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1. Assessing Soft-Tissue Shrinkage Estimates in Museum Specimens Imaged With Diffusible Iodine-Based Contrast-Enhanced Computed Tomography (diceCT)

   https://doi.org/10.1017/S1431927618000399  

   Hedrick, Brandon P.; Yohe, Laurel; Vander Linden, Abby; Dávalos, Liliana M.; Sears, Karen; Sadier, Alexa; Rossiter, Stephen J.; Davies, Kalina T.; Dumont, Elizabeth (June 2018, Microscopy and Microanalysis)

   null (Ed.)

   Abstract The increased accessibility of soft-tissue data through diffusible iodine-based contrast-enhanced computed tomography (diceCT) enables comparative biologists to increase the taxonomic breadth of their studies with museum specimens. However, it is still unclear how soft-tissue measurements from preserved specimens reflect values from freshly collected specimens and whether diceCT preparation may affect these measurements. Here, we document and evaluate the accuracy of diceCT in museum specimens based on the soft-tissue reconstructions of brains and eyes of five bats. Based on proxies, both brains and eyes were roughly 60% of the estimated original sizes when first imaged. However, these structures did not further shrink significantly over a 4-week staining interval, and 1 week in 2.5% iodine-based solution yielded sufficient contrast for differentiating among soft-tissues. Compared to six “fresh” bat specimens imaged shortly after field collection (not fixed in ethanol), the museum specimens had significantly lower relative volumes of the eyes and brains. Variation in field preparation techniques and conditions, and long-term storage in ethanol may be the primary causes of shrinkage in museum specimens rather than diceCT staining methodology. Identifying reliable tissue-specific correction factors to adjust for the shrinkage now documented in museum specimens requires future work with larger samples. 

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2. A 3D ontogenetic atlas ofAlligator mississippiensiscranial nerves and their significance for comparative neurology of reptiles

   https://doi.org/10.1002/ar.24550  

   Lessner, Emily_J; Holliday, Casey_M (November 2020, The Anatomical Record)

   Abstract Cranial nerves are key features of the nervous system and vertebrate body plan. However, little is known about the anatomical relationships and ontogeny of cranial nerves in crocodylians and other reptiles, hampering understanding of adaptations, evolution, and development of special senses, somatosensation, and motor control of cranial organs. Here we share three dimensional (3D) models an of the cranial nerves and cranial nerve targets of embryonic, juvenile, and adult American Alligators (Alligator mississippiensis) derived from iodine‐contrast CT imaging, for the first time, exploring anatomical patterns of cranial nerves across ontogeny. These data reveal the tradeoffs of using contrast‐enhanced CT data as well as patterns in growth and development of the alligator cranial nervous system. Though contrast‐enhanced CT scanning allows for reconstruction of numerous tissue types in a nondestructive manner, it is still limited by size and resolution. The position of alligator cranial nerves varies little with respect to other cranial structures yet grow at different rates as the skull elongates. These data constrain timing of trigeminal and sympathetic ganglion fusion and reveal morphometric differences in nerve size and path during growth. As demonstrated by these data, alligator cranial nerve morphology is useful in understanding patterns of neurological diversity and distribution, evolution of sensory and muscular innervation, and developmental homology of cranial regions, which in turn, lead to inferences of physiology and behavior. 

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3. Unveiling the elusive: X‐rays bring scolecophidian snakes out of the dark

   https://doi.org/10.1002/ar.24729  

   Bell, Christopher J.; Daza, Juan D.; Stanley, Edward L.; Laver, Rebecca J. (October 2021, The Anatomical Record)

   Abstract Scolecophidian snakes have long posed challenges for scholars interested in elucidating their anatomy. The importance, and relative paucity, of high‐quality anatomical data pertaining to scolecophidians was brought into sharp focus in the late 20 th century as part of a controversy over the phylogeny and ecological origin of snakes. The basal position of scolecophidians in the phylogeny of snakes makes their anatomy, behavior, ecology, and evolution especially important for such considerations. The depauperate fossil record for the group meant that advances in understanding their evolutionary history were necessarily tied to biogeographic distributions and anatomical interpretations of extant taxa. Osteological data, especially data pertaining to the skull and mandible, assumed a dominant role in shaping historical and modern perspectives of the evolution of scolecophidians. Traditional approaches to the exploration of the anatomy of these snakes relied heavily upon serial‐sectioned specimens and cleared‐and‐stained specimens. The application of X‐ray computed tomography (CT) to the study of scolecophidians revolutionized our understanding of the osteology of the group, and now, via diffusible iodine‐based contrast‐enhanced computed tomography (diceCT), is yielding data sets on internal soft anatomical features as well. CT data sets replicate many aspects of traditional anatomical preparations, are readily shared with a global community of scholars, and now are available for unique holotype and other rare specimens. The increasing prevalence and relevance of CT data sets is a strong incentive for the establishment and maintenance of permanent repositories for digital data. 

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4. A comparison of diceCT and histology for determination of nasal epithelial type

   https://doi.org/10.7717/peerj.12261  

   Smith, Timothy D.; Corbin, Hayley M.; King, Scot E.; Bhatnagar, Kunwar P.; DeLeon, Valerie B. (January 2021, PeerJ)

   Diffusible iodine-based contrast-enhanced computed tomography (diceCT) has emerged as a viable tool for discriminating soft tissues in serial CT slices, which can then be used for three-dimensional analysis. This technique has some potential to supplant histology as a tool for identification of body tissues. Here, we studied the head of an adult fruit bat ( Cynopterus sphinx ) and a late fetal vampire bat ( Desmodus rotundus ) using diceCT and µCT. Subsequently, we decalcified, serially sectioned and stained the same heads. The two CT volumes were rotated so that the sectional plane of the slice series closely matched that of histological sections, yielding the ideal opportunity to relate CT observations to corresponding histology. Olfactory epithelium is typically thicker, on average, than respiratory epithelium in both bats. Thus, one investigator (SK), blind to the histological sections, examined the diceCT slice series for both bats and annotated changes in thickness of epithelium on the first ethmoturbinal (ET I), the roof of the nasal fossa, and the nasal septum. A second trial was conducted with an added criterion: radioopacity of the lamina propria as an indicator of Bowman’s glands. Then, a second investigator (TS) annotated images of matching histological sections based on microscopic observation of epithelial type, and transferred these annotations to matching CT slices. Measurements of slices annotated according to changes in epithelial thickness alone closely track measurements of slices based on histologically-informed annotations; matching histological sections confirm blind annotations were effective based on epithelial thickness alone, except for a patch of unusually thick non-OE, mistaken for OE in one of the specimens. When characteristics of the lamina propria were added in the second trial, the blind annotations excluded the thick non-OE. Moreover, in the fetal bat the use of evidence for Bowman’s glands improved detection of olfactory mucosa, perhaps because the epithelium itself was thin enough at its margins to escape detection. We conclude that diceCT can by itself be highly effective in identifying distribution of OE, especially where observations are confirmed by histology from at least one specimen of the species. Our findings also establish that iodine staining, followed by stain removal, does not interfere with subsequent histological staining of the same specimen. 

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5. 3D Digitization in Functional Morphology: Where is the Point of Diminishing Returns?

   https://doi.org/10.1093/icb/icz101  

   Santana, Sharlene E; Arbour, Jessica H; Curtis, Abigail A; Stanchak, Kathryn E (June 2019, Integrative and Comparative Biology)

   Abstract Modern computational and imaging methods are revolutionizing the fields of comparative morphology, biomechanics, and ecomorphology. In particular, imaging tools such as X-ray micro computed tomography (µCT) and diffusible iodine-based contrast enhanced CT allow observing and measuring small and/or otherwise inaccessible anatomical structures, and creating highly accurate three-dimensional (3D) renditions that can be used in biomechanical modeling and tests of functional or evolutionary hypotheses. But, do the larger datasets generated through 3D digitization always confer greater power to uncover functional or evolutionary patterns, when compared with more traditional methodologies? And, if so, why? Here, we contrast the advantages and challenges of using data generated via (3D) CT methods versus more traditional (2D) approaches in the study of skull macroevolution and feeding functional morphology in bats. First, we test for the effect of dimensionality and landmark number on inferences of adaptive shifts during cranial evolution by contrasting results from 3D versus 2D geometric morphometric datasets of bat crania. We find sharp differences between results generated from the 3D versus some of the 2D datasets (xy, yz, ventral, and frontal), which appear to be primarily driven by the loss of critical dimensions of morphological variation rather than number of landmarks. Second, we examine differences in accuracy and precision among 2D and 3D predictive models of bite force by comparing three skull lever models that differ in the sources of skull and muscle anatomical data. We find that a 3D model that relies on skull µCT scans and muscle data partly derived from diceCT is slightly more accurate than models based on skull photographs or skull µCT and muscle data fully derived from dissections. However, the benefit of using the diceCT-informed model is modest given the effort it currently takes to virtually dissect muscles from CT scans. By contrasting traditional and modern tools, we illustrate when and why 3D datasets may be preferable over 2D data, and vice versa, and how different methodologies can complement each other in comparative analyses of morphological function and evolution. 

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