Genetic footprints of a rapid and large-scale range expansion: the case of cyclic common vole in Spain.

In the Anthropocene, many species are rapidly shifting their ranges in response to human-driven habitat modifications. Studying patterns and genetic signatures of range shifts helps to understand how species cope with environmental disturbances and predict future shifts in the face of global environmental change. We investigated the genetic signature of a contemporary wide-range expansion observed in the Iberian common vole Microtus arvalis asturianus shortly after a colonization event. We used mtDNA and microsatellite data to investigate patterns of genetic diversity, structure, demography, and gene flow across 57 localities covering the historical range of the species and the newly colonized area. The results showed a genetic footprint more compatible with a true range expansion (i.e. the colonization of previously unoccupied areas), than with a model of "colonization from within" (i.e. local expansions from small, unnoticed populations). Genetic diversity measures indicated that the source population was likely located at the NE of the historical range, with a declining gradient of genetic diversity towards the more recently invaded areas. At the expansion front, we observed the greatest gene flow and smallest pairwise differences between nearby localities. Both natural landscape features (rivers) and recent anthropogenic barriers (roads, railways) explained a large proportion of genetic variance among populations and had a significant impact on the colonization pathways used by voles.

Direct evidence of poison-driven widespread population decline in a wild vertebrate.

Toxicants such as organochlorine insecticides, lead ammunition, and veterinary drugs have caused severe wildlife poisoning, pushing the populations of several apex species to the edge of extinction. These prime cases epitomize the serious threat that wildlife poisoning poses to biodiversity. Much of the evidence on population effects of wildlife poisoning rests on assessments conducted at an individual level, from which population-level effects are inferred. Contrastingly, we demonstrate a straightforward relationship between poison-induced individual mortality and population changes in the threatened red kite (Milvus milvus). By linking field data of 1,075 poisoned red kites to changes in occupancy and abundance across 274 sites (10 × 10-km squares) over a 20-y time frame, we show a clear relationship between red kite poisoning and the decline of its breeding population in Spain, including local extinctions. Our results further support the species listing as endangered, after a breeding population decline of 31% to 43% in two decades of this once-abundant raptor. Given that poisoning threatens the global populations of more than 2,600 animal species worldwide, a greater understanding of its population-level effects may aid biodiversity conservation through increased regulatory control of chemical substances. Our results illustrate the great potential of long-term and large-scale on-ground monitoring to assist in this task.

Enhancing monitoring and transboundary collaboration for conserving migratory species under global change: The priority case of the red kite.

Calls for urgent action to conserve biodiversity under global change are increasing, and conservation of migratory species in this context poses special challenges. In the last two decades the Convention on the Conservation of Migratory Species of Wild Animals (CMS) has provided a framework for several subsidiary instruments including action plans for migratory bird species, but the effectiveness and transferability of these plans remain unclear. Such laws and policies have been credited with positive outcomes for the conservation of migratory species, but the lack of international coordination and on-ground implementation pose major challenges. While research on migratory populations has received growing attention, considerably less emphasis has been given to integrating ecological information throughout the annual cycle for examining strategies to conserve migratory species at multiple scales in the face of global change. We fill this gap through a case study examining the ecological status and conservation of a migratory raptor and facultative scavenger, the red kite (Milvus milvus), whose current breeding range is limited to Europe and is associated with agricultural landscapes and restricted to the temperate zone. Based on our review, conservation actions have been successful at recovering red kite populations within certain regions. Populations however remain depleted along the southern-most edge of the geographic range where many migratory red kites from northern strongholds overwinter. This led us to a forward-looking and integrated strategy that emphasizes international coordination involving researchers and conservation practitioners to enhance the science-policy-action interface. We identify and explore key issues for conserving the red kite under global change, including enhancing conservation actions within and outside protected areas, recovering depleted populations, accounting for climate change, and transboundary coordination in adaptive conservation and management actions. The integrated conservation strategy is sufficiently general such that it can be adapted to inform conservation of other highly mobile species subject to global change.

The fractal dimension of a conspicuous ornament varies with mating status and shows assortative mating in wild red-legged partridges (Alectoris rufa).

Complex body designs, such as plumage ornaments in birds, can be described by fractal geometry. These complex patterns could have a role as visual signals during courtship and social interactions, but an empirical validation in the wild is currently lacking. Here, we investigated whether the fractal dimension (FD) of a complex plumage pattern displayed by red-legged partridges Alectoris rufa could function as a potential sexual signal. We captured wild birds early in the breeding season and tested if mated and unmated birds differed in the FD of their conspicuous melanin-based black bib. We also tested if the FD of the black bib was correlated within the pair, looking for evidence of assortative mating based on the expression of this trait. We simultaneously assessed similar effects in other ornamental traits (black bib size, white throat patch and black flank band surface, redness of the eye rings and bill). Mated birds showed higher black bib FD values than unmated ones. Mated males, but not females, also displayed a larger black bib. Moreover, the black bib FD (but not the trait size) and the white throat patch surface showed assortative mating. Finally, females with higher black bib FD showed smaller black flank band surface, suggesting a trade-off in the expression of the two melanin-pigmented plumage traits. This provides unique and novel indication for the shape complexity of a pigmented trait, here described by its fractal dimension, to be potentially under sexual selection in a wild animal.

Changes in behaviour and faecal glucocorticoid levels in response to increased human activities during weekends in the pin-tailed sandgrouse.

Human recreational activities are becoming increasingly widespread and frequent, a fact that may potentially exacerbate their effects on wildlife. These human-related disturbances on animals may induce behavioural and physiological changes that can ultimately affect their fitness, showing a similar anti-predator response that against natural predator or other threats. Here, we combine the use of behavioural and physiological approaches to assess the potential effect of winter human activities on a threatened farmland bird in Europe, the pin-tailed sandgrouse (Pterocles alchata). We compared before, during and after weekend variations in human activity rates, pin-tailed sandgrouse behaviour (flocking and flying behaviour, interspecific association in mixed flocks and habitat use) and faecal glucocorticoid metabolite concentrations. Human disturbances, in particular those associated with hunting activities, peaked during weekends. Sandgrouse showed significant behavioural changes (increased sandgrouse-only flock sizes, increased proportion of birds flying and changes in habitat use) during weekends and higher faecal glucocorticoid metabolite concentrations after the weekends compared with during or before weekends. Therefore, physiological stress levels could be modulated by behavioural adjustments such as increased flock sizes and changes in habitat use that may allow sandgrouse to cope with increased human disturbance rates during weekends. Nevertheless, temporal and spatial organization of hunting days among groups of estates might be good strategies to buffer these potential adverse effects on wintering pin-tailed sandgrouse and other steppe species of conservation concern, while preserving a socio-economically important activity such as hunting.

Detection of methicillin-resistant Staphylococcus aureus (MRSA) carrying the mecC gene in wild small mammals in Spain.

OBJECTIVES: To determine the rate of Staphylococcus aureus faecal carriage in 101 wild small mammals in Spain and to characterize the isolates obtained. METHODS: Faecal samples were seeded on mannitol salt agar and ORSAB plates. The presence of the resistance genes mecA, mecC and blaZ and the new blaZ allotype associated with staphylococcal cassette chromosome mec (SCCmec) XI (blaZ-SCCmecXI) was studied by PCR. S. aureus isolates were characterized by spa typing, agr typing and multilocus sequence typing. The presence of immune evasion cluster (IEC) genes and virulence genes was analysed by PCR. RESULTS: S. aureus was detected in 13/101 studied faecal samples and one isolate per positive sample was further studied. Two S. aureus isolates were methicillin-resistant S. aureus (MRSA) (recovered from wood mice, Apodemus sylvaticus) and 11 were methicillin-susceptible S. aureus (MSSA). Both MRSA isolates harboured the mecC gene and the novel blaZ-SCCmecXI, were typed as spa-t1535/agrIII/ST1945(CC130)/SCCmecXI (where ST stands for sequence type and CC stands for clonal complex), carried the exfoliative toxin etd2 gene and were IEC type E. Eight different spa types were identified among the 11 MSSA isolates (five new) and six different sequence types were identified (two new). All MSSA strains were susceptible to the antibiotics tested except one blaZ-positive penicillin-resistant isolate (spa-t120/agrII/ST15). MSSA isolates were ascribed to the CCs (number of strains) CC5 (1), CC1956 (4) and singleton (6). Nine of 11 MSSA isolates carried the cna virulence gene. Only one MSSA isolate carried IEC genes (type C). CONCLUSIONS: This is the first report of MRSA carrying mecC in faecal samples of wild small mammals in Spain. These resistant isolates carried genes of the IEC system, unusual in S. aureus from animals. Wild small mammals could be a reservoir of the mecC gene with important implications for public health.

Niche-habitat mechanisms and biotic interactions explain the coexistence and abundance of congeneric sandgrouse species.

Ascertaining which niche processes allow coexistence between closely related species is of special interest in ecology. We quantified variations in the environmental niches and densities of two congeneric species, the pin-tailed and the black-bellied sandgrouse (Pterocles alchata and Pterocles orientalis) in allopatry and sympatry under similar abiotic, habitat and dispersal contexts to understand their coexistence. Using principal component analysis, we defined environmental gradients (niche dimensions) including abiotic, habitat and anthropogenic variables, and calculated niche breadth, position and overlap of both species in sympatry and allopatry. Additionally, sandgrouse density was modelled as a function of the niche dimensions and the density of the other species. We found evidence that each species occupies distinct environmental niches in sympatry and in allopatry. The black-bellied sandgrouse exploits a broader range of environmental conditions (wider niche breadth) while the pin-tailed sandgrouse reaches high densities where conditions seem to match its optimum. In sympatry, both species shift their niches to intermediate positions, indicating the importance of abiotic factors in setting coexistence areas. Environmental conditions determine regional densities of pin-tailed sandgrouse whereas biotic interactions explain the density of the black-bellied sandgrouse in areas with abiotic conditions similarly conducive for both species. Highly suitable areas for the pin-tailed sandgrouse fall beyond the upper thermal limit of the black-bellied sandgrouse, leading to niche segregation and low densities for the latter. Finally, local niche shift and expansion plus possible heterospecific aggregation allow the pin-tailed sandgrouse to thrive in a priori less favourable environments. This work provides insight into how different mechanisms allow species coexistence and how species densities vary in sympatry compared to allopatry as a result of environmental filtering and biotic interactions.

Habitat type modulates sharp body mass oscillations in cyclic common vole populations.

Cyclic rodent populations exhibit pronounced changes in body mass associated with the population cycle phase, long-known as Chitty effect. Although Chitty effect is a common epiphenomenon in both America and Europe, there is still incomplete evidence about the generality of these patterns across the entire range of most species. Moreover, despite decades of research, the underlying factors driving Chitty effect remains poorly understood. Here, we examined the influence of intrinsic and extrinsic factors that may underlie observed patterns in vole size variation in the Iberian common vole Microtus arvalis asturianus. We weighed and measured 2816 adult voles that were captured during 6 trapping periods. Vole numbers and body mass showed strong period- and phase-related variation both in females and males, demonstrating marked Chitty effect in the studied population. Body mass of adult males correlated with body length, evidencing that heavier males are also structurally larger. Statistical models showed that probability of occurrence of large-sized vole (> 37 g) was significantly more likely in reproductive males, during increase and peak phases, and it was modulated by habitat, with crop fields and field margins between crops showing an increased likelihood. We suggest an effect of the habitat on vole body mass mediated by predation.

Comparing sagittal heights calculated using corneal parameters and those measured with profilometry.

PURPOSE: To compare the sagittal height of the anterior eye (OC-SAG) calculated using corneal parameters with the OC-SAG measured by profilometry. METHOD: Seventy right eyes of soft contact lens wearers measured with the ESP (Eaglet Eye, The Netherlands) after lens removal were retrospectively analyzed for this study. The OC-SAG of the eyes was calculated using mean k-values, eccentricity and the inner (corneal) radius obtained with the ESP for an 11-mm cord diameter. It was then extrapolated to chord diameters of 14, 14.5 and 15 mm. These values were compared with OC-SAG values obtained with the ESP for the same chord diameters. Additionally, the OC-SAG was calculated through the formula used by a lab that manufactures custom soft lenses (mark'ennovy, Madrid, Spain) and compared again with the values obtained using the ESP. RESULTS: Differences between calculated OC-SAG obviating the shape factor were 121 ± 44, 155 ± 105, 172 ± 117 and 189 ± 129 µm for chord diameters of 11, 14, 14.5 and 15 mm, respectively (p < 0.001). When the shape factor was included in the calculation, differences were 28 ± 48, 62 ± 102, 79 ± 113 and 96 ± 123 µm (p < 0.001). When the inner best fit sphere was used to estimate OC-SAG, differences were 34 ± 11, 0 ± 72, 17 ± 86 and 34 ± 99, respectively, with no significant differences for the 14 and 14.5 mm-chord diameters (p = 0.99 and 0.11, respectively). Correlation coefficients between OC-SAG calculated and measured OC-SAG ranged from 0.53 to 0.90 depending on the chord diameter used. When the mark'ennovy formula was used to calculate the OC-SAG as the lens diameter proposed by the formula, the difference was -47 ± 147 µm (p < 0.01). CONCLUSIONS: Differences between the OC-SAG calculated using corneal parameters and that measured with a profilometer are statistically and clinically significant, especially for large chord diameters. The impact of this on contact lens fitting should be addressed in future studies.

Local Effects of Nest-Boxes for Avian Predators over Common Vole Abundance during a Mid-Density Outbreak.

At the end of the 20th century, the common vole (Microtus arvalis) colonized the practical totality of agricultural ecosystems in the northern sub-plateau of the Iberian Peninsula. To prevent crop damage, chemical control campaigns using anticoagulant rodenticides have been employed. This approach has a high environmental impact, and it has been banned in most countries in the European Union, including Spain. It is therefore essential to analyze alternative methods with lower environmental impacts. Here we explored the efficacy of biological control by avian predators to reduce vole abundance by providing nest-boxes in croplands. We used an indirect index based on the presence/absence of vole activity signs to measure the effect of nest-boxes on common vole abundance. We found that vole abundance was significantly lower near occupied nest-boxes at distances less than 180 m, where vole abundance increases progressively with increasing distance to the nearest nest-box. We also observed that the predatory pressure negatively affects the vole abundance at the end of the breeding period, considering the total number of fledglings. However, the effect of nest-boxes was highly variable depending on the study area and more limited in alfalfa fields, the optimal habitat for voles in agrarian ecosystems. Thus, nest-box supplementation would be a feasible measure for the biological control of the common vole in Mediterranean ecosystems, but it needs improvements for vole control in alfalfa fields within an integrated pest control program. We provide several recommendations to improve the performance of biological control in alfalfa fields.

What we know about the scleral profile and its impact on contact lens fitting.

Scleral contact lens fitting has provided practitioners with one of the earliest pieces of evidence that the sclera was more often asymmetric than symmetric. Some preliminary observations such as different haptic alignment patterns over the sclera in 360 degrees, the appearance of fogging in the fluid chamber and excessive tear out-in flow through specific meridians, quadrants, or areas of the haptic provide evidence of some scleral asymmetry. The advent of technologies that allowed measurement of the scleral profile led to formal research confirming that only about 6% of scleras are symmetric, while the rest are toric, quad-specific, or irregular. This has an evident impact on how to focus scleral lens fitting. Measuring the scleral profile also provided us with true ocular sagittal height data for cord diameters beyond the cornea. Although high variability was expected in pathological eyes, healthy eyes also showed a wide range of ocular sagittal heights. Due to this variability among healthy eyes, a discussion has emerged on whether the one-size-fits-all approach to soft lens fitting is a proper strategy to fit the whole spectrum of sagittal heights. The traditional mode of selecting the parameters for custom soft lenses through corneal parameters is also questioned.

Corneoscleral junction angle in healthy eyes assessed objectively.

PURPOSE: To introduce a fully objective method to measure corneoscleral junction (CSJ) angle and evaluate the CSJ angle in healthy eyes. METHODS: Corneoscleral topography (Eye Surface Profiler, ESP) was acquired from the right eye of 105 healthy Caucasian subjects, ranging from 18 to 59 years old. From the raw anterior eye height data, the topographic limbus was automatically demarcated in 360 semi-meridians. Further, in limbal location, the CSJ angle was automatically calculated from corneoscleral height data using neighbouring auxiliary points for angle calculation. Additionally, CSJ angle was statistically analysed regionally. RESULTS: The mean CSJ angle was 177.5 ± 1.1°. There is a mean difference of 7.7 ± 3.7° between the steepest (smallest) and flattest (largest) CSJ angle within the same eye. There exist statistically significant differences between temporal (178.2 ± 1.4°) and nasal (176.4 ± 1.1°) regions (paired t-test, p < 0.001), and between superior (178.1 ± 1.1°) and inferior (177.9 ± 1.1°) regions (p = 0.038). CSJ angle is correlated with limbus position (r = 0.43, p < 0.001). CONCLUSION: CSJ angle is rotationally asymmetric. CSJ varies regionally, being the smallest (steepest) in the nasal region. Significant rough changes in CSJ angle were observed for some healthy individuals.

Harvest of Red-Legged Partridge in Central Spain.

A basic rule to attain sustainable use of harvested resources is to adjust take to availability. Populations of red-legged partridges in Spain have decreased in recent decades, and releases of farm-bred partridges to improve short-term availability are increasingly common. We used questionnaires and bird surveys to assess whether harvest was related to availability of wild partridges or intensity of farm-bred partridge releases. We studied 50 hunting estates, including 6 administratively labeled as intensive (with few numerical and temporal restrictions to releases). In addition, we considered hunting pressure (number of hunters × hunting days/km(2)) and habitat as explanatory variables in the analyses. In intensive estates, annual harvest was exclusively related to release intensity, indicating that in these estates hunting is detached from natural resources and approaches an industrial activity based on external inputs. In non-intensive estates, harvest was affected by wild stock availability, walked-up shooting pressure, and habitat (greater harvest in estates with more Mediterranean shrubland). In these estates, releases did not increase annual harvest, and can be considered an inefficient practice. Additionally, the relationship between abundance estimates and harvest disappeared in estates with low partridge abundance estimates, suggesting possibilities for overharvesting in a large proportion of estates. Increasing the abundance of wild red-legged partridge through techniques like habitat management, and improving the adjustment of harvest to availability, may be a good strategy to increase long-term harvest in non-intensive estates. Additionally, Government and managers must create ways to segregate and label the estates where only wild red-legged partridges are managed from those where releases are used, to reduce ecological costs of management decisions. © 2012 The Wildlife Society.

Weasel exposure to the anticoagulant rodenticide bromadiolone in agrarian landscapes of southwestern Europe.

Bromadiolone is an anticoagulant rodenticide (AR) commonly used as a plant protection product (PPP) against rodent pests in agricultural lands. ARs can be transferred trophically to predators/scavengers when they consume intoxicated live or dead rodents. ARs exposure in weasels Mustela nivalis, small mustelids specialized on rodent predation, is poorly known in southern Europe. Moreover, in this species there is no information on bioaccumulation of AR diastereomers e.g., cis- and trans-bromadiolone. Trans-bromadiolone is more persistent in the rodent liver and thus, is expected to have a greater probability of trophic transfer to predators. Here, we report on bromadiolone occurrence, total concentrations and diastereomers proportions (trans- and cis-bromadiolone) in weasels from Castilla y León (north-western Spain) collected in 2010-2017, where bromadiolone was irregularly applied to control outbreaks of common voles Microtus arvalis mainly with cereal grain bait distributed by the regional government. We also tested variables possibly associated with bromadiolone occurrence and concentration, such as individual features (e.g., sex), spatio-temporal variables (e.g., year), and exposure risk (e.g., vole outbreaks). Overall bromadiolone occurrence in weasels was 22% (n = 32, arithmetic mean of concentration of bromadiolone positives = 0.072 mg/kg). An individual showed signs of bromadiolone intoxication (i.e., evidence of macroscopic hemorrhages or hyperaemia and hepatic bromadiolone concentration > 0.1 mg/kg). All the exposed weasels (n = 7) showed only trans-bromadiolone diastereomer in liver, whilst a single analyzed bait from those applied in Castilla y León contained trans- and cis-bromadiolone at 65/35%. Bromadiolone occurrence and concentration in weasels varied yearly. Occurrence was higher in 2012 (100% of weasels), when bromadiolone was widely distributed, compared to 2016-2017 (2016: 20%; 2017: 8.33%) when bromadiolone was exceptionally permitted. The highest concentrations happened in 2014 and 2017, both years with vole outbreaks. Our findings indicate that specialist rodent predators could be exposed to bromadiolone in areas and periods with bromadiolone treatments against vole outbreaks.

Wild micromammal host spectrum of zoonotic eukaryotic parasites in Spain. Occurrence and genetic characterisation.

Micromammals have historically been recognized as highly contentious species in terms of the maintenance and transmission of zoonotic pathogens to humans. Limited information is currently available on the epidemiology and potential public health significance of intestinal eukaryotes in wild micromammals. We examined 490 faecal samples, grouped into 155 pools, obtained from 11 micromammal species captured in 11 Spanish provinces for the presence of DNA from Cryptosporidium spp., Giardia duodenalis, Enterocytozoon bieneusi and Blastocystis sp. The presence of Leishmania spp. was investigated in individual spleen samples. All micromammal species investigated harboured infections by at least one eukaryotic parasite, except Apodemus flavicollis, Myodes glareolus, Sorex coronatus and Sciurus vulgaris, but the sample size for these host species was very low. Cryptosporidium spp. was the most prevalent species found (3.7%, 95% confidence interval [CI]: 2.2-5.7), followed by G. duodenalis (2.8%, 95% CI: 1.6-4.6) and E. bieneusi (2.6%, 95% CI: 1.4-4.3). All pooled faecal samples tested negative for Blastocystis sp. Leishmania infantum was identified in 0.41% (95% CI: 0.05-1.46) of the 490 individual spleen samples analysed. Sequence analyses allowed the identification of Cryptosporidium andersoni (5.9%), C. ditrichi (11.7%), C. muris (5.9%), C. parvum (5.9%), C. tyzzeri (5.9%), rat genotypes CR97 (5.9%) and W19 (5.9%), vole genotypes V (11.7%) and VII (5.9%) and Cryptosproridium spp. (35.3%) within Cryptosporidium (n = 17). Known genotypes C (66.7%) and Peru11 (25.0%) and a novel genotype (named MouseSpEb1, 8.3%) were detected within E. bieneusi (n = 12). None of the G. duodenalis-positive samples could be genotyped at the assemblage level. Molecular data indicate that wild micromammals were primarily infected by rodent-adapted species/genotypes of eukaryotic pathogens and thereby have a limited role as a source of human infections. The presence of ruminant-adapted species C. andersoni along with finding C. parvum is indicative of an overlap between domestic/peri-domestic and sylvatic transmission cycles of these agents.

Investigating the Role of Micromammals in the Ecology of Coxiella burnetii in Spain.

Coxiella burnetii, the causal agent of human Q fever and animal Coxiellosis, is a zoonotic infectious bacterium with a complex ecology that results from its ability to replicate in multiple (in)vertebrate host species. Spain notifies the highest number of Q fever cases to the ECDC annually and wildlife plays a relevant role in C. burnetii ecology in the country. However, the whole picture of C. burnetii hosts is incomplete, so this study seeks to better understand the role of micromammals in C. burnetii ecology in the country. Spleen samples from 816 micromammals of 10 species and 130 vaginal swabs from Microtus arvalis were analysed by qPCR to detect C. burnetii infection and shedding, respectively. The 9.7% of the spleen samples were qPCR positive. The highest infection prevalence (10.8%) was found in Microtus arvalis, in which C. burnetii DNA was also detected in 1 of the 130 vaginal swabs (0.8%) analysed. Positive samples were also found in Apodemus sylvaticus (8.7%), Crocidura russula (7.7%) and Rattus rattus (6.4%). Positive samples were genotyped by coupling PCR with reverse line blotting and a genotype II+ strain was identified for the first time in one of the positive samples from M. arvalis, whereas only partial results could be obtained for the rest of the samples. Acute Q fever was diagnosed in one of the researchers that participated in the study, and it was presumably linked to M. arvalis handling. The results of the study are consistent with previous findings suggesting that micromammals can be infected by C. burnetii. Our findings additionally suggest that micromammals may be potential sources to trace back the origin of human Q fever and animal Coxiellosis cases in Europe.

Lack of detectable genetic isolation in the cyclic rodent Microtus arvalis despite large landscape fragmentation owing to transportation infrastructures.

Although roads are widely seen as dispersal barriers, their genetic consequences for animals that experience large fluctuations in population density are poorly documented. We developed a spatially paired experimental design to assess the genetic impacts of roads on cyclic voles (Microtus arvalis) during a high-density phase in North-Western Spain. We compared genetic patterns from 15 paired plots bisected by three different barrier types, using linear mixed models and computing effect sizes to assess the importance of each type, and the influence of road features like width or the age of the infrastructure. Evidence of effects by roads on genetic diversity and differentiation were lacking. We speculate that the recurrent (each 3-5 generations) episodes of massive dispersal associated with population density peaks can homogenize populations and mitigate the possible genetic impact of landscape fragmentation by roads. This study highlights the importance of developing spatially replicated experimental designs that allow us to consider the large natural spatial variation in genetic parameters. More generally, these results contribute to our understanding of the not well explored effects of habitat fragmentation on dispersal in species showing "boom-bust" dynamics.

Testosterone-mediated trade-offs in the old age: a new approach to the immunocompetence handicap and carotenoid-based sexual signalling.

The immunocompetence handicap hypothesis proposes that testosterone mediates a trade-off between sexual signalling and immunocompetence in males. Such a trade-off could favour the reliability of sexual signals on the basis that testosterone required for signal expression also promotes immunosuppression. However, the immunosuppressive activity of testosterone has not been convincingly demonstrated. We propose that the optimal solution to the testosterone-mediated trade-off should change with age, explaining ambiguous results in the past. Testosterone and ageing would promote two simultaneous immunosuppressive challenges unaffordable for low-quality males. Oxidative stress, as intimately related to ageing and immunosenescence, could contribute to enhance signal reliability. In this context, traits coloured by carotenoids (yellow-red traits) could play a crucial role due to the immunostimulatory and antioxidant properties of these pigments. Here, old and middle-aged male red-legged partridges were treated with testosterone or manipulated as controls. In the presence of high-testosterone levels, middle-aged males increased both circulating carotenoid levels and colour expression, whereas their cell-mediated immunity was not significantly altered. However, in old males, neither circulating carotenoids nor sexual signalling increased when treated with testosterone, but immunosuppression was detected. The link between testosterone and carotenoids could favour the reliability of sexual signals throughout the life.

Repeated sampling but not sampling hour affects plasma carotenoid levels.

Carotenoid pigments have become a central subject of research on animal signaling systems during the past decade. Thus, measurement of plasma carotenoid levels is widespread in the literature. Many plasma biochemical parameters tend to vary with the hour of sampling, which may be an important source of sampling error. However, little is known about this kind of variation for circulating carotenoids. With a sampling protocol that allowed us to separately analyze the effect of sampling hour and repeated blood extraction at the within-individual level, we evaluated the effect of these two parameters on plasma carotenoid concentration in captive red-legged partridges (Alectoris rufa). The hour of sampling did not show a significant influence on carotenoid variability. Nevertheless, carotenoid levels significantly decreased as a result of repeated sampling. In fact, carotenoid variability was positively related to hematocrit values, which also decreased throughout the experiment. Furthermore, the effect was evident from the second sampling event. These results suggest that blood samples for carotenoid analysis may be obtained at random during the daytime with no serious risk of adding variance to data obtained. However, the effect of hemodilution associated with repeated blood extraction should be carefully considered in those studies involving repeated sampling.

Exposure of birds to cholinesterase-inhibiting pesticides following a forest application for tick control.

A mixture of malathion (4%), carbaryl (2%) and cypermethrin (4%) was applied as dry powder to forest and grasslands at a rate of 5kg/ha for tick control in 1500ha of a deer hunting estate in the south of Spain. The effect on plasma cholinesterase (ChE) of house sparrow and nightingale, and brain ChE of red-legged partridge was studied. Plasma ChE was lower after the treatment in passerine birds, but brain AChE was not affected in partridges. The body condition of house sparrows was significantly lower after the treatment.