Chemical, experimental, and morphological evidence for diagenetically altered melanin in exceptionally preserved fossils.

In living organisms, color patterns, behavior, and ecology are closely linked. Thus, detection of fossil pigments may permit inferences about important aspects of ancient animal ecology and evolution. Melanin-bearing melanosomes were suggested to preserve as organic residues in exceptionally preserved fossils, retaining distinct morphology that is associated with aspects of original color patterns. Nevertheless, these oblong and spherical structures have also been identified as fossilized bacteria. To date, chemical studies have not directly considered the effects of diagenesis on melanin preservation, and how this may influence its identification. Here we use time-of-flight secondary ion mass spectrometry to identify and chemically characterize melanin in a diverse sample of previously unstudied extant and fossil taxa, including fossils with notably different diagenetic histories and geologic ages. We document signatures consistent with melanin preservation in fossils ranging from feathers, to mammals, to amphibians. Using principal component analyses, we characterize putative mixtures of eumelanin and phaeomelanin in both fossil and extant samples. Surprisingly, both extant and fossil amphibians generally exhibit melanosomes with a mixed eumelanin/phaeomelanin composition rather than pure eumelanin, as assumed previously. We argue that experimental maturation of modern melanin samples replicates diagenetic chemical alteration of melanin observed in fossils. This refutes the hypothesis that such fossil microbodies could be bacteria, and demonstrates that melanin is widely responsible for the organic soft tissue outlines in vertebrates found at exceptional fossil localities, thus allowing for the reconstruction of certain aspects of original pigment patterns.

Water supply and demand remain coordinated during breakdown of the global scaling relationship between leaf size and major vein density.

Vein networks that disobey the global scaling of major vein density with leaf size shed light on functional constraints of vein network formation in dicotyledons. Understanding their evolution, distribution and impact on vein-stomata-climate associations is an important contribution to our global view of vein network organization. Based on vein traits of 55 species of pantropical Ochnaceae, stomata and climatic niche data, and a dated molecular phylogeny, we unveil major structural shifts in vein networks through deep time, relationships between leaf size, vein and stomata traits, and their interplay with climate. Dense 2° veins, reduction of minor veins and the associated breakdown of vein-leaf size scaling evolved multiple times independently in Ochnaceae. In spite of the drastic changes in vein architecture in this venation type, vein and stomatal densities remain correlated. Our study demonstrates that shortening the major vein-stomata distance is economically not less advantageous than by increasing minor vein density, as illustrated by the same degree of coordination between vein and stomatal densities and the similar construction costs across networks with dense 2° veins and those with 'normally' spaced 2° veins.

Inferring echolocation in ancient bats.

Laryngeal echolocation, used by most living bats to form images of their surroundings and to detect and capture flying prey, is considered to be a key innovation for the evolutionary success of bats, and palaeontologists have long sought osteological correlates of echolocation that can be used to infer the behaviour of fossil bats. Veselka et al. argued that the most reliable trait indicating echolocation capabilities in bats is an articulation between the stylohyal bone (part of the hyoid apparatus that supports the throat and larynx) and the tympanic bone, which forms the floor of the middle ear. They examined the oldest and most primitive known bat, Onychonycteris finneyi (early Eocene, USA), and argued that it showed evidence of this stylohyal-tympanic articulation, from which they concluded that O. finneyi may have been capable of echolocation. We disagree with their interpretation of key fossil data and instead argue that O. finneyi was probably not an echolocating bat.

Primitive Early Eocene bat from Wyoming and the evolution of flight and echolocation.

Bats (Chiroptera) represent one of the largest and most diverse radiations of mammals, accounting for one-fifth of extant species. Although recent studies unambiguously support bat monophyly and consensus is rapidly emerging about evolutionary relationships among extant lineages, the fossil record of bats extends over 50 million years, and early evolution of the group remains poorly understood. Here we describe a new bat from the Early Eocene Green River Formation of Wyoming, USA, with features that are more primitive than seen in any previously known bat. The evolutionary pathways that led to flapping flight and echolocation in bats have been in dispute, and until now fossils have been of limited use in documenting transitions involved in this marked change in lifestyle. Phylogenetically informed comparisons of the new taxon with other bats and non-flying mammals reveal that critical morphological and functional changes evolved incrementally. Forelimb anatomy indicates that the new bat was capable of powered flight like other Eocene bats, but ear morphology suggests that it lacked their echolocation abilities, supporting a 'flight first' hypothesis for chiropteran evolution. The shape of the wings suggests that an undulating gliding-fluttering flight style may be primitive for bats, and the presence of a long calcar indicates that a broad tail membrane evolved early in Chiroptera, probably functioning as an additional airfoil rather than as a prey-capture device. Limb proportions and retention of claws on all digits indicate that the new bat may have been an agile climber that employed quadrupedal locomotion and under-branch hanging behaviour.

Improved non-destructive 2D and 3D X-ray imaging of leaf venation.

BACKGROUND: Leaf venation traits are important for many research fields such as systematics and evolutionary biology, plant physiology, climate change, and paleoecology. In spite of an increasing demand for vein trait data, studies are often still data-limited because the development of methods that allow rapid generation of large sets of vein data has lagged behind. Recently, non-destructive X-ray technology has proven useful as an alternative to traditional slow and destructive chemical-based methods. Non-destructive techniques more readily allow the use of herbarium specimens, which provide an invaluable but underexploited resource of vein data and related environmental information. The utility of 2D X-ray technology and microfocus X-ray computed tomography, however, has been compromised by insufficient image resolution. Here, we advanced X-ray technology by increasing image resolution and throughput without the application of contrast agents. RESULTS: For 2D contact microradiography, we developed a method which allowed us to achieve image resolutions of up to 7 µm, i.e. a 3.6-fold increase compared to the industrial standard (25 µm resolution). Vein tracing was further optimized with our image processing standards that were specifically adjusted for different types of leaf structure and the needs of higher imaging throughput. Based on a test dataset, in 91% of the samples the 7 µm approach led to a significant improvement in estimations of minor vein density compared to the industrial standard. Using microfocus X-ray computed tomography, very high-resolution images were obtained from a virtual 3D-2D transformation process, which was superior to that of 3D images. CONCLUSIONS: Our 2D X-ray method with a significantly improved resolution advances rapid non-destructive bulk scanning at a quality that in many cases is sufficient to determine key venation traits. Together with our high-resolution microfocus X-ray computed tomography method, both non-destructive approaches will help in vein trait data mining from museum collections, which provide an underexploited resource of historical and recent data on environmental and evolutionary change. In spite of the significant increase in effective image resolution, a combination of high-throughput and full visibility of the vein network (including the smallest veins and their connectivity) remains challenging, however.

The Only Known Jawed Vertebrate with Four Eyes and the Bauplan of the Pineal Complex.

The pineal and parapineal organs are dorsal outpocketings of the vertebrate diencephalon that play key roles in orientation and in circadian and annual cycles. Lampreys are four eyed in that both the pineal and parapineal form eyelike photosensory structures, but the pineal is the dominant or sole median photosensory structure in most lower vertebrate clades. The pineal complex has been thought to evolve in a single direction by losing photosensory and augmenting secretory function in the transitions from three-eyed lower vertebrates to two-eyed mammals and archosaurs [1-3]. Yet the widely accepted elaboration of the parapineal instead of the pineal as the primary median photosensory organ [4] in Lepidosauria (lizards, snakes, and tuataras) hints at a more complex evolutionary history. Here we present evidence that a fourth eye re-evolved from the pineal organ at least once within vertebrates, specifically in an extinct monitor lizard, Saniwa ensidens, in which pineal and parapineal eyes were present simultaneously. The tandem midline location of these structures confirms in a striking fashion the proposed homology of the parietal eye with the parapineal organ and refutes the classical model of pineal bilaterality. It furthermore raises questions about the evolution and functional interpretation of the median photosensory organ in other tetrapod clades.

Description of a Well Preserved Fetus of the European Eocene Equoid Eurohippus messelensis.

The early Middle Eocene locality of Grube Messel, near Darmstadt (Germany), is famous for its complete vertebrate skeletons. The degree of preservation of soft tissues, such as body silhouettes, internal organs and gut contents, is frequently remarkable. The present specimen was analyzed for remnants of the reproductive system. Classic anatomy and osteology and high-resolution micro-x-ray were applied to describe the fetus of the European Eocene equoid Eurohippus messelensis. Scanning electronic microscopy (SEM) was used for determination of soft tissue remnants. The fetus is the earliest and best-preserved fossil specimen of its kind. The postcranial fetal skeleton is almost complete and largely articulated, allowing the conclusion that the pregnant mare was in late gestation. The apparent intrauterine position of the fetus is normal for the phase of pregnancy. Death of mare and fetus were probably not related to problems associated with parturition. Soft tissue interpreted as the uteroplacenta and a broad uterine ligament are preserved due to bacterial activity and allow considerations on the evolutionary development of the structures.

Controls on gut phosphatisation: the trilobites from the Weeks Formation Lagerstätte (Cambrian; Utah).

Despite being internal organs, digestive structures are frequently preserved in Cambrian Lagerstätten. However, the reasons for their fossilisation and their biological implications remain to be thoroughly explored. This is particularly true with arthropods--typically the most diverse fossilised organisms in Cambrian ecosystems--where digestive structures represent an as-yet underexploited alternative to appendage morphology for inferences on their biology. Here we describe the phosphatised digestive structures of three trilobite species from the Cambrian Weeks Formation Lagerstätte (Utah). Their exquisite, three-dimensional preservation reveals unique details on trilobite internal anatomy, such as the position of the mouth and the absence of a differentiated crop. In addition, the presence of paired pygidial organs of an unknown function is reported for the first time. This exceptional material enables exploration of the relationships between gut phosphatisation and the biology of organisms. Indeed, soft-tissue preservation is unusual in these fossils as it is restricted to the digestive structures, which indicates that the gut played a central role in its own phosphatisation. We hypothesize that the gut provided a microenvironment where special conditions could develop and harboured a source of phosphorus. The fact that gut phosphatization has almost exclusively been observed in arthropods could be explained by their uncommon ability to store ions (including phosphorous) in their digestive tissues. However, in some specimens from the Weeks Formation, the phosphatisation extends to the entire digestive system, suggesting that trilobites might have had some biological particularities not observed in modern arthropods. We speculate that one of them might have been an increased capacity for ion storage in the gut tissues, related to the moulting of their heavily-mineralised carapace.

Complete primate skeleton from the Middle Eocene of Messel in Germany: morphology and paleobiology.

BACKGROUND: The best European locality for complete Eocene mammal skeletons is Grube Messel, near Darmstadt, Germany. Although the site was surrounded by a para-tropical rain forest in the Eocene, primates are remarkably rare there, and only eight fragmentary specimens were known until now. Messel has now yielded a full primate skeleton. The specimen has an unusual history: it was privately collected and sold in two parts, with only the lesser part previously known. The second part, which has just come to light, shows the skeleton to be the most complete primate known in the fossil record. METHODOLOGY/PRINCIPAL FINDINGS: We describe the morphology and investigate the paleobiology of the skeleton. The specimen is described as Darwinius masillae n.gen. n.sp. belonging to the Cercamoniinae. Because the skeleton is lightly crushed and bones cannot be handled individually, imaging studies are of particular importance. Skull radiography shows a host of teeth developing within the juvenile face. Investigation of growth and proportion suggest that the individual was a weaned and independent-feeding female that died in her first year of life, and might have attained a body weight of 650-900 g had she lived to adulthood. She was an agile, nail-bearing, generalized arboreal quadruped living above the floor of the Messel rain forest. CONCLUSIONS/SIGNIFICANCE: Darwinius masillae represents the most complete fossil primate ever found, including both skeleton, soft body outline and contents of the digestive tract. Study of all these features allows a fairly complete reconstruction of life history, locomotion, and diet. Any future study of Eocene-Oligocene primates should benefit from information preserved in the Darwinius holotype. Of particular importance to phylogenetic studies, the absence of a toilet claw and a toothcomb demonstrates that Darwinius masillae is not simply a fossil lemur, but part of a larger group of primates, Adapoidea, representative of the early haplorhine diversification.