Biology subjects

Fuller, D.

Publications and source records attributed to Fuller, D..

4 recordsLinked to original sources

bioRxiv2023

Elevated peripheral and nervous system inflammation is associated with decreased short-chain fatty acid levels in Zika-virus infected macaques

Zika virus (ZIKV) infection of central nervous system (CNS) tissue is associated with CNS inflammation, which contributes to ZIKV pathology. Similarly, ZIKV infection has been associated with increased vaginal and rectal mucosal inflammation. As mucosal dysfunction may contribute to elevated systemic inflammation, ZIKV-induced mucosal alterations could potentiate CNS disruptions, leading to ZIKV pathogenesis. However, the potential link between mucosal dysfunction, CNS inflammation and the underlying mechanisms causing these disruptions in ZIKV infection has not been well described. Here, we assessed plasma and CSF indicators of inflammation, including neopterin, tryptophan, kynurenine and serotonin by liquid chromatography tandem mass spectrometry. We observed significant increases in neopterin formation, tryptophan catabolism and serotonin levels in the plasma and CSF of ZIKV-infected pigtail macaques (PTM), rhesus macaques (RM) and in the plasma of ZIKV-infected humans. We next examined whether ZIKV infection resulted in microbial translocation across mucosal surfaces by evaluating plasma and cerebrospinal fluid (CSF) levels of soluble CD14 (sCD14) and lipopolysaccharide-binding protein (LBP) by enzyme-linked immunosorbent assay (ELISA). Increased sCD14 was observed in the CSF of PTM and rhesus macaque (RM), while increased LBP was observed in pigtail macaque (PTM) plasma. Finally, to examine whether ZIKV-induced microbial dysbiosis could underlie increased microbial translocation and inflammation, we characterized intestinal microbial communities by 16s rRNA gene sequencing and microbial functional changes by quantifying short-chain fatty acid (SCFA) concentrations by gas chromatography mass spectrometry. We observed that although ZIKV infection of PTM did not result in significant taxonomic shifts in microbial communities, there were significant reductions in SCFA levels. Loss of microbial function in ZIKV infection could cause decreased intestinal integrity, thereby contributing to elevated microbial translocation and systemic and CNS inflammation, providing a possible mechanism underlying ZIKV pathogenesis. Further, this may represent a mechanism underlying inflammation and pathogenesis in other diseases. Author SummaryZika virus (ZIKV) can be transmitted to humans via the bite of an infected mosquito or between humans during sexual intercourse, typically resulting in mild symptoms, which has been linked to elevated inflammation in the CNS and the development of more serious conditions, including severe neurological syndromes. Previous studies have observed that ZIKV infection is associated with increased mucosal dysfunction, including elevated inflammation in rectal and vaginal mucosal tissue. However, the mechanism of ZIKV-induced mucosal dysfunction may contribute to systemic and CNS inflammation has not been previously investigated. Here, we used the non-human primate (NHP) model and clinical specimens from ZIKV-infected humans to examine markers of systemic and CNS inflammation and microbial translocation. We observed elevated markers indicative of microbial translocation and inflammation in the CNS of ZIKV-infected macaques and humans. A potential association with mucosal dysfunction in ZIKV infection is shifts in microbial dysbiosis. We also observed that there were no significant overall taxonomic shifts in microbial communities, but a reduction of bacterial-derived short-chain fatty acid (SCFA) levels. Finally, we observed that the decrease in SCFA levels significantly negatively correlated with the elevated peripheral and CNS inflammatory markers, suggesting a link between ZIKV-driven disease pathology and microbial function. Taken together, our study provides new insight into a previously unconsidered mechanism underlying ZIKV pathogenesis.

microbiology↗

bioRxiv2022

Sequencing by avidity enables high accuracy with low reagent consumption

We present avidity sequencing - a novel sequencing chemistry that separately optimizes the process of stepping along a DNA template and the process of identifying each nucleotide within the template. Nucleotide identification uses multivalent nucleotide ligands on dye-labeled cores to form polymerase-polymer nucleotide complexes bound to clonal copies of DNA targets. These polymer-nucleotide substrates, termed avidites, decrease the required concentration of reporting nucleotides from micromolar to nanomolar, and yield negligible dissociation rates. We demonstrate the use of avidites as a key component of a sequencing technology that surpasses Q40 accuracy and enables a diversity of applications that include single cell RNA-seq and whole human genome sequencing. We also show the advantages of this technology in sequencing through long homopolymers.

genomics↗

bioRxiv2022

Uman Type NF-L Antibodies Are Effective Reagents for the Imaging of Neurodegeneration

Recent work shows that certain immunological assays for the neurofilament light chain NF-L detect informative signals in the CSF and blood of human and animals affected by a variety of CNS injury and disease states. Much of this work has been performed using two mouse monoclonal antibodies to NF-L, UD1 and UD2, also known as 2.1 and 47.3 respectively. These are the essential components of the Uman Diagnostics NF-Light ELISA kit, the Quanterix Simoa bead based NF-L assay and others. We show here that the antibodies bind to neighboring epitopes in a short, conserved and unusual peptide in the NF-L "rod" Coil 2 region. We also describe a surprising and useful feature of Uman and similar reagents. While other well characterized NF-L antibodies show robust staining of countless cells and processes in CNS sections from healthy rats, both Uman antibodies reveal only a minor subset of presumably spontaneously degenerating or degenerated neurons and their processes. However following experimental mid-cervical injuries to rat spinal cord both Uman antibodies recognize numerous profiles in tissue sections. The Uman positive material was associated with fiber tracts expected to be damaged by the injury administered and the profiles had the swollen, beaded, discontinuous and sinusoidal morphology expected for degenerating and degenerated processes. We also found that several antibodies to the C terminal "tail" region of NF-L stain undamaged axonal profiles but fail to recognize the Uman positive material. The unmasking of the Uman epitopes and the loss of the NF-L tail epitopes can be mimicked by treating sections from healthy animals with proteases suggesting that the immunological changes we have discovered are due to neurodegeneration induced proteolysis. We have also generated a novel panel of monoclonal and polyclonal antibody reagents directed against the region of NF-L including the Uman epitopes which have staining properties identical to the Uman reagents. Using these we show that the NF-L region to which the Uman reagents bind contains further hidden epitopes distinct from those recognized by the two Uman reagents. We speculate that the Uman type epitopes are part of a binding region important for higher order neurofilament assembly. The work provides important insights into the properties of the NF-L biomarker, describes novel and useful properties of Uman type and NF-L tail binding antibodies and provides a hypothesis relevant to further understanding of neurofilament assembly.

neuroscience↗

bioRxiv2022

Discovery of Potent Pyrazoline-Based Covalent SARS-CoV-2 Main Protease Inhibitors

While vaccines and antivirals are now being deployed for the current SARS-CoV-2 pandemic, we require additional antiviral therapeutics to not only effectively combat SARS-CoV-2 and its variants, but also future coronaviruses. All coronaviruses have relatively similar genomes that provide a potential exploitable opening to develop antiviral therapies that will be effective against all coronaviruses. Among the various genes and proteins encoded by all coronaviruses, one particularly "druggable" or relatively easy-to-drug target is the coronavirus Main Protease (3CLpro or Mpro), an enzyme that is involved in cleaving a long peptide translated by the viral genome into its individual protein components that are then assembled into the virus to enable viral replication in the cell. Inhibiting Mpro with a small-molecule antiviral would effectively stop the ability of the virus to replicate, providing therapeutic benefit. In this study, we have utilized activity-based protein profiling (ABPP)-based chemoproteomic approaches to discover and further optimize cysteine-reactive pyrazoline-based covalent inhibitors for the SARS-CoV-2 Mpro. Structure-guided medicinal chemistry and modular synthesis of di- and tri-substituted pyrazolines bearing either chloroacetamide or vinyl sulfonamide cysteine-reactive warheads enabled the expedient exploration of structure-activity relationships (SAR), yielding nanomolar potency inhibitors against Mpro from not only SARS-CoV-2, but across many other coronaviruses. Our studies highlight promising chemical scaffolds that may contribute to future pan-coronavirus inhibitors.

biochemistry↗