gattii strains for additional assay validation Culture collection ID Geographic origin Sample type MLST Year of isolation B4501 Australia Human VGI unknown B4503 Australia Human VGI unknown B4504 Australia Human VGI unknown B4516 Australia Human VGI unknown B5765 India Environmental VGI unknown B9018 California Human

VGI 2011 B9019 New Mexico Human VGI 2011 B9021 Rhode Island Human VGI 2011 B9142 Georgia Human PR-171 manufacturer VGI 2011 B9149 California Human VGI 2011 B8508 Oregon Human VGIIa 2009 B8512 Oregon Alpaca VGIIa 2009 B8558 Washington Human VGIIa 2010 B8561 Washington Human VGIIa 2010 B8563 Washington Human VGIIa 2010 B8567 Washington Dog VGIIa 2010 B8854 Washington Human VGIIa 2010 B8889 Oregon Environmental VGIIa 2010 B9077 Washington

Environmental VGIIa 2011 B9296 British Columbia Environmental VGIIa 2011 B8211 Oregon Selleckchem SB431542 Human VGIIb 2009 B8966 Oregon Horse VGIIb 2010 B9076 Washington Environmental VGIIb 2011 B9157 Washington Horse VGIIb 2011 B9170 Washington Porpoise VGIIb 2011 B9234 Washington Cat VGIIb 2011 B9290 British Columbia Cat VGIIb 2011 B9241 Oregon Human VGIIb 2011 B9428 Washington Cat VGIIb 2012 B9159 Washington Sheep VGIIc 2011 B9227

Oregon Cat VGIIc 2011 B9235 Oregon Human VGIIc 2011 B9244 Oregon Human VGIIc 2011 B9245 Oregon Human VGIIc 2011 B9295 British Columbia Environmental VGIIc 2011 B9302 Oregon Environmental VGIIc 2011 B9374 Oregon Human VGIIc 2011 B8965 New Mexico Human VGIII 2010 B9148 California Human VGIII 2011 B9151 Michigan Human VGIII 2011 B9163 New Mexico Human VGIII 2011 B9237 New Mexico Cat VGIII 2011 B9372 Cediranib (AZD2171) California Cow VGIII 2011 B9422 Oregon Cat VGIII 2012 B9430 Alaska Cat VGIII 2012 B7238 Botswana Human VGIV 2005 B7240 Botswana Human VGIV 2005 B7243 Botswana Human VGIV 2005 B7247 Botswana Human VGIV 2005 B7249 Botswana Human VGIV 2005 B7260 Botswana Human VGIV 2006 B7262 Botswana Human VGIV 2006 B7263 Botswana Human VGIV 2006 B7264 Botswana Human VGIV 2006 B7265 Botswana Human VGIV 2006 Isolate culturing and DNA extraction Isolates were grown on Yeast Peptone Glucose (YPD) agar plus 0.5% NaCl at 37°C for 24 hours; and DNA was prepared using an UltraClean DNA Isolation Kit as described by the manufacturer, with some modifications (MO BIO Laboratories, Carlsbad, CA). Briefly, ~0.

A single crossover between the regions of homology leads to a functional tetA gene. Plasmids pYA4463 and pYA4590 were constructed to test intraplasmid recombination (Figure 1 panel A). Plasmid pYA4463 carries two truncated tetA genes (5′ end and 3′end), which have this website 466-bp of tandemly repeated sequence. An intramolecular recombination event can delete one of the repeats resulting in an intact tetA gene, thereby recreating the structure of plasmid pACYC184 (Figure 1 panel A). Theoretically, intermolecular recombination may occur between two pYA4463 molecules to form a plasmid dimer with a functional tetA gene (Figure 1 panel C). Plasmid pYA4590 contains a 602-bp tetA sequence duplication separated by a

1041-bp kan cassette. The intramolecular recombination product is equivalent to pACYC184. The intermolecular recombination product is a dimer plasmid containing an intact tetA gene (Figure 1 panel C). Plasmids pYA4464 and pYA4465 carry the 3′tet gene and 5′tet gene, respectively (Figure 1). The Rec+ Salmonella strain χ3761 carrying either plasmid individually was sensitive to tetracycline. There is 751-bp of tetA DNA in common between the two truncated tetA genes. Recombination between the two plasmids creates a hybrid plasmid containing an intact BKM120 tetA gene (Figure 1 panel C). Intraplasmid recombination products To verify the recombination products, plasmid DNA was prepared

from tetracycline resistant (TcR) single colonies derived from χ3761(pYA4463), χ3761(pYA4590) and χ3761(pYA4464, pYA4465). Plasmids extracted from TcR clones of χ3761(pYA4463) were digested with XbaI and SalI. Theoretically, XbaI/SalI digestion of pYA4463 will yield two fragments (3524 bp and 1187 bp), pACYC184 will yield two fragments (3524 bp and 721 bp) and pYA4463 dimer will yield four fragments (3524 bp, 3524 bp, 1653 bp and 721 bp). The results (Figure 3A) showed that digestion of all 16 TcR clones yielded a 721-bp band, indicating either a pYA4463 dimer or a plasmid equivalent to cAMP pACYC184. Three clones (lane 1, 5 and 10) yielded the pYA4463 dimer-specific 1653-bp band. Therefore, we conclude that the other 13 clones recombined to form the pACYC184-like

structure. Of note, several clones (2, 13-16) also yielded the 1187-bp pYA4463-specific band, suggesting that the original plasmid (pYA4463) and its recombination product (pACYC184-like) could coexist in the same bacterial cell. Figure 3 Verification of plasmid recombination product by agarose gel separation. (A) Plasmid DNA was isolated from TcR clones derived from χ3761(pYA4463) and digested by XbaI and SalI. (B) Plasmid DNA was isolated from TcR clones of χ3761(pYA4590) and digested by KpnI and EcoRI. (C) Plasmid DNA was isolated from TcR or TcS clones of χ3761(pYA4464, pYA4465). The purified plasmids were digested with NcoI and BglII. Plasmids extracted from TcR clones of χ3761(pYA4590) were digested with KpnI and EcoRI.

albicans, and as a consequence, reduce biofilm formation. However, our results suggest that the compound in serum that inhibits C. albicans biofilm formation is not proteinaceous. Abraham et al.[15] found that a low molecular weight component of human serum inhibits biofilm formation in Staphylococcus aureus, and the component was protease-resistant and heat stable. We conclude here that human serum may also contain non-protein component(s) that can inhibit the adhesion and biofilm formation TSA HDAC chemical structure of fungi and bacteria. To confirm this hypothesis, future studies are needed to identify this component of human serum. In this study, planktonic growth of C. albicans was not inhibited by human serum,

indicating that inhibition of biofilm formation Selleckchem PXD101 was not due solely to growth inhibition. Biofilm formation of C. albicans, a process that depends upon both cell-cell and cell-substrate adherence, is controlled by a tightly woven network of genes [10]. Among this gene network, BCR1 is one of the best-characterized biofilm regulators [11–13, 29]. Through its adhesin targets ALS1, ALS3, HWP1 and ECE1, BCR1 mediates cell-substrate and cell-cell interactions in biofilms [30, 31]. In this study, at the adhesion stage of biofilm formation (60 min, 90 min), the

expression of BCR1 went from less than to significantly higher than that of the control group. This may be due to the promoting effect of serum on hypha growth, as BCR1 RNA accumulation depends on the hyphal developmental activator TEC1[32]. ALS1 and ALS3 are members of the agglutinin-like sequence (ALS) gene family that encodes cell-wall glycoproteins [33]. Most Als proteins have adhesin functions [34, 35]. Mutational analysis indicates

that strains lacking all functional ALS1 and ALS3 alleles (als1Δ/als1Δ als3Δ/als3Δ) failed to produce any detectable adherent cells in biofilm models both in vivo and in vitro[30], or in actual biofilm formation. The als1Δ/als1Δ mutants produced substantial biofilms, but the biofilms often sloughed Tenofovir mouse off the substrate, while the als3Δ/als3Δ mutant only produced scant, disorganized biofilms on catheter material in vitro[12]. Our data on transcript analysis showed that the expression of ALS1 and ALS3 were reduced at different time points in the biofilm adhesion stage. Therefore, we supposed that the anti-adhesion effect of human serum might occur via inhibition of the expression of ALS1 and ALS3, and therefore affect biofilm formation. Previous studies have shown that a bcr1Δ/ bcr1Δ mutant, which has reduced expression of ALS1, ALS3, and other adhesins, has defective biofilm formation in both an in vitro and in vivo catheter model [12]. In this study, at 90 min of growth, the change in the levels of BCR1 level was different from ALS1 and ALS3, indicating that ALS1 and ALS3 are also affected by other factors [8, 36]. Interestingly, human serum promotes the expression of HWP1 and ECE1. HWP1 is a well-characterized hypha-specific gene that can mediate C.

Bennett DE, Cafferkey MT: Multilocus restriction typing: A tool for Neisseria meningitidis strain discrimination. J Med Microbiol 2003, 52:781–787.PubMedCrossRef 29. Helgerson AF, Sharma V, Dow AM, Schroeder R, Post K, Cornick NA: Edema disease caused by a clone of Escherichia coli O147. J Clin Microbiol 2006, 44:3074–3077.PubMedCrossRef 30. Singh I, Virdi JS: Isolation biochemical characterization and in vitro tests of pathogenicity of Yersinia enterocolitica isolated mTOR inhibitor review from pork. Curr Sci 1999, 77:1019–1021. 31. Sinha I, Choudhary I, Virdi JS: Isolation of Yersinia enterocolitica and Yersinia intermedia from wastewaters and their biochemical and serological characteristics. Curr Sci 2000, 79:510–513.

32. Singh I, Bhatnagar S, Virdi JS: Isolation and characterization of Yersinia enterocolitica from diarrheic human subjects and other sources. Curr Sci 2003, 84:1353–1355. 33. Nei M: Estimation of average heterozygosity and genetic distance from a small sample of individuals.

Genetics 1978, 89:583–590.PubMed 34. Brown AH, Feldman MW, Nevo E: Multilocus structure check details of natural populations of Hordeum spontaneum . Genetics 1980, 96:523–536.PubMed 35. Maynard Smith J, Smith NH, O’Rourke M, Spratt BG: How clonal are bacteria? Proc Nat Acad Sci USA 1993, 90:4384–4388.CrossRef 36. Souza V, Nguyen TT, Hudson RR, Piñero D, Lenski RE: Hierarchical analysis of linkage disequilibrium in Rhizobium populations: Evidence for sex? Proc Natl Acad Sci USA 1992, 89:8389–8393.PubMedCrossRef 37. Haubold H, Hudson RR: LIAN 3.0: detecting linkage disequilibrium in multilocus data. Bioinformatics 2000, 16:847–848.PubMedCrossRef 38. Hunter PR, Gaston MA: Numerical index of the discriminatory ability of typing systems. An application of Simpson’s index of diversity. J Clin Microbiol 1988, 26:2465–2466.PubMed 39. Fearnley C, On SLW, Kokotovic B, Manning G, Cheasty T, Newell DG:

Application of fluorescent amplified fragment length polymorphism for comparison of human and animal isolates of Yersinia enterocolitica . Appl Environ Microbiol 2005, 71:4960–4965.PubMedCrossRef 40. Tauxe RV, Vandepitte J, Wauters G, Martin SM, Goossens V, DeMol P, Van Noyen R, Thiers G: Yersinia enterocolitica infections and pork: the missing link. Lancet 1987, 1:1129–1132.PubMedCrossRef 41. Muller-Graf CDM, Whatmore AM, King SJ, Trzcinski K, Pickerill AP, Doherty N, Paul J, Griffiths Selleck Rapamycin D, Crook D, Dowson CG: Population biology of Streptococcus pneumoniae isolated from oropharyngeal carriage and invasive disease. Microbiology 1999, 145:3283–3293.PubMed 42. Dyet KH, Simmonds RS, Martin DR: Multilocus restriction typing method to predict the sequence type of meningococci. J Clin Microbiol 2004, 42:1742–1745.PubMedCrossRef 43. Coenye T, Spilker T, Martin A, LiPuma JJ: Comparative assessment of genotyping methods for epidemiologic study of Burkholderia cepacia genomovar III. J Clin Microbiol 2002, 40:3300–3307.PubMedCrossRef 44.

Aquat Microb Eco 2008, 52:69–82.CrossRef 14. Chen M, Chen F, Zhao B, Wu QL, Kong FX: Genetic diversity of eukaryotic microorganisms in Lake Taihu, a large shallow subtropical lake in China. Microb Ecol 2008,56(3):572–583.PubMedCrossRef 15. Masquelier S, Foulon E, Jouenne F, Ferréol M, Brussard CPD, Vaulot D: Distribution of eukaryotic in the English Channel and North Sea in summer. J Sea Res 2011, 66:111–122.CrossRef 16. Petchey OL, McPhearson PT,

Casey TM, Morin PJ: Environmental warming alters food-web structure and ecosystem function. Nature 1999, 402:69–72.CrossRef 17. Mostajir B, Sime-Ngando T, Demers S, Belzile C, et al.: Ecological implications of changes in cell size and photosynthetic capacity of marine Prymnesiophyceae

induced by ultraviolet-B radiation. Mar Ecol Prog Ser 1999, 187:89–100.CrossRef 18. Sommaruga R, Hofer www.selleckchem.com/products/mek162.html JS, Alonso-Saez L, Gasol JM: Differential Sunlight Sensitivity of Picophytoplankton from Surface Mediterranean Coastal Waters. Appl Environ Microb 2005,71(4):2154–2157.CrossRef 19. Ferreyra GA, Mostajir B, Schloss IR, Chatila K, Ferrario ME, Sargian P, Roy S, Prod’homme J, Demers S: Ultraviolet-B radiation effects on the structure and function of lower trophic levels of the marine planktonic food web. Photochem Photobiol 2006,82(4):887–897.PubMedCrossRef 20. Conan P, Joux F, Torréton JP, Pujo-Pay M, Rochelle-Newall E, Mari X: Impact of solar ultraviolet radiation on bacterio- and phytoplankton activity in a large coral reef lagoon (SW New Caledonia). Aquat Microb Ecol 2008, 52:83–98.CrossRef 21. Christensen MR, Graham MD, Vinebrooke RD, Findlay DL, Paterson MJ, Turner MA: Multiple 4EGI-1 solubility dmso anthropogenic stressors cause ecological surprises in boreal lakes. Global Change Biol 2006,12(12):2316–2322.CrossRef 22. Vidussi F, Mostajir B, Fouilland E, Le Floc’h E, et al.: Effects of experimental warming and increased ultraviolet B radiation on the Mediterranean plankton food web. Limnol Oceanogr 2011,56(1):206–218.CrossRef 23. Doyle SA, Saros JE, Williamson CE: Interactive effects

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Typically, the self-assembly this website of noble-metal nanoparticles has attracted much attention because of their unique plasmon resonance and their tremendous applications in the area of optical waveguides [6], superlensing [7], photon detection [8], and surface-enhanced Raman scattering (SERS) [9–12].

Recently, the SERS effect based on noble-metal ensembles is of particular interest because of its extraordinary ability to detect a wide variety of chemical/biological species at extremely low concentrations even down to the single-molecule level [9]. Gold nanoparticles (GNPs) have been widely used as Raman active substrates because of their good biocompatibility and strong SERS enhancement [13–18]. However, it should be mentioned that the particles tend to aggregate during aging, which results in an unwanted reduction of the active surface area [19, 20]. To address this issue, the fixation of GNPs in one-dimensional (1D), 2D, or 3D spaces can avoid the aggregation of the particles as SERS substrates. Tsukruk et al. assembled GNPs onto 1D silver nanowires and 2D silver nanoplates to create bimetallic

nanostructures as efficient single-nanoparticle Raman markers [21]. Li et al. developed a 2D GNP monolayer film as SERS substrate by the self-assembly of nanoparticles at a liquid/liquid interface [22]. Zhang et al. reported that GNPs dispersed on the grapheme oxide (GO, 2D) and reduced graphene oxide (RGO, 2D) supports exhibit excellent SERS and catalytic performance compared click here with the metal nanoparticles alone [23]. Qian et al. prepared the self-assembled 3D-ordered GNP precursor composite (SiO2/GNPs) arrays as SERS

nanoprobes [24]. Choi et al. reported a highly ordered SERS-active surface that is provided by a 3D GNP array based on thermal evaporation of gold onto an indium tin oxide (ITO) surface through a nanoporous alumina mask [25]. This SERS-active surface was applied to analyze the intracellular state. Therefore, the development of appropriate support materials to fix GNPs is very important in practical SERS detection applications. Recently, 3D Ag microspheres (AgMSs), which contain special fine structure, large specific surface area, and click here micron-sized particles, have been applied as SERS substrates [19, 26]. For example, Zhao et al. prepared 3D AgMSs with nanotextured surface morphology by a simple, sonochemical, surfactant-free method. Due to their special structural features with nanoscale corrugations, the obtained 3D silver microstructures showed a structurally enhanced SERS performance [19]. Zhang et al. developed hierarchical assemblies of silver nanostructures as highly sensitive SERS platforms by an acid-directed assembly method [26]. Our group also used proteins [27] and microorganisms [28] as templates to synthesize AgMSs and hollow porous AgMSs, respectively. However, the controlled synthesis of AgMSs with clean rough surface is still a significant challenge.

The observation that this short sunitinib treatment did not affect tumor growth is in line with our previous experience with tumors of the same melanoma line growing in dorsal window chambers [11]. In JQEZ5 molecular weight that study, we observed that 4-days with sunitinib treatment did not affect tumor growth, whereas

tumor growth was reduced when the treatment was continued for 8 days. Treatment-induced reductions in tumor size generally occur late after antiangiogenic treatment [5]. If non-responding patients could be identified shortly after treatment initiation, any ineffective treatment could be stopped to avoid toxicity, and other treatments could be considered. In the current study, a short treatment period was chosen deliberately to investigate whether DW-MRI and DCE-MRI can detect treatment-induced effects occurring before reductions in tumor size. Our study suggests that these MR techniques may be used to identify patients that respond to antiangiogenic treatment before treatment-induced reductions in tumor size can be detected. Sunitinib-treated tumors showed reduced K trans and increased RG7420 ADC values.

The reduction in K trans could be attributed to several vascular effects, but sunitinib-induced reduction in microvascular density was probably the dominating effect. We have previously shown that K trans reflects vessel density in untreated A-07 tumors [24, 28], and in the current study sunitinib-treated tumors showed significantly lower microvascular density than untreated tumors. Sunitinib-induced inhibition of VEGFR-2 may also have reduced vessel permeability, because VEGF-A signaling is known to increase vessel permeability [29]. The reduction in K trans may thus also be influenced by reduced vessel permeability. The increase in ADC was probably a result of sunitinib-induced necrosis. Sunitinib-treated tumors showed massive necrosis whereas untreated tumors did not show necrotic regions. Elevated ADC values have been found in necrotic tissue in untreated tumors [12, 13], and increases

in ADC reflecting treatment-induced necrosis have been reported after chemotherapy, radiation therapy, and treatment with vascular disrupting agents [6]. In the current study, DW-MRI was performed by choosing b values of 200-800 s/mm2 to avoid confounding effects of Janus kinase (JAK) blood perfusion, as recommended by Padhani et al. [30]. It is therefore unlikely that the ADC values reported here were significantly influenced by vascular effects. The present study thus strongly suggests that ADC and K trans reflected different physiological parameters, illustrating that it may be beneficial to combine DW-MRI and DCE-MRI when evaluating effects of antiangiogenic treatment. It has been suggested that antiangiogenic agents including sunitininib can normalize tumor vasculature and microenvironment and hence sensitize tumors to conventional therapy [4, 31].

This suggests that Ge/GeO x layers are observed rather than pure Ge NWs, which should help to obtain good resistive switching memory characteristics. To observe the defects in the Ge/GeO x NWs, we recorded PL spectra of the NWs, as shown in Figure 3a. To understand the temperature dependence of the PL spectra, the peak was normalized with respect to PL at 300 K. No significant shift of the emission peak with temperature learn more was observed. However, the PL intensity gradually increases as the temperature increases from 10 to 300

K, revealing that more defect states are activated as the temperature is raised. To identify the defects inside the Ge/GeO x NWs, the PL spectrum measured at 300 K was decomposed into four component peaks using Gaussian fitting, as shown in Figure 3b. The peaks are centered around 387 nm (3.2 eV), 402 nm (3.1 eV), 433 nm (2.9 eV), and 483

nm (2.6 eV). Violet-blue emission is observed from these Ge/GeO x NWs. Because of their large diameter of approximately 100 nm, the quantum confinement effect is not the origin of this broad emission spectrum [41]. Therefore, selleck compound the PL peaks probably originate from oxygen vacancies (V o), oxygen-germanium vacancy pairs (V Ge, V o), and related defects. The broad violet-blue emission can be explained by a simple mechanism. It is assumed that acceptors will form (V Ge, V o), and the donors will form V o. After the excitation of acceptors/donors, a hole (h o) and electron (e) are created on the acceptor and donor, respectively, forming (V Ge, V o) and (V o) according to the following equation [42]: (1) where h is Plank’s constant and

ν is frequency. The violet-blue emission occurs via the reverse reaction. This suggests that the vacancies exist in the Ge/GeO STK38 x NWs, which may improve their resistive switching memory performance. A schematic diagram of the NW-embedded MOS capacitor in an IrO x /Al2O3/Ge NWs/p-Si structure is shown in Figure 4a. The capacitance (C)-voltage (V) hysteresis characteristics of the Ge/GeO x NW capacitors with different sweeping voltages from ±1 to ±5 V were investigated, as shown in Figure 4b. Memory windows of 1.7 and 3.1 V are observed under small sweeping gate voltages of ±3 and ±5 V, respectively. In contrast, a small memory window of 1.2 V under a sweeping gate voltage of ±7 V was observed for the device without Ge/GeO x NW capacitors because of the degradation of the GeO x film (data not shown here). The larger memory window of the device containing Ge/GeO x NW capacitors compared with those without the capacitors may be caused by effective charge trapping on the surface of the Ge/GeO x NWs. Defects on the surface of the Ge/GeO x NWs will trap holes rather than electrons because the C-V signal shifted towards the negative side, which was also observed in the PL spectrum of the NWs.

The C. albicans sur7Δ mutant has an abnormal response to induction of filamentation and hyphal cells are markedly defective in plasma membrane structure An important virulence attribute in

C. albicans is the ability to switch between yeast, pseudohyphal, and filamentous forms [25–27]. When spotted onto M199 agar, hyphal structures were formed from each colony (Fig. 4A). However, the extent of filamentation was reduced in the sur7Δ null mutant compared to DAY185 and the SUR7 complemented strain. Similar results were observed when grown on Spider agar medium at 37°C (Fig. 4A). When BSA agar plates were incubated for an extended period of time, filamentous structures emerged from the edge of each colony except in the sur7Δ null mutant (Fig. 4A). This reduced filamentation in response to inducing conditions was also seen on solid media containing VRT752271 in vivo fetal calf serum (Fig. 4A). In YH25448 molecular weight liquid media (YPD supplemented with 10% FCS, high glucose D-MEM with 10% FCS, or RPMI-1640), time of germination and the extent of filament elongation of the C. albicans sur7Δ mutant were grossly similar to the wild-type and SUR7 complemented strains (data not shown). However, when grown in weak hyphal-inducing liquid Spider medium, a population of yeast cells and hyphae with aberrant morphology and branching was observed (Fig. 4B). Figure 4 Filamentation assays on various media.

(A) Overnight cultures were spotted onto weak-inducing media such as M199 agar plates, Spider agar, and BSA plates, and monitored daily. Overnight cultures were also spotted onto YPD containing 10% (v/v) fetal calf serum (FCS), a strong inducer of filamentation. Representative figures at the indicated times and incubation temperatures are shown. (B) Filamentation was also assayed in liquid media. Inoculums of 5 × 106 cells ml-1 were incubated at 37°C with constant shaking at 200 rpm. The time of germination, extent of elongation,

and overall Tyrosine-protein kinase BLK hyphal morphology were observed and compared between each strain at given time points using standard light microscopy. Results from growth in weak-inducing medium (Spider medium) are shown here at 2 and 4 hours where aberrant branching is evident at the latter timepoint. Standard light microscopy was performed using a 60× and 40× objective for the 2 and 4 hour timepoint, respectively. Next, structures of the filamentous form were compared using light microscopy. After 24 hours of growth, the wild-type (DAY185; Table 1) and SUR7 complemented strains produced mature, elongated hyphal cells with clear septa, whereas the sur7Δ null mutant produced irregularly shaped hyphae with obvious intracellular invaginations (Fig. 5A). Thin-section electron microscopy demonstrated subcellular structures in the filaments formed by the sur7Δ null mutant strain (Fig.

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