Search

BackgroundMalaria elimination using current tools has stalled in many areas. Ivermectin (IVM) is a broad-antiparasitic drug and mosquitocide that has been proposed as a tool for reaching malaria elimination. Under laboratory conditions, IVM has been shown to reduce the survival ofAnophelespopulations that have fed on IVM-treated mammals. Treating cattle with IVM has been proposed as an important contribution to malaria vector management, however, the impacts of IVM in this animal health use-case had been untested in field trials in Southeast Asia.MethodsThrough a randomized village-based trial, this study aimed to quantify the effect of IVM-treated cattle on anopheline populations in treated vs. untreated villages in Central Vietnam. Local zebu cattle in six rural villages were included in this study. Cattle were treated with IVM at established veterinary dosages in three villages and in three additional villages, cattle were untreated as controls. The mosquito populations in all villages were sampled using cattle-baited traps for six days before, and six days after a 2-day treatment IVM-administration (intervention) period. Vector species were characterized using taxonomic keys. The impact of the intervention was analyzed using a difference-in-differences (DID) approach with generalized estimating equations (Poisson distribution with bootstrapped errors).ResultsAcross the treated villages, 1,112 of 1,527 censused cows (73% overall; range 67% to 83%) were treated with IVM. In both control and treated villages, there was a 30% to 40% decrease in total anophelines captured in the post-intervention period as compared to the pre-intervention period. In the control villages, there were 1873 captured pre-intervention and 1079 captured during the post-intervention period. In the treated villages, there were 1594 captured pre-intervention, and 1101 captured during the post-intervention period. The DID model analysis comparing total captures between arms was not statistically significant (p = 0.67). Secondary outcomes of vector diversity found that in four villages (two treated and two control) there were statistically significant changes in the anopheline population diversity (p < 0.05) based on Shannon’s diversity index. Two villages (one treated and one control) had a statistically significant increase in diversity and two villages (one treated and one control) had a significant decrease in population diversity (p < 0.05). There were no clear trends in treated or untreated vector population evenness or richness estimates.ConclusionsUnexpectedly large decreases in trapping counts post-intervention across all study villages impacted the ability of this study to quantify any differential impacts. As such, the results of this study do not provide evidence that treating cattle in villages with IVM reduces nightly captures from cattle-baited traps of female anopheles mosquitoes when compared to control villages. The lack of differential impacts may be due to several factors including the short half-life of IVM, crossover in mosquito populations between treated and control villages, feeding preferences of the mosquitoes, and mass-action effects from extensive mosquito trapping. Future studies should plan to treat at least 80% of the cattle in the village and evaluate the relationship between dose-density and mosquito prevalence. Additional studies should investigate whether IVM differentially impacts vector species at a population level.

Background: Strongyloidiasis, a neglected disease caused by intestinal nematodes of the genus Strongyloides, is endemic to tropical and subtropical areas such as Vietnam. The morphological diagnosis of larvae by Hara Mori culture technique and microscopicare are considered the standard diagnostic procedures in the endemic areas of Strongyloides spp. However, they could only identify the genus, not the species of Strongyloides. DNA-molecular techniques which are highly sensitive and more cost-effective have been increasingly utilized in detection of Strongyloides species. This study aims to determine prevalence and the species of Strongyloides among resident population in Duc Hoa district, Long An province, Southern Vietnam. Methods: A cross-sectional study was conducted using 1,190 stool samples collected in Duc Hoa district, Long An province, Vietnam, from July, 2017 to November 2018. The stool specimens were transported to the Laboratory of Medical Parasitology, Pham Ngoc Thach University of Medicine within two hours of collection at an appropriate temperature of 25 oC. All samples were stored at 2 - 8°C and processed within 48 hours for microscopic examination. Molecular detection was carried out at Laboratory of Molecular Biology, Pham Ngoc Thach University of Medicine, Hochiminh city, VietNam. Results: Of the 1,190 samples tested, Strongyloides spp. larvae were detected in 79 specimens (6.6%) by two classical parasitological methods, namely direct microscopy and the modified Harada-Mori filter paper culture. DNA was extracted from 70 of the 79 samples of Strongyloides spp. larvae, which was subsequently characterized by real-time PCR amplification of the 18S and 28S regions of the rDNA gene. The results showed that 97.1% of the DNA samples were S. stercoralis, 2.9% were co-infections with S. ratti and S. stercoralis, and 2.9% belonged to S. ratti. For all 14 isolates, nucleotide sequencing was compared with other human pathogenic species of Strongyloides whose sequences are available in GenBank. The identity of 12/14 sequences were confirmed as S. stercoralis with a high level of similarity (91.3% - 100%) and over 98% for S. ratti. Between the two co-infection samples, the higher similarity belonged to S. stercoralis. Conclusion: A molecular amplification of small subunit ribosome RNA followed by sequence analysis has been proved to be a suitable method for discrimination of Strongyloides spp. retrieved from stool samples.

Toxocara canis, a common roundworm that mainly causes toxocariasis, is a zoonotic parasite found worldwide. Humans, an accidental host, can acquire T. canis infection through accidental ingestion of T. canis-embryonated egg-contaminated food, water, and soil, and by encapsulated larvae in a paratenic host's viscera or meat. Long-term residence of T. canis larvae in a paratenic host's lungs may induce pulmonary inflammation that contributes to lung injury, airway inflammatory hyperresponsiveness, and collagen deposition in mice and clinical patients. This study intended to investigate the relationship between T. canis infection and allergic asthma in BALB/c mice inoculated with high, moderate, and low doses of T. canis eggs for a 13-week investigation. The airway hyperresponsiveness (AHR) to methacholine, collagen deposition, cytokine levels, and pathological changes in lung tissues was assessed in infected mice at weeks 1, 5, and 13 postinfection. The cell composition in bronchoalveolar lavage fluid of infected mice was assessed at weeks 5 and 13 postinfection. Compared with uninfected control mice, all groups of T. canis-infected mice exhibited significant AHR, a dose-dependent increase in eosinophilic infiltration leading to multifocal interstitial and alveolar inflammation with abundant mucus secretion, and collagen deposition in which the lesion size increased with the infective dose. Infected mice groups also showed significant expressions of eotaxin and type 2 T-helper-dominant cytokines such as interleukin (IL)-4, IL-5, and IL-13. Overall, these results suggest that T. canis larval invasion of the lungs may potentially cause pulmonary inflammatory injury and could subsequently contribute to the development of allergic manifestations such as asthma.

Background: Human toxocariasis is prevalent in many countries but this disease has been rarely reported from Vietnam. We aimed to investigate the clinical and laboratory findings and assess possible association between these findings in patients with toxocariasis in Vietnam. Methods: A prospectively study, between October 2017 and June 2019 was performed involving 120 toxocariasis patients at Medic Medical Center, Ho Chi Minh City, Vietnam. The diagnosis of toxocariasis was established based on clinical, laboratory (eosinophilia, raised IgE concentration) and serological (positive Toxocara IgG ELISA test) evaluation as well as the exclusion of other helminthic coinfection. Results: The most frequently reported manifestation was of skin (n = 93, 77.5%), including urticarial (n= 69, 57.5%) followed by neurologic, gastrointestinal and pulmonary signs/symptoms. Hepatic involvement occurred in 8.3% of the patients. No significant relationship between clinical findings and laboratory parameters was found except the higher values of eosinophil count and IgE concentration among patients with liver involvement. There was a significant relationship between eosinophil count and IgE concentration (r=0.389, P

Apical membrane antigen-1 of Plasmodium falciparum (PfAMA-1) is a leading malaria vaccine candidate antigen. However, the genetic diversity of pfama-1 and associated antigenic variation in global P. falciparum field isolates are major hurdles to the design of an efficacious vaccine formulated with this antigen. Here, we analyzed the genetic structure and the natural selection of pfama-1 in the P. falciparum population of Vietnam. A total of 37 distinct haplotypes were found in 131 P. falciparum Vietnamese isolates. Most amino acid changes detected in Vietnamese pfama-1 were localized in the ectodomain, domains I, II, and III. Overall patterns of major amino acid changes in Vietnamese pfama-1 were similar to those of global pfama-1, but the frequencies of the amino acid changes slightly differed by country. Novel amino acid changes were also identified in Vietnamese pfama-1. Vietnamese pfama-1 revealed relatively lower genetic diversity than currently analyzed pfama-1 in other geographical regions, and suggested a distinct genetic differentiation pattern. Evidence for natural selection was detected in Vietnamese pfama-1, but it showed purifying selection unlike the global pfama-1 analyzed so far. Recombination events were also found in Vietnamese pfama-1. Major amino acid changes that were commonly identified in global pfama-1 were mainly localized to predicted B-cell epitopes, RBC-binding sites, and IUR regions. These results provide important information for understanding the genetic nature of the Vietnamese pfama-1 population, and have significant implications for the design of a vaccine based on PfAMA-1.

The rise in Plasmodium falciparum resistance to dihydroartemisinin-piperaquine in Vietnam justifies the need to evaluate alternative artemisinin-based combination therapies. Between July 2018 and October 2019, a single-arm trial of pyronaridine-artesunate (Pyramax, PA) was conducted in Dak Nong province, Vietnam. PA (3-day course) was administered to adults and children infected with P. falciparum. PA was well tolerated by the participants. The proportion of patients with Day 42 PCR-corrected adequate clinical and parasitological response was 95.2% (95% confidence interval [CI], 82.3 to 98.8, n = 40/42) for treating falciparum malaria. The median parasite clearance half-life was 6.7 h (range, 2.6 to 11.9) and the median parasite clearance time was 72 h (range, 12 to 132) with 44.9% (22/49) of patients having positive blood films at 72 h. The two patients that recrudesced had comparable Day 7 blood pyronaridine concentrations (39.5 and 39.0 ng/ml) to the 40 patients who did not recrudesce (median 43.4 ng/ml, 95% CI, 35.1 to 54.9). Ring-stage and piperaquine survival assays revealed that of the 29 P. falciparum isolates collected from the patients before PA treatment, 22 (75.9%) had reduced susceptibility to artemisinins and 17 (58.6%) were resistant to piperaquine. Genotyping confirmed that 92.0% (46/50) of falciparum patients were infected with parasites bearing the Pfkelch13 C580Y mutation associated with artemisinin resistance. Of these, 56.0% (28/50) of the isolates also had multiple copies of the plasmepsin 2/3 genes responsible for piperaquine resistance. Overall, PA was effective in treating P. falciparum in the Central Highlands of Vietnam. (This study has been registered at AustralianClinicalTrials.gov.au under trial ID ACTRN12618001429246.).

Apical membrane antigen-1 of Plasmodium falciparum (PfAMA-1) is a leading malaria vaccine candidate antigen. However, the genetic diversity of pfama-1 and associated antigenic variation in global P. falciparum field isolates are major hurdles to the design of an efficacious vaccine formulated with this antigen. Here, we analyzed the genetic structure and the natural selection of pfama-1 in the P. falciparum population of Vietnam. A total of 37 distinct haplotypes were found in 131 P. falciparum Vietnamese isolates. Most amino acid changes detected in Vietnamese pfama-1 were localized in the ectodomain, domains I, II, and III. Overall patterns of major amino acid changes in Vietnamese pfama-1 were similar to those of global pfama-1, but the frequencies of the amino acid changes slightly differed by country. Novel amino acid changes were also identified in Vietnamese pfama-1. Vietnamese pfama-1 revealed relatively lower genetic diversity than currently analyzed pfama-1 in other geographical regions, and suggested a distinct genetic differentiation pattern. Evidence for natural selection was detected in Vietnamese pfama-1, but it showed purifying selection unlike the global pfama-1 analyzed so far. Recombination events were also found in Vietnamese pfama-1. Major amino acid changes that were commonly identified in global pfama-1 were mainly localized to predicted B-cell epitopes, RBC-binding sites, and IUR regions. These results provide important information for understanding the genetic nature of the Vietnamese pfama-1 population, and have significant implications for the design of a vaccine based on PfAMA-1.

Author(s): Jacob, Christopher G; Thuy-Nhien, Nguyen; Mayxay, Mayfong; Maude, Richard J; Quang, Huynh Hong; Hongvanthong, Bouasy; Vanisaveth, Viengxay; Ngo Duc, Thang; Rekol, Huy; van der Pluijm, Rob; von Seidlein, Lorenz; Fairhurst, Rick; Nosten, Francois; Hossain, Md Amir; Park, Naomi; Goodwin, Scott; Ringwald, Pascal; Chindavongsa, Keobouphaphone; Newton, Paul; Ashley, Elizabeth; Phalivong, Sonexay; Maude, Rapeephan; Leang, Rithea; Huch, Cheah; Dong, Le Thanh; Nguyen, Kim-Tuyen; Nhat, Tran Minh; Hien, Tran Tinh; Nguyen, Hoa; Zdrojewski, Nicole; Canavati, Sara; Sayeed, Abdullah Abu; Uddin, Didar; Buckee, Caroline; Fanello, Caterina I; Onyamboko, Marie; Peto, Thomas; Tripura, Rupam; Amaratunga, Chanaki; Myint Thu, Aung; Delmas, Gilles; Landier, Jordi; Parker, Daniel M; Chau, Nguyen Hoang; Lek, Dysoley; Suon, Seila; Callery, James; Jittamala, Podjanee; Hanboonkunupakarn, Borimas; Pukrittayakamee, Sasithon; Phyo, Aung Pyae; Smithuis, Frank; Lin, Khin; Thant, Myo; Hlaing, Tin Maung; Satpathi, Parthasarathi; Satpathi, Sanghamitra; Behera, Prativa K; Tripura, Amar; Baidya, Subrata; Valecha, Neena; Anvikar, Anupkumar R; Ul Islam, Akhter; Faiz, Abul; Kunasol, Chanon; Drury, Eleanor; Kekre, Mihir; Ali, Mozam; Love, Katie; Rajatileka, Shavanthi; Jeffreys, Anna E; Rowlands, Kate; Hubbart, Christina S; Dhorda, Mehul; Vongpromek, Ranitha; Kotanan, Namfon; Wongnak, Phrutsamon; Almagro Garcia, Jacob; Pearson, Richard D; Ariani, Cristina V; Chookajorn, Thanat; Malangone, Cinzia; Nguyen, T; Stalker, Jim; Jeffery, Ben | Abstract: BackgroundNational Malaria Control Programmes (NMCPs) currently make limited use of parasite genetic data. We have developed GenRe-Mekong, a platform for genetic surveillance of malaria in the Greater Mekong Subregion (GMS) that enables NMCPs to implement large-scale surveillance projects by integrating simple sample collection procedures in routine public health procedures.MethodsSamples from symptomatic patients are processed by SpotMalaria, a high-throughput system that produces a comprehensive set of genotypes comprising several drug resistance markers, species markers and a genomic barcode. GenRe-Mekong delivers Genetic Report Cards, a compendium of genotypes and phenotype predictions used to map prevalence of resistance to multiple drugs.ResultsGenRe-Mekong has worked with NMCPs and research projects in eight countries, processing 9623 samples from clinical cases. Monitoring resistance markers has been valuable for tracking the rapid spread of parasites resistant to the dihydroartemisinin-piperaquine combination therapy. In Vietnam and Laos, GenRe-Mekong data have provided novel knowledge about the spread of these resistant strains into previously unaffected provinces, informing decision-making by NMCPs.ConclusionsGenRe-Mekong provides detailed knowledge about drug resistance at a local level, and facilitates data sharing at a regional level, enabling cross-border resistance monitoring and providing the public health community with valuable insights. The project provides a rich open data resource to benefit the entire malaria community.FundingThe GenRe-Mekong project is funded by the Bill and Melinda Gates Foundation (OPP11188166, OPP1204268). Genotyping and sequencing were funded by the Wellcome Trust (098051, 206194, 203141, 090770, 204911, 106698/B/14/Z) and Medical Research Council (G0600718). A proportion of samples were collected with the support of the UK Department for International Development (201900, M006212), and Intramural Research Program of the National Institute of Allergy and Infectious Diseases.

Objectives Human toxocariasis is a widespread zoonosis for which a chemotherapy decision and therapy effectiveness are difficult to determine. We aimed to investigate the kinetic profile of clinical and laboratory findings and treatment outcome of patients with toxocariasis in Vietnam. Methods The prospective study was conducted between October 2017 and June 2019. The diagnosis of toxocariasis was established based on clinical, laboratory (eosinophilia, raised IgE concentration) and serological (positive Toxocara IgG ELISA) evaluation as well as the exclusion of another helminthic coinfection. The patients were followed up after seven days, then one, three, and six months after chemotherapy by thiabendazole. Results The study involved 80 patients with a mean age of 41.6 ± 15.2 years of whom 58.8% were female. At three and six months after chemotherapy, most patients demonstrated resolution of clinical signs and symptoms, eosinophil count and IgE concentration but not in the proportion of IgG seropositivity. Skin lesions and eosinophilia resolved earlier than the other symptoms (one month after treatment). About four-fifths of the patients were "cured" after three and six months of follow-up. 33.8% showed side effects to thiabendazole therapy but no severe events were reported. The most common adverse reaction was neurologic symptoms followed by gastrointestinal or skin manifestations which lasted as long as 4 days. Conclusions In toxocariasis patients, cutaneous manifestations and eosinophilia resolve more rapidly than other clinical and laboratory findings while IgG titre has a very slow kinetic after therapy. Thiabendazole seems to be a potential alternative for the treatment of human toxocariasis.

Fasciola hepatica, F. gigantica and “intermediate” forms of Fasciola species, are responsible for fascioliasis in ruminant animals and humans in many countries. There are two forms: “natural/admixed” and “introgressive” hybridization. Vietnam is a "hot spot" country, where the “hybrid” Fasciola forms have been found nationwide. Determination of phylogenetic relationships and genetic distance of these hybrid forms with those in family Fasciolidae and class Trematoda (Phyllum Platyhelminthes) is necessary to confirm their taxonomic classification. In this study, we sequenced the ITS1 and ITS2 for discrimination of two kinds of hybrid Fasciola mentioned above. The deduced amino acid sequences of the entire mitochondrial genes, cytochrome b (cob), nicotinamide dehydrogenase 1 (nad1) and cytochrome oxidase 1 (cox1), from two hybrid Fasciola spp. including Fsp-DL11-VN (from buffalo, admixed hybrid), Fsp-FH1-VN (from human, introgressive hybrid) and “pure” F. gigantica (Fgig-T4V-VN, from cattle) were obtained and combined (cob+nad1+cox1). These sequences were used as markers for phylogenetic analysis together with 28 representative isolates/species of Fasciolidae and Trematoda. Genetic distances between 13 isolates/species in the family Fasciolidae have also been determined to accurately account for their species-relationships. As results, pairwise genetic distance calculation showed that the distance rate was only 0.4% – 0.7% between Fasciola spp. hybrids of Vietnam and China, slightly higher, 1.3 – 2.0% with “pure” F. gigantica, whilist this rate was quite high compared with F. hepatica (5.7% – 5.9%), with Fasciolopsis buski (20.6% – 21.0%), and two other fasciolids, Fasciola jacksoni and Fascioloides magna (11.0% – 12.6%). The phylogenetic tree of 31 strains/species showed 5 distinct groups, corresponding to 5 families, ie., Fasciolidae, Echinostomatidae, Echinochasmidae, Heterophyidae, Opisthorchiidae and an outgroup, Schistosomatidae. Two "hybrid" Fasciola species of Vietnam (Fsp-FH1-VN and Fsp-DL11-VN) grouped with the reference "hybrid" Fsp-GHL-CN strain of China; and “pure” Fgig-T4V-VN with the two “pure” F. gigantica, Fgig-Bali-ID (Indonesia) and Fgig-GX-CN (China). These results confirmed that “hybrid” Fasciola spp. were of the maternally inherited mitochondrial lineage from F. gigantica.

Plasmodium falciparum resistance to dihydroartemisinin-piperaquine has spread through the Greater Mekong Subregion to southwestern Vietnam. In 2018 to 2019, we collected 127 P. falciparum isolates from Dak Nong (36), Dak Lak (55), Gia Lai (13), and Kon Tum (23) provinces in Vietnam’s Central Highlands and found parasites bearing the Pfkelch13 C580Y mutation and multiple plasmepsin 2/3 genes (mean prevalence, 17.9%; range, 4.3% to 27.8%), conferring resistance to dihydroartemisinin-piperaquine.

Dihydroartemisinin-piperaquine (DHA-PPQ) is a current frontline drug recommended in global by WHO for the treatment ofPlasmodium falciparummalaria (WHO, 2015), but is now failing in Vietnam’s provinces where border Cambodia, and has emerged and spread. The purpose of this study was to evaluate the efficacy and molecular markers of DHA-PPQ failures in Dak Lak province. Methods: A study design of non-randomized controlled study design for the 42 day-course follow-up in vivo test, and the molecular markers analysis. Findings: The data showed that adequate clinical and parasitological response was sharply declined to 12,1%, the proportion of late clinical failure was 51.5%, and 36.4% of patients had late parasitological. The proportion of positive parasitemia at D3 was 37%, the slope half-life was 5.36 hrs, and the progressive parasite clearance (PC) PC50, PC75, PC 90, PC95, and PC99 were 13.24; 19.29; 25.69; 29.97 and 39.15 hours, respectively. Molecular markers of C580Y Kelch mutation were observed in 100% (50/50) of the patients, the increased of Plasmepsine 2 copy number variation (CNV) was 72% (36/50), and the proportion of patients who had both K13 and increased Plasmepsine 2 CNV was 72% (36/50). Conclusions: The DHA-PPQ efficacy severely decreased to 12.1%, overall treatment failure was 87.9% with the prominent C580Y mutant plus increased Plasmepsine 2 CNV in delayed asexualP. falciparumparasite clearance. These obvious data suggest the urgency to change antimalarial policy in DHA-PPQ resistance zones, especially in Dak Lak province.

Toxocara canis, a common roundworm that mainly causes toxocariasis, is a zoonotic parasite found worldwide. Humans, an accidental host, can acquire T. canis infection through accidental ingestion of T. canis–embryonated egg-contaminated food, water, and soil, and by encapsulated larvae in a paratenic host’s viscera or meat. Long-term residence of T. canis larvae in a paratenic host’s lungs may induce pulmonary inflammation that contributes to lung injury, airway inflammatory hyperresponsiveness, and collagen deposition in mice and clinical patients. This study intended to investigate the relationship between T. canis infection and allergic asthma in BALB/c mice inoculated with high, moderate, and low doses of T. canis eggs for a 13-week investigation. The airway hyperresponsiveness (AHR) to methacholine, collagen deposition, cytokine levels, and pathological changes in lung tissues was assessed in infected mice at weeks 1, 5, and 13 postinfection. The cell composition in bronchoalveolar lavage fluid of infected mice was assessed at weeks 5 and 13 postinfection. Compared with uninfected control mice, all groups of T. canis–infected mice exhibited significant AHR, a dose-dependent increase in eosinophilic infiltration leading to multifocal interstitial and alveolar inflammation with abundant mucus secretion, and collagen deposition in which the lesion size increased with the infective dose. Infected mice groups also showed significant expressions of eotaxin and type 2 T-helper–dominant cytokines such as interleukin (IL)-4, IL-5, and IL-13. Overall, these results suggest that T. canis larval invasion of the lungs may potentially cause pulmonary inflammatory injury and could subsequently contribute to the development of allergic manifestations such as asthma.

Ivermectin is a low-cost and nontoxic mosquitocide that may have a role in malaria elimination. However, the extent to which this drug impacts the mortality of Anopheles dirus and Anopheles epiroticus, two important malaria vectors in Southeast Asia, is unknown. This study compared and quantified anopheline mortality after feeding on ivermectin-treated cattle and control cattle in Vietnam. Local anopheline colonies fed on cattle 1 to 3, 6 to 8, 13 to 15, 20 to 22, and 28 to 30 days after injection (DAI) with ivermectin (intervention) or saline (control). An. dirus that fed on ivermectin-treated cattle had higher mortality rates than controls for up to 20 DAI (P < 0.05); An. epiroticus that fed on ivermectin-treated cattle had consistently higher mortality rates than controls for up to 8 DAI (P < 0.05). Feeding on ivermectin-treated cattle increased the mortality rate of these vector species for biologically relevant time periods. Therefore, ivermectin has the potential to become an important tool for integrated vector management.

The Asia–Pacific region faces formidable challenges in achieving malaria elimination by the proposed target in 2030. Molecular surveillance of Plasmodium parasites can provide important information on malaria transmission and adaptation, which can inform national malaria control programmes (NMCPs) in decision-making processes. In November 2019 a parasite genotyping workshop was held in Jakarta, Indonesia, to review molecular approaches for parasite surveillance and explore ways in which these tools can be integrated into public health systems and inform policy. The meeting was attended by 70 participants from 8 malaria-endemic countries and partners of the Asia Pacific Malaria Elimination Network. The participants acknowledged the utility of multiple use cases for parasite genotyping including: quantifying the prevalence of drug resistant parasites, predicting risks of treatment failure, identifying major routes and reservoirs of infection, monitoring imported malaria and its contribution to local transmission, characterizing the origins and dynamics of malaria outbreaks, and estimating the frequency of Plasmodium vivax relapses. However, the priority of each use case varies with different endemic settings. Although a one-size-fits-all approach to molecular surveillance is unlikely to be applicable across the Asia–Pacific region, consensus on the spectrum of added-value activities will help support data sharing across national boundaries. Knowledge exchange is needed to establish local expertise in different laboratory-based methodologies and bioinformatics processes. Collaborative research involving local and international teams will help maximize the impact of analytical outputs on the operational needs of NMCPs. Research is also needed to explore the cost-effectiveness of genetic epidemiology for different use cases to help to leverage funding for wide-scale implementation. Engagement between NMCPs and local researchers will be critical throughout this process.

The aim of this study is to delineate 'admixed hybrid' and 'introgressive' Fasciola genotypes present in the Fasciola population in Vietnam. Adult liver flukes collected from ruminants in 18 Provinces were morphologically sorted out by naked eyes for small (S), medium (M) and large (L) body shapes; and human samples (n=14) from patients. Nuclear ribosomal (rDNA) ITS1 and ITS2, and mitochondrial (mtDNA) nad1 markers were used for determination of their genetic status. Total 4,725 worm samples of ruminants were tentatively classified by their size: 6% (n=284) small (S)-, 13% (n=614) medium (M)-, and 81% (n=3,827) large (L)-forms. All the representative (n=120, as 40 each group) and 14 human specimens, possessed maternal mtDNA of only F. gigantica and none of F. hepatica. Paternally, all (100%) of the L-(n=40) and 77.5% (n=31) of the M-flukes had single F. gigantica rDNA indicating 'pure' F. gigantica. A majority (90%, n=36) of the S- and 15% (n=6) of the M-worms had single F. hepatica rDNA, indicating their introgressive; the rest (10%, n=4) of the S- and 7.5% (n=3) of the M-flukes had mixture of both F. gigantica and F. hepatica rDNAs, confirming their admixed hybrid genetic status. Fourteen human samples revealed 9 (64%) of pure F. gigantica, 3 (22%) of introgressive and 2 (14%) of admixed hybrid Fasciola spp. By the present study, it was confirmed that the small worms, which are morphologically identical with F. hepatica, are admixed and/or introgressive hybrids of Fasciola spp., and able to be the pathogens of human fascioliasis.

Asymptomatic parasite carriers represent a “silent” infective reservoir for malaria transmission and contributes to malaria persistence. However, limited data are available on asymptomatic malaria in Vietnam. Between November 2018 and March 2019, we conducted a malaria epidemiological survey of asymptomatic people (children ≥ 10 years old and adults ≥18 years old, n = 2,809) residing in three communes in Tuy Duc district, Dak Nong province in the Central Highlands of Vietnam. Based on the national stratification of malaria risk, Dak Buk So, Dak Ngo and Quang Truc communes were classified by the National Malaria Control Programme as low, moderate and high malaria endemic areas, respectively. Using participants’ finger prick blood samples, malaria parasites were detected by one-step reverse transcription-quantitative polymerase chain reaction (RT-qPCR). The median age (Interquartile Range) for adults and children were 35 years (26–50) and 12 years (11–14), respectively. The prevalence of asymptomatic malaria was 1.7% (22/1,328), 3.5% (31/890) and 12.2% (72/591) for participants from Dak Buk So, Dak Ngo and Quang Truc, respectively. The prevalence of asymptomatic malaria was lower in children compared to adults: 2.6% (9/352) versus 4.7% (116/2,457) (Odds Ratio 0.53, 95% Confidence Interval 0.28 to1.02). Ownership of long-lasting insecticide-treated bed nets and hammocks was 97.1%, 99.0% and 94.7% for participants in Dak Buk So, Dak Ngo and Quang Truc, respectively, however, only 66.0%, 57.3% and 42.8% of the participants reported using bed nets every night. Of the several risk factors examined, going to the forest two weeks prior to enrolment into the study and sleeping in the forest had a significant association with participants being infected with asymptomatic malaria in Quang Truc, but not in the other two communes. Knowledge of the prevalence and distribution of asymptomatic malaria will help design and evaluate future intervention strategies for malaria elimination in Vietnam.

Vietnam achieved outstanding success against malaria in the last few decades. The mortality and morbidity of malaria in Vietnam have decreased remarkably in recent years, but malaria is still a major public health concern in the country, particularly in the Central Highlands region. In this study, molecular analyses of malaria parasites in the Central Highlands were performed to understand the population structure and genetic diversity of the parasites circulating in the region. Plasmodium falciparum (68.7%) and P. vivax (27.4%) along with mixed infections with P. falciparum/P. vivax (3.9%) were detected in 230 blood samples from patients with malaria. Allele-specific nested-polymerase chain reaction (PCR) or PCR-restriction fragment length polymorphism (PCR-RFLP) analyses of pfmsp-1, pfama-1, pvcsp, and pvmsp-1 revealed complex genetic makeup in P. falciparum and P. vivax populations of Vietnam. Substantial multiplicity of infection (MOI) was also identified, suggesting significant genetic diversity and polymorphism of P. falciparum and P. vivax populations in the Central Highlands of Vietnam. These results provide fundamental insight into the current patterns of dispersion and genetic nature of malaria parasites as well as for the development of malaria elimination strategies in the endemic region.

SummaryBackgroundA multidrug resistant co-lineage of Plasmodium falciparum malaria, named KEL1/PLA1, spread across Cambodia c.2008-2013, causing high treatment failure rates to the frontline combination therapy dihydroartemisinin-piperaquine. Here, we report on the evolution and spread of KEL1/PLA1 in subsequent years.MethodsWe analysed whole genome sequencing data from 1,673 P. falciparum clinical samples collected in 2008-2018 from northeast Thailand, Laos, Cambodia and Vietnam. By investigating genome-wide relatedness between parasites, we inferred patterns of shared ancestry in the KEL1/PLA1 population.FindingsKEL1/PLA1 spread rapidly from 2015 into all of the surveyed countries and now exceeds 80% of the P. falciparum population in several regions. These parasites maintained a high level of genetic relatedness reflecting their common origin. However, several genetic subgroups have recently emerged within this co-lineage with diverse geographical distributions. Some of these emerging KEL1/PLA1 subgroups carry recent mutations in the chloroquine resistance transporter (crt) gene, which arise on a specific genetic background comprising multiple genomic regions.InterpretationAfter emerging and circulating for several years within Cambodia, the P. falciparum KEL1/PLA1 co-lineage diversified into multiple subgroups and acquired new genetic features including novel crt mutations. These subgroups have rapidly spread into neighbouring countries, suggesting enhanced fitness. These findings highlight the urgent need for elimination of this increasingly drug-resistant parasite co-lineage, and the importance of genetic surveillance in accelerating elimination efforts.FundingWellcome Trust, Bill & Melinda Gates Foundation, UK Medical Research Council, UK Department for International Development.Research in contextEvidence before this studyThis study updates our previous work describing the emergence and spread of a multidrug resistant P. falciparum co-lineage (KEL1/PLA1) within Cambodia up to 2013. Since then, a regional genetic surveillance project, GenRe-Mekong, has reported that markers of dihydroartemisinin-piperaquine (DHA-PPQ) resistance have increased in frequency in neighbouring countries. A PubMed search (terms: “artemisinin”, “piperaquine”, “resistance”, “southeast asia”) for articles listed since our previous study (from 30/10/2017 to 05/01/2019) yielded 28 results, including reports of a recent sharp decline in DHA-PPQ clinical efficacy in Vietnam; the spread of genetic markers of DHA-PPQ resistance into neighbouring countries by Imwong and colleagues; and multiple reports associating mutations in the crt gene with piperaquine resistance, including newly emerging crt variants in Southeast Asia.Added value of this studyWe analysed P. falciparum whole genomes collected up to early 2018 from Eastern Southeast Asia (Cambodia and surrounding regions), describing the fine-scale epidemiology of multiple KEL1/PLA1 genetic subgroups that have spread out from Cambodia since 2015 and taken over indigenous parasite populations in northeastern Thailand, southern and central Vietnam and parts of southern Laos. Several newly emerging crt mutations accompanied the spread and expansion of KEL1/PLA1 subgroups, suggesting an active proliferation of biologically fit, multidrug resistant parasites.Implications of all the available evidenceThe problem of P. falciparum multidrug resistance has dramatically worsened in Eastern Southeast Asia since previous reports. KEL1/PLA1 has diversified and spread widely across Eastern Southeast Asia since 2015, becoming the predominant parasite group in several regions. This may have been fuelled by continued parasite exposure to DHA-PPQ, resulting in sustained selection after KEL1/PLA1 became established. Continued drug pressure enabled the acquisition of further mutations, resulting in higher levels of resistance. These data demonstrate the value of pathogen genetic surveillance and the urgent need to eliminate these dangerous parasites.

Background Reduced artemisinin susceptibility and artemisinin-based combination therapy (ACT)-resistance against Plasmodium falciparum and chloroquine (CQ)-resistant P. vivax malaria has been reported in Vietnam. Two therapeutic efficacy studies were conducted in Thuan Bac District (Ninh Thuan Province, Vietnam) in 2015 and 2016 to determine the extent of reduced artemisinin susceptibility and ACT resistant falciparum malaria, and CQ-resistant vivax malaria were present. Methods Twenty-seven patients with falciparum malaria were randomized to receive artesunate alone (AS ~ 4 mg/kg/day) for 4 days followed by dihydroartemisinin (DHA) (2.2 mg/kg)–piperaquine (PPQ) (18 mg/kg) daily for 3 days or artemether (AM) (1.7 mg/kg)–lumefantrine (LUM) (12 mg/kg) twice daily for 3 days. Sixteen subjects with vivax malaria received CQ (total 25 mg/kg over 3 days). The therapeutic efficacy study for treating falciparum malaria was complemented with molecular analysis for artemisinin and piperaquine resistance, and in vitro drug susceptibility testing. Patient’s drug exposure following both falciparum and vivax treatment studies was determined. Results Twenty-five of 27 patients treated with the artemisinin regimens completed the 42-day follow-up period. None had parasites present on day 3 after commencing treatment with no incidence of recrudescence (100% curative rate). One patient on AS + DHA–PPQ was lost to follow-up and one patient had Plasmodium falciparum and Plasmodium vivax infection on day 0 by PCR. Of the vivax patients, 15 of 16 completed CQ treatment and two had a recurrence of vivax malaria on day 28, a failure rate of 13.3% (2/15). No mutations in the Pfkelch-13 gene for artemisinin resistance or exo-E415G gene polymorphism and amplification in plasmepsins 2 and 3 for piperaquine resistance were observed. In vitro testing of patient’s falciparum parasites indicated susceptibility (low IC50 nM values) to dihydroartemisinin, lumefantrine, piperaquine and pyronaridine. Patient’s drug exposure to artesunate and lumefantrine was comparable to published data, however, blood CQ concentrations were lower. Conclusions Clinical findings, molecular analysis and in vitro testing revealed that the falciparum parasites at Phuoc Chien Commune were artemisinin susceptible. The clinical failure rate of the 15 vivax patients who completed CQ treatment was 13%. Further studies are required to determine whether CQ-resistant vivax malaria is present at the commune. Electronic supplementary material The online version of this article (10.1186/s12936-019-2640-2) contains supplementary material, which is available to authorized users.

Background As part of a targeted malaria elimination project, mass drug administrations (MDAs) were conducted in Vietnam. The impact of MDAs on malaria transmission depends largely on the efficacy of the anti-malarial drug regimen, the malaria epidemiology in the site and the population coverage. To explore why some people participate in MDAs and others do not, a quantitative survey of the villagers’ perceptions was undertaken in Vietnam. Methods In 2013/2014 MDAs were conducted in a village in Binh Phuoc province and a village in Ninh Thuan province. Within three months of the drug administration, 59 respondents in a village in Binh Phuoc and 79 respondents in a village in Ninh Thuan were randomly selected and interviewed. Results Comprehension of the purpose of the intervention was of paramount importance for participation in the intervention. Respondents aware that the intervention aims to protect against malaria were significantly more likely to participate than respondents who were unaware of the MDA’s purpose. Secondly, how and by whom villagers were informed was critical for participation. There was a strong association between sensitization by an informant such as a member of the local health team with participation in the intervention. Conclusions The study suggests several approaches to increase participation in mass drug administration campaigns. Training trustworthy informants to sensitize the study population is critical to maximize village participation in this setting. To achieve high coverage the entire community must understand and agree with the intervention. Electronic supplementary material The online version of this article (doi:10.1186/s12936-016-1662-2) contains supplementary material, which is available to authorized users.

Background Artemisinin resistant Plasmodium falciparum has emerged in the countries of the Greater Mekong sub-region posing a serious threat to global malaria elimination efforts. The relationship of artemisinin resistance to treatment failure has been unclear. Methods In annual studies conducted in three malaria endemic provinces in the south of Vietnam (Binh Phuoc, Ninh Thuan and Gia Lai) between 2011 and 2015, 489 patients with uncomplicated P. falciparum malaria were enrolled in detailed clinical, parasitological and molecular therapeutic response assessments with 42 days follow up. Patients received the national recommended first-line treatment dihydroartemisinin-piperaquine for three days. Results Over the 5 years the proportion of patients with detectable parasitaemia on day 3 rose steadily from 38 to 57% (P

The spread of artemisinin-resistant Plasmodium falciparum compromises the therapeutic efficacy of artemisinin combination therapies (ACTs) and is considered the greatest threat to current global initiatives to control and eliminate malaria. This is particularly relevant in Vietnam, where dihydroartemisinin-piperaquine (DP) is the recommended ACT for P. falciparum infection. The propeller domain gene of K13, a molecular marker of artemisinin resistance, was successfully sequenced in 1,060 P. falciparum isolates collected at 3 malaria hot spots in Vietnam between 2009 and 2016. Eight K13 propeller mutations (Thr474Ile, Tyr493His, Arg539Thr, Ile543Thr, Pro553Leu, Val568Gly, Pro574Leu, and Cys580Tyr), including several that have been validated to be artemisinin resistance markers, were found. The prevalences of K13 mutations were 29% (222/767), 6% (11/188), and 43% (45/105) in the Binh Phuoc, Ninh Thuan, and Gia Lai Provinces of Vietnam, respectively. Cys580Tyr became the dominant genotype in recent years, with 79.1% (34/43) of isolates in Binh Phuoc Province and 63% (17/27) of isolates in Gia Lai Province carrying this mutation. K13 mutations were associated with reduced ring-stage susceptibility to dihydroartemisinin (DHA) in vitro and prolonged parasite clearance in vivo . An analysis of haplotypes flanking K13 suggested the presence of multiple strains with the Cys580Tyr mutation rather than a single strain expanding across the three sites.

Safe and effective antimalarial drugs are required for the treatment of pregnant women. We report a 3-day regimen of artesunate (4 mg/kg/day)-azithromycin (25 mg/kg/day) (ASAZ) to be efficacious (polymerase chain reaction-corrected cure rate of 96.7%) and well tolerated in the treatment of Plasmodium falciparum malaria in children (N = 11) and adults (N = 19), in Vietnam in 2010. In comparison, the cure rate for artesunate (4 mg/kg on day 0, 2 mg/kg on days 1-6) was 90.0% in children (N = 7) and adults (N = 23). Because azithromycin is considered safe in pregnancy, our findings provide further evidence that ASAZ should be evaluated for the treatment of pregnant women with malaria.

Background In Vietnam, the artemisinin-based combination therapy (ACT) of dihydroartemisinin-piperaquine is currently used for first-line treatment of uncomplicated Plasmodium falciparum malaria. However, limited efficacy and tolerability data are available on alternative forms of ACT in Vietnam in case there is a reduction in the susceptibility of dihydroartemisinin-piperaquine. A study was conducted to compare the efficacy and tolerability of two fixed-dose formulations of ACT, artemisinin–piperaquine (Artequick®, ARPQ) and artesunate-amodiaquine (Coarsucam™, ASAQ) for the treatment of P. falciparum malaria in south-central Vietnam. Methods A randomized, open-label trial was conducted comparing the efficacy of a two-day regimen of ARPQ (~2.8 mg/kg artemisinin plus ~17.1 mg/kg of piperaquine per day) and a three-day regimen of ASAQ (~4.7 mg/kg of artesunate plus ~12.6 mg/kg of amodiaquine per day) for the treatment of children and adults with uncomplicated falciparum malaria. Primary efficacy endpoint was day 42, PCR-corrected, parasitological cure rate. Secondary endpoints were parasite and fever clearance times and tolerability. Results Of 128 patients enrolled, 63 were administered ARPQ and 65 ASAQ. Of the patients who completed the 42 days follow-up period or had a recurrence of malaria, 55 were on ARPQ (30 children, 25 adults) and 59 were on ASAQ (31 children, 28 adults). Recrudescent parasitaemia was PCR-confirmed for one patient in each treatment group, with cure rates at day 42 of 98% (95% CI: 88–100) for both forms of ACT. The median parasite clearance time was significantly slower in the ARPQ group compared with the ASAQ group (48 h vs. 36 h, Pvs. 24 h, P = 0.07). The two forms of ACT were well tolerated with no serious adverse events. Conclusion Both forms of ACT were highly efficacious in the treatment of uncomplicated P. falciparum malaria. Although the two-day course of ARPQ was equally as effective as the three-day course of ASAQ, parasite and fever clearance times were shorter with ASAQ. Further studies are warranted in different regions of Vietnam to determine the nationwide efficacy of ASAQ. Trial registration Australian New Zealand Clinical Trials Registry Number, ACTRN12609000816257

To assess the in vivo sensitivity of Plasmodium falciparum to mefloquine and artesunate in a hyperendemic area of southern Viet Nam, we studied 41 children and 21 adults from a remote commune who had uncomplicated falciparum malaria without previous treatment. Patients were randomly allocated to artesunate (4 mg/kg on day 0 and 2 mg/kg on days 1-4) or mefloquine (10 mg/kg followed by 5 mg/kg at 6 h). Serial assessments were performed over 28 days. Of 31 patients allocated artesunate, nine (29%) redeveloped parasitaemia during follow-up compared with 23% (seven of 30) who received mefloquine. Of the 41 children, 15 (37%) had recrudescence/re-infection compared with only one of 20 adults (5%; p < 0.001). Significantly more children than adults failed on mefloquine treatment (37% vs 0%; p = 0.021) and one case showed RIII resistance. There was no significant difference in the case of artesunate. In regression analysis, parasitaemia was an independent predictor of recrudescence/re-infection after mefloquine (p = 0.02). These data support the use of combination therapy such as artesunate plus mefloquine for falciparum malaria in a hyperendemic area of Viet Nam. Primarily because of their greater parasite densities, children should be given higher doses of mefloquine (e.g. 25 mg/kg).