The prognostic role of PD-L1 expression and the presence of polyomavirus in Merkel cell carcinoma cases.

BACKGROUND: Merkel cell carcinoma (MCC) comprises a rare malignant primary skin tumor presenting neuroendocrine differentiation. Recently, agents blocking the programmed cell death protein 1 and programmed cell death protein ligand 1 pathway (PD-1/PD-L1) have demonstrated objective and durable tumor regressions in patients presenting advanced MCC. This study aimed to describe the sociodemographic, clinical, and histopathological characteristics of MCC patients, also assessing the prevalence of PD-L1 expression and Merkel cell Polyomavirus (MCPyV), as well as their prognostic roles. METHODS: Data from patients diagnosed with MCC between 1996 and 2019 at a reference cancer center in Rio de Janeiro, southeastern Brazil, were evaluated in a retrospective study. Tumor samples were tested for MCPyV and PD-L1 employing immunohistochemistry. Survival analyses were carried out employing the Kaplan-Meier method and curves were compared using the log-rank test. A multiple semiparametric Cox model was used. Values p < 0.05 were considered significant. RESULTS: A total of 65 patients were included in the study, with a mean age at diagnosis of 72 (standard deviation 13.9). A total of 56.9% (37/65) of the patients were male, 86.2% (56/65) were white, and 56.9% (37/64) were illiterate or with incomplete elementary school. MCPyV immunohistochemistry was positive in 29 cases (44.6%) and PD-L1 positivity was ≥ 1% in 42 cases (64.6%). Significant associations between MCPyV and PD-L1 expression ≥ 1% (p = 0.003) and PD-L1 expression ≥ 5% (p = 0.005) were noted. Concerning the multivariate analysis, only education level and advanced MCC stage indicated statistically significant worse progression-free survival. Regarding overall survival (OS), being male, education level and advanced stage comprised risk factors. The estimated OS at 60 months for stages I to III was of 48.9% and for stage IV, 8.9%. CONCLUSIONS: This is the first large Brazilian cohort to assess the prevalence of MCPyV in MCC tumors, as well as PD-L1 expression and their associations. No correlations were noted between MCPyV infection or PD-L1 expression and survival rates.

Evaluation of Two Configurations of Hydroxyapatite and Beta-Tricalcium Phosphate in Sinus Grafts with Simultaneous Implant Installation: An Experimental Study in Rabbits.

BACKGROUND: This study aimed to evaluate peri-implant bone formation in rabbits after sinus grafting mediated by hydroxyapatite and beta-tricalcium phosphate (HA + ß-TCP) in granule or paste configurations, concomitant with immediate implant installation. MATERIAL & METHODS: Thirty-four rabbit maxillary sinuses were grafted with HA + ß-TCP, half of which were applied in a granule and half in a paste composition. Implant placement was performed simultaneously. At 7 and 40 days postoperatively, the animals were euthanized, and samples were prepared for tomographic, microtomographic, histological, histometric (hematoxylin and eosin staining, HE), and immunohistochemical (labeling of transcription factor Runx-2 [RUNX2], vascular endothelial growth factor [VEGF], osteocalcin [OCN], and tartrate-resistant acid phosphatase [TRAP]) analysis. Implant removal torque was also measured. RESULTS: On tomography, maintenance of sinus membrane integrity was observed in both the groups. Higher values of morphometric parameters evaluated by micro-CT were found in the "paste group" after seven days. At 40 days, there were no significant differences between the groups in most of the microtomographic parameters evaluated. In histological sections stained with HE, a higher percentage of newly formed bone was observed in the "granule group" after 40 days. Similar positive immunolabeling was observed for both RUNX2 and OCN in both the experimental groups. TRAP immunolabeling was similar in both groups as well. VEGF labeling increased in the "granule group", indicating a higher osteoconductive potential in this biomaterial. Similar removal torque values were observed in both groups. Thus, the two HA + ß-TCP configurations showed similar healing patterns of simultaneously installed implants adjacent to sinus floor elevation. However, significantly higher bone values were observed for the "granule configuration". CONCLUSIONS: The HA + ß-TCP granules and paste presentations showed favorable long-term healing results, with bone formation in similar quantities and quality adjacent to the implants.

Determinants of COVID-19 vaccine hesitancy among pregnant persons.

BACKGROUND: Vaccines reduce the risk of contracting and developing complications from coronavirus disease 2019 (COVID-19). Pregnant people are at increased risk of disease-related complications but have a higher prevalence of vaccine hesitancy (VH) than their nonpregnant counterparts. OBJECTIVE: This study aimed to describe risk factors and COVID-19- and vaccine-related perspectives that lead to VH among pregnant people in Mexico to target strategies to increase vaccine acceptance in this population. METHODS: A cross-sectional survey-based study to evaluate risk factors and COVID-19- and vaccine-related perspectives associated with VH among pregnant people was conducted. Respondents were pregnant people of all ages attending a regular follow-up visit or admitted to labor and delivery in a third-level maternity hospital in Mexico. VH was defined as not having received a COVID-19 vaccine and either declining or being undecided to accept a vaccine during their pregnancy. We used bivariate and multivariable logistic regression models to estimate assess the relationship among demographic factors, COVID-19- and vaccine-related perspectives, and VH. RESULTS: A total of 1475 respondents completed the questionnaire; 216 (18%) were under the age of 18 years, and 860 (58%) had received at least one dose of a COVID-19 vaccine. In this sample, 264 (18%) were classified as vaccine hesitant. Key factors associated with VH were adolescence, having family as a primary source of information, first pregnancy, and history of vaccines in previous pregnancies. COVID-19 perspectives were also strongly associated with VH. CONCLUSIONS: Among pregnant people in Mexico, VH is associated with demographic factors, vaccination history, sources of information, and perceived risks to the fetus. This information is relevant to policy makers and health care professionals to identify those more likely to be hesitant and to inform strategies to increase vaccine uptake among pregnant people.

Environmental performance of phytosanitary control techniques on soybean crop estimated by life cycle assessment (LCA).

Phytosanitary control is essential to ensure agricultural productivity and quality. However, approaches based on scheduled use of pesticides, overuse of harmful molecules produce impacts on different types of living organisms. Integrated Pest and Disease Management (IPM-IDM) may significantly reduce the burden of pesticides in the environment. Plant resistance may also be included in the IPM-IDM and even in conventional management due to little requirement of additional knowledge and changes in agricultural practices. Robust environmental assessments using methodology of universal use, life cycle assessment (LCA), may estimate the impacts of specific pesticides that cause major damages, including remarkable category impacts. Therefore the objective of this study was to determine the impacts and (eco)toxicological effects of phytosanitary strategies (IPM-IDM including or not lepidopteran resistant transgenic cultivars) vs. the scheduled approach. Two inventory modeling methods were also applied to gather information on the use and applicability of these methods. Life cycle assessment (LCA) was applied using two inventory modeling methods: 100%Soil and PestLCI (Consensus) using data from Brazilian croplands under tropical conditions, by combining phytosanitary approaches (IPM-IDM, IPM-IDM + transgenic cultivar, conventional, conventional + transgenic cultivar) and modeling methods. Hence, eight soybean production scenarios were established. The IPM-IDM was efficient to reduce the (eco)toxicity impacts of soybean production mainly for freshwater ecotoxicity category. Due to the dynamic character of IPM-IDM approaches, the inclusion of recently introduced strategies (plant resistant and biological control to stink bugs and plant fungal diseases) may diminish even more the principal impacting substances throughout the Brazilian croplands. The PestLCI Consensus method, although its development is yet in progress, to date can be suggested to estimate the agriculture environmental impacts more properly under tropical conditions.

Urbanization promotes specific bacteria in freshwater microbiomes including potential pathogens.

Freshwater ecosystems are characterized by complex and highly dynamic microbial communities that are strongly structured by their local environment and biota. Accelerating urbanization and growing city populations detrimentally alter freshwater environments. To determine differences in freshwater microbial communities associated with urbanization, full-length 16S rRNA gene PacBio sequencing was performed in a case study from surface waters and sediments from a wastewater treatment plant, urban and rural lakes in the Berlin-Brandenburg region, Northeast Germany. Water samples exhibited highly habitat specific bacterial communities with multiple genera showing clear urban signatures. We identified potentially harmful bacterial groups associated with environmental parameters specific to urban habitats such as Alistipes, Escherichia/Shigella, Rickettsia and Streptococcus. We demonstrate that urbanization alters natural microbial communities in lakes and, via simultaneous warming and eutrophication and creates favourable conditions that promote specific bacterial genera including potential pathogens. Our findings are evidence to suggest an increased potential for long-term health risk in urbanized waterbodies, at a time of rapidly expanding global urbanization. The results highlight the urgency for undertaking mitigation measures such as targeted lake restoration projects and sustainable water management efforts.

Vasopressin as rescue therapy in pediatric catecholamine-refractory shock / Vasopressina como terapia de resgate em choque refratário à catecolamina em crianças

Introduction: There is scarce evidence on the efficacy of vasopressin as a vasopressor agent in pediatric catecholamine-refractory shock. The aim of this study is to describe hemodynamic changes during the first hours of administration vasopressin as rescue therapy in patients with catecholamine-refractory shock. Methods: This is a retrospective study including children from 1 month to 18 years admitted to the PICU who received vasopressin as rescue therapy for catecholamine-refractory shock (noradrenalin ≥1µg/ kg/min and variable doses of other inotropic / vasopressor agents). For analysis purposes, blood pressure means and modified vasoactive scores were calculated at two time periods: 2 hours prior to vasopressin therapy (T-2) and within the first 10 hours of vasopressin therapy (T10). Results: Using the paired Students' t test, mean blood pressure and vasoactive-inotropic scores modified by Wernovsky in the 2 hours before the use of vasopressin (T-2) were compared with the means of the first 10 hours of vasopressin administration (T10). The sample consisted of 16 patients. Median initial dose of vasopressin was 0.0005U/kg/min (interquartile range 0.00024-0.00168). Mean blood pressure and diastolic blood pressure increased with the use of vasopressin (p=0.0267 and p=0.0194, respectively). There was no reduction in vasoactive-inotropic score or increased diuresis. Conclusion: The administration of vasopressin increased blood pressure in this sample but did not promote a reduction in catecholamine scores nor increased diuresis.

Introdução: Há escassez de evidências sobre a eficácia da vasopressina como agente vasopressor no choque refratário à catecolamina em crianças. O objetivo foi descrever as alterações hemodinâmicas que ocorrem nas primeiras horas de administração da vasopressina, como terapia de resgate em pacientes com choque refratário à catecolamina. Métodos: Estudo retrospectivo. Foram incluídas crianças de 1 mês a 18 anos internadas na UTIP que receberam vasopressina como terapia de resgate para choque refratário à catecolamina (noradrenalina ≥1µg/kg/min e doses variáveis de outros inotrópicos /vasopressores). Para fins de análise, as medidas da pressão arterial e os escores vasoativos modificados foram calculados em dois períodos: nas 2 horas anteriores ao início da terapia com vasopressina (T-2) e nas primeiras 10 horas da terapia com vasopressina (T10). Resultados: Utilizando o teste t de Student pareado, a pressão arterial média e o escore vasoativo-inotrópico modificado por Wernovsky nas 2 horas anteriores ao uso de vasopressina (T-2) foram comparados com as médias dessas variáveis durante as primeiras 10 horas de administração de vasopressina (T10). A amostra foi composta por 16 pacientes. A dose mediana inicial de vasopressina foi de 0,0005U/kg/min (intervalo interquartil 0,00024-0,00168). A pressão arterial média e a pressão arterial diastólica aumentaram com o uso de vasopressina (p=0,0267 e p=0,0194, respectivamente). Não houve redução no escore vasoativo-inotrópico ou aumento na diurese. Conclusão: A administração de vasopressina aumentou a pressão arterial nesta amostra, mas não teve efeitos na redução dos escores de catecolaminas ou no aumento da diurese.

Effect of nutrition status and inflammatory stimuli on ghrelin and peptide-YY levels among critically ill children: A prospective and observational study.

BACKGROUND: Ghrelin and peptide-YY (PYY) are two gut peptides with apparent opposing actions. In normal conditions, ghrelin and PYY work together in synergy to regulate energy homeostasis. During critical illness, series of metabolic, endocrine, and inflammatory changes take place in response to a severe insult. Emerging studies recorded alterations in gut hormone levels in critically ill adults. This study aims to assess the effect of inflammation, nutrition, and feeding status on ghrelin and PYY levels in critically ill children. METHODS: In this prospective study, we collected blood samples from critically ill children on days 2 or 3 of pediatric intensive care unit (PICU) admission for the analysis of serum ghrelin, PYY, and inflammatory markers. Data related to the intake anthropometry, as well as other clinical data, were collected from patients' records. Multiple linear regression analysis was used to identify factors affecting serum levels of these hormones. RESULTS: Forty-two children admitted to the PICU were included in this study. Ghrelin level was influenced by admission nutrition status of the children and age. PYY was influenced by macronutrient intake and age. Inflammatory markers also showed an association with the measured levels of these hormones, with C-reactive protein being positively associated with ghrelin levels and tumor necrosis factor alpha showing a positive association with PYY levels. CONCLUSION: Although ghrelin and PYY have been linked to feeding status in healthy patients, during critical illness there might be other factors, such as inflammation and nutrition status, that might contribute to the changes observed in ghrelin/PYY profiles.

The effectiveness of hydroxyapatite-beta tricalcium phosphate incorporated into stem cells from human exfoliated deciduous teeth for reconstruction of rat calvarial bone defects.

OBJECTIVE: To investigate the effects of stem cells from the pulp of human exfoliated deciduous teeth (SHED) on biphasic calcium phosphate granules (BCP) to repair rat calvarial defects as compared to autogenous bone grafting. MATERIALS AND METHODS: A defect with a 6-mm diameter was produced on the calvaria of 50 rats. BCP granules were incorporated into SHED cultures grown for 7 days in conventional (CM) or osteogenic (OM) culture media. The animals were allocated into 5 groups of 10, namely: clot, autogenous bone, BCP, BCP+SHED in CM (BCP-CM), and BCP+SHED in OM (BCP-OM). The presence of newly formed bone and residual biomaterial particles was assessed by histometric analysis after 4 and 8 weeks. RESULTS: The autogenous group showed the largest newly formed bone area at week 8 and in the entire experimental period, with a significant difference in relation to the other groups (P < 0.05). At week 8, BCP-CM and BCP-OM groups showed homogeneous new bone formation (P = 0.13). When considering the entire experimental period, the BCP group had the highest percentage of residual particle area, with no significant difference from the BCP-CM group (P = 0.06) and with a significant difference from the BCP-OM group (P = 0.01). BCP-CM and BCP-OM groups were homogeneous throughout the experimental period (P = 0.59). CONCLUSIONS: BCP incorporated into SHED cultures showed promising outcomes, albeit less pronounced than autogenous grafting, for the repair of rat calvarial defects. CLINICAL RELEVANCE: BCP incorporated into SHED cultures showed to be an alternative in view of the disadvantages to obtain autogenous bone graft.

Results of Preimplantation Genetic Testing for Aneuploidy (PGT-A) in a Cohort of 319 Embryos: Experience in a Fertility Clinic in Colombia.

OBJECTIVE: To analyze the results of the preimplantation genetic testing for aneuploidy at the Instituto de Fertilidad Humana - Inser Bogotá, Colombia, from 2016 to 2020. METHODS: This study is an observational, retrospective, and correlative analysis of biopsies from 319 embryos (from 54 patients) submitted to preimplantation genetic testing for aneuploidy by different molecular techniques. RESULTS: Of the 54 patients included in the study, 42 provided their own oocytes, and 12 used donated oocytes. The main indication to perform the preimplantation genetic testing was advanced maternal age. We obtained 319 embryos: Ninety-one (28.5%) euploid, 197 (61.8%) aneuploid and 31 (9.7%) with no detectable DNA. The highest rate of aneuploid embryos was found in patients over 40 years (72.7%), and the euploidy rate in patients under 35 years was 37.1%. After the transfer of euploid embryos, the rates for implantation, ongoing pregnancy, live birth, and miscarriage were 40%, 50%, 40.6%, and 0%, respectively. Older maternal age correlated with higher numbers of aneuploid embryos and lower numbers of both euploid and 5-day embryos. CONCLUSIONS: There was a positive correlation between maternal age and aneuploidy rate. Complex chromosomal abnormalities were the most frequent aneuploidies, followed by mosaicism and double aneuploidies. The miscarriage rate after the transfer of euploid embryos was 0 %.

Automated ELISA on-chip for the detection of an-ti-SARS-CoV-2 antibodies

The COVID-19 pandemic has been the most critical public health issue in modern his-tory due to its highly infectious and deathly potential; and the limited access to massive, low-cost, and reliable testing has significantly worsened the crisis. The recovery and the vaccination of millions of people against COVID-19, have made serological tests highly relevant to identify the presence and levels of SARS-CoV-2 antibodies. Due to its advantages, microfluidic-based technologies represent an attractive alternative to the conventional testing methodologies used for these purposes. In this work, we describe the development of an automated ELISA on-chip capable of detecting anti-SARS-CoV-2 antibodies in serum samples from COVID-19 patients and vaccinated individuals. The colorimetric reactions were analyzed with a microplate reader. No statistically significant differences were observed when comparing the results of our automated ELISA on-chip against the ones obtained from a traditional ELISA on a microplate. Moreover, we demonstrated that it is possible to carry out the analysis of the colorimetric reaction by performing basic image analysis of photos taken with a smartphone, which constitutes a useful alternative when lacking specialized equipment or a laboratory setting. Our automated ELISA on-chip has the potential to be used in a clinical setting and mitigate some of the burden caused by testing deficiencies.

Accuracy of cerebrospinal fluid ferritin for purulent meningitis.

OBJECTIVE: To evaluate the use of cerebrospinal fluid (CSF) ferritin levels in the diagnosis of purulent meningitis (PM). METHOD: We studied 81 children between 28 days and 12 years of age who presented with clinical suspicion of meningitis to the emergency department. CSF ferritin levels were measured and compared between diagnostic groups (PM, aseptic meningitis (AM) and no meningitis). RESULTS: The median age was 24 (IQR 8-69) months. There were 32 patients with AM (39%), 23 with PM (28%) and 26 with no meningitis (32%). Median CSF ferritin was 4.2 ng/mL (IQR 3.0-6.5), 52.9 ng/mL (IQR 30.7-103 ng/mL) and 2.4 ng/mL (IQR 2-4), respectively. CSF ferritin was higher in children with PM compared with AM (p<0.001) or no meningitis (p<0.001). There was no difference between AM and no meningitis. CONCLUSION: CSF ferritin may be a useful biomarker to discriminate PM in children with clinical symptoms of this disease.

Statistical approach to Trichinella infection in horses handled by rural slaughterhouses across five distinctive socioeconomic regions in Mexico

Abstract This work aimed to investigate the prevalence of Trichinella infection in horses (Equus ferus caballus) handled by rural slaughterhouses across five distinctive socioeconomic regions in Mexico. Serum samples were obtained by non-probabilistic convenience sampling in the Eastern, Southern Central and Western regions (100 samples of each). Additionally, muscle tissue samples were collected from the East (n = 45), Southeastern (n = 88), Southern Central (n = 39) and Southwestern (n = 11) regions. Antibodies were determined by Western blot and the muscle tissue was examined by artificial digestion. A global antibody prevalence of 2% was obtained. Regionally, a prevalence of 5% was observed in the East and 1% in the Southern Central region. No antibodies were detected in the West region and no larvae were found in the muscle tissue samples. These findings support the low presence of Trichinella in Mexican horses, which can positively impact the Mexican horse meat trade.

Resumen Se investigó la prevalencia de Trichinella en caballos (Equus ferus caballus) demataderos rurales en cinco regiones socioeconómicas de México. Los sueros se obtuvieron por Trichinella muestreo de conveniencia no probabilístico en las regiones Este, Centrosur y Oeste (100 muestras de cada una). Además se colectaron muestras de tejido muscular de animales de las regionesEste (n = 45), Centro Sur (n = 39), Sudeste (n = 88) y Sudoeste (n = 11). Los anticuerpos se determinaron por Western blot y el tejido muscular se examinó por digestión artificial. Se obtuvo una prevalencia global de anticuerpos del 2%. A nivel regional se observó una prevalencia del 5% enel Este y del 1% en el Centrosur. No se detectaron anticuerpos en el Oeste y no se encontraron larvas en el tejido muscular. Los datos respaldan la baja presencia de Trichinella en caballos mexicanos, lo que puede impactar positivamente en su comercio.

SARS-CoV-2 protein Nsp1 alters actomyosin cytoskeleton and phenocopies arrhythmogenic cardiomyopathy-related PKP2 mutant

Mutations in desmosomal Plakophilin-2 (PKP2) are the most prevalent drivers of arrhythmogenic cardiomyopathy (ACM) and a common cause of sudden cardiac death in young athletes. However, partner proteins that elucidate PKP2 cellular mechanism to understand cardiac dysfunction in ACM are mostly unknown. Here we identify the actin-based motor proteins Myh9 and Myh10 as key PKP2 interactors, and demonstrate that the expression of the ACM-related PKP2 mutant R735X alters actin fiber organization and cell mechanical stiffness. We also show that SARS-CoV-2 Nsp1 protein acts similarly to this known pathogenic R735X mutant, altering the actomyosin component distribution on cardiac cells. Our data reveal that the viral Nsp1 hijacks PKP2 into the cytoplasm and mimics the effect of delocalized R735X mutant. These results demonstrate that cytoplasmic PKP2, wildtype or mutant, induces the collapse of the actomyosin network, since shRNA-PKP2 knockdown maintains the cell structure, validating a critical role of PKP2 localization in the regulation of actomyosin architecture. The fact that Nsp1 and PKP2 mutant R735X share similar phenotypes also suggests that direct SARS-CoV-2 heart infection could induce a transient ACM-like disease in COVID-19 patients, which may contribute to right ventricle dysfunction, observed in patients with poor survival prognosis. HighlightsThe specific cardiac isoform Plakophilin-2a (PKP2) interacts with Myh9 and Myh10. PKP2 delocalization alters actomyosin cytoskeleton component organization. SARS-CoV-2 Nsp1 protein hijacks PKP2 from the desmosome into the soluble fraction where it is downregulated. Viral Nsp1 collapses the actomyosin cytoskeleton and phenocopies the arrhythmogenic cardiomyopathy-related mutant R735X.

A randomized controlled study comparing guided bone regeneration with connective tissue graft to reestablish buccal convexity at implant sites: A 1-year volumetric analysis.

OBJECTIVES: To volumetrically compare guided bone regeneration (GBR) with connective tissue graft (CTG) to reestablish convexity at the buccal aspect of single implants. MATERIALS AND METHODS: Patients with a single tooth gap in the anterior maxilla and horizontal alveolar defect were enrolled in a single-blind randomized clinical trial (RCT). All sites had a buccopalatal bone dimension of at least 6 mm, received a single implant, and were randomly allocated to the control (GBR) or test group (CTG) to reestablish buccal soft tissue convexity. Patients received a provisional crown at 3 months and a permanent crown at 6 months. Primary outcomes were volumetric increase (mm3 ) and linear increase (mm) in buccal soft tissue profile (BSP) within a well-defined area of interest at fixed time points. Alveolar process deficiency was a secondary outcome. RESULTS: Twenty-one patients were included per group (control: 11 females, mean age 51; test: 9 females, mean age 48). After 1 year, GBR resulted in a significant volumetric increase of 20.74 mm3 (P < .001) corresponding to linear increase in BSP of 1.30 mm (P < .001). For CTG, this was 15.86 (P < .001) and 1.19 mm (P < .001), respectively. The changes over time in volume (P = .173) and BSP (P = .241) were not significantly different between the groups. Twenty-nine percentage and 26% of the final volumetric increase was the result of installing and altering prosthetic components in the control and test groups, respectively. Alveolar process deficiency significantly reduced from pre-op to 1 year following GBR (P < .001) and CTG (P < .001). The difference between the groups was not significant (P = .342). However, 58% of the patients treated with GBR and 38% treated with CTG failed to show perfect soft tissue convexity at the buccal aspect. CONCLUSION: GBR as well as CTG are effective in reducing horizontal alveolar defects for aesthetic purposes. However, in about half of the cases, either strategy failed to optimally reestablish buccal convexity.

Should we wait 3 months for a repeat aspiration in non-diagnostic/indeterminate thyroid nodules? A cancer centre experience.

INTRODUCTION: The Bethesda System recommends repeat fine needle aspiration (rFNA) as a management option for nodules classified under the non-diagnostic (ND) and atypia of undetermined significance (AUS/FLUS) categories. We evaluated the impact of an rFNA in diagnostic resolution and the role of early (≤3 months) vs delayed (more than 3 months) rFNA of nodules initially diagnosed as ND and AUS/FLUS. METHODS: We retrospectively collected all thyroid FNA performed in a 4-year period with repeat aspiration. For cases initially signed out as ND or AUS/FLUS, diagnostic resolution was defined as a change to a Bethesda System category other than these two on rFNA. Comparison and regression models were fitted to identify the impact of time of rFNA on diagnostic resolution. RESULTS: In total, 184 cases were initially assigned as ND and 143 as AUS/FLUS, with overall diagnostic resolution rates for the reassessment of these nodules calculated at 70.1% and 62.9%, respectively. For ND cases, time of rFNA was not significantly associated with diagnostic resolution (P > .05). For AUS/FLUS nodules, however, repeat aspiration performed in more than 3 months after the initial diagnosis was 2.5 times more likely to achieve a resolution in diagnosis than early rFNA (P = .024). CONCLUSIONS: Repeat aspiration of ND and AUS/FLUS nodules helped define diagnosis for the majority of cases, being highly effective in determining correct patient management. For AUS/FLUS nodules, repeat aspiration performed more than 3 months after the initial diagnosis was associated with a higher diagnostic resolution.

Methodological Considerations for Using Fecal Glucocorticoid Metabolite Concentrations as an Indicator of Physiological Stress in the Brown Bear (Ursus arctos).

Reliable methods to measure stress-related glucocorticoid responses in free-ranging animals are important for wildlife management and conservation. Such methods are also paramount for our ability to improve our knowledge of the ecological consequences of physiological processes. The brown bear (Ursus arctos) is a large carnivore of ecological and cultural importance and is important for management. Here, we provide a physiological validation for an enzyme immunoassay (EIA) to quantify glucocorticoid metabolites in brown bear feces. We also provide an evaluation of the effects of sample exposure to ambient temperature on measured fecal glucocorticoid metabolite (fGCM) concentrations. We evaluated three EIA systems: a cortisol assay, an 11-oxoetiocholanolone assay, and an 11ß-hydroxyetiocholanolone assay. Of these, the cortisol assay provided the best discrimination between peak fGCM concentrations detected 1-4 d after injections of synthetic adrenocorticotrophic hormone and preinjection baseline concentrations in four individual brown bears. The time of exposure to ambient temperature had substantial but variable effects on measured fGCM concentrations, including variation both between samples from the same individual and among samples from different bears. We propose that the validated EIA system for measuring fGCM concentrations in the brown bear could be a useful noninvasive method to monitor stress in this species. However, we highlight that this method requires that fecal samples be frozen immediately after defecation, which could be a limitation in many field situations.