Shixia Xu

Phylogenomic analysis resolves the interordinal relationships and rapid diversification of the Laurasiatherian mammals

Abstract Although great progress has been made in resolving the relationships of placental mammals, the position of several clades in Laurasiatheria remain controversial. In this study, we performed a phylogenetic analysis of 97 orthologs (46,152 bp) for 15 taxa, representing all laurasiatherian orders. Additionally, phylogenetic trees of laurasiatherian mammals with draft genome sequences were reconstructed based on 1608 exons (2,175,102 bp). Our reconstructions resolve the interordinal relationships within Laurasiatheria and corroborate the clades Scrotifera, Fereuungulata, and Cetartiodactyla. Furthermore, we tested alternative topologies within Laurasiatheria, and among alternatives for the phylogenetic position of Perissodactyla, a sister-group relationship with Cetartiodactyla receives the highest support. Thus, Pegasoferae (Perissodactyla + Carnivora + Pholidota + Chiroptera) does not appear to be a natural group. Divergence time estimates from these genes were compared with published estimates for splits within Laurasiatheria. Our estimates were similar to those of several studies and suggest that the divergences among these orders occurred within just a few million years.

Baiji genomes reveal low genetic variability and new insights into secondary aquatic adaptations

The baiji, or Yangtze River dolphin (Lipotes vexillifer), is a flagship species for the conservation of aquatic animals and ecosystems in the Yangtze River of China; however, this species has now been recognized as functionally extinct. Here we report a high-quality draft genome and three re-sequenced genomes of L. vexillifer using Illumina short-read sequencing technology. Comparative genomic analyses reveal that cetaceans have a slow molecular clock and molecular adaptations to their aquatic lifestyle. We also find a significantly lower number of heterozygous single nucleotide polymorphisms in the baiji compared to all other mammalian genomes reported thus far. A reconstruction of the demographic history of the baiji indicates that a bottleneck occurred near the end of the last deglaciation, a time coinciding with a rapid decrease in temperature and the rise of eustatic sea level.

Phylogenomic analyses and improved resolution of Cetartiodactyla

The remarkable antiquity, diversity, and significance in the ecology and evolution of Cetartiodactyla have inspired numerous attempts to resolve their phylogenetic relationships. However, previous analyses based on limited samples of nuclear genes or mitochondrial DNA sequences have generated results that were either inconsistent with one another, weakly supported, or highly sensitive to analytical conditions. Here, we present strongly supported results based upon over 1.4 Mb of an aligned DNA sequence matrix from 110 single-copy nuclear protein-coding genes of 21 Cetartiodactyla species, which represent major Cetartiodactyla lineages, and three species of Perissodactyla and Carnivora as outgroups. Phylogenetic analysis of this newly developed genomic sequence data using a codon-based model and recently developed models of the rate autocorrelation resolved the phylogenetic relationships of the major cetartiodactylan lineages and of those lineages with a high degree of confidence. Cetacea was found to nest within Artiodactyla as the sister group of Hippopotamidae, and Tylopoda was corroborated as the sole base clade of Cetartiodactyla. Within Cetacea, the monophyletic status of Odontoceti relative to Mysticeti, the basal position of Physeteroidea in Odontoceti, the non-monophyly of the river dolphins, and the sister relationship between Delphinidae and Monodontidae+Phocoenidae were strongly supported. In particular, the groups of Tursiops (bottlenose dolphins) and Stenella (spotted dolphins) were validated as unnatural groups. Additionally, a very narrow time frame of ∼3 My (million years) was found for the rapid diversification of delphinids in the late Miocene, which made it difficult to resolve the phylogenetic relationships within the Delphinidae, especially for previous studies with limited data sets. The present study provides a statistically well-supported phylogenetic framework of Cetartiodactyla, which represents an important step toward ending some of the often-heated, century-long debate on their evolution.

The loss of taste genes in cetaceans

BackgroundFive basic taste modalities, sour, sweet, bitter, salt and umami, can be distinguished by humans and are fundamental for physical and ecological adaptations in mammals. Molecular genetic studies of the receptor genes for these tastes have been conducted in terrestrial mammals; however, little is known about the evolution and adaptation of these genes in marine mammals.ResultsHere, all five basic taste modalities, sour, sweet, bitter, salt and umami, were investigated in cetaceans. The sequence characteristics and evolutionary analyses of taste receptor genes suggested that nearly all cetaceans may have lost all taste modalities except for that of salt.ConclusionsThis is the first study to comprehensively examine the five basic taste modalities in cetaceans with extensive taxa sampling. Our results suggest that cetaceans have lost four of the basic taste modalities including sour, sweet, umami, and most of the ability to sense bitter tastes. The integrity of the candidate salt taste receptor genes in all the cetaceans examined may be because of their function in Na+ reabsorption, which is key to osmoregulation and aquatic adaptation.

Positive selection at the ASPM gene coincides with brain size enlargements in cetaceans

The enlargement of cetacean brain size represents an enigmatic event in mammalian evolution, yet its genetic basis remains poorly explored. One candidate gene associated with brain size evolution is the abnormal spindle-like microcephaly associated (ASPM), as mutations in this gene cause severe reductions in the cortical size of humans. Here, we investigated the ASPM gene in representative cetacean lineages and previously published sequences from other mammals to test whether the expansion of the cetacean brain matched adaptive ASPM evolution patterns. Our analyses yielded significant evidence of positive selection on the ASPM gene during cetacean evolution, especially for the Odontoceti and Delphinoidea lineages. These molecular patterns were associated with two major events of relative brain size enlargement in odontocetes and delphinoids. It is of particular interest to find that positive selection was restricted to cetaceans and primates, two distant lineages both characterized by a massive expansion of brain size. This result is suggestive of convergent molecular evolution, although no site-specific convergence at the amino acid level was found.

Sequence variability at three MHC loci of finless porpoises (Neophocaena phocaenoides)

Major histocompatibility complex (MHC) class II DQB and DRA genes and class I gene of finless porpoises (Neophocaena phocaenoides) were investigated by single-strand conformation polymorphism and sequence analysis. The DRA, DQB, and MHC-I loci each contained 5, 14, and 34 unique sequences, respectively, and considerable sequence variation was found at the MHC-I and DQB loci. Gene duplication was manifested as three to five distinct sequences at each of the DQB and MHC-I loci from some individuals, and these sequences at each of the two loci separately clustered into four groups (cluster A, B, C, and D) based on the phylogenetic trees. Phylogenetic reconstruction revealed a trans-species pattern of evolution. Relatively high rates of non-synonymous (dN) vs synonymous (dS) substitution in the peptide-binding region (PBR) suggested balancing selection for maintaining polymorphisms at the MHC-I and DQB loci. In contrast, one single locus with little sequence variation was detected in the DRA gene, and no non-synonymous substitutions in the PBR indicated no balancing selection on this gene.

Adaptive evolution of the osmoregulation-related genes in cetaceans during secondary aquatic adaptation

BackgroundOsmoregulation was a primary challenge for cetaceans during the evolutionary transition from a terrestrial to a mainly hyperosmotic environment. Several physiological mechanisms have been suggested to maintain the water and salt balance in cetaceans, but their genetic and evolutionary bases remain poorly explored. The current study investigated the genes involved in osmoregulation in cetaceans and compared them with their counterparts in terrestrial mammals to test whether adaptive evolution occurred during secondary aquatic adaptation.ResultsThe present study analyzed the molecular evolution of 11 osmoregulation-related genes in 11 cetacean species, which represented all of the major cetacean clades. The results demonstrated positive selection acting on angiotensin converting enzyme (ACE), angiotensinogen (AGT), SLC14A2, and aquaporin 2 (AQP2). This evidence for the positive selection of AQP2 and SLC14A2 suggests that the adaptive evolution of these genes has helped to enhance the capacity for water and urea transport, thereby leading to the concentration of urine, which is an efficient mechanism for maintaining the water balance. By contrast, a series of positively selected amino acid residues identified in the ACE and AGT (two key members of the renin-angiotensin-aldosterone system, RAAS) proteins of cetaceans suggests that RAAS might have been adapted to maintain the water and salt balance in response to a hyperosmotic environment. Radical amino acid changes in positively selected sites were distributed among most internal and terminal branches of the cetacean phylogeny, which suggests the pervasively adaptive evolution of osmoregulation since the origin of cetaceans and their subsequent diversification.ConclusionsThis is the first comprehensive analysis of the molecular evolution of osmoregulation-related genes in cetaceans in response to selection pressure from a generally hyperosmotic environment. Four genes, i.e., AQP2, SLC14A2, ACE, and AGT were subject to positive selection in cetaceans, which suggests that cetaceans may have adapted to maintain their water and salt balance. This also suggests that cetaceans may have evolved an effective and complex mechanism for osmoregulation.

Adaptive evolution and functional constraint at TLR4 during the secondary aquatic adaptation and diversification of cetaceans

BackgroundCetaceans (whales, dolphins and porpoises) are a group of adapted marine mammals with an enigmatic history of transition from terrestrial to full aquatic habitat and rapid radiation in waters around the world. Throughout this evolution, the pathogen stress-response proteins must have faced challenges from the dramatic change of environmental pathogens in the completely different ecological niches cetaceans occupied. For this reason, cetaceans could be one of the most ideal candidate taxa for studying evolutionary process and associated driving mechanism of vertebrate innate immune systems such as Toll-like receptors (TLRs), which are located at the direct interface between the host and the microbial environment, act at the first line in recognizing specific conserved components of microorganisms, and translate them rapidly into a defense reaction.ResultsWe used TLR4 as an example to test whether this traditionally regarded pattern recognition receptor molecule was driven by positive selection across cetacean evolutionary history. Overall, the lineage-specific selection test showed that the dN/dS (ω) values along most (30 out of 33) examined cetartiodactylan lineages were less than 1, suggesting a common effect of functional constraint. However, some specific codons made radical changes, fell adjacent to the residues interacting with lipopolysaccharides (LPS), and showed parallel evolution between independent lineages, suggesting that TLR4 was under positive selection. Especially, strong signatures of adaptive evolution on TLR4 were identified in two periods, one corresponding to the early evolutionary transition of the terrestrial ancestors of cetaceans from land to semi-aquatic (represented by the branch leading to whale + hippo) and from semi-aquatic to full aquatic (represented by the ancestral branch leading to cetaceans) habitat, and the other to the rapid diversification and radiation of oceanic dolphins.ConclusionsThis is the first study thus far to characterize the TLR gene in cetaceans. Our data present evidences that cetacean TLR4 has undergone adaptive evolution against the background of purifying selection in response to the secondary aquatic adaptation and rapid diversification in the sea. It is suggested that microbial pathogens in different environments are important factors that promote adaptive changes at cetacean TLR4 and new functions of some amino acid sites specialized for recognizing pathogens in dramatically contrasted environments to enhance the fitness for the adaptation and survival of cetaceans.

Whale phylogeny and rapid radiation events revealed using novel retroposed elements and their flanking sequences

BackgroundA diversity of hypotheses have been proposed based on both morphological and molecular data to reveal phylogenetic relationships within the order Cetacea (dolphins, porpoises, and whales), and great progress has been made in the past two decades. However, there is still some controversy concerning relationships among certain cetacean taxa such as river dolphins and delphinoid species, which needs to be further addressed with more markers in an effort to address unresolved portions of the phylogeny.ResultsAn analysis of additional SINE insertions and SINE-flanking sequences supported the monophyly of the order Cetacea as well as Odontocete, Delphinoidea (Delphinidae + Phocoenidae + Mondontidae), and Delphinidae. A sister relationship between Delphinidae and Phocoenidae + Mondontidae was supported, and members of classical river dolphins and the genera Tursiops and Stenella were found to be paraphyletic. Estimates of divergence times revealed rapid divergences of basal Odontocete lineages in the Oligocene and Early Miocene, and a recent rapid diversification of Delphinidae in the Middle-Late Miocene and Pliocene within a narrow time frame.ConclusionsSeveral novel SINEs were found to differentiate Delphinidae from the other two families (Monodontidae and Phocoenidae), whereas the sister grouping of the latter two families with exclusion of Delphinidae was further revealed using the SINE-flanking sequences. Interestingly, some anomalous PCR amplification patterns of SINE insertions were detected, which can be explained as the result of potential ancestral SINE polymorphisms and incomplete lineage sorting. Although a few loci were potentially anomalous, this study demonstrated that the SINE-based approach is a powerful tool in phylogenetic studies. Identifying additional SINE elements that resolve the relationships in the superfamily Delphinoidea and family Delphinidae will be important steps forward in completely resolving cetacean phylogenetic relationships in the future.

High similarity at three MHC loci between the baiji and finless porpoise: trans-species or convergent evolution?

Two DRA alleles and six MHC-I alleles were identified from a group of 15 baiji (Lipotes vexillifer), the most threatened cetacean in the world. Little sequence variation was detected at the DRA locus but extensive variation at the MHC-I locus. In combination with data at the DQB locus previously reported, three MHC loci exon 2 of the baiji all revealed striking similarity with those of the finless porpoise. Especially, some identical alleles shared by both species at the MHC-I and DQB loci suggested the convergent evolution as a consequence of common adaptive solutions to similar environmental pressures in the Yangtze River. As for DRA locus, the identity alleles were shared not only by baiji and finless porpoise but by some other cetacean species of the families Phocoenidae and Delphinidae, suggesting trans-species evolution on this gene.