Discovering disease-causing pathogens in resource-scarce Southeast Asia using a global metagenomic pathogen monitoring system.

SignificanceMetagenomic pathogen sequencing offers an unbiased approach to characterizing febrile illness. In resource-scarce settings with high biodiversity, it is critical to identify disease-causing pathogens in order to understand burden and to prioritize efforts for control. Here, metagenomic next-generation sequencing (mNGS) characterization of the pathogen landscape in Cambodia revealed diverse vector-borne and zoonotic pathogens irrespective of age and gender as risk factors. Identification of key pathogens led to changes in national program surveillance. This study is a "real world" example of the use of mNGS surveillance of febrile individuals, executed in-country, to identify outbreaks of vector-borne, zoonotic, and other emerging pathogens in a resource-scarce setting.

Molecular markers of artemisinin resistance during falciparum malaria elimination in Eastern Myanmar.

BACKGROUND: Artemisinin resistance in Plasmodium falciparum threatens global malaria elimination efforts. To contain and then eliminate artemisinin resistance in Eastern Myanmar a network of community-based malaria posts was instituted and targeted mass drug administration (MDA) with dihydroartemisinin-piperaquine (three rounds at monthly intervals) was conducted. The prevalence of artemisinin resistance during the elimination campaign (2013-2019) was characterized. METHODS: Throughout the six-year campaign Plasmodium falciparum positive blood samples from symptomatic patients and from cross-sectional surveys were genotyped for mutations in kelch-13-a molecular marker of artemisinin resistance. RESULT: The program resulted in near elimination of falciparum malaria. Of 5162 P. falciparum positive blood samples genotyped, 3281 (63.6%) had K13 mutations. The prevalence of K13 mutations was 73.9% in 2013 and 64.4% in 2019. Overall, there was a small but significant decline in the proportion of K13 mutants (p < 0.001). In the MDA villages there was no significant change in the K13 proportions before and after MDA. The distribution of different K13 mutations changed substantially; F446I and P441L mutations increased in both MDA and non-MDA villages, while most other K13 mutations decreased. The proportion of C580Y mutations fell from 9.2% (43/467) before MDA to 2.3% (19/813) after MDA (p < 0.001). Similar changes occurred in the 487 villages where MDA was not conducted. CONCLUSION: The malaria elimination program in Kayin state, eastern Myanmar, led to a substantial reduction in falciparum malaria. Despite the intense use of artemisinin-based combination therapies, both in treatment and MDA, this did not select for artemisinin resistance.

Use of High-Resolution Geospatial and Genomic Data to Characterize Recent Tuberculosis Transmission, Botswana.

Combining genomic and geospatial data can be useful for understanding Mycobacterium tuberculosis transmission in high-burden tuberculosis (TB) settings. We performed whole-genome sequencing on M. tuberculosis DNA extracted from sputum cultures from a population-based TB study conducted in Gaborone, Botswana, during 2012-2016. We determined spatial distribution of cases on the basis of shared genotypes among isolates. We considered clusters of isolates with ≤5 single-nucleotide polymorphisms identified by whole-genome sequencing to indicate recent transmission and clusters of ≥10 persons to be outbreaks. We obtained both molecular and geospatial data for 946/1,449 (65%) participants with culture-confirmed TB; 62 persons belonged to 5 outbreaks of 10-19 persons each. We detected geospatial clustering in just 2 of those 5 outbreaks, suggesting heterogeneous spatial patterns. Our findings indicate that targeted interventions applied in smaller geographic areas of high-burden TB identified using integrated genomic and geospatial data might help interrupt TB transmission during outbreaks.

International border malaria transmission in the Ethiopian district of Lare, Gambella region: implications for malaria spread into South Sudan.

BACKGROUND: Despite notable progress in the control and prevention of malaria in the Horn of Africa, the disease continues to cause significant morbidity and mortality in various regions of Ethiopia, and elsewhere in the region. The transmission of malaria is affected by genetic, sociocultural, and ecological factors. Lare is an Ethiopian district adjacent to the Ethio-South Sudan border, in Gambella region. The region currently has the highest prevalence of malaria in Ethiopia. This study assesses the burden and spatiotemporal patterns of disease transmission, including the effect of climatic factors on the occurrence of malaria, across an international border crossing. This understanding can assist in crafting informed programmatic and policy decisions for interventions. METHODS: This study was conducted in Lare district, Southwest Ethiopia, a temperate zone. A retrospective descriptive analysis was conducted using clinical service data collected between 2011 and 2021 from the 9 health facilities of the district. Both clinically diagnosed patients and those identified using microscopy and rapid diagnostic testing (RDT) were included in the study. Additionally, climate data was incorporated into analyses. Examples of analyses include malaria burden, positivity rate, incidence, species frequency, and an ANOVA to assess inter-annual case number and meteorological factor variation. RESULTS: Between 2011 and 2021, a total of 96,616 suspected malaria cases were tested by microscopy or RDT, and 39,428 (40.8%) of these cases were reported as positive. There were 1276 patients admitted with 22 deaths recorded. There were further more significant fluctuations in positivity rates across years, the highest being 74.5% in 2021. Incidence varied from 18.0% in 2011 to 151.6% in 2016. The malaria parasite species most detected was Plasmodium falciparum, followed by a smaller proportion of Plasmodium vivax. The greatest proportions of P. falciparum cases were observed in 2018 and 2019, at 97.4% and 97.0% prevalence, respectively. There was significant seasonal variation in case number, the highest observed in July through September of each year. Climatic conditions of annual rainfall, temperature and humidity favored the increment of malaria cases from June until October. CONCLUSION: The study shows that the burden, i.e. morbidity and mortality (with fluctuating patterns) of malaria are still significant public health problems and can pose serious consequences in the district. This has implication for cross-border malaria transmission risk due to considerable border crossings. The predominant cause of the disease is P. falciparum, which causes severe complications in patients. The district has to prepare to deal with such complications for better patient care and outcomes.

Community acceptability, participation, and adherence to mass drug administration with primaquine for Plasmodium vivax elimination in Southern Thailand: a mixed methods approach.

BACKGROUND: Mass drug administration (MDA) with primaquine (PQ) is being considered for accelerating Plasmodium vivax elimination in remaining active foci. This study aimed to determine the acceptability of MDA with PQ in malaria endemic villages in a malarious setting in the South of Thailand undergoing MDA with PQ. METHODS: A cross-sectional mixed-methods approach was conducted in seven malaria endemic villages where MDA with PQ was implemented. The data were collected from community villagers and health workers using structured questionnaires, in-depth interviews, and focus group discussions. Descriptive statistics and logistic regression models were used for quantitative data analysis. Thematic analysis was applied for qualitative data. RESULTS: Among a total of 469 participants from the MDA villages, 293 participants were eligible for MDA with PQ and 79.86% (234) completed 14-days of PQ. The logistic regressions indicated that males (adjusted odds ratio: 2.52 [95% confidence interval: 1.33-4.81]) and those who are farmers (2.57 [1.12-5.90]) were most likely to participate in the MDA. Among 293 participants in the post-MDA study, 74.06% had originally agreed to participate in the MDA with PQ while 25.94% had originally reported not wanting to participate in the MDA. Of those who originally reported being willing to participate in the MDA, 71.23% followed through with participation in the first or second round. Conversely, 93.24% of those who originally reported not being willing to participate in the MDA did in fact participate in the MDA. Factors contributing to higher odds of agreeing to participate and following through with participation included being male (1.98 [1.06-3.69]) and correctly responding that malaria is preventable (2.32 [1.01-5.35]) with some differences by village. Five key themes emerged from the qualitative analyses: concern about side effects from taking PQ; disbelief that malaria could be eliminated in this setting; low overall concern about malaria infections; misunderstandings about malaria; and a general need to tailor public health efforts for this unique context. CONCLUSION: While the reported likelihood of participating in MDA was high in this setting, actual follow-through was relatively moderate, partially because of eligibility (roughly 71% of those in the follow-up survey who originally agreed to participate actually followed through with participation). One of the largest concerns among study participants was PQ-related side effects-and these concerns likely heavily influenced participant adherence to the MDA. The results of this study can be used to tailor future MDAs, or other public health interventions, in this and potentially other similar settings.

Development of Inapparent Dengue Associated With Increased Antibody Levels to Aedes aegypti Salivary Proteins: A Longitudinal Dengue Cohort in Cambodia.

BACKGROUND: We established the first prospective cohort to understand how infection with dengue virus is influenced by vector-specific determinants such as humoral immunity to Aedes aegypti salivary proteins. METHODS: Children aged 2-9 years were enrolled in the PAGODAS (Pediatric Assessment Group of Dengue and Aedes Saliva) cohort with informed consent by their guardians. Children were followed semi-annually for antibodies to dengue and to proteins in Ae. aegypti salivary gland homogenate using enzyme-linked immunosorbent assays and dengue-specific neutralization titers. Children presented with fever at any time for dengue testing. RESULTS: From 13 July to 30 August 2018, we enrolled 771 children. At baseline, 22% (173/770) had evidence of neutralizing antibodies to 1 or more dengue serotypes. By April 2020, 51 children had symptomatic dengue while 148 dengue-naive children had inapparent dengue defined by neutralization assays. In a multivariate model, individuals with higher antibodies to Ae. aegypti salivary proteins were 1.5 times more likely to have dengue infection (hazard ratio [HR], 1.47 [95% confidence interval {CI}, 1.05-2.06]; P = .02), particularly individuals with inapparent dengue (HR, 1.64 [95% CI, 1.12-2.41]; P = .01). CONCLUSIONS: High levels of seropositivity to Ae. aegypti salivary proteins are associated with future development of dengue infection, primarily inapparent, in dengue-naive Cambodian children. CLINICAL TRIALS REGISTRATION: NCT03534245.

Mobile phone handover data for measuring and analysing human population mobility in Western Ethiopia: implication for malaria disease epidemiology and elimination efforts.

BACKGROUND: Human mobility behaviour modelling plays an essential role in the understanding and control of the spread of contagious diseases by limiting the contact among individuals, predicting the spatio-temporal evolution of an epidemic and inferring migration patterns. It informs programmatic and policy decisions for effective and efficient intervention. The objective of this research is to assess the human mobility pattern and analyse its implication for malaria disease epidemiology. METHODS: In this study, human mobility patterns in Benishangul-Gumuz and Gambella regions in Western Ethiopia were explored based on a cellular network mobility parameter (e.g., handover rate) via real world data. Anonymized data were retrieved for mobile active users with mobility related information. The data came from anonymous traffic records collected from all the study areas. For each cell, the necessary mobility parameter data per hour, week and month were collected. A scale factor was computed to change the mobility parameter value to the human mobility pattern. Finally, the relative human mobility probability for each scenario was estimated. MapInfo and Matlab softwares were used for visualization and analysis purposes. Hourly travel patterns in the study settings were compared with hourly malaria mosquito vector feeding behaviour. RESULTS: Heterogeneous human movement patterns were observed in the two regions with some areas showing typically high human mobility. Furthermore, the number of people entering into the two study regions was high during the highest malaria transmission season. Two peaks of hourly human movement, 8:00 to 9:00 and 16:00 to 18:00, emerged in Benishangul-Gumuz region while 8:00 to 10:00 and 16:00 to 18:00 were the peak hourly human mobility time periods in Gambella region. The high human movement in the night especially before midnight in the two regions may increase the risk of getting mosquito bite particularly by early biters depending on malaria linked human behaviour of the population. CONCLUSIONS: High human mobility was observed both within and outside the two regions. The population influx and efflux in these two regions is considerably high. This may specifically challenge the transition from malaria control to elimination. The daily mobility pattern is worth considering in the context of malaria transmission. In line with this malaria related behavioural patterns of humans need to be properly addressed.

Surveillance to achieve malaria elimination in eastern Myanmar: a 7-year observational study.

BACKGROUND: The collection and utilization of surveillance data is essential in monitoring progress towards achieving malaria elimination, in the timely response to increases in malaria case numbers and in the assessment of programme functioning. This paper describes the surveillance activities used by the malaria elimination task force (METF) programme which operates in eastern Myanmar, and provides an analysis of data collected from weekly surveillance, case investigations, and monitoring and evaluation of programme performance. METHODS: This retrospective analysis was conducted using data collected from a network of 1250 malaria posts operational between 2014 and 2021. To investigate changes in data completeness, malaria post performance, malaria case numbers, and the demographic details of malaria cases, summary statistics were used to compare data collected over space and time. RESULTS: In the first 3 years of the METF programme, improvements in data transmission routes resulted in a 18.9% reduction in late reporting, allowing for near real-time analysis of data collected at the malaria posts. In 2020, travel restrictions were in place across Karen State in response to COVID-19, and from February 2021 the military coup in Myanmar resulted in widescale population displacement. However, over that period there has been no decline in malaria post attendance, and the majority of consultations continue to occur within 48 h of fever onset. Case investigations found that 43.8% of cases travelled away from their resident village in the 3 weeks prior to diagnosis and 36.3% reported never using a bed net whilst sleeping in their resident village, which increased to 72.2% when sleeping away from their resident village. Malaria post assessments performed in 82.3% of the METF malaria posts found malaria posts generally performed to a high standard. CONCLUSIONS: Surveillance data collected by the METF programme demonstrate that despite significant changes in the context in which the programme operates, malaria posts have remained accessible and continue to provide early diagnosis and treatment contributing to an 89.3% decrease in Plasmodium falciparum incidence between 2014 and 2021.

Therapeutic efficacy of chloroquine for uncomplicated Plasmodium vivax malaria in southeastern and western border areas of Myanmar.

BACKGROUND: In the Greater Mekong Subregion of Southeast Asia, Plasmodium vivax malaria is endemic and causes significant morbidity. In this study, the efficacy of chloroquine for treating uncomplicated P. vivax malaria at the eastern and western borders of Myanmar was investigated. METHODS: A total of 197 participants with microscopically confirmed P. vivax infection were enrolled from three townships of the southeastern (Thanbyuzayat and Kawthoung) and western (Kyauktaw) borders of Myanmar. Patients were treated with chloroquine according to the national malaria treatment guidelines and followed for 28 days. RESULTS: Among the 197 enrollments, 172 completed the 28-day follow-up. Twelve recurrent P. vivax infections, all occurring in the third and fourth week, were detected, resulting in an overall cumulative rate of recurrence of 4.7% [95% confidence interval (CI) 1.5-7.8]. The incidence rate of recurrence varied among the three sites. In Thanbyuzayat township, no patients had recurrent parasitemia between days 7 and 28. In contrast, Kyauktaw township had a day 28 cumulative incidence rate of recurrence of 7.2% (95% CI 0.6-13.9%) compared to 6.9% (95% CI 0.6-13.2) in Kawthoung township. CONCLUSION: While this study confirmed the relatively high clinical efficacy of chloroquine for treating P. vivax in Myanmar with modest rates of recurrent infections within 28 days of the treatment, it also revealed considerable geographical heterogeneity of chloroquine efficacy, which warrants continuous surveillance efforts.

Spatiotemporal dynamics of malaria in Banmauk Township, Sagaing region of Northern Myanmar: characteristics, trends, and risk factors.

BACKGROUND: While national malaria incidence has been declining in Myanmar, some subregions within the nation continue to have high burdens of malaria morbidity and mortality. This study assessed the malaria situation in one of these regions, Banmauk Township, located near the Myanmar-India border. Our goal was to provide a detailed description of the malaria epidemiology in this township and to provide some evidence-based recommendations to formulate a strategy for reaching the national malaria elimination plan. Banmauk consistently has one of the highest malaria burdens in Myanmar. METHODS: With the implementation of strengthened malaria control and surveillance activities after the endorsement of a national malaria elimination plan in 2015, detailed incidence data were obtained for 2016-2018 for Banmauk Township. The data include patient demographics, parasite species, disease severity, and disease outcome. Data were analyzed to identify characteristics, trends, distribution, and risk factors. RESULTS: During 2016-2018, 2,402 malaria cases were reported, with Plasmodium falciparum accounting for 83.4% of infections. Both P. falciparum and P. vivax were transmitted more frequently during the rainy season (May-October). Despite intensified control, the annual parasite incidence rate (API) in 2017 (11.0) almost doubled that in 2016 (6.5). In total, 2.5% (59/2042) of the cases, of which 54 P. falciparum and 5 P. vivax, were complicated cases, resulting in 5 deaths. Malaria morbidity was high in children < 15 years and accounted for 33.4% of all cases and about 47% of the complicated cases. Older age groups and males living with poor transportation conditions were more likely to test positive especially in rainy and cold seasons. Despite the clear seasonality of malaria, severe cases were found among young children even more common in the dry season, when malaria incidence was low. CONCLUSIONS: Despite the declining trend, the malaria burden remained high in Banmauk Township. Our study also documented severe cases and deaths from both falciparum and vivax malaria. P. falciparum remained the predominant parasite species, demanding increased efforts to achieve the goal of elimination of P. falciparum by 2025. As P. falciparum cases decreased, the proportion of cases attributable to P. vivax increased. In order to eliminate malaria, it will likely be important to increasingly target this species as well.

Burden of malaria, impact of interventions and climate variability in Western Ethiopia: an area with large irrigation based farming.

BACKGROUND: Land use change has increasingly been expanding throughout the world in the past decades. It can have profound effects on the spatial and temporal distribution of vector borne diseases like malaria through ecological and habitat change. Understanding malaria disease occurrence and the impact of prevention interventions under this intense environmental modification is important for effective and efficient malaria control strategy. METHODS: A descriptive ecological study was conducted by reviewing health service records at Abobo district health office. The records were reviewed to extract data on malaria morbidity, mortality, and prevention and control methods. Moreover, Meteorological data were obtained from Gambella region Meteorology Service Center and National Meteorology Authority head office. Univariate, bivariate and multivariate analysis techniques were used to analyze the data. RESULTS: For the twelve-year time period, the mean annual total malaria case count in the district was 7369.58. The peak monthly malaria incidence was about 57 cases per 1000 people. Only in 2009 and 2015 that zero death due to malaria was recorded over the past 12 years. Fluctuating pattern of impatient malaria cases occurrence was seen over the past twelve years with an average number of 225.5 inpatient cases. The data showed that there is a high burden of malaria in the district. Plasmodium falciparum (Pf) was a predominant parasite species in the district with the maximum percentage of about 90. There was no statistically significant association between season and total malaria case number (F3,8: 1.982, P:0.195). However, the inter-annual total case count difference was statistically significant (F11,132: 36.305, p < 0001). Total malaria case count had shown two months lagged carry on effect. Moreover, 3 months lagged humidity had significant positive effect on total malaria cases. Malaria prevention interventions and meteorological factors showed statistically significant association with total malaria cases. CONCLUSION: Malaria was and will remain to be a major public health problem in the area. The social and economic impact of the disease on the local community is clearly pronounced as it is the leading cause of health facility visit and admission including the mortality associated with it. Scale up of effective interventions is quite important. Continuous monitoring of the performance of the vector control tools needs to be done.

Predictors of Test Positivity, Mortality, and Seropositivity during the Early Coronavirus Disease Epidemic, Orange County, California, USA.

We conducted a detailed analysis of coronavirus disease in a large population center in southern California, USA (Orange County, population 3.2 million), to determine heterogeneity in risks for infection, test positivity, and death. We used a combination of datasets, including a population-representative seroprevalence survey, to assess the actual burden of disease and testing intensity, test positivity, and mortality. In the first month of the local epidemic (March 2020), case incidence clustered in high-income areas. This pattern quickly shifted, and cases next clustered in much higher rates in the north-central area of the county, which has a lower socioeconomic status. Beginning in April 2020, a concentration of reported cases, test positivity, testing intensity, and seropositivity in a north-central area persisted. At the individual level, several factors (e.g., age, race or ethnicity, and ZIP codes with low educational attainment) strongly affected risk for seropositivity and death.

The assembly effect: the connectedness between populations is a double-edged sword for public health interventions.

BACKGROUND: Many public health interventions lead to disruption or decrease of transmission, providing a beneficial effect for people in the population regardless of whether or not they individually participate in the intervention. This protective benefit has been referred to as a herd or community effect and is dependent on sufficient population participation. In practice, public health interventions are implemented at different spatial scales (i.e., at the village, district, or provincial level). Populations, however defined (i.e., neighbourhoods, villages, districts) are frequently connected to other populations through human movement or travel, and this connectedness can influence potential herd effects. METHODS: The impact of a public health intervention (mass drug administration for malaria) was modelled, for different levels of connectedness between populations that have similar disease epidemiology (e.g., two nearby villages which have similar baseline malaria incidences and similar malaria intervention measures), or between populations of varying disease epidemiology (e.g., two nearby villages which have different baseline malaria incidences and/or malaria intervention measures). RESULTS: The overall impact of the interventions deployed could be influenced either positively (adding value to the intervention) or negatively (reducing the impact of the intervention) by how much the intervention units are connected with each other (e.g., how frequent people go to the other village or town) and how different the disease intensity between them are. This phenomenon is termed the "assembly effect", and it is a meta-population version of the more commonly understood "herd effect". CONCLUSIONS: The connectedness of intervention units or populations is an important factor to be considered to achieve success in public health interventions that could provide herd effects. Appreciating the assembly effect can improve the cost-effective strategies for global disease elimination projects.

Ownership and utilization of bed nets and reasons for use or non-use of bed nets among community members at risk of malaria along the Thai-Myanmar border.

BACKGROUND: With the goal for malaria elimination in Thailand set for 2024, increased coverage and utilization of bed net, especially insecticide-treated net (ITN) or long-lasting insecticidal net (LLIN) is a key strategy. This study aims to provide the necessary information about bed net ownership and utilization among the population at risk of malaria living along the Thai-Myanmar border in Tak province. METHODS: A cross-sectional study was conducted using a mixed-method approach in 331 households from 5 hamlets in the villages of the Thai-Myanmar border. The research tools included a questionnaire, bed net inspection, and semi-structured interviews. Logistic regression was used to explore the sociodemographic factors associated with bed net utilization. The qualitative analysis employed a thematic analysis approach. RESULTS: This survey found that 98.5% of households had at least one bed net per household, and 74.3% had at least one ITN/LLIN. However, only 30.8% of households reached the standard policy set by the Minister of Public Health of one ITN/LLINs per two persons. Most residents used bed net (92.1% used in the previous night and 80.9% used every day). For those using bed nets, however, 61.9% used ITNs or LLINs the night before and 53.1% used them every day. Nonetheless, the usage rates of bed nets (any type) in the previous night among children and pregnant women were high, reaching 95.3% and 90.0%, respectively. Seven explanatory variables showed statistically significant associations with bed net use every day, including: "not staying overnight in the forest or the field", "sleeping pattern based on gender", "sufficient numbers of bed nets to cover all sleeping spaces", "preference for free bed nets", "age", "gender", and "SES score" showed statistically significant association with bed net use every day. The major reasons for the regular use of bed nets in both household and the forest were to prevent mosquito biting. The reasons for not using bednets in the household were discomfort feelings from heat, perception of unnecessity due to low mosquito density, whereas the reason for not using bed nets in the forest was inconvenience. CONCLUSION: Despite that overall coverage and usage of bed nets was high, only one third reached the standard level specified by the policy. Overnight in the forest, the dissatisfaction with the quality of free bed nets, insufficient number of bed nets, sleeping alone, male gender, age more than 10 years, low socioeconomic status, discomfort from heat, perception of no benefits of bed nets due to low mosquito density, and inconvenience were factors influencing bed net use. Maintaining high coverage and utility rate of bed nets should be a priority for the malaria high-risk population.

Clustering of malaria in households in the Greater Mekong Subregion: operational implications for reactive case detection.

BACKGROUND: Malaria reactive case detection is the testing and, if positive, treatment of close contacts of index cases. It is included in national malaria control programmes of countries in the Greater Mekong Subregion to accelerate malaria elimination. Yet the value of reactive case detection in the control and elimination of malaria remains controversial because of the low yield, limited evidence for impact, and high demands on resources. METHODS: Data from the epidemiological assessments of large mass drug administration (MDA) studies in Myanmar, Vietnam, Cambodia and Laos were analysed to explore malaria infection clustering in households. The proportion of malaria positive cases among contacts screened in a hypothetical reactive case detection programme was then determined. The parasite density thresholds for rapid diagnostic test (RDT) detection was assumed to be > 50/µL (50,000/mL), for dried-blood-spot (DBS) based PCR > 5/µL (5000/mL), and for ultrasensitive PCR (uPCR) with a validated limit of detection at 0.0022/µL (22/mL). RESULTS: At baseline, before MDA, 1223 Plasmodium infections were detected by uPCR in 693 households. There was clustering of Plasmodium infections. In 637 households with asymptomatic infections 44% (278/637) had more than one member with Plasmodium infections. In the 132 households with symptomatic infections, 65% (86/132) had more than one member with Plasmodium infections. At baseline 4% of households had more than one Plasmodium falciparum infection, but three months after MDA no household had more than one P. falciparum infected member. Reactive case detection using DBS PCR would have detected ten additional cases in six households, and an RDT screen would have detected five additional cases in three households among the 169 households with at least one RDT positive case. This translates to 19 and 9 additional cases identified per 1000 people screened, respectively. Overall, assuming all febrile RDT positive patients would seek treatment and provoke reactive case detection using RDTs, then 1047 of 1052 (99.5%) Plasmodium infections in these communities would have remained undetected. CONCLUSION: Reactive case detection in the Greater Mekong subregion is predicted to have a negligible impact on the malaria burden, but it has substantial costs in terms of human and financial resources.

Distance matters: barriers to antenatal care and safe childbirth in a migrant population on the Thailand-Myanmar border from 2007 to 2015, a pregnancy cohort study.

BACKGROUND: Antenatal care and skilled childbirth services are important interventions to improve maternal health and lower the risk of poor pregnancy outcomes and mortality. A growing body of literature has shown that geographic distance to clinics can be a disincentive towards seeking care during pregnancy. On the Thailand-Myanmar border antenatal clinics serving migrant populations have found high rates of loss to follow-up of 17.4%, but decades of civil conflict have made the underlying factors difficult to investigate. Here we perform a comprehensive study examining the geographic, demographic, and health-related factors contributing to loss to follow-up. METHODS: Using patient records we conducted a spatial and epidemiological analysis looking for predictors of loss to follow-up and pregnancy outcomes between 2007 and 2015. We used multivariable negative binomial regressions to assess for associations between distance travelled to the clinic and birth outcomes (loss to follow-up, pregnancy complications, and time of first presentation for antenatal care.) RESULTS: We found distance travelled to clinic strongly predicts loss to follow-up, miscarriage, malaria infections in pregnancy, and presentation for antenatal care after the first trimester. People lost to follow-up travelled 50% farther than people who had a normal singleton childbirth (a ratio of distances (DR) 1.5; 95% confidence interval (CI): 1.4 - 1.5). People with pregnancies complicated by miscarriage travelled 20% farther than those who did not have miscarriages (DR: 1.2; CI 1.1-1.3), and those with Plasmodium falciparum malaria in pregnancy travelled 60% farther than those without P. falciparum (DR: 1.6; CI: 1.6 - 1.8). People who delayed antenatal care until the third trimester travelled 50% farther compared to people who attended in the first trimester (DR: 1.5; CI: 1.4 - 1.5). CONCLUSIONS: This analysis provides the first evidence of the complex impact of geography on access to antenatal services and pregnancy outcomes in the rural, remote, and politically complex Thailand-Myanmar border region. These findings can be used to help guide evidence-based interventions to increase uptake of maternal healthcare both in the Thailand-Myanmar region and in other rural, remote, and politically complicated environments.

Optimising the cost of roadkill surveys based on an analysis of carcass persistence.

Reliable estimates of wildlife mortality due to wildlife-vehicle collisions are key to understanding its impact on wildlife populations and developing strategies to prevent or reduce collisions. Standardised approaches for monitoring roadkill are needed to derive robust and unbiased estimates of mortality that are comparable across different study systems and ecological contexts. When designing surveys, there is a trade-off between survey frequency (and hence logistical effort and financial cost) and carcass detection. In this regard, carcass persistence (the period a carcass remains detectable before being removed by decomposition or scavengers) is important; the longer a carcass persists, the greater the likelihood it will be detected with lower survey effort by conducting more infrequent surveys. Using multi-taxon carcass data collected over a month of repeated driven surveys, combined with five covariates (species functional group, body weight, carcass position on road, carcass condition [either flattened or not after impact], and rainfall prior to each survey), we explored the drivers of carcass persistence with the overall aim of providing information to optimise the design of carcass surveys along linear infrastructure. Our methodological approach included a survival analysis to determine carcass persistence, linear regressions to test the effect of covariates, a subsampling analysis (using field data and a simulation exercise) to assess how the proportion of carcasses detected changes according to survey frequency, and an analysis to compare the costs of surveys based on study duration, transect length and survey frequency. Mean overall carcass persistence was 2.7 days and was significantly correlated with position on road and within-functional group body weight. There was no evidence for a significant effect of rainfall, while the effect of carcass condition was weakly non-significant. The proportion of carcasses detected decreased sharply when survey intervals were longer than three days. However, we showed that survey costs can be reduced by up to 80% by conducting non-daily surveys. Expanding on the call for a standardised methodology for roadkill surveys, we propose that carcass persistence be explicitly considered during survey design. By carefully considering the objectives of the survey and characteristics of the focal taxa, researchers can substantially reduce logistical costs. In addition, we developed an R Shiny web app that can be used by practitioners to compare survey costs across a variety of survey characteristics. This web app will allow practitioners to easily assess the trade-off between carcass detection and logistical effort.

Contribution of Asymptomatic Plasmodium Infections to the Transmission of Malaria in Kayin State, Myanmar.

BACKGROUND: The objective of mass antimalarial drug administration (MDA) is to eliminate malaria rapidly by eliminating the asymptomatic malaria parasite reservoirs and interrupting transmission. In the Greater Mekong Subregion, where artemisinin-resistant Plasmodium falciparum is now widespread, MDA has been proposed as an elimination accelerator, but the contribution of asymptomatic infections to malaria transmission has been questioned. The impact of MDA on entomological indices has not been characterized previously. METHODS: MDA was conducted in 4 villages in Kayin State (Myanmar). Malaria mosquito vectors were captured 3 months before, during, and 3 months after MDA, and their Plasmodium infections were detected by polymerase chain reaction (PCR) analysis. The relationship between the entomological inoculation rate, the malaria prevalence in humans determined by ultrasensitive PCR, and MDA was characterized by generalized estimating equation regression. RESULTS: Asymptomatic P. falciparum and Plasmodium vivax infections were cleared by MDA. The P. vivax entomological inoculation rate was reduced by 12.5-fold (95% confidence interval [CI], 1.6-100-fold), but the reservoir of asymptomatic P. vivax infections was reconstituted within 3 months, presumably because of relapses. This was coincident with a 5.3-fold (95% CI, 4.8-6.0-fold) increase in the vector infection rate. CONCLUSION: Asymptomatic infections are a major source of malaria transmission in Southeast Asia.

Extended and Continuous Decline in Effective Population Size Results in Low Genomic Diversity in the World's Rarest Hyena Species, the Brown Hyena.

Hyenas (family Hyaenidae), as the sister group to cats (family Felidae), represent a deeply diverging branch within the cat-like carnivores (Feliformia). With an estimated population size of <10,000 individuals worldwide, the brown hyena (Parahyaena brunnea) represents the rarest of the four extant hyena species and has been listed as Near Threatened by the IUCN. Here, we report a high-coverage genome from a captive bred brown hyena and both mitochondrial and low-coverage nuclear genomes of 14 wild-caught brown hyena individuals from across southern Africa. We find that brown hyena harbor extremely low genetic diversity on both the mitochondrial and nuclear level, most likely resulting from a continuous and ongoing decline in effective population size that started ∼1 Ma and dramatically accelerated towards the end of the Pleistocene. Despite the strikingly low genetic diversity, we find no evidence of inbreeding within the captive bred individual and reveal phylogeographic structure, suggesting the existence of several potential subpopulations within the species.

The impact of targeted malaria elimination with mass drug administrations on falciparum malaria in Southeast Asia: A cluster randomised trial.

BACKGROUND: The emergence and spread of multidrug-resistant Plasmodium falciparum in the Greater Mekong Subregion (GMS) threatens global malaria elimination efforts. Mass drug administration (MDA), the presumptive antimalarial treatment of an entire population to clear the subclinical parasite reservoir, is a strategy to accelerate malaria elimination. We report a cluster randomised trial to assess the effectiveness of dihydroartemisinin-piperaquine (DP) MDA in reducing falciparum malaria incidence and prevalence in 16 remote village populations in Myanmar, Vietnam, Cambodia, and the Lao People's Democratic Republic, where artemisinin resistance is prevalent. METHODS AND FINDINGS: After establishing vector control and community-based case management and following intensive community engagement, we used restricted randomisation within village pairs to select 8 villages to receive early DP MDA and 8 villages as controls for 12 months, after which the control villages received deferred DP MDA. The MDA comprised 3 monthly rounds of 3 daily doses of DP and, except in Cambodia, a single low dose of primaquine. We conducted exhaustive cross-sectional surveys of the entire population of each village at quarterly intervals using ultrasensitive quantitative PCR to detect Plasmodium infections. The study was conducted between May 2013 and July 2017. The investigators randomised 16 villages that had a total of 8,445 residents at the start of the study. Of these 8,445 residents, 4,135 (49%) residents living in 8 villages, plus an additional 288 newcomers to the villages, were randomised to receive early MDA; 3,790 out of the 4,423 (86%) participated in at least 1 MDA round, and 2,520 out of the 4,423 (57%) participated in all 3 rounds. The primary outcome, P. falciparum prevalence by month 3 (M3), fell by 92% (from 5.1% [171/3,340] to 0.4% [12/2,828]) in early MDA villages and by 29% (from 7.2% [246/3,405] to 5.1% [155/3,057]) in control villages. Over the following 9 months, the P. falciparum prevalence increased to 3.3% (96/2,881) in early MDA villages and to 6.1% (128/2,101) in control villages (adjusted incidence rate ratio 0.41 [95% CI 0.20 to 0.84]; p = 0.015). Individual protection was proportional to the number of completed MDA rounds. Of 221 participants with subclinical P. falciparum infections who participated in MDA and could be followed up, 207 (94%) cleared their infections, including 9 of 10 with artemisinin- and piperaquine-resistant infections. The DP MDAs were well tolerated; 6 severe adverse events were detected during the follow-up period, but none was attributable to the intervention. CONCLUSIONS: Added to community-based basic malaria control measures, 3 monthly rounds of DP MDA reduced the incidence and prevalence of falciparum malaria over a 1-year period in areas affected by artemisinin resistance. P. falciparum infections returned during the follow-up period as the remaining infections spread and malaria was reintroduced from surrounding areas. Limitations of this study include a relatively small sample of villages, heterogeneity between villages, and mobility of villagers that may have limited the impact of the intervention. These results suggest that, if used as part of a comprehensive, well-organised, and well-resourced elimination programme, DP MDA can be a useful additional tool to accelerate malaria elimination. TRIAL REGISTRATION: ClinicalTrials.gov NCT01872702.