Divergent RNA localisation patterns of maternal genes regulating embryonic patterning in the butterfly Pararge aegeria

The maternal effect genes responsible for patterning the embryo along the antero-posterior(AP) axis are broadly conserved in insects. The precise function of these maternal effect genes is the result of the localisation of their mRNA in the oocyte. The main developmental mechanisms involved have been elucidated in Drosophila melanogaster, but recent studies have shown that other insect orders often diverge in RNA localisation patterns. A recent study has shown that in the butterfly Pararge aegeria the distinction between blastodermal embryonic (i.e. germ band) and extra-embryonic tissue (i.e. serosa) is already specified in the oocyte during oogenesis in the ovariole, long before blastoderm cellularisation. To examine the extent by which a female butterfly specifies and patterns the AP axis within the region fated to be the germ band, and whether she specifies a germ plasm, we performed in situ hybridisation experiments on oocytes in P. aegeria ovarioles and on early embryos. RNA localisation of the following key maternal effect genes were investigated: caudal (cad),orthodenticle (otd), hunchback (hb) and four nanos (nos) paralogs, as well as TDRD7 a gene containing a key functional domain (OST-HTH/LOTUS) shared with oskar. TDRD7 was mainly confined to the follicle cells, whilst hb was exclusively zygotically transcribed. RNA of some of the nos paralogs, otd and cad revealed complex localisation patterns within the cortical region prefiguring the germ band (i.e. germ cortex). Rather interestingly, otd was localised within and outside the anterior of the germ cortex. Transcripts of nos-O formed a distinct granular ring in the middle of the germ cortex possibly prefiguring the region where germline stem cells form. These butterfly RNA localisation patterns are highly divergent with respect to other insects, highlighting the diverse ways in which different insect orders maternally regulate early embryogenesis of their offspring

Development on drought-stressed host plants affects life history, flight morphology and reproductive output relative to landscape structure

With global climate change, rainfall is becoming more variable. Predicting the responses of species to changing rainfall levels is difficult because, for example in herbivorous species, these effects may be mediated indirectly through changes in host plant quality. Furthermore, species responses may result from a simultaneous interaction between rainfall levels and other environmental variables such as anthropogenic land use or habitat quality. In this eco-evolutionary study, we examined how male and female Pararge aegeria (L.) from woodland and agricultural landscape populations were affected by the development on drought-stressed host plants. Compared with individuals from woodland landscapes, when reared on drought-stressed plants agricultural individuals had longer development times, reduced survival rates and lower adult body masses. Across both landscape types, growth on drought-stressed plants resulted in males and females with low forewing aspect ratios and in females with lower wing loading and reduced fecundity. Development on drought-stressed plants also had a landscape-specific effect on reproductive output; agricultural females laid eggs that had a significantly lower hatching success. Overall, our results highlight several potential mechanisms by which low water availability, via changes in host plant quality, may differentially influence P. aegeria populations relative to landscape structure

Seasonal environments drive convergent evolution of a faster pace-of-life in tropical butterflies

New ecological niches that may arise due to climate change can trigger diversification, but their colonisation often requires adaptations in a suite of life-history traits. We test this hypothesis in species-rich Mycalesina butterflies that have undergone parallel radiations in Africa, Asia, and Madagascar. First, our ancestral state reconstruction of habitat preference, using c. 85% of extant species, revealed that early forest-linked lineages began to invade seasonal savannahs during the late Miocene-Pliocene. Second, rearing replicate pairs of forest and savannah species from the African and Malagasy radiation in a common garden experiment, and utilising published data from the Asian radiation, demonstrated that savannah species consistently develop faster, have smaller bodies, higher fecundity with an earlier investment in reproduction, and reduced longevity, compared to forest species across all three radiations. We argue that time-constraints for reproduction favoured the evolution of a faster pace-of-life in savannah species that facilitated their persistence in seasonal habitats.Peer reviewe

Variation in life history and flight morphological traits in Speckled Wood (Pararge aegeria) butterflies infected with a baculovirus.

Sub-lethal impacts are known to affect the insect-host relationship and have an important role in describing host dynamics. The impact of sub-lethal infections of pathogens on life history traits of affected hosts has been understudied in natural or semi-natural systems. The Speckled Wood (Pararge aegeria) is a satyrine butterfly that is common in temperate zones and has been extensively used as a model system for evolutionary ecology studies. It is known that the deployment of the immune system within this species, as with other invertebrates, is energetically costly and may result in trade-offs with fitness-related traits. In this study, we investigated the sub-lethal effect of exposure to Autographa californica multiple nucleopolyhedrovirus (AcMNPV) on life history and flight morphological traits of P. aegeria. Larvae were inoculated with increasing doses of AcMNPV and measurements made of life history and flight morphology traits. Generally, larvae exposed to virus took longer to develop to pupae and larval mass acquisition per day was significantly reduced in viral exposed larvae. However, viral exposed larvae were able to attain the same pupal mass and their duration as pupae was the same as controls. Forewing length, forewing aspect ratio, dry thorax mass and forewing loading were related to sex and bioassay differences but there was no evidence of any viral impact on these measures. Adult male butterflies had significantly less basal wing melanisation when exposed to virus compared to control males but there was no difference between females. Implications for population dynamics of P. aegeria are discussed

Flight-induced transgenerational maternal effects influence butterfly offspring performance during times of drought

Maternal condition can generate resource-related maternal effects through differential egg provisioning that can negatively affect offspring performance especially when offspring growth occurs in stressful or sub-optimal environments. Using the Speckled Wood butterfly, Pararge aegeria (L.) we tested the hypothesis that repeated periods of intensive flight during female oviposition affects egg provisioning and reduces offspring performance when larval development occurs under stressful conditions on drought stressed host plants. We investigated whether (after controlling for egg size) maternal age and flight treatment resulted in changes in egg provisioning and whether this contributed to variation in offspring traits across life stages. Age-related changes in maternal condition were found to generate resource-related maternal effects that influenced offspring traits across all life stages. Flight-induced changes in maternal egg provisioning were found to have direct consequences for offspring development in the egg and larval stages.. There were significant interactive effects between maternal age and flight on larval development and growth. Compared to offspring from forced flight mothers, offspring from control (no forced flight) mothers that hatched from eggs laid early in the oviposition period (i.e. by younger mothers) had shorter larval development times and heavier pupal masses, suggesting that offspring from mothers in relatively good condition may be able to buffer some of the costs associated with growth on drought stressed host plants. Our multi-factor study demonstrates the importance of considering the various, and often interacting, mechanisms by which maternal effects may influence offspring performance in stressful environments

Why small is beautiful: wing colour is free from thermoregulatory constraint in the small lycaenid butterfly, Polyommatus icarus

We examined the roles of wing melanisation, weight, and basking posture in thermoregulation in Polyommatus Icarus, a phenotypically variable and protandrous member of the diverse Polyommatinae (Lycaenidae). Under controlled experimental conditions, approximating to marginal environmental conditions for activity in the field (= infrequent flight, long duration basking periods), warming rates are maximised with fully open wings and maximum body temperatures are dependent on weight. Variation in wing melanisation within and between sexes has no effect on warming rates; males and females which differ in melanisation had similar warming rates. Posture also affected cooling rates, consistent with cooling being dependent on convective heat loss. We hypothesise that for this small sized butterfly, melanisation has little or no effect on thermoregulation. This may be a factor contributing to the diversity of wing colours in the Polyommatinae. Because of the importance of size for thermoregulation in this small butterfly, requirements for attaining a suitable size to confer thermal stability in adults may also be a factor influencing larval feeding rates, development time and patterns of voltinism. Our findings indicate that commonly accepted views of the importance of melanisation, posture and size to thermoregulation, developed using medium and large sized butterflies, are not necessarily applicable to small sized butterflies

Ancient expansion of the Hox cluster in Lepidoptera generated four homeobox genes implicated in extra-embryonic tissue formation

Gene duplications within the conserved Hox cluster are rare in animal evolution, but in Lepidoptera an array of divergent Hox-related genes (Shx genes) has been reported between pb and zen. Here, we use genome sequencing of five lepidopteran species (Polygonia c-album, Pararge aegeria, Callimorpha dominula, Cameraria ohridella, Hepialus sylvina) plus a caddisfly outgroup (Glyphotaelius pellucidus) to trace the evolution of the lepidopteran Shx genes. We demonstrate that Shx genes originated by tandem duplication of zen early in the evolution of large clade Ditrysia; Shx are not found in a caddisfly and a member of the basally diverging Hepialidae (swift moths). Four distinct Shx genes were generated early in ditrysian evolution, and were stably retained in all descendent Lepidoptera except the silkmoth which has additional duplications. Despite extensive sequence divergence, molecular modelling indicates that all four Shx genes have the potential to encode stable homeodomains. The four Shx genes have distinct spatiotemporal expression patterns in early development of the Speckled Wood butterfly (Pararge aegeria), with ShxC demarcating the future sites of extraembryonic tissue formation via strikingly localised maternal RNA in the oocyte. All four genes are also expressed in presumptive serosal cells, prior to the onset of zen expression. Lepidopteran Shx genes represent an unusual example of Hox cluster expansion and integration of novel genes into ancient developmental regulatory networks

Historical and current patterns of gene flow in the butterfly Pararge aegeria

Aim. We have investigated the phylogeography and genetic structure of the Speckled Wood butterfly (Pararge aegeria) across its entire distribution range and studied its dispersal both on mainland and across sea straits. The apparent lack of gene flow between Sardinia and Corsica was further investigated by means of mating experiments. Location. Europe and North Africa. Methods. We sampled 345 individuals and sequenced one mitochondrial gene (Cytochrome c Oxidase subunit I, COI) for all samples and two nuclear genes (wingless and zerknullt) for a subset of the specimens. A total of 22 females from Corsica and Sardinia were used to establish a series of crosses to investigate reproductive compatibility and were screened for the presence of Wolbachia. Bayesian inference (BI) and haplotype networks were employed to infer phylogenetic relationships and a Principal Coordinate Analysis (PCoA) was used to represent 4 geographical patterns of genetic diversity. Mating and courtship data were analysed using linear mixed effect models. Results. We detected two main COI lineages separated by the Mediterranean Sea and maintained over relatively short sea straits. While nuclear gene variation was generally in agreement with that of COI, this was not the case in all areas (e.g. Iberian Peninsula and Corsica/Sardinia). Mating experiments revealed no evidence of reproductive isolation between the lineages, nor clear relation to Wolbachia infection status. Main conclusions. We propose that following the post-glacial recolonisation of Europe, the ancestral COI lineage of P. aegeria was maintained in North Africa and Mediterranean islands, while a new lineage colonised from Eastern Europe, replacing and apparently outcompeting the ancestral variant. Several hypotheses are discussed that may explain the local discordance between the nuclear genes and COI, including sex-specific dispersal, selection and differential rates of gene evolution

A Single Basis for Developmental Buffering of Drosophila Wing Shape

The nature of developmental buffering processes has been debated extensively, based on both theoretical reasoning and empirical studies. In particular, controversy has focused on the question of whether distinct processes are responsible for canalization, the buffering against environmental or genetic variation, and for developmental stability, the buffering against random variation intrinsic in developmental processes. Here, we address this question for the size and shape of Drosophila melanogaster wings in an experimental design with extensively replicated and fully controlled genotypes. The amounts of variation among individuals and of fluctuating asymmetry differ markedly among genotypes, demonstrating a clear genetic basis for size and shape variability. For wing shape, there is a high correlation between the amounts of variation among individuals and fluctuating asymmetry, which indicates a correspondence between the two types of buffering. Likewise, the multivariate patterns of shape variation among individuals and of fluctuating asymmetry show a close association. For wing size, however, the amounts of individual variation and fluctuating asymmetry are not correlated. There was a significant link between the amounts of variation between wing size and shape, more so for fluctuating asymmetry than for variation among individuals. Overall, these experiments indicate a considerable degree of shared control of individual variation and fluctuating asymmetry, although it appears to differ between traits