Unique Locomotory Mechanism of Mermis Nigrescens, a Large Nematode That Crawls Over Soil and Climbs Through Vegetation

Home , Ascaris suum, Mermis nigrescens, Mermithidae, Nematomorpha

JOURNAL OF MORPHOLOGY 222:133-148 (1994)

Unique Locomotory Mechanism of Mermis nigrescens, a Large Nematode That Crawls Over Soil and Climbs Through Vegetation

CARL GANS AND A. H. JAY BURR Department of Biology, University of Michigan, Ann Arbor, Michigan 48109 (C.G.); Department of Biological Sciences, Simon Fraser University, Burnaby, British Columbia, V5A 15'6 Canada (A.H.J.B.) ABSTRACT Females of Mermis nigrescens, a nematode parasitic on grasshoppers, climb through terrestrial vegetation where they lay their eggs. The 100-mm-long body of these nematodes bridges gaps in this three-dimensional substratum, and crawls efficiently over planar surfaces. The nematodes do not use the classical undulant pattern of nematode locomotion as one coordinated unit; instead they propel themselves in several independent, locally controlled zones that propagate posteriorly. A repeated motion of their anterior end laces the body around fixed objects at which force may be applied. Propulsive force is applied to objects as the body glides past the contact site. Intermediate loops are elevated above the surface where they cannot contribute to propulsion. These loops rise and fall with time due to varying differences in propulsive forces between the contact sites. Forces are applied to the objects by internally generated bending couples that are propagated along the trunk, propelling the body in a cam-follower mechanism. Bending couples are generated by the contraction of ventral or dorsal longitudinal muscle bands that apply compressive force to the cuticle. The muscle bands, consisting of a single layer of obliquely striated muscle cells, are closely applied to the cuticle and are separated from it only by a fibrous basal lamina and a thin extension of a hypodermal cell. The myofilaments of each sarcomere are parallel to the body axis and attached perpendicularly via dense bodies (z-line equivalents) to the basal lamina, which in turn is fixed to the cuticle via filaments passing through the hypodermal cytoplasm. Conse- quently, forces are transmitted laterally to the cuticle over the entire length of the muscle, compressing it parallel to the surface without need for attachment to the terminal ends of the muscle cells. Thus the muscles are engineered for local control of bending and avoidance of buckling. There is evidence that the motor nervous system of Mermis may not be as simple as in classical nematode examples, which may explain why Mermis is capable of a much more localized control of ~ocomotorymotion. Q 1994 Wiley-Liss, Inc.

After mating, the nonfeeding female of the been analyzed or described, although other parasitic nematode Mermis nigrescens lies nonclassical locomotory patterns of nema- dormant in the soil for almost 2 years while todes have been noticed (Stauffer, '24; Streu her eggs develop. Then, during wet weather, et al., '61; Reed and Wallace, '65; Croll, '70; she migrates to terrestrial vegetation on Crofton, '71; Lee and Atkinson, '77). which she lays her eggs (Cobb, '26, '29; Chris- The movements involved may be compared tie, '37). Unlike the well-studied nematodes not only to those of typical nematodes, but Caenorhabditis elegans and Ascaris suum, also to those of elongate aquatic and terres- adult female Mermis do not move by the classical undulant pattern (Gray and Liss- mann, '64). Instead, they locomote with a Address reprint requests to Carl Gans, Dept. of Biology, Univer- variable, puzzling wave motion that has never sity of Michigan, Ann Arbor, MI 48109.

0 1994 WILEY-LISS, INC. 134 C. GANS AND A.H.J. BURR trial vertebrates (Gans, '86). The compari- al.,'90). This study is based on - 14 hours of sons must take into account the architec- VHS and 8-mm videotape, plus additional tures of the internal force production and direct observations. The tapes were analyzed transmission systems. Nematodes transmit and figures prepared using a Panasonic AG forces to external objects via a thick cuticle. 6300 single frame video analyzer. Some speci- The obliquely striated muscles of nematodes, mens were allowed to climb vertical surfaces described previously (Rosenbluth, '65, '67; (e.g., felt, glass panes, and chicken-wire Hope, '69; Valvassori et al., '81; Waterston, mesh). Movements on these substrates were '88), must somehow link longitudinal forces compared with videotapes of travel over the generated by the contractile filaments to the surface of the soil, as well as amid grass and local cuticle. other vegetation in the field and in the labora- As important as architecture are compari- tory. The sites and magnitudes of force appli- sons of scale. Not only do vertebrates com- cation were established by having Mermis monly have diameters at least an order of cross gaps in the substrate from the main magnitude greater, but at a diameter of 0.4 body of felt to and from a patch of felt that mm and a mass of 15 mg, Mermis falls into a was suspended from a sling, for which the size range in which capillary adhesion effects force required to achieve a unit of displace- can be significant (Denny, '93). The length-to- ment could be calibrated using Pesola scales. diameter ratio of the nematode is - 300 (to- The continuous application of force in grass tal length -120 mm), much greater than was monitored by video recording the deflec- that of vertebrates and typical nematodes. In tion of grass blades. Ascaris, the ratio is 50 (Johnson and Stret- Some of the anatomical description derives ton, '80). Evidently nematode architecture from preparations made earlier for other must somehow overcome the tendency for a studies (Burr and Harosi, '85; Burr et al., long narrow tube to buckle under longitudi- '89; Burr and Babinszki, '90).Specimens pre- nal compression. pared for transmission electron microscopy This report describes the basic motor archi- (TEM)were killed instantaneously by immer- tecture and locomotor patterns of Mermis. It sion in cold 3% glutaraldehyde in 0.05 M stresses kinematics, but also provides infor- phosphate buffer at pH 7.2, cut into 1-mm mation about the production of internal forces pieces, and fixed - 16 hr at FC, then washed and the nature and sites of their exertion in buffer. They were further fixed in 2% onto the environment. It attempts to match Os04 in the same buffer for 2 hr, then washed the peculiar morphology observed with the in buffer. They were transferred through an functional differences seen in this species. ethanol series to propylene oxide and embedded in Araldite: 1 day in a 50% mixture with MATERIALS AND METHODS propylene oxide and 4 days in 100%. The Gravid female Mermis nigrescens, avail- Araldite mixture contained Araldite 6005 able once per year, were collected in the Fra- (37.5%), RD-2 (12.5%) and dodecenyl suc- ser Valley near Vancouver, British Colum- cinic anhydride (50%),and DMP-30 accelera- bia, Canada, and maintained in their dormant tor (1 drop per 5 ml). After polymerizing at state in soil in the dark at temperatures of 65°C and remounting, silver-gray transverse 10°C. Others were removed from artificially or longitudinal sections were cut on a Rei- infected Shistocerca gregaria hosts at full chert Om U2 ultramicrotome with a dia- size and maintained in soil at greenhouse mond knife and mounted on uncoated copper temperatures until molted to adult females grids. These were stained 15 min in 5% ura- and eye color was just visible, -4 months. nyl acetate and 5 min in lead citrate and Prior to behavioral observations, collected observed in a Philips 300 electron microscope Mermis were transferred to a humid cham- at 80 kV.As well, 1-pm sections were photo- ber with horizontal illumination and the tem- graphed under phase illumination. Several perature was raised to 21°C. Normally, 5-20% specimens of Mermis were fixed in 3% para- of worms can be activated on a given day formaldehyde, wax embedded, and cross- under these conditions. sectioned at 7 pm. They were stained either Animals were videotaped using several with hematoxylin or with DAPI (4',6'-diami- standard cameras with a macro lens at 30 fps dino-2-phenylindoleHCl), a fluorescent DNA at various magnifications and angles while stain. In addition, wholemounts were ob- the worms were traveling horizontally over served by dlfferential interference contrast the surface of black felt that was intermit- or fluorescence illumination to demonstrate tently sprayed with distilled water (Burr et commissures and nuclei of the motor ner- LOCOMOTORY MECHANISM OF MERMIS 135 vous system. Muscle cell length was mea- sured in polarized light micrographs.

RESULTS Morphology Mermis is very slender and outwardly cylindrical. The length and diameter of female specimens range from 50 to 150 mm, and .25-.5 mm, respectively and their mass is 5-15 mg just before the start of egg deposi- tion (N 2 20). Normally smooth, the surface of desiccating animals forms accordionlike rings, possibly suggesting that normal body form reflects internal hydrostatic pressure. Similar corrugations are observed on the in- side of sharp bends. As is true for Ascaris (Harris and Crofton, '571, transection of the Mermis body produces immediate discharge of fluid and release of pressure.

Cuticle The transparent cuticle is extremely thick, measuring 1/5th-%th of the radius of the nematode (Fig. 1).The typical nematode cuticle is ?/I5 the radius (Bird and Bird, '91). Lacking the external lateral ridges (alae) ob- Fig. 1. Mermis nigrescens. Cross section showing structure of the body wall and location of neurons. A thin served in other nematodes, the cuticle is extension of hypodermal ceIIs separates the cuticle and markedly thickened along inward lateral longitudinal muscle bands. Interneurons and ventral mo- ridges, as is typical of mermithids (Kaiser, tor neurons are contained within the ventral cord; dorsal '91). The structural function may be like that motor neurons originate in the ventral cord and send their axon or dendrite to the dorsal nerve cord via commis- of alae, to stiffen the body against lateral sures (not shown). From a nearly transverse section. bending, In a region of the head in which small radius lateral bending is known to occur during scanning motion (Burr et al., '901, basal zone containing many layers of smaller these ridges are diminished. The ridge is fibers (Fig. 2). In longitudinal sections these present in the 2-cm neck region behind the are seen to travel longitudinally before bend- head where larger radius lateral bending oc- ing toward a radial orientation and interdigi- curs as well as in the rest of the body where tating with other fiber layers (Lee '70; Burr, lateral bending is not observed. unpub.). The nematode cuticle is basically a three- layered cylinder composed of various cross- Muscles linked collagens and covered by a thin epicu- Nematode body-wall muscle is longitudi- ticle of another material (Wharton, '86; nal and is separated from the cuticle only by Wright, '91; Bird and Bird, '91). The cuticle a basal lamina and a 1.5-pm-thick extension ofMermis, described by Lee ('70),is shown in of the hypodermis (Figs. 1-3). The longitudi- transverse section in Figure 2. The osmio- nal muscle bands lie in subdorsal and subven- philic, 10-nm epicuticle is not resolved at this tral quadrants separated by cords of the hypo- magnification. The outermost layer (cortical dermis. The dorsal quadrants of Mermis and zone) is composed of a homogeneous-appear- other mermithids (Kaiser, '91) are unusual ing material penetrated by radial canals in having only a single muscle band; the through which electron-dense material of the ventral quadrants have the normal two on surface coat is secreted. Next, the median each side (Fig. 1). The greater cross-sectional zone is a spongy material through which two area of muscle in the ventral quadrant sug- layers of large, oval "giant" fibers wind heli- gests that there is more force available for cally around the body with opposite handed- ventral than for dorsal bending. ness. The crossing angles range from 111" to In adult female Mermis nigrescens, the 122"in wholemounts. Innermost is the thick arrangement of the muscle cells and contrac- 136 C. GANS AND A.H.J. BURR

Fig. 2. Mermis nigrescens. Transverse section showing structural zones of the cuticle. gf, giant fiber. Transmission electron micrograph. tile apparatus within the muscle bands is (Figs. 1, 3). The roughly 2-mm-long, 2-pm- very similar to the pattern in other large wide, cells overlap obliquely at their longitu- nematodes, e.g., Ascaris (Rosenbluth, '65, dinal ends and the myofilaments are joined '67). This is classified as polymyarian (more by half-desmosome connections to the basal than five longitudinal rows of muscle cells in lamina separating them. Similar observa- each band) and coelomyarian (having the con- tions are reported in larvae of an unidentified tractile region of each cell folded longitudi- Mermis species (Valvassori et al., '81). nally to form two parallel radial plates) (Hope, The radial plates of longitudinal contrac- '69; Bird and Bird, '91). In Mermis, each tile filaments are closely apposed to the band is composed of 5-30 rows of narrow, plasma membrane and the basal lamina; as ribbon-like cells surrounded by basal lamina described below, the myofilaments are con- LOCOMOTORY MECHANISM OF MERMIS 137

Fig. 3. Memis nigrescens. Transverse section of body wall muscle, basal lamina and hypodermis. bl, basal lamina; cu, cuticle; db, dense body; hyp, hypodermis; mi, mitochondrion; tf, region containing thick myofilaments. nected perpendicularly to the plasma mem- dense bodies, cytoskeletal components equiva- brane via dense bodies (Fig. 3). The fibrous lent to Z-lines of cross-striated muscle, are basal lamina between the cells is continuous similarly offset, making the sarcomere ar- with the basal lamina enclosing the muscle rangement appear parallelogramic in radial- band (Fig. 3), which, in turn, is connected plane sections through the contractile appa- across the thin hypodermal cytoplasm to the ratus. The ends of the filaments in adjacent cuticle at closely-spaced intervals via half- radial planes are aligned along a perpendicu- desmosomes and intermediate filaments lar to the filaments which is also normal to (Francis and Waterston, '91). Thus, this con- the surface of the radial basal lamina sheath tractile apparatus, as in other nematodes, between the cells. The ends of the adin fila- differs fundamentally from that of verte- ments are connected along this direction to brates by being linked to the cuticle all along the sarcolemma by dense bodies. These are the length of the muscle cell, rather than tied across the sarcolemma to the basal only at its ends. lamina. Another cytoskeletal structure (M- Unlike cross-striated muscle of vertebrates, line analog), similarly links the centers of the nematode locomotor muscle cells are ob- thick filaments to the cell membrane and liquely striated (Toida et al., '75; Lanzavec- basal lamina (Waterston, '88). The basal chia, '77). The myofilaments lie parallel to lamina is structured as if to transmit forces the long axis of the worm. However, in any parallel to its surface. Fibrous strands run in radial plane, adjacent myofilament rows are multiple directions in the lamina (Rosen- offset longitudinally. Their attachment to bluth, '67) and evidently are attached across 138 C. GANS AND A.H.J. BURR the sarcolemma to the intracellular dense bodies (Francis and Waterston, '91). In the coelomyarian type of nematode muscle, these attachments connect the contractile filaments perpendicularly to the radial sheath between cells, then radially to the cuticle. Local contraction shortens the dis- tance between dense bodies and the force is transmitted radially and probably obliquely via the fibrous basal lamina sheath to shorten a local region of the attached cuticle. Thus, contractile forces are applied to the cuticle locally and continuously along each muscle band, rather than only at the ends of the muscle fibers as in vertebrates and arthro- Fig. 4. Mermis nigrescens. Distribution of nuclei along pods. In Mermis, as in Ascaris (Rosenbluth, ventral cord. Ventral view of DAPI-stained, glutaralde- '67), localized compression of the cuticle can hyde-fixed wholemount of a region anterior to the vulva. produce an accordion-like, 500-pm corrugation in its outer surface. This is to be distin- guished from the fine, 0.33-pm-spaced annu- identifying sensory neurons by light micros- lations resolved by the electron microscope in COPY. the cuticle of Mermis (Lee, '70; Burr, unpub- In both Caenorhabditis elegans (White et lished) and other nematodes and also thought al., '76, '86) and Ascaris suum (Stretton et to provide flexibility (Bird and Bird, '91). al., '78) the motor neurons of the ventral cord are arranged in five units, each with a Motor system basic set (ignoring the extra neurons in the The muscles in the dorsal and ventral quad- anteriormost units) of 11 cells and seven rants of the nematode body wall are con- commissures. There are no sensory neurons trolled by motor neurons in the dorsal and in either the dorsal or ventral cords. In adult ventral cords respectively. Each muscle cell hermaphrodites of Caenorhabditis, 57 motor sends a process to the nearest nerve cord neurons with 36 commissures innervate 79 where it receives excitatory and inhibitory muscle cells, and in adult females of Ascaris, innervation (Rosenbluth, '65; Stretton, '76; 59 motor neurons with 36 commissures in- White et al., '76; Walrond et al., '85). The cell nervate - 50,000 muscle cells (Stretton, '76; bodies and nuclei of hth the dorsal and Sulston and Horvitz, '77). By comparison, ventral motor neurons lie in the ventral cord, the total number of muscle cells in adult and some send an axon or dendrite to the female Mermis is -10,000 (length of each dorsal cord via circumferential commissures muscle cell, 2 mm; number per cross section, (Stretton et al., '78; Johnson and Stretton, 200; and length of the worn, 100 mm). The '80; White et al., '86). This pattern also oc- number of muscle cells appears to corre- curs in Mermis: Nomarski differential inter- spond to the length of the nematode, but the ference contrast discloses commissures un- number of neurons clearly does not. The der the transparent cuticle and DAPI much larger motor system of Mermis may be fluorescent staining shows no nuclei in the related to the more complex locomotory be- dorsal cord. Unlike Caenorhabditis elegans havior we observe. and Ascaris, numerous (80/mm) large hypodermal cell nuclei are located in the ventral Locomotion cord (Fig. 4) and the numbers of commis- Movement patterns sures are uniquely high. This prevented the The travel of Mermis over horizontal sur- enumeration of motor neuron nuclei. On the faces proceeds by a unique combination of other hand, we counted 11.6 commissures body movements that are most clearly re- per mm in one worm, suggesting a total of vealed in video recordings of locomotion on approximately 1,000. In Caenorhabditis and moist felt, such as the sequence shown in Ascuris, seven out of 11 motor neurons send Figure 5. The crawling motion appears slug- commissures to the dorsal cord (White et al., gish. Movement between contact sites re- '76; Stretton et al., '78); thus the number of quires six or more seconds and movement of motor neurons in Mermis must be more than one 100 mm body length requires N 60 sec in a thousand. As yet we have not succeeded an active individual crawling effectively on LOCOMOTORY MECHANISM OF MERMIS 139

lime 5 rnm (sac) - 0 See 5 0 10 I5 20 4

30 11 35 40 13 45

50 15 55

60 65 18 70

75 24 80 Fig. 6. Mermis nigrescens. Propagation of bends along 85 body, slow change in wave shapes, and slippage of contact 90 points along surface (arrows). Tracings from video tape 95 of side view during phototaxis on felt surface. The position of the vulval region, shown in solid black, indicates the motion of the trunk through the wave trajectory.

Fig. 5. Mermis nigrescens. Body wave fluctuations in a side view. Tracings from video recordings during photo- tween contact sites. The motor system of taxis on moist felt surface. Sequence begins with tail out Mermis must be capable of accurately propa- of the field of view to left. gating a local curved region along the body as it moves past a point in space. Video recordings indicate that the shape felt (Burr et al., '90). The locomotion illus- and sizes of the waves between the fixed trated in Figure 5 is not that of Thomas, '90). lacing sites change gradually (Figs. 5, 6). Note, however, that Mermis adults neither Whenever the worm travels concertedly to- feed nor defecate; they live on food stored dur- ward a particular goal such as a light source, ing a parasitic larval stage and lack an anus. the vertical waves between contact points Typically, during steady locomotion, Mer- show minimal elevation as in Figure 5, secs 0, mis contacts a flat horizontal surface at 6-8 40, 90-95. At other times, these portions ventral points with an elevated wave be- between contact sites can gradually form large tween each point (Burr et al., '90). At any vertical and horizontal waves, which some- instant, the curves formed by the body are times twist to one side or fold into helical stiff; they are conserved for a short time loops. We show below that the various el- whenever a portion of an animal is pushed or evated wave shapes are not directly related to lifted from the surface. propulsion, but are a consequence of differ- Observation of the movement of the body ing propulsive velocities at the ventral con- trunk through the curved trajectory is facili- tact points. tated by the obvious dark egg masses and the Locomotion has to be analyzed in terms of vulva visible through the transparent cuticle, various movement routines. Key among them e.g., in the locomotion sequence on felt repre- are lacing, head elevation bouts including sented by Figure 6. Over periods of 2-4 sec, scanning and orientational motions, and nic- the position of the bends remains stationary tating. Reverse locomotion is also observed. in space while the body flows through the The subroutines not only establish the sites bend. Thus over short times, the wave or loop of force transmission, but also establish the of body created between two contact points path that will be traversed by the animal. establishes an aerial trajectory through which all of the following portions of the body travel. Lacing and other contact sites The body even passes through irregularities A worm progressing over a horizontal felt in the curvature of the elevated portions be- surface terminates a head elevatior, bout with 140 C. CANS AND A.H.J. BURR a unique lacing motion of the anterior few millimeters of the body. It pokes the anterior tip into the substrate (Fig. 7a), then slides the tip along the surface in a hooking motion (Fig. 7a-c) that is actually helical. This fre- quently succeeds in lacing the body under one or more fibers on the surface of the felt. After the head advances forward from the lacing site, the body just emerging from the site bends upward as the head is elevated, thus forming a tight curve at the site (Fig. 7d-f). The increased curvature at the lacing site is maintained as the body glides through the site. This is shown below to be instrumen- tal in the application of a propulsive force to the fiber. The head remains elevated until the next lacing site is probed. Lacing is more successful on felt than on the many other surfaces observed and tested; however, on other surfaces the general pattern is equivalent. Movement patterns in an array of three- dimensional surfaces such as encountered in grass is more complex, but with the observations on planar surfaces analyzed, they can be readily understood. For example, in the Fig. 8. Mermis nigrescens. Lacing sequence and force sequence illustrated in Figure 8a-c, one can application while climbing through grass. (a)Touching grass blade; (b)lacing around blade. (c) force application recognize lacing and tight curve formation bending blade to right. Tracing of anterior portion from about a grass blade. As well, the bending of videotape. Movement of grass blade (arrow) is not to scale. the grass blade indicates the concomitant application of force. Propulsive forces are strongest when gen- tact points or zones along the body. When- erated at the lacing sites as these are the ever lacing is not possible, the velocity is most secure positions of contact with the lower and one commonly sees slippage at the substrate. Whenever the substrate makes lac- zones of contact with the substratum (Figs. ing difficult, the animals appear to propel 5,6). themselves by use of other contacts with Scanning and orientation surface irregularities. On moist surfaces including felt, soil and grass, advantage is often A head elevation bout begins with the emer- taken of adherence by surface tension at con- gence of the head from a lacing site (Fig. 7c) and continues until the next contact with a surface. Throughout the head elevation bout, the anterior 2 mm, or “head” of the worm, oscillates in a scanning motion involving both vertical and sideways swinging motions (Fig. 7d-0. The next 10-20 mm, or “neck,” are involved in orientation to light or gravity (Burr et al., ’90). The scanning motion of the head modulates the photoreceptor illumination during phototaxis, and it is independent of the orientational motion of the neck (Burr and Babinszki, ’90). Orientation of the neck establishes the direction of locomotion and is Fig. 7. Mermis nigrescens. Lacing sequence and begin- modulated by the sensed direction of gravity ning of head elevation bout on felt. (a) poking into or light (Burr et al., ’90). The neck can be surface; (b)sliding along surface; (c)just after lacing bent sideways or in the vertical plane. The body under surface fiber; (d,e,f) head elevation with anterior tip scanning direction of light source. Tracing neck can also be involved in generation of from videotape of phototaxis. propulsive forces; however, the body behind LOCOMOTORY MECHANISM OF MERMIS 141 the neck is primarily involved in locomotion as a grass stalk, whereupon the worm tends and only dorsoventral bending is observed to bend or lace here and locomotion transfers (Burr et al., ’90). onto the new site. Nictation when crawling on soil makes it possible to discover plants Reversing and other objects that may be climbed. How- Nematodes commonly reverse locomotion, ever, their selection is by chance collision thereafter turning and resuming forward during the process of nictation. Nictation movement. Caenorhabditis elegans reverse similarly facilitates the discovery of new sur- spontaneously every 90 sec on average (Burr, faces after encountering an opening in vegeta- ’85) and reversal is elicited within 2 sec after tion. A nictation bout can terminate in a touching the anterior end, or more weakly by collision with the previous surfacethe usual light and changes in temperature, osmolar- case on a horizontal surface such as soil or ity, or concentration of odorants (reviewed the experimental felt. by Chalfie and White, ’88).The neural circuit Whenever the nidating portion of the ani- for the touch reflex is well worked out for mal makes a contact, it commonly tends to Caenorhabditis elegans at the level of identi- start a very different body orientation and fied nerve cells (Chalfie et al., ’85; Rankin, the subsequent movements indicate the inde- ’911. pendence of different regions of the trunk. Reversing is observed occasionally in Mer- The elevated body anterior to the new con- mis, but it nearly always occurs following the tact site reorients, bending sharply in the cessation of light during phototaxis. De- region of contact while retaining its previ- scribed initially as a stop response (Burr et ously existing curves anterior to the bend. al., ,901, full reverse locomotion has now These remain frozen in position for a while, been observed as a continuation of this re- although the more posterior portions of the sponse. Remarkably, it only starts some body keep on flowing through the new bend, 11-30 sec after the light source has been propelled by forces generated at posterior turned off. It begins with the halt in forward lacing sites. The new contact site and bend locomotion, then, the body simultaneously can be made anywhere along the reach of the forms a series of waves of shorter wavelength nictating portion whereupon additional pro- than usual-the “stop” response observed pulsive forces may develop at these sites. If previously. This simultaneously involves the the anterior tip makes contact, a new lacing entire body, rather than successively local- site may be formed by a normal lacing bout. ized regions, as in forward locomotion. Then, The various movement routines in different reverse locomotion begins. Unlike that in body regions appear independent, which is Caenorhabditis elegans, the tail does not completely untypical of classical nematode move as fast as the body. As a result the body locomotory motion. coils as it moves posteriorly. After 60 sec, - Evidence of cantilevering the forward locomotion resumes, but in a different direction than previously. Several observations show that Mermis is capabie of cantilevering-elevating its ante- Nictation rior or posterior body above the substrate by Nictating behavior might be deemed a loco- applying a torque at the nearest contact, motor subroutine. In Mermis it facilitates which acts as the fulcrum. For example, when navigation through a three-dimensional ar- the anterior tip is elevated during a head ray of surfaces, such as the terrestrial vegeta- elevation bout (Fig. 7d-0, contraction of dor- tion on which it lays its eggs. Nictation oc- sal muscles in the region of contact would curs whenever >12 sec elapse without the cause upward bending and thus provide the head coming into contact with a surface. This torque, whereas the posterior body is held can occur during a head elevation bout when down by laced regions, gravity, or surface crawling on soil or when encountering open- tension. Another example occurs during pho- ings in vegetation. Nictation consists of wide, totaxis to a light source elevated by 45” above horizontal, and circular swings of the ante- a horizontal felt substrate (Burr et al., ’90). rior body. As the body is propelled past the Remarkably, the anterior portion is held ori- anteriormost contact point, a continuously ented toward the source while the body glides increasing portion of the animal, up to half through the anteriormost contact point and, its length, becomes involved in the swinging. as a result, as much as a third of the body Nictation terminates whenever the swinging becomes cantilevered and aimed at the source. anterior portion contacts another object, such Nictation is an extension of this example, 142 C. GANS AND AH.J. BURR with an extensive length of the body not only would have exerted a propulsive force at that suspended anterior to a contact, but also site. being rocked and twirled. What is striking in The application of forces also was moni- all of these cases is the ability of the region in tored by having worms travel from a fixed which torque is developed to be propagated surface to a floating or suspended platform, posteriorly as the body glides past the ful- the excursion of which is very sensitive to crum. The activity in muscles and local mo- horizontal force applications. Mermis travers- tor neurons is continually replaced by equiva- ing from lacing sites on the fixed substratum lent activity in adjacent posterior regions. to new ones on the floating platform exerted horizontal forces of <0.1 g, the minimum Generation of propulsive force detectable by our force transducer. These The lacing sites clearly are the primary forces or deflections occurred irregularly with sites of force transmission, as documented by differing amplitudes and at differing rates. several incidental observations. On felt, the Apparently the stiffened body transmits caudal portion of a Mermis commonly may be forces between the lacing sites. In field videos seen suddenly to accelerate away from the of Mermis climbing amid grass blades, simi- restraint of a surface fiber whenever the cau- lar force application is seen to cause variable dal tip passes out from under the lacing site. but continuous deflection of a grass blade This movement must have reflected the stress while the body slides past the blade (Fig. 10). incorporated in the last loop. This is seen The sequential inserts show that, during force even more graphically whenever the fiber ofa application, the large curvature of the body newly formed lacing site suddenly ruptures, in the region of the contact point is main- for the anterior tip then makes a sudden up- tained while the body anterior to this region ward motion (Fig. 9). The upward force would is waved about independently, probably in appear to be due to torque transmitted along search of a new lacing site. the body from the posterior cantilevering In closeups of lacing sites on felt where one site. The tension developed by contraction of can discern the threads under which the the dorsal muscle bands during cantilevering worm looped, one can see these threads being is evident. As the tension is relieved, 1/15 sec stretched and can determine the times and after the break, the curvature of the regions vectors along which forces are being applied. anterior and posterior to the fulcrum changes The force application appears to be continu- in the manner expected if upward bending ous. Whenever the worm applies substantial couples had been present in these regions. forces, its curvature along the lacing site has Whenever Mermis does not manage to lace a characteristic shape; its radius is lowest under a fiber at a contact point, it will still just anterior to the contact with the fiber and form a downwardly bent profile and touch much greater posterior to the fixed site (Fig.9). the surface but the contact point is observed to slip backward with time. This suggests Surface tension effects that if a fixed site had been secured, the body Mermis are small enough to be affected by surface tension, and this affects their propulsive patterns. For instance, if tossed against a vertical plastic mesh that has been sprayed with water, the body will adhere to it without slipping. The body is smooth and naturally slippery, not sticky; therefore, the adherence must be due to the forces of surface tension. The forces of surface tension are sufficient to maintain the 5-15-mg animal in the vertical plane and to facilitate its vertical locomotion. Whenever the body contacts a wet grass blade during nictation, it obviously sticks to it. The body adheres to plant stems at right angles to it. When a portion of the worm contacts a Fig. 9. Mermis nigrescens. Evidence of force applica- portion of the sprayed felt, one can then tion to a lacing site on felt and of cantilevering at a more see deformation of the droplet meniscus at posterior contact point. 1, force being applied to fiber prior to breaking; 2, release of tension just after fiber the contact site, indicating that forces can be breaks; 3, further dorsal flexure. Note changes in curva- transferred between the body and the surface ture. Tracings from videotape of phototaxis. film. LOCOMOTORY MECHANISM OF MERMIS 143

15 sec lasec 24 sec I osec 4 sec 7 sec 12 sec ,

-2 I I I I I I 0 5 10 15 20 25 TIME (sec)

Fig. 10. Mermis nigrescens. Application of force to a lacing site in grass. Inserts: Sequential body positions and deflection of grass blade in side view. Tracings from videotape. Arrows indicate original blade position. Graph: Deflection of grass blade maintained during sliding contact with blade.

A similar force transfer occurs whenever phorea; Ascaris and Caenorhabditis belong the animals traverse smooth planar surfaces, in the subclass Rhabditia of the class Secern- such as wetted vertical glass or broad leaves. entea (Maggenti, '83). Adult Mermis re- In such cases the body waves are flattened semble Nematomorpha, which also have an against the surface by the action of surface extremely elongated, free-living, nonfeeding tension, although the head and neck are re- adult stage (Hyman, '51). There are similari- peatedly raised away from the surface and ties in the cuticle, particularly the presence locomotion is slower and less effective. Such of helical crossed-fiber layers (Zapotsky, '71; film-embedded animals show at least two Cham et al., '83;Seymour, '831, and in the movements. The first is slippage of the flat- body wall muscle, with the coelomyarian tened body waves over the surface, primarily muscle structure and plate-like muscle cells in the anterior half. In the second, the poste- separated by sheets of basal lamina (Lanza- rior body breaks away from the surface and is vecchia, '79; Bresciani, '91). Although similar- placed into a more advanced position. On ity of architecture suggests that force genera- vertical wetted glass, the second motion can tion also should be similar, Nematomorpha be more effective in gaining altitude than the differ from nematodes in fundamental ways first. In both cases, propulsive forces are gen- including their lack of a dorsal nerve cord erated by pushing against the resistance of (Cham et al., '83). The cuticular structure is surface tension, sliding friction and/or plant quite different in detail from that of nema- hairs. todes, and Valvassori et al. ('81) state that the sarcomeres have a completely different DISCUSSION ultrastructure. There appear to be no descrip- General tions of the motor behavior in Nematomorpha. Nematoda are divided into two major Many features of Mermis are shared by groups representing two independent clades: other mermithid genera (Kaiser, 1991): in- Mermithidae are included in the class Adeno- cluding the extremely threadlike body, 144 C. GANS AND A.H.J. BURR smooth cuticle containing two layers of est radius) just anterior to the contact point crossed helical fibers, six instead of eight and decreases posteriorly to it (Figs. 9, 10). muscle bands, a parasitic larval stage which As already demonstrated by Gray ('53) and absorbs nutrients through its cuticle, non- used much earlier in the engineering so- feeding adults, and a greatly modified diges- called cam follower design (Ham et al., '58; tive tract for food storage. However, only in Shigley, ,611, a movable track will slide past a Mermis and Pheromermis, are the hosts in- fixed resistance site so that travel is from a fected by ingestion of eggs; in the other gen- zone of small curvature to one of large curva- era, second stage larvae actively invade the ture. In this interpretation, the body of the host. Only in Mermis nigrescens is the adult worm represents a muscularly deformable female specialized for navigating through veg- track that will travel past the fixed lacing etation. Larvae of other nematodes, includ- site. The body will move in the direction that ing some Mermithidae, can climb over vegeta- decreases the curvature at the contact. As tion; however, their unusual length, makes the zone of muscular contraction creating female Mermis the only forms of nematodes the small localized curvature moves posteri- that can bridge major discontinuities in the orly along the body, there is continual for- environment. Toward this end, nematode nic- ward sliding of the body. tating behavior has become adapted in Mer- The velocity with which the worm then mis for the search for new stems of vegeta- slides past a site reflects a number of factors: tion not continuous with the surface being the force applied to the site, forces transmit- traversed. Moving over soil and climbing veg- ted axially from other sites, the momentum etation appears to be associated with the of the traveling object, and the friction at the need to migrate from its over wintering site site. The small mass of a worm (5-15 mg) in soil to distribute eggs among the terminal and low velocity indicate that momentum leaves and shoots on which grasshoppers feed. will be negligible relative to the potentially Locomotion in Mermis is seen to have a large forces of surface tension. The net force number of unique features. Mermis forms contact sites alternating with aerial curves; propelling a given segment of the body must crawling with body waves elevated above the be the sum of varying forces then being ap- substratum is very rare. It is unlike "typical" plied to the several individual contact points nematode locomotion in which the dorso- anterior and posterior to the region. ventral waves lie in plane of the surface film The waves do not obviously relate to the and the worms travel on their sides; nor does pattern of propulsion. The shape of such it reflect any of the special modes of nema- loops is even more misleading whenever the tode locomotion (Stauffer, '24; Reed and Wal- animals cannot fix the body at lacing sites, lace, '65; Wallace, '59, '68; Seymour, '73; whereupon the contact points slide, the wave Keller and Falkovits, '83; Dobrolyukov, '86; form changes, and the positions of force appli- Lee and Biggs, '90; Niebur and Erdos, '91). cation shift among successful lacing sites. Adult female Mermis do not move by the The body passes continuously past a contact classical undulant pattern of other nema- site or through the path of an aerial curve, but todes (Gray and Lissmann, '64) or swimming the velocity can vary between regions. For snakes (Gray and Lissmann, '501, in which each pair of lacing (force-transmitting) sites, the waveform along the body is controlled as the length of body between the sites and a single propulsive unit. The localized control consequently the shape of the aerial segment over curvature observed for Mermis is not represents a balance between the velocities of evident in Caenorhabditis elegans, nor has it travel through the anterior and posterior been reported in other nematodes described. sites. If the velocities differ, the shape will change. If the leading velocity is greater than External force transmission the following one, the body loop will shrink A major question is how a propulsive force and may shift from a compressive to a tensile can be applied to substrate at the contact state. If the reverse is true, the body will be sites. The force transmission to a lacing site under compression and the loop will increase is similar to that utilized in propelling a in size. Additional loops may form as more of snake past a single peg (Gray and Lissmann, the body is shifted into the interval; the length '50; Gasc et al., '89; Gans, '94). The body of stiffened body loaded in compression will bends around a fixed object such as a felt first of all deflect from the line connecting the fiber or a grass stalk. The local curvature is lacing sites and next twist to form a new asymmetrical; it attains a maximum (small- helical loop. These external manifestations of LOCOMOTORY MECHANISM OF MERMIS 145 forces raise questions as to internal force primary difference between oblique-striated production and coordination. and typical vertebrate cross-striated muscle. A body segment in one of the curves be- The combination of lateral attachment and tween contact sites will be in compression if staggering of the myofilaments allows the the axial forces pushing it from behind ex- worm to engage in very continuous, yet lo- ceed the axial forces pulling it forward. Evi- cally adjustable, shortening of the attach- dence of this occurs when the tip is poked ment sites, i.e. the longitudinal strips of cu- against a felt surface during lacing: the size ticle. This potentially provides highly localized of the loop in the body increases behind this control of the shape and movement of the point (Fig. 7a). The axial forces being trans- cuticular envelope. It should permit bending mitted by the stiff body derive from forces with minimal risk of major buckling of the applied at one or more posterior contact sites. cuticular surface. Thus it compensates for If, later, force is applied at the lacing site, the the absence of other skeletal structures. loop is diminished as tension develops in the Cuticular structure same segment. The cuticle of Mermis has several features Muscle architecture unique among nematodes that may be related to the unusual locomotory motion. The locomotor muscles of the various Crossed helical fiber layers are found in some nematodes for which we have descriptions all other nematodes, e.g., in Ascaris (Bird, '71). have oblique striations. This pattern may be However, in Mermis and other mermithids, geometrically confusing, which led to old ar- this trellislike structure lies in the outer cu- guments about whether obliquely striated ticle rather than near the inner surface of the muscle represented twitch or slow muscle cuticle near the muscles. Fiber crossing angles (Toida et al., '75; Lanzavecchia, '77; Hoyle, in Mermis range from 111"to 122"in whole- '83).There are also differences in the way the mounts, slightly larger than the angle of 110" contractile regions are packed within the cells, found by Lee ('70). A crossing angle of 110" e.g., between coelomyarian and platymyarian corresponds approximately to the critical spi- muscle types (Hope, '69; Bird, '711, differ- ral angle of 54.7"for a spiral-wrapped hydro- ences that may reflect the diameter of the skeleton for which there is no change in worms. However, there is an overwhelming volume or pressure with muscle-induced number of common denominators for ob- shortening; pressure would increase with liquely striated muscle. Whatever the cell shortening at the considerably larger angle, types, shapes and attachments, the myofila- 150", found for adult Ascaris (Harris and ments of the locomotor muscles always lie in Crofton, '57). At the crossing angles observed parallel to the long axis of the worm and in Mermis, a crossed helical structure ap- define the direction of shortening. The con- proaches the functional limit of a spiral tractile region is clearly striated in that it has wrapped hydroskeleton (O'Grady, '83) and it repeated sarcomeres with thick and thin fila- is doubtful that it would provide the force nec- ments that slide past each other in shorten- essary to restore the effect of muscular con- ing. The sarcomeres are staggered relative to traction. However, cutting of the cuticle evi- each other, with adjacent sarcomeres ad- dently releases an internal pressure. The thick vanced so that they lie at -6", rather than fibrous basal zone may provide the needed go", to the longitudinal axis. The staggering anisometric elasticity, and it is also better lo- produces the characteristic oblique striation cated nearer the muscle layer. This network observed in sections parallel to the cuticle in of many smaller and shorter fibers is arrayed Caenorhabditis, or parallel to the intercellu- differently than in the equivalent layer of lar sheets of basal lamina in Ascaris and other nematodes. The cuticle of Mermis also Mermis. The myofilaments are longer than lacks struts in the median zone, which in their equivalents in vertebrates (Rosenbluth, many nematode species provides a compliant '67; Waterston and Francis, '85). connection between the basal and cortical In another distinction from the vertebrate layers (Inglis, '64; Wright, '91).However, the model, the contractile forces developed in the spongy underlying material of the median myofilaments are not transferred at the ends layer of Mermis may serve this purpose. of the fibers to connective tissues and ten- dons. Instead, the forces are transmitted lat- Stiffness and internal forces erally to the cell membrane and a fibrous Anesthetized Mermis and Ascaris are limp, basal lamina. This lateral connection is the yet, Mermis can stiffen short sections of its 146 C. GANS AND A.H.J. BURR body so that these may be pushed along by insufficient to prevent buckling during long- forces originating at posterior contact sites. column compression. Such stiffening is observed whenever Mermis It is clear that the body-wall muscle must is poking the felt substrate during lacing. provide most of the needed stiffness. Fur- Also, in the pendulum platform experiment, ther, the structure of obliquely striated the body is stiffened so that the initial con- muscles provides a unique mechanism for tact of the head deflects the platform away applying bending couples over long distances from the fixed site on which the trunk of the without causing buckling. Because the longi- worm rested. Further, Mermis can cantilever tudinal force required for cantilevering is up to a third of its length as during head applied locally, rather than at the ends of a elevation bouts or nictating. Thus its very muscle band, the required compressive force thin body not only withstands a limited is distributed over continuous short local re- amount of longitudinal compression, but it gions of the cuticle. In effect, the long- can also resist bending. This observation is column compression ratio is reduced to a truly remarkable because the long-column length that can be accommodated by the local compression ratio to which the cantilevered cuticle without buckling. portion is exposed is above 75 (30 mml > .4 Our finding that Mermis is able continu- mm), well within the buckling range. The ously to maintain a bending couple at a local lack of long-columnbuckling with force appli- site of force application while its body glides cation at the ends of such an elongate column by is unique for the nematodes studied. This needs an explanation as it is at the limits of ability demands a control of the body muscles technological capability. unlike that for classical nematode locomo- Alexander ('87) has predicted that cuticles tion. Rather than being coordinated as one with helical fibers are under tension when unit by signals originating in the anterior or the body is straight and lose strain energy posterior end, the muscles must be con- when it bends, and therefore nematodes trolled locally, probably with local sensory should normally be bent. However, Mermis feedback. Obliquely striated muscle fibers are straightens if anesthetized by immersion in a well suited to the formation of local bending couples. Our evidence for a large number of saturated solution of phenoxypropenol in sa- motor neurons, perhaps two orders of magni- line. Also, Ascaris straightens whenever its tude above that ofAscaris or Caenorhabditis, muscles are relaxed by an injection of GABA suggests that the motor system of Mermis (Burr, unpub. obs.). would appear to be capable of local control. What, then, stiffens the body of Mermis so The locomotor mechanism appears to incor- that it can transmit collateral forces without porate several behavioral subroutines that buckling? Three nonexclusive possibilities ex- each deserve independent study. The cues for ist. The first is that stiffness is due to a lacing, and for nictating need to be estab- hydrostatic mechanism, the second assumes lished, as do those that establish the relative it to be inherent in the thick cuticle, and the magnitude of force application and velocity third is that it is due to muscle stiffness. The at each of the several lacing sites. Locomo- occurrence of a hydrostatic component is re- tion in a three-dimensional medium such as flectedby the corrugation of the cuticle when- soil has yet to be examined in nematodes. ever the animal is shortened by dehydration. One could look for similar locomotor behav- A component due to thick cuticle is indicated ior and mechanisms in other extremely elon- by the capacity of the dehydrated animal to gate worms, such as other mermithids and in move by bending. That the musculature con- nematomorphs. tributes a large component to stiffness is shown by the limpness of anesthetized Mer- ACKNOWLEDGMENTS mis. This study was reported in part at the The thick-walled cuticle is clearly flexible December 1992 Annual Meeting of the Ameri- enough to allow bending. Its internal mesh- can Society of Zoologists. The work was sup- work of small fibers and helical crossing gi- ported by a grant from the Leo Leeser Foun- ant fibers will stiffen it locally and the inter- dation for Tropical Biology (to C.G.) and from nal lateral ridges may stiffen it further to the Natural Sciences and Engineering Re- restrict bending to the lateral plane. How- search Council of Canada (to A.H.J. B.). The ever, these structures and hydrostatic pres- authors are grateful for the assistance of sure provide only a fraction of the stiffness Gwen Bollerup for collection of Mermis, Ed observed in unanaesthetized worms and are North with some of the behavioral observa- LOCOMOTORY MECHANISM OF MERMIS 147 tions, Hella Prochaska with the electron mi- Gans, C. (1986) Locomotion of limbless vertebrates: Pat- croscopy, Jennifer Wu with preparation of tern and evolution. Herpetologica 42/1):3346. Gans, C. (1994) Approaches to the evolution of limbless the micrographs, and Elizabeth Carefoot with locomotion. Cuadernos de Herpetol. 8(1):12-17. the artwork. Gasc, J.-P., D. Cattaert, C. Chasserat, and F. Clarac (1989) Propulsive action of a snake pushing against a single site: its combined analysis. J. Morph. 201:315- LITERATURE CITED 329. Alexander, R. McN. (1987) Bending of cylindrical ani- Gray, J. (1953)Undulatory propulsion. Quart. J. Microsc. mals with helical fibres in their skin or cuticle. J. Sci. 94(4):551-578. Theor. Biol. 124:97-110. Gray, J.E. and H. Lissmann (1950) The kinetics of loco- Bird, A.F. (1971) The Structure of Nematodes. London: motion of the grass snake. J. Exp. Biol. 26:354-367. Academic Press. Gray, J.E. and H. Lissmann (1964) The locomotion of Bird, A.F. and J. Bird (1991) The Structure of Nema- nematodes. J. Exp. Biol. 41:135-154. todes, 2nd ed. San Diego: Academic Press. Ham, C.W., E.J. Crane, and W.L. Rogers (1958) Mechan- Bresciani, J. (1991) Nematomorpha. In F.W. Harrison ics of Machinery, 4th ed. New York McGraw-Hill. and E.E. Ruppert (eds): MicroscopicAnatomy of Inver- Harris, J.E., and H.D. Crofton (1957) Structure and tebrates, Vol. 4. Aschelminthes. New York Wiley-Liss, function in nematodes: internal pressure and cuticular pp. 197-218. structure in Ascaris. J. Exper. Biol. 34t116-130. Burr, A.H. (1985) The photomovement of Caenorhabdi- Hope, W.D. (1969) Fine structure of the somatic muscles tis eleguns, a nematode which lacks ocelli. Proof that ofthe free-living marine nematode Deontostorna califor- the response is to light and not radiant heating. Photo- nicum Steiner and Albin, 1933 (Leptosomatidae).Proc. chem. Photobiol. 41.577-582. Burr, A.H.J., D.K. Eggleton, R. Patterson, and J.T. Leut- Helminthol. SOC.36:lO-29. scher-Hazelhoff (1989) The role of hemoglobin in the Hoyle, G. (1983) Muscles and Their Neural Control. New phototaxis of the nematode Mermis nigrescens. Photo- York: Wiley-Interscience. chem. Photobiol. 49r89-95. Hyman, L.H. (1951) The Invertebrates. Vol. 3. Acantho- Burr. A.H.J. and F.I. Harosi (1985) Naturallv crvstalline cephala, Aschelminthes and Entoproeta. The Pseudo- hemoglobin of the nematode Mermis nig&c&s. Bio- coelomate Bilateria. New York: McGraw-Hill. phys. J. 47:527-536. Inglis, W.G. (1964) The structure of the nematode cu- Burr, A.H.J. and C.P.F. Babinszki (1990) Scanning mo- ticle. J. 2001. (Lond.) 143:465-502. tion, ocellar morphology and orientation mechanGm in Johnson, C.D., and A.O.W. Stretton (1980) Neural con- the phototaxis of the nematode Mermis nigrescens. J. trol of locomotion in Ascaris: Anatomy, electrophysiol- Comp. Physiol. A 167:257-268. ogy and biochemistry. In B.M. Zuckerman (ed): Nema- Burr, A.H.J., C.P.F. Babinszki, and A.J. Ward (1990) todes as Biological Models, Vol. 1. New York: Academic Components of phototaxis of the nematode Mermis Press, pp. 159-195. nigrescens. J. Comp. Physiol. A 167t245-255. Kaiser, H. (1991) Terrestrial and semiterrestrial Mermi- Chalfie, M. and J. White (1988) The nervous system. In thidae. In W.R. Nickle (ed): Manual of Agricultural W.B. Wood (ed): The Nematode Caenorhabditis el- Nematology. New York: Marcel Dekker, pp. 899-965. egans. Cold Spring Harbor, NY: Cold Spring Harbor Keller, J.B. and M.S. Falkovits (1983)Crawling of worms. Laboratory, pp. 337-391. J. Theor. Biol. 104:417492. Chalfie, M., J.E. Sulston, J.G. White, E. Southgate, J.N. Lanzavecchia, G. (1977) Morphological modulations in Thomson, and S. Brenner (1985) The neural circuit for helical muscles (Aschelminthes and Annelida). Inter- touch sensitivity in C. elegans. J. Neurosci. 5:956-964. nat. Rev. Cytol. 51r133-186. Cham, S.A., M.K. Seymour, and D.J. Hooper (1983) Lanzavecchia, G. (1979) Three-dimensional reconstruc- Observations on a British hairworm, Parachordodes tion of the contractile system of the Nematomorpha wolterstorfii (Nematomorpha: Gordidae). J. Zool. muscle fiber. J. Ultrastruct. Res. 66:201-223. (Lond.) 199.275-285. Lee, D.L. (1970)The ultrastructure of the cuticle of adult Christie, J.R. (1937) Mermis subnigrescens, a nematode female Mermis nigrescens (Nematoda).J. Zool. (Lond.) parasite of grasshoppers. J. Agnc. Res. 55(5):353-364. Cobb, N.A. (1926) The species ofMermis, a very remark- 161513-518. able group of nemas infesting insects. J. Parasitol. Lee, D.L. and H.J. Atkinson (1977) The Physiology of 8:66-72. Nematodes, 2nd ed. New York: Columbia University Cobb, N.A. (1929) The chromatropism of Merrnis subni- Press. grescens, a nemic parasite of grasshoppers. J. Wash. Lee, D.L. and W.D. Biggs. (1990) Two- and three- Acad. Sci. 19:159-166. dimensional locomotion of the nematode Nippostrongy- Crofton, H.D. (1971) Form, Function and Behaviour. In lus brasiliensis. Parasitol. 101:301-308. B.M. Zuckerman, W.F. Mai, and R.A. Rohde (eds): Maggenti, A.R. (1983) Nematode higher classification as Plant Parasitic Nematodes, Vol. 1. New York: Aca- influenced by species and family concepts. In A.R. demic Press, pp. 83-113. Stone, H.M. Platt, and L.F. Khalil (eds): Concepts in Croll, N.A. (1970) The Behavior of Nematodes: Their Nematode Systematics. London: Academic Press, pp. Activity, Senses and Responses. London: Edward Ar- 2540. nnia Niehur, E. and P. Erdos (1991) Theory of the locomotion Denny, M.W. (1993) &r and Water: The Biology and of nematodes. Dynamics of undulatory progression on Physics of Life’s Media. Princeton, NJ; Princeton Uni- a surface. Biophys. J. 60:1132-1146. versity Press. O’Grady, R.T. (1983) Cuticular changes and structural Dobrolyukov, A.I. (1986) The mechanism of locomotion dynamics in the fourth-stage larvae and adults ofhca- of some terrestrial animals by traveling waves of defor- ris suum Goeze, 1782 (Nematoda: Ascaridaidea) devel- mation. J. Theor. Biol. 119:457-466. opingin swine. Canad. J. Zool. 61:1293-1303. Francis,R. and R.H. Waterston (1991) Muscle cell attach- Rankin, C.H. (1991) Interactions between two antagonis- ment in Cuenorhabditis elegans. J. Cell Biol. 114:465- tic reflexes in the nematode Cuenorhabditis elegans. J. 479. Comp. Physiol., A 169:59-67. 148 C. GANS AND A.H.J. BURR Reed, E.M. and H.R. Wallace (1965) Leaping locomotion VIII. Ultrastructural analysis of body wall muscles by an insect parasitic nematode. Nature, Lond. 206: from Mermis sp. J. Ultrastruct. Res. 76.82-88. 210-21 1. Wallace, H.R. (1959) Movement of eelworms in water Rosenbluth, J. (1965) Ultrastructural organization of films. Ann. Appl. Biol. 47(2):366-370. obliquely striated muscle fibers in Ascaris Zumbricoi- Wallace, H.R. (1968) Dynamics of nematode movement. des. J. Cell. Biol. 25:495-515. A. Rev. Phytopath. 6t91-114. Rosenbluth, J. (1967) Obliquely striated muscle. 111. Con- Walrond, J.P., IS. Kass, A.O.W. Stretton, and J.E. Don- traction mechanism of Ascuris body muscle. J. Cell. moyer (1985) Identification of excitatory and inhibi- Biol. 34:15-33. tory motoneurons in the nematode Ascuris by electro- Seymour, M.K. (1973) Motion and the skeleton in small physiological techniques. J. Neurosci. 5:l-8. nematodes. Nematologica 19:4348. Seymour, M.K. (1983) Some implications of helical fibers Waterston, R.H. (1988) Muscle. In W.B. Wood (ed): The in worm cuticles. J. Zool. (Lond.) 199:287-295. Nematode Caenorhabditis elegans. Cold Spring Har- Shigley, J.E. (1961) Theory of Machines. New York bor, Ny: Cold Spring Harbor Laboratory, pp. 281335. McGraw-Hill. Waterston, R.H. and G.R. Francis (1985) Genetic analy- Stauffer, H. (1924) Die Lokomotion der Nematoden. sis of muscle development in Cuenorhubditis eleguns. Beitrage zur Kausalmorphologie der Fadenwiirmer. Trends Neurosci. 8.270-276. Zool. Jahrb. (Syst.) 49:l-118. Wharton, D.A. (1986) A Functional Biology of Nema- Stretton, A.O.W. (1976) Anatomy and development of todes. London: Croon Helm. the somatic musculature of the nematode Ascaris. J. White, J.G., E. Southgate, J.N. Thomson, andS. Brenner Exp. Biol. 64.773-788. (1976) The structure of the ventral nerve cord of Cue- Stretton,A.O.W., R.M. Fishpool, E. Southgate, J.E. Don- norhabditis elegans. Phil. Trans. Roy. SOC.Lond. B moyer, J.P. Walrond, J.E.R.Moses, and1.S. Kass(1978) 2 75: 327-348. Structure and physiological activity of the motoneu- White, J.G., E. Southgate, J.N. Thomson, and S. Brenner rons of the nematode Ascaris. Proc. Natl. Acad. Sci. (1986) The structure of the nervous system of Cue- 753493-3497. norhabditis elegans. Phil. Trans. Roy. SOC. Lond. B Streu, H.T., W.R. Jenkins, and M.T. Hutchinson (1961) 314:l-340. Nematodes associated with carnations. New Jersey Wright, K.A. (1987) The nematode’s cuticle-its surface Agric. Exp. Sta. Rutgers Bull. 800. Sulston, J.E. and H.R. Horvitz (1977) Post-embryonic and the epidermis: Function, homology, analogy-a cell lineages of the nematode Caenorhabditis eleguns. current consensus. J. Parasitol. 73:1077-1083. Develop. Biol. 56:llO-156. Wright, K.A. (1991) Nematoda. In F.W. Harrison, and Thomas, J.H. (1990) Genetic analysis of defecation in E.E. Ruppert (eds): Microscopic Anatomy of Inverte- Caenorhabditis elegans. Genetics 124:855-872. brates, Vol. 4. Aschelminthes. New York Wiley-Liss, Toida, N., H. Kuriyama, N. Tashiro, and Y. Ito (1975) pp. 111-195. Obliquely striated muscle. Physiol. Revs. 55:700-756. Zapotsky, J.E. (1971) The cuticular ultrastructure of Valvassori, R., M. de Equileor, and G. Lanzavecchia Paragordius uurius (Leidy, 1851) (Gordioidea: Chor- (1981) Studies on the helical and paramyosinic muscle. dodidae). Proc. Helminthol. SOC.Wash. 38:228-236.