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First published online November 14, 2008
Journal of Experimental Biology 211, 3698-3702 (2008)
Published by The Company of Biologists 2008
doi: 10.1242/jeb.021980
Better mate in the shade: enhancement of male mating behaviour in the cabbage butterfly, Pieris rapae crucivora, in a UV-rich environment
1 Behavioural Biology, Department of Agriculture, Tokyo University of
Agriculture and Technology, Fuchu, Tokyo 183-8509, Japan
2 Graduate School of Integrated Science, Yokohama City University, 22-2 Seto,
Kanazawa-ku, Yokohama 236-0027, Japan
3 Laboratory of Neuroethology, The Graduate University for Advanced Studies
(Sokendai), Shonan Village, Hayama 240-0193, Japan
* Author for correspondence (e-mail: arikawa{at}soken.ac.jp)
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Summary |
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 TOP Summary INTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONReferences |
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Key words: mate recognition, UV vision, photoreceptor, compound eye, colour vision, wing reflection spectra
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INTRODUCTION |
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TOPSummaryINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONReferences |
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;
Kelber et al., 2003). UV light
is often indispensable for certain behaviours, such as foraging
(Barth, 1982;
Church et al., 2001), mate
recognition (Obara, 1970;
Silberglied and Taylor, 1973)
and mate choice (Rutowski,
1985; Bennett et al.,
1996). However, little is known about whether and how these
UV-related behaviours are affected by the temporal as well as local variations
of UV conditions in the field. For example, the relative content of UV light
is higher in shaded places than in direct sunlight because of scattering
(Kok, 1972).
The Japanese subspecies of cabbage butterfly, Pieris rapae
crucivora, is subjected to such changes in the UV environment. Its eyes
contain at least six classes of spectral receptors, including UV receptors
(Wakakuwa et al., 2006), and
the males and females distinctly differ in wing reflectance in both the UV and
visible wavelength range (Giraldo and
Stavenga, 2007). The males visually recognize the females as
potential mates based on the UV reflection of the wings
(Obara and Hidaka, 1968;
Obara, 1970). It is therefore
probable that the visibility of female wings for males varies with the weather
conditions or local light environment, i.e. depending upon whether the females
are exposed to the sun or are in the shade. This raises the question of
whether the light environment affects the UV-guided mating behaviour of males
that compete for access to virgin females
(Ohsaki, 1980;
Suzuki, 1980;
Thornhill and Alcock, 1983).
To answer this we first confirmed that the relative UV content is much higher
in shaded than in sunlit places, and investigated whether and how the UV
content affects the mating behaviour of male Pieris rapae
crucivora.
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MATERIALS AND METHODS |
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TOPSummaryINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONReferences |
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Spectral measurements
A calibrated spectrophotometer (HSU-100S, Asahi Spectra) was used to
measure spectra. The spectral composition of sunlight was measured with a
MgO-coated reflection standard placed on a table in the field on a fine day.
The reflection standard was shaded with cardboard when measuring the spectrum
in the shade. The reflection spectra of wings of Pieris rapae
crucivora were measured with the same method using an isolated hind wing
of a freshly emerged female, ventral side up, instead of the reflection
standard. The transmittance spectra of UV-opaque and UV-transparent plastic
sheets (see below) were measured with the same spectrophotometer.
Behavioural experiments
Experiment 1: large cage experiments
We tested the female-locating behaviour of males in the sunlight and in the
shade in a large outdoor cage measuring 9 mx9 mx2 m (experiment
1). Females that were presented to males were freshly emerged individuals
killed by freezing, wings closed, and laid flat with the ventral side of the
wings up. Such female dummies effectively elicit mating behaviour of males,
which even respond to a piece of paper that reflects UV
(Obara and Hidaka, 1968;
Obara, 1970). To stimulate
mating behaviour even further, we added cabbage plants as well as buckwheat
plants as nectar sources.
In the first experiment (experiment 1a), conducted in the large cage, we placed two female dummies laid flat, 15 cm apart, in the centre of a piece of black cardboard measuring 25 cmx30 cm (Fig. 1A). One dummy was shaded by placing a 60 cmx60 cm wooden board 1.8 m above the ground. The black cardboard with the female dummies was rotated every 15 min by 180° to eliminate any possible differences due to local factors. In this experimental arrangement, we assumed that the males visually compared the two female dummies at the same time.
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In each experiment, we released about 280 virgin males in the cage, and observed their mating behaviour to the female dummies. The observed behaviours included: approach to the dummies as close as about 5 cm, contact with the dummies, and copulatory attempt with the dummies. We counted the number of males that expressed any one of these behaviours. The counted males were immediately removed from the cage during or after the behaviour; consequently, the data obtained are all derived from males naïve to the experiment.
We carried out experiment 1a from 07:15 h to 08:15 h on a sunny day in mid
summer (16th July, 2004) and experiment 1b from 08:30 h to 11:00 h on the same
day. We designed this procedure based on the previous findings that the males
start searching females around 06:30 h on a sunny day and that the number of
female-searching males increases over time, peaking between 09:00 h and 11:00
h; at the peak, nearly three times more male flies are searching for females
(Hirota and Obara, 2000).
Experiment 2: open field experiment
We designed experiment 2 to test whether and how the male mating behaviour
was affected by a varying UV environment. Here we observed the male mating
behaviour in the presence or absence of UV. For this purpose, we covered an 8
mx8 m area of a cabbage field (20 mx30 m) with a UV-opaque plastic
sheet (Cut-Ace-Clean, MKV Platech, Tokyo Japan), which transmits visible light
but not UV light (Fig. 1C). We
covered another 8 mx8 m area of the same field similarly with a
UV-transparent plastic sheet (Clean-Ace-Daichi, MKV Platech, Tokyo Japan;
Fig. 1C). The two areas were 2
m apart and we hereafter call these areas the UV-absent and the UV-present
areas, respectively.
The sheets were placed 1.4 m above the ground, and because there were no walls, the butterflies could freely move into and out of the areas; the temperatures of these areas were virtually the same (30.5–31.5°C and 30.0–31.0°C in the UV-present and -absent areas, respectively). In an `inter-generations period', when wild individuals were virtually absent, we released about 300 fresh males into the cabbage field in the evening 1 day before the experiment. The released males stayed around the cabbage field for several days, thus allowing observation of their mating behaviour for an extended period. The experiment was carried out from 08:00 h to 12:00 h on 22 July, 2005.
In the morning of the next day, the released males started to search for females by flying zigzag among the cabbage leaves, although there were no females. The time of female-searching behaviour was measured by visually following a flying male and recording the time the male entered the UV-present area, apparently then searching for females, and subsequently recording the time the male left the area. Some males took up a resting position on a substrate within the area and stayed there for a prolonged period. In such cases, we concluded that the males were no longer performing the female searching behaviour. After the male left the area or alighted on a substrate, we repeated the same observation in the UV-absent area. We performed these observations in the two areas alternately, and thus recorded in each area the behaviour of total 79 cases (not necessarily 79 individuals, because some individuals may have flown a number of times in and out of the areas).
Experiment 3: small arena experiment
To measure the frequency of actual copulation, we carried out experiment 3.
We prepared two frames, each measuring 45 cmx5 cmx55 cm, one with
the ceiling and four walls covered with the UV-opaque plastic sheet, and the
other covered with the UV-transparent plastic sheet
(Fig. 1C). We hereafter refer
to the former and the latter as the UV-absent and UV-present arenas,
respectively. We put 47 males and 64 females in each arena, and counted the
number of mated pairs in a period of 75 min. The arenas were placed in the
shade to avoid excessive warming. The experiment was carried out from 12:30 h
to 13:45 h on the same day as experiment 2.
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RESULTS |
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TOPSummaryINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONReferences |
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Experiment 1: higher detectability of females in the shade
In Experiment 1, where males were allowed to choose between sunlit and
shaded dummies, males significantly preferred the female dummy placed in the
shade. When the dummies were placed 15 cm apart in experiment 1a
(Fig. 3A), a total of 21 males
responded, 17 of which responded to the female in the shade, whereas four
responded to the female in the sun (
2=8.05,
P<0.05). When the dummies were placed 2 m apart in experiment 1b
(Fig. 3B), 128 males responded:
80 responded to the dummy in the shade, and 48 to the dummy in direct sunlight
(2=8.00, P<0.05). The number of responding males
in experiment 1b was larger, because the experiment was conducted for a longer
period than experiment 1a and also later in the morning, when more males were
actively searching females (Hirota and
Obara, 2000).
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Experiment 2: active female search in the UV-rich environment
In experiment 2, 76 of 79 males in the UV-present area performed active
zigzag flight and left the area while continuing the search for females; only
three males alighted on a substrate within the area and stopped the search for
females (Fig. 4A). In the
UV-absent area, 58 of 79 males performed active zigzag flight, whereas 21
males sat on a substrate within the area and stopped the search behaviour
(2=14.20, P<0.001). However, the average duration
of the female-searching flight was virtually the same in the UV-present (mean
± s.d.; 16.7±9.1 s, N=76) and the UV-absent areas
(16.7±12.1 s, N=58, Mann-Whitney U-test,
P=0.587).
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Experiment 3: enhanced mating in the UV-rich environment
Within 75 min after introducing the individuals, 28 pairs successfully
copulated in the UV-present arena, whereas only eight pairs copulated in the
UV-absent arena (Fig. 4B;
2=16.25, P<0.001).
Fig. 4C presents the reflection
spectra of female wings in two arenas set in the shade. The wings in the
UV-absent area reflect very little UV.
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DISCUSSION |
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TOPSummaryINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONReferences |
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) (Fig. 2A,B),
and revealed for the first time that the relative UV reflection of female
wings is much higher in the shade, although the absolute UV reflection is 10
times stronger in the sunlight (Fig.
3A,B). The illumination-dependent difference in the relative UV
reflection of female wings affected the mating behaviour of male Pieris
rapae crucivora: female-searching males responded more frequently to
female wings placed in the shade (Fig.
3A,B).
The possibility that males prefer the shade itself is considered unlikely,
as in experiment 1 carried out in a large cage, the males were allowed to
visually compare two dummies while flying in the direct sunshine (cf.
Fig. 1), particularly when the
dummies were placed only 15 cm apart. Moreover, in experiment 2, carried out
in an open field, more males searched for females actively in the UV-present
area (Fig. 4A), which must be
brighter than the UV-absent area for Pieris, because their eyes are
furnished with a number of photoreceptors that are sensitive in the UV
wavelength region (
400 nm) (Qiu and
Arikawa, 2003; Arikawa et al.,
2005).
The males prefer to locate mates in the shade and respond more readily to
shaded females. The possible reasons for this include that the shaded females
are more visible to the males. Also this is reasonable from the adaptive point
of view. Newly emerged virgin females stay for a few hours on the underside of
cabbage leaves, where they are shaded from direct sunlight
(Hirota et al., 2001). This may
promote female location by males and account for the facts that most females
copulate before their initial flight after emergence, and that females flying
in sunshine are most likely already mated and refuse approaching males by the
characteristic mate refusal posture
(Obara, 1964).
In addition, the UV environment also seemed to affect the male mating behaviour. UV-rich environments promoted the female-searching behaviour (Fig. 4A) and copulation (Fig. 4B). Such behavioural plasticity dependent on the UV environment will contribute to the enhanced mating success of males and therefore be favoured by natural selection.
Visual mechanisms underlying male mating behaviour
Most probably, the females are more visible when staying in the shade. The
richness of UV in the shade significantly enhances the UV reflection of female
wings (Fig. 3D), which
effectively stimulates the UV, DB and B receptors, i.e. the receptors with
pronounced sensitivity in the wavelength region shorter than 400 nm
(Qiu and Arikawa, 2003). As in
other lepidopteran species (Kelber and
Henique, 1999; Kelber and
Pfaff, 1999; Kinoshita et al.,
1999; Kelber et al.,
2002), Pieris rapae also has colour vision (K.A. and Y.
Nakatani, manuscript in preparation), which is the ability to discriminate
visual stimuli based on the spectral contents regardless of the brightness.
Therefore, the males of Pieris must be able to detect different
reflection spectra as different colours. In fact, males preferred females with
stronger UV reflection (summer form) to those with weaker UV reflection
(spring/autumn form), both placed in direct sunlight
(Obara et al., 2008). In the
present case, the males selected the shaded female wings in preference to
those under the sun even though the shaded ones were 10 times darker
(Fig. 3A). Such discriminations
must be achieved by their colour vision. Note that the DB receptor, which has
a secondary sensitivity band peaking at 380 nm, is male-specific
(Arikawa et al., 2005). Whether
this receptor has any crucial function or not in mate detection is an
interesting topic for further study.
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Acknowledgments |
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