Characterization of organic fouling on thermal bubble-driven micro-pumps.

Thermal bubble-driven micro-pumps are an upcoming micro-actuator technology that can be directly integrated into micro/mesofluidic channels, have no moving parts, and leverage existing mass production fabrication approaches. These micro-pumps consist of a high-power micro-resistor that boils fluid in microseconds to create a high-pressure vapor bubble which performs mechanical work. As such, these micro-pumps hold great promise for micro/mesofluidic systems such as lab-on-a-chip technologies. However, to date, no current work has studied the interaction of these micro-pumps with biofluids such as blood and protein-rich fluids. In this study, the effects of organic fouling due to egg albumin and bovine whole blood are characterized using stroboscopic high-speed imaging and a custom deep learning neural network based on transfer learning of RESNET-18. It was found that the growth of a fouling film inhibited vapor bubble formation. A new metric to quantify the extent of fouling was proposed using the decrease in vapor bubble area as a function of the number of micro-pump firing events. Fouling due to egg albumin and bovine whole blood was found to significantly degrade pump performance as well as the lifetime of thermal bubble-driven micro-pumps to less than 104 firings, which may necessitate the use of protective thin film coatings to prevent the buildup of a fouling layer.

Chromosomal instability can favor macrophage-mediated immune response and induce a broad, vaccination-like anti-tumor IgG response.

Chromosomal instability (CIN), a state in which cells undergo mitotic aberrations that generate chromosome copy number variations, generates aneuploidy and is thought to drive cancer evolution. Although associated with poor prognosis and reduced immune response, CIN generates aneuploidy-induced stresses that could be exploited for immunotherapies. In such contexts, macrophages and the CD47-SIRPα checkpoint are understudied. Here, CIN is induced pharmacologically induced in poorly immunogenic B16F10 mouse melanoma cells, generating persistent micronuclei and diverse aneuploidy while skewing macrophages towards an anti-cancer M1-like phenotype, based on RNA-sequencing profiling, surface marker expression and short-term antitumor studies. These results further translate to in vivo efficacy: Mice bearing CIN-afflicted tumors with wild-type CD47 levels survive only slightly longer relative to chromosomally stable controls, but long-term survival is maximized when combining macrophage-stimulating anti-tumor IgG opsonization and some form of disruption of the CD47-SIRPα checkpoint. Survivors make multi-epitope, de novo anti-cancer IgG that promote macrophage-mediated phagocytosis of CD47 knockout B16F10 cells and suppress tumoroids in vitro and growth of tumors in vivo . CIN does not greatly affect the level of the IgG response compared to previous studies but does significantly increase survival. These results highlight an unexpected therapeutic benefit from CIN when paired with maximal macrophage anti-cancer activity: an anti-cancer vaccination-like antibody response that can lead to more durable cures and further potentiate cell-mediated acquired immunity.

Genetic heterogeneity in p53-null leukemia increases transiently with spindle assembly checkpoint inhibition and is not rescued by p53.

Chromosome gains or losses often lead to copy number variations (CNV) and loss of heterozygosity (LOH). Both quantities are low in hematologic "liquid" cancers versus solid tumors in data of The Cancer Genome Atlas (TCGA) that also shows the fraction of a genome affected by LOH is ~ one-half of that with CNV. Suspension cultures of p53-null THP-1 leukemia-derived cells conform to these trends, despite novel evidence here of genetic heterogeneity and transiently elevated CNV after perturbation. Single-cell DNAseq indeed reveals at least 8 distinct THP-1 aneuploid clones with further intra-clonal variation, suggesting ongoing genetic evolution. Importantly, acute inhibition of the mitotic spindle assembly checkpoint (SAC) produces CNV levels that are typical of high-CNV solid tumors, with subsequent cell death and down-selection to novel CNV. Pan-cancer analyses show p53 inactivation associates with aneuploidy, but leukemias exhibit a weaker trend even though p53 inactivation correlates with poor survival. Overexpression of p53 in THP-1 does not rescue established aneuploidy or LOH but slightly increases cell death under oxidative or confinement stress, and triggers p21, a key p53 target, but without affecting net growth. Our results suggest that factors other than p53 exert stronger pressures against aneuploidy in liquid cancers, and identifying such CNV suppressors could be useful across liquid and solid tumor types.

Optimizing environmental viral surveillance: bovine serum albumin increases RT-qPCR sensitivity for high pathogenicity avian influenza H5Nx virus detection from dust samples.

IMPORTANCE: With the circulation of high pathogenicity avian influenza viruses having intensified considerably in recent years, the European Union is considering the vaccination of farmed birds. A prerequisite for this vaccination is the implementation of drastic surveillance protocols. Environmental sampling is a relevant alternative to animal sampling. However, environmental samples often contain inhibitory compounds in large enough quantities to inhibit RT-qPCR reactions. As bovine serum albumin is a molecule used in many fields to overcome this inhibitory effect, we tested its use on dust samples from poultry farms in areas heavily affected by HPAIV epizootics. Our results show that its use significantly increases the sensitivity of the method.

Differences in cell shape, motility, and growth reflect chromosomal number variations that can be visualized with live-cell ChReporters.

Chromosome numbers often change dynamically in tumors and cultured cells, which complicates therapy as well as understanding genotype-mechanotype relationships. Here we use a live-cell "ChReporter" method to identify cells with a single chromosomal loss in efforts to better understand differences in cell shape, motility, and growth. We focus on a standard cancer line and first show clonal populations that retain the ChReporter exhibit large differences in cell and nuclear morphology as well as motility. Phenotype metrics follow simple rules, including migratory persistence scaling with speed, and cytoskeletal differences are evident from drug responses, imaging, and single-cell RNA sequencing. However, mechanotype-genotype relationships between fluorescent ChReporter-positive clones proved complex and motivated comparisons of clones that differ only in loss or retention of a Chromosome-5 ChReporter. When lost, fluorescence-null cells show low expression of Chromosome-5 genes, including a key tumor suppressor APC that regulates microtubules and proliferation. Colonies are compact, nuclei are rounded, and cells proliferate more, with drug results implicating APC, and patient survival data indicating an association in multiple tumor-types. Visual identification of genotype with ChReporters can thus help clarify mechanotype and mechano-evolution.

Mathematical modeling at the livestock-wildlife interface: scoping review of drivers of disease transmission between species.

Modeling of infectious diseases at the livestock-wildlife interface is a unique subset of mathematical modeling with many innate challenges. To ascertain the characteristics of the models used in these scenarios, a scoping review of the scientific literature was conducted. Fifty-six studies qualified for inclusion. Only 14 diseases at this interface have benefited from the utility of mathematical modeling, despite a far greater number of shared diseases. The most represented species combinations were cattle and badgers (for bovine tuberculosis, 14), and pigs and wild boar [for African (8) and classical (3) swine fever, and foot-and-mouth and disease (1)]. Assessing control strategies was the overwhelming primary research objective (27), with most studies examining control strategies applied to wildlife hosts and the effect on domestic hosts (10) or both wild and domestic hosts (5). In spatially-explicit models, while livestock species can often be represented through explicit and identifiable location data (such as farm, herd, or pasture locations), wildlife locations are often inferred using habitat suitability as a proxy. Though there are innate assumptions that may not be fully accurate when using habitat suitability to represent wildlife presence, especially for wildlife the parsimony principle plays a large role in modeling diseases at this interface, where parameters are difficult to document or require a high level of data for inference. Explaining observed transmission dynamics was another common model objective, though the relative contribution of involved species to epizootic propagation was only ascertained in a few models. More direct evidence of disease spill-over, as can be obtained through genomic approaches based on pathogen sequences, could be a useful complement to further inform such modeling. As computational and programmatic capabilities advance, the resolution of the models and data used in these models will likely be able to increase as well, with a potential goal being the linking of modern complex ecological models with the depth of dynamics responsible for pathogen transmission. Controlling diseases at this interface is a critical step toward improving both livestock and wildlife health, and mechanistic models are becoming increasingly used to explore the strategies needed to confront these diseases.

Emotion regulation during encoding reduces negative and enhances neutral mnemonic discrimination in individuals with depressive symptoms.

Individuals with depression exhibit dysfunctional emotion regulation, general episodic memory deficits, and a negativity bias, where negative experiences are better remembered. Recent work suggests that the negativity bias in depression may be driven by enhanced mnemonic discrimination, a memory measure that relies on hippocampal pattern separation - a computation that processes experiences with overlapping features as unique. Previously, we found that individuals with depressive symptoms show enhanced negative and impaired neutral mnemonic discrimination. The current study aimed to investigate emotion regulation as an approach toward modifying memory encoding of negative and neutral events in individuals with depressive symptoms. Here we show that applying psychological distancing (a cognitive reappraisal strategy characterized by taking a third-person perspective toward negative events) during encoding was associated with reduced negative and enhanced neutral mnemonic discrimination during retrieval in individuals with depressive symptoms. These results suggest that applying emotion regulation techniques during encoding may provide an effective approach toward altering dysfunctional memory in those with depressive symptoms. Given that pharmacological treatments often fail to treat depression, emotion regulation provides a powerful and practical approach toward modifying cognitive and emotional processes. Future neuroimaging studies will be important to determine how emotion regulation impacts the neural mechanisms underlying these findings.

Structural insights into functional properties of the oxidized form of cytochrome c oxidase.

Cytochrome c oxidase (CcO) is an essential enzyme in mitochondrial and bacterial respiration. It catalyzes the four-electron reduction of molecular oxygen to water and harnesses the chemical energy to translocate four protons across biological membranes. The turnover of the CcO reaction involves an oxidative phase, in which the reduced enzyme (R) is oxidized to the metastable OH state, and a reductive phase, in which OH is reduced back to the R state. During each phase, two protons are translocated across the membrane. However, if OH is allowed to relax to the resting oxidized state (O), a redox equivalent to OH, its subsequent reduction to R is incapable of driving proton translocation. Here, with resonance Raman spectroscopy and serial femtosecond X-ray crystallography (SFX), we show that the heme a3 iron and CuB in the active site of the O state, like those in the OH state, are coordinated by a hydroxide ion and a water molecule, respectively. However, Y244, critical for the oxygen reduction chemistry, is in the neutral protonated form, which distinguishes O from OH, where Y244 is in the deprotonated tyrosinate form. These structural characteristics of O provide insights into the proton translocation mechanism of CcO.

Titrating CD47 by mismatch CRISPR-interference reveals incomplete repression can eliminate IgG-opsonized tumors but limits induction of antitumor IgG.

Phagocytic elimination of solid tumors by innate immune cells seems attractive for immunotherapy, particularly because of the possibilities for acquired immunity. However, the approach remains challenging, with blockade of the macrophage checkpoint CD47 working in immunodeficient mice and against highly immunogenic tumors but not in the clinic where tumors are poorly immunogenic. Even when mouse tumors of poorly immunogenic B16F10 melanoma are opsonized to drive engulfment with a suitable monoclonal antibody (mAb), anti-CD47 blockade remains insufficient. Using both in vitro immuno-tumoroids and in vivo mouse models, we show with CRISPR interference (CRISPRi) that a relatively uniform minimum repression of CD47 by 80% is needed for phagocytosis to dominate net growth when combined with an otherwise ineffective mAb (anti-Tyrp1). Heterogeneity enriches for CD47-high cells, but mice that eliminate tumors generate prophagocytic IgGs that increase in titer with CD47 repression and with tumor accumulation of macrophages, although deeper repression does not improve survival. Given well-known limitations of antibody permeation into solid tumors, our studies clarify benchmarks for CD47 disruption that should be more clinically feasible and safer but just as effective as complete ablation. Additionally, safe but ineffective opsonization in human melanoma trials suggests that combinations with deep repression of CD47 could prove effective and initiate durable immunity.

Confinement plus myosin-II suppression maximizes heritable loss of chromosomes, as revealed by live-cell ChReporters.

The mechanical environment of a cell can have many effects, but whether it impacts the DNA sequence of a cell has remained unexamined. To investigate this, we developed a live-cell method to measure changes in chromosome numbers. We edited constitutive genes with GFP or RFP tags on single alleles and discovered that cells that lose Chromosome reporters (ChReporters) become non-fluorescent. We applied our new tools to confined mitosis and to inhibition of the putative tumor suppressor myosin-II. We quantified compression of mitotic chromatin in vivo and demonstrated that similar compression in vitro resulted in cell death, but also rare and heritable ChReptorter loss. Myosin-II suppression rescued lethal multipolar divisions and maximized ChReporter loss during three-dimensional (3D) compression and two-dimensional (2D) lateral confinement, but not in standard 2D culture. ChReporter loss was associated with chromosome mis-segregation, rather than just the number of divisions, and loss in vitro and in mice was selected against in subsequent 2D cultures. Inhibition of the spindle assembly checkpoint (SAC) caused ChReporter loss in 2D culture, as expected, but not during 3D compression, suggesting a SAC perturbation. Thus, ChReporters enable diverse studies of viable genetic changes, and show that confinement and myosin-II affect DNA sequence and mechano-evolution.

Cooperative phagocytosis of solid tumours by macrophages triggers durable anti-tumour responses.

In solid tumours, the abundance of macrophages is typically associated with a poor prognosis. However, macrophage clusters in tumour-cell nests have been associated with survival in some tumour types. Here, by using tumour organoids comprising macrophages and cancer cells opsonized via a monoclonal antibody, we show that highly ordered clusters of macrophages cooperatively phagocytose cancer cells to suppress tumour growth. In mice with poorly immunogenic tumours, the systemic delivery of macrophages with signal-regulatory protein alpha (SIRPα) genetically knocked out or else with blockade of the CD47-SIRPα macrophage checkpoint was combined with the monoclonal antibody and subsequently triggered the production of endogenous tumour-opsonizing immunoglobulin G, substantially increased the survival of the animals and helped confer durable protection from tumour re-challenge and metastasis. Maximizing phagocytic potency by increasing macrophage numbers, by tumour-cell opsonization and by disrupting the phagocytic checkpoint CD47-SIRPα may lead to durable anti-tumour responses in solid cancers.

Structural basis for functional properties of cytochrome c oxidase.

Cytochrome c oxidase (CcO) is an essential enzyme in mitochondrial and bacterial respiration. It catalyzes the four-electron reduction of molecular oxygen to water and harnesses the chemical energy to translocate four protons across biological membranes, thereby establishing the proton gradient required for ATP synthesis1. The full turnover of the CcO reaction involves an oxidative phase, in which the reduced enzyme (R) is oxidized by molecular oxygen to the metastable oxidized OH state, and a reductive phase, in which OH is reduced back to the R state. During each of the two phases, two protons are translocated across the membranes2. However, if OH is allowed to relax to the resting oxidized state (O), a redox equivalent to OH, its subsequent reduction to R is incapable of driving proton translocation2,3. How the O state structurally differs from OH remains an enigma in modern bioenergetics. Here, with resonance Raman spectroscopy and serial femtosecond X-ray crystallography (SFX)4, we show that the heme a3 iron and CuB in the active site of the O state, like those in the OH state5,6, are coordinated by a hydroxide ion and a water molecule, respectively. However, Y244, a residue covalently linked to one of the three CuB ligands and critical for the oxygen reduction chemistry, is in the neutral protonated form, which distinguishes O from OH, where Y244 is in the deprotonated tyrosinate form. These structural characteristics of O provide new insights into the proton translocation mechanism of CcO.

Room-temperature structural studies of SARS-CoV-2 protein NendoU with an X-ray free-electron laser.

NendoU from SARS-CoV-2 is responsible for the virus's ability to evade the innate immune system by cleaving the polyuridine leader sequence of antisense viral RNA. Here we report the room-temperature structure of NendoU, solved by serial femtosecond crystallography at an X-ray free-electron laser to 2.6 Å resolution. The room-temperature structure provides insight into the flexibility, dynamics, and other intrinsic properties of NendoU, with indications that the enzyme functions as an allosteric switch. Functional studies examining cleavage specificity in solution and in crystals support the uridine-purine cleavage preference, and we demonstrate that enzyme activity is fully maintained in crystal form. Optimizing the purification of NendoU and identifying suitable crystallization conditions set the benchmark for future time-resolved serial femtosecond crystallography studies. This could advance the design of antivirals with higher efficacy in treating coronaviral infections, since drugs that block allosteric conformational changes are less prone to drug resistance.

Heterogeneity in the M. tuberculosis ß-Lactamase Inhibition by Sulbactam.

For decades, researchers have been determined to elucidate essential enzymatic functions on the atomic lengths scale by tracing atomic positions in real time. Our work builds on new possibilities unleashed by mix-and-inject serial crystallography (MISC) 1-5 at X-ray free electron laser facilities. In this approach, enzymatic reactions are triggered by mixing substrate or ligand solutions with enzyme microcrystals 6 . Here, we report in atomic detail and with millisecond time-resolution how the Mycobacterium tuberculosis enzyme BlaC is inhibited by sulbactam (SUB). Our results reveal ligand binding heterogeneity, ligand gating 7-9 , cooperativity, induced fit 10,11 and conformational selection 11-13 all from the same set of MISC data, detailing how SUB approaches the catalytic clefts and binds to the enzyme non-covalently before reacting to a trans- enamine. This was made possible in part by the application of the singular value decomposition 14 to the MISC data using a newly developed program that remains functional even if unit cell parameters change during the reaction.

Rapid Fabrication of Low-Cost Thermal Bubble-Driven Micro-Pumps.

Thermal bubble-driven micro-pumps are an upcoming actuation technology that can be directly integrated into micro/mesofluidic channels to displace fluid without any moving parts. These pumps consist of high power micro-resistors, which we term thermal micro-pump (TMP) resistors, that locally boil fluid at the resistor surface in microseconds creating a vapor bubble to perform mechanical work. Conventional fabrication approaches of thermal bubble-driven micro-pumps and associated microfluidics have utilized semiconductor micro-fabrication techniques requiring expensive tooling with long turn around times on the order of weeks to months. In this study, we present a low-cost approach to rapidly fabricate and test thermal bubble-driven micro-pumps with associated microfluidics utilizing commercial substrates (indium tin oxide, ITO, and fluorine doped tin oxide, FTO, coated glass) and tooling (laser cutter). The presented fabrication approach greatly reduces the turn around time from weeks/months for conventional micro-fabrication to a matter of hours/days allowing acceleration of thermal bubble-driven micro-pump research and development (R&D) learning cycles.

Effects of habitat fragmentation and hunting activities on African swine fever dynamics among wild boar populations.

African Swine Fever (ASF) has been slowly but steadily increasing its endemic range throughout Europe, posing an imminent risk to the pig industry. ASF transmission among wild boar occurs mainly through wild boar population movements, hence wild boar presence and density are important risk factors for introducing, maintaining, and spreading the disease. The understanding of wild boar population dynamics and their role in ASF transmission and persistence remains limited. It is crucial to gain knowledge in this area to improve wildlife management while minimizing the risks for ASF introduction and spread. We adapted an individual-based spatio-temporal stochastic model developed by Halasa et al. (2019) and tailored it to two regions in France. The model assessed yearly hunting activity, the carcass persistence seasonality, and the specific landscape characteristics of the Franco-Belgian border region and the Pyrénées-Atlantiques department. Following the establishment of local population dynamics through preliminary runs of the model, the model was run 100 iterations over 8 years in the two study areas where ASF was randomly seeded after the 2nd year of simulation. For each scenario, the model was initiated with 500 wild boar groups randomly spread across the study areas. Hunting activities were included and excluded to assess the impact on population growth and ASF spread. Results showed an ever-growing wild boar population for all scenarios, which was balanced when hunting activities were included. When introducing ASF, the wild boar populations were dramatically impacted in both areas with a decrease of 63 % of the population at the Franco-Belgian border and 86 % in the Pyrénées-Atlantiques department. Habitat fragmentation and landscape connectivity were highlighted as important factors shaping ASF propagation. The Franco-Belgian border, which had the most fragmented habitat with unsuitable areas for wild boars, was shown to limit wild boar movements, reducing the probability, and spread of ASF across the landscape. The lack of connectivity was reflected in a less effective transmission and lower number of infected groups (406 versus 467). In contrast, the epidemic duration was lengthened in the fragmented habitat compared to the homogenous area (2.6 years vs 1.6 years). This study provided information on defining and implementing control measures in case of an ASF incursion, since delimitation of the area via fences artificially induces landscape fragmentation, which is important for controlling ASF outbreaks.

Intersubunit Coupling Enables Fast CO2-Fixation by Reductive Carboxylases.

Enoyl-CoA carboxylases/reductases (ECRs) are some of the most efficient CO2-fixing enzymes described to date. However, the molecular mechanisms underlying the extraordinary catalytic activity of ECRs on the level of the protein assembly remain elusive. Here we used a combination of ambient-temperature X-ray free electron laser (XFEL) and cryogenic synchrotron experiments to study the structural organization of the ECR from Kitasatospora setae. The K. setae ECR is a homotetramer that differentiates into a pair of dimers of open- and closed-form subunits in the catalytically active state. Using molecular dynamics simulations and structure-based mutagenesis, we show that catalysis is synchronized in the K. setae ECR across the pair of dimers. This conformational coupling of catalytic domains is conferred by individual amino acids to achieve high CO2-fixation rates. Our results provide unprecedented insights into the dynamic organization and synchronized inter- and intrasubunit communications of this remarkably efficient CO2-fixing enzyme during catalysis.

Endogenous Retroviral Elements Generate Pathologic Neutrophils in Pulmonary Arterial Hypertension.

Rationale: The role of neutrophils and their extracellular vesicles (EVs) in the pathogenesis of pulmonary arterial hypertension is unclear. Objectives: To relate functional abnormalities in pulmonary arterial hypertension neutrophils and their EVs to mechanisms uncovered by proteomic and transcriptomic profiling. Methods: Production of elastase, release of extracellular traps, adhesion, and migration were assessed in neutrophils from patients with pulmonary arterial hypertension and control subjects. Proteomic analyses were applied to explain functional perturbations, and transcriptomic data were used to find underlying mechanisms. CD66b-specific neutrophil EVs were isolated from plasma of patients with pulmonary arterial hypertension, and we determined whether they produce pulmonary hypertension in mice. Measurements and Main Results: Neutrophils from patients with pulmonary arterial hypertension produce and release increased neutrophil elastase, associated with enhanced extracellular traps. They exhibit reduced migration and increased adhesion attributed to elevated ß1-integrin and vinculin identified by proteomic analysis and previously linked to an antiviral response. This was substantiated by a transcriptomic IFN signature that we related to an increase in human endogenous retrovirus K envelope protein. Transfection of human endogenous retrovirus K envelope in a neutrophil cell line (HL-60) increases neutrophil elastase and IFN genes, whereas vinculin is increased by human endogenous retrovirus K deoxyuridine triphosphate diphosphatase that is elevated in patient plasma. Neutrophil EVs from patient plasma contain increased neutrophil elastase and human endogenous retrovirus K envelope and induce pulmonary hypertension in mice, mitigated by elafin, an elastase inhibitor. Conclusions: Elevated human endogenous retroviral elements and elastase link a neutrophil innate immune response to pulmonary arterial hypertension.

CD47-SIRPα Checkpoint Disruption in Metastases Requires Tumor-Targeting Antibody for Molecular and Engineered Macrophage Therapies.

The macrophage checkpoint interaction CD47-SIRPα is an emerging target for cancer therapy, but clinical trials of monoclonal anti-CD47 show efficacy only in liquid tumors when combined with tumor-opsonizing IgG. Here, in challenging metastatic solid tumors, CD47 deletion shows no effect on tumor growth unless combined with otherwise ineffective tumor-opsonization, and we likewise show wild-type metastases are suppressed by SIRPα-blocked macrophages plus tumor-opsonization. Lung tumor nodules of syngeneic B16F10 melanoma cells with CD47 deletion show opsonization drives macrophage phagocytosis of B16F10s, consistent with growth versus phagocytosis calculus for exponential suppression of cancer. Wild-type CD47 levels on metastases in lungs of immunocompetent mice and on human metastases in livers of immunodeficient mice show that systemic injection of antibody-engineered macrophages also suppresses growth. Such in vivo functionality can be modulated by particle pre-loading of the macrophages. Thus, even though CD47-SIRPα disruption and tumor-opsonizing IgG are separately ineffective against established metastatic solid tumors, their combination in molecular and cellular therapies prolongs survival.

Modelling African swine fever virus spread in pigs using time-respective network data: Scientific support for decision makers.

African swine fever (ASF) represents the main threat to swine production, with heavy economic consequences for both farmers and the food industry. The spread of the virus that causes ASF through Europe raises the issues of identifying transmission routes and assessing their relative contributions in order to provide insights to stakeholders for adapted surveillance and control measures. A simulation model was developed to assess ASF spread over the commercial swine network in France. The model was designed from raw movement data and actual farm characteristics. A metapopulation approach was used, with transmission processes at the herd level potentially leading to external spread to epidemiologically connected herds. Three transmission routes were considered: local transmission (e.g. fomites, material exchange), movement of animals from infected to susceptible sites, and transit of trucks without physical animal exchange. Surveillance was represented by prevalence and mortality detection thresholds at herd level, which triggered control measures through movement ban for detected herds and epidemiologically related herds. The time from infection to detection varied between 8 and 21 days, depending on the detection criteria, but was also dependent on the types of herds in which the infection was introduced. Movement restrictions effectively reduced the transmission between herds, but local transmission was nevertheless observed in higher proportions highlighting the need of global awareness of all actors of the swine industry to mitigate the risk of local spread. Raw movement data were directly used to build a dynamic network on a realistic timescale. This approach allows for a rapid update of input data without any pre-treatment, which could be important in terms of responsiveness, should an introduction occur.