In this way, the ability of the device to measure real-time variations of the hemodynamics parameters has been tested. The obtained high sensitivity of 0.012 V Pa−1 m−2, joint to the inherent biocompatibility and non-toxicity of the used materials, demonstrates that the device can successfully monitor the prosthesis functioning under different conditions, opening new perspectives for real-time vascular graft surveillance.Vascular grafts are artificial conduits properly designed to substitute a diseased blood vessel. However prosthetic fail can occur without premonitory symptoms. Continuous monitoring of the system can provide useful information not only to extend the graft’s life but also to optimize the patient’s therapy. In this respect, various techniques have been used, but all of them affect the mechanical properties of the artificial vessel. To overcome these drawbacks, an ultrathin and flexible smart patch based on piezoelectric Aluminum Nitride (AlN) integrated on the extraluminal surface of the prosthesis is presented. The sensor can be conformally wrapped around the external surface of the prosthesis. Its design, mechanical properties and dimensions are properly characterized and optimized in order to maximize performances and to avoid any interference with the graft structure during its activity. The sensorized graft is tested  In fact, despite the “failing-graft” has a hemodynamically compromised blood flow, no evident clinical signs are observed, and its continuous asymptomatic deterioration can result in thrombosis and possible ischemia of the treated area. Of note, the treatment of a thrombosed graft is a really complex process, which can lead to an increased postoperative morbidity and, in the worst case scenario, to patient mortality. A bypass surgery is mandatory when the graft is completely occluded, whereas an easier endovascular procedure for a local angioplasty is sufficient to restore the physiological blood flow in case of only partial occlusion2. However, even if most surgical practices have incorporated a follow up program of graft surveillance to increase the “failing-graft” recognition3, no single blood flow sensing technique has been adopted, in clinical practice, for real time monitoring of the prosthetic graft4. The follow-up surveillance after the implantation is currently based on ultrasound protocols, computed tomography scan and angiograms at defined time intervals in order to evaluate parameters such as graft flow velocity, variation in lumen dimensions and blood flow pressure5. However, the prevention of failures is not exceptionally efficient nowadays7. In this respect, the current techniques come with procedural morbidity, possible exposure to nephrotoxic contrast agent and are time consuming and expensive. All together, these factors limit their wide application and the potential benefit for early diagnosis4. To resolve these issues, a continuous monitoring of the vascular graft is extremely important to extend its lifetime, as well as to provide vital information for early failure warning. In this respect new solutions based on different analysis techniques have been investigated to monitor the graft status9. One approach involves the use a Micro-Electro-Mechanic System (MEMS) based pressure sensor, inserted in direct contact with the blood-stream, which causes the deformation of a strain gauge9. Although these pressure sensors can detect the hemodynamic variation with a good sensitivity, they require a voltage supply and traditional electronic components which alters the structure of the graft itself. Furthermore, the direct contact of the sensor with the blood can induce localized turbulences in the blood flow and can generate response to the “foreign body”. More recently, a new class of flexible sensors has been developed to be directly mounted on the external graft surface, so to follow the vascular graft expansion due to the pulsatile blood flow. To do this, a possible approach uses a flexible strain sensor composed of structured polydimethylsiloxane (PDMS) with an embedded conductive PDMS to measure graft wall deflection8. Yet, the dimensions of this sensor still affect the mechanical properties of the graft. An innovative approach is based on the piezoelectric effect. Indeed, the ability of piezoelectric based sensors to instantaneously generate an output voltage as an effect of response to rapid variations of dynamic pressure makes them very appealing for monitoring the blood flow. Different types of piezoelectric sensors have been used to evaluate different hemodynamic parameters for both on skin and implantable applications11. In particular, a sensor unit based on two polyvinylidene difluoride (PVDF) sensors (28 μm) encapsulated in Mylar® foil has been developed2. The system wrapped around the graft evaluates flow variations in both in vitro and in vivo models. The reported results show how the piezoelectric transduction is really useful to monitor hemodynamic variations and the presence of small occlusions in the graft, but the sensor dimensions with its high thickness still affects the mechanical behavior of the graft structure.A prosthetic vascular graft is an artificial conduit, designed on the patient’s arterial anatomy, to bypass a diseased vessel. Although recent technological advances have allowed improvements of prosthetic graft performances, failures can occur without any premonitory symptoms12, with high electromechanical coupling13 and intrinsic high resistivity14, making it one of the primary choices for the development of an implantable piezoelectric sensor. The whole fabrication process trades on traditional microfabrication techniques, allowing a simple engineering of the geometry and definition of the mechanical properties of the final device, to improve the matching with the structure of the artificial vessel, without affecting its behavior.Here we demonstrate an ultrathin, lightweight and flexible smart microsystem, based on piezoelectric Aluminum Nitride (AlN), integrated on the extraluminal surface of prosthesis, for the continuous monitoring of the vascular graft. The piezoelectric device, wrapped on the extraluminal surface of the graft, generates an electric response due to the wall deformation, providing important information about the blood flow hemodynamics in the artificial vessel. The proposed sensor is developed as a smart patch composed by the sensitive piezoelectric area, embedded between two electrodes and inserted in an ultrathin, flexible polymeric matrix made of polyimide (PI) and a thin layer of PDMS. Importantly, the AlN is inherently biocompatible and non-toxicad-hoc measurement set-up.This work encompasses the fabrication and characterization of the flexible smart patch, the finite element modeling (FEM) and shows the experimental results derived from the study of the graft-sensor system, exploiting an piezo) is then acquired to extract information about the status of the graft. In this work, the quality of the piezoelectric AlN deposited on polyimide was verified by means of morphological analysis and piezo force microscopy. The representative AlN flexible sensor – named PSP-20 (piezoelectric smart patch-20 mm2) –, having a piezoelectric area of about 4 × 5 mm2, was mechanically and electrically tested. Finally, the sensor was used to continuously monitor the status of a prosthetic graft.The flexible piezoelectric smart patch consists of a sensing element with a rectangular shape whose active area is composed of a multilayered stack consisting of aluminum nitride-interlayer (AlN-IL), molybdenum bottom electrode (Mo), piezoelectric aluminum nitride (AlN) and molybdenum top electrode (Mo). A dynamic external stress, applied to the sensing element by the cyclic expansion and contraction of the graft walls, allows the smart patch to generate a proportional electrical peak voltage by virtue of the direct piezoelectric effect. This voltage output  and scanning electron microscopy  were used. Figure θ–2θ scan mode, in which a sharp peak at 36° is observed, corresponding to (002) crystal orientation of the AlN hexagonal structure. A second, lower peak is also evident at about 33°corresponding to the (100) crystal orientation. The intensity ratio between these two peaks confirms the preferential growth direction along the c-axis, suggesting a well oriented AlN, grown by sputtering on flexible polyimide substrate and good piezoelectric properties of the thin film.The morphological structure and crystalline orientation of the AlN thin film deposited on polyimide substrate after the transfer process from the rigid substrate, were evaluated with particular attention to the grain orientation along the Visual inspection with SEM shows a good continuity through the different grains, as well as a clean separation between the layers Fig. .Piezo force microscopy  was used to evaluate the piezoelectric response of the deposited AlN. This technique exploits the application of a modulated AC voltage between the bottom Mo electrode of the tested sample and the conductive tip of the instrument, to measure the elastic deformation of thin film. Figure d33eff is measured as the slope of the piezoresponse amplitude versus applied voltage. The measured value of 5.0 ± 0.2 pm/V is higher than that reported for AlN sputtered on polymeric substrates16.From the piezoresponse amplitude curve Fig.  - carriegraft = 0.5 GPa), as desired17. Finally, using the DMA compression clamp tool, a periodic and impulsive normal force was applied in order to measure the response time of the PSP-20. A constant pressure of 50 kPa was applied by the use of an hemispherical tip whereas the sensor was connected to the oscilloscope . The corresponding generated voltage acquired showed that the sensor exhibited a very fast response to the impulsive applied force, whose value, being 0.84 µs, has been evaluated as the rise time between the 10% and 90% of the ascending edge , with an uniaxial tensile testing. In this test, the sensor has been subjected to a controlled tension, with a force range varied from 0 up to 1 N, with a ramp of 0.1 N/min. The considered sample has a rectangular shape with a thickness of 17 µm, a width of 9.57 mm and a measured length of about 17 mm. The Young’s modulus, extrapolated from the slope of the linear zone of the stress-strain curve  - previously measured by the DMA tensile testing - has been introduced in the FEM description in each PDMS/PI/AlN-IL/Mo/AlN/Mo stack layer materials data. Similarly, the PDMS membrane material data with a Young’s modulus of EPDMS ≈ 2 MPa, measured by DMA, was used. PVA material data were acquired from literature18 while its layer thickness was measured by profilometer (55 μm). Because of its wurzite structure, AIN has been assumed to be in-plane isotropic and the compliance matrix has been formulated accordingly. Consequently, the effective piezoelectric coefficient d33eff of the AlN layers, measured by PFM, has been inserted in the Strain-Charge matrix. Table Subsequently, a FEM model (built by COMSOL Multiphysics software) has been developed to verify the responsivity of the sensor to a pressure variation. The simulations have been set up combining structural mechanics with electrodynamics, describing the material piezoelectricity through a strain-charge matrix. The structure was defined, following the real sensor structure, by a superposition of the AlN-IL/Mo/AlN/Mo stack, embedded between two polyimide layers on a PDMS structural layer, with a thickness of about 20 μm. The whole patch is, then subsequnetly, attached to a PDMS deformable membrane (thickness 500 μm), used as substrate mimicking the soft system . Between the PDMS membrane and the smart piezoelectric patch, an adhesion layer of Poly (PVA) has been inserted in the simulations, in order to account for the glue to which the smart patch is attached. Nonlinearity in PDMS behavior has been introduced in FEM simulations. To better validate the simulations, the effective Young’s modulus of the stack  corresponds to an increase of the electrical response in terms of output voltage generated by the AlN, as reported in Fig. 20:contact is the contact area between the smart patch and the PDMS pushing membrane and α is a parameter that depends on the materials’ constant and on the stack’s deformation mechanics. These conditions have been replicated in the laboratory by applying the same pressure range on the PDMS supporting membrane, deforming the piezoelectric sensor and directly measuring the generated open circuit voltage with an oscilloscope. The obtained results, reported in Fig. −1 m−2.According to the literature21. The simulations have been set up exploiting structural mechanics describing the mechanical behavior of the patch using the “plate theory”22. In this respect the bending rigidity of the PSP-20 was described by the Eq.  ring with a thickness of 1 mm. To provide the model with realistic features, the effective mechanical properties of the stack  in order to perform a comprehensive characterization of the device response as a function of the graft behavior.23. As a matter of fact, this signal is useful for recording information about the blood pressure and the analysis of its waveform provides important clinical information. The pulse wave signal is commonly divided in two main parts defined as anacrotic phase (the rising edge of the pulse) and catacrotic phase  which corresponds to different moments of the cardiac cycle24. The corresponding curve is characterized by the presence of two peaks, related to systole (SP) and diastole (DP), respectively, and a dicrotic notch (DN) between them. A distortion in the wave profile and, in particular, a variation of amplitude and position of these parameters are significant signs of an undesired variation in the patient hemodynamics, such as atherosclerosis or different heart dysfunctions23. In this respect, recording specific alterations of these signals on vascular grafts allows retrieving information on the graft status and highly predictive signs of prosthetic failures. According to these considerations, an ad hoc measurement set up has been developed, in order to pump a flow into a sensorized graft and mimic the behavior of a real arterial blood flow. In particular, the continuous sequence of systolic and diastolic peaks with the corresponding dicrotic notch in between have been successfully simulated. Moreover, their specific amplitude and generation time have been varied properly. The setup, presented in Fig. 25. The solenoid valve was controlled by a power supply driven by a LabVIEW© script. In this way, a cyclic routine was achieved that controlled the water flow by properly varying the opening percentage and relative time interval of the valve channel, inducing the creation of previously mentioned parameters (both systolic and diastolic peaks and dicrotic notch). The output response of the sensor as a voltage signal has been acquired and recorded with an oscilloscope.Generally, the most common way to evaluate the hemodynamics in a vessel is to study the pulse wave signal Fig.  recorded(i)26.the variation of the peak-to-peak interval, closely related to the R-R interval of the electrocardiogram (ECG) and useful to define the duration of a complete heart cycle and the corresponding flow rate. Moreover, a continuous control of this parameter is fundamental because its variation can influence the usefulness of some indexes (to be discussed later)(ii)26 – allowing the recognition of modifications in the graft wall compliance due to the formation of a plaque inside the vessel. This index is defined as the ratio between the diastolic and systolic peaks, respectively defined as P2 and P1, (Eq. (24.the variation of the Augmentation Index (AI); this parameter provides information on the stiffness of a vessel – indeed, arterial stiffness is an important marker of cardiovascular risk and atherosclerosisP1, (Eq. ) of the (iii)DVP). This index is related to the transit time of pressure wave, from the central cardiovascular system to the periphery and its reflection. It is strictly correlated to AI and provides additional information about the mechanical properties of the vessel and the blood pressure  20,24,4(iv)27.the presence of different percentages of occlusion in the vessel, since restenosis appears to be the main cause of vascular graft failing after one yearThe PSP-20 output has been acquired in different hemodynamics conditions, properly replicated by the measurement set up by controlling the following parameters:Flow signals are reproducibly obtained from the sensor on the graft as a function of flow parameters. Firstly, the PSP-20 device attached to the graft Fig.  generate11.Despite the sensor measured response time (0.84 µs), the whole system (PSP-20 and graft) is able to detect frequency variations with an overall response time of about 250 μs, due to the damping effects induced by the graft deformation. This response time respects the common requirements for flow rate monitoring and it is comparable with the recently reported results about flexible piezoelectric sensors for blood flow analysis2/P1 ratio has been imposed inside the graft. For this purpose, the opening percentage of the solenoid valve has been varied accordingly to the peak amplitude variation of the pulse wave shape. In particular, a variation of P2 amplitude has been enforced. The variation of this parameter is more indicative in terms of clinical evaluation. Indeed, reduced compliance causes an earlier return of the reflected wave, and this phenomenon is commonly translated in a modification of the P2 amplitude. Considering that, when the elastic arteries become stiffer there is a consequent increase of the augmentation index value28 – which is expected for the artificial vessel that mimics the arterial properties and behavior - the P2 amplitude has been varied in order to modify the AI, starting from a value of about 0.45, compatible with a safe vessel, until 0.80, typical of a highly stiff one. The plot reported in Fig. In order to simulate the variation of the augmentation index, an induced change of the PDVP. A variation of the time interval between the P1 and P2 peaks has been imposed by modifying the opening time of the outlet channel of the solenoid valve. From the analysis of the shifted peaks, it is shown that the sensor can detect even small variations of the time interval between P1 and P2, without any undesired variation in the output voltage amplitude of the sensor, in good agreement with the imposed time step of the solenoid valve of about 50 ms  has been employed, allowing to strictly control the properties of the deposited film in terms of thickness and smoothness.The piezoelectric sensor used for the vascular graft monitoring is based on a thin films heterostructure Fig.  and fabr30, promoting a well oriented growth and a high continuity through the single nanograin of the piezoelectric AlN.A 4 μm-thick layer of polyimide  has been used as a substrate for the multilayered structure consisting of aluminum nitride-interlayer , molybdenum bottom electrode , piezoelectric aluminum nitride  and molybdenum top electrode  stack. The adhesion of the structure and texture orientation on the polymeric substrate has been guaranteed by the presence of the thin interlayer of AlN under the Mo bottom electrode2:Ar) at a working pressure of 2.8 × 10−3 mbar, with a deposition rate of 17 nm/min, under a DC-pulsed power of 750 W and 1000 W for AlN-IL and piezoelectric AlN, respectively. Mo layers are sputtered using a pure Mo target (99.95%) in Ar atmosphere under DC power of 200 W and a working pressure of 5 × 10−3 mbar, with a deposition rate of 10 nm/min. AlN interlayer and bottom Mo electrode are sputtered directly on the polymeric substrate in the first single deposition run. The thin films AlN-IL/Mo has been patterned in rectangular geometries by dry etching, using inductively coupled plasma (ICP) etching system with defined gas mixtures of BCl3 and Ar, properly optimized to selectively etch these two materials. AlN piezoelectric thin film and Mo top electrode are then deposited by the second deposition run and patterned by ICP, exploiting the same BCl3 – Ar gas mixtures of the previous etching processes.The main steps of the micro-fabrication protocol of the sensor are illustrated in Fig. An insulating film of polyimide is then conformally deposited to mechanically protect the active area and make it waterproof, preventing short-circuits. At this stage, the device is detached from the rigid support by dipping the sample in an acetone bath for 10 minutes in order to melt the PMMA sacrificial layer. The freestanding structure Fig.  is final2 plasma, through the top polyimide coating, in order to guarantee electrical connections. The freestanding sensor, is then glued on the graft using a PVA-based adhesive layer.To guarantee a perfect adhesion between this additional flexible layer and the PI based transducer, both surfaces are chemically treated: the PDMS top surface is functionalized in a solution of adhesion promoter ; in parallel, the device is dipped in potassium hydroxide (KOH) and hydrochloric acid (HCl) solutions to chemically activate the PI surface, and to promote the adhesion to the PDMS substrate. The total thickness of the final device, including the PDMS sheet, is about 20 µm, with an active area extension of 4 mm × 5 mm. Vias are opened, using O


Living donor kidneys with two arteries can be revascularized using various techniques depending on anatomy. We hypothesized that the revascularization technique could impact long-term outcomes. We retrospectively analyzed 1714 living donor renal transplants at our institution between 1999 and 2015. Three hundred and eleven kidneys had dual arteries, and these were categorized into 5 groups; end-to-side (n = 18), inferior epigastric artery (n = 21), direct anastomosis (n = 65), side-to-side (n = 126) and ligated (n = 81). We then compared the outcomes with that of a control group  using Kaplan-Meier and Cox regression analyses. Cox regression was adjusted by age, sex and race/ethnicity of donor and recipient, side of kidney, transplant period and recipient surgeon. Compared to the control group, the end-to-side group had increased all-cause graft loss  and death-censored graft loss , whereas the other groups did not. Our study shows that 10-year overall survival and death-censored graft survival were significantly worse for end-to-side arterial reconstruction than for other techniques. Alternative techniques to the end-to-side method should be used for accessory arteries that require revascularization. In living donor renal transplantation, kidneys with multiple arteries are encountered in about 7–28% of cases . AlthougVarious techniques are used to revascularize multiple arteries depending on the anatomy and institutional or surgeon preference , 3. In oHowever, whether the type of arterial reconstruction method for multiple renal arteries has an influence on long-term graft and patient survival is not known . We hypoWe retrospectively analyzed 1750 living donor renal transplantations in which kidneys were retrieved by laparoscopic donor nephrectomy at our institution between 1999 and 2015. To analyze the pure impact of each reconstruction technique, we excluded 36 cases from the analysis: 30 with more than three arteries, five with an arterial graft, and one for which recipient data was unavailable. Two artery cases (n = 311) were divided into five groups for analysis: end-to-side (n = 18), inferior epigastric artery (n = 21), direct anastomosis (n = 65), side-to-side (n = 126) and ligated (n = 81), and compared with control group . Demographic data were collected from patients’ electronic medical records. This study was approved by the Institutional Review Board of the University of California, San Francisco . The review board waived the requirement for informed consent for this research. None of the transplant donors were from a vulnerable population and all donors or next of kin provided written informed consent that was freely given.All nephrectomies were done using an intraperitoneal pure laparoscopic donor nephrectomy approach as previously described –7. The dThe Chi-squared or Fisher’s exact test was used for categorical data analysis and the Mann-Whitney U test or Kruskal-Wallis H test was used for continuous data. Post-hoc analyses for categorical and continuous data were all adjusted using the Bonferroni correction and Z-test. Two-tailed p-values ≤0.05 were considered statistically significant. Values are expressed as median [interquartile range] unless otherwise specified.The Kaplan-Meier method was used to analyze overall graft survival , death-censored graft survival, and patient survival. Cox regression analysis was used to analyze the impact of each reconstruction technique on survival, adjusted for age, sex and race/ethnicity of donor and recipient, side of kidney, transplant period and recipient surgeon. Interaction analysis was done thereafter, using recipient sex and body mass index (BMI).All analyses were done using SPSS version 22 . The authors have followed the suggestions of the Strengthening the Reporting of Observational Studies in Epidemiology (STROBE) statement guidelines for reporting observational studies.Baseline characteristics, including age, race, sex and BMI for both donor and recipient did not differ among donors and recipients across all the groups . In addiThe groups did not significantly differ with respect to the frequency of acute rejection and delayed graft function. The end-to-side group had the highest rate of ureteral complications . Critical graft thrombosis causing immediate graft loss  occurred only in the direct anastomosis and side-to-side groups, as well as the control group . PartialThe 10-year survival rates for the end-to-side group were significantly lower than those for the control group for overall graft survival , death-censored graft survival  and patient survival ; this was not true for the other techniques . The altThe causes of graft loss within 10 years are shown in To our knowledge, this is the first report showing significantly inferior long-term graft survival of a specific arterial reconstruction method—end-to-side—for living donor kidneys with two arteries. Long-term graft survival was previously reported to be similar for single and multiple arteries , 4, 9–20In our study, arterial thrombosis rates were lower than reported by other groups . Our cenA unique aspect of the end-to-side technique is that the main artery diameter is presumably sufficient to supply the original distribution; the main artery of the kidney with two arteries is smaller than that of the kidney with a single artery. Adding another smaller artery upstream of this main artery is technically difficult and would potentially cause stenosis and thrombosis in the smaller artery. The end-to-side technique is also more vulnerable to stenosis, as the diameter of the larger artery is generally smaller and needs to supply the entire kidney. This contrasts with the inferior epigastric reconstruction or separate anastomosis techniques, which provide an independent blood supply, and the larger artery only needs to supply its original portion of the kidney. Furthermore, the thrombus and/or hyperplasia of the smaller artery could spread in the main artery, producing future stenosis and chronic ischemia of the main artery as well. However, without angiography, the relative contribution of thrombosis or stenosis cannot be determined. Importantly, post-transplant renal artery stenosis is strongly associated with inferior overall graft survival .Several ancillary findings in our study support the concept of arterial insufficiency and/or insufficient renal mass as a contributor to the increased graft loss in end-to-side reconstruction. First, a significantly higher incidence of non-specific chronic failure was observed among the reasons for graft loss for end-to-side group, possibly due to the chronic ischemic injury to the graft. Secondly, adjusted hazard ratios of all-cause graft loss and death-censored graft loss were higher in patients who would benefit from more renal mass, namely male patients and patients with BMI ≥30.Both the inferior epigastric artery method and the direct anastomosis to recipient iliac artery appear to be good alternatives to the end-to-side method because long-term graft survival was similar to that in the control group. The inferior epigastric artery is often available and can be anastomosed after the main renal artery is reperfused, which contributes to reduced ischemia time and reduced incidence of delayed graft function . HoweverA recent study of 27 kidneys  with ligOur study has the usual limitations of a retrospective, single-institution series. Additional limitations included the lack of immunosuppressive drug information  for recipients. The cumulative incidence of acute rejection was the same among the groups, but we could not exclude the possible confounding effect due to long-term immunological damage. However, the cause of graft loss was not clearly immunologic as discussed above. In general, most kidney transplant patients at our center receive aspirin perioperatively, especially after any complex vascular reconstruction, unless there is a contraindication. We do not use heparin or any other form of anticoagulation on a routine basis. Further statistical analysis such as propensity-score matching to mitigate the long-term survival data was not done because the baseline characteristics of the five groups were all similar to those of the control group.In conclusion, our study shows that 10-year overall survival and death-censored graft survival were significantly worse for end-to-side arterial reconstruction than for single artery group. While we acknowledge the pitfalls of retrospective surgical studies, these types of data are not available in registry data. Our results clearly suggest that, if possible, the end-to-side reconstruction technique should not be used. Further prospective and multi-center studies would be ideal but are extremely difficult to conduct; however, addition of more granular reconstruction data in large databases such as the SRTR database may be useful to examine whether outcomes differ for specific arterial reconstruction methods for kidneys with two arteries.


The molecular combing technique was used to dissociate the nanostructural units of starch granules from the starch fragments after a gelatinization process. With the help of atomic force microscopy (AFM), we observed that some nanostructural chains were just ﬂowing out of the granules. It proves that there are substantive nanostructural units in the starch granules, a phenomenon not previously observed, so these nanostructural units were defined as suspected intermediates. Furthermore, we conclude that blocklets of starch granules are formed through twisting or distortion of nanochains. D-glucopyranosyl units; in the macrocosm, the diameter of starch granules falls in the micron size range [In the microcosm, natural starch granules are composed of 1/4- and 1/6-linked α-ze range . In foodze range ,4,5,6,7.et al. [D-glucosidic bonds) form the amorphous layer. Many data about starch structure have been obtained previously due to the continuous research for a long time. On the basis of these data Gallant et al.  put forwWhen observing the surface of starch granule with AFM, Baldwin found some protrusions , so in GMolecular manipulation is a new technology in nanoscience. It is a kind of ﬁxed-point operation to change the uniformity of the whole macromolecule. Its main aim is to stretch the bio- and poly-macromolecules from the clew or the leptospira into straight chains in order to directly observe their structure using some special instrumentation such as AFM ,11. One In this study, AFM was used to observe the chains of starch nanounits during the gelatinization process. The corresponding images can be seen in In These six figures reveal the whole gelatinization process of starch nanochains. Before describing the gelatinization process from these six figures, the nanounit chains, the salient points and conjunction areas should be made clear. From the present data, the height of starch nanounit chains should be below 3.0 nm, and the diameter of blocklets should be 20–500 nm ,16,17,18The salient points of starch nanounit chains should be the crystalline areas, which are formed by the helixes of side chains of amylopectin macromolecules. The conjunction areas of the starch nanochains should be amorphous areas, which are formed by the branch points  of amylopectin macromolecules. This proves that starch nanounit chains contain crystalline areas and amorphous areas, alternated with each other. Finally we deduced the mechanism of the gelatinization process of starch nanounit chains: First the structures of the starch nanounit chains that are just flowing out of granules are very tight. The amorphous areas could not be observed by AFM and the whole nanounit chain has a uniform height and width. Next, when the nanounit chains come into contact with hot water, they can begin swelling, just like the starch granule swelling process. As a result, the structure of the nanounit chains becomes loose, and the amorphous areas can be observed with AFM. Second, gelatinization begins to happen in the amorphous areas. Water molecules penetrate slowly into the crystalline areas from the amorphous areas and untie the helixes in the crystalline locations. As a result, the crystalline areas becomes slowly disorganized and its structure becomes loose. This can explain why the height of salient points becomes lower and lower from -7 Ω·cm-1); Atomic Force Microscope (AFM) .Cornstarch ; double distilled water (quartz evaporator, conductivity < 1.0 × 101. Gelatinization : a starc2. Molecule combing : starch 3. Test: all samples ready to be tested are observed using the AFM, working in tapping mode in air. In this paper the technology of molecular combing has been used to dissociate starch nanounit chains from starch fragments and these nanostructural unit chains were defined as suspected intermediates because they distributed widely in the starch fragments. From the analysis, we put forward a gelatinization model for starch nanounit chains which proves that these suspected intermediates contain crystalline areas and amorphous areas. It deduces that blocklets should be the result of the twisting or distortion of nanochains.


Ficus species were sequenced using next‐generation methods, and the results indicated that closely related host species had clearly differentiated FEF assemblages. Furthermore, host phylogenetic proximity was significantly correlated with the similarity of their FEF assemblages. In the canonical correspondence analysis, eleven leaf traits explained 32.9% of the total variation in FEF assemblages, whereas six traits  were significant in the first two dimensions of ordination space. In the multiple regression on distance matrix analysis, 21.0% of the total variance in FEF assemblage was explained by both host phylogeny and leaf traits while phylogeny alone explained 7.9% of the variance. Thus, our findings suggest that both evolutionary and ecological processes are involved in shaping FEF assemblages.Foliar endophytic fungi (FEF) are diverse and ubiquitously associated with photosynthetic land plants. However, processes shaping FEF assemblages remain poorly understood. Previous studies have indicated that host identity and host habitat are contributing factors, but these factors are often difficult to disentangle. In this study, we studied FEF assemblages from plants grown in a botanical garden, enabling us to minimize the variation in abiotic environmental conditions and fungal dispersal capacity. FEF assemblages from 46  Host plant plays a key role in shaping the foliar endophytic fungi assemblage, host phylogenetic proximity was significantly correlated with the similarity of their FEF assemblages, and 21.0% of the total variance in FEF assemblage was explained by both host phylogeny and leaf traits while phylogeny alone explained 7.9% of the variance. They haNicotiana species and their endophytes  and found no significant influence. Thirdly, which leaf traits contribute more to FEF assemblage is still unclear. Leaf traits are key factors in host filtering for foliar endophytic fungi. Arnold et al.  contains about 80 species and varieties of the genus in an area of 1.3 ha, including all of the six subgenera, multiple life forms, and two sexual systems (monoecious and dioecious). The collection is managed without the use of pesticides or fungicides; thus, all plants are exposed to natural fungi and herbivorous insects. In addition, most of plants have historic records, including provenance and propagation history, most of the species used in this study are native to the Yunnan province, and all the species were introduced into the garden before 2011 Table .2.2Ficus individual during the wet season in 2015, when most fig species produce new leaves. Ten well‐expanded mature and healthy leaves were sampled from each individual , leaf water content (%), total C content (g/kg), N content (g/kg), pH, toughness (g), leaf tannin content per mass (%), latex volume per area (μl/cm2), latex water content (%), latex tannin content (g/L), and latex alkaloid content (g/L)  Table . Leaf trL) Table ; plant iL) Table , and posL) Table ; secondaL) Table ; leaf pHL) Table ; and theL) Table . Other l2) was measured with digital photographs using ImageJ (Version 1.46r). Leaf material and latex were oven‐dried at 40°C until a constant mass was reached. SLA was estimated as the ratio of leaf area to leaf dry mass. Water content was estimated as the difference between the wet and dry mass for both leaf and latex. Dry samples were then sent to the XTBG Central Laboratory for measuring chemical properties. Total C and N were determined by Dumas combustion analysis . The pH meter was calibrated against buffer solutions  before each measurement series. Toughness was measured with a device modeled after a design by Feeny  was collected from the leaves, dissolved in 90 μl of purified water, and diluted 10 times. The mixture was filtered through a 0.45‐µm membrane. The extract was fully blended with 50 ml acetone for 40 min and filtered through a medium speed quantitative filter. Then, 1 ml of filtrate was mixed with 2.5 ml of sodium tungstate–phosphorus‐molybdenum acid mixed solution and 5 ml of carbonate. The tannin content was measured by a colorimetric method using the Folin–Denis reagent  ethanol (5 s), 0.5% NaClO (2 min), and 70% vol/vol ethanol (2 min), and finally rinsed three times for 1 min in di H2O  HTotal DNA was extracted from the subsamples of pooled leaves using the HP Plant DNA Mini Kit (OMEGA Bio‐tek) and stored at −20°C. Then, the ITS1 region  and PCR (1 μl sterilized Milli‐Q water substituted for the DNA template); no amplicons were detected by electrophoresis. Each triplicate reaction was pooled, and PCR cleanup was performed using Bioteke purification kits (BioTeke) according to the manufacturer's instructions. After testing the cleaned PCR products for concentration using a DNA assay 2100, and for size via gel electrophoresis, amplicon samples were standardized by molarity and pooled into a single sample. Libraries were built using a combination of PKR Truseq Adapter Kit  and KAPA Hyper Prep Kit (Roche) and then quantified by quantitative real‐time PCR (QPCR) according to the KAPA Library Quantification Kit's protocol (Roche). The PKR Truseq Adapter Kit provides 12 unique adapters, each containing one unique index, which are compatible with Illumina's MiSeq platform and allow demultiplexing the data postsequencing by the platform software. The libraries were pooled following Illumina's guidelines and sequenced on Illumina's MiSeq platform, using the MiSeq Reagent Kit V2 (Illumina) which allows a maximum read length of 2 × 250 bp by paired‐end sequencing. In this study, we did not conduct negative controls for the sequencing, due to our pilot study indicated a very limited influence of extremely rare sequences in downstream analysis, compared with abundant sequences in samples with much higher concentration of amplicons.2.5http://hannonlab.cshl.edu/fastx_toolkit/). Primers and barcodes were trimmed using USEARCH 10.0 and served as raw sequences to produce the operational taxonomic units (OTU) table. Amplicon sequence data were deposited in a Sequence Read Archive under accession number SRR8675934  and Dryad Digital Repository. To obtain high‐quality sequences for clustering OTUs, the reads were further filtered by USEARCH 10.0 with a constant maximum expected error threshold of one. Sequences with homopolymers with length >10 bp or with a total length of <100 bp were discarded according to variance of ITS1 length  and permutational multivariate analysis of variance (PERMANOVA)  was used to visualize the dissimilarity among subgenera, using the “capscale” function.The Mantel test was used to test the relationship between the dissimilarities in FEF assemblages and the phylogenetic distance among host species, using the “mantel” function in the ecodist package  from 8,096,909 raw sequences were used to construct the OTU list, and 5,353 OTUs were grouped at 97% similarity. All the raw sequences were used to build the OTU table. The sequence‐based rarefaction curves based on the observed OTUs were asymptotic Figure , indicat3.1Conocycea, subsection Urostigma, and the F. auriculata species complex , especially the FEF assemblages of all individuals for 13 host species clustered in separate groups.The results of both the ANOSIM and PERMANOVA analyses showed that FEF assemblages differed significantly among host species in each of the three groups including subsection ex Table . Pairwisea Table  and Urosma Table , and homex Table . DCA orda Figure a, subseca Figure b, and thx Figure c. Hierarx Figure  showed t3.2r = .25, p = .001, n = 46; Figure R = 0.32, p = .001, Figure The Mantel test showed that the dissimilarity in FEF assemblages was significantly correlated with the phylogenetic distance separating host plants . The other 10 traits did not present significant phylogenetic signals (Table The CCA was used to test and illustrate whether leaf traits mediate the variation in FEF assemblages Figure . In totals Table .We determined which single leaf trait could predict the similarity between FEF assemblages. The Euclidean distance of SLA, leaf pH, and toughness was significantly correlated with the similarity among FEF assemblages Table , both be4Ficus species and that the leaf traits studied explained a considerable proportion of the variation in FEF assemblages among host species. Thus, this study suggests that both evolutionary and ecological processes shape FEF assemblages in Ficus.This study provides evidence that both host species identity and phylogenetic signal play a significant role in the assemblages of FEF in different F. auriculata complex. While genetically distinct (Wei, Kobmoo, Cruaud, & Kjellberg, Conocycea and Urostigma. For each Ficus species sampled in this study, different individuals were usually characterized by very similar FEF assemblages (Figure Different host species harbored different FEF assemblages Figure . This was Figure . This res Figure , in whicThe Mantel test revealed a significant correlation between FEF similarity and the phylogenetic proximity of host species Figure . A phyloThe distinct chemical composition and physical traits of leaves might underlie the influence of hosts on FEF assemblages. In this study, 32.9% of the variation in FEF assemblages was explained by the 11 leaf traits studied. This result is consistent with that of the culture‐based study by Vincent et al. , in whicFicus individuals have been introduced into the garden for at least four years (Table Although most fig species included in this study are native to the Yunnan province in China, many of them were introduced into the botanical garden as seedlings collected from different sites in the wild. Thus, there is a possibility that FEF recruited from native habitats have influenced the FEF assemblages in our study. However, the influence might be extremely limited. rs Table , providirs Table  FEF fromFicus species, suggesting that FEF are very diverse in the garden, which is consistent with findings in the tropics (Arnold, More than 5,000 nonsingleton OTUs were observed from 46  Arnold, . However Arnold, . In thisHost‐mediated filtering is the cornerstone of research on plant–FEF interactions. By characterizing FEF assemblages of 46 congeneric species in a small area, and thereby minimizing the variation in abiotic environmental conditions and fungal dispersal capacity, this study provides strong evidence that host identity has an important influence on shaping FEF assemblages and that leaf traits contribute to explaining the effect of host identity. Host phylogeny was also found to exert a significant impact on shaping FEF assemblages. Our findings suggest that both evolutionary and ecological processes play a role in shaping FEF assemblages, which is fundamental to understanding the mechanisms for FEF assemblage establishment.Authors declare no conflict of interest.J. L. and J. C. designed the study, J. L. and J. Z. collected the data, J. L. performed the statistical analyses, J. L. and J. C. drafted the manuscript, and all the authors approved the final version. Click here for additional data file.


On receiver operating characteristic curve analysis, a graft weight of 580 g was identified the cutoff for stratifying outcomes. Recipients transplanted with a graft weighing ≥580 g had significantly better outcome as compared with other recipients (p = 0.001). Moreover, SPLT remarkably provided a better survival benefit for recipients than those on the LT wait-list (p < 0.0001). Conclusions: Given the considerable incidence of wait-list mortality, SPLT for two adult recipients should be encouraged whenever possible to increase the donor pool and benefit patients awaiting LT. Nonetheless, caution should be taken with a smaller graft weight owing to the risk of early graft loss.Background: Organ demand for liver transplantation (LT) is constantly increasing. Split liver transplantation (SPLT) is an ideal option for increasing the number of available liver grafts for transplantation and ameliorating organ shortage to a certain degree. However, SPLT for two adult recipients is still not broadly applied. Methods: We retrospectively analyzed the outcomes of SPLT for adult recipients at a single center. All donor, recipient, and transplantation factors were thoroughly investigated to clarify factors affecting patient outcomes after LT. Results: One hundred consecutive adult SPLTs were performed during the study period. Early mortality and 1-year mortality occurred in 21 and 31 recipients, respectively. On multivariate analysis, graft weight ( Since the first liver transplantation (LT) a half century ago, LT has evolved into a common operation in many transplantation centers nowadays. ,2 With tSPLT was initially proposed as a method in which an adult liver graft is divided into a smaller graft for a pediatric recipient and a bigger graft for an adult recipient . To dateNonetheless, deceased organ donations for LT are extremely scarce in Asian countries, including Taiwan . TherebyAmong 1093 LTs, a total of 100 consecutive adult SPLTs were performed between July 2003 and June 2019 at our transplantation institute. All medical records of donors and recipients were retrospectively reviewed, and data were collected for analysis with approval from the Institutional Review Board of Chang Gung Memorial Hospital . Owing to the retrospective nature of the study, the need for informed consent from patients was waived. The study protocol complied with the ethical standards established by the Declaration of Helsinki with respect to confidentiality of patient data. Importantly, no organs from executed prisoners were used in this study. Further, patients registered in the wait-list for LT and patients transplanted with a whole liver graft during the corresponding period were analyzed for survival comparison.Generally, the potential donor should be hemodynamically stable with acceptable cardiopulmonary function, no uncontrollable bacterial infection, and no malignant neoplasm. All potential donors were thoroughly evaluated for eligibility for liver donation with biochemical tests and hepatic ultrasonography. The donor should also have acceptable liver functional reserve and only mild fatty change in the hepatic parenchyma.The decision to split liver grafts was based on the evaluation of the donor, evaluation of the liver graft, and recipient allocation from the national organ sharing program, as previously described ,14. BrieSeparation of the hepatic parenchyma for split liver grafts was generally performed in situ, as previously described, except for one case in which hemodynamic instability occurred in the donor during graft procurement . During All hepatic parenchyma transections were performed using the Cavitron Ultrasonic Surgical Aspirator , and the Pringle maneuver for inflow vascular control was not applied during hepatectomy. All vessels between the right and left lobes were divided, and the main tributary veins of segments 5 and 8 in the right lobe were temporarily clamped with bulldog clamps and released when graft perfusion was commenced. The hepatic parenchyma was split down to the anterior surface of the inferior vena cava (IVC) when possible, and the hepatic duct was divided. Subsequently, the liver graft was perfused with histidine-tryptophan-ketoglutarate  solution and allowed to recover.The right and left hemi-liver grafts were completely separated on the back table, and all vascular structures were divided A. GeneraGenerally, all potential recipients were routinely evaluated for laboratory tests of infectious microorganisms, color Doppler cardiac sonography, whole-body computed tomography (CT), and panendoscopy prior to list on the wait-list or transplantation. An informed consent document that detailed the indication and risk of LT were well explained to the recipient and were to be signed before transplantation. All recipient operations started with total hepatectomy with IVC preservation. The right hemi-liver graft was orthotopically implanted using the piggyback technique B. The leAfter transplantation, all recipients were transferred to the intensive care unit for postoperative management. Biochemical tests and hepatic ultrasonography were performed at regular intervals. The postoperative immunosuppressive regimen consisted of a combination of methylprednisolone, tacrolimus, and mycophenolate mofetil and was adjusted according to the clinical status of the recipient.2 test or Fisher’s exact test, as appropriate. In order to include all important factors, variables with a p-value < 0.10 from univariate analysis were selected for multivariate analyses by enter procedure using logistic regression models. All statistical analyses were performed using the statistical software SPSS version 20.0  for Windows. A p-value of <0.05 was defined as statistically significant.The endpoint outcome measures were early mortality within 3 months, 1-year mortality, and overall survival (OS). The OS was calculated from the date of LT to the date of death or the end of the study, and all causes of deaths were counted as events. Survival curves were generated using the Kaplan–Meier method and compared using the log-rank test. Continuous variables were compared using the Mann–Whitney U test, and categorical variables were compared using the χIn accordance with the organ donation regulation in Taiwan, all livers were procured after the donors were declared brain dead by specialists. None of the liver grafts in this study were procured from a donor with a non-beating heart. After SPLT, the median follow-up period was 17.2 months . During the follow-up period, 54 (54%) recipients were still alive by the end of this study. Early hospital mortality (from 1 to 86 days after LT) occurred in 21 (21%) recipients. Ten recipients died during the period from 3.6 to 11.8 months after LT. The remaining 15 deaths occurred 1 year after SPLT.p-value < 0.1 in univariate group comparison analysis, including graft type (p = 0.004), graft weight (p = 0.004), and GRWR (p = 0.054), were selected into multivariate analysis. The results of multivariate analysis showed that graft weight  was the only independent factor for the occurrence of early mortality. p < 0.1)—graft type (p < 0.001), graft weight (p < 0.001), cold ischemia time (p = 0.039), and GRWR (p = 0.007)—were strongly related to 1-year mortality. Furthermore, multivariate regression analysis of these four factors revealed that only two factors—graft weight (p = 0.070) and cold ischemia time (p = 0.069)—showed borderline correlation with 1-year mortality  curve to determine the best cutoff point related to the outcome of SPLT. A graft weight of 580 g was identified as the most optimal cutoff point for stratifying outcomes. The area under the ROC curve was 0.71 (95% CI = 0.59–0.82) for early mortality. The sensitivity and 1-specificity were 71.4% and 30.3%, respectively. Furthermore, recipients who were transplanted with grafts weighing <580 g and encountered early mortality were analyzed. The clinical features and major events leading to early mortality for these patients are summarized in p < 0.0001). However, the survival curve of SPLT was comparable with whole liver transplantation, in which the 1-, 3-, and 5-year survival rates for whole liver recipients were 73.4%, 66.9%, and 66.9%. (p = 0.198) The clinical features of patients who had listed in the wait-list or underwent LT is summarized in The 1-, 3-, and 5-year survival rates in all recipients after SPLT were 68.3%, 60.5%, and 59.0%, respectively. In the corresponding period, 165 adult patients (132 men and 33 women) who had undergone LT with a whole liver graft and 995 adult patients (750 men and 245 women) who were registered in the wait-list for LT and had not received any type of LT were analyzed for survival comparison. Of those, 816 patients (82%) had no opportunity to undergo LT and died during the waiting period , among whom 593 patients (59.6%) died within 1 year of being in the wait-list. The remaining 179 patients were still awaiting LT  by the end of this study. The 1-, 3-, and 5-year survival rates of patients in the wait-list were 40.2%, 16.3%, and 8.0%, respectively. The comparison of survival curves showed that SPLT remarkably provided a survival benefit for patients who had been indicated for LT and registered in the wait-list (p = 0.001).The comparison of survival curves in patients after SPLT based on graft weight is illustrated in This study describes the experience of 100 SPLTs performed in adult recipients at a single center in the setting of organ donor shortage. With the achievement of this milestone after a long evolution of the procedure, we aimed to thoroughly assess the beneficial effects and risk factors of SPLT in order to provide additional information for optimizing the strategies in this approach. Importantly, the results showed that graft weight was strongly associated with the initial success of SPLT. Further, SPLT for two adult recipients saved patients from the long waiting period for LT, which is characterized by a high mortality rate.It is well known that extreme organ shortage is the most important hindrance to organ transplantation worldwide. Specifically, deceased organ donors are extremely scarce in the East Asian region. As a result, LDLT is increasingly performed nowadays to overcome the shortage of liver donations for transplantation . HoweverHistorically, a number of reasons may be responsible for the less frequent use of SPLT for two adult recipients. First, splitting the liver into two hemi-liver grafts during donor operation is technically demanding ,25. SecoAdditionally, the allocation system and accessibility of the transplantation facilities are important considerations with respect to the practicability of SPLT. Generally, SPLT for a pediatric recipient and an adult recipient is better accepted by most transplantation programs when the split liver grafts are allocated to two different centers. On the other hand, most transplantation centers are reluctant to use leftover hemi-liver grafts gained from SPLT for two adult recipients. In such circumstances, trust and collaboration between different centers are essential for successfully sharing hemi-liver grafts. Currently, there is no formal allocation system, particularly for sharing hemi-liver grafts from SPLT between transplantation centers in Taiwan. As a result, 46 pairs of SPLTs were performed at our institute and only 8 hemi-liver grafts were shared with other transplantation centers.However, the accessibility of the facilities in terms of space and manpower might provide a large challenge to performing SPLT for two recipients at the same time. Accordingly, three teams need to be activated for simultaneous organ procurement and recipient operations. There were only four transplantation surgeons at our institute by the year 2008; however, the number has doubled to date. Although the outcomes of SPLT were obviously not affected by the different eras before and after 2008 in this study, the greater number of transplantation team members largely provided relief to surgeons from the stress of the transplantation operations. Moreover, sleep deprivation and fatigue have been concerns with respect to physician performance and patient outcomes. ,33. ThusNonetheless, although the surgical technique may be the most important factor for the accomplishment of LT, there remain several concerns affecting the long-term graft and patient survival. Generally, determining the feasibility of LT usually starts from donor and recipient matching, and GRWR has been a common indicator for assessing the suitability of a partial liver graft for transplantation. Although a minimum GRWR of 0.6% could be successfully utilized in LDLT, a GRWR ranging from at least 0.8% to 1.0% is recommended for split liver grafts from deceased donors ,24,34. HOn the other hand, graft weight was a significant factor leading to early mortality in adult SPLT in this study. Although many equations were utilized for calculating the SLV, none of them were able to estimate the true liver volume and weight. Generally, liver volume is not equal to liver weight because many factors could affect the specific gravity of the liver. This discrepancy was observed not only in this study but also in other previous studies ,38. To oThe limitation of the study might be its retrospective nature with a small number of patients. Although SPLT for two adult recipients was performed whenever possible, the number of patients remain small because of the scarcity of deceased donors during the long study period. Additionally, this study might be considered impractical in the clinical setting despite establishing a cutoff graft weight (580 g). Meanwhile, the odds ratio of a smaller graft for early mortality is only 1% to 2% difference in the study. Although the study cautions that a graft weight less than the cutoff value may pose a risk of early graft loss, SPLT should not be discouraged under this concern. Moreover, the small-for-size problem might be overcome in the near future owing to the growing knowledge and experience of small grafts. Although the outcome of SPLT was comparable with that of whole liver transplantation in this study, the overall outcome might be criticized for the inferior survival as compared with the outcomes of other reports from whole liver transplantation. However, the important observations might provide additional information for guiding decision-making in the selection of a suitable deceased donor for SPLT to two adult recipients.In summary, LT has evolved into a common operation with satisfactory outcomes nowadays. However, SPLT for two adult recipients is still not a popular practice worldwide. Given the considerably high incidence of wait-list mortality, SPLT for two adult recipients should be encouraged to increase the donor pool and benefit patients awaiting LT.


High phenotypic variation was recorded for the two traits, which positively correlated. Moreover, both the parameters correlated with seed length (positive correlation) and seed width (negative correlation). A correlational selection according to human preferences has been hypothesized for the two starch traits and grain size. In addition, human selection has been proposed as the causal agent even for the different phenotypes related to starch and grain size showed by the tropical and temperate japonica accessions utilized in this study. The present GWAS led to the identification of 11 associations for RS on seven chromosomes and five associations for AAC on chromosome 6. Candidate genes and co-positional relationships with quantitative trait loci (QTLs) previously identified as affecting RS and AAC were identified for 6 associations. The candidate genes and the new RS- and/or AAC-associated regions detected provide valuable sources for future functional characterizations and for breeding programs aimed at improving rice grain quality. Rice quality is mainly related to the following two starch components, apparent amylose content (AAC) and resistant starch (RS). The former affects grain cooking properties, while RS acts as a prebiotic. In the present study, a Genome Wide Association Scan (GWAS) was performed using 115 rice  Rice quality is primarily related to the composition of the main component of grains, starch. The percentage of amylose on total starch, referred as apparent amylose content (AAC), is the first factor affecting the cooking properties of rice grains [Waxy (Wx) gene and, as extensively reported, different alleles at this locus correspond to different AAC percentages [Wx locus, as follows: aWx, associated with high AAC levels and bWx, carried by low AAC rice cultivars [Wx intron 1, which causes an alternative splicing, thus reducing the production of functional enzymes in rice endosperm and determining the occurrence of low AAC amounts [Wx allele associated with AAC variation is represented by inWx, frequently in accessions belonging to an aromatic group and to tropical japonica exhibiting an intermediate AAC [Wx exon 6, which determines a Tyr to Ser substitution in the active site of the enzyme, reducing the specificity for its substrate [Rice provides more than 1/5 of calories for human worldwide populations  is a dietary fiber representing the part of starch that is not digestible by the enzymes of the stomach and the small intestine, but remains intact until reaching the colon where it acts as a prebiotic . The benindica varieties have higher levels of RS (from 0.1 to 3.2%) with respect to japonica varieties (from 0 to 1.6%). In addition, in rice, as in other cereals, RS is affected by starch composition; for example, it positively correlates with AAC [amylose extender mutant IR36ae, exhibiting an inactive SBEIIb that results in alterations in the amylopectin fine structure, has a higher level of RS in cooked grains with respect to the wild type [There is a wide genetic diversity for RS in rice. RS levels vary from 1.2% to 9.0% of cooked polished rice dry weight in commercial normal-starch rice varieties, across different growing locations . Evaluatwith AAC ,23,24. Mwith AAC , whereaswith AAC . In factild type . A higheild type . A negatild type .amiRNA) was associated with increased AAC, with only modest changes in the total starch (TS) content and with an increase in the proportion of long and intermediate amylopectin chains [Since for most rice cultivars the RS content is generally under 3% in milled grains, which is not enough to confer the associated health benefits ,29, manyn chains . This len chains . In accon chains . Correspn chains .SBE loci in determining RS content in rice was confirmed by the mapping of sbe3-rs, a quantitative trait locus (QTL) for RS on rice chromosome 2, using a mapping population obtained from the cross between the high RS japonica mutant Jiangtangdao 1 and the indica cultivar Miyang 23 [SBE3, which encodes for a SBE, was suggested as the candidate gene for the QTL region [The importance of the iyang 23 . In thisL region .indica rice [ssIIIa allele, resulting from the introduction of a premature stop codon, which was responsible for an increased RS production. Furthermore, it was shown that the high RS phenotype was also dependent on the high expression of the aWx allele [In addition to SBEs, other enzymes implicated in starch metabolism have been demonstrated to affect RS in rice. High RS rice mutants, indeed, displayed lower activity of SBEs and ADP-glucose pyrosphorylase (AGPase), catalyzing the first rate-limiting reaction of starch biosynthesis, and higher activities of starch synthases (SSs) and starch de-branching enzymes (DBEs), with both affecting the number of short-chain amylopectins . A map-bica rice  identifia allele .2 populations [qRS7-1 and qRS7-2) in brown and polish rice, using a F2–3 family derived from the cross between the two rice genotypes Gongmi No. 3 and Diantun 502 [japonica and indica genotypes, led to the discovery of four RS-related QTLs, explaining from 10% to 13% of total phenotypic variation. Two QTLs were located on chromosome 6 and corresponded to the Wx and the SSIIa loci, respectively, while the other two QTLs were localized to chromosomes 8 and 9, proximal to the SBEI and ADPase small subunit I encoding loci, respectively [SSIIa gene results into a shortened amylopectin chain length distribution and negative pleiotropic effects on SBEIIa and SBEIIb activity [Despite the evidence described above, to date, limited information is available on the genetic bases controlling the variability of RS in rice. By segregation analysis of a population using the RS111 mutant as the common high-RS parent, SSR markers on chromosomes 8 and 6 were found to be associated with RS in two Fulations . Moreoventun 502 . More reectively . In barlactivity , therebyjaponica rice population, including 115 accessions genotyped by the genotyping by sequencing (GBS) method, with the aim of expanding the understanding about the genetic basis of the determination of RS content and AAC.In the present study, a GWAS approach was performed on a Non-cooked brown rice of the 115 rice genotypes were analyzed for TS and RS content. On average, the TS content was 73.43 ± 1.69%, with Graal showing the lowest value (70.40%) and Lagrue the highest one 81.73%; . RS contA hierarchical clustering, based on the two variables, RS and RSTS, and using Euclidean distances, separated the 115 rice genotypes into three main clusters , as follThe AAC values were evaluated in a previous work  on the sp < 0.001) positive correlations were detected between RS and RSTS (R = 0.999), RS and AAC (R = 0.464), and RSTS and AAC  was higher with respect to the other clusters, while the lowest AAC mean value (18.52 ± 2.85%) was recorded for cluster 1, corresponding to the low RS class . The WilAccording to previous results , AAC corSignificant differences were recorded for some grain shape related traits calculated for the low and medium AAC classes, particularly for SWSL and NSWNSL, for which higher values were calculated for the low AAC class, with respect to the medium one .p-values (The same type of correlation as for AAC was found between RS (and RSTS) and the grain shape-related traits, even considering the lower p-values . Accordip-values .No significant correlations were recorded between TS and the traits related to grain shape .japonica accessions (45 accessions) with respect to the temperate ones (70 accessions)  , while nate ones .2 values were calculated for the two traits (0.97 for RS and 0.99 for AAC). Together with the continuous distribution of the phenotypic data related to RS and AAC, this makes our dataset particularly relevant for studying the genetic bases of the two starch traits in japonica rice through a GWAS approach. A lower genetic effect was instead recorded for TS (H2 = 0.33).The ANOVA analysis revealed significant genotypic variability for RS and AAC, with no substantial effect of replicates . As confR = 0.805, p ≤ 0.001), confirming overall, that the determined hierarchy based on RS content in uncooked rice was also maintained after cooking.To assess relationships among RS evaluations in brown non-cooked and brown cooked kernels, twenty genotypes, randomly selected among the low, medium, and high RS classes, were subjected to cooking tests to evaluate the RS content after the cooking process. The range of variability was from 0.026 to 0.241% and from 0.185 to 1.223% in not cooked and cooked samples, respectively . ResultsWx gene [Wx intron 1, and TBGI270316, lying inside the Wx exon 6. Based on the rice genome size of 373 Mb [The panel of 115 rice accessions was genotyped by GBS. After pruning, 28,445 SNPs were utilized for the GWA mapping of RS, AAC, and TS-related regions. The SNP panel was enriched by the addition of the following two SNPs identified within the f 373 Mb , the avejaponica subgroups  of a PCA analysis, which accounted for 24.31% of total genetic variability . The secnd Graal . All thend Graal  and comeA mean LD decay of 1,247 kb  was calculated over the physical distance in the whole panel of 115 accessions .A GWAS experiment was conducted to identify the loci associated with the variation in AAC, RS, and TS content among the 115 rice accessions.No associated regions were identified for TS, likely because of the low genetic variability related to this trait in our genetic material.The list of all the RS- or AAC-associated SNPs is reported in 2) from 11% to 29%  were discovered on chromosomes 1, 2, 3, 5, 6, 8, and 11 , with −l% to 29% .Only three RS_MTAs were defined by single SNPs, as follows: RS_MTA2, related to SNP S2_9401106 on chromosome 2; RS_MTA3, defined by S3_15491784 on chromosome 3; and RS_MTA6_2, corresponding to S6_24395396 on chromosome 6 . The oth2 = 0.29), defined by 167 significant SNPs (peak marker S1_32380039), located from 31,035,257 to 35,233,919 bp, and belonging to a LD block from 30,645,440 to 33,485,305 bp (The most significantly RS-associated MTA was the chromosome 1 RS_MTA1 (−log10(p) = 9.15 and R5,305 bp .Chromosomes 5 and 6 contained the highest number of RS-related MTAs  and TBGI270316  = 5.99 and R2 = 0.19; For AAC, 5 MTAs were discovered on chromosome 6 (−log10(p) from 5.99 to 3.90 and Rt of AAC , as follWx SNPs revealed the presence of three haplotypes in our rice collection, as follows: GA , GC (28 accessions), and TA  and grain shape was previously observed [aWx (G) allele were significantly different from those recorded for the bWx (T) accessions, particularly for SWSL that was higher for aWx genotypes, in comparison to the bWx ones  were included in the one recorded for RS_MTA6_1 .https://shigen.nig.ac.jp/rice/oryzabase/) was conducted for the genes annotated as implicated in starch metabolism and in the determination of amylose content present in and located at the level or in proximity (±1.20 kbp) of the RS- and/or AAC-related MTAs and their LD blocks. The analysis was enriched by literature searching. Results are summarized in To find out candidate genes affecting RS and/or AAC levels, a search on the Oryzabase database  was located within the RS_MTA1 associated region on chromosome 1 , encoding for an adenylate translocator which facilitates the transfer of extraplastidial synthesized ADPglucose into amyloplasts in maize [Among the candidate loci affecting RS content, mosome 1 ; Table 2mosome 1 ; Table 2in maize , was incqAC5 QTL, affecting the amylose content in rice [indica rice genotypes and, among these, a SNP in the promoter region of the Wx gene contributed to haplotypes discriminating samples into different amylose classes [indica varieties, represents a major hot spot associated to the rice glycaemic index (GI), involving 26 genes including Wx [A searching for previously mapped QTLs involved in the determination of starch properties and co-localized with the RS- and/or AAC-related MTAs detected in the present work revealed a positional correspondence between RS_MTA6_1 and AAC_MTA6_4 and the RS-associated locus chr06_6168586 detected by Bao et al.  Figure . The qAC in rice , was fou classes . In addiuding Wx . Both thuding Wx ,46  Protein). AGPase, which catalyzes the first rate-limiting reaction of starch biosynthesis, was demonstrated to have an active role in RS determination by the fact that, as mentioned above, high RS rice mutants showed lower activities of SBE and AGPase and higher activities of SS and DBE [CRCT expression and the starch content in the leaf sheath increased in CRCT overexpression lines while they decreased in knockdown lines [CRCT might indirectly affect the RS content because of its effect on starch synthesis genes.The analysis of genes implicated in starch metabolism and located at the level or in proximity of the RS-associated QTLs led to the identification of three candidate genes affecting the RS content. The following two of them were positioned inside RS_MTA5_3: The  and DBE . CRCT waOsBT1-3  was located at the level of the RS_MTA6_2 LD block. The BRITTLE1 gene encodes for an adenylate translocator, which facilitates the transfer of extraplastidial synthesized ADPglucose, the substrate of SSs and GBSSI, into amyloplasts in maize [BRITTLE1 caused a reduction in the amount of starch produced in the endosperm [OsBT1-3 locus might reduce the ADPglucose available for SSs in amyloplasts, with alteration of the amylose/amylopectin ratio and consequently changing in the RS amount. On these bases, OsBT1-3 might represent a genuine candidate for RS determination.in maize . Mutationdosperm ,55. ThusDRUS2  gene, encoding a receptor-like kinase [DRUS2, together with its closely related gene DRUS1, affects sugar utilization during rice reproductive development, probably by stimulating the expression of the gene encoding for the UGP2 (UDP-glucose pyrophosphorylase 2) enzyme, which catalyzes the interconversion between starch and sucrose metabolites [DRUS2 on starch metabolism and RS can be also postulated on rice seeds.The e kinase , was fouabolites , and thuabolites . By thisWx gene, the main determinant of AAC [Wx was also found in a recent GWAS study on rice eating and cooking characteristics , using 174 and 84 indica and japonica rice genotypes, respectively [qAC5, which was located in the LD block recorded for RS_MTA5_3 [Wx gene contributing to haplotypes related to different amylose classes [Wx [Wx in determining rice grain healthy properties.As expected, the most significant association with AAC was discovered for the SNPs located at the level of intron 1  and exon 6 , respectively, of the t of AAC ,6,7. In ectively . This anS_MTA5_3 , thus su classes  and a masses [Wx  were botAn additional positional correspondence with known starch-related QTLs present in literature was observed between RS_MTA6_1 and AAC_MTA6_4, which co-localized with the RS-associated putative locus chr06_6168586, detected in Bao et al. , furtherWx SNPs utilized in the present study were demonstrated to be reliable tools for AAC prediction in breeding programs [In a previous research aimed at improving the marker-based predictability of AAC, the two programs . Three dprograms . AccordiWx intron 1 SNP (TBGI270314) was strongly associated with SW and LW as aWx genotypes  showed thinner grains with a higher length/width ratio, with respect to bWx  [aWx genotypes with respect to the ones recorded for the accessions carrying the T allele. All these data represent a further confirmation of the hypothesis that correlational selection for grain shape and AAC potently marked the evolution of rice according to customer’s requests [In addition, the As previously indicated, in the present work, significant correlations were recorded between RS and grain shape-related traits. These observations were enforced by the discovery of correspondences between two of the RS_MTAs and QTLs associated with grain shape-related traits mapped in previous research, as follows: RS_MTA5_3 co-localized with the NSL5-1 and NSWLR5-2 QTLs, associated with naked seed length and the ratio between naked seed width/length, respectively ; RS_MTA6No candidate genes or co-localizations with known starch-related QTLs were discovered for RS_MTA2, RS_MTA3, RS_MTA5_2, RS_MTA6_3, RS_MTA8, and RS_MTA11 or for AAC_MTA6_5, indicating that they could represent new QTLs, contributing to the variation of grain starch-related traits, and can be considered new sources for RS or AAC determination.japonica genotypes well adapted to European agro-climatic conditions, even though they were bred in different countries, as follows: A total of 40 accessions in Italy, 25 in the USA, 8 in Argentina, 11 in Portugal, and the remaining in other locations . All analytical measurements were performed in two replicates and, because of the predominant genetic influence on starch properties , they weRS content was determined on 100 mg of milled sample (particle size < 1 mm) according to the AOAC (2005) Official Method 2002.02, using the Resistant Starch Assay Kit . After the hydrolysis with KOH 2M at 4 °C, RS was measured by adding the GOPOD reagent to 100 μL of the sample and following the procedure adopted for TS, described above.The cooking test was performed following the procedure of Ranghino , slightlAAC values used in the present work were determined in a previous study on the same materials here used, where the adopted procedures for AAC determination are also detailed .The traits related to seed morphology used here were recorded on the same materials in a previous work , in whic2) was estimated from the variance components, obtained by fitting both replications and genotypes as random terms as H2 = σ2g/(σ2g + σ2e), where σ2g is the genotypic variance component and σ2e is the residual variance component.Frequency distributions of phenotypic data were tested for normality using the Shapiro–Wilk test in R environment. Two-factor ANOVA was carried out using JMP v.8 software  to evaluate differences for TS, RS, and AAC between genotypes. Broad sense heritability , was utilized to compare the mean values of the traits related to starch or grain shape among the different RS clusters, the temperate and tropical Hierarchical clustering was used to create relatively homogenous groups of rice genotypes using two variables, RS and the ratio between RS and TS (RSTS), as classification criteria. The cluster analysis, based on Euclidean distances, was carried out by the JMP v.8 software .https://rapdb.dna.affrc.go.jp/download/irgsp1.html), was filtered using the PLINK program [All the accessions were genotyped by sequencing according to a pipeline described previously . The ini program . MarkersWx gene , were added, for a total of 28,447 SNP markers. The genotyping of the two SNPs was performed by direct sequencing as described by Biselli et al. [Two additional SNPs, associated with the A random subset of 9996 SNPs (833 markers/chromosome) was utilized for the analysis of genetic stratification in the rice panel. Two different methods were adopted, as follows: A principal component analysis (PCA) performed by the Tassel v5.2.0 software  and a ph2) analysis was performed, as described by Volante et al. [2 between unlinked loci was set as 0.2. The physical distance corresponding to a LOESS value of 0.2 was assumed as the extent of LD.The linkage disequilibrium  method [https://cran.r-project.org/web/packages/qqman/index.html) was utilized to draw Manhattan plots and Q-Q plots for each trait. Single-SNP associations were considered true positive when a peak of multiple SNPs was detected at lower −log10  in the Manhattan plot. SNP clusters in full LD with the same p-value were considered as peak regions. The regions associated to each trait were aligned with the results of the Haploview analysis described above.GWAS analyses were performed for the three starch-related traits, RS, AAC, and TS, according to Volante et al. . Tassel ) method . The R phttps://shigen.nig.ac.jp/rice/oryzabase/), considering the gene onthologies “starch metabolism” and “amylose content”, and genes described in literature to affect RS or AAC, located at the level of or in proximity to the MTAs detected.For each trait, the regions ranging from −120 kb to +120 kb around each associated marker (corresponding to an average LD decay of 0.5 estimated on the LOESS curve) were screened for candidate genes. The identification of candidate genes for each trait was performed by selecting the genes annotated in the Oryzabase database (japonica rice collection. A significant positive correlation was found between AAC and RS, according to the observation that AAC represents one of the main determinants of RS in rice seeds [japonica accessions belonging to our rice collection indicate that the correlational selection for AAC and grain shape, and maybe RS, had probably marked the different grain characteristic between tropical and temperate japonica cultivars according to human preferences.Considerable phenotypic variation for three grain quality-related traits, represented by RS, AAC, and TS, was present in the analyzed ce seeds , and betce seeds . The assThe GWAS analyses conducted for the starch-related parameters considered in this study identified 11 and 5 MTAs related to RS and AAC, respectively. Two AAC-associated MTAs corresponded to one RS_MTA, thus highlighting the correlation between AAC and RS. Candidate genes affecting starch properties were located at the level of or in proximity to five of the identified starch-related MTAs, thus providing a list of genes that can be exploited to dissect the genetic bases of the MTAs. The new candidate genes and the new RS- and/or AAC-associated regions detected provide valuable sources for future functional characterization of starch-related genetic determinants and for breeding programs aimed at improving rice grain quality.


The results show that whether or not the four large air holes far away from the geometry center of the PCF are filled with analyte has no influence on the wavelength sensitivity of the sensor which means those holes can be replaced by small air holes. The wavelength sensitivity can be tuned by adjusting the sizes of the other large air holes which are as for liquid holes. The dynamic detection range of the refractive index (RI) is from 1.33 to 1.51. In particular, high linearity is obtained in the range of 1.44 to 1.51. The sensitivity is as high as 6021 nm/RIU when the liquid holes are the smallest. When liquid holes are tangential with the envelope of first layer air holes, the wavelength sensitivity is 4028 nm/RIU, and the coefficient of determination (R The surface plasma is excited on the surface of the metal–dielectric interface which is exceedingly sensitive to changes in the refractive index (RI) of the analyte. A. Hassani and M. Skorobogatiy proposed that the photonic crystal fiber (PCF) sensor plated gold film can detect minute changes in the RI of the analyte . The polZhou et al.  studied 2 is 0.99822 when the RI of analyte varies from 1.44 to 1.51, so that the sensor has both high sensitivity and good linearity.In this paper, a dual-core PCF structure is proposed, and the influence of parameters is studied and then the structure is improved. The numerical simulation results show it has no effect on wavelength sensitivity whether the four large air holes far away from the PCF geometry center are filled with analyte or not. As well, the sizes of those holes have no impact on the sensor. The sizes of the other large air holes which are used for liquid holes has an effect on wavelength sensitivity. The smaller the liquid holes are, the higher the sensitivity is. To make the sensor desirable for applications, adjusting the diameter of liquid holes is essential to reduce the filling difficulty. A solution is that liquid holes are tangent with the envelope of the first layer air holes. The wavelength sensitivity is 4028 nm/RIU and Rc) from the center is 3 μm. The diameter of the B holes nSiO2λ is the wavelength of the incident electromagnetic wave, and A1 = 0.696166300, A2 = 0.407942600, A3 = 0.897479400, B1 = 4.67914826 × 10−3 μm2, B2 = 1.35120631 × 10−2 μm2, B3 = 97.9340025 μm2, respectively. The dielectric constant of gold is given by the Drude model [n) varies from 1.33 to 1.51.Here, n = 1.46 (left) and n = 1.49 (right) when the A holes are filled with analyte and the B holes are empty, and the diameters of them are d1 = d2 = 2.6 μm. When the RI of an analyte is 1.46, the dispersion relationships intersect at point b , which results in incomplete coupling. In contrast, when the RI of the analyte is 1.49, the imaginary part curves of them are very close, and the real part difference is minimum at point e, which is called the avoided crossing point or the anti-crossing point. It can be considered that they undergo energy exchange intensively at point e and the anti-crossing effect takes place; this is known as complete coupling. It should be noted that the curves are depicted in the same color in the right illustration of The energy loss is caused by the appearance of surface plasmon resonance (SPR) between the gold film and the analyte. The absorption peaks change with the RI of an analyte. The analyte can be measured by judging the position of the absorption peak. The two adjacent core modes interact with each other, which is described as superimposition of electric fields, so they are called supermodes. The in-phase supermodes for which electric field distribution is in same direction are selected as the fundamental mode because their loss is less than that of other supermodes . Figure n) led to the shift of the resonant peak. Loss is defined, according to ref. [λ is the wavelength of the incident electromagnetic wave and effn is the effective RI of the fundamental mode.Variation in RI αWhen the A holes were filled with analyte and the B holes were empty and both had diameters of 2.6 μm, and the change in loss curves with n varied from 1.33 to 1.46 and from 1.47 to 1.51, as shown as n = 1.40 was not consistent around the phase matching point (653 nm), which shows that the energy in one core was greater than that in the other core. The coupling between the two adjacent fundamental modes led to a change in the complex amplitude of the light field in each core, so one of the two mutual coupling fundamental modes was disturbed by the other fundamental mode, especially when energy transferred from the fundamental mode to the SPP mode. Consequently, the two power flow distributions of the two cores were different, which is depicted as n = 1.45 and n = 1.50 in n ≥ 1.47) was much greater than the loss in incomplete coupling. For example, the fundamental mode energy was converted into the plasma mode with n = 1.50 at the avoided crossing point (λres = 969 nm) in the third column of As is shown in the first column of 1 values on the sensor are discussed. As shown in 1 was 2.6 μm, which was used as a reference to compare with other cases. For cases where the d1 was fixed at 2.6 μm, the scatter plots of the RI wavelength for B holes were filled with analyte (green) and not filled with analyte  are coincident in the left of 1 = 1.2 μm (blue) and d1 = 2.6 μm  were almost coincident as well. So, the impact of the B holes on the resonant wavelength is negligible; that is, the holes far away from the geometry center of the PCF have an insignificant influence on the wavelength sensitivity.The effects of filling the analyte and having different dlossα1 is the loss peak of the reference when the B holes are not filled with analyte, and the d1 is 2.6 μm.Here, the relative difference of the loss peak is defined as1 was 2.6 μm (green). The relative loss difference value was less than 5% in the case of the incomplete coupling (n ≤ 1.46), and the loss with analyte was greater than without analyte in the simulated B holes. Similarly, the relative difference in the loss peak was larger in the complete coupling than in the incomplete coupling when the B holes did not contain analyte and the d1 was 1.2 μm (blue). Therefore, the B holes have an effect on the loss peak only to a small extent in complete coupling, and their effect on the resonant wavelength is negligible. From the viewpoint of fiber fabrication, it is clear that the diameter of the B holes is 1.2 μm, in which case the B holes are the same as other small air holes of cladding. The following analysis is based on the condition where the B holes did not contain analyte and the d1 was 1.2 μm.The right diagram in λres is the resonance wavelength.The A holes, as in liquid holes, have a great influence on the sensitivity. Using wavelength interrogation, the wavelength sensitivity can be calculated in accordance with ref. :(4)SλR(RIU2) was 2.6 μm and 1.2 μm, respectively, are shown in 2 = 2.6 μm was smaller than that at d2 = 1.2 μm—the larger the d2, the smaller the sensitivity. The illustrations describe the dispersion relationships between the fundamental mode and the SPP mode at n = 1.47. The resonance peak occurred in the avoided crossing point at d2 = 2.6 μm, while the peak appeared in the phase matching point at d2 = 1.2 μm. In fact, the resonance peak at d2 = 1.2 μm appeared merely in the phase matching point with the RI of analyte increasing from 1.33 to 1.51. That means that the strength of coupling between the fundamental mode and the SPP mode of the sensor with small liquid holes was much less than that with larger liquid holes. Supercontinuum broadband source (SBS) was used as the light source because the designed sensor works at visible and near infrared wavelengths. As shown in The RI wavelength curves for when the diameter of the A holes (d2) was 1.2 μm, the A holes were the same as other small air holes. The sensitivity was 6021 nm/RIU and the R2 was 0.99841. When the d2 was 2.6 μm, the A holes were nearly tangential with the other cladding air holes. The sensitivity was 3539 nm/RIU and the R2 was 0.99873. The linearity of the two was good, and the sensitivity of the former was higher than that of the latter. However, the smaller diameter made filling with analyte difficult. Taking into account the reduced filling difficulty and improving the sensitivity of the sensor, the value of d2 was determined to be between 1.2 μm and 2.6 μm. As shown in the right of 2 was 1.72 μm; that is, the A holes were tangential with the envelope of the first layer cladding air hole. The diameters of d1 and d3 were both 1.2 μm in this setting. The d2 is increased by 43.33% compared to the smallest liquid holes. The wavelength sensitivity was as high as 4028 nm/RIU and the R2 was 0.99822, with n changing from 1.44 to 1.51, which shows that the sensor has high sensitivity and good linearity. If the spectral minimum resolution (Δλmin) is 0.1 nm, the resolution of the sensor is 2.48 × 10−5 RIU. This structure not only makes it easier to fill with analyte, but it also has high sensitivity.Several types of sensors that have been investigated in recent years are listed in 2 was 0.99822 when the n varied from 1.44 to 1.51, which means it has both high sensitivity and good linearity.The influence of the parameters of the dual-core PCF was numerically simulated, and an optimized structure was proposed. The dynamic detection range of the sensor was from 1.33 to 1.51, in which the optimum detection range was shown to be from 1.44 to 1.51 because of the high sensitivity and good linearity in this range. The holes far away from the geometry center of the PCF had little impact on the performance of the sensor, and the diameter of the liquid holes affected the sensitivity of the sensor—the smaller the diameter of the liquid holes, the higher the sensitivity, and the maximum sensitivity was 6021 nm/RIU. It is convenient to fill analyte by adjusting the diameter of liquid holes so that they are tangential to the envelope of the first layer cladding holes. In this case, the sensitivity was 4028 nm/RIU and the R


Distinguished colleaguesWe founded the Euroasian Gastroenterological Association 23 years ago in Ankara. Our aim has been to introduce scientists and the science of Central Asian countries with the West, to form a Euroasia science bridge and create a new scientific platform, in other words a new international scientific association. The first event of this new scientific vision took place on May 21-25, 1997 in Baku, the capital of Azerbaijan. We brought together many scientists from Turkey, Azerbaijan, Russia, Uzbekistan, Kazakhstan, Kyrgyzstan, Turkmenistan and Georgia. When we look back after all those years, we are happy to see the importance of our initiative in 1997.That was just a dream 23 years ago. Today we see that it has come through. We decided to organise international scientific congresses with a view to achieve our goals and objectives. We have organized 16 international congresses, three single topic conferences and three international conferences so far. These are as follows:1. 1997 Baku-Azerbaijan, 2. 1998 Almati-Kazakhstan, 3. 1999 Antalya-Turkey, 4. 2000 Tashkent-Uzbekistan, 5. 2001 Bishkek-Krygyzhstan, 6. 2003 Baku-Azerbaijan, 7. 2004 Ankara-Turkey, 8. 2005 Tbilisi-Georgia, 9. 2006 Baku-Azerbaijan,10. 2007 Isfahan-Iran, 11. 2008 Baku-Azerbaijan, 12. 2011 Baku-Azerbaijan, I. STC 2013 Yalta, Crimea, 13. 2013 Baku-Azerbaijan, II. STC 2014 Ljubljana-Slovenia, 14. 2014 Magusa-Nothern Cyprus, III. STC 2015 Bucharest - Romania, 15. 2016 Antalya-Turkey, 1st Conference of EGA 2016, 16. 2017 Mostar-Bosnia Herzegovina, 2nd Conference of EGA 2017 Almaty - Kazakhstan, 3rd Conferance of EGA 2017 Antalya-Turkey.We are going to organize two Euroasian congresses this year. These will be held in Split-Croatia (17th) and Baku-Azerbaijan (18th).We have achieved the impossible with scarce resources. We will see the importance of these congresses clearly in the next years. We believe that our young colleagues will take this further.A few years ago we decided to publish a journal in order to achieve our objectives. To this end Dr. Sheikh Mohammad Fazle Akbar from Japan, Prof. Mamun Al Mahtab from Bangladesh and I began to publish Euroasian Journal of Hepato-Gastroenterology with Jaypee Brothers Medical Publishers (P) Ltd. . Numerous articles have been published among papers submitted by scientists from 37 different countries in the world. We applied for the well-known American index, PubMed and finally entered PubMed and PMC in December 2017. This was yet-another achievement for our association. Lately, we applied for International Scientific Index (ISI) at the end of 2018. I believe that the application will be accepted shortly.We organized a very fruitful congress in Mostar-Bosnia Herzegovina last year with the help of Prof. Enver Zerem and Prof. Milenko Bevanda. This year we are going to organize the 17th Euroasian Congress of Hepato-Gastroenterology and Surgery at Split, Crotia on March 27-31 with the kind support of Prof. Neven Ljubicic, Prof. Miroslav Smunic and many other Croatian scientists.This year also the 18th Euroasian Congress of Hepato-Gastroenterology and Surgery (Azerbaijan) will take place in the beginning of October at Baku in the memory of Prof. Büyükkişi Agayev, one of the pionners of our Euroasian vision. I would like to thank our respected Azerbaijani colleagues in advance, including Prof. Geray Geraybeyli, the rector of the Azerbaijan University of Medicine and Prof. Rauf Agayev.I will be expecting your support and contributions in taking our scientific vision further and offer my kind regards to all of my colleagues in Euroasia.Prof. Hasan ÖzKANPresident of Euroasian Gastroenterological AssociationEditor in Chief of Euroasian Journal of Hepato-GastroenterologyHead of Hepato-Gastroenterology DepartmentAnkara University School of Medicine, Turkey