DOI: 10.5061/dryad.3bk3j9kft
title: The roles of temperature, nest predators and information parasites for geographical variation in egg covering behaviour of tits (Paridae)

description: Aim: Nest building is widespread among animals. Nests may provide
 receptacles for eggs, developing offspring and the parents, and protect
 them from adverse environmental conditions. Nests may also indicate the
 quality of the territory and its owner and can be considered as an
 extended phenotype of its builder(s). Nests may, thus, function as a
 sexual and social signal. Here, we examined ecological and abiotic
 factors—temperature, nest predation and interspecific information
 utilization—shaping geographical variation in a specific nest
 structure—hair and feather cover of eggs— and its function as an extended
 phenotype before incubation in great (Parus major) and blue tits
 (Cyanistes caeruleus) across Europe. We also tested whether egg covering
 is associated with reproductive success of great tits. Location: Fourteen
 different study sites and 28 populations across Europe. Taxon: Parus
 major, Cyanistes caeruleus. Methods: We recorded clutch coverage estimates
 and collected egg covering nest material from the tit nests. We also
 measured nest specific breeding parameters and phenotypic measurements on
 adults. We tested whether mean spring temperatures, nest predation rates
 and flycatcher (Ficedula spp) densities in the study areas explain the
 large-scale geographical variation of clutch coverage and reproductive
 success of tits. Results: The degree of egg coverage of great tits
 increased with lower mean spring temperature, higher nest predation rate
 and higher flycatcher density. We did not find egg covering of blue tits
 to be associated with any of the ecological or abiotic factors. Moreover,
 egg covering of great tits was not associated with reproductive success in
 our cross-sectional data, yet a rigorous assessment of fitness effects
 would require long-term data. Main conclusions: Our findings suggest that,
 in great tits, egg covering may simultaneously provide thermal insulation
 against cold temperatures during egg-laying in spring and also represent a
 counter-adaptation to reduce information parasitism by flycatchers and
 nest predation. Hence, geographical variation in interspecific
 interactions, and consequently in co-evolutionary processes, may affect
 the evolution of nest characteristics besides environmental conditions.
description: <strong>Study areas.</strong> The great tit data for
 this study were collected in spring 2013 from 10 different countries, 14
 study areas (Figure 1, Table S1) and 28 populations in Europe. The blue
 tit data were collected in the same year from 6 of the 10 countries, 8 of
 the 14 study areas and 22 of the 28 populations. All study populations
 breed in nest boxes. Research was carried out in accordance with
 legislation of each country. <strong>Field
 procedure.</strong> Nest building state and the beginning of the
 egg-laying (laydate) were checked during regular field observations. Use
 of egg cover tends to increase during the first day of the laying stage
 (Haftorn & Slagsvold, 1995). During the egg-laying stage, when
 tits had laid their fourth to eighth egg, (a) the nest was photographed to
 get a measurement of the extent of the clutch coverage, that is, the
 proportion of the visible clutch surface (%) and (b) all the hair and
 other material that covered tit eggs and nest cup was removed to expose
 the eggs and placed in a zip lock bag for later measurement of hair mass
 and the nest was photographed again. After photographing the nest, the
 removed material was replaced by same quantity of sheep hair. The onset of
 incubation was determined by observing the presence of female on the nest
 and touching eggs to determine whether the eggs were cold or warm. Nest
 specific breeding parameters (number of hatched eggs and fledglings) and
 phenotypic measurements on adults (Table 1) were also collected. We
 recorded clutch coverage estimates and mean spring temperatures (from the
 nearest available meteorological stations to each of the study area) from
 476 great tit nests and 123 bluetit nests and nest predation rates from
 345 great tit nests and 74 blue tit nests. Flycatcher (either Ficedula
 hypoleuca or Ficedula albicollis) density was measured in the end of
 breeding season as the proportion of nest boxes occupied by flycatchers in
 the study population. <strong>Measurement of nest
 characteristics</strong>. The clutch coverage rate was measured by
 comparing the proportions of the visible clutch surfaces from the digital
 photographs taken from the nest before and after cover removals using
 ImageJ software (US National Institutes of Health,
 http://imagej.nih.gov/ij). The clutch surface was measured using freehand
 tracing and area calculator tools. Clutch surfaces were measured twice
 from each picture to minimize measurement error and average values were
 used in the analyses. Masses of the collected hair samples were weighed to
 the nearest 0.0001 g by using an Ohaus AS120S analytical balance.
 Phenotypic measurements on adult tits were obtained when they were
 captured during food provisioning. Age was classified in the field as
 1-year-old (second calendar year) or older (at least third calendar year)
 (Jenni & Winkler, 1994). Adult and young birds were handled under
 the ringing licenses of the authors. Hence, our study complied with the
 national legislation of Belgium, Czech Republic, Denmark, Estonia,
 Finland, Hungary, Italy, Spain, Sweden and Switzerland concerning handling
 wild animals. Variables used in statistical analyses are listed in Table
 1. <strong>Statistical methods.</strong>
 The distribution of clutch covering rate (proportion of covered eggs) was
 slightly U-shaped with a high peak at one (all eggs covered), which is
 problematic for analysis. Therefore, we measured clutch coverage by
 combining clutch covering rate and the mass of material used to cover the
 eggs, because these variables together measure the investment of the tit
 parents in covering their clutch. For this purpose, we ran principal
 component analysis for the data on egg coverage and the mass of the
 covering material and used the first principal component (‘clutch
 coverage’ hereafter, explains 72.3% of the variance, eigenvalue = 1.0) as
 a response variable when analysing variation in clutch covering behaviour.
 Clutch coverage variable was symmetrically (approximately normally)
 distributed, and positively correlated with both clutch covering rate and
 mass of the covering material, higher values, thus, indicating higher
 investment in clutch covering (Figure S1). All statistical analyses were
 conducted with R version 3.4.3 (R Core Team, 2017). Linear mixed-effects
 models (LMMs; function lme in package ‘nlme’ (Pinheiro et al., 2017)) were
 used to analyse variation in the clutch covering of great (Model sets 1–4)
 and blue tits (Model set 5). Generalized linear mixed-effects models
 (function glmer in package ‘lme4’; Bates, Mächler, Bolker, &
 Walker, 2014) with Poisson distribution and a logarithmic link function
 were used to analyse variation in the number of hatched eggs and
 fledglings of the great tits (Model sets 6–9). In model sets 6–9, we
 standardized all continuous explanatory variables of the model.
 Standardization makes the quantitative interpretation of model parameters
 less intuitive, which is the reason why standardization was used only when
 it was really needed for aiding/facilitating model convergence. We used
 multi-model inference; effects of analysed variables were summarized by
 model averaging (Burnham & Anderson, 2002) (function model.avg in
 package ‘MuMIn’; Barton, 2009). We derived 10 model sets. Model set 1
 (Table 2a) tested if the alternative hypotheses (i.e. insulation, nest
 predation, information parasitism), nest floor surface area or forest type
 (dominant tree genus) in the study site explain variation in clutch
 coverage in great tits. This model set was fitted to data (Nobservations =
 341) including observations from all the study sites, also including sites
 where the flycatcher density was zero. Mean spring temperature, nest
 predation rate, flycatcher density, nest floor surface area, first
 principal component of geographical variables (altitude, latitude and
 longitude) and dominant tree genus in the study site were set as fixed
 effects, and population as a random effect in the global model. Time of
 the year (Laydate) was not included in the analysis because it is strongly
 negatively correlated with the first principal component of geographical
 variables (Pearson's correlation, r = −.61, t = −3.41, df = 343, p
 &lt; .001) (see Laydate in Table S3). Nest predation rate positively
 correlates with mean spring temperature (r = .67) but both of these
 variables were retained in all models because of their importance for
 assessing the study hypotheses. No interactions were included in any
 model. The set of all meaningful models simpler than the global model was
 derived with the function ‘dredge’ (package ‘MuMIn’; Bartón &
 Barton, 2017) for model averaging, the global model being included in
 model averaging (see Table S2a for the set of averaged models). Bivariate
 correlations between the study variables are provided in Table S3. Model
 set 2 (Table 2b) was derived otherwise similarly to model set 1, but all
 possible interactions among the nest predation rate, mean spring
 temperature and flycatcher density were included in the global model (see
 Table S2b for the set of averaged models). Model set 3 (Table 3a) was
 derived otherwise similarly to model set 1, but these models were fitted
 to data including only areas where the flycatchers were present
 (flycatcher density &gt; 0, Nobservations = 169, see Table S4a for the
 set of averaged models). This was done to reliably estimate the effect of
 flycatcher density. In the full data the high number of zeros (flycatchers
 not present) might confound the estimation (underestimation) of flycatcher
 density effect on great tit egg covering behaviour. Model set 4 (Table 3b)
 was derived otherwise similarly to model set 3, but all two-way
 interactions among the nest predation rate, mean spring temperature and
 flycatcher density were included in the global model (the three-way
 interaction was ignored as the model-fitting failed when it was included
 in the global model; see Table S4b for the set of averaged models). Model
 set 5 (Nobservations = 74; Table S5) was otherwise similar to model set 1,
 but it focused on variation in clutch coverage in blue tits (see Table S6
 for the set of averaged models). Because of the low number of blue tit
 observations, we did not conduct any further analyses for this species.
 Model sets 6 and 7 tested whether the clutch coverage of great tits or the
 ecological and abiotic environment explain the number of hatched eggs of
 great tits by using all great tit data. In model set 6 (Table S7), number
 of hatched eggs was used as a dependent variable and clutch size was added
 as a covariate in the global model in both model sets to take the effect
 of clutch size variation into account. In addition to clutch size, fixed
 effects of the global models included also clutch coverage, mean spring
 temperature, nest predation rate, flycatcher density, as well as female
 age and its interaction with clutch size, because female age affects
 clutch size (Perrins & Mccleery, 1985). Population was set as a
 random effect. Except clutch size (a non-negative integer), continuous
 variables were standardized (by subtracting mean from each observation and
 dividing this difference by standard deviation, see Table S8 for the set
 of averaged models) to aid model convergence. Model set 7 (Table S9) was
 derived otherwise similarly to model set 6, but all possible interactions
 among the nest predation rate, mean spring temperature and flycatcher
 density were included in the global model (see Table S10 for the set of
 averaged models). Model sets 8 (Table S11; see Table S12, for the set of
 averaged models) and 9 (Table S13; see Table S14, for the set of averaged
 models)) tested whether the clutch coverage of tits or the ecological and
 abiotic environment explain the number of fledged offspring of great tits
 and were derived otherwise similarly to model sets 6 and 7, respectively,
 but number of fledged offspring was used as a dependent variable.
 Population was set as a random effect in all models. Model set 10 (Table
 S15; see Table S16, for the set of averaged models) was otherwise similar
 to model set 1, but flycatcher density was replaced with flycatcher
 presence (binary variable; flycatcher present or not) for checking whether
 the results are sensitive to the way how flycatcher density is handled
 (continuous vs. presence/ absence).