A recently described species of black fly, Simulium wayani Takaoka and Chen, from the island of Timor was chromosomally mapped to provide insights into its evolutionary and biogeographic history. The morphologically based species status of S. wayani is supported by a suite of fixed chromosomal rearrangements and unique sex chromosomes derived primarily from a large pool of polymorphisms in the S. ornatipes complex in Australia. The banding patterns of its polytene chromosomes indicate that S. wayani is closely related to a pair of homosequential cryptic species (S. norfolkense Dumbleton and S. ornatipes cytoform A2) in the S. ornatipes Skuse complex on mainland Australia; all three species uniquely share the same amplified band in their chromosomal complement. The low level of polymorphism and heterozygosity in S. wayani, relative to Australian populations of the S. ornatipes complex, suggests few colonization events from the larger land mass.
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.
Three new species of black flies from Thailand, Simulium wangkwaiense, S. tadtonense and S. maeklongkeense, are described based on their adults, pupal exuviae and cocoons. All three new species are assigned to the Simulium (Simulium) striatum species-group, bringing its total number in Thailand to seven. Simulium wangkwaiense sp. nov. is the species formerly called S. quinquestriatum in Thailand. Certain male and pupal morphological characteristics are shown to separate all seven Thai species of this species-group. All of the three new species have been analyzed genetically for their phylogenetic relationships, with three known related species (except for S. thailandicum), by using the fast-evolving nuclear big zinc finger (BZF) gene.
A new black fly species, Simulium (Gomphostilbia) kiewlomense, is described from females, males, pupae and mature larvae in Thailand. This new species is placed in the S. asakoae species-group and is characterized by having a combination of the elongate female sensory vesicle, widened male hind basitarsus, which is much wider than the hind femur, small pupal terminal hooks, and light greenish larval abdominal segments 1-3. Taxonomic notes are given to separate this new species from other related species. A DNA analysis using the COI gene shows that this new species has two genoforms with 1.21% difference. This is the 28th species of the S. asakoae species-group in Thailand, strengthening the evidence for high species diversity of this species-group.
Most female black flies in the genus Simulium are blood-sucking flies and they can cause various parasitic diseases in human and animal. A total of 94 species of black flies have been reported in Malaysia, however, their biting behavior and role as vector of infectious agents remain understudied. To fill in this knowledge gap, we attempted to survey adult black flies from field populations in Peninsular Malaysia. In a survey carried out in 2017 at Tasik Kenyir, Terengganu, three females were caught while attracted and landed on human skin. Further morphological and molecular analyses showed that the specimens were identical to Simulium (Gomphostilbia) aziruni Takaoka, Hashim & Chen of the Simulium gombakense species-group. This is the first report on a black fly species attracted to human in Malaysia which serves as a steppingstone towards in-depth studies for black flies in this region.
Larvae and adults reared from pupae of Simulium (Gomphostilbia) gombakense Takaoka & Davies from eight streams in five provinces of Thailand were genetically analyzed by using cytochrome c oxidase subunit 1 sequences. The material was composed of four lineages, of which lineages 2, 3 and 4 are genetically closely related to one another, but more distantly related to lineage 1, which is equivalent to typical S. (G.) gombakense from Peninsular Malaysia. Lineages 2, 3 and 4, which are morphologically indistinguishable, are designated as genoforms A, B and C within a species complex of S. (G.) paiense sp. nov., which is described based on specimens of lineage 2 (=genoform A). Simulium (G.) paiense sp. nov. is similar to S. (G.) gombakense from Peninsular Malaysia but appears to be barely distinguished by the relative length of the female fore basitarsus. Simulium (G.) thuathienense Takaoka & Sofian-Azirun is newly recorded from Thailand and its male is described for the first time. The female of S. (G.) gombakense is briefly described based on females reared from pupae collected from Peninsular Malaysia.
A black fly species of the Simulium feuerborni species-group of Simulium (Nevermannia) from Cameron Highland, Peninsular Malaysia, previously regarded as S. feuerborni Edwards, originally described from East Java, is described as Simulium pairoti sp. nov. based on complete life stages. High intraspecific variations in the arrangement of the six pupal gill filaments, length of the stalk of the ventral paired filaments, and length of the anterodorsal projection of the cocoon, are noted in this species. This new species is readily distinguished from its congeners by having the characters of male genitalia with simple lamellate ventral plate, short inwardly-twisted styles, several parameral hooks, and a simple narrow median sclerite. Morphological data reported herein plus the chromosomal and molecular data presented elsewhere support S. pairoti as a novel pseudocryptic species.
In recent decades, the numbers of cryptic taxa have increased significantly with current progress in DNA barcoding, yet, most of these cryptic taxa have not been formally named and recognized as valid species. To address this issue, we provide a guide for applying the procedure of describing new cryptic species in the family Simuliidae. Simulium (Simulium) vanluni from Pahang, Peninsular Malaysia, previously treated as S. nobile De Meijere, is described as a new species by using an integrated morpho-taxonomical and genetic approach. This new species is morphologically identical to S. nobile from Java and S. kiuliense Smart & Clifford from Borneo, but their distinctiveness is supported by an expanded multigene phylogeny analysis.
Surveys of pupae and larvae of black flies were carried out in Bali, Lombok, Sumbawa and Flores in the Lesser Sunda Archipelago, Indonesia, where 10 species were known. A total of 14 simuliid species including four new species and five new records of the genus Simulium were collected, bringing the number of species from the Lesser Sunda Archipelago to 19. They are classified into four subgenera: two in Nevermannia, nine in Gomphostilbia, seven in Simulium and one in Wallacellum. One of four new species, Simulium (Simulium) baliense, is described based on females, males, pupae and larvae from Bali and Lombok. This new species, which is placed in the Simulium striatum species-group of the subgenus Simulium, is closely related to S. (S.) argyrocinctum De Meijere from Java and Sumatra, but it is distinguished from the latter species by the smaller number of the male enlarged upper-eye facets and larval abdomen lacking dorsal pairs of conical protuberances. The distribution record of S. (S.) upikae Takaoka & Davies from Flores is corrected as that of S. (S.) eximium De Meijere. Some aberrant characters of the pupal gill filaments of S. (G.) atratum De Meijere, S. (G.) floresense Takaoka, Hadi & Sigit and S. (G.) sunapii Takaoka, Sofian-Azirun & Suana are illustrated. Characteristics of the fauna of black flies in this archipelago are briefly noted. Keys to all 19 species are provided for females, males, pupae and larvae.
Simulium pumatense sp. nov. is described from Vietnam, and is placed in the Simulium feuerborni species-group of the subgenus Simulium (Nevermannia) Enderlein. Its morphological characteristics include the relatively smaller numbers of the following three numerical features: inner teeth of the female mandible (15-18), minute conical processes (16) on the female cibarium, and male upper-eye facets (in 15 vertical columns and 16 horizontal rows). Keys are constructed to distinguish this species from four species of the same group in Vietnam. Our molecular analysis of the DNA barcoding COI gene shows that this species is most closely related to cytoform A of the S. feuerborni complex from Thailand.
The population genetic structures of S. vanluni, S. cheongi and S. jeffreyi were determined from mitochondria-encoded sequences of cytochrome c oxidase subunits I (COI) across different states in Peninsular Malaysia. High levels of genetic diversity and genetic differentiation were observed among three species. All three species revealed an intermediate level of gene flow among the populations. Negative values of Fu's Fs and low values of Raggedness index supported the hypothesis of population expansion in S. vanluni, S. cheongi and S. jeffreyi.
Simulium (Gomphostilbia) aziruni Takaoka, Hashim & Chen was described initially based only on a pupa and a mature larva collected from Peninsular Malaysia. Herein, we describe the morphological characters of the female of S. aziruni for the first time. It resembles those of the other members of the Simulium gombakense species-group by the genital fork with a distinct projection directed medioposteriorly from each arm and claw with a large basal tooth. Cytochrome c oxidase I (COI) barcoding analysis indicates that S. aziruni is the sister species of S. maleewongae, but both are distantly separated by a genetic distance of 4.9%.
A new species of black fly, Simulium (Gomphostilbia) isanense, is described based on females, males, pupae and mature larvae from Thailand. This new species is placed in the Simulium epistum species-group of the subgenus Gomphostilbia Enderlein. It is characterized by the pupal gill with eight filaments arranged as 3+3+2 from dorsal to ventral, of which an inner filament of the ventral pair is slightly longer than its counter filament. Taxonomic notes are provided to distinguish this new species from S. (G.) angulistylum Takaoka & Davies from Peninsular Malaysia, and three other related species. The difference between this new species and S. (G.) angulistylum is supported by genetic distances using the mitochondrial COI gene.
Five new species of black flies, Simulium (Gomphostilbia) chiangraiense, S. (G.) huaikaeoense, S. (G.) khaokhoense, S. (G.) maeklangense, and S. (G.) pamiangense, are described based on adults, pupae and mature larvae from Thailand. These five species are similar to one another and are placed in the S. decuplum subgroup of the S. batoense species-group by having the male fore coxae darkened, ventral plate flat and transverse; pupal gill with 10 short filaments; and larval postgenal cleft deep approaching or reaching the posterior margin of the hypostoma. Simulium (G.) chiangraiense sp. nov. and S. (G.) pamiangense sp. nov. differ from the three other new species by the much longer pupal gill filaments and darkened dorsal surface of abdominal segments 1 and 2. Taxonomic notes are provided to distinguish these five new species from five other related species. The phylogenetic positions of these new species in the S. decuplum subgroup are presented based on the mitochondrial COI gene. Simulium (G.) pamiangense sp. nov. and S. (G.) huaikaeoense sp. nov. are highly similar to each other genetically, showing a sister relationship, though they are clearly different morphologically. On the contrary, S. (G.) khaokhoense sp. nov. is distantly positioned from S. (G.) huaikaeoense nov., though it is almost indistinguishable morphologically from the latter.
Simulium feuerborni Edwards is geographically widespread in Southeast Asia. Previous cytogenetic study in Thailand revealed that this species is a species complex composed of two cytoforms (A and B). In this study, we cytologically examined specimens obtained from the Cameron Highlands, Malaysia, and Puncak, Java, Indonesia. The results revealed two additional cytoforms (C and D) of S. feuerborni. Specimens from Malaysia represent cytoform C, differentiated from other cytoforms by a fixed chromosome inversion on the long arm of chromosome III (IIIL-5). High frequencies of the B chromosome (33-83%) were also observed in this cytoform. Specimens from Indonesia represent the cytoform D. This cytoform is differentiated from others by a fixed chromosome inversion difference on the long arm of chromosome II (IIL-4). Mitochondrial DNA sequences support genetic differentiation among cytoforms A, B, and C. The pairwise F(ST) values among these cytoforms were highly significantly consistent with the divergent lineages of the cytoforms in a median-joining haplotype network. However, a lack of the sympatric populations prevented us from testing the species status of the cytoforms.
We access the molecular diversity of the black fly Simulium nobile De Mejiere, using the universal cytochrome c oxidase subunit I (COI) barcoding gene, across its distributional range in Southeast Asia. Our phylogenetic analyses recovered three well-supported mitochondrial lineages of S. nobile, suggesting the presence of cryptic species. Lineage A is composed of a population from Sabah, East Malaysia (Borneo); lineage B represents the type population from Java, Indonesia; and lineage C includes populations from the mainland of Southeast Asia (Peninsular Malaysia and Thailand). The genetic variation of lineage C on the mainland is greater than that of lineages A and B on the islands of Borneo and Java. Our study highlights the value of a molecular approach in assessing species status of simuliids in geographically distinct regions.
Simulium (Gomphostilbia) yvonneae sp. nov. is described based on adults, pupae, and mature larvae from Vietnam. This new species belongs to the Simulium duolongum subgroup in the S. batoense species-group of the subgenus Gomphostilbia Enderlein. It is distinguished by having a relatively larger number of male upper-eye facets in 16 vertical columns and 16 horizontal rows and a pupal gill with eight filaments arranged as 3+(1+2)+2 from dorsal to ventral, of which two filaments of the ventral pair are 1.8 times as long as the longest filament of the middle and dorsal triplets. Morphological comparisons are made to distinguish this new species from all 22 related species. The genetic distinctiveness of this new species in the S. duolongum subgroup is also presented based on the DNA barcoding COI gene.
Seven populations of Simulium parahiyangum Takaoka & Sigit (Diptera: Simuliidae), a geographically widespread nominal species of black fly in Southeast Asia, were morphologically and molecularly studied. Three morphoforms based on male and pupal morphological features, and two primary lineages based on the COI gene sequence analysis were recognized. Morphoform 1 (lineage 1) from Sarawak, Malaysia, is identified as S. parahiyangum sensu stricto and morphoform 2 (lineage 2) from Thailand and Vietnam, and morphoform 3 (lineage 1) from Peninsular Malaysia are each regarded as distinct species, although morphoform 3 is partially homosequential for the COI gene with morphoform 1. Morphoforms 2 and 3 are described as Simulium ngaoense sp. nov. and Simulium sazalyi sp. nov., respectively. Overall, S. parahiyangum is not a single geographic generalist but a composite of multiple species.
Simulium undecimum sp. nov. is described from Thailand. This new species is assigned to the Simulium multistriatum species-group, one of the 20 species-groups of the subgenus Simulium in the Oriental Region. It is characterized by the female cibarium with minute processes, male ventral plate with a narrow body having two vertical rows of distinct teeth on the posterior surface and without setae on the anterior and lateral surfaces, pupal gill with eight short filaments decreasing in length from dorsal to ventral, and divergent at an angle of around 90 degrees when viewed laterally, spine-combs only on abdominal segments 7 and 8, and cocoon wall-pocket shaped with anterolateral windows. Taxonomic notes to separate this new species from related species in Thailand and other countries are given. This new species is the 11th nominal member of this species-group recorded in Thailand. An analysis of the COI gene sequences shows that it is most closely related with S. malayense Takaoka & Davies (cytoform A) from Thailand but they are distantly separated by 3.01-8.87%.
A man-biting black fly species so far regarded as Simulium rufibasis Brunetti in Thailand was morphologically analyzed. It was found to be almost indistinguishable in the female from S. rufibasis sensu stricto but distinguishable in the male by the slender fore basitarsus and abdominal segments 2, 6, and 7 each with a pair of shiny dorsolateral patches, and in the pupa by the absence of spine-combs on abdominal segment 7 and terminal hooks on segment 9. It is described as a new species, Simulium tenebrosum. Taxonomic notes are given to separate it from all 11 known species of the S. rufibasis subgroup of the Simulium (Simulium) tuberosum species-group. An analysis of the COI gene sequences shows that this new species is distantly separated from the closely related species.