Complex enameloid microstructure of †Ischyrhiza mira rostral denticles.

Serving in a foraging or self-defense capacity, pristiophorids, pristids, and the extinct sclerorhynchoids independently evolved an elongated rostrum lined with modified dermal denticles called rostral denticles. Isolated rostral denticles of the sclerorhynchoid Ischyrhiza mira are commonly recovered from Late Cretaceous North American marine deposits. Although the external morphology has been thoroughly presented in the literature, very little is known about the histological composition and organization of these curious structures. Using acid-etching techniques and scanning electron microscopy, we show that the microstructure of I. mira rostral denticles are considerably more complex than that of previously described dermal denticles situated elsewhere on the body. The apical cap consists of outer single crystallite enameloid (SCE) and inner bundled crystallite enameloid (BCE) overlying a region of orthodentine. The BCE has distinct parallel bundled enameloid (PBE), tangled bundled enameloid (TBE), and radial bundled enameloid (RBE) components. Additionally, the cutting edge of the rostral denticle is produced by a superficial layer of SCE and a deeper ridges/cutting edge layer (RCEL) of the BCE. The highly organized enameloid observed in the rostral denticles of this batomorph resembles that of the multifaceted tissue architecture observed in the oral teeth of selachimorphs and demonstrates that dermal scales have the capacity to evolve histologically similar complex tooth-like structures both inside and outside the oropharyngeal cavity.

Exquisite air sac histological traces in a hyperpneumatized nanoid sauropod dinosaur from South America.

This study reports the occurrence of pneumosteum (osteohistological structure related to an avian-like air sac system) in a nanoid (5.7-m-long) saltasaurid titanosaur from Upper Cretaceous Brazil. We corroborate the hypothesis of the presence of an air sac system in titanosaurians based upon vertebral features identified through external observation and computed tomography. This is the fifth non-avian dinosaur taxon in which histological traces of air sacs have been found. We provided a detailed description of pneumatic structures from external osteology and CT scan data as a parameter for comparison with other taxa. The camellate pattern found in the vertebral centrum (ce) of this taxon and other titanosaurs shows distinct architectures. This might indicate whether cervical or lung diverticula pneumatized different elements. A cotylar internal plate of bone tissue sustains radial camellae (rad) in a condition similar to Alamosaurus and Saltasaurus. Moreover, circumferential chambers (cc) near the cotyle might be an example of convergence between diplodocoids and titanosaurs. Finally, we also register for the first time pneumatic foramina (fo) and fossae connecting camellate structures inside the neural canal in Titanosauria and the second published case in non-avian dinosaurs. The extreme pneumaticity observed in this nanoid titanosaur contrasts with previous assumptions that this feature correlates with the evolution of gigantic sizes in sauropodomorphs. This study reinforces that even small-bodied sauropod clades could present a hyperpneumatized postcranial skeleton, a character inherited from their large-bodied ancestors.

Preservation of collagen in the soft tissues of frozen mammoths.

We investigated the characteristics of extracellular matrix (ECM) in the soft tissue of two frozen baby woolly mammoths (Mammuthus primigenius) that died and were buried in Siberian permafrost approximately 40,000 years ago. Morphological and biochemical analyses of mammoth lung and liver demonstrated that those soft tissues were preserved at the gross anatomical and histological levels. The ultrastructure of ECM components, namely a fibrillar structure with a collagen-characteristic pattern of cross-striation, was clearly visible with transmission and scanning electron microscopy. Type I and type IV collagens were detected by immunohistochemical observation. Quantitative amino acid analysis of liver and lung tissues of the baby mammoths indicated that collagenous protein is selectively preserved in these tissues as a main protein. Type I and type III collagens were detected as major components by means of liquid chromatography-mass spectrometry analysis after digestion with trypsin. These results indicate that the triple helical collagen molecule, which is resistant to proteinase digestion, has been preserved in the soft tissues of these frozen mammoths for 40,000 years.

A potential cephalopod from the early Cambrian of eastern Newfoundland, Canada.

Although an early Cambrian origin of cephalopods has been suggested by molecular studies, no unequivocal fossil evidence has yet been presented. Septate shells collected from shallow-marine limestone of the lower Cambrian (upper Terreneuvian, c. 522 Ma) Bonavista Formation of southeastern Newfoundland, Canada, are here interpreted as straight, elongate conical cephalopod phragmocones. The material documented here may push the origin of cephalopods back in time by about 30 Ma to an unexpected early stage of the Cambrian biotic radiation of metazoans, i.e. before the first occurrence of euarthropods.

On the Planar Geometric Patterns of the Exoskeletal Relief Formation in Early Vertebrates (Osteostraci, Agnatha).

A study of the diversity of sculpture and histological structure of the exoskeleton in various osteostracan taxa (Osteostraci, Agnatha) enabled the first characterization of the main elements (geometric modules) of the planar organization of the complex relief on their armor surface. The armor relief was analyzed using a circular model of the formation of exoskeletal hard structures. The model was applied to unique material, fragments of a shield of the osteostracan Oeselaspis pustulata (Patten, 1931) from the Silurian of the Severnaya Zemlya Archipelago (Russia).

Fossil evidence for vampire squid inhabiting oxygen-depleted ocean zones since at least the Oligocene.

A marked 120 My gap in the fossil record of vampire squids separates the only extant species (Vampyroteuthis infernalis) from its Early Cretaceous, morphologically-similar ancestors. While the extant species possesses unique physiological adaptations to bathyal environments with low oxygen concentrations, Mesozoic vampyromorphs inhabited epicontinental shelves. However, the timing of their retreat towards bathyal and oxygen-depleted habitats is poorly documented. Here, we document a first record of a post-Mesozoic vampire squid from the Oligocene of the Central Paratethys represented by a vampyromorph gladius. We assign Necroteuthis hungarica to the family Vampyroteuthidae that links Mesozoic loligosepiids with Recent Vampyroteuthis. Micropalaeontological, palaeoecological, and geochemical analyses demonstrate that Necroteuthis hungarica inhabited bathyal environments with bottom-water anoxia and high primary productivity in salinity-stratified Central Paratethys basins. Vampire squids were thus adapted to bathyal, oxygen-depleted habitats at least since the Oligocene. We suggest that the Cretaceous and the early Cenozoic OMZs triggered their deep-sea specialization.

Application of mid-infrared free-electron laser for structural analysis of biological materials.

A mid-infrared free-electron laser (MIR-FEL) is a synchrotron-radiation-based femto- to pico-second pulse laser. It has unique characteristics such as variable wavelengths in the infrared region and an intense pulse energy. So far, MIR-FELs have been utilized to perform multi-photon absorption reactions against various gas molecules and protein aggregates in physical chemistry and biomedical fields. However, the applicability of MIR-FELs for the structural analysis of solid materials is not well recognized in the analytical field. In the current study, an MIR-FEL is applied for the first time to analyse the internal structure of biological materials by using fossilized inks from cephalopods as the model sample. Two kinds of fossilized inks that were collected from different strata were irradiated at the dry state by tuning the oscillation wavelengths of the MIR-FEL to the phosphoryl stretching mode of hydroxyapatite (9.6 µm) and to the carbonyl stretching mode of melanin (5.8 µm), and the subsequent structural changes in those materials were observed by using infrared microscopy and far-infrared spectroscopy. The structural variation of these biological fossils is discussed based on the infrared-absorption spectral changes that were enhanced by the MIR-FEL irradiation, and the potential use of MIR-FELs for the structural evaluation of biomaterials is suggested.

Using tooth enamel microstructure to identify mammalian fossils at an Eocene Arctic forest.

Lower Eocene (Wasatchian-aged) sediments of the Margaret Formation on Ellesmere Island in Canada's High Arctic preserve evidence of a rainforest inhabited by alligators, turtles, and a diverse mammalian fauna. The mammalian fossils are fragmentary and often poorly preserved. Here, we offer an alternative method for their identification. Among the best preserved and extensive of the Eocene Arctic forests is the Strathcona Fiord Fossil Forest, which contains permineralized in situ tree stumps protruding from a prominent coal seam, but a paucity of vertebrate fossils. In 2010 and 2018, we recovered mammalian tooth fragments at the fossil forest, but they are so incomplete as to be undiagnostic by using their external morphology. We used a combination of light microscopy and SEM analysis to study the enamel microstructure of two tooth fragments from the fossil forest-NUFV2092B and 2092E. The results of our analysis indicate that NUFV2092B and 2092E have Coryphodon-enamel, which is characterized by vertical bodies that manifest as bands of nested chevrons or treelike structures visible in the tangential section under light microscopy. This enamel type is not found in other mammals known from the Arctic. Additionally, when studied under SEM, the enamel of NUFV2092B and 2092E has rounded prisms that open to one side and are surrounded by interprismatic matrix that is nearly parallel to the prisms, which also occurs in Coryphodon enamel, based on prior studies. The tooth fragments reported here, along with some poorly preserved bone fragments, thus far are the only documented vertebrate fossils from the Strathcona Fiord Fossil Forest. However, fossils of Coryphodon occur elsewhere in the Margaret Formation, so its presence at the fossil forest is not surprising. What is novel in our study is the way in which we identified the fossils using their enamel microstructure.

Genomic and fossil windows into the secret lives of the most ancient fungi.

Fungi have crucial roles in modern ecosystems as decomposers and pathogens, and they engage in various mutualistic associations with other organisms, especially plants. They have a lengthy geological history, and there is an emerging understanding of their impact on the evolution of Earth systems on a large scale. In this Review, we focus on the roles of fungi in the establishment and early evolution of land and freshwater ecosystems. Today, questions of evolution over deep time are informed by discoveries of new fossils and evolutionary analysis of new genomes. Inferences can be drawn from evolutionary analysis by comparing the genes and genomes of fungi with the biochemistry and development of their plant and algal hosts. We then contrast this emerging picture against evidence from the fossil record to develop a new, integrated perspective on the origin and early evolution of fungi.

The cono-dos and cono-dont's of phosphatic microfossil preparation and microanalysis.

Scanning electron microscope (SEM) imaging of fossils allows unlocking ultrastructural information about their skeletal tissues, but sample preparation of biominerals forming their skeletons requires time, patience, and knowledge. SEM and associated analytical methods allow the observation of internal microstructure, shedding light on function, growth and chemistry. Sample preparation is the process by which material is fixed within a medium (e.g. epoxy resin), a transect created and surface defects removed. This step is arguably the most important in any SEM-based analysis, allowing for the acquisition of reliable, high quality data sets. When conducting any SEM-based technique, the presence of a flat surface is needed to collect consistent and reliable data. Surfaces with topography will both induce charging effects but will also compromise the reliability of data acquired. Techniques from material science are continuously adapted to palaeontological applications, in particular with respect to calcareous microfossils. However, similar studies have not been extensively conducted on bioapatite, owing in part to the difficulties faced in sample preparation alongside its susceptibility to electron beam damage. This case study focuses on conodonts, a marine vertebrate group ranging from the late Cambrian to the Late Triassic. They have been chosen as a model due to the abundance of material, complexity of internal tissues and previous work focused on histological features. With these phosphatic microfossils, we attempt to outline the process of sample preparation and provide information on how to avoid and overcome common pitfalls.

Nanoscale trace metal imprinting of biocalcification of planktic foraminifers by Toba's super-eruption.

Bioactive metal releases in ocean surface water, such as those by ash falls during volcanic super-eruptions, might have a potentially toxic impact on biocalcifier planktic microorganisms. Nano-XRF imaging with the cutting-edge synchrotron hard X-ray nano-analysis ID16B beamline (ESRF) revealed for the first time a specific Zn- and Mn-rich banding pattern in the test walls of Globorotalia menardii planktic foraminifers extracted from the Young Toba Tuff layer, and thus contemporaneous with Toba's super-eruption, 74,000 years ago. The intra-test correlation of Zn and Mn patterns at the nanoscale with the layered calcareous microarchitecture, indicates that the incorporation of these metals is syngenetic to the wall growth. The preferential Mn and Zn sequestration within the incipient stages of chamber formation suggests a selective incorporation mechanism providing a resilience strategy to metal pollution in the test building of planktic foraminifers.

A Tube-Dwelling Early Cambrian Lobopodian.

Facivermis yunnanicus [1, 2] is an enigmatic worm-like animal from the early Cambrian Chengjiang Biota of Yunnan Province, China. It is a small (<10 cm) bilaterian with five pairs of spiny anterior arms, an elongated body, and a swollen posterior end. The unusual morphology of Facivermis has prompted a history of diverse taxonomic interpretations, including among annelids [1, 3], lophophorates [4], and pentastomids [5]. However, in other studies, Facivermis is considered to be more similar to lobopodians [2, 6-8]-the fossil grade from which modern panarthropods (arthropods, onychophorans, and tardigrades) are derived. In these studies, Facivermis is thought to be intermediate between cycloneuralian worms and lobopodians. Facivermis has therefore been suggested to represent an early endobenthic-epibenthic panarthropod transition [6] and to provide crucial insights into the origin of paired appendages [2]. However, the systematic affinity of Facivermis was poorly supported in a previous phylogeny [6], partially due to incomplete understanding of its morphology. Therefore, the evolutionary significance of Facivermis remains unresolved. In this study, we re-examine Facivermis from new material and the holotype, leading to the discovery of several new morphological features, such as paired eyes on the head and a dwelling tube. Comprehensive phylogenetic analyses using parsimony, Bayesian inference, and maximum likelihood all support Facivermis as a luolishaniid in a derived position within the onychophoran stem group rather than as a basal panarthropod. In contrast to previous studies, we therefore conclude that Facivermis provides a rare early Cambrian example of secondary loss to accommodate a highly specialized tube-dwelling lifestyle.

Growing up Tyrannosaurus rex: Osteohistology refutes the pygmy "Nanotyrannus" and supports ontogenetic niche partitioning in juvenile Tyrannosaurus.

Despite its iconic status as the king of dinosaurs, Tyrannosaurus rex biology is incompletely understood. Here, we examine femur and tibia bone microstructure from two half-grown T. rex specimens, permitting the assessments of age, growth rate, and maturity necessary for investigating the early life history of this giant theropod. Osteohistology reveals these were immature individuals 13 to 15 years of age, exhibiting growth rates similar to extant birds and mammals, and that annual growth was dependent on resource abundance. Together, our results support the synonomization of "Nanotyrannus" into Tyrannosaurus and fail to support the hypothesized presence of a sympatric tyrannosaurid species of markedly smaller adult body size. Our independent data contribute to mounting evidence for a rapid shift in body size associated with ontogenetic niche partitioning late in T. rex ontogeny and suggest that this species singularly exploited mid- to large-sized theropod niches at the end of the Cretaceous.

Earth's earliest and deepest purported fossils may be iron-mineralized chemical gardens.

Recognizing fossil microorganisms is essential to the study of life's origin and evolution and to the ongoing search for life on Mars. Purported fossil microbes in ancient rocks include common assemblages of iron-mineral filaments and tubes. Recently, such assemblages have been interpreted to represent Earth's oldest body fossils, Earth's oldest fossil fungi, and Earth's best analogues for fossils that might form in the basaltic Martian subsurface. Many of these putative fossils exhibit hollow circular cross-sections, lifelike (non-crystallographic, constant-thickness, and bifurcate) branching, anastomosis, nestedness within 'sheaths', and other features interpreted as strong evidence for a biological origin, since no abiotic process consistent with the composition of the filaments has been shown to produce these specific lifelike features either in nature or in the laboratory. Here, I show experimentally that abiotic chemical gardening can mimic such purported fossils in both morphology and composition. In particular, chemical gardens meet morphological criteria previously proposed to establish biogenicity, while also producing the precursors to the iron minerals most commonly constitutive of filaments in the rock record. Chemical gardening is likely to occur in nature. Such microstructures should therefore not be assumed to represent fossil microbes without independent corroborating evidence.

Chemical characterization of pterosaur melanin challenges color inferences in extinct animals.

Melanosomes (melanin-bearing organelles) are common in the fossil record occurring as dense packs of globular microbodies. The organic component comprising the melanosome, melanin, is often preserved in fossils, allowing identification of the chemical nature of the constituent pigment. In present-day vertebrates, melanosome morphology correlates with their pigment content in selected melanin-containing structures, and this interdependency is employed in the color reconstruction of extinct animals. The lack of analyses integrating the morphology of fossil melanosomes with the chemical identification of pigments, however, makes these inferences tentative. Here, we chemically characterize the melanin content of the soft tissue headcrest of the pterosaur Tupandactylus imperator by alkaline hydrogen peroxide oxidation followed by high-performance liquid chromatography. Our results demonstrate the unequivocal presence of eumelanin in T. imperator headcrest. Scanning electron microscopy followed by statistical analyses, however, reveal that preserved melanosomes containing eumelanin are undistinguishable to pheomelanin-bearing organelles of extant vertebrates. Based on these new findings, straightforward color inferences based on melanosome morphology may not be valid for all fossil vertebrates, and color reconstructions based on ultrastructure alone should be regarded with caution.

Organically-preserved multicellular eukaryote from the early Ediacaran Nyborg Formation, Arctic Norway.

Eukaryotic multicellularity originated in the Mesoproterozoic Era and evolved multiple times since, yet early multicellular fossils are scarce until the terminal Neoproterozoic and often restricted to cases of exceptional preservation. Here we describe unusual organically-preserved fossils from mudrocks, that provide support for the presence of organisms with differentiated cells (potentially an epithelial layer) in the late Neoproterozoic. Cyathinema digermulense gen. et sp. nov. from the Nyborg Formation, Vestertana Group, Digermulen Peninsula in Arctic Norway, is a new carbonaceous organ-taxon which consists of stacked tubes with cup-shaped ends. It represents parts of a larger organism (multicellular eukaryote or a colony), likely with greater preservation potential than its other elements. Arrangement of open-ended tubes invites comparison with cells of an epithelial layer present in a variety of eukaryotic clades. This tissue may have benefitted the organism in: avoiding overgrowth, limiting fouling, reproduction, or water filtration. C. digermulense shares characteristics with extant and fossil groups including red algae and their fossils, demosponge larvae and putative sponge fossils, colonial protists, and nematophytes. Regardless of its precise affinity, C. digermulense was a complex and likely benthic marine eukaryote exhibiting cellular differentiation, and a rare occurrence of early multicellularity outside of Konservat-Lagerstätten.

Morphological variation suggests that chitinozoans may be fossils of individual microorganisms rather than metazoan eggs.

Chitinozoans are organic-walled microfossils widely recorded in Ordovician to Devonian (ca 485-359 Mya) marine sediments and extensively used in high-resolution biostratigraphy. Their biological affinity remains unknown, but most commonly, they are interpreted as eggs of marine metazoans. Here, we provide new insights into their palaeobiology from three lines of inquiry. We examine morphological variation of a new, well-preserved Late Ordovician species, Hercochitina violana; analyse a compiled dataset of measurements on 378 species representing all known chitinozoan genera; and compare these data with the size variation of eggs of both extinct and extant aquatic invertebrates. The results indicate that the magnitude of size variation within chitinozoan species is larger than observed in fossil and modern eggs. Additionally, delicate morphological structures of chitinozoans, such as prosome and complex ornamentation, are inconsistent with the egg hypothesis. Distinct and continuous morphological variation in H. violana is more plausibly interpreted as an ontogenetic series of individual microorganisms, rather than as eggs.

An Akania (Akaniaceae) inflorescence with associated pollen from the early Miocene of New Zealand.

PREMISE OF THE STUDY: An Akania-like inflorescence, including flowers with in situ pollen was recovered from the remarkable Konservat-Lagerstätte lacustrine diatomite deposit at Foulden Maar, Otago indicating the presence of Akaniaceae in southern New Zealand during the early Miocene. The flowers, although slightly smaller than the sole modern Australian species, A. bidwillii, contain pollen grains that are very like that taxon. The pollen also resembles that of the monospecific sister genus Bretschneidera from Southeast Asia and India, although that taxon has flowers with very different morphology from this genus. METHODS: The floral morphology of the fossil and in situ pollen grains were compared with flowers and pollen grains from extant species of Akaniaceae and related taxa. KEY RESULTS: The fossil inflorescence and associated pollen are referred to a new, extinct species of Akania: Akania gibsonorum. The floral structures and pollen resemble those of the modern Australian Akania species. CONCLUSIONS: The discovery of fossil flowers of Akania in an early Miocene lake deposit in New Zealand, coupled with earlier recognition of Akaniaceae leaves from the Paleocene epoch and wood from the Miocene epoch in South America suggests that the genus was once widespread in former Gondwana landmasses. The extinction of Akaniaceae in New Zealand and South America, and its present relictual distribution in eastern Australia, is most likely related to post-Miocene climatic cooling.

Combined microscopy and spectroscopy techniques to characterize a fossilized feather with minimal damage to the specimen.

The study of fossil feathers has been revitalized in the last few decades and has contributed significantly to paleontological studies of dinosaurs and birds. Specific morphological and physicochemical characteristics of the microscale structures of feathers and the protein keratin are key targets when preserved during the fossilization process. Keratin is a fibrous protein that composes some hard tissues such as hair, nails and feathers. It is part of the so called intermediate filaments inside keratinocyte cells and is rich in sulfur containing amino acid cysteine. To date, different microscopy and analytical methods have been used for the analysis and detailed characterization and classification of feathers. However, in this work we showed that analytical optical and electron microscopies can be quick and precise methods with minimal effects on the sample during analysis. This association of different approaches on the same sample results in correlative data albeit in different length scales. Intracellular bodies called melanosomes originally present in melanocyte cells were identified with Scanning Electron Microscopy (SEM) and Transmission Electron Microscopy (TEM), and had well-defined orientation and a mean aspect ratio comparable to melanosomes extant in dark feathers. The detection of sulphur in melanosomes via Energy Dispersive Spectroscopy both in SEM and TEM shows that, along the fossilization process, sulphur from the degraded keratin matrix could have been trapped inside the melanosomes. Chemical groups that make up keratin and melanin in the fossil sample were detected via FT-IR Spectroscopy and Confocal Laser Scanning Microscopy (CLSM). The use of combined analytical microscopy techniques can contribute significantly to the study of fossils generating precise results with minimum damage to the original sample.

Morphological characteristics of preparator air-scribe marks: Implications for taphonomic research.

Taphonomic analyses of bone-surface modifications can provide key insights into past biotic involvement with animal remains, as well as elucidate the context(s) of other biostratinomic (pre-burial) processes, diagenesis, excavation, preparation and storage. Such analyses, however, first require researchers to rigorously disambiguate between continuums of damage morphologies prior to attributing individual marks to specific actors and effectors (e.g., carnivore tooth, stone tool cutting edge, etc.). To date, a number of bone-modifying agents have been identified, and criteria for identifying their traces have been published. Relatively little research, however, has focused on bone-surface modifications imparted during specimen preparation. Herein we report that air scribes, small pneumatic tools commonly used for preparation in museum contexts, can generate unintentional marks that may mimic surficial modification caused by carnivores. To aid investigators in assessing the hypothesis that a mark in question is derived from air-scribe preparation activities, we provide high-resolution, detailed morphological information imaged with scanning electron microscopy (SEM). The main diagnostic characteristic of air-scribe damage is the occurrence of sequential, variously spaced, sub-millimeter scallop-like stepped bone removals. This morphology can resemble damage imparted by carnivore teeth. In contrast to marks produced by trampling, stone tools and carnivores, however, no continuous internal features, such as linear microstriations, were observed within grooves produced by the air scribe. Thus, the presence of such features can be used to disprove an air-scribe origin. A culmination of the morphological criteria presented herein, cross-cutting relationships with other surficial features (e.g., diagenetic discoloration, weathering textures), the position of occurrence, and an overall contextual framework for the assemblage is suggested for accurate identification of such traces. The ability to recognize or disprove air-scribe damage will allow researchers to confidently proceed with interpreting past biological and sedimentological interactions with animal remains.