Unmasking microsatellite deceptiveness and debunking hybridization with SNPs in four marine copepod species of Calanus.

Interspecific hybridization events are on the rise in natural systems due to climate change disrupting species barriers. Across taxa, microsatellites have long been the molecular markers of choice to identify admixed individuals. However, with the advent of high-throughput sequencing easing the generation of genome-wide datasets, incorrect reports of hybridization resulting from microsatellite technical artefacts have been uncovered in a growing number of taxa. In the marine zooplankton genus Calanus (Copepoda), whose species are used as climate change indicators, microsatellite markers have suggested hybridization between C. finmarchicus and C. glacialis, while other nuclear markers (InDels) never detected any admixed individuals, leaving the scientific community divided. Here, for the first time, we investigated the potential for hybridization among C. finmarchicus, C. glacialis, C. helgolandicus and C. hyperboreus using two large and independent SNP datasets. These were derived firstly from a protocol of target-capture applied to 179 individuals collected from 17 sites across the North Atlantic and Arctic Oceans, including sympatric areas, and second from published RNA sequences. All SNP-based analyses were congruent in showing that Calanus species are distinct and do not appear to hybridize. We then thoroughly re-assessed the microsatellites showing hybrids, with the support of published transcriptomes, and identified technical issues plaguing eight out of 10 microsatellites, including size homoplasy, paralogy, potential for null alleles and even two primer pairs targeting the same locus. Our study illustrates how deceptive microsatellites can be when applied to the investigation of hybridization.

Characteristics of the spiny dogfish (Squalus acanthias) nuclear genome.

Sequenced shark nuclear genomes are underrepresented, with reference genomes available for only four out of nine orders so far. Here, we present the nuclear genome, with annotations, of the spiny dogfish (Squalus acanthias), a shark of interest to biomedical and conservation efforts, and the first representative of the second largest order of sharks (Squaliformes) with nuclear genome annotations available. Using Pacific Biosciences Continuous Long Read data in combination with Illumina paired-end and Hi-C sequencing, we assembled the genome de novo, followed by RNA-Seq-supported annotation. The final chromosome-level assembly is 3.7 Gb in size, has a BUSCO completeness score of 91.6%, and an error rate of less than 0.02%. Annotation predicted 33,283 gene models in the spiny dogfish's genome, of which 31,979 are functionally annotated.

Evolutionary dynamics of sex-biased gene expression in a young XY system: insights from the brown alga genus Fucus.

Sex-biased gene expression is considered to be an underlying cause of sexually dimorphic traits. Although the nature and degree of sex-biased expression have been well documented in several animal and plant systems, far less is known about the evolution of sex-biased genes in more distant eukaryotic groups. Here, we investigate sex-biased gene expression in two brown algal dioecious species, Fucus serratus and Fucus vesiculosus, where male heterogamety (XX/XY) has recently emerged. We find that in contrast to evolutionary distant plant and animal lineages, male-biased genes do not experience high turnover rates, but instead reveal remarkable conservation of bias and expression levels between the two species, suggesting their importance in sexual differentiation. Genes with consistent male bias were enriched in functions related to gamete production, along with sperm competition and include three flagellar proteins under positive selection. We present one of the first reports, outside of the animal kingdom, showing that male-biased genes display accelerated rates of coding sequence evolution compared with female-biased or unbiased genes. Our results imply that evolutionary forces affect male and female sex-biased genes differently on structural and regulatory levels, resulting in unique properties of differentially expressed transcripts during reproductive development in Fucus algae.

Insights into the species evolution of Calanus copepods in the northern seas revealed by de novo transcriptome sequencing.

Copepods of the zooplankton genus Calanus play a key role in marine ecosystems in the northern seas. Although being among the most studied organisms on Earth, due to their ecological importance, genomic resources for Calanus spp. remain scarce, mostly due to their large genome size (from 6 to 12 Gbps). As an alternative to whole-genome sequencing in Calanus spp., we sequenced and de novo assembled transcriptomes of five Calanus species: Calanus glacialis, C. hyperboreus, C. marshallae, C. pacificus, and C. helgolandicus. Functional assignment of protein families based on clusters of orthologous genes (COG) and gene ontology (GO) annotations showed analogous patterns of protein functions across species. Phylogenetic analyses using maximum likelihood (ML) of 191 protein-coding genes mined from RNA-seq data fully resolved evolutionary relationships among seven Calanus species investigated (five species sequenced for this study and two species with published datasets), with gene and site concordance factors showing that 109 out of 191 protein-coding genes support a separation between three groups: the C. finmarchicus group (including C. finmarchicus, C. glacialis, and C. marshallae), the C. helgolandicus group (including C. helgolandicus, C. sinicus, and C. pacificus) and the monophyletic C. hyperboreus group. The tree topology obtained in ML analyses was similar to a previously proposed phylogeny based on morphological criteria and cleared certain ambiguities from past studies on evolutionary relationships among Calanus species.

Population and seascape genomics of a critically endangered benthic elasmobranch, the blue skate Dipturus batis.

The blue skate (Dipturus batis) has a patchy distribution across the North-East Atlantic Ocean, largely restricted to occidental seas around the British Isles following fisheries-induced population declines and extirpations. The viability of remnant populations remains uncertain and could be impacted by continued fishing and by-catch pressure, and the projected impacts of climate change. We genotyped 503 samples of D. batis, obtained opportunistically from the widest available geographic range, across 6 350 single nucleotide polymorphisms (SNPs) using a reduced-representation sequencing approach. Genotypes were used to assess the species' contemporary population structure, estimate effective population sizes and identify putative signals of selection in relation to environmental variables using a seascape genomics approach. We identified genetic discontinuities between inshore (British Isles) and offshore (Rockall and Faroe Island) populations, with differentiation most pronounced across the deep waters of the Rockall Trough. Effective population sizes were largest in the Celtic Sea and Rockall, but low enough to be of potential conservation concern among Scottish and Faroese sites. Among the 21 candidate SNPs under positive selection was one significantly correlated with environmental variables predicted to be affected by climate change, including bottom temperature, salinity and pH. The paucity of well-annotated elasmobranch genomes precluded us from identifying a putative function for this SNP. Nevertheless, our findings suggest that climate change could inflict a strong selective force upon remnant populations of D. batis, further constraining its already-restricted habitat. Furthermore, the results provide fundamental insights on the distribution, behaviour and evolutionary biology of D. batis in the North-East Atlantic that will be useful for the establishment of conservation actions for this and other critically endangered elasmobranchs.

Adaptation of Temperate Seagrass to Arctic Light Relies on Seasonal Acclimatization of Carbon Capture and Metabolism.

Due to rising global surface temperatures, Arctic habitats are becoming thermally suitable for temperate species. Whether a temperate species can immigrate into an ice-free Arctic depends on its ability to tolerate extreme seasonal fluctuations in daylength. Thus, understanding adaptations to polar light conditions can improve the realism of models predicting poleward range expansions in response to climate change. Plant adaptations to polar light have rarely been studied and remain unknown in seagrasses. If these ecosystem engineers can migrate polewards, seagrasses will enrich biodiversity, and carbon capture potential in shallow coastal regions of the Arctic. Eelgrass (Zostera marina) is the most widely distributed seagrass in the northern hemisphere. As the only seagrass species growing as far north as 70°N, it is the most likely candidate to first immigrate into an ice-free Arctic. Here, we describe seasonal (and diurnal) changes in photosynthetic characteristics, and in genome-wide gene expression patterns under strong annual fluctuations of daylength. We compared PAM measurements and RNA-seq data between two populations at the longest and shortest day of the year: (1) a Mediterranean population exposed to moderate annual fluctuations of 10-14 h daylength and (2) an Arctic population exposed to high annual fluctuations of 0-24 h daylength. Most of the gene expression specificities of the Arctic population were found in functions of the organelles (chloroplast and mitochondrion). In winter, Arctic eelgrass conserves energy by repressing respiration and reducing photosynthetic energy fluxes. Although light-reactions, and genes involved in carbon capture and carbon storage were upregulated in summer, enzymes involved in CO2 fixation and chlorophyll-synthesis were upregulated in winter, suggesting that winter metabolism relies not only on stored energy resources but also on active use of dim light conditions. Eelgrass is unable to use excessive amounts of light during summer and demonstrates a significant reduction in photosynthetic performance under long daylengths, possibly to prevent photoinhibition constrains. Our study identified key mechanisms that allow eelgrass to survive under Arctic light conditions and paves the way for experimental research to predict whether and up to which latitude eelgrass can potentially migrate polewards in response to climate change.

The microbiome of the habitat-forming brown alga Fucus vesiculosus (Phaeophyceae) has similar cross-Atlantic structure that reflects past and present drivers1.

Latitudinal diversity gradients have provided many insights into species differentiation and community processes. In the well-studied intertidal zone, however, little is known about latitudinal diversity in microbiomes associated with habitat-forming hosts. We investigated microbiomes of Fucus vesiculosus because of deep understanding of this model system and its latitudinally large, cross-Atlantic range. Given multiple effects of photoperiod, we predicted that cross-Atlantic microbiomes of the Fucus microbiome would be similar at similar latitudes and correlate with environmental factors. We found that community structure and individual amplicon sequencing variants (ASVs) showed distinctive latitudinal distributions, but alpha diversity did not. Latitudinal differentiation was mostly driven by ASVs that were more abundant in cold temperate to subarctic (e.g., Granulosicoccus_t3260, Burkholderia/Caballeronia/Paraburkholderia_t8371) or warm temperate (Pleurocapsa_t10392) latitudes. Their latitudinal distributions correlated with different humidity, tidal heights, and air/sea temperatures, but rarely with irradiance or photoperiod. Many ASVs in potentially symbiotic genera displayed novel phylogenetic biodiversity with differential distributions among tissues and regions, including closely related ASVs with differing north-south distributions that correlated with Fucus phylogeography. An apparent southern range contraction of F. vesiculosus in the NW Atlantic on the North Carolina coast mimics that recently observed in the NE Atlantic. We suggest cross-Atlantic microbial structure of F. vesiculosus is related to a combination of past (glacial-cycle) and contemporary environmental drivers.

The Seagrass Methylome Is Associated With Variation in Photosynthetic Performance Among Clonal Shoots.

Evolutionary theory predicts that clonal organisms are more susceptible to extinction than sexually reproducing organisms, due to low genetic variation and slow rates of evolution. In agreement, conservation management considers genetic variation as the ultimate measure of a population's ability to survive over time. However, clonal plants are among the oldest living organisms on our planet. Here, we test the hypothesis that clonal seagrass meadows display epigenetic variation that complements genetic variation as a source of phenotypic variation. In a clonal meadow of the seagrass Zostera marina, we characterized DNA methylation among 42 shoots. We also sequenced the whole genome of 10 shoots to correlate methylation patterns with photosynthetic performance under exposure to and recovery from 27°C, while controlling for somatic mutations. Here, we show for the first time that clonal seagrass shoots display DNA methylation variation that is independent from underlying genetic variation, and associated with variation in photosynthetic performance under experimental conditions. It remains unknown to what degree this association could be influenced by epigenetic responses to transplantation-related stress, given that the methylomes showed a strong shift under acclimation to laboratory conditions. The lack of untreated control samples in the heat stress experiment did not allow us to distinguish methylome shifts induced by acclimation from such induced by heat stress. Notwithstanding, the co-variation in DNA methylation and photosynthetic performance may be linked via gene expression because methylation patterns varied in functionally relevant genes involved in photosynthesis, and in the repair and prevention of heat-induced protein damage. While genotypic diversity has been shown to enhance stress resilience in seagrass meadows, we suggest that epigenetic variation plays a similar role in meadows dominated by a single genotype. Consequently, conservation management of clonal plants should consider epigenetic variation as indicator of resilience and stability.

Proteomic and Transcriptomic Patterns during Lipid Remodeling in Nannochloropsis gaditana.

Nutrient limited conditions are common in natural phytoplankton communities and are often used to increase the yield of lipids from industrial microalgae cultivations. Here we studied the effects of bioavailable nitrogen (N) and phosphorus (P) deprivation on the proteome and transcriptome of the oleaginous marine microalga Nannochloropsis gaditana. Turbidostat cultures were used to selectively apply either N or P deprivation, controlling for variables including the light intensity. Global (cell-wide) changes in the proteome were measured using Tandem Mass Tag (TMT) and LC-MS/MS, whilst gene transcript expression of the same samples was quantified by Illumina RNA-sequencing. We detected 3423 proteins, where 1543 and 113 proteins showed significant changes in abundance in N and P treatments, respectively. The analysis includes the global correlation between proteomic and transcriptomic data, the regulation of subcellular proteomes in different compartments, gene/protein functional groups, and metabolic pathways. The results show that triacylglycerol (TAG) accumulation under nitrogen deprivation was associated with substantial downregulation of protein synthesis and photosynthetic activity. Oil accumulation was also accompanied by a diverse set of responses including the upregulation of diacylglycerol acyltransferase (DGAT), lipase, and lipid body associated proteins. Deprivation of phosphorus had comparatively fewer, weaker effects, some of which were linked to the remodeling of respiratory metabolism.

Novel genomic resources for shelled pteropods: a draft genome and target capture probes for Limacina bulimoides, tested for cross-species relevance.

BACKGROUND: Pteropods are planktonic gastropods that are considered as bio-indicators to monitor impacts of ocean acidification on marine ecosystems. In order to gain insight into their adaptive potential to future environmental changes, it is critical to use adequate molecular tools to delimit species and population boundaries and to assess their genetic connectivity. We developed a set of target capture probes to investigate genetic variation across their large-sized genome using a population genomics approach. Target capture is less limited by DNA amount and quality than other genome-reduced representation protocols, and has the potential for application on closely related species based on probes designed from one species. RESULTS: We generated the first draft genome of a pteropod, Limacina bulimoides, resulting in a fragmented assembly of 2.9 Gbp. Using this assembly and a transcriptome as a reference, we designed a set of 2899 genome-wide target capture probes for L. bulimoides. The set of probes includes 2812 single copy nuclear targets, the 28S rDNA sequence, ten mitochondrial genes, 35 candidate biomineralisation genes, and 41 non-coding regions. The capture reaction performed with these probes was highly efficient with 97% of the targets recovered on the focal species. A total of 137,938 single nucleotide polymorphism markers were obtained from the captured sequences across a test panel of nine individuals. The probes set was also tested on four related species: L. trochiformis, L. lesueurii, L. helicina, and Heliconoides inflatus, showing an exponential decrease in capture efficiency with increased genetic distance from the focal species. Sixty-two targets were sufficiently conserved to be recovered consistently across all five species. CONCLUSION: The target capture protocol used in this study was effective in capturing genome-wide variation in the focal species L. bulimoides, suitable for population genomic analyses, while providing insights into conserved genomic regions in related species. The present study provides new genomic resources for pteropods and supports the use of target capture-based protocols to efficiently characterise genomic variation in small non-model organisms with large genomes.

The complete mitochondrial genome of the blue skate Dipturus batis.

The complete mitochondrial genome of the blue skate Dipturus batis is described from shotgun sequencing on an Illumina next-generation sequencing platform. We report a 16,911 bp long sequence similar in size to other members of the genus, containing 13 protein-coding regions, 22 tRNA genes, 2 rRNA genes, and 2 non-coding areas. Phylogenetic analysis was performed using the complete mitochondrial genomes of 17 related species, placing D. batis within the Rajini tribe of the Rajidae family, consistent with current taxonomy. The new resource adds to a growing database of rajid mitogenomes which will help resolve phylogenetic relationships within the family.

Towards population genomics in non-model species with large genomes: a case study of the marine zooplankton Calanus finmarchicus.

Advances in next-generation sequencing technologies and the development of genome-reduced representation protocols have opened the way to genome-wide population studies in non-model species. However, species with large genomes remain challenging, hampering the development of genomic resources for a number of taxa including marine arthropods. Here, we developed a genome-reduced representation method for the ecologically important marine copepod Calanus finmarchicus (haploid genome size of 6.34 Gbp). We optimized a capture enrichment-based protocol based on 2656 single-copy genes, yielding a total of 154 087 high-quality SNPs in C. finmarchicus including 62 372 in common among the three locations tested. The set of capture probes was also successfully applied to the congeneric C. glacialis. Preliminary analyses of these markers revealed similar levels of genetic diversity between the two Calanus species, while populations of C. glacialis showed stronger genetic structure compared to C. finmarchicus. Using this powerful set of markers, we did not detect any evidence of hybridization between C. finmarchicus and C. glacialis. Finally, we propose a shortened version of our protocol, offering a promising solution for population genomics studies in non-model species with large genomes.

Decadal stability in genetic variation and structure in the intertidal seaweed Fucus serratus (Heterokontophyta: Fucaceae).

BACKGROUND: The spatial distribution of genetic diversity and structure has important implications for conservation as it reveals a species' strong and weak points with regard to stability and evolutionary capacity. Temporal genetic stability is rarely tested in marine species other than commercially important fishes, but is crucial for the utility of temporal snapshots in conservation management. High and stable diversity can help to mitigate the predicted northward range shift of seaweeds under the impact of climate change. Given the key ecological role of fucoid seaweeds along rocky shores, the positive effect of genetic diversity may reach beyond the species level to stabilize the entire intertidal ecosystem along the temperate North Atlantic. In this study, we estimated the effective population size, as well as temporal changes in genetic structure and diversity of the seaweed F. serratus using 22 microsatellite markers. Samples were taken across latitudes and a range of temperature regimes at seven locations with decadal sampling (2000 and 2010). RESULTS: Across latitudes, genetic structure and diversity remained stable over 5-10 generations. Stable small-scale structure enhanced regional diversity throughout the species' range. In accordance with its biogeographic history, effective population size and diversity peaked in the species' mid-range in Brittany (France), and declined towards its leading and trailing edge to the north and south. At the species' southern edge, multi-locus-heterozygosity displayed a strong decline from 1999 to 2010. CONCLUSION: Temporally stable genetic structure over small spatial scales is a potential driver for local adaptation and species radiation in the genus Fucus. Survival and adaptation of the low-diversity leading edge of F. serratus may be enhanced by regional gene flow and 'surfing' of favorable mutations or impaired by the accumulation of deleterious mutations. Our results have clear implications for the conservation of F. serratus at its genetically unique southern edge in Northwest Iberia, where increasing temperatures are likely the major cause for the decline not only of F. serratus, but also other intertidal and subtidal macroalgae. We expect that F. serratus will disappear from Northwest Iberia by 2100 if genetic rescue is not induced by the influx of genetic variation from Brittany.

Genetics redraws pelagic biogeography of Calanus.

Planktonic copepods of the genus Calanus play a central role in North Atlantic/Arctic marine food webs. Here, using molecular markers, we redrew the distributional ranges of Calanus species inhabiting the North Atlantic and Arctic Oceans and revealed much wider and more broadly overlapping distributions than previously described. The Arctic shelf species, C. glacialis, dominated the zooplankton assemblage of many Norwegian fjords, where only C. finmarchicus has been reported previously. In these fjords, high occurrences of the Arctic species C. hyperboreus were also found. Molecular markers revealed that the most common method of species identification, prosome length, cannot reliably discriminate the species in Norwegian fjords. Differences in degree of genetic differentiation among fjord populations of the two species suggested that C. glacialis is a more permanent resident of the fjords than C. finmarchicus We found no evidence of hybridization between the species. Our results indicate a critical need for the wider use of molecular markers to reliably identify and discriminate these morphologically similar copepod species, which serve as important indicators of climate responses.

The complete mitochondrial genome of the copepod Calanus glacialis.

Calanus glacialis, a marine planktonic copepod, is a keystone species in the Arctic Ocean. In this study, we shotgun sequenced the total DNA of one C. glacialis individual, using the NextSeq® Illumina platform, in order to determine its mitochondrial genome sequence. We successfully assembled and annotated this 20,674 bp long sequence, which included 13 protein-coding genes, 2 rRNA genes and 22 tRNA genes. Common gene-coding regions of 19 other species were used to reconstruct a phylogenetic tree, using mitogenomes of the phylogenetically closest copepods available. The new resource described here constitutes a tool of interest for better understanding the structure and dynamics of C. glacialis populations.

Dispersal similarly shapes both population genetics and community patterns in the marine realm.

Dispersal plays a key role to connect populations and, if limited, is one of the main processes to maintain and generate regional biodiversity. According to neutral theories of molecular evolution and biodiversity, dispersal limitation of propagules and population stochasticity are integral to shaping both genetic and community structure. We conducted a parallel analysis of biological connectivity at genetic and community levels in marine groups with different dispersal traits. We compiled large data sets of population genetic structure (98 benthic macroinvertebrate and 35 planktonic species) and biogeographic data (2193 benthic macroinvertebrate and 734 planktonic species). We estimated dispersal distances from population genetic data (i.e., FST vs. geographic distance) and from ß-diversity at the community level. Dispersal distances ranked the biological groups in the same order at both genetic and community levels, as predicted by organism dispersal ability and seascape connectivity: macrozoobenthic species without dispersing larvae, followed by macrozoobenthic species with dispersing larvae and plankton (phyto- and zooplankton). This ranking order is associated with constraints to the movement of macrozoobenthos within the seabed compared with the pelagic habitat. We showed that dispersal limitation similarly determines the connectivity degree of communities and populations, supporting the predictions of neutral theories in marine biodiversity patterns.

The fate of the Arctic seaweed Fucus distichus under climate change: an ecological niche modeling approach.

Rising temperatures are predicted to melt all perennial ice cover in the Arctic by the end of this century, thus opening up suitable habitat for temperate and subarctic species. Canopy-forming seaweeds provide an ideal system to predict the potential impact of climate-change on rocky-shore ecosystems, given their direct dependence on temperature and their key role in the ecological system. Our primary objective was to predict the climate-change induced range-shift of Fucus distichus, the dominant canopy-forming macroalga in the Arctic and subarctic rocky intertidal. More specifically, we asked: which Arctic/subarctic and cold-temperate shores of the northern hemisphere will display the greatest distributional change of F. distichus and how will this affect niche overlap with seaweeds from temperate regions? We used the program MAXENT to develop correlative ecological niche models with dominant range-limiting factors and 169 occurrence records. Using three climate-change scenarios, we projected habitat suitability of F. distichus - and its niche overlap with three dominant temperate macroalgae - until year 2200. Maximum sea surface temperature was identified as the most important factor in limiting the fundamental niche of F. distichus. Rising temperatures were predicted to have low impact on the species' southern distribution limits, but to shift its northern distribution limits poleward into the high Arctic. In cold-temperate to subarctic regions, new areas of niche overlap were predicted between F. distichus and intertidal macroalgae immigrating from the south. While climate-change threatens intertidal seaweeds in warm-temperate regions, seaweed meadows will likely flourish in the Arctic intertidal. Although this enriches biodiversity and opens up new seaweed-harvesting grounds, it will also trigger unpredictable changes in the structure and functioning of the Arctic intertidal ecosystem.

Variation in thermal stress response in two populations of the brown seaweed, Fucus distichus, from the Arctic and subarctic intertidal.

It is unclear whether intertidal organisms are 'preadapted' to cope with the increase of temperature and temperature variability or if they are currently at their thermal tolerance limits. To address the dichotomy, we focused on an important ecosystem engineer of the Arctic intertidal rocky shores, the seaweed Fucus distichus and investigated thermal stress responses of two populations from different temperature regimes (Svalbard and Kirkenes, Norway). Thermal stress responses at 20°C, 24°C and 28°C were assessed by measuring photosynthetic performance and expression of heat shock protein (HSP) genes (shsp, hsp90 and hsp70). We detected population-specific responses between the two populations of F. distichus, as the Svalbard population revealed a smaller decrease in photosynthesis performance but a greater activation of molecular defence mechanisms (indicated by a wider repertoire of HSP genes and their stronger upregulation) compared with the Kirkenes population. Although the temperatures used in our study exceed temperatures encountered by F. distichus at the study sites, we believe response to these temperatures may serve as a proxy for the species' potential to respond to climate-related stresses.

Biodiversity mediates top-down control in eelgrass ecosystems: a global comparative-experimental approach.

Nutrient pollution and reduced grazing each can stimulate algal blooms as shown by numerous experiments. But because experiments rarely incorporate natural variation in environmental factors and biodiversity, conditions determining the relative strength of bottom-up and top-down forcing remain unresolved. We factorially added nutrients and reduced grazing at 15 sites across the range of the marine foundation species eelgrass (Zostera marina) to quantify how top-down and bottom-up control interact with natural gradients in biodiversity and environmental forcing. Experiments confirmed modest top-down control of algae, whereas fertilisation had no general effect. Unexpectedly, grazer and algal biomass were better predicted by cross-site variation in grazer and eelgrass diversity than by global environmental gradients. Moreover, these large-scale patterns corresponded strikingly with prior small-scale experiments. Our results link global and local evidence that biodiversity and top-down control strongly influence functioning of threatened seagrass ecosystems, and suggest that biodiversity is comparably important to global change stressors.

A dated molecular phylogeny of manta and devil rays (Mobulidae) based on mitogenome and nuclear sequences.

Manta and devil rays are an iconic group of globally distributed pelagic filter feeders, yet their evolutionary history remains enigmatic. We employed next generation sequencing of mitogenomes for nine of the 11 recognized species and two outgroups; as well as additional Sanger sequencing of two mitochondrial and two nuclear genes in an extended taxon sampling set. Analysis of the mitogenome coding regions in a Maximum Likelihood and Bayesian framework provided a well-resolved phylogeny. The deepest divergences distinguished three clades with high support, one containing Manta birostris, Manta alfredi, Mobula tarapacana, Mobula japanica and Mobula mobular; one containing Mobula kuhlii, Mobula eregoodootenkee and Mobula thurstoni; and one containing Mobula munkiana, Mobula hypostoma and Mobula rochebrunei. Mobula remains paraphyletic with the inclusion of Manta, a result that is in agreement with previous studies based on molecular and morphological data. A fossil-calibrated Bayesian random local clock analysis suggests that mobulids diverged from Rhinoptera around 30 Mya. Subsequent divergences are characterized by long internodes followed by short bursts of speciation extending from an initial episode of divergence in the Early and Middle Miocene (19-17 Mya) to a second episode during the Pliocene and Pleistocene (3.6 Mya - recent). Estimates of divergence dates overlap significantly with periods of global warming, during which upwelling intensity - and related high primary productivity in upwelling regions - decreased markedly. These periods are hypothesized to have led to fragmentation and isolation of feeding regions leading to possible regional extinctions, as well as the promotion of allopatric speciation. The closely shared evolutionary history of mobulids in combination with ongoing threats from fisheries and climate change effects on upwelling and food supply, reinforces the case for greater protection of this charismatic family of pelagic filter feeders.