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Mosquitoes have profoundly affected human history and continue to threaten human health through the transmission of a diverse array of pathogens. The phylogeny of mosquitoes has remained poorly characterized due to difficulty in taxonomic sampling and limited availability of genomic data beyond the most important vector species. Here, we used phylogenomic analysis of 709 single copy ortholog groups from 256 mosquito species to produce a strongly supported phylogeny that resolves the position of the major disease vector species and the major mosquito lineages. Our analyses support an origin of mosquitoes in the early Triassic (217 MYA [highest posterior density region: 188–250 MYA]), considerably older than previous estimates. Moreover, we utilize an extensive database of host associations for mosquitoes to show that mosquitoes have shifted to feeding upon the blood of mammals numerous times, and that mosquito diversification and host-use patterns within major lineages appear to coincide in earth history both with major continental drift events and with the diversification of vertebrate classes.

 

As a widespread vector of disease with an expanding range, the mosquitoAedes albopictusSkuse (Diptera: Culicidae) is a high priority for research and management.A. albopictushas a complex life history with aquatic egg, larval and pupal stages, and a terrestrial adult stage. This requires targeted management strategies for each life stage, coordinated across time and space. Population genetics can aid inA. albopictuscontrol by evaluating patterns of genetic diversity and dispersal. However, how life stage impacts population genetic characteristics is unknown. We examined whether patterns ofA. albopictusgenetic diversity and differentiation changed with life stage at a spatial scale relevant to management efforts. We first conducted a literature review of field‐caughtA. albopictuspopulation genetic papers and identified 101 peer‐reviewed publications, none of which compared results between life stages. Our study uniquely examines population genomic patterns of egg and adultA. albopictusat five sites in Wake County, North Carolina, USA, using 8425 single nucleotide polymorphisms. We found that the level of genetic diversity and connectivity between sites varied between adults and eggs. This warrants further study and is critical for research aimed at informing local management.

 

Aedes aegyptiis among the best‐studied mosquitoes due to its critical role as a vector of human pathogens and ease of laboratory rearing. Until now, this species was thought to have originated in continental Africa, and subsequently colonized much of the world following the establishment of global trade routes. However, populations of this mosquito on the islands in the southwestern Indian Ocean (SWIO), where the species occurs with its nearest relatives referred to as the Aegypti Group, have received little study. We re‐evaluated the evolutionary history ofAe. aegyptiand these relatives, using three data sets: nucleotide sequence data, 18,489 SNPs and 12 microsatellites. We found that: (a) the Aegypti Group diverged 16 MYA (95% HPD: 7–28 MYA) from its nearest African/Asian ancestor; (b) SWIO populations ofAe. aegyptiare basal to continental African populations; (c) after diverging 7 MYA (95% HPD: 4–15 MYA) from its nearest formally described relative (Ae. mascarensis),Ae. aegyptimoved to continental Africa less than 85,000 years ago, where it recently (<1,000 years ago) split into two recognized subspeciesAe. aegypti formosusand a human commensal,Ae. aegypti aegypti; (d) the Madagascar samples form a clade more distant from all otherAe. aegyptithan the named speciesAe. mascarensis, implying that Madagascar may harbour a new cryptic species; and (e) there is evidence of introgression betweenAe. mascarensisandAe. aegyption Réunion, and between the two subspecies elsewhere in the SWIO, a likely consequence of recent introductions of domesticAe. aegypti aegyptifrom Asia.

 

Abstract Background The most species-rich radiation of animal life in the 66 million years following the Cretaceous extinction event is that of schizophoran flies: a third of fly diversity including Drosophila fruit fly model organisms, house flies, forensic blow flies, agricultural pest flies, and many other well and poorly known true flies. Rapid diversification has hindered previous attempts to elucidate the phylogenetic relationships among major schizophoran clades. A robust phylogenetic hypothesis for the major lineages containing these 55,000 described species would be critical to understand the processes that contributed to the diversity of these flies. We use protein encoding sequence data from transcriptomes, including 3145 genes from 70 species, representing all superfamilies, to improve the resolution of this previously intractable phylogenetic challenge. Results Our results support a paraphyletic acalyptrate grade including a monophyletic Calyptratae and the monophyly of half of the acalyptrate superfamilies. The primary branching framework of Schizophora is well supported for the first time, revealing the primarily parasitic Pipunculidae and Sciomyzoidea stat. rev. as successive sister groups to the remaining Schizophora. Ephydroidea, Drosophila ’s superfamily, is the sister group of Calyptratae. Sphaeroceroidea has modest support as the sister to all non-sciomyzoid Schizophora. We define two novel lineages corroborated by morphological traits, the ‘Modified Oviscapt Clade’ containing Tephritoidea, Nerioidea, and other families, and the ‘Cleft Pedicel Clade’ containing Calyptratae, Ephydroidea, and other families. Support values remain low among a challenging subset of lineages, including Diopsidae. The placement of these families remained uncertain in both concatenated maximum likelihood and multispecies coalescent approaches. Rogue taxon removal was effective in increasing support values compared with strategies that maximise gene coverage or minimise missing data. Conclusions Dividing most acalyptrate fly groups into four major lineages is supported consistently across analyses. Understanding the fundamental branching patterns of schizophoran flies provides a foundation for future comparative research on the genetics, ecology, and biocontrol. 

Abstract Target enrichment (such as Hyb-Seq) is a well-established high throughput sequencing method that has been increasingly used for phylogenomic studies. Unfortunately, current widely used pipelines for analysis of target enrichment data do not have a vigorous procedure to remove paralogs in target enrichment data. In this study, we develop a pipeline we call Putative Paralogs Detection (PPD) to better address putative paralogs from enrichment data. The new pipeline is an add-on to the existing HybPiper pipeline, and the entire pipeline applies criteria in both sequence similarity and heterozygous sites at each locus in the identification of paralogs. Users may adjust the thresholds of sequence identity and heterozygous sites to identify and remove paralogs according to the level of phylogenetic divergence of their group of interest. The new pipeline also removes highly polymorphic sites attributed to errors in sequence assembly and gappy regions in the alignment. We demonstrated the value of the new pipeline using empirical data generated from Hyb-Seq and the Angiosperm 353 kit for two woody genera Castanea (Fagaceae, Fagales) and Hamamelis (Hamamelidaceae, Saxifragales). Comparisons of datasets showed that the PPD identified many more putative paralogs than the popular method HybPiper. Comparisons of tree topologies and divergence times showed evident differences between data from HybPiper and data from our new PPD pipeline. We further evaluated the accuracy and error rates of PPD by BLAST mapping of putative paralogous and orthologous sequences to a reference genome sequence of Castanea mollissima. Compared to HybPiper alone, PPD identified substantially more paralogous gene sequences that mapped to multiple regions of the reference genome (31 genes for PPD compared with 4 genes for HybPiper alone). In conjunction with HybPiper, paralogous genes identified by both pipelines can be removed resulting in the construction of more robust orthologous gene datasets for phylogenomic and divergence time analyses. Our study demonstrates the value of Hyb-Seq with data derived from the Angiosperm 353 probe set for elucidating species relationships within a genus, and argues for the importance of additional steps to filter paralogous genes and poorly aligned regions (e.g., as occur through assembly errors), such as our new PPD pipeline described in this study. 

Abstract Container Aedes mosquitoes are the most important vectors of human arboviruses (i.e., dengue, chikungunya, Zika, or yellow fever). Invasive and native container Aedes spp. potentially utilize natural and artificial containers in specific environments for oviposition. Several container Aedes spp. display ‘skip-oviposition’ behavior, which describes the distribution of eggs among multiple containers during a single gonotrophic cycle. In this study, we compared individual skip-oviposition behavior using identical eight-cup testing arenas with three container Aedes species: Aedes aegypti (Linnaeus), Aedes albopictus (Skuse), and Aedes triseriatus (Say). We applied the index of dispersion, an aggregation statistic, to individual mosquitoes’ oviposition patterns to assess skip-oviposition behavior. Aedes aegypti and Ae. albopictus utilized more cups and distributed eggs more evenly among cups than Ae. triseriatus under nutritionally enriched oviposition media (oak leaf infusion) conditions. When presented with a nutritionally unenriched (tap water) oviposition media, both Ae. aegypti and Ae. albopictus increased egg spreading behavior. Aedes albopictus did not modify skip-oviposition behavior when reared and assessed under fall-like environmental conditions, which induce diapause egg production. This study indicates specific oviposition site conditions influence skip-oviposition behavior with ‘preferred’ sites receiving higher amounts of eggs from any given individual and ‘non-preferred’ sites receive a limited contribution of eggs. A further understanding of skip-oviposition behavior is needed to make the best use of autodissemination trap technology in which skip-ovipositing females spread a potent larvicide among oviposition sites within the environment. 

Scientific collections such as the U.S. National Museum (USNM) are critical to filling knowledge gaps in molecular systematics studies. The global taxonomic impediment has resulted in a reduction of expert taxonomists generating new collections of rare or understudied taxa and these large historic collections may be the only reliable source of material for some taxa. Integrated systematics studies using both morphological examinations and DNA sequencing are often required for resolving many taxonomic issues but as DNA methods often require partial or complete destruction of a sample, there are many factors to consider before implementing destructive sampling of specimens within scientific collections. We present a methodology for the use of archive specimens that includes two crucial phases: 1) thoroughly documenting specimens destined for destructive sampling—a process called electronic vouchering, and 2) the pipeline used for whole genome sequencing of archived specimens, from extraction of genomic DNA to assembly of putative genomes with basic annotation. The process is presented for eleven specimens from two different insect subfamilies of medical importance to humans: Anophelinae (Diptera: Culicidae)—mosquitoes and Triatominae (Hemiptera: Reduviidae)—kissing bugs. Assembly of whole mitochondrial genome sequences of all 11 specimens along with the results of an ortholog search and BLAST against the NCBI nucleotide database are also presented. 

Abstract Although parasites are by definition costly to their host, demonstrating that a parasite is regulating its host abundance in the field can be difficult. Here we present an example of a gregarine parasite, Ascogregarina taiwanensis Lien and Levine (Apicomplexa: Lecudinidae), regulating its mosquito host, Aedes albopictus Skuse (Diptera: Culicidae), in Bermuda. We sampled larvae from container habitats over 2 yr, assessed parasite prevalence, and estimated host abundance from egg counts obtained in neighboring ovitraps. We regressed change in average egg count from 1 yr to the next on parasite prevalence and found a significant negative effect of parasite prevalence. We found no evidence of host density affecting parasite prevalence. Our results demonstrate that even for a parasite with moderate virulence, host regulation can occur in the field.