High-resolution visualisation of antisense oligonucleotide release from polymers in cells.

Antisense oligonucleotides (ASOs) are a well-established therapeutic modality based on RNA interference, but low cellular uptake, limited ability to direct ASO trafficking, and a range of intracellular barriers to successful activity compromise both gene silencing outcomes and clinical translations. Herein, we demonstrate that polymers can increase ASO internalisation via intracellular trafficking pathways that are distinct from lipid-based delivery reagents. For the first time, we spatially define internalisation and dissociation stages in the polymer-mediated cytosolic delivery of ASOs using Nanoscale Secondary Ion Mass Spectrometry (NanoSIMS), which enables visualisation of ASO localisation at the organelle level. We find that polymer-ASO complexes are imported into cells, from which free ASO enters the cytosol following complex dissociation. This information enables a better understanding of the intracellular trafficking pathways of nucleic acid therapeutics and may be exploited for therapeutic delivery to enhance the effectiveness of nucleic acid therapeutics in the future.

Dynamic primary resources, not just wild prey availability, underpin lion depredation of livestock in a savanna ecosystem.

Because it can lead to retaliatory killing, livestock depredation by large carnivores is among the foremost threats to carnivore conservation, and it severely impacts human well-being worldwide. Ongoing climate change can amplify these human-wildlife conflicts, but such issues are largely unexplored, though are becoming increasingly recognized. Here, we assessed how the availability of primary resources and wild prey interact to shape large carnivore selection for livestock rather than wild prey (i.e., via prey switching or apparent competition). Specifically, we combined remotely sensed estimates of primary resources (i.e., water availability and primary productivity), wild prey movement, and 7 years (2015-2021) of reports for livestock depredation by African lions (Panthera leo) in the Makgadikgadi Pans ecosystem, Botswana. Although livestock depredation did not vary between wet versus dry seasons, analyses at finer temporal scales revealed higher incidences of livestock depredation when primary production, water availability, and wild prey availability were lower, though the effects of wild prey availability were mediated by water availability. Increased precipitation also amplified livestock depredation events despite having no influence on wild prey availability. Our results suggest that livestock depredation is influenced by the diverse responses of livestock, wild prey, and lions to primary resource availability, a driver that is largely overlooked or oversimplified in studies of human-carnivore conflict. Our findings provide insight into tailoring potential conflict mitigation strategies to fine-scale changes in resource conditions to efficiently reduce conflict and support human livelihoods.

"Because that is the right thing to do": A focus group study of Australian expert perspectives on offering smoking cessation support in lung cancer screening.

INTRODUCTION: Lung cancer screening (LCS) trials, targeting people with smoking history, have demonstrated reduced mortality. How to optimally embed evidence-based smoking cessation support in LCS, including in Australia, needs to be better understood. We sought experts' perspectives to identify potential barriers and effective implementation strategies. METHODS: Perceptions of providing smoking cessation support in LCS were elicited in 24 focus groups and three individual interviews with clinicians, cancer screening program managers/policymakers, and researchers during 2021. We conducted framework analysis and mapped key topics to the updated Consolidated Framework for Implementation Research. RESULTS: Experts (N=84 participants) strongly supported capitalising on an "opportune time" for smoking cessation and new LCS participant contact opportunities throughout the screening and assessment pathway. Many advocated for adapting existing cessation resources to the LCS setting and providing support without participant costs. Experts generally considered referral alone to established programs (e.g., telephone Quitline) as insufficient, but likely helpful in follow-up, and dedicated cessation specialist roles as essential. Broader cessation messaging (via mass media/community channels) was also suggested to reinforce individualised support. Experts described inherent alignment, and an ethical responsibility, to deliver smoking cessation as a core LCS component. It was suggested that LCS-eligible participants' varied experiences of stigma, health literacy, and motivation, be considered in cessation supports. Primary care support and individualised interventions were suggested to facilitate implementation. CONCLUSIONS: Experts considered smoking cessation support essential in LCS. The expert-identified and multi-level implementation strategies described here can directly inform smoking cessation-specific planning for Australia's forthcoming National LCS Program. IMPLICATIONS: The international literature includes few examples considering how best to provide smoking cessation support within a lung cancer screening (LCS) program in advance of program commencement. Our analysis, using the updated Consolidated Framework for Implementation Research, is one of the first to explore experts' perspectives within this context. Experts identified multiple implementation barriers to providing smoking cessation support within and outside of an Australian LCS program, including key work infrastructure barriers, and advocated for providing tailored interventions within this program. Our foundational work in a new targeted screening program's pre-implementation phase will allow international comparisons to be made.

Engineering Planar Gram-Negative Outer Membrane Mimics Using Bacterial Outer Membrane Vesicles.

Antibiotic resistance is a major challenge in modern medicine. The unique double membrane structure of gram-negative bacteria limits the efficacy of many existing antibiotics and adds complexity to antibiotic development by limiting transport of antibiotics to the bacterial cytosol. New methods to mimic this barrier would enable high-throughput studies for antibiotic development. In this study, we introduce an innovative approach to modify outer membrane vesicles (OMVs) from Aggregatibacter actinomycetemcomitans, to generate planar supported lipid bilayer membranes. Our method first involves the incorporation of synthetic lipids into OMVs using a rapid freeze-thaw technique to form outer membrane hybrid vesicles (OM-Hybrids). Subsequently, these OM-Hybrids can spontaneously rupture when in contact with SiO2 surfaces to form a planar outer membrane supported bilayer (OM-SB). We assessed the formation of OM-Hybrids using dynamic light scattering and a fluorescence quenching assay. To analyze the formation of OM-SBs from OM-Hybrids we used quartz crystal microbalance with dissipation monitoring (QCM-D) and fluorescence recovery after photobleaching (FRAP). Additionally, we conducted assays to detect surface-associated DNA and proteins on OM-SBs. The interaction of an antimicrobial peptide, polymyxin B, with the OM-SBs was also assessed. These findings emphasize the capability of our platform to produce planar surfaces of bacterial outer membranes, which in turn, could function as a valuable tool for streamlining the development of antibiotics.

Comparing suicide risk screening strategies in Spanish-speaking pediatric patients.

BACKGROUND: Suicide and suicidal behaviors pose significant global public health challenges, especially among young individuals. Effective screening strategies are crucial for addressing this crisis, with depression screening and suicide-specific tools being common approaches. This study compares their effectiveness by evaluating the Ask Suicide-Screening Questions (ASQ) against item 9 of the Patient Health Questionnaire-A (PHQ-A). METHODS: This study is a secondary analysis of the Argentinean-Spanish version of the ASQ validation study, an observational, cross-sectional, and multicenter study conducted in medical settings in Buenos Aires, Argentina. A convenience sample of pediatric outpatients/inpatients aged 10 to 18 years completed the ASQ, PHQ-A, and Suicide Ideation Questionnaire (SIQ) along with clinical and sociodemographic questions. RESULTS: A sample of 267 children and adolescents were included in this secondary analysis. Results show that the ASQ exhibited higher sensitivity (95.1%; 95% CI: 83% - 99%) compared to PHQ-A item 9 (73.1%; 95% CI: 57% - 85%), and superior performance in identifying suicide risk in youth. LIMITATIONS: The study included a convenience sampling and was geographically restricted to Buenos Aires, Argentina. The study also lacked longitudinal follow-up to assess the predictive validity of these screening tools for suicide risk. CONCLUSION: The study highlights the ASQ's effectiveness in identifying suicide risk among youth, emphasizing the importance of specialized screening tools over depression screening tools alone for accurate risk assessment in this population.

Re-entrant percolation in active Brownian hard disks.

Non-equilibrium clustering and percolation are investigated in an archetypal model of two-dimensional active matter using dynamic simulations of self-propelled Brownian repulsive particles. We concentrate on the single-phase region up to moderate levels of activity, before motility-induced phase separation (MIPS) sets in. Weak activity promotes cluster formation and lowers the percolation threshold. However, driving the system further out of equilibrium partly reverses this effect, resulting in a minimum in the critical density for the formation of system-spanning clusters and introducing re-entrant percolation as a function of activity in the pre-MIPS regime. This non-monotonic behaviour arises from competition between activity-induced effective attraction (which eventually leads to MIPS) and activity-driven cluster breakup. Using an adapted iterative Boltzmann inversion method, we derive effective potentials to map weakly active cases onto a passive (equilibrium) model with conservative attraction, which can be characterised by Monte Carlo simulations. While the active and passive systems have practically identical radial distribution functions, we find decisive differences in higher-order structural correlations, to which the percolation threshold is highly sensitive. For sufficiently strong activity, no passive pairwise potential can reproduce the radial distribution function of the active system.

The value of error-correcting responses for cognitive assessment in games.

Traditional conflict-based cognitive assessment tools are highly behaviorally restrictive, which prevents them from capturing the dynamic nature of human cognition, such as the tendency to make error-correcting responses. The cognitive game Tunnel Runner measures interference control, response inhibition, and response-rule switching in a less restrictive manner than traditional cognitive assessment tools by giving players movement control after an initial response and encouraging error-correcting responses. Nevertheless, error-correcting responses remain unused due to a limited understanding of what they measure and how to use them. To facilitate the use of error-correcting responses to measure and understand human cognition, we developed theoretically-grounded measures of error-correcting responses in Tunnel Runner and assessed whether they reflected the same cognitive functions measured via initial responses. Furthermore, we evaluated the measurement potential of error-correcting responses. We found that initial and error-correcting responses similarly reflected players' response inhibition and interference control, but not their response-rule switching. Furthermore, combining the two response types increased the reliability of interference control and response inhibition measurements. Lastly, error-correcting responses showed the potential to measure response inhibition on their own. Our results pave the way toward understanding and using post-decision change of mind data for cognitive measurement and other research and application contexts.

ClearScope: a fully integrated light sheet theta microscope for sub-cellular resolution imaging without lateral size constraints.

Three-dimensional (3D) ex vivo imaging of cleared intact brains of animal models and large human and non-human primate postmortem brain specimens is important for understanding the physiological neural network connectivity patterns and the pathological alterations underlying neuropsychiatric and neurological disorders. Light-sheet microscopy has emerged as a highly effective imaging modality for rapid high-resolution imaging of large cleared samples. However, the orthogonal arrangements of illumination and detection optics in light sheet microscopy limits the size of specimen that can be imaged. Recently developed light sheet theta microscopy (LSTM) technology addressed this by utilizing a unique arrangement of two illumination light paths oblique to the detection light path, while allowing perpendicular arrangement of the detection light path relative to the specimen surface. Here, we report development of a next-generation, fully integrated, and user-friendly LSTM system for rapid sub-cellular resolution imaging uniformly throughout a large specimen without constraining the lateral (XY) size. In addition, we provide a seamlessly integrated workflow for image acquisition, data storage, pre- and post-processing, enhancement, and quantitative analysis. We demonstrate the system performance by high-resolution 3D imaging of intact mouse brains and human brain samples, and complete data analysis including digital neuron tracing, vessel reconstruction and design-based stereological analysis in 3D. This technically enhanced and user-friendly LSTM implementation will enable rapid quantitative mapping of molecular and cellular features of interests in diverse types of very large samples.

Robot-Assisted Spine Surgery: The Pearls and Pitfalls.

Robot-assisted spine surgery has gained notable popularity among surgeons because of recent advancements in technology. These innovations provide several key benefits, including high screw accuracy rates, reduced radiation exposure, customized preoperative and intraoperative planning options, and improved ergonomics for surgeons. Despite the promising outcomes reported in literature, potential technical challenges remain across various robotic platforms. It is crucial for surgeons to remember that robotic platforms are shared-control systems, requiring the surgeon to maintain primary control throughout the procedure. To ensure patient safety, surgeons should be well versed in common technical pitfalls and strategies to mitigate these limitations.

An intrinsic mechanism of metabolic tuning promotes cardiac resilience to stress.

Defining the molecular mechanisms underlying cardiac resilience is crucial to find effective approaches to protect the heart. A physiologic level of ROS is produced in the heart by fatty acid oxidation, but stressful events can boost ROS and cause mitochondrial dysfunction and cardiac functional impairment. Melusin is a muscle specific chaperone required for myocardial compensatory remodeling during stress. Here we report that Melusin localizes in mitochondria where it binds the mitochondrial trifunctional protein, a key enzyme in fatty acid oxidation, and decreases it activity. Studying both mice and human induced pluripotent stem cell-derived cardiomyocytes, we found that Melusin reduces lipid oxidation in the myocardium and limits ROS generation in steady state and during pressure overload and doxorubicin treatment, preventing mitochondrial dysfunction. Accordingly, the treatment with the lipid oxidation inhibitor Trimetazidine concomitantly with stressful stimuli limits ROS accumulation and prevents long-term heart dysfunction. These findings disclose a physiologic mechanism of metabolic regulation in the heart and demonstrate that a timely restriction of lipid metabolism represents a potential therapeutic strategy to improve cardiac resilience to stress.

Relationship Between Neighborhood-Level Socioeconomic Status and Functional Outcomes After Hip Arthroscopy.

BACKGROUND: Despite the growing volume of neighborhood-level health disparity research, there remains a paucity of prospective studies investigating the relationship between Area Deprivation Index (ADI) and functional outcomes for patients undergoing hip arthroscopy. PURPOSE: To investigate the relationship between neighborhood-level socioeconomic status and functional outcomes after hip arthroscopy. STUDY DESIGN: Cohort study; Level of evidence, 3. METHODS: A retrospective analysis of prospectively collected data was performed on patients aged ≥18 years with minimum 1-year follow-up who underwent hip arthroscopy for the treatment of symptomatic labral tears. The study population was divided into ADILow and ADIHigh cohorts according to ADI score: a validated measurement of neighborhood-level socioeconomic status standardized to yield a score between 1 and 100. Patient-reported outcome measures (PROMs) included the modified Harris Hip Score, Nonarthritic Hip Score, Hip Outcome Score-Activities of Daily Living, Hip Outcome Score-Sports-Specific Subscale, 33-item International Hip Outcome Tool, visual analog scale for pain, and patient satisfaction. RESULTS: A total of 228 patients met inclusion criteria and were included in the final analysis. After patients were stratified by ADI score (mean ± SD), the ADILow cohort (n = 113; 5.8 ± 3.0; range, 1-12) and ADIHigh cohort (n = 115; 28.0 ± 14.5; range, 13-97) had no differences in baseline patient demographics. The ADIHigh cohort had significantly worse preoperative baseline scores for all 5 PROMs; however, these differences were not present by 1-year follow-up. Furthermore, the 2 cohorts achieved similar rates of the minimal clinically important difference for all 5 PROMs and the Patient Acceptable Symptom State for 4 PROMs. When controlling for patient demographics, patients with higher ADI scores had greater odds of achieving the minimal clinically important difference for all PROMs except the 33-item International Hip Outcome Tool. CONCLUSION: Although hip arthroscopy patients experiencing a greater neighborhood-level socioeconomic disadvantage exhibited significantly lower preoperative baseline PROM scores, this disparity resolved at 1-year follow-up. In fact, when adjusting for patient characteristics including ADI score, more disadvantaged patients achieved greater odds of achieving the minimal clinically important difference. The present study is merely a first step toward understanding health inequities among patients seeking orthopaedic care. Further development of clinical guidelines and health policy research is necessary to advance care for patients from disadvantaged communities.

Chaotropic Agent-Assisted Supported Lipid Bilayer Formation.

Supported lipid bilayers (SLBs) are useful structures for mimicking cellular membranes, and they can be integrated with a variety of sensors. Although there are a variety of methods for forming SLBs, many of these methods come with limitations in terms of the lipid compositions that can be employed and the substrates upon which the SLBs can be deposited. Here we demonstrate the use of an all-aqueous chaotropic agent exchange process that can be used to form SLBs on two different substrate materials: SiO2, which is compatible with traditional SLB formation by vesicle fusion, and Al2O3, which is not compatible with vesicle fusion. When examined with a quartz crystal microbalance with dissipation monitoring, the SLBs generated by chaotropic agent exchange (CASLBs) have similar frequency and dissipation shifts to SLBs formed by the vesicle fusion technique. The CASLBs block nonspecific protein adsorption on the substrate and can be used to sense protein-lipid interactions. Fluorescence microscopy was used to examine the CASLBs, and we observed long-range lateral diffusion of fluorescent probes, which confirmed that the CASLBs were composed of a continuous, planar lipid bilayer. Our CASLB method provides another option for forming planar lipid bilayers on a variety of surfaces, including those that are not amenable to the widely used vesicle fusion method.

Lessons learned on the feasibility of coastal wetland restoration for blue carbon and co-benefits in Australia.

Diverse types of saline coastal wetlands contribute significantly to global biodiversity, carbon stocks, and ecosystem functions. Opportunities to incentivise coastal wetland restoration from carbon markets is growing across the world. However, little is known of the economic feasibility of blue carbon restoration across different regions, or the quantities of ecological and social co-benefits that accompany restoration. We explored the opportunities for tidal restoration of coastal wetlands for blue carbon projects in three regions across Australia. We identified biophysically suitable potential restoration sites for mangroves, saltmarshes and supratidal forests, estimated their carbon abatement over 25 years, and undertook a cost-benefit analysis under the carbon market. Potential co-benefits of restoration sites for biodiversity, fisheries, water quality and coastal protection were measured to identify economically feasible sites that maximise the provision of co-benefits. Cultural benefits were identified as the potential for leadership and collaboration by Traditional Custodians at sites. We found that the extent of restoration opportunities varied among regions, with variation in tidal range, extent of agricultural land-use, and the type of hydrological modifications influencing carbon abatement forecasts. The presence of threatened species in hydrologically modified wetlands reduced the amount of land available for restoration, however the restoration of remaining areas could produce rich ecological and cultural benefits. A high carbon price was needed to make blue carbon restoration profitable on land used for beef production. We found sites where carbon credits can be bundled with co-benefits to possibly attain higher carbon prices. Traditional Custodians were interested in leading blue carbon projects, however the opportunity is dependent on Native Title rights. Through comparison of case studies, we developed a regional approach to identify coastal wetland restoration sites for blue carbon and co-benefits that can incorporate local knowledge and data availability, engage with Traditional Custodians, and adapt to the unique characteristics of regions.

An assessment of the Midwest climate adaptation network: A call for improved coordination and collaboration.

Climate adaptation and the management of climate impacts require cross-sectoral and regional coordination and collaboration, but presently there is no thorough assessment of the adaptation network in the Midwest United States to evaluate how well it achieves such collaboration. We investigated the climate adaptation network across the Midwest to inform the strategic agenda for a climate adaptation boundary organization in Minnesota - the University of Minnesota Climate Adaptation Partnership (MCAP). We identified 150 organizations and more than 500 unique connections between them. About ten organizations with more than 25 connections each link the existing Midwest climate adaptation network, but most organizations have fewer than five connections. This asymmetry can affect the flow of resources such as information, technical assistance, and financial support. It can also hinder coordination and collaboration as called for by the Intergovernmental Panel on Climate Change (IPCC et al., 2019). The Midwest adaptation network is not well-balanced with respect to the adaptation cycle: many organizations focus on understanding or planning for climate change, with few organizations focused on problem identification, plan implementation, or monitoring. The gaps identified here suggest that MCAP and other regional adaptation organizations can (1) improve cross-sectoral and intraregional coordination and collaboration, and (2) fill gaps in the adaptation cycle, particularly implementation and monitoring. As more communities and jurisdictions move beyond climate planning toward adaptation implementation and management, and as an increasing number of state, federal and private sector funds become available to support implementation, climate service providers such as MCAP should evaluate their services and capacities and adapt alongside the communities they support.

Inequity in cardiometabolic hospital admissions and blood screening in New Zealand Indigenous Maori with psychosis.

BACKGROUND: People with psychosis experience worse cardiometabolic health than the same-aged general population. In New Zealand, Indigenous Maori experiencing psychosis have greater risk of cardiometabolic and other physical health problems. AIMS: To identify a cohort of adults accessing secondary mental health and addiction services in New Zealand, with a previous psychosis diagnosis as of 1 January 2018, and compare odds of hospital admission outcomes, mortality and receipt of cardiometabolic blood screening between Maori and non-Maori in the following 2 years. METHOD: Crude and adjusted logistic regression models compared odds of hospital admission outcomes, mortality and receipt of cardiometabolic blood screening (lipids and haemoglobin A1c) between Maori and non-Maori, occurring between 1 January 2018 and 31 December 2019. RESULTS: A cohort (N = 21 214) of Maori (n = 7274) and non-Maori (n = 13 940) was identified. Maori had higher adjusted risk of mortality (odds ratio 1.26, 95% CI 1.03-1.54), and hospital admission with diabetes (odds ratio 1.64, 95% CI 1.43-1.87), cardiovascular disease (odds ratio 1.54, 95% CI 1.25-1.88) and any physical health condition (odds ratio 1.07, 95% CI 1.00-1.15) than non-Maori. Around a third of people did not receive recommended cardiometabolic blood screening, with no difference between Maori and non-Maori after covariate adjustment. CONCLUSIONS: Maori experiencing psychosis are more likely to die and be admitted to hospital with cardiovascular disease or diabetes than non-Maori. Because of the higher cardiometabolic risk borne by Maori, it is suggested that cardiometabolic screening shortfalls will lead to worsening physical health inequities for Maori experiencing psychosis.

Templated 3' terminal fluorescent labeling of RNA using Klenow DNA polymerase.

A long-standing challenge in the study of RNA structure-function dynamics using fluorescence-based methods has been the precise attachment of fluorophores to structured RNA molecules. Despite significant advancements in the field, existing techniques have limitations, especially for 3' end labeling of long, structured RNAs. In response to this challenge, we developed a chemo-enzymatic method that uses Klenow DNA polymerase to label RNAs. In this method:•Klenow DNA polymerase adds an amino-modified nucleotide to the 3' end of the RNA, guided by the DNA oligonucleotide template.•An NHS-ester dye is then conjugated to the amino-modified RNA, forming a covalent amide bond.•For highly structured RNAs, DNA oligonucleotides complementary to the RNA disrupt pre-existing intramolecular RNA structures. This methodological advancement enables site-specific incorporation of a single modified nucleotide at the 3' terminus of various RNA substrates, irrespective of their length or secondary structure. The user-friendly nature of the technique, with minimal modifications required for different RNA targets, makes it readily adaptable by a broad range of researchers. This approach has the potential to significantly improve the development of functionalized RNA for various applications.

Temperature dependence of leaf breakdown in streams differs between organismal groups and leaf species.

Increased temperatures are altering rates of organic matter (OM) breakdown in stream ecosystems with implications for carbon (C) cycling in the face of global change. The metabolic theory of ecology (MTE) provides a framework for predicting temperature effects on OM breakdown, but differences in the temperature dependence of breakdown driven by different organismal groups (i.e., microorganisms vs. invertebrate detritivores) and litter species remain unresolved. Over two years, we conducted 12 60-day leaf litterbag incubations in 20 headwater streams in the southern Appalachian Mountains (USA). We compared temperature dependence (as activation energy, Ea) between microbial and detritivore-mediated breakdown, and between a highly recalcitrant (Rhododendron maximum) and a relatively labile (Acer rubrum) leaf species. Detritivore-mediated breakdown had a higher Ea than microbial breakdown for both leaf species (Rhododendron: 1.48 > 0.56 eV; Acer: 0.97 > 0.29 eV), and Rhododendron breakdown had a higher Ea than Acer breakdown for both organismal groups. Similarly, the Ea of total (coarse-mesh) Rhododendron breakdown was higher than the Ea of total Acer breakdown (0.89 > 0.52 eV). These effects for total breakdown were large, implying that the number of days to 95% mass loss would decline by 40% for Rhododendron and 26% for Acer between 12°C (our mean temperature value) and 16°C (+4°C, reflecting projected increases in global surface temperature due to climate change). Despite patterns in Ea, overall breakdown rates were higher for microbes than detritivores, and for Acer than Rhododendron over most of our temperature gradient. Additionally, the Ea for a subset of the microbial breakdown data declined from 0.40 to 0.22 eV when fungal biomass was included as a model predictor, highlighting the key role of fungi in determining the temperature dependence of litter breakdown. Our results imply that, as streams warm, routing of leaf litter C to detritivore-mediated fates will increase faster than predicted by previous studies and MTE, especially for labile litter. As temperatures rise, earlier depletion of autumn-shed, labile leaf litter combined with rapid breakdown rates of recalcitrant litter could exacerbate seasonal resource limitation and alter carbon storage and transport dynamics in temperate headwater stream networks.

Real-world performance of a single-use, analyser-free, molecular point-of-care test for COVID-19 used in the emergency department: Results of a prospective trial (ED-POC).

BACKGROUND: A novel single-use, analyser-free, molecular point-of-care test for SARS-CoV-2 (Veros COVID-19 test, Sherlock Biosciences) could reduce time to results and improve patient care and flow in the emergency department (ED), but its performance in this setting is unknown. METHODS: Adults aged ≥18 years presenting to Southampton General Hospital (UK) with suspected COVID-19 were tested with the Veros COVID-19 test in addition to standard of care near-patient PCR. Measures of diagnostic accuracy were calculated for the Veros COVID-19 test stratified by Ct value. Discrepant results underwent viral culture. FINDINGS: Between Jan 16 and May 2, 2023, 400 patients were enrolled with a median (IQR) age of 60 (34-77) and 141 (35·3%) were SARS-CoV-2 positive by PCR. The Veros test gave valid results on the first test in 384 (96·0%), and sensitivity and specificity were 127/141 (90·1%, 95%CI 83·9-94·5) and 258/259 (99·6%, 95%CI 97·9-100) overall. For those with high or moderate viral load (Ct ≤30), sensitivity was 125/129 (96·9%, 95%CI 92·3-99·2). One (7·1%) of 14 PCR positive/Veros test negative samples was culture positive. Median (IQR) time from sample collection to result was 19 (18-20) mins with the Veros test versus 73 (59-92) mins with PCR (p < 0·0001). INTERPRETATION: The Veros COVID-19 test generated results in near real-time, around 1 h sooner than rapid, near-patient, analyser-based PCR, and accuracy was excellent for samples with moderate and high viral loads. The Veros test represents a step-change in molecular diagnostics for infection and could significantly reduce time to results and improve patient management in EDs and other settings.

Cryptic species complex shows population-dependent, rather than lineage-dependent tolerance to a neonicotinoid.

Cryptic species are rarely considered in ecotoxicology, resulting in misleading outcomes when using a single morphospecies that encompasses multiple cryptic species. This oversight contributes to the lack of reproducibility in ecotoxicological experiments and promotes unreliable extrapolations. The important question of ecological differentiation and the sensitivity of cryptic species is rarely tackled, leaving a substantial knowledge gap regarding the vulnerability of individual cryptic species within species complexes. In times of agricultural intensification and the frequent use of pesticides, there is an urgent need for a better understanding of the vulnerability of species complexes and possible differences in adaptive processes. We used the cryptic species complex of the aquatic amphipod Gammarus roeselii, which comprises at least 13 genetic mtDNA lineages and spans from small-scale endemic lineages in Greece to a large-scale widely distributed lineage in central Europe. We exposed eleven populations belonging to four lineages to the neonicotinoid thiacloprid in an acute toxicity assay. We recorded various environmental variables in each habitat to assess the potential pre-exposure of the populations to contaminants. Our results showed that the populations differed up to 4-fold in their tolerances. The lineage identity had a rather minor influence, suggesting that the cryptic species complex G. roeselii does not differ significantly in tolerance to the neonicotinoid thiacloprid. However, the observed population differentiation implies that recent pre-exposure to thiacloprid (or similar substances) or general habitat contamination has triggered adaptive processes. Though, the extent to which these mechanisms are equally triggered in all lineages needs to be addressed in the future. Our study provides two key findings: Firstly, it shows that observed phylogenetic differences within the G. roeselii species complex did not reveal differences in thiacloprid tolerance. Second, it confirms that differentiation occurs at the population level, highlighting that susceptibility to toxicants is population-dependent. The population-specific differences were within the range of accepted intraspecific variability from a regulatory standpoint. From an evolutionary-ecological perspective, it remains intriguing to observe how persistent stresses will continue to influence tolerance and whether different populations are on distinct pathways of adaptation. Given that the potential selection process has only lasted a relatively short number of generations, it is crucial to monitor these populations in the future, as even brief exposure periods significantly impact evolutionary responses.