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Simple Summary: The concentration of larval black flies in well-defined aquatic habitats makes knowledge of the breeding sites critical for the efficient management of pest species. Black flies can achieve pest status either as generalists by developing in many types of streams and rivers or as specialists by developing in one or a few types of flowing water, such as large rivers. These two developmental strategies are evident in pest and vector species of the Simulium (Gomphostilbia) varicorne species group in Thailand. Simulium chumpornense is a habitat generalist, whereas S. khelangense is a habitat specialist, developing in the large Mekong River, where we discovered its immature stages. The first descriptions of the larva and pupa of S. khelangense, along with mitochondrial and nuclear genetic markers, allow accurate identification and comparisons with structurally similar species in the S. varicorne species group, thus aiding the ability to monitor the pest and vector status of black flies in Southeast Asia. Two species of black flies (Simuliidae) in Thailand, Simulium chumpornense Takaoka and Kuvangkadilok, 2000, and S. khelangense Takaoka, Srisuka & Saeung, 2022, are potent vectors of avian blood protozoa of the genera Leucocytozoon and Trypanosoma and are pests of domestic avian species. Although the adults are abundant throughout Thailand, information on their breeding habitats is limited, and the immature stages of S. khelangense are unknown. We collected the larvae and pupae of S. khelangense from the Mekong River, the first-ever record of Simuliidae from this large continental river. Mitochondrial cytochrome c oxidase I and internal transcribed spacer 2 were used to associate the larvae and pupae with known adults. Both genetic markers strongly supported their identity as S. khelangense. The larvae and pupa of S. khelangense are described. The pupal gill filaments, larval abdominal protuberances, and setae distinguish this species from other members of the S. varicorne species group. The immature stages of S. chumpornense inhabit a wide variety of flowing waters, from small streams (3 m wide) to enormous continental rivers (400 m wide); thus, S. chumpornense is a habitat generalist. In contrast, S. khelangense was found only in the large Mekong River and is, therefore, a habitat specialist. Both species can exploit their principal habitats and produce abundant adult populations. [ABSTRACT FROM AUTHOR]

Brugia malayi and B. pahangi, potential zoonotic pathogens transmitted by mosquitoes, are believed to primarily infect dogs and cats as reservoir hosts. Although previous studies have indicated nematode infections in lions, particularly in zoo environments where human contact with these reservoirs is possible, limited documentation exists regarding Brugia sp. infections in lions in Thailand. This study aims to investigate a case of Brugia infection in a lion from a zoo in Thailand. The blood sample was collected and examined from a female lion, using staining methods to morphologically identify microfilaria at the genus level. Subsequently, the PCR was employed targeting specific genes, including mitochondrial 12S rDNA, 18S rDNA, cytochrome oxidase I (COI) and Wolbachia surface protein (wsp), to confirm the species of the filarial nematode parasite. The genetic sequencing results revealed a high similarity (99-100%) to B. malayi for the 12S rDNA, 18S rDNA, COI and wsp genes. Phylogenetic analysis based on nucleotide sequences from the 12S rDNA, 18S rDNA, COI and wsp genes showed that the sequences from this study belong to different clusters. This marks the inaugural documentation of molecular identification of Brugia infection in a lion, signifying that lions could function as reservoirs for this parasite and present a potential public health risk in the region. Our research underscores the effectiveness of molecular techniques and phylogenetic analysis in discerning and comprehending the evolution of filarial parasites. Additionally, it emphasizes the significance of these methods in enhancing the diagnosis, control, and prevention of zoonotic filarial nematode infections. [ABSTRACT FROM AUTHOR]

Background and Aim: Filarial nematode typically produces a larval stage (microfilariae) in the bloodstream of vertebrate hosts, where microfilariae reside in the blood or subcutaneous tissues. Filarial nematodes cause human diseases, such as river blindness and elephantiasis, which are widely studied. However, in avian species, they are overlooked because they are nonpathogenic. In Thailand, microfilaria can be found in wild birds and domestic chickens. Recently, an increase in the number of blood samples submitted to veterinary diagnostic laboratories may have increased the number of microfilariae. Therefore, knowledge about filarial species and reliable detection methods are important. Therefore, this study aimed to investigate the efficacy of buffy coat smear and polymerase chain reaction (PCR)-based methods for the detection of microfilaria in domestic chickens. In addition, parasites were identified using the sequence of the cytochrome c oxidase subunit 1 (COX1) gene. Materials and Methods: Giemsa-stained buffy coat smears from a previous study were reanalyzed. These available buffy coat smears were prepared from 55 domestic chickens raised as backyard free-ranging in Southern Thailand. Fifty-seven frozen genomic DNA extracted from chicken blood were used to detect the presence of the COX1 gene in Onchocercidae nematodes. The nested PCR protocol for amplification of the OnchoCOI_R2-OnchoCOI_R2 fragment of the COX1 gene was applied from a previous report. Sequences of COX1 were analyzed to identify Onchocercidae nematodes and if they were single or mixed infections. We constructed Bayesian phylogenetics to identify parasites and assessment of the relationship between filarial nematodes in avian species and other vertebrate hosts. Results: Buffy coat smears from 15 samples revealed microfilaria. Of these 15 samples, only eight were positive for COX1 nested-PCR amplification. The other two buffy coat-negative samples were also positive for nested-PCR. Sequencing of these 11 nested PCR-positive samples revealed that almost all of them were Onchocercidae nematodes. Bayesian phylogenetic analysis showed that chicken Onchocercidae spp. were grouped with other avian filarial nematodes. However, all chickens Onchocercidae spp. showed a double peak in the sequencing chromatogram, indicating mixed filarial infection (species or haplotypes). Therefore, no chicken Onchocercidae sequence was deposited on National Center for Biotechnology Information, GenBank. Conclusion: Giemsa-stained buffy coat smear was a reliable method for the detection of chicken microfilaria in routine veterinary diagnostic laboratories. Development of a new PCR-based method is necessary. This method may provide greater sensitivity and specificity of detection. In addition, the PCR method allowed us to access the genetic characteristics of nematodes, which helped us maximize our knowledge of nematodes. Further investigations, such as the pathogenicity of filarial nematodes in chickens and their potential vectors, are required. [ABSTRACT FROM AUTHOR]

Simple Summary: The chigger mite Neoschoengastia gallinarum (Hatori, 1920) is a parasite that feeds on the skin tissue of birds across multiple countries in Southeast and East Asia. In domestic chickens, heavy infestations with this mite can lead to skin irritation and damage to the carcass, reducing economic value. In this study, we collected N. gallinarum samples from wild birds of conservation concern and domestic chickens in Peninsular Malaysia and Thailand. Sequence analyses of three genes from the mites were compared across four Malaysian populations, one Thai population, and previously published sequences from southeastern China. A variety of methods were applied to classify these sequences and determine the extent of interbreeding between populations. These methods agreed in identifying three clusters of sequences by country of origin, although there was partial overlap between Thailand and China. The populations from Malaysia and Thailand appear to be reproductively isolated from one another and may represent distinct species with almost identical morphological features, except for leg length. Further studies are required to determine if these genetic dissimilarities are accompanied by distinct ecological, behavioural, or pathological differences in N. gallinarum in different regions of Asia. Neoschoengastia gallinarum is widely distributed in Asia, preferentially parasitising birds, and heavy infestations have clinical impacts on domestic fowl. In common with other trombiculid mites, the genetic diversity and potential variation in host preferences or pathology induced by N. gallinarum are poorly understood. This study aimed to unravel the geographical variation and population structure of N. gallinarum collected from galliform birds in Peninsular Malaysia and Thailand by inference from concatenated mitochondrial-encoded cytochrome c oxidase subunit I (COI), and nuclear-encoded internal transcribed spacer 2 (ITS2) and 18S ribosomal DNA gene sequences, including a comparison with previously published data from southeastern China. Our multi-locus sequence analysis revealed three monophyletic clades comprising (A) specimens from Peninsular Malaysia, (B) the samples from Thailand together with a minority of Chinese sequences, and (C) the majority of sequences from China. Similarly, most species delimitation approaches divided the specimens into three operational taxonomic units. Analysis of molecular variance revealed 96.41% genetic divergence between Malaysian and Thai populations, further supported by the absence of gene flow (Nm = 0.01). In conclusion, despite the two countries sharing a land border, populations of N. gallinarum from Peninsular Malaysia and Thailand appear to be genetically segregated and may represent distinct cryptic species. [ABSTRACT FROM AUTHOR]

Species identification is an essential and vital basis for biological research. Since several fish species have similar appearances and shapes at different developmental stages and researchers lack experience identifying these fish species, molecular techniques are needed. In Thailand, fish species of the family Notopteridae consist of two genera and four species, including Notopterus notopterus , Chitala ornata , Chitala blanci , and Chitala lopis. The combination of the cytochrome c oxidase I (COI) gene sequences with a nonsequencing approach, high-resolution melting (HRM), was developed to discriminate four target species in a rapid, reliable and cost-effective manner. One of three designed primer pair sets, Notop-HRMF1/Notop-HRMR1, was the most appropriate and provided four distinct melting curves. In addition, ten fish species of other families were used for the specificity test, and only the four target species of Notopteridae were specific for this primer pair set. The results of this study indicated that COI-HRM analysis is an efficient approach for identifying species of Notopteridae in Thailand. Applications of this technique may be used to investigate species of this family in processed fish products or larval fish species in several spawning grounds for sustainable fishery resource management. [ABSTRACT FROM AUTHOR]

Background and Aim: Freshwater snails belonging to the family Planorbidae serve as the first intermediate hosts of many species of important parasitic flukes of animals and humans. Information regarding the occurrence of planorbid snail larval trematode infection is limited in Northern Thailand. Thus, this study aimed to estimate the prevalence of larval trematode infection of the freshwater snail Indoplanorbis exustus in Uttaradit, Thailand, and to identify trematode species based on their morphological and molecular characteristics. Materials and Methods: Planorbid snail specimens were collected from a water reservoir in Uttaradit, Thailand, from June to August 2023. Snails were assessed for larval trematode infection through cercarial shedding and crushing methods. The released cercariae were preliminarily identified on the basis of their morphological characteristics. In addition, species identification of the detected cercariae was conducted using 28S ribosomal RNA and cytochrome c oxidase subunit 1 gene sequence analyses. Results: The overall prevalence of cercarial infection was 61.5% (107/174) in planorbid snails in Uttaradit province. Two species of cercarial trematodes, Schistosoma spindale (106/174, 60.9%) and Artyfechinostomum malayanum (1/174, 0.6%), were identified using morphological and molecular analyses, of which S. spindale was the most abundant species. Our studied snails did not have mixed infection with more than two cercarial species. Conclusion: Our findings reveal a remarkably high prevalence of S. spindale cercariae infecting planorbid snails in Uttaradit, indicating that humans and animals across the study area are at risk of infection. Our data may contribute to the development of effective strategies to control this zoonotic infectious disease. [ABSTRACT FROM AUTHOR]

Hookworm infections remain a significant public health concern in tropical and subtropical regions, including Thailand. This study investigated the species and genetic diversity of hookworm infections in domestic dogs from northeastern Thailand. The molecular analysis focused on amplifying and sequencing specific regions of ribosomal RNA genes (ITS1-5.8S-ITS2 region) and the mitochondrial cytochrome c oxidase subunit 1 (cox1) gene in hookworm larvae recovered from 21 domestic dog stool samples. Among 21 larvae (one larva per infected dog) analyzed, 14 had sequences identical to Ancylostoma caninum, and 7 showed sequences almost identical to Ancylostoma ceylanicum. Phylogenetic analysis of cox1 sequences placed A. caninum and A. ceylanicum in separate clades. The median-joining network of A. caninum cox1 sequences from Thailand showed high haplotype diversity and belonged to the same cluster as sequences from Australia while forming separate clusters from those of A. caninum samples from the USA. The available published A. ceylanicum cox1 sequences (n = 33), in combination with seven sequences in the present study, represented 15 haplotypes distributed among three clusters. Interestingly, A. ceylanicum sequences from dogs and humans shared the same haplotypes. These findings are crucial for recognizing the potential for zoonotic transmission, highlighting the necessity for targeted control measures, and increasing awareness among pet owners and healthcare professionals to mitigate the risk of hookworm transmission to humans. [ABSTRACT FROM AUTHOR]

We described a new species of elapid snake genus Bungarus from the Tenasserim Mountain Range in Ratchaburi Province, western Thailand. Bungarus sagittatus sp. nov. can be distinguished from all congeners by having the combination of 15 dorsal scale rows; 215–217 ventral scales; 48–56 undivided subcaudal; prefrontal suture 2.4–2.6 times length of internasal suture; anterior chin shields larger than posterior chin shields; head of adult uniform black while juvenile black with small dim white patches on temporal and parietal areas; dorsal body black, with 25–31 white narrow bands, white and black bands at midbody covering 1.5–3.0 and 4.5–6.0 vertebral scales, respectively; dorsal body black bands not intruding ventrals or intruding ventrals less than 0.5 times of width of outer dorsal scales; ventral surface of body immaculate white; ventral side of tail white with a row of dark brown triangular patches on middle pointing posteriorly; tail relatively long, tail length/total length 0.140–0.143. Genetically, the new species has uncorrected pairwise divergences of ≥ 8.29% of the mitochondrial cytochrome b from other Bungarus species. Currently, the new species is only known from the type locality. [ABSTRACT FROM AUTHOR]

Background: Babesia is a protozoal, tick-borne parasite that can cause life-threatening disease in humans, wildlife and domestic animals worldwide. However, in Southeast Asia, little is known about the prevalence and diversity of Babesia species present in wildlife and the tick vectors responsible for its transmission. Recently, a novel Babesia species was reported in confiscated Sunda pangolins (Manis javanica) in Thailand. To investigate the presence of this parasite in Singapore, we conducted a molecular survey of Babesia spp. in free-roaming Sunda pangolins and their main ectoparasite, the Amblyomma javanense tick. Methods: Ticks and tissue samples were opportunistically collected from live and dead Sunda pangolins and screened using a PCR assay targeting the 18S rRNA gene of Babesia spp. DNA barcoding of the cytochrome oxidase subunit I (COI) mitochondrial gene was used to confirm the species of ticks that were Babesia positive. Results: A total of 296 ticks and 40 tissue samples were obtained from 21 Sunda pangolins throughout the 1-year study period. Babesia DNA was detected in five A. javanense ticks (minimum infection rate = 1.7%) and in nine different pangolins (52.9%) located across the country. Phylogenetic analysis revealed that the Babesia 18S sequences obtained from these samples grouped into a single monophyletic clade together with those derived from Sunda pangolins in Thailand and that this evolutionarily distinct species is basal to the Babesia sensu stricto clade, which encompasses a range of Babesia species that infect both domestic and wildlife vertebrate hosts. Conclusions: This is the first report documenting the detection of a Babesia species in A. javanense ticks, the main ectoparasite of Sunda pangolins. While our results showed that A. javanense can carry this novel Babesia sp., additional confirmatory studies are required to demonstrate vector competency. Further studies are also necessary to investigate the role of other transmission pathways given the low infection rate of ticks in relation to the high infection rate of Sunda pangolins. Although it appears that this novel Babesia sp. is of little to no pathogenicity to Sunda pangolins, its potential to cause disease in other animals or humans cannot be ruled out. [ABSTRACT FROM AUTHOR]

Previously, the euthyplociid mayfly from Thailand was reported as Polyplocia spp. without designation to any nominal species. In this study, the genus Polyplocia Lestage, 1921 in Thailand was reviewed. Two species are recognized: Polyplocia orientalis Nguyen & Bae, 2003 and P. nebulosa Gonçalves & Peters, 2016. This is the first report of P. nebulosa in Continental Asia. Cytochrome c oxidase subunit I (COI) data and illustrations based on nymphal characters were used to confirm two different species in Thailand. Additional morphological characteristics of the nymph and eggs of the two species from Thailand are also shown in detail. The taxonomic status of Polyplocia in the Oriental region is reviewed. Keys to known nymphal and imaginal stages are provided and the distribution of Polyplocia in the Oriental region is discussed. [ABSTRACT FROM AUTHOR]

Angiostrongylus cantonensis, or the rat lungworm, is the causative agent of human angiostrongyliasis associated with eosinophilic meningitis or meningoencephalitis. Additionally, this nematode can cause ocular angiostrongyliasis, though this is rare. The worm can cause permanent damage to the affected eye and sometimes even blindness. Genetic characterization of the worm from clinical samples is limited. In the present study, we investigated the genetics of A. cantonensis recovered from a patient's eye in Thailand. We sequenced two mitochondrial genes (cytochrome c oxidase subunit I, or COI, and cytochrome b, or cytb) and nuclear gene regions (66-kDa protein and internal transcribed spacer 2, or ITS2) from a fifth-stage larva of Angiostrongylus sample that was surgically removed from the human eye. All sequences of the selected nucleotide regions were highly similar (98–100%) to the sequences of A. cantonensis in the GenBank database. The maximum likelihood and neighbor-joining trees of the COI gene indicated that A. cantonensis was closely related to the AC4 haplotype, whereas the cytb and 66-kDa protein genes were closely clustered with the AC6 and Ac66-1 haplotypes, respectively. In addition, the phylogeny of the concatenated nucleotide datasets of the COI and cytb revealed that the worm was closely related to the Thai strain and strains from other countries. This study confirms the identification and genetic variation of the fifth-stage larvae of A. cantonensis recovered from a patient's eye in Thailand. Our findings are important for future research on the genetic variation of A. cantonensis that causes human angiostrongyliasis. [ABSTRACT FROM AUTHOR]

The Euconulidae is a globally distributed land snail family but there is no record of this family from Thailand. In this study, we describe a new genus, Siamoconus gen. nov., based on comparative studies of the shell characteristics, external morphology of animals, radula, genital and spermatophore structures, and molecular phylogeny. We performed phylogenetic analyses of two mitochondrial gene fragment (cytochrome c oxidase I, COI; and 16S rRNA) and one nuclear (28S rRNA) gene fragment to clarify the relationships with other euconulid genera and its position in the superfamily Trochomorphoidea. We also analysed shell morphometrics, male genitalia and mantle pigmentation of Siamoconus gen. nov., confirming the status of three new species with keeled shells (S. boreas sp. nov., S. coleus sp. nov. and S. geotrochoides sp. nov.) and one new species with a rounded shell (S. destitutus sp. nov.). These new species are restricted to limestone areas in northern and north-eastern Thailand. We also re-examined the status of other genera in the superfamily Trochomorphoidea based on shell features, body pigmentation, radula, genital anatomy, spermatophore morphology, and a phylogenetic analysis of all available trochomorphoidean DNA sequences. Our analysis suggests that the family Geotrochidae, previously synonymised with the Trochomorphidae, should be resurrected to represent the genus Geotrochus from Borneo, and this family is retrieved as the sister clade of the Dyakiidae. ZooBank: urn:lsid:zoobank.org:pub:C5205F4E-5DDE-42E6-A532-761CAFE099C3 Euconulidae is one of the most diverse families in the superfamily Trochomorphoidea with a distribution that is almost world-wide but there is no record of this family from Thailand. We describe four new species of Siamoconus gen. nov., a new euconulid genus and also perform phylogenetic analyses of one nuclear and two mitochondrial gene fragments to clarify the systematic position of this new genus and the relationships to other genera in the superfamily Trochomorphoidea. In addition, the family Geotrochidae, previously synonymised with the Trochomorphidae, is resurrected as sister clade of the Dyakiidae within the superfamily Trochmorphoidea. [ABSTRACT FROM AUTHOR]

Shark fins are among the most highly prized seafood products in the world with massive consumption in Asia over the past several decades. The demand for shark fins is a major driver of the enormous population declines of elasmobranchs that are generally vulnerable to overexploitation. This study aims to better understand the species composition of shark fin products in Thailand and their conservation statuses by using DNA-based species identification. Various types and sizes of shark fins were collected from 4 locations in Thailand. DNA barcoding method based on a fragment of the cytochrome c oxidase subunit I (COI) gene was applied to species identification. Fins from at least 15 shark species were found from Thailand's markets. The spottail shark (Carcharhinus sorrah) and the night shark (Carcharhinus signatus) were the two dominant species presented in this study. 34% of identifiable samples are the species that have not been record in this region. 62% of species detected from the fin samples are categorized under the threatened categories of IUCN Red List. Species composition reported in shark fin products potentially helps indicate the appropriate conservation action and increases awareness from monitoring the trade in elasmobranch products. [ABSTRACT FROM AUTHOR]

Rapid genetic improvements from selective breeding are anticipated in many aquaculture species and mitogenome is used to complement the morphological taxonomy of hybrids to evaluate its genetic structures. A study of the genetic variations within the two strains (Malaysian strain and Golden Thai strain) of Anabas testudineus (Bloch, 1792) and their hybrids, phylogenetic trees based on cytochrome oxidase subunit 1 (CO1) of the partial mitochondrial DNA gene were constructed using Maximum Likelihood and Neighbor Joining approaches. The findings support the monophyletic status of the genus with only one haplogroup from which other haplotypes were descended and of a single common ancestor. The individual fishes’ phylogenetic relationships revealed two major clades and Saurida undosquamis as an outgroup. All the groups had high haplotype diversity, except for the hybrid Malaysian strain × golden Thai strain (0.1540±0.126). This suggests that different fish species in each of these studies had different nucleotide compositions in their mitochondrial genomes. The highest number of haplotypes and the presence of distinct haplotypes in the Golden Thai strain × Malaysian strain hybrid point to the absence of recent or regular gene flow as well as high genetic diversity within the hybrids. This study demonstrated that the mtDNA diversity of Anabas testudineus from Malaysia and Thailand had been preserved. Studies on the population genetic diversity and molecular evolution of anabas fish species can benefit from the data provided by this work. [ABSTRACT FROM AUTHOR]

To establish sustainable resources and founder populations for genetic improvement of the Siamese fighting fish Betta splendens, genetic diversity in wild and hatchery stocks was examined using mitochondrial (mt) DNA genes cytochrome b (cytb), 16S ribosomal DNA (16S rDNA), and cytochrome oxidase subunit I (COI), and eight microsatellite loci. Based on mtDNA sequences, restrictive levels of polymorphism (0, 3, and 1 substitutions) were observed in this study. For analysis of microsatellites, fluorescent multiplex PCR was developed, and subsequently identifying moderate levels of observed (Ho = 0.4488) and expected (He = 0.6627) heterozygosities and a high number of alleles per locus (15.125 alleles) for overall samples. Comparison of Siamese fighting fish from different sources revealed large genetic differences between pairs of farmed fish (eight groups) and between wild (three geographic locations) and farmed fish (P < 0.0031 following Bonferroni correction). This suggested limited exchanges of genetic resources between commercial farms. When different color varieties of B. splendens were compared, large genetic distances and significant FST estimates and genetic heterogeneity were found (P < 0.0031). Effective population sizes (Ne) were estimated and two farms (NP2-2BS and BK1-4BS) showed Ne greater than 10. Among color varieties, Multi-colors and Blue revealed reasonable Ne (large and 27.9), but lower Ne values (3.6–8.4) were found for the remaining color varieties. These results indicate an urgent need for the establishment of gene pool resources of B. splendens for effective genetic improvement of Siamese fighting fish in Thailand. [ABSTRACT FROM AUTHOR]

Unlike malaria parasites in humans, non-human primates, rodents, and birds, ungulate malaria parasites and their vectors have received little attention. As a result, understanding of the hosts, vectors, and biology of ungulate malaria parasites has remained limited. In this study, we aimed to identify the vectors of the goat malaria parasite Plasmodium caprae. A total of 1019 anopheline and 133 non-anopheline mosquitoes were collected from goat farms in Thailand, where P. caprae-infected goats were discovered. Anopheline mosquitoes were identified using molecular biological methods that target the cytochrome c oxidase subunit 1 (cox1), the cytochrome c oxidase subunit 2 (cox2) genes, and the internal transcribed spacer 2 (ITS2) region. Pool and individual mosquitoes were tested for P. caprae using the head-thorax parts that contain the salivary glands, with primers targeting three genetic markers including cytochrome b, cytochrome c oxidase subunit 1, and 18S small subunit ribosomal RNA genes. Additionally, goat blood samples were collected concurrently with mosquito surveys and screened to determine the status of malaria infection. This study revealed nine mosquito species belonging to six groups on goat farms, including Hyrcanus, Barbirostris, Subpictus, Funestus, Tessellatus, and Annularis. The DNA of P. caprae was detected in Anopheles subpictus and Anopheles aconitus. This is the first time An. subpictus and An. aconitus have been implicated as probable vectors of P. caprae. [ABSTRACT FROM AUTHOR]

• This study successfully identified 21 fly species among three dipteran families (Calliphoridae, Muscidae, and Sarcophagidae) collected from 18 provinces in Thailand using DNA barcoding. • This study confirmed 100 % alignment between morphological identification and molecular characteristics, thus validating reliability of both methods. • New information of 15 species, including their morphologies, COI gene sequences, and locations of collection, from Thailand was deposited firstly in the BOLD system. Forensic entomology plays a crucial role in criminal investigations by providing vital insights into minimum postmortem interval (PMI min) and corpse relocation by identifying insect species that colonize in decomposing remains. This study aimed to identify and analyze the genetic variation of forensically significant fly species in Thailand, using DNA barcoding of the mitochondrial cytochrome c oxidase subunit I COI gene. A total of 3,220 fly specimens were collected from 18 provinces across six regions of Thailand from October 2017 to September 2022. These specimens were classified by morphological identification into 21 species among three Dipteran families: Calliphoridae, Muscidae, and Sarcophagidae, with Chrysomya megacephala Diptera: Calliphoridae being the most abundant species. DNA barcoding confirmed the morphological identifications with 100 % accuracy, showing low intraspecific K2P distances0.0 to 1.1 %) and significant interspecific K2P distances 2.5 % to 17.2 %. A Neighbour-Joining (NJ) analysis was conducted to assess the molecular identification capabilities of the barcoding region. This analysis successfully recovered nearly all species as distinct monophyletic groups. The species groupings obtained were generally consistent with both morphological and molecular identifications. These findings underscore the effectiveness of DNA barcoding for precise species identification and contribute to a comprehensive database of forensically important flies in Thailand, thus facilitating improved forensic investigations and biodiversity studies. [Display omitted] [ABSTRACT FROM AUTHOR]

The first Panorpa was recently discovered in Thailand even though the genus had not been thought to occur south of the Himalaya. We now add a second new species, Panorpa apscisaceran. sp., also from Northern Thailand, which has nearly immaculate pale-yellow wings with only a pronounced stigma. The male has a blunt, vestigial bifid anal horn, and thin, bare ventral parameres. Additionally, we describe Neopanorpa appendiceman. sp. with mesad-curled male hypovalve apices and large, basomesal lobes, a relatively large species, N. inchoatan. sp., with overlapping elliptical hypovalves and incomplete apical bands, N. setosiloban. sp. with huge bristly lobes on the mesal margin of the male gonostyles, and N. mandangensisn. sp. with a narrow 3rd tergum process extending to the posterior margin of the 4th tergum. Neopanorpa nielseniByers, 1965, previously known from northern Vietnam and southern China, has recently been discovered in northern Thailand. Bittacus leptocaudusByers, 1965, known from a lone male has been rediscovered and locality details and photo of the habitat provided. These bring the number of species of Mecoptera occurring in Thailand to 22 (19 Neopanorpa, two Panorpa, and one Bittacus) and the number of Mecoptera species known from Indochina to 56 (48 Neopanorpa, three Panorpa, 4 Bittacus, and one Bicaubittacus). A key to the species of Mecoptera in Thailand with illustrations is provided. DNA was extracted from specimens and the DNA Barcode, mitochondrial cytochrome oxidase I (COI) gene fragment sequenced and analyzed for 19 of the 22 species of Thai Mecoptera. The results are figured as a Neighbor-Joining tree. แมลงแมงป่องสกุล Panorpa ไม่เคยมีรายงานจากทางตอนใต้ของเทือกเขาฮิมาลายา (Himalaya) มาก่อน จนกระทั่งในปี พ.ศ. 2562 แมลงแมงป่องสกุลนี้ได้ถูกรายงานเป็นครั้งแรกในประเทศไทย งานวิจัยนี้ได้ค้นพบ แมลงแมงป่องชนิดใหม่ Panorpa apscisacera n. sp. จากภาคเหนือและเป็นชนิดที่สองของประเทศไทย ลักษณะ เด่นของปีกสีเหลืองอ่อนเกือบทั่วปีก มีตาปีก (stigma) ชัดเจน anal horn ในเพศผู้สั้นและแยกเป็นสองแฉกแต่ ไม่ชัดเจน ventral paramere บางและไม่มีขนปกคลุม นอกจากนี้ยังค้นพบแมลงแมงป่องในสกุล Neopanorpa 4 ชนิดใหม่ ได้แก่ ชนิด N. appendicema n. sp. เป็นชนิดที่มีขนาดค่อนข้างใหญ่ ส่วนกลางของ hypovalve ม้วนขด และ basomesal lobe ขนาดใหญ่ ชนิด N. inchoata n. sp. มี hypovalve รูปร่างยาวและวางซ้อนทับ กัน แถบสีที่ปลายปีกไม่สมบูรณ์ ชนิด N. setosiloba n. sp. ในเพศผู้มี bristly lobes ขนาดใหญ่ที่ gonostyles และชนิด N. mandangensis n. sp. ด้านบนของท้องปล้องที่สามมี notal organ เรียวและยื่นยาวไปถึงส่วนท้าย ของปล้องที่สี่ แมลงแมงป่องชนิด N. nielseni Byers, 1965 ถูกค้นพบเป็นครั้งแรกในประเทศไทยจากภาคเหนือ ซึ่งเคยมีรายงานจากภาคเหนือของประเทศเวียดนามและทางตอนใต้ของประเทศจีน และได้มีการค้นพบชนิด Bittacus leptocaudus Byers, 1965 อีกครั้งจากที่เคยรายงานเพศผู้เพียงหนึ่งตัว มีการเพิ่มเติมข้อมูลของพื้นที่ เก็บตัวอย่างและภาพถ่ายในธรรมชาติ การรายงานนี้ทําาให้จําานวนชนิดของแมลงแมงป่องในประเทศไทยเพิ่ม ขึ้นเป็น 22 ชนิด ใน 3 สกุล ได้แก่ สกุล Neopanorpa จําานวน 19 ชนิด สกุล Panorpa จําานวน 2 ชนิด และสกุล Bittacus จําานวน 1 ชนิด และทําาให้ในภูมิภาคอินโดจีน (Indochina) มีจําานวนชนิดเพิ่มขึ้นเป็น 56 ชนิด (สกุล Neopanorpa 48 ชนิด, Panorpa 3 ชนิด, Bittacus 4 ชนิด, และ Bicaubittacus 1 ชนิด) ทั้งนี้ยังได้จัดทําารูปวิธาน และรูปภาพประกอบการระบุชนิดของแมลงแมงป่องในประเทศไทย ทําาการวิเคราะห์ลําาดับยีนของไมโตคอนเด รียในส่ วนของ cytochrome oxidase I (COI) ของแมลงแมงป่ องจ าานวน 19 ชนิด จาก 22 ชนิดของประเทศไทย และรายงานผลโดยใช้ Neighbor Joining tree. [ABSTRACT FROM AUTHOR]

Genetic information is crucial to manage fish resources, and a good case in point is the population of silver croakers, Pennahia argentata (Houttuyn, 1782), which is economically important for Thailand. In the present study, 102 silver croaker samples were collected along the coast of the Gulf of Thailand and analysed for genetic variation based on nucleotide sequences in the cytochrome oxidase subunit I gene (510 bp). Of these, 33 haplotypes were examined, and 21 were singleton haplotypes, indicating a historical pattern of large female effective population sizes (female reproductive success). An analysis of molecular variance (AMOVA) and pairwise FST analysis showed that the geographic barrier did not affect the genetic structure of the silver croakers in the Gulf of Thailand. The minimum spanning network and phylogenetic tree revealed that the silver croaker population in the Gulf of Thailand separated into two haplogroups. Various methods to examine demographic history showed that the silver croaker population in the Gulf of Thailand had expanded. This study's findings can guide the management of silver croaker populations in the Gulf of Thailand to conserve genetic diversity. [ABSTRACT FROM AUTHOR]

Enterobius vermicularis, a nematode parasite with a global distribution causes enterobiasis in schoolchildren and is considered a neglected parasite. An understanding of the prevalence and genetic diversity of enterobiasis is crucial for appropriate control measures. Therefore, the objectives of this research were to study the prevalence and genetic diversity of E. vermicularis in schoolchildren from lower northern Thailand, based on cytochrome c oxidase subunit I (COI) and internal transcribed spacer 2 (ITS2) sequences. Using the scotch tape technique, 7.4% (188/2544) of schoolchildren from 21 primary schools were found positive for E. vermicularis eggs, which is a relatively low infection rate. Phylogenetic trees of partial COI sequences (397 bp) revealed similar topologies using maximum likelihood (ML) and neighbor-joining (NJ) methods and identified E. vermicularis type A (105 sequences) and B (1 sequence). Haplotype network analysis of the COI sequences demonstrated a high haplotype diversity (Hd = 0.9028). In contrast, phylogenetic analysts of a 343 bp region of the ITS2 locus (52 sequences) revealed a monophyletic group. More sequence analyses of E. vermicularis from humans and other hosts in Thailand are necessary to better understand the genetic diversity of this parasite. [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]

The giant water bug, Lethocerus indicus (Lepeletier and Serville, 1825), is one of the most important edible insects in Thailand. This study investigated the genetic diversity of L. indicus from natural resources in northeast Thailand by using mitochondrial DNA sequences. Ninety adults of L. indicus were collected from 12 different localities by light trap and hand-net methods. Nucleotide and haplotype diversity of cytochrome c oxidase subunit 1 (CO1) sequences were relatively high. The 43 CO1 haplotypes (Li1–Li43) with 26 unique haplotypes were classified and used to generate a star-like haplotype network. The most common haplotypes were Li6 and Li2 shared between 19 and 15 sequences, respectively from nine different localities. A phylogenetic tree clearly separated L. indicus from the other species and closely clustered with a sibling species, L. patruelis (Stål, 1854). However, the comprehensive genetic variation of L. indicus covering broad range of areas needs to be further explored. [ABSTRACT FROM AUTHOR]

Anopheles (Cellia) dirus Peyton & Harrison and Anopheles baimaii Sallum & Peyton are sibling species within the Dirus complex belonging to the Leucosphyrus group, and have been incriminated as primary vectors of malaria in Thailand. In the present study, DNA barcoding and geometric morphometrics were used to distinguish between An. dirus and An. baimaii in the international border areas, Trat Province, eastern Thailand. Our results revealed that DNA barcoding based on the cytochrome c oxidase subunit I gene could not be used to distinguish An. dirus from An. baimaii. The overlapping values between intra- and interspecific genetic divergence indicated no barcoding gap present for An. dirus and An. baimaii (ranging from 0 to 0.99%). However, the results of the geometric morphometric analysis based on the wing shape clearly distinguished An. dirus and An. baimaii, with 92.42% of specimens assigned to the correct species. We concluded that geometric morphometrics is an effective tool for the correct species identification of these two malaria vectors. Our findings could be used to make entomological surveillance information more accurate, leading to further effective mosquito control planning in Thailand and other countries in Southeast Asia. [ABSTRACT FROM AUTHOR]

Simple Summary: Biting midges are important blood sucking insects. Many species are pests of humans and other animals, including economically important livestock. A new biting midge species, Culicoides mahasarakhamense, was recently described in Thailand. This species is a pest and a potential vector of blood protozoa transmitted to domestic chickens and other avian species. Knowledge of genetic diversity and genetic differentiation among populations of vector species is of paramount importance as it can be used to design effective control programs. However, this information is lacking for C. mahasarakhamense. In this study, we used mitochondrial cytochrome c oxidase I (COI) sequences as genetic markers to evaluate the level of genetic diversity and the genetic structure of C. mahasarakhamense from Thailand. The level of genetic diversity was relatively high, but most populations were not genetically different. However, one population from northern Thailand was found to be genetically highly different from others. Historical population isolation and geographic distance separation are possible explanations for the high level of genetic differentiation. This information will help in monitoring the spread of the parasitic disease agents carried by C. mahasarakhamense from different populations. Biting midges of the genus Culicoides Latreille are significant pests and vectors of disease agents transmitted to humans and other animals. Understanding the genetic structure and diversity of these insects is crucial for effective control programs. This study examined the genetic diversity, genetic structure, and demographic history of Culicoides mahasarakhamense, a possible vector of avian haemosporidian parasites and Leishmania martiniquensis, in Thailand. The star-like shape of the median joining haplotype network, a unimodal mismatch distribution, and significant negative values for Tajima's D and Fu's FS tests indicated that populations had undergone recent expansion. Population expansion time was estimated to be 2000–22,000 years ago. Population expansion may have been triggered by climatic amelioration from cold/dry to warm/humid conditions at the end of the last glaciations, resulting in the increased availability of host blood sources. Population pairwise FST revealed that most (87%) comparisons were not genetically different, most likely due to a shared recent history. The exception to the generally low level of genetic structuring is a population from the northern region that is genetically highly different from others. Population isolation in the past and the limitation of ongoing gene flows due to large geographic distance separation are possible explanations for genetic differentiation. [ABSTRACT FROM AUTHOR]

Anisakis nematodes infecting Indian mackerel (Rastrelliger kanagurta) were initially discovered in Thailand in our preliminary investigation. Nevertheless, the species of Anisakis collected has not been determined nor has its genetic variation been researched. Thus, this study aimed to molecularly identify the species of Anisakis specimens using the internal transcribed spacer (ITS) region of ribosomal DNA sequences. In addition, the intraspecific genetic variation was also determined using mitochondrial cytochrome oxidase subunit II (COII) gene sequences. The phylogenetic relationships of the ITS region classified all samples into Anisakis typica; however, the genetic variation between them could not be distinguished. By contrast, the phylogenetic tree analysis of the COII region identified all samples as A. typica, with 17 different haplotypes by 66 polymorphic sites and five of the substitutions resulted in amino acid change. Additionally, the distribution pattern of the COII region can be separated into two groups between South America and Asian countries. All our haplotypes belong to Asian countries. Compared with the two genetic markers used in this investigation, COII appears to be a better candidate for studying genetic variation sensitive to environmental changes and intermediate or definitive host behavioral changes. [ABSTRACT FROM AUTHOR]

Chamberlainia hainesiana (Lea, 1856) (Unionidae) is the most economically important freshwater mussel in Thailand and is commonly used in food, spiritual ceremonies and pearl culture. Despite the clear economic importance of this monotypic genus, the distribution and diversity of Chamberlainia Simpson, 1900 are poorly understood. We set out to re-evaluate the taxonomic and geographic boundaries of Chamberlainia using molecular and morphological data gathered from recently and historically collected material. Our cytochrome c oxidase subunit I gene tree recovered three divergent species-level lineages in the genus Chamberlainia. Fourier shape analysis of 60 digitized Chamberlainia shell outlines revealed evidence of two morphologically divergent groups that could be statistically distinguished 88% of the time. We used this evidence to recognize two genetically and morphologically divergent Chamberlainia species, C. hainesiana and C. duclerci (Rochebrune, 1882) stat. res. Geographical information from 46 museum records was used to map the distribution of Chamberlainia. These results are discussed in the context of their taxonomic, biogeographic and conservation implications. [ABSTRACT FROM AUTHOR]

Pseudocypretta maculata Klie, 1932 , type species of the genus, is redescribed based on new material from Thailand. The main diagnostic features of the species are the presence of marginal septa, presence of Wouters organ on the first antenna, strongly serrated claw G2 of the second antenna (A2), small β seta on the mandibular palp, elongated terminal segment of the maxillular palp, smooth and large bristles on the third endite of the maxillula, undivided penultimate segment of the second thoracopod (T2), absence of d1 seta on the T2, completely separated terminal segment of the third thoracopod, absence of a pincer organ, and reduced caudal ramus with a triangular base. The taxonomic position of Pseudocypretta Klie, 1932 is also revised. Based on molecular analysis using the cytochrome c oxidase subunit I (COI), Pseudocypretta appears closer to Cypridopsinae Kaufmann, 1900 than to Cyprettinae Hartmann, 1971. This result is congruent with morphological evidence and, among other cypridopsine genera, Pseudocypretta shows highest similarity with Cyprettadopsis Savatenalinton, 2020 given its reduced caudal ramus, the strongly serrated claw G2 of the A2, absence of pincer organ, and presence of marginal septa. Pseudocypretta is therefore allocated to the tribe Cyprettadopsini Savatenalinton, 2020. [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]

• Cryptic genetic lineages of Simulium chumpornense were evaluated. • Morphology and molecular data indicated the lineages are different species. • ITS2 sequences accurately identified cryptic species of S. chumpornense. • Two Leucocytozoon and one Trypanosoma species were found in S. khelangense. Species of the Simulium varicorne group in Thailand have veterinary significance as vectors of haemosporidian parasites. Accurate identification is, therefore, critical to the study of vectors and parasites. We used morphology and molecular markers to investigate cryptic genetic lineages in samples identified as Simulium chumpornense Takaoka & Kuvangkadilok, 2000. We also tested the efficiency of the nuclear internal transcribed spacer 2 (ITS2) marker for the identification of species in this group. Morphological examinations revealed that S. chumpornense lineage A is most similar to S. khelangense Takaoka, Srisuka & Saeung, 2022, with minor morphological differences. They are also genetically similar based on mitochondrial cytochrome c oxidase I (COI) sequences. Geographically, the sampling site where paratypes of S. khelangense were originally collected is <50 km from where S. chumpornense lineage A was collected. We concluded that cryptic lineage A of S. chumpornense is actually S. khelangense. COI sequences could not differentiate S. kuvangkadilokae Pramual and Tangkawanit, 2008 from S. chumpornense and S. khelangense. In contrast, ITS2 sequences provided perfect accuracy in the identification of these species. Molecular analyses of the blood protozoa Leucocytozoon and Trypanosoma demonstrated that S. khelangense carries L. shoutedeni, Leucocytozoon sp., and Trypanosoma avium. The Leucocytozoon sp. in S. khelangense differs genetically from that in S. asakoae Takaoka & Davies, 1995, signaling the possibility of vector-parasite specificity. [ABSTRACT FROM AUTHOR]

Ungulate malaria parasites and their vectors are among the least studied when compared to other medically important species. As a result, a thorough understanding of ungulate malaria parasites, hosts, and mosquito vectors has been lacking, necessitating additional research efforts. This study aimed to identify the vector(s) of Plasmodium bubalis. A total of 187 female mosquitoes (133 Anopheles spp., 24 Culex spp., 24 Aedes spp., and 6 Mansonia spp. collected from a buffalo farm in Thailand where concurrently collected water buffalo samples were examined and we found only Anopheles spp. samples were P. bubalis positive. Molecular identification of anopheline mosquito species was conducted by sequencing of the PCR products targeting cytochrome c oxidase subunit 1 (cox1), cytochrome c oxidase subunit 2 (cox2), and internal transcribed spacer 2 (ITS2) markers. We observed 5 distinct groups of anopheline mosquitoes: Barbirostris, Hyrcanus, Ludlowae, Funestus, and Jamesii groups. The Barbirostris group (Anopheles wejchoochotei or Anopheles campestris) and the Hyrcanus group (Anopheles peditaeniatus) were positive for P. bubalis. Thus, for the first time, our study implicated these anopheline mosquito species as probable vectors of P. bubalis in Thailand. [ABSTRACT FROM AUTHOR]

In total, 160 ticks infesting cattle in the northeast region of Thailand were collected and used for molecular investigation. Three tick species—Rhipicephalus microplus Canestrini, Rhipicephalus haemaphysaloides Supino and Haemaphysalis bispinosa Neumann—were identified based on morphology and DNA sequences of mitochondrial cytochrome c oxidase subunit 1 (CO1) and 16S ribosomal RNA (16S rRNA). In total, 26 and seven unique haplotypes of the CO1 and 16S rRNA genes, respectively, were recovered. Phylogenetic analysis using the CO1 sequence revealed that the R. microplus from northeastern Thailand were grouped into the previously described clades A and C, whereas the 16S rRNA phylogenetic tree assigned all isolates of R. microplus from Northeast Thailand into the previously described clade B. Clade C of the CO1 phylogenetic tree is a new genetic assemblage recently discovered from India and Malaysia, which has now been detected in our study. The haplotype network also demonstrated that R. microplus is divided into two haplogroups corresponding to the assemblage of the CO1 phylogenetic tree. Our findings strongly support the previous genetic assemblage classification and evidence that R. microplus from Northeast Thailand is a species complex comprising at least two genetic assemblages, i.e., clades A and C. However, further investigation is needed and should involve more comprehensive genetic and morphological analyses and cover a larger part of their distributional range throughout Southeast Asia. [ABSTRACT FROM AUTHOR]

Objective: Rare disease Background: Neurocysticercosis is the most common central nervous system infection in developing countries. A wide array of clinical manifestations, ranging from asymptomatic to severe neurological symptoms, is observed in patients diagnosed with neurocysticercosis, depending on the number of lesions, cyst location, cyst stage, parasite genotype, and host immunity. Case Report: We report the case of a 25-year-old Burmese man who presented with focal seizure and secondary generalized tonic-clonic seizure. Brain imaging studies revealed a 1-cm cyst, which showed rim enhancement, an eccentric scolex, and surrounding brain edema at the left superior frontal gyrus. His serum cysticercus antibody was positive. Thus, the patient was diagnosed with solitary neurocysticercosis based on clinical manifestations, neuroimaging findings, and positive serology. The patient received anti-parasitic and anti-seizure medications before surgical excision of the cyst via computed tomography (CT) scan navigation. Stereomicroscopic examination of the cyst revealed a parasite larva in a fluid-filled cyst, containing a scolex with hooks and 4 suckers, identical to that of Taenia solium. Molecular characterization of the parasite based on T. solium cytochrome c oxidase subunit 1 (COX-1) gene identified the species as being 99.7% identical to T. solium Asia genotype previously reported from pigs in Thailand. Conclusions: Although the prevalence of neurocysticercosis seems to be declining, sporadic cases have been reported throughout the world and the prevalence may be underestimated. Differential diagnosis of neurocysticercosis in patients presenting with adult-onset epilepsy should be considered in disease-endemic areas. [ABSTRACT FROM AUTHOR]

Hitherto, the millipede genus Apeuthes (family Pachybolidae, subfamily Trigoniulinae) was only known from three species described in Vietnam based on morphological characters. The present study uses two partial mitochondrial gene fragments (cytochrome c oxidase I (COI) and 16S ribosomal RNA) and morphology to define four new Apeuthes species from Malaysia, Thailand and Vietnam: A. fimbriatus, sp. nov., A. longeligulatus, sp. nov., A. pollex, sp. nov. and ?A. spininavis, sp. nov. The intraspecific COI sequence divergence of two Apeuthes species is 3–7% (mean: 5%) and the interspecific divergence of five species is 11–16% (mean: 13.7%). All members of the genus share unique male characters, viz the posterior gonopod telopodite with several dentate, serrate or tuberculate lamellae in a boat-like cavity or a boat-like cavity covered with spines. The delimitation of the four new species is supported by the congruence between mitochondrial DNA and morphological data. However, while the monophyly of Trigoniulinae is well supported, the relationships within this subfamily, and particularly among Apeuthes species, including the monophyly of Apeuthes, lack strong support. Therefore assignment of the four new species, and particularly of ?A. spininavis sp. nov., to the genus Apeuthes is tentative and awaits a comprehensive revision of the group. [ABSTRACT FROM AUTHOR]

The assassin snail genus Anentome is widespread in Southeast Asia, and is distributed all over the world via the aquarium trade. One species of genus Anentome, Anentome helena, is known to act as intermediate host of parasitic trematodes. This study investigates the taxonomic diversity of larval trematodes infecting A. helena and Anentome wykoffi in Thailand. Larval trematodes were identified by combining morphological and DNA sequence data (cytochrome c oxidase I and internal transcribed spacer 2). Species delimitation methods were used to explore larval trematode species boundaries. A total of 1107 specimens of Anentome sp. were collected from 25 localities in Thailand. Sixty-two specimens of A. helena (n = 33) and A. wykoffi (n = 29) were infected by zoogonid cercariae, heterophyid metacercariae and echinostome metacercariae, with an overall prevalence of 5.6% (62/1107) and population-level prevalences in the range of 0.0–22.3%. DNA sequence data confirmed that the larval trematodes belong to the families Echinostomatidae, Heterophyidae and Zoogonidae. As such, this is the first report of zoogonid cercariae and heterophyid metacercariae in A. helena, and echinostome metacercariae in A. wykoffi. Moreover, this study provides evidence of tentative species-level differentiation between Thai Echinostoma sp. and Cambodian Echinostoma mekongi, as well as within Echinostoma caproni, Echinostoma trivolvis and Echinostoma revolutum. [ABSTRACT FROM AUTHOR]

Biting midges of genus Culicoides (Diptera: Ceratopogonidae) are the vectors of several pathogenic arboviruses and parasites of humans and animals. Several reports have suggested that biting midges might be a potential vector of Leishmania parasites. In this study, we screened for Leishmania and Trypanosoma DNA in biting midges collected from near the home of a leishmaniasis patient in Lamphun province, northern Thailand by using UV-CDC light traps. The identification of biting midge species was based on morphological characters and confirmed using the Cytochrome C oxidase subunit I (COI) gene. The detection of Leishmania and Trypanosoma DNA was performed by amplifying the internal transcribed spacer 1 (ITS1) and small subunit ribosomal RNA (SSU rRNA) genes, respectively. All the amplified PCR amplicons were cloned and sequenced. The collected 223 biting midges belonged to seven species (Culicoides mahasarakhamense, C. guttifer, C. innoxius, C. sumatrae, C. huffi, C. oxystoma, and C. palpifer). The dominant species found in this study was C. mahasarakhamense (47.53%). Leishmania martiniquensis DNA was detected in three samples of 106 specimens of C. mahasarakhamense tested indicating a field infection rate of 2.83%, which is comparable to reported rates in local phlebotomines. Moreover, we also detected Trypanosoma sp. DNA in one sample of C. huffi. To our knowledge, this is the first molecular detection of L. martiniquensis in C. mahasarakhamense as well as the first detection of avian Trypanosoma in C. huffi. Blood meal analysis of engorged specimens of C. mahasarakhamense, C. guttifer, and C. huffi revealed that all specimens had fed on avian, however, further studies of the host ranges of Culicoides are needed to gain a better insight of potential vectors of emerging leishmaniasis. Clarification of the vectors of these parasites is also important to provide tools to establish effective disease prevention and control programs in Thailand. Author summary: Culicoides are known to be vectors of several pathogens, including arboviruses and parasites. However, other investigations have demonstrated that Culicoides could be involved in the transmission of Leishmania and Trypanosoma parasites. Our studies demonstrated the first detection of L. martiniquensis in Culicoides mahasarakhamense and Trypanosoma sp., which is closely related to avian Trypanosome in C. huffi from an endemic area of leishmaniasis in northern Thailand. The finding of this work suggested that Culicoides biting midges are suspected to be the potential vector of L. martiniquensis and Trypanosoma parasites in Thailand. [ABSTRACT FROM AUTHOR]

Cassava plantations in an area of 458 hectares spanning five provinces along the Thailand–Cambodia border were surveyed from October 2018 to July 2019 to determine the prevalence of cassava mosaic disease (CMD) caused by Sri Lankan cassava mosaic virus (SLCMV) in the region. CMD prevalence was 40% in the whole area and 80% in Prachinburi, 43% in Sakaeo, 37% in Burium, 25% in Surin, and 19% in Sisaket provinces. Disease incidence of CMD was highest 43.08% in Sakaeo, followed by 26.78% in Prachinburi, 7% in Burium, 2.58% in Surin, and 1.25% in Sisaket provinces. Disease severity of CMD symptoms was mild chlorosis to moderate mosaic (2–3). The greatest disease severity was recorded in Prachinburi and Sakaeo provinces. Asymptomatic plants were identified in Surin (12%), Prachinburi (5%), Sakaeo (0.2%), and Buriram (0.1%) by PCR analysis. Cassava cultivars CMR-89 and Huai Bong 80 were susceptible to CMD. In 95% of cases, the infection was transmitted by whiteflies (Bemisia tabaci), which were abundant in Sakaeo, Buriram, and Prachinburi but were sparse in Surin; their densities were highest in May and June 2019. Nucleotide sequencing of the mitochondrial cytochrome oxidase 1 (mtCO1) gene of whiteflies in Thailand revealed that it was similar to the mtCO1 gene of Asia II 1 whitefly. Furthermore, the AV1 gene of SLCMV—which encodes the capsid protein—showed 90% nucleotide identity with SLCMV. Phylogenetic analysis of completed nucleotide sequences of DNA-A and DNA-B components of the SLCMV genome determined by rolling circle amplification (RCA) indicated that they were similar to the nucleotide sequence of SLCMV isolates from Thailand, Vietnam, and Cambodia. These results provide important insights into the distribution, impact, and spread of CMD and SLCMV in Thailand. [ABSTRACT FROM AUTHOR]

Aim: The aim of this study was to characterize Leucocytozoon caulleryi from backyard chickens in Khon Kaen Province, Thailand. Materials and Methods: Tissue samples were collected from backyard chickens suspected to have leucocytozoonosis and subjected to histopathology examination. The BLAST Tool at NCBI GenBank (Basic Local Alignment Research Tools) (http://www.ncbi.nlm.nih.gov/BLAST) was used to identify the nucleotide sequence of the partial cytochrome c oxidase subunit I (cox I) gene. A Phylogenetic tree for analysis of L. caulleryi was constructed by using MEGA 7.0 software (https:// www.megasoftware.net/). Results: The necropsy results revealed the subcutaneous hemorrhages of pectoral muscles, multifocal hemorrhages of the thymus and pectoral muscles, hemorrhage of the proventriculus and peritoneal cavity, and megaloschizonts of the pancreas and intestine, including subcapsular hemorrhages of the liver. Microscopic examination revealed numerous megaloschizonts of Leucocytozoon spp. in the pectoral muscles, intestine, pancreas, and thymus. Molecular analysis of the partial cox I gene showed that the causal agent was closely related to L. caulleryi reported in Japan. Conclusion: From these results, L. caulleryi was determined to be the causal agent of leucocytozoonosis and was closely associated with L. caulleryi reported in Japan. [ABSTRACT FROM AUTHOR]

Background and Aim: Cercarial dermatitis or swimmer's itch is an allergic skin reaction caused by penetrating cercaria of animal blood flukes. It is considered as a zoonotic water-borne skin condition that is found globally. Among the schistosomatid trematodes, avian schistosomes are the most responsible for cercarial dermatitis. Very little is known regarding the occurrence of dermatitis-causing cercariae in Thailand. Therefore, the objective of this study was to preliminarily investigate the presence of larval blood fluke infection among local lymnaeidae snails in Phayao by the incorporation of morphological and molecular methods. Materials and Methods: Overall 500 Radix (Lymnaea) rubiginosa (Michelin, 1831) were collected from freshwater reservoirs near Phayao Lake in San Kwan village in Phayao, Thailand, from October to December 2020. The snails were examined for avian blood fluke infection by the cercarial shedding technique followed by morphological and molecular characterization. Results: Only one type of furcocercous cercaria was observed to emerge from six infected snails (1.2%). Our molecular analyses demonstrated that the emerging cercariae showed most similarity to either the 28S ribosomal RNA gene (28S rDNA) or cytochrome oxidase C subunit 1 gene (cox1 or COI) sequences to those of Trichobilharzia species. In addition, phylogenetic tree analyses of both loci revealed similar results; the emerging cercariae were consistently clustered together with Trichobilharzia regenti. Conclusion: Our results clearly confirmed that the detected furcocercous cercariae belonged to the genus Trichobilharzia and displayed the highest homology to T. regenti. This study provides important data on the occurrence of dermatitis causing cercariae infection among local lymnaeidae snails, encouraging effective management, and control measures for this zoonotic infectious disease. [ABSTRACT FROM AUTHOR]

Background: The malaria vector Anopheles minimus has been influenced by external stresses affecting the survival rate and vectorial capacity of the population. Since An. minimus habitats have continuously undergone ecological changes, this study aimed to determine the population genetic structure and the potential gene flow among the An. minimus populations in Thailand. Methods: Anopheles minimus was collected from five malaria transmission areas in Thailand using Centers for Disease Control and Prevention (CDC) light traps. Seventy-nine females from those populations were used as representative samples. The partial mitochondrial cytochrome c oxidase subunit I (COI), cytochrome c oxidase subunit II (COII) and cytochrome b (Cytb) gene sequences were amplified and analyzed to identify species and determine the current population genetic structure. For the past population, we determined the population genetic structure from the 60 deposited COII sequences in GenBank of An. minimus collected from Thailand 20 years ago. Results: The current populations of An. minimus were genetically divided into two lineages, A and B. Lineage A has high haplotype diversity under gene flow similar to the population in the past. Neutrality tests suggested population expansion of An. minimus, with the detection of abundant rare mutations in all populations, which tend to arise from negative selection. Conclusions: This study revealed that the population genetic structure of An. minimus lineage A was similar between the past and present populations, indicating high adaptability of the species. There was substantial gene flow between the eastern and western An. minimus populations without detection of significant gene flow barriers. [ABSTRACT FROM AUTHOR]

Cattle and buffaloes, popular protein sources worldwide, are intermediate hosts for several Sarcocystis species. These coccidian protozoans cause sarcocystosis resulting in subclinical and chronic infections in striated muscles by forming macrocysts or microcysts. In Thailand, Lao People's Democratic Republic, and Cambodia, Sarcocystis species have been reported, but molecular identification has been lacking. This study investigates the prevalence of infection, histo-morphology, and molecular identification of Sarcocystis species in hearts of cattle and buffalo sold in local markets. A phylogenetic tree inferred from a portion of the 18S ribosomal (r) RNA gene was used to identify the genus and species of Sarcocystis. The mitochondrial cytochrome c oxidase subunit 1 (cox-1 gene) was sequenced to confirm the species of host tissue. In Thailand, Sarcocystis was detected in 66.7% (14/21) of samples. In Lao People's Democratic Republic, 90% (9/10) of samples were infected and in Cambodia 100% (8/8). For the first time from these countries, we report Sarcocystis cruzi, Sarcocystis heydorni, and Sarcocystis levinei found in taurine cattle (Bos taurus) and water buffalo (Bubalus bubalis). Zoonotic protozoan transmission needs to be controlled by inspection activities by local health inspectors, and appropriate action is required at all points in the food chain by competent authorities to protect consumer health and prevent sarcocystosis in cattle and water buffaloes. [ABSTRACT FROM AUTHOR]

Background: Phlebotomine sand flies are vectors of Leishmania spp. At least 27 species of sand flies have been recorded in Thailand. Although human leishmaniasis cases in Thailand are mainly imported, autochthonous leishmaniasis has been increasingly reported in several regions of the country since 1999. Few studies have detected Leishmania infection in wild-caught sand flies, although these studies were carried out only in those areas reporting human leishmaniasis cases. The aim of this study was therefore to identity sand fly species and to investigate Leishmania infection across six provinces of Thailand. Methods: Species of wild-caught sand flies were initially identified based on morphological characters. However, problems identifying cryptic species complexes necessitated molecular identification using DNA barcoding in parallel with identification based on morphological characters. The wild-caught sand flies were pooled and the DNA isolated prior to the detection of Leishmania infection by a TaqMan real-time PCR assay. Results: A total of 4498 sand flies (1158 males and 3340 females) were caught by trapping in six provinces in four regions of Thailand. The sand flies were morphologically classified into eight species belonging to three genera (Sergentomyia, Phlebotomus and Idiophlebotomus). Sergentomyia iyengari was found at all collection sites and was the dominant species at most of these, followed in frequency by Sergentomyia barraudi and Phlebotomus stantoni, respectively. DNA barcodes generated from 68 sand flies allowed sorting into 14 distinct species with 25 operational taxonomic units, indicating a higher diversity (by 75%) than that based on morphological identification. Twelve barcoding sequences could not be assigned to any species for which cytochrome c oxidase subunit I sequences are available. All tested sand flies were negative for Leishmania DNA. Conclusions: Our results confirm the presence of several sand fly species in different provinces of Thailand, highlighting the importance of using DNA barcoding as a tool to study sand fly species diversity. While all female sand flies tested in this study were negative for Leishmania, the circulation of Leishmania spp. in the investigated areas cannot be ruled out. [ABSTRACT FROM AUTHOR]

In Thailand, there are at least ten species of freshwater snails of the family Bithyniidae, most of which can act as intermediate hosts of veterinary and medically important parasites. The genetic variation, geographical distribution, and taxonomic status of some species are obscure, including for Hydrobioides nassa. Thus, this study aims to explore the genetic variation and distribution of H. nassa in north, west, and central Thailand. We collected 264 specimens of H. nassa from 46 localities in 16 provinces using mitochondrial cytochrome c oxidase subunit I (COI) and 16S ribosomal DNA sequences to determine variation. Genetic diversity of H. nassa is relatively high, with 65 and 11 haplotypes of COI and 16S rDNA observed, respectively. The phylogenetic tree and haplotype network analyses classified H. nassa into three haplogroups (haplogroup I–III). Haplogroup I and II belonged to clade A, whereas haplogroup III belonged to clade B. Interestingly, haplogroup III or clade B contained the specimens from Yom river basin in Phrae province, which was the most genetically distinct. Thus, this study suggests that H. nassa in Thailand is a complex of phenotypically similar but genetically distinct species. [ABSTRACT FROM AUTHOR]

Fungus growing termites is a unique group of termites that can grow mutualistic fungi from Basidiomycota: Termitomyces inside their nest. This group of termites has an important ecological role as a major decomposer. However, comprehensive study of fungus-growing termites is limited in Thailand and the complex termites-fungus relationship is a challenging issue to study. This research was aimed to identify the species of termites and fungal symbionts using DNA barcoding, to compare the molecular results with morphological identification, and to gain insights into the co-evolutionary relationship by constructing co-cladogenesis analysis from the termites and the fungal symbionts. Four termite species (Macrotermes annandalei, Odontotermes feae, Ancistrotermes pakistanicus and Ancistrotermes sp.) from four Sakaerat and two Chulabhorn colonies were identified based on the BLAST-N of cytochrome oxidase sub-unit I (COI) sequence and morphological identification. The partial COI sequence of O. feae identified in this study is the first record of this species deposited into the NCBI nucleotide database. Molecular identification of the fungi were conducted by using nuclear ribosomal internal transcribed spacer (ITS) region. The results showed that those six termite colonies grew specifically the fungal genus Termitomyces. Three genetic groups of Termitomyces i.e. Group 1, Group 2 (T. cylindricus), and Group 6, were present among the six colonies. Molecular identification was confidently supported by morphological identification of termite samples, at least up to the genus level. High host-symbionts specificity is considerably observed within some clades on paired termite-fungus phylogeny. However, more sampling is needed for the more confident result. [ABSTRACT FROM AUTHOR]

The population structure of a fruit fly, Zeugodacus tau, was studied by a Single-Strand Conformation Polymorphism (SSCP) technique. Twenty haplotypes of Cytochrome Oxidase unit I (COI) sequence were found in flies collected from Southern Thailand. The phylogenetic tree and haplotype network revealed gene flow across a large geographic range. With the aid of winds, their gene flow diminished population structure. The population size of Z. tau in Southern Thailand seemed to be large and stable, but the other populations in some locations had experienced a bottleneck effect, leading to local genetic differentiation. Fruit flies from the eastern areas had large effective population sizes, whereas the populations from other areas were smaller. This pattern matched the ecological niche centroid model, in which fruit flies disperse from high population areas to lower ones. [ABSTRACT FROM AUTHOR]

Ornate threadfin bream (Nemipterus hexodon) is an economically important fishery species in Southeast Asia. In Thailand, N. hexodon decreased dramatically due to overexploitation for commercial purposes. To construct an effective sustainable management plan, genetic information is necessary. Thus, in our study, the population genetic structure and demographic history of N. hexodon were investigated using 419 bp of the mitochondrial DNA sequence in cytochrome oxidase subunit I gene (mtDNA COI). A total of 142 samples was collected from nine localities in the Gulf of Thailand (Chonburi, Samut Songkhram, Surat Thani, Nakhon Si Thammarat, Songkhla), and the Andaman Sea (Satun, Trang, Krabi, Phang Nga). Fourteen polymorphic sites defined 18 haplotypes, revealing a high haplotype diversity and low nucleotide diversity among nine localities. The analysis of molecular variance (AMOVA) analysis, pairwise FST, and minimum spanning network result revealed that the genetic structure of N. hexodon was separated into two populations: the Gulf of Thailand and the Andaman Sea population. The genetic structure of N. hexodon can be explained by a disruption of gene flow from the geographic barrier and the Pleistocene isolation of the marine basin hypothesis. Neutrality tests, Bayesian skyline analysis, mismatch distribution, and the estimated values of population growth suggested that N. hexodon had experienced a population expansion. The genetic information would certainly help us gain insight into the population genetic structure of N. hexodon living on the coast of Thailand. [ABSTRACT FROM AUTHOR]

The endemic terrestrial Crown Snail from Southern Thailand formerly in Macrochlamys Gray, 1847 is now described as Taphrenalla Pholyotha & Panha gen. nov., based on comparative morphology and molecular data. Overall, eleven Taphrenalla species are now recognized, including two nominal species: T. asamurai and T. diadema. A total of nine new species are proposed: T. alba sp. nov., T. conformis sp. nov., T. corona sp. nov., T. dalli sp. nov., T. incilis sp. nov., T. macrosulcata sp. nov., T. parversa sp. nov., T. pygmaea sp. nov. and T. zemia sp. nov. The molecular phylogeny constructed from the mitochondrial cytochrome oxidase c subunit I (COI) and 16S rRNA gene fragments plus the nuclear 28S rDNA gene fragments revealed that Taphrenalla gen. nov. is monophyletic with a well-supported clade. The diagnostic characters of Taphrenalla gen. nov. are the shell sculpture with several radial grooves, body with well-developed colourful stripes running from the head to tail, and genitalia similar to Macrochamys but with an un-coiling epiphallic caecum. The spermatophore has one or two spines near the sperm sac and a spineless tail. [ABSTRACT FROM AUTHOR]

The wedge clam (Donax scortum) is an important commercial species in Thailand. In our study, we performed an analysis of the genetic structure of the wedge clam population in the Andaman Sea along the coast of Thailand to provide information for developing a management plan. Partial sequences of the mitochondrial DNA cytochrome oxidase subunit I gene (mtDNA COI) with a size of 426 base pairs were investigated in 115 individuals, collected from four sampling sites within fishing grounds along the Andaman Sea coast of Thailand. Thirty-two haplotypes were observed. Haplotype diversity and nucleotide diversity were 0.887 and 0.00541, respectively. The population genetic analysis shows a lack of genetic structure, possibly caused by a high level of gene flow due to the high dispersal ability of wedge clam larvae. Demographic history tests reveal that the wedge clam population experienced expansion approximately 1,000 years ago. The results of our study can be used to guide the management of the wedge clam in this part of the Andaman Sea to maintain genetic diversity. [ABSTRACT FROM AUTHOR]

Ampullariidae include the largest of all freshwater snails and are of ecological, evolutionary and anthropogenic importance in Southeast Asia (SEA). Native ampullariids belonging to the genus Pila face various threats but are understudied, with their species taxonomy being confused and data on their distributions being scarce. We provide a comprehensive update on the nomenclature, status and distribution of Pila species in Thailand, based on DNA barcoding and geometric morphometric analysis of recently collected material. We confirm that at least five Pila species are extant in Thailand: Pila virescens, P. celebensis, P. turbinis, P. gracilis and P. pesmei. Pila celebensis , which has distinctive egg masses among all the known Southeast Asian Pila , appears to be sister to a clade comprising other SEA and some African Pila. Our results suggest that Pila may have dispersed into SEA on at least two separate occasions. Two singletons collected from northern and eastern Thailand may constitute separate species, but this requires further study. Intraspecific diversity of the mitochondrial cytochrome c oxidase subunit I gene was relatively high for P. celebensis, P. gracilis and P. pesmei (maximum uncorrected p-distances varied from 7% to 9%), and may include cryptic species. Conversely, P. virescens showed low intraspecific p-distances (c. 0%) among clades collected from different localities. This strongly suggests that introductions by humans may be the major cause of this pattern, and our own observations—we found that this species is being cultured and is commonly sold in markets for human consumption—are consistent with this. Pila turbinis was the rarest species, with live snails being collected from only two localities. Throughout Thailand, invasive confamilial Pomacea species appear to be replacing native Pila species, particularly in the Chao Phraya basin. While Thai Pila exhibit surprisingly high genetic diversity, with cryptic species likely being present, widespread invasive snails pose a major threat to their survival and urgent conservation action is needed. [ABSTRACT FROM AUTHOR]