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The transmission of pathogens to native species has been highlighted as one of the most important impacts of biological invasions. In this study, we evaluated the presence of psittacine beak and feather disease virus (BFDV) and other circoviruses in native bird species cohabiting with invasive populations of wild rose-ringed (Psittacula krameri) and monk parakeets (Myiopsitta monachus) that were found positive for a particular BFDV genotype in Sevilla, southern Spain. None of the 290 individuals from the 18 native bird species captured showed typical signs of disease caused by BFDV. A sample of 79 individuals from 15 native species showed negative results for the presence of the BFDV genotype previously detected in the sympatric invasive parakeets, as well as any other of the circoviruses tested. Although preliminary, this study suggests a lack of circovirus transmission from invasive parakeets to native birds at the study site. Further research is needed to determine if this apparent absence in transmission depends on the BFDV genotype present in the parakeets, which requires additional screening in other invasive and native populations living in sympatry.

A simple methodology was developed to select new sex-specific primers for bird sexing from degraded and low-quantity DNA sources. The strategy was validated using highly degraded DNA extracted from Giemsa-stained blood smears of common cranes (Grus grus). The new primers allowed the accurate molecular sexing using (i) a classic approach of PCR followed by agarose gel electrophoresis and (ii) an advanced real-time PCR method. The simplicity, speed and low cost make this methodology a versatile molecular tool for selection of novel markers/primers for bird sex differentiation from complex DNA sources, which can be used as basis or complement in several fields of ornithological research.

Current bird populations in southern temperate latitudes often represent relicts of glacial refugia from which northern populations expanded as the climate became suitable following the last glacial maximum, 18 000 years before present. Alternatively, these southern populations could be the result of the fragmentation of large distributions and other processes not related to glaciations, like recent recolonization from northern populations and human impact in historical times. Here, we investigate the origin of a small, isolated population of rooks Corvus frugilegus in north-western Iberia. We use genetic data from mitochondrial sequence markers and seven micro-satellite loci to assess levels of genetic diversity, structure and gene flow among extant populations in Iberia and its broad distribution across western Europe. Microsatellite markers revealed the existence of two genetic clusters corresponding to Iberia and the remaining European populations, respectively. Haplotype networks based on mtDNA markers revealed a marked star-like phylogenetic pattern and evidence of a recent population expansion in northern Europe, but not in the Iberian population. Our results suggest that contemporary gene flow between Iberia and western Europe is restricted, and that breeding recruitment over recent generations in the Iberian population is local. The results are consistent with a relatively recent post-glacial colonization of Europe and western Siberia by rooks surviving the last glacial maximum in an Iberian refugium, and likely from refugia in other southern peninsulas. The unique ecological features and genetic diferentiation of the Iberian rooks underscore the importance of ensuring the long-term conservation of this declining population.

Behavioural and socio‑cultural traits are recognized in the restriction of gene flow in species with high cognitive capacity and complex societies. This isolation by social barriers has been generally overlooked in threatened species by assuming disrupted gene flow due to population fragmentation and decline. We examine the genetic structure and ecology of the global population of the Critically Endangered red‑fronted macaw (Ara rubrogenys), an endemic species to the inter‑Andean valleys of Bolivia. We found a fine‑scale genetic structuring in four genetic clusters. Genetic diversity was higher in wild compared to captive‑bred macaws, but similar to that of captive wild‑caught macaws. We found no clear evidence of severe genetic erosion in the population in recent decades, but it was patent in historic times, overlapping with drastic human habitat transformation and macaw persecution over millennia. We found no evidence of geographical and ecological barriers, owing to the high dispersal ability, nesting and foraging habits between genetic clusters. The lack of genetic intermixing despite long‑distance foraging and seasonal movements suggests recruitment in natal colonies and other social factors reinforcing philopatry‑related genetic structure. Conservation efforts should be specifically focussed on major threats in each genetic cluster as independent conservation units, and also considered in ex‑situ management.

The assessment of temporal variation in genetic features can be particularly informativeon the factors behind demography and viability of wildlife populations and species. Weused molecular methods to evaluate neutral genetic variation, relatedness, bottlenecks,and inbreeding in a declining population of Egyptian vulture (Neophron percnopterus)in central Spain. The results show that the genetic diversity remained relatively stableover a period of twelve years despite the decline in census and effective population sizesin the last decades. A relatively high proportion of nestlings from different and distantterritories showed high relatedness in each study year. We also found support for anincreasing impact of severe recent (contemporary) rather than distant (historical) pastdemographic bottlenecks, and the first evidence of inbred mating between full siblingscoinciding with lethal malformations in offspring. The inbred nestling with feathermalformations was positive to beak and feather disease virus recorded for the first timein this species. These results alert on recent and novel threats potentially affecting healthand reducing the adaptive potential of individuals in this threatened species.

Telomere length has been used as a proxy of fitness, aging and lifespan in vertebrates. In the last decade, dozens of articles reporting on telomere dynamics in the fields of ecology and evolution have been published for a wide range of taxa. With this growing interest, it is necessary to ensure the accuracy and reproducibility of telomere length measurement techniques. Real-time quantitative PCR (qPCR) is routinely applied to measure relative telomere length. However, this technique is highly sensitive to several methodological variables and the optimization of qPCR telomere assays remains highly variable between studies. Therefore, standardized guidelines are required to enable the optimization of robust protocols, and to help in judging the validity of the presented results. This review provides an overview of preanalytical and analytical factors that can lead to qPCR inconsistencies and biases, including: (a) sample type, collection and storage; (b) DNA extraction, storage and quality; (c) qPCR primers, laboratory reagents, and assay conditions; and (d) data analysis. We propose a minimum level of information for publication of qPCR telomere assays in evolutionary ecology considering the methodological pitfalls and sources of error. This review highlights the complexity of the optimization and validation of qPCR for telomere measurement per se, demonstrating the importance of transparency and clarity of reporting methodological details required for reliable, reproducible and comparable qPCR telomere assays. We encourage efforts to implement standardized protocols that ensure the rigour and quality of telomere dynamics studies.

The role of wild birds in the carriage and transmission of human and food animal bacteria with resistant genotypes has repeatedly been highlighted. However, few studies have focussed on the specific exposure sources and places of acquisition and selection for antimicrobial-resistant bacteria in vultures relying on livestock carcasses across large areas and different continents. The occurrence of bacterial resistance to antimicrobial agents was assessed in the faecal microbiota of sedentary Griffon vultures (Gyps fulvus) and trans-Saharan migratory Egyptian vultures (Neophron percnopterus) in central Spain. High rates (generally >50%) of resistant Escherichia coli and other enterobacteria to amoxicillin, cotrimoxazole and tetracycline were found. About 25–30% of samples were colonised by extended-spectrum beta-lactamases (ESBL) producing bacteria, while 5–17% were positive for plasmid mediated quinolone resistance (PMQR) phenotypes, depending on vulture species and age. In total, nine ESBL types were recorded (7 in griffon vultures and 5 in Egyptian vultures), with CTX-M-1 the most prevalent in both species. The most prevalent PMQR was mediated by qnrS genes. We found no clear differences in the occurrence of antimicrobial resistance in adult vultures of each species, or between nestling and adult Egyptian vultures. This supports the hypothesis that antimicrobial resistance is acquired in the European breeding areas of both species. Bacterial resistance can directly be driven by the regular ingestion of multiple active antimicrobials found in medicated livestock carcasses from factory farms, which should be not neglected as a contributor to the emergence of novel resistance clones. The One Health framework should consider the potential transboundary carriage and spread of epidemic resistance from high-income European to low-income African countries via migratory birds.

Parthenogenesis is a biological process of asexual reproduction. Recent studies have highlighted the significance of this fascinating phenomenon in the vertebrate evolution. Although parthenogenetic reproduction appears to be widespread among reptiles, a restricted number of cases were reported in captivity and wild. Here, we studied and reported an intriguing case of a 20-year old captive female Cuban boa (Chilabothrus angulifer), from the Zoo da Maia (Maia, Portugal) collection, isolated from conspecifics males, that gave birth twice in 4 years. The neonates from both deliveries, one fresh and the other fixed in formalin, were submitted to histopathological and molecular genetic analysis. Both neonates were homozygous for the loci analyzed, carrying only mother alleles. Furthermore, morphological abnormalities (anophthalmia) were observed in the second neonate. Our data support a pattern of parthenogenetic reproduction. This is the first documented case of facultative parthenogenesis in a Cuban boa, which can be of great interest for further research on ecology, evolution, captive breeding and conservation of the species.

The analysis of telomere dynamics in birds is a growing research field providing important findings on ecological and environmental variations in individuals’ aging, fitness and lifespan. Real-time quantitative PCR (qPCR) has gained much interest for the evaluation of telomere length in birds. However, the assessment of several key preanalytical and analytical factors to optimize the method for achieving reproducible results, and the influence of these factors on the conclusions of each study, have been generally overlooked. In this study, we assessed the performance of eight commercially-available qPCR master mixes in the amplification of telomere fragments in two bird species (zebra finch Taeniopygia guttata and red-billed chough Pyrrhocorax pyrrhocorax). We observed that qPCR master mixes influence the telomere primer binding to target sequences and the amplification specificity within and between bird species, although PCR amplification efficiencies were very close to 100% for all master mixes. These findings indicated that the suitability of the master mix and other analytical factors must be carefully evaluated before starting a telomere dynamics study, especially when the technique has not previously been used in the species. We also showed that optimal PCR amplification efficiencies do not translate to good qPCR telomere assays, and that inference about amplification specificity based only on melting curve data can lead to misleading conclusions. Overall, this work highlights the complexity of qPCR optimization for the study of telomere length in birds.

[Aim]: Oceanic islands have often been colonized by small groups of individuals dispersing from the nearest mainland, giving rise to insular populations characterized by locally adapted phenotypes and low genetic diversity. Alternatively, due to past geo‐climatic changes, the present‐day distribution of the species may not correspond to that found at the time of the original colonization so that the current mainland distribution may not include the original source area, leading to erroneous assumptions regarding colonization history. Here, we use patterns of genetic variation to evaluate alternative colonization scenarios of an insular passerine in the Canary Islands., [Location]: La Palma (Canary Islands), Northern Africa and the Iberian Peninsula., [Taxon]: Red‐billed chough (Pyrrhocorax pyrrhocorax)., [Methods]: We use phylogeographical and coalescent analyses of mitochondrial DNA sequences and 10 microsatellite loci, together with Bayesian demographic modelling, to determine whether choughs on the island of La Palma originate from (a) present‐day populations in Iberia, (b) present‐day populations in the mountains of inland Morocco or (c) former populations in coastal Morocco, where suitable habitat existed in the past., [Results]: Both the mitochondrial and nuclear datasets indicate that the chough population on La Palma is genetically well differentiated from those in Iberia and Morocco, and that La Palma choughs are more closely related to choughs in Iberia than to those in Morocco. Genetic diversity in La Palma is lower than that of mainland populations, but shows no evidence of past bottlenecks. The best supported demographic model to explain the origin of La Palma choughs that is congruent with both genetic datasets includes a ‘ghost’ population closely related to Iberia, from which the insular population diverged within the last 30,000 years., [Main conclusions]: Our results are most consistent with the existence of a former connection between La Palma and Iberia along the North African coast, when suitable habitat was found there. Subsequent desertification of these coastal areas led to local extinctions that restricted gene flow between Iberia and the islands, promoting genetic differentiation. Our results provide a counterintuitive solution to a biogeographical enigma, and could help resolve the colonization history of other systems with similarly complex climatic pasts.

The psittacine beak and feather disease (PBFD) is a globally widespread infectious bird disease that mainly affects species within the Order Psittaciformes (parrots and allies). The disease is caused by an avian circovirus (the beak and feather disease virus, BFDV), which is highly infectious and can lead to severe consequences in wild and captive populations during an outbreak. Both legal and illegal trading have spread the BFDV around the world, although little is known about its prevalence in invasive parrot populations. Here, we analyze the BFDV prevalence in sympatric invasive populations of rose-ringed (Psittacula krameri) and monk parakeets (Myiopsitta monachus) in Southern Spain. We PCR-screened 110 blood samples (55 individuals from each species) for BFDV and characterized the genotypes of five positives from each species. About 33% of rose-ringed parakeets and 37% of monk parakeets sampled were positive for BFDV, while neither species showed disease symptoms. The circovirus identified is a novel BFDV genotype common to both species, similar to the BFDV genotypes detected in several parrot species kept in captivity in Saudi Arabia, South Africa and China. Our data evidences the importance of an accurate evaluation of avian diseases in wild populations, since invasive parrots may be bringing BFDV without showing any visually detectable clinical sign. Further research on the BFDV prevalence and transmission (individual–individual, captive–wild and wild–captive) in different bird orders and countries is crucial to understand the dynamics of the viral infection and minimize its impact in captive and wild populations

Social barriers have been shown to reduce gene flow and contribute to genetic structure among populations in species with high cognitive capacity and complex societies, such as cetaceans, apes and humans. In birds, high dispersal capacity is thought to prevent population divergence unless major geographic or habitat barriers induce isolation patterns by dispersal, colonization or adaptation limitation. We report that Iberian populations of the red-billed chough, a social, gregarious corvid with high dispersal capacity, show a striking degree of genetic structure composed of at least 15 distinct genetic units. Monitoring of marked individuals over 30 years revealed that long-distance movements over hundreds of kilometres are common, yet recruitment into breeding populations is infrequent and highly philopatric. Genetic differentiation is weakly related to geographic distance and habitat types used are overall qualitatively similar among regions and regularly shared by individuals of different populations, so that genetic structure is unlikely to be due solely to isolation by distance or isolation by adaptation. Moreover, most population nuclei showed relatively high levels of genetic diversity, suggesting a limited role for genetic drift in significantly differentiating populations. We propose that social mechanisms may underlie this unprecedented level of genetic structure in birds through a pattern of isolation by social barriers not yet described, which may have driven this remarkable population divergence in the absence of geographic and environmental barriers.

Social barriers have been shown to reduce gene flow and contribute to genetic structure among populations in species with high cognitive capacity and complex societies, such as cetaceans, apes and humans. In birds, high dispersal capacity is thought to prevent population divergence unless major geographical or habitat barriers induce isolation patterns by dispersal, colonization or adaptation limitation. We report that Iberian populations of the red-billed chough, a social, gregarious corvid with high dispersal capacity, show a striking degree of genetic structure composed of at least 15 distinct genetic units. Monitoring of marked individuals over 30 years revealed that long-distance movements over hundreds of kilometres are common, yet recruitment into breeding populations is infrequent and highly philopatric. Genetic differentiation is weakly related to geographical distance, and habitat types used are overall qualitatively similar among regions and regularly shared by individuals of different populations, so that genetic structure is unlikely to be due solely to isolation by distance or isolation by adaptation. Moreover, most population nuclei showed relatively high levels of genetic diversity, suggesting a limited role for genetic drift in significantly differentiating populations. We propose that social mechanisms may underlie this unprecedented level of genetic structure in birds through a pattern of isolation by social barriers not yet described, which may have driven this remarkable population divergence in the absence of geographical and environmental barriers.