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Genetic colour variation visible for predators and conspecifics is concealed from humans in a polymorphic moth



The definition of colour polymorphism is intuitive: genetic variants express discretely coloured phenotypes. This classification is, however, elusive as humans form subjective categories or ignore differences that cannot be seen by human eyes. We demonstrate an example of a 'cryptic morph' in a polymorphic wood tiger moth (Arctia plantaginis), a phenomenon that may be common among well-studied species. We used pedigree data from nearly 20,000 individuals to infer the inheritance of hindwing colouration. The evidence supports a single Mendelian locus with two alleles in males: WW and Wy produce the white and yy the yellow hindwing colour. The inheritance could not be resolved in females as their hindwing colour varies continuously with no clear link with male genotypes. Next, we investigated if the male genotype can be predicted from their phenotype by machine learning algorithms and by human observers. Linear discriminant analysis grouped male genotypes with 97% accuracy, whereas humans could only group the yy genotype. Using vision modelling, we also tested whether the genotypes have differential discriminability to humans, moth conspecifics and their bird predators. The human perception was poor separating the genotypes, but avian and moth vision models with ultraviolet sensitivity could separate white WW and Wy males. We emphasize the importance of objective methodology when studying colour polymorphism. Our findings indicate that by-eye categorization methods may be problematic, because humans fail to see differences that can be visible for relevant receivers. Ultimately, receivers equipped with different perception than ours may impose selection to morphs hidden from human sight.
J Evol Biol. 2022;35:467–478.
Received: 24 June 2021 
Revised: 31 Januar y 2022 
Accepted: 6 February 2022
DOI : 10.1111/j eb.13994
Genetic colour variation visible for predators and conspecifics
is concealed from humans in a polymorphic moth
Ossi Nokelainen1,2 | Juan A. Galarza1,2| Jimi Kirvesoja1| Kaisa Suisto1|
Johanna Mappes1,2
This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium,
provided the original work is properly cited.
© 2022 The Authors. Journal of Evoluti onary Biolog y published by John Wiley & Sons Ltd on behalf of European Society for Evolutionar y Biology
Ossi Noke lainen and Juan A. Gal arza author s are contributed equally to thi s work.
1Department of Biological and
Environmental Science, University of
Jyväskylä, Jyväskylä, Finland
2Organismal and Evolutionary Biology
Research Program, Faculty of Biological
and Environmental Sciences, University
of Helsinki, Helsinki Universit y, Helsinki,
Ossi Nokelainen, Depar tment of Biological
and Environmental Science, Universit y of
Jyväskylä, P.O. Box 35, Jyväskylä 4 0014,
Funding information
This work was supported by the Academy
of Finland to JM (#320438) and the grant
(#21000 038821) to ON.
The definition of colour polymorphism is intuitive: genetic variants express discretely
coloured phenotypes. This classification is, however, elusive as humans form subjective
categories or ignore differences that cannot be seen by human eyes. We demonstrate
an example of a ‘cryptic morph’ in a polymorphic wood tiger moth (Arctia plantaginis),
a phenomenon that may be common among well- studied species. We used pedigree
data from nearly 20,000 individuals to infer the inheritance of hindwing colouration.
The evidence supports a single Mendelian locus with two alleles in males: WW and
Wy produce the white and yy the yellow hindwing colour. The inheritance could not
be resolved in females as their hindwing colour varies continuously with no clear link
with male genotypes. Next, we investigated if the male genotype can be predicted
from their phenotype by machine learning algorithms and by human observers. Linear
discriminant analysis grouped male genotypes with 97% accuracy, whereas humans
could only group the yy genotype. Using vision modelling, we also tested whether the
genotypes have differential discriminability to humans, moth conspecifics and their
bird predators. The human perception was poor separating the genotypes, but avian
and moth vision models with ultraviolet sensitivity could separate white WW and
Wy males. We emphasize the importance of objective methodology when studying
colour polymorphism. Our findings indicate that by- eye categorization methods may
be problematic, because humans fail to see differences that can be visible for relevant
receivers. Ultimately, receivers equipped with different perception than ours may im-
pose selection to morphs hidden from human sight.
aposematism, Arctia plantaginis, discriminant analysis, multispectral imaging, polymorphism,
wood tiger moth
Colour polymorphism, the occurrence of multiple discrete colour phe-
notypes within a population (Ford, 1945; Huxley, 1955; White & Kemp,
2016), is a flagship topic of evolutionary biology (Gray & McKinnon,
2007; McKinnon & Pierotti, 2010; Svensson, 2017). The study of co-
lour polymorphism has traditionally been a very popular topic among
evolutionary biologists (Brakefield & Liebert, 1985; Cain & Sheppard,
1954; Fisher & Ford, 1947; Kettlewell, 1955), because as a visible
trait, colouration enables scientists to study evolution in action in a
tractable manner. Importantly, colouration is a composite trait that
has multiple fitness- linked functions (Cuthill et al., 2017), including
thermoregulation (e.g. Stuart- Fox et al., 2017), immune defence (e.g.
Freitak et al., 2005), sexual signalling (e.g. Tibbetts et al., 2017) and
avoiding predation either through camouflage, mimicry or warning sig-
nalling (e.g. Ruxton et al., 2019). The diversity of colours, as well as co-
lour polymorphism, is therefore valuable to understand the processes
generating and maintaining genetic variation in the wild.
Classically, a genetic polymorphism is defined as: ‘the occurrence
together of two or more discontinuous forms of a species in the
same habitat in such proportions that the rarest of them cannot be
maintained merely by recurrent mutation’ (Ford, 1945, 1965; Huxley,
1955). This definition has remained virtually unchanged over the last
75 years (Nokelainen et al., 2018; Svensson, 2017; White & Kemp,
2016). While the concept of colour polymorphism may be rather
intuitive, its quantification is not, mainly because the difference
between colour variants is not always clear- cut. For example, phe-
notypic plasticity can provide discrete appearances as if they were a
result of polymorphism (Price, 2006), such as the density- dependent
colour change (i.e. polyphenism) in a desert locust (Schistocerca
gregaria) (Sword et al., 2000). On the other hand, sometimes a ge-
netically polymorphic trait may show overlapping phenotypic dis-
tribution (Kappers et al., 2018; Nokelainen et al., 2018), such as the
extravagant colour polymorphism of the Hawaiian happy- face spi-
der (Theridion grallator) that shows high phenotypic variation among
populations (Gillespie & Oxford, 2009).
In Lepidoptera, one of the potential caveats of early polymor-
phism studies is that colouration was mostly quantified through
human vision and thus included a source of subjectivity (Brakefield
& Liebert, 1985; Endler, 1990; Fisher & Ford, 1947). This may not
always be a problem as humans are good in categorizing colours
across a broad visible wavelength spectrum from 400 to 70 0 nm
(Bergeron & Fuller, 2017) and have excellent visual acuity (Caves
et al., 2018). However, our perception excludes the near ultravio-
let part of the spectrum (300– 400 nm), which is important to many
animals in signalling (Kelber et al., 2003; Osorio & Vorobyev, 2005).
Also, humans may not be able to detect nuances in patterns or judge
colour polymorphism based only on a single key trait (e.g. hindwing
‘base’ colour), or see subjective categories where they do not exist.
Colouration must therefore be objectively quantified using either
spectrometry or multispectral imaging approaches (Endler, 1990,
Troscianko & Stevens, 2015, van den Berg et al., 2020). As such
there is a clear need for studies that can link the phenotypes to their
genotypes, because this can illuminate our understanding of how
selection of allele frequencies that constitute the genotypes operate
in the wil d (C uthill et al., 2017; Svens son, 2017; Tib bet ts et al., 2017).
We investigated genotype– phenotype associations in the wood
tiger moth (Arctia plantaginis), a widely distributed member of the
Erebidae family (Rönkä et al., 2016) found across the Northern
hemisphere (Hegna et al., 2015). It is known that its polymorphic
hindwing colour is heritable in males (Nokelainen et al., 2013) and
in females (Lindstedt et al., 2016). In general, males have either yel-
low or white hindwings (Nokelainen et al., 2012, 2013; Suomalainen,
1938). Female hindwings, on the other hand, vary continuously in
the yellow- orange- red range (Lindstedt et al., 2011).
First, we explored the heritability of the hindwing colouration. It
has been suggested, although with a very limited data set originating
from a single brood, that the inheritance of male hindwing coloura-
tion follows a Mendelian one- locus two- allele model where the yel-
low allele is recessive (Suomalainen, 1938). The genetic mechanism
of female wing colouration is largely unknown and some plasticity
in female colouration has been reported (Lindstedt et al., 2010). To
confirm the mode of inheritance, we compared human- visible hind-
wing colour (and only hindwing colour as genotype proxy) frequen-
cies from 452 laboratory- reared families (i.e. with pedigree) against
those predicted by the one- locus two- allele model. Second, focussing
in the males, we tested further whether their genotype can be pre-
dicted by their phenotype. We examined if the colour morphs could
be assigned to their genotype (i.e. an information derived from the
pedigree, Box 1) by human observers through sequential and simul-
taneous sorting tasks, as well as by machine learning algorithms (i.e.
discriminant functions). We used linear discriminant function analysis
as part of the machine learning realm, as we wanted to explicitly un-
derstand what are the parameters that may allow visual separation
of the genotypes. It can be expected that the computational meth-
ods should outperform human sorting skills, because the algorithms
can take into account combined nuances in phenotypic variation, in-
cluding those beyond the human- visible spectrum (Høye et al., 2021;
Wilkins & Osorio, 2019). Lastly, we asked whether these genotype–
phenotype associations may have ecological relevance beyond the
human- visible spectrum. Using vision modelling, Henze et al. (2018)
investigated the differences in the discriminability of the wood tiger
moth colour morphs by moth conspecifics and bird predators. Here,
we used receptor- noise- limited vision modelling (Maia et al., 2013;
Vorobyev & Osorio, 1998) to test pairwise genotype chromatic con-
trasts of hindwing colour using human, avian and moth vision models.
2.1  |  Moth pedigree rearing protocol
Altogether we used pedigree data from 15 generations of the wood
tiger moth reared in the laboratory over the course of 6- years. As a
gene ral rearin g proto col, two adults (male and female) are put tog eth er
in a plastic box (Huhtamäki, 1000 ml, transparent casing) with mesh
for ventilation at the top and allowed to mate under natural lighting.
The eggs are laid inside the box where after ~6 days the larvae hatch
and are kept for another ~14 days inside the box as they are too deli-
cate to be moved. The larvae are then separated into rearing contain-
ers (max. 30 lar vae/container) to continue growth at approximately
25°C under natural light conditions and are fed with fresh dandelion
leaves (Tara xa cum ssp.) until pupation. The pupae are then moved
into individual jars and sex and wing colour are recorded from the
emerging adults. In males, colour classification is conventionally done
by- eye (white or yellow hindwing ‘base colour’) and in females, a six-
step (yellow- orange- red) scale is used as described by Lindstedt et al.
(2010) and Nokelainen et al. (2012): in 1 to 6 scale yellow- orange- red
gradient yellows are 1– 2, oranges 3– 4 and reds 5– 6 (Figure 1).
2.2  |  Inheritance of hindwing colour based
on the pedigree
We specifically tested a Mendelian inheritance model where a
single locus with two- alleles controls the white- yellow polymor-
phism. Here, the yellow allele (y) is recessive to the dominant
white (W) allele, as suggested by Suomalainen (1938) who used
a single brood of individuals originating from Finland. Under this
model, genotypes with the W allele (i.e. WW, Wy) exhibit white
hindwings, whereas only the homozygote recessive genotype (i.e.
yy) exhibit yellow hindwings. We tested the one- locus two- allele
model (Box 1), by comparing the expected model's frequencies
to those obser ved from 452 families with pedigree data (each
offspring was considered as independent data point) using a Chi
Square test for independence. The expected frequencies under
this model with their resulting hindwing colour are presented in
Tables S1 and S2. Only families with at least 10 male and female
offspring were used to ensure reliable frequency distribution.
From the pedigree data, the parental genotypes were inferred
from the phenotype distribution of the F1 offspring. For instance,
a 100% yellow mal e off spring wo uld ind icate that both parents are
homozygous for the y allele (Box 1). The reasoning uses the same
approach as myriad classic studies of ecological genetics decipher-
ing Mendelian ratios. The key point with this pedigree back- trace
approach is that by producing consecutive generations it is pos-
sible to match the observed F1 phenotype frequencies to those
expected by Mendelian inheritance, and thus, the genotype of the
parental generation can be inferred.
2.3  |  Objective genotype– phenotype associations
using image analysis techniques
The pedigree back- trace approach above was used to unravel the
inheritance of hindwing colouration using hindwing colouration as
BOX 1  Pedigree crossing design and determination of the wood tiger moth genotypes with respect to hind wing
colour. The first panel shows the classic Mendelian one locus two allele segregation (A); we expect that white is
dominant trait over yellow (Suomalainen, 1938). Each homozygous parent in the parental generation produces one
type of gamete (W or y). The following generation heterozygous offspring produces again two types of gametes. In
the second panel (B), the next generation produces offspring with a 3:1 ratio of dominant allele to recessive. The
third panel, shows the crossing design followed to mate selection lines of known genotypes and their expected
phenotype frequencies. Fifteen generations were produced over the course of 6- years. The colours in the bars
indicate the hind wing colour of the offspring. An important point with this classic approach is that by producing
consecutive generations and following the logic of expected offspring phenotype frequencies, it is possible to
back- trace pedigree and determine putative genotypes of earlier generations.
(a) (b) (c)
perceived by humans. To investigate genotype– phenotype associa-
tions further than simply using a human- visible hindwing colour, we
used a subset of laboratory- reared adults with a known pedigree in
our image analyses. The total sample size of the photographed in-
dividuals was 292: where of the males 37 were WW, 88 were Wy,
and 42 were yy, while of the females, 33 were WW, 68 were Wy
and 24 were yy. The image calibration and analysis broadly followed
previously established methods (Troscianko & Stevens, 2015, van
den Berg et al., 2020). Briefly, photography was undertaken with a
Samsung NX1000 digital camera converted to full spectrum with no
quartz filter to enable UV sensitivity fitted with a Nikon EL 80 mm
lens. For the photos in the human- visible range, we used a UV and
infrared (IR) blocking filter on the lens, which passes wavelengths
only between 400 and 680 nm (Baader UV/IR Cut Filter). For the UV
images, a UV pass filter was used (Baader U filter), which transmits
wavelengths between 320 and 380 nm. Grey reflectance standards,
which reflect light equally at 7% and 93% between 300 and 750 nm,
were used for image calibration. A standard light source 75W Exo-
terra Sunray (mimicking sunlight across the spectrum) was used.
To obtain colour and pattern metrics, we measured the entire
dorsal view of the forewings (FW), hindwings (HW), thorax (TH)
and abdomen (AB) of the mounted and spread adult as regions of
interest (ROI). For reflectance data, we used normalized camera re-
sponses of red, green, blue and the UV channel. To extract pattern
information, we applied a pattern analysis technique (a ‘granularity
analysis), which decomposed the image into a series of spatial fre-
quencies (‘granularity bands’) using Fourier analysis and band pass
filtering, followed by determining the relative contribution of differ-
ent marking sizes to the overall pattern (Barbosa et al., 2008; Hanlon
et al., 2009; Stoddard & Stevens, 2010). The analysis calculated the
amount of light information (or pixel energy) corresponding to mark-
ings of different sizes, starting with small markings (we used a pixel
start size of 2) and increased in size to larger markings (we used a
pixel end size of 100). Increase in pixel step size was set to multiply
each step by 1.414, thus representing exponential growth. The lu-
minance was measured over 20 bands from lowest luminance (0) to
highest luminance (65535), the maximum dynamic range of a 32- bit
TIFF image. The luminance channel was set to longwave channel (R).
For the pattern data variables, we used dominance (i.e. maxPower—
the energy at the spatial frequency with the highest pixel energy),
diversity (i.e. propPower— maximum or peak energy value divided by
the summed energy) and marking size (i.e. maxFreq— the spatial fre-
quency with peak energy).
Prior to testing, colour metrics were filtered for correlations to
avoid multicollinearity. The following variables were retained: area,
three pattern variables (pattern size, contrast and diversity) and four
bandpass channels (UV, blue, green, red channels, i.e. uv, sw, mw, lw
respectively). All values were separately measured for the four re-
gions of interest (forewing, hindwing, thorax and abdomen). In addi-
tion, the following allometric measurements of size were calculated
by dividing areas of the ROIs: forewing to abdomen (FW/AB), fore-
wing to thorax (FW/TH), forewing to hindwing (FW/AB) and thorax
to abdomen (TH/AB).
2.4  |  Discreteness of colour morphs— subjective
genotype discrimination using human observers
Next, we evaluated human sorting accuracy of male genotypes
through sequential (‘sequence’) and simultaneous (‘sorting’)
FIGURE 1 Pedigree information of the wood tiger moth
genotype crossings and their human- visible hindwing coloration.
The figure shows relative frequencies of offspring phenotypes
with respect to their parental genotype crosses. The numbers
above the bars indicate the sample size. The colours in the bars
indicate the subjective by- eye hindwing colour of the offspring.
Notice the dichotomous yellow- white hindwing categorization
in males (a), whereas the females are more converged to similar
orange coloration (b); the scale depicts the visually scaled yellow-
orange- red colour gradient used to categorize female coloration.
The images were gamma corrected for better screen imaging
and are meant to illustrate representative examples of the wood
tiger moth colour variation. In males, hindwing colour shows
statistically nonsignificant difference from predicted one locus two
allele model, whereas females deviate significantly from the same
predicted outcome of phenotypes
tasks. Participants were familiar with the wood tiger moth. In
both tasks, we showed participants 10 images per genotype (i.e.
3 genotypes by 10 replicates) and asked them to sort the images
according to their genotype. As the mechanism controlling for
the hindwing colouration in females is currently unknown and
warrants further investigation, we used only males in the sorting
tasks due to their discrete hindwing colouration that allows for
tractable inheritance.
In the sequential sor ting task, 12 par tici pa nt s we re asked to clas-
sify moth photographs by their genotype. The photographs were
of a mounted specimen with wings spread out. The photographs
were shown in a randomized order via the Google Docs ‘Forms’ plat-
form. The participants were asked to pay attention to the appear-
ance, wing and body colouration. The following cues derived from
the image analysis (see above) to classify the male genotypes (WW,
Wy, yy) were given as training instructions. White hindwings, large
forewing patterning and pale abdomen are typical to WW. White
hindwings and a yellow tinge in forewings and abdomen are typical
to Wy. Yellow hindwings, variable wing patterning, yellow abdominal
colour is typical to yy. The participants were instructed to classify
moths using these cues (Fig. S3).
In the simultaneous sorting task, 10 participants (a subset of the
former group) were asked to sort the genotypes into three clusters
based on similarities in their appearance; no further instructions
were given to accomplish this task. All moths were visible at the
same time and the test was done using PowerPoint slides with moth
photographs (i.e. 30 images were simultaneously presented, 3 geno-
types by 10 replicates). The percentage of correct answers was then
calculated (Fig. S4).
2.5  |  Vision modelling
The vision modelling we carried out largely followed established
methods (Stevens et al., 2007, Troscianko & Stevens, 2015, van
den Berg et al., 2020). To gain insight into how well different vi-
sion systems can recognize the colour differences between geno-
types, we used a receptor- noise limited (RNL) visual discrimination
model (Vorobyev et al., 1998). We compared trichromatic human,
tetrachromatic avian (Blue tit; Cyanistes caeruleus) and trichromatic
moth (wood tiger moth) vision models. This allowed us to mecha-
nistically understand human sorting accuracy of genotypes and to
compare this with more ecologically relevant vision systems of con-
specifics (moths) and predators (birds). We used 0.05 Weber fraction
for most abundant cone type for all vision models. The cone ratios
were: avian cone ratios 1:1.92:2.68:2.7 uv:sw:mw:lw (Hart, 2001b),
human cone ratios 0.057:0.314:0.629 sw:mw:lw (Hofer et al., 2005).
For moth vision model, spectral sensitivities of cone cells (uv, sw,
mw) were obtained from (Henze et al., 2018), and cone ratios 1:1:1,
were used as the specific ratio is unknown. As we were interested
in differences in chromatic contrast (dS), we excluded the achro-
matic contrast (dL) from the vision model analysis. The vision model
yields discrimination values in ‘just noticeable differences’ (JNDs), al-
though before behavioural validation these should be considered as
predicted contrast values (dS). By definition, values lower than one
(<1 JND) are considered indistinguishable, whereas larger values are
discriminable for the receiver (Kang et al., 2015; Nokelainen et al.,
2019; Siddiqi et al., 2004).
2.6  |  Statistical analyses
First, we tested whether the wood tiger moth hindwing colour
follows a simple Mendelian one- locus two- allele inheritance pat-
tern using Chi Square test for independence. We would expect
that the observed phenotype frequencies from the crossing de-
signs do not deviate significantly from the predicted phenotype
frequencies. We tested the expected versus observed subjective
colour morph frequencies separately for males (Table S1) and fe-
males (Table S2).
Second, we tested the discriminability of genotype– phenotype
associations. The success rate of cor rect genotype designation was
tested with a general linear model (GLM with a Poisson distribu-
tion), where the success rate of visually genotyping each moth was
set as the dependent variable and method of genotyping (human
sequential task, human simultaneous task, or computer algorithm)
as the explanatory variable. We used a linear discriminant function
analysis to evaluate whether the computer algorithm can outper-
form human observers in the genotype sorting task. The analysis
was carried out as a 3- group problem. The genotype (derived from
the pedigree data) was set as the predicted group membership
and regions of interest (ROI) were selected form the digital image
namely; area, pattern size, pattern contrast, pattern diversity, uv,
sw, mw, lw, forewing to abdomen, forewing to thorax, forewing to
hindwing and thorax to abdomen were set as predictor variables.
All colour and pattern metrics were investigated separately for the
forewing, hindwing, thorax and abdomen. We also used the Boruta
feature selection algorithm to provide additional information on
which feature s are th e bes t predi c tor s of th e prio r gen otyp e gro ups
(under R- package ‘Boruta’). Briefly, Boruta is a random forest al-
gorithm that compares the significance of each variable against
random noise data created from all variables of interest (Kursa &
Rudnicki, 2010). The significance is then determined based on the
relative difference against the random noise. Generally, variables,
which fall in between lower- and upper- bound significance of the
random generated noise reference are flagged as non- significant
(Table S3, Figs. S1– S2).
Third, we tested the discriminability of the genotypes using
three different vision models. For this, we conducted a linear
mixed effects model (lmer- function) with the lmerTest R- package
(Kuznetsova et al., 2017). The colour contrast (dS) was set as the
dependent variable and genotype (WW, Wy, yy), ROI (abdomen,
forewing, hindwing), vision model (avian, human, moth) and their in-
teractions were set as the explanatory variables. The moth ID was
set as a random variable to control for data structure. All analyses
were conducted using RStudio, version 1.1.447 and R, version 3.5.0
(R Core Team, 2018; RStudio Team, 2016).
3.1  |  Mendelian inheritance of the hindwing colour
Of the individuals used in the pedigree analysis 10911 were males
and 8295 were females (Figure 1, Tables S1 and S2). In the males, the
frequencies of white and yellow offspring were in close agreement
to the expected phenotype frequencies under one- locus two- allele
Mendelian inheritance where white dominates over yellow (Box 1,
Figure 1a). Thus, the one- locus two- allele inheritance mode with
dominance of the W allele over y was confirmed by the pedigree
data for males. Whether the established locus also controls hindwing
colour in the females was less clear. In females, the spread of the
hindwing colour phenotypes showed an apparent normal distribu-
tion (Figure 1b) and indicated phenotypic tendency towards orange
hindwing colouration (by- eye classification; yellow- orange- red). Out
of all crossings, the emerged females on average, were 3% yellow,
68% orange and 29% red regardless of the parental genotype. Thus,
there was no obvious correlation between the male (white- yellow)
and female (yellow- orange- red) hindwing colour within the brood
(Figure 1a- b) when using the subjective hindwing colour classifica-
tion made by human observers.
3.2  |  Discriminability of genotype– phenotype
associations— human versus algorithm
A computer- based discrimination algorithm outperformed subjec-
tive sorting accuracy of humans using linear combinations of col-
our and pattern data (ANOVA: F2,48 = 7.10 , p < 0.001). In males, the
linear discriminant analysis reached 96.80% accuracy for predicting
the correct genotype membership. Within test data, 88.88% of WW,
98.11% of Wy and 100% of yy were correctly classified (Figure 2a).
The discriminability of genotypes using colour metrics was also as-
sessed using the Boruta feature extraction algorithm (Table S3). The
most important variables to separate white and yellow morphs are
all hindwing features and include: UV reflectance, short wavelength
reflectance, pattern diversity, pattern contrast and long wavelength
reflectance. The most important variables to separate white geno-
types (WW, Wy) are: thorax UV reflectance, thorax to abdomen size
ratio, abdomen UV, forewing to abdomen size ratio and forewing
pattern diversity.
In females, the linear discriminant analysis reached 65.65% ac-
curacy for predicting the correct genotype membership. Within test
data, 83.33% of WW, 58.62% of Wy and 69.56% of yy were cor-
rectly classified (Figure 2b). The most important variables to make
a distinction between white homozygotes (WW) and yellow allele
bearers (Wy or y y) are: thorax UV reflectance, abdomen marking
size, abdomen UV reflectance, forewing UV reflectance, hindwing
short wavelength reflectance (Table S3). The most important vari-
ables to separate white heterozygotes (Wy) from yellow homozy-
gotes (y y) are: abdomen marking size, hindwing short wavelengths,
FIGURE 2 Objective phenotype quantification using colour and pattern metrics. Linear discriminant analysis for wood tiger moth male
(a) and female (b) genotypes. Notice that here genotypes refer to one locus two allele model, where there are two predominantly white
morphs and one yellow in males and red, orange, yellow morphs in females. In males, the yy (yellow) genotype is clustered easily as its own
subgroup, but linear combinations also separates WW (white) and Wy (white) genotypes with good degree of certainty along a second axis
that combines parts of the full spectrum (invisible to us) as well as pattern metrics. In females, the three genotypes seemingly cluster into
three subgroups; however, there is much more grouping overlap (i.e. phenotypic convergence) than in males
hindwing medium wavelengths, forewing short wavelengths and
hindwing long wavelengths.
We next focussed on discriminability of genotypes for human
observers only using males as we detected a close phenotypic
similarity in females. Human participants were not able to reliably
categorize the genotypes (Figure 3, Fig. S5). Of the male moths, par-
ticipants were only able to distinguish yellow (yy) genotype from the
whites, but not the two white male genotypes (W W, Wy). In the si-
multaneous sorting task where all moths were presented together,
participants were able to sort the homozygous and heterozygous
males only slightly better than in the sequential sorting task, yet only
60% was the highest success rate in sorting white males based on
3.3  |  Chromatic discriminability of the genotypes
to ecologically relevant receivers
The discriminability of the genotypes was measured pairwise using
hindwing chromatic contrast (dS) of the two moths being compared
(Figure 4). The vision modelling results indicate that detectability of
the genotypes was different for human, bird and moth vision mod-
els, as the three- way interaction of vision model, ROI and genotype
was significant (lmer ANOVA, F4,23 028.1 = 506.11, p < 0.001). Also,
vision modelling results suggest that human perception is poor at
separating WW and Wy male genotypes, whereas avian and moth
vision systems with UV sensitivity could separate white WW and Wy
male genotypes (Figure 4). Thus, when viewed through ecologically
FIGURE 3 The success of objective
versus subjective sorting of male
genotypes. Colour metric super vised
computer algorithm (LDA, linear
discriminant analysis) outperforms human
sorting accuracy of genotypes in both
sequential (‘sequential’) and simultaneous
(‘sorting’) tasks. In sequential sorting
people were asked to classify moth
pictures by their genotypes; only basic
information of the best describing colour
metrics were given as instructions. In
simultaneous sorting, people were asked
to sort the genotypes into three clusters
based on their superficial appearance;
no further instructions were given, but
all moths were visible at the same time.
Percentage indicates the number of
correct answers by genotype
relevant receivers’ vision, the genotypes may have nuanced pheno-
typic differences beyond human perception (Figure 5).
Our results highlight that genetic polymorphism expressed at the
phenotypic level is not always clear- cut to define as categorization
depends on the perception. With a wood tiger moth stock originating
from north Europe, we validate that male hindwing colour is geneti-
cally controlled by a single Mendelian locus, where the white allele
(W) is dominant to the yellow (y) allele. Male genotypes can be told
apart using colour metrics with high accuracy by machine learning al-
gorithms, but not by human obser vers, because white heterozygous
males can be separated from white homozygotes by differences in
ultraviolet reflectance. In turn, female genotypes are currently in-
separable by their phenotypes to us. It seems plausible that female
colour is controlled to some extent by the same genetic loci as the
male colour, although with different dominance relationships and
other interacting loci. Although we have less- extensive data about
colour variation in females, it seems that in many localities a yellow-
orange- red continuum is common (Lindstedt et al., 2011).
Generally, functional genes in the melanin biosynthetic path-
way can affect both wing scale pigmentation and morphology in
Lepidoptera (Matsuoka & Monteiro, 2018). Our preliminary pig-
ment analyses in the wood tiger moth indicate that the white pig-
mentation in the wings is produced by N- acetyldopamine (NADA)
sclerotin, whereas the yellow pigment is derived from a mix of N-
β- alanyldopamine (NBAD) sclerotin and pheomelanin and the red
pigment results from a dopamine- derived pheomelanin (Brien et al.
In Prep.). Also, differential expression in melanin- promoting and
melanin- inhibiting genes impacts black colouration in the cuticle and
in the hairs of wood tiger moth caterpillars (Galarza, 2021). Plausibly,
differential regulation in genes involved in the melanin pathway
could contribute to the colour differences between the sexes (Gazda
et al., 2020). Whether an upregulation of a single gene that causes
FIGURE 4 Wood tiger moth genotype separability through ecologically relevant vision systems compared to human vision. We tested
the discriminability of the genotypes using three different vision models (human, avian and moth vision). In all images, x- axis represents
pairwise genotype comparisons and y- axis shows chromatic contrast (dS) of the two moths being compared. The region of interest (ROI)
indicates the comparison between abdomen (AB), forewing (FW) and hindwing (HW) colour. The panels separate vision modelling results for
human (a), avian (b) and moth (c) vision models. Contrast values <1 are considered indistinguishable and values above this are increasingly
easy to distinguish (outliers not shown). The black horizontal line indicates dS = 1 corresponding to the perception threshold of the contrast
pigment degradation in the other sex could take place in wood tiger
moth's melanin- based colouration is currently unknown but war-
rants further investigation.
The machine learning algorithm was more efficient at assign-
ing individuals to known group memberships based on the subtle,
but consistent phenotypic differences between the genotypes in
comparison to human observers. The finding that computational
approach outperforms human perception is not surprising, but still
the majority of the colour polymorphism research relies on classic
approach using human- visible categorization. In males, the linear
discriminant analysis separated the three- group problem with high
accuracy. Successful discrimination between the three groups of
males was expected due to differences in short and long wavelength
reflectance between the two white morphs (Henze et al., 2018;
Nokelainen et al., 2012). Also, the white homozygotes have a lower
thorax UV reflectance, smaller thorax by abdomen ratio (i.e. larger
abdomens), smaller forewing by abdomen ratio, lower abdominal
UV reflectance and less variable forewing patterning. In females,
the sorting accuracy was not any better than from expected ran-
dom chance frequency. Genotypes clustered more closely together
in phenotypic space and the three- group problem was separated
with low accuracy. It may be possible to improve this prediction ac-
curacy using different boundary selection protocol; however, it will
not change the fact that the females are phenotypically more similar
than males. The covariation of some of these phenotypic differences
is still unclear, however, it may be possible to use these phenotypic
associations in combination to predict genotypes of wild caught in-
dividuals. It will be our future task to investigate whether increasing
data over the years and developing methods (e.g. convolutional neu-
ral networks) will enhance the prediction accuracy.
From an evolutionary standpoint, it is plausible that interplay
between natural and sexual selection facilitates polymorphism in
this species (Gordon et al., 2015, 2018; Nokelainen et al., 2012;
Rönkä et al., 2020). Since male wood tiger moths, which are ac-
tively searching for females in the vegetation, have limited ability
to see differences in yellow- orange- red hues (Henze et al., 2018),
it is unlikely that sexual selection alone would be responsible for
the colouration of females, but we cannot exclude the possibil-
ity that male colouration could be used in intraspecific commu-
nication. Moreover, recent studies in other species have shown
that UV may facilitate separation of incipient species as recently
demonstrated in Colias butter flies (Fic ar rotta et al., 2022) and that
the differences in UV reflection may arise from novel duplication
of the gene producing sex- specific differences in reflectance as
in Zerene cesonia butterfly (Rodriguez- Caro et al., 2021). Previous
experiments have shown that birds learn to avoid red wood tiger
moths more effectively than yellow or white ones (Ham et al.,
2006; Lindstedt et al., 2011; Rönkä et al., 2018), but the selection
FIGURE 5 Representation of the wood
tiger moth genotypes to illustrate how
they may appear to ecologically relevant
receivers. These false image examples
show genotypes of both sexes organized
in vertical columns and human, avian and
moth false colour images in horizontal
groups. For trichromatic human vision
sw, mw and lw sensitivities were used for
blue, green and red channels respectively.
For tetrachromatic avian vision uv, sw
and lw sensitivities were used for blue,
green and red channels respectively. For
trichromatic moth vision uv, sw and mw
sensitivities were used for blue, green and
red channels respectively. Images were
corrrected for better image screening
for visual signals may be altered due to multimodal signalling
(Rojas et al., 2018; Winters et al., 2021). Also, avian predators may
distinguish between the nuances in colouration among the gen-
otypes as they, and wood tiger moths, perceive UV wavelengths
that are beyond human perception (Henze et al., 2018). Thus, eco-
logically relevant receivers, predators and conspecifics, may exert
different selection pressures on visual signals beyond our percep-
tion (Endler, 1978) and maintain colour polymorphism in natural
conditions (Galarza et al., 2014; Mochida, 2011; Nokelainen et al.,
2014; Rönkä et al., 2020).
Conclusively, determining genotypes based on their pheno-
typic characteristics is important in any species, because it al-
lows studying allele dynamics in the wild (e.g. as in tiger moths
(Brakefield & Liebert, 1985; Fisher & Ford, 1947; Liebert &
Brakefield, 1990), lizards (Sinervo & Calsbeek, 2006; Sinervo &
Lively, 1996) and damselflies (Le Rouzic et al., 2015; Svensson
& Abbott, 20 05)). Colour polymorphisms have several fitness
consequences in the maintenance of genetic variation (Galarza
et al., 2014; Gray & McKinnon, 2007; McKinnon & Pierotti, 2010).
They can influence intraspecific variation in mating cues (Merrill
et al., 2012; Nokelainen et al., 2012), fitness of colour morphs
in different light environments due to increased predation risk
(Nokelainen et al., 2014, 2021; Rojas et al., 2014) and divergence
in thermoregulatory capabilities (Forsman, 2000; Hegna et al.,
2013; Lindstedt et al., 2009). As it has become possible to model
the conspicuousness of different genotypes to different receivers
(Endler & Basolo, 1998; Hart, 2001a; Henze et al., 2018), we may
soon be able to estimate how their appearance shapes the fate
of allelic combinations using long- term data sets (Le Rouzic et al.,
2015; Svensson & Abbott, 2005). Ultimately, this will broaden our
understanding of how genetic variation underlying phenotypic
evolution is shaped in nature.
We thank the members of the plantaginis research group and helping
to sort the phenotypes in the quiz, as well as University of Jyväskylä
Department of Biological and Environmental Science Darwin Club
for discussions.
Authors have no conflict of interest to declare.
ON wrote the fi r st dr a f t of the man u s c r ipt, con d u c ted th e im a g e an al-
ysis and analysed frequency data; JAG devised the crossing- design
and inheritance analyses; JK helped with genotype– phenotype fre-
quencies analyses; KS reared tens of thousands wood tiger moths
over consecutive years and JM contributed substantially to project
management and manuscript editing.
The peer review history for this article is available at https://publo n/10.1111/je b.13994.
The supporting data are archived in a public repository (https://jyx. e/12345 6789/79808): jyx/
datas et/79808.
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... The wood tiger moth (Arctia plantaginis) represents a compelling study species to investigate how different selective pressures can act on a single color locus and maintain within-population trait variation. In this system, male hindwing coloration is determined by a simple genetic basis (Suomalainen 1938;Nokelainen et al., 2022b;Brien et al., 2022): a one locustwo allele polymorphism (dominant W allele and recessive y allele), which translates into white (genotype: WW, Wy) and yellow (genotype: yy) males. Because this is an aposematic moth species, the color trait is not only used for intraspecific communication (i.e., sexual selection) but also to advertise their unpalatability to predators (i.e., interspecific communication). ...
... The wood tiger moth (Arctia plantaginis) (formerly Parasemia plantaginis; Rönkä et al., 2016) is a polymorphic and aposematic moth species. The male hindwing coloration is determined by a simple genetic mechanism where a one locus-two allele (W and y allele) polymorphism translates into white (WW or Wy genotype) or yellow (yy genotype) male morphs (Suomalainen 1938;Nokelainen et al., 2022b) (Fig. 1). Females do not phenotypically express the male color alleles as their hindwing coloration varies continuously from yellow to red but pass the color alleles to their offspring (Nokelainen et al., 2022b) (Fig. 1). ...
... The male hindwing coloration is determined by a simple genetic mechanism where a one locus-two allele (W and y allele) polymorphism translates into white (WW or Wy genotype) or yellow (yy genotype) male morphs (Suomalainen 1938;Nokelainen et al., 2022b) (Fig. 1). Females do not phenotypically express the male color alleles as their hindwing coloration varies continuously from yellow to red but pass the color alleles to their offspring (Nokelainen et al., 2022b) (Fig. 1). The wood tiger moth is a capital breeder; it does not feed at the adult stage, making the larval diet very important for both their development and the adult stage (e.g., sperm quality, egg numbers) (Tammaru and Haukioja 1996). ...
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The persistence of intrapopulation phenotypic variation typically requires some form of balancing selection since drift and directional selection eventually erode genetic variation. Heterozygote advantage remains a classic explanation for the maintenance of genetic variation in the face of selection. However, examples of heterozygote advantage, other than those associated with disease resistance are rather uncommon. Across most of its distribution, males of the aposematic moth Arctia plantaginis have two hindwing phenotypes determined by a heritable one locus-two allele polymorphism (genotypes: WW/Wy = white morph, yy = yellow morph). Using genotyped moths we show that the presence of one or two copies of the yellow allele affects several life-history traits. Reproductive output of both males and females, and female mating success are negatively affected by two copies of the yellow allele. Females carrying one yellow allele (i.e. Wy) have higher fertility, hatching success, and offspring survival than either homozygote, thus leading to strong heterozygote advantage. Our results indicate strong female contribution especially at the postcopulatory stage in maintaining the color polymorphism. The interplay between heterozygote advantage, yellow allele pleiotropic effect and morph-specific predation pressure may exert balancing selection on the color locus, suggesting that color polymorphism may be maintained through complex interactions between natural and sexual selection. This article is protected by copyright. All rights reserved.
Behavioral ecologists have long studied the role of coloration as a defense against natural enemies. Recent reviews of defensive coloration have emphasized that these visual signals are rarely selected by single predatory receivers. Complex interactions between signaler, receiver, and environmental pressures produce a striking array of color strategies—many of which must serve multiple, sometimes conflicting, functions. In this review, we describe six common conflicts in selection pressures that produce multifunctional color patterns, and three key strategies of multifunctionality. Six general scenarios that produce conflicting selection pressures on defensive coloration are: (1) multiple antagonists, (2) conspecific communication, (3) hunting while being hunted, (4) variation in transmission environment, (5) ontogenetic changes, and (6) abiotic/physiological factors. Organisms resolve these apparent conflicts via (1) intermediate, (2) simultaneous, and/or (3) plastic color strategies. These strategies apply across the full spectrum of color defenses, from aposematism to crypsis, and reflect how complexity in sets of selection pressures can produce and maintain the diversity of animal color patterns we see in nature. Finally, we discuss how best to approach studies of multifunctionality in animal color, with specific examples of unresolved questions in the field.
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Colour is often used as an aposematic warning signal, with predator learning expected to lead to a single colour pattern within a population. However, there are many puzzling cases where aposematic signals are also polymorphic. The wood tiger moth, Arctia plantaginis , uses bright hindwing colours as a signal of unpalatability, and males have discrete colour morphs which vary in frequency geographically. In Finland, both white and yellow morphs can be found, and these colour morphs also differ in behavioural and life-history traits. Complex polymorphisms such as these are often explained by supergenes. Here, we show that male colour is linked to an extra copy of a yellow family gene that is only present in the white morphs. This white-specific duplication, which we name valkea , is highly upregulated during wing development, and could act to reduce recombination, thus potentially representing a supergene. We also characterise the pigments responsible for yellow, white and black colouration, showing that yellow is partly produced by pheomelanins, while black is dopamine-derived eumelanin. The yellow family genes have been linked to melanin synthesis and behavioural traits in other insect species. Our results add to only a few examples of seemingly paradoxical and complex polymorphisms which are associated with single genes.
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Significance Incipient species are at an intermediate stage of speciation where reproductive isolation is counteracted by the homogenizing effects of gene flow. Human activity sometimes leads such species to reunite, as seen in the Orange Sulphur butterfly, which forms large hybridizing populations with the Clouded Sulphur in alfalfa fields. Here we show that sex chromosomes maintain these species as distinct, while the rest of their genome is admixed. Sex chromosomes notably determine which males display to females a bright, iridescent UV signal on their wings. Genetic mapping, antibody stainings, and CRISPR knockouts collectively indicate that the gene bric a brac controls whether UV-iridescent nanostructures develop in each species, illustrating how a master switch gene modulates a male courtship signal.
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A big question in behavioral ecology is what drives diversity of color signals. One possible explanation is that environmental conditions, such as light environment, may alter visual signaling of prey, which could affect predator decision-making. Here, we tested the context-dependent predator selection on prey coloration. In the first experiment, we tested detectability of artificial visual stimuli to blue tits (Cyanistes caeruleus) by manipulating stimulus luminance and chromatic context of the background. We expected the presence of the chromatic context to facilitate faster target detection. As expected, blue tits found targets on chromatic yellow background faster than on achromatic grey background whereas in the latter, targets were found with smaller contrast differences to the background. In the second experiment, we tested the effect of two light environments on the survival of aposematic, color polymorphic wood tiger moth (Arctia plantaginis). As luminance contrast should be more detectable than chromatic contrast in low light intensities, we expected birds, if they find the moths aversive, to avoid the white morph which is more conspicuous than the yellow morph in low light (and vice versa in bright light). Alternatively, birds may attack first moths that are more detectable. We found birds to attack yellow moths first in low light conditions, whereas white moths were attacked first more frequently in bright light conditions. Our results show that light environments affect predator foraging decisions, which may facilitate context-dependent selection on visual signals and diversity of prey phenotypes in the wild.
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Aposematic organisms warn predators of their unprofitability using a combination of defenses, including visual warning signals, startling sounds, noxious odors, or aversive tastes. Using multiple lines of defense can help prey avoid predators by stimulating multiple senses and/or by acting at different stages of predation. We tested the efficacy of three lines of defense (color, smell, taste) during the predation sequence of aposematic wood tiger moths ( Arctia plantaginis ) using blue tit ( Cyanistes caeruleus ) predators. Moths with two hindwing phenotypes (genotypes: WW/Wy = white, yy = yellow) were manipulated to have defense fluid with aversive smell (methoxypyrazines), body tissues with aversive taste (pyrrolizidine alkaloids) or both. In early predation stages, moth color and smell had additive effects on bird approach latency and dropping the prey, with the strongest effect for moths of the white morph with defense fluids. Pyrrolizidine alkaloid sequestration was detrimental in early attack stages, suggesting a trade-off between pyrrolizidine alkaloid sequestration and investment in other defenses. In addition, pyrrolizidine alkaloid taste alone did not deter bird predators. Birds could only effectively discriminate toxic moths from non-toxic moths when neck fluids containing methoxypyrazines were present, at which point they abandoned attack at the consumption stage. As a result, moths of the white morph with an aversive methoxypyrazine smell and moths in the treatment with both chemical defenses had the greatest chance of survival. We suggest that methoxypyrazines act as context setting signals for warning colors and as attention alerting or “go-slow” signals for distasteful toxins, thereby mediating the relationship between warning signal and toxicity. Furthermore, we found that moths that were heterozygous for hindwing coloration had more effective defense fluids compared to other genotypes in terms of delaying approach and reducing the latency to drop the moth, suggesting a genetic link between coloration and defense that could help to explain the color polymorphism. Conclusively, these results indicate that color, smell, and taste constitute a multimodal warning signal that impedes predator attack and improves prey survival. This work highlights the importance of understanding the separate roles of color, smell and taste through the predation sequence and also within-species variation in chemical defenses.
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Sexually dimorphic development is responsible for some of the most remarkable phenotypic variation found in nature. Alternative splicing of the transcription factor gene doublesex is a highly conserved developmental switch controlling the expression of sex specific pathways. Here, we leverage sex-specific differences in butterfly wing color pattern to characterize the genetic basis of sexually dimorphic development. We use RNA-seq, immunolocalization, and motif binding site analysis to test specific predictions about the role of Doublesex in the development of structurally-based ultraviolet (UV) wing patterns in Zerene cesonia (Southern Dogface). Unexpectedly, we discover a novel duplication of Doublesex that shows a sex-specific burst of expression associated with the sexually dimorphic UV coloration. The derived copy consists of a single exon that encodes a DNA binding but no protein binding domain, and has experienced rapid amino-acid divergence. We propose the novel dsx paralog may suppress UV scale differentiation in females, which is supported by an excess of Dsx binding sites at cytoskeletal and chitin-related genes with sex-biased expression. These findings illustrate the molecular flexibility of the dsx gene in mediating the differentiation of secondary sexual characteristics.
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Coloration is perhaps one of the most prominent adaptations for survival and reproduction of many taxa. Coloration is of particular importance for aposematic species, which rely on their coloring and patterning acting as a warning signal to deter predators. Most research has focused on the evolution of warning coloration by natural selection. However, little information is available for color mutants of aposematic species, particularly at the genomic level. Here, I compare the transcriptomes of albino mutant caterpillars of the aposematic wood tiger moth (Arctia plantaginis) to those of their full sibs having their distinctive orange‐black warning coloration. The results showed >290 differentially expressed genes genome‐wide. Genes involved in the immune system, structural constituents of cuticular, and immunity were mostly downregulated in the albino caterpillars. Surprisingly, higher expression was observed in core melanin genes from albino caterpillars, suggesting that melanin synthesis may be disrupted in terminal ends of the pathway during its final conversion. Taken together, these results suggest that caterpillar albinism may not be due to a depletion of melanin precursor genes. In contrast, the albino condition may result from the combination of faulty melanin conversion late in its synthesis and structural deficiencies in the cuticular preventing its deposition. The results are discussed in the context of how albinism may impact individuals of aposematic species in the wild. Albinism in warningly colored caterpillars.
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Significance Arthropods are excellent indicators for studying global change in the rapidly changing climate of the Arctic. We used the most comprehensive standardized dataset on Arctic arthropods to quantify diversity and abundance variation over 24 y in an area that is warming rapidly. Overall arthropod abundance and diversity showed opposing nonlinear trends, with a sharp increase in overall abundance in recent years. However, trends varied substantially among taxa and habitats and several groups declined in abundance. We found strong evidence of conditions outside the growing season and density-dependent feedbacks affecting abundance. Our results emphasize the need for a more integrated approach to investigating arthropod responses to environmental stressors at finer taxonomic resolution and by incorporating time-lagged effects.
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Warning signals are predicted to develop signal monomorphism via positive frequency‐dependent selection (+FDS) albeit many aposematic systems exhibit signal polymorphism. To understand this mismatch, we conducted a large‐scale predation experiment in four countries, among which the frequencies of hindwing warning coloration of the aposematic moth, Arctia plantaginis, differ. Here we show that selection by avian predators on warning colour is predicted by local morph frequency and predator community composition. We found +FDS to be the strongest in monomorphic Scotland and lowest in polymorphic Finland, where the attack risk of moth morphs depended on the local avian community. +FDS was also found where the predator community was the least diverse (Georgia), whereas in the most diverse avian community (Estonia), hardly any models were attacked. Our results support the idea that spatial variation in predator communities alters the strength or direction of selection on warning signals, thus facilitating a geographic mosaic of selection. A geographic mosaic of selection by predators could explain the paradoxical maintenance of warning signal variation, but direct ecological evidence is scarce and focused on tropical systems. We monitored local avian predators and attacks on 4000 + moth models representing red, yellow or white warning colour morphs in a temperate moth system with natural variation in local morph frequencies. We found positive frequency‐dependent selection to be strongest in monomorphic populations and the direction and strength of selection to be significantly associated with local predator community composition and diversification, which can explain not only geographic variation (polytypism) but also local polymorphism when coupled with gene flow.
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To understand the function of colour signals in nature, we require robust quantitative analytical frameworks to enable us to estimate how animal and plant colour patterns appear against their natural background as viewed by ecologically relevant species. Due to the quantitative limitations of existing methods, colour and pattern are rarely analysed in conjunction with one another, despite a large body of literature and decades of research on the importance of spatio‐chromatic colour pattern analyses. Furthermore, key physiological limitations of animal visual systems such as spatial acuity, spectral sensitivities, photoreceptor abundances and receptor noise levels are rarely considered together in colour pattern analyses. Here, we present a novel analytical framework, called the Quantitative Colour Pattern Analysis (QCPA). We have overcome many quantitative and qualitative limitations of existing colour pattern analyses by combining calibrated digital photography and visual modelling. We have integrated and updated existing spatio‐chromatic colour pattern analyses, including adjacency, visual contrast and boundary strength analysis, to be implemented using calibrated digital photography through the Multispectral Image Analysis and Calibration (MICA) Toolbox. This combination of calibrated photography and spatio‐chromatic colour pattern analyses is enabled by the inclusion of psychophysical colour and luminance discrimination thresholds for image segmentation, which we call ‘Receptor Noise Limited Clustering’, used here for the first time. Furthermore, QCPA provides a novel psycho‐physiological approach to the modelling of spatial acuity using convolution in the spatial or frequency domains, followed by ‘Receptor Noise Limited Ranked Filtering’ to eliminate intermediate edge artefacts and recover sharp boundaries following smoothing. We also present a new type of colour pattern analysis, the ‘local edge intensity analysis’ as well as a range of novel psycho‐physiological approaches to the visualization of spatio‐chromatic data. QCPA combines novel and existing pattern analysis frameworks into what we hope is a unified, free and open source toolbox and introduces a range of novel analytical and data‐visualization approaches. These analyses and tools have been seamlessly integrated into the MICA toolbox providing a dynamic and user‐friendly workflow.
Most animal species on Earth are insects, and recent reports suggest that their abundance is in drastic decline. Although these reports come from a wide range of insect taxa and regions, the evidence to assess the extent of the phenomenon is sparse. Insect populations are challenging to study, and most monitoring methods are labor intensive and inefficient. Advances in computer vision and deep learning provide potential new solutions to this global challenge. Cameras and other sensors can effectively, continuously, and noninvasively perform entomological observations throughout diurnal and seasonal cycles. The physical appearance of specimens can also be captured by automated imaging in the laboratory. When trained on these data, deep learning models can provide estimates of insect abundance, biomass, and diversity. Further, deep learning models can quantify variation in phenotypic traits, behavior, and interactions. Here, we connect recent developments in deep learning and computer vision to the urgent demand for more cost-efficient monitoring of insects and other invertebrates. We present examples of sensor-based monitoring of insects. We show how deep learning tools can be applied to exceptionally large datasets to derive ecological information and discuss the challenges that lie ahead for the implementation of such solutions in entomology. We identify four focal areas, which will facilitate this transformation: 1) validation of image-based taxonomic identification; 2) generation of sufficient training data; 3) development of public, curated reference databases; and 4) solutions to integrate deep learning and molecular tools.
Canaries changing colors Many animals are sexually dimorphic, with different phenotypes in males and females. To identify the genetic basis of sexual differences in bird coloration, Gazda et al. investigated red coloration in mosaic canaries and related species (see the Perspective by Chen). Using a combination of genetic crosses, genomic mapping, transcriptomics, and comparative analyses, the authors show that trans-regulation of the carotenoid-processing gene BCO2 is involved in sexual dichromatism. Although such variation in coloration among the sexes is common, particularly in birds, there are few candidate genes known to be involved. This study helps to elucidate the molecular mechanisms that underlie the evolution of dichromatism and may aid in uncovering sexually selected traits. Science , this issue p. 1270 ; see also p. 1185