Handling missing disease information due to death in diseases that need two visits to diagnose.

In studies that assess disease status periodically, time of disease onset is interval censored between visits. Participants who die between two visits may have unknown disease status after their last visit. In this work, we consider an additional scenario where diagnosis requires two consecutive positive tests, such that disease status can also be unknown at the last visit preceding death. We show that this impacts the choice of censoring time for those who die without an observed disease diagnosis. We investigate two classes of models that quantify the effect of risk factors on disease outcome: a Cox proportional hazards model with death as a competing risk and an illness death model that treats disease as a possible intermediate state. We also consider four censoring strategies: participants without observed disease are censored at death (Cox model only), the last visit, the last visit with a negative test, or the second last visit. We evaluate the performance of model and censoring strategy combinations on simulated data with a binary risk factor and illustrate with a real data application. We find that the illness death model with censoring at the second last visit shows the best performance in all simulation settings. Other combinations show bias that varies in magnitude and direction depending on the differential mortality between diseased and disease-free subjects, the gap between visits, and the choice of the censoring time.

Advancing randomized controlled trial methodologies: The place of innovative trial design in eating disorders research.

Randomized controlled trials can be used to generate evidence on the efficacy and safety of new treatments in eating disorders research. Many of the trials previously conducted in this area have been deemed to be of low quality, in part due to a number of practical constraints. This article provides an overview of established and more innovative clinical trial designs, accompanied by pertinent examples, to highlight how design choices can enhance flexibility and improve efficiency of both resource allocation and participant involvement. Trial designs include individually randomized, cluster randomized, and designs with randomizations at multiple time points and/or addressing several research questions (master protocol studies). Design features include the use of adaptations and considerations for pragmatic or registry-based trials. The appropriate choice of trial design, together with rigorous trial conduct, reporting and analysis, can establish high-quality evidence to advance knowledge in the field. It is anticipated that this article will provide a broad and contemporary introduction to trial designs and will help researchers make informed trial design choices for improved testing of new interventions in eating disorders. PUBLIC SIGNIFICANCE: There is a paucity of high quality randomized controlled trials that have been conducted in eating disorders, highlighting the need to identify where efficiency gains in trial design may be possible to advance the eating disorder research field. We provide an overview of some key trial designs and features which may offer solutions to practical constraints and increase trial efficiency.

Oleanane-type saponins from Lysimachia laxa Baudo and their anti-bacterial activities.

Seven new oleanane-type triterpene saponins, lysimaponins A-G, were isolated from aerial parts of Lysimachia laxa Baudo. Their chemical structures have been elucidated by analysis of spectroscopic and chemical methods. All compounds were evaluated for their anti-bacterial effects against Microcystis aeruginosa, Vibrio parahaemolyticus, V. harveyi, V. vulinificus, V. cholerae, and V. alginolyticus. All compounds showed potent anti-bacterial activities against the cyanobacteria M. aeruginosa with IC50 values ranging from 14.4 ± 1.2 to 35.3 ± 2.2 µg/mL. Compounds 1, 2, 4-7 inhibited V. parahaemolyticus with MIC values ranging from 64 to 256 µg/mL. The results suggested that saponins from L. laxa could be potential anti-cyanobacteria agents.

Effect of Low-Dose Aspirin Versus Placebo on Incidence of Anemia in the Elderly : A Secondary Analysis of the Aspirin in Reducing Events in the Elderly Trial.

BACKGROUND: Daily low-dose aspirin increases major bleeding; however, few studies have investigated its effect on iron deficiency and anemia. OBJECTIVE: To investigate the effect of low-dose aspirin on incident anemia, hemoglobin, and serum ferritin concentrations. DESIGN: Post hoc analysis of the ASPREE (ASPirin in Reducing Events in the Elderly) randomized controlled trial. (ClinicalTrials.gov: NCT01038583). SETTING: Primary/community care in Australia and the United States. PARTICIPANTS: Community-dwelling persons aged 70 years or older (≥65 years for Black persons and Hispanic persons). INTERVENTION: 100 mg of aspirin daily or placebo. MEASUREMENTS: Hemoglobin concentration was measured annually in all participants. Ferritin was measured at baseline and 3 years after random assignment in a large subset. RESULTS: 19 114 persons were randomly assigned. Anemia incidence in the aspirin and placebo groups was 51.2 events and 42.9 events per 1000 person-years, respectively (hazard ratio, 1.20 [95% CI, 1.12 to 1.29]). Hemoglobin concentrations declined by 3.6 g/L per 5 years in the placebo group and the aspirin group experienced a steeper decline by 0.6 g/L per 5 years (CI, 0.3 to 1.0 g/L). In 7139 participants with ferritin measures at baseline and year 3, the aspirin group had greater prevalence than placebo of ferritin levels less than 45 µg/L at year 3 (465 [13%] vs. 350 [9.8%]) and greater overall decline in ferritin by 11.5% (CI, 9.3% to 13.7%) compared with placebo. A sensitivity analysis quantifying the effect of aspirin in the absence of major bleeding produced similar results. LIMITATIONS: Hemoglobin was measured annually. No data were available on causes of anemia. CONCLUSION: Low-dose aspirin increased incident anemia and decline in ferritin in otherwise healthy older adults, independent of major bleeding. Periodic monitoring of hemoglobin should be considered in older persons on aspirin. PRIMARY FUNDING SOURCE: National Institutes of Health and Australian National Health and Medical Research Council.

Anthropogenic particle abundance and characteristics in seawater and intertidal sediments of the Tonkin Bay Coast (North Vietnam).

Microplastics (MPs, plastic items from 1 µm to 5 mm in size) are present in all environmental compartments. The evaluation of their concentration, fate, and spatial distribution is still a challenge for the scientific community. This concern is just debuting in developing countries, (i.e., Asia, South America, and Africa). This study deals with the MP contamination in the abiotic marine compartments of Northern Vietnam: seawater and intertidal sediments. Four sites located in the intertidal zone or near the coastline in Tonkin Bay, Vietnam were studied. A total of 16 samples (eight for each compartment) were collected in July 2020 (rainy season) and January 2021 (dry season). Anthropogenic particles (total observed fibers and fragments) were found at levels ranging from 3 to 303 particles/m3 in seawater and from 63 to 955 particles/kg dry weight in sediments. Most of these were fibers less than 300-µm long. Higher levels of seawater at the Nam Dinh site were found in the rainy season compared to the dry one. As the river flow was estimated six times higher during the rainy season than during the dry season, these results suggest the river discharge is a potential source of contamination for the coastal zone. The temporal variability was lower for the sediments than for the seawater, suggesting the long-term integration of the anthropogenic particles in this compartment. A small portion of sorted particles were analyzed by µFTIR (8.35%), and this sub-sample was only composed of fragments. Still, fragments were mostly composed of polypropylene (PP, 82%), polyethylene (PE, 9%), and polystyrene (PS, 9%). The fragment size was similar in the two studied compartments, but it was dependent on polymer types since PS fragments (140 ± 17 µm) were smaller than those made of PE (622 ± 123 µm) and PP (869 ± 905 µm). Future works should investigate the smallest fraction of MP (even nanoplastics) as well as find solutions in order to mitigate MP contamination in the marine environment.

Microplastic accumulation in bivalves collected from different coastal areas of Vietnam and an assessment of potential risks.

Microplastic (MP) pollution is an emerging problem in many areas around the world and in coastal areas of Vietnam, requiring more studies dedicated to the accumulation of this pollutant in the food chain as well as its potential risk to human health. This study investigated MP levels in tissues of five common bivalve species collected from aquaculture areas along the coast of Vietnam. MPs were found in all bivalve samples, with average values of 10.84 ± 2.61 items/individual or 2.40 ± 1.34 items/g wet weight. Impacts of feeding habits of bivalves showed influences on MP abundance in the samples. Fibers were the dominant shape of MPs recorded, most of which accumulated in the gills and digestive glands of all bivalve samples, with the majority falling within the size range of 300-2000 µm. MPs found in all studied species had relatively similar chemical compositions, mainly composed of polypropylene (PP) and polyethylene (PE). In this study, a diverse diet consisting of different bivalve species and detailed data on the consumption rate of these species were used to assess the human health risk of MPs dedicated to the coastal communities of Vietnam. The results suggested a significant part of MP uptake by human could be via bivalve consumption, in which removing viscera and proper depuration should be applied prior to eating, thereby reducing the risk.

Benchmark Dynamics of Dipolar Molecular Rotors in Fluorinated Metal-Organic Frameworks.

Fluorinated Metal-Organic Frameworks (MOFs), comprising a wheel-shaped ligand with geminal rotating fluorine atoms, produced benchmark mobility of correlated dipolar rotors at 2 K, with practically null activation energy (Ea =17 cal mol-1 ). 1 H T1 NMR revealed multiple relaxation phenomena due to the exchange among correlated dipole-rotor configurations. Synchrotron radiation X-ray diffraction at 4 K, Density Functional Theory, Molecular Dynamics and phonon calculations showed the fluid landscape and pointed out a cascade mechanism converting dipole configurations into each other. Gas accessibility, shown by hyperpolarized-Xe NMR, allowed for chemical stimuli intervention: CO2 triggered dipole reorientation, reducing their collective dynamics and stimulating a dipole configuration change in the crystal. Dynamic materials under limited thermal noise and high responsiveness enable the fabrication of molecular machines with low energy dissipation and controllable dynamics.

A case-control study of agricultural and behavioral factors associated with leptospirosis in Vietnam.

INTRODUCTION: Leptospirosis is a neglected disease in Vietnam. Until now, there has been limited knowledge about risk factors of this disease in Vietnam. The study was carried out to identify agricultural and behavioral factors associated with the transmission of leptospirosis in Vietnam. METHODS: This matched retrospective hospital-community-based case-control study was conducted from 1 October 2018 to 31 October 2019. We recruited cases from 11 selected government hospitals in three provinces of Vietnam, while controls were selected from the same communes of cases and matched by age (± 2 years) and sex. Microscopic agglutination test (MAT) and enzyme-linked immunosorbent assay (ELISA) were applied to determine confirmed cases, while only MAT was used to identify controls with a single high MAT titer < 1:100. RESULTS: 504 participants (252 cases and 252 controls) were identified. Cultivating (OR 2.83, CI 1.38-5.79), animal farming (OR 8.26, CI 2.24-30.52), pig owners (OR 10.48, CI 5.05-21.73), cat owners (OR 2.62, CI 1.49-4.61) and drinking unboiled water (OR 1.72, CI 1.14 -2.59, p = 0.010) were significantly associated with human leptospirosis in Vietnam. Hand washing after farming/ gardening (OR 0.57, CI 0.38-0.86, p = 0.007) and bathing after farming, gardening, contact with cattle and poultry (OR 0.33, CI 0.19-0.58, p = 0.000) were determined as protective factors for this disease. CONCLUSIONS: In short, the case-control study has revealed the risks in agricultural and animal practices and protective behavioral factors related to human leptospirosis in Vietnam. The findings suggested promotion of communication and health education programs targeting health behaviors in daily life and agricultural practices. Using personal protective equipment such as gowns, gloves, and boots during agricultural practices, especially cultivating and animal farming, is most recommended.

Distribution and ecological risk assessment of phthalic acid esters in surface sediments of three rivers in Northern Vietnam.

Pollution status and distribution characteristics of ten typical phthalic acid esters (PAEs) were investigated in 36 sediment samples collected from three rivers in Northern Vietnam from June to October 2020. The total concentrations of PAEs in sediment samples collected from the To Lich River (n = 9), the Nhue River (n = 12), and the Day River (n = 15) were in ranges of 11,000-125,000 ng/g-dwt (mean/median: 50,000/42,200 ng/g-dwt), 2140-89,900 ng/g-dwt (mean/median: 29,300/20,700 ng/g-dwt), and 1140-43,100 ng/g-dwt (mean/median: 13,800/10,400 ng/g-dwt), respectively. Among ten PAEs studied, di-(2-ethylhexyl) phthalate (DEHP) was found at the highest levels in all samples meanwhile dimethyl phthalate (DMP), diethyl phthalate (DEP), and dipropyl phthalate (DPP) were detected at low frequency and concentration. Significant correlations have existed between the median-chain (C4-C7) PAE pairs in sediment samples. Due to the high accumulation in the sediments, the median-chain PAEs had a higher ecological risk than the short-chain (C1-C3) PAEs. These contaminants may present a longstanding influence on organisms and ecosystems.

Fecal indicator bacteria diversity and decay in an estuarine mangrove ecosystem of the Xuan Thuy National Park, Vietnam.

Mangroves are complex and dynamic ecosystems that are highly dependent on diverse microbial activities. In this study, laboratory experiments and field studies for fecal indicator bacteria (FIB) decay rates are carried out for the first time in the Xuan Thuy Mangrove Forest Reserve of Vietnam. Results show that there are significant differences in bacterial diversity in the water of mangrove areas that have been deforested compared to those which have been planted. The highest mean total coliform (TC) and Escherichia coli (EC) values were found in the natural mangroves (3,807±2,922 and 964±1133 CFU 100 ml-1, respectively). The results indicated that the source of contamination and seasonal changes affect the abundance of fecal bacteria. These results were exceeding by far the safety guidelines for individual, non-commercial water supplies in most of the samples. In the planted mangrove sampling sites, the highest mean Fecal streptococci (FS) values of 1,520±1,652 CFU 100 ml-1 were found. Microbial die-off rates were calculated over 5 days, and observed to be systematically higher for TC than for EC.

Selection of a density separation solution to study microplastics in tropical riverine sediment.

Microplastics (MPs) are small (< 5 mm) plastic particles that are widely found in marine, freshwater, terrestrial and atmospheric environments. Due to their prevalence and persistence, MPs are considered an emerging contaminant of environmental concern. The separation and quantitation of MPs from freshwater sediments is a challenging and critical issue. It is necessary to identify the fate and sources of MPs in the environment, minimise their release and adverse effects. Compared to marine sediments, standardised methods for extracting and estimating the amount of MPs in freshwater sediments are relatively limited. The present study focuses on MP recovery efficiency of four commonly used salt solutions (NaCl, NaI, CaCl2 and ZnCl2) for isolating MPs during the density separation step from freshwater sediment. Known combinations of artificial MP particles (PS, PE, PVC, PET, PP and HDPE) were spiked into standard river sediment. Extraction using NaI, ZnCl2 and NaCl solutions resulted in higher recovery rates from 37 to 97% compared to the CaCl2 solution (28-83%) and varied between polymer types. Low-density MPs (PE, HDPE, PP and PS) were more effectively recovered (> 87%) than the denser polymers (PET and PVC: 37 to 88.8%) using NaCl, NaI and ZnCl2 solutions. However, the effective flotation of ZnCl2 and NaI solutions is relatively expensive and unsafe to the environment, especially in the context of developing countries. Therefore, considering the efficiency, cost and environmental criteria, NaCl solution was selected. The protocol was then tested by extracting MPs from nine riverine sediment samples from the Red River Delta. Sediments collected from urban rivers were highly polluted by MPs (26,000 MPs items·kg-1 DW) compared to sediments located downstream. Using a NaCl solution was found to be effective in this case study and might also be used in long-term and large-scale MP monitoring programmes in Vietnam.

Polyhalogenated Bicyclo[1.1.1]pentane-1,3-dicarboxylic Acids.

Herein we report the highly selective radical chlorination of 2,2-difluorobicyclo[1.1.1]pentane-1,3-dicarboxylic acid. Together with radical hydrodechlorination by TMS3SiH, four new bicyclo[1.1.1]pentane cages carrying two fluorine and one to three chlorine atoms in bridge positions have been obtained. The exact positions of all halogen atoms have been confirmed by X-ray diffraction. The acidity constants (pKa) for all new derivatives have been determined by capillary electrophoresis, and these experimental values show excellent agreement with pKas predicted by DFT methods. Extensive DFT calculations have been used to rationalize the selective formation of four out of nine possible F2Cl1-4 isomers of bridge-halogenated bicyclo[1.1.1]pentanes and to obtain relative strain energies for all possible isomers.

Diterpenoids and Diacetylenes from the Roots of Aralia cordata with Inhibitory Effects on Nitric Oxide Production.

Bioactivity-guided isolation of a MeOH extract of Aralia cordata led to the isolation of four new ent-pimarane diterpenoids (1-4) and a diacetylene (5) together with 21 known compounds (6-26). Their structures were established based on the interpretation of one- and two-dimensional NMR and HRESIMS data. The absolute configurations of the new isolates were determined by electronic circular dichroism data analysis, single crystal X-ray diffraction, and Mosher's esterification method. All compounds exhibited inhibitory effects on lipopolysaccharide-induced nitric oxide production in RAW 264.7 macrophages with IC50 values ranging from 1.1 to 69.4 µM.

Sesquiterpenoids from Chrysanthemum indicum with Inhibitory Effects on NO Production.

Three new guaianolide lactones (1-3) and four new 9-oxonerolidol glucosides (5-8) together with 20 known compounds were isolated from the MeOH extract of the flowers of Chrysanthemum indicum. Their structures were elucidated based on the interpretation of NMR, HRESIMS, and electronic circular dichroism (ECD) data along with acid hydrolysis. Of the isolates, sesquiterpenoids 1-4 and 15 and flavones 17 and 18 exhibited inhibitory effects on lipopolysaccharide (LPS)-induced nitric oxide production in RAW 264.7 cells with IC50 values in the range 0.2-27.0 µM.

Pentacyclic triterpenes with nitric oxide inhibitory activity from Potentilla chinensis.

Three new ursane-type triterpenes (1-3) and twenty-one known triterpenoids (4-24) were isolated from the methanolic extract of the whole plants of Potentilla chinensis. Their structures were elucidated by extensive spectroscopic analysis of 1D and 2D NMR (HSQC, HMBC, COSY and ROESY) and HRESIMS data. The bioassay screening revealed the inhibitory effects on nitric oxide production of compounds 2, 5, 7, 9, 10, and 13-24 in LPS-induced RAW264.7 macrophages.

Comprehensive determination of polychlorinated biphenyls and brominated flame retardants in surface sediment samples from Hanoi urban area, Vietnam: Contamination status, accumulation profiles, and potential ecological risks.

Comprehensive and updated information about polychlorinated biphenyls (PCBs) and brominated flame retardants (BFRs) in surface sediments from Hanoi, the capital city of Vietnam, is rather scarce. In this study, concentrations and profiles of 209 PCBs, 41 polybrominated diphenyl ethers (PBDEs), 2,2',4,4',5,5'-hexabromobiphenyl (BB-153), hexabromocyclododecane (HBCD), pentabromoethylbenzene (PBEB), 1,2-bis(2,4,6-tribromophenoxy)ethane (BTBPE), and decabromodiphenyl ethane (DBDPE) were determined in sediment samples collected from the Red River and some inner-city rivers of Hanoi. Concentrations (ng/g dry weight, median and range) of pollutants decreased in the order: DBDPE (28; not detected ND - 59) ≈ PCBs (27; 1.7-50) > PBDEs (23; 0.20-61) > HBCD (1.2; ND - 5.2) > BTBPE (0.46; ND - 3.6) > BB-153 (0.004; ND - 0.014) > PBEB (ND). Pollutant levels in the inner-city river sediments were about one to two orders of magnitude higher than those measured in the Red River main stream sediments. Tri-to hexa-CBs are major homologs but detailed profiles vary between individual samples, reflecting source and/or seasonal variations. CB-11 and CB-209 were found at higher proportions in sediments than in technical PCB mixtures, suggesting their novel sources from pigments. Deca-BDE and DBDPE are the most predominant BFRs with an increasing trend predicted for DBDPE. A preliminary ecological risk assessment was conducted for these pollutants in sediments. Total PCBs and deca-BDE in a few inner-city river sediments may exhibit adverse effects on benthic organisms, but no serious risk was estimated in general.

Supramolecular Gels Incorporating Cordyline terminalis Leaf Extract as a Polyphenol Release Scaffold for Biomedical Applications.

Cordyline terminalis leaf extract (aqCT) possesses abundant polyphenols and other bioactive compounds, which are encapsulated in gelatin-polyethylene glycol-tyramine (GPT)/alpha-cyclodextrin (α-CD) gels to form the additional functional materials for biomedical applications. In this study, the gel compositions are optimized, and the GPT/α-CD ratios equal to or less than one half for solidification are found. The gelation time varies from 40.7 min to 5.0 h depending on the increase in GPT/α-CD ratios and aqCT amount. The aqCT extract disturbs the hydrogen bonding and host-guest inclusion of GPT/α-CD gel networks, postponing the gelation. Scanning electron microscope observation shows that all gels with or without aqCT possess a microarchitecture and porosity. GPT/α-CD/aqCT gels could release polyphenols from 110 to 350 nmol/mL at the first hour and sustainably from 5.5 to 20.2 nmol/mL for the following hours, which is controlled by feeding the aqCT amount and gel properties. GPT/α-CD/aqCT gels achieved significant antioxidant activity through a 100% scavenging DPPH radical. In addition, all gels are non-cytotoxic with a cell viability more than 85%. Especially, the GPT3.75α-CD10.5aqCT gels with aqCT amount of 3.1-12.5 mg/mL immensely enhanced the cell proliferation of GPT3.75α-CD10.5 gel without extract. These results suggest that the inherent bioactivities of aqCT endowed the resulting GPT/α-CD/aqCT gels with effective antioxidant and high biocompatibility, and natural polyphenols sustainably release a unique platform for a drug delivery system or other biomedical applications.

Unsubstituted and Methylated PAHs in Surface Sediment of Urban Rivers in the Red River Delta (Hanoi, Vietnam): Concentrations, Profiles, Sources, and Ecological Risk Assessment.

Unsubstituted and methylated polycyclic aromatic hydrocarbons (22 PAHs and 17 Me-PAHs) were examined in surface sediments collected from the Red River and four inner-city rivers of Hanoi City, Vietnam. Concentrations of total PAHs and Me-PAHs ranged from 52 to 920 (median 710) and from 70 to 2600 (median 1000) ng/g dry weight in samples of dry and wet seasons, respectively. Significant correlation was observed between total PAHs and organic carbon contents (Spearman's ρ = 0.782; p < 0.05). PAHs were more abundant than Me-PAHs in all samples and dominated by 4-6 ring compounds. The most predominant PAHs were benzo[ghi]perylene, benzo[b/j]fluoranthene, chrysene, pyrene, fluoranthene, and phenanthrene. Methylated derivatives of naphthalene, phenanthrene, anthracene, and benz[a]anthracene were frequently detected. The patterns of PAHs indicated principal pyrogenic sources (notably gasoline exhaust) in this highly urbanized area. The occurrence of several PAHs were occasionally associated with adverse effects on benthic organisms of the inner-city rivers.

Dynamic Prediction of Death in Patients With Tuberculous Meningitis Using Time-updated Glasgow Coma Scale and Plasma Sodium Measurements.

BACKGROUND: Pretreatment predictors of death from tuberculous meningitis (TBM) are well established, but whether outcome can be predicted more accurately after the start of treatment by updated clinical variables is unknown. Hence, we developed and validated models that dynamically predict mortality using time-updated Glasgow Coma Scale (GCS) and plasma sodium measurements, together with patient baseline characteristics. METHODS: We included 1048 adults from 4 TBM studies conducted in southern Vietnam from 2004 to 2016. We used a landmarking approach to predict death within 120 days after treatment initiation using time-updated data during the first 30 days of treatment. Separate models were built for patients with and without human immunodeficiency virus (HIV) infection. We used the area under the receiver operating characteristic curve (AUC) to evaluate performance of the models at days 10, 20, and 30 of treatment to predict mortality by 60, 90, and 120 days. Our internal validation was corrected for overoptimism using bootstrap. We provide a web-based application that computes mortality risk within 120 days. RESULTS: Higher GCS indicated better prognosis in all patients. In HIV-infected patients, higher plasma sodium was uniformly associated with good prognosis, whereas in HIV-uninfected patients the association was heterogeneous over time. The bias-corrected AUC of the models ranged from 0.82 to 0.92 and 0.81 to 0.85 in HIV-uninfected and HIV-infected individuals, respectively. The models outperformed the previously published baseline models. CONCLUSIONS: Time-updated GCS and plasma sodium measurements improved predictions based solely on information obtained at diagnosis. Our models may be used in practice to define those with poor prognosis during treatment.

Novel multiplex real-time PCR assays reveal a high prevalence of diarrhoeagenic Escherichia coli pathotypes in healthy and diarrhoeal children in the south of Vietnam.

BACKGROUND: Diarrhoeagenic Escherichia coli (DEC) infections are common in children in low-middle income countries (LMICs). However, detecting the various DEC pathotypes is complex as they cannot be differentiated by classical microbiology. We developed four multiplex real-time PCR assays were to detect virulence markers of six DEC pathotypes; specificity was tested using DEC controls and other enteric pathogens. PCR amplicons from the six E. coli pathotypes were purified and amplified to be used to optimize PCR reactions and to calculate reproducibility. After validation, these assays were applied to clinical samples from healthy and diarrhoeal Vietnamese children and associated with clinical data. RESULTS: The multiplex real-time PCRs were found to be reproducible, and specific. At least one DEC variant was detected in 34.7% (978/2815) of the faecal samples from diarrhoeal children; EAEC, EIEC and atypical EPEC were most frequent Notably, 41.2% (205/498) of samples from non-diarrhoeal children was positive with a DEC pathotype. In this population, only EIEC, which was detected in 34.3% (99/289) of diarrhoeal samples vs. 0.8% (4/498) non-diarrhoeal samples (p < 0.001), was significantly associated with diarrhoea. Multiplex real-time PCR when applied to clinical samples is an efficient and high-throughput approach to DEC pathotypes. CONCLUSIONS: This approach revealed high carriage rates of DEC pathotypes among Vietnamese children. We describe a novel diagnostic approach for DEC, which provides baseline data for future surveillance studies assessing DEC burden in LMICs.