German Ixodes inopinatus samples may not actually represent this tick species.

Ticks are important vectors of human and animal pathogens, but many questions remain unanswered regarding their taxonomy. Molecular sequencing methods have allowed research to start understanding the evolutionary history of even closely related tick species. Ixodes inopinatus is considered a sister species and highly similar to Ixodes ricinus, an important vector of many tick-borne pathogens in Europe, but identification between these species remains ambiguous with disagreement on the geographic extent of I. inopinatus. In 2018-2019, 1583 ticks were collected from breeding great tits (Parus major) in southern Germany, of which 45 were later morphologically identified as I. inopinatus. We aimed to confirm morphological identification using molecular tools. Utilizing two genetic markers (16S rRNA, TROSPA) and whole genome sequencing of specific ticks (n = 8), we were able to determine that German samples, morphologically identified as I. inopinatus, genetically represent I. ricinus regardless of previous morphological identification, and most likely are not I. ricinus/I. inopinatus hybrids. Further, our results showed that the entire mitochondrial genome, let alone singular mitochondrial genes (i.e., 16S), is unable to distinguish between I. ricinus and I. inopinatus. Our results suggest that I. inopinatus is geographically isolated as a species (northern Africa and potentially southern Spain and Portugal) and brings into question whether I. inopinatus exists in central Europe. Our results highlight the probable existence of I. inopinatus and the power of utilizing genomic data in answering questions regarding tick taxonomy.

Are animal personality, body condition, physiology and structural size integrated? A comparison of species, populations and sexes, and the value of study replication.

A major question in behavioural ecology is why behaviour, physiology and morphology are often integrated into syndromes. In great tits, Parus major, for example, explorative males are larger (vs. smaller) and leaner (vs. heavier) compared to less explorative individuals. Unfortunately, considerable debate exists on whether patterns found in specific studies are replicable. This debate calls for study replication among species, populations and sexes. We measured behavioural (exploration), physiological (breathing rate) and morphological traits (body mass, tarsus length, wing length, bill length) in two species (great vs. blue tits Cyanistes caeruleus), two populations (Forstenrieder Park vs. Starnberg) and two sexes (males vs. females). We then tested whether the same pattern of integration characterized all unique combinations of these three biological categories (hereafter called datasets). We used a multi-year repeated measures set-up to estimate among-individual trait correlation matrices for each dataset. We then used structural equation modelling to test for size-dependent behaviour and physiology, size-corrected (i.e. size-independent) behaviour-physiology correlations and size-corrected body mass-dependent behaviour and physiology. Finally, we used meta-analyses to test which structural paths were generally (vs. conditionally) supported (vs. unsupported). We found general and consistent support for size-dependent physiology and size-corrected body mass-dependent physiology across datasets: faster breathers were smaller but heavier for their size. Unexpectedly, condition-dependent behaviour was not supported: explorative birds were neither leaner, nor was this relationship heterogeneous across datasets. All other hypothesized patterns were dataset-specific: the covariance between size and behaviour, and between behaviour and physiology differed in sign between datasets, and both were, on average, not supported. This heterogeneity was not explained by any of our moderators: species, population or sex. The specific pattern of size- and condition-dependent physiology reported for a unique combination of species, population, and sex, thus generally predicted those in others. Patterns of size- or condition-dependent behaviour (i.e. 'personality'), or behaviour-physiology syndromes reported in specific datasets, by contrast, did not. These findings call for studies revealing the ecological background of this variation and highlight the value of study replication to help understand whether patterns of phenotypic integration reported in one study can be generalized.

Male song stability shows cross-year repeatability but does not affect reproductive success in a wild passerine bird.

Predictable behaviour (or 'behavioural stability') might be favoured in certain ecological contexts, for example when representing a quality signal. Costs associated with producing stable phenotypes imply selection should favour plasticity in stability when beneficial. Repeatable among-individual differences in degree of stability are simultaneously expected if individuals differ in ability to pay these costs, or in how they resolve cost-benefit trade-offs. Bird song represents a prime example, where stability may be costly yet beneficial when stable singing is a quality signal favoured by sexual selection. Assuming energetic costs, ecological variation (e.g. in food availability) should result in both within- and among-individual variation in stability. If song stability represents a quality signal, we expect directional selection favouring stable singers. For a 3-year period, we monitored 12 nest box plots of great tits Parus major during breeding. We recorded male songs during simulated territory intrusions, twice during their mate's laying stage and twice during incubation. Each preceding winter, we manipulated food availability. Assuming that stability is costly, we expected food-supplemented males to sing more stable songs. We also expected males to sing more stable songs early in the breeding season (when paternity is not decided) and stable singers to have increased reproductive success. We found strong support for plasticity in stability for two key song characteristics: minimum frequency and phrase length. Males were plastic because they became more stable over the season, contrary to expectations. Food supplementation did not affect body condition but increased stability in minimum frequency. This treatment effect occurred only in 1 year, implying that food supplementation affected stability only in interaction with (unknown) year-specific ecological factors. We found no support for directional, correlational or fluctuating selection on the stability in minimum frequency (i.e. the song trait whose stability exhibited cross-year repeatability): stable singers did not have higher reproductive success. Our findings imply that stability in minimum frequency is not a fitness quality indicator unless males enjoy fitness benefits via pathways not studied here. Future studies should thus address the mechanisms shaping and maintaining individual repeatability of song stability in the wild.

Heterogeneous selection on exploration behavior within and among West European populations of a passerine bird.

Heterogeneous selection is often proposed as a key mechanism maintaining repeatable behavioral variation ("animal personality") in wild populations. Previous studies largely focused on temporal variation in selection within single populations. The relative importance of spatial versus temporal variation remains unexplored, despite these processes having distinct effects on local adaptation. Using data from >3,500 great tits (Parus major) and 35 nest box plots situated within five West-European populations monitored over 4 to 18 y, we show that selection on exploration behavior varies primarily spatially, across populations, and study plots within populations. Exploration was, simultaneously, selectively neutral in the average population and year. These findings imply that spatial variation in selection may represent a primary mechanism maintaining animal personalities, likely promoting the evolution of local adaptation, phenotype-dependent dispersal, and nonrandom settlement. Selection also varied within populations among years, which may counteract local adaptation. Our study underlines the importance of combining multiple spatiotemporal scales in the study of behavioral adaptation.

Individual variation in age-dependent reproduction: Fast explorers live fast but senesce young?

Adaptive integration of life history and behaviour is expected to result in variation in the pace-of-life. Previous work focused on whether 'risky' phenotypes live fast but die young, but reported conflicting support. We posit that individuals exhibiting risky phenotypes may alternatively invest heavily in early-life reproduction but consequently suffer greater reproductive senescence. We used a 7-year longitudinal dataset with >1,200 breeding records of >800 female great tits assayed annually for exploratory behaviour to test whether within-individual age dependency of reproduction varied with exploratory behaviour. We controlled for biasing effects of selective (dis)appearance and within-individual behavioural plasticity. Slower and faster explorers produced moderate-sized clutches when young; faster explorers subsequently showed an increase in clutch size that diminished with age (with moderate support for declines when old), whereas slower explorers produced moderate-sized clutches throughout their lives. There was some evidence that the same pattern characterized annual fledgling success, if so, unpredictable environmental effects diluted personality-related differences in this downstream reproductive trait. Support for age-related selective appearance was apparent, but only when failing to appreciate within-individual plasticity in reproduction and behaviour. Our study identifies within-individual age-dependent reproduction, and reproductive senescence, as key components of life-history strategies that vary between individuals differing in risky behaviour. Future research should thus incorporate age-dependent reproduction in pace-of-life studies.

Sources of (co)variation in alternative siring routes available to male great tits (Parus major).

Males of socially monogamous species can increase their siring success via within-pair and extra-pair fertilizations. In this study, we focused on the different sources of (co)variation between these siring routes, and asked how each contributes to total siring success. We quantified the fertilization routes to siring success, as well as behaviors that have been hypothesized to affect siring success, over a five-year period for a wild population of great tits Parus major. We considered siring success and its fertilization routes as "interactive phenotypes" arising from phenotypic contributions of both members of the social pair. We show that siring success is strongly affected by the fecundity of the social (female) partner. We also demonstrate that a strong positive correlation between extra-pair fertilization success and paternity loss likely constrains the evolution of these two routes. Moreover, we show that more explorative and aggressive males had less extra-pair fertilizations, whereas more explorative females laid larger clutches. This study thus demonstrates that (co)variation in siring routes is caused by multiple factors not necessarily related to characteristics of males. We thereby highlight the importance of acknowledging the multilevel structure of male fertilization routes when studying the evolution of male mating strategies.