively small genome that has facilitated genomic research. Over the past 20 years, the sorghum research community characterized the cytogenetic and recombinant landscapes of sorghum's 10 chromosomes, sequenced and annotated the sorghum genome, and used that information to identify genes/alleles that modulate flowering time, plant height, seed shattering, and other important traits. More recently, >1000 RNA-seq transcriptome profiles were collected from 15 sorghum genotypes to help understand the genetic basis of variation in growth and development of sorghum stems, tillers, roots, and leaves, and the regulation of biosynthetic pathways that produce epicuticular wax, dhurrin, and RFOs, compounds that contribute to sorghum's resilience. Transcriptome studies were designed to identify differentially expressed genes that are co-expressed during development or in response to a treatment to enable construction of gene regulatory networks. Co-expression and network analysis identified transcription factors and their cognate binding sites in target gene promoters and signaling pathways that modulate gene regulatory networks providing gene editing targets for further trait optimization. RNA-seq data from >20 experiments targeting sorghum organs, tissues, cell types, developmental stages, and responses to environmental conditions (i.e., diel, day-length, shading, water-deficit, temperature) has been compiled in a sorghum transcriptome compendium. The goal of this resource paper is to describe compendium content, accessibility, and a compendium data analysis pipeline and to illustrate the types of information that can be derived from the compendium with a focus on the elucidation of gene regulatory networks useful for guiding the improvement of sorghum traits through gene editing.

RNA-seq↗
NASA NTRS2001

Genomic instability in human lymphoid cells exposed to 1 GeV/amu Fe ions

The goal of this study was to assess whether charged particle radiations of importance to spaceflight elicit genomic instability in human TK6 lymphoblasts. The incidence of genomic instability in TK6 cells was assessed ~21 days after exposure to 2, 4, or 6 Fe ions (1 GeV/amu, LET= 146 keV/micrometers). Three indices of instability were used: intraclonal karyotypic heterogeneity, mutation rate analysis at the thymidine kinase (TK1) locus, and re-cloning efficiency. Fifteen of sixty clones demonstrated karyotypic heterogeneity. Five clones had multiple indicators of karyotypic change. One clone was markedly hypomutable and polyploid. Six clones were hypomutable, while 21 clones were mutators. Of these, seven were karyotypically unstable. Six clones had low re-cloning efficiencies, one of which was a mutator. All had normal karyotypes. In summary, many clones that survived exposure to a low fluence of Fe ions manifested one or more forms of genomic instability that may hasten the development of neoplasia through deletion or by recombination.

Non-NASA Center↗
NASA NTRS1997

Mechanical continuity and reversible chromosome disassembly within intact genomes removed from living cells

Chromatin is thought to be structurally discontinuous because it is packaged into morphologically distinct chromosomes that appear physically isolated from one another in metaphase preparations used for cytogenetic studies. However, analysis of chromosome positioning and movement suggest that different chromosomes often behave as if they were physically connected in interphase as well as mitosis. To address this paradox directly, we used a microsurgical technique to physically remove nucleoplasm or chromosomes from living cells under isotonic conditions. Using this approach, we found that pulling a single nucleolus or chromosome out from interphase or mitotic cells resulted in sequential removal of the remaining nucleoli and chromosomes, interconnected by a continuous elastic thread. Enzymatic treatments of interphase nucleoplasm and chromosome chains held under tension revealed that mechanical continuity within the chromatin was mediated by elements sensitive to DNase or micrococcal nuclease, but not RNases, formamide at high temperature, or proteases. In contrast, mechanical coupling between mitotic chromosomes and the surrounding cytoplasm appeared to be mediated by gelsolin-sensitive microfilaments. Furthermore, when ion concentrations were raised and lowered, both the chromosomes and the interconnecting strands underwent multiple rounds of decondensation and recondensation. As a result of these dynamic structural alterations, the mitotic chains also became sensitive to disruption by restriction enzymes. Ion-induced chromosome decondensation could be blocked by treatment with DNA binding dyes, agents that reduce protein disulfide linkages within nuclear matrix, or an antibody directed against histones. Fully decondensed chromatin strands also could be induced to recondense into chromosomes with pre-existing size, shape, number, and position by adding anti-histone antibodies. Conversely, removal of histones by proteolysis or heparin treatment produced chromosome decondensation which could be reversed by addition of histone H1, but not histones H2b or H3. These data suggest that DNA, its associated protein scaffolds, and surrounding cytoskeletal networks function as a structurally-unified system. Mechanical coupling within the nucleoplasm may coordinate dynamic alterations in chromatin structure, guide chromosome movement, and ensure fidelity of mitosis.

NASA Discipline Cell Biology↗
DOE OSTI2024

Improving I/O-aware Workflow Scheduling via Data Flow Characterization and trade-off Analysis

The scientific computing paradigm has transitioned from compute-intensive to I/O-intensive and memory-intensive in the past decade, especially when data-driven science has become common practice. Numerous empirical I/O-aware scheduling optimizations have been developed by incorporating I/O capacity and bandwidth as constraints into scheduling. Unfortunately, there is a lack of data flow (I/O) characterization tool and an understanding of trade-offs between concurrency, locality, and I/O bandwidth. To bridge the gap, this work 1) presents a set of descriptors to characterize, organize, and visualize I/O profiles, including flow size, I/O bandwidth, and operation count, which group data flows by I/O types, tasks, and files; 2) proposes an I/O Roofline model-based trade-off analysis to find the optimal trade-off between flow operational intensity, concurrency, and flow performance. The I/O descriptors generate useful insights into complicated I/O behaviors, suggesting distinct concurrency, storage, and scheduling to be used by types, tasks, and files. The proposed trade-off analysis guides scheduling decisions that generate resource assignment with the best flow parallelism. We evaluate our I/O-aware scheduling methodology on a highly I/O-intensive workflow–1000 Genomes. The experimental results demonstrate speedups of up to 2.4× compared to the state-of-the- art methods.

Guo, Luanzheng [BATTELLE (PACIFIC NW LAB)]↗
DOE OSTI2025

Hybridization capture sequencing for Vibrio spp. and associated virulence factors

ABSTRACT Proliferation ofVibriospp. in aquatic ecosystems is associated with climate change and, concomitantly, increased incidence of vibriosis. They are autochthonous to aquatic environments globally, but traditional metagenomic methods for detecting and typing pathogenicVibriospp. are challenged by their presence in relatively low abundance and ability to persist in a viable but nonculturable state. In the study reported here, hybridization capture sequencing (HCS) was employed to profile low-abundanceVibriospp. in environmental samples. The HCS panel targeted a family of molecular chaperones (CPN60) specific to 69Vibriospp. and 162Vibrio-specific virulence factors. This approach was evaluated in parallel with traditional whole-community shotgun sequencing in a metagenomic analysis of water and oyster samples collected from the Chesapeake Bay. In addition,Vibrio parahaemolyticusandVibrio vulnificusstrains isolated from the samples were subjected to whole-genome sequencing to determine the genetic characteristics of pathogenicVibriospp. circulating in an aquatic environment. HCS, employed to determine the incidence and characterization of specificVibriospp., yielded significantly greater metagenomic insight, notably a variety of otherVibriospp., including detection ofVibrio cholerae,Vibrio fluvialis, andVibrio aestuarianus, in addition toVibrio parahaemolyticusandVibrio vulnificus, and also important virulence factors not detectable using traditional molecular methods. Thus, pathogenicVibriospp. in aquatic ecosystems may be far more common than currently understood. It is concluded that environmental surveillance should include HCS, a valuable tool for the detection and characterization of pathogenic agents in aquatic ecosystems, notably vibrios. IMPORTANCE The increasing prevalence of pathogenicVibriospp. in aquatic ecosystems, driven by climate change, is closely linked to a rise in cholera and vibriosis cases, emphasizing the need for improved environmental surveillance. Vibrios are naturally occurring in aquatic environments globally, but traditional metagenomic methods for detecting and typing pathogenicVibriospp. are challenged by their presence in relatively low abundance and ability to persist in a viable but nonculturable state. In the study reported here, hybridization capture sequencing was employed to profile low-abundanceVibriospp. in metagenomic samples, namely water and oysters collected from the Chesapeake Bay. This approach was evaluated in parallel with traditional whole-community shotgun sequencing and whole-genome sequencing ofVibrio parahaemolyticusandVibrio vulnificusstrains isolated from the samples. Results suggest pathogenicVibriospp. in aquatic ecosystems may be far more common than currently understood, when multiple methods are considered for environmental surveillance.

Microbiology↗
DOE OSTI2025

Visolis Microbial Chemical Intermediate Library Screening (CRADA Final Report)

Visolis is developing a commercial process for the bioproduction of a chemical intermediate, towards the derivative production of a portfolio of bio-based chemicals with large application potential, from drop-in commodity chemicals, to innovative specialty chemical offerings, to materials for multiple end markets and industries, which will ultimately benefit consumers and the public. Visolis already has several variants of a microbe that produce the chemical intermediate at distinct levels (i.e. high, medium, and low). They are currently pursuing transcriptomics analysis for some of these strains, towards a better understanding of the biology behind what makes certain strains perform better than others. There exist genome-scale library approaches for the microbe that could be used to screen gene disruption, overexpression, or repression candidates for perturbations to the production of the chemical intermediate. These approaches often reveal opportunities for further production improvement that are not otherwise accessible using hypothesis-driven metabolic engineering approaches. However, Visolis, while it could generate or procure such genome-scale libraries, does not have the automated strain engineering workflows required to screen thousands of variants, obtaining a production phenotype and a genotype for each. The purpose of this collaboration is to use LBNL and SNL capabilities to enable Visolis to effectively screen thousands of genome-scale library strain variants for phenotype/genotype relationships that will complement Visolis’ transcriptomic investigations.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
NASA NTRS1995

Sequence, molecular properties, and chromosomal mapping of mouse lumican

PURPOSE. Lumican is a major proteoglycan of vertebrate cornea. This study characterizes mouse lumican, its molecular form, cDNA sequence, and chromosomal localization. METHODS. Lumican sequence was determined from cDNA clones selected from a mouse corneal cDNA expression library using a bovine lumican cDNA probe. Tissue expression and size of lumican mRNA were determined using Northern hybridization. Glycosidase digestion followed by Western blot analysis provided characterization of molecular properties of purified mouse corneal lumican. Chromosomal mapping of the lumican gene (Lcn) used Southern hybridization of a panel of genomic DNAs from an interspecific murine backcross. RESULTS. Mouse lumican is a 338-amino acid protein with high-sequence identity to bovine and chicken lumican proteins. The N-terminus of the lumican protein contains consensus sequences for tyrosine sulfation. A 1.9-kb lumican mRNA is present in cornea and several other tissues. Antibody against bovine lumican reacted with recombinant mouse lumican expressed in Escherichia coli and also detected high molecular weight proteoglycans in extracts of mouse cornea. Keratanase digestion of corneal proteoglycans released lumican protein, demonstrating the presence of sulfated keratan sulfate chains on mouse corneal lumican in vivo. The lumican gene (Lcn) was mapped to the distal region of mouse chromosome 10. The Lcn map site is in the region of a previously identified developmental mutant, eye blebs, affecting corneal morphology. CONCLUSIONS. This study demonstrates sulfated keratan sulfate proteoglycan in mouse cornea and describes the tools (antibodies and cDNA) necessary to investigate the functional role of this important corneal molecule using naturally occurring and induced mutants of the murine lumican gene.

NASA Discipline Cell Biology↗
NASA NTRS2006

Anaerovirgula multivorans gen. nov., sp. nov., a Novel Spore-Forming, Alkaliphilic Anaerobe Isolated from Owens Lake, California, USA

A novel, alkaliphilic, obligately anaerobic bacterium, strain SCAT, was isolated from mud sediments of a soda lake in California, USA. The rod-shaped cells were motile, Gram-positive, formed spores and were 0.4-0.5x2.5-5.0 micrometers in size. Growth occurred within the pH range 6.7-10.0 and was optimal at pH 8.5. The temperature range for growth was 10-45 degrees C, with optimal growth at 35 degrees C. NaCl was required for growth. Growth occurred at 0.5-9.0% (w/v) NaCl and was optimal at 1-2% (w/v). The novel isolate was a catalase-negative chemo-organoheterotroph that fermented sugars, proteolysis products, some organic and amino acids, glycerol, d-cellobiose and cellulose. It was also capable of growth by the Stickland reaction. Strain SCAT was sensitive to tetracycline, chloramphenicol, rifampicin and gentamicin, but it was resistant to ampicillin and kanamycin. The G+C content of the genomic DNA was 34.2 mol%. Major fatty acid components were C14:0, iso-C15:0, C16:1omega9c and C16:0. 16S rRNA gene sequence analysis of strain SCAT showed a similarity of approximately 97% with the type strains of Clostridium formicaceticum and Clostridium aceticum in clostridial cluster XI and a similarity of less than 94.2% to any other recognized Clostridium species and those of related genera in this cluster. Strain SCAT was clearly differentiated from C. formicaceticum and C. aceticum based on comparison of their phenotypic properties and fatty acid profiles, as well as low levels of DNA-DNA relatedness between strain SCAT and the type strains of these two species. Therefore, strain SCAT is considered to represent a novel species of a new genus, Anaerovirgula multivorans gen. nov., sp. nov., in clostridial cluster XI. The type strain is SCAT (=ATCC BAA-1084T=JCM 12857T=DSM 17722T=CIP 107910T).

Pikuta, Elena V.↗
NASA NTRS2016

GeneLab Phase 2: Integrated Search Data Federation of Space Biology Experimental Data

The GeneLab project is a science initiative to maximize the scientific return of omics data collected from spaceflight and from ground simulations of microgravity and radiation experiments, supported by a data system for a public bioinformatics repository and collaborative analysis tools for these data. The mission of GeneLab is to maximize the utilization of the valuable biological research resources aboard the ISS by collecting genomic, transcriptomic, proteomic and metabolomic (so-called omics) data to enable the exploration of the molecular network responses of terrestrial biology to space environments using a systems biology approach. All GeneLab data are made available to a worldwide network of researchers through its open-access data system. GeneLab is currently being developed by NASA to support Open Science biomedical research in order to enable the human exploration of space and improve life on earth. Open access to Phase 1 of the GeneLab Data Systems (GLDS) was implemented in April 2015. Download volumes have grown steadily, mirroring the growth in curated space biology research data sets (61 as of June 2016), now exceeding 10 TB/month, with over 10,000 file downloads since the start of Phase 1. For the period April 2015 to May 2016, most frequently downloaded were data from studies of Mus musculus (39) followed closely by Arabidopsis thaliana (30), with the remaining downloads roughly equally split across 12 other organisms (each 10 of total downloads). GLDS Phase 2 is focusing on interoperability, supporting data federation, including integrated search capabilities, of GLDS-housed data sets with external data sources, such as gene expression data from NIHNCBIs Gene Expression Omnibus (GEO), proteomic data from EBIs PRIDE system, and metagenomic data from Argonne National Laboratory's MG-RAST. GEO and MG-RAST employ specifications for investigation metadata that are different from those used by the GLDS and PRIDE (e.g., ISA-Tab). The GLDS Phase 2 system will implement a Google-like, full-text search engine using a Service-Oriented Architecture by utilizing publicly available RESTful web services Application Programming Interfaces (e.g., GEO Entrez Programming Utilities) and a Common Metadata Model (CMM) in order to accommodate the different metadata formats between the heterogeneous bioinformatics databases. GLDS Phase 2 completion with fully implemented capabilities will be made available to the general public in September 2017.

Space Biology↗
NASA NTRS2019

Nasa GeneLab Computomics Reveal Horizontal Gene Transfer on International Space Station Environmental Metagenomes

Prokaryotic lifeforms can be observed to demonstrate many keen adaptive advantages, perhaps facilitated by a nature simplistic relative to divergent domains of life. In particular, decompartmentalized gene expression facilitates adaptation by allowing free exchange of genetic material, albeit at the cost of increased susceptibility to genetic damage. Thus, these lifeforms must compensate by embracing diverse investment strategies in an attempt to “brute force” the evolvability equation through precipitous genesis, lean metabolic efficiency, and sheer population. This prokaryotic archetype also enables symbiotic relationships with secondary mobile genetic elements known as plasmids, which have been shown to drive evolution on rapid temporal scales through processes such as conjugation and transformation. This study attempts to decipher whether these mechanisms of horizontal gene transfer (HGT) are major factors in determining prokaryote fitness within a unique isolated environment, the International Space Station (ISS). The ISS Microbial Tracking (MT) project has generated a wealth of data concerning the successive reigns of microbial genera that appear to thrive amidst harsh conditions for life. Despite relatively higher doses of ionizing radiation as compared to Earth, complications associated with microgravity, and the anti-microbial mélange deployed, microbial life still persists in this environment. The NASA GeneLab serves as a data repository and analysis platform to enable researchers to access space flight factor related data. With the use of GeneLab’s modern computational suites (computomics), phylogenetic and functional genomic investigations of HGT events were conducted on the data generated from the MT-1 project. The putative data concerning the plasmid population (plasmidome) of the ISS was algorithmically derived and compared to those of habitats with similar environmental dynamics- such as living quarters and hospitals- to investigate whether these HGT elements may play crucial role(s) in shaping the microbiome of this closed habitat that serves as the only inhabited structure in space.

Bense, Nicholas↗
NASA NTRS2018

Detection of Antimicrobial Resistance Genes Associated With the International Space Station Environmental Surfaces

Antimicrobial resistance (AMR) is a global health issue. In an effort to minimize this threat to astronauts, who may be immunocompromised and thus at a greater risk of infection from antimicrobial resistant pathogens, a comprehensive study of the ISS “resistome’ was conducted. Using whole genome sequencing (WGS) and disc diffusion antibiotic resistance assays, 9 biosafety level 2 organisms isolated from the ISS were assessed for their antibiotic resistance. Molecular analysis of AMR genes from 24 surface samples collected from the ISS during 3 different sampling events over a span of a year were analyzed with Ion AmpliSeq™ and metagenomics. Disc diffusion assays showed that Enterobacter bugandensis strains were resistant to all 9 antibiotics tested and Staphylococcus haemolyticus being resistant to none. Ion AmpliSeq™ revealed that 123 AMR genes were found, with those responsible for beta-lactam and trimethoprim resistance being the most abundant and widespread. Using a variety of methods, the genes involved in antimicrobial resistance have been examined for the first time from the ISS. This information could lead to mitigation strategies to maintain astronaut health during long duration space missions when return to Earth for treatment is not possible.

Antimicrobial resistance↗
DOE OSTI2024

Systematic identification of transcriptional activation domains from non-transcription factor proteins in plants and yeast

Transcription factors can promote gene expression through activation domains. Whole-genome screens have systematically mapped activation domains in transcription factors but not in non-transcription factor proteins (e.g., chromatin regulators and coactivators). To fill this knowledge gap, we employed the activation domain predictor PADDLE to analyze the proteomes of Arabidopsis thaliana and Saccharomyces cerevisiae. We screened 18,000 predicted activation domains from >800 non-transcription factor genes in both species, confirming that 89% of candidate proteins contain active fragments. Our work enables the annotation of hundreds of nuclear proteins as putative coactivators, many of which have never been ascribed any function in plants. Analysis of peptide sequence compositions reveals how the distribution of key amino acids dictates activity. Finally, we validated short, "universal" activation domains with comparable performance to state-of-the-art activation domains used for genome engineering. Our approach enables the genome-wide discovery and annotation of activation domains that can function across diverse eukaryotes.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

SARS-CoV-2 wastewater variant surveillance: pandemic response leveraging FDA’s GenomeTrakr network

ABSTRACT Wastewater surveillance has emerged as a crucial public health tool for population-level pathogen surveillance. Supported by funding from the American Rescue Plan Act of 2021, the FDA‘s genomic epidemiology program, GenomeTrakr, was leveraged to sequence SARS-CoV-2 from wastewater sites across the United States. This initiative required the evaluation, optimization, development, and publication of new methods and analytical tools spanning sample collection through variant analyses. Version-controlled protocols for each step of the process were developed and published on protocols.io. A custom data analysis tool and a publicly accessible dashboard were built to facilitate real-time visualization of the collected data, focusing on the relative abundance of SARS-CoV-2 variants and sub-lineages across different samples and sites throughout the project. From September 2021 through June 2023, a total of 3,389 wastewater samples were collected, with 2,517 undergoing sequencing and submission to NCBI under the umbrella BioProject,PRJNA757291. Sequence data were released with explicit quality control (QC) tags on all sequence records, communicating our confidence in the quality of data. Variant analysis revealed wide circulation of Delta in the fall of 2021 and captured the sweep of Omicron and subsequent diversification of this lineage through the end of the sampling period. This project successfully achieved two important goals for the FDA’s GenomeTrakr program: first, contributing timely genomic data for the SARS-CoV-2 pandemic response, and second, establishing both capacity and best practices for culture-independent, population-level environmental surveillance for other pathogens of interest to the FDA. IMPORTANCE This paper serves two primary objectives. First, it summarizes the genomic and contextual data collected during a Covid-19 pandemic response project, which utilized the FDA’s laboratory network, traditionally employed for sequencing foodborne pathogens, for sequencing SARS-CoV-2 from wastewater samples. Second, it outlines best practices for gathering and organizing population-level next generation sequencing (NGS) data collected for culture-free, surveillance of pathogens sourced from environmental samples.

Microbiology↗