This unexpected coupling between polarized light and ferroelectric polarization modifies the stress induced in the BaTiO3 at the domain wall, which is observed using in situ confocal Raman spectroscopy. This effect potentially leads to the non-contact remote control of ferroelectric domain walls by light.Ferroelectric materials exhibit spontaneous and stable polarization, which can usually be reoriented by an applied external electric field. The electrically switchable nature of this polarization is at the core of various ferroelectric devices. The motion of the associated domain walls provides the basis for ferroelectric memory, in which the storage of data bits is achieved by driving domain walls that separate regions with different polarization directions. Here we show the surprising ability to move ferroelectric domain walls of a BaTiO Domain walls between ferroelectric domains are of interest for ferroelectric memory and to achieve a better control of the switching process. Here, the authors induce the motion of ferroelectric domains by light, creating a new possibility to control ferroelectrics. Ferroelectric devices, as nonvolatile ferroelectric random access memory (FeRAM)3 (ferroelectric polarization ~100 μC cm−2 and saturation magnetization of 150 emu cm−3)567For this reason, the ability of moving domain walls in ferroelectric materials without a physical contact fascinates the scientific community. This motivation has induced an important research activity in multiferroic materials, combining the advantages of both ferroelectricity and ferromagnetism for new nonvolatile memories, which can be written electrically, but read magnetically. Recently significant room temperature coupling was demonstrated by monitoring changes in ferroelectric domain patterns induced by magnetic fieldset al.3 (BTO) showing electron gas-like conductivity, while the individual domains remained excellent insulators. The discovery of this stimulant behaviour from the sCDW described for the BTO-based material123 thin filmsRecently, in a pioneering study, SlukaThe interaction of light with matter above described for the photovoltaic effect can be further exploited by the generation of photostriction on ferroelectric materials. The photostrictive effect is a phenomenon in which strain is induced in the sample when illuminated151615161819Here we have made a breakthrough that could lead to devices without electrical connections, directly converting energy from light into ferroelectric domain wall motion. To achieve this purpose, we have correlated an unexpected coupling between polarized light and ferroelectric polarization, which modifies the stress induced at the domain wall. As a result, these properties are particularly attractive, because domain walls move without applying an electrical field and the process is reversible accordingly with the intrinsic ferroic nature of the crystal. This new effect potentially leads to remote control of ferroelectric domain walls by light, which opens a framework for micro and nanoscale devices with non-contact domain engineering.c-plane and (h00) or a-plane. Study of the domain structure was performed by optical microscopy and atomic force microscope (AFM). A basic identification of the structure and of the crystalline orientation of the single crystal was performed using X-Ray Diffraction see . The sam6 octahedra and by the cationic displacementsA (red) and C (blue) points.The AFM technique gives topographical information of the domain structure but yields no structural information. Recent methods based on confocal Raman microscopy (CRM), give the possibility to study at a local scale the structural deformations of perovskites, induced both by the tilting of BO−1 region  band . Raman peaks at 520 and 560 cm−1 are attributed to the same A1(TO3) Raman mode, even if the physical nature of band asymmetry is not yet established. Consequently, the red spectrum of the a-domain. Thus, red and blue spectra correspond, respectively, to a- and c-domains.Clear differences can be observed in the shape of both spectra, particularly in the 440 to 640 cmand 5 in  ~520 cm−c-domain regions (out-of-plane polarizations) and the protrusion evidenced by AFM, as well as between a-domain areas (in-plane polarization) with the trough determined by AFM. Generally, the plane of a–c-domain walls is {110}pc at 45° to the top surface22Pm3m cubic→tetragonal P4mm). The 90° ferroelastic domains are generated, each of them gives rise to two 180° ferroelectric domains. As a consequence, domain boundaries are along {110}Cubic lattice planes, with ≈45° angle between the boundary and the (100)Tetragonala-plane or the (001)Tetragonalc-plane of the tetragonal cella–c-subdomains observed for the single crystal are due to twinning, induced either by synthesis (phase transition) or by the cutting step (ferroelastic transformation)a–c-domain and c–a-domain walls slopes ~20° to the top surface. Further experiments are in progress to confirm this result, even if this result seems to be in agreement with recent studiesIn addition, there is a clear correlation between B in a–c-domain wall and is clearly located at the step detected from the topographical analysis , is carried out , there is a slight increase of the background spectra for the b-domains, in which incipient Raman peaks at 166 and 199 cm−1 seems to appear. The appearance of nanometer-sized local structures with rhombohedral symmetry was recently evidenced in both the orthorhombic and tetragonal phases of BTO single crystal by convergent-beam electron diffraction−1 are attributed to the presence in the lattice of different kinds of BO6 octahedra. Nevertheless, other Raman peaks could not be associated with the rhombohedral phase−1, as reported in monoclinic single crystal of BaTi2O5 TixO3 (PZT) thin films is particularly sensitive to embedded ferroelastic domains and can be dramatically enhanced by manipulating the domain structure to either remove these domains or increase their mobility3 thin films, as predicted by Meyer et al.Taking into account their size, orientation and morphology, the b-domain would allow the rotation of the polarization. Consequently, this b-domain is associated to a non 90° polarization vector and probably electrically charged.The twinned tetragonal model only allows ≈90° and ≈180° angles between polarization vectors of neighbouring domains23b-domains clearly appear in the a–c-domain wall due to higher crystalline stress in the a–c-domain wall than in the c–a one. Due to the boundary conditions of the crystal surface, the c-domain protrudes accordingly with out-of-plane distortion, while the a-domain is depleted. The a-domain partly overlays the c-domain and limits such protrusion. As a result, the a–c-domain wall is under compressive stress, minimized by the complex domain structure, which increases the domain density. In the c–a-domain wall, the protrusion of the c-domain region is favoured because in such case the c-domain is located at the surface and the a-domain is below it. Thus the c–a-domain wall possesses less stress and the complex domain structure is not favoured.The b-domain structure is built by combining the AFM and Raman mapping information (a-domains (red) and c-domains (blue). The b-domains (green) appear in the a–c-domain wall within the ({101}pc) plane. The a–c-domain wall is associated with a head-to-head 90° domain wall, where the mechanical stress is enhanced. We also envisage that the head-to-head configuration maximizes the bound charge close to the domain wall. The b-domains minimize the internal stress by increasing the domain wall density,  seems to be behind such behaviourb-domain structure must also affect the charge distribution at the domain wall.Despite the fact that the existence of stress is resolved by increasing domain wall density, b-domains are associated with a significant stress degree and according to the literature by an uncompensated chargein situ observations of polarization switching are carried out by applying polarized light. In all cases the incidence is normal to the sample surface and the polarization direction of light parallel to the surface. The angle between the polarization of light and the depth scan direction varies by using Δθ=15° between θ=0° and θ=90°. The scanning direction is always perpendicular to the a–c-domain wall. Thus, in the a-domain case the polarization of light and thus its associated electric field (E) is parallel to in-plane polarization for θ=0°  and perpendicular for θ=90°. Consequently, the electric field have no effect on the polarization vector of BTO cell for θ=0°  and acts with a maximum force for θ=90° (E and P are perpendicular). Thus, for the a-domain, the effect of the electric field implies in-plane rotation of the polarization vector. The a-direction remains parallel to the surface of the sample, which means that this domain remains to be a-domain even with re-orientation of its polarization direction under polarized light.Following the above analysis, the c-domain, the polarization of light is always perpendicular to the out-of-plane polarization. Whatever the θ angle, the electric field associated to polarized light have the same effect. For the b-domain, a combination of both behaviours is expected as a result of the bundle of a–c-subdomains. Thus b-domains are probably the most sensitive to electric field and thus to the polarized light.For θ angles. The line D→E in x-axis for both the b-domains (green) and the c-domains (blue). When the polarized light reaches θ=90°, the Raman spectrum of the c-domain evolves towards b-domain (D and E in a-domain (red), located on top surface of the sample, appears as unchanged. The local change of domain configuration seems to imply the displacement of the b-domain (green) to the detriment of the c-domain (blue). This observation supports our previous hypothesis of charged b-domains: these charges are highly sensitive to electric field and associated to the motion of domain walls.b-domain ., accoun6 octahedra, associated with the crystalline stress: We used confocal Raman mapping to understand this complex domain structure. The A1(TO2) Raman active phonon is a symmetrical mode, which is detected as relatively strong scattering signals in BTO because of a near-perfect equilateral octahedral symmetry. 1(TO2) Raman mode, for the different light polarization angles. Raman shifts as large as ~15 cm−1 (from 205 to 220 cm−1) are determined in the complex domain structure.However, the domain wall motion observed here implies a modification of tetragonality and polarization, at least at local scale. The Raman shift is an indicator of the crystal stress and correlated with both tetragonality and polarization40c-domains (blue) near the a–c-domain wall. This implies a higher tetragonality in this region and as a consequence a higher off centering of the Ti-cation, which means higher polarization. The surprising increase of polarizability could be explained by the presence of local charge accumulation at the 90° domain wall. The higher Raman shift is thus an evidence of the enhancement of the local electric field provided by the head-to-head a–c-domain wall. Besides, the local electrical field also competes with the compressive stress. On the contrary, the b-domain  Raman mode. When the polarization light rotates between 45 and 90°, the redshift is three times higher than in the 0°–45° range. This fact clearly shows a correlation between the domain wall relative motion and the structural changes. The point 2  presents a higher stress and higher tetragonality, less sensitive to the light polarization angle. The presence of a local electric field or stress concentration effects seem to be more relevant than the out-of-plane polarization in the c-domain region close to a–c-domain wall.−1, that is, ≈6 J cm−3 or 6 pJ μm−3). Since the angular dependence of polarized light is observed, the origin of such phenomena is clearly correlated to the interaction between the electric field of polarized light and charges or electrical dipoles, such as the bound charge associated to the head-to-head 90° domain wall or the polarization vector of tetragonal BTO. To analyze such coupling, we studied the evolution of the Raman spectra of surface a-domain, b-domain and c-domain as a function of the light polarization angle (−1) and appearance of new Raman modes  at 170 cm−1, E(TO2)/E(LO) at 180 cm−1 and A1(LO) at 185 cm−1 /E(LO) at 178 cm−1 is almost missing, which are sensible to the polarization direction. Thus, the lower presence in c-domain denotes an out-of-plane polarizationθ=0° is incipient because of the Raman mode splitting at 192 cm−1 (associated with blueshift of A1(LO) for tetragonal symmetry) and 201 cm−1. This effect is less clear in b-domain and absent in a-domain.First, at low wave number, tetragonal BTO presents Raman modes, assigned to Am−1 refs , 47, 48,θ=90° Raman spectrum shows a different scenario in which Raman modes ascribed to rhombohedral structure gained intensity. Raman modes are evidenced at 154, 193 and 199 cm−1 /E(LO). This means that the c-domain changes from out-of-plane to in-plane polarization as a consequence of the rotation of the polarization direction. However, there is a coexistence between <001>pc and <111>pc polarization directions, probably associated with nanometer-size regions2)/E(LO) Raman mode for the c-domain, in comparison with a-domain, indicates a structural change as a consequence of the same polarization rotation. It is worth remarking that this situation occurs in the c-domain and is related to the application of light polarized perpendicularly to the polarization direction. Thus, the rotation of the polarization from <001>pc to <111>pc is a direct effect of the energy transfer from the polarized light to the crystalline structure. The stress reduction is, however, limited because the out-of-plane polarization is in compressive stress and the in-plane switching is mechanically clamped. The b-domain produces always an intermediate response between a-domain and c-domain response. The b-domain Raman redshift denotes a stress reduction as a consequence of the higher domain density of the subdomain structure.The 199 cm−1 . The c-da-domain, the increase of asymmetry observed in Raman mode at 560 cm−1  Raman mode is a feature of the appearance of polarization rotation towards the <111>pc direction. The distorted structure possesses higher bonding constant force for such Raman mode and higher blueshift as well. From a structural point of view, this distorted structure appears as a local symmetry, allowing the rotation of the polarization vector during the domain wall motion. The concomitant observation of Raman modes characteristic of BTO rhombohedral polymorph allows thinking that this local structure is probably of rhombohedral symmetry , thus with polarization along the <111>pc direction. This agrees well with the recently reported observation by transmission electron microscopy of the local structure of rhombohedral symmetry in the orthorhombic and tetragonal polymorphs of BTOFor a–c-domain wall is under compressive stress. This stress promotes a complex domain structure, in which a-domain, c-domain and a–c-subdomain regions (named b-domain in this work) increase the domain density to minimize stress. We observed different BO6 octahedra by the appearance of forbidden phonons in the Raman spectra as a signature of the occurrence of nanometer-sized local structures with <111>pc polarization direction. The <001>pc polarization rotation towards <111>pc occurs through the coupling of the polarized light with the crystalline structure. Thus the polarized light transfers its energy to the crystalline structure to effectively reduce the local mechanical stress. There is a clear correlation between the domain wall relative motion and the light polarization angle. The relaxation of the crystal structure is evidenced by the Raman modes redshift for the different domain regions. The domain wall motion is constricted by local compression stresses and in the case of out-of-plane polarization the relaxation is limited, while the in-plane is more favoured.In summary, the ferroelectric domain wall motion induced by a polarized coherent light source at room temperature has been successfully demonstrated in BTO single crystal. The differences between ferroelastic and ferroelectric directions yield a complex domain structure with characteristic 90° domain walls forming <45° from the top surface. On this 90° domain wall, the To conclude, the implication derived from this study is that we can surprisingly modulate the motion of the ferroelectric domain walls by varying the polarization angle of a coherent light source. The BTO single crystal is relatively simple in domain structure when compared with ferroelectric ceramics and in particular with morphotrophic phase boundary piezoelectric, as PZT ceramics, in which coexistence of different symmetries are on the base of superior piezoelectric properties. It is worth pointing out that this stimulant behaviour observed here could have potential technological applications leading to a non-contact read-out method in FeRAM devices or remote control of piezoelectric actuators. These findings undoubtedly open questions to be solved by the scientific community as well new application possibilities for ferroelectric materials.a-plane. The single-crystal was polished using 1 μm diamond paste, then cleaned with acetone and ethanol before characterization. No further thermal and/or chemical etching was used to reveal the domain structure, thus avoiding topographic artifacts due nanoroughness. The sample was maintained at T>25 °C during the AFM and CRM using a temperature controller. In addition, the reader can find more information about the full domain structure of the BTO single crystal in c-plane and a-plane) was performed using X-Ray Diffraction . The lattice parameters were refined by a global simulation of the full diagram . The orientation degree was determined using the intensity ratio of the two characteristic diffraction peaks I(002)/I(200).The BTO single crystal used in this study, hereafter referred as ‘BTO’, was produced by top-seeded solution growth and provided by PIKEM Ltd (UK). The 5 × 5 × 1 mm BTO crystal was grown with (100) orientation or r=50%, recording both topography and phase images simultaneously. Raman spectra were obtained using a frequency-doubled Nd-YAG laser operating at 532 nm and a × 100 objective lens . The incident laser power was 40 mW. The optical diffraction resolution of the confocal microscope was limited to about ~250 nm laterally and ~500 nm vertically. Raman spectral resolution of the system was down to 0.02 cm−1. The microscopy sample was mounted on a piezo-driven scan platform having 4 nm lateral and 0.5 mm vertical positional accuracy. The piezoelectric scanning table allows steps of 3 nm , giving a very high spatial resolution for both the AFM and the CRM. The microscope base was also fitted with an active vibration isolation system, active 0.7–1000 Hz. Collected spectra were analyzed by using Witec Control Plus Software.The experiments were performed using a CRM coupled with AFM . The sample was imaged in AFM AC mode using ArrowFM cantilevers  with a resonance frequency in the range of 70–90 kHz and damping of y-axis of the piezo-driven scan table and perpendicular to the x-axis. The light incidence was normal to the sample surface, the polarization direction of light was parallel to the surface, and the scan direction, denoted as x-axis, was maintained perpendicular to the a–c-domain wall. The in situ domain switching was monitored taking subsequent depth scans at different polarization angles of the same area, using as position reference a significant point of the sample surface visible with the optical microscope. Depth scan Raman images had 10 μm of width, 4 μm of height, 100 × 40 spectra of 0.2 s of integration time at 40 mW of laser power. Thus, the Raman image consisting of 100 × 40 pixels  required ~14 min for acquisition of the whole cross-section. In addition, to avoid the photoconductivity effects present in BTO, in our experiments a polarized light with a wavelength (at λ=532 nm) outside the two absorption maxima18θ=15° between θ=0° and θ=90° was rotated. Experiments were attempted in two ways: first, the light polarization angle varying in steps from 0°→90° and thereafter from 90°→0°, and second the light polarization angle randomly switching from selected steps. Collected spectra were analyzed by using Witec Control Plus Software.The sample was mounted under the microscope objective with the domains walls aligned parallel to the F.R.-M. designed and performed the experiments, analyzed the results and wrote the manuscript. A.D.C. carried out AFM and Raman Image experiments. P.M. assisted in experiments and contributed to data analysis and the writing of the manuscript. J.F.F. initiated and supervised the project and contributed to the writing of the manuscript.How to cite this article: Rubio-Marcos, F. et al. Ferroelectric domain wall motion induced by polarized light. Nat. Commun. 6:6594 doi: 10.1038/ncomms7594 (2015).Supplementary Figures 1-4, Supplementary Tables 1-2 and Supplementary References


The critical ratio t/d for the worst filling ability increases with cavity width t and tends to disappear when the cavity width t increases to 300 μm. A polycrystalline filling model considering the friction size effect, effect of constrained grains by the tools, grain size, cavity width and ratio of cavity width to grain size is proposed to reveal the filling size effect in micro coining. A quasi in-situ Electron Backscatter Diffraction (EBSD) method is proposed to investigate filling mechanism in micro coining. When several grains across the cavity width, each grain deforms heterogeneously to ordinate the deformation compatibility. When there is only one grain across the cavity width, the grain is fragmented into several smaller grains with certain prolongation along the extrusion direction to coordinate the deformation in the cavity. This is different from the understandings before. Then the filling deformation mechanism is revealed by a proposed model considering the plastic flow in micro coining.In this study, interactive effects of microstructure and cavity dimension on the filling behaviors in micro coining were investigated. The results indicate that the filling ability is dependent on both the cavity width  To trace the deformation behaviors of individual grain in micro coining, a quasi in-situ EBSD method is proposed in micro forming, as shown in kN and velocity of 0.36 mm/min. The deformed sample is taken out from the dies and scanned using EBSD again on the same area above, as shown in in-situ EBSD method, the evolution of individual grain in the deformation region is traced. The deformation behaviors of the individual grain can be analyzed by comparison the microstructural characteristics before and after deformation. From In micro coining, there are usually only several grains or even only one grain across the cavity width. The deformation behaviors dependent on the microstructure distribution are inhomogeneous because of heterogeneous materials in the deformation region. It is difficult to analyze the filling behavior using the traditional models. Thus, the Kernel average misorientation (KAM) as characterization of local misorientation is calculated by averaging the misorientation among the point at the center of the kernel to its nearest neighbors. KAM is commonly used as a tool in OIM (orientation imaging microscopy) to assess the magnitude of the residual plastic strain in the grains of the deformed metals313334Based on the analysis above, when there are several grains across the cavity width, the filling behavior is complicated because of the heterogeneous microstructure coupling the effect of processing parameters. The deformation degree is dependent on the grain distribution strongly. The constrained grains by the tools deformed heavily, the grains with free surface just changing their orientations to coordinate the inhomogeneous intergranular deformation with slightly plastic deformation. Grains in the inner part across the cavity width, the role the grains is to coordinate the inhomogeneous intergranular deformation by rotation and slip with their neighbouring grains. Therefore, the filling behavior is accomplished through intergranular and intragranular heterogeneous deformation to maintain the deformation compatibility and fill the cavity when there are several grains across the cavity width.Vx) and (2) parallel to the extrusion direction (Vy). The flowing velocity Vx at the region B will induce the transverse flow of the materials in the region A. The main and second slip planes are corresponding to the longitudinal and transverse directions, as shown in t and the ratio of cavity width to grain size t/d strongly. The coupling effects of microstructure and cavity dimension results in the worst filling ability when t is 50 μm and t/d is 1.04. The critical ratio t/d increases with the cavity width t and tends to disappear when the cavity width t increases to 300 μm, because of the transformation of the deformation patterns from micro coining to micro extrusion.The filling size effect is dependent on both the cavity width A polycrystalline filling model in micro coining is proposed considering the friction size effect, the effect of constrained grains by the tools and grain size, cavity width, ratio of cavity width to grain size. These parameters contribute negative or positive effect individually on the filling ability and their competition leads to the filling size effect in micro coining.in-situ EBSD method is proposed to investigate the filling process in micro coining. The intergranular and intragranular deform heterogeneously to ordinate the compatibility of different grains in the cavity when several grains across the cavity width. However, smaller grains with certain prolongation distribute along the extrusion direction are formed when there is only one grain across the cavity width because of severe plastic deformation in micro coining. This finding is different from the previous reports and interpreted by a proposed model considering the severe shearing deformation in the cavity.A quasi In this study, the interactive effects of microstructure and cavity width on the filling ability of pure nickel polycrystals in micro coining were investigated. The conclusions are drawn as follows:kN. To reduce the effect of friction on the deformation behaviors, all the tests were carried out with lubricant of castor oil. The specimens were pressed with a load of 10 to 40 kN and a low punch velocity of 0.36 mm/min was used for all tests in this research.In this research, a female die with U-shape micro groove is designed to conduct the micro coining process and to investigate the deformation behaviors in micro coining. The dimensions of the micro grooves are 50, 100, 200, 300, 400 and 500 μm in width and with a specific ratio of groove depth to width of 3. Pure nickel with purity of 99.8% is selected to conduct the micro coining tests. The specimens were manufactured by a precision machining process. The dimensions of the rectangular specimen are 5.5 × 3.5 × 2.0 mm with a corner of R0.5. To homogenize the microstructure of the as-received materials, the material was treated at temperature of 500, 650, 900, 1000, 1025 and 1100 °C for 3 h, and then cooled in air. The grain sizes ranges from 13, 23, 48, 107, 267 and 490 μm, respectively. The micro coining tests were carried out by a Zwick testing machine with a load cell of 100 How to cite this article: Wang, C. et al. Interactive effect of microstructure and cavity dimension on filling behavior in micro coining of pure nickel. Sci. Rep.6, 23895; doi: 10.1038/srep23895 (2016).


It is believed that the failure mode of metal nanowires under tensile loading is the result of the nucleation and propagation of dislocations. Such failure modes can be slip, partial slip or twinning and therefore they are regarded as local deformation. Here we provide numerical and theoretical evidences to show that global deformation is another predominant failure mode of nanowires under tensile loading. At the global deformation mode, nanowires fail with a large contraction along a lateral direction and a large expansion along the other lateral direction. In addition, there is a competition between global and local deformations. Nanowires loaded at low temperature exhibit global failure mode first and then local deformation follows later. We show that the global deformation originates from the intrinsic instability of the nanowires and that temperature is a main parameter that decides the global or local deformation as the failure mode of nanowires. Mechanical responses of metal nanowires have been extensively studied since they show unusual behaviors such as very large elastic range and higher elastic moduli compared to their bulk counterparts. In particular, failure event of metal nanowires under uniaxial tensile loading has been of great interest, because it gives us fundamental understanding about what makes the nanowires fails, when the nanowire fails, and what deformation the nanowire undergoes in that moment. Therefore, a lot of simulations123567Most predominant origin of local deformations that makes nanowires fail is the nucleation and propagation of dislocations. Since the energy barrier for the nucleation of dislocation is lower at edges than on surfaces or in bulk, the dislocation initiates from one of edges of the nanowire, propagates through the cross-section, and finally forms one of local deformations4151718x-direction to be the length direction (the [100]-direction), and y- and z-directions to be the lateral directions . Periodic boundary condition was applied in the length direction. To study the temperature effect on the tensile tests of nanowires, we assigned the temperature to the nanowires ranged from 0.01 K to 300 K using Nosé-Hoover thermostat238 s-1. Timestep was 1 femtosecond and all calculations were conducted by LAMMPSIn order to find the failure mode of metal nanowires, we performed uniaxial tension tests for metal nanowires by using molecular dynamics (MD) simulation. The face-centered-cubic (FCC) (001) nanowires of seven metals were modelled with embedded atom method (EAM) potentials2021ε = 0) to the configuration where the stress is the maximum (ε = 0.108), the square nanowire is contracting in both lateral directions (the y- and z-directions) by the same amount, and thus the shape of cross-section maintains square. In many previous reports22627ε = 0.113) in y-direction) and sudden contraction in the other lateral direction (here in the z-direction) occurs simultaneously. Now, the shape of cross-section is not square but a similar shape of a rectangle. At ε = 0.113, the ratio of a/b becomes 1.1 as shown in ε = 0.120). The changes in the sizes of nanowire in the lateral directions are shown in We plot the stress and energy curves to strain of a Au (001) nanowire in B is the elastic stiffness matrix in the Voigt notation. When σ1 > 0), the general condition is simplified to C22−C23 > 0, where C is the elastic modulus matrix of the structure. The corresponding eigenstate to this instability condition is  = , i.e. the deformation with a sudden expansion in a lateral direction and a sudden contraction in the other lateral direction. The magnitude of expansion is the same with the magnitude of contraction  bulk material. Again, the stress drops at this strain but the energy does not change. The elastic instability theory predicts well the failure behavior of cubic bulk materials.This global deformation has been predicted in many cubic bulk materials under uniaxial loading. This phenomenon in cubic bulk materials is called as branching or bifurcation of crystal structures and has been explained by the elastic instability theory. The theory was initially developed by Born151718For both FCC bulk and nanowire structures under uniaxial tensile loading, they eventually undergo the bifurcation: a contraction in one lateral direction and an expansion in the other lateral direction. However, details of their deformations at the bifurcation are different from each other, because there are substantial effects of free surfaces and edges for nanowires. Under uniaxial tensile loading, only one normal component of stress is nonzero (positive) and the others are zero. Under this loading condition, all the atoms in a bulk material have the same stress-state and the total stress is also the same: only one nonzero normal component of the stress. However, for nanowires, whereas the total stress has only one nonzero component, the stress state of each atom is different depending on its position in the nanowires. For FCC nanowires, there is a large relaxation of free surfaces and edges, and thus each atom is redistributed according to the relaxation. As a result, all atoms of the nanowires have three or more nonzero components of stress, and further the magnitudes of nonzero component are different. Stress is usually more tensile at the outmost atomic layers and less tensile at the interior part. Therefore, atomic displacements of the nanowires before and after the bifurcation are dependent on the positions of atoms, and thus they are different from those of bulk materials. Consequently, after the failure, the cross-section of the nanowire is not exactly rectangular like the case of bulk, as shown in C22-C23 > 0, by the condition, N is the total number of atoms and k. Therefore These different displacements or stresses of atoms in nanowires make it difficult to predict the instability moment by adopting the elastic instability theory, because atomic elastic moduli of the nanowires also become functions of atomic position. The elastic moduli at surfaces or edges are different from those in the interior part. Therefore, we need to develop an equivalent elastic modulus that can predict the instability phenomenon of the nanowires. Among possible choices, we replaced the condition, et al.et al.The global deformation (bifurcation) takes place not only in Au [100]/(001) nanowires but also in other FCC metal [100]/(001) nanowires. We performed the same MD simulations to study the failure mode of other nanowires, namely Al, Ni, Cu, Pd, Ag, and Pt. All of the metal nanowires considered showed the same global deformation as their failure modes. Furthermore, in order to confirm the generality of the global deformation of FCC (001) nanowires as the failure mode, we employed three different interatomic potentials such as Cai and Yeδε1,δε2,δε3,δε4,δε5,δε6) =  can only be found in cubic materials under uniaxial stress or hydrostatic loading condition. In BCC (001) structures, this deformation mode takes place under uniaxial compressive stress condition. It is noteworthy that when we consider FCC structures with other crystallographic directions such as (111) or (110), the deformation mode becomes a function of elastic moduli unlike (001) structuresThe global deformation, which is the simultaneous occurring of the sudden expansion in a lateral direction and sudden contraction in the other lateral direction, is the unique property of FCC (001) nanowires under tension. This is because the deformation mode at onset of instability  nanowires regardless of strain rate.So far, we considered the simulations with the strain rate of 10ε = 0.090) at higher temperature (100 K), as shown in et al.So far, we have shown that the global deformation induced by the elastic instability is the failure mode of FCC (001) nanowires. Then, a couple of questions arise: why do the simulations show global deformation as the failure mode instead of the local deformation like many other reports? Do the observed results violate the previous studies? In order to answer the important questions, we performed the tension tests of a Au [100]/(001) nanowire at different temperatures: 0.01 K, 10 K, and 100 K. Local deformations are mainly the results of the nucleation and propagation of dislocations. The required stress for the nucleation of dislocations is critical for the occurrence of the local deformation, because the required stress for the propagation of dislocation is usually very low in metals. It has been known that the nucleation stress for dislocation is dependent on temperature: increase of temperature lowers the nucleation stress4, the yield stress of a circular nanowire was found to be 50% larger than that of a square nanowire with a similar size. However, the yield stresses, under which a local deformation occurs, were compared in their study. And, the nucleation stress of the dislocations that are responsible for the local deformation is different depending on the nucleation sites: edges for the square nanowire and surfaces for the circular one. Therefore, the yield stresses of the two nanowires should be different. In contrast to their study, here, both nanowires failed with a global deformation when their equivalent elastic moduli satisfied the instability condition, We have discussed that the nanowires with a square cross-section exhibit the global failure mode at low temperature. As temperature increase, local failure mode becomes dominant. The temperature at which the failure mode changes is approximately 12 K for a square Au nanowire  with a size of 3 nm × 3 nm. This transient temperature depends on the cross-sectional shape of nanowires. In order to investigate the effects of the shape of nanowires on the failure mode or the transient temperature, we performed the same MD simulations for a circular Au nanowire with a diameter of 3 nm. We changed the temperature of the circular nanowire from 0.01 K to 200 K. First, at 0.01 K, the circular nanowire failed with a global deformation at the critical strain of 0.108, which was the same with the critical strain of the square nanowire as shown in 4 on the effects of the shapes of nanowires on the yield strength of the nanowires. Using a strain shear invariant, they found that the edges of nanowires are the easiest sites for the nucleation of dislocations. Further, Zhu et al.Next, we found that the transient temperature of the circular nanowire is approximately 100 K, which is much higher than that of the square nanowire, 12 K. As discussed earlier, the nucleation stress of dislocations on the edges of a nanowire is lower than that on the surfaces of the nanowire. This is also supported by the study of Cao and MaEven though nanowires fail with a global deformation at low temperature, a local deformation takes place at last following the global deformation when more tensile loading is applied to the nanowires. Interestingly, at near the transient temperature which was approximately 12 K for this nanowire, we observed that the global and local deformations occurred simultaneously. At this temperature, the required stresses for the nucleation of dislocation and for the elastic instability are very similar to each other, and thus the nanowire undergoes both deformation modes simultaneously during the failure. At low temperatures (<12 K), global deformation takes place first and thus mechanical properties of the nanowire changes significantly during deformation. For examples, two extended surfaces by bifurcation of the nanowire are mechanically weaker than the other two contracted surfaces. As a result, later local deformation is determined by new mechanical properties. At near the transient temperature (~12 K), we observed that a dislocation initiated from one edge of the nanowire and propagated while the expansion and contraction of the nanowire along the lateral directions were taking place.et al. was used for the Ni nanowires. The transient temperatures of circular nanowires were much higher than those of square nanowires for both metals, because of the absence of edges in the circular nanowires as discussed previously. And, although the transient temperatures slightly differed as thickness increases, they did not show a clear dependency on the thickness. On the other hand, it is remarkable that the transient temperatures of the square Au nanowires were extremely low (<15 K) whereas those of the square Ni nanowires were much higher (~120 K). The transient temperatures of circular Ni nanowires were approximately 200 K which was approximately 100 K higher than those of circular Au nanowires. The transient temperatures we obtained through MD simulations are much lower than room temperature and thus it is hard to observe the global deformation as a failure mode because it takes place at temperatures lower than the transient temperature. However, we believe that it is still meaningful that there is another failure mode for FCC (001) nanowires at extremely low temperature.For all the seven metal (001) nanowires and the three different interatomic potentials we employed, there always exist the transient temperatures under which the global deformation is the failure mode for the nanowires and near which the global and local deformations occur simultaneously. However the transient temperatures strongly depends on the FCC metals and the interatomic potentials. For examples, we compared the transient temperatures of Au and Ni nanowires with different lateral sizes and cross-sectional shapes in We performed MD simulations to study the failure behavior of FCC (001) metal nanowires under uniaxial loading. We found that global deformation is a mechanical failure mode of nanowires under tension besides the commonly believed local deformation mode. At the failure of nanowires with the global deformation, a large expansion along a lateral direction and a large contraction along the other lateral direction take place simultaneously in the nanowires. Such phenomenon is driven by the elastic instability of materials. We also found that there is a competition between the global deformation mode induced by the elastic instability and the local deformation mode induced by the nucleation and propagation of dislocations. At low temperatures, the global deformation mode is dominant because the thermal activation is not enough to nucleate dislocations. At high temperatures, nanowires meet the condition for the nucleation of dislocations before they meet the elastic instability conditions and thus local deformation becomes dominant. At near the transient temperatures, nanowires fail by the simultaneous occurrence of the global and local deformations.Although we found that there are large differences in the transient temperatures of FCC (001) nanowires depending on the metals, all transient temperatures we obtained in MD simulations were much lower than room temperature. Therefore, we expect that FCC (001) nanowires under tensile loading will always fail by local deformations in real experiments at room temperature. However, it would be possible that one verify the global deformation as a failure mode of FCC (001) nanowires by experiments at extremely low-temperature.How to cite this article: Ho, D. T. et al. Mechanical Failure Mode of Metal Nanowires: Global Deformation versus Local Deformation. Sci. Rep.5, 11050; doi: 10.1038/srep11050 (2015).


P < 0.05) in taste, crispness, flavor and texture of the biscuits while significant differences (P < 0.05) existed in color and overall acceptability. The proximate composition of the biscuits ranged from 1.84% to 2.55% for moisture, protein 8.03–9.26%, fat 30.07–35.81%, ash 2.94–3.68%, crude fiber 0.47–0.80%, carbohydrate 48.74–55.96%, and energy 526.53–554.21 kcal/100 g. The microbial count of the best biscuit after 20 days of storage was 4.0 × 103 cfu/g for bacteria and mould contained 5.0 × 104 cfu/g. This study forms a basis for new product development for the biscuit food industry.The consumption of cereal foods such as biscuit has become very popular globally. Partial replacement of wheat flour with beniseed and unripe plantain flours rich in protein, vitamins, and minerals will increase nutrient, diversify utilization of beniseed and unripe plantain, and increase biscuit variety. Wheat composite biscuit was produced from wheat, beniseed, and unripe plantain flours. The composite flour was mixed in the proportion of 100:0:0, 80:10:10, 70:20:10, 60:30:10, and 50:40:10% of wheat, beniseed, and unripe plantain, respectively. The physical, sensory, chemical, and microbial properties of the biscuits were determined. The physical properties ranged from 6.80 g to 8.30 g for weight, spread ratio 6.93–7.38, and break strength 500–690 g. There was no significant difference ( Biscuit is a small thin crispy cake made from unleavened dough. Biscuits have been suggested as a better use of composite flour than bread due to their ready to eat form, wide consumption, relatively long shelf life, and good eating quality  is a cereal grain grown all over the world for its highly nutritious and useful gain. It is one of the top three most produced crops in the world, along with corn and rice. According to Okaka  or over ripe (and therefore sweet) , unripe plantain (Musa paradisiaca), fat , salt , eggs, and baking powder . The equipment and chemicals were of analytical grade.The consumable materials were purchased from Muda Lawal market, Bauchi, Bauchi State and they are as follows: wheat flour , beniseed  and nine (9) representing “extremely dislike” and “extremely like”, respectively, was used. The qualities assessed include color, texture, taste, flavor, crispness, and general acceptability. Coded samples of the same size and temperature (29 ± 3°C) were served on clean white plates to judges in different boots under the florescent light. Only one sensory attribute was tasted at one sitting.The moisture, protein, fat, ash, and crude fiber contents were determined by the AOAC . The totThe pour plate method of knowing and counting the number of viable bacteria present in the sample was adopted as described by Jideani and Jideani . Two graThe various results obtained were subjected to Analysis of Variance (ANOVA) and Duncan  MultiplePhysical analysis of the biscuit is shown in Table The spread ratio of the biscuit increased from 7.00 to 7.38. The increase is an indication of the binding properties of the flour and of the texture of the biscuits. This increase may be as a result of increase in beniseed flour as both unripe plantain and beniseed flour has poor binding properties.The break strength value ranged from 500 g to 690 g with sample 80:10:10% wheat, beniseed, and unripe plantain flour having the highest and sample 50:40:10% wheat, beniseed and unripe plantain flour having the lowest. This may be due to poor quality of beniseed and unripe plantain flour to form a strong network in both flours in a molten state at baking temperature.P < 0.05) in taste, crispness, flavor, and texture of the biscuits while significant differences (P < 0.05) existed in color and overall acceptability. Sample AMA  had the highest mean value of 6.85 when compared to 7.30 for 100% wheat flour biscuit. Sample OKO  had the lowest mean value of 5.50. This may be as a result of addition of beniseed and unripe plantain flour. This showed that sample AMA  compared favorably with the 100% wheat flour biscuit followed by sample MAK  and then sample AKA . There was a general decrease in the acceptability of the biscuits with increase in beniseed flour.The mean scores for the sensory evaluation of the biscuits are shown in Table The proximate composition of the best three biscuit samples, beniseed flour, and 10% unripe plantain are shown in Table The protein content of the biscuits increased from 8.03% to 9.26% with increase in the percentage of beniseed flour. This was due to the addition of the defatted beniseed flour which is a good source of protein supplement. The fat content increased from 30.07% to 35.81% with sample AMA  having the highest. This increase may be due to the addition of beniseed flour which is a good source of oil  after 20 days of storage in polyethylene pack with paperboard is shown in Table An acceptable biscuit was formulated from wheat flour, beniseed flour, and unripe plantain flour. The composition of beniseed flour and unripe plantain made them dietary items. Analysis carried out showed that the formulated biscuit is rich in protein, carbohydrate, ash, and energy. The formulation of the biscuit has provided another means of utilizing our abundant beniseed seed and unripe plantain fruit. The biscuit produced has low microbial load and therefore has a long shelf life.


Clostridium difficile is presented for the first time. The cathepsin L-like fold of cysteine protease domain, a newly observed ‘lectin-like’ domain and several other features are described.The crystal structure of Cwp84, an S-layer protein from  Clostridium difficile is a major problem as an aetiological agent for antibiotic-associated diarrhoea. The mechanism by which the bacterium colonizes the gut during infection is poorly understood, but undoubtedly involves a myriad of components present on the bacterial surface. The mechanism of C. difficile surface-layer (S-layer) biogenesis is also largely unknown but involves the post-translational cleavage of a single polypeptide  into low- and high-molecular-weight subunits by Cwp84, a surface-located cysteine protease. Here, the first crystal structure of the surface protein Cwp84 is described at 1.4 Å resolution and the key structural components are identified. The truncated Cwp84 active-site mutant  exhibits three regions: a cleavable propeptide and a cysteine protease domain which exhibits a cathepsin L-like fold followed by a newly identified putative carbohydrate-binding domain with a bound calcium ion, which is referred to here as a lectin-like domain. This study thus provides the first structural insights into Cwp84 and a strong base to elucidate its role in the C. difficile S-layer maturation mechanism. Even so, it has been speculated that the interruption of S-layer biogenesis may make the bacterium more susceptible to antibiotics  and structures based on solution-scattering (SAXS) experiments of both full-length LMW SLP and the complex formed by LMW SLP and HMW SLP  crystal structure of the N-terminal cysteine protease domain of Cwp84. Interestingly, the hitherto uncharacterized 170-residue ‘linker’ region between the cysteine protease domain and putative location of the first Pfam 04122 repeat exhibits a lectin-like domain structure with a bound calcium ion.To further the understanding of 2.2.1.C. difficile Cwp84 residues 33–497  with a C116A mutation  was cloned by PCR into the GST expression vector pGEX-6P-1. The mutation was introduced to potentially circumvent problems with poor expression and degradation or problems with purification . Of the two constructs produced with the mutation, neither had the problems discussed above and one was purified to near-homogeneity in one step (see below). The structure presented in this manuscript made use of this particular construct.A synthetically synthesized gene encoding BamHI at the 5′ end and NotI at the 3′ end preceded by a TAA stop codon . The PCR products were digested with BamHI and NotI (NEB) and cloned into pGEX-6P-1 using T4 DNA ligase (New England Biolabs) to generate pGEX-6P-1-Cwp8433–497C116A.The gene was amplified from the stock pMA vector by PCR with Expand High Fidelity polymerase (Roche) utilizing primers incorporating cleavage sites for E. coli BL21*(DE3) cells. Cultures were grown from glycerol stocks in 5 ml LB supplemented with 100 µg ml−1 ampicillin for 17 h and centrifuged . The cell pellets were washed with water, centrifuged a second time, resuspended in water and used to inoculate 500 ml selenomethionine medium (Molecular Dimensions) supplemented with 100 µg ml−1 ampicillin. These cultures were grown with shaking  to an OD600 of 0.7. The temperature was reduced to 16°C and methionine production was inhibited by the addition of 100 µg ml−1 lysine, phenylalanine and threonine and 50 µg ml−1 leucine, isoleucine and valine. 60 µg ml−1 selenomethionine was also added and the cultures were incubated for 15 min before expression was induced with 1 mM IPTG. The cultures were incubated for a further 18 h and harvested by centrifugation .The plasmid was transformed into M NaCl, 2.7 mM KCl, 5 mM DTT, 10 mM Na2HPO4, 1.8 mM KH2PO4 pH 7.3), lysed in a French press and clarified by centrifugation . The supernatant was loaded onto a GSTrap column  and washed with PBS, and tagged protein was eluted with 10 mM glutathione, 50 mM Tris–HCl pH 8.0. PreScission protease (80 µl) was added and the eluted protein was dialyzed overnight into cleavage buffer . The dialyzed sample was then reloaded onto the GSTrap column to separate the unbound protein from the tag.The cell pellets were resuspended in PBS (140 mM Tris–HCl pH 8.0 (using a Superdex 200 16/600 column); fractions containing Cwp8433–497C116A were pooled and concentrated to 11.9 mg ml−1.Unbound protein was concentrated to a volume of roughly 1 ml and further purified by size-exclusion chromatography (SEC) into 50 m2.2.33–497 was incubated with trypsin at a molar ratio of approximately 10:1 for 45 min. Following purification by SEC in 25 mM MOPS pH 7.0, the resulting single species (Cwp8492–497) was analysed by electrospray ionization mass spectrometry. Cwp8492–497 was also transferred onto PVDF and sent for N-terminal sequencing (AltaBioscience).GST-Cwp842.3.M ammonium sulfate, 30% PEG 4K; Molecular Dimensions Structure Screen 1 & 2, solution D7). Crystals took between 3 d and a week to grow.Crystallization-condition screening was performed with a range of pre-prepared 96-well screens (Molecular Dimensions) using an Art Robbins Phoenix nanodispensing robot. Optimal conditions were reproduced with 0.3 µl drops with a 1:1 ratio of protein to reservoir solution , CRUNCH2 . The diffraction data were recorded with 1.0° oscillation on a Pilatus 6M detector from four crystals to obtain maximum redundancy. Selenium-fluorescence peak and inflection data were collected from all four crystals (to a maximum resolution of 1.73–1.87 Å), while high and low remote data were collected from two crystals (to a maximum resolution of 1.94–2.16 Å). 1120 peak images were collected at 12 660 eV, 1120 inflection images at 12 656 eV, 540 low-remote images at 12 550 eV and 540 high-remote images at 12770 eV. The data were automatically indexed and integrated with http://www.pdb.org) under PDB accession code 4ci7.The atomic coordinates and structure-factor amplitudes have been deposited with the RCSB Protein Data Bank .The high-resolution structure was solved in the monoclinic space group 3.2.B but not in chain A: our secondary-structure numbering assumes that this helix is not present.The propeptide largely consists of loop regions with a central helix (α1) and short β-strand (β1). The poorly defined region was determined to contain a short helix in chain b and 1c) and does not exhibit similarities to propeptides from other papain proteases, which commonly form a small globular domain covering the top of the active site and are stabilized by a β-sheet formed by interaction with the prosegment binding loop . This novel conformation leaves the S′ end of the active-site groove .The N-terminal portion of the Cwp84 propeptide (His33–Gly65) wraps around the lectin-like domain  that precedes this loop is considerably removed from the active site, around 7–8 Å away from its location in both cathepsin L and cathepsin B . Residues Asn64–Ile67 form a hydrogen-bond network with the cysteine protease domain. These interactions are mainly with Met160–Ser164, but hydrogen bonds are also formed to Asn114 and Leu260. After this, the propeptide enters the active-site groove, with Pro70–Glu72 forming hydrogen bonds to the N-terminal part of the propeptide. Thr76–Arg79 form a large number of hydrogen bonds to the lectin-like domain and the cysteine protease domain. Close interactions between the propeptide and the cysteine protease domain are seen in many other proteins  forms an extended loop that sits in the active-site cleft. The poorly defined helix . Given the apparent lack of PBL stabilization and the shorter Val66, the propeptide is likely to be stabilized through other multi-domain interactions.There are usually two main points on a cysteine protease to which its propeptide is anchored: the surface-exposed PBL (prosegment-binding loop), which the propeptide of Cwp84 does not approach, and the Set Fig. 3d. Given33–497 protein (78.5 kDa) with trypsin was found to result in the loss of approximately 33.5 kDa, giving a single band of 45 kDa. The mass of this protein, as confirmed by mass-spectrometric analysis, was 45 058 Da, and therefore the loss of 33.5 kDa from the protein is consistent with removal of the proregion and GST. The N-terminal sequencing determined that the remaining 45 kDa protein had an N-terminus of SSVAY, confirming that the proregion up to Ser92 had been removed. These data suggest that the proregion is folded in Cwp8433–497 in such a way that it is accessible for cleavage by trypsin and that artificial maturation has replicated the removal of the proregion up to Ser92 as observed in C. difficile , one helix (α5) and several loop regions, and the other formed by a central 15-residue-long α-helix (α2) surrounded by two short α-helices (α3 and α4), an antiparallel β-sheet containing two strands (β3 and β9) and several loop regions Fig. 2.et al., 2003The active site of the cysteine protease domain of Cwp84 is similar to those of other cysteine proteases with regard to the positions of the active-site residues Cys116 , His262 and Gln110. Asn287, which has previously been suggested to be an active-site residue  consisting of 13 β-strands (β10–β22), eight of which form a twisted antiparallel β-sandwich with a hydrophobic core. Proteins with similar folds to this domain were determined using a The lectin-like domain contains a hydrophobic core that opens at the surface of the protein, producing a hydrophobic pocket formed by residues Ile347, Ile468, Ile477 and Phe483. Interestingly, both Leu36 and Val39 from the propeptide insert into this pocket, with Lys34 hydrogen bonding to Thr479, suggesting that these interactions may provide stabilizing roles through hydrophobic interactions.BLASTP results  are both sequentially and spatially close to the calcium ion-binding site .4.C. difficile which plays a key role in the maturation of the S-layer protein SlpA. The high-resolution structural data presented here will improve the understanding of the role of Cwp84 in S-layer biogenesis. In addition, the discovery of a newly identified calcium-binding lectin-like (putative carbohydrate-binding) domain raises exciting possibilities with regard to the potential role(s) that this region may have in S-layer biogenesis in C. difficile and also in other species, such as those presented in Fig. 2C. difficile . This loop is poorly conserved among closely related proteins . Indeed, mutation of the P2 lysine to alanine has been shown to abolish the cleavage of an SlpA fragment by Cwp84 in co-expression studies, suggesting its significance in SlpA cleavage  has a fold almost identical to that of sialidases, but the carbohydrate-binding region is replaced by a region that interacts with a helix from IFT27 to form the IFT25/27 complex (Bhogaraju et al., 2011The carbohydrate-binding region seen in many of the We believe the close interactions between the cysteine protease domain, the lectin-like domain and the propeptide are likely to be essential to the initial folding of the protein and will mediate substrate binding and specificity.5.We have determined the structure of the Cwp84 cysteine protease domain with its bound propeptide and a newly discovered lectin-like domain. The propeptide sits in the active-site groove and wraps itself around the lectin-like domain, closely interacting with both domains, a feature that is likely to be important in the initial folding of the protein. The cysteine protease domain, although similar to many previously determined cathepsin L-like structures, bears significant differences; namely, the active-site groove is deepened by the lectin-like domain, the PBL is not present and the would-be occluding loop is slightly longer. The lectin-like domain bears a similar β-sandwich fold to that seen in many carbohydrate-binding proteins, but it is currently unclear what function it possesses. If it does bind a carbohydrate, it is possible that the lectin-like domain may be involved in substrate recognition or attachment to the cell wall, resulting in correct orientation of the cysteine protease domain for cleavage of SlpA.C. difficile surface in growth and colonization, the potential development of anti-colonization inhibitors or vaccines is significantly aided by structural data such as that presented here.Further structural and functional studies are necessary to elucidate the exact mechanism of Cwp84-mediated SlpA cleavage and how this contributes to overall S-layer biosynthesis. Given the likely key role of the Cwp84, 4ci7PDB reference:


High density magnetic domain wall gratings are imprinted in ferromagnetic-antiferromagnetic thin films by local ion irradiation by which alternating head-to-tail-to-head-to-tail and head-to-head-to-tail-to-tail spatially overlapping domain wall networks are formed. Unique magnetic domain processes result from the interaction of anchored domain walls. Non-linear magnetization response is introduced by the laterally distributed magnetic anisotropy phases. The locally varying magnetic charge distribution gives rise to localized and guided magnetization spin-wave modes directly constrained by the narrow domain wall cores. The exchange coupled multiphase material structure leads to unprecedented static and locally modified dynamic magnetic material properties. Magnetic domains are generated in order to reduce the overall free energy, mainly the magneto-static energy, in magnetic systems1146912To intentionally tailor the static and dynamic magnetization behavior of extended unpatterned magnetic thin films by domain walls, domain walls need to be positioned in a controlled way within a magnetic thin film structure. Artificial domain wall structures can be achieved by strain mediation in multiferroic ferroelectric-ferromagnetic heterostructures incorporating ferromagnetic magnetostrictive layers in a bottom-up approach14171981Fe19/Ir23Mn77 thin films is set during film deposition in the presence of a magnetic in-plane field and an orthogonal alignment of exchange bias is obtained by subsequent light He-ion irradiation. A magnetization modulated landscape from domain walls is attained by varying the exchange bias in thin films in a stripe-like fashion with different orientation  responses from non-irradiated and irradiated full films with the exchange bias direction oriented at ±/4 ≈ 0.7  for the Hext perpendicular and parallel to the long axis of the stripes are shown in μm switches at Hc ≈ 1.5 kA/m /AMRmax, implying an alignment of magnetization along the magnetic grating at zero magnetic field for the 〈+ + − −〉 configuration. Yet the AMR value is slightly higher and the magnetization is aligned nearly to the stripe axis for the 1 μm stripes. From the AMR response, the tilting of magnetization away from the imprinted ±45° is estimated to be on average ±33° for a stripe width of 6 μm and down to at least ±40° for the 1 μm grating, but the opposite for the 〈+ − + −〉 case.Different modifications of magnetization reversal behavior are obtained for the highly magnetically charged 〈+ + − −〉 configurations as shown in  stripes . The revpes π in . For thepes π in . The cormy across the domain wall and the stripe border reduces. Correspondingly, the mx-component of magnetization  see fres,w in the core of the domain walls at the stripe borders becomes obvious. The mode localization results from the formation of a potential well for the dynamic modes in the inhomogeneous internal magnetic field of the domain wall, similar to the edge regions of nearly saturated magnetic stripesfres,b, clearly located in the domain wall tail region in the center of the individual stripes, is significantly lower. An analysis similar to fres,b to the overall dynamic behavior fres,w due to the limited volume of the constricted domain walls. Thus, in agreement with the experiments, the contributions from the domain walls increase and dominate for high domain wall densities. A conclusive comparison of the exhibited frequencies of the modes is displayed in Clarity on the bi-modal dynamic behavior is obtained from direct comparison of dynamic inductive and modeling data. Agreement of simulation and experiments over the whole frequency range is demonstrated in e widths . Both moArtificial Néel domain walls with different magnetostatic contributions can be generated by anisotropy patterning in exchange biased thin films. Longitudinal modulated head-to-tail-to-head-to-tail 〈+ − + −〉 and magnetically charged transversal head-to-head-to-tail-to-tail 〈+ + − −〉 domain structures are formed. The altered magnetic structure results in magnetic domain wall dominated magnetization behavior for high packing densities. The overlapping domain wall structures give rise to reorientation of the magnetization away from the imprinted magnetic anisotropy direction and to formation of magnetic superdomains. The nucleation and annihilation of the dense domain wall structures result in highly non-linear magnetization processes. By varying the domain wall density, the domain wall properties can be probed by integral magnetic measurement techniques. Magnetization reversal and magnetoresistive response are tailored with the inclusion of domain wall networks.δc,w for soft magnetic materials is typically below the magnetic film thickness, corresponding to δc,w ≤ 50 nm in our case. Yet due to magneto-static energy contributions long-range magnetic tail structures with a tail width δt,w extending over several micrometers exist. Estimating the effective domain wall core width δFWHM,w from the full width at half maximum (FWHM) of the obtained Néel wall profiles from micromagnetic simulations leads to an effective width of between δFWHM,w ≈ 105 nm and δFWHM,w ≈ 150 nm for a stripe width of 500 nm and 6 μm /Ir23Mn77(7 nm)/TaN(5 nm) thin films were deposited by DC magnetron sputtering under UHV conditions with a base pressure below 2 × 10−8 Torr by using 6N Ar at a pressure of 10−3 Torr on oxidized Silicon substrates. An initial unidirectional anisotropy in the layer stack was set during the sputtering process by a homogeneous magnetic field of 4 kA/m. No post annealing steps were performed. The Ta buffer layer acts as a seed layer for the Ni81Fe19 to guarantee a well-defined 〈111〉 texture. The top TaN layer prevents corrosion of the thin film stack. The sample size is (1 cm × 1 cm), allowing for complementary integral as well as laterally resolved local analysis.Extended Ta(5 nm)/Niπ/4 or −π/4 to the initial exchange bias direction was introduced. The stripe width was varied from 6 μm down to 500 nm, being well below the distance of the extended magnetic Néel domain wall tails15 ions/cm2 was performed by which a reorientation of exchange bias in accordance with the state of magnetization in the non-masked regions was achieved. To alter the initial unidirectional anisotropy in the ion irradiated areas, an external in-plane magnetic saturation field (Hsat = 26 kA/m) was applied during irradiation. The field was oriented π/2 to the initial exchange bias direction. By doing this, artificial magnetic domain patterns were introduced, resulting in stripe-like two-dimensional structures with alternating directions of exchange biasThrough photolithographic masking, photoresist stripes aligned either at +m||Quasi-static magnetization curves were obtained by inductive magnetometry carried out 1 cm × 1 cm at a frequency of 10 Hz. From this the component of magnetization parallel to the applied magnetic field was obtained. m⊥ relative to the applied magnetic field was derived. The AMR amplitude was calibrated against the AMR response of stripe structures measured in a linear four probe setup on structured elongated stripe structures. The magnetic field resolution ΔH for the magnetometry and the AMR measurement is better than ΔH = 30 A/m. Laterally resolved information of the magnetization reversal was studied by high resolution magneto-optical Kerr effect microscopy in the longitudinal modeby which information about the transverse magnetization component Hpulse ≈ 1.5 A/m and a rise time of trt,10−90 ≤ 50 ps. The measurements were performed with varying bias field Hext. The time domain data was transferred into the frequency domain by fast fourier transformation, from which the dynamic magnetic permeability spectra were obtained.The dynamic magnetic characteristics, e.g. magnetic permeability spectra, were obtained by pulsed inductive microwave magnetometry (PIMM)mi of the ith discretization cell in the formIn order to resolve the different behavior of the 〈+ − + −〉 and the 〈+ + − −〉 domain wall phases, the dynamics of the two configurations were calculated by micromagnetic simulationsγ being the gyromagnetic ratio and α the magnetic damping parameter of the material. No thermal effects were taken into account for the simulations. The in-plane cell size for the calculations ranged from 1.25 nm for the narrow stripe width of 500 nm to 15 nm for the wide stripe width of 6 μm. The vertical dimension of the cell was the film thickness. A saturation magnetization of μ0Ms = 1 T and a magnetic damping parameter α = 0.0181Fe19 layer in the calculations. The exchange stiffness constant was set to A = 1.05 × 10−11 J/mwith How to cite this article: Trützschler, J. et al. Magnetic domain wall gratings for magnetization reversal tuning and confined dynamic mode localization. Sci. Rep.6, 30761; doi: 10.1038/srep30761 (2016).


Nucleotide-binding oligomerization domain (NOD)-like receptors (NLRs) are intracellular receptors that control innate immunity and other biotic interactions in animals and plants. NLRs have been characterized in plant and animal lineages, but in fungi, this gene family has not been systematically described. There is however previous indications of the involvement of NLR-like genes in nonself recognition and programmed cell death in fungi. We have analyzed 198 fungal genomes for the presence of NLRs and have annotated a total of 5,616 NLR candidates. We describe their phylogenetic distribution, domain organization, and evolution. Fungal NLRs are characterized by a great diversity of domain organizations, suggesting frequently occurring combinatorial assortments of different effector, NOD and repeat domains. The repeat domains are of the WD, ANK, and TPR type; no LRR motifs were found. As previously documented for WD-repeat domains of fungal NLRs, TPR, and ANK repeats evolve under positive selection and show highly conserved repeats and repeat length polymorphism, suggesting the possibility of concerted evolution of these repeats. We identify novel effector domains not previously found associated with NLRs, whereas others are related to effector domains of plant or animals NLRs. In particular, we show that the HET domain found in fungal NLRs may be related to Toll/interleukin-1 receptor domains found in animal and plant immune receptors. This description of fungal NLR repertoires reveals both similarities and differences with plant and animals NLR collections, highlights the importance of domain reassortment and repeat evolution and provides a novel entry point to explore the evolution of NLRs in eukaryotes. Detection and adequate response to nonself is essential for survival and development in all multicellular organisms. An important part of the innate immune detection in plants and animal lineages is ensured by a class of signal transducing proteins known as NB-LRR proteins in plants and nucleotide-binding oligomerization domain (NOD)-like receptors (NLRs) in animals .Plant NB-LRR proteins sense the presence of fungal, oomycete, nematode, bacterial, or viral pathogens and trigger an immune response in the form of a localized cell death reaction termed the hypersensitive response . NB-LRR Hydra and the coral Acropora digitifera  and at least in mammals, the number of NLRs is more limited than in plant genomes . Animal gitifera . The N-tgitifera , whereasgitifera . In addigitifera . In spitgitifera . It has gitifera .Podospora anserina, one of the founding members defining the NACHT domain, is involved in a fungal nonself recognition and programmed cell death process termed heterokaryon incompatibility  family is, in addition, characterized by a remarkable diversification mechanism based on alternative splicing of multiple codon-sized microexons , and then combined. The candidate set was pruned from sequences with multiple disjoint matches to the queries and from very short sequences (below 100 amino acids). Then it was limited to sequences from complete or draft whole-genome sequencing and resequencing projects, according to Genome OnLine Database  queries were obtained by extraction of NACHT and NB-ARC domains from the full-length sequences according to PfamA PF05729.7 and PF00931.17 profile matches . PSI-BLAatabase  to builE value was chosen, except for the P-loop NTP-ase clan (CL0023), where NACHT or NB-ARC annotations were always preferred (if above the PfamA threshold). Annotations from repeat-containing clans: Ankyrin (CL0465), Beta propeller (CL0186), and TPR  were merged to three main categories: ANK, WD40, and TPR, respectively. Highly overlapping N-terminal annotations, as well as three prion-forming domain annotations were merged (see Results). Conflicting annotations from PfamA HeLo and in-house HeLo-like signatures were resolved in the favor of the former. Numerical suffixes of signature names were truncated and sequential occurrences of identical annotations were squeezed. Domain associations were visualized using the graphviz package  in which a given architecture was found. Paralog index summed the number of sequences with a given architecture in all species .All phylogenetic trees were calculated through the maximum likelihood estimation based on alignments of NB or N-terminal domains extracted from nonidentical sequences. In each case, the best alignment was selected according to the norMD score out of alignments generated by the same MSA tools as above. Then, the alignment was pruned from columns with more than 50% of gaps (using trimAl ) and subGenes with no clear ortholog in all (“orphans”) or in some other strains (“semiorphans”) from the same species were identified according to co-phenetic distances between leaves in multistrain phylogenetic trees (using the R package ape). To detect highly homologous pairs of NB domain sequences associated with different N-terminal domains, BLASTP scores  were calHighly internally conserved repeats were detected using T-reks  with cusP. anserina ANK and TPR motifs, genes-encoding STAND proteins with individual repeats displaying over 85% internal conservation were analyzed, excluding hnwd family members, resulting in a set of ten genes. They code for TPR, ankyrin, or HEAT repeats. For each gene, the repeat-encoding DNA was polymerase chain reaction (PCR)-amplified from five wild isolates from the Wageningen collection   , gel-purProtein repeats were identified using RADAR . IndividP. anserina HET-E, HET-D, and HET-R incompatibility genes and the fungal STAND proteins comprising a putative prion-forming domain , comprised an ensemble of STAND proteins with NACHT and NB-ARC Pfam-A annotations with a large phylogenetic distribution, ranging from bacteria to plants and animals and included sequences from different major lineages . The NB-ARC and NACHT sequences were extracted from the different query set and used in PSI-BLAST searches with three iterations and an E value cut-off of 10−5 on the complete annotated genome sequences of 198 strains of 164 fungal species . The IR, FV, and PD query sets recovered 5,571, 1,053, and 4,657 hits, respectively . The IR recovered the most hits, whereas the FV set led to the lowest number of hits, but FV hits were almost entirely included in the IR and PD sets. The FV query did recover only a very limited number of NACHT domain STAND proteins, but was more efficient in the identification of NB-ARC STAND proteins . We included all hits in our candidate set, which thus adds up to 5,616 sequences  corresponding to 4,476 (79.7%) and 1,144 (20.4%) NACHT and NB-ARC hits, respectively (four sequences were hit by both NACHT and NB-ARC queries). Hits were found in 122(101) of the 198(164) strains (species). In these 122 strains, there is mean number of STANDs per genome of 46, with a median of 37.To identify NLR-like proteins in the different complete fungal genomes, we have used NACHT and NB-ARC NOD domains from previously identified STAND proteins as queries. We have defined three different query sets. The first set comprised a list of fungal STAND proteins previously identified in the context of the study of fungal incompatibility. This query set we termed IR (incompatibility-related) includes the g domain . A seconsupplementary fig. S2, Supplementary Material online). The NACHT to NB-ARC ratio is 5:1. This contrasts with the situation observed in viridiplantae, where NB-ARC largely predominates (NACHT to NB-ARC ratio based on Pfam annotations is 1:180). In bacterial STAND proteins, both types are common; however, NB-ARC domains are also more abundant than NACHT domains (approximately in a 2:1 ratio). The higher occurrence of NACHT versus NB-ARC makes the fungal NLR candidate set more animal-like, because in metazoans NACHT domains are more frequent than NB-ARC (in a 17:1 ratio). A remaining 28% of the NOD regions picked up in the BLAST searches were neither annotated as NACHT nor NB-ARC by Pfam. Among the candidates were also a number of sequences showing noncanonical P-loop motifs with recurrent variations around the canonical GXXXXGKT/S motif , a situation also described in plant NLRs  and were not analyzed further. We end up this way with 12 annotation categories for the N-terminal domains . The PNP_UDP domain has been previously identified as an N-terminal effector domain in NLR proteins from the coral A. digitifera .NLRs have a typical tripartite domain organization, with a central NOD flanked N-terminally by an effector domain and C-terminally by an autoinhibitory/ligand-binding domain, often composed of superstructure-forming repeats . For ann domains . Among tgitifera , and a sgitifera . Globallsupplementary fig. S2, Supplementary Material online). In ascomycota, ANK repeats were more abundant whereas WD40 repeats prevailed in basidiomycota. No LRR motifs were found in agreement with a previous study  have also been found in NLRs from other phyla. A large fraction (roughly 50%) of the N-terminal and C-terminal domains do not respond to known annotations.supplementary fig. S3, Supplementary Material online). These preferential association trends always suffer exceptions, as a small fraction of the NB-ARC domains are associated with ANK or WD, and a small fraction of the NACHTs is followed by TPRs. The fact that in our sequence set some domain architectures are encountered only once suggests that some of the missing architectures might be identified by analyzing additional species.Next, we analyzed the domain architectures of the fungal NLR candidate set. Globally, there is a great diversity of domain architectures. To illustrate this aspect, we focused our analysis on the 1,228 sequences for which all three domains  have an annotation. The 12 annotated effector domains and NACHT and NB-ARC NOD domains can in principle lead to 24 (12 × 2) domain associations, and of those, 21 occur in our candidate set. Similarly, all six combinations of NACHT and NB-ARC with WD, TPR, and ANK motifs are found in the set. Globally, of the 72 possible tripartite domain architectures (12 effector domains × 2 NOD domains × 3 repeat domain), 32 are actually found in the set . In genesupplementary fig. S4, Supplementary Material online). Then, when generating phylogenetic trees from the NLR complement from a given species based on the NOD sequences, domain architectures based on N-terminal domains do no form monophyletic groups but rather are to some extent scattered in different branches of the tree. For instance, in the phylogenetic tree based on the NOD domain of the NLR complement of the species Bipolaris maydis, the HET domain is found in different branches of the tree. The same is true for PNP_UDP, Goodbye, and HeLo-like domains . This distribution and the observed combinatorial domain association suggest that de novo generation of specific domain architectures can occur by domain fusion events between N-terminal domains and a different lineage of NODs. In order to explore this aspect, we analyzed our NLR candidate set for situations in which a given NOD is highly similar to a NOD embedded in a distinct domain architecture. When inspecting the distribution of annotated N-terminal domains in phylogenetic trees based on the NOD domains, it appears that phylogeny of the N-terminal domains is frequently distinct from that of the NODs. This is apparent in two ways. First, in the global candidate set, the phylogenic trees based on the N-terminal domains are not congruent with the phylogenies of the NODs , characterized by WD-repeats showing a high level of internal repeat conservation, meaning that the individual WD-repeats of a given gene are highly similar to each other (with about 85% identity at the amino acid level) (A). These observations indicate that the internal repeat conservation noted for WD repeats in P. anserina is a common property of a significant proportion of the NLR-like proteins and that this phenomenon is also encountered with ANK and TPR motifs both in ascomycetes and basidiomycetes. We have analyzed the occurrence of such highly conserved repeats in ANK, TPR, and WD-type repeats in plants, metazoan, and fungi . We found that the fraction of repeats with high internal conservation is globally very low . There is thus a specific enrichment for highly conserved repeats in fungal NLR proteins. In dikarya, occurrence of highly conserved ANK, TPR, and WD repeats occurs mainly in NLR-like proteins, which globally account for 60-70% of the occurrence of highly conserved repeats. We conclude that highly conserved ANK, TPR, and WD repeats are highly enriched in fungal NLRs, as compared with their global occurrence.The analysis of STAND protein evolution in d level) . This ind level) . To deted level) A. These B). This difference might be related to the fact that ANK and TPR motifs form open-ended superstructures (fig. 4C).The distribution of the number of repeats per gene was different in ANK and TPR, compared with WD repeats. There was a gradual decrease in the class size with increasing number of repeats per protein in the case of ANK and TPR, while in the case of WD, class sizes were relatively constant from 1 to 14 repeats but then dropped sharply above 14 repeats .Next, we analyzed whether or not highly conserved repeats are randomly associated with the different N-terminal effector domains. All frequent N-terminal domains can be found associated with highly conserved repeats, but it appears that certain N-terminal domains are preferentially associated with highly conserved repeats, as for instance the HET domain but also the prion-forming domains, whereas others like the Goodbye domains are very seldom associated with this type of repeats . NLRs were absent from certain lineages; in particular, no hits were found in any of the 38 analyzed Saccharomycotina genomes, or in the Schizosaccharomycetes. Similarly, we found no hits in early branching lineages of the microsporidia, chytrids, and mucorales. In contrast, hits were abundant in major basidiomycetes  and ascomycetes lineages . When comparing the annotation of the ascomycota and basidiomycota ensembles, three main trends are apparent. The ratio of NACHT to NB-ARC is slightly different in both lineages, with NB-ARC being rarer in basidiomycota . The abundance of the different types of repeat motifs also differs in both lineages: WD, ANK, and TPR account for 27, 9 and 8% of the C-terminal domain annotations in basidiomycota, compared with 10, 27, and 14% in ascomycota. The higher abundance of NB-ARC and TPR motifs in ascomycotina is expected, considering the preferential association of NB-ARC with TPR motifs . The level of annotation of the N-terminal domains is very different in both lineages, with only 13% of the sequences receiving an annotation in the basidiomycota, compared with 63% for the ascomycota. This difference is probably related to the fact that our in-house annotations derive from ascomycete sequences.Next, we analyzed the phylogenetic distribution of NLRs in fungi . The onygenales group containing several dermatophytes shows less hits than the rest of the pezizomycotina .Variation in the number of NLRs per genome is extreme, ranging from 1 (or 0) to 274 in the endophytic basidiomycetes species Aspergillus genus. Some domains show a strong tendency for marked expansions, while other are usually found as a single occurrence. We calculated a paralog-to-ortholog index, corresponding to the ratio of number of occurrences of the domain to the number of species in which the domain is encountered. The domains showing the highest number of occurrences per species were PNP_UDP and Goodbye, with a mean occurrence of 7.5 and 7.4 per species, respectively, while in contrast HeLo and Patatin domains showed the lowest occurrence  . These two domains are most generally found as one or two occurrences per species, but some rare exceptions of marked expansion occur as for instance for the HeLo domain in the fusaria.We compared the occurrence of the 12 different N-terminal domains in the different species and there again the diversity between species is considerable. None of the 12 annotated domains has a universal distribution in all species displaying NLRs but some are found in a large fraction of species like the Goodbye-like, HeLo-like, and sesB-like domains found in NLRs of 88, 73, and 75 species, respectively. Other domains are found in a narrow species range, like the C2 domain found only in a few basidiomycota. As already noted for the total NLRs numbers, there is a high variability in the number of domain occurrences, even for closely related species, with for instance the number of PNP_UDP NLRs ranging from 2 to 23 in different species of the Laccaria bicolor. 72 strains, among the 122 displaying NLRs proteins, have at least one gene with internally conserved WD, TPR, or ANK repeats . Species in which such NLR-like proteins with high conserved repeats are particularly abundant are L. bicolor, B. maydis, and Talaromyces stipitatus.When considering the C-terminal repeat domains, the fraction of repeats with high internal conservation varies dramatically between species from 0 to up to 58% in HSP90 and its co-chaperones SGT1 and RAR1 play important roles in NLR function both in plants and animals . We analsupplementary fig. S6, Supplementary Material online). In all 15 analyzed species, some level of polymorphism in the NLR complement is observed. A variable fraction of the NLR sequences lack a clear ortholog in the other analyzed strain(s). Penicillium digitatum to 100% in Rhizoctonia solani). In addition, in many species, a significant proportion of the NLR candidates do not have a conserved ortholog in the other strain(s) . For instance, in Aspergillus niger CBS 513.88, ten sequence show no ortholog in the other strain (ATCC 1015) with a cut-off distance value of 1, which corresponds to about 50% identity. Inspection of the phylogenetic trees reveals the existence of numerous such orphans as well as strain-specific expansion in certain branches of the tree, arguing for a birth-and-death evolution of these sequences . This strain-specific expansion is for instance evident Fusarium oxysporum Fo5176.Previous reports suggest that fungal STAND proteins show high level of intraspecific variation . In addihet-e, het-d, and het-r incompatibility genes. Functional studies have identified this domain as being a cell death and incompatibility effector domain in P. anserina (The HET domain acquired this designation because it was found in different proteins involved in nonself recognition in the form of heterokaryon incompatibility in fungi . In partanserina . We now Paracoccus denitrificans (Brucella melitensis (We first conducted PSI-BLAST searches in the nr (“nonredundant”) database with the HET-e1 HET domain by excluding fungal sequences and found that homologs of this domain are also found outside of the fungal kingdom in Stramenopiles, Haptophyceae, Choanoflagellates, green algae, and bryophytes. Next, we used Hidden Markov model searches to identify remote homologs of the HET-e1 HET domain. Both algorithms that we used (HHpred and JackHHmer) identified similarity to TIR domains. In particular, the two best hits in HHpred were to structure-based profiles constructed from the TIR domain of PdTIR from rificans  and of tlitensis . Figure litensis . This poPodospora and are subject to concerted evolution and positive diversifying selection   . ANK repsm (RNP) .Table 4Next, we selected one ANK repeat locus and one TPR motif locus for which we had sequenced the highest number of repeats  and analysed the variability of the repeats from individual loci. For each locus, individual repeat sequences were aligned and analysed for position under positive selection  . Five poChaetomium globosum (PNP_UDP/NACHT/ANK) and gi255934897 from Penicillium chrysogenum (UNK/AAA/TPR), with, respectively, 21 ANK repeats and 21 TPR motifs. By comparing the repeats and mapping the variable positions onto a homology model (PDB ID 4GPM for ANK and 3EDT for TPR), we found that polymorphisms map to the same positions in the α-helices of the concave surface of the TPR domain and to the inner helices and β-hairpin/loop region of the concave interface of the ANK domain . Based on the localization of these polymorphic sites, it can be inferred that if repeat contraction/expansion/shuffling occurs in these genes, these events will lead to ANK and TPR arrays with modified binding interfaces.We also analysed two putative proteins from different species to determine whether this localization of the polymorphisms might be common to other ANK and TPR motifs. We chose the ANK and TPR proteins with the highest number of highly conserved ANK and TPR motifs, gi116208038 from Podospora NLR candidates is analogous to the evolution of highly conserved WD repeats of NLR-like proteins, which has been described previously . This can only be made with extreme caution, because in many cases, the information available on the species ecology is at best fragmentary and many species have multiple habitats and life-styles. In some groups, there is a significant enrichment or scarcity of NLRs. For instance, animal dermatophytes of the onygenales have in general few NLR genes. But it is difficult to determine whether this is related to the phylogenetic position or to ecology. If the function of NLR homologs in fungi is related to innate immunity, the prediction might be that fungi potentially in relation to diverse pathogens or competitors or hosts should be particularly enriched in terms of NLR repertoire, and reciprocally, that in fungi living in less populated niches, smaller repertoires could be sufficient. This prediction might be verified in some instances as, for example, in the case of the highly versatile pathogens like Fusarium species or mycoparasitic Trichoderma species, in which the repertoire is large. In the thermophile Chaetomium thermophilum, the citrus fruit pathogen Pe. digitatum or the “whisky fungus” Baudonia compniacensis have small repertoires and inhabit restrictive niches. Similarly, specialized pathogens, such as Claviceps purpurea, might be protected against microbial competitors by their host immune system, which could explain the low number of NLRs.Within the filamentous agaricomycotina and pezizomycotina, the number of NLR homologs varies dramatically between species. One may attend to establish a relationship between the species ecology and the constitution of the NLR homolog repertoire .