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Journal of Chemical Ecology, Vol. 30, No. 6, June 2004 (°
CHEMICAL SIGNALING IN A WOLF SPIDER: A TEST
OF ETHOSPECIES DISCRIMINATION
J. ANDREW ROBERTS∗ and GEORGE W. UETZ
Department of Biological Sciences, University of Cincinnati, Cincinnati, Ohio 45221, USA
(Received August 12, 2003; accepted February 16, 2004)
Abstract—Chemical signals from female wolf spiders that elicit exploratory
behavior and courtship in males are often assumed to be species-specific, but
males of some species court in response to silk cues deposited by closely related
heterospecific females. Such is the case with the wolf spiders Schizocosa ocreata
and S. rovneri, ethospecies reproductively isolated on the basis of differences in
behavioral mechanisms during courtship. We explored whether male S. ocreata
and S. rovneri reciprocally discriminate species-specific chemical or mechanical
cues associated with female silk by using male behavioral response as an assay.
Males were exposed to stimulus treatment categories including silk, washed silk,
silk extract, and appropriate controls within conspecific or heterospecific female
stimulus categories. Male S. ocreata and S. rovneri did not discriminate between
conspecific or heterospecific female stimuli, and courtship intensity was greatest on untreated silk. There were no differences in latency to begin courtship
or in rates of courtship behaviors attributed to species origin of silk. However,
silk treatment (washed silk, extract) had a significant effect on display and exploratory behaviors (e.g., chemoexplore) in both species. Methanol extraction of
female silk successfully removed or inactivated a component necessary to elicit
active courtship, but extraction did not significantly reduce exploratory behavior, suggesting that a separate compound may be responsible for releasing this
behavior. Together, these experiments support the characterization of S. ocreata
and S. rovneri as ethospecies reproductively isolated only by female discrimination of species-specific male courtship, and indicate that chemical, but not
mechanical cues associated with silk are critical for eliciting male courtship in
both species.
Key Words—Behavioral assay, chemical communication, Schizocosa, Lycosidae, wolf spider, species discrimination.
∗
To whom correspondence should be addressed. E-mail: [email protected]
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INTRODUCTION
Spiders, like most arthropods, use chemical signals to mediate many types of
interactions. These interactions may include prey detection, predator avoidance,
social recognition, and courtship (reviews in Tietjen and Rovner, 1982; Uetz,
2000; Barth, 2002; Uetz and Roberts, 2002). Male spiders may produce compounds that alter the behavior of conspecifics, either in the context of male/male
competition by suppression of activity in other males (Tietjen, 1979; Ayyagari
and Tietjen, 1987), or in mating behavior by either releasing female receptivity
displays (Ross and Smith, 1979) or inducing a cataleptic state in females prior to
copulation (Singer et al., 2000). However, most studies of chemical signaling in
spiders have focused on compounds associated with the silk and/or cuticle of females and the role these putative pheromones play in mate attraction and release of
exploratory and courtship behaviors (Tietjen and Rovner, 1982; Pollard et al., 1987;
Riechert and Singer, 1995; Trabalon et al., 1997; Prouvost et al., 1999; Papke et al.,
2001).
In wolf spiders (Araneae: Lycosidae), there is evidence that females can attract males with airborne chemical signals (Tietjen, 1979; Searcy et al., 1999), but
most chemical signaling appears to be via contact chemical compounds associated
with silk. Conspecific female silk and associated pheromones are sufficient to elicit
normal courtship behavior in males of many species, even in the absence of all
visual and vibratory cues (den Hollander et al., 1973; Tietjen and Rovner, 1982;
Stratton and Uetz, 1983; Costa and Capocasale, 1984). While the chemical signals
from female wolf spiders that elicit exploratory behavior and release courtship in
males are thought to be species-specific (Hegdekar and Dondale, 1969; Tietjen,
1977; Costa and Capocasale, 1984), males of closely-related species will sometimes court the silk of heterospecifics (Kaston, 1936; Hegdekar and Dondale, 1969;
den Hollander, 1971; Miller et al., 1998).
Such is the case with males of the two wolf spider species used in this study,
Schizocosa ocreata (Hentz) and S. rovneri (Uetz and Dondale). These species can
be considered “ethological species” (Dobzhansky, 1970) and have been described
as “ethospecies” (Uetz and Denterlein, 1979), that is, species that are isolated reproductively only on the basis of differences in behavioral mechanisms occurring
during courtship (den Hollander and Dijkstra, 1974), with no other prezygotic
isolating mechanisms (Uetz and Denterlein, 1979; Stratton and Uetz, 1981, 1983,
1986). Schizocosa ocreata and S. rovneri prefer different microhabitats but overlap in geographical range and habitat use (Uetz and Denterlein, 1979; Stratton
and Uetz, 1981; McClintock and Uetz, 1996), and are essentially seasonally synchronous (Uetz and Denterlein, 1979; Roberts, J. A., unpublished data). Male S.
ocreata do not show evidence of territoriality, and may travel great distances each
day during some portions of the breeding season (linear distance of up to 15 m/hr—
Roberts, J. A. and Uetz, G. W., unpublished data), increasing chances of encounter
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with both conspecifics and heterospecifics. In fact, males allocate much of their
time to searching for females in the complex habitat (Cady, 1984).
There is no evidence of mechanical or gametic incompatibility, as hybrids of
these two species have been successfully produced in “forced” copulation studies in laboratory experiments (Stratton and Uetz, 1986; Orr, M. and Uetz, G. W.,
unpublished data). The genes that code for courtship behavior do not assort independently so that otherwise viable hybrids suffer behavioral sterility, and no
known natural hybrids have yet been identified (Stratton and Uetz, 1986). Females of these species are receptive only to species-specific visual and vibratory
courtship cues (Uetz and Denterlein, 1979; Stratton and Uetz, 1981, 1983, 1986;
but see Uetz and Roberts, 2002), a finding that is not surprising as females mating with heterospecifics would suffer fitness consequences related to behaviorally
sterile offspring.
Males of both species have been demonstrated to be equally likely to court
both conspecific and heterospecific silk (Uetz and Denterlein, 1979; Stratton and
Uetz, 1983), and S. ocreata males court with similar intensity on both silks (Roberts
and Uetz, unpublished data), but there has not been a rigorous, reciprocal test of
male discrimination of female cues. Male courtship is costly in these species, with
recent studies indicating a high energetic expense (Cady, A. B., Uetz, G. W., and
Delaney, K. J., unpublished data), and, for S. ocreata, that courtship characteristics
important in mate choice also increase predation (Roberts, J. A., Taylor, P. W., and
Uetz, G. W., unpublished data). Intraguild predation has a large impact on survival
in wolf spiders (Polis and McCormick, 1986; Wise and Chen, 1999), and males
making discrimination mistakes may become prey if they court heterospecifics.
We predict that if there are detectable differences in female cues associated
with silk, there should be differences in the intensity of courtship (i.e., rate of behaviors) for males of both species when exposed to conspecific versus heterospecific
cues. In addition, male courtship response is thought to be elicited by chemical
compounds associated with silk (contact pheromones), but this has not been confirmed using silk extractions. While unlikely, it is possible that male courtship
response in these species could be due to mechanical properties of silk. In this
study, we explored whether males are able to discriminate species-specific chemical cues associated with female silk, testing the null hypothesis of no behavioral
differences (nondiscrimination). Any significant differences in courtship intensity
would suggest species discrimination and could indicate species recognition via
chemical or mechanical cues associated with silk.
METHODS AND MATERIALS
General Methods. The wolf spider ethospecies used in this study are common in deciduous forest habitats of eastern and mid-western North America.
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All spiders used in this study were collected as juveniles or subadults in the
field and then raised to adulthood under identical laboratory conditions. Juveniles
and adult females of these species are morphologically indistinguishable. Adult
males of each species can be identified by the presence/absence of large tufts
of bristles found only on the forelegs of S. ocreata. As the experiments described here required prior knowledge of species identity, all individuals were
collected from unique species-pure localities surrounded by the zone of sympatry.
Schizocosa ocreata were collected from the Cincinnati Nature Center, Rowe
Woods, Clermont County, Ohio, and the University of Cincinnati Benedict
Nature Preserve, Hamilton County, Ohio. Schizocosa rovneri were collected from
the Ohio River flood plain at Sandy Run, Boone County, Kentucky. Individuals
were visually isolated from other spiders in opaque plastic containers (deli-dish,
10-cm diam), fed (as appropriate for their size/age) assorted collembolans, fruit
flies (Drosophila), or two–three 10-day-old cricket nymphs (Acheta domesticus)
twice weekly, and supplied with water ad libitum from a cotton wick suspended in
a water reservoir below the container. All spiders were maintained at room temperature (22–25◦ C) with stable humidity and a 13:11-hr light:dark photoperiod.
Males and females were unmated and ranged from 1 to 4-wk postfinal molt at the
time of the study.
Collection of Silk and Extraction of Chemical Cues. Female S. ocreata deposit dragline silk as they walk, so in treatments where female silk was necessary
for study, it was collected by placing single females on filter paper (Fisherbrand
90-mm diam) in a glass petri-dish (90-mm diam) allowing each to locomote for
24 hr and deposit dragline silk. Females were not used on the same day they had
been fed to reduce contamination from feces (Ayyagari and Tietjen, 1987). Fecal
material is quickly absorbed by filter paper allowing any fecal stains to easily be
excised prior to chemical extraction or use in experiments. Filter paper was handled with stainless steel forceps, and stains were excised by using a stainless steel
dissecting blade. Any filter paper disk with three or more fecal stains was not used
in this study. To reduce the likelihood of contamination, silk collection equipment
was cleaned in Alconox detergent and rinsed in double deionized water and 70%
ethanol between filter paper manipulations.
Preliminary behavioral assays were conducted with S. ocreata to identify
an effective solvent for extraction of chemical cues associated with female silk.
We exposed males (5 min) to three stimulus treatments including untreated silk
(N = 10) and silk treated in 3-min extractions with pentane (N = 12, Fisher Scientific, HPLC grade) or methanol (N = 10, Pharmco Products, Inc., HPLC grade).
Each of these solvents has been used previously to successfully extract behaviorally
active chemical components associated with spider silk (Ayyagari and Tietjen,
1987; Trabalon et al., 1997). We used total duration of active courtship (jerky tap)
exhibited by male S. ocreata on washed silk as a measure of extraction effectiveness. Methanol was the best solvent, and this result was consistent with a previous
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study in which male S. ocreata pheromone compounds were extracted (Ayyagari
and Tietjen, 1987).
For the collection of silk, 120 females (60 of each of the two ethospecies) were
used. The female stimulus treatments for the S. ocreata and S. rovneri discrimination experiments were prepared as follows; (1) Untreated Silk– filter paper disks
containing silk (and associated chemical cues) of conspecific or heterospecific females were used directly to elicit male behavior, (2) Washed Silk– following silk
collection, filter paper disks with bound silk were soaked in 50 ml of methanol for
30 min, removed from the solvent bath, and allowed to air-dry for 30 min, and (3)
Silk Extract– filter papers were soaked in 50 ml of methanol for 30 min, discarded,
and the remaining solvent allowed to completely evaporate on a clean piece of
filter paper for 30 min. Previous experiments involving the extraction of silk for
behavioral assays have used extraction periods ranging from 15 min (Jackson,
1987) to 180 min (Suter and Renkes, 1982). Ayyagari and Tietjen (1987) found
that a 15-min extraction in only 10 ml of solvent was sufficient to remove behaviorally active components from the silk of male S. ocreata, but we chose 30 min
in 50 ml of solvent (enough to completely submerge the filter paper and silk) as a
more conservative estimate to ensure more complete extraction (Suter, R. B., personal communication). The Untreated Control (4) treatment (filter paper alone)
was prepared by duplicating the silk collection treatments but without female silk,
including placing each filter paper in a glass petri-dish for 24 hr. All prepared
filter paper disks were used in behavioral trials within 4 hr of completion of the
preparation.
Behavioral Assays. We used male behavioral response to chemical and/or
mechanical stimuli as an assay of male discrimination of female species identity.
Behavioral trials were conducted during the period of peak activity for males
(1000–1600 hr, Roberts, J. A. and Uetz, G. W., unpublished data), in clear plastic
containers (13 × 7 × 7 cm) that provided a visual arena suitable for videotaping.
Filter paper disks were handled only with fine, stainless steel forceps during all
stages of pre- and posttreatment manipulation. Each disk was cut in half with
stainless steel dissecting scissors, and both halves were placed into the arena no
more than 5 min prior to the start of a trial. Males were placed gently onto the
filter paper stimulus from above and videotaped (RCA Autoshot VHS video
camera, Model CC4352) for later behavioral analysis with a software package for
the detailed analysis of behavioral data (The Observer , version 4.1). The plastic
arenas, dissecting scissors, and forceps were cleaned prior to each trial with a
Kimwipe and 70% ethanol to remove any trace of silk and pheromone deposited
in previous trials and were allowed to air-dry.
A total of 160 males (80 of each species) were chosen randomly from the
laboratory population and assigned to one of four stimulus treatment groups (described above) within the species categories S. ocreata or S. rovneri (a two-factor
ANOVA with treatment and species as main effects and individuals as replicates).
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TABLE 1. ETHOGRAM OF MALE BEHAVIORAL ELEMENTS FOR THE WOLF SPIDERS
Schizocosa ocreata AND Schizocosa rovneria
Schizocosa ocreata
Jerky tap
Tap
Schizocosa rovneri
Bounce
Shared behaviors
Chemoexplore
Leg raise
Grooming
Locomotion
Stationary
a Behavioral
Active courtship characterized by a combination of jerky forward
walking motion and tapping of the forelegs against the substrate, and
often ending with the chelicerae striking the substrate.
One or both forelegs are raised off the substrate and then lowered, lightly
striking the substrate.
While stationary, all legs are contracted simultaneously causing the
ventral surface of the body to strike the substrate.
Active exploratory behavior where the anteriolateral surfaces of the
pedipalps are brushed on the surface of the substrate in rapid
succession.
One or more legs are raised off the substrate above parallel with the body
and then lowered without striking the substrate. This is a combination
of “arch” (Stratton and Uetz, 1986) and all other leg waving.
The legs or pedipalps are drawn between the chelicerae and/or lateral
pairs of legs are brushed together rapidly.
Walking with no other elements expressed.
Motionless with no other elements expressed.
elements have been adapted from Stratton and Uetz (1986) and Delaney (1997).
Shared and species-specific male behavioral elements were scored for each trial
(Table 1). The behaviors critical in discrimination and courtship in these species
are chemoexplore (both species), jerky tap (male S. ocreata), and bounce (male
S. rovneri) (Stratton and Uetz, 1986; Stratton, 1997; see ethogram, Table 1), and
results for these behaviors are presented for each species.
Two parameters were determined for each key behavior; (1) Total Number –
a count of the number of bouts (or events) of each behavior during the trial period,
and (2) Latency – the time from introduction of the male into the arena to the initiation of a behavior (a score of 300 sec was used if a behavior was not seen during the
5-min trial period). The active courtship element of S. rovneri (“bounce”) occurs
as an event (an instantaneous behavior with each occurrence having a duration
less than 1 sec; measured in terms of rate) and not a behavioral state (behavioral
elements with a measurable duration), so parameters including a term for duration of behavior could not be calculated for S. rovneri courtship and were not
calculated for either species. Each parameter scored was square root transformed
for analysis, and for each species a separate, Bonferroni-corrected critical value
(α = 0.025) was used in significance tests of the ANOVA results to account for
multiple comparisons (Shaffer, 1995). Post hoc analyses of the preliminary assay
results and pooled data from the behavioral assays, where appropriate, were made
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with Tukey–Kramer HSD tests (Zar, 1999). All statistical analyses were performed
with JMP version 4.02 (SAS Institute).
RESULTS
Preliminary Assay. Total duration of male courtship behavior on washed silk
varied significantly with extraction treatment (ANOVA, F2,29 = 9.4, P < 0.001).
Male S. ocreata behavior on female silk washed in pentane was not significantly
different from untreated silk, but behavior was significantly reduced on silk washed
in methanol.
Schizocosa ocreata, Chemoexploratory Behavior. The total number of bouts
and latency of chemoexploratory behavior both varied significantly with stimulus
treatment, but there was no effect of species (conspecific or heterospecific source
of silk) on male behavior, and no interaction between factors (Figure 1). After
pooling the species data for each treatment group, we found that males exhibited
the most bouts of chemoexploratory behavior on untreated and washed silk, with
frequency of bouts significantly reduced on silk extract, and lowest in the control
treatments (Figure 1A). Latency to chemoexplore was shortest on untreated silk
followed by washed silk, then silk extract, and longest in the control treatment
(Figure 1B).
Schizocosa ocreata, Courtship Behavior. As in the data for chemoexploratory
behavior, the total number and latency of courtship bouts in S. ocreata varied
significantly with stimulus treatment, and there was again no significant effect of
the source of silk (species) and no significant interaction (Figure 2). The number of
bouts of jerky tap was highest on untreated silk, significantly reduced in response
to washed silk, and not different from untreated control for the extract treatment
(Figure 2A). Males began actively courting on untreated silk faster than on any of
the other stimulus treatments (Figure 2B).
Schizocosa rovneri, Chemoexploratory Behavior. For S. rovneri, both the
total number and latency of bouts of chemoexploratory behavior varied with stimulus treatment, and while there was no effect of species for either parameter, the
interaction was significant for total number of bouts of chemoexplore behavior
(Figure 3). This interaction, presumably due to response in the silk extract treatment where the species difference is not consistent with the pattern of differences
in other treatments, precluded a post hoc analysis of number of bouts of chemoexplore behavior by treatment stimulus (Figure 3A). Latency to chemoexplore was
shortest on untreated silk, intermediate on washed silk and silk extract, and longest
for the control treatment (Figure 3B).
Schizocosa rovneri, Courtship Behavior. The total number of bounce bouts
was significant by treatment, as was the latency of bounce behavior, with no effect
of species or interaction term for either measure (Figure 4). Males exhibited more
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FIG. 1. Schizocosa ocreata chemoexploratory behavior. For male spiders exposed to each
treatment within the conspecific (S. ocreata) or heterospecific (S. rovneri) species categories; (A) mean total number of behavioral bouts (+SE) (ANOVA, treatment: F3,72 = 40.2,
P < 0.001a ; species: F1,72 = 0.8, P = 0.365; interaction: F3,72 = 0.4, P = 0.748), and (B)
mean latency (s) of behavioral bouts (+SE) (ANOVA, treatment: F3,72 = 64.5, P < 0.001a ;
species: F1,72 = 0.0, P = 0.829; interaction: F3,72 = 0.7, P = 0.560) (a Indicates significance using Bonferroni-corrected critical value α = 0.025 to account for multiple comparisons). Shared letters above the bars indicate no significant difference between treatment
categories by Tukey–Kramer HSD post hoc analysis (with species category data pooled).
bounce bouts on untreated silk than on any other treatment (Figure 4A), and latency
to begin bounce behavior was shortest on untreated silk and not different from
untreated control for the other treatments (Figure 4B).
DISCUSSION
According to Dobzhansky (1970), “ethological species” are those in which
there is a breakdown in some component of mutual attraction between the sexes in
heterospecific pairings that prevents reproduction. In spiders, at least, species that
meet this definition have been called “ethospecies” (den Hollander and Dijkstra,
1974; Stratton and Uetz, 1981, 1983, 1986). Stratton and Uetz (1981) presented
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FIG. 2. Schizocosa ocreata jerky tap behavior. For male spiders exposed to each treatment
within the conspecific (S. ocreata) or heterospecific (S. rovneri) species categories; (A) mean
total number of behavioral bouts (+SE) (ANOVA, treatment: F3,72 = 33.8, P < 0.001a ;
species: F1,72 = 0.0, P = 0.999; interaction: F3,72 = 0.1, P = 0.933), and (B) mean latency
(s) of behavioral bouts (+SE) (ANOVA, treatment: F3,72 = 31.6, P < 0.001a ; species:
F1,72 = 0.0, P = 0.887; interaction: F3,72 = 0.2, P = 0.923) (a Indicates significance using Bonferroni-corrected critical value α = 0.025 to account for multiple comparisons).
Significant differences indicated as in Figure 1.
evidence of reproductive isolation between S. ocreata and S. rovneri based on
species-specific male courtship behaviors without a rigorous test of reciprocal
chemical cue discrimination. In this study, males of both S. rovneri and S. ocreata
responded equivalently with respect to courtship and exploratory behaviors on
untreated conspecific and heterospecific female silk cues. It appears that either
there are no differences between female signals, or that there are no speciesspecific differences recognizable by males of either species. Therefore, female
discrimination of species-specific male courtship appears to be the only component
of attraction preventing reproduction in these ethospecies, supporting previous
studies (Stratton and Uetz, 1981, 1983, 1986). The results of experiments presented
here also suggest that chemical, and not mechanical cues associated with silk
are critical for eliciting male courtship in both S. ocreata and S. rovneri (Uetz
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FIG. 3. Schizocosa rovneri chemoexploratory behavior. For male spiders exposed to each
treatment within the conspecific (S. rovneri) or heterospecific (S. ocreata) species categories; (A) mean total number of behavioral bouts (+SE) (ANOVA, treatment: F3,72 = 25.9,
P < 0.001a ; species: F1,72 = 1.6, P = 0.208; interaction: F3,72 = 7.0, P < 0.001a ), and
(B) mean latency (s) of behavioral bouts (+SE) (ANOVA, treatment: F3,72 = 45.0, P <
0.001a ; species: F1,72 = 1.7, P = 0.195; interaction: F3,72 = 3.0, P = 0.036) (a Indicates
significance using Bonferroni-corrected critical value α = 0.025 to account for multiple
comparisons). The significant interaction effect in (A) prohibited a post hoc analysis by
treatment category, however significant differences for (B) indicated as in Figure 1.
and Denterlein, 1979; Stratton and Uetz, 1983). Even so, while we were able to
successfully remove a compound on female silk that elicits male courtship, extracts
were not sufficient to stimulate males to court at levels equivalent to untreated silk.
There may be more than one potential explanation for this finding.
One possible explanation relates to stimulus strength. Ayyagari and Tietjen
(1987) extracted behaviorally active components from male silk in S. ocreata, but
in that study, the silk of multiple individuals was extracted simultaneously to arrive
at a product that would elicit male response. This method has the undesirable side
effect of potentially creating a “super-stimulus” (a chemical stimulus far stronger
than any that would be encountered in the natural habitat) that could result in
atypical behavioral response in stimulated individuals. In an attempt to arrive at
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FIG. 4. Schizocosa rovneri bounce behavior. For male spiders exposed to each treatment
within the conspecific (S. rovneri) or heterospecific (S. ocreata) species categories; (A) mean
total number of behavioral bouts (+SE) (ANOVA, treatment: F3,72 = 23.8, P < 0.001a ;
species: F1,72 = 0.9, P = 0.346; interaction: F3,72 = 1.0, P = 0.391), and (B) mean latency
(s) of behavioral bouts (+SE) (ANOVA, treatment: F3,72 = 17.3, P < 0.001a ; species:
F1,72 = 0.3, P = 0.599; interaction: F3,72 = 0.8, P = 0.512) (a Indicates significance using Bonferroni-corrected critical value α = 0.025 to account for multiple comparisons).
Significant differences indicated as in Figure 1.
more natural dose levels, we used extracts of the silk of single individuals. It seems
likely that the low courtship response in our study may have been due to diffusion
of a finite amount of chemical stimulus over a greater area, reducing point-specific
concentration to levels below the threshold necessary for release of courtship.
Another possibility is that there may be multiple compounds affected differently by solvent type. Ayyagari and Tietjen (1987) used methanol to extract a
behaviorally active component from male S. ocreata silk, and in part for this reason
methanol was selected as a potential solvent for female chemical compounds in
the work presented here. In previous attempts to study potential pheromones associated with the silk of female wolf spiders, several organic and inorganic solvents
were found to produce a pattern of inactivation similar to that found using methanol
in the current study (Kaston, 1936; Hegdekar and Dondale, 1969). This evidence
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is suggestive that male courtship response is elicited by multiple compounds that
are differentially extracted in various solvents but are necessary in combination to
elicit courtship.
In contrast to the results for male courtship, males exhibited a similar number of bouts of chemoexploratory behavior on untreated silk and silk washed in
methanol (Figures 1 and 3). This suggests that the chemical compound(s) that elicits courtship behavior plays only a limited role for release of chemoexploratory
behavior. Either there is a separate compound associated with silk that releases
chemoexploratory behavior but is not extracted by methanol, or the term “chemo”
exploratory behavior may be a misnomer for these species. Wolf spiders are known
to have chemosensory sensilla concentrated on the anterior dorsal and lateral surfaces of the pedipalps (Kronestedt, 1979; Tietjen and Rovner, 1982; Barth, 2002),
which are actively rubbed on the substrate during exploratory behavior presumably to collect chemical information (Tietjen, 1977; Stratton and Uetz, 1986). In
another study of S. ocreata (Roberts, J. A. and Uetz, G. W., unpublished data),
total number of bouts of chemoexploratory behavior was similar on the silk of
female S. crassipes, a member of the ocreata clade along with S. ocreata and
S. rovneri (Miller et al., 1998), but decreased in species outside the clade despite
the presence of silk. Together this evidence suggests that a chemical associated
with silk elicits chemoexploration, and that this compound may be independent
of, or redundant to, the chemical that releases courtship behavior. Of note is the
unexpected result for S. rovneri males of greater number of bouts of chemoexploratory behavior on extract of heterospecific silk, compared to conspecific extract
(Figure 3A). This suggests that there may be slight differences in the composition
or ratios of chemical compounds on the silk of these species, but those differences
are not behaviorally relevant on untreated silk. Further chemical analysis will be
necessary to fully explain this difference.
Acknowledgments—This work was supported by the American Arachnological Society (JAR),
the University of Cincinnati Research Council and Department of Biological Sciences (JAR), and the
National Science Foundation (IBN 9906446 and IBN 9414239 to GWU). This research was submitted
in partial fulfillment of the requirements for completion of the PhD in Biological Sciences at the
University of Cincinnati. We thank the Cincinnati Nature Center, Rowe Woods, and the residents of
Sandy Run, KY, for permission to collect spiders on their properties. Voucher specimens are on deposit
at the National Museum of Natural History in Washington, DC, and in the personal collection of
the corresponding author (JAR). We are especially grateful to J. Rovner (for access to his reference
collection), and K. Roberts, A. Cady, C. Harris, J. Hinn, B. Jayne, K. Petren, M. Polak, J. Shann, and
S. Walker for their advice and editorial comments.
REFERENCES
AYYAGARI, L. R. and TIETJEN, W. J. 1987. Preliminary isolation of male-inhibitory pheromone of the
spider Schizocosa ocreata (Araneae, Lycosidae). J. Chem. Ecol. 13:237–245.
P1: JLS
Journal of Chemical Ecology [joec]
pp1208-joec-487114
CHEMICAL SIGNALING IN A WOLF SPIDER
May 22, 2004
9:35
Style file version June 28th, 2002
1283
BARTH, F. G. 2002. A Spider’s World: Senses and Behavior. Springer, Heidelberg, Germany.
CADY, A. B. 1984. Microhabitat selection and locomotor activity of Schizocosa ocreata (Walckenaer)
(Araneae: Lycosidae). J. Arachnol. 11:297–307.
COSTA, F. G. and CAPOCASALE, R. M. 1984. Lycocsa carbonelli, sp. nov.: una etoespecie simpatrida,
sibilina de Lycosa thorelli (Keyserling) (Araneae, Lycosidae). J. Arachnol. 11:423–431.
DELANEY, K. J. 1997. Communication in the contexts of courtship and aggression in two species on
wolf spider (Araneae: Lycosidae). MS Dissertation, University of Cincinnati, Cincinnati.
DEN HOLLANDER, J. 1971. Species barriers in the Pardosa pullata group/Araneae, Lycosidae/. Proceedings of 5th International Arachnological Congress, Brno, pp. 129–142.
DEN HOLLANDER, J. and DIJKSTRA, H. 1974. Pardosa vlijmi sp. Nov., a new ethospecies sibling Pardosa proxima (C. L. Koch, 1948), from France, with description of courtship display (Araneae,
Lycosidae). Beaufortia 22:57–65.
DEN HOLLANDER, J., DIJKSTRA, H., ALLEMAN, H., and VLIJM, L. 1973. Courtship behaviour as species
barrier in the Pardosa pullata group (Araneae, Lycosidae). Tijdschr. Entomol. 116:1–22.
DOBZHANSKY, T. 1970. Genetics of the Evolutionary Process. Columbia University Press, New York.
HEGDEKAR, B. M. and DONDALE, C. D. 1969. A contact sex pheromone and some response parameters
in lycosid spiders. Can. J. Zool. 47:1–4.
JACKSON, R. R. 1987. Comparative study of releaser pheromones associated with silk of jumping
spiders (Araneae, Salticidae). N. Z. J. Zool. 14:1–10.
KASTON, B. J. 1936. The senses involved in the courtship of some vagabond spiders. Entomol. Am.
16:97–167.
KRONESTEDT, T. 1979. Study on chemosensitive hairs in wolf spiders (Araneae, Lycosidae) by scanning
electron microscopy. Zool. Scr. 8:279–285.
MCCLINTOCK, W. J. and UETZ, G. W. 1996. Female choice and pre-existing bias: visual cues during
courtship in two Schizocosa wolf spiders (Araneae: Lycosidae). Anim. Behav. 52:167–181.
MILLER, G. L., STRATTON, G. E., MILLER, P. R., and HEBETS, E. 1998. Geographic variation in male
courtship behaviour and sexual isolation in wolf spiders of the genus Schizocosa. Anim. Behav.
56:937–951.
PAPKE, M. D., RIECHERT, S. E., and SCHULZ, S. 2001. An airborne female pheromone associated with
male attraction and courtship in a desert spider. Anim. Behav. 61:877–886.
POLIS, G. A. and MCCORMICK, S. J. 1986. Scorpions, spiders, and solpugids: Predation and competition
among distantly related taxa. Oecologia 71:111–116.
POLLARD, S. D., MACNAB, A. M., and JACKSON, R. R. 1987. Communication with chemicals:
Pheromones and spiders, pp. 133–141, in W. Nentwig (ed.). Ecophysiology of Spiders. Springer,
Berlin.
PROUVOST, O., TRABALON, M., PAPKE, M., and SCHULZ, S. 1999. Contact sex signals on web and
cuticle of Tegenaria atrica (Araneae, Agelenidae). Arch. Insect Biochem. Physiol. 40:194–202.
RIECHERT, S. E. and SINGER, F. D. 1995. Investigation of potential male mate choice in a monogamous
spider. Anim. Behav. 49:715–723.
ROSS, K. and SMITH, R. L. 1979. Aspects of the courtship behavior of the black widow spider, Latrodectus hesperus (Araneae: Theridiidae), with evidence for the existence of a contact sex pheromone.
J. Arachnol. 7:69–77.
SEARCY, L. E., RYPSTRA, A. L., and PERSONS, M. H. 1999. Airborne chemical communication in the
wolf spider Pardosa milvina. J. Chem. Ecol. 25:2527–2533.
SHAFFER, J. P. 1995. Multiple hypothesis testing. Annu. Rev. Psychol. 46:561–584.
SINGER, F., RIECHERT, S. E., XU, H., MORRIS, A. W., BECHER, E., HALE, J. A., and NOUREDDINE,
M. A. 2000. Analysis of courtship success in the funnel-web spider Agelenopsis aperta. Behaviour
137:93–117.
STRATTON, G. E. 1997. Investigation of species divergence and reproductive isolation of Schizocosa
stridulans (Araneae: Lycosidae) from Illinois. Bull. Br. Arachnol. Soc. 10:313–321.
P1: JLS
Journal of Chemical Ecology [joec]
1284
pp1208-joec-487114
May 22, 2004
9:35
Style file version June 28th, 2002
ROBERTS AND UETZ
STRATTON, G. E. and UETZ, G. W. 1981. Acoustic communication and reproductive isolation in two
species of wolf spiders. Science 214:575–577.
STRATTON, G. E. and UETZ, G. W. 1983. Communication via substratum-coupled stridulation and
reproductive isolation in wolf spiders (Araneae: Lycosidae). Anim. Behav. 31:164–172.
STRATTON, G. E. and UETZ, G. W. 1986. The inheritance of courtship behavior and its role as a
reproductive isolating mechanism in two species of Schizocosa wolf spiders (Araneae; Lycosidae).
Evolution 40:129–141.
SUTER, R. B. and RENKES, G. 1982. Linyphiid spider courtship: releaser and attractant functions of a
contact sex pheromone. Anim. Behav. 30:714–718.
TIETJEN, W. J. 1977. Dragline-following by male lycosid spiders. Psyche 84:165–178.
TIETJEN, W. J. 1979. Is the sex pheromone of Lycosa rabida (Araneae: Lycosidae) deposited on a
substratum? J . Arachnol. 6:207–212.
TIETJEN, W. J. and ROVNER, J. S. 1982. Chemical communication in lycosids and other spiders, pp.
249–279, in P. N. Witt and J. S. Rovner (eds.). Spider Communication. Mechanisms and Ecological
Significance. Princeton University Press, Princeton, NJ.
TRABALON, M., BAGNERES, A. G., and ROLAND, C. 1997. Contact sex signals in two sympatric spider
species, Tegenaria domestica and Tegenaria pagana. J. Chem. Ecol. 23:747–758.
UETZ, G. W. 2000. Signals and multi-modal signaling in spider communication, pp. 387–405, in Y.
Espmark, T. Amundsen, and G. Rosenqvist (eds.). Animal Signals: Signalling and Signal Design
in Animal Communication. Tapir Academic Press, Trondheim, Norway.
UETZ, G. W. and DENTERLEIN, G. 1979. Courtship behavior, habitat, and reproductive isolation in
Schizocosa rovneri Uetz and Dondale (Araneae: Lycosidae). J. Arachnol. 7:121–128.
UETZ, G. W. and ROBERTS, J. A. 2002. Multisensory cues and multimodal communication in spiders:
Insights from video/audio playback studies. Brain Behav. Evol. 59:222–230.
WISE, D. H. and CHEN, B. 1999. Impact of intraguild predators on survival of a forest-floor wolf spider.
Oecologia 121:129–137.
ZAR, J. H. 1999. Biostatistical Analysis, 4th edn. Prentice Hall, Upper Saddle River, NJ.