The genetic signature left by the range expansion of red foxes in Australia is detectable after more than 80 years of population stability.

Reconstructing biological invasions from historical sources can provide insights into how they occur but are difficult to do when invasions are poorly documented. Genetic signatures left by invaders can also offer insights into invasion routes, points of origin and general biology but often present conclusions that are contradictory to expectations. Here, we test the ability of continental-wide microsatellite genotype data from 29 loci and 3122 samples to reconstruct the well-documented invasion of red foxes Vulpes vulpes from the United Kingdom into Australia over 150 years ago, an invasion that has led to the extinction of many native species. Our analysis reveals several key signals of invasion evident in Australian foxes. They display lower levels of diversity than foxes sampled from the UK, exhibit clines in diversity from the point of introduction (south-east Australia) to the edge of their range, and show strong evidence of allele surfing in westerly and north-easterly directions. These characteristics are consistent with a single point of origin followed by rapid expansion in westerly and north-easterly directions as suggested by historical records. We also find little genetic structure in foxes across Australia with only the vast Nullarbor Plains and Great Victoria Desert region presenting a detectable barrier to their dispersal. As such, no mainland region within the current range of foxes can be considered genetically isolated and therefore appropriate for localised eradication efforts. Overall, our analyses demonstrate the ability of comprehensive population genetic studies to reconstruct invasion histories even after more than 80 years since colonisation was stabilised.

Working dogs in dynamic on-duty environments: The impact of dark adaptation, strobe lighting and acoustic distraction on task performance.

Sudden changes in sound and light (e.g., sirens and flashing police beacons) are a common component of working dogs' on-duty environment. Yet, how such stimuli impact dogs' ability to perform physical and cognitive tasks has not been explored. To address this shortcoming, we compared the accuracy and time taken for twelve dogs to complete a complex physical and cognitive task, before, during and after exposure to three 'real-world' stimuli: an acoustic distractor (85dB), white strobe lighting (5, 10 & 15 Hz), and exposure to a dazzling white, red, or blue lights. We found that strobe lighting, and to a greater extent, acoustic distraction, significantly reduced dogs' physical performance. Acoustic distraction also tended to impair dogs' cognitive performance. Dazzling lights had no effect on task performance. Most (nine out of twelve) dogs sensitised to the acoustic distraction to the extent of non-participation in the rewarded task. Our results suggest that without effective distractor response training, sudden changes in noise and flickering lights are likely to impede cognitive and physical task performance in working dogs. Repeated uncontrolled exposure may also amplify these effects.

Consumption of Rodenticide Baits by Invertebrates as a Potential Route into the Diet of Insectivores.

Non-target species are commonly exposed to anticoagulant rodenticides worldwide, which may pose a key threat to declining species. However, the main pathway of exposure is usually unknown, potentially hindering conservation efforts. This study aimed to examine whether baits mixed with the biomarker rhodamine B can be used to track invertebrate consumption of rodenticides in a field environment, using this to observe whether invertebrate prey are a potential vector for anticoagulant rodenticides in the diet of insectivores such as the European hedgehog (Erinaceus europaeus). Rhodamine B baits were found to create an observable response. Uptake was negligible in captured insects; however, 20.7% of slugs and 18.4% of snails captured showed uptake of bait. Maximum temperature, distance from bait, proximity to buildings, and the addition of copper tape to bait boxes all influenced the rate of bait uptake in molluscs. Based on these data, it seems likely that molluscs could be a source of rodenticide poisoning in insectivores. This research demonstrates which prey may pose exposure risks to insectivores and likely environmental factors, knowledge of which can guide effective mitigation measures. We suggest that further investigation into using mollusc repellents around bait boxes should be considered.

Identifying and locating carotenoids in supra-orbital combs of male black grouse (Lyurus tetrix) using Raman and transmission electron microscopy: A histological study using rehydrated tissue samples.

Colourful signals have long been implicated as indicators of individual quality in animals. Bare-skin signals are an understudied aspect of avian colouration compared with plumage studies, despite displaying rapid changes in size and colour in response to different environmental or physiological stressors. Even fewer studies have focused on the underlying histology of these structures and the importance this plays in the resulting skin colour. Using the Black Grouse (Lyrurus tetrix), we identified the underlying structure of individual dermal spikes, which make up the red supra-orbital comb (a known integumentary signal of male quality), and highlight visual structural differences between combs of different sizes. In addition, we used Raman spectroscopy to indicate the presence of carotenoids within the tissue, something that had previously only been inferred through characteristic reflectance patterns. An increased understanding of the structural basis of colour of featherless parts of the skin opens up exciting new avenues for interpreting the information content of integumentary signals.

Effects of hunting on genetic diversity, inbreeding and dispersal in Finnish black grouse (Lyrurus tetrix).

Intensive hunting activities such as commercial fishing and trophy hunting can have profound influences on natural populations. However, less intensive recreational hunting can also have subtle effects on animal behaviour, habitat use and movement, with implications for population persistence. Lekking species such as the black grouse (Lyrurus tetrix) may be especially prone to hunting as leks are temporally and spatially predictable, making them easy targets. Furthermore, inbreeding in black grouse is mainly avoided through female-biased dispersal, so any disruptions to dispersal caused by hunting could lead to changes in gene flow, increasing the risk of inbreeding. We therefore investigated the impact of hunting on genetic diversity, inbreeding and dispersal on a metapopulation of black grouse in Central Finland. We genotyped 1065 adult males and 813 adult females from twelve lekking sites (six hunted, six unhunted) and 200 unrelated chicks from seven sites (two hunted, five unhunted) at up to thirteen microsatellite loci. Our initial confirmatory analysis of sex-specific fine-scale population structure revealed little genetic structure in the metapopulation. Levels of inbreeding did not differ significantly between hunted and unhunted sites in neither adults nor chicks. However, immigration rates into hunted sites were significantly higher among adults compared to immigration into unhunted sites. We conclude that the influx of migrants into hunted sites may compensate for the loss of harvested individuals, thereby increasing gene flow and mitigating inbreeding. Given the absence of any obvious barriers to gene flow in Central Finland, a spatially heterogeneous matrix of hunted and unhunted regions may be crucial to ensure sustainable harvests into the future.

Biophysical Determinants and Constraints on Sperm Swimming Velocity.

Over the last 50 years, sperm competition has become increasingly recognised as a potent evolutionary force shaping male ejaculate traits. One such trait is sperm swimming speed, with faster sperm associated with increased fertilisation success in some species. Consequently, sperm are often thought to have evolved to be longer in order to facilitate faster movement. However, despite the intrinsic appeal of this argument, sperm operate in a different biophysical environment than we are used to, and instead increasing length may not necessarily be associated with higher velocity. Here, we test four predictive models (ConstantPower Density, Constant Speed, Constant Power Transfer, Constant Force) of the relationship between sperm length and speed. We collated published data on sperm morphology and velocity from 141 animal species, tested for structural clustering of sperm morphology and then compared the model predictions across all morphologically similar sperm clusters. Within four of five morphological clusters of sperm, we did not find a significant positive relationship between total sperm length and velocity. Instead, in four morphological sperm clusters we found evidence for the Constant Speed model, which predicts that power output is determined by the flagellum and so is proportional to flagellum length. Our results show the relationship between sperm morphology (size, width) and swimming speed is complex and that traditional models do not capture the biophysical interactions involved. Future work therefore needs to incorporate not only a better understanding of how sperm operate in the microfluid environment, but also the importance of fertilising environment, i.e., internal and external fertilisers. The microenvironment in which sperm operate is of critical importance in shaping the relationship between sperm length and form and sperm swimming speed.

Ear pinnae in a neotropical katydid (Orthoptera: Tettigoniidae) function as ultrasound guides for bat detection.

Early predator detection is a key component of the predator-prey arms race and has driven the evolution of multiple animal hearing systems. Katydids (Insecta) have sophisticated ears, each consisting of paired tympana on each foreleg that receive sound both externally, through the air, and internally via a narrowing ear canal running through the leg from an acoustic spiracle on the thorax. These ears are pressure-time difference receivers capable of sensitive and accurate directional hearing across a wide frequency range. Many katydid species have cuticular pinnae which form cavities around the outer tympanal surfaces, but their function is unknown. We investigated pinnal function in the katydid Copiphora gorgonensis by combining experimental biophysics and numerical modelling using 3D ear geometries. We found that the pinnae in C. gorgonensis do not assist in directional hearing for conspecific call frequencies, but instead act as ultrasound detectors. Pinnae induced large sound pressure gains (20-30 dB) that enhanced sound detection at high ultrasonic frequencies (>60 kHz), matching the echolocation range of co-occurring insectivorous gleaning bats. These findings were supported by behavioural and neural audiograms and pinnal cavity resonances from live specimens, and comparisons with the pinnal mechanics of sympatric katydid species, which together suggest that katydid pinnae primarily evolved for the enhanced detection of predatory bats.

Ecological and life-history correlates of erythrocyte size and shape in Lepidosauria.

Blood oxygen-carrying capacity is shaped both by the ambient oxygen availability as well as species-specific oxygen demand. Erythrocytes are a critical part of oxygen transport and both their size and shape can change in relation to species-specific life-history, behavioural or ecological conditions. Here, we test whether components of the environment (altitude), life history (reproductive mode, body temperature) and behaviour (diving, foraging mode) drive erythrocyte size variation in the Lepidosauria (lizards, snakes and rhynchocephalians). We collected data on erythrocyte size (area) and shape (L/W: elongation ratio) from Lepidosauria across the globe (N = 235 species). Our analyses show the importance of oxygen requirements as a driver of erythrocyte size. Smaller erythrocytes were associated with the need for faster delivery (active foragers, high-altitude species, warmer body temperatures), whereas species with greater oxygen demands (diving species, viviparous species) had larger erythrocytes. Erythrocyte size shows considerable cross-species variation, with a range of factors linked to the oxygen delivery requirements being major drivers of these differences. A key future aspect for study would include within-individual plasticity and how changing states, for example, pregnancy, perhaps alter the size and shape of erythrocytes in Lepidosaurs.

Floral ultraviolet absorbance area responds plastically to ultraviolet irradiance in Brassica rapa.

Solar ultraviolet (UV) radiation is known to have significant effects on the development and performance of plants, including flowers. In multiple species, UV-absorbing floral patterns are associated with environmental conditions such as the solar UV exposure they typically receive. However, it is not known whether plants can increase the UV-absorbing areas found on petals plastically when in a high-UV environment. We grew Brassica rapa at three different UV radiation intensities (control, low, and high) and under two exposure duration regimes. We removed petals from flowers periodically during the flowering period and measured the proportion of the petal that absorbed UV. UV-absorbing areas increased when plants were exposed to longer periods of UV radiation, and at high UV radiation intensities. UV-absorbing area of petals of the UV intensity treatments decreased over time in long exposure plants. This study demonstrates that flowers can potentially acclimate to different UV radiation intensities and duration of exposure through an increase in UV-absorbing areas even after a relatively short exposure time to UV. Such a rapid plastic response may be especially beneficial for dynamically changing UV conditions and in response to climate change.

Energetic Lifestyle Drives Size and Shape of Avian Erythrocytes.

The size and shape of red blood cells (erythrocytes) is determined by key life history strategies in vertebrates. They have a fundamental role to deliver oxygen to tissues, and their ability to do so is shaped by the tissue's need and their shape. Despite considerable interest in how other components of blood are shaped by ecology and life history, few studies have considered erythrocytes themselves. We tested how erythrocyte size and shape varied in relation to energetically demanding activities using a dataset of 631 bird species. We found that in general, birds undergoing greater activities such as long distance migration had smaller and more elongated cells, while those with greater male-male competition had smaller and rounder cells. Smaller, more elongated erythrocytes allow more rapid oxygenation/deoxygenation and support greater aerobic activity. The rounder erythrocytes found in species with strong male-male competition may stem from younger erythrocytes deriving from androgen-induced erythropoiesis rates. Finally, diving species of bird had larger erythrocytes, indicating that erythrocytes are acting as a vital oxygen store. In summary, erythrocyte size and shape in birds are driven by the need to deliver oxygen during energetically costly activities.

Differentiation between left and right wing stridulatory files in the field cricket Gryllus bimaculatus (Orthoptera: Gryllidae).

Male crickets produce acoustic signals by wing stridulation, attracting females for mating. A plectrum on the left forewing's (or tegmen) anal margin rapidly strikes along a serrated vein (stridulatory file, SF) on the opposite tegmen as they close, producing vibrations, ending in a tonal sound. The tooth strike rate of the plectrum across file teeth is equal to the sound frequency produced by the cricket (i.e., ∼5k teeth/s for ∼5 kHz in field crickets) and is specific to the forewing's resonant frequency. Sound is subsequently amplified using specialised wing cells. Anatomically, the forewings appear to mirror each other: both tegmina bear a SF and plectrum; however, most cricket species stridulate using right-over-left wing overlap making the stridulatory mechanism asymmetrical by default, rendering the left tegmen's SF unused. Therefore, we hypothesised structural differences between functional and unfunctional SFs. Three-dimensional mapping was used to accurately measure SF structures in Gryllus bimaculatus wings. We found that the left SF shows significantly greater variation in inter-tooth distance than the right, but less variation within the first sixty teeth (the functional part) than the right file. The left SF's slow evolutionary change over millions of years is discussed considering modern molecular phylogenies and fossil records.

A narrow ear canal reduces sound velocity to create additional acoustic inputs in a microscale insect ear.

Located in the forelegs, katydid ears are unique among arthropods in having outer, middle, and inner components, analogous to the mammalian ear. Unlike mammals, sound is received externally via two tympanic membranes in each ear and internally via a narrow ear canal (EC) derived from the respiratory tracheal system. Inside the EC, sound travels slower than in free air, causing temporal and pressure differences between external and internal inputs. The delay was suspected to arise as a consequence of the narrowing EC geometry. If true, a reduction in sound velocity should persist independently of the gas composition in the EC (e.g., air, [Formula: see text]). Integrating laser Doppler vibrometry, microcomputed tomography, and numerical analysis on precise three-dimensional geometries of each experimental animal EC, we demonstrate that the narrowing radius of the EC is the main factor reducing sound velocity. Both experimental and numerical data also show that sound velocity is reduced further when excess [Formula: see text] fills the EC. Likewise, the EC bifurcates at the tympanal level (one branch for each tympanic membrane), creating two additional narrow internal sound paths and imposing different sound velocities for each tympanic membrane. Therefore, external and internal inputs total to four sound paths for each ear (only one for the human ear). Research paths and implication of findings in avian directional hearing are discussed.

Income and capital breeding in males: energetic and physiological limitations on male mating strategies.

Income and capital breeding describe two dichotomous breeding strategies that characterise the allocation of resources to reproduction. Capital breeders utilise stored endogenous resources (typically lipids) to finance reproduction, whereas income breeders use exogenous resources (typically carbohydrates). The basis for such characterisation has mainly come from studying females, yet for many species, male and female reproductive success may be determined by substantially different factors. Females allocate resources to offspring production, whereas males typically allocate resources to accessing mating opportunities, e.g. from contests or displays. The primary metabolic fuel (lipids or carbohydrates) in males appears to be dependent on the type of activity being performed (i.e. high versus low intensity or long versus short duration), rather than capital or income breeding strategy per se. Males performing sustained, long-duration effort typically utilise lipids, whereas those undergoing intense activity more often utilise carbohydrates. As a result, either fuel type can be used in either strategy. Breeding season duration can constrain strategy choice; lipids and carbohydrates can be used in short breeding season species, but only lipids provide a viable fuel source for long breeding season capital breeders. Both capital- and income-breeding males must manage their resource use during the breeding season, but capital breeders must also cope with physiological stressors associated with extended fasting. Overall, the capital-income breeding concept applies equally to male reproduction, but compared with females, there are different physical and physiological constraints that shape choice of strategy. This Commentary also highlights some key future areas that need to be investigated to further understand how capital-income breeding strategies shape male mating strategies.

Age- and quality-dependent DNA methylation correlate with melanin-based coloration in a wild bird.

Secondary sexual trait expression can be influenced by fixed individual factors (such as genetic quality) as well as by dynamic factors (such as age and environmentally induced gene expression) that may be associated with variation in condition or quality. In particular, melanin-based traits are known to relate to condition and there is a well-characterized genetic pathway underpinning their expression. However, the mechanisms linking variable trait expression to genetic quality remain unclear. One plausible mechanism is that genetic quality could influence trait expression via differential methylation and differential gene expression. We therefore conducted a pilot study examining DNA methylation at a candidate gene (agouti-related neuropeptide: AgRP) in the black grouse Lyrurus tetrix. We specifically tested whether CpG methylation covaries with age and multilocus heterozygosity (a proxy of genetic quality) and from there whether the expression of a melanin-based ornament (ultraviolet-blue chroma) correlates with DNA methylation. Consistent with expectations, we found clear evidence for age- and heterozygosity-specific patterns of DNA methylation, with two CpG sites showing the greatest DNA methylation in highly heterozygous males at their peak age of reproduction. Furthermore, DNA methylation at three CpG sites was significantly positively correlated with ultraviolet-blue chroma. Ours is the first study to our knowledge to document age- and quality-dependent variation in DNA methylation and to show that dynamic sexual trait expression across the lifespan of an organism is associated with patterns of DNA methylation. Although we cannot demonstrate causality, our work provides empirical support for a mechanism that could potentially link key individual factors to variation in sexual trait expression in a wild vertebrate.

Stabilising selection on immune response in male black grouse Lyrurus tetrix.

Illnesses caused by a variety of micro- and macro- organisms can negatively affect individuals' fitness, leading to the expectation that immunity is under positive selection. However, immune responses are costly and individuals must trade-off their immune response with other fitness components (e.g. survival or reproductive success) meaning that individuals with intermediate response may have the greatest overall fitness. Such a process might be particularly acute in species with strong sexual selection because the condition-dependence of male secondary sexual-traits might lead to striking phenotypic differences amongst males of different immune response levels. We tested whether there is selection on immune response by survival and reproduction in yearling and adult male black grouse (Lyrurus tetrix) following an immune challenge with a novel antigen and tested the hypothesis that sexual signals and body mass are honest signals of the immune response. We show that yearling males with highest immune response to these challenges had higher survival, but the reverse was true for adults. Adults with higher responses had highest mass loss and adult males with intermediate immune response had highest mating success. Tail length was related to baseline response in adults and more weakly in yearlings. Our findings reveal the complex fitness consequences of mounting an immune response across age classes. Such major differences in the direction and magnitude of selection in multiple fitness components is an alternative route underpinning the stabilising selection of immune responses with an intermediate immune response being optimal.

Structural biomechanics determine spectral purity of bush-cricket calls.

Bush-crickets (Orthoptera: Tettigoniidae) generate sound using tegminal stridulation. Signalling effectiveness is affected by the widely varying acoustic parameters of temporal pattern, frequency and spectral purity (tonality). During stridulation, frequency multiplication occurs as a scraper on one wing scrapes across a file of sclerotized teeth on the other. The frequency with which these tooth-scraper interactions occur, along with radiating wing cell resonant properties, dictates both frequency and tonality in the call. Bush-cricket species produce calls ranging from resonant, tonal calls through to non-resonant, broadband signals. The differences are believed to result from differences in file tooth arrangement and wing radiators, but a systematic test of the structural causes of broadband or tonal calls is lacking. Using phylogenetically controlled structural equation models, we show that parameters of file tooth density and file length are the best-fitting predictors of tonality across 40 bush-cricket species. Features of file morphology constrain the production of spectrally pure signals, but systematic distribution of teeth alone does not explain pure-tone sound production in this family.

Paternal arbuscular mycorrhizal fungal status affects DNA methylation in seeds.

Most land plants grow in association with arbuscular mycorrhizal fungi (AMF) in their roots and these fungi can cause transgenerational effects on plants' offspring. These may be caused by changes in DNA methylation of the offspring. In this study, we compared the amount of global DNA methylation in seeds of the gynodioecious plant Geranium sylvaticum in relation to the gender and the AMF status of the parents producing the seeds. The amount of DNA methylated was positively related to seed mass. Seeds produced by females had a similar proportion of methylated DNA regardless of the AMF status of the father siring the seed. By contrast, seeds from hermaphrodites had higher DNA methylation when sired by AMF fathers. We show to the best of our knowledge for the first time, that the AMF status of fathers can affect DNA methylation in seeds and that these changes in DNA methylation are further dependent on the gender of the mother producing the seeds.

Non-invasive biophysical measurement of travelling waves in the insect inner ear.

Frequency analysis in the mammalian cochlea depends on the propagation of frequency information in the form of a travelling wave (TW) across tonotopically arranged auditory sensilla. TWs have been directly observed in the basilar papilla of birds and the ears of bush-crickets (Insecta: Orthoptera) and have also been indirectly inferred in the hearing organs of some reptiles and frogs. Existing experimental approaches to measure TW function in tetrapods and bush-crickets are inherently invasive, compromising the fine-scale mechanics of each system. Located in the forelegs, the bush-cricket ear exhibits outer, middle and inner components; the inner ear containing tonotopically arranged auditory sensilla within a fluid-filled cavity, and externally protected by the leg cuticle. Here, we report bush-crickets with transparent ear cuticles as potential model species for direct, non-invasive measuring of TWs and tonotopy. Using laser Doppler vibrometry and spectroscopy, we show that increased transmittance of light through the ear cuticle allows for effective non-invasive measurements of TWs and frequency mapping. More transparent cuticles allow several properties of TWs to be precisely recovered and measured in vivo from intact specimens. Our approach provides an innovative, non-invasive alternative to measure the natural motion of the sensilla-bearing surface embedded in the intact inner ear fluid.

Sexually-trimorphic interactions with colour polymorphism determine nectar quality in a herbaceous perennial.

Amongst gynodioecious plant breeding systems, there can exist intermediate morphs with a reduction in their male function (i.e. reduced number of functional anthers). Along with this sexual trimorphism, plants can also show floral colour polymorphism. Such intricate mixtures of phenotypes within a species may have complex effects on floral rewards. Floral rewards are known to vary between sexually dimorphic species and to a lesser extent between colour morphs. However, the interactive effect of sexual trimorphism and colour polymorphism is unexplored. We measured nectar's sugar content in the sexually trimorphic Geranium sylvaticum, a gynodioecious plant with a light/dark floral polymorphism. We found that nectar reward differed across genders and colour morphs. Results were not however consistent within the three genders; dark female and hermaphrodite flowers had higher sugar content than light morphs, whereas intermediate flowers did not. As expected, females and hermaphrodites had different nectar reward, with intermediate morphs being midway between the other genders. In intermediates, the sugar content was not related to the number of functional stamens. We show for the first time the existence of sex-specific differences between flower gender and colour morphs in nectar rewards. Our results demonstrate the importance of considering multiple and conflicting selection pressures to explain rewards.

Auditory mechanics in a bush-cricket: direct evidence of dual sound inputs in the pressure difference receiver.

The ear of the bush-cricket, Copiphora gorgonensis, consists of a system of paired eardrums (tympana) on each foreleg. In these insects, the ear is backed by an air-filled tube, the acoustic trachea (AT), which transfers sound from the prothoracic acoustic spiracle to the internal side of the eardrums. Both surfaces of the eardrums of this auditory system are exposed to sound, making it a directionally sensitive pressure difference receiver. A key feature of the AT is its capacity to reduce the velocity of sound propagation and alter the acoustic driving forces at the tympanum. The mechanism responsible for reduction in sound velocity in the AT remains elusive, yet it is deemed to depend on adiabatic or isothermal conditions. To investigate the biophysics of such multiple input ears, we used micro-scanning laser Doppler vibrometry and micro-computed X-ray tomography. We measured the velocity of sound propagation in the AT, the transmission gains across auditory frequencies and the time-resolved mechanical dynamics of the tympanal membranes in C. gorgonensis Tracheal sound transmission generates a gain of approximately 15 dB SPL, and a propagation velocity of ca 255 m s-1, an approximately 25% reduction from free field propagation. Modelling tracheal acoustic behaviour that accounts for thermal and viscous effects, we conclude that reduction in sound velocity within the AT can be explained, among others, by heat exchange between the sound wave and the tracheal walls.