Synthetic sub-genomic transcript promoter from Horseradish Latent Virus (HRLV).

MAIN CONCLUSION: We characterized an efficient chimeric sub-genomic transcript promoter from Horseradish Latent Virus, FHS4, active in both dicot and monocot plants, and it could be a potential tool for plant biotechnology. Plant pararetroviruses are a rich source of novel plant promoters widely used for biotechnological applications. Here, we comprehensively characterized a unique sub-genomic transcript (Sgt) promoter of Horseradish Latent Virus (HRLV) and identified a fragment (HS4; - 340 to + 10; 351 bp) that showed the highest expression of reporter genes in both transient and transgenic assays as evidenced by biochemical, histochemical GUS reporter assay and transcript analysis of uidA gene by qRT-PCR. Phylogenetic analysis showed that the HSgt promoter was closely related to the sub-genomic promoter of the Cauliflower Mosaic Virus (CaMV19S). We found that the as-1 element and W-box played an important role in the transcriptional activity of the HS4 promoter. Furthermore, the HS4 promoter was also induced by salicylic acid. Alongside, we enhanced the activity of the HS4 promoter by coupling the enhancer region from Figwort Mosaic Virus (FMV) promoter to the upstream region of it. This hybrid promoter FHS4 was around 1.1 times stronger than the most commonly used promoter, 35S (Cauliflower Mosaic Virus full-length transcript promoter), and was efficient in driving reporter genes in both dicot and monocot plants. Subsequently, transgenic tobacco plants expressing an anti-microbial peptide BrLTP2.1 (Brassica rapa lipid transport protein 2.1), under the control of the FHS4 promoter, were developed. The in vitro anti-fungal assay revealed that the plant-derived BrLTP2.1 protein driven by an FHS4 promoter manifested increased resistance against an important plant fungal pathogen, Alternaria alternata. Finally, we concluded that the FHS4 promoter can be used as an alternative to the 35S promoter and has a high potential to become an efficient tool in plant biotechnology.

Inbreeding depression and its effect on sperm quality traits in Pietrain pigs.

In most cases, inbreeding is expected to have unfavourable effects on traits in livestock. The consequences of inbreeding depression could be substantial, primarily in reproductive and sperm quality traits, and thus lead to decreased fertility. Therefore, the objectives of this study were (i) to compute inbreeding coefficients using pedigree (FPED ) and genomic data based on runs of homozygosity (ROH) in the genome (FROH ) of Austrian Pietrain pigs, and (ii) to assess inbreeding depression on four sperm quality traits. In total, 74,734 ejaculate records from 1034 Pietrain boars were used for inbreeding depression analyses. Traits were regressed on inbreeding coefficients using repeatability animal models. Pedigree-based inbreeding coefficients were lower than ROH-based inbreeding values. The correlations between pedigree and ROH-based inbreeding coefficients ranged from 0.186 to 0.357. Pedigree-based inbreeding affected only sperm motility while ROH-based inbreeding affected semen volume, number of spermatozoa, and motility. For example, a 1% increase in pedigree inbreeding considering 10 ancestor generations (FPED10 ) was significantly (p < 0.05) associated with a 0.231% decrease in sperm motility. Almost all estimated effects of inbreeding on the traits studied were unfavourable. It is advisable to properly manage the level of inbreeding to avoid high inbreeding depression in the future. Further, analysis of effects of inbreeding depression for other traits, including growth and litter size for the Austrian Pietrain population is strongly advised.

Individual- and Community-Level Factors Associated with Diarrhea in Children Younger Than Age 5 Years in Bangladesh: Evidence from the 2014 Bangladesh Demographic and Health Survey.

Background: Diarrheal disease is among the leading causes of morbidity and mortality among children younger than age 5 years in Bangladesh. Objective: The objective of this study is to assess the prevalence of diarrhea among children younger than age 5 years and its associated risk factors. Methods: Data were sourced from the Bangladesh Demographic and Health Survey, a nationally representative study conducted in 2014. We used multilevel logistic regression models to identify factors associated with diarrheal disease. Results: Children aged 6 to11 months (odds ratio = 2.26; 95% CI, 1.50-3.42), and 12 to 23 months (odds ratio = 2.31; 95% CI, 1.62-3.31) were more likely to have diarrhea than older children. Other significant risk factors for diarrheal infection included households without access to drinking water (odds ratio = 1.39; 95% CI, 1.03-1.88) and mothers lacking mass media access (odds ratio = 1.32; 55% CI, 1.01-1.73). Conclusions: Childhood diarrhea in Bangladesh was associated with individual- and community-level factors. The finding of this study suggests that diarrhea prevention programs in the country can effectively be delivered by targeting young children through expanding community-based education and increasing access to health information through mass media.

Characteristics and Population Size Estimation of Female Sex Workers in Bhutan.

BACKGROUND: Knowing the number of persons at risk for sexually transmitted diseases (STDs) and their risk behaviors are needed to allocate limited resources, set targets for prevention and care activities, gauge the reach of programs, and assess their impact. Female sex workers (FSWs) are a population at high risk for STD worldwide; little is known about FSW in Bhutan. METHODS: We conducted a community-based survey and population size exercise of FSW in Bhutan. The survey used a hybrid venue-based/peer-referral sampling method. Population size estimation methods were key informant mapping, census and enumeration, unique object multiplier, and capture-recapture. RESULTS: Of 517 women surveyed, 67.5% provided sex for money in the last year. Compared with FSWs at venues, FSWs referred by peers were more likely to have sex with alcohol (80.1% vs. 51.5%, P < 0.001) and more paying partners (mean, 3.5 vs. 2.0; P = 0.001), and less likely to have tested for an STD (28.3% vs. 51.0%, P < 0.001) or have outreach worker contact (27.6% vs. 41.5%, P = 0.007). The estimated number of FSWs in the 9 districts was 353 (95% confidence interval, 345-362). Extrapolation to the whole country projected 597 (417-777) FSWs nationally. CONCLUSIONS: Our estimate of the number of FSW in Bhutan corresponds to 0.71% of adult urban women, a figure in line with other countries in South and Southeast Asia. Our data highlight the need for outreach beyond venues where women are employed to reach FSW at higher risk for STD but who have less access to interventions.

Clinical features and outcomes of COVID-19 and dengue co-infection: a systematic review.

BACKGROUND: Dengue is the most common arboviral disease in the tropical and sub-tropical regions of the world. Like other regions, dengue-endemic areas have faced the additional public health and socio-economic impact of the ongoing coronavirus disease 2019 (COVID-19) pandemic. COVID-19 and dengue co-infections have been reported, with complicated patient management and care requirements. This review aimed to collate and synthesise current knowledge on the clinical features and outcomes of COVID-19 and dengue virus co-infection, a potentially important new dimension to be considered in public health management of the COVID-19 pandemic. METHODS: A systematic literature review was conducted using PubMed, Web of Science and Scopus databases from 1st January to 21st November 2020. The key search terms used were "dengue" and "coronavirus". Descriptive analysis with graphical illustrations were used to present the clinical and laboratory parameters of the co-infection. RESULTS: Thirteen published papers and four news articles were included in the review. Most studies were case reports with a detailed description of the clinical and laboratory characteristics of the co-infection. All cases were in adults with the exception of a six-year old child. The common symptoms of co-infection were fever, dyspnea, headache, and cough. Common laboratory results included thrombocytopenia, lymphocytopenia, elevated transaminases, and leukopenia. Serious outcomes of co-infection included septic shock, acute respiratory disease syndrome and multi-organ failure, leading to death in some patients. CONCLUSIONS: COVID-19 and dengue co-infection was associated with severe disease and fatal outcomes. The correct diagnosis and treatment of co-infection poses a substantial challenge due to the overlapping clinical and laboratory parameters. Therefore, confirmative diagnostic tests are necessary for accurate and timely diagnosis and patient management.

Analysis of plant pararetrovirus promoter sequence(s) for developing a useful synthetic promoter with enhanced activity in rice, pearl millet, and tobacco plants.

Promoters are one of the most important components for many gene-based research as they can fine-tune precise gene expression. Many unique plant promoters have been characterized, but strong promoters with dual expression in both monocot and dicot systems are still lacking. In this study, we attempted to make such a promoter by combining specific domains from monocot-infecting pararetroviral-based promoters sugarcane bacilliform virus (SCBV) and banana streak virus (BSV) to a strong dicot-infecting pararetroviral-based promoter mirabilis mosaic virus (MMV). The generated chimeric promoters, MS, SM, MB, and BM, were tested in monocot and dicot systems and further validated in transgenic tobacco plants. We found that the developed chimeric promoters were species-specific (monocot or dicot), which depended on their respective core promoter (CP) region. Furthermore, with this knowledge, deletion-hybrid promoters were developed and evaluated, which led to the development of a unique dual-expressing promoter, MSD3, with high gene expression efficiency (GUS and GFP reporter genes) in rice, pearl millet, and tobacco plants. We conclude that the MSD3 promoter can be an important genetic tool and will be valuable in plant biology research and application.

Risk mapping and socio-ecological drivers of soil-transmitted helminth infections in the Philippines: a spatial modelling study.

Background: The Philippines reports a high prevalence of soil-transmitted helminth (STH) infections despite the implementation of nationwide mass drug administration since 2006. The spatial variation of STH infections in the Philippines was last described using the 2005-2007 national STH and schistosomiasis survey. This study aimed to identify sociodemographic and environmental factors that drive STH transmission and predict high-risk areas in the Philippines. Methods: Epidemiological data on STH for students aged 5-16 years were obtained from the 2015 Philippines National Prevalence survey, while environmental data were extracted from satellite images and publicly available sources. Model-based geostatistics, implemented in a Bayesian framework, was used to identify sociodemographic and environmental correlates and predict high-risk areas for STH across the Philippines. The best-fitting model with the lowest deviance information criterion (DIC) was used to interpret the findings of the model and predict STH infection risk for the entire country. Risk maps were developed for each STH infection using the posterior means derived from the model. Findings: The prevalence of Ascaris lumbricoides (20.0%) and Trichuris trichiura (29.3%) was higher in the Visayas Island than in the Luzon and Mindanao Islands. Hookworm prevalence was highest in Mindanao Island (1.3%). Risk of A. lumbricoides was positively associated with males (odds ratio [OR]: 1.197; 97.5% Credible Interval [CrI]: 1.114, 1.286) and temperature (OR: 1.148; 97.5% CrI: 1.033, 1.291), while normalized difference vegetation index (OR: 0.354; 97.5% CrI: 0.138, 0.930) and soil pH (OR: 0.606; 97.5% CrI: 0.338, 0.949) were negatively associated with the transmission. T. trichiura risk was positively associated with males (OR: 1.261; 97.5% CrI: 1.173, 1.341), temperature (OR: 1.153; 97.5% CrI: 1.001, 1.301), and rainfall (OR: 1.004; 97.5% CrI: 1.011, 1.069). Hookworm risk was positively associated with males (OR: 2.142; 97.5% CrI: 1.537, 2.998), while children aged ≤12 years (OR: 0.435; 97.5% CrI: 0.252, 0.753) had a negative association with risk compared to those over 12 years. Focal areas of high risk were identified for A. lumbricoides and T. trichiura in the Visayas Island, and hookworm in the Mindanao Island. Interpretation: The spatial distribution of all three STH infections has considerably decreased since a previous national risk-mapping exercise. The high-risk areas identified in the study can be used to strategically target deworming and health education activities to further reduce the burden of STH and support progress toward elimination. Funding: The Australian Centre for the Control and Elimination of Neglected Tropical Diseases and the Australian National Health and Medical Research Council.

Protocol for spatial prediction of soil transmitted helminth prevalence in the Western Pacific region using a meta-analytical approach.

BACKGROUND: Soil transmitted helminth (STH) infections are estimated to impact 24% of the world's population and are responsible for chronic and debilitating morbidity. Disadvantaged communities are among the worst affected and are further marginalized as infection prevalence fuels the poverty cycle. Ambitious targets have been set to eliminate STH infections, but accurate epidemiological data will be required to inform appropriate interventions. This paper details the protocol for an analysis that aims to produce spatial prediction mapping of STH prevalence in the Western Pacific Region (WPR). METHODS: The protocol follows the Preferred Reporting Items for Systematic Review and Meta-Analysis Protocol (PRISMA-P) guidelines. The study design will combine the principles of systematic review, meta-analysis, and geospatial analysis. Systematic searches will be undertaken in PubMed, Scopus, ProQuest, Embase, and Web of Science for studies undertaken post 2000, to identify surveys that enable the prevalence of human STH infection within the WPR to be calculated. Covariate data for multivariable analysis will be obtained from publicly accessible sources. Survey data will be geolocated, and STH prevalence and covariates will be linked to produce a spatially referenced dataset for analysis. Bayesian model-based geostatistics will be used to generate spatially continuous estimates of STH prevalence mapped to a resolution of 1 km2. A separate geospatial model will be constructed for each STH species. Predictions of prevalence will be made for unsampled locations and maps will be overlaid for each STH species to obtain co-endemicity maps. DISCUSSION: This protocol facilitates study replication and may be applied to other infectious diseases or alternate geographies. Results of the subsequent analysis will identify geographies with high STH prevalence's and can be used to inform resource allocation in combating this neglected tropical disease. TRIAL REGISTRATION: Open Science Framework: osf.io/qmxcj.

Evaluating Country Performance After Transitioning From Gavi Assistance: An Applied Synthetic Control Analysis.

INTRODUCTION: Over the past decade, international development assistance for health has slowed. As donors seek to increase domestic cofinancing and ultimately transition countries from donor aid dependence, COVID-19 has severely constrained public budgets. The evaluation of sustainability and longer-term impacts of donor withdrawal is increasingly important. We assess vaccination coverage and post-neonatal mortality to estimate country performance of these outcomes among countries that no longer received assistance from Gavi, the Vaccine Alliance (Gavi) between 2000 and 2018. METHODS: Using data from all countries receiving Gavi support between 2000 and 2020, we employed a synthetic control method to generate a pre-transition counterfactual with the same characteristics as the observation of interest to predict a future that empirically never existed. The synthetic unit is constructed from the weighted average of other units with good fit to the unit of interest before transition but did not transition. RESULTS: We found substantial heterogeneity after transitioning from Gavi assistance. China, Guyana, and Turkmenistan overperformed their expected coverage rates; Albania, Bhutan, China, Guyana, and Turkmenistan maintained coverage over 90%; and Bosnia and Herzegovina and Ukraine reported precipitous drop-offs that fell well below their synthetic controls. We also observed a vaccination coverage decline in 2020 for several countries after transitioning and most synthetic controls, which we attribute to COVID-19-related service disruptions. CONCLUSIONS: We recommend that Gavi adjust its transition model to systematically assess contextual externalities and risk. In addition, countries that no longer receive Gavi assistance can leverage technical assistance and communities of practice to mutually assist each other and other countries advancing toward transition. This could also foster intracountry accountability after transition. We also recommend that Gavi systematize post-transition assessments and evaluations that leverage the expertise and experience of graduated countries to encourage cross-learning.

Effectiveness of nutritional support to improve treatment adherence in patients with tuberculosis: a systematic review.

CONTEXT: Nutritional interventions substantially improve tuberculosis (TB) treatment outcomes and prevent complications. However, there is limited evidence about the connections between having nutritional support and TB treatment adherence. OBJECTIVE: The aim of this study was to determine the effectiveness of nutritional support in improving treatment adherence among patients with TB. DATA SOURCES: Databases, including PubMed, Embase (Ovid), Web of Science, and Scopus, were comprehensively reviewed to identify relevant studies reporting the impacts of nutritional support on TB treatment adherence. DATA EXTRACTION: Two authors independently screened the title, abstracts, and full article texts to identify eligible studies and assess the risk of bias. Observational and interventional studies were included. DATA ANALYSIS: A narrative synthesis approach was used to summarize the findings qualitatively. RESULTS: From the search, 3059 publications were identified; of these, 8 studies were included in this systematic review. Three types of nutritional interventions were identified: food baskets (eg, energy, micronutrient- or macronutrient-enriched food support), nutritional advice and guidance, and incentives for buying foods. Although 5 studies reported that nutritional support significantly improved treatment adherence in patients with TB, 3 studies showed that nutritional support had no effect on TB treatment adherence. CONCLUSIONS: Providing nutritional support may improve adherence to TB treatment. However, more well-powered, high-quality trials are warranted to demonstrate the effect of nutrition support on cost-effectively improving adherence to TB treatment. SYSTEMATIC REVIEW REGISTRATION: PROSPERO registration no. CRD42023392162.

Impact of COVID-19 on mental health in Bhutan: a way forward for action.

The coronavirus disease 2019 (COVID-19) pandemic continues to impact mental health and wellbeing globally. There is a lack of scientific documentation highlighting the mental health impact of COVID-19 in Bhutan. We present the mental health burdens and control measures taken, and suggest ways to further strengthen mental health services in Bhutan. During the pandemic, a rise in depression and anxiety had been reported in Bhutan. Depression rose from an average prevalence of 9 per 10,000 between 2011 and 2019 to 16 per 10,000 in 2020 and 32 per 10,000 in 2021. Similarly, anxiety rose from an average prevalence of 18 per 10,000, to 29 per 10,000 in 2020, and 55 per 10,000 in 2021. Psychological impacts related to isolation due to lockdowns, economic losses, and poor coping abilities were associated with negative outcomes. Stigma and discrimination towards mental health disorders discouraged mentally distressed people from seeking care. In response to increased demand, Bhutan's government initiated a range of interventions including home delivery of medicines and tele-counselling to people in need of urgent mental health care. Mental health care in Bhutan can be further improved through investment in services and human resources, and decentralization of services to the community.

Stigma and discrimination against transgender men in Bhutan.

BACKGROUND: While transgender people worldwide face high rates of stigma and discrimination, there are few studies of transgender men (also "trans men") in Asia. We measured the prevalence of, and factors associated with, stigma and discrimination faced by trans men in Bhutan to bring visibility to their experiences and inform health and social policy changes. METHODS: This cross-sectional survey was conducted in nine regions in Bhutan from November 2019 to January 2020. A total of 124 trans men were recruited using a hybrid venue-based and peer-referral approach. Data were collected using an interviewer-administered questionnaire. Multivariate logistic regression characterized associations with experiencing stigma and discrimination when accessing health services. FINDINGS: Participants were young (48.0% 18-24 years) and 48.4% had migrated from a rural to an urban area. The majority (95.2%) experienced stigma because people knew or thought they were trans men. Associations with frequent experiences of stigma were living with their partner as a couple (adjusted odds ratio [AOR] 3.07, 95% CI 1.27-7.44) and being unemployed or a student (3.22, 1.44-7.19). Nearly half (47.6%) said they experienced discrimination when accessing health care because people knew or thought they were a trans man; this experience was associated with migration (2.42, 1.08-5.39) and having >15 trans men in their social network (3.73, 1.69-8.26). Most (94.4%) experienced verbal violence, 10.5% experienced physical violence, and 4.8% experienced sexual violence. INTERPRETATION: Our study found high rates of stigma, discrimination, and interpersonal violence due to being a trans man in Bhutan. Findings highlight the urgent need for strengthening laws and regulations to protect the rights of transgender persons, particularly when accessing health services, recognizing partnerships, and preventing violence in public spaces.

Risk Factors of Symptomatic COVID-19 in Samtse District, Bhutan.

Introduction: All Coronavirus disease 2019 (COVID-19) positive cases in Samtse District, Bhutan were isolated in the isolation facilities managed by the government hospitals. This study aimed to identify the socio-demographic risk factors for developing COVID-19 symptoms amongst these patients. Methods and Materials: A secondary data of the COVID-19 positive cases from isolation facilities of Samtse District from 5 May to 7 September 2021 was used for this study. Survival analysis was carried out to estimate the cumulative probability of symptom onset time by each risk factor. Kaplan-Meier curves were used to estimate the probabilities for the onset of symptoms at different time points and a log-rank test was employed to assess the differences between covariates. Results: A total of 449 patients were included, of which 55.2% were males and 73.3% (328) were aged >18 years. The mean age was 42 years with a range of 3 months to 83 years. Forty-seven percent (213) reported at least one symptom. Common symptoms were fever (32.3%, 145), headache (31.6%, 142), and cough (30.1%, 135), respectively. Males were 64% less likely to be symptomatic than females [adjusted hazard ratio (aHR) = 0.36, 95% confidence interval (CI) 0.183-0.917]. Farmers (aHR = 3.17, 95% CI 1.119-8.953), and drivers and loaders (aHR = 3.18, 95% CI 1.029-9.834) were 3 times more likely to be symptomatic compared to housewives. Residents of Samtse sub-districts were 5 times more likely to be symptomatic than those living in other sub-districts (aHR = 5.16, 95% CI 2.362-11.254). Conclusion: The risk of developing COVID-19 symptoms was being fe male, farmers, drivers and loaders, and residents of the Samtse sub-district. These high-risk groups should be provided additional care when in isolation facilities.

Aetiologies and Risk Factors of Prolonged Fever Admission in Samtse Hospital, Bhutan, 2020.

Febrile illness is a common cause of hospital admission in developing countries, including Bhutan. Prolonged fever admission can add considerable strain on healthcare service delivery. Therefore, identifying the underlying cause of prolonged hospital stays can improve the quality of patient care by providing appropriate empirical treatment. Thus, the study's aims were to evaluate the aetiologies and factors of prolonged fever admission in Samtse Hospital, Bhutan. Fever admission data from 1 January to 31 December 2020 were retrieved from the Samtse Hospital database. Prolonged hospital stay was defined as those with >5 days of hospital admission. Univariable and multivariable logistic regression was used to identify risk factors for a prolonged hospital stay. Of 290 records, 135 (46.6%) were children (≤12 years), 167 (57.6%) were males, and 237 (81.7%) patients were from rural areas. The common aetiologies for fever admissions were respiratory tract infection (85, 29.3%) and acute undifferentiated febrile illness (48, 16.6%). The prolonged stay was reported in 87 (30.0%) patients. Patients from rural areas (adjusted odds ratio [AOR] = 4.02, 95% CI = 1.58−10.24) and those with respiratory tract infections (AOR = 5.30, 95% CI = 1.11−25.39) and urinary tract infections and kidney disease (AOR = 8.16, 95% CI = 1.33−49.96) had higher odds of prolonged hospital stay. This epidemiological knowledge on prolonged hospital stay can be used by the physician for the management of fever admission in Samtse Hospital.

Identifying socio-ecological drivers of common cold in Bhutan: a national surveillance data analysis.

The common cold is a leading cause of morbidity and contributes significantly to the health costs in Bhutan. The study utilized multivariate Zero-inflated Poisson regression in a Bayesian framework to identify climatic variability and spatial and temporal patterns of the common cold in Bhutan. There were 2,480,509 notifications of common cold between 2010 and 2018. Children aged < 15 years were twice (95% credible interval [CrI] 2.2, 2.5) as likely to get common cold than adults, and males were 12.4% (95 CrI 5.5%, 18.7%) less likely to get common cold than females. A 10 mm increase in rainfall lagged one month, and each 1 °C increase of maximum temperature was associated with a 5.1% (95% CrI 4.2%, 6.1%) and 2.6% (95% CrI 2.3%, 2.8%) increase in the risk of cold respectively. An increase in elevation of 100 m and 1% increase in relative humidity lagged three months were associated with a decrease in risk of common cold by 0.1% (95% CrI 0.1%, 0.2%) and 0.3% (95% CrI 0.2%, 0.3%) respectively. Seasonality and spatial heterogeneity can partly be explained by the association of common cold to climatic variables. There was statistically significant residual clustering after accounting for covariates. The finding highlights the influence of climatic variables on common cold and suggests that prioritizing control strategies for acute respiratory infection program to subdistricts and times of the year when climatic variables are associated with common cold may be an effective strategy.

The COVID-19 vaccination campaign in Bhutan: strategy and enablers.

Bhutan has reported a total of 2596 COVID-19 cases and three deaths as of September 15, 2021. With support from India, the United States, Denmark, the People's Republic of China, Croatia and other countries, Bhutan was able to conduct two rounds of nationwide vaccination campaign. While many countries struggle to overcome vaccine refusal or hesitancy due to complacency, a lack of trust, inconvenience and fear, escalated in some countries by anti-vaccine groups, Bhutan managed to inoculate more than 95% of its eligible populations in two rounds of vaccination campaign. Enabling factors of this successful vaccination campaign were strong national leadership, a well-coordinated national preparedness plan, and high acceptability of vaccine due to effective mass communication and social engagement led by religious figures, volunteers and local leaders. In this short report, we described the national strategic plan and enabling factors that led to the success of this historical vaccination campaign.

Further Characterization of MUAS35SCP and FUAS35SCP Recombinant Promoters and Their Implication in Translational Research.

Recombinant promoters are of high value in translational research. Earlier, we developed two recombinant promoters, namely MUAS35SCP and FUAS35SCP, and their transcriptional activities were found to be stronger than that of the most widely used CaMV35S promoter in dicot plants. Presently, we are reporting constitutive expression of both GUS and GFP reporters under the control of these promoters in several monocots, including rice, wheat, and pearl millet. We observed that these promoters could express the reporter genes constitutively, and their expression abilities were almost equal to that of the CaMV35S2 promoter. Plant-derived enriched PaDef (Persea americana var. drymifolia defensin) and NsDef2 (Nigella sativa L. defensin 2) antimicrobial peptides expressed under the control of these promoters arrest the growth of devastating phytopathogens like Pseudomonas syringae, Rhodococcus fascians, and Alternaria alternata. We observed that plant-derived NsDef2 and PaDef under control of these promoters showed approximately 80-90% inhibitory activity against Pseudomonas syringae. Hence, these promoters were constitutive and universal, as they can drive the expression of transgenes in both dicot and monocot plants. Alongside, these promoters could become a valuable tool for raising genetically modified plants with in-built resistance toward phytopathogens.

A cross-sectional study to assess the epidemiological situation and associated risk factors of dengue fever; knowledge, attitudes, and practices about dengue prevention in Khyber Pakhtunkhwa Province, Pakistan.

Background: Dengue fever has been responsible for around 12 countrywide large outbreaks in Pakistan, resulting in 286,262 morbidities and 1,108 deaths. Khyber Pakhtunkhwa (KP) is the most recently impacted province. This study aimed to investigate the molecular, epidemiological, and potential elements that contribute to increasing dengue transmission patterns, and knowledge, attitude, and practice (KAP) toward dengue in KP province. Method: This cross-sectional community-based study was conducted (June-December, 2021) in two phases. Phase I involved the epidemiological (n = 5,242) and molecular analysis of DENV in 500 randomly collected blood samples of the 2021 dengue outbreak in KP. Phase II focused on assessing dengue-KAP levels in healthy communities (n = 14,745, aged >18 years), adopting a cross-sectional clustered multistage sampling in eight districts (dengue-hotspot vs. non-hotspot) of KP. Chi-square tests and logistic regression analysis were applied. Results: Peshawar district had the highest dengue cases (60.0%) associated with the predominant co-circulation of DENV-2 (45.8%) and DENV-3 (50.4%) serotypes. A rise in cases was reported in October (41.8%) followed by September (27.9%) and August (14.4%; p < 0.001). Males (63.7%, p < 0.001) and individuals aged 16-30 years (37.0%, p < 0.001) were highly affected. General workers (18.0%), families with a monthly income of 10,000-20,000 Pak rupees (50.5%), unmarried (71.0%), uneducated (31%), families with higher human density (>10 individuals per household), and those (29.0%) who faced power outages for more than 7/24 h were the most affected. Moreover, co-morbidities like renal failure and bronchial asthma were associated with disease severity. A community survey on KAP revealed that an average of 74, 60, and 43% of the participants demonstrated good knowledge, attitudes, and dengue preventive practices, respectively. Conclusion: Multiple poor socioeconomic elements are influencing dengue fever transmission in the province. Higher KAP levels may explain the low frequency of dengue in non-hotspot districts. Our study emphasizes the need for effective and long-term public health education, strengthened vector surveillance, and expanded laboratory capacity for better diagnosis and management of dengue cases to better predict the burden and seasonality of disease in the country.

Population size estimation of transgender women and men in Bhutan.

INTRODUCTION: Transgender persons experience health disparities and are marginalized in many societies worldwide. Even their numbers are unknown in many countries. We conducted the first effort to estimate the population size of transgender women (TGW) and transgender men (TGM) in Bhutan from November 2019 to January 2020. METHODS: Community-based surveys of TGW and TGM integrated several methods to estimate the size of hidden populations, including key informant mapping, wisdom-of-the-crowd, the service multiplier, and the unique object multiplier. Results of the several methods were synthesized using a Bayesian approach. RESULTS: Surveys included 34 TGW and 124 TGM. TGW was persons assigned to the male sex at birth and currently self-identified as "trans women" (91%), "women" (6%), or another gender (3%). TGM were persons assigned female sex at birth and self-identified as "trans men" (100%). Bayesian synthesis of the multiple methods estimated 84 TGW (credible interval 61-110) and 166 TGM (credible interval 124-211) in Bhutan. CONCLUSIONS: Our study documented that TGW and TGM are part of Bhutanese society, with TGW constituting 0.03% of adult women and TGM 0.06% of adult men. Estimates can help advocate for resources and programs to address the health and well-being of these communities.

Survival Analysis of Symptomatic COVID-19 in Phuentsholing Municipality, Bhutan.

COVID-19 is a disease that is caused by a highly transmissible and pathogenic novel coronavirus: severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). All of the COVID-19 positive cases in Bhutanese travellers returning via the Phuentsholing point of entry, the local population, and Indian nationals were isolated in the Phuentsholing COVID-19 isolation ward, Bhutan. This study aimed to identify the risk factors for developing symptoms among COVID-19 positive patients in this ward. A retrospective cohort study was conducted using the data regarding COVID-19 positive cases in the Phuentsholing COVID-19 isolation ward from 28 May 2020 to 31 May 2021. The Cox proportional hazards regression model was used to identify the risk factors of developing COVID-19 symptoms. There were 521 patients in the study; 368 (70.6%) were males and 153 (29.4%) were females. The mean age was 32 years (with a range of 1-78 years), and 290 (56.0%) reported at least one symptom. The median length of isolation was eight days (with a range of 3-48 days). The common symptoms were: cough (162, 31.0%), fever (135, 26.0%), and headache (101, 19.0%). In the multivariable Cox regression, vaccinated patients were 77.0% (p = 0.047) less likely to develop symptoms compared to those who were not vaccinated. The front line workers and the mini-dry port (MDP) workers were 15 (p = 0.031) and 41 (p < 0.001) times more likely to be symptomatic compared to returning travellers. The young and economically active population group was most commonly affected by COVID-19. The presence of risk factors, such as being front line workers, MDP workers, or not being vaccinated against COVID-19, meant that patients had a higher probability of developing symptoms of COVID-19.