Here, we present a decadally resolved record of sea-salt sodium (a proxy for open-ocean area) and non-sea salt calcium  from the well-dated Roosevelt Island Climate Evolution (RICE) core, focusing on the time period between 40–26 ka BP. The RICE dust record exhibits an abrupt shift towards a higher mean dust concentration at 32 ka BP. Investigating existing ice-core records, we find this shift is a prominent feature across Antarctica. We propose that this shift is linked to an equatorward displacement of Southern Hemisphere westerly winds. Subsequent to the wind shift, data suggest a weakening of Southern Ocean upwelling and a decline of atmospheric CO Antarctic ice core records provide insights into past environmental conditions. Here, an abrupt, synchronous increase in dust from ice cores in Antarctica is identified that suggests a sudden equatorward shift of westerly winds and coincides with a reduction in atmospheric CO2. The evolution of these coupled climate events is primarily controlled by oceanic teleconnections driven by the variations in the strength of overturning circulation in the North Atlantic4, with far-reaching consequences such as changes in the Southern Ocean (SO) sea-ice conditions5. While the DO events were abrupt, the evolution of AIM events was gradual1. This implies the presence of a large ocean heat reservoir with high thermal inertia that modifies the signal in the Southern Hemisphere (SH)3. A recent study suggests that this heat reservoir could be the global interior ocean, located north of the Antarctic Circumpolar Current6.During the last ice age and especially during Marine Isotope Stage (MIS) 3 (~60–27 ka Before Present (BP)), ice cores from Greenland and Antarctica document the presence of millennial-scale warm events known as Dansgaard—Oeschger (DO) events and Antarctic Isotope Maxima (AIM) respectively8. The latitudinal positional of SHWW plays an important role in the rate of Southern Ocean (SO) upwelling10 and the amount of atmospheric dust entrainment from mid-latitude sources11. Consequently, changes in the SHWW during AIM-DO cycles are thought to have impacted atmospheric CO2 concentrations12, SO productivity9, and continental dust loading in Antarctica14. In addition, local insolation is also an important driver for climatic changes during MIS 3, particularly with its influence on SO conditions15. Sea-ice in some sectors of the SO started to advance by ~35 ka and expanded as far north as ~55°S by ~32 ka16, markedly before the Global Last Glacial Maximum  or the local Antarctic LGM (29–19 ka BP)17.However, the events also involved rapid and large-scale atmospheric re-organizations. In the SH, these changes are documented to cause a latitudinal displacement of Southern Hemisphere Westerly Winds (SHWW)2+- a proxy for continental dust) and sea-salt sodium (ssNa+ –a proxy for sea-ice/open-ocean changes) have been widely used to reconstruct environmental changes18. Here we present a new, well-dated, and decadally resolved record of ssNa+ and nssCa2+ from the Roosevelt Island Climate Evolution (RICE) ice core, an intermediate-depth ice core obtained from Roosevelt Island, West Antarctica . We focus on the section of the record that captures a period between ~40 and 26 ka BP, spanning from mid-MIS 3 to early MIS 2.Impurities measured in Antarctic ice cores such as non-sea salt calcium  climate variability and environmental conditions. The nssCa2+ record provides an opportunity to examine the dust transport and draw conclusions on the latitudinal position of SHWW, the ssNa+ record is used to infer the extent of large-scale sea-ice cover in the Ross Sea, and δD is used to understand the regional temperature and sea-ice conditions. Here we focus on a pan-Antarctic climate event that occurred at 32 ka BP and is marked by a significant change in RICE nssCa2+, RICE ssNa+, and RICE δD. We define the event based on RICE nssCa2+ and use ssNa+ and δD to understand the drivers.Roosevelt Island is an independent ice dome grounded below sea level, located at the northern edge of the Ross Ice Shelf near the Ross Sea polynya, an important location for sea-ice generation2+ and δD are published previously20 while ssNa+ is presented with this study.In the second step, we compare RICE with existing ice cores and SH mid-latitude terrestrial records for spatial context, marine sediments, and climate model outputs to interrogate plausible hypotheses for the causes of this event and to investigate potential consequences, particularly on SO upwelling and global carbon cycle. RICE nssCa2+ shows an abrupt transition across ~300 years from lower (~7.3 ppb) to higher mean values . During the glacial period, nssCa2+ measured in Antarctic ice cores is widely regarded as a proxy for Southern Hemisphere continental dust23. At RICE, the dominant source of Ca2+ has been shown to be marine during the Holocene24. In addition, minimal contributions from local Antarctic sources such as Marie Byrd Land is also possible25. We argue that the marine contribution would be modest during the glacial period as the ice-sheet extended until the continental shelf26 and SO sea-ice became more extensive16. Local Antarctic sources are also less likely to be prominent during the glacial period due to thick ice-cover. Hence, as we discussed in the previous study20, the dominant source of Ca2+ to Roosevelt Island during the glacial is likely to be dominated by terrestrial sources, originating from the SH mid-latitude sources, similar to cores of the Antarctic interior.At 32 ka BP, RICE nssCa2+. Type 1: AIM events 8, 7, 5.2, 5.1, and 4, are characterized by a reduction in nssCa2+ concentration  in the nssCa2+ concentration   and Methane Sulfonic Acid, a complex proxy for biological productivity27 shows a weak relationship  , ssNa+ shows a decreasing trend from AIM 8 towards MIS 2/GLGM  fifth-generation reanalysis products (ERA-5) sea-ice concentration28 for the period between 1951–2011 using Climate Reanalyzer (www.climatereaanlzyer.org) shows a statistically significant negative correlation between RICE Na+ and winter (June-August (JJA)) sea ice in the Ross Sea sector , the comparison of RICE with high-resolution records from East and West Antarctica shows that AIM events are well captured  Fig. 19. δD is1. AIM 7, 6, 5.2, 5.1, and 3 are identified as smaller climate fluctuations with a warm period duration of ~1000 years and a temperature change of ~1 °C1. Concurrent with nssCa2+ and ssNa+, the RICE δD record also shows a significant shift to more depleted values at ~32 ka BP   Fig. . A polew36. Trajectory modeling suggests that in addition to Patagonia, continental dust may be transported to West Antarctic ice core sites from Australia and New Zealand37. Isotopic signatures of dust deposited in marine sediments from the South Pacific also confirm a non-negligible contribution from Australia during the glacial period38. Dust entrainment to the South Pacific from New Zealand is identified for the Holocene and it is suggested that it may have been more extensive during the glacial period due to favorable environmental conditions39. However, provenance studies are yet to confirm and quantify contributions from Australia and New Zealand in Antarctica during the glacial. MD07-2611 sediment record   record from Lake Laguna Potrok Aike in southern South America  which suggests that the gyre was displaced northwards from at least 30 ka BP. This includes during Heinrich stadials 3 and 2, when the gyre is typically expected to shift poleward8. While a DO-stadial results in poleward displacement of SHWW, during interstadials the SHWW shifts equatorward8. To assess the climatic condition in the North Atlantic, we examine the δ18O record (proxy for temperature) from the high-resolution North Greenland Ice Core Project (NGRIP) ice core2  and Interdecadal Pacific Oscillation56 have been shown to be potent drivers of variations in Antarctic conditions. However, their impacts are asymmetric and regionally distinct, which stands in stark contrast to the remarkably concurrent dust shift at 32 ka across East and West Antarctica. Next, we investigate potential SH drivers.We also assess tropical-extratropical teleconnections as potential drivers for the 32 ka event. The Pacific South American mode58. Integrated summer insolation for 65°S, where summer is defined as days with insolation above a threshold of 275 W/m2, was at a minimum between ~32 and 28 ka, with a major reduction starting from ~39 ka  and lower compared to earlier time intervals (e.g. 0.03 GJ/m2 between 39 and 38 ka). Therefore, it is less likely that the insolation changes had any direct influence on the abrupt SHWW wind shift.Late MIS 3 involved a gradual reduction in local insolation in the SH ka Fig. 57. Howev+ show a significant change at ~32 ka BP, with δD showing a major depletion and ssNa+ exhibiting a lowered mean concentration  and external factors .The abrupt nature of the SHWW shift observed in the nssCa2 shows a significant drop at 32 ka BP and remained at low values through the LGM  and non-climatic  factors can contribute to glacial advance and retreat and a single driver alone may not be sufficient to explain the changes63. However, it is possible that the increased frequency of cold and moist SO air masses associated with an equatorward shift of SHWW may have helped with the glacier advance, as noted during ACR in an earlier study, when SHWW is understood to have shifted equatorward53.Finally, studies show evidence of maximum advance of multiple glaciers in New Zealand around ~31.5 ± 3 ka BP64. As it is known that changes in the strength and/or position of SHWW can evoke large impacts on global and regional climate, understanding the changes in the past can provide crucial insights into the response of SHWW and its consequences under future climate scenarios.Coupled Model Intercomparison Project 6 models indicate that the position and strength of SHWW varies with the level of greenhouse gas forcing applied, where medium to high forcing leads to stronger and poleward shifted westerly winds, while low forcing causes the winds to weaken and shift equatorward2 following the 32 ka BP event. Advancing our understanding of the complex relationships and feedback between these important drivers and large-scale consequences is critical to improve future climate change projections.In summary, Antarctic ice cores identify a continent-wide, abrupt shift in atmospheric dust concentration at ~32 ka BP. We provide evidence that links the change in dust loading to an equatorward shift of the SHWW. A reduction in the rate of SO upwelling is independently observed after the SHWW shift. This weakening of the overturning circulation in the SO may have contributed to the reported long-term drop in atmospheric CO59. The bedrock base of the dome lies ~214 m below sea level. The overlying ice has a surface elevation of ~550 m above sea level, with a total thickness of ~764 m at the crest. The drilling site  has favourable temperatures  and high accumulation ) to preserve high-resolution records of the climate and was chosen close to the ice divide to minimize layer distortion and advection at depth59. The upper 130 m of the core was recovered during the 2011–2012 field season, while the remaining section of the core (130–764.60 m) was retrieved during the following season.The RICE project is a 9-nation collaboration that drilled and recovered the RICE ice core from Roosevelt Island, a local ice dome in the Ross Ice Shelf, West Antarctica24, and gas synchronization for the remaining section of the core19. For the time interval between 30.6 ka BP-1971 CE, RICE CH4, and δ18Oatm profiles are matched to the West Antarctic Ice Sheet (WAIS) Divide ice core (WDC) CH4 and δ18Oatm records using an automated algorithm. For older sections of the core, CH4 and δ18Oatm profiles in RICE were instead visually matched due to the limited resolution of the RICE CH4 record19. A dynamic version of the Herron–Langway model has been used to simulate the Δage and the RICE 17 ice-age scale is derived by adding Δage to gas-ages. RICE discrete and continuous CH4 and cumulative ice-age uncertainty for ~40–26 ka BP are shown in Fig. The RICE 17 chronology is a combination of annual layer counting for the top 343 m, covering the past 2700 years2H/1H, expressed as δD) in RICE samples were measured at the National Ice Core Laboratory, GNS Science, New Zealand using a continuous-flow melting system with an off-axis integrated cavity output spectroscopy (OA-ICOS) analyzer, manufactured by Los Gatos Research. The experimental setup has been discussed elsewhere65. Combined uncertainty for δD for 2 cm depth-averaged resolution is estimated as ±0.85‰59. The δD record discussed here is sampled at 50-yr intervals, calculated from decadal means, and smoothed with a 500-yr low pass filter19. Data was calibrated using in-house standards and normalized against the V-SMOW2/SLAP2 scale.The stable water isotopic ratio (2+ and Na+ presented in this study were measured using a reagent-free Dionex Ion Chromatography System (ICS – 5000), with a 2 mm column and flowrate of 0.25 ml/min20. Samples were melted in a class-100 clean room facility at room temperature prior to analysis. Internal standards and an international standard, Atmospheric Environment Service (AES: Environment Canada), were inter-dispersed regularly within the samples for quality control. Precision is better than ~5% for both Ca2+ and Na+ based on internal standards. A total of 525 samples were measured for the period between ~40–26 ka BP.Ca2+) and sea-salt fraction of sodium (ssNa+) are calculated using the linear Eqs. ,,2+) and findchangepts function in MATLAB. The function identifies the location in a record where a significant change in the mean occurs by employing a global parametric method22. According to this method, the function chooses a point in the record and splits it into two sections, computes the empirical mean for both sections, and calculates the total residual error. The function then moves the location of the division point, with the minimum number of samples between division points set to one, until the total residual error is at a minimum22.The mean shift in the chemical and isotopic records of ice cores mentioned in this study are identified using the 2+ at 32160 years BP, RICE ssNa+ at 32,096 years BP, and RICE δD at 32,200 years BP. The ice-age uncertainty (1σ) in RICE around 32,200 years BP is ~500 years , EPICA Dome C (EDC), Talos Dome (TLD), and WDC. Continental dust records from low-accumulation East Antarctic ice cores  are presented as fluxes as accumulation in these locations is dominated by dry deposition, for which flux data more appropriately represent the atmospheric concentration13. In contrast, the accumulation in RICE and WDC are dominated by wet deposition59. For such regions, concentration values are more appropriate32. Synchronization of age-scales for these ice cores was achieved by identifying peaks of volcanic sulfur that exist in the ice-matrix and are placed on WD 2014 chronology7. Absolute ice-age uncertainty in each core is a combination of the uncertainty of the original WD 2014 chronology and the uncertainty associated with matching volcanic events and is ~40 years (1σ)7. Sample resolution may introduce additional uncertainty to the change point. To quantify this uncertainty associated with sample resolution, we performed a sensitivity study by resampling WDC nssCa2+  to the minimum resolution represented by Dome Fuji (~200 years) and computed the change point . In addition to the mean, we also observe that root mean square/variance abruptly changes in RICE nssCa2+ at 32,160 years BP (not shown). This suggests that the change we observe at 32,160 years BP is significant. We also examined the change point by extending the record to 60 ka BP and observe that the changepoint would still be detected at 32160 years BP (not shown). The same approach of extending the record and performing the analysis was also employed on WDC nssCa2+. The changepoint was detected at ~32 ka BP (not shown), suggesting that the analysis is robust.To understand the sensitivity of the changepoint analysis, we carried out an additional test on RICE nssCaSupplementary InformationSource DataSource Codes


Vernal keratoconjunctivitis (VKC) is a severe ocular allergic disease characterized by chronic inflammation of the cornea and conjunctiva that may lead to loss of visual acuity and blindness. The disease occurs primarily in children and is more common in geographical regions characterized by warm temperatures and high humidity. The clinical manifestations of VKC, when inadequately treated, may lead to severe complications and corneal damage. The prevalence of allergen sensitization, specific serum immunoglobulin E (IgE), and specific tear IgE was reported in approximately 55%–60% of patients with VKC, confirming the involvement of IgE-mediated and non−IgE-mediated mechanisms in the pathophysiology of the condition. This article explores current knowledge on the immunological pathways of VKC and the role of the monoclonal anti-IgE antibody, omalizumab, in its management. The review evaluated the effects of omalizumab beyond the direct IgE-mediated reactions and discusses its potential as a therapeutic target for VKC. Multiple retrospective analyses, case series, and case reports have reported the effectiveness of omalizumab in the management of VKC. A summary of the clinical data from these studies revealed that in children with VKC omalizumab treatment was well tolerated with improvement or resolution of ocular symptoms, reduction in steroid use, and enhancement of quality of life. Omalizumab may serve as a promising treatment option for VKC due to its ability to target both IgE-mediated and non−IgE-mediated pathophysiological pathways. Larger, controlled clinical trials are needed to support these findings. Limbal VKC is the most frequent form (54%) and more common in warmer climates; it presents with limbal swelling and the characteristic Trantas dots at the pericorneal limbus A. Tarsal,Patients with VKC often present with symptoms of intense itching, excessive tear production, foreign body sensation, sticky eyes, and conjunctival hyperemia. Intense photophobia and ocular pain have also been reported, which can impact quality of life (QoL).,,,Clinical findings of VKC include conjunctival giant papillae B, thickeComplications of VKC include keratoconus, shield ulcers, pseudogerontoxon, corneal scars and neovascularization, and microbial keratitis. In some patients, prolonged use of topical steroids may lead to iatrogenic complications, including cataracts, ocular hypertension, glaucoma, and corneal infection .10, 11, 12Although the clinical findings and complications of VKC are well characterized, there is a lack of standardized criteria for diagnosis and no uniform definition to grade disease severity. Several methods of grading disease severity and corneal involvement are found in the literature, and each clinician selects a method based on specificity, ease of use, and suitability for their clinical practice.This state-of-the-art review on the immunological mechanisms involved in the pathophysiology of VKC highlights the IgE-mediated and non−IgE-mediated pathways and the available treatment options, focusing on the ability of the anti-IgE monoclonal antibody, omalizumab, to target IgE and beyond.,The mechanisms governing the pathogenesis of VKC are still under investigation,,VKC is predominantly a type 2 helper T (Th2) cell-mediated disease,,,27Environmental airborne allergens, such as dust mites or pollen, stimulate the production of IgE, mainly via plasma cells. The allergen-specific IgE binds to high-affinity IgE receptors (FcεRI) on the surface of mast cells, basophils, and dendritic cells, causing degranulation and the release of pro-inflammatory mediators, which are responsible for the symptoms of VKC.,Additionally, cytokines such as IL-4 and IL-13 activate corneal fibroblasts to secrete chemokines, matrix metalloproteinases (MMPs), and intercellular adhesion molecule-1 (ICAM-1), which results in tissue damage in VKC.,Eosinophils release mediators that contribute to type 2 immune responses in VKC. Eosinophil cationic protein (ECP) and eosinophil major basic protein (EMBP) can disrupt the ocular epithelial barrier. Activated eosinophils secrete chemokines (eotaxin), MMP9, and collagenases that degrade the extracellular matrix, resulting in ocular surface remodeling in VKC.,,Although IgE is considered to be a key mediator in ocular inflammation, IgE sensitization (or atopy), assessed by a skin prick test or serum-specific IgE measurements, is reported only in approximately half of the patients with VKC.,,,Several types of non−IgE-mediated reactions occur in VKC, including direct activation of T cells, dendritic cells, enzymes, and MMPs, and non-specific hyper-reactivity.It has also been proposed that VKC may not be an isolated ocular disease but rather a systemic inflammatory disease based on the influx of inflammatory proteins and mediators (high-mobility group box-1 [HMGB1] and soluble receptor for advanced glycation end products [sRAGE]) from outside the eye. The concentration of these inflammatory mediators in the serum of children with VKC correlates with the levels in the lacrimal fluid and is also associated with the severity of the disease.,The main goals of VKC treatment are to interrupt and/or prevent the pro-inflammatory immune response and to stop remodeling to prevent long-term corneal damage and vision loss.Currently, there are no well-established, standardized treatment algorithms or guidelines for VKC, but some pharmacological options are available that can be individualized based on disease severity.VKC is a persistent and severe allergic corneal disease that might require initial treatment by ophthalmologists in an emergency with subsequent follow-up monitoring and maintenance treatment on a regular basis. Corneal disease experts generally agree that aggressive intervention early in the course of the disease is instrumental in avoiding complications and vision loss.,A multidisciplinary team approach is often recommended, involving an ophthalmologist or a corneal specialist; an allergist to manage specific allergen immunotherapy, if applicable; a pediatrician, considering the age group commonly impacted; and a pneumologist, dermatologist, internist, or an ear, nose, and throat (ENT) specialist to manage any concomitant dermatitis and asthma, rhinitis and any systemic immunosuppressive drugs and their associated complications.Symptomatic treatment includes avoidance of exposure to bright light, especially sunlight, using sunglasses and a cap; frequent ocular rinsing with saline; and cold compress applications.Common pharmacological therapies for VKC include anti-allergic and anti-inflammatory treatments such as topical antihistamines, mast-cell stabilizers, non-steroidal anti-inflammatory drugs (NSAIDs), and topical or systemic corticosteroids, depending on disease severity. These medications largely represent palliative therapy rather than targeting the complex immune mechanisms underlying the inflammation. Although relief of symptoms is achieved with steroids, their long-term use is associated with severe ocular and systemic complications, such as increased intraocular pressure, with a tendency toward evolution to glaucoma; cataract formation; and incidence of bacterial, fungal, and viral infections.,Alternative therapies include topical immunosuppressive drugs, such as cyclosporin A and tacrolimus. These agents are effective in the treatment of VKC symptoms but their long-term use may be associated with tolerability issues as they are frequently reported to have a “burning sensation” and could lead to patients being more prone to opportunistic infections such as papillomavirus, herpes, and molluscum contagiosum.,,Novel therapies targeting immunological pathways have been used off-label for VKC, including those targeting IgE. Omalizumab is a humanized monoclonal antibody that binds to free serum IgE and inhibits the binding of IgE to FcεRI on the surface of mast cells, basophils, and dendritic cells. This inhibits the synthesis and release of inflammatory mediators and reduces the presentation of antigens by dendritic cells. In addition, omalizumab inhibits the expression of FcεRI on mast cells and basophils, thereby limiting their reactivity to allergens.,,The effectiveness and tolerability of omalizumab are well established for approved indications such as moderate to severe allergic asthma, nasal polyposis, and chronic spontaneous urticaria , resulting in >1.86 million patient-years of estimated exposure.VKC is an allergic disease, and omalizumab has proven efficacy in targeting the allergic type 2 inflammatory pathways.,+, CD4+, and CD8+) cells, B cells, and IL-4-positive cells.In vitro, omalizumab decreased the deposition of type 1 collagen and fibronectin, which are implicated in the pathophysiology of VKC.Pathophysiological mechanisms of inflammation in VKC that are not directly mediated by IgE also appear to be targeted by omalizumab, as has been reported in other atopic diseases .32,34 FoIn patients with severe asthma, serum MMP9 levels decreased significantly at 12, 24, and 36 months after initiating omalizumab therapy.The anti-IgE treatment restored the capacity of plasmacytoid dendritic cells (pDCs), impaired by cross-linking of IgE–FcεRI, to induce regulatory T cells (Tregs) in vitro in atopic patients.,Biologics other than omalizumab  that target immune mechanisms have not been studied extensively in VKC.A comprehensive literature search was performed using PubMed and EMBASE databases for articles that reported the use of omalizumab in VKC, published from inception to June 27, 2022 . The medical subject headings and search terms used were: “omalizumab”, “Xolair”, “anti-IgE”, “vernal keratoconjunctivitis”, “VKC”, and “allergic conjunctival diseases”. The studies were filtered by clinical trial, publication type, and language (English). Any other potential studies were identified by manually searching based on the eligibility criteria. The search found a total of 18 articles on the use of omalizumab in 46 patients with VKC, and this included 4 retrospective analyses, 2 case series, and 12 case reports .Table 2RThe clinical data from the results of the literature search showed that, in patients with severe VKC unresponsive to standard therapies, omalizumab treatment was well tolerated and improved or resolved ocular symptoms; this was followed by complete or partial remission, gradual topical steroid dose reduction and termination, improvement of any atopic comorbidities in most cases, and enhancement of QoL in children . It is n,VKC is associated with several atopic comorbid conditions, including asthma, rhinitis, allergic rhinoconjunctivitis, AD, eczema, and food allergy. In patients with VKC, around 85% of patients had allergic comorbidity.,,,,,,In most of the studies reviewed, the outcomes of the use of omalizumab in patients with VKC were evaluated using several methods; these included the ocular visual analog scale (VAS),Multiple small studies have established the important role of IgE in VKC and the beneficial effect of omalizumab in VKC, prompting larger studies and further exploration of the underlying disease mechanisms in VKC and the role of anti-IgE therapy, mainly in patients without high systemic IgE. Since locally high levels of IgE have been reported in tears, detection of tear IgE levels may help guide treatment decisions in patients with a negative skin prick test and low serum-specific IgE levels. The absence of allergic sensitization as a risk factor for treatment failure with omalizumab also warrants further research.Large, prospective, randomized, and controlled trials to confirm the efficacy of omalizumab in VKC patients are warranted, especially to help determine the optimal dose and duration to achieve maximal benefit. Other research areas in VKC where there are data gaps with omalizumab include long-term effectiveness and safety in children <6 years old, health-economic analyses, and evaluations of possible relapse after discontinuation.Given the unmet therapeutic need to manage moderate and severe VKC in addition to the complications associated with this condition, omalizumab appears to benefit pediatric patients and is a promising treatment option based on its ability to target IgE-mediated and non–IgE-mediated pathways. Larger, controlled clinical trials are needed to support these positive findings from small studies of omalizumab in the treatment of VKC.AD, atopic dermatitis; CCR7, CC motif chemokine receptor 7; CD, cluster of differentiation; CXCR4, CXC motif chemokine receptor 4; ECP, eosinophil cationic protein; EMBP, eosinophil major basic protein; ENT, ear, nose, and throat; GM-CSF, granulocyte-macrophage colony-stimulating factor; HMGB1, high-mobility group box-1; ICAM-1, intercellular adhesion molecule-1; IFN-γ, interferon gamma; IgE, immunoglobulin E; IL, interleukin; MCP-1, monocyte chemoattractant protein-1; MMP, matrix metalloproteinase; NSAID, non-steroidal anti-inflammatory drug; OX40, synonym of CD134 ; pDC, plasmacytoid dendritic cell; QoL, quality of life; sRAGE, soluble receptor for advanced glycation end products; TARC, thymus- and activation-regulated chemokine; Th1 cell, type 1 helper T cell; Th2 cell, type 2 helper T cell; TLR, toll-like receptor; TNF-α, tumor necrosis factor-alpha; TNF-ꞵ, tumor necrosis factor-beta; Tregs, regulatory T cells; TSLP, thymic stromal lymphopoietin; US, United States; VAS, visual analog scale;VKC, vernal keratoconjunctivitis.The authors thank Preethi B and Ian Wright  for providing medical writing support in accordance with the Good Publication Practice 2022 (GPP2022) guidelines (https://www.ismpp.org/gpp-2022). The study and the writing support was funded by Novartis Pharma AG, Basel, Switzerland).Not applicable as this is a review article and not an original research.All authors have contributed equally to the design, review, and preparation of the manuscript.Not applicable, as this is a review article and not an original research.I confirm that all authors listed on the title page have agreed to the submission and consequent publication of the manuscript, post-review by the editorial board.Serge Doan reports the receipt of consulting fees from 10.13039/100007816Alcon, Bausch & Lomb, Horus, 10.13039/100004339Sanofi, 10.13039/501100004286Santen, and Thea.Nikolaos G. Papadopoulos reports the receipt of grants from Capricare, Nestle, Numil, and Vianex; and consulting fees from Abbott, 10.13039/100006483AbbVie, 10.13039/100004325AstraZeneca, 10.13039/100004330GlaxoSmithKline, HAL, Medscape, Menarini/Faes Farma, 10.13039/100016259Mylan, 10.13039/100004336Novartis, Nutricia, OM Pharma, and 10.13039/100009857Regeneron/10.13039/100004339Sanofi.Jason K. Lee reports the receipt of grants from 10.13039/100004339Sanofi, 10.13039/100009857Regeneron, 10.13039/100006483AbbVie, Astra Zeneca, and 10.13039/100004330GlaxoSmithKline. He has received consulting fees, speaker payments, and travel and meeting support from Astra Zeneca, 10.13039/100009857Regeneron, 10.13039/100004330GlaxoSmithKline, Pfizer, 10.13039/501100014565Bausch Health, Medexus, 10.13039/100004339Sanofi, 10.13039/100006483AbbVie, 10.13039/100004336Novartis, ALK, Valeo, Miravo, and 10.13039/100004331Johnson and Johnson. He has served on advisory boards for Sanofi, GlaxoSmithKline, Novartis, Regeneron, and AstraZeneca.Susanne Lau reports the receipt of consulting fees and payments for lectures from Sanofi-Aventis and has participated in advisory board meetings for 10.13039/100009946Allergopharma, 10.13039/501100011831DBV, 10.13039/100004339Sanofi, and Leo Pharma.Vibha Sharma reports the receipt of grants for conducting research, payments for expert testimony, honoraria, speaker fees, travel support, and medical writing payments from Aimmune Therapeutics. She has also received payment from Santen for expert testimony. She is a member of the Paediatric Steering Group within the British Society of Allergy and Clinical Immunology, and the European Academy of Allergy and Clinical Immunology, Ocular Allergy Working Group.Uwe Pleyer reports the receipt of grants from the 10.13039/501100000780European Union. He has received honoraria for lectures from AbbVie, Allergan, Alimera, Bayer, Novartis, Pfizer, Roche, Santen, and Thea. He is a board member of Deutsche UVEITIS Arbeitsgemeinschaft DUAG e. V and Head of the board of Uveitis/DOG section.Carmen Rondon, Salvatore Leonardi, and Sara Manti have no conflicts of interest to disclose.Xavier Jaumont and Slawomir B. Lazarewicz are employees of Novartis.


Takifugu flavidus protein hydrolysate (<1 kDa). Twenty-four peptides with an average local confidence score (ALC) ≥ 80% from bounded components (eluted by 1 M NaCl) were identified by LC-MS/MS. Among them, a novel peptide, TLRFALHGME, with ACE-inhibitory activity (IC50 = 93.5 µmol·L−1) was selected. Molecular docking revealed that TLRFALHGME may interact with the active site of ACE through H-bond, hydrophobic, and electrostatic interactions. The total binding energy (ΔGbinding) of TLRFALHGME was estimated to be −82.7382 kJ·mol−1 by MD simulations, indicating the favorable binding of peptides with ACE. Furthermore, the binding affinity of TLRFALHGME to ACE was determined by surface plasmon resonance (SPR) with a Kd of 80.9 µmol, indicating that there was a direct molecular interaction between them. TLRFALHGME has great potential for the treatment of hypertension.An affinity chromatography filler of CNBr-activated Sepharose 4B-immobilized ACE was used to purify ACE-inhibitory peptides from  The angiotensin-I-converting enzyme (ACE), a zinc-dependent dipeptidyl carboxypeptidase, plays a vital role in regulating blood pressure . ACE catThe preparation of ACE-inhibitory peptides from protein hydrolysates has always been an intense focus. ACE-inhibitory peptides can be separated using multiple techniques based on their physicochemical properties, such as molecular weight, charge difference, binding affinity, and polar nature . The con50 50 and 150 times lower than those of the original sunflower and rapeseed hydrolysates, respectively ), and the Km and Vmax were calculated.The inhibitory kinetics of TLRFALHGME were analyzed using Lineweaver–Burk plots according to the protocol described by Lin et al. . The reaDiscovery Studio 2019  was employed to analyze the binding affinities and modes of interaction between ACE and peptide . The molhttps://www.rcsb.org/structure/1O8A (accessed on 15 August 2023)). Molecular and protein files were converted into PDBQT format, where water molecules were excluded, and polar hydrogen atoms were added for docking analysis. Molecular docking results were selected based on their docking scores and binding energies. According to docking scores and binding energies, the best docking result was selected.The 3D coordinates of the structure of human ACE (PDB: 108A) were downloaded from the PDB (https://cloud.yinfotek.com/ (accessed on 20 August 2023)) [MD simulations were conducted on the Yinfo Cloud Computing Platform (CCP) using AmberTools 20 (t 2023)) . The ACEt 2023)) . The bint 2023)) . The fol−1 of TLRFALHGME were injected sequentially into the chamber in PBS running buffer. The SPR assay was performed at 25 °C on an OpenSPRTM . The conditions for the analysis were set at a flow rate of 20 μL·min−1, the binding time was 240 s, and the disassociation time was 480 s. Chips were regenerated using a regeneration buffer (0.25% SDS). A one-to-one diffusion-corrected model was fitted to the wavelength shifts corresponding to the series of TLRFALHGME concentrations. The kinetic constant analysis was performed using TraceDrawer 1.9.1 software  to determine the association constant (Ka) and dissociation constant (Kd). The affinity constant (KD) was calculated as the ratio kd/ka.The affinity interaction between TLRFALHGME and ACE was verified by SPR experiments . In brieMale SHRs  were purchased from Vital River Laboratory Animal Technology Co., Ltd. . Following an acclimation period of one week, SHRs with systolic blood pressure (SBP) higher than 180 mmHg were randomly divided into groups of ten rats. Animals were housed at 25 °C under a light–dark cycle of 12 h/12 h. TLRFALHGME was injected via tail vein at a dose of 4 mg/kg body weight.−1) as the positive control. The SBP of all rats was measured using a BP-98A blood pressure monitor  before and after intravenous administration. The rats were all treated humanely according to the guidelines of the National Institutes of Health and Use of Laboratory Animals and approved by the Ethics Committee of Guangdong Medical Laboratory Animal Center .Under the same conditions, a saline group was used as the negative control and captopril , and mean comparisons were carried out by Duncan’s multiple range test using SPSS statistics software v.20.0 .T. flavidus. In total, 24 peptide sequences of eluted fractions were identified by LC-MS/MS and subjected to ACE-inhibitory activity assay. One potential ACE-inhibitory peptide (TLRFALHGME) possessed the highest ACE-inhibitory activities with IC50 values of 93.5 µmol·mL−1. Kinetic studies indicated that TLRFALHGME possessed a competitive inhibition mode. The results of molecular docking, molecular dynamics, and SPR analysis confirmed the direct interaction between the peptide and ACE. This is also in contrast to our previous work, in which we purified a novel ACE-inhibitory peptide PPLLFAAL using semi-preparative RP-HPLC, sephadex G-15 gel chromatography, and RP-HPLC [In this study, an affinity chromatography of Sepharose-immobilized ACE was applied for the purification of ACE-inhibitory peptides from  RP-HPLC . The aff


Intraoperative pedicle screw depth adjustment after initial insertion, including both forward and backward adjustments, is sometimes necessary to facilitate rod application and ensure that the screw is in the correct position, which is determined by intraoperative fluoroscopy. Adjusting the screw with forward turns has no negative influence on the screw fixation stability; however, screw turnback may weaken the fixation stability. The aim of this study is to evaluate the biomechanical properties of screw turnback and demonstrate the reduction in the fixation stability after the screw is turned 360° from its full insertion position. Commercially available synthetic closed-cell polyurethane foams with three different densities simulating various degrees of bone density were utilized as substitutes for human bone. Two different screw shapes  together with two different pilot hole profiles  were tested. Following specimen preparation, screw pullout tests were conducted using a material test machine. The mean maximal pullout strength between full insertion and 360-degree turnback from full insertion in each setting was statistically analyzed. The mean maximal pullout strength after 360-degree turnback from full insertion was generally lower than that at full insertion. The reduced mean maximal pullout strength after turnback increased with decreasing bone density. Conical screws had significantly lower pullout strength after 360-degree turnback than cylindrical screws. The mean maximal pullout strength was reduced by up to approximately 27% after 360-degree turnback when using a conical screw in a low bone density specimen. Additionally, specimens treated with a conical pilot hole presented a less reduction in pullout strength after screw turnback as compared to those with a cylindrical pilot hole. The strength of our study was that we systematically investigated the effects of various bone densities and screw shapes on screw stability after turnback, which has rarely been reported in the literature. Our study suggests that pedicle screw turnback after full insertion should be reduced in spinal surgeries, particularly procedures that use conical screws in osteoporotic bone. Pedicle screw secured with a conical pilot hole might be beneficial for screw adjustment. The pedicle screw-rod system is commonly used in various spine surgeries to stabilize vertebrae, correct spine alignment, and accomplish arthrodesis . DespiteScrew depth adjustment, which involves turning the screw forward or backward through the track after the initial insertion, is sometimes necessary during surgery to facilitate subsequent rod application. In some cases, the screw is inserted too far, exceeding the anterior cortex according to intraoperative fluoroscopy results, and the screw must be turned backward for safety. Screw depth adjustment is usually needed in procedures that require long instrumentation, such as scoliosis surgery, because the rod is difficult to connect to the screws. In this situation, we prioritize inserting screws that could be later adjusted to deeper locations since this method does not have a negative influence on the screw fixation stability. However, there is a limit to how far the screw can be inserted forward, as the tulip of the screw will be blocked by the bony edge of the facet joint. At this point, the only option is to choose another screw and turn it back . TherefoAccording to the literature review, a study  reportedin vitro biomechanical research to investigate the difference in the pullout strengths of pedicle screws between full insertion and 360-degree turnback from full insertion using three commercially available test blocks to mimic different degrees of bone quality and two pedicle screws with distinct shapes .Because few studies have systematically explored these effects, we conduct 3, which simulated cadaveric bones with osteoporosis, healthy bones, and high bone quality, respectively, were chosen  that mimic various bone density grades were utilized as experimental substitutes for human bone. . Three te chosen . The foae chosen .This study employed two types of screws, namely, cylindrical and conical screws. The cylindrical screws had a consistent diameter of 6.0 mm throughout, while the conical screws’ diameter gradually decreased from 6.0 mm at the hub to 5.0 mm at the tip. Both screw designs had a thread depth of 1.0 mm and a thread pitch of 1.5 mm. Moreover, the thread coverage length was standardized at 40 mm, and The experiment was divided into two major parts: full screw insertion and 360-degree turnback from full insertion. First, the pilot hole was created, and the screw was directly inserted into the test block until all the threads were in the block, which was assigned as the full insertion group. The second experimental group was similar to the first group, except the screw was reversed 360° after full insertion. Three test block densities and two screw designs were used in the two groups, and the tests were repeated six times for each setting. The experimental flowchart is shown in In the two experimental groups, a consistent insertion depth and a trajectory axis that was perpendicular to the insertion plane of the test block were confirmed using X-ray imaging  before pullout testing . The prop < 0.05.To assess the impact of screw turnback, the ultimate pullout strengths of the pedicle screws between full insertion and 360-degree turnback from full insertion in each setting were statistically compared. All measurements were expressed as the mean ± standard deviation (SD). Statistical analyses were conducted using the SPSS software . Differences between groups were assessed using Mann-Whitney U tests, with a significance level of In the 3.6-mm cylindrical pilot hole group, which simulated the minimally invasive fluoroscopy-guided insertion method, the cylindrical screw at full insertion, cylindrical screw after 360-degree turnback, conical screw at full insertion, and conical screw after 360-degree turnback in the 7.5-pcf group showed mean maximal pullout strengths of 525.19 ± 21.02 N, 458.73 ± 60.02 N, 548.71 ± 67.73 N, and 433.48 ± 79.97 N, respectively, compared with mean maximal pullout strengths of 1396.05 ± 38.15 N, 1325.71 ± 117.22 N, 1652.44 ± 140.93 N, and 1464.42 ± 88.86 N, respectively, in the 15-pcf group and 4822.96 ± 105.87 N, 4717.91 ± 121.77 N, 5936.98 ± 150.43 N, and 5550.35 ± 136.66 N in the 30-pcf group .In the 3.2- to 5.0-mm conical pilot hole group, which simulated the traditional freehand insertion technique, the mean maximal pullout strengths of the cylindrical screw at full insertion, cylindrical screw after 360-degree turnback, conical screw at full insertion, and conical screw after 360-degree turnback in the 7.5-pcf group were 493.91 ± 34.30 N, 506.55 ± 54.87 N, 563.52 ± 56.67 N, and 487.25 ± 58.42 N, respectively, compared with mean maximal pullout strengths of 1417.33 ± 70.11 N, 1429.86 ± 74.78 N, 1619.65 ± 70.13 N, and 1505.98 ± 50.87 N, respectively, in the 15-pcf group, and 5513.35 ± 90.90 N, 5469.19 ± 162.18 N, 5760.98 ± 143.08 N, and 5640.11 ± 173.84 N in the 30-pcf group .These data have been organized in p = 0.023), 13% (p = 0.015), and 7% (p < 0.01), respectively, after conical screw turnback. In contrast, the mean maximal pullout strengths were reduced by 13% (p = 0.028), 5% (p = 0.192), and 2% (p = 0.342), respectively, after cylindrical screw turnback. In the 3.2- to 5-mm conical pilot hole group, the mean maximal pullout strengths in the 7.5-pcf, 15-pcf, and 30-pcf groups were reduced by 16% (p = 0.023), 8% (p = 0.015), and 2% (p = 0.213), respectively, after conical screw turnback, while these values were reduced by −2% (p = 0.642), −1% (p = 0.771), and 2% (p = 0.873), respectively, after cylindrical screw turnback. Here, the minus sign indicates an increased value.In the 3.6-mm cylindrical pilot hole group, the mean maximal pullout strengths in the 7.5-pcf, 15-pcf, and 30-pcf groups were reduced by 27%  and 13% (p = 0.028), respectively, in the 7.5-pcf group. In contrast, the mean maximal pullout strengths were reduced by 13% (p = 0.015) and 5% (p = 0.192), respectively, in the 15-pcf group and 7% (p < 0.01) and 2% (p = 0.342), respectively, in the 30-pcf group. In the 3.2- to 5.0-mm conical pilot hole group, the mean maximal pullout strengths after conical screw turnback and cylindrical screw turnback were reduced by 16% (p = 0.023) and −2% (p = 0.642), respectively, in the 7.5-pcf group, compared with 8% (p = 0.015) and −1% (p = 0.771) reductions, respectively, in the 15-pcf group and 2% (p = 0.213) and 2% (p = 0.873) reductions in the 30-pcf group. Here, the minus sign indicates an increased value.After 360-degree turnback from full insertion, in the 3.6-mm cylindrical pilot hole group, the mean maximal pullout strengths after conical screw turnback and cylindrical screw turnback were reduced by 27% . The strength of our study was that we systematically investigated the effects of various bone densities and screw shapes on screw stability after turnback, an aspect that has received little attention in the available literature. These findings provide spine surgeons with valuable insights into adjusting screw depth during spine surgery.Numerous studies have demonstrated that utilizing screw pullout testing in a laboratory setting is a dependable approach for assessing the effectiveness of a novel spinal fusion technique or instrumentation . While tp < 0.1) and 50% higher initial stiffness (p < 0.05) at full insertion. After the conical and cylindrical screws were turned back 180 or 360° from full insertion, the pullout strengths, stiffness and failure rates remained constant. Three hypothetical mechanisms were proposed, including trabecular bone elastic deformation, slight pedicle expansion, and a specific screw design that held a considerable amount of cancellous bone under compression without crushing it. They concluded that appropriately designed conical screws can be backed out 180–360° for intraoperative adjustment without loss of pullout strength, stiffness or failure. Amaritsakul et al.  cement augmentation. Ying et al.  reportedSome biomechanical studies have discussed pedicle screw reinsertion using the previous pilot hole and trajectory . These sIn this study, we found that compared with cylindrical screws, conical screws resulted in significantly decreased pullout strength after 360-degree turnback. This finding can be explained by the geometric configuration of the screw shape. The contact between the conical screws and the surrounding bone became progressively weaker during screw turnback. In the same process, the decrease in the bone contact area is not as apparent with the cylindrical screws. In term of pilot hole profile, our results also demonstrated that, for a given bone density , specimens treated with a conical pilot hole (3.2- to 5.0-mm) presented a less reduction in pullout strength after screw turnback as compared to those with a cylindrical pilot hole (3.6 mm). This implied that pedicle screw secured with a conical pilot hole might be beneficial for screw adjustment.3, are widely used as standard test materials to mimic human trabecular bone, as stated in ASTM F1839-01 standard specification for rigid polyurethane foam for use as a standard material for testing orthopaedic devices and instruments, published by the American Society for Testing and Materials (ASTM) in 2001 in Pennsylvania. The second limitation was that the cortical bone was not considered in the test block, which results in differences between the experimental pullout force values and clinically determined values. In fact, while the presence of cortical bone would improve the pullout strength, it may not change the comparative results. The average thickness of the pedicular cortical wall in a cadaveric study was 0.6–1.7 mm (The present study had some limitations. First, the test blocks were simply homogeneous material and thus cannot represent real vertebrae conditions. However, the test block was made of uniform polyurethane foam, which reduces the impact of the variability of cadaveric bones and provides an effective and reproducible platform for different degrees of bone mineral density. Therefore, polyurethane foams, with densities typically ranging from 0.16 to 0.64 g/cm6–1.7 mm . The scrThe mean maximal pullout strength after 360-degree turnback from full insertion was generally lower than that at full insertion. The reduction in the mean maximal pullout strength after turnback increased with decreasing bone density. Conical screws had significantly lower pullout strengths after 360-degree turnback than cylindrical screws. The mean maximal pullout strength was reduced by up to approximately 27% after 360-degree turnback when using a conical screw in a low bone density specimen. Therefore, our study suggests that pedicle screw turnback after full insertion should be reduced in spinal surgeries, particularly when conical screws are used in osteoporotic bone. Pedicle screw secured with a conical pilot hole might be beneficial for screw adjustment.


Oceanographic changes adjacent to Antarctica have global climatic and ecological impacts. However, this is the most challenging place in the world to obtain marine data due to its remoteness and inhospitable nature, especially in winter. Here, we present more than 2000 Conductivity-Temperature-Depth (CTD) profiles and associated water sample data collected with  full year-round coverage from the British Antarctic Survey Rothera Research Station at the west Antarctic Peninsula. Sampling is conducted from a small boat or a sled, depending on the sea ice conditions. When conditions allow, sampling is twice weekly in summer and weekly in winter, with profiling to nominally 500 m and with discrete water samples taken at 15 m water depth. Daily observations are made of the sea ice conditions in the area. This paper presents the first 20 years of data collection, 1997-2017. This time series represents a unique and valuable resource for investigations of the high-latitude ocean’s role in climate change, ocean/ice interactions, and marine biogeochemistry and carbon drawdown. The water column is highly biologically productive due to the input of micronutrients from sediments and glaciers, in addition to the high macronutrient concentrations from the upwelling of circumpolar deep water; this structures the marine ecosystem and plays a leading role in ocean carbon cycling4. These oceans lie adjacent to, and beneath, Antarctic ice shelves and marine terminating glaciers, which are being melted by ocean heat, with consequences for sea level rise globally7. It is thus of key importance to obtain sustained oceanographic measurements here, however this is especially challenging due to the remoteness and inhospitable nature of the environment. This is particularly the case during the austral winter, when extensive sea ice cover and shortened daylength (with up to 24-hour darkness) hamper operations. As such, year-round observations have great scientific value, given the systematic seasonal biases in nearly all existing datasets.The seas around Antarctica are globally important, being intimately connected to the rest of the Earth System. Processes on the Antarctic shelves lead to the formation of the densest waters that participate in the global ocean overturning circulation, and hence play a large role in setting planetary climateS, 68°7′29″W on the eastern side of Adelaide Island at the west Antarctic Peninsula  following the completion of the runway and the start of regular flights to Rothera from South America and the Falkland Islands in 1994. This led to the inception of the Rothera Oceanographic and Biological Time Series (RaTS) in 19978. The first twenty years of oceanographic and water sampling data from RaTS are presented here. Observations are ongoing, with the intention that this dataset will be updated on a five-yearly basis.Here we report such a year-round dataset collected at Rothera Research Station, which is located adjacent to Ryder Bay, and embayment within Marguerite Bay, at 67°34′8″C in winter. Sea ice forms in winter but is not always present or persistent, depending on variability in air temperature and wind strength and direction. Deep waters are relatively warm (≈1 °C) and some of this heat can be vented to the atmosphere, especially in winters with low sea ice cover and hence sustained periods where deep mixing and air-sea fluxes can persist . This derives from the CDW that flows along the shelf break at the southern edge of the Antarctic Circumpolar Current (ACC); this crosses the physical and dynamical barrier of the shelf break to flow onto the WAP shelf. The water reaching the area around Rothera is particularly strongly modified due to the convoluted route through narrow deep channels, with sills blocking the densest water and creating localised mixing due to the overflows12, with less sea ice formation due to relatively warm air temperatures13.Whilst these deeper waters (~150 m and below) show relatively little seasonal variability in temperature and salinity Figs. , 4, the 17. Chlorophyll a concentrations can exceed 20 mg m−3 at the peak of the spring and summer phytoplankton blooms, in contrast to midwinter values well below 0.1 mg m−3  mean that conditions are sometimes near optimal for phytoplankton growthm−3 Fig. .Fig. 5. The RaTS sampling has captured the variability throughout the annual cycle, made possible due to sampling in the polar winter and spring. In addition, the time series highlights interannual variability, which is influenced strongly by winter sea ice changes and, therefore, the meteorological forcings of that variability14. The changes in water mass characteristics from particularly low sea ice years can be seen through the following summer in a range of parameters, and at depth can persist on a decadal timescale when sea ice anomalies are more widespread14. The level to which significant decadal variability/trends exist in the data depends on multiple factors, including the nature and forcings of each variable being considered and the level of seasonal and interannual variability of that variable (subject explored in separate publications14). The addition of autonomous underwater gliders to the RaTS program has provided an increase in temporal and spatial resolution of physical and bio-optical parameters, and these data are reported separately11.The RaTS program has clearly shown the value in sustained observations and sampling in the marginal ice zone on the WAP. The time series has been supported by the US Antarctic Program Palmer Long Term Ecological Research (LTER) study20; inorganic carbon chemistry21; macronutrients and trace metals23; climate active gases26; sediment trap moorings and export29; and phytoplankton community dynamics30.The RaTS program sampling has provided a foundation for other scientific research projects to run alongside the core time-series sampling, for varying periods according to purpose. The additional expertise and equipment that these projects have brought have greatly increased the measurements achieved by the wintering scientists running the time series from Rothera. These projects have been run through BAS core funding, three individual UK Natural Environment Research Council (NERC) research fellowships, Antarctic Funding Initiative, Collaborative Gearing Scheme (CGS) and Collaborative Antarctic Science Scheme (CASS) projects (where BAS hosts visiting UK-based scientists) and a collaboration with the Netherlands Polar Programme. These include (but are not limited to) projects focused on: internal tides and coastal upwellingMeasurements are led by a wintering scientist (Marine Assistant) based at Rothera Research Station, supported by the boating team and other personnel on station. In the early part of the record, Marine Assistants would typically spend two consecutive winters (spanned by three summers) in Antarctica, though this has since reduced to one winter and two summers. Overlap of personnel in summer allows for training and handover of duties.Sampling occurs throughout the year, with typically more events in the austral summer compared with winter due to weather and sea ice conditions Fig. . Most ofThe deepest part of the bay at site 1 is approximately 4 km from Rothera Research Station , samples are transferred to the Bonner Laboratory for analysis, or processing/storage before transferring to the UK for analysis.a concentrations to 250 ml in summer and 2000 ml in winter) were gravity-filtered immediately on return to the research station through sequential 47 mm filters as follows: i) microphytoplankton (>20 μm size fraction via nylon mesh), ii) large nanophytoplankton (5 to 20 μm size fraction via membrane filters), iii) small nanophytoplankton (2 to 5 μm size fraction via membrane filters), iv) picophytoplankton (0.2 to 2 μm size fraction via membrane filters). Pigments were extracted into chloroform/methanol31 and measured by fluorometry (Turner AU-10 fluorometer) before and after addition of two drops of 0.1 N HCl under low light levels. Data density of chlorophyll a and other discrete water sample measurements are shown in Figs. Collected water samples were mixed gently by inversion, and triplicate samples . Ammonium  and fluorometry34. Sample measurements were carried out in triplicate using a Turner TD-700 fluorometer8.Water samples for the determination of macronutrient concentrations were filtered as soon as possible after collection (GF/C until 2015 and a PALL Acrodisc 32 mm Syringe Filter with 0.8/0.2 4)) were measured using standard wet chemistry approaches in Rothera Research Station33. After 2002, these macronutrient samples were frozen at −20 °C and measured in the UK using a standard nutrient autoanalyser approach35. From 2002 to 2017, the samples were measured at the National Oceanography Centre, Southampton ; from 2017 onwards, the samples were measured at the Plymouth Marine Laboratory .Before 2002, other macronutrients (nitrate (δ18O) inform on the relative prevalence of sea ice melt in the water sampled compared with freshwater from meteoric sources ; it thus has great value in determining the origin of substances delivered to the ocean via the freshwater system37. Unfiltered water samples were stored in capped and sealed glass bottles with rubber inserts and minimal head space and kept in the dark at +4 °C during transport to the UK37. The samples were measured for δ18O using the CO2 equilibration method for oxygen38 in triplicate . Before 2012, the δ18O measurements were made with a SIRA 10 mass spectrometer plus Isoprep18 device. From 2012 to 2017, the δ18O measurements were made with an Isoprime 100 mass spectrometer plus Aquaprep device.As a tracer measured in tandem with salinity, seawater oxygen isotopes (denoted by Direct (human) observations of ice type and coverage are made on a daily basis by the Marine Assistant, with up to three types of ice observed recorded at any one time. Ice coverage is given in tenths. Ice categories are given in Table 39. The data coverage of CTD casts is shown in Fig. The Data Records are provided in comma-separated values (.csv) and NetCDF (.nc) formats and are held by the UK Polar Data Centre39, with each record listing the Event number, Date (YYYY-MM-DD), the parameter (and units), and any additional comments. The data coverage of the discrete water samples, and seawater parameters measured from the samples, is shown in Figs. The Data Records are provided as comma separated variable , the site, ice observations, and any additional comments. Abbreviations used in the ice observation dataset are shown in Table The Data Records are provided as comma separated variable . Spikes were then identified and removed manually in salinity in the initial processing. This is usually between 1 and 7 metres of data, though some profiles are completely removed where pump problems make all data invalid. The precision of the salinity data is ensured by discrete samples being collected and by joint casts between the RaTS CTD(s) and that used by the Palmer LTER program, with adjustments applied in initial processing.In polar waters, with temperatures normally below 4 °C, density profiles are largely governed by the shape of the salinity profile. This means that salinity checks can also include density profile checks and the dynamical feasibility of density overturns. There are a limited number of casts with significant density overturns. As these would make the profiles statically unstable (dense water above less dense), in almost all cases they can be ascribed to sensor problems. They can happen throughout a profile but are more common at the surface or bottom of the profile. They were screened by detecting overturns of >0.05 kg mTemperature has little effect on density in the range encountered and is therefore free to vary up and down with depth such that there is no way to ascribe a profile to be physically implausible. The temperature data have been very robust, with no suspicious profiles and very tight matches in all joint casts, it is therefore presented as recorded, except for profiles with pump problems.From 2017 there have been repeating problems with the Photosynthetically Active Radiation (PAR) sensors, despite servicing and replacement. Some values at depth are easily filtered as impossible but other times the values are within bounds but the shape of the profile is implausible. There are standard sampling issues, caused by shading from the boat, ice and varying cloud coverage, such that mean light can increase rapidly with time and/or depth. This makes filtering the problem profiles harder, without removing data where the sensor is working well. Often the shape of the profile is more important than the absolute values, as it is a direct measure of attenuation, so these profiles that increase with depth are of reduced value.The first filtering is to use a mask created from the first 700 events and remove values <−1. This removes fliers, accounting for the variability driven by weather and attenuation (which can vary considerably with phytoplankton concentration). Away from changing shading/cloud conditions the expectation is for an exponential decline of PAR with depth, and so significant deviations from this can flag up potentially problematic profiles. Estimations of attenuation from PAR profiles are calculated by fitting a regression line to the logarithm of PAR in overlapping 5 m depth intervals down the profile. Checking profiles with negative “attenuation” catches further profiles that are judged to be problematic due to the sensor  and these PAR profiles are removed after individual checking. Two profiles in 2015  show very unlikely increases at depth. Given the similarities and closeness of the profiles these have also been deemed a sensor problem and blanked out.−3 density difference criterion (referenced to either the surface density or density at 10 m) gives a good match to the reduction of chlorophyll with depth, for the period of the year where the mixed layer exceeds the photic zone. This is a good indication of the mixed layer depth that is calculated describing the depth that is, or has very recently been, connected to the surface through vertical mixing.Quantification of mixed layer depth (MLD) is sensitive to the definition used, which is inevitably a processing choice, with no universally correct answer. Profile-by-profile checking has shown that the use of a 0.05 kg mA time where this has been found to be too tight a criterion is during winter or early spring where spring melt can produce a very shallow layer of fresher, less dense water, above a homogenous zone that has clearly mixed recently. This is exacerbated by our inability to take profiles in the windy conditions that drive the mixing, due to boating safety reasons. To counter this a mixed layer depth relative to 10 m is also calculated, which has use in identifying (for example) deep mixing events that can happen between sampling events.−2 40. This is considered more relevant than a profile of (for example) Brunt Väisälä frequency.There are frequent times through summer where there is strong stratification through the surface depths  due to the large input of meltwater from sea ice, icebergs and the glaciers. In these circumstances, a mixed layer depth of 1 m or 2 m does little to describe the conditions for mixing of phytoplankton. Due to this, stratification is calculated from the surface to 10 m, 20 m, 30 m through to 100 m in 10 m increments. This is calculated as the additional potential energy that is required to homogenise the depth interval, which gives a physically based metric, with units of J ma standards, with samples diluted as required during strong phytoplankton blooms to reduce the range of values measured. The ratio of fluorescence before and after acidification measured each sampling day and is used to assess the reliability of the phaeopigement data. All data are reported as chlorophyll a . Due to a quality control issue, chlorophyll values after Event 1930 (15 NOV 2017) are currently blanked; it is hoped to incorporate these in a future RaTS data release once resolved.Calibration is carried out twice a year using chlorophyll 8. The detection limit is 0.01 μM. Any negative values were reset at 0.001 μM.Ammonium measurements were carried out in triplicate using a Turner TD-700 fluorometer, and calibrated using standard addition comprising four concentrations, also in triplicateμM for nitrate, 0.1 μM for nitrite, 0.2 μM for orthophosphate, and 1.2 μM for silicic acid from 1998 to 2017 (OSIL/NOCS). From 2017 onwards, detection limits for nitrate and orthophosphate were 0.02 μM, and 0.01 μM for nitrite, and 0.02–0.03 μM for silicic acid (PML). The typical uncertainty of the analytical results was between 2 and 3%. Clean sampling and handling techniques were employed during the sampling, processing, defrosting , and manipulations within the laboratory, and where possible carried out according to the International GO-SHIP nutrient manual recommendations41. After 2017, seawater nutrient reference materials (KANSO Ltd. Japan) were analysed to assess analyser performance and for quality control purposes. Spikes were removed from the macronutrient dataset as follows: Nitrite – Events 502 and 504 (APR 2003); Nitrate – Event 328 (15 JUN 2001); Orthophosphate – Event 486 (21 FEB 2003); Silicic acid – Event 314 (18 APR 2001).For the other inorganic macronutrients, detection limits are 0.3 δ18O.Isotope measurements used internal standards calibrated against the international standards VSMOW and VSLAP (before 2012) and VSMOW2 and VSLAP2 (after 2012), with errors typically <0.05‰ for


Accurate phylogenies are fundamental to our understanding of the pattern and process of evolution. Yet, phylogenies at deep evolutionary timescales, with correspondingly long branches, have been fraught with controversy resulting from conflicting estimates from models with varying complexity and goodness of fit. Analyses of historical as well as current empirical datasets, such as alignments including Microsporidia, Nematoda, or Platyhelminthes, have demonstrated that inadequate modeling of across-site compositional heterogeneity, which is the result of biochemical constraints that lead to varying patterns of accepted amino acids along sequences, can lead to erroneous topologies that are strongly supported. Unfortunately, models that adequately account for across-site compositional heterogeneity remain computationally challenging or intractable for an increasing fraction of contemporary datasets. Here, we introduce “compositional constraint analysis,” a method to investigate the effect of site-specific constraints on amino acid composition on phylogenetic inference. We show that more constrained sites with lower diversity and less constrained sites with higher diversity exhibit ostensibly conflicting signals under models ignoring across-site compositional heterogeneity that lead to long-branch attraction artifacts and demonstrate that more complex models accounting for across-site compositional heterogeneity can ameliorate this bias. We present CAT-posterior mean site frequencies (PMSF), a pipeline for diagnosing and resolving phylogenetic bias resulting from inadequate modeling of across-site compositional heterogeneity based on the CAT model. CAT-PMSF is robust against long-branch attraction in all alignments we have examined. We suggest using CAT-PMSF when convergence of the CAT model cannot be assured. We find evidence that compositionally constrained sites are driving long-branch attraction in two metazoan datasets and recover evidence for Porifera as the sister group to all other animals.  Understanding the biological foundations of contemporary life on Earth requires detailed knowledge of evolutionary history. The history of speciation events informs us about the appearance of advantageous innovations and the loss of dispensable traits in a continuously changing environment. Consequently, the development of phylogenetic models inferring the history of speciation events has continued at an impressive pace during the past decades.Models of sequence evolution are inevitably simplifications of the complex processes that generate real-life biological sequences. Unfortunately, overly simplistic models can lead to model misspecification and long-branch attraction  symmetric exchangeabilities, describing differences in rates of exchange between pairs of states, and (ii) stationary frequencies of the target states e.g., . FiguratWe can distinguish between general distribution mixture models estimated from curated training databases, and distribution mixture models directly estimated from the datasets at hand. For example, Finite mixtures can be used in the context of maximum likelihood (ML) phylogenetic inference. However, statistical analyses of model fit and investigation of known cases of LBA indicate that a large number of components are necessary for robustness against LBA  which isBayesian approaches can more easily accommodate richer mixtures. Nonparametric Bayesian methods do not require explicit specification of the number of mixture components nor their stationary distributions. In particular, the CAT model  uses a DIn all cases, mixture modeling approaches accounting for across-site compositional heterogeneity are complex and require considerable computational resources e.g., . In ordeIn this work, we follow a multistep procedure similar to the PMSF model and address two points: First, on the computational side, we extend the PMSF approach by using the CAT model instead of an empirical profile mixture model as the reference model for computing the profiles. Importantly, the CAT model is used under a fixed guide topology. Thus, the problem of simultaneous inference of both the site-specific stationary distributions and the tree is reduced to a search of site-specific stationary distributions, and tree branch lengths only. We termed our approach CAT-PMSF.Second, we investigate the contribution of sites with different degrees of compositional constraints to LBA. In particular, we test to what extent more severely constrained sites exhibit bias toward LBA trees under models that do not adequately account for across-site compositional heterogeneity. We employ “compositional constraint analysis,” which examines phylogenetic signal for alternative topologies as a function of per-site compositional diversity measured by the effective number of amino acids .Examining simulated alignments as well as classic empirical datasets including Microsporidia , Nematodsite-homogeneous and site-heterogeneous when referring to models ignoring and accounting for across-site compositional heterogeneity, respectively. Furthermore, we use the term tree to denote a directed acyclic graph with node labels and branch lengths, in which exactly one branch connects any two nodes. We use the term topology to denote a tree without information about branch lengths but with node labels. We specify an evolutionary model with exchangeabilities, EX, and across-site compositional heterogeneity model, ASCH as EX + ASCH. All discussed evolutionary models used for simulations as well as inferences implicitly use discrete gamma rate heterogeneity with four components. We add a flag +PMSF to denote usage of the posterior mean site frequency model . The scripts and the simulated data are available in the Dryad Digital Repository: https://doi.org/10.5061/dryad.g79cnp5rh and at https://github.com/drenal/cat-pmsf-paper.We used Felsenstein-type trees with four leaves , let us denote them topology A positive value of Supplementary Figs. S13–S15 available in the Supplementary material which can be found in the Dryad Digital Repository: https://doi.org/10.5061/dryad.g79cnp5rh). For the simulation study, we used the actual http://phylopic.org/), the silhouette for Microsporidia is based on We ordered and binned the sites according to their Supplementary Figs. S1 and S2 show simplified and complete species trees, respectively.Supplementary Fig. S3 show simplified and complete species trees, respectively.The dataset provided by Supplementary Fig. S4 show simplified and complete species trees, respectively.The placement of Ctenophora on the tree of Metazoa is still a matter of debate. We apply CAT-PMSF to two datasets. First, the alignment provided by https://github.com/lsjermiin/Homo.v2.1) performs this test for all pairs of sequences in an alignment. We applied Homo v2.1 to the simulated as well as empirical alignments.CAT-PMSF assumes homogeneity of the evolutionary process across the branches of the tree. The matched-pairs test of symmetry  tests foIn brief, CAT-PMSF comprises three steps: (i) estimate a guide topology using a site-homogeneous model, (ii) estimate site-specific stationary distributions with the CAT model in PhyloBayes  using thWe assessed and compared the accuracy of CAT-PMSF with other site-homogeneous and site-heterogeneous models. To this end, we simulated amino acid sequence alignments with a length of 10,000 sites along Felsenstein-type quartet trees . Supplementary Figs. S7 and S8 show results for other values of The true topology was not recovered with site-homogeneous models when Supplementary Figs. S7 and S8). For site-homogeneous evolutionary models, more constrained sites with a lower value of At odds with this expectation, site-homogeneous evolutionary models exhibit conflicting phylogenetic signal between sites with lower and higher Supplementary Tables S3, S5, and S7). For the Pearson’s correlation coefficient, site-homogeneous models exhibit large and significant correlation for To ascertain the statistical significance of the compositional constraint analysis we calculated Pearson’s correlation coefficients, as well as Spearman’s and Kendall’s rank correlation coefficients and associated p-values between the log-likelihood differences and the site-specific Supplementary Table S9). AU tests of the Poisson + CAT-PMSF model show similar results in that the topology exhibiting LBA is rejected for Supplementary Table S10). The LG + CAT-PMSF model only rejects the topology exhibiting LBA for Supplementary Table S10).Approximately unbiased (AU) tests  of ML trSupplementary Figs. S7 and S8). In all cases excluding the LG + CAT-PMSF and GTR + CAT-PMSF models for large Supplementary Figs. S7 and S8).Finally, we note that site-heterogeneous models with GTR exchangeabilities perform well if Supplementary Figures S9 and S11 show the cumulative site-specific log-likelihood differences and Supplementary Figures S13–S15 provide results for alternative bin sizes.Similar to the simulation study above, for empirical alignments the site-specific stationary distributions obtained in Step 2 of the CAT-PMSF pipeline can be used to quantify the strength of compositional constraint measured by For site-homogeneous models, the site-specific log-likelihood differences between the ML trees constrained to the two competing topologies . For Platyhelminthes, the bias is strong enough that the total likelihood across all sites is higher for the LBA topology, while for the datasets involving Nematoda and Microsporidia, the LG + C10 + PMSF model recovers the correct topology, albeit with reduced support. In general, the results of the LG + C10 + PMSF and LG + C60 + PMSF models are consistent with the observation .The site-heterogeneous LG + C60 + PMSF model shows reduced, but still apparent conflict compared to site-homogeneous models and the LG + C10 + PMSF model (ervation  that incSupplementary Table S4). The ML trees inferred by CAT-PMSF are consistent with the accepted phylogenetic relationships and AU tests confirm the rejection of trees with LBA topologies (Supplementary Tables S12–S14).In contrast, CAT-PMSF exhibits consistent signal toward the assumed-to-be-correct topologies across all sites and datasets with no significant correlation between log-likelihood difference and site-specific Finally, we used CAT-PMSF on two metazoan datasets  to invesSupplementary Figs. S10 and S12) for both the alignments from Supplementary Table S4).Compositional constraint analysis under site-homogeneous models, as well as combinations of PMSF and site-heterogeneous mixture models with 20 and 60 components exhibit patterns of conflicting phylogenetic signal for sites with different degrees of compositional constraints . Sites with higher Supplementary Table S3) favoring a Porifera-sister topology and rejecting the Ctenophora-sister topology  with the closest outgroup, Choanoflagellatea . For the alignment published by Under site-homogeneous models, sites with We introduce CAT-PMSF, a method for phylogenetic inference from alignments exhibiting across-site compositional heterogeneity. The CAT-PMSF pipeline uses the site-specific amino acid preferences estimated by a nonparametric Bayesian approach in the context of a downstream ML analysis. Doing so combines the benefits of both approaches: a more accurate inference of the patterns across sites with a computationally more efficient and more reproducible inference of the tree topology. In addition to phylogenetic inference, the CAT-PMSF pipeline can also be used to investigate the consistency of phylogenetic signal for sites under different degrees of compositional constraints. Compositional constraint analyses on both simulated and empirical datasets exhibiting across-site compositional heterogeneity show that site-homogeneous and some site-heterogeneous mixture models indeed have inconsistent signal which contributes to topological bias and LBA artifacts.In the simulation study , site-hoThe threshold Supplementary Figs. S7 and S8).In our simulations, support of site-homogeneous models shifted from the true topology toward the incorrect topology when increasing CAT-PMSF . AlthougWe discovered bias toward one of the topologies in simulation studies because we know the true parameters and trees. Bias is harder to detect in analyses of empirical data. Compositional constraint analysis detects conflicting signal between more and less constrained sites. Detection of such inconsistencies is a strong indicator for bias: Knowing the stationary distribution of a site alone should not provide us with information about the favored evolutionary history. In mathematical terms, the log-likelihood difference of a site between two hypotheses should be conditionally independent given the stationary distribution of that site. In contrast, we expect the signal obtained from more and less constrained sites to be consistent up to the random statistical error.P-values confirm this observation across a wide range of simulated and empirical datasets (Supplementary Tables S3 and S4).In our analyses of empirical data, we observed strong inconsistencies between more and less constrained sites for site-homogeneous models and hardly any inconsistencies when using CAT-PMSF . PearsonSupplementary Table S15). If we add more distant outgroups, the results are less conclusive (Supplementary Table S15). LBA provides an explanation for this observation: outgroups more distant to the ingroup  attract outgroups closer to the ingroup .Compositional constraint analysis seeks to decompose the log-likelihood contributions of more and less constrained sites. We decided to measure how constrained sites are by computing the effective number of amino acids per site based on the concept of homoplasy, because it is inherently related to LBA: A model that underestimates the probability of homoplasy will incorrectly attribute sequence similarities due to homoplasy to a putative close evolutionary distance. Other measures of how constrained sites are or any monotonic transformation of the function computing eristics . To reiteristics –4. More We also note that CAT-PMSF assumes homogeneity of the evolutionary process across branches of the tree. The simulation study conforms to this assumption. Tests for across-branch homogeneity were less conclusive for the empirical datasets  than for the simulation study. If the evolutionary process is stationary and homogeneous, the CAT model should perform well in estimating site-specific amino acid compositions. Even if the evolutionary process is non-stationary or heterogeneous, the site-specific amino acid compositions inferred by the CAT model will capture the spectrum of compositions attained at least somewhere on the tree. In this case, the inferred compositions will not be stationary in the mathematical sense, but still should have a positive impact on the detection of genuine convergent evolution. Ideally, we should model across-site and across-branch compositional heterogeneity for amino acid sequences in a combined way. For example, there has been work in this direction on nucleotide sequences e.g., . In the Supplementary Table S1), and trees that we are convinced to be free from LBA artifacts in the analyses comprising empirical datasets . This observation justifies the usage of site-homogeneous models in Step 1 of the CAT-PMSF pipeline.The results of CAT-PMSF are conservative because the CAT model estimates the site-specific stationary distributions using guide topologies prone to LBA artifacts. That is, the guide topologies are obtained with site-homogeneous models. Even so, CAT-PMSF correctly infers the true trees in the simulation study . In practice, many sites may have evolved under different conditions, so we cannot expect all sites to share a universal stationary distribution. In fact, we do not even expect stationarity. In our opinion, we should analyze data using complex models and decide about which parameters are necessary to grasp the complexities of evolution. With CAT-PMSF we further explored this path. The CAT-PMSF method uses site-specific stationary distributions and therefore is a parameter-rich model.In comparison, the site-specific posterior mean stationary distributions of the classical PMSF approach are a superposition of a finite set of stationary distributions of the underlying mixture model. Consequently, the Finally, the speed benefit of CAT-PMSF originates from fixing the topology during the Bayesian analysis with the CAT model. Of course, estimating the site-specific stationary distributions is still by far the most time-consuming step. In the future, we aim to design improved methods for estimating site-specific stationary distributions. Specifically, we are thinking about methods based on machine learning such as AlphaFold .In conclusion, compositional constraint analyses show evidence for a potential LBA caused by model misspecification and an argument that careful model choice as well as validation is important in phylogenetic inference. We also propose a method, CAT-PMSF, with the potential to produce more accurate phylogenetic estimates using a site heterogeneous, but branch homogeneous, substitution process.https://doi.org/10.5061/dryad.g79cnp5rh.Supplementary material, including data files and/or online-only appendices, can be found in the Dryad Digital Repository:


Dry ice is widely used in the chemistry research settingsas anexcellent coolant. Herein, we report a case study of a graduate studentresearcher who lost consciousness while retrieving 180 lbs of dryice from a deep dry ice container. We share the details of the incidentand the lessons learned from it to promote safer handling of dry icein these situations. We have includeda brief overview of safety incidents with dry ice, as well as thephysical and mental health effects from such events.Dry ice is the solid form of carbon dioxide(CAS No. 124-38-9)that sublimes into gaseous carbon dioxide, which is heavier than air,at −78.5 °C at atmospheric pressure.Prior to the COVID-19 pandemic, dryice was delivered from thesupply company to individual research laboratories in the Departmentof Chemistry at UC Irvine. However, since COVID-19, the lab-specificdelivery service stopped, and the laboratories are now largely dependentupon the purchase of dry ice from the School of Physical Science Stores(PS Stores). Each dry ice pack thus supplied weighs 30–50 lb(14–23 kg) and is wrapped in brown paper for handling.On Monday, May 30th, 2022 , around 8 a.m., a graduatestudent researcher visited PS Stores with a cart to retrieve dry icefor their research group. The general protocol for the lab was that∼200 lb (90.7 kg) of dry ice would be purchased from the storeand brought up to the lab to store it in separate, enclosed storageboxes distributed among the different bays of the laboratory. Thisdry ice would be used by about a dozen people in the synthetic chemistrylab over the course of a couple of days. At the time of the incident,the graduate student was wearing full personal protection equipment,including closed-toe shoes, long pants, a lab coat, nitrile and cryogenicgloves, and safety goggles. Because of the holiday, PS Stores staffmembers were not on duty, and the graduate student opted to do a self-check-outof the dry ice.The dry ice packs in the PS Stores are heldin a container 1 with hoThe graduate student repeated this procedure severaltimes, placing∼180 lb (81.6 kg) of dry ice onto the cart. After placing thefinal block onto the cart, they became dizzy, had shortness of breath,and lost consciousness 2. The grIt is evident that, had the person lost consciousnesswhile theirtorso was extended into the container, the outcome would have beendisastrous. Luckily, in this case, the exposed graduate student hada medical check-up, including a chest X-ray and bloodwork, and allresults were normal. The student did not experience any burn injuries.After this safety incident, the exposed student was clinicallydiagnosed with post-traumatic stress disorder (PTSD). The person continuesto have strong negative reactions when encountering situations reminiscentof the incident, such as when entering areas with a cold, positiveairflow or handling dry ice for use in cold traps.The container is too deep for a researcher to retrievedry ice safely when the supplies are nearly exhausted.A buddy system was not in place for the job.There were no tools provided to expeditethe retrievalprocess.There was no oxygen or carbondioxide oxygen monitoringsystem, nor enhanced indoor ventilation system.There was no Standard Operating Procedure, or warningsign for retrieving dry ice.Because dry ice, especially in large quantities, isan asphyxiation hazard in a confined space, dry ice storage must beappropriately sized for those working with it to minimize asphyxiationrisks.A buddy system is critical forprevention of any adverseevents from exposure to carbon dioxide and for effective responsein the event of any such event.A StandardOperating Procedure must be carefully designedwith respect to the working environment and tasks. Signs and warningsmust be clearly visible.Warning signs and informational posters were postedon the walls near the dry ice container and on bathroom stalls toeducate researchers of risks associated with dry ice.A large metal clamp tool was provided to help peoplelift dry ice blocks from the bottom of the container. However, thistool is challenging to use with heavy blocks of dry ice.PS Stores implemented a process to regularly monitorthe level of dry ice; once below half full, the container is exchangedfor a full one such that the dry ice is more easily accessible. Thecontainer was also moved away from one wall, affording access to thecontainer from three usable sides.Critically,a mandatory buddy system was introducedfor when dry ice is retrieved from PS Stores and transported to researchlaboratories.After the incident,there were multiple discussions within the Graduate Student SafetyTeam, faculty advisors, and the environmental health and safety (EH&S)team about how to enable safer retrieval of dry ice for all researchers,with an obvious preference for engineering controls that would diminishor eliminate risk. The following measures were implemented as a result:We believe that these solutions, in aggregate, significantlyreduce the risk associated with retrieval of dry ice blocks from UCIPS Stores. Unfortunately, the Chemistry department, EH&S, andPS Stores staff concluded that it was not plausible to change thesize of the container that the supplier uses to provide dry ice, becausemost bins that are overall smaller in volume are either just as tallor taller than the one currently in use. Dry ice pellets were consideredas a possible alternative; however, they would be stored in similarcontainers and likely require more time within the depths of the containerfor retrieval.2 Fifteen 1 L dry ice blocks had been deliveredto the room 3 h prior to his entry. About 4 h after last seen alive,the decedent was found on the floor of the cold room. The autopsyreport concluded that the cause of death was excessive inhalationof carbon dioxide. A similar incident occurred in 2009, as a 59-year-oldfreezer technician entered a broken walk-in freezer filled with dryice.3 The autopsy report suggested thatthe decedent likely sought to exit the freezer room based on the contusionson his right arm and hands and the dents inside the freezer room door.A similar incident occurred in 2021 to a 35-year-old man who workedat a fast-food chain.4 He initially experiencedacute hypoxic respiratory failure and metabolic acidosis, but wasdischarged within 24 h subsequent to supplemental oxygen and fluidtreatment.There are many examplesof asphyxiation injuries caused by carbondioxide released by sublimation of dry ice in enclosed spaces. In2002, a 51-year-old medical research scientist with a history of mildcold-induced bronchial asthma entered an 8 ft × 8 ft × 14ft (2.4 m × 2.4 m × 4.3 m) poorly ventilated cold room.5 In 2017, a 62-year-old woman wasdriving with a container of dry ice and ice cream present with herin the vehicle.6 After 15 min of experiencinglightheadedness and shortness of breath, she was found unconsciouswith her vehicle overturned. Similar asphyxiation events caused deathsin an ice cream truck7 and a restaurantdelivery truck.8Dry ice asphyxiations in cars have also been reportednumeroustimes.9 transportationof a liquid CO2 tank that affected 25 people,10 a volcanic explosion,11 and onion fermentations.12 Most deathsoccurred in confined indoor spaces.Other examples ofasphyxiation injuries by carbon dioxide includethe injuries from a fire extinguisher system,2 is present in the arterialbloodstream, the “Bohr effect” occurs, where an elevatedlevel of CO2 causes hemoglobin to have a lower affinityfor O2.13 The increased partialpressure of CO2 in alveoli leads to reaction with waterto generate carbonic acid, which triggers respiratory acidosis. Thus,inhalation of significant quantities of CO2 lowers theblood pH and impacts cardiological, respiratory, and nervous systems.12 Common symptoms of CO2 exposure atlower concentrations are hearing loss, tachycardia, hypertension,hyperventilation, headache, and dizziness.14 At higher concentrations, convulsion, loss of consciousness, anddeath have also been reported. Langford published an excellent in-depthreview on the mechanism and toxicokinetic and clinical features ofCO2 poisoning.14Carbon dioxide gas is a normal byproduct of the human respiratorysystem. But when excess COAs noted above,the graduate student involved in the incident wasdeemed physically healthy, but was diagnosed with PTSD and has lingeringmental health problems nearly a year after the incident. While preparingthis manuscript, we noticed the lack of literature reports on themental health of researchers after experiencing safety incidents.We would like to encourage those involved in lab safety incidentsto share holistic stories of how such events can impact researchers’lives.1 source in organic reactions, dry ice must,like all chemicals, be handled carefully and with adequate personalprotection equipment and safety measures. This report was disclosedin the hope of preventing dangerous situations related to retrievingdry ice from large containers in laboratory settings. Indeed, in aninformal poll by the corresponding author of a number of chemistryfaculty members at representative research active institutions, thesame dangerous situation existed in many cases. In some instances,direct delivery of smaller quantities of dry ice to laboratories wasavailable; even so, these institutions almost surely have a centralizedstockroom or store from which dry ice can be retrieved by less regularusers, likely from containers of the sort involved in the incidentwe describe. We worry that with such a routinely used substance asdry ice, researchers might become desensitized to its potential dangers,particularly with respect to asphyxiation.While it is an excellent coolant for cryogenicbaths and servesas a valuable C