Species-specific effects of single sensillum ablation on mating position in Drosophila
1 UMR-CNRS 5548, Faculté des Sciences, Université de
Bourgogne, 6 Boulevard Gabriel, F-21000 Dijon, France
2 School of Biological Sciences, University of Manchester, Oxford Road,
Manchester, M13 9PL, UK
* Author for correspondence: (e-mail: jean-francois.ferveur{at}u-bourgogne.fr)
Accepted 3 June 2003
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Summary |
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Key words: lock and key hypothesis, sexual selection, mechanoreceptor, copulation, fruit fly, Drosophila melanogaster
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Introduction |
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This is particularly apparent in the case of the Diptera, which show the
widest range of mating positions of any pterygote insect, with at least ten
recorded positions (Alexander,
1964; McAlpine,
1981
). Species-specific mating positions, which are frequently
constant within a given genus, are thought to be driven by two factors:
ecological selective pressures relating to the evolution of courtship and
mating on a substrate rather than in the air
(McAlpine, 1981
) and, in some
species, the twisting of the final segments ('hypopygium') of the Dipteran
male abdomen by 180° or even, bizarrely, by 360° in the Cyclorrhapha,
which makes flexion of the final segments easier to effect, like a twist in a
long balloon (Bickel, 1990
). In
virtually all Dipteran species, the initial coupling position involves both
male and female facing in the same direction; the pair may then make
synchronised movements to arrive at the species' typical final mating
position. These movements, and indeed mating itself, require both partners to
be able to detect both their partner's position and their own
(proprioception). The most likely sources of this positional information are
the genital sensilla. In crickets, for example, mechanoreceptors on the male
cerci detect the female's position and form part of the neural network
underlying spermatophore transfer (Sakai
et al., 1991
; Snell and
Killian, 2000
).
Functional studies of insect male external genital structures are rare
(Eberhard, 1993,
2001
) and there have been no
experimental studies of Dipteran mating position. To investigate the sensory
mechanics of insect mating, we studied three closely related species of
fruitfly, Drosophila melanogaster, D. simulans and D.
sechellia, all of which mate in the classic Cyclorrhaphan position with
the male on the female's back and both partners facing the same way, following
substrate-based courtship behaviours. Once mating has taken place, there is no
subsequent change in position. Males of these species show specific
differences in their genital structures, and in particular the shape of the
claspers (CLs), the lateral plates (LPs) and the genital arch
(Ashburner, 1989
). In all three
species the CLs and the LPs are covered with similar sex-specific
mechanoreceptor sensilla, which in D. melanogaster show afferent
projections to different levels of the abdominal ganglion
(Taylor, 1989
). To discover
whether the information encoded by these sensilla is related to mating, they
were ablated and the male's resultant behaviour was observed. The results were
surprising.
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Materials and methods |
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Sensilla ablation and scanning electron microscopy
Flies less than 2 h old were sexed under light CO2 anaesthesia
and subsequently kept in food vials until the behavioural test. Males were
kept individually, and females were placed in groups of five. 1 day before the
behavioural assay, 3- to 4-day-old males were CO2 anesthetized and
immobilized in a pipette tip with their abdomen protruding. Bristles were
removed from the claspers or from the lateral plates with fine forceps (ref.
9980, Moria, France). Operated males were isolated in a fresh food vial until
the next day when their behaviour was studied. Micrographs of male genitalia
from control and ablated flies were taken using a scanning electron microscope
(ESEM XL30; Philips, Eindoven, The Netherlands).
Behavioural tests
Each male (4 or 5 days old) was tested once with a virgin intact female (3
days old) in a circular observation chamber (2.8 cm diameter, 0.5 cm height).
Single pairs were observed for 60 min
(Savarit et al., 1999). In the
case of hybrid males, however, which were difficult to obtain, behaviour was
monitored with an alternative procedure. Instead of pairs of flies confined in
a small mating chamber, 3-5 hybrid males (3- to 5-day-old control or operated
flies) were placed with 10-20 females (2 days old) in a Petri dish (5 cm
diameter), and their behaviour was monitored for 120 min. Moreover, each
hybrid male was tested on two following days consecutively with D.
melanogaster and with D. simulans females (or vice
versa). All tests were performed at 25±0.5°C, at 65±5%
humidity. In all cases, we recorded the time at which copulation occurred
(copulation latency) and its duration. We noted the total number of mating
pairs, and from that figure we calculated the frequency of abnormal and normal
mating positions. Digital images of mating position were taken using a MZ8
binocular microscope (Leica, Munich, Germany) connected to a digital video
cassette recorder (Panasonic AJ-D230E, Tokyo, Japan) coupled to a colour
digital video camera (Sony SSC-DC38P, Tokyo, Japan). Flies were placed in
specially designed plastic cells (internal dimensions: 16 mm x 12 mm
x 4 mm).
Statistics
For each series of experiments, the significance level is indicated in the
corresponding table. Mating frequency was tested by a 2-test.
The data for copulation latency, which were not normally distributed, were
logarithmically (ln) transformed prior to being tested with an analysis of
variance (ANOVA). Data obtained for copulation duration were normally
distributed and were compared by means of an ANOVA.
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Results |
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... But not in all species
The shapes of the claspers on D. simulans and D.
sechellia males are different from those of D. melanogaster, but
both species possess a pair of long sensilla located at a similar position to
CLlbs in D. melanogaster males
(Tsacas et al., 1971;
Tsacas and Bachli, 1981
)
(Fig. 1E,F), suggesting that
these structures are homologous in all three species. Surprisingly, unilateral
and bilateral ablation of CLlbs in D. simulans and D.
sechellia males had no effect on mating posture, courtship success as
measured by mating frequency or copulation latency in either species
(Table 2). These results
contrast with those obtained for D. melanogaster, which demonstrates
interspecies differences in the sensory systems underlying mating and mating
position and may indicate that these genital sensilla have different functions
in the three species. The CLlbs are not involved in maintaining mating posture
in D. simulans and D. sechellia; other structures presumably
are involved.
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To investigate the underlying genetic control of this effect, interspecific hybrid males were produced by crossing D. melanogaster males with D. simulans females. Hybrids showed a single pair of long clasper bristles, as in both parental species. Hybrid males were observed with virgin females of both parental species. Unilateral ablation of CLlb sensilla in hybrid males produced unambiguous results, although only a few pairs copulated (Table 2): like their D. simulans parents, operated hybrid males adopted a normal mating posture with females of both species, showing that the control of mating posture does not depend upon female type. No systematic differences were found for other mating or courtship features.
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Discussion |
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The mechanisms by which these effects occur was further investigated, with
the most striking result for mating position. Given the mechanosensory nature
of the bristles, one possibility is that the male uses proprioceptive
information to determine his position on the female, perhaps coded
via the sexually dimorphic neurons and axonal arborisation that have
been observed in the central nervous system
(Taylor, 1989). In the absence
of such information, he may incline his position as far as is possible, or
until he obtains similar information from other sensory sources. It is also
possible that the female reacts to the absence of information normally
provided by these bristles, and changes her behaviour accordingly, leading to
the pair adopting a lateral mating position.
The effects on mating frequency are more complex. Ablation may have affected the quantity or the quality of male courtship. In particular, during the final stages of courtship, males of all three species sometimes make what appear to be attempts at copulation. Failure to detect the female, or to provide her with appropriate stimuli, or both, may lead to a decrease in the frequency with which these attempts at copulation were transformed into successful matings, as shown by lower mating frequencies observed in bilaterally ablated flies. Similar effects may produce the increased mating latency observed in unilaterally ablated flies. These sensilla may therefore play a role in both mating and the final, decisive stages of courtship.
Surprisingly, ablation of the apparently homologous sensilla in D.
simulans and D. sechellia did not produce the same effect:
ablation of the CLlbs had no effect on mating position in these species. There
are several possible explanations for this interspecific difference: the
sensilla may have different functions, the neural networks to which they are
connected may have different functions, female criteria may show interspecific
differences or males may be differentially able to compensate for the absence
of specific stimuli from their genitalia. Whatever the precise interaction
that is taking place here, this result is of fundamental importance for
studies that attempt to interpret the evolution of insect genitalia in terms
of either the `lock and key' hypothesis or sexual selection
(Arnqvist, 1998;
Alexander et al., 1997
). It
suggests that from a functional point of view rather than from a simple
anatomical standpoint, the diversity of insect genitalia may be greater than
hitherto suspected. Furthermore, the mating pair must be considered as an
interacting pair, and not an active male with a passive female. Stimulation,
response and interactions are taking place during courtship and mating.
Functional studies will be required to make real progress in this field, as
apparently identical structures in closely related species can form sensory
systems with radically different functional characteristics.
It is striking that many other species of Diptera show similar long
bristles on the male's claspers (e.g. Diopsidae, Mycetophilidae, Sciardiae and
Tanypezidae (McAlpine et al.,
1981,
1987
). This suggests that the
effects observed here might apply to all Diptera and that changes in the
detection or expression of sensory information in either or both sexual
partners may have contributed to the changes in mating position that have
taken place in Dipteran species. For example, some Empid flies couple with the
male on the female's back, then during the course of mating the male bends his
genitalia down and to one side of the female
(McAlpine, 1981
). This clearly
requires proprioceptive information of the type that is apparently coded by
the male clasper long bristles in D. melanogaster. The decrease in
mating frequency shown by unilaterally ablated males suggests, however, that
any mutant lacking that particular mechanoreceptor would be at a clear
selective disadvantage. Coadaptive change by both the sender and receiver of
the information associated with these mechanoreceptors would clearly be
necessary to ensure the adoption of a new mating position by such a
mechanism.
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Acknowledgments |
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