Engineering Papers⌕ Search

SEARCH · Engineering Papers

Results for “Virus”

Search indexed NASA NTRS and DOE OSTI research on propulsion, heat transfer, battery materials and energy systems. Follow report and document links to the original sources.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 631 records · Page 35

Characterization of a monoclonal antibody specific to California serogroup orthobunyaviruses and development as a chimeric immunoglobulin M-positive control in human diagnostics

ABSTRACT California serogroup viruses (CSGVs) of medical importance in the United States include La Crosse virus, Jamestown Canyon virus (JCV), California encephalitis virus, and snowshoe hare virus. Current diagnosis of CSGVs relies heavily on serologic techniques for detecting immunoglobulin M (IgM), an indication of a recent CSGV infection. However, human-positive control sera reactive to viruses in the serogroup are scarce because detection of recent infections is rare. Here, we describe the development of new murine monoclonal antibodies (MAbs) reactive to CSGVs and the engineering of a human-murine chimeric antibody by combining the variable regions of the broadly CSGV cross-reactive murine MAb, 3-3B6/2-3B2 and the constant region of the human IgM. MAb 3-3B6/2-3B2 recognizes a tertiary epitope on the Gn/Gc heterodimer, and epitopes important in JCV neutralization were mapped to the Gc glycoprotein. This engineered human IgM constitutively expressed in a HEK-293 stable cell line can replace human-positive control sera in diagnostic serological techniques such as IgM antibody capture enzyme-linked immunosorbent assay (MAC-ELISA). Compared to the parent murine MAbs, the human-chimeric IgM antibody had identical serological activity to CSGVs in ELISA and demonstrated equivalent reactivity compared to human immune sera in the MAC-ELISA. Importance Orthobunyaviruses in the California serogroup cause severe neurological disease in children and adults. While these viruses are known to circulate widely in North America, their occurrence is rare. Serological testing for CSGVs is hindered by the limited availability and volumes of human-positive specimens needed as controls in serologic assays. Here, we described the development of a murine monoclonal antibody cross-reactive to CSGVs engineered to contain the variable regions of the murine antibody on the backbone of human IgM. The chimeric IgM produced from the stably expressing HEK293 cell line was evaluated for use as a surrogate human-positive control in a serologic diagnostic test.

Microbiology↗

Sticky roots--implications of widespread, cryptic, viral infection of plants in natural and managed ecosystems for soil carbon processing in the rhizosphere

Plants strongly influence soil properties through rhizodeposition, in which exudates diffuse from roots, additional secretions are actively released, and root cells are sloughed into the soil. This contribution by plants of carbon compounds belowground is at the core of soil health, water holding capacity, and the soil carbon storage that pulls carbon dioxide out of the atmosphere. Once in soil, organic matter can bind with minerals such as iron hydroxides, where it can be protected from microbial attack for millenia, preserving very large terrestrial soil carbon pools. However, those same compounds contributed by roots to soil may also destabilize the long-term protective associations of SOM with minerals, making that soil organic matter (SOM) more vulnerable to microbial attack and decomposition. Plant roots thus influence both the buildup and breakdown of soil carbon pools. DOE’s E3SM Land Model (ELM) includes a representation of soil carbon storage on minerals, but the potential vulnerability of SOM–mineral associations to effects of rhizodeposition is not yet represented in ELM. To begin testing for this effect of rhizodeposition on soil carbon storage and decomposition, we worked to develop a novel approach during this TES Exploratory project DE-SC0019142 – we harnessed the power of plant viral infection. We examined whether plant virus infection can serve as a tool to intensify rhizodeposition at the root surface, and therefore possibly intensify mobilization of SOM from minerals making it visible to our analytical techniques. Viral infection is widespread in terrestrial ecosystems; 25-70% of plants have virus infection, yet the influence of such infection on root traits and terrestrial soil carbon dynamics remains largely unexplored. We used two plant hosts: the annual Avena sativa (oats) and the genetically tractable, model grass Brachypodium distachyon. These grasses were infected with the broad host range virus Barley Yellow Dwarf Virus (BYDV) via aphids (Rhopalosiphum padi). BYDV infects at least 150 grass species in agricultural and natural ecosystems, and in previous experiments, oats infected with BYDV had roots that were very sticky to the touch, strongly suggesting that infection altered rhizodeposition. We developed this new experimental approach mostly in a one virus (Barley Yellow Dwarf Virus)–one plant (Avena sativa) system. (Several effects of infection in a Brachypodium-BYDV system were similar in nature to effects on Avena sativa, but were more variable.) In the BYDV-Avena system, we developed protocols for consistently infecting target plants (and avoiding infection of control plants) using aphid caging on leaves. We measured that infected plants exhibited reduced photosynthesis, plant (including root) biomass, and root:shoot ratio, as well as simplified root system architecture. We established procedures for sampling the organic compounds carried specifically in phloem (vascular tissue) of leaves and roots, using aphid stylectomy. We used FTICR-MS, Orbitrap GC-MS, and LC-MS/MS to analyze organic compounds in phloem, liquid around roots of plants grown hydroponically, and pore water around roots in soil, and found differences in the compounds in solution bathing roots when infected and uninfected plants were grown hydroponically. Finally, we synthesized isotopically-labeled mineral–organic matter (MAOM) associations in the lab and developed assays using them in solution and in soil. Assays quantified the extent and rate of mineralization of labeled MAOM that was mobilized by functionally distinct rhizodeposits and then attacked by microbes. Two mechanisms for MAOM mobilization emerged, with distinct dynamics. During “direct” mobilization, rhizodeposits such as the strong ligand oxalic acid could drive rapid dissolution of minerals, mobilizing MAOM. During “indirect” mobilization, rhizodeposits such as the simple sugar glucose did not attack minerals directly but instead intensified microbial activity, which led to mobilization via changes in e.g. pH, Eh, and microbial metabolite production (Li et al. 2021). Mechanistic understanding derived from these data and our ongoing experiments using these techniques will inform future development of ELM. Plant roots not only contribute newly fixed organic compounds to soils, but also root activities can drive mineralization of the carbon and nutrients mobilized off minerals via “indirect” or “direct” mechanisms. Using viral infection as a new tool, ongoing combined experimentation and modeling will explore the strength and larger-scale significance of the cascade of processes from rhizodeposition to MAOM mobilization for soil carbon storage and nutrient cycling in terrestrial ecosystems. And if viral infection leads quite generally to “sticky roots”, our perception of the potential importance of prevalent virus infection in terrestrial landscapes will be transformed.

54 ENVIRONMENTAL SCIENCES↗

Multimerization of Ebola GPΔmucin on protein nanoparticle vaccines has minimal effect on elicitation of neutralizing antibodies

Ebola virus (EBOV), a member of the Filoviridae family of viruses and a causative agent of Ebola Virus Disease (EVD), is a highly pathogenic virus that has caused over twenty outbreaks in Central and West Africa since its formal discovery in 1976. The only FDA-licensed vaccine against Ebola virus, rVSV-ZEBOV-GP (Ervebo®), is efficacious against infection following just one dose. However, since this vaccine contains a replicating virus, it requires ultra-low temperature storage which imparts considerable logistical challenges for distribution and access. Additional vaccine candidates could provide expanded protection to mitigate current and future outbreaks. Here, we designed and characterized two multimeric protein nanoparticle subunit vaccines displaying 8 or 20 copies of GPΔmucin, a truncated form of the EBOV surface protein GP. Single-dose immunization of mice with GPΔmucin nanoparticles revealed that neutralizing antibody levels were roughly equivalent to those observed in mice immunized with non-multimerized GPΔmucin trimers. These results suggest that some protein subunit antigens do not elicit enhanced antibody responses when displayed on multivalent scaffolds and can inform next-generation design of stable Ebola virus vaccine candidates.

60 APPLIED LIFE SCIENCES↗

Proteome-wide characterization of PTMs reveals host cell responses to viral infection and identifies putative antiviral drug targets

Post-translational modifications (PTMs) are biochemical modifications that can significantly alter protein structure, function, stability, localization, and interactions with other molecules, thereby activating or inactivating intracellular processes. A growing body of research has begun to highlight the role of PTMs, including phosphorylation, ubiquitination, acetylation, and redox modifications, during virus-host interactions. Collectively, these PTMs regulate key steps in mounting the host immune response and control critical host pathways required for productive viral replication. This has led to the conception of antiviral therapeutics that focus on controlling host protein PTMs, potentially offering pathogen-agnostic treatment options and revolutionizing our capacity to prevent virus transmission. On the other hand, viruses can hijack the host cellular PTM machinery to modify viral proteins in promoting viral replication and evading immune surveillance. PTM regulation during virus-host interactions is complex and poorly mapped, and the development of effective PTM-targeted antiviral drugs will require a more comprehensive understanding of the cellular pathways essential for virus replication. In this review, we discuss the roles of PTMs in virus infection and how technological advances in mass spectrometry-based proteomics can capture systems-level PTM changes during viral infection. Additionally, we explore how such knowledge is leveraged to identify PTM-targeted candidates for developing antiviral drugs. Looking ahead, studies focusing on the discovery and functional elucidation of PTMs, either on the host or viral proteins, will not only deepen our understanding of molecular pathology but also pave the way for developing better drugs to fight emerging viruses.

Immunology↗

Expansion of the global RNA virome reveals diverse clades of bacteriophages

High-throughput RNA sequencing offers broad opportunities to explore the Earth RNA virome. Mining 5,150 diverse metatranscriptomes uncovered >2.5 million RNA virus contigs. Analysis of >330,000 RNA-dependent RNA polymerases (RdRPs) shows that this expansion corresponds to a 5-fold increase of the known RNA virus diversity. Gene content analysis revealed multiple protein domains previously not found in RNA viruses and implicated in virus-host interactions. Extended RdRP phylogeny supports the monophyly of the five established phyla and reveals two putative additional bacteriophage phyla and numerous putative additional classes and orders. The dramatically expanded phylum Lenarviricota, consisting of bacterial and related eukaryotic viruses, now accounts for a third of the RNA virome. Identification of CRISPR spacer matches and bacteriolytic proteins suggests that subsets of picobirnaviruses and partitiviruses, previously associated with eukaryotes, infect prokaryotic hosts.

59 BASIC BIOLOGICAL SCIENCES↗

The protein structurome of Orthornavirae and its dark matter

Metatranscriptomics is uncovering more and more diverse families of viruses with RNA genomes comprising the viral kingdom Orthornavirae in the realm Riboviria. Thorough protein annotation and comparison are essential to get insights into the functions of viral proteins and virus evolution. In addition to sequence- and hmm profile-based methods, protein structure comparison adds a powerful tool to uncover protein functions and relationships. We constructed an Orthornavirae “structurome” consisting of already annotated as well as unannotated (“dark matter”) proteins and domains encoded in viral genomes. We used protein structure modeling and similarity searches to illuminate the remaining dark matter in hundreds of thousands of orthornavirus genomes. The vast majority of the dark matter domains showed either “generic” folds, such as single α-helices, or no high confidence structure predictions. Nevertheless, a variety of lineage-specific globular domains that were new either to orthornaviruses in general or to particular virus families were identified within the proteomic dark matter of orthornaviruses, including several predicted nucleic acid-binding domains and nucleases. In addition, we identified a case of exaptation of a cellular nucleoside monophosphate kinase as an RNA-binding protein in several virus families. Notwithstanding the continuing discovery of numerous orthornaviruses, it appears that all the protein domains conserved in large groups of viruses have already been identified. The rest of the viral proteome seems to be dominated by poorly structured domains including intrinsically disordered ones that likely mediate specific virus-host interactions.

59 BASIC BIOLOGICAL SCIENCES↗

Comparing variability in diagnosis of upper respiratory tract infections in patients using syndromic, next generation sequencing, and PCR-based methods

Early and accurate diagnosis of respiratory pathogens and associated outbreaks can allow for the control of spread, epidemiological modeling, targeted treatment, and decision making–as is evident with the current COVID-19 pandemic. Many respiratory infections share common symptoms, making them difficult to diagnose using only syndromic presentation. Yet, with delays in getting reference laboratory tests and limited availability and poor sensitivity of point-of-care tests, syndromic diagnosis is the most-relied upon method in clinical practice today. Here, we examine the variability in diagnostic identification of respiratory infections during the annual infection cycle in northern New Mexico, by comparing syndromic diagnostics with polymerase chain reaction (PCR) and sequencing-based methods, with the goal of assessing gaps in our current ability to identify respiratory pathogens. Of 97 individuals that presented with symptoms of respiratory infection, only 23 were positive for at least one RNA virus, as confirmed by sequencing. Whereas influenza virus (n = 7) was expected during this infection cycle, we also observed coronavirus (n = 7), respiratory syncytial virus (n = 8), parainfluenza virus (n = 4), and human metapneumovirus (n = 1) in individuals with respiratory infection symptoms. Four patients were coinfected with two viruses. In 21 individuals that tested positive using PCR, RNA sequencing completely matched in only 12 (57%) of these individuals. Few individuals (37.1%) were diagnosed to have an upper respiratory tract infection or viral syndrome by syndromic diagnostics, and the type of virus could only be distinguished in one patient. Thus, current syndromic diagnostic approaches fail to accurately identify respiratory pathogens associated with infection and are not suited to capture emerging threats in an accurate fashion. We conclude there is a critical and urgent need for layered agnostic diagnostics to track known and unknown pathogens at the point of care to control future outbreaks.

60 APPLIED LIFE SCIENCES↗

Morphological and Genomic Features of the New Klosneuvirinae Isolate Fadolivirus IHUMI-VV54

Since the discovery of Mimivirus, viruses with large genomes encoding components of the translation machinery and other cellular processes have been described as belonging to the nucleocytoplasmic large DNA viruses. Recently, genome-resolved metagenomics led to the discovery of more than 40 viruses that have been grouped together in a proposed viral subfamily named Klosneuvirinae. Members of this group had genomes of up to 2.4Mb in size and featured an expanded array of translation system genes. Yet, despite the large diversity of the Klosneuvirinae in metagenomic data, there are currently only two isolates available. Here, we report the isolation of a novel giant virus known as Fadolivirus from an Algerian sewage site and provide morphological data throughout its replication cycle in amoeba and a detailed genomic characterization. The Fadolivirus genome, which is more than 1.5Mb in size, encodes 1,452 predicted proteins and phylogenetic analyses place this viral isolate as a near relative of the metagenome assembled Klosneuvirus and Indivirus. The genome encodes for 66 tRNAs, 23 aminoacyl-tRNA synthetases and a wide range of transcription factors, surpassing Klosneuvirus and other giant viruses. The Fadolivirus genome also encodes putative vacuolar-type proton pumps with the domains D and A, potentially constituting a virus-derived system for energy generation. The successful isolation of Fadolivirus will enable future hypothesis-driven experimental studies providing deeper insights into the biology of the Klosneuvirinae.

59 BASIC BIOLOGICAL SCIENCES↗

Deletion of the EP402R Gene from the Genome of African Swine Fever Vaccine Strain ASFV-G-ΔI177L Provides the Potential Capability of Differentiating between Infected and Vaccinated Animals

The African swine fever virus (ASFV) mutant ASFV-G-ΔI177L is a safe and efficacious vaccine which induces protection against the challenge of its parental virus, the Georgia 2010 isolate. Although a genetic DIVA (differentiation between infected and vaccinated animals) assay has been developed for this vaccine, still there is not a serological DIVA test for differentiating between animals vaccinated with ASFV-G-ΔI177L and those infected with wild-type viruses. In this report, we describe the development of the ASFV-G-ΔI177L mutant having deleted the EP402R gene, which encodes for the viral protein responsible for mediating the hemadsorption of swine erythrocytes. The resulting virus, ASFV-G-ΔI177L/ΔEP402R, does not have a decreased ability to replicates in swine macrophages when compared with the parental ASFV-G-ΔI177L. Domestic pigs intramuscularly (IM) inoculated with either 10 2 or 10 6 HAD 50 of ASFV-G-ΔI177L/ΔEP402R remained clinically normal, when compared with a group of mock-vaccinated animals, indicating the absence of residual virulence. Interestingly, an infectious virus could not be detected in the blood samples of the ASFV-G-ΔI177L/ΔEP402R-inoculated animals in either group at any of the time points tested. Furthermore, while all of the mock-inoculated animals presented a quick and lethal clinical form of ASF after the intramuscular inoculation challenge with 10 2 HAD 50 of highly virulent parental field isolate Georgia 2010 (ASFV-G), all of the ASFV-G-ΔI177L/ΔEP402R-inoculated animals were protected, remaining clinically normal until the end of the observational period. Most of the ASFV-G-ΔI177L/ΔEP402R-inoculated pigs developed strong virus-specific antibody responses against viral antigens, reaching maximum levels at 28 days post inoculation. Importantly, all of the sera collected at that time point in the ASFV-G-ΔI177L/ΔEP402R-inoculated pigs did not react in a direct ELISA coated with the recombinant EP402R protein. Conversely, the EP402R protein was readily recognized by the pool of sera from the animals immunized with recombinant live attenuated vaccine candidates ASFV-G-ΔI177L, ASFV-G-ΔMGF, or ASFV-G-Δ9GL/ΔUK. Therefore, ASFV-G-ΔI177L/ΔEP402R is a novel, safe and efficacious candidate with potential to be used as an antigenically DIVA vaccine.

59 BASIC BIOLOGICAL SCIENCES↗

Expanding standards in viromics: in silico evaluation of dsDNA viral genome identification, classification, and auxiliary metabolic gene curation

Viruses influence global patterns of microbial diversity and nutrient cycles. Though viral metagenomics (viromics), specifically targeting dsDNA viruses, has been critical for revealing viral roles across diverse ecosystems, its analyses differ in many ways from those used for microbes. To date, viromics benchmarking has covered read pre-processing, assembly, relative abundance, read mapping thresholds and diversity estimation, but other steps would benefit from benchmarking and standardization. Here we use in silico-generated datasets and an extensive literature survey to evaluate and highlight how dataset composition (i.e., viromes vs bulk metagenomes) and assembly fragmentation impact (i) viral contig identification tool, (ii) virus taxonomic classification, and (iii) identification and curation of auxiliary metabolic genes (AMGs). The in silico benchmarking of five commonly used virus identification tools show that gene-content-based tools consistently performed well for long (≥3 kbp) contigs, while k -mer- and blast-based tools were uniquely able to detect viruses from short (≤3 kbp) contigs. Notably, however, the performance increase of k -mer- and blast-based tools for short contigs was obtained at the cost of increased false positives (sometimes up to ~5% for virome and ~75% bulk samples), particularly when eukaryotic or mobile genetic element sequences were included in the test datasets. Furthermore, for viral classification, variously sized genome fragments were assessed using gene-sharing network analytics to quantify drop-offs in taxonomic assignments, which revealed correct assignations ranging from ~95% (whole genomes) down to ~80% (3 kbp sized genome fragments). A similar trend was also observed for other viral classification tools such as VPF-class, ViPTree and VIRIDIC, suggesting that caution is warranted when classifying short genome fragments and not full genomes. Finally, we highlight how fragmented assemblies can lead to erroneous identification of AMGs and outline a best-practices workflow to curate candidate AMGs in viral genomes assembled from metagenomes. Together, these benchmarking experiments and annotation guidelines should aid researchers seeking to best detect, classify, and characterize the myriad viruses ‘hidden’ in diverse sequence datasets.

59 BASIC BIOLOGICAL SCIENCES↗

Evolutionary history, potential intermediate animal host, and cross-species analyses of SARS-CoV-2

To investigate the evolutionary history of the recent outbreak of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in China, a total of 70 genomes of virus strains from China and elsewhere with sampling dates between 24 December 2019 and 3 February 2020 were analyzed. To explore the potential intermediate animal host of the SARS-CoV-2 virus, we reanalyzed virome data sets from pangolins and representative SARS-related coronaviruses isolates from bats, with particular attention paid to the spike glycoprotein gene. We performed phylogenetic, split network, transmission network, likelihood-mapping, and comparative analyses of the genomes. Additionally, based on Bayesian time-scaled phylogenetic analysis using the tip-dating method, we estimated the time to the most recent common ancestor and evolutionary rate of SARS-CoV-2, which ranged from 22 to 24 November 2019 and 1.19 to 1.31 × 10 -3 substitutions per site per year, respectively. Our results also revealed that the BetaCoV/bat/Yunnan/RaTG13/2013 virus was more similar to the SARS-CoV-2 virus than the coronavirus obtained from the two pangolin samples (SRR10168377 and SRR10168378). We also identified a unique peptide (PRRA) insertion in the human SARS-CoV-2 virus, which may be involved in the proteolytic cleavage of the spike protein by cellular proteases, and thus could impact host range and transmissibility. Interestingly, the coronavirus carried by pangolins did not have the RRAR motif. Therefore, we concluded that the human SARS-CoV-2 virus, which is responsible for the recent outbreak of COVID-19, did not come directly from pangolins.

59 BASIC BIOLOGICAL SCIENCES↗

Viral Inhibitors Derived from Macroalgae, Microalgae, and Cyanobacteria: A review of antiviral potential throughout pathogenesis

Viruses are abiotic obligate parasites utilizing complex mechanisms to hijack cellular machinery and reproduce, causing multiple harmful effects in the process. Viruses represent a growing global health concern; at the time of writing, COVID-19 has killed at least two million people around the world and devastated global economies. Lingering concern regarding the virus’ prevalence yet hampers return to normalcy. While catastrophic in and of itself, COVID-19 further heralds in a new era of human-disease interaction characterized by the emergence of novel viruses from natural sources with heretofore unseen frequency. Due to deforestation, population growth, and climate change, we are encountering more viruses that can infect larger groups of people with greater ease and increasingly severe outcomes. The devastation of COVID-19 and forecasts of future human/disease interactions call for a creative reconsideration of global response to infectious disease. There is an urgent need for accessible, cost-effective antiviral (AV) drugs that can be mass-produced and widely distributed to large populations. Development of AV drugs should be informed by a thorough understanding of viral structure and function as well as human biology. To maximize efficacy, minimize cost, and reduce development of drug-resistance, these drugs would ideally operate through a varied set of mechanisms at multiple stages throughout the course of infection. Due to their abundance and diversity, natural compounds are ideal for such comprehensive therapeutic interventions. Promising sources of such drugs are found throughout nature; especially remarkable are the algae, a polyphyletic grouping of phototrophs that produce diverse bioactive compounds. While not much literature has been published on the subject, studies have shown that these compounds exert antiviral effects at different stages of viral pathogenesis. In this review, we follow the course of viral infection in the human body and evaluate the AV effects of algae-derived compounds at each stage. Specifically, we examine the AV activities of algae-derived compounds at the entry of viruses into the body, transport through the body via the lymph and blood, infection of target cells, and immune response. We discuss what is known about algae-derived compounds that may interfere with the infection pathways of SARS-CoV-2; and review which algae are promising sources for AV agents or AV precursors that, with further investigation, may yield life-saving drugs due to their diversity of mechanisms and exceptional pharmaceutical potential.

Algae, antiviral, SARS-CoV-2, seaweed, cyanobacter↗

Efficacy of an inactivated Senecavirus A vaccine in weaned pigs and mature sows

Senecavirus A (SVA), commonly known as Seneca Valley virus (SVV) is a causative agent for vesicular disease in swine. It has been found across the globe including the United States, Brazil, and China. Clinical disease caused by this virus is identical to foot-and-mouth disease virus (FMDV). Since FMDV has the potential to cause severe economic consequences in FMDV-free countries, those countries are on high alert for signs of vesicles in swine and an investigation is performed to rule out the presence of FMDV if observed. In countries where SVA cases have continued to occur, investigations and testing can cause a burden on personnel and resources. The objectives of this study were to test the efficacy of a whole-virus inactivated SVA vaccine against challenge in nursery-aged pigs, mature sows, and to assess the protection of passive maternal immunity generated by immunized dams. Animals were given two doses of the vaccine intramuscularly three weeks apart and challenged intranasally two weeks after the second dose. Non-vaccinated animals challenged with SVA developed clinical signs of disease, replicated virus, and developed a neutralizing antibody response. Vaccinated animals had robust neutralizing titers after two doses; and after challenge, did not develop vesicular disease and had limited rectal shedding. Piglets suckling immunized dams and challenged with SVA at 3–6 days-of-age had neutralizing titers prior to challenge and did not replicate or shed virus. An efficacious vaccine could improve swine welfare and reduce the economic consequences of continued foreign animal disease investigations.

60 APPLIED LIFE SCIENCES↗

Viral elements and their potential influence on microbial processes along the permanently stratified Cariaco Basin redoxcline

Little is known about viruses in oxygen-deficient water columns (ODWCs). In surface ocean waters, viruses are known to act as gene vectors among susceptible hosts. Some of these genes may have metabolic functions and are thus termed auxiliary metabolic genes (AMGs). AMGs introduced to new hosts by viruses can enhance viral replication and/or potentially affect biogeochemical cycles by modulating key microbial pathways. Here we identify 748 viral populations that cluster into 94 genera along a vertical geochemical gradient in the Cariaco Basin, a permanently stratified and euxinic ocean basin. The viral communities in this ODWC appear to be relatively novel as 80 of these viral genera contained no reference viral sequences, likely due to the isolation and unique features of this system. We identify viral elements that encode AMGs implicated in distinctive processes, such as sulfur cycling, acetate fermentation, signal transduction, [Fe–S] formation, and N-glycosylation. These AMG-encoding viruses include two putative Mu-like viruses, and viral-like regions that may constitute degraded prophages that have been modified by transposable elements. Our results provide an insight into the ecological and biogeochemical impact of viruses oxygen-depleted and euxinic habitats.

59 BASIC BIOLOGICAL SCIENCES↗

Pollen antigens and atmospheric circulation driven seasonal respiratory viral outbreak and its implication to the Covid-19 pandemic

Abstract The patterns of respiratory virus illness are expressed differently between temperate and tropical climates. Tropical outbreaks often peak in wet seasons. Temperate outbreaks typically peak during the winter. The prevailing causal hypotheses focus on sunlight, temperature and humidity variations. Yet no consistent factors have been identified to sufficiently explain seasonal virus emergence and decline at any latitude. Here we demonstrate close connections among global-scale atmospheric circulations, IgE antibody enhancement through seasonal pollen inhalation, and respiratory virus patterns at any populated latitude, with a focus on the US. Pollens emerge each Spring, and the renewed IgE titers in the population are argued to terminate each winter peak of respiratory illness. Globally circulated airborne viruses are postulated to subsequently deposit across the Southern US during lower zonal geostrophic winds each late Summer. This seasonally refreshed viral load is postulated to trigger a new influenza outbreak, once the existing IgE antibodies diminish to a critical value each Fall. Our study offers a new and consistent explanation for the seasonal diminishment of respiratory viral illnesses in temperate climates, the subdued seasonal signature in the tropics, the annually circulated virus phenotypes, and the northerly migration of influenza across the US every year. Our integrated geospatial and IgE hypothesis provides a new perspective for prediction, mitigation and prevention of the outbreak and spread of seasonal respiratory viruses including Covid-19 pandemic.

60 APPLIED LIFE SCIENCES↗

Pleiotropic Effects of Influenza H1, H3, and B Baloxavir-Resistant Substitutions on Replication, Sensitivity to Baloxavir, and Interferon Expression

Baloxavir is an anti-influenza endonuclease inhibitor that targets the polymerase acidic (PA) protein of influenza A and B viruses. Our knowledge regarding the pleiotropic effects of baloxavir resistance-associated substitutions is limited. We generated recombinant A/California/04/09 (H1N1)-, A/Hong Kong/218849/2006 (H3N2)-, and B/Victoria/504/2000-like viruses that contained PA substitutions identified in baloxavir clinical trials and surveillance that could potentially be associated with baloxavir resistance. We characterized their susceptibility to baloxavir, impact on polymerase activity, viral growth, and ability to induce interferon (IFN) and IFN-stimulated genes expression in vitro. Four PA substitutions, H1N1 I38L/T, E199D, and B G199R, significantly reduced the sensitivity of the recombinant viruses to baloxavir (14.1-fold). We confirmed our findings by using the luciferase-based ribonucleoprotein minigenome assay and by using virus yield reduction assay in Calu-3 and normal human bronchial epithelial (NHBE) cells. We observed that I38L and E199D resulted in decreased viral replication of the H1N1 wild-type virus (1.4-fold) but the H1N1 I38T and B G199R substitutions did not significantly alter replication capacity in Calu-3 cells. In addition, H1N1 variants with PA I38L/T and E199D induced significantly higher levels of IFNB1 gene expression compared to the wild-type virus (4.2-fold). In contrast, the B variant, G199R, triggered the lowest levels of IFN genes in Calu-3 cells (1.6-fold). Because baloxavir is a novel anti-influenza therapeutic agent, identifying and characterizing substitutions associated with reduced sensitivity to baloxavir, as well as the impact of these substitutions on viral fitness, is paramount to the strategic implementation of this novel countermeasure.

54 ENVIRONMENTAL SCIENCES↗

Comparative Assessment of Severe Acute Respiratory Syndrome Coronavirus 2 Variants in the Ferret Model

The continued spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in humans necessitates evaluation of variants for enhanced virulence and transmission. We used the ferret model to perform a comparative analysis of four SARS-CoV-2 strains, including an early pandemic isolate from the United States (WA1), and representatives of the Alpha, Beta, and Delta lineages. While Beta virus was not capable of pronounced replication in ferrets, WA1, Alpha, and Delta viruses productively replicated in the ferret upper respiratory tract, despite causing only mild disease with no overt histopathological changes. Strain-specific transmissibility was observed; WA1 and Delta viruses transmitted in a direct contact setting, whereas Delta virus was also capable of limited airborne transmission. Viral RNA was shed in exhaled air particles from all inoculated animals but was highest for Delta virus. Prior infection with SARS-CoV-2 offered varied protection against reinfection with either homologous or heterologous variants. Notable genomic variants in the spike protein were most frequently detected following WA1 and Delta virus infection.

59 BASIC BIOLOGICAL SCIENCES↗

Single-molecule 3D imaging of HIV cellular entry by liquid-phase electron tomography

Enveloped viruses, including human immunodeficiency virus (HIV) and SARS-CoV-2, target cells through membrane fusion process. The detailed understanding of the process is sought after for vaccine development but remains elusive due to current technique limitations for direct three-dimensional (3D) imaging of an individual virus during its viral entry. Recently, we developed a simple specimen preparation method for real-time imaging of metal dynamic liquid-vaper interface at nanometer resolution by transmission electron microscopy (TEM). Here, we extended this method to study biology sample through snapshot 3D structure of a single HIV (pseudo-typed with the envelope glycoprotein of vesicular stomatitis virus, VSV-G) at its intermediate stage of viral entry to HeLa cells in a liquid-phase environment. By individual-particle electron tomography (IPET), we found the viral surface release excess lipids with unbound viral spike proteins forming ~50-nm nanoparticles instead of merging cell membrane. Moreover, the spherical-shape shell formed by matrix proteins underneath the viral envelope does not disassemble into a cone shape right after fusion. Further, the snapshot 3D imaging of a single virus provides us a direct structure-based understanding of the viral entry mechanism, which can be used to examine other viruses to support the development of vaccines combatting the current ongoing pandemic.

Kong, Lingli↗