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RESEARCH ARTICLE
Attachment performance of stick insects (Phasmatodea) on
convex substrates
Thies H. Bu
̈scher*, Martin Becker and Stanislav N. Gorb
ABSTRACT
Phasmatodea (stick and leaf insects) are herbivorous insects well
camouflaged on plant substrates as a result of cryptic masquerade.
Also, their close association with plants has allowed them to adapt to
different substrate geometries and surface topographies of the plants
they imitate. Stick insects are gaining increasing attention in
attachment- and locomotion-focused research. However, most
studies experimentally investigating stick insect attachment have
been performed either on single attachment pads or on flat surfaces.
In contrast, curved surfaces, especially twigs or stems of plants, are
dominant substrates for phytophagous insects, but not much is
known about the influence of curvature on their attachment. In this
study, by combining analysis of tarsal usage with mechanical traction
and pull-off force measurements, we investigated the attachment
performance on curved substrates with different diameters in two
species of stick insects with different tarsal lengths. We provide the
first quantitative data for forces generated by stick insects on convex
curved substrates and show that the curvature significantly influences
attachment ability in both species. Within the studied range of
substrate curvatures, traction force decreases and pull-off force
increases with increasing curvature. Shorter tarsi demonstrate
reduced forces; however, tarsus length only has an influence for
diameters thinner than the tarsal length. The attachment force
generally depends on the number of tarsi/tarsomeres in contact,
tarsus/leg orientation and body posture on the surface. Pull-off force
is also influenced by the tibiotarsal angle, with higher pull-off force for
lower angles, while traction force is mainly influenced by load, i.e.
adduction force.
KEY WORDS: Adhesion, Friction, Biomechanics, Euplantula,
Arolium, Tarsal pads
INTRODUCTION
Attachment during locomotion on different kinds of surfaces is a
widespread phenomenon in the animal kingdom. Especially in
insects, various types of reversible attachment structures, which
allow them to hold themselves and to walk onthe majority of natural
surfaces (Beutel and Gorb, 2001, 2006, 2008), evolved in response
to the great variability of natural surfaces (Gorb, 2001; Gorb and
Gorb, 2004). Phasmatodea Jacobson and Bianchi 1902, or stick and
leaf insects, are a mesodiverse group of large terrestrial insects
(Bradler and Buckley, 2018) inhabiting various habitats worldwide
(Bedford, 1978; Brock et al., 2019). These herbivorous insects are
strongly adapted to foraging on plants, e.g. exhibiting extreme forms
of masquerade crypsis (Robertson et al., 2018), and presumably
have co-evolved with plants since pre-angiosperm times (Wang
et al., 2014). This process shaped the evolution of the attachment
apparatus of phasmids and resulted in diverse adaptations to
different surfaces in their habitats (Büscher et al., 2019). As a result,
the tarsal attachment apparatus of most stick insects consists of a
flexible chain of five tarsomeres, bearing two types of attachment
pads and two claws. The tarsal euplantulae, present on the proximal
four/five tarsomeres, are used to generate friction and withstand
shear forces, and the pretarsal arolium provides adhesion to
withstand pull-off forces (Labonte and Federle, 2013). On smooth
or micro-rough surfaces, these pads provide attachment due to
adhesion and friction forces, whereas on rough surfaces the claws
additionally achieve mechanical interlocking (Büscher and Gorb,
2019). Species-specific functional attachment microstructures
(AMS) are present on the euplantulae of stick insects and
correlate with the different substrates in their natural habitats
(Büscher and Gorb, 2017; Büscher et al., 2018a,b, 2019). These
structures are reported for at least one species to change during post-
embryonic development, in which the animals change their habitat
(Gottardo et al., 2015). Previous studies on the attachment
performance of stick insects have shown that euplantulae with
smooth AMS generate higher forces on smooth surfaces, whereas
nubby euplantulae are adapted to micro-rough surfaces (Bußhardt
et al., 2012; Büscher and Gorb, 2019).
Most studies on the attachment properties of stick insects have
focused on the performance of single attachment pads (Bennemann
et al., 2011; Bußhardt et al., 2012; Labonte and Federle, 2013;
Labonte et al., 2014) and examined the adhesive and frictional
components of attachment in detail. Other studies, focusing on the
attachment performance of whole stick insects, investigated the
combination of the different pads and claws and their contribution to
the overall attachment (Büscher and Gorb, 2019; Labonte et al.,
2019). In regard to measurements of actual attachment forces, these
studies are limited to investigations of (1) size dependence of
attachment in different instars of one species ofstick insects (Labonte
et al., 2019) and (2) complementary effects of different components
of the attachment apparatus in two other species (Büscher and Gorb,
2019). Labonte et al. (2019) have shown that during the ontogeny of
stick insects, adhesion scales with contact area in whole-animal
measurements, as attachment performance compensates for weight in
larger stick insects by the weight-related shear forces produced by
sliding of the pads. Büscher and Gorb (2019) investigated the use of
all tarsal attachment structures in two species and showed a flexible
usage of the attachment pads according to the animal’s orientation to
the substrate. This study revealed stronger attachment forces on rough
substrates by the contribution of claws and on smooth substrates by
the contribution of tarsal attachment pads.
Although they provide insight into the functionality of the whole
attachment system, all these studies have been carried out on flat
Received 8 April 2020; Accepted 20 July 2020
Department of Functional Morphology and Biomechanics, Institute of Zoology, Kiel
University, Am Botanischen Garten 9, 24118 Kiel, Germany.
*Author for correspondence (tbuescher@zoologie.uni-kiel.de)
T.H.B., 0000-0003-0639-4699; S.N.G., 0000-0001-9712-7953
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© 2020. Published by The Company of Biologists Ltd
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Journal of Experimental Biology (2020) 223, jeb226514. doi:10.1242/jeb.226514
Journal of Experimental Biology
surfaces. In nature, stick insects face a lot of differently structured
surfaces. In particular, curved surfaces, such as branches or twigs of
plants, are among the most abundant substrates for stick and leaf
insects (Bedford, 1978; Gottardo et al., 2015; Büscher et al., 2019)
and have even shaped the appearance of most stick insects (Bedford,
1978; Wang et al., 2014; Bradler and Buckley, 2018). Locomotion
on curved substrates implicates different mechanics in comparison
to upright walking on flat substrates. Previous studies on other
insects have examined some aspects of locomotion on twigs and
curved substrates. Studies of different insect species walking on thin
stems revealed several strategies of attachment, including the use of
pads, claws and additional structures on the tibia enabling
attachment to and propulsion along the stems (Gladun and Gorb,
2007; Bußhardt et al., 2014). Measurements of attachment forces
showed remarkable differences between the maximal forces on
curved surfaces and flat ones for beetles and true bugs (Bußhardt
et al., 2014; Voigt et al., 2017, 2019). In these studies, the measured
attachment forces were higher on rods in comparison to flat plates of
the same material. The attachment performance on curved substrates
is different for different radii of curvature in relation to the
dimensions of the animal, as it has been shown for frogs that traction
is dependent on the diameter of the cylinder the frog tries to ascend
(Bijma et al., 2016; Hill et al., 2018).
On smooth flat surfaces, it is known that wide tarsi generate
higher attachment forces than slender tarsi in Stenus rove beetles
(Betz, 2002), demonstrating an influence of tarsus geometry on
attachment performance. The performance on different diameters of
curved substrates should be dependent on the length of the
attachment system instead. The length of the tarsal chain might
have an effect on how many attachment pads can be brought into
contact with a curved substrate and can further influence the
orientation of the attachment pads on the substrate. Friction
experiments on the euplantulae of two species of stick insects
revealed frictional anisotropy for both smooth and nubby
euplantulae (Bußhardt et al., 2012). At a load of 500 µN, the
species Medauroidea extradentata and Carausius morosus both
show higher friction in the proximal direction (pull) than in the
distal direction (push). The diameter of the structure in relation to
the length of the tarsal chain determines how the tarsi can be placed
on it (Gladun and Gorb, 2007; Voigt et al., 2017) and, hence,
influences the orientation of the animal and the direction of the
forces acting on the attachment system.
In this study, we examined the attachment capability of stick
insects on surfaces with different levels of curvature. We measured
the pull-off and traction forces of the species Sungaya inexpectata
and Orestes mouhotii. These two species differ primarily in body
size and in the length of their tarsi (Büscher et al., 2019).
Specifically, we aimed to answer the following questions. (1) How
does the attachment system of stick insects perform in traction and
pull-off on curved substrates with different diameters? (2) Does the
length of the tarsus influence attachment ability on surfaces with
different degrees of curvature? (3) How do stick insects use the
tarsal attachment system during locomotion on curved substrates?
MATERIALS AND METHODS
Animals
The species Sungaya inexpectata Zompro 1996 and Orestes
mouhotii (Bates 1865) were chosen (Fig. 1), as their tarsi differ in
the length of the tarsal chain, but not in either their overall tarsal
morphology (see Fig. 2) or their euplantular microstructures
(Büscher et al., 2019). Females were selected to avoid differences
caused by sexual dimorphism. All animals examined in this study
were obtained from the laboratory cultures of the Department of
Functional Morphology and Biomechanics (Kiel University,
Germany). They were fed with leaves of bramble (Rubus spp.)
and hazel (Corylus avellana)ad libitum. All animals retained a
natural day/night cycle. Adult individuals with six intact legs and no
visible damage on the attachment pads (using light microscopy)
were used. All insects were weighed prior to the measurements to
the nearest 100 µg (AG204 Delta Range, Mettler Toledo,
Columbus, OH, USA). We used the same specimens for pull-off
and traction force measurements and measured their mass on the
particular day of the measurement. As the mass of the specimens
changed over the course of the experiment, the values reported
below correspond to the masses of the individual insects at the time
of the measurements.
Scanning electron microscopy
Tarsi of both species were cut off from adult females and fixed in
2.5% glutaraldehyde in PBS buffer for 24 h on ice on a shaker.
Afterwards, samples were dried in an ascending ethanol series,
critical-point dried and sputter-coated with a 10 nm layer of gold-
palladium. The samples were mounted on a rotatable specimen
holder (Pohl, 2010) and overview images were obtained using a
scanning electron microscope (TM3000, Hitachi High-technologies
Corp., Tokyo, Japan) at 15 kV acceleration voltage. The
micrographs were processed and measurements of the tarsal
segments were conducted using the software Photoshop CS6
(Adobe Systems Inc., San Jose, CA, USA). The tarsi were
visualized from the lateral view to measure the length of all
tarsomeres (see Fig. 2).
Substrate preparation
As substrates, we used custom-made tubes and plates of acrylic
glass. We used four tubes with diameters of 24.8, 11.9, 5 and
2.9 mm to provide substrates that correspond to different abilities of
the animals to embrace the tubes. These cylinders correspond to the
different geometric characteristics of twigs in the natural habitats,
O. mouhotii
S. inexpectata
Fig. 1. Species examined in this study. Adult females of Sungaya
inexpectata and Orestes mouhotii. Scale bar: 2 cm.
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RESEARCH ARTICLE Journal of Experimental Biology (2020) 223, jeb226514. doi:10.1242/jeb.226514
Journal of Experimental Biology
but exclude parameters influencing attachment, like differing
roughness, stiffness or surface chemistry. For comparison, we
used a flat square plate of acrylic glass. All substrates were cleaned
with ethanol prior to the experiment. During the experiments, the
substrates were cleaned between measurements to remove potential
residuals of attachment pad fluids from the substrates. All substrates
had a length of 25 cm, in order to provide sufficient latitude for
horizontal pulling.
Force measurements
Attachment forces were measured with a BIOPAC model MP100
and TCI-102 system (BIOPAC Systems, Inc., Goleta, CA, USA),
connected to a 100 gforce transducer (FORT100, World Precision
Instruments, Sarasota, FL, USA). A horse hair was attached to the
force transducer and glued with Plasticine to the metanotum of the
stick insect. The attachment forces of the insects were measured in
the following two directions: vertically (pull-off force) and
horizontally (traction force).
Pull-off forces
Specimens of both species (S. inexpectata 1.18±0.26 g mean±s.d.,
N=14; O. mouhotii 0.91±0.22 g, N=14) were attached to the force
transducer as described above. To pull the insects off the substrate at
an angle of 90 deg, the force transducer was mounted on a
micromanipulator (DC 3001R, World Precision Instruments Inc.)
with the same setup described by Wohlfart et al. (2014) and
employed for stick insects by Büscher and Gorb (2019). One glass
tube at a time, or the flat plate, respectively, was fixed below the
animal using two clamps. The micromanipulator was manually
moved upwards, with a retraction velocity of 2–4cms
−1
, and
stretched the horse hair in the vertical direction until the animal
detached from the surface (see Fig. 3B).
Traction forces
Specimens of both species (S. inexpectata 1.32±0.31 g, N=14;
O. mouhotii 1.05±0.13 g, N=14) were attached to the force transducer
in the same manner as described above. To measure passive traction
force, the force transducer was aligned horizontally at the same height
as the insect (see Wolff and Gorb, 2012; Büscher and Gorb, 2019).
Each insectwasthen manually pulled backwards (along its bodyaxis)
in the traction direction without detaching it from the surface with a
retraction velocity of 2–5cms
−1
(see Fig. 3A). As both species
preferred to walk upright on the flat surface and on the tube with the
largest diameter, but walked hanging upside down on the other tubes,
the traction forces were measured in accordance with the naturally
preferred animal orientation (see below).
O. mouhotii S. inexpectata
Total length (mm)
2
3
4
5
6
7
8
9*
Ta1
Ta2 Ta3 Ta 4
Ta5
Pt
Eu1 Cl
Ar
Cl
Cl
Ar
Cl
Eu2 Eu3 Eu4 Eu5
Eu1 Eu2 Eu3 Eu4 Eu5
A
B
C
D
E
F
Fig. 2. Tarsal morphology of the examined species. (A–C) Scanning electron micrographs of the right metatarsus of an adult female O. mouhotii (A,B) and
Sungaya inexpectata (C) (A, lateral overview; B,C, ventral overviews). (D) Total tarsal length of the two species ( paired t-test, t=−24.26, d.f.=10, N
1,2
=11,
P<0.001). Boxes indicate the 25th and 75th percentiles, whiskers are the 10th and 90th percentiles and the line within the boxes shows the median. (E,F) The
nubby microstructure on the euplantulae (4th euplantula) of the right metatarsus of an adult female O. mouhotii (E) and S. inexpectata (F). Ar, arolium; Cl, claw;
Eu, euplantula; Pt, pretarsus; Ta, tarsomere. *P<0.001. Scale bars: 1 mm (A–C), 4 µm (E,F).
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RESEARCH ARTICLE Journal of Experimental Biology (2020) 223, jeb226514. doi:10.1242/jeb.226514
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Force data analysis
In both directions, force–time curves were recorded and visualized
with the software Acqknowledge 3.7.0 (BIOPAC Systems Inc.,
Goleta, CA, USA). The maximum peak of the force–time curve was
extracted for data analysis (see Fig. 3C,D for example curves). For
every direction and every species, the attachment forces for 14
individuals were measured. Every insect was measured 3 times and
the median of all three maximum attachment forces was taken. The
order of surfaces was randomized for each animal and the order of
measured individuals was also randomized during the experiment.
All measurements were performed in daylight at 19–21°C and 50–
60% relative humidity. Additionally, the behaviour of animals was
documented during the measurements.
Videography
To evaluate walking behaviour and leg positioning, adult females of
both species were filmed on each surface (S. inexpectata 1.24±
0.27 g, N=10; O. mouhotii 1.04±0.13 mg, N=10). The animals were
filmed with a Nikon D5300 digital camera (Nikon Corp., Tokyo,
Japan) equipped with a macro lens (Canon Macro Lens EF 100 mm,
Canon Inc., Tokyo, Japan) from the lateral side on all substrates. We
analysed (1) the walking gait pattern, (2) the orientation of the
specimen in relation to the substrate and (3) the positioning of the
tarsi. Therefore, sequences of at least 10 steps were analysed and
sequences with disruptions (turns, falls, etc.) were excluded. For the
flat substrate, only sequences where the animal walked in a straight
line were used. For analysis of the videos, Adobe Premiere Pro CS6
software (Adobe Systems Inc.) was used. To evaluate the walking
gait patterns, three general patterns were identified (tripod,
quadrupedal/tetrapod, waive gait; see Grabowska et al., 2012;
Büscher and Gorb, 2019), based on the number of feet
simultaneously in contact with the substrate, with a maximum of
10% of steps deviating from the ground pattern. If more than 10% of
steps were observed differing from the ground pattern (e.g.
transitions between two gait patterns), we considered these gait
pattern sequences as irregular.
Statistical analysis
SigmaPlot 12.0 (Systat Software Inc., San Jose, CA, USA) was used
to perform the statistical analyses. Initially, normal distribution
(Shapiro–Wilk test) and homoscedasticity (Levene’s test) were
tested. The tarsal length and the body weight of both species were
compared using a two-tailed t-test. The traction and pull-off forces
on the five different substrate radii were compared for both species
separately using one-way ANOVA and Holm–Šidák’spost hoc test,
if the data were normally distributed and showed homoscedasticity.
Kruskal–Wallis ANOVA on ranks and Tukey’spost hoc tests were
used if the data were not normally distributed. The safety factors
(attachment force per body mass) of the two species on substrates
with different radii were compared using t-test. For better
understanding, the exact test used and the sample size are
specified in the relevant sections.
RESULTS
Tarsal morphology
As for the majority of extant Phasmatodea, the tarsi of both species
examined consisted of five tarsomeres (see Büscher et al., 2019),
each equipped with one euplantula (Fig. 2A–C). The pretarsus bears
two claws and an arolium. The tarsi of S. inexpectata were
significantly longer than the tarsi of O. mouhotii ( paired t-test,
t=−24.26, d.f.=10, N
1,2
=11, P<0.001). The measurements of the
length of single tarsomeres and the total length of the tarsus in all
specimens are available in Table S1. We did not find significant
differences in the total tarsal length between different leg pairs in the
d1
d2
d3
d4
d5
d=24.8 mm
d=11.9 mm
d=5.0 mm
d=2.9 mm
Flat
A
B
EC
D
Time (s)
mtf
mpf
0 5 10 15 20 25 30
0
0
20
40
60
80
100
120
0
50
100
150
200
5 101520253035
Traction force (mN)Pull-off force (mN)
Fig. 3. Experimental setup for traction and pull-off force measurements. (A,B) Schematic setup of force measurements in the traction (A) and
pull-off (B) direction.(C,D) Example curves for traction (C) and pull-off force (D).(E) Cross-sections of the flat plate (d
1
) and cylindrical substrates used in the
experiments: 24.8 mm (d
2
), 11.9 mm (d
3
), 5 mm (d
4
) and 2.9 mm diameter (d
5
) tube. mtf, maximum traction force; mpf, maximum pull-off force.
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RESEARCH ARTICLE Journal of Experimental Biology (2020) 223, jeb226514. doi:10.1242/jeb.226514
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two species examined (see Table S2; S. inexpectata: Kruskal–Wallis
one-way ANOVA; H=2.455, d.f.=2; P=0.293, N=15; O. mouhotii:
Kruskal–Wallis one-way ANOVA; H=0.0614, d.f.=2, P=0.97,
N=15). The total length of the S. inexpectata tarsus (all three leg
pairs pooled) was 7.87±0.36 mm (median±s.d.) and 3.47±0.34 mm
for O. mouhotii (see Fig. 2). The major difference in the
tarsal morphology between both species was the length of the
single tarsomeres. In particular, the basitarsus (Ta1) was longer in
S. inexpectata (2.91±0.13 mm) than in O. mouhotii (1.0±0.1 mm).
Both species possess a nubby microstructure on the euplantulae (see
Fig. 2), as previously reported (e.g. Büscher et al., 2019). Although
the length of the tarsi of the same leg pairs differed between the
species, the dimensions of the surface microstructure on the
euplantulae were similar (Fig. 2E,F).
Locomotion on curved substrates
The locomotory behaviour on the test substrates differed depending
on the degree of their curvature. All tested individuals were analysed
to obtain the following data: (1) walking gait pattern, (2) orientation
on the substrate, (3) angle of attachment (AoA) and (4) leg
configuration (see Fig. 4). The AoA is defined by the centre of the
animal in the frontal projection and the distal-most part of the tarsus
being in contact with the substrate (Fig. 4A). Both species showed
rather similar behaviour on each test substrate (Table 1). On nearly
all substrates, insects employed a tripod walking gait pattern (three
legs at once in swing phase). Only on the tube with the smallest
diameter did O. mouhotii walk more slowly and just moved two legs
at once (tetrapod gait) in 80% of the recorded sequences. The same
species used tripod gait in 100% of the sequences on all other
substrates except the tube with the largest diameter, where it used
tetrapod gait in 25% of the sequences. Sungaya inexpectata used
tripod gait less often, but still in the majority of sequences (Table 1).
On the smallest tube (2.9 mm diameter), S. inexpectata moved more
carefully and used tripod gait in only 40% of the sequences and
tetrapod gait in all of the others. On both the 5.0 and 24.8 mm tubes,
tripod gait was used in 90% of the sequences and on the flat
substrate in 60% of the sequences. All S. inexpectata sequences
which did not show tripedal walking gait patterns revealed
tetrapedal gait patterns.
The same orientation to the substrate was recorded in all
individuals of both species. On the 24.8 mm tube, both species
walked upright on the tube, attaching at an AoA of <90 deg
(Fig. 4B). On the three other tubes, both species walked hanging
upside down. This agrees to observations on the species Aretaon
asperrimus (Frantsevich and Cruse, 1997), which is quite closely
related to the species examined herein (Brock et al., 2019). While S.
A
B
d=24.8 mm d=11.9 mm d=5.0 mm d=2.9 mm
d=24.8 mm d=11.9 mm d=5.0 mm d=2.9 mm
C
E
G
D
F
H
<90 deg >90 deg >180 deg >180 deg
<90 deg <90 deg =90 deg >90 deg
Fig. 4. Posture and leg conformations of both species on the different substrates. (A,B) Body and leg positioning on the different substrate diameters (d)in
(A) O. mouhotii (short tarsus) and (B) S. inexpectata (long tarsus). AoA, angle of attachment, i.e. angle between the centre of the animal (in frontal
projection) and the distal-most part of the tarsus (indicated below each animal scheme). (C–H) Images of example tarsal contacts taken in experiments:
S. inexpectata on d=2.9 mm (C); O. mouhotii on d=2.9 mm (D), d=5.0 mm (E), d=11.9 mm (F,G); and (H) S. inexpectata on d=11.9 mm.
Table 1. Walking pattern and body posture on the test substrates
Substrate diameter (mm)
Sungaya inexpectata Orestes mouhotii
Gait pattern Body posture Gait pattern Body posture
2.9 Tripod (40%) Tetrapod (60%) Hanging down Tetrapod (80%) Irregular (20%) Hanging down
5.0 Tripod (90%) Tetrapod (10%) Hanging down Tripod (100%) Hanging down
11.9 Tripod (100%) Hanging down Tripod (100%) Hanging down
24.8 Tripod (90%) Tetrapod (10%) Standing Tripod (75%) Tetrapod (25%) Standing
Plane Tripod (60%) Tetrapod (40%) Standing Tripod (100%) Standing
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RESEARCH ARTICLE Journal of Experimental Biology (2020) 223, jeb226514. doi:10.1242/jeb.226514
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inexpectata attached at the downside of the 11.9 mm tube at an AoA
of <90 deg, O. mouhotii revealed AoAs of >90 deg on the same
tube. On the smaller tubes, O. mouhotii embraced the tubes with
higher AoAs than S. inexpectata as well. While O. mouhotii
attached with AoAs of >180 deg on both 5.0 and 2.9 mm diameter
tubes, S. inexpectata revealed AoAs of 90 deg on 5.0 mm and
>90 deg but <180 deg on the 2.9 mm tube. While hanging down,
both species attached with AoAs of >90 deg on all substrates with
one exception: on the tube with a diameter of 11.9 mm,
S. inexpectata did not embrace the tube, but attached hanging on
the downside. Although the animals kept some distance from the
tube while hanging down, their legs were still angled (tibiofemoral
joint 90–160 deg). In this posture, only the arolia were employed for
attachment (Fig. 4H). In all other situations in which the animals
were hanging upside down (11.9 mm for O. mouhotii, 5.0 and
2.9 mm for both species), the insects embraced the tubes with AoAs
of 90 deg or more. In these postures, animals employed all
attachment pads (arolia and euplantulae), except for the fifth
euplantula (see Fig. 2A–C), which was used in only a few
sequences: on the 5 and 2.9 mm substrates by S. inexpectata and
only on the 2.9 mm substrate by O. mouhotii (Fig. 4C–G). The four
proximal euplantulae were used on all curved substrates by both
species, except for all individuals of S. inexpectata, which used the
arolium only on the 11.9 mm substrate (Fig. 4H) and the arolium
and the distal-most three euplantulae on the 2.9 mm substrate.
Generally, on the flat substrate, both species brought only the
arolium and euplantulae of the third and fourth tarsomeres into
contact.
Traction forces
The traction forces, measured via horizontal pulls, revealed lower
values for smaller tube diameters for both species. Generally,
S. inexpectata showed the highest traction force (Fig. 5C) on the
24.8 mm diameter tube (272.38±111.40 mN , median±s.d.), lower
force on the flat substrate (230.05±97.87 mN) and further
decreasing values on decreasing tube diameters (11.9 mm:
220.17±77.92 mN, 5.0 mm: 146.07±53.67 mN, 2.9 mm: 126.97±
67.06 mN). Statistical analysis, however, revealed significant
differences primarily between the traction forces on larger tube
diameters and the flat substrate in comparison with smaller tube
diameters (one-way ANOVA, F=7.3, N=14, P<0.001). In detail, the
traction forces on the smallest tube (2.9 mm diameter) were
significantly lower than those on the flat substrate (Holm–Šidák
test, P=0.015), the 24.8 mm diameter tube (Holm–Šidák test,
P<0.001) and the 11.9 mm diameter tube (Holm–Šidák test,
P=0.032), but not different from those on the 5.0 mm diameter
tube (Holm–Šidák test, P=0.798). The traction forces generated on
the 5.0 mm diameter tube were significantly lower in comparison to
those on the largest tube (Holm–Šidák test, P=0.002).
A similar trend was observed in O. mouhotii (Fig. 5A), with the
highest mean traction forces of 210.21±71.96 mN on the 24.8 mm
diameter tube, 188.28±90.47 mN on the flat substrate, 177.97±
d1
d2
d3
d4
d5
d=24.8 mm
d=11.9 mm
d=5.0 mm
d=2.9 mm
Flat
E
A
BD
C
Traction force (mN)
d1d2d3d4d5
d1d2d3d4d5
d1d2d3d4d5
d1d2d3d4d5
Substrates
0
100
200
300
400
500
600
0
100
200
300
400
500
600
0
50
100
150
200
250
300
Pull-off force (mN)
0
50
100
150
200
250
300
a a
b b b
a
cb,c
a,b a,b
b b
aaa
a
c
b b b
Traction
Pull-off
Fig. 5. Traction and pull-off force for the two species on the differently curved substrates. (A,B) Orestes mouhotii (N=14) traction force (A) and
pull-off force (B). (C,D) Sungaya inexpectata (N=14) traction force (C) and pull-off force (D). Groups with different lowercase letters are statistically different. Boxes
indicate the 25th and 75th percentiles, whiskers are the 10th and 90th percentiles and the line within the boxes shows the median. (E) The different substrates.
6
RESEARCH ARTICLE Journal of Experimental Biology (2020) 223, jeb226514. doi:10.1242/jeb.226514
Journal of Experimental Biology
65.08 mN on the 11.9 mm diameter tube and 93.20±37.48 mN on
the 5.0 mm diameter tube. The lowest traction force of 60.12±
31.74 mN was measured on the 2.9 mm tube. In general,
O. mouhotii generated higher traction on the substrates with the
largest diameters (i.e. lowest curvature) and lower traction on the
smallest diameter tube. Kruskal–Wallis one-way ANOVA
(H=41.82, d.f.=4, N=14, P=0.001) revealed a significantly higher
traction force on the flat substrate and the two largest tubes
(24.8 mm diameter and 11.9 mm diameter) in comparison to the
5.0 mm and 2.9 mm tubes (Tukey’s test, P<0.05).
Pull-off forces
The pull-off forces generated by S. inexpectata (Fig. 5D) were
highest on the 5.0 mm tube with 166.41±34.93 mN (mean±s.d.)
(one-way ANOVA, F=30.65, N=14, P<0.001; Holm–Šidák test,
P<0.001). The performance on this tube was significantly higher
than on the other tubes (2.9 mm: 99.73±37.57 mN, 11.9 mm:
119.61±26.39 mN, 24.8 mm: 98.08±33.02 mN), which were not
significantly different from each other (one-way ANOVA, F=30.65,
N=14, P<0.001; Holm–Šidák test, P>0.05). The pull-off forces on
the flat substrate were significantly lower than on all other substrates
(40.14±17.42 mN; one-way ANOVA, F=30.65, N=14, P<0.001;
Holm–Šidák test, P<0.001). For O. mouhotii, the pull-off forces
(Fig. 5B) were significantly higher on the three smallest tubes than
on the largest diameter tube and the flat substrate (2.9 mm: 111.68±
50.94 mN, 5 mm: 129.49±47.87 mN, 11.9 mm: 125.96±61.62 mN;
Kruskal–Wallis ANOVA on ranks, H=40.05, d.f.=4, P≤0.001,
N=14; Tukey’s test, P<0.05). The median of the pull-off forces was
highest on the 5.0 mm diameter tube, similar to S. inexpectata, but
the difference between the forces on 5.0 mm and the other small
substrate diameters was not significant (Tukey’s test, P>0.05). The
mean pull-off force on the 24.8 mm diameter tube (67.41±
31.30 mN) was higher than that on the flat substrate (35.27±
14.62 mN); however, this difference was also statistically not
significant (Kruskal–Wallis ANOVA on ranks, H=40.05, d.f.=4,
P≤0.001, N=14; Tukey’s test, P>0.05). In contrast to the traction
forces, both species revealed increasing pull-off forces on
decreasing tube diameter, with the maximum on the 5.0 mm tube
and decreasing pull-off forces for tubes thinner than 5.0 mm.
Comparison of attachment forces between species
The adult females of O. mouhotii (0.91±0.22 g, N=14) were lighter
than those of S. inexpectata (1.18±0.26 g, N=14; t-test, t=3.09,
d.f.=13, P=0.009). Therefore, for comparing traction and pull-off
forces of the two species with different tarsal lengths, safety factors
(attachment force/body weight) were used (Fig. 6A,B). The safety
factor in the traction direction was higher for S. inexpectata (9.17±
4.13) in comparison to O. mouhotii (5.89±3.22) on the 2.9 mm tube
(t-test, t=2.337, d.f.=26, N
1,2
=14, P=0.014). On all other substrates,
the differences between the specific safety factors were not
0
10
20
30
40
50
0
5
10
15
20
25
30
A
B
d=24.8 mm
d=11.9 mm
d=5.0 mm
d=2.9 mm
Flat
Sungaya inexpectata
Orestes mouhotii
Substrates
Traction
Pull-off
Safety factorSafety factor
*
*
d1d2d3d4d5
d1d2d3d4d5
d1
d2
d3
d4
d5
Fig. 6. Comparison of safety factors in the traction and pull-off direction between the two species on the differently curved substrates. (A) Traction
(N=14 for both). (B) Pull-off (N=14 for both). Boxes indicate the 25th and 75th percentiles, whiskers are the 10th and 90th percentiles and the line within
the boxes shows the median. *P<0.05, only significant comparisons are displayed. (C) The different substrates.
7
RESEARCH ARTICLE Journal of Experimental Biology (2020) 223, jeb226514. doi:10.1242/jeb.226514
Journal of Experimental Biology
significant. In contrast, comparison of safety factors in the pull-off
direction revealed a significantly higher safety factor on the 2.9 mm
tube for O. mouhotii (t-test, t=−2.149, d.f.=26, N
1,2
=14, P=0.021)
(Fig. 6B). Although the safety factors in the pull-off direction on the
11.9 mm diameter tube were different for the species (O. mouhotii:
14.27±5.89, S. inexpectata: 11.05±4.12), the t-test probability value
was slightly higher than 0.05 (t-test, t=−1.666, d.f.=26, N
1,2
=14,
P=0.054).
DISCUSSION
Tarsal morphology
In some stick insect species, the length of the tarsi differs between
the three leg pairs (Poinar, 2011; Vallotto et al., 2016, measured
from fig. 2 therein). We did not find a significant difference in the
total tarsal length between different leg pairs in the two species of
Heteropterygidae examined herein, but difference occurred between
the species. In the following, we discuss the functional relevance of
the different tarsal lengths in regard to the attachment performance
on curved substrates. Other insects use tibial spurs for locomotion
on curved substrates, either for interlocking with the stem (Gladun
and Gorb, 2007) or in combination with the pretarsal claws to
generate traction even on smooth curved substrates (Bußhardt et al.,
2014). Such structures are not present in the species examined
herein.
Attachment performance on curved substrates
In the vertical direction, the generated pull-off force of both species
increased with the degree of curvature, from the flat substrate to the
5.0 mm diameter tube. On the thinnest tube, the pull-off forces were
again reduced (see Fig. 4B,D). Accordingly, there is an optimal
range of stem diameter, where the stick insect generates the strongest
attachment, and despite the different tarsal lengths, this range was
surprisingly similar for the two species. Furthermore, in both
species, the generated pull-off force on the three smallest tubes, even
on the thinnest one (2.9 mm diameter), was significantly higher than
that on the flat plate.
On curved substrates, the increasing attachment performance can
be caused by generation of active adduction forces. When climbing
on thin rods, compared with their own body size, animals can utilize
clamping grips and generate active adduction forces to increase
normal load on the surface. This has been shown for tree frogs
(Endlein et al., 2017; Hill et al., 2018) and was additionally assumed
for sawyer beetles (Voigt et al., 2017) and other insect species from
different lineages (Gladun and Gorb, 2007). The angle between the
substrate surface and the direction of pulling obviously influences
the pull-off force. According to Kendall’s peeling model, the
pulling force is dependent on the peeling angle (Kendall, 1975).
Insect attachment systems are usually more complex than Kendall’s
classical peeling model predicts (Gorb and Heepe, 2017; Heepe
et al., 2017); still, there is a significant influence of the angle
between substrate and the detaching tarsus in many animals (Gu
et al., 2016). The relationship between peeling angle and attachment
force has been previously shown in Hymenoptera (Federle et al.,
2001), Diptera (Niederegger et al., 2002) and Blattodea (Clemente
and Federle, 2008), as well as in vertebrates, such as geckos
(Autumn et al., 2000; Persson and Gorb, 2003) and frogs (Hanna
and Barnes, 1991).
In the species examined herein, leg and tarsus orientation on a
curved tube caused a lower angle between the attachment pads and
the direction of pulling with increasing curvature of the substrate
(Fig. 4), resulting in a higher amount of force, which is necessary for
detachment of the attachment pad. This relationship is also reflected
by the AoA: a larger AoA of the animal on the substrate results in
more individual attachment pads in contact with the surface, with
the pulling force acting at a correspondingly lower angle.
Consequently, a higher AoA results in a higher pull-off force,
except for the smallest tube. The pull-off force on the smallest tube
(2.9 mm diameter) was significantly lower than on the 5.0 mm
diameter tube in S. inexpectata (in O. mouhotii, the median of the
pull-off force was lower as well, but not statistically significant).
Additionally, the walking gait pattern of both species revealed a
larger percentage of the tetrapod gait. The tetrapod gait in
comparison to the tripod gait often indicates a less secure
attachment to the substrate in insects (Gorb and Heepe, 2017;
Büscher and Gorb, 2019). On curved substrates with a very low
diameter, the flexion of the tarsal chain is not sufficient to bring all
attachment pads into contact with the substrate and hence results in a
lower pull-off force. However, S. inexpectata is able to contact the
2.9 mm diameter tube with the arolium and two euplantulae
(Fig. 4C).
In contrast to the pull-off forces, the traction forces were higher
for larger tube diameters and decreased with decreasing tube
diameter for both species (Fig. 5A,C). The traction forces were
highest on curved substrates with large diameters (24.8 mm). The
curvature of the substrate potentially enables the tarsi to better adapt
to the surface profile than on flat substrates. Additionally, the
convex substrate curvature provides better grasping of the substrate
and the generation of higher load on the tarsi in comparison to flat
substrates (Frantsevich and Cruse, 1997; Gladun and Gorb, 2007;
Voigt et al., 2017). On very thin stems, with similar or lower
diameter than the tarsal length, the examined species provide lower
traction force than on surfaces with a lower degree of curvature.
Generally, on the smaller tubes, especially on the thinnest ones,
fewer attachment pads were brought into contact and, hence, less
actual contact area was provided, resulting in lower attachment force
(Arzt et al., 2003; Persson and Gorb, 2003). Presumably, on smaller
diameter tubes, less contact area is available to distribute the
resulting forces, especially in the lateral dimension. This probably
results in a reduced stabilization on the thinner tubes in comparison
with the larger ones, leading to a faster detachment of single legs
and a lower attachment force in general. Additionally, in animals
that clamp their tarsi around the tubes, the traction force vector along
the substrate is oriented perpendicular to the orientation of the
tarsus. Accordingly, the tarsi are sliding in the lateral direction in
most cases. If the tarsi were arranged differently, the animals
actively positioned them along the tube and tried to counteract the
pulling direction. Previous studies have reported higher friction for
stick insect euplantuae while being pulled compared with being
pushed (Bußhardt et al., 2012). In the lateral direction of the
euplantula, to the best of our knowledge, nothing is known about the
frictional properties of the attachment pad in stick insects. The
generated traction force, however, is significantly lower on the
smaller tubes, where the tarsi cannot be arranged along the tube
length, in comparison to the flat substrate and the tubes with larger
diameters (Fig. 5A,C).
Influence of tarsal length on attachment on curved
substrates
In both directions (vertical and horizontal pulling), the two species
showed similar trends, i.e. higher pull-off force on thinner stems and
higher traction on thicker stems and flat substrates. The safety
factors of both species were similar and not statistically different on
all substrates except the 2.9 mm diameter tube. Hence, the length of
the tarsus essentially does not matter for stems with larger diameters
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Journal of Experimental Biology
than the tarsal length. Only on the thinnest tube (2.9 mm diameter)
was the safety factor of S. inexpectata in the traction direction higher
than for O. mouhotii and lower in the pull-off direction (Fig. 6). The
shorter tarsi of O. mouhotii can generate a larger AoA in comparison
to the longer tarsi of S. inexpectata. This results in a lower angle
between the tarsus and tibia and, hence, could explain the higher
safety factor of O. mouhotii in the pull-off direction (e.g. Gu et al.,
2016). In spite of the longer tarsus of S. inexpectata, it does not
allow for a proper grasping around the rod, as the tarsus is not
flexible enough to follow the perimeter of the substrate. The tarsus
of O. mouhotii, however, is able to bring all attachment pads into
contact.
Opiliones (Arachnida), or harvestmen, are able to provide friction
using their elongated and highly articulated prehensile feet (Wolff
et al., 2019). In contrast to harvestmen, stick insects, like most other
insects, are not able to hyperflex the tarsus and therefore need to
provide load by the coordinated action of contralateral legs (Gladun
and Gorb, 2007; Bußhardt et al., 2014; Voigt et al., 2017). The
safety factor in the traction direction was higher for S. inexpectata
on the substrate with the smallest diameter. The tarsi of this species
made contact with the substrate at a lower AoA than those of O.
mouhotii (Fig. 4), i.e. >90 deg but <180 deg. This results in
potentially better clamping and higher adduction forces of the
contralateral legs, and consequently, higher traction, as the traction
is dependent on the load on the attachment pads (e.g. Gorb and
Scherge, 2000; Gorb et al., 2000; Bußhardt et al., 2012; Gorb, 2007;
Labonte and Federle, 2013; Labonte et al., 2014). In O. mouhotii,in
contrast, the high AoA (>180 deg) on the small tube results in lower
adduction forces, as the tarsus grasps around the tube. The force
transmitted to the attachment pads is, in this conformation, not
generating load, as the tarsus is under tension. This is in agreement
with observations previously made on other insects, which clasp
stems with an equal or slightly larger diameter than the tarsal length,
but clutch stems with a smaller diameter with the contralateral tarsi
(Gladun and Gorb, 2007), and, as shown for the two species
examined herein, this clutching results in higher traction.
Usage of the attachment system for locomotion on curved
substrates
According to Federle and Endlein (2004), attachment is regulated
on four different hierarchical levels, ranging from control of the
attachment system itself, over movements of the tarsomeres/tarsi and
the legs, to the body kinematics. Some aspects of the tarsus, leg and
body positioning have already been discussed above. Furthermore,
there are some behavioural aspects influencing body kinematics in
addition to the walking gait patterns and posture (Table 1). Generally,
the clamping efficiency is dependent on morphological
characteristics; for example, the ratios of body width and leg length
(Voigt et al., 2017). The same ratios are reported to show a correlation
of behaviour and morphology, i.e. while climbing on stairs, stick
insect species with shorter legs show different behaviour from species
with long legs (Theunissen et al., 2015).
With respect to their body lengths, the legs of S. inexpectata are
longer than those of O. mouhotii. Both are associated with low-
growing vegetation and show quite similar feeding habits and bury
their eggs in the soil (Büscher et al., 2019). However, there are no
studies that have investigated details of their natural behaviour in the
field. Still, the tarsi of O. mouhotii are shorter and therefore perform
better on thin branches than those of S. inexpectata. Presumably, the
tarsi of O. mouhotii are adapted to locomote on low-growing
vegetation with small stem diameter, rather than larger plant
diameters (bushes and trees).
In lizards (Goodmann et al., 2008) and Stenus rove beetles (Betz,
2006) limb length and habitat correlate in some species, e.g. limb
length increases with perch diameter or habitat openness ( presence
of less suitable refugia in the habitat). Likewise, openness in the
natural habitats could be higher for S. inexpectata than for O.
mouhotii and therefore longer legs (and tarsi) could be a result of the
presence of larger substrate diameters and less-suitable twigs for
imitation in the natural habitats of S. inexpectata.
Ground-dwelling spiders with longer legs walk faster on curved
substrates with large diameter compared with flat substrates and thin
stems (Prenter et al., 2010). Similarly, S. inexpectata has longer
limbs and does not show any cataleptic defensive reaction and
instead tries to escape. Orestes mouhotii in contrast relies on its
camouflage in combination with thanatosis (Bedford, 1978). For the
employment of visual camouflage, next to the animal’s size,
structure and colour of the habitat play a major role, as shown for
chameleons (Cuadrado et al., 2001) and moths (Kang et al., 2015).
Orestes mouhotii combines smaller size and stronger crypsis
compared with S. inexpectata, which might be a part of the predator
avoidance strategy of this species. Traction forces, however, were
larger for longer tarsi on some substrates. Higher speed during
locomotion and active escape reactions might be assumed as
potential behavioural traits associated with the longer tarsal length.
Conclusions
This study sheds light on the attachment performance of stick
insects on curved substrates and provides, to the best of our
knowledge, the first quantitative data on traction and pull-off forces
for this group of insects on smooth rods of different diameter. Stick
and leaf insects face several different surface topographies in their
natural habitats, and dominant among these are twigs and branches
of different diameter. The degree of substrate curvature has a
significant influence on the attachment ability of both examined
species, namely higher traction on curved substrates with larger
diameters and higher pull-off force on substrates with lower
diameter. On flat substrates and curved substrates with large
diameters, the two species showed similar traction performance and
much lower pull-off forces than on curved substrates. The traction
force decreases with decreasing stem diameter and the pull-off force
increases with decreasing stem diameter, unless the substrate is
thinner than the tarsal length. On flat substrates and curved
substrates with large diameters, the tarsus length does not
significantly influence the attachment ability. On stems with
diameters lower than the length of the tarsus, the pull-off forces
are reduced, as the contact formation between tarsal attachment pads
and the substrate is reduced. Traction forces, however, are larger for
longer tarsi on stems with a diameter falling below the length of the
tarsus, as the longer tarsi can better clamp onto the substrate between
the contralateral tarsi and produce higher adduction forces, while
shorter tarsi clasp around the stem. The attachment performance is
also dependent on the number of tarsi and tarsomeres in contact,
tarsus position on the tube (e.g. AoA), tarsus and leg orientation, as
well as leg and body configuration on the surface.
Corroborating earlier studies on other insects (e.g. Federle et al.,
2001; Niederegger et al., 2002; Clemente and Federle, 2008), the
pull-off force is dependent on the tibiotarsal angle. On rods, the
pull-off force is higher for higher AoA (or lower tibiotarsal angle).
Furthermore, the AoA probably influences adduction forces,
leading to proper load transmission on the euplantulae, which in
turn influences the traction force. Additionally, the contribution of
claws (Song et al., 2016; Büscher and Gorb, 2019) as well as
differences in the euplantular attachment microstructures present in
9
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Journal of Experimental Biology
other stick insect species (Büscher et al., 2018a,b, 2019) might also
influence the attachment performance on curved rough substrates.
Surfaces with different degrees of curvature play an important
role for stick insects. Referring to the results of this study, this
induces a much more complex interaction of various phenomena.
The results presented herein further stress the importance of
evaluation of attachment performance on different surface
geometries (Gladun and Gorb 2007; Bußhardt et al., 2014; Voigt
et al., 2017, 2019), besides commonly investigated flat substrates.
Furthermore, insights generated from the present experiments on
curved substrates can be potentially useful for the design of bio-
inspired robotic grippers (e.g. Gorb et al., 2007; Voigt et al., 2012).
Acknowledgements
We thank Chuchu Li and Dennis Petersen (Department of Functional Morphology
and Biomechanics, Kiel University, Germany) for fruitful discussions and Femke
Vogel (same affiliation) for assistance with force measurements. Furthermore, we
thank Holger Dra
̈ger (Schwerin, Germany) and Daniel Dittmar (Berlin, Germany) for
providing the breeding stocks used for the laboratory cultures.
Competing interests
The authors declare no competing or financial interests.
Author contributions
Conceptualization: T.H.B., S.N.G.; Methodology: T.H.B., S.N.G.; Validation: T.H.B.,
S.N.G.; Formal analysis: T.H.B., M.B.; Investigation: T.H.B., M.B.; Resources:
T.H.B., S.N.G.; Writing - original draft:T.H.B.; Writing - review & editing: T.H.B., M.B.,
S.N.G.; Visualization: T.H.B., M.B.; Supervision: T.H.B., S.N.G.; Project
administration: T.H.B., S.N.G.; Funding acquisition: S.N.G.
Funding
Funding was provided by the Deutsche Forschungsgemeinschaft (grant GO 995/34-1).
Supplementary information
Supplementary information available online at
https://jeb.biologists.org/lookup/doi/10.1242/jeb.226514.supplemental
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RESEARCH ARTICLE Journal of Experimental Biology (2020) 223, jeb226514. doi:10.1242/jeb.226514
Journal of Experimental Biology
Table S1. Measurements of all tarsomeres and the total length of the hind tarsus for Orestes
mouhotii and Sungaya inexpectata (N1,2 = 11).
species
ID
T1 [mm]
T2 [mm]
T3 [mm]
T4 [mm]
T5 [mm]
Lenght
[mm]
Orestes mouhotii
#OM1
1.00
0.47
0.45
0.50
1.06
3.48
Orestes mouhotii
#OM2
0.90
0.42
0.41
0.45
0.95
3.13
Orestes mouhotii
#OM3
1.09
0.51
0.50
0.55
1.16
3.81
Orestes mouhotii
#OM4
0.88
0.41
0.40
0.44
0.93
3.06
Orestes mouhotii
#OM5
1.02
0.48
0.46
0.51
1.08
3.54
Orestes mouhotii
#OM6
0.80
0.37
0.36
0.40
0.85
2.79
Orestes mouhotii
#OM7
1.05
0.49
0.48
0.52
1.11
3.65
Orestes mouhotii
#OM8
1.09
0.51
0.50
0.55
1.16
3.80
Orestes mouhotii
#OM9
1.00
0.47
0.45
0.50
1.06
3.47
Orestes mouhotii
#OM10
0.82
0.38
0.37
0.41
0.87
2.85
Orestes mouhotii
#OM11
0.92
0.43
0.42
0.46
0.98
3.21
Sungaya inexpectata
#SI1
2.91
0.96
0.88
0.92
2.20
7.87
Sungaya inexpectata
#SI2
2.88
0.95
0.88
0.91
2.18
7.80
Sungaya inexpectata
#SI3
2.72
0.90
0.83
0.85
2.05
7.35
Sungaya inexpectata
#SI4
2.98
0.99
0.91
0.94
2.26
8.07
Sungaya inexpectata
#SI5
2.76
0.91
0.84
0.87
2.09
7.46
Sungaya inexpectata
#SI6
2.93
0.97
0.89
0.92
2.22
7.94
Sungaya inexpectata
#SI7
2.90
0.96
0.88
0.91
2.20
7.85
Sungaya inexpectata
#SI8
3.09
1.02
0.94
0.97
2.34
8.35
Sungaya inexpectata
#SI9
2.80
0.93
0.85
0.88
2.12
7.58
Sungaya inexpectata
#SI10
3.16
1.04
0.96
0.99
2.39
8.54
Sungaya inexpectata
#SI11
3.05
1.01
0.93
0.96
2.31
8.26
Journal of Experimental Biology: doi:10.1242/jeb.226514: Supplementary information
Journal of Experimental Biology • Supplementary information
Table S2. Measurements of the total length of the tarsi of all three leg pairs for Orestes
mouhotii and Sungaya inexpectata (N1,2 = 15).
Total length [mm]
species
ID
foreleg
midleg
hindleg
Sungaya inexpectata
#SI12
6.6
6.3
7.2
Sungaya inexpectata
#SI13
6.8
6.6
6.7
Sungaya inexpectata
#SI14
6.6
6.8
6.9
Sungaya inexpectata
#SI15
7.0
6.8
7.0
Sungaya inexpectata
#SI16
6.2
6.2
6.2
Sungaya inexpectata
#SI17
6.8
6.7
7.0
Sungaya inexpectata
#SI18
6.8
6.3
6.5
Sungaya inexpectata
#SI19
6.9
6.5
6.8
Sungaya inexpectata
#SI20
6.2
6.7
6.8
Sungaya inexpectata
#SI21
6.3
6.7
6.7
Sungaya inexpectata
#SI22
7.0
6.5
6.9
Sungaya inexpectata
#SI23
7.1
6.8
7.1
Sungaya inexpectata
#SI24
6.8
6.9
6.3
Sungaya inexpectata
#SI25
6.6
6.8
6.4
Sungaya inexpectata
#SI26
6.5
6.7
6.8
Orestes mouhotii
#OM12
3.1
2.8
2.9
Orestes mouhotii
#OM13
3.3
3.7
3.1
Orestes mouhotii
#OM14
3.2
3.8
3.8
Orestes mouhotii
#OM15
3.5
3.5
3.5
Orestes mouhotii
#OM16
2.9
2.9
3.5
Orestes mouhotii
#OM17
3.0
3.2
3.2
Orestes mouhotii
#OM18
3.1
3.5
2.8
Orestes mouhotii
#OM19
3.5
3.7
3.6
Orestes mouhotii
#OM20
2.9
2.9
3.5
Orestes mouhotii
#OM21
3.8
3.0
3.4
Orestes mouhotii
#OM22
3.3
3.0
3.1
Orestes mouhotii
#OM23
3.4
3.4
2.9
Orestes mouhotii
#OM24
3.7
3.2
3.2
Orestes mouhotii
#OM25
3.5
3.3
3.6
Orestes mouhotii
#OM26
3.3
3.4
3.5
Journal of Experimental Biology: doi:10.1242/jeb.226514: Supplementary information
Journal of Experimental Biology • Supplementary information
