The contribution of incomplete lineage sorting and introgression to the evolutionary history of the fast-evolving genus Ctenomys (Rodentia, Ctenomyidae).

Incomplete lineage sorting (ILS) and introgression have been increasingly recognized as important processes involved in biological differentiation. Both ILS and introgression result in incongruences between gene trees and species trees, consequently causing difficulties in phylogenetic reconstruction. This is particularly the case for rapid radiations, as short internodal distances and incomplete reproductive isolation increase the likelihood of both ILS and introgression. Estimation of the relative frequency of these processes requires assessments across many genomic regions. We use transcriptomics to test for introgression and estimate the frequency of ILS in a set of three closely related and geographically adjacent South American tuco-tucos species (Ctenomys), a genus comprising 64 species resulting from recent, rapid radiation. After cleaning and filtering, 5764 orthologous genes strongly support paraphyly of C. pearsoni relative to C. brasiliensis (putatively represented by the population of Villa Serrana). In line with earlier phylogenetic work, the C. pearsoni - C. brasiliensis pair is closely related to C. torquatus, whereas C. rionegrensis is more distantly related to these three nominal species. Classical Patterson's d-statistic shows significant signals of introgression from C. torquatus into C. brasiliensis. However, a 5-taxon test shows no significant results. ILS was estimated to have involved about 9% of the loci, suggesting it represents an important process in the incipient diversification of tuco-tucos.

Living on the edge: Exploring the role of coastal refugia in the Alexander Archipelago of Alaska.

Although islands are of long-standing interest to biologists, only a handful of studies have investigated the role of climatic history in shaping evolutionary diversification in high-latitude archipelagos. In this study of the Alexander Archipelago (AA) of Southeast Alaska, we address the impact of glacial cycles on geographic genetic structure for three mammals co-distributed along the North Pacific Coast. We examined variation in mitochondrial and nuclear loci for long-tailed voles (Microtus longicaudus), northwestern deermice (Peromyscus keeni), and dusky shrews (Sorex monticola), and then tested hypotheses derived from Species Distribution Models, reconstructions of paleoshorelines, and island area and isolation. In all three species, we identified paleoendemic clades that likely originated in coastal refugia, a finding consistent with other paleoendemic lineages identified in the region such as ermine. Although there is spatial concordance at the regional level for endemism, finer scale spatial and temporal patterns are less clearly defined. Demographic expansion across the region for these distinctive clades is also evident and highlights the dynamic history of Late Quaternary contraction and expansion that characterizes high-latitude species.

An association between differential expression and genetic divergence in the Patagonian olive mouse (Abrothrix olivacea).

Recent molecular studies have found striking differences between desert-adapted species and model mammals regarding water conservation. In particular, aquaporin 4, a classical gene involved in water regulation of model species, is absent or not expressed in the kidneys of desert-adapted species. To further understand the molecular response to water availability, we studied the Patagonian olive mouse Abrothrix olivacea, a species with an unusually broad ecological tolerance that exhibits a great urine concentration capability. The species is able to occupy both the arid Patagonian steppe and the Valdivian and Magellanic forests. We sampled 95 olive mouse specimens from four localities (two in the steppe and two in the forests) and analysed both phenotypic variables and transcriptomic data to investigate the response of this species to the contrasting environmental conditions. The relative size of the kidney and the ratio of urine to plasma concentrations were, as expected, negatively correlated with annual rainfall. Expression analyses uncovered nearly 3,000 genes that were differentially expressed between steppe and forest samples and indicated that this species resorts to the "classical" gene pathways for water regulation. Differential expression across biomes also involves genes that involved in immune and detoxification functions. Overall, genes that were differentially expressed showed a slight tendency to be more divergent and to display an excess of intermediate allele frequencies, relative to the remaining loci. Our results indicate that both differential expression in pathways involved in water conservation and geographical allelic variation are important in the occupation of contrasting habitats by the Patagonian olive mouse.

Molecular adaptive convergence in the α-globin gene in subterranean octodontid rodents.

Tuco-tucos (Ctenomys) and related coruros (Spalacopus) are South American subterranean rodents. An energetically demanding lifestyle within the hypoxic/hypercapnic underground atmosphere may change the selective regime on genes involved in O2 transport in blood. In addition, some species of tuco-tucos may be found at high altitude, thus facing additional reductions in changes O2 availabily. We examined sequence variation in the alpha globin subunit gene of hemoglobine in these lineages, within a robust phylogenetic context. Using different approaches (classical and Bayesian maximum likelihood (PAML/Datamonkey) and alternatives methods (TreeSAAP)) we found at least 2 sites with evidence of positive selection in the basal branch of Octodontidae, but not in tuco-tucos. These results suggest some adaptive changes associated to fossoriality, but not strictly to life underground.

Testing for the Occurrence of Selective Episodes During the Divergence of Otophysan Fishes: Insights from Mitogenomics.

How natural selection shapes biodiversity constitutes a topic of renewed interest during the last few decades. The division Otophysi comprises approximately two-thirds of freshwater fish diversity and probably underwent an extensive adaptive radiation derived from a single invasion of the supercontinent Pangaea, giving place to the evolution of the main five Otophysan lineages during a short period of time. Little is known about the factors involved in the processes that lead to lineage diversification among this group of fishes and identifying directional selection acting over protein-coding genes could offer clues about the processes acting on species diversification. The main objective of this study was to explore the otophysan mitochondrial genome evolution, in order to account for the possible signatures of selective events in this lineage, and to explore for the functional connotations of these molecular substitutions. Mainly, three different approaches were used: the "ω-based" BS-REL and MEME methods, implemented in the DATAMONKEY web server, and analysis of selection on amino acid properties, implemented in the software TreeSAAP. We found evidence of selective episodes along several branches of the evolutionary history of othophysan fishes. Analyses carried out using the BS-REL algorithm suggest episodic diversifying selection at basal branches of the otophysan lineage, which was also supported by analyses implemented in MEME and TreeSAAP. These results suggest that throughout the Siluriformes radiation, an important number of adaptive changes occurred in their mitochondrial genome. The metabolic consequences and ecological correlates of these molecular substitutions should be addressed in future studies.

Scorched mussels (Brachidontes spp., Bivalvia: Mytilidae) from the tropical and warm-temperate southwestern Atlantic: the role of the Amazon River in their speciation.

Antitropicality is a distribution pattern where closely related taxa are separated by an intertropical latitudinal gap. Two potential examples include Brachidontes darwinianus (south eastern Brazil to Uruguay), considered by some authors as a synonym of B. exustus (Gulf of Mexico and the Caribbean), and B. solisianus, distributed along the Brazilian coast with dubious records north of the intertropical zone. Using two nuclear (18S and 28S rDNA) and one mitochondrial gene (mtDNA COI), we aimed to elucidate the phylogeographic and phylogenetic relationships among the scorched mussels present in the warm-temperate region of the southwest Atlantic. We evaluated a divergence process mediated by the tropical zone over alternative phylogeographic hypotheses. Brachidontes solisianus was closely related to B. exustus I, a species with which it exhibits an antitropical distribution. Their divergence time was approximately 2.6 Ma, consistent with the intensification of Amazon River flow. Brachidontes darwinianus, an estuarine species is shown here not to be related to this B. exustus complex. We suspect ancestral forms may have dispersed from the Caribbean to the Atlantic coast via the Trans-Amazonian seaway (Miocene). The third species, B rodriguezii is presumed to have a long history in the region with related fossil forms going back to the Miocene. Although scorched mussels are very similar in appearance, their evolutionary histories are very different, involving major historical contingencies as the formation of the Amazon River, the Panama Isthmus, and the last marine transgression.

Brain size and thermoregulation during the evolution of the genus Homo.

Several hypotheses have been proposed to explain the evolution of an energetically costly brain in the genus Homo. Some of these hypotheses are based on the correlation between climatic factors and brain size recorded for this genus during the last millions of years. In this study, we propose a complementary climatic hypothesis that is based on the mechanistic connection between temperature, thermoregulation, and size of internal organs in endothermic species. We hypothesized that global cooling during the last 3.2 my may have imposed an increased energy expenditure for thermoregulation, which in the case of hominids could represent a driver for the evolution of an expanded brain, or at least, it could imply the relaxation of a negative selection pressure acting upon this costly organ. To test this idea, here we (1) assess variation in the energetic costs of thermoregulation and brain maintenance for the last 3.2 my, and (2) evaluate the relationship between Earth temperature and brain maintenance cost for the same period, taking into account the effects of body mass and fossil age. We found that: (1) the energetic cost associated with brain enlargement represents an important fraction (between 47.5% and 82.5%) of the increase in energy needed for thermoregulation; (2) fossil age is a better predictor of brain maintenance cost than Earth temperature, suggesting that (at least) another factor correlated with time was more relevant than ambient temperature in brain size evolution; and (3) there is a significant negative correlation between the energetic cost of brain and Earth temperature, even after accounting for the effect of body mass and fossil age. Thus, our results expand the current energetic framework for the study of brain size evolution in our lineage by suggesting that a fall in Earth temperature during the last millions of years may have facilitated brain enlargement.

Characterization of the kidney transcriptome of the long-haired mouse Abrothrix hirta (Rodentia, Sigmodontinae) and comparison with that of the olive mouse A. olivacea.

To understand how small mammals cope with the challenge of water homeostasis is a matter of interest for ecologists and evolutionary biologists. Here we take a step towards the understanding of the transcriptomic functional response of kidney using as a model the long-haired mouse (Abrothrix hirta) a species that distributes across Patagonian steppes and Austral temperate rainforests in Argentina and Chile. Specifically, we i) characterize the renal transcriptome of A. hirta, and ii) compare it with that-already available-of the co-generic and co-distributed A. olivacea. Renal mRNA transcripts from 16 specimens of A. hirta from natural populations were analyzed. Over 500 million Illumina paired-end reads were assembled de novo under two approaches, an individual assembly for each specimen, and a single in-silico normalized joint assembly including all reads from all specimens. The total number of annotated genes was similar with both strategies: an average of 14,956 in individual assemblies and 14,410 in the joint assembly. Overall, 15,463 distinct genes express in the kidney of A. hirta. Transcriptomes of A. hirta and A. olivacea were similar in terms of gene abundance and composition: 95.6% of the genes of A. hirta were also found in A. olivacea making their functional profiles also similar. However, differences in the transcriptome of these two species were observed in the set of highly expressed genes, in terms of private genes for each species and the functional profiles of highly expressed genes. As part of the novel transcriptome characterization, we provide distinct gene lists with their functional annotation that would constitute the basis for further research on these or any other species of the subfamily Sigmodontinae, which includes about 400 living species distributed from Tierra del Fuego to southern United States.

Characterization of the kidney transcriptome of the South American olive mouse Abrothrix olivacea.

BACKGROUND: The olive mouse Abrothrix olivacea is a cricetid rodent of the subfamily Sigmodontinae that inhabits a wide range of contrasting environments in southern South America, from aridlands to temperate rainforests. Along its distribution, it presents different geographic forms that make the olive mouse a good focal case for the study of geographical variation in response to environmental variation. We chose to characterize the kidney transcriptome because this organ has been shown to be associated with multiple physiological processes, including water reabsorption. RESULTS: Transcriptomes of thirteen kidneys from individuals from Argentina and Chile were sequenced using Illumina technology in order to obtain a kidney reference transcriptome. After combining the reads produced for each sample, we explored three assembly strategies to obtain the best reconstruction of transcripts, TrinityNorm and DigiNorm, which include its own normalization algorithms for redundant reads removal, and Multireads, which simply consist on the assembly of the joined reads. We found that Multireads strategy produces a less fragmented assembly than normalization algorithms but recovers fewer number of genes. In general, about 15000 genes were annotated, of which almost half had at least one coding sequence reconstructed at 99% of its length. We also built a list of highly expressed genes, of which several are involved in water conservation under laboratory conditions using mouse models. CONCLUSION: Based on our assembly results, Trinity's in silico normalization is the best algorithm in terms of cost-benefit returns; however, our results also indicate that normalization should be avoided if complete or nearly complete coding sequences of genes are desired. Given that this work is the first to characterize the transcriptome of any member of Sigmodontinae, a subfamily of cricetid rodents with about 400 living species, it will provide valuable resources for future ecological and evolutionary genomic analyses.

Dietary and flight energetic adaptations in a salivary gland transcriptome of an insectivorous bat.

We hypothesized that evolution of salivary gland secretory proteome has been important in adaptation to insectivory, the most common dietary strategy among Chiroptera. A submandibular salivary gland (SMG) transcriptome was sequenced for the little brown bat, Myotis lucifugus. The likely secretory proteome of 23 genes included seven (RETNLB, PSAP, CLU, APOE, LCN2, C3, CEL) related to M. lucifugus insectivorous diet and metabolism. Six of the secretory proteins probably are endocrine, whereas one (CEL) most likely is exocrine. The encoded proteins are associated with lipid hydrolysis, regulation of lipid metabolism, lipid transport, and insulin resistance. They are capable of processing exogenous lipids for flight metabolism while foraging. Salivary carboxyl ester lipase (CEL) is thought to hydrolyze insect lipophorins, which probably are absorbed across the gastric mucosa during feeding. The other six proteins are predicted either to maintain these lipids at high blood concentrations or to facilitate transport and uptake by flight muscles. Expression of these seven genes and coordinated secretion from a single organ is novel to this insectivorous bat, and apparently has evolved through instances of gene duplication, gene recruitment, and nucleotide selection. Four of the recruited genes are single-copy in the Myotis genome, whereas three have undergone duplication(s) with two of these genes exhibiting evolutionary 'bursts' of duplication resulting in multiple paralogs. Evidence for episodic directional selection was found for six of seven genes, reinforcing the conclusion that the recruited genes have important roles in adaptation to insectivory and the metabolic demands of flight. Intragenic frequencies of mobile- element-like sequences differed from frequencies in the whole M. lucifugus genome. Differences among recruited genes imply separate evolutionary trajectories and that adaptation was not a single, coordinated event.

Two mitochondrial genes under episodic positive selection in subterranean octodontoid rodents.

Tuco-tucos (Ctenomys) and related coruros (Spalacopus) are South American subterranean rodents. An energetically demanding lifestyle within the hypoxic, underground atmosphere may change the selective regime on oxidative phosphorylation. We examined whether weak and/or episodic positive directional selection affected the evolution of two mitochondrial genes (COX2, CytB), in a background of purifying selection in these lineages. We estimated rates of synonymous (dS) and non-synonymous (dN) substitutions and found: 1) significantly higher dN/dS ratio in subterranean groups relative to non-subterranean related species, and 2) two codons in each gene under episodic selection: 94 and 277 of COX2 and 269 and 307 of CytB.

Population genetic structure and social kin associations of franciscana dolphin, Pontoporia blainvillei.

We investigated population and social structure of the franciscana dolphin, Pontoporia blainvillei, an endemic and the most endangered cetacean of the southwestern Atlantic Ocean. We analyzed samples from the Rio de la Plata estuary obtained in Uruguayan waters and from the Atlantic Ocean obtained in both Uruguayan and Brazilian waters. Mitochondrial and microsatellite DNA markers were used to study differentiation between the estuary and the ocean and the association between kinship and social group structure. Although multilocus analyses suggested that franciscanas are structured into 2 subpopulations (K = 2, divergence among clusters: F(ST) = 0.06, P = 0.002; R(ST) = 0.3, P = 0.001), mitochondrial markers did not support such divergence (F(ST) = 0.02, P = 0.12; Ф(ST) = 0.06, P =0.06). However, these units are not entirely segregated geographically. Regarding social structure, some groups are composed by first-order related individuals (R ≥ 0.5, P < 0.5). Overall, the data suggest that matrilines could be the social unit in this species. We argue that the divergence found could be associated to local adaptation and social structure, resulting from either feature leading to a recent divergence or reflecting equilibrium between local differentiation and gene flow. This evidence supports considering franciscanas from the Rio de la Plata estuary a discrete management unit.

The evolution of mitochondrial genomes in subterranean caviomorph rodents: adaptation against a background of purifying selection.

South American tuco-tucos (Ctenomys) and the related coruro (Spalacopus) are two rodent lineages that have independently colonised the subterranean niche. The energetically demanding lifestyles of these species, coupled with the hypoxic atmospheres characteristic of subterranean environments, may have altered the selective regimes on genes encoding proteins related to cellular respiration. Here, we examined the molecular evolution of 13 protein-coding genes in the mitochondrial genome of seven caviomorph rodents, including these two subterranean genera and their above-ground relatives. Using maximum-likelihood and Bayesian approaches, we estimated rates of synonymous (dS) and nonsynonymous (dN) substitutions. We found a significantly higher ω ratio (dN/dS) in subterranean groups as compared to their non-subterranean counterparts in 11 of 13 genes, although no ω ratio was larger than 1. Additionally, we applied a method based on quantitative physicochemical properties to test for positive selection. Amino acid changes implicated in radical structural or functional shifts in the protein property were found to be ubiquitous across the phylogeny, but concentrated in the subterranean lineages. Convergent changes were also found between the subterranean genera used in this study and other mammals adapted to hypoxia. The results of this study suggest a link between niche shifts and weak directional (or episodic) selection at the molecular level against a background of purifying selection.

Genetic footprints of late Quaternary climate change in the diversity of Patagonian-Fueguian rodents.

Species are impacted by climate change at both ecological and evolutionary time scales. Studies in northern continents have provided abundant evidence of dramatic shifts in distributions of species subsequent to the last glacial maximum (LGM), particularly at high latitudes. However, little is known about the history of southern continents, especially at high latitudes. South America is the only continent, other than Antarctica, that extends beyond 40 degrees S. Genetic studies of a few Patagonian species have provided seemingly conflicting results, indicating either postglacial colonization from restricted glacial refugia or persistence through glacial cycles and in situ differentiation. Using mitochondrial DNA sequences of 14 species of sigmodontine rodents, a major faunal ensemble of Patagonia and Tierra del Fuego, we show that at least nine of these species bear genetic footprints of demographic expansion from single restricted sources. However, timing of demographic expansion precedes the LGM in most of these species. Four species are fragmented phylogeographically within the region. Our results indicate that (i) demographic instability in response to historical climate change has been widespread in the Patagonian-Fueguian region, and is generally more pronounced at high latitudes in both southern and northern continents; (ii) colonization from lower latitudes is an important component of current Patagonian-Fueguian diversity; but (iii) in situ differentiation has also contributed to species diversity.

Dispersal and population structure at different spatial scales in the subterranean rodent Ctenomys australis.

BACKGROUND: The population genetic structure of subterranean rodent species is strongly affected by demographic (e.g. rates of dispersal and social structure) and stochastic factors (e.g. random genetic drift among subpopulations and habitat fragmentation). In particular, gene flow estimates at different spatial scales are essential to understand genetic differentiation among populations of a species living in a highly fragmented landscape. Ctenomys australis (the sand dune tuco-tuco) is a territorial subterranean rodent that inhabits a relatively secure, permanently sealed burrow system, occurring in sand dune habitats on the coastal landscape in the south-east of Buenos Aires province, Argentina. Currently, this habitat is threatened by urban development and forestry and, therefore, the survival of this endemic species is at risk. Here, we assess population genetic structure and patterns of dispersal among individuals of this species at different spatial scales using 8 polymorphic microsatellite loci. Furthermore, we evaluate the relative importance of sex and habitat configuration in modulating the dispersal patterns at these geographical scales. RESULTS: Our results show that dispersal in C. australis is not restricted at regional spatial scales (approximately 4 km). Assignment tests revealed significant population substructure within the study area, providing support for the presence of two subpopulations from three original sampling sites. Finally, male-biased dispersal was found in the Western side of our study area, but in the Eastern side no apparent philopatric pattern was found, suggesting that in a more continuous habitat males might move longer distances than females. CONCLUSIONS: Overall, the assignment-based approaches were able to detect population substructure at fine geographical scales. Additionally, the maintenance of a significant genetic structure at regional (approximately 4 km) and small (less than 1 km) spatial scales despite apparently moderate to high levels of gene flow between local sampling sites could not be explained simply by the linear distance among them. On the whole, our results support the hypothesis that males disperse more frequently than females; however they do not provide support for strict philopatry within females.

Phylogeographical structure in the subterranean tuco-tuco Ctenomys talarum (Rodentia: Ctenomyidae): contrasting the demographic consequences of regional and habitat-specific histories.

In this work we examined the phylogeography of the South American subterranean herbivorous rodent Ctenomys talarum (Talas tuco-tuco) using mitochondrial DNA (mtDNA) control region (D-loop) sequences, and we assessed the geographical genetic structure of this species in comparison with that of subterranean Ctenomys australis, which we have shown previously to be parapatric to C. talarum and to also live in a coastal sand dune habitat. A significant apportionment of the genetic variance among regional groups indicated that putative geographical barriers, such as rivers, substantially affected the pattern of genetic structure in C. talarum. Furthermore, genetic differentiation is consistent with a simple model of isolation by distance, possibly evidencing equilibrium between gene flow and local genetic drift. In contrast, C. australis showed limited hierarchical partitioning of genetic variation and departed from an isolation-by-distance pattern. Mismatch distributions and tests of neutrality suggest contrasting histories of these two species: C. talarum appears to be characterized by demographic stability and no significant departures from neutrality, whereas C. australis has undergone a recent demographic expansion and/or departures from strict neutrality in its mtDNA.

Phylogeography of the deer mouse (Peromyscus maniculatus) provides a predictive framework for research on hantaviruses.

Phylogeographical partitioning of Sin Nombre and Monongahela viruses (hantaviruses) may reflect that of their primary rodent host, the deer mouse (Peromyscus maniculatus). Lack of a comprehensive assessment of phylogeographical variation of the host has precluded the possibility of predicting spatial limits of existing strains of these viruses or geographical regions where novel viral strains might emerge. The complete cytochrome b gene was sequenced for 206 deer mice collected from sites throughout North America to provide a foundation for future studies of spatial structure and evolution of this ubiquitous host. Bayesian analyses of these sequences partitioned deer mice into six largely allopatric lineages, some of which may represent unrecognized species. The geographical distributions of these lineages were probably shaped by Quaternary climatic events. Populations of mice were apparently restricted to refugia during glacial advances, where they experienced genetic divergence. Expansion of these populations, following climatic amelioration, brought genetically distinctive forms into contact. Occurrence of parallel changes in virus strains can now be explored in appropriate regions. In New Mexico, for example, near the location where Sin Nombre virus was first discovered, there are three genetically distinctive lineages of deer mice whose geographical ranges need to be delineated precisely. The phylogeography of P. maniculatus provides a framework for interpreting geographical variability, not only in hosts, but also in associated viral variants and disease transmission, and an opportunity to predict the potential geographical distribution of newly emerging viral strains.

Adaptive evolution of the insulin gene in caviomorph rodents.

Insulin is a conservative molecule among mammals, maintaining both its structure and function. Rodents that belong to the Suborder Hystricognathi represent an exception, having a very divergent molecule with unusual physiological properties. In this work, we analyzed the evolutionary pattern of the insulin gene in caviomorph rodents (South American hystricomorph rodents). We found that these rodents have higher rates of nonsynonymous:synonymous substitutions (d(N)/d(S)) than nonhystricomorph rodents and that values are heterogeneous inside the group. We estimated codons under positive selection, specifically the second binding site (A13 and B17) and others related with hexamerization (B18, B20, and B22). In the monomer structure, all selected sites formed a single patch around the second binding site. In the hexamer structure, these amino acids were grouped into three major patches. In this structure, contacts between B chains involved all selected sites (except B18), and between faces in the center of the molecule, all contacts were among selected sites. While there is no clear hypothesis regarding the cause of this drastic change, experimental evidence does show that this group of rodents has some peculiarities in growth function, and, whether coincidental or not, these changes appeared together with important changes in life-history traits.