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Fuels-reduction treatments are commonly implemented in the western U.S. to reduce the risk of high-severity fire, but they may have negative short-term impacts on species associated with older forests. Therefore, we modeled the effects of a completed fuels-reduction project on fire behavior and California Spotted Owl (Strix occidentalis occidentalis) habitat and demography in the Sierra Nevada to assess the potential short- and long-term trade-offs. We combined field-collected vegetation data and LiDAR data to develop detailed maps of forest structure needed to parameterize our fire and forest-growth models. We simulated wildfires under extreme weather conditions (both with and without fuels treatments), then simulated forest growth 30 years into the future under four combinations of treatment and fire: treated with fire, untreated with fire, treated without fire, and untreated without fire. We compared spotted owl habitat and population parameters under the four scenarios using a habitat suitability index developed from canopy cover and large-tree measurements at nest sites and from previously derived statistical relationships between forest structure and fitness (λ) and equilibrium occupancy at the territory scale. Treatments had a positive effect on owl nesting habitat and demographic rates up to 30 years after simulated fire, but they had a persistently negative effect throughout the 30-year period in the absence of fire. We conclude that fuels-reduction treatments in the Sierra Nevada may provide long-term benefits to spotted owls if fire occurs under extreme weather conditions, but can have long-term negative effects on owls if fire does not occur. However, we only simulated one fire under the treated and untreated scenarios and therefore had no measures of variation and uncertainty. In addition, the net benefits of fuels treatments on spotted owl habitat and demography depends on the future probability that fire will occur under similar weather and ignition conditions, and such probabilities remain difficult to quantify. Therefore, we recommend a landscape approach that restricts timber harvest within territory core areas of use (∼125 ha in size) that contain critical owl nesting and roosting habitat and locates fuels treatments in the surrounding areas to reduce the potential for high-severity fire in territory core areas.

A cell-free culture system was designed for human embryo development to the blastocyst stage by testing a range of culture conditions in a series of protocols. The culture system that was evolved has a brief 1 h exposure to spermatozoa and then culture of the pronucleate zygote for 2 days in IVF-50 medium. Two or three embryos were cultured together in 20 μl microdrops of medium under oil. Embryos were then regrouped and two or three at a similar stage were cultured together in 50 μl microdrops of Gardner's G2 medium under oil from days 3 to 5. Embryos were transferred to fresh G2 medium on day 5 and cultured for a further 1 or 2 days (day 6 or 7). No serum was used in any of the cultures. The embryo transfer medium and G2 medium were supplemented with human serum albumin. The zonae of all blastocysts to be transferred to patients were completely removed enzymatically. Using this protocol, 52% of zygotes developed to blastocysts and 34 out of 35 patients treated received 82 blastocysts and 11 morulae on day 5 or 6. Twenty-one fetal sacs with positive heartbeats (23% implantation rate) were detected in 13 ongoing pregnancies (38% pregnancy rate/transfer or 37%/patient treated). We anticipate that further improvements in embryo development and the selection of viable embryos can be achieved using this simple and effective culture system. [ABSTRACT FROM PUBLISHER]

Background: Hypertension is a major source of morbidity and mortality worldwide, particularly for racial and ethnic minorities who face higher rates of hypertension and worse health-related outcomes. Recent research has reported on protective associations between classic psychedelics and hypertension; however, there is a need to explore how race and ethnicity may moderate such associations., Methods: We used data from the National Survey on Drug Use and Health (2005-2014) to assess whether race and ethnicity moderate the associations between classic psychedelic use - specifically psilocybin - and past year hypertension., Results: Hispanic identity moderated the associations between psilocybin use and past year hypertension. Furthermore, individuals who used psilocybin and identified as Non-Hispanic White had reduced odds of hypertension (aOR: 0.83); however, these associations were not observed for any other racial or ethnic groups in our study for individuals who used psilocybin., Conclusion: Overall, our results demonstrate that the associations between psychedelics and hypertension may vary by race and ethnicity. Longitudinal studies and clinical trials can further advance this research and determine whether such differences exist in causal contexts., Project Registration: https://osf.io/xsz2p/?view_only=0bf7b56749034c18abb2a3f8d3d4bc0b., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Jones, Ricard and Nock.)

Fire activity during 2020 to 2021 in California, USA, was unprecedented in the modern record. More than 19,000 km 2 of forest vegetation burned (10× more than the historical average), potentially affecting the habitat of 508 vertebrate species. Of the >9,000 km 2 that burned at high severity, 89% occurred in large patches that exceeded historical estimates of maximum high-severity patch size. In this 2-y period, 100 vertebrate species experienced fire across >10% of their geographic range, 16 of which were species of conservation concern. These 100 species experienced high-severity fire across 5 to 14% of their ranges, underscoring potentially important changes to habitat structure. Species in this region are not adapted to high-severity megafires. Management actions, such as prescribed fires and mechanical thinning, can curb severe fire behavior and reduce the potential negative impacts of uncharacteristic fires on wildlife., Competing Interests: Competing interests statement:The authors declare no competing interest.

Fire regimes are a major agent of evolution in terrestrial animals. Changing fire regimes and the capacity for rapid evolution in wild animal populations suggests the potential for rapid, fire-driven adaptive animal evolution in the Pyrocene. Fire drives multiple modes of evolutionary change, including stabilizing, directional, disruptive, and fluctuating selection, and can strongly influence gene flow and genetic drift. Ongoing and future research in fire-driven animal evolution will benefit from further development of generalizable hypotheses, studies conducted in highly responsive taxa, and linking fire-adapted phenotypes to their underlying genetic basis. A better understanding of evolutionary responses to fire has the potential to positively influence conservation strategies that embrace evolutionary resilience to fire in the Pyrocene., Competing Interests: Declaration of interests No interests are declared., (Published by Elsevier Ltd.)

The exponential computational cost of describing strongly correlated electrons can be mitigated by adopting a reduced-density matrix (RDM)-based description of the electronic structure. While variational two-electron RDM (v2RDM) methods can enable large-scale calculations on such systems, the quality of the solution is limited by the fact that only a subset of known necessary N -representability constraints can be applied to the 2RDM in practical calculations. Here, we demonstrate that violations of partial three-particle (T1 and T2) N -representability conditions, which can be evaluated with knowledge of only the 2RDM, can serve as physics-based features in a machine-learning (ML) protocol for improving energies from v2RDM calculations that consider only two-particle (PQG) conditions. Proof-of-principle calculations demonstrate that the model yields substantially improved energies relative to reference values from configuration-interaction-based calculations.

Lacosamide (LCM) is an antiseizure medication used to manage status epilepticus (SE). Previous retrospective analyses have demonstrated safety and efficiency in intravenous push (IVP) administration at 80 mg per minute. Quick administration can be achieved in a high-acuity setting without the additional time required for compounding. However, previous literature only partially represents high doses of IVP LCM, which limits the understanding of the safety profile of these doses. Our study was a single-center, retrospective, single-arm analysis of patients who received IVP LCM 300 mg or 400 mg during admission. The primary outcome was the incidence of infusion site reactions, hypotension, and bradycardia within 2 hours of IVP administration. Secondary outcomes included the incidence of PR prolongation. A total of 113 patients were evaluated for infusion site reactions. Of these, 108 patients had vital signs assessed within 2 hours of IVP LCM and could be evaluated for hypotension and bradycardia. The sample primarily consisted of LCM 400 mg IVP (85.8%). The primary outcome consisted of 7 (6.2%) infusion reactions, 12 (11.1%) hypotensive events, and no reports of bradycardia. Each adverse event was assessed using the Naranjo Adverse Drug Probability Scale. All events scored less than two, suggesting the possibility was likely related to factors other than the medication. In conclusion, LCM 300 mg and 400 mg IVP administration have the potential to facilitate more rapid treatment of seizures without additional risk of infusion site reactions, hypotension, and bradycardia., Competing Interests: The author(s) declared the following potential conflicts of interest with respect to the research, authorship, and/or publication of this article: Dr. Torian has no financial disclosures or conflicts of interest to report. When the research was conducted, Dr. Jones was a full-time employee of Methodist University Hospital. He is currently a full-time employee of Chiesi USA, which has no conflicts of interest related to this research., (© The Author(s) 2023.)

Mature forests provide important wildlife habitat and support critical ecosystem functions globally. Within the dry conifer forests of the western United States, past management and fire exclusion have contributed to forest conditions that are susceptible to increasingly severe wildfire and drought. We evaluated declines in conifer forest cover in the southern Sierra Nevada of California during a decade of record disturbance by using spatially comprehensive forest structure estimates, wildfire perimeter data, and the eDaRT forest disturbance tracking algorithm. Primarily due to the combination of wildfires, drought, and drought-associated beetle epidemics, 30% of the region's conifer forest extent transitioned to nonforest vegetation during 2011-2020. In total, 50% of mature forest habitat and 85% of high density mature forests either transitioned to lower density forest or nonforest vegetation types. California spotted owl protected activity centers (PAC) experienced greater canopy cover decline (49% of 2011 cover) than non-PAC areas (42% decline). Areas with high initial canopy cover and without tall trees were most vulnerable to canopy cover declines, likely explaining the disproportionate declines of mature forest habitat and within PACs. Drought and beetle attack caused greater cumulative declines than areas where drought and wildfire mortality overlapped, and both types of natural disturbance far outpaced declines attributable to mechanical activities. Drought mortality that disproportionately affects large conifers is particularly problematic to mature forest specialist species reliant on large trees. However, patches of degraded forests within wildfire perimeters were larger with greater core area than those outside burned areas, and remnant forest habitats were more fragmented within burned perimeters than those affected by drought and beetle mortality alone. The percentage of mature forest that survived and potentially benefited from lower severity wildfire increased over time as the total extent of mature forest declined. These areas provide some opportunity for improved resilience to future disturbances, but strategic management interventions are likely also necessary to mitigate worsening mega-disturbances. Remaining dry mature forest habitat in California may be susceptible to complete loss in the coming decades without a rapid transition from a conservation paradigm that attempts to maintain static conditions to one that manages for sustainable disturbance dynamics., (© 2022 The Ecological Society of America.)

Background: Hands-on culinary medicine education for medical trainees has emerged as a promising tool for cardiovascular health promotion., Purpose: To determine whether virtual culinary medicine programming associates with Mediterranean diet (MedDiet) adherence and lifestyle medicine competencies among medical trainees across the USA., Method: A total of 1433 medical trainees across 19 sites over a 12-month period were included. The Cooking for Health Optimisation with Patients-Medical Trainees survey composed of 61 questions regarding demographics, nutritional attitudes, dietary habits including MedDiet score and lifestyle medicine counselling competencies. Multivariable logistic regression assessed the association of virtual culinary medicine education with MedDiet intake and nutritional attitudes., Results: There were 519 medical trainees who participated in virtual culinary medicine education and 914 medical trainees who participated in their standard nutrition curricula. More than one-half of participants were women (n=759) and the mean age was 27 years old. Compared with students enrolled in traditional nutrition curricula, participants in virtual culinary medicine education were 37% more likely to adhere to MedDiet guidelines for fruit intake (OR 1.37, 95% CI 1.03 to 1.83, p=0.03). Virtual culinary medicine education was associated with higher proficiency in lifestyle medicine counselling categories, notably recommendations involving fibre (OR 4.03; 95% CI 3.05 to 5.34), type 2 diabetes prevention (OR 4.69; 95% CI 3.51 to 6.27) and omega fatty acids (OR 5.21; 95% CI 3.87 to 7.02). Virtual culinary medicine education had a similar, although higher magnitude association with MedDiet counselling competency (OR 5.73, 95% CI 4.26 to 7.70) when compared with historical data previously reported using hands-on, in-person culinary medicine courseware (OR 4.97, 95% CI 3.89 to 6.36)., Conclusions: Compared with traditional nutritional educational curricula, virtual culinary medicine education is associated with higher MedDiet adherence and lifestyle medicine counselling competencies among medical trainees. Both virtual and hands-on culinary medicine education may be useful for cardiovascular health promotion., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2023. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Background: Psychedelics are receiving renewed attention within Western medicine as they represent potential treatments for many difficult-to-treat mental health disorders. However, psychedelic science is limited in its focus and inclusion of racial and ethnic minorities. Hence, this study examines whether race and ethnicity moderate the associations that naturalistic 3,4-methylenedioxymethamphetamine (MDMA)/ecstasy use and psilocybin use share with major depressive episodes (MDEs)., Method: Data for this project are from The National Survey on Drug Use and Health (2005-2019). Participants were adults aged 18 years and older (unweighted N   =  596,187). This study used multivariable logistic regression to test the interaction between race and ethnicity and MDMA/ecstasy use and psilocybin use for predicting lifetime, past year, and past year severe MDEs., Results: Race and ethnicity significantly moderated the associations between MDMA/ecstasy use and psilocybin use and MDEs. For White participants, MDMA/ecstasy use and psilocybin use each were associated with lowered odds of all three MDE outcomes (adjusted odds ratio (aOR) range: 0.82-0.92). For Hispanic participants, MDMA/ecstasy use and psilocybin use each conferred lowered odds of only a past year MDE (MDMA/ecstasy aOR: 0.82; psilocybin aOR: 0.79). For Non-Hispanic Racial Minority participants, MDMA/ecstasy and psilocybin use did not confer lowered odds of any MDE outcomes., Conclusion: Race and ethnicity have an impact on the associations that psychedelics share with mental health outcomes. Future research should explore the impact of identity and discrimination on the effects of psychedelics and should explore whether these substances can serve as effective treatments for minorities when used in culturally informed contexts.

Cerebrospinal fluid (CSF) is a source of diagnostic biomarkers for a range of neurological conditions. Cell-free DNA (cfDNA) is detected in CSF and differences in the concentration of cell-free mitochondrial DNA have been reported in studies of neurodegenerative disorders including Alzheimer's disease (AD). However, the influence of pre-analytical steps has not been investigated for cfDNA in CSF and there is no standardised approach for quantification of total cfDNA (copies of nuclear genome or mitochondria-derived gene targets). In this study, the suitability of four extraction methods was evaluated: QIAamp Circulating Nucleic Acid (Qiagen), Quick-cfDNA Serum & Plasma (Zymo), NucleoSnap® DNA Plasma (Macherey-Nagel) and Plasma/Serum Circulating DNA Purification Mini (Norgen) kits, for cfDNA extraction from CSF of controls and AD dementia patients, utilising a spike-in control for extraction efficiency and fragment size. One of the optimal extraction methods was applied to a comparison of cfDNA concentrations in CSF from control subjects, AD dementia and primary and secondary brain tumour patients. Extraction efficiency based on spike-in recovery was similar in all three groups whilst both endogenous mitochondrial and nucleus-derived cfDNA was significantly higher in CSF from cancer patients compared to control and AD groups, which typically contained < 100 genome copies/mL. This study shows that it is feasible to measure low concentration nuclear and mitochondrial gene targets in CSF and that normalisation of extraction yield can help control pre-analytical variability influencing biomarker measurements., Competing Interests: Declaration of Competing Interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Robert Perneczky reports a relationship with Janssen that includes: consulting or advisory and speaking and lecture fees. Robert Perneczky reports a relationship with Biogen that includes: consulting or advisory and speaking and lecture fees. Robert Perneczky reports a relationship with Takeda that includes: funding grants. Robert Perneczky reports a relationship with Schwabe that includes: consulting or advisory and speaking and lecture fees. Robert Perneczky reports a relationship with Grifols that includes: consulting or advisory and speaking and lecture fees. Robert Perneczky reports a relationship with Roche that includes: consulting or advisory and speaking and lecture fees. Robert Perneczky reports a relationship with Eli Lilly that includes: consulting or advisory and speaking and lecture fees. Robert Perneczky reports a relationship with Bayer that includes: consulting or advisory and speaking and lecture fees. Robert Perneczky reports a relationship with Novo Nordisk that includes: consulting or advisory and speaking and lecture fees. Alison Devonshire reports a relationship with Roche that includes: paid expert testimony. Declaration of Competing Interest RP received speaker honoraria and consultancy fees from Janssen, Biogen, Schwabe, Grifols, Roche, Eli Lilly, Bayer and Novo Nordisk, and research support from Takeda. AD received paid expert testimony from Roche., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

Objectives: Alteplase, a tissue-type plasminogen activator, is recommended for ischemic stroke patients presenting within 4.5 h. Due to bleeding risks, current guidelines advise delaying antiplatelet therapy for 24 h after alteplase. However, specific scenarios may require antiplatelet therapy to be given within the 24 h window. This study aimed to examine the safety of early antiplatelet therapy administration within the first 24 h after alteplase., Materials and Methods: This study is a retrospective, observational study of adult patients with acute ischemic stroke who received alteplase across a multi-hospital system. Patients were grouped based on early antiplatelet therapy (within 24 h window) or as recommended per guidelines. The occurrence of bleeding events, including symptomatic intracranial hemorrhage and/or gastrointestinal bleeding, in-hospital mortality, unfavorable outcomes (modified Rankin score 3-6), and hospital length of stay, were compared between groups., Results: Patients were predominantly African American (72%) and female (53%) with a median age of 62 years. Median baseline NIHSS scores were higher in the early group (5 vs. 7; p = 0.04), and patients in the early group were more likely to undergo endovascular therapy (26% vs. 8%, p < 0.0001). In patients treated with alteplase only and who did not undergo endovascular therapy, there was no difference in symptomatic intracranial hemorrhage (1.4% vs. 0%, p = 0.1), gastrointestinal bleeding, in-hospital mortality, unfavorable outcomes, or length of stay., Conclusions: In our retrospective analysis, early administration of antiplatelet therapy (< 24 h post-alteplase) did not increase the risk of symptomatic intracranial hemorrhage, gastrointestinal bleeding, or unfavorable outcomes in patients who received alteplase alone for management of acute ischemic stroke. Prospective studies are needed to validate these findings., Competing Interests: Declaration of Competing Interest The authors declare that they have no conflict of interest., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Psychedelic compounds have been linked to salutary mental health outcomes in both naturalistic and clinical settings; however, current research on psychedelics suffers from a lack of inclusion and focus on racial and ethnic minorities. Thus, the goal of our study was to assess whether race and ethnicity moderate the associations that naturalistic lifetime MDMA (3,4-Methylenedioxymethamphetamine) use and psilocybin use share with past month psychological distress and past year suicidality (ideation and planning). Using data from the National Survey on Drug Use and Health (NSDUH) (2008-2019) (N = 484,732), we conducted survey-weighted multivariable logistic regression to conduct interaction tests and to assess the associations that MDMA use and psilocybin use share with the aforementioned outcomes for each racial and ethnic group. Race and ethnicity significantly moderated the associations between MDMA and psilocybin use and psychological distress and suicidality. For White participants, MDMA and psilocybin use conferred lowered odds of all distress and suicidality outcomes. For racial and ethnic minority participants, the associations between psychedelic use and suicidality were far fewer. These findings invite further research into the impact of race, ethnicity, and other identity factors (e.g., socioeconomic status, sexual/gender minority status) on the effects of psychedelic substances., (© 2022. The Author(s).)

Purpose of Review: Gambling disorder is a serious mental health condition and the only behavioral addiction currently recognized by the American Psychiatric Association. Though prevalence and treatment seeking for gambling disorder have been stable for decades, the global COVID-19 pandemic has created a seismic shift in societal factors that influence gambling disorder and treatment availability. At the same time, the increasing availability of telehealth therapy for gambling disorder offers a new way to provide specialized treatment nationwide., Recent Findings: The present review discusses the seismic shifts in underlying risk factors, gambling availability, and reduced access to treatment. It also covers the emergence of telehealth treatment for gambling disorder, including its benefits and pitfalls., Summary: In separate commentaries, several experts hypothesized that the pandemic was likely to increase mental health risk factors in society such as social isolation, job loss, and stress. In addition, access to online gambling also increased, while countries that locked down more strictly (like Singapore) saw a reduction in terrestrial gambling, countries that did not lock down strictly (like Sweden) did not see changes in gambling behavior. Telehealth was almost universally recommended as a good option for treatment in the studies we reviewed, though proper training and staffing are needed for telehealth to prove effective. Gambling disorder was only included in nationwide addiction telehealth programs in two countries., (Copyright © 2022 Wolters Kluwer Health, Inc. All rights reserved.)

International demand for wood and other forest products continues to grow rapidly, and uncertainties remain about how animal communities will respond to intensifying resource extraction associated with woody bioenergy production. We examined changes in alpha and beta diversity of bats, bees, birds, and reptiles across wood production landscapes in the southeastern United States, a biodiversity hotspot that is one of the principal sources of woody biomass globally. We sampled across a spatial gradient of paired forest land-uses (representing pre and postharvest) that allowed us to evaluate biological community changes resulting from several types of biomass harvest. Short-rotation practices and residue removal following clearcuts were associated with reduced alpha diversity (-14.1 and -13.9 species, respectively) and lower beta diversity (i.e., Jaccard dissimilarity) between land-use pairs (0.46 and 0.50, respectively), whereas midrotation thinning increased alpha (+3.5 species) and beta diversity (0.59). Over the course of a stand rotation in a single location, biomass harvesting generally led to less biodiversity. Cross-taxa responses to resource extraction were poorly predicted by alpha diversity: correlations in responses between taxonomic groups were highly variable (-0.2 to 0.4) with large uncertainties. In contrast, beta diversity patterns were highly consistent and predictable across taxa, where correlations in responses between taxonomic groups were all positive (0.05-0.4) with more narrow uncertainties. Beta diversity may, therefore, be a more reliable and information-rich indicator than alpha diversity in understanding animal community response to landscape change. Patterns in beta diversity were primarily driven by turnover instead of species loss or gain, indicating that wood extraction generates habitats that support different biological communities., (© 2021 Society for Conservation Biology.)

Effective conservation requires understanding the processes that determine population outcomes. Too often, we assume that protected areas conserve wild populations despite evidence that they frequently fail to do so. Without large-scale studies, however, we cannot determine what relationships are the product of localized conditions versus general patterns that inform conservation more broadly. Leopards' (Panthera pardus) basic ecology is well studied but little research has investigated anthropogenic effects on leopard density at broad scales. We investigated the drivers of leopard density among 27 diverse protected areas in northeastern South Africa to understand what conditions facilitate abundant populations. We formulated 10 working hypotheses that considered the relative influence of bottom-up biological factors and top-down anthropogenic factors on leopard density. Using camera-trap survey data, we fit a multi-session spatial capture-recapture model with inhomogenous density for each hypothesis and evaluated support using an information theoretic approach. The four supported hypotheses indicated that leopard density is primarily limited by human impacts, but that habitat suitability and management conditions also matter. The proportion of camera stations that recorded domestic animals, a proxy for the extent of human impacts and protected area effectiveness, was the only predictor variable present in all four supported models. Protected areas are the cornerstone of large felid conservation, but only when the human-wildlife interface is well managed and protected areas shelter wildlife populations from anthropogenic impacts. To ensure the long-term abundance of large carnivore populations, reserve managers should recognize the ineffectiveness of "paper parks" and promote contiguous networks of protected areas that offer leopards and other large mammal populations greater space and reduced human impacts., (© 2022 The Ecological Society of America.)

Esophageal adenocarcinoma (EAC) develops from a chronic inflammatory environment across four stages: intestinal metaplasia, known as Barrett's esophagus, low- and high-grade dysplasia, and adenocarcinoma. Although the genomic characteristics of this progression have been well defined via large-scale DNA sequencing, the dynamics of various immune cell subsets and their spatial interactions in their tumor microenvironment remain unclear. Here, we applied a sequential multiplex immunohistochemistry (mIHC) platform with computational image analysis pipelines that allow for the detection of 10 biomarkers in one formalin-fixed paraffin-embedded (FFPE) tissue section. Using this platform and quantitative image analytics, we studied changes in the immune landscape during disease progression based on 40 normal and diseased areas from endoscopic mucosal resection specimens of chemotherapy treatment- naïve patients, including normal esophagus, metaplasia, low- and high-grade dysplasia, and adenocarcinoma. The results revealed a steady increase of FOXP3 + T regulatory cells and a CD163 + myelomonocytic cell subset. In parallel to the manual gating strategy applied for cell phenotyping, we also adopted a sparse subspace clustering (SSC) algorithm allowing the automated cell phenotyping of mIHC-based single-cell data. The algorithm successfully identified comparable cell types, along with significantly enriched FOXP3 T regulatory cells and CD163 + myelomonocytic cells as found in manual gating. In addition, SCC identified a new CSF1R + CD1C + myeloid lineage, which not only was previously unknown in this disease but also increases with advancing disease stages. This study revealed immune dynamics in EAC progression and highlighted the potential application of a new multiplex imaging platform, combined with computational image analysis on routine clinical FFPE sections, to investigate complex immune populations in tumor ecosystems., Competing Interests: LC reports consulting services for Cell Signaling Technologies, AbbVie, the Susan G Komen Foundation, and Shasqi, received reagent and/or research support from Cell Signaling Technologies, Syndax Pharmaceuticals, and Acerta Pharma, and has participated in advisory boards for Pharmacyclics, Syndax, Carisma, Verseau, CytomX, Kineta, Hibercell, Cell Signaling Technologies, Alkermes, Zymeworks, the AstraZeneca Partner of Choice Network, the Lustgarten Foundation, and the NIH/NCI-Frederick National Laboratory Advisory Committee. RF holds patents related to Cytosponge-TFF3 and related assays that have been licensed by the Medical Research Council to Covidien (now Medtronic). RF is a co-founder and shareholder in an early detection and digital pathology company Cyted Ltd. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Sundaram, Kim, Jones, Sivagnanam, Tripathi, Miremadi, Di Pietro, Coussens, Fitzgerald, Chang and Zhuang.)

A candidate digital PCR (dPCR)-based reference measurement procedure for quantification of human cytomegalovirus (hCMV) was evaluated in 10 viral load comparison schemes (seven external quality assessment (EQA) and three additional training schemes) organized by INSTAND e.V. over four years (between September 2014 and March 2018). Four metrology institutes participated in these schemes using the same extraction method and dPCR measurement procedure for the hCMV specific target sequence of UL54 gene. The calibration independent reference measurement procedure results from the metrology institutes were compared to the results of the clinical diagnostic laboratories applying hCMV qPCR measurement procedures calibrated to reference materials. While the criteria for the acceptable deviation from the target value interval for INSTAND's EQA schemes is from -0.8 log 10 to +0.8 log 10 , the majority of dPCR results were between -0.2 log 10 to +0.2 log 10 . Only 4 out of 45 results exceeded this interval with the maximum deviation of -0.542 log 10 . In the training schemes containing samples with lower hCMV concentrations, more than half of the results deviated less than ±0.2 log 10 from the target value, while more than 95% deviated less than ±0.4 log 10 from the target value. Evaluation of intra- and inter-laboratory variation of dPCR results confirmed high reproducibility and trueness of the method. This work demonstrates that dPCR has the potential to act as a calibration independent reference measurement procedure for the value assignment of hCMV calibration and reference materials to support qPCR calibration as well as ultimately for routine hCMV load testing., (Copyright © 2021 The Author(s). Published by Elsevier Inc. All rights reserved.)