Our knowledge about how the environment influences sexual selection regimes and how ecology and sexual selection interact is still limited. We performed an integrative study of wing pigmentation in calopterygid damselflies, combining phylogenetic comparative analyses, field observations and experiments. We investigated the evolutionary consequences of wing pigmentation for sexual dimorphism, speciation and extinction and addressed the possible thermoregulatory benefits of pigmentation. First, we reconstructed ancestral states of male and female phenotypes and traced the evolutionary change of wing pigmentation. Clear wings are the ancestral state and that pigmentation dimorphism is derived, suggesting that sexual selection results in sexual dimorphism. We further demonstrate that pigmentation elevates speciation and extinction rates. We also document a significant biogeographic association with pigmented species primarily occupying northern temperate regions with cooler climates. Field observations and experiments on two temperate sympatric species suggest a link between pigmentation, thermoregulation and sexual selection, although body temperature is also affected by other phenotypic traits such as body mass, microhabitat selection and thermoregulatory behaviors. Taken together, our results suggest an important role for wing pigmentation in sexual selection in males and in speciation. Wing pigmentation might not increase ecological adaptation and species longevity, and its primary function is in sexual signalling and species recognition.
Data on latitude and wing length of calopterygid damselflies (including Tables S1 and S2)
"WingLengthAndLatitudinalData.xlsx" is an Exel file consisting of three sheets with data used in the comparative analyses of this paper. The data sheet "Wing Pigment" has wing pigment data from males and females (Table S1). The data sheet "Range Info" has range information for the damselfly species used (Table S2). Finally, the data sheet "Body Size" lists the average total body size of the male species in mm. Total body size is measured from the tip of the head to the tip of the abdomen. Body size data were taken from field guides.
File containing information about the phylogenetic tree used in this paper (in NEXUS-format)
"TreeFile" is a nexus format tree file of Calopterygoid damselflies (Odonata: Zygoptera). The tree includes branch labels and lengths, male pigment state, and latitude state. The phylogenetic tree is based on Dumont et al. 2005 Systematic Biology.
Table S3: Additional and complementary BiSSE diversification analyses, excluding different genera
Estimated rate parameters from BiSSE diversification analyses when temperate taxa and the ambiguous tropical genera Hetaerina and Mnesarete were excluded (Table 4; Fig. S1). “Temperate taxa excluded” means that those taxa with a ranges above 23° N were excluded from the tree. “All genera, Ambiguous taxa of Mnesarete clear” means no genera were excluded but taxa in the genus Mnesarete with very small (< 5 cells) wing spots were classified as clear-winged. “Temperate taxa, Hetearina and Mnesarete excluded” means these genera and taxa were deleted from the tree. “Temperate taxa excluded”, Hetearina and Mnesarete as clear-winged” means that temperate taxa were excluded and the genera Hetearina and Mnesarete were classified as clear-winged.
Table S4: Analysis of species-specific thermal reaction norms of Calopteryx splendens and C. virgo
General Linear Model (GLM), involving three factors (Year, Species, Sex) one continuous covariate (Ambient temperature) and all two-way interactions. Data from Klingavälsåns Naturreservat from the summers of 2009 and 2010 (N = 200). The significant Species * Ambient temperature interaction in the table is illustrated in Fig. 6C (see Results). This interaction show that the two species (C. splendens and C. virgo) differ significantly in the slopes of their thermal reaction norms. Full model: r2 = 0.216; F10,189 = 6.481; P < 0.001.
Table S5: Laboratory heating experiment of species-specific thermal reaction norms in Calopteryx splendens and C. virgo
Repeated-measures analysis of laboratory heating experiments of C. splendens (N = 40) and C. virgo (N = 31) of both sexes. The dependent variables (=the repeat) was the thorax temperature at the start of the experiment experiment (“Pre-heating”) and at the end of the experiment, after three minutes under a heat lamp (“Post-heating”). The independent variables were Year (2009 and 2009) Species and Sex, which were factors, and ambient temperature at the start of the experiment was a continuous covariate. The significant Thorax temperature * Species interaction is illustrated in Supporting Figure S3. Pre-heating thorax temperature: r2 = 0.494, F6,64 = 12.402; P < 0.001. Post-heating thorax temperature: r2 = 0.373, F6,64 = 7.926; P < 0.001.
Figure S1: Unresolved phylogenetic tree used in the BiSSE analyses.
Unresolved tree used BiSSE analysis. Large horizontal triangles represent the size and divergence timings of the terminally unresolved clades. Gray triangles are clades with majority clear-winged taxa, black triangles are clades with majority pigmented taxa. Black and white squares on the tips represent binary wing states (transparent = white, pigmented = black), used in the analysis. Red squares mark taxa with ranges north of 23° N.
Figure S2: Phylogenetic trees illustrating the correlation between male wing pigmentation and latitude
Phylogenetic trees showing the correlation of A. male wing pigment and B. latitude. Trees have been pruned of subspecies and had ranges altered so as to create a more representative sampling from those genera. Ranges that have been altered have been marked “Range changed to tropical” and have been filled with a red square. Taxa with wing pigment are filled in with black, transparent wings with white. Temperate ranges are filled in with black, tropical with white. Ancestral reconstruction method is maximum parsimony with branch lengths equal to 1. Gray ancestral nodes represent an ambiguous trait reconstruction for that node.
Figure S3: Heating rates of Calopteryx splendens and C. virgo in a laboratory experiment
Result of from a repeated-measures analysis of a laboratory experiments aimed to investigate species-differences in heating rates of Calopteryx splendens (open symbols, dashed line) and C. virgo (closed symbols, solid line). Least square means ± 95 Confidence Intervals (CI:s) are depicted and have been estimated from the model in Supporting Table S5. Pre-heating and post-heating show the thorax temperatures of both species at the start of the experiment and after three minutes under a heat lamp.
Mass data for field-caught male and female Calopteryx splendens and C. virgo
Data in this file are the mass data (in g) obtained in 2011 and 2012 of field-caught male and female Calopteryx splendens and C. virgo from Klingavälsåns Naturreservat (Scania, Lunds Kommun, Sweden). These data are shown in Fig. 6B in the article.
Field body temperature data obtained from IR-photography for male and female Calopteryx splendens and C. virgo
Data are field body temperatures and ambient temperatures of 200 individual male and female Calopteryx splendens and C. virgo during the summers of 2009 and 2010 at "Klingavälsåns Naturreservat" (Lunds Kommun, Scania, Sweden). These data are shown in Figure 6C in the article, where the species-specific regression lines are illustrated, and the statistical analyses are shown in Figure S4.
Laboratory experiments on heating rate differences of male and female Calopteryx splendens and C. virgo
Data come from a laboratory heating experiment on field-caught male and female Calopteryx splendens and C. virgo during 2009 and 2010, from two field localities: "Klingavälsåns Naturreservat" and "Höje Å, Värpinge", both in southern Sweden. Thorax temperatures were measured using IR-camera prior to heating and after three minutes under a heating lamp. Statistical analyses are provided in Table S5 and the species-specific differences in heating rates are illustrated in Figure S3.
Phenology (date of emergence) for Calopteryx splendens and C. virgo at "Klingavälsåns Naturreservat" in 2002 and 2003
Data are individual emergence records (daily counts) of male and female Calopteryx splendens ("CSMale" and "CSFemale", respectively) and C. virgo ("CVMale" and "CVFemale", respectively) at "Klingavälsåns Naturreservat" (Scania, Lunds Kommun, Sweden) during the summers of 2002 and 2003. These data are shown in the article in Figure 6A.