72 (1): 11 – 21
23.4.2014© Senckenberg Gesellschaft für Naturforschung, 2014.

Mating and genital coupling in the primitive earwig
species Echinosoma denticulatum (Pygidicranidae):
implications for genital evolution in dermapteranphylogeny
Yoshitaka Kamimura *, 1, 2 & Chow-Yang Lee 1
Urban Entomology Laboratory, Vector Control Research Unit, School of Biological Sciences, Universiti Sains Malaysia, Minden 11800, Penang, Malaysia — 2 Department of Biology, Keio University, 4-1-1 Hiyoshi, Yokohama 223-8521, Japan; Yoshitaka Kamimura * [kamimura@fbc.keio.ac.jp] — * Corresponding author

1

Accepted 11.iii.2014.
Published online at www.senckenberg.de/arthropod-systematics on 8.iv.2014.

Abstract
Dermaptera (earwigs) shows much diversity in the genital structures, the presence of either one or two male intromittent organs (penes)
being one striking aspect. The members of several groups (Karschiellidae, Eudermaptera, Arixeniina, and Hemimerina) possess a single
functional penis, while others have a pair of penes. The latter condition is considered to be plesiomorphic in Dermaptera. Despite its importance for inferring the phylogeny of Dermaptera, it is presently unclear how the ancestor of earwigs acquired paired penes. To estimate themode of mating and sperm transfer in the common ancestor of extant earwigs, this study examines the mating behavior and genital couplingof the primitive earwig species Echinosoma denticulatum Hincks, 1959 as a representative of Pygidicranidae, one of the basal-most assemblages of earwigs. Staged mating experiments, including surgical manipulation of male penes revealed the following characteristics forthis species: (1) males use only one of the paired penes for a single genital coupling; (2) both penes are likely functional; (3) there are no
consistent biases in usage of the penes; (4) laterality of the penis-use pattern is not related to the direction of rotation of the male abdomen to
establish genital coupling; (5) sperm are transferred directly into the spermatheca, a female sperm storage organ; (6) the spined area of thepenis inflicts wounds on the vagina around the spermathecal opening, which bears many setae, during copulation. Characteristics (1)–(4)are considered to be plesiomorphic and may represent the condition of the common ancestor. Traumatic penetration during copulation isreported for earwigs for the first time. Together with intermittent acceptance of courting males by females only after a certain interval, theseresults suggest sexual conflict over mating in this species.

Key words
Evolution of laterally paired structures, dermapteran phylogeny, Pygidicranidae, genital evolution, mating behavior, insemination process,traumatic mating.

1.

Introduction

The insect order Dermaptera (earwigs) shows much diversity in the genital structures, including size, number
and laterality of both male and female genital components, and the degree of differentiation of female com-

ISSN 1863-7221 (print)| eISSN 1864-8312 (online)

ponents, making them valuable study models of genital
evolution (e.g., Popham 1965; Steinmann 1986, 1989,
1990, 1993; Kamimura 2000, 2004a,b, 2006, 2007b,
2013, 2014; Kamimura & Matsuo 2001; Klass 2001,

11

Kamimura & Lee: Reproductive biology of Echinosoma

2003; Kamimura & Iwase 2010; Schneider & Klass
2013). The presence of either a single or a pair of male
intromittent organs (penes) is an especially striking feature of earwigs.
Dermaptera is generally subdivided into three taxa:
Hemimerina, Arixeniina and Forficulina. Members of theformer two taxa live on mammals (hamster rats [Crice­
tomys spp.] in Africa and bats in Asia, respectively) andshow many adaptations to their phoretic-epizoic lifestyle(Nakata & Maa 1974). According to the classificationof Sakai (1982), Forficulina comprises eight families oftypical free-living earwigs: Pygidicranidae, Diplatyidae,
Anisolabididae, Apachyidae, Labiduridae, Spongiphoridae, Chelisochidae, and Forficulidae. Karschiellinae of
the Pygidicranidae in Sakai’s system is sometimes treated as the ninth family Karschiellidae (e.g., Popham 2000;
Haas & Klass 2003; Jarvis et al. 2005); here we follow
this view. Figure 1 shows the currently most elaborate
phylogenetic hypothesis for Forficulina. Several studies
have suggested that both Hemimerina and Arixeniina are
in-group members of Forficulina (Klass 2001; Haas &Klass 2003; Jarvis et al. 2005; Tworzydlo et al. 2012;
Kocarek et al. 2013), but their phylogenetic placements
in Forficulina have not been settled.
Males of all Dermaptera have a genital (phallic) organ placed at the midline of the body in the reflected
membrane above the coxosternum of abdominal segment
IX (subgenital plate). In six families the genitalic organ
includes a pair of penes, with few exceptions (Fig. 1;
for genital terminology, see Fig. 2). Among the “basal”
dermapteran families (Karschiellidae, Pygidicranidae,
and Diplatyidae), the right and left penes are symmetrical in Pygidicranidae and Diplatyidae, both pointing to
the head when not in copulation, while in Karschiellidae
the left penis is strongly reduced (e.g., Steinmann 1986).
In Apachyidae, Anisolabisidae, and Labiduridae of the“higher Forficulina”, penes are asymmetric when in repose: the left and right penes point in opposite directions(e.g., Popham 1965; Steinmann 1989; Kamimura & Matsuo 2001; Kamimura 2006; Fig. 1). The members of theother three families of the higher Forficulina, which arecollectively termed Eudermaptera, have only one penis
without exception (Fig. 1; Steinmann 1990, 1993), as do
the phoretic earwigs (Hemimerina and Arixeniina; Nakata & Maa 1974). Importantly, however, in these taxa
(Eudermaptera, Hemimerina and Arixeniina) either the
ejaculatory ducts and gonopores are still paired, or there
is at least some reminiscence of a formerly paired condition by the presence of a vestigial ejaculatory duct on
the non-functional side (e.g., Snodgrass 1959: fig. 4H,I;
Popham 1965: fig. 4; Kamimura 2006: fig. 3C, table 4).
Although the detailed relationships among the majorityof the families of earwigs are still unsettled, all previous studies have assumed or estimated that paired penes
represent the ancestral state (Popham 1985; Sakai 1987;
Haas 1995; Haas & Kukalová-Peck 2001; Colgan et al.
2003; Haas & Klass 2003; Kamimura 2004b; Jarvis etal. 2005). In addition, to our knowledge, no examples ofreversals from the status ‘asymmetrically paired’ to the12status ‘symmetrically paired’ or from ‘single penis’ tothe status of ‘asymmetrically paired’ have been reported.
Thus, the number and direction of penes are consideredstable characters and are used to define higher taxa in
Dermaptera (Fig. 1; Burr 1915a,b, 1916; Popham 1965;
Steinmann 1986, 1989, 1990, 1993).
Irrespective of the importance for inferring the phylogeny of Dermaptera and its placement in the polyneopteran insect orders, the evolutionary reason how theancestor of earwigs acquired paired penes has remained
unexplained. In anisolabidids such as Anisolabis mar­itima (Bonelli, 1832) and Euborellia plebeja (Dohrn,1863), males have two functionally competent penes although they use only one of them during a single genital
coupling (Kamimura 2000; Kamimura & Matsuo 2001).
Each penis bears an elongated virga, which is a heavily sclerotized process which contains the terminal partof the ejaculatory duct and has the gonopore on its tip.
Since the virgae are fragile, they sometimes break off
during mating; therefore, the remaining counterpart inpaired penises functions as a spare (Kamimura & Matsuo 2001). This phenomenon can explain the modern
function of paired penes in this specialized group. However, because not all earwigs, especially those in basalgroups, have thin and elongated virgae, this spare function is not likely sufficient to explain the evolution ofpaired penes and virgae in the ancestor of earwigs. The
usage pattern of paired penes has been investigated fora few representative species of Diplatyidae, Anisolabididae and Labiduridae. In Diplatys flavicollis Shiraki
1907 (Diplatyidae), Anisolabis littorea (White, 1846)(Anisolabididae), A. maritima and E. plebeja, males use
or are ready to use the right or left penes without great
bias (Giles 1961; Kamimura & Matsuo 2001; Kamimura
2003b, 2004a). In contrast, males of Labidura riparia(Pallas, 1773) (Labiduridae) predominantly use the rightpenis for insemination, although both penes are functional without any detectable morphological differentiations between them (Kamimura 2006). Previous phylogenetic studies based on morphology and/or molecular
data have suggested a sister-group relationship between
the Labiduridae and Eudermaptera (Spongiphoridae +Chelisochidae + Forficulidae), which is characterized
by a single penis (Sakai 1987; Haas 1995; Wirth et al.
1999; Haas & Kukalová-Peck 2001; Haas & Klass
2003; Colgan et al. 2003; Kamimura 2004b; Jarvis etal. 2005; Fig. 1). Therefore, extant labidurids may haveretained morphological and/or behavioral charactersthat led eventually to the loss of the less frequently used
left penis in the common ancestor of the Eudermaptera(Kamimura, 2006, 2007b).
Despite its importance for estimating the evolutionary history of earwig genitalia, the detailed functional
morphology of male and female genitalia has been studied for only one species of the basal Dermaptera, D. fla­
vicollis (Diplatyidae; Kamimura 2004a). The membersof Diplatyidae have paired penes, each of which bears
a bifurcated virga (e.g., Steinmann 1986). In several
diplatyids, each of bifurcated virgae bears two independ-

ARTHROPOD SYSTEMATICS & PHYLOGENY — 72 (1) 2014

Fig. 1. The most elaborate phylogenetic hypothesis for the Forficulina (modified fromHaas & Klass 2003) with the characteristicsof the male genitalia of each group. AlthoughHaas & Kukalová-Peck (2001) originally proposed that Karschiellidae is the first offshootof the present Dermaptera, while paraphyletic
Diplatyidae is basal-most to the other extant
Forficulina, subsequent detailed examinationof female genitalia by Klass (2003) supported
the monophyly of two taxa, Diplatyidae (Ha­plodiplatys and Diplatys) and Karschiellidae +Diplatyidae (Haas & Klass 2003).

ent gonopores, yielding four exits of ejaculates per male
(Popham 1965; Steinmann 1986; Kamimura 2004a). Female diplatyids also have multiple spermathecae (Popham 1965; Klass 2003; Kamimura 2004a). Fixation andexamination of copulating pairs of D. flavicollis revealed
that males inserted only one of the paired virgae into thefemales during a single genital coupling. To determine
whether this method of penis use actually represents a
plesiomorphy in the extant earwigs, it is especially important to examine the functional aspects of the genitalmorphologies in the members of Pyginicranidae, whose
subgroups (“subfamilies” and “genera”) represent themost basal clades of earwigs together with Diplatyidae(+ Karschiellidae) in the estimated earwig phylogenies(Haas & Klass 2003; Fig. 1).
In this study, we examined the mating behavior,
genital morphology and genital coupling in the earwig
Echinosoma denticulatum Hincks, 1959 (Pygidicranidae: Echinosomatinae). For this species, taxonomists
described the presence of a pair of symmetrical penes,
each bearing a single slender virga, and a well-developed
spined area on each penis (Hincks 1959). The internal
genitalia of females has not been described, but female
genitalia including cuticulized parts of the gonoducts are
known for the congener E. yorkense Dohrn, 1869 (Klass
2003). Courtship and mating behaviors of pygidicranids
have been reported for only one species, Tagalina papua(Bormans, 1903) by Matzke & Klass (2005), in which
both males and females show no apparent precopulatory
courtship and copulation is long (14 – 20 hours). During
the course of the present study, we found that the stout
spines of the penis inflict wounds on female genitalia
during copulation. This type of mating, termed traumatic
mating by Lange et al. (2013), has been reported in several insect taxa but not previously from earwigs (Langeet al. 2013). We here report the occurrence of traumatic
mating in earwigs for the first time and discuss the significance of this phenomenon.

2.

Materials and methods

2.1.

Insects and rearing

Two males and about fifty nymphs of Echinosoma dentic­
ulatum were collected from forested areas of Bukit Jambul and Bukit Kukus, Penang Island (05°30′N 100°28′E),
Malaysia, during May – November 2012. Field-collected
adults were individually placed in a separate plastic vessel (60 mm diameter, 40 mm high) with plaster of Paris
at its base and kept at 26 ± 1°C (12 h photoperiod). All
animals were provided with water, unlimited amounts ofcommercial cat food and a small piece of peeled tree barkfor harborage (and an oviposition site for mated females;
see below). Nymphs were kept in a group (up to 40 individuals) in vessels of a similar setting, and checked for
imaginal eclosion every three days. Laboratory-raised
virgins of both sexes (6 – 44 days old; including F1 andF2 generations) were used for various types of observation and experiments described below. All of the following observations and experiments were conducted under
the same laboratory conditions.

2.2.

Frequency and duration of copulation

To observe courtship and mating behaviors, a virgin male(but in two cases, field-collected males of unknown age)
and a virgin female were introduced into a mating arena(identical to the rearing vessel but without harborage)
with a transparent plastic lid. To avoid possible effects
of inbreeding and kin-recognition, pairing of a male anda female from the same full-sib family (i.e., offspring of
13

Kamimura & Lee: Reproductive biology of Echinosoma

the same male-female pair) was avoided. Their behavior was recorded for 24 – 72 hours by a video camera(GZ-MG980S; Victor, Kanagawa, Japan) using a timelapse recording function (one frame per two seconds).
Observations in dark periods (1900 – 0700 hour) wereconducted under a dim red light. The number of pairs
observed simultaneously ranged from one to three, basedon the availability of adults. Because of the limitation in
the number of video recorders, the starting time, endingtime and duration of video recording varied according
to the schedule of other experiments. After video recording, the females were individually reared for two months
to produce offspring. After two months, the spermatheca
was dissected out from the females in insect Ringer’s solution (0.9 g NaCl, 0.02 g CaCl2, 0.02 g KCl and 0.02
g NaHCO3 in 100 ml water) under a stereo microscope(EZ vision, Saxon, Guangzhou, China) and then examined under light microscopes (BX53 and CX21, Olympus, Tokyo; 40 – 400×). A total of 21 pairs were observed.
The subsequent dissection revealed that one female had
malformed genitalia and one female that died before twomonths after the recording could not be dissected becauseof the bad condition of the body. These two females were
removed from the subsequent analysis. Genitalia from
fifteen laboratory reared virgin females were also examined using the same method.

2.3. Penis-use patterns and surgical		 treatment
Two mating experiments were conducted to examine theuse pattern and the insemination potential of the paired
penes of E. denticulatum. In the first experiment, adultpairs were released into a mating arena (see 2.2.). Ten
minutes after the initiation of copulation, the mating
pairs were instantaneously fixed by pouring liquid nitrogen into the vessel. These samples were later dissected
to observe the genital coupling and to determine which
penis had been used for insemination.
For the second experiment, either the right or left penis, randomly determined, was excised from males thatwere lightly anaesthetized with carbon dioxide using fineforceps (n = 23 in total; Fig. 2D). After at least six days
following surgery, the males were individually pairedwith a virgin female. All of the pairs copulated at least 10
min. When copulation lasted over 60 min, the pairs wereartificially dislodged. At the initiation of copulation, themale mating posture (direction of abdominal rotation;
Fig. 2A) was also recorded. As in Kamimura (2006),
the direction of this abdominal rotation was defined asclockwise (CW) or counterclockwise (CCW), as viewed
from the head of the male (see Fig. 2A for an example of
CCW rotation). The insects were then stored in a freezer(– 20°C) for later dissection, in which the success of thesurgical treatment and insemination were examined forthe male and female samples, respectively. All statisticalanalyses of the data were conducted using the R 3.0.1
software (R Core Team 2013).

3.

Results

3.1.

Courtship and mating behavior

When a virgin female and virgin male pair was placedin a mating arena (n = 19), the male actively courted thefemale by directing his abdomen and forceps toward thefemale abdomen, simultaneously rotating the abdomen
180° around the anterior-posterior axis and retreating
to establish genital contact. The first copulation usually
took place within a few minutes (up to 38 minutes), irrespective of the start time of the observation (Fig. 3).
In 12 of the 19 cases, repeated matings were observedin the same pairs. Later rearing and dissection revealed
that two females did not lay eggs and had no detectable
sperm in the spermatheca. These two uninseminated females (the two samples at bottom of Fig. 3) mated two
and six times, respectively, while the 17 inseminated females mated up to four times (Fig. 3). Copulation lasted
153 ± 94 min (mean ± SD, n = 41 including all copulations of all pairs). The average duration of the first copulation was longer than those of the second and third copu-→ Fig. 2. Mating posture (A), male genitalia (B – D) and female genitalia (E – G) of Echinosoma denticulatum. In this paper, the terminologies of Klass (2003) and Kamimura (2014) are followed for female and male genital structures, respectively. The spined areas of the malepenis and the setaceous parts of the female genitalia are indicated by the open (in B and C) and filled arrowheads (in E – G), respectively.
A: Mating posture. The male (left) has the abdomen twisted in a counterclockwise (CCW) direction (indicated by the gray curved arrow)to establish genital coupling with the female (right). B: Male genitalia, entire view from dorsally, penes extended and endophallic sac ofleft penis (lp) evaginated. C: The spined area of the penis. D: Male genitalia with the right penis excised. E: Female genitalia. F: The setaceous area in the vagina of a virgin female. G: The setaceous area of a mated female with many melanized patches indicated by the arrows.
Similar patches were not detected in the same area of virgin females, indicating that these are repaired wounds inflicted by the male penial
spines. All photographs except for (A) are shown in the direction indicated in (B). – Abbreviations: gl9, gonoplac IX; gp8, gonapophysis
VIII; gp9, gonapophysis IX; lp, left penis; pm, paramere; rp, right penis; so, spermathecal opening; sp, spermatheca; vir, virga. – Scale bars:
200 µm in B – E, 100 µm in F, G.

14

ARTHROPOD SYSTEMATICS & PHYLOGENY — 72 (1) 2014

lations (Table 1). However, there were no significant differences among them when we analyzed only the cases
in which the male successfully inseminated the female(Table 1). Although the males continued to court the female frequently after their first mating, the females usually escaped from the courting male and accepted them

only after a certain interval (Fig. 3). In the cases in which
the female was inseminated, copulations were repeatedat 8−24 hour intervals with a few exceptions (Fig. 4). In
contrast, the two uninseminated females repeatedly mated with the same male at intervals of less than four hours
with one exception (Fig. 4).

15

Kamimura & Lee: Reproductive biology of Echinosoma

Table 1. Copulation durations (in min; mean ± SD [n]) in Echinosoma denticulatum. There were significant differences among the durations of the first, second and third copulations when all samples observed were included (Kruskal-Wallis test; χ22 = 6.80, P = 0.033), but thedifferences were insignificant when only pairs in which the females were inseminated were analyzed (χ22 = 5.03, P = 0.080).

12

1st

2nd

3rd

4th

5th

6th

All cases

194 ± 72[19]

117 ± 99[12]

115 ± 99[5]107 ± 98
[3]

12
[1]

259
[1]

Females
inseminated201 ± 59[17]

128 ± 105[10]139 ± 97[4]

57 ± 62
[2]

—

—

18

0

6

12

18

0

6

12

18

0

6

12

18

0

Hours
Fig. 3. Copulation duration and frequencies in 19 pairs of Echinosoma denticulatum. Each rectangle represents the time window of video
recording for each pair, in which the filled parts indicate copulation. Later dissection revealed that the two bottom-most females were notinseminated, while the others were inseminated. Dark periods are indicated by gray shading.

Frequency

3.2.
		

–4

4–8

8–12 12–16 16–20 20–24 24–
Copulation interval (hr)Fig. 4. Intervals between successive matings in repeatedly mating
pairs of Echinosoma denticulatum. The data for pairs where females were inseminated (10 pairs, 16 intervals) and for pairs wherefemales remained uninseminated (2 pairs, 6 intervals) are shown by
open and filled bars, respectively.

16

Genitalia, genital coupling andcopulatory wounds

As described by Hincks (1959), each of the paired penes of E. denticulatum bears an area of many strong
spines, each about 100 µm long, and a slender virga in
the endophallus, which is evaginated during copulation(Fig. 2B,C). The female genital structure is essentiallysimilar to what Klass (2003) described for Echinosoma
yorkense: anteriorly to two pairs of the major ovipositor
components (a pair of gonapophyses VIII and a pair ofgonoplacs IX [= coxal lobes IX]), a single, long, tubular
spermatheca without internal branching opens into thedorsal side of the vagina (Fig. 2E). The vagina of E. den­ticulatum is membranous, concave to the dorsal side, andcovered with many long (ca. 150 µm) setae (Fig. 2E,F).

ARTHROPOD SYSTEMATICS & PHYLOGENY — 72 (1) 2014

Table 2. The relationship between the direction of abdomen rotation during copulation (CW or CCW) and the penis used in Echi­
nosoma denticulatum. P = 0.41 (Fisher’s exact probability test).
Clockwise(CW)

Counterclockwise(CCW)

Right penis used
(Left penis excited)

5

6

Left penis used
(Right penis excited)8

4

All 19 females used for the observation of normal
mating behavior, including the two uninseminated ones,
showed a number of melanized patches around the setaceous area posterior to the spermathecal orifice on which
the spines of the male penis pressed during genital coupling (Fig. 2G). None of the virgin females examined(n = 15) had detectable melanized patches in the area,
indicating that these were repaired wounds inflicted by
the male penial spines during copulation.

3.3.
With
sperm

Without
sperm

0

20
40
Copuration duration (min)60

Fig. 5. The relationship between insemination success and copulation duration revealed by surgical experiment in Echinosoma den­ticulatum. The matings were artificially terminated at 60 min after
initiation. The open circles and open diamonds represent the data
for the right- and left-penis-excised males, respectively. The curved
line shows predictions of probability of insemination from logistic
regression (χ21 = 4.64, P = 0.031).

The length and orientation of the setae varied with the region in the vagina, but most setae are directed posteriorly(from inner parts towards the entrance of vagina).
In the freeze-fixation experiment of mating pairs(n = 12), five males used the right penis. In these males,
the right penis was directed posteriorly for genital coupling, while the left penis pointed toward the head as
when in repose. For the other seven males, the opposite
condition was observed. There was no significant bias in
the laterality of penis use (5 right : 7 left; binomial test,
P = 0.77). The penes used were dilated (endophallic sac
evaginated, possibly caused by increased hemolymph
pressure) and the virga and spined area were exposed
outside (as shown in Fig. 2B). When the right (left) penis
was used, it was inserted into the female genital chamber
through the membranous region on the right (left) sideof the right (left) gonoplac IX (note that because of theend-to-end mating posture and 180° rotation of the maleabdomen around the anterior-posterior axis, the rightside of the male contacts the right side of the female).
Although the male penis was easily separated after melting of the samples, in the frozen samples, the spined areaof the penis in use and the setaceous area of the vagina
were in contact, and the virga was inserted directly into
the spermatheca.

Surgical experiment

As described above, the freeze-fixation experiment revealed that there was no consistent bias in the lateralityof penis use (five and seven males used the right and leftpenes, respectively). However, this fact does not necessarily indicate that a single male of E. denticulatum canuse both the right and left penes. A surgical experiment
was conducted to examine whether only one of the twopenes is functional or not. The males from which one ofthe paired penes had been artificially excised (12 and11 cases for the right and left, respectively), courted
and copulated easily with a virgin female, just as normal males. Fifteen pairs copulated for 60 min and wereartificially dislodged. All but one females of these pairs
had detectable sperm in the spermatheca (Fig. 5). The
other eight pairs spontaneously terminated copulation
before 60 min after initiation (Fig. 5); among these, theprobability of insemination increased with the durationof copulation, indicating a variation in the intervals between establishment of genital coupling and the initiationof sperm transfer (Fig. 5). If only one of the paired penes
is functional, only 50% of the penis-excised males are
expected to successfully inseminate the female. However, the observed insemination success (17 of 23 cases) ishigher than expected from this null hypothesis (binomial
test, P = 0.035). Considering that several pairs copulated
only once for a short time in this experiment, and that ina few cases females remained uninseminated even after
prolonged mating with a normal male (Fig. 3), it is highly
likely that both right and left penes of E. denticulatum arefunctional. The direction of abdominal rotation was biased toward neither the CW nor CCW direction (13 : 10;
binomial test, P = 0.68), with no significant relationshipto the laterality of penis use (Table 2).

4.

Discussion

This study showed the following characteristics of matingin E. denticulatum: (1) males use only one of the paired
penes for a single copulation; (2) both the right and left
17

Kamimura & Lee: Reproductive biology of Echinosoma

penes are likely functional; (3) there is no consistent bias
in the usage of the right or left penes; (4) the lateralityof penis-use pattern (right or left) is not related to thedirection of abdominal rotation (CW or CCW); (5) one
virga is inserted directly into the spermatheca to transfer
sperm; (6) the spined area of the penis (endophallus) inflicts wounds on the vaginal wall around the spermathecal
opening, on which females bear many setae, during copulation. Among these characteristics, (1) is shared by allother earwigs examined to date (Kamimura 2004a, 2006)
and is likely to be a common feature of earwigs. Apparently, the vagina of E. denticulatum can only accommodate one penis at a time. Characteristic (2), the insemination competency of both paired penes, was also confirmed for D. flavicollis (Diplatyidae; Kamimura 2004a),
E. plebeja (Anisolabididae; Kamimura & Matsuo 2001)
and L. riparia (Labiduridae; Kamimura 2006) and is also
likely to be common among earwigs with paired penes.
In contrast, the lack of consistent lateral bias in penis use[characteristic (3)] observed in E. denticulatum is likely
shared by diplatyids and anisolabidids. This supports theview that the conspicuous right-handedness in L. riparia
is an apomorphy, which may have resulted in the reduction of the left side penis in the sister clade Eudermaptera(Kamimura 2006; see Introduction for details).
How did the ancestor acquire a paired condition ofpenes and gonopores, and how has the paired state been
maintained in many extant earwigs? In several earwig
species with paired penes, damaged penes or virgae aresporadically observed and considered to have occurred
accidentally during copulation (Kamimura & Matsuo
2001; Kamimura 2003a, 2006). Although further scrutiny is needed for Echinosoma, we failed to detect any
naturally caused genital damage in the males used for themating experiments. In many insect taxa, the evolutionof asymmetric genitalia is associated with asymmetriccopulatory positions (Ludwig 1970; Huber et al. 2007).
Thus, another possibility is that the direction of abdominal rotation is related to the laterality of penis use, that is,
one penis for CW and another for CCW rotation of themale abdomen (Kamimura 2003b). To our knowledge,
the present study is the first attempt to address laterality in the mating posture of basal Dermaptera. However,as in the two species of the higher Forficulina studied todate (E. plebeja, Kamimura 2003b; L. riparia, Kamimura
2006), no consistent correlation was observed between
the two asymmetric traits in E. denticulatum (Table 2).
Several recent studies have supported a sister relationshipbetween Dermaptera and Plecoptera (stoneflies) among
the polyneopteran insect orders (Kjer 2004; Yoshizawa& Johnson 2005; Misof et al. 2007; Ishiwata et al. 2011;
Yoshizawa 2011; Wan et al. 2012). Moreover, several
stonefly species also have paired orifices of ejaculatoryducts that apparently function as sperm exits (Brink
1956a,b). Thus, the origin of paired earwig penes may
date back to the common ancestor of these two orders.
This study revealed the occurrence of a closed spermtransfer; i.e., direct insertion of the virga into the spermatheca for sperm transfer [characteristic (5)], in a mem18

ber of the basal Dermaptera for the first time. This feature
differs from the open system of the diplatyid D. flavicol­
lis, in which two male gonopores of a bifurcated virga areclosely pressed against a row of two or three spermathecal openings, but are not directly inserted into the spermathecae (Kamimura 2004a). According to the phylogenetic relationships proposed by Haas & Klass (2003),
it is presently equivocal whether the common ancestorsof the extant earwigs possessed a closed or an open system of transfer (Fig. 1). The virgae of many membersof two subfamilies of Pygidicranidae, Pyragrinae andEsphalmeninae, are bifurcated, suggesting the presenceof four exits for sperm per male as in some diplatyids(e.g., Brindle 1984; Sakai 1985; Steinmann 1986). The
genus Pyragra Audinet-Serville, 1831 (Pyragrinae) is estimated to be the basal-most clade in the paraphyletic Pygidicranidae, while the placement of Esphalmeninae has
not been settled (Haas & Klass 2003). Thus, possibly inassociation with the evolution of non-bifurcated virgae,
the closed system of sperm transfer may had evolved in aclade of the paraphyletic Pygidicranidae and be presently
shared by “the Pygidicranidae (not necessarily all members, but including Echinosoma) + higher Forficulina.”
Genital couplings in other representatives of the basal
Dermaptera should be examined to confirm this hypo­thesis.
Functional aspects of associated sclerites or spines ofthe earwig penis have been examined in only two earwig
species to date. The male penis of Paralabellula dorsalis
Burmeister, 1838 (Spongiphoridae; referred to as Para­labella dorsalis in Briceño 1997; cf. Kevan & Vickery1997) possesses several areas with hardened spines (referred to as “toothed plates” in Briceño 1997). During
copulation, the membranous penis is inflated and thesespines contact areas around the female gonopore andthe lateral wall of the vagina (Briceño 1997). Males ofD. flavicollis possess three differently shaped accessory
sclerites, specifically U-, rod- and saber-shaped sclerites,in each penis (Kamimura 2004a). In an established genital coupling, two lateral pouches in the female genitalchamber receive the U- and rod-shaped sclerites, while
the saber-shaped sclerite contacts the female subgenital plate (Kamimura 2004a). Although these structuresare supposed to function as anchorage devices during
copulation, occurrence of copulatory trauma has notbeen reported for these two species (re-examination by
Y. Kamimura of the female samples used in Kamimura
2004a failed to reveal any traces of wounds).
Traumatic mating has been reported for members ofseven insect orders (Orthoptera, Hemiptera, Coleoptera,
Strepsiptera, Diptera, Lepidoptera and Hymenoptera;
Lange et al. 2013) and the present study adds Dermaptera
to the list as the eighth order. Lange et al. (2013) divided
the examples of traumatic mating into three categories:
traumatic insemination, traumatic secretion transfer andtraumatic penetration. In cases of the former two categories, wounds function as the entrance of semen andseminal secretions (without sperm) into the female body,respectively (see Hotzy et al. 2012; Lange et al. 2013;

ARTHROPOD SYSTEMATICS & PHYLOGENY — 72 (1) 2014

Tatarnic et al. 2014 for examples of these categories). In
E. denticulatum, because the wounds are distributed outside of the spermatheca, while after dissection semen isfound inside the sclerotized spermatheca, the copulatory
wounds are unlikely to function as an entrance of either
sperm or seminal fluid into the female hemocoel. The
case of E. denticulatum thus represents an example oftraumatic penetration. There is continuing debate about
the function and significance of this type of traumatic
mating (Lange et al. 2013). Although many experimental
or observational studies support the view that wounding
is a side effect of mate-anchoring during copulation (e.g.,
Teuschl et al. 2007; Kamimura & Mitsumoto 2011; Okuzaki et al. 2012), the wound itself or its associated damage may increase the paternity gain of the focal male via
stimulatory functions (Eberhard 1996, 2010). Damaging
the female itself can produce similar effects via induction of immediate reproductive output (termed terminal
reproductive output) or reduction of the probability of remating (Johnstone & Keller 2000; Morrow et al. 2003).
Although there is no confirmative empirical support forthese hypotheses, the observed repeated copulation byfemale E. denticulatum that took place only after a certain interval is a possible candidate phenomenon that canbe explained by the latter hypothesis. Future studies are
warranted to substantiate this possibility as well as thecircadian rhythms of both sexes. Moreover, two females
remained uninseminated after prolonged repeated matings by apparently normal males (Fig. 3) and frequentpremature dislodgement of matings with the surgicallytreated males (Fig. 5), both of which have not been reported for other earwig species, also indicate conflict between the sexes with respect to control over the outcomeof mating.
Female insects can develop specialized structures,such as pouches, screlotized plates and thickened walls,
in their genitalia where male genitalia inflict wounds;
these are considered counteradaptations for traumatic
mating (e.g., Rönn et al. 2007; Kamimura 2007a, 2012).
Detailed female genital structures were reported for only
two other species of the genus Echinosoma: E. afrum(Palisot de Beauvois, 1805) (Giles 1963) and E. yorkense(Klass 2003). In these species, males lack conspicuous
penial spines (Giles 1963; Sakai 1985; Steinmann 1986).
In E. yorkense much of the vaginal walls show a dense
cover of minute hair-like elements, which are much
shorter than the setae of E. denticulatum (Klass 2003;
K.-D. Klass, pers. comm.), while detailed description of
E. afrum by (Giles 1963) lacks any mentions of setae orbristles on the vaginal wall. Thus, the long setae on thevaginal wall likely represents a counteradaptation of female E. denticulatum to the male genital spines. Interestingly, female Paralabellula dorsalis also bear sclerotized
spines around the gonopore (the orifice of the common
oviduct) and some of the teeth of the male toothed platemesh with them during copulation (Briceño 1997).
However, such spiny structures are totally absent fromboth male and female genital structures of the congeners Paralabellula curvicauda (Motschulsky, 1863) and

Paralabellula rotundifrons (Hincks, 1954) (Steinmann
1990; Sakai 1993; Y. Kamimura, unpublished data).
Thus, such a possible counteradaptation is likely to have
evolved independently multiple times in distantly related
earwig lineages (Pygidicranidae in the basal Dermaptera
and Spongiphoridae in the higher Forficulina). To date,no comprehensive study has been conducted on sexual
conflict over mating in any species of earwig (Kamimura
2014). These groups should be excellent study models
for this topic of evolutionary biology.

5.

Acknowledgements

This study was conducted with the approval of the Economic Planning Unit, Malaysia (Reference No. UPE:
40/200/19/2844). We thank H.-S. Tee for his assistancein insect rearing, M. Nishikawa for identification of species and providing important references, G. Veera Singham for his help in the freeze-fixation experiment andthe staff of the Electron Microscope Unit, School of Biological Science, Universiti Sains Malaysia for their help
in microscopic observations. We are also grateful to K.D. Klass and F. Haas for their helpful comments on an
earlier version of the manuscript. This study was partly
supported by a grant for overseas research from Keio
University and a Grant-in-Aid for Scientific Research(Kakenhi, No. 22770058) from the Japan Society for the
Promotion of Science to Y.K.

6.

References

Briceño R.D. 1997. Genitalic structure and copulation in Parala­
bella dorsalis (Dermaptera: Labbidae). – Revista de Biologia
Tropical 45: 1107 – 1116.
Brindle A. 1984. The Esphalmeninae (Dermaptera: Pygidicranidae): a group of Andean and South African earwigs. – Systematic Entomology 9: 281 – 292.
Brink P. 1956a. Reproductive system and mating in Plecoptera I. –Opsucula Entomologica 21: 57 – 96.
Brink P. 1956b. Reproductive system and mating in Plecoptera II. –Opsucula Entomologica 21: 97 – 128.
Burr M. 1915a. On the male genital armature of the Dermaptera
Part 1: Protodermaptera (except Psalidae). – Journal of theRoyal Microscopical Society 1915: 413 – 447.
Burr M. 1915b. On the male genital armature of the Dermaptera
Part 2: Psalidae. – Journal of the Royal Microscopical Society
1915: 521 – 546.
Burr M. 1916. On the male genital armature of the Dermaptera
Part 3: Eudermaptera. – Journal of the Royal MicroscopicalSociety 1916: 1 – 18.
Colgan D.J., Cassis G., Beacham E. 2003. Setting the molecularphylogenetic framework for the Dermaptera. – Insect Systematics and Evolution 34: 65 – 79.

19

Kamimura & Lee: Reproductive biology of Echinosoma

Eberhard W.G. 1996. Female Control: Sexual Selection by Cryptic
Female Choice. – Princeton University Press, Princeton, New
Jersey. 501 pp.
Eberhard W.G. 2010. Rapid divergent evolution of genitalia: theory and data updated. Pp. 40 – 78 in: Leonard J.L., CórdobaAguilar A. (eds), The Evolution of Primary Sexual Charactersin Animals. – Oxford University Press, Oxford, UK.
Giles E.T. 1961. The male reproductive organs and genitalia of
Anisolabis littorea (White) (Dermaptera: Labiduridae). –Transactions of the Royal Society of New Zealand (Zoology)
1: 203 – 213.
Giles E.T. 1963. The comparative external morphology and affinities of the Dermaptera. – Transactions of the Royal Entomological Society, London 115: 95 – 164.
Haas F. 1995. The phylogeny of the Forficulina, a suborder of theDermaptera. – Systematic Entomology 20: 85 – 98.
Haas F., Klass K.-D. 2003. The basal phylogenetic relationships in
the Dermaptera. In: Klass K.-D. (ed.), Proceedings of the 1st
Dresden meeting on insect phylogeny: “Phylogenetic Relationships within the Insect Orders” (Dresden, September 19 – 21,2003). – Entomologische Abhandlungen 61: 138 – 142.
Haas F., Kukalová-Peck J. 2001. Dermaptera hindwing structureand folding: New evidence for familial, ordinal and superordinal relationships within Neoptera (Insecta). – European Journalof Entomology 98: 445 – 509.
Hincks W.D. 1959. A Systematic Monograph of the Dermapteraof the World. Part II. Pygidicranidae excluding Diplatyinae. –British Museum of Natural History, London. 218 pp.
Hotzy C., Polak M., Rönn J.L., Arnqvist G. 2012. Phenotypic engineering unveils the function of genital morphology. – Current
Biology 22: 2258 – 2261.
Huber B.A., Sinclair B.J., Schmitt M. 2007. The evolution ofasymmetric genitalia in spiders and insects. – Biological Reviews 82: 647 – 698.
Ishiwata K., Sasaki G., Ogawa J., Miyata T., Su Z.-H. 2011. Phylogenetic relationships among insect orders based on three nuclear protein-coding gene sequences. – Molecular Phylogenetics
and Evolution 58: 169 – 180.
Jarvis K.J., Haas F., Whiting M.F. 2005. Phylogeny of earwigs(Insecta: Dermaptera) based on molecular and morphologicalevidence: reconsidering the classification of Dermaptera. –Systematic Entomology 30: 442 – 453.
Johnstone R.A., Keller L. 2000. How males can gain by harming
their mates: sexual conflict, seminal toxins, and the cost of mating. – American Naturalist 156: 368 – 377.
Kamimura Y. 2000. Possible removal of rival sperm by the elongated genitalia of the earwig, Euborellia plebeja. – Zoological
Science 17: 667 – 672.
Kamimura Y. 2003a. Effects of broken male intromittent organs on
the sperm storage capacity of female earwigs, Euborellia ple­beja. – Journal of Ethology 21: 29 – 35.
Kamimura Y. 2003b. Genital structure and asymmetry in matingbe­havior of earwigs. – Nature & Insects 38: 42 – 45. [in Japanese]
Kamimura Y. 2004a. Mating behaviour and insemination in Diplat­ys flavicollis, an earwig with double-barrelled penises and avariable number of female sperm-storage organs. – Journal of
Zoology 262: 37 – 46.

20

Kamimura Y. 2004b. In serach of the origin of twin penises: molecular phylogeny of earwigs (Dermaptera: Forficulina) basedon mitochondrial and nuclear ribosomal RNA genes. – Annalsof the Entomological Society of America 97: 903 – 912.
Kamimura Y. 2006. Right-handed penises of the earwig Labidura
riparia (Insecta, Dermaptera, Labiduridae): Evolutionary relationships between structural and behavioral asymmetries. –Journal of Morphology 267: 1381 – 1389.
Kamimura Y. 2007a. Twin intromittent organs of Drosophila fortraumatic insemination. – Biology Letters 3: 401 – 404.
Kamimura Y. 2007b. Possible atavisms of genitalia in two speciesof earwig (Dermaptera), Proreus simulans (Chelisochidae) andEuborellia plebeja (Anisolabididae). – Arthropod Structure &Development 36: 361 – 368.
Kamimura Y. 2012. Correlated evolutionary changes in Drosophila
female genitalia reduce the possible infection risk caused bymale copulatory wounding. – Behavioral Ecology and Sociobiology 66: 1107 – 1114.
Kamimura Y. 2013. Promiscuity and elongated sperm storage organs work cooperatively as a cryptic female choice mechanism
in an earwig. – Animal Behaviour 85: 377 – 383.
Kamimura Y. 2014. Pre- and postcopulatory sexual selection andthe evolution of sexually dimorphic traits in earwigs (Dermaptera). – Entomological Science. doi:10.1111/ens.12058
Kamimura Y., Iwase R. 2010. Evolutionary genetics of genital sizeand lateral asymmetry in the earwig Euborellia plebeja (Dermaptera: Anisolabididae). – Biological Journal of the Linnean
Society 101: 103 – 112.
Kamimura Y., Matsuo Y. 2001. A “spare” compensates for the riskof destruction of the elongated penis of earwigs (Insecta: Dermaptera). – Naturwissenschaften 88: 468 – 471.
Kamimura Y., Mitsumoto H. 2011. Comparative copulation anatomy of the Drosophila melanogaster species complex (Diptera:
Drosophilidae). – Entomological Science 14: 399 – 410.
Kevan D.K.McE., Vickery V.R. 1977. An annotated provisional listof non-saltatorial orthopteroid insects of Micronesia, compiled
mainly from the literature. – Micronesica 30: 269 – 353.
Kjer K.M. 2004. Aligned 18S and insect phylogeny. – Systematic
Biology 53: 506 – 514.
Klass K.-D. 2001. The female abdomen of the viviparous earwig
Hemimerus vosseleri (Insecta: Dermaptera: Hemimeridae),with a discussion of the postgenital abdomen of Insecta. – Zoological Journal of the Linnean Society 131: 251 – 307.
Klass K.-D. 2003. The female genitalic region in basal earwigs(Insecta: Dermaptera: Pygidicranidae s.l.). – Entomologische
Abhandlungen 61: 173 – 225.
Kocarek P., John V., Hulva P. 2013. When the body hides the ancestry: Phylogeny of morphologically modified epizoic earwigs on molecular evidence. – PLoS ONE 8: e66900.
Lange R., Reinhardt K., Michels N.K., Anthes N. 2013. Functions, diversity, and evolution of traumatic mating. – Biological
Reviews 88: 585 – 601.
Ludwig W. 1970. Das Rechts-Links-Problem im Tierreich und beim
Menschen. Reprint of 1932 edition. – Springer-Verlag, Berlin.
496 pp.
Matzke D., Klass K.-D. 2005. Reproductive biology and nymphal
development in the basal earwig Tagalina papua (Insecta: Dermaptera: Pygidicranidae), with a comparison of brood care inARTHROPOD SYSTEMATICS & PHYLOGENY — 72 (1) 2014

Dermaptera and Embioptera. – Entomologische Abhandlungen
62: 99 – 116.
Misof B., Niehuis O., Bischoff I., Rickert A., Erpenbeck D., Stan
­ i­czek A. 2007. Towards an 18S phylogeny of hexapods: ac­count­ing for group-specific character covariance in optimized
mixed nucleotide/doublet models. – Zoology 110: 409 – 429.
Morrow E.H., Arnqvist G., Pitnick S. 2003. Adaptation versus
pleiotropy: why do males harm their mates. – Behavioral Eco­logy 14: 802 – 806.
Nakata S., Maa T.C. 1974. A review of the parasitic earwigs. – Pacific Insects 16: 307 – 374.
Okuzaki Y., Takami Y., Tsuchiya Y., Sota T. 2012. Mating behavior
and the function of the male genital spine in the ground beetle
Carabus clathratus. – Zoological Science 29: 428 – 432.
Popham E.J. 1965. The functional morphology of the reproductive
organs of the common earwig (Forficula auricularia) and otherDermaptera with reference to the natural classification of theorder. – Journal of Zoology 146: 1 – 43.
Popham E.J. 1985. The mutual affinities of the major earwig taxa
(Insecta, Dermaptera). – Zeitschrift für Zoologische Systematik und Evolutionforschung 23: 199 – 214.
Popham E.J. 2000. The geographical distribution of the Dermaptera(Insecta) with reference to continental drift. – Journal of Natural History 34: 2007 – 2027.
R Core Team 2013. R: A Language and Environment for StatisticalComputing. – R Foundation for Statistical Computing, Vienna,
Austria.
Rönn J., Katvala M., Arnqvist G. 2007. Coevolution between
harmful male genitalia and female resistance in seed beetles. –Proceedings of the National Academy of Sciences USA 95:
6217 – 6221.
Sakai S. 1982. A new proposed classification of the Dermapterawith special reference to the check list of the Dermaptera of theworld. – Bulletin of Daito Bunka University 20: 1 – 108.
Sakai S. 1985. Dermapterorum Catalogus. XVI – XVIII: Icono­gra­phia I – III. Pygidicranidae and Diplatyidae. – Daito Bunka
University, Tokyo. Pp. 698 – 1080.
Sakai S. 1987. Phylogenetic and evolutionary information on Dermaptera from the point of view of insect integrated taxonomy.
Pp. 496 – 513 in: Bacetti B.M. (ed.), Evolutionary Biology of
Orthopteroid Insects. – Ellis Horwood, Chichester.
Sakai S. 1993. Dermapterorum Catalogus. XXV. Iconographia
IX. Spongiphoridae. – Daito Bunka University, Tokyo. Pp.
4691 – 5285.
Schneider K., Klass K.-D. 2013. The female genitalic region in
Eu­dermaptera (Insecta: Dermaptera). – Zoologischer Anzeiger
252: 183 – 203.
Snodgrass R.E. 1959. A revised interpretation of the external reproductive organs of male insects. – Smithsonian Miscellaneous Collections 135(6): 1 – 60.
Steinmann H. 1986. Dermaptera. Catadermaptera I. – Das Tier­reich 102: 1 – 343.
Steinmann H. 1989. Dermaptera. Catadermaptera II. – Das Tier­reich 105: 1 – 504.
Steinmann H. 1990. Dermaptera. Eudermaptera I. – Das Tierreich
106: 1 – 558.
Steinmann H. 1993. Dermaptera. Eudermaptera II. – Das Tierreich
108: 1 – 711.

Tatarnic N.J., Cassis G., Siva-Jothy M.T. 2014. Traumatic insemination in terrestrial arthropods. – Annual Review of Entomo­
logy 59: 245 – 261.
Teuschl Y., Hosken D.J., Blanckenhorn W.U. 2007. Is reduced
female survival after mating a by-product of male-male competition in the dung fly Sepsis cynipsea? – BMC Evolutionary
Biology 7: 94.
Tworzydlo W., Lechowska-Liszka A., Kocarek P., Bilinski S.M.
2013. Morphology of the ovarioles and the mode of oogenesisof Arixenia esau support the inclusion of Arixeniina to the Eudermaptera. – Zoologischer Anzeiger 252: 410 – 416.
Wan X., Kim M.I., Kim M.J., Kim I. 2012. Complete mitochondrial genome of the free-living earwig, Challia fletcheri (Dermaptera: Pygidicranidae) and phylogeny of polyneoptera. –PLoS ONE 7: e42056.
Wirth T.R., Guellec L., Veuille M. 1999. Directional substitution
and evolution of nucleotide content in the cytochrome oxidase
II gene in earwigs (dermapteran insects). – Molecular Biology
and Evolution 16: 1645 – 1653.
Yoshizawa K. 2011. Monophyletic Polyneoptera recovered bywing base structure. – Systematic Entomology 36: 377 – 394.
Yoshizawa K., Johnson K.P. 2005. Aligned 18S for Zoraptera (Insecta): phylogenetic position and molecular evolution. – Molecular Phylogenetics and Evolution 37: 572 – 580.

21