My doctoral thesis aimed to contribute to our understanding of the evolution in songbirds. What are the ecological and behavioral forces that shape the extraordinary diversity in songbirds? I specifically focused on the role of seasonality, song, and their underlying endocrine regulations as potential drivers leading to population divergence, and eventually speciation.
Evolution requires inter-individual variation, which is subject to natural and sexual selection, and can thereby lead to population divergence. As a first step to understand reproductive isolation mechanisms it is crucial to evaluate the perception of geographic variation in sexually selected traits.
Specifically, signaling in the context of resource defense or mate attraction may enhance reproductive isolation (Grant and Grant 2002; Edwards et al. 2005; Price and Sol 2008; Podos 2010). Therefore, I first examined the geographic variation of song and morphological traits in Stonechats, and then tested their behavioral response to phenotypes from different populations. European stonechats discriminated phenotypic traits from different populations during playback and decoy experiments, and this was separately confirmed for African stonechats. Furthermore, both sexes discriminated consistently between stimuli from different populations, responding more strongly to song and morphological traits of local than foreign birds. This finding corresponds well with the local adaptation hypothesis, i.e. birds, which select mates from their natal regions, may gain fitness benefits because their young will more likely express adaptations to local ecological conditions (Marler and Tamura 1962; Nottebohm 1969; Baker 1975; Searcy 1992; Kawecki and Ebert 2004). Interestingly, my results further suggest that song may be indeed the stronger signal for Stonechats than genetically inherited morphological traits. Culturally transmitted song may evolve more quickly, and by promoting reproductive isolation could play a more essential role in incipient speciation (Grant and Grant 2012).
The relationship of local adaptation with associated signaling and female preference may also be affected by a species’ life history (Helbig 2003). To gain insights into the geographic variation of song on a larger geographical scale, I therefore studied song traits in relation to the diverse life histories of Stonechats. I focused on a pace of life axis, which includes both demographic characteristics, such as longevity and reproductive output, and seasonal life history traits such as migration or temporary territoriality. In particular, life histories associated with the seasonality of the environment may have striking implications for evolutionary dynamics. My background findings that tropical stonechats follow a slower pace of life than temperate stonechats are concordant with previous studies in Stonechats. Intriguingly, some aspects of song may be indeed linked to different life histories along a latitudinal gradient. In the northern hemisphere, migratory stonechats increased their song activity
during the beginning of the breeding season, the time of year when it is most essential to establish territories and attract prospecting females. In sedentary African stonechats, on the other hand, territory establishment and mate attraction only seem to play a minor role for reproduction, since they generally occupy the same territories year-round and form long-term pair bonds. During the short period of the females’ maximum fecundity, i.e. nestbuilding, afrotropical males adjusted their song traits to accommodate short-range signaling within close proximity to fertile females. Thereby, they could avoid eavesdropping of other males, and thus prevent extra pair paternity of their mates. Taken together, these findings imply that song of temperate stonechats may be under stronger sexual selection than the sedentary afrotropical populations ((Collins et al. 2009, 2011) but see (Byers 2011)).
Seasonal activities such as migration of temperate species may elaborate song as a sexually selected trait because migrants are typically more time constrained for breeding. In migratory populations the strong pressure to find a mate, breed quickly and make the most of the brief breeding season may drive the evolution of more elaborated song than in sedentary populations (Read and Weary 1992;
Irwin et al. 2001, 2008; Mountjoy and Leger 2001).
I am also keenly interested in the underlying endocrine regulation of song in birds with diverse life histories. In general, testosterone is known to activate reproductive behaviors, such as song and territoriality (reviewed in (Hau 2007)). Phenotypic variation in testosterone levels seems to follow a latitudinal trend along the pace of life axis. Tropical birds typically lack the dramatic seasonal fluctuations in testosterone shown by males from northern latitudes (Dittami and Gwinner 1985;
Reyer et al. 1986; Wingfield et al. 1991; Wikelski et al. 2003a; Onofrei et al. 2004; Garamszegi et al.
2008). In the tropics, testosterone can be involved in regulating song and territorial behavior in year-round territorial birds, although it may remain at low plasma levels throughout the year (Hau et al.
2008). In the stonechats I studied, testosterone was elevated during breeding in all populations regardless of latitude. Compared to other tropical study species, African stonechats experience a rather fluctuating environment with reoccurring patterns of seasonality. They adjust their reproductive timing to such seasonal patterns to optimize their fitness. I suggest that in seasonally breeding tropical species, such as the stonechat, testosterone is elevated during breeding much as in higher latitude birds to amplify territorial behavior and mate guarding during the females’ fertile period. Thus, testosterone may facilitate the seasonal timing of reproductive behaviors in tropical species with short, synchronized breeding seasons (Goymann et al. 2004; Goymann and Landys 2011). A previous study found similar seasonal testosterone patterns in Stonechats and proposed that such brief, but pronounced peaks of testosterone may be common in tropical birds, but they may simply be missed if the exact breeding stage of individual birds is not known (Goymann et al. 2006). In light of the seasonal diversity of tropical environments, the pace of life theory and its underlying endocrine control mechanisms should be reconsidered in future studies.
Based on this knowledge, the question arises why birds do not express elevated testosterone throughout the entire year. Testosterone is one important physiological factor proposed to mediate the trade-off between reproduction and survival (Stearns 1989). In particular, it has important pleiotropic effects on resource allocation for mate attraction, territorial behavior, reproductive effort, and associated signal plasticity, while often simultaneously decreasing fitness by suppressing traits such as immune function and parental care. The seasonal adaptation of endocrine regulation may have evolved to circumvent these negative implications of tonically elevated testosterone levels.
In this context, a very astonishing life history trait of some temperate species is the extension of territoriality and associated song into the non-breeding season (Hau et al. 2000). I evaluated the associations of seasonality, song and the endocrine regulation of territoriality in the Black redstart.
Using phenotypic engineering, i.e. hormonal manipulation, during simulated territorial intrusions (Ketterson et al. 1996; Hau 2007), I found that during breeding placebo implanted but not testosterone blocked males displayed substantial changes in song structure in response to a territorial intruder.
During the non-breeding season, when testosterone levels are naturally low, placebo implanted and testosterone implanted males sang a reduced song, similar to breeding males with blocked testosterone. These results imply that during breeding testosterone seems to facilitate context-dependent changes in song structures. While song activity and structure were regulated by testosterone, non-vocal territorial behaviors were independent of testosterone. The behavioral response to simulated territorial intrusions was similar during breeding and non-breeding. Previous studies observed that in several species outside the breeding season, males defend territories with extended borders, overlapping boundaries or even share these territories with other birds (Wingfield and Monk 1992). Thus, territorial behavior may be only activated by testosterone in a reproductive context, when it is crucial to defend breeding territories (Logan 1992; Schwabl 1992; Wingfield and Monk 1992; Onofrei et al. 2004).
These findings are relevant for my studies of African stonechats, which are well suited for understanding the temporal plasticity of endocrine mechanisms in diversifying seasonal environments. In East Africa, stonechats exhibit considerable differences in timing of breeding between populations on a local scale. In late April 2013, I compared testosterone levels of geographically close populations with asynchronous breeding onsets. Testosterone levels of breeding birds in Kinangop peaked during this time, whereas levels of non-breeding birds from the allochronic population, Mount Meru, were extremely low or could not be detected at all. The highest peak of testosterone levels of this population occurred during their breeding season in late October.
Testosterone increase strongly depends on testicular growth previous to breeding. It has been shown that testicular size underlies a rigid circannual cycle and takes about four weeks to fully grow into
breeding condition (Helm et al. 2009). These circannual patterns of testicular size and associated testosterone secretion may limit the opportunity for allochronic populations to interbreed, and thus may promote population divergence in African stonechats.
A central aim of the thesis is to contribute to the understanding of the evolutionary dynamics of allochronic populations in African stonechats. I set out to investigate the genetic, song and morphology divergence, and specifically tested for relative contributions of allochronic, spatial and behavioral isolation leading to the present population structure. The results revealed that allochronic populations within the Great Rift Valley, and geographically separated populations from the Eastern Arc Mountains, are genetically differentiated. I examined the environmental connectivity between populations to quantify the relative temporal and spatial contributions to reproductive isolation.
Synchronic and allochronic populations along the escarpment of the Great Rift Valley showed similar spatial connectivity, which strongly indicates genetic divergence as the result of allochronic isolation.
In contrast, the higher genetic differentiation between populations of the two mountain ranges is rather caused by strong spatial separation. The maximum-likelihood population-effects model, which explained the genetic structure of all twelve populations best, included both temporal and spatial ecological factors. However, the allochronic diversification may have occurred rather recently compared to the more ancient geographic diversification between populations of the two mountain ranges. A preliminary estimate of the divergence time based on the mitochondrial cytochrome b indicates that the genetic divergence between allochronic populations may have commenced at least 100,000 years ago, in contrast to the considerably earlier divergence time between spatially separated populations dating to about 600,000 years ago.
A previous study on captive Saxicola species revealed that birds did not modify timing in response to a mate, resulting in decreased reproductive success (Gwinner et al. 1995b; Helm et al. 2009; Bradshaw and Holzapfel 2010). Thus, species such as Stonechats, in which timing of breeding is highly heritable, are expected to have limited phenological plasticity, implying that allochronic populations would experience highly reduced effective gene flow even if they were connected by dispersal (Hendry and Day 2005; Helm et al. 2009). To our knowledge this is the second study providing evidence for incipient allochronic isolation in tetrapods. Such allochronic reproductive isolation mechanisms may drive evolutionary diversification and ultimately speciation, and may be more common than currently recognized.
Remarkably, the genetic structure is strongly concordant with patterns of song and morphology divergence. Concordant with the genetic structure, song traits of all three regions differed from each other, and followed a similar pattern as the genetic structure, whereas morphology traits of allochronic populations from the Great Rift Valley were similar and only differed from the spatially isolated
populations of the Eastern Arc Mountains. As mentioned previously, the allochronic diversification may have occurred rather recently compared to the more ancient geographic diversification between populations of the two mountain ranges. The high diversification of song may indicate that the cultural inheritance of song may be a stronger driver for population divergence than genetically inherited morphology, and thus may play a key role in incipient speciation.
In this study, both sexes distinguished local morphological and especially acoustic phenotypes from those of allochronic populations consistently. Females showed preference for song of mates with synchronized reproductive timing. Female preference for familiar vocalizations has been shown in some captive and field experiments by increased copulation-solicitation displays to standardized playback (Baker 1982, 1983; Searcy et al. 2002; Danner et al. 2011), whereas in Stonechats it was measured by approach of the stimulus and associated behaviors. Although the fine acoustic discrimination ability of Stonechats suggests potential behavioral barriers, its implications for geographic isolation are not fully clear, partly depending on song plasticity, and ultimately on the mechanisms involved in song learning. Birdsong is typically learned during a sensitive period early in life. In species that show geographic discrimination, males that subsequently disperse into ranges of other populations would face reduced mating prospects if an acoustic signature of the natal population remains in their repertoire (reviewed in (Podos and Warren 2007). In Stonechats, both sexes had similar sensitivity to incipient behavioral barriers between allochronic populations. The results demonstrate that variation in sexually selected traits may contribute to geographic isolation over relatively short distances, and thereby foster local adaptation.