Reciprocal interactions between neuropeptide F and RYamide regulate host attraction in the mosquito Aedes aegypti.

Female mosquitoes produce eggs in gonadotrophic cycles that are divided between a previtellogenic and vitellogenic phase. Previtellogenic females consume water and sugar sources like nectar while also being attracted to hosts for blood feeding. Consumption of a blood meal activates the vitellogenic phase, which produces mature eggs and suppresses host attraction. In this study, we tested the hypothesis that neuropeptide Y-like hormones differentially modulate host attraction behavior in the mosquito Aedes aegypti. A series of experiments collectively indicated that enteroendocrine cells (EECs) in the posterior midgut produce and release neuropeptide F (NPF) into the hemolymph during the previtellogenic phase which stimulates attraction to humans and biting behavior. Consumption of a blood meal, which primarily consists of protein by dry weight, down-regulated NPF in EECs until mature eggs developed, which was associated with a decline in hemolymph titer. NPF depletion depended on protein digestion but was not associated with EEC loss. Other experiments showed that neurons in the terminal ganglion extend axons to the posterior midgut and produce RYamide, which showed evidence of increased secretion into circulation after a blood meal. Injection of RYamide-1 and -2 into previtellogenic females suppressed host attraction, while coinjection of RYamides with or without short NPF-2 also inhibited the host attraction activity of NPF. Overall, our results identify NPF and RYamide as gut-associated hormones in A. aegypti that link host attraction behavior to shifts in diet during sequential gonadotrophic cycles.

Identification of a viral gene essential for the genome replication of a domesticated endogenous virus in ichneumonid parasitoid wasps.

Thousands of endoparasitoid wasp species in the families Braconidae and Ichneumonidae harbor "domesticated endogenous viruses" (DEVs) in their genomes. This study focuses on ichneumonid DEVs, named ichnoviruses (IVs). Large quantities of DNA-containing IV virions are produced in ovary calyx cells during the pupal and adult stages of female wasps. Females parasitize host insects by injecting eggs and virions into the body cavity. After injection, virions rapidly infect host cells which is followed by expression of IV genes that promote the successful development of wasp offspring. IV genomes consist of two components: proviral segment loci that serve as templates for circular dsDNAs that are packaged into capsids, and genes from an ancestral virus that produce virions. In this study, we generated a chromosome-scale genome assembly for Hyposoter didymator that harbors H. didymator ichnovirus (HdIV). We identified a total of 67 HdIV loci that are amplified in calyx cells during the wasp pupal stage. We then focused on an HdIV gene, U16, which is transcribed in calyx cells during the initial stages of replication. Sequence analysis indicated that U16 contains a conserved domain in primases from select other viruses. Knockdown of U16 by RNA interference inhibited virion morphogenesis in calyx cells. Genome-wide analysis indicated U16 knockdown also inhibited amplification of HdIV loci in calyx cells. Altogether, our results identified several previously unknown HdIV loci, demonstrated that all HdIV loci are amplified in calyx cells during the pupal stage, and showed that U16 is required for amplification and virion morphogenesis.

On the Origin and Evolution of the Mosquito Male-determining Factor Nix.

The mosquito family Culicidae is divided into 2 subfamilies named the Culicinae and Anophelinae. Nix, the dominant male-determining factor, has only been found in the culicines Aedes aegypti and Aedes albopictus, 2 important arboviral vectors that belong to the subgenus Stegomyia. Here we performed sex-specific whole-genome sequencing and RNAseq of divergent mosquito species and explored additional male-inclusive datasets to investigate the distribution of Nix. Except for the Culex genus, Nix homologs were found in all species surveyed from the Culicinae subfamily, including 12 additional species from 3 highly divergent tribes comprising 4 genera, suggesting Nix originated at least 133 to 165 million years ago (MYA). Heterologous expression of 1 of 3 divergent Nix open reading frames (ORFs) in Ae. aegypti resulted in partial masculinization of genetic females as evidenced by morphology and doublesex splicing. Phylogenetic analysis suggests Nix is related to femaleless (fle), a recently described intermediate sex-determining factor found exclusively in anopheline mosquitoes. Nix from all species has a conserved structure, including 3 RNA-recognition motifs (RRMs), as does fle. However, Nix has evolved at a much faster rate than fle. The RRM3 of both Nix and fle are distantly related to the single RRM of a widely distributed and conserved splicing factor transformer-2 (tra2). The RRM3-based phylogenetic analysis suggests this domain in Nix and fle may have evolved from tra2 or a tra2-related gene in a common ancestor of mosquitoes. Our results provide insights into the evolution of sex determination in mosquitoes and will inform broad applications of mosquito-control strategies based on manipulating sex ratios toward nonbiting males.

Functional characterization of Microplitis demolitor bracovirus genes that encode nucleocapsid components.

IMPORTANCE: Understanding how bracoviruses (BVs) function in wasps is of broad interest in the study of virus evolution. This study characterizes most of the Microplitis demolitor bracovirus (MdBV) genes whose products are nucleocapsid components. Results indicate several genes unknown outside of nudiviruses and BVs are essential for normal capsid assembly. Results also indicate most MdBV tyrosine recombinase family members and the DNA binding protein p6.9-1 are required for DNA processing and packaging into nucleocapsids.

Riboflavin instability is a key factor underlying the requirement of a gut microbiota for mosquito development.

We previously determined that several diets used to rear Aedes aegypti and other mosquito species support the development of larvae with a gut microbiota but do not support the development of axenic larvae. In contrast, axenic larvae have been shown to develop when fed other diets. To understand the mechanisms underlying this dichotomy, we developed a defined diet that could be manipulated in concert with microbiota composition and environmental conditions. Initial studies showed that axenic larvae could not grow under standard rearing conditions (27 °C, 16-h light: 8-h dark photoperiod) when fed a defined diet but could develop when maintained in darkness. Downstream assays identified riboflavin decay to lumichrome as the key factor that prevented axenic larvae from growing under standard conditions, while gut community members like Escherichia coli rescued development by being able to synthesize riboflavin. Earlier results showed that conventional and gnotobiotic but not axenic larvae exhibit midgut hypoxia under standard rearing conditions, which correlated with activation of several pathways with essential growth functions. In this study, axenic larvae in darkness also exhibited midgut hypoxia and activation of growth signaling but rapidly shifted to midgut normoxia and arrested growth in light, which indicated that gut hypoxia was not due to aerobic respiration by the gut microbiota but did depend on riboflavin that only resident microbes could provide under standard conditions. Overall, our results identify riboflavin provisioning as an essential function for the gut microbiota under most conditions A. aegypti larvae experience in the laboratory and field.

Toll9 from Bombyx mori functions as a pattern recognition receptor that shares features with Toll-like receptor 4 from mammals.

Toll/Toll-like receptors (TLRs) are key regulators of the innate immune system in both invertebrates and vertebrates. However, while mammalian TLRs directly recognize pathogen-associated molecular patterns, the insect Toll pathway is thought to be primarily activated by binding Spätzle cytokines that are processed from inactive precursors in response to microbial infection. Phylogenetic and structural data generated in this study supported earlier results showing that Toll9 members differ from other insect Tolls by clustering with the mammalian TLR4 group, which recognizes lipopolysaccharide (LPS) through interaction with myeloid differentiation-2 (MD-2)-like proteins. Functional experiments showed that BmToll9 from the silkmoth Bombyx mori also recognized LPS through interaction with two MD-2-like proteins, previously named BmEsr16 and BmPP, that we refer to in this study as BmMD-2A and BmMD-2B, respectively. A chimeric BmToll9-TLR4 receptor consisting of the BmToll9 ectodomain and mouse TLR4 transmembrane and Toll/interleukin-1 (TIR) domains also activated LPS-induced release of inflammatory factors in murine cells but only in the presence of BmMD-2A or BmMD-2B. Overall, our results indicate that BmToll9 is a pattern recognition receptor for LPS that shares conserved features with the mammalian TLR4-MD-2-LPS pathway.

The Complete Genome of Chelonus insularis Reveals Dynamic Arrangement of Genome Components in Parasitoid Wasps That Produce Bracoviruses.

Bracoviruses (BVs) are endogenized nudiviruses in parasitoid wasps of the microgastroid complex (family Braconidae). Microgastroid wasps have coopted nudivirus genes to produce replication-defective virions that females use to transfer virulence genes to parasitized hosts. The microgastroid complex further consists of six subfamilies and ∼50,000 species but current understanding of BV gene inventories and organization primarily derives from analysis of two wasp species in the subfamily Microgastrinae (Microplitis demolitor and Cotesia congregata) that produce M. demolitor BV (MdBV) and C. congregata BV (CcBV). Notably, several genomic features of MdBV and CcBV remain conserved since divergence of M. demolitor and C. congregata ∼53 million years ago (MYA). However, it is unknown whether these conserved traits more broadly reflect BV evolution, because no complete genomes exist for any microgastroid wasps outside the Microgastrinae. In this regard, the subfamily Cheloninae is of greatest interest because it diverged earliest from the Microgastrinae (∼85 MYA) after endogenization of the nudivirus ancestor. Here, we present the complete genome of Chelonus insularis, which is an egg-larval parasitoid in the Cheloninae that produces C. insularis BV (CinsBV). We report that the inventory of nudivirus genes in C. insularis is conserved but are dissimilarly organized compared to M. demolitor and C. congregata. Reciprocally, CinsBV proviral segments share organizational features with MdBV and CcBV but virulence gene inventories exhibit almost no overlap. Altogether, our results point to the functional importance of a conserved inventory of nudivirus genes and a dynamic set of virulence genes for the successful parasitism of hosts. Our results also suggest organizational features previously identified in MdBV and CcBV are likely not essential for BV virion formation. IMPORTANCE Bracoviruses are a remarkable example of virus endogenization, because large sets of genes from a nudivirus ancestor continue to produce virions that thousands of wasp species rely upon to parasitize hosts. Understanding how these genes interact and have been coopted by wasps for novel functions is of broad interest in the study of virus evolution. This work characterizes bracovirus genome components in the parasitoid wasp Chelonus insularis, which together with existing wasp genomes captures a large portion of the diversity among wasp species that produce bracoviruses. Results provide new information about how bracovirus genome components are organized in different wasps while also providing additional insights on key features required for function.

An aphid symbiont confers protection against a specialized RNA virus, another increases vulnerability to the same pathogen.

Insects often harbour heritable symbionts that provide defence against specialized natural enemies, yet little is known about symbiont protection when hosts face simultaneous threats. In pea aphids (Acyrthosiphon pisum), the facultative endosymbiont Hamiltonella defensa confers protection against the parasitoid, Aphidius ervi, and Regiella insecticola protects against aphid-specific fungal pathogens, including Pandora neoaphidis. Here, we investigated whether these two common aphid symbionts protect against a specialized virus A. pisum virus (APV), and whether their antifungal and antiparasitoid services are impacted by APV infection. We found that APV imposed large fitness costs on symbiont-free aphids and these costs were elevated in aphids also housing H. defensa. In contrast, APV titres were significantly reduced and costs to APV infection were largely eliminated in aphids with R. insecticola. To our knowledge, R. insecticola is the first aphid symbiont shown to protect against a viral pathogen, and only the second arthropod symbiont reported to do so. In contrast, APV infection did not impact the protective services of either R. insecticola or H. defensa. To better understand APV biology, we produced five genomes and examined transmission routes. We found that moderate rates of vertical transmission, combined with horizontal transfer through food plants, were the major route of APV spread, although lateral transfer by parasitoids also occurred. Transmission was unaffected by facultative symbionts. In summary, the presence and species identity of facultative symbionts resulted in highly divergent outcomes for aphids infected with APV, while not impacting defensive services that target other enemies. These findings add to the diverse phenotypes conferred by aphid symbionts, and to the growing body of work highlighting extensive variation in symbiont-mediated interactions.

Insulin-like peptide 3 stimulates hemocytes to proliferate in anautogenous and facultatively autogenous mosquitoes.

Most mosquito species are anautogenous, which means they must blood feed on a vertebrate host to produce eggs, while a few are autogenous and can produce eggs without blood feeding. Egg formation is best understood in the anautogenous mosquito Aedes aegypti, where insulin-like peptides (ILPs), ovary ecdysteroidogenic hormone (OEH) and 20-hydroxyecdysone (20E) interact to regulate gonadotrophic cycles. Circulating hemocytes also approximately double in abundance in conjunction with a gonadotrophic cycle, but the factors responsible for stimulating this increase remain unclear. Focusing on Ae. aegypti, we determined that hemocyte abundance similarly increased in intact blood-fed females and decapitated blood-fed females that were injected with ILP3, whereas OEH, 20E or heat-killed bacteria had no stimulatory activity. ILP3 upregulated insulin-insulin growth factor signaling in hemocytes, but few genes - including almost no transcripts for immune factors - were differentially expressed. ILP3 also stimulated circulating hemocytes to increase in two other anautogenous (Anopheles gambiae and Culex quinquefasciatus) and two facultatively autogenous mosquitoes (Aedes atropalpus and Culex pipiens molestus), but had no stimulatory activity in the obligately autogenous mosquito Toxorhynchites amboinensis. Altogether, our results identify ILPs as the primary regulators of hemocyte proliferation in association with egg formation, but also suggest this response has been lost in the evolution of obligate autogeny.

MdBVe46 is an envelope protein that is required for virion formation by Microplitis demolitor bracovirus.

Bracoviruses (BVs) are endogenized nudiviruses that braconid parasitoid wasps have coopted for functions in parasitizing hosts. Microplitis demolitor is a braconid wasp that produces Microplitis demolitor bracovirus (MdBV) and parasitizes the larval stage of the moth Chrysodeixis includens. Some BV core genes are homologs of genes also present in baculoviruses while others are only known from nudiviruses or other BVs. In this study, we had two main goals. The first was to separate MdBV virions into envelope and nucleocapsid fractions before proteomic analysis to identify core gene products that were preferentially associated with one fraction or the other. Results indicated that nearly all MdBV baculovirus-like gene products that were detected by our proteomic analysis had similar distributions to homologs in the occlusion-derived form of baculoviruses. Several core gene products unknown from baculoviruses were also identified as envelope or nucleocapsid components. Our second goal was to functionally characterize a core gene unknown from baculoviruses that was originally named HzNVorf64-like. Immunoblotting assays supported our proteomic data that identified HzNVorf64-like as an envelope protein. We thus renamed HzNVorf64-like as MdBVe46, which we further hypothesized was important for infection of C. includens. Knockdown of MdBVe46 by RNA interference (RNAi) greatly reduced transcript and protein abundance. Knockdown of MdBVe46 also altered virion morphogenesis, near-fully inhibited infection of C. includens, and significantly reduced the proportion of hosts that were successfully parasitized by M. demolitor.

Evolutionary genomics of APSE: a tailed phage that lysogenically converts the bacterium Hamiltonella defensa into a heritable protective symbiont of aphids.

BACKGROUND: Most phages infect free-living bacteria but a few have been identified that infect heritable symbionts of insects or other eukaryotes. Heritable symbionts are usually specialized and isolated from other bacteria with little known about the origins of associated phages. Hamiltonella defensa is a heritable bacterial symbiont of aphids that is usually infected by a tailed, double-stranded DNA phage named APSE. METHODS: We conducted comparative genomic and phylogenetic studies to determine how APSE is related to other phages and prophages. RESULTS: Each APSE genome was organized into four modules and two predicted functional units. Gene content and order were near-fully conserved in modules 1 and 2, which encode predicted DNA metabolism genes, and module 4, which encodes predicted virion assembly genes. Gene content of module 3, which contains predicted toxin, holin and lysozyme genes differed among haplotypes. Comparisons to other sequenced phages suggested APSE genomes are mosaics with modules 1 and 2 sharing similarities with Bordetella-Bcep-Xylostella fastidiosa-like podoviruses, module 4 sharing similarities with P22-like podoviruses, and module 3 sharing no similarities with known phages. Comparisons to other sequenced bacterial genomes identified APSE-like elements in other heritable insect symbionts (Arsenophonus spp.) and enteric bacteria in the family Morganellaceae. CONCLUSIONS: APSEs are most closely related to phage elements in the genus Arsenophonus and other bacteria in the Morganellaceae.

Hypoxia-induced transcription factor signaling is essential for larval growth of the mosquito Aedes aegypti.

Gut microbes positively affect the physiology of many animals, but the molecular mechanisms underlying these benefits remain poorly understood. We recently reported that bacteria-induced gut hypoxia functions as a signal for growth and molting of the mosquito Aedes aegypti In this study, we tested the hypothesis that transduction of a gut hypoxia signal requires hypoxia-induced transcription factors (HIFs). Expression studies showed that HIF-α was stabilized in larvae containing bacteria that induce gut hypoxia but was destabilized in larvae that exhibit normoxia. However, we could rescue growth of larvae exhibiting gut normoxia by treating them with a prolyl hydroxylase inhibitor, FG-4592, that stabilized HIF-α, and inhibit growth of larvae exhibiting gut hypoxia by treating them with an inhibitor, PX-478, that destabilized HIF-α. Using these tools, we determined that HIF signaling activated the insulin/insulin growth factor pathway plus select mitogen-activated kinases and inhibited the adenosine monophosphate-activated protein kinase pathway. HIF signaling was also required for growth of the larval midgut and storage of neutral lipids by the fat body. Altogether, our results indicate that gut hypoxia and HIF signaling activate multiple processes in A. aegypti larvae, with conserved functions in growth and metabolism.

Blood feeding activates the vitellogenic stage of oogenesis in the mosquito Aedes aegypti through inhibition of glycogen synthase kinase 3 by the insulin and TOR pathways.

Most mosquitoes, including Aedes aegypti, only produce eggs after blood feeding on a vertebrate host. Oogenesis in A. aegypti consists of a pre-vitellogenic stage before blood feeding and a vitellogenic stage after blood feeding. Primary egg chambers remain developmentally arrested during the pre-vitellogenic stage but complete oogenesis to form mature eggs during the vitellogenic stage. In contrast, the signaling factors that maintain primary egg chambers in pre-vitellogenic arrest or that activate vitellogenic growth are largely unclear. Prior studies showed that A. aegypti females release insulin-like peptide 3 (ILP3) and ovary ecdysteroidogenic hormone (OEH) from brain neurosecretory cells after blood feeding. Here, we report that primary egg chambers exit pre-vitellogenic arrest by 8 h post-blood meal as evidenced by proliferation of follicle cells, endoreplication of nurse cells, and formation of cytoophidia. Ex vivo assays showed that ILP3 and OEH stimulate primary egg chambers to exit pre-vitellogenic arrest in the presence of nutrients but not in their absence. Characterization of associated pathways indicated that activation of insulin/insulin growth factor signaling (IIS) by ILP3 or OEH inactivated glycogen synthase kinase 3 (GSK3) via phosphorylation by phosphorylated Akt. GSK3 inactivation correlated with accumulation of the basic helix-loop-helix transcription factor Max and primary egg chambers exiting pre-vitellogenic arrest. Direct inhibition of GSK3 by CHIR-99021 also stimulated Myc/Max accumulation and primary egg chambers exiting pre-vitellogenic arrest. Collectively, our results identify GSK3 as a key factor in regulating the pre- and vitellogenic stages of oogenesis in A. aegypti.

Toll family members bind multiple Spätzle proteins and activate antimicrobial peptide gene expression in Drosophila.

The Toll signaling pathway in Drosophila melanogaster regulates several immune-related functions, including the expression of antimicrobial peptide (AMP) genes. The canonical Toll receptor (Toll-1) is activated by the cytokine Spätzle (Spz-1), but Drosophila encodes eight other Toll genes and five other Spz genes whose interactions with one another and associated functions are less well-understood. Here, we conducted in vitro assays in the Drosophila S2 cell line with the Toll/interleukin-1 receptor (TIR) homology domains of each Toll family member to determine whether they can activate a known target of Toll-1, the promoter of the antifungal peptide gene drosomycin. All TIR family members activated the drosomycin promoter, with Toll-1 and Toll-7 TIRs producing the highest activation. We found that the Toll-1 and Toll-7 ectodomains bind Spz-1, -2, and -5, and also vesicular stomatitis virus (VSV) virions, and that Spz-1, -2, -5, and VSV all activated the promoters of drosomycin and several other AMP genes in S2 cells expressing full-length Toll-1 or Toll-7. In vivo experiments indicated that Toll-1 and Toll-7 mutants could be systemically infected with two bacterial species (Enterococcus faecalis and Pseudomonas aeruginosa), the opportunistic fungal pathogen Candida albicans, and VSV with different survival times in adult females and males compared with WT fly survival. Our results suggest that all Toll family members can activate several AMP genes. Our results further indicate that Toll-1 and Toll-7 bind multiple Spz proteins and also VSV, but they differentially affect adult survival after systemic infection, potentially because of sex-specific differences in Toll-1 and Toll-7 expression.

Polydnaviruses: Evolution and Function.

Polydnaviruses (PDVs) were originally viewed as large DNA viruses that are beneficial symbionts of parasitoid wasps. Two groups of PDVs were also recognized: bracoviruses (BVs), which are associated with wasps in the family Braconidae, and ichnoviruses (IVs), which are associated with wasps in the family Ichneumonidae. Results to date indicate that BVs are endogenous virus elements (EVEs) that evolved from an ancient betanudivirus. IVs are also likely EVEs but are unrelated to BVs. BVs and IVs are very unusual relative to most known EVEs because they retain many viral functions that benefit wasps in parasitizing hosts. However, BVs and IVs cannot be considered beneficial symbionts because all components of their genomes are fixed in wasps. Recent studies indicate that other nudiviruses have endogenized in insects. Each exhibits a different functional fate from BVs but shares certain architectural features. We discuss options for classifying BVs and other endogenized nudiviruses. We also discuss future directions.

Predaceous Toxorhynchites mosquitoes require a living gut microbiota to develop.

Most species of mosquitoes are detritivores that feed on decaying plant and animal materials in their aquatic environment. Studies of several detritivorous mosquito species indicate that they host relatively low diversity communities of microbes that are acquired from the environment while feeding. Our recent results also indicate that detritivorous species normally require a living gut microbiota to grow beyond the first instar. Less well known is that some mosquitoes, including those belonging to the genus Toxorhynchites, are predators that feed on other species of mosquitoes and nektonic prey. In this study, we asked whether predaceous Toxorhynchites amboinensis larvae still require living microbes in their gut in order to develop. Using the detritivorous mosquito Aedes aegypti as prey, we found that T. amboinensis larvae harbour bacterial communities that are highly similar to that of their prey. Functional assays showed that T. amboinensis first instars provided axenic (i.e. bacteria-free) prey failed to develop, while two bacterial species present in gnotobiotic (i.e. colonized by one or more known bacterial species) prey successfully colonized the T. amboinensis gut and rescued development. Axenic T. amboinensis larvae also displayed defects in growth consistent with previously identified roles for microbe-mediated gut hypoxia in nutrient acquisition and assimilation in A. aegypti. Collectively, these results support a conserved role for gut microbes in regulating the development of mosquitoes with different feeding strategies.

Bacteria-mediated hypoxia functions as a signal for mosquito development.

Mosquitoes host communities of microbes in their digestive tract that consist primarily of bacteria. We previously reported that several mosquito species, including Aedes aegypti, do not develop beyond the first instar when fed a nutritionally complete diet in the absence of a gut microbiota. In contrast, several species of bacteria, including Escherichia coli, rescue development of axenic larvae into adults. The molecular mechanisms underlying bacteria-dependent growth are unknown. Here, we designed a genetic screen around E. coli that identified high-affinity cytochrome bd oxidase as an essential bacterial gene product for mosquito growth. Bioassays showed that bacteria in nonsterile larvae and gnotobiotic larvae inoculated with wild-type E. coli reduced midgut oxygen levels below 5%, whereas larvae inoculated with E. coli mutants defective for cytochrome bd oxidase did not. Experiments further supported that hypoxia leads to growth and ecdysone-induced molting. Altogether, our results identify aerobic respiration by bacteria as a previously unknown but essential process for mosquito development.

The genomes of two parasitic wasps that parasitize the diamondback moth.

BACKGROUND: Parasitic insects are well-known biological control agents for arthropod pests worldwide. They are capable of regulating their host's physiology, development and behaviour. However, many of the molecular mechanisms involved in host-parasitoid interaction remain unknown. RESULTS: We sequenced the genomes of two parasitic wasps (Cotesia vestalis, and Diadromus collaris) that parasitize the diamondback moth Plutella xylostella using Illumina and Pacbio sequencing platforms. Genome assembly using SOAPdenovo produced a 178 Mb draft genome for C. vestalis and a 399 Mb draft genome for D. collaris. A total set that contained 11,278 and 15,328 protein-coding genes for C. vestalis and D. collaris, respectively, were predicted using evidence (homology-based and transcriptome-based) and de novo prediction methodology. Phylogenetic analysis showed that the braconid C. vestalis and the ichneumonid D. collaris diverged approximately 124 million years ago. These two wasps exhibit gene gains and losses that in some cases reflect their shared life history as parasitic wasps and in other cases are unique to particular species. Gene families with functions in development, nutrient acquisition from hosts, and metabolism have expanded in each wasp species, while genes required for biosynthesis of some amino acids and steroids have been lost, since these nutrients can be directly obtained from the host. Both wasp species encode a relative higher number of neprilysins (NEPs) thus far reported in arthropod genomes while several genes encoding immune-related proteins and detoxification enzymes were lost in both wasp genomes. CONCLUSIONS: We present the annotated genome sequence of two parasitic wasps C. vestalis and D. collaris, which parasitize a common host, the diamondback moth, P. xylostella. These data will provide a fundamental source for studying the mechanism of host control and will be used in parasitoid comparative genomics to study the origin and diversification of the parasitic lifestyle.

Bacteriophage acquisition restores protective mutualism.

Insects are frequently infected with inherited facultative symbionts known to provide a range of conditionally beneficial services, including host protection. Pea aphids (Acyrthosiphon pisum) often harbour the bacterium Hamiltonella defensa, which together with its associated bacteriophage A. pisum secondary endosymbiont (APSE) confer protection against an important natural enemy, the parasitic wasp Aphidius ervi. Previous studies showed that spontaneous loss of phage APSE resulted in the complete loss of the protective phenotype. Here, we demonstrate that APSEs can be experimentally transferred into phage-free (i.e. non-protecting) Hamiltonella strains. Unexpectedly, trials using injections of phage particles alone failed, with successful transfer occurring only when APSE and Hamiltonella were simultaneously injected. After transfer, stable establishment of APSE fully restored anti-parasitoid defenses. Thus, phages associated with heritable bacterial symbionts can move horizontally among symbiont strains facilitating the rapid transfer of ecologically important traits although natural barriers may preclude regular exchange.

Ovary ecdysteroidogenic hormone requires a receptor tyrosine kinase to activate egg formation in the mosquito Aedes aegypti.

Mosquitoes are major disease vectors because most species must feed on blood from a vertebrate host to produce eggs. Blood feeding by the vector mosquito Aedes aegypti triggers the release of two neurohormones, ovary ecdysteroidogenic hormone (OEH) and insulin-like peptides (ILPs), which activate multiple processes required for egg formation. ILPs function by binding to the insulin receptor, which activates downstream components in the canonical insulin signaling pathway. OEH in contrast belongs to a neuropeptide family called neuroparsins, whose receptor is unknown. Here we demonstrate that a previously orphanized receptor tyrosine kinase (RTK) from A. aegypti encoded by the gene AAEL001915 is an OEH receptor. Phylogenetic studies indicated that the protein encoded by this gene, designated AAEL001915, belongs to a clade of RTKs related to the insulin receptor, which are distinguished by an extracellular Venus flytrap module. Knockdown of AAEL001915 by RNAi disabled OEH-mediated egg formation in A. aegypti. AAEL001915 was primarily detected in the mosquito ovary in association with follicular epithelial cells. Both monomeric and dimeric AAEL001915 were detected in mosquito ovaries and transfected Drosophila S2 cells. Functional assays further indicated that OEH bound to dimeric AAEL001915, which resulted in downstream phosphorylation of Ak strain transforming factor (Akt). We hypothesize that orthologs of AAEL001915 in other insects are neuroparsin receptors.