Selection on Visual Opsin Genes in Diurnal Neotropical Frogs and Loss of the SWS2 Opsin in Poison Frogs.

Amphibians are ideal for studying visual system evolution because their biphasic (aquatic and terrestrial) life history and ecological diversity expose them to a broad range of visual conditions. Here, we evaluate signatures of selection on visual opsin genes across Neotropical anurans and focus on three diurnal clades that are well-known for the concurrence of conspicuous colors and chemical defense (i.e., aposematism): poison frogs (Dendrobatidae), Harlequin toads (Bufonidae: Atelopus), and pumpkin toadlets (Brachycephalidae: Brachycephalus). We found evidence of positive selection on 44 amino acid sites in LWS, SWS1, SWS2, and RH1 opsin genes, of which one in LWS and two in RH1 have been previously identified as spectral tuning sites in other vertebrates. Given that anurans have mostly nocturnal habits, the patterns of selection revealed new sites that might be important in spectral tuning for frogs, potentially for adaptation to diurnal habits and for color-based intraspecific communication. Furthermore, we provide evidence that SWS2, normally expressed in rod cells in frogs and some salamanders, has likely been lost in the ancestor of Dendrobatidae, suggesting that under low-light levels, dendrobatids have inferior wavelength discrimination compared to other frogs. This loss might follow the origin of diurnal activity in dendrobatids and could have implications for their behavior. Our analyses show that assessments of opsin diversification in across taxa could expand our understanding of the role of sensory system evolution in ecological adaptation.

Sensory evolution in a cavefish radiation: patterns of neuromast distribution and associated behaviour in Sinocyclocheilus (Cypriniformes: Cyprinidae).

The genus Sinocyclocheilus, comprising a large radiation of freshwater cavefishes, are well known for their presence of regressive features (e.g. variable eye reduction). Fewer constructive features are known, such as the expansion of the lateral line system (LLS), which is involved in detecting water movements. The precise relationship between LLS expansion and cave adaptation is not well understood. Here, we examine morphology and LLS-mediated behaviour in Sinocyclocheilus species characterized by broad variation in eye size, habitat and geographical distribution. Using live-staining techniques and automated behavioural analyses, we examined 26 Sinocyclocheilus species and quantified neuromast organ number, density and asymmetry within a phylogenetic context. We then examined how these morphological features may relate to wall-following, an established cave-associated behaviour mediated by the lateral line. We show that most species demonstrated laterality (i.e. asymmetry) in neuromast organs on the head, often biased to the right. We also found that wall-following behaviour was distinctive, particularly among eyeless species. Patterns of variation in LLS appear to correlate with the degree of eye loss, as well as geographical distribution. This work reveals that constructive LLS evolution is convergent across distant cavefish taxa and may mediate asymmetric behavioural features that enable survival in stark subterranean microenvironments.

Evolving in the darkness: Phylogenomics of Sinocyclocheilus cavefishes highlights recent diversification and cryptic diversity.

Troglomorphism-any morphological adaptation enabling life to the constant darkness of caves, such as loss of pigment, reduced eyesight or blindness, over-developed tactile and olfactory organs-has long intrigued biologists. However, inferring the proximate and ultimate mechanisms driving the evolution of troglomorphism (stygomorphism) in freshwater fish requires a sound understanding of the evolutionary relationships between surface and stygomorphic lineages. We use Restriction Site Associated DNA Sequencing (RADseq) to better understand the evolution of the Sinocyclocheilus fishes of China. With a remarkable array of derived stygomorphic traits, they comprise the largest cavefish diversification in the world, emerging as a multi-species model system to study evolutionary novelty. We sequenced a total of 120 individuals throughout the Sinocyclocheilus distribution. The data comprised a total of 646,497 bp per individual, including 4378 loci and 67,983 SNPs shared across a minimum of 114 individuals at a given locus. Phylogenetic analyses using either the concatenated RAD loci (RAxML) or the SNPs under a coalescent model (SVDquartets, SNAPP) showed a high degree of congruence with similar topologies and high node support (>95 for most nodes in the phylogeny). The major clades recovered conform to a pattern previously established using Sanger-based mt-DNA sequences, with a few notable exceptions. We now recognize six major clades in this group, elevating the blind cavefish S. tianlinensis and the micro-eyed S. microphthalmus as two new distinct clades due to their deep divergence from other clades. PCA plots of the SNP data also support the recognition of six major clusters of species congruent with the identified clades in ordination space. A Bayes factor delimitation (BFD) analysis showed support for 21 species, recognizing 19 previously described species and two putative new cryptic ones. Two species whose identities were previously disputed, S. furcodorsalis and S. tianeensis, are supported here as distinct species. In addition, our multi-species calibrated tree in SNAPP suggests that the genus Sinocyclocheilus originated around 10.16 Mya, with most speciation events occurring in the last 2 Mya, likely favored by the uplift of the Qinghai-Tibetan Plateau and cave occupation induced by climate-driven aridification during this period. These results provide a firm basis for future comparative studies on the evolution of Sinocyclocheilus and its adaptations to cave life.

Phylogenomic species delimitation in microendemic frogs of the Brazilian Atlantic Forest.

The advent of next-generation sequencing allows researchers to use large-scale datasets for species delimitation analyses, yet one can envision an inflection point where the added accuracy of including more loci does not offset the increased computational burden. One alternative to including all loci could be to prioritize the analysis of loci for which there is an expectation of high informativeness. Here, we explore the issue of species delimitation and locus selection with montane species from two anuran genera that have been isolated in sky islands across the southern Brazilian Atlantic Forest: Melanophryniscus (Bufonidae) and Brachycephalus (Brachycephalidae). To delimit species, we obtained genetic data using target enrichment of ultraconserved elements from 32 populations (13 for Melanophryniscus and 19 for Brachycephalus), and we were able to create datasets that included over 800 loci with no missing data. We ranked loci according to their number of parsimony-informative sites, and we performed species delimitation analyses using BPP with the most informative 10, 20, 40, 80, 160, 320, and 640 loci. We identified three types of phylogenetic node: nodes with either consistently high or low support regardless of the number of loci or their informativeness and nodes that were initially poorly supported where support became stronger as we included more data. When viewed across all sensitivity analyses, our results suggest that the current species richness in both genera is likely underestimated. In addition, our results show the effects of different sampling strategies on species delimitation using phylogenomic datasets.

Evaluating methods for phylogenomic analyses, and a new phylogeny for a major frog clade (Hyloidea) based on 2214 loci.

Phylogenomic approaches offer a wealth of data, but a bewildering diversity of methodological choices. These choices can strongly affect the resulting topologies. Here, we explore two controversial approaches (binning genes into "supergenes" and inclusion of only rapidly evolving sites), using new data from hyloid frogs. Hyloid frogs encompass ∼53% of frog species, including true toads (Bufonidae), glassfrogs (Centrolenidae), poison frogs (Dendrobatidae), and treefrogs (Hylidae). Many hyloid families are well-established, but relationships among these families have remained difficult to resolve. We generated a dataset of ultraconserved elements (UCEs) for 50 ingroup species, including 18 of 19 hyloid families and up to 2214 loci spanning >800,000 aligned base pairs. We evaluated these two general approaches (binning, rapid sites only) based primarily on their ability to recover and strongly support well-established clades. Data were analyzed using concatenated likelihood and coalescent species-tree methods (NJst, ASTRAL). Binning strongly affected inferred relationships, whereas use of only rapidly evolving sites did not (indicating ∼87% of the data contributed little information). The optimal approaches for maximizing recovery and support of well-established clades were concatenated likelihood analysis and the use of a limited number of naive bins (statistical binning gave more problematic results). These two optimal approaches converged on similar relationships among hyloid families, and resolved them with generally strong support. The relationships found were very different from most previous estimates of hyloid phylogeny, and a new classification is proposed. The new phylogeny also suggests an intriguing biogeographical scenario, in which hyloids originated in southern South America before radiating throughout the world.

The evolution of climatic niches in squamate reptiles.

Despite the remarkable diversity found in squamate reptiles, most of their species tend to be found in warm/dry environments, suggesting that climatic requirements played a crucial role in their diversification, yet little is known about the evolution of their climatic niches. In this study, we integrate climatic information associated with the geographical distribution of 1882 squamate species and their phylogenetic relationships to investigate the tempo and mode of climatic niche evolution in squamates, both over time and among lineages. We found that changes in climatic niche dynamics were pronounced over their recent squamate evolutionary history, and we identified extensive evidence for rate heterogeneity in squamate climatic niche evolution. Most rate shifts involved accelerations, particularly over the past 50 Myr. Most squamates occupy similar regions of the climatic niche space, with only a few lineages diversifying into colder and humid climatic conditions. The changes from arid to mesic conditions in some regions of the globe may have provided opportunities for climatic niche evolution, although most lineages tended to remain near their ancestral niche. Variation in rates of climatic niche evolution seems common, particularly in response to the availability of new climatic conditions over evolutionary time.

Development of a real-time PCR assay for the detection of the golden mussel ( Limnoperna fortunei , Mytilidae) in environmental samples.

The golden mussel, Limnoperna fortunei, is among the most devastating invasive species in freshwater habitats worldwide, leading to severe environmental disturbances and economic losses. Therefore, management efforts would be greatly improved by methods that efficiently detect and quantify the abundance of the golden mussel in freshwater habitats, particularly in early stages of colonization. In this study, we describe a highly-sensitive real-time PCR assay targeting a 100-bp region of the COI mitochondrial gene of the golden mussel. The method was able to detect as little as 0.225 pg of target DNA. This assay represents an important contribution to surveillance methods, as well as to optimize field measures to contain and manage populations of the golden mussel in its introduced range.

Species delimitation, phylogeny and evolutionary demography of co-distributed, montane frogs in the southern Brazilian Atlantic Forest.

The Brazilian Atlantic Forest (BAF) is recognized as one of the world's biodiversity hotspots, with even more species per unit of area than the Amazon, however the mechanisms that led to such astonishing diversity are yet to be fully understood. In this study, we investigate the diversification of two co-distributed frog genera associated with montane areas of southern BAF: Melanophryniscus (Bufonidae) and Brachycephalus (Brachycephalidae). Species delimitation methods using mitochondrial and nuclear loci supported the existence of a remarkable number of highly endemic species in each genus, most of which occupy only one or a few adjacent mountaintops. Their timing of diversification was highly congruent, supporting recent speciation events within the past 600 thousand years. Extended Bayesian skyline plots indicate that most populations have remained relatively stable in size across the evolutionary past, with recent growth after 0.15My, suggesting that the drastic changes found in previous studies on lowland frog species were not shared by these montane taxa. These results are consistent with the existence of a montane refugium in southern BAF, allowing species persistence through the climatic shifts experienced along the BAF during the Quaternary.

Prediction of phylogeographic endemism in an environmentally complex biome.

Phylogeographic endemism, the degree to which the history of recently evolved lineages is spatially restricted, reflects fundamental evolutionary processes such as cryptic divergence, adaptation and biological responses to environmental heterogeneity. Attempts to explain the extraordinary diversity of the tropics, which often includes deep phylogeographic structure, frequently invoke interactions of climate variability across space, time and topography. To evaluate historical versus contemporary drivers of phylogeographic endemism in a tropical system, we analyse the effects of current and past climatic variation on the genetic diversity of 25 vertebrates in the Brazilian Atlantic rainforest. We identify two divergent bioclimatic domains within the forest and high turnover around the Rio Doce. Independent modelling of these domains demonstrates that endemism patterns are subject to different climatic drivers. Past climate dynamics, specifically areas of relative stability, predict phylogeographic endemism in the north. Conversely, contemporary climatic heterogeneity better explains endemism in the south. These results accord with recent speleothem and fossil pollen studies, suggesting that climatic variability through the last 250 kyr impacted the northern and the southern forests differently. Incorporating sub-regional differences in climate dynamics will enhance our ability to understand those processes shaping high phylogeographic and species endemism, in the Neotropics and beyond.

Historical biogeography of the neotropical Diaptomidae (Crustacea: Copepoda).

INTRODUCTION: Diaptomid copepods are prevalent throughout continental waters of the Neotropics, yet little is known about their biogeography. In this study we investigate the main biogeographical patterns among the neotropical freshwater diaptomid copepods using Parsimony Analysis of Endemicity (PAE) based on species records within ecoregions. In addition, we assess potential environmental correlates and limits for species richness. RESULTS: PAE was efficient in identifying general areas of endemism. Moreover, only ecoregion area showed a significant correlation with diaptomid species richness, although climatic factors were shown to provide possible upper limits to the species richness in a given ecoregion. CONCLUSION: The main patterns of endemism in neotropical freshwater diaptomid copepods are highly congruent with other freshwater taxa, suggesting a strong historical signal in determining the distribution of the family in the Neotropics.

Assessing the exposure of lion tamarins (Leontopithecus spp.) to future climate change.

Understanding how biodiversity will respond to climate change is a major challenge in conservation science. Climatic changes are likely to impose serious threats to many organisms, especially those with narrow distribution ranges, small populations and low dispersal capacity. Lion tamarins (Leontopithecus spp.) are endangered primates endemic to Brazilian Atlantic Forest (BAF), and all four living species are typical examples of these aggravating conditions. Here, we integrate ecological niche modeling and GIS-based information about BAF remnants and protected areas to estimate the exposure (i.e., the extent of climate change predicted to be experienced by a species) of current suitable habitats to climate change for 2050 and 2080, and to evaluate the efficacy of existing reserves to protect climatically suitable areas. Niche models were built using Maxent and then projected onto seven global circulation models derived from the A1B climatic scenario. According to our projections, the occurrence area of L. caissara will be little exposed to climate change. Western populations of L. chrysomelas could be potentially exposed, while climatically suitable habitats will be maintained only in part of the eastern region. Protected areas that presently harbor large populations of L. chrysopygus and L. rosalia will not retain climatic suitability by 2080. Monitoring trends of exposed populations and protecting areas predicted to hold suitable conditions should be prioritized. Given the potential exposure of key lion tamarin populations, we stress the importance of conducting additional studies to assess other aspects of their vulnerability (i.e., sensitivity to climate and adaptive capacity) and, therefore, to provide a more solid framework for future management decisions in the context of climate change.

Wall-following - Phylogenetic context of an enhanced behaviour in stygomorphic Sinocyclocheilus (Cypriniformes: Cyprinidae) cavefishes.

With 75 known species, the freshwater fish genus Sinocyclocheilus is the largest cavefish radiation in the world and shows multiple adaptations for cave-dwelling (stygomorphic adaptations), which include a range of traits such as eye degeneration (normal-eyed, micro-eyed and eyeless), depigmentation of skin, and in some species, the presence of "horns". Their behavioural adaptations to subterranean environments, however, are poorly understood. Wall-following (WF) behaviour, where an organism remains in close contact with the boundary demarcating its habitat when in the dark, is a peculiar behaviour observed in a wide range of animals and is enhanced in cave dwellers. Hence, we hypothesise that wall-following is also present in Sinocyclocheilus, possibly enhanced in eyeless species compared to eye bearing (normal-/micro-eyed species). Using 13 species representative of Sinocyclocheilus radiation and eye morphs, we designed a series of assays, based on pre-existing methods for Astyanax mexicanus behavioural experiments, to examine wall-following behaviour under three conditions. Our results indicate that eyeless species exhibit significantly enhanced intensities of WF compared to normal-eyed species, with micro-eyed forms demonstrating intermediate intensities in the WF distance. Using a mtDNA based dated phylogeny (chronogram with four clades A-D), we traced the degree of WF of these forms to outline common patterns. We show that the intensity of WF behaviour is higher in the subterranean clades compared to clades dominated by normal-eyed free-living species. We also found that eyeless species are highly sensitive to vibrations, whereas normal-eyed species are the least sensitive. Since WF behaviour is presented to some degree in all Sinocyclocheilus species, and given that these fishes evolved in the late Miocene, we identify this behaviour as being ancestral with WF enhancement related to cave occupation. Results from this diversification-scale study of cavefish behaviour suggest that enhanced wall-following behaviour may be a convergent trait across all stygomorphic lineages.

Clade density and the evolution of diversity-dependent diversification.

The assumption of an ecological limit to the number of species in a given region is frequently invoked in evolutionary studies, yet its empirical basis is remarkably meager. We explore this assumption by integrating data on geographical distributions and phylogenetic relationships of nearly six thousand terrestrial vertebrate species. In particular, we test whether sympatry with closely-related species leads to decreasing speciation rates. We introduce the concept of clade density, which is the sum of the areas of overlap between a given species and other members of its higher taxon, weighted by their phylogenetic distance. Our results showed that, regardless of the chosen taxon and uncertainty in the phylogenetic relationships between the studied species, there is no significant relationship between clade density and speciation rate. We argue that the mechanistic foundation of diversity-dependent diversification is fragile, and that a better understanding of the mechanisms driving regional species pools is sorely needed.

Mechanical demands of bite in plane head shapes of ant (Hymenoptera: Formicidae) workers.

Food processing can exert significant evolutionary pressures on the morphological evolution of animal appendages. The ant genus Pheidole displays a remarkable degree of morphological differentiation and task specialization among its workers. Notably, there is considerable variation in head shape within worker subcastes of Pheidole, which could affect the stress patterns generated by bite-related muscle contraction. In this study, we use finite element analysis (FEA) to investigate the effect of the variation in head plane shape in stress patterns, while exploring the morphospace of Pheidole worker head shapes. We hypothesize that the plane head shapes of majors are optimized for dealing with stronger bites. Furthermore, we expect that plane head shapes at the edges of each morphospace would exhibit mechanical limitations that prevent further expansion of the occupied morphospace. We vectorized five head shapes for each Pheidole worker type located at the center and edges of the corresponding morphospaces. We conducted linear static FEA to analyze the stresses generated by mandibular closing muscle contraction. Our findings indicate that plane head shapes of majors exhibit signs of optimization to deal with stronger bites. Stresses are distinctly directed along the lateral margins of the head, following the direction of muscle contraction, whereas the stresses on the plane head shapes of minors tend to concentrate around the mandibular articulations. However, the comparatively higher stress levels observed on majors' plane head shapes suggest a demand for cuticular reinforcement, like increased cuticle thickness or sculpturing pattern. Our results align with the expectations regarding the main colony tasks performed by each worker subcaste, and we find evidence of biomechanical limitations on extreme plane head shapes for majors and minors.

The evolutionary history of bees in time and space.

Bees are the most significant pollinators of flowering plants. This partnership began ca. 120 million years ago, but the uncertainty of how and when bees spread across the planet has greatly obscured investigations of this key mutualism. We present a novel analysis of bee biogeography using extensive new genomic and fossil data to demonstrate that bees originated in Western Gondwana (Africa and South America). Bees likely originated in the Early Cretaceous, shortly before the breakup of Western Gondwana, and the early evolution of any major bee lineage is associated with either the South American or African land masses. Subsequently, bees colonized northern continents via a complex history of vicariance and dispersal. The notable early absences from large landmasses, particularly in Australia and India, have important implications for understanding the assembly of local floras and diverse modes of pollination. How bees spread around the world from their hypothesized Southern Hemisphere origin parallels the histories of numerous flowering plant clades, providing an essential step to studying the evolution of angiosperm pollination syndromes in space and time.

Phylogenetic diversity and the structure of host-epiphyte interactions across the Neotropics.

Understanding the mechanisms driving community assembly has been a major focus of ecological research for nearly a century, yet little is known about these mechanisms in commensal communities, particularly with respect to their historical/evolutionary components. Here, we use a large-scale dataset of 4,440 vascular plant species to explore the relationship between the evolutionary distinctiveness (ED) (as measured by the 'species evolutionary history' (SEH)) of host species and the phylogenetic diversity (PD) of their associated epiphyte species. Although there was considerable variation across hosts and their associated epiphyte species, they were largely unrelated to host SEH. Our results mostly support the idea that the determinants of epiphyte colonization success might involve host characteristics that are unrelated to host SEH (e.g., architectural differences between hosts). While determinants of PD of epiphyte assemblages are poorly known, they do not appear to be related to the evolutionary history of host species. Instead, they might be better explained by neutral processes of colonization and extinction. However, the high level of phylogenetic signal in epiphyte PD (independent of SEH) suggests it might still be influenced by yet unrecognized evolutionary determinants. This study highlights how little is still known about the phylogenetic determinants of epiphyte communities.

Geographical range overlap networks and the macroecology of species co-occurrence.

Direct interactions among species are only possible if there is some overlap in their geographical distributions. However, despite intense focus of macroecological research on species geographical ranges, relatively little theoretical and empirical work has been done on the evolution of range overlap. In this study we explore a simple model of range overlap based on a log-normal distribution of species range sizes along a one-dimensional domain, with or without absorbing boundary conditions. In particular, we focus on the mean and variance of range overlap distributions, as well as the topology of the resulting overlap networks with respect to their degree distribution, evenness, and betweenness scores. According to the model, there is an approximately linear relationship between many aspects of the distribution of range overlaps and their underlying species distributions, such as their mean and variance. However, the expected mean number of non-zero range overlaps for a given species varied from linear to convex depending on the variance of the underlying geographical range distribution. The expected topology of range overlap networks varied substantially depending on the mean and variance in the corresponding geographical distributions, particularly in the case of the degree and closeness distributions. Finally, we test the expectations of our model against five datasets of altitudinal distributions of Neotropical birds. We found strong departures from the expectations based on our model, which could potentially result from phylogenetic niche conservatism related to altitudinal gradients in environmental conditions, or from the asymmetric colonization of mountains by species from lowlands. Potential applications of range overlap networks to a variety of ecological and evolutionary phenomena are discussed.

The role of climate and islands in species diversification and reproductive-mode evolution of Old World tree frogs.

Large diversifications of species are known to occur unevenly across space and evolutionary lineages, but the relative importance of their driving mechanisms, such as climate, ecological opportunity and key evolutionary innovations (KEI), remains poorly understood. Here, we explore the remarkable diversification of rhacophorid frogs, which represent six percent of global amphibian diversity, utilize four distinct reproductive modes, and span a climatically variable area across mainland Asia, associated continental islands, and Africa. Using a complete species-level phylogeny, we find near-constant diversification rates but a highly uneven distribution of species richness. Montane regions on islands and some mainland regions have higher phylogenetic diversity and unique assemblages of taxa; we identify these as cool-wet refugia. Starting from a centre of origin, rhacophorids reached these distant refugia by adapting to new climatic conditions ('niche evolution'-dominant), especially following the origin of KEIs such as terrestrial reproduction (in the Late Eocene) or by dispersal during periods of favourable climate ('niche conservatism'-dominant).

Alpha and beta phylogenetic diversities jointly reveal ant community assembly mechanisms along a tropical elevational gradient.

Despite the long-standing interest in the organization of ant communities across elevational gradients, few studies have incorporated the evolutionary information to understand the historical processes that underlay such patterns. Through the evaluation of phylogenetic α and ß-diversity, we analyzed the structure of leaf-litter ant communities along the Cofre de Perote mountain in Mexico and evaluated whether deterministic- (i.e., habitat filtering, interspecific competition) or stochastic-driven processes (i.e., dispersal limitation) were driving the observed patterns. Lowland and some highland sites showed phylogenetic clustering, whereas intermediate elevations and the highest site presented phylogenetic overdispersion. We infer that strong environmental constraints found at the bottom and the top elevations are favoring closely-related species to prevail at those elevations. Conversely, less stressful climatic conditions at intermediate elevations suggest interspecific interactions are more important in these environments. Total phylogenetic dissimilarity was driven by the turnover component, indicating that the turnover of ant species along the mountain is actually shifts of lineages adapted to particular locations resembling their ancestral niche. The greater phylogenetic dissimilarity between communities was related to greater temperature differences probably due to narrow thermal tolerances inherent to several ant lineages that evolved in more stable conditions. Our results suggest that the interplay between environmental filtering, interspecific competition and habitat specialization plays an important role in the assembly of leaf-litter ant communities along elevational gradients.