Hadrosauroid eggs and embryos from the Upper Cretaceous (Maastrichtian) of Jiangxi Province, China.

BACKGROUND: Dinosaur eggs containing embryos are rare, limiting our understanding of dinosaur development. Recently, a clutch of subspherical dinosaur eggs was discovered while blasting for a construction project in the Upper Cretaceous red beds (Hekou Formation) of the Ganzhou Basin, Jiangxi Province, China. At least two of the eggs contain identifiable hadrosauroid embryos, described here for the first time. RESULTS: The eggs, attributable to Spheroolithidae indet., are thin-walled and small (~ 660 mL) compared to those of Lambeosaurinae. The shape of the embryonic squamosal is reminiscent of that seen in the Late Cretaceous hadrosauroids Levnesovia transoxiana, Tanius sinensis, and Nanningosaurus dashiensis, suggestive of possible affinities. CONCLUSION: The small size of the eggs and embryos, similar to those of Hadrosaurinae, indicates that the larger eggs and hatchlings typical of Lambeosaurinae are evolutionarily derived.

An exquisitely preserved in-ovo theropod dinosaur embryo sheds light on avian-like prehatching postures.

Despite the discovery of many dinosaur eggs and nests over the past 100 years, articulated in-ovo embryos are remarkably rare. Here we report an exceptionally preserved, articulated oviraptorid embryo inside an elongatoolithid egg, from the Late Cretaceous Hekou Formation of southern China. The head lies ventral to the body, with the feet on either side, and the back curled along the blunt pole of the egg, in a posture previously unrecognized in a non-avian dinosaur, but reminiscent of a late-stage modern bird embryo. Comparison to other late-stage oviraptorid embryos suggests that prehatch oviraptorids developed avian-like postures late in incubation, which in modern birds are related to coordinated embryonic movements associated with tucking - a behavior controlled by the central nervous system, critical for hatching success. We propose that such pre-hatching behavior, previously considered unique to birds, may have originated among non-avian theropods, which can be further investigated with additional discoveries of embryo fossils.

A possible brachiosaurid (Dinosauria, Sauropoda) from the mid-Cretaceous of northeastern China.

Brachiosauridae is a lineage of titanosauriform sauropods that includes some of the most iconic non-avian dinosaurs. Undisputed brachiosaurid fossils are known from the Late Jurassic through the Early Cretaceous of North America, Africa, and Europe, but proposed occurrences outside this range have proven controversial. Despite occasional suggestions that brachiosaurids dispersed into Asia, to date no fossils have provided convincing evidence for a pan-Laurasian distribution for the clade, and the failure to discover brachiosaurid fossils in the well-sampled sauropod-bearing horizons of the Early Cretaceous of Asia has been taken to evidence their genuine absence from the continent. Here we report on an isolated sauropod maxilla from the middle Cretaceous (Albian-Cenomanian) Longjing Formation of the Yanji basin of northeast China. Although the specimen preserves limited morphological information, it exhibits axially twisted dentition, a shared derived trait otherwise known only in brachiosaurids. Referral of the specimen to the Brachiosauridae receives support from phylogenetic analysis under both equal and implied weights parsimony, providing the most convincing evidence to date that brachiosaurids dispersed into Asia at some point in their evolutionary history. Inclusion in our phylogenetic analyses of an isolated sauropod dentary from the same site, for which an association with the maxilla is possible but uncertain, does not substantively alter these results. We consider several paleobiogeographic scenarios that could account for the occurrence of a middle Cretaceous Asian brachiosaurid, including dispersal from either North America or Europe during the Early Cretaceous. The identification of a brachiosaurid in the Longshan fauna, and the paleobiogeographic histories that could account for its presence there, are hypotheses that can be tested with continued study and excavation of fossils from the Longjing Formation.

First report of leopard fossils from a limestone cave in Kenting area, southern Taiwan.

Longshia-dong Cave, a limestone cave located in the Kenting area within the Kenting National Park of southern Taiwan, yields numerous terrestrial mammalian fossils. Many of them were not reported in historical literature and are neither present in Taiwan. For instance, no historical literature mentioned leopards inhabited in Taiwan, and thus their existence remained unknown. This study describes three fossil leopard (Panthera pardus) teeth uncovered from Longshia-dong Cave. Two isolated lower premolars and one lower molar, respectively p3, p4 and m1, were discovered in a very small area (11 × 6 cm) and show a series of progressive increase in size. Thus, the three teeth should have been belonging to the same individual from the subfamily of Pantherinae. Traditional linear measurements and two-dimensional geometric morphometric analysis for the occlusal surface outlines were conducted on the fossil teeth and extant pantherines inhabited in Asia such as clouded leopards (Neofelis nebulosa), leopards (Panthera pardus), and tigers (Panthera tigiris). Results show that the fossil teeth are similar both in size and morphology to the teeth of extant leopards, suggesting the assignment of the fossil teeth to leopards. This study, for the first time, reported the presence of leopards in the Late Pleistocene of Taiwan. In addition, the smaller size of the fossil teeth in comparison with Chinese fossil leopards is putatively attributed to insular dwarfism or individual size variability, yet more studies are required.

An oviraptorid preserved atop an embryo-bearing egg clutch sheds light on the reproductive biology of non-avialan theropod dinosaurs.

Recent studies demonstrate that many avialan features evolved incrementally prior to the origin of the group, but the presence of some of these features, such as bird-like brooding behaviours, remains contentious in non-avialan dinosaurs. Here we report the first non-avialan dinosaur fossil known to preserve an adult skeleton atop an egg clutch that contains embryonic remains. The preserved positional relationship of the adult to the clutch, coupled with the advanced growth stages of the embryos and their high estimated incubation temperatures, provides strong support for the brooding hypothesis. Furthermore, embryos in the clutch are at different developmental stages, suggesting the presence of asynchronous hatching-a derived feature even among crown-group birds-in non-avialan theropods. These findings demonstrate that the evolution of reproductive biology along bird-line archosaurs was a complex rather than a linear and incremental process, and suggest that some aspects of non-avialan theropod reproduction were unique to these dinosaurs.

Dinosaur egg colour had a single evolutionary origin.

Birds are the only living amniotes with coloured eggs1-4, which have long been considered to be an avian innovation1,3. A recent study has demonstrated the presence of both red-brown protoporphyrin IX and blue-green biliverdin5-the pigments responsible for all the variation in avian egg colour-in fossilized eggshell of a nonavian dinosaur6. This raises the fundamental question of whether modern birds inherited egg colour from their nonavian dinosaur ancestors, or whether egg colour evolved independently multiple times. Here we present a phylogenetic assessment of egg colour in nonavian dinosaurs. We applied high-resolution Raman microspectroscopy to eggshells that represent all of the major clades of dinosaurs, and found that egg colour pigments were preserved in all eumaniraptorans: egg colour had a single evolutionary origin in nonavian theropod dinosaurs. The absence of colour in ornithischian and sauropod eggs represents a true signal rather than a taphonomic artefact. Pigment surface maps revealed that nonavian eumaniraptoran eggs were spotted and speckled, and colour pattern diversity in these eggs approaches that in extant birds, which indicates that reproductive behaviours in nonavian dinosaurs were far more complex than previously known3. Depth profiles demonstrated identical mechanisms of pigment deposition in nonavian and avian dinosaur eggs. Birds were not the first amniotes to produce coloured eggs: as with many other characteristics7,8 this is an attribute that evolved deep within the dinosaur tree and long before the spectacular radiation of modern birds.

Fossilization transforms vertebrate hard tissue proteins into N-heterocyclic polymers.

Vertebrate hard tissues consist of mineral crystallites within a proteinaceous scaffold that normally degrades post-mortem. Here we show, however, that decalcification of Mesozoic hard tissues preserved in oxidative settings releases brownish stained extracellular matrix, cells, blood vessels, and nerve projections. Raman Microspectroscopy shows that these fossil soft tissues are a product of diagenetic transformation to Advanced Glycoxidation and Lipoxidation End Products, a class of N-heterocyclic polymers generated via oxidative crosslinking of proteinaceous scaffolds. Hard tissues in reducing environments, in contrast, lack soft tissue preservation. Comparison of fossil soft tissues with modern and experimentally matured samples reveals how proteinaceous tissues undergo diagenesis and explains biases in their preservation in the rock record. This provides a target, focused on oxidative depositional environments, for finding cellular-to-subcellular soft tissue morphology in fossils and validates its use in phylogenetic and other evolutionary studies.

Fossil eggshell cuticle elucidates dinosaur nesting ecology.

The cuticle layer consisting mainly of lipids and hydroxyapatite (HAp) atop the mineralized avian eggshell is a protective structure that prevents the egg from dehydration and microbial invasions. Previous ornithological studies have revealed that the cuticle layer is also involved in modulating the reflectance of eggshells in addition to pigments (protoporphyrin and biliverdin). Thus, the cuticle layer represents a crucial trait that delivers ecological signals. While present in most modern birds, direct evidence for cuticle preservation in stem birds and non-avian dinosaurs is yet missing. Here we present the first direct and chemical evidence for the preservation of the cuticle layer on dinosaur eggshells. We analyze several theropod eggshells from various localities, including oviraptorid Macroolithus yaotunensis eggshells from the Late Cretaceous deposits of Henan, Jiangxi, and Guangdong in China and alvarezsaurid Triprismatoolithus eggshell from the Two Medicine Formation of Montana, United States, with the scanning electron microscope (SEM), electron probe micro-analysis (EPMA), and Raman spectroscopy (RS). The elemental analysis with EPMA shows high concentration of phosphorus at the boundary between the eggshell and sediment, representing the hydroxyapatitic cuticle layer (HAp). Depletion of phosphorus in sediment excludes the allochthonous origin of the phosphorus in these eggshells. The chemometric analysis of Raman spectra collected from fossil and extant eggs provides further supportive evidence for the cuticle preservation in oviraptorid and probable alvarezsaurid eggshells. In accordance with our previous discovery of pigments preserved in Cretaceous oviraptorid dinosaur eggshells, we validate the cuticle preservation on dinosaur eggshells through deep time and offer a yet unexplored resource for chemical studies targeting the evolution of dinosaur nesting ecology. Our study also suggests that the cuticle structure can be traced far back to maniraptoran dinosaurs and enhance their reproductive success in a warm and mesic habitat such as Montana and southern China during the Late Cretaceous.

The origin of the bird's beak: new insights from dinosaur incubation periods.

The toothless beak of modern birds was considered as an adaption for feeding ecology; however, several recent studies suggested that developmental factors are also responsible for the toothless beak. Neontological and palaeontological studies have progressively uncovered how birds evolved toothless beaks and suggested that the multiple occurrences of complete edentulism in non-avian dinosaurs were the result of selection for specialized diets. Although developmental biology and ecological factors are not mutually exclusive, the conventional hypothesis that ecological factors account for the toothless beak appears insufficient. A recent study on dinosaur incubation period using embryonic teeth posited that tooth formation rate limits developmental speed, constraining toothed dinosaur incubation to slow reptilian rates. We suggest that selection for tooth loss was a side effect of selection for fast embryo growth and thus shorter incubation. This observation would also explain the multiple occurrences of tooth loss and beaks in non-avian dinosaur taxa crownward of Tyrannosaurus Whereas our hypothesis is an observation without any experimental supports, more studies of gene regulation of tooth formation in embryos would allow testing for the trade-off between incubation period and tooth development.

Dinosaur origin of egg color: oviraptors laid blue-green eggs.

Protoporphyrin (PP) and biliverdin (BV) give rise to the enormous diversity in avian egg coloration. Egg color serves several ecological purposes, including post-mating signaling and camouflage. Egg camouflage represents a major character of open-nesting birds which accomplish protection of their unhatched offspring against visually oriented predators by cryptic egg coloration. Cryptic coloration evolved to match the predominant shades of color found in the nesting environment. Such a selection pressure for the evolution of colored or cryptic eggs should be present in all open nesting birds and relatives. Many birds are open-nesting, but protect their eggs by continuous brooding, and thus exhibit no or minimal eggshell pigmentation. Their closest extant relatives, crocodiles, protect their eggs by burial and have unpigmented eggs. This phylogenetic pattern led to the assumption that colored eggs evolved within crown birds. The mosaic evolution of supposedly avian traits in non-avian theropod dinosaurs, however, such as the supposed evolution of partially open nesting behavior in oviraptorids, argues against this long-established theory. Using a double-checking liquid chromatography ESI-Q-TOF mass spectrometry routine, we traced the origin of colored eggs to their non-avian dinosaur ancestors by providing the first record of the avian eggshell pigments protoporphyrin and biliverdin in the eggshells of Late Cretaceous oviraptorid dinosaurs. The eggshell parataxon Macroolithus yaotunensis can be assigned to the oviraptor Heyuannia huangi based on exceptionally preserved, late developmental stage embryo remains. The analyzed eggshells are from three Late Cretaceous fluvial deposits ranging from eastern to southernmost China. Reevaluation of these taphonomic settings, and a consideration of patterns in the porosity of completely preserved eggs support an at least partially open nesting behavior for oviraptorosaurs. Such a nest arrangement corresponds with our reconstruction of blue-green eggs for oviraptors. According to the sexual signaling hypothesis, the reconstructed blue-green eggs support the origin of previously hypothesized avian paternal care in oviraptorid dinosaurs. Preserved dinosaur egg color not only pushes the current limits of the vertebrate molecular and associated soft tissue fossil record, but also provides a perspective on the potential application of this unexplored paleontological resource.