Spectroscopy on the wing: Investigating possible differences in protein secondary structures in feather shafts of birds using Raman spectroscopy.

The central shaft of a bird's flight feather bears most of the aerodynamic load during flight and exhibits some remarkable mechanical properties. The shaft comprises two parts, the calamus and the rachis. The calamus is at the base of the shaft, while the rachis is the longer upper part which supports the vanes. The shaft is composed of a fibrous outer cortex, and an inner foam-like core. Recent nanoindentation experiments have indicated that reduced modulus values, Er, for the inner and outer regions of the cortex can vary, with the Er values of the inner region slightly greater than those of the outer region. In this work, Raman spectroscopy is used to investigate the protein secondary structures in the inner and outer regions of the feather cortex. Analysis of the Amide I region of Raman spectra taken from four birds (Swan, Gull, Mallard and Kestrel) shows that the ß-sheet structural component decreases between the inner and outer region, relative to the protein side-chain components. This finding is consistent with the proposal that Er values are greater in the inner region than the outer region. This work has shown that Raman spectroscopy can be used effectively to study the change in protein secondary structure between the inner and outer regions of a feather shaft.

Imaging techniques for observing laminar geometry in the feather shaft cortex.

Bird feather shafts are light, stiff and strong, but the fine details of how their structure, mechanics and function relate to one another remains poorly understood. The missing piece in our understanding may be the various fibrous layers that make up the shaft's cortex. Detailed imaging techniques are needed to enable us to capture, analyse and quantify these layers before we can begin to unravel the relationship between their structure, mechanics and function. We show that Serial-Block-Face scanning electron microscopy, scanning confocal polarised microscopy and synchrotron-based computed tomography are three suitable techniques to investigate layer thickness and fibre orientation in the feather cortex. These techniques and other are discussed in terms of their ability to resolve the fibrous laminar structure of the feather cortex, on sample preparation, and on throughput. Annotated images are presented for each and less suitable techniques are presented in the Supplementary Material. LAY DESCRIPTION: Bird feathers have a light, stiff and strong central shaft. However, the fine details of how their structure, mechanics and function relate to one another remains poorly understood. The missing piece in our understanding may have to do with how fibrous layers within the shaft vary in thickness and alignment. Detailed imaging techniques are needed so that we can quantify some of this variation before we can revisit some long-unanswered questions about the feather shaft's structure, mechanics and function. We investigate a number of microscopy techniques and show that three techniques are suitable for the sort of investigation that is required. These techniques and others are discussed in terms of their ability to resolve the layers' thickness and alignment, on sample preparation, and on the sample sizes they are able to process. Annotated images are presented and discussed for each of the three techniques and unsuitable techniques receive the same examination in the Supplementary Material.

Skin pigmentation provides evidence of convergent melanism in extinct marine reptiles.

Throughout the animal kingdom, adaptive colouration serves critical functions ranging from inconspicuous camouflage to ostentatious sexual display, and can provide important information about the environment and biology of a particular organism. The most ubiquitous and abundant pigment, melanin, also has a diverse range of non-visual roles, including thermoregulation in ectotherms. However, little is known about the functional evolution of this important biochrome through deep time, owing to our limited ability to unambiguously identify traces of it in the fossil record. Here we present direct chemical evidence of pigmentation in fossilized skin, from three distantly related marine reptiles: a leatherback turtle, a mosasaur and an ichthyosaur. We demonstrate that dark traces of soft tissue in these fossils are dominated by molecularly preserved eumelanin, in intimate association with fossilized melanosomes. In addition, we suggest that contrary to the countershading of many pelagic animals, at least some ichthyosaurs were uniformly dark-coloured in life. Our analyses expand current knowledge of pigmentation in fossil integument beyond that of feathers, allowing for the reconstruction of colour over much greater ranges of extinct taxa and anatomy. In turn, our results provide evidence of convergent melanism in three disparate lineages of secondarily aquatic tetrapods. Based on extant marine analogues, we propose that the benefits of thermoregulation and/or crypsis are likely to have contributed to this melanisation, with the former having implications for the ability of each group to exploit cold environments.

A Jurassic avialan dinosaur from China resolves the early phylogenetic history of birds.

The recent discovery of small paravian theropod dinosaurs with well-preserved feathers in the Middle-Late Jurassic Tiaojishan Formation of Liaoning Province (northeastern China) has challenged the pivotal position of Archaeopteryx, regarded from its discovery to be the most basal bird. Removing Archaeopteryx from the base of Avialae to nest within Deinonychosauria implies that typical bird flight, powered by the forelimbs only, either evolved at least twice, or was subsequently lost or modified in some deinonychosaurians. Here we describe the complete skeleton of a new paravian from the Tiaojishan Formation of Liaoning Province, China. Including this new taxon in a comprehensive phylogenetic analysis for basal Paraves does the following: (1) it recovers it as the basal-most avialan; (2) it confirms the avialan status of Archaeopteryx; (3) it places Troodontidae as the sister-group to Avialae; (4) it supports a single origin of powered flight within Paraves; and (5) it implies that the early diversification of Paraves and Avialae took place in the Middle-Late Jurassic period.

The four-flipper swimming method of plesiosaurs enabled efficient and effective locomotion.

The extinct ocean-going plesiosaurs were unique within vertebrates because they used two flipper pairs identical in morphology for propulsion. Although fossils of these Mesozoic marine reptiles have been known for more than two centuries, the function and dynamics of their tandem-flipper propulsion system has always been unclear and controversial. We address this question quantitatively for the first time in this study, reporting a series of precisely controlled water tank experiments that use reconstructed plesiosaur flippers scaled from well-preserved fossils. Our aim was to determine which limb movements would have resulted in the most efficient and effective propulsion. We show that plesiosaur hind flippers generated up to 60% more thrust and 40% higher efficiency when operating in harmony with their forward counterparts, when compared with operating alone, and the spacing and relative motion between the flippers was critical in governing these increases. The results of our analyses show that this phenomenon was probably present across the whole range of plesiosaur flipper motion and resolves the centuries-old debate about the propulsion style of these marine reptiles, as well as indicating why they retained two pairs of flippers for more than 100 million years.

Irregularly calcified eggs and eggshells of Caiman latirostris (Alligatoridae: Crocodylia).

We describe irregularly calcified egg and eggshell morphologies for the first time in nests of the broad-snouted caiman, Caiman latirostris. Research is based on detailed descriptions of 270 eggs from a total sample of 46,800 collected between 2005 and 2011 in Santa Fe Province, Argentina, and encompasses animals from both natural habitats and held in captivity. We discuss possible reasons for the occurrence of eggs with different mineralisation patterns in our extensive C. latirostris field sample and its conservation significance; the chemistry of egg laying in amniotes is sensitive to environmental contamination which, in turn, has biological implications. Based on our egg sample, we identify two caiman eggshell abnormalities: (1) regularly calcified eggs with either calcitic nodules or superficial wrinkles at one egg end and (2) irregularly calcified eggs with structural gaps that weaken the shell. Some recently laid clutches we examined included eggs with most of the shell broken and detached from the flexible membrane. Most type 1 regularly calcified eggs lost their initial calcified nodules during incubation, suggesting that these deposits do not affect embryo survival rates. In contrast, irregularly calcified caiman eggs have a mean hatching success rate of 8.9% (range 0-38%) across our sample compared to a mean normal success of 75%. Most irregularly calcified caiman eggs probably die because of infections caused by fungi and bacteria in the organic nest material, although another possible explanation that merits further investigation could be an increase in permeability, leading to embryo dehydration.

Constraints on the wing morphology of pterosaurs.

Animals that fly must be able to do so over a huge range of aerodynamic conditions, determined by weather, wind speed and the nature of their environment. No single parameter can be used to determine-let alone measure-optimum flight performance as it relates to wing shape. Reconstructing the wings of the extinct pterosaurs has therefore proved especially problematic: these Mesozoic flying reptiles had a soft-tissue membranous flight surface that is rarely preserved in the fossil record. Here, we review basic mechanical and aerodynamic constraints that influenced the wing shape of pterosaurs, and, building on this, present a series of theoretical modelling results. These results allow us to predict the most likely wing shapes that could have been employed by these ancient reptiles, and further show that a combination of anterior sweep and a reflexed proximal wing section provides an aerodynamically balanced and efficient theoretical pterosaur wing shape, with clear benefits for their flight stability.

A gigantic bird from the Upper Cretaceous of Central Asia.

We describe an enormous Late Cretaceous fossil bird from Kazakhstan, known from a pair of edentulous mandibular rami (greater than 275 mm long), which adds significantly to our knowledge of Mesozoic avian morphological and ecological diversity. A suite of autapomorphies lead us to recognize the specimen as a new taxon. Phylogenetic analysis resolves this giant bird deep within Aves as a basal member of Ornithuromorpha. This Kazakh fossil demonstrates that large body size evolved at least once outside modern birds (Neornithes) and reveals hitherto unexpected trophic diversity within Cretaceous Aves.

A drowned Mesozoic bird breeding colony from the Late Cretaceous of Transylvania.

Despite a rapidly improving fossil record, the reproductive biology of Mesozoic birds remains poorly known: only a handful of undisputed, isolated Cretaceous eggs (some containing embryonic remains) are known. We report here the first fossil evidence for a breeding colony of Mesozoic birds, preserved at the Late Cretaceous (Maastrichtian) Oarda de Jos (Od) site in the Sebes area of Transylvania, Romania. A lens of calcareous mudstone with minimum dimensions of 80 cm length, 50 cm width and 20 cm depth contains thousands of tightly packed, morphologically homogenous eggshell fragments, seven near-complete eggs and neonatal and adult avialan skeletal elements. Eggshell forms 70-80 % of the matrix, and other fossils are entirely absent. The bones exhibit clear characters of the Cretaceous avialan clade Enantiornithes, and the eggshell morphology is also consistent with this identification. Both taphonomy and lithology show that the components of this lens were deposited in a single flood event, and we conclude that it represents the drowned remains of a larger enantiornithine breeding colony, swamped by rising water, washed a short distance and deposited in a shallow, low-energy pond. The same fate often befalls modern bird colonies. Such a large concentration of breeding birds suggests aquatic feeding in this species, augments our understanding of enantiornithine biology and shows that colonial nesting was not unique to crown birds.

Biomechanics of the unique pterosaur pteroid.

Pterosaurs, flying reptiles from the Mesozoic, had wing membranes that were supported by their arm bones and a super-elongate fourth finger. Associated with the wing, pterosaurs also possessed a unique wrist bone--the pteroid--that functioned to support the forward part of the membrane in front of the leading edge, the propatagium. Pteroid shape varies across pterosaurs and reconstructions of its orientation vary (projecting anteriorly to the wing leading edge or medially, lying alongside it) and imply differences in the way that pterosaurs controlled their wings. Here we show, using biomechanical analysis and considerations of aerodynamic efficiency of a representative ornithocheirid pterosaur, that an anteriorly orientated pteroid is highly unlikely. Unless these pterosaurs only flew steadily and had very low body masses, their pteroids would have been likely to break if orientated anteriorly; the degree of movement required for a forward orientation would have introduced extreme membrane strains and required impractical tensioning in the propatagium membrane. This result can be generalized for other pterodactyloid pterosaurs because the resultant geometry of an anteriorly orientated pteroid would have reduced the aerodynamic performance of all wings and required the same impractical properties in the propatagium membrane. We demonstrate quantitatively that the more traditional reconstruction of a medially orientated pteroid was much more stable both structurally and aerodynamically, reflecting likely life position.

An abelisaurid (Dinosauria: Theropoda) ilium from the Upper Cretaceous (Cenomanian) of the Kem Kem beds, Morocco.

Abelisaurid theropods first appear in the fossil record in the early Jurassic and survived at least until the end of the Mesozoic. They were known to have dominated South America, India and Madagascar but were not so abundant in North America or Asia. Much less is known about their presence in Africa, although there has been several recent discoveries of abelisaurid material in Morocco. Here we add a partially preserved ilium to a growing body of evidence that suggests abelisaurs might also have dominated Africa.

Ultraviolet light illuminates the avian nature of the Berlin Archaeopteryx skeleton.

The question of whether the iconic avialan Archaeopteryx was capable of active flapping flight or only passive gliding is still unresolved. This study contributes to this debate by reporting on two key aspects of this fossil that are visible under ultraviolet (UV) light. In contrast to previous studies, we show that most of the vertebral column of the Berlin Archaeopteryx possesses intraosseous pneumaticity, and that pneumatic structures also extend beyond the anterior thoracic vertebrae in other specimens of Archaeopteryx. With a minimum Pneumaticity Index (PI) of 0.39, Archaeopteryx had a much more lightweight skeleton than has been previously reported, comprising an air sac-driven respiratory system with the potential for a bird-like, high-performance metabolism. The neural spines of the 16th to 22nd presacral vertebrae in the Berlin Archaeopteryx are bridged by interspinal ossifications, and form a rigid notarium-like structure similar to the condition seen in modern birds. This reinforced vertebral column, combined with the extensive development of air sacs, suggests that Archaeopteryx was capable of flapping its wings for cursorial and/or aerial locomotion.

Older birds have better feathers: A longitudinal study on the long-distance migratory Sand Martin, Riparia riparia.

Feather quality is of critical importance to long-distance migratory birds. Here, we report a series of analyses of a unique data set encompassing known-age individuals of the long-distance migratory Sand Martin (Riparia riparia). Sampling over 17 years along the Tisza River, eastern Hungary, has resulted in the recapture of numerous individuals enabling longitudinal and cross-sectional investigation of the role of adaptation to variable environmental conditions on feather morphology. We show that older individuals tend to possess better quality feathers, measured using bending stiffness, feather length and thickness as proxies. Bending stiffness and feather thickness do not change with individual age, in contrast with increases in feather length and declines in daily feather growth versus age of individual alongside moult duration. Individuals who live to older ages tend to have similar, or higher, feather growth rates and better feather quality than individuals captured at younger ages. Thus, on the basis of strong selection against individuals with slow feather growth, as seen in other species of swallows and martins, which causes a delay in moult completion, the results of this analysis highlight the potential cost of producing better quality feathers when this depends on moult duration. Feather length also does change during the lifetime of the individual and thus enabled us to further investigate influence of individual and environmental conditions during the moult. The results of this analysis provide important insights on the adaptive significance of these traits, and the potential use of physical characteristics in unravelling the reasons why long distance migratory bird populations are in global decline.

A mixed vertebrate eggshell assemblage from the Transylvanian Late Cretaceous.

A Late Cretaceous-aged multi-taxon nesting site from Romania preserved in three dimensions reveals the earliest example of nest site sharing yet known from the vertebrate fossil record. Eggshell and osteological evidence combined in this single accumulation demonstrate that at least four vertebrate taxa including enantiornithine birds and another avian of indeterminate affinities as well as crocodylomorphs and gekkotan squamates nested together in the same place. Colonial nesting in enantiornithines was previously described from this site; here, we present the first fossil evidence that other vertebrates also nested in the same place, perhaps exploiting the presence of the large bird colony. We describe four distinct eggshell morphotypes that have been collected from this site and draw palaeoecological inferences based on this unique multi-taxon nesting association.

Complex neuroanatomy in the rostrum of the Isle of Wight theropod Neovenator salerii.

The discovery of large, complex, internal canals within the rostra of fossil reptiles has been linked with an enhanced tactile function utilised in an aquatic context, so far in pliosaurids, the Cretaceous theropod Spinosaurus, and the related spinosaurid Baryonyx. Here, we report the presence of a complex network of large, laterally situated, anastomosing channels, discovered via micro-focus computed tomography (µCT), in the premaxilla and maxilla of Neovenator, a mid-sized allosauroid theropod from the Early Cretaceous of the UK. We identify these channels as neurovascular canals, that include parts of the trigeminal nerve; many branches of this complex terminate on the external surfaces of the premaxilla and maxilla where they are associated with foramina. Neovenator is universally regarded as a 'typical' terrestrial, predatory theropod, and there are no indications that it was aquatic, amphibious, or unusual with respect to the ecology or behaviour predicted for allosauroids. Accordingly, we propose that enlarged neurovascular facial canals shouldn't be used to exclusively support a model of aquatic foraging in theropods and argue instead that an enhanced degree of facial sensitivity may have been linked with any number of alternative behavioural adaptations, among them defleshing behaviour, nest selection/maintenance or social interaction.