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Background: Influenza A viruses pose a significant public health threat globally and are characterized by rapid evolution of the hemagglutinin (HA) gene causing seasonal epidemics. The aim of this study was to investigate the evolutionary dynamics of A(H3N2) circulating in Bhutan during 2022 and 2023., Methods: We analysed 166 whole-genome sequences of influenza A(H3N2) from Bhutan, obtained from the GISAID database. We employed a Bayesian Markov Chain Monte Carlo (MCMC) framework, with a curated global dataset of HA sequences from regions with significant migration links to Bhutan. Phylogenetic, temporal, and phylogeographic analyses were conducted to elucidate the evolutionary dynamics and spatial dissemination of the viruses., Results: Our phylogenetic analysis identified the circulation of influenza A(H3N2) Clade 3C.2a1b.2a.2 in Bhutan during 2022 and 2023, with viruses further classified into three subclades: 2a.3 (39/166), 2a.3a.1 (58/166) and 2a.3b (69/166). The TMRCA estimates suggest that these viral lineages originated approximately 1.93 years prior to their detection. Phylogeographic analysis indicates introductions from the United States in 2022 and Australia in 2023. The mean evolutionary rate across all gene segments was calculated to be 4.42 × 10 -3 substitutions per site per year (95% HPD: 3.19 × 10 -3 to 5.84 × 10 -3 ), with evidence of purifying selection and limited genetic diversity. Furthermore, reassortment events were rare, with an estimated rate of 0.045 events per lineage per year., Conclusion: Our findings show that primary forces shaping the local evolution of the influenza A(H3N2) in Bhutan are largely stochastic, with only sporadic instances of adaptive change, and thus underscore the importance of continuous surveillance to mitigate the impact of evolving strains., (© 2024 The Author(s). Influenza and Other Respiratory Viruses published by John Wiley & Sons Ltd.)

Introduction: In any healthcare setting, cardiopulmonary resuscitations (CPRs) stand out as demanding and chaotic resuscitation endeavours. Emergency departments (EDs) witness a significant volume of CPRs. Given the critical nature of CPR, content knowledge and procedural skills alone fall short in delivering optimal care. Effective teamwork, complemented by a well-coordinated response, is imperative for achieving favourable patient outcomes. A survey conducted in our ED highlighted that while the majority of staff acknowledged the significance of teamwork in CPR and were aware of the whiteboard for assigning team roles, only 19% were familiar with their individual roles during CPR., Methods: To address this gap, our project aimed to increasing the role delegation for CPR from 19% to 80% within 2 months. We formed an interprofessional team and implemented strategies through four plan-do-study-act cycles. Interventions encompassed increasing sensitisation, creating a simplified format for assigning team roles and entrusting the nursing team leader of each shift with the responsibility of role assignment for accountability. The sharing of progress charts for acknowledgment served as a motivating factor, leading to sustained adherence to the project goals without necessitating reminders in the final two weeks., Results: This project proved to be highly successful as our process indicator steadily increased and remained above the target for 4 consecutive weeks., Conclusion: Our results underscore the importance of patience and teamwork in achieving project objectives. It serves as a good example of the efficiency of simple and cost-effective interventions, one that can be replicated and implemented in other EDs., 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: Influenza (Flu) causes considerable morbidity and mortality globally, and in Bhutan, Flu viruses are a leading cause of acute respiratory infection and cause outbreaks during Flu seasons. In this study, we aim to analyze the epidemiology and the genetic characterization of Flu viruses circulated in Bhutan in 2022., Method: Respiratory specimens were collected from patients who meet the case definition for influenza-like illness (ILI) and severe acute respiratory infection (SARI) from sentinel sites. Specimens were tested for Flu and SARS-CoV-2 viruses by RT-PCR using the Multiplex Assay. Selected positive specimens were utilized for Flu viral genome sequencing by next-generation sequencing. Descriptive analysis was performed on patient demographics to see the proportion of Flu-associated ILI and SARI. All data were analyzed using Epi Info7 and QGIS 3.16 software., Result: A weekly average of 16.2 ILI cases per 1000 outpatient visits and 18 SARI cases per 1000 admitted cases were reported in 2022. The median age among ILI was 12 years (IQR: 5-28) and SARI was 6.2 (IQR: 2.5-15) years. Flu A(H3N2) (70.2%) subtype was the most predominant circulating strain. Flu A(H1N1)pdm09 and Flu B viruses belonged to subclades that were mismatched to the vaccine strains recommended for the 2021-2022 season but matched the vaccine strain for the 2022-2023 season with vaccine efficacy 85.14% and 88.07% respectively. Flu A(H3N2) virus belonged to two subclades which differed from the vaccine strains recommended in both the 2021-2022 and 2022-2023 seasons with vaccine efficacy 68.28%., Conclusion: Flu virus positivity rates were substantially elevated during the Flu season in 2022 compared to 2021. Flu A(H3N2) subtype was the most predominant circulating strain in the country and globally. Genetic characterization of the Flu viruses in Bhutan showed a close relatedness of high vaccine efficacy with the vaccine strain that WHO recommended for the 2022-23 season., Competing Interests: Authors have declared that no competing interest exist., (Copyright: This is an open access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication.)

Bovine tuberculosis (bTB) is a chronic zoonotic disease affecting cattle of all age groups including wild animals. It poses a significant threat to public health and high economic losses to dairy farmers. While the disease has been eradicated from most of the developed countries through extensive surveillance, testing and culling strategy, it is endemic in Africa, Asia, and the Middle East countries. Currently, there is limited research regarding the prevalence of bTB in cattle in Bhutan. This study aimed to determine the seroprevalence of bTB in cattle in six districts of eastern Bhutan. A two-stage probability proportional to size (PPS) sampling strategy was used to determine the number of animals from which serum samples needed to be collected in each district and sub-district. All farms and cattle for sampling were randomly selected from the data in the annual livestock census of 2020. The samples were tested using bTB ELISA test kit. The seroprevalence and their 95% confidence intervals were calculated. Logistic regression models were constructed to assess the influence of various individual animal and environmental risk factors (breed, age, sex, source of animal, body condition scores of animals, respiratory system status) associated with sero-positivity in animals. The study revealed an apparent seroprevalence of 2.57% (25/971 cattle; 95% CI:1.58-3.57), with an estimated true seroprevalence of 0.91% (95% CI: 0.0-2.81). However, none of the variables were found to be significantly associated with bTB seroprevalence in cattle. We recommend, further sampling and employment of confirmatory testing to fully ascertain the extent of bTB in the cattle herds in eastern Bhutan for prevention and control., Competing Interests: ‘The authors have declared that no competing interests exist., (Copyright: © 2024 Wangmo et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Soft tissue reconstruction plays an integral part in orthopaedic surgery. For developing country like Bhutan, where no micro-surgical or plastic surgeons are available, orthopaedic surgeons perform the local or regional flaps for the soft tissue defects. In this paper, we describe the use of different kinds of local and regional flaps and its outcome at Eastern Regional Referral Hospital, Bhutan., (© 2024 The Authors. International Wound Journal published by Medicalhelplines.com Inc and John Wiley & Sons Ltd.)

Introduction: Respiratory syncytial virus (RSV) is a leading cause of lower respiratory tract infections worldwide, particularly in young children. In Bhutan, respiratory disease continues to be among the top 10 diseases of morbidity for several years. This study aimed to estimate the prevalence of RSV among hospitalized patients with severe acute respiratory infection (SARI) in Bhutan., Method: Respiratory specimens were collected from SARI patients of all ages in 2016 and 2018 following influenza surveillance guidelines. Specimens were tested for influenza and RSV, human metapneumovirus, adenovirus, and human parainfluenza virus types 1, 2, and 3 using real-time reverse-transcription polymerase chain reaction assay. Descriptive statistics were used to analyze the result in STATA 16.1., Result: Of the 1339 SARI specimens tested, 34.8% were positive for at least one viral pathogen. RSV was detected in 18.5% of SARI cases, followed by influenza in 13.4% and other respiratory viruses in 3%. The median age of SARI cases was 3 (IQR: 0.8-21 years) years. RSV detection was higher among children aged 0-6 (Adj OR: 3.03; 95% CI: 1.7-5.39) and 7-23 months (Adj OR: 3.01; 95% CI: 1.77-5.12) compared with the children aged 5-15 years. RSV was also associated with breathing difficulty (Adj OR: 1.73; 95% CI: 1.17-2.56) and pre-existing lung disease, including asthma (Adj OR: 2.78; 95% CI: 0.99-7.8)., Conclusion: Respiratory viruses were detected in a substantial proportion of SARI hospitalizations in Bhutan., Competing Interests: All authors declared that they have no conflict of any financial interest to disclose., (© 2024 The Authors. Influenza and Other Respiratory Viruses published by John Wiley & Sons Ltd.)

Background: The coronavirus disease 2019 (COVID-19) pandemic, caused by a dynamic virus, has had a profound global impact. Despite declining global COVID-19 cases and mortality rates, the emergence of new severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants remains a major concern. This study provides a comprehensive analysis of the genomic sequences of SARS-CoV-2 within the Bhutanese population during the pandemic. The primary aim was to elucidate the molecular epidemiology and evolutionary patterns of SARS-CoV-2 in Bhutan, with a particular focus on genetic variations and lineage dynamics., Methods: Whole-genome sequences of SARS-CoV-2 collected from Bhutan between May 2020 and February 2023 (n=135) were retrieved from the Global Initiative on Sharing All Influenza Database., Results: The SARS-CoV-2 variants in Bhutan were predominantly classified within the Nextstrain clade 20A (31.1%), followed by clade 21L (20%) and clade 22D (15.6%). We identified 26 Pangolin lineages with variations in their spatial and temporal distribution. Bayesian time-scaled phylogenetic analysis estimated the time to the most recent common ancestor as February 15, 2020, with a substitution rate of 0.97×10-3 substitutions per site per year. Notably, the spike glycoprotein displayed the highest mutation frequency among major viral proteins, with 116 distinct mutations, including D614G. The Bhutanese isolates also featured mutations such as E484K, K417N, and S477N in the spike protein, which have implications for altered viral properties., Conclusion: This is the first study to describe the genetic diversity of SARS-CoV-2 circulating in Bhutan during the pandemic, and this data can inform public health policies and strategies for preventing future outbreaks in Bhutan.

Objectives: This study examines the status of implementation of the International Code of Marketing of Breast-milk Substitutes of eight countries in the South Asia region (Afghanistan, Bangladesh, Bhutan, India, Maldives, Nepal, Pakistan, and Sri Lanka), and describes the sales value and volume of commercial milk formula (CMF) marketed as breastmilk substitutes (BMS) and baby food in four countries (Bangladesh, India, Pakistan, and Sri Lanka)., Design: A mix of descriptive methods is used to assess national status of Code implementation, including a desk review of the 2022 WHO/UNICEF/IBFAN Code Status Report, systematic content analysis of national Code measures, and insights generated from the participation of key government and UNICEF/WHO actors in a regional workshop that aimed to identify each country's barriers, gaps, and the status of Code implementation. Data on the sales value and volume of CMF and baby food between 2007 to 2021 and with the prediction to 2026 in Bangladesh, India, Pakistan, and Sri Lanka were obtained from Global Data., Findings: There are major gaps in Code implementation in countries even with legal measures considered substantially aligned with the Code, such as the inadequate age range of CMF covered in the scope, insufficient safeguards against conflicts of interest in the health system, lack of warning of risks of intrinsic contamination of powdered milk formula, and an absence of effective monitoring and enforcement mechanisms. Data on CMF sales shows health facilities and pharmacies sustain the highest sales. Lower sales volume of infant formula (including special formula), compared to other CMF such as follow-up formula and growing-up milk, has been observed in three of the four countries (Bangladesh, India, and Sri Lanka). Overall, GUM, followed by baby cereals, accounted for a large portion of CMF and baby foods sales in the same three countries., Recommended Actions Include: (1) Closing the gaps between national measures and the Code, (2) Ensuring effective monitoring and enforcement mechanisms, (3) Strengthening conflicts of interest safeguards in the health system, (4) Tackling digital marketing, and (5) Galvanizing political support and support from in-country public health and women's rights jurist networks., Competing Interests: 4.1.8.Conflicts of interest within the health system and government, such as industry sponsorship of health programs and industry’s participation in policymaking and monitoring, can impede the adoption of strong laws and proper enforcement. This study reveals two main problems. Firstly, for some countries, their law specifies that financial or material inducements are not allowed only if they are used for promoting products covered in the law. This creates situations where financial or materials inducements could arguably be allowed so long as they are not explicitly specified with a purpose to promote designated products. Secondly, the definition of conflicts of interest may not be clear, allowing room for companies to establish ambiguous ‘partnerships’ or ‘corporate social responsibility’ campaigns that are fraught with conflicts of interest. The 2016 WHO Guidance reinforces with clearer conflicts of interest safeguards within the health systems.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Ching, Sethi, Nguyen, Murira, Shats, Rowel, Ahmed, Dorji, Chakma, Haag, Singh, Khatoon, Bukhari, Aminee, Ghosh, Forissier, Kappos, Zambrano and Khan.)

Underdiagnosis and underreporting of scrub typhus has increasingly affected public health in Bhutan since its initial detection in 2008. Identifying scrub typhus risk factors would support early diagnosis and treatment for this nonspecific febrile disease, reducing the incidence of potentially fatal complications. We conducted a hospital-based, case‒control study during October‒December 2015 in 11 scrub typhus‒prone districts. We identified harvesting cardamom as the major risk factor (odds ratio 1,519; p<0.001); other factors were traditional housing, largely caused by an outside toilet location, as well as owning a goat and frequently sitting on grass. Harvesting vegetables, herding cattle in the forest, and female sex were protective. Age had a nonlinear effect; children and the elderly were more likely to seek treatment for clinical scrub typhus. This study has informed public health policies and awareness programs for healthcare workers through development of National Guidelines for Prevention, Treatment and Control of Scrub Typhus in Bhutan.

Laboratory medicine plays a critical role in the modern healthcare system, and it is reported to influence 60-70% of clinical decision makings. The quantitative laboratory test results are interpreted by comparing to the Reference Intervals (RIs) and therefore the use of appropriate RIs is critical. Clinical laboratories in Bhutan have been randomly using RIs from textbooks and manufacturer's package inserts without even verifying their applicability and therefore lessening their contribution to clinical decision makings. To improve the healthcare service delivery in Bhutan, this study aims to establish routine clinical chemistry and haematological test RIs for healthy adults in the Bhutanese population. Out of 1150 (male, n = 570; female, n = 580) healthy Bhutanese adults listed for the study through a simple random sampling technique, 1002 (male, n = 405; female, n = 597) individuals were assessed and 815 (male, n = 372; female, n = 443) individuals were enrolled in the study. An adequate volume of venous blood was drawn from these participants with the use of standard phlebotomy technique for clinical chemistry and haematological analysis. The laboratory data were analysed with the use of statistical methods recommended by the International Federation of Clinical Chemistry and Laboratory Medicine and Clinical and Laboratory Standards Institute. After excluding the test results indicating underlying pathology and statistically detected outliers, a maximum of 775 (male, n = 346; female, n = 429) and 784 (male, n = 351; female, n = 433) individuals test values were eligible for clinical chemistry and haematology RIs establishment respectively. Statistically, there were no significant differences between age groups of same-sex for both test categories; however, significant differences between sex were observed for various test parameters in both test categories. Our RIs are generally comparable to other published literature. The established RIs are applicable to all the adult Bhutanese population; however, clinical laboratories should validate the transference of these RIs before using them for clinical purposes., Competing Interests: The authors have declared that no competing interests exist.

Objective: To determine the effectiveness of ultrasound/phonophoresis as an adjuvant to exercise or manual therapy for the improvement of patient-centred outcomes in adults with non-specific neck pain (NSNP)., Methods: Seven electronic databases were systematically searched up to September 2020. The Grading of Recommendations, Assessment, Development and Evaluation (GRADE) approach was used to judge the Quality of Evidence (QoE)., Results: Six studies involving 249 participants were included. The QoE was very low GRADE. Phonophoresis with capsaicin plus exercise improved pain at immediate post-treatment (MD: -3.30 [-4.05, -2.55]) but not with diclofenac sodium plus exercise as compared to exercise. Continuous ultrasound (CUS) plus exercise improved pain and pressure pain threshold (PPT) at immediate post-treatment (pain: MD: -3.42 [-4.08, -2.7]); (PPT: MD: 0.91 [0.68, 1.14]) and at intermediate-term as compared to exercise. CUS or high power pain threshold (HPPT) ultrasound plus manual therapy and exercise showed no benefit for pain reduction (MD: -0.75 [-2.08, 0.58]) did not improve function/disability (MD: -1.05 [-4.27, 2.17]) at immediate or short-term as compared to manual therapy and exercise., Conclusions: Due to high risk of bias, inconsistency, and indirectness the QoE is very low in support of benefit of ultrasound/phonophoresis as an adjuvant treatment for NSNP.Implication for rehabilitationDue to high risk of bias, inconsistency, and indirectness the quality of evidence (QoE) is very low in support of benefit of adding ultrasound or phonophoresis to exercise or manual therapy for pain reduction or improvement in function/disability for those with sub-acute and chronic myofascial associated neck pain. However, our confidence in the findings is very low and conclusions are likely to change as more evidences emerges.Clinicians using ultrasound therapy as an adjuvant intervention for management of chronic myofascial associated neck pain should carefully consider the available evidence on ultrasound, including the benefits and costs involved.

This study investigated Bhutanese science teachers' conceptions of the nature of science (NOS). The study recruited 225 Bhutanese science teachers based on convenient and snowball sampling techniques. The data was collected using the Myths of Science Questionnaire (MOSQ). The MOSQ was designed on Google Forms and administered through the online survey mode. The data was analysed using descriptive statistics in terms of the measure of frequency supported by science teachers' open-ended written responses, Independent Sample t -test, and One-way Analysis of Variance (ANOVA). Findings from descriptive statistics showed that Bhutanese science teachers considerably lacked clear understanding of the NOS in terms of scientific knowledge, scientific method, scientists' work, and scientific enterprise. The Independent Sample t -test showed that there was no statistically significant difference between Bhutanese male and female science teachers' perceptions of the NOS with p  > .05. The One-way ANOVA test revealed statistically significant differences amongst Bhutanese science teachers' perceptions of the NOS based on their academic qualifications with p <  .05. The Tukey Post-hoc test, however, revealed that Bhutanese science teachers' perceptions of the NOS based on academic qualifications was significant only between teachers with postgraduate diploma and doctor of philosophy., Competing Interests: Competing interestsThe authors declare that they have no competing interests., (© The Author(s) 2021.)

Background: Alcohol use is a major public health problem in Bhutan. Compliance with regulations at the point of sale is an important strategy in alcohol control. Retail outlets were briefed on sale regulations and provided notification of rules, which they were directed to display on the premises. The extent to which licensed alcohol outlets responded to possible alcohol purchases was assessed through the use of young proxy-purchasers, adults feigning alcohol intoxication and sober adults. A total of 854 visits (pre versus post visits) were made across four district towns. Two towns (Damphu town in Tsirang district and Pema Gatshel town in Pema Gatshel district) received pre- and post-intervention purchase surveys, while the other two neighbouring towns (Khuruthang town in Punakha and Bajo town in Wangdue) were administered only baseline surveys., Method: We used a pre- and post-test community intervention design covering all alcohol retailers both on premise (bar, hotel, restaurant, karaoke bars) and off premise (grocery shops). Compliance with alcohol regulations at the point of sale was assessed through the use of young proxy-purchasers, adults feigning alcohol intoxication and sober adults., Results: Retailers rarely checked the age and/or identification (ID) of the proxy-purchasers before the intervention. There was a 22.7% (8.6, 37) percent increase in compliance with laws after the intervention. While some strategies are suggested, the strongest and most effective measure to prevent under-age drinking and service to intoxicated individuals is more rigorous enforcement of existing liquor laws., Conclusion: Alcohol control requires ongoing government enforcements, supplemented by public awareness and knowledge., (© 2021. The Author(s).)

Purpose: To determine the outcome and predictive factors of clinical success of bronchial artery embolization in life-threatening hemoptysis., Material and Methods: We reviewed all bronchial artery embolization procedures performed for life-threatening hemoptysis between January 2008 and December 2018. The outcomes and predictive factors of clinical success following embolization were evaluated., Results: One hundred and eighty-four bronchial artery embolization procedures performed in 145 patients were eligible for the study. The technical and clinical success rates of the procedures were 170/184 (92.4%) and 129/184 (70.1%), respectively. The unstable hemodynamics and prothrombin time/international normalized ratio >1.5 was associated with lower clinical success rate, while embolization of more than one vessel was associated with higher clinical success rate., Conclusion: Bronchial artery embolization is a safe and effective procedure for controlling bleeding in life-threatening hemoptysis. However, low clinical success rate was noted in patients with unstable hemodynamics and coagulopathy, while multiple vessel embolization was associated with higher clinical success., Competing Interests: The authors have no competing interests to declare., (Copyright: © 2021 The Author(s).)

Purpose: Bhutan, known as a country of happiness, has experienced rapid social changes and the increasing burden of non-communicable diseases (NCDs) that can impact health and happiness. To inform future NCD prevention programs in Bhutan, this study explores knowledge, perception, and the practices of Bhutanese related to NCDs in the context of the philosophy of happiness., Methodology: Research was conducted in rural and urban communities of Bhutan in 2017 among 79 inhabitants of both genders, aged ≥18. Participants were recruited through purposive sampling with the data collected by in-depth interviews, participatory observation, and anthropometric measurements. Data were analyzed by thematic analysis., Results/discussion: Across participants, health was considered as an important element of "happiness". However, lifestyle-related NCD risk factors prevailed due to the lack of effective education programs on NCDs and thus the lack of practical knowledge for NCD prevention across society. We further found that the value of happiness "finding happiness in any situation is virtue" was universal as well as other traditional values and customs, shaping people's health behaviors. From these observations, it is recommended that more practical NCD education/prevention programs should be urgently introduced in Bhutan that involve multiple generations, religion authorities, educational settings, and medical services., Originality: This is the first comprehensive qualitative study on the NCD-related lifestyle risks among Bhutanese concerning the concept of happiness., Competing Interests: The authors have declared that no competing interests exist.

Background: Direct observed treatment (DOT) has been implemented in Bhutan since 1997 and currently, it is offered in various model of delivery including a combination of hospital based, home based DOT and ambulatory DOT. Overall, treatment success rate for tuberculosis cases is higher than the global target; however, it is still need to be improved. Evaluation to the implementation fidelity of DOT is important to identify potential rooms for improvement. This study aimed to assess two major components of the program's implementation fidelity: to assess patient's adherence to DOT and explore factors for adherence; to assess provider's compliance with DOT guideline and explore factors for compliance., Methods: This research used a sequential explanatory mixed method. The conceptual framework of implementation fidelity was adopted to guide this study design. The cross-sectional study of TB patients was enrolled in two hospitals with highest TB load, between September to November 2017 in Bhutan. Interviewer assisted survey was conducted with 139 TB patients who visited the hospital in continuation phase. In-depth interview was then conducted with nine TB patients and four health staffs to explore the barriers and enablers of DOT., Results: Total of 61.9% (86/139) of patients has received DOT at intensive phase. Proportion was higher among MDR-TB cases (100%), and smear sputum positive TB cases (84.7%). In the continuation phase, 5.8% of patients took medicine at hospital, 48.9% at home and the rest 45.3% no longer practiced DOT. More than 90% of patient received correct dosage and standard regimen of anti-TB drugs according to the guideline. The key factors affecting poor adherence to DOT as perceived by patients were; lack of willingness to visit the clinic on daily basis due to long distance, financial implications and family support. However, patient's satisfaction to the quality of TB treatment service delivery was high (98.6%). Providing incentives to the patient was most agreed enabler felt by both health workers and patients., Conclusion: In the selected hospital sites, the patient's adherence to DOT and provider's compliance with DOT guideline is partially implemented; the coverage and the duration of DOT is very low, therefore, need to revise and improve DOT model and structure.

Threat assessment is critical to species conservation and management planning, because prior identification and assessment of key threats to conservation planning can assist in developing appropriate interventions or strategies. Comprehensive threat assessments are currently lacking for many threatened primates. In this paper, we classify and rank all direct threats to the endangered golden langur (Trachypithecus geei) in Bhutan in order to provide a practical guide to future conservation of the species. Information on threats was based on interviews with local people, discussion with field forestry staff, and social media interaction. We classified threats to golden langur habitats and populations, and ranked them using Miradi™, an analytical software for the adaptive management of conservation projects. We identified five habitat threats: (1) hydropower development, (2) road development, (3) housing development, (4) resource extraction, and (5) agricultural expansion. We also identified seven population threats: (1) electrocution, (2) road kill, (3) road injury, (4) dog kill, (5) retaliatory killing, (6) illegal pet keeping, and (7) hybridization with capped langurs. We rated the overall threat to golden langurs in Bhutan as 'medium'. Hydropower, road, and housing development constituted 'high' impact, while agricultural expansion, resource extraction, electrocution, and road kill had 'medium' impact; the remaining threats had 'low' impact. To immediately mitigate threats to golden langurs, we recommend: (a) installing speed limit signage and speed breakers with strict enforcement of speed limits; (b) installing insulated electric cables and fencing around power transformers; and (c) reducing and restraining domestic dog populations.

In 2018, Bhutan reported 54 cases of malaria, of which six were indigenous, 14 introduced and 34 imported. Considering the continuous reduction in the number of indigenous cases, Bhutan plans to eliminate malaria by 2025 under the Bhutan Malaria Elimination Strategy. The study was conducted to assess the presence of asymptomatic plasmodial infection in both, Bhutanese population living in malaria-risk areas and in migrant workers to guide the elimination strategies. A cross-sectional study was conducted from April to May 2016 in 750 Bhutanese people and 473 migrant workers. Plasmodium falciparum and Plasmodium vivax infections were investigated by using a rapid diagnostic test (RDT) and the polymerase chain reaction (PCR). Prevalence of asymptomatic plasmodial infection based on PCR was 0.27% (95% CI: 0.05-1.07%) among Bhutanese people with a mean age of 43 years old. The proportions of males and females were 45% and 55%, respectively. Among migrant workers, the prevalence of asymptomatic plasmodial infection was 0.42% (95% CI: 0.07-1.69%) with a mean age of 30 years old. The majority of migrant workers were from the neighboring Indian State of West Bengal (57.51%), followed by Assam (12.26%). RDT in both study groups did not detect any plasmodial infection. The presence of a low prevalence of asymptomatic plasmodial infection indicates that the current elimination strategies and interventions are effective.

Scrub typhus (ST) is a vector-borne rickettsial infection causing acute febrile illness. The re-emergence of ST in the Asia-Pacific region represents a serious public health threat. ST was first detected in Bhutan in 2008. However, the disease is likely to be under-diagnosed and under-reported, and the true impact is difficult to estimate. At the end of 2014, the SD Bioline Tsutsugamushi Test TM rapid diagnostic test (RDT) kits became available in all hospitals to assist clinicians in diagnosing ST. We conducted a retrospective descriptive study, reviewing records from all hospitals of Bhutan to identify all RDT-positive clinical cases of ST in Bhutan in 2015. The aim was to evaluate the burden of ST in Bhutan, describe the demographic, spatial and temporal patterns of disease, and identify the typical clinical presentations. The annual incidence of RDT-positive cases of ST reporting to Bhutanese hospitals in 2015 was estimated to be 62 per 100,000 population at risk. The incidence of disease was highest in the southern districts with a subtropical climate and a high level of agricultural production. The highest proportion of cases (87%) was rural residents, with farmers being the main occupational category. The disease was strongly seasonal, with 97% of cases occurring between June and November, coinciding with the monsoon and agricultural production seasons. Common ST symptoms were not specific, and an eschar was noted by clinicians in only 7.4% of cases, which is likely to contribute to an under-diagnosis of ST. ST represents an important and neglected burden, especially in rural communities in Bhutan. The outcomes of this study will inform public health measures such as timely-awareness programmes for clinicians and the public in high-risk areas, to improve the diagnosis, treatment and clinical outcomes of this disease.

Objective: This study aims to identify the required management competencies, current competency levels, and strategies for improving the management competencies of Bhutanese primary health care (PHC) managers., Methods: A quantitative method with a cross-sectional survey using self-administered questionnaires. This study recruited 339 PHC managers across Bhutan. The data were analyzed using statistical software., Results: This study identified three competency domains and seven key sub-domain competencies. People domain was perceived to be the highest required competency with a mean score of 4.2376, followed by execution (4.1851), and the transformation (4.0501) domains. For the seven key sub-domains, the communication sub-domain (4.3220) was perceived as the highest required competency, followed by professionalism (4.2967), managing change (4.1776), relationship building (4.1686), analytical thinking (4.1091), leadership (4.0980), and innovative thinking (3.9794). The current competency levels of PHC managers in domains and sub-domain competencies were the people domain (3.7322), execution (3.6471), and the transformation (3.5554). For the sub-domains, communication (3.8092), professionalism (3.7939), relationship building (3.6603), analytical thinking (3.6396), leadership (3.5805), managing change (3.5723), and innovative thinking (3.4543)., Conclusion: Findings of Bhutan health managers' competencies are consistent with the findings of other international contexts. This study suggests that agencies responsible for health system need to focus more on the competencies defined by the study to positively influence health leadership and management development interventions., Competing Interests: Disclosure The authors report no conflicts of interest in this work.

Rodents are well-known reservoirs and vectors of many emerging and re-emerging infectious diseases, but little is known about their role in zoonotic disease transmission in Bhutan. In this study, a cross-sectional investigation of zoonotic disease pathogens in rodents was performed in Chukha district, Bhutan, where a high incidence of scrub typhus and cases of acute undifferentiated febrile illness had been reported in people during the preceding 4-6 months. Twelve rodents were trapped alive using wire-mesh traps. Following euthanasia, liver and kidney tissues were removed and tested using PCR for Orientia tsutsugamushi and other bacterial and rickettsial pathogens causing bartonellosis, borreliosis, human monocytic ehrlichiosis, human granulocytic anaplasmosis, leptospirosis, and rickettsiosis. A phylogenetic analysis was performed on all rodent species captured and pathogens detected. Four out of the 12 rodents (33.3%) tested positive by PCR for zoonotic pathogens. Anaplasma phagocytophilum, Bartonella grahamii, and B. queenslandensis were identified for the first time in Bhutan. Leptospira interrogans was also detected for the first time from rodents in Bhutan. The findings demonstrate the presence of these zoonotic pathogens in rodents in Bhutan, which may pose a risk of disease transmission to humans.

Background: Due to competing health priorities and limited resources, many low-income countries, even those with a high disease burden, are not able to introduce pneumococcal conjugate vaccines., Objective: To determine the cost-utility of 10- and 13-valent pneumococcal conjugate vaccines (PCV10 and PCV13) compared to no vaccination in Bhutan., Methods: A model-based cost-utility analysis was performed in the Bhutanese context using a government perspective. A Markov simulation model with one-year cycle length was used to estimate the costs and outcomes of three options: PCV10, PCV13 and no PCV programmes for a lifetime horizon. A discount rate of 3% per annum was applied. Results are presented using an incremental cost-effectiveness ratio (ICER) in United State Dollar per quality-adjusted life year (QALY) gained (USD 1 = Ngultrum 65). A one-way sensitivity analysis and a probabilistic sensitivity analysis were conducted to assess uncertainty., Results: Compared to no vaccination, PCV10 and PCV13 gained 0.0006 and 0.0007 QALYs with additional lifetime costs of USD 0.02 and USD 0.03 per person, respectively. PCV10 and PCV13 generated ICERs of USD 36 and USD 40 per QALY gained compared to no vaccination. In addition, PCV13 produced an ICER of USD 92 compared with PCV10. When including PCV into the Expanded Programme on Immunization, the total 5-year budgetary requirement is anticipated to increase to USD. 3.77 million for PCV10 and USD 3.75 million for PCV13. Moreover, the full-time equivalent (FTE) of one health assistant would increase by 2.0 per year while the FTE of other health workers can be reduced each year, particularly of specialist (0.6-1.1 FTE) and nurse (1-1.6 FTE)., Conclusion: At the suggested threshold of 1xGDP per capita equivalent to USD 2708, both PCVs are cost-effective in Bhutan and we recommend that they be included in the routine immunization programme., (Copyright © 2018 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Background: Human beings use plants for a multitude of purposes of which a prominent one across the globe is for their medicinal values. Medicinal plants serve as one of the major sources of income for high altitude inhabitants in the Himalaya, particularly in countries like Nepal, and Bhutan. People here harvest huge volumes of medicinal plants indiscriminately, risking their sustainability. This paper attempts to identify some of the priority medicinal plant species harvested in the wild and assess their ecological status for their judicious utilization, and to help provide policy guidance for possible domestication and support strategic conservation frameworks., Results: Out of the 16 priority species identified by the expert group, collectors' perception on ecological status of the priority species differed from survey findings. Chrysosplenium nudicaule (clumps) ranked as most threatened species followed by Corydalis dubia, and Meconopsis simplicifolia. Onosma hookeri, Corydalis crispa and Delphinium glaciale were some of the species ranked as threatened species followed by Halenia elliptica (not in priority list). Percent relative abundance showed irregular pattern of species distribution. High species evenness was recorded among Nardostachys grandiflora, Chrysosplenium nudicaule, Saussurea gossypiphora and Aconitum orochryseum with average species density of 8 plant m -2 . Rhodiola crenulata, and Dactylorhiza hatagirea followed by Meconopsis horridula and Meconopsis simplicifolia were ranked as most threatened species with average species density of 0.4, 0.4, 5.6 and 6.0 plant m -2 , respectively. The most abundant (common) species was Onosma hookeri (plant m -2 ). Species composition and density also differed with vegetation, altitude, slope and its aspects., Conclusion: Priority species identified by expert group were found vulnerable and patchy in distribution. Survey results and collectors' perceptions tally to an extent. Some of the species (Dactylorhiza hatagirea, Rhodiola crenulata, Meconopsis simplicifolia and Meconopsis horridula) were critically low in plant density with less than a plant per m 2 while Delphinium glaciale, Fritillaria delavayi and Aconitum orochryseum were confined to narrow altitude range. Collectors were aware that most species identified in priority list are threatened and existing harvesting plan are hardly implemented as it is not pragmatic. Moreover, major chunk of medicinal plants harvested remain unaccounted as illegal harvest and marketing seemed to occur across the borders. Policing and monitoring would continue to be a challenge given the rugged terrain and harsh climate. In-depth study and further monitoring of low density species is suggested to ensure its sustainability through long term strategy development.

Background: Bhutan has made substantial progress in reducing malaria incidence. The national guidelines recommend chloroquine (CQ) and primaquine (PQ) for radical cure of uncomplicated Plasmodium vivax, but the local efficacy has not been assessed. The impact of cases imported from India on the genetic make-up of the local vivax populations is currently unknown., Methods: Patients over 4 years of age with uncomplicated P. vivax mono-infection were enrolled into a clinical efficacy study and molecular survey. Study participants received a standard dose of CQ (25 mg/kg over 3 days) followed by weekly review until day 28. On day 28 a 14-day regimen of PQ (0.25 mg/kg/day) was commenced under direct observation. After day 42, patients were followed up monthly for a year. The primary and secondary endpoints were risk of treatment failure at day 28 and at 1 year. Parasite genotyping was undertaken at nine tandem repeat markers, and standard population genetic metrics were applied to examine population diversity and structure in infections thought to be acquired inside or outside of Bhutan., Results: A total of 24 patients were enrolled in the clinical study between April 2013 and October 2015. Eight patients (33.3 %) were lost to follow-up in the first 6 months and another eight patients lost between 6 and 12 months. No (0/24) treatment failures occurred by day 28 and no (0/8) parasitaemia was detected following PQ treatment. Some 95.8 % (23/24) of patients were aparasitaemic by day 2. There were no haemolytic or serious events. Genotyping was undertaken on parasites from 12 autochthonous cases and 16 suspected imported cases. Diversity was high (H E 0.87 and 0.90) in both populations. There was no notable differentiation between the autochthonous and imported populations., Conclusions: CQ and PQ remains effective for radical cure of P. vivax in Bhutan. The genetic analyses indicate that imported infections are sustaining the local vivax population, with concomitant risk of introducing drug-resistant strains.

Introduction: Determining the optimal time to vaccinate is important for influenza vaccination programmes. Here, we assessed the temporal characteristics of influenza epidemics in the Northern and Southern hemispheres and in the tropics, and discuss their implications for vaccination programmes., Methods: This was a retrospective analysis of surveillance data between 2000 and 2014 from the Global Influenza B Study database. The seasonal peak of influenza was defined as the week with the most reported cases (overall, A, and B) in the season. The duration of seasonal activity was assessed using the maximum proportion of influenza cases during three consecutive months and the minimum number of months with ≥80% of cases in the season. We also assessed whether co-circulation of A and B virus types affected the duration of influenza epidemics., Results: 212 influenza seasons and 571,907 cases were included from 30 countries. In tropical countries, the seasonal influenza activity lasted longer and the peaks of influenza A and B coincided less frequently than in temperate countries. Temporal characteristics of influenza epidemics were heterogeneous in the tropics, with distinct seasonal epidemics observed only in some countries. Seasons with co-circulation of influenza A and B were longer than influenza A seasons, especially in the tropics., Discussion: Our findings show that influenza seasonality is less well defined in the tropics than in temperate regions. This has important implications for vaccination programmes in these countries. High-quality influenza surveillance systems are needed in the tropics to enable decisions about when to vaccinate.

Background: Comprehensive biotic surveys, or 'all taxon biodiversity inventories' (ATBI), have traditionally been limited in scale or scope due to the complications surrounding specimen sorting and species identification. To circumvent these issues, several ATBI projects have successfully integrated DNA barcoding into their identification procedures and witnessed acceleration in their surveys and subsequent increase in project scope and scale. The Biodiversity Institute of Ontario partnered with the rare Charitable Research Reserve and delegates of the 6th International Barcode of Life Conference to complete its own rapid, barcode-assisted ATBI of an established land trust in Cambridge, Ontario, Canada., New Information: The existing species inventory for the rare Charitable Research Reserve was rapidly expanded by integrating a DNA barcoding workflow with two surveying strategies - a comprehensive sampling scheme over four months, followed by a one-day bioblitz involving international taxonomic experts. The two surveys resulted in 25,287 and 3,502 specimens barcoded, respectively, as well as 127 human observations. This barcoded material, all vouchered at the Biodiversity Institute of Ontario collection, covers 14 phyla, 29 classes, 117 orders, and 531 families of animals, plants, fungi, and lichens. Overall, the ATBI documented 1,102 new species records for the nature reserve, expanding the existing long-term inventory by 49%. In addition, 2,793 distinct Barcode Index Numbers (BINs) were assigned to genus or higher level taxonomy, and represent additional species that will be added once their taxonomy is resolved. For the 3,502 specimens, the collection, sequence analysis, taxonomic assignment, data release and manuscript submission by 100+ co-authors all occurred in less than one week. This demonstrates the speed at which barcode-assisted inventories can be completed and the utility that barcoding provides in minimizing and guiding valuable taxonomic specialist time. The final product is more than a comprehensive biotic inventory - it is also a rich dataset of fine-scale occurrence and sequence data, all archived and cross-linked in the major biodiversity data repositories. This model of rapid generation and dissemination of essential biodiversity data could be followed to conduct regional assessments of biodiversity status and change, and potentially be employed for evaluating progress towards the Aichi Targets of the Strategic Plan for Biodiversity 2011-2020.

Background: Bhutan is a small Himalayan country lying within the region considered to be the origin of citrus. Diverse citrus wild types grow naturally in different climates, elevations and edaphic conditions, but only mandarin is cultivated commercially. The first report of Huanglongbing (also known as greening disease) in Bhutan in 2003, and the threat it posed to the country's citrus orchards prompted the collection of mandarin germplasm from across the country. This paper describes the genetic diversity of mandarin accessions in Bhutan using amplified fragment length polymorphic (AFLP) markers., Results: Twenty three accessions of Bhutanese mandarin were analyzed using AFLP markers to assess the genetic variability that is believed to exist only in Bhutan and some parts of North East India and South China. Five primer pairs (E-ACA/M-CAG, E-ACG/M-CAT, E-ACC/M-CTT, E-AAG/M-CAA and E-ACA/M-CTC) were identified (based on the number and quality of polymorphic bands produced) and used for the analyses. A total of 244 bands were scored visually of which 126 (52%) were polymorphic with an average polymorphism information content of 0.95 per marker. A cluster dendrogram based on multiscale bootstrap sampling categorized twenty three accessions into two broad groups containing eight and 14 accessions, respectively. Group A consisted accessions (Tsirang1, Tsirang3, Sarpang1, Dagana4, Samtse4, Dagana1, and Trongsa2) from five districts (Tsirang, Sarpang, Samtse, Dagana and Trongsa) and their grouping was strongly supported by bootstrap analysis (B p-value = 96%, AU p-value = 86%). Cluster B consisted of 14 accessions divided into three sub-groups (1, 2 and 3). However, bootstrap value supported significantly for subgroup1 (containing accessions: Tsirang4, Sarpang5, and Tsirang2) and subgroup3 (with accessions - Zhemgang2, Zhemgang3 and Zhemgang4)., Conclusion: This study indicates that Bhutanese mandarin germplasm collected from across the country are genetically diverse although the level of variability differed among the accessions assessed. The variation in genetic variability was observed irrespective of where the accessions were collected suggesting that phenotype and geographical location can serve a basis for future germplasm collection in Bhutan. Further, five primer pair combinations could separate 23 mandarins accessions considered in this study, suggesting that AFLP markers can be a useful tool for future identification.

Trade-offs in allocation to resprouting vs. seedling regeneration in plants are predicted to occur along fire disturbance gradients. Increased resprouting ability should be generally favored in plant communities with a high probability of crown fire return. Hence, communities dominated by resprouters are predicted to have smaller seed banks than those dominated by species killed by fire. We tested whether there were trait shifts in resprouting ability among growth forms (short-lived herbaceous vs. ground-dwelling perennials vs. shrubs) and among communities (rocky outcrop vs. sclerophyll forest) with contrasting probabilities of crown fire return. Resprouting was more common in ground-dwelling perennials and in the sclerophyll forest community with a high probability of crown fire. Soil seed banks were sampled in rocky outcrop and sclerophyll forest communities in recently burned (18 months) and long-since-burned (12 years) locations at interspersed replicated sites. Collected seed banks were treated with orthogonal treatments of fire stimuli or no stimuli, and seedling emergence was measured in controlled conditions. Seed bank composition reflected the pattern of extant vegetation, with resprouting species being more common in the community with a higher probability of crown fire. Overall, however, resprouting species were poorly represented in the seed bank compared to those species killed by fire. Predicted shifts in allocation to seed production were strongly manifested in community seed banks across the disturbance gradient. Fewer species, seedlings, and seedlings per adult emerged from seed banks in the sclerophyll forest. This suggests that the dominance of resprouting species influences recruitment at the community scale. Community patterns in the seed bank also reflected predicted trade-offs with plant size and growth rate. Short-lived species that are killed by fire dominated the seed bank on rocky outcrops, while longer-lived resprouting species were found in low abundance. Life history trade-offs in persistence and regeneration strongly contribute to coexistence patterns between and within communities with contrasting probabilities of fire return.