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Respiration in intertidal salt marshes generates dissolved inorganic carbon (DIC) that is exported to the coastal ocean by tidal exchange with the marsh platform. Understanding the link between physical drivers of water exchange and chemical flux is a key to constraining coastal wetland contributions to regional carbon budgets. The spatial and temporal (seasonal, annual) variability of marsh pore water exchange and DIC export was assessed from a microtidal salt marsh (Sage Lot Pond, Massachusetts). Spatial variability was constrained from 224Ra : 228Th disequilibria across two hydrologic units within the marsh sediments. Disequilibrium between the more soluble 224Ra and its sediment‐bound parent 228Th reveals significant pore water exchange in the upper 5 cm of the marsh surface (0–36 L m−2 d−1) that is most intense in low marsh elevation zones, driven by tidal overtopping. Surficial sediment DIC transport ranges from 0.0 to 0.7 g C m−2 d−1. The sub‐surface sediment horizon intersected by mean low tide was disproportionately impacted by tidal pumping (20–80 L m−2 d−1) and supplied a seasonal DIC flux of 1.7–5.4 g C m−2 d−1. Export exceeded 10 g C m−2 d−1 for another marsh unit, demonstrating that fluxes can vary substantially across salt marshes under similar conditions within the same estuary. Seasonal and annual variability in marsh pore water exchange, constrained from tidal time‐series of radium isotopes, was driven in part by variability in mean sea level. Rising sea levels will further inundate high marsh elevation zones, which may lead to greater DIC export. [ABSTRACT FROM AUTHOR]

The impacts of ongoing climate warming on cold‐regions hydrogeology and groundwater resources have created a need to develop groundwater models adapted to these environments. Although permafrost is considered relatively impermeable to groundwater flow, permafrost thaw may result in potential increases in surface water infiltration, groundwater recharge, and hydrogeologic connectivity that can impact northern water resources. To account for these feedbacks, groundwater models that include the dynamic effects of freezing and thawing on ground properties and thermal regimes have been recently developed. However, these models are more complex than traditional hydrogeology numerical models due to the inclusion of nonlinear freeze–thaw processes and complex thermal boundary conditions. As such, their use to date has been limited to a small community of modeling experts. This article aims to provide guidelines and tips on cold‐regions groundwater modeling for those with previous modeling experience. This article is categorized under:Engineering Water > MethodsScience of Water > Hydrological Processes [ABSTRACT FROM AUTHOR]

While river water temperatures are a strong control on instream processes and aquatic ecosystems, monitoring networks for river water temperatures are often sparse. Despite recent advancements in water temperature modeling strategies, current models struggle to provide real-time and reach-specific predictions across broad spatial domains. We developed a physically-based water temperature model coupled to the National Water Model (NWM) to assess the potential for water temperature prediction to be incorporated into the NWM at the continental scale. Using model forcings and outputs from the NWM v2.1 retrospective, we evaluated the ability of four model configurations of increasing complexity to simulate hourly water temperatures in the forested headwaters of H.J. Andrews Experimental Forest, Oregon, USA during a six-week summer period. Our modeling framework, representing a first effort at pairing water temperature simulation with predictions from the NWM, suggests that the NWM can be leveraged to give insight into other water quality variables. • NWM-WT simulates hourly river water temperatures using National Water Model data. • Models of increasing complexity were calibrated and validated in a montane headwater catchment. • Select model configurations produced temperature predictions with RMSE values under 1 °C during the validation period. • Calibrated runs highlight the need to better constrain streambed and groundwater heat fluxes. [ABSTRACT FROM AUTHOR]

Oceanic storms can strongly disturb the physical and biogeochemical conditions of transitional coastal waters. Impacts of extreme oceanic storms on coastal ecosystems have received limited attention worldwide, with no studies at higher latitudes (> 45°) where tropical cyclones have usually abated. This study investigates the combined impacts from marine and atmospheric forcing on a coastal lagoon in Prince Edward Island, Canada, during and after Extratropical Cyclone Fiona in September 2022. Physical (water levels and temperature) and biogeochemical (dissolved oxygen [DO], electrical conductivity, pH, nitrate–nitrogen concentrations, total suspended solids [TSS]) datasets from the lagoon and the watershed's tributaries, groundwater springs, and piezometers were used to assess ecosystem disturbance and recovery timelines following the storm. Fiona resulted in a 1.6 m storm surge into the lagoon that elevated water temperatures by up to 6°C, disturbed the density‐dependent stratification of salinity and temperature, and reduced the diel amplitude of DO, indicating a reduction in plant respiration due to ecosystem disturbance. The freshwater tributaries revealed sharp changes in flow (30‐fold increase), nitrate–nitrogen (NO3‐N) concentrations and loading (70‐fold increase), and TSS loading (40‐fold increase) to the lagoon during and immediately following the storm. The lagoon rapidly recovered (hours) from the hydraulic disturbance of the storm surge, but elevated nutrient levels persisted for months. The intensity and frequency of extratropical cyclones is projected to increase in the Northwest Atlantic, making field‐based studies of cyclone impacts on coastal waters critical for understanding future coastal ecosystem disturbance and recovery periods relative to the timing of future storms. [ABSTRACT FROM AUTHOR]

Human activities and climate change threaten coldwater organisms in freshwater ecosystems by causing rivers and streams to warm, increasing the intensity and frequency of warm temperature events, and reducing thermal heterogeneity. Cold‐water refuges are discrete patches of relatively cool water that are used by coldwater organisms for thermal relief and short‐term survival. Globally, cohesive management approaches are needed that consider interlinked physical, biological, and social factors of cold‐water refuges. We review current understanding of cold‐water refuges, identify gaps between science and management, and evaluate policies aimed at protecting thermally sensitive species. Existing policies include designating cold‐water habitats, restricting fishing during warm periods, and implementing threshold temperature standards or guidelines. However, these policies are rare and uncoordinated across spatial scales and often do not consider input from Indigenous peoples. We propose that cold‐water refuges be managed as distinct operational landscape units, which provide a social and ecological context that is relevant at the watershed scale. These operational landscape units provide the foundation for an integrated framework that links science and management by (1) mapping and characterizing cold‐water refuges to prioritize management and conservation actions, (2) leveraging existing and new policies, (3) improving coordination across jurisdictions, and (4) implementing adaptive management practices across scales. Our findings show that while there are many opportunities for scientific advancement, the current state of the sciences is sufficient to inform policy and management. Our proposed framework provides a path forward for managing and protecting cold‐water refuges using existing and new policies to protect coldwater organisms in the face of global change. [ABSTRACT FROM AUTHOR]

Surface effects of sea‐level rise (SLR) in permafrost regions are obvious where increasingly iceless seas erode and inundate coastlines. SLR also drives saltwater intrusion, but subsurface impacts on permafrost‐bound coastlines are unseen and unclear due to limited field data and the absence of models that include salinity‐dependent groundwater flow with solute exclusion and freeze‐thaw dynamics. Here, we develop a numerical model with the aforementioned processes to investigate climate change impacts on coastal permafrost. We find that SLR drives lateral permafrost thaw due to depressed freezing temperatures from saltwater intrusion, whereas warming drives top‐down thaw. Under high SLR and low warming scenarios, thaw driven by SLR exceeds warming‐driven thaw when normalized to the influenced surface area. Results highlight an overlooked feedback mechanism between SLR and permafrost thaw with potential implications for coastal infrastructure, ocean‐aquifer interactions, and carbon mobilization. Plain Language Summary: Along coastlines globally, sea‐level rise is causing saltwater to intrude into terrestrial environments and freshwater reservoirs (i.e., saltwater intrusion). The impact of saltwater intrusion on temperate and tropical environments has been extensively studied, but assessment of saltwater intrusion impacts on high‐latitude permafrost environments is lacking due to limited field data and appropriate models. This knowledge gap limits projections of climate change impacts to coastal Arctic ecosystems and communities. In this study, we develop and use a mathematical model that incorporates multiple, interrelated processes, including how salt content affects the freezing temperature of water, to evaluate the impacts of sea‐level rise and associated saltwater intrusion on coastal permafrost. Results show that sea‐level rise causes saltwater to intrude into the unfrozen pore space of permafrost. With a lower freezing temperature than freshwater, saltwater intrusion triggers permafrost thaw and lateral retreat. The combination of atmospheric and oceanic warming and sea‐level rise has the potential to drive extensive permafrost loss along Arctic coastlines. Key Points: Newly developed cryohydrogeological model incorporates salinity‐dependent freeze‐thaw processes and solute exclusionSubsurface saltwater intrusion drives lateral thaw of coastal ice‐saturated permafrostCombination of sea‐level rise and land surface warming results in extensive ice‐saturated permafrost loss [ABSTRACT FROM AUTHOR]

Freshwater lenses underlying small ocean islands exhibit spatial variability and temporal fluctuations in volume, influencing ecologic management. For example, The Palmyra Atoll National Wildlife Refuge harbors one of the few surviving native stands of Pisonia grandis in the central Pacific Ocean, yet these trees face pressure from groundwater salinization, with little basic groundwater data to guide decision making. Adding to natural complexity, the geology of Palmyra was heavily altered by dredge and fill activities. Our study based at this atoll combines geophysical and hydrological field measurements from 2008 to 2019 with groundwater modeling to study the drivers of observed freshwater lens dynamics. Electromagnetic induction (EMI) field data were collected on the main atoll islands over repeat transects in 2008 following 'strong' La Niña conditions (wet) and in 2016 during 'very strong' El Niño conditions (dry). Shallow monitoring wells were installed adjacent to the geophysical transects in 2013 and screened within the fresh/saline groundwater transition zone. Temporal EMI and monitoring well data showed a strong contraction of the freshwater lens in response to El Niño conditions, and indicated a thicker lens toward the ocean side, an opposite spatial pattern to that observed for many other Pacific islands. On an outer islet where a stand of mature Pisonia trees exist, EMI surveys revealed only a thin (<3 m from land surface) layer of brackish groundwater during El Niño. Numerical groundwater simulations were performed for a range of permeability distributions and climate conditions at Palmyra. Results revealed that the observed atypical lens asymmetry is likely due to more efficient submarine groundwater discharge on the lagoon side as a result of lagoon dredging and filling with high-permeability material. Simulations also predict large decreases (40%) in freshwater lens volume during dry cycles and highlight threats to the Pisonia trees, yielding insight for atoll ecosystem management worldwide. Unlabelled Image • Limited fresh groundwater lenses support surface ecology on small atolls. • Human alteration of geology and natural climate cycles may impact lens dynamics. • Numerical modeling was used to explain unexpected geophysical field data. • Asymmetric lens weighting toward the ocean side driven by dredge and fill activities. • Small-island groundwater is highly sensitive to climate cycles, influencing surface ecology. [ABSTRACT FROM AUTHOR]

Global social and economic changes, alongside climate change, are affecting the operating environment for agriculture, leading to efforts to increase production and yields, typically through the use of agrochemicals like pesticides and fertilizers, expanded irrigation, and changes in seed varieties. Intensification, alongside the expansion of agriculture into new areas, has increased harvest, but has also had numerous well-known impacts on the environment, ultimately resulting in a loss of resilience and lack of sustainability in agro-ecosystems. Combined with features of agricultural systems such as the differential movement of ecosystem services, and interactions among ecosystem services driven in part by management choices, such intensification has disrupted key feedbacks in agricultural systems. These changes have tended to perpetuate the management choices that have led to efficient, productive agriculture, often at the expense of nature and the provision of important nonfood ecosystem services. Here, we explore how agriculture functions as a complex adaptive system. We assess how recent changes have interacted with agro-ecosystem features to result in a loss of resilience, and suggest key research directions to help harmonize production and ecosystem function, drawing primarily on Canadian examples. Enhancing the resilience of agricultural landscapes is critical to the long-term sustainability of agriculture in a rapidly changing world. [ABSTRACT FROM AUTHOR]

Recent waste rock pile designs have been proposed to incorporate a fine-grained layer to create a capillary barrier to prevent surface water from draining into the pile interior. This study analyses active fibre optic distributed temperature sensing (FO-DTS) as a tool to measure the effectiveness a capillary barrier system following an infiltration test. A laboratory waste rock column was built with anorthosite waste rock overlain by sand. Volumetric water content is calculated during heat cycles lasting 15 min powered at 15 W/m in the column. A new algorithm is employed to circumvent several requirements for soil specific calibration. The inferred moisture contents were verified by soil moisture probes located adjacent to the cable. The FO-DTS data indicate, at vertical resolutions up to 2 cm, that water is retained in the sand and does not drain into the anorthosite following the infiltration test. The coefficient of determination, R2, between the inferred and measured volumetric water content in the fine cover sand layer is 0.90, while the screened anorthosite maintained an R2 of 0.94 with constant moisture content throughout the test. This study will ultimately help guide future waste rock storage design initiatives incorporating fibre optic sensors, leading to improved environmental mine waste management. [ABSTRACT FROM AUTHOR]

Streams strongly influenced by groundwater discharge may serve as “climate refugia” for sensitive species in regions of increasingly marginal thermal conditions. The main goal of this study is to develop paired air and stream water annual temperature signal analysis techniques to elucidate the relative groundwater contribution to stream water and the effective groundwater flowpath depth. Groundwater discharge to streams attenuates surface water temperature signals, and this attenuation can be diagnostic of groundwater gaining systems. Additionally, discharge from shallow groundwater flowpaths can theoretically transfer lagged annual temperature signals from aquifer to stream water. Here we explore this concept using multi-year temperature records from 120 stream sites located across 18 mountain watersheds of Shenandoah National Park, VA, USA and a coastal watershed in Massachusetts, USA. Both areas constitute important cold-water habitat for native brook trout ( Salvelinus fontinalis ). Observed annual temperature signals indicate a dominance of shallow groundwater discharge to streams in the National Park, in contrast to the coastal watershed that has strong, apparently deeper, groundwater influence. The average phase lag from air to stream signals in Shenandoah National Park is 11 d; however, extended lags of approximately 1 month were observed in a subset of streams. In contrast, the coastal stream has pronounced attenuation of annual temperature signals without notable phase lag. To better understand these observed differences in signal characteristics, analytical and numerical models are used to quantify mixing of the annual temperature signals of surface and groundwater. Simulations using a total heat budget numerical model indicate groundwater-induced annual temperature signal phase lags are likely to show greater downstream propagation than the related signal amplitude attenuation. The measurement of multi-seasonal paired air and water temperatures offers great promise toward understanding catchment processes and informing current cold-water habitat management at ecologically-relevant scales. [ABSTRACT FROM AUTHOR]

• SEAWAT is used to model coastal groundwater sensitivity to climate change. • Groundwater inundates up to 27% of OWTS in modelled climate change scenario. • Saltwater-freshwater interface movement depends on climate change scenario. • Recommendations for climate-resilient septic system guidelines are proposed. An often-overlooked impact of rising sea levels is the concurrent rise in coastal groundwater tables, which can result in subsurface inundation of below-ground infrastructure. This is an important consideration for rural coastal communities, many of which rely on on-site wastewater treatment systems (OWTS). Subsurface inundation and flooding of OWTS leads to treatment performance issues, potentially resulting in enhanced contaminant transport to coastal groundwater resources and the ocean via submarine groundwater discharge. We use SEAWAT to develop a variable-density groundwater flow model for the groundwater system underlying a densely populated rural community on the north shore of Nova Scotia, Canada. We then impose various climate change scenarios (sea-level rise, changes in recharge) to evaluate the risk of OWTS inundation from rising groundwater. We find that under current conditions as many as 9 % of OWTS in this small but densely populated watershed are either inundated or completely flooded. This number that could grow to 27 % of OWTS for the climate change scenario with the highest recharge and sea-level rise. As a secondary objective, we track the location of the modeled saltwater-freshwater interface, and investigate the potential salinization of groundwater resources used for drinking water supply. The modeled interface moved landward by ≤ 20 m and proved to be less of a concern than OWTS inundation, except for shoreline dwellings. We also use the modelling results to develop adaptation recommendations for OWTS installation regulations. This research contributes to an increasing number of groundwater modelling studies focusing on the impacts of sea-level rise on coastal subsurface infrastructure and provides important insight for rural coastal communities. [ABSTRACT FROM AUTHOR]

Aims: This study aimed at comparing the cerebral cytotoxicity of ethanol and its main metabolite acetaldehyde after acute or chronic exposures of rat astrocytes in primary culture. Methods: Cytotoxicity was evaluated on the cell reduction of viability (MTT reduction test) and on the characterization of DNA damage by single cell gel electrophoresis (or comet assay). Results: Changes in astrocyte survival and in DNA integrity only occurred when the astrocytes were chronically exposed to ethanol (20 mM; 3,6 or 9 days). On the other hand, viability and DNA integrity were deeply affected by acute exposure to acetaldehyde. Both effects were dependent on the concentration of acetaldehyde. The cytotoxic effect of acetaldehyde was also indirectly evaluated after modifications of the normal ethanol metabolism by the use of different inducers or inhibitors. In presence of ethanol, the concomitant induction of catalase (i.e. by glucose oxidase) and inhibition of aldehyde dehydrogenase (i.e. by methylene blue) led to acetaldehyde accumulation within cells. It was followed by both a reduction in viability and a substantial increase in DNA strand breaks. Conclusions: These data were thus consistent with a possible predominant role of acetaldehyde during brain ethanol metabolism. On the other hand, the effects observed after AMT could also suggest a possible direct ethanol effect and a role for free radical attacks. These data were thus consistent with a possible predominant role of acetaldehyde during brain ethanol metabolism. On the other hand, the effects observed after AMT could also suggest a possible direct ethanol effect and a role for free radical attacks. [ABSTRACT FROM AUTHOR]

Objectives: To describe the priors and decision thresholds in phase 2 and 3 randomized controlled trials (RCTs) evaluating drug efficacy using Bayesian methods., Study Design and Setting: A systematic review of phase 2 and 3 RCTs evaluating drug efficacy through Bayesian inference was conducted across the MEDLINE, EMBASE, and Cochrane databases, with no date restrictions until September 2022. The type of prior used for the analysis of the primary endpoint and its characteristics (type and parameters of the distribution, justification, and sensitivity analysis), the use of a posterior probability decision threshold defined a priori, and its value, were extracted., Results: From 1161 articles screened, 69 articles were ultimately included, encompassing a total of 91 comparisons, as some trials assessed multiple primary endpoints or treatments. The prior was assigned to treatment effect in 51% of the cases (n = 46) to each arm in 37% (n = 34) and was not explicitly defined in 12% (n = 11). Prior distribution was described (with its parameters) in 59% of cases (n = 54). A decision threshold was set a priori in 68% of the results (n = 62), and its value ranged from 70% to 99% (median 95%)., Conclusion: The inconsistent description of priors, along with the wide variation and occasional absence of decision thresholds, underscore the need for clear guidelines on the use and reporting of Bayesian methods., Plain Language Summary: Bayesian methods are being used more frequently in clinical trials to assess drug's efficacy. These methods offer flexibility by incorporating prior knowledge into the analysis. However, the use of Bayesian approaches is still not widespread, and there are challenges with how results are interpreted, partly due to a lack of clear standards. We conducted a systematic review to describe how Bayesian methods were used and reported in phase 2 and 3 clinical trials assessing drug efficacy. We looked at the types of prior information used in the analyses and how decisions about efficacy were made based on the results. Out of 1161 studies reviewed, 69 were included in the analysis, covering 91 drug comparisons. The priors in Bayesian drug trials were assigned to treatment effects or each arm, often justified but variably describe. Similarly, decision thresholds for determining drug efficacy were preset in most studies, but with heterogeneity in the thresholds used to conclude. Our findings highlight the need for clearer guidelines on using Bayesian methods in clinical trials to improve transparency and consistency in how results are reported., Competing Interests: Declaration of competing interest None declared. All authors have completed the ICMJE uniform disclosure form at http://www.icmje.org/disclosure-of-interest/and declare: no support from any organization for the submitted work; no financial relationships with any organisations that might have an interest in the submitted work in the previous 3 years; no other relationships or activities that could appear to have influenced the submitted work., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Objectives: To evaluate whether there is evidence of efficacy of the most commonly used medications in their primary indications., Study Design and Setting: This scoping review was executed using the Cochrane Library and MEDLINE databases up to May 2023. The 10 most prescribed medications in England, France, and the United States were identified using country-specific public databases. Up to 3 common indications in primary care were defined for each medication, based on a survey of general practitioners. The outcomes were determined by the authors to be patient-important outcomes, with placebo as the comparator. Two investigators independently conducted searches, following a predefined algorithm, to identify randomized controlled trials or meta-analyses of randomized controlled trials assessing the efficacy of these medications for each indication. The risk of bias was assessed using the ROBIS or ROB 2.0 tools., Results: We identified 21 drugs, covering 56 indications and 114 outcomes. Sixty-seven percent of the evaluated medications demonstrated efficacy for at least one outcome in at least one of the sought indications. Overall, evidence of efficacy was found for 48% of the indications. There was no study evaluating the efficacy of amoxicillin and salbutamol. For other drugs such as phloroglucinol or cholecalciferol, available studies suggested an absence of efficacy in the most common indications., Conclusion: This study underscores the lack of data regarding the level of evidence for the most prescribed medications. Limitations include the choice of outcomes, and the understanding that the absence of evidence is not synonymous with the absence of efficacy., Competing Interests: Declaration of competing interest All authors have completed the ICMJE uniform disclosure form at http://www.icmje.org/disclosure-of-interest/and declare: no support from any organization for the submitted work, no financial relationships with any organizations that might have an interest in the submitted work in the previous 3 years, and no other relationships or activities that could appear to have influenced the submitted work., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Estuaries are vulnerable to oceanic and atmospheric climate change. Much of the research investigating climate change impacts on estuaries is focused on saltwater intrusion within surface water due to drought and rising sea levels, with implications for ecosystems and humans. Groundwater and soil near estuaries may also be influenced, as estuary salinity and hydraulic head changes can impact soils and aquifers not previously at risk of salinization. This study was conducted to address knowledge gaps related to present and future groundwater salinity distribution in a groundwater system connected to a macro-tidal estuary. The studied estuary experiences a tidal bore due to its hydraulic connection to the Bay of Fundy in Nova Scotia, Canada. A parcel of agricultural land adjacent to the estuary was selected to assess the groundwater response to episodic fluctuations in estuary water levels and salinity. Groundwater monitoring and electromagnetic surveys were conducted to map soil and groundwater salinity patterns. A numerical model of groundwater flow and solute transport informed by field data was used to investigate how varying estuary salinity due to droughts and sea-level rise could impact groundwater salinity. Results showed that, in contrast to salt wedges observed along marine coasts, the saline groundwater existed as a plume immediately around the estuary. Model simulations showed that short-term droughts had an insignificant impact on the adjacent groundwater salinity. However, permanent increases in salinity caused by sea-level rise increased the plume volume by 86 %, or an additional ∼11 m horizontally and ∼ 4.5 m vertically. Our results suggest that increased river salinity in this setting would not result in widespread salinization of porewater and agricultural soils, but more extensive salinization may be experienced in permeable aquifers or along more saline estuarine zones. Findings may inform land management decisions in regions exposed to increased salinity in the future., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Coastal defense structures (e.g., dikes, seawalls) protect vulnerable communities along marine coastlines and estuaries from the physical and chemical influences of adjacent water bodies. These structures are susceptible to overtopping or breaching by tides and waves, with risks amplified by climate change-induced sea-level rise. Repeated inundation by saline water can contaminate freshwater resources and salinize soil, impacting land-use activities, including agricultural productivity. Managed ecosystem-based dike realignment and salt marsh restoration can provide alternatives to traditional coastal adaptation approaches. We assess the changes to soil salinity at a managed dike realignment project prior to the transformation from a diked terrestrial environment to an estuarine environment. Baseline data are compared to conditions following 8-10 months of intermittent flooding at spring tides. Results show that an increase in salinity occurred over the entire site in the shallow subsurface, with the most significant contamination occurring in low-lying areas. Bulk soil electrical conductivity (salinity proxy) measured from geophysical surveys increased from the previous freshwater condition of ∼300 μS/cm to over 6000 μS/cm following <20 flood events, while successive flooding resulted in increased soil moisture as infiltrated floodwater propagated to greater depths. Sediment deposition occurred at high rates, with up to 4 cm of sediment deposited per flood, converting much of the previously cultivated land into tidal mudflats. Deeper sediments and groundwater (i.e., >1.8 m depth) were not impacted over the time scale of this research. This study demonstrates that intermittent shallow flooding can rapidly increase moisture content and soil salinity in surficial sediments and, in turn, adversely impact conditions suitable for agricultural crop production. The realignment zone serves as an engineered analog of coastal flooding, presenting an opportunity to investigate how low-lying coastal environments may experience regular flooding in the future due to sea-level rise and intensifying coastal storms., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

The homogeneous labeling of antibodies and their fragments is a critical step for the generation of robust probes used in immuno-detection applications. To date, numerous chemical, genetic and peptide-based site-specific coupling methods have been developed. Among these methods, co-assembling peptide-tags is one of the most straightforward and versatile solutions. Here, we describe site-specific labeling of nanobodies through the use of two co-associating peptides tags, E3 and K3, originating from the tetramerization domain of p53. These E3 and K3-tags provide a simple and robust method for associating stoichiometric amount of V H H and fluorescent probes, either fluorescent proteins or fluorochromes, at specific positions. As a proof of concept, a nanobody targeting the human epidermal growth factor receptor 2 (HER2), the nano-HER2 was genetically fused to the E3 and associated with different fluorescent K3-derivates. Entities were produced separately in Escherichia coli in soluble forms at high yields and co-assembled in vitro. These molecular probes present high binding specificity on HER2-overexpressing cells in flow-cytometry with relative binding constants in the low nanomolar range and are stable enough to stain HER2-receptor on living cells followed detection using fluorescent confocal microscopy. Altogether, our results demonstrate that the non-covalent conjugation method using these two co-associating peptides can be easily implemented for the modular engineering of molecular probes for cell immuno-staining.

Despite the global interest in green energy alternatives, little attention has focused on the large-scale viability of recycling the ground heat accumulated due to urbanization, industrialization and climate change. Here we show this theoretical heat potential at a multi-continental scale by first leveraging datasets of groundwater temperature and lithology to assess the distribution of subsurface thermal pollution. We then evaluate subsurface heat recycling for three scenarios: a status quo scenario representing present-day accumulated heat, a recycled scenario with ground temperatures returned to background values, and a climate change scenario representing projected warming impacts. Our analyses reveal that over 50% of sites show recyclable underground heat pollution in the status quo, 25% of locations would be feasible for long-term heat recycling for the recycled scenario, and at least 83% for the climate change scenario. Results highlight that subsurface heat recycling warrants consideration in the move to a low-carbon economy in a warmer world., (© 2022. The Author(s).)

Membrane proteins (MPs) comprise about one-third of the human proteome, playing critical roles in many physiological processes and associated disorders. Consistently, they represent one of the largest classes of targets for the pharmaceutical industry. Their study at the molecular level is however particularly challenging, resulting in a severe lack of structural and dynamic information that is hindering their detailed functional characterization and the identification of novel potent drug candidates.Magic Angle Spinning (MAS) NMR is a reliable and efficient method for the determination of protein structures and dynamics and for the identification of ligand binding sites and equilibria. MAS-NMR is particularly well suited for MPs since they can be directly analysed in a native-like lipid bilayer environment but used to require aggravating large amounts of isotope enriched material. The frequent toxicity of human MP overexpression in bacterial cultures poses an additional hurdle, resulting in the need for alternative (and often more costly) expression systems. The recent development of very fast (up to 150 kHz) MAS probes has revolutionized the field of biomolecular solid-state NMR enabling higher spectral resolution with significant reduction of the required sample, rendering eukaryotic expression systems cost-effective.Here is presented a set of accessible procedures validated for the production and preparation of eukaryotic MPs for Fast-MAS 1 H-detected NMR analysis. The methodology is illustrated with the human copper uptake protein hCTR1 recombinantly produced and 13 C- 15 N uniformly labeled with the versatile and affordable Pichia pastoris system. Subsequent purification procedures allow the recovery of mg amounts that are then reconstituted into liposome formulations compatible with solid-state NMR handling and analysis., (© 2022. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Immunotoxins are emerging candidates for cancer therapeutics. These biomolecules consist of a cell-targeting protein combined to a polypeptide toxin. Associations of both entities can be achieved either chemically by covalent bonds or genetically creating fusion proteins. However, chemical agents can affect the activity and/or stability of the conjugate proteins, and additional purification steps are often required to isolate the final conjugate from unwanted byproducts. As for fusion proteins, they often suffer from low solubility and yield. In this report, we describe a straightforward conjugation process to generate an immunotoxin using coassociating peptides (named K3 and E3), originating from the tetramerization domain of p53. To that end, a nanobody targeting the human epidermal growth factor receptor 2 (nano-HER2) and a protein toxin fragment from Pseudomonas aeruginosa exotoxin A (TOX) were genetically fused to the E3 and K3 peptides. Entities were produced separately in Escherichia coli in soluble forms and at high yields. The nano-HER2 fused to the E3 or K3 helixes (nano-HER2-E3 and nano-HER2-K3) and the coassembled immunotoxins (nano-HER2-K3E3-TOX and nano-HER2-E3K3-TOX) presented binding specificity on HER2-overexpressing cells with relative binding constants in the low nanomolar to picomolar range. Both toxin modules (E3-TOX and K3-TOX) and the combined immunotoxins exhibited similar cytotoxicity levels compared to the toxin alone (TOX). Finally, nano-HER2-K3E3-TOX and nano-HER2-E3K3-TOX evaluated on various breast cancer cells were highly potent and specific to killing HER2-overexpressing breast cancer cells with IC 50 values in the picomolar range. Altogether, we demonstrate that this noncovalent conjugation method using two coassembling peptides can be easily implemented for the modular engineering of immunotoxins targeting different types of cancers.

In the presence of ciprofloxacin (CPFX), free radical adduct formation was demonstrated in rat cerebral microsomes using a spin trap α-(4-pyridyl-1-oxide)-N- tert -butyl-nitrone by electron spin resonance spectroscopy. Active microsomes, dihydronicotinamide-adenine dinucleotide phosphate, and ciprofloxacin were necessary for the formation of a spin trap/radical adduct. Adduct formation increased dose-dependently at 0.5-1 mM CPFX concentration for 180 min, and 0.3-1 mM concentration level for 240 min. The addition of SKF 525A, ZnCl 2 or desferrioxamine to the incubation system caused complete inhibition of the radical formation. However, pretreatment of microsomal system with superoxide dismutase (SOD) did not induce any protective effect. Induction of lipid peroxidation, and depletion of thiol levels by CPFX were also shown in the system. These results strongly suggested that CPFX produces free radical(s) in the cerebral microsomes of rats.

Cocaine abuse induces brain injury and neurodegeneration by a mechanism that has not yet been fully elucidated. Mitochondria play a key role in cell death processes, notably through the opening of the permeability transition pore (PTP). In this work, we examined the involvement of the PTP in cocaine-induced toxicity in PC12 cell lines. We used two different PTP inhibitors -i.e. cyclosporin A (CsA) and metformin-to assess their ability to counteract the cocaine induced effects. We first observed that a 48 h exposure to cocaine strongly sensitized cells to calcium overload, as measured by the calcium retention capacity. CsA and metformin significantly decreased the cocaine-induced PTP opening sensitization. We next showed by confocal microscopy that cocaine induced a permanent PTP opening in intact living cells, a phenomenon characterized by the collapse of the mitochondrial membrane potential and the relocation of the NAD(P)H from the mitochondrial matrix to the cytosol. As expected, a cocaine-induced PTP opening was prevented by PTP inhibitors. Finally, a flow cytometry analysis revealed that cocaine induced cell death while CsA and metformin promoted cell survival. Our results demonstrate that cocaine induces PC12 cell death through a mechanism involving permanent PTP opening., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Obtaining reliable estimates of vertical groundwater flows remains a challenge but is of critical importance to the management of groundwater resources. When large scale land clearing or groundwater extraction occurs, methods based on water table fluctuations or water chemistry are unreliable. As an alternative, a number of methods based on temperature-depth (T-z) profiles are available to provide vertical groundwater flow estimates from which recharge rates may be calculated. However, methods that invoke steady state assumptions have been shown to be inappropriate for sites that have experienced land surface warming. Analytical solutions that account for surface warming are available, but they typically include unrealistic or restrictive assumptions (e.g. no flow initial conditions or linear surface warming). Here, we use a new analytical solution and associated computer program (FAST) that provides flexible initial and boundary conditions to estimate fluxes using T-z profiles from the Willunga Super Science Site, a complex, but densely instrumented groundwater catchment in South Australia. T-z profiles from seven wells (ranging from high elevation to near sea level) were utilised, in addition to mean annual air temperatures at nearby weather stations to estimate boundary conditions, and thermal properties were estimated from down borehole geophysics. Temperature based flux estimates were 5 to 23mmy -1 , which are similar to those estimated using chloride mass balance. This study illustrates that T-z profiles can be studied to estimate recharge in environments where more commonly applied methods fail., (Copyright © 2016 Elsevier B.V. All rights reserved.)

AMP-activated protein kinase (AMPK) and cytosolic brain-type creatine kinase (BCK) cooperate under energy stress to compensate for loss of adenosine triphosphate (ATP) by either stimulating ATP-generating and inhibiting ATP-consuming pathways, or by direct ATP regeneration from phosphocreatine, respectively. Here we report on AMPK-dependent phosphorylation of BCK from different species identified by in vitro screening for AMPK substrates in mouse brain. Mass spectrometry, protein sequencing, and site-directed mutagenesis identified Ser6 as a relevant residue with one site phosphorylated per BCK dimer. Yeast two-hybrid analysis revealed interaction of active AMPK specifically with non-phosphorylated BCK. Pharmacological activation of AMPK mimicking energy stress led to BCK phosphorylation in astrocytes and fibroblasts, as evidenced with a highly specific phospho-Ser6 antibody. BCK phosphorylation at Ser6 did not affect its enzymatic activity, but led to the appearance of the phosphorylated enzyme at the endoplasmic reticulum (ER), close to the ER calcium pump, a location known for muscle-type cytosolic creatine kinase (CK) to support Ca²⁺-pumping., (Copyright © 2014 Elsevier B.V. All rights reserved.)