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Abstract Background The rapidly growing availability of diverse full genome sequences from across the world is increasing the feasibility of studying the large-scale population processes that underly observable pattern of virus diversity. In particular, characterizing the genetic structure of virus populations could potentially reveal much about how factors such as geographical distributions, host ranges and gene flow between populations combine to produce the discontinuous patterns of genetic diversity that we perceive as distinct virus species. Among the richest and most diverse full genome datasets that are available is that for the dicotyledonous plant infecting genus, Begomovirus, in the Family Geminiviridae. The begomoviruses all share the same whitefly vector, are highly recombinogenic and are distributed throughout tropical and subtropical regions where they seriously threaten the food security of the world's poorest people. Results We focus here on using a model-based population genetic approach to identify the genetically distinct sub-populations within the global begomovirus meta-population. We demonstrate the existence of at least seven major sub-populations that can further be sub-divided into as many as thirty four significantly differentiated and genetically cohesive minor sub-populations. Using the population structure framework revealed in the present study, we further explored the extent of gene flow and recombination between genetic populations. Conclusions Although geographical barriers are apparently the most significant underlying cause of the seven major population sub-divisions, within the framework of these sub-divisions, we explore patterns of gene flow to reveal that both host range differences and genetic barriers to recombination have probably been major contributors to the minor population sub-divisions that we have identified. We believe that the global Begomovirus population structure revealed here could facilitate population genetics studies into how central parameters of population genetics namely selection, recombination, mutation, gene flow, and genetic drift shape the global begomovirus diversity.

Abstract Background Tomato-infecting begomoviruses are widely distributed across the world and cause diseases of high economic impact on wide range of agriculturally important crops. Though recombination plays a pivotal role in diversification and evolution of these viruses, it is currently unknown whether there are differences in the number and quality of recombination events amongst different tomato-infecting begomovirus species. To examine this we sought to characterize the recombination events, estimate the frequency of recombination, and map recombination hotspots in tomato-infecting begomoviruses of South and Southeast Asia. Results Different methods used for recombination breakpoint analysis provided strong evidence for presence of recombination events in majority of the sequences analyzed. However, there was a clear evidence for absence or low Recombination events in viruses reported from North India. In addition, we provide evidence for non-random distribution of recombination events with the highest frequency of recombination being mapped in the portion of the N-terminal portion of Rep. Conclusion The variable recombination observed in these viruses signified that all begomoviruses are not equally prone to recombination. Distribution of recombination hotspots was found to be reliant on the relatedness of the genomic region involved in the exchange. Overall the frequency of phylogenetic violations and number of recombination events decreased with increasing parental sequence diversity. These findings provide valuable new information for understanding the diversity and evolution of tomato-infecting begomoviruses in Asia.

Abstract Background Inflammation is believed to play an important role in the pathology of Alzheimer's disease (AD) and cytokine production is a key pathologic event in the progression of inflammatory cascades. The current study characterizes the cytokine expression profile in the brain of two transgenic mouse models of AD (TgAPPsw and PS1/APPsw) and explores the correlations between cytokine production and the level of soluble and insoluble forms of Aβ. Methods Organotypic brain slice cultures from 15-month-old mice (TgAPPsw, PS1/APPsw and control littermates) were established and multiple cytokine levels were analyzed using the Bio-plex multiple cytokine assay system. Soluble and insoluble forms of Aβ were quantified and Aβ-cytokine relationships were analyzed. Results Compared to control littermates, transgenic mice showed a significant increase in the following pro-inflammatory cytokines: TNF-α, IL-6, IL-12p40, IL-1β, IL-1α and GM-CSF. TNF-α, IL-6, IL-1α and GM-CSF showed a sequential increase from control to TgAPPsw to PS1/APPsw suggesting that the amplitude of this cytokine response is dependent on brain Aβ levels, since PS1/APPsw mouse brains accumulate more Aβ than TgAPPsw mouse brains. Quantification of Aβ levels in the same slices showed a wide range of Aβ soluble:insoluble ratio values across TgAPPsw and PS1/APPsw brain slices. Aβ-cytokine correlations revealed significant relationships between Aβ1–40, 1–42 (both soluble and insoluble) and all the above cytokines that changed in the brain slices. Conclusion Our data confirm that the brains of transgenic APPsw and PS1/APPsw mice are under an active inflammatory stress, and that the levels of particular cytokines may be directly related to the amount of soluble and insoluble Aβ present in the brain suggesting that pathological accumulation of Aβ is a key driver of the neuroinflammatory response.

Background: A retrospective chart audit was performed to review biliary stent utilisation from January 2020 to January 2021. Non-guideline-based stent insertion was identified in 16% of patients with common bile duct (CBD) stones presenting for endoscopic retrograde cholangiopancreatography (ERCP). To improve this knowledge-practice gap, a quality improvement (QI) intervention was devised and trialled., Aim: To synchronise clinical indications for biliary stent insertion in patients with CBD stones in accordance with published guidelines., Methods: Using a QI pre-post study design, chart audits were completed and shared with the ERCP team (n=6). Indication for biliary stent insertion was compared to published guidelines assessed by two reviewers independently ( kappa statistic calculated). The QI intervention included an education session and quarterly practice audits. An interrupted time series with segmented regression was completed., Results: A total of 661 patients (337 F), mean age 59±19 years (range 12-98 years), underwent 885 ERCPs during this postintervention period. Of 661 patients, 384 (58%) were referred for CBD stones. A total of 192 biliary stents (105 plastic, 85 metal) were placed during the first ERCP (192/661, 29%), as compared with the preintervention year (223/598, 37%, p=0.2). Furthermore, 13/192 stents (7%) were placed not in accordance with published guidelines ( kappa =0.53), compared with 63/223 (28%) in the preintervention year (p<0.0001). A 75% reduction in overall avoidable stent placement was achieved with a direct cost avoidance of $C97 500. For the CBD stone subgroup, there was an 88% reduction in avoidable biliary stent placement compared with the preintervention year (8/384, 2% vs 61/375, 16%, p<0.0001)., Conclusions: Education with audit and feedback supported the closing of a knowledge-to-practice gap for biliary stent insertion during ERCP, especially in patients with CBD stones. This has resulted in a notable reduction of avoidable stent placements and additional follow-up ERCPs and an overall saving of healthcare resources., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2024. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Introduction: Acute kidney injury (AKI) is common in the perioperative setting and associated with poor outcomes. Whether clinical decision support improves early management and outcomes of AKI on surgical units is uncertain., Methods: In this cluster-randomized, stepped-wedge trial, 8 surgical units in Alberta, Canada were randomized to various start dates to receive an education and clinical decision support intervention for recognition and early management of AKI. Eligible patients were aged ≥18 years, receiving care on a surgical unit, not already receiving dialysis, and with AKI., Results: There were 2135 admissions of 2038 patients who met the inclusion criteria; mean (SD) age was 64.3 (16.2) years, and 885 (41.4%) were females. The proportion of patients who experienced the composite primary outcome of progression of AKI to a higher stage, receipt of dialysis, or death was 16.0% (178 events/1113 admissions) in the intervention group; and 17.5% (179 events/1022 admissions) in the control group (time-adjusted odds ratio, 0.76; 95% confidence interval [CI], 0.53-1.08; P  = 0.12). There were no significant differences between groups in process of care outcomes within 48 hours of AKI onset, including administration of i.v. fluids, or withdrawal of medications affecting kidney function. Both groups experienced similar lengths of stay in hospital after AKI and change in estimated glomerular filtration rate (eGFR) at 3 months., Conclusion: An education and clinical decision support intervention did not significantly improve processes of care or reduce progression of AKI, length of hospital stays, or recovery of kidney function in patients with AKI on surgical units., (© 2024 International Society of Nephrology. Published by Elsevier Inc.)

Background: Indiscriminate use of laboratory blood testing in hospitals contributes to patient discomfort and healthcare waste. Patient engagement in low-value healthcare can help reduce overuse. Understanding patient experience is necessary to identify opportunities to improve patient engagement with in-hospital laboratory testing., Objectives: To understand patient experience with the process of in-hospital laboratory blood testing., Methods: We used a qualitative study design via semistructured interviews conducted online or over the phone. Participants were adult patients or family members/caregivers (≥18 years of age) with a recent (within 12 months of interview) experience of hospitalization in Alberta or British Columbia, Canada. We identified participants through convenience sampling and conducted interviews between May 2021 and June 2022. We analysed transcripts using thematic content analysis. Recruitment was continued until code saturation was reached., Results: We interviewed 16 participants (13 patients, 1 family member and 2 caregivers). We identified four themes from patients' experiences of in-hospital laboratory blood testing: (i) patients need information from healthcare teams about expected blood testing processes, (ii) blood draw processes should consider patient comfort and preferences, (iii) patients want information from their healthcare teams about the rationale and frequency of blood testing and (iv) patients need information on how their testing results affect their medical care., Conclusion: Current laboratory testing processes in hospitals do not facilitate shared decision-making and patient engagement. Patient engagement with laboratory testing in hospitals requires an empathetic healthcare team that provides clear communication regarding testing procedures, rationale and results, while considering patient preferences and offering opportunities for involvement., Patient or Public Contribution: We interviewed 16 patients and/or family members/caregivers regarding their in-hospital laboratory blood testing experiences. Our findings show correlations between patient needs and patient recommendations to make testing processes more patient-centred. To bring a lived-experience lens to this study, we formed a Patient Advisory Council with 9-11 patient research partners. Our patient research partners informed the research design, co-developed participant recruitment strategies, co-conducted data collection and informed the data analysis. Some of our patient research partners are co-authors of this manuscript., (© 2023 The Authors. Health Expectations published by John Wiley & Sons Ltd.)

Background: Ensuring language concordant care through medical interpretation services (MIS) allows for accurate information sharing and positive healthcare experiences. The COVID-19 pandemic led to a regional halt of in-person interpreters, leaving only digital MIS options, such as phone and video. Due to longstanding institutional practices, and lack of accessibility and awareness of these options, digital MIS remained underused. A Multimodal Medical Interpretation Intervention (MMII) was developed and piloted to increase digital MIS usage by 25% over an 18-month intervention period for patients with limited English proficiency., Methods: Applying quality improvement methodology, an intervention comprised digital MIS technology and education was trialled for 18 months. To assess intervention impact, the number of digital MIS minutes was measured monthly and compared before and after implementation. A questionnaire was developed and administered to determine healthcare providers' awareness, technology accessibility and perception of MIS integration in the clinical workflow., Results: Digital MIS was used consistently from the beginning of the COVID-19 pandemic (March 2020) and over the subsequent 18 months. The total number of minutes of MIS use per month increased by 44% following implementation of our intervention. Healthcare providers indicated that digital MIS was vital in facilitating transparent communication with patients, and the MMII ensured awareness of and accessibility to the various MIS modalities., Conclusion: Implementation of the MMII allowed for an increase in digital MIS use in a hospital setting. Providing digital MIS access, education and training is a means to advance patient-centred and equitable care by improving accuracy of clinical assessments and communication., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2024. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Background: A coalition (Strategic Clinical Improvement Committee), with a mandate to promote physician quality improvement (QI) involvement, identified hospital laboratory test overuse as a priority. The coalition developed and supported the spread of a multicomponent initiative about reducing repetitive laboratory testing and blood urea nitrogen (BUN) ordering across one Canadian province. This study's purpose was to identify coalition factors enabling medicine and emergency department (ED) physicians to lead, participate and influence appropriate BUN test ordering., Methods: Using sequential explanatory mixed methods, intervention components were grouped as person focused or system focused. Quantitative phase/analyses included: monthly total and average of the BUN test for six hospitals (medicine programme and two EDs) were compared pre initiative and post initiative; a cost avoidance calculation and an interrupted time series analysis were performed (participants were divided into two groups: high (>50%) and low (<50%) BUN test reduction based on these findings). Qualitative phase/analyses included: structured virtual interviews with 12 physicians/participants; a content analysis aligned to the Theoretical Domains Framework and the Behaviour Change Wheel. Quotes from participants representing high and low groups were integrated into a joint display., Results: Monthly BUN test ordering was significantly reduced in 5 of 6 participating hospital medicine programmes and in both EDs (33% to 76%), resulting in monthly cost avoidance (CAN$900-CAN$7285). Physicians had similar perceptions of the coalition's characteristics enabling their QI involvement and the factors influencing BUN test reduction., Conclusions: To enable physician confidence to lead and participate, the coalition used the following: a simply designed QI initiative, partnership with a coalition physician leader and/or member; credibility and mentorship; support personnel; QI education and hands-on training; minimal physician effort; and no clinical workflow disruption. Implementing person-focused and system-focused intervention components, and communication from a trusted local physician-who shared data, physician QI initiative role/contribution and responsibility, best practices, and past project successes-were factors influencing appropriate BUN test ordering., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2023. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Background: Attempts have been made to reduce excessive laboratory test ordering; however, the problem persists and barriers to physician involvement in quality improvement (QI) remain. We sought to understand physician participation experience following a laboratory test overuse initiative supported by a QI coalition., Methods: As part of a larger mixed-methods study, structured virtual interviews were conducted with 12 physicians. The Theoretical Domains Framework (TDF) and the Behavioural Change Wheel (BCW) were used to identify characteristics that influence physician behaviour change for QI leadership and participation and appropriate blood urea nitrogen (BUN) test ordering. A content analysis of physicians' statements to the TDF was performed, resulting in overarching themes; relevant TDF domains were mapped to the intervention functions of the BCW., Results: Nine overarching themes emerged from the data. Eight of 14 TDF domains influence QI leadership and participation, and 10 influence appropriate BUN-test ordering behaviours. The characteristics participants described that promoted a change in their QI participation, leadership and appropriate BUN-test ordering were: QI education with hands-on training; physician peer mentorship/support; personnel assistance (QI and analytics) and communication from a trusted/credible physician leader who shares data and insights about the physician role in the initiative, clinical best practice and past project success. Other elements included: a simply designed initiative requiring minimal effort and no clinical workflow disruptions; revised order forms/panels and limiting test-order frequency when laboratory tests are normal. Additionally, various future intervention strategies were identified. For their initial initiative participation, physicians acknowledged coalition leader or member credibility was more important than awareness of the coalition., Conclusions: Based on physicians' described perceptions and experiences, coalition characteristics that influenced their QI leadership and participation, and appropriate BUN-test ordering behaviours were revealed; these characteristics aligned to several TDF domains. The findings suggest that these behaviours are multidimensional, requiring a multistrategy approach to change behaviour., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2023. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Introduction: The objective of this quality improvement, interventional study regarding patients with diabetes undergoing diabetic ophthalmology outpatient surgery aimed to develop, implement, and evaluate a new diabetic algorithm to improve safety, operating room efficiency, and decrease supply cost., Methods: A multidisciplinary study team was assembled, including ophthalmologists, endocrinologists, anesthesiologists, management, and nurses to review the current diabetic protocol. From August 2016 to July 2017, 13 patient safety concerns or incident reports were reviewed that identified two serious cases of hypoglycemia. Using the concerns data, frontline perspectives, and reviewing best practice guidelines, a new diabetic algorithm was developed and trialed for 24 months. The new algorithm limited the use of an existing preoperative insulin protocol and reduced the number of nurses required. The number of adverse events, nursing setup process steps, setup time, and preoperative insulin infusion protocols used were collected. An evaluation of the supply costs was performed., Results: After implementing the new diabetic algorithm, zero safety incidents were reported, and a 97.5% reduction in the use of preoperative insulin protocol resulted. Nursing staff perceived that the new diabetic algorithm was easier to configure, 23 minutes faster to set up, and required one nursing staff member. Supply cost was reduced by $30.63 (Canadian Dollars, CAD) per patient., Conclusion: Perioperative glucose irregularities may threaten patient safety and surgical outcomes. Healthcare professionals must improve patient safety, decrease healthcare expenditure, and prevent unnecessary delays. Multidisciplinary frontline staff experiential knowledge aided in the recognition of potential problems and comprehensive solutions to optimize patient care., Competing Interests: Conflict of Interest: None.

Urine testing on asymptomatic patients is not aligned with guidelines; however, stroke survivors have trouble communicating symptoms, and urinary tract infections (UTIs) are a recognised poststroke complication. All stroke inpatients at a tertiary rehabilitation hospital underwent urine testing on admission. We led a quality improvement (QI) project on one stroke rehabilitation unit aimed to reduce admission urine testing from 100% to 0%. Baseline audit representing 2 weeks of admissions identified 27 of 28 patients had urine tests; however, none required UTI treatment despite 3 positive culture results. Estimated cost of testing was $C675. QI tools identified that a standardised paper-based admission form facilitated automatic urine testing. Project intervention strategies included education, clinicians crossing off urine orders and unit clerks flagging unaddressed orders for reassessment. A chart audit after 4 weeks and prescriber survey after 6 months assessed impact. Postintervention audit (n=23) revealed 1 patient had admission urine tests, 22 orders were crossed out, 1 chart was flagged and estimated testing cost declined from $C675 to $C25. Six urine tests were completed after admission and two patients required UTI treatment. Post 6 months, unit clerks assumed the role to cross out the order on the standardised form, and no patient had routine admission urine testing. There was no clinical benefit in screening for UTIs prior to stroke rehabilitation. This project is a practical example of deadopting a practice promoted by standardised order forms., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2022. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Objective: The spread of the COVID-19 virus has caused an unforeseen strain on the healthcare system and particularly on healthcare workers (HCW). In this study, 1 year after the COVID-19 pandemic began, we used photovoice, a visual photographic approach, to understand HCW needs, concerns and resilience and to determine improvement strategies aligned with the HCW-described challenges., Methods: Using a qualitative design, HCW were recruited from a single Western Canadian hospital, voluntarily submitting a photographic image and narrative that depicts their experiences. An artist artistically enhanced the photovoice submissions, which were then displayed at the hospital-based art gallery for public display. A survey was used to collect feedback from gallery viewers. Inductive thematic analysis was completed identifying themes from the photovoice narratives and survey comments, aiding the identification of recommendations., Results: There were 25 submissions, and 1281 individuals viewed the art exhibit. Six themes emerged: (1) hopeful and resilient, (2) pandemic fatigue-negative mental and physical states, (3) personal protective equipment is our armour but masks who we are, (4) human connection, (5) responsibility, preparation and obligation and (6) technology surge. According to survey results from the art exhibit, the use of photovoice was a creative method that personalised the HCW experience and validated viewers' perceptions of the difficulties faced by HCW. Ten improvement strategies that were aligned with the described challenges were identified., Conclusion: The ongoing COVID-19 pandemic continues to strain HCW. Photovoice has great potential in the professional clinical setting to provide unique insights that narrative language alone cannot capture. Future research exploring the longitudinal impact of COVID-19, reviewing photographs at different timepoints could be beneficial. Using this method as a creative outlet intervention and evaluating participation artistic experience may offer additional insights to further support both HCW and patients., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2022. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Background: Most undergraduate medical students (UMS) do not receive any formal exposure to quality improvement (QI) efforts in healthcare during the entirety of their undergraduate programme. This is despite the rising interest amongst UMS and the unique potential that UMS hold as an innovator unencumbered by previous biases. To explore this, we implemented an undergraduate training programme that provides experiential QI education., Approach: The 15-week Summer Healthcare Improvement Programme (SHIP) was established in 2017, supported by a regional physician QI leadership coalition, a QI consultant preceptor who is linked to both the local university and health organisation and an UMS leadership group. Students were assigned QI projects that were aligned with the health organisation's purpose and scope. Students co-led the project to completion with mentorship from both physician QI leaders, and residents. Student competencies were formatively assessed by completing QI activities and a programme survey., Results: From 2017 to 2019, 19 students completed 22 QI projects, academic posters and publications, and all received QI certification. The majority (72%) of students felt involvement in SHIP increased their QI knowledge and skills, 90% believed SHIP would benefit their peers, and 71% of students felt it directly applied to their future careers., Discussion: Benefits of the programme were threefold: provided students with early experiential QI exposure, provided student QI leaders who possess dedicated time and effort to complete projects over the summer months and provided a physician QI learning continuum implemented with minimal to no additional cost to either the university or health organisation., (© 2022 John Wiley & Sons Ltd and The Association for the Study of Medical Education.)

Background: Patients in remote communities who risk premature delivery require transfer to a tertiary care centre for obstetric and neonatal care. Following stabilisation, many patients are candidates for outpatient management but cannot be discharged to their home communities due to lack of neonatal intensive care unit (ICU) support., Problem: Without outpatient accommodation proximal to neonatal ICU, these patients face prolonged hospitalisation-an expensive option with medical, social and psychological consequences. Therefore, we sought to establish an alternative accommodation for out-of-town stable antepartum patients., Methods: Quality Improvement approaches were used to identify process strengths and opportunities for improvement on the antepartum ward in a tertiary care centre. Physician and patient surveys informed outpatient accommodation programme development by a multidisciplinary team. The intervention was implemented using a plan-do-study-act cycle. Barriers to patient discharge and enrolment in the programme were analysed by completing thematic and strengths-weaknesses-opportunities-threats (SWOT) analysis., Results: Physicians broadly supported safe outpatient management, whereas patients were hesitant to leave the hospital even when physicians assured safety. Our alternative accommodation was pre-existing and cost-effective, however, we encountered significant barriers. The physical space limited family visits and social interaction, lacked desired amenities,and the programme proved inconvenient to patients. The thematic and SWOT analysis identified aspects of the intervention which can be optimised to develop future actionable strategies., Conclusion: The utilisation of acute care beds is costly for the healthcare system and must be allocated judiciously. Patient needs, experience and health system barriers need to be considered when establishing alternative outpatient accommodations and strategies for stable antepartum patients., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2022. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Healthcare providers and administrators are incorporating patient feedback to drive local health system improvement. Improvement interventions, including patient feedback, guided a novel approach for rheumatology patient appointment preparedness. We tested the interventions in a single rheumatology clinic. A comprehensive patient questionnaire was developed by assessing patients' clinic access using patient journey mapping. The questionnaire was administered to a random cohort of 125 rheumatology patients about their feedback on pre-clinic practices. From the responses, modifications were made to existing administrative practices within the clinic. The modified practices were tested with an additional cohort of 10 patients aligned to one rheumatologist, assessing overall patients' preparedness and clinic visit cycle time. It was identified from the questionnaire that during appointment booking, inconsistent pre-clinic planning information was communicated to patients and the appointment package did not support patient preparedness, resulting in extending clinic visits. Standardization of the appointment phone call script, updating the appointment package, and inclusion of a clinic visit preparation checklist, reduced the clinic visit cycle time by 10 minutes for new patients, and 5 minutes for existing patients. The participating clinic rheumatologist also perceived improved patient preparedness and clinic visit flow based on the modified practices. In this study, patient feedback was used to identify patient-centred interventions to improve patient preparedness at clinic visits. The interventions developed were simple and easy to incorporate into practice. Systematic collection of and strategies based on patient feedback was determined to be a valid, meaningful method for incorporating clinical quality improvement. KEY POINTS : • Structured patient feedback can inform quality improvement practices in a rheumatological clinic setting. • A patient journey map outlining healthcare clinic access can help to understand patient experiences and needs. • Simple, patient-centred interventions, such as an appointment package and a consistent telephone reminder script, improved patient preparedness and reduced average clinic cycle time., (© 2021. International League of Associations for Rheumatology (ILAR).)

Although biologics are well-studied, expertise regarding their use is often lacking. Many biologics have been added to the market in recent years with distinctive characteristics. This study was designed to create a tool to assist physicians involved in the care of patients with psoriasis undergoing biologic treatment. We used a quality improvement approach to develop and trial an educational visual aid to deliver comprehensive information about biologics in a convenient manner. As a pilot study, trialing this tool was carried out on a small scale to test the feasibility of both the study design and the visual aid itself, with 8 physician and 8 patients completing questionnaires evaluating the visual aid. From our results, the tool was helpful for improving patient knowledge of biologic treatment and their engagement in clinical decision-making. This visual aid may serve as a central convenient biologic resource for physicians., Competing Interests: All of the authors declare no potential conflicts of interest with respect to their research, authorship and/or publication of this article.

Background: Trauma of hospitalization is characterized by patient-reported disturbances in sleep, mobility, nutrition, and/or mood and one study suggested it was associated with more 30-day readmissions., Objective: To define the trauma of hospitalization in medical inpatients and determine whether higher rates of disturbance correlate with adverse post-discharge outcomes., Design: A prospective cohort study was conducted between June 2018 and August 2019 with patients reporting disturbances in sleep, mobility, nutrition, and/or mood. High trauma of hospitalization was defined as disturbance in 3 or 4 domains., Participants: General medicine inpatients at an academic hospital in Edmonton, Canada., Main Measures: 7-day, 30-day, and 90-day rates of death, unplanned hospital readmission, or emergency department (ED) visit., Key Results: Of 299 patients (mean age 65.9 years, 47.8% female, mean Charlson score 3.6, and mean length of stay 8.2 days), 260 (87.0%) reported disturbance in at least one domain (most commonly nutrition or mobility) during their hospitalization, 179 (59.9%) reported disturbances in multiple domains, and 87 (29.1%) met the criteria for high trauma of hospitalization. Patients who reported a high trauma of hospitalization did not differ from those reporting less hospitalization disturbances in terms of demographics, burden of comorbidities, or length of stay, but did report higher rates of pre-hospital disturbances in sleep (32.3% vs. 14.4%, p = 0.03), nutrition (77.4% vs. 54.4%, p = 0.02), and mood (41.9% vs. 13.3%, p = 0.0007). High trauma of hospitalization was not significantly associated with death, readmission, or ED visit at 7 days (12.6% vs. 11.3%, aOR 1.13 [95% CI 0.52-2.46]), 30 days (31.0% vs. 32.1%, aOR 1.03 [95% CI 0.59-1.79]), or 90 days (52.9% vs. 50.9%, aOR 1.16 [95% CI 0.69-1.94]) after discharge., Conclusions: In-hospital disturbances in sleep, mobility, nutrition, and mood are common in medical inpatients but were not associated with post-discharge outcomes., (© 2020. Society of General Internal Medicine.)

Introduction: Laboratory blood testing is one of the most high-volume medical procedures and continues to increase steadily with instances of inappropriate testing resulting in significant financial implications. Studies have suggested that the design of a standard hospital admission order form and laboratory request forms influence physician test ordering behaviour, reducing inappropriate ordering and promoting resource stewardship., Aim/method: To redesign the standard medicine admission order form-laboratory request section to reduce inappropriate blood urea nitrogen (BUN) testing., Results: A redesign of the standard admission order form used by general internal medicine physicians and residents in two large teaching hospitals in one health zone in Alberta, Canada led to a significant step reduction in the ordering of the BUN test on hospital admission., Conclusions: Redesigning the standard medicine admission order form-laboratory request section can have a beneficial effect on the reduction in BUN ordering altering physician ordering patterns and behaviour., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2021. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Background: A high-quality colonoscopy bowel prep is vital to completing the procedure. Adequate inpatient bowel preparation has been consistently difficult to achieve because of multiple factors. Incomplete bowel prep can lead to repeated colonoscopies, poor patient experience, increased costs, and prolonged hospitalization. This study aimed to develop patient-centered interventions to optimize bowel prep for inpatients undergoing colonoscopy., Methods: The Model for Improvement and Donabedian frameworks guided this project. An interdisciplinary team compiled quality improvement tools that identified areas for improvement. Interventions development included a nursing tip sheet for troubleshooting symptoms, a standardized order label and a patient educational placemat. Plan-Do-Study-Act (PDSA) cycles were carried out to test and analyze the effects of the interventions. The project aim was a 30% reduction in incomplete inpatient colonoscopies from poor bowel prep. Process measures included the number of colonoscopy split prep order labels, and placemats used. The balancing measure was the number of repeat colonoscopies., Results: Prior to the intervention, 44% (44/99) of inpatient colonoscopies had poor bowel prep resulting in 10 repeat procedures (10%). Post intervention, 60% (28/47) of the colonoscopies used the standardized label, 66% of physician orders used 2-L split prep, and 80% of patients were provided with the educational placemat. Of the 47 colonoscopies audited post intervention, there was a significant decrease in poor prep (27.7% [13/47], P=0.038) for colonoscopies. The percentage of repeated colonoscopies decreased to 4% (2/47)., Conclusion: Developing simple and easy-to-use patient-centered interventions can effectively improve colonoscopy preparation for hospitalized patients., Competing Interests: Conflict of Interest: None, (Copyright: © Hellenic Society of Gastroenterology.)

Objectives: We aimed to assess the perception of psychodermatology, practice patterns, and challenges reported by Canadian dermatologists., Methods: We designed an online questionnaire based on previous literature, including questions about practitioners' perceptions, practice patterns, training, and challenges in psychodermatology. We solicited their opinions on desired training, research needs, and clinical approach recommendations. Our survey was distributed nationally by the Canadian Dermatology Association (CDA)., Results: Of the total of 78 participating dermatologists, >75% reported treating patients with psychodermatological conditions, with higher frequencies of secondary than primary psychodermatological conditions. While practitioners had some confidence in their understanding of psychodermatology (median = 4 on a 5-point scale), their comfort levels to approach these patients were lower (median = 3), and their confidence in prescribing psychotropic medication was markedly low (median = 2). A total of 50% reported that a "multidisciplinary approach" would be best for these patients. Poor access to psychiatry was the most reported (26.9%) challenge, together with time constraints, lack of training, poor communication with patients, and lack of patient insight and resources. While 46.2% reported having never participated in psychodermatology training, 55.1% expressed interest in doing so., Conclusion: We identified several challenges with knowledge, awareness, and healthcare delivery in psychodermatological practice in Canada. Increasing dermatologists' access to psychiatric consultations/services, a multidisciplinary approach with dermatologists and psychiatrists co-providing care, and more specialized training in this area are recommended to narrow the identified gaps.

Background and Purpose: Quality Improvement (QI) science is a burgeoning component of healthcare systems and the practice of pharmacy. There is limited published literature on the implementation of QI training in undergraduate pharmacy education. We describe the development and implementation of QI training in an experiential course., Educational Activity and Setting: A QI curriculum was developed for an existing eight-week pharmacy practice elective experience for year four pharmacy students. The curriculum was divided into two, four-week blocks, combining didactic instruction and applied learning activities. A partnership between university departments and a local health organization provided a QI preceptor along with an interdisciplinary clinical team in an acute care setting to improve clinical services., Findings: Six students have completed the elective along with completing three QI projects. The Kirkpatrick 4 level model guided course evaluation. A curriculum strength was no additional cost or educational burden on the faculty of pharmacy and pharmaceutical sciences. Students recognized benefit from (1) course flexibility and independence to learn, (2) clear expectations and weekly guidance from the preceptor, (3) alignment of weekly readings and real-time application of QI concepts, and (4) the Institute for Healthcare Improvement education modules. Reducing the number of assigned tasks per week and improving availability of the clinical QI team to support student learning were identified as areas for improvement., Summary: This elective course demonstrated a starting point from which QI education can be formally included in pharmacy undergraduate education., Competing Interests: Declaration of Competing Interest None, (Copyright © 2020 Elsevier Inc. All rights reserved.)

Background: Prior to 2017, internal medicine (IM) residents at the University of Alberta did not have a standardised quality improvement (QI) educational curriculum. Our goal was to use QI principles to develop a resident sustained curriculum using the Evidence-based Practice for Improving Quality (EPIQ) training course., Methods: Three one-year Plan-Do-Study-Act (PDSA) cycles were conducted. The EPIQ course was delivered to postgraduate year (PGY) 1-3 residents (n=110, PDSA 1) in 2017, PGY-1 residents (n=27, PDSA 2) in 2018 and PGY-1 residents (n=28, PDSA 3) in 2019. Trained residents were recruited as facilitators for PDSA 2 and 3. Residents worked through potential QI projects that were later presented for evaluation. Precourse and postcourse surveys and tests were conducted to assess knowledge acquisition and curriculum satisfaction. Process, outcome and balancing measures were also evaluated., Results: In PDSA 1, 98% felt they had acquired understanding of QI principles (56% increase), 94% of PGY-2 and PGY-3 residents preferred this QI curriculum compared with previous training, and 65% of residents expressed interest in pursuing a QI project (15% increase). In PDSA 2, tests scores of QI principles improved from 77.6% to 80%, and 40% of residents expressed interest in becoming a course facilitator. In PDSA 3, self-rated confidence with QI methodology improved from 53% to 75%. A total of 165 residents completed EPIQ training and 11 residents became course facilitators., Conclusions: Having a structured QI curriculum and working through practical QI projects provided valuable QI training for residents. Feedback was positive, and with each PDSA cycle there was increased resident interest in QI. Developing this curriculum using validated QI tools highlighted areas of change opportunity thereby enhancing acceptance. As more cycles of EPIQ are delivered and more residents become facilitators, it is our aim to have this curriculum sustained by future residents., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2021. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Background: Psychodermatologic disorders are difficult to identify and treat. Knowledge about the prevalence of these conditions in dermatological practice in Canada is scarce. This hampers our ability to address potential gaps and establish optimal care pathways., Objectives: To provide an estimate of the frequencies of psychodermatologic conditions in dermatological practice in Alberta, Canada., Methods: Two administrative provincial databases were used to estimate the prevalence of potential psychodermatological conditions in Alberta from 2014 to 2018. Province-wide dermatology claims data were examined to extract relevant International Classification of Disease codes as available. Claims were linked with pharmacy dispensation data to identify patients who received at least 1 psychoactive medication within 90 days of the dermatology claim., Results: Of 243 963 patients identified, 28.6% had received at least 1 psychotropic medication (mean age: 47.9 years; 67.5% female). Rates of concurrent psychotropic medications were highest for pruritus and related conditions (46.7%), followed by urticaria (44.5%) and hyperhidrosis (32.8%). Among patients with psychotropic medications, rates of antidepressants were highest (56.3%), followed by anxiolytics (37.1%). Across billing codes, besides hyperhidrosis (71.2%), diseases of hair (61.4%) and psoriasis (59.1%) had the highest rates of antidepressant dispensations. Patients with atopic dermatitis had the highest rates for anxiolytic prescriptions (54.3%)., Conclusion: In a 5-year window, more than a quarter of the identified dermatology patients in Alberta received at least 1 psychotropic medication, pointing to high rates of potential psychodermatologic conditions and/or concurrent mental health issues in dermatology. Diagnostic and care pathways should include a multidisciplinary approach to better identify and treat these conditions.

Competing Interests: Competing interests: None declared.

Background: Appropriate, timely trauma team activation (TTA) can directly affect outcomes for patients with trauma. A review of quality-performance indicators at our Canadian level 1 trauma centre showed a high level of undertriage, with TTA compliance rates less than 60% for major trauma. A quality-improvement project was undertaken, targeting a sustained goal of at least 90% TTA compliance based on Accreditation Canada guidelines., Methods: Quality-improvement action followed a well-defined process. Baseline data collection was performed, and, in keeping with the Donabedian approach, we brought together stakeholders to collectively review and understand the reasons behind poor TTA compliance; and root-cause analysis. This was followed by rapid change cycles that focused on structure and processes with ongoing audits to support and sustain change., Results: Trauma team activation compliance improved from 58.8% to more than 90% over 2 years. Quality indicators showed a statistically significant reduction in the time to computed tomography scanner, time in the acute care region of the emergency department and total time in the emergency department, with improved TTA compliance., Conclusion: Compliance with TTA protocols improved to more than 90% over a 2-year period, which shows the benefit of having a clearly outlined qualityimprovement process. This well-defined quality-improvement method provides a framework for use by other institutions that seek to improve their processes of trauma care, including activation rates., Competing Interests: V. Fawcett and S. Widder are members of the CJS editorial board, but they were not involved in the review or acceptance of this manuscript. No other competing interests declared., (© 2019 Joule Inc. or its licensors)

Benzodiazepines are recognised as being potentially inappropriate medications for seniors due to their considerable side-effect profile, yet they are commonly prescribed and infrequently discontinued (deprescribed). The study's primary objective was the deprescription or the dose reduction of benzodiazepines among newly hospitalised seniors using a patient education intervention. A 3-month duration quality improvement study based on the plan-do-study-act model was conducted across two units (3C and 4D) in the Glenrose Rehabilitation Hospital to improve benzodiazepine deprescribing among newly admitted seniors (65 years or older) who were using benzodiazepines. The primary outcome measure was the number of eligible patients who had benzodiazepine deprescribing initiated. A patient education intervention comprising a structured medication review, written patient education (the Eliminating Medications Through Patient Ownership of End Results (EMPOWER) brochure) and at least one brief supportive counselling session by the clinical pharmacist or physician was applied to all eligible patients. All 12 eligible patients consented to benzodiazepine deprescribing; however, only 11 of them (92%) initiated benzodiazepine deprescribing. Six of the 11 patients (55%) had their benzodiazepines discontinued, with the 5 remaining patients (45%) achieving greater than 50% dosage reduction. Seven patients (64%) experienced side effects during the deprescribing process, with over half (57%, n=4) of these seven patients experiencing worsening anxiety symptoms. Five of the 11 patients (45%) required benzodiazepine substitute medications. The use of a structured patient education intervention involving the use of a structured medication review, written patient education material and one-on-one patient counselling can promote benzodiazepine deprescribing. Although worsening anxiety was frequently observed, this was easily managed by the substitution of a more appropriate and clinically indicated medication, which was well tolerated and acceptable by all of our participants. Targeted screening for the presence of anxiety would help to guide the deprescribing process and the need for medication substitution., Competing Interests: Competing interests: None declared.

Ensuring adequate vascular access in major trauma patients prior to decompensative physiological processes is crucial to patient outcomes. Most protocols suggest achieving two 18-gauge or larger intravenous lines immediately in patients with major trauma. We discuss a quality improvement approach to ensure that >90% of patients with major trauma (as defined by an injury severity score ≥12) at a level one trauma centre receive timely and adequate fluid access. Applying Donabedian principles for process improvement, we used the Alberta Trauma Registry to perform a 4-month chart audit on patients with major trauma at the University of Alberta Hospital. Background data were supported with a formal root cause analysis to outline the problems and generate plan, do, study and act (PDSA) rapid change cycles. These PDSA cycles were then implemented over the course of 2 months to alter system and personnel barriers to care, thereby ensuring that patients with major trauma received adequate vascular access for fluid resuscitation. This was followed by a 6-month sustainability assessment. The percentage of patients with major trauma who received adequate fluid access went from a mean of 55.5% to >90% in 2 months and was sustained at or greater than 90% for 6 consecutive months. The formal application of quality improvement processes is uncommon in trauma care but is much needed to ensure success and sustainability of quality initiatives. Planning including engagement and prechange awareness is crucial to staff engagement, change, and sustainment. Formal quality improvement and change management techniques can elicit rapid and sustainable changes in trauma care. We provide a framework for change to increase compliance with fluid access in patients with major trauma., Competing Interests: Competing interests: None declared.

Objective: To develop a standardized process for reviewing daily patient lists and identifying potential risks of misidentification. Our goal was to develop a proactive approach to identify and eliminate risks of patient misidentification., Methods: Assessment of current patient identification practices took place over a period of 4 weeks. Using a process map, a patient survey was developed to determine the encounter points when patient identification was confirmed. This information was used to develop a standardized protocol for review of daily appointment lists., Results: Review of daily appointment lists was completed to identify potential similar/same name risks. A standardized manual process of chart review, flagging, and tracking was developed., Conclusions: The name alert process resulted in a simple manual process for identifying which patients have a higher name risk and allowed care providers to take preventative action to decrease potential risk of incorrect diagnostic testing, procedure, or medication administration., (Copyright © 2017 Canadian Ophthalmological Society. Published by Elsevier Inc. All rights reserved.)

Background: Ophthalmology residents on call at the Royal Alexandra Hospital identified workplace disorganization and lack of standardization in emergency eye examination rooms as an impediment to efficient patient treatment., Objective: The aim of the study was to use the "6S Lean" model to improve workflow in eye examination rooms at the Royal Alexandra Hospital., Methods: With the assistance of quality improvement consultants, the "6S Lean" model was applied to the current operation of the emergency eye clinic examination rooms. This model, considering 8 waste categories, was then used to recommend and implement changes to the examination rooms and to workplace protocols to enhance efficiency and safety., Results: Eye examination rooms were improved with regards to setup, organization of supplies, inventory control, and maintenance. All targets were achieved, and the 5S audit checklist score increased by 33 points from 44 to 77., Conclusions: Implementation of the 6S methodology is a simple approach that removes inefficiencies from the workplace. The ophthalmology clinic removed waste from all 8 waste categories, increased audit results, mitigated patient and resident safety risks, and ultimately redirected resident time back to patient care delivery., (Copyright © 2017 Canadian Ophthalmological Society. Published by Elsevier Inc. All rights reserved.)

Background: Cancer is a major disease worldwide, and many patients use complementary and alternative treatments. The purpose of this study was to identify the herbal remedies and functional foods used as complementary medicine by prostate, breast and colorectal cancer patients at speciality care facilities in Trinidad. We also sought to determine how patients rated the efficacy of these modalities compared with conventional treatment., Methods: A descriptive, cross-sectional survey was conducted using an interviewer-administered pilot-tested de novo questionnaire during the period June to August 2012 at two speciality treatment centres on the island. Data was analysed using χ 2 analyses., Results: Among the 150 patients who reported use of herbal remedies/functional foods, soursop (Annona muricata L.) was the most popular; with 80.7 % using the leaves, bark, fruit and seeds on a regular basis. Other common herbal remedies/functional foods included wheatgrass (Triticum aestivum L.), saffron (Crocus sativus L.) and Aloe vera (L.) Burm. f. The most commonly used functional foods were beetroot (Beta vulgaris L.), carrots (Daucus carata L.) and papaya (Carica papaya L.) used by 43.3 % of patients; and these were mostly blended as a mixture. Herbal remedies and functional foods were used on a daily basis and patients believed that this modality was equally (32.0 %) or more efficacious (14.7 %) than conventional treatment., Conclusions: This survey identified the most common herbal remedies and functional foods used among prostate, breast and colorectal cancer patients in Trinidad. Although functional foods rarely pose a problem, herbs may interact with conventional chemotherapy and physicians need to inform patients regarding probable herb-drug interactions.

The increased transmission and geographic spread of dengue fever (DF) and its most severe presentations, dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS), make it one of the most important mosquito-borne viral disease of humans. Four distinct serotypes of dengue viruses are transmitted to humans through the bites of the mosquitoes. Currently there is no vaccine or antiviral drug against DV infections. Cross-protection between dengue virus serotypes is limited and antibody dependent enhancement (ADE) contributes significantly to the severity of the disease. The major challenge is to induce a broad durable immune response against all four serotypes of dengue virus simultaneously while avoiding the possible exacerbation of risk of developing the severe forms of disease through incomplete or modified responses. In order to address this worldwide concern, we present a synthetic consensus (SynCon) human codon optimized DNA vaccine that elicits immunity against all four dengue serotypes. We cloned consensus DIII domain of E protein from all serotypes and expressed them as a single open reading frame in a mammalian expression vector, called pDV-U-DIII (dengue-vaccine universal). In mice, this dengue-universal construct elicits significant level of anti-DIII antibody that neutralizes all four dengue subtypes and prevents cell death induced by dengue infection. This is the first SynCon DNA vaccine that provides tetravalent immunity against all four serotypes of dengue virus.

Dengue virus serotype 3 (DENV-3) was associated with severe dengue epidemics in Thailand during 1973-1999. We studied Thai DENV-3 viruses isolated from hospitalized children in Bangkok with differing disease severity during that period. Viruses were sequenced at their 5' and 3' untranslated regions (UTRs), which are regions that play a pivotal role in viral replication. Our results indicated that the primary sequences as well as the secondary structures at both ends of Thai DENV-3 viruses were highly conserved over almost three decades. We found nucleotide insertions and deletions at the variable region (VR) that is located just downstream of the nonstructural protein 5 (NS5) stop codon among these viruses. The phylogenetic tree derived from the size heterogeneity of VR in the 3' UTR divided DENV-3 into four genotypes, and Thai DENV-3 viruses in this study belonged to genotype II. The replication efficiency of the candidate viruses with different lengths at the VR were assessed in the mosquito (C6/36) and human (HepG2) cell lines. Our results show that the viruses with nucleotide insertions at VR replicated better than the virus that contained deletions. Our findings indicate that Thai DENV-3 demonstrated a remarkable conservation of nucleotides over 28 years. Correlation with disease severity suggests that both primary sequences and secondary structures of the 3' UTR do not appear correlated with disease severity in humans.

The Japanese encephalitis virus (JEV) and West Nile virus (WNV) are responsible for a large proportion of viral encephalitis in humans. Currently, there is no FDA approved specific treatment for either, though there are attempts to develop vaccines against both viruses. In this study, we proposed novel genetically engineered DNA vaccines against these two neurotrophic flaviviruses. The structural domain III (DIII) of E protein from these viruses is reported to carry dominant epitopes that induce neutralizing antibodies. Therefore we created consensus sequence of DIII domain across numerous strains of JEV and WNV. Based on the consensus amino acid sequence, synthetic codon and RNA optimized DIII-expressing DNA vaccine constructs with an efficient leader sequence were synthesized for immunization studies. In addition, we also constructed a genetically engineered IL15 DNA vaccine molecular adjuvant for co-stimulating the immune response against DIII clones. Vaccine constructs were delivered into BALB/C mice intramuscularly followed by electroporation using the CELLECTRA in vivo electroporator. We have observed that the combined delivery of both WNV DIII and IL15-ECRO DNA vaccine constructs resulted in not only the highest level of antibody against DIII, but also enhanced cross reactivity with two other antigens tested. Also, coimmunization with IL15 plasmid further increased the immune response by four- to five-fold. Importantly, we have shown that IL15 coimmunization adjuvanted humoral responses against DIII antigens by elevating the level of antibody secreting B cells. Such a DNA vaccine approach may better help to control potential travel related infectious agents such as JEV.

Human papillomavirus type 16 (HPV-16) infection is associated with a majority of cervical cancers and a significant proportion of head and neck cancers. Here, we describe a novel-engineered DNA vaccine that encodes a HPV-16 consensus E6/E7 fusion gene (pConE6E7) with the goal of increasing its antitumor cellular immunity. Compared to an early stage HPV-16 E7 DNA vaccine (pE7), this construct was up to five times more potent in driving E7-specific cellular immune responses. Prophylactic administration of this vaccine resulted in 100% protection against HPV E6 and E7-expressing tumors. Therapeutic studies indicated that vaccination with pConE6E7 prevented or delayed the growth of tumors. Moreover, immunization with pConE6E7 could also partially overcome immune tolerance in E6/E7 transgenic mice. Such DNA immunogens are interesting candidates for further study to investigate mechanisms of tumor immune rejection in vivo.

An important goal for human immunodeficiency virus (HIV) vaccines is to develop immunogens that induce broader and more potent cellular immune responses. In this study of DNA vaccine potency, we constructed a novel subtype B env gene (EY2E1-B) with the goal of increasing vaccine antigen immune potency. The vaccine cassette was designed based on subtype B-specific consensus sequence with several modifications, including codon optimization, RNA optimization, the addition of a Kozak sequence, and a substituted immunoglobulin E leader sequence. The V1 and V2 loops were shortened and the cytoplasmic tail was truncated to prevent envelope recycling. Three different strains of mice (BALB/c, C57BL/6, and HLA-A2 transgenic mice) were immunized three times with pEY2E1-B or the primary DNA immunogen pEK2P-B alone. The analysis of specific antibody responses suggested that EY2E1-B could induce a moderate subtype B-specific antibody response. Moreover, this construct was up to four times more potent at driving cellular immune responses. Epitope mapping results indicated that there is an increase in the breadth and magnitude of cross-reactive cellular responses induced by the EY2E1-B immunogen. These properties suggest that such a synthetic immunogen deserves further examination for its potential to serve as a component antigen in an HIV vaccine cocktail.

DNA vaccines expressing the envelope (env) of the human immunodeficiency virus type 1 (HIV-1) have been relatively ineffective at generating strong immune responses. In this study, we described the development of a recombinant plasmid DNA (pEK2P-B) expressing an engineered codon-optimized envelope gp140 gene of primary (nonrecombinant) HIV-1 subtype B isolate 6101. Codon usage and RNA optimization of HIV-1 structural genes has been shown to increase protein expression in vitro as well as in the context of DNA vaccines in vivo. To further increase the expression, a synthetic IgE leader with kozak sequences were fused into the env gene. The cytoplasmic tail of the gene was also truncated to prevent recycling. The expression of env by the recombinant pEK2P-B was evaluated using T7 coupled transcription/translation. The construct demonstrated high expression of the HIV-1 env gene in eukaryotic cells as demonstrated in transfected 293-T and RD cells. Immunogenicity of pEK2P-B was evaluated in mice using IFN-gamma ELISpot assay, and the construct was found to be highly immunogenic and crossreactive with HIV-1 clade C env peptides. Three immunodominant peptides were also mapped out. Furthermore, by performing a CFSE flow cytometry-based proliferation assay, 2.4 and 1.5% proliferation was observed in CD4+, CD8+, and CCR+ memory T cells, respectively. Therefore, this engineered synthetic optimized env DNA vaccine may be useful in DNA vaccine and other studies of HIV-1 immunogenicity.

The West Nile Virus (WNV) non-structural proteins 2B and 3 (NS2B-NS3) constitute the proteolytic complex that mediates the cleavage and processing of the viral polyprotein. NS3 recruits NS2B and NS5 proteins to direct protease and replication activities. In an effort to investigate the biology of the viral protease, we cloned cDNA encoding the NS2B-NS3 proteolytic complex from brain tissue of a WNV-infected dead crow, collected from the Lower Merion area (Merion strain). Expression of the NS2B-NS3 gene cassette induced apoptosis within 48 h of transfection. Electron microscopic analysis of NS2B-NS3-transfected cells revealed ultra-structural changes that are typical of apoptotic cells including membrane blebbing, nuclear disintegration and cytoplasmic vacuolations. The role of NS3 or NS2B in contributing to host cell apoptosis was examined. NS3 alone triggers the apoptotic pathways involving caspases-8 and -3. Experimental results from the use of caspase-specific inhibitors and caspase-8 siRNA demonstrated that the activation of caspase-8 was essential to initiate apoptotic signaling in NS3-expressing cells. Downstream of caspase-3 activation, we observed nuclear membrane ruptures and cleavage of the DNA-repair enzyme, PARP in NS3-expressing cells. Nuclear herniations due to NS3 expression were absent in the cells treated with a caspase-3 inhibitor. Expression of protease and helicase domains themselves was sufficient to trigger apoptosis generating insight into the apoptotic pathways triggered by NS3 from WNV.

The authors report a new strain of West Nile virus (WNV) with the expression analysis of its individual open reading frames. Since its sudden appearance in the summer of 1999 in New York City, the virus has spread rapidly across the continental United States into Canada and Mexico. Besides, its rapid transmission by various vectors, the spread of this virus through organ transplantation, blood transfusion, and mother-child transmission through breast milk is of concern. In order to understand molecular variations of WNV in North America and to generate new tools for understanding WNV biology, a complete clone of WNV has been constructed. Investigations so far have focused only on half of its genes products and a detailed molecular and cell biological aspects on all of WNV gene have yet to be clearly established. The open reading frames of WNV were recovered through an reverse transcriptase-polymerase chain reaction (RT-PCR)-PCR using brain tissue from a dead crow collected in Merion, PA, and cloned into a mammalian expression vector. The deduced amino acid sequences of individual open reading frames were analyzed to determine various structural motifs and functional domains. Expression analysis shows that in neuronal cells, C, NS1, and NS5 proteins are nuclear localized whereas the rest of the antigens are confined to the cytoplasm when they are expressed in the absence of other viral antigens. This is the first report that provides an expression analysis as well as intracellular distribution pattern for all of WNV gene products, cloned from an infected bird. Evolutionary analysis of Merion strain sequences indicates that this strain is distinct phylogenetically from the previously reported WNV strains.

DNA vaccines are a promising technology for the induction of Ag-specific immune responses, and much recent attention has gone into improving their immune potency. In this study we test the feasibility of delivering a plasmid encoding IL-15 as a DNA vaccine adjuvant for the induction of improved Ag-specific CD8(+) T cellular immune responses. Because native IL-15 is poorly expressed, we used PCR-based strategies to develop an optimized construct that expresses 80-fold higher than the native IL-15 construct. Using a DNA vaccination model, we determined that immunization with optimized IL-15 in combination with HIV-1gag DNA constructs resulted in a significant enhancement of Ag-specific CD8(+) T cell proliferation and IFN-gamma secretion, and strong induction of long-lived CD8(+) T cell responses. In an influenza DNA vaccine model, coimmunization with plasmid expressing influenza A PR8/34 hemagglutinin with the optimized IL-15 plasmid generated improved long term CD8(+) T cellular immunity and protected the mice against a lethal mucosal challenge with influenza virus. Because we observed that IL-15 appeared to mostly adjuvant CD8(+) T cell function, we show that in the partial, but not total, absence of CD4(+) T cell help, plasmid-delivered IL-15 could restore CD8 secondary immune responses to an antigenic DNA plasmid, supporting the idea that the effects of IL-15 on CD8(+) T cell expansion require the presence of low levels of CD4 T cells. These data suggest a role for enhanced plasmid IL-15 as a candidate adjuvant for vaccine or immunotherapeutic studies.

Objective: The food consumption and serum nutritional status of people living in the Kathmandu valley, Nepal, were examined to obtain actual data for comparison with our previous findings., Methods: A dietary survey of 45 males and 60 females was carried out in March 1997 by the 24-hr dietary recall method and nutrient intake was calculated from food tables of India and Japan. Fasting venous blood samples were obtained and serum biochemical parameters were measured using clinical kits., Results: The mean body mass index values was at about the same level in both sexes, although the mean percentage body fat of females was higher than that of males, and vice versa for packed red cell volume. The main foods consumed by both sexes, were rice, potatoes, meats, milk & dairy products and vegetables, with a difference in the amounts consumed. Females did not drink alcoholic beverages. The mean daily intakes of energy, protein, lipids, iron and vitamin B group for the males were higher than those for the females, while those of vitamins A and C for the males were lower than those for the females. The mean values of serum biochemical parameters for both sexes were generally at the normal levels, but those of ALT and TG were at the higher end of the normal range. Differences of correlation between food groups and between serum parameters were observed depending on the sex, however, no clear relationship between food and nutrient intake and serum biochemical parameters were observed., Conclusions: The present food intake study revealed that the amounts of food consumption for both sexes, especially for the females, were mostly insufficient although the serum parameters were at the normal levels. The energy intake of both sexes was lower than that of estimated requirements and those in Terai region. The relatively high serum TG level of the subjects may be due to the consumption of large amounts of cereals containing much carbohydrate. Our findings suggested a marked influence on food consumption by food price and income in spite of the easier food availability in the city, and also lack of knowledge about nutrients and health, thus there is need for improvement of the nutritional status of this group of people.

Objective: The present study examined the levels of serum α-Tocopherol (Toc), retinol (Ret), cholesterol (Chol) and triglycerides (TG), and their correlations in the sera of people in Nepal., Methods: The survey was conducted on the general populace in the agricultural Terai region in southern Nepal. The study population consisted of 93 males and 83 females aged 10-68 years. Serum Toc and Ret were measured by high-performance liquid chromatography., Results: No significant differences were observed between the genders for the average of total Chol (T-Chol) (140 and 145 mg/100 ml, respectively), HDL-C (45 and 47 mg/100 ml), LDL-C (94 and 97 mg/100 ml), and TG (106 and 110 mg/100 ml), and the ratio of LDL/HDL (2.16). The levels of mean Toc (4.32 and 4.27 μg/ml) were about the same for both genders, while the mean Ret levels were significantly higher for males (624 ng/ml) than for females (535 ng/ml) (p<0.001). A direct relationship was found between the levels of Toc and Ret (r=0.46, p<0.001 and r=0.28, p<0.05 for males and females, respectively). Serum levels of Toc and Ret were positively related to the levels of Chol (r=0.48 and r=0.58, p<0.001 for males and r=0.49, p<0.01 and r-0.28, p<0.05 for females, respectively). The ratio of Toc/TG normalized to serum TG was directly correlated to the ratio of Ret/TG (r=0.79 for males, and r=0.72 for females, p<0.001, respectively) and the ratios of Toc/TG and Ret/TG were negatively related to the LDL/HDL levels (r=-0.49 and r=-0.43, for males, and r=-0.46 and r=-0.57 for females, p<0.001, respectively)., Conclusion: The levels of Toc and Ret were low in the sera of people living in the southern rural Terai region in Nepal, and it was found that lower levels of Toc and Ret normalized to TG increased the ratio of LDL/HDL. These results suggest that greater intake of foods rich in Toc and Ret should be encouraged to reduce the erisk of coronary heart disease.

About 47% of Nepal's total population is living in Terai region and 90% of them are relying on groundwater as their major source of drinking water. About 200,000 shallow tubewells have been installed by different agencies in 20 Terai districts, serving 11 million people. Recently, arsenic contamination of groundwater has been recognized as a public health problem in Nepal. This has sensitized government, national and international nongovernment organizations working on water quality sector to carry out water quality assessment for arsenic in the affected communities. So far, 15,000 tubewells has been tested where 23% samples exceeded World Health Organization guideline value of 10 microg/L and 5% exceeded "Nepal Interim Arsenic Guideline" of 50 microg/L. It is estimated that around 0.5 million people in Terai are living at risk of arsenic poisoning (>50 microg/L). Some recent studies have reported the prevalence of dermatosis related to arsenicosis from 1.3 to 5.1% and the accumulation of arsenic in biological samples like hair and nail much higher than the acceptable level. Though some steps are being taken by government and private organizations to combat the problem, it has not been able to cover all the affected communities. Nepal still needs more research work on arsenic occurrence and effects and mitigation programs simultaneously.

Human immunodeficiency virus, type 1 (HIV-1) vpr is a highly conserved gene among lentiviruses. The diverse functions of Vpr support interactions of this HIV accessory protein with host cell partners of important pathways. hVIP/mov34 (human Vpr Interacting Protein) is one of these identified Vpr ligands. hVIP is a 34-kDa member of the eIF3 family that is vital for early embryonic development in transgenic mice and important in cell cycle regulation. Its interaction with Vpr, however, is not yet clearly defined. Therefore, we constructed a panel of deletion mutants of this cytoplasmic cellular ligand to map the protein domain that mediates its interaction with Vpr. We observed that the carboxyl-terminal region of hVIP is critical for its interaction with Vpr. In the absence of Vpr or HIV infection, full-length hVIP is expressed in the cytoplasm. The cytoplasmic localization pattern of full-length hVIP protein, however, is shifted to a clear nuclear localization pattern in cells expressing both hVIP and Vpr. In contrast, Vpr did not alter the localization pattern of hVIP mutants, which have their carboxyl-terminal domain deleted. The movement of hVIP supported prior work that suggested that Vpr triggers activation of the GR receptor complex. In fact, we also observed that dexamethasone moves hVIP into the nucleus and that glucocorticoid antagonists inhibit this effect. Interestingly, the expression of an hVIP carboxyl-terminal mutant, which is not responsive to Vpr, is also not responsive to dexamethasone. These data illustrate that the carboxyl-terminal domain of hVIP is critical for mediating hVIP-Vpr interaction as well as for its glucocorticoid response. These results support the view that hVIP is a member of the complex array of nucleocytoplasmic shuttling proteins that are regulated by HIV infection and glucocorticoids.

West Nile virus (WNV) is a member of the Flaviviridae family of vector-borne pathogens. Clinical signs of WNV infection include neurologic symptoms, limb weakness, and encephalitis, which can result in paralysis or death. We report that the WNV-capsid by itself induces rapid nuclear condensation and cell death in tissue culture. Apoptosis is induced through the mitochondrial pathway resulting in caspase-9 activation and downstream caspase-3 activation. Capsid gene delivery into the striatum of mouse brain or interskeletal muscle resulted in cell death and inflammation, likely through capsid-induced apoptosis in vivo. These studies demonstrate that the capsid protein of WNV may be responsible for aspects of viral pathogenesis through induction of the apoptotic cascade.

Objective: This study examined the levels of serum protein fractionation, immunoglobulin (Ig) and antistreptolysin O (ASO) of people (91 males and 84 females aged 10-68 years) living in the Terai region of southern Nepal, as there has previously been no information available about them., Methods: Blood samples were collected early in the morning after overnight fasting. Serum protein fractionation was carried out by cellulose-acetate electrophoresis. IgG, IgA, IgM and ASO were measured by immuno-turbidimetry with clinical kits., Results: The mean proportion of albumin (Alb) was rather low due to increased globulin (Glb). The Alb level of males was significantly higher than that of females, while the male γ-Glb level was significantly lower than that of females. The mean values of IgG, IgM and ASO for males were lower than those values for females, although the difference was significant only for the IgM value. The mean values of IgM for female age groups of 30-39 years or less were significantly higher than those for the corresponding male age groups. Age correlated positively with IgA, and negatively with ASO in both sexes. γ-Glb and IgG correlated significantly with TP, AlB, α(1)-Glb and IgA in both sexes. ASO correlated with β-Glb and IgA in males, and with γ-Glb and IgG in females., Conclusions: The fact that the level of γ-Glb, a major component of serum globulin, was high suggested exposure to a highly bacterial and viral environment. These results point to the need to prevent infectious diseases as well as improve their nutritional status, especially for children and young adults.