Population Genetic Structure and Population History of the Biting Midge Culicoides mahasarakhamense (Diptera: Ceratopogonidae).

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.

Population structure and population history of the black fly Simulium chumpornense (Diptera: Simuliidae) from Thailand.

Understanding genetic structure and diversity of insect vectors is crucial for disease epidemiology. In this study, mitochondrial cytochrome c oxidase I sequences were used to infer genetic diversity, genetic structure and population history of the black fly, Simulium chumpornense Takaoka and Kuvangkadilok, a suspected vector of blood protozoa of the genus Leucocytozoon and Trypanosoma. High intraspecific genetic divergence (max. 3.76%) was found among 142 specimens obtained from 19 locations across Thailand. A median joining network revealed two genetic lineages (A and B) that were geographically associated. Lineage A is representative of central and northeastern regions. Lineage B represents specimens from diverse locations in northern, western, and southern Thailand, including the type locality. Mismatch distribution and the neutrality tests provided signals of past population expansions in both lineages. The expansion time dating back to the end of last glaciations at 12,000 - 15,000 years ago is possibly related to increasing of precipitation at the end of last glacial period. Despite recent population history, population pairwise FST analysis revealed that almost all population comparisons were genetically significantly different. The high level of genetic structuring is possibly a result of historical isolation of the population that survived in different refugia sites during the dry conditions during glaciations.

Molecular detection of avian haemosporidian parasites in biting midges (Diptera: Ceratopogonidae) from Thailand.

Culicoides biting midges are vectors of many haemosporidian parasite species. These parasites are found in several wild and domestic avian species in Thailand but knowledge of the insect vectors is very limited. In this study, a molecular approach was used to detect haemosporidian parasites in six biting midge species in Thailand. A total of fifteen cytochrome b haplotypes of three haemosporidian parasites were detected from 1,165 specimens of six different Culicoides species. Comparisons of these sequences with those recorded in a public database revealed that they were comprised sequences of three species, Leucocytozoon sp., Plasmodium juxtanucleare and P. gallinaceum. All of these haemosporidian parasite species were detected in Culicoides mahasarakhamense Pramual, Jomkumsing, Piraonapicha, & Jumpato while P. juxtanucleare was also detected in C. huffi Causey and C. guttifer Meijere and Leucocytozoon sp. were also detected in C. guttifer. All of these parasites are commonly found in chickens in agreement with information that these biting midge species will bite chickens. Detection DNA of Leucocytozoon sp. in biting midges reported here provides the second record of this parasitic genus infecting Culicoides. This study also provides the first records of P. juxtanucleare and P. gallinaceum in biting midge species. Further investigation is required to determine whether Culicoides biting midge species are competent vectors of these parasites.

DNA barcoding of tropical black flies (Diptera: Simuliidae) in Thailand: One decade of progress.

Black flies (Diptera: Simuliidae) are important blood sucking insects because they are the vectors of disease agents transmitted to human and other animals. Rapid and correct species identifications are necessary for all aspects of the study of black flies. DNA barcodes based on mitochondrial cytochrome c oxidase I (COI) have been effectively used for the determination of black fly species. However, the success of this method requires a large and reliable COI sequence library. In this study, 171 DNA barcoding sequences from 17 black fly species were added to NCBI GenBank database, six of these species were reported for the first time. Efficacy of DNA barcodes for species identification was examined using 1,286 sequences representing 89 nominal species of black flies in Thailand. A considerable level of success (90%) was achieved but efficiency of COI sequences for species identification was very low in the following species-groups; Simulium asakoae, S. feuerborni, S. multistriatum and S. striatum. Incomplete lineage sorting or inadequate variation of this genetic marker for differentiation of recently diverged species are the more likely explanations, and thus, more variable genetic markers are needed. Other reasons for unsuccessful DNA barcoding are imperfect taxonomy and the misidentification of sources of reference sequences. Because many new black fly species in Thailand were described recently, a reassessment of the COI sequences reported previously is necessary.

Integrative taxonomy uncovers a new Culicoides (Diptera: Ceratopogonidae) biting midge species from Thailand.

Fully understanding biodiversity often requires an integrated approach especially for small insects because species diagnostic morphological characters are limited. In this study, morphological characters and DNA barcodes were used to examine previously recognized genetically divergent lineages detected in the biting midge, Culicoides arakawae (Arakawa), from Thailand. Morphological examinations revealed that specimens belonging to one lineage are morphologically different from C. arakawae in shape of the paramere in males, and in the leg color pattern of both sexes. Therefore, a formal description is provided for this new species, Culicoides mahasarakhamense sp. nov. Based on morphological characters including a large and shallow palpal pit, one sac like spermatheca and male with parameres bent at base with large basal knob, the new species was assigned into the subgenus Meijerehelea Wirth and Hubert. Morphological differentiation including wing pattern and shape of spermatheca of the new species are discussed and compared with other members of this subgenus. Mitochondrial cytochrome c oxidase I sequence analysis indicated that this new species is different from other members of the subgenus Meijerehelea with minimum interspecific genetic divergence of 3.92%. Automatic Barcode Gap Discovery species delimitation analysis also supported the recognition of a new species. Phylogenetic analyses revealed that the new species is closely related to C. arakawae, consistent with morphological similarity of these species. Results of this study highlight the necessity of using integrated approach for Culicoides taxonomy.

Genetic variation, DNA barcoding and blood meal identification of Culicoides Latreille biting midges (Diptera: Ceratopogonidae) in Thailand.

Biting midges of the genus Culicoides Latreille are blood sucking insects of medical and veterinary importance. Many species are vectors of disease agents transmitted to humans and other animals. Therefore, rapid and accurate species identification is essential for appreciation of all aspects of these insects. In this study, DNA barcode efficacy and molecular identification of host blood sources were examined in biting midges from Thailand. A total of 203 barcoding sequences were obtained from 16 Culicoides taxa. Intraspecific genetic divergence varied from 0.28% to 9.90% for specimens collected in Thailand. Despite this high level of genetic variation, DNA barcode identifications in the Barcoding of Life Data System had a considerable success rate (90%). Phylogenetic analyses and distance-based species delimitation methods indicated the possibility of cryptic species in four taxa, namely, Culicoides actoni Smit, C. arakawae Arakawa, C. huffi Causey and C. jacobsoni Macfie. Further investigations will be required to examine the species status of these lineages. Host blood meal identifications from 42 blood engorged females of 10 Culicoides taxa revealed three animal hosts: chicken, cattle and buffalo. Most of this information agrees with previous knowledge but this is the first report of C. actoni, C. fulvus and C. huffi feeding on chicken.

Who is biting you? DNA barcodes reveal cryptic diversity in human-biting black flies (Diptera: Simuliidae).

Black flies (Simuliidae) are important biting insects and vectors of diseases agents of humans and livestock. Thus, understanding the taxonomy and biodiversity of these insects is crucial for control and management of these diseases. In this study, we used mitochondrial cytochrome c oxidase I sequences to examine genetic diversity of three human-biting and possible vector black fly taxa; the Simulium asakoae species-complex, S. chamlongi and S. nigrogilvum. High levels of genetic diversity (>3.5% intraspecific genetic divergence) were found in all three taxa. Phylogenetic analyses indicated that the S. asakoae complex can be divided into seven groups with the largest group consisting of specimens from Thailand, Malaysia and Myanmar. This group most likely represents true S. asakoae. The remaining haplotypes formed groups with conspecific haplotypes or with other closely related species. Among these groups, one including S. monglaense and another including S. myanmarense suggest that certain specimens identified as S. asakoae most likely belong to those species. Therefore, they constitute new locality records for Thailand and also represent new records of anthropophily. Members of S. chamlongi are not monophyletic as its clade also included S. hackeri. A median joining network revealed strong geographic associations of the haplotypes of S. nigrogilvum suggesting limitation of gene flow. Because this species occurs mainly in high elevation habitats, low land areas could present a barrier to gene flow.