Anatomical changes of Tenebrio molitor and Tribolium castaneum during complete metamorphosis.

In holometabolous insects, extensive reorganisation of tissues and cells occurs at the pupal stage. The remodelling of the external exoskeleton and internal organs that intervenes during metamorphosis has been traditionally studied in many insect species based on histological or ultrastructural methods. This study demonstrates the use of synchrotron X-ray phase-contrast micro-computed tomography as a powerful, non-destructive tool for in situ morphological observation of anatomical structures at the pupal stage in two Tenebrionid beetles, i.e. Tribolium castaneum and Tenebrio molitor, known as important pests, as well as emerging and promising models in experimental biology. Virtual sections and three-dimensional reconstructions were performed on both males and females at early, intermediate, and late pupal stage. The dataset allowed us to observe the remodelling of the gut and nervous system as well as the shaping of the female and male reproductive system at different pupal ages in both mealworm and red flour beetles. Moreover, we observed that the timing and duration pattern of organ development varied between the species analysed, likely related to the species-specific adaptations of the pre-imaginal stages to environmental conditions, which ultimately affect their life cycle. This research provides new knowledge on the morphological modifications that occur during the pupal stage of holometabolous insects and provides a baseline set of information on beetle metamorphosis that may support future research in forensics, physiology, and ecology as well as an image atlas for educational purposes.

Circulating profile of the appetite-regulating hormone ghrelin during moult-fast and chick provisioning in southern rockhopper penguins (Eudyptes chrysocome chrysocome).

A multitude of animal species undergo prolonged fasting events at regularly occurring life history stages. During such periods of food deprivation, individuals need to suppress their appetite. The satiety signalling gut hormone ghrelin has received much attention in this context in studies looking at mammalian systems. In wild birds, however, knowledge on the ghrelin system and its role during extended fasts is still scarce. In this study, we collected plasma samples for measurements of circulating ghrelin concentrations from adult southern rockhopper penguins (Eudyptes chrysocome chrysocome) during the three to four week-long moult-fast that they repeat annually to replace their feathers. We further sampled chicks before and after feeding bouts and non-moulting adults. Circulating ghrelin levels did not differ significantly between fed and unfed chicks but chicks had significantly lower plasma ghrelin levels compared to adults. Furthermore, penguins in late moult (i.e. individuals at the end of the prolonged fasting bout) had higher ghrelin levels compared to non-moulting adults. Our results show elevated levels of circulating ghrelin during moult and generally lower levels of ghrelin in chicks than in adults regardless of feeding state. Given the scarcity or absence of knowledge on the function of ghrelin in seabirds and in fasting birds in general, our results add greatly to our understanding of the avian ghrelin system.

Changes in body surface temperature reveal the thermal challenge associated with catastrophic moult in captive gentoo penguins.

Once a year, penguins undergo a catastrophic moult, replacing their entire plumage during a fasting period on land or on sea-ice during which time individuals can lose 45% of their body mass. In penguins, new feather synthesis precedes the loss of old feathers, leading to an accumulation of two feather layers (double coat) before the old plumage is shed. We hypothesized that the combination of the high metabolism required for new feather synthesis and the potentially high thermal insulation linked to the double coat could lead to a thermal challenge requiring additional peripheral circulation to thermal windows to dissipate the extra heat. To test this hypothesis, we measured the surface temperature of different body regions of captive gentoo penguins (Pygoscelis papua) throughout the moult under constant environmental conditions. The surface temperature of the main body trunk decreased during the initial stages of the moult, suggesting greater thermal insulation. In contrast, the periorbital region, a potential proxy of core temperature in birds, increased during these same early moulting stages. The surface temperature of the bill, flipper and foot (thermal windows) tended to initially increase during the moult, highlighting the likely need for extra heat dissipation in moulting penguins. These results raise questions regarding the thermoregulatory capacities of penguins in the wild during the challenging period of moulting on land in the current context of global warming.

Evolutionary trade-off between male colouration and feather moult extent also indirectly determines female moult.

Males and females are often influenced by different selective forces, frequently resulting in diverging phenotypes, for example in colouration. Since an animal's colouration may strongly influence its fitness, causes and consequences of sexual dichromatism in birds could aid in understanding important factors affecting sexual and natural selection. Variation in plumage ornamentation may affect mate attraction or intraspecific antagonistic behaviour. In most passerines, body plumage colouration of juveniles is obtained through the process of feather moult. The number of moulted wing and tail feathers, which also influences the bird's appearance, may affect its fitness. Here, we show that body plumage colouration of male, but not female, passerines is correlated with the number of moulted wing and tail feathers in the early stage of the bird's life for both sexes. Thus, the extent of wing and tail moult in females is not modulated by the female's colouration and can prevent females from reaching their sex-specific optima. This result could be explained by high intersexual genetic correlations, which might make it impossible for the sexes to reach their own trait fitness optima. Our findings may indicate that species-specific, rather than sex-specific, internal correlations shaped bird moult strategy, an important avian life-history trait.

Differential changes in life cycle-event phenology provide a window into regional population declines.

Climate change affects the phenology of annual life cycle events of organisms, such as reproduction and migration. Shifts in the timing of these events could have important population implications directly, or provide information about the mechanisms driving population trajectories, especially if they differ between life cycle event. We examine if such shifts occur in a declining migratory passerine bird (willow warbler, Phylloscopus trochilus), which exhibits latitudinally diverging population trajectories. We find evidence of phenological shifts in breeding initiation, breeding progression and moult that differ across geographic and spring temperature gradients. Moult initiation following warmer springs advances faster in the south than in the north, resulting in proportionally shorter breeding seasons, reflecting higher nest failure rates in the south and in warmer years. Tracking shifts in multiple life cycle events allowed us to identify points of failure in the breeding cycle in regions where the species has negative population trends, thereby demonstrating the utility of phenology analyses for illuminating mechanistic pathways underlying observed population trajectories.

Colour moult phenology and camouflage mismatch in polymorphic populations of Arctic foxes.

Species that seasonally moult from brown to white to match snowy backgrounds become conspicuous and experience increased predation risk as snow cover duration declines. Long-term adaptation to camouflage mismatch in a changing climate might occur through phenotypic plasticity in colour moult phenology and or evolutionary shifts in moult rate or timing. Also, adaptation may include evolutionary shifts towards winter brown phenotypes that forgo the winter white moult. Most studies of these processes have occurred in winter white populations, with little attention to polymorphic populations with sympatric winter brown and winter white morphs. Here, we used remote camera traps to record moult phenology and mismatch in two polymorphic populations of Arctic foxes in Sweden over 2 years. We found that the colder, more northern population moulted earlier in the autumn and later in the spring. Next, foxes moulted earlier in the autumn and later in the spring during colder and snowier years. Finally, white foxes experienced relatively low camouflage mismatch while blue foxes were mismatched against snowy backgrounds most of the autumn through the spring. Because the brown-on-white mismatch imposes no evident costs, we predict that as snow duration decreases, increasing blue morph frequencies might help facilitate species persistence.

Varying aerobic capacity in relation to breeding stage and reproductive success in giant petrels (Macronectes spp.).

Reproduction, and parental care in particular, are among the most energy-demanding activities within the annual cycle of adult birds. Parents that cannot meet the metabolic demands and other physiological costs of raising offspring may opt to abandon chicks in favour of self-maintenance and future reproduction. Recent work examining reproductive trade-offs in birds revealed an important role of oxygen carrying capacity in mediating variation in parental effort. This study explores the aerobic factors underlying the success or failure of parental care in two closely-related petrel species during their breeding season on Bird Island, South Georgia: northern giant petrels (Macronectes halli) and southern giant petrels (M. giganteus). Failed breeders of both sexes and species had significantly lower hematocrit levels (by 5.48 ± 0.64%) than successful breeders, and reticulocyte counts also tended to be lower in failed males, consistent with the hypothesis that parental care and workload depend on aerobic capacity. We discuss these results in relation to differences in the foraging ecology of both species and sexes.

Metabolic heat loss in southern elephant seals (Mirounga leonina) differs with stage of moult and between habitats.

The moult in southern elephant seals (Mirounga leonina) represents an especially energetically demanding period during which seals must maintain high skin temperature to facilitate complete replacement of body fur and upper dermis. In this study, heat flux from the body surface was measured on 18 moulting southern elephant seals to estimate metabolic heat loss in three different habitats (beach, wallow and vegetation). Temperature data loggers were also deployed on 10 southern elephant seals to monitor skin surface temperature. On average, heat loss of animals on the beach was greater than in wallows or vegetation, and greater in wallows than in vegetation. Heat loss across all habitats during the moult equated to 1.8 x resting metabolic rate (RMR). The greatest heat loss of animals was recorded in the beach habitat during the late moult, that represented 2.3 x RMR. Mass loss was 3.6 ± 0.3 kg day-1, resulting in changes in body condition as the moult progressed. As body condition declined, skin surface temperature also decreased, suggesting that as animals approached the end of the moult blood flow to the skin surface was no longer required for hair growth.

A novel approach to co-expression network analysis identifies modules and genes relevant for moulting and development in the Atlantic salmon louse (Lepeophtheirus salmonis).

BACKGROUND: The salmon louse (Lepeophtheirus salmonis) is an obligate ectoparasitic copepod living on Atlantic salmon and other salmonids in the marine environment. Salmon lice cause a number of environmental problems and lead to large economical losses in aquaculture every year. In order to develop novel parasite control strategies, a better understanding of the mechanisms of moulting and development of the salmon louse at the transcriptional level is required. METHODS: Three weighted gene co-expression networks were constructed based on the pairwise correlations of salmon louse gene expression profiles at different life stages. Network-based approaches and gene annotation information were applied to identify genes that might be important for the moulting and development of the salmon louse. RNA interference was performed for validation. Regulatory impact factors were calculated for all the transcription factor genes by examining the changes in co-expression patterns between transcription factor genes and deferentially expressed genes in middle stages and moulting stages. RESULTS: Eight gene modules were predicted as important, and 10 genes from six of the eight modules have been found to show observable phenotypes in RNA interference experiments. We knocked down five hub genes from three modules and observed phenotypic consequences in all experiments. In the infection trial, no copepodids with a RAB1A-like gene knocked down were found on fish, while control samples developed to chalimus-1 larvae. Also, a FOXO-like transcription factor obtained highest scores in the regulatory impact factor calculation. CONCLUSIONS: We propose a gene co-expression network-based approach to identify genes playing an important role in the moulting and development of salmon louse. The RNA interference experiments confirm the effectiveness of our approach and demonstrated the indispensable role of a RAB1A-like gene in the development of the salmon louse. We propose that our approach could be generalized to identify important genes associated with a phenotype of interest in other organisms.

Effects of experimental increase in insulin-like growth factor 1 on feather growth rate, moult intensity and feather quality in a passerine bird.

Moulting is a crucial, yet often overlooked life-history stage in many animals, when they renew their integumental structures. This life-history stage is an energetically demanding somatic growth event that has particular importance in birds because feathers play a crucial role in flight, insulation and communication. Somatic growth processes are regulated by the evolutionarily conserved peptide hormone insulin-like growth factor 1 (IGF-1). However, the role of IGF-1 in feather growth remains unknown. In this study, we captured 41 juvenile free-living bearded reedlings (Panurus biarmicus) that had started their first complete moult and brought them into captivity. Then, we manipulated their circulating IGF-1 levels using poly-(lactic-co-glycolid acid) microparticles (microspheres) that provide a sustained release of IGF-1. The treatment increased IGF-1 levels but did not affect the feather growth rate. However, 2 weeks after the treatment, birds in the increased IGF-1 group were moulting more feathers simultaneously than the controls and were at a more advanced stage of moult. Birds with experimentally increased IGF-1 levels had better quality feathers (measured by a lower number of fault bars) than the controls. These results suggest that an increase in IGF-1 does not speed up feather growth, but may alter moult intensity by initiating the renewal of several feathers simultaneously. This may shorten the overall moulting time but may imply costs in terms of IGF-1-induced oxidative stress.

N-acetyl-ß-d-glucosaminidase activity in Palaemon serratus - Methodological optimisation and intrinsic variability.

Chitinolytic enzymes fulfil a key role in the moulting process of crustaceans, in degrading the endocuticle during apolysis. Measuring the enzyme activity is an interesting manner to monitor the moult process at sub-individual level, complementary to the classical observation of the integument morphogenesis, ecdysis success, or moult cycle duration. The present study aimed to optimise the methodology of using N-acetyl-ß-D-glucosaminidase (NAGase) activity to monitor moulting in the marine prawn Palaemon serratus, and to compare NAGase activity levels along the moult cycle of both male and female specimens. First, to optimise protocols for five different organs, different reaction medium compositions were tested, considering the type buffer, concentration of the substrate, and the load in enzymatic extract. Second, levels of NAGase activity were closely monitored during eight moulting stages in male prawns. Variations in NAGase activity were observed during the moult cycle, with an increase in activity in the late premoult phase of approximately 2.4-fold the level of the intermoult phase. This response profile was observed for each tested organ. The levels of NAGase activity of male and female specimens were compared during three stages of the premoult phase. The patterns observed for both sexes were similar for all the tested organs.

Mechanical and structural adaptations to migration in the flight feathers of a Palaearctic passerine.

Current avian migration patterns in temperate regions have been developed during the glacial retreat and subsequent colonization of the ice-free areas during the Holocene. This process resulted in a geographic gradient of greater seasonality as latitude increased that favoured migration-related morphological and physiological (co)adaptations. Most evidence of avian morphological adaptations to migration comes from the analysis of variation in the length and shape of the wings, but the existence of intra-feather structural adjustments has been greatly overlooked despite their potential to be under natural selection. To shed some light on this question, we used data from European robins Erithacus rubecula overwintering in Campo de Gibraltar (Southern Iberia), where sedentary robins coexist during winter with conspecifics showing a broad range of breeding origins and, hence, migration distances. We explicitly explored how wing length and shape, as well as several functional (bending stiffness), developmental (feather growth rate) and structural (size and complexity of feather components) characteristics of flight feathers, varied in relation to migration distance, which was estimated from the hydrogen stable isotope ratios of the summer-produced tail feathers. Our results revealed that migration distance not only favoured longer and more concave wings, but also promoted primaries with a thicker dorsoventral rachis and shorter barb lengths, which, in turn, conferred more bending stiffness to these feathers. We suggest that these intra-feather structural adjustments could be an additional, largely unnoticed, adaptation within the avian migratory syndrome that might have the potential to evolve relatively quickly to facilitate the occupation of seasonal environments.

Partial or complete? The evolution of post-juvenile moult strategies in passerine birds.

Moulting strategies in birds have evolved to avoid overlap with, or prepare for, other demanding parts of the annual cycle, such as reproduction or migration. When moulting for the first time after leaving the nest, young birds replace their typically poor-quality plumage during the post-juvenile moult. The extent of this moult varies between species from partial to complete. Earlier studies, restricted to Western Palearctic birds, suggest that in most species a complete post-juvenile moult may not be possible simply because young birds are constrained by not having the same access to resources as adults, unless environmental conditions are favourable. These studies also show that complete post-juvenile moult is more common in species with poor-quality nest-grown plumage. We expanded the spatial and taxonomic scope of previous studies to 1,315 species of passerines from across the world and considered both the role of constraints, plumage quality and other selective pressures favouring a complete post-juvenile moult. Thus, we test whether complete moult is more prevalent in species where nest-grown feathers are presumably of poor quality (shorter nestling period), that live in environments that foster quick plumage degradation (open habitats, high insolation and humidity), and where males are under strong sexual selection. Our data reveal that 24% of species carry out a complete post-juvenile moult, and that this trait has a strong phylogenetic signal. Complete moult is more common in species that inhabit warmer regions and open habitats, show no delayed plumage maturation and have higher levels of sexual dichromatism (indicative of strong sexual selection). Neither the presumed quality of the nest-grown plumage nor living in regions with high insolation correlates with complete moult. In conclusion, the evolution of complete post-juvenile moult not only depends on whether birds can perform a complete moult (i.e. suitable environmental conditions) but also on the strength of selection associated with the need of a complete moult. In particular, the necessity to keep the plumage in good condition in challenging environments and the benefits associated with producing adult-like plumage colours to attract mates or deter rivals seem to play an important role.

Evolution of moult-migration is directly linked to aridity of the breeding grounds in North American passerines.

To avoid energy allocation conflicts, birds generally separate breeding, migration and moult during the annual cycle. North American passerines typically moult on the breeding grounds prior to autumn migration. However, some have evolved a moult-migration strategy in which they delay moult until stopping over during autumn migration. Rohwer et al. (2005) proposed the 'push-pull hypothesis' as an explanation for the evolution of this moult strategy, but it has not been empirically tested. Poor conditions on the breeding grounds at the end of the summer would push birds to depart prior to moult, while productive stopover locations would pull them. We tested for a relationship between moult-migration and breeding grounds aridity as measured by the normalized difference vegetation index. Our results strongly support the 'push' aspect of the push-pull hypothesis and indicate that arid breeding grounds, primarily in western North America, would drive species to evolve stopover moult-migration, although this relationship may depend upon migration distance.

The effect of temperature on ability of Lepeophtheirus salmonis to infect and persist on Atlantic salmon.

The salmon louse (Lepeophtheirus salmonis) is an ecologically and economically important parasite of salmonid fish. Temperature is a strong influencer of biological processes in salmon lice, with development rate increased at higher temperatures. The successful attachment of lice onto a host is also predicted to be influenced by temperature; however, the correlation of temperature with parasite survival is unknown. This study describes the effects of temperature on infection success, and survival on the host during development to the adult stage. To accurately describe infection dynamics with varying temperatures, infection success was recorded on Atlantic salmon (Salmo salar) between 2 and 10°C. Infection success ranged from 20% to 50% and was strongly correlated with temperature, with the highest success at 10°C. Parasite loss was monitored during development at eight temperatures with high loss of lice at 3 and 24°C, whilst no loss was recorded in the temperature range from 6 to 21°C. Sea temperatures thus have large effects on the outcome of salmon louse infections and should be taken into account in the management and risk assessment of this parasite. Improving understanding of the infection dynamics of salmon lice will facilitate epidemiological modelling efforts and efficiency of pest management strategies.

Evolutionary drivers of seasonal plumage colours: colour change by moult correlates with sexual selection, predation risk and seasonality across passerines.

Some birds undergo seasonal colour change by moulting twice each year, typically alternating between a cryptic, non-breeding plumage and a conspicuous, breeding plumage ('seasonal plumage colours'). We test for potential drivers of the evolution of seasonal plumage colours in all passerines (N = 5901 species, c. 60% of all birds). Seasonal plumage colours are uncommon, having appeared on multiple occasions but more frequently lost during evolution. The trait is more common in small, ground-foraging species with polygynous mating systems, no paternal care and strong sexual dichromatism, suggesting it evolved under strong sexual selection and high predation risk. Seasonal plumage colours are also more common in species predicted to have seasonal breeding schedules, such as migratory birds and those living in seasonal climates. We propose that seasonal plumage colours have evolved to resolve a trade-off between the effects of natural and sexual selection on colouration, especially in seasonal environments.

Origin of ecdysis: fossil evidence from 535-million-year-old scalidophoran worms.

With millions of extant species, ecdysozoans (Scalidophora, Nematoida and Panarthropoda) constitute a major portion of present-day biodiversity. All ecdysozoans secrete an exoskeletal cuticle which must be moulted periodically and replaced by a larger one. Although moulting (ecdysis) has been recognized in early Palaeozoic panarthropods such as trilobites and basal groups such as anomalocaridids and lobopodians, the fossil record lacks clear evidence of ecdysis in early scalidophorans, largely because of difficulties in recognizing true exuviae. Here, we describe two types of exuviae in microscopic scalidophoran worms from the lowermost Cambrian Kuanchuanpu Formation ( ca 535 Ma) of China and reconstruct their moulting process. These basal scalidophorans moulted in a manner similar to that of extant priapulid worms, extricating themselves smoothly from their old tubular cuticle or turning their exuviae inside out like the finger of a glove. This is the oldest record of moulting in ecdysozoans. We also discuss the origin of ecdysis in the light of recent molecular analyses and the significance of moulting in the early evolution of animals.

Timing manipulations reveal the lack of a causal link across timing of annual-cycle stages in a long-distance migrant.

Organisms need to time their annual-cycle stages, like breeding and migration, to occur at the right time of the year. Climate change has shifted the timing of annual-cycle stages at different rates, thereby tightening or lifting time constraints of these annual-cycle stages, a rarely studied consequence of climate change. The degree to which these constraints are affected by climate change depends on whether consecutive stages are causally linked (scenario I) or whether the timing of each stage is independent of other stages (scenario II). Under scenario I, a change in timing in one stage has knock-on timing effects on subsequent stages, whereas under scenario II, a shift in the timing of one stage affects the degree of overlap with previous and subsequent stages. To test this, we combined field manipulations, captivity measurements and geolocation data. We advanced and delayed hatching dates in pied flycatchers (Ficedula hypoleuca) and measured how the timing of subsequent stages (male moult and migration) were affected. There was no causal effect of manipulated hatching dates on the onset of moult and departure to Africa. Thus, advancing hatching dates reduced the male moult-breeding overlap with no effect on the moult-migration interval. Interestingly, the wintering location of delayed males was more westwards, suggesting that delaying the termination of breeding carries over to winter location. Because we found no causal linkage of the timing of annual-cycle stages, climate change could shift these stages at different rates, with the risk that the time available for some becomes so short that this will have major fitness consequences.

Morph- and sex-specific effects of challenging conditions on maintenance parameters in the Gouldian finch.

Intraspecific discrete polymorphism is associated with the use of alternative life-history strategies, reflected by distinct reproductive or copying strategies in individuals of different morphs. Yet, morph-specific costs and benefits related to different life-history strategies remain unclear. Here, we examined in the polymorphic Gouldian finch (Erythrura gouldiae) whether markers of somatic maintenance (body mass, oxidative status and telomere length) differed between red- and black-headed birds under energetically demanding conditions (during heatwaves of different intensity, and during moult or breeding following heatwaves). During heatwaves, red-headed birds showed a homogeneous response, as males and females initially tended to gain mass and had reduced plasma hydroperoxide levels (a marker of oxidative damage) irrespective of heatwave intensity. In contrast, black-headed birds showed a stronger and more heterogeneous response, as black-headed males gained mass at the beginning of the thermoneutral heatwave and showed stable oxidative status, whereas black-headed females lost mass and tended to show higher hydroperoxide levels at the end of the thermocritical heatwave. Following heatwaves, we found morph-specific oxidative costs owing to moult or reproduction, with oxidative markers varying only in black-headed birds. Again, oxidative markers varied differently in black-headed males and females, as plasma antioxidant capacity decreased in moulting or breeding females, whereas males showed higher hydroperoxide levels with larger broods. For the first time, our study highlights that within polymorphic species, some individuals appear more vulnerable than others when coping with energetically demanding conditions. In the context of climate change, such differential effects may ultimately alter the currently observed balance between morphs and sexes within natural populations.

Differential gene transcription across the life cycle in Daphnia magna using a new all genome custom-made microarray.

BACKGROUND: Unravelling the link between genes and environment across the life cycle is a challenging goal that requires model organisms with well-characterized life-cycles, ecological interactions in nature, tractability in the laboratory, and available genomic tools. Very few well-studied invertebrate model species meet these requirements, being the waterflea Daphnia magna one of them. Here we report a full genome transcription profiling of D. magna during its life-cycle. The study was performed using a new microarray platform designed from the complete set of gene models representing the whole transcribed genome of D. magna. RESULTS: Up to 93% of the existing 41,317 D. magna gene models showed differential transcription patterns across the developmental stages of D. magna, 59% of which were functionally annotated. Embryos showed the highest number of unique transcribed genes, mainly related to DNA, RNA, and ribosome biogenesis, likely related to cellular proliferation and morphogenesis of the several body organs. Adult females showed an enrichment of transcripts for genes involved in reproductive processes. These female-specific transcripts were essentially absent in males, whose transcriptome was enriched in specific genes of male sexual differentiation genes, like doublesex. CONCLUSION: Our results define major characteristics of transcriptional programs involved in the life-cycle, differentiate males and females, and show that large scale gene-transcription data collected in whole animals can be used to identify genes involved in specific biological and biochemical processes.