Asymmetric Limb Control in Steady Bipedal Locomotion Robert E

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RESEARCH ARTICLE Guineafowl with a twist: asymmetric limb control in steady bipedal locomotion Robert E. Kambic1,2,*, Thomas J. Roberts1 and Stephen M. Gatesy1

ABSTRACT imbalance. These sources include deviations from bilateral In avian bipeds performing steady locomotion, right and left limbs are symmetry in the skeleton and soft tissues (e.g. Van Valen, 1962), typically assumed to act out of phase, but with little kinematic shifting of the center of mass due to uneven visceral loads or from – disparity. However, outwardly appearing steadiness may harbor changes in head and tail position, left right differences in step previously unrecognized asymmetries. Here, we present marker- length or duty factor when one leg differentially accelerates or based XROMM data showing that guineafowl on a treadmill routinely decelerates the body, and irregular cycles of yaw, pitch and roll yaw away from their direction of travel using asymmetrical limb among strides (Gatesy, 1999a; Rubenson et al., 2007; Jindrich et al., kinematics. Variation is most strongly reflected at the hip joints, where 2007; Abourachid et al., 2011; Stoessel and Fischer, 2012). patterns of femoral long-axis rotation closely correlate to degree of However, these factors are not usually considered significant yaw divergence. As yaw deviations increase, hip long-axis rotation enough to warrant special attention or to override the general angles undergo larger excursions and shift from biphasic to assumption of symmetry. For instance, many studies measure the monophasic patterns. At large yaw angles, the alternately striding motion, muscle activity or mechanics of a single limb under the limbs exhibit synchronous external and internal femoral rotations of premise that these data are representative of both limbs. Asymmetry substantial magnitude. Hip coordination patterns resembling those in these cases will be a component of the calculated standard used during sidestep maneuvers allow birds to asymmetrically deviations, undifferentiated from measurement error, stride-to-stride modulate their mediolateral limb trajectories and thereby advance variation in speed, and individual variation. using a range of body orientations. While collecting data for a 3D kinematic study of avian walking and running, we were surprised to find major deviations in the KEY WORDS: Locomotion, Bipedalism, Kinematics, Avian, XROMM, orientation of the pelvis relative to the direction of movement. Our Three-dimensional, Numida meleagris, X-ray, Animation preliminary results revealed that birds could hold their position on a treadmill while maintaining a net yaw. The implications of deviating INTRODUCTION the body’s longitudinal axis away from the direction of travel led us Analyses of avian bipedalism typically focus on steady locomotion. to a new set of questions about symmetry and differential limb Birds are predominantly studied moving forward over level ground at control. How asymmetrical is avian bipedalism during steady relatively constant speeds in both kinematic (Cracraft, 1971; Jacobson locomotion? When symmetry is broken, how are the many degrees and Hollyday, 1982; Muir et al., 1996; Gatesy and Biewener, 1991; of freedom (DoF) within joints, among joints and among limbs Gatesy, 1999a; Abourachid, 2000, 2001; Reilly, 2000; Verstappen coordinated to move the two legs differently? et al., 2000; Rubenson et al., 2007; Nyakatura et al., 2012; Provini Until recently, such questions were difficult to answer. With the et al., 2012; Stoessel and Fischer, 2012) and kinetic (Clark and development of XROMM (X-ray reconstruction of moving Alexander, 1975; Alexander et al., 1979; Roberts et al., 1998; morphology; Brainerd et al., 2010; Gatesy et al., 2010) we now Hancock et al., 2007; Goetz et al., 2008; Nudds et al., 2010; Rubenson have the ability to measure six degree of freedom skeletal et al., 2011; Andrada et al., 2013, 2014) analyses. Compared with the kinematics at high resolution. Our 3D analysis of maneuvering uniformity of locomotion on a treadmill or straight trackway, the locomotion in helmeted guineafowl (Numida meleagris) revealed inherent variability of unsteady behaviors is much more difficult to the critical role of femoral and tibiotarsal long-axis rotation (LAR; characterize. Consequently, examinations of birds accelerating Kambic et al., 2014). Hip and knee LAR were critical for achieving (Roberts and Scales, 2002, 2004), maneuvering/turning (Jindrich the non-planar poses required to transversely shift and reorient the et al., 2007; Kambic et al., 2014), and running over uneven terrain body. We hypothesized that a closer look inside steady, yet (Daley and Biewener, 2006; Daley et al., 2009; Birn-Jeffery et al., asymmetrical, forward locomotion might reveal comparable non- 2014; Gordon et al., 2015) remain relatively uncommon. sagittal foot motions governed by similar joint control strategies. Steady locomotion is typically assumed to be symmetrical in striding bipeds. Although out of phase, right and left limbs are MATERIALS AND METHODS thought to mirror each other to a large degree. Most researchers All procedures conducted with animals were approved by the Institutional would likely concur that absolutely symmetrical operation of the Animal Care and Use Committee at Brown University. Six helmeted limbs is rare, as there are many potential sources of functional guineafowl, Numida meleagris (Linnaeus 1758), individuals (1.4±0.19 kg) were acquired from a local source and housed as a group with food and water available ad libitum. 1Department of Ecology and Evolutionary Biology, Brown University, Providence, 2 The methods used for analyzing X-ray videos, animating bone models RI 02912, USA. Museum of Comparative Zoology and Department of Organismic and applying joint coordinate systems in this paper are identical to those and Evolutionary Biology, Harvard University, Cambridge, MA 02138, USA. described in Kambic et al. (2014). Briefly, birds were surgically implanted *Author for correspondence ([email protected]) with multiple conical carbide markers in the pelvis, femur, tibiotarsus and tarsometatarsus. The first two birds were implanted only in the right limb,

Received 31 May 2015; Accepted 4 October 2015 the third in the right limb and left femur. The final three individuals were Journal of Experimental Biology

3836 RESEARCH ARTICLE Journal of Experimental Biology (2015) 218, 3836-3844 doi:10.1242/jeb.126193 bilaterally implanted with markers in both limbs. After sufficient recovery derived from a hospital scanner (Philips Medical System, Best, The time, individuals were recorded in the W. M. Keck Foundation XROMM Netherlands) at 512×512 pixel resolution and 0.625 mm intervals, with one facility located at Brown University. This system uses two EMD exception in which a Fidex micro-CT scanner (Animage, LLC, Pleasanton, Technologies 425 model EPS 45-80 X-ray generators connected to Varian CA, USA) at 512×512 pixel resolution and 0.456 mm intervals was used. model G-1086 X-ray tubes suspended on ceiling-mounted telescoping Geomagic Studio 2013 (3D Systems, Morrisville, NC, USA) was used to cranes. The X-rays are captured by Dunlee model TH9447QXH590 image clean bone models. XrayProject, a set of XROMM tools for Matlab intensifiers (40.64 cm diameter) mounted on mobile-arm bases. The image (MathWorks, Natick, MA, USA), was used for undistorting X-ray videos, intensifiers are backed with Phantom v10 high-speed digital video cameras calibrating cameras, and 3D marker tracking. These tools were also used to (Vision Research, Wayne, NJ, USA). We recorded at 1760×1760 pixel generate transformation matrices to position and orient the bone models in resolution. Additional Phantom v9.1 cameras were used to capture light 3D space (Fig. 1). Bone models were animated in Maya 2010 (Autodesk video at 1600×1200 pixel resolution. All videos were recorded at Inc., San Rafael, CA, USA). The raw data, including X-ray and light videos, 250 frames s−1 and 1/2000 s shutter speed. and calibration and distortion grid images, were uploaded to the X-ray Three individuals were recorded within the acrylic enclosure (29.5 cm Motion Analysis Research Portal (xmaportal.org) and will be made publicly wide×100 cm long×48 cm high) of a custom-built hand-crank treadmill at available on publication. As a measure of marker tracking precision, the speeds up to 1 m s−1. Bilaterally marked individuals were recorded on a standard deviation of intermarker distances within single bones (Tashman DC5 model Jog-a-Dog motorized treadmill (JOG A DOG, LLC, Ottawa and Anderst, 2003; Brainerd et al., 2010) averaged ±0.222 mm. Lake, MI, USA) within a similar enclosure (35 cm wide×106 cm Joint angles were calculated according to joint coordinate systems (Grood long×48 cm high). The treadmill was oriented between the horizontally and Suntay, 1983; Wu et al., 2002) that were set up identically to those arranged X-ray systems, providing two 45 deg oblique views (Fig. 1A,B). described in the appendices of Kambic et al. (2014). The sign conventions Standard light video cameras recorded anterior and lateral perspectives were as follows. Pelvic yaw was positive to the left, pelvic pitch was positive (Fig. 1C,D). Individuals were run for short durations (∼15–30 s) and had when raising the cranial end, and pelvic roll was positive when rolling the several minutes of rest in between trials while the files were evaluated and right hip higher than the left. At the hip, extension was positive, abduction saved. Trials were deemed suitable for keeping if the sequence included a was positive and external rotation was positive. At the knee, extension was number of consecutive strides where all markers were visible in both X-ray positive, adduction was positive (note that this differs from the hip and videos. If individuals appeared to be tiring, data collection would be ankle), and external rotation was positive. Finally, at the ankle, extension postponed until the next day. Birds were generally run without interference. was positive, abduction was positive and external rotation was positive. To prevent slowing birds from dropping out of the X-ray volume, an Stance and swing phases were initially determined using light videos, operator standing behind the treadmill would lightly contact the tail with a with stance being defined as the first frame showing ground contact by digit nitrile glove hanging loosely from a dowel. The exception to this protocol III, and swing beginning with the first frame where the foot was not in was for a single trial where a wedge of foam was placed at the front of the contact with the ground. These sequences were used to guide the creation of running box to induce a larger yaw. The foam formed a 45 deg angle with rules to infer stance and swing phases from knee joint rotations for the the treadmill, and the treadmill speed was lowered such that the individual sequences without light video. In the test sequences used to create these could maintain a large yaw while progressing forwards. When data rules, the knee method results were typically within 1–2 frames of the light collection for an individual was completed, the individual was induced with video results. We use the term ‘excursion’ to refer to the overall range 5% isoflurane and killed with Beuthanasia. (maximum minus minimum) of angles during a stride or phase. A ‘net Bilaterally marked individuals were also recorded running over a excursion’ is the difference in angle over a phase (end minus start). trackway (34 cm wide×310 cm long) positioned the same way as the To characterize mediolateral displacements of the foot relative to the treadmills between the X-ray systems. The sides of the trackway were pelvis, a transverse distance (Kambic et al., 2014) was calculated that constructed from opaque siding, with a 70 cm section of acrylic in the measured the distance from the distal end of the tarsometatarsus to middle of the trackway to allow light video recording. This transparent the median sagittal plane. For both limbs, transverse distance was zero at the section coincided with the X-ray field of view. midline, negative medial to the median plane, and positive lateral to For treadmill data, trials to be analyzed were chosen for their length and the plane. consistency in keeping markers in view. The longest trials in which the individual remained within the X-ray volume were preferred. Light video RESULTS was not used to identify trials for marker tracking, and we made no attempt Steady locomotion and pelvic kinematics ‘ ’ to infer pelvic yaw before a trial was analyzed. Steady locomotion was None of the six individuals studied showed any signs of lameness, inferred based on the limitations of the X-ray volume coupled with the limping or obvious external gait asymmetry following surgery. The consistency of the treadmill speed. Trackway sequences were chosen for – analysis without regard for speed or pelvic yaw. subjects moved steadily over a range of treadmill speeds (0.6 −1 Data were analyzed using the marker-based XROMM workflow 1.9 m s ). Duty factor approached 0.5 at the highest speed, but (Brainerd et al., 2010; xromm.org). Bone models were segmented using aerial phases were seldom observed in these walking and ‘grounded OsiriX (v.4.1.2, Geneva, Switzerland; Rosset et al., 2006) from CT scans running’ gaits (Rubenson et al., 2004; Hancock et al., 2007; Nudds

A BCD

Fig. 1. Reconstructing bone positions by marker-based XROMM. Four synchronized video frames of a bird yawed 16 deg to the right. (A,B) Bone models registered to X-ray video frames based on rigid body kinematics of tracked marker clusters. (C,D). Bone models rendered with respect to calibrated light cameras.

Data from this sequence are plotted in Fig. 3. Journal of Experimental Biology

3837 RESEARCH ARTICLE Journal of Experimental Biology (2015) 218, 3836-3844 doi:10.1242/jeb.126193 et al., 2011). We were only able to analyze sequences in which the values. In contrast, there were persistent deviations in yaw that subjects successfully maintained their position within the limited exceeded intra-stride variation. The bird began the sequence closely biplanar X-ray volume (ca. 9800 cm3, equivalent to a cube with parallel to the tread, but yawed slightly to the left (Fig. 3, stride 1). 21.4 cm sides). Additionally, recording time was limited to a Soon after, a yaw to the right (negative) accrued until the bird’s maximum of 10 s per trial. Despite these challenges, full sequences body was facing more than 20 deg away from its direction of travel containing as many as 23 consecutive strides were recorded. (Fig. 3, strides 2–3). The individual then maintained a right yaw Measurements of pelvic yaw confirmed the initial observation that over several strides (Fig. 3, strides 4–7). birds routinely deviate substantially from their direction of travel. The direction and magnitude of this deviation can be difficult to detect Limb kinematics from a lateral view (Fig. 2A–C). Data from six birds for 10 treadmill The hip, knee and ankle (intertarsal) joints underwent substantial sequences composed of over 12,000 frames from 197 steps (116 right, flexion/extension (FE). The hips flexed briefly at the beginning of 81 left) were widely distributed (Fig. 2D). For unknown reasons, all stance and then extended through most of the rest of the phase. individuals had a tendency to yaw to the right (see Discussion). Yaws Flexion began just prior to toe off and then the hips re-extended just of 3–5 deg to the right were most common, but values in the teens and before toe down. The knees flexed considerably throughout the twenties were regularly present across the trials analyzed. The broad majority of stance, with a variable degree of late-stance extension. distribution was not driven solely by the slowest walking sequences Flexion continued past toe off until the knees reversed direction and (Fig. S1). The bird in the fastest trial we recorded (1.9 m s−1) reached extended for the rest of swing. The ankles underwent flex–extend– a yaw of greater than 11 deg to the right. Although both overall flex sequences during stance, followed by large flex–extend magnitude and right skewing of yaws may have been an artefact of our excursions in swing. Abduction/adduction (ABAD) angle two different treadmills, birds moving freely down a trackway also changed comparatively little at any of the joints, similar to the exhibited yaw variation (Fig. 2D). minimal ABAD excursions during maneuvering (Kambic et al., Within an individual sequence, three-rotational DoF kinematic 2014). data from the pelvis and both limbs of a bird moving at 0.86 m s−1 LAR decreased progressively from hips to knees to ankles. At revealed consistent patterns over seven and a half strides (Fig. 3; the hips, LAR excursions showed high variability (N=15, 9.8± Movie 1). Variability in pelvic kinematics occurred at two major 8.5 deg, mean±s.d.). Strides earlier in the sequence (Fig. 3, strides scales. At the level of the stride, yaw, pitch and roll all showed 1–2) had less femoral rotation than later strides, in which hip LAR repeating patterns. During a given stance phase, the pelvis pitched excursions equaled or even exceeded hip FE excursions for several down for the first quarter, pitched up for the middle two quarters, steps (Fig. 3, strides 3–7). At the knees, LAR excursions were and then pitched down for the last quarter (best seen in Fig. 3, strides more consistent (N=14, 8.6±4.1 deg, mean±s.d.), but differed 4, 5). The body initially rolled towards the stance limb. Then, about slightly between the right and left sides. The right knee internally halfway through stance, the body began rolling away from the rotated through most of stance, whereas the left knee first stance limb (best seen in Fig. 3, strides 2, 4). Finally, the pelvis externally and then internally rotated. Both knees underwent net initially yawed away from the stance limb and then yawed toward it external rotation in swing. Some LAR occurred at the ankles, but for the remainder of the phase (best seen in Fig. 3, strides 3, 7). excursions during both stance and swing were small compared with Over the course of the 4 s sequence, intra-stride patterns were those for the knees and hips and were less variable (N=8, mean 3.9± repeated, with pitch and roll oscillating about relatively consistent 3.8 deg, mean±s.d.).

A BCFig. 2. Yaw during steady locomotion. (A–C) Light video frames from a treadmill sequence while the individual was yawed 7.1 deg to the left, 1.8 deg to the right and 14.1 deg to the right, respectively. (D) Histogram of pelvic yaw values for 10 steady treadmill sequences. Superimposed renderings show the orientation of the pelvis in dorsal view at the maximum yaw to the left and right, with the pelvic long axis (black) and the direction of treadmill travel (red). Arrows show ranges of yaw values for 12 short D sequences from the same birds moving freely down a trackway. Arrowheads indicate the animal’s direction of movement; those pointing to the right were facing in the 800 same direction as the treadmill trials. 11 deg 33 deg 700

600 B 500

Frame 400 A 300 C 200 100 0 21 18 15 12 9 6 3 0 3 6 9 12 15 18 21 24 27 30

Pelvic yaw (deg) Journal of Experimental Biology

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123 4 567 Fig. 3. Pelvis and joint angles over time for a fast 20 walking sequence. Pelvic angles are shown during right stance (solid), left stance (short dash) and double- 10 support (long dash) phases of the stride. Joint angles are shown for right (solid) and left (dashed) limbs 0 during stance (thick lines) and swing (thin lines). Gray –10 vertical lines separate strides, which are numbered at the top of the plot, based on the swing/stance Pelvis angle (deg) transitions of the right limb. Flexion/extension (FE) and yaw are blue, abduction/adduction (ABAD) and pitch are green, and internal/external rotation (LAR) and roll are red. 60 40 20

Hip angle (deg) 0

120

40 Knee angle (deg)

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Ankle angle (deg) 0

0 0.51 1.52 2.53 3.5 Time (s)

Variation in hip LAR monophasic U shapes (Fig. 4A, strides 3–7). LAR traces from the We compared limb kinematics from low-yaw and high-yaw right and left hips were synchronized in time because the hips sequences to determine whether persistent pelvic yaws of large rotated in opposite directions at equivalent points in their stride absolute value were associated with specific joint rotations. Only cycles. A 0.92 m s−1 sequence in which the bird maintained a right hip LAR underwent large-scale changes in pattern and magnitude yaw (Fig. 4B) shows the monophasic hip LAR pattern very clearly. that echoed those of yaw. Here, the right and left hips were even more synchronized in time, The relationship between hip LAR and pelvic yaw is more easily with traces that were frequently superimposed. observed if the two are plotted together in isolation (Fig. 4). Data Even in a faster (1.59 m s−1), relatively low-yaw trial (Fig. 4C), from the sequence plotted in Fig. 3 (Fig. 4A) show that the hips used body orientation and hip LAR were correlated. Throughout the final a biphasic LAR pattern initially, when the bird’s pelvis and body two-thirds of the sequence, when the bird maintained a low yaw, the deviated only a few degrees to the left from its direction of travel. hips exhibited a biphasic LAR pattern with little net LAR in either During stance, hip LAR curves were W-shaped as both limbs stance or swing (Fig. 4C, strides 5–9). The first third of the sequence underwent small internal–external–internal–external excursions showed LAR asymmetry at yaws of only ca. 10 deg (Fig. 4C, strides (Fig. 4A, strides 1–2). In swing, an external–internal sequence 1–4). Despite being biphasic, the hips underwent significant net gave each hip LAR curve an upside-down U shape (Fig. 4A, strides LAR excursions during stance and swing. Patterns of low-yaw, 1–2), but there was little or no net excursion during either phase. biphasic LAR and high-yaw, monophasic LAR were consistent Later in the sequence (Fig. 4A, strides 3–7), when the bird yawed across the hips of all individuals. well to the right, the biphasic hip LAR patterns shifted to more The relationship between hip LAR and yaw can be summarized monophasic waves. During this portion of the sequence, the hips by plotting net LAR excursion against average yaw during each rotated through large net excursions, but in opposite directions. The stance phase for all treadmill sequences (Fig. 5). Larger net stance result of these changes is that stance and swing together created hip LAR excursions were correlated with greater deviation from the Journal of Experimental Biology

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Fig. 4. Pelvic yaw and hip long-axis rotation for A 12 3 4 567 sample strides from three sequences. Line 0 weights and dashes follow Fig. 3. Gray vertical lines separate strides, which are numbered at the –10 top of the plots, based on the swing/stance transitions of the right limbs. (A) Data from the − –20 sequence plotted in Fig. 2 (0.86 m s 1). (B) Data −1

Pelvic yaw (deg) from a more yawed sequence (0.92 m s ). Gray 25 box highlights a single stance phase of the right leg. (C) Data from a higher speed sequence 20 − (1.59 m s 1). 15

5 Hip LAR (deg) LAR Hip 0

B 123456 7 8 0

(deg) –10 Pelvic yaw

10 Hip LAR (deg) LAR Hip 5

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(deg) –10 Pelvic yaw

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5 Hip LAR (deg) LAR Hip 0 0 1 2 3 Time (s) direction of travel. For example, during stride 2 of Fig. 4B, the right Previously, we found that LAR at these joints was crucial for hip underwent 19 deg of net internal rotation while at an average sidesteps, yaw maneuvers and slow turns (Kambic et al., 2014). The right yaw of 12 deg (Fig. 5, red circle). Right and left hips responded results presented here demonstrate that the role of LAR is not limited to yaw similarly in net LAR excursion magnitude, but in opposite to maneuvering; LAR is an essential component of avian steady directions. Yaws to the right co-occurred with net internal rotation at locomotion as well. the right hip during stance, but net external rotation at the left hip during stance, and vice versa. Variation in foot trajectory Maintaining a non-zero pelvic yaw on a treadmill has important DISCUSSION consequences for the relative motion of the body and feet. Low and Here, we report that guineafowl can substantially vary their yaw high yaw steps (strides 1 and 4 from the sequence plotted in Fig. 3, during steady locomotion. In sequences in which birds maintained Fig. 4A) are compared in Fig. 6. In lab space, a stance foot stays position within a limited X-ray volume, yawing occurred over a fixed on the tread, which it follows regardless of pelvic orientation range of speeds. Multiple individuals in multiple runs surpassed (Fig. 6A,C). If pelvic yaw is small, the foot’s trajectory closely 20 deg of yaw compared with their direction of travel. This yaw parallels the body’s longitudinal axis (Fig. 6A). The more the pelvis offset was overlaid on the normal, rhythmic, intra-stride yaw is yawed away from the direction of travel, the more the foot’spath pattern. Guineafowl varied pelvic yaw both on a treadmill and when is skewed relative to the body (Fig. 6C). This difference in moving over a trackway. Marker-based XROMM analysis revealed trajectory is emphasized when viewed from a pelvic reference measurable LAR at the hip and knee regardless of yaw (Figs 3, 4). frame, as if looking down the body’s median plane from above Journal of Experimental Biology

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Left Right i 20 AB

10 External

Internal –10

ii Net stance hip LAR excursion (deg) –20 CDiii

5 0 –5 –10 –15 –20 –25 Average stance pelvic yaw (deg)

Fig. 5. Net hip LAR excursion during stance versus average pelvic yaw for 10 sequences. Net hip excursions were calculated as the difference of values at the beginning and end of stance, while yaw at the beginning and end of stance was averaged to calculate average stance yaw. Right hips are represented by filled circles while left hips are represented by open circles. Right yaws and internal LAR excursions are negative. Left yaws and external LARs are positive. The red circle shows data for the right stance phase of stride 2 highlighted in Fig. 4B.

(Fig. 6B,D). At low yaw, the transverse distance from the distal iv tarsometatarsus to the median plane is small and changes little over the course of stance (Fig. 6E). In contrast, at a larger yaw, the foot EF takes a diagonal path relative to the body midline, moving from 5 iii medial to lateral for the right leg and a right yaw (Fig. 6E). The 15 transverse distance from the median plane starts small and increases 4 over 5 cm through stance (Fig. 6E). 10 Hip LAR patterns closely match transverse distance patterns. 3 ii During the low yaw step, hip LAR forms a W with similar beginning 5 i 2 and ending values (Fig. 6F, i and ii). During the high yaw step, the Hip LAR (deg) LAR Hip 0 hip begins more externally rotated, internally rotates through stance, 1

and ends at a much lower angle (Fig. 6F, iii and iv). The similarity of (cm) distance Transverse iv the transverse distance and hip LAR traces suggests that hip LAR is 0 0.1 0.2 0 0.1 0.2 used to modulate transverse distance during forward locomotion. Time (s) Time (s) Pooled data from several steady treadmill trials confirm this kinematic relationship (Fig. 7, filled circles). Larger net hip LAR Fig. 6. Modulation in foot trajectory and hip LAR for two different steps. Right stance phases from strides 1 and 4 of Fig. 3 and Fig. 4A are compared. excursions are associated with foot paths that deviate further away (A,B) Path of the distal tarsometatarsus in lab co-ordinate space (A, treadmill from or towards the median plane. Even more extreme transverse oriented vertically on the page) and pelvic co-ordinate space (B) when the distance and hip LAR excursions (Fig. 7, open squares) were pelvis is at low yaw (blue). (C,D) Path of the distal tarsometatarsus in lab co- achieved when an angled foam barrier at the front of the treadmill ordinate space (C) and pelvic co-ordinate space (D) when the pelvis is at high induced a bird to walk slowly at yaws of 35–65 deg. yaw (red). Pelvis and right limb poses are shown at touchdown. Limb poses are shown at toe on (solid, i and iii) and toe off (transparent, ii and iv). The dashed Asymmetric limb control line shows the pelvic midline. (E) Transverse distance (perpendicular distance from the distal metatarsus to the pelvic midline) over time for the steps from Yaws of sufficient magnitude correlate with a monophasic hip LAR A–D. (F) Hip LAR angle over time for the steps from A–D. pattern, in which the femora undergo substantial net LAR excursion during stance and swing (Fig. 4B, Fig. 5). At high yaws, the right time. Plots of hip, knee and ankle FE (Fig. 3), and hip LAR at low and left hips internally and externally rotate together, generating yaws (Fig. 4C, Fig. 5) demonstrate this kinematic symmetry. In similar curves when plotted versus time. Perhaps counter- contrast, at high yaws the limbs both retract in stance and protract in intuitively, these shared monophasic patterns are evidence of swing, but undergo hip LAR in opposite directions. kinematic asymmetry. Right and left hips internally and externally When walking with a yawed pelvis, the pattern of femoral LAR rotate simultaneously, but the two limbs are 180 deg out of phase resembles the coordination reported for guineafowl executing with respect to the stride cycle. One hip internally rotates in stance sidestep maneuvers (Kambic et al., 2014). During a sidestep, birds while the other internally rotates in swing. When both hips sequentially spread and converge the feet transversely (Fig. 8A; externally rotate, the side that was in stance has switched to swing Movie 2). Mediolateral motion of the limbs is similarly required for and the side that was in swing has switched to stance. In order to act forward movement while yawed, but the amount of mediolateral symmetrically, both limbs would need to undergo similar LAR skewing required depends on pelvic orientation. When the excursions during the same phase of the stride, thereby alternating in individual is induced to walk slowly at extreme yaws around Journal of Experimental Biology

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is not ‘natural’. We do not know whether or how often birds yaw 20 while walking and running in the wild. In the lab, one concern with marker-based XROMM is surgery-induced asymmetry. Yet our 10 subjects yawed in similar ways whether one or both limbs had been implanted. Based on head orientation, birds appear to have been 0 looking toward the same side they were yawing most of the time. Head deviation may provide evidence that the body is yawed, although it is not yet clear whether head motion precedes or follows –10 body rotation. Further analysis of the visual saccades that transition between ‘fixed’ head orientations relative to the body and direction –20 of travel is warranted (Dunbar, 2004).

Net stance hip LAR excursion (deg) The consistent treadmill bias to the right is suggestive that one or –6 –4 –2 0 2 4 6 more features of the lab – personnel, lighting, equipment, reflection, Stance transverse distance excursion (cm) etc. – were drawing the individuals’ attention, although data from a Fig. 7. Net hip LAR excursion versus distal tarsometatarsal transverse trackway in the same location and orientation do not show the same distance excursion for stance phases from 10 sequences. Filled circles pattern. It is possible that the researcher operating the treadmill from represent normal trials. Open squares represent steps from the induced yaw behind influenced asymmetry, although both hand-cranked and trial. Excursions were calculated as the difference of values at the beginning motorized systems yielded similar results. Regardless of the cause and end of stance. Only steps with complete data are plotted. The two steps from Fig. 6 are shown with their respective colors. of the right bias, yawing behavior was performed at will by all individuals on two different treadmills as well as on a trackway. If 45 deg, mediolateral and craniocaudal foot excursions are close to solely a lab-based phenomenon, orienting the body away from the equal (Fig. 8B). At progressively lower yaws, less and less skewing direction of travel may be common in other studies of avian of the foot trajectories is required (Fig. 8C,D). Therefore, forward kinematics as well. progression with substantial yaw (Fig. 8B,C) can be seen as For 2D analyses of motion from lateral perspective recordings, intermediate between the two extremes of a crab-like sidestep errors in FE joint angles will be minimal when yaws are small. (Fig. 8A) and symmetrical striding with little yaw (Fig. 8D). However, knowing that this behavior occurs should advise caution. The sidestep model of yawed forward progression explains the If the behavior is steady and cyclical, it is common to measure synchronized, monophasic pattern of hip LAR characteristic of high kinematics and physiological parameters (muscle activity, muscle yaw locomotion (Fig. 4B; Fig. 7, red circle). Net hip LAR produces strain, bone strain, etc.) for a single limb and assume that the other mediolateral excursions that spread and converge the feet to limb is behaving similarly at equivalent points in the stride. Indeed, generate appropriately skewed trajectories. For example, when our initial XROMM markers were implanted in only one limb. yawed to the right (Fig. 8C), the right limb moves from medial to Given our results, restraint is warranted in assuming symmetrical lateral in stance by net internal hip LAR. Simultaneously, the left limb motion if yaw is not measured. Yaw variation should also be limb moves from medial to lateral in swing, again using net considered when summarizing angular data as averages. Assuming internal hip LAR to widen the transverse distance between the feet. symmetry may mask explainable inter-stride variation, as numerous Both femora then externally rotate together to return to their researchers working on human locomotion have found (see narrower configuration. Rather than alternating, the striding limbs Hausdorff, 2007, for a review). exhibit hip LAR angles that are coordinated in time. Right–left Despite these cautions, there are potential benefits to individuals asymmetry is actually increased by making hip LAR patterns more routinely orienting away from their direction of travel. The patterns of identical. kinematic variation with yaw are intriguing from a motor control standpoint. Birds graded smoothly between biphasic and monophasic Implications and future work hip LAR patterns, and blended elements of each depending on the Our finding that guineafowl routinely yaw their bodies compared circumstances of the stride. The modulation of LAR, combined with with their direction of travel may be less significant if this behavior the similarity of yawed locomotion patterns to sidestep patterns, may

A BCD Fig. 8. Sidestep and yawed treadmill trials in overhead view. (A) Sidestep to the left. (B) Forward progression at a large induced yaw. (C) Forward progression at a large natural yaw. (D) Forward progression at low yaw. Arrows show the approximate direction of travel. Red lines show paths of the distal tarsometatarsi during stance relative to the pelvis for right and left steps. Journal of Experimental Biology

3842 RESEARCH ARTICLE Journal of Experimental Biology (2015) 218, 3836-3844 doi:10.1242/jeb.126193 be evidence that cranial–caudal limb motion and mediolateral limb Abourachid, A., Hackert, R., Herbin, M., Libourel, P. A., Lambert, F., Gioanni, H., motion are controlled by separate sets of muscle activation patterns, Provini, P., Blazevic, P. and Hugel, V. (2011). Bird terrestrial locomotion as ’ revealed by 3D kinematics. Zoology 114, 360-368. termed muscle synergies (e.g. d Avella et al., 2003; Ting and Alexander, R. McN., Maloiy, G. M. O., Njau, R. and Jayes, A. S. (1979). Mechanics MacPherson, 2005). Many researchers have found evidence of joint of running of the ostrich (Struthio camelus). J. Zool. 187, 169-178. coordination in steady locomotion (e.g. Ogihara et al., 2014), and we Andrada, E., Nyakatura, J. A., Bergmann, F. and Blickhan, R. (2013). Adjustments of global and local hindlimb properties during terrestrial locomotion identified similar integration of hip and knee LAR during of the common quail (Coturnix coturnix). J. Exp. Biol. 216, 3906-3916. maneuvering (Kambic et al., 2014). However, in this study we did Andrada, E., Rode, C., Sutedja, Y., Nyakatura, J. A. and Blickhan, R. (2014). not find a strong correlation between hip and knee LAR patterns – a Trunk orientation causes asymmetries in leg function in small bird terrestrial result that should be investigated further. locomotion. Proc. R. Soc. B Biol. Sci. 281, 20141405. Birn-Jeffery, A. V., Hubicki, C. M., Blum, Y., Renjewski, D., Hurst, J. W. and We seek to learn more about the muscles responsible for Daley, M. A. (2014). Don’t break a leg: running birds from quail to ostrich prioritise controlling femoral LAR (Hutchinson and Gatesy, 2000) as well as leg safety and economy on uneven terrain. J. Exp. Biol. 217, 3786-3796. the contribution of muscles having both FE and LAR moments Brainerd, E. L., Baier, D. B., Gatesy, S. M., Hedrick, T. L., Metzger, K. A., Gilbert, about the hip. Based on their stance phase activity pattern, muscles S. L. and Crisco, J. J. (2010). X-ray Reconstruction of Moving Morphology (XROMM): precision, accuracy and applications in comparative biomechanics such as the iliotrochantericus caudalis and medius appear to be research. J. Exp. Zool. 313A, 262-279. ideally situated to induce internal hip LAR (Gatesy, 1994, 1999b). Clark, J. and Alexander, R. McN. (1975). Mechanics of running by quail (Coturnix). Yet, Hutchinson et al. (2015) found that several of the most J. Zool. 176, 87-113. Cracraft, J. (1971). The functional morphology of the hind limb of the domestic significant hip extensors have large internal rotation (called medial pigeon, Columba livia. Bull. Am. Mus. Nat. Hist. 144, 175-265. rotation in their paper) moment arms at relevant joint angles as well. Daley, M. A. and Biewener, A. A. (2006). Running over rough terrain reveals limb Determining how these muscles (and active antagonists) interact control for intrinsic stability. Proc. Natl. Acad. Sci. USA 103, 15681-15686. with inertial loads and passive forces to modulate LAR during Daley, M. A., Voloshina, A. and Biewener, A. A. (2009). The role of intrinsic muscle mechanics in the neuromuscular control of stable running in the guinea fowl. stance will require a more sophisticated, fully 3D approach. J. Physiol. 587, 2693-2707. Likewise, the control of LAR in swing awaits further d’Avella, A., Saltiel, P. and Bizzi, E. (2003). Combinations of muscle synergies in experimental and dynamic modeling work. the construction of a natural motor behavior. Nat. Neurosci. 6, 300-308. Dunbar, D. C. (2004). Stabilization and mobility of the head and trunk in vervet The resolution of our XROMM approach has revealed how the monkeys (Cercopithecus aethiops) during treadmill walks and gallops. J. Exp. outwardly appearing ‘steady’ locomotion of striding birds can be Biol. 207, 4427-4438. asymmetrical in previously under-appreciated dimensions. Joints Gatesy, S. M. (1994). Neuromuscular diversity in archosaur deep dorsal thigh such as the avian hip and knee operate in 3D and cannot be reduced muscles. Brain Behav. Evol. 43, 1-14. Gatesy, S. M. (1999a). Guineafowl hind limb function. I: cineradiographic analysis to hinges. However, our understanding of the kinematic and speed effects. J. Morphol. 240, 115-125. mechanisms for controlling yaw during forward locomotion also Gatesy, S. M. (1999b). Guineafowl hind limb function. II: electromyographic analysis remains incomplete. Although we have identified patterns of LAR and motor pattern evolution. J. Morphol. 240, 127-142. excursion associated with yaw deviations, the coordination among Gatesy, S. M. and Biewener, A. A. (1991). Bipedal locomotion: effects of speed, size and limb posture in birds and humans. J. Zool. 224, 127-147. joints and degrees of freedom used to change yaw await further Gatesy, S. M., Baier, D. B., Jenkins, F. A. and Dial, K. P. (2010). Scientific analysis. rotoscoping: a morphology-based method of 3-D motion analysis and visualization. J. Exp. Zool. 313A, 244-261. Acknowledgements Goetz, J. E., Derrick, T. R., Pederson, D. R., Robinson, D. A., Conzemius, M. G., We thank Elizabeth Brainerd, David Baier and the Brown Morphology Group for their Baer, T. E. and Brown, T. D. (2008). Hip joint contact force in the emu (Dromaius work on XROMM. Ariel Camp, Peter Falkingham, Erika Giblin and Angela Horner novaehollandiae) during normal level walking. J. Biomech. 41, 770-778. Gordon, J. C., Rankin, J. W. and Daley, M. A. (2015). How do treadmill speed assisted with data collection. John Hutchinson, Danny Miranda and Mike Rainbow and terrain visibility influence neuromuscular control of guinea fowl locomotion? provided advice on joint coordinate systems. We extend thanks to Kia Huffman for J. Exp. Biol. 218, 3010-3022. help with the XMA Portal. Farish Jenkins, Jr and William Amaral originally designed Grood, E. S. and Suntay, W. J. (1983). A joint coordinate system for the clinical conical markers that were adapted for this study, with additional fabrication advice description of three-dimensional motions: application to the knee. J. Biomech. from Amy Davidson. Comments and suggestions from two anonymous reviewers Eng. 105, 136-144. significantly improved the final manuscript. Hancock, J. A., Stevens, N. J. and Biknevicius, A. R. (2007). Whole-body mechanics and kinematics of terrestrial locomotion in the Elegant-crested Competing interests Tinamou Eudromia elegans. Ibis 149, 605-614. 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