Cynthia C. Steiner

Impacts of the Cretaceous Terrestrial Revolution and KPg extinction on mammal diversification.

Molecular phylogenetic analysis, calibrated with fossils, resolves the time frame of the mammalian radiation. Previous analyses of relations, divergence times, and diversification patterns among extant mammalian families have relied on supertree methods and local molecular clocks. We constructed a molecular supermatrix for mammalian families and analyzed these data with likelihood-based methods and relaxed molecular clocks. Phylogenetic analyses resulted in a robust phylogeny with better resolution than phylogenies from supertree methods. Relaxed clock analyses support the long-fuse model of diversification and highlight the importance of including multiple fossil calibrations that are spread across the tree. Molecular time trees and diversification analyses suggest important roles for the Cretaceous Terrestrial Revolution and Cretaceous-Paleogene (KPg) mass extinction in opening up ecospace that promoted interordinal and intraordinal diversification, respectively. By contrast, diversification analyses provide no support for the hypothesis concerning the delayed rise of present-day mammals during the Eocene Period.

Macroevolutionary Dynamics and Historical Biogeography of Primate Diversification Inferred from a Species Supermatrix

Phylogenetic relationships, divergence times, and patterns of biogeographic descent among primate species are both complex and contentious. Here, we generate a robust molecular phylogeny for 70 primate genera and 367 primate species based on a concatenation of 69 nuclear gene segments and ten mitochondrial gene sequences, most of which were extracted from GenBank. Relaxed clock analyses of divergence times with 14 fossil-calibrated nodes suggest that living Primates last shared a common ancestor 71–63 Ma, and that divergences within both Strepsirrhini and Haplorhini are entirely post-Cretaceous. These results are consistent with the hypothesis that the Cretaceous-Paleogene mass extinction of non-avian dinosaurs played an important role in the diversification of placental mammals. Previous queries into primate historical biogeography have suggested Africa, Asia, Europe, or North America as the ancestral area of crown primates, but were based on methods that were coopted from phylogeny reconstruction. By contrast, we analyzed our molecular phylogeny with two methods that were developed explicitly for ancestral area reconstruction, and find support for the hypothesis that the most recent common ancestor of living Primates resided in Asia. Analyses of primate macroevolutionary dynamics provide support for a diversification rate increase in the late Miocene, possibly in response to elevated global mean temperatures, and are consistent with the fossil record. By contrast, diversification analyses failed to detect evidence for rate-shift changes near the Eocene-Oligocene boundary even though the fossil record provides clear evidence for a major turnover event (“Grande Coupure”) at this time. Our results highlight the power and limitations of inferring diversification dynamics from molecular phylogenies, as well as the sensitivity of diversification analyses to different species concepts.

Evolutionary Genomics and Conservation of the Endangered Przewalski’s Horse

Przewalskis horses (PHs, Equus ferus ssp. przewalskii) were discovered in the Asian steppes in the 1870s and represent the last remaining true wild horses. PHs became extinct in the wild in the 1960s but survived in captivity, thanks to major conservation efforts. The current population is still endangered, with just 2,109 individuals, one-quarter of which are in Chinese and Mongolian reintroduction reserves [1]. These horses descend from a founding population of 12 wild-caught PHs and possibly up to four domesticated individuals [2-4]. With a stocky build, an erect mane, and stripped and short legs, they are phenotypically and behaviorally distinct from domesticated horses (DHs, Equus caballus). Here, we sequenced the complete genomes of 11 PHs, representing all founding lineages, and five historical specimens dated to 1878-1929 CE, including the Holotype. These were compared to the hitherto-most-extensive genome dataset characterized for horses, comprising 21 new genomes. We found that loci showing the most genetic differentiation with DHs were enriched in genes involved in metabolism, cardiac disorders, muscle contraction, reproduction, behavior, and signaling pathways. We also show that DH and PH populations split ∼45,000 years ago and have remained connected by gene-flow thereafter. Finally, we monitor the genomic impact of ∼110 years of captivity, revealing reduced heterozygosity, increased inbreeding, and variable introgression of domestic alleles, ranging from non-detectable to as much as 31.1%. This, together with the identification of ancestry informative markers and corrections to the International Studbook, establishes a framework for evaluating the persistence of genetic variation in future reintroduced populations.

Conservation genomics of threatened animal species.

The genomics era has opened up exciting possibilities in the field of conservation biology by enabling genomic analyses of threatened species that previously were limited to model organisms. Next-generation sequencing (NGS) and the collection of genome-wide data allow for more robust studies of the demographic history of populations and adaptive variation associated with fitness and local adaptation. Genomic analyses can also advance management efforts for threatened wild and captive populations by identifying loci contributing to inbreeding depression and disease susceptibility, and predicting fitness consequences of introgression. However, the development of genomic tools in wild species still carries multiple challenges, particularly those associated with computational and sampling constraints. This review provides an overview of the most significant applications of NGS and the implications and limitations of genomic studies in conservation.

Characterization of Prdm9 in Equids and Sterility in Mules

Prdm9 (Meisetz) is the first speciation gene discovered in vertebrates conferring reproductive isolation. This locus encodes a meiosis-specific histone H3 methyltransferase that specifies meiotic recombination hotspots during gametogenesis. Allelic differences in Prdm9, characterized for a variable number of zinc finger (ZF) domains, have been associated with hybrid sterility in male house mice via spermatogenic failure at the pachytene stage. The mule, a classic example of hybrid sterility in mammals also exhibits a similar spermatogenesis breakdown, making Prdm9 an interesting candidate to evaluate in equine hybrids. In this study, we characterized the Prdm9 gene in all species of equids by analyzing sequence variation of the ZF domains and estimating positive selection. We also evaluated the role of Prdm9 in hybrid sterility by assessing allelic differences of ZF domains in equine hybrids. We found remarkable variation in the sequence and number of ZF domains among equid species, ranging from five domains in the Tibetan kiang and Asiatic wild ass, to 14 in the Grevy’s zebra. Positive selection was detected in all species at amino acid sites known to be associated with DNA-binding specificity of ZF domains in mice and humans. Equine hybrids, in particular a quartet pedigree composed of a fertile mule showed a mosaic of sequences and number of ZF domains suggesting that Prdm9 variation does not seem by itself to contribute to equine hybrid sterility.

Molecular phylogeny of extant equids and effects of ancestral polymorphism in resolving species-level phylogenies

Short divergence times and processes such as incomplete lineage sorting and species hybridization are known to hinder the inference of species-level phylogenies due to the lack of sufficient informative genetic variation or the presence of shared but incongruent polymorphism among taxa. Extant equids (horses, zebras, and asses) are an example of a recently evolved group of mammals with an unresolved phylogeny, despite a large number of molecular studies. Previous surveys have proposed trees with rather poorly supported nodes, and the bias caused by genetic introgression or ancestral polymorphism has not been assessed. Here we studied the phylogenetic relationships of all extant species of Equidae by analyzing 22 partial mitochondrial and nuclear genes using maximum likelihood and Bayesian inferences that account for heterogeneous gene histories. We also examined genetic signatures of lineage sorting and/or genetic introgression in zebras by evaluating patterns of intraspecific genetic variation. Our study improved the resolution and support of the Equus phylogeny and in particular the controversial positions of the African wild ass (E. asinus) and mountain zebra (E. zebra): the African wild ass is placed as a sister species of the Asiatic asses and the mountain zebra as the sister taxon of Grevys and Burchells zebras. A shared polymorphism (indel) detected among zebra species in the Estrogen receptor 1 gene was likely due to incomplete lineage sorting and not genetic introgression as also indicated by other mitochondrial (Cytochrome b) and nuclear (Y chromosome and microsatellites) markers. Ancestral polymorphism in equids might have contributed to the long-standing lack of clarity in the phylogeny of this highly threatened group of mammals.

Molecular characterization and mitochondrial sequence variation in two sympatric species of Proechimys (Rodentia: Echimyidae) in French Guiana

Variations in mitochondrial DNA characters were used to characterize two morphologically similar and sympatric species of Neotropical terrestrial rodents of the genus Proechimys (Mammalia: Echimyidae). We sequenced both cytochrome b (1140pb) and part of the control region (445pb) from four individuals of P. cuvieri and five of P. cayennensis from French Guiana, which allowed us to depict intra- and inter-specific patterns of variation. The phylogenetic relationships between the nine sequences evidence the monophyly of each species, and illustrate that more polymorphism might exist within P. cuvieri than within P. cayennensis. By developing species-specific primers to amplify a fragment of the cytochrome b gene, we were able to identify 50 individuals of Proechimys spp. caught in two localities of French Guiana. In both sites of primary rainforests, we showed that the two species live in syntopy, and this observation emphasizes the need to document ecological differences which should exist in order to diminish inter-specific competition.

MITOCHONDRIAL DIVERSITY AND MORPHOLOGICAL VARIATION OF MARMOSA MURINA(DIDELPHIDAE) IN FRENCH GUIANA

Abstract The murine mouse opossum Marmosa murina is a widely distributed species in tropical South America. In northeastern South America, authors propose the recognition of several subspecies, of which 2 might be found in French Guiana: a paler taxon (M. murina murina) inhabiting the narrow coastal strip and a darker one (M. m. muscula) occurring in the inner rainforests. We present here results of molecular analyses for 2 mitochondrial fragments (cytochrome-b gene and control region) and morphological examination of 3 external and 15 cranial measurements. These data support the presence of only 1 subspecies (M. m. murina) in French Guiana based on the absence of genetic structure and morphological divergence. Nevertheless, it appears that M. murina shows substantial morphometric variation across the Guianan shield with respect to the dental measurement M1–M4. This suggests that M. murina could be a differentiated taxon in the Guianas, likely comprising more than one biological entity.

Species identification and chromosome variation of captive two-toed sloths.

Two-toed sloth species, Linnaeuss and Hoffmmans, are frequent residents of zoo collections in North America. However, species identification has always been problematic because of their large overlap in external morphology, which represents an obstacle to the captive breeding program. We describe here a PCR-based technique that allows species identification of two-toed sloths without requiring sequencing, by using a mitochondrial marker (COI gene) and restriction enzyme assay. We also report intra- and inter-specific patterns of chromosome variation in captive two-toed sloths. Molecularly, we identified 22 samples of Linnaeuss and Hoffmmans two-toed sloths corresponding to 14 and 8 individuals, respectively. One animal was identified as a hybrid using the nuclear gene Enam having alleles derived from both species. The chromosome number in Hoffmans two-toed sloths showed low variation ranging only between 50 and 51. In contrast, Linnaeuss two-toed sloths appeared to vary widely, with diploid numbers ranging from 53 to 67, suggesting distinct geographic groups. The species identification method presented here represents a low-cost easy-to-use tool that will help to improve management of the captive population of two-toed sloths.

Evaluating recovery potential of the northern white rhinoceros from cryopreserved somatic cells

The critically endangered northern white rhinoceros is believed to be extinct in the wild, with the recent death of the last male leaving only two remaining individuals in captivity. Its extinction would appear inevitable, but the development of advanced cell and reproductive technologies such as cloning by nuclear transfer and the artificial production of gametes via stem cells differentiation offer a second chance for its survival. In this work, we analyzed genome-wide levels of genetic diversity, inbreeding, population history, and demography of the white rhinoceros sequenced from cryopreserved somatic cells, with the goal of informing how genetically valuable individuals could be used in future efforts toward the genetic rescue of the northern white rhinoceros. We present the first sequenced genomes of the northern white rhinoceros, which show relatively high levels of heterozygosity and an average genetic divergence of 0.1% compared with the southern subspecies. The two white rhinoceros subspecies appear to be closely related, with low genetic admixture and a divergent time <80,000 yr ago. Inbreeding, as measured by runs of homozygosity, appears slightly higher in the southern than the northern white rhinoceros. This work demonstrates the value of the northern white rhinoceros cryopreserved genetic material as a potential gene pool for saving this subspecies from extinction.