Ice Age effects on genetic divergence of the American crocodile (Crocodylus acutus) in Panama: reconstructing limits of gene flow and environmental ranges: a reply to O'Dea et al.

O'Dea et al. (2022) (Pleistocene sea level changes and crocodile population histories on the isthmus of panama: a comment on Avila-Cervantes et al. (2020). Evolution, 76(11), 2778-2783. https://doi.org/10.1111/evo.14610) question our hypothesis that sea-level changes during the past glaciation played a role in restricting gene flow between Pacific and Caribbean Crocodylus acutus in Panama. They argue that an error in sea-level high-stand reconstruction during the last interglacial period (118-130 ka) does not support our hypothesis. Although they are correct in our high-stand reconstruction error, overlooked the point in that we were presenting a model of restricted gene flow across the Panamanian Isthmus during low sea levels. We review the assumptions of gene demographic methods, emphasizing that we were focusing on times of genetic divergence. We expand here why gene flow between these coastal populations could have been restricted during the last glacial maximum (19-26.5 ka) and the 50,000 years preceding it when sea levels were lower than today. O'Dea et al. suggest local climates may have played larger roles than sea levels. We demonstrate that paleoclimate estimates for the past 3.3 Ma in Panama are within the bounds of extant C. acutus. The importance of Ice Age Sea-level dynamics on Neotropical species was likely profound and should be incorporated into evolutionary studies of these taxa.

The Development of the Chimaeroid Pelvic Skeleton and the Evolution of Chondrichthyan Pelvic Fins.

Pelvic girdles, fins and claspers are evolutionary novelties first recorded in jawed vertebrates. Over the course of the evolution of chondrichthyans (cartilaginous fish) two trends in the morphology of the pelvic skeleton have been suggested to have occurred. These evolutionary shifts involved both an enlargement of the metapterygium (basipterygium) and a transition of fin radial articulation from the pelvic girdle to the metapterygium. To determine how these changes in morphology have occurred it is essential to understand the development of extant taxa as this can indicate potential developmental mechanisms that may have been responsible for these changes. The study of the morphology of the appendicular skeleton across development in chondrichthyans is almost entirely restricted to the historical literature with little contemporary research. Here, we have examined the morphology and development of the pelvic skeleton of a holocephalan chondrichthyan, the elephant shark (Callorhinchus milii), through a combination of dissections, histology, and nanoCT imaging and redescribed the pelvic skeleton of Cladoselache kepleri (NHMUK PV P 9269), a stem holocephalan. To put our findings in their evolutionary context we compare them with the fossil record of chondrichthyans and the literature on pelvic development in elasmobranchs from the late 19th century. Our findings demonstrate that the pelvic skeleton of C. milii initially forms as a single mesenchymal condensation, consisting of the pelvic girdle and a series of fin rays, which fuse to form the basipterygium. The girdle and fin skeleton subsequently segment into distinct components whilst chondrifying. This confirms descriptions of the early pelvic development in Scyliorhinid sharks from the historical literature and suggests that chimaeras and elasmobranchs share common developmental patterns in their pelvic anatomy. Alterations in the location and degree of radial fusion during early development may be the mechanism responsible for changes in pelvic fin morphology over the course of the evolution of both elasmobranchs and holocephalans, which appears to be an example of parallel evolution.

Paratympanic sinuses in juvenile Alligator.

Crocodylia has an extensive epithelial pneumatic space in the middle ear, paratympanic sinus system. Although fossil and extant crocodylian paratympanic sinus systems have been studied recently using the computed tomography (CT) and three-dimensional (3D) reconstruction data, due to the soft tissue nature of the pneumatic system and presence of its surrounding soft tissue structures, some boundaries, and definitions of each extension remain ambiguous. We describe the comprehensive paratympanic sinus system in posthatched alligator using soft tissue enhanced CT data with 3D reconstructions. The data are compared to the available data to discuss the ontogenetic pattern in alligator. We introduce further divided entities of the pneumatic system based on their associated bony and soft tissue structures and epithelial membrane and clarify the pneumatic terminologies. We then re-visit the potential homology of the paratympanic sinus in Archosauria. Epithelial boundaries of the ventral portion of the pneumatic system from the histological data suggest that the dual origin of the basioccipital diverticulum derived from the tympanic sinus and basicranial diverticulum medially. The presence of the epithelial boundary and pneumatic changes in ontogeny suggests that the middle ear may function differently in developmental stages. Lastly, a morphogenetic tree is constructed to help future work of comparative developmental studies of the paratympanic sinus system between crocodiles and birds.

Docosahexaenoic acid normalizes QT interval in long QT type 2 transgenic rabbit models in a genotype-specific fashion.

AIM: Long QT syndrome (LQTS) is a cardiac channelopathy predisposing to ventricular arrhythmias and sudden cardiac death. Since current therapies often fail to prevent arrhythmic events in certain LQTS subtypes, new therapeutic strategies are needed. Docosahexaenoic acid (DHA) is a polyunsaturated fatty acid, which enhances the repolarizing IKs current. METHODS AND RESULTS: We investigated the effects of DHA in wild type (WT) and transgenic long QT Type 1 (LQT1; loss of IKs), LQT2 (loss of IKr), LQT5 (reduction of IKs), and LQT2-5 (loss of IKr and reduction of IKs) rabbits. In vivo ECGs were recorded at baseline and after 10 µM/kg DHA to assess changes in heart-rate corrected QT (QTc) and short-term variability of QT (STVQT). Ex vivo monophasic action potentials were recorded in Langendorff-perfused rabbit hearts, and action potential duration (APD75) and triangulation were assessed. Docosahexaenoic acid significantly shortened QTc in vivo only in WT and LQT2 rabbits, in which both α- and ß-subunits of IKs-conducting channels are functionally intact. In LQT2, this led to a normalization of QTc and of its short-term variability. Docosahexaenoic acid had no effect on QTc in LQT1, LQT5, and LQT2-5. Similarly, ex vivo, DHA shortened APD75 in WT and normalized it in LQT2, and additionally decreased AP triangulation in LQT2. CONCLUSIONS: Docosahexaenoic acid exerts a genotype-specific beneficial shortening/normalizing effect on QTc and APD75 and reduces pro-arrhythmia markers STVQT and AP triangulation through activation of IKs in LQT2 rabbits but has no effects if either α- or ß-subunits to IKs are functionally impaired. Docosahexaenoic acid could represent a new genotype-specific therapy in LQT2.

First fossil representative of Cerylonidae (Coleoptera: Coccinelloidea) described using X-ray micro-computed tomography, from Eocene Baltic amber.

Based on material originating from five amber collections of Eocene Baltic amber, Protostomopsis pandema gen. et sp. nov. is described and illustrated using X-ray micro-computed tomography. It is the first formally described extinct species of Cerylonidae, and the first known Palaearctic representative of the subfamily Ostomopsinae. As such, the new species extends the temporal range of the family Cerylonidae by approximately 45 Ma.

Response to Serrano and Chiappe.

In the recent study in Current Biology by Pei and colleagues1, we used two proxies - wing loading and specific lift - to reconstruct powered flight potential across the vaned feathered fossil pennaraptorans. The results recovered multiple origins of powered flight. We respectfully disagree with the criticism raised by Serrano and Chiappe2 that wing loading and specific lift, used in sequence, fail to discriminate between powered flight and gliding. We will explain this in reference to our original conservative approach.

Locally Adapted and Organically Grown Landrace and Ancient Spring Cereals-A Unique Source of Minerals in the Human Diet.

Consumer interest in local and organic produce, sustainability along the production chain and food products contributing to health, are laying the foundation for local and organic-based diets using nutrient-dense food. Here, we evaluated 25 locally adapted landrace and ancient spring cereal genotypes per location over four locations and three years, for mineral content, nutritional yield and nutrient density. The results showed a large variation in minerals content and composition in the genotypes, but also over cultivation locations, cultivation years and for genotype groups. Highest minerals content was found in oats, while highest content of Zn and Fe was found in ancient wheats. The wheat Diamant brun, the wheat landrace Öland and naked barley showed high mineral values and high content of Zn and Fe when grown in Alnarp. Nutritional yield, of the cereals evaluated here, was high related to values reported internationally but lower than those found in a comparable winter wheat material. The nutrient density was generally high; less than 350 g was needed if any of the evaluated genotype groups were to be used in the daily diet to reach the recommended value of Zn and Fe, while if the suggested Novel Nordic Diet mix was used, only 250 g were needed. A transfer from currently consumed cereals to those in the present study, along the New Nordic Diet path, showed their potential to contribute as sustainable and nutrient-rich sources in the human diet.

Effect of the Central American Isthmus on gene flow and divergence of the American crocodile (Crocodylus acutus).

The final formation of the Central American Isthmus (CAI) about 3.5 million years ago altered global ocean circulation, connected North and South America terrestrial biotas, and established the Caribbean Sea. The nature of this event creates a natural scenario to test vicariance, divergence, and speciation by allopatry. Studies have shown the effect of the CAI on marine and terrestrial species, but none have examined a large-bodied amphibious taxon. We used RAD sequencing on populations of the American crocodile (Crocodylus acutus) to study the genomic variation of C. acutus on both sides of the CAI, infer its demographic history, and measure the effect of the opening of the Panama Canal. Our results showed three genomic clusters: (1) Caribbean and the Panama Canal, (2) Pacific coast, and (3) Coiba island. The estimated divergence times between the Caribbean and Pacific populations are about 20,000 years ago, which is younger than the formation of the CAI, coinciding with the Last Glacial Maximum. We hypothesize the glacial/interglacial cycles facilitated gene flow between the Caribbean and Pacific crocodile populations after the formation of the CAI, masking any genomic divergence the CAI may have caused. There is no evidence of gene flow associated with the opening of the Panama Canal.

Adding the third dimension to studies of parallel evolution of morphology and function: An exploration based on parapatric lake-stream stickleback.

Recent methodological advances have led to a rapid expansion of evolutionary studies employing three-dimensional landmark-based geometric morphometrics (GM). GM methods generally enable researchers to capture and compare complex shape phenotypes, and to quantify their relationship to environmental gradients. However, some recent studies have shown that the common, inexpensive, and relatively rapid two-dimensional GM methods can distort important information and produce misleading results because they cannot capture variation in the depth (Z) dimension. We use micro-CT scanned threespine stickleback (Gasterosteus aculeatus Linnaeus, 1758) from six parapatric lake-stream populations on Vancouver Island, British Columbia, to test whether the loss of the depth dimension in 2D GM studies results in misleading interpretations of parallel evolution. Using joint locations described with 2D or 3D landmarks, we compare results from separate 2D and 3D shape spaces, from a combined 2D-3D shape space, and from estimates of biomechanical function. We show that, although shape is distorted enough in 2D projections to strongly influence the interpretation of morphological parallelism, estimates of biomechanical function are relatively robust to the loss of the Z dimension.

Aerodynamics Show Membrane-Winged Theropods Were a Poor Gliding Dead-end.

The bizarre scansoriopterygid theropods Yi and Ambopteryx had skin stretched between elongate fingers that form a potential membranous wing. This wing is thought to have been used in aerial locomotion, but this has never been tested. Using laser-stimulated fluorescence imaging, we re-evaluate their anatomy and perform aerodynamic calculations covering flight potential, other wing-based behaviors, and gliding capabilities. We find that Yi and Ambopteryx were likely arboreal, highly unlikely to have any form of powered flight, and had significant deficiencies in flapping-based locomotion and limited gliding abilities. Our results show that Scansoriopterygidae are not models for the early evolution of bird flight, and their structurally distinct wings differed greatly from contemporaneous paravians, supporting multiple independent origins of flight. We propose that Scansoriopterygidae represents a unique but failed flight architecture of non-avialan theropods and that the evolutionary race to capture vertebrate aerial morphospace in the Middle to Late Jurassic was dynamic and complex.

Potential for Powered Flight Neared by Most Close Avialan Relatives, but Few Crossed Its Thresholds.

Uncertainties in the phylogeny of birds (Avialae) and their closest relatives have impeded deeper understanding of early theropod flight. To help address this, we produced an updated evolutionary hypothesis through an automated analysis of the Theropod Working Group (TWiG) coelurosaurian phylogenetic data matrix. Our larger, more resolved, and better-evaluated TWiG-based hypothesis supports the grouping of dromaeosaurids + troodontids (Deinonychosauria) as the sister taxon to birds (Paraves) and the recovery of Anchiornithinae as the earliest diverging birds. Although the phylogeny will continue developing, our current results provide a pertinent opportunity to evaluate what we know about early theropod flight. With our results and available data for vaned feathered pennaraptorans, we estimate the potential for powered flight among early birds and their closest relatives. We did this by using an ancestral state reconstruction analysis calculating maximum and minimum estimates of two proxies of powered flight potential-wing loading and specific lift. These results confirm powered flight potential in early birds but its rarity among the ancestors of the closest avialan relatives (select unenlagiine and microraptorine dromaeosaurids). For the first time, we find a broad range of these ancestors neared the wing loading and specific lift thresholds indicative of powered flight potential. This suggests there was greater experimentation with wing-assisted locomotion before theropod flight evolved than previously appreciated. This study adds invaluable support for multiple origins of powered flight potential in theropods (≥3 times), which we now know was from ancestors already nearing associated thresholds, and provides a framework for its further study. VIDEO ABSTRACT.

Geology and paleontology of the Upper Cretaceous Kem Kem Group of eastern Morocco.

The geological and paleoenvironmental setting and the vertebrate taxonomy of the fossiliferous, Cenomanian-age deltaic sediments in eastern Morocco, generally referred to as the "Kem Kem beds", are reviewed. These strata are recognized here as the Kem Kem Group, which is composed of the lower Gara Sbaa and upper Douira formations. Both formations have yielded a similar fossil vertebrate assemblage of predominantly isolated elements pertaining to cartilaginous and bony fishes, turtles, crocodyliforms, pterosaurs, and dinosaurs, as well as invertebrate, plant, and trace fossils. These fossils, now in collections around the world, are reviewed and tabulated. The Kem Kem vertebrate fauna is biased toward large-bodied carnivores including at least four large-bodied non-avian theropods (an abelisaurid, Spinosaurus, Carcharodontosaurus, and Deltadromeus), several large-bodied pterosaurs, and several large crocodyliforms. No comparable modern terrestrial ecosystem exists with similar bias toward large-bodied carnivores. The Kem Kem vertebrate assemblage, currently the best documented association just prior to the onset of the Cenomanian-Turonian marine transgression, captures the taxonomic diversity of a widespread northern African fauna better than any other contemporary assemblage from elsewhere in Africa.

The fast and the frugal: Divergent locomotory strategies drive limb lengthening in theropod dinosaurs.

Limb length, cursoriality and speed have long been areas of significant interest in theropod paleobiology, since locomotory capacity, especially running ability, is critical in the pursuit of prey and to avoid becoming prey. The impact of allometry on running ability, and the limiting effect of large body size, are aspects that are traditionally overlooked. Since several different non-avian theropod lineages have each independently evolved body sizes greater than any known terrestrial carnivorous mammal, ~1000kg or more, the effect that such large mass has on movement ability and energetics is an area with significant implications for Mesozoic paleoecology. Here, using expansive datasets that incorporate several different metrics to estimate body size, limb length and running speed, we calculate the effects of allometry on running ability. We test traditional metrics used to evaluate cursoriality in non-avian theropods such as distal limb length, relative hindlimb length, and compare the energetic cost savings of relative hindlimb elongation between members of the Tyrannosauridae and more basal megacarnivores such as Allosauroidea or Ceratosauridae. We find that once the limiting effects of body size increase is incorporated there is no significant correlation to top speed between any of the commonly used metrics, including the newly suggested distal limb index (Tibia + Metatarsus/ Femur length). The data also shows a significant split between large and small bodied theropods in terms of maximizing running potential suggesting two distinct strategies for promoting limb elongation based on the organisms' size. For small and medium sized theropods increased leg length seems to correlate with a desire to increase top speed while amongst larger taxa it corresponds more closely to energetic efficiency and reducing foraging costs. We also find, using 3D volumetric mass estimates, that the Tyrannosauridae show significant cost of transport savings compared to more basal clades, indicating reduced energy expenditures during foraging and likely reduced need for hunting forays. This suggests that amongst theropods, hindlimb evolution was not dictated by one particular strategy. Amongst smaller bodied taxa the competing pressures of being both a predator and a prey item dominant while larger ones, freed from predation pressure, seek to maximize foraging ability. We also discuss the implications both for interactions amongst specific clades and Mesozoic paleobiology and paleoecological reconstructions as a whole.

The tissues and regulatory pattern of limb chondrogenesis.

Initial limb chondrogenesis offers the first differentiated tissues that resemble the mature skeletal anatomy. It is a developmental progression of three tissues. The limb begins with undifferentiated mesenchyme-1, some of which differentiates into condensations-2, and this tissue then transforms into cartilage-3. Each tissue is identified by physical characteristics of cell density, shape, and extracellular matrix composition. Tissue specific regimes of gene regulation underlie the diagnostic physical and chemical properties of these three tissues. These three tissue based regimes co-exist amid a background of other gene regulatory regimes within the same tissues and time-frame of limb development. The bio-molecular indicators of gene regulation reveal six identifiable patterns. Three of these patterns describe the unique bio-molecular indicators of each of the three tissues. A fourth pattern shares bio-molecular indicators between condensation and cartilage. Finally, a fifth pattern is composed of bio-molecular indicators that are found in undifferentiated mesenchyme prior to any condensation differentiation, then these bio-molecular indicators are upregulated in condensations and downregulated in undifferentiated mesenchyme. The undifferentiated mesenchyme that remains in between the condensations and cartilage, the interdigit, contains a unique set of bio-molecular indicators that exhibit dynamic behaviour during chondrogenesis and therefore argue for its own inclusion as a tissue in its own right and for more study into this process of differentiation.

Unique Tooth Morphology and Prismatic Enamel in Late Cretaceous Sphenodontians from Argentina.

Mammals and reptiles have evolved divergent adaptations for processing abrasive foods. Mammals have occluding, diphyodont dentitions with taller teeth (hypsodonty), more complex occlusal surfaces, continuous tooth eruption, and forms of prismatic enamel that prolong the functional life of each tooth [1, 2]. The evolution of prismatic enamel in particular was a key innovation that made individual teeth more resilient to abrasion in early mammals [2-4]. In contrast, reptiles typically have thin, non-prismatic enamel, and shearing, polyphyodont dentitions with multi-cusped or serrated tooth crowns, multiple tooth rows, rapid tooth replacement rates, or batteries made of hundreds of teeth [5-9]. However, there are rare cases where reptiles have evolved alternative solutions to cope with abrasive diets. Here, we show that the combined effects of herbivory and an ancestral loss of tooth replacement in a lineage of extinct herbivorous sphenodontians, distant relatives of the modern tuatara (Sphenodon punctatus) [10], are associated with the evolution of wear-resistant and highly complex teeth. Priosphenodon avelasi, an extinct sphenodontian from the Cretaceous of Argentina, possesses a unique cone-in-cone dentition with overlapping generations of teeth forming a densely packed tooth file. Each tooth is anchored to its predecessor via a rearrangement of dental tissues that results in a novel enamel-to-bone tooth attachment. Furthermore, the compound occlusal surfaces, thickened enamel, and the first report of prismatic enamel in a sphenodontian are convergent strategies with those in some mammals, challenging the perceived simplicity of acrodont dentitions [11-15] and showcasing the reptilian capacity to produce complex and unusual dentitions.

Polyunsaturated fatty acid-derived IKs channel activators shorten the QT interval ex-vivo and in-vivo.

AIM: We aimed to assess the ability of natural and modified polyunsaturated fatty acids (PUFAs) to shorten QT interval in ex-vivo and in-vivo guinea pig hearts. METHODS: The effect of one natural (docosahexaenoic acid [DHA]) and three modified (linoleoyl glycine [Lin-GLY], docosahexaenoyl glycine [DHA-GLY], N-arachidonoyl taurine [N-AT]) PUFAs on ventricular action potential duration (APD) and QT interval was studied in a E4031 drug-induced long QT2 model of ex-vivo guinea pig hearts. The effect of DHA-GLY on QT interval was also studied in in-vivo guinea pig hearts upon intravenous administration. The effect of modified PUFAs on IKs was studied using Xenopus laevis oocytes expressing human KCNQ1 and KCNE1. RESULTS: All tested PUFAs shortened ADP and QT interval in ex-vivo guinea pig hearts, however, with different ability in restoring baseline APD/QT interval with specific modified PUFAs being most efficacious. Despite comparable ability in activating the human KCNQ1/KCNE1 channel, Lin-GLY was not as effective in shortening APD/QT interval as DHA-GLY in ex-vivo hearts. By constructing a guinea pig-like KCNE1, we found Lin-GLY to induce less activating effect compared with DHA-GLY on human KCNQ1 co-expressed with guinea pig-like KCNE1. Docosahexaenoyl glycine was studied in more detail and was found to shorten QT interval in in-vivo guinea pig hearts. CONCLUSION: Our results show that specific PUFAs shorten QT interval in guinea pig hearts. The tendency of modified PUFAs with pronounced IKs channel activating effect to better restore QT interval suggests that modifying PUFAs to target the IKs channel is a means to improve the QT-shortening effect.

The benefits differential equations bring to limb development.

Systems biology is a large field, offering a number of advantages to a variety of biological disciplines. In limb development, differential-equation based models can provide insightful hypotheses about the gene/protein interactions and tissue differentiation events that form the core of limb development research. Differential equations are like any other communicative tool, with misuse and limitations that can come along with their advantages. Every theory should be critically analyzed to best ascertain whether they reflect the reality in biology as well they claim. Differential equation-based models have consistent features which researchers have drawn upon to aid in more realistic descriptions and hypotheses. Nine features are described that highlight these trade-offs. The advantages range from more detailed descriptions of gene interactions and their consequence and the capacity to model robustness to the incorporation of tissue size and shape. The drawbacks come with the added complication that additional genes and signaling pathways that require additional terms within the mathematical model. They also come in the translation between the mathematical terms of the model, values and matrices, to the real world of genes, proteins, and tissues that constitute limb development. A critical analysis is necessary to ensure that these models effectively expand the understanding of the origins of a diversity of limb anatomy, from evolution to teratology. This article is categorized under: Vertebrate Organogenesis > Musculoskeletal and Vascular Gene Expression and Transcriptional Hierarchies > Regulatory Mechanisms Establishment of Spatial and Temporal Patterns > Repeating Patterns and Lateral Inhibition.

New skulls and skeletons of the Cretaceous legged snake Najash, and the evolution of the modern snake body plan.

Snakes represent one of the most dramatic examples of the evolutionary versatility of the vertebrate body plan, including body elongation, limb loss, and skull kinesis. However, understanding the earliest steps toward the acquisition of these remarkable adaptations is hampered by the very limited fossil record of early snakes. Here, we shed light on the acquisition of the snake body plan using micro-computed tomography scans of the first three-dimensionally preserved skulls of the legged snake Najash and a new phylogenetic hypothesis. These findings elucidate the initial sequence of bone loss that gave origin to the modern snake skull. Morphological and molecular analyses including the new cranial data provide robust support for an extensive basal radiation of early snakes with hindlimbs and pelves, demonstrating that this intermediate morphology was not merely a transient phase between limbed and limbless body plans.

Artificial turf infill associated with systematic toxicity in an amniote vertebrate.

Artificial athletic turf containing crumb rubber (CR) from shredded tires is a growing environmental and public health concern. However, the associated health risk is unknown due to the lack of toxicity data for higher vertebrates. We evaluated the toxic effects of CR in a developing amniote vertebrate embryo. CR water leachate was administered to fertilized chicken eggs via different exposure routes, i.e., coating by dropping CR leachate on the eggshell; dipping the eggs into CR leachate; microinjecting CR leachate into the air cell or yolk. After 3 or 7 d of incubation, embryonic morphology, organ development, physiology, and molecular pathways were measured. The results showed that CR leachate injected into the yolk caused mild to severe developmental malformations, reduced growth, and specifically impaired the development of the brain and cardiovascular system, which were associated with gene dysregulation in aryl hydrocarbon receptor, stress-response, and thyroid hormone pathways. The observed systematic effects were probably due to a complex mixture of toxic chemicals leaching from CR, such as metals (e.g., Zn, Cr, Pb) and amines (e.g., benzothiazole). This study points to a need to closely examine the potential regulation of the use of CR on playgrounds and artificial fields.