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Insects geographically separated into island and mainland populations often exhibit phenotypic variations, a phenomenon known as insular conditions. These conditions can lead to rapid evolutionary changes that affect the morphological characteristics of mosquito vectors. Nevertheless, studies that specifically examine phenotype differences between island and mainland mosquito populations have been limited. In this study, wing variation in size and shape was investigated using the geometric morphometric (GM) technique in two dominant mosquito vectors, Aedes albopictus and Armigeres subalbatus, in the Ranong and Trat archipelagos of Thailand. Significant differences in average wing centroid size (CS) were found in 6 out of 15 population pairs for Ae. albopictus (p < 0.05) and in 5 population pairs for Ar. subalbatus (p < 0.05). After removing the allometric effect, canonical variate analyses (CVA) based on wing shape analysis revealed overlap across all populations for both Ae. albopictus and Ar. subalbatus. However, the statistical analysis indicated that Ar. subalbatus exhibited wing shape differences across all populations (p < 0.05), and most Ae. albopictus populations also displayed distinct wing shapes (p < 0.05), except for the populations from Chang Island and the mainland of Ranong, which showed no significant differences (p > 0.05). These findings enhance our understanding of mosquito adaptability in island regions and provide valuable data for the surveillance and monitoring of vector evolution. [ABSTRACT FROM AUTHOR]

Simple Summary: Stomoxys flies (Diptera: Muscidae) are bloodsucking insects that feed on both animals and humans. They are a major vector of a variety of animal pathogens, including agents of trypanosomosis, anaplasmosis, bovine leucosis, African horse sickness, lumpy skin disease, etc. This study investigated the effectiveness of landmark-based geometric morphometrics of wings to discriminate among three morphologically similar species of Stomoxys in Thailand: Stomoxys bengalensis, Stomoxys calcitrans, and Stomoxys sitiens. Our study reveals that geometric morphometrics can accurately discriminate the three species of Stomoxys based on wing shape. Therefore, wing geometric morphometrics can be used in addition to traditional morphological methods to identify these flies as species. The flies of the genus Stomoxys Geoffroy, 1762 (Diptera: Muscidae), are regarded as pests of veterinary and medical importance. In Thailand, Stomoxys calcitrans (Linnaeus, 1758) is the most abundant species and is widely distributed throughout the country. This Stomoxys species can coexist with two other morphologically similar species: Stomoxys bengalensis Picard, 1908, and Stomoxys sitiens Rondani, 1873. Hence, discriminating using morphological characteristics is difficult, especially if the specimen is damaged or loses its diagnostic characteristics. In this study, we evaluated the effectiveness of the landmark-based geometric morphometric (GM) approach to discriminate among the three Stomoxys spp.: S. bengalensis, S. calcitrans, and S. sitiens. Left-wing images of S. bengalensis (n = 120), S. calcitrans (n = 150), and S. sitiens (n = 155) were used for the GM analyses. The results of the wing shape analyses revealed that the GM approach was highly effective for discriminating three Stomoxys, with high accuracy scores ranging from 93.75% to 100%. This study adds to the evidence that landmark-based GM is an excellent alternative approach for discriminating Stomoxys species. [ABSTRACT FROM AUTHOR]

Simple Summary: There are four species of Lutzia mosquitoes in Thailand, including Lt. chiangmaiensis, Lt. fuscana, Lt. halifaxii, and Lt. vorax. However, it is difficult to identify damaged specimens when some of their morphological character is missing. Thus, we evaluated the efficacy of the geometric morphometric (GM) approach and DNA barcoding for the identification of four Lutzia species. Our results showed that DNA barcoding is poorly effective in identifying Lt. fuscana and Lt. halifaxii based on their low interspecific genetic differences. On the other hand, the GM approach based on wing shape analyses successfully identified the four Lutzia species in Thailand. There are four species of Lutzia mosquitoes in Thailand, including Lutzia chiangmaiensis, Lt. fuscana, Lt. halifaxii, and Lt. vorax. The accurate species identification of adult Lutzia mosquitoes based on morphological features requires many body parts, including the abdominal terga and wing. However, species identification is difficult in the case of damaged specimens when some of their morphological character is missing due to transit or gathering in the field. Thus, we evaluated the efficacy of the landmark-based geometric morphometric (GM) approach for the discrimination of Lutzia species in Thailand. In addition, DNA barcoding was also used in parallel with the GM approach to identify the species. Larvae of Lutzia were collected, raised into adults, and identified based on their morphological characteristics. The validated reclassification test results clearly demonstrated that wing shape resulted in a high level of success in identification (correct identifications ranged from 92.50% to 100%); however, based on the DNA barcoding analyses, our results showed that it was poorly effective in identifying Lt. fuscana and Lt. halifaxii based on an overlap between the intraspecific and interspecific divergence. Moreover, our survey results provide updates on the distribution of Lt. chiangmaiensis and Lt. vorax in Thailand. This research will help medical entomologists more efficiently identify mosquitoes in the genus Lutzia, resulting in more effective mosquito control and surveillance. [ABSTRACT FROM AUTHOR]

The correct identification of mosquito species is important for effective mosquito vector control. However, the standard morphological identification of mosquito species based on the available keys is not easy with specimens in the field due to missing or damaged morphological features during mosquito collections, often leading to the misidentification of morphologically indistinguishable. To resolve this problem, we collected mosquito species across Thailand to gather genetic information, and evaluated the DNA barcoding efficacy for mosquito species identification in Thailand. A total of 310 mosquito samples, representing 73 mosquito species, were amplified using mitochondrial cytochrome c oxidase subunit I (COI) primers. The average maximum intraspecific genetic variation of the 73 mosquito species was 1% ranged from 0–5.7%. While, average minimum interspecific genetic variation (the distance to the nearest neighbour) of the 73 mosquito species was 7% ranged from 0.3–12.9%. The identification of success rates based on the "Best Match," "Best Close Match," and "All Species Barcodes" methods were 97.7%, 91.6%, and 81%, respectively. Phylogenetic analyses of Anopheles COI sequences demonstrated a clear separation between almost all species (except for those between An. baimaii and An. dirus), with high bootstrap support values (97%–99%). Furthermore, phylogenetic analyses revealed potential sibling species of An. annularis, An. tessellatus, and An. subpictus in Thailand. Our results indicated that DNA barcoding is an effective molecular approach for the accurate identification of mosquitoes in Thailand. [ABSTRACT FROM AUTHOR]

Culex gelidus (Diptera: Culicidae), an important vector of the Japanese encephalitis virus (JEV), contributes to human viral encephalitis in many Asian countries, including Thailand. This study represents the first investigation of the demographic patterns of Cx. gelidus populations in Thailand using cytochrome c oxidase subunit I (COI) gene analysis and wing geometric morphometrics (GM). Mosquitoes were collected from 10 provinces across six regions of Thailand in 2022. Analysis of the COI sequences (n = 182) indicated high haplotype diversity (0.882) and low nucleotide diversity (0.006), with 72 haplotypes identified. The haplotype network demonstrated no profound splits among the geographic populations. Neutral tests, including Tajima's D and Fu's Fs , displayed negative values, with a significant result observed for Fu's Fs (−33.048, p < 0.05). The mismatch distribution analysis indicated that the population does not statistically deviate from a model of sudden population expansion (SSD = 0.010, p > 0.05; Rg = 0.022, p > 0.05). The estimations suggest that the Cx. gelidus population in Thailand began its expansion approximately between 459,243 and 707,011 years ago. The Mantel test showed no significant relationship between genetic and geographic distances (r = 0.048, p > 0.05). Significant phenotypic differences (based on wing shape) were observed among most populations. Additionally, in this study, we found no significant relationships between phenotypic and genetic distances (r = 0.250, p > 0.05). Understanding the genetic and morphological dynamics of Cx. gelidus is vital for developing targeted surveillance and vector control measures. This knowledge will also help to predict how future environmental changes might affect these populations, thereby informing long-term vector management strategies. [ABSTRACT FROM AUTHOR]

Simple Summary: The stable fly, Stomoxys calcitrans (Diptera: Muscidae), is the predominant Stomoxys spp. in Thailand and is considered a pest for livestock, pets, wildlife, and occasionally humans. This study investigated the phenotypic variation in the wing size and shape of S. calcitrans populations from different geographical regions in Thailand using landmark-based geometric morphometric analysis. Results showed strong spatial variation in wing shape among S. calcitrans populations and thus suggested the existence of phenotypic plasticity in this fly. Stomoxys calcitrans (Linnaeus, 1758) (Diptera: Muscidae) is a cosmopolitan hematophagous ectoparasite of veterinary and medical importance. It is an important mechanical vector of several animal pathogens and can cause significant economic losses. However, the morphological variation of this species remains unknown. This study aimed to investigate the phenotypic variation in the wing size and shape of S. calcitrans populations in Thailand based on a landmark-based geometric morphometric approach. Specimens were collected from five populations in five geographical regions in Thailand. A total of 490 left wings of S. calcitrans (245 female and 245 male individuals) were used for geometric morphometric analysis. Wing size differences were detected between some populations of S. calcitrans, whereas wing shape differences were found among populations. Therefore, the phenotypic variation in S. calcitrans populations indicated that these populations are adaptive responses to local environmental pressures, suggesting the presence of phenotypic plasticity in this species. [ABSTRACT FROM AUTHOR]

Objective: The objective of this study was to determine the comparative larvicidal efficacy of commercial ylang-ylang (Cananga odorata) essential oils from India and Thailand against the Aedes aegypti mosquito to be used as a guideline for Aedes mosquito control. Materials and Methods: The bioassay for the larvicidal activity of commercial ylang-ylang essential oils in this experiment was modified from the World Health Organization standard protocols. The concentration ranges at 0.025, 0.050, 0.075, 0.100, 0.125, and 0.150 ppm in each treatment were used for testing, and four replicates were used per concentration. The larval mortality was observed and recorded 24- and 48-h after exposure. Results: The results of this study clearly revealed that commercial ylang-ylang essential oils from India and Thailand were highly toxic to the larvae of the dengue vector Ae. aegypti, and Indian ylang-ylang had an LC50 value of 0.064 ppm, whereas Thai ylang-ylang had an LC50 value of 0.042 ppm after 24-h exposure. Conclusion: This study revealed the efficacy of commercial Indian and Thai ylang-ylang essential oils as natural vector control for the larval stage of the dengue vector Ae. aegypti. Usually, natural larvicide products are not commonly found in the market due to complex production processes. The results of this research support the use of commercial essential oils to aid in further control of Aedes mosquito larvae populations in the community. [ABSTRACT FROM AUTHOR]

Culex sitiens Wiedemann (Diptera, Culicidae) is a mosquito vector that is found in coastal areas. Effective control of mosquitoes requires knowledge of the biology, ecology, and behavior of the vector as well as of various other aspects, including its morphology. Currently, variations in the wing size and shape of coastal Cx. sitiens have not been described. Here, morphological changes were studied in the wings of Cx. sitiens from a coastal area of Samut Songkhram Province, Thailand. Samples were collected at night (6:00 pm–6:00 am) during single weeks of September in the years 2015–2017 using Center for Disease Control light traps with dry ice as bait. Eighteen landmarks of each individual were selected and digitized for landmark-based geometric morphometric analyses. Wing size variability was estimated using the isometric estimator of centroid size. Wing-shape variables were computed as Procrustes superimposition with residual coordinates of the 18 landmarks following a Generalized Procrustes Analysis and the principal components of residual coordinates. Degrees of wing-shape dissimilarity among individuals were analyzed using discriminant analysis or canonical variate analysis, which was illustrated in a discriminant space of canonical variables. Differences in wing size and shape among populations were calculated using nonparametric permutations based on 1000 runs with Bonferroni correction tests at a p-value of <0.05. The wing sizes and shapes of the mosquitoes differed significantly between observation years in all population groups, as indicated by nonparametric tests (1000 runs) with the Bonferroni correction. Differing rainfall between observation years was related to morphological changes in mosquito populations, presumably reflecting environmental adaptation. Differences in the wing morphology of Cx. sitiens between annual populations reflect adaptation to environmental variables such as rainfall and may affect the potential to act as insect vectors of human disease. These observations may facilitate the development of tools for managing mosquito-borne disease. [ABSTRACT FROM AUTHOR]

Objective: The aim of this research is to study the effectiveness of mosquito magnet (MM) for reducing mosquitoes (Diptera) populations in coastal areas. Materials and methods: The study sites are in the coastal area of Samut Songkhram province, Thailand, which is divided into two locations; one that is 2 km and another that is 4 km in distance from the sea. We used the Mosquito Magnet® Independence (MMI) trap for effective field testing in Samut Songkhram Province, Thailand. Traps were placed 100 m away from the house (one trap per location) and mosquitoes were collected at night from 6 PM to 6 AM during September and October 2017 (30 days). Results: A total of 2,561 adult mosquitoes, including Anopheles epiroticus Linton & Harbach, Culex quinquefasciatus Say, Cx. sitiens Wiedmann, and Cx. gelidus Theobald were collected by MMI. At a 2-km distance from the sea were captured more mosquitoes per night more than at a 4-km distance (63.63 ± 42.30 vs. 21.70 ± 12.42). The comparison of effectiveness of MMI in two locations of the coastal area was shown to have a statistically significant difference (p < 0.05) and analysis of the correlation between the number of mosquitoes caught in coastal areas, including at a 2- and 4-km distance from the sea, accounting for weather factors, we found that the effectiveness of MMI was not correlated with weather (p > 0.05). Conclusion: Overall, this study demonstrated that MM can be used to control mosquitoes in coastal areas with high efficiency, especially 2 km away from the sea. [ABSTRACT FROM AUTHOR]

Stomoxys flies (Diptera: Muscidae) are hematophagous ectoparasites of medical and veterinary importance. In this study, three Stomoxys species, i.e. S. bengalensis , S. calcitrans , and S. sitiens , were collected from three provinces in Central Thailand with the aim of estimating the genetic divergence between species, for species identification, as well as within species, for a genetic diversity study based on the cytochrome c oxidase subunit I (COI) gene. Our results showed that the average intraspecific genetic divergences of Stomoxys flies ranged from 0.11% in S. sitiens to 0.98% in S. calcitrans , whereas the average interspecific genetic divergences ranged from 5.24% between S. sitiens and S. bengalensis to 6.69% between S. calcitrans and S. bengalensis. In addition, there was no overlap between the intraspecific and interspecific genetic divergences. The COI sequence analysis revealed a high haplotype diversity and low nucleotide diversity, reflecting a rapid population expansion after past bottlenecks. Moreover, there was no significant difference (P > 0.05) in the pairwise population differentiation (Fst) among Stomoxys flies in Central Thailand, because of the lack of natural barriers, thus allowing genetic exchange between them. The monitoring of the haplotype network revealed that two lineages of S. calcitrans in Central Thailand were distributed in all study areas, including the Nakhon Pathom, Pathum Thani, and Saraburi Provinces. These findings may improve our understanding of the genetic patterns of these three Stomoxys flies, as well as the underlying biological mechanisms, which is knowledge that can be used for further effective control of these flies. [Display omitted] • This study was the first to report COI barcode sequences for species identification in Stomoxys bengalensis. • Stomoxys bengalensis , Stomoxys calcitrans , and Stomoxys sitiens COI sequences indicated a rapid population expansion in Central Thailand after historical bottlenecks. • The monitoring of the haplotype network revealed the existence of two lineages of Stomoxys calcitrans in Central Thailand. [ABSTRACT FROM AUTHOR]

Simple Summary: Tabanus spp. (Diptera: Tabanidae) are blood-sucking parasites of animals and humans. The accurate identification of these flies is very important for determining the vector species involved in disease transmission and for planning effective vector control and management strategies. We explored the effectiveness of landmark-based geometrics at distinguishing and identifying morphologically similar species of Tabanus (T. megalops, T. rubidus, and T. striatus) in Thailand. Our study reveals that geometric morphometrics is effective at distinguishing between the three species of Tabanus. Furthermore, our study material can be used as reference material for species identification. Tabanus spp., also known as horse flies (Diptera: Tabanidae), are important vectors of several animal pathogens. Adult females of Tabanus megalops and Tabanus striatus, which are members of the T. striatus complex, are morphologically similar and hence difficult to distinguish using morphological characteristics. In addition, molecular identification by DNA barcoding is also unable to distinguish these species. These two species can occur sympatrically with Tabanus rubidus, which is morphologically similar to T. megalops and T. striatus. Wing geometric morphometrics has been widely used in various insects to distinguish morphologically similar species. This study explored the effectiveness of landmark-based geometrics at distinguishing and identifying T. megalops, T. rubidus, and T. striatus in Thailand. Specimens were collected from different geographical regions of Thailand, and only unambiguously identified specimens were used for geometric morphometric analyses. Left wings of females of T. megalops (n = 160), T. rubidus (n = 165), and T. striatus (n = 85) were photographed, and 22 wing landmarks were used for the analysis. Wing shape was able to distinguish among species with high accuracy scores, ranging from 94.38% to 99.39%. We showed that morphologically very close species of Tabanus can be reliably distinguished by the geometry of their wing venation, and we showed how our experimental material could be used as a reference to tentatively identify new field collected specimens. [ABSTRACT FROM AUTHOR]

Background: Anopheles sawadwongporni Rattanarithikul & Green, Anopheles maculatus Theobald and Anopheles pseudowillmori (Theobald) of the Anopheles maculatus group (Diptera: Culicidae) are recognized as potential malaria vectors in many countries from the Indian subcontinent through Southeast Asia to Taiwan. A number of malaria vectors in malaria hotspot areas along the Thai-Myanmar border belong to this complex. However, the species distribution and dynamic trends remain understudied in this malaria endemic region. Methods: Mosquitoes of the Maculatus group were collected using CDC light traps every other week from four villages in Tha Song Yang District, Tak Province, Thailand from January to December 2015. Adult female mosquitoes were morphologically identified on site using taxonomic keys. Molecular species identification was performed by multiplex PCR based on the internal transcribed spacer 2 (ITS2) region of ribosomal DNA (rDNA) and sequencing of the cox1 gene at a DNA barcoding region in a subset of 29 specimens. Results: A total of 1328 An. maculatus (sensu lato) female mosquitoes were captured with An. maculatus, An. sawadwongporni and An. pseudowilmori accounting for 75.2, 22.1 and 2.7% respectively. The field captured mosquitoes of the Maculatus group were most abundant in the wet season and had a preferred distribution in villages at higher elevations. The phylogenetic relationships of 29 cox1 sequences showed a clear-cut separation of the three member species of the Maculatus group, with the An. pseudowillmori cluster being separated from An. sawadwongporni and An. maculatus. Conclusions: This study provides updated information for the species composition, seasonal dynamics and microgeographical distribution of the Maculatus group in malaria-endemic areas of western Thailand. This information can be used to guide the planning and implementation of mosquito control measures in the pursuance of malaria transmission. [ABSTRACT FROM AUTHOR]

Armigeres subalbatus , a mosquito species widely found in Thailand and other Asian countries, serves as a vector for filarial parasites, affecting both humans and animals. However, the surveillance of this vector is complicated because of its morphological similarity to two other species, Armigeres dohami and Armigeres kesseli. To differentiate these morphologically similar species, our study employed both wing geometric morphometrics (GM) and DNA barcoding, offering a comprehensive approach to accurately identify these closely related Armigeres species in Thailand. Our GM analyses based on shape demonstrated significant accuracy in differentiating Armigeres species. Specifically, the outline-based GM method focusing on the 3rd posterior cell exhibited an accuracy rate of 82.61%, closely followed by the landmark-based GM method with 81.54%. Both these GM techniques effectively distinguished Ar. subalbatus from Ar. dohami and Ar. kesseli. Regarding DNA barcoding, our investigation of pairwise intra- and interspecific divergences revealed a "barcoding gap". Furthermore, the results of species confirmation using both species delimitation methods including the automatic barcode gap discovery method (ABGD) and the Multi-rate Poisson tree process (mPTP) were consistent with those of morphological identification, sequence comparisons with the GenBank and Barcode of Life Data System (BOLD) databases, and the neighbor-joining tree construction. These consistent results emphasize the efficacy of DNA barcoding in the precise identification of Armigeres species. • New identification approach for three Armigeres speciesvia wing geometric morphometrics (GM) and DNA barcoding. • GM methods show ∼80% accuracy in differentiating Armigeres species. • DNA barcoding effective, validated by barcoding gap and ABGD/mPTP methods. [ABSTRACT FROM AUTHOR]

Coquillettidia mosquitoes are important nuisance-biting pests and a vector of brugian filariasis in Thailand. However, comprehensive information about these mosquitoes remains unavailable such as molecular and morphometric differences among species. The lack of vector knowledge on Coquillettidia species could affect future disease control. This study aims to investigate differences in molecular variations based on mitochondrial cytochrome oxidase subunit I (COI) gene and wing geometric traits of three Coquillettidia species, namely Cq. crassipes, Cq. nigrosignata, and Cq. ochracea in Thailand. The results of molecular analyses revealed the differences among three Coquillettidia species. The genetic difference measure based on the Kimura two-parameter model among three Coquillettidia species showed low intraspecific distances (0%–3.05%) and large interspecific distances (10.10%–12.41%). The values of intra- and inter-genetic differences of three Coquillettidia species did not overlap which showed the existence of a barcoding gap indicating the efficiency of the identification based on the COI gene. As with molecular analysis, the landmark-based geometric morphometrics approach based on wing shape analysis indicated three distinct species groups which were supported by the high total performance score of cross-validated classification (97.16%). These results provide the first evidence of taxonomic signal based on molecular and wing geometric differences to support species identification and biological variations of Coquillettidia mosquitoes in Thailand for understanding these rare vector mosquitoes in depth and leading to effective further mosquito control. • This study provides the first evidence of molecular and wing geometric differences to support species identification of Coquillettidia crassipes, Cq. nigrosignata , and Cq. ochracea. • The intra- and inter-genetic difference values between the three Coquillettidia species did not overlap, showing the presence of a barcoding gap. • Wing geometric morphometrics analyses based on shape indicated three distinct species groups which were supported by the high total performance score (97.16%). [ABSTRACT FROM AUTHOR]