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Previous work suggests that sometimes the more people value happiness, the less happy they are. For whom and why is this the case? To answer these questions, we examined a model of happiness pursuit that disentangles two previously conflated individual differences related to valuing happiness. The first individual difference operates at the strength of the value itself and involves viewing happiness as a very important goal (i.e., aspiring to happiness). The second individual difference occurs later in the process of pursuing happiness and involves judging one's levels of happiness (i.e., concern about happiness). This model predicts that aspiring to happiness is relatively innocuous. Conversely, being concerned about happiness leads people to judge their happiness, thereby infusing negativity (i.e., negative meta-emotions) into potentially positive events, which, in turn, interferes with well-being. We tested these hypotheses using cross-sectional, daily-diary, and longitudinal methods in student and community samples, collected between 2009 and 2020, which are diverse in gender, ethnicity, age, and geographic location (Ntotal = 1,815). In Studies 1a and 1b, aspiring to happiness and concern about happiness represented distinct individual differences. In Study 2, concern about happiness (but not aspiring to happiness) was associated with lower well-being cross-sectionally and longitudinally. In Study 3, these links between concern about happiness and worse well-being were partially accounted for by experiencing greater negative meta-emotions during daily positive events. These findings suggest that highly valuing happiness is not inherently problematic; however, concern and judgment about one's happiness can undermine it. (PsycInfo Database Record (c) 2024 APA, all rights reserved).
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Progressive retinal atrophies (PRAs) are a genetically heterogeneous group of inherited eye diseases that affect over 100 breeds of dog. The initial clinical sign is visual impairment in scotopic conditions, as a consequence of rod photoreceptor cell degeneration. Photopic vision degeneration then follows, due to progression of the disease to the cone photoreceptors, and ultimately results in complete blindness. Two full-sibling English Shepherds were diagnosed with PRA at approximately 5 years old and tested clear of all published PRA genetic variants. This study sought to identify the novel PRA-associated variant segregating in the breed. We utilised a combined approach of whole genome sequencing of the probands and homozygosity mapping of four cases and 22 controls and identified a short interspersed nuclear element within an alternatively spliced exon in FAM161A. The XP_005626197.1 c.17929_ins210 variant was homozygous in six PRA cases and heterozygous or absent in control dogs, consistent with a recessive mode of inheritance. The insertion is predicted to extend exon 4 by 39 aberrant amino acids followed by an early termination stop codon. PRA is intractable to treatment, so the development of a genetic screening test, based on the associated variant, is significant, because it provides dog breeders/owners with a means of reducing the frequency of the disease variant within this breed as well as minimising the risk of breeding puppies that will develop this blinding disease.
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Doenças do Cão , Éxons , Animais , Cães , Éxons/genética , Doenças do Cão/genética , Doenças do Cão/patologia , Elementos Nucleotídeos Curtos e Dispersos/genética , Degeneração Retiniana/genética , Degeneração Retiniana/veterinária , Degeneração Retiniana/patologia , Feminino , Masculino , Linhagem , Genes Recessivos , Mutagênese Insercional , Proteínas do Olho/genética , Sequenciamento Completo do GenomaRESUMO
Single-cell RNA sequencing (scRNAseq) and Variable, Diversity, Joining (VDJ) profiling have improved our understanding of B-cells. Recent scRNAseq-based approaches have led to the discovery of intermediate B-cell states, including preplasma cells and pregerminal centre B-cells, as well as unveiling protective roles for B-cells within tertiary lymphoid structures in respiratory infections and cancers. These studies have improved our understanding of transcriptional and epigenetic control of B-cell development and of atypical and memory B-cell differentiation. Advancements in temporal profiling in parallel with transcriptomic and VDJ sequencing have consolidated our understanding of the trajectory of B-cell clones over the course of infection and vaccination. Challenges remain in studying B-cell states across tissues in humans, in relating spatial location with B-cell phenotype and function, in examining antibody isotype switching events, and in unequivocal determination of clonal relationships. Nevertheless, ongoing multiomic assessments and studies of cellular interactions within tissues promise new avenues for improving humoral immunity and combatting autoimmune conditions.
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Linfócitos B , Análise de Célula Única , Transcriptoma , Humanos , Análise de Célula Única/métodos , Linfócitos B/imunologia , Animais , Perfilação da Expressão Gênica , Diferenciação Celular/genética , Diferenciação Celular/imunologiaRESUMO
Birds maintain some of the highest body temperatures among endothermic animals. Often deemed a selective advantage for heat tolerance, high body temperatures also limits birds' thermal safety margin before reaching lethal levels. Recent modelling suggests that sustained effort in Arctic birds might be restricted at mild air temperatures, which may require reductions in activity to avoid overheating, with expected negative impacts on reproductive performance. We measured within-individual changes in body temperature in calm birds and then in response to an experimental increase in activity in an outdoor captive population of Arctic, cold-specialised snow buntings (Plectrophenax nivalis), exposed to naturally varying air temperatures (- 15 to 36 °C). Calm buntings exhibited a modal body temperature range from 39.9 to 42.6 °C. However, we detected a significant increase in body temperature within minutes of shifting calm birds to active flight, with strong evidence for a positive effect of air temperature on body temperature (slope = 0.04 °C/ °C). Importantly, by an ambient temperature of 9 °C, flying buntings were already generating body temperatures ≥ 45 °C, approaching the upper thermal limits of organismal performance (45-47 °C). With known limited evaporative heat dissipation capacities in these birds, our results support the recent prediction that free-living buntings operating at maximal sustainable rates will increasingly need to rely on behavioural thermoregulatory strategies to regulate body temperature, to the detriment of nestling growth and survival.
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Temperatura Baixa , Aves Canoras , Animais , Regiões Árticas , Aves Canoras/fisiologia , Regulação da Temperatura Corporal/fisiologia , Temperatura Corporal/fisiologia , Cruzamento , Reprodução/fisiologia , Feminino , Masculino , TemperaturaRESUMO
Creativity is core to being human. Generative artificial intelligence (AI)-including powerful large language models (LLMs)-holds promise for humans to be more creative by offering new ideas, or less creative by anchoring on generative AI ideas. We study the causal impact of generative AI ideas on the production of short stories in an online experiment where some writers obtained story ideas from an LLM. We find that access to generative AI ideas causes stories to be evaluated as more creative, better written, and more enjoyable, especially among less creative writers. However, generative AI-enabled stories are more similar to each other than stories by humans alone. These results point to an increase in individual creativity at the risk of losing collective novelty. This dynamic resembles a social dilemma: With generative AI, writers are individually better off, but collectively a narrower scope of novel content is produced. Our results have implications for researchers, policy-makers, and practitioners interested in bolstering creativity.
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Inteligência Artificial , Criatividade , Humanos , Idioma , RedaçãoRESUMO
INTRODUCTION: Dance Medicine is a growing discipline, and healthcare for dancers requires appreciating particularities of their artistic craft. Manual therapists (MTs) are often dancers' first choice when seeking care. Whilst dancers' experiences of injury and healthcare have been extensively researched, little is known of MTs' perspectives of caring for them. AIM: This study aimed to gain an understanding of MTs' perspectives of caring for dancers. Communication, treatment strategies and obstacles to a successful therapeutic alliance were examined to broaden knowledge and improve care. METHODS: This was a qualitative study using Grounded Theory as a framework for data collection and analysis. A purposive sample of 8 participants was recruited-physiotherapists, chiropractors, and osteopaths with 2+ years' experience of treating dancers. Data collection was conducted via semi-structured interviews, which were transcribed verbatim, anonymized, member checked, and later analyzed. RESULTS: Data analysis constructed 3 categories which described the range of participants' experiences. Two categories were identified-(i) A Strangled Industry and (ii) Be-all End-all Patients-and found to culminate in a core category: (iii) Beyond Hands-on, which exemplified MTs' attempts to address clinical challenges. CONCLUSIONS: MTs experience several challenges when providing healthcare to dancers. These encompass psychosocial aspects relating to dancers' internal world and the dance industry environment. It suggests that caring for dancers demands comprehensive, collaborative, and psychologically informed approaches. Emphasis should also be given to developing and implementing dance-specific pain science for all stakeholders.
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Children in malaria-endemic regions can experience repeated Plasmodium infections over short periods of time. Effects of re-infection on multiple co-existing CD4+ T cell subsets remain unresolved. Here, we examine antigen-experienced CD4+ T cells during re-infection in mice, using scRNA-seq/TCR-seq and spatial transcriptomics. TCR transgenic TEM cells initiate rapid Th1/Tr1 recall responses prior to proliferating, while GC Tfh counterparts are refractory, with TCM/Tfh-like cells exhibiting modest non-proliferative responses. Th1-recall is a partial facsimile of primary Th1-responses, with no upregulated effector-associated genes being unique to recall. Polyclonal, TCR-diverse, CD4+ T cells exhibit similar recall dynamics, with individual clones giving rise to multiple effectors including highly proliferative Th1/Tr1 cells, as well as GC Tfh and Tfh-like cells lacking proliferative capacity. Thus, we show substantial diversity in recall responses mounted by multiple co-existing CD4+ T cell subsets in the spleen, and present graphical user interfaces for studying gene expression dynamics and clonal relationships during re-infection.
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Linfócitos T CD4-Positivos , Malária , Reinfecção , Animais , Malária/imunologia , Malária/parasitologia , Linfócitos T CD4-Positivos/imunologia , Camundongos , Reinfecção/imunologia , Células Th1/imunologia , Camundongos Endogâmicos C57BL , Baço/imunologia , Baço/parasitologia , Receptores de Antígenos de Linfócitos T/metabolismo , Receptores de Antígenos de Linfócitos T/imunologia , Receptores de Antígenos de Linfócitos T/genética , Camundongos Transgênicos , Feminino , Memória ImunológicaRESUMO
Naive CD4+ T cells must differentiate in order to orchestrate immunity to Plasmodium, yet understanding of their emerging phenotypes, clonality, spatial distributions, and cellular interactions remains incomplete. Here, we observe that splenic polyclonal CD4+ T cells differentiate toward T helper 1 (Th1) and T follicular helper (Tfh)-like states and exhibit rarer phenotypes not elicited among T cell receptor (TCR) transgenic counterparts. TCR clones present at higher frequencies exhibit Th1 skewing, suggesting that variation in major histocompatibility complex class II (MHC-II) interaction influences proliferation and Th1 differentiation. To characterize CD4+ T cell interactions, we map splenic microarchitecture, cellular locations, and molecular interactions using spatial transcriptomics at near single-cell resolution. Tfh-like cells co-locate with stromal cells in B cell follicles, while Th1 cells in red pulp co-locate with activated monocytes expressing multiple chemokines and MHC-II. Spatial mapping of individual transcriptomes suggests that proximity to chemokine-expressing monocytes correlates with stronger effector phenotypes in Th1 cells. Finally, CRISPR-Cas9 gene disruption reveals a role for CCR5 in promoting clonal expansion and Th1 differentiation. A database of cellular locations and interactions is presented: https://haquelab.mdhs.unimelb.edu.au/spatial_gui/.
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Linfócitos T CD4-Positivos , Diferenciação Celular , Malária , Animais , Camundongos , Linfócitos T CD4-Positivos/citologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Malária/imunologia , Malária/parasitologia , Camundongos Endogâmicos C57BL , Fenótipo , Receptores de Antígenos de Linfócitos T/metabolismo , Receptores CCR5/metabolismo , Receptores CCR5/genética , Baço/imunologia , Células Th1/imunologiaRESUMO
The clinical impact of platelet-rich fibrin (PRF) and plasma rich in growth factors (PRGF®) respectively has been studied extensively in the field of regenerative dentistry during the last two decades. Literature supports evidence for additional benefits in regenerative periodontal therapy, alveolar ridge preservation, management of extraction sockets, implantology including guided bone regeneration as well as defect management in oral surgery. Regarding gingival wound healing and soft tissue regeneration, there is sufficient evidence for their positive effects which have been confirmed in several systematic reviews. The effects seem less clear in conjunction with osseous regenerative treatments, where the inter-study heterogenity in terms of different PRF-protocols, indications and application forms might hinder a systematic comparison. Nevertheless there is evidence that PRF might have beneficial effects on hard-tissue or its regeneration respectively.For being able to facilitate conclusions in systematic reviews, precise reporting of the used PRF-protocols is mandatory for future (clinical) research in the field of autologous platelet concentrates.
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Fibrina Rica em Plaquetas , Plasma Rico em Plaquetas , Humanos , Regeneração Tecidual Guiada Periodontal/métodos , Plaquetas/fisiologia , Regeneração Óssea/fisiologia , Regeneração Óssea/efeitos dos fármacos , Cicatrização/fisiologia , Cicatrização/efeitos dos fármacos , Medicina Regenerativa/métodosRESUMO
The use of autologous platelet concentrates (APC) such as platelet-rich fibrin (PRF) and/or plasma rich in growth factors (PRGF®) is considered an established treatment modality in re-generative dentistry. The possibility of delivering growth factors over aclinically relevant time of several days seems particularly interesting in the context of wound healing.The growing body of evidence in the field of APC requires a continuous and actual knowledge of the literature for being able to make evidence-based treatment recommendations with a realistic assessment of possible advantages of this technology.PR(G)F can be applied in solid or liquid form, pure or in combination with other biomaterials. Both appear to be reasonable, depending on the clinical indication and/or desired treatment outcomes. Because of the many different factors that can affect the PR(G)F products final characteristics, a basic understanding of these parameters is desirable for choosing the most suitable product and/or optimizing its clinical application. This review aims to provide an over-view of relevant theoretical, practical, legal and biologic aspects of APCs.
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Fibrina Rica em Plaquetas , Humanos , Plasma Rico em Plaquetas , Plaquetas/fisiologia , Peptídeos e Proteínas de Sinalização Intercelular/uso terapêutico , Cicatrização/fisiologiaRESUMO
Attention-deficit/hyperactivity disorder (ADHD) is a childhood-onset neurodevelopmental condition characterized by developmentally extreme and impairing symptoms of inattention and/or hyperactivity/impulsivity. Great interest has emerged in the ways ADHD and its underlying symptom dimensions relate to the development of personality traits. Much extant research on this topic is cross-sectional, relying on self-report measures and male samples. Herein, we present data from a prospective, longitudinal study of a socioeconomically and racially diverse sample of girls, including those with ADHD and a matched neurotypical comparison sample. We examined how parent- and teacher-reported ADHD in middle childhood relate to self-reported Big Five personality traits in adolescence. As expected, childhood ADHD diagnosis prospectively predicted lower self-reported Conscientiousness, lower Agreeableness, and higher Neuroticism in adolescence. With ADHD diagnosis covaried, Inattention (IA) predicted only low Conscientiousness, Hyperactivity/Impulsivity (HI) predicted only low Agreeableness, and neither predicted adolescent Neuroticism. An exploratory moderator analysis showed that family income moderated the effects of IA and HI on the negativity of adolescent self-descriptions of their own personalities, with more pronounced negative effects for girls in families with higher (rather than lower) income. Familial pressures to achieve in higher-income families may be linked to more pronounced negative ramifications of ADHD on personality development.
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Cats with a distinctive white hair pattern of unknown molecular cause have been discovered in the Finnish domestic cat population. Based on the unique appearance of these cats, we have named this phenotype salmiak ("salty licorice"). The use of a commercially available panel test to genotype four salmiak-colored cats revealed the absence of all known variants associated with white-haired phenotypic loci: full White (W), Spotting (Ws) and the Birman white Gloves associated (wg) allele of the KIT proto-oncogene (KIT) gene. Whole-genome sequencing on two salmiak-colored cats was conducted to search for candidate causal variants in the KIT gene. Despite a lack of coding variants, visual inspection of the short read alignments revealed a large ~95 kb deletion located ~65 kb downstream of the KIT gene in the salmiak cats. Additional PCR genotyping of 180 domestic cats and three salmiak-colored cats confirmed the homozygous derived variant genotype fully concordant with the salmiak phenotype. We suggest the newly identified variant be designated as wsal for "w salmiak".
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Cor de Cabelo , Proteínas Proto-Oncogênicas c-kit , Animais , Gatos/genética , Cor de Cabelo/genética , Proteínas Proto-Oncogênicas c-kit/genética , Fenótipo , Deleção de Sequência , Finlândia , Genótipo , Sequenciamento Completo do Genoma/veterináriaRESUMO
BACKGROUND: Individuals from minority groups have historically faced social injustices. Those from underrepresented groups have been less likely to access both healthcare services and higher education. Little is known about the experiences of underrepresented students during their undergraduate studies in osteopathy in the UK. The aim of this project was to explore awareness of cultural diversity and beliefs about patients from underrepresented groups in current osteopathic educational environments and evaluate students' preparedness to manage patients from diverse groups. The project also aimed to investigate the educational experiences of students from underrepresented backgrounds during their training and their opinions on changes that could support better levels of recruitment and achievement. The findings were discussed with stakeholders in interactive workshops with the aim to develop recommendations for action and change. METHODS: A transformative action research paradigm informed this mixed methods project. It included: 1/ a survey of students from all seven osteopathic educational providers in the UK using the Multidimensional Cultural Humility Scale (MCHS); 2/ a series of focus groups with students from underrepresented groups (women, students with disabilities, students from minority ethnic backgrounds, and students identifying as LGBTQIA+); and 3/ a workshop forum to discuss findings. RESULTS: A total of 202 participants completed the MCHS and demographic questionnaire and seven focus groups were conducted. A model was developed to describe participants' training experiences comprising two main themes: institutional contextual obstacles (with four sub-themes) and underrepresented students' conceptual understanding of Equity, Diversity and Inclusion (EDI). Recommendations for change identified in the workshops were based on three topics: institutions, staff, and students. CONCLUSION: Our findings confirm conclusions from other institutions that staff education is urgently needed to create and maintain equitable, inclusive environments in osteopathic educational institutions in the UK to support all students, particularly those from underrepresented groups. Institutional EDI processes and policies also need to be clarified or modified to ensure their usefulness, accessibility, and implementation.
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Diversidade Cultural , Grupos Focais , Grupos Minoritários , Medicina Osteopática , Humanos , Medicina Osteopática/educação , Feminino , Masculino , Reino Unido , Estudantes de Medicina/psicologia , Adulto , Inquéritos e Questionários , Adulto JovemRESUMO
In the pursuit of terminal tin chalcogenides, heteroleptic stannylenes bearing terphenyl- and hexamethyldisilazide ligands were reacted with carbodiimides to yield the respective guanidinato complexes. Further supported by quantum chemical calculations, this revealed that the iso-propyl-substituted derivative provides the maximum steric protection achievable. Oxidation with elemental selenium produced monomeric terminal tin selenides with four-coordinate tin centers. In reactions with N2O as oxygen transfer reagent, silyl migration toward putative terminal tin oxide intermediates gave rise to tin complexes with terminal âOSiMe3 functionality. To prevent silyl migration, the silyl groups were substituted with cyclohexyl moieties. This analogue exhibited distinctively different reactivities toward selenium and N2O, yielding a 1,2,3,4,5-tetraselenastannolane and chalcogenide-bridged dimeric compounds, respectively.
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OBJECTIVE: To organize the experience and international knowledge in the surgical management and staging of colorectal endometriosis, with a management proposal in stages. METHOD: An extensive non-systematic review of the literature was carried to organize the disease in stages (limited, intermediate and advanced) according to a scoring system, which considers the characteristics of the endometrioma, the personal history and surgical findings. We tested the proposed staging in a retrospective group of patients. RESULTS: From January 2017 to April 2023, we collected 19 patients with a confirmed diagnosis of colorectal endometriosis, treated laparoscopically, by the same group of surgeons, in whom we found a strong correlation between the stage of the disease and the presence of complications that required reinterventions. CONCLUSIONS: We suggest a sequence of colorectal surgical management in stages according to the staging of the disease and we hope that this work will be followed by joint efforts to test it prospectively in order to compare results between hospital centers and make planned decisions.
OBJETIVO: Organizar la experiencia y el conocimiento internacional en el manejo quirúrgico y la estadificación de la endometriosis colorrectal, con una propuesta de manejo por etapas. MÉTODO: Se realizó una revisión amplia no sistemática de la literatura para organizar la enfermedad en etapas (limitada, intermedia y avanzada) de acuerdo con un sistema de puntuación que considera las características del endometrioma, los antecedentes personales y los hallazgos en la cirugía. La estatificación propuesta se probó en un grupo retrospectivo de pacientes. RESULTADOS: De enero de 2017 a abril de 2023 recopilamos 19 pacientes con diagnóstico confirmado de endometriosis colorrectal, tratadas por vía laparoscópica, por el mismo grupo de cirujanos, en las que encontramos una fuerte correlación entre el estadio de la enfermedad y la presencia de complicaciones que requirieron reintervenciones. CONCLUSIONES: Sugerimos una secuencia de manejo quirúrgico colorrectal en etapas de acuerdo con la estadificación de la enfermedad y esperamos que el presente trabajo sea seguido de esfuerzos compartidos por probarla de manera prospectiva para poder comparar resultados entre centros hospitalarios y tomar decisiones planificadas.
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Neoplasias Colorretais , Endometriose , Laparoscopia , Feminino , Humanos , Endometriose/cirurgia , Endometriose/complicações , Estudos Retrospectivos , Neoplasias Colorretais/cirurgia , Laparoscopia/métodosRESUMO
BACKGROUND: Chiropractors, osteopaths and physiotherapists (COPs) can assess and manage musculoskeletal conditions with similar manual or physical therapy techniques. This overlap in scope of practice raises questions about the boundaries between the three professions. Clinical settings where they are co-located are one of several possible influences on professional boundaries and may provide insight into the nature of these boundaries and how they are managed by clinicians themselves. OBJECTIVES: To understand the nature of professional boundaries between COPs within a co-located clinical environment and describe the ways in which professional boundaries may be reinforced, weakened, or navigated in this environment. METHODS: Drawing from an interpretivist paradigm, we used ethnographic observations to observe interactions between 15 COPs across two clinics. Data were analysed using reflexive thematic analysis principles. RESULTS: We identified various physical and non-physical 'boundary objects' that influenced the nature of the professional boundaries between the COPs that participated in the study. These boundary objects overall seemed to increase the fluidity of the professional boundaries, at times simultaneously reinforcing and weakening them. The boundary objects were categorised into three themes: physical, including the clinic's floor plan, large and small objects; social, including identities and discourse; and organisational, including appointment durations and fees, remuneration policies and insurance benefits. CONCLUSIONS: Physical, social, organisational related factors made the nature of professional boundaries between COPs in these settings fluid; meaning that they were largely not rigid or fixed but rather flexible, responsive and subject to change. These findings may challenge patients, clinicians and administrators to appreciate that traditional beliefs of distinct boundaries between COPs may not be so in co-located clinical environments. Both clinical practice and future research on professional boundaries between COPs may need to further consider some of these broader factors.
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Doenças Musculoesqueléticas , Médicos Osteopáticos , Fisioterapeutas , Humanos , Atitude do Pessoal de Saúde , Antropologia CulturalRESUMO
Interspecific foraging associations (IFAs) are biological interactions where two or more species forage in association with each other. Climate-induced reductions in Arctic sea ice have increased polar bear (Ursus maritimus) foraging in seabird colonies, which creates foraging opportunities for avian predators. We used drone video of bears foraging within a common eider (Somateria mollissima) colony on East Bay Island (Nunavut, Canada) in 2017 to investigate herring gull (Larus argentatus) foraging in association with bears. We recorded nest visitation by gulls following n = 193 eider flushing events from nests during incubation. The probability of gulls visiting eider nests increased with higher number of gulls present (ß = 0.14 ± 0.03 [SE], p < .001) and for nests previously visited by a bear (ß = 1.14 ± 0.49 [SE], p < .02). In our model examining the probability of gulls consuming eggs from nests, we failed to detect statistically significant effects for the number of gulls present (ß = 0.09 ± 0.05 [SE], p < .07) or for nests previously visited by a bear (ß = -0.92 ± 0.71 [SE], p < .19). Gulls preferred to visit nests behind bears (χ2 = 18, df = 1, p < .0001), indicating gulls are risk averse in the presence of polar bears. Our study provides novel insights on an Arctic IFA, and we present evidence that gulls capitalize on nests made available due to disturbance associated with foraging bears, as eiders disturbed off their nest allow gulls easier access to eggs. We suggest the IFA between gulls and polar bears is parasitic, as gulls are consuming terrestrial resources which would have eventually been consumed by bears. This finding has implications for estimating the energetic contribution of bird eggs to polar bear summer diets in that the total number of available clutches to consume may be reduced due to avian predators.
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Enzymes of the enolase superfamily share a conserved structure and a common partial reaction (i.e., metal-assisted, Brønsted base-catalyzed enol(ate) formation). The architectures of the enolization apparatus at the active sites of the mandelate racemase (MR)-subgroup members MR and l-fuconate dehydratase (FucD) are almost indistinguishable at the structural level. Tartronate and 3-hydroxypyruvate (3-HP) recognize the enolization apparatus and can be used to interrogate the active sites for differences that may not be apparent from structural data. We report a circular dichroism-based assay of FucD activity that monitors the change in ellipticity at 216 nm (Δ[Θ]S-P = 8985 ± 87 deg cm2 mol-1) accompanying the conversion of l-fuconate to 2-keto-3-deoxy-l-fuconate. Tartronate was a linear mixed-type inhibitor of FucD (Ki = 8.4 ± 0.7 mM, αKi = 63 ± 11 mM), binding 18-fold weaker than l-fuconate, compared with 2-fold weaker binding of tartronate by MR relative to mandelate. 3-HP irreversibly inactivated FucD (kinact/KI = 0.018 ± 0.002 M-1s-1) with an efficiency that was â¼4.6 × 103-fold less than that observed with MR. The inactivation arose predominantly from modifications at multiple sites and Tris-HCl, but not l-fuconate, afforded protection against inactivation. Similar to the reaction of 3-HP with MR, 3-HP modified the Brønsted base catalyst (Lys 220) at the active site of FucD, which was facilitated by the Brønsted acid catalyst His 351. Thus, the interactions of tartronate and 3-HP with MR and FucD revealed differences in binding affinity and reactivity that differentiated between the enzymes' enolization apparatuses.
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Fosfopiruvato Hidratase , Tartronatos , Fosfopiruvato Hidratase/química , Fosfopiruvato Hidratase/metabolismo , Hidroliases/química , Racemases e Epimerases/metabolismo , CinéticaRESUMO
The compound [5,10,15,20-tetrakis(4-fluoro-2,6-dimethylphenyl)porphyrinato]platinum(II), [Pt(C52H40F4N4)] or Pt(II)TFP, has been synthesized and structurally characterized by single-crystal X-ray crystallography. The Pt porphyrin exhibits a long-lived phosphorescent excited state (τ0 = 66â µs), which has been characterized by transient absorption and emission spectroscopy. The phosphorescence is extremely sensitive to oxygen, as reflected by a quenching rate constant of 5.0 × 108â M-1â s-1, and as measured by Stern-Volmer quenching analysis.