We aimed to assess food intake and adherence to the Mediterranean Diet in patients with T1D compared with nondiabetic individuals.This was an observational, multicenter study in 262 T1D subjects and 254 age- and sex-matched nondiabetic subjects. A validated food-frequency questionnaire was administered. The alternate Mediterranean Diet Score (aMED) and alternate Healthy Eating Index (aHEI) were assessed. The clinical variables were also collected. The analysis of data included comparisons between groups and multivariate models.p < 0.001), processed meat (p = 0.001), fatty fish (p = 0.009), fruits and vegetables (p < 0.001), nuts (p = 0.011), legumes (p < 0.001), potatoes (p = 0.045), and bread (p = 0.045), and a lower intake of seafood (p = 0.011), sweets (p < 0.001), and alcohol drinks (p = 0.025). This intake pattern resulted in a higher consumption of complex carbohydrates (p = 0.049), fiber (p < 0.001), protein (p < 0.001), polyunsaturated fatty acids (PUFA) (p = 0.007), antioxidants (p < 0.001), vitamins (p < 0.001), and minerals (p < 0.001). The frequency of patients with T1D and low aMED score (23.2%) was lower than that of the controls . The overall multivariate analysis showed that, among other factors, being a T1D subject was associated with improved aMED and aHEI scores (p = 0.006 and p < 0.001). In patients with T1D, residing in a nonurban area was associated with improved aMED and aHEI scores (p = 0.001 and p < 0.001).Compared to the controls, the patients with T1D had a higher intake of dairy products  contains supplementary material, which is available to authorized users. Medical nutrition therapy (MNT) and physical activity, in addition to insulin therapy, are cornerstones in the management of type 1 diabetes (T1D). MNT produces health benefits on glucose control, lipid profile, weight management, and maintenance of muscle mass . DiffereIn our region, the dietary habits are identified by the Mediterranean Diet (MedDiet). This dietary pattern includes moderate intake of energy, low intake of animal fat, high intake of fruits, vegetables, whole grains, legumes, olive oil and moderate intake of fish, poultry, and red wine, together with regular physical activity (25–30 min every day) , 7. A nuSeveral studies in T1D have assessed the comparison between the dietary habits and the American Diabetes Association (ADA) or the European Association for the Study of Diabetes (EASD) nutritional recommendations –15. HighBecause patients with T1D receive regular nutritional education, we hypothesized that they would have healthier eating habits than nondiabetic subjects. In a Mediterranean country like ours, MNT includes recommendations for adhering to a MedDiet pattern . HoweverThis was an observational and two-center study. A sample of 513 participants was recruited: 259 patients with T1D and 254 nondiabetic subjects matched for age and sex in each location [healthcare areas of Lleida (center 1) and Barcelona (center 2)] between January 2013 and May 2015. The location of Lleida is a mixed rural and semi-urban area, and Barcelona is a fully urban area. The cases were patients diagnosed with T1D who were regularly cared for at their reference hospital . Participation in the study was offered to outpatients with T1D in both departments until the study sample was complete. The inclusion criteria of cases were as follows: diagnosis of T1D with a duration of more than 1 year; age > 18 years for both cases and control participants. The exclusion criteria for both groups were as follows: being a healthcare professional, participants who showed physical and cognitive deterioration as dementia, mental diseases, and known cardiovascular diseases  or previous diabetic foot disease, pregnancy, and renal insufficiency (estimated glomerular filtration rate < 60 mL/min). Furthermore, for T1D, we excluded patients with a condition that requires additional MNT measures, i.e., macroalbuminuria . The control participants were also excluded if they had an HbA1c value ≥ 6.5% [In center 1, from an initial sample of 170 patients with T1D who were contacted to participate in the study, 128 subjects accepted to participate, and 3 were excluded because of the exclusion criteria, resulting in a target sample of 125 . However, 3 additional subjects were excluded after their inclusion because of the identification of exclusion criteria, resulting in a final sample of 122 cases. The corresponding control group was identified from a population-based study, which was performed simultaneously in that center [In center 2, a total of 160 patients with T1D were contacted; 148 agreed to participate. From this group, 11 were excluded, because they had met the exclusion criteria. The final sample was 137 patients with T1D. The control group was recruited at the same time as the cases; from a sample of 160 controls, 151 accepted and 22 were excluded because of the exclusion criteria, resulting in a final sample of 129 controls .The study was approved by the local Ethics Committee of both centers, and written informed consent was obtained from all the participants.The clinical variables collected for each study group are shown in Table The Food-Frequency Questionnaire Consumption (FFQC) was administered by personal interview by specialized and trained researchers. This is a semiquantitative questionnaire, validated and adapted for the Spanish population and based on the Nurses’ Health Study , 29. Theα = 0.05) and a statistical power of 80% (β = 0.2), a sample of 125 cases and 125 controls was necessary to detect a statistically significant difference in the aMED score. Finally, based on the availability of a sufficient number of potential participants, we aimed to conduct the study in two different geographical areas, which also allowed us to increase the statistical power of the study.The sample size was calculated to detect the differences between the study groups based on previous results from a pilot study performed to assess the potential output (60 cases and 60 controls) in which the mean and standard deviation values of aMED were estimated in both groups: T1D 4.2 ± 1.4 and controls 3.7 ± 1.4. Based on this difference, using a significance level of 5% (n) and relative frequencies (%). Statistical significance was assessed using the Chi-squared test to assess the differences in the frequencies. The mean of clinical determinations, daily food and nutrient intake, and the dietary quality index were compared using Student’s t test. The “BH” (aka “fdr”) and “BY” methods of Benjamini, Hochberg, and Yekutieli were performed; they control the false discovery rate and the expected proportion of false discoveries among the rejected hypotheses. The false discovery rate is a less stringent condition than the family-wise error rate, so these methods are more powerful than the others. The multivariate regression models were developed to analyze the relationship between group (case and control) and dietary quality index adjusted by the potential confounders. The models were adjusted by variables that were statistically significant in the bivariate analysis or were clinically associated with diabetes. The conditional logistic regression models were designed to explain low aMED adherence (aMED low: 0–2) and adjusting linear regression models were performed for aHEI. The goodness-of-fit assumption using the Hosmer–Lemeshow test for logistic models was assessed, and for the linear regression model, the Kolmogorov–Smirnov test was performed. The estimate measures were the odds ratio (OR) with 95% confidence interval (95% CI) and with logistic regression models, and the effect (β) and standard error (± SE) with linear regression. In all the tests, a p < 0.05 was considered to be statistically significant.An initial descriptive comparison between groups of all variables was performed. The quantitative variables were summarized using the mean and standard deviation (SD) values, and the qualitative variables were summarized using the absolute (p < 0.001) than the control group. However, the patients with T1D performed regular physical activity more frequently (p < 0.001), and had a high frequency of treatment for hypertension (p < 0.001) and dyslipidemia (p < 0.001). Furthermore, the patients with T1D showed a better lipid profile.The clinical and demographic characteristics of the study groups are shown in Table p < 0.001), processed meat (p = 0.001), fatty fish (p = 0.009), fruits and vegetables (p < 0.001), nuts (p = 0.011), legumes (p < 0.001), potatoes (p = 0.045) and bread (p = 0.045), and a lower intake of seafood (p = 0.011), sweets (p < 0.001) and alcohol drinks (p = 0.025).The patients with T1D had a healthier food intake pattern than the control group, as shown in Table p = 0.049), soluble and insoluble fiber (p < 0.001), protein (p < 0.001), and PUFA (p = 0.007); the latter included intake of omega 3 (p < 0.001), omega 6 (p = 0.016), linoleic acid (p = 0.016), α-linolenic acid (p < 0.001), eicosapentaenoic acid (EPA) (p = 0.049), and docosahexaenoic acid (DHA) (p = 0.016). Furthermore, subjects with T1D also showed a higher consumption of all vitamins , carotenoids (p < 0.001), and minerals . In contrast, the control group showed a higher intake of energy (p = 0.011), sugar (p < 0.001), stearic acid (p = 0.021), and alcohol (p = 0.002); they also had a higher dietary glycemic index (p = 0.002).Consistent with this food pattern, there was also a differential intake of nutrients . The patients with T1D showed a higher intake of complex carbohydrates (p = 0.009). Furthermore, the frequency of patients with T1D with poor adherence to a MedDiet (low aMED score) was lower (p = 0.019). However, both groups had a moderate mean aMED score . The multivariate logistic analysis showed that being a subject with T1D (p = 0.006), increased physical activity (p = 0.017) and increasing age (p = 0.008) were negatively related to a low aMED score; thus, all these variables were associated with a higher adherence to a MedDiet  (Table p = 0.011). Moreover, the mean of aHEI was relatively low in both groups . The multivariate linear analysis revealed that the factors associated with higher aHEI scores were the T1D group (p < 0.001), high educational level (p = 0.016), residing in the region of Lleida (p < 0.001), increased physical activity (p = 0.033), and increasing age (p = 0.006) .Patients with T1D also showed a higher aHEI score than the controls (1) Table . In addip = 0.001) , high educational level (p = 0.025), and age (p = 0.006). The male sex was negatively associated with aHEI (p = 0.008)   showing a high Canadian Healthy Eating Index (C-HEI); this finding is similar to our results , 22. HowWe must underscore that the fact of not residing in an urban area was the factor that was closely and consistently associated with improved dietary quality indexes in patients with T1D. This points to a relevant influence of the socio-geographical context in determining the dietary pattern of any given subject. In addition, discordant results found by different studies in T1D may, in part, be attributed to this factor. This is also an important finding for future studies, as the control group should be carefully selected.In our study, the subjects with a high educational level, regular physical activity, and older age showed an improved healthy lifestyle behavior independent of the disease duration. It is reasonable to think that a healthy lifestyle is more likely to be adopted in older patients and that it is maintained over time . In addiThe current study has several limitations. No relationship of direct causality can be established between the different variables associated with the MedDiet, because the changes in lifestyle habits produced over time cannot be addressed with a study like the current one. Although the current results point to the probable influence on the better dietary habits of T1D patients of the educational dietary intervention  that they receive from the treating healthcare professionals, we cannot conclude that this is the case. Unfortunately, we did not assess the knowledge of patients about the dietary management of diabetes. This study has several strengths. The large number of participants, the multicenter design, and a well-characterized sample allow us to establish the variability of different populations and lifestyles associated with them. Furthermore, we could study the different pattern in two different areas of the same region, North-Eastern Spain, that differed by their urban and rural/semi-urban locations. Moreover, this is the first study that assessed the adherence to the MedDiet between adults with T1D and a nondiabetic control group.In conclusion, in our Mediterranean region, adults with T1D showed a healthier food intake pattern and, specifically, an improved adherence to the MedDiet than their nondiabetic counterparts. However, additional research is warranted in this field and in the identification of the educational strategies that need to be adopted to enhance the adherence of patients with T1D to a healthy diet.Below is the link to the electronic supplementary material.Supplementary material 1 (DOCX 36 KB)Supplementary material 2 (DOCX 26 KB)Supplementary material 3 (DOCX 18 KB)Supplementary material 4 (DOCX 17 KB)


After 9.5 ± 6.9 years of follow-up, 87.29% of patients were disease-free and 12.71% had persistent disease; no patient had deceased. Patients aged <45 years old with extrathyroidal invasion tumor had greater baseline body weight and BSA than those without extrathyroidal invasion (median 79.5 kg versus 67 kg and 1.85 m2 versus 1.74 m2). Women with poorly differentiated tumor and patients aged ≥45 years old with distant metastasis presented greater weight loss during follow-up compared to patients without such characteristics . The relationship between body size and DTC evolution was not observed. In conclusion, higher weight and BSA were associated with a greater chance of extrathyroidal tumor invasion in younger patients. Specific subgroups of patients with aggressive disease presented higher weight loss. Young patients with higher BSA should be carefully treated due to possible worse prognosis related to increased incidence of extrathyroid invasion. Findings related to tumor aggressiveness and weight loss in specific groups deserve further mechanistic studies.Considering controversial data about the relationship between body size and prognosis of differentiated thyroid cancer (DTC), the current study aimed to assess the influence of body weight, body mass index (BMI), and body surface area (BSA) on DTC. We conducted a retrospective analysis of patients' records from the Thyroid Cancer Unit, assessing body size measures, clinical and laboratory prognostic factors, and disease evolution. 337 patients, aged 45.95 ± 13.04 years old, with BMI of 27.87 ± 5.13 kg/m Differentiated thyroid cancer (DTC) is a neoplasia with raising incidence and patients usually present long survival , 2. OverSome publications suggested an association of higher BMI and characteristics of thyroid tumor aggressiveness. Trésallet et al.  evaluateConsidering obesity is associated to higher incidence of malignant tumors and also to worse prognosis of different kinds of neoplasia, but there are no conclusive data regarding DTC, the current study was performed with the objective of evaluating the impact of body size measures in prognostic factors and evolution of disease, involving a greater number of different clinical and histological parameters and longer follow-up compared to previously published studies.This was a retrospective analysis of the hospital files from the Thyroid Cancer Unit of the University of Campinas. All patients were submitted to total thyroidectomy and were treated according to thyroid cancer guidelines between 1980 and 2015 . Patient2), overweight (25 ≤ BMI < 30 kg/m2), or obese (BMI ≥ 30 kg/m2) [2) = 0.007184 × weight (kg)0.425 × height (cm)0.725 [BMI (kg/m2) was calculated according to the guidelines from the World Health Organization using the formula weight (kg)/height (m) squared. Patients were categorized as normal weight (18.5 ≤ BMI < 25 kg/m0 kg/m2) . BSA wascm)0.725 . Since nWe also evaluated the following clinical and histological parameters of the cancer: age at diagnosis, ethnicity, family history of DTC, smoking history, follow-up time, presence of thyroid antibodies, size and number of malignant nodules, histology type and variant, vascular and lymphatic invasion, extracapsular or extrathyroidal invasion, histological differentiation grade, metastasis to lymph nodes and to distant organs, and tumor staging according to the tumor node and metastasis system (TNM) . PresencData were tabulated, and we performed an exploratory analysis through summary measures  in order to evaluate the impact of body size measures on parameters of DTC. Comparative analyses were performed by comparing median values of applicable variables using Mann–Whitney, Kruskal-Wallis, Chi-square, or Fisher exact tests. A significance level of 5% was considered significant in the analysis. All analyses involving anthropometry data were planned to be done by sex, because men and women's body composition is known to be different . The spl2 and mean initial BSA was 1.37 ± 0.21 m2. One-hundred women (33.9%) had initial BMI < 25 kg/m2, 107 (36.1%) presented 25 ≤ BMI <30 kg/m2, and 89 (30%) had BMI ≥ 30 kg/m2. The respective numbers for male patients were 9 (21.9%), 20 (48.8%), and 12 (29.3%). After mean follow-up time of 9.5 ± 6.9 years, mean BMI increased to 28.51 ± 5.34 kg/m2 and mean BSA to 1.76 ± 0.18 m2. Regarding the evolution of the disease, it was not possible to get accurate information in 38 files. From the remaining 299, 87.29% of patients were classified as disease-free state and 12.71% as persistent disease state. No patients included in the analysis have died due to DTC or were classified as recurrent disease state. Detailed clinical and laboratory characteristics of patients are described in Tables From 939 patients in the Thyroid Cancer Unit's database, 337 presented all applicable anthropometry information in their files and were considered to the study. Majority (87.8%) of patients was female and had tumor histology of PTC 89.29%); mean age at diagnosis was 45.95 ± 13.04 years old. Mean initial BMI was 27.87 ± 5.13 kg/m9%; mean p = 0.0253 and 1.85 m2 versus 1.74 m2, p = 0.0096, respectively). During follow-up, such difference was sustained, considering last assessment of weight and BSA . Such results are in line with the association of higher body measures and occurrence of more aggressive cancer. Female patients with poorly differentiated tumors have lost more weight compared to patients with well-differentiated tumors  from the same gender. Also, patients aged 45 years old or more and who presented distant metastasis during follow up presented greater weight loss during evolution than patients of the same age without such metastasis . This was an unexpected finding, which was not related to the initial aim of assessing the relationship between higher body size measures and tumor evolution. Those were the only significant findings, and they are shown in details in The analysis according to age group evidenced patients aged less than 45 years old who presented tumors with extrathyroidal invasion had higher weight and BSA at diagnosis than those patients without such invasion , follicular thyroid cancer (FTC) histology , distant metastasis at diagnosis or during evolution (p < 0.0001 for both), metastasis for lymph nodes at diagnosis (p = 0.0002), vascular and lymphatic tumor invasion (p = 0.0008), extracapsular invasion (p < 0.0001), and extrathyroidal invasion (p = 0.0036). The multivariate analysis of the same tumor factors showed only initial lymph nodes metastasis to be correlated with CDT prognosis .The analysis of prognosis considering comparison of median values between disease-free and persistent disease groups of patients evidenced that the following variables were associated with higher occurrence of persistent disease: size of nodules  by sex, there were no significant differences between disease-free patients group and persistent disease patients group . Also, cOur group of patients is a representative sample of DTC, with baseline characteristics which are comparable to previous publications –24. TypiRegarding survival rates, few patients deceased in all those published studies and survival analysis could not be properly done, such as in our study , 10. ForAlthough our sample size was smaller than those in some previous studies, our follow-up time was the longest (9.5 years). Even so, it was not possible to observe an effect of body size measures in prognosis. This could be explained because DTC mortality rates are generally much lower than other tumors . Then we2 increase in BMI was associated with microscopic extrathyroid invasion (odds ratio of 1.23), independent of age and gender [Despite that, we found one important significant association regarding body size measures and one DTC prognostic factor in our study, which was the link between higher weight and BSA with higher chance of extrathyroid tumor invasion in patients aged <45 years old. The study led by Kim et al. evidenced association between BMI and extrathyroid invasion, by showing that a 5 kg/md gender . Other Sd gender . In lined gender , and thed gender , 27. Extd gender , 25.α) with concomitant state of TNFα resistance and increase in interleukin-6 (IL-6) and leptin levels [The mechanism for the relationship between higher body size and extrathyroid invasion, which is one feature associated to bad prognosis, could maybe be explained by the same factors that have been hypothesized to explain the association between obesity and thyroid cancer. One of them is the increased serum level of TSH, frequently observed in obese patients, since TSH is a growth factor for thyroid cells and a predictor of malignancy in thyroid nodules . Other in levels . Those sn levels , 32.However, the association between higher body size and higher chance of extrathyroid invasion was only evidenced in patients aged <45 years old. Such occurrence could be explained by the fact that older patients often present other features associated with higher chances of extrathyroid invasion, which reduces the power of influence of body size measures on the feature in this age group. It is already known that older patients have a higher risk for extrathyroid invasion compared to younger patients . Then, iOther finding in our analysis was the greatest weight loss over time in women presenting poorly differentiated tumors and in patients aged 45 years old or more who presented distant metastasis during follow-up. Both cases reveal the association of more aggressive cases of DTC and reduction of weight. Our study is the only one that evaluated this aspect of weight change over time, and, therefore, it is not possible to make a comparison with previous results –12, 27. The present study found weight and BSA to be associated with a higher chance of extrathyroidal tumor invasion in patients aged <45 years old. Nevertheless, it was not possible to observe the influence of body size on DTCs prognosis or evolution. Considering extrathyroidal tumor invasion is an important risk factor for DTC-related mortality, younger patients with higher weight and BSA should be carefully evaluated and treated to avoid worse evolution of the disease.On the other hand, findings related to tumor aggressiveness and weight loss in specific groups deserve further mechanistic studies, since it is an unprecedented and unexpected report in the field of DTC.


Prostatic diseases are common and mostly associated with enlargement of the accessory gland. Thus, determining the prostate size has become a main criterion for evaluating prostate health status. Computed tomography (CT) is recommended as a beneficial tool for evaluating prostate size, morphology and surrounding tissues. The purpose of this study was to establish an accurate procedure for volume estimation and afterwards evaluate the prostate volume in CT. Data of 95 dogs were analysed  using the slice addition technique with the Amira program. Accuracy of volumetric measurements by CT was validated by comparing them with those of phantoms of known volume. Patients were grouped according to age  and prostate morphology in CT . The length of the sixth lumbar vertebra was measured to relate prostate volume to body size. This ratio was generated to compare prostate volume between the groups, irrespective of body size (ratio volume = Rv).3. The mean ratio volume was 1.3 in intact male dogs, this being significantly higher than in neutered dogs (0.7). The lowest ratio volume values were found in group H for intact (Rv = 0.9) and neutered dogs (Rv = 0.6), followed by group I  and C . The length of the sixth lumbar vertebra was well correlated with the prostate volume , while age exhibited a correlation only in intact dogs .A high correlation between the CT-derived and phantom volume was found. Overall, the mean prostate volume was 58.6 cmThe present study is pioneering in applying a slice addition technique to volumetric measurements of the prostate gland in CT, resulting in a highly precise method. Volumetric measurements of the canine prostate gland in CT images provide information about the prostate structure, castration status, age and body size of the patients. Therefore, prostate volume is a relevant parameter for evaluating prostate health status.The online version of this article (10.1186/s12917-019-1778-z) contains supplementary material, which is available to authorized users. Prostatic diseases including benign prostatic hyperplasia (BPH), prostatitis, abscesses and cysts are common, especially in older dogs . SymptomSchulze et al. measured prostate volume in CT with a formula of an ellipsoid body but accuracy was not verified . HoweverLee et al. and Choi et al. used a volume-rendering software tool to measure the prostate volume in CT data. The actual prostate volume was unknown in these studies , 19.The slice addition technique is recommended for measuring the volume of different organs like canine kidney, liver and spleen . NonetheThe aim of the present study is to measure the canine prostate volume using a slice addition technique. The accuracy of this technique shall be verified by the usage of phantoms with known volume as well as by volumetric measurements of canine prostates ex vivo. Furthermore, the volume of the prostate gland in dogs shall be analysed with the validated technique regarding differences due to age, body size and prostate structure. In past literature, the length of the sixth lumbar vertebra was used to relate prostate dimensions to the body size of the examined dogs –19, but In this retrospective study, CT data sets of dogs that were presented to the Small Animal Clinic, University of Veterinary Medicine Hannover, Foundation between October 2007 and August 2017 were reviewed. The following criteria were established for patients to be included in this study: male intact dogs aged eight months or older were included if abdominal CT examination data sets with and without contrast agent and no imaging artifacts such as high density streaks caused by metal implants existed. The scans should include the whole prostate as well as the sixth lumbar vertebra to put the prostate size in relation to body size. Dogs castrated by hormone substitution were excluded. Ninety-five patients met the criteria. The mean age was 7.6 years and mean body weight 28.4 kg. Fifty-eight male intact (mean age 7.3 years/mean weight 30.8 kg) and 37 neutered male dogs (mean age 8.2 years/mean weight 25.5 kg) were included in this study . Abdominal CT scans were performed in dorsal or ventral recumbency with a voltage of 120 kV, slice thickness of 2 mm, pixel sizes ranging from 0.15 × 0.15 mm to 0.84 × 0.84 mm and a pitch of 1.171. Depending on the patient’s body symmetry change, an automatic current selection function (DoseRight-D) modulated current during tube rotation, which resulted in different mAs-products between the dogs. Patients were anaesthetised with levomethadon , diazepam  and propofol . Anaesthesia was maintained by inhalation with isoflurane  during CT examination. First, a native scan was performed. A second scan was carried out after a non-ionic iodinated contrast agent  had been injected into the vena cephalica antebrachii or vena saphena lateralis via a power injector . The automatic scan in the prostate region of interest (ROI) was started 49 s after a threshold of 150 Hounsfield units in the aorta had been exceeded, which was tracked by a local ROI positioned in the aorta in transversal view. Afterwards, CT data sets were stored in DICOM format.Morphological analysis of the prostates was carried out with an image-processing workstation . Each prostate was analysed by the same observer (first author) to avoid inter-observer variability. The prostates of male intact and neutered dogs were assigned to one of three groups depending on the morphological appearance of the prostate in CT imaging: prostates with homogenous tissue structure (H), prostates with inhomogeneous tissue structure (I) and prostates with cystic tissue structure  , four-eight years (group 2) and older than eight years (group 3) .The allocation to age groups and structure groups are shown in Table The prostate volume was determined by a specific software . In order to determine the prostate volume by means of Amira, it was necessary to segment the gland from the surrounding tissue. Due to low contrast between the prostate and surrounding structures, no automatic segmentation was feasible and manual segmentation was performed in scans with contrast agent. Therefore, the prostate gland was encircled manually with the mouse curser tool in transversal image view within all slices. The urethra .To evaluate the accuracy of the CT-based volume measurement method, two different types of phantoms with known water-filled or water displacement volume were scanned , one irregular (ca. 4 × 4.5 × 3 cm3) and one with a “paraprostatic cyst” (ca. 5.5 × 4 × 3.5 cm3)  shaped like a prostate Figs.  and 5, o3 measuring cup filled with 100 cm3 water. The volume was directly read off the measuring scale (5 cm3 steps), subtracting 100 cm3. The phantoms were scanned in a water-filled bucket (100 mL) and CT scans were analysed with Amira as described above.The volume of the phantoms from this group was measured by water displacement, positioning the phantoms in a 250 cm3 measuring cup filled with 100 cm3 water. The volume was directly read off the measuring scale (5 cm3 steps), subtracting 100 cm3. Measurements were repeated twice and the mean value (+/− standard deviation) was determined.Furthermore, five  canine cadavers  which had been euthanised due to other diseases at Small Animal Clinic, University of Veterinary Medicine Hannover, Foundation, underwent CT examination without applying contrast medium . Differences in volume of the prostates in regard to castration or prostate structure were examined with a two-way analysis of variance and post-hoc Tukey test. The evaluation of accuracy of the assessment method was performed by Bland-Altman plots with GraphPad Prism  and linear regression analysis by SAS.Statistical analysis was carried out with SAS® Enterprise Guide® 7.1 . Normal distribution was analysed with the Kolmogorov- Smirnov test or Shapiro-Wilk test. Differences of the mean values between the groups were tested with the Ryan-Einot-Gabriel-Welsch multiple range-test. p = 0.766). Furthermore, linear regression exhibited a significant relation  between volumetric measurements and actual prostate values ) Fig. . The ANO3 (ranging from 0.6 to 1600.5 cm3). The mean ratio volume measured was 1.1 (ranging from 0.3 to 3.4). The ratio volume was statistically significantly affected by status of castration and prostate structure. The mean ratio volume was 1.3 in intact male dogs and 0.7 in neutered dogs. The prostate ratio volume of intact male dogs was significantly higher than that of neutered dogs (p < 0.001)  as well as between structure I and C (p = 0.020). No statistically significant differences in volume were found between the structures H and I. In neutered dogs, the volume of prostates grouped as structure H and I as well as structure H and C differed statistically significantly . No significant difference in prostate volume was found between structure I and C  and castrated dogs (Rv = 0.6), followed by inhomogeneous prostates . Canine prostates with cystic structures exhibited the highest mean volume values in intact and neutered dogs . In intact dogs, differences in volume were significant between structure H and C ( and castp < 0.001), but no significances were found in castrated male dogs , but not in neutered dogs (p = 0.407) (Table p < 0.001) and castrated dogs . The correlation was higher in intact male dogs Additional file 2:Phantom and cadaver measurements PVC = polyvinyl chloride; m = male intact, n = male neutered; H = homogeneous; I = inhomogeneous; C = cystic; − = no data available. (XLS 26 kb)


In this study, we investigated how vection strength depends on motion direction. Participants observed random-dot optical flow in a circular field and rated the perceived vection strength. Dot movement was systematically controlled using the following angles: 0° (up), 30°, 45°, 60°, 90°, 120°, 135°, 150°, and 180° (down). We found that vection strength depended on motion direction and was weaker in the oblique directions than cardinal directions. Thus, the effect of motion direction on vection strength was variable, as seen in the shape of the oblique effect.In some phenomena of visual perception, the motion direction of visual stimuli can affect perception. In particular, asymmetries between oblique directions and cardinal  directions have been reported and are known as  Numerous characteristics of vection have been reported since Brandt, Dichgans, and Koenig first investigated this interesting illusion in 1973.Stationary observers perceive self-motion when a moving visual stimulus is presented in their visual fields. This illusory self-motion is termed cardinal directions along with forward and backward) are very important for controlling the body, such as when standing vertically or walking on a horizontal floor. Especially for vection, the direction of gravity is an important factor. Several studies have shown that vection is closely related to and depends on the posture of the observer in relation to the direction of gravity, such as sitting, lying down  induced stronger vection than forward self-motion , objects also move in oblique directions. The visual system in the brain sometimes performs differently in oblique and cardinal directions, and this asymmetry is known as the Many studies have shown that oblique effects occur when the head or body is tilted. Examples can be found in visual search , 1999, oThese studies indicate that in addition to vection, oblique effects also depend on the direction of gravity and imply that vection strength has an oblique effect. Although to our knowledge, none have compared vection strength between oblique and cardinal directions. However, some studies have focused on the asymmetry of oblique directions in vection. To investigate the effect of visual field, Vection is closely related to perception of visual motion. Global motion of optical flow which induces vection is initially processed in low-level motion detector, which is common between vection and visual motion perception. Past studies have argued that some types of motion perception imply the existence of oblique effects. For example, In this study, our primary goal was to investigate the effect of motion direction on vection. Vection strength was measured in various directions, including oblique and cardinal directions. To determine whether the direction effect can be modified by oblique motion, vection strength was compared between cardinal and oblique directions (Experiment 1). We also investigated whether the effect of motion direction on vection is caused by the intensity of the motion signal (Experiment 2). In both experiments, visual stimuli were presented in a circular field and were symmetric in all directions. This allowed us to directly compare motion-direction conditions without any effects related to the asymmetry of stimuli. In addition, we employed commonly used indices of vection strength, which differed from the unconventional measures found in Stimuli were generated and controlled with Matlab  and Psychotoolbox-3  on a perEach pixel of the plasma display was a rectangle and arranged on a square-grid pattern as is usual on most displays. This means that the stimuli that were intended to be symmetrical were slightly asymmetric. However, the pixels were very small and stimuli were drawn by antialiased subpixels. Furthermore, the experimental stimuli induced suprathreshold motion signals. Therefore, any pixel asymmetry had very weak or no effect.http://www.senotake.jp/stimulus/2016/). White dots were placed in a circular field at the center of a black background. Dot luminance was 16.2 cd/m2 and that for the background was less than 0.01 cd/m2. Dot density was 0.54/deg2. The diameter of the central dot field was 82° and the size of each dot was 0.30°. Dots were antialiased. In addition, red fixation point was placed at the center.Stimuli were random-dot optical flows (All dots moved coherently in a single direction at a constant speed (33 deg/s), with a duration of 40 seconds. The angle of motion direction was varied over 16 conditions; 0° (up), ±30°, ±45°, ±60°, ±90°, ±120°, ±135°, ±150°, and 180° (down). The positive values indicate rightward motion, and negative values indicate leftward motion.no vection) to 100 (very strong vection). Very strong vection meant that observers perceived vection as if they were actually moving in the scene very naturally by real locomotion. Thus, if participants perceived that they were moving about half as much as natural self-motion, they would rate vection strength as “50.” They were allowed to choose a rating greater than 100 if necessary, but none of them did so. This method is the same as we have used in several studies . All participants were healthy, had a normal or corrected-to-normal vision, and no history of vestibular system diseases. No one (aside from the authors) was aware of the purpose of the experiment.Thirteen adult volunteers participated in this experiment . In addition, post hoc multiple comparison analyses (Ryan’s method) were performed to determine whether any differences between pairs of direction conditions were significant. To further investigate the oblique effect of vection strength, The results  were almF = 6.63, p < .001, η2 = .21; latency: F = 5.18, p < .001, η2 = .10; and duration: F = 10.48, p < .001, η2 = .24. In addition, Ryan’s test for multiple comparisons indicated significant differences (p < .05) for magnitude , latency , and duration . The other comparisons were not statistically significant.t(12) = −3.60, p = .004, r = .33; latency: t(12) = 3.48, p = .005, r = .84; and duration: t(12) = −5.50, p = .000, r = .59.The initial analyses indicate that vection strength depends on motion direction and the post hoc analyses support the oblique-effect hypothesis such that vection strength during the oblique conditions was weaker than during the cardinal conditions, as evidenced by the lower magnitudes, longer latency, and shorter duration. Next, we directly compared the averages between diagonal oblique data (45° and 135°) and cardinal data  for each vection index. The analyses confirmed that vection strength differed significantly between oblique and cardinal conditions: magnitude: t(11) = 4.64, p = .001, r = .76; latency: t(11) = 1.80, p = .050, r = .98; and duration: t(11) = 3.62, p = .004, r = .90. Therefore, we do not think that the imbalanced design was the source of the oblique effect in the initial experiment.Although the results from the statistical analyses support the hypothesis that vection strength has an oblique effect, this conclusion remains somewhat debatable because oblique motion stimuli  were more frequently presented to participants than cardinal motion stimuli . This imbalance in oblique/cardinal condition frequency might itself have biased perception through accumulation effects and led to the observed oblique effect. To address this issue, we repeated the initial experiment but included an equal number of oblique/cardinal conditions (one each: 0° and 30°). Nevertheless, as in the initial experiment, the results from 12 observers  clearly showed a significant oblique effect: magnitude: The results from Experiment 1 confirmed the effect of motion direction on vection and revealed that this effect varied depending on whether the direction of stimulus motion was cardinal or oblique. However, the effect of motion direction on vection strength could be just reflected the effect of motion direction on motion signal intensity, a finding that has been reported in previous studies e.g.,  regardleStimuli were the same as in Experiment 1, and motion directions were 0° (straight up) and −45° (upper left). Twelve naive observers  participated, and each condition was repeated 4 times for each individual. Participants were instructed to estimate the perceived smoothness and speed of stimuli via visual analogue scales. The values on the analogue scales were translated values between 1 and 100 and averaged across participants.t(11) = 0.59, p = .57, r = .17; speed: t(11) = 0.93, p = .37, r = .27. Thus, oblique effects were not observed for perception of smoothness or speed of motion.The results are shown in In this study, we examined the effect of motion direction on vection strength. The stimuli were symmetric, and unlike previous studies, the directions of optical flow motion were systematically controlled to cover numerous directions.The results of Experiment 1 showed a significant effect of motion direction on vection strength. In addition, further statistical analysis and the results from the additional experiment clearly revealed an oblique effect of vection strength which vection strength was weaker in the oblique conditions. Why is vection weaker in oblique directions? One possible explanation is related to the difference in subjective speed. Previous vection studies indicate that the speed of optical flow is a major factor that contributes to vection strength; However, the results of Experiment 2 showed that motion signals (subjective smoothness and speed) did not exhibit any oblique effect, despite using the same stimuli as Experiment 1. We can therefore conclude that the oblique effect on vection strength that we observed in Experiment 1 is a specific effect on vection and that the effect cannot be explained by the oblique effects of motion signals that has been shown in previous studies.The lack of an oblique effect of motion signal in Experiment 2 deserves consideration. One reason for this difference from other studies could be that while our stimuli induced suprathreshold motion signals, those in other studies were near threshold. Our suprathreshold signal likely masked small differences between oblique and cardinal directions.This physiological background should be related to the oblique effect in vection because some studies previously suggested that low-level processing in the visual cortex has an important role in producing vection strength. Vection studies have indicated that some vection processes are independent from motion perception processes e.g., . For exaOur suprathreshold optical flow induced an oblique effect on vection strength but did not induce effects on other motion signals (as shown by Experiment 2). This indicates that the optical flow was processed almost symmetrically between oblique and cardinal directions in terms of motion but asymmetrically in terms of vection. Therefore, long exposure to the flow might induce symmetrical inhibition in motion processing but asymmetrical inhibition in vection processing. This difference can be observed behaviorally as symmetry in MAEs and asymmetry in VAEs and is a very interesting topic for future study.When looking at the results for the cardinal directions, we did not find any differences in vection strength between them. This contrasts with previous studies, which have reported that vertical motion induced stronger vection than horizontal motion  and thatWe found that vection strength depends on the direction of optical flow motion; optical flow in oblique directions induced weaker vection than vertical or horizontal flow. This oblique effect could not be explained by any differences in the motion signal, which itself did not show any oblique effect. Thus, vection strength depends on direction, while motion signals do not—a distinction reflected in the different types of processing needed for motion perception and vection. Although this kind of dual measurement with the same stimuli has not been common in previous vection studies, we think that in the future, this simple method has the potential to reveal additional properties of the vection process.


Gastrointestinal (GI) cryptococcosis is rarely reported. Most cases were diagnosed during evaluation of comorbid conditions, incidental findings, or postmortem. Here, we present a case of Crohn’s disease with gastrointestinal cryptococcosis that resembled exacerbation of Crohn’s disease.Cryptococcus species. Disseminated cryptococcosis was diagnosed due to gastrointestinal involvement and presumed pulmonary involvement regarding the presence of an oval-shaped cavitary lesion on chest X-ray and computed tomography of the lung. Patient was successfully treated with amphotericin B followed by fluconazole with satisfactory result.A 64-year-old woman with Crohn’s disease (CD) was referred to Siriraj Hospital due to worsening of abdominal pain and watery diarrhea for 2 weeks. The dose of immunosuppressive agents was increased for presumed exacerbation of CD. Pathologic examination of tissue obtained from polypoid mass at ileocecal valve and multiple clean-based ulcers at cecum revealed active ileitis and colitis with multiple round shape organisms with capsule, which was compatible with Early diagnosis of gastrointestinal cryptococcosis in Crohn’s disease is difficult due to the lack of specific symptoms and sign or mimicking an exacerbation of Crohn’s disease. Seeking for other site of involvement in disseminated cryptococcosis including lung or central nervous system as well as detection of serum cryptococcal antigen would be helpful for early diagnosis and management. Cryptococcosis is a significant opportunistic infection in non-HIV immunocompromised individuals, including those with solid organ transplantation, malignancy, diabetes, rheumatologic disease, or chronic liver disease –3. Clini2 ulcer that was covered with white exudate at the cecum, and multiple polypoid masses at the ileocecal (IC) valve. The histopathologic findings revealed a chronic ulcer with marked active inflammation, a hyperplastic polyp with chronic ileitis, and no viral inclusion, dysplasia, or malignancy. Eight weeks ago, she started complaining of intermittent cramping at the RLQ of her abdomen. The dose of azathioprine was increased to 175 mg/day for presumed exacerbation of CD; however, her pain worsened, and she eventually developed a low-grade fever and watery diarrhea for 2 weeks before admission. She denied having any pulmonary symptoms. On examination, her body temperature was 38.6 °C and she had abdominal tenderness and a palpable mass at the RLQ of her abdomen. All other examination findings were unremarkable.A 64-year-old woman with a history of Crohn’s disease (CD) was referred to Siriraj Hospital located in Bangkok, Thailand in June 2017 due to worsening of abdominal pain and watery diarrhea for 2 weeks. She was diagnosed with CD in 2010 after she presented with abdominal pain and palpable mass at the right lower quadrant (RLQ) of her abdomen. She was treated successfully with prednisolone 40 mg/day, azathioprine 50 mg/day, and mesalazine 2400 mg/day. Her disease was then controlled with prednisolone 5 mg/day and the same dose of azathioprine and mesalazine. One year after her CD diagnosis, she developed severe abdominal pain and diarrhea. A colonoscopy was performed, which showed inflamed mucosa and a clean-based ulcer at the cecum. Biopsy revealed an intracytoplasmic inclusion body, which is consistent with cytomegalovirus (CMV) colitis. Her symptoms gradually subsided after 3 weeks treatment of intravenous ganciclovir 5 mg/kg twice daily. Two years after her CD diagnosis, she began experiencing recurrent abdominal pain. Abdominal computed tomography (CT) and colonoscopic findings revealed a large polypoid mass at the ascending colon with partial colonic obstruction that was suspected to be caused by active CD. She was managed conservatively and was discharged home with prednisolone 30 mg/day, azathioprine 150 mg/day, and mesalazine 2400 mg/day after making substantial improvement. Even though she remained stable for about 4 years after her most recent discharge, she still required prednisolone 5 mg/day, azathioprine 150 mg/day, and mesalazine 2400 mg/day to control her disease activity. Her last colonoscopy with biopsy, which was performed 6 months earlier to monitor her disease activity, revealed a shallow 1 cm3 with 90.5% neutrophils and 8.5% lymphocytes, and a hemoglobin level of 8.2 g/dl. Chest X-ray showed an oval-shaped cavitary lesion at the right lower lung  count of 2010 cells/mmThe final diagnosis was disseminated cryptococcosis due to gastrointestinal and presumed pulmonary involvement. Initial induction therapy with amphotericin B deoxycholate (AmBD) 0.7 mg/kg per day was given for 6 weeks, followed by consolidation therapy with fluconazole 800 mg/day for 8 weeks. After consolidation, maintenance treatment with fluconazole 200 mg/day was prescribed for up to 1 year. After diagnosis of disseminated cryptococcosis, the immunosuppressive agents were adjusted as follow: discontinued azathioprine, decreased the dose of prednisolone to 15 mg/day, and maintained mesalazine at 2400 mg/day. Her clinical status was gradually improved, and the colonoscopy that was performed at 10 weeks after the start of treatment showing marked improvement in the polypoid mass-like lesion at the cecum  are at increased risk for developing opportunistic infections due to factors that include receiving immunosuppressive and immunomodulatory therapy [particularly anti-tumor necrosis factor-α (anti-TNF-α) agents], severity of disease, and malnutrition –10. A syd Europe . More stThere is no direct relationship between CD and cryptococcosis. Among 9 reported cases of cryptococcosis in CD patients, 6 had pulmonary cryptococcosis –14, and According to the practice guidelines for the management of cryptococal diseases, non-CNS, non-severe pulmonary, localized cryptococcal infection can be treated with fluconazole alone. However, patients with disseminated disease (involvement at ≥2 noncontiguous sites) in non-HIV and non-transplant patients should be treated as CNS disease, as follows: amphotericin B deoxycholate (AmBD) 0.7–1 mg/kg per day for at least 6 weeks or AmBD 0.7 mg/kg per day plus flucytosine (100 mg/kg/day) for at least 4 weeks, followed by consolidation therapy with fluconazole  for 8 weeks, and maintenance therapy with fluconazole (200 mg/day) for a total duration of 6–12 months . Since fEarly diagnosis of GI cryptococcosis in CD is difficult due to the lack of specific symptoms and signs, and the fact that GI cryptococcosis tends to mimic exacerbation of CD. Investigation for other common sites of involvement in disseminated cryptococcosis (including lung and CNS), as well as detection of serum cryptococcal antigen, would help to facilitate early diagnosis, appropriate management, and more favorable outcomes.


Methylmalonic acidemia (MMA) is an autosomal recessive genetic disorder involving the metabolism of organic acids.cblA type MMA after hospitalization.Here, we report the case of a patient who developed acute metabolic crisis after vaccination and was diagnosed with MMAA gene that caused the disease in the patient but did not conform to Mendelian inheritance. Using chromosomal microarray analysis, maternal uniparental disomy (UPD) was found on chromosome 4q26‐q35.2 of the patient. The MMAA gene of the patient was inherited only from the mother and carried the same pathogenic variant on both alleles of chromosome 4. MMAA gene expression levels in whole blood were detected by real‐time PCR.Further examination revealed a homozygous pathogenic variant in the The nonsense pathogenic variant, NM_172250.2:c.742C>T (p.Gln248*), carried by the patient leads to a premature termination of transcription of the gene, thereby resulting in partial loss of protein function while retaining some others. Segmental UPD 4 is rare, and to our knowledge, has not been reported previously. We report a case that developed acute metabolic crisis after vaccination, and was diagnosed with cblA‐type of methylmalonic acidemia (MMA) after hospitalization. Using chromosomal microarray analysis, maternal uniparental disomy (UPD) was found in chromosome 4q26‐q35.2 of the patient. MMA due to UPD 4 had not been reported earlier. Segmental UPD 4 is rare, and MMA due to segmental UPD 4 has not been reported previously.Here, we report on a patient diagnosed with MMA. In order to identify the etiology, we employed exome sequencing and it was identified that the patient had a pathogenic homozygous pathogenic variant, NM_172250.2:c.742C>T (p.Gln248*), in the 22.1The patient was an 8‐month‐old girl with no report of abnormalities or delays in early motor milestones prior to vaccination. Approximately an hour after vaccination with the hepatitis B and meningococcal group A vaccines, the patient presented with vomiting, fever, cough, and opisthotonos. The patient's condition did not improve after upper respiratory tract treatment at a local hospital, and subsequently worsened after 10 days. She was lethargic and refused to eat, therefore, she was transferred to a superior hospital for treatment and was diagnosed with viral encephalitis and possible genetic metabolic disease. After treatment with oseltamivir, methylprednisolone, and immunoglobulin, fever reduced and her responses improved. Gas chromatography‐mass spectrometry (GS‐MS) and tandem mass spectrometry (MS/MS) performed on urine samples suggested the possibility of MMA. The patient was then transferred to our hospital for further treatment.Routine laboratory examinations included blood and urine biochemistry. No obvious abnormality was noted in liver function. Plasma homocysteine was in the normal range: 4.46 µmol/L . Urine GC‐MS confirmed elevated methylmalonic and methylcitrate, confirming the diagnosis of isolated MMA. The concentrations of blood propionyl carnitine (C3) and the C3 to acetylcarnitine (C2) ratio (C3/C2) were analyzed using MS/MS. C3 was 6.853 µmol/L  and C3/C2 was 0.85 µmol/L , suggesting propionic or MMA. A head MRI showed suspicious spotty and linear T2‐FLAIR hyperfocal lesions in the white matter of the brain. Both lateral ventricles and the third ventricle were dilated. The cerebral sulcus was widened.12 (1 mg/day), an intravenous infusion of l‐carnitine (100 mg kg day−1), a low protein diet (1–2 g kg day−1), and a special formula free from methionine, valine, and threonine. After treatment, she improved, and long‐term oral administration of l‐carnitine (1 g/day) and Cyanocobalamin (0.5 mg/day), along with maintenance of the low protein diet and formula continued. At the 6‐month follow‐up, the patient was in stable condition with no obvious abnormalities in growth and development. And C3, C3/C2 level decreased and only a small amount of methylmalonic acid was excreted in urine organic acid.Then she was treated with an intramuscular injection of vitamin B2.2https://decipher.sanger.ac.uk), DGV (http://dgv.tcag.ca/dgv/app/home), ClinVar (http://www.ncbi.nlm.nih.gov/clinvar/), and a local database. After obtaining the MMAA gene pathogenic variant, Sanger sequencing was performed to verify the pathogenic locus of the single gene. GenBank reference sequence and version number:MMAA .This study was performed in accordance with the principles of the Declaration of Helsinki and was approved by the Ethics Committee of the Sixth Affiliated Hospital, Sun Yat‐sen University .Written informed consent was obtained from the parents of the patient. After obtaining written informed consent, peripheral blood was drawn from the patient and her parents using the Agilent capture kit and Illumina SBS kit. High‐throughput sequencing was performed on the Illumina HiSeq platform and the Clinical Sequence Analyzer (CSA) from WuXi NextCODE was used for quantitative analysis. The Sentieon software suite was mainly used for secondary analysis of the sequencing data. Sequencing data were compared to the UCSC hg19 reference genome by Sentieon BWA. Variants were annotated with a process developed using WuXi NextCODE, and each variant was evaluated using the CSA, developed and validated by WuXi NextCODE. Sequencing results were compared with DECIPHER  was used to detect the mRNA expression level of MMAA gene. The pre‐sequence and post‐sequence of the variant site c.742C>T were amplified respectively, and the primer sequences respectively were: (a) Sequence name: c.742C>T, forward primer: 5’‐AGGAGTTAGCCCAGGTGCTTC‐3’, reverse primer: 5’‐CCAGACAATCCTACTCGAAATGCT‐3’; (b) Sequence Name: c.742C>T, forward primer: 5’‐AGGAGATGAGCTGCAGGGTATC‐3’, reverse primer: 5’‐TATCCTTCGAGCTGGCACAA‐3’. Forty replicates were used and all amplified fragments were 103 bp. The relative expression of the sample was calculated using the 2-ΔΔCT method, where ΔTC equals the target gene TC‐internal reference gene TC, and ΔΔTC equals ΔTC of patient group − ΔTC of the control group/maximum value. TC is the cycle number that correlates to the fluorescence signal as it reaches the set domain value.3MMAA gene was found in the patient and mother, confirming cblA type MMA. In the sequencing data of the family, the patient was diagnosed with a homozygous pathogenic variant, the mother with a heterozygous pathogenic variant, and the father had no pathogenic variants . The molecular karyotype was 46,XX,arr[hg19] 4q26q35.2 x2 hmz  pathogenic variant in the MMAA gene in whole blood samples were detected by real‐time PCR. The results showed that the relative expression of the pre‐variant sequence and sequences after the pathogenic variant of the MMAA gene was 1.000 in the homozygous sample. The relative expression was 1.1865 and 1.1674 in the pre‐variant and post variant sequence, respectively, in the maternal mutant. The relative expression was 1.0669 and 1.5052 in the pre‐variant and post variant sequence, respectively, in wild‐type samples  homozygous pathogenic variant of the MMAA gene on segmental UPD 4 with the molecular karyotype of 46,XX,arr[hg19]4q26q35.2 x2 hmz. The nonsense pathogenic variant, NM_172250.2:c.742C>T (p.Gln248*), carried by the patient is associated with premature termination of the gene, synergistically the decrease trend in gene expression, could lead to partial lost function and decreased expression of the protein. When the human body is in stress, caused by fever, improper diet, fatigue, drugs, and hunger, its metabolic homeostasis may be destroyed causing acute onset of the disease. In general, the clinical phenotype manifested in patients is relatively mild, and prognosis is mostly good, as shown in previous studies  and heterodisomy (inheritance of a pair of homologous chromosomes from one parent). In many UPD cases, chromosomes of both isodisomy and heterodisomy may be present  homozygous pathogenic variant of the MMAA gene. The chromosome molecular karyotype is 46,XX,arr[hg19]4q26q35.2 x2 hmz. The effects of this variant need to be further studied in future. We suggest that SNP or microsatellite genotyping would be useful in diagnose patients with UPD.This report suggests that UPD may be a cause of autosomal recessive disorders, especially when patients have a rare homozygous pathogenic variant. Here, we have reported the first case of All authors have no conflicting financial interests to declare.http://www.wanfangdata.com.cn/details/detail.do?_type=perio%26xml:id=syeklczz201621010 provided molecular analysis other than genetic sequence analysis and prepared the manuscript. Yao Cai provided the clinical follow‐up. Hu http://www.wanfangdata.com.cn/details/detail.do?_type=perio%26xml:id=syeklczz201621010 and Hui Xiong edited the paper.Fei Ma and Congcong Shi performed analysis of the sequence data. Min


We aimed to assess longitudinal changes of B‐type natriuretic peptide (BNP) in aortic stenosis (AS) patients treated by transcatheter aortic valve replacement (TAVR).From our TAVR database, we identified 193 consecutive patients with severe symptomatic AS who underwent TAVR and were prospectively followed using serial BNP levels and echocardiography. Patients were divided into subgroups according to type of left ventricular (LV) remodeling as having normal LV mass and relative wall thickness, or showing concentric remodeling (CR), concentric hypertrophy (CH), and eccentric hypertrophy (EH).P < 0.01) and a greater subsequent decrease (P < 0.001). During the median follow‐up of 1331 days (interquartile range: 632‐1678), 119 (62%) patients died. BNP showed a time‐dependent association with all‐cause mortality both in a univariable , and in a multivariable model with Society of Thoracic Surgeons score and baseline BNP forced into the analysis . Elevated BNP was associated with a larger LV end‐diastolic volume index (P < 0.001) and shorter 6‐minute walk test distance (P = 0.013) throughout follow‐up.At baseline, 30 patients (16%) had EH, 115 (60%) had CH, 37 (19%) had CR, and 11 (6%) had normal LV geometry. After TAVR, BNP decreased in the first 30 days, with further improvement during follow‐up. Patients with EH had higher BNP at baseline (In patients with AS, BNP was associated with LV remodeling phenotypes and functional status before and after TAVR. Elevated BNP levels were associated with poor prognosis. We also assessed the determinants of BNP change and its association with specific LV remodeling patterns.22.1We reviewed our series of 237 consecutive high risk or surgically inoperable patients with severe AS who underwent TAVR through transfemoral access with the Edwards SAPIEN valve at Cleveland Clinic between May 2006 and December 11, 2012.2.2Baseline and subsequent echocardiographic measurements were systematically reviewed and measured by an experienced reader according to the current guidelines blinded to clinical, other variables, and outcome.13 with LVH was defined as LVMi > 95 g/m2 in female and LVMi > 115 g/m2 in male. RWT was calculated as: LV remodeling was classified according to the combination of presence of LV hypertrophy (LVH) and relative wall thickness (RWT) values as concentric hypertrophy (CH), eccentric hypertrophy (EH), concentric remodeling (CR), and normal  and they were coded as zero meters. Data were obtained from research database.2.4t test, Mann‐Whitney test, One‐way anova, and Kruskal‐Wallis test to compare the data between two or more groups as appropriate. To assess changes in BNP after TAVR over time, and possible impact of LV remodeling pattern, we applied longitudinal data analysis using a mixed effect model with unstructured covariance for random and fixed effects,P‐value of <0.05 was considered statistically significant. All statistical analyses were performed with JMP 10.0 , SPSS 23.0 software , and R software version 3.2.2 .Continuous data are expressed as mean ± SD when normally distributed, or median (interquartile range). Categorical data are presented as an absolute number and percentages. BNP values were logarithmically transformed prior to analysis. We used the unpaired 3We identified 193 patients who fulfilled inclusion criteria Table . Six pat3.1P < 0.001)  to 262 (144‐461) pg/mL in the first 30 days in the entire group with further improvement during follow‐up . This indicates that decrease in BNP during follow‐up was not related to the informative censoring .In a next step, we divided patients into three groups according to their survival: patients who died during first 450 days, patients who died between 450 and 1200 days, and patients who survived for more than 1200 days. All three groups experienced a similar drop in BNP after TAVR (interaction between groups with time: 3.2P < 0.001). There was interaction between LV remodeling pattern and time, suggesting that patients with EH demonstrated a greater subsequent decrease in BNP levels after TAVR compared to those with CH, CR, and normal patterns . While significant drop in BNP was observed immediately after TAVR but continued through 6 months to 1 year after TAVR. When we compare the %BNP change according to the remodeling pattern after TAVR, EH group showed greatest %BNP decrease at 100 to 300 days after TAVR and continued with slow decrease afterwards   .To assess the association between functional class and BNP levels, we collected NYHA class and 6MWT results. At baseline, 49 patients were unable to walk because of medical conditions and 6MWT distance was considered as 0 m. After TAVR, among those 49 patients, 19 patients were able to walk. The mean distance of the 6MWT was 222 ± 116 m at baseline and improved to 264 ± 121 m at 30 days after TAVR, followed by further subsequent improvement to 271 ± 124 m at 6 months after TAVR (3.4P = 0.017). In a multivariable model, BNP as a time‐dependent covariate remained significant after adjusting for STS score , and baseline BNP levels , or both . Kaplan‐Meier curves show that patients with high BNP (>400 pg/mL) at 1 year post‐TAVR had poorer survival after TAVR (Log‐rank P = 0.03) , 119 (62%) patients died of any cause during follow‐up, with 48 patients (25%) dying during first 2 years. Of those 119 patients who died during follow‐up, 81 died because of cardiac cause. Furthermore, 74 patients died at 5 years of follow‐up, and the risk of cardiovascular‐related mortality at 5 years was 38%. To explore the impact of changing BNP levels during follow‐up, we performed uni‐ and multivariable Cox proportional hazards model with post‐TAVR BNP levels as a segmental time‐dependent covariate. In a univariable model, BNP as a time‐dependent covariate was a significant predictor of survival  Figure . These dP = 0.036). On the other hand, BNP was not significantly associated with survival in patients with EH .Moreover, when we repeated the analysis after excluding patients with EH, BNP was still a significant predictor of death with HR of 1.40  over the intervals of 0 to 100 days, 101 to 300 days, 301 to 600 days, 601 to 1000 days, 1001 to 1300 days, and > 1300 days. Error bars represent 95% confidence intervals. BNP, B‐type natriuretic peptide; CH, concentric hypertrophy; CR, concentric remodeling; EH, eccentric hypertrophy.Click here for additional data file.FIGURE S3 Association between BNP and GLS during follow‐up. BNP and GLS decreased in parallel throughout follow‐up (P < 0.001 for the association). Ln BNP, natural logarithm of B‐type natriuretic peptide concentration; GLS, global longitudinal strain; TAVR, transcatheter aortic valve replacement.Click here for additional data file.FIGURE S4 Kaplan‐Meier curves according to BNP level at 1‐year after TAVR. Survival curves showed patients with high plasma BNP level at 1 year after TAVR associated with higher mortality (Log‐rank P < 0.001). BNP, B‐type natriuretic peptide; TAVR, transcatheter aortic valve replacement.Click here for additional data file.TABLE S1 Patient clinical characteristicsClick here for additional data file.


Nepenthes pitcher plants. We genotyped wild populations of flowering males and females of three Nepenthes taxa using ddRAD‐seq and sequenced a male inflorescence transcriptome. We developed a statistical tool (privacy rarefaction) to distinguish true sex specificity from stochastic noise in read coverage of sequencing data from wild populations and identified male‐specific loci and XY‐patterned single nucleotide polymorphsims (SNPs) in all three Nepenthes taxa, suggesting the presence of homomorphic XY sex chromosomes. The male‐specific region of the Y chromosome showed little conservation among the three taxa, except for the essential pollen development gene DYT1 that was confirmed as male specific by PCR in additional Nepenthes taxa. Hence, dioecy and part of the male‐specific region of the Nepenthes Y‐chromosomes likely have a single evolutionary origin.Species with separate sexes (dioecy) are a minority among flowering plants, but dioecy has evolved multiple times independently in their history. The sex‐determination system and sex‐linked genomic regions are currently identified in a limited number of dioecious plants only. Here, we study the sex‐determination system in a genus of dioecious plants that lack heteromorphic sex chromosomes and are not amenable to controlled breeding:  Nepenthes. We surveyed wild populations of three species by genotyping a large number of loci throughout their genomes. Because such data are noisy, we propose a solution to the common problem of distinguishing true signal from noise in presence–absence data by generating null distributions through permutations of the observed data. We discovered loci that occur only in males and reveal an XY sex chromosome system. One gene on the Nepenthes Y chromosome is particularly interesting, because it is presumably essential for pollen development and present only in males, and thus can be used to diagnose the sex of nonflowering plantsOne of the most striking polymorphisms observed in organismal populations is the existence of male and female individuals. In contrast to animals, where this condition is common, plants are usually functional hermaphrodites. Some plants, however, are dioecious, that is, individuals are either of male or female sex. Dioecy has evolved hundreds of times independently in plants, which offers the potential for comparative studies of sex chromosome evolution and for investigating the genetic basis of transitions between hermaphroditism and dioecy , and female heterogamety when the fully sex‐linked region is transmitted via females . The sex‐specific, fully nonrecombining regions  show a number of special properties. First, an MSY may contain sequences that are absent from the X, and thus male specific . As X and Y chromosomes are thought to evolve from a pair of autosomes, the gain of male‐specific sequences can be explained by several mechanisms, such as the rise of a new male‐determining mutation, or the translocation of a male‐determining cassette  includes at least 160 species of perennial vines and shrubs occurring mostly in Southeast Asia  Are there sex‐linked loci in Nepenthes? (2) Are the same sex‐linked loci shared among different Nepenthes species? (3) Which expressed genes are sex linked? We discovered fully male‐specific and XY‐patterned loci and developed a molecular sexing assay for Nepenthes. The identified markers include two candidate sex‐determination genes, and these suggest that part of the Y chromosome is ancestral in this genus.Here, we investigated the previously undescribed sex‐determination system of multiple Nepenthes were sampled in Brunei Darussalam (Borneo), Singapore, and the Seychelles. Fresh leaf material was stored in a nucleic acid preserving buffer  were sequenced on an Illumina HiSeq 2500. Bioinformatic data filtering, de novo assembly of reference contigs , read mapping, genotype calling, and quality filtering followed a modified dDocent pipeline . Further details, including a performance analysis with simulations, are presented in Texts We distinguished between sex‐linked loci showing sex‐specificity, meaning without homology between the two sex chromosomes , and loci that are present on both sex chromosomes but whose allele frequencies diverged between the sex chromosomes, called ZW‐ or XY‐patterned variants  was calculated exclusively for SNPs from different contigs using PLINK. The same contrasts were made for nucleotide diversity π, which was averaged per contig for all SNP‐containing contigs (including singleton SNPs) in VCFtools version 0.1.15 , that is, the nonrandom association of alleles at separate loci, differed in male samples between sex‐linked regions and the genomic background . Stringent quality filters were applied: singleton SNPs were removed, SNPs in male‐specific contigs were excluded if any heterozygous genotypes were called in the males , and for non‐sex‐linked contigs any excessively heterozygous SNPs (Hardy–Weinberg test with significance level 5%) were excluded. Excessively heterozygous SNPs were retained for XY‐patterned contigs. LD . PFAM domains were detected using hmmer 3.1b1  and the nonredundant protein collection . Transposable elements (TEs) were detected using RepeatMasker 4.0.6 , taxonomic overlap, and the quality of annotation of matching transcripts. PCR primers were designed in Geneious R6 , and tested according to the protocol described in Text S. latifolia GBS data using privacy rarefaction. When small numbers of individuals of each sex were analyzed, the procedure yielded similar numbers of male‐ and female‐specific candidates , 0.11% for N. rafflesiana s.l. , and 0.06% for N. gracilis , versus 1.52% for S. latifolia .Qualitatively consistent signatures of male‐specific contigs were detected independently in .l. Fig. . The proN. pervillei and N. rafflesiana s.l., as well as in S. latifolia, we also detected XY‐patterned SNPs, but not in N. gracilis, while none of the species yielded any ZW‐patterned SNPs. The XY‐patterned SNPs for S. latifolia recovered 289 contigs already known to be sex linked in that species, which, however, represent only a small fraction of the known and theoretically expected S. latifolia sex‐linked sequences : the estimates were, respectively, 0.25% and 0.017% for N. pervillei  and N. rafflesiana s.l. .In N. pervillei showed perfect LD . Likewise, the male‐specific contigs of N. rafflesiana s.l. (containing 82 SNP pairs) had an r2 that was on average c. 0.4 above the genomic mean (P < 10−5); the median value was complete LD (r2 = 1). Among contigs with XY‐patterned SNPs, r2 was 0.15 higher than the background mean in N. pervillei , but was no different from the mean in N. rafflesiana s.l. . Our observation of some low LD values between sex‐specific contigs is likely due to the discreteness of allele frequency estimates from small sample sizes, and the allele frequency dependence of LD metrics, whose maximum possible value is frequently much less than 1.0 , but not for N. pervillei or N. gracilis . In contrast, mean π of males in contigs with XY‐patterned SNPs was higher than the genomic mean for both N. pervillei  contigs tended to be lower than the genomic background in all three taxa Fig. , consist. P < 10−, but not. P < 10−, but notN. gracilis and N. rafflesiana s.l. No sex‐specific candidates were shared between N. pervillei and the other species  to the same inflorescence transcript containing a DUF4283 .Six candidate sex‐specific contigs were found at privacy rarefaction stringency level ≥5 that were shared between es Table . There wN. pervillei aligned to the transcript of a bHLH transcription factor, and the best matches in all accessed databases were consistently to predicted orthologs of the Arabidopsis gene DYSFUNCTIONAL TAPETUM1 (DYT1). A further XY‐patterned contig of N. pervillei matched a transcript annotated as A. thalianaSEPALLATA‐1 (SEP1), which aligned to the predicted 3′‐UTR of the putative N. pervillei SEP1‐ortholog, and contained two SNPs that were both homozygous in 95% of females and heterozygous in 96% of males. No estimate of SEP1 X–Y divergence was possible because the male inflorescence transcriptome reads were not heterozygous. In N. gracilis, a male‐specific contig matched a long transcript similar to a mitochondrial NADH‐ubiquinone oxidoreductase from Beta vulgaris (Swiss‐Prot). This finding was unexpected and may represent either an unspecific match of the short (96 bp) contig to the inflorescence transcript, or else a cyto‐nuclear transfer to the sex chromosomes. The occurrence of organellar genes on plant sex chromosomes has been documented in other species . Nine out of 13 sex‐linked TE‐transcripts annotated as gypsy‐like retrotransposons, a significant overrepresentation relative to nonsex‐linked TE transcripts .All other candidate loci either included traces of TEs, or had no known sequence motifs Table . In partN. rafflesiana s.l. but could not be annotated. Four close transcript “isoforms” . These two transcripts appear to be close isoforms (putative intron presence–absence), and both annotated as DYT1 (see above).Complementary to the sex‐specificity scan on the ddRAD‐de novo reference, we repeated privacy rarefaction by directly mapping the ddRAD reads to the male inflorescence transcriptome to find further annotated sex‐linked genes. This identified seven transcripts Table  that mapDYT1‐ortholog is male specific in a broad range of Nepenthes species. A single PCR product of approximately 290 bp length was observed exclusively and consistently in phenotypically sexed male Nepenthes but never in females  million years ago but followed the split between Nepenthaceae and hermaphroditic Droseraceae at least 44.2 million years ago  have lost sequence similarity in the former but not in the latter species.During the radiation of lei Fig. . Male‐spNepenthes, most sex‐linked genomic regions detected by our approach were noncoding sequences and TEs and only a few genes with putative developmental functions were identified. Of these, a Nepenthes homolog of DYT1 appears to be located in the MSY of all Nepenthes species. DYT1 is essential for tapetum development and thus pollen fertility in A. thaliana . The sex chromosomes are homomorphic with a small Y‐specific region, which has a relatively old core that is shared between distinct species. The nonrecombining region is enriched for noncoding sequences and TEs, but also contains several expressed genes with putative developmental functions.This study reports the discovery of an XY sex‐determination system in dioecious pitcher plants (Associate Editor: S. WrightText S1. Preliminary molecular sexing assay for Nepenthes rafflesiana s.l.Text S2. A molecular sexing assay for the genus Nepenthes.Text S3. Analyses of sex‐linkage in Silene latifolia.Text S4. Privacy rarefaction.Text S5. Performance analysis of privacy rarefaction on simulated RAD data.Text S6. Male inflorescence transcriptome of N. khasiana.Text S7. Phylogenetic dating of Nepenthes.Click here for additional data file.Table S8. Result tables of privacy rarefaction for three Nepenthes and Silene.Click here for additional data file.Table S9. Result tables of association tests for SNPs with sex and heterozygosities.Click here for additional data file.Table S10. Summary table of sex‐linked loci including overlap between species and annotations for RAD‐tag and transcriptome references.Click here for additional data file.Table S11. Data table for S. latifolia sex‐linkage comparison of our results to previous studies.Click here for additional data file.


Solidago canadensis L., a widespread wild hexaploid invader with localized cultivated diploid populations. Our analysis of the leaf transcriptome revealed 116,801 unigenes, of which 12,897 unigenes displayed significant differences in expression levels. A substantial number of these differentially expressed unigenes (DEUs) were significantly associated with the biosynthesis of secondary metabolites, carbohydrate metabolism, lipid metabolism, and environmental adaptation pathways. Gene Ontology term enrichment‐based categorization of DEU‐functions was consistent with this observation, as terms related to single‐organism, cellular, and metabolic processes including catalytic, binding, transporter, and enzyme regulator activity were over‐represented. Concomitantly, 186 miRNAs belonging to 44 miRNA families were identified in the same leaf tissues, with 59 miRNAs being differentially expressed. Furthermore, we discovered 83 miRNA‐target interacting pairs that were oppositely regulated, and a meticulous study of these targets depicted that several unigenes encoding transcription factors, DNA methyltransferase, and leucine‐rich repeat receptor‐like kinases involved in the stress response were greatly influenced. Collectively, these transcriptional and epigenetic data provide new insights into miRNA‐mediated gene expression regulatory mechanisms that may operate in hexaploid cytotypes to favor successful invasion.Dissecting complex connections between cytogenetic traits (ploidy levels) and plant invasiveness has emerged as a popular research subject in the field of invasion biology. Although recent work suggests that polyploids are more likely to be invasive than their corresponding diploids, the molecular basis underlying the successful invasion of polyploids remains largely unexplored. To this end, we adopted an RNA‐seq and sRNA‐seq approach to describe how polyploids mediate invasiveness differences in two contrasting cytotypes of  To this end, we adopted an RNA‐seq and sRNA‐seq approach to investigate the divergences of gene and miRNA expression between diploid and hexaploid cytotypes of S. canadensis. Furthermore, we constructed a co‐expression network of differentially expressed genes and miRNAs to shed light on the regulatory action of miRNAs. Taken together, our work provides new insights into miRNA‐mediated gene expression regulatory mechanisms that may be useful to explain the successful invasion of hexaploid cytotypes.Herein, we evaluate the effect of polyploids on gene and miRNA expression while also considering the potential roles of miRNA‐mediated gene expression regulation in driving differences in invasiveness between diploid and hexaploid cytotypes of 22.1Solidago canadensis (Asteraceae), a perennial weed native to North America  and hexaploid cytotypes (HS 3) were selected as the experimental materials for comparison to investigate gene and miRNA expression profiling in this work Figure a,b. Leav2.4S. canadensis were pooled together and further clustered to generate “All‐Unigene” for subsequent assembly evaluation, unigene annotation, and expression analysis. The “All‐Unigene” sequences were aligned by BLASTx to a series of protein databases to gain unigene annotation. See Supporting Information Appendix Raw reads were filtered by removing those reads that contained adaptors, unknown nucleotides (more than 5%), and low‐quality bases . De novo assembly of all processed reads was performed by Trinity  setting of ≤0.05. DEUs were further assigned KO (KEGG Orthology) numbers using the KEGG database, and their enrichment was performed as mentioned for GO.The Bowtie2 program  available in the miRBase 22.0 database using the BLASTn program to identify the known miRNAs. Transcripts per million was used to normalize the read count of each identified miRNA based on the following formula: Normalized expression = Actual miRNA count × 106/Total count of clean reads. After normalization, differential expression analysis of miRNA was performed using DEGseq as described by Wang, Feng, Wang, Wang, and Zhang  with default parameters. Additionally, the expression level of predicted miRNA targets was taken from the inventory of assembled “All‐Unigene.” GO terms were also evaluated using a similar method.To predict the potential genes targeted by miRNAs, the Targetfinder ; and (b) miRNA‐target interaction pairs that were negatively correlated in expression levels. qRT‐PCR was implemented in triplicate on an ABI Step One Plus Real‐Time PCR System (Applied Biosystems) with unigene‐ and miRNA‐specific sense and anti‐sense primer (Table GAPDH (Unigene25510_All) was co‐amplified to normalize the expression values of unigenes and miRNAs in each sample using the double‐standard curve method.Eighteen promising candidate unigenes and six miRNAs observed to be differentially expressed were chosen for qRT‐PCR to validate the reliability of RNA‐seq and sRNA‐seq results with the following selection criteria: (a) up‐ or down‐regulated unigenes discussed in this paper  raw reads were produced and subjected to Seq‐QC collating, which resulted in 289.45 M (86%) clean reads with Q20 values ranging from 98.88% to 98.94%. Then, clean reads from six libraries were de novo assembled separately into unigenes by Trinity. These assembled unigenes were pooled together and further clustered into a reference transcriptome  with an average length of 1,056 bp, a N50 value of 1,610 bp, and a GC content of 39.20%  down‐regulated in hexaploid cytotypes  were over‐represented, followed by MYB‐related (270), AP2‐EREBP (211), bHLH (134), and WRKY family members (120). In addition, we discovered 381 TF‐encoding genes that were differentially expressed, with the majority being up‐regulated in hexaploid cytotypes. Notably, among these differentially expressed TF‐encoding genes, almost all the members of the bHLH group were found to be up‐regulated (12/15 genes) in hexaploid cytotypes. In addition, MYB (32/53), MYB‐related (31/47), ARF (10/16) and Trihelix (9/12) members exhibited a similar trend. However, the majority of TF‐encoding genes belonging to the WRKY (18/26) and NAC (13/17) families were down‐regulated in hexaploid cytotypes  unigenes from the 12,897 DEUs were successfully classified into three major functional categories: biological process , molecular function , and cellular component . Then, the three major categories were further assigned to 50 terms  single‐end raw reads were produced and subjected to Seq‐QC collating, which resulted in 166.6 M (92.6%) clean reads with lengths ranging from 18 to 30 nt Table , Oryza sS. canadensis and found that the identified families included a changing count of miRNA members , −3.15 (q = 1.40e−04) and −2.45 (q = 6.35e−11), respectively, and sca‐miR166p, sca‐miR528, and sca‐miR396a were markedly up‐regulated with log2fold‐change values of 5.16 (q = 7.39e−12), 5.13 (q = 9.36e−07), and 5.04 (q = 5.30e−11), respectively. Notably, for the miR160 and miR169 family, sca‐miR160e, sca‐miR169b, sca‐miR169e, sca‐miR169f, sca‐miR169g, and sca‐miR169h were up‐regulated specifically in hexaploid cytotypes, while sca‐miR160b and sca‐miR169d were up‐regulated specifically in diploid cytotypes. These observations suggested that different members from the same miRNA family had different regulatory modes, probably associated with the cooperative and redundant regulation activity of miRNAs. qRT‐PCR analysis performed for six DEMs confirmed the miRNA changes detected by sRNA‐seq   miR5139a had the highest target abundance (179), and (b) the genes such as CL10163.Contig1_All, CL13112.Contig1_All, and Unigene2861_All had the highest miRNA abundance (4). To understand in depth the group of unigenes targeted by DEMs, GO functional analysis of the predicted targets was carried out. Under the biological process category of GO classification, unigenes involved in terms such as “cellular process,” “metabolic process,” “single‐organism process,” “response to stimulus,” etc. were abundantly enriched as the targets of DEMs. Under the molecular function category, unigenes displaying “catalytic activity,” “binding,” “transporter activity,” etc. were targeted by miRNAs. Moreover, “cell,” “membrane,” “organelle,” etc. related unigenes were discovered to be clustered into the cellular component category as targets , ubiquitin‐protein ligase (CL2235.Contig13_All), U‐box domain‐containing protein (CL2207.Contig4_All), and F‐box protein (Unigene1223_All), that were predicted to be targets of sca‐miR396d, sca‐miR444a, sca‐miR393d, and sca‐miR5139a, suggesting that these unigenes may be subjected to miRNA‐mediated DNA methylation and ubiquitination; and (d) two dirigent protein genes (CL6884.Contig1_All and CL6884.Contig3_All) that were predicted to be targets of sca‐miR169d. This target was an unspecific oxidizing enzyme gene for radical formation that functions in lignan biosynthesis, which was previously reported to be an integral regulator of plant secondary metabolism  and diploid Aegilops tauschii (DD)  was markedly greater than those in its two diploid ancestors, G. raimondii (DD), and G. arboreum (AA)  and Brassica nigra (BB). In the present work, the number and expression levels of miRNAs in hexaploid cytotypes were greater than those in their diploids, which was consistent with the findings of the above‐mentioned studies. These results suggest that an increase in ploidy was generally coupled with an obvious increase in the percentages and expression levels of miRNAs.Likewise, the relative amount of miRNAs was higher in a derived hexaploid wheat (BBAADD) than in the parental tetraploid 4.2How does hexaploid cytotypes regulate the differential expression of unigenes? Several mechanisms could be associated with this regulation. miRNAs work as regulators for controlling target‐gene expression, thereby affecting a variety of aspects of phenotype, growth, development, and stress response  annotated as GIF were found to be differentially expressed and may form functional complexes potentially implicated in leaf size and growth. Furthermore, other transcription factors, such as the TCP transcription factor , were also identified as regulators of leaf size. Except for TFs, regulatory proteins act as important regulators of leaf size and growth by influencing cell proliferation. EBP1, an ortholog of ErbB3‐binding protein from humans, regulates leaf size and growth by cell proliferation. Some studies highlight that the expression of EBP1 correlates with plant organ size, growth, and stress tolerance  to affect plant organ size and growth , a gene deeply induced by auxin, participate in organ size regulation. Wang, Zhou, et al. (Brassica rapa) BrARGOS gene in Arabidopsis elevates the size of plant organs. In this work, an ortholog of BrARGOS, CL15040.Contig2_All was detected, and the up‐regulated expression may have similar functions in the organ giantism observed in hexaploid cytotypes. Auxin Response Factor (ARF), a transcription factor, functions in plant size, growth, and stress adaptation by transcriptionally activating and repressing the expression of auxin response genes ” was found to be the most significantly enriched pathway with a Q value far below 0.05 in the pathway enrichment analysis of DEUs in the present work. Taking the above into account, we propose that polyploids are more likely to remodel the transcriptome and metabolome in hexaploid cytotypes, resulting in ploidy‐specific metabolic adaptation. Moreover, a marked number of DEUs encoding enzymes related to plant metabolism were observed, which further supports this plausible explanation. The synthesis of secondary metabolites primarily contains the oxidation, reduction, and cyclization steps, in which unigenes encoding enzymes of cytochrome P450 (CYPs) and uridine diphosphate glucuronosyl transferases (UGTs) play crucial roles in catalyzing these reactions , CYP76B , and CYP71 , which respectively participated in the biosynthesis of isoflavonoids, flavonoids, and sesquiterpenoids and triterpenoids, which may act as defensive compounds that protect against oxidative damage under abiotic stress. Furthermore, a great deal of UGT genes that participated in flavonoid biosynthesis, such as UGT73, UGT74, UGT76, UGT83, UGT85, and UGT89, were also identified (Table S. canadensis.Furthermore, polyploids can also profoundly affect plant metabolism qualitatively and quantitatively, furnishing the chance for increased metabolic activity through transcriptional divergence, which eventually results in alterations in the levels of secondary metabolites (Fasano et al., S. canadensis have been investigated in the current work, and we have also further inferred ploidy‐related regulation of DNA methylation as an additional modulatory event that occurs to modulate transcriptome reprogramming to drive invasion success. Furthermore, a model for depicting the events involved in ploidy alteration in S. canadensis is summarized in Figure In conclusion, important candidate unigenes and miRNA regulators that contributed to the successful invasion of hexaploid cytotypes of None declared.C.C.X. collected plants, performed greenhouse experiment, analyzed the collected data, and wrote this manuscript; Y.M.G. participated in data analysis; J.B.W. conceived the research and contributed to data interpretation and revisions of this manuscript. Click here for additional data file. Click here for additional data file. Click here for additional data file. Click here for additional data file.