The evolution of lncRNA repertoires and expression patterns in tetrapods.

Only a very small fraction of long noncoding RNAs (lncRNAs) are well characterized. The evolutionary history of lncRNAs can provide insights into their functionality, but the absence of lncRNA annotations in non-model organisms has precluded comparative analyses. Here we present a large-scale evolutionary study of lncRNA repertoires and expression patterns, in 11 tetrapod species. We identify approximately 11,000 primate-specific lncRNAs and 2,500 highly conserved lncRNAs, including approximately 400 genes that are likely to have originated more than 300 million years ago. We find that lncRNAs, in particular ancient ones, are in general actively regulated and may function predominantly in embryonic development. Most lncRNAs evolve rapidly in terms of sequence and expression levels, but tissue specificities are often conserved. We compared expression patterns of homologous lncRNA and protein-coding families across tetrapods to reconstruct an evolutionarily conserved co-expression network. This network suggests potential functions for lncRNAs in fundamental processes such as spermatogenesis and synaptic transmission, but also in more specific mechanisms such as placenta development through microRNA production.

Retroviral envelope gene captures and syncytin exaptation for placentation in marsupials.

Syncytins are genes of retroviral origin captured by eutherian mammals, with a role in placentation. Here we show that some marsupials-which are the closest living relatives to eutherian mammals, although they diverged from the latter ∼190 Mya-also possess a syncytin gene. The gene identified in the South American marsupial opossum and dubbed syncytin-Opo1 has all of the characteristic features of a bona fide syncytin gene: It is fusogenic in an ex vivo cell-cell fusion assay; it is specifically expressed in the short-lived placenta at the level of the syncytial feto-maternal interface; and it is conserved in a functional state in a series of Monodelphis species. We further identify a nonfusogenic retroviral envelope gene that has been conserved for >80 My of evolution among all marsupials (including the opossum and the Australian tammar wallaby), with evidence for purifying selection and conservation of a canonical immunosuppressive domain, but with only limited expression in the placenta. This unusual captured gene, together with a third class of envelope genes from recently endogenized retroviruses-displaying strong expression in the uterine glands where retroviral particles can be detected-plausibly correspond to the different evolutionary statuses of a captured retroviral envelope gene, with only syncytin-Opo1 being the present-day bona fide syncytin active in the opossum and related species. This study would accordingly recapitulate the natural history of syncytin exaptation and evolution in a single species, and definitely extends the presence of such genes to all major placental mammalian clades.

Comparative anatomy of neonates of the three major mammalian groups (monotremes, marsupials, placentals) and implications for the ancestral mammalian neonate morphotype.

The existing different modes of reproduction in monotremes, marsupials and placentals are the main source for our current understanding of the origin and evolution of the mammalian reproduction. The reproductive strategies and, in particular, the maturity states of the neonates differ remarkably between the three groups. Monotremes, for example, are the only extant mammals that lay eggs and incubate them for the last third of their embryonic development. In contrast, marsupials and placentals are viviparous and rely on intra-uterine development of the neonates via choriovitelline (mainly marsupials) and chorioallantoic (mainly placentals) placentae. The maturity of a newborn is closely linked to the parental care strategy once the neonate is born. The varying developmental degrees of neonates are the main focus of this study. Monotremes and marsupials produce highly altricial and nearly embryonic offspring. Placental mammals always give birth to more developed newborns with the widest range from altricial to precocial. The ability of a newborn to survive and grow in the environment it was born in depends highly on the degree of maturation of vital organs at the time of birth. Here, the anatomy of four neonates of the three major extant mammalian groups is compared. The basis for this study is histological and ultrastructural serial sections of a hatchling of Ornithorhynchus anatinus (Monotremata), and neonates of Monodelphis domestica (Marsupialia), Mesocricetus auratus (altricial Placentalia) and Macroscelides proboscideus (precocial Placentalia). Special attention was given to the developmental stages of the organs skin, lung, liver and kidney, which are considered crucial for the maintenance of vital functions. The state of the organs of newborn monotremes and marsupials are found to be able to support a minimum of vital functions outside the uterus. They are sufficient to survive, but without capacities for additional energetic challenges. The organs of the altricial placental neonate are further developed, able to support the maintenance of vital functions and short-term metabolic increase. The precocial placental newborn shows the most advanced state of organ development, to allow the maintenance of vital functions, stable thermoregulation and high energetic performance. The ancestral condition of a mammalian neonate is interpreted to be similar to the state of organ development found in the newborns of marsupials and monotremes. In comparison, the newborns of altricial and precocial placentals are derived from the ancestral state to a more mature developmental degree associated with advanced organ systems.

The evolution of gene expression levels in mammalian organs.

Changes in gene expression are thought to underlie many of the phenotypic differences between species. However, large-scale analyses of gene expression evolution were until recently prevented by technological limitations. Here we report the sequencing of polyadenylated RNA from six organs across ten species that represent all major mammalian lineages (placentals, marsupials and monotremes) and birds (the evolutionary outgroup), with the goal of understanding the dynamics of mammalian transcriptome evolution. We show that the rate of gene expression evolution varies among organs, lineages and chromosomes, owing to differences in selective pressures: transcriptome change was slow in nervous tissues and rapid in testes, slower in rodents than in apes and monotremes, and rapid for the X chromosome right after its formation. Although gene expression evolution in mammals was strongly shaped by purifying selection, we identify numerous potentially selectively driven expression switches, which occurred at different rates across lineages and tissues and which probably contributed to the specific organ biology of various mammals.

Morphology and ultrastructure of the chorioallantoic placenta of the Iberian mole (Talpa occidentalis) with special reference to heterophagous areolas and the nature of interhaemal barrier.

This study provides a contribution to the reconstruction of the eulipotyphlan placental morphotype and also may help resolving a long-standing conflict about the interhaemal barrier in moles. As detailed descriptions of talpid placentation, only available for Talpa europaea and Scalopus aquaticus, led to a controversial debate on the nature of interhaemal barrier, the collection of more placental data of further mole species was strongly desired. Hence, the placentas of six gestational stages of Talpa occidentalis have been studied concerning their morphogenesis and ultrastructure with special focus on the structure of the interhaemal barrier and heterophagous regions. Generally, the mode of placentation in T. occidentalis resembles that of T. europaea, including a broad, discoid, antimesometrial, definitive chorioallantoic placenta of labyrinthine type being still villous in earlier stages. Within the labyrinth, the zona intima shows an endotheliochorial interhaemal barrier with a two-layered trophoblast. This clearly contradicts former statements on the S. aquaticus placenta made by Prasad et al. (1979), although their findings cannot exclude a totally different interpretation. Regardless, the placenta of moles represents the least invasive mode of placentation among Eulipotyphla, which otherwise have highly invasive placentas. Although the phagocytic areolas situated above uterine gland openings are heterophagous, they mainly seem to serve fetal histiotrophic nutrition, at least early in pregnancy. In later stages the number of glands and areolas decreases. This special type of additional phagocytic region is usually most common in species with noninvasive, epitheliochorial placentation, which suggests a correlation between placental invasiveness and the occurrence and type of phagocytic placental structures. The compact and invasive mode of placentation of Talpidae and all other Eulipotyphla seems to be plesiomorphic within Laurasiatheria and is always correlated with an altricial neonate.

Spatial Behavior and Habitat Use of Two Sympatric Bat Species.

Movement behavior and habitat use of the long-eared bat species Plecotus auritus and Plecotus austriacus were studied in the Havelland region in Brandenburg (Germany). Data collection included mist-netting, radiotelemetry, reconstruction of prey items, and monitoring of roosting sites. Body measurements confirm a high degree of phenotypic similarity between the two species. Total activity areas (100% Minimum Convex Polygons, MCPS) of Plecotus austriacus (2828.3 ± 1269.43 ha) were up to five-fold larger compared to Plecotus auritus (544.54 ± 295.89 ha). The activity areas of Plecotus austriacus contained up to 11 distinct core areas, and their mean total size (149.7 ± 0.07 ha) was approximately three-fold larger compared to core areas of Plecotus auritus (49.2 ± 25.6 ha). The mean distance between consecutive fixes per night was 12.72 ± 3.7 km for Plecotus austriacus and 4.23 ± 2.8 km for Plecotus auritus. While Plecotus austriacus was located most frequently over pastures (>40%) and meadows (>20%), P. auritus was located mostly within deciduous (>50%) and mixed forests (>30%) in close vicinity to its roosts. Roost site monitoring indicates that the activity of P. austriacus is delayed relative to P. auritus in spring and declined earlier in autumn. These phenological differences are probably related to the species' respective diets. Levins' measure of trophic niche breadth suggests that the prey spectrum for P. auritus is more diverse during spring (B = 2.86) and autumn (B = 2.82) compared to P. austriacus (spring: B = 1.7; autumn: B = 2.1). Our results give reason to consider these interspecific ecological variations and species-specific requirements of P. auritus and P. austriacus to develop adapted and improved conservation measures.

Transfrontier Conservation Areas and Human-Wildlife Conflict: The Case of the Namibian Component of the Kavango-Zambezi (KAZA) TFCA.

Our study deals with human-wildlife conflicts in the Namibian component of the Kavango-Zambezi Transfrontier Conservation Area (KAZA TFCA). The study reconstructs the historical occurrence of selected mammal species and adopts a socio-ecological approach to assess the impact of human dimensions in the KAZA TFCA. Our results reveal pronounced human-wildlife conflicts with considerable impacts on the livelihoods of communities. Human-wildlife conflict has the potential to become a significant contributor to the failure of the TFCA concept. Conflicts are influenced by a growing human population and large mammal species re-colonising formerly abandoned areas. Mapping the occurrence of selected mammal species over time reveals an interesting picture: although conservation initiatives have led to an increase in the population size of selected species, their occurrence is more restricted than in times of heavily decimated wildlife populations. The increasing restriction of wildlife to protected areas reduces the resilience of the ecosystem. To sustainably manage and conserve wildlife populations, a bigger picture including areas outside of the current borders of KAZA TFCA should be considered. This could support re-connecting ecologically important areas for congested populations to move to and reduces the concentration of wildlife and pressure on the land and people of the region.

Lung and metabolic development in mammals: contribution to the reconstruction of the marsupial and eutherian morphotype.

Marsupials represent only 6% of all living mammals. Marsupialia and Placentalia are distinguished mainly by their modes of reproduction. In particular, the differences in the stage of development of the neonates may be one explanation for the divergent evolutionary success. In this respect one important question is whether the survivability of the neonate depends on the degree of maturation of the respiratory system relative to the metabolic capacity at the time of birth. Therefore, this review highlights the differences in lung morphology and metabolic development of extant Marsupialia and Placentalia. The Marsupial neonate is born with a low birth weight and is highly immature. The neonatal lung is characterized by large terminal sacs, a poorly developed bronchial system and late formation of alveoli. Marsupialia have a low metabolic rate at birth and attain adult metabolic rate and thermoregulatory capacity late in postnatal development. In contrast, the eutherian neonate is born with a relative high birth weight and is always more mature than marsupial neonates. The neonatal lung has small terminal sacs, the bronchial system is well developed and the formation of alveoli begins few days after birth. Placentalia have a high metabolic rate at birth and attain adult metabolic rate and thermoregulatory capacity early in postnatal development. The differences in the developmental degree of the newborn lung between Marsupialia and Placentalia have consequences for their metabolic and thermoregulatory capacity. These differences could be advantageous for Placentalia in the changing environments in which they evolved.

Postnatal lung and metabolic development in two marsupial and four eutherian species.

Two marsupial species (Monodelphis domestica, Macropus eugenii) and four eutherian species (Mesocricetus auratus, Suncus murinus, Tupaia belangeri and Cavia aperea) were examined to compare and contrast the timing of lung and metabolic development during the postnatal maturation of the mammalian respiratory apparatus. Using light, scanning and transmission electron microscopy, the lung structural changes were correlated with indirect calorimetry to track the metabolic development. Marsupial and eutherian species followed the same pattern of mammalian lung development, but differed in the developmental pace. In the two newborn marsupial species, the lung parenchyma was at the early terminal sac stage, with large terminal air sacs, and the lung developed slowly. In contrast, the newborn eutherian species had more advanced lungs at the late terminal sac stage in altricial species (M. auratus, S. murinus) and at the alveolar stage in precocial species (T. belangeri, C. aperea). Postnatal lung development proceeded rapidly in eutherian species. The marsupial species had a low metabolic rate at birth and achieved adult metabolism late in postnatal development. In contrast, newborn eutherian species had high metabolic rates and reached adult metabolism during the first week of life. The time course of the metabolic development is thus tightly linked to the structural differentiation of the lungs and the timing of postnatal lung development. These differences in the neonatal lung structure and the timing of postnatal lung maturation between marsupial and eutherian species reflect their differing reproductive strategies.

Inner ear labyrinth anatomy of monotremes and implications for mammalian inner ear evolution.

The monophyletic clade Monotremata branches early from the rest of the mammalian crown group in the Jurassic and members of this clade retain many ancestral mammalian traits. Thus, accurate and detailed anatomical descriptions of this group can offer unique insight into the early evolutionary history of Mammalia. In this study, we examine the inner ear anatomy of two extant monotremes, Ornithorhynchus anatinus and Tachyglossus aculeatus, with the primary goals of elucidating the ancestral mammalian ear morphology and resolving inconsistencies found within previous descriptive literature. We use histological serial sections and high-resolution microcomputed tomography (µCT) for correlating soft tissue features of the vestibule and cochlea to the osseous labyrinth endocast. We found that in both monotremes the scala tympani coils to a lesser degree than scala vestibuli and scala media, although all three scalae show an apical coil inside the osseous cochlear tube. The helicotrema (conduit between scala tympani and scala vestibuli) is in subapical position, and the cochlear and lagenar ganglia and their associated nerve fibers are not enclosed by bone. In comparison, in extant therian mammals (i.e., marsupials and placentals) the helicotrema is located at the apex of the osseous cochlear canal, the three scalae coil to the same degree and the cochlear ganglion is enclosed by the primary bony lamina. Whether the lagenar ganglion is lost in therian mammals or integrated into the cochlear ganglion is still debated. The presence of a sensory lagenar macula at the apex of the membranous cochlear duct, innervated by a separate lagenar nerve and ganglion is a plesiomorphic condition of amniotes that monotremes share. A separate osseous lagenar canaliculus for the lagenar nerve, and the coiling of the distended lagenar sac at the end of the cochlear duct are autapomorphies of monotremes. Based on our findings we hypothesize that the ancestral inner ear of stem mammaliaforms is characterized by a straight or slightly curved osseous cochlear canal, a lagenar macula, lagenar nerve fibers separated from a larger bundle of cochlear nerve fibers, the presence of an organ of Corti and an intra-otic cochlear ganglion suspended by membranous connective tissue. Among the major Mesozoic clades of crown mammals, cladotherians and gondwanatherians most likely acquired a fully functioning organ of Corti but lost the sensory lagenar macula, like extant therians. However, Mesozoic spalacotherioids, multituberculates and eutriconodonts likely retained the mammaliaform condition. J. Morphol. 278:236-263, 2017. © 2016 Wiley Periodicals,Inc.

The placentation of eulipotyphla-reconstructing a morphotype of the Mammalian placenta.

Placentation determines the developmental status of the neonate, which can be considered as the most vulnerable stage in the mammalian life cycle. In this respect, the different evolutionary and ecological adaptations of marsupial and placental mammals have most likely been associated with the different reproductive strategies of the two therian clades. The morphotypes of marsupial and placental neonates, as well as the placental stem species pattern of Marsupialia, have already been reconstructed. To contribute to a better understanding of the evolution of Placentalia, a histological and ultrastructural investigation of the placenta in three representatives of Eulipotyphla, that is, core insectivores, has been carried out in this study. We studied the Musk shrew (Suncus murinus), the four-toed hedgehog (Atelerix albiventris), and the Iberian mole (Talpa occidentalis). As a result, a eulipotyphlan placental morphotype consisting of a compact and invasive placenta was reconstructed. This supports the widely accepted hypothesis that the stem lineage of Placentalia is characterized by an invasive, either endothelio- or hemochorial placenta. Evolutionary transformations toward a diffuse, noninvasive placenta occurred in the stem lineages of lower primates and cetartiodactyles and were associated with prolonged gestation and the production of few and highly precocial neonates. Compared to the choriovitelline placenta of Marsupialia, the chorioallantoic placenta of Placentalia allows for a more intimate contact and is associated with more advanced neonates.

Lung development of monotremes: evidence for the mammalian morphotype.

The reproductive strategies and the extent of development of neonates differ markedly between the three extant mammalian groups: the Monotremata, Marsupialia, and Eutheria. Monotremes and marsupials produce highly altricial offspring whereas the neonates of eutherian mammals range from altricial to precocial. The ability of the newborn mammal to leave the environment in which it developed depends highly on the degree of maturation of the cardio-respiratory system at the time of birth. The lung structure is thus a reflection of the metabolic capacity of neonates. The lung development in monotremes (Ornithorhynchus anatinus, Tachyglossus aculeatus), in one marsupial (Monodelphis domestica), and one altricial eutherian (Suncus murinus) species was examined. The results and additional data from the literature were integrated into a morphotype reconstruction of the lung structure of the mammalian neonate. The lung parenchyma of monotremes and marsupials was at the early terminal air sac stage at birth, with large terminal air sacs. The lung developed slowly. In contrast, altricial eutherian neonates had more advanced lungs at the late terminal air sac stage and postnatally, lung maturation proceeded rapidly. The mammalian lung is highly conserved in many respects between monotreme, marsupial, and eutherian species and the structural differences in the neonatal lungs can be explained mainly by different developmental rates. The lung structure of newborn marsupials and monotremes thus resembles the ancestral condition of the mammalian lung at birth, whereas the eutherian newborns have a more mature lung structure.

Evolutionary transformations of fetal membranes and reproductive strategies.

Fetal membranes (such as the chorioallantois and yolk sac) are essential to embryonic development, and have contributed importantly to the evolutionary and ecological diversity of vertebrates. Since the mid-19th century, many scientific careers have been devoted to investigations of their structure, function, and development. However, significant gaps remain in our understanding of the diversity and evolution of fetal membranes. This symposium volume focuses on the use of cladistic principles and phylogenetic relationships to reconstruct the evolutionary morphology of fetal membranes. The main goal of the present paper is to introduce the application of such methods to the evolution of vertebrate fetal membranes, as well as to provide the reader with background information on relevant concepts and terminology. Contributions within this journal issue draw upon studies of metatherian and eutherian mammals, as well as sauropsid reptiles (notably squamates). Particular attention is given to historical transformations of fetal membranes associated with the evolution of such phenomena as placentation and matrotrophy, and reproductive strategies such as viviparity. What emerges from the contributed papers is a broad sampling of contemporary research on fetal membranes, and an overview of how these membranes have evolved to support embryonic life in diverse terrestrial and intra-uterine environments.

The marsupial placenta: a phylogenetic analysis.

The structure, physiology, and endocrinology of the yolk sac placenta of different marsupial groups is compared and phylogenetically analyzed to provide information on placental characters in the marsupial stem species. We conclude that the marsupial stem species possessed a functional yolk sac placenta. Histotrophic nutrition by uterine secretion decreased during late pregnancy and at least half of the yolk sac was vascularized at the time of shell coat rupture. Due to yolk sac fusion, the larger part of the avascular, bilaminar yolk sac could not serve as a placenta at late gestation in the polyovular marsupial stem species. The bilaminar yolk sac gained a relatively greater importance for nutrition in monovular australidelphians. In macropodids a greater proportion of the yolk sac remained bilaminar at the time of shell coat rupture than in the stem species. Another derived feature of macropodids is the sustained plasma progesterone synthesis that is in turn responsible for an extended secretory phase of the uterus and a lengthened gestation. The placenta of the marsupial stem species was probably capable of metabolising histo- and hemotrophes. Recognition of pregnancy during early stages of development is a derived character of macropodids that we suggest did not occur in the marsupial stem species. However, birth and birth behaviour were apparently induced by prostaglandins in the marsupial stem species. Although the yolk sac formed the definitive placenta, it is likely that the allantois provided a supplementary placental function in the marsupial stem species, but that the role of the allantois became progressively less important during the evolution of marsupial placentation.

Ultrastructure of the placenta of the tammar wallaby, Macropus eugenii: comparison with the grey short-tailed opossum, Monodelphis domestica.

The ultrastructure of the tammar placenta was studied throughout pregnancy. The uterine epithelium grows from a columnar to an enlarged, undulating epithelium between early gestation and mid-gestation when the shell coat that surrounds the marsupial conceptus ruptures. Trophectoderm and uterine epithelium do not form syncytia, nor does invasion of the endometrium occur at any stage of pregnancy. Uterine secretion is provided to both the bilaminar and the trilaminar side of the yolk sac placenta up to birth. Fenestrations, abundant vesicles and lumenal processes of maternal capillaries, as well as deep basal folds of the uterine epithelium, suggest that there is transfer of hemotrophes adjacent to both parts of the yolk sac. In contrast, in the grey short-tailed opossum, these structures are lacking. The yolk sacs of adjacent embryos fuse to form a common yolk sac cavity, thus losing most of the bilaminar yolk sac. The bilaminar and trilaminar components of the yolk sac placenta of the tammar are less different in structure and function than those of the grey short-tailed opossum, but both types are fully functional placentas. The extended secretory phase of the tammar uterus and the maternal recognition of early pregnancy appear to be derived characters of macropodid marsupials.