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Evolutionary developmental biology
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UV wavelengths experienced during
development affect larval newt visual
sensitivity and predation efficiency
Research
Mélissa Martin1,2, Marc Théry1, Gwendolen Rodgers1, Delphine Goven3,
Stéphane Sourice2,4, Pascal Mège1,2 and Jean Secondi2,4
Cite this article: Martin M, Théry M, Rodgers
G, Goven D, Sourice S, Mège P, Secondi J. 2016
UV wavelengths experienced during
development affect larval newt visual
sensitivity and predation efficiency. Biol. Lett.
12: 20150954.
http://dx.doi.org/10.1098/rsbl.2015.0954
Received: 13 November 2015
Accepted: 7 January 2016
Subject Areas:
developmental biology, ecology,
evolution, behaviour
Keywords:
ultraviolet vision, development, SWS1 opsin
gene, foraging, Lissotriton vulgaris, water
transmission
Author for correspondence:
Jean Secondi
e-mail: [email protected]
Electronic supplementary material is available
at http://dx.doi.org/10.1098/rsbl.2015.0954 or
via http://rsbl.royalsocietypublishing.org.
1
UMR 7179 CNRS-MNHN, Mécanismes Adaptatifs et Evolution, Brunoy, France
GECCO, 3UPRES EA 2647/USC INRA 1330, Récepteurs et Canaux Ioniques Membranaires, and 4UMR 6554
Littoral, Environnement, Télédétection, Géomatique, Université d’Angers, France
2
MM, 0000-0002-4496-2450; JS, 0000-0001-8130-1195
We experimentally investigated the influence of developmental plasticity of
ultraviolet (UV) visual sensitivity on predation efficiency of the larval
smooth newt, Lissotriton vulgaris. We quantified expression of SWS1 opsin
gene (UV-sensitive protein of photoreceptor cells) in the retinas of individuals
who had developed in the presence (UVþ) or absence (UV2) of UV light
(developmental treatments), and tested their predation efficiency under
UVþ and UV2 light (testing treatments). We found that both SWS1 opsin
expression and predation efficiency were significantly reduced in the UV2
developmental group. Larvae in the UV2 testing environment displayed
consistently lower predation efficiency regardless of their developmental treatment. These results prove for the first time, we believe, functional UV vision
and developmental plasticity of UV sensitivity in an amphibian at the larval
stage. They also demonstrate that UV wavelengths enhance predation efficiency and suggest that the magnitude of the behavioural response depends
on retinal properties induced by the developmental lighting environment.
1. Introduction
Many animals possess ultraviolet (UV, 300–400 nm) vision and use UV cues
(reflectance or absorbance) from food items or the environment while foraging
for food (reviewed in [1]). When UV cues are not accessible, animals’ foraging
efficiency is directly negatively affected [2,3]. A recent study demonstrated that,
at least in zooplankton predators, UV vision allows location of UV-absorbing
food items at greater distances and angles [4].
Variation in lighting environment directly affects development of visual systems [5]. Development in UV-poor environments results in a decreased
expression of SWS1 opsin protein, the UV-sensitive molecule of retinal photoreceptor cells [6,7]. Differential expression of the opsin gene should result in
differences in different cone type densities or in visual pigments within
cones. This could impact colour discrimination [8], visual sensitivity and
speed of phototransduction [9]. Interestingly, in the bluefin killifish, developmental plasticity in the retina induces variations in how individuals perceive
colours in different lighting environments [10]. The critical question is whether
such plasticity also has implications for fitness-related behaviours like foraging.
This is a major issue for freshwater-foraging UV-sensitive species because
they frequently experience heterogeneous UV transmission conditions owing
to strong spatial and temporal variation in UV-absorbing organic molecules
[11]. Here, we investigated the importance of UV vision plasticity on foraging
efficiency in the larval stage of the smooth newt, Lissotriton vulgaris, a species
that breeds and develops in lentic habitats. Larvae are aquatic and feed on
& 2016 The Author(s) Published by the Royal Society. All rights reserved.
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relative LWS mRNA quantity (%)
6
4
70
60
Biol. Lett. 12: 20150954
relative SWS1 mRNA quantity (%)
8
50
40
2
DUV+
DUV–
DUV+
DUV–
Figure 1. Percentage relative mRNA quantity of SWS1 and LWS opsin genes for newt larvae that developed in UVþ and UV2 lighting conditions. The individual
points are outliers.
UV-absorbing zooplankton, whereas adults have a diversified carnivorous diet and alternate aquatic and terrestrial
phases. Adults possess a retina with photoreceptor cells sensitive to UV, short and long wavelengths [12], and use UV
signals in sexual selection [13].
We tested (i) whether the SWS1 opsin gene is expressed in
the retina of newt larvae, (ii) whether UV light conditions
experienced during development modulate SWS1 expression,
and (iii) whether the effects of immediate lighting environment and developmental plasticity interact to affect
foraging efficiency. To do so, we reared larvae from eggs in
the presence (UVþ) or absence (UV2) of UV light (developmental environments), measured the expression of the SWS1
gene, and assessed predation efficiency of each individual
under both UVþ and UV2 light (testing environments).
We predicted lower SWS1 expression level and predation
efficiency in the UV2 developmental group as well as
lower predation efficiency in the UV2 testing environment
regardless of the developmental group.
(b) Predation tests
At the final stage before metamorphosis (i.e. 58 – 64 days after
hatching), visual foraging performance of 29 larvae from 11
females was tested. Tests took place in the darkroom with the
lighting system described above after a 48 h fasting period.
Before the test began, each larva was placed in a Petri dish
(40 mm diameter 17.4 mm height, 1 cm water column) under
the neon light and, after 9 min of habituation, the light was
turned on. One minute later, eight large Daphnia magna were
gently introduced in the Petri dish and the test begun. To optimize detectability and attractivity of prey, daphnia had been
preliminarily fed with a lutein – yeast solution to enhance their
reflectance in the visible and UV ranges (details in the electronic
supplementary material, S3 and S4; [3]). The number of remaining live prey items was recorded every 5 min for 90 min. Each
larva was tested twice, once under UVþ visible light (UVþ
testing group, TUVþ) and once under visible light (UV2 testing
group, TUV2) in a random presentation order, with a minimum
2 day interval between tests.
(c) Quantification of opsin gene expression
2. Material and methods
(a) Sampling, rearing conditions and lighting
treatments
Larvae from 11 gravid female L. vulgaris captured on April 2012
in a pond near Angers (western France) were reared from eggs
laid in a UV-free lighting environment (see details in the electronic supplementary material, S1). From hatching, each larva
was exposed daily to 15 min supplementary lighting. Half the
larvae from each female were exposed to UV þ visible light
(UVþ developmental group, DUVþ), while the other half was
exposed to visible light only (UV2 developmental group,
DUV2). To do so, rearing containers were moved into a darkroom
equipped with a UV-B neon light (Repti Glo 5.0 UVB/T8, Exo
Terraw), located at 20 cm above the water surface. This light
was left uncovered for the DUVþ group, or covered with a
UV-blocking filter (3 mm layer of clear DuraLarTM polyester
film, Grafixw) for the DUV2 group (see electronic supplementary
material, S2, for spectra of lighting conditions).
After behavioural tests, we selected two larvae at the final larval
stage, one from each developmental group, from nine females
(for two females, all larvae of one group had metamorphosed
before analysis). After euthanasia and enucleation, relative
opsin mRNA expression of SWS1 opsin gene in the eye was
quantified using methods described by Dupuis et al. [14] and
Pfaffl et al. [15] (see electronic supplementary material, S5 and
S6 for method details). In order to ensure that experimental conditions affected SWS1 opsin expression only, we also quantified
the long-wavelength-sensitive (LWS) opsin expression.
(d) Statistical analyses
All analyses were carried out using R v. 3.1.1 [16]. The differences in relative mRNA expression levels of SWS1 and LWS
genes between the two developmental treatments were tested
using paired Wilcoxon tests. To analyse predation tests, we
tested the effects of developmental and testing treatments and
their interaction on daphnia survival using a mixed-effects Cox
model (COXME package [17]), with larva identity nested within
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prey survival probability
1.0
0.8
0.6
0.4
0
0
20
40
60
80
time (min)
Figure 2. Survival probability curves of daphnia prey depending on UVþ and UV2 lighting conditions during the larval development (D) and test (T).
female identity as a random factor. Test order was introduced as
covariate. Surviving daphnia were treated as right-censored (i.e.
still alive) cases. Analysis began with a full model including all
experimental effects as fixed effects. The minimum adequate
model was obtained using likelihood ratio tests (LRTs).
3. Results
Developmental treatment strongly affected SWS1 gene
expression ( paired Wilcoxon test, V ¼ 45, N ¼ 9, p ¼ 0.004)
but not LWS opsin gene expression (V ¼ 35, N ¼ 9, p ¼ 0.16,
figure 1). UV light deprivation during development resulted
in a 53.1% average decrease in SWS1 opsin expression at the
final larval stage.
Predation tests reveal that daphnia prey survival
was significantly affected by both developmental (x 2 ¼ 6.59,
p ¼ 0.010) and testing treatment (x 2 ¼ 10.83, p ¼ 0.001). Survival probability of daphnia was 71.1 + 2.7% higher in
tests with a DUV2 newt predator than in tests with a DUVþ
newt (figure 2). Regardless of developmental treatment,
daphnia survival was 46.6 + 1.4% higher during TUV2 tests
than during TUVþ tests (figure 2; see also the electronic
supplementary material, S7, for a representation of the
inter-individual variation of the total number of eaten prey
at the end of the test). Prey survival did not depend on test
order (TUVþ or TUV2 in first test, LRT: x 2 ¼ 0.14, p ¼ 0.91).
UV light conditions experienced during development
can affect behaviour [10,20]. For example, bluefin killifish
reared in a UV-rich environment shifted colour-guided foraging preferences when tested in a UV-poor environment,
whereas those reared in a UV-poor environment did not
shift when tested in a UV-rich environment [10]. By contrast,
we found additive, not interactive, effects of the immediate
and developmental lighting environment on predation efficiency in newts. Regardless of developmental group, prey
survival was higher in the UV2 testing environment, meaning that, as in fishes [2–4], the use of UV cues enhances
predation efficiency in newt larvae. We also found that
UV-deprived larvae foraged less efficiently than larvae
reared with UV light. Thus, developing in UV-poor environments such as coloured water strongly reduces the
performance of visually guided behaviours like foraging,
which potentially negatively affects fitness components at
the larval or later stage.
Overall, this study suggests that the magnitude of visually guided behavioural response depends on the retinal
properties induced by the developmental lighting environment. We can hypothesize that early UV light exposure is
required for the proper development of UV-sensitive visual
systems; otherwise the performance of visually guided
tasks can be jeopardized. Thus, UV transmission properties
of the environmental medium might be an unrecognized
but important factor for the selection of breeding sites in
UV-sensitive animals.
4. Discussion
We found low, but substantial levels of SWS1 gene expression in
the eyes of larvae, of the same order as those found in several fish
species (e.g. [11,18]). This is the first demonstration to our knowledge that UV cones are present and functional in the retina of an
amphibian at the larval stage. In addition, we demonstrated
developmental plasticity in the expression of the SWS1 gene.
Expression of SWS1 was halved in larvae deprived of UV light
during development, whereas LWS expression remained
unchanged. SWS1 opsin was still expressed in UV-deprived
larvae, which suggests that its expression is partly genetically
determined. Our results are consistent with other findings in
vertebrates [6,7,18], and support the idea that the visual
system is plastic and changes with the quantity and quality of
wavelengths available in the environment [19].
Ethics. All experiments were conducted under the approval of the
Préfecture de Maine-et-Loire ( permit 04/2012) and in accordance
with the current laws in France.
Data accessibility. Data are available at Dryad: http://dx.doi.org/10.
5061/dryad.444s1.
Authors’ contribution. J.S., G.R., S.S., M.T. conceived and designed the
study. G.R., S.S., J.S. performed the behavioural experiment. M.M.,
G.R., J.S. analysed data. D.G. carried out the genetic labwork. P.M.
developed primers. M.M., J.S., G.R., M.T., D.G., S.S., P.M. drafted
and revised the manuscript. All authors gave final approval for publication and agree to be held accountable for the work performed.
Competing interests. Authors have no conflict of interests.
Funding. This study was funded by the Agence National de la
Recherche grant no. ANR-2011-BSV7-001 to M.T.
Acknowledgements. We thank Mark Briffa and two anonymous referees
for their constructive comments on the manuscript.
Biol. Lett. 12: 20150954
DUV– TUV–
DUV– TUV+
DUV+ TUV–
DUV+ TUV+
0.2
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