Chinese yam polysaccharides (CYP) on myogenic differentiation 1 (MYOD1), myogenin (MYOG), and myostatin (MSTN) mRNA expression of breast and thigh muscle tissues in broilers. A total of 360  crossbred broilers chicks with similar body weight (BW) were randomly distributed into four groups, with three replicates in each group and each replicate included 30 broilers. The feeding trial lasted for 48 days. Experimental broilers were fed 0.00 mg/kg basal diet (control group), 250 mg/kg, 500 mg/kg, and 1000 mg/kg CYP, respectively. The results showed that CYP250 and CYP500 groups had higher thigh muscle percentage (TMP) compared to the control group (p < 0.05). Meanwhile, the expression of MYOD1, MYOG mRNA in breast muscle tissues of CYP500 and CYP1000 groups was higher (p < 0.05), and the expression of MSTN mRNA in thigh muscle of CYP250, CYP500, and CYP1000 groups was lower than that of the control group (p < 0.05). In addition, there was no significant difference in the expression of MYOD1 mRNA in the thigh muscle tissue of each group (p > 0.05). Bivariate correlation analysis showed that the expression levels of MYOD1, MYOG, and MSTN mRNA in the thigh muscle tissue of broiler chickens in the CYP500 group were positively correlated with TMP. However, the expression of MYOG mRNA in thigh muscle tissue of the CYP1000 group was negatively correlated with TMP. In general, this study indicated that appropriate dietary CYP supplementation influenced the growth and development of thigh muscle tissue in broilers by altering TMP and muscle tissue development-related genes expression. Therefore, CYP could be used as a potential feed additive to promote the development of muscle tissues in broilers.This experiment was conducted to evaluate the effects of dietary  Chinese yam (CY) and Lycium barbarum. They have a variety of biological functions, such as regulating intestinal microflora, antioxidant and anti-inflammatory responses, anti-tumor, and so on [Dioscorea opposita Thunb [The Pharmacopoeia of the People’s Republic of China clearly records that yam, as one of the traditional Chinese herbs, has edible and medicinal value in China for more than 1500 years [As the biological resources of natural plants, Chinese herbs are a source of green feed additive with no residue, wide source, and low cost, which contains polysaccharides, organic acids, flavonoids, and other bioactive substances . Chinesend so on ,3,4, whind so on , therefota Thunb , which ita Thunb . The 20100 years . It cont00 years ,10, amon00 years . Animal 00 years . CYP sup00 years . Adding 00 years . Our pre00 years .MYOD1, MYOG, MSTN, and other regulatory factors [MYOD1 is a skeletal muscle-specific transcription factor, which can act on activation, proliferation, and differentiation of satellite cells, and plays an important biological regulatory role in fast- and slow-muscle formation [MYOG is a key regulator of skeletal muscle differentiation and plays an important role in terminal myoblast differentiation [MSTN, also known as growth differentiation factor 8 (GDF-8), negatively regulates skeletal muscle growth by blocking myoblast proliferation and differentiation to control the number of muscle fibers [MYOD1, MYOG, and MSTN gene mRNA in broiler breast muscle and thigh muscle tissue. In order to explore the relationship between CYP and the growth and development of muscle tissues in broilers, the purpose of this experiment is to study the effect of adding CYP in the diet on broiler breast muscle percentage (BMP), TMP and gene expression related to muscle development, and the relationship between gene expression and BMP or TMP. These results will help us understand the regulatory function of CYP in the growth and development of muscle tissue and the expression of related genes in broilers.Skeletal muscle is one of the important tissues for weight gain in broilers, including breast and thigh muscles , with it factors ,17. MYODormation ,19. MYOGntiation ,21. MSTNe fibers ,23. Howe250, CYP500, and CYP1000, respectively) with 3 replicates in each group and each replicate included 30 broilers . The control group was fed a basal diet without CYP, and the CYP250, CYP500, and CYP1000 groups received the same basal diets included 250, 500, and 1000 mg/kg CYP, respectively. Feeding trials were divided into early (1–28 days) and late (29–48 days). The basal diet meeting the National Research Council  requirements is shown in All procedures involving broilers feeding were reviewed and approved by the Animal Protection and Utilization Committee of Henan Institute of Science and Technology . A total of 360  crossbred broilers chicks with similar initial BW (39.54 ± 0.51 g) were randomly allotted to 4 groups . After the neck was bled and slaughtered, breast and thigh muscle were harvested and weighed respectively, and used for carcass performance determination. The percentage of breast and thigh muscle tissues are calculated as a percentage of the eviscerated weight. Meanwhile, breast and thigh muscle tissue were collected from the same site, preserved in liquid nitrogen, and then stored at −80 °C in a freezer until further analysis.β-actin, MYOD1, MYOG, and MSTN was determined by the 2−ΔΔCt method.Total RNA was detached from breast and thigh muscle tissues by employing TRizol reagent . After extraction, total RNA utilizing a spectrophotometer  measured the concentration of RNA within the A260:A280 ratio range. The RNA was reverse transcribed using test kits by Thermo Fisher Scientific to make cDNA. Each sample was measured in triplicate. Relative mRNA expression of https://www.ncbi.nlm.nih.gov/genbank/) known sequences were designed with Primer 5.0 software β-actin, MYOD1, MYOG, and MSTN mRNA primers [Based on GenBank . Muscles percentage and gene expression data were analyzed using a one-way ANOVA test. Prism 8  was used to generate graphs. SPSS 26.0 for Windows  was used to perform the Pearson’s correlation coefficient two tail test to evaluate the correlation between muscle development-related genes expression and BMP or TMP. p > 0.05). However, compared with the control group, the TMP of broilers in the CYP250 and CYP500 groups was higher (p < 0.05).Effects of dietary CYP on the BMP and TMP in broilers are shown in 500 and CYP1000 groups had higher MYOD1 mRNA expression compared to the control group (p < 0.05). Meanwhile, compared to the control group, the CYP500 and CYP1000 groups had higher MYOG expression, CYP250 and CYP500 groups had higher MSTN expression (p < 0.05).As shown in MYOD1 mRNA in the thigh muscle tissues of each group (p > 0.05), while MYOG and MSTN mRNA expression in CYP250, CYP500, and CYP1000 groups was significantly lower than that in the control group (p < 0.05). In addition, MYOG and MSTN mRNA expression of the CYP500 group was higher than that in the CYP250 and CYP1000 groups (p < 0.05).As shown in MYOD1, MYOG, and MSTN mRNA expression showed no significant correlation with BMP, TMP in each group (p > 0.05).As shown in MYOD1, MYOG, and MSTN mRNA expression was significantly positively correlated with TMP in the CYP500 group , while the MYOG mRNA expression was negatively correlated with TMP in the CYP1000 group .As shown in Astragalus membranaceus polysaccharides (AMP) significantly enhanced the TMP of chickens. Zhang et al. [L. barbarum polysaccharides (LBP) additives could improve TMP and enhance the muscle tissue development of early chickens. The present study found that dietary appropriate CYP supplementation could significantly increase TMP of broilers. Our study was similar to previous reports showing that CHPs can increase TMP and promote muscle tissues development in broiler chickens.Previous study showed that BMP and TMP are important traits of carcass yield, which are closely related to the increase of broiler growth and carcass yield ,26. Manyg et al.  found thg et al.  also fouMYOD1, MYOG, and MSTN play key roles in the muscle growth and development. MYOD1 and MYOG are members of the myogenic regulatory factors (MRFs) family, and their expression levels are closely related to the growth rate of muscle tissues [MSTN has been shown to be involved in inhibiting muscle precursor cell proliferation, which determines muscle tissue growth and development [MSTN is also involved in the formation of skeletal muscle in broilers [500 and CYP1000 significantly enhanced the mRNA expression of the MYOD1 and MYOG in breast muscle tissues. In addition, a diet which is CYP supplemented significantly downregulated the mRNA expression of the MSTN in thigh muscle tissues of broilers. These results indicate that dietary appropriate CYP supplementation has a positive effect on the expression of MYOD1 and MYOG genes related to muscle development in broilers, and can also inhibit the expression of the negative regulator MSTN. All of these results suggest that the CYP promotes broiler muscle development by regulating the expression of muscle-related genes, which may be related to intestinal absorption and utilization of nutrients or digestive enzyme activities. However, how the mechanisms CYP regulate gene expression need to be further studied.Numerous evidence demonstrated that  tissues ,31. As aelopment . In addibroilers . In the MYOD1 is a primary myogenic regulator, which is expressed before differentiation of myoblasts [MYOG is a secondary myogenic regulator, which is expressed during and after differentiation of myoblasts [MSTN is an important negative regulator of muscle tissue growth and negatively regulates muscle growth in animals [MYOD1, MYOG, and MSTN mRNA expression in breast muscle tissues showed no significant correlation with BMP and TMP of the broilers. Moreover, the MYOD1, MYOG, and MSTN mRNA expression in thigh muscle tissues of the CYP500 supplemented group was significantly positively correlated with TMP of broilers. Meanwhile, the MYOG mRNA expression in thigh muscle tissues of the CYP1000 supplementation group was negatively correlated with TMP of broilers. The results demonstrated that diets supplemented with CYP500 and CYP1000 might affect the growth and development of thigh muscle tissues in broilers. Overall, the MYOD1, MYOG, and MSTN mRNA expression was correlated with TMP of broilers in thigh muscle tissues. Additionally, dietary appropriate CYP supplementation may affect muscle tissue development of broilers by regulating the expression of MYOD1, MYOG, and MSTN mRNA. However, the exact molecular mechanism needs further study.During muscle development, yoblasts , and MYOyoblasts , both of animals ,37. AccoMYOD1, MYOG, and MSTN in thigh muscle tissue was significantly positively correlated with TMP in the CYP500 supplemented group. MYOG mRNA expression in thigh muscle tissues of the dietary CYP1000 supplementation group was significantly negatively correlated with TMP. At the same time, adding appropriate CYP to the diet can increase the expression of MYOD1 and MYOG and reduce the expression of MSTN mRNA, thereby improving the breast and thigh muscles yield in broilers. Therefore, CYP could be used as a potential feed additive to promote the development of muscle tissues in broilers.In this study, the mRNA expression of


BRAF V600E mutation and a homozygous deletion in PTEN, without changes in other relevant genes like EGFR, CDKN2A, TP53, NFKBIA, CDK4, MDM2 and PDGFRA. Although PTEN homozygous deletions are described in gliomas, the present case constitutes the first report of a PTEN mutation in a DIA, and this genetic feature may be related to the malignant behavior of a usually benign tumor. These genetic findings may point at the need of further and deeper genetic characterization of DIAs, in order to better understand the biology of this tumor and to obtain new prognostic approaches, a better clinical management and targeted therapies, especially in malignant cases of DIA.Desmoplastic infantile astrocytoma (DIA) is rare, cystic and solid tumor of infants usually found in superficial cerebral hemispheres. Although DIA is usually benign, uncommon cases bearing malignant histological and aggressive clinical features have been described in the literature. We report a newborn patient who was diagnosed with a DIA and died postresection. Pathologic examination revealed that the main part of the tumor had benign features, but the internal region showed areas with a more aggressive appearance, with higher-proliferative cells, anaplastic GFAP positive cells with cellular polymorphism, necrosis foci, vascular hyperplasia with endothelial proliferation and microtrombosis. Genetic study, performed in both regions of the tumor, showed a  Desmoplastic infantile astrocytoma (DIA) is a meningocerebral neuroepithelial tumor of infancy defined by a combination of distinctive clinicopathologic features . DIA wasBRAF V600E mutation and PTEN homozygous deletion. Genetic studies of DIA are very scarce. Since its first description in 1984, only twelve works performed genetic search of DIA mutations, and most of them focused exclusively on the detection of the well characterized BRAF V600E mutation [BRAF mutation have been described [PTEN have never been reported. Table Here, we report a rare case of DIA in a male newborn, with histological characteristics of malignancy, mutation , 18–26. escribed , 23, 26,The patient was a newborn male, product of the first pregnancy of a 27-year-old mother. Ultrasonography controls during pregnancy reported an adequate gestational age, with a 88 mm biparietal diameter at week 34. However, a subsequent control at week 39 showed a high biparietal diameter, 109 mm and an intracranial hypoechoic image. An emergency caesarean was performed. External analysis showed a cephalic perimeter of 38 cm, cephalic suture softening and prominent fontanels. A MRI scan on day 1 showed a supratentorial tumor of 75 × 63 × 76 mm with heterogenic signal and cystic areas. The tumor produced a midline deviation, hydrocephaly and subfalcian herniation  was used . Results showed a BRAF mutation that led to a substitution of valine by glutamic acid at position 600 (V600E), in both parts of the tumor  was found in both areas  including those in ALK/ROS1/NTRK/MET were found in 21 cases of infantile hemispheric glioma and four DIGs, but in none of the DIA cases. From this series, only three cases were diagnosed as DIAs, without BRAF mutations, and one of them with losses in chromosomes 2, 9 and 22, and gains in 5 and 10 [BRAF were found [BRAF V600E mutation. The study concluded that no recurrent genetic alterations were identified in the series, and high-grade and low-grade areas did not have significant genetic differences that could explain their distinct morphology or biological behavior.Knowledge of genetic alterations in these tumors is limited. Table istently . From tht of all , 19. BRArocytoma , but its DIA-DIG . Three irocytoma , 19, 20.rocytoma . In 2018ic tumor . ChaterjA tumors . Two morpositive , 23. In  gliomas . In 20205 and 10 . Imperatre found . Finallyre found . From thBRAF V600E mutation is now considered relatively common in DIA [Taken together, although n in DIA , 12, 19,n in DIA , 12, 36.BRAF V600E mutation and a PTEN homozygous deletion. Both mutations were present in the different areas of the tumor, apparently showing a lack of correlation between the aggressiveness of the area and the genetic findings. Interestingly, although PTEN homozygous deletions are found in high-grade gliomas [PTEN mutation in a DIA. PTEN is a haploinsufficient gene tumor suppressor which regulates various cellular processes, including genomic stability, survival, proliferation and metabolism. Due to its role, a subtle decline or a partial inactivation of PTEN functions substantially induces susceptibility to cancer and tumorigenesis [PTEN allele contributes to increase the aggressiveness of gliomas, being involved in the transition from the low to the high grade of these tumors [PTEN could explain, at least partially, the malignant transformation of a typical benign tumor like DIA. Elucidating the mechanisms underlying tumorigenesis mediated by PTEN loss in gliomas would be important and can reveal its potential role in atypical DIAs with this mutation, like the present case.In the present work, we show a rare case of a DIA from a newborn, with histological features of malignancy,  gliomas , the preigenesis , 40. Seve tumors –43 and te tumors , 45. SimPTEN alterations in DIA, it is unknown whereas PTEN mutational status may provide useful information for the prognosis of this tumor, and distinguish a new PTEN-deficient category of aggressive DIAs. These genetic findings, concurrent with the anaplastic histological characteristics of the tumor, point at the possibility of analyze the mutational status of PTEN, together with BRAF and other still undiscovered gene mutations in new cases of DIA, in order to better understand the behavior of this rare tumor, provide a new classification of DIAs, achieve a better management of the patients and obtain new targeted molecular therapies.Since no previous studies reported data about


Belgian endive is grown in a two-step cultivation processthatinvolves growing of the plants in the field, cold storage of the taproots,and a second growth period in dark conditions called forcing to yieldthe witloof heads. In this study, the changes in the carbohydratecontent and the secondary metabolite composition were studied in differenttissues of Belgian endive during the cultivation process. Belgianendive heads contain between 336–388 mg/g DW of total solublecarbohydrates, predominantly fructose and glucose. The heads alsocontain phenolic compounds and terpenoids that give Belgian endiveits characteristic bitter taste. The terpenoid and phenolic compoundcomposition of the heads was found to be constant during the cultivationseason, regardless of the root storage time. In roots, the main storagecarbohydrate, inulin, was degraded during storage and forcing processes;however, more than 70% of total soluble carbohydrates remained unusedafter forcing. Additionally, high amounts of phenolics and terpenoidswere found in the Belgian endive taproots, predominantly chlorogenicacid, isochlorogenic acid A, and sesquiterpene lactones. As shownin this study, Belgian endive taproots, which are currently discardedafter forcing, are rich in carbohydrates, terpenes, and phenolic compoundsand therefore have the potential for further valorization. This systematicstudy contributes to the understanding of the carbohydrate and secondarymetabolite metabolism during the cultivation process of Belgian endive. Cichorium intybus is a perennialplant, which is grown as a vegetable (C. intybus L. var. foliosum) and is appreciatedfor its bitter taste. Examples are Belgian endive and radicchio. Anothervariety of chicory C. intybus L. var. sativum, also called industrial chicory, is cultivatedfor the extraction of the food fiber inulin. Belgian endive or witloofis an important vegetable crop in Belgium, France, and the Netherlands.The estimated total yearly EU production of Belgian endive amountsto 350,000 ton.12 The second cultivation process called forcingstarts with defrosting of the taproots, that are subsequently cultivatedon hydroponics in specialized chambers in the dark at elevated temperatures.The temperature choice depends on the Belgian endive variety used.After 3 weeks, a blanched, densely packed head is formed on the taproot.The head is harvested, and the remaining forced taproot is discardedas waste. The storage conditions of the taproots can influence thequality of the produced head.4 However, after manyyears of optimization, the current hydroponics-based production isrobust and guarantees a year-round production of fresh Belgian endive.For the productionof Belgian endive two cultivation phases areneeded 1.First,1 Currently,the roots do not have a high value and are discarded or used as animalfeed. The Belgian endive taproot is rich in carbohydrates. Inulin,a linear polymer of fructose linked to a terminal glucose moiety,is the major storage carbohydrate in chicory roots.5 In industrial chicory, about 70% of the taproot dry weight(DW) is composed of inulin.7 The degree of polymerizationof inulin typically ranges from 2 to 60; however, fractions with alow degree of polymerization are removed when inulin is extractedto be used as food fibre.8The discarded forced taproots form the major wastestream of theBelgian endive cultivation, constituting more than half of the harvestedbiomass. In Europe approximately 300,000–400,000 tons of forcedroots (FR) are produced annually.9 During storage and forcingof the Belgian endive taproots, the quantity of inulin is decreasedas it is converted into monosaccharides to be used for the basic metabolismin the storage phase, as well as for the development of the head inthe forcing phase.10In Belgianendive, typically inulin of low degree of polymerizationis present; therefore, its taproots are not exploited as a sourceof dietary fibre.12 Industrial chicory is particularly rich in hydroxycinnamic acidderivatives, including chlorogenic acid and isochlorogenic acid A . Inaddition, two caffeic acid esters of tartaric acid, that is, caftaricacid and chicoric acid, are accumulating in industrial chicory butare predominantly found in the leaves.13 In a study on different varieties of C. intybus, it was shown that also Belgian endive contains phenolic acids, namelychlorogenic acid, caftaric acid, and chicoric acid, like many otherleafy chicory vegetables.14 The phenoliccompounds are thought to be involved in the plant’s defenseagainst biotic and abiotic stress.15 Theirproposed biological functions in plants include antibacterial, antiviral,and antifungal activities, and they also serve as antioxidants andoxygen free radical scavengers. In particular, the di-CQA’shave potent antioxidant capacity, for example, in coffee (Coffea canephora)16 andare increased upon UV exposure, as shown in Vitis vinifera leaves17 and artichoke.18 Also, for humans, the phenolics have potential health effects.Epidemiological studies suggest correlations between the intake ofhigh levels of phenolics and the prevention of some diseases,19 such as thrombose and inflammation.20 It is generally believed that the health-promotingactivities of the phenolics are due to the ability to scavenge freeradicals, as shown by Kono et al. for chlorogenic acid and caffeicacid.21In additionto carbohydrates, the taproots of both industrial chicoryand Belgian endive are rich in phenolic compounds and terpenoids 2.11,12 IC. intybus plants are also rich in sesquiterpene lactones (STLs).22 STLs are sesquiterpenes consisting of a 15-carbonskeleton with a characteristic lactone ring containing a conjugatedexomethylene group (α-methylene-γ-lactone). The majorSTLs reported in chicory are the guaianolide-type STLs lactucin, 8-deoxylactucin,and lactucopicrin, predominantly present in their oxalated form.22 In plants, the STLs function in resistance againstseveral pathogens25 and play a role in resistance to herbivore attack.26 The oxalate esters of STLs are thought to havean antifeedant role. As they are unstable and may decompose upon tissuedamage, both oxalic acid and the STLs will be released and may contributeto the antifeedant properties.22 In chicory,as well as in many other plant species, STLs are stored in latex inspecialized organs present in the leaves and the roots called laticifers.27 STLs are known to have several health benefitsfor humans.28 The sesquiterpene α-methylene-γ-lactone-ringof the molecules is thought to be essential for their functionality.This ring structure facilitates the alkylation of cellular thiol-groups,found in, for example, glutathione and in cysteine-containing proteins.In this manner, STLs influence the functioning of thiols in regulatingcellular mechanisms, for example, by sensitizing cancer cells forchemical treatments30 or lowering blood pressure inhumans.31 Chicory STLs show analgesic andsedative activities in mice.32 An interventionstudy showed the anti-inflammatory potential of chicory STLs againstosteoarthritis in humans.33The roots and leaves of In thiswork, metabolic changes in carbohydrate, phenolic compound,and sesquiterpene lactone profiles were studied in Belgian endivetaproots and heads during its cultivation cycle, including field harvest,storage, and forcing. The carbohydrate and secondary metabolite compositionof heads, which are used as a vegetable, was studied through the cultivationseason of 1 year. Additionally, the metabolic composition of taproots,which are currently discarded after forcing, was studied to determinethe potential for their upcycling.C. intybus L. var. foliosum) cultivars “SweetLady” and “Flexine” were grown on experimentalfields at Praktijkpunt Landbouw Vlaams-Brabant vzw in year 2019. Both cultivars are considered late cultivars that canproduce high-quality heads after prolonged storage for up to 1 year.Field cultivation was performed according to general crop husbandrycomparable to commercial fields. The USDA soil classification forthis location was Alfisol.34 In October2019, the taproots were harvested, cleaned, and stored in wooden containersin storage compartments at −1 °C. Samples of taproots(field roots) and field-grown leaves (field leaves) were sampled atharvest, processed as described below, and stored until metaboliteanalysis. Second cultivation, also called forcing, was performed after1, 3, 6, and 12 months of root storage for cultivar “SweetLady” and after 12 months for cultivar “Flexine.”In parallel, the Belgian endive cultivar “Sweet Lady”was cultivated, stored for 6 months, and forced at a second locationat Inagro in Rumbeke-Beitem, Belgium. Taproots were defrosted, placedin trays, and cultivated in hydroponic cultures in darkness for 3weeks at 15–16 °C. The hydroponics solution was supplementedwith 50% Kristalon Label Blue (Yara), 25% potassium nitrate, and 25%calcium nitrate to a final EC  of about 1.8mS/cm at pH = 6.8. Non forced roots (NFR) are roots that were defrostedand sampled before the second cultivation phase started. Forced roots(FR) are roots collected after the cultivation in hydroponic cultureand harvesting of Belgian endive heads. FR, NFR, and heads were sampledfor carbohydrate- and secondary metabolite analysis in triplicate.For each replicate, 20 roots or heads were collected, washed undercold tap water, and cut into 1 cm3 cubes using a RobotCoupe cutter, model CL 50 Ultra . The samples were subsequently freeze-dried using an Epsilon2–10 D LSC freeze dryer and milled to a fine powder using an Ultra centrifugal mill ZM 200. The resulting dry powders were stored inairtight sealed aluminum-foil-laminated plastic bags  at room temperature until analysis.Belgian endive  in all samples of the cultivar “SweetLady” cultivated at the location Herent. The analysis was performedaccording to van Waes et al.35The total carbohydrate and inulin content was analyzed in hydrolyzedextracts of the freeze-dried plant material. Hydrolysis was performedin water with the addition of 5 mL of 3 M hydrochloric acid at 85°C for 1 h. After cooling, neutralization, and filtration, thesamples were analyzed with a Dionex ICS 3000 chromatography systemon a Metacarb 67C (Agilent Technologies) column at 90 °C withdeionized water as a mobile phase at a flow rate of 0.5 mL/min. Peakdetection was performed using a Perkin Elmer 2414 refractive indexdetector. The total carbohydrate content was calculated as the sumof fructose and glucose in the hydrolyzed plant extract divided by1.1. The inulin content was calculated as the total carbohydrate contentfrom which the concentrations of free glucose, fructose, and sucrosewere subtracted. The mDP was calculated as the ratio of fructose toglucose in the hydrolyzed plant extract.36 In brief,100 mg (+/–3 mg) of freeze-dried plant material was extractedusing 1400 μL of 80% methanol supplemented with 0.13% formicacid. The samples were incubated for 15 min in an ultrasonic bath.Next, the debris was separated from the extract by centrifugation.LC–MS analysis was performed using an LC-PDA-LTQ-Orbitrap FTMSsystem (Thermo Scientific), which consisted of an Acquity HPLC withan Acquity eLambda photodiode array detector (220–600 nm) connectedto an LTQ/Orbitrap XL hybrid mass spectrometer equipped with an electrosprayionization source (ESI). Chromatographic separation was on a reversed-phasecolumn at 40 °C. Degassed eluent A [ultra-pure water: formic acid ] and eluent B [acetonitrile: formic acid ] wereused at a flow rate of 0.19 mL min–1. A linear gradientfrom 5 to 75% acetonitrile (v/v) in 45 min was applied, which wasfollowed by 15 min of washing and equilibration. FTMS full scans (m/z 90.00–1350.00) were recordedwith a mass resolution of 60,000 FWHM.Semipolar metaboliteswere analyzed by LC–MS.Quantification of selectedmetabolites was performed by comparison of LCMS peak areas to a standardcurve prepared from authentic standards of chlorogenic acid (Sigma-Aldrich),caftaric acid (Sigma-Aldrich), chicoric acid (Sigma-Aldrich), isochlorogenicacid A (Sigma-Aldrich), lactucin (Extrasynthese), and lactucopicrin(Extrasynthese). For compounds for which authentic standards are notavailable, the MS detector response (ion counts/scan) of the basepeak in the negative ionization mode is presented as the relativepeak area.37 . Baseline correction and noise determination were performed, andsuccessively, the m/z values werealigned. After removing low and inconsistent signals, that is, presentin <3 samples or with an ion intensity lower than 5000 in all samples,the remaining mass signals were subjected to MSClust software38 in order to cluster mass signals derived fromthe same metabolite based on their corresponding retention time andabundance pattern across samples. This resulted in the relative peakintensities of 554 and 295 mass clusters, respectively, for the datagenerated on root and head samples of the cultivar “Sweet Lady,”respectively, each representing a putative metabolite present in atleast 3 samples.For the targeted metaboliteanalysis, mass-specific peak areas were integrated using Xcalibursoftware version 4.1 (Thermo Scientific). For untargeted data processing,the LC–MS data files were processed using Metalign softwarep ≤ 0.05), we followed up with Bonferroniposthoc tests between time points within each tissue type and betweentissues within each timepoint. For those compounds with a nonsignificantinteraction (p > 0.05), the term was dropped fromthe model, and posthoc tests (Tukey’s test) focused on differencesbetween levels of one factor only . Proportions, such as the total carbohydrate,were analyzed analogously using logistic regression with a beta distributionusing R package GlmmTMB. A type III Wald test was used to assess thesignificance of main effects and interactions, such as tissue typeand storage time, for all components using R package Car. For themDP, a two-way ANOVA was performed to reveal the interaction betweentissue type and storage time for these compounds. Finally, the R packageEmmeans was used to carry out pairwise comparisons by an approximate t-test in combination with the multiple testing correctionsmentioned above .Principal component analysis (PCA)was performed on the log2-transformed and mean-centered data matrixusing the software package SIMCAp . Statisticalanalysis was performed on individual metabolites and attributes thatwere further studied using SPSS software  and R (version 4.0.3). Two-way analysis of variance (ANOVA) wascarried out to assess the differences in metabolite levels betweenthe tissues , the effect of forcing on the roots,and the effect of root storage time on the roots and head in “SweetLady.” The model comprised fixed effects for tissue type, storagetime , and their interaction.An F-test employing type III sums of squares was used to assess thesignificance of each effect. In case of a significant interactioneffect  enzyme depolymerization activity that is induced at harvest.39 and the level of total carbohydrates in theroots.43 The inulin mDP measured in rootswas low, ranging from 3 to 8. This is in line with previous observationof inulin mDP in Belgian endive.44 Owingto low mDP, Belgian endive roots are not suitable for inulin extractionas a dietary fiber.9 While inulin contentdecreases during forcing, unexpectedly, the mDP value seems to beincreased upon forcing. This is most probably the result of the formationof inulo-n-oses, polymers of fructose not containingthe terminal glucose, which disturb the mDP calculation, as previouslydiscussed by van Arkel.6An additional decrease in inulin content in taproots is observedupon forcing when the carbohydrates are mobilized to form the head.Simultaneously, the level of free fructose in FR is increased three-foldas compared to NFR. This finding is supported by a previous studywhere the effect of forcing was studied on the different inulin polymers45 Indeed, expression of vacuolar invertase hasbeen reported in the Belgian endive head as well.46 The duration of storage of taproots did not influence thecarbohydrate profiles found in the head. This supports the ratherconstant head quality throughout the cultivation season, as the freesugar levels might influence the palatability of the head. The headsshow a high carbohydrate content compared to the field leaves. Thisdifference could be explained by the functional differences betweenthe two tissues, the field leaves are source organs for sugars thatare designed to export the sugars to the roots. The leaves of theBelgian endive head are a sink organ that uses the carbon from thetaproot for its fast outgrowth.The Belgianendive heads contain between 336 and 388 mg/gDW oftotal soluble carbohydrates 3. In con47 However, our finding is in contrast with an earlier studyperformed on the storage of chicory taproots for 6 weeks, where anearly complete breakdown of inulin was found upon forcing.48 An explanation for the observed difference couldbe the storage temperature: in our experiments, a temperature of −1°C was applied in contrast to the +3 and +5 °C that Rutherfordand Weston used. We demonstrate here that after storage of roots at−1 °C, high amounts of soluble carbohydrates are stillpresent in the FR. The free sugars and inulin left in the FR, therefore,could be an interesting source for the production of biogas or platformchemicals, such as HMF.49Belgian endive heads are consumedas a vegetable, while the rootsare discarded after forcing. As shown here, the taproots contain 634mg/g DW of soluble carbohydrates at harvest, and after forcing, thecarbohydrate content is only partially reduced to about 476–509mg/g DW. In a previous study, the amount of stored sugar was not deemedlimiting for the quality and yield of the crop.The forcing process significantlymobilizes the carbohydrates reserves in chicory roots, as shown hereand in earlier studies. However, the effect of forcing and storageduration on the profile of secondary metabolites of the Belgian endiveis not well studied. Untargeted LCMS-based metabolomics analysis wasperformed to see the effect of the storage and forcing on the metabolitefingerprints. The PCA of the root samples based on the relative levelsof 554 putative compounds indicated clear differences between samplesof field roots, NFR, and FR. The PCA showed that the main variation in the data set is caused by the process of forcing 4A. The eNext, targeted quantitative analysis of phenoliccompounds andSTLs was performed. The major phenolic compounds identified in rootsand heads of Belgian endive were chlorogenic acid, isochlorogenicacid A, caftaric acid, and chicoric acid, and the major STLs foundwere lactucin, dihydrolactucin, lactucopicrin, lactucopicrin 15-oxalate,lactucin 15-oxalate, dihydrolactucin 15-oxalate, and 8-deoxylactucin15-oxalate. For example, LC–MS profiles of the Belgian endivehead and NFR are shown in 13 In contrast to the observationthat industrial chicory leaves (var. sativum) contain low levels of chlorogenic acid and isochlorogenic acidA, both taproots and heads of Belgian endive (var. foliosum) accumulate significant levels of thesecompounds in our study. The levels of most phenolic compounds in thetaproots were not influenced by forcing, with the exception of isochlorogenicacid A, which showed between a 2.1 and 4.6 fold increase upon forcingat different time points. Forcing may have induced conversion of theactivity of the so far uncharacterized enzyme in chicory,13 which converts chlorogenic acid into isochlorogenicacid A; however, only a small reduction in chlorogenic acid was observed.Next, the amount of chlorogenic acid, isochlorogenicacid A, caftaricacid, and chicoric acid was quantified 1. The le51Quantification was also performed for seven majorSTLs 6. TheamTable S4). The level of the STLs in taprootswas affected by cold storage (table S4). At different timepointsduring storage, root samples were transferred to the hydroponic cultureand forced to produce the heads. The levels of phenolic compoundsin the FR were not affected upon prolonged storage of the roots .In order to investigatewhether the findings from our analysison the Belgian endive cultivar “Sweet Lady” would alsohold true for another Belgian endive cultivar, the cultivar ‘Flexine’was taken along in this study. Taproots of the cultivar “Flexine”were stored and forced after 12 months of storage rather than at differenttime points, as described for “Sweet Lady.” The targetedmetabolomics analysis of ‘Flexine’ revealed a comparableoverall effect of forcing on the target metabolites and a significantincrease in the taproot upon forcing of isochlorogenic acid A, lactucin,and lactucopicrin 15-oxalate, as was found for “Sweet Lady”. Inan earlier study, it was shown that the cultivation location can havea pronounced effect.4 This latter studywas performed on many more cultivars and five different locations,giving the opportunity to better study the effect in detail. It couldalso be that the two crop cultivation facilities used in the presentstudy are more comparable to each other than the locations used inthe study by Peters et al.In order to study the effect of the location of cultivation andforcing on the metabolite composition, a second location was includedin this study. The cultivar “Sweet Lady” was grown,stored, and successively forced in Beitem, Belgium, in a comparablesetup as in location Herent. At this location, the taproots were storedfor 6 months and then forced. The stored roots, FR, and heads werecollected and included in this analysis. The conclusion from the targetedanalysis of the phenolic compounds and STLs was that for these threetissues, most of the studied compounds were not significantly differentbetween the two locations, with the exception of a significantly lowerlevel (50%) of dihydrolactucin in heads harvested al the locationBeitem (This study demonstrates a stable carbohydrate compositionof Belgianendive heads throughout the cultivation season. Additionally, thecomposition of major STLs and phenolics, important for the bittertaste of the heads, remains largely unaltered during the entire cultivationseason until up to 12 months. Importantly, this study also demonstratesthat the Belgian endive roots are potentially a valuable source ofcarbohydrates, terpenes, and phenolics. This highlights the potentialof Belgian endive roots for valorization, whereas they are currentlylargely unused, and 400.000 tons are discarded in the EU each year.Further studies into the extraction and processing of the roots, aswell as on bioactivity of extracted compounds, are needed to supportthe development of valorization strategies.


They also showed a higher number of bruxism episodes with arousal compared with nonsmokers , indicating increased sleep fragmentation. However, no significant effect of alcohol on SB intensity was observed, and the bruxism episode index was similar in alcohol drinkers and nondrinkers. In addition, electrolyte disturbances and lipid disorders were evaluated. Compared with nonsmokers, tobacco smokers showed a higher concentration of plasma triglycerides (177.67 ± 106.9 vs. 129.18 ± 65.61) and lower levels of iron and magnesium . Conclusions: In summary, this study showed that tobacco smoking, but not alcohol consumption, is related to bruxism intensity and lipid and electrolyte disturbances in individuals with sleep disorders.Background: Sleep bruxism (SB) is a common sleep-related movement behavior with a complex etiology. A recent hypothesis suggests psychoactive substance usage, tobacco smoking, and alcohol intake are risk factors for SB. This study aimed to evaluate SB intensity in tobacco smokers and alcohol drinkers. Methods: A total of 133 adults underwent full-night audio- and video-polysomnography, and the polysomnograms were evaluated using the American Academy of Sleep Medicine guidelines. The study group was divided into smoker and nonsmoker groups as well as drinker and non-drinker groups. Results: The results of the polysomnographic analysis confirmed that tobacco smoking has a significant effects on SB. Tobacco smokers showed increased bruxism intensity (5.50 ± 4.71 vs. 3.83 ± 3.26,  Sleep bruxism (SB) is a major sleep-related movement behavior with a complex etiology and an uncertain and insufficiently understood pathophysiology .Various As bruxism has become an emerging problem worldwide and attracted research interest in the medical field, Lobbezoo et al. established a new definition of bruxism, which was extended without the prior restrictions of earlier definitions . They deSymptoms of SB such as jaw clenching and tooth grinding are risk factors for several health problems . MasticaSB episodes can be analyzed using polysomnographic examination—the gold-standard tool for SB diagnosis—which is based on night video and audio recordings, along with bioelectric signals from electroencephalograms, electromyography, electrocardiograms, and airflow detectors ,16.The Lancet in 2019, the major risk factor for death in countries with a middle to high socio-demographic index is the usage of tobacco products, which causes, among other effects, high systolic blood pressure, dietary risks, overweight, and high fasting plasma glucose levels. The primary risk factors have changed from 1990 to 2019, but the pattern remains comparable. In the 25–49 year age group, alcohol use is the most common risk factor, but smoking has become a less significant risk factor and has steadily declined, mainly due to widespread government actions  that resulted in a behavior change in populations [Cigarettes and alcohol are legal psychoactive substances that affect the central nervous system (CNS) and can also affect the behavior, cognitive functions, and mood of individuals who consume them. Alcohol contains ethanol, which is a CNS depressant that acts as a gamma-aminobutyric acid receptor agonist and an N-methyl-D-aspartate receptor antagonist. Tobacco products contain nicotine, which is an acetylcholine agonist that can stimulate or depress the CNS depending on the amount consumed . The mosulations . Recentlulations . Cardiovulations ,24,25.An unhealthy lifestyle, including the consumption of stimulants, can lead to significant health problems that affect the general health of patients and is a risk factor for bruxism.Based on this knowledge and due to the lack of data on the cause–effect relationship, the effects of psychoactive substances on SB intensity were hypothesized in this study. The null hypothesis was that increased bruxism intensity occurs in tobacco smokers and alcohol drinkers compared with controls. The aim of the study was to investigate the relationship between bruxism intensity in tobacco smokers and alcohol drinkers using polysomnographic assessments. An additional aim of the study was to evaluate the lipid profile and concentrations of electrolytes and iron in blood samples of tobacco smokers and alcohol drinkers.This was a prospective, observational study carried out on 133 adult Caucasian individuals in the Sleep Laboratory of the Department and Clinic of Internal Medicine, Occupational Diseases, Hypertension and Clinical Oncology at Wroclaw Medical University. This study was approved by the Ethical Committee of Wroclaw Medical University (no. KB-790/2022). All patients signed voluntary informed consent prior to their participation in the study.n = 82) used legal stimulants, including tobacco (n = 24) and alcohol (n = 77). Those who consumed both alcohol and tobacco contributed to 14% of the study group (n = 19). Alcohol drinkers who did not smoke contributed to 44% of the study group, and smokers who did not drink contributed to 4% of the study group. The following comorbidities were observed among the participants: myocardial infarction (n = 7), stroke history (n = 6), hypertension (n = 52), diabetes (n = 24), and ischemic heart disease (n = 10). The characteristics of the study group are presented in Among the 133 participants, 62%  was smoked per day. Smoking status was categorized as smoker or nonsmoker. There were no former smokers in the study group, and all of the smokers were current smokers. Patients were classified as alcohol drinkers or nondrinkers based on self-reporting of any amount of alcohol consumption. Patients were divided into four groups: smokers To confirm SB diagnosis and investigate sleep architecture, an instrumental approach was followed using video-polysomnography (PSG) with a NoxA1  device. Recordings were made between 22:00 and 06:00, in accordance with the natural circadian rhythm of the patients. Electromyographic electrodes were placed according to the recommendations of the American Academy of Sleep Medicine (AASM). The PSG examination included video and audio recording, along with electrocardiographic, electroencephalographic, electrooculographic, and electromyographic recordings , body position detection, and thoracic and abdominal breathing activity. Saturation level (SpO2%), pulse, and plethysmography data were recorded using a NONIN WristOx2 3150 pulse oximeter . The recordings were stored using Noxturnal software . The obtained PSG recordings were analyzed and interpreted in 30 s epochs in accordance with the guidelines for sleep scoring by a qualified physician in the Sleep Laboratory .Rhythmic activities of masseter muscles occurring at least 3 s after the previous muscle activity were recognized as SB episodes. These episodes were frequently accompanied by specific movements in the orofacial region and grinding sounds. The bruxism episode index (BEI) was used to measure the bruxism intensity by counting the number of bruxism episodes per hour of sleep. Three phenotypes of SB were distinguished using electromyographic assessments: (1) phasic, characterized by ≥3 bursts of muscle contractions at 1 Hz frequency; (2) tonic, characterized by a contraction that lasts >2 s; and (3) mixed, including both phasic and tonic movements . After on = 133) was recruited in the present study.The sample size was evaluated using a sample size calculator based on the following standard assumptions: population size, 3,000,000; population proportion, 8%; confidence level, 0.95; and maximum error, 5%. The required group size was found to be 114. Thus, a larger group  in the case of parametric data and the Mann–Whitney U test or Kruskal–Wallis ANOVA in the case of nonparametric data. Correlations were evaluated using Pearson correlation coefficients. A p value < 0.05 was considered to be statistically significant.Statistical analysis of the data was carried out using the statistical analysis program Statistica 13.3 . The data are presented as the mean ± standard deviation. Differences between the groups were analyzed using Student’s n = 60) of the study group and men comprising 55% (n = 73). The mean pack-years in the smoker group was 13.60 ± 2.77. The polysomnographic parameters are presented in The average age of the study population was 47.89 ± 16.57 years, with women comprising 45% , phasic BEI , mixed BEI , tonic BEI , and bruxism with arousal .Statistically significant positive correlations were observed between pack-years and BEI  content was observed among smokers compared with nonsmokers; however, no significant differences were observed between alcohol drinkers and nondrinkers. Noticeable changes were observed in the electrolyte and blood composition of smokers, which were characterized by lower magnesium (Mg) and iron (Fe) levels in the blood. No statistical differences were observed in sodium, potassium, calcium, total cholesterol, low-density lipoprotein, and high-density lipoprotein levels in smokers compared with nonsmokers. The blood parameters are presented in Although many research studies on SB are available in the literature, the majority of studies are based only on non-instrumental approaches using self-reported or telephone surveys ,31,32,33In an international consensus, Lobbezoo et al. established that an instrumental approach for PSG assessment is the gold-standard for “definite” diagnosis of SB . Non-insThe most important finding of this study was the higher BEI in smokers compared with nonsmokers. Taking into account the electromyographic phenotype of episodes, a higher number of mixed bruxism episodes was observed in smokers compared with nonsmokers. As reported in a previous study, bruxism episodes are strongly associated with the sleep stages of non-rapid eye movement sleep, including the N1 and N2 stages, which are light sleep stages . To the According to the AASM guidelines, arousals are “sleep perturbations characterized by a transient increase (for 3–15 s) in electroencephalography fast wave activity, with or without an increase in EMG activity and cardiac rhythm” ,36. Sleen = 133) to analyze SB-related movements and sleep architecture in smokers and alcohol drinkers. These results support the findings of previous studies, indicating that SB can be related to smoking [To the best of our knowledge, this is the first study to use PSG in a large study group , and increases the index of microarousals . FurtherSeveral studies have reported the deleterious effects of tobacco on general health. A previous study investigated the influence of tobacco smoking on iron homeostasis dysregulation . AnotherThe effect of smoking on dyslipidemia is a well-researched topic. Increased serum lipid profiles, especially TG, are strongly correlated with obesity and smoking, which is consistent with the findings of the present study ,67,68,69The finding that the magnesium content was lower in smokers is consistent with the findings of previous studies ,71. ThisIn the present study, no significant differences in bruxism intensity and sleep architecture were observed between alcohol drinkers and nondrinkers. It is well established that alcohol consumption has adverse health effects on health and sleep ,74. AlcoThis study included a large study sample of 133 patients. Furthermore, this study was based on polysomnography, which is the “gold-standard tool” for SB diagnosis according to Lobbezoo et al. . MoreoveThere were a few limitations to this study. The small size of the smoker group is correlated with a higher risk of errors than in a larger sample group. In addition, patients were admitted to the Sleep Laboratory with no adaptive night before the avPSG examination due to the organization of the Polish health service and the limited capabilities of the laboratory. The group of nonsmokers could include alcohol drinkers, which may cause bias.Tobacco smokers have a higher bruxism intensity, especially the mixed phenotype, in N1 sleep and in the nonsupine position.Tobacco smokers have a higher number of bruxism episodes with arousal than nonsmokers, which suggests increased sleep fragmentation.Smokers with comorbid sleep-related disorders tend to have electrolyte disturbances and lipid disorders.Alcohol consumption has no significant influence on bruxism parameters.


In recent decades, populations of several game species in the wild have decreased, which has given rise to the creation of game farms that produce birds to be released in, and to restock, hunting preserves. Some game farming sectors are well known and developed, but in Spain, quail game farms have received little attention in terms of research. To address this gap in the literature, we characterised the Spanish quail game farm sector by administering a survey to farmers. We found that complete-cycle farms, of very different sizes, coexist with a majority of farms that do not have breeding flock, and which carry out only the finishing phase of quail raising. These farms mainly produce quails with good flying abilities that are primarily intended to be released for intensive hunting and shooting practices, rather than for restocking hunting preserves. Quails are sold almost all year round, and the farms also offer complementary services to clients, such as transporting birds to their destination, or organising bird releases at the customer’s hunting preserve. Quail game farms use various channels to promote themselves in a competitive market. These game farms originated in Spain five decades ago, which has led to the creation of a consolidated sector.Coturnix genus) game farms were characterised in Spain using variables related to their age, geographical distribution, farmed species, structure, size, and commercialisation, using a survey conducted on 21 farms in 2018. It was found that 38.1% of the sample represented complete-cycle farms, and 61.9%, which have no breeding flocks, undertook only the finishing phase of quail raising. The average size of the breeding flock, with regard to complete-cycle farms, was 1096 males and 3735 females, with a female/male ratio of 3.6, and 75% of these farms carried out self-replacement of breeding quails. The most farmed species was European quail (Coturnix coturnix), followed by Japanese quail (Coturnix japonica), and hybrids of both species. In addition to quail, all farms produced other gamebird species such as pheasant (Phasianus colchicus) and red-legged partridge (Alectoris rufa). The rearing phase, which began when the chicks were one-day-old, lasted 35.3 days on average (range: 25–49 days), with an average stocking density of 47.2 birds/m2 in the brooder house. The finishing phase, which was carried out in flying pens at a stocking density of 9.5 birds/m2, ended when the quails’ average age was 60.5 days. All farms sold quails for release in hunting reserves (EUR 1.54 per bird) and for dog training (EUR 1.65 per bird) for almost 10 months of the year; only 62% sold quails for shooting after being thrown with an adapted clay-pigeon shooting machine (EUR 1.49 per bird). Transporting quails to their destination (95.2% of the farms) and releasing the birds in the client’s hunting preserve (52.4% of the farms) were services offered by the farms, among others. The main advertising and promotional strategies undertaken by the quail game farms to gain a share of the market included maintaining a business website (85.7% of the farms) and attending game and agricultural fairs (47.6% of the farms). In conclusion, this alternative poultry sector has been consolidated in Spain, five decades since the establishment of the first game farms, and it satisfies a relevant part of the demand for quail hunting.Quail ( Coturnix genus  are raised in game farms for hunting purposes, among other uses. In several European countries such as Italy [Coturnix coturnix Linnaeus, 1758), that is autochthonous to Spain, has been identified as a bird species with an unfavourable conservation status [Quails of the as Italy , Greece as Italy , the Repas Italy ,3,4, andas Italy ,6. Put-aCoturnix japonica, Temminck and Schlegel, 1849). Hybrids of both species are also being marketed more frequently, which are usually obtained by crossing female Japanese quails with common quail males, and successive generations are backcrossed with common species males. The latter two types of quail are more easily farmed than the European quail because of their better reproductive performance [The main species of quails that are being farmed for the hunting market are the common or European quail ,10 and tformance ,12,13,14formance ,12,15. Dformance .The game quail farming model ,11,14,17Game quail farming and hunting-based activities are complementary resources for the rural sector ,11 with The study was conducted in Spain by administering a survey to quail game farmers. It followed the same methodology that has been previously applied to other Spanish game farming sectors in order to characterise them ,20,21. SThe information was obtained using a survey carried out in May and June 2018, wherein farmers were directly interviewed. The structured questionnaire included qualitative and quantitative variables  arranged into the following groups: (i) geographic location; (ii) year of establishment; (iii) farm size and breeding flock structure; iv) birds produced per year; (v) reproductive management; (vi) breeding flock replacement practices; (vii) facilities and management during the rearing and finishing phases; (viii) farmers’ knowledge of the types of quail they raise from a genetic standpoint; (ix) game species other than quail raised on the farm; (x) animal products supplied by the farm; (xi) additional services offered by the farm; (xii) market’s geographical area; and (xiii) farm advertising channels.All variables were selected on the basis of a review concerning previous knowledge of the quail game farm subsector ,11,14,23The mean, standard error (SE), minimum and maximum were calculated for (i) the number of females, number of males, and the female-to-male ratio variables in the breeding flock; (ii) the number of housing facilities; (iii) stocking densities; (iv) bird ages during the different breeding and raising stages; and (v) mortalities that occurred during the two raising phases. Regarding the other variables of the survey, the number and percentage of farms showing those particular attributes were calculated. Chi-square tests were performed on contingency tables to analyse the differences between complete-cycle farms and farms that carry out only finishing phase, regarding the variables that were related to: (i) game species other than quail produced by the farm, (ii) additional services, other than the sale of birds, offered by the farm, and (iii) advertising and promotional channels chosen by the farm. Pearson correlations were performed in order to analyse the relationship between stocking density, on the one hand, and the length of artificial lighting, on the other, and mortality during the rearing phase. Moreover, Pearson correlations were also performed to assess the relationship between stocking density and mortality during the finishing phase of quail raising. Statistical analyses were performed using SPSS v.15.0 software .Only 38.1% (n = 8) of the 21 surveyed farms were complete-cycle farms. The remaining 61.9% (n = 13) of farms raised quails by only carrying out the finishing phase of quail-rearing in flight pens. Most of the finishing farms (n = 12) reared quails by buying chicks aged 25 to 60 days old  from other farms, and the remaining one finishing farm bought one-day-old chicks and undertook also the starting or rearing phase in brooder houses. It is relevant to point out that the biggest quail farm in Spain, located in the region of Catalonia, supplied quail chicks for most of the finishing farms . Some of these farmers preferred not to raise their own chicks, starting from own breeding flocks, because they consider the reproduction, incubation, and starting phases of the quail-rearing process to be unprofitable, due to the low market prices of one-day-old chicks.When considering only complete-cycle farms, the average breeding flock was composed of 1096 males and 3735 females; the average female/male ratio was 3.6 .The average farm production was 76,824 ± 30,015.2 quails per year, though there was a great difference between the farm that produced the lowest number of quails (5000 quails) and the farm that produced the most quails, which sold around 580,000 birds per year. The number of quails produced by the latter complete-cycle farm is more than one third of the total number of quails produced by all of the surveyed farms . Therefore, it clearly creates an unbalanced view of the sector because, when this big complete-cycle farm was excluded from the analysis, the average production decreased to 58,200 ± 18,953.6 birds/year for the complete-cycle farms (n = 5) or 45,308 ± 14,565.2 birds/year in the case of the finishing farms (n = 13).The surveys showed that reproductive life of breeding quails begins at an average age of 49 ± 2.3 days old, and it ranges between 39 (minimum) and 60 days old (maximum). The end of the reproductive life of quails occurs at an average age of 10.4 ± 1.07 months old, and it ranges between the ages of 5 and 12 months old. Most of the complete-cycle farms  reported that the reproductive life of quails ends at 12 months of age, whereas the remaining two complete-cycle farms (25%) stated that the reproductive life of quails ends at 5 and 6 months, respectively; these farms were the smallest in terms of size and they produced purebred European quails. Two farms (25%) that raised Japanese quails or the hybrid quails reported that the reproductive life of quails ends at 12 months of age. However, four farms (50%) reported that the reproductive life of their quails ends at 12 months of age, and they raised the European quail species.Most of the complete-cycle farms (75%) carried out self-replacement of their breeding flock, whereas the rest of the farms, in addition to the system of self-replacement, also bought breeding quails from other farms.All complete-cycle farms housed breeding quails in battery cages that were kept indoors. In order to improve laying performance, all complete-cycle farms, except one, used artificial lighting during the laying period; however, each farm adopted different lighting schedules and durations.2, with a minimum of 20 and maximum of 80 quails/m2. The average number of brooder houses was 12.4, and it ranged between one and 38, depending on farm size. The mortality during this phase averaged at 4.4%, and it ranged between 2 and 8%. No correlation was found between stocking density and mortality during the rearing phase in the brooder house . The use of artificial lighting programs during this phase in complete-cycle farms (n = 8) ranged between no artificial light implementation (12.5%) and 24 h of artificial light (62.5%); other farmers supplied 16 h (12.5%) or 12 h (12.5%) of artificial light per day during this period. No correlation was observed between the length of the artificial lighting period and mortality rate during the rearing phase in the brooder house .The rearing phase began when the chicks were one-day-old, and it lasted between 25 and 49 days , and it finished when the quails were sold, when they were an average age of 60.5 days old (range: 45–85 days). The average number of flight pens, where quails were housed during this phase, was 5.2, and it ranged between one and 38, depending on farm size. The mortality during the finishing phase was 1.9%, on average, and it ranged between 1 and 9%. No correlation was found between the stocking density and mortality during the finishing phase in the flight pens .The finishing phase is implemented by housing the quails in open-air flight pens, and it lasted, on average, 21.1 days, and it varied between 7 and 30 days . The avep > 0.05), three quarters of the farms also produced red-legged partridges (Alectoris rufa) or pheasants (Phasianus colchicus) in addition to quails, and half of them produced all three species concurrently (All of the surveyed farms produced game species other than quails. Regardless of whether they were complete-cycle farms or carried out only the finishing phase of quail-raising (urrently .p > 0.05) between farm types (complete-cycle or finishing farms) were found in terms of the services offered.p < 0.1) to be complete-cycle farm than finishing farm.p > 0.05) between complete-cycle farms and farms carrying out only the finishing phase, in terms of promotional and advertising activities.Nearly 90% of the quail game farms had their own website ; websiteThere is little scientific ,25,26,27Although quail farming for hunting purposes began earlier in other countries, such as the USA and Italy , some ofAlternative poultry farming was considered to be an available option for farmers with few economic resources since it enables them to reuse abandoned lands and old facilities  to develRegarding the average size of complete-cycle farms, the data obtained in this study present a broad range of breeding flocks, ranging from one hundred up to 15,000 females . This diThe beginning and end of the breeding quails’ reproductive life reported in this study for the quail game farms are in accordance with the reviewed literature on general quail farming ,34. HoweBreeding quail self-replacement, which is a strategy followed by all quail farms (and that is the only replacement practice in three quarters of the surveyed farms), is also the same self-replacement strategy used in most game farms raising red-legged partridge , pheasan2 [2 [Data obtained for the rearing phase length in the brooder house  are in a2  to 50 qu2 [2 . Nationa2 [2  does not2 [2  for Japa2 [2  for game2 [2 ,39, the 2, thus permitting to improve birds’ physical condition to achieve a vigorous and fast fly and to minimise feather pecking, which devalues birds [Most farms decide the beginning of finishing phase  in accores birds . The morAs releasing Japanese or hybrid quails for restocking purposes is forbidden in most European countries because they are not native species ,11,15, mThe main product offered by all game quail farms is a bird with excellent flying abilities, to be released and shot, mainly in private hunting preserves, throughout the year . In addiAccording to statistical data from MITECO , in 2018Compared with red-legged partridge farms , game quServices supplied to customers by the quail game farms, as evidenced in the present study , coincidCompared with other game farming sectors in Spain (73.0% of red-legged partridge , 64.0% oThe creation and maintenance of a business website appears to be the main promotion channel used by quail game farms in Spain , probablThis research shows that the production of quails in game farms in Spain is a half-century old, and it has become a consolidated sector. Quail game farms vary markedly in terms of size, with one of them producing more than a third of all quails released in Spain, and they have evolved into two differentiated typologies. Some of the farms are complete-cycle farms, which involves breeding flocks, the artificial incubation of eggs, chick rearing in brooder houses, and finally, they are brought to flying pens; however, the majority of farms only carry out the finishing phase of chick raising in the flight pens. The main product is a bird that can be released in hunting preserves for intensive shooting, rather than for restocking. Moreover, it is marketed practically all year round, all over Spain, and several farms export quails to Portugal and France. Many of the farms offer services in addition to selling birds, which mainly involves the transportation of quails to the hunting preserves. The farms use various promotional and advertising channels, mainly their websites, which allows them to compete in a sector that has already matured.


To alleviate the rising mortality burden due to hypertension and other non-communicable diseases, a new public health policy initiative in 2009 called the Basic Public Health Services (BPHS). Program was introduced by the Chinese government. The goal of the study is to assess the feasibility and impact of a nationwide health care service—the “BPHS”.From January to December 2021, a stratified multistage random sampling method in the survey was conducted to select 6,456 people from 8 cities/districts in Yunnan Province, China, who were above the age of 35 years. 1,521 hypertensive patients were previously aware of their high blood pressure status were matched to the BPHS program database based on ID number and then further divided into BPHS group and non-BPHS (control) group. The results of the current study are based on their responses to a short structured questionnaire, a physical examination, and laboratory tests. The association between BPHS management and its effect on the control of hypertension was estimated using multivariable logistic regression models. We evaluated the accessibility and efficacy of BPHS health care services by analyzing various variables such as blood pressure, BMI, lifestyle modification, anti-hypertensive drugs taken, and cardiovascular risk factors.Among the 1,521 hypertensive patients included in this study, 1,011 (66.5%) were managed by BPHS programme. The multivariable logistic regression model demonstrated that the BPHS facilitated hypertension control . A higher proportion of participants receiving lifestyle guidance from the BPHS management showed lowering of total cholesterol. In comparison to the non-BPHS group, those under BPHS management adhered better to antihypertensive medications either single drug (54.3%) or in combination (17.3%) of drugs. Additionally, we also noticed that urban areas with centralized and well-established digital information management system had better hypertension treatment and control.Nearly two-thirds of the hypertensive patients in Yunnan Province were included in BPHS management. The impact of the national BPHS program was evident in lowering risk factors for cardiovascular diseases, promoting healthy lifestyles, lowering blood pressure, increasing medication adherence, and the better control rate of hypertension. The number of people with hypertension increases from 650 to 1.28 billion worldwide from 1990 to 2019 in the age group of 30–79 years , and hypControlling blood pressure to a normal state reduces the risk of cardiovascular events and all-cause mortality –8. The tIn 2009, China's new healthcare reform introduced the “National Basic Public Health Service Program” (BPHS), which provides free of cost health services throughout the country by partnering with community health organizations. The management of hypertensive patients aged 35 and over was one of the 12 kinds of voluntary free services contents in BPHS , 16, incThis study comprehensively evaluated the BPHS hypertension management in Yunnan Province, a relatively economically backward in southwest China. Hypertensive patients who were aware of their high blood pressure from a representative sampling survey and BPHS system matched, to compare matched BPHS group and unmatched non-BPHS groups' coverage and current hypertension status, CVD risk factors, management and effectiveness, blood pressure control rate and antihypertensive drug use, etc. This study supports the World Health Organization's (WHO) suggestion that developing nations should increase their access to management-controlled hypertension-related healthcare services . Rarely The survey was conducted in Yunnan province from January to December 2021, and samples were chosen using a stratified multistage random sampling method. As shown in The target population for BPHS hypertension management is patients who are residents of the jurisdiction aged 35 years and older , so we evia face-to-face short structured questionnaires that included details of their socio-demographics like age, gender, ethnicity, place of residence, educational attainment, occupation, annual household income, and wellness behaviors like smoking, drinking, dieting, exercising, and finally about hypertension perse like the history of hypertension, treatment followed from the time of diagnosis. Additionally, we measured height with RGZ-160 measuring instrument , and body weight was measured with an InBody H20B  removing shoes, hats, coats, or weight in pockets. Body mass index (BMI) is a person's weight (kg) divided by the square of height (m)  .Diastolic blood pressure (DBP) and systolic blood pressure (SBP) were measured in the right upper arm using an OMRON HBP-1,300  at 5 min intervals; three readings were recorded and the average of the three measurements was used for this study. In the meantime, participants were asked to fast for more than 8 h before collecting 8 ml of blood samples for cryopreservation and then sent to Beijing ZhongtongLanbo Medical Test Laboratory for measurement of triglycerides (TG), high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein cholesterol (LDL-C), and total cholesterol (TC). The findings and readings were recorded on the iPad in specially designed survey tables see , and theAccording to the 2018 Chinese Guidelines  and 20202, obesity was a BMI of 25 kg/m2 and above. Risk factors for cardiovascular disease (CVD) as per 2020 ISH Guidelines  were skewed distribution data by Kolmogorov-Smirnov test, described by median and inter quartile range, and the differences between groups were tested by Wilcoxon rank sum test.All statistical analyses use IBM SPSS 22.0 . Categorical variables from the BPHS and non-BPHS groups were presented as numbers (proportions), and differences between groups were compared using the chi-square test or Fisher's exact test. Mean ± standard deviation (SD) and Eleven characteristic variables as age , residence , gender , ethnicity (Han or other minorities), annual household income , education attainment , current smoker (yes or no), current drinker (yes or no), insufficient vegetable and fruit intake (yes or no), insufficient physical activity (yes or no), overweight and obesity (yes or no) were compared between the two groups.p < 0.05 was considered statistically significant.The ggplot2 package of R 4.0.5  was used to construct the visible distribution of blood pressure and a stacked column chart of “controlled with treatment”, “treated but not controlled”, and “diagnosed but untreated”. Unadjusted and full adjusted binary logistic regression models were conducted to assess the association between hypertension control with BPHS, the first model did not adjust for potential influencing factors. Next, second model was performed to adjust for potential influencing factors, including age, gender, ethnicity, residence, household income and educational level. Then, the odds ratio (OR) and 95% confidence interval (CI) were estimated. All tests were two-sided and The mean age of the 1,521 hypertensive patients was 62.6 ± 12.6 years old, with 737 men (48.5%) and 858 people living in urban areas (56.4%), 1,118 Han people (73.5%), 1,104 people (72.6%) with an annual family income of <4,487 USD/year, and 898 people (59.0%) with primary school education or below .p < 0.05). Furthermore, the BPHS group had a lower proportion of current smokers and their consumption of fruits and vegetables was inadequate .1011 (66.5%) and 510 (33.5%) hypertensive patients came from the BPHS and non-BPHS groups. Hypertensive patients included in BPHS management were more likely to be older, female, non-Han Chinese, and had lower family economics and literacy , the average SBP was (146.8±18.8 vs. 149.7 ± 19.1) mm Hg, and DBP (85.7 ± 12.5 vs. 91.2 ± 12.7) mm Hg, respectively . The non-BPHS group showed greater variation in SBP and DBP distribution than the BPHS group (As shown in HS group . FurtherThe BPHS group indicated higher hypertension control rate (33.2%) than the non-BPHS group (22.2%) shown in p < 0.05)  .Further analysis proved that patients with BPHS had lower TC after receiving weight reduction guidance, smoking cessation guidance, and alcohol consumption restriction guidance than those who did not receive such services. In addition, patients who received weight loss, smoking cessation, alcohol consumption restriction, and stress guidance also had lower HDL-C, however within the normal range. No significant differences were observed in TG and LDL-C levels between those who received lifestyle modification services and those who did not .p < 0.001).In A lower blood pressure control rate was observed from male patients aged 65 years and above, especially at Dayao, Xinping, Anning, and Chengjiang counties in rural areas. In general, it was observed that, there was a higher prevalence of hypertension treatment and better control among women than men. Patients with hypertension of any gender who are older than 65 can access antihypertensive medications and have better control over their condition. There is no significant difference noticed between subjects with different income levels among the groups of “diagnosed but untreated,” “treated but not controlled,” and “controlled with treatment.” .This study showed the status of BPHS coverage and the effectiveness in the management of hypertensive patients in Southwest China (Yunnan Province) was corroborated by provincial-specific investigation results. Patients' ID in the survey matched with the BPHS system and found that 66.5% of hypertensive patients were included in BPHS and received related management services. The accessibility of BPHS for hypertensive patients in Yunnan Province was higher than the national accessibility rate . BPHS inUncontrolled hypertension may result in ~24,914,000 years of life lost and 28,657,000 quality-adjusted life years lost in 1.7 million Chinese adults . TherefoThe study also found, in the BPHS group, 71.6% of hypertensive patients used antihypertensive medication (vs. non-BPHS 45.9%), which was higher than the national treatment rate of 61.3% . It may Generally, the control rate of hypertension is the standard for evaluating the effectiveness of interventions, resulting in effectively preventing or delaying the occurrence of stroke, myocardial infarction, heart failure, renal insufficiency, and other complications . Our stuLimitations include the design method in this survey, this might not substantiate causation, particularly the link between the services offered by the BPHS program and the management of hypertension without these services. Recall bias can also be a challenge with self-reported data. Some subjects lacked information, such as height or weight, which can skew the data. Therefore, information collection and interpretation of risk factor changes should be conducted with utmost care.Since its launch in 2009, the BPHS program run by provincial authority healthcare providers has made a substantial contribution to bettering hypertension management and balancing access to healthcare across inhabitants of urban and rural areas as well as among various socioeconomic strata.Results showed that program participants kept healthy lifestyles, had lower blood pressure, higher control rate of hypertension, and took their medications more consistently, which could account for some of the benefits of this program. In districts or counties with fully operational digitalized and centralized data management systems, it was easier to include the participants compared to “non-matched ID” in rural areas, therefore health information management may further be improved to address this limitation in rural areas. In order to better diagnose and treat hypertension at its early stages, a customized program has to be designed to cater to the requirements of hypertension control in the younger population through the BPHS program.The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.The studies involving human participants were reviewed and approved by the central Ethics Committee at the Fuwai Hospital, CAMS/National Center for Cardiovascular Diseases approved this project . The survey was conducted in accordance with the ethical principles of the Declaration of Helsinki, and the informed consent and electronic signature of the participants were obtained before the investigation. The patients/participants provided their written informed consent to participate in this study.LP and LK were co-first authors and drafted the manuscript. ZG and YZ initiated, conceived, and supervised the study. LD, WL, and MF guided the analysis and modified the article. LP, JF, and HF were involved in data curation and analysis. YS and MT completed field execution and coordinated all divisions. XW, MM, and HW checked the integrity of the data. LK and DM reviewed and edited the manuscript. LD conceptualized and supervised this project. All authors have read and approved the final version of the manuscript.This work was supported by Key Research and Development Program from Yunnan Province Science and Technology Department (Grant No. 202103AF140002); Yunnan Provincial Clinical Research Center for Cardiovascular Diseases-New Technology Research and Development Project for Diagnosis and Treatment of Major Cardiovascular Diseases (Grant No. 202102AA310002); and Provincial Innovation Team Project of Heart Failure Diagnosis and Treatment in Fuwai Yunnan Cardiovascular Hospital (Grant No. 202005AE160020).The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. 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