Heritability and adaptive significance of the number of egg-dummies in the cichlid fish Astatotilapia burtoni.

Abstract

Cichlid fishes are a textbook example of rapid speciation and exuberant diversity--this applies especially to haplochromines, a lineage with approximately 1800 species. Haplochromine males uniquely possess oval, bright spots on their anal fin, called 'egg-spots' or 'egg-dummies'. These are presumed to be an evolutionary key innovation that contributed to the tribe's evolutionary success. Egg-spots have been proposed to mimic the ova of the mouthbrooding females of the corresponding species, contribute to fertilization success and even facilitate species recognition. Interestingly, egg-spot number varies extensively not only between species, but also within some populations. This high degree of intraspecific variation may appear to be counterintuitive since selection might be expected to act to stabilize traits that are correlated with fitness measures. We addressed this 'paradox' experimentally, and found that in the haplochromine cichlid Astatotilapia burtoni, the number of egg-spots was related to male age, body condition and dominance status. Intriguingly, the egg-spot number also had a high heritable component (narrow sense heritability of 0.5). These results suggest that the function of egg-spots might have less to do with fertilization success or species recognition, but rather relate to mate choice and/or male-male competition, helping to explain the high variability in this important trait.

Full text

Page 1

Heritability and adaptive significance of the
number of egg-dummies in the cichlid
fish Astatotilapia burtoni
Topi K. Lehtonen1,2and Axel Meyer1,*
1Lehrstuhl fu ¨r Zoologie und Evolutionsbiologie, Department of Biology, University of Konstanz,
78457 Konstanz, Germany
2Section of Ecology, Department of Biology, University of Turku, 20014 Turku, Finland
Cichlid fishes are a textbook example of rapid speciation and exuberant diversity—this applies especially
to haplochromines, a lineage with approximately 1800 species. Haplochromine males uniquely possess
oval, bright spots on their anal fin, called ‘egg-spots’ or ‘egg-dummies’. These are presumed to be an evol-
utionary key innovation that contributed to the tribe’s evolutionary success. Egg-spots have been
proposed to mimic the ova of the mouthbrooding females of the corresponding species, contribute to fer-
tilization success and even facilitate species recognition. Interestingly, egg-spot number varies extensively
not only between species, but also within some populations. This high degree of intraspecific variation
may appear to be counterintuitive since selection might be expected to act to stabilize traits that are cor-
related with fitness measures. We addressed this ‘paradox’ experimentally, and found that in the
haplochromine cichlid Astatotilapia burtoni, the number of egg-spots was related to male age, body con-
dition and dominance status. Intriguingly, the egg-spot number also had a high heritable component
(narrow sense heritability of 0.5). These results suggest that the function of egg-spots might have less
to do with fertilization success or species recognition, but rather relate to mate choice and/or male–
male competition, helping to explain the high variability in this important trait.
Keywords: body condition; dominance hierarchy; narrow sense heritability; intraspecific variation;
key innovation; signal value
1. INTRODUCTION
A famously high number of species, and unsurpassed
rates of evolution, have made cichlid fishes of the great
East African lakes one of the most remarkable examples
of adaptive radiation [1–4]. The great majority of these
species (approximately 1800 of the 2500 species in the
family Cichlidae) belong to a single lineage, the haplo-
chromine cichlids [5]. The anal fin of a sexually mature
haplochromine male carries bright yellow or orange
ovoid spots, which have been suggested to mimic real
eggs of the species, and have therefore been called egg-
dummies [6]. Females of some species also have spots,
but these are less pronounced and on average lower in
number. Interestingly, the less diverse lineages of cichlids
do not posses similar egg-spots [7], and indeed, posses-
sion of egg-dummies may have promoted speciation
within the group [5,8] and have been considered to be a
key innovation that has contributed to the evolutionary
success of haplochromines [5,7]. As a consequence of
the particular mating system of female mouthbrooding
and the concomitant evolution of egg-spots, these fish
are limited by the number of eggs that fit into a female’s
mouth (typically only 20–40), which might have contrib-
uted to smaller populations sizes and hence faster rates of
evolution [5,7].
In haplochromines, fertilization typically takes place in
the mouth of the female after she has nipped at the male’s
egg-dummies, which are located near the male’s genital
opening (figure 1; see the electronic supplementary
material, video clip; see also [7]). Hence, egg-spots have
been suggested to enhance fertilization success and facili-
tate spawning ([6], but see [9]). A few other functions for
the spots have also been proposed (e.g. [8,10]), such as
certainty of paternity and species recognition. Indeed,
arrangement and number of egg-spots differs between
haplochromine species [7,10–12].
Interestingly, in some species the number of egg-spots
is not fixed, but varies among individuals of a population.
For example, in a natural breeding colony of A. burtoni,
the number of egg-spots ranged between five and nine
[13]. Similarly, sibling males of a Lake Malawi haplochro-
mine species (Labeotropheus) were reported to have from
two to six egg-spots [14]. This type of variation calls for
an explanation, as traits important for successful fertili-
zation, species recognition or fitness, in general, are
expected to be under stabilizing selection, which should
erode heritable genetic variation behind these traits (e.g.
[15]). However, so far the sources for this variation in
egg-spots, an evolutionary key trait [5], have not been
investigated.
We examined within-species variation in the number of
egg-spots in the haplochromine cichlid A. burtoni, a
widely used model system in behavioural endocrinology
and genomics [16–22]. The species lives in ponds and
riversaroundLakeTanganyika,EastAfrica. As
* Author for correspondence (axel.meyer@uni-konstanz.de).
Electronic supplementary material is available at http://dx.doi.org/
10.1098/rspb.2010.2483 or via http://rspb.royalsocietypublishing.org.
Proc. R. Soc. B (2011) 278, 2318–2324
doi:10.1098/rspb.2010.2483
Published online 5 January 2011
Received 13 November 2010
Accepted 6 December 2010
2318
This journal is q 2011 The Royal Society
on June 27, 2011rspb.royalsocietypublishing.org Downloaded from

Page 2

characteristic for haplochromines, females of the species
are mouth brooders, whereas males do not contribute to
the care of offspring. Males are highly aggressive and
only successful territory holders are able to attract females
to spawn [13]. A manipulation of egg-spot number in a
haplochromine species closely related to A. burtoni
suggested that females discriminate against males with a
reduced number of spots [9]. This is notable since the
number of egg-spots is highly variable among males of
the species (among males used in this study 4–15; see
also [11,13]). We tested five mutually non-exclusive
explanations that could contribute to the high variation
in egg-spot number, observed within A. burtoni popu-
lations: (1) egg-spot number is a function of body size
[11], e.g. because large males also have larger anal fins
that can potentially accommodate a larger number of
egg-dummies of a given size, (2) the number of egg-
dummies increases over time [14], which could predict
the number of spots better than body size per se, (3) the
number of egg-spots depends on body condition of the
male, suggesting that they are costly to produce or main-
tain [23,24], (4) the number of egg-dummies is a badge of
status, or more generally, dominant males have more
spots [14,25], and finally, (5) explanations 3 and 4
include the possibility that the number of spots has a sig-
nificant genetic component and it may therefore signal
heritable differences between the males that selection
can act on. We designed a series of laboratory experiments
to test each of these possibilities.
2. MATERIAL AND METHODS
The experiments were conducted in the animal care facility at
the University of Konstanz, Germany, under a 12 L:12 D
cycle and water temperature of 25+28C. The fish were fed
daily with commercial fish foods if nototherwise noted. Juven-
iles were also fed with newly hatched brine shrimps (Artemia).
Under these conditions, A. burtoni reaches sexual maturity
and starts to breed at the age of approximately four months,
with even the smallest and most subdominant individuals
maturing in six months (T. K. Lehtonen & A. Meyer, 2008,
personal observations), having attained a total length of
6–6.5 cm ([26]; T. K. Lehtonen & A. Meyer, 2008, personal
observations). In the wild, the exact population structure
seems to depend on the habitat [26,27], but most breeding
populations are likely to consist of young individuals due to
heavy natural predation and fishery [26,27]. Under these con-
ditions, males do not seem to survive long enough to exceed
the total length of 10–12 cm [26], although in captivity the
species can ultimately attain a larger size (15 cm or even
larger) and live for several years. However, many individuals
start to show signs of senescence (ragged fins, deformations,
increased mortality and decreased rate of breeding) even
before the age of 2 years (T. K. Lehtonen & A. Meyer,
2008, personal observations). Accordingly, we used young,
mature males to assess the following five explanations for the
high within-population variation in the number of egg-spots
in male A. burtoni.
(a) Explanation 1: body size
To assess whether there is a relationship between male size
and the number or size of his egg-spots, we photographed
four different groups of males: (1) 64 males of the age of
7.5 months (the second generation from ‘explanation 5’,
see below), (2) 12 relatively young, mature males (age 10.5
months), (3) another of group of males of the same age
(10.5 months, n ¼ 15) and (4) 12 older, large males (16
months). At least two photographs of each male were taken
on a grid paper that was covered with a smooth, transparent
plastic sheet. Two photographs of each male were later ana-
lysed using Sigma Scan Pro 5.0 (SPSS Inc.) software, and
average values from these measurements were used for stat-
istical analyses. Where relevant, this same photography
procedure was also used in other experiments, described
below. We counted as an egg-spot a bright, ovoid-shaped
spot on the male’s anal fin with a rim of lighter coloration
around it (figure 1, see also [7]).
(b) Explanation 2: changes over time
Different stocking conditions (most importantly tank size, com-
petitive regime and social background) of the different (age)
groups of ‘explanation 1’ did not allow formal tests over these
groups. Hence, in order to investigate whether the number
(or size) of egg-spots changes over time (and hence presumably
with age), males of the groups 2–4 were photographed again
after a period of time. Specifically, we kept the males of
group 2 singly for two months in tanks measuring 50 ? 25 ?
30 cm (length ? width ? height), the males of group 3 in con-
tact with 2–4 conspecifics in tanks measuring 60 ? 40 ? 35 cm
for 1–2 months, and the males of group 4 singly for three
months in the 50 ? 25 ? 30 cm tanks.
(c) Explanation 3: condition dependence
In this experiment, we examined whether the number or size
of egg-spots depends on body condition of the male. Imma-
ture juveniles that did not yet possess egg-spots (age under
three months) were divided into two treatments: food
restricted individuals (n ¼ 18) were given food on 3 days a
week, whereas control individuals were fed daily (n ¼ 18).
The same number (1–5) of individuals from any one family
was used for both treatments. All male offspring (n ¼ 10 in
both treatments) were photographed at the age of 7.5 months.
Besides comparing egg-spot number and size directly
between the treatments, we also ran a general linear model
(GLM) on egg-spot number, with total length as a
10 cm
Figure 1. Male A. burtoni and a close-up of his anal fin with
egg-spots. A 1 mm grid on the background provides a scale
for assessing the size of the egg-spots.
Variation in evolutional key trait
T. K. Lehtonen & A. Meyer 2319
Proc. R. Soc. B (2011)
on June 27, 2011rspb.royalsocietypublishing.orgDownloaded from

Page 3

continuous factor and treatment as a categorical factor. After
the interaction between variables ‘total length’ and ‘treat-
ment’ was found to be non-significant, we removed it from
the model and proceeded with assessing the main effects.
(d) Explanation 4: dominance hierarchy
Earlier work on A. burtoni has shown that dominant, territor-
ial males have higher circulating plasma concentrations of
androgens than subdominant, non-territorial males [17],
but males are nevertheless able to switch between the two
territorial stages within minutes depending on the immediate
social environment [16]. We had two different set-ups to test
for any relationship between male dominance and the
number or size of his egg-spots. First, the most dominant
male in 14 different family groups (see ‘explanation 5’,
below) was identified (as distinguished by coloration and
interactions among the individuals in the tank) and com-
pared with the rest of the males of that family. In one case
we took an average over three males that appeared to be
equally dominant. The sample size for this measurement
was 13 because one of the 14 families had only a single
male. Secondly, in a separate experiment, two visually size-
matched (for length and condition) males (age 16 months)
were placed in a tank measuring 100 ? 40 ? 40 cm. The
tank contained a single 20 cm long and 8 cm wide clay
arch as a shelter. Behaviour of the males was monitored to
assess their relative dominance positions and also to abort
the replicate before males were able to cause unnecessary
stress or any physical damage to each other. In all 20 repli-
cates, only one of the two males claimed the shelter and
the same male was also behaviourally dominant in behaviour-
al interactions observed between the two males. The males
were photographed after the experiment.
(e) Explanation 5: heritable differences
To estimate narrow-sense heritability (h2) of the number and
size of egg-spots, 14 males aged 10.5 months were each
placed into one tank with three females. At least one of the
three females was considered ready to spawn, and the role
of the two other was to diffuse aggression. As soon as a
female was mouthbrooding, the rest of them were returned
to stock tanks and the male was used for the group 3 of
‘explanations 1 and 2’. When the juveniles started to swim
freely, the female was removed from the tank. Each batch
of juveniles (families) was raised in equal-sized partitions
measuring 40 ? 60 ? 50 cm. One large tank (120 ? 60 ?
50 cm) contained three such enclosures, partitioned with
opaque dividers. The enclosure for each batch of juveniles
was assigned randomly. After three months, the maximum
number of juveniles in each family was limited to 14 by ran-
domly removing any number of individuals exceeding 14, to
assure as homogenized conditions across the families as
possible. At the age of 7.5 months, all remaining male
offspring (1–14 per family, average 4.6) were photographed.
Narrow-sense heritability (h2) measures the proportion of
phenotypic variance explained by additive genetic variance; a
high value of h2indicates a high degree of additive genetic
influence. To yield an estimate for h2, and its standard
error, the slope of father–male offspring regression analysis
and its standard error were multiplied by 2 [28]. In the
regression, each offspring value consisted of the mean over
the family. All statistical tests were conducted using Systat
12.0 (SPSS Inc.) software.
3. RESULTS
(a) Explanation 1: body size
Despite the large range in the egg-spot number and total
length among males of each group (table 1), these two
variables correlated only within one of the four male
groups (table 1). In none of the groups male total
length and egg-spot size (area) correlated significantly
(p . 0.10 in all cases).
(b) Explanation 2: changes over time
All values below are given as ‘mean+standard error of
the mean’, if not otherwise noted. In all cases, males
developed 0–2 new egg-spots during the experimental
period and none of them lost any. The change in average
spot number was 0.75+0.18 (Wilcoxon signed-rank test,
z ¼ 2.71, n ¼ 12, p ¼ 0.007), 0.67+0.21 (Wilcoxon
signed-rank test, z ¼ 2.43, n ¼ 15, p ¼ 0.015) and from
0.42+0.19 (Wilcoxon signed-rank test, z ¼ 1.89, n ¼
12, p ¼ 0.059) for the groups 2, 3 and 4, respectively
(figure 2). The corresponding changes in the size of the
spots were 0.40+0.14 mm2(paired t-test, t ¼ 2.81,
d.f. ¼ 11,
t-test, t ¼ 0.040, d.f. ¼ 14, p ¼ 0.97) and 20.031+
0.12 mm2(paired t-test, t ¼ 0.260, d.f. ¼ 11, p ¼ 0.80).
Young males alsogrew
(group 2: paired t-test, t ¼ 8.24, d.f. ¼ 11, p , 0.001;
group 3: paired t-test, t ¼ 6.11, d.f. ¼ 14, p , 0.001),
whereastherewasonlyamarginallynon-significanttendency
for older males to gain length during the study period
(group 4: paired t-test, t ¼ 2.18, d.f. ¼ 11, p ¼ 0.052).
p ¼ 0.017),0.005+0.12 mm2
(paired
during thestudyperiod
(c) Explanation 3: condition dependence
Our food restriction treatment was effective in manipulat-
ing condition: well-fed males were longer (two sample
t-test, t ¼ 3.16, d.f. ¼ 18, p ¼ 0.005) and visually clearly
more ‘bulky’ than food-restricted males of the same age.
Furthermore, at the age of 7.5 months, well-fed males
(6.2+0.9) had on average a larger number of egg-spots
than food-restricted males (4.9+0.9) (two sample
Table 1. Total lengths, egg-spot numbers and correlations between these two traits in four experimental groups of A. burtoni
males.
male group age (months)
total length (mm) egg-spots (n)
nrp
mean+s.e.range mean+s.e. range
1
2
3
4
7.5
10.5
10.5
16
81+0.7
86+1.2
91+1.3
114+1.3
70–100
78–95
80–98
106–121
7.2+0.2
6.4+0.3
9.5+0.6
10.3+0.8
5–11
5–8
6–14
7–14
64
12
15
12
0.265
0.331
20.234
20.173
0.034
0.29
0.40
0.59
2320T. K. Lehtonen & A. Meyer
Variation in evolutional key trait
Proc. R. Soc. B (2011)
on June 27, 2011 rspb.royalsocietypublishing.org Downloaded from

Page 4

t-test, t ¼ 3.40, d.f. ¼ 18, p ¼ 0.003). However, overall
length differences among these 20 males seemed to over-
rule the effect of experimental treatment on egg-spot
number (GLM, treatment effect: F1,17¼ 2.46, p ¼ 0.14;
body size effect: F1,17¼ 4.59, p ¼ 0.047). The size of
egg-spots did not differ between the males of the
two treatments (two sample t-test, t ¼ 0.848, d.f. ¼ 18,
p ¼ 0.41).
(d) Explanation 4: dominance hierarchy
In the groups of individuals raised together until the age
of 7.5 months, the most dominant males were always
larger than the average of the rest of the males (paired
t-test, t ¼ 4.63, d.f.¼12, p ¼ 0.001). However, the
number (7.3+0.4) or size (2.0+0.1 mm2) of egg-
spots of these dominant males did not significantly
differ from the rest of the males (number: 7.0+0.3,
paired t-test, t ¼ 0.898, d.f. ¼ 12, p ¼ 0.39, figure 3;
size: 1.9+0.1 mm2, paired t-test, t ¼ 0.683, d.f. ¼ 12,
p ¼ 0.51). Nevertheless, among visually size-matched
male pairs, the male that exhibited dominant behaviour
and acclaimed the single shelter (which as the only avail-
able structure typically provided the centre for the new
territory) had more egg-spots (10.0+0.8) than the sub-
dominant male (7.9+0.4) (paired t-test, t ¼ 2.57,
d.f. ¼ 19, p ¼ 0.019; figure 3). The average size of
egg-spots was not different between dominant and sub-
dominantmales(dominant
subdominant males 2.1+0.1 mm2; paired t-test, t ¼
0.284, d.f. ¼ 19, p ¼ 0.78). The small size differences
(0–6.5 mm in total length) between the males in a pair,
dueto imperfect visual
dominance (dominant male: 109+1.4 mm; subdomi-
nant: 109+1.5 mm; paired t-test, t ¼ 0.040, d.f. ¼ 19,
p ¼ 0.97).
For the two set-ups combined, dominant males were
larger than subdominants in 24 cases out of 33 (binomial
distribution: p ¼ 0.014), had more egg-spots in 18 cases
out of 25 (with eight cases of equal egg-spot number,
binomial distribution: p ¼ 0.043), and had larger average
spot size in 17 cases out of 33 (binomial distribution: p ¼
1.0). Hence, it seems that when there are large size differ-
ences between males, body size is the most important
males:2.1+0.1 mm2;
matching, didnot affect
determinant
spot number being a better predictor among males of
approximately similar size.
of dominancehierarchy,withegg-
(e) Explanation 5: heritable differences
The number of egg-spots was significantly heritable (h2¼
0.50+0.20); males that had manyegg-spots (at the age of
10.5 months) tended to have sons with many egg-spots
(figure 4). The estimates for narrow sense heritability
of egg-spot size (0.20+0.51) and male total length
(20.32+0.44) did not significantly differ from zero.
4. DISCUSSION
It has been reasoned that the evolution of the egg-spots
and concomitant evolution of the maternal mouthbrood-
ing mating system might have been a key innovation that
permitted the evolution of extremely species-rich cichlid
fish lineage, the haplochromines, which is composed of
about 1800 species [5]. In this study, we set out to explore
the sources of intra-population variation in this key trait.
We found, among A. burtoni males of a certain age,
mixed evidence regarding the relationship between the
number of egg-dummies and body size. We propose that
age (Henning & Meyer, 2009, unpublished manuscript)
or social background of the males may determine whether
such a link between the two traits exists. Heterogeneous
male ages may also explain why an earlier study suggests
the traits to correlate [11]. In any case, the results of the
current study indicate that egg-spot number is not
unequivocally determined
among males of a certain age. Furthermore, males devel-
oped additional egg-spots over time and males in good
condition had more spots than low condition males.
Taken together, these findings suggest that egg-spot
number has potential to purport somewhat different
information than male size per se, giving the female haplo-
chromine cichlids the potential to use egg-spot number as
a mate choice cue. By preferring males that have high-
number of egg-spots, females could mate with older
males (for their size) or males that are in good condition.
In some systems, age correlates with a male’s survival
skills and high immune resistance [29], and females
would therefore benefit by choosing old males that pass
bybody size, especially
0
0.2
0.4
0.6
0.8
1.0
group 2group 3 group 4
change in egg-spot number ± s.e.
Figure 2. The number of egg-spots at the beginning of a
study period subtracted from the egg-spot number at the
end of the period. The numbers of egg-spots have increased
over time in all instances.
–0.5
0
0.5
1.0
1.5
2.0
2.5
3.0
difference in egg-spot number ±s.e.
family groups
size-matched males
Figure 3. The number of egg-spots of the subdominant
male(s) subtracted from the egg-spot number of the domi-
nant male. Positive values indicate that the males with
more egg-spots dominated males with fewer spots.
Variation in evolutional key trait
T. K. Lehtonen & A. Meyer 2321
Proc. R. Soc. B (2011)
on June 27, 2011rspb.royalsocietypublishing.org Downloaded from

Page 5

their good genes to their offspring. Similarly, mating with
a male in good condition is generally thought to provide
corresponding indirect (and sometimes direct) benefits
[23,24,30]. Indeed, females of some haplochromine
cichlid species have been reported to use egg-spot
number as a mate choice cue [9,31,32], although this is
not necessarily the case in A. burtoni (Henning &
Meyer, 2009, unpublished manuscript) or some other
haplochromine species [10]. Furthermore, condition
dependence of a trait (here egg-spot number) as such
may also help to explain how genetic variability in that
trait persists ([23,30], see the discussion below).
We also found that the number of egg-spots has rel-
evance for the dominance hierarchy: although larger and
heavier males generally tend to be more dominant
([10,33]; T. K. Lehtonen & A. Meyer, 2008, personal
observations), among males of approximately the same
size, individuals with many spots dominated those with
a lower egg-spot number. However, in sister groups with
high variation in body size, largest males, more often
than those with the highest egg-spot number, were
socially dominant. Nevertheless, our results suggest that
besides potential relevance of egg-spot number to
female choice, the trait is also important in male–male
competition, predicting dominance status among males
of the same size class. Due to a short lifespan and size
assortative mortality [26], rival males are likely to be
often of similar size in the wild. This is important also
because if the egg-spots are ‘badges of status’ by honestly
indicating a male’s position in dominance hierarchy (rela-
tive to his size) [25], the observed variation in the number
of egg-dummies among males could be more readily
understandable. In such a system, cheaters would be
punished by their rivals [34–36].
A dominant A. burtoni male usually spreads his anal fin
during courtship and male–male interactions, suggesting
that the fish are likely to use the signal potential of egg-
spots in the context of sexual selection. However, even
if the high success of males with a large number of
egg-spots was purely countable to correlates of the trait,
some disadvantages associated with a high egg-spot
number (or its correlates) should be expected, given the
observed high within-population variation in the trait.
These potential disadvantages
mentioned social costs, physiological costs of producing
or maintaining spots [23,37] and higher predation
pressure on males with many bright spots [38–42]. For
example, in small tropical fishes called swordtails (genus
Xiphophorus), females prefer males with a brightly
coloured and long extension of their caudal fin (i.e.
‘sword’), but these males are also preferably targeted
by predatory fish [43,44]. However, we are not aware
of any empirical tests on the potential costs of high
egg-spot number in A. burtoni or other haplochromines.
Given that individual differences in egg-spot number
can be related to passing of time, as well as a male’s dom-
inance status (which may, in turn, change quickly and be
under hormonal control, see [16,17]), the question arises
whether these within-population differences are solely
under environmental control and strongly affected by
phenotypic plasticity. Interestingly, our results suggest
that this is not the case, since the differences among
males in egg-spot number have a significant heritable
component. Specifically, a male with many spots tended
to sire male offspring with a high egg-spot number,
resulting in a high estimate for narrow sense heritability
(h2¼ 0.50) of this conspicuous trait. This finding is
notable because, obviously, traits can respond to selection
only if they have a heritable component. In this respect,
egg-spot number seems to be similar to the length of
caudal fin extensions in male swordtails, a trait under
sexual selection and clear genetic control [45]. More gen-
erally, although mating success and fecundity may cause
more persistent directional evolution than selection
through survival [46], and most traits associated with
fitness are expected to have low heritability values
[15,47,48], traits involved in sexual selection have recur-
rently been reported to have relatively high heritabilities
(reviewed in [49]). For example, in collared flycatchers
(Ficedula albicollis), the size of a male’s forehead patch
had narrow sense heritability of h2¼ 0.381 [50]. In a
population of guppies (Poecilia reticualata) the number
of orange-coloured later spots, a trait relevant to female
mate choice, had an estimated heritability as high as
h2¼ 0.62 [51]. Regarding these comparisons, it is
worth noting that our estimate (h2¼ 0.50) might have
been somewhat influenced by the fact that, for logistical
reasons, the male offspring needed to be photographed
at a slightly younger age than their fathers; it is possible
that the lower range of trait values in the former than
the latter (figure 4) underestimated the strength of
regression.
The large within-population variation in the number of
egg-spots found in this study is also relevant regarding
earlier research suggesting that the number of egg-spots
plays a role in species recognition [8]. Specifically, the
large range of egg-spot number in A. burtoni, and some
other species [14], means that there is likely to be con-
siderable overlap in the number of egg-spots among
many species, albeit males of some species always have
a lower and more consistent egg-spot number [5,7,12].
Our results may therefore call for re-evaluation of egg-
spots as a species recognition cue, although we also note
that it is still possible that the arrangement or appearance
(such as size) of egg-spots is more species-specific
include the above-
4
5
6
7
8
9
10
678910 11 12 1314
father
male offspring
Figure 4. The relationshipbetween the numberofegg-spotsof
fathers and their sons based on mean offspring values (+ s.e.)
over 14 families.
2322 T. K. Lehtonen & A. Meyer
Variation in evolutional key trait
Proc. R. Soc. B (2011)
on June 27, 2011rspb.royalsocietypublishing.orgDownloaded from

Page 6

than their number [5,7,8]. Accordingly, in the current
study, we also assessed the average size of each male’s
egg-spots. When males at the age of 10.5 months were
isolated from other individuals, the size of spots slightly
increased over time. We found no other consistent patterns
concerning the size of the egg-spots in relation to passing
of time, body size, dominance status or condition of
males. Similarly, heritability of the average size of
egg-spots was not significant. These findings are not sur-
prising since new, emerging spots are initially small, and
we also found that large spots sometimes ended up divid-
ing into two smaller spots in older fish. Furthermore, one
might expect that the final size of the spots could be under
stabilizing selection, for example if they truly functioned as
egg mimics [6] or if any sexual selection pressures on egg-
spot size are counter-selected by natural selection [52,53].
However, neither hypothesis has been rigorously tested in
cichlid fishes. Moreover, females do not necessarily
prefer egg-spots that are of the same size as the ova they
produce [8,12].
Here, we have provided insights into the proximate and
ultimate factors that contribute to the variation in the
number of egg-spots on anal fins of male haplochromine
cichlid fish. Multiple, potentially interacting, mechanisms
seem to be involved: dominant males in good condition
are likely to have many egg-spots, the number of which
increases over time. Interestingly, the differences in the
egg-spot number are highly heritable. Hence, it seems
that the primary function of egg-spot number could lie
in sexual selection rather than species recognition or fer-
tility assurance. Future work will also have to further
investigate the developmental proximate mechanisms of
this key trait with special evolutionary importance [5,7].
We thank Janine Sieling for logistical help, Birgit Gogol for
help with measuring the photographs, Hysing Bolstad and
Carla Sgro for their advice on statistical matters, and
Frederico Henning, Matthias Sanetra and two anonymous
referees for helpful comments and discussions. All animal
experiments were approved by the German authorities. The
funding was provided by the Alexander von Humboldt
Foundation fellowship and the Academy of Finland grant
to T.K.L. and by the Deutsche Forschungsgemeinschaft
and the University of Konstanz to A.M.
REFERENCES
1 Meyer,A.1993Phylogeneticrelationshipsandevolutionary
processes in East African cichlid fishes. Trends Ecol. Evol. 8,
279–284. (doi:10.1016/0169-5347(93)90255-N)
2 Stiassny, M. L. J. & Meyer, A. 1999 Cichlids of the rift
lakes. Sci. Am. 280, 64–69. (doi:10.1038/scientificamer-
ican0299-64)
3 Kocher, T. D. 2004 Adaptive evolution and explosive
speciation: the cichlid fish model. Nat. Rev. Genet. 5,
288–298. (doi:10.1038/nrg1316)
4 Seehausen, O. 2006 African cichlid fish: a model system
in adaptive radiation research. Proc. R. Soc. B 273, 1987–
1998. (doi:10.1098/rspb.2006.3539)
5 Salzburger, W., Mack, T., Verheyen, E. & Meyer, A. 2005
Out of Tanganyika: genesis, explosive speciation, key-
innovations and phylogeography of the haplochromine
cichlid fishes. BMC Evol. Biol. 5, 17. (doi:10.1186/
1471-2148-5-17)
6 Wickler, W. 1962 ‘Egg-dummies’ as natural releasers in
mouth-breedingcichlids.
(doi:10.1038/1941092a0)
Nature
194, 1092–1093.
7 Salzburger, W., Braasch, I. & Meyer, A. 2007 Adaptive
sequence evolution in a color gene involved in the for-
mation of the characteristic egg-dummies of male
haplochromine cichlid fishes. BMC Biol. 5, 51. (doi:10.
1186/1741-7007-5-51)
8 Goldschmidt, T. & de Visser, J. 1990 On the possible role
of egg mimics in speciation. Acta Biotheor. 38, 125–134.
(doi:10.1007/BF00047549)
9 Hert, E. 1989 The function of egg-spots in an African
mouth-brooding cichlid fish. Anim. Behav. 37, 726–
732. (doi:10.1016/0003-3472(89)90058-4)
10 Tobler, M. 2006 The eggspots of cichlids: evolution
through sensory exploitation? Z. Fischkunde 8, 39–46.
11 Capron de Caprona, M.-D. & Fritzsch, B. 1984
Interspecific fertile hybrids of haplochromine cichlidae
(teleostei) and their possible importance for specia-
tion.
Neth.J.Zool.
34,
002829684X00272)
12 Couldridge, V. C. K. 2002 Experimental manipulation of
male eggspots demonstrates female preference for one
large spot in Pseudotropheus lombardoi. J. Fish Biol. 60,
726–730. (doi:10.1111/j.1095-8649.2002.tb01696.x)
13 Ferno ¨, A. 1987 Aggressive behaviour between territorial
cichlids (Astatotilapia burtoni) in relation to rank and ter-
ritorial stability. Behaviour 103, 241–258. (doi:10.1163/
156853987X00189)
14 Balon, E. K. 1977 Early ontogeny of Labeotropheus Ahl,
1927 (Mbuna, Cichlidae, Lake Malawi), with a discus-
sion on advanced protective styles in fish reproduction
and development. Environ. Biol. Fish 2, 147–176.
(doi:10.1007/BF00005370)
15 Merila ¨, J. & Sheldon, B. C. 2000 Lifetime reproductive
success and heritability in nature. Am. Nat. 155,
301–310. (doi:10.1086/303330)
16 Burmeister, S. S., Jarvis, E. D. & Fernald, R. D. 2005
Rapid behavioral and genomic responses to social oppor-
tunity. PLoS Biol. 3, e363. (doi:10.1371/journal.pbio.
0030363)
17 Parikh, V. N., Clement, T. S. & Fernald, R. D. 2006
Androgen level and male social status in the African
cichlid, Astatotilapia burtoni. Behav. Brain Res. 166,
291–295. (doi:10.1016/j.bbr.2005.07.011)
18 Grosenick, L., Clement, T. S. & Fernald, R. D. 2007
Fish can infer social rank by observation alone. Nature
445, 429–432. (doi:10.1038/nature05511)
19 Hoegg, S., Boore, J. L., Kuehl, J. V. & Meyer, A. 2007
Comparative phylogenomic analyses of teleost fish Hox
gene clusters: lessons from the cichlid fish Astatotilapia
burtoni. BMC Genom. 8, 317. (doi:10.1186/1471-2164-
8-317)
20 Siegel, N., Hoegg, S., Salzburger, W., Braasch, I. &
Meyer, A. 2007 Comparative genomics of ParaHox clus-
ters of teleost fishes: gene cluster breakup and the
retention of gene sets following whole genome dupli-
cations. BMC Genom. 8, 312. (doi:10.1186/1471-2164-
8-312)
21 Salzburger,W.,Renn,S.C.P.,Steinke,D.,Braasch,I.,Hof-
mann, H. A. & Meyer, A. 2008 Annotation of expressed
sequence tags for the East African cichlid fish Astatotilapia
burtoni and evolutionary analyses of cichlid ORFs. BMC
Genom. 9, 96. (doi:10.1186/1471-2164-9-96)
22 Sanetra, M., Henning, F., Fukamachi, S. & Meyer, A.
2009 A microsatellite-based genetic linkage map of the
cichlid fish, Astatotilapia burtoni (Teleostei): a comparison
of genomic architectures among rapidly speciating
cichlids. Genetics 182, 387–397. (doi:10.1534/genetics.
108.089367)
23 Jennions, M. D., Møller, A. P. & Petrie, M. 2001 Sexually
selected traits and adult survival: A meta-analysis.
Q. Rev. Biol. 76, 3–36. (doi:10.1086/393743)
503–538.(doi:10.1163/
Variation in evolutional key trait
T. K. Lehtonen & A. Meyer2323
Proc. R. Soc. B (2011)
on June 27, 2011 rspb.royalsocietypublishing.orgDownloaded from

Page 7

24 Johnstone, R. A., Rands, S. A. & Evans, M. R. 2009
Sexual selection and condition-dependence. J. Evol.
Biol. 22, 2387–2394. (doi:10.1111/j.1420-9101.2009.
01822.x)
25 Rohwer, S. 1982 The Evolution of reliable and unreliable
badges of fighting ability. Am. Zool. 22, 531–546.
(doi:10.1093/icb/22.3.531)
26 Mulimbwa, N., Sarvala,J. & vander Knaap,M. 2010 The
effect ofintensivefishing ontwocichlids (Oreochromisnilo-
ticus and Astatotilapia burtoni) in Nyangara wetland at the
north end of Lake Tanganyika (D. R. Congo) with fishery
management implications. Aquat. Ecosyst. Health 13,
11–19. (doi:10.1080/14634981003627342)
27 Fernald, R. D. & Hirata, N. R. 1977 Field study of Hap-
lochromis burtoni: habitats and co-habitant. Environ. Biol.
Fish 2, 299–308. (doi:10.1007/BF00005997)
28 Lynch, M. & Walsh, B. 1998 Genetics and analysis of
quantitative traits. Sunderland, MA: Sinauer.
29 Brooks, R. & Kemp, D. J. 2001 Can older males deliver
the good genes? Trends Ecol. Evol. 16, 308–313. (doi:10.
1016/S0169-5347(01)02147-4)
30 Rowe, L. & Houle, D. 1996 The lek paradox and the cap-
ture of genetic variance by condition dependent traits.
Proc. R. Soc. Lond. B 263, 1415–1421. (doi:10.1098/
rspb.1996.0207)
31 Hert, E. 1991 Female choice based on egg-spots in Pseu-
dotropheus aurora Burgess 1976, a rock-dwelling cichlid of
Lake Malaw ˆi, Africa. J. Fish Biol. 38, 951–953. (doi:10.
1111/j.1095-8649.1991.tb03634.x)
32 Couldridge, V. C. K. & Alexander, G. J. 2001 Does the
time spent near a male predict female mate choice in a
Malawian cichlid? J. Fish Biol. 59, 667–672. (doi:10.
1006/jfbi.2001.1679)
33 Korzana, W. J., Robison, R. R., Zhao, S. & Fernald, R. D.
2008 Color change as a potential behavioral strategy.
Horm. Behav. 54, 463–470. (doi:10.1016/j.yhbeh.2008.
05.006)
34 Candolin, U. 2000 Increased signalling effort when survi-
val prospects decrease: male–male competition ensures
honesty. Anim. Behav. 60, 417–422. (doi:10.1006/anbe.
2000.1481)
35 Candolin, U. 2000 Male–male competition ensures
honest signaling of male parental ability in the three-
spined stickleback (Gasterosteus aculeatus). Behav. Ecol.
Sociobiol. 49, 57–61. (doi:10.1007/s002650000267)
36 Parker, T. H. & Ligon, J. D. 2002 Dominant male red
junglefowl (Gallus gallus) test the dominance status of
other males. Behav. Ecol. Sociobiol. 53, 20–24. (doi:10.
1007/s00265-002-0544-5)
37 Hill, G. E. 1999 Is there an immunological cost to caro-
tenoid-based ornamental coloration? Am. Nat. 154, 589–
595. (doi:10.1086/303264)
38 Grether, G. F. 1997 Survival cost of an intrasexually
selected ornament in a damselfly. Proc. R. Soc. Lond. B
264, 207–210. (doi:10.1098/rspb.1997.0029)
39 Godin, J.-G.-J. & McDonough, H. E. 2003 Predator pre-
ference for brightly colored males in the guppy: a viability
cost for a sexually selected trait. Behav. Ecol. 14, 194–
200. (doi:10.1093/beheco/14.2.194)
40 Pruden, A. J. & Uetz, G. W. 2004 Assessment of potential
predation costs of male decoration and courtship display
in wolf spiders using video digitization and playback.
J. InsectBehav.
17, 67–80.
0000025133.54118.1e)
41 Langerhans, R. B., Layman, C. A. & DeWitt, T. J. 2005
Male genital size reflects a tradeoff between attracting
mates and avoiding predators in two live-bearing fish
species. Proc. Natl Acad. Sci. 102, 7618–7623. (doi:10.
1073/pnas.0500935102)
42 Hurtado-Gonzales, J. L. & Baldassarre, J. A. C. 2010
Interaction between female mating preferences and pre-
dation may explain the maintenance of rare males in
the pentamorphic fish Poecilia parae. J. Evol. Biol. 23,
1293–1301. (doi:10.1111/j.1420-9101.2010.01995.x)
43 Rosenthal, G. G., Flores Martinez, T. Y., Garcı ´a de
Leo ´n, F. J. & Ryan, M. J. 2001 Shared preferences by
predators and females for male ornaments in swordtails.
Am. Nat. 158, 146–154. (doi:10.1086/321309)
44 Basolo, A. L. & Wagner Jr, W. E. 2004 Covariation
between predation risk, body size and fin elaboration in
the green swordtail, Xiphophorus helleri. Biol. J. Linn.
Soc.
83,87–100.(doi:10.1111/j.1095-8312.2004.
00369.x)
45 Meyer, A., Salzburger, W. & Schartl, M. 2006 Hybrid
origin of a swordtail species (Teleostei: Xiphophorus
clemenciae) driven by sexual selection. Mol. Ecol. 15,
721–730. (doi:10.1111/j.1365-294X.2006.02810.x)
46 Siepielski, A. M., DiBattista, J. D., Evans, J. A. &
Carlson, S. M. 2011 Differences in the temporal
dynamics of phenotypic selection among fitness com-
ponents in the wild. Proc. R. Soc. B 277, 1572–1580.
(doi:10.1098/rspb.2010.1973)
47 Blows, M. W. & Hoffmann, A. A. 2005 A reassessment of
genetic limits to evolutionary change. Ecology 86, 1371–
1384. (doi:10.1890/04-1209)
48 Carlson, S. M. & Seamons, T. R. 2008 A review of quan-
titative genetic components of fitness in salmonids:
implications for adaptation to future change. Evol. Appl.
1, 222–238. (doi:10.1111/j.1752-4571.2008.00025.x)
49 Bakker, T. C. M. 1999 The study of intersexual selection
using quantitative genetics. Behaviour 136, 1237–1265.
(doi:10.1163/156853999501748)
50 Qvarnstro ¨m, A., Brommer, J. E. & Gustafsson, L. 2006
Testing the genetics underlying the co-evolution of mate
choice and ornament in the wild. Nature 441, 84–86.
(doi:10.1038/nature04564)
51 Karino, K. & Haijima, Y. 2001 Heritability of male sec-
ondary sexual traits in feral guppies in Japan. J. Ethol.
19, 33–37. (doi:10.1007/s101640170015)
52 Endler, J. A. 1982 Convergent and divergent effects of
natural selection on colour patterns in two fish faunas.
Evolution 36, 178–188. (doi:10.2307/2407979)
53 Andersson, M. 1994 Sexual selection. Priceton, NJ:
Priceton University Press.
(doi:10.1023/B:JOIR.
2324T. K. Lehtonen & A. Meyer
Variation in evolutional key trait
Proc. R. Soc. B (2011)
on June 27, 2011 rspb.royalsocietypublishing.org Downloaded from