Comparative proteomics of a versatile, marine, iron-oxidizing chemolithoautotroph.

This study conducted a comparative proteomic analysis to identify potential genetic markers for the biological function of chemolithoautotrophic iron oxidation in the marine bacterium Ghiorsea bivora. To date, this is the only characterized species in the class Zetaproteobacteria that is not an obligate iron-oxidizer, providing a unique opportunity to investigate differential protein expression to identify key genes involved in iron-oxidation at circumneutral pH. Over 1000 proteins were identified under both iron- and hydrogen-oxidizing conditions, with differentially expressed proteins found in both treatments. Notably, a gene cluster upregulated during iron oxidation was identified. This cluster contains genes encoding for cytochromes that share sequence similarity with the known iron-oxidase, Cyc2. Interestingly, these cytochromes, conserved in both Bacteria and Archaea, do not exhibit the typical ß-barrel structure of Cyc2. This cluster potentially encodes a biological nanowire-like transmembrane complex containing multiple redox proteins spanning the inner membrane, periplasm, outer membrane, and extracellular space. The upregulation of key genes associated with this complex during iron-oxidizing conditions was confirmed by quantitative reverse transcription-PCR. These findings were further supported by electromicrobiological methods, which demonstrated negative current production by G. bivora in a three-electrode system poised at a cathodic potential. This research provides significant insights into the biological function of chemolithoautotrophic iron oxidation.

Split fertilizer nitrogen application with a cereal rye cover crop reduces tile nitrate loads in a corn-soybean rotation.

Splitting fertilizer nitrogen (N) applications and using cover crops are management strategies to reduce nitrate in tile drainage water. We investigated split fertilizer N applications to corn (Zea mays L.) on crop yields and tile nitrate loss in both corn and soybean (Glycine max L.) in rotation from 2016 through 2019. We evaluated the inclusion of cover crops in a split-N treatment. Fertilizer N treatments included 100% in the fall; 50% in the fall + 25% at planting + 25% at side-dress; 100% as spring preplant; 75% as spring preplant (reduced N rate); 50% as spring preplant + 50% at side-dress; and 50% as spring preplant + 50% at side-dress with a cover crop. We did not find significant differences between split and single full rate N application treatments for corn yields or tile nitrate loss; however, the reduced N rate treatment significantly decreased corn yield by 10%. Cumulative tile nitrate losses (over four seasons) ranged from 115 kg ha-1 for all of the N in the fall to 65 kg ha-1 for 50% as spring preplant + 50% at side-dress with a cover crop, a decrease of 43%. Tile nitrate loss responded similarly to (corn) N treatments under both corn and soybean, with 64% of the loss under corn and 36% under soybean. Our results suggest that decreasing the fertilizer N rate may impact corn yield more than nitrate loss, while split fertilizer N application with a cover crop has potential to reduce tile nitrate loss without decreasing crop yield.

State of maternal–fetal medicine in the Philippines / Philippine Journal of Obstetrics and Gynecology

@#The practice of maternal–fetal medicine (MFM) in the Philippines originated within the broader scope of obstetrics and gynecology. The specialty started to emerge to address the need for specialized care for high‑risk pregnancies and the management of fetal complications. Over the years, the development of MFM as a distinct subspecialty has evolved through advancements in medical technology, training programs, and the increasing complexity of pregnancy management. The Philippine Society of MFM (PSMFM), established in 1996, is the primary organization representing MFM specialists in the country. It is dedicated to advancing the field through education, training, service, and research. PSMFM provides its members a platform to enhance their knowledge, improve their skills, foster collaboration, and share expertise.

E-cigarettes and arrhythmogenesis: a comprehensive review of pre-clinical studies and their clinical implications.

Electronic cigarette use has grown exponentially in recent years, and while their popularity has increased, the long-term effects on the heart are yet to be fully studied and understood. Originally designed as devices to assist with those trying to quit traditional combustible cigarette use, their popularity has attracted use by teens and adolescents who traditionally have not smoked combustible cigarettes. Acute effects on the heart have been shown to be similar to traditional combustible cigarettes, including increased heart rate and blood pressure. The main components of electronic cigarettes that contribute to these arrhythmic effects are found in the e-liquid that is aerosolized and inhaled, comprised of nicotine, flavourings, and a combination of vegetable glycerin (VG) and propylene glycol (PG). Nicotine can potentially induce both ventricular and atrial arrhythmogenesis, with both the atrial and ventricular effects resulting from the interactions of nicotine and the catecholamines they release via potassium channels. Atrial arrhythmogenesis, more specifically atrial fibrillation, can also occur due to structural alterations, which happens because of nicotine downregulating microRNAs 133 and 590, both post-transcriptional growth factor repressors. Liquid flavourings and the combination of PG and VG can possibly lead to arrhythmic events by exposing users to acrolein, an aldehyde that stimulates TRPA1 that in turn causes a change towards sympathetic activation and autonomic imbalance. The design of these electronic delivery devices is constantly changing; therefore, it has proven extremely difficult to study the long-term effects on the heart caused by electronic cigarettes but will be important to understand given their rising popularity. The arrhythmic effects of electronic cigarettes appear similar to traditional cigarettes as well; however, a comprehensive review has not been compiled and is the focus of this article.

The past, present, and future of immunotherapy for endometrial adenocarcinoma.

Incidences of endometrial adenocarcinoma are increasing in the USA with poor prognosis for patients with advanced disease. The current treatment standard is surgery including total hysterectomy and bilateral oophorectomy with surgical staging and adjunct treatment, such as chemotherapy or radiation. However, these methods do not present as an effective treatment option for poorly differentiated advanced cancers. Advancements in immunotherapy now offer a new approach for various types of cancer and specifically show promise in the treatment of endometrial adenocarcinoma. This review summarizes immunotherapeutic treatment options relevant to endometrial adenocarcinoma, such as immune checkpoint blockades, bispecific T-cell engager antibodies, vaccinations, and adoptive cell transfer. This study could be helpful for clinicians to identify treatment options more suitable for women with late-stage endometrial adenocarcinoma.

All-Optical GeV Electron Bunch Generation in a Laser-Plasma Accelerator via Truncated-Channel Injection.

We describe a simple scheme, truncated-channel injection, to inject electrons directly into the wakefield driven by a high-intensity laser pulse guided in an all-optical plasma channel. We use this approach to generate dark-current-free 1.2 GeV, 4.5% relative energy spread electron bunches with 120 TW laser pulses guided in a 110 mm-long hydrodynamic optical-field-ionized plasma channel. Our experiments and particle-in-cell simulations show that high-quality electron bunches were only obtained when the drive pulse was closely aligned with the channel axis, and was focused close to the density down ramp formed at the channel entrance. Start-to-end simulations of the channel formation, and electron injection and acceleration show that increasing the channel length to 410 mm would yield 3.65 GeV bunches, with a slice energy spread ∼5×10^{-4}.

The past, present, and future of immunotherapy for bladder tumors.

Bladder cancer is a prominent cancer worldwide with a relatively low survival rate for patients with increased stage and metastasis. Current treatments are based on surgical removal, bacillus Calmette-Guerin (BCG) Immunotherapy, and platinum-based chemotherapy. However, treatment resistance due to genetic instability of bladder tumors, as well as intolerance to treatment adverse effects leads to the necessity to further treatment options. New advancements in immunotherapy are on the rise for treatment of various cancers and specifically has shown promise in the treatment of bladder cancer. This review summarizes these new advancements in treatment options involving cytokines and cytokine blockade. Such a study might be helpful for urologists to manage patients with bladder cancer more effectively.

Maize Leaf Appearance Rates: A Synthesis From the United States Corn Belt.

The relationship between collared leaf number and growing degree days (GDD) is crucial for predicting maize phenology. Biophysical crop models convert GDD accumulation to leaf numbers by using a constant parameter termed phyllochron (°C-day leaf-1) or leaf appearance rate (LAR; leaf oC-day-1). However, such important parameter values are rarely estimated for modern maize hybrids. To fill this gap, we sourced and analyzed experimental datasets from the United States Corn Belt with the objective to (i) determine phyllochron values for two types of models: linear (1-parameter) and bilinear (3-parameters; phase I and II phyllochron, and transition point) and (ii) explore whether environmental factors such as photoperiod and radiation, and physiological variables such as plant growth rate can explain variability in phyllochron and improve predictability of maize phenology. The datasets included different locations (latitudes between 48° N and 41° N), years (2009-2019), hybrids, and management settings. Results indicated that the bilinear model represented the leaf number vs. GDD relationship more accurately than the linear model (R 2 = 0.99 vs. 0.95, n = 4,694). Across datasets, first phase phyllochron, transition leaf number, and second phase phyllochron averaged 57.9 ± 7.5°C-day, 9.8 ± 1.2 leaves, and 30.9 ± 5.7°C-day, respectively. Correlation analysis revealed that radiation from the V3 to the V9 developmental stages had a positive relationship with phyllochron (r = 0.69), while photoperiod was positively related to days to flowering or total leaf number (r = 0.89). Additionally, a positive nonlinear relationship between maize LAR and plant growth rate was found. Present findings provide important parameter values for calibration and optimization of maize crop models in the United States Corn Belt, as well as new insights to enhance mechanisms in crop models.

A Genus Definition for Bacteria and Archaea Based on a Standard Genome Relatedness Index.

Genus assignment is fundamental in the characterization of microbes, yet there is currently no unambiguous way to demarcate genera solely using standard genomic relatedness indices. Here, we propose an approach to demarcate genera that relies on the combined use of the average nucleotide identity, genome alignment fraction, and the distinction between type- and non-type species. More than 3,500 genomes representing type strains of species from >850 genera of either bacterial or archaeal lineages were tested. Over 140 genera were analyzed in detail within the taxonomic context of order/family. Significant genomic differences between members of a genus and type species of other genera in the same order/family were conserved in 94% of the cases. Nearly 90% (92% if polyphyletic genera are excluded) of the type strains were classified in agreement with current taxonomy. The 448 type strains that need reclassification directly impact 33% of the genera analyzed in detail. The results provide a first line of evidence that the combination of genomic indices provides added resolution to effectively demarcate genera within the taxonomic framework that is currently based on the 16S rRNA gene. We also identify the emergence of natural breakpoints at the genome level that can further help in the circumscription of taxa, increasing the proportion of directly impacted genera to at least 43% and pointing at inaccuracies on the use of the 16S rRNA gene as a taxonomic marker, despite its precision. Altogether, these results suggest that genomic coherence is an emergent property of genera in Bacteria and ArchaeaIMPORTANCE In recent decades, the taxonomy of Bacteria and Archaea, and therefore genus designation, has been largely based on the use of a single ribosomal gene, the 16S rRNA gene, as a taxonomic marker. We propose an approach to delineate genera that excludes the direct use of the 16S rRNA gene and focuses on a standard genome relatedness index, the average nucleotide identity. Our findings are of importance to the microbiology community because the emergent properties of Bacteria and Archaea that are identified in this study will help assign genera with higher taxonomic resolution.

Simulating nitrogen management impacts on maize production in the U.S. Midwest.

Nutrient loss reduction strategies have recently been developed in the U.S. Midwest to decrease the environmental footprint associated with nitrogen (N) fertilizer use. Although these strategies generally suggest decreasing N rates and shifting the timing of N application from fall to spring, the spatiotemporal impacts of these practices on maize yield and fertilizer N use efficiency (NUE, kg grain yield increase per kg N applied) have not been assessed at the watershed scale using crop simulation models. We simulated the effects of N fertilizer rate (0, 168, 190, 224 kg N ha-1) and application timing [fall-applied N (FN): 100% N applied on 1 December; spring-applied N (SN): 100% N applied 10 days before planting; split N: 66% N applied on 1 December + 34% N applied 10 days before planting] on maize grain yield (GY) across 3042 points in Illinois during 2011-2015 using the DSSAT-CERES-Maize model. When simulations were scaled up to the watershed level, results suggest that increases in average maize GY for SN compared to FN occurred in years with higher than average winter rainfall (2011, 2013), whereas yields were similar (+/- 4%) in 2012, 2014, and 2015. Accordingly, differences in NUE for SN compared to FN were small (0.0-1.4 kg GY/kg N) when cumulative winter rainfall was < 300 mm, but increased to 0.1-9.2 kg GY/kg N when winter rainfall was > 500 mm at both 168 kg N ha-1 and 224 kg N ha-1. The combined practice of reducing N fertilizer amounts from 224 kg N ha-1 to 190 kg N ha-1 and shifting from FN to SN resulted in a wide range of yield responses during 2011-2015, with the probability of increasing yields varying from <10% to >70% of simulation points within a watershed. Positive impacts on both GY and NUE occurred in only 60% of simulations for this scenario, highlighting the challenge of simultaneously improving yield and NUE with a 15% N rate reduction in this region.

Insights into the Fundamental Physiology of the Uncultured Fe-Oxidizing Bacterium Leptothrix ochracea.

Leptothrix ochracea is known for producing large volumes of iron oxyhydroxide sheaths that alter wetland biogeochemistry. For over a century, these delicate structures have fascinated microbiologists and geoscientists. Because L. ochracea still resists long-term in vitro culture, the debate regarding its metabolic classification dates back to 1885. We developed a novel culturing technique for L. ochracea using in situ natural waters and coupled this with single-cell genomics and nanoscale secondary-ion mass spectrophotometry (nanoSIMS) to probe L. ochracea's physiology. In microslide cultures L. ochracea doubled every 5.7 h and had an absolute growth requirement for ferrous iron, the genomic capacity for iron oxidation, and a branched electron transport chain with cytochromes putatively involved in lithotrophic iron oxidation. Additionally, its genome encoded several electron transport chain proteins, including a molybdopterin alternative complex III (ACIII), a cytochrome bd oxidase reductase, and several terminal oxidase genes. L. ochracea contained two key autotrophic proteins in the Calvin-Benson-Bassham cycle, a form II ribulose bisphosphate carboxylase, and a phosphoribulose kinase. L. ochracea also assimilated bicarbonate, although calculations suggest that bicarbonate assimilation is a small fraction of its total carbon assimilation. Finally, L. ochracea's fundamental physiology is a hybrid of those of the chemolithotrophic Gallionella-type iron-oxidizing bacteria and the sheathed, heterotrophic filamentous metal-oxidizing bacteria of the Leptothrix-Sphaerotilus genera. This allows L. ochracea to inhabit a unique niche within the neutrophilic iron seeps.IMPORTANCELeptothrix ochracea was one of three groups of organisms that Sergei Winogradsky used in the 1880s to develop his hypothesis on chemolithotrophy. L. ochracea continues to resist cultivation and appears to have an absolute requirement for organic-rich waters, suggesting that its true physiology remains unknown. Further, L. ochracea is an ecological engineer; a few L. ochracea cells can generate prodigious volumes of iron oxyhydroxides, changing the ecosystem's geochemistry and ecology. Therefore, to determine L. ochracea's basic physiology, we employed new single-cell techniques to demonstrate that L. ochracea oxidizes iron to generate energy and, despite having predicted genes for autotrophic growth, assimilates a fraction of the total CO2 that autotrophs do. Although not a true chemolithoautotroph, L. ochracea's physiological strategy allows it to be flexible and to extensively colonize iron-rich wetlands.

Quantitative analysis of O2 and Fe2+ profiles in gradient tubes for cultivation of microaerophilic Iron(II)-oxidizing bacteria.

The classical approach for the cultivation of neutrophilic microaerophilic Fe(II)-oxidizing bacteria is agar-based gradient tubes where these bacteria find optimal growth conditions in opposing gradients of oxygen (O2) and dissolved Fe(II) (Fe2+). The goals of this study were to quantify the temporal development of O2 and Fe2+ concentrations over time, to compare abiotic and microbially inoculated tubes and to test the suitability of different Fe(II)-sources for the cultivation of freshwater and marine microaerophilic Fe(II)-oxidizers. O2 and Fe2+ gradients were monitored on a high spatial resolution as a function of time applying amperometric and voltammetric microsensors. Fe(II)-oxidizers could be cultivated well with FeS and zero-valent iron powder as Fe(II)-source, but FeCO3 and FeCl2 are extremely sensitive for this application. Fe(III) minerals accumulated in inoculated tubes within the first days in regions with an O2 concentration of 20-40 µM and were confirmed to be related to bacterial growth. Microbial Fe(II) oxidation could compete only for the first days with the abiotic reaction after which heterogeneous Fe(II) oxidation, catalyzed by Fe(III) minerals, dominated. Our results imply that transfer of cultures to fresh tubes within 48-72 h is crucial to provide optimal growth conditions for microaerophilic Fe(II)-oxidizers, particularly for the isolation of new strains.

Tile Drainage Nitrate Losses and Corn Yield Response to Fall and Spring Nitrogen Management.

Nitrogen (N) management strategies that maintain high crop productivity with reduced water quality impacts are needed for tile-drained landscapes of the US Midwest. The objectives of this study were to determine the effect of N application rate, timing, and fall nitrapyrin addition on tile drainage nitrate losses, corn ( L.) yield, N recovery efficiency, and postharvest soil nitrate content over 3 yr in a corn-soybean [ (L.) Merr.] rotation. In addition to an unfertilized control, the following eight N treatments were applied as anhydrous ammonia in a replicated, field-scale experiment with both corn and soybean phases present each year in Illinois: fall and spring applications of 78, 156, and 234 kg N ha, fall application of 156 kg N ha + nitrapyrin, and sidedress (V5-V6) application of 156 kg N ha. Across the 3-yr study period, increases in flow-weighted NO concentrations were found with increasing N rate for fall and spring N applications, whereas N load results were variable. At the same N rate, spring vs. fall N applications reduced flow-weighted NO concentrations only in the corn-soybean-corn rotation. Fall nitrapyrin and sidedress N treatments did not decrease flo8w-weighted NO concentrations in either rotation compared with fall and spring N applications, respectively, or increase corn yield, crop N uptake, or N recovery efficiency in any year. This study indicates that compared with fall N application, spring and sidedress N applications (for corn-soybean-corn) and sidedress N applications (for soybean-corn-soybean) reduced 3-yr mean flow-weighted NO concentrations while maintaining yields.

A Vision for Incorporating Environmental Effects into Nitrogen Management Decision Support Tools for U.S. Maize Production.

Meeting crop nitrogen (N) demand while minimizing N losses to the environment has proven difficult despite significant field research and modeling efforts. To improve N management, several real-time N management tools have been developed with a primary focus on enhancing crop production. However, no coordinated effort exists to simultaneously address sustainability concerns related to N losses at field- and regional-scales. In this perspective, we highlight the opportunity for incorporating environmental effects into N management decision support tools for United States maize production systems by integrating publicly available crop models with grower-entered management information and gridded soil and climate data in a geospatial framework specifically designed to quantify environmental and crop production tradeoffs. To facilitate advances in this area, we assess the capability of existing crop models to provide in-season N recommendations while estimating N leaching and nitrous oxide emissions, discuss several considerations for initial framework development, and highlight important challenges related to improving the accuracy of crop model predictions. Such a framework would benefit the development of regional sustainable intensification strategies by enabling the identification of N loss hotspots which could be used to implement spatially explicit mitigation efforts in relation to current environmental quality goals and real-time weather conditions. Nevertheless, we argue that this long-term vision can only be realized by leveraging a variety of existing research efforts to overcome challenges related to improving model structure, accessing field data to enhance model performance, and addressing the numerous social difficulties in delivery and adoption of such tool by stakeholders.

Insulin signaling misregulation underlies circadian and cognitive deficits in a Drosophila fragile X model.

Fragile X syndrome (FXS) is an undertreated neurodevelopmental disorder characterized by low intelligence quotent and a wide range of other symptoms including disordered sleep and autism. Although FXS is the most prevalent inherited cause of intellectual disability, its mechanistic underpinnings are not well understood. Using Drosophila as a model of FXS, we showed that select expression of dfmr1 in the insulin-producing cells (IPCs) of the brain was sufficient to restore normal circadian behavior and to rescue the memory deficits in the fragile X mutant fly. Examination of the insulin signaling (IS) pathway revealed elevated levels of Drosophila insulin-like peptide 2 (Dilp2) in the IPCs and elevated IS in the dfmr1 mutant brain. Consistent with a causal role for elevated IS in dfmr1 mutant phenotypes, the expression of dfmr1 specifically in the IPCs reduced IS, and genetic reduction of the insulin pathway also led to amelioration of circadian and memory defects. Furthermore, we showed that treatment with the FDA-approved drug metformin also rescued memory. Finally, we showed that reduction of IS is required at different time points to rescue circadian behavior and memory. Our results indicate that insulin misregulation underlies the circadian and cognitive phenotypes displayed by the Drosophila fragile X model, and thus reveal a metabolic pathway that can be targeted by new and already approved drugs to treat fragile X patients.

The role of microaerophilic Fe-oxidizing micro-organisms in producing banded iron formations.

Despite the historical and economic significance of banded iron formations (BIFs), we have yet to resolve the formation mechanisms. On modern Earth, neutrophilic microaerophilic Fe-oxidizing micro-organisms (FeOM) produce copious amounts of Fe oxyhydroxides, leading us to wonder whether similar organisms played a role in producing BIFs. To evaluate this, we review the current knowledge of modern microaerophilic FeOM in the context of BIF paleoenvironmental studies. In modern environments wherever Fe(II) and O2 co-exist, microaerophilic FeOM proliferate. These organisms grow in a variety of environments, including the marine water column redoxcline, which is where BIF precursor minerals likely formed. FeOM can grow across a range of O2 concentrations, measured as low as 2 µm to date, although lower concentrations have not been tested. While some extant FeOM can tolerate high O2 concentrations, many FeOM appear to prefer and thrive at low O2 concentrations (~3-25 µm). These are similar to the estimated dissolved O2 concentrations in the few hundred million years prior to the 'Great Oxidation Event' (GOE). We compare biotic and abiotic Fe oxidation kinetics in the presence of varying levels of O2 and show that microaerophilic FeOM contribute substantially to Fe oxidation, at rates fast enough to account for BIF deposition. Based on this synthesis, we propose that microaerophilic FeOM were capable of playing a significant role in depositing the largest, most well-known BIFs associated with the GOE, as well as afterward when global O2 levels increased.

Climate-induced reduction in US-wide soybean yields underpinned by region- and in-season-specific responses.

The United States is one of the largest soybean exporters in the world. Production is concentrated in the upper Midwest(1). Much of this region is not irrigated, rendering soybean production systems in the area highly sensitive to in-season variations in weather. Although the influence of in-season weather trends on the yields of crops such as soybean, wheat and maize has been explored in several countries(2-6), the potentially confounding influence of genetic improvements on yields has been overlooked. Here we assess the effect of in-season weather trends on soybean yields in the United States between 1994 and 2013, using field trial data, meteorological data and information on crop management practices, including the adoption of new cultivars. We show that in-season temperature trends had a greater impact on soybean yields than in-season precipitation trends over the measurement period. Averaging across the United States, we show that soybean yields fell by around 2.4% for every 1 °C rise in growing season temperature. However, the response varied significantly among individual states, ranging from -22% to +9%, and also with the month of the year in which the warming occurred. We estimate that year-to-year changes in precipitation and temperature combined suppressed the US average yield gain by around 30% over the measurement period, leading to a loss of US$11 billion. Our data highlight the importance of developing location-specific adaptation strategies for climate change based on early-, mid- and late-growing season climate trends.

Metodologia da análise seminal para pacientes azoospérmicos no laboratório fleury / Methodology of seminal analysis for azoospermicos patients in the laboratório fleury

A azoospermia é definida como a ausência de espermatozoide no líquido seminal ejaculado pelo homem depois de aplicada a técnica de centrifugação em pelo menos duas amostras. Dada a importância de um diagnóstico correto da análise seminal para os casais, toda amostra que não apresentar espermatozoides no exame a fresco deve seguir em avaliação laboratorial. Com isso, o presente estudo tem como objetivo analisar os resultados de centrifugação de uma alíquota do sêmen ejaculado ou de todo o volume ejaculado de pacientes com diagnóstico de azoospermia para determinar qual o melhor método a ser empregado na análise seminal para esse grupo de pacientes.

The azoospermia is defined as the absence of sperm in the ejaculate by the seminal fluid man after centrifugation technique conducted in at least two samples. Given the importance of a correct diagnosis of the seminal analysis for couples, all sample no sperm present in fresh examination should follow in laboratory tests. Thus the present study aims to analyze the results of a spin rate of ejaculate or all of the ejaculate volume of patients with azoospermia to determine the best method to be used in semen analysis for this group of patients.

Trends in use in a Canadian pediatric emergency department.

INTRODUCTION: Emergency department (ED) crowding is a significant problem in Canada and has been associated with decreased quality of care in general and pediatric emergency departments (PEDs). Although boarding of admitted patients in the ED is the main contributor to adult ED overcrowding, factors involved in PED crowding may be different. The objective of this study was to report the trend in PED services use and to document the degree of overcrowding experienced in a Canadian PED. METHODS: A retrospective cohort study was conducted using administrative data from a tertiary care PED from 2002 to 2011. The primary outcome was PED use (total volume of visits and case severity per triage levels using the Canadian Triage and Acuity Scale [CTAS] score and admissions). Secondary outcomes included measures of PED overcrowding, such as rates of patients leaving without being seen (LWBS) and length of stay (LOS). RESULTS: Total volumes increased by 30% over the 10-year study period, whereas hospitalizations remained stable at approximately 10%. Trends in CTAS levels did not indicate meaningful changes in the severity of cases treated at our PED. LWBS proportions among CTAS 3, CTAS 4, and CTAS 5 groups and LOS for all CTAS groups progressively and statistically increased from year to year. CONCLUSIONS: Over the course of the study period, there was a substantial increase in PED visits, which likely contributed to the worsening markers of PED flow outcomes. Further study into the effects of PED crowding on patient outcomes is warranted.