Freundlich isotherm well described the adsorption of VOCs on the considered materials. According to the obtained results, a multilayer, physical, and cooperative adsorption process was hypothesized, particularly evident when the VOCs’ concentrations are high. This was confirmed by the high adsorption efficiency percentages (E% > 80%) of VOCs from a real polluted matrix (urban solid waste leachate), containing high concentrations of total organic content.The aim of this work was to study the potential of food waste materials  as low-cost adsorbents for the removal of aliphatic and aromatic volatile organic compounds (VOCs) from wastewater. The ability of examined food waste materials to adsorb VOCs from synthetic multi-component standard solutions was evaluated and the examined food waste materials showed high removal efficiency. Performances of coffee waste, grape waste, and lemon peel were detailed by using Trichloroethylene and p-Xylene in mono-component standard solutions. The adsorption capacity of the three selected food wastes was determined by using linear Langmuir and Freundlich isotherm models. Two errors functions, average percentage error (APE) and the chi-square test (χ Volatile organic compounds (VOCs), aliphatic and aromatic, are a wide class of organic pollutants extensively released in the environment from a variety of anthropogenic sources such as fuel storage and transport, industrial operations , manufacBTEXs, which have been found mainly in sites contaminated by petrol, gasoline, and petrochemical products ,14, are Several methods for wastewater decontamination from organic pollutants are reported ,20,21,22Among the biosorbents, food waste materials appeared to be efficient for the removal of inorganic pollutants from wastewaters ,39,40,41Since the high danger of volatile organic compounds for humans and the environment along with the scarce amount of studies regarding the biosorption of VOCs by food waste materials, the evaluation of the performances of these materials as VOCs adsorbents constitutes an interesting issue that still should be investigated.In this study, eight food waste materials  were evaluated as potential low-cost biosorbents for the removal of VOCs from wastewater. With this aim, their adsorption efficiency and capacity of aliphatic/aromatic VOCs from multi-component and mono-component synthetic solutions and from a real polluted matrix were explored.Food waste powders (n = 8) were employed as low-cost adsorbents: banana peel, potato peel, apple peel, lemon peel, coffee waste, decaf coffee waste, grape waste and carob peel . PreparaFirst, removal efficiency of organic pollutants was explored by exposing each biosorbent to a synthetic multi-standard solution containing both aliphatic and aromatic VOCs . In these preliminary trials, 100 mg of each adsorbent was exposed to 10 mL multi-standard solutions at 20 µg/L by using 20 mL glass vials, hermetically sealed. Multi-standard solutions were obtained by diluting 2 mg/L multi-component solution, prepared by mixing proper aliquots of aliphatic and aromatic standard stock solutions  in methanol.The adsorption kinetic was assessed by keeping both the mass of each waste material (100 mg) and the VOCs concentration (20 μg/L) constant, while increasing exposure time from 1 h to 24 h .eC), which is the amount of adsorbate remaining in solution after adsorption processes. Multi-component standard solutions without biosorbent were always treated together with each sample set, to trace and control possible VOCs’ concentration variability due to loss by evaporation during treatment; VOCs’ concentration of these solutions was used as initial concentration (0C).All the samples were analysed to determine the equilibrium concentration  of this solution was measured by a TOC analyzer .The performances of the food waste materials as biosorbents in a real polluted matrix were tested by exposing 100 mg of each food waste sorbent to 10 mL of an urban solid waste leachate. The same procedure described for the synthetic multi-standard solutions was applied to perform the exposure experiments and to determine The pH of both multi-standard solutions and real polluted matrix, with and without biosorbents, was measured using a pH meter .2/g. These values were close to the detection limit of the technique and the high uncertainties (about 0.05 m2/g) did not allow a reliable comparison among materials.According to results obtained from the just described preliminary trials, two selected target VOCs were chosen for deepening the study of adsorption behavior in synthetic mono-component solutions. Trichloroethylene and p-Xylene were representatives of aliphatic and aromatic classes, respectively. In these trials, adsorption capacity was calculated for three selected food wastes by varying VOCs’ concentration in the range 25–2000 µg/L and by keeping constant at 100 mg the adsorbent amount. Coffee waste, grape waste, and lemon peel were chosen for this experimental procedure as biosorbents, with reference to a recent paper in which all the considered food waste materials were characterized in detail by scanning electronic microscopy (SEM), Fourier transform infrared spectroscopy (FTIR), and principal component analysis (PCA) . SpecifiAromatic and aliphatic VOCs considered in this work were identified by comparison with the analytes registered in NIST Mass Spectral Library (1.6 NIST MS Software Version 2.3) of static headspace gas chromatography coupled to mass spectrometry (SHS-GC-MS) (probability of more than 70%) and quantified.Static headspace gas chromatography coupled to mass spectrometry  was used for all the VOCs’ analyses. After having chosen the exposure time, the glass vials containing the solutions (with or without the biosorbent) were maintained under stirring for 23 min at the constant temperature of 80 °C, then 1 µL of the vapor phase was injected in a capillary column  with a Hamilton syringe . Helium was used as carrier gas at the flow rate of 1 mL/min. The temperature was kept at 35 °C for 2 min, then it was increased to 50 °C at the rate of 6 °C per minute and, lastly, to 180 °C at the rate of 12 °C per minute. The total duration of the chromatographic run was 16 min.Chromatograms were registered in selective ion monitoring mode (SIM) by selecting specific mass to charge ratios (m/z) for each analyte. The conditions employed to mass spectrometry and selected fragments and retention times of the considered VOCs and are summarized in Final volume of each analysed sample was 10 mL and all the experiments were performed in triplicate. External standard solutions were used for quantification.E%) was calculated by the following equation:0C (µg/L) is the inlet VOCs’ concentration and eC (µg/L) is the equilibrium concentration after the adsorption process.Adsorption efficiency (eA) and in the absence (0A) of the biosorbent  was determined by following the procedure described in Wang et al. [V is the volume of solution (L) inside the vial and m is the amount (g) of food waste powder.Adsorption capacity g et al. , using aThe characterization of adsorption isotherms is considered one of the most appropriate methods for the assessment of the adsorbent capacity . AdsorpteC (mg/L) is the equilibrium concentration after adsorption process, eQ (mg/g) is the adsorption capacity, mq (mg/g) and LK (in L/mg) are sorption equilibrium constants. The constant LK is related to the free energy of adsorption and indicates affinity between the VOCs and the adsorbents [For Langmuir isotherms, sorbents .FK (mg/L) and n (dimensionless) are both constants indicative of sorption capacity and sorption intensity, respectively [n indicates the favorability of the adsorption process [n value represents good adsorption capacity, meaning VOCs’ adsorption is favorable [2R) is useful to predict which model best matches with the experimental data. In addition, the average percentage error (APE) and chi-square test (χ2) values were calculated in order to verify the validity of the isotherm models. APE and χ2, defined ad distribution functions, indicate the fit between the experimental and the predicted values of the adsorption capacity [2 were calculated as follows:e,expQ, e,preQ are experimental and predicted adsorption capacity, respectively; N is the number of observations in the experimental data.In Freundlich isotherm, ectively ,53. In p process ; a closeavorable . The evacapacity  and werecapacity : if dataPreliminary tests on synthetic multi-component solutions were performed in order to verify the general behavior of the considered food waste materials as VOCs’ biosorbents. First, the rate of the adsorption process was evaluated by exposing the materials to a synthetic mixture of aromatic and aliphatic VOCs for increasing times . As shown in E% > 50%); within the aliphatic compounds, Trichloroethylene, Tetrachloroethylene, and 1,1,2-Tetrachloroethane showed more than 40% of adsorption efficiency in presence of coffee and decaf coffee waste. Moreover, coffee waste was able to adsorb more than 90% of the Trans-1,2-Dichloroethene in multi-component solution. Apple peel seemed to be efficient for 1,1,2-Tetrachloroethane (E% > 50%) and for 1,2-Dichlorobenzene, 1,3-Dichlorobenzene and 1,4-Dichlorobenzene (E% > 30%). Lastly, it is worth noting that also grape waste adsorption efficiency is considerable for 1,2-Dichlorobenzene, 1,3-Dichlorobenzene and 1,4-Dichlorobenzene (E% > 40%) and 1,1,2-Tetrachloroethane (E% > 50%).Adsorption efficiency of each examined food waste material (100 mg) from a mixture of aliphatic and aromatic VOCs at 20 µg/L is reported in In general, looking at Considering the efficiency of the examined low-cost adsorbents for the removal of VOCs from synthetic solutions, other experimental tests were conducted to confirm the potential of the investigated food waste materials to adsorb organic compounds from a real polluted matrix. The chosen real polluted matrix had a very high total organic content (TOC = 7500 mg/L), and a spontaneous pH of 6.5. In E%) of Trichloroethylene was 43 ± 2 and 99 ± 12; lemon peel showed a Chlorobenzene E% of 10 ± 1 and 86 ± 12 and grape waste showed a m, p-Xylene E% of 34 ± 3 and 96 ± 5 in synthetic and real polluted matrix, respectively. This behavior may be due to the different VOCs’ concentration range in real polluted sample compared to the synthetic one, and seems to indicate a cooperative mechanism of adsorption, meaning an increase in the amount of pollutants adsorbed when initial VOCs’ concentration was increased. Coffee waste, decaf coffee waste, and grape waste were confirmed to be the most efficient biosorbents for the removal of both aliphatic and aromatic VOCs.The real polluted matrix (a leachate of urban solid waste) contained a wide variety of VOCs; only some of them were identified by comparison with NIST library (>70% probability). The estimated adsorption efficiencies for these compounds are reported in Adsorption processes were further investigated by studying the adsorption isotherms of selected biosorbents and VOCs. Two selected target VOCs were chosen for deepening the study of adsorption behavior in synthetic mono-component solutions. Trichloroethylene and p-Xylene were chosen as representative of aliphatic and aromatic classes, respectively. In these trials, VOCs’ concentration was varied in the range of 25–2000 µg/L, keeping constant at 100 mg the adsorbent amount. Adsorption capacity was calculated for three selected food waste biosorbents  according to the results obtained from a previous study, in which all the considered food waste materials were analysed by scanning electronic microscopy (SEM) and Fourier transform infrared spectroscopy (FTIR) . In thisSecond cluster included coffee and decaf coffee waste, apple, and banana peel, and was characterized by the highest amount of CH bonds of methyl and methylene groups of lipids; coffee waste was chosen as representative of this cluster. Third cluster included grape waste and carob peel and was characterized by the lowest amount of the identified functional groups; grape waste was chosen as representative of this cluster.eQ (µg/g) in presence of coffee waste, while grape waste and lemon peel were found to be less efficient. Adsorption capacity curves show a quite complex trend and seem to indicate that the adsorbent saturation was reached at the tested conditions. e value reached with Trichloroethylene and p-Xylene was 30 µg/g and 80 µg/g, respectively.Adsorption capacity of coffee waste, grape waste and lemon peel towards Trichloroethylene and p-Xylene is graphically represented in eQ recorded value of 10 µg/g and 20 µg/g, respectively, probably because of its less porous surface and the presence of polar functional groups [These results agree with those obtained from the adsorption experiments in multi-component standard solutions. Coffee waste was found to be the most efficient biosorbent for the removal of both Trichloroethylene and p-Xylene. The most plausible explanation comes from the chemical-physical properties of Trichloroethylene, p-Xylene, and coffee waste composition. FTIR spectra of coffee waste showed peaks of asymmetric and symmetric stretching of hydrophobic C-H bonds of methyl and methylene groups, attributed to the presence of lipids that are available in coffee samples in large amount ,55. Consl groups .Sorption equilibrium data for the two VOCs and the three food waste materials were fitted to the Langmuir and Freundlich isotherm models. Although many adsorption models were developed and applied in the literature ,59, the 2R ≥ 0.9) obtained by plotting logeQ versus logeC for both Trichloroethylene and p-Xylene indicate that these adsorption processes are well represented by the Freundlich model. Lemon peel showed a lower correlation, but it’s still significant.The constants obtained by applying the Freundlich isotherm model are summarized in n and FK of the Freundlich model indicate good adsorption intensity and capacity, respectively: n was close to unity for all the examined food waste adsorbents and for both Trichloroethylene and p-Xylene, indicating that VOCs adsorption is favorable [APE and 2χ in association with higher 2R generate a satisfactory fit of Freundlich model to the experimental data, even if they are little higher for lemon peel. To explain these contrasting results, the adsorption processes simulated by the two models are to be considered. Langmuir model represents only monolayer adsorption processes [n values reported in n < 1, the presence of an isotherm called “solvent-affinity isotherm” is suggested [Moreover, the values of avorable . Furtherrocesses  on the orocesses . Insteadrocesses ,62 in aqrocesses , in whicE%).Food waste materials, that are available in large amount and at very low costs, appeared to be promising biosorbents for both aliphatic and aromatic VOCs in aqueous solutions. In general, the aromatic VOCs were better adsorbed than the aliphatic by most of the considered materials. This is confirmed by the high percentages of adsorption efficiency (eQ) of three chosen food waste materials  with Trichloroethylene (chosen as representative of aromatic VOCs) and p-Xylene  were investigated; coffee waste resulted the most efficient sorbent for the removal of both considered VOCs. This is probably due to the presence of lipids in coffee samples on which Trichloroethylene and p-Xylene compounds are efficiently adsorbed, due to their lipophilic nature.Adsorption capacities (Adsorption equilibrium isotherms showed that coffee waste is the most efficient material for both p-Xylene and Trichloroethylene adsorption. In all the examined systems, the sorption processes followed much more closely the Freundlich than the Langmuir isotherm model. This indicates a physical adsorption mechanism thanks to London-Van der Waals forces between solute and sorbent, through a cooperative adsorption mechanism, in which the already adsorbed molecules on the sorbent’s surface facilitate the adsorption of other molecules. This kind of physical adsorption may be particularly beneficial in field applications, because it gives the adsorbents an opportunity to have great adsorption performance at high pollutant concentrations.Evaluation of food waste performances as biosorbents in field still needs further deepening, such as the effects of pH on removal capacity, the assessment of the effects of sorbents’ particle size, the investigation of repetitive adsorption desorption cycles for material reusability investigation, or even the comparison between other food waste materials’ adsorption capacity. Taking into account all the considerations, the results of this preliminary study suggest further investigations about the possible applicative use of these low-cost adsorbents for VOCs’ removal from wastewater.In this context, the efficient adsorption capacity of food waste materials at high VOCs’ concentration was preliminarily confirmed by the excellent adsorption efficiencies obtained in a real polluted matrix.


Herein, feasibility of supplementing wheat flour with Chinese yam powder (CYP) for noodle preparation was assessed. After supplementation with CYP, the alterations in chemical, texture, cooking, rheological, and microstructure attributes of noodles were observed. Due to higher protein and lower gluten, 20% of CYP promoted the stable network of gluten and starch particles. However, the excessive addition reduced the flexibility and the chewiness. The adverse changes were observed at 40% substitution level in texture profile analysis (TPA) and rheological parameters due to disrupted gluten–protein network which accelerated the exposure of starch particles. The CYP incorporation up to 20% showed better mouthfeel but further addition lowered the total sensory scores. Scanning Electronic Microscopy (SEM) analysis confirmed the modifications in noodles microstructure as CYP addition affected starch granule structure. In general, 30% substitution significantly improved the textural and rheological properties of noodles, indicating the potential of Chinese yam powder for industrial application. Noodles are rich in nutrition, easy to cook, and delicious traditional foods in Asian countries besides their increasing demand in many Western countries . TraditiIn the past years, researchers have been engaged to enrich noodles by fortifying them with various ingredients. The attempts have been made to use wheat noodles as a resource and then supplement them by adding broken rice, oat flour, barley flour, sweet potato flour, and buckwheat flour ,2,6,7. TDioscorea Polystachya (Turczaninow), is one of the important edible starch plants widely cultivated in China [Yam is recognized as a traditional staple food in many tropical regions in the world particularly in South Asia . Chinesein China  and it iin China  with genin China . Yam tubin China . Among tin China . Previous studies were conducted to evaluate the functional properties of noodles and the effects of various compounds to improve their final quality . Noodle Wheat flour  and table salt were purchased from Carrefour supermarket in Hefei city, China. The dried Chinese yam thin piece was procured from Tanan Local Wholesale Shop . At the laboratory, Chinese yam powder (CYP) was ground using Xichu Food Grinder , and passed through 100 mesh sieve.The proximate composition  of purchased wheat flour and CYP were tested by using the standard methods of Association of official analytical chemists (AOAC) . Ash conw/w). The aluminum crucible was stored under refrigerated conditions (4 °C) for 12 h. The empty crucible was taken as control. Initially, the starting temperature of the sample was 30 °C. After holding at 30 °C for 1 min, sample was heated to 90 °C at a rate of 10 °C/min [The starch gelatinization properties of wheat flour supplemented with different concentrations  of CYP were determined by Differential Scanning Calorimetry (DSC) . Briefly0 °C/min . The folTp: gelatinization temperature peak; To: onset gelatinization temperature; Tc: conclusion gelatinization temperature; R: gelatinization temperature range. R was calculated following Formula (1).According to a previous study with few modifications , the far−1 at 20 °C and as a function of frequency (0.1 to 10 Hz) using a Discovery Hybrid Rheometer-3  equipped with a cone-and-plate geometry . All parameters were tested as per the above-mentioned experimental conditions.Rheological evaluation, especially in the linear viscoelastic region, has been used for unraveling the structure and properties of dough in order to study the functions of dough ingredients . Elasticw/w)-supplemented wheat flours mixed with 50% distilled water, 2% NaCl, and 0.1% NaHCO3. To prepare the dough, 100 g of supplemented powder (different yam levels) was mixed with required amount of water in an automatic noodle machine . The dough was sheeted to about 2 mm thickness and the resulting fresh noodles were cut into 15 cm.Based on an earlier study , noodles1) and put into a beaker with 400 mL of boiled water. After cooking for 2 min, the noodles were allowed to cool down by placing them in cold water for 30 s followed by drying for 3 min and weighed (W2). The cooking yield of the noodles was calculated using the following Formula (2)For this, noodles (15 cm) strips were cooked for 2 min. According to the method of Lu et al. (2009) with few modifications , cooking2 = Weight of cooked noodle after drying; W1 = Weight of raw noodle.W3) is recorded as the weight of beaker after drying. The cooking loss of the noodles was calculated using the following Formula (3)For the cooking loss, the final weight  , three nFor sensory evaluation, five types of noodles samples (100 g) were cut into 15 cm length and cooked up to their optimal time (the time when white core in the cross section disappeared) in 100 °C water. The samples were dried for not more than 10 min in tightly covered plastic food containers before testing. Sample noodles were evaluated by 20 students and staff of the Food Science and Engineering College, Hefei University of Technology. All the samples were coded in a randomized order and evaluated by a 9-point hedonic scale . The different sensory attributes included color, appearance, taste, flavor, hardness, toughness, and overall acceptability. The raw and cooked samples were cut into 3 cm and frozen at −80 °C. After freeze-drying, the dried samples were cut into cross-section and placed on a specimen holder for coating with gold for 90 s. The microstructure of samples was observed by Scanning Electron Microscope , at 22 KV with 250× and 1000× magnification.All the measurements were performed in triplicate and the obtained results were expressed as the mean values ± standard deviation (SD). The results were statistically analyzed using SPSS . For the data analysis, one-way analysis of variance (ANOVA) was performed using the Origin Lab  software.The chemical compositions of original wheat flour, CYP, and the mixed flour with different amounts of CYP was presented in p < 0.05) as compared to CYP. From At first, lower gelatinization peak temperature in wheat flour was observed . The addition of CYP decreased the springiness of wheat flour noodles which might be due to decreased gluten E. No gluThe hardness initially increased from 25,504.35 g to 26,440.21 g after the addition of CYP (up to 30%) F which mIt can be seen from the The CYP-fortified cooked noodles were evaluated for their sensory properties and the results are presented in Up to 20% concentration of CYP, some sensory properties were satisfactory such as hardness, mouthfeel, and springiness, which was also revealed in TPA and SEM properties. Incorporation of more than 20% of CYP resulted in low total score and the noodles were not acceptable due to diluted gluten. Similarly, in an earlier study by Sanju and Kawaljit [The SEM microstructures of raw noodles and cooked noodles are shown in Our study explored the possibility of preparing noodles with acceptable quality from wheat flour supplemented with different levels of Chinese yam powder. Rheological, farinograph, and gelatinization texture properties were evaluated. It can be concluded that the addition of CYP to wheat flour can affect the overall properties of dough and noodles. When supplemented, the texture of noodles as evidenced by several parameters such as water absorption, cooking loss and firmness were improved. For sensory evaluation, some properties such as hardness, mouthfeel, and springiness were improved up to 20% incorporation of CYP, which supported the TPA properties. For microstructure, SEM revealed that addition of CYP could promote the close binding of starch particles to gluten protein and thereby resulted in relatively smooth surface. The above results encourage further studies to confirm the beneficial effects in terms of improved nutritional values of the noodles prepared from wheat flour supplemented with acceptable levels of CYP. They can in fact be advantageous for consumers seeking alternatives containing natural healthy ingredients in foods. The results of the current study demonstrate the potential for extending the use of Chinese yam powder in wheat noodle for industrial application.


To determine the differences in preterm infants’ stool microbiota considering the use of exclusive own mother’s milk and formula in different proportions in the first 28 days of life.The study included newborns with GA ≤ 32 weeks divided in 5 group according the feeding regimen: 7 exclusive own mother’s milk, 8 exclusive preterm formula, 16 mixed feeding with >70% own mother’s milk, 16 mixed feeding with >70% preterm formula, and 15 mixed 50% own mother’s milk and preterm formula. Exclusion criteria: congenital infections, congenital malformations and newborns of drug addicted mothers. Stools were collected weekly during the first 28 days. Microbial DNA extraction, 16S rRNA amplification and sequencing were performed.Escherichia and Clostridium was always greater in those containing formula than in the those with maternal milk only.All groups were similar in perinatal and neonatal data. There were significant differences in microbial community among treatments. Approximately 37% of the variation in distance between microbial communities was explained by use of exclusive own mother´s milk only compared to other diets. The diet composed by exclusive own mother´s milk allowed for greater microbial richness (average of 85 OTUs) while diets based on preferably formula, exclusive formula, preferably maternal milk, and mixed of formula and maternal milk presented an average of 9, 29, 23, and 25 OTUs respectively. The mean proportion of the genus Fecal microbiota in the neonatal period of preterm infants fed with exclusive own mother’s milk presented increased richness and differences in microbial composition from those fed with different proportions of formula. The intestinal microbiota is very important for human metabolism, development and behavior ,2. DespiThe immature intestinal microbiota of premature newborns is influenced by factors such as postnatal age, gestational age, birth weight and nutritional exposures ,13,14,15Therefore, the aim of this study is to describe the intestinal microbiota of very low birth weight infants in the first 28 days depending on use of mother’s own milk or use of formula in different proportions.th week of life. All samples were immediately stored in liquid nitrogen until DNA extraction.This study used a convenience sampling strategy with patients recruited from the Neonatology Section of Hospital de Clínicas de Porto Alegre (HCPA), Brazil from May 2014 to January 2017. Pregnant women with gestation age ≤ 32 weeks that provided written informed consent were enrolled at hospital admission for their delivery. The study protocol was approved by the Ethics Committee of Hospital de Clínicas de Porto Alegre (HCPA), and the guardians signed an informed consent form. Exclusion criteria were: 1) HIV or congenital infections, 2) mothers with substance abuse and 3) neonatal congenital malformations. Infants´ weekly stool samples were collected from diapers beginning with the first stool until the 4Newborns were divided in five groups according to the feeding: exclusive own mother’s milk (LME), exclusive formula (PFL), mixed 50% own mother’s milk and 50% formula (MFLM), mixed with formula and 70% or more of own mother’s milk (PLM), and mixed with own mother´s milk and 70% or more of formula (PFL). The newborns received daily the same diet for up to 28 days. The different amounts of formula and own mother´s milk was offered separately, at different times of the day, so that at the end of the day the ratio was maintained. The ratio of breast milk to formula was the limitation of the amount of breast milk available. The preterm formula used was Pre Nan. This formula does not contain neither probiotics nor prebiotics.Obstetrical data and neonatal data were collected prospectively.The laboratory technique as described in detail previously .5’-CCATCTCATCCCTGCGTGTCTCCGACTCAG-3’) and Ion adapter P1 sequence (5’-CCTCTCTATGGGCAGTCGGTGAT-3’) to obtain a sequence of primer composed for A-barcode-806R and P1-515F adapter and primers. Each of the 25μL of PCR mixture consisted of 2U of Platinum Taq DNA High Fidelity Polymerase , 4μL 10X High Fidelity PCR Buffer, 2 mM MgSO4, 0.2 mM dNTP's, 0.1 μM both the 806R barcoded primer and the 515F primer, 25μg of Ultrapure BSA  and approximately 50 ng of DNA template. PCR conditions used were: 95°C for 5 min, 35 cycles of denaturation at 94°C for 45s; annealing at 56°C for 45s and extension at 72°C for 1 min; followed by a final extension step at 72°C for 10 min.Microbial DNA was isolated from samples using the QIAmp Fast DNA Stool Mini Kit  following manufacturer’s instructions with modifications. All DNA samples were kept at -80°C until use. The V4 region of the 16S rRNA gene was amplified and sequenced using the PGM Ion Torrent platform  with the barcoded bacterial/archaeal primers 515F and 806R . PCR ampThe resulting PCR products were purified with the Agencourt AMPure XP Reagent  and the final concentration of the PCR product was quantified by using the Qubit Fluorometer kit  following manufacturer's recommendations. Finally, the reactions were combined in equimolar concentrations to create a mixture composed by 16S gene amplified fragments of each sample. This composite sample was used for library preparation with Ion OneTouch 2 System with the Ion PGM Template OT2 400 Kit Template . The sequencing was performed using Ion PGM Sequencing 400 on Ion PGM System using Ion 318 Chip v2 with a maximum of 40 samples per microchip.The 16S rRNA raw sequences were analyzed following the recommendations of the Brazilian Microbiome Project . BrieflyAll clinical data was analyzed using the software SPSS 21.0 at the significance level of 5%. Quantitative variables with normal distribution were described through means/SD and compared by ANOVA with Tukey test. Quantitative variables with asymmetric distribution were described through median/interquartile range and compared by the Kruskall-Wallis test with Dunn Test. For comparison among proportions Pearson's chi-square test was used in conjunction with residue analysis adjusted.For all 16S rRNA downstream analysis, the data set was filtered, keeping only OTUs that were present in at least 30% of the samples in each treatment and rarefied to the same number of sequences . The bioadonis function implemented in the vegan package [To accesses the main differences among treatments in this study, a Permutational Multivariate Analysis of Variance (PERMANOVA)  as perfo package .Differences between treatments were accessed through the STAMP software . P-valueAnalysis along the 28-day period was performed using meta-analysis of effect sizes reported at multiple points using general linear mixed mode .A total of 175 samples from 62 preterm newborns divided in five groups  were collected and analyzed. The five groups were similar in respect to maternal and obstetrical data .Neonatal data are shown in The microbial community differences at OTU level (97% similarity cutoff for grouping definition) found among fecal samples from preterm newborns’ fed with five different diets during a period of 28 days are presented in The overall analyses  indicate that approximately 31% of the variation in distances among treatments was explained by the different diets provided in each treatment. Pairwise comparisons revealed that diets based on maternal milk assembled microbial communities with large variation within group  while diets based on formula created more similar microbial communities  Table 3Table 3. Besides, the greatest variations in distances between treatments were observed in those comparisons involving the use maternal milk only. Approximately 37% of the variation in distance between microbial communities could be explained by the treatment with maternal milk only compared to the diet based preferentially in formula .In agreement with the ordination analysis, alpha diversity measurements indicated significant differences  among the richness of OTUs within treatments . The dieOnce overall differences among microbial communities found in fecal samples from preterm newborns fed with different diets during 28 days were detected, the next step was to identify the microbial taxa responsible for that difference. To detect differences among treatments at genus level, a pairwise differential abundance based on a two-sided White’s non-parametric t-test followed by the Benjamini-Hochberg FDR correction was performed. As LME treatment presented the greatest difference in microbial community among diets, the pairwise comparisons were performed between LME and the other diets .Escherichia was always greater in treatments containing formula  than in the treatment with maternal milk only. Particularly, the diet based on maternal milk presented an increased abundance of Acinetobacter, Bradyrhizobium, Caulobacter, Corynebacterium and Paenibacillus, Burkholderia, Faecalibacterium, Sphingomonas and the unknown genus from the Microbacteriaceae family as compared with the other treatments. Compared to the diet based on maternal milk, the diet preferably based on maternal milk (PLM) and the diet composed by a mixture of formula and maternal milk increased the abundance of Clostridium and Escherichia. The fecal samples from newborns fed with a diet preferable based on formula (PFL) presented greater abundance of Escherichia, Salmonella, Enterococcus and the unknown genus from the Enterobacteriaceae family when compared to the LME treatment  during the first 60 days of life fed with maternal breast milk, pasteurized donor human milk and preterm infant formula. It was found that those fed with maternal breast milk presented higher diversity . Cacho eEscherichia and Clostridium genus were always higher in treatments containing formula than in treatment which provided breast milk exclusively. This finding has been reported previously [In our study the average proportion of the eviously . A possieviously ,34. Besieviously , 36.In spite of several studies on the topic and the subject’s relation with diseases of high complexity ,3,4, sucBesides contributing to the microbial richness, breast milk also favors the prevention of sepsis, necrotizing enterocolitis (NEC) and other diseases ,31,33,37Other studies emphasize the influence of diet according to the ethnic and/or geographic characteristics as variables that influence the development of newborn intestinal microbiota ,40,41. IEscherichia and Clostridium have been associated to NEC as well as a high proportion of Proteobacteria with few numbers with Firmicutes [The microbiome found in breast milk contributes in the short and long term to the prevention of colonization by pathogens, as it stimulates the production of reactive antibodies and establishes a healthy intestinal microbiome capable of preventing long-term morbidities such as obesity, type 2 diabetes, chronic intestinal inflammation, autoimmune disorders, allergies, irritable bowel syndrome and allergic gastroenteritis ,42,43,44rmicutes .Some situations, however, crossed this research, such as the loss of fecal samples; the difficulty of mothers in breastfeeding preterm infants with breast milk exclusively due to socioeconomic difficulties, for example; and the low fecal volume produced by premature infants. Another important point to consider when analyzing the intestinal microbiota of the newborn, we analyze only the stools and we do not know the microbiota of the proximal colon neither of the small bowel. We could not adjust microbial analysis for birth weight and gestational age because the number of subjects in exclusive breast milk group was not big enough. It is very difficult to have extreme premature newborns fed exclusively with breast milk during the whole period in NICU hence we were not allowed to study different variables. Our objective was to compare exclusive own mother´s milk feeding with different proportion of formula feeding. We studied just extreme premature newborns, all of them included in the same category of prematurity, and those small differences in birth weight and gestational age probably do not have major repercussion on microbioma.Many studies have focused on the issue of breastfeeding, particularly regarding the newborns' microbiota, to identify its benefits for the development and prevention of diseases throughout their lives. Considering the importance of the topic, this study aimed to describe the intestinal microbiota of preterm newborns according to their nutritional habits establishing modifications of the intestinal microbiota according to the type of enteral diet administered.Based on our data, it is noticed that global differences of the microbial community are found among the types of diets administered to preterm infants, showing that the greatest microbial richness was found in those who received exclusive own mother’s milk in comparison with those that received different proportion of formula.


We previously described that loss of chromosome 10 and phosphatase and tensin homolog (PTEN), and associated pathological activation of the Akt/mammalian target of rapamycin (mTOR) pathway, is a hallmark of PTPR.3 Here, we for the first time report a patient case with relapsing and rapidly progressing PTPR that showed loss of chromosome 10 and PTEN, phosphatidylinositol-3 kinase (PI3K)/Akt/mTOR hyperactivity, and a dramatic and durable response to mTOR inhibition with everolimus.Papillary tumor of the pineal region (PTPR) was first described as a distinct tumor entity by Jouvet et al in 20034,5 More importantly, a copy number profile extracted from the DNA methylation profile demonstrated several chromosomal aberrations, most notably loss of chromosome 10 . In July 2016, MRI showed increased tumor volume, intensified contrast enhancement, and enhanced relative cerebral blood volume (rCBV) consistent with recurrent disease . EveroliNineteen months after initiation of everolimus treatment, the volume of contrast enhanced tumor in the pineal region had decreased by ~75% (from 30 × 24 × 22 mm to 19 × 16 × 14 mm), and contrast enhanced areas in the thalamus, mesencephalon, and splenium corpus callosum showed partial to complete regression . This waThis case indicates that PTPR—even after repeated relapses—depends on pathological mTOR activity and that mTOR inhibition represents a feasible option for targeted treatment in these patients. Based on the biological rationale and the well-documented response of the presented case, we conclude that everolimus should be considered as first-line treatment in patients with recurrent PTPR.The study received grant support from the Skåne University Hospital donation funds, the Medical Faculty, Lund University, governmental funding of clinical research within the national health services (ALF) (to M.B.), and the German Childhood Cancer Foundation (Deutsche Kinderkrebsstiftung) (to T.P.)Conflict of interest statement. The authors have no conflicts of interest to declare.


Determining the correct saddle fit is essential in order to optimise the interaction between the horse and rider dyad, and to reduce the risk of back-related problems or loss of performance as a result of incorrect saddle fit. Although there are industry guidelines (Society of Master Saddlers) on correct saddle fit, some saddle fitters (and others) choose to fit saddles that are wider than industry guidelines on the assumption that increased saddle width will enhance equine locomotion and allow the horses’ thoracolumbar spine to function unhindered. This study quantified the effect that a saddle that was one width fitting wider and narrower (based on the Society of Master Saddlers industry guidelines) had on the kinematics of the thoracolumbar spine, thoracolumbar epaxial musculature profiles, equine locomotion, and saddle pressure distribution. It was found that a saddle that was one width fitting wider and narrower affected the kinematics of the thoracolumbar spine, resulting in concavities in epaxial musculature at T13 when using the wide saddle and at T18 when using the narrow saddle. The wide saddle caused areas of high pressures in the cranial region of the saddle and the narrow saddle caused areas of high pressures in the caudal region of the saddle. It is essential that the correct saddle fit is achieved for each horse and rider combination in order to optimise the horse-rider system and reduce the risk of back-related problems or loss of performance that may occur as a result of incorrect saddle fit.p ≤ 0.05). Compared with the correct saddle width, in trot, in the wide saddle, an 8.5% increase in peak pressures was found in the cranial region of the saddle (p = 0.003), a 14% reduction in thoracolumbar dimensions at T13 (p = 0.02), and a 6% decrease in the T13 range of motion in the mediolateral direction (p = 0.02). In the narrow saddle, a 14% increase in peak pressures was found in the caudal region of the saddle (p = 0.01), an 8% decrease in the range of motion of T13 in the mediolateral direction (p = 0.004), and a 6% decrease in the vertical direction (p = 0.004) of T13. Compared with the correct saddle width, in canter, in the wide saddle, axial rotation decreased by 1% at T5 (p = 0.03) with an 5% increase at T13 (p = 0.04) and a 5% increase at L3 (p = 0.03). Peak pressures increased by 4% (p = 0.002) in the cranial region of the wide saddle. Altering the saddle fit had an effect on thoracolumbar kinematics and saddle pressure distribution; hence, correct saddle fit is essential to provide unhindered locomotion.This study evaluated the effect of saddle tree width on thoracolumbar and limb kinematics, saddle pressure distribution, and thoracolumbar epaxial musculature dimensions. Correctly fitted saddles were fitted by a Society of Master Saddler Qualified Saddle Fitter in fourteen sports horses , and were altered to one width fitting wider and narrower. Horses were equipped with skin markers, inertial measurement units, and a pressure mat beneath the saddle. Differences in saddle pressure distribution, as well as limb and thoracolumbosacral kinematics between saddle widths were investigated using a general linear model with Bonferroni adjusted alpha ( Correct saddle fit should enhance the athletic performance  of both Generally, saddle fitting is subjective and relies on the skills of an individual to assess the suitability of a saddle for both the horse and rider. In an attempt to standardise saddle fitting, the Society of Master Saddlers (SMS) has produced guidelines that qualified saddle fitters follow. Initially, a static saddle fitting assessment is performed featuring several criteria, as described elsewhere . Once eaUnder laboratory conditions, compared with a saddle that was of the correct tree width , saddles that were one width fitting narrower, one width fitting wider, and two width fittings wider have been reported . CompareThe pitch of a correctly fitted saddle should remain unchanged with a rider, and during locomotion. In a wide saddle, the cranial aspect of the saddle pitches ventrally and induces areas of high pressure beneath the cranial region of the saddle . In a naThe aim of this experiment was to quantify the effect of a wide saddle (one width fitting wider than required) and a narrow saddle (fitted one width fitting narrower) on limb kinematics, thoracolumbar kinematics, epaxial musculature dimensions, and saddle pressure distribution, when compared with a saddle fitted following industry guidelines   in horseIt was hypothesised that, in trot and canter during ridden exercise, a wide saddle would result in the following: (1) an increase in range of motion in the mediolateral direction of the thoracolumbar area; (2) areas of high pressures beneath the cranial portion of the saddle; (3) an increase in front and hind maximum fetlock hyperextension; (4) reduced carpal and tarsal flexion; and (5) concavities in the thoracolumbar musculature in the region of T13. It was hypothesised that the narrow saddle will result in the following: (1) areas of high pressure in the caudal region of the saddle; (2) increased axial rotation of the thoracolumbar region; and (3) concavities in the thoracolumbar musculature in the region of T18.The study was approved by the Royal Veterinary College ethics and welfare committee, project number URN 20181785-2.Fourteen adult non-lame sports horses were used in this study. All horses were warmbloods and of a similar type and conformation, all competing at affiliated dressage (elementary-advanced medium) and showjumping (British Novice—Foxhunter). All horses displayed good muscle definition with a well-defined musculature of the thoracolumbar region. Horses were housed at the same facility. All horses were geldings. They ranged in height at the withers (mean ± SD: of 1.67 ± 0.07 m), body mass (mean ± SD: 553 ± 24 kg), and age (mean ± SD: 9 ± 1 years). Horses underwent a subjective veterinary assessment performed by two veterinary surgeons, including flexion tests of all four limbs, and no lameness was observed. In addition, the horses’ gait asymmetry was quantified using a validated sensor system  . Two expTo limit the effect that a dressage or showjumping saddle design had on the rider, ten new general-purpose saddles (Kent & Masters), with an interchangeable gullet system, were used. For all conditions, a standard high withered saddle cloth  was positioned beneath the saddle. Each saddle was fitted by a SMS qualified saddle fitter and ridden in for ten hours prior to the study to allow the flocking (Jacob wool) to settle. Flocking and panel construction were evaluated between each saddle and horse by an SMS qualified saddle fitter. If the flocking had become irregular or compressed, then it was adjusted; these adjustments were performed by a single SMS qualified saddle fitter. The seat size remained the same (17.5″) for all saddles and the horse’s own girth was used throughout, which was a standard non-anatomically shaped girth (Atherstone) with elastic on one side. The girth was fastened, ensuring that the elastic was on the right side. The same stirrup leathers and stirrups were used and the stirrup length remained the same throughout. The order of saddle fit was randomized and blinded to all technicians, riders, and veterinarians . At someEach horse underwent a 15 min warm up protocol self-prescribed by the rider for each condition, which included walk, trot, and canter on both the left and right rein. This was followed by a prescribed trot and canter protocol, during which horse kinematics were quantified along with saddle-horse kinetics. Each saddle was evaluated independently by five SMSQSFs following the SMS static and dynamic evaluation guidelines. Data were collected during straight line locomotion in sitting trot and canter on both the left and right rein. All measurements were performed on the same indoor (60 × 20 m) arena on the same surface. The arena dimensions allowed for eleven repeated straight strides, hence eleven were included in the kinematic and kinetic analysis, with both the start and end points being determined using two cones . The surHorses were instrumented with eight MTw inertial measurement units (IMUs) (Xsens) using a validated sensor based system  ,19. By mrange of motion: maximum–minimum value over a stride cycle for x, y, and z displacement for trot and canterhip hike difference (HHD): difference between vertical upward movement amplitude of left and right tuber coxae during contra-lateral stance .IMU data were processed following published protocols . In brieFlexion-extension range of motion (ROM)—the body’s rotation about the transverse  axis.Axial rotation ROM—the body’s rotation about the longitudinal  axis.Lateral bending—the body’s rotation about the vertical  axis.-Vertical direction—up and down movement of the whole horse.-Lateral–lateral direction—side to the side movement of the whole horse.Translational movement at T5, T18, and L3 was measured in millimetres in two directions:Angular movement (a change in orientation) of T5, T18, and L3 was assessed in three planes; all data outcomes were measured in degrees.In order to allow interpretation of the effect of tree width, IMU derived kinematic variables were summarized between reins. Range of motion variables were subtracted from each other  and movement symmetry values  were added up . This procedure ensured that for horses performing symmetrically between reins, values near zero are expected, as head and pelvic movement symmetry values show directional circle dependent tendencies   and rangc) connected to a laptop via a fire wire was used to synchronize video and pressure measurements. The video was used to identify the maximal protraction of the inside hindlimb on both reins, at which point pressure data were extracted for analysis.Kinetic data under the saddle were recorded using a pressure mapping system . The pressure mat consisted of 256 sensors arranged into 8 columns  and 16 rows (transverse), left and right sides of the pressure mat . The mat was divided into two halves with no sensors over the midline of the back. Prior to the study, the pressure mat was calibrated, and during testing (for each horse), the pad was zeroed without the saddle, girth, or rider . It was c (30 mm) placed on each horse using double-sided tape. Marker locations have been described elsewhere [Kinematic data were recorded with a high-speed video camera system, using twenty-four half-sphere skin markerslsewhere . In the Maximum carpal flexion  lateral condyle of humerus, (4) lateral metacarpal condyles, and (5) distal aspect of the metacarpus over the LCL of the MCP joint).Maximum tarsal flexion  talus, and (11) distal aspect of the metatarsus over the LCL of the MTP joint) during the swing phase.Maximum forelimb fetlock extension during stance  LCL, (5) distal aspect of the metacarpus over the LCL of the MCP joint, and (6) origin of the LCL of the DIP joint)Maximum hindlimb fetlock extension during stance  talus, (11) distal aspect of the metatarsus over the LCL of the MTP joint, and (12) origin of the LCL of the DIP joint).Two high speed cameras  were positioned at a distance of ten metres from the experiment track, simultaneously capturing the left and right sides of the horse at 400 Hz , with a field of view capturing two complete trot or canter motion cycles. A halogen light was used to illuminate the markers. High speed video data were recorded and downloaded to a laptop  and processed using two-dimensional motion capture (Quintic Biomechanics). Described elsewhere ,24,25,27All raw data were smoothed using a Butterworth low-pass filter with a cut off frequency of 10 Hz .p < 0.04 for all measurements using three repeats per measurement.Thoracolumbar epaxial musculature dimensions were obtained using a new Flexi Curve Ruler , which was shaped around the dorsum, perpendicular to the dorsal midline at three levels of the vertebral column: T10, T13, and T18. Locations  were identified by manual palpation by the same technician. For repeatability, once identified, T10, T13, and T18 were referenced with skin paint. A single SMSQSF performed all measurements, with the horse standing square on a hard, level surface. Measurements of dorsal thoracolumbar body shape of each horse were obtained following the SMS guidelines . The thoFrom the two-dimensional kinematic analysis, data were collected from two consecutive strides with three repeats each, that is, six strides were used for analysis for trot and six strides for canter (three on each rein) for each horse and saddle condition .Maximum elbow flexion during the swing phase.Maximum MCP and MTP hyperextension during stance.Maximum carpal flexion during the swing phase.Maximum tarsal flexion during the swing phase.The outcome parameters for limb kinematics between saddle conditions  were as follows:From IMU and pressure distribution, data were matched in relation to movement condition and collected from eleven consecutive strides per repeat totaling a mean ± SD of 33 ± 3 strides, which was used for analysis of the trot and canter on both the left and the right rein for each horse and saddle condition .Craniocaudal, vertical, and mediolateral range of motion for T5, T13, T18, and L3 TS.Flexion and extension range of motion of T5, T13, T18, and L3 TS.Axial rotation range of motion of T5, T13, T18, L3 and TS.Lateral bending range of motion of T5, T13, T18, and L3 TS.The outcome parameters for the IMU derived data between saddle conditions  were as follows:Pressures beneath the cranial aspect of the saddle (kPa).Pressures beneath the caudal aspect of the saddle (kPa).Maximum overall force (N).The outcome parameters for the pressure distribution between saddle conditions  were as follows:p ≥ 0.05.As many kinematic parameters are influenced by speed, we tested for differences in speed using a repeated measures analysis of variance (ANOVA) by obtaining stride length and stride time from the two-dimensional analysis for each trial. This was repeated for each saddle condition , and no significant differences were found at p values were found to be ≥0.05), left and right rein data were pooled. A general linear mixed model was used for kinetic and kinematic data, with tree width  defined as a fixed factor and horse defined as a random factor. Two general linear models were created; one for trot and one for canter. For all outcome parameters, the significance level was set at p ≤ 0.05. A Bonferroni post hoc analysis was carried out to determine differences between pairs of conditions . Instead of applying the Bonferoni correction on the significance level, alpha, this study reported the Bonferroni adjusted p-values  multiplied by the number of comparisons done). This allowed us to assess the significance of our results  to the traditional alpha of 5%, without increasing type II errors.Statistical analysis was performed in SPSS . After checking to see if direction (left/right rein) had an effect  asymmetry values: HDmin, 1.47 ± 2.21 mm and HDmax 1.55 ± 2.72 mm, Pelvis Min Figure T5p = 0.02). Post hoc analysis showed an increase in craniocaudal ROM between the correct and wide saddle . Post hoc analysis showed no significant differences between the correct and narrow saddle widths (p ≥ 0.05). Craniocaudal range of motion (ROM) differed according to saddle widths (T13p = 0.03). Post hoc analysis showed an increase in flexion-extension between the correct and narrow saddle widths . Post hoc analysis showed no significant differences between the correct and wide saddle widths (p ≥ 0.05).Flexion-extension differed according to saddle widths (p = 0.03). Post hoc analysis showed an increase in axial rotation between the correct and narrow saddle widths . Post hoc analysis showed no significant differences between the correct and wide saddle widths (p ≥ 0.05).Axial rotation differed according to saddle widths (p = 0.003). Post hoc analysis showed decreased vertical movement between the correct and narrow saddle widths . Post hoc analysis showed no significant differences between the correct and wide saddle widths (p ≥ 0.05).Vertical movement differed according to saddle widths (p = 0.001). Post hoc analysis showed decreased vertical movement between the correct and wide saddle widths  and increased vertical movement between the correct and narrow saddle widths .Mediolateral ROM differed according to saddle widths  Bonferroni corrected post hoc analysis did not identify any pairwise significant differences Axial rotation differed according to saddle widths . Post hoc analysis showed an increase in axial rotation between the correct and wide saddle . Post hoc analysis showed no significant differences between the correct and narrow saddle widths (p ≥ 0.05).Axial rotation differed according to saddle widths (T13p = 0.01). Post hoc analysis showed a decrease in axial rotation between the correct and wide saddle .Axial rotation differed according to saddle widths (p ≤ 0.05). Post hoc analysis showed no significant differences between saddle width conditions (p ≥ 0.05).Vertical movement of T13 was different between saddle widths (T18p = 0.03). Post hoc analysis showed a decrease in flexion-extension between the wide and the correct saddle widths . Post hoc analysis showed no significant differences between the correct and narrow saddle widths (p ≥ 0.05).Flexion-extension differed according to saddle widths (p = 0.03). Post hoc analysis showed no significant differences between saddle width conditions (p ≥ 0.14).Lateral bending was affected by saddle widths (p ≤ 0.05). Post hoc analysis showed no significant differences between saddle width conditions (p ≥ 0.05).In the mediolateral direction, the ROM of T18 differed according to saddle widths (L3p = 0.007). Post hoc analysis showed a decrease in axial rotation between the correct and wide saddle width  and no significant difference between the correct and narrow saddle (p = 0.10).Axial rotation differed according to saddle width for both the narrow and wide saddle (p ≤ 0.05). Bonferroni corrected post hoc analysis did not identify any pairwise significant differences In the mediolateral direction, the L3 ROM differed according to saddle widths . Post hoc analysis showed an increase in peak pressures (kPa) in the cranial region of the saddle between the correct and wide saddle widths . Post hoc analysis showed no significant differences between peak pressures (kPa) in the cranial region of the correct and narrow saddle widths (p ≥ 0.05).Peak pressures (kPa) differed between saddles in the cranial region of the saddle (p = 0.003). Post hoc analysis showed an increase in peak pressures (kPa) in the caudal region of the saddle between the correct and narrow saddle widths  and narrow and wide saddle widths .Peak pressures (kPa) differed between saddles in the caudal region of the saddle (p = 0.11). Overall, force differed in the caudal region between saddle widths (p = 0.02). Post hoc analysis showed an increase in overall force (N) in the caudal region of the saddle between the wide and narrow saddle widths   in the cranial region differed between saddles, but was not found to be significant (= 0.002) .p = 0.04). Post hoc analysis showed an increase in peak pressures (kPa) in the cranial region of the saddle between the correct and wide saddle widths . Post hoc analysis showed no significant differences between peak pressures (kPa) in the cranial region of the correct and narrow saddle widths (p ≥ 0.05).Peak pressures (kPa) in thirteen horses differed between saddles in the cranial region of the saddle (p = 0.001). Post hoc analysis showed a decrease in overall force (N) in the cranial region of the saddle between the correct and narrow saddle widths  (p ≥ 0.05).The overall force (N) in the cranial region differed between saddles (= 0.002) . Post hop = 0.03). Post hoc analysis showed an increase in maximum tarsal flexion between the narrow and wide saddle widths . All other parameters were p ≥ 0.05 . Post hoc analysis showed a decrease at the level of T13  between the correct and wide saddle widths . Post hoc analysis showed no significant differences between the narrow and correct saddle widths (p ≥ 0.05).Thoracolumbar epaxial musculature profiles differed between saddle widths (p = 0.03). Post hoc analysis showed a decrease at the level of T18  between the correct and narrow saddle widths . Post hoc analysis showed no significant differences between the wide and correct saddle widths (p ≥ 0.05)  .This experiment quantified thoracolumbar kinematics during ridden exercise in trot and canter locomotion with the use of inertial measuring units. Following published protocols , this stThe cranial region of the thoracic spine (T10–T13) is of great interest; it is where the rider’s centre of mass aligns approximately above the horse’s centre of mass. It is the area where the saddle is at its narrowest and it is also a region that has been reported to be associated with muscle activity  and imprThere is limited evidence on thoracolumbar kinematics in canter during ridden exercise. In this experiment, it was observed that the wide saddle reduced flexion-extension at T18, which could be the result of the saddles’ balance and stability. Additionally, in the wide saddle, axial rotation was increased at T5 and decreased at T13 and L3. These findings would be in contrast to the belief by some saddle fitters that increased tree width allows for increased spinal kinematics. In the wide saddle, the cranial aspect of the saddle is pitched ventrally and it is speculated that this positioning (and increased pressures) limits the spinal range of motion. Our values are slightly higher for flexion-extension and axial rotation than those reported for horses cantering on a treadmill . This coFurther support that tree width affects equine locomotion arises from the saddle pressure distribution. In accordance with the hypothesis, in this group of horses, changes in saddle pressures and location of pressure distribution when fitted with a wide and narrow saddle were found. The findings reported here are in accordance with those reported elsewhere . In a wiSimilar to previous findings , the resWhen riding with the narrow saddle, the cranial part of the saddle is pitched dorsally (front rising up) and, as a result, the seat of the saddle is not parallel to the horse’s back, causing the saddle to rock backwards ,7. SaddlFinally, further support that saddle width has an effect on the thoracolumbar spine arises from our thoracolumbar dimensions. The thoracolumbar musculature has been reported to change over a year as a result of changes in body weight, seasonal variations, and saddle fit , as wellp ≤ 0.05), are in some cases of a small magnitude. Although some of the changes reported are small, it is speculated over time that these changes may have an effect on the locomotor apparatus of the horse; hence, a longitudinal study is warranted. Studies have reported changes in equine locomotion when pressures beneath a saddle [Although differences have been reported in this study, the authors appreciate that this study is limited by the number and type of horses used and that only two riders were used. The use of the Pliance pressure mat beneath the saddle may have influenced saddle fit, although this was consistent for all conditions. The effect that the Pliance mat may have on saddle fit should be considered when interpreting the results. The use of a girth with elastic on one side may have contributed to saddle instability and, if the study were to be repeated, then a non-elasticated girth should be considered. The changes observed are immediate and, although statistically significant (a saddle , bridle a saddle , and gira saddle  have beeCorrect saddle fit is essential to provide unhindered back function and optimise the interaction between the horse and rider dyad. This study has investigated the effect of tree width on thoracolumbar kinematics, limb kinematics, saddle pressure distribution, and thoracolumbar dimensions. A saddle that is fitted too wide or too narrow resulted in altered pressure distribution and, in some regions, changes in thoracolumbar spinal kinematics. Here, immediate changes were investigated and a longitudinal study would be useful in determining the long-term effect of varying tree widths. Horse owners should consider the effects that tree width has on the horse and seek regular professional advice to ensure optimal saddle fit. It is hoped that the findings from this study and those presented elsewhere ,50 will


Alternaria toxins are mycotoxins produced by various Alternaria species which, besides the Fusarium species, represent the principal contaminants of wheat worldwide. As currently, only limited information on the behaviour of Alternaria toxins during processing of cereals is available, the objective of this study was to investigate the effect of the dry milling process of wheat on Alternaria toxins distribution. Alternariol (AOH), alternariol monomethyl ether (AME) and tenuazonic acid (TeA) content were analysed by high performance liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) in all milling fractions of untreated (control), fungicide-treated, Alternaria tenuissima inoculated and commercial wheat sample. After dry milling process, in last break and milling flows and by-products, increased concentration of examined Alternaria toxins was detected. TeA was quantified in almost all milling fractions in all tested wheat samples, while AOH and AME were detectable mostly in last break and milling flows and by-products. In respect to the contamination with Alternaria toxins, white flour can be considered as relatively safe product. Since Alternaria toxins are concentrated mainly in the peripheral parts of the kernel, a special attention should be given to their content in low-grade flours and milling by-products. Tritricum aestivum) is the second most produced grain worldwide [Fusarium toxins, especially to deoxynivalenol (DON) [Fusarium spp., fungi of the genus Alternaria are considered as another group of ubiquitous pathogens on wheat leaves and ears. They are widespread in both humid and semi-arid regions and can infect growing plants in the field [Alternaria species, especially Alternaria (A.) alternata and A. tenuissima are frequently associated with several plant diseases of small grain cereals  [Alternaria toxins can be grouped into five different structural classes: dibenzopyrone derivatives—alternariol (AOH), alternariol monomethyl ether (AME) and altenuene (ALT); tetramic acid derivatives—tenuazonic acid (TeA) and iso-tenuazonic acid (iso-TeA); perylene derivatives—altertoxins I (ALX-I), altertoxin II (ALX-II) and alertoxin III (ALX-III) and stemphyltoxin III, a cyclic tetrapeptide—tentoxin (TEN) and Alternaria alternata f. sp. lycopersici toxins (AAL-toxins) [Alternaria toxins—AOH, AME, TeA and TEN—are generally found in certain grains and grain-based products, tomato and tomato products, sunflower seeds and sunflower oil, fruits and fruit products including fruit juices and in beer and wine [Alternaria toxins is an increasing concern for human health due to their possible harmful effects [Alternaria toxins have been described to possess genotoxic and mutagenic properties. In addition, they show cytotoxic, fetotoxic and/or teratogenic activity; they are mutagenic, clastogenic and oestrogenic in microbial and mammalian cell systems and they inhibit the cell proliferation [Alternaria toxins can be found in almost all food and feed products and that they might exhibit harmful effects on human and animal health, currently there are no specific international regulations or any national regulation in the world for any of the Alternaria toxins in food and feed, with the exception of Bavarian health and food safety authority who decided to limit the TeA content in sorghum/millet-based infant food at 500 µg/kg [Alternaria toxins worldwide. In order to enable better understanding of the real risk for public health, opinions of different European scientific committees [Alternaria toxins during storage and processing of food and feed. However, only limited information on the stability and fate of Alternaria toxins during storage and processing of food and feed is available [Alternaria toxins was observed [Alternaria toxins  by extrusion processing of whole wheat flour was investigated. The highest reduction of all three Alternaria toxins was achieved when high raw material moisture (w = 24 g/100 g), high feeding rate (q = 25 kg/h) and medium screw speed (v = 390 rpm) were applied. Under these extrusion conditions, a reduction of 65.6, 87.9 and 94.5%, was achieved for TeA, AOH and AME, respectively.Cereals and cereal by-products play an important role in the daily human diet, as well as in animal nutrition. Cereals are exposed to numerous biotic and abiotic stress factors during all phases, from cultivation to processing. The most common chemical contaminants of cereal crops are mycotoxins—secondary metabolites produced by toxigenic filamentous fungi as their natural protection either in the field or during storage . Among corldwide ,3 and thorldwide . After horldwide . During orldwide . Regardiol (DON) . Besideshe field . Among A blight) ,8,9,10 a blight) ,12,13. A-toxins) . The mosand wine . Since t effects . Some offeration . Despite00 µg/kg . This ismmittees ,16 recogvailable . After dobserved . In receobserved  the possAlternaria toxins during wheat milling process. Therefore, the main objective of this study was to evaluate the effect of wheat milling process on the distribution of AOH, AME and TeA in processed wheat samples.To the best of our knowledge, there is no data about the fate of −1, while in other examined wheat samples AOH was absent, as can be seen from the A. tenuissima with the initial concentration of 0.61 and 0.46 µg × kg−1, respectively. After cleaning, content of AME was below of limit of detection (LOD) in inoculated wheat sample, while in commercial wheat sample its content was reduced only by 8.2%. TeA was quantified in all the investigated samples and it was the most abundant of the monitored mycotoxins. After cleaning of the control, fungicide-treated, inoculated and commercial wheat grain samples, the content of TeA were reduced by 39.5, 40.7, 53.6 and 45.9%, respectively, in comparison to the initial TeA concentrations in the uncleaned wheat samples.The initial concentration of AOH in an uncleaned commercial wheat sample was 1.67 µg × kgAlternaria toxins, the results obtained in this study are in agreement with the results published so far regarding effect of wheat cleaning process to decrease the content of present mycotoxins before milling, summarized by Schaarschmindt and Fauhl-Hassek [Although there are no published data regarding the effects of wheat cleaning process on the content of l-Hassek  and confA. tenuissima inoculated wheat, as well as commercial wheat, were ground in the pilot scale mill (Bühler MLU 202). From the point of the interpretation of the results regarding the presence of Alternaria toxins in milling fractions of wheat, it is important to look at the yield of the obtained fractions . There is a significant share of the first two breaking fractions as well , while the third breaking flour (break flour III) is represented with a smaller share. All these flours belong to a group of white flours , which mFlours from the tail-end breaking runs  and reducing runs  belong to darker flours which make up a composition of low-grade flour  with a hAlternaria toxins content. Distribution of AOH, AME and TeA in milling fractions for all applied treatments is given in The obtained milling fractions were analysed for Only in commercial wheat sample concentration of AOH was above the LOD. Moreover, in the milling fractions obtained from all examined samples AOH was quantified in at least one of the fractions. The wheat milling fractions with the parts of wheat kernel where AOH is concentrated are primarily later breaking and reducing runs (tail-end passages), as well as milling by-products (shorts and bran). This result indicates that AOH is concentrated mainly in the peripheral parts of the kernel, which predominantly get into these fractions during the milling process.In the initial wheat samples, AME was present only in commercial wheat sample. Despite this, similar to AOH, AME was found in certain milling fractions  in all tParticularly interesting results of toxin distribution in milling fractions  were obtAlternaria toxins in flours and milling by-products a multi factorial analysis of variation was conducted  was calculated from the combination of the yield data during the milling process and the AOH, AME and TeA content. The white flours consist of the following milling fractions: reduction flours 1, 2, 3 and break flours I, II and III, while the composition of low-grade flours (dark flours) makes up reduction flours 4, 5, 6 and break flours IV, V and VI.The milling yields and AOH, AME and TeA mass balance of each milling product are reported in Alternaria toxins in white flour, absence of AOH was recorded in all cases, while AME was present only in white flour obtained from commercial wheat sample and its content amounted to 8.0% of the total contamination of the kernels. On the other hand, TeA was present in each of the white flour obtained and its content amounted from 7.65  to 19.8% (wheat sample inoculated whit A. tenuissima) of the total contamination of the kernel. However, the recorded concentration of AME and TeA in white flours intended for human consumption was relatively low. In case of contamination and distribution of examined Alternaria toxins in obtained low-grade flour (dark flour), it can be noted that only low-grade flour obtained from wheat samples which was treated with fungicide was not contaminated by AOH and AME. In other cases, content of AOH and AME amounted from 8 to 21.1% and from 8.72 to 64.2% of the total contamination of the kernel, respectively. The quantified concentration of these mycotoxins was low, while significantly higher content of TeA (37.0–73.8 µg × kg−1) was recorded in all low-grade flour obtained. Of the total contamination of the kernel, content of TeA in low-grade flours amounted from 9.7 (wheat treated with fungicide) to 10.9% (non-treated wheat sample). Since, low-grade flour (dark flour) which mainly contained end-tail breaking fractions  and reducing fractions  in which the content of TeA is higher , as the main milling product of wheat for human consumption ranged from 66.9 to 75.1%, while for low-grade flour that is, dark flour  the milling yield ranged from 7.4 to 8.7%. Regarding distribution of examined s higher  in relatAlternaria toxins in the milling by-products, it can be noted from the A. tenuissima the largest share of AME was found in shorts and its content amounted 100 and 41.9% of the total contamination of the kernel, respectively. Milling by-products are very often used for animal feed but can also be used for human consumption. Therefore, great attention should be given to the fact that Alternaria toxins are concentrated in these fractions of the milling process. The obtained results in this study could not be completely compared to the published data, since to the best of authors’ knowledge there is no previously published study regarding the fate of Alternaria toxins during dry milling process of wheat and their distribution in obtained milling fractions. However, comparing the results obtained in this study with the distribution of DON, the most studied regulated mycotoxin in wheat-based foods during dry milling process , summarized by Schaarschmindt and Fauhl-Hassek [Alternaria toxins takes place according to the same principle as in the case of DON.Regarding distribution of the analysed l-Hassek , the simAlternaria toxins in commercial milling products. One of the few studies refers to the presence of Alternaria toxins  in samples of commercial wheat flour from China [−1), AOH (30.2 µg × kg−1) and AME (27.1 µg × kg−1) were observed in these wheat flour samples, while the maximum content of AOH, AME, TeA and TEN were 98.7, 61.8, 520 and 129 µg × kg−1, respectively. Furthermore, currently published scientific report related to the dietary exposure assessment to Alternaria toxins in the European population [Alternaria toxins  are present in unspecified wheat milling products. According this scientific report among mentioned Alternaria toxins, TeA is the most frequent contaminant in commercial wheat milling products and this toxin is present with higher content in relation to others Alternaria toxins. Further, as already mentioned, published data around the world , followed by ALT-I (96.4%) and TeA (21.4%), while the highest median (116 µg × kg−1) and maximum content (305 µg × kg−1) were observed for AOH [−1, while AOH and AME rarely occurred in German wheat. ALT was present only in 7 out 267 wheat samples from Germany [−1, 48.9 and 18.6 μg × kg−1 and 70.2 and 39.0 μg × kg−1 for TeA, AOH and AME, respectively [Alternaria toxin in terms of either frequency or concentration in wheat from China and it was detected in 100% (370/370) samples with the highest mean (289 µg × kg−1) and maximum concentration (3330.7 µg × kg−1). Further, wheat from China were frequently contaminated with TEN (77%), followed by AOH (47%) and AME (15.1%). It should be noted, that TEN occurred with high mean (43.8 µg × kg−1) and maximum content (258.6 µg × kg−1) in Chinese wheat, also. Among 71 harvested wheat samples (2014–2015) from Albania, 82.9% in 2014 and 86.1% in 2015 were contaminated with some of the investigated Alternaria toxins  and the major detected Alternaria toxin was TeA with maximum, mean and median concentration of 175.7, 29.9 and 16.5 µg × kg−1, respectively [Alternaria toxins are frequently detected in wheat and therefore there is a great possibility for their presence in wheat-based product.Furthermore, there is also a lack of data on the presence of om China . Among 1 by EFSA , Germany by EFSA , Serbia  by EFSA , China [ by EFSA  and Alba by EFSA ) indicat and TeA .4%, whil Germany . Regardiectively . TeA wasectively . AccordiAlternaria toxins content in wheat milling fractions. In wheat milling process, even for wheat with concentration of Alternaria toxins under the LOD of the analytical method used, increased concentration of Alternaria toxins can be expected in fraction containing peripheral kernel parts, including last break and milling flows and by-products. In white flours, which represent the main part of the final products of wheat milling, the level of TeA was reduced by (83.6 to 90.4%) of the initial concentration in wheat before cleaning. In relation to TeA concentration in wheat after cleaning, in white flours the level of TeA was reduced by 72.3 to 82.3%. Therefore, white flour can be considered relatively safe product in respect to the contamination with TeA. In the by-products of wheat milling (bran and shorts), TeA was present in 2 to 4 times and in 3 to 6 times higher concentrations than in uncleaned and cleaned wheat, respectively. The wheat milling fractions with the parts of wheat kernel where AOH and AME were concentrated were primarily later breaking and reducing runs (tail-end passages), as well as milling by-products (shorts and bran). This result indicates that AOH and AME were also concentrated mainly in the peripheral parts of the kernel, which predominantly get into these fractions during the milling process. The future research should be related to the behaviour of Alternaria toxins under industrial conditions, in terms of managing the milling process of wheat in order to obtain safe final milling products. In addition, results of this first report may also represent useful support in the future consideration of the legal maximum limit for most common Alternaria toxins.Based on results obtained it can be concluded that both investigated factors  and their interaction express significant influence on the level of Alternaria toxins in wheat and wheat processing products. Experiment was carried out in the 2012/2013 season on the wheat (Triticum aestivum cv. Sirtaki). Samples of (1) wheat treated by fungicide,(2) wheat inoculated by A. tenuissima  and collection of wheat samples, which were also used in this investigation. Furthermore, (4) commercial wheat sample with visible discoloration called black points, which indicate potential fungal infestation by Alternaria spp., was procured (about 60 kg) from the warehouse for this study. The batches of all four wheat samples were well mixed in a NAUTA mixer  before taking samples for analysis and before dry milling process. Mixing homogeneity of wheat samples was assured by taking of four subsamples for analysis of investigated Alternaria toxins levels before cleaning of each examined wheat sample.The presented research represents one segment of comprehensive research targeted at investigations of incidence and possibilities of reduction of  Simmons .) and (3ublished ). FurtheWheat samples (500 g) were ground to a 1 mm particle size using a laboratory mill  to obtain the milled whole wheat from uncleaned and cleaned wheat samples.After cleaning of 10 kg of each wheat sample by a Laboratory Grain Cleaner 5150 , a portion of 3 kg was milled using a Bühler MLU 202  pilot scale mill. The wheat preparation for milling included wheat grain tempering  to 13.5% of moisture with a holding period of 24 h. This was followed by the second tempering up to the moisture content in wheat grain of 15% with an additional holding period of 30 min. Milling was carried out according to an extended scheme, presented in Each of the obtained milling fraction of each wheat sample were separately collected, weighed and kept at −20 °C before examination.Moisture content in wheat samples and in all milling fractions was determined according to ISO 712  and was The modified method from Siegel et al. , describAlternaria toxins  were quantified by high performance liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) using our previously published method without any modifications, including the equipment and materials [aterials .r2). To differentiate between extraction efficiency and matrix-induced signal suppression/enhancement, the slope ratios of the linear calibration functions were calculated to yield the apparent recovery (AR), that is, the overall method recovery and the signal suppression/enhancement (SSE) due to matrix effects. The recovery of the extraction step (ER), that is, sample preparation recovery, was calculated by dividing the overall recovery by the matrix effect as follows:MMCa is the slope of matrix-matched calibration, SCa is the slope of solvent calibration and SPa is the slope of spiked sample-prepared curve. For overall method recovery assessment, the wheat flour fractions were spiked in triplicate, over the ranges present in AR), respectively. Spiked samples were left overnight at room temperature to allow solvent evaporation and equilibration between analytes and matrix and were analysed using matrix-matched calibration curve. For the matrix-matched calibration curves (MMC), the wheat flour fractions were enriched with working standard solutions at the final reconstitution step, confirming linearity over the range presented in AR and MMC for all of the compounds and was used for matrix effects evaluation.The method was validated by in-house quality control procedure following the guidelines of Commission Decision EC 657/2002 . Method The precision of the method was expressed in terms of repeatability, that is, as the relative standard deviation (%RSD) of 6 replicates at three concentration levels  using thAlternaria toxins were quantified by external matrix-matched calibration procedure. Two matrix-matched calibration curves were constructed for TeA in the concentration range from LOD to 100 µg × kg−1 and from 100 to 250 µg × kg−1. For AOH and AME, one calibration curve was constructed in the concentration range from LOD to 10 µg × kg−1 and from LOD to 5 µg × kg−1, respectively. Linearity testing gave values of correlation coefficients (r2) above 0.9910 in all the investigated ranges. The obtained results were corrected for sample preparation recovery (ER) and were expressed on a dry matter basis. All samples were prepared and analysed in triplicates.In order to eliminate the effect of matrix, Alternaria toxins content in these composite flours, as the products that would be actually placed at the market by the mills, was calculated according to the following equation:i weight (g) of the fractions that make up the composition of white and low-grade flours; TiAlternaria toxins  content (µg × kg−1) in the fractions that make up the composition of white and low-grade flours.Further, based on the obtained data for weight of each flour fraction and its AOH, AME and TeA content, according to the scheme for obtaining of composite white and low-grade flours provided in P values <0.05 were regarded as significant.One way and multifactorial analysis of variance of obtained data were carried out. For determination of significance of differences among the mean values Duncan’s multiple comparison tests was used. For not detectable values (under LOD) of AOH and AME, data were adopted at the level of half of LOD. STATISTICA software v13.3 was used . P value


Ursus thibetanus) and wild boar (Sus scrofa) from May 2011 to March 2016 to examine seasonal variation in radiocesium (137Cs) concentrations in muscle tissues  of these important game species in Fukushima Prefecture. We measured muscle 137Cs of bears and wild boars killed by hunters or in animal control culls. First, using a linear mixed model (LMM), we tested for a positive relationship between muscle 137Cs and 137Cs in the soil at the site of capture  estimated from a soil 137Cs deposition map produced by the Japan Atomic Energy Agency. In the LMM, muscle 137Cs was positively related to estimated soil 137Cs, which corroborates the results of previous studies. The LMM regression coefficients differed between the two species, with wild boar muscle 137Cs being higher than that of bears sampled at the same locations. We then employed a generalized additive mixed model (GAMM) to estimate seasonal variation in the muscle 137Cs of the target species. GAMM showed that muscle 137Cs varied seasonally and that this seasonal variation also differed between the two species. In bears, muscle 137Cs decreased from spring to early autumn, before increasing in winter. Wild boar muscle 137Cs remained low during spring and summer and was high during autumn and early spring. These patterns are likely influenced by differences in diet, habitat use, and physiology between these two species.To elucidate and reduce the risk of radionuclide contamination in wildlife caused by the Tokyo Electric Power Company Fukushima Dai-ichi Nuclear Power Station accident, it is important to understand radionuclide variations in the wild animal population. Here, we used environmental monitoring data and muscle samples collected from Asian black bear ( We calculated the muscle 137Cs value on the animal capture date for each sample using the physical half-life of 137Cs (30.1 years). For statistical analysis, we used muscle 137Cs value samples for each species for which 137Cs concentration was above the detection limit  and the geographic location of each capture site  for analysis of seasonal variation in the muscle 137Cs for each species, we estimated soil 137Cs (Bq/m2) based on the 137Cs ground deposition map of the Japan Atomic Energy Agency’s (JAEA) 5th Airborne Monitoring Survey, which covered all of Fukushima Prefecture  and ending in March [month 12] because the sampling started in April each year) as the explanatory variable to estimate seasonal variation in muscle 137Cs. Animal species (Asian black bear or wild boar) was used as an interaction term of the explanatory variable to show statistical differences in the seasonal variation between the two species. We used sampling year as a random factor, for the same reason given for the LMM above. We removed the effect of positive relationships between muscle 137Cs and soil 137Cs on seasonal variation in muscle 137Cs by using soil 137Cs as the offset term to estimate pure seasonal variation in muscle 137Cs. Because the seasonal variation showed a cyclic pattern, we used the cyclic cubic spline smooth as the smoothing term, and estimated the degree of smoothing using the restricted maximum likelihood method [In the next stage, we analyzed seasonal variation in muscle d method . Then, w137Cs and soil 137Cs with log10 transformation. We used R3.3.1 [In the statistical analyses, we used muscle d R3.3.1 , mgcv pad R3.3.1 , lme4 pad R3.3.1 , and MuM137Cs and soil 137Cs was 4–1,080 Bq/kg FM and 1,563–227,118Bq/m2, respectively (Tag (m2kg-1FM: muscle 137Cs / soil 137Cs) for Asian black bear was 2.2×10−4–5.3×10−2 m2kg-1FM. In wild boar, the range of muscle 137Cs and soil 137Cs was 7–40,200 Bq/kg FM and 4,691–1,135,776 Bq/m2, respectively. The range of Tag for wild boar was 9.2×10−5–9.1×10−1 m2 kg-1FM.In Asian black bear, the range of muscle ectively . The ranUsing the ERICA tool, the mean and the range of total dose rate of Asian black bear was 0.103 μGy/h and 0.007–0.477 μGy/h, respectively. In wild boar, the mean and the range of total dose rate was 0.479 μGy/h and 0.010–12.700 μGy/h, respectively.137Cs and soil 137Cs  in Fukushima Prefecture was lower than that of brown bear (7.0 x 10−2 m2kg-1FM) in Finland [Tag from wild boar in Fukushima Prefecture after the FDNPS accident (3.2×10−3 m2kg-1FM) was lower than that in Europe after the Chernobyl Nuclear Power Station accident (8.0×10−3 m2kg-1FM in 2003 and 6.2×10−2 m2kg-1FM in 2004) [Tag from wild boar in Fukushima Prefecture (9.2×10−5 to 9.1×10−1 m2kg-1FM) was larger than that in Europe (4.0×10−3 to 1.5×10−1 m2kg-1FM) [When we compared the  Finland . The samin 2004) . HoweverWhen we calculated the total dose rate from our data using the ERICA tool –35, tota137Cs of wild boar and Asian black bear varied by season, and the pattern of seasonal variation differed by species. In Europe, seasonal variation in muscle 137Cs was observed in roe deer (Capreolus capreolus) and wild boar, and muscle 137Cs of those species was high from summer to autumn when fungi and underground food items accounted for a greater proportion of the diet [137Cs was not observed in red deer (Cervus elaphus), which did not forage fungi [137Cs in wild animals is affected by the food habits of the animals. In wild boar in Fukushima, muscle 137Cs was low from spring to summer and high from autumn to winter. This seasonal variation pattern supported our hypothesis 1, but it differed from seasonal patterns observed in Europe. Wild boar in Japan mainly forage the leaves of grasses and woody species in spring and summer, and mainly forage roots, tubers and hard masts in autumn and winter [137Cs in wild boar in Fukushima. However, because wild boar are omnivores, their food habits change with changing habitat conditions, resulting in seasonal variation patterns in muscle 137Cs. For example, when hard masts are abundant, muscle 137Cs decreases during part of the peak season of muscle 137Cs because wild boar forage large amount of hard masts in autumn [137Cs concentration in food items in the diets of wild animals in Japan are lacking. Further study of the seasonal food habits of wild boar in Fukushima and 137Cs concentrations in food items are needed to further elucidate the reasons for seasonal variations in muscle 137Cs.Our study revealed that muscle the diet ,16,17, athe diet –25. On tge fungi . Therefod winter –11. Thusn autumn ,25. Furt137Cs increased from autumn to winter. This result did not support our hypothesis 2, but our hypothesis 3 was supported. The autumn and winter seasons coincide with large intakes of hard masts and denning [137Cs was low when foraging on hard masts was higher because hard masts contained low 137Cs [137Cs concentration in food items are likely to be responsible for the elevated muscle 137Cs of Asian black bear. The muscle 137Cs of roe deer increased during the season in which food intake was higher [137Cs concentration in food items, but also the effects of intake rate on seasonal variation in muscle 137Cs of the Asian black bear. However, the food habits of the Asian black bear in Fukushima and 137Cs concentration in food items has not been reported, and these remain important issue for understanding the seasonal variation in muscle 137Cs.In Asian black bear, muscle  denning ,26,27. Bow 137Cs –25, facts higher . Therefo137Cs increased in autumn when bears prepared for denning and peaked in the winter denning season. Other bear species such as the American black bear (U. americanus) and brown bear (U. arctos) also have a denning season in winter [137Cs, and physiological changes in bear species in preparation for denning begin to take place in autumn [137Cs from autumn to winter. To understand the mechanisms of seasonal change in muscle 137Cs in bear species, understanding physiological changes during the denning season is also important.Our hypothesis 3 was supported by the study results, however, understanding the physiological characteristics of the target species is also important to revealing the mechanism of seasonal variation in radionuclide concentrations. Particularly for Asian black bear, muscle n winter , and then winter . Suppresn autumn ,51. Thus137Cs of Asian black bear changed most drastically during autumn when they mainly foraged on hard masts and had the lowest muscle 137Cs in September followed by the highest muscle 137Cs of the year in November and December. It is possible that changes in the location of foraging as hard masts drop from treetops to the ground during late autumn may play a role in the increase in muscle 137Cs of Asian black bear. In the forest environment, radionuclides are more heavily concentrated at the forest floor, which is the habitat of the Asian black bear and wild boar [137Cs in Asian black bear in autumn, it is necessary to show the relationships between intake of radionuclides and changes in foraging location. Recently, bio-logging technology  to monitor animal behavior has advanced and the application of such technology to study the relationships between radionuclide intake and animal behavior, especially changes in foraging location, is promising.The muscle ild boar ,53. Thus137Cs of Asian black bear during February to March, which is after the end of the hunting season on February 15 and when bears enter denning. Obtaining muscle samples during this period will be difficult but important to elucidating the relationships between radionuclide concentrations in the muscle of the Asian black bear and physiological changes due to denning.In this study, no data were obtained for muscle 137Cs exceeding the standard limits for general foodstuffs (100 Bq/kg). From our findings, sampling for radiocesium monitoring during winter when was the muscle 137Cs is likely to be at its maximum will be necessary to lift the transportation restriction. Radiocesium monitoring in Asian black bear and wild boar that excludes winter sampling would result in underestimation, and we therefore need to develop methods for radionuclide concentration monitoring that take into consideration the seasonal variation in radionuclide concentrations in wild animals.This is one of the few studies to report seasonal variation in radionuclide concentration in wild animals after the FDNPS accident. Notably, to the best of our knowledge, this is the first report on seasonal variation in radionuclide concentration in a bear species. The findings of our study provide useful information not only about the radionuclide dynamics of wild animals but also monitoring radionuclide concentration in wild animals in Japan. Such information may be helpful in developing hunting management strategies based on monitoring. In Fukushima Prefecture, transport of Asian black bear and wild boar was restricted following the report of muscle Our study showed that radionuclide concentrations and seasonal variation differed between two large omnivorous mammal species in a region impacted by radionuclide contamination. To elucidate the mechanism of radionuclide transfer from the environment to wild animals, it is necessary to study factors that affect the specific differences that were revealed in our study. Taking concepts from previous studies and the ecological traits of both species into consideration, we suggest that the following factors are the most important when evaluating a target species: 1) food habits, 2) habitat use, and 3) seasonal changes in physiology.