Data validation shows that the ice phenology data derived by our method is highly consistent with that based on existing approaches . The dataset is valuable to investigate the lake-atmosphere interactions and long-term hydrothermal change of lakes across the Tibetan Plateau.The Tibetan Plateau (TP) is a region sensitive to global climate change and has been experiencing substantial environmental changes in the past decades. Lake ice phenology (LIP) is a perceptible indicator reflecting changes of lake thermodynamics in response to global warming. Lake ice phenology over the Tibetan Plateau is however rarely observed and recorded. This research presents a dataset containing 39-year (1978–2016) lake ice phenology data of 132 lakes (each with area >40 km  Change of lake ice phenology in the Cryosphere is considered as one of the most reliable pieces of evidence reflecting global warming5. It could subsequently lead to substantial alteration in biogeochemical processes . Research efforts on investigating changes of lake ice phenology are mainly based on ground observation, satellite observation or modelling10. Most of the research, however, are for lakes locating in North America or Northern Europe as there are relatively abundant in ground observations10.The formation and duration of ice cover for lakes in the Cryosphere plays an important role in thermodynamics of lakes. It interacts with lake water temperature and shapes aquatic ecosystems as well. Lake ice phenology  are closely related to temperature variationet al. (2021) reported that 41% of inland water area around the globe were seasonally frozen11. Though with uncertainties12 from both data sources15 and extraction methods, various satellite-based observations have shown their merits in tracking the freezing-thawing cycle of lake ice cover16. The most commonly used satellite data includes that from the optical sensors like Advanced Very High Resolution Radiometer (AVHRR)17 and Moderate-resolution Imaging Spectroradiometer (MODIS)18, the passive microwave sensors like Scanning Multichannel Microwave Radiometer (SMMR)3, the Advanced Microwave Scanning Radiometer for EOS (AMSR-E)19 and Special Sensor Microwave/Imager (SSM/I)20. Active microwave also shows capabilities in monitoring lake ice status based on sensors like European Remote Sensing Satellite (ERS)-1/2 synthetic aperture radar (SAR)21 and Radarsat-1/2 SAR22, but the narrow swath width and the relatively low temporal resolution limited the application of the technology to monitor lake ice phenology at a daily scale and for a large area. It is also noted that the satellite-based observations of lake ice phenology mostly start from the late 1990s and could be with considerable data gaps. For example, the currently available dataset for lake ice phenology in the TP is that derived from microwave brightness temperature measured by AMSR-E, the Advanced Microwave Scanning Radiometer 2 (AMSR2) and Micro-Wave Radiation Imager (MWRI). The dataset however is limited to 51 lakes on the TP covering the period from 2002 to 201523.Lake ice phenology observations are not always widely available particularly for regions like the Tibetan Plateau (TP), where ground-based observation is challenging and costly. Observation from space therefore has been becoming more attractive in investigating the lake ice phenology. For instance, with stratified random sample from Sentinel-2 and PlanetScope, Pickens 24. The model-based approach goes beyond reproducing data on lake ice phenology and can be used to quantify the response of lake ice phenology to climate change or climate variation. The models may cover different level of details on lake thermodynamics, subject to their research focuses29. It is recognized that a well simplified process-based model (like air2water31) could perform as good as a more complicated model  if the research interest is mainly on the timing of freezing and thawing of lake ice. The simplified model could be more competitive and applicable for regions with little in-situ observations (like the TP) as it requires much less data for running and calibration.In the literature of limnology, numerous research efforts also have been invested on developing mathematical models to reconstruct lake ice phenology for the historical period or to predict the response of lake ice phenology to a warming climate32 is applied to calibrate and validate the numerical model in reproducing the consecutive daily surface water temperature for the studied period, based on which algorithm has been developed to determine the ice phenology indices (including dates of freezing-up and breaking-up and durations). Lake ice phenology derived directly from satellite observations in this research and by other researchers is used to validate the reconstructed LIP33.The dataset herein provides complete, consistent and continuous time series 1978–2017) of reconstructed ice phenology for 132 lakes in the TP . The daily air temperature from the dataset is only available at 31 meteorological stations located in the TP. The spread and density of the meteorological stations are big constraints for earth system research in the region. We noticed that quite a few research efforts have been invested to provide reliable reanalysis climate datasets for the TP . The public available climate reanalysis datasets however are found not necessarily of high good quality for our simulation when we compare them against the observations from the meteorological stations. Hence, we use the air temperature interpolated from the nearest station with altitude adjustment (by a lapse rate of 0.65 °C 100 m−1)34 to provide air temperature input to drive the modified air2water model.The modified air2water model requires solely air temperature as input. For the TP, the best publicly accessible ground-based observation of daily air temperature is that from the dataset of daily climate data from Chinese surface stations (V3.0) provided by China Meteorological Data Service Centre of National Meteorological Information Centre . The surface water temperature of a specific lake is the lake-wide mean temperature based on all pixels within the lake. Boundaries of the lakes (shapefiles included in the dataset) are mainly from National Tibetan Plateau Data Center  and are cross-checked by maps from HydroLAKES (http://www.hydrosheds.org/)35 and Google Earth. The lake-wide MODIS-based surface water temperature is the mean of daytime and night-time LST in the MOD11A2. All the data processing procedures are implemented via Google Earth Engine (GEE)36.The direct output of the model is lake surface water temperature. Hence, the model is firstly calibrated and validated against the satellite-based observation of lake surface water temperature deriving from the MOD11A237. The MOD10A1 dataset is a snow cover product with a spatial resolution of 500 m containing daily, gridded snow cover and albedo derived from radiance data acquired by MODIS on board the Terra satellite. Snow cover is identified using the Normalized Difference Snow Index (NDSI) and a series of screens designed to alleviate errors and flag uncertain snow cover detections37.Prior to determine the lake ice phenology given the modelled lake surface temperature, temperature thresholds should be defined, which is done herein with the aid of remotely sensed snow cover product, i.e., MOD10A128 is a semi-physical model designed to simulate lake surface temperature principally based on lake surface energy balance expressed as:Tw is water surface temperature at time t (day step in this study), Hnet is the net heat flux per unit surface, A is the surface area of the lake, Vs is the volume of water involved in the heat exchange with the atmosphere, ρ and cp is the density and specific heat capacity of water. The model considers all contributions in the energy balance (Hnet) as a function of air temperature or is included in the parameters of the model. The original air2water model has simplified the lake energy balance by introducing eight calibratable parameters a1–8 with air temperature as the only required input, which can be expressed as:Ta is air temperature, ty is days of a year. δ = D/Dr is normalized well-mixed depth, where D is the depth of well-mixed surface layer (the epilimnion thickness), Dr is the maximum thickness. Th is the deep water temperature assumed to be 4 °C.We used a slightly modified air2water model to simulate the daily lake surface temperature for the period 1978–2017. The air2water39. However, the original air2water is limited to simulate the surface temperature of open water, which results in difficulties for applications in lakes with long ice cover duration. For this sake, we assume that when the lake is completely covered by ice, the heat exchange between air and water is blocked and surface energy balance would be replaced by land surface energy balance without mixing depth in water36, which indicates δ = 1 and Eq.  can be expressed as:The model has been tested efficient in different regions40. The objective function of model calibration is the root mean square error (RMSE). To ensure the rationality of the calibrated model parameters, a physically consistent priori range are assigned to each parameter31 and Dr is bounded by the average depth of the lake obtained from Records of Lakes in China41 and HydroLAKES35. The calibration period is 2000–2012, while 2013–2017 is for validation.The model is calibrated against the LSWT derived from MOD11A2 using the PSO (Particle Swarm Optimization) approachRwater) to total lake area is firstly calculated as: Nwater is the pixel number of “inland water”, while Nice is the pixel number of “ice”. The lake is assumed to be open when Rwater > 90% and frozen when Rwater < 10%43. As shown in Fig. Rwater) drops down below 10% or rises above 90%. The multi-year medians of the temperature thresholds are finally considered as the overall thresholds to determine the starting and ending dates of freezing-up (TFUS and TFUE) and breaking-up of lake ice phenology (TBUS and TBUE) separately for each lake.The lake surface temperature output from air2water model is used to derive from lake ice phenology given prior defined temperature thresholds. The temperature thresholds are defined with the aid of the snow cover products (MOD10A1), where the MOD10A1 images with cloud proportion over the lake exceeding 20% are neglected. For each lake, the ratio of “inland water” . Since the lake ice phenology could spans two consecutive years, we assign herein the value of t to range from 1 to 730, where t>365 represents the date in the next consecutive calendar year.The long-term lake ice phenology for the period 1978–2016 is derived from the simulated lake surface temperature given the temperature thresholds. The lake ice phenology considered herein consists of 6 indicators reflecting the timing or duration of different freezing/thawing phases. In addition to starting and ending dates of freezing-up (FUS and FUE) and breaking-up (BUS and BUE), the ice-covered duration  and the fully ice-covered duration  are also presented in the dataset. The indicators are defined as:44 via the link: 10.6084/m9.figshare.18852338.v2. Description about the 132 lakes is shown in the lake_info.csv file, including lake ID, name, latitude, longitude, lake area, water level, depth, water volume, freezing property, and glacial replenishment. The information about the lakes is mainly collected from Records of Lakes in China41, HydroLAKES35, and Map of Glaciers and Lakes on the Tibetan Plateau and Adjoining Regions. The Phenology folder in the dataset consists of 132 CSV files. Each CSV file is named according to the lake ID presented in lake_info.csv. The columns of each CSV file are listed in Table The lake ice phenology data across Tibetan Plateau is archived and openly accessible via Figshare45 above 0.6) when the simulated lake surface temperature is compared against the MODIS-based one. Suspicious outliers  are eliminated from satellite-based lake surface temperature and snow product in providing reliable data for robust model calibration and validation. The MOD10A1 is used as an aid in determining the temperature thresholds of LIP. It is noted that the MOD10A1 is not always of good quality in continuously retrieving the ice-covered ratio of a lake due to accidental misclassification and effects of cloud cover. To alleviate the uncertainties induced by the optical RS data, we use open water ratio (see Methods) instead of directly using ice-covered ratio of a lake in determining the temperature threshold, and the medians of the temperature thresholds are finally accepted for robust lake ice phenology identification. The reconstructed LIP dataset is validated by comparing against the published LIP data as described below.The reconstructed LIP dataset is produced with strict quality control throughout the procedures. The boundaries of the lakes are examined via professional GIS tools (both ArcGIS and Google Map) to ensure the consistency of their geographic locations, and topography. Lake-wide mean surface temperature instead of that of lake centroid is derived from MOD11A2 for model calibration. The 132 lakes presented in the dataset are all with good model performance  of the estimated FUE in Nam Co is around 7 days, RMSE of the estimated BUE in Lesser Slave Lake is near 9 days and RMSE of the estimated in Qinghai Lake is around 12 days. It should be noted that the bias could be due to both the uncertainties in the model and in the climate input to the model as well.There is little et al.32 and Qiu et al.23 respectively. The dataset published by Cai et al.32 is retrieved from MOD10A1 and Landsat data, providing mean lake ice phenology of 44 TP lakes for the period 2000–2017. Figure et al. for the 44 lakes. It can be seen that the LIP dataset produced by our approach is in high agreement with that solely based on MOD10A1, where the R2 for all the four phenology indices  are higher than 0.96 and the root mean squared errors  are all within 5 days.We further compare the reconstructed LIP against two existing datasets developed based on remotely sensed data, which were published by Cai et al.23, which was derived from microwave brightness temperature measured by AMSR-E, AMSR2 and MWRI. The dataset from Qiu et al. provides LIP of 51 lakes in High Asia for the period from 2002 to 2016, among which there are 35 lakes identical to the lakes in our dataset. For the 35 lakes in the period 2002–2015 (sample size = 35 × 14), the statistically significant correlations are found between the two datasets for all the six phenology indices with R2 all above 0.75  or from 85 to 255 days (IceD). The latitude gradients of CID and IceD are 17.5 days/degree and 15.2 days/degree respectively. Lake ice phenology is also affected by altitude (Fig. Figure Supplementary of Arcticle


Introduction: Bioactive molecules from natural sources having contraceptive properties were excellent alternatives for modern hormonal contraceptives. Researchers around the world were working on identifying contraceptive leads targeting the male reproductive system rather than the usual female contraceptives. The lack of proper understanding on male contraceptive protein drug targets leads to insufficient evidence on activities of identified contraceptive compounds. The proteins specific to the male reproductive system and involved in sperm-egg fusion will be an excellent drug target to identify the male non-hormonal, reversible contraceptive leads. Inhibiting sperm hyaluronidase activity by natural non-hormonal compounds will lead to reversible and non-hormonal male contraception. The Aegle marmelos Linn. is one such important medicinal plant with valuable phytocompounds, used traditionally as a potential contraceptive measure. The in vivo experiments on leaf extracts of Aegle marmelos. Linn containing terpenes, sterols, and alkaloids shows prominent contraceptive activities. Moreover, this study explores the potential ability of the leaf extract on inhibiting the sperm hyaluronidase action with additional molecular details on the interaction between sperm hyaluronidases and three phytocompounds such as aegeline, marmin, and marminol.Material and methods: The in vitro hyaluronidase inhibition assay and Computer Assisted Sperm Analysis (CASA) were used to evaluate the male contraceptive properties of the Aegle marmelos Linn. leaf extract. To identify the interaction profile of aegeline, marmin, and marmenol on sperm cell hyaluronidases the in-silico methods such as molecular docking, Non-Covalent Interaction analysis, Molecular dynamics, and Molecular Mechanics Poisson Boltzmann Surface Area were used.Results and discussion: The results of in vitro hyaluronidase inhibition assay and Computer Assisted Sperm Analysis shows the inhibition of hyaluronidase enzymatic activity and reduced sperm activities in the presence of leaf extracts. After incubation with leaf extracts for about 30 minutes time intervals show, the motility drops from progressive to non-progressive and ended up with complete immotile in 100 μg/ml concentration of leaf extract. The results of molecular docking, Non-Covalent Interaction analysis, Molecular dynamics, and Molecular mechanics Poisson Boltzmann Surface Area show that the phytocompounds marmin, and aegeline have the potential ability to inhibit sperm hyaluronidase. After 30 min the test solution-treated semen samples were carefully placed over cover slides and analyzed for sperm motility changes using a Magnus MX21I LED Binocular microscope. The live changes in the motility of the sperm cells treated with AMLE with respect to untreated sperm cells were monitored using the commercial version of the MMC  software package . Since the hyaluronidase enzyme used in this assay was bovine testicular hyaluronidase, the results bring maximum probability similar to human sperm hyaluronidase inhibition. The assay was performed based on the modified protocol given in the Sigma Aldrich hyaluronidase inhibition assay manual  plots were used to specify the hydrogen bonding, van der Waals interactions, and steric effects that arise within the hyaluronidase—inhibitor complex. Finally, the QTAIM analysis was used to confirm the strong and weak hydrogen bonding between the amino acids of hyaluronidases and phytocompounds. Multiwfn software version 3.3.1  was usedhttps://simlab.uams.edu/index.php)   and ligadex.php) . All theThe MM-PBSA calculations were carried out using g_mmpbsa version 5.1.2 tool  which waThe computer-assisted sperm analysis method was used to investigate the contraceptive properties such as affecting the motility of the sperm cells and involvement in remodeling the morphology of sperm cells. To ensure the reversible contraceptive nature of the plant extracts, the vitality of the immotile cells was evaluated. The minimum inhibitory concentration (MIC) for plant extracts of different concentrations diluted in DMSO to inhibit sperm motility was 100 μg/ml. To ensure the effectiveness of the leaf extracts without DMSO the extracts are diluted in a saline medium and the semen samples of healthy individuals were incubated with and without the plant extract for 5 min of incubation time. The results show a considerable decrease in motility parameter values such as VCL, VSL, VAP, BCF, and MAD when compared to semen samples without leaf extracts . The speThe results of the hyaluronidase inhibition assay using bovine testicular hyaluronidase show that complete enzyme inhibition was achieved for 3000 μg/ml of leaf extracts. The enzyme inhibition begins at 3 μg/ml (26%) but there was no increase in enzyme inhibition percentage for 30 μg/ml (26%). The increase in inhibition starts at 300 μg/mL of an extract with 34 percent of enzyme inhibition. The results show that the concentration of plant extracts required for the minimum inhibition was 30 μg/ml and 3,000 μg/ml for complete inhibition of enzyme activity .The aegeline, marmin, and marmenol are docked in the binding site surrounded by catalytic and substrate binding residues of human sperm hyaluronidases. The binding site residues were chosen based on literature studies and sequence alignment of human sperm hyaluronidases with human hyaluronidase-1 involved in tumor angiogenesis . The resThe docking results of aegeline, marmin and marmenol with hyaluronidase-2 show the aegeline-HYAL-2 complex has the least binding affinity of −7.0 kcal/mol with the highest number of hydrogen and hydrophobic interaction in the active site cleft. The aegeline interacts with active site residue Glu 135 through hydrogen bond interaction of 2.8 Å and interacts through hydrophobic interaction with substrate binding residues Asp 133, Tyr 206, Tyr 253, Trp 328 . The resExploring the van der Waals interaction energy, the average binding energy of Hyaluronidase-aegeline, marmin, and marmenol complexes reveals there was a string interaction of marmin with the Hyal4 enzyme. From Based on the results of molecular docking the molecular dynamics for hylaluronidase-2- aegeline, hyaluronidase-3-marmenol, hyaluronidase-4-marmin, and hyaluronidase-ph20- marmin complexes were simulated for 100 ns . AnalyziA. marmelos Linn., leaf extract show the leaf extract creates temporary inactivation of sperm activity and changes in head morphology. The changes in the motility of the sperm cells due to leaf extracts were due to changes produced in the cytoskeletal and sperm head structure. The hyaluronidase inhibition assay also supports the CASA results of plant extracts. The results of molecular docking with sperm hyaluronidase show the compounds aegeline and marmin have a high capability of inhibiting the enzyme activity when compared to apigenin a natural hyaluronidase inhibitor. The molecular dynamics results such as complex RMSD, backbone RMSD and RMSF values confirm that the aegeline, marmin complex has a high probability to produce enzyme inhibition. The NCI and MM-PBSA results are key results in establishing the potential ability of marmin and aegeline to bind with hyaluronidase isoforms. Especially, marmin shows prominent results in all four insilico characterizations, where the marmin-HYAL4 complex shows the least binding values among all in MM-PBSA results and this result supports the NCI topological parameter results where the marmin-HYAL4 has strong interactions. The above results are based on the studies targeting the hyaluronidase enzyme in the active and substrate-binding sites, which ensures that the results are more reliable in the terms of the potential ability of AMLE and its compounds in enzyme-inhibiting activity. Further, future studies to understand the effect of marmin and aegeline in sperm cells will be carried out.The results of CASA studies on the effect of A. marmelos. Linn was analyzed and the results from the CASA analysis of motility parameters, vitality, and morphological characterization show leaf extracts of A. marmelos. Linn has the potential ability in inhibiting the sperm vital parameter necessary for male fertility. The sperm vitality results confirm the A. marmelos. Linn leaves and their compounds have reversible contraceptive effects on sperm cell activity. The hyaluronidase inhibition assay and in silico analysis such as molecular docking, molecular dynamics, MM-PBSA, and NCI analysis confirms that the marmin and aegeline have the potential ability to inhibit the enzyme activity. The results also implicate sperm hyaluronidases as a potential target for male contraceptive drug development. Further research in the future will focus on analyzing the effect of these compounds on hyaluronidase and other important enzymes essential for sperm activity by understanding the protein-ligand dynamics.The male contraceptive effect for leaf extracts of


Climate is currently warming due to anthropogenic impact on the Earth’s atmosphere. To better understand the processes and feedbacks within the climate system that underlie this accelerating warming trend, it is useful to examine past periods of abrupt climate change that were driven by natural forcings. Glaciers provide an excellent natural laboratory for reconstructing the climate of the past as they respond sensitively to climate oscillations. Therefore, we study glacier systems and their behavior during the transition from colder to warmer climate phases, focusing on the period between 15 and 10 ka. Using a combination of geomorphological mapping and beryllium-10 surface exposure dating, we reconstruct ice extents in two glaciated valleys of the Silvretta Massif in the Austrian Alps. The mountain glacier record shows that general deglaciation after the Last Glacial Maximum (LGM) was repeatedly interrupted by glacier stabilization or readvance, perhaps during the Oldest Dryas to Bølling transition (landform age: 14.4 ± 1.0 ka) and certainly during the Younger Dryas  and the Early Holocene . The oldest landform age indicates a lateral ice margin that postdates the ‘Gschnitz’ stadial (ca. 17–16 ka) and predates the YD. It shows that local inner-alpine glaciers were more extensive until the onset of the Bølling warm phase (ca. 14.6 ka), or possibly even into the Bølling than during the subsequent YD. The second age group, ca. 80 m below the (pre-)Bølling ice margin, indicates glacier extents during the YD cold phase and captures the spatial and temporal fine structure of glacier retreat during this period. The ice surface lowered approximately 50–60 m through the YD, which is indicative of milder climate conditions at the end of the YD compared to its beginning. Finally, the third age group falls into a period of more substantial warming, the YD–EH transition, and shows discontinuous glacier retreat during the glacial to interglacial transition. The new geochronologies synthesized with pre-existing moraine records from the Silvretta Massif evidence multiple cold phases that punctuated the general post-LGM warming trend and illustrate the sensitive response of Silvretta glaciers to abrupt climate oscillations in the past. This sensitivity is evident in the accelerating deglaciation of alpine regions caused by rapid warming due to increasing greenhouse gas emissions in recent decades3. Deglaciation affects mountain regions in various ways; ice retreat alters the hydrological regime in these areas and downstream, impacts ecosystems, and increases the frequency of natural hazards4. Improving our understanding of interactions between the climate system and the cryosphere in the past helps project the magnitude and impact of environmental change in the future.Mountain glaciers are highly sensitive to climate variations, most importantly to changes in summer temperatures and to a lesser extent to changes in precipitation6 enable us to explore the climate system and how it operates with and without anthropogenic impact. Detailed geomorphological mapping and direct dating of former ice margins allow us to reconstruct glaciers across time and space and to draw conclusions about climate conditions that drove glacier advance or retreat. Here, we present ice-margin reconstructions of two glaciated valleys in the Austrian Alps, Jamtal and Fimbatal, covering the Late Glacial (LG) and the Early Holocene (EH)—a period when the climate was transitioning from a glacial to an interglacial mode.Glaciers and their shaping of many parts of the Earth’s surface in the past7. Other periods of climate transition before and after the YD remain more controversial in terms of glacier extents and ice dynamics, for instance, the Bølling–Allerød period  and the EH (ca. 11.7–9 ka). Although glaciers were presumably much more extensive at the beginning of the B–A compared to the beginning of the EH, they were driven by a similar climatic pattern during both periods: an abrupt temperature increase which led to rapid deglaciation. For the period between ca. 12 and 10 ka, moraine chronologies from different places in the Alps, including data presented in this study, have shown that ice retreat and therefore warming was not linear but interrupted by centennial-scale cooling11. For the (pre-)Bølling period, it is still unclear whether ice retreat was steady and of such magnitude that most glaciers in the European Alps retreated to the highest cirques or disappeared altogether7. The objective of this study is to contribute to these less resolved periods and to view climate transitions from glacial to interglacial conditions through the lens of glacial geomorphology. Therefore, we mapped moraine sequences at two alpine valleys, the Jamtal and the Fimbatal, and applied 10Be surface exposure dating to selected landforms to produce direct spatial and temporal information of former ice extents in the Silvretta Massif.Late Glacial moraines in the European Alps, especially those deposited during the Younger Dryas  termed ‘Egesen’ moraines, are subject to numerous geochronological studies that provide valuable insights into this last phase of prolonged cooling before Holocene warming12. Lithologies in the valleys include amphibolites, different types of metasediments, and gneisses13. Glaciers in the region are temperate and are sensitive to climate oscillations  and Fimbatal n = 6). Ages are stratigraphically in order and are presented from old to young, beginning with the Jamtal (JAM) record, followed by the Fimbatal (FMB) record. . Ages ar10Be surface exposure dating is J7 at an elevation of ca. 2520 m a.s.l.  valley flank that features a right-lateral moraine set that was shaped by a former tributary glacier  deposited at ca. 2040 m a.s.l. Fig. . The out29. For reference, the LIA terminal moraine in this valley is located at an elevation of ca. 2520 m a.sl., i.e,. around 500 m higher and almost 7 km upstream of F5a/b  is preserved on both sides of the creek and exhibits a blocky structure in its frontal section caused by the outwash of fine-grained material. Exposure ages of the four boulders along F5a range between 11.9 ± 0.2 ka and 9.1 ± 0.2 ka. The age of FMB-18-09 (9.1 ± 0.2 ka) falls within a period during the Holocene when most glaciers had probably retreated inboard the subsequent LIA ice marginBoulder ages in both valleys fall into three periods: (1) the Oldest Dryas to Bølling (J7), (2) the Younger Dryas , and (3) the Early Holocene (J3–4), and are discussed in this order.The deposition of boulders featured by landform J7 (14.4 ± 1.0 ka) occurred during a period when regional climate was transitioning from stadial to interstadial conditions, i.e., from the Oldest Dryas cold phase to the Bølling warm phase. We review the deglaciation of the European Alps in the millennia before the exposure of J7 boulders to place the new moraine records at Jamtal and Fimbatal in a coherent temporal and spatial context of the Alpine LG.30. This phase of readvance that is often associated with Heinrich event 1 in the North Atlantic31 was followed by a period of still relatively low mean annual temperatures but slightly increasing summer temperatures paralleling increasing solar insolation  glacier by the main  glacier Bølling moraines.Since the hypothesis of glacier stagnation after the Gschnitz and before the YD stadial has been proposed and discussed in the past in the context of the European Alps10 investigated an ice margin indicative of glacier extents that exceed the Egesen extent. They obtained a  age of 14.4 ± 0.7 ka (their sample VBA-7) for a boulder sitting on top of a bedrock section that deglaciated at the same time. The authors interpret the site as an LG ice margin of the Great Aletsch glacier. Böhlert, et al.38 yielded a  age of 15.2 ± 1.9 ka for a boulder (their sample VM7) embedded in a lateral moraine outboard the presumable Egesen moraine at Val Mulix (Switzerland) and ascribed the landform to the Daun stadial. Rolland, et al.39 combined the analysis of proglacial lake sediments in the Argentera-Mercantour Massif with exposure dating of glacial features in the region and identified a LG moraine that was deposited ca. 14.6 ± 0.9 ka . These dated boulders and landforms, albeit limited in number, are in good agreement with the age of J7 and suggest moraine formation between ca. 16 and 14 ka in the Alps.In a geochronological study at the Great Aletsch glacier, Schindelwig, et al.55. Its onset around 14.6 ka is characterized by an abrupt temperature increase that induced substantial changes in environmental and vegetational conditions57. Summer temperatures increased by several degrees in the Alps54, transitioning from cooler stadial to warmer interstadial levels 56, and perhaps also to a cold spell, a few centuries earlier suggested by a potential J7 moraine.Considering the uncertainty calculated for the age of J7 (14.4 ± 1.0), the landform could also have been deposited at the onset of the Bølling interstadial, during the Older Dryas cold snap (ca. 14 ka), or during the subsequent Allerød interstadial (until 12.9 ka). However, we emphasize that the most likely age of deposition is 14.4 ka, hence the earliest part of this warm phase Fig. a. The B–62. Moraine formation as a result of this teleconnection on the LG to Holocene time scale has recently been proposed by Young, et al.63 for the Greenland ice sheet, by Protin, et al.8 for mountain glaciers in the French Alps, by Braumann, et al.9 for the Austrian Alps, and by Jomelli, et al.64 for glaciers in the tropical Andes and the greater North Atlantic region. Comparing the warming during the pre-Bølling to Bølling transition and during the YD–Holocene transition , a millennial-scale cold phase documented in Antarctic ice cores79. The extent to which this cold phase propagated from Antarctica and the Southern Hemisphere further to the North remains controversial, but we note that moraine formation indicated by J7 falls within the early phase of the ACR.Both scenarios—moraine deposition prior to or during the Bølling—are plausible, but more direct age data covering that period are needed to better constrain moraine deposition and thus cooling between 16 and 14 ka in the European Alps. Interestingly, when comparing the (pre-)Bølling moraine age of J7 with mountain glacier records beyond the Alps, we find similar intervals of moraine formation along (former) ice sheets and in alpine catchments in Norway80.The next LG time slice that is captured by the new moraine chronologies of both valleys is the YD period. We interpret moraines J6 and J5 at Jamtal and F5a and F5b at Fimbatal as Egesen moraines that portray the fine structure of glacier advance or stabilization during this final stadial before Holocene warming Fig. e. The J681. Glacier retreat through the YD with intermittent phases of glacier stabilization is observed in the European Alps83 and in other glaciated regions of the Northern Hemisphere84, and coincides with slightly increasing summer temperatures  to J5 (2385 m a.sl.) within a few centuries glaciers and address the resilience of mountain glaciers under rapidly warming climate conditions. The response of Silvretta glaciers to natural warming during recent geological periods shows their sensitive response to climate change and places the magnitude and impact of anthropogenic climate forcing in a natural context.The A comparison of moraine records of ice sheets and alpine glaciers on a global scale suggests that glaciers in both hemispheres deposited moraines around 15–14 ka, during the YD, and during the EH. The next step is to investigate whether these similarities are coincidental or due to large-scale climatic forcing.80 developed a relative moraine stratigraphy for the region this work is based on. The pre-existing geomorphological maps were updated and supplemented with information gained during several field campaigns in the summers of 2018 to 2021, from remote sensing data92 and drone imagery. Glacier reconstructions of Jamtal glacier for the Holocene, including corresponding maps, are presented in Braumann, et al.9. This study extends the Jamtal glacier chronology into the LG and focuses on landforms that evidence glacier oscillations during this period by mapping and dating ice margins outboard the Holocene moraines. Reconstructions of paleoglaciers are complemented with glaciological data of modern, annual- to decadal-scale glacier change, including observations of glacier mass balances, front variation, and ice-covered area94.Hertl10Be begins according to the nuclide-specific production rate (ca. 4 atoms g−1 year−1 at sea level and high latitude). The longer a surface has been exposed, the more 10Be has accumulated95. Thus, the nuclide content in rock surfaces is a function of exposure time. This principle is used in the application of surface exposure dating to glacial landforms. A moraine boulder sampled for 10Be analysis has ideally been eroded from bedrock beneath the glacier, has been transported sub- or englacially, and has finally been deposited at a stable ice margin. It has not been exposed to cosmic radiation prior to its deposition along the ice margin, so its radionuclide inventory is ‘zeroed’. If these assumptions are valid, the 10Be inventory measured in a rock surface after exposure will produce an age that reflects the boulder’s melt out of glacial ice, or in other words, the onset of ice retreat, hence warming.When fresh quartz-containing rock surfaces are exposed to cosmic radiation, the production of the cosmogenic radionuclide Rock samples were collected using an electric saw, and hammer and chisel. The sample location was measured using a hand-held GPS device. Strike and dip angles of sampled rock surfaces were quantified using a geological compass. Samples were preferably collected from boulder tops at windswept locations to minimize shielding effects due to snow and/or sediment cover. We avoided surfaces affected by exfoliation and prioritized boulder locations with striations preserved on the surface, which indicates low postglacial erosion. For more details on our sample selection criteria, we refer to Braumann, et al.’s supplement, their Table 96. Purified quartz yields, the target mineral for 10Be extraction, ranged from 1.1 % to 11.3 % (Table 9Be carrier (#7) made of deep-mine beryl, which has a concentration of approximately 1000 ppm. The extraction of 10Be was accomplished following the LDEO protocol97. Isotope ratios in samples (10Be/9Be) were measured at the Center for Accelerator Mass Spectrometry (CAMS) facility, Lawrence Livermore National Laboratory (LLNL) using the 07KNDSTD3110 standard with a 10Be/9Be ratio of 2.85 × 10–1225.Mechanical sample preparation was accomplished at the Department of Lithospheric Research of the University of Vienna and at the Lamont-Doherty Earth Observatory (LDEO). Whole-rock samples were crushed to grain sizes between 63 and 500 µg using a mill or a jaw crusher. Mineral separation strategies included magnetic separation, boiling with phosphoric acid, froth flotation, density separation and repeated (> 3) leaches with hydrofluoric acid and nitric acid at concentrations between 1 and 5 % % Table . Samples % Table  with qua2 test) were performed using the online calculator formerly known as the CRONUS–Earth online calculator v326. We used the local ‘Swiss’ production rate27 and chose the time-dependent ‘Lm’ scaling scheme98. In the data presentation and discussion, we distinguish between exposure ages of individual boulders and moraine ages. Boulder ages are reported with 1σ analytical uncertainties and a 1 % error on the carrier concentration but without uncertainties on the production rate as this source of uncertainty is constant for all boulders sampled from the same area. For moraine ages (n ≥ 3), the production rate error is propagated in quadrature to analytical and carrier uncertainties to allow the correlation of landform ages with moraine records from other regions.Exposure age calculations including statistical outlier identification (ΧSupplementary Information.


Among the E3 ubiquitin ligase families, the cullin-RING E3 ubiquitin ligase (CRL) family is the largest. Cullins are molecular scaffold proteins that recruit multiple subunits to form the CRL complex. We hypothesized that a CRL complex mediates the cGMP-dependent increase in NKCC2 ubiquitination in TALs. Cullin-1, cullin-2, cullin-3, cullin-4A, and cullin-5 were expressed at the protein level, whereas the other members of the cullin family were expressed at the mRNA level, in rat TALs. CRL complex activity is regulated by neuronal precursor cell-expressed developmentally downregulated protein 8 (Nedd8) to cullins, a process called neddylation. Inhibition of cullin neddylation blunted the cGMP-dependent increase in ubiquitinated NKCC2 while increasing the expression of cullin-1 by threefold, but this effect was not seen with other cullins. CRL complex activity is also regulated by cullin-associated Nedd8-dissociated 1 (CAND1). CAND1 binds to cullins and promotes the exchange of substrate-recognition proteins to target different proteins for ubiquitination. CAND1 inhibition exacerbated the cGMP-dependent increase in NKCC2 ubiquitination and decreased surface NKCC2 expression. Finally, cGMP increased neddylation of cullins. We conclude that the cGMP-dependent increase in NKCC2 ubiquitination is mediated by a CRL complex. To the best of our knowledge, this is the first evidence that a CRL complex mediates NKCC2 ubiquitination in native TALs.The NaNEW & NOTEWORTHY The Na+/K+/2Cl− cotransporter (NKCC2) reabsorbs NaCl by the thick ascending limb. Nitric oxide and atrial natriuretic peptide decrease NaCl reabsorption in thick ascending limbs by increasing the second messenger cGMP. The present findings indicate that cGMP increases NKCC2 ubiquitination via a cullin-RING ligase complex and regulates in part surface NKCC2 levels. Identifying the E3 ubiquitin ligases that regulate NKCC2 expression and activity may provide new targets for the development of specific loop diuretics. NKCC2,  (NKCC2) –6 mainta (NKCC2) –7.cGMP, the second messenger of nitric oxide and atrial natriuretic peptide, exerts potent diuretic and natriuretic effects along the nephron –12. In tPhosphorylation and glycosylation are posttranslational mechanisms known to participate in the regulation of surface NKCC2 expression and therefore NKCC2 activity , 13, 14.2-terminal regions, whereas in the COOH-terminus, all members of the cullin family contain a lysine residue that is targeted by Nedd8  with members of the cullin family –38. The by Nedd8 , 35. Culby Nedd8 , which iby Nedd8 , leadingby Nedd8 , 44. TheOn the other hand, CRL complex activity is also regulated by cullin-associated Nedd8-dissociated 1 (CAND1). CAND1 binds unneddylated cullin and promotes the exchange of substrate-recognition proteins to target different proteins for ubiquitination –48. CAND+ channel (ENaC) prevents ENaC degradation  phosphorylation, leading to exacerbated NCC-dependent NaCl reabsorption in the distal nephron . The reagents and antibodies used are shown in The composition of the physiological solution (PS) was as follows: 130 mM NaCl, 2.5 mM NaHg for 2 min, suspended in chilled PS, and stirred on ice for 30 min to detach the tubules from each other. The suspension was filtered through 250-μm nylon mesh and centrifuged at 120 g for 2 min. The pellet was washed, centrifuged again, and finally suspended in 0.4 mL of chilled PS. In general, TAL suspensions were split into four aliquots for ubiquitin pulldown assays or experiments to study the biotinylation of plasma membrane proteins. All animal protocols were approved by the Institutional Animal Care and Use Committee of Henry Ford Hospital and Wayne State University.Rat medullary TAL suspensions were obtained as previously described , 7. Maleg for 2 min at 4°C. The PS was discarded, and TALs were incubated with lysis buffer . TALs were vigorously vortex three times for 3 s each. Each tube was spun at 12,000 g for 2 min at 4°C. The undissolved pellet was discarded. The protein content in each sample was measured in duplicate with a colorimetric assay using Bradford’s method . The TAL lysate (150 μg protein) was incubated on a rocking platform at 4°C overnight with 40 μL of a 50% slurry containing Ubiqapture-Q beads in a final volume of 400 μL. The beads were centrifuged at 12,000 g for 2 min at 4°C. The supernatant was separated from the beads and saved for later measurement of nonubiquitinated NKCC2 and was also used as a loading control. The beads were washed twice with high-salt buffer  and twice with no-salt buffer . Proteins were eluted from the beads by boiling in 60 μL of SDS Laemmli loading buffer containing 50 μM dl-dithiothreitol and 5% β-mercaptoethanol. Proteins from the supernatant and proteins eluted from the beads were separated by SDS-PAGE (6% gels) and transferred to Immobilon PVDF membranes . NKCC2 was detected by Western blot analysis.Ubiquitinated NKCC2 was measured as previously described . Brieflyg for 2 min at 4°C. TAL samples were then incubated with 0.75 mL of chilled biotinylation solution  containing 1.2 mg/mL NHS-SS-biotin (Pierce Biotechnology) on a rocking platform at 4°C for 15 min. Then, 0.75 mL of freshly prepared NHS-SS-biotin (1.2 mg/mL) was added to the top of the solution, and samples were incubated for an additional 15 min. After biotinylation, tubules were washed three times at 4°C with PS containing 100 mM glycine to remove excess NHS-SS-biotin. TALs were then centrifuged at 120 g for 2 min at 4°C and lysed in buffer (composition listed in a previous protocol). The protein content in each sample was measured in triplicate by colorimetric assay following the Bradford method (Pierce Biotechnology). Equal amounts of protein (50–75 μg) were incubated overnight at 4°C with streptavidin-coated agarose beads (10%) in lysis buffer. The beads were pulled down by centrifugation, and the supernatant was incubated with streptavidin-coated agarose beads (10%) for 2 h at 4°C. The supernatant was saved for use in determining intracellular NKCC2 levels, and the beads were centrifuged and pooled with the beads obtained from the first preparation. All beads were then washed twice in lysis buffer, twice in high-salt buffer , and twice in no-salt buffer . Biotinylated proteins were extracted from the beads by boiling in 60 μL of SDS-loading buffer containing 50 μM dl-dithiothreitol and 5% β-mercaptoethanol. The proteins were separated by SDS-PAGE (6% gels), and NKCC2 in the membrane was detected by Western blot analysis.Cell surface biotinylation of TAL suspensions was performed as previously described –7. Briefg, loaded into each lane of a 6% SDS-polyacrylamide (PAGE) gel, separated by electrophoresis, and transferred to Immobilon-P PVDF membranes. Membranes were blocked with 50 mM Tris, 150 mM NaCl, 5% nonfat dried milk, and 0.1% Tween 20 at pH 7.4 for 60 min and then incubated with primary antibodies . These data indicate that the cGMP-dependent increase in NKCC2 ubiquitination is mediated by a CRL complex.The second messenger cGMP decreases NaCl reabsorption by decreasing surface NKCC2 levels . We haven = 6, P < 0.05; We have previously shown that inhibition of proteasomal degradation blunted the cGMP-dependent decrease in surface NKCC2 levels. However, whether preventing the ubiquitination of NKCC2 would blunt the cGMP-dependent decrease in surface NKCC2 levels was unknown. Therefore, we sought to determine whether inhibition of cullin neddylation would prevent the cGMP-dependent decrease in surface NKCC2 levels. First, TALs were divided into two groups and treated with vehicle or MLN4924. TALs were then divided again into two groups and treated with saline or db-cGMP (500 µM). We found that inhibition of cullin neddylation prevented the cGMP-dependent decrease in surface NKCC2 levels  was added to the PS. To obtain the required amount of protein for this protocol, TAL suspensions from two Sprague–Dawley rats were combined and mixed. The TAL suspension was then split into two aliquots and treated with either vehicle or 1 µM C60. Later, each TAL sample was further split into three groups and treated with saline, 100 µM db-cGMP, or 500 µM db-cGMP. We found that 1 µM C60 increased NKCC2 ubiquitination as well as the cGMP-dependent increase in NKCC2 ubiquitination . cGMP also increased cullin-1 expression in TALs . We found that cGMP decreased CAND1 expression in TALs  and the CAND1 inhibitor C60 (1 µM). TALs were equilibrated for 10 min and then treated with vehicle or 100, 500, or 1,000 µM cGMP for 50 min. We found that cGMP increased neddylation of members of the cullin family in TALs . Taken together, these previous reports align with our findings in which members of the cullin family are expressed at the mRNA and protein level and support our findings that a CRL mediates ubiquitination of NKCC2 in TALs.We also examined expression of each member of the cullin family at the mRNA level. By performing RT-PCR, we found that all members of the cullin family  were expressed at the mRNA level in TALs. Therefore, expression of cullin-4B, cullin-7, and cullin-9 in the TAL cannot be ruled out based on our Western blot data. Using a proteomic approach, Limbutara et al.  dissecteHowever, the role of each member of the cullin family in TAL-dependent NaCl reabsorption remains unknown since the function of each member of the cullin family has not been explored.Cullins act as scaffold proteins that recruit members of the CRL complex . A critiThe regulation of NKCC2 ubiquitination remains poorly understood. Since inhibition of CAND1 activity increased baseline NKCC2 ubiquitination, these data show, for the first time, that the ubiquitination of NKCC2 can be stimulated. Since NKCC2 ubiquitination regulates surface NKCC2 levels, we examined whether the increase in NKCC2 ubiquitination would be accompanied by exacerbation of the cGMP-dependent decrease in surface NKCC2 levels. Our data show that inhibition of CAND1 with the compound C60 decreased surface NKCC2 levels and that cGMP was not able to further decrease surface NKCC2 levels. To the best of our knowledge, these data show that the ubiquitination of NKCC2 regulates surface NKCC2 levels in TALs.+ intake stimulates NCC phosphorylation while decreasing ubiquitination. On the other hand, Murali et al. (+ intake was attenuated when cullin activity was inhibited. Taken together, these studies suggest that an increase in cullin activation is indirectly related to NCC phosphorylation. Furthermore, Murali et al. (+ can still reduce phosphorylation of NCC, suggesting that another member of the cullin family may regulate the K+-dependent regulation of NCC phosphorylation. Based on the proximity and similarities between NCC and NKCC2, the phosphorylation and ubiquitination of NKCC2 may be regulated in a manner similar to those of NCC. This is based on the fact that although the role of K+ on cullin activity in TALs has not yet been studied, Stoke et al. (+ diet decreased TAL-mediated NaCl reabsorption, which is in agreement with the recent aforementioned reports. Since NCC and NKCC2 share several conserved motifs, the ubiquitination and phosphorylation of NKCC2 may also be related. However, despite the considerable amount of work done to examine NKCC2 phosphorylation, no study has focused on whether NKCC2 phosphorylation affects the NKCC2 ubiquitination state. Therefore, the interaction between the ubiquitination and phosphorylation of NKCC2 deserves attention.The regulation of renal transporters is dynamic and involves more than one posttranslational modification. This has been observed in work done with NCC. Rosenbaek et al.  showed ti et al.  showed ti et al.  showed te et al.  showed tThe data show that cGMP stimulates NKCC2 ubiquitination via a CRL complex. However, the site of action of cGMP is unknown. We found that cGMP increased neddylation of members of the cullin family in rat TALs. We detected an increase in Nedd8 expression, which corresponded to a molecular weight of ∼90–100 kDa, which is the expected molecular weight of cullin family members; since Nedd8 is a small peptide, Nedd8 is detected only with neddylated proteins. Next, we focused on cullin-1, which was expressed at higher levels in cGMP-treated samples. Since the samples were treated with cGMP for only 20 min, we speculate that neddylated cullin-1 was likely protected from degradation and that synthesis of cullin-1 at the protein level was not stimulated. It is also possible that the increase in cullin-1 expression observed may have been due to inhibition of its autoregulation. We studied the effect of cGMP on CAND1 expression. Our data show that cGMP decreased CAND1 expression in native TALs. Currently, little is known about how CAND1 is regulated in mammals. Finally, the data show that the effect of cGMP on neddylation of members of the cullin family was exacerbated when CAND1 was inhibited. Control experiments showed that inhibition of CAND1 did not change baseline neddylation of members of the cullin family nor affect CAND1 expression or total NKCC2 expression. Taken together, the data show that cGMP increases neddylation of members of the cullin family in combination with a decrease in CAND1 expression promoting the activation of a CRL complex, which stimulates ubiquitination of NKCC2. Moreover, the data shown here and in our previous publication demonstrate that the cGMP-dependent increase in NKCC2 ubiquitination leads to the degradation of NKCC2 by the proteasome, decreasing surface NKCC2 levels . Up to dOverall, our data show that the cGMP-dependent increase in NKCC2 ubiquitination is mediated by a CRL complex. To the best of our knowledge, this is the first evidence pointing to a specific E3 ubiquitin ligase family that is stimulated by cGMP and has a direct effect on a renal NaCl transporter.Data will be made available on reasonable request.This work was supported in part by the National Institute of Diabetes and Digestive and Kidney Diseases Grant 1K01DK123192 and Mentored Scientist Institutional support from the Henry Ford Health System.No conflicts of interest, financial or otherwise, are declared by the author.G.R.A. conceived and designed research; performed experiments; analyzed data; interpreted results of experiments; prepared figures; drafted manuscript; edited and revised manuscript; approved final version of manuscript.


Candidatus Nitrosopumilus and bacteria belonging to the Gammaproteobacteria , indicating that nitrogen-based metabolisms, particularly ammonia oxidation to nitrite by Cand. Nitrosopumilus was prevalent. At the onset of the vernal sea-ice algae bloom, the community shifted to the dominance of Gammaproteobacteria  and Bacteroidia (Polaribacter), while Cand. Nitrosopumilus almost disappeared. The bioinformatically predicted carbohydrate-active enzymes increased during spring and summer, indicating that sea-ice algae-derived carbon sources are a strong driver of bacterial and archaeal community succession in Arctic sea ice during the change of seasons. This implies a succession from a nitrogen metabolism-based winter community to an algal-derived carbon metabolism-based spring/ summer community.The Arctic is warming 2–3 times faster than the global average, leading to a decrease in Arctic sea ice extent, thickness, and associated changes in sea ice structure. These changes impact sea ice habitat properties and the ice-associated ecosystems. Sea-ice algal blooms provide various algal-derived carbon sources for the bacterial and archaeal communities within the sea ice. Here, we detail the transition of these communities from winter through spring to early summer during the Norwegian young sea ICE (N-ICE2015) expedition. The winter community was dominated by the archaeon  The increase in global average temperature has pronounced effects on the Arctic , leadingPhaeocystis pouchetii [Alpha- and Gammaproteobacteria and changed from a dominance of Candidatus Pelagibacter during winter to the dominance of Gammaproteobacteria in spring and early summer [Candidatus Nitrosopumilus, a member of the Nitrososphaeria (formerly classified as both Marine Group I Crenarchaeota and Marine Group I Thaumarchaeota), in the surface samples in winter and a near absence in the summer samples. This decrease is most likely due to photo-inhibition of members of this group at high light regimes in spring and summer and competition with phytoplankton for ammonium in the surface ocean [During the N-ICE expedition, a pelagicphytoplankton bloom was observed under the ice from late May to late June . This blouchetii . The bacouchetii . The bacy summer . This chy summer ,8. The mce ocean ,9,10.Nitzschia frigida was found at the bottom of second-year ice (SYI) [In addition to the under-ice phytoplankton bloom, the sea-ice algae bloom dominated by ce (SYI) . Such blce (SYI) ,8. Flavobacteriia and Gammaproteobacteria, but also Alphaproteobacteria, Verrucomicrobia, and Bacilli [Crenarchaea of the Marine Group I was found in ice and the underlying water, suggesting an abundance of the ammonia-oxidizing Candidatus Nitrosopumilus, which have been classified in this group previously [Gammaproteobacteria  and Flavobacteriia , have been found in high abundance in connection to phytoplankton blooms in marine environments and similar blooms of sea-ice algae, e.g., Fragilariopsis cylindrus, Nitzschia frigida, or Melosira arctica, which occur in the bottom layers of sea ice in spring/summer [Bacteroidia and Gammaproteobacteria have been found in high abundance in Arctic sea ice and brine samples [Despite the low temperatures and high salinity in the brine channels, bacterial and archaeal communities thrive in these environments, both in Arctic and Antarctic sea ice. The communities are mostly dominated by  Bacilli ,14,15,16eviously ,18. Thisg/summer ,20. Gene samples ,22. Howe samples ,22. ThisThe transition of bacterial and archaeal communities in sea ice from winter to summer is widely unknown. Such studies could elaborate on the status of algal-fueled microbial ecosystems and potentially enable conclusions about the changes in these ecosystems regarding the ongoing effects of global warming. Here, we present a seasonal investigation of microbes in Arctic sea ice from winter to spring and early summer of 2015, hypothesizing an algal-derived carbon sources triggered the succession of the bacterial and archaeal communities.RV Lance [Ice cores were sampled during the Norwegian young sea ICE N-ICE2015) expedition from 15 January to 22 June 2015 in the Arctic sea ice pack north of Svalbard, Norway, on board  expeditiRV Lance . Due to RV Lance .−1 and HNF at 500 µL min−1, following protocols modified from Marie et al. [The triplicates of 1.8 mL FCM samples were fixed with glutaraldehyde  at 4 °C for a minimum of 2 h and then flash frozen in liquid nitrogen and stored at −80 °C until later analyses. Abundance of bacteria was determined on an Attune Acoustic Flow cytometer  with a syringe-based fluidic system using a 20 mW 488 nm blue laser after SYBR-Green staining. Prior to analyses, the samples were thawed and stained with SYBR Green I  for a minimum of 1 h. Heterotrophic bacteria and archaea were analyzed using a flow rate of 25 µL ming, saving the flow-through for subsequent RNA extraction. Centrifugation steps were repeated for the remaining lysate volume, as necessary. The extracted RNA was treated using the DNA-free DNA Removal kit  prior to reverse transcription using the SuperScript III First-Strand Synthesis kit for Real-Time PCR (Invitrogen), according to the manufacturer’s instructions. The DNA was quantified using a Qubit 3.0 Fluorometer . Primer Both cDNA and DNA were then used to prepare sequencing libraries targeting the V4 region of the 16S rRNA genes, using multiplex identifier tagged primers 519F—5′-CAGCMGCCGCGGTAA-3′ and 806RB—5′-GGACTACNVGGGTWTCTAAT-3′ [For phylogenetic analyses of the bacterial and archaeal diversity and a corresponding inference of their activity, the 16S rRNA genes were sequenced from DNA and RNA extracted from the samples. For that, the Sterivex filters were thawed on ice, and subsequently, a lysis solution was added. DNA and RNA were extracted from filters using the AllPrep DNA/RNA Mini Kit  with a modified protocol as described in Wilson et al. . BrieflyCTAAT-3′ ,28. The CTAAT-3′ . In the nifH, nirS/nirK, norB, nosZ, narB, napA, hzsA, amoA-pmoA, nrfAhao, , aprA, dsrA, cysH, soxB, mmoX, mcrA, psaA, and psbA, were used for metabolic analyses (The retrieved sequences were stored in the European Nucleotide Archive PRJEB47256) and processed using DADA2 in R . Briefly256 and panalyses . CAZymes5 cells mL−1 on 21 May and 2.1 × 106 ± 1.3 × 104 cells mL−1 in June, dominated by the pennate diatom Nitzschia frigida/neofrigida [a concentration in the lower part of the ice cores from <0.05 mg m−3 in winter to 6.25 mg m−3 in May, 3.15 mg m−3 on 4 June, and 1.04 mg m−3 on 17 June , Bacteroidia (20.6 ± 12.9% rel. abundance), and Alphaproteobacteria (16.7 ± 3.2% rel. abundance), complemented with Verrucomicrobiae (3.2 ± 3.9% rel. abundance), members of the SAR324 clade (0.9 ± 0.9% rel. abundance), and several other classes with <0.5% average rel. abundance  to a spring/ early summer community with high rel. abundance of Bacteroidia (33.6 ± 3.5% rel. abundance) was visible. Gammaproteobacteria showed higher rel. abundance in spring/early summer (41.2 ± 4.8% rel. abundance) as compared to winter (33.2 ± 4.7% rel. abundance), while Alphaproteobacteria remained constant in rel. abundance ~18.6%  showed differences between the samples, confirming the differences of the communities between seasons and in correlation to the sea-ice algae bloom . The icep-value = 0.012), whereas the main ASV of Nitrincolaceae showed significantly higher rel. abundance in summer with 8.2 ± 0.3% . In addition, one ASV assigned to Sulfitobacter was significantly higher in spring and summer with >1% rel. abundance . In winter, 736 ASVs were found (293 unique ASVs), as compared to 173 ASVs (15 unique ASVs) in spring and 351 ASVs (25 unique ASVs) in summer, with 121 ASVs being present in all communities . Using ICand. Nitrosopumilus from winter to spring, when the overall active Kangiellaceae and Nitrincolaceae dominated, and early summer, when Polaribacter, Lewinella, and Lentimonas were most active. The overall activity of the bacterial community was dominated by Gammaproteobacteria (63.3 ± 7.0% rel. cDNA abundance), Bacteroidia (12.0 ± 6.8% rel. cDNA abundance), and Alphaproteobacteria (8.9 ± 2.5% rel. cDNA abundance), complemented by Verrucomicrobiae (1.7 ± 2.8% rel. cDNA abundance), Nitrospinia (1.2 ± 1.2% rel. cDNA abundance), Desulforomonadia (0.8 ± 1.5% rel. cDNA abundance), Dehalococcoida (0.7 ± 0.8% rel. cDNA abundance), Bdellovibrionia (0.7 ± 0.7% rel. cDNA abundance), and members of the SAR324 clade  from 0.41 ± 0.11% to 0.95% in spring and 0.80 ± 0.18% in early summer  from 0.17 ± 0.03% and 0.23 ± 0.03% to 0.29% and 0.34% in spring and 0.24 ± 0.01% and 0.31 ± 0.05% in early summer , amoA (Nitrification), aprA (Dissimilatory sulphate reduction), and cysH (Assimilatory sulfate reduction) reached a rel. abundance of >0.05%, while nifH (Nitrogen fixation), norB (Denitrification), nosZ (Denitrification), narB (Assimilatory nitrate reduction), hao (Nitrification), nrfA (Dissimilatory nitrate reduction to ammonium), and dsrA (Dissimilatory sulfite reduction) were predicted in low abundance (<0.01%), and hzsA (Annamox), soxB (Sulphur oxidation), mmoX (Aerobic methane oxidation), mcrA (Methanogenesis), psaA (Photosynthesis), and psbA (Photosynthesis) were not predicted  to <0.01% in spring and early summer  was found in Polaribacter, Arcobacteraceae, and Synechococcus, while nitrogen fixation (nifH) was predicted for Bdellovibrionia (members of the OM27 clade), Campylobacteria (Arcobacteraceae), and Desulfuromonadia  to 0.01% in spring and increased again to 0.03 ± 0.01% in early summer. The marker genes for dissimilatory sulfate reduction and oxidation (aprA and dsrA) were predicted for members of the OM27 clade (Bdellovibrionia), Desulfofrigus (Desulfobacteria), Desulforhopalus (Desulfobulbia), and PB19 (Desulfuromonadia), and members of the SAR11 clades Ia, II, and further unspecified members, as well as members of the AEGEAN-169 Marine Group only aprA. The cysH gene was predicted stable at 0.11 ± 0.01% rel. abundance in all samples and for a large variety of classes, dominated by Cryomorphaceae and members of the NS9 Marine Group in the Bacteroidia, while Magnetospiraceae dominated the abundance in the Alphaproteobacteria, members of the OM182 clade the Gammaproteobacteria, and Lentimonas and Roseibacillus the Verrucomicrobia.The Here, we describe the transition of the bacterial and archaeal communities of Arctic sea ice from winter to spring and early summer. Even though this is not a true time series due to the necessary relocation in drifting ice, the samples taken on Floe 3 (April to June) show a clear succession of the bacterial and archaeal community. The community from April matches the communities from January and March (Floes 1 and 2), while the community from 17 June (Floe 4) is similar to 4 June (Floe 3). Thus, suggesting comparability of samples. In addition, the high taxonomic resolution of the ASVs supports the comparability since most ASVs were found in most or all samples. Nitzschia frigida dominated the bottom ice algal assemblage of both FYI and SYI in May and June. Other pennate diatoms, unidentified flagellates, and resting cells  came next in importance, while centric diatoms and ciliates occurred at low abundances  may also use nitrite oxidation as the main energy source, thus explaining the relatively high abundance during winter and adding to the production of nitrate [Cand. Nitrosopumilus mortality and would explain the near absence in light conditions in spring and early summer [Cand. Nitrosopumilus is adapted to very low ammonia concentrations, the competition could add to the explanation of the decreasing rel. cDNA abundance of Cand. Nitrosopumilus in April and the near disappearance of this taxon during the sea-ice algae bloom. However, this would not explain the decreased rel. cDNA abundance in January, which might correlate to the lower abundance in this sample. In addition, competition with fast-growing members of the Gammaproteobacteria and Bacteroidia could lead to a decrease in Cand. Nitrosopumilus in carbon-replenished spring and summer samples [The most drastic change was the vanishing of e waters ,26,51,52fication , thus ex sea ice . The nitopumilus ,55,56, w nitrate ,58. Photy summer ,59,60. Iy summer . Even th samples ,63. aprA gene in winter samples is due to the abundance of members of SAR11 clades. AprA/B has been found in SAR11 previously, and dissimilatory sulfate oxidation has been suspected [Desulfuromonadia [Members of the alphaproteobacterial SAR11 clade are resilient, slow-growing organisms with low requirements for carbon sources and high abundance in oligotrophic polar oceans ,65,66,67uspected ,69. The uspected ,58,70. Muspected  and haveomonadia  also poiGammaproteobacteria and Bacteroidia are usually found in eutrophic, carbon-rich marine environments, often in correlation with phytoplankton blooms, thus explaining their abundance in the spring and early summer samples [Nitrincolaceae, members of the OM182 and SAR92 clades, and the Alteromonadaceae groups Arenicella, Paraglaciecola, and Colwellia, have been found in association with phytoplankton blooms and polar waters [Nitrincolaceae harbor a high diversity of carbon metabolisms, explaining their abundance during the sea-ice algae bloom [Kangiellaceae were suggested to extracellularly utilize amino acid as carbon and nitrogen sources, explaining the abundance of Kangiellaceae in early winter and summer samples [Bacteroidia harbor a multitude of carbon degradation pathways, leading to successions of different groups after phytoplankton blooms [Polaribacter co-occur with phytoplankton blooms [Verrucomicrobiae, specifically Lentimonas, degrade complex carbon sources, and hence have been found in later stages of phytoplankton blooms [Rhodobacteraceae, such as Sulfitobacter and Octadecabacter, are often correlated to diatom blooms [Nitrincolaceae and Sulfitobacter being indicator species for spring and early summer. This succession might even go on after early summer, as samples from Arctic sea ice and brines with a high abundance of different taxa within the Gammaproteobacteria and Bacteroidia indicate [Gammaproteobacteria  and Bacteroidia . However, due to the lack of fall samples in this study and the geographical distances between the studies, this has to remain a hypothesis. samples ,77,78,79r waters ,81,82,83ae bloom . Kangiel samples . Bacteron blooms ,67,86,87n blooms ,89. In an blooms ,91,92. Rm blooms . Thus, aindicate ,21,22. OP. pouchetii bloom observed below the ice starting on 25 May [Alphaproteobacteria and Cand. Nitrosopumilus in winter to Gammaproteobacteria and Bacteroidia in spring and early summer was found [The pattern of increased predicted CAZyme abundance in Bacteroidia and Gammaproteobacteria in spring and early summer, as compared to winter, is based on the abundance of these taxa. Therefore, an increase in CAZymes indicates the potential of the various genera within the Bacteroidia and Gammaproteobacteria to utilize different carbon sources and, consequently, different ecological niches. These community patterns are also congruent with the finding in a sample from adjacent first-year ice (FYI), where a similar bloom occurred , and then 25 May , where aas found . This coas found . This coas found ,8,77.Bacteria and Archaea during the transition from winter to early summer in 2015 north of Svalbard. We show the transition of this community from a winter state with decreasing cell numbers, mostly nitrogen-based metabolisms, and dominance of Cand. Nitrosopumilus from January to an early spring state in March. There, nitrification potentially replenished the nitrate pool, which could then be used by other organisms during spring and summer. In spring, Bacteroidia and Gammaproteobacteria increased in abundance, while Cand. Nitrosopumilus nearly vanished. The succession of Gammaproteobacteria and Bacteroidia is driven by season and the corresponding availability of carbon sources derived from the light-induced, diatom-dominated ice algal spring bloom, which leads to the establishment of different ecological niches, subsequently exploited by the best-adapted groups. Given the changes in arctic environments based on global warming, this seasonal succession from dominantly nitrogen-based to dominantly carbon-based cycling could be interrupted or accelerated by alteration in the sea-ice environment, potentially leading to changes in the microbial sea-ice ecosystem.Here, we report the community composition and activity of Arctic sea-ice


Staphylococcus aureus. The production of local IgE is modulated by lymphocytes, cytokines, transcription factors, and B cell-intrinsic factor. Due to the central role of IgE in NPs, it is regarded as an ideal target for therapy and has been proved to be clinically successful. Based on this knowledge, we believe that exploring the trigger and regulatory factors for the activation of local B cells and CSR to IgE will provide more valuable information for us to recognize the pathological mechanisms of local IgE and offer the possible option for new therapeutic targets of nasal polyps.In the airway, IgE is traditionally regarded as a key mediator in allergic diseases, such as AR and allergic asthma. However, growing evidence demonstrates the importance of local IgE in airway inflammatory diseases, irrespective of the presence of allergy. In this review, we discuss the most recent evidence for IgE in chronic rhinosinusitis with nasal polyps(CRSwNP), including the local IgE’s characteristics, the modulation of its synthesis, and its function. The levels of local IgE are significantly elevated in polyps independently of IgE serum levels and atopic status. Local IgE, which is correlated with type 2 inflammation, is polyclonal and functional. IgE is produced by active B cells and is dependent on the class switch recombination(CSR). In NPs, this process is triggered by not only allergens but also microbial colonization, especially the superantigen-  Immunoglobulin E(IgE) has an essential function in immunity to parasites and type I hypersensitivity. Although IgE is typically the least abundant isotype, it is one of the responsible antibodies for the classical adaptive immune response and is capable of triggering the most powerful inflammatory reactions after binding to its specific receptors .Traditionally, IgE is regarded as a key mediator in the pathophysiology of allergic diseases, such as allergic rhinitis(AR), allergic asthma, anaphylaxis, and food allergy. People with a tendency to develop allergic reactions when exposed to antigens initially produce antigen-specific IgE during sensitization. Subsequent exposure to its specific sensitizing allergen, IgE mediates antigen cross-linking of the high-affinity IgE receptor (FcϵRI) on a variety of cells . Release of mediators by mast cells and basophils follows, triggering classic symptoms of allergy However, our concept of the role of IgE in disease states is undergoing reassessment and re-evaluation., Growing evidence shows that IgE also plays a role in non-allergic airway diseases. Increased local IgE production was found in nasal polyps(NPs) regardless of systemic atopic status –4. ClassEven if IgE is involved in both allergic and nonallergic airway diseases, its role may vary in these diseases. In AR, IgE concentrations in serum and mucosal tissue homogenates are highly correlated, while they fail to show relevant correlations in chronic rhinosinusitis with nasal polyps(CRSwNP) and comorbid asthma . AlthougIn this review, we focused on the role of IgE in NPs. We summarized the characteristics of local IgE in NPs, then analyzed the possible pathway of local IgE synthesis and its modulation, and finally discussed the function of IgE in NPs, including recent information from targeting IgE.in vitro  on basophil, finally contributing to the persistent local inflammation in CRSwNP.IgE antibody in CRSwNP appear to be polyclonal and covers a broad spectrum of allergens/antigens rather than oligoclonal ones like in AR , 19. LocThe Type2(T2) inflammation in NPs includes higher levels of local IgE, IL-5 , IL-4, I+CD27+CD38hi plasmablasts were dramatically increased in CRSwNPs, which also had a higher frequency of expression of Epstein Barr Virus-induced protein 2(EBI2), compared to tonsils samples , a specific circle transcript of Iϵ-Cm (408 bp) produced during CSR from IgM to IgE was detected to be increased in eosinophilic NPs, meaning Ig class switching to IgE in eosinophilic polyps with ectopic lymphoid tissues , Staphylococcus pyogenes, and Haemophilus influenzae, were found in polyps of patients with CRSwNP  cells can antagonize ϵCSR in activated B cells , composed of galectin-10 , are proteins produced by eosinophils, basophils, and some T cells . They arThe binding of phosphorylated signal transducer and activator of transcription 6(pSTAT6) to the Iϵ promoter is essential for promoter activation. Transcription factors including E2A, NF-κB, PU.1, Pax5, AP-1, C/EBP and SWAP-70 synergizes with pSTAT6 to activate germline Iϵ transcription , 97–101.In studies focusing on B cell-intrinsic factors, three different IgE reporter mice were used to explore the regulation of IgE class switching and production. With M1 prime GFP knock-in IgE reporter mice, Talay O described that M1, an extracellular 52-amino-acid segment present in membrane IgE, induced CSR to ϵ . With VeIn addition, tumor necrosis factor receptor superfamily(TNFRSF) members play key roles in B cell development, maturation, homeostasis, activation, and differentiation, also influencing the ability of B cells to synthesize IgE antibodies and act as regulators of immune responses, such as CD40(TNFRSF5) and OX40L(TNFSF4), BAFF(TNFSF13B) and APRIL(TNFSF13) . BAFF, avia local induction of IgA  (2020MSXM035).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. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Fisher’s exact tests were used to compare individual genotype frequencies, whereas unweighted and weighted total genotype scores  were computed to assess differences in polygenic profiles. In isolation, the IL6 (rs1800795) G allele was overrepresented in PDP players (90.5%) compared to YDP players , whereby PDP players had nearly three times the odds of possessing a G allele . The TGS (p = 0.001) and TWGS (p < 0.001) were significant, but poor, in distinguishing YDP and PDP players (AUC = 0.643–0.694), with PDP players exhibiting an overall more power-orientated polygenic profile. If validated in larger independent youth football cohorts, these findings may have important implications for future studies examining genetic associations in youth football.The purpose of this study was to examine differences in the genotype frequency distribution of thirty-three single nucleotide variants (SNVs) between youth development phase (YDP) and professional development phase (PDP) academy football players. One hundred and sixty-six male football players from two Category 1 and Category 3 English academies were examined within their specific age phase: YDP ( The process of athlete development and ultimately reaching senior professional status in a sport such as football (soccer) is both dynamic and multifactorial . Indeed,The failure to clearly identify a set of variables that uniformly predicts performance levels is, in part, due to methodological issues identified throughout talent identification and development research in football . ProspecAlthough under-researched within a football context, inter-individual genetic variation also appears to influence performance and development in football  were significantly different between pre- (aged 10.6 ± 1.4 years) and post- (aged 16.8 ± 2.3 years) peak height velocity academy football players [Recent studies have begun to explore which specific genetic variants may explain some of the genetic influence on performance and development in football  , with agn = 92; aged 13.84 ± 1.63 years) and PDP . Informed assent from all players, consent from parents/guardians, and gatekeeper consent from each academy were collected prior to the commencement of the study. All experimental procedures were conducted in accordance with the guidelines in the Declaration of Helsinki and ethical approval was granted by the corresponding author’s institutional Ethics Committee. This study was conducted in accordance with the recommendations for reporting the results of genetic association studies defined by the Strengthening the Reporting of Genetic Association studies (STREGA) statement.One hundred and sixty-six male football players from two Category 1 and Category 3 English academies participated within their specific age phase: YDP  for DNA extraction. Using Qiagen chemistry, DNA was extracted on an automated Kingfisher FLEX instrument . To measure the quality and quantity of extracted DNA, PicoGreen and Nanodrop measurements were taken. Input to the custom testing array occurs at 200 ng in 20 µL. Amplification, fragmentation, and resuspension were performed using Biomek FXP. GeneTitan instrumentation  was used to stain and scan the arrays, with hybridization performed in a Binder oven at 48 degrees for 24 h, following the Affymetrix Axiom high throughput 2.0 protocol. Data analysis was then performed using raw CEL file data input into the Affymetrix Axiom Analysis Suite . Procedures were in accordance with previous studies ,20,24.ACTN3, AMPD1, ADRB2, ACE, AGT, CPNE5, CKM, FTO, HSD17B14, HIF1A, IGF1, IGF2, IL6, NOS3, PPARA, PPARG, GALNT13, SOD2, TRHR, UCP2) and psychological/technical  phenotypes in previous studies with academy football players [https://www.genenames.org). Standard genomic quality control (QC) procedures and thresholds were applied when selecting genetic variants: SNV call rate (>95), sample call rate (>95), Fisher’s linear discriminant (>3.6), and minor allele frequency (>0.05).The SNVs in 33 genes see  were sel players ,21,24,25Unweighted and weighted total genotype scores  were calculated to assess the differences in polygenic profiles between YDP and PDP players . BotFor the TGS, the original procedure of Williams and Folland  was follFor the TWGS, a similar procedure to Varillas Delgado et al.  was usedt-test was used to assess differences in the TGS and TWGS between YDP and PDP players. Additionally, receiver operating characteristic (ROC) curves and area under the curve (AUC) were used to evaluate the discriminatory power of the TGS and TWGS to distinguish YDP and PDP players with threshold values of: >0.5–0.7 = poor, >0.7–0.8 = acceptable, >0.8–0.9 = excellent, and >0.9 = outstanding [p < 0.05.Data were analyzed using Jamovi version 1.8.1 and IBM SPSS version 25. Fisher’s exact tests were used to test SNVs for adherence with Hardy–Weinberg equilibrium (HWE) and to compare genotype frequencies between YDP and PDP players. Akaike information criterion (AIC) was used to select which genetic model  best fit the data and would be subjected to hypothesis testing. However, if MAF ≤ 0.25, a dominant model was utilized to retain statistical power . An indestanding . Odds raGALNT13 (p < 0.001) and UCP2 (p = 0.010) in the PDP group (see IL6 was significantly different between YDP and PDP players (p = 0.023)  see . More sp023) see .t (164) = 3.26, p = 0.001). The YDP tertile distribution was: lower = 27, middle = 43, and higher = 22, whereas the PDP tertile distribution was: lower = 16, middle = 24, and higher = 34. Compared to YDP players, PDP players had 2.61 times the odds of having a TGS in the higher third  than a TGS in the lower third , as well as 2.77 times the odds of having a TGS in the higher third than a TGS in the middle third . The ROC analysis determined that TGS frequency distribution showed significant, but poor, discriminatory power in distinguishing YDP and PDP players .The TGS of players ranged from 31 to 69 in the YDP group and 38 to 73 in the PDP group see . The meat (164) = 4.34, p < 0.001). The YDP tertile distribution was: lower = 39, middle = 29, and higher = 24, whereas the PDP tertile distribution was: lower = 11, middle = 24, and higher = 39. Compared to YDP players, PDP players had 5.76 times the odds of having a TWGS in the higher third  than a TWGS in the lower third , as well as 2.93 times the odds of having a TWGS in the middle third  than a TWGS in the lower third . The ROC analysis determined that TWGS frequency distribution showed significant, but poor, discriminatory power in distinguishing YDP and PDP players .The TWGS of players ranged from 31 to 74 in the YDP group and 24 to 78 in the PDP group see . The meaIL6 (rs1800795) G allele in PDP players compared to YDP players. In addition, the TGS and TWGS models demonstrated that the combination of these thirty-three SNVs was effective in differentiating YDP and PDP players. As such, these results suggest there is significant genetic variation between youth football players of distinct age groups. To our knowledge, this is the first assessment of genotype frequency distribution in isolation, and as part of a polygenic profile, between two age-specific phases of academy football players in England. Therefore, these findings may have important implications for future studies examining genetic associations in youth football.This study examined differences in the genotype frequency distribution of thirty-three SNVs, both individually and collectively, between YDP and PDP English academy football players. The key findings showed an overrepresentation of the IL6 gene encodes for the pleiotropic cytokine interleukin-6 (IL-6), which has previously been associated with multiple biological processes relevant to sport performance  [IL6 (rs1800795) SNV alter promoter activity and consequently result in higher and lower IL-6 levels, respectively [The  muscle) . The cirectively . Higher ectively ,33. In cectively .IL6 (rs1800795) G allele in those who take part in power-based sports  compared to controls [IL6 (rs1800795), the G allele may better protect skeletal muscle and aid in repair during powerful muscle contractions, which subsequently allows for a higher volume of training that stimulates favorable adaptations and ultimately results in superior performance in high-intensity activities.More recent sport-specific research has shown that IL-6 may be an important biomarker in power-orientated sports and performance phenotypes. Studies assessing Polish and Spanish high performing athletes have reported an overrepresentation of the controls ,35. Croscontrols . TherefoAlthough power-orientated phenotypes such as acceleration, speed, and vertical jumps are important across all youth football age groups , they apIL6 (rs1800795) G allele in the PDP group compared to the YDP group due to its association with several power-orientated phenotypes. However, recent research in academy football has also shown that the G allele may protect PDP players from injury. More specifically, Hall et al. [IL6 (rs1800795) C/C genotype suffered significantly more injuries than G allele carriers. The authors noted that the association was possibly due to the combination of greater muscle damage and inflammation experienced by C allele carriers, alongside the higher intensity of match actions and increased frequency of training and/or competitive match-play in older age groups. As such, the overrepresentation of the G allele in the PDP group may be explained by a pleiotropic effect of IL6 (rs1800795) on power and injury.As competitive match-play demands shift more towards anaerobic capacities, academy recruitment teams may choose to retain players displaying superior power rather than endurance capabilities . This mal et al.  reportedIL6 (rs1800795) findings. Specifically, PDP players had a greater proportion of alleles previously associated with power-orientated phenotypes . This indicates PDP players may have an overall more power-orientated polygenic profile, which corresponds with similar findings reported in post-peak height velocity (aged 16.8 ± 2.3 years) academy football players using only four of these SNVs: ACTN3 (rs1815739), AGT (rs699), PPARA (rs4253778), and NOS3 (rs2070744) [The TGS and TWGS models showed that YDP and PDP football players have distinct polygenic profiles, with the TWGS demonstrating greater discriminatory accuracy. This suggests that whilst each SNV has a small additive effect, favorable alleles of individual SNVs have different degrees of influence. This corresponds with previous research in academy football players on physiological, psychological, and technical phenotypes that underpin differences in these age-specific phases ,20,21. THTR2A rs6311 T allele, ADBR2 rs1042714 C allele, BDNF rs6265 T allele, DBH rs1611115 C allele, DRD1 rs4532 C allele, DRD4 rs1800955 C allele, GABRA6 rs3219151 C allele) in academy footballers [The polygenic models also showed that in general YDP players had a greater proportion of favorable alleles in SNVs previously associated with psychological and technical phenotypes  used in this study was relatively small. However, this was still larger than the median sample size (N = 60) reported in a recent review of eighty genetic association studies in football [GALNT13 and UCP2), which can indicate genotyping error and may have influenced the findings.Nevertheless, this study does have important limitations that should be considered. For instance, we did not make adjustments for multiple comparisons, which may have increased type 1 errors. However, due to the exploratory nature of this study, in regard to the novel experimentation methods employed and the unique cohort, reducing type 2 errors was considered a priority. This is recommended in exploratory research, as a main aim is to ensure an important discovery is not missed in the first instance, which can be validated in subsequent dedicated replication studies . In addifootball . There wIL6 (rs1800795) G allele was overrepresented in PDP players compared to YDP players, possibly due to its theorised pleiotropic effect on power and injury phenotypes. Moreover, the TGS and TWGS models derived from all thirty-three SNVs effectively distinguished YDP and PDP players, with PDP players exhibiting an overall more power-orientated polygenic profile. As such, this study has shown for the first time that there is significant inter-individual genetic variation between youth football players of specific age phases in English academies. If validated in larger independent youth football cohorts, these findings may have important implications for future studies examining genetic associations in youth football.This study has presented novel evidence with regard to the genetic profiles of YDP and PDP male academy football players in England. To be specific, the