U-Net Convolutional Neural Network for Mapping Natural Vegetation and Forest Types from Landsat Imagery in Southeastern Australia.

Accurate and comparable annual mapping is critical to understanding changing vegetation distribution and informing land use planning and management. A U-Net convolutional neural network (CNN) model was used to map natural vegetation and forest types based on annual Landsat geomedian reflectance composite images for a 500 km × 500 km study area in southeastern Australia. The CNN was developed using 2018 imagery. Label data were a ten-class natural vegetation and forest classification (i.e., Acacia, Callitris, Casuarina, Eucalyptus, Grassland, Mangrove, Melaleuca, Plantation, Rainforest and Non-Forest) derived by combining current best-available regional-scale maps of Australian forest types, natural vegetation and land use. The best CNN generated using six Landsat geomedian bands as input produced better results than a pixel-based random forest algorithm, with higher overall accuracy (OA) and weighted mean F1 score for all vegetation classes (93 vs. 87% in both cases) and a higher Kappa score (86 vs. 74%). The trained CNN was used to generate annual vegetation maps for 2000-2019 and evaluated for an independent test area of 100 km × 100 km using statistics describing accuracy regarding the label data and temporal stability. Seventy-six percent of pixels did not change over the 20 years (2000-2019), and year-on-year results were highly correlated (94-97% OA). The accuracy of the CNN model was further verified for the study area using 3456 independent vegetation survey plots where the species of interest had ≥ 50% crown cover. The CNN showed an 81% OA compared with the plot data. The model accuracy was also higher than the label data (76%), which suggests that imperfect training data may not be a major obstacle to CNN-based mapping. Applying the CNN to other regions would help to test the spatial transferability of these techniques and whether they can support the automated production of accurate and comparable annual maps of natural vegetation and forest types required for national reporting.

Global groundwater droughts are more severe than they appear in hydrological models: An investigation through a Bayesian merging of GRACE and GRACE-FO data with a water balance model.

Realistic representation of hydrological drought events is increasingly important in world facing decreased freshwater availability. Index-based drought monitoring systems are often adopted to represent the evolution and distribution of hydrological droughts, which mainly rely on hydrological model simulations to compute these indices. Recent studies, however, indicate that model derived water storage estimates might have difficulties in adequately representing reality. Here, a novel Markov Chain Monte Carlo - Data Assimilation (MCMC-DA) approach is implemented to merge global Terrestrial Water Storage (TWS) changes from the Gravity Recovery And Climate Experiment (GRACE) and its Follow On mission (GRACE-FO) with the water storage estimations derived from the W3RA water balance model. The modified MCMC-DA derived summation of deep-rooted soil and groundwater storage estimates is then used to compute 0.5∘ standardized groundwater drought indices globally to show the impact of GRACE/GRACE-FO DA on a global index-based hydrological drought monitoring system. Our numerical assessment covers the period of 2003-2021, and shows that integrating GRACE/GRACE-FO data modifies the seasonality and inter-annual trends of water storage estimations. Considerable increases in the length and severity of extreme droughts are found in basins that exhibited multi-year water storage fluctuations and those affected by climate teleconnections.

Untangling fuel, weather and management effects on fire severity: Insights from large-sample LiDAR remote sensing analysis of conditions preceding the 2019-20 Australian wildfires.

Evaluation of fire severity reduction strategies requires the quantification of intervention outcomes and, more broadly, the extent to which fuel characteristics affect fire severity. However, investigations are currently limited by the availability of accurate data on fire severity predictors, particularly relating to fuel. Here, we used airborne LiDAR data collected before the 2019-20 Australian Black Summer fires to investigate the contribution of fuel structure to fire severity under a range of weather conditions. Fire severity was estimated using the Relative Burn Ratio calculated from Sentinel-2 optical remote sensing imagery. We modelled the effects of various fuel structure estimates and other environmental predictors using Random Forest models. In addition to variables estimated at each observation point, we investigated the influence of surrounding landscape characteristics using an innovative method to estimate fireline progression direction. Our models explained 63-76% of fire severity variance using parsimonious predictor sets. Fuel cover in the understorey and canopy, and vertical vegetation heterogeneity, were positively associated with fire severity. Up-fire burnt area and recent planned and unplanned fire reduced fire severity, whereby unplanned fire provided a longer-lasting reduction of fire severity (up to 15 years) than planned fire (up to 10 years). Although fuel structure and land management effects were important predictors, weather and canopy height effects were dominant. By mapping continuous interactions between weather and fuel-related variables, we found strong evidence of diminishing fuel effects below 20-40% relative air humidity. While our findings suggest that land management interventions can provide meaningful fire severity reduction, they also highlight the risk of warmer and drier future climates constraining these advantages.

High-resolution (1 km) Köppen-Geiger maps for 1901-2099 based on constrained CMIP6 projections.

We introduce Version 2 of our widely used 1-km Köppen-Geiger climate classification maps for historical and future climate conditions. The historical maps (encompassing 1901-1930, 1931-1960, 1961-1990, and 1991-2020) are based on high-resolution, observation-based climatologies, while the future maps (encompassing 2041-2070 and 2071-2099) are based on downscaled and bias-corrected climate projections for seven shared socio-economic pathways (SSPs). We evaluated 67 climate models from the Coupled Model Intercomparison Project phase 6 (CMIP6) and kept a subset of 42 with the most plausible CO2-induced warming rates. We estimate that from 1901-1930 to 1991-2020, approximately 5% of the global land surface (excluding Antarctica) transitioned to a different major Köppen-Geiger class. Furthermore, we project that from 1991-2020 to 2071-2099, 5% of the land surface will transition to a different major class under the low-emissions SSP1-2.6 scenario, 8% under the middle-of-the-road SSP2-4.5 scenario, and 13% under the high-emissions SSP5-8.5 scenario. The Köppen-Geiger maps, along with associated confidence estimates, underlying monthly air temperature and precipitation data, and sensitivity metrics for the CMIP6 models, can be accessed at www.gloh2o.org/koppen .

Evolutionary diversification of defensins and cathelicidins in birds and primates.

Divergent evolution for more than 310 million years has resulted in an avian immune system that is complex and more compact than that of primates, sharing much of its structure and functions. Not surprisingly, well conserved ancient host defense molecules, such as defensins and cathelicidins, have diversified over time. In this review, we describe how evolution influenced the host defense peptides repertoire, its distribution, and the relationship between structure and biological functions. Marked features of primate and avian HDPs are linked to species-specific characteristics, biological requirements, and environmental challenge.

Avian surfactant protein (SP)-A2 first arose in an early tetrapod before the divergence of amphibians and gradually lost the collagen domain.

The air-liquid interface of the mammalian lung is lined with pulmonary surfactants, a mixture of specific proteins and lipids that serve a dual purpose-enabling air-breathing and protection against pathogens. In mammals, surfactant proteins A (SP-A) and D (SP -D) are involved in innate defence of the lung. Birds seem to lack the SP-D gene, but possess SP-A2, an additional SP-A-like gene. Here we investigated the evolution of the SP-A and SP-D genes using computational gene prediction, homology, simulation modelling and phylogeny with published avian and other vertebrate genomes. PCR was used to confirm the identity and expression of SP-A analogues in various tissue homogenates of zebra finch and turkey. In silico analysis confirmed the absence of SP-D-like genes in all 47 published avian genomes. Zebra finch and turkey SP-A1 and SP-A2 sequences, confirmed by PCR of lung homogenates, were compared with sequenced and in silico predicted vertebrate homologs to construct a phylogenetic tree. The collagen domain of avian SP-A1, especially that of zebra finch, was dramatically shorter than that of mammalian SP-A. Amphibian and reptilian genomes also contain avian-like SP-A2 protein sequences with a collagen domain. NCBI Gnomon-predicted avian and alligator SP-A2 proteins all lacked the collagen domain completely. Both avian SP-A1 and SP-A2 sequences form separate clades, which are most closely related to their closest relatives, the alligators. The C-terminal carbohydrate recognition domain (CRD) of zebra finch SP-A1 was structurally almost identical to that of rat SP-A. In fact, the CRD of SP-A is highly conserved among all the vertebrates. Birds retained a truncated version of mammalian type SP-A1 as well as a non-collagenous C-type lectin, designated SP-A2, while losing the large collagenous SP-D lectin, reflecting their evolutionary trajectory towards a unidirectional respiratory system. In the context of zoonotic infections, how these evolutionary changes affect avian pulmonary surface protection is not clear.

Innate Immune Training of Human Macrophages by Cathelicidin Analogs.

Trained innate immunity can be induced in human macrophages by microbial ligands, but it is unknown if exposure to endogenous alarmins such as cathelicidins can have similar effects. Previously, we demonstrated sustained protection against infection by the chicken cathelicidin-2 analog DCATH-2. Thus, we assessed the capacity of cathelicidins to induce trained immunity. PMA-differentiated THP-1 (dTHP1) cells were trained with cathelicidin analogs for 24 hours and restimulated after a 3-day rest period. DCATH-2 training of dTHP-1 cells amplified their proinflammatory cytokine response when restimulated with TLR2/4 agonists. Trained cells displayed a biased cellular metabolism towards mTOR-dependent aerobic glycolysis and long-chain fatty acid accumulation and augmented microbicidal activity. DCATH-2-induced trained immunity was inhibited by histone acetylase inhibitors, suggesting epigenetic regulation, and depended on caveolae/lipid raft-mediated uptake, MAPK p38 and purinergic signaling. To our knowledge, this is the first report of trained immunity by host defense peptides.

Novel insights in antimicrobial and immunomodulatory mechanisms of action of PepBiotics CR-163 and CR-172.

OBJECTIVES: Our group recently developed a new group of antimicrobial peptides termed PepBiotics, of which peptides CR-163 and CR-172 showed optimized antibacterial activity against Pseudomonas aeruginosa and Staphylococcus aureus without inducing antimicrobial resistance. In this study, the antibacterial mechanism of action and the immunomodulatory activity of these two PepBiotics was explored. METHODS: RAW264.7 cells were used to determine the ability of PepBiotics to neutralize Lipopolysaccharide (LPS)-and Lipoteichoic acid (LTA)-induced activation of macrophages. Isothermal titration calorimetry and competition assays with dansyl-labeled polymyxin B determined binding characteristics to LPS and LTA. Combined bacterial killing with subsequent macrophage activation assays was performed to determine so-called 'silent killing'. Finally, flow cytometry of peptide-treated genetically engineered Escherichia coli expressing Green Fluorescent Protein (GFP) and mCherry in the cytoplasm and periplasm, respectively, further established the antimicrobial mechanism of PepBiotics. RESULTS: Both CR-163 and CR-172 were shown to have broad-spectrum activity against ESKAPE pathogens and E. coli using a membranolytic mechanism of action. PepBiotics could exothermically bind LPS/LTA and were able to replace polymyxin B. Finally, it was demonstrated that bacteria killed by PepBiotics were less prone to stimulate immune cells, contrary to gentamicin and heat-killed bacteria that still elicited a strong immune response. CONCLUSIONS: These studies highlight the multifunctional nature of the two peptide antibiotics as both broad-spectrum antimicrobial and immunomodulator. Their ability to kill bacteria and reduce unwanted subsequent immune activation is a major advantage and highlights their potential for future therapeutic use.

d-enantiomers of CATH-2 enhance the response of macrophages against Streptococcus suis serotype 2.

Introduction: Due to the increase of antibiotic resistant bacterial strains, there is an urgent need for development of alternatives to antibiotics. Cathelicidins can be such an alternative to antibiotics having both a direct antimicrobial capacity as well as an immunomodulatory function. Previously, the full d-enantiomer of chicken cathelicidin-2 (d-CATH-2) has shown to prophylactically protect chickens against infection 7 days post hatch when administered in ovo three days before hatch. Objectives: To further evaluate d-CATH-2 in mammals as a candidate for an alternative to antibiotics.In this study, the prophylactic capacity of d-CATH-2 and two truncated derivatives, d-C(1-21) and d-C(4-21), was determined in mammalian cells. Methods: Antibacterial assays; immune cell differentiation and modulation; cytotoxicity, isothermal titration calorimetry; in vivo prophylactic capacity of peptides in an S. suis infection model. Results: d-CATH-2 and its derivatives were shown to have a strong direct antibacterial capacity against four different S. suis serotype 2 strains (P1/7, S735, D282, and OV625) in bacterial medium and even stronger in cell culture medium. In addition, d-CATH-2 and its derivatives ameliorated the efficiency of mouse bone marrow-derived macrophages (BMDM) and skewed mouse bone marrow-derived dendritic cells (BMDC) towards cells with a more macrophage-like phenotype. The peptides directly bind lipoteichoic acid (LTA) and inhibit LTA-induced activation of macrophages. In addition, S. suis killed by the peptide was unable to further activate mouse macrophages, which indicates that S. suis was eliminated by the previously reported silent killing mechanism. Administration of d-C(1-21) at 24 h or 7 days before infection resulted in a small prophylactic protection with reduced disease severity and reduced mortality of the treated mice. Conclusion: d-enantiomers of CATH-2 show promise as anti-infectives against pathogenic S. suis for application in mammals.

Increased livestock weight gain from improved water quality in farm dams: A cost-benefit analysis.

Access to water is a critical aspect of livestock production, although the relationship between livestock weight gain and water quality remains poorly understood. Previous work has shown that water quality of poorly managed farm dams can be improved by fencing and constructing hardened watering points to limit stock access to the dam, and revegetation to filter contaminant inflow. Here we use cattle weight gain data from three North American studies to develop a cost-benefit analysis for the renovation of farm dams to improve water quality and, in turn, promote cattle weight gain on farms in south-eastern Australia. Our analysis indicated a strong likelihood of positive results and suggested there may be substantial net economic benefit from renovating dams in poor condition to improve water quality. The average per-farm Benefit-Cost Ratios based on deterministic assumptions was 1.5 for New South Wales (NSW) and 3.0 for Victoria in areas where rainfall exceeds 600mm annually. Our analyses suggested that cattle on farms in NSW and Victoria would need to experience additional weight gain from switching to clean water of at least 6.5% and 1.8% per annum respectively, to break even in present value terms. Monte Carlo simulation based on conservative assumptions indicated that the probability of per-farm benefits exceeding costs was greater than 70%. We recommend localised experiments to assess the impact of improved water quality on livestock weight gain in Australian conditions to confirm these expectations empirically.

Outer Membrane Vesicles Protect Gram-Negative Bacteria against Host Defense Peptides.

Host defense peptides (HDPs) are part of the innate immune system and constitute a first line of defense against invading pathogens. They possess antimicrobial activity against a broad spectrum of pathogens. However, pathogens have been known to adapt to hostile environments. Therefore, the bacterial response to treatment with HDPs was investigated. Previous observations suggested that sublethal concentrations of HDPs increase the release of outer membrane vesicles (OMVs) in Escherichia coli. First, the effects of sublethal treatment with HDPs CATH-2, PMAP-36, and LL-37 on OMV release of several Gram-negative bacteria were analyzed. Treatment with PMAP-36 and CATH-2 induced release of OMVs, but treatment with LL-37 did not. The OMVs were further characterized with respect to morphological properties. The HDP-induced OMVs often had disc-like shapes. The beneficial effect of bacterial OMV release was studied by determining the susceptibility of E. coli toward HDPs in the presence of OMVs. The minimal bactericidal concentration was increased in the presence of OMVs. It is concluded that OMV release is a means of bacteria to dispose of HDP-affected membrane. Furthermore, OMVs act as a decoy for HDPs and thereby protect the bacterium. IMPORTANCE Antibiotic resistance is a pressing problem and estimated to be a leading cause of mortality by 2050. Antimicrobial peptides, also known as host defense peptides (HDPs), and HDP-derived antimicrobials have potent antimicrobial activity and high potential as alternatives to antibiotics due to low resistance development. Some resistance mechanisms have developed in bacteria, and complete understanding of bacterial defense against HDPs will aid their use in the clinic. This study provides insight into outer membrane vesicles (OMVs) as potential defense mechanisms against HDPs, which will allow anticipation of unforeseen resistance to HDPs in clinical use and possibly prevention of bacterial resistance by the means of OMVs.

PepBiotics, novel cathelicidin-inspired antimicrobials to fight pulmonary bacterial infections.

BACKGROUND: Antimicrobial peptides are considered potential alternatives to antibiotics. Here we describe the antibacterial properties of a family of novel cathelicidin-related (CR-) peptides, which we named PepBiotics, against bacteria typically present in cystic fibrosis (CF) patients. METHODS: Broth dilution assays were used to determine antibacterial activity of PepBiotics under physiological conditions, as well as development of bacterial resistance against these peptides. Toxicity was tested in mice and cell cultures while molecular interactions of PepBiotics with bacterial membrane components was determined using CD, ITC and LPS/LTA induced macrophage studies. RESULTS: A relatively small number of PepBiotics remained highly antibacterial against CF-related respiratory pathogens Pseudomonas aeruginosa and Staphylococcus aureus, at high ionic strength and low pH. Interestingly, these PepBiotics also prevented LPS/LTA induced activation of macrophages and was shown to be non-toxic to primary human nasal epithelial cells. Furthermore, both P. aeruginosa and S. aureus were unable to induce resistance against CR-163 and CR-172, two PepBiotics selected for their excellent antimicrobial and immunomodulatory properties. Toxicity studies in mice indicated that intratracheal administration of CR-163 was well tolerated in vivo. Finally, interaction of CR-163 with bacterial-type anionic membranes but not with mammalian-type (zwitterionic lipid) membranes was confirmed using ITC and 31P solid state NMR. CONCLUSIONS: PepBiotics are a promising novel class of highly active antimicrobial peptides, of which CR-163 showed the most potential for treatment of clinically relevant (CF-) pathogens in physiological conditions. GENERAL SIGNIFICANCE: These observations emphasize the therapeutic potential of PepBiotics against CF-related bacterial respiratory infections.

Chicken CATH-2 Increases Antigen Presentation Markers on Chicken Monocytes and Macrophages.

BACKGROUND: Cathelicidins are a family of Host Defense Peptides (HDPs), that play an important role in the innate immune response. They exert both broad-spectrum antimicrobial activity against pathogens, and strong immunomodulatory functions that affect the response of innate and adaptive immune cells. OBJECTIVE: The aim of this study was to investigate immunomodulation by the chicken cathelicidin CATH-2 and compare its activities to those of the human cathelicidin LL-37. METHODS: Chicken macrophages and chicken monocytes were incubated with cathelicidins. Activation of immune cells was determined by measuring surface markers Mannose Receptor Ctype 1 (MRC1) and MHC-II. Cytokine production was measured by qPCR and nitric oxide production was determined using the Griess assay. Finally, the effect of cathelicidins on phagocytosis was measured using carboxylate-modified polystyrene latex beads. RESULTS: CATH-2 and its all-D enantiomer D-CATH-2 increased MRC1 and MHC-II expression, markers for antigen presentation, on primary chicken monocytes, whereas LL-37 did not. D-CATH- 2 also increased the MRC1 and MHC-II expression if a chicken macrophage cell line (HD11 cells) was used. In addition, LPS-induced NO production by HD11 cells was inhibited by CATH-2 and D-CATH-2. CONCLUSION: These results are a clear indication that CATH-2 (and D-CATH-2) affect the activation state of monocytes and macrophages, which leads to optimization of the innate immune response and enhancement of the adaptive immune response.

Cathelicidins PMAP-36, LL-37 and CATH-2 are similar peptides with different modes of action.

Host defense peptides (HDPs) play a pivotal role in innate immunity and have, in addition to antimicrobial activity, also important immunomodulatory functions. Bacteria are less likely to develop resistance against HDPs because these peptides target and kill bacteria in multiple ways, as well as modulate the immune system. Therefore, HDPs, and derivatives thereof, are promising alternatives to traditional antibiotics. Hardly anything is known about the immunomodulatory functions of porcine cathelicidin PMAP-36. In this study, we aimed to determine both antibacterial and immunomodulatory activities of PMAP-36 comparing the properties of PMAP-36 analogs with two well-studied peptides, human LL-37 and chicken CATH-2. Transmission electron microscopy revealed different killing mechanisms of E. coli for PMAP-36, CATH-2 and LL-37. LL-37 binds LPS very weakly in contrast to PMAP-36, but it inhibits LPS activation of macrophages the strongest. The first 11 amino acids of the N-terminal side of PMAP-36 are dispensable for E. coli killing, LPS-neutralization and binding. Deletion of four additional amino acids resulted in a strong decrease in activity. The activity of full length PMAP-36 was not affected by monomerization, whereas the shorter analogs require dimerization for proper immunomodulatory activity but not for their antibacterial activity.

Forecasting dryland vegetation condition months in advance through satellite data assimilation.

Dryland ecosystems are characterised by rainfall variability and strong vegetation response to changes in water availability over a range of timescales. Forecasting dryland vegetation condition can be of great value in planning agricultural decisions, drought relief, land management and fire preparedness. At monthly to seasonal time scales, knowledge of water stored in the system contributes more to predictability than knowledge of the climate system state. However, realising forecast skill requires knowledge of the vertical distribution of moisture below the surface and the capacity of the vegetation to access this moisture. Here, we demonstrate that contrasting satellite observations of water presence over different vertical domains can be assimilated into an eco-hydrological model and combined with vegetation observations to infer an apparent vegetation-accessible water storage (hereafter called accessible storage). Provided this variable is considered explicitly, skilful forecasts of vegetation condition are achievable several months in advance for most of the world's drylands.

Cathelicidins: Immunomodulatory Antimicrobials.

Cathelicidins are host defense peptides with antimicrobial and immunomodulatory functions. These effector molecules of the innate immune system of many vertebrates are diverse in their amino acid sequence but share physicochemical characteristics like positive charge and amphipathicity. Besides being antimicrobial, cathelicidins have a wide variety in immunomodulatory functions, both boosting and inhibiting inflammation, directing chemotaxis, and effecting cell differentiation, primarily towards type 1 immune responses. In this review, we will examine the biology and various functions of cathelicidins, focusing on putting in vitro results in the context of in vivo situations. The pro-inflammatory and anti-inflammatory functions are highlighted, as well both direct and indirect effects on chemotaxis and cell differentiation. Additionally, we will discuss the potential and limitations of using cathelicidins as immunomodulatory or antimicrobial drugs.

The potential for immunoglobulins and host defense peptides (HDPs) to reduce the use of antibiotics in animal production.

Innate defense mechanisms are aimed at quickly containing and removing infectious microorganisms and involve local stromal and immune cell activation, neutrophil recruitment and activation and the induction of host defense peptides (defensins and cathelicidins), acute phase proteins and complement activation. As an alternative to antibiotics, innate immune mechanisms are highly relevant as they offer rapid general ways to, at least partially, protect against infections and enable the build-up of a sufficient adaptive immune response. This review describes two classes of promising alternatives to antibiotics based on components of the innate host defense. First we describe immunoglobulins applied to mimic the way in which they work in the newborn as locally acting broadly active defense molecules enforcing innate immunity barriers. Secondly, the potential of host defense peptides with different modes of action, used directly, induced in situ or used as vaccine adjuvants is described.

Immunomodulation and effects on microbiota after in ovo administration of chicken cathelicidin-2.

Host Defense Peptides (HDPs) such as cathelicidins are multifunctional effectors of the innate immune system with both antimicrobial and pleiotropic immunomodulatory functions. Chicken cathelicidin-2 (CATH-2) has multiple immunomodulatory effects in vitro and the D-amino acid analog of this peptide has been shown to partially protect young chicks from a bacterial infection. However, the mechanisms responsible for CATH-2 mediated in vivo protection have not been investigated so far. In this study, D-CATH-2 was administered in ovo and the immune status and microbiota of the chicks were investigated at 7 days posthatch to elucidate the in vivo mechanisms of the peptide. In three consecutive studies, no effects on numbers and functions of immune cells were found and only small changes were seen in gene expression of Peripheral Blood Mononuclear Cells (PBMCs). In two studies, intestinal microbiota composition was determined which was highly variable, suggesting that it was strongly influenced by environmental factors. In both studies, in ovo D-CATH-2 treatment caused significant reduction of Ruminococcaceae and Butyricicoccus in the cecum and Escherichia/Shigella in both ileum and cecum. In conclusion, this study shows that, in the absence of an infectious stimulus, in ovo administration of a CATH-2 analog alters the microbiota composition but does not affect the chicks' immune system posthatch.

A new and efficient culture method for porcine bone marrow-derived M1- and M2-polarized macrophages.

BACKGROUND: Macrophages play an important role in the innate immune system as part of the mononuclear phagocyte system (MPS). They have a pro-inflammatory signature (M1-polarized macrophages) or anti-inflammatory signature (M2-polarized macrophages) based on expression of surface receptors and secretion of cytokines. However, very little is known about the culture of macrophages from pigs and more specific about the M1 and M2 polarization in vitro. METHODS: Porcine monocytes or mononuclear bone marrow cells were used to culture M1- and M2-polarized macrophages in the presence of GM-CSF and M-CSF, respectively. Surface receptor expression was measured with flow cytometry and ELISA was used to quantify cytokine secretion in response to LPS and PAM3CSK4 stimulation. Human monocyte-derived macrophages were used as control. RESULTS: Porcine M1- and M2-polarized macrophages were cultured best using porcine GM-CSF and murine M-CSF, respectively. Cultures from bone marrow cells resulted in a higher yield M1- and M2-polarized macrophages which were better comparable to human monocyte-derived macrophages than cultures from porcine monocytes. Porcine M1-polarized macrophages displayed the characteristic fried egg shape morphology, lower CD163 expression and low IL-10 production. Porcine M2-polarized macrophages contained the spindle-like morphology, higher CD163 expression and high IL-10 production. CONCLUSION: Porcine M1- and M2-polarized macrophages can be most efficiently cultured from mononuclear bone marrow cells using porcine GM-CSF and murine M-CSF. The new culture method facilitates more refined studies of porcine macrophages in vitro, important for both porcine and human health since pigs are increasingly used as model for translational research.

Land use change and El Niño-Southern Oscillation drive decadal carbon balance shifts in Southeast Asia.

An integrated understanding of the biogeochemical consequences of climate extremes and land use changes is needed to constrain land-surface feedbacks to atmospheric CO2 from associated climate change. Past assessments of the global carbon balance have shown particularly high uncertainty in Southeast Asia. Here, we use a combination of model ensembles to show that intensified land use change made Southeast Asia a strong source of CO2 from the 1980s to 1990s, whereas the region was close to carbon neutral in the 2000s due to an enhanced CO2 fertilization effect and absence of moderate-to-strong El Niño events. Our findings suggest that despite ongoing deforestation, CO2 emissions were substantially decreased during the 2000s, largely owing to milder climate that restores photosynthetic capacity and suppresses peat and deforestation fire emissions. The occurrence of strong El Niño events after 2009 suggests that the region has returned to conditions of increased vulnerability of carbon stocks.