Resultados de investigación

[Usage Notes] This files contains all DNA and topoclimate data from the manuscript. The file README contains additional data for each individual., Geographic isolation substantially contributes to species endemism on oceanic islands when speciation involves the colonisation of a new island. However, less is understood about the drivers of speciation within islands. What is lacking is a general understanding of the geographic scale of gene flow limitation within islands, and thus the geographic scale and drivers of geographical speciation within insular contexts. Using a community of beetle species, we show that when dispersal ability and climate tolerance are restricted, microclimatic variation over distances of only a few kilometres can maintain strong geographic isolation and drive incipient speciation. Further to this, we demonstrate congruent diversification with gene flow across species, mediated by Quaternary climate oscillations that have facilitated a dynamic of isolation and secondary contact. The unprecedented scale of parallel species responses to a common environmental driver for evolutionary change has profound consequences for understanding past and future species responses to climate variation., Peer reviewed

[Methods] Using a sample of Atlantic rainforest avian frugivores we analyze fruit effectiveness as a nutritional food resource for seed dispersing birds, i.e., Resource-Provisioning Effectiveness (RPE), an overlooked aspect of seed dispersal effectiveness. We found ample variation in RPE among plant species, given not just variation in fruit design and nutrient content but also in fruit availability, accessibility and other features that determine how rewarding a fruit resource is for the frugivore. Yet fruit species show consistent RPE values despite highly variable outcomes of interactions with different avian frugivores. The rationale for our study includes four steps: 1) selecting a representative sample of frugivore species illustrating both the full range of body mass and phylogenetic diversity of the Brazilian Southeastern Atlantic forest frugivores; 2) compiling a literature dataset on fruit foraging and usage data for these frugivore species; 3) analyze variation in RPE across selected plant species by building effectiveness landscapes from the frugivore’s perspective; and 4) experimentally assess RPE subcomponents (i.e. fruit pulp digestibility) that refine the estimates of RPE variation among plant species. To estimate the RPE for fruit species consumed by each frugivore species we have defined its two components: the rate at which the fruits are ingested/handled by the animal (feeding rate) as the quantity component, and the energy gained from eating the fruit (fruit energy) as the quality component. The feeding rate is calculated by multiplying the bird visitation frequency to the plant by the number of fruits consumed in each visit; from a frugivore perspective these subcomponents of RPE-quantity illustrate potential encounter rates with the fruit resource (visitation) and potential handling/ingestion rates once the fruits are encountered (fruits/visit). Fruit-pulp energy yield is assessed using the specific energy (KJ/g) multiplied by the fruit pulp dry mass. Since the nutrient proportional contents of the fruit pulp are calculated based on its dry mass, we use this value as the profitable fruit mass per fruit (Herrera 1981). Specific energy was quantified using the following energy conversion factors for fruits (FAO, 2002): 14.1 KJ/g for proteins, 35 KJ/g for lipids and 15.1 KJ/g for carbohydrates. Ultimately, by multiplying the two components we obtained the total effect value for each specific interaction (RPE; Schupp, 1993; Schupp, Jordano, & Gómez, 2017): RPE[g energy assimilated] = [no fruits consumed/obs time (h)] × [g energy accumulated/fruit consumed] (1) Note that the RPE estimate for a given fruit species is specific for its interaction with a frugivore, so we used the average RPE values of a given fruit of all the frugivore species it provides food to, to characterize its overall or total effectiveness (RPET): RPET= sum(RPEi)/N (2) where, for a given focal fruit species, N is the number of pairwise interactions with RPE data available, and RPEi is the RPE value for a specific pairwise interaction i. Thus, RPET is fruit species-specific, while each pairwise interaction has a specific RPE value (RPEi) depending on the identity of the specific frugivore partner. Data collection RPE Quantity component Most of the data used to calculate the quantitative component have been obtained from available bibliographic sources. Data compilation comes from a total of 51 studies and four databases for frugivore-plant interactions from the Brazilian Atlantic Forest (see Data Sources section for a list of data sources used and Supp. Mat. Fig. S1 for a map of study locations). Variables collected from the bibliography were: number of visits, observation time and number of fruits consumed per visit. To reduce bias we divided the total number of visits to the plants by observation time to control for the different duration of each study. The fruit mass ingested per visit was positively correlated with frugivore body mass (Pearson’s correlation r = 0.588, p < 0.001, n = 541 distinct pairwise interactions; Supp. Mat. Fig. S2). Fruit mass-body mass correlation allowed the estimation of the number of fruits consumed per visit for avian species with no data on fruit consumption rates available; this was done for special cases when avian species had limited number of records in most studies (n = 7 fruit species for Aburria jacutinga, n = 2 fruit species for Penelope obscura, n = 3 fruit species for Procnias nudicollis). We have referred to a fruit as all the dispersing and consequently ingested units (i.e. diaspora). Diaspora and fruit are often the same thing; however, in some cases such as Cabralea canjerana or Virola spp., an aril (i.e. a fleshy covering in some seeds) acts as a diaspore, being smaller than the actual fruit. Therefore, when the diaspore was actually different from the fruit, the energy in the qualitative component has been measured accordingly. In other cases frugivores may peck pieces and ingest just a part of the whole fruit or infructescence (e.g. catkins of Cecropia spp. or syconia from Ficus spp.). For those large infructescences that birds do not consume whole, we used the number of pecks and assessed the percentage of a single fruit actually consumed, corresponding to a given number of pecks and the beak size of the frugivore. Data on each species’ gape size and fruit length (obtained from Bello et al., 2017 and Galetti et al., 2013) allowed us to estimate the total number of pecks needed to consume an infructescence (Supp. Mat. Table S1). RPE Quality component All fruit pulp dry masses and nutrient proportions were obtained from available databases (Pizo, 2017 unpublished; Jordano 1995, 2007; Bello et al., 2017). In a few cases, pulp dry mass values were missing; these were estimated using additional species-specific data such as pulp fresh mass or water proportion (6 species records imputed in this way; see Supp. Mat. Table S2). However, when species-specific data were unavailable, we estimated the specific values by averaging the data available for congeneric species. This type of data imputation was done for a few cases and only when essential to retain a given species in the dataset (9 species records were imputed this way; see Supp. Mat. Table S3). When calculating specific energy, we preferably used the non-structural carbohydrate content for the carbohydrate value; however, when this information was not available total carbohydrate content or total sugar content was used (see Suppl. Mat. Table S4). Due to the diverse origin of nutritional data, variation in the techniques and devices used for their analyses is likely. This variation in methods together with the use of different carbohydrate content variables have limitations when comparing energetic values among fruits. We therefore want to highlight these possible limitations and advise to take results cautiously. While the quality component gives us an approximate idea of the nutritional value of each fruit it does not offer a detailed ranking. A total of 315 different fruit species were reported to be eaten by the frugivore species considered in this study, of which 36 fruits had enough data to calculate the quantity and quality component. [Usage Notes] Please visit the GitHub repository for this dataset to find updates and additional information: https://github.com/PJordano-Lab/MS_JEcol_Resource-provisioning-effectiveness., 1. Mutualistic interactions between frugivorous birds and fleshy fruited plants are key processes for both natural plant regeneration and the maintenance of birds relying on fruit resources. However, seed dispersal effectiveness (SDE) has been frequently studied only from the plant’s perspective, i.e. the contribution of animals to plant fitness. 2. Using a sample of Atlantic rainforest avian frugivores, this study presents a first comparative, empirical study of fruit effectiveness as a nutritional food resource for seed dispersing birds through the use of Resource-Provisioning Effectiveness (RPE) models. RPE is the product of the amount of fruits a frugivore can consume (quantity component) and the quality of the ingesta in terms of energy and nutrients obtained (quality component). 3. Our results show wide variation in Resource-Provisioning Effectiveness among fleshy-fruited plant species. Energy-rich fruits consistently show a smaller quantity component, while energetically-poor fruits are consistently the most consumed, with fruit species spanning a gradient from these two extremes. 4. The specific RPEi resulting from a pairwise fruit-frugivore interaction is positively correlated to the total RPE (RPET) that a given fruit species has for the whole frugivore assemblage. RPE therefore appears to be a characteristic feature of the fruit species, rather than of the specific frugivore partner. 5. Only the fruit’s specific energy content showed a significant phylogenetic signal, suggesting potential constraints for free covariation between RPE and SDE of fruits and frugivores. 6. Synthesis: We analyze variation in the effectiveness of fleshy fruit food provisioning to avian frugivores by explicitly redefining Resource-Provisioning Effectiveness within the Seed Dispersal Effectiveness framework. We found ample variation in RPE among plant species, showing differences in both quantity and quality components of fruit resources rewards for the frugivores. Our findings help unravel how seed-dispersing birds may discriminate among alternative fruit resources and to understand the configuration of mutual dependencies among mutualistic partners., Agencia Estatal de Investigación, Award: CGL2017-82847-P. Fundación La Caixa, Award: LCF/BQ/ES18/11670007. Consejería de Economía, Innovación, Ciencia y Empleo, Junta de Andalucía, Award: RNM–5731., Peer reviewed

[Usage Notes] Dataset consists of genomic data generated in vcf format., [Aim] Differentiation of island lineages from mainland relatives and radiation after initial colonization are two important processes generating island diversity. Both of these processes are influenced by colonization dynamics and contemporary connections between island species and their source populations. The classic island progression rule model, i.e., dispersal from mainland to older islands and subsequently to younger islands, can be considered a null hypothesis, particularly for islands that are geographically aligned perpendicular to the mainland coast with ages inversely proportional to distance from the mainland. Alterations to this pattern have been reported, particularly in archipelagoes that are geographically closely adjacent to mainland source populations. Here we aim to integrate genomic and environmental niche data to infer the colonization history of a Canary endemic species and to understand its current diversity patterns., [Location] Canary Islands., [Taxon] Lavatera acerifolia (Malvaceae)., [Methods] We used high-throughput genotyping-by-sequencing (GBS) combined with species distribution modelling (SDM) projected onto past conditions. Genetic structure (clustering methods), relatedness (coalescent and ML trees), nucleotide diversity and differentiation (population genetics) were assessed based on SNPs obtained from three alternative bioinformatics pipelines. The influence of environmental variables over time was assessed with a GLM in which the response variable was amount of heterozygous sites per individual., [Results] Four genetic groups were identified arranged along a longitudinal gradient, and the earliest diverging coincides with the older, and easternmost, islands (Lanzarote and Fuerteventura). Genetic diversity is reduced in the westernmost islands, which are more distant from the mainland, host few populations and yet apparently offer more suitable habitats., [Main conclusions] The inferred colonization scenario generally fits the progression rule model, but suggests a more complex pattern for the central islands. For the westernmost islands, the contrast between high availability of suitable habitats and reduced genetic diversity and number of populations suggests a colonization front moving at a slow pace, rather than local extinctions, as an explanation for the scarcity of populations in those islands. Historical projections of SDM lend support to this interpretation., Spanish Ministry of Science, Technology and Innovation, Award: CGL2010-16138. Spanish Ministry of Economy and Competitivity, Award: CGL2013-49097-C2-1-P, CGL2017-88500-P (AEI/FEDER, UE). FPI fellowship, Award: BES-2011-049852., Peer reviewed

[Methods] Overall design: We used multiplexed bisulfite sequencing (MBS) to measure the DNA methylation of specific regions in tissues of sea bass from different age classes. Two MBS libraries were prepared and sequenced. In all cases, each tissue and age class were represented by 4 replicate fish. The first MBS library consisted of 22 valid amplicons amplified in ovary, testis and muscle of 1.28, 3.07 and 10.5 years sea bass. The second MBS library consisted of 4 valid amplicons amplified in muscle of a) 0.55, 0.96, 1, 1.1, 1.37, 1.64, 4.17, 5.83 and 6.75 years old sea bass reared at natural (low-17ºC) temperature and b) 0.48, 0.55, 0.82, 0.96, 1.1, 1.37 years old sea bass reared at high (21ºC) temperature from 7 to 68 days post fertilization. Description of protocols: Fish were raised at the Aquarium facilities of the Institute of Marine Sciences or the Institute of Aquaculture Torre de La Sal (Spanish National Research Council, CSIC) following standard procedures until sampling. Samples were snap frozen in liquid nitrogen and kept at -80ºC until DNA extraction. Genomic DNA was extracted by the Phenol/Chlorofom/Isoamyl-alcohol (25:24:1) extraction protocol. MBS libraries were prepared as described in Anastasiadi et al 2018 (https://doi.org/10.1080/15592294.2018.1529504). Briefly, 2 μg of DNA were bisulfite-converted and PCRs with primers targeting specific bisulfite-converted regions were performed. Bead-based normalization of DNA quantities was followed by pooling of amplicons per sample. Indices were added to samples following a dual-index strategy by PCRs. Equal quantities of samples were pooled into a single final library and sequenced in an Illumina MiSeq using the 300 bp paired-end protocol. Description of data processing: Raw reads were quality trimmed by the Trim Galore for MBS 1 and Trimmomatic for MBS 2 Trimmed reads were aligned to the reference genome dicLab (v1.0c, June 2012; http://seabass.mpipz.mpg.de/) using Bismark with --non_directional --phred33-quals --score_min L,0,-0.6. For samples of 1.28, 3.07 and 10.5 years alignments were performed in three steps: 1) paired reads were aligned, 2) unmapped reads from the first step were aligned as single reads, and 3) unpaired reads from the first step of trimming were aligned like the unmapped reads. Paired-end and single-end alignments are provided as .bam files. Methylation calling was performed by the bismark_methylation_extractor. For samples of 1.28, 3.07 and 10.5 years extraction was performed separately for paired-end and for single-end reads. Methylation files were merged into a single file containing all samples. Methylation values were read into R. CpGs with less than 5 coverage were eliminated. In this data package, two types of data are included per sample: alignmnet files (.bam) and methylation value of each CpG (.txt). For the MBS1, paired-end and single-end alignments (.bam) are provided. The processed data files are tab-delimited and contain: 1) the name of the sample, 2) the genomic position of the CpG (in the format chr, cpg start.cpg end), 3) the percent methylation and 4) the age in years. [Usage Notes] A spreadsheet is included in this data package. It includes as columns the name of each sample, the age, tissue, developmental temperature, sex and the names of the corresponding alignment (for MBS1, both paired-end and single-end) and methylation values files., Age-related changes in DNA methylation do occur. Taking advantage of this, mammalian and avian epigenetic clocks have been constructed to predict age. In fish, studies on age-related DNA methylation changes are scarce and no epigenetic clocks are available. However, in fisheries and population studies there is a need for accurate estimation of age, something that is often impossible for some economically important species with the currently available methods. Here, we used the European sea bass, a marine fish where age can be known with accuracy, to construct a piscine epigenetic clock, the first one in a cold-blooded vertebrate. We used targeted bisulfite sequencing to amplify 48 CpGs from four genes in muscle samples and applied penalized regressions to predict age. We, thus, developed an age predictor in fish that is highly accurate (0.824) and precise (2.149 years of error). In juvenile fish, accelerated growth due to elevated temperatures had no effect in age prediction, indicating that the clock is able to predict the chronological age independently of environmentally-driven perturbations. An epigenetic clock developed using muscle samples accurately predicted age in samples of testis but not ovaries, possibly reflecting the reproductive biology of fish. In conclusion, we report the development of the first piscine epigenetic clock, paving the way for similar studies in other species. Piscine epigenetic clocks should be of great utility for fisheries management and conservation purposes, where age determination is of crucial importance., Ministerio de Economía, Industria y Competitividad, Gobierno de España, Award: AGL2016–78710–R .Ministerio de Economía, Industria y Competitividad, Gobierno de España, Award: BES–2011–044860., Peer reviewed

Ozone concentrations in ambient air collected using low-cost sensor technologies, in the framework of EU project CAPTOR. Data collected during summer 2018 in NE Spain and N Italy. Sensors are metal-oxide. Data are calibrated using multiple linear regression, and validated against official reference data from each local air quality monitoring network. More details on the calibration and data validation may be found in A. Ripoll et al. / Science of the Total Environment 651 (2019) 1166–1179., European Commission: CAPTOR - Collective Awareness Platform for Tropospheric Ozone Pollution (688110), Peer reviewed

Ozone concentrations in ambient air collected using low-cost sensor technologies, in the framework of EU project CAPTOR. Data collected during summer 2017 in NE Spain and N Italy. Sensors are metal-oxide. Data are calibrated using multiple linear regression, and validated against official reference data from each local air quality monitoring network. More details on the calibration and data validation may be found in A. Ripoll et al. / Science of the Total Environment 651 (2019) 1166–1179., European Commission: CAPTOR - Collective Awareness Platform for Tropospheric Ozone Pollution (688110), Peer reviewed

Time Series water turbidity derived from Landsat TM, ETM+ & OLI in the Path 202 Row 34 (Doñana). Also, the product and its metadata are freely available to consult or downloaded in the LAST-EBD Cartography Server: http://mercurio.ebd.csic.es/imgs/ Teh methodology is described in the paper: Empirical models to estimate water turbidity from reflectance data from TM or ETM+ Landsat sensors in shallow wetlands such as Doñana marshes. See the reference: Bustamante, J. et al. 2009. Predictive models of turbidity and water depth in the Doñana marshes using Landsat TM and ETM+ images. Journal of Environmental Management. 90:2219-2225.https://doi.org/10.1016/j.jenvman.2007.08.021, European Commission: ECOPOTENTIAL - ECOPOTENTIAL: IMPROVING FUTURE ECOSYSTEM BENEFITS THROUGH EARTH OBSERVATIONS (641762), Peer reviewed

Time Series of NDVI derived from Landsat TM, ETM+ & OLI in the Path 202 Row 34 (Doñana). Also, the product and its metadata are freely available to consult or downloaded in the LAST-EBD Cartography Server: http://mercurio.ebd.csic.es/imgs/, European Commission: ECOPOTENTIAL - ECOPOTENTIAL: IMPROVING FUTURE ECOSYSTEM BENEFITS THROUGH EARTH OBSERVATIONS (641762), Peer reviewed

Time Series of annual Hydroperiods derived from Landsat MSS, TM, ETM+ & OLI in the Path 202 Row 34 (Doñana) cover the period 1974-2018., European Commission: ECOPOTENTIAL - ECOPOTENTIAL: IMPROVING FUTURE ECOSYSTEM BENEFITS THROUGH EARTH OBSERVATIONS (641762), Peer reviewed

Time Series of flooded areas derived from Landsat TM, ETM+ & OLI in the Path 202 Row 34 (Doñana). Also, these products and its metadata are freely available to consult or downloaded in the LAST-EBD Cartography Server: http://mercurio.ebd.csic.es/imgs/ Methodology is described in this paper: Remote Sensing 8(9):775 · September 2016. DOI: 10.3390/rs8090775, European Commission: ECOPOTENTIAL - ECOPOTENTIAL: IMPROVING FUTURE ECOSYSTEM BENEFITS THROUGH EARTH OBSERVATIONS (641762), Peer reviewed