Danielle J. Whittaker

Natural Selection on Testosterone Production in a Wild Songbird Population

Because of their role in mediating life‐history trade‐offs, hormones are expected to be strongly associated with components of fitness; however, few studies have examined how natural selection acts on hormonal variation in the wild. In a songbird, the dark‐eyed junco (Junco hyemalis), field experiments have shown that exogenous testosterone alters individuals’ resolution of the survival‐reproduction trade‐off, enhancing reproduction at the expense of survival. Here we used standardized injections of gonadotropin‐releasing hormone (GnRH) to assay variation in the testosterone production of males. Using measurements of annual survival and reproduction, we found evidence of strong natural selection acting on GnRH‐induced increases in testosterone. Opposite to what would be predicted from the survival‐reproduction trade‐off, patterns of selection via survival and reproduction were remarkably similar. Males with GnRH‐induced testosterone production levels that were slightly above the population mean were more likely to survive and also produced more offspring, leading to strong stabilizing selection. Partitioning reproduction into separate components revealed positive directional selection via within‐pair siring success and stabilizing selection via extrapair mating success. Our data represent the most complete demonstration of natural selection on hormones via multiple fitness components, and they complement previous experiments to illuminate testosterone’s role in the evolution of life‐history trade‐offs.

Hormonal, Behavioral, and Life-History Traits Exhibit Correlated Shifts in Relation to Population Establishment in a Novel Environment

Climate change, habitat alteration, range expansions, and biological invasions are all predicted to require rapid shifts in multiple traits including behavior and life history, both for initial population establishment and subsequent adaptation. Hormonal mechanisms likely play a key role in facilitating or constraining plastic and genetic responses for suites of traits, but few studies have evaluated their role in shaping contemporary adaptation or diversification. We examined multiple phenotypic adjustments and associated hormonal changes following a recent (early 1980s) colonization event, in which a temperate-breeding songbird, the dark-eyed junco (Junco hyemalis), became established in the Mediterranean climate of San Diego, California. The milder climate has led to an extended breeding season and year-round residency, and we document shifts in multiple sexually selected behaviors and plumage traits. Testosterone titers in San Diego were elevated for longer but with a lower peak value compared to a nearby native-range population, and correlations between testosterone and related traits were similar within and among populations. A common garden study indicated that changes in testosterone likely represent plastic responses to the less seasonal environment of the city, providing the context against which subsequent genetic changes in morphology likely occurred. We argue that correlated shifts in multiple traits, organized by underlying physiology, may be a generally important element of many successful adjustments to changing environments.

Bird odour predicts reproductive success

Although the importance of chemical communication in birds has long been overlooked or doubted, volatile compounds in avian preen secretions have been shown to covary with traits including species, sex and breeding condition, and thus may be useful mate recognition cues. Here we demonstrate for the first time that these compounds may reliably predict reproductive success in a North American songbird, the dark-eyed junco, Junco hyemalis. Several compounds associated with sex differences in this species varied with reproductive success, such that females with a more ‘female-like’ volatile profile and males with a more ‘male-like’ profile produced more genetic offspring. A males preen oil volatile compounds also predicted his success in rearing offspring in his home nest: males with a higher abundance of ‘male-like’ compounds had more surviving nestlings, including offspring sired by extrapair males. Finally, males with a higher abundance of ‘female-like’ compounds had more extrapair offspring in their home nests. Our results suggest that odours correlate with reproductive success and thus have qualities that could allow them to serve as reliable mate assessment cues in birds.

De novo transcriptome sequencing in a songbird, the dark-eyed junco (Junco hyemalis): genomic tools for an ecological model system

BackgroundThough genomic-level data are becoming widely available, many of the metazoan species sequenced are laboratory systems whose natural history is not well documented. In contrast, the wide array of species with very well-characterized natural history have, until recently, lacked genomics tools. It is now possible to address significant evolutionary genomics questions by applying high-throughput sequencing to discover the majority of genes for ecologically tractable species, and by subsequently developing microarray platforms from which to investigate gene regulatory networks that function in natural systems. We used GS-FLX Titanium Sequencing (Roche/454-Sequencing) of two normalized libraries of pooled RNA samples to characterize a transcriptome of the dark-eyed junco (Junco hyemalis), a North American sparrow that is a classically studied species in the fields of photoperiodism, speciation, and hormone-mediated behavior.ResultsFrom a broad pool of RNA sampled from tissues throughout the body of a male and a female junco, we sequenced a total of 434 million nucleotides from 1.17 million reads that were assembled de novo into 31,379 putative transcripts representing 22,765 gene sets covering 35.8 million nucleotides with 12-fold average depth of coverage. Annotation of roughly half of the putative genes was accomplished using sequence similarity, and expression was confirmed for the majority with a preliminary microarray analysis. Of 716 core bilaterian genes, 646 (90 %) were recovered within our characterized gene set. Gene Ontology, orthoDB orthology groups, and KEGG Pathway annotation provide further functional information about the sequences, and 25,781 potential SNPs were identified.ConclusionsThe extensive sequence information returned by this effort adds to the growing store of genomic data on diverse species. The extent of coverage and annotation achieved and confirmation of expression, show that transcriptome sequencing provides useful information for ecological model systems that have historically lacked genomic tools. The junco-specific microarray developed here is allowing investigations of gene expression responses to environmental and hormonal manipulations – extending the historic work on natural history and hormone-mediated phenotypes in this system.

Role of Testosterone in Stimulating Seasonal Changes in a Potential Avian Chemosignal

Songbird preen oil contains volatile and semivolatile compounds that may contain information about species, sex, individual identity, and season. We examined the relationship between testosterone (T) and the amounts of preen oil volatile and semivolatile compounds in wild and captive dark-eyed juncos (Junco hyemalis). In wild males and females, we observed an increase in volatile compound relative concentration early in the breeding season. This increase mirrored previously described seasonal elevation in T levels in wild males and females, suggesting a positive relationship between hormone levels and preen gland secretions, and a possible role for these secretions in signaling receptivity. In females, the greatest relative concentrations of most compounds were observed close to egg laying, a time when steroid hormones are high and also the only time that females respond to an injection of gonadotropin-releasing hormone with a short-term increase in T. In a study of captive juncos held on short days, we asked whether the seasonal increases observed in the wild could be induced with experimental elevation of T alone. We found that exogenous T stimulated the production of some volatile compounds in non-breeding individuals of both sexes. However, of the 15 compounds known to increase during the breeding season, only four showed an increase in relative concentration in birds that received T implants. Our results suggest that testosterone levels likely interact with other seasonally induced physiological changes to affect volatile compound amounts in preen oil.

Robust behavioral effects of song playback in the absence of testosterone or corticosterone release.

Some species of songbirds elevate testosterone in response to territorial intrusions while others do not. The search for a general explanation for this interspecific variation in hormonal response to social challenges has been impeded by methodological differences among studies. We asked whether song playback alone is sufficient to bring about elevation in testosterone or corticosterone in the dark-eyed junco (Junco hyemalis), a species that has previously demonstrated significant testosterone elevation in response to a simulated territorial intrusion when song was accompanied by a live decoy. We studied two populations of juncos that differ in length of breeding season (6-8 vs. 14-16 weeks), and conducted playbacks of high amplitude, long-range song. In one population, we also played low amplitude, short-range song, a highly potent elicitor of aggression in juncos and many songbirds. We observed strong aggressive responses to both types of song, but no detectable elevation of plasma testosterone or corticosterone in either population. We also measured rise in corticosterone in response to handling post-playback, and found full capacity to elevate corticosterone but no effect of song class (long-range or short-range) on elevation. Collectively, our data suggest that males can mount an aggressive response to playback without a change in testosterone or corticosterone, despite the ability to alter these hormones during other types of social interactions. We discuss the observed decoupling of circulating hormones and aggression in relation to mechanisms of behavior and the cues that may activate the HPA and HPG axes.

Chemosignaling diversity in songbirds: chromatographic profiling of preen oil volatiles in different species.

Large foraging seabirds are known to navigate to food sources using their excellent sense of smell, but much less is known about the use of olfaction by the songbirds (passerine birds). Some evidence of individual recognition based on the bird preen oil volatile organic compound (VOC) compositions, which is the main odor source in birds, have been reported for dark-eyed junco and house finch. In this study we have investigated preen oil VOCs in 16 different songbird species and two other small bird species in order to determine whether the VOC compositions follow phylogenetic and evolutionary relatedness. We have used the stir bar sorptive extraction (SBSE) methodology followed by gas chromatography-mass spectrometry (GC-MS) to determine preen oil VOCs during the long light summer conditions for mostly wild caught birds. Large diversity among the VOC compositions was observed, while some compound classes were found in almost all species. The divergent VOC profiles did not follow the phylogenetic family lines among the bird species. This suggests that songbirds may use VOC odors as a mate recognition cue.

The Diversity of Small Apes and the Importance of Population-Level Studies

Most primatologists, biologists, and laypeople agree that gibbons, with their incredible acrobatic displays and haunting duets, are absolutely marvelous animals. For all of their beauty and grace, however, they have received relatively little attention from the scientific community and the public alike. This volume is an attempt to begin addressing this problem by summarizing the progress of gibbon studies to date, identifying the key areas for future research, and cautioning against the belief that we already know everything worth knowing about gibbons. Over two decades have passed since the publication of the seminal volume The Lesser Apes: Evolutionary and Behavioural Biology (Preuschoft et al. 1984). That book was based on a conference, the first of its kind focusing on gibbons, held in 1980 inUlm,Germany.The Lesser Apes comprises a thorough summary of progress in gibbon studies up to that time, focusing on conservation, functional morphology, ecology, social behavior, and evolutionary biology. The contributors identified several areas that required additional study, including calls and songs; the basic behavioral biology of little understood species (Hoolock spp., Nomascus spp.); molecular phylogenetic studies, particularly of Hoolock and Hylobates klossii; and the fossil record. In the decades since the publication of The Lesser Apes, progress toward many of these goals has been made. Twenty years later, gibbonologists gathered again, at two International Primatological Society symposia: ‘‘Gibbon Diversity and Conservation’’ in Beijing in 2002 and ‘‘Wild Gibbons as Members of Populations’’ in Torino in 2004. This book is the product of those two symposia and has been assembled in recognition of the fact that a great deal of progress has been made in the field since 1984, allowing new perspectives on gibbon socioecology.

Bacterial Communities Associated with Junco Preen Glands: Preliminary Ramifications for Chemical Signaling

Intraspecific chemical communication may play a significant role in avian behavior. Preen oil secreted by the uropygial gland emits volatile compounds that vary with many aspects of bird biology and may figure prominently in mate choice. Many of these compounds are known end products of bacterial metabolism in other environments. The fermentation hypothesis for chemical recognition suggests that symbiotic bacteria in mammalian scent glands produce volatile odorants that are used as recognition cues by the host animals and that variation in these bacterial communities contributes to variation in the animal scents. We preliminarily evaluated whether this hypothesis could apply to birds by sampling and sequencing bacterial communities associated with the preen gland of adult breeding dark-eyed juncos (Junco hyemalis). The most common operational taxonomic units (OTUs), defined by the V4 region of the 16S rRNA gene, belonged to the phyla Actinobacteria, Firmicutes, and Proteobacteria. Most of the identified bacterial genera contain species that are known odor producers. Most notably, Burkholderia and Pseudomonas species consistently produce 9 of the 17 volatile compounds whose production is either upregulated during the breeding season or differs between the sexes in juncos. Other genera have documented antifungal or antibacterial properties and may provide other valuable services to the birds. We found no effect of host sex on bacterial community composition or structure. Instead, paired males and females clustered together, suggesting that individuals that have frequent contact may develop similar microbial communities quickly. Our study suggests that the fermentation hypothesis for chemical recognition, originally formulated for mammals, may apply to birds as well, opening new pathways for avian-microbial research.

Social Environment Has a Primary Influence on the Microbial and Odor Profiles of a Chemically Signaling Songbird

Chemical signaling is an underappreciated means of communication among birds, as may be the potential contributions of symbiotic microbes to animal chemical communication in general. The dark-eyed junco (Junco hyemalis) produces and detects volatile compounds that may be important in reproductive behavior. These compounds are found in preen oil secreted by the uropygial gland, and this gland supports diverse bacterial communities including genera known to produce some of these volatile compounds. We investigated the relative contributions of shared environments and genetic relatedness in shaping juncos’ symbiotic bacterial communities, and investigated whether these bacterial communities underlie juncos’ chemical signaling behavior. We sampled parents and nestlings at 9 junco nests during one breeding season at Mountain Lake Biological Station in Virginia, USA. From each individual, we collected swabs of the uropygial gland and the cloaca, preen oil, and a small blood sample for paternity testing. We characterized junco bacterial communities through 16S rRNA gene surveys and preen oil volatile compounds via gas chromatography-mass spectrometry. Nest membership and age class had the strongest influence on the structure of bacterial and volatile profiles. We compared father-offspring similarity based on paternity, and nestling similarity in nests containing full siblings and half siblings, and found that relatedness did not noticeably affect bacterial or volatile profiles. While we cannot rule out an influence of genetic relatedness on these profiles, it is clear that shared environments are more influential in shaping bacterial and volatile profiles among juncos. We did not find significant covariation between individual bacterial and volatile profiles. Possible explanations for this result include: 1) bacteria do not underlie volatile production; 2) ample redundancy in volatile production among bacterial types obscures covariation; or 3) the relationship is confounded by the fact that, unlike glands exclusively dedicated to chemical communication, uropygial glands have multiple functions, and symbiotic bacteria are hypothesized to contribute to each of these. Therefore, different bacteria may contribute to different phenotypes of the avian holobiont. Future work will include cultivation, metabolomic, genomic, and behavioral assay approaches to tease these scenarios apart.