DOI: 10.3897/asp.84.e176547

Authors: Yang Liu, Ruoqian Sun, Cong Li, Ruyue Zhang, Liming Wang, Ding Yang, Yuyu Wang

Abstract: Abstract Mosquitoes rank among the most deadly organisms worldwide, facilitating >700,000 human deaths annually through transmission of vector-borne pathogens. Culex are famous as vectors of multiple pathogens affecting both animals and humans. This study presents the first mitogenome sequencing and comparative analysis of seven species within Culex. Our findings demonstrated conserved structural features and nucleotide composition across the mitogenomes of these species. This study performed phylogenetic analysis of Culex based on mitochondrial genome data under both homogeneous and heterogeneous models separately, and estimated the divergence times. Phylogenetic analyses revealed that Culex is paraphyletic, with Lutzia nested within it. Both Cx. (Neoculex) and Cx. (Culex) were non-monophyletic. The two species of Cx. (Neoculex) were placed in separate lineages, with Cx. fergusoni as the sister group to all other Culex. Meanwhile, Cx. (Culex) was rendered paraphyletic by the inclusion of Cx. (Culiciomyia) and Cx. (Oculeomyia) within its clade. Divergence time estimation placed the basal split of Culicidae in Late Triassic, followed by the Culicinae-Anophelinae divergence in Late Jurassic (~147 Mya), with all speciation events within Culex postdating these splits and clustering in Neogene. This study provides a fundamental basis for understanding the mitogenomic architecture and phylogenetic relationships within the genus Culex, and also establishes a theoretical foundation for transmission mechanisms and control strategies of common mosquito-borne diseases.

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DOI: 10.3897/asp.82.e111985

Authors: Carol-Anne Villeneuve, Louwrens P. Snyman, Emily J. Jenkins, Nicolas Lecomte, Isabelle Dusfour, Patrick A. Leighton

Abstract: Abstract Arctic ecosystems face increasing risks from vector-borne diseases due to climate-driven shifts in disease patterns and vector distribution. However, species identification challenges impact vector-borne disease surveillance, necessitates accurate identification. Aedes species are predominant among Arctic mosquitoes and pose health risks, with some species potentially carrying Jamestown Canyon and Snowshoe hare viruses. However, identifying Aedes species is challenging, especially under Arctic conditions and with complex adult traits. This study assessed the suitability of DNA barcoding (COI and ITS2 regions) and morphological characteristics for the identification of Arctic black-legged Aedes. It also aimed to evaluate the reliability of publicly available sequences. Our analysis focused on Aedes impiger, Aedes nigripes, and two species from the Punctor subgroup – Aedes hexodontus and Aedes punctor. In our study, the COI barcoding region distinguished Ae. impiger and Ae. nigripes but not within the species of the Punctor subgroup. In addition, the ITS2 barcoding region did not differentiate the species. When we evaluated GenBank and BOLD sequences, we found issues of under-representation and misidentifications, particularly within the Punctor subgroup. Based on these results, we recommend addressing identification difficulties, particularly within the Punctor subgroup, and advocate for more comprehensive morphological and molecular identification strategies. Integrating morphology and DNA barcoding holds promise for robust disease surveillance in Arctic regions, yet challenges persist, especially in complex species groups like the Punctor subgroup. Tackling these issues is pivotal to ensuring accurate vector status determination and reliable disease risk assessments in a rapidly changing Arctic ecosystem.

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DOI: 10.3897/asp.82.e109995

Authors: Daubian Santos, Guilherme Cunha Ribeiro

Abstract: Abstract Eloeophila Rondani, 1856 and Idioptera Macquart, 1834 are two genera of the family Limoniidae (Diptera) distinguished by the presence of a supernumerary cross-vein m-cu. Although these genera were previously combined, there has been a lack of phylogenetic tests to investigate the evolutionary relationship between them. In this study, we conducted a cladistic analysis that indicates that Idioptera form a clade within Eloeophila, and therefore the two genera should be synonymized under Idioptera. Consequently, 87 species of Eloeophila are transferred to Idioptera.

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DOI: 10.3897/asp.81.e104426

Authors: Noor Izwan-Anas, Van Lun Low, Zubaidah Ya’cob, Emmanuel Y. Lourdes, Mohamad Rasul Abdullah Halim, Mohd Sofian-Azirun, Hiroyuki Takaoka, Peter H. Adler

Abstract: Black flies play a prominent role in public health and the epidemiology of parasitic diseases of humans, domesticated and wild animals. Correct identification and a comprehensive survey are required to identify vector and pest species and thus understand their biological attributes which play a vital role in the monitoring program. DNA barcoding is an established molecular tool that provides rapid and accurate species identification. Our study strengthens the molecular database for black flies in Malaysia by adding 59 cytochrome c oxidase I sequences for 22 species, of which 14 are included for the first time. These sequences, combined with those in public databases, represent a total of 338 sequences for 52 Malaysian species, nearly 50% of which were collected from type localities. At the subgeneric level, barcode gap analysis most accurately identified species in the subgenus Nevermannia (92%), followed by Simulium s. l. (91%), and Gomphostilbia (81%). The remaining sequences were ambiguous and could not be distinguished from those of nearest neighbour species due to an overlap in genetic divergence and low genetic diversity, especially between insular species. Tree analyses indicate that certain species had incomplete lineage sorting and low mitochondrial signals. Possible cryptic species were indicated in the Simulium (Gomphostilbia) batoense and S. (G.) epistum species groups. Species delimitations were consistent with morphological identifications except in large species groups such as the S. (G.) asakoae, S. (G.) batoense, S. (G.) epistum, and S. (Simulium) melanopus groups. The use of type specimens or specimens collected from type localities (topotypes) in barcoding is strongly recommended for reference sequences to increase the reliability of the molecular database.

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DOI: 10.3897/asp.81.e97946

Authors: Zehui Kang, Yuanyuan Xu, Guoquan Wang, Ding Yang, Xiao Zhang

Abstract: Limoniidae, the most speciose family in the superfamily Tipuloidea, consists of four subfamilies and more than 11,000 species. However, mitochondrial (mt) genome sequences, which have been widely used for phylogenetic study, are available for only 11 species across three subfamilies. Thus, a larger variety of mt genome sequences in Limoniidae are required to improve our understanding of tipuloid phylogeny and genomic evolution. Here we present mt genomes of Elephantomyia (Elephantomyia) inulta Alexander, 1938 and Helius (Helius) pluto Alexander, 1932, representing the first mt genomes of the tribe Elephantomyiini (Limoniidae). The two mt genomes are typical circular DNA molecules and show similar gene order, nucleotide composition and codon usage. Standard ATN start and TAR stop codons are present in most protein-coding genes. All transfer RNA (tRNA) genes exhibited the cloverleaf secondary structure typical for metazoans except in tRNASer(AGN), which lacks the dihydrouridine arm. Phylogenetic analyses were performed based on four nucleotide matrixes for the currently sequenced species of Tipuloidea using Bayesian inference and maximum likelihood methods. Four-cluster likelihood mapping was used to study incongruent signals between different topologies. Pediciidae is supported as the earliest lineage in Tipuloidea, and the sister-group relationship between Cylindrotomidae and Tipulidae is also supported, but the monophyly of Limoniidae is not supported. Our study also supports the monophyly of Elephantomyiini (Elephantomyia + Helius), as one of origins of flower-visiting in Limoniidae. Although Elephantomyiini is sister to Limoniinae + Epiphragma (Limnophilinae) in our study, a more precise understanding of its phylogenetic position in Tipuloidea will require additional studies that include a broader species sample.

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DOI: 10.3897/asp.81.e86898

Authors: Ayman Khamis Elsayed, Tadao Ichita, Makoto Tokuda

Abstract: We describe a gall midge Parampelomyia yukawai Elsayed and Tokuda gen. nov. sp. nov. belonging to the subtribe Schizomyiina (Diptera: Cecidomyiidae: Asphondyliini) based on an integrative taxonomic study. This species forms barely-swollen flower bud galls on the porcelain berry Ampelopsis brevipedunculata var. heterophylla (Vitaceae) in Japan. The new genus is distinguishable from all known schizomyiine genera by tarsomere I lacking a ventroapical extension, the bulbous base of the protrusible portion of the ovipositor, the fused and sclerotized female cerci, the bidentate gonostylus, and the larval terminal abdominal segment that bears two corniform, two asetose and six setose papillae. The new genus is compared with and separated from the similar genera Schizomyia and Ampelomyia morphologically and phylogenetically.

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DOI: 10.3897/asp.80.e83035

Authors: Daubian Santos, Rodrigo dos Reis Santos, Guilherme Cunha Ribeiro

Abstract: The subgenus Amphineurus (Ramphoneurus Alexander) (Diptera: Limoniidae), a group of craneflies endemic to southern South America, is revised. The previously described species are redescribed and illustrated, along with fifteen new species: A. (R.) alexanderi sp. nov., A. (R.) amorimi sp. nov., A. (R.) anchoralis sp. nov., A. (R.) anfractus sp. nov., A. (R.) billinghami sp. nov., A. (R.) caleuchus sp. nov., A. (R.) deceptus sp. nov., A. (R.) falcatus sp. nov., A. (R.) immaculatus sp. nov., A. (R.) morphyi sp. nov., A. (R.) oosterbroeki sp. nov., A. (R.) podenasi sp. nov., A. (R.) theischingeri sp. nov., A. (R.) triangularis sp. nov., and A. (R.) stigmaticus sp. nov. In addition, A. (R.) fuscifusus Alexander is considered a nomen dubium. A phylogenetic analysis with a broad taxonomic sampling with all valid species of the subgenus A. (Rhamphoneurus Alexander) and species from all the subgenera of Amphineurus Skuse is performed. The broad outgroup sampling used in the study (including taxa from the entire range of distribution of Amphineurus) suggests that the subgenus A. (Rhamphoneurus) is a monophyletic taxon, that its closest relatives are taxa currently distributed in New Zealand, and that Australasian taxa are paraphyletic with respect to the Neotropical Clade.

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DOI: 10.3897/asp.80.e78315

Authors: Christoforos Pavlou, Emmanouil Dokianakis, Nikolaos Tsirigotakis, Vasiliki Christodoulou, Yusuf Özbel, Maria Antoniou, Nikos Poulakakis

Abstract: The genus Phlebotomus (Diptera: Psychodidae: Phlebotominae) comprises a group of small winged insect species of medical importance. To date, ten species of Phlebotomus are known to be present in Greece; yet their evolutionary history is poorly studied due to the lack of comprehensive phylogenetic and phylogeographic studies. Herein, we aim to clarify the phylogenetic relationships amongst the local species collected from 12 Aegean Islands, Cyprus and Turkey; and to identify which of the palaeogeographic events may have influenced their biogeographic history. Our analyses revealed for the first time the presence of P. cf. major and P. sergenti in the Aegean Islands. All studied local species were retrieved as monophyletic and the mtDNA and nDNA phylogenetic trees indicated a plausible mitochondrial introgression between the closely related species of the P. major complex. From a palaeogeographic viewpoint, the major driving force that shaped the biogeographic history of the studied Phlebotomus species seems to be the dispersal that started in the Oligocene epoch, followed by several speciation events that occurred at the end of Miocene and the Plio-Pleistocene, including multiple dispersal events of Asiatic origin. The Messinian Salinity Crisis, the bimodal Mediterranean climate, and the glacial and interglacial periods were identified as key drivers for the diversification of the local species of Phlebotomus.

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