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The North American Animal Disease Spread Model ( NAADSM) is a stochastic model framework developed to simulate the spread of highly contagious diseases of livestock and poultry, such as foot-and-mouth disease and highly pathogenic avian influenza ( HPAI). The objective of this study was to make recommendations on the most effective HPAI control policy for Canada, specifically, on the effect of different speeds of detection, effectiveness of movement restrictions and stamping-out and ring-culling strategies on the magnitude of an HPAI outbreak. In addition, the effect of introduction of infection in a range of multiple farms simultaneously was also evaluated. A total of 21 060 scenarios, defined as different combinations of parameters for various epidemiological conditions and control measures, were created to simulate the number of poultry flocks that would become infected as a result of an incursion of HPAI. Each scenario was parameterized in NAADSM and replicated 1000 times, generating the median number of flocks infected at the end of the simulated outbreak for each scenario. Negative binomial regression analysis was used to model significant explanatory variables of the median number of flocks infected at the end of each simulated outbreak for each of the 21 060 scenarios. The final model included the following explanatory variables: number and type initially infected flock(s), density of flocks within the county where the initially infected flock(s) was located, probability of transmission through indirect contact, subclinical spread of the infection, speed of detection and a two-way interaction between intensity of bird destruction strategy and movement restriction effectiveness to reduce transmission through direct and indirect contacts. The modelling results suggested that stamping out of the detected infected flocks, without ring culling, in combination with effective movement restrictions on direct and indirect contacts, would be the most appropriate policy for Ontario. [ABSTRACT FROM AUTHOR]

Background: The association between covid-19 vaccine and menstrual disturbance is unclear., Methods: An in-person cross-sectional survey among female members ≥ 18 years enrolled in an ongoing Zero TB prospective cohort in Northern India who had received one or two doses of covid-19 vaccine was conducted to study the characteristics and association of menstrual disturbance within six months of receiving Covishield., Results: Between June 29 and September 5, 2021, 339 females ≥ 18 years of age were administered the survey. Median age was 30 (IQR: 22-39) years; 84 % were between 18 and 49 and 16 % were ≥ 50 years old. There were 152 college students, 27 healthcare workers, and 160 nuns. Forty-two women (12 %) had received one dose and 297 (88 %) had received two doses of Covishield. Overall, 66 (20 %) women reported experiencing menstrual disturbance after receiving Covishield vaccine. The problems included early menstruation: 6 % (n = 19/339); late menstruation: 4 % (n = 14/339); and heavier bleeding: 5 % (n = 17/339). Disturbances lasted for less than seven days and cycles normalized in 1-3 months. There was no post-menopausal bleeding. There was no significant difference in menstrual disturbance based on receiving one vs. two doses of Covishield (OR: 1.58; 95 % CI: 0.55-4.57; p = 0.381). History of SARS-CoV-2 infection was not associated with the development of menstrual disturbance among the vaccinees (OR: 0.63; 95 % CI: 0.24-1.73; p = 0.379). Presence of emotional disturbance at baseline (OR: 31; 95 % CI: 3.52-267; p = 0.002) or previous history of dysmenorrhea (OR: 41; 95 % CI: 8.7-196; p < 0.001) was associated with menstrual disturbance in the vaccinees, indicating their potential to confound or bias study results., Conclusion: Menstrual problems were reported by Covishield vaccinees, but they were minor and reversible within three months and do not constitute a ground for vaccine hesitancy. Studies designed to assess causal link taking care to avoid selection bias or confounding are needed., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Leishmaniasis is considered one of the neglected tropical diseases in the world. Although Bhutan is a member of the visceral leishmaniasis elimination consortium in South Asia, not much attention has been accorded to the disease because of its low incidence. The vector that transmits Leishmania remains poorly understood. In this backdrop, sand-fly surveys were regularly conducted at multiple sites where leishmaniasis cases have been reported in Bhutan. Collections were made using CDC light traps and cattle-baited net traps in 15 villages from February 2019 to May 2022. Six species of Phlebotomus and four species of Sergentomyia were identified from these sites that included two Phlebotomus and three Sergentomyia species discovered for the first time in Bhutan. Sand-fly density varied significantly from village to village, and it showed strong seasonality with peak numbers collected from June to October and almost zero from December to February. Overall, sand-fly density was highest in the basements of the houses and were higher outdoors than indoors. Cattle-baited net traps collected few sand flies during the surveillance period. This work constituted the first systematic sand-fly population surveillance conducted in Bhutan and will provide a baseline for future vector ecology and Leishmania epidemiological studies.

Background: Recently, extra-pulmonary tuberculosis (EPTB) has been increasingly reported in Bhutan, rising from 26% in 2001 to 39% in 2010. In 2016, almost half (49%) of all reported TB cases were classified as EPTB. Thus, this study was conducted to understand the epidemiology and identify risk factors contributing to increasing notification of EPTB in Bhutan., Methods: A case-control study was conducted by recruiting all 110 cases of nationally notified EPTB (Extrapulmonary bacteriologically confirmed/EPBC) as cases and 235 Pulmonary TB (Pulmonary bacteriologically confirmed/PBC) as controls. Data were collected on socio-demography, clinical and diagnostic, behavioral and lifestyle and environmental exposures using a structured questionnaire between April and September, 2018., Results: The median age of the cases was 25 years, ranged 9-79 years. Lymphatic TB was predominant (n = 78; 70.91%) followed by genitourinary TB (n = 4 (3.64%). The likelihood of EPTB decreased with increase in age (p = 0.023). The odds of EPTB in females was 1.65 times higher than the males (p = 0.038). Increase in Body Mass Index (adjusted odds ratio (AOR): 1.1; 95% confidence interval (CI): 1.052-1.200) and urban residency were (AOR 1.6; 95% CI 1.016-2.805) were found to have higher odds of developing EPTB., Conclusion: Females, urban residents, and those with higher BMI are at increased risk of developing EPTB. These at-risk groups can be used to target limited public health resources to control EPTB in Bhutan., Competing Interests: Conflict of interest The authors have none to declare., (Copyright © 2023 Tuberculosis Association of India. Published by Elsevier B.V. All rights reserved.)

Leishmaniases are a group of diseases under the category of neglected tropical diseases targeted for global elimination. However, they continue to pose major clinical and public health problems, especially among those living in poor socioeconomic conditions. Here, we summarize leishmaniasis elimination efforts in Bhutan. Between 1994 and 2022, Bhutan recorded 54 cases of leishmaniasis across 14 of its 20 districts. There are seven species of Phlebotomus and three species of Sergentomyia sand flies documented in the country. At a subnational level, all endemic districts recorded a visceral leishmaniasis annual incidence <1 per 10,000 population, meeting the regional elimination targets. Serological testing with ELISA and molecular testing with polymerase chain reaction were established at the Royal Center for Disease Control in 2022. A leishmaniasis prevention and management guideline was adopted in 2023 to aid clinicians in diagnosis and management. Active and passive case surveillance was integrated with the national infectious disease early warning and response system. Risk-based entomological surveillance and control have also been prioritized. Climate change may play a major role in rendering districts in the temperate zone favorable for vector proliferation. The country's medical university introduced a diploma course in medical entomology in 2023 to augment the human resources needed for vector surveillance efforts. However, leishmaniasis elimination lacks dedicated programmatic management amid competing priorities for resources against other infectious diseases. Leishmaniasis elimination requires a targeted and programmatic approach in Bhutan, including cross-border collaborative efforts with neighboring Indian states. Bhutan remains highly committed to achieving leishmaniasis elimination targets.

Background: Rabies continues to pose significant public health challenges in many developing countries including Bhutan. A probable case of rabies was admitted to our hospital and its reporting led to the uncovering of an outbreak in domestic and wild animals. We discuss the challenges in the diagnosis and management of rabies in a resource-limited setting., Case Presentation: A 35-year-old male presented with intermittent fever, bilateral lower limb weakness that was rapidly progressive, urinary incontinence with episodes of palpitations and sweating. He had sustained a Category III bite on the right lower thigh with four bite marks, inflicted by a stray dog. He had received post-exposure prophylaxis with intra-dermal anti-rabies vaccine. On initial examination, the patient was in distress but cooperative for the interview. He had pulse rate ranging from 60 to 100/min with episodes of diaphoresis and palpitations, but with normal capillary blood glucose. In the lower limb, the muscle power was zero with absent tendon reflexes in the lower limb and impaired abdominal reflex below T 10 level. He had hyperaesthesia below T 8 , hydrophobia, aerophobia and photophobia. He had multiple spontaneous fasciculations in both the thighs and right deltoid and these later involved the intercostal muscles, neck and face muscles. He had altered sensorium and desaturation for which he required mechanical ventilation. Polymerase chain reaction for rabies virus was negative in cerebrospinal fluid and saliva. Rabies virus neutralizing antibody was negative in cerebrospinal fluid but had high titres in the serum. He received Human Rabies Immunoglobulin after admission. He was managed in the intensive care unit and died 23 days later. After this case was notified, a rapid response team was deployed in the field, and uncovered rabies outbreak in animals in the locality., Conclusions: This case called for a serious evaluation of the country's efforts in achieving zero rabies deaths by 2030. The management of this case identified several critical areas of context-specific interventions in Bhutan. There is also an urgent need to improve diagnostic capabilities at the national reference laboratory and enhance the technical competencies of healthcare workers in the management of dog bite cases., (© 2023. National Institute of Parasitic Diseases.)

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.

Poor maternal mental health negatively impacts cognitive development from infancy to childhood, affecting both behavior and brain architecture. In a non-western context (Thimphu, Bhutan), we demonstrate that culturally-moderated factors such as family, community social support, and enrichment may buffer and scaffold the development of infant cognition when maternal mental health is poor. We used eye-tracking to measure early building blocks of cognition: attention regulation and social perception, in 9-month-old Bhutanese infants (N = 121). The cognitive development of Bhutanese infants in richer social environments was buffered from poor maternal mental health, while for infants in environments with lower rates of protective social environment factors, worse maternal mental health significantly predicted greater costs for infant attention, a fundamental building block cognition. International policies and interventions geared to improve maternal mental health and child health outcomes should incorporate each regions' unique family, cultural, and community support structures., (© 2021 The Authors. Developmental Science published by John Wiley & Sons Ltd.)

Background: Delayed childbearing has been noted in a high percentage of women with a previous Caesarean section (CS). Many women with CS scar defects (CSDs) present with clinical symptoms of irregular vaginal bleeding. The present study aimed to investigate bacterial colonies at CSDs in women suffering from secondary infertility., Methods: This observational study included 363 women with secondary infertility who visited the Assisted Reproduction Unit between 2008 and 2013. Among them, 172 women with a previous CS and 191 women with no previous CS were approached. The women with a previous CS had their CS operations in the past 1 to 14 years, with a mean of 3.5 years. The presence of CSDs was detected by vaginal ultrasonography. Bacteriology cultures of specimens taken from the uterine niches in those with CSDs were collected during Day 7 to Day 10 of the follicular phase. Specimens were obtained from the endocervical canal for bacterial culture in those without CSDs. The main outcome measure was the detection of the growth of bacterial colonies., Results: CSDs were found in 60.4% (96 of 159) of women with a previous CS. In women with a previous CS, bacterial colonies were identified in 89.6% (86 of 96) and 69.8% (44 of 63) of women with and without CSDs, respectively. In women with no previous CS, 49.7% (88 out of 177) of bacterial cultures of endocervical samples showed bacterial colony growth. Gram-positive cocci (P = 0.0017, odds ratio (OR) = 1.576, 95% confidence intervals (CI) -22.5 to - 5.4) and Gram-negative rods (P = 0.0016, OR = 1.74, CI - 20.8 to - 5.0) were the most commonly isolated bacteria and contributed to approximately 90% of all microorganisms found in those with a previous CS. In women with a previous CS, more Gram-negative rods were isolated (P = 0.01, OR = 1.765, CI - 27.2 to - 3.8), especially Pseudomonas species (P = 0.02, OR = 1.97, CI - 16.7 to - 1.0), in those with visible CSDs than in those without CSDs., Conclusions: Bacterial colonization at CSDs was found in a high percentage of women with secondary infertility., (© 2022. The Author(s).)

Context: Gonadotropins can be administered every 5 days under intradermal injection in in vitro fertilization (IVF) treatment., Objective: To explore the effectiveness of intradermal injection of recombinant human FSH (rhFSH) for women undergoing IVF., Methods: Women who received their first IVF treatment enrolled in this prospective intervention in 2018. All women received a bolus of 900 IU rhFSH intradermally at day 2 of the treatment cycle followed by additional dosage of rhFSH at day 7 and/or day 10. The main outcome measures included the total dose of rhFSH and number of injections required, sequential serum FSH level detected, and number of mature oocytes retrieved., Results: Seventy women completed the study. On average, 2.31 ± 0.73 injections and 1662 ± 397 IU of rhFSH were administered. While the baseline FSH level was 5.6 ± 2.2 IU/L, the serum concentrations of FSH after rhFSH administration were 35.3 ± 7.0 on the first day (24 hours) and 10.7 ± 3.7 IU/L on the fifth day (120 hours). A total of 10.5 ± 6.6 mature oocytes were retrieved, resulting in 7.3 ± 5.1 pronuclear embryos; 1.8 ± 0.6 embryos were transferred to the uterus. Our findings resulted in 72% fertilization, 91% cleavage, 31% implantation, and 36% live birth rates. Although fewer larger follicles were found, noninferiority results were noted in the mature oocytes retrieved, good embryos available, and clinical pregnancy rate compared with those received conventional daily subcutaneous rhFSH administration., Conclusion: Intradermal administration of rhFSH, with a smaller dose of rhFSH and fewer injections, may achieve the goal of a cost-effective and more patient-friendly regimen., (© The Author(s) 2021. Published by Oxford University Press on behalf of the Endocrine Society. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

The veterinary prescription of antibiotics in low- and middle-income countries (LMIC) remains largely undocumented. In Bhutan, however, the national veterinary service keeps records of their activities and prescriptions, which offer an opportunity to establish a benchmark to assess the use of these agents in this and other LMIC. A cross-sectional retrospective study was designed and 2,266 handwritten veterinary records from 2017 were sampled from 23 animal health premises (AHPs) to estimate individual and an overall proportion of consultations that resulted in an antibiotic prescription. The frequency of antibiotic prescription per species, type of AHP, and according to WHO's AWaRe index and OIE list of priority antimicrobials were also explored. It was estimated that 31% (95% confidence interval: 29-33%; intracluster correlation: 0.03) of the veterinary consultations resulted in an antibiotic prescription. The incidence of antibiotic prescription was highest in consultations of poultry across AHP. Across species, diarrhea and wounds were frequently treated with broad-spectrum antibiotics including sulfonamides, tetracyclines, trimethoprim + sulfa, and penicillin. Between 45% and 70% antibiotics prescribed correspond to AWaRe's access group and up to 25% to AWaRe's watch group . Over 70% of antibiotics dispensed in veterinary consultations for any species correspond to the OIE's veterinary critically important antimicrobial agents . Overall, the study demonstrated positive features of veterinary antimicrobial stewardship in Bhutan, given the conservative proportion of consultation that results in this type of prescription and the type of antibiotic prescribed. Although the veterinary service closely follows the Bhutanese Standard Treatment Guidelines, the prescription of antibiotics to key species should be closely monitored. Our study suggests that further improvements of antibiotic stewardship can be achieved through standardisation of antibiotic prescription to some species, a revision of the guidelines toward reducing the prescription of antibiotics of high relevance for human medicine, and by including details of clinical investigation, use of tests, and treatment outcomes in veterinary consultation records., Competing Interests: 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 © 2021 Villanueva-Cabezas, Rinzin, Dorjee, Tshewang, Namgyel, Sharma, Stevenson and McVernon.)

Objective: This study was designed to investigate if whole dimension subcortical ovarian administration of platelet-rich plasma with gonadotropin, in proximity to most ovarian follicles, is effective in restoring ovarian functions in women during early menopause., Methods: Platelet-rich plasma, prepared from 40 mL of autologous peripheral blood using the buffy coat method, was injected into extended subcortical area of bilateral ovaries along with recombinant follicle-stimulating hormone (rFSH) (Gonal-F 300IU) under laparoscopic guidance. The posttreatment ovarian folliculogenesis and serum levels of FSH, luteinizing hormone (LH), and estradiol were followed up for 6 months at weekly to monthly intervals. IVF was carried out in women resuming ovulatory functions., Results: Twelve early menopausal women with mean age of 44.42 ± 2.84 were enrolled. After treatment, 11 women resumed their menstrual period in 37.1 ± 23.5 days. Their average serum FSH was 70.47 ± 20.92 and 26.22 ± 17.55 IU/L, luteinizing hormone was 34.81 ± 11.86 and 14.3 ± 12.8 IU/L, before and after treatment, respectively. The mid-cycle E2 was 251.1 ± 143.8 pg/mL. Ten oocyte retrievals were carried out among six participants, four of them received controlled ovarian stimulation and another two using natural ovulation cycles. Thirteen mature eggs were retrieved which were then ICSI fertilized to obtain 10 normally fertilized 2PN oocytes. Two participants had cleavage stage embryos transferred of which one achieved clinical pregnancy., Conclusions: Whole dimension subcortical ovarian administration of platelet-rich plasma with gonadotropin was shown to restore ovarian functions, at least temporarily, and could increase the probability of pregnancy using autologous oocytes in women with early menopause., Competing Interests: Financial disclosures/conflicts of interest: None reported., (Copyright © 2021 by The North American Menopause Society.)

Livestock farming plays an important role in supporting the livelihood of resource-poor subsistence farmers in Bhutan. However, ticks and tick-borne diseases (TBDs) are one of the major constraints to livestock farming due to their negative effect on health and production. To date, no study has been conducted in Bhutan to assess farmers' knowledge, attitude, and practices (KAP) about ticks and TBDs in cattle, although such information is essential in ensuring the development and adoption of effective prevention and control measures. Therefore, a KAP survey was conducted among 246 cattle owners in the Samkhar sub-district of eastern Bhutan in June 2019, using a structured questionnaire. Based on our scoring criteria, 52% [95%CI: 45.5-58.4] had adequate knowledge about ticks as potential vectors of diseases. Logistic regression analysis showed that the individuals who practiced a stall-feeding system of cattle rearing were 2.8 times [OR = 2.8 (95%CI: 1.66-4.78)] more likely to have adequate knowledge than others. Sixty-eight percent [95%CI: 62.5-74.4] had a favorable attitude toward tick prevention and control programs. Men were 1.95 times [OR = 1.95 (95%CI: 1.09-3.55)] more likely to have a favorable attitude than women, and the individuals who practiced a stall-feeding system were 2.59 times [OR = 2.59 95%CI: 1.45-4.78)] more likely to have a favorable attitude than others, after adjusting for the effect of other variables in the model. Overall, only 38% [95%CI 32.5-45] of the respondents reported tick infestation as one of the most important animal health problems, but 100% reported using acaricides to control ticks in cattle. Despite a high level of acaricide usage, the level of knowledge was low among the farmers interviewed. Findings from this study underline the importance of considering identified knowledge gaps and initiating education efforts to improve the adoption of effective tick prevention and control measures among farmers., Competing Interests: The authors have declared that no competing interests exist.

Tick infestation is the most reported parasitological problem in cattle in Bhutan. In May and June 2019, we collected ticks from 240 cattle in two districts of Eastern Bhutan. Tick presence, diversity, and infestation prevalence were examined by morphological identification of 3600 live adult ticks. The relationships between cattle, geographic factors, and infestation prevalence were assessed using logistic regression analyses. Habitat suitability for the tick species identified was determined using MaxEnt. Four genera and six species of ticks were found. These were Rhipicephalus microplus (Canestrini) (70.2% (95% confidence interval (CI): 68.7-71.7)), Rhipicephalus haemaphysaloides Supino (18.8% (95% CI: 17.5-20.1)), Haemaphysalis bispinosa Neumann (8.2% (95% CI: 7.3-9.1)), Haemaphysalis spinigera Neumann (2.5% (95% CI: 2-3)), Amblyomma testudinarium Koch (0.19% (95% CI: 0.07-0.4)), and a single unidentified Ixodes sp. Logistic regression indicated that the variables associated with infestation were: longitude and cattle age for R. microplus ; latitude for R. haemaphysaloides ; and altitude and cattle breed for H. bispinosa and H. spinigera . MaxEnt models showed land cover to be an important predictor for the occurrence of all tick species examined. These findings provide information that can be used to initiate and plan enhanced tick surveillance and subsequent prevention and control programs for ticks and tick-borne diseases in cattle in Bhutan.

Leishmaniasis is a neglected tropical disease with an annual estimated 700,000 to 1 million new cases globally. The diseases affect the rural community and people living in poverty. It is transmitted by female phlebotomine sand-fly; a tiny 2-3 mm long insect vector. Along with the regional countries, Bhutan has embarked on the elimination goal. However, several challenges lies ahead on its path to elimination. There is no elimination strategy and actions. The reporting, surveillance system, control and prevention mechanisms are inadequate. Further, there is lack of knowledge among the health care providers that impedes elimination goals. Therefore, if Bhutan is serious about its elimination goal, all the gaps and current challenges needs to be addressed appropriately.

Background: Head and neck cancers are among the commonest cancers in the developing world. Personal habits, such as the use of tobacco, betel nut and alcohol are strongly associated with the development of head and neck cancers at certain sites. Therefore, they are among the preventable cancers. In Bhutan, there has not yet been a study conducted on head and neck cancers., Objective: To describe baseline epidemiological and histopathological characteristics of head and neck cancers in Bhutan., Methods: This is a 7-year descriptive study of all cases of head and neck cancers presented at the Jigme Dorji Wangchuk National Referral Hospital from 2011 to 2017. The data were collected from the hospital's medical records section, histopathology records, patient referral unit and some treatment centres in India. Prior approval was sought from the Research and Ethics Board for Health, the Ministry of Health and the hospital management., Results: There were a total of 515 cases of head and neck cancers from 2011 to 2017. The crude incidence rate was 10 per 100,000 and the overall age adjusted rate was 12.3 (95% CI 9.5-15.1) per 100,000 population. The prevalence during this 7-year period was 69.1 per 100,000 population. The commonest cancers are thyroid, oral cavity, hypopharyngeal, laryngeal and nasopharyngeal cancer in decreasing order. Head and neck cancers are more common in males than females in the majority of sites except thyroid, salivary gland and sinonasal malignancies. Thyroid cancers and nasopharyngeal cancers are found to affect younger age groups. Tashigang (48) followed by Paro (43) recorded the highest number of cases. Squamous cell carcinoma is the commonest histopathology type in almost all the cases, while papillary carcinoma is the commonest among thyroid cancers. Personal habits, such as smoking, chewing tobacco, betel nut and alcohol consumption, were found to be more common among patients suffering from oral cavity, laryngeal, hypopharyngeal and oropharyngeal cancers., Conclusion: Head and neck cancers are the third most common cancer in Bhutan after stomach cancer and cervical cancer. Thyroid, oral cavity and hypopharynx are the top three anatomical sites for head and neck cancers in Bhutan. The current epidemiological and histopathological profile of head and neck cancers will form a baseline of information and basis for further research on head and neck cancers in Bhutan., Competing Interests: None declared., (© the authors; licensee ecancermedicalscience.)

Background: Rabies is endemic in southern Bhutan, associated with 1-2 human deaths and high post exposure prophylaxis (PEP) costs annually. Evaluation of clinicians' management of human cases potentially exposed to rabies could contribute to improving PEP prescribing practices to both reduce unnecessary costs associated with PEP and reach the target of zero human deaths due to rabies by 2023., Methods: A cross-sectional survey of 50 clinicians' management of human cases potentially exposed to rabies was conducted in 13 health centers in high-rabies-risk areas of Bhutan during February-March 2016., Results: Data were collected on clinicians' management of 273 human cases potentially exposed to rabies. The 50 clinicians comprised health assistants or clinical officers (55%) and medical doctors (45%) with a respective median of 19, 21 and 2 years' experience. There was poor agreement between clinicians' rabies risk assessment compared with an independent assessment for each case based on criteria in the National Rabies Management Guidelines (NRMG). Of the 194 cases for which clinicians recorded a rabies risk category, only 53% were correctly classified when compared with the NRMG. Clinicians were more likely to underestimate the risk of exposure to rabies and appeared to prescribe PEP independently of their risk classification.. Male health assistants performed the most accurate risk assessments while female health assistants performed the least accurate. Clinicians in Basic Health Units performed less accurate risk assessments compared with those in hospitals., Conclusions: This study highlights important discrepancies between clinicians' management of human cases potentially exposed to rabies and recommendations in the NRMG. In particular, clinicians were not accurately assessing rabies risk in potentially exposed cases and were not basing their PEP treatment on the basis of their risk assessment. This has significant implications for achieving the national goal of eliminating dog-mediated human rabies by 2030 and may result in unnecessary costs associated with PEP. Recommendations to improve clinician's management of human cases potentially exposed to rabies include: reviewing and updating the NRMG, providing clinicians with regular and appropriately targeted training about rabies risk assessment and PEP prescription, and regularly reviewing clinicians' practices.

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.

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.

Introduction: Considerable advocacy, funding, training, and technical support have been provided to South Asian countries to strengthen One Health (OH) collaborative approaches for controlling diseases with global human pandemic potential since the early 2000s. It is essential that the OH approach continues to be strengthened given South Asia is a hot spot for emerging and endemic zoonotic diseases. The objectives of this article are to describe OH research and training and capacity building activities and the important developments in government support for OH in these countries to identify current achievements and gaps., Materials and Methods: A landscape analysis of OH research, training, and government support in South Asia was generated by searching peer-reviewed and grey literature for OH research publications and reports, a questionnaire survey of people potentially engaged in OH research in South Asia and the authors' professional networks., Results: Only a small proportion of zoonotic disease research conducted in South Asia can be described as truly OH, with a significant lack of OH policy-relevant research. A small number of multisectoral OH research and OH capacity building programmes were conducted in the region. The governments of Bangladesh and Bhutan have established operational OH strategies, with variable progress institutionalising OH in other countries. Identified gaps were a lack of useful scientific information and of a collaborative culture for formulating and implementing integrated zoonotic disease control policies and the need for ongoing support for transdisciplinary OH research and policy-relevant capacity building programmes., Discussion: Overall we found a very small number of truly OH research and capacity building programmes in South Asia. Even though significant progress has been made in institutionalising OH in some South Asian countries, further behavioural, attitudinal, and institutional changes are required to strengthen OH research and training and implementation of sustainably effective integrated zoonotic disease control policies., Competing Interests: and funding The authors have not received any funding or benefits from industry or elsewhere to conduct this study.

In Bhutan, Capture-Neuter-Vaccinate-Release (CNVR) programs have been implemented to manage the dog population and control rabies, but no detailed evaluation has been done to assess their coverage and impact. We compared estimates of the dog population using three analytical methods: Lincoln-Petersen index, the Chapman estimate, and the logit-normal mixed effects model, and a varying number of count periods at different times of the day to recommend a protocol for applying the mark-resight framework to estimate free-roaming dog population abundance. We assessed the coverage of the CNVR program by estimating the proportion of dogs that were ear-notched and visually scored the health and skin condition of free-roaming dogs in Gelephu and Phuentsholing towns in south Bhutan, bordering India, in September-October 2012. The estimated free-roaming dog population in Gelephu using the Lincoln-Petersen index and Chapman estimates ranged from 612 to 672 and 614 to 671, respectively, while the logit-normal mixed effects model estimate based on the combined two count events was 641 (95% CI: 603-682). In Phuentsholing the Lincoln-Petersen index and Chapman estimates ranged from 525 to 583 and 524 to 582, respectively, while the logit-normal mixed effects model estimate based on the combined four count events was 555 (95% CI: 526-587). The total number of dogs counted was significantly associated with the time of day (AM versus PM; P=0.007), with a 17% improvement in dog sightings during the morning counting events. We recommend to conduct a morning marking followed by one count event the next morning and estimate population size by applying the Lincoln-Peterson corrected Chapman method or conduct two morning count events and apply the logit-normal mixed model to estimate population size. The estimated proportion of vaccinated free-roaming dogs was 56% (95% CI: 52-61%) and 58% (95% CI: 53-62%) in Gelephu and Phuentsholing, respectively. Given coverage in many neighbourhoods was below the recommended threshold of 70%, we recommend conducting an annual "mass dog vaccination only" campaign in southern Bhutan to create an immune buffer in this high rabies-risk area. The male-to-female dog ratio was 1.34:1 in Gelephu and 1.27:1 in Pheuntsholing. Population size estimates using mark-resight surveys has provided useful baseline data for understanding the population dynamics of dogs at the study sites. Mark-resight surveys provide useful information for designing and managing the logistics of dog vaccination or CNVR programs, assessing vaccination coverage, and for evaluating the impact of neutering programs on the size and structure of dog populations over time., (Copyright © 2015. Published by Elsevier B.V.)

Increasing incidences of emerging and re-emerging diseases that are mostly zoonotic (e.g. severe acute respiratory syndrome, avian influenza H5N1, pandemic influenza) has led to the need for a multidisciplinary approach to tackling these threats to public and animal health. Accordingly, a global movement of 'One-Health/One-Medicine' has been launched to foster collaborative efforts amongst animal and human health officials and researchers to address these problems. Historical evidence points to the fact that pandemics caused by influenza A viruses remain a major zoonotic threat to mankind. Recently, a range of mathematical and computer simulation modelling methods and tools have increasingly been applied to improve our understanding of disease transmission dynamics, contingency planning and to support policy decisions on disease outbreak management. This review provides an overview of methods, approaches and software used for modelling the spread of zoonotic influenza viruses in animals and humans, particularly those related to the animal-human interface. Modelling parameters used in these studies are summarized to provide references for future work. This review highlights the limited application of modelling research to influenza in animals and at the animal-human interface, in marked contrast to the large volume of its research in human populations. Although swine are widely recognized as a potential host for generating novel influenza viruses, and that some of these viruses, including pandemic influenza A/H1N1 2009, have been shown to be readily transmissible between humans and swine, only one study was found related to the modelling of influenza spread at the swine-human interface. Significant gaps in the knowledge of frequency of novel viral strains evolution in pigs, farm-level natural history of influenza infection, incidences of influenza transmission between farms and between swine and humans are clearly evident. Therefore, there is a need to direct additional research to the study of influenza transmission dynamics in animals and at the animal-human interface., (© 2012 Blackwell Verlag GmbH.)

Problem: To develop an immunohistochemical assay for determination of acrosome-reacted human sperm and to study the effects of progesterone and cholesterol treatment on human sperm acrosome reaction., Method of Study: Three distinct anti-sperm monoclonal antibodies were biotinylated and used as probes for assessment of acrosome reaction in a 30-min immunohistochemical assay. Progesterone and/or cholesterol were added to sperm preparation to influence the acrosome reaction in different experimental conditions., Results: Percentages of acrosome-intact sperm decreased significantly during the 18-hr incubation. Acrosome reaction could be induced by progesterone as early as 2 hr after sperm incubation in human tubal fluid. The degree of progesterone-induced acrosome reaction was time dependent and the optimal effect was reached by adding 10 micrograms/ml progesterone for 30 min incubation. Progesterone-induced acrosome reaction was shown to be hormone-concentration dependent with 50% stimulation at 1 microgram/ml. Cholesterol (1 microgram/ml) was found to inhibit progesterone-induced acrosome reaction either by co-incubation with human sperm during capacitation, or by simultaneous incubation with progesterone during acrosome reaction induction., Conclusions: Assessment of human sperm acrosomal status by avidin-biotin immunohistochemical assay can be a routine in clinical laboratories for male infertility services. Cholesterol can inhibit progesterone-induced acrosome reaction, possibly by its modifications of sperm plasma membrane and/or interference of progesterone binding to its surface receptors.

Among the numerous anti-sperm monoclonal antibodies generated in our laboratory, HS-33 was shown to react with a conserved antigen on the acrosome of spermatozoa from human and mouse. By using indirect immunofluorescent assay, it was demonstrated that HS-33 did not bind to live human sperm. However, this antibody was found to react with the methanol-fixed acrosome-intact, but not with acrosome-reacted sperm. The human sperm antigen recognized by this antibody was purified from human sperm extract by immunoaffinity chromatography. The purified cognate human sperm antigen designated as HSAg-33 was found to be a protein with a molecular weight of approximately 72 kDa on sodium dodecyl sulfate polyacrylamide gel electrophoresis under reducing conditions. The tissue-specificity and the developmental expression of this sperm antigen were examined using frozen sections of various human and mouse tissues. The antigen was shown to be expressed specifically in the testicular sperm at the postmeiotic stages of spermatogenesis but not in any other somatic tissues. "Spontaneous" acrosome reaction was determined following 18 hours of incubation in Biggers, Whitten, and Whittingham (BWW) medium by using HS-33 monoclonal antibody and Pisum sativum agglutinin (PSA) as probes. The number of sperm stained positively with this antibody decreased significantly following overnight incubation, indicating the occurrence of an acrosome reaction. The results of this study suggest that HSAg-33 is a potentially useful sperm-specific acrosome marker for studies of sperm capacitation and acrosome reaction.