Jianfengia Hou 1987

Results External attributes and diagnostic traits Two Jianfengia specimens of similar size, YKLP11117 (Fig. 1a – e) and NIGPAS 100123b (Supplementary Fig. 1), provide views of the external morphology of this species, with an emphasis on its rostral carapace and its associated skeletal attributes. Jianfengia multisegmentalis has a segmented trunk, approximately 2.5 cm in length comprising 27-28 homonomous segments (Fig. 1a). A rostral carapace 5 mm in length and barely 3 mm in width covers the cerebrum and the first three postcerebral segments (Supplementary Table 1). Paired eyestalks extend laterally from the carapace immediately in front of a pair of uniramous postocular ‘ great appendages ’ that originate from beneath the carapace (Supplementary Fig. 1a – c). Each ‘ great appendage ’ comprises six podomeres of which the second and third provide an elbow-like articulation 7. The elongated shaft of the third podomere terminates as three articulating blades that together furnish a stubby chela. These appendicular attributes conform to the general category of ‘ great appendages ’ ascribed to members of the paraphyletic clade Megacheira (Supplementary Table. 2), the significance of which is discussed later. Rostrally, a pair of protrusions extending from the front of the carapace flank a substantial forward-projecting anterior sclerite,which arises from beneath the edge of the carapace with which it likely articulates (Figs. 1b, 2a – c). Despite considerable flattening typical of Chengjiang fossils, features of the anterior sclerite indicate that it is capped by three areas we interpret as ocellar-like lenses comparable to those described for the anterior sclerites of the deuteropodian Odaraia alata 8, thereby corresponding to frontal eyes of trilobites 9 and the nauplius eye/ocelli of extant pancrustaceans 10, 11. As shown in Fig. 2 a-c, these putative ocelli overlie a palisade of rod-like components here interpreted as photoreceptors at the base of the anterior sclerite. Superimposition of neural traces from the anterior sclerite of Jianfengia specimen YKLP17299 onto the corresponding exoskeleton of YKLP11117 (Fig. 2d) further supports the presence of ocelli represented by a system of diverging axons from the anterior sclerite.These extend into the most rostral neuropil of the brain in a manner identical to axonal projections from a pancrustacean ’ s ocelli into its rostral cerebrum 10, 11. Proven by developmental genetics of extant pancrustaceans, the combinatorial activity of Six3, FoxQ2 and hbn gene homologs is required for the development of rostral ocellar photoreceptors whose axons project into the prosocerebrum of the forebrain 12 – 14. The corresponding organization in Jianfengia of axonal projections from the anterior sclerite (Fig. 2d) thus identifies the prosocerebral domain of its cerebrum. Conspicuously absent in extant Myriapoda (and in Fuxianhuiidae), the nauplius/ocellar visual system therefore appears to be restricted to certain lineages of Artiopoda 8, Trilobita 9, and Pancrustacea 10, 11. Jianfengia further possesses compound eyes that crown each of the paired eyestalks extending laterally from beneath the carapace (Fig. 1b – e). This organization corresponds to eyestalk morphology typifying eumalacostracan crustaceans 15 and other members of Jianfengiidae 16 whose eyestalks evince two articles connected by a suture (Fig. 1c – f). In extant taxa, the combinatorial activity of Six3, Otx and Pax 6 defines the protocerebral domain of the pancrustacean forebrain 12, 17 and its nested optic neuropils, which receive information from the eye ’ s ommatidial array 18. Compound eyes on stalks and the areas to which their associated fossilized optic neuropils project thus indicate the protocerebral domain of the jianfengiid cerebrum. Reconstructing the brain of Jianfengia Our initial procedure for reconstructing the cerebrum of Jianfengia (Fig. 3) followed a routine used previously on the large fuxianhuiid cerebrum where incomplete neural traces from several specimens were mapped as mirror symmetric profiles within an envelope representing the bilaterally symmetric fuxianhuiid head shield 1. For Jianfengia, the neural traces, manifested as blue-black residues or as clusters of grey to near-black granules, were mapped onto a mirror-symmetric envelope for each of four specimens. The initial envelope was defined by the outline of the right frontal carapace and eyestalk of specimen YKLP11117 (from Fig. 1b). Flipping a copy of the outline across the animal ’ s midline provided the mirror symmetric envelope, as shown in Fig. 3b. The envelope was correspondingly adjusted to map neural traces in specimens that showed evidence of taphonomic misalignments, such as lateral displacements of the eyestalks or medial areas (e.g. specimen YKLP1367: Fig. 3c). These adjustments facilitate mapping of neural traces irrespective of such distortions, as shown for YKLP11368 (Fig. 3d). Likewise, specimen YKLP11369, in which only half the frontal volume could be retrieved, provided neural traces that mapped into the eyestalk and laterally alongside the esophageal foramen (Fig. 3e). To obtain the final reconstruction, the envelopes and their traces of each specimen were readjusted and registered in the summary envelope as shown in Fig. 3g. Finally, each tracing in one half of the envelope was mirrored in the other (Fig. 3h). The symmetrical profile from each of the four specimens was then filled and made 33% opaque. All four profiles were superimposed to provide a summed reconstruction of the Jianfengia cerebrum and its connection to the first three contiguous trunk ganglia T1 – T3 (Fig. 3h). With manual tracing, however, it is impossible to exclude anticipatory bias 19. To meet this challenge, we generated a second reconstruction using a minimum of human intervention. Two specimens were selected,YKLP 11367 and YKLP17299,both of which provide evidence of bilaterally preserved neuropils and tracts. Unlike specimen YKLP17299, whose mirror-symmetric neural traces suggest it is preserved flat (Fig. 3o), the asymmetric disposition of traces in specimen YKLP 11367 (Fig. 3i) indicates a slight taphonomic rotation around the fossil ’ s anteroposterior axis. Assuming that Jianfengia was a member of Bilateria, then these left and right traces in YKLP 11367 each represents a different depth within the specimen and are present on the contralateral side. These neural traces contribute to a mirror-symmetric organization as do the reflected outlines obtained by tracing. Reconstitution required four actions: the elimination of structures beneath a defined gray level (here 87% black of the CMYK scale), thus retaining the darkest puncta; inversion of the image to provide white profiles on a black background followed by the imposition of a Gaussian blur function (here radius expansion R = 10px) that expands and runs together puncta (compare Fig. 3j, k). To counteract asymmetry around the midline (Fig. 3k), the right side of the processed image beneath its ipsilateral eyestalk was isolated (Fig. 3l), flipped to the other side and merged with the left half, guided by matching the perimeters of the esophageal foramen (Fig.3m). The left half was then copied, flipped back to become the right half with the two halves joined at the midline (Fig. 3n). This provides a mirror-symmetric depiction of all traces present in the original specimen (Fig. 3n). Finally, tracings pertaining to the ocellar/naupliar system of specimen YKLP17299 (Fig. 3o) were identically processed (Fig. 3p). The mirror-symmetric image of YKLP17299 was superimposed onto that of YKLP11367, aligning their prominent ocellar features to provide the final reconstituted view of the jianfengiid cerebrum (Fig. 3q). This second procedure precludes subjective bias, yet the resultant image corresponds well with that obtained by tracing. Corresponding branchiopod and malacostracan cerebral organization in Jianfengia Both reconstruction methods resolved cerebral neuropils rostral to the esophageal foramen and fossilized nerve cords that extend around the esophageal foramen giving rise to neuropils disposed lateral to it. These nerves converge caudally to provide a synganglion immediately posterior to the foramen. The dispositions of these neuropils as well as neuropils defining the lateral expansion of the protocerebrum into the eyestalks correspond to traits typifying the brains described for the crown groups Branchiopoda and Decapoda 5, 18, 20 – 22. As would be expected from observations of extant eucrustaceans, traces of neural tissue within the jianfengiid esophageal foramen (Fig. 3h, q) also align with the standard locations of the appendicular labrum 15. In both reconstructions fossilized axon bundles diverge laterally from the anterior sclerite ’ s ocellar/nauplius eyes to supply the rostral neuropil of the prosocerebrum (Fig. 2d) where they merge with bilateral neuropils anterior and lateral to the esophageal foramen.These protocerebral areas receive the eyestalks ’ nerves which connect a series of three nested neuropils originating from beneath the compound eyes. (Fig. 3h, q). Next, we determined whether the reconstructed cerebra may correspond to the optic lobes and other neuropils that exist in today ’ s eucrustaceans.Both reconstructions of Fig. 3h, q allow an interpretive view of the jianfengiid brain (Fig. 4a – c). The lateral extension of the brain into the eyestalks comprises a small first optic neuropil (ON1) beneath the compound retina. This center is contiguous with a voluminous second optic neuropil (ON2) nestled against a smaller third neuropil (ON3). That the three neuropils are homologues of the lamina, medulla and lobula typifying pancrustaceans is supported by their alignment and similarity to the eye stalk neuropils exemplified here by the extant decapod malacostracan Astacus astacus 23 (upper left in Fig. 4a). The organization of the circumstomodeal nerve cords and their