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Controlling mosquitoes is vital for counteracting the rising number of mosquito-borne illnesses. Vector control requires the implementation of various measures; however, current methods lack complete effectiveness, and new control agents or substances are urgently needed. Therefore, this study developed a nonwoven fabric sheet coated with hydroxyapatite-binding silver/titanium dioxide compound (hydroxyapatite-binding silver/titanium dioxide sheet [HATS])and evaluated its effectiveness on all stages of laboratory Aedes aegypti (Linnaeus); Diptera: Culicidae and Anopheles dirus (Peyton & Harrison); Diptera: Culicidae. We reared larvae with HATS and control sheets and assessed their mortality, emergence, and hatching rates. The submersion rates of engorged female mosquitoes in submerged HATS and control sheets were also compared. The HATS strongly affected mosquito development, resulting in high mortality rates (mean ± SE) of 99.66 ± 0.58% (L1–L2) and 91.11 ± 9.20% (L3–L4) for Ae. aegypti and 100% of both stages for An. dirus. In contrast, mosquitoes raised in the control sheet showed relatively high survival rates of 92.33 ± 3.21% (L1–L2) and 95.67 ± 0.58% (L3–L4) for Ae. aegypti and 86.07 ± 3.53% (L1–L2) and 92.01 ± 8.67% (L3–L4) for An. dirus. Submersion of engorged females was found in the HATS oviposition cup, leading to a decreased number of eggs and a low hatching rate compared to that of the control. Overall, HATS may be a useful new control method for Ae. aegypti and An. dirus.

The malaria landscape in the Greater Mekong Subregion has experienced drastic changes with the ramp-up of the control efforts, revealing formidable challenges that slowed down the progress toward malaria elimination. Problems such as border malaria and cross-border malaria introduction, multidrug resistance in Plasmodium falciparum, the persistence of Plasmodium vivax, the asymptomatic parasite reservoirs, and insecticide resistance in primary vectors require integrated strategies tailored for individual nations in the region. In recognition of these challenges and the need for research, the Southeast Asian International Center of Excellence for Malaria Research has established a network of researchers and stakeholders and conducted basic and translational research to identify existing and emerging problems and develop new countermeasures. The installation of a comprehensive disease and vector surveillance system at sentinel sites in border areas with the implementation of passive/active case detection and cross-sectional surveys allowed timely detection and management of malaria cases, provided updated knowledge for effective vector control measures, and facilitated the efficacy studies of antimalarials. Incorporating sensitive molecular diagnosis to expose the significance of asymptomatic parasite reservoirs for sustaining transmission helped establish the necessary evidence to guide targeted control to eliminate residual transmission. In addition, this program has developed point-of-care diagnostics to monitor the quality of artemisinin combination therapies, delivering the needed information to the drug regulatory authorities to take measures against falsified and substandard antimalarials. To accelerate malaria elimination, this program has actively engaged with stakeholders of all levels, fostered vertical and horizontal collaborations, and enabled the effective dissemination of research findings.

In the course of malaria elimination in the Greater Mekong Subregion (GMS), malaria epidemiology has experienced drastic spatiotemporal changes with residual transmission concentrated along international borders and the rising predominance of Plasmodium vivax. The emergence of Plasmodium falciparum parasites resistant to artemisinin and partner drugs renders artemisinin-based combination therapies less effective while the potential spread of multidrug-resistant parasites elicits concern. Vector behavioral changes and insecticide resistance have reduced the effectiveness of core vector control measures. In recognition of these problems, the Southeast Asian International Center of Excellence for Malaria Research (ICEMR) has been conducting multidisciplinary research to determine how human migration, antimalarial drug resistance, vector behavior, and insecticide resistance sustain malaria transmission at international borders. These efforts allow us to comprehensively understand the ecology of border malaria transmission and develop population genomics tools to identify and track parasite introduction. In addition to employing in vivo, in vitro, and molecular approaches to monitor the emergence and spread of drug-resistant parasites, we also use genomic and genetic methods to reveal novel mechanisms of antimalarial drug resistance of parasites. We also use omics and population genetics approaches to study insecticide resistance in malaria vectors and identify changes in mosquito community structure, vectorial potential, and seasonal dynamics. Collectively, the scientific findings from the ICEMR research activities offer a systematic view of the factors sustaining residual malaria transmission and identify potential solutions to these problems to accelerate malaria elimination in the GMS.

Ivermectin mass drug administration to humans or livestock is a potential vector control tool for malaria elimination. The mosquito-lethal effect of ivermectin in clinical trials exceeds that predicted from in vitro laboratory experiments, suggesting that ivermectin metabolites have mosquito-lethal effect. The three primary ivermectin metabolites in humans (i.e., M1 (3″-O-demethyl ivermectin), M3 (4-hydroxymethyl ivermectin), and M6 (3″-O-demethyl, 4-hydroxymethyl ivermectin) were obtained by chemical synthesis or bacterial modification/metabolism. Ivermectin and its metabolites were mixed in human blood at various concentrations, blood-fed to Anopheles dirus and Anopheles minimus mosquitoes, and mortality was observed daily for fourteen days. Ivermectin and metabolite concentrations were quantified by liquid chromatography linked with tandem mass spectrometry to confirm the concentrations in the blood matrix. Results revealed that neither the LC50 nor LC90 values differed between ivermectin and its major metabolites for An. dirus or An. minimus., Additionally, there was no substantial differences in the time to median mosquito mortality when comparing ivermectin and its metabolites, demonstrating an equal rate of mosquito killing between the compounds evaluated. These results demonstrate that ivermectin metabolites have a mosquito-lethal effect equal to the parent compound, contributing to Anopheles mortality after treatment of humans.

The occurrence and spread of insecticide resistance has had a negative effect on the efficacy of insecticide-based tools and is distributed worldwide, including the Greater Mekong Subregion (GMS). This study aims to determine the insecticide susceptibility of malaria and dengue vectors in malaria and dengue hotspots on the Thai-Myanmar border. Mosquito larvae and pupae were obtained from water sources from December 2019 to April 2020 in Tha Song Yang District, Tak province, western Thailand. WHO bioassay susceptibility tests were conducted with three classes of insecticides to evaluate the knockdown and mortality rates of

Mosquito repellents reduce human-vector contact of vector-borne diseases. We compared the repellent activity of 10 undiluted essential oils (anise, basil, bergamot, coriander, patchouli, peppermint, petitgrain, rosemary, sage and vetiver) against

Anopheles minimus Theobald 1901 and An. harrisoni Harbach & Manguin 2007 belong to the same species complex. They are morphologically similar and can exist in sympatry but have blood host preferences. The most accurate method for their identification is based on molecular techniques. Here, we measure the level of interspecific discrimination by geometric morphometry. Sixty-seven An. minimus and 22 An. harrisoni specimens were selected based on their morphological integrity and confirmed by identification polymerase chain reaction of internal transcribed spacer 2. These samples were used as reference data allowing for a morphometric identification based on geometric shape. Despite size overlap between the two species, there was a significant shape divergence allowing for differentiation of An. minimus and An. harrisoni with 90% accuracy. An intraspecific study of An. minimus showed a summer period associated to the reducing of wing size, which did not influence the shape-based differentiation of An. harrisoni. Wing venation geometry can be used to distinguish between these cryptic species mainly based on shaped divergence. This study suggests that geometric morphometrics represent a convenient low-cost method to complement morphological identification, especially concerning damaged specimens, i.e., insects having accidentally lost the anatomical features allowing a reliable morphological identification.

All members of the ant genus

Diseases transmitted by mosquito vectors such as malaria, dengue, and Zika virus are amongst the largest healthcare concerns across the globe today. To tackle such life-threatening diseases, it is vital to evaluate the risk of transmission. Of critical importance in this task is the estimation of vector species populations in an area of interest. Traditional approaches to estimating vector populations involve physically collecting vector samples in traps and manually classifying species, which is highly labor intensive. A promising alternative approach is to classify mosquito species based on the audio signal from their wingbeats. Various traditional machine learning and deep learning models have been developed for such automated acoustic mosquito species classification. But they require data preprocessing and significant computation, limiting their suitability to be deployed on low-cost sensor devices. This paper presents two lightweight deep learning models for mosquito species and sex classification from wingbeat audio signals which are suitable to be deployed on small IoT sensor devices. One model is a 1D CNN and the other combines the 1D CNN with an LSTM model. The models operate directly on a low-sample-rate raw audio signal and thus require no signal preprocessing. Both models achieve a classification accuracy of over 93% on a dataset of recordings of males and females of five species. In addition, we explore the relation between model size and classification accuracy. Through model tuning, we are able to reduce the sizes of both models by approx. 60% while losing only 3% in classification accuracy.

Members of the Maculatus group are important malaria vectors in the border regions of Thailand. However, the role of each species in malaria transmission remains unclear because of their highly similar morphologies, making them difficult to be differentiated. Whereas An. pseudowillmori may be identified by the color pattern of some scales on abdomen and wings, the distinction between An. maculatus and An. sawadwongporni relies on the wings only. Scales are labile structures, as they may be accidentally removed during capture and transportation to the laboratory. To discriminate among the species of this complex, we tested the suitability of geometric techniques. Shape variables were used as input for discriminant analyses and validated reclassification. Both landmark- and outline-based geometric techniques disclosed significant differences between the three species. For the delicate An. maculatus - An. sawadwongporni distinction, the outline-based approach appeared as the most promising, with validated reclassification scores reaching 93%, as compared to 77% obtained by landmark data. For An. pseudowillmori, reclassification scores were 100% and 94%, respectively. Geometric morphometrics may provide an alternative and useful complement for discriminating members of the Maculatus group.

All members of the ant genus Odontomachus Latreille, 1804 are venomous ants. Four species in this genus have been identified from Thailand: Odontomachus latidens Mayr, 1867; O. monticola Emery, 1892; O. rixosus Smith, 1757; and O. simillimus Smith, 1758. The three latter species are available and have been used for an outline morphometric study. They display similar morphology, which makes their distinction very difficult except for highly qualified individuals. A total of 80 worker specimens were studied, exploring the contour shapes of their head and pronotum as possible taxonomic characters. The size of each body part was estimated determining the contour perimeter, the values for which were largely overlapping between O. rixosus and O. simillimus; most O. monticola specimens exhibited a significantly larger size. In contrast to the size, each contour shape of the head or pronotum established O. rixosus as the most distinct species. An exploratory data analysis disclosed the higher taxonomic signal of the head contour relative to the pronotum one. The scores obtained for validated reclassification were much better for the head (99%) than for the pronotum (82%). This study supports outline morphometrics of the head as a promising approach to contribute to the morphological identification of ant species, at least for monomorphic workers.

Crude extracts and essential oils of A. conyzoides were tested with larva and adult stages of Ae. aegypti mosquitoes to determine their insecticidal properties. The crude extracts and essential oils came from three varieties of A. conyzoides (with white flowers, purple flowers, or white-purple flowers) and from two places on each plant (leaves and flowers), giving six types overall: leaf-white (LW); leaf-purple (LP); leaf white-purple (LW-P); flower-white (FW); flower-purple (FP); and flower white-purple (FW-P). Chemical constituents and components of the essential oils were identified using gas chromatography-mass spectrometry (GC-MS). Electron microscopic and histopathological studies were performed to determine the toxicological effects on mosquitoes in terms of morphological alterations. The six types of crude extracts exhibited no activity against individuals in the larval stages. However, six types of essential oils were effective against adult Ae. aegypti females. The mortality of adult Ae. aegypti females was higher from leaf extracts, particularly LP (median lethal dose, LD50 = 0.84%). The number of chemical constituents identified by GC-MS was high in flowers, especially W-P. Precocene I was the most abundant chemical component among the five types of essential oils, except in LP, in which precocene II was the most abundant. Histopathological alterations in adult Ae. aegypti females included compound eye degeneration, muscular damage with cellular infiltration, gut epithelial degeneration and necrosis, pyknotic nuclei in the malpighian epithelium and ovarian cell degeneration. FW and FP plant types exhibited the highest severity of histopathological alterations in mosquitoes compared with other plants, probably owing to the presence of monoterpene compounds in their tissues. The present study demonstrated LP plant extracts from A. conyzoides could be effective adulticides against adult Ae. aegypti. As natural products are biodegradable and exhibit low toxicity to mammalian and non-target organisms, they are suitable candidates for use in vector control programmes.

Additional file 1.

Background A mixed methods study was conducted to look at the magnitude of residual malaria transmission (RMT) and factors contributing to low (

Geographic populations of the two main sandflies genera present in Thailand were studied for species and population identification. Size and shape of Phlebotomus stantoni and Sergentomyia hodgsoni from different island and mainland locations were examined by landmark-based geometric morphometrics. Intraspecific and interspecific wing comparison was carried out based on 12 anatomical landmarks. The wing centroid size of P. stantoni was generally larger than that of S. hodgsoni. Within both species, wings from the continent were significantly larger than those from island populations. Size variation could be significant between geographic locations, but could also overlap between genera. The wing venation geometry showed non-overlapping differences between two species. The within-species variation of geometric shape between different geographical locations was highly significant, but it could not interfere with the interspecies difference. The lack of species overlapping in shape, and the high discrimination between geographic populations, make geometric shape a promising character for future taxonomic and epidemiological studies.

Background Giardia duodenalis causes giardiasis in humans, particularly in developing countries. Despite the availability of treatments, resistance to some of the commercial anti-Giardia drugs has been reported in addition to their harmful side effects. Therefore, novel treatments for giardiasis are required. In this study, we aimed to assess the in vitro activity of crude extracts of Ageratum conyzoides against G. duodenalis trophozoites. Methods Plants were classified into three groups based on their flower colors: white (W), purple (P), and white–purple (W–P). Plants were separately cut into leaf (L) and flower (F) parts. Changes in internal organelle morphology of trophozoites following exposure to crude extracts were assessed using transmission electron microscopy (TEM). In subsequent experiments, efficacy of the most active essential oils from crude extracts [half maximal inhibitory concentrations (IC50) ≤ 100 μg/mL] against G. duodenalis trophozoites was tested. In vitro anti-Giardia assays using essential oils were performed in the same way as those performed using crude extracts. Results LW–P and FP extracts showed high activity (IC50 ≤ 100 μg/mL) against G. duodenalis trophozoites, with IC50 ± SD values of 45.67 ± 0.51 and 96.00 ± 0.46 μg/mL, respectively. In subsequent experiments, IC50 ± SD values of LW–P and FP essential oils were 35.00 ± 0.50 and 89.33 ± 0.41 μg/mL, respectively. TEM revealed the degeneration of flagella and ventral discs of G. duodenalis trophozoites following exposure to crude extracts. Conclusion Crude LW–P and FP extracts of A. conyzoides showed the highest activity against G. duodenalis. Exposure to crude extract induced changes in the flagella and ventral discs of G. duodenalis trophozoites, which play important roles in attachment to the surface of mucosal cells. Our results suggest that the tested extracts warrant further research in terms of their efficacy and safety as giardiasis treatment.

Additional file 3: Table S3. Calculation of indoor and outdoor exposure risk for users and non-users of LLINs in Pha Man.

Additional file 2: Table S2. Utilization of bed nets the previous night by demographic factor among the surveyed adult population.

Additional file 1: Table S1. Demographics of the surveyed adult (aged 18Â years and over) population in the three study villages of Tha Song Yang district.

There was recently an outbreak of malaria in Ubon Ratchathani Province, northeastern Thailand. In the absence of information on malaria vector transmission dynamics, this study aimed to identify the anopheline vectors and their role in malaria transmission. Adult female Anopheles mosquitoes were collected monthly by human-landing catch in Na Chaluai District of Ubon Ratchathani Province during January 2014−December 2015. Field-captured mosquitoes were identified to species using morphology-based keys and molecular assays (allele-specific polymerase chain reaction, AS-PCR), and analysed for the presence of Plasmodium falciparum and Plasmodium vivax using an enzyme-linked immunosorbent assay (ELISA) for the detection of circumsporozoite proteins (CSP). A total of 1,229 Anopheles females belonging to 13 species were collected. Four anopheline taxa were most abundant: Members of the Anopheles barbirostris complex, comprising 38% of the specimens, species of the Anopheles hyrcanus group (18%), Anopheles nivipes (17%) and Anopheles philippinensis (12%). The other nine species comprised 15% of the collections. Plasmodium infections were detected in two of 668 pooled samples of heads/thoraces, Anopheles dirus (1/29) and An. philippinensis (1/97). The An. dirus pool had a mixed infection of P. vivax-210 and P. vivax-247, whereas the An. philippinensis pool was positive only for the latter protein variant. Both positive ELISA samples were confirmed by nested PCR. This study is the first to incriminate An. dirus and An. philippinensis as natural malaria vectors in the area where the outbreak occurred. This information can assist in designing and implementing a more effective malaria control programme in the province.

Background Cross-border malaria transmission is an important problem for national malaria control programmes. The epidemiology of cross-border malaria is further complicated in areas where Plasmodium falciparum and Plasmodium vivax are both endemic. By combining passive case detection data with entomological data, a transmission scenario on the northwestern Thai–Myanmar border where P. falciparum is likely driven by importation was described, whereas P. vivax is also locally transmitted. This study highlights the differences in the level of control required to eliminate P. falciparum and P. vivax from the same region. Methods Malaria case data were collected from malaria clinics in Suan Oi village, Tak Province, Thailand between 2011 and 2014. Infections were diagnosed by light microscopy. Demographic data, including migrant status, were correlated with concomitantly collected entomology data from 1330 mosquito trap nights using logistic regression. Malaria infection in the captured mosquitoes was detected by ELISA. Results Recent migrants were almost four times more likely to be infected with P. falciparum compared with Thai patients (OR 3.84, p

Comptes Rendus Biologies - In Press.Proof corrected by the author Available online since jeudi 9 fevrier 2017

Understanding specific gene regulation during responses to malaria infection is key to dissecting the mosquito defense mechanisms and host/parasite interactions. A full-length serine protease cDNA was isolated from the fat body of female Anopheles dirus, a major malaria vector in Thailand. The predicted amino acid sequence of SERF4 identifies it as a member of the serine protease family containing a single trypsin-like protease domain. Digestive trypsins function in the female mosquito midgut and are inducible in two phases in this tissue upon blood intake. However, the gene was highly upregulated in the midgut at day 3 postinfection by Plasmodium falciparum. In situ hybridization confirmed that SERF4 transcripts were located in the midgut epithelial cells rather than hemocytes or other tissues associated with the midgut. SERF4 was also strongly downregulated in the whole insects at day 16 after infection in comparison with the blood-fed control. Changes in the expression of the SERF4 gene in response to infection with this human malaria parasite suggest a role in parasite-specific innate immunity.

Aedes aegypti (Diptera: Culicidae) (L.), Ae. albopictus (Skuse), and Ae. scutellaris (Walker) are important mosquito vectors of dengue and chikungunya viruses. They are morphologically similar and sympatric in some parts of their distribution; therefore, there is a risk of incorrect morphological identification. Any confusion could have a negative impact on epidemiological studies or control strategies. Therefore, we explored two modern tools to supplement current morphological identification: DNA barcoding and geometric morphometric analyses. Field larvae were reared to adults and carefully classified based on morphological traits. The genetic analysis was based on the 658 bp each of 30COI sequences. Some Culex spp., Mansonia bonneae, were included as outgroups, and inclusion of a few other Aedes spp. facilitated phylogenetic inference of the relationship between Ae. albopictus and Ae. scutellaris. The two species were separated by an average interspecific divergence of 0.123 (0.119-0.127). Morphometric examination included landmark- (392 specimens) and outline-based (317 specimens) techniques. The shape of the wing showed different discriminating power based on sex and digitizing technique. This is the first time that Ae. scutellaris and Ae. albopictus have been compared using these two techniques. We confirm that these morphologically close species are valid, and that geometric morphometrics can considerably increase the reliability of morphological identification.

Approximately 1.4 billion people are at some risk of malaria infection in the South East Asia region, with 352 million at high risk. The most common and effective malaria vector control strategies currently in use are based on insecticides: indoor residual spraying (IRS) and long-lasting insecticide-treated nets. However, these interventions were designed for indoor-biting vectors, and hence are not necessarily effective in all regions. Although there are many nongenetic tools for vector control, a major limitation is the research-based evaluation of efficiency of those tools in many areas. Another limitation for research of malaria vectors in Asia is due to the high complexity of many vector species and the difficulty to maintain most of the important vectors in the laboratory, as it is very labor intensive. Here, we review the ecological and behavioral characteristics of only the most important anthropophilic vectors within this region: Anopheles culicifacies , Anopoheles fluviatilis , and Anopheles stephensi across the Indian subcontinent; and Anopheles dirus and Anopheles minimus within Southeast Asia. The challenges in applying genetic control methods to malaria vectors in Asia will be similar to the other nongenetic tools that have been used to control vectors in this region: species complexity, multiple vectors, and potential vectors in each endemic area and only a few species that can be successfully maintained in the laboratory.

Stomoxyini flies (Diptera: Muscidae) include species of parasitic flies of medical and veterinary importance. The adult flies feed on the blood of mammals and may transmit several parasites and pathogens. We conducted an entomological survey of Stomoxyini flies from different sites in Thailand. Stomoxyini flies were collected at four major types of sites: zoos, livestock farms, wildlife conservation areas and a national park using vavoua traps between November 2010 and April 2011. A total of 3,314 Stomoxyini flies belonging to the genera Stomoxys, Haematobosca, Haematostoma and Haematobia were collected. Eight species were identified: S. calcitrans (46.6%), S. uruma (26.8%), S. pulla (4.3%), S. indicus (0.7%), S. sitiens (0.1%), H. sanguinolenta (11.2 %), H. austeni (0.5%) and H. irritans exigua (9.8%). The diversity of Stomoxyini flies in the livestock farms was higher than the other sites. Altitude correlated with the number of flies. This study provides information that may be useful for Stomoxyini flies control.

Understanding specific gene regulation during responses to malaria infection is key to dissecting the mosquito defense mechanisms and host/parasite interactions. A full-length serine protease cDNA was isolated from the fat body of female Anopheles dirus, a major malaria vector in Thailand. The predicted amino acid sequence of SERF4 identifies it as a member of the serine protease family containing a single trypsin-like protease domain. Digestive trypsins function in the female mosquito midgut and are inducible in two phases in this tissue upon blood intake. However, the gene was highly upregulated in the midgut at day 3 postinfection by Plasmodium falciparum. In situ hybridization confirmed that SERF4 transcripts were located in the midgut epithelial cells rather than hemocytes or other tissues associated with the midgut. SERF4 was also strongly downregulated in the whole insects at day 16 after infection in comparison with the blood-fed control. Changes in the expression of the SERF4 gene in response to infection with this human malaria parasite suggest a role in parasite-specific innate immunity.

Three serine protease cDNA clones were isolated from Anopheles dirus, a major vector of malaria in Southeast Asia. Transcript abundance was examined following infection by Plasmodium falciparum by RT-PCR analysis. SerF3 exhibited increased transcript abundance in the whole body at 10 days post-infection with P. falciparum. All three genes are candidates for further investigation to determine their roles in mosquito immune responses.

Trap-nights and mosquitoes caught per trap and year in each of the seven villages for the most abundant Anopheles mosquito species. (DOCX 25 kb)

Additional file 3: Table S1. Patient characteristics for the M2 migrant population*. Table S2. Patient characteristics for the M1 migrant population*. Table S3. Patient characteristics for the Thai national population. Table S4. Occupation distribution among the different population strata.

Additional file 1. Histogram of number of days spent in trips.

All mosquitoes captured in the present study, their blood-fed status, percentage of females and gravidity status. (DOCX 18 kb)

Additional file 2. Counts of people with SRM by gender and village.

Trap-nights and mosquitoes caught per trap and year in each of the seven villages for the most abundant mosquito genera and species other than Anopheles. (DOCX 19 kb)

Additional file 2. Counts of people with SRM by gender and village.

Trap-nights and mosquitoes caught per trap and year in each of the seven villages for the most abundant mosquito genera and species other than Anopheles. (DOCX 19 kb)

Additional file 1. Histogram of number of days spent in trips.

Trap-nights and mosquitoes caught per trap and year in each of the seven villages for the most abundant Anopheles mosquito species. (DOCX 25 kb)

Additional file 3: Table S1. Patient characteristics for the M2 migrant population*. Table S2. Patient characteristics for the M1 migrant population*. Table S3. Patient characteristics for the Thai national population. Table S4. Occupation distribution among the different population strata.

All mosquitoes captured in the present study, their blood-fed status, percentage of females and gravidity status. (DOCX 18 kb)