Forest reorganisation effects on fuel moisture content can exceed changes due to climate warming in wet temperate forests.

The distributions of vegetation and fire activity are changing rapidly in response to climate warming. In many regions, climate effects on dead fuel moisture content (FMC) are expected to increase future wildfire activity. However, forest FMC is largely driven by microclimate conditions, which are moderated from open weather by vegetation canopies. As shifts in vegetation increase under climate warming, the extent to which future fire activity will be driven by climate directly or associated vegetation shifts remains unresolved. Here, we present a study aimed at quantifying the relative magnitudes of (i) direct climate warming, and (ii) vegetation change, on FMC. Field sites to evaluate these effects were established in a natural laboratory of altered forest states to mature wet temperate forest in south-eastern Australia. FMC was estimated using a process-based model and 48 years of reconstructed climate data. Canopy effects on microclimate were captured by transferring inputs from climate to microclimate using models parameterised with field observations. To evaluate the relative magnitude of climate and vegetation effects, we calculated the maximum difference in mean annual FMC across annual climate replicates and compared this to FMC differences across reorganising forest sites. Our results show vegetation effects on FMC can exceed those related to expected climate change. Changes to forest structure and composition increased (+15.7%) and decreased (-12.3%) mean annual FMC, with a larger negative effect when forest cover was completely removed (-18.5%). In contrast, the largest climate effect on FMC was -6.6% across 48-years of data. Our study demonstrates that the magnitude of vegetation effects on FMC can exceed expected climate change effects. Models of future fire activity that do not account for changing vegetation effects on microclimate are omitting a key biophysical control on FMC and therefore may not be accurately predicting future fire activity.

Exploring antiviral and anti-inflammatory effects of thiol drugs in COVID-19.

The redox status of the cysteine-rich SARS-CoV-2 spike glycoprotein (SARS-2-S) is important for the binding of SARS-2-S to angiotensin-converting enzyme 2 (ACE2), suggesting that drugs with a functional thiol group ("thiol drugs") may cleave cystines to disrupt SARS-CoV-2 cell entry. In addition, neutrophil-induced oxidative stress is a mechanism of COVID-19 lung injury, and the antioxidant and anti-inflammatory properties of thiol drugs, especially cysteamine, may limit this injury. To first explore the antiviral effects of thiol drugs in COVID-19, we used an ACE-2 binding assay and cell entry assays utilizing reporter pseudoviruses and authentic SARS-CoV-2 viruses. We found that multiple thiol drugs inhibit SARS-2-S binding to ACE2 and virus infection. The most potent drugs were effective in the low millimolar range, and IC50 values followed the order of their cystine cleavage rates and lower thiol pKa values. To determine if thiol drugs have antiviral effects in vivo and to explore any anti-inflammatory effects of thiol drugs in COVID-19, we tested the effects of cysteamine delivered intraperitoneally to hamsters infected with SARS-CoV-2. Cysteamine did not decrease lung viral infection, but it significantly decreased lung neutrophilic inflammation and alveolar hemorrhage. We speculate that the concentration of cysteamine achieved in the lungs with intraperitoneal delivery was insufficient for antiviral effects but sufficient for anti-inflammatory effects. We conclude that thiol drugs decrease SARS-CoV-2 lung inflammation and injury, and we provide rationale for future studies to test if direct (aerosol) delivery of thiol drugs to the airways might also result in antiviral effects.

Thiol drugs decrease SARS-CoV-2 lung injury in vivo and disrupt SARS-CoV-2 spike complex binding to ACE2 in vitro.

Neutrophil-induced oxidative stress is a mechanism of lung injury in COVID-19, and drugs with a functional thiol group ("thiol drugs"), especially cysteamine, have anti-oxidant and anti-inflammatory properties that could limit this injury. Thiol drugs may also alter the redox status of the cysteine-rich SARS-CoV-2 spike glycoprotein (SARS-2-S) and thereby disrupt ACE2 binding. Using ACE2 binding assay, reporter virus pseudotyped with SARS-CoV-2 spikes (ancestral and variants) and authentic SARS-CoV-2 (Wuhan-1), we find that multiple thiol drugs inhibit SARS-2-S binding to ACE2 and virus entry into cells. Pseudoviruses carrying variant spikes were less efficiently inhibited as compared to pseudotypes bearing an ancestral spike, but the most potent drugs still inhibited the Delta variant in the low millimolar range. IC50 values followed the order of their cystine cleavage rates and lower thiol pKa values. In hamsters infected with SARS-CoV-2, intraperitoneal (IP) cysteamine decreased neutrophilic inflammation and alveolar hemorrhage in the lungs but did not decrease viral infection, most likely because IP delivery could not achieve millimolar concentrations in the airways. These data show that thiol drugs inhibit SARS-CoV-2 infection in vitro and reduce SARS-CoV-2-related lung injury in vivo and provide strong rationale for trials of systemically delivered thiol drugs as COVID-19 treatments. We propose that antiviral effects of thiol drugs in vivo will require delivery directly to the airways to ensure millimolar drug concentrations and that thiol drugs with lower thiol pKa values are most likely to be effective.

Plant traits linked to field-scale flammability metrics in prescribed burns in Eucalyptus forest.

Vegetation is a key determinant of wildfire behaviour at field scales as it functions as fuel. Past studies in the laboratory show that plant flammability, the ability of plants to ignite and maintain combustion, is a function of their traits. However, the way the traits of individual plants combine in a vegetation community to affect field flammability has received little attention. This study aims to bridge the gap between the laboratory and field by linking plant traits to metrics of field-scale flammability. Across three prescribed burns, in Eucalyptus-dominated damp and dry forest, we measured pre-burn plant species abundance and post-burn field flammability metrics (percentage area burnt, char and scorch height). For understory species with dominant cover-abundance, we measured nine traits that had been demonstrated to influence flammability in the laboratory. We used fourth-corner ordination to evaluate covariation between the plant traits, species abundance and flammability. We found that several traits covaried at the species level. In some instances, these traits (e.g. specific leaf area and bulk density) could have cumulative effects on the flammability of a species while in other instances (e.g. moisture and specific leaf area) they may have counteractive effects, assuming trait effects on flammability are akin to previous research. At field scales, species with similar traits tended to co-occur, suggesting that the effects of individual traits accumulate within a plant community. Fourth-corner analyses found the trait-field flammability relationship to be statistically significant. Traits significantly associated with increasing field flammability metrics were: bulk density (negatively associated) and hydrocarbon quantity, specific leaf area and surface area to volume ratio (all positively associated). Our study demonstrates that some traits known to influence flammability in the laboratory can be associated with field-scale flammability metrics. Further research is needed to isolate the contributions of individual traits to understand how species composition drives forest flammability.

Modulation of Protein Fouling and Interfacial Properties at Carbon Surfaces via Immobilization of Glycans Using Aryldiazonium Chemistry.

Carbon materials and nanomaterials are of great interest for biological applications such as implantable devices and nanoparticle vectors, however, to realize their potential it is critical to control formation and composition of the protein corona in biological media. In this work, protein adsorption studies were carried out at carbon surfaces functionalized with aryldiazonium layers bearing mono- and di-saccharide glycosides. Surface IR reflectance absorption spectroscopy and quartz crystal microbalance were used to study adsorption of albumin, lysozyme and fibrinogen. Protein adsorption was found to decrease by 30-90% with respect to bare carbon surfaces; notably, enhanced rejection was observed in the case of the tested di-saccharide vs. simple mono-saccharides for near-physiological protein concentration values. ζ-potential measurements revealed that aryldiazonium chemistry results in the immobilization of phenylglycosides without a change in surface charge density, which is known to be important for protein adsorption. Multisolvent contact angle measurements were used to calculate surface free energy and acid-base polar components of bare and modified surfaces based on the van Oss-Chaudhury-Good model: results indicate that protein resistance in these phenylglycoside layers correlates positively with wetting behavior and Lewis basicity.

Enhanced Antifouling Properties of Carbohydrate Coated Poly(ether sulfone) Membranes.

Poly(ether sulfone) membranes (PES) were modified with biologically active monosaccharides and disaccharides using aryldiazonium chemistry as a mild, one-step, surface-modification strategy. We previously proposed the modification of carbon, metals, and alloys with monosaccharides using the same method; herein, we demonstrate modification of PES membranes and the effect of chemisorbed carbohydrate layers on their resistance to biofouling. Glycosylated PES surfaces were characterized using spectroscopic methods and tested against their ability to interact with specific carbohydrate-binding proteins. Galactose-, mannose-, and lactose-modified PES surfaces were exposed to Bovine Serum Albumin (BSA) solutions to assess unspecific protein adsorption in the laboratory and were found to adsorb significantly lower amounts of BSA compared to bare membranes. The ability of molecular carbohydrate layers to impart antifouling properties was further tested in the field via long-term immersive tests at a wastewater treatment plant. A combination of ATP content assays, infrared spectroscopic characterization and He-ion microscopy (HIM) imaging were used to investigate biomass accumulation at membranes. We show that, beyond laboratory applications and in the case of complex aqueous environments that are rich in biomass such as wastewater effluent, we observe significantly lower biofouling at carbohydrate-modified PES than at bare PES membrane surfaces.

Changes in soil moisture drive soil methane uptake along a fire regeneration chronosequence in a eucalypt forest landscape.

Disturbance associated with severe wildfires (WF) and WF simulating harvest operations can potentially alter soil methane (CH4 ) oxidation in well-aerated forest soils due to the effect on soil properties linked to diffusivity, methanotrophic activity or changes in methanotrophic bacterial community structure. However, changes in soil CH4 flux related to such disturbances are still rarely studied even though WF frequency is predicted to increase as a consequence of global climate change. We measured in-situ soil-atmosphere CH4 exchange along a wet sclerophyll eucalypt forest regeneration chronosequence in Tasmania, Australia, where the time since the last severe fire or harvesting disturbance ranged from 9 to >200 years. On all sampling occasions, mean CH4 uptake increased from most recently disturbed sites (9 year) to sites at stand 'maturity' (44 and 76 years). In stands >76 years since disturbance, we observed a decrease in soil CH4 uptake. A similar age dependency of potential CH4 oxidation for three soil layers (0.0-0.05, 0.05-0.10, 0.10-0.15 m) could be observed on incubated soils under controlled laboratory conditions. The differences in soil CH4 uptake between forest stands of different age were predominantly driven by differences in soil moisture status, which affected the diffusion of atmospheric CH4 into the soil. The observed soil moisture pattern was likely driven by changes in interception or evapotranspiration with forest age, which have been well described for similar eucalypt forest systems in south-eastern Australia. Our results imply that there is a large amount of variability in CH4 uptake at a landscape scale that can be attributed to stand age and soil moisture differences. An increase in severe WF frequency in response to climate change could potentially increase overall forest soil CH4 sinks.

Expedited solid-phase synthesis of fluorescently labeled and biotinylated aminoalkane diphenyl phosphonate affinity probes for chymotrypsin- and elastase-like serine proteases.

In this study, we report on a novel, expedited solid-phase approach for the synthesis of biotinylated and fluorescently tagged irreversible affinity based probes for the chymotrypsin and elastase-like serine proteases. The novel solid-phase biotinylation or fluorescent labeling of the aminoalkane diphenyl phosphonate warhead using commercially available Biotin-PEG-NovaTag or EDANS NovaTag resin permits rapid, facile synthesis of these reagents. We demonstrate the kinetic evaluation and utilization of a number of these irreversible inactivators for chymotrypsin-like (chymotrypsin/human cathepsin G) and elastase-like serine proteases. Encouragingly, these compounds display comparable potency against their target proteases as their N-benzyloxycarbonyl (Cbz)-protected parent compounds, from which they were derived, and function as efficient active site-directed inactivators of their target proteases. We subsequently applied the biotinylated reagents for the sensitive detection of protease species via Western blot, showing that the inactivation of the protease was specifically mediated through the active site serine. Furthermore, we also demonstrate the successful detection of serine protease species with the fluorescently labeled derivatives "in-gel", thus avoiding the need for downstream Western blotting. Finally, we also show the utility of biotinylated and pegylated affinity probes for the isolation/enrichment of serine protease species, via capture with immobilized streptavidin, and their subsequent identification via de novo sequencing. Given their selectivity of action against the serine proteases, we believe that these reagents can be exploited for the direct, rapid, and selective identification of these enzymes from biological milieu containing multiple protease subclasses.