Hydrodynamic model for renal microvascular filtration: Effects of physiological and hemodynamic changes on glomerular size-selectivity.

OBJECTIVE: The first step in renal urine formation is ultrafiltration across the glomerular barrier. The change in its nanostructure has been associated with nephrotic syndromes. Effects of physiological and hemodynamic factor alterations associated with diabetic nephropathy (DN) on glomerular permselectivity are examined through a mathematical model employing low-Reynolds-number hydrodynamics and hindered transport theory. METHODS: Glomerular capillaries are represented as networks of cylindrical tubes with multilayered walls. Glomerular basement membrane (GBM) is a fibrous medium with bimodal fiber sizes. Epithelial slit fiber spacing follows a lognormal distribution based on reported electron micrographs with the highest resolution. Endothelial fenestrae are filled with fibers the size of glycosaminoglycans (GAGs). Effects of fiber-macromolecule steric and hydrodynamic interactions are included. Focusing on diabetic nephropathy, the physiological and hemodynamic factors employed in the computation are those reported for healthy humans and patients with early-but-overt diabetic nephropathy. The macromolecule concentration is obtained as a finite element solution of the convection-diffusion equation. RESULTS: Computed sieving coefficients averaged along the capillary length agree well with ficoll sieving coefficients from studies in humans for most solute radii. GBM thickening and the loss of the slit diaphragm hardly affect glomerular permselectivity. GAG volume fraction reduction in the endothelial fenestrae, however, significantly increases macromolecule filtration. Increased renal plasma flow rate (RPF), glomerular hypertension, and reduction of lumen osmotic pressure cause a slight sieving coefficient decrease. These effects are amplified by an increased macromolecule size. CONCLUSION: Our results indicate that glomerular hypertension and the reduction in the oncotic pressure decreases glomerular macromolecule filtration. Reduction of RPF and changes in the glomerular barrier structure associated with DN, however, increase the solute sieving. Damage to GAGs caused by hyperglycemia is likely to be the most prominent factor affecting glomerular size-selectivity.

Advances in rare cell isolation: an optimization and evaluation study.

BACKGROUND: Rare nucleated CD45 negative cells in peripheral blood may be malignant such as circulating tumor cells. Untouched isolation thereof by depletion of normal is favored yet still technological challenging. We optimized and evaluated a novel magnetic bead-based negative selection approach for enhanced enrichment of rare peripheral blood nucleated CD45 negative cells and investigated the problem of rare cell contamination during phlebotomy. METHODS: Firstly, the performance of the magnetic cell separation system was assessed using leukocytes and cultivated fibroblast cells in regard to depletion efficiency and the loss of cells of interest. Secondly, a negative selection assay was optimized for high performance, simplicity and cost efficiency. The negative selection assay consisted of; a RBC lysis step, two depletion cycles comprising direct magnetically labelling of leukocytes using anti-CD45 magnetic beads followed by magnetic capture of leukocytes using a duopole permanent magnet. Thirdly, assay evaluation was aligned to conditions of rare cell frequencies and comprised cell spike recovery, cell viability and proliferation, and CD45 negative cell detection. Additionally, the problem of CD45 negative cell contamination during phlebotomy was investigated. RESULTS: The depletion factor and recovery of the negative selection assay measured at most 1600-fold and 96%, respectively, leaving at best 1.5 × 104 leukocytes unseparated and took 35 min. The cell viability was negatively affected by chemical RBC lysis. Proliferation of 100 spiked ovarian cancer cells in culture measured 37% against a positive control. Healthy donor testing revealed findings of nucleated CD45 negative cells ranging from 1 to 22 cells /2.5 × 107 leukocytes or 3.5 mL whole blood in 89% (23/26) of the samples. CONCLUSION: Our assay facilitates high performance at shortest assay time. The enrichment assay itself causes minor harm to cells and allows proliferation. Our findings suggest that rare cell contamination is unavoidable. An unexpected high variety of CD45 negative cells have been detected. It is hypothesized that a rare cell profile may translate into tumor marker independent screening.

Transport of Spherical Particles Through Fibrous Media and a Row of Parallel Cylinders: Applications to Glomerular Filtration.

Viewed in renal physiology as a refined filtration device, the glomerulus filters large volumes of blood plasma while keeping proteins within blood circulation. Effects of macromolecule size and macromolecule hydrodynamic interaction with the nanostructure of the cellular layers of the glomerular capillary wall on the glomerular size selectivity are investigated through a mathematical simulation based on an ultrastructural model. The epithelial slit, a planar arrangement of fibers connecting the epithelial podocytes, is represented as a row of parallel cylinders with nonuniform spacing between adjacent fibers. The mean and standard deviation of gap half-width between its fibers are based on values recently reported from electron microscopy. The glomerular basement membrane (GBM) is represented as a fibrous medium containing fibers of two different sizes: the size of type IV collagens and that of glycosaminoglycans (GAGs). The endothelial cell layer is modeled as a layer full of fenestrae that are much larger than solute size and filled with GAGs. The calculated total sieving coefficient agrees well with the sieving coefficients of ficolls obtained from in vivo urinalysis in humans, whereas the computed glomerular hydraulic permeability also falls within the range estimated from human glomerular filtration rate (GFR). Our result indicates that the endothelial cell layer and GBM significantly contribute to solute and fluid restriction of the glomerular barrier, whereas, based on the structure of the epithelial slit obtained from electron microscopy, the contribution of the epithelial slit could be smaller than previously believed.

Kinetics of domain registration in multicomponent lipid bilayer membranes.

The kinetics of registration of lipid domains in the apposing leaflets of symmetric bilayer membranes is investigated via systematic dissipative particle dynamics simulations. The decay of the distance between the centres of mass of the domains in the apposing leaflets is almost linear during early stages, and then becomes exponential during late times. The time scales of both linear and exponential decays are found to increase with decreasing strength of interleaflet coupling. The ratio between the time scales of the exponential and linear regimes decreases with increasing domain size, implying that the decay of the distance between the domains' centres of mass is essentially linear for large domains. These numerical results are largely in agreement with the recent theoretical predictions of Han and Haataja [Soft Matter, 2013, 9, 2120-2124]. We also found that the domains become elongated during the registration process.

Modeling the dynamics of backyard chicken flows in traditional trade networks in Thailand: implications for surveillance and control of avian influenza.

In Southeast Asia, traditional poultry marketing chains have been threatened by epidemics caused by the highly pathogenic avian influenza H5N1 (HPAI H5N1) virus. In Thailand, the trade of live backyard chickens is based on the activities of traders buying chickens from villages and supplying urban markets with chicken meat. This study aims to quantify the flows of chickens traded during a 1-year period in a province of Thailand. A compartmental stochastic dynamic model was constructed to illustrate trade flows of live chickens from villages to slaughterhouses. Live poultry movements present important temporal variations with increased activities during the 15 days preceding the Chinese New Year and, to a lesser extent, other festivals (Qingming Festival, Thai New Year, Hungry Ghost Festival, and International New Year). The average distance of poultry movements ranges from 4 to 25 km, defining a spatial scale for the risk of avian influenza that spread through traditional poultry marketing chains. Some characteristics of traditional poultry networks in Thailand, such as overlapping chicken supply zones, may facilitate disease diffusion over longer distances through combined expansion and relocation processes. This information may be of use in tailoring avian influenza and other emerging infectious poultry disease surveillance and control programs provided that the cost-effectiveness of such scenarios is also evaluated in further studies.

New inflammatory indicators for cell-based liquid biopsy: association of the circulating CD44+/CD24- non-hematopoietic rare cell phenotype with breast cancer residual disease.

BACKGROUND: Breast cancer residual disease assessment in early-stage patients has been challenging and lacks routine identification of adjuvant therapy benefit and objective measure of therapy success. Liquid biopsy assays targeting tumor-derived entities are investigated for minimal residual disease detection, yet perform low in clinical sensitivity. We propose the detection of CD44-related systemic inflammation for the assessment of residual cancer. METHODS: Circulating CD44+/CD45- rare cells from healthy, noncancer- and cancer-afflicted donors were enriched by CD45 depletion and analyzed by immuno-fluorescence microscopy. CD44+ rare cell subtyping was based on cytological feature analysis and referred to as morphological index. AUC analysis was employed for identification of the most cancer-specific CD44+ subtype. RESULTS: The EpCam-/CD44+/CD24-/CD71-/CD45-/DNA+ phenotype alludes to a distinct cell type and was found frequently at concentrations below 5 cells per 5 mL in healthy donors. Marker elevation by at least 5 × on average was observed in all afflicted cohorts. The positive predicted value for the prediction of malignancy-associated systemic inflammation of a CD44+ rare cell subtype with a higher morphological index was 87%. An outlook for the frequency of sustained inflammation in residual cancer may be given to measure 78%. CONCLUSION: The CD44+ rare cell and subtype denotes improvement in detection of residual cancer disease and may provide an objective and alternative measure of disease burden in early-stage breast cancer.

Zinc Oxide Synthesis from Extreme Ratios of Zinc Acetate and Zinc Nitrate: Synergistic Morphology.

The synthesis of ZnO comprising different ratios of zinc acetate (ZA) and zinc nitrate (ZN) from the respective zinc precursor solutions was successfully completed via a simple precipitation method. Zinc oxide powders with different mole ratios of ZA/ZN were produced-80/1, 40/1, and 20/1. The crystallinity, microstructure, and optical properties of all produced ZnO powders were characterized using X-ray diffraction (XRD), scanning electron microscopy (SEM), and UV-Vis-NIR spectrophotometry. The average agglomerated particle sizes of ZnO-80/1, ZnO-40/1, and ZnO-20/1 were measured at 655, 640, and 620 nm, respectively, using dynamic light scattering (DLS). The optical properties of ZnO were significantly affected by the extreme ratio differences in the zinc precursors. ZnO-80/1 was found to have a unique coral-sheet structure morphology, which resulted in its superior ability to reflect near-infrared (NIR) radiation compared to ZnO-40/1 and ZnO-20/1. The NIR-shielding performances of ZnO were assessed using a thermal insulation test, where coating with ZnO-80/1 could lower the inner temperature by 5.2 °C compared with the neat glass substrate. Due to the synergistic effects on morphology, ZnO-80/1 exhibited the property of enhanced NIR shielding in curtailing the internal building temperature, which allows for its utilization as an NIR-reflective pigment coating in the construction of building envelopes.

Systemic cytology. A novel diagnostic approach for assessment of early systemic disease.

Recognition of low grade or asymptomatic systemic diseases suggests prevention of the worst, yet has been proven challenging ever since. Biomarker-based liquid biopsy has emerged in recent years as a practical platform for the assessment of systemic diseases yet, technical realizations were mainly focused on cancer, faced challenges in accuracy at early stage and are lacking provision of sufficient evidence of disease. In particular in cell-based cancer liquid biopsy, obstacles are rarity and heterogeneity of circulating tumor and tumor-associated rare cells. Evidence is mounting about an entire spectrum of distinct circulating rare cell types that denotes the systemic component of a certain physiological state. Therefore, circulating rare cells in combination may arise from yet, also account for systemic diseases, which we denote as multi-rare cell association and involves foremost bone marrow-derived progenitor and stem cells yet, also matured somatic cell types. One would expect immense diagnostic value in the read-out of the so called rare cell population which represents cytological evidence of abnormality. We hypothesize that comprehensive rare cell population profiling as contrasted to the biomarker screening approach may realize the premise of a biopsy as to confirm, characterize, grade, stage or predict a systemic disease. This novel approach represents the "missing link" in diagnostic care of in particular early or residual systemic disease and presumes a steady gain in knowledge about the clinical interpretation of rare cell population profiles thus, expecting the knowledge-driven transformation of cell-based liquid biopsy from suggestion to confirmation. We support our hypothesis by past findings made by others and us and provide insights how to interpret a certain rare cell population profile.

A comparison of deterministic and stochastic simulations of neuronal vesicle release models.

We study the calcium-induced vesicle release into the synaptic cleft using a deterministic algorithm and MCell, a Monte Carlo algorithm that tracks individual molecules. We compare the average vesicle release probability obtained using both algorithms and investigate the effect of the three main sources of noise: diffusion, sensor kinetics and fluctuations from the voltage-dependent calcium channels (VDCCs). We find that the stochastic opening kinetics of the VDCCs are the main contributors to differences in the release probability. Our results show that the deterministic calculations lead to reliable results, with an error of less than 20%, when the sensor is located at least 50 nm from the VDCCs, corresponding to microdomain signaling. For smaller distances, i.e. nanodomain signaling, the error becomes larger and a stochastic algorithm is necessary.

The Blood Circulating Rare Cell Population. What is it and What is it Good For?

Blood contains a diverse cell population of low concentration hematopoietic as well as non-hematopoietic cells. The majority of such rare cells may be bone marrow-derived progenitor and stem cells. This paucity of circulating rare cells, in particular in the peripheral circulation, has led many to believe that bone marrow as well as other organ-related cell egress into the circulation is a response to pathological conditions. Little is known about this, though an increasing body of literature can be found suggesting commonness of certain rare cell types in the peripheral blood under physiological conditions. Thus, the isolation and detection of circulating rare cells appears to be merely a technological problem. Knowledge about rare cell types that may circulate the blood stream will help to advance the field of cell-based liquid biopsy by supporting inter-platform comparability, making use of biological correct cutoffs and "mining" new biomarkers and combinations thereof in clinical diagnosis and therapy. Therefore, this review intends to lay ground for a comprehensive analysis of the peripheral blood rare cell population given the necessity to target a broader range of cell types for improved biomarker performance in cell-based liquid biopsy.

Leptospirosis research: fast, easy and reliable enumeration of mobile leptospires.

Leptospirosis caused by Leptospira interrogans is the most widespread zoonosis and a major public health problem worldwide. Based on light-scattering and absorption, quantification of leptospires using UV-VIS spectroscopy was used as an indirect counting technique by measuring the optical density and comparing this to automated direct counting using a counting chamber in combination with imaging and analyzing software. Two serovars, Bangkok and Copenhagenii, from log-phase growth were used for the establishment of standard curves. They were found to be linear and slightly different in gradient for each serovar. The ease, rapidity, and reliability of these two adapted and optimized counting techniques may provide a useful alternative enumeration technique for leptospirosis research.

Chemical force mapping of phosphate and carbon on acid-modified tapioca starch surface.

Surface chemical microstructure of hydrochloric acid hydrolyzed tapioca starch producing different amylose:amylopectin (Am:Ap) ratios were studied with scanning chemical force microscopy (CFM). The chemical force probes were functionalized of two types with -OH (phosphate specific) and -CH3 (carbon specific). Lateral force trace-minus-retrace (TMR) images from -OH and -CH3 probes revealed changes in the phosphate domains and the carbon backbone for the varying acid hydrolyzed tapioca starch compared to that of the native tapioca starch. Scanning electron micrographs (SEM) showed different degree of the granule surface disruption before and after hydrolysis. The exterior structures of the acid hydrolyzed starch granules were chemically investigated with CFM to study the relationships of the surface molecular structures and the Am:Ap ratios.

Mathematical modeling and application of genetic algorithm to parameter estimation in signal transduction: trafficking and promiscuous coupling of G-protein coupled receptors.

G-protein-coupled receptors (GPCRs) constitute a large and diverse family of proteins whose primary function is to transduce extracellular stimuli into intracellular signals. These receptors play a critical role in signal transduction, and are among the most important pharmacological drug targets. Upon binding of extracellular ligands, these receptor molecules couple to one or several subtypes of G-protein which reside at the intracellular side of the plasma membrane to trigger intracellular signaling events. The question of how GPCRs select and activate a single or multiple G-protein subtype(s) has been the topic of intense investigations. Evidence is also accumulating; however, that certain GPCRs can be internalized via lipid rafts and caveolae. In many cases, the mechanisms responsible for this still remain to be elucidated. In this work, we extend the mathematical model proposed by Chen et al. [Modelling of signalling via G-protein coupled receptors: pathway-dependent agonist potency and efficacy, Bull. Math. Biol. 65 (5) (2003) 933-958] to take into account internalization, recycling, degradation and synthesis of the receptors. In constructing the model, we assume that the receptors can exist in multiple conformational states allowing for a multiple effecter pathways. As data on kinetic reaction rates in the signalling processes measured in reliable in vivo and in vitro experiments is currently limited to a small number of known values. In this paper, we also apply a genetic algorithm (GA) to estimate the parameter values in our model.

An update of circulating rare cell types in healthy adult peripheral blood: findings of immature erythroid precursors.

BACKGROUND: Circulating rare cells (CRCs) are benign or malignant minuscule events in the peripheral blood or other bodily fluids. The detection and quantification of certain CRC types is an invaluable or proposed candidate biomarker for diagnosis, prognosis and prediction of various pathological conditions. The list of CRC types and biomarker applicability thereof continues to expand along with improvements in cell selection technology. Past findings may suggest commonness of healthy donor peripheral blood circulating mature erythroblasts. This work suggests the occurrence of morphologically distinct bone marrow native circulating early erythroid precursors that we intend to add to the list of CRCs. METHODS: We tested 15 healthy individuals that varied in age and gender employing a negative cell selection assay based on magnetic bead technology to characterize healthy adult circulating CD45 negative cell events using cell surface markers CD71 and glycophorin-A. RESULTS: Positive events were detected and varied in cell and nuclear size ranging between 7.5 µm till 15 µm and 4.5 till 9.2 µm, respectively with distinct appearance under bright field microscope. Cell rarity increased with cell and nuclear size. Largest cells exceeded 13.5 µm in cell diameter and were found in 7 out of 15 donors. CONCLUSIONS: Circulating erythroid precursors occur at different stages of maturation and may be part of the benign CRC spectrum.

Effect of lipid peroxidation on the properties of lipid bilayers: a molecular dynamics study.

Lipid peroxidation plays an important role in cell membrane damage. We investigated the effect of lipid peroxidation on the properties of 1-palmitoyl-2-linoleoyl-sn-glycero-3-phosphatidylcholine (PLPC) lipid bilayers using molecular dynamics simulations. We focused on four main oxidation products of linoleic acid with either a hydroperoxide or an aldehyde group: 9-trans, cis-hydroperoxide linoleic acid, 13-trans, cis-hydroperoxide linoleic acid, 9-oxo-nonanoic acid, and 12-oxo-9-dodecenoic acid. These oxidized chains replaced the sn-2 linoleate chain. The properties of PLPC lipid bilayers were characterized as a function of the concentration of oxidized lipids, with concentrations from 2.8% to 50% for each oxidation product. The introduction of oxidized functional groups in the lipid tail leads to an important conformational change in the lipids: the oxidized tails bend toward the water phase and the oxygen atoms form hydrogen bonds with water and the polar lipid headgroup. This conformational change leads to an increase in the average area per lipid and, correspondingly, to a decrease of the bilayer thickness and the deuterium order parameters for the lipid tails, especially evident at high concentrations of oxidized lipid. Water defects are observed in the bilayers more frequently as the concentration of the oxidized lipids is increased. The changes in the structural properties of the bilayer and the water permeability are associated with the tendency of the oxidized lipid tails to bend toward the water interface. Our results suggest that one mechanism of cell membrane damage is the increase in membrane permeability due to the presence of oxidized lipids.

The stochastic model of non-equilibrium mutagen-induced alterations of DNA: implication to genetic instability in cancer.

Genetic alterations such as point mutations, chromosomal rearrangements, modification of DNA methylation and chromosome aberrations accumulate during the lifetime of an organism. They can be caused by intrinsic errors in the DNA replication and repair as well as by external factors such as exposure to mutagenic substances or radiation. The main purpose of the present work is to begin an exploration of the stochastic nature of non-equilibrium DNA alteration caused by events such as tautomeric shifts. This is done by modeling the genetic DNA code chain as a sequence of DNA-bit values ('1' for normal bases and '-1' for abnormal bases). We observe the number of DNA-bit changes resulting from the random point mutation process which, in the model, is being induced by a stochastic Brownian mutagen (BM) as it diffuses through the DNA-bit systems. Using both an analytical and Monte Carlo (MC) simulation techniques, we observe the local and global number of DNA-bit changes. It is found that in 1D, the local DNA-bit density behaves like 1/t, the global total number of the switched (abnormal) DNA-bit increases as t. The probability distribution P(b, 0, t) of b(0, t) is log-normal. It is also found that when the number of mutagens is increased, the number of the total abnormal DNA-bits does not grow linearly with the number of mutagens. All analytic results are in good agreement with the simulation results.

Stochastic Models of Emerging Infectious Disease Transmission on Adaptive Random Networks.

We presented adaptive random network models to describe human behavioral change during epidemics and performed stochastic simulations of SIR (susceptible-infectious-recovered) epidemic models on adaptive random networks. The interplay between infectious disease dynamics and network adaptation dynamics was investigated in regard to the disease transmission and the cumulative number of infection cases. We found that the cumulative case was reduced and associated with an increasing network adaptation probability but was increased with an increasing disease transmission probability. It was found that the topological changes of the adaptive random networks were able to reduce the cumulative number of infections and also to delay the epidemic peak. Our results also suggest the existence of a critical value for the ratio of disease transmission and adaptation probabilities below which the epidemic cannot occur.

Mathematical modeling of diphtheria transmission in Thailand.

In this work, a mathematical model for describing diphtheria transmission in Thailand is proposed. Based on the course of diphtheria infection, the population is divided into 8 epidemiological classes, namely, susceptible, symptomatic infectious, asymptomatic infectious, carrier with full natural-acquired immunity, carrier with partial natural-acquired immunity, individual with full vaccine-induced immunity, and individual with partial vaccine-induced immunity. Parameter values in the model were either directly obtained from the literature, estimated from available data, or estimated by means of sensitivity analysis. Numerical solutions show that our model can correctly describe the decreasing trend of diphtheria cases in Thailand during the years 1977-2014. Furthermore, despite Thailand having high DTP vaccine coverage, our model predicts that there will be diphtheria outbreaks after the year 2014 due to waning immunity. Our model also suggests that providing booster doses to some susceptible individuals and those with partial immunity every 10 years is a potential way to inhibit future diphtheria outbreaks.

Investigation on predominant Leptospira serovars and its distribution in humans and livestock in Thailand, 2010-2015.

BACKGROUND: Leptospirosis is a worldwide zoonotic bacterial disease caused by infection with leptospires. Leptospirosis in humans and livestock is an endemic and epidemic disease in Thailand. Livestock may act as reservoirs for leptospires and source for human infection. METHODOLOGY/PRINCIPAL FINDINGS: Data on leptospirosis infection in humans and livestock (Buffaloes, Cattle, and Pigs) species during 2010 to 2015 were analyzed. Serum samples were examined using Microscopic Agglutination Test (MAT) to identify antibodies against Leptospira serovars using a cut-off titer ≥ 1:100. The seroprevalence was 23.7% in humans, 24.8% in buffaloes, 28.1% in cattle, and 11.3% in pigs. Region specific prevalence among humans and livestock was found in a wide range. The most predominant serovars were Shermani, followed by Bratislava, Panama, and Sejroe in human, Shermani, Ranarum, and Tarassovi in buffaloes, and Shermani and Ranarum in cattle and pigs. Equally highest MAT titers against multiple serovars per one sample were found mainly in buffaloes and cattle showing equally titers against Ranarum and Shermani. The correlations of distribution of serovars across Thailand's regions were found to be similar in pattern for cattle but not for buffaloes. In humans, the serovar distribution in the south differed from other regions. By logistic regression, the results indicated that livestock is more susceptible to infection by serovar Shermani when compared to humans. CONCLUSIONS/SIGNIFICANCE: This study gives a detailed picture of the predominance of Leptospira serovars in relation to region, humans and typical livestock. The broad spatial distribution of seroprevalence was analyzed across and within species as well as regions in Thailand. Our finding may guide public health policy makers to implement appropriate control measures and help to reduce the impact of leptospirosis in Thailand.

A one-year effective reproduction number of the 2014-2015 Ebola outbreaks in the widespread West African countries and quantitative evaluation of air travel restriction measure.

BACKGROUND: The 2014-2015 Ebola outbreak in West Africa is the largest and longest Ebola Virus Disease (EVD) outbreak in the history, and the virus has escaped across countries and continents via air travel in this outbreak. METHOD: The interpolated data from WHO Ebola situation reports were used to estimate number of weekly infectious individuals and daily effective reproduction numbers (Rt) in Guinea, Liberia and Sierra Leone. A stochastic dynamic model was performed to estimate the risk of EVD importation into the top 20 final destination countries of air travelers departing from within the three epidemic countries, and the effectiveness of air travel restriction was subsequently evaluated. RESULTS: The daily Rt was estimated at 0.72-1.32 in Guinea, 0.62-1.38 in Liberia and 0.81-1.38 in Sierra Leone. The peak of EVD importation probability was observed in early November 2014 and the restriction of air travel may mitigate the risk up to 67.7% (95% CI 66.6-68.7). CONCLUSIONS: Our results suggest that restriction of air travels is effective in reducing the risk of EVD importation but controlling of the virus at the original affected countries is vitally more important for preventing inter-terrestrial dissemination of EVD.