Transplantation represents the optimal treatment for many patients with end-stage kidney disease. When a donor kidney is available to a waitlisted patient, clinicians responsible for the care of the potential recipient must make the decision to accept or decline the offer based upon complex and variable information about the donor, the recipient and the transplant process. A clinical prediction model may be able to support clinicians in their decision-making. The Kidney Donor Risk Index (KDRI) was developed in the United States to predict graft failure following kidney transplantation. The survival process following transplantation consists of semi-competing events where death precludes graft failure, but not vice-versa.We externally validated the KDRI in the UK kidney transplant population and assessed whether validation under a semi-competing risks framework impacted predictive performance. Additionally, we explored whether the KDRI requires updating. We included 20,035 adult recipients of first, deceased donor, single, kidney-only transplants between January 1, 2004, and December 31, 2018, collected by the UK Transplant Registry and held by NHS Blood and Transplant. The outcomes of interest were 1- and 5-year graft failure following transplantation. In light of the semi-competing events, recipient death was handled in two ways: censoring patients at the time of death and modelling death as a competing event. Cox proportional hazard models were used to validate the KDRI when censoring graft failure by death, and cause-specific Cox models were used to account for death as a competing event.The KDRI underestimated event probabilities for those at higher risk of graft failure. For 5-year graft failure, discrimination was poorer in the semi-competing risks model , but predictions were more accurate . Calibration plots were similar regardless of whether the death was modelled as a competing event or not. Updating the KDRI worsened calibration, but marginally improved discrimination.Predictive performance for 1-year graft failure was similar between death-censored and competing event graft failure, but differences appeared when predicting 5-year graft failure. The updated index did not have superior performance and we conclude that updating the KDRI in the present form is not required.The online version contains supplementary material available at 10.1186/s41512-023-00159-9. For many patients with end-stage kidney disease, transplantation represents the optimal treatment. The demand for deceased donor kidneys in the United Kingdom (UK) greatly outweighs availability . It is tIn the United States (US), the Kidney Donor Risk Index (KDRI), proposed by Rao et al. , is usedThe scientific and clinical practices underpinning the delivery of transplantation services have evolved over time. Further variation exists between different units and countries. As such, prediction models developed in a particular country may not be reliably applicable to populations in other countries in the future. It is therefore essential to externally validate proposed prediction models when considering their use in different populations and to revisit these validations over time –5.We sought to validate the predictive performance of the KDRI in the UK kidney transplantation population. In our systematic review , we founThe survival process following transplantation consists of semi-competing events, where a terminal event precludes the observation of a non-terminal event, but not vice-versa. Specifically, in the context of kidney transplant survival outcomes, once a patient has died, we can no longer observe whether they experience graft failure. However, if a patient suffered graft failure, then we could still observe their death. In the existing literature on prediction models for graft failure, death is often not treated as a competing event, rather graft failure is censored by death or they are combined to predict a composite event.The original KDRI defined graft failure as the earliest of graft failure or death. Predicting a composite outcome assumes that predictors have the same effect on both outcomes of interest , and in The aim of this study was to externally validate the KDRI in the UK kidney transplantation adult population. Additionally, we aimed to explore whether modelling death as a competing event, rather than censoring recipients at the time of death, influences the predictive performance of the KDRI. Furthermore, we assessed whether updating the KDRI was required to improve predictions for graft failure.This study was reported in accordance with the Transparent reporting of a multivariable prediction model for Individual Prognosis or Diagnosis (TRIPOD) statement .This was a cohort study based on a registry database collected by the UK Transplant Registry (UKTR) and held by NHS Blood and Transplant (NHSBT). Recipients were transplanted in the UK between January 1, 2004, and December 31, 2018. Recipients were followed-up until March 31, 2021.Adult recipients (aged 18 years and above) of a first, deceased donor, single, kidney-only transplant were included. Where recipients have had multiple transplants within the study period, only their first one was used for analysis. Recipients of en bloc, or multiple organ transplants (such as combined kidney and pancreas transplants), were not included.We assessed the performance of the KDRI for predicting graft failure 1 year and 5 years following kidney transplantation. Graft failure was defined as the time from transplantation until either return to dialysis or re-transplantation. In light of the semi-competing events, recipient death was handled in two ways: censoring patients at the time of death and modelling death as a competing event. The original KDRI was intended to predict graft failure; however, the model was developed to predict a composite outcome of time to the earliest of death or graft failure.Recipients with both event time and indicator missing were excluded from the analysis. Missing values for donor height, weight, ethnicity, history of hypertension, history of diabetes, cause of death (cerebrovascular accident or not), creatinine value and hepatitis C virus status were imputed using multiple imputations with chained equations , assumin12.78% of the patients had incomplete information for calculating the KDRI; therefore, we determined at least thirteen imputed data sets were required . FifteenParameter estimates and model performance measures, along with the associated standard errors, were pooled across the imputed data sets according to Rubin’s rules . These pThe suitability of the sample size was determined according to the methods of Riley et al. . While tIn the development article , the KDRFor 1- and 5-year graft failure, the mean standard error for the calibration slope varied between 0.053 and 0.092, and 0.036 and 0.051, respectively, for the survival probabilities under consideration  . Values Calibration is used to measure the agreement between observed and predicted risks. Here, we used the observed event proportion as a proxy for observed risk. As some patients were censored prior to the event time horizon, it was not possible to calculate the observed event proportion. To overcome this, we used a jack-knife approach to calculate pseudo-observations, which were then used as proxy measures of event indicators for censored patients .Calibration plots using pseudo-observations with local weighted regression smoothing were assessed in each imputed dataset. For models that are well calibrated, the smoothed curve lies on the diagonal line that runs through the origin. Further, we calculated the calibration slope, where a value equal to one indicates perfect calibration. A calibration slope less than 1 suggests that predictions are too high for recipients with high event probabilities and too low for those with low probabilities. Conversely, a calibration slope greater than 1 suggests that recipients with high observed risk are under-estimated, and those with low observed risk are over-estimated. The calibration slope was calculated using a generalised linear model with pseudo-observations as the outcome and the complementary log–log transformed predicted risks as both an offset and covariate , 28. TheGiven that the baseline survival value for 1 and 5 years following transplantation was not reported in the original article, it was calculated within the UKTR data. Consequently, the calibration may appear more optimistic since the baseline cumulative incidence, which is required to calculate the absolute risk of graft failure, was estimated in the same cohort.The Brier score measures prediction error by estimating the squared difference between the event indicator and estimated risk . Values The KDRI  can be cthe KDRI  were usethe KDRI  or the Fthe KDRI . Howeverthe KDRI . TherefoTo assess whether the KDRI required updating, we re-estimated the coefficients used in the original index. For the KDRI to be applicable in the UK cohort, we substituted African American ethnicity for Black ethnic origin. Variables were centred in the same way that they were in the original KDRI, and no further variable selection was undertaken. We re-estimated the coefficients by censoring graft failure at the time of death using a Cox proportional hazards model and accounting for death as a competing event using a cause-specific Cox model.The coefficients were re-estimated in each of the 15 imputed datasets, and the performance of those updated models was individually assessed. The re-estimated coefficients and performance measures were then pooled according to Rubin’s rules.When updating the KDRI, we assessed the predictive performance in the same group of recipients that were used to update the KDRI. This will naturally produce optimistic results. To account for optimism in the numerical summaries of predictive performance, we used Harrell’s bias correction method , with 10Multiple imputation was performed using Stata/MP 16.1 . All othIn total 20,134 deceased donor single kidney-only recipients who received a transplant between January 1, 2004, and December 31, 2018, in the UK were eligible for inclusion  recipients were alive with a functioning graft. 2675 (13.35%) recipients experienced graft failure only, and 904 (4.51%) died following graft failure. 2732 (13.64%) recipients died with a functioning graft  recipients had experienced graft failure only, and 186 (0.93%) died following graft failure. A total of 497 (2.48%) recipients had died with a functioning graft. By 5 years, 1936 (9.66%) recipients experienced graft failure alone, and 456 (2.28%) died following graft failure. 1509 (7.53%) transplant recipients died with a functioning graft.A summary of the donor characteristics used to calculate the KDRI, including the number of missing values, is presented in Table No values were missing for donor age and type of donor. 1688 (8.43%) donors had missing values for creatinine, the most of any variables required for calculating the KDRI. 2560 (12.78%) were missing at least one value required to calculate the KDRI.The distribution of the KDRI in the original article by Rao et al.  was simiThe KDRI discriminated moderately well for predicting 1-year graft failure. The time-dependent AUC was 0.607 (95% CI 0.589 to 0.625) and 0.610 (95% CI 0.592 to 0.628) with and without accounting for competing events, respectively Table .Table 3Calibration plots and slopes were similar when modelling graft failure while censoring for death and death as a competing event Fig. . The KDRPredictive accuracy was the same regardless of whether death was handled as a competing event or not with reported Brier scores equal to 0.058 for both types of outcomes.Five years following kidney transplantation the time-dependent AUC was slightly lower when predicting graft failure with death as a competing event  as opposed to censoring at the time of death .Using calibration plots, predicted risks using the KDRI were generally similar to the observed proportion of recipients who experienced graft failure Fig. a, b. CalPredictions were less accurate for 5 years compared with a 1-year graft failure. Brier scores differed for death-censored and competing event graft failure .To update the KDRI, we re-estimated the coefficients used in the original index in the UK kidney transplant population. No additional predictors were considered. In the updated models, the estimates were similar regardless of whether the death was modelled as a competing event or not Table . ConfideThe effect of age for those under 18 and over 50 years and height was not found to be associated with graft failure. Additionally, donor ethnicity and donor HCV status were not significantly associated with graft survival.Discrimination was similar for predicting death-censored graft failure (time-dependent AUC 0.614) and graft failure with death as a competing event (time-dependent AUC 0.608) Table .Table 5Calibration slopes were 1.096 when censoring at the time of death and 1.068 when modelling death as a competing event. Calibration plots were similar for both types of graft failure and clearly showed that low risks were over-estimated and high risks were under-estimated Fig. c, d. TheDiscrimination was lower when modelling graft failure with death as a competing event  for the updated index Table .We found calibration slopes were 1.016 for death-censored graft failure and 1.002 when modelling death as a competing event. The calibration plots Fig. c, d showPrediction accuracy was slightly improved when modelling death as a competing event compared to censoring at the time of death with Brier scores equal to 0.114 and 0.117, respectively.In external validation, the KDRI had moderate discrimination and was generally well calibrated for predicting graft failure 1 year and 5 years following kidney transplantation. For predicting 1-year graft failure discrimination, calibration and predictive accuracy did not differ depending on how death prior to graft failure was handled. Discrimination was higher for predicting 5-year graft failure. Predictions were more accurate for early graft failure compared to those at 5 years following transplantation.Calibration slopes indicated miscalibration in the KDRI; however, the corresponding 95% confidence intervals were wide. In calibration plots for both outcomes, miscalibration was mainly driven by recipients at higher risk, where the event probabilities were generally underestimated. However, it should be noted that the baseline survival was not reported in the original article and as such has been estimated within the same cohort as it is being validated in. Therefore, the calibration of the KDRI in the UK kidney transplant population may be more optimistic.Updating the KDRI in the UK kidney transplant population yielded similar coefficients, but some prognostic factors were no longer associated with graft failure. Given that the coefficient estimates did not differ between the Cox and the cause-specific Cox models, it is unsurprising that there was little difference between the predictive performance of those models.To our knowledge, this is the first study to assess the performance of the KDRI under a semi-competing risks framework for first, deceased donor, single, kidney-only and adult transplants. Our work included all eligible kidney transplants that occurred in the UK during the study period, with a long follow-up period. The KDRI was previously validated in the UK by Watson et al.  using inThe baseline survivor function was recalibrated for our cohort; therefore, calibration may seem more optimistic in this external validation of the KDRI. From the current analyses, we cannot comment on the clinical utility of the index in the UK kidney transplant population. Further work is required to determine whether the KDRI is clinically relevant in practice.Few recipients experienced a competing event (died with a functioning graft), which may explain why little difference was found in predictive performance when considering death as a competing event and censoring graft failure at the time of death. There is a lack of guidance concerning under what situation ignoring the competing risk elements can impact the performance of the prediction models. Externally validating and updating a model under the competing risk framework serves as a sensitivity analysis to evaluate the developed models which ignore the competing risk elements. Future work could explore to what extent the proportion of non-terminal events censored by the terminal events impacts the predictive performance. This can potentially lead to recommendations for practice for when it is necessary to account for competing events and when traditional methods, such as the Cox proportional hazards model, might suffice.The KDRI only considers donor-related variables to predict graft failure in the recipient of the kidney transplant. Additional donor variables may improve predictive performance. The Maryland Aggregate Pathology Index (MAPI) , for exaA validation study in the US  evaluateZhong et al.  also assThe Kidney Donor Risk Index, originally developed in the US population, showed moderate predictive performance overall in our external validation in the UK kidney transplant population. The use of a semi-competing risks framework made a slight difference when predicting 5-year graft failure compared to censoring for death. The updated index had slightly improved discrimination but was poorly calibrated for those with the highest and lowest risk of graft failure. Therefore, we conclude that updating the KDRI in the present form is not required.Additional file 1: Fig. S1. Imputed datasets: Summary of continuous variables. Tables S1. Imputed datasets: Summary of categorical variables. Table S2. Sample size calculations. Table S3. Summary of recipient and transplant related factors.


Postoperative patient satisfaction after anterior cruciate ligament reconstruction (ACL-R) is influenced mainly by the degree of pain, the need for reoperation, and functional performance in daily activities and sports. Graft choice has shown to have an influence on postoperative outcomes after ACL-R. While patient reported outcomes measurements do not differ between graft options, evidence shows that normal knee kinematics is not fully restored after ACL-R with an increase in postoperative anterior tibial translation (ATT). Postoperative graft rupture rates seem to favor bone-patella-tendon-bone (BPTB) and quadriceps tendon (QT) autografts over HT or allografts. While return to sports rates seem comparable between different graft types, postoperative extensor strength is reduced in patients with BPTB and QT whereas flexion strength is weakened in patients with HT. Postoperative donor site morbidity is highest in BPTB but comparable between HT and QT. With all graft options having advantages and drawbacks, graft choice must be individualized and chosen in accordance with the patient. Pain, graft survival, and functional performance during daily activity and sport all significantly affect patient satisfaction following anterior cruciate ligament (ACL) reconstruction (ACL-R). Details about anatomy, biomechanics, graft fixation and incorporation commonly used autograft and allografts are reviewed in part I of this current concept paper. The following review will further highlight in-vivo analyses, patient reported outcomes (PROs), re-rupture rates, flexion and extension strength recovery, return to sport, and complications of the quadriceps tendon (QT), bone-patella-tendon-bone (BPTB) and hamstring tendon (HT) autograft as well as allografts. Unless otherwise specified, for the purposes of uniform comparison only studies using anteromedial portal drilling technique were included, as clinical and functional outcomes may differ with more traditional techniques .Measuring in-vivo knee kinematics during daily and athletic activities is essential to detect abnormal joint mechanics and microinstability which may not present during routine clinical testing, yet may lead to accelerated joint degeneration .ACL-R has been shown to have a significant impact on knee kinematics, with reconstructed knees more externally rotated and less flexed than the contralateral limb in the early stance phase of the running cycle one year postoperatively , 122. AdOverall, the effect of different graft types on in-vivo kinematics remains inconclusive. For HT ACL-R an increased ATT during activity was reported and linked to a reduction in hamstring force . SimilarPostoperative patient satisfaction is undoubtedly the most important outcome when it comes to ACL-R. While there is an abundance of short-, mid- and long-term literature comparing BPTB and HT, little is known about postoperative outcomes of QT. Although BPTB autograft has long been the gold standard in ACL–R, QT is gaining in popularity, especially among patients injured in pivoting sports and in those with concomitant medial collateral ligament injuries , 108.To date, only two randomized controlled trials (RCTs) have compared clinical outcomes of BPTB and QT. Randomizing 51 patients using a transtibial ACL-R technique revealed no statistically significant difference in any of the reported PROs at two years postoperative . SimilarWhen comparing BPTB to HT, three recent RCTs demonstrated no significant differences between subjective IKDC and Lysholm scores , 88, 112The reported results of QT and HT are similar to those of BPTB and HT. In a recent prospective RCT, Lind et al.  comparedWhile allografts were historically associated with inferior clinical and patient reported outcomes, recent studies using non-irradiated and non-chemically treated allografts produce comparable patient satisfaction rates and PROs to autografts , 135.Graft failure is multifactorial. Risk factors include male gender , youngerWhen comparing graft failure rates, care must be taken with terminology, as the terms "graft rupture," “failure rates,” and "revision surgery" are often used interchangeably and interpreted inconsistently. Particularly in registry studies, “revision surgery” may be reported rather than graft ruptures, as determined by postoperative MRI or clinical examination. This may lead to underestimation of true re-rupture rates. In terms of re-rupture, BPTB has long been considered the gold standard, demonstrating decreased rates compared to HT and allograft , 137. HoThere is extensive evidence on ACL revision surgery rates between BPTB and HT. Out of eleven registry studies, nine reported a significant relationship between revision rate and graft choice, with patients undergoing HT ACL-R having an up to two times higher risk of revision –79, 124.When comparing failure rates of QT to HT, high-level evidence is still lacking. Two RCTs including 99 and 51 patients respectively, found no significant difference between both graft options in the short term , 71. TheThere is extensive but contradicting evidence comparing graft rupture rates between allograft and autograft. Allografts are thought to have higher rupture and reoperation rates, with an up to sixfold increased risk of failure when compared to autograft, especially in young and active patients , 96, 126In more recent studies comparing non-irradiated or fresh frozen allograft to autograft, these higher failure rates have not been consistently reported , 68, 135Regaining normal extensor and flexor muscle strength after ACL-R, measured by a limb symmetry index (LSI) of > 90%, is a key focus of rehabilitation. The goal is to ensure safe return to sport and work, as inadequate strength has been associated with poorer function, altered biomechanics, and an increased risk of further knee injury , 138. IsComparing QT- to BPTB and HT, significantly increased isometric quadriceps weakness at 5–8 months postoperatively with QT, but no significant difference between groups at 9 to 15 months has been demonstrated . ConversWhen using HT, isokinetic flexor strength is significantly reduced compared to QT, and the deficit may persist for up to two years , 29, 70.Return to sport (RTS) following ACL-R is a commonly utilized and clinically important outcome measure. Despite its prevalence, this outcome is often reported in a variety of ways, making it difficult to compare patient subgroups. A meta-analysis found an overall 82% RTS rate following ACL-R, however the rate dropped to 63% when looking at RTS at the same level . Many faThere are few studies in the literature specifically comparing graft choice and its impact on successful RTS, but the consensus appears to find no difference between various graft types. Currently, the literature shows no difference between BPTB and HT in RTS rates. A study focusing on 100 soccer players who underwent ACL-R with either BPTB or HT revealed an overall return to play rate of 72% at 1 year follow up with 85% of those patients returning at the same level or higher . This stIn regard to QT, a retrospective study looking at 5-year follow up for 291 young active patients demonstrated a 73% RTS at preinjury level with a mean time of 8 months to return . AlthougWhile allograft is an uncommon graft choice in young athletes, the literature frequently reports no difference in RTS rates between autograft and allograft. A recent study compared 78 collegiate level soccer players who underwent ACLR with BPTB (66%), HT (17%), allograft (10%), and QT (1%). The overall mean RTS time was 6 months. There was no difference in RTS rates based on graft selection when comparing all autograft and allograft patients  . ConversWhile the current literature highlights that there may be no difference in RTS following ACL-R with various graft types, there is a need for further research on how to improve rates of return to the same level of sport amongst all graft types.Surgical techniques continuously evolve not only to improve functional postoperative outcomes, but also to decrease complications and donor site morbidity. Knowledge of the various advantages and disadvantages of each graft option is fundamental to individualized ACL-R. Of course, one of the primary benefits of allograft use is the avoidance of donor site morbidity.When considering complications and donor site morbidity related to graft choice, it is important to distinguish between minor and major complications. Minor donor site morbidities include persistent anterior knee pain, sensory loss of the lower leg, donor-site tendinopathy, scarring, cosmetic issues, and discomfort during kneeling (in patients without daily kneeling activities). Major complications besides graft rupture and contralateral ACL rupture include kneeling pain in patients who kneel during daily living, patellar fracture, extensor tendon rupture, and infection.Anterior knee and kneeling pain is the most common postoperative complication related to graft choice, reported in up to 21.5% of patients , 125. WhWhile minor donor site morbidities are irritating, severe complications like patellar fracture or extensor tendon rupture have a major impact on a patient's life and recovery. Patella fracture after ACL-R with autograft using bone blocks ranges between 0.1% and 2% , 61, 123Superficial and deep surgical site infection (SSI) after ACL-R is a rare but major complication, with an incidence between 0.32% and 1.1% , 75, 80.Compared to autografts, allografts have the advantage of reduced surgical time, lower donor site morbidity, and more predictable graft size but are believed to have a higher infection rate compared to autografts , 50. AltWith all graft options having advantages and drawbacks Table , graft cGraft choice affects postoperative outcomes after ACL-R and normal knee kinematics is not fully restored after surgery. Patients with hamstring tendon autograft may experience an increase in ATT and a decrease in flexion strength compared to those treated with BPTB or QT. Contrary, extensor strength is affected in patients with BPTB and QT. While patient reported outcomes are not influenced by graft choice, evidence suggests favorable postoperative graft rupture rates in patients treated with BPTB and QT autografts over HT or allografts. With regards to return to sports the consensus appears to find no difference between various graft types. Postoperative donor site morbidity is highest in BPTB, comparable between HT and QT and absent in allografts. With all graft options having advantages and drawbacks, graft choice must be individualized and chosen in accordance with the patient.


High amylose wheat (HAW) has potential health benefits but its dough structure is usually inferior. Wheat dough is a complex mixture and its structure is influenced by the physicochemical properties of gluten and starch. In this study, we investigated the starch granule development, gluten structure, starch properties, pasting, and thermal properties of flour, as well as the rheological properties of dough in wheat variety Xinong 836 with high amylose content (33.57%) and its parents. The results showed that Xinong 836 wheat starch contained more small starch granules, which was consistent with the microstructural results of starch granules in grain filling stage. Moreover, Xinong 836 wheat starch showed highest swelling power and water solubility. Importantly, the flour of Xinong 836 wheat had the highest protein content and wet gluten content and Xinong 836 wheat gluten showed highest β-sheets content and disulfide bond content than its parents Zhengmai 7698 and Xinong 979, which conferring to more compact microscopic networks of dough, thereby contributing to the higher peak viscosity (PV), final viscosity (FV), and setback viscosity (SB) in the flour of Xinong 836. Our finding elucidated that the stability of gluten and properties of starch synergistically affected the pasting and thermal properties of the flour paste, and the presence of more small starch granules contributed to dough with a rather dense structure in HAW Xinong 836. Thus, superior gluten structure and more small starch granules have synergistic effects on enhancing the gluten–starch interaction, thereby contributing to better dough quality. Storage proteins and starch are the main components of wheat flour, and they are accumulated in the endosperm during grain development and stored in protein bodies (PBs) and starch granules (SGs), respectively. Upon addition of water and next stirring, wheat flour can form the dough with a unique three-dimensional structure and viscoelasticity woven by gluten and starch. Gluten proteins constitute 80–85% of wheat flour proteins and are considered to have crucial effects on the properties of dough , Shaanxi Province, China, during two consecutive wheat growing seasons (2018–2019 and 2019–2020). The plot size was 20 × 2 m × 0.23 m and the row spacing was 0.23 m. Three biological replicates were performed for each wheat variety. The spikes obtained from three varieties were marked at the flowering stage. About 50 labeled spikes were randomly collected from each wheat variety at 7, 16, and 28 days after anthesis (DAA). The labeled spikes were used for cytological observations. After harvesting, the wheat grains were milled using a multifunctional pulverizer , and passed through a 100-mesh screen.2.2.Wheat kernels were collected at 7, 16, and 28 DAA to observe the microstructure of the caryopses. Slices of the wheat endosperm were prepared and immersed immediately in 4% glutaraldehyde fixative at 4°C for 8 h. The samples were subsequently washed three times with 0.1 M sodium phosphate buffer (pH 6.8). Finally, the slices were finally embedded in white resin, then carefully cut into 1-μm slices by using a histotome  and stained with toluidine blue (0.03 M) for 10 s. The samples were observed under a Leica microscope .2.3.A near-infrared reflectance spectrometer  was used to obtain the quality-related parameters for the mature wheat grains as described by Gao et al. . As the 2.4.Dough samples were prepared by mixing flour with water [flour (g): water (mL) = 5:3], followed by recovery for 15 min. Gluten and starch were separated by hand washing. The gluten was lyophilized in a freeze dryer , ground into a fine powder, passed through a 100-mesh sieve, and finally stored at 4°C. The starch slurry was successively washed with 2 M sodium chloride, 2% sodium dodecyl sulfate (SDS), and distilled water and centrifuged. And this process was repeated three times. Finally, the pure starch was oven-dried, passed through a 100-mesh sieve and stored at 4°C.2.5.−1) in the Fourier transform infrared (FTIR) spectra as reported by Song et al. , where 1.36 × 104 is the molar absorptivity. The experiments were repeated independently in triplicate.Where A412 is the absorbance at 412 nm, D is the dilution factor, i.e., 5.02 for free SH and 10 for total SH, C is the sample concentration (mg/mL), and 73.53 is derived from 102.6.2.6.1.The amylose contents were determined using the dual-wavelength iodine binding method reported by Liu et al.  with a s2.6.2.The morphology of SGs was observed by scanning electron microscopy . Starch samples were placed on a metal stub with double-sided adhesive tape and sprayed with gold particles for 90s in a vacuum evaporator. Scanning electron micrographs were captured using SEM at a magnification of 1,000×. The number and volume distribution were determined for SGs using a Microtrac S3500 laser diffraction instrument  and laser diffraction particle size analyzer , respectively.2.6.3.g for 10 min, and the sediment was weighed. The sediments in the tubes were then dried to a constant weight and weighed again. The swelling power and solubility were calculated as described by Zhu et al. . The starch solution was heated in water at 95°C for 30 min and shaken gently. The tubes were cooled to room temperature, centrifuged at 8000 × u et al. . The exp2.6.4.−1. The relative crystallinity was calculated by using MDI Jade 6.0 software .XRD patterns were acquired for starch samples using an X-ray diffractometer . The starch samples were scanned at 30 mA and 40 kV, with diffraction angles (2θ) ranging from 4° to 40° at a scan rate of 2° min2.7.The pasting properties of flour were determined using a rapid viscosity analyzer . A sample weighing 3 mg and 25 g of ultrapure water were added to the canister. The pasting parameters including the peak viscosity (PV), breakdown viscosity (BD), final viscosity (FV), setback viscosity (SB), and pasting temperature (PT) were obtained using the pasting program described by Shang et al. .o, Tp, and Tc), and enthalpy of thermal transition (ΔH) were obtained using TA7000 software . All of these measurements were repeated three times.The thermal properties of the samples were determined using a simultaneous thermogravimetric analyzer  according to the method described by Song et al. . The ons2.8.The dough development time and stability time were analyzed using a Mixolab2 system , and calculated using the Mixolab “Chopin +” protocol. Each flour sample was tested independently in triplicate.2.9.The dough samples were post-stained using a dye mixture comprising 0.010% (w/w) rhodamine B solution in water. The microstructure of each dough sample was observed by CLSM as described by Gao et al. . For eac2.10.p < 0.05 with Duncan’s multiple range test. Pearson’s correlation coefficients were calculated between the parameters.Differences among data were detected using one-way analysis of variance , and significant differences among parameters were compared at 3.3.1.The basic components of the three wheat varieties are shown in In general, it is considered that higher protein content improves the end-product quality. According to previous studies, gluten proteins comprise 80–85% of wheat flour proteins, and greater amounts of gluten may enhance the formation of the gluten network . Thus, t3.2.The accumulated PBs and SGs in endosperm cells in the three wheat varieties were observed at 7, 16, and 28 DAA using optical microscopy. During the early filling stage (7 DAA), PBs did not appear to be accumulated in the three varieties, but SGs formed in the endosperm and they were distributed along the cell wall –C. PBs w3.3.Glu-B1 and Glu-D1 loci. Relating the subunit composition to certain quality traits is a frequently used methodology in wheat. The Payne score provided a single-number parameter to estimate the dough strength of a sample as a function of its HMW-GS allelic composition (−1). It is widely accepted that β-sheets and intermolecular β-sheets contribute to a more stable gluten network conformation , followed by Zhengmai 7698 and Xinong 979. The results may be explained by the higher B-type starch content in Xinong 836. A positive correlation was also observed previously between the small SGs content and starch swelling power . According to Wang et al.  obtained between the gluten structure, starch properties, and dough-related parameters are summarized in The A-type and B-type SGs contents were correlated with the dough lacunarity, thereby indicating that differences in the SGs contents could have affected the filling state of the dough. The presence of more small SGs contributes to a rather dense dough structure and affects the dough quality. The CLSM images also confirmed the compact structure . In addir = −0.984, p < 0.01), FV , SB , and To , but positively correlated with ΔH , thereby confirming that the relative crystallinity affected the pasting and thermal properties of flour. The swelling power and relative crystallinity had opposite effects on the flour pasting properties, mainly because the swelling power is related to interactions between amorphous and crystalline structures in starch, and negatively related to the relative crystallinity , the Special funds for Nanyang Wheat Experimental and Demonstration Station (No. 2021NY), and the Program of Introducing Talents of Innovative Discipline to Universities (Project 111) from the State Administration of Foreign Experts Affairs (No. #B18042) “Crop breeding for disease resistance and genetic improvement.”The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. 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Our results reveal that the diatomite-based ceramic thermal insulator obtained by starch consolidation is effective for use on roofs to improve thermal comfort in dwellings located in cold regions.Researchers are continuously seeking to develop new materials to protect against inclement weather and thus optimize energy efficiency in housing. This research aimed to determine the influence of corn starch percentage on the physicomechanical and microstructural properties of a diatomite-based porous ceramic. The starch consolidation casting technique was applied to fabricate a diatomite-based thermal insulating ceramic with hierarchical porosity. Diatomite mixtures with 0%, 10%, 20%, 30%, and 40% starch were consolidated. The results show that starch content significantly influences apparent porosity, and this, in turn, influences several parameters, such as thermal conductivity, diametral compressive strength, microstructure, and water absorption of diatomite-based ceramics. The porous ceramic processed by the starch consolidation casting method corresponding to the mixture of diatomite with 30% starch obtained the best properties, with a thermal conductivity of 0.0984 W/m·K, an apparent porosity of 57.88%, a water absorption of 58.45%, and a diametral compressive strength of 35.18 kg/cm As mentioned in [Humans have always sought out materials to protect themselves from inclement weather. Environmental pollution and energy-related challenges have both experienced progressive worsening in recent years . Accordiioned in , the tenioned in  states, Porous ceramics play an important role in our daily lives; they are used in applications such as thermal insulation and barriers, acoustic absorption, vibration attenuation, energy storage, and fluid filtration . As arguThere are currently two main areas of concentration for insulating material research and development. The goal of encouraging the use of environmentally friendly building materials and products is becoming increasingly important, which, in turn, is encouraging the development of insulation materials with a lower environmental impact throughout their entire life cycle, favoring those that are quickly renewable, derived from plants or animals, potentially recyclable, non-toxic, and have low energy production methods .Diatomite is a naturally occurring sedimentary rock made of the siliceous remains of fossilized diatoms . In addiOne of the commonly used pore-forming substances in ceramic technology is starch. The ceramic forming technology that uses starch is known as starch consolidation casting, in which starch additionally acts as a binder . A high-Thermal insulators are highly resistant to the passage of heat, reducing heat transfer; i.e., they insulate against cold or heat, resulting in savings in energy use by increasing their thermal resistance . There aFor this reason, this research seeks to develop a diatomite-based material that has a hierarchical pore structure and can be used as an efficient thermal insulator to improve thermal comfort in houses located in cold regions. This research work is justified because it develops a new material for obtaining an efficient thermal insulator to help address frost and cold. In this way, it optimizes the conservation of internal heat in the house by reducing thermal conductivity, which will improve people’s thermal comfort and quality of life. Based on the above, we formulate the following question: What is the influence of the percentage of corn starch on the microstructure, apparent porosity, water absorption, diametral compressive strength, and thermal conductivity of a diatomite-based ceramic?3, BET surface area 12.99 m2/g, and particle size less than 45 μm (325 mesh), as guaranteed by the supplier. The chemical composition of diatomite was obtained by X-ray fluorescence in the Bizalab laboratory in Lima, Peru. The identification of the crystalline phases present in the diatomite prior to processing was carried out using a Rigaku Mini-flex 600 X-ray diffractometer and QualX 2.0 software. The operating conditions to obtain the diffractometry were a Kα line of copper, wavelength 1.54 A°, with a step of 0.02°, a sweep between 10 and 80° at a speed of 2° per minute, a voltage of 40 kV, and a current intensity of 15 milliamperes.The main material used in this research was diatomite obtained from the company Silicis Perú, whose deposits are in Piura, Peru. This diatomite has the following characteristics: density 2.06 g/cmThe material used as a pore-forming agent was commercial corn starch. Its morphology and chemical composition were determined with a Tescan Vega 3 scanning electron microscope with an Oxford EDS attached. Corn starch is a granular polysaccharide composed of two glucose polymers, amylose (amorphous and soluble) and amylopectin ; the amylose/amylopectin ratio determines the different degrees of crystallinity of the grain .2). All tests were performed in triplicate, except for diametral compressive strength, which was performed in quintuplicate.The experimental design was unifactorial, having as an independent variable the percentage of starch in the diatomite–starch mixture, which was investigated at 5 levels: 0%, 10%, 20%, 30%, and 40% starch. The dependent variables were apparent porosity (%), water absorption (%), and diametral compressive strength  plate of known thermal conductivity. InstaCal’s software version 6.73 was used for data acquisition, and TracerDAQ software version 2.3.4.0 was used for plotting the graph, which was used to verify that the temperature measurements were taken in a steady state.The Tescan Vega 3 XMU scanning electron microscope was used to obtain the microstructure that allowed us to differentiate the porosities obtained at the different scales.Finally, the variation in the formation of crystalline phases in the unsintered diatomite with the diatomite sintered at 1000 °C was determined. Likewise, they were compared with the best mixture of diatomite and starch sintered at the same temperature.2).The chemical composition of diatomite is shown in 2/Na2O ratio of 3.35 was used as a deflocculant, in an amount determined by its flocculation curve.Finally, to stabilize the slips, which are aqueous suspensions of diatomite–starch, liquid sodium silicate with a SiOOn the other hand, in the diatomite–starch materials , it is observed that mesopores increasingly form with greater amounts of added starch. These formed pores are of the order of 10 to 15 microns, as observed in the microphotographs at 500 magnification. In the 7500-magnification micrograph e2, a large pore occupies almost the entire microphotograph.Finally, From the deflocculation curves carried out in the rheological study , it was Regarding the behavior observed in Analyzing the water absorption curve b, we canThe behavior observed in the thermal conductivity curve as a function of the percentage of starch added c is expl2, which is equivalent to 3.45 MPa. These findings are consistent with those of other authors, as reported in their studies [The behavior of diametrical compressive strength d is expl studies ,13,29,312 in closed pores, which has a lower thermal conductivity than atmospheric air [From the SEM micrograph in eric air . Therefo(1)“Influence of starch content”: Starch content significantly influences the apparent porosity, and this, in turn, influences several parameters, such as thermal conductivity, diametral compressive strength, microstructure, and water absorption of the diatomite-based ceramic. As starch content increases, apparent porosity and water absorption increase, while thermal conductivity and diametral compressive strength decrease. This is important because the lower the thermal conductivity of a material, the better it is as a thermal insulator, and the greater the thermal comfort inside the house.(2)2 (3.45 MPa). These values of thermal conductivity and thermal resistance are adequate for a thermal insulating material.“Optimal diatomite–starch mixture”: The optimal mixture was diatomite with 30% starch. With this mixture, ceramic tiles can be manufactured to provide thermal insulation for residential roofs in cold regions. This mixture had a thermal conductivity of 0.0984 W/m·K, an apparent porosity of 57.88%, a water absorption of 58.45%, and a diametral compressive strength of 35.18 kg/cm(3)“Thermal insulator with hierarchical porosity”: A cellular ceramic with hierarchical porosity at the nanoscale, microscale, and mesoscale was obtained, making it suitable for application as an effective thermal insulator. It is important for thermal insulators to have pores of different sizes, because this will make it more difficult for heat to flow through them.This research work presents the successful synthesis of a diatomite-based porous ceramic, obtained by the starch consolidation method, to be used as thermal insulation in roofs to improve thermal comfort in cold areas. The main findings are summarized as follows:In summary, our findings reveal that the developed thermal insulator is suitable to be used to improve thermal comfort in houses located in cold regions. To improve its mechanical strength, it is recommended that in future work, kaolin be incorporated as the third component of the solid mixture to act as the main binder, leaving starch only as a pore-forming agent. In addition, a more exhaustive study of the closed porosity is recommended, because the carbon dioxide generated by the starch combustion could also influence the thermal conductivity by being trapped in the closed pores and having a thermal conductivity lower than that of air.


Disgust and neophobia elicited by whole insect products, has necessitated the need to mask insect-based food products. The physico-chemical parameters, sensory acceptance, and willingness to pay (WTP) for wheat biscuits supplemented with cricket powder was evaluated. The biscuits’ color intensity correlated with the cricket inclusion levels. Spread ration of cricket-enriched-biscuits increased (1.0–1.2-folds), while the hardness and fracturability decreased  compared to the control biscuit. Cricket-biscuits exhibited 1.2–1.7, 1.1–3.7, 1.2–3.0 and 1.1–1.2-folds higher (p < 0.05) protein, ash, fiber, and fat, respectively. Ca, Fe, and Zn were 1.1–3.7, 1.1–1.2 and 1.4–4.0-folds higher, respectively, for cricket-based biscuits. Monounsaturated and polyunsaturated fatty acids proportionally increased with increasing cricket flour. On a likert scale, 71.4%, 71.9%, 38.4% and 57.5% of the caregivers and 38.6%, 58.3%, 40.0% and 34.0% for children (3–5 years) strongly preferred the color, texture, taste and aroma, respectively, of the cricket-based biscuits. Forty-seven (47%) of the caretakers were WTP a premium of 37 Kenyan shillings (0.34 USD) for cricket-based biscuits. Our findings demonstrated that integration of cricket flour into existing market-driven consumer familiar food products significantly increased acceptability and WTP, thus promising potential to contribute to improved food and nutritional security. •Childhood malnutrition is an important public health problem.•Protein and fat in cricket-enriched biscuits exceeded the daily recommended amount.•Cricket-enriched biscuits can tackle protein-energy malnutrition, Fe, and Zn deficiencies.•Caregivers expressed increased acceptability and WTP for cricket-enriched biscuits.•More studies are needed to examine their effect on nutritional status in children. Around Entomophagy (the consumption of insects) has been practiced in many civilizations around the world and is re-emerging as a source of high-quality, cost-effective, and sustainable alternative food sources to meat . Edible Bifidobacterium animalis in the digestive tract  is one of the most widespread species worldwide and has been shown to be a viable source of food and feed , fed and bred on green leafy vegetables and chicken feed, were obtained from the Jomo Kenyatta University of Agriculture and Technology Insect Farm, Nairobi, Kenya and temporarily frozen-stored at −30 °C. The crickets were thawed, oven-dried at 180 °C for 20 min, ground in an electric grinder  and sieved through 200-micron-sized mesh to attain consistent particle size. The cricket flour obtained was used to substitute wheat flour at varying proportions: 5% (CBB5), 10% (CBB10), and 20% (CBB20) with 0% (w/w) serving as control (no insect flour included). The basic nutritional characteristics of G. bimaculatus meal and wheat flour used in the biscuit's formulation are presented in All the ingredients used  were purchased from the local market. Six kilograms of crickets  ranging from 0 (black) to 100 (white) and green-red (a*) and blue-yellow (b*) chromaticity. The overall color difference (ΔE) was computed using the formula below:ΔL*, Δa*, and Δb* are the differences in L*, a*, and b* values between the reference and test samples, respectively.Within 24 h of preparation, the weight, diameter, thickness, spread ratio, textural quality, and color of the different biscuits products were determined following standard methods described by Association of Official Agricultural Chemists . BiscuitA snap test using a texture analyzer  for a 50 kg load cell was adopted for determination of the biscuit hardness, with the application of the following settings; 5 mm/s pre-test, 3 mm/s test, and 10 mm/s post-test.2.32.3.1The moisture, total ash, crude fiber, fat, crude protein, and carbohydrate contents of formulated biscuits were determined using standard laboratory methods . Fat con2.3.2Mineral analysis  of the formulated biscuits was carried out using a standard method . Eight-m2.3.3Fatty acids determination followed the procedures described by 2.3.4This study was conducted in Keiyo South sub-county, Elgeyo Marakwet County, Kenya. A preliminary sensory test, recruiting children and caregivers from the same demographic area but from a different locale (n = 86), suggested higher consumer preferability of biscuits formulated with 20% cricket flour (CBB20) and the control, justifying their use in the subsequent test. The study targeted 214 children aged between 5 and 10 years and 146 caregivers, tallying to 360 participants. The demographic characteristics of the adult participants were as follows: Gender distribution was 83% female and 17% male. The average monthly income was Kenya shillings (KES) 10,000. Adults aged 18 to 34 constituted the majority of the population, accounting for 84% of all adults, and the average household size was between 5 and 10 people. The majority had completed primary school (48.3%), secondary school (35.2%), and college (16%).Before the assessment, the contents of the biscuit samples were disclosed to all the participants verbally and in writing for consent. Further, participants with history of self-reported allergy to any ingredient in the biscuits, as well as any medical contraindication, were deemed ineligible. The participants were each randomly served with two coded biscuits samples and clean water for mouth cleansing in between the tests with children participating first followed by the caregivers. The panelists were instructed to rate the color, flavor, taste, texture, and overall acceptability of the biscuits against a 5-point hedonic scale . As a reward for participating in the experiment, participants were asked to keep the remaining biscuits. This was done to reduce the risk of in-kind endowment effects, which could have distorted the actual liking . Ethical2.3.5A double-bounded dichotomous choice questionnaire, applying the contingent valuation method, was used to determine the consumer willingness to pay (WTP). The caregivers and children were recruited to provide a yes or no response based on their WTP. Depending on the responses given, follow-up questions were posed i.e., ‘yes’ respondents were offered a higher price while the ‘no’ respondents were given a lower price. Participants were also assigned different starting bids at random to reduce starting point bias. The forms were collected, data compiled and analyzed. A Stata double-bounded command was utilized to create the econometric WTP estimate using a linear function  as shownt1 and the second bid amount being marked as t2. The WTP was then reduced to one of the four following categories of each respondent:In the double-bounded model, the respondents provided two responses to the closed willingness to pay (WTP) questions, with the first bid amount being marked as i replies were coded with the dichotomous variables y1 1 (response to the first WTP question) and y2 1 (response to the second WTP question), respectively, indicating that the variables were given the value 1 if the individual's answer was yes and 0 if no. Because of the normal distribution of the WTP and error term ui, the probability of the four cases is defined as follows  = Pr  Where Double-bounded model the probability that an individual answers ‘Yes’ to the first question and ‘No’ to the second can be expressed as i  = z0iβ + ui and ui ∼ N , Therefore, the probability of each one of the four cases is given by:When WTPi = 1,. …., n, whereas The maximum likelihood method was used to estimate β and μ. The function that needs to be maximized in order to determine the model's parameter is described using equation 2.4All data were analyzed using R Studio version 2022.12.0 + 353.pro3 . One-way33.1The physical characteristics of the cricket-based cookies and the control biscuits are indicated in 3.2Present results display significantly increase in redness (a*) of the biscuits while reducing their lightness (L*) and yellowness (b*) color parameters with increasing cricket flour inclusion .Table 3C3.33.3.1The proximate composition and energy values exhibited significant variabilities with increasing cricket flour incorporation . The lev3.3.2Substantial significant differences were discerned in the concentrations of all the minerals with increasing cricket flour supplementation . Supplem3.3.3p < 0.05) altered as a result of the use of cricket flour in the recipe (Z)-octadecenoate and methyl (11Z)-octadecenoate of the MUFAs and methyl -octadecadienoate of the PUFAs were the predominant fatty acids (FAs) detected in the biscuit samples.The fatty acid composition of the biscuits was significantly  was unwilling to consume the biscuits, citing “it is not usual to consume insects in my society,” for the rejection. Approximately 23.1% of the respondents rejected the cricket biscuits because of lack of interest in the insect proteins and aversion. A group, accounting for 15.4% of the total respondents avoided the cricket biscuits due to financial constraints. This group may have liked the cricket biscuits but were unable to afford them. Another group comprising 7.6% of the respondents were skeptical of the new food. All these reasons for negative WTP could be a precedent for future actions, such as educating consumers about the benefits of edible insects.3.4.4The respondents' WTP was a significantly influenced by age, income, education, gender, and employment . There a3.4.5To get the mean WTP , the coe4The higher diameter of cricket-based biscuits could be attributed to the low gluten content caused by the dilution effect of cricket flour in the dough, which could have altered the consistency and structure of the dough , resultiWhile there was no significant difference in weight, the developed biscuits' thickness varied significantly between formulations. The progressive increase in thickness with inclusion levels may be attributed to the addition of cricket flour which reduced gluten and starch levels. The alteration of such components of wheat has a substantial impact on dough rheology and product quality. Compared to wheat dough, low gluten dough has a reduced cohesion and elasticity . These pThe increasing cricket flour levels also reduced the hardness and fracturability of the biscuits rendering them softer and less brittle. Incorporation of cricket flour into the biscuit dough introduced novel proteins which suppressed the levels of gluten and starch , resultiΔE) of the cricket-cookies with 20% cricket biscuits exhibiting the highest value. These differences portray color changes from slightly dark color to darker colors with increasing cricket inclusion. This can be attributed to the possible addition of highly reactive free amino acids and proteins introduced from cricket flour, which may have caused intense Maillard reactions, generating dark colored biscuits after baking  but reducing their lightness (L*) and yellowness (b*). r baking .The progressive increase in the proportions of protein, ash, fiber, fats, and energy of the biscuits with cricket substitution levels may be linked to their additive supplies from cricket flour, previously reported to be endowed with appreciable amounts of these proximate parameters . In partThe minerals phosphorous, calcium, potassium, magnesium, and sodium were the most predominant minerals, which have also been previously reported in different cricket species . EvidentThe dominant SFAs, MUFAs and PUFAs detected in this study are consistent with studies which identified palmitic acid, stearic acid, oleic acid and linoleic acids as the abundant fatty acids in cricket flour, considerably enhancing their respective levels in the bakery products they are used to formulate . CertainThe low rating of color among the children can be attributed to an unexpectedly dark color change, occasioned by Maillard reaction between dough sugars and free amino acids from cricket flour, which did not match the snack they are accustomed to. The higher desirability of the color and texture of CBB20 among the caregivers is a demonstration that the dark color and gritty texture had no effect on their opinions. This could be due to the notion that brown-colored and fibrous foods are more nutritious and wholesome . As estaIn general, slightly more than half of consumers were willing to pay a premium price. The significant numbers of consumers prepared to pay low premiums indicate that consumers may have lacked confidence in the usefulness of cricket-based biscuits in combating malnutrition and food insecurity, as mentioned in our study information. In addition, consumers may not have trusted the nutritional information given considering the fact that protein in the biscuits is an invisible component. Our initial price of Kenya shillings (KES) 30 (USD 0.98) may also have been too high for local consumers to afford at that time of the study.p < 0.05) WFP with the middle-aged consumers (23–35 years) being more willing to pay than older consumers. Younger parents prefer quality food for their children to consume than older respondents with grown children. Similarly, readiness to adopt insects was found to be stronger among younger consumers compared to older consumers in a previous study (p<0.005). This may be attributed to the fact that employed individuals are mostly educated and may view insect products as inferior foods meant for the poor. The computed premium price for cricket-based biscuits superseded the price of the familiar product selling at KES 30 (USD 0.27) for 100 g. Concurring results were reported by This study revealed that females would be willing to pay more for cricket-based biscuits than males. This is because women are responsible for purchasing food and cooking for the entire home and would sacrifice to give out more of their income to ensure a healthy diet for their households . Conversus study . IncomesG. bimaculatus in enhancing the nutritional levels of the cricket-based biscuits as demonstrated in this study, their contribution to health through the purveyance of therapeutic compounds cannot be trivialized. Earlier reports have implicated G. bimaculatus extracts as effective remedies to certain medical disorders and gut health improvement (G. bimaculatus as a novel functional food with immense potential for alleviating malnutrition as well as advancing human health simultaneously.Despite the effectiveness of rovement . These a5This study has demonstrated that the substitution of wheat flour with cricket flour yielded nutritionally enriched biscuits with good sensory acceptability. The use of cricket flour to improve familiar products like biscuits could increase the diversification of insect food products, promote insect consumption, and improve household nutrition and health. However, amidst global exponential rise in adoption of insects and insect-derived products, particular attention should be paid to food frauds with regards to mislabeling of insect-enriched products. Therefore, proper control methods such as the use of near infrared spectroscopy for authentication of the insect-based products before introduction to the market should be employed to curb such vice in the food industry. Further studies on cost benefit analysis of these novel products would be crucial.Divina Arama: Conceptualization, Methodology, Formal analysis, Investigation, Data curation, Writing – review & editing, Visualization. John Kinyuru: Conceptualization, Methodology, Visualization, Validation, Investigation, Writing – review & editing, Supervision. Jeremiah Ng'ang'a: Conceptualization, Validation, Writing – review & editing, Supervision. Beatrice Kiage-Mokua: Conceptualization, Methodology, Investigation, Writing – review & editing, Supervision. Brian O. Ochieng: Methodology, Formal analysis, Writing – review & editing. Chrysantus Mbi Tanga: Conceptualization, Methodology, Investigation, Validation, Resources, Writing – review & editing, Visualization, Supervision, Project administration, Funding acquisition, All authors read, reviewed, and approved the final manuscript.The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.


LATE EMBROYGENESIS ABUNDANT gene, OsLEA1b, a heat-stress-responsive gene in rice grain filling. OsLEA1b is highly expressed in the endosperm, and its coding protein localizes to the nucleus and cytoplasm. Knock-out mutants of OsLEA1b had abnormal compound starch granules in endosperm cells and chalky endosperm with significantly decreased grain weight and grain number per panicle. The oslea1b mutants exhibited a lower proportion of short starch chains with degrees of polymerization values from 6 to 13 and a higher proportion of chains with degrees from 14 to 48, as well as significantly lower contents of starch, protein, and lipid compared to the wild type. The difference was exacerbated under high temperature conditions. Moreover, OsLEA1b was induced by drought stress. The survival rate of oslea1b mutants decreased significantly under drought stress treatment, with significant increase in ROS levels. These results indicate that OsLEA1b regulates starch biosynthesis and influences rice grain quality, especially under high temperatures. This provides a valuable resource for genetic improvement in rice grain quality.High temperatures accelerate the accumulation of storage material in seeds, often leading to defects in grain filling. However, the mechanisms regulating grain filling at high temperatures remain unknown. Here, we want to explore the quality factors influenced by the environment and have identified a  Oryza sativa) is an important crop consumed as a staple food by more than half of the world’s population. The rice endosperm is the main storage organ for nutrients, including starch, proteins, and lipids [Rice  likely stabilizes amylose content in rice seeds under high temperatures [The environment also affects endosperm starch biosynthesis. Heat-induced crop damage is becoming increasingly common as a result of global warming, severely limiting high and stable rice yields . Rice cae stress . Inhibitnditions . The higeratures . MoreoveArabidopsis thaliana), LEA proteins are associated with seed dehydration tolerance [OsLEA promoter to activate its expression to high levels in embryonic tissues during endosperm development [There are several endosperm-specific genes that were identified through a genomic survey . These golerance . Four NAelopment . Howeveroslea1b mutants were more sensitive to high-temperature stress and drought stress than the wild type. Milled rice from oslea1b mutants showed significantly lower starch content and abnormal structure under a high-temperature environment, which resulted in significantly greater chalkiness and chalky grain rate than that from the wild type. A series of assays were utilized to determine the effects of OsLEA1b on starch biosynthesis under high-temperature conditions. These findings enhance our understanding of OsLEA proteins and grain filling in rice, providing useful information for genetic improvement of rice grain yield and quality.Here, we want to explore the quality factors influenced by environment and have identified an embryonic protein homologous to OsLEA3-2 , which wOsLEA1b (LOC_Os01g16920), which is predominantly expressed during the grain-filling stage and is homologous to OsLEA3-2  reporter gene driven by the OsLEA1b promoter into rice. Histochemical analysis corroborated that GUS activity in transgenic plants was strongest in the endosperm /CRISPR-associated nuclease 9 (Cas9)-mediated genome editing. We successfully obtained two independent homozygous mutants, oslea1b-1 and oslea1b-2. RT-qPCR analysis showed that OsLEA1b expression in the two mutants was significantly lower than that in the wild type  than that of the wild type  revealed that wild-type starch granules were polygonal in shape and densely arranged. In contrast, starch granules in the chalky endosperm of the mutants were round and had numerous air spaces  and normal temperatures (28 °C for 12 h in light and 22 °C for 12 h in darkness). Under high-temperature conditions, mature wild-type grains were slightly chalky, but mature oslea1b grains were floury with a significantly lower 1000-grain weight  and BRITTLE 1 (OsBT1) were significantly lower in the mutant than in the wild type under high temperature conditions. Conversely, the expression levels of SOLUBLE STARCH SYNTHASE IIA (OsSSIIa), PULLULANASE (PUL), ISOAMYLASE 3 (ISA3), STARCH BRANCHING ENZYME IIb (OsBEIIb), GRANULE-BOUND STARCH SYNTHASE I (GBSS1), RAG2, and GLUTELIN TYPE-D 1 (GLUD1) were significantly higher in the mutant than in the wild type under high temperatures  activity, hydrogen peroxide (H2O2) content, proline content, and malondialdehyde (MDA) content after PEG treatment compared to the wild type (oslea1b mutants after PEG treatment but not in the control (oslea1b mutants are more sensitive to drought stress than the wild type.Reactive oxygen species (ROS) levels were determined in leaves of the wild type and the ild type G–J. Gene control K,L. Thesoslea1b mutants exhibit high chalkiness rates under normal temperatures and chalky endosperm under high temperatures. The abnormally developed starch granules in the mutant endosperm cells were loosely arranged rather than whole complex granules. The oslea1b mutants were more sensitive to high temperature during endosperm development than wild-type plants. Moreover, the chalkiness rate and chalkiness degree of the mutants were dramatically higher than those of the wild type under both natural and artificial high temperatures. By contrast, the gelatinization and gel consistency of the two mutants were significantly lower than those in the wild type at high temperatures. Recent studies have shown that in addition to starch content, the arrangement of starch granules also affects rice grain cooking and eating qualities. It is likely that the abnormal starch granule development induced by high temperature in oslea1b mutants would affect the cooking and eating qualities. The higher proportion of short starch chain length and easier pasting properties of oslea1b may confer it greater sensitivity to high temperature tolerance.High temperature during the rice grain-filling stage often results in lower quantity and worse quality, such as higher chalkiness and lower grain-setting rate . Grain cChalk5, the major QTL control chalky grain rate of rice, two polymorphic nucleotides on the promoter of Chalk5 affect its expression level. The lower expression level could decrease the chalkiness of rice and increase the percentage of milled rice. Total starch and amylose content in the oslea1b mutants were significantly lower than those in the wild type, especially under high temperatures. Moreover, a significantly different distribution of amylose chain lengths in the mutant led to an altered starch structure. The expression of BE1, SSIIa, PUL, ISA3, and BEIIb showed significant differences between the wild type and mutants under high-temperature treatment, which resulted in differences in the fine structure of starch [SSIIa was significantly higher in the mutant than in the wild type under high temperatures, which may be responsible for the reduced viscosity. Although the effects of high temperature on rice quality are well known, additional research is needed to explore the underlying regulatory network to improve rice quality.Starch content and structure directly influence grain appearance such as chalkiness . And manf starch ,28. The f starch . GelatinZea mays), cotton (Gossypium hirsutum), potato (Solanum tuberosum), and other plant species [Glycine max) PM2 (LEA3) protects proteins and maintains enzyme activity under unfavorable temperatures [Phylogenetic analysis showed that the embryonic protein OsLEA1b is homologous to the late embryogenesis abundant protein OsLEA3-2. LEA proteins are widely distributed in rice, Arabidopsis, maize  plants heterologously expressing melon (Cucumis melo) CmLEA-S display greater tolerance to drought and salt compared to wild-type plants [oslea1b mutants were more sensitive to drought stress than wild-type plants, showing extremely low survival rates and significantly increased cellular ROS levels. CATALASE B (OsCATB), ASCORBATE PEROXIDASE 1 (OsAPX1), PEROXIDASE 1 (OsPOX1), 2+/H+ EXCHANGER 1aCa (OsCAX1a), OsP5CR, OsNAC106, HIGH-AFFINITY K+ TRANSPORTER 1;3 , CYCLIC NUCLEOTIDE-GATED CHANNEL 9 (OsCNGC9), RESPONSIVE TO ABA 21 (OsRAB21), OsRAB16a, and AMMONIUM TRANSPORTER 3 (OsAMT3) are stress-related genes [oslea1b confers the sensitivity of the mutants to drought stress. This validates that OsLEA1b responds to drought stress by influencing the expression of those genes. Our results indicate that OsLEA1b regulates stress tolerance. We will further investigate the effects of overexpression of OsLEA1b on drought tolerance.Furthermore, LEA proteins enhance plant salt and drought tolerance. LEA proteins also play an important role in the adaptive responses to water deficit in higher plants . Transgee plants . In thised genes ,49,50,51OsLEA1b is highly expressed in seeds yet barely detectable in other rice tissues. However, the drought sensitivity of oslea1b mutants was observed in other parts of the plant, such as the leaves. Additional studies are needed to explain how OsLEA1b, which is enriched in the endosperm, regulates plant stress tolerance. Overall, our research reveals that OsLEA1b regulates starch biosynthesis in the grain endosperm and plays an important role in rice drought tolerance. This study provides useful information for potential genetic improvement of rice yield and grain quality.oslea1b mutants were generated by CRISPR/Cas9-mediated genome editing in the japonica rice (Oryza sativa ssp. japonica) of ‘Zhonghua 11′ (ZH11) background. Homozygous T2 lines of oslea1b were used in experiments. Rice plants were grown in a paddy field in Hangzhou, Zhejiang province, China, during the regular growing season. Wild-type and oslea1b plants (pre-flowering) were subjected to normal temperatures  and high temperatures  in plant growth chambers. The average outdoor daytime temperature in Hangzhou was 37 °C from 20 July 2020 to 20 August 2020.The OsLEA1b using CRISPR/Cas9, single guide RNAs (sgRNA) targeting OsLEA1b (Os01g0276300) were cloned into the BGK03 vector . The CRISPR/Cas9–OsLEA1b construct was transformed into ZH11 calli via Agrobacterium tumefaciens (Agrobacterium)-mediated transformation. Primer sequences used for vector construction and validation are listed in To knock out −ΔΔCT method [OsLEA1b (~2 kb upstream of ATG) was amplified by PCR and cloned into the EcoRI and NcoI sites of pCAMBIA1305. This construct was transformed into ZH11 calli through Agrobacterium-mediated transformation. Positive T0 transgenic progeny were used to detect GUS activity as previously described [Total RNA was isolated from roots, stems, leaves, flowers, and developing grains at 5, 10, 15, 20, 25, and 30 days after fertilization (DAF) using an RNA extraction kit  following the manufacturer’s instructions. Total RNA was reverse transcribed by priming with oligo (dT18) based on the instructions from the ReverTra Ace qPCR RT Kit . Gene expression was quantified by reverse transcription quantitative PCR (RT-qPCR). RT-qPCR was performed using SYBR Green Real-time PCR Master Mix (Toyobo) and a Light Cycler 480 system . Relative gene expression was calculated using the 2T method . Primer escribed .OsLEA1b coding sequence was cloned into the pAN580-GFP vector capable of generating an OsLEA1b-GFP fusion protein. The fusion construct and an empty vector control (PAN580-GFP) were separately transformed into rice protoplasts [The toplasts . GFP fluoslea1b mutants and the wild type were randomly selected. Images were obtained using a HITACHI S3400N scanning electron microscope . Scanning electron microscopy was performed as described previously [oslea1b endosperms at 10 DAF were used to prepare semi-thin sections (800 nm). Samples were stained with I2-KI for 5 s and subsequently examined under a light microscope . All treatments were performed as described previously [oslea1b plants were analyzed using transmission electron microscopy . Samples were treated as described previously [Dry and completely brown rice grains of eviously . To exameviously . To obseeviously .oslea1b mutant and wild type were quantified following a previously established method [Pseudomonas amyloderamosa isoamylase (Megazyme) and then analyzed by capillary electrophoresis . Total starch and amylose contents were measured using the Megazyme K-TSTA and K-AMYL  kits, respectively. Lipid and protein contents in d method . Gel cond method . To dete2O2 levels were determined by visible spectrophotometry using a malondialdehyde (MDA) assay kit , proline assay kit (A084-3-1), and hydrogen peroxide assay kit , respectively. Peroxidase activity in leaves was measured using an Amplex™ Red Hydrogen Peroxide/Peroxidase Assay Kit (A22180). MDA, proline, and H2) test and independent samples t-test  were used for statistical analysis using IBM SPSS Statistics 26 software.Data are presented as the means ± standard deviation (SD), shown by error bars. The chi-square (χOsLEA1b, a heat-stress-responsive gene in rice grain filling. OsLEA1b is highly expressed in the endosperm, and its coding protein localizes to the nucleus and cytoplasm. The oslea1b mutants had abnormal compound starch granules in the endosperm cells and chalky endosperm with significantly decreased grain weight and grain number per panicle, as well as significantly lower contents of starch, protein, and lipid compared to the wild type. Moreover, OsLEA1b was induced by high temperature and drought stress. The oslea1b mutants showed more sensitivity to high temperatures. These results indicate that OsLEA1b regulates starch biosynthesis and influences rice grain quality, especially under high temperatures. In conclusion, this research provides a valuable gene resource and strategy for genetic improvement of rice grain quality.In summary, we identified


Aquatic animals such as fish and cetaceans can actively modulate their body stiffness with muscle to achieve excellent swimming performance under different situations. However, it is still challenging for a robotic swimmer with bionic propulsion mode to dynamically adjust its body stiffness to improve the swimming speed due to the difficulties in designing an effective stiffness adjustment structure. In this paper, based on the special torque mode of a motor, we propose an active variable stiffness control method for a robotic dolphin to pursue better swimming speed. Different from a variable stiffness structure design, a torque control strategy for the caudal motor is employed to imitate the physical property of a torsion spring to act as the variable stiffness component. In addition, we also establish a dynamic model with the Lagrangian method to explore the variable stiffness mechanism. Extensive experiments have validated the dynamic model, and then the relationships between frequency and stiffness on swimming performance are presented. More importantly, through integrating the dynamic model and torque actuation mode-based variable stiffness mechanism, the online performance optimization scheme can be easily realized, providing valuable guidance in coordinating system parameters. Finally, experiments have demonstrated the stiffness adjustment capability of the caudal joint, validating the effectiveness of the proposed control method. The results also reveal that stiffness plays an essential role in swimming motion, and the active stiffness adjustment can significantly contribute to performance improvement in both speed and efficiency. Namely, with the adjustment of stiffness, the maximum speed of our robotic dolphin achieves up to 1.12 body length per second (BL/s) at 2.88 Hz increasing by 0.44 BL/s. Additionally, the efficiency is also improved by 37%. The conducted works will offer some new insights into the stiffness adjustment of robotic swimmers for better swimming performance. Over the eons of evolutionary time, marine animals have adapted perfectly to the complex aquatic environment with their unique body structure and motion mechanism. Thereinto, fish and some cetacean animals with undulatory propulsion mode, such as Tuna and dolphin, are among the best candidates in terms of underwater swimming performance including speed, maneuverability, efficiency, and stealthiness, which provides extensive inspiration for the development of underwater robots ,2,3,4,5.From the perspective of motion mechanism, underwater swimming robots are usually designed with multi-joint rigid discrete links to pursue better swimming performance. The optimization control of joints was conducted to imitate the propulsion mode with the fish body wave model. For example, Yu et al. combined the dynamic model and particle swarm optimization (PSO) algorithm to optimize the central pattern generators (CPG) parameters for an enhanced performance . Hong etIn nature, the body of fish or dolphins is actuated by continuous flexible muscles that can regulate the body stiffness actively to achieve outstanding swimming performance at different locomotions. Inspired by this phenomenon, many researchers have been devoted to the investigation of variable stiffness mechanisms for better swimming performance . Park etIn this paper, we mainly focus on the performance optimization of a self-propelled robotic dolphin with a torque control-based variable stiffness mechanism. To achieve this goal, a robotic dolphin with powerful dorsoventral propulsion is introduced. Different from the traditional variable stiffness structure, we take advantage of a unique torque mode of the actuation motor to imitate the compliant component in the caudal joint. Namely, according to the properties of torsion springs, a control method is employed to adjust the torque output of the joint at different joint angles. Therefore, the stiffness adjustment of the joint can be also achieved easily by setting different stiffness coefficients. A dynamic model of the robotic dolphin with the active variable stiffness mechanism is built to analyze the influence of control parameters and stiffness on swimming performance, and an online performance optimization scheme is proposed to improve the swimming speed. Finally, extensive experiments have validated the effectiveness of the proposed variable stiffness mechanism on performance improvement.The rest of this paper is organized as follows. Drawing from a dolphin, a novel robotic dolphin has been developed. As for the developed robotic dolphin, the waist joint and caudal joint are responsible for the dolphin-like propulsion. The motor of the waist joint plays a key role in overcoming the drag, in this paper, the waist joint is actively actuated by position control mode  to oscillate in reciprocal sinusoidal form. To imitate the flexible propulsion, the caudal joint is chosen as a compliant joint. A flexible component in a swimming robot usually performs as a passive bent, which provides a torque to keep its original shape. Inspired by this, by controlling the output torque of the caudal motor, the caudal joint can imitate a compliant joint. Owing to the excellent effect in speed enhancing , the cauTo imitate the torsion spring’s flexible physical property, we can employ the cycle synchronous torque mode (CST) of the motor driver to control the output torque . As illuTo optimize the stiffness of the caudal joint at different undulation frequencies, a dynamic model is established. With the specific mechanical design, the developed robotic dolphin is not proficient in yaw maneuvers. In addition, the main goal is to explore the optimal stiffness with frequency, therefore, only the locomotion in the vertical plane is considered. In this study, the caudal joint is taken as a compliant joint with a torque control strategy to explore the variable stiffness mechanism of the robotic dolphin, therefore, the joint angle is unknown and the Lagrangian dynamic approach is employed ,31.ith link and axis As illustrated in According to the definitions of frames and notations, the kinematics can be derived easily. The position vector of an arbitrary point in link The translational velocity and angular velocity can be obtained by taking the time derivative of the position vector and angle vector: ith link, In the design of the robotic dolphin, the caudal joint is utilized to imitate a compliant joint with the capability of variable stiffness. Concretely, in the proposed torque control strategy, a torque mode is employed to actuate the caudal joint which can be regarded as a passive joint with a torsion spring. To describe the robot system completely, the generalized coordinates are selected as As for the caudal joint, a torque control strategy is proposed to perform passive rotation with a torsion spring-like compliant component. Namely, the variable torque acting on the caudal joint can be expressed in the following form:L is the Lagrangian, Ultimately, the Lagrange equations can be expressed as follows:The free-swimming motion is achieved through the interaction between the robotic dolphin and the surrounding fluid. Therefore, the generalized forces are produced by the hydrodynamic forces. In this paper, for the structure characteristics of each link, the Morrison equation and lift/drag model are utilized to evaluate the hydrodynamic forces ,33, respAs for the body, the Morrison equation where the hydrodynamic forces include added mass forces and drag forces is adopted . The addith link by integrating the forces along The drag force exerted on the per unit length of the link is generated by the pressure difference and friction viscosity, which is given as follows: Additionally, the lift and drag model is employed to calculate the hydrodynamic forces acting on the flukes with airfoil profile and given as below.ith link in the direction of ith link in the direction of ith link and relating to ith link and relating to Finally, the generalized forces and moments can be calculated as follows: In the established dynamic model, many parameters including physical parameters and hydrodynamic parameters should be identified to simulate the swimming motion. By using the software of SolidWorks, we can directly measure or calculate the physical parameters of the robotic dolphin, as tabulated in V indicates the speed of robot’s center mass (CM) and can be easily obtained with the velocity To identify the hydrodynamic parameters of the built dynamic model, some experiments were conducted in an indoor pool whose size is 5 m in length, 4 m in width, and 1.2 m in depth. During the experiments, the swimming motions of the robotic dolphin can be recorded by a global camera located above the pool. With the help of a customized motion measurement system in our previous work , we can The obtained experimental results from a small pool are more accurate and are suitable for the parameter identification of the built dynamic model. In this paper, we primarily concentrate on the swimming speed, therefore, we employ the measured speed as the adopted experimental result. Concretely, the swimming speed curves with a frequency of 2.53 Hz and a stiffness of 0.31 NThe torque control of the caudal motor is the foundation of the proposed active variable stiffness mechanism. The drive board provided by the Maxon motor manufacturer is feature-rich and can provide extensive information, such as current, voltage, position, torque, and so on. The output torque of torsion springs is determined by both the joint angle and stiffness coefficient  and three stiffness including  is selected as an indicator and used to analyze the effect of frequency and stiffness. The COT indicating the consumed power during the robotic dolphin swims a unit distance is defined as the following form:The evaluated COTs are presented in During swimming motion, the interaction between a robotic dolphin and water is complicated, therefore, the relationship between frequency, stiffness, and swimming speed is challenging to clarify. The means of experiment is usually time-consuming and toilsome. In this study, we take advantage of the validated dynamic model to evaluate the swimming performance in different frequencies and stiffness, which provides a simple and convenient approach. To analyze the influence of frequency and stiffness on swimming motion, we estimate the swimming speed with the established dynamic model. The frequency in the range of  with an interval of 0.5 Hz and the stiffness in the range of [0.2 NIn nature, biological dolphins can modulate their body stiffness to obtain better swimming performance in time. As for the robotic dolphin with a compliant joint, there also exists an optimal stiffness at a certain condition to achieve maximum swimming speed. The optimal stiffness is mainly determined by oscillation frequency. To explore the proposed variable stiffness mechanism, the optimal coefficient can be estimated with a dynamic model, which helps to achieve high performance including speed and efficiency under a complex environment. Different from the existing variable stiffness mechanism with special structure design, the torque control-based variable stiffness mechanism only requires the parameter adjustment of the motor, which simplifies the performance optimization significantly. In this paper, we mainly focus on pursuing better speed, therefore, by integrating the established dynamic model and the proposed stiffness variable mechanism, we can achieve performance optimization online easily, as shown in In this paper, we adopt the torque control strategy for a caudal motor to imitate the torsion spring characteristics. Namely, the motor outputs a torque that is relevant to the joint angle with a given stiffness coefficient. Therefore, in the whole rotation cycle of flukes, the motor always consumes energy. Compared to the robot with a real torsion spring, our robotic dolphin with a variable stiffness mechanism is deficient in efficiency. A compliant component with a single stiffness can store and release potential energy, where it plays a role of power modulation to improve the speed and efficiency simultaneously. However, to possess the ability of stiffness adjustment, it is inevitable to require extra energy. Additionally, different from the traditional variable structure, the proposed active variable stiffness mechanism is essentially a torque control of a caudal motor that conforms to the conventional system design, therefore, it is easy to design a robotic fish. The torque control accuracy has been validated by the error analysis between target torque and real torque, which can guarantee the reliability of the proposed variable stiffness mechanism. More importantly, in this paper, only a simple situation imitating the stiffness variation characteristic of torsion springs is considered. Based on the proposed active variable stiffness mechanism with torque control of the motor, more torque control strategies, such as changing the stiffness coefficients in one oscillation cycle can be performed to pursue better swimming performance. Through integrating the advantages of the adopted actuation motor in drive mode and the propulsion mechanism of the caudal fin, there is still great exploration space in terms of torque control strategies of the caudal joint to improve the robotic dolphin’s performance, which exhibits strong adaptability.The interaction between a robotic dolphin and surrounding fluid is extremely complicated. To simplify the analyses, the hydrodynamic forces are estimated with some assumptions, such as fewer fluid factors and hydrodynamic coefficients are considered, which mainly causes the errors between experiment results and simulated results. In this study, only the waist joint with frequency, caudal joint stiffness, and swimming speed are considered. Therefore, based on the dynamic model, we directly present the relationships among these parameters. Towards a multi-joint robotic fish with multi-performances, some intelligent optimization algorithms , such asFinally, we compare the swimming speeds of some robotic fishes with variable stiffness mechanisms, as tabulated in In this paper, based on a special torque drive mode, we have proposed an active variable stiffness control method for a robotic dolphin to improve swimming performance. Inspired by the locomotion characteristic of a flexible joint with torsion springs, a torque control method is proposed for the caudal joint of a robotic dolphin to imitate the variable stiffness mechanism. In addition, a dynamic model for the robotic dolphin is established with the variable stiffness caudal joint, and its effectiveness is also validated. Extensive experiments and model-based simulations are all conducted to explore the influences of key parameters (frequency and stiffness) on swimming performance. These results reveal that the frequency and stiffness play important role in swimming performance. At different actuation frequencies, the stiffness can be optimized to improve swimming speed and efficiency. In addition, the results also demonstrate the capability of variable stiffness, validating the effectiveness of the proposed control method. By adjusting the stiffness of the caudal joint, our robotic dolphin achieves a maximum speed of 1.12 BL/s at a frequency of 2.88 Hz. Finally, combining the built dynamic model and the variable stiffness control method, an online performance optimization scheme for the robotic dolphin is proposed to guide performance improvement.In future work, we will design a tendon-driven body with tensegrity structure to further enrich the stiffness modulation capability. In addition, we will utilize the technology of computational fluid dynamics (CFD) to further explore the variable stiffness mechanisms of swimming robots in performance improvement.