In order to exploit full potential of materials, it is necessary to fundamentally control their microstructures through grain refinement and orientation. Deformation and recrystallization are means to control the microstructure. Some recent examples of research on deformation and recrystallization which make use of advanced analytical techniques and computational materials science are examined and current limitations are identified. Finally, the potential for future developments is considered with respect to the unresolved technical problems that must be addressed as part of the development of new steels.  Therefore, further research is required to increase their strength ,2. At thThe controls of deformation and recrystallization are important as a way to refine grains but also as a means to control the crystal orientation . For example, texture controls are used in automotive panels as well as the motors of hybrid or electric vehicles, and they contribute to the high performance of the final products.The thermo-mechanical controlled process (TMCP) is the basic concept underlying grain refinement. A combination of controlled rolling and controlled cooling is used to produce refined α grains through (1) γ recrystallization/non-recrystallization and (2) γ to α phase transformation. Since the recrystallization and phase transformation occur heterogeneously, local variations in strain and orientation are essential. The basic method for controlling the texture is as follows. Based on the generally accepted orientated nucleation and selected growth theory, both the heterogeneous deformation structure and deformed specific structure are crucial. The heterogeneous deformation structure is required for the recrystallized grains to nucleate with a preferred orientation and the deformed specific structure is required for the nuclei to grow in a selective manner. Consequently, it is necessary to understand and control the heterogeneity of the deformation structure in order to achieve grain refinement and control the texture. However, due to their complexity the technology required to evaluate and predict heterogeneous deformation structures has not yet been fully established. Therefore, rigorous microstructure controls exploiting cutting-edge technologies are anticipated.In this overview, recent research into deformation and recrystallization using advanced novel techniques and computational materials science are introduced. In addition, potential future developments are discussed.2.2.1.The deformation structure of polycrystal metallic materials after rolling is extremely complicated. In particular, the area surrounding the initial grain boundary is heterogeneously deformed due to the constraints from neighboring grains. In addition, the heterogeneous structure in the grain interior includes microbands, shear bands, and transition bands depending on the initial orientation, grain size, chemical composition, reduction rate etc. Moreover, a locally heterogeneous structure develops at the interface between the matrix and the hard second phase. Thus, attention should be given to heterogeneous structures such as the initial grain boundary, shear bands, and the interface between the soft matrix and the hard second phase.2.1.1.Abe at el. investigated heterogeneity in the deformation structure focusing on the initial grain boundary in pure polycrystalline Fe after it was cold rolled by 70% . In gene2.1.2.Small local fluctuations in crystal orientation within grains are generally expected after rolling. Furthermore, heterogeneously deformed regions occur within grains where there is immense shear strain; for example, shear bands are associated with significant crystal rotation. Recently another type of shear band is reported by Nguyen-Minh et al.  when matThe transition band parallel to the rolling direction is another example of heterogeneous deformation caused by rolling material with initial {100}<001> orientation . The tra2.1.3.The presence of a hard second phase in the relatively soft matrix generally leads to locally complex severe deformation at the interface. Kimura et al.  preparedpatterns .Figure 52.2.−1, as the first deformation. They were then held at 800°C until the second compression test with ε = 0.3 at the same temperature and strain rate. The αʹ microstructure was observed after quenching specimens immediately before the first and second compression tests. y1, σy2σ, and εσ represent the yield stress during the first and second compression tests, and the flow stress at ε = 0.3 during the first compression test, respectively. Immediately before the first and second compression tests, the samples were quenched and subjected to EBSD observation. As shown in 3 s . The EBn in 3 s c2, is to3 s (4 s ). In add3 s  has revealed the actual state of alloying elements in view of the solute drag effect using B bearing Ti-IF steel ,26. Figu[11ˉ0]. This sh2.4.In the process of producing high strength steel sheets consisting of α and αʹ through cold rolling and subsequent intercritical annealing, both α recrystallization and the α→γ reverse phase transformation take place in a competitive manner. Ogawa et al.  exploredDannoshita et al.  investigWhen 0.02% and 0.05%Nb is added to such a steel, three phenomena occur simultaneously: α recrystallization, α→γ reverse phase transformation, and NbC precipitation . First, 2.5.Rigorous X-ray line profile analysis has made it possible to evaluate the dislocation density, the fraction of screw/edge dislocation components, and the degree of dislocation rearrangement . The app3.3.1.There have been many studies on the prediction of the heterogeneous deformation structure and texture development of poly-crystal materials when they are plastically deformed. However, these predictions are not easy since the deformation is not only affected by the presence of neighboring grains, but also by the presence of a hard second phase. This is because the neighboring grains and hard second phase constrain the deformation of a particular grain. Furthermore, the addition of solute elements affects the slip behavior. In recent years, various approaches have been tested including crystal plasticity (CP)-finite element method (FEM) (CP-FEM) , CP-fast3. The heterogeneous spatial distribution of the stored energy and the misorientation were obtained as a function of the applied strain after the material was deformed to relatively low strain under plane strain conditions. The results predict that the stored energy is heterogeneous and is locally high at certain grain boundaries and that there is large misorientation within a grain. This is an important result in terms of reproducing the three dimensionally heterogeneous deformation structures; therefore, it is expected to have many applications. Yamanaka [Yamanaka  simulateYamanaka  used theYamanaka ,37.3.2.Heterogeneous deformation structures are important as they act as preferential nucleation sites for recrystallization and phase transformation. Research attempting to construct a reliable model that can predict the microstructure and texture evolution from upstream to downstream has been advanced by the coupling model of deformation and recrystallization/phase transformation ,35,38,393.3.In order to understand the evolution of heterogeneous deformation structures, it is necessary to know how solute atoms affect dislocation glide behavior. Waketa et al. systematically investigated the interaction between screw dislocation core and various substitutional atoms using first principle calculations combined with molecular dynamics . They foThe interaction between the dislocation core and solute atoms not only affects the dislocation glide in the early stage of deformation, but also has a significant effect on the collective dislocation motion related to work hardening and dynamic recovery in the later stages of deformation and subsequent plastic instability. Based on this, we anticipate the development of a multi-scale approach that relates atomic-scale calculations to macro-scale properties.4.The recent development of advanced analytical techniques is remarkable, and it is now possible to clearly observe phenomena that were previously invisible. Furthermore, the progress in computational materials science has enabled us to conduct multi-scale calculations and to understand phenomena at a level of detail that used to be un-calculable and speculative. The accurate prediction of phenomena, which cannot be tested experimentally, is also becoming possible and has various implications.The understanding and prediction of heterogeneous deformation structures, both experimentally and theoretically, are still insufficient and further advancement is required.The mechanism underlying orientation selection during recrystallization following deformation must be clarified, especially focusing on the recovery and nucleation of recrystallization.The effect of alloying elements on the dislocation core structure must be fundamentally elucidated in conjunction with the movement of single dislocations and the collective movement of dislocations.The interface energy and mobility should be systematically elucidated together with the effect of alloying elements while taking into consideration the interface character.Finally, advanced analytical techniques should be combined with computational materials science for further materials research in order to solve crucial technical problems, and to realize the profound capabilities of materials.However, many problems remain outstanding:


Detailed knowledge on the formation of biomembrane domains, their structure, composition, and physical characteristics is scarce. Despite its frequently discussed importance in signaling, e.g., in obtaining localized non-homogeneous receptor compositions in the plasma membrane, the nanometer size as well as the dynamic and transient nature of domains impede their experimental characterization. In turn, atomistic molecular dynamics (MD) simulations combine both, high spatial and high temporal resolution. Here, using microsecond atomistic MD simulations, we characterize the spontaneous and unbiased formation of nano-domains in a plasma membrane model containing phosphatidylcholine (POPC), palmitoyl-sphingomyelin (PSM), and cholesterol (Chol) in the presence or absence of the neurotransmitter serotonin at different temperatures. In the ternary mixture, highly ordered and highly disordered domains of similar composition coexist at 303 K. The distinction of domains by lipid acyl chain order gets lost at lower temperatures of 298 and 294 K, suggesting a phase transition at ambient temperature. By comparison of domain ordering and composition, we demonstrate how the domain-specific binding of the neurotransmitter serotonin results in a modified domain lipid composition and a substantial downward shift of the phase transition temperature. Our simulations thus suggest a novel mode of action of neurotransmitters possibly of importance in neuronal signal transmission. In particular the so-called coarse-grained (CG) simulations characterized by condensation of the degrees of freedom by introduction of super-atoms or super-beads enabled the study of spontaneous domain formation in biomembrane models. The popular MARTINI model provided a first molecular view on domain formation of model membranes with fully saturated and polyunsaturated phospholipids as well as cholesterol, with well distinguished ordered  and disordered domains (LD)  domains introduced early for phosphatidylcholine-cholesterol systems  and sterol-enriched domains that compartmentalize cellular processes  exposure to xenobiotics or endogenous molecules. Of particular interest is the interaction of neurotransmitters with membranes. These are released to the synaptic cleft in response to a signal. Their concentration in synaptic vesicles is as high as 270 mM  and 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) for both charged and neutral forms of serotonin (pKa = 9.97 for primary amino group) on the 100 ns timescale. A protonation-dependent binding orientation of serotonin was reported, with the charged amine interacting with the lipid phosphate moiety and the deprotonated serotonin adopting a reversed orientation with the primary amine pointing toward the membrane core. The cationic serotonin form was independently used to investigate the effect of lipid acyl chain unsaturation , 8:61:31 (LO system), and 69:23:8 (LD system) in the absence or presence of 10 mol% serotonin (5-HT). The simulations were performed at a temperature of 303 K  and at 294 and 298 K (LO/D). The lipid compositions for the LO and LD systems were selected according to a FRET-based phase diagram for a brain SM:DOPC:cholesterol composition using the tie line ends  and an aromatic hydroxyl moiety (OH) , with pKa values of 9.97 and 10.73, respectively  analysis, similar to the approach of Park and Im , with means μL, μO and standard deviations σL, σO  are assigned when μL + (i−1)ΔP2 ≤ P2 ≤ μL + iΔP2, where ΔP2 = (μO − μL)/7. For the low and high hidden ordered states, the initial emission probability matrix was set up making use of the integrals of the normal distributions over the subspace of the observables. For the intermediate ordered state, we considered integrals over a normal distribution with variance μI = (σLμO + σOμL)/(σO + σL), and σL = min/3 to ensure that the intermediate states initial emission probabilities are localized within 3σ between the upper, μO, and the lower, μL, bounds , the emission states and the initial transition and emission probabilities were based on the POPC/PSM/Chol 4:4:2 simulation at 303 K.For better comparison of the hidden ordered, disordered, and intermediate states between simulations with/without serotonin and between simulations at different temperatures . Angular brackets denote an average over all sampled configurations. The order parameters were calculated by first averaging over time separately for each lipid in the system and then calculating the average and the standard error of the mean over the different lipids.where θ is the angle between the C-H bond and the membrane normal  and sphingosine backbones (PSM), and serotonin centers of mass along the bilayer normal, F(z) = −kBTlnp(z). p(z) is the probability of finding serotonin at position z, and z is the position along the bilayer normal with respect to the membrane center, T is the absolute temperature, and kB the Boltzmann constant. The probability density p(z) was estimated using a Gaussian mixture model  for the bilayers was calculated as the area of the simulation cell A divided by the number of lipids per leaflet. This assumes that undulations are small so the difference in projected and local areas is negligible. The area compressibility KA is evaluated from:The instantaneous area per lipid A〉 is the average total area, 〈δA2〉 is the mean square fluctuation, kB the Boltzmann constant, and T the temperature. The estimation of the standard error was performed using the block average method  to 150 ns (LO), corresponding to twice the correlation time of the time series fluctuations, estimated from the normalized correlation function of the average total area fluctuations.Where 〈KC, was determined using the method developed by Watson et al.  are observed only over wavelengths shorter than 3 bilayer thicknesses , 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC), N-palmitoyl-D-erythro-sphingosylphosphorylcholine (PSM) of highest quality were purchased from Avanti Polar Lipids . 5-HT was purchased from Merck . All other chemicals were of highest purity with exception of organic solvents which were of spectroscopic grade.2PO4 buffer , freeze thawed and finally extruded through two 100 nm polycarbonate filters at 40° C to make unilamellar vesicles (LUV). Lipid samples were diluted from a LUV stock solution to a concentration of 0.3 mM for the zeta potential measurements. Concentration of 5-HT in the samples varied between 0 and 10 mM. Serotonin was added externally from a K2PO4 buffer . The samples were measured at 25° C after 10 min incubation after externally adding 5-HT. Zeta potential was measured using a Zeta potential analyzer (Brookhaven) with 10 runs and 10 cycles.Lipids were mixed in organic solvents at the molar ratio POPC/PSM/Chol 4/4/2 and evaporated at 40° C in a rotary evaporator. Afterwards the lipids were hydrated at 40° C for 30 min using KLO/D system) in the absence or presence of serotonin at three different temperatures . In addition, for comparison of the domain characteristics and quantitative binding analysis of serotonin, MD simulations of previously in silico characterized LO and LD domain compositions . Similarly, we observe small drifts in the number of lipid-lipid contacts as a measure of the formation of differently composed domains. Here, we thus focus on the initial steps of domain formation in ternary mixtures containing spingomyelin in the presence or absence of the neurotransmitter serotonin, domain composition, and domain influence on the mechanical membrane properties. The simulation system size and simulation length (10 μs) does not allow to conclude on the formation of large domains beyond 10 nm. Also domain compositions and sizes analyzed below may likely further slowly adapt on the 100 μs to millisecond timescale.The spontaneous formation of biomembrane domains was addressed in atomistic MD simulations for a POPC/PSM/Chol mixture  employing P2 order parameter-distinguished emission states (see section 2 for details). For an unambigous assignment and for quantitative comparison of systems with/without serotonin as well as for comparison of simulations at different temperatures, the emission states were defined based on the As a measure of the charge of serotonin bound to phospholipid membranes we determined the zeta potential for a POPC/PSM/Chol (4:4:2) mixture in the absence or presence of varying concentrations of serotonin. The zeta potential see  was hardLO/D simulation (POPC/PSM/Chol 4:4:2 mixture). The HMM analysis revealed spontaneously formed ordered, sphingomyelin-enriched domains  is observed to be enriched in POPC (52%). That is, the sphingomyelin-enriched LO domains are surrounded by PC-enriched intermediate regions that transition into a more mixed POPC/PSM LD domain. That indeed coexisting domains are formed is clearly discernible from representative snapshots displayed in yellow) with only small more disordered patches, however at a comparably high lipid packing density. These differences in the formation of loosely packed LD domains are as well reflected in the average area per lipid. At 294 K, the area per lipid is ≈ 53 Å2 and increases to ≈ 56.5 Å2 at 303 K . Also, the domain composition changed upon addition of serotonin, in particular a significant change in the PC content is observed: While the PC and PSM content in the absence of serotonin is similar for the LD domain, addition of serotonin led to a significant enrichment of PC within the disordered domain at all investigated temperatures .To further assess the dynamical domain properties we analyzed the lifetime of POPC and PSM lipids within the ordered and disordered hidden states for the e lipids . The timnd 99 ns . Note, himescale . SimilarLO and LD phases as determined from the HMM differed substantially for the POPC/PSM/Chol (4:4:2) lipid membranes at 303 K, the average POPC sn-1-chain order 〈|SCH|〉 being 0.347 ± 0.008 for the LO phase and 0.224 ± 0.014 for the LD phase (see LDsn-1-chain order increases to 0.294 ± 0.017 at 294 K while the LOsn-1-order stays approximately constant (0.367 ± 0.006). The comparably high order for lipids assigned a disordered state by the HMM at low temperature is related to the small LD domain size and hence increased packing density of the lipids  at onin see . While iree case .Our results thus suggest a significant serotonin-induced decrease in the phase transition temperature for the POPC/PSM/Chol mixture.LO phase (POPC:PSM:Chol at a concentration of 8:61:31) binding almost exclusively occurred to the lipid headgroup region  profiles. The profiles are shown in  profile  at the pz axis of the indole and the bilayer normal; and α, the angle that is assumed when the bilayer normal is projected into the molecular x-y plane (compare section 2). right panel). However, orientations with θ = 50o or θ = 135o are preferred. This can be related to a tendency to form cation-pi stacking interactions with the choline headgroups, experimentally observed for indoles binding to PC lipids . The orientation is dominated by structures with θ near 90o—indicating that the molecular plane of the indole is orthogonal to the plane of the bilayer. This is particularly the case for serotonin binding to the LO system, while a much broader profile is observed for the LD and the mixed LO/D systems  is in good agreement with a previous simulation for the same composition  with respect to those of POPC-PSM mixtures (781 mN/m) have been experimentally reported by Needham and Nunn . The average area per lipid for this system is also in full agreement with that reported for the same composition by Sodt et al. domains differing in composition, structure, and ordering i.e., dynamics, and mechanics, starting from a random mixture of the membrane constituents. Additionally, we demonstrate that a high neurotransmitter concentration may heavily affect the equilibrium between disordered and ordered lipid membrane domains by preferential binding to LO) were defined in literature as domains containing lipids with extended and ordered acyl chains similar to the gel phase or Lβ-phase, and at the same time display a high mobility more similar to the disordered LD-phase . The latter may be explained by changed domain compositions as observed in the simulations displaying an increased PC content of the LD domains and a thus likely decreased PC content for (larger) ordered domains and thereby increased ordering. Interestingly, in good agreement with our simulation data, a recent work reports a significant softening of lipid bilayers containing POPC/POPG/Chol (1:1:1) by serotonin  is observed on the microsecond timescale. Despite small differences in compositions, formed nano-domains display a huge difference in order resembling those for PC-rich disordered and PSM/Chol-enriched raft-like ordered domains. The neurotransmitter serotonin reshapes in particular the disordered membrane domains resulting in a PC-enrichment and a decreased membrane elasticity and probably in increased membrane domain sizes as suggested by comparison to LD-domains with a concomitant change in composition, ordering, and reduced phase transition temperature likely as well alters the membrane lateral pressure profile as reported before also for ethanol (Griepernau and Böckmann, Regarding its action as neurotransmitter, our results suggest a fast binding of serotonin to the postsynaptic membrane following synaptic vesicle fusion and thereby a fast and efficient mechanism for neurotransmitter removal from the synaptic cleft (Postila et al., The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.RB designed the research. ABo setup and carried out the simulations. ABo and ABr analyzed the simulations. OE performed the zeta potential measurements and analysis. RB wrote the paper with contributions from all coauthors. All authors contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Al-Mg-Si based alloys are popular alloys used in the automotive industry. However, limited studies have been performed to investigate the microstructure, deformation characteristics, and deformation mechanism for the semi-solid 6063 alloys. In this study, the cold radial forging method and semi-solid isothermal treatment (SSIT) are proposed in the semi-solid isothermal compression (SSIC) process to fabricate high-quality semi-solid 6063 billets. The effects of deformation temperature, strain rate, and strain on the microstructure, deformation characteristics, and deformation mechanism of the SSIC of cold radial forged 6063 alloys were investigated experimentally. Constitutive equations were established based on the measured data in experiments to predict the flow stress. Results show that an average grain size in the range from 59.22 to 73.02 μm and an average shape factor in the range from 071 to 078 can be obtained in the microstructure after the cold radial forged 6063 alloys were treated with SSIT process. Four stages  were observed in the true stress- true strain curve. The correlation coefficient of the constitutive equation was obtained as 0.9796 while the average relative error was 5.01%. The deformation mechanism for SSIC of cold radial forged aluminum alloy 6063 mainly included four modes: The liquid phase flow, grain slide or grain rotation along with the liquid film, slide among solid grains, and the plastic deformation of solid grains. Al-Mg-Si based aluminium alloys are popular materials applied in various engineering applications due to their outstanding physical properties, which are usually used to fabricate products via hot forging and machining processes. However, some drawbacks are observed during the hot forging manufacturing process, which includes low efficiency, high production cost, and low material usage coefficient ,2,3. TheThese disadvantages including low efficiency, high production cost, and low material usage coefficient, gas and shrinkage porosity, can be effectively eliminated by the semi-solid metal forming (SSMF) process, which integrates the easy fluidity of liquid casting process and the high mechanical properties of solid-state plastic forming and is considered one of the most promising metal processing technologies in the 21st century ,7,8. In In the SSMF process, the deformation characteristics and mechanism of the semi-solid billets are vital for the prediction of the flow stress, based on the temperature, material structure, and strain rate, through which the optimisation of the process parameters can be achieved ,11. TherThe preparation of the semisolid billet is an indispensable step for the SSIC process. The SIMA, as one of the most popularly studied process methods for the preparation of the billet with fine and spheroidal solid particles, includes two stages namely the plastic deformation and the semi-solid isothermal treatment (SSIT). In the SIMA process, plastic deformation is the most critical stage as it significantly influences the grain size and grain distribution uniformity in the microstructure of the semi-solid alloy. Therefore, different deformation methods applied in the SIMA process, such as compression ,17, rollHowever, to the best knowledge of the authors, the aluminium alloy directly treated by cold radial forging in the SSIC process has not been reported. Moreover, little research has been performed on the microstructure, deformation characteristics, and deformation mechanism for the semi-solid 6063 alloys. In this study, the cold radial forging method is proposed in the SIMA process for the preparation of high-quality semi-solid 6063 billets for the SSIC process. The effects of deformation temperature , strain rate, and strain on the microstructure, deformation characteristics, and deformation mechanism of the cold radial forged 6063 alloys were investigated experimentally. Furthermore, the constitutive equation of the semi-solid 6063 alloys was established based on the experimental results to predict the flow stress based on the effective liquid phase fraction, the deformation temperature, strain rate, and strain.The chemical composition of 6063 aluminium alloy investigated in this study is summarized in ϕ60 mm). The reduction in the area is calculated by the ratio of (A0–A1) to A0, where A0 and A1 are the cross-sectional area of the initial alloy, and the deformed alloy, respectively. Then, the action of sampling was finished from the cold radial forged alloy along the direction of radial forging, which had the diameter and height of 10 mm and 15 mm, respectively. Stage II: The SSIC process including SSIT and the deformation of isothermal compression were carried out with the SSIC- equipment which mainly included the top pressing head, bottom pressing head, insulation material, resistance furnace, and thermocouples. During the SSIC process, the resistance furnace was heated up to the setting deformation temperature, and then the cold radial forged sample was placed between the top and bottom pressing head for SSIT. The semi-solid samples can be obtained when the SSIT process had been executed for 10 min at different isothermal temperatures. After that, these samples were compressed with the true strain of 0.7 to obtain the compressed semi-solid sample by the top and bottom pressing head at different deformation temperatures and different strain rates. For all the experiments, the deformation temperature was selected the same as the isothermal temperature in SSIT. Stage III: The compressed semi-solid sample was cooled rapidly by water quenching for preserving the microstructures of the sample.The SSIC process of the cold radial forged 6063 aluminium alloy in this paper mainly includes three stages as shown in −1, respectively. Similarly, in Experiments 7–9 and Experiments 10–12, the isothermal temperatures were the same in each group with the strain rate varying from 0.01 to 1 s−1.In this paper, there was a total of 12 semi-solid isothermal compression experiments performed to examine the effects of the operating parameters on the microstructure, deformation characteristics, and the deformation mechanism of the cold radial forged 6063 aluminum alloy. The details of the experiments are summarized in D) of solid grains can be calculated by Equation (1) while the shape factor (F) can be obtained by Equation (2) based on the number, area, and perimeter of solid grains, which were measured and calculated by the Image-Pro Plus 6.0 software  [A, N, P are the area, number, perimeter of solid grains, respectively; AL and AS are the area of the liquid phase, and the area of the solid phase, respectively.During the semi-solid isothermal compression experiments, the true stress and true strain are determined in-situ by the SSIC-equipment . Samples were taken along the axial direction, first ground through the sandpapers with the grits of 200, 400, 600, 800, 1000, 1200, and 1500 in sequence and then polished through the diamond paste with the particle size of 0.1 μm. Samples were etched in the aqueous HF solution with a concentration of 5% for about 90 s at room temperature. Finally, the observation of sample microstructure was carried out by optical microscope . The average grain size  ,25,26,27The microstructures of the radially forged 6063 aluminum alloy without and with SSIT are shown in The characteristics of the microstructure can be obtained by the quantitative analysis of samples processed at different isothermal temperatures, which includes the average grain size, the shape factor of solid grains, and the effective liquid fraction. As shown in The true stress-true strain curves of the radial wrought aluminium alloy 6063 during the semi-solid isothermal compression deformation procedure at different isothermal temperatures  are shown in It can be concluded from Stage I: Sharp increase. Semi-solid materials can be regarded as the material with solid grains suspended in the liquid phase, where the solid phase grains or solid-phase grain aggregates contact locally to form a spatial framework structure when the solid phase ratio is large ,10,30. AStage II: Decrease. As the SSIC continues, when the space framework structure reaches a certain degree of deformation, its deformation cannot be restored even if the load is removed. In other words, plastic deformation is achieved in the sample. At this stage, the spatial framework structure begins to be destroyed and is gradually and completely surrounded by the liquid phase when the deformation degree increased. The slip and partial rotation of the solid grains then occur at this time. The slip among the solid grains is realized by the shearing force, which promotes the occurrence of holes at the boundary of the grains. These holes would expand rapidly along with the liquid phase, resulting in the damage of samples. This sample damage causes the flow stress to drop rapidly, that is, the true stress drops.Furthermore, the plastic deformation will also promote the disaggregation of larger solid grain agglomerates, so that the liquid phase wrapped by the agglomerates is released, thereby increasing the liquid content inside the sample. This is helpful for the slippage and rotation of the solid grains, thereby contributing to the reduction of true stress.Stage III: Steady state. At this stage, the spatial framework structure formed by the solid grains is basically destroyed. The solid particles in the semi-solid material are surrounded by the liquid phase and undergo plastic flow. The deformation resistance is mainly attributed to the slippage and rotation among the solid grains as well as the liquid phase flow. Therefore, the deformation resistance remains basically unchanged, that is, the true stress is at a steady state as the displacement increases.Stage IV: Slow increase. The main reason for the increase in true stress at this stage is that the true strain is too large, which causes the semi-solid material to roll over during the SSIC process, gradually increasing the contact surface between the pressing head and the sample. This in turn increases the friction between the pressing head and the sample. Furthermore, during the SSIC process of the sample, as the true strain increases, most of the liquid is separated from the solid phases inside the sample and squeezed to the edge of the sample, resulting in an increase in the local solid fraction of the sample. This consequently causes the solid grains to be squeezed and bonded together and therefore generates the plastic deformation . The preThe effects of the deformation temperature on the peak stress and steady-state stress are shown in (a)At the beginning of the SSIC deformation process, the deformation speed of the sample was small because the strain rate was low, which provided enough time for the liquid phase to participate in repairing the liquid phase film among the deformed solid grains. The deformation of the spatial framework structure of the sample can take place with the slippage and rotation among the solid grains, resulting in a small deformation resistance and small peak stress. On the contrary, the liquid phase did not have enough time to repair the liquid film among the deformed solid grains when the strain rate was high, which caused the difficulty of solid grains to slip and rotate. The solid grains squeezed with each other in this early stage, which hindered the deformation of the sample, thereby, increasing the peak stress.(b)The spatial framework structure of the sample was gradually and completely destroyed with the increase in the semi-solid isothermal compression deformation displacement, which formed solid grains or solid grain agglomerates wrapped by the liquid phase. This influence of the liquid phase on the solid grains or solid grain agglomerates contributed to the occurrence of compression deformation, so the true stress gradually decreased and stabilized. When the strain rate was low, the movement speed of the liquid phase film among the solid grains was small, resulting in the weak shearing effect among the solid grains. Consequently, it was difficult to achieve the disaggregation and destruction of solid grain aggregates. Therefore, higher steady-state stress was required to promote the coordinated deformation between the solid grain agglomerates after entering the steady state flow. On the contrary, when the strain rate was high, the strong shearing effect between the solid grains can promote the disaggregation and destruction of some solid grain aggregates, so that the liquid phase wrapped by the aggregates was released. The liquid content inside the sample, thereby, increased, which was beneficial for the slippage and rotation of the solid grains, so the steady-state stress was observed relatively low.The effects of the strain rate on the peak and steady-state stresses of the semi-solid isothermal compression curve of aluminum alloy 6063 are presented in −1.The constitutive model expresses the material’s dynamic response to the deformation amount, deformation temperature, and the strain rate during the plastic processing, which is crucial for the numerical simulation of the plastic deformation process using the finite element method and can be illustrated by Equation (4) ,(4)σ=f–(8) ,33,34. IL is the liquid phase factor; The magnitude of the peak stress is related to the selection of processing equipment and moulds in the semi-solid forming process. In this study, the peak stress is taken as the research object, the liquid phase correction term is introduced into the Arrhenius equation to modify Equation (8). Consequently, the Arrhenius equation adapted to the semi-solid temperature zone can be obtained as Equation (9),In order to obtain the value of α, take the logarithm of Equations (6) and (7) to convert into the linear Equations (10), and (11), respectively:Introducing the peak stress and the corresponding strain rate at different deformation temperatures into Equations (10) and (11), the relationship between the strain rate and the peak stress at different deformation temperatures can be obtained, as shown in n, Equation (12) can be obtained by taking the logarithm of both sides of Equation (9):In order to calculate Importing the peak stress and the corresponding strain rate at different deformation temperatures into Equation (12), the relationship between Based on the results in A, L, and Q are unknown. The least-squares solution was used to solve the overdetermined equation in this study. These 9 equations can then be written as the following matrix illustrated by Equation (14),Therefore, one Equation (13) can be obtained at a certain deformation temperature and a certain strain rate. Correspondingly, 9 equations can be obtained for 3 deformation temperatures and 3 strain rates, where only three parameters A, L, and Q, Equation (14) is multiplied by To calculate the values of parameters Introducing the corresponding parameters into Equation (13), matrixesX can be obtained by performing transposition and inverse matrix operations using Matlab software, which is Introducing the above matrixes into Equation (16), the matrix It is worth noting that the applicable range of N is the statistical number of data points.The correlation coefficient R and the average absolute relative error (AARE) were used in the study to indicate the accuracy performance of the established peak stress equation of the aluminium alloy in the semi-solid region, which are expressed by Equations (18), and (19), respectively,As shown in −1 with different deformation temperatures. It was found that the compressed sample gradually broke with the elevation in the deformation temperature. The higher the deformation temperature, the more obvious the degree of crushing of the compressed sample. This was because, at the beginning of the semi-solid isothermal compression deformation, the liquid phase already existed in the aluminium alloy 6063 semi-solid material, the fraction of which elevated with the enlargement in the deformation temperature. The liquid phase in the compressed sample was squeezed and flew to the sidewall of the sample at the central region and caused the sidewall to rupture during the SSIC deformation process. This liquid phase in the compressed sample continued to flow along the rupture of the sidewall in the final stage of the SSIC deformation, resulting in the fragmentation of the entire compressed sample. Nevertheless, the phenomenon aforementioned did not occur in the actual SSMF process. This was mainly because the mould cavity in the actual SSMF process was closed, which consequently avoided the outflow of the liquid phase [The macroscopic photo of the radially forged aluminium alloy 6063 semi-solid isothermal compression samples is shown in id phase ,39.The macroscopic photo of the samples processed at the deformation temperature of 630 °C under different strain rates is shown in −1, the deformation temperature was set as 630 °C, and the strain was 0.7. It was found that similar to the compression deformation of solid metal, the semi-solid isothermal compression deformation sample also exhibited three different deformation regions, which are the hard deformation area near the end (Region A), the huge deformation area in the centre (Region B), and the free deformation region (Region C) in the convex region.According to the principle of pressure processing ,32, the The microstructures from different positions of the semi-solid compressed sample are shown in The liquid phase of the semi-solid material was mainly distributed at the boundary of the solid grains, which has the effect of isolating solid grains. When the compression force was applied to the semi-solid material, the separated flow of liquid and solid phases would take place. Generally, the liquid content has a tendency to move before the solid grains. The reduction of the liquid phase would cause the subsequent flow of solid grains to contact with each other. If the deformation continues, severe plastic deformation would take place in the solid grains. Furthermore, the flow of the liquid phase would also cause the liquid phase segregation. The deformation was restricted in the deformation region. Therefore, the microstructure basically remained the original semi-solid microstructure without obvious segregation, as shown in The nearly spherical phenomenon was observed in solid grains in the free deformation region (see Positions 4 and 5), but a more serious liquid phase segregation was also found as shown in Based on the analysis mentioned above, the deformation mechanism of the radially forged aluminium alloy 6063 material during the SSIC mainly included the grain slide or grain rotation along with the liquid film, the sliding among solid grains, and the plastic deformation of solid grains. Moreover, according to the analysis of the microstructure in different deformation regions, during the SSIC, different degrees of liquid phase flow occurred in all these three deformation regions. Considering this liquid phase flow, there were four deformation mechanisms in SSIC, which is presented in As presented in It can be found in In summary, the phase fraction of liquid determines the deformation mode of the semi-solid material during the isothermal compression deformation process. When the phase fraction of liquid is large, the content of the liquid phase in the microstructure is high and thus, solid grains would not completely contact each other. The flow channel of the liquid phase is therefore unobstructed. The deformation mode mainly contains the liquid phase flow and the sliding or rotation of the solid grain along with the liquid film. When the phase fraction of liquid is small, the content of the liquid phase in the microstructure reduces, resulting in the local contact of solid grains. The flow channel of the liquid phase is then hindered. The deformation mode then mainly includes the sliding among solid grains and the plastic deformation of solid grains. Furthermore, the order of the main deformation mechanism with the decrease in the liquid phase fraction is from the liquid phase flow to the grain slide or grain rotation along with the liquid film, then to the sliding among the solid grains, and finally to the plastic deformation of solid particles. In terms of a specific compressed sample, the grain slide or grain rotation along the liquid phase is the main deformation mode in the hard deformation region (Region A); the plastic deformation of solid grains is the main deformation mode in the huge deformation region (Region B), while a certain degree of sliding among the solid grains also appears in this region; the liquid phase flow and the grain slide or grain rotation along the liquid film are the main deformation modes in the free deformation region (Region C).In this work, SSIC is proposed to investigate the microstructure, deformation characteristics, and deformation mechanism of semi-solid alloy, where the cold radial forging is introduced in the SIMA process to prepare the high-quality semi-solid billet. The main conclusions drawn from this study are shown as follows.(1) The average grain size, shape factor, and effective liquid phase fraction increase with the enhancement in the deformation temperature. The high-quality semi-solid alloy, which has the average grain size in the range of 59.22–73.02 μm while the average shape factor in the range of 0.71–0.78, can be obtained when the cold radial forged 6063 aluminum alloy is treated by SSIT process at the deformation temperature in the range of 625–635 °C.(2) The true stress-true strain curve of the cold radial forged 6063 aluminium alloy SSIC mainly includes four deformation stages: sharp increase stage, decrease stage, steady state stage, and slow increase stage. The true stress decreases when the deformation temperature increases at a certain strain rate, which means that both the peak and steady-state stresses decrease with the enlargement in the deformation temperature. The peak stress elevates while the steady-state stress decreases with the enlargement in the strain rate at a certain deformation temperature.(3) Introducing the liquid phase correction term (4) The deformation mechanisms of cold radial forged aluminium alloy 6063 during SSIC mainly include four modes: liquid phase flow, grain slide or grain rotation along with the liquid film, sliding among solid grains, and the plastic deformation of solid grains. For a specific sample, the grain slide or grain rotation along the liquid film is the main deformation mode in the hard deformation region. In the huge deformation region, the plastic deformation of solid grains is the main deformation mode while a certain degree of sliding among solid grains also exists. The liquid phase flow and the grain slide or grain rotation along the liquid film are the main deformation mode in the free deformation region.


Rice flour increased RKB batter consistency, solid- and liquid-like viscoelastic behavior and revealed a heterogeneous structure, based on the sweep frequency test. Rice flour at the 25% level increased the shear modulus and activation energy of gelatinization, compared to 0% rice flour addition. Rice flour levels in the RKB batter decreased the inflection gelation temperature from 63 to 56 °C. In addition, the texture of RKB cupcakes with 25% rice flour were 46% softer, compared to the control. The scores from all sensory attributes of cupcakes increased with the addition of rice flour. Rice flour addition improved solid- and liquid-like behavior of the RKB batter and improved the cupcake’s macro-structural characteristics. Overall, 25% rice flour addition performed better than the lower levels. This study confirmed the potential of RKB as a functional ingredient and its improvement in cupcake application with the addition of rice flour. Red kidney bean (RKB) flour is a nutrient-rich ingredient with potential use in bakery products. The objective of this study was to investigate the viscoelastic properties and key quality parameters of a functional RKB flour in gluten-free cupcakes with different rice flour levels. A 10 g model batter was developed for analyzing the viscoelastic properties of RKB with rice incorporation, in a formula containing oil, liquid eggs, and water. Rice flour was added at five levels 0%, 5%, 10%, 15%, and 25% ( Phaseolus vulgaris L.) is a good source of vegetable protein, insoluble and soluble fiber, and is overall a nutrient-rich food ingredient with a great potential for different food applications [Red kidney bean  were rinsed, boiled in water for 20 min, strained, and dried for 4 h at 80 °C using a convection oven. These conditions were selected from Chompoorat et al. (2018) . The resGluten-free RKB cupcakes were prepared with ingredients purchased from a local market and RKB flour produced in the laboratory. The cupcake formula expressed as a weight percent of RKB flour base (100%) contained granulated white sugar (65%), baking powder (1.9%), baking soda (1.1%), cocoa powder (10%), salt (1%), guar gum (0.5%), vegetable oil (37%), egg (65%), water (150%), and vanilla extract (3%). Commercial rice flour was added at 0%, 5%, 10%, 15%, and 25% based on RKB flour. The control sample contained 100% RKB flour.The cupcakes were prepared by mixing all dry ingredients  for 1 min at the lowest speed (setting# 1), using a paddle attachment. Wet ingredients  were added, mixed for 2 min at low speed (setting #1), followed by 5 min at high speed (setting # 6). Cupcake batter aliquots (40 g) were placed into paper-lined molds and baked in a convection oven at 195 °C for 20 min. The cupcakes were removed from the mold, allowed to cool for 60 min at room temperature, and immediately transferred for analysis of firmness inside a cupcake cardboard box.Batter aliquot (100 g) was used for consistency measurements with a Bostwick consistometer , following manufacturer’s directions. The cupcake batter was transferred to the consistometer chamber, the gate was released, and the batter flow rate (cm per min) was recorded. Batter flow rate was inversely proportional to consistency. Measurements were made in three replicates per treatment.G′ (storage modulus), G″ (loss modulus), and tan delta (δ) were determined by this dynamic oscillatory measurement. Analyses were performed in three independent samples.A model batter sample was prepared containing all the ingredients listed in G′ , the stored energy in each batter sample or recoverable cycle of deformation and G″ , the loss energy in each batter sample or viscous dissipation per cycle of deformation. Complex shear modulus (G*) was calculated  at the minimum value of dG′/dt, a, b, and c are the constants of the model, and c can be used to calculate the Ea (kJ.mol−1) activation energy (Equation (3)), where R is 8.314 J mol−1 K−1 . Activation energy (Ea) revealed the energy required for gelatinization [The kinetic parameters of the model (Equation (2)) were obtained by numerical methods, according to Alvarez et al. (2017); it was performed with the numerical derivative of the elastic moduli with respect to time. Where Cupcake firmness is among the key texture parameters that influence consumer sensory perception and acceptance. After the cupcake samples were cooled to ambient temperature (25 °C), firmness was measured using a TA-XT2 Texture Analyzer . Analysis was performed following the Approved Method outlined by the American Association of Cereal Chemists (AACC 74-10.02) for bread, using a 25-mm diameter cylindrical probe and 40% compression. Cupcake firmness was performed with 12 independent replicates per sample. Organoleptic properties of red kidney bean-rice cupcakes were evaluated with a 9-point hedonic scale ranging from ‘dislike extremely’ (1) to ‘like extremely’ (9). Cupcakes were tested after being cooled for 60 min. The samples labeled with random 3-digit codes were scored by twenty untrained panelists, students of the Postharvest Technology at Maejo University, Thailand. The sensory attributes included flavor, taste, texture, and overall quality.n = 12) and sensory evaluation (n = 20). Mean comparison for significant differences of each treatment was tested by Duncan’s New Multiple Range, using the SAS program . Correlations of the rice flour level effect on batter and cupcake properties were estimated with Principal Component Analysis (PCA) by Canoco for the Windows 5 software .Results are expressed as mean values ± standard deviation of the minimum three independent analyses, unless otherwise described, such as cupcake firmness  reflecting negative slopes starting at 10% rice level. Increased consistency of the batter matrix could be explained in part by the increase in air entrainment forming air bubbles, which increase the elastic character of the batter [Consistency was estimated with a 100 g-aliquot of full formula batter by measuring the flow rate in a consistometer, as used in the industry for quality control. The test was simple, fast, and provided enough information for consistency standardization. The levels of rice flour significantly decreased the flow rate (increased consistency) of gluten-free RKB cupcake batter, starting at 10% rice flour addition . Rice fle batter . This mie batter . The vale batter . Starch e batter ,26. The e batter ,28. Preve batter ,30,31. Re batter ,33. ThusG′ (storage modulus) and G″ (loss modulus) increased as the levels of rice flour increased in the frequency range of 0.1 to 10 Hz (G′ > G″). Cake batter is considered a wet foam with contributions to its bulk rheology from the continuous viscous phase and the discontinuous bubble phase (elastic effect) introduced during mixing and the production of gas from leavening agents [G′) of the batter (22.4 Pa for control up to 50.7 Pa for 25% rice flour) was higher than the 1.3 fold increase of the liquid-like behavior (G″) (17.6 Pa for control and up to 22.1 Pa for 25% rice flour) (G′) for the batter; when the rice flour content varied from 5% to 15%, the rice flour showed a similar behavior. to 10 Hz . Elasticg agents . At 1 Hze flour) . Adequate flour) . We specG′ and G″ that were used to evaluate if the elastic and storage moduli followed a power law relationship in the RKB batter samples, with different rice flour levels. If the response was linear, then the relationship followed a power law, suggesting that a constant increase of G′ led to a constant increase in G″. The same plot, known as the Han plot, was used to determine the compatibility of polymer blends [G’), compared to the control (p = 0.05) , as compared to the control RKB flour and other rice levels. Our previous study of RKB batter with different heat-moisture treatments showed that the high temperature altered the viscoelastic properties of the RKB batter; however, their structures were homogenous [r blends ,35. Over = 0.05) . The tramogenous . Thus, FG*) increased as the tan δ decreased (G*) trends after 50 °C  lower tan δ at the initial stages of heating before the gelation onset, compared to the rest of the samples, suggesting a higher solid-like character (G′) in this sample. That higher solid-like behavior was conserved up to around 50 °C, where the decrease of tan δ accelerated, suggesting a rapid increase in solid-like behavior. This was attributed to the starch and protein hydration, resulting in gel rigidity onset. Tan δ curves crossed-over the G* curves at around 60 °C (rice flour additions) and 65 °C (RKB flour control), suggesting more rapid changes occurring in the structures. Most likely these changes were related to the denaturation of proteins and the rapid starch granule swelling at those temperatures [G* and decrease tan δ (The tracing curves of tan δ showed two (rice flour additions) and three (RKB flour) clear inflection points, suggesting different changes of structures (phase transitions) as a function of increasing temperature . Rice fleratures ,30. The se tan δ , in the aE) was used as an indication of the energy used for the critical gel rigidity during gelatinization, while the inflection temperature (0T) was used for the point at which the batter viscoelasticity increased at the initial stage of gel point. 0T is normally associated with batter density and level of gelatinization [G′ was higher than the values of our study [0T) from 63 to 56 °C, with no significant (p = 0.05) change in the activation energy (aE). Significant difference (p = 0.05) in activation energy was observed only between the control RKB flour and 25% rice addition, where a 44% increase was needed to reach the critical gel rigidity (p < 0.05). This could be partially explained by the smaller starch granule size of rice flour (3–8 µm), compared to the kidney bean starch (16–60 µm), increase in overall starch content in the system, and RKB flour protein reduction [Ea) from this non-isothermal heating of small amplitude oscillatory shear test could be useful to indicate the changes during gelatinization.Activation energy  moved slightly to the horizontal PC2, meaning it had some variations explained by it. The fact that 10% rice addition was almost in a direction opposite to the viscous behavior also suggest that it was inversely correlated to 10% rice addition. However, as the contribution to the variance was low—only 12.1%—compared to 80.3% contribution to the variance of PC1, it was better to highlight the contributions of PC1. Notice how all parameters were along PC1. The vectors aligned in opposite direction revealed negative correlation, while the vectors in the same direction were positively correlated. The results from the loading plot (0T) of gelation, and the treatments associated with those parameters were 0% and 5% rice levels. Complex modulus G*, solid (G′) and viscous (G″) moduli were positively correlated with the activation energy (Ea) of gelatinization and the highest rice flour level (25%), which was the farthest away from the center of the graph. Rice flour addition to RKB flour was negatively correlated to cupcake firmness. These results confirmed that the most variation was observed at the highest rice level (25%). This study also revealed that the gelation of the batter and its consistency were good indicators of quality control for gluten-free RKB–rice flour cupcakes. Total explained variance from all variables was 92.4% . Principing plot  revealedG′ vs G″. In addition, the activation energy of gelatinization increased with 25% rice incorporation. This study described an improvement of the RKB batter and cupcake macro-structural characteristics with addition of rice. The level of rice flour addition improved the viscoelastic properties of gluten-free RKB batter and cupcake softness. With 25% of rice flour addition in gluten-free RKB flour, the system increased the batter’s solid-like and viscous behavior, batter consistency, inflection of gelatinization and temperature, and produced a softer cupcake texture. The RKB batter with rice followed a power law and had a heterogeneous structure based on the log–log plot of


Based on a national survey of program directors we developed a letter of recommendation (LOR) scoring rubric (SR) to assess LORs submitted to a pediatric residency program. The objective was to use the SR to analyze: the consistency of LOR ratings across raters and LOR components that contributed to impression of the LOR and candidate.We graded 30 LORs submitted to a pediatric residency program that were evenly distributed based on final rank by our program. The SR contained 3 sections  and 2 questions about the quality of the LOR (LORQ) and impression of the candidate (IC) after reading the LOR on a 5-point Likert scale. Inter-rater reliability was calculated with intraclass correlation coefficients ). Pearson (r) correlations and stepwise multivariate linear regression modeling predicted LORQ and IC. Mean scores of phrases, features, and applicant abilities were analyzed with ANOVA and Bonferroni correction.2 = 0.75, p<0.001) and IC  were best predicated by: writing about candidates’ abilities, strength of recommendation, and depth of interaction with the applicant. For abilities, LORQ  and IC  were best predicted by: clinical reasoning, leadership, and communication skills (0.2). There were significant differences for phrases and features (p<0.05).Phrases  = 0.82, p<0.001)) and features  = 0.60, p<0.001)) were rated consistently, while applicant abilities were not  = 0.28, p<0.001)). For features, LORQ (RThe SR was consistent across raters and correlates with impression of LORQ and IC. This rubric has potential as a faculty development tool for writing LORS. The letter of recommendation (LOR) is valued by program directors (PDs) when making decisions about candidates to interview and rank in their programs, yet little is known about how these documents sway PDs and contribute to those decisions –3. Whilehow LORs are used make decisions about applicants. While writing LORs is a time-honored tradition and it is hard to imagine the application without them, they take time to both read and write and are fraught with challenges [What is evident after reviewing the literature is there is limited research reporting allenges . The litallenges –14. Themallenges , 14. It allenges –19.To address this gap in the literature we sought to build on work that was previously published reporting how residency PDs in pediatrics, surgery, and internal medicine interpreted three components of LORs , 14. We Using this data we developed a LOR scoring rubric to assess LORs submitted to a pediatric residency program. The objectives of this study were to use the rubric to analyze (1) the consistency of LOR ratings across raters, and (2) LOR components that contributed to overall impression of the LOR and candidate. One aim was to develop a tool that could be used in faculty development to write a more informative LOR.Six medical educators with leadership positions in the pediatric residency program, clerkship program, and intern selection committee developed a LOR scoring rubric based on results of previously published studies looking at how PDs interpret components of LORs , 14. As Before finalizing the rubric, we read additional LORs to mine for any additional phrases or applicant abilities that were not included in the original survey using LORs of applicants on our final rank list from our 2016–2017 application season. They were randomly selected by our program coordinator and all identifying information  was redacted. For each round of development each letter evaluator was sent the same 5 LORs to evaluate using the scoring rubric. Using an iterative process we discussed letter phrases, features, and applicant abilities that were identified in the LORs and they were added if consensus was achieved. We reviewed 30 LORs for a total of six iterations until no further feedback was generated and theme saturation was achieved. The final rubric contained 21 letter phrases, 4 letter features, and 14 applicant abilities with 34 synonyms. There were two additional questions asking letter evaluators to rate (1) the overall quality of the LOR and (2) the overall impression of the quality of the applicant after reading the LOR on a 5-point Likert scale . A final copy of the rubric is available Using the finalized scoring rubric we graded 30 new and randomly selected LORs submitted to a moderate-sized pediatric residency program from our final rank list of 265 students. The LORs were evenly distributed based on final rank by our program with 10 top tertile, 10 middle tertile, and 10 lowest tertile. Inter-rater reliability was calculated with intraclass correlation coefficients ). Pearson (r) correlations and stepwise multivariate linear regression modeling predicted the overall quality of the letter (LORQ) and impression of the quality of the applicant (IC) after reading the LOR. The mean scores of letter phrases, features, applicant abilities, and position on final rank list were analyzed with ANOVA and the Bonferroni correction for multiple comparisons. A waiver of informed consent was approved and this study was approved by the Institutional Review Board of the Medical College of Wisconsin.2 = 0.75, P < 0.001) was best predicted by writing about candidates’ specific abilities (b = 0.5), including a summative statement on the strength of the recommendation (0.3), and describing the depth of interaction with the applicant (0.2). The overall IC  was best predicted by the same 3 features as depicted in There was strong inter-rater reliability between letter evaluators for LOR commonly used phrases  = 0.82, P < 0.001)) and features  = 0.60, P <0.001)), but not applicant abilities  = 0.28, P < 0.001)). LORQ and IC scores were strongly correlated with commonly used phrases, letter features, and applicant abilities . For letter features, LORQ  was best predicted by clinical reasoning (beta = 0.5), leadership (0.3), and communication skills (0.2). Trustworthiness, maturity, enthusiastic, team player, professionalism, compassionate, resilience, resourcefulness, inquisitiveness, and efficient did not factor into the model. The overall IC  was best predicted by the same 3 abilities. Team player, professionalism, compassionate, resilience, resourcefulness, inquisitiveness, and efficient did not factor into the model as shown in For applicant abilities, LORQ  as shown in In this study looking at components of LORs that contributed to overall impression of the letter quality and impression of the applicant, we developed a scoring rubric that demonstrated good inter-rater reliability. Top tier LORs contained significantly more positive phrases, described the applicant better, and achieved a higher position on the final rank list than middle and lowest tier LORs. Select letter features and applicant abilities best predicted the strongest LORs and most favorable impression of the applicant. These results may help letter writers craft more informative LORs.Developing a tool to reliably rate LORs was an important goal of this study. Prior research has cautioned that an objective system to evaluate LORs might prove too challenging to develop given the unique characteristics of the letter writer, applicant, and written language . We wereOur results also shed some light into what pediatric residency program leadership may value in terms of applicant abilities when making selections about who to interview and rank in their programs. In the LORs reviewed for this study, clinical reasoning, leadership, and communication skills emerged as the top predictors of best letter quality and impression of the applicant. It is interesting that many other desirable abilities did not factor into the model, such as professionalism, when results of the 2020 National Resident Matching Program (NRMP) survey indicate that PDs in all specialties value professionalism as an important predictor of resident success in their program . There wOur results may best be suited for applications in faculty development as a majority of faculty report they receive little training for this important part of their job . BecauseThis study was conducted at a single pediatric residency program, so may not be generalizable to other programs and specialties. Because the initial rubric was developed using the results of a survey of program PDs in pediatrics, surgery and internal medicine we know that PDs in all three specialties rated commonly used phrases, letter features, and applicant abilities very similarly so could speculate they would value a very similar LOR , 14. AreThe scoring rubric was consistent across raters and correlates with raters’ overall impression of the letter quality and impression of the applicant. Our results show promise for faculty development in quality letter writing.S1 File(PDF)Click here for additional data file.


It was first identified as being induced upon injury to rat arteries and was found to be highly expressed in multiple human cancer types. Here, we explore the phylogenetic and evolutionary trends of this metazoan gene family, previously studied only in vertebrates. We identify Cthrc1 orthologs in two distant cnidarian species, the sea anemone Nematostella vectensis and the hydrozoan Clytia hemisphaerica, both of which harbor multiple copies of this gene. We find that Cthrc1 clade-specific diversification occurred multiple times in cnidarians as well as in most metazoan clades where we detected this gene. Many other groups, such as arthropods and nematodes, have entirely lost this gene family. Most vertebrates display a single highly conserved gene, and we show that the sequence evolutionary rate of Cthrc1 drastically decreased within the gnathostome lineage. Interestingly, this reduction coincided with the origin of its conserved upstream neighboring gene, Frizzled 6 (FZD6), which in mice has been shown to functionally interact with Cthrc1. Structural modeling methods further reveal that the yet uncharacterized C-terminal domain of Cthrc1 is similar in structure to the globular C1q superfamily domain, also found in the C-termini of collagens VIII and X. Thus, our studies show that the Cthrc1 genes are a collagen-like family with a variable short collagen triple helix domain and a highly conserved C-terminal domain structure resembling the C1q family.Collagen triple helix repeat containing protein 1 (Cthrc1) is a secreted glycoprotein reported to regulate collagen deposition and to be linked to the Transforming growth factor  Cthrc1 ant line . It was ant line , severalant line , gastroiant line , and in ant line . The expant line .Cthrc1 gene was first reported only in vertebrates and in the ascidian Ciona intestinalis  on the JGI genome database , Stellabase (Nve-Cthrc1 genes were assembled by extracting for each at least one full-length transcript from the above databases and verifying its sequence by using BLAST (BlastN) on the Nematostella JGI genome database; the other JGI genomic entries which encompassed identical albeit partial sequences of these genes were discarded. The Clytia hemisphaerica Cthrc1 genes were retrieved by BLAST (TBlastN) from the current genome/transcriptome assemblies  were excluded.The database  using thellabase , the traellabase , and theellabase . Eight fsemblies  using the-value < 0.01) on available proteins, genomes, and transcriptomes  as well as on dedicated databases  analyses were performed using RaxML v8.2.9  on a LinMus musculus, Gallus gallus, Xenopus laevis, Cyprinus carpio, Danio rerio, Salmo salar, Takifugu rupestris, Callorhinchus milii, Petromyzon marinus, and Ci. intestinalis using the Ensembl, NCBI, and ANISEED databases (FZD6 and Slc25a32 genes (see above) was determined using the corresponding genome databases.The genes located immediately upstream and downstream of Cthrc1 were identified in human, atabases  . We then searched the genome of the recently sequenced hydrozoan model C. hemisphaerica (Che-Cthrc1’s). As in vertebrates, the predicted proteins comprise a signal peptide, a CTHR domain, and a conserved C-terminal domain , hinders a precise reconstruction of the evolutionary history of cnidarian Cthrc1 genes. We could nevertheless conclude with confidence that the Cthrc1 paralogs found in Clytia and Nematostella originated from distinct diversification events  (100% BP for each) were found in close proximity on the same genomic scaffold and likely resulted from tandem duplications (Cthrc1 sequences between N. vectensis and its close relative Edwardsiella lineata (Edw) allowed defining orthology groups   ((Nve-Cthrc086 and Spi_XP_022784555–66% BP)   online, 4–213 Ma . Within c1 genes  (Nve-Cth–48% BP)  online. –66% BP)  online, Cthrc1 repertoire across eukaryotes, by means of broad genome and transcriptome analyses, revealed at first that Cthrc1 is a metazoan-specific gene . Sequences resembling Cthrc1 C-terminal domain—but lacking the CTHR domain—could nevertheless be identified in several choanoflagellate transcriptomes , strongly arguing in favor of an independent combination of the CTHR and Cthrc1 C-terminal domains in Metazoa and Bacteria.Interestingly, we could find Cthrc1 C-terminal domain in several bacteria metagenomes, including some predicted proteins which also contain a CTHR domain  online. Cthrc1 genes in several poriferan, cnidarian, and bilaterian clades but none in Ctenophora and Placozoa. The patchy distribution of the Cthrc1 genes across Metazoa suggests multiple loss events  as well as losses in annelids and in Rotifera. In addition, we could not identify Cthrc1 in any platyhelminth, molluskan, and nemertean available genomes. In urochordates, Cthrc1 was lost in appendicularians as well as in the colonial species Botryllus schlosseri and Botrylloides leachii, whereas it was found in the transcriptome of closely related ascidian species . Only a minority of protostomes possess Cthrc1 genes (up to 9 in Priapulus), whereas loss of this gene family was rare among deuterostomes , with vertebrates displaying a unique phylogenetic pattern.The phylogenetic distribution of Cthrc1 gene family in gnathostomes. In lamprey, only a partial Cthrc1 sequence could be found. In most vertebrate species, only one Cthrc1 gene could be identified, with up to four in teleost fishes. Vertebrate Cthrc1 phylogeny reconstructed using the NJ algorithm is overall congruent with the accepted vertebrate species phylogeny, with monophyly recovered for the major vertebrate groups, such as teleosts, tetrapods, amniotes, frogs, mammals, and birds  also likely resulted from more recent whole genome duplication events , show markedly longer branches  and Slc25a23  and zebrafish (7 Mb apart). The mitochondrial folate carrier gene Slc25a32 was found downstream of Cthrc1 in all gnathostome genomes except in carps , but not from other primates and other mammalian genomes  and more “conservative” groups losing it . This phylogenetic pattern, nevertheless, suggests a rapid evolution of the function of Cthrc1 in Metazoa.In spite of the uncertainty about the origin of Cthrc1, we could confidently infer many losses of this gene family during metazoan evolution as well as gene family expansion in other groups. Several of the groups that lost e losses . Conversfamilies . The numHydra, planaria, Hofstenia, and Botryllus) have nevertheless lost Cthrc1. These highly regenerative animals, however, all rely on specialized stem cell populations, which might have disengaged them from the putative Cthrc1-dependent remodeling. Functional data on Cthrc1 genes among different metazoan groups are required for a better understanding of the repeated diversifications and losses of this gene family and to assess its signaling function and contribution to body patterning and regeneration.The conserved structure of Cthrc1, together with our current knowledge of the molecular activity of these proteins in vertebrates, suggests a conserved signaling function across metazoans. Interestingly, many of the groups harboring multiple Cthrc1 display indirect development, with an intermediate larval-like form before the adult stage, and high regenerative capabilities. This would correlate with the epithelial–mesenchymal transition and migration promoting abilities of Cthrc1 shown in vertebrates  and the Cthrc1 genes were remarkably conserved in sequence and structure, with identical size for the CTHR domain in all vertebrates, in contrast to the high variation observed between nonvertebrate groups  and a C1q/TNF domain (Our structural analyses showed that the hitherto “orphan” C-terminal domain of Cthrc1 proteins bears similarity to collagen and collagen-like protein domains for which structural data exist. We therefore concluded that the C-terminal domain of Cthrc1 shows the highest similarity to the globular C1q domain of collagens VIII and X and less to the COLFI domain of fibrillar collagens and to the C-terminal domain of collagen IV. In the original description of F domain . They, hF domain . The assF domain . The higCthrc1 was retained and duplicated in several animal clades, lost from many others, and was highly conserved in terms of sequence, domain size, and copy number in the vertebrate lineage.Is Cthrc1 a bona fide collagen? The term collagen refers to a diversity of extracellular matrix protein families all sharing a CTHR domain (reviewed by evaa020_Supplementary_DataClick here for additional data file.


Grus japonensis) and how this subsequently influences crane diet and use of wetland sites. Pearson's correlations and generalized linear models revealed that mudflat crabs became significantly more surface active with increasing burrow ambient temperature. Piecewise regression analysis revealed that crab surface activity was largely limited to a burrow ambient temperature threshold between 12 and 13℃ after which activity significantly increased. Crab activity declining temporally during the crane's autumn migration period but increased during spring migration. Crabs accounted for a significant proportion of crane diet at two of three sites; however, the frequency of crab remains was significantly different between sites, and between autumn and spring migration. Analyses of crane count data revealed a degree of congruence between the migration timing of Red‐crowned cranes with periods of warmer ambient temperature, and a significant, positive correlation between the percentage of crab remains in crane feces and site ambient temperature. Collectively, our data suggest that temperature‐related mudflat crab activity may provide an important time window for migratory Red‐crowned cranes to utilize critical stopover sites and the crabs’ food resources.For migratory birds that specialize on particular benthic macroinvertebrate species, the timing of migration is critical since prey availability may be temporally limited and a function of local ambient temperature. Hence, variation in local ambient temperature can influence the diet composition of migrant birds, and, consequently, they may be constrained by which stopover and wintering sites they are able to utilize during periods of colder temperatures. Here, we use fecal analysis, observer‐based population counts, digital video recordings, and temperature data to test five predictions regarding the influence of local ambient temperature on the activity and availability of mudflat crabs—a key prey resource at three staging/wintering sites in eastern China, for migratory Red‐crowned cranes ( Mudflat crabs became significantly more surface active with increasing burrow ambient temperature, and the crab surface activity was largely limited to a burrow ambient temperature threshold between 12 and 13°C after which activity significantly increased. Crabs accounted for a significant proportion of crane diet at two of three sites; however, the frequency of crab remains was significant different between sites, and between autumn and spring migration. A degree of congruence between the migration timing of Red‐crowned cranes with periods of warmer ambient temperature, and a significant, positive correlation between the percentage of crab remains in crane feces and site ambient temperature. Li et al.  and Yancheng Nature Reserve (YNR) and mudflat crab populations occur at both these sites.The Red‐crowned crane  mudflat crabs become more surface active with increasing burrow ambient temperatures; (2) crab surface activity would be limited to a temperature threshold; (3) crab activity would decline temporally during the crane's autumn migration period but increase during the spring migration period with increasing temperature; (4) crane diet will vary across different stopover/wintering sites and between autumn and spring; (5) cranes select and use staging/wintering sites with mudflat crabs when ambient temperature provides them with the opportunity to do so.22.1S. salsa saltmarshes, intertidal mudflats, reed marsh habitat, and artificial aquaculture fishponds , the Yellow River Delta Nature Reserve , and the Yancheng Nature Reserve .The activities of mudflat crabs around their burrows were filmed using two digital cameras . We randomly selected up to two crab burrows per day to film, with the camera positioned 0.5 m above the ground, to ensure an unobstructed view to the burrow entrance. Each recording lasted 8–9 hr from 8:00 a.m. to 16.00 hr or 17.00 hr. Recordings were made at YRD from November 7 to December 30 in 2014, and February 26 to March 28 in 2015, and at LRD from November 7 to November 29, 2014. No recordings were made during the spring of 2015 at LRD, because crabs were still in hibernation due to low ambient temperatures during Red‐crowned crane migration. In addition, no recordings were made at YNR because of limited logistical resources, but previous data have revealed that the Red‐crowned crane populations wintering there do feed on the crabs during the winter time . These site‐related daily maximum ambient temperatures were used to explore the relationship between diet composition and migration timing of Red‐crowned cranes.Burrow ambient temperature was recorded at the entrance of each crab burrow at a depth of 2 cm using the Tinytag Plus 2 temperature data logger . We used one data logger per site, with LRD temperature sampling conducted during September 16 to December 24, 2014 (this logger failed to work during the following spring season) and YRD sampling conducted during November 20 to April 4, 2015. We selected one burrow for temperature data recording and did not move the data logger to a different burrow each day as we suspected there would be minimal variation in ambient temperature between different burrows. Crab activity at the burrow entrance  was extracted from the video recordings using the Baofeng 5.0 digital player. Daily maximum and mean burrow ambient temperatures for each day were calculated to examine the influence of temperature on crab activity. Daily maximum ambient temperature data for the time period October 1, 2014, to April 30, 2015, for each site were downloaded from the website  or Siberian crane (Leucogeranus leucogeranus), and only on these occasions we did not collect any fecal samples to avoid any error with allocating samples to the different crane species. We limited the number of fecal samples to ≤3 samples collected from each foraging or roost site to reduce the potential for pseudo‐replication. Only newly defecated samples were collected from the ground using a sterilized spoon, and these were subsequently stored in a sample tube and taken back to the field station (<8 hr travel time) and stored in refrigerator at −20°C before analysis.We collected a total of 902 fresh fecal samples of Red‐crowned cranes from their foraging or roosting sites between 2011 and 2015  and LRD . Counts were conducted within the coastal 2.5r = .899, p < .001), we built these variables into the different models separately. We ran the GLMs with Poisson error structure and logit link function using the glm function included in the MASS package. We examined Wald test z scores to make inferences about each parameter estimate.Crab activity datasets from all sites were pooled for the analyses without considering inter‐site differences. To test our first prediction, that mudflat crabs become more surface active with increasing burrow ambient temperature, we first used Pearson's correlations to examine the relationships between maximum daily and mean daily burrow ambient temperature with the onset (time) of crab activity time, and with the mean percentage of active crab burrows. In addition, we fitted two generalized linear models (GLMs) with the percentage time of crab activity at burrow entrances and percentage time crab activity outside of burrows on the mudflat as the response variables, with site , season , daily maximum burrow ambient temperature and mean burrow ambient temperature included as predictor variables. Since both burrow ambient temperature response variables were positively correlated , we fitted the data using Pearson's correlations. To test our fourth prediction (crane diet would vary across different stopover/wintering sites and between autumn and spring migration seasons), we examined seasonal site differences in the frequency of the presence of crab remains and the percentage of crab remains in crane fecal samples using chi‐square tests. We pooled all fecal sample data since we hypothesized that there was little variation in the foraging microhabitat of Red‐crowned cranes across the S. salsa wetland and that crab specialization was a function of ambient temperature. Finally, to test our fifth prediction, that cranes select and use staging/wintering sites with mudflat crabs when the site ambient temperature provides them with the opportunity to do so, we examine the percentage of crab remains in all fecal samples with daily maximum and daily mean site ambient temperatures with Pearson's correlations. All statistical analyses were conducted using R 3.6.0 .To test our second prediction (that crab surface activity would be limited to an ambient temperature threshold), we used a piecewise linear regression model to explore the relationships between percentage time of crab surface activity (combining the activity time both on the burrow entrance and outside the burrow on the mudflat) and maximum or mean burrow ambient temperature using the R package  Muggeo, . Differe3Do mudflat crabs become more surface active with increasing burrow ambient temperature?n = 98) of the video samples of crab burrows revealed crab activity, either being recorded present on the entrance or walking outside of the burrow. Mean day time of the onset of crab activity was 11.41 ± 1.48 a.m.  and varied from 09.16 am to 15.07 p.m. The onset of crab activity‐starting time was or very nearly negatively associated with the respective of daily maximum burrow ambient temperature  and daily mean burrow ambient temperature  and daily mean burrow ambient temperature  but these increases were not significantly correlated , which was significantly lower in the autumn (11.1°C ± 4.2) than in spring . The break point estimation of the piecewise linear regression shown that the percentage of time that crabs were recorded as being active at the burrow entrance and also outside of the burrows increased when daily mean ambient temperature was 2.8 ± 2.4°C and the daily maximum ambient temperature reached 11.9 ± 2.2°C  in autumn (November 11–December 20) and a positive significant relationship between mean percentage of active crab burrows and date  in spring (March 4–April 2) at YRD. This suggests that crab activity decreased from early December and increased again from early March in the following year  and in the percentage of crab remains in crane fecal samples . Crab remains were the dominant food type from crane feces collected in the YRD both in spring and autumn, and in the autumn of LRD (>90% in the percentage of prey remains), but not in the spring at LRD  were found in fecal samples during the crane's spring staging stage at LRD . There was some variation in the diet composition of Red‐crowned cranes between the winter of 2011/2012 and 2014/2015 at YNR. At this site, crab remains only accounted for a small percentage (18.5%) of crane's fecal samples and less than 7% of the fecal contents in 2011/2012 winter, and no crab remains were found in crane feces in the YNR 2014/2015 samples. The dominant fecal remains were rice, snail, and shrimp in the 2011/2012 and 2014/2015 winters respectively at YNR  and daily mean site ambient temperature  and crab activity significantly increased at a burrow ambient temperature threshold of 12 ~ 13°C (prediction 2). Crab activity tended to advance with warmer ambient temperature, with the percentage of active burrows declining in the autumn and increasing again during the spring migration (prediction 3). Temperature‐related crab availability also has a significant influence on variation in the amount of crab prey found in crane feces between different staging/wintering sites and migration seasons (prediction 4). Finally, we also found a significant, positive relationship between the percentage of crab remains in the crane feces and local daily ambient temperature, and a degree of congruence between local ambient temperature and the arrival and departure timing of Red‐crowned cranes at our three sites. This suggests that temperature‐depended mudflat crab activity may provide an important time window for migratory Red‐crowned cranes (prediction 5).Limulus polyphemus) at Delaware Bay, USA, provides a critical nutrient‐rich dietary resource (eggs) for migratory populations of red knot Calidris canutus, ruddy turnstone Arenaria interpres, sanderling C. alba, and semipalmated sandpiper C. pusilla  for their spring department from Banc d'Arguin, Mauritania  is the only other crane species currently known to depend on crabs  . Clearly, both representing critical resources for migratory Red‐crowned cranes and identifying links between prey availability and migration timing across multiple stopover sites would provide more comprehensive data for improved migratory bird habitat conservation management in the region. Since mudflat crab represents a temperature‐related food resource, it will be critical to determine whether Red‐crowned cranes wintering in coastal China can adjust their migration timing to response to further variation in weather and/or phenology along their migration routes due to the predicted impacts of global climate change. Further data from their breeding ground regarding arrival times, territory establishment, and breeding success are needed to shed more light on these trade‐offs.The migratory timing of birds is multi‐mechanistic, determined by numerous biological, ecological, and environmental factors ; Data curation ; Formal analysis ; Funding acquisition ; Investigation ; Methodology ; Project administration ; Resources ; Software ; Supervision ; Validation ; Visualization ; Writing‐original draft ; Writing‐review & editing . Jing Zhang: Investigation ; Methodology ; Validation . Lingyu Chen: Investigation ; Methodology ; Validation . Huw Lloyd: Formal analysis ; Software ; Validation ; Visualization ; Writing‐review & editing . Zhengwang Zhang: Conceptualization ; Funding acquisition ; Project administration ; Validation; Writing‐review & editing.Figure S1Click here for additional data file.Figure S2Click here for additional data file.Tables S1‐S2Click here for additional data file.