Early astogeny in Hornera (; Cyclostomata; )

PETER B. BATSON, PAUL D. TAYLOR & ABIGAIL M. SMITH

BATSON, P.B., TAYLOR, P.D. & SMITH, A.M., 2019:11:15. Early astogeny in Hornera (Bryozoa; Cyclostomata; Cancellata). Australasian Palaeontological Memoirs 52, 23–30. ISSN 2205–8877.

In this contribution we document early astogeny in the cancellate cyclostome bryozoan Hornera cf. robusta. Colonies were collected by dredge from the mid-continental shelf off southeastern New Zealand and larvae settled in the laboratory. Developmental milestones of a single, newly metamorphosed protoecium were monitored over 18 days in seawater ranging from 14–16°C. After 48 hours the ancestrular tube had begun to appear. After five days the polypide had developed fully and the lophophore was extended, bearing nine tentacles. Days four and five marked the first steps in the formation of the second and third zooids. These initially sac-like chambers budded directly from the roof of the completed interior-walled protoecial dome, adjacent to the ancestrular tube. By day 16, divergence of the distal portions of the three zooids had begun, initiating development of the branch crown. Comparison with wild-caught Hornera cf. robusta ancestrulae showed that, overall, colony development in the laboratory-reared colony had proceeded normally. Direct budding of epi-protoecial zooids in Hornera parallels early astogeny in several genera of , including Lyroporella and Adlatipora. Our findings suggest that Borg’s (1926) description of early hornerid development may be more typical for the group than his revised model (Borg 1944).

Peter Batson ([email protected]). Department of Marine Science, University of Otago, Dunedin, 9054, New Zealand; Paul D. Taylor ([email protected]). Departments of Earth and Life Sciences, Natural History Museum, London SW7 5BD, United Kingdom; Abigail M. Smith ([email protected]). Department of Marine Science, University of Otago, Dunedin, 9054, New Zealand

Keywords: Hornera cf. robusta, , Cancellata, Cyclostomata, early astogeny, protoecium, ancestrula, interior-walled.

CYCLOSTOME bryozoan colonies arise from a free- vertically (Borg 1926) and there is no adnate stage swimming larva, which metamorphoses into a calcified, separating it from the developing main branch system hemispherical protoecium (= proancestrula, primary disc) (cf. Heteropora, Cinctipora and most other erect branching soon after settlement. In all taxa that have been studied, the cyclostomes). Consequently the ancestrula and early protoecium extends a calcified tube, develops a tentaculate zooidal growth stages are soon immured in the massive polypide, and becomes a functioning ancestrula within a secondary calcification typical of cancellates. week of larval metamorphosis (Nielsen 1970). Despite these challenges, Borg (1926) was able to Early astogeny in cyclostome bryozoans is poorly deduce aspects of cancellate early astogeny. Using sections known, despite indications that it has potential utility in of decalcified colonies ofHornera antarctica Waters, 1904, understanding the phylogeny of this group (Taylor et al. he described the distinctive vertical ancestrular tube and 2015; Jenkins & Taylor 2017). Ancestrula formation, further noted: ‘the zoids [sic] immediately succeeding the morphology and early zooid budding patterns have been primary zooid also grow straight upwards. It is remarkable documented in a handful of species across four of the five that at least the first two of these zoids take their origin extant cyclostome suborders (e.g. Nielsen 1970; Jenkins about as far proximally as the primary zoid; that is, in the & Taylor 2014, 2017; Taylor et al. 2015). Stages of early immediate neighborhood of the primary disc.’ (Borg 1926, astogeny in the remaining living suborder, the Cancellata, p. 305, fig. 49). has not been described in equivalent detail in the electron- Later Borg presented a different depiction of early microscopy era, even though the unjointed and vertical astogeny in Hornera, one in which the first daughter zooids ancestrular tube is one of the defining characters of this appeared to arise via septum formation (zooidal body wall group (Borg 1926, 1941, 1944). ‘fission’) from the distal part of the ancestrular tube, well The recent discovery by one of us (PDT) that the roof above the protoecial disc (Borg 1944, p. 176, fig. 23). This of the Hornera protoecium is fully interior-walled further interpretation has been widely used as a model for early stimulated our interest in tracking the earliest developmental hornerid development. stages in living cancellate colonies. In this contribution we The cancellate protoecium has been described briefly describe development of the ancestrula and early budding as part of broader species descriptions or comparative in the cancellate Hornera cf. robusta MacGillivray, 1883 surveys. Weedon (1998) described but did not illustrate using larvae spawned and settled in the laboratory. the protoecial skeletal ultrastructural fabric of an unknown cancellate species. Taylor & Gordon (2003) illustrated the PREVIOUS STUDIES OF HORNERID EARLY interior of the protoecium roof of the hornerid Calvetia DEVELOPMENT osheai Taylor & Gordon, 2003, showing it to be porous Cancellate early development is not easy to observe and unusually large (~0.75 mm). McKinney et al. (1993) directly without access to very small colonies (in Hornera, and Weedon (1998) described the hornerid protoecium as those less than ~2 mm high). The ancestrular tube grows ‘fixed-walled’ and ‘exterior-walled’ respectively. 24 AP Memoirs 52 (2019)

MATERIAL AND METHODS days with unfiltered ‘natural’ seawater from Portobello Living colonies of Hornera cf. robusta, H. foliacea and Point, Otago Harbour. Cultured phytoplankton was not Hornera sp. were collected by box dredge or beam trawl at provided. Gentle cleaning of any accumulated detritus was 80–95 m depth on the mid shelf off Otago Peninsula (46°S done using water jets from a soft plastic pipette. No attempt 171°E), southeastern New Zealand, by the University of was made to remove mobile epifauna occupying the clump. Otago research vessel RV Polaris II over an 18 month Regular observations and images were made over the next period (August 2014, December 2015, August 2015, 18 days using an optical microscope. January 2016). Biogenic substrata such as horse mussels (Atrina), other bivalve shells, and large frame-building bryozoans (Hippomenella, Celleporaria) were also Imaging collected in order to search for naturally settled ancestrulae Digital imaging of live larvae and ancestrulae was and early colonies. undertaken using a microscope-mounted digital camera. For some specimens, focus-stacking of multiple digital micrographs was used. Focus-stacked images were pre- Larval settlement and ancestrula rearing trials sharpened, then integrated using the auto-align and auto- Trials 1 & 2: collections of live specimens were made blend functions of Adobe Photoshop CS6. For SEM, organic in August 2014 and December 2015. Fertile Hornera material was removed from colonies using bleach solution colonies could be identified by their bright-orange brood (10 g/l sodium hypochlorite). Lightly calcified protoecia chambers. Colonies were transferred to Portobello Marine were bleached using a single drop of solution from a pipette Laboratory, where eight of them, representing three species under the microscope for a few minutes. Some specimens (H. cf. robusta, H. foliacea, Hornera sp.) were maintained fragmented upon drying using this method. Specimens were in individual containers (modified 2 L glass beakers) in sputter coated in gold palladium and scanned using a JEOL a controlled-temperature room at 11°C for six weeks. To 6700F FE-SEM (JEOL Ltd, Tokyo, Japan) at the Otago avoid removal or bubble entrainment of spawned larvae in Centre for Electron Microscopy, University of Otago. the tanks, no mechanical filtration or aeration systems were used, necessitating regular water changes. Water changes took place every 24–48 hours. Pre-cooled filtered seawater RESULTS was added once most of the water had been drained via a Across the five trials, fewer than 20 metamorphosed larvae drain at the bottom of the beaker, which also removed most developed beyond the one-day protoecium stage, and only accumulated detritus (and potentially some larvae). Water one developed significantly further (trial 5). Lightly to very motion was generated using motor-driven mechanical lightly calcified protoecia were produced by three different stirrers, and the parent colonies were mounted on racks hornerid species (H. cf. robusta, H. foliacea and Hornera midway up the sides of each container. Parent colonies and sp.; comparative data are not presented here). any feeding progeny were provided a mixture of cultured microalgae, Tetraselmis sp. and Pavlova sp. (2–4 mls of Trials 1 & 2 (August and December, 2014) mature culture after the water change), and maintained Using the ‘stirrer’ method over two trials, ~6–10 larvae of in darkness when unattended. The bottom and sides of Hornera sp. metamorphosed successfully to the calcified the containers were lined with acetate sheet that had protoecium stage. None developed beyond the early been pre-conditioned in seawater for 48 hours as a larval protoecium stage. Termination of development coincided settlement substratum. Containers and settlement sheets with a slightly ragged appearance and the attendance of were periodically inspected for free-swimming larvae and numerous motile microscopic organisms (probably ciliates protoecia. from the algal culture used in feeding). Some protoecia appeared cracked or fragmented when examined, probably Trials 3 & 4 (August 2015): these were similar to Trials 1 owing to handling damage. Three larvae from trial 1 were & 2, except that: (1) unlined pre-conditioned 2 L plastic successfully removed and mounted on SEM stubs. containers were used, (2) stirrers were not used, and (3) there was no feeding protocol, only regular changes of filtered seawater pre-cooled to 11°C. A week later after trial 3 Trials 3 & 4 (August, 2015) commenced, naturally spawned larvae were supplemented Outcomes were similar to trials 1 & 2. Trial 3 yielded with larvae liberated from the now-moribund colonies by three 24-hr+ stage protoecia: one of H. foliacea, one breaking open the brood chambers (trial 4). of H. cf. robusta, plus one more Hornera protoecium of an unknown species. Trial 4, using larvae liberated from Trial 5: The final ‘trial’ was serendipitous. A different broken brood chambers, did not yield any protoecia of rearing method was attempted owing to the poor outcomes ‘normal’ appearance. of the previous trials. Trial 5 began when a dredged bryozoan clump containing three species of Hornera was Trial 5 placed in a tank overnight on the day of collection (January Commencing January 17, 2016, a single specimen was 17, 2016). The next day a single early-stage protoecium spawned from a H. cf. robusta colony dredged from was observed attached to the branch of a fertile H. cf. 92 m depth, off Taiaroa Head, Otago, New Zealand robusta colony, about six millimetres away from a brood (45°47'S 170°55'E). The protoecium grew for 18 days and chamber. This colony was subsequently moved to a 20 L its study forms the main basis of this paper. bucket, which was kept in a seawater bath at 14–16°C (the ambient temperature of the reticulated seawater system at PML during mid January, 2016, measured daily at ~0900 Larvae and larval metamorphosis of Hornera cf. robusta hrs). Once a feeding lophophore had emerged from the Naturally released H. cf. robusta larvae (Figs 1A, 2A) are ancestrula, seawater in the bucket was replaced every 2–3 bright orange and densely ciliated, and are very large by AP Memoirs 52 (2019) 25 cyclostome standards. Equatorial diameters of the three mode of later zooids. The incipient bud of Z3 first appeared larvae measured were 250–260 μm. Larval behaviour on day five. It appeared as a low vertical buttress at the appeared consistent with that reported for other suborders junction of the ancestrular tube and the top of the proto- (see Nielsen 1970): alternating periods of free-swimming ecial dome, adjacent to Z2 in an anticlockwise position and slow creeping over the substratum were observed. (Fig. 1F). Metamorphosis in H. cf. robusta outwardly resembles that of other cyclostomes, beginning with eversion of Seven Days: The ancestrular tube was now ~200 μm long. the adhesive sac, which becomes a flattened protoecium Regular growth rings and a strongly dentate peristome (Figs 1B, C). The next phase of metamorphosis was not were increasingly evident, and the ancestrular tentacles observed directly in larvae that went on to form a calcified had grown in length (Figs 1G, H). The bud of Z2 began protoecium. a distal extension phase, ‘climbing’ up the ancestrular tube. As with the ancestrular tube, the growth of Z2 was punctuated by strong growth rings. However, these rings Ancestrula growth and early astogeny replicated the contour of the original budding plane on the The following account describes the first 18 days of protoecium roof, leading to an obliquely truncated growing development of the only Hornera cf. robusta protoecium tip and giving the zooid a hanging, sack-like appearance. to be successfully reared through to the multizooidal stage Also visible on day seven was the complete bud outline of (trial 5). Based on the timing of collection of the parent Z3. Completion of the proximal part of Z2 was required colony by dredging, it is inferred that the trial 5 protoecium for distal growth of Z3 to commence because Z3 emerged was ~24 hrs old, and probably no more than 36 hours old, partly from its wall. when first examined. This interpretation is consistent with the rate of protoecium development observed by Nielsen Ten days: The ancestrular tube had extended to ~300 μm (1970) in other suborders, but verification is needed (see (Fig. 1I). Rapid extension of Z2 and Z3 had also occurred, Discussion). and these zooids were close to ‘catching up’ with the ancestrular tube (although both their apertures were still ~24–36 hours: After one day, secretion of the skeleton obliquely truncated at ~45° at this stage). Developing was evident from the appearance of a pale, weakly lophophores were clearly visible in these budded zooids. corrugated ring ~120 μm in diameter, delineating the base of the future ancestrular tube, and also from the presence Twelve days: The three lophophores — those of the of faintly visible radiating lines around the periphery ancestrula, plus Z2 and Z3 — were protruded and actively of the protoecial disc (Figs 1D, 2D). Emerging from the feeding (Fig. 1J). At ~100 μm in diameter, the Z2 and Z3 base of the ancestrular tube and onto one one side of the zooids were much smaller than the flared orifice of the upper surface of the protoecium were two incipient walls ancestrula (~145 μm). Longitudinal ridges, corresponding that extended towards the edge of the disc (Fig. 1D). These to the individual teeth of the dentate peristome, were now curved ridges coincided with a gentle lateral bulge in the a prominent feature of the ancestrular tube, crossing the underlying contours of the protoecial dome, marking the transverse rings to create a grid-like pattern around the future position of the second zooid (Z2). It is inferred from tube. Z2 and Z3 lacked this feature, as they did not have the ridges that an interior wall had already developed on the dentate peristomes at this stage. upper surface of the protoecium. Fourteen days: By day 14, the peristomes of Z2 and Z3 Two days: The only visible changes were the first vertical had become level with the ancestrular tube, and were no extension of the ancestrular tube and slight lengthening of longer obliquely truncated. Growth increments in these the walls of Z2. In a 24-hour period the tube extended about zooids were now annular and transversely oriented, and 25 μm distally from the pale ring at the top of the protoecial their peristomes were dentate. dome (Fig. 1E). Sixteen days: Total height of the colony was now ~600 μm, Five days: The ancestrular tube was about 100 μm long by and the entire colony had changed to a pale cream colour day five, growing at ~10° off the vertical. The protruded (Figs 1K, 2G). The tentacle crowns of the three tightly lophophore was observed for the first time, bearing nine packed zooids were now approximately equal in size, short tentacles, the same number as zooids from later and distal growth of the three zooids became divergent, ontogeny in Hornera cf. robusta (Figs 1F, 2E). Viewed in thus preventing lophophore interference. This divergence profile, the extending zooidal tube appeared thin-walled initiated formation of the branch crown. In between the relative to the short collar at the base of the tube, the divergent zooids three new septa were formed, creating a boundary of which was marked by a sharp discontinuity. polygonal space in the incipient axil. As these septa were By day five, the ring-like walls visible at 24 hours had porous, the newly formed space seemingly had the potential formed a conspicuous zooidal bud (Z2) on the domed roof to develop into an autozooid. By day 16, a series of small, of the protoecium, extending from the edge of the basal buttress-like kenozooidal supports had begun to grow at the disc to the base of the ancestrular tube (Fig. 1F). About base of Z1/Z2 zooids, connecting the proximal portion of one-fifth of the bud’s circumference was shared with the these zooids to the protoecium roof (Figs 2G, 3). wall of the ancestrular tube. The bud’s ‘footprint’ when mapped onto the surface of the protoecial dome was egg- Eighteen days: No new growth was observed. The colony shaped, but viewed from above, the developing bud was appeared moribund, with loss of membrane integrity close to circular and matched the full zooidal diameter of and low lophophore activity. This corresponded with later zooids (Fig. 1F), suggesting morphogenetic control an unanticipated spike in temperature to >20°C during a of epi-protoecial budding differs from the septate budding heatwave. 26 AP Memoirs 52 (2019) AP Memoirs 52 (2019) 27

Figure 2. Early development in laboratory-reared Hornera cf. robusta: A, oblique silhouette showing relative size of free-swimming larva (cilia not shown); B, metamorph with everted adhesive sac (based on a specimen metamorphosing while unattached to substratum); C, early uncalcified protoecium;D , ~two day old interior-walled calcified protoecium with incipient zooid Z2 visible as curved ridges; E, ~five-day old ancestrula with functioning lophophore and epi-protoecial buds of zooids Z2 and Z3;F , ~10-day old colony, with sac- like daughter zooids and two functioning lophophores; G, at ~16 days, view rotated 180° from previous: colony has three functioning lophophores, growing tips of Z2/Z3 are level with the ancestrular zooid, and development of the basal branch crown has begun via zooid divergence and septum formation. Incipient kenozooids and other secondary thickening are visible at the colony base. All images shown to the same scale.

Comparison of cultured and wild-caught ancestrulae and kenozooid growth. This variability is perhaps due Nielsen (1970) reported cases of suspected abnormal to a combination of species-specific development and growth in cyclostome ancestrulae raised in the laboratory. environmental factors. To gauge whether growth in the laboratory-raised colony To reduce the risk of comparing early stages of two had developed normally, we compared it with wild-caught different species we selected a wild-grown 2–3 week-old specimens. Much variation in size and form exists among early colony growing on the brood chamber of a dredged wild-caught Hornera ancestrulae and early colonies, Hornera cf. robusta colony — inferred to be its progeny complicated by the fact that at least three Hornera species — for a more direct comparison (Fig. 3). In structure and occur on the Otago shelf. Among the variability encountered development the two young colonies were similar (although in wild-caught early colonies were: (1) diameter of the the distal portion of Z2 and all the peristomial denticles of protoecium; (2) number of autozooids budding directly the lab-cultured colony were accidentally broken off during from the protoecial roof (ranging from two to six); (3) the preparation for SEM). The wild-grown specimen was larger relationship between ancestrular tube and surrounding peri- in diameter relative to its height (Fig. 3). The diameter of ancestrular autozooids during early development (ranging the laboratory-cultured protoecium was 396 μm, while from a shared wall, to fully independent zooidal tubes with the wild-caught ancestrula was 470 μm. These respective a septate connection); (4) patterns of secondary calcification sizes do not reflect different developmental stages because

Figure 1 (opposite). Micrographs of early astogeny in living Hornera cf. robusta: A, larva; B, early metamorphosis (unattached); C, uncalcified protoecium; D, ~one day protoecium with incipient buds of second zooid arrowed; E, ~two day protoecium; F, five day ancestrula with feeding lophophore, circular Z2 bud and the beginning of the Z3 bud (arrowed); G, H, seven day ancestrula with developing sac-like periancestrular zooids; I, at ten days the distal extension of daughter zooids is approaching tip of ancestrular tube; J, at 14 days three zooids are feeding; K, colony at 16 days, with incipient branch crown formation via zooid divergence. (Scale bar =100 μm; all images are Hornera cf. robusta except 1C, which is H. robusta sensu strictu, from the Otago shelf; images 1D–1K are of the same individual.) 28 AP Memoirs 52 (2019)

margin in Disporella hispida shown in Jenkins & Taylor 2014, fig. 4). Consequently, the entire protoecium roof in H. cf. robusta is functionally interior-walled and available to support epi-protoecial budding of zooids. Evidence for this interpretation includes spinose pustules developed at the disc edge in the laboratory-grown early colony, and the radiating vertical buttresses (incipient interzooidal walls between kenozooids) emerging from the protoecium roof and extending to within 5–10 μm of the disc perimeter. Note that the roof of the hornerid protoecium is not fully calcified and the ancestrular tube is contiguous with the protoecium.

DISCUSSION Recent investigations of cyclostome ancestrulae and early astogenetic stages have revealed new skeletal characters within the (Jenkins & Taylor 2014, 2017; Figure 3. Comparison of cultured (left) and wild-caught (right) Taylor et al. 2015), adding to those described by Weedon ancestrulae of Otago shelf Hornera cf. robusta. Note that the distal (1998), Taylor & Weedon (2000) and others. Such studies portion of one autozooid was broken-off during SEM preparation highlight the potential of early developmental traits to of the cultured specimen. A shred of material, possibly cuticle, be informative in phylogenetic studies of this character- obscures part of the distal zooids on the right-hand ancestrula. poor group, especially when used in conjunction with molecular data from living clades (e.g. Waeschenbach et al. 2009; Jenkins & Taylor 2014; Taylor et al. 2015). This the protoecium remains fixed in diameter from its first emphasizes the need for further surveys of early astogeny calcification. Ancestrular tube diameter was greater than within and across cyclostome clades. the diameters of later zooids in both colonies (146 μm lab- This contribution describes the early astogeny of reared; 185 μm wild-grown). Hornera cf. robusta, supporting the findings made by Both the laboratory-raised and wild-caught colonies had Borg (1926). His observations on the close relationship the beginnings of kenozooidal supports (evident as low between the protoecium and early zooids in other species buttresses) emerging from the base of the zooidal tubes. The of Hornera (Borg 1926, p. 305, fig. 49) can be explained by laboratory-reared colony had a more pustulose protoecium, the epi-protoecial budding documented in this study. It is with pustules becoming almost spinose towards the edge unclear whether this pattern of astogeny is universal within of the protoecium. However, the laboratory-raised colony extant Horneridae; if it is, Borg’s reinterpretation of early had less conspicuous transverse ridging on the ancestrular astogeny in a young Hornera zoarium via septate budding tube. Another difference was the pronounced band of (Borg 1944, p. 176, fig. 23) may be misleading. secondary thickening at the junction of the protoecium and Our results provide an interesting contrast to the the base of the ancestrular tube in the cultured specimen general mode of early skeleton formation in cyclostomes. — this was absent from the wild-caught specimen. The Development of interior walls in the early astogeny of relative distribution of ultrastructures and mural pores was qualitatively similar between the two colonies. Overall, it appears that development of the laboratory- reared H. cf. robusta colony proceeded normally in terms of expected colony architecture and skeletal microstructure. One possible artifact of the rearing conditions was the reduced intensity of transverse growth increments, which are usually expressed as ‘pustule rings’ in developing zooids. This difference may be due to the absence of daily light-dark cycles, or the absence of tidal currents in the laboratory.

Interior-walled protoecium The protoecium roof of H. cf. robusta is entirely interior- walled. This agrees with observations made previously by Jenkins & Taylor (2017). The radiating ridges across the dorsal surfaces, the ongoing growth of pustules, and the presence of scattered mural pores, are all consistent with this interpretation (Fig. 4). Loose sheets of detached cuticle, most likely derived from the outer, uncalcified body wall wall were observed in SEMs of early (~24 hr) Figure 4. SEM of the disc edge of a cultured Hornera sp. Hornera protoecia. protoecium at about the same developmental stage as the The transition between interior and exterior walls on protoecium shown in Fig. 1D. Presence of an interior wall is the protoecium is probably located at the very edge of evident from the ridging, pustules and fragments of the outer the protoecial disc (cf. the position well inwards of the membranous wall. AP Memoirs 52 (2019) 29 extant cyclostomes typically occurs either by septum mechanism by which this state was achieved? And by formation at the extending tip of the ancestrular tube or by what means are the epi-protoecial autozooids budded in reflection of the growing wall back upon itself to form a Hornera: was it similar, for example, to that inferred by compound wall over the protoecium (rectangulates) (Borg Tavener-Smith (1969) for ? 1926). Until Jenkins & Taylor (2014) reported the partially The trials performed in this study show that it is possible interior-walled protoecium of Disporella hispida, it was to settle and raise hornerid larvae in the laboratory. widely thought that the newly formed protoecia of all extant Future studies could employ these methods, combined Cyclostomata were exterior-walled (e.g. McKinney 1978, with histological and improved imaging techniques, to p. 90; McKinney et al. 1993, p. 352; Weedon 1998). The examine how the hornerid interior wall is formed, how fully interior-walled hornerid protoecium and the direct budding occurs, and what morphogenetic and life-history budding of daughter zooids onto its calcified roof adds consequences arise from them. Such studies could aid another variation to known astogenetic pathways within understanding of morphogenetic patterns in some Paleozoic the Cyclostomata. clades, as well as in the wider Cancellata. The research reported here is preliminary, and two potential sources of error must be highlighted. First, the timing and order of developmental milestones described CONCLUSIONS above are based on a single colony grown in a laboratory 1. Patterns of larval metamorphosis and protoecium setting. Second, the protoecium was inferred to be ~24– formation in Hornera are basically similar to those reported 36 hours old when first examined, based on documented for other cyclostome suborders. development rates for other cyclostomes (Nielsen 1970). 2. Developmental rate, measured from calcification of the However, early development of the hornerid protoecium protoecium until first tentacle protrusion, was in line with (i.e. prior to ancestrular tube extension) may take relatively that reported for other taxa. longer than other cyclostomes because an entirely interior- walled roof is formed during this period. This possibility 3. Unlike other cyclostomes, however, the large protoecia requires further investigation. of Hornera have an entirely interior-walled roof, which develops during early calcification. Early astogeny in Hornera: analogy with Paleozoic 4. Direct epi-protecial budding of secondary zooids occurs stenolaemates on the surface of the protoecium. This development pattern Outwardly, striking similarities exist in the early astogeny parallels that of some Paleozoic stenolaemates, such as the of Hornera and Paleozoic Fenestrata. A number of fenestrates Lyroporella and Adlatipora. families, such as the fenestellids and acanthocladiids, 5. The number of autozooids budded on the protoecium share the hornerid traits of an interior-walled ancestula, a roof is variable in Hornera, ranging from two to six. This vertical ancestrular tube, and peri-ancestrular budding of variability may influence the number of primary branches primary zooids directly on the surface of the protoecium comprising the basal branch crown. (e.g. Tavener-Smith 1969; McKinney 1978; Gautier et al. 2013). It is notable that the broader colony bauplan in 6. Borg’s (1926) description of early hornerid development these taxa is also commonly similar to Hornera, including appears more compatible with our findings than his revised the presence of a stout, secondarily calcified primary stem schematic (Borg 1944), in which earliest zooidal budding leading to a multi-branched crown, from which unilaminate occurred by septum formation. branches typically arise (see McKinney et al. 1993 for a discussion of homeomorphy among reticulate forms in 7. Cancellate larvae from a mid-continental shelf setting can these groups). be settled and ongrown in the laboratory but improvements Ancestrulae, including the protoecium, of Paleozoic in cultivation methods are needed. stenolaemates are primitively exterior-walled (i.e. fixed- walled), as is evident in the basal stenolaemate Corynotrypa ACKNOWLEDGEMENTS (Taylor & Wilson 1994). Additional steps are required for We thank the Master and Crew of RV Polaris II (University the colony to become free-walled. In some trepostomes, of Otago), Dennis Gordon (NIWA), and Doug Mackie, this entailed ‘doubling back’ and self-overgrowth of Reuben Pooley, and Linda Groenewegan of the Portobello the exterior wall after septate multizooidal budding had Marine Laboratory. Special thanks to Kim Currie commenced (as described for Rhombotrypa by Boardman (Department of Chemistry, OU) for sharing vessel time, and & McKinney 1976, text-fig. 2). In fenestrate stenolaemates Elizabeth Girvan (Otago Centre for Electron Microscopy) a similar process occurred, but much earlier in development for her patience and efficiency. We also thank Helen Jenkins (Tavener-Smith 1969; Gautier et al. 2013). For example, for advice on culturing bryozoans, and reviewers Andrew in the acanthocladiid, Adlatipora, transition to an interior- Ostrovsky and Juan Cancino, whose comments greatly walled state via deflection was completed prior to the improved this manuscript. extension of the ancestrular tube (Gautier et al. 2013, fig. 5). In Hornera, the process of protoecial ‘doubling back’ REFERENCES to form an interior wall either does not occur, or does Boardman, R.S. & McKinney, F.K., 1976. Skeletal architecture so early in protoecium development (i.e. prior to tube and preserved organs of four-sided zooids in convergent extension). The fully formed, interior-walled, protoecial genera of Paleozoic Trepostomata (Bryozoa). Journal of roof appeared under light microscopy to be present from Paleontology 50, 25–78. the time calcification was first visible. Was this interior Borg, F., 1926. 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