The gut microbiome is impacted by certain types of dietary fibre. However, the type, duration and dose needed to elicit gut microbial changes and whether these changes also influence microbial metabolites remain unclear. This study investigated the effects of supplementing healthy participants with two types of non-digestible carbohydrates (resistant starch (RS) and polydextrose (PD)) on the stool microbiota and microbial metabolite concentrations in plasma, stool and urine, as secondary outcomes in the Dietary Intervention Stem Cells and Colorectal Cancer (DISC) Study. The DISC study was a double-blind, randomised controlled trial that supplemented healthy participants with RS and/or PD or placebo for 50 d in a 2 × 2 factorial design. DNA was extracted from stool samples collected pre- and post-intervention, and V4 16S rRNA gene sequencing was used to profile the gut microbiota. Metabolite concentrations were measured in stool, plasma and urine by high-performance liquid chromatography. A total of fifty-eight participants with paired samples available were included. After 50 d, no effects of RS or PD were detected on composition of the gut microbiota diversity (alpha- and beta-diversity), on genus relative abundance or on metabolite concentrations. However, Drichlet’s multinomial mixture clustering-based approach suggests that some participants changed microbial enterotype post-intervention. The gut microbiota and fecal, plasma and urinary microbial metabolites were stable in response to a 50-d fibre intervention in middle-aged adults. Larger and longer studies, including those which explore the effects of specific fibre sub-types, may be required to determine the relationships between fibre intake, the gut microbiome and host health.

OBJECTIVES/GOALS: Hypertension is a major risk factor for coronavirus disease 2019 (COVID-19) severity. Our goal was to determine if hypertension worsens lung pathology induced by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection in hamsters. METHODS/STUDY POPULATION: Male hamsters (7-8 weeks old) were infused with angiotensin II (AII; 200 ng/kg/min via osmotic minipump) for 4 weeks to induce hypertension. During the last week of the infusion, the hamsters were inoculated intranasally with vehicle (V) or SARS-CoV-2 (S; 1 x 105 plaque forming units/ml). Half of the hamsters were sacrificed 1 day post-inoculation (dpi-1) and the other half on dpi-6. Two scoring systems were applied to lung tissue sections stained with hematoxylin and eosin to determine the degree and severity of lung pathology: the first system assessed all pertinent alterations in the lungs, while the second system only assessed the pathology related to the pulmonary vasculature. Lung histopathology scores were calculated as the sum of the airway and lung alveolar scores in arbitrary units (AU). RESULTS/ANTICIPATED RESULTS: Studies revealed that the SARS-CoV-2-infected hamsters exhibited a 76-fold higher total airway score compared to vehicle controls [(AU): V, 0.25 ± 0.1; S, 19.00 ±1.35; p<0.05; n=4]. Total lung alveolar scores (27-fold) [(AU): V, 0.30 ± 0.11; S, 8.0 ± 4.1; p<0.05; n=4] and total vascular scores (17-fold) [(AU): V, 0.35 ± 0.2; S, 6.0 ±1.4; p<0.05; n=4] were also markedly higher compared to controls on dpi-1. AII increased blood pressure, which was sustained through the 4-week infusion period. Under these conditions, body weight slightly dropped by 4.5%. Ongoing studies are assessing the effect of hypertension on the % of airway, alveoli and vessels affected, airway and alveolar severity, and bronchiolar epithelial and type II pneumocyte hyperplasia. DISCUSSION/SIGNIFICANCE: Establishing the hypertensive hamster as a small animal model of COVID-19 will facilitate investigations into why preexisting hypertension is a risk factor for disease severity. These studies could lead to the development of novel therapeuticsfor treating COVID-19 patients with hypertension.

OBJECTIVES/GOALS: Hamsters develop COVID-19 similarly to people because the SARS-CoV-2 spike protein binds with high affinity to hamster ACE2 resulting in host cell entry and replication. Our goal was to establish a hamster model that mirrors the lung and brain pathophysiology observed in COVID-19. METHODS/STUDY POPULATION: Hamsters infected with SARS CoV-2 are sacrificed on day 1 and day 6 postinfection. Lung histopathology scoring model was implemented for assessment all pathological relevant changes in the lungs of infected animals on tissue sections stained with hematoxylin and eosin. To quantify the extent and severity of lung pathology, two scoring systems were used: the first evaluated all relevant changes in the lungs of the infected animals and the second evaluated only the pathology associated with the pulmonary vasculature. Percentage of airway affected, airway severity, bronchiolar epithelial hyperplasia, alveoli affected, alveolar severity, type II pneumocyte hyperplasia and vessels affected were analyzed. Total airway score plus total lung alveolar score give lung histopathology score. RESULTS/ANTICIPATED RESULTS: Compared to the control hamster, the hamsters day 1 postinfection, exhibited a higher total airway score [9.00 ± 1.35 vs. 0.25 ± 0.1; p DISCUSSION/SIGNIFICANCE: Establishing this outstanding small animal model of COVID-19 will facilitate studies investigating diagnostics, prognosis and response to treatment in COVID-19 disease. These studies will provide insights that will complement on-going clinical trials on angiotensin type 1 receptor (AT1R) blockers (ARBs) in COVID-19.

OBJECTIVES/GOALS: Childhood psychiatric symptoms are highly comorbid. Their co-occurrence and association with negative life outcomes is partially explained by deficits in executive control, or processes enabling self-regulation. Here, we test a novel executive neural target in three fMRI tasks and its relevance to shared psychopathology. METHODS/STUDY POPULATION: We studied 60 children [15 F/45 M; mean age (SD)=11.6 years (1.62)] with diverse diagnoses including attention deficit disorder (n=26) and autism spectrum disorder (n=22). We extracted a latent general factor of psychopathology using principal component analyses applied to parent-report Child Behavior Checklist syndrome scores. Subjects completed 3 executive control fMRI probes, tapping adaptive control, working memory, and inhibition. Correlational psychophysiological interaction (cPPI) analysis measured correlations between executive control-related modulations of activity in 414 network-affiliated parcels. We selected parcels exhibiting control-related cross-network correlations as well as control-related activity across all tasks and tested them for association with psychopathology. RESULTS/ANTICIPATED RESULTS: cPPI connectivity matrices were thresholded and graphs were identified using the Network-Based Statistic toolbox (p90th percentile PC) as well as control-related activation (>10% activated voxels; p DISCUSSION/SIGNIFICANCE: Our results examine cross-network interactions between brain regions during 3 fMRI tasks and their role in explaining individual variation in psychopathology. As executive control links to both comorbidity and life outcomes, identifying the clinically-relevant neural correlates of controlled behavior may lead to transdiagnostic treatments.

OBJECTIVES/GOALS: Rodents are the most widely used experimental animals to study disease mechanisms due to their availability and cost-effectiveness. An international drive to investigate the pathophysiology of COVID-19 is inhibited by the resistance of rats and mice to SARS-CoV-2 infection. Our goal was to establish an appropriate small animal model. METHODS/STUDY POPULATION: To recreate the cytokine storm that is associated with COVID-19, we injected angiotensin converting enzyme 2 knockout (ACE2KO) mice (C57BI/6 strain) with lipopolysaccharide (LPS) intraperitoneally and measured the expression of multiple cytokines as a function of time and LPS dose. We then chose a minimum dose (500ug/kg) and time (3h) when multiple cytokines were elevated to measure lung injury scores using a point-counting technique on tissue sections stained with hematoxylin and eosin. The data are expressed as mean percentage of grid points lying within the peribronchial and superficial area in up to 20 fields. Percentage of peribronchial and superficial intrapulmonary hemorrhage, congestion, neutrophil infiltration and area of alveolar space were all assessed. RESULTS/ANTICIPATED RESULTS: Compared to the wildtype group (WT-G), the LPS-injected ACE2KO mice (LPS-G) exhibited a higher percentage of peribronchial intrapulmonary hemorrhage [(%): LPS-G, 10.56 ± 2.06 vs. WT-G, 5.59 ± 0.53; p DISCUSSION/SIGNIFICANCE: Establishing this novel mouse model of COVID-19 will facilitate studies investigating tissue-specific mechanisms of pathogenesis in this disease. This model can also be used to discover novel therapeutic targets and the design of clinical trials focusing on diagnostics, treatments and outcomes in COVID-19.

OBJECTIVES/GOALS: The SARS-CoV-2 (Severe Acute Respiratory Syndrome CoronaVirus-2), which underlies the current COVID-19 pandemic, among other tissues, also targets the central nervous system (CNS). The goal of this study is to investigate mechanisms of neuroinflammation in Lipopolysaccharides (LPS)-treated mouse model and SARS-CoV-2-infected hamsters. METHODS/STUDY POPULATION: In this research I will assay vascular reactivity of cerebral vessels to assess vascular dysfunction within the microcirculation. I will determine expression of proinflammatory cytokines, coagulation factors and AT1 receptors (AT1R) in isolated microvessels from the circle of Willis to assess inflammation, thrombosis and RAS activity in the microvasculature. LPS and SARS-CoV-2, are both associated with coagulopathies and because of that I will measure concentration of PAI-1, von Willebrand Factor, thrombin and D-dimer to assess the thrombotic pathway in the circulation. Histology and immunohistochemistry will assess immune cell type infiltration into the brain parenchyma, microglia activation and severity of neuroinflammation and neural injury. RESULTS/ANTICIPATED RESULTS: We hypothesize that under conditions of reduced ACE2 (e.g., SARS-CoV-2 infection), AT1R activity is upregulated in the microvasculature. In the presence of an inflammatory insult, these AT1Rs promote endothelialitis and immunothrombosis through pro-thrombotic pathways and pro-inflammatory cytokine production leading to endothelial dysfunction in the microvasculature, blood brain barrier (BBB) injury, deficits in cognition and increased anxiety. We will test this hypothesis through 2 aims: Aim 1: Determine the role of the pro-injury arm of the RAS in the pathophysiology of the brain in animal models of neuroinflammation and COVID-19. Aim 1: Determine the role of the protective arm of the RAS in the pathophysiology of the brain in animal models of neuroinflammation and COVID-19. DISCUSSION/SIGNIFICANCE: This study will provide insights that will complement on-going clinical trials on angiotensin type 1 receptor (AT1R) blockers (ARBs) in COVID-19. This research is a necessary first step in understanding mechanisms of brain pathogenesis that can set the groundwork for future studies of more complex models of disease.

Frequent freezing injury greatly influences winter wheat production; thus, effective prevention and a command of agricultural production are vital. The freezing injury monitoring method integrated with ‘3S’ (geographic information systems (GIS), global positioning system (GPS) and remote sensing (RS)) technology has an unparalleled advantage. Using HuanJing (HJ)-1A/1B satellite images of a winter wheat field in Shanxi Province, China plus a field survey, crop types and winter wheat planting area were identified through repeated visual interpretations of image information and spatial analyses conducted in GIS. Six vegetation indices were extracted from processed HJ-1A/1B satellite images to determine whether the winter wheat suffered from freezing injury and its degree of severity and recovery, using change vector analysis (CVA), the freeze injury representative vegetation index and the combination of the two methods, respectively. Accuracy of the freezing damage classification results was verified by determining the impact of freezing damage on yield and quantitative analysis. The CVA and the change of normalized difference vegetation index (ΔNDVI) monitoring results were different so a comprehensive analysis of the combination of CVA and ΔNDVI was performed. The area with serious freezing injury covered 0.9% of the total study area, followed by the area of no freezing injury (3.5%), moderate freezing injury (10.2%) and light freezing injury (85.4%). Of the moderate and serious freezing injury areas, 0.2% did not recover; 1.2% of the no freezing injury and light freezing injury areas showed optimal recovery, 15.6% of the light freezing injury and moderate freezing injury areas showed poor recovery, and the remaining areas exhibited general recovery.

Schistosomiasis has been subjected to extensive control efforts in the People's Republic of China (China) which aims to eliminate the disease by 2030. We describe baseline results of a longitudinal cohort study undertaken in the Dongting and Poyang lakes areas of central China designed to determine the prevalence of Schistosoma japonicum in humans, animals (goats and bovines) and Oncomelania snails utilizing molecular diagnostics procedures. Data from the Chinese National Schistosomiasis Control Programme (CNSCP) were compared with the molecular results obtained.

Sixteen villages from Hunan and Jiangxi provinces were surveyed; animals were only found in Hunan. The prevalence of schistosomiasis in humans was 1.8% in Jiangxi and 8.0% in Hunan determined by real-time polymerase chain reaction (PCR), while 18.3% of animals were positive by digital droplet PCR. The CNSCP data indicated that all villages harboured S. japonicum-infected individuals, detected serologically by indirect haemagglutination assay (IHA), but very few, if any, of these were subsequently positive by Kato-Katz (KK).

Based on the outcome of the IHA and KK results, the CNSCP incorporates targeted human praziquantel chemotherapy but this approach can miss some infections as evidenced by the results reported here. Sensitive molecular diagnostics can play a key role in the elimination of schistosomiasis in China and inform control measures allowing for a more systematic approach to treatment.

Scanning transmission electron microscopy (STEM) allows for imaging, diffraction, and spectroscopy of materials on length scales ranging from microns to atoms. By using a high-speed, direct electron detector, it is now possible to record a full two-dimensional (2D) image of the diffracted electron beam at each probe position, typically a 2D grid of probe positions. These 4D-STEM datasets are rich in information, including signatures of the local structure, orientation, deformation, electromagnetic fields, and other sample-dependent properties. However, extracting this information requires complex analysis pipelines that include data wrangling, calibration, analysis, and visualization, all while maintaining robustness against imaging distortions and artifacts. In this paper, we present py4DSTEM, an analysis toolkit for measuring material properties from 4D-STEM datasets, written in the Python language and released with an open-source license. We describe the algorithmic steps for dataset calibration and various 4D-STEM property measurements in detail and present results from several experimental datasets. We also implement a simple and universal file format appropriate for electron microscopy data in py4DSTEM, which uses the open-source HDF5 standard. We hope this tool will benefit the research community and help improve the standards for data and computational methods in electron microscopy, and we invite the community to contribute to this ongoing project.

An acute gastroenteritis (AGE) outbreak caused by a norovirus occurred at a hospital in Shanghai, China, was studied for molecular epidemiology, host susceptibility and serological roles. Rectal and environmental swabs, paired serum samples and saliva specimens were collected. Pathogens were detected by real-time polymerase chain reaction and DNA sequencing. Histo-blood group antigens (HBGA) phenotypes of saliva samples and their binding to norovirus protruding proteins were determined by enzyme-linked immunosorbent assay. The HBGA-binding interfaces and the surrounding region were analysed by the MegAlign program of DNAstar 7.1. Twenty-seven individuals in two care units were attacked with AGE at attack rates of 9.02 and 11.68%. Eighteen (78.2%) symptomatic and five (38.4%) asymptomatic individuals were GII.6/b norovirus positive. Saliva-based HBGA phenotyping showed that all symptomatic and asymptomatic cases belonged to A, B, AB or O secretors. Only four (16.7%) out of the 24 tested serum samples showed low blockade activity against HBGA-norovirus binding at the acute phase, whereas 11 (45.8%) samples at the convalescence stage showed seroconversion of such blockade. Specific blockade antibody in the population played an essential role in this norovirus epidemic. A wide HBGA-binding spectrum of GII.6 supports a need for continuous health attention and surveillance in different settings.

OBJECTIVES/GOALS: Flavorings differ between brands and tobacco products, potentially altering the sensory perceptions. This study aimed to examine discrepancies in flavor preference across various non-cigarette tobacco products among a national representative sample of US adult regular tobacco users. METHODS/STUDY POPULATION: Data from the Population Assessment of Tobacco and Health (PATH) Study Wave 3 (W3) were used. Weighted prevalence of flavor preference for various tobacco products, including electronic nicotine delivery systems (ENDS), traditional cigars, cigarillos/filtered cigars, hookah and snus/smokeless, was presented for 9,037 adult current and new former users of multiple flavored tobacco products. Within-subject flavor discrepancies were assessed using generalized estimating equations (GEE) models considering the complex sampling design of the PATH study. RESULTS/ANTICIPATED RESULTS: Most regular users of a flavored tobacco products reported using one flavor category per product. Fruit flavors, followed by tobacco, were the most common flavor categories among ENDS (32% and 25%, respectively) and hookah users (44% and 36%, respectively). Tobacco flavor was the most common among regular users of traditional cigars (80%), cigarillos/filtered cigars (55%), and smokeless tobacco (79%). Polytobacco users of ENDS and traditional cigars had the largest discrepancy, where about 68-76%% used different flavor categories when switching products. Conversely, polytobacco users of traditional cigars and cigarillos/filtered cigars had the lowest discrepancy (23-25%). DISCUSSION/SIGNIFICANCE OF IMPACT: Many consumers of multiple tobacco products had different flavor preferences when switching between products. In the event of a partial or full flavor ban for ENDS, these findings raise questions about consumer loyalty to a particular tobacco product or a particular flavor category. Conflict of Interest Description: MLG serves as a paid consultant for Johnson & Johnson and has received research grant from Pfizer, manufacturers of smoking cessation medications. The other authors have no conflicts to declare. CONFLICT OF INTEREST DESCRIPTION: MLG serves as a paid consultant for Johnson & Johnson and has received research grant from Pfizer, manufacturers of smoking cessation medications. The other authors have no conflicts to declare.

OBJECTIVES/GOALS: Wheezing has been shown to be associated with use of cigarettes, and more recently, electronic nicotine delivery systems (ENDS). This study assessed the association of poly use of tobacco products with wheezing among a national representative sample of US adult current tobacco users. METHODS/STUDY POPULATION: Data from the Population Assessment of Tobacco and Health (PATH) Study Wave 3 (W3) were used. Weighted prevalences of self-reported wheezing and related respiratory symptoms for non-users compared to users of cigarettes, ENDS, cigars, and any combination of these products (poly use of tobacco products) were presented for 28,082 adults. The cross-sectional association of tobacco use with self-reported wheezing and other related respiratory symptoms was assessed using weighted multivariable and ordinal logistic regression with consideration of complex sampling design. RESULTS/ANTICIPATED RESULTS: Most adults who reported on wheezing symptoms did not currently use cigarettes, ENDS or cigars (79%), 15% used cigarettes, 3% used a combination of cigarettes, ENDS and cigars, 1% used ENDS, and 1% used cigars. Significantly higher odds of ever had wheezing or whistling in chest at any time in the past was observed among current cigarette (adjusted OR: 2.62, 95%CI: 2.35, 2.91), ENDS (1.49, 95%CI: 1.14, 1.95), and poly users (2.67, 95%CI: 2.26, 3.16) compared to non-users. No differences were seen for cigar use. Polytobacco use was associated with a higher odds of ever wheezing when compared to ENDS (1.61, 95%CI: 1.19, 2.17) and cigar use (2.87, 95%CI: 1.93, 4.26), but not cigarettes. DISCUSSION/SIGNIFICANCE OF IMPACT: Wheezing is associated with the use of cigarettes, ENDS, or any combination of cigarette, ENDS and cigars likely due to the inhalation of noxious chemicals and gases found in the smoke of cigarettes and ENDS that are likely to increase the odds of experiencing wheezing. CONFLICT OF INTEREST DESCRIPTION: MLG serves as a paid consultant for Johnson & Johnson and has received research grant from Pfizer, manufacturers of smoking cessation medications. The other authors have no conflicts to declare.

Several autonomous phase-sensitive radio-echo sounders (ApRES) were deployed at Greenland glaciers to investigate ice deformation. Different attenuation settings were tested and it was observed that, in the presence of clipping of the deramped ApRES signal, each setting produced a different result. Specifically, higher levels of clipping associated with lower attenuation produced an apparent linear increase of diurnal vertical cumulative displacement with depth, and obscured the visibility of the basal reflector in the return amplitude. An example with a synthetic deramped signal confirmed that these types of artifacts result from the introduction of harmonics from square-wave-like features introduced by clipping. Apparent linear increase of vertical displacement with depth occurs when the vertical position of a near-surface internal reflector changes in time. Artifacts in the return amplitude may obscure returns from internal reflectors and the basal reflector, making it difficult to detect thickness evolution of the ice and to correctly estimate vertical velocities. Variations in surface melt during ApRES deployments can substantially modulate the received signal strength on short timescales, and we therefore recommend using higher attenuator settings for deployments in such locations.

A new developed spatially targeted mollusciciding technology for snail control was utilised in a research site. This study aims to analyse whether this technology can achieve rational effectiveness compared with the routine method. Snail density was monitored every spring and autumn from 2010 to 2017 at the research site and routine mollusciciding for snail control was then performed. After snail density monitoring in spring 2018, spatially targeted mollusciciding technology was adopted. Log-linear regression and nonlinear regression models were used for snail density prediction in autumn 2018 and the predicted value was compared with the actual snail density in autumn 2018 to verify the effectiveness of the spatially targeted mollusciciding. Monitoring results showed that overall snail density in the research site decreased from 2010 to 2018. The monitored snail density in autumn 2018 was 0.014/0.1 m2. Predicted by the log-linear regression model, the snail density in autumn 2018 would be 0.028 (95% CI 0.11–0.072)/0.1 m2. Predicted by the nonlinear regression model, the snail density growth in autumn 2018 in contrast to spring 2018 would be 79.79% (95% CI 54.81%–104.77%) and the actual value was 55.56%. Therefore, the effectiveness of the first application of spatially targeted mollusciciding was acceptable. However, the validation of its sustainable effectiveness still needs a replicated study comparing areas where targeted and untargeted methods are applied simultaneously and both snail abundance and human infection are monitored.

Correct diagnosis of cause of death is necessary to suggest the most effective management interventions to reduce perinatal lamb mortality. Haemorrhage on the surface of the brain has been used as a field diagnostic tool to allocate lambs to a cause of death category, but the usefulness of this method was unclear. This study aimed to evaluate whether gross pathology was related to neuronal death and whether haemorrhage of the central nervous system (CNS) was distinct between differing causes of death, enabling indicators to be used in field diagnoses. Lambs dying from natural causes (n = 64) and from euthanasia (n = 7) underwent postmortem examination, then the brain and spinal cord were extracted and examined histologically. Histological changes consistent with neuronal death were not detected in any lamb. Haemorrhage of the meninges and/or parenchyma of the CNS occurred in all lambs. The age of the haemorrhage indicated that it occurred near the time of death in most lambs. Dilation of blood vessels varied in severity but appeared to be unrelated to causal diagnosis, severity of subcutaneous oedema, breathing or milk status. Moderate or severe dilation of blood vessels and haemorrhage of the CNS did not occur in all lambs with alternative clear indicators of dystocia and occurred in all death classifications, so it could not be used as diagnostic indicators for classification of cause of death. Dilation and haemorrhage were unrelated to neuronal damage and may have been artefactual. In conclusion, haemorrhage of the CNS was not indicative of neuronal damage and could not be used to distinguish between lambs with clear indicators of differing causes of death, so it is not recommended as a field diagnostic tool.

This study aimed to investigate the clinical characteristics and to analyse the epidemiological features of coronavirus disease 2019 (COVID-19) patients during convalescence. In this study, we enrolled 71 confirmed cases of COVID-19 who were discharged from hospital and transferred to isolation wards from 6 February to 26 March 2020. They were all employees of Zhongnan Hospital of Wuhan University or their family members of which three cases were <18 years of age. Clinical data were collected and analysed statistically. Forty-one cases (41/71, 57.7%) comprised medical faculty, young and middle-aged patients (aged ⩽60 years) accounted for 81.7% (58/71). The average isolation time period for all adult patients was 13.8 ± 6.1 days. During convalescence, RNA detection results of 35.2% patients (25/71) turned from negative to positive. The longest RNA reversed phase time was 7 days. In all, 52.9% of adult patients (36/68) had no obvious clinical symptoms, and the remaining ones had mild and non-specific clinical symptoms (e.g. cough, sputum, sore throat, disorders of the gastrointestinal tract etc.). Chest CT signs in 89.7% of adult patients (61/68) gradually improved, and in the others, the lesions were eventually absorbed and improved after short-term repeated progression. The main chest CT manifestations of adult patients were normal, GGO or fibre streak shadow, and six patients (8.8%) had extrapulmonary manifestations, but there was no significant correlation with RNA detection results (r = −0.008, P > 0.05). The drug treatment was mainly symptomatic support therapy, and antibiotics and antiviral drugs were ineffective. It is necessary to re-evaluate the isolation time and standard to terminate isolation for discharged COVID-19 patients.

Introduction: Improved access to HIV testing would benefit the one in six Canadians living with undiagnosed HIV, and potentially reduce transmission. Emergency departments may be the first or only point of contact with the healthcare system for people exposed to HIV; however, HIV testing remains inaccessible in many EDs in Canada. Methods: We used a grounded theory approach to characterize the experiences and context of HIV testing in Canadian EDs. We conducted semi-structured phone interviews with ED and public health practitioners from a purposive sample of urban, rural, academic, and community ED catchment areas. Thematic analysis was performed through iterative readings by two authors. Results were triangulated through consultation with public health and HIV experts. Results: Data were obtained from 16 ED physicians and 8 public health practitioners. HIV tests were infrequently performed in the EDs of our sample. Informants from higher incidence regions believed that greater availability of HIV tests in the ED would benefit the populations they serve. In half of the sample, rapid HIV tests were available. However, indications for testing were most often occupational or known high-risk exposure. Notably, two urban EDs in British Columbia screened all patients who otherwise needed blood tests (opt-out), but had shifted to opt-in testing at the time of this study. Consent practices and perceived requirements varied widely between sites; this confused or frustrated physicians. Most EDs were unable to offer a test result to patients during their visit as results were not available until days to weeks later. Commonly, the ordering physician was responsible for communicating results. Some EDs had an assigned physician managing all results on a given day while others relied on public health units for follow-up. All EDs reported access to public health clinics for ongoing care. Barriers to offering a test in the ED included time required for consent, discomfort with pre-test counselling, delay in results availability and unclear processes for follow-up. Conclusion: We describe substantial regional and within-site variation in HIV testing practices across a diverse sample of EDs across Canada. These findings highlight disparities in access to HIV testing and warrant a national discussion on best practices for testing in EDs with an emphasis on reducing barriers for high-risk populations and addressing unmet needs.