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At least 649 records · Page 36

Targeted metabolomic analysis identifies increased serum levels of GABA and branched chain amino acids in canine diabetes

Introduction Dogs with naturally occurring diabetes mellitus represent a potential model for human type 1 diabetes, yet signifcant knowledge voids exist in terms of the pathogenic mechanisms underlying the canine disorder. Untargeted metabolomic studies from a limited number of diabetic dogs identifed similarities to humans with the disease. Objective To expand and validate earlier metabolomic studies, identify metabolites that difer consistently between diabetic and healthy dogs, and address whether certain metabolites might serve as disease biomarkers. Methods Untargeted metabolomic analysis via liquid chromatography-mass spectrometry was performed on serum from diabetic (n=15) and control (n=15) dogs. Results were combined with those of our previously published studies using identical methods (12 diabetic and 12 control dogs) to identify metabolites consistently diferent between the groups in all 54 dogs. Thirty-two candidate biomarkers were quantifed using targeted metabolomics. Biomarker concentrations were compared between the groups using multiple linear regression (corrected P<0.0051 considered signifcant). Results Untargeted metabolomics identifed multiple persistent diferences in serum metabolites in diabetic dogs compared with previous studies. Therefore, targeted metabolomics showed increases in gamma amino butyric acid, valine, leucine, isoleucine, citramalate, and 2-hydroxyisobutyric acid in diabetic versus control dogs while indoxyl sulfate, N-acetyl-L-aspartic acid, kynurenine, anthranilic acid, tyrosine, glutamine, and tauroursodeoxycholic acid were decreased. Conclusion Several of these fndings parallel metabolomic studies in both human diabetes and other animal models of this disease. Given recent studies on the role of GABA and branched chain amino acids in human diabetes, the increase in serum concentrations in canine diabetes warrants further study of these metabolites as potential biomarkers, and to identify similarity in mechanisms underlying this disease in humans and dogs.

59 BASIC BIOLOGICAL SCIENCES↗

A review on bacteria-derived antioxidant metabolites: their production, purification, characterization, potential applications, and limitations

Abstract Antioxidants are organic molecules that scavenge reactive oxygen species (ROS) and reactive nitrogen species (RNS), thereby maintaining cellular redox balance in living organisms. The human body synthesizes endogenous antioxidants, whereas humans obtain exogenous antioxidants from other organisms such as plants, animals, fungi, and bacteria. This review primarily focuses on the antioxidant potential of natural metabolites and extracts from five major bacterial phyla, including the well-studiedActinobacteriaandCyanobacteria, as well as less-studiedBacteroides,Firmicutes, andProteobacteria.The literature survey revealed that the metabolites and the extracts with antioxidant activity can be obtained from bacterial cells and their culture supernatants. The metabolites with antioxidant activity include pigments, phycobiliproteins, polysaccharides, mycosporins-like amino acids, peptides, phenolic compounds, and alkaloids. Both metabolites and extracts demonstrate in vitro antioxidant capacity through radical-scavenging, metal-reducing, and metal-chelating activity assays. In in vivo models, they can scavenge ROS and RNS directly and/or indirectly eliminate them by enhancing the activities of antioxidant enzymes, such as catalase, superoxide dismutase, and glutathione peroxidase. Due to their antioxidant activities, they may find applications in the cosmetic industry as anti-aging agents for the skin and in medicine as drugs or supplements for combating oxidative stress-related disorders, such as neurodegenerative diseases and diabetes. The literature survey also elucidated that some metabolites and extracts with antioxidant activity also exhibited strong antimicrobial properties. Therefore, we consider that they may have future applications in the treatment of infectious diseases, the preparation of pathogen-free healthy foods, and the extension of food shelf life.

Pharmacology & Pharmacy↗

Reused cultivation water from a self-inhibiting alga does not inhibit other algae but alters their microbiomes

Economical production of algal commodities (food, feed, and fuels) requires reusing cultivation water to reduce operating costs. While some algae strains show growth inhibition in reused water, other strains appear un-affected. Reusing water to grow different strains (i.e., crop rotation) could potentially improve overall biomass production compared to water reuse with the same strain by reducing accumulation of strain-specific waste products, limiting carryover of population-specific pathogens such as viruses, and introducing bacteria that degrade residual organic matter. Here, batch culture experiments tested the effects of reused cultivation water from the self-inhibiting diatom Staurosira sp. C323 on the diatom Navicula sp. SFP and green alga Chlorella sp. D046, as well as their microbiomes. Navicula sp. and Chlorella sp. grew well in Staurosira sp. reused water, indicating that previously observed self-inhibition was likely specific to this Staurosira strain. While algal mi-crobiomes were not significantly different between controls and reused water treatments, specific bacteria taxa were differentially abundant in reused treatments, suggesting that some taxa responded to compounds re-maining in the reused water. Finally, results suggest that algae growth responses in reused water may depend largely on strain-specific factors, and that crop rotation can support biomass production.

09 BIOMASS FUELS↗

Grazer-induced changes in molecular signatures of cyanobacteria

Algal biomass production is an emerging renewable source of fuels, nutrients, manufacturing materials, and pharmaceuticals. Industrial-scale production is predominantly performed in open raceway ponds that are inexpensive to build, operate, and maintain compared to closed bioreactors. However, these open pond systems suffer from increased opportunities for biological contamination from predators, pathogens, and competitors, which result in reduced biomass quality and yields and often the complete destruction of the crop over a short period of time. Early detection of contaminants is a necessary step of integrated pest management for triggering and informing interventions to prevent crop losses. To develop a sensitive method of detection utilizing mass spectrometry (MS), we used three methods – imaging mass spectrometry (imaging MS), liquid chromatography MS/MS (LC-MS/MS) combined with molecular networking, and gas chromatography MS/MS (GC–MS/MS) – to observe and identify molecular signatures from a model predator-prey system of the heterolobosean amoeba HGG1 preying on the filamentous cyanobacterium Anabaena sp. PCC 7120. Imaging MS enabled the association of molecules with the crop, the predator, or the activity of protozoan grazing, while LC-MS/MS-based molecular networking identified a subset of the grazing-specific signals as the chlorophyll breakdown products pyropheophytin, pheophorbide A, and pyropheophorbide. Application of MS techniques to other amoebacyanobacterial predator-prey pairs allowed categorization of grazing signatures as universal or specific to the species of prey, predator, or the prey-predator pairing, indicating that MS-based techniques can distinguish crops from competitors and potentially identify predators. Finally, GCMS/MS was shown to be capable of monitoring novel volatile organic compounds (VOCs), including those predicted to be released through chlorophyll breakdown, in the headspace over algal cultures under predation. Furthermore, these results demonstrate that the combination of multiple MS technologies creates a predictive framework to identify and catalog relevant molecular signatures for informing crop protection strategies.

09 BIOMASS FUELS↗

Tissue replication and mucosal swab detection of Sosuga virus in Syrian hamsters in the absence of overt tissue pathology and clinical disease

Human infection with Sosuga virus (SOSV), a recently discovered pathogenic paramyxovirus, has been reported in one individual to date. No animal models of disease are currently available for SOSV. Here, we describe initial characterization of experimental infection in Syrian hamsters, including kinetics of virus dissemination and replication, and the corresponding clinical parameters, immunological responses, and histopathology. Here, we demonstrate susceptibility of hamsters to infection in the absence of clinical signs or significant histopathologic findings in tissues.

60 APPLIED LIFE SCIENCES↗

A review of experimental methods to determine bioaerosol transfer in energy recovery ventilators

Increasing ventilation is an effective method to reduce indoor airborne disease transmission. An Energy recovery ventilator (ERV) is a passive energy recovery device used to reduce the energy consumption of heating, ventilation and air-conditioning (HVAC) systems for conditioning the ventilation air. It preconditions the ventilation air by transferring energy from building exhaust air. Therefore, the ventilation and exhaust air streams interact directly/indirectly in the ERV for energy transfer. It is surmised that the ERV may transfer bioaerosols (with pathogens) from the exhaust air to ventilation air, resulting in the spread of infectious diseases. Consequently, many pandemic HVAC guidelines recommend that the use of ERVs be limited. This is a highly unsustainable direction given the increased energy requirements associated with the high ventilation provision advocated for pandemic operation. It must be noted that no validated experimental evidence exists in literature for bioaerosol transfer in ERVs. Hence, it is necessary to conduct extensive bioaerosol transfer research before adopting the unsustainable practice of limiting the utilization of ERVs. The main objective of this review study is to summarize the experimental methods and instrumentation for bioaerosol transfer research in ERVs. This comprehensive article provides a detailed overview of the generation, sampling, and analysis of bioaerosols for conducting the experiments. Further, it explains the possible mechanisms for bioaerosol transfer in various types of ERVs based on which the ERVs that need immediate attention are identified. The main contribution of this research paper is that it provides a novel experimental method which encompasses the biosafety aspects, instrumentation, performance parameters and uncertainties in conducting virus contained bioaerosol transfer study in ERVs. The findings from this review will be helpful in designing bioaerosol transfer experiments and developing future ERV test standards for such experiments.

42 ENGINEERING↗

Evaluation of artificial neural network performance for classification of potato plants infected with potato virus Y using spectral data on multiple varieties and genotypes

Potato virus Y (Potyviridae, PVY) is a plant virus that poses a significant threat to potato producers on a global basis. The pathogen has disrupted seed potato supplies and negatively impacted yield and quality of commercial potato crops. The potato industry currently manages PVY infection levels via insecticide applications, regional seed certification programs that rely on field scouting to visually assess individual plants for infection status, and destructive and costly tissue sampling coupled with laboratory assays. Despite these efforts, PVY continues to confound potato industry stakeholders resulting in economic harm. Remote sensing and machine learning provide for the development of new tools to more accurately detect and spatially quantify PVY-infected plants versus the current state of the art. However, there is a need to understand how the occurrence of many different potato varieties impact the dynamics of developing models to detect potato plants impacted with PVY and their potential effectiveness. This study evaluates classification modelling outcomes using spectral datasets collected in different temporal and spatial environments (greenhouse and a production field) on multiple potato varieties consisting of labelled instances of plants infected with PVY and those not infected with the virus. A modelling framework was developed to support iterative modelling runs using artificial neural network (ANN) architectures configured as binary classifiers to develop sample populations to support statistical analysis on model performance using specific spectral subsets. When using spectral data to detect PVY-infected plants, ANN models achieved the highest mean accuracy of 0.894 on a single variety. Conversely, the same ANN model architecture only achieved a mean accuracy of 0.575 on a spectral data set representing 29 potato breeding lines. Additionally, statistical analysis indicates spectral regions including the red edge, near infrared and shortwave infrared contain more important spectral features for the ANN classifier introduced in this research.

60 APPLIED LIFE SCIENCES↗

Discovery of multidrug efflux pump inhibitors with a novel chemical scaffold

Multidrug efflux is a major contributor to antibiotic resistance in Gram-negative bacterial pathogens. Inhibition of multidrug efflux pumps is a promising approach for reviving the efficacy of existing antibiotics. Previously, inhibitors targeting both the efflux transporter AcrB and the membrane fusion protein AcrA in the Escherichia coli AcrAB-TolC efflux pump were identified. In this work we use existing physicochemical property guidelines to generate a filtered library of compounds for computational docking. We then experimentally test the top candidate coumpounds using in vitro binding assays and in vivo potentiation assays in bacterial strains with controllable permeability barriers. We thus identify a new class of inhibitors of E. coli AcrAB-TolC. Six molecules with a shared scaffold were found to potentiate the antimicrobial activity of erythromycin and novobiocin in hyperporinated E. coli cells. Importantly, these six molecules were also active in wild-type strains of both Acinetobacter baumannii and Klebsiella pneumoniae, potentiating the activity of erythromycin and novobiocin up to 8-fold.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗

A Trichomonas vaginalis C2-XYPPX-repeat protein with a structured C2 domain displaying dampened flexibility upon binding calcium

C2 domains are ubiquitous membrane-binding modules of ∼130 residues in eukaryotes that are often associated with proteins involved in membrane trafficking and lipid modification. The genome of Trichomonas vaginalis, the most common, non-viral, sexually transmitted human pathogen, encodes eight genes that contain a N-terminal C2 module linked to a XYPPX-repeat domain of more than four XYPPX repeats (C2-XYPPX). While the function of the XYPPX-repeat domain remains unknown, its multiple association with C2 domains in T. vaginalis suggests it is important. Here, the C2 domain from one of these C2-XYPPX-repeat proteins, Tv-C2-1, was structurally and physically characterized using X-ray crystallography and NMR spectroscopy. The crystal structure for Tv-C2-1 shows that this domain shares a fold common to all C2 domains, a compact Greek-key motif composed of eight anti-parallel β-strands in the type-2 topology. An NMR chemical shift perturbation study with Ca 2+ showed that Tv-C2-1 bound two Ca 2+ atoms primarily via two loops (loop-1 and loop-3) on the predicted calcium binding face of the protein with K d s of 58.0 ± 0.1 μM and 232 ± 6 μM. Estimations of the overall rotational correlation time, τ c , in the apo (11.1 ns) and Ca 2+ -bound (9.2 ns) state suggests the protein becomes more compact upon Ca 2+ binding, consistent with a decrease in dynamics in loop-3 and marginally in loop-1 suggested by amide 15 N heteronuclear steady-state { 1 H}- 15 N NOEs. Showing Tv-C2-1 binds calcium and adopts a compact Greek-key motif structure, two primary features of C2 domains, suggests understanding the function of the XYPPX-repeat domain may be warranted.

NMR spectroscopy↗

Tolerance to allelopathic inhibition by free fatty acids in five biofuel candidate microalgae strains

Contaminating organisms (grazers, pathogens, competitors) and self-inhibition by algae-produced allelopathic chemicals are two issues that may limit the productivity of algal cultivation for bioproducts. One potential solution is to identify algal strains that are not affected by allelopathic inhibition even while undesirable organisms are suppressed. Here we used two experiments to test how sensitivity to allelopathy varies across algae. In the first experiment, we tested the sensitivity of five biofuel candidate green algae strains to two allelopathic compounds (i.e., free fatty acids) and found that the degree of inhibition depends strongly on both the species and specific compound. In the second experiment, we exposed one alga (Chlorella) to the sterile-filtered medium of each species, and found that the concentration of free fatty acids released into the media predicted Chlorella’s growth response. This provides a better understanding of how the production of, and sensitivity to, allelopathic compounds determines algal productivity.

09 BIOMASS FUELS↗

Effects of pH on an IDP conformational ensemble explored by molecular dynamics simulation

The conformational ensemble of intrinsically disordered proteins, such as α-synuclein, are responsible for their function and malfunction. Misfolding of α-synuclein can lead to neurodegenerative diseases, and the ability to study their conformations and those of other intrinsically disordered proteins under varying physiological conditions can be crucial to understanding and preventing pathologies. In contrast to well-folded peptides, a consensus feature of IDPs is their low hydropathy and high charge, which makes their conformations sensitive to pH perturbation. We examine a prominent member of this subset of IDPs, α-synuclein, using a divide-and-conquer scheme that provides enhanced sampling of IDP structural ensembles. We constructed conformational ensembles of α-synuclein under neutral (pH ~ 7) and low (pH ~ 3) pH conditions and compared our results with available information obtained from smFRET, SAXS, and NMR studies. Specifically, α-synuclein has been found to in a more compact state at low pH conditions and the structural changes observed are consistent with those from experiments. We also characterize the conformational and dynamic differences between these ensembles and discussed the implication on promoting pathogenic fibril formation. We find that under low pH conditions, neutralization of negatively charged residues leads to compaction of the C-terminal portion of α-synuclein while internal reorganization allows α-synuclein to maintain its overall end-to-end distance. Here, we also observe different levels of intra-protein interaction between three regions of α-synuclein at varying pH and a shift towards more hydrophilic interactions with decreasing pH.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗

Investigation of potential aerosol transmission and infectivity of SARS-CoV-2 through central ventilation systems

Here we evaluate the concentrations and probabilities of infection for both building interior and exterior exposure sources using a well-mixed model in a connected multizone building. As a central hub of many community and economic activities, buildings provide social connectivity, but the COVID-19 pandemic has reduced social connectivity due to concerns of viral spread within buildings. Although single zone models of infectious spread are well studied, the impact of aerosolized spread of SARS-CoV-2 via air handling systems in multizone buildings remains unexplored. Here we evaluate the influence of filtration, air exchange rates, and the fraction of outdoor air on the probability of infection using the well-known well-mixed modeling approach for a multizone. We find filtration lowers the concentration and probability of infection in both source and connected rooms provided at least some air is recirculated, but that probability is not zero. Filtration has no influence without recirculation or unless the outdoor air contains virus. We find that increasing the air exchange rate removes virus from the source room faster but also increases the rate of exposure to connected rooms. Therefore, slower air exchange rates reduce infectivity in connected rooms at shorter durations, but higher air exchange rates reduce infectivity at longer durations. We further find that when outdoor air is virus free, increasing the fraction of outdoor air is helpful, but, when outdoor air is infective, pathogen exposure inside can persist for hours after a short-term release.

60 APPLIED LIFE SCIENCES↗

Intricate Genetic Programs Controlling Dormancy in Mycobacterium tuberculosis

Mycobacterium tuberculosis (MTB) displays the remarkable ability to transition in and out of dormancy, a hallmark of the pathogen’s capacity to evade the immune system and exploit susceptible individuals. Uncovering the gene regulatory programs that underlie the phenotypic shifts in MTB during disease latency and reactivation has posed a challenge. We develop an experimental system to precisely control dissolved oxygen levels in MTB cultures in order to capture the transcriptional events that unfold as MTB transitions into and out of hypoxia-induced dormancy. Using a comprehensive genome-wide transcription factor binding map and insights from network topology analysis, we identify regulatory circuits that deterministically drive sequential transitions across six transcriptionally and functionally distinct states encompassing more than three-fifths of the MTB genome. The architecture of the genetic programs explains the transcriptional dynamics underlying synchronous entry of cells into a dormant state that is primed to infect the host upon encountering favorable conditions.

59 BASIC BIOLOGICAL SCIENCES↗

Formin nanoclustering-mediated actin assembly during plant flagellin and DSF signaling

Plants respond to bacterial infection acutely with actin remodeling during plant immune responses. The mechanisms by which bacterial virulence factors (VFs) modulate plant actin polymerization remain enigmatic. Here, we show that plant-type-I formin serves as the molecular sensor for actin remodeling in response to two bacterial VFs: Xanthomonas campestris pv. campestris (Xcc) diffusible signal factor (DSF), and pathogen-associated molecular pattern (PAMP) flagellin in pattern-triggered immunity (PTI). Both VFs regulate actin assembly by tuning the clustering and nucleation activity of formin on the plasma membrane (PM) at the nano-sized scale. By being integrated within the cell-wall-PM-actin cytoskeleton (CW-PM-AC) continuum, the dynamic behavior and function of formins are highly dependent on each scaffold layer’s composition within the CW-PM-AC continuum during both DSF and PTI signaling. Our results reveal a central mechanism for rapid actin remodeling during plant-bacteria interactions, in which bacterial signaling molecules fine tune plant formin nanoclustering in a host mechanical-structure-dependent manner.

59 BASIC BIOLOGICAL SCIENCES↗

Zika-specific neutralizing antibodies targeting inter-dimer envelope epitopes

Zika virus (ZIKV) is an emerging pathogen that causes devastating congenital defects. The overlapping epidemiology and immunologic cross-reactivity between ZIKV and dengue virus (DENV) pose complex challenges to vaccine design, given the potential for antibody-dependent enhancement of disease. Therefore, classification of ZIKV-specific antibody targets is of notable value. From a ZIKV-infected rhesus macaque, we identify ZIKV-reactive B cells and isolate potent neutralizing monoclonal antibodies (mAbs) with no cross-reactivity to DENV. We group these mAbs into four distinct antigenic groups targeting ZIKV-specific cross-protomer epitopes on the envelope glycoprotein. Co-crystal structures of representative mAbs in complex with ZIKV envelope glycoprotein reveal envelope-dimer epitope and unique dimer-dimer epitope targeting. All four specificities are serologically identified in convalescent humans following ZIKV infection, and representative mAbs from all four groups protect against ZIKV replication in mice. These results provide key insights into ZIKV-specific antigenicity and have implications for ZIKV vaccine, diagnostic, and therapeutic development.

59 BASIC BIOLOGICAL SCIENCES↗

Rapid design of bacteriophage cocktails to suppress the burden and virulence of gut-resident carbapenem-resistant Klebsiella pneumoniae

Antibiotic use can lead to the expansion of multi-drug-resistant pathobionts within the gut microbiome that can cause life-threatening infections. Selective alternatives to conventional antibiotics are in dire need. Here, in this work, we describe a Klebsiella PhageBank for the tailored design of bacteriophage cocktails to treat multi-drug-resistant Klebsiella pneumoniae. Using a transposon library in carbapenem-resistant K. pneumoniae, we identify host factors required for phage infection in major Klebsiella phage families. Leveraging the diversity of the PhageBank, we formulate phage combinations that eliminate K. pneumoniae with minimal phage resistance. Optimized cocktails selectively suppress the burden of K. pneumoniae in the mouse gut and drive the loss of key virulence factors that act as phage receptors. Phage-mediated diversification of bacterial populations in the gut leads to co-evolution of phage variants with higher virulence and broader host range. Altogether, the Klebsiella PhageBank charts a roadmap for phage therapy against a critical multidrug-resistant human pathogen.

60 APPLIED LIFE SCIENCES↗

A k-mer based approach for classifying viruses without taxonomy identifies viral associations in human autism and plant microbiomes

Viruses are an underrepresented taxa in the study and identification of microbiome constituents; however, they play an essential role in health, microbiome regulation, and transfer of genetic material. Only a few thousand viruses have been isolated, sequenced, and assigned a taxonomy, which limits the ability to identify and quantify viruses in the microbiome. Additionally, the vast diversity of viruses represents a challenge for classification, not only in constructing a viral taxonomy, but also in identifying similarities between a virus’ genotype and its phenotype. However, the diversity of viral sequences can be leveraged to classify their sequences in metagenomic and metatranscriptomic samples, even if they do not have a taxonomy. To identify and quantify viruses in transcriptomic and genomic samples, we developed a dynamic programming algorithm for creating a classification tree out of 715,672 metagenome viruses. To create the classification tree, we clustered proportional similarity scores generated from the k-mer profiles of each of the metagenome viruses to create a database of metagenomic viruses. The resulting Kraken2 database of the metagenomic viruses can be found here: https://www.osti.gov/biblio/1615774 and is compatible with Kraken2. We then integrated the viral classification database with databases created with genomes from NCBI for use with ParaKraken (a parallelized version of Kraken provided in Supplemental Zip 1), a metagenomic/transcriptomic classifier. To illustrate the breadth of our utility for classifying metagenome viruses, we analyzed data from a plant metagenome study identifying genotypic and compartment specific differences between two Populus genotypes in three different compartments. We also identified a significant increase in abundance of eight viral sequences in post mortem brains in a human metatranscriptome study comparing Autism Spectrum Disorder patients and controls. We also show the potential accuracy for classifying viruses by utilizing both the JGI and NCBI viral databases to identify the uniqueness of viral sequences. Finally, we validate the accuracy of viral classification with NCBI databases containing viruses with taxonomy to identify pathogenic viruses in known COVID-19 and cassava brown streak virus infection samples. Our method represents the compulsory first step in better understanding the role of viruses in the microbiome by allowing for a more complete identification of sequences without taxonomy. Better classification of viruses will improve identifying associations between viruses and their hosts as well as viruses and other microbiome members. Despite the lack of taxonomy, this database of metagenomic viruses can be used with any tool that utilizes a taxonomy, such as Kraken, for accurate classification of viruses.

59 BASIC BIOLOGICAL SCIENCES↗

Fungal endophytes

Organisms are commonly grouped into ecological guilds that reflect their shared resource use and similar ecological roles. The guild concept has been used to categorize the vast diversity of the fungal kingdom (estimated at 2.2–5 million species) into groups of fungi with common lifestyles, such as mycorrhizal symbionts, pathogens of animals and plants, and the group that is the focus of this primer — fungal endophytes of plants. Fungal endophytes have resisted scientists’ attempts to silo organisms into neat assemblages. In conclusion, scattered across the fungal tree of life and lacking few diagnostic characteristics, they are defined less by what they are than by what they are not.

Cosner, Julian [Oak Ridge National Laboratory (ORN↗