DOI: 10.5061/dryad.0zpc866xv
title: Synchrotron imagery of phosphatized eggs in Waptia cf. W. fieldensis from the middle Cambrian (Miaolingian; Wuliuan) Spence Shale of Utah

description: Exceptionally preserved fossil eggs and embryos provide critical
 information regarding paleoembryogenesis, reproductive strategies, and the
 early ontogeny of early arthropods, but the rarity of preservation of both
 eggs and egg-bearing organisms in situ limits their use in detailed
 evo-devo studies. Burgess Shale-type deposits preserve rare instances of
 egg-bearing arthropods as carbonaceous compressions, however, the eggs are
 usually poorly preserved with no compelling evidence of embryos. We
 describe the first record of a brooding specimen of Waptia cf. fieldensis
 from the Spence Shale, a Cambrian (Wuliuan stage) Burgess Shale-type
 deposit in northeastern Utah and southeastern Idaho. This is the first
 record of an egg-bearing arthropod from the Spence Shale and it exhibits
 two distinct modes of preservation among eggs within the single clutch:
 carbonization and phosphatization. Unlike the egg-bearing Burgess Shale
 specimens, many eggs of this Utah specimen are also preserved
 three-dimensionally. In addition, synchrotron radiation X-ray tomographic
 microscopy reveals internal distributions of mineral phases, along with
 potential remnants of the egg membrane and attachment structures but, as
 in the Burgess Shale, no explicit traces of developing embryos. The
 distinct modes of preservation highlight the existence of diagenetic
 microenvironments within some eggs but not in others during fossilization.
description: The <i>Waptia </i>cf.
 W. <i>fieldensis</i> Walcott, 1912 specimen (KUMIP 314032)
 (Fig. 1.2–1.3, 2.4–2.7) was collected by the Gunther Family at Miners
 Hollow (Section 14, T10N, R02W; 41.6023°N, 112.0334°W), Wellsville
 Mountains, Langston Formation, Spence Shale, Utah, United States (Kimmig
 et al., 2019, p. 610, fig. 1). The specimen was photographed with a Canon
 EOS 5D Mark II (f/6.3, 1/100 sec, ISO 2500, 100 mm focal length) using
 different illumination techniques (Fig. 1). Dimensions of eggs were
 measured using Dragonfly version 2021.1 for Windows (Object Research
 Systems Inc., Montreal, Canada, 2021), where max length and diameter
 measurements were taken from the dorsal point of view to keep the
 orientation consistent (Table 1). Portions of eggs cropped out of the
 synchrotron images were estimated based on Figure 2.7 using FIJI ImageJ
 software for Windows (Schindelin et al., 2012). Synchrotron images were
 taken at beamline ID 19 of the European Synchrotron Radiation Facility,
 Grenoble, France, to investigate the ultrastructural details of the eggs.
 One of the eggs (referred to as Egg One) (Fig. 1.7, 3.2–3.4) became loose
 and was analyzed separately from the matrix. Egg One was imaged using 26.5
 keV monochromatic beam at 0.65 μm resolution. All other eggs were scanned
 using 110 keV monochromatic beam at 0.64–0.66 μm resolution. Synchrotron
 images were converted from .TIFF to grayscale .JPEG using the FIJI ImageJ
 software for Windows (Schindelin et al., 2012) batch convert tool to
 accommodate hardware limitations. The converted data were volume-rendered
 in Dragonfly version 2021.1 (Object Research Systems Inc., Montreal,
 Canada, 2021). All eggs were segmented by: (1) enforcing contrast
 thresholds to isolate the fossils from the matrix (Fig. 4.2–4.3); (2)
 creating regions of interest; (3) and applying false colors for
 segmentation (Fig. 4.4). Backscatter Electron image (BSE) and
 Energy-dispersive X-ray spectroscopy (EDS) was conducted on a single
 isolated egg (Egg One) using a FEI Quanta 200 FEG Environmental SEM, with
 an EDAX Octane Plus SDD x-ray detector in Low Vacuum mode with chamber
 pressure of 70 Pa to analyze the surface composition of Egg One at 80x
 magnification with a scanning energy of 12 kv at the University of
 Windsor, Toronto, Canada. The single egg had initially been glued using
 cyanoacrylate on a toothpick for analysis at the synchrotron. Some
 residues of cyanoacrylate could not be removed, explaining local
 concentrations of carbon.
description: Moon et al. Supp_1.mp4 is a
 video file of Synchrotron analysis of the Spence Shale specimen, showing
 the lateral cross-section of three-dimensionally preserved eggs. A
 composite image of these eggs is shown as Figure 4.1 Moon et al. Supp_2.mp4 is a
 rendered animation of Egg 1 using Dragonfly ORS software to illustrate the
 overall morphology of the egg. The surrounding matrix has been removed and
 different colors represent the different phases shown in the analysis.
 Yellow represents the low-density phase, and light blue represents the
 high-density phase.