Oily secretions produced in the uropygial gland of incubating female hoopoes contain antimicrobial-producing bacteria that prevent feathers from degradation and eggs from pathogenic infection. Using the beak, females collect the uropygial gland secretion and smear it directly on the eggshells and brood patch. Thus, some bacterial strains detected in the secretion should also be present on the eggshell, beak, and brood patch. To characterize these bacterial communities, we used Automatic Ribosomal Intergenic Spacer Analysis (ARISA), which distinguishes between taxonomically different bacterial strains (i.e. different operational taxonomic units [OTUs]) by the size of the sequence amplified.We identified a total of 146 different OTUs with sizes between 139 and 999 bp. Of these OTUs, 124 were detected in the uropygial oil, 106 on the beak surface, 97 on the brood patch, and 98 on the eggshell. The highest richness of OTUs appeared in the uropygial oil samples. Moreover, the detection of some OTUs on the beak, brood patch, and eggshells of particular nests depended on these OTUs being present in the uropygial oil of the female. These results agree with the hypothesis that symbiotic bacteria are transmitted from the uropygial gland to beak, brood patch, and eggshell surfaces, opening the possibility that the bacterial community of the secretion plays a central role in determining the communities of special hoopoe eggshell structures (i.e., crypts) that, soon after hatching, are filled with uropygial oil, thereby protecting embryos from pathogens., Support by funding was provided by Spanish Ministerio de Economía y Competitividad, European funds (FEDER) (CGL2013-48193-C3-1-P, CGL2013-48193-C3-3-P) and Junta de Andalucía (P09-RNM-4557).AM-G had a predoctoral grant from the Junta de Andalucía (P09-RNM-4557)., Peer reviewed

18 files are available from the NCBI GenBank database (accession number(s) KR076707 to KR076724, Molecular methods have revealed that symbiotic systems involving bacteria are mostly based on whole bacterial communities. Bacterial diversity in hoopoe uropygial gland secretion is known to be mainly composed of certain strains of enterococci, but this conclusion is based solely on culture-dependent techniques. This study, by using culture-independent techniques (based on the 16S rDNA and the ribosomal intergenic spacer region) shows that the bacterial community in the uropygial gland secretion is more complex than previously thought and its composition is affected by the living conditions of the bird. Besides the known enterococci, the uropygial gland hosts other facultative anaerobic species and several obligated anaerobic species (mostly clostridia). The bacterial assemblage of this community was largely invariable among study individuals, although differences were detected between captive and wild female hoopoes, with some strains showing significantly higher prevalence in wild birds. These results alter previous views on the hoopoe-bacteria symbiosis and open a new window to further explore this system, delving into the possible sources of symbiotic bacteria (e.g. nest environments, digestive tract, winter quarters) or the possible functions of different bacterial groups in different contexts of parasitism or predation of their hoopoe host., This work was supported by the Ministerio de Ciencia y Tecnología (projects CGL2005-06975/ BOSFEDER; CGL2007-61251/BOSFEDER), the Ministerio de Ciencia e Innovación (projects CGL2009-14006/BOSFEDER; CGL2010-19233-C03- 01/BOSFEDER; CGL2010-19233-C03-03/ BOSFEDER), the Ministerio de Economía y Competitividad (projects CGL2013-48193-C3-1-P/BOSFEDER; CGL2013-4819-C3-2-P/BOSFEDER), and the Junta de Andalucía (RNM 345, P09-RNM- 4557). SMRR received a grant from the Ministerio de Ciencia e Innovación (FPI program, BES-2011- 047677), Peer reviewed

Honesty of offspring begging behaviours is the keystone to understanding the evolution of parent
eoffspring communication. Three main begging costs have been traditionally advocated that ensure the
reliability of offspring signalling: energy expenditure, loss of inclusive fitness and attraction of predators.
Here, we propose that ectoparasites may eavesdrop on begging signals, especially acoustic signals, for
host detection, a never considered but potentially generalized cost of begging that will constrain the
evolution of exaggerated begging displays. Ectoparasitic insects possess a diversity of auditory systems
for intraspecific communication that may be used to detect begging calls of host offspring. The use of
auditory cues for host detection offers some advantages to ectoparasites, particularly in environments in
which long-distance detection of hosts is necessary. There are well-known examples of interspecific
eavesdropping on host auditory signals by parasites that include parasitoid flies attracted to calling
crickets and cicadas, and frog-biting midges and mosquitoes attracted to frog calls. Eavesdropping on
begging signals may have evolved in those parasites searching for hosts that display begging behaviours,
which include not only birds but also mammals and some reptiles and insects with parental care of
juveniles. Considering begging costs due to detection by ectoparasites may help us understand the
reliability, and therefore the evolution, of signals of need and parenteoffspring communication., We were supported by Spanish MINECO/FEDER (CGL2013-48193-C3-1-P), and partially by Ram on y Cajal programme (Spain) and by Secretaría de Educaci on Superior, Ciencia, Tecnología e Innovaci on del Ecuador (SENESCYT) through a Prometeo research grant., Peer reviewed

Selective pressures imposed by pathogenic microorganisms to embryos have selected in hosts for a battery of antimicrobial lines of defenses that includes physical and chemical barriers. Due to the antimicrobial properties of volatile compounds of green plants and of chemicals of feather degrading bacteria, the use of aromatic plants and feathers for nest building has been suggested as one of these barriers. However, experimental evidence suggesting such effects is scarce in the literature. During two consecutive years, we explored experimentally the effects of these nest materials on loads of different groups of bacteria (mesophilic bacteria, Enterobacteriaceae, Staphylococcus and Enterococcus) of eggshells in nests of spotless starlings (Sturnus unicolor) at the beginning and at the end of the incubation period. This was also explored in artificial nests without incubation activity. We also experimentally increased bacterial density of eggs in natural and artificial nests and explored the effects of nest lining treatments on eggshell bacterial load. Support for the hypothetical antimicrobial function of nest materials was mainly detected for the year and location with larger average values of eggshell bacterial density. The beneficial effects of feathers and plants were more easily detected in artificial nests with no incubation activity, suggesting an active role of incubation against bacterial colonization of eggshells. Pigmented and unpigmented feathers reduced eggshell bacterial load in starling nests and artificial nest boxes. Results from artificial nests allowed us to discuss and discard alternative scenarios explaining the detected association, particularly those related to the possible sexual role of feathers and aromatic plants in starling nests. All these results considered together confirm the antimicrobial functionality mainly of feathers but also of plants used as nest materials, and highlight the importance of temporally and geographically environmental variation associated with risk of bacterial proliferation determining the strength of such effects. Because of costs associated to nest building, birds should adjust nest building effort to expected bacterial environments during incubation, a prediction that should be further explored., This work was financed by Spanish Ministerio de Ciencia e Innovación, European funds (FEDER) (CGL2010-19233-C03-01, CGL2010-19233-C03-03, CGL2013-48193-C3-1-P, CGL2013-48193-C3-3-P). MRR and DMG received a postdoc from the programmes “JAE-Doc” and CRC had a predoctoral grant from the Spanish Government. GT was supported by Juan de la Cierva programme (Spain)
and by Secretaría de Educación Superior, Ciencia, Tecnología e Innovación del Ecuador (SENESCYT)
through a Prometeo research grant., Peer reviewed

The study of associations between symbiotic bacterial communities of hosts and those of
surrounding environments would help to understand how bacterial assemblages are
acquired, and how they are transmitted from one to another location (i.e. symbiotic bacteria
acquisition by hosts). Hoopoes (Upupa epops) smear their eggshells with uropygial secretion
(oily secretion produced in their uropygial gland) that harbors antibiotic producing bacteria.
Trying to elucidate a possible role of nest material and cloaca microbiota in
determining the bacterial community of the uropygial gland and the eggshells of hoopoes,
we characterized bacterial communities of nest material, cloaca, uropygial gland and eggshells
by the ARISA fingerprinting. Further, by adding material with scarce bacteria and antimicrobial
properties, we manipulated the bacterial community of nest material and thus
tested experimentally its effects on the microbiomes of the uropygial secretion and of the
eggshells. The experiment did not influence the microbiome of the uropygial secretion of
females, but affected the community established on eggshells. This is the first experimental
evidence indicating that nest material influences the bacterial community of the eggshells
and, therefore, probability of embryo infection. Some of the bacterial strains detected in the
secretion were also in the bacterial communities of the nest material and of cloaca, but their
occurrence within nests was not associated, which suggests that bacterial environments of
nest material and cloaca are not sources of symbiotic bacteria for the gland. These results
do not support a role of nest environments of hoopoes as reservoirs of symbiotic bacteria.
We discuss possible scenarios explaining bacterial acquisition by hoopoes that should be
further explored, Support by funding was provided by Spanish Ministerio de Economía y Competitividad, European funds (FEDER) (CGL2013-48193-C3-1-P, CGL2013-48193-C3-2-P) and Junta de Andalucía (P09-RNM-4557). AM-G had a predoctoral grant from the Junta de Andalucía (P09-RNM-4557)., Peer reviewed

Exploring the links between parental supply and nestling demands and between nestling demand and food supply is of central importance for understanding the evolution of parent-offspring communication. It has been suggested that optimal food supply by parents and begging effort of nestlings are context dependent, and we here test some predictions of this hypothesis. House sparrow (Passer domesticus) nestlings were experimentally fed with a pharmacological appetitive stimulant (cyproheptadine) that increases nestling demands, and explore its effect on nestling growth (i.e. body mass and tarsus length), which can be considered as the net payoff of inflated and costly offspring demand. As assumed by the experimental protocol, nestlings with an exaggerated demand were preferentially fed by parents. In accordance with the hypothesis, net benefits in terms of growth were mainly detected in first breeding attempt of parents that successfully reared three broods. Because costs associated with parental feeding should be lower for first breeding attempts and for parents of higher phenotypic quality (those able to successfully rear three clutches), our results provide to our knowledge, the first experimental evidence supporting a dynamic role of costs of food supply affecting net payoff of offspring demands, which may help to understand the mechanisms allowing the evolutionary equilibrium between intensities of offspring begging and parental provisioning., This work was financed by Spanish Ministerio de Ciencia e Innovación, Ministerio de Economía y Competitividad, European funds (FEDER) (CGL2013-48193-C3-1-P to JJS and CGL2007-61940/BOS To MS). DM-G received a postdoc from the programme “JAE-Doc” of the CSIC, and CR-C had a pre-doctoral grant from the Spanish Government., Peer reviewed

How microbial symbionts are established and maintain on their hosts is a leading question with important consequences for the understanding of the evolution and functioning of mutualistic relationships. The acquisition by hosts of mutualistic microbial symbionts can be considered as colonization processes of environments (i.e., host) by symbionts. Colonization processes can be explored by characterizing the nestedness of communities, but this approach has rarely been applied to communities of microbial symbionts. We used this approach here, and estimated the nestedness of bacterial communities of hoopoes (Upupa epops), a species with symbiotic bacteria in their uropygial gland that are expected to colonize eggshells where they protect embryos from pathogens. Bacterial communities were characterized by ARISA (Automated rRNA Intergenetic Spacer Region) and studied the nestedness characteristics of bacterial communities living in the uropygial secretion, bill, belly and eggshells of each sampled female hoopoes. We detected a consistent nested pattern of bacterial communities of hoopoes; from the uropygial gland to the eggshell. We also found evidence of study year and reproductive events influencing the level of nestedness of bacterial communities of hoopoes. These results indicate that bacterial communities of eggshells and body parts of female hoopoes are at least partially conditioned by the symbiotic community in the uropygial gland. We discuss the importance of these results for understanding this host–microbial mutualism functioning and evolution, Support by funding was provided by Spanish Ministerio de Economía y Competitividad, European funds (FEDER) (CGL2013-48193-C3-1-P, CGL2013-48193-C3-2-P) and Junta de Andalucía (P09-RNM-4557). AMG had a PhD grant from the Junta de Andalucía (P09-RNM-4557)., Peer reviewed

Data available from the Dryad Digital Repository:
< http://dx.doi.org/10.5061/dryad.768vg > (Martín-Gálvez
and Soler 2016), The colouration of some traits in nestlings of altricial birds may influence parental food allocation as it may reflect physical condition or hunger. There is increasing evidence of the relationship between colouration of begging traits and nestling performance. However, evidence of the influence of hunger level on nestling colouration is scarce, mainly because of difficulty of distinguishing between the effects of physical condition and hunger levels. Here, we used the appetite stimulant cyproheptadine hydrochloride to increase the sensation of hunger of magpie Pica pica nestlings for eight days and assessed the effect on the colouration of rictal flanges, mouth and body skin. We found that nestlings administered with cyproheptadine had flanges more conspicuous (chromatic visual contrast), more UV coloured and less yellow coloured than their control nestmates. Conversely, mouths of experimental nestlings were more yellow coloured and less UV coloured
than controls. Our pharmacological experiment afected the strength of the relationship between body mass and some colour components of body skin (chromatic and achromatic visual contrasts, UV – chroma and yellow – chroma) and of rictal flanges (chromatic visual contrasts, UV – chroma and yellow – chroma), but not for mouth colouration. These results taken together suggest that the effect of the cyproheptadine on nestling colourations is probably mediated by an increase in hunger levels of nestlings for rictal flanges and body skin colourations, and by an increase in physical condition in the case of mouth coloration., Th is research was funded by a postdoctoral fellowship to DM-G from Ministerio de Educación y Ciencia, and by Ministerio de Ciencia e Innovación/FEDER (project CGL2013-48193-C3-1-P), Peer reviewed

Microorganisms have shaped the evolution of a variety of defense mechanisms against pathogenic infections. Radioactivity modifies bacterial communities and, therefore, bird hosts breeding in contaminated areas are expected to adapt to the new bacterial environment. We tested this hypothesis in populations of barn swallows (Hirundo rustica) from a gradient of background radiation levels at Chernobyl and uncontaminated controls from Denmark. Investment in defenses against keratinolytic bacteria was measured from feather structure (i.e., susceptibility to degradation) and uropygial secretions. We studied degradability of tail feathers from areas varying in contamination in laboratory experiments using incubation of feathers with a feather-degrading bacterium, Bacillus licheniformis, followed by measurement of the amount of keratin digested. The size of uropygial glands and secretion amounts were quantified, followed by antimicrobial tests against B. licheniformis and quantification of wear of feathers. Feathers of males, but not of females, from highly contaminated areas degraded at a lower rate than those from medium and low contamination areas. However, feathers of both sexes from the Danish populations showed little evidence of degradation. Individual barn swallows from the more contaminated areas of Ukraine produced the largest uropygial secretions with higher antimicrobial activity, although wear of feathers did not differ among males from different populations. In Denmark, swallows produced smaller quantities of uropygial secretion with lower antimicrobial activity, which was similar to swallow populations from uncontaminated areas in Ukraine. Therefore, barn swallows breeding in contaminated areas invested more in all defenses against keratinolytic bacteria than in uncontaminated areas of Ukraine and Denmark, although they had similar levels of feather wear. Strong natural selection exerted by radioactivity may have selected for individuals with higher defense capacity against bacterial infections during the 30 years since the Chernobyl disaster., M. Ruiz-Rodríguez had a Postdoc from the program “Andalucía Talent Hub” (Agencia Andaluza del Conocimiento, Junta de Andalucía). Funding was provided in part from the Samuel Freeman Charitable Trust, the US Fulbright Program, the CNRS (France), the American Council of Learned Societies, the University of South Carolina College of Arts and Sciences, and the Spanish Ministerio de Economía y Competitividad (European funds (FEDER)) (CGL2013-48193-C3-1-P)., Peer reviewed

Intraspecific brood parasitism is widespread among birds and provides clues for elucidating the evolutionary origin of interspecific brood parasitism. Studies suggest that brood parasitism does not occur at random, but that parasitic females select nests with advantages such as higher physical stability, reduced predation risk, or lower ectoparasite infestations. However, this evidence is sparse and mainly correlative. By experimentally increasing the abundance of Carnus hemapterus (a common, generalist and widespread ectoparasitic nest fly of a multitude of bird species) in half of the nests, we show that parasitic Spotless Starlings (Sturnus unicolor) avoid conspecific nests infested by ectoparasites. Since Carnus ectoparasites impinge costs on their avian nestling hosts, this avoidance response would be adaptive for parasitic Starlings. Further, we suggest a mechanism by which parasitic females may assess the level of ectoparasite infestation to select host nests accordingly: by using parasite cues such as faeces and blood remains. Additionally, these cues may be used by parasitic females for synchronization with the reproductive cycle of host females because ectoparasite cues also indicate that incubation has already commenced. Whatever the functionality, the mechanism suggested here may be employed by intra- and interspecific brood parasites, so it might represent a widespread strategy in nature., Financial support was provided by the Spanish
Ministerio de Economía y Competitividad and FEDER (CGL2013-
48193-C3-1-P), the JAE programme to D. M. G. and M. R. R., and
the Ramo´n y Cajal programme (Spain) and the Secretarı´a de Educacio
´n Superior, Ciencia, Tecnologı´a e Innovacio´n del Ecuador
(SENESCYT) through a Prometeo research grant to G. T, Peer reviewed

Nest sanitation-related traits have often been explained at the intraspecific level as reducing the probability
of infection or detection by predators and parasites, but its evolution within the avian phylogeny is
still poorly understood. We compiled detailed information of such traits for more than 400 bird species
and, by means of modern comparative methodologies, we reconstructed the evolution of adults'
contribution to removing their offspring's faeces and the production of faecal sacs by nestlings.
Furthermore, because the functional hypotheses used to explain nest sanitation behaviour assume potential
effects of brood size, body mass, nestling period and diet, we explored the association between
these traits and those related to nest sanitation in a phylogenetically controlled framework. Our results
suggest that parental removal of nestling faeces has driven the evolution of faecal sacs, while the
ancestral states involved birds with faecal sacs removed by parents. These results support the long-held
idea that faecal sacs facilitate the removal of faeces by parents. Moreover, we found that animal diets and
small body sizes have favoured the evolution of faecal sacs suggesting the existence of some chemical
and physical constraints in relation to the evolution of the mucous covering. Our results highlight the
importance of nest sanitation in the evolution of birds and their life history characteristics, J.D.I. was
financed by a postdoctoral contract (TAHUB-104) from the program
‘Andalucía Talent Hub’ (cofunded by the European Union's Seventh
Framework Program Marie Skłodowska-Curie actions (COFUND)
and the regional Government of Andalucía). Funding was partially
provided by the Spanish Ministerio de Economía y Competitividad
(European funds (FEDER)) (CGL2013-48193-C3-1-P)., Peer reviewed

Nest materials used by animals can have profound effects on developing offspring. They can modify the bacterial and parasitic environment of the nest, and can influence parental investment through sexual signalling processes. In spotless starlings, Sturnus unicolor, green plants and feathers are known nest materials with such functions. The aim of our study was to experimentally assess their influence on nestlings' telomere length and attrition, which are good predictors of their survival prospects. In a full-factorial experiment, we explored these effects in two different populations, together with the potential effects of hatching date, ectoparasitism, bacterial environment and nestling growth. Telomere length and attrition largely depended on population identity and hatching date. After correcting for these effects, the addition of feathers resulted in higher rates of telomere attrition. The addition of plants did not affect nestling telomeres in general, but did in interaction with location: in Hueneja, the experimental addition of green plants resulted in longer telomeres. Feather pigmentation also did not affect telomere length or attrition in general, but did in interaction with location: in Hueneja, the experimental addition of unpigmented feathers resulted in nestlings with longer telomeres and lower attrition rates. Moreover, prevalence of staphylococci on the skin of 8-day-old nestlings was negatively related to telomere lengths of fledglings. Taken together, these results suggest a direct link between nest material composition and nestling telomere length and dynamics. This relationship could be partially mediated by the antimicrobial and/or antiparasitic properties of nest materials or by sexual signalling processes. We discuss possible roles of maternal effects, parasites, immunity and nestling growth in explaining these experimental effects., Funding was provided by the Spanish Ministerio de Economia y Competitividad and European funds (FEDER) (CGL2013-48193-C3-1-P, CGL2013-48193-C3-2-P, CGL2012-30759, CGL2015-65055-P). J.M.P., G.T. and M.R.R. were supported respectively by Juan de la Cierva, Ramón & Cajal and Talentia postdoctoral contracts., Peer reviewed

The avian nest is an essential structure for offspring development. For adults, nest building entails costs in terms of time, energy, and exposure to predators and parasites. Amount and diversity of materials used for nest building depend on their availability and functionality in scenarios of sexual selection and parasitism. Green plants and feathers of different colors have been hypothesized to play key roles in offspring protection against pathogens, and we here experimentally assessed spotless starling (Sturnus unicolor) preferences for pigmented versus unpigmented feathers and for different green plants (aromatic vs. non-aromatic plants) as nest materials. We predicted a preferential selection of unpigmented feathers and aromatic plants according to the antimicrobial properties of these materials described in the literature. We evaluated these predictions during nest building and during egg-laying stages. As expected, starlings preferentially selected unpigmented feathers both before and during egg laying, while aromatic plants were preferentially selected only during the egg-laying stage. These results suggest that starlings prefer nest materials that enhance antimicrobial protection of their offspring. We also discuss some other, non-exclusive functions that might explain the observed preference for nest materials, especially with regard to their potential role in sexual signaling, This work was supported by Spanish Ministerio de Ciencia e Innovación, European funds (FEDER) (CGL2010-19233-C03-01, CGL2010-19233-C03-03, CGL2013-48193-C3-1-P, CGL2013-48193-C3-2-P). M.R.R. received a postdoc from the program “JAE-Doc” and C.R.C. had a predoctoral grant from the Spanish Government. G.T. was supported by Ramón y Cajal program (Spain)., Peer reviewed

Nestlings of most avian species produce faecal sacs, which facilitate the removal of nestlings’ excrements by parents, thereby reducing proliferation of potentially pathogenic microorganisms and/or detectability by predators and parasites. The nest microbial environment that birds experience during early life might also affect their development and thus, faecal sacs facilitating parental removal may be a strategy to decrease bacterial contamination of nests that could harm developing nestlings. Here, we tested this hypothesis by experimentally broken faecal sacs and spreading them in nests of spotless starlings (Sturnus unicolor), thereby avoiding their removal by adults. In accordance with the hypothesis, experimental nests harboured higher bacterial density than control nests. Nestlings in experimental nests were of smaller size (tarsus length) and experienced lower probability of survival (predation) than those in control nests. Moreover, nestlings in experimental nests tended to suffer more from ectoparasites than those in control nests. We discuss the possible pivotal role of bacteria producing chemical volatiles that ectoparasites and predators might use to find avian nests, and that could explain our experimental results in starlings, Ministerio de Ciencia, Innovación y Universidades and European (FEDER) funds (CGL2013-48193-C3-1-P, CGL2017-83103-P), Peer reviewed

Factors affecting the evolution of plumage conspicuousness in females are nowadays the focus of debate, and here we explore the possibility that the conspicuousness of female plumage and male participation in nest building are associated in birds. We hypothesize that males that participate in nest building will gain higher fecundity from high-quality ornamented females, whereas ornamented females will adjust fecundity to the costly nest-building behaviour of males. Large-sized species might experience higher costs of nest building and, thus, body size should affect the scenario described above. We used information on male contribution to nest construction (yes/no), male and female conspicuousness (conspicuous or cryptic plumage) and body size of Western Palaearctic passerines. In accordance with the hypothesis, we found that female conspicuousness, in interaction with body mass, was strongly associated with male participation in nest building. For large-sized species, female conspicuousness was positively associated with male participation in nest building. Discrete analyses of correlated evolution rendered evidence of female conspicuousness determining the evolution of male contributions to nest building, with the loss of female conspicuousness occurring more frequently before the loss of male participation in nest building. We discuss possible adaptive scenarios explaining the detected evidence because of mutual sexual selection in males and females, The study was financed by the Ministerio de Economía y Competitividad MINECO
(CGL2013-48193-C3-1-P, CGL2013-48193-C3-2-P, CGL2013-48193-C3-3-P and CGL2017-83103-P) and the European Regional Development Fund (FEDER), Peer reviewed

Sexually selected signals reliably reflect individual phenotypic or genetic quality and, thus, survival prospects of holders. Telomere length is considered a good predictor of life expectancy and, consequently, exploring the links between telomere length and sexually selected traits is much needed to better understand the mechanisms that maintain the honesty of sexual signals. We manipulated the length of throat feathers in spotless starling (Sturnus unicolor) males (a sexually selected signal) before reproduction and explored its effects on telomere shortening and breeding performance in subsequent reproductive events. We did not detect an effect of the feathers clipping manipulation, but males with longer throat feathers before the experiment had shorter telomeres that also shortened more slowly than those of males with shorter throat feathers did. Moreover, length of throat feathers of males before manipulation was positively related to hatching success of second clutches. Thus, correlative but not experimental results support the expected associations between sexually selected signals, telomeres and reproduction. We discuss such results in scenarios of sexual selection where feather length reflects, but does not directly cause, telomere attrition and enhanced reproductive success. Males with longer throat feathers might be older, more experienced males (i.e., with shorter telomeres), able to buffer telomere shortening between reproductive events. Because of the absence of experimental effects, differential incubation effort of females cannot explain the detected association with hatching success, but other sexually selected traits that covary with throat-feather length could be responsible. Exploring those physiological and/or morphological characteristics related to throat-feather length should therefore be the matter of future research, This work was supported by the currently named Ministerio de Ciencia, Innovación y Universidades and European (FEDER) funds (CGL2013-48193-C3-1-P, CGL2017-83103-P, CGL2017-89063-P, CGL2015-65055-P and PGC2018-095704-B-100). MAG was financed by a predoctoral contract (BES-2014-068661) from the Spanish Ministerio de Economía y Competitividad, and GT by the Ramón y Cajal Programme., Reproductive data; Telomere data, Peer reviewed

Data file includes (i) average values of melanised (M) and non-melanised (NM) feather wear of nestling birds that grew in different nests. The file also included (ii) information of feather mass degradability of nestlings and adult birds from different nests and species estimated as the olipeptide concentration (μg x mL1) of tubes containing 4mL of PBS buffer and a colony Bacillus licheniformis D13 after 21 days of incubation at 37 ºC in constant agitation at 120 rpm in an orbital agitator (VWR, Spain). The third excel sheet contain information of bacterial loads in terms of terms of mesophilic bacteria (TSA), Enterobacteriaceae (HK), Staphylococcus (VJ), and Enterococcus (KF), and average wear of melanised (M) and non-melanised (NM) wing feathers of nestlings that grew in the same nests where bacterial loads were estimated. The last excel file contain information on feather wear of melanised and non-melanised feathers of each sampled nestlings and adult. All these data were collected the Guadix region (southern Spain) and used to perform the statistical analyses in Tables 2 and 3 in Azcárate et al. 2020 (Doi: 10.1111/jav.02320)., This work was supported by the currently named Ministerio de Ciencia, Innovación y Universidades and European (FEDER) funds (CGL2013-48193-C3-1-P, CGL2013-48193-C3-2-P, CGL2013-48193-C3-3-P, CGL2017-83103-P). MAG and SDL were financed by predoctoral contracts (BES-2014-068661, BES-2014-069116) from the Spanish Ministerio de Economía y Competitividad., Peer reviewed

Feathers are essential for avian life, and factors affecting their integrity are important to understand their evolution. These factors should depend on, among other traits, species‐specific bacterial environments and life‐history characteristics. However, interspecific variation in feather deterioration, feather susceptibility to degradation by keratinolytic bacteria (degradability), and bacterial environment, have rarely been quantified. Here, we did so by measuring deterioration and degradability of wing feathers of fledglings in 16 bird species, and characterizing the bacterial environment where they developed. We found statistically significant interspecific variation for all considered variables. On average, non‐melanised were more deteriorated than melanised feathers, but differences depended on the species. Moreover, nest bacterial loads were related to feathers wear, but the sign of the association depended on the bacterial group considered and on feather pigmentation. We also found a positive association of feather degradability with wear of non‐melanised feathers, and with bacterial loads. These results suggest that bacterial environments determine the integrity of fledgling feathers as well as their resistance to bacterial degradation, which implies a preponderant role of bacteria in driving the evolution of avian feathers., This work was supported by the currently named Ministerio de Ciencia, Innovación y Universidades and European (FEDER) funds (CGL2013-48193-C3-1-P, CGL2013-48193-C3-2-P, CGL2013-48193-C3-3-P, CGL2017-83103-P). MAG and SDL were financed by predoctoral contracts (BES-2014-068661, BES-2014-069116) from the Spanish Ministerio de Economía y Competitividad., Peer reviewed

Nest re-use in birds has the potential cost of infection by parasites and pathogens but may also be a source of beneficial symbiotic bacteria transmitted horizontally. Eurasian hoopoes Upupa epops host antibiotic-producing bacteria in their uropygial gland but only while breeding, which suggests that the nest-hole may be a source of those symbionts. Interestingly, hoopoes do not build nests, thus might prefer for reproduction nest holes with soft materials from previous reproductions. Here, we tested experimentally this preference by installing in the field new nest boxes that were left empty or filled with either sawdust or a mixture of sawdust and hoopoe's nest material from the previous year. We explored the experimental effect on the composition of the uropygial secretion bacterial community, on eggshell bacterial loads, and on several proxies of reproductive success. Hoopoes bred significantly more often in nest boxes with nest material than in empty ones, but the type of nest material did not affect nest box occupancy. Eggs in nest boxes with old-soft material harbored higher bacterial density on their shells, and the microbiota of the uropygial secretion of nestlings and females in these nest boxes differed from those in nest boxes without old-soft material. Moreover, although the experiment did not affect breeding success or related proxies, several operational taxonomic units from female uropygial secretions were positively associated with hatching success. This is the first experimental evidence showing that re-used nest material affects the bacterial community of the uropygial secretions of hoopoe females. This suggests that the nest material can be a source of strains for their incorporation to both the uropygial gland and eggshell communities, highlighting a possible advantage of nest re-use previously unconsidered., Silvia Díaz Lora was financed by a predoctoral contract (BES‐2014‐069116) and research by three projects from the Spanish Ministerio de Economía y Competitividad, Ministerio de Ciencia, Innovacion y Universidades, and European (FEDER) funds (CGL2013‐48193‐C3‐1‐P, CGL2013‐48193‐C3‐2‐P and CGL2017‐83103‐P).

Avian feathers need to be replaced periodically to fulfill their functions, with natural, social, and sexual selection presumably driving the evolution of molting strategies. In temperate birds, a common pattern is to molt feathers immediately after the breeding season, the pre- basic molt. However, some species undergo another molt in winter- spring, the pre-alternate molt. Using a sample of 188 European passerine species, Bayesian phylogenetic mixed models, and correlated evolution analyses, we tested whether the occurrence of the pre-alternate molt was positively associated with prox-ies for sexual selection (sexual selection hypothesis) and nonsexual social selection (social selection hypothesis), and with factors related to feather wear (feather wear hypothesis) and time constraints on the pre- basic molt (time constraints hypothesis). We found that the pre-alternate molt was more frequent in migratory and less gre-garious species inhabiting open/xeric habitats and feeding on the wing, and margin-ally more frequent in species with strong sexual selection and those showing a winter territorial behavior. Moreover, an increase in migratory behavior and sexual selection intensity preceded the acquisition of the pre-alternate molt. These results provide support for the feather wear hypothesis, partial support for the sexual selection and time constraints hypotheses, and no support for the social selection hypothesis.KEYWORDSbirds, feather wear, pre-alternate molt, sexual selection, social selection, time constraintsTAXONOMY CLASSIFICATIONBehavioural ecology; Evolutionary ecology; Life history ecology, European Regional Development Fund; Spanish Ministry of Economy and Competitiveness, Grant/Award Number: CGL2013-48193-C3-1-P, CGL2013-48193-C3-3-P, CGL2017-83103-P, CGL2017-83843-C2-1-P and RYC-2014-15145; Spanish Ministry of Science and Innovation, Grant/Award Number: PID2019-106032GB-I00, Peer reviewed

Avian brood parasites are expected to select host nests according to characteristics that maximize offspring fitness, such as reduced probability of ectoparasitism. Spotless starlings, Sturnus unicolor, lay immaculate blue eggs that sometimes become brownish-spotted due to the activity of the ectoparasitic fly Carnus hemapterus on incubating birds. Therefore, conspecific parasitic females should adaptively avoid parasitizing nests with spotted eggshells. Here, we manipulated perceived risk of ectoparasitism by painting the eggs with either brown spots (similar to those due to ectoparasite activity) or blue spots. A third group of nests was maintained with immaculate eggshells. Nests with nonspotted eggshells showed the highest rate of brood parasitism, while nests with brownish or bluish spots on the eggshells were parasitized at a similar lower rate. These results suggest that brood-parasitic females use the presence of spots on the eggshells in their selection of host nests. This study adds to the scarce evidence showing that brood-parasitic birds select host nests with a low risk of ectoparasitism and demonstrates that colour patterns of the eggshell (i.e. spottiness) of their potential hosts represent a major cue employed to appraise the risk of ectoparasitism., This work was supported by the currently named Ministerio de Ciencia, Innovación y Universidades and European (FEDER) funds (CGL 2013-48193-C3-1-P, CGL 2017-83103-P, CGL 2017-89063-P). M.A.G. was financed by a predoctoral contract (BES-2014-068661) from the Spanish Ministerio de Economía y Competitividad, and G.T.y the Ramón y Cajal Programme., Peer reviewed