Patricia A. Holroyd

Best Practices for Justifying Fossil Calibrations

Our ability to correlate biological evolution with climate change, geological evolution, and other historical patterns is essential to understanding the processes that shape biodiversity. Combining data from the fossil record with molecular phylogenetics represents an exciting synthetic approach to this challenge. The first molecular divergence dating analysis (Zuckerkandl and Pauling 1962) was based on a measure of the amino acid differences in the hemoglobin molecule, with replacement rates established (calibrated) using paleontological age estimates from textbooks (e.g., Dodson 1960). Since that time, the amount of molecular sequence data has increased dramatically, affording ever-greater opportunities to apply molecular divergence approaches to fundamental problems in evolutionary biology. To capitalize on these opportunities, increasingly sophisticated divergence dating methods have been, and continue to be, developed. In contrast, comparatively, little attention has been devoted to critically assessing the paleontological and associated geological data used in divergence dating analyses. The lack of rigorous protocols for assigning calibrations based on fossils raises serious questions about the credibility of divergence dating results (e.g., Shaul and Graur 2002; Brochu et al. 2004; Graur and Martin 2004; Hedges and Kumar 2004; Reisz and Muller 2004a, 2004b; Theodor 2004; van Tuinen and Hadly 2004a, 2004b; van Tuinen et al. 2004; Benton and Donoghue 2007; Donoghue and Benton 2007; Parham and Irmis 2008; Ksepka 2009; Benton et al. 2009; Heads 2011). The assertion that incorrect calibrations will negatively influence divergence dating studies is not controversial. Attempts to identify incorrect calibrations through the use of a posteriori methods are available (e.g., Near and Sanderson 2004; Near et al. 2005; Rutschmann et al. 2007; Marshall 2008; Pyron 2010; Dornburg et al. 2011). We do not deny that a posteriori methods are a useful means of evaluating calibrations, but there can be no substitute for a priori assessment of the veracity of paleontological data. Incorrect calibrations, those based upon fossils that are phylogenetically misplaced or assigned incorrect ages, clearly introduce error into an analysis. Consequently, thorough and explicit justification of both phylogenetic and chronologic age assessments is necessary for all fossils used for calibration. Such explicit justifications will help to ensure that divergence dating studies are based on the best available data. Unfortunately, the majority of previously published calibrations lack explicit explanations and justifications of the age and phylogenetic position of the key fossils. In the absence of explicit justifications, it is difficult to distinguish between correct and incorrect calibrations, and it becomes difficult to reevaluate previous claims in light of new data. Paleontology is a dynamic science, with new data and perspectives constantly emerging as a result of new discoveries (see Kimura 2010 for a recent case where the age of the earliest known record of a clade was more than doubled). Calibrations based upon the best available evidence at a given time can become inappropriate as the discovery of new specimens, new phylogenetic analyses, and ongoing stratigraphic and geochronologic revisions refine our understanding of the fossil record. Our primary goals in this paper are to establish the best practices for justifying fossils used for the temporal calibration of molecular phylogenies. Our examples derive mainly, but not exclusively, from the vertebrate fossil record. We hope that our recommendations will lead to more credible calibrations and, as a result, more reliable divergence dates throughout the tree of life. A secondary goal is to help the community (researchers, editors, and reviewers) who might be unfamiliar with fossils to understand and overcome the challenges associated with using paleontological data. In order to accomplish these goals, we present a specimen-based protocol for selecting and documenting relevant fossils and discuss future directions for evaluating and utilizing phylogenetic and temporal data from the fossil record. We likewise encourage biologists relying on nonfossil calibrations for molecular divergence estimates (e.g., ages of island or mountain range formations, continental drift, and biomarkers) to develop their own set of rigorous guidelines so that their calibrations may also be evaluated in a systematic way.

Cretaceous Extinctions: Multiple Causes

![Figure][1] Deccan plateau basalts. Lava from Deccan volcanism formed distinct layering. CREDIT: GSFC/NASA In the Review “The Chicxulub Asteroid Impact and Mass Extinction at the Cretaceous-Paleogene boundary” (P. Schulte et al. , 5 March, p. [1214][2]), the terminal Cretaceous

Identifying Aquatic Habits Of Herbivorous Mammals Through Stable Isotope Analysis

Abstract Large-bodied, semiaquatic herbivorous mammals have been a recurring component of most continental ecosystems throughout the Cenozoic. Identification of these species in the fossil record has largely been based on the morphological similarities with present-day hippopotamids, leading to the designation of this pairing of body type and ecological niche as the hippo ecomorph. These morphological characters, however, may not always be diagnostic of aquatic habits. Here, enamel δ13C and δ18O values from living hippopotamuses were examined to define an isotopic signature unique to the hippo ecomorph. Although δ13C values do not support unique foraging habits for this ecomorph, living and fossil hippopotamids typically have low mean δ18O values relative to associated ungulates that fit a linear regression (δ18Ohippopotamids = 0.96 ± 0.09·δ18Ofauna − 1.67 ± 2.97; r2 = 0.886, p < 0.001). Modeling of oxygen fluxes in large mammals suggests that high water-turnover rates or increased water loss through feces and urine may explain this relationship. This relationship was then used to assess the aquatic adaptation of four purported hippo ecomorphs from the fossil record: Coryphodon (early Eocene), Moeritherium and Bothriogenys (early Oligocene), and Teleoceras (middle–late Miocene). Only fossil specimens of Moeritherium, Bothriogenys, and large species of Coryphodon had δ18O values expected for hippo ecomorphs; δ18O values for Teleoceras and a small species of Coryphodon were not significantly different from those of the associated fauna. These results show that the mean δ18O value of fossil specimens is an effective tool for assessing the aquatic habits of extinct species.

The Asian Origin of Anthropoidea Revisited

The concept of an Asian origin for the Anthropoidea has surfaced repeatedly in our attempts to discern the biogeographic and phyletic origins of the suborder (e.g., Pilgrim, 1927; Ba Maw et al., 1979; Gingerich, 1980; Ciochon and Ghiarelli, 1980; Ciochon et al., 1985). The primary evidence for such an Asian origin traditionally lies in the poorly known and phylogenetically enigmatic primate genera Amphipithecus and Pondaungia from the late middle Eocene Pondaung deposits of Burma (now Myanmar).

Relative Ages of Eocene Primate-Bearing Deposits of Asia

Paleontologists have often looked to Asia as a center of origin for anthropoid primates (e.g., Pilgrim, 1927; Colbert, 1937, 1938; Ba Maw et al., 1979; Gingerich, 1980; Ciochon and Chiarelli, 1980; Ciochon et al., 1985; Ciochon and Etler, 1994). In part, this notion has been founded in the accepted earlier occurrence of putative anthropoids or “protoanthropoids” in the Asian faunal record. In particular, the occurrence of characters similar to those found in African anthropoids in the Pondaung primates Amphipithecus and Pondaungia from presumed upper Eocene deposits in the Pondaung Hills of Burma (now Myanmar) has led to conjecture that this area represented the center of Origin for higher primates. However, the criterion of earlier occurrence has been eliminated by the redating of the earliest Fay urn anthropoid fauna to the late Eocene (Kappelman, 1992; Rasmussen et al., 1992) such that the Fayum anthropoids would be penecontemporaneous with the Asian primates Amphipithecus, Pondaungia, Hoanghonius, and Rencunius (see Chapter 7, this volume) based on current age assessments.

First report of snakes (Serpentes) from the Late Middle Eocene Pondaung Formation, Myanmar

Department of Anthropology, University of Iowa, Iowa City, IA 52242, USAThe reptile fauna of the Eocene Pondaung Formation of central My-anmar (Fig. 1) has received little attention compared to its well-knownmammalian fauna (e.g., Colbert, 1938; Tsubamoto et al., 2000). In priorreports, Buffetaut (1978) described indeterminate crocodylians and dy-rosaurids. Hutchison et al. (in press) described carettochelyid, trionychid,testudinoid, and pelomedusoid turtles from the formation. They addi-tionally noted the presence of agamid lizards, a pristichampsine croco-dilian, and snakes. The snakes are described here.The occurrence of snakes in the Pondaung Formation is significantbecause the fossil history of Paleogene South Asian snakes has histori-cally been under studied relative to the North American and Europeanrecords (e.g., Rage, 1984; Szyndlar, 1984; Holman, 2000), despite hypoth-eses predicting the region as the origin of extant clades (e.g., Underwoodand Stimson, 1990; Rage et al., 1992). The South Asian record consistsprimarily of marine palaeophiid taxa as well as terrestrial/terrigenousspecimens referred to Boidae (Boinae + Erycinae, Table 1). The onlyderived snakes from the South Asian Paleogene are six colubrid verte-brae from the late Eocene Krabi Basin of Thailand (Rage et al., 1992)and a single vertebra referred to Colubroidea (possibly Colubridae) fromthe early Eocene of India (Rage et al., 2003). The absence of coevalcolubroids elsewhere (Rage, 1988), combined with the occurrence of theKrabi Basin record, led Rage et al. (1992) to conclude that Asia repre-sents the center of origin for Colubridae, the most speciose and diverseextant snake clade.The Pondaung snake record consists of two specimens derived fromterrestrial sediments occurring in the upper 100+ meters of the otherwisemarine Pondaung Formation as it crops out to the west and northwest ofMogaung village, Myaing Township, central Myanmar (Fig. 1). Thesnake localities are interpreted as swale-fills and/or paleosols depositedin an ancient floodplain (Soe et al., 2002; see also Ciochon and Gunnell,2002, and Gunnell et al., 2002 for more detailed discussions of the lithol-ogy and stratigraphy of these localities). Traditionally, the age of thePondaung fauna was considered to be late Eocene (e.g., Pilgrim, 1928;Bender, 1983); however, Holroyd and Ciochon (1994) concluded that thePondaung fauna is latest middle Eocene (Bartonian) in age and broadlycontemporaneous with Asian faunas assigned to the SharamurunianLand Mammal Age, a finding confirmed by fission-track dates of 37.2 ±1.3 Ma (Tsubamoto et al., 2002). These findings indicate that thePondaung fauna is slightly older than the Krabi Basin record, which hasbeen dated between 33.54 and 34.65 Ma in age based on paleomagneticcorrelations (Benammi et al., 2001).Here we describe the Pondaung snakes and discuss their relativeimplications for paleoecology, divergence timings, and biogeographichistories. We refrain from erecting new taxa for the Pondaung speci-mens because the record is limited to just the two elements and neitheris complete. Additional material will be necessary to determine wheth-er or not the Pondaung record represents new, distinct taxa, or indi-vidual or intracolumnar variants of previously known South Asiansnakes.

Morphology and Evolution of Turtles

The unquestioned unity of the Chelonia provides a necessary basis for establishing their interrelationships and determining the evolutionary history within the group. On the other hand, the host of uniquely derived features of the oldest known turtles make it extremely difficult to establish their ancestry among more primitive amniotes. This is illustrated by the great diversity of taxa that continue to be proposed as putative sister-taxa of turtles without general acceptance of any. Nearly every major clade of early amniotes from the late Paleozoic and early Mesozoic has been proposed as a possible sister-taxon of turtles, from synapsids to anapsids and diapsids, including pelycosaurs, captorhinomorphs, procolophonids, pareiasaurs, aquatic placodonts and crocodiles, but none possess derived characters that could be synapomorphic with the unique skeletal structure and patterns of development of the chelonian skull, carapace or plastron, which had reached an essentially modern configuration by the Late Triassic. Numerous molecular biologists have attempted to establish the closest sister-group of turtles through analyses of a host of living species, but there is no way for them to preclude turtles from having evolved from one or another of the Paleozoic or early Mesozoic clades that have become extinct without leaving any other living descendants. On the other hand, recent studies of the genetic and molecular aspects of the development of the carapace and plastron imply unique patterns of evolutionary change that cannot be recognized in any of the other amniote lineages, living or dead. This, together with the retention of a skull without temporal fenestration implies a very early divergence from a lineage that probably retained an anapsid skull configuration. This problem may be resolved by more detailed study of the enigmatic genus Eunotosaurus, from the Late Permian of South Africa.

Rencunius zhoui, New Primate from the Late Middle Eocene of Henan, China, and a Comparison with Some Early Anthropoidea

Late Eocene primates of Asia are often mentioned in discussions of anthropoid origins. This is in part because of the distinctive morphologies of Asian Eocene primates that document an otherwise hidden diversity of potential anthropoid ancestors. Asia also draws our attention as a large, centrally placed geographic region that is still inadequately known paleontologically. Asia and its Eocene primates are important for understanding both the phylogenetic and biogeographic history of primate and anthropoid diversification.

Paleogeography, Paleobiogeography, and Anthropoid Origins

The study of anthropoid origins has long been tied to hypotheses postulating Eocene dispersal of early anthropoids or their precursors into Africa from Europe (e.g., Gingerich, 1975; Rasmussen and Simons, 1988), from Asia (e.g., Gingerich, 1980; Ciochon and Ghiarelli, 1980; Giochon et al., 1985; Rosenberger, 1986), or from South America (e.g., Szalay, 1976). Similarly, investigations of platyrrhine origins have focused on either a North American (e.g., Gingerich, 1980; Hoffstetter, 1972; Wood, 1980; Rosenberger, 1986) or an African source (e.g., Lavocat, 1974, 1980; Giochon and Chiarelli, 1980; Fleagle, 1986). These different scenarios have been based in large part on putative ancestor-descendant relationships and the identification of early anthropoids or protoanthropoids in the presumed source areas but also have relied on current understanding of temporal relationships between faunas and reconstructions of Eocene paleogeography and paleobiogeography to determine the probable timing, mode, and route of dispersal.

THE ANTIQUITY OF AFRICAN TORTOISES

Tortoises (Testudinidae) are a diverse and highly specialized clade of terrestrial turtles that currently inhabit five continents. The global radiation of tortoises in the Paleogene, part of an explosive radiation of testudinoid turtles out of Asia, is poorly understood. The oldest known tortoises are from the late Paleocene of Mongolia (Parham, pers. obs. at PIN), and early in the Eocene they are known to have colonized North America and Europe (e.g., Hutchison, 1998; Lapparent de Broin, 2001). At some point in the early Paleogene they marched or floated to Africa. Today, the ancestors of those first invaders have evolved into the most diverse tortoise fauna in the world; Africa is home to 10 of the 13 extant genera (Iverson, 1992; Lapparent de Broin, 2000). One of the most poorly understood episodes in the early testudinid range expansion is the dispersal of tortoises into Africa. Fossil tortoises have been known from Africa since the beginning of the last century, when Andrews (1902) noted that a ‘‘gigantic land-tortoise’’ had been found by H. J. L. Beadnell. However, the age of these tortoises has never been firmly established, because early collecting records were not precise with regard to the specific quarries from which they were collected. Three species of Testudo were described (Andrews and Beadnell, 1903; Andrews, 1906) as having come from the upper Eocene ‘‘Fluviomarine’’ deposits of the Jebel Qatrani Formation in the Fayum Province of Egypt. In the intervening years, the Jebel Qatrani Formation came to be regarded as early Oligocene in age (e.g., Simons, 1968), then partly late Eocene and partly early Oligocene (Kappelman, 1992; Kappelman et al., 1992). No further testudinids have been reported from Egypt, although fragmentary testudinid fossils possibly close to the Egyptian taxon have been recovered from lower Oligocene sediments in Oman (Thomas et al., 1989; Lapparent de Broin, 2000). Most recently, Lapparent de Broin (2000) reviewed the African fossil record of turtles and conservatively reported the age of the Fayum tortoises as early Oligocene. Resolution of the age of Africa’s oldest tortoises has been difficult because specimens housed in European institutions (Natural History Museum, London, and Staatliches Museum, Stuttgart) lack detailed locality data, and more recent fieldwork in the area by E. L. Simons and crews (materials housed at Yale Peabody Museum, Cairo Geological Museum, and Duke University Primate Center, Durham, North Carolina) has not yielded remains of these comparatively rare reptiles. Reevaluation of older collections from the Jebel Qatrani Formation has uncovered the only African tortoise with associated stratigraphic data indicating a late Eocene age, allowing us to place this taxon in an updated geochronologic context and providing us with the opportunity to resolve several taxonomic issues regarding these specimens. Abbreviations Used AMNH, American Museum of Natural History, New York; BMNH, The Natural History Museum, London; CGM, Cairo Geological Museum, Cairo; PIN, Paleontological Institute, Moscow.