N6-methyladenosine modification of RNA controls dopamine synthesis to influence labour division in ants.

The N6-methyladenosine (m6A) modification of RNA has been reported to remodel gene expression in response to environmental conditions; however, the biological role of m6A in social insects remains largely unknown. In this study, we explored the role of m6A in the division of labour by worker ants (Solenopsis invicta). We first determined the presence of m6A in RNAs from the brains of worker ants and found that m6A methylation dynamics differed between foragers and nurses. Depletion of m6A methyltransferase or chemical suppression of m6A methylation in foragers resulted in a shift to 'nurse-like' behaviours. Specifically, mRNAs of dopamine receptor 1 (Dop1) and dopamine transporter (DAT) were modified by m6A, and their expression increased dopamine levels to promote the behavioural transition from foragers to nurses. The abundance of Dop1 and DAT mRNAs and their stability were reduced by the inhibition of m6A modification caused by the silencing of Mettl3, suggesting that m6A modification in worker ants modulates dopamine synthesis, which regulates labour division. Collectively, our results provide the first example of the epitranscriptomic regulation of labour division in social insects and implicate m6A regulatory mechanism as a potential novel target for controlling red imported fire ants.

Exploring the evolutionary dynamics of myrmecophytism: Perspectives from the Southeast Asian Macaranga ant-plant symbiosis.

Myrmecophytic plants utilise defensive services offered by obligate ant partners nesting in their domatia in a novel means of survival in tropical habitats. Although much is known about the ecology of myrmecophytism, there aren't enough empirical examples to demonstrate whether it substantially influences evolutionary patterns in host plant lineages. In this study, we make use of the species-rich Macaranga (Euphorbiaceae) ant-plant symbiosis distributed in the Southeast Asian Sundaland to delve into the evolutionary dynamics of myrmecophytism in host plants. We generated the most comprehensive dated phylogeny of myrmecophytic Macaranga till date using genotyping-by-sequencing (GBS). With this in hand, we traced the evolutionary history of myrmecophytism in Macaranga using parametric biogeography and ancestral state reconstruction. Diversification rate analysis methods were employed to determine if myrmecophytism enhanced diversification rates in the genus. Our results demonstrate that myrmecophytism is labile and easily lost. Ancestral state reconstruction supported a single origin of myrmecophytism in Macaranga âˆ¼18 mya on Borneo followed by multiple losses. Diversification rate analysis methods did not yield sufficient evidence to support the hypothesis that myrmecophytism enhanced diversification rates in Macaranga; we found that topographical features on Borneo may have played a more direct role in the divergence of clades instead. Our study provides evidence that while the acquisition of domatia clearly functions as a key innovation that has enabled host plants to exploit the environment in novel ways, it may not necessarily enhance diversification rates. In fact, we hypothesise that overly specialised cases of myrmecophytism may even be an evolutionary dead end.

De novo assembly of the complete mitochondrial genome of Mycetophylax simplex Emery, 1888 through organelle targeting revels no substantial expansion of gene spacers, but rather some slightly shorter genes.

Mitochondria play a key role in cell biology and have their own genome, residing in a highly oxidative environment that induces faster changes than the nuclear genome. Because of this, mitochondrial markers have been exploited to reconstruct phylogenetic and phylogeographic relationships in studies of adaptation and molecular evolution. In this study, we determined the complete mitogenome of the fungus-farming ant Mycetophylax simplex (Hymenoptera, Formicidae) and conducted a comparative analysis among 29 myrmicine ant mitogenomes. Mycetophylax simplex is an endemic ant that inhabits sand dunes along the southern Atlantic coast. Specifically, the species occur in the ecosystem known as "restinga", within the Atlantic Forest biome. Due to habitat degradation, land use and decline of restinga habitats, the species is considered locally extinct in extremely urban beaches and is listed as vulnerable on the Brazilian Red List (ICMBio). We employed a mitochondrion-targeting approach to obtain the complete mitogenome through high-throughput DNA sequencing technology. This method allowed us to determine the mitogenome with high performance, coverage and low cost. The circular mitogenome has a length of 16,367 base pairs enclosing 37 genes (13 protein-coding genes, 22 tRNAs and 2 rRNAs) along with one control region (CR). All the protein-coding genes begin with a typical ATN codon and end with the canonical stop codons. All tRNAs formed the fully paired acceptor stems and fold into the typical cloverleaf-shaped secondary structures. The gene order is consistent with the shared Myrmicinae structure, and the A + T content of the majority strand is 81.51%. Long intergenic spacers were not found but some gene are slightly shorter. The phylogenetic relationships based on concatenated nucleotide and amino acid sequences of the 13 protein-coding genes, using Maximum Likelihood and Bayesian Inference methods, indicated that mitogenome sequences were useful in resolving higher-level relationship within Formicidae.

Positive selection has shaped the evolution of Argentine ant immune genes both in native and introduced supercolonies.

The highly invasive Argentine ant (Linepithema humile) started its colonisation from the species' native range in South America approximately 150 years ago and has since become one of the major pests in the world. We investigated how the shifts into new ranges have affected the evolution of Argentine ants' immune genes. To the best of our knowledge, this is the first broadscale population genetic study focusing on ants' immune genes. We analysed comprehensive targeted-seq data of immune and non-immune genes containing 174 genes from 18 Argentine ant supercolonies covering the species' native and introduced ranges. We predicted that the immune gene evolution of introduced supercolonies differs from that of the native supercolonies and proposed two different, non-mutually exclusive hypotheses for this: 1) the enemy release hypothesis and 2) the higher pathogen pressure hypothesis - both of which seem to explain the observed evolutionary patterns on their behalf. Our results show that the introduced supercolonies were targeted by weaker selection than natives, but positive selection was evident among supercolonies of both ranges. Moreover, in some cases, such as the antiviral RNAi genes, introduced range supercolonies harboured a higher proportion of positively selected genes than natives. This observation was striking, knowing the recent demographic history and the detected generally lower selection efficacy of introduced supercolonies. In conclusion, it is evident that pathogen pressure is ubiquitous and strongly affects the immune gene evolution in Argentine ants.

Ant backbone phylogeny resolved by modelling compositional heterogeneity among sites in genomic data.

Ants are the most ubiquitous and ecologically dominant arthropods on Earth, and understanding their phylogeny is crucial for deciphering their character evolution, species diversification, and biogeography. Although recent genomic data have shown promise in clarifying intrafamilial relationships across the tree of ants, inconsistencies between molecular datasets have also emerged. Here I re-examine the most comprehensive published Sanger-sequencing and genome-scale datasets of ants using model comparison methods that model among-site compositional heterogeneity to understand the sources of conflict in phylogenetic studies. My results under the best-fitting model, selected on the basis of Bayesian cross-validation and posterior predictive model checking, identify contentious nodes in ant phylogeny whose resolution is modelling-dependent. I show that the Bayesian infinite mixture CAT model outperforms empirical finite mixture models (C20, C40 and C60) and that, under the best-fitting CAT-GTR + G4 model, the enigmatic Martialis heureka is sister to all ants except Leptanillinae, rejecting the more popular hypothesis supported under worse-fitting models, that place it as sister to Leptanillinae. These analyses resolve a lasting controversy in ant phylogeny and highlight the significance of model comparison and adequate modelling of among-site compositional heterogeneity in reconstructing the deep phylogeny of insects.

The genome of the ant Tetramorium bicarinatum reveals a tandem organization of venom peptides genes allowing the prediction of their regulatory and evolutionary profiles.

BACKGROUND: Venoms have evolved independently over a hundred times in the animal kingdom to deter predators and/or subdue prey. Venoms are cocktails of various secreted toxins, whose origin and diversification provide an appealing system for evolutionary researchers. Previous studies of the ant venom of Tetramorium bicarinatum revealed several Myrmicitoxin (MYRTX) peptides that gathered into seven precursor families suggesting different evolutionary origins. Analysis of the T. bicarinatum genome enabling further genomic approaches was necessary to understand the processes underlying the evolution of these myrmicitoxins. RESULTS: Here, we sequenced the genome of Tetramorium bicarinatum and reported the organisation of 44 venom peptide genes (vpg). Of the eleven chromosomes that make up the genome of T. bicarinatum, four carry the vpg which are organized in tandem repeats. This organisation together with the ML evolutionary analysis of vpg sequences, is consistent with evolution by local duplication of ancestral genes for each precursor family. The structure of the vpg into two or three exons is conserved after duplication events while the promoter regions are the least conserved parts of the vpg even for genes with highly identical sequences. This suggests that enhancer sequences were not involved in duplication events, but were recruited from surrounding regions. Expression level analysis revealed that most vpg are highly expressed in venom glands, although one gene or group of genes is much more highly expressed in each family. Finally, the examination of the genomic data revealed that several genes encoding transcription factors (TFs) are highly expressed in the venom glands. The search for binding sites (BS) of these TFs in the vpg promoters revealed hot spots of GATA sites in several vpg families. CONCLUSION: In this pioneering investigation on ant venom genes, we provide a high-quality assembly genome and the annotation of venom peptide genes that we think can fosters further genomic research to understand the evolutionary history of ant venom biochemistry.

Exploring horizontal transfer of mariner transposable elements among ants and aphids.

Aphids and ants are mutualistic species with a close space-time relationship, which may facilitate the occurrence of horizontal transfer events between these insect groups. Myrmar-like mariner elements were previously isolated from two ant (Myrmica ruginodis and Tapinoma ibericum) and two aphid species (Aphis fabae and Aphis hederae). The aim of this work is to determine the presence of Myrmar-like mariner elements in new ant and aphid species, as well as to analyze the likelihood of horizontal transfer events between these taxa. To accomplish this, the Myrmar-like element has been isolated from five aphid species and six ant species. Among these new analyzed species, full-length Myrmar-like mariner elements with very high sequence similarity have been isolated from the aphids Aphis nerii, Aphis spiraecola, Brachycaudus cardui, and Rhopalosiphum maidis as well as from the ants Lasius grandis and Lasius niger, even though aphids and ants belong to two insect orders (Hemiptera and Hymenoptera) that have evolved independently for at least 300 million-years. Both Lasius species establish frequent mutualistic relationships with multiple aphid species, including A. nerii, A. spiraecola, and B. cardui. The study of the putative protein encoded by them and the phylogenetic analysis suggests that they could be active transposons shared by aphids and ants through horizontal transfer events. Additionally, mariner elements with internal deletion were found in several aphids and one ant species, showing a high degree of sequence similarity among them. The characteristics of these elements with internal deletion suggest a complex origin involving various evolutionary processes, possibly including also horizontal transfer events. Myrmar-like elements have also been isolated from the other ant species, although without similarity with the aphid mariner sequences. Myrmar-like elements are also present in phylogenetically distant insect species, as well as in one crustacean species. The phylogenetic study carried out with all Myrmar-like elements suggests the probable occurrence of horizontal transfer events.

Out of the desert: Paleoclimatic changes drove the diversification of arid-adapted Ocymyrmex ants in southern Africa.

A highly endemic ant fauna is found in the arid regions of southern Africa, including species in the genus Ocymyrmex. This genus of ants has higher species richness in the western arid regions of southern Africa compared to tropical and subtropical parts of the continent. The processes that have produced these patterns of diversity and distribution of arid adapted ants in southern Africa have never been investigated. The diversification of many other taxa in the region has been associated with past climate fluctuations that occurred during the Miocene epoch. In this study, the nature and timing of historical processes that may have led to the diversification within Ocymyrmex were assessed. We hypothesized that past climate oscillations, characterized by long periods of aridification, have driven the current distribution of Ocymyrmex species that resulted in the highest species richness of the genus in the Deserts & xeric shrublands biome in southern Africa. Ninety-four Ocymyrmex worker specimens from Botswana, Kenya, Namibia, South Africa, Tanzania and Zimbabwe, representing 21 currently described species and six morphospecies, were included in a phylogenomic analysis. Phylogenies for the genus, based on next generation sequencing data from ultraconserved elements, were inferred using Maximum Likelihood, and a dating analysis was performed using secondary age estimates as calibration points. A distribution database of Ocymyrmex records was used to assign species ranges, which were then coded according to major biomes in southern Africa and used as input for biogeographical analysis. We explored the phylogenomic relationships of Ocymyrmex and analysed these within a biogeographical and paleoclimatic framework to disentangle the potential processes responsible for diversification in this group. Dating analyses estimated that the crown age of Ocymyrmex dates to the Oligocene, around 32 Ma. Diversification within this group occurred between the mid-Miocene (∼12.5 Ma) and Pleistocene (∼2 Ma). Our biogeographic analyses suggest that Ocymyrmex species originated in the south-western region of southern Africa, which is now part of the Deserts & xeric shrublands biome and diversified into eastern subtropical areas during the Pliocene. Paleoclimatic changes resulting in increased aridity during the Miocene likely drove the diversification of the genus Ocymyrmex. It is most likely that the diversification of grasslands, because of historical climate change, facilitated the diversification of these ants to the eastern parts of southern Africa when open grasslands replaced forests during the early Miocene.

Reference genome of the bicolored carpenter ant, Camponotus vicinus.

Carpenter ants in the genus Camponotus are large, conspicuous ants that are abundant and ecologically influential in many terrestrial ecosystems. The bicolored carpenter ant, Camponotus vicinus Mayr, is distributed across a wide range of elevations and latitudes in western North America, where it is a prominent scavenger and predator. Here, we present a high-quality genome assembly of C. vicinus from a sample collected in Sonoma County, California, near the type locality of the species. This genome assembly consists of 38 scaffolds spanning 302.74 Mb, with contig N50 of 15.9 Mb, scaffold N50 of 19.9 Mb, and BUSCO completeness of 99.2%. This genome sequence will be a valuable resource for exploring the evolutionary ecology of C. vicinus and carpenter ants generally. It also provides an important tool for clarifying cryptic diversity within the C. vicinus species complex, a genetically diverse set of populations, some of which are quite localized and of conservation interest.

Transcriptional and post-transcriptional control of odorant receptor choice in ants.

Insects and mammals have independently evolved odorant receptor genes that are arranged in large genomic tandem arrays. In mammals, each olfactory sensory neuron chooses to express a single receptor in a stochastic process that includes substantial chromatin rearrangements. Here, we show that ants, which have the largest odorant receptor repertoires among insects, employ a different mechanism to regulate gene expression from tandem arrays. Using single-nucleus RNA sequencing, we found that ant olfactory sensory neurons choose different transcription start sites along an array but then produce mRNA from many downstream genes. This can result in transcripts from dozens of receptors being present in a single nucleus. Such rampant receptor co-expression at first seems difficult to reconcile with the narrow tuning of the ant olfactory system. However, RNA fluorescence in situ hybridization showed that only mRNA from the most upstream transcribed odorant receptor seems to reach the cytoplasm where it can be translated into protein, whereas mRNA from downstream receptors gets sequestered in the nucleus. This implies that, despite the extensive co-expression of odorant receptor genes, each olfactory sensory neuron ultimately only produces one or very few functional receptors. Evolution has thus found different molecular solutions in insects and mammals to the convergent challenge of selecting small subsets of receptors from large odorant receptor repertoires.

Comparative Analyses of Reproductive Caste Types Reveal Vitellogenin Genes Involved in Queen Fertility in Solenopsis invicta.

The red imported fire ant (Solenopsis invicta Buren) is a social pest species with a robust reproductive ability that causes extensive damage. Identification of the genes involved in queen fertility is critical in order to better understand the reproductive biology and screening for the potential molecular targets in S. invicta. Here, we used the mRNA deep sequencing (RNA-seq) approach to identify differentially expressed genes (DEGs) in the transcriptomes of three reproductive caste types of S. invicta, including queen (QA) and winged female (FA) and male (MA) ants. The genes that were specific to and highly expressed in the queens were then screened, and the Vg2 and Vg3 genes were chosen as targets to explore their functions in oogenesis and fertility. A minimum of 6.08 giga bases (Gb) of clean reads was obtained from all samples, with a mapping rate > 89.78%. There were 7524, 7133, and 977 DEGs identified in the MA vs. QA, MA vs. FA, and FA vs. QA comparisons, respectively. qRT-PCR was used to validate 10 randomly selected DEGs, including vitellogenin 2 (Vg2) and 3 (Vg3), and their expression patterns were mostly consistent with the RNA-seq data. The S. invicta Vgs included conserved domains and motifs that are commonly found in most insect Vgs. SiVg2 and SiVg3 were highly expressed in queens and winged females and were most highly expressed in the thorax, followed by the fat body, head, and epidermis. Evaluation based on a loss-of-function-based knockdown analysis showed that the downregulation of either or both of these genes resulted in smaller ovaries, less oogenesis, and less egg production. The results of transcriptional sequencing provide a foundation for clarifying the regulators of queen fertility in S. invicta. The functions of SiVg2 and SiVg3 as regulators of oogenesis highlight their importance in queen fecundity and their potential as targets of reproductive disruption in S. invicta control.

Microbial symbionts are shared between ants and their associated beetles.

The transmission of microbial symbionts across animal species could strongly affect their biology and evolution, but our understanding of transmission patterns and dynamics is limited. Army ants (Formicidae: Dorylinae) and their hundreds of closely associated insect guest species (myrmecophiles) can provide unique insights into interspecific microbial symbiont sharing. Here, we compared the microbiota of workers and larvae of the army ant Eciton burchellii with those of 13 myrmecophile beetle species using 16S rRNA amplicon sequencing. We found that the previously characterized specialized bacterial symbionts of army ant workers were largely absent from ant larvae and myrmecophiles, whose microbial communities were usually dominated by Rickettsia, Wolbachia, Rickettsiella and/or Weissella. Strikingly, different species of myrmecophiles and ant larvae often shared identical 16S rRNA genotypes of these common bacteria. Protein-coding gene sequences confirmed the close relationship of Weissella strains colonizing army ant larvae, some workers and several myrmecophile species. Unexpectedly, these strains were also similar to strains infecting dissimilar animals inhabiting very different habitats: trout and whales. Together, our data show that closely interacting species can share much of their microbiota, and some versatile microbial species can inhabit and possibly transmit across a diverse range of hosts and environments.

Social antagonism facilitates supergene expansion in ants.

Antagonistic selection has long been considered a major driver of the formation and expansion of sex chromosomes. For example, sexually antagonistic variation on an autosome can select for suppressed recombination between that autosome and the sex chromosome, leading to a neo-sex chromosome. Autosomal supergenes, chromosomal regions containing tightly linked variants affecting the same complex trait, share similarities with sex chromosomes, raising the possibility that sex chromosome evolution models can explain the evolution of genome structure and recombination in other contexts. We tested this premise in a Formica ant species, wherein we identified four supergene haplotypes on chromosome 3 underlying colony social organization and sex ratio. We discovered a novel rearranged supergene variant (9r) on chromosome 9 underlying queen miniaturization. The 9r is in strong linkage disequilibrium with one chromosome 3 haplotype (P2) found in multi-queen (polygyne) colonies. We suggest that queen miniaturization is strongly disfavored in the single-queen (monogyne) background and is thus socially antagonistic. As such, divergent selection experienced by ants living in alternative social "environments" (monogyne and polygyne) may have contributed to the emergence of a genetic polymorphism on chromosome 9 and associated queen-size dimorphism. Consequently, an ancestral polygyne-associated haplotype may have expanded to include the polymorphism on chromosome 9, resulting in a larger region of suppressed recombination spanning two chromosomes. This process is analogous to the formation of neo-sex chromosomes and consistent with models of expanding regions of suppressed recombination. We propose that miniaturized queens, 16%-20% smaller than queens without 9r, could be incipient intraspecific social parasites.

Scrutinising an inscrutable bark-nesting ant: Exploring cryptic diversity in the Rhopalomastix javana (Hymenoptera: Formicidae) complex using DNA barcodes, genome-wide MIG-seq and geometric morphometrics.

Overlooking cryptic species diversity has grave implications on assessments of climate change impacts on biodiversity, ecosystems and organismal populations. Discriminating between cryptic species has long been challenging even for seasoned taxonomists, as interspecies morphological differences are often indiscernible by visual observation. Multi-disciplinary methods involving genetic analyses in conjunction with quantitative morphological data, should therefore be used to investigate boundaries between cryptic species. We adopted an integrated approach combining analyses of mitochondrial COI barcodes, a genome-wide dataset obtained via multiplexed inter-simple sequence repeats (ISSRs) genotyping by sequencing (MIG-seq), and geometric morphometrics to investigate species divergences in the inscrutable Rhopalomastix javana species complex. Objective clustering of COI suggested five putative molecular species units divergent from each other by thresholds within 4.2-10.6% uncorrected pairwise distance. Phylogenetic analyses based on concatenated MIG-seq data also recovered and strongly supported the monophyly of five major lineages in agreement with COI clusters. Co-ancestry analyses based on MIG-seq data using fineRADstructure resolved variable patterns of admixture linked to geography, and potential genetic drift within some putative species. Geometric morphometric analyses of specimen images further detected statistically significant differences in at least one of three anatomical aspects (Head, Meso, Profile) between all pairs of putative species. Head shape (full-face view) was determined to be the most informative character for species diagnosis, with relatively high classification accuracy. Thin-plate spline deformation grids highlighted areas of high variation between species in each shape for deeper taxonomic scrutiny. The presence of species from multiple distinct lineages existing in near-sympatry firmly demonstrates that R. javana comprises more than one closely-related species, but exact species boundaries are difficult to ascertain. Differences in elevation and its associated abiotic effects on ant adaptations and reproductive phenology may contribute to restricting gene flow and maintaining species boundaries between sympatric populations of the R. javana complex. We further assess the advantages and limitations of geometric morphometrics as a taxonomic tool. Despite its drawbacks, our combined approach has helped draw important insights on cryptic diversity in R. javana, and also identified gaps of knowledge that await address. Results from this study will inform and prime future in-depth taxonomic investigation on the R. javana complex, including formal descriptions and establishment of the five putative species.

Task-specific odorant receptor expression in worker antennae indicates that sensory filters regulate division of labor in ants.

Division of labor (DOL) is a characteristic trait of insect societies, where tasks are generally performed by specialized individuals. Inside workers focus on brood or nest care, while others take risks by foraging outside. Theory proposes that workers have different thresholds to perform certain tasks when confronted with task-related stimuli, leading to specialization and consequently DOL. Workers are presumed to vary in their response to task-related cues rather than in how they perceive such information. Here, we test the hypothesis that DOL instead stems from workers varying in their efficiency to detect stimuli of specific tasks. We use transcriptomics to measure mRNA expression levels in the antennae and brain of nurses and foragers of the ant Temnothorax longispinosus. We find seven times as many genes to be differentially expressed between behavioral phenotypes in the antennae compared to the brain. Moreover, half of all odorant receptors are differentially expressed, with an overrepresentation of the 9-exon gene family upregulated in the antennae of nurses. Nurses and foragers thus apparently differ in the perception of their olfactory environment and task-related signals. Our study supports the hypothesis that antennal sensory filters predispose workers to specialize in specific tasks.

Long live the host! Proteomic analysis reveals possible strategies for parasitic manipulation of its social host.

Parasites with complex life cycles often manipulate the phenotype of their intermediate hosts to increase the probability of transmission to their definitive hosts. Infection with Anomotaenia brevis, a cestode that uses Temnothorax nylanderi ants as intermediate hosts, leads to a multiple-fold extension of host lifespan and to changes in behaviour, morphology and colouration. The mechanisms behind these changes are unknown, as is whether the increased longevity is achieved through parasite manipulation. Here, we demonstrate that the parasite releases proteins into its host with functions that might explain the observed changes. These parasitic proteins make up a substantial portion of the proteome of the hosts' haemolymph, and thioredoxin peroxidase and superoxide dismutase, two antioxidants, exhibited the highest abundances among them. The largest part of the secreted proteins could not be annotated, indicating they are either novel or severely altered during recent coevolution to function in host manipulation. We also detected shifts in the hosts' proteome with infection, in particular an overabundance of vitellogenin-like A in infected ants, a protein that regulates division of labour in Temnothorax ants, which could explain the observed behavioural changes. Our results thus suggest two different strategies that might be employed by this parasite to manipulate its host: secreting proteins with immediate influence on the host's phenotype and altering the host's translational activity. Our findings highlight the intricate molecular interplay required to influence the phenotype of a host and point to potential signalling pathways and genes involved in parasite-host communication.

Age- and caste-independent piRNAs in the germline and miRNA profiles linked to caste and fecundity in the ant Temnothorax rugatulus.

Social insects are models for studies of phenotypic plasticity. Ant queens and workers vary in fecundity and lifespan, which are enhanced and extended in queens. Yet, the regulatory mechanisms underlying this variation are not well understood. Ant queens live and reproduce for years, so that they need to protect their germline from transposable element (TE) activity, which may be redundant in short-lived, often sterile workers. We analysed the expression of two protective classes of small RNAs, microRNAs (miRNAs) and Piwi-interacting RNAs (piRNAs), in various tissues, castes and age classes of the ant Temnothorax rugatulus. In queens, piRNAs were highly abundant in ovaries with TEs being their clear targets, with reduced but still detectable piRNA-specific ping-pong signatures in thorax and brains. piRNA pathway activity varied little with age in queens. Moreover, the reduced ovaries of workers also exhibited similar piRNA activity and this not only in young, fertile workers, but also in older foragers with regressed ovaries. Therefore, these ants protect their germline through piRNA activity, regardless of ovarian development, age or caste, even in sterile workers often considered the soma of the superorganism. Our tissue-specific miRNA analysis detected the expression of 304 miRNAs, of which 105 were expressed in all tissues, 10 enriched in the brain, three in the thorax, whereas 83 were ovarian-specific. We identified ovarian miRNAs whose expression was related to caste, fecundity and age, and which likely regulate group-specific gene expression. sRNA shifts in young- to middle-aged queens were minor, suggesting delayed senescence in this reproductive caste.

The invasive ant Solenopsis invicta is established in Europe.

The red imported fire ant (Solenopsis invicta) is classified as one of the worst invasive alien species1 and as the fifth costliest worldwide2, impacting ecosystems, agriculture and human health3. We report the establishment of S. invicta in Europe for the first time, documenting a mature population in Sicily. We use genetic analyses to assess its putative origin, as well as wind tracking and species distribution modelling to predict its potential range on the continent. We show that half of the urban areas in Europe are already suitable and that climate warming expected under current trends will favor the expansion of this invasive ant.

Estrogen-related receptor functions via the 20-hydroxyecdysone and IIS/TOR signaling pathways to regulate the development and morphology changes of ant Polyrhachis vicina Roger (Hymenoptera, Formicidae).

Estrogen-related receptor (ERR) is a key regulator of insect growth, development, and metabolic processes in insects; however, the molecular mechanisms underlying its effects are not fully understood. We investigated roles of 20-hydroxyecdysone (20E) and insulin/insulin-like signaling/target of rapamycin (IIS/TOR) signaling pathways in the effects of PvERR on larval development, metamorphosis, and adult growth in ant Polyrhachis vicina Roger. PvFOXO expression levels depended on caste and developmental stage. PvERR RNAi significantly reduced the expression levels of IIS/TOR signaling pathway genes and 20E signaling pathway genes in fourth-instar larvae, pupae, females, and workers and significantly increased the expression levels of IIS/TOR signaling pathway genes PvFOXO and PvAkt in males. PvFOXO RNAi resulted in developmental defects and increased mortality. After PvFOXO RNAi, the expression of PvERR, 20E signaling pathway genes, and IIS/TOR signaling pathway genes decreased significantly in pupae, females, and workers and increased significantly in fourth-instar larvae. Exogenous 20E attenuated expression changes induced by PvFOXO RNAi in a sex- and stage-specific manner. These results indicate that ERR interacts with 20E and IIS/TOR signaling pathways to regulate caste determination, metamorphosis, and male fertility in P. vicina and that correlations between PvERR and PvFOXO are caste- and stage-specific.

The First Complete Mitochondrial Genome of the Genus Pachycondyla (Formicidae, Ponerinae) and Insights into the Phylogeny of Ants.

Ants are the standout group among eusocial insects in terms of their exceptional species richness and ecological dominance. The phylogenetic relationships among the group remain elusive. Mitochondrial genome sequences, as a kind of molecular marker, have been widely utilized in the phylogenetic analysis of insects. However, the number of ant mitogenomes published is still very limited. In this study, we utilized next-generation sequencing to determine the complete mitogenome of Pachycondyla annamita (Formicidae, Ponerinae). This is the first mitogenome from the genus Pachycondyla. Two gene rearrangements were identified in the mitogenome, the transposition of trnQ and trnM and the transposition of trnV and rrnS. The secondary structures of tRNAs were predicted. The tRNA genes trnR and trnS1 lacked the dihydrouridine (DHU) arm, and the trnE lacked the TΨC (T) arm. Phylogenetic analyses of the mitochondrial protein-coding genes under maximum likelihood (ML) and Bayesian inference (BI) criteria resulted in conflicting hypotheses. BI analysis using amino acid data with the site-heterogeneous mixture model produced a tree topology congruent with previous studies. The Formicidae was subdivided into two main clades, namely the "poneroid" clade and the "formicoid" clade. A sister group relationship between Myrmicinae and Formicinae was recovered within the "formicoid" clade.