Tiny Individuals Attached to a New Silurian Arthropod Suggest a Unique Mode of Brood Care

Tiny individuals attached to a new Silurian arthropod suggest a unique mode of brood care

Derek E. G. Briggsa,b,1, Derek J. Siveterc,d, David J. Sivetere, Mark D. Suttonf, and David Leggc

aDepartment of Geology and Geophysics, Yale University, New Haven, CT 06520-8109; bYale Peabody Museum of Natural History, Yale University, New Haven, CT 06520-8109; cOxford University Museum of Natural History, Oxford OX1 3PW, United Kingdom; dDepartment of Earth Sciences, University of Oxford, Oxford OX1 3AN, United Kingdom; eDepartment of Geology, University of Leicester, Leicester LE1 7RH, United Kingdom; and fDepartment of Earth Sciences and Engineering, Imperial College London, London SW7 2BP, United Kingdom

Edited by Alessandro Minelli, University of Padova, Padova, Italy, and accepted by the Editorial Board March 7, 2016 (received for review January 13, 2016) The ∼430-My-old Herefordshire, United Kingdom, Lagerstätte has Diagnosis. Features include a head shield with rostrum-like an- yielded a diversity of remarkably preserved invertebrates, many of terior projection, large uniramous antenna, chelate limb, and two which provide fundamental insights into the evolutionary history other biramous appendages in the head, the last similar to those and ecology of particular taxa. Here we report a new arthropod of the trunk; an elongated trunk with long, slender lateral spines with 10 tiny arthropods tethered to its tergites by long individual on the 11 tergites, with all trunk somites bearing limbs of which threads. The head of the host, which is covered by a shield that all but the last are biramous; and a short telson and long cerci. projects anteriorly, bears a long stout uniramous antenna and a chelate limb followed by two biramous appendages. The trunk Description. The head shield is subtriangular in dorsal view (Fig. 1 comprises 11 segments, all bearing limbs and covered by tergites A and J); the margins are incompletely preserved. The posterior with long slender lateral spines. A short telson bears long parallel area is raised medially into a broad axial ridge, which is also cerci. Our phylogenetic analysis resolves the new arthropod as a present along the length of the trunk (Fig. 1 J and K). An an- stem-group mandibulate. The evidence suggests that the tethered terior rostrum-like projection extends forward and somewhat individuals are juveniles and the association represents a complex ventrally a distance similar to the length of the rest of the head brooding behavior. Alternative possibilities—that the tethered indi-

shield (Fig. 1 J and K). An apparent series of four or five short EVOLUTION viduals represent a different epizoic or parasitic arthropod—appear less likely. slender lateral spines near the base of this projection are artifacts of preservation (Fig. 1J). The sides of the head shield bear a arthropod | Silurian | brood care | juvenile | Herefordshire Lagerstätte paired series of at least four long slender spines, projecting antero-laterally and curved convex dorsally (Fig. 1J). The spines increase slightly in length from anterior to posterior and are vidence of brooding in fossil arthropods is unusual and nor- Emally confined to eggs and early juveniles: later stage juve- similar in morphology to those on the trunk tergites. A swelling niles are rarely encountered. Among the highlights described from in the axial area on the ventral side of the head, which is aligned with the attachment of appendage 3, is interpreted as a hypo-

the Silurian Herefordshire Lagerstätte are ostracods preserving EARTH, ATMOSPHERIC, soft parts, including evidence of a brooding strategy that persists stome (Fig. 1 B and C). AND PLANETARY SCIENCES today: eggs and possible early juveniles are held within the space at There is no evidence of eyes. The first three head appendages the rear of the carapace (1). Here we report a new larger arthropod are morphologically differentiated, whereas the fourth appears – from the same fauna, with smaller arthropods attached to the very similar to those of the trunk (Fig. 1 B D and H). tergites by means of long threads. These smaller individuals lie Due to incomplete preservation proximally and lack of infor- within or are associated with a cuticular capsule, the largest about mation on the interior morphology of the head, it is not possible to 2 mm in length, with a gape through which the appendages determine the sequence in which the first two head appendages emerged. They preserve evidence of ∼6 pairs of appendages in insert. The relative position of antenna and chelate appendages in contrast to 15 (four of them in the head) in the adult. The evidence suggests that the attached individuals are juveniles that must have Significance added segments during the transition to an adult morphology, a strategy established in trilobites, eucrustaceans, pycnogonids, and The paper reports a remarkable arthropod from the Silurian other “Orsten” forms and in short great appendage arthropods by Herefordshire Lagerstätte of England. The fossil reveals a unique the early Cambrian (2–4). If so, the parent may be a female, al- association in an early Paleozoic arthropod involving tethering though male brood care is known in arthropods (in pycnogonids of 10 tiny individuals each by a single thread to the tergites so eggs are carried by the male, which is equipped with ovigers). that their appearance is reminiscent of kites. The evidence suggests that these are juveniles and that the specimen records a Results unique brooding strategy. This is part of a diversity of complex Aquilonifer spinosus is a new genus and species of arthropod from brooding behaviors in early arthropods heralding the variety the Herefordshire Lagerstätte, a late Wenlock (mid-Silurian) that occurs today. The possibility that the small individuals volcaniclastic deposit in Herefordshire, United Kingdom (5, 6). It represent a different arthropod, possibly parasitic, which colo- is preserved, as are the other fossils from this Lagerstätte, in three nized the larger individual, seems less likely. dimensions as a calcitic void fill in a carbonate concretion (7). The name of the new taxon refers to the fancied resemblance between Author contributions: D.E.G.B., Derek J. Siveter, David J. Siveter, and M.D.S. designed research; D.E.G.B., Derek J. Siveter, David J. Siveter, M.D.S., and D.L. performed research; the tethered individuals and kites, and echoes the title of the 2003 and D.E.G.B. wrote the paper with input from the other authors. novel The Kite Runner by Khaled Hosseini (aquila, eagle or kite; The authors declare no conflict of interest. -fer, suffix meaning carry; thus aquilonifer,kitebearer;spinosus, This article is a PNAS Direct Submission. A.M. is a guest editor invited by the Editorial spiny, referring to the long lateral spines on the tergites). The Board. material is a single specimen, the holotype OUMNH C.29695, 1To whom correspondence should be addressed. Email: [email protected]. registered at the Oxford University Museum of Natural History This article contains supporting information online at www.pnas.org/lookup/suppl/doi:10. (Fig. 1 and Movie S1). 1073/pnas.1600489113/-/DCSupplemental.

www.pnas.org/cgi/doi/10.1073/pnas.1600489113 PNAS Early Edition | 1of6 Downloaded by guest on September 30, 2021 Fig. 1. Holotype of A. spinosus gen. et sp. nov., “virtual” reconstructions. (A) Dorsal view. (B) Ventral view with juveniles omitted. (C) Ventral oblique view of right head appendages and hypostome (stereo pair). (D) Ventral–oblique (stereo pair). (E) Juvenile 5, oblique view with associated capsule. (F) Juvenile 5, lateral view. (G) Lateral view with juveniles removed. (H) Anterior–oblique view (stereo pair) of posteriormost head appendage and anterior trunk appendages showing exopods. (I) Proximal part of antenna showing spines (stereo pair). (J) Dorsal view without appendages (stereo pair) with juveniles numbered as referred to in text. (K) Anterodorsal–oblique view. (L) Anterior view (stereo pair). (M) Dorsal view of posterior of trunk (stereo pair). (N)Anterior view of trunk limb 9 (stereo pair). ap, juvenile appendages; b, basis; c, claw; ca, capsule; e, endopod; g, gut; h1–4, head appendages; hy, hypostome; t, telson; t1–11, trunk appendages; x, exopod. Numbers refer to trunk tergites, attached juveniles, or appendage podomeres as appropriate. (Scale bars, 1 mm.)

other Paleozoic arthropods, however, suggests that the uniramous enumerate—neither spines nor podomere boundaries are evi- nonchelate appendage (the antenna) is anteriormost (8). dent more distally. Spine bases are evident on the left appendage Head appendage 1 (green; Fig. 1 A, B, D, G, I, and K), as but not the spines themselves. Extrapolation suggests that the designated here, is uniramous, antenniform, and large. The right total number of podomeres is about 25. This first appendage is appendage is the better preserved (the reconstruction of the left about the same length as the body, including the “rostrum” but is incomplete distally). The angle of the slices (see Methods), excluding the cerci. subparallel to the length of the appendage, makes the proximal Head appendage 2 (pink; Fig. 1 A–D, K, and L) extends for- part difficult to interpret, but it may consist of three or four ward but not beyond the anterior projection of the head shield. segments similar in length to the more distal ones or, perhaps Subtle changes in direction along the length of the right limb less likely, a long basal segment. The appendage tapers gradually to a point. The individual podomeres are narrower proximally suggest that there may be as many as five proximal podomeres, and expand distally (Fig. 1I) to a point about their midlength but this is not certain (the slices run along the length of the limb, where they bear two short narrow spines, which project dorso- rather than transverse to it, obscuring details). The appendage laterally relative to the orientation of the trunk; more spines may terminates in a laterally directed swollen chela-like structure, have been present. The podomeres taper distally beyond the which terminates distally in two slender curved finger-like pro- spines to their articulation with the next podomere. Only the jections. A poorly preserved laterally directed projection from near segments in the proximal half of the appendage are easy to the base of appendage 2 (better preserved on the left example but

2of6 | www.pnas.org/cgi/doi/10.1073/pnas.1600489113 Briggs et al. Downloaded by guest on September 30, 2021 difficult to reconstruct; Fig. 1 A and B) may represent a slender The gut is preserved as an impersistent sediment fill; it exopod, but its nature is uncertain. becomes visible dorso-medially in the head because it lies too Appendage 3 (blue; Fig. 1 A–D, G, K, and L) is biramous. A close to the head shield for the intervening material to be visu- broad basis expands adaxially and may project into a gnathobase. alized (Fig. 1 A, J, and K). The position of the anus is unknown. It gives rise to an endopod, which extends abaxially and then The length of the body from the tip of the rostrum-like pro- curves axially so that the distal and proximal podomeres are jection of the head shield to the posterior margin of the telson is subparallel. Geniculations suggest the presence of five or six 9.5 mm. The large first antenniform appendage is about the same podomeres and a terminal spine. The exopod is much longer, is length (9.5 mm), and the cerci are about 7.3 mm long. more slender, and projects laterally. That on the right appendage Apart from its unusual morphology, the other remarkable fea- bends sharply ventrally and curves outward distally; it may end in ture of the arthropod is the attachment of multiple individuals to a series of short podomeres (Fig. 1L). the trunk tergites (Fig. 1 A, D,andJ–L). These 10 individuals, Appendage 4 (yellow–green; Fig. 1 A–D, G, H, K, and L) bears which are best seen when the trunk limbs are removed (Fig. 1J), an endopod similar to that of appendage 3, likewise with evi- are enumerated clockwise in what follows starting from the dence of five or six podomeres. The exopod is evident on the left anteriormost on the right side (Fig. 1J). They are shaped like side, where it is very short and projects just a short distance flattened lemons. They consist of an outer “shell” (here referred anteriorly (this ramus is incompletely preserved and has been to as a capsule) that does not appear to be calcified. The shell is lost on the right limb, but the data available are consistent with generally ∼15–20 μm thick where it is thinnest (Fig. S1B) but may the morphology of the trunk appendage exopods). be thicker in places perhaps as a result of soft tissue adhering to The trunk consists of 11 divisions (tergites) of similar length; the inner surface or the orientation of the capsule to the grinding the first two and the last one are slightly shorter than the rest plane. The capsule opens distally exposing filamentous structures (Fig. 1A), which may reflect a gradient in growth rate along the within. Some capsules, such as that of individual 3, show a narrow trunk axis (9). The trunk is near parallel sided, tapering markedly ridge along one margin, which may represent a kind of hinge (Fig. only in the last three tergites (Fig. 1M). Each tergite is comprised 1J and Fig. S1A). The largest capsules (individuals 3, 6, and 9) are of a broad, gently convex axial ridge occupying about half its about 2 mm in length (Fig. 1J). In some cases the filamentous width (excluding the long slender lateral spines), flanked by internal structures are separated from the capsule, particularly in lateral areas that are slightly concave dorsally (Fig. 1 J–L). Two individuals 1 and 5 (Fig. 1J). The smallest capsules (individuals 4 and 10) are less than 0.6 mm long (Fig. 1J). Thus, the capsules are

– EVOLUTION short triangular lateral projections of trunk tergites 1 10 bear ∼ × long slender spines, curved concave dorsally. These lateral spines characterized by a significant size range (the largest is 4 the are approximately evenly spaced along the length of the trunk. length of the smallest). Most individuals preserve a mass of tissue Only the posterior spine is preserved on the left side of tergite associated with the capsule, and individuals 2, 3, and 5 in particular preserve evidence of multiple paired slender projections 10, and the anterior spine, together with a hint of the posterior that represent limbs (Fig. 1 E, F,andJ and Fig. S1), although the one, on the right side. Tergite 11 appears to bear just one spine details are difficult to interpret due to their small size relative to on each side, which projects posteriorly (Fig. 1M). The bound- the spacing of slices. Individual 5, which is preserved outside its aries between the tergites are marked by transverse grooves in capsule, shows at least six pairs, some of them evident as curved the axial area (Fig. 1 J and K). The position of the maximum lines on a surface exposed during grinding (Fig. 1 E and F and height of a tergite lies progressively further posteriorly in tergites EARTH, ATMOSPHERIC, Fig. S1B). The body extends and tapers beyond the obvious ap- AND PLANETARY SCIENCES – K 7 9 (Fig. 1 ). pendages through a length similar to the appendage-bearing part. The first trunk appendage (appendage 5, blue–green) is similar – Individual 3 sits within its capsule and shows at least three pairs of to the posteriormost appendage of the head (Fig. 1 B D and H). A limbs projecting out of the gape (Fig. S1C). The smallest capsules broad basis expands adaxially; it may project into a gnathobase, (individuals 4 and 10) preserve hints of soft tissue within the but there is a significant gap between the opposing members of the capsule but no evidence of specific structures. pair here and in successive limbs. The basal podomere gives rise to Each capsule is borne by a slender flexible thread that originates an endopod, which extends abaxially and then curves axially so where the capsule tapers to a point. This proximal area of the that the distal and proximal podomeres are subparallel. Genicu- capsule is thickened (Fig. S1 A and B). The thread expands lations in the right appendage suggest the presence of six podo- abruptly just beyond the capsule and tapers gradually to a long meres and a terminal spine. The right appendage preserves a short slender portion that affixes to the host (Fig. 1J). Some of the incompletely preserved exopod projecting forward. threads appear to be discontinuous (e.g., that of individual 1), but – – Trunk appendages 2 10 (appendages 6 14) are similar in this is interpreted as a reconstruction artifact. The threads vary in morphology to the first trunk appendage. They increase slightly in length from about 1.5 mm (that of individual 5) to 3.3 mm (that of size to trunk appendage 6 and decrease slightly in the more pos- individual 9) (Fig. 1J). The threads are attached to the slender terior appendages (Fig. 1 B and D). The basis projects adaxially, lateral spines on the tergites, except for those of capsules 4 and 10, and the right limb of trunk appendage 7 preserves delicate spines. which are attached to the main part of a tergite (Fig. 1J). Left trunk appendages 8 and 9 preserve possible evidence of segmentation in the endopod (Fig. 1N). Four stout proximal Discussion podomeres are evident followed by a distal section of apparently Phylogenetic Position of the New Genus and Species. The combi- two podomeres (left trunk endopod appendage 9) terminating in a nation of characters in A. spinosus differs from that in any other slender claw (i.e., six podomeres + claw). The exopod is a long, known arthropod, living or fossil, and we therefore assign it to a flat, forward projection. The orientation of the slices combined new genus and species. Aquilonifer shows some similarity to with indifferent preservation makes it appear filamentous, but its Artiopoda, but when added to the analysis of Legg et al. (10) (with structure is unknown. This exopod is not evident in appendage 11 minor modifications; see Methods), it falls out as a stem-group (this is unlikely to be a preservational artifact, as the exopod is mandibulate lying above the Marrellomorpha and below those clearly present in appendage 10). “Orsten” forms that cannot be placed in crown-group Crustacea The trunk terminates in a small conical projection that extends (Fig. 2). Transposing the order of head appendages 1 and 2 (see beyond the last trunk tergite (Fig. 1M). This projection (referred Description) yields longer trees (142.60540 steps versus 142.16612 to here as the telson) bears a pair of slender parallel cerci that steps) with a largely unresolved topology. Modes of development are about three-quarters the length of the rest of the body (Fig. 1 are coded in the phylogenetic analysis (see Methods and citations A and G). It is unclear whether these structures are annulated. therein), but more derived brooding strategies are very diverse,

Briggs et al. PNAS Early Edition | 3of6 Downloaded by guest on September 30, 2021 Fig. 2. Cladogram showing the phylogenetic position of A. spinosus gen. et sp. nov. Shown is a strict consensus of the 12 most parsimonious trees of 142.16612 steps (consistency index = 0.513; retention index = 0.870), produced using New Technology search options in TNT (tree analysis using new technology) and using implied character weighting with a concavity constant of three. Numbers above nodes are GC support values. 1, Euarthropoda (crown- group); 2, total-group Chelicerata; 3, Artiopoda; 4, total-group Mandibulata; 5, Mandibulata (crown-group).

particularly among Eucrustacea (11), and provide little constraint epizoans, thoracican cirripedes such as Octolasmis,whichinfest on phylogenetic position. larger crustaceans today (15), are similar to these attached individuals. Some thoracicans, such as Pagurolepas, which live in association The Nature of the Attached Individuals. The very small size and with hermit crabs, have reduced the calcified plates that armor the consequent lack of detail revealed by the grinding technique make capitulum (16). The threads that tether the capsules to Aquilonifer, the individuals attached to Aquilonifer difficult to interpret. How- however, are much more slender and longer than the robust mus- ever, their size and morphology are inconsistent with protozoan cular peduncle of thoracican cirripedes. The attached individuals are ciliates such as peritrichs or with epiphytic algae. The outer covering also different to the larval stages of the cirripede Rhamphoverritor of the capsules resembles a carapace that encloses the body and reduncus from the Herefordshire Lagerstätte, which are about twice opens at one extremity. The absence of a mineralized shell, and the size, even though they represent developmental stages before presence of soft tissue beyond the capsule, together with the ap- attachment to a substrate (17). Given the potential for diversification parent symmetry, eliminates brachiopods. The serially arranged among arthropods, as exemplified by living crustaceans, the indi- paired structures within the capsules, about six in number (Fig. 1 E viduals attached to Aquilonifer could represent an unknown type of and F and Fig. S1) and sometimes projecting out or separated from epizoan; other lines of evidence, however, argue against this possi- the capsule, represent segmented appendages. Thus, the evidence bility. Epizoans have been reported from the Herefordshire biota— indicates that the attached individuals are arthropods. on brachiopods (18, 19)—but similar capsules to those described Arthropods attached by a thread are likely to represent one of here have not been observed tethered to any other animal from the three possible strategies: they are either parasites, epizoans, or fauna. Furthermore, the “host” arthropod was clearly living when the brooded juveniles. Comparative behaviors are most readily sought capsules became attached: it is unlikely to have tolerated the pres- among living crustaceans because they are by far the most diverse ence of so many drag-inducing epizoans, and head appendage 1 is group of aquatic arthropods today. Parasitic forms may retain ap- long enough to have cleaned the trunk tergites (“general body pendages for a motile phase in the life cycle. Living tantulocarids grooming” as in some living crustaceans) (20). Thus, the attached develop in a sac-like structure derived from the tantalus larva to individuals are more likely to be juveniles. which they are connected by a kind of umbilical cord, and the larva Tethering of capsule-like structures containing tiny individuals in turn is attached to the host crustacean (12, 13). Parasitic tho- is consistent with a brooding strategy, albeit one with no exact racican barnacles may retain cirri even though they feed by absorp- parallel among living arthropods; it would have protected the tion through the peduncle (14). A variety of parasitic copepods use juveniles from predation by keeping them close to the parent. a system of rootlets, some threadlike, to absorb nutrients from a Attachment by a stalk occurs in the embryos of freshwater variety of different hosts (13). The individuals attached to Aquilonifer, crayfish (Astacida), for example, which are tethered to the adult however, are unlikely to be parasitic because there would be no (21, 22). Some of the individuals attached to Aquilonifer show advantage in such long threads for absorbtion, and their most com- evidence of limbs: about six pairs are evident in individual 5, for mon attachment position, on the slender lateral spines of the host, example (Fig. 1 E and F). The length of the body in individual 5 is not a favorable site for accessing nutrients. would accommodate sufficient pairs to make up the number in The gape at the distal end of the capsules attached to Aquilonifer the host: they may not be preserved or have not yet developed would have facilitated feeding with the appendages. Among living fully. Release of the juveniles would have to have occurred

4of6 | www.pnas.org/cgi/doi/10.1073/pnas.1600489113 Briggs et al. Downloaded by guest on September 30, 2021 within a molt cycle of the adult, but this may have been extended care of eggs and early juveniles (1, 24, 29). All examples reported to avoid them being discarded. to date in early Paleozoic arthropods involve protection within a The size of the capsules varies from ∼0.5–2.00 mm. A diversity bivalved carapace, a strategy that evolved independently in of larval sizes is also known in recent ostracods (23), and eggs bradoriids (30), Waptia (29), and myodocope ostracods (1, 24). and juveniles have been reported together in individuals of the Extended parental care (31) has yet to be clearly demonstrated ostracod Nymphatelina from the Herefordshire Lagerstätte (1) in invertebrate fossils. and in ostracods from the Ordovician Beecher’s Bed (24). Em- Analogs for brood care in aquatic arthropods today are found in bryos brooded by the living crayfish Procambarus pass through crustaceans and pycnogonids. Several strategies exist: enclosure by the earliest stages “rather synchronously,” whereas rates of de- the thoracopods, by attaching eggs to the pleopods or ovigers (in velopment vary thereafter so that stage 3, 4, and 5 juveniles from the case of pycnogonids), within a dorsal brood pouch, within a the same batch may occur together under a mother’s abdomen marsupium formed by oostegites, and protection using an elon- (21, p. 573). Similar patterns could explain the variation in size of gated first pleopod (32). The distribution of these methods among the capsules attached to Aquilonifer. Alternatively the range in crustaceans suggests that most or all of them may have evolved size may indicate that the breeding adult accommodated more independently (32). Brooding in pycnogonids is different in that than one generation by molting at long intervals. the male rather than the female carries the eggs. Aquilonifer The correlation between the size of each capsule and the adopted yet another strategy that includes a dorsal position and length of its thread is not statistically significant (Figs. S2 and S3 attachment by a thread to a tergite. Among living crustaceans, a and Table S1). The correlation becomes stronger (although it is dorsal position for the embryos is confined to Thermosbaenacea, only significant at P < 0.10) when the thread length is augmented blind shrimp-like forms that live in caves and other underground by the distance between its point of attachment and the lateral systems (33). Their dorsal brood pouch is formed from an extension margin of the trunk of the host (i.e., the base of the slender tergal of the carapace in the female, and the embryos are transported spines). Thus, molting may have included lengthening of the there by currents generated by the thoracopods or transferred thread in a manner similar to epizoic thoracican barnacles (25) within a membrane that subsequently dissolves. The embryos of perhaps to improve access to particulate food (Table S1). Thermosbaenacea are free within the dorsal brood pouch (33). The embryos of freshwater crayfish (Astacida) are tethered to the adult Mode of Life. The morphology of the adult Aquilonifer provides by a stalk (21, 22). The egg cases are attached to the pleopods by a limited evidence of mode of life. The first head appendage shows a stalk secreted by cement glands on the sternum and pleopods. superficial similarity to that of the Cambrian arthropod Kiisortoqia When the hatchling emerges, it remains tethered to the egg case by EVOLUTION soperi from Sirius Passet, Greenland, which is antenna-like but a telson thread composed of the inner lining of the egg capsule. This armed with paired spines along its adaxial margin interpreted as maintains the attachment to the parent until the hatchling can use “possibly suitable for capturing prey” (26, p. 495). The spines on the hooks on the first pereiopod to grip the adult. In some crayfish, the equivalent appendage in Aquilonifer, however, are relatively an anal thread performs the same function as the telson thread. short, are more widely spread, and do not face adaxially. Fur- Thus, among the diversity of brooding strategies in living aquatic thermore the appendage in Aquilonifer tapers to a slender ex- arthropods are devices analogous, but very different, to that in tremity and does not appear suitable for a grasping function. This A. spinosus. first appendage may, in contrast, have been sensory or functioned Our interpretation of this remarkable specimen as representing 10 EARTH, ATMOSPHERIC,

in sweeping sediment in search of food. The second appendage is juveniles tethered to the parent A. spinosus, combined with its phy- AND PLANETARY SCIENCES chelate and presumably functioned in manipulating food. logenetic position among early arthropods, indicates that a com- Both right and left (in a less pronounced fashion) biramous plexity of brooding strategies evolved early in the history of the group. limbs curve adaxially at their distal extremity (Fig. 1 B and D). This position may be a response to burial. However, the seg- Methods mented nature and flexibility of the endopods suggest that they The holotype of A. spinosus (OUMNH C. 29695) was ground at 30-μm inter- could have functioned as walking limbs. Neither their morphol- vals, in two separate pieces. Surfaces were imaged digitally and image stacks ogy, nor that of the exopods, appear to be primarily adapted for used to generate a 3D “virtual fossil” using the custom SPIERS software suite swimming, indicating that Aquilonifer was benthic. The basipods (www.spiers-software.org) (34, 35). The virtual fossil (VAXML) was studied on- screen using the manipulation, virtual dissection, and stereoscopic-viewing were weakly spinose, but there is no evidence that they met in the capabilities of SPIERS. Images in Fig. 1 were rendered as ray-traced virtual midline. Food was presumably transferred directly to the mouth photographs using the open-source Blender package (https://www.blender. rather than transported anteriorly by the trunk limbs. The long org). The data are housed at OUMNH. cerci that project from the telson were presumably sensory. The holotype of A. spinosus (OUMNH C.29695) was studied as an interactive The juveniles would have operated at low Reynolds numbers and virtual model, in VAXML format. VAXML models (36) consist of a series of STL- likely used movement of the appendages to elevate them during or PLY-format files describing morphology, together with an XML-based file feeding (27) rather than relying on forward locomotion of the adult providing metadata. They can be imported into any 3D graphics package that to generate lift. The vast majority of crustacean larvae, for example, supports STL/PLY files or more conveniently can be viewed directly using the filter phyto- or zooplankton from the surrounding water (28). It is SPIERSview component of the freely available SPIERS software suite. To understand the affinities of Aquilonifer, it was coded into the extensive less likely that the juveniles attached to Aquilonifer were feeding on phylogenetic dataset of Legg et al. (10), including subsequent modifications the sediment surface, as there would be no obvious advantage in a by Siveter et al. (37), and a single additional character from Legg (38): the longer thread once the substrate was reached. Although the threads possession of an extensive posterior transverse ridge on the trunk tergites, are preserved curving ventrally, none of them reaches below the which was coded as present in some cheloniellids (see the supplemental sec- appendages; their arrangement may be partly a result of the parent tion in ref. 38 for discussion). This new dataset of 315 taxa and 754 characters being overwhelmed by sediment (capsules 7 and 8 are in a position (Dataset S1) was analyzed under general parsimony in TNT (tree analysis using where they might impede movement of the trunk appendages). new technology) v.1.1 (39). All characters were unordered and weighted using There is no evidence of the position of the oviduct in Aquilonifer or implied weighting with a concavity constant of three. Tree searches used 100 how the arthropod transferred or attached the offspring to its dorsal Random Addition Sequences with Parsimony Ratchet (40), Sectorial Searches, Tree Drifting, and Tree Fusing (41). Nodal support was measured using Sym- side. The long antenna may have been involved, or one parent may metric Resampling (each search used New Technology Searches with a change have attached the eggs to the other. probability of 33%) and is reported as GC values.

Brooding in Early Arthropods. Evidence of parental care is rarely ACKNOWLEDGMENTS. C. Lewis provided technical assistance, and David preserved in fossil taxa and is largely restricted, as here, to brood Edwards and other staff of Tarmac Western and the late R. Fenn facilitated

Briggs et al. PNAS Early Edition | 5of6 Downloaded by guest on September 30, 2021 fieldwork. Part of D.E.G.B.’s contribution was written while he was a by two reviewers, J. T. Høeg and N. C. Hughes. We thank the Natural sabbatical visitor at Stanford University. We are grateful to E. Lazo- Environmental Research Council (Grant NE/F018037/1), the John Fell Ox- Wasem for discussion and comments and S. McMahon for help in present- ford University Press Fund, the Leverhulme Trust (Grant EM-2014-068), ing the Supporting Information. J. B. Solodow assisted in coining the and the Yale Peabody Museum of Natural History Invertebrate Paleontol- taxon name. The paper benefited from insightful suggestions offered ogy Division for support.

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