Diet and host genetic or evolutionary history are considered the two main factors determining gut microbiota of animals, although studies are scarce in natural populations. The system of great spotted cuckoos (Clamator glandarius) parasitizing magpies (Pica pica) is ideal to study both effects since magpie adults feed cuckoo and magpie nestlings with the same diet and, consequently, differences in gut microbiota of nestlings of these two species will mainly reflect the importance of genetic components. Moreover, the diet of adults and of nestling cuckoos drastically differ from each other and, thus, differences and similarities in their microbiotas would respectively reflect the effect of environmental and genetic factors. We used next-generation sequencing technologies to analyze the gut microbiota of cuckoo adults and nestlings and of magpie nestlings. The highest α-diversity estimates appeared in nestling cuckoos and the lowest in nestling magpies. Moreover, despite the greatest differences in the microbiome composition of magpies and cuckoos of both ages, cuckoo nestlings harbored a mixture of the Operational Taxonomic Units (OTUs) present in adult cuckoos and nestling magpies. We identified the bacterial taxa responsible for such results. These results suggest important phylogenetic components determining gut microbiome of nestlings, and that diet might be responsible for similarities between gut microbiome of cuckoo and magpie nestlings that allow cuckoos to digest food provided by magpie adults., This work was supported by the Ministerio de Ciencia, Innovacion y Universidades and European (FEDER) funds (CGL2017-83103-P)., Peer reviewed

The uropygial gland of hoopoe nestlings and nesting females hosts bacterial symbionts that cause changes in the characteristics of its secretion, including an increase of its antimicrobial activity. These changes occur only in nesting individuals during the breeding season, possibly associated with the high infection risk experienced during the stay in the hole-nests. However, the knowledge on hoopoes uropygial gland microbial community dynamics is quite limited and based so far on culture-dependent and molecular fingerprinting studies. In this work, we sampled wild and captive hoopoes of different sex, age, and reproductive status, and studied their microbiota using quantitative polymerase chain reaction (qPCR), fluorescence in situ hybridization (FISH) and pyrosequencing. Surprisingly, we found a complex bacterial community in all individuals (including non-nesting ones) during the breeding season. Nevertheless, dark secretions from nesting hoopoes harbored significantly higher bacterial density than white secretions from breeding males and both sexes in winter. We hypothesize that bacterial proliferation may be host-regulated in phases of high infection risk (i.e., nesting). We also highlight the importance of specific antimicrobial-producing bacteria present only in dark secretions that may be key in this defensive symbiosis. Finally, we discuss the possible role of environmental conditions in shaping the uropygial microbiota, based on differences found between wild and captive hoopoes., This work was supported by the Spanish Ministry of Economy and Competitiveness and European
(FEDER) funds (CGL2013-48193-C3-1-P/BOS, CGL2013-48193-C3-2-P/BOS, CGL2017-83103-P), and the Junta de Andalucía (RNM 339, RNM 340). S. M. Rodríguez-Ruano received a grant from the Spanish Ministry of Science and Innovation (FPI program)., Peer reviewed

Diet and host genetic or evolutionary history are considered the two main factors determining gut microbiota of animals, although studies are scarce in natural populations. The system of great spotted cuckoos (Clamator glandarius) parasitizing magpies (Pica pica) is ideal to study both effects since magpie adults feed cuckoo and magpie nestlings with the same diet and, consequently, differences in gut microbiota of nestlings of these two species will mainly reflect the importance of genetic components. Moreover, the diet of adults and of nestling cuckoos drastically differ from each other and, thus, differences and similarities in their microbiotas would respectively reflect the effect of environmental and genetic factors. We used next-generation sequencing technologies to analyze the gut microbiota of cuckoo adults and nestlings and of magpie nestlings. The highest α-diversity estimates appeared in nestling cuckoos and the lowest in nestling magpies. Moreover, despite the greatest differences in the microbiome composition of magpies and cuckoos of both ages, cuckoo nestlings harbored a mixture of the Operational Taxonomic Units (OTUs) present in adult cuckoos and nestling magpies. We identified the bacterial taxa responsible for such results. These results suggest important phylogenetic components determining gut microbiome of nestlings, and that diet might be responsible for similarities between gut microbiome of cuckoo and magpie nestlings that allow cuckoos to digest food provided by magpie adults., This work was supported by the Ministerio de Ciencia, Innovacion y Universidades and European (FEDER) funds (CGL2017-83103-P)., 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

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., 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).
MAG was financed by a predoctoral contract (BES-2014-068661)
from the Spanish Ministerio de Economía y Competitividad
(CGL2013-48193-C3-1-P)., Peer reviewed

The offspring of many animals are conspicuous during parental dependence, despite juveniles generally suffering from high predation risk. However, to date, it is unclear whether offspring structural ornaments play a role in intrafamily communication. This is the case of conspicuous plumage in young birds, which is worn unchanged during a long period after fledging, when they still depend on their parents. If plumage color facilitates intrafamily interactions, its role should be more important in large-brooded species, where the strength of intrafamily conflict is potentially stronger. We therefore performed a comparative study in 210 passerine bird species to test whether an offspring structural trait, white plumage, evolves more frequently in lineages with larger clutches. We also explored the number of broods raised per year as another source of intrafamily conflict. First, we found that juvenile whiteness was more frequent in open-nesting species. Moreover, in agreement with our prediction, the presence of juvenile white tail/wing patches was strongly and positively associated with clutch size. This relationship was not due to the strong resemblance between offspring and adult plumage, which was controlled for in the statistical analyses. Moreover, the association remained significant after taking into account predation risk, for which there was information for a subset of species. In contrast, juvenile whiteness was not associated with the number of broods raised per year. These results may suggest that the evolution of juvenile conspicuousness is favored in species with potentially stronger intrabrood sibling conflict., This work was financed by the Ministerio de Economía, Industria y
Competitividad MINECO (CGL2016-79390-P, CGL2017‐83103‐P,
CGL2017-83843-C2-1-P and CGL2013-48193-C3-1-P) and the European
Regional Development Fund (FEDER)., 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).

Within the context of complex sexual signalling, most research has focussed on exploring the associations between several signals and/or their relationships with different proxies of individual quality. However, very few studies have focused on checking whether the expression of one signal is conditioned by the expression of the others. Here, by experimentally shortening the throat feathers of male spotless starlings (Sturnus unicolor), we evaluated the influence of this trait on the colour expression of the bea19 base. In addition, we tested the relationship between these two sexually dimorphic characters with traits indicating individual quality such as body condition and colour reflectance at the wavelength related to carotenes in the tip of the beak. Our results show that the colouration of the beak base in males, but not in females, is positively related to body condition and to the length of ornamental throat feathers. Moreover, the experimental shortening of throat feathers in males had a negative effect on the blue chroma intensity of their beak base one year after manipulation. These results support for the first time a causal link between the expression of two sexually dimorphic characters, which is essential to understand their functionality in a multiple signalling framework, 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). 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., Peer reviewed

Determining costs associated with nest‐building behaviour and nest characteristics is of prime importance to understand the evolution of such extended phenotypic traits. We explored a cost of nest building experimentally to determine whether large nests attracted more ectoparasites. This cost may be especially important to cavity‐nesting species where larger nests likely reduce the distance between nest contents and the cavity entrance, making the host nest more vulnerable to ectoparasite detection. We manipulated the amount of nest material and the distance between nest and cavity entrance and estimated the abundance of ectoparasites (Carnus hemapterus flies) before and after manipulation in nests of Spotless Starlings (Sturnus unicolor) breeding in nestboxes. Nest size was enlarged and elevated in a first group of nests by adding nest material to the base of the nest (enlarged nests); a second group of nests was elevated with a piece of plastic but without adding nest material (elevated nests); a third group of nests was not manipulated (control nests). Number of flies per nest increased significantly in enlarged and elevated nests compared to control nests. Moreover, distance to cavity entrance after manipulation was negatively associated with the increase in the number of flies per nest (i.e., nests closer to the entrance had more parasites). Thus, nest size, through its effect on distance between nest and cavity entrance, affected fly abundance. Cues produced by nest contents located closer to the cavity entrance may be detected more readily by ectoparasitic flies searching for hosts. These results suggest a novel constraint for the evolution of nest size in cavity‐nesting birds., Financial support was provided by grants from Spanish Ministerio de Ciencia e Innovación (MCI) and FEDER (CGL2010-19233-C03-01), from Ministerio de Economía, Industria y Competitividad (CGL2017-89063-P, CGL2017-83103-P), by a predoctoral fellowship from MCI to CRC, by Junta para Ampliación de Estudios programme to MRR, and by Juan de la Cierva and Ramón y Cajal programmes to GT. Authorization to conduct the study was granted by the Dirección General de Gestión del Medio Natural, Consejería de Medio Ambiente, Junta de Andalucía., Peer reviewed

Dataset used in Tomás et al. (2020): (DOI: 10.1111/jav.02427). Data were collected in the Hoya de Guadix in 2012 in spotless starling (Sturnus unicolor) nests in the framework of an experiment to evaluate whether distance to the cavity entrance and nest size influence the risk of ectoparasitism in avian nests. Dataset includes hatching date, brood size, mean nestling body mass, microclimate of the cavity, as well as ectoparasite abundance and distance between the nest and the cavity entrance before and after experimental manipulation., Dataset used in Tomás et al. (2020): (DOI: 10.1111/jav.02427). It contains data collected in Guadix in 2012 in the framework of an experiment to evaluate whether distance to the cavity entrance and nest size influence the risk of ectoparasitism in avian nests., Financial support was provided by grants from Spanish Ministerio de Ciencia e Innovación (MCI) and FEDER (CGL2010-19233-C03-01), from Ministerio de Economía, Industria y Competitividad (CGL2017-89063-P, CGL2017-83103-P), by a predoctoral fellowship from MCI to CRC, by Junta para Ampliación de Estudios programme to MRR, and by Juan de la Cierva and Ramón y Cajal programmes to GT., Peer reviewed

It contains data collected in Guadix, Granada (southeast Spain) in 2019, in the framework of a research to evaluate the effects of chemical and auditory cues of hoopoes (Upupa epops) in repellence and attraction of blood-feeding flies. Dataset includes number of mosquitoes, blackflies and biting midges collected in insect traps in the field in four habitat types and two sampling periods, in relation to different chemical and auditory stimuli derived from nestlings of this bird species., Financial support was provided by grants from the Spanish Ministerio de Economía, Industria y Competitividad (CGL2017-89063-P, CGL2017-83103-P). GT was also supported by the Ramón y Cajal Programme. We acknowledge support of the publication fee by the CSIC Open Access Publication Support Initiative through its Unit of Information Resources for Research (URICI)., Peer reviewed

Data sampled in “Hoya de Guadix”, Granada, Spain, during breeding seasons (March-July) 2016 and 2019., Data file includes information about identity of each sampled starling nestling, the experiment and treatment assigned to them, biometric measurements on day 10 and day 14, and measurements of colour variables. Colour variables include colouration of eggshells (measured as Brightness and Blue-green chroma for eggs laid on the nest where each nestling was born (nest of origin), and the eggs laid on the nest where each nestling grew (nest of rearing)), and colouration of begging related traits (measured as Mean bright, Total hue, UV hue, Yellow-red hue, UV croma, Yellow-red chroma and Carotenoid chroma) for each body part (mouth, flanges and skin). The data set also includes the plasma carotenoid concentration of each sampled nestling., Ester Martínez Renau was financed by a predoctoral contract (PRE2018-085378), while the whole research group received funds from the Spanish Ministerio de Ciencia, Innovacion y Universidades, and European (FEDER) (CGL2017-83103-P)., Peer reviewed

Research on the mechanisms involved in host location by parasites is of paramount importance and may aid in developing protective measures against them. This topic attains far-reaching repercussions for human and animal welfare regarding parasites transmitting vector-borne pathogens, such as blood-feeding flies. Very few studies have evaluated the effect of bird-derived cues on attraction of vectors in field conditions. We here explored the attraction of different groups of blood-feeding flies (mosquitoes, blackflies and biting midges) to auditory cues produced by begging hoopoe (Upupa epops) nestlings, and to three chemical cues derived from hoopoe nestlings or nests (uropygial secretion, symbiotic bacteria isolated from the secretion, and nest material) in the field. We deployed insect traps baited with the different stimuli at the beginning and at the end of the hoopoe breeding-season in four different habitats. Abundance of blood-feeding flies varied depending on habitat and sampling period. Begging auditory cues of nestling hoopoes did not affect abundance of flies. However, chemical stimuli affected abundance of mosquitoes, which were less abundant in traps baited with bacteria or with nest material than in control traps. Abundance of biting midges in traps also depended on the chemical stimulus but in interaction with sampling period or habitat. Fewer biting midges were collected in traps baited with bacteria and with secretion in the habitats where abundance of biting midges is higher. Our results suggest that uropygial secretion of hoopoes, and symbiotic bacteria living in this secretion, may repel blood-feeding flies from their nests., Financial support was provided by grants from the Spanish Ministerio de Economía, Industria y Competitividad (CGL2017-89063-P and CGL2017-83103-P). GT was also supported by the Ramón y Cajal Program. We acknowledge support of the publication fee by the CSIC Open Access Publication Support Initiative through its Unit of Information Resources for Research (URICI)., Peer reviewed

This article belongs to the Special Issue Cues Followed by Parasites and Predators in Detecting Their Victims., [Simple Summary] Microorganisms may have direct negative effects on their animal hosts and cause diseases, but some others provide animals with protection against infections, parasites, and even predators. However, parasites or predators might take their cue from odors produced by bacteria, even those from protecting microorganisms, which could turn net benefits of bacteria that are a priori considered beneficial into a neutral or even negative outcome. This possibility has scarcely been studied in wildlife populations; we manipulated the bacterial community of nest-material of hoopoes and detected a negative effect in terms of the intensity of parasitism by blood-sucking flies that nestlings suffer. We also detected a positive link between the bacterial density of the nest-materials and the intensity of ectoparasitism, which further points at the importance of the bacteria determining the level of parasitism. Blood-sucking ectoparasites are also disease vectors, affecting both humans and livestock, and considering the role of the bacterial environment might help to establish new transmission control protocols., Nest bacterial environment influences avian reproduction directly because it might include pathogenic- or antibiotic-producing bacteria or indirectly because predators or ectoparasites can use volatile compounds from nest bacterial metabolism to detect nests of their avian hosts. Hoopoes (Upupa epops) do not build nests. They rather reuse holes or nest-boxes that contain remains of nest-materials from previous breeding seasons. Interestingly, it has been recently described that the nest’s bacterial environment partly affects the uropygial gland microbiota of hoopoe females and eggshells. Blood-sucking ectoparasites use chemical cues to find host nests, so we experimentally tested the hypothetical effects of microorganisms inhabiting nest-material remains before reproduction regarding the intensity of ectoparasitism suffered by 8-day-old nestling hoopoes. In accordance with the hypothesis, nestlings hatched in nest-boxes with autoclaved nest-material remains from the previous reproductive seasons suffered less from ectoparasites than those hatched in the control nest-boxes with nonautoclaved nest-material. Moreover, we found a positive association between the bacterial density of nest-material during the nestling phase and ectoparasitism intensity that was only apparent in nest-boxes with autoclaved nest-material. However, contrary to our expectations, nest bacterial load was positively associated with fledgling success. These results suggest a link between the community of microorganisms of nest-material remains and the intensity of ectoparasitism, and, on the other hand, that the nest bacterial environment during reproduction is related to fledging success. Here, we discuss possible mechanisms explaining the experimental and correlative results, including the possibility that the experimental autoclaving of nest material affected the microbiota of females and nestlings’ secretion and/or nest volatiles that attracted ectoparasites, therefore indirectly affecting both the nest bacterial environment at the nestling stage and fledging success., This research was funded by Ministerio de Ciencia, Innovación y Universidades, and European Regional Development Fund (FEDER; CGL2017-83103-P)., Peer reviewed

[Simple Summary] Symbiotic bacteria and fungi facilitate the acquisition of nutrients to their animal hosts, protect them against predators, parasites and diseases, and, in some ways, modulate complex animal behavior, including communication, by means of chemical signaling. However, odors of symbiotic bacterial origin would not only inform conspecifics of their animal host, but parasites and/or predators may also use those odors to detect their victims. We here review the role of bacterial symbionts on animal communication, and on interactions of their animal hosts with parasites and predators. Moreover, because microbial symbionts can have negative effects on their hosts facilitating predation and parasitism, these enemies could modulate the microbial community of animals, and we reviewed the available evidence supporting this idea. The inclusion of microorganisms in scenarios of communication, parasitism, and predation opens up new avenues of research that will contribute to understanding such interactions. We here elaborate some predictions and provide some guidance for future research., Microbial symbionts are nowadays considered of pivotal importance for animal life. Among the many processes where microorganisms are involved, an emerging research avenue focuses on their major role in driving the evolution of chemical communication in their hosts. Volatiles of bacterial origin may underlie chemical communication and the transfer of social information through signals, as well as inadvertent social information. We reviewed the role of microorganisms in animal communication between conspecifics, and, because the microbiome may cause beneficial as well as deleterious effects on their animal hosts, we also reviewed its role in determining the outcome of the interactions with parasites and predators. Finally, we paid special attention to the hypothetical role of predation and parasitism in driving the evolution of the animal microbiome. We highlighted the novelty of the theoretical framework derived from considering the microbiota of animals in scenarios of communication, parasitism, and predation. We aimed to encourage research in these areas, suggesting key predictions that need to be tested to better understand what is one of the main roles of bacteria in animal biology., This research was funded by Ministerio de Ciencia, Innovación y Universidades and European Regional Development Fund (FEDER) (CGL2017-83103-P, CGL2017-89063-P), by a predoctoral fellowship (MMA, FPU15/03420), and by the Ramón y Cajal programme (GT, RYC-2013-13040)., Peer reviewed

Animal coloration results from pigments, nanostructures, or the cosmetic use of natural products, and plays a central role in social communication. The role of cosmetic coloration has traditionally been focused in scenarios of sexual selection, but it could also take place in other contexts. Here, by using spotless starlings (Sturnus unicolor) as a model system, we explore the possibility that nestlings cosmetically use their intensely yellow-colored uropygial secretion to signal their genetic and/or phenotypic quality. In agreement with the hypothetical cosmetic use of the uropygial secretion, (i) video recorded nestlings collected secretion with the bill at the age
of feathering, (ii) cotton swabs turned to the color of secretion after rubbing with them nestlings’ gape, and (iii) gape and skin colorations correlated positively with that of secretion. Furthermore, we found that (iv) secretion coloration has a genetic component, and (v) associated positively with Vitamin E supplementation and (vi) with plasma carotenoid concentration, which highlights the informative value of nestling secretion. Finally, (vii) coloration of begging-related traits and of secretion of nestlings predicted parental feeding preferences. Consequently, all these results strongly suggest that the cosmetic use of colored uropygial secretion might also play a role in parent-offspring communication, complementing or amplifying information provided by the flamboyant colored gapes and skin of nestlings. The use of makeups by offspring for communication with relatives has been scarcely explored and we hope that these results will encourage further investigations in birds and other taxa with parental care., Ester Martínez Renau was financed by a predoctoral contract (PRE2018-085378, funded by Ministerio de Ciencia e Innovación/Agencia Estatal de Investigación/10.13039/501100011033 and by “European Science Fundation Investing in your future”), while the whole research group was supported by the project+ CGL2017-83103-P unded by Ministerio de Ciencia e Innovación/Agencia Estatal de Investigación/10.13039/501100011033 and by “Fondo Europeo de Desarrollo Regional, a way of making Europe”., Peer reviewed

Data file includes information about identity of each sampled individual, the species id, nest type, age, biometric measurements, the mean of the antagonistic halos shown against each 9 indicator bacteria, the antagonistic activity (average values of the width of antagonistic halos (mm) when tested against each of the nine indicator bacteria), the antagonistic range (Shannon index of the antagonistic activity) and the total density of bacteria on the gland., Ester Martínez Renau was financed by a predoctoral contract PRE2018-085378) while the whole research group received funds from the projects CGL2017-83103-P, PID2020-117429GB-C21 and PID2020-117429GB-C22, funded by the Ministerio de Ciencia e Innovación/Agencia Estatal de Investigación/10.13039/501100011033 and by “Fondo Europeo de Desarrollo Regional, a way of making Europe, Peer reviewed

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

Dataset 1 includes weekly estimations of median, maximum, minimum and range values of temperature and humidity in nest boxes, for three locations, two experimental treatments and in relation to dormice presence in February., Dataset 2 includes daily differences (between 9:00 am and 3:00 am) in temperature and humidity of open nest boxes occupied or not by dormice., Dataset 3 describes Carnus emergence patterns from each sample of nest material, including number of emerged flies (Carnus abundance), date of first emergence and emergence duration., Dataset 4 includes intensity of parasitism suffered by hoopoe nestlings during the breeding season after the experiment, showing number of Carnus, hatching date, brood size and nestling age., The dataset contains data collected in Guadix, Granada (southeast Spain) in 2020, in the framework of a research to evaluate the effects of overwinter occupancy of nest cavities on the incidence of ectoparasitism suffered by hoopoes (Upupa epops) in the next breeding season., The research group benefited from grants CGL2017-83103-P, CGL2017-89063-P, PGC2018-097426-B-C22, and PID2020-117429GB-C21 funded by MCIN/AEI/ 10.13039/501100011033 and by “ERDF A way of making Europe”. AJGN was funded by the Spanish Ministerio de Ciencia, Innovación y Universidades (Programa de Formación de Personal Investigador JAE Intro). GT was supported by the Ramón y Cajal Programme (RYC-2013-13040)., Peer reviewed

Cavities used for avian reproduction in spring often host other organisms for roosting during winter, which should affect their microclimatic conditions and, then, the patterns of emergence of nest-dwelling ectoparasites of birds and successful parasitism. To prove these cascade effects, we experimentally blocked the entrance of half of nest boxes previously used for reproduction by hoopoes (Upupa epops), recorded temperature and humidity during the fall–winter period, and evaluated abundance, emergence of ectoparasitic flies (Carnus hemapterus), and intensity of parasitism in hoopoe nestlings growing in open and blocked nest boxes the next spring. As expected, experimental treatment as well as the occupancy of open nest boxes by rodents (i.e., mainly dormice [Eliomys quercinus]) affected temperature and humidity, but did not predict the onset or duration of Carnus emergence. Moreover, flies were more abundant in open nest boxes and, among them, those occupied by dormice showed the lowest abundance. Finally, hoopoe nestlings developing in nest boxes blocked during winter experienced lower intensity of ectoparasitism than those in open nest boxes. These hitherto undescribed cryptic effects of overwinter occupants of nest cavities on subsequent emergence and viability of nest-dwelling ectoparasites would profoundly impact on the interaction with their avian hosts., The research group benefited from grants CGL2017-83103-P, CGL2017-89063-P, PGC2018-097426-B-C22, and PID2020-117429GB-C21 funded by MCIN/AEI/10.13039/501100011033 and by “ERDF: A way of making Europe.” We also benefit from facilities, including an apartment, provided by the city authorities of Guadix. Antonio José García-Núñez was funded by the Spanish Ministerio de Ciencia, Innovación y Universidades (Programa de Formación de Personal Investigador JAE Intro). Gustavo Tomás was supported by the Ramón y Cajal Programme (RYC-2013-13040). We thank the reviewers for their constructive comments., Peer reviewed

Similar to other signals, coloration of traits expressed by nestlings while they are begging for food are assumed to reflect their phenotypic conditions, and, therefore, they should be environmentally determined. However, these colorations are also species specifics and, thus, a genetic component explaining the evolution of nestling coloration should exist; a possibility rarely explored in the literature. Cross-fostering experiments allowed us to quantify genetic and environmental components of coloration of mouth, flanges and skin in a spotless starling Sturnus unicolor population. In addition, by supplementing some nestlings within each brood with non-pigmented antioxidants (Vitamin E or Vitamin complex), we explored the effects of food supplementation on coloration of begging-related traits and plasma carotenoid concentration. We detected significant genetic components of colorations of skin and mouth, but not of flanges or plasma carotenoid concentration. Moreover, experimental supplementation with Vitamin E affected UV coloration of mouth, flanges and skin, while vitamin-complex supplementation exclusively affected flange coloration. Furthermore, coloration of flanges, but not that of other traits, predicted plasma carotenoid concentration suggesting a direct link between these two traits. All these results suggest that coloration of begging-related traits is a multifaceted signal informing parents about genetic characteristics, and of short-term needs of nestlings, including antioxidative capacity, that parents could use to adjust feeding efforts, and deciding which chick to feed. Future research should focus on the parents' perspective, taking into account their feeding effort and feeding preferences in relation to genetically and/or environmentally determined coloration of begging-related traits. A free Plain Language Summary can be found within the Supporting Information of this article., We thank Manuel Martín-Vivaldi for his contribution to the experimental design. The research group benefits from facilities, including an apartment, provided by the city hall of Guadix where a small laboatory to quickly process the samples was installed. Ester Martínez Renau was financed by a predoctoral contract (PRE2018-085378) while the whole research group received funds from the Spanish Ministerio de Ciencia, Innovacion y Universidades and Fondo Europeo de Desarrollo Regional (FEDER; CGL2017-83103-P)., Peer reviewed

Symbiotic bacteria on animal hosts can prevent pathogenic bacterial infections by several mechanisms. Among them, symbiotic bacteria can indirectly enhance host's immune responses or, directly, produce antimicrobial substances against pathogens. Due to differences in life-style, different host species are under different risks of microbial infections. Consequently, if symbiotic bacteria are somewhat selected by genetically determined host characteristics, we would expect the antimicrobial properties of bacterial symbionts to vary among host species and to be distributed according to risk of infection. Here we have tested this hypothesis by measuring the antimicrobial ability of the bacterial strains isolated from the uropygial-gland skin of 19 bird species differing in nesting habits, and, therefore, in risk of microbial infection. In accordance with our predictions, intensity and range of antimicrobial effects against the indicator strains assayed varied among bird species, with hole-and open-nesters showing the highest and the lowest values, respectively. Since it is broadly accepted that hole-nesters have higher risks of microbial infection than open nesters, our results suggest that the risk of infection is a strong driver of natural selection to enhance immunocompetence of animals through selecting for antibiotic-producing symbionts. Future research should focus on characterizing symbiotic bacterial communities and detecting coevolutionary processes with particular antibiotic-producing bacteria within-host species., EM-R was financed by a predoctoral contract (PRE2018-085378) while the whole research group received funds from the projects CGL2017-83103-P, PID2020-117429GB-C21, and PID2020-117429GB-C22, funded by the Ministerio de Ciencia e Innovación/Agencia Estatal de Investigación/10.13039/501100011033 and by “Fondo Europeo de Desarrollo Regional, a way of making Europe.” The research group also benefits from facilities, including accommodation, provided by the City Hall of Guadix, where a small lab to quickly process the samples was installed., Peer reviewed

Parents might use signals of need or of quality to decide food provisioning among their offspring, while the use of one or another signal might depend on food availability. Begging success of nestlings of different quality (i.e., body size) would also depend on food availability, and we here explore the effect of experimental food supply in begging success of nestlings and in provisioning of female hoopoes (Upupa epops), a species with extreme hatching asynchrony and nestlings size hierarchy. We video-recorded food allocation of females, begging success of nestlings of different size, and the social context (i.e., the size category of the other nestlings that were begging for food) during periods when experimental food supply was or was not available in the same nests. We found that when experimental food supplementation was present, begging success of the intermediate, but not that of large or small-sized nestlings, increased. The experiment, however, did not affect the feeding preferences of females toward nestlings of different size. Moreover, when small nestlings were the only ones that were begging for food, their begging success decreased in the experimental period, and females used supplemented prey to feed themselves. Those results, on one hand, confirm the importance of food availability for the begging success of nestlings of particular sizes and, on the other, indicate that females prefer to use extra food for their own rather than for the smallest nestlings. We discuss possible mechanisms explaining the detected experimental effects and the adaptive and nonadaptive explanations of mothers ignoring the small nestlings., The research group received funds from the projects CGL2017-83103-P, PID2020-117429GB-C21 and PID2020-117429GB-C22, funded by the Ministerio de Ciencia e Innovación/Agencia Estatal de Investigación/10.13039/501100011033 and by “Fondo Europeo de Desarrollo Regional, a way of making Europe.” We also benefited from facilities, including an apartment, provided by the city authorities of Guadix., Peer reviewed

Sibling cannibalism is relatively common in nature, but its evolution in birds and certain other vertebrates with extended parental care had been discarded. Here, however, we demonstrate its regular occurrence in two European populations of the Eurasian hoopoe (Upupa epops) and explore possible adaptive and non-adaptive explanations. Results showed that sibling cannibalism was more frequently detected in Spain (51.7%) than in Austria (5.9%). In these two populations, the hoopoes laid similar clutch sizes, resulting in similar fledging production, but hatching failures were more frequent in the northern population. Consequently, having more nestlings condemned to die in the southern population may explain the higher incidence of sibling cannibalism. In accordance with this interpretation, hatching span and failure, but not breeding date, explained the probability of sibling cannibalism in the Spanish hoopoes, while all three variables predicted brood reduction intensity. Furthermore, experimental food supply reduced the probability of sibling cannibalism, but not the intensity of brood reduction. Finally, females allocated fewer resources to the smallest nestlings when they were going to starve, but not necessarily when they were going to be used as food for their siblings. These results suggest that hoopoes produce extra eggs that, in the case of reduced hatching failure and food scarcity, produce nestlings that are used to feed older siblings. These findings provide the first evidence that sibling cannibalism occurs regularly in a bird species, thus expanding our evolutionary understanding of clutch size, hatching asynchrony, parent-offspring conflict, infanticide, and sibling cannibalism in the animal kingdom., This study was supported by the Spanish Ministerio de Ciencia, Innovación y Universidades and European (FEDER) Funds (CGL2017-83103-P), Peer reviewed

Background: Extra-pair paternity (EPP) in birds provides benefits in terms of more offspring, and characteristics for maintenance of this behaviour have been the subject of investigation. Microorganisms are known to be transmitted during mating, especially when mating with multiple partners, and factors reducing this cost of multiple mating are expected. Further, plumage brightness and colour intensity have been shown to be important traits to benefits from multiple mating as predicted by sexual selection. The aim of this study was to investigate the relationship between the rate of extra-pair paternity and the relative size of the uropygial gland at the interspecific level, as the uropygial gland is an exocrine gland hypothesized to produce antiparasitic substances and further identified to affect plumage brightness. Because of the expected benefits of large uropygial gland in scenarios of sexual selection, we predicted a positive correlation with EPP. Methods: We collected information from the literature of uropygial gland size and frequency of extra-pair paternity of 60 avian species of different families and explored the predicted positive correlation between them. We did so with means of comparative analyses that considered phylogenetic relationship as random factor and included body mass as covariate. We used Markov chain Monte Carlo generalized linear mixed models that were weighted by number of nests used to estimate extra-pair paternity. Results: We detected a positive relationship between level of extra-pair paternity and uropygial gland size at an interspecific level. This finding is consistent with the prediction. Conclusions: We discuss the importance of this result in scenarios of sexual selection and argue that the detected relationship may have arisen by utilizing antiparasitic secretions through secondary sexual characters indicating parasite resistance., JJS received financial support by the Spanish Ministerio de Ciencia, Innovación y Universidades and European (FEDER) funds (CGL2017-83103-P)., Peer reviewed

Gut microbiota are essential for host health and survival, but we are still far from understanding the processes involved in shaping their composition and evolution. Controlled experimental work under lab conditions as well as human studies pointed at environmental factors (i.e., diet) as the main determinant of the microbiota with little evidence of genetic effects, while comparative interspecific studies detected significant phylogenetic effects. Different species, however, also differ in diet, feeding behavior, and environmental characteristics of habitats, all of which also vary interspecifically, and, therefore, can potentially explain most of the detected phylogenetic patterns. Here, we take advantage of the reproductive strategy of avian brood parasites and investigate gut microbiotas (esophageal (food and saliva) and intestinal) of great spotted cuckoo (Clamator glandarius) and magpie (Pica pica) nestlings that grow in the same nests. We also estimated diet received by each nestling and explored its association with gut microbiota characteristics. Although esophageal microbiota of magpies and great spotted cuckoos raised within the same environment (nest) did not vary, the microbiota of cloacal samples showed clear interspecific differences. Moreover, diet of great spotted cuckoo and magpie nestlings explained the microbiota composition of esophageal samples, but not of cloaca samples. These results strongly suggest a genetic component determining the intestinal microbiota of host and parasitic bird species, indicating that interspecific differences in gut morphology and physiology are responsible for such interspecific differences., We thank Manuel Martin-Vivaldi Martínez and Jarkko Rutila for discussion
on the interest of exploring genetic and environmental factors of gut
microbiotas of brood parasitic and host nestlings. The research group
benefits from facilities, including an apartment, provided by the city hall of
Guadix where a small lab to quickly process the samples was installed.
Financial support was provided by the Spanish Ministerio de Ciencia,
Innovación y Universidades and European (FEDER) funds (CGL2017-
83103-P). All procedures were conducted under licence from the
Environmental Department of the Regional Government of Andalucía, Spain
(reference SGYB/FOA/AFR). All applicable international, national, and/or
institutional guidelines for the care and use of animals were followed., 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

Background: Some parasites use olfactory cues to detect their hosts and, since bacterial symbionts are partially responsible for animal odours, they could influence host parasitism. By autoclaving nest materials of hoopoe (Upupa epops) nests before reproduction started, we explored the hypothetical links between host-associated bacteria, volatiles and parasitism. During the nestling stage, we (i) estimated the level of ectoparasitism by chewing lice (Suborder Mallophaga) in adult hoopoe females and by Carnus haemapterus flies in nestlings, and (ii) characterized microbial communities and volatile profiles of nest environments (nest material and nest cavity, respectively) and uropygial secretions. Results: Experimental nests had less diverse bacterial communities and more diverse volatile profiles than control nests, while occupants experienced lower intensity of parasitism in experimental than in control nests. The experiment also affected beta diversity of the microbial communities of nest material and of the volatiles of the nestling uropygial secretions. Moreover, microbial communities of uropygial secretions and of nest materials covaried with their volatile profiles, while the volatile profile of the bird secretions explained nest volatile profile. Finally, a subset of the volatiles and bacteria detected in the nest material and uropygial secretions were associated with the ectoparasitism intensity of both adult females and nestlings, and with fledging success. Conclusions: These results show that a component of animal odours is linked with the microbial communities of the host and its reproductive environment, and emphasize that the associations between bacteria, ectoparasitism and reproductive success are partially mediated by volatiles of bacterial origin. Future work should focus on mechanisms underlying the detected patterns., Open Access funding provided thanks to the CRUE-CSIC agreement with Springer Nature. The research group was supported by the projects CGL2017-83103-P, PID2020-117429 GB-C21 and PID2020-117429 GB-C22, funded by the Ministerio de Ciencia e Innovación/Agencia Estatal de Investigación/https://doi.org/10.13039/501100011033 and by “Fondo Europeo de Desarrollo Regional, a way of making Europe”. The research group also benefits from facilities, including accommodation, provided by the City Hall of Guadix, where a small lab to quickly process the samples was installed. JMPS enjoined a Ramon y Cajal scholarship (RYC2022-036147-I) funded by Ministerio de Ciencia, Innovación y Universidades (MCIN/AEI/https://doi.org/10.13039/501100011033 )., Peer reviewed

Bacteria have been suggested as being partially responsible for avian nest odours and, thus, volatiles from their metabolism could influence the intensity of selection pressures due to parasites detecting olfactory cues of their hosts. Here, we tested this hypothesis by exploring intraspecific and interspecific variability in microbial environments, volatile profiles and intensity of ectoparasitism by Carnus hemapterus in the nests of 10 avian species. As expected, we found that (i) alpha and beta diversity of microbial and volatile profiles were associated with each other. Moreover, (ii) alpha diversity of bacteria and volatiles of the nest environment, as well as some particular bacteria and volatiles, was associated with the intensity of parasitism at early and late stages of the nestling period. Finally, (iii) alpha diversity of the nest microbiota, as well as some particular bacteria and volatiles, was correlated with fledging success. When considering them together, the results support the expected links between the microbial environment and nest odours in different bird species, and between the microbial environment and both ectoparasitism intensity and fledging success. Relative abundances of particular volatiles and bacteria predicted ectoparasitism and/or fledging success. Future research should prioritise experimental approaches directed to determine the role of bacteria and volatiles in the outcomes of host-ectoparasite interactions., The research group was supported by the projects CGL2017-83103-P, PID2020-117429GB-C21 and PID2020-117429GB-C22, funded by the Ministerio de Ciencia e Innovación/Agencia Estatal de Investigación/10.13039/501100011033 and by ’Fondo Europeo de Desarrollo Regional, a way of making Europe’. The research group also benefits from facilities, including accommodation, provided by the City Hall of Guadix, where a small laboratory to quickly process the samples was installed. Juan M. Peralta-Sánchez enjoined a Ramon y Cajal scholarship (RYC2022-036147-I) funded by Ministerio de Ciencia, Innovación y Universidades (MCIN/AEI/10.13039/501100011033) at University of Seville.

The use of cosmetic substances in communication is widespread in animals. Birds, for instance, use their uropygial secretion as a cosmetic in scenarios of sexual selection and parent-offspring communication. This secretion harbours symbiotic bacteria that could mediate the synthesis of pigments for cosmetic colouration. Here, we investigate the association between bacteria and the conspicuous yellow secretion used by spotless starling (Sturnus unicolor) nestlings to stain their mouths, and hypothesize a possible role of bacteria in the colour production. We also experimentally explore how nestling oxidizing condition influences the microbiota, suggesting that the possible bacterial-mediated coloured secretion acts as a reliable honest signal. An antioxidant supplementation experiment, previously known to affect secretion and mouth colouration, was conducted to assess its impact on the microbial community of secretions from control and experimental siblings. Antioxidant supplementation increased richness and phylogenetic diversity of the secretion’s microbiota. Moreover, the microbiota’s alpha and beta diversity, and the abundance of two bacterial genera (Parabacteroides and Pseudogracilibacillus), correlated with secretion colour. These findings demonstrate that antioxidant condition influences the gland microbiota of starling nestlings, and suggest a link between bacteria and the colouration of their cosmetic secretion. Alternative explanations for the detected links between bacteria and colouration are discussed., E.M.-R. was financed by a predoctoral contract (PRE2018-085378) while the whole research group received funds from grants CGL2017-83103-P, PID2020-117429GB-C21 and PID2020-117429GB-C22 funded by MICIU/AEI /10.13039/501100011033., Peer reviewed

[Background]: Animal bacterial symbionts are established early in life, either through vertical transmission and/or by horizontal transmission from both the physical and the social environment, such as direct contact with con- or heterospecifics. The social environment particularly can influence the acquisition of both mutualistic and pathogenic bacteria, with consequences for the stability of symbiotic communities. However, segregating the effects of the shared physical environment from those of the social interactions is challenging, limiting our current knowledge on the role of the social environment in structuring bacterial communities in wild animals. Here, we take advantage of the avian brood-parasite system of Eurasian magpies (Pica pica) and great spotted cuckoos (Clamator glandarius) to explore how the interspecific social environment (magpie nestlings developing with or without heterospecifics) affects bacterial communities on uropygial gland skin., [Results]: We demonstrated interspecific differences in bacterial community compositions in members of the two species when growing up in monospecific nests. However, the bacterial community of magpies in heterospecific nests was richer, more diverse, and more similar to their cuckoo nest-mates than when growing up in monospecific nests. These patterns were alike for the subset of microbes that could be considered core, but when looking at the subset of potentially pathogenic bacterial genera, cuckoo presence reduced the relative abundance of potentially pathogenic bacterial genera on magpies., [Conclusions]: Our findings highlight the role of social interactions in shaping the assembly of the avian skin bacterial communities during the nestling period, as exemplified in a brood parasite—host system., Open Access funding provided thanks to the CRUE-CSIC agreement with Springer Nature. EM-R was financed by a predoctoral contract (PRE2018-085378) while the whole research group received funds from the projects CGL2017-83103-P, PID2020-117429GB-C21, and PID2020-117429GB-C22, funded by the Ministerio de Ciencia e Innovación/Agencia Estatal de Investigación/https://doi.org/10.13039/501100011033 and by “Fondo Europeo de Desarrollo Regional, a way of making Europe.”, Peer reviewed

Dataset and R scripts used in the manuscript "Environmental factors associated with nesting habits and age shape the composition and connection between skin and uropygial gland microbiomes of birds", currently under review in Journal of Animal Ecology. It includes the R scripts used for data analysis, along with all necessary data to reproduce the results. The study investigates how species, age and environmental factors (nesting habits), influence the microbiota of the uropygial gland (skin and secretion) across 26 bird species, using 16S rRNA amplicon sequencing to characterise bacterial communities., Ester Martínez-Renau was financed by a predoctoral contract (PRE2018-085378) while the whole research group received funds from the projects CGL2017-83103-P, PID2020-117429GB-C21, and PID2020-117429GB-C22, funded by the Ministerio de Ciencia e Innovación/Agencia Estatal de Investigación/10.13039/501100011033., A. ID - Identification name of the fastq files B. Correspondence - Short identification name of the fastq files C. sample_type - Whether the sample is from the uropgyial gland skin or the uropygial secretion D. Species - Host species name E. Ring - Individual ring number for each bird sampled F. Adults_nd_nestl - Whether we've got adult and nestling samples of the same species G. same_indiv_gland_secre - Wether we've got samples of the same individual for both gland skin and uropygial secretion H. Nest - Nest identification I. Locality - Sampling locality J. Nest_type - Whether the sampled species is open-cup nester or cavity-nester K. Age - Whether the bird sampled is an adult or a nestling L. Age_Nest - Combinación of Age and Nest_type columns M. Date - Sampling date N. Tarsus - Tarsus length O. Weight - Weight P. Height gl - Gland height Q. Length gl - Gland length R. Width gl - Gland width, Peer reviewed

Bacterial communities on skin and feathers can act as a critical line of defence against pathogenic infections in birds and may originate from secretions produced by the uropygial gland. These secretions reach the bird integument during preening, with the preening effort possibly determining the connectivity between uropygial gland and integument microbiomes.
The risk of pathogen infections depends on a number of variables, including environmental conditions (i.e. temperature and humidity), species identity, life-history traits (i.e. cavity vs. open-cup nesters) and life stage (i.e. age). Bacterial symbionts of the host, particularly those of the uropygial gland, may counter such pathogenic infections. We therefore hypothesise that bacterial communities of the uropygial gland differ among host species, age and nesting habits, with higher bacterial diversity in nestlings due to their immature immune system, and in cavity nesters due to potentially increased pathogen exposure.
We examined this using 16S rRNA metabarcoding of bacterial communities of the uropygial secretion (N = 352) and uropygial gland skin (N = 339) of nestlings and adults of 26 bird species from 14 families in southern Spain.
In accordance with the hypotheses, we find species-specific differences in bacterial communities of uropygial gland skin and secretion, as well as an effect of age, with nestlings showing a higher bacterial diversity, especially in the uropygial gland skin. Additionally, the microbiotas of cavity-nesting species are more diverse and heterogeneous than those of open nesters, with these effects more pronounced in adult and uropygial secretions. Finally, the uropygial gland is relatively larger in cavity- than in open-nester species, which suggests that cavity nesters preen more often than the open nesters. Moreover, we found a stronger sharing of secretion and skin microbes in cavity nesters and nestlings compared to adults and open nesters.
Overall, our findings on the effects of age and nest type on structuring bird uropygial gland skin and secretion microbiota imply that age and pathogen risks related to nest environment could drive the external microbiome assembly in birds., Ester Martínez-Renau was financed by a predoctoral contract (PRE2018-085378) while the whole research group received funds from the projects CGL2017-83103-P, PID2020-117429GB-C21, PID2020-117429GB-C22, PID2024-159017NB-C31 and PID2024-159017NB-C32 funded by the Ministerio de Ciencia e Innovación/Agencia Estatal de Investigación/10.13039/501100011033., Peer reviewed