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At least 1,387 records · Page 77
All sourcesNASA NTRSDOE OSTI
DOE OSTI2024

Relationships between Habitual Polyphenol Consumption and Gut Microbiota in the INCLD Health Cohort

While polyphenol consumption is often associated with an increased abundance of beneficial microbes and decreased opportunistic pathogens, these relationships are not completely described for polyphenols consumed via habitual diet, including culinary herb and spice consumption. This analysis of the International Cohort on Lifestyle Determinants of Health (INCLD Health) cohort uses a dietary questionnaire and 16s microbiome data to examine relationships between habitual polyphenol consumption and gut microbiota in healthy adults (n = 96). In this exploratory analysis, microbial taxa, but not diversity measures, differed by levels of dietary polyphenol consumption. Taxa identified as exploratory biomarkers of daily polyphenol consumption (mg/day) included Lactobacillus, Bacteroides, Enterococcus, Eubacterium ventriosum group, Ruminococcus torques group, and Sutterella. Taxa identified as exploratory biomarkers of the frequency of polyphenol-weighted herb and spice use included Lachnospiraceae UCG-001, Lachnospiraceae UCG-004, Methanobrevibacter, Lachnoclostridium, and Lachnotalea. Several of the differentiating taxa carry out activities important for human health, although out of these taxa, those with previously described pro-inflammatory qualities in certain contexts displayed inverse relationships with polyphenol consumption. Our results suggest that higher quantities of habitual polyphenol consumption may support an intestinal environment where opportunistic and pro-inflammatory bacteria are represented in a lower relative abundance compared to those with less potentially virulent qualities.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2024

Evaluation of Inactivation Methods for Rift Valley Fever Virus in Mouse Microglia

Rift Valley fever phlebovirus (RVFV) is a highly pathogenic mosquito-borne virus with bioweapon potential due to its ability to be spread by aerosol transmission. Neurological symptoms are among the worst outcomes of infection, and understanding of pathogenesis mechanisms within the brain is limited. RVFV is classified as an overlap select agent by the CDC and USDA; therefore, experiments involving fully virulent strains of virus are tightly regulated. Here, we present two methods for inactivation of live virus within samples derived from mouse microglia cells using commercially available kits for the preparation of cells for flow cytometry and RNA extraction. Using the flow cytometry protocol, we demonstrate key differences in the response of primary murine microglia to infection with fully virulent versus attenuated RVFV.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2025

Virulence and Genetic Diversity of Puccinia spp., Causal Agents of Rust on Switchgrass (Panicum virgatum L.) in the USA

Switchgrass (Panicum virgatum L.) is an important cellulosic biofuel grass native to North America. Rust, caused by Puccinia spp. is the most predominant disease of switchgrass and has the potential to impact biomass conversion. In this study, virulence patterns were determined on a set of 38 switchgrass genotypes for 14 single-spore rust isolates from 14 field samples collected in seven states. Single nucleotide polymorphism (SNP) variation was also assessed in 720 sequenced cloned amplicons representing 654 base pairs of the elongation factor 1-α gene from the field samples. Five major haplotypes were identified differing by 11 out of the 39 SNP positions identified. STRUCTURE, Principal Coordinate Analysis, and phylogenetic analyses divided the rust population into two genetic clusters. Virginia and Georgia had the highest and lowest rust genetic diversity, respectively. Only nine accessions showed a differential disease response between the 14 isolates, allowing the identification of eight races, differing by 1–3 virulence factors. Overall, the results suggested clonal reproduction of the pathogen and a North–South differentiation via local adaptation. However, similar haplotypes and races were also recovered from several states, suggesting migration events, and highlighting the need to further investigate the switchgrass rust population structure and evolution in the USA.

Bahri, Bochra A. (ORCID:0000000159055880)↗
DOE OSTI2026

In Vitro Selection of Antibodies Targeting Yersinia pestis Membrane Lipids Using Nanodisc-Based Antigen Presentation

Proteins are the most common targets for antibody discovery and vaccine development, but their sequence variability can limit the breadth of resulting antigens. Lipids represent an alternative class of antigens due to their structural conservation and roles in host–pathogen interactions. Here, we describe the development and optimization of an in vitro antibody selection workflow using lipid-containing nanodiscs as antigen presentation platforms to enable phage and yeast display selections under conditions adapted for these non-protein targets. Lipopolysaccharide (LPS) nanodiscs were first used as a model system to evaluate selection strategies, including competitive and subtractive approaches to reduce non-specific binders, yielding peptide and single-chain variable fragment (scFv) binders that were affinity matured to improve binding signals. The same approach was subsequently used to select scFv antibodies that recognize lipid nanodiscs prepared from Yersinia pestis membrane lipid extracts. These antibodies show binding to lipid nanodiscs derived from Y. pestis, with evidence of selectivity relative to control nanodiscs. Overall, this work establishes a workflow for antibody selection against lipid-containing nanodisc antigens and highlights practical considerations associated with these targets. The approach may be useful for generating affinity reagents to membrane-associated lipids, although further characterization is required to define antigen specificity and functional activity.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2020

Comparison of the Transcriptome Response within the Swine Tracheobronchial Lymphnode Following Infection with PRRSV, PCV-2 or IAV-S.

Porcine reproductive and respiratory syndrome virus (PRRSV) is a major respiratory pathogen of swine that has become extremely costly to the swine industry worldwide, often causing losses in production and animal life due to their ease of spread. However, the intracellular changes that occur in pigs following viral respiratory infections are still scantily understood for PRRSV, as well as other viral respiratory infections. The aim of this study was to acquire a better understanding of the PRRS disease by comparing gene expression changes that occur in tracheobronchial lymph nodes (TBLN) of pigs infected with either porcine reproductive and respiratory syndrome virus (PRRSV), porcine circovirus type 2 (PCV-2), or swine influenza A virus (IAV-S) infections. The study identified and compared gene expression changes in the TBLN of 80 pigs following infection by PRRSV, PCV-2, IAV-S, or sham inoculation. Total RNA was pooled for each group and time-point (1, 3, 6, and 14 dpi) to make 16 libraries—analyses are by Digital Gene Expression Tag Profiling (DGETP). The data underwent standard filtering to generate a list of sequence tag raw counts that were then analyzed using multidimensional and differential expression statistical tests. The results showed that PRRSV, IAV-S and PCV-2 viral infections followed a clinical course in the pigs typical of experimental infection of young pigs with these viruses. Gene expression results echoed this course, as well as uncovered genes related to intersecting and unique host immune responses to the three viruses. By testing and observing the host response to other respiratory viruses, our study has elucidated similarities and differences that can assist in the development of vaccines and therapeutics that shorten or prevent a chronic PRRSV infection.

Miller↗
DOE OSTI2021

Variation in Gene Expression between Two Sorghum bicolor Lines Differing in Innate Immunity Response

Microbe associated molecular pattern (MAMPs) triggered immunity (MTI) is a key component of the plant innate immunity response to microbial recognition. However, most of our current knowledge of MTI comes from model plants (i.e., Arabidopsis thaliana) with comparatively less work done using crop plants. In this work, we studied the MAMP triggered oxidative burst (ROS) and the transcriptional response in two Sorghum bicolor genotypes, BTx623 and SC155-14E. SC155-14E is a line that shows high anthracnose resistance and the line BTx623 is susceptible to anthracnose. Our results revealed a clear variation in gene expression and ROS in response to either flagellin (flg22) or chitin elicitation between the two lines. While the transcriptional response to each MAMP and in each line was unique there was a considerable degree of overlap, and we were able to define a core set of genes associated with the sorghum MAMP transcriptional response. The GO term and KEGG pathway enrichment analysis discovered more immunity and pathogen resistance related DEGs in MAMP treated SC155-14E samples than in BTx623 with the same treatment. The results provide a baseline for future studies to investigate innate immunity pathways in sorghum, including efforts to enhance disease resistance.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

Auxin-Producing Bacteria from Duckweeds Have Different Colonization Patterns and Effects on Plant Morphology

The role of auxin in plant–microbe interaction has primarily been studied using indole-3-acetic acid (IAA)-producing pathogenic or plant-growth-promoting bacteria. However, the IAA biosynthesis pathway in bacteria involves indole-related compounds (IRCs) and intermediates with less known functions. Here, we seek to understand changes in plant response to multiple plant-associated bacteria taxa and strains that differ in their ability to produce IRCs. We had previously studied 47 bacterial strains isolated from several duckweed species and determined that 79% of these strains produced IRCs in culture, such as IAA, indole lactic acid (ILA), and indole. Using Arabidopsis thaliana as our model plant with excellent genetic tools, we performed binary association assays on a subset of these strains to evaluate morphological responses in the plant host and the mode of bacterial colonization. Of the 21 tested strains, only four high-quantity IAA-producing Microbacterium strains caused an auxin root phenotype. Compared to the commonly used colorimetric Salkowski assay, auxin concentration determined by LC–MS was a superior indicator of a bacteria’s ability to cause an auxin root phenotype. Studies with the auxin response mutant axr1-3 provided further genetic support for the role of auxin signaling in mediating the root morphology response to IAA-producing bacteria strains. Interestingly, our microscopy results also revealed new evidence for the role of the conserved AXR1 gene in endophytic colonization of IAA-producing Azospirillum baldaniorum Sp245 via the guard cells.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

An Improved Sampling and Baiting Method for Phytophthora tropicalis and P. heveae Detection in Macadamia integrifolia

Macadamia nuts are, economically, the second most important crop in the state of Hawai’i. A recent decline in yield and acreage has been attributed to insect damage and diseases such as Macadamia Quick Decline (MQD) caused by Phytophthora tropicalis and P. heveae. To develop an improved methodology for the diagnosis and treatment of MQD, investigations were undertaken to better understand the pathosystem of the disease. These investigations included sampling from multiple locations from sectioned trees utilizing two methods of tissue collection and isolations using two baiting techniques. The collection of tissue from the cambium and phloem of trees after scraping away the bark and in locations of recent or current sap exudation using a narrow diameter steel awl proved to be an efficient means for the molecular detection of the MQD pathogens from infected trees exhibiting MQD symptoms. In addition, a more efficient and cost-effective baiting method using apple puree was developed.

Plant Sciences↗
DOE OSTI2024

Chemical Characterization and Antimicrobial Activity of Green Propolis from the Brazilian Caatinga Biome

Green propolis, particularly from the unique flora of the Brazilian Caatinga biome, has gained significant interest due to its diverse chemical composition and biological activities. This study focuses on the chemical characterization and antimicrobial evaluation of Caatinga green propolis. Twelve compounds were isolated through different chromatographic techniques, including flavanones (naringenin, 7-O-methyleriodictyol, sakuranetin), flavones (hispidulin, cirsimaritin), flavonols (quercetin, quercetin-3-methyl ether, kaempferol, 6-methoxykaempferol, viscosine, penduletin), and one chalcone (kukulkanin B). Using liquid chromatography–quadrupole time-of-flight tandem mass spectrometry (LC-QToF-MS), a total of 55 compounds excluding reference standards were tentatively identified, which include flavonoids, phenolic acids derivatives, and alkaloids, with flavonols, flavanones, and flavones being predominant. Antimicrobial testing against pathogens revealed that the crude extract exhibited low inhibitory activity, against Gram-positive bacteria, including methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant Enterococcus faecium (VRE) (IC50: 148.4 and 120.98 µg/mL, respectively). Although the isolated compounds showed limited individual activity, a fraction containing sakuranetin and penduletin (Fraction 8) exhibited moderated activity against Cryptococcus neoformans (IC50: 47.86 µg/mL), while a fraction containing quercetin and hispidulin showed moderated activity against VRE (IC50: 16.99 µg/mL). These findings highlight the potential application of Caatinga green propolis as an antimicrobial agent, particularly against resistant bacterial strains, and underscore the importance of synergistic interactions between compounds in enhancing biological effects.

Aldana-Mejía, Jennyfer A. (ORCID:0000000307361899)↗
DOE OSTI2020

Post-Synthetic Reduction of Pectin Methylesterification Causes Morphological Abnormalities and Alterations to Stress Response in Arabidopsis thaliana

Pectin is a critical component of the plant cell wall, supporting wall biomechanics and contributing to cell wall signaling in response to stress. The plant cell carefully regulates pectin methylesterification with endogenous pectin methylesterases (PMEs) and their inhibitors (PMEIs) to promote growth and protect against pathogens. We expressed Aspergillus nidulans pectin methylesterase (AnPME) in Arabidopsis thaliana plants to determine the impacts of methylesterification status on pectin function. Plants expressing AnPME had a roughly 50% reduction in methylester content compared with control plants. AnPME plants displayed a severe dwarf phenotype, including small, bushy rosettes and shorter roots. This phenotype was caused by a reduction in cell elongation. Cell wall composition was altered in AnPME plants, with significantly more arabinose and significantly less galacturonic acid, suggesting that plants actively monitor and compensate for altered pectin content. Cell walls of AnPME plants were more readily degraded by polygalacturonase (PG) alone but were less susceptible to treatment with a mixture of PG and PME. AnPME plants were insensitive to osmotic stress, and their susceptibility to Botrytis cinerea was comparable to wild type plants despite their compromised cell walls. This is likely due to upregulated expression of defense response genes observed in AnPME plants. These results demonstrate the importance of pectin in both normal growth and development, and in response to biotic and abiotic stresses.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

One Health Approach to Tackle Microbial Contamination on Poultries—A Systematic Review

This study reports the search of available data published regarding microbial occupational exposure assessment in poultries, following the PRISMA methodology. Air collection through filtration was the most frequently used. The most commonly used passive sampling method was material collection such as dust, cages, soils, sediment, and wastewater. Regarding assays applied, the majority of studies comprised culture-based methods, but molecular tools were also frequently used. Screening for antimicrobial susceptibility was performed only for bacteria; cytotoxicity, virological and serological assays were also performed. Most of the selected studies focused on bacteria, although fungi, endotoxins, and β-glucans were also assessed. The only study concerning fungi and mycotoxins reported the carcinogenic mycotoxin AFB1. This study gives a comprehensive overview of microbial contamination in the poultry industry, emphasizing this setting as a potential reservoir of microbial pathogens threatening human, animal, and environmental health. Additionally, this research helps to provide a sampling and analysis protocol proposal to evaluate the microbiological contamination in these facilities. Few articles were found reporting fungal contamination in poultry farms worldwide. In addition, information concerning fungal resistance profile and mycotoxin contamination remain scarce. Overall, a One Health approach should be incorporated in exposure assessments and the knowledge gaps identified in this paper should be addressed in further research.

Gomes, Bianca (ORCID:0000000262646072)↗
DOE OSTI2020

Portable Rabies Virus Sequencing in Canine Rabies Endemic Countries Using the Oxford Nanopore MinION

As countries with endemic canine rabies progress towards elimination by 2030, it will become necessary to employ techniques to help plan, monitor, and confirm canine rabies elimination. Sequencing can provide critical information to inform control and vaccination strategies by identifying genetically distinct virus variants that may have different host reservoir species or geographic distributions. However, many rabies testing laboratories lack the resources or expertise for sequencing, especially in remote or rural areas where human rabies deaths are highest. We developed a low-cost, high throughput rabies virus sequencing method using the Oxford Nanopore MinION portable sequencer. A total of 259 sequences were generated from diverse rabies virus isolates in public health laboratories lacking rabies virus sequencing capacity in Guatemala, India, Kenya, and Vietnam. Phylogenetic analysis provided valuable insight into rabies virus diversity and distribution in these countries and identified a new rabies virus lineage in Kenya, the first published canine rabies virus sequence from Guatemala, evidence of rabies spread across an international border in Vietnam, and importation of a rabid dog into a state working to become rabies-free in India. Taken together, our evaluation highlights the MinION’s potential for low-cost, high volume sequencing of pathogens in locations with limited resources.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2020

NCBI’s Virus Discovery Codeathon: Building “FIVE” —The Federated Index of Viral Experiments API Index

Viruses represent important test cases for data federation due to their genome size and the rapid increase in sequence data in publicly available databases. However, some consequences of previously decentralized (unfederated) data are lack of consensus or comparisons between feature annotations. Unifying or displaying alternative annotations should be a priority both for communities with robust entry representation and for nascent communities with burgeoning data sources. To this end, during this three-day continuation of the Virus Hunting Toolkit codeathon series (VHT-2), a new integrated and federated viral index was elaborated. This Federated Index of Viral Experiments (FIVE) integrates pre-existing and novel functional and taxonomy annotations and virus–host pairings. Variability in the context of viral genomic diversity is often overlooked in virus databases. As a proof-of-concept, FIVE was the first attempt to include viral genome variation for HIV, the most well-studied human pathogen, through viral genome diversity graphs. As per the publication of this manuscript, FIVE is the first implementation of a virus-specific federated index of such scope. FIVE is coded in BigQuery for optimal access of large quantities of data and is publicly accessible. Many projects of database or index federation fail to provide easier alternatives to access or query information. To this end, a Python API query system was developed to enhance the accessibility of FIVE.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

Single-Dose Immunogenic DNA Vaccines Coding for Live-Attenuated Alpha- and Flaviviruses

Single-dose, immunogenic DNA (iDNA) vaccines coding for whole live-attenuated viruses are reviewed. This platform, sometimes called immunization DNA, has been used for vaccine development for flavi- and alphaviruses. An iDNA vaccine uses plasmid DNA to launch live-attenuated virus vaccines in vitro or in vivo. When iDNA is injected into mammalian cells in vitro or in vivo, the RNA genome of an attenuated virus is transcribed, which starts replication of a defined, live-attenuated vaccine virus in cell culture or the cells of a vaccine recipient. In the latter case, an immune response to the live virus vaccine is elicited, which protects against the pathogenic virus. Unlike other nucleic acid vaccines, such as mRNA and standard DNA vaccines, iDNA vaccines elicit protection with a single dose, thus providing major improvement to epidemic preparedness. Still, iDNA vaccines retain the advantages of other nucleic acid vaccines. In summary, the iDNA platform combines the advantages of reverse genetics and DNA immunization with the high immunogenicity of live-attenuated vaccines, resulting in enhanced safety and immunogenicity. This vaccine platform has expanded the field of genetic DNA and RNA vaccines with a novel type of immunogenic DNA vaccines that encode entire live-attenuated viruses.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2024

Recommendations for Uniform Variant Calling of SARS-CoV-2 Genome Sequence across Bioinformatic Workflows

Genomic sequencing of clinical samples to identify emerging variants of SARS-CoV-2 has been a key public health tool for curbing the spread of the virus. As a result, an unprecedented number of SARS-CoV-2 genomes were sequenced during the COVID-19 pandemic, which allowed for rapid identification of genetic variants, enabling the timely design and testing of therapies and deployment of new vaccine formulations to combat the new variants. However, despite the technological advances of deep sequencing, the analysis of the raw sequence data generated globally is neither standardized nor consistent, leading to vastly disparate sequences that may impact identification of variants. Here, we show that for both Illumina and Oxford Nanopore sequencing platforms, downstream bioinformatic protocols used by industry, government, and academic groups resulted in different virus sequences from same sample. These bioinformatic workflows produced consensus genomes with differences in single nucleotide polymorphisms, inclusion and exclusion of insertions, and/or deletions, despite using the same raw sequence as input datasets. Here, we compared and characterized such discrepancies and propose a specific suite of parameters and protocols that should be adopted across the field. Consistent results from bioinformatic workflows are fundamental to SARS-CoV-2 and future pathogen surveillance efforts, including pandemic preparation, to allow for a data-driven and timely public health response.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2023

The Polymorphic Membrane Protein G Has a Neutral Effect and the Plasmid Glycoprotein 3 an Antagonistic Effect on the Ability of the Major Outer Membrane Protein to Elicit Protective Immune Responses against a Chlamydia muridarum Respiratory Challenge

Chlamydia trachomatis is the most common bacterial sexually transmitted pathogen. The number of chlamydial infections continuous to increase and there is an urgent need for a safe and efficacious vaccine. To assess the ability of the Chlamydia muridarum polymorphic membrane protein G (PmpG) and the plasmid glycoprotein 3 (Pgp3) as single antigens, and in combination with the major outer-membrane protein (MOMP) to induce protection, BALB/c mice were immunized utilizing CpG-1826 and Montanide ISA 720 VG as adjuvants. Following vaccination with MOMP, significant humoral and cell-mediated immune responses were observed, while immunization with PmpG, or Pgp3, elicited weaker immune responses. Weaker immune responses were induced with MOMP+Pgp3 compared with MOMP alone. Following the intranasal challenge with C. muridarum, mice vaccinated with MOMP showed robust protection against body-weight loss, inflammatory responses in the lungs and number of Chlamydia recovered from the lungs. PmpG and Pgp3 elicited weaker protective responses. Mice immunized with MOMP+PmpG, were no better protected than animals vaccinated with MOMP only, while Pgp3 antagonized the protection elicited by MOMP. In conclusion, PmpG and Pgp3 elicited limited protective immune responses in mice against a respiratory challenge with C. muridarum and failed to enhance the protection induced by MOMP alone. The virulence of Pgp3 may result from its antagonistic effect on the immune protection induced by MOMP.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2024

CT584 Is Not a Protective Vaccine Antigen against Respiratory Chlamydial Challenge in Mice

Background: Chlamydia trachomatis is the most prevalent bacterial sexually transmitted pathogen in humans worldwide. Since chlamydial infection is largely asymptomatic with the potential for serious complications, a preventative vaccine is likely the most viable long-term answer to this public health threat. Cell-free protein synthesis (CFPS) utilizes the cellular protein manufacturing machinery decoupled from the requirement for maintaining cellular viability, offering the potential for flexible, rapid, and decentralized production of recombinant protein vaccine antigens. Methods: Here, we use CFPS to produce the full-length putative chlamydial type three secretion system (T3SS) needle-tip protein, CT584, for evaluation as a vaccine antigen in mouse models. High-speed atomic force microscopy (HS-AFM) (RIBM, Tsukuba, Japan) imaging and computer simulations confirm that CFPS-produced CT584 retains a native-like structure prior to immunization. Female mice were primed with CT584 adjuvanted with CpG-1826 intranasally (i.n.) or CpG-1826 + Montanide ISA 720 intramuscularly (i.m.), followed four weeks later by an i.m. boost before respiratory challenge with 10 4 inclusion forming units (IFU) of Chlamydia muridarum. Results: Immunization with CT584 generated robust antibody responses but weak cell-mediated immunity and failed to protect against i.n. challenge as demonstrated by body weight loss, increased lung weights, and the presence of high numbers of IFUs in the lungs. Conclusion: While CT584 was not a protective vaccine candidate, the speed and flexibility with which CFPS can be used to produce other potential chlamydial antigens make it an attractive technique for antigen production.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2020

Vaccine Advances against Venezuelan, Eastern, and Western Equine Encephalitis Viruses

Vaccinations are a crucial intervention in combating infectious diseases. The three neurotropic Alphaviruses, Eastern (EEEV), Venezuelan (VEEV), and Western (WEEV) equine encephalitis viruses, are pathogens of interest for animal health, public health, and biological defense. In both equines and humans, these viruses can cause febrile illness that may progress to encephalitis. Currently, there are no licensed treatments or vaccines available for these viruses in humans. Experimental vaccines have shown variable efficacy and may cause severe adverse effects. Here, we outline recent strategies used to generate vaccines against EEEV, VEEV, and WEEV with an emphasis on virus-vectored and plasmid DNA delivery. Despite candidate vaccines protecting against one of the three viruses, few studies have demonstrated an effective trivalent vaccine. We evaluated the potential of published vaccines to generate cross-reactive protective responses by comparing DNA vaccine sequences to a set of EEEV, VEEV, and WEEV genomes and determining the vaccine coverages of potential epitopes. Finally, we discuss future directions in the development of vaccines to combat EEEV, VEEV, and WEEV.

59 BASIC BIOLOGICAL SCIENCES↗