Microbial competition for phosphorus limits the CO2 response of a mature forest.

The capacity for terrestrial ecosystems to sequester additional carbon (C) with rising CO2 concentrations depends on soil nutrient availability1,2. Previous evidence suggested that mature forests growing on phosphorus (P)-deprived soils had limited capacity to sequester extra biomass under elevated CO2 (refs. 3-6), but uncertainty about ecosystem P cycling and its CO2 response represents a crucial bottleneck for mechanistic prediction of the land C sink under climate change7. Here, by compiling the first comprehensive P budget for a P-limited mature forest exposed to elevated CO2, we show a high likelihood that P captured by soil microorganisms constrains ecosystem P recycling and availability for plant uptake. Trees used P efficiently, but microbial pre-emption of mineralized soil P seemed to limit the capacity of trees for increased P uptake and assimilation under elevated CO2 and, therefore, their capacity to sequester extra C. Plant strategies to stimulate microbial P cycling and plant P uptake, such as increasing rhizosphere C release to soil, will probably be necessary for P-limited forests to increase C capture into new biomass. Our results identify the key mechanisms by which P availability limits CO2 fertilization of tree growth and will guide the development of Earth system models to predict future long-term C storage.

CD22L Conjugation to Insulin Attenuates Insulin-Specific B Cell Activation.

Pancreatic islet-reactive B lymphocytes promote Type 1 diabetes (T1D) by presenting an antigen to islet-destructive T cells. Teplizumab, an anti-CD3 monoclonal, delays T1D onset in patients at risk, but additional therapies are needed to prevent the disease entirely. Therefore, bifunctional molecules were designed to selectively inhibit T1D-promoting anti-insulin B cells by conjugating a ligand for the B cell inhibitory receptor CD22 (i.e., CD22L) to insulin, which permit these molecules to concomitantly bind to anti-insulin B cell receptors (BCRs) and CD22. Two prototypes were synthesized: 2:2 insulin-CD22L conjugate on a 4-arm PEG backbone, and 1:1 insulin-CD22L direct conjugate. Transgenic mice (125TgSD) expressing anti-insulin BCRs provided cells for in vitro testing. Cells were cultured with constructs for 3 days, then assessed by flow cytometry. Duplicate wells with anti-CD40 simulated T cell help. A 2-insulin 4-arm PEG control caused robust proliferation and activation-induced CD86 upregulation. Anti-CD40 further boosted these effects. This may indicate that BCR-cross-linking occurs when antigens are tethered by the PEG backbone as soluble insulin alone has no effect. Addition of CD22L via the 2:2 insulin-CD22L conjugate restored B cell properties to that of controls without an additional beneficial effect. In contrast, the 1:1 insulin-CD22L direct conjugate significantly reduced anti-insulin B cell proliferation in the presence of anti-CD40. CD22L alone had no effect, and the constructs did not affect the WT B cells. Thus, multivalent antigen constructs tend to activate anti-insulin B cells, while monomeric antigen-CD22L conjugates reduce B cell activation in response to simulated T cell help and reduce pathogenic B cell numbers without harming normal cells. Therefore, monomeric antigen-CD22L conjugates warrant futher study and may be promising candidates for preclinical trials to prevent T1D without inducing immunodeficiency.

Multivalent Scaffolds to Promote B cell Tolerance.

Autoimmune diseases are characterized by aberrant immune responses toward self-antigens. Current treatments lack specificity, promoting adverse effects by broadly suppressing the immune system. Therapies that specifically target the immune cells responsible for disease are a compelling strategy to mitigate adverse effects. Multivalent formats that display numerous binding epitopes off a single scaffold may enable selective immunomodulation by eliciting signals through pathways unique to the targeted immune cells. However, the architecture of multivalent immunotherapies can vary widely, and there is limited clinical data with which to evaluate their efficacy. Here, we set forth to review the architectural properties and functional mechanisms afforded by multivalent ligands and evaluate four multivalent scaffolds that address autoimmunity by altering B cell signaling pathways. First, we address both synthetic and natural polymer backbones functionalized with a variety of small molecule, peptide, and protein ligands for probing the effects of valency and costimulation. Then, we review nanoparticles composed entirely from immune signals which have been shown to be efficacious. Lastly, we outline multivalent liposomal nanoparticles capable of displaying high numbers of protein antigens. Taken together, these examples highlight the versatility and desirability of multivalent ligands for immunomodulation and illuminate strengths and weaknesses of multivalent scaffolds for treating autoimmunity.

Novel insight into ammonium, phosphate, and iron(II) dynamics in the sediment porewater of a constructed wetland under artificial aeration through the diffusive equilibrium in thin films technique.

The analysis of porewater concentrations in constructed wetland sediments could help to understand biogeochemical processes, the sources and sinks of nutrients, and their effect on overlying water quality. In this study, we measured high-resolution porewater concentration profiles of ammonium (NH4+-N), nitrate (NO3N), phosphate (PO43--P), and ferrous iron (Fe(II)) in-situ in the Laratinga constructed wetland in Mount Barker (South Australia) using diffusive equilibration in thin films (DET) techniques. Measurements were taken under light and dark conditions, and non-aerated and aerated conditions to determine the effect on sediment porewater nutrient concentrations. Baseline surface water nutrient concentrations (NH4+-N > 36 mg L-1, PO43--P > 0.43 mg L-1) greatly exceeded water quality guideline criteria. Aeration of the water column alleviated night-time hypoxic conditions (i.e. dissolved oxygen increased from a minimum of 0.7 mg L-1 to a minimum of 4 mg L-1), and increased the redox potential in the sediment. Significant differences were present for NH4+-N, PO43--P, and Fe(II) concentrations with depth in the sediment. Ammonium concentrations in the sediment reduced under aerated conditions, presumably due to enhanced nitrification. However it was observed that PO43--P and Fe(II) concentrations increased significantly with aeration, especially under dark conditions, and were strongly correlated (R2>0.8). This was not what was hypothesised and points to complex interactions between Fe and P in the sediment. Nitrate concentrations in the sediment were below the detection limit (<0.9 mg L-1) which suggests limited nitrification-denitrification is occurring. Overall the results suggest that DET techniques are useful tools for quantifying porewater concentrations of nutrients in constructed wetlands under various environmental conditions.

Artificial aeration of an overloaded constructed wetland improves hypoxia but does not ameliorate high nitrogen loads.

High organic loadings to constructed wetlands can result in water quality issues such as low dissolved oxygen and high ammonium concentrations, with artificial aeration a potential mitigation option. This study compared baseline (no aeration - NA), continuous aeration (CA), and intermittent aeration (IA) conditions to improve water quality in a tertiary treatment free water surface constructed wetland (FWS CW) with night time hypoxia/anoxia, and high nutrient concentrations. The response variables included dissolved oxygen (DO), total nitrogen (TN), ammonium nitrogen (NH4+-N), nitrate nitrogen (NO3--N), total phosphorus (TP), phosphate (PO43--P), and dissolved organic carbon (DOC). In situ aeration and monitoring was performed from April to June 2021 in a large, field-scale FWS CW, the Laratinga wetlands Mount Barker, South Australia. The results demonstrated that DO increased by an average 2.11 mg L-1 from NA to CA during the night and 1.26 mg L-1 and 1.84 mg L-1 from NA to IA during the night and day respectively when averaging over the basins. The C/N ratio was very low and there was no significant influence of DO on DOC concentrations. There was no significant difference in TN concentrations with the application of aeration aside from a decrease in the channel at night from NA to IA, and an increase in NH4+-N resulted under IA compared with NA in Basin 1 and 2 during the day. This implies that the N loadings exceeded the wetland's ability to complete nutrient conversions at a rate that aligns with input rate. The concentrations of NO3--N increased at night under CA and IA treatments suggesting that some nitrification was promoted, or there was inhibition of dissimilatory nitrate reduction to ammonium. The concentrations of TP and PO43--P significantly increased with the aeration compared with no aeration, however there was no difference between the aeration treatments. This suggested that increased sediment resuspension during aeration increased P in the water. There was no change in DOC with the application of aeration. Overall, the DO increased with aeration application and may be able to better support the wetland ecology; however, the Laratinga wetland is overloaded and the capacity of the wetland to effectively transform and remove nutrients is inhibited, even with the application of artificial aeration.

Lectin Drug Conjugates Targeting High Mannose N-Glycans.

Cancer-associated alterations to glycosylation have been shown to aid cancer development and progression. An increased abundance of high mannose N-glycans has been observed in several cancers. Here, we describe the preparation of lectin drug conjugates (LDCs) that permit toxin delivery to cancer cells presenting high mannose N-glycans. Additionally, we demonstrate that cancer cells presenting low levels of high mannose N-glycans can be rendered sensitive to the LDCs by co-treatment with a type I mannosidase inhibitor. Our findings establish that an increased abundance of high mannose N-glycans in the glycocalyx of cancer cells can be leveraged to enable toxin delivery.

Ericoid shrubs shape fungal communities and suppress organic matter decomposition in boreal forests.

Mycorrhizal fungi associated with boreal trees and ericaceous shrubs are central actors in organic matter (OM) accumulation through their belowground carbon allocation, their potential capacity to mine organic matter for nitrogen (N) and their ability to suppress saprotrophs. Yet, interactions between co-occurring ectomycorrhizal fungi (EMF), ericoid mycorrhizal fungi (ERI), and saprotrophs are poorly understood. We used a long-term (19 yr) plant functional group manipulation experiment with removals of tree roots, ericaceous shrubs and mosses and analysed the responses of different fungal guilds (assessed by metabarcoding) and their interactions in relation to OM quality (assessed by mid-infrared spectroscopy and nuclear magnetic resonance) and decomposition (litter mesh-bags) across a 5000-yr post-fire boreal forest chronosequence. We found that the removal of ericaceous shrubs and associated ERI changed the composition of EMF communities, with larger effects occurring at earlier stages of the chronosequence. Removal of shrubs was associated with enhanced N availability, litter decomposition and enrichment of the recalcitrant OM fraction. We conclude that increasing abundance of slow-growing ericaceous shrubs and the associated fungi contributes to increasing nutrient limitation, impaired decomposition and progressive OM accumulation in boreal forests, particularly towards later successional stages. These results are indicative of the contrasting roles of EMF and ERI in regulating belowground OM storage.

Determination of Bio-Based Fertilizer Composition Using Combined NIR and MIR Spectroscopy: A Model Averaging Approach.

Application of bio-based fertilizers is considered a practical solution to enhance soil fertility and maintain soil quality. However, the composition of bio-based fertilizers needs to be quantified before their application to the soil. Non-destructive techniques such as near-infrared (NIR) and mid-infrared (MIR) are generally used to quantify the composition of bio-based fertilizers in a speedy and cost-effective manner. However, the prediction performances of these techniques need to be quantified before deployment. With this motive, this study investigates the potential of these techniques to characterize a diverse set of bio-based fertilizers for 25 different properties including nutrients, minerals, heavy metals, pH, and EC. A partial least square model with wavelength selection is employed to estimate each property of interest. Then a model averaging, approach is tested to examine if combining model outcomes of NIR with MIR could improve the prediction performances of these sensors. In total, 17 of the 25 elements could be predicted to have a good performance status using individual spectral methods. Combining model outcomes of NIR with MIR resulted in an improvement, increasing the number of properties that could be predicted from 17 to 21. Most notably the improvement in prediction performance was observed for Cd, Cr, Zn, Al, Ca, Fe, S, Cu, Ec, and Na. It was concluded that the combined use of NIR and MIR spectral methods can be used to monitor the composition of a diverse set of bio-based fertilizers.

Long-term water quality response to increased hydraulic loadings in a field-scale free water surface constructed wetland treating domestic effluent.

There is limited understanding of how constructed wetland (CW) water quality may change over time in response to increased wastewater nutrient and hydraulic loadings. We evaluated long-term water quality trends and drivers for a full-scale (8.19 ha) free water surface CW that was developed in 2001 for the treatment of increasing amounts of pre-treated domestic wastewater from the township of Mount Barker, South Australia. Water quality parameter concentrations and loads, hydraulic loadings rates, trend direction assessments (TDAs), and water quality parameter removal efficiencies were analysed over the study period. The wetland received an annual average loading rate of 947, 19644, 31039, 18140, 2985, and 807 kg year-1 for BOD5, TN, NH4-N, TKN-N, NOx-N, and TP respectively and removed on average 8%, 72%, 73%, 78%, 12% and -246% of these loadings respectively. The average influent concentrations for the study period were 2.6, 42.3, 40.6, 35.9, 9.0, and 1.9 mg L-1 for BOD5, TN, NH4-N, TKN-N, NOx-N, and TP respectively. Average concentration removal rates over the study period were 50%, 39%, 40%, 15%, -216% and -600.5% for TN, NH4-N, TKN-N, NOx-N, BOD5 and TP respectively, suggesting that nitrogen was only partly assimilated by the wetland and it was a source of organic material and phosphorus. Using seasonally and inflow rate adjusted data, TDAs predicted virtually certain increases in TN, NH4-N, and TKN-N influent concentrations over time, a decline in NOx-N, no trend in BOD5, and a possible decreasing trend in TP. The inflow explained variance accounted for approximately 50% of the variation in TN, NH4-N and TKN-N effluent concentrations. Annual removal efficiencies of N declined with increasing hydraulic loads, and hydraulic loading rates varied with management practices. Seasonal analysis showed that N removal was greater during summer and lower in winter. Due to local population growth and various management practices, hydraulic loading is variable and has often exceeded design targets. Our findings indicate the long-term performance of CWs need to be closely monitored, as water quality can deteriorate due to increased hydraulic loadings.

Long-Acting BMS-378806 Analogues Stabilize the State-1 Conformation of the Human Immunodeficiency Virus Type 1 Envelope Glycoproteins.

During human immunodeficiency virus type 1 (HIV-1) entry into cells, the viral envelope glycoprotein (Env) trimer [(gp120/gp41)3] binds the receptors CD4 and CCR5 and fuses the viral and cell membranes. CD4 binding changes Env from a pretriggered (state-1) conformation to more open downstream conformations. BMS-378806 (here called BMS-806) blocks CD4-induced conformational changes in Env important for entry and is hypothesized to stabilize a state-1-like Env conformation, a key vaccine target. Here, we evaluated the effects of BMS-806 on the conformation of Env on the surface of cells and virus-like particles. BMS-806 strengthened the labile, noncovalent interaction of gp120 with the Env trimer, enhanced or maintained the binding of most broadly neutralizing antibodies, and decreased the binding of poorly neutralizing antibodies. Thus, in the presence of BMS-806, the cleaved Env on the surface of cells and virus-like particles exhibits an antigenic profile consistent with a state-1 conformation. We designed novel BMS-806 analogues that stabilized the Env conformation for several weeks after a single application. These long-acting BMS-806 analogues may facilitate enrichment of the metastable state-1 Env conformation for structural characterization and presentation to the immune system.IMPORTANCE The envelope glycoprotein (Env) spike on the surface of human immunodeficiency virus type 1 (HIV-1) mediates the entry of the virus into host cells and is also the target for antibodies. During virus entry, Env needs to change shape. Env flexibility also contributes to the ability of HIV-1 to evade the host immune response; many shapes of Env raise antibodies that cannot recognize the functional Env and therefore do not block virus infection. We found that an HIV-1 entry inhibitor, BMS-806, stabilizes the functional shape of Env. We developed new variants of BMS-806 that stabilize Env in its natural state for long periods of time. The availability of such long-acting stabilizers of Env shape will allow the natural Env conformation to be characterized and tested for efficacy as a vaccine.

The Road to Structurally Defined ß-Glucans.

ß-glucans are polymers of glucose that have been isolated from a variety of organisms. Isolated ß-glucans have been used for medical purposes for centuries; however, efforts to define the biological activities of ß-glucans experimentally were initiated in the 1940's. The diversity of structure associated with isolated ß-glucans has impeded said investigations, and efforts to leverage the biological activity of ß-glucans for clinical applications. In recognition of the need for defined ß-glucans that retain the biological activity of isolated ß-glucans, considerable investment has been made to facilitate the synthesis of structurally defined ß-glucans. Here, we review the different approaches that have been applied to prepare ß-glucans. In addition, we summarize the approaches that have been utilized to conjugate ß-glucans to proteins.

Functionalized High Mannose-Specific Lectins for the Discovery of Type†I Mannosidase Inhibitors.

An engineered cyanovirin-N homologue that exhibits specificity for high mannose N-glycans has been constructed to aid type I α 1,2-mannosidase inhibitor discovery and development. Engineering the lectins C-terminus permitted facile functionalization with fluorophores via a sortase and click strategy. The resulting lectin constructs exhibit specificity for cells presenting high mannose N-glycans. Importantly, these lectin constructs can also be applied to specifically assess changes in cell surface glycosylation induced by type I mannosidase inhibitors. Testing the utility of these lectin constructs led to the discovery of type I mannosidase inhibitors with nanomolar potency. Cumulatively, these findings reveal the specificity and utility of the functionalized cyanovirin-N homologue constructs, and highlight their potential in analytical contexts that require high mannose-specific lectins.

Strain-Dependent Activation and Inhibition of Human Immunodeficiency Virus Entry by a Specific PF-68742 Stereoisomer.

Human immunodeficiency virus (HIV-1) entry into cells is mediated by the viral envelope glycoprotein (Env) trimer, which consists of three gp120 exterior glycoproteins and three gp41 transmembrane glycoproteins. When gp120 binds sequentially to the receptors CD4 and CCR5 on the target cell, the metastable Env trimer is triggered to undergo entry-related conformational changes. PF-68742 is a small molecule that inhibits the infection of a subset of HIV-1 strains by interfering with an Env function other than receptor binding. Determinants of HIV-1 resistance to PF-68742 map to the disulfide loop and fusion peptide of gp41. Of the four possible PF-68742 stereoisomers, only one, MF275, inhibited the infection of CD4-positive CCR5-positive cells by some HIV-1 strains. MF275 inhibition of these HIV-1 strains occurred after CD4 binding but before the formation of the gp41 six-helix bundle. Unexpectedly, MF275 activated the infection of CD4-negative CCR5-positive cells by several HIV-1 strains resistant to the inhibitory effects of the compound in CD4-positive target cells. In contrast to CD4 complementation by CD4-mimetic compounds, activation of CD4-independent infection by MF275 did not depend upon the availability of the gp120 Phe 43 cavity. Sensitivity to inhibitors indicates that MF275-activated virus entry requires formation/exposure of the gp41 heptad repeat (HR1) as well as CCR5 binding. MF275 apparently activates a virus entry pathway parallel to that triggered by CD4 and CD4-mimetic compounds. Strain-dependent divergence in Env conformational transitions allows different outcomes, inhibition or activation, in response to MF275. Understanding the mechanisms of MF275 activity should assist efforts to optimize its utility.IMPORTANCE Envelope glycoprotein (Env) spikes on the surface of human immunodeficiency virus (HIV-1) bind target cell receptors, triggering changes in the shape of Env. We studied a small molecule, MF275, that also induced shape changes in Env. The consequences of MF275 interaction with Env depended on the HIV-1 strain, with infection by some viruses inhibited and infection by other viruses enhanced. These studies reveal the strain-dependent diversity of HIV-1 Envs as they undergo shape changes in proceeding down the entry pathway. Appreciation of this diversity will assist attempts to develop broadly active inhibitors of HIV-1 entry.

Effects of plant functional group removal on structure and function of soil communities across contrasting ecosystems.

Loss of plant diversity has an impact on ecosystems worldwide, but we lack a mechanistic understanding of how this loss may influence below-ground biota and ecosystem functions across contrasting ecosystems in the long term. We used the longest running biodiversity manipulation experiment across contrasting ecosystems in existence to explore the below-ground consequences of 19 years of plant functional group removals for each of 30 contrasting forested lake islands in northern Sweden. We found that, against expectations, the effects of plant removals on the communities of key groups of soil organisms (bacteria, fungi and nematodes), and organic matter quality and soil ecosystem functioning (decomposition and microbial activity) were relatively similar among islands that varied greatly in productivity and soil fertility. This highlights that, in contrast to what has been shown for plant productivity, plant biodiversity loss effects on below-ground functions can be relatively insensitive to environmental context or variation among widely contrasting ecosystems.

Type†II Anion Relay Chemistry: Exploiting Bifunctional Weinreb Amide Linchpins for the One-Pot Synthesis of Differentiated 1,3-Diketones, Pyrans, and Spiroketals.

The design, synthesis, and validation of new highly effective bifunctional linchpins for type II anion relay chemistry (ARC) has been achieved. The mechanistically novel negative-charge migration that comprises the Brook rearrangement is now initiated by a stabilized tetrahedral intermediate, which is generated by nucleophilic addition to a Weinreb amide, rather than by a simple oxyanion that is generated from an epoxide. As a result, the linchpin preserves the carbonyl functionality in the ARC adducts, thus permitting access to functionally complex systems in a single flask without the need for further chemical manipulations. This tactic was validated with the one-pot preparation of monoprotected 1,3-diketones as well as pyran and spiroketal scaffolds, depending on the choice of nucleophile, electrophile, and work-up conditions.

Network analysis reveals that bacteria and fungi form modules that correlate independently with soil parameters.

Network and multivariate statistical analyses were performed to determine interactions between bacterial and fungal community terminal restriction length polymorphisms as well as soil properties in paired woodland and pasture sites. Canonical correspondence analysis (CCA) revealed that shifts in woodland community composition correlated with soil dissolved organic carbon, while changes in pasture community composition correlated with moisture, nitrogen and phosphorus. Weighted correlation network analysis detected two distinct microbial modules per land use. Bacterial and fungal ribotypes did not group separately, rather all modules comprised of both bacterial and fungal ribotypes. Woodland modules had a similar fungal : bacterial ribotype ratio, while in the pasture, one module was fungal dominated. There was no correspondence between pasture and woodland modules in their ribotype composition. The modules had different relationships to soil variables, and these contrasts were not detected without the use of network analysis. This study demonstrated that fungi and bacteria, components of the soil microbial communities usually treated as separate functional groups as in a CCA approach, were co-correlated and formed distinct associations in these adjacent habitats. Understanding these distinct modular associations may shed more light on their niche space in the soil environment, and allow a more realistic description of soil microbial ecology and function.

C/N ratio drives soil actinobacterial cellobiohydrolase gene diversity.

Cellulose accounts for approximately half of photosynthesis-fixed carbon; however, the ecology of its degradation in soil is still relatively poorly understood. The role of actinobacteria in cellulose degradation has not been extensively investigated despite their abundance in soil and known cellulose degradation capability. Here, the diversity and abundance of the actinobacterial glycoside hydrolase family 48 (cellobiohydrolase) gene in soils from three paired pasture-woodland sites were determined by using terminal restriction fragment length polymorphism (T-RFLP) analysis and clone libraries with gene-specific primers. For comparison, the diversity and abundance of general bacteria and fungi were also assessed. Phylogenetic analysis of the nucleotide sequences of 80 clones revealed significant new diversity of actinobacterial GH48 genes, and analysis of translated protein sequences showed that these enzymes are likely to represent functional cellobiohydrolases. The soil C/N ratio was the primary environmental driver of GH48 community compositions across sites and land uses, demonstrating the importance of substrate quality in their ecology. Furthermore, mid-infrared (MIR) spectrometry-predicted humic organic carbon was distinctly more important to GH48 diversity than to total bacterial and fungal diversity. This suggests a link between the actinobacterial GH48 community and soil organic carbon dynamics and highlights the potential importance of actinobacteria in the terrestrial carbon cycle.

Embolic activity during in vivo cardiopulmonary bypass.

Neurologic injury after cardiac surgery is principally associated with emboli. Although much work has focused on surgical sources of emboli, less attention has been focused on emboli associated with the heart-lung machine. We tested whether emboli are associated with discrete processes during cardiopulmonary bypass (CPB). One hundred patients undergoing cardiothoracic surgery were enrolled between April 2008 and May 2011 at a single medical center. During each surgical procedure, emboli were counted in three CPB locations: the venous side (Channel 1), before the arterial line filter (Channel 2), and after the arterial line filter (Channel 3). We used prespecified event markers to identify perfusionist interventions. Identical circuits were used on all patients. Of the 100 patients enrolled, 62 underwent isolated coronary artery bypass grafting (CABG), 17 underwent isolated valve operations, and 21 underwent CABG plus valve. Median counts across Channels 1, 2, and 3 were 69,853, 3,017, and 1,251, respectively. The greatest contributor to emboli in Channels 1, 2, and 3, respectively, were achieving the calculated CPB flow, opening of the electronic arterial line clamp, and introducing a hemofilter. The circuit technology was efficient in reducing total emboli counts from Channels 1-2 irrespective of the size of the emboli. Nearly 71% of all emboli 30-100 microm in size were removed from the circuit between Channels 2 and 3. No significant association was found between emboli counts and S100B release. Emboli occur frequently during CPB and are predominantly associated with the initiation of bypass, operation of the electronic arterial line clamp, and the initiation of a hemofilter. Continued work to reduce the occurrence of emboli is warranted.

Carbon-phosphorus cycle models overestimate CO2 enrichment response in a mature Eucalyptus forest.

The importance of phosphorus (P) in regulating ecosystem responses to climate change has fostered P-cycle implementation in land surface models, but their CO2 effects predictions have not been evaluated against measurements. Here, we perform a data-driven model evaluation where simulations of eight widely used P-enabled models were confronted with observations from a long-term free-air CO2 enrichment experiment in a mature, P-limited Eucalyptus forest. We show that most models predicted the correct sign and magnitude of the CO2 effect on ecosystem carbon (C) sequestration, but they generally overestimated the effects on plant C uptake and growth. We identify leaf-to-canopy scaling of photosynthesis, plant tissue stoichiometry, plant belowground C allocation, and the subsequent consequences for plant-microbial interaction as key areas in which models of ecosystem C-P interaction can be improved. Together, this data-model intercomparison reveals data-driven insights into the performance and functionality of P-enabled models and adds to the existing evidence that the global CO2-driven carbon sink is overestimated by models.