Caracterització histològica i genètica de Perkinsus mediterraneus de les Illes Balears a diferents espècies de mol·luscs bivalves

[cat]Perkinsus mediterraneus infecta, sense mortalitats associades, a una gran varietat de mol·luscs bivalves a l’arxipèlag balear: Ostrea edulis, Mimachlamys varia, Arca noae, Chamelea gallina, Pinna nobilis i Venus verrucosa. La detecció de Perkinsus spp. s’ha realitzat mitjançant RFTM i la determinació de l’espècie per PCR-RFLP i seqüenciació. Els nostres resultats han indicat l’existència a Balears de 12 haplotips de P. mediterraneus amb una elevada similitud genètica. Les anàlisis filogenètiques han detectat tres grups diferents d’O. edulis a l’illa de Menorca, que es diferencien d’altres llinatges coespecífics. Les anàlisis recolzen aquesta diferenciació entre les poblacions de Menorca i Mallorca, la qual sembla, en bona part deguda a l’aïllament geogràfic del port de Maó. Malgrat això, altres factors, com la variabilitat ambiental, diferents localitats i dates de detecció, la translocació d’animals, l’activitat humana, etc. poden tenir certa influència. Malgrat que hi ha co-infeccions a l’escopinya gravada del port de Maó amb P. olseni i P. mediterraneus, no n’hem trobat cap cas, ni tampoc bivalves afectats per P. chesapeaki, espècie que recentment s’ha detectat al delta de l'Ebre. Amb aquest treball es determina la distribució geogràfica de P. mediterraneus, les espècies afectades, la seva variabilitat genètica, la seva prevalença i la dinàmica de la infecció.[eng]A wide bivalve mollusc variety is infected by Perkinsus mediterraneus in Balearic Islands: Ostrea edulis, Mimachlamys varia, Arca noae, Chamelea gallina, Pinna nobilis and Venus verrucosa. Perkinsus spp. search was performed using RFTM and species was established by PCR-RFLP and sequencing. We have found 12 P. mediterraneus haplotypes, all of them sharing a high similarity. Three groups of O. edulis from Minorca were revealed by phylogenetic analyses which are different from other co-specifics lineages. Analysis supported this differentiation among populations from Minorca and Majorca. This differentiation could be due to Mahon harbour geographic isolation. Nevertheless, other factors, such as environmental variability, different detection locations and dates, animal translocations, human activity, etc., might also have some influence. Although it is known that co-infections between P. olseni and P. mediterraneus can happen in Venus verrucosa from Mahon harbour, we have not found any occurrence. Furthermore, we have not detected infection by P. chesapeaki, although it has been found in the Ebro delta. This work establishes distribution, affected species, their genetic variability, its prevalence and the infection dynamics by P. mediterraneus

Environmental DNA: State-of-the-art of its application for fisheries assessment in marine environments

Fisheries management involves a broad and complex set of tasks that are necessary to prevent overfishing and to help the recovery of overfished stock. Monitoring fishing activities based on two main sources, landings data and scientific surveys, is a challenging task. Fisheries collection data is often limited, which compromises the accuracy of the results obtained. Therefore, several emerging applications of molecular methods have the potential to provide unique understanding of ecological processes in marine environments and to build stronger empirical underpinnings for the Ecosystem-Based Fisheries Management. Environmental DNA (eDNA) is a complex mixture of genetic material shed by those organisms that inhabit a given environment, whereby DNA is extracted from an environmental sample without accessing the target organism. eDNA studies can be categorized into two main approaches, i) eDNA metabarcoding or semi-targeted (community) approaches and ii) species-specific or targeted approaches (single). Although both categories are often discussed, they differ drastically in their methodology, interpretations and accuracy. Both approaches involve a series of steps that include eDNA capture, preservation, extraction and amplification. This detection will depend on the affinity to the targeted taxa sequences and completeness and accuracy of DNA reference collection databases. The eDNA method applied in marine environments are probably the most challenging aquatic environments for applying this technique. This is because of the extreme relationship between water-volume to biomass, dynamics and the physical and chemical properties of seawater that affect dispersion, dilution and preservation. Here, we review the present application of this novel method in fishery assessment in marine environments. To date, many studies suggest that this method offers the potential to revolutionize fisheries monitoring, which will contribute to improving the range of tasks involved in fisheries management. The compelling conclusion is that the methodological steps including in eDNA surveys should be standardized and that research efforts should focus on developing appropriately validated tests to address environmental and sampling factors that may affect eDNA detection in marine environments in order to draw reliable conclusions. This bioassessment tool can assist fisheries professionals in achieve their research, management, and conservation objectives, but not as a replacement for time-proven assessment methods

Relating the outcome of HCV infection and different host SNP polymorphisms in a Majorcan population coinfected with HCV–HIV and treated with pegIFN-RBV

Hepatitis C virus (HCV) is one of the major causes of chronic hepatitis, cirrhosis, and hepatocellular carcinoma, and the development of HCV-related disease is accelerated in individuals coinfected with human immunodeficiency-1 virus (HIV). In the present study, we correlated different host single-nucleotide polymorphisms (SNPs) in the IL28B, CTLA4, LDLr, and HFE genes and mitochondrial DNA (mtDNA) haplogroups with the outcome of HCV infection and the response to pegylated-interferon plus ribavirin (pegIFN-RBV) treatment. Our study population consisted of 63 Majorcan patients coinfected with HCV and HIV and 59 anonymous unrelated controls. Whereas the population frequency of IL28B alleles was similar to that found in a North-American cohort of European descent, the frequency of the rs12979860 C allele was lower than that determined in other cohorts from Spain. The frequencies of CTLA4 and LDLr polymorphisms were comparable to those reported in other populations. Significant differences between cases and control cohorts occurred only for the H63D mutation of the HFE gene. There were no other differences in the frequencies of other polymorphisms or mtDNA haplogroups. The IL28B rs12979860 CC genotype was shown to be associated with a rapid virological response, and the spontaneous viral clearance rate for HCV was higher in patients with the CTLA4+49 G allele. There was no relationship between SNPs in the LDLr and HFE genes and mtDNA haplogroups and the response to treatment. Our results suggest that the host genetic background plays a significant role in the pegIFN-RBV response of patients coinfected with HCV and HIV. [Int Microbiol 2014; 17(1):11-20]Keywords: HCV–HIV co-infection · mtDNA haplogroups · SNP polymorphism

New morphological and molecular evidence confirm the presence of the Norwegian skate Dipturus nidarosiensis (Storm, 1881) in the Mediterranean Sea and extend its distribution to the western basin

The present study confirms the presence of the Norwegian skate Dipturus nidarosiensis (Storm, 1881) in the Mediterranean Sea, by means of morphological traits and molecular markers providing the first record of this species in the Alboran Sea. Cannas et al. (2010) reported D. nidarosiensis for the first time in the Mediterranean from specimens captured in the central western basin, but Ebert & Stehmann (2013) and Stehmann et al. (2015) considered these records "likely refer to the smaller morphotype, Dipturus sp.", a species not yet described. Eight specimens of the Dipturus genus (Rajiformes: Rajidae) were caught off the Alboran Island (western Mediterranean) in 2012, 2013 and 2016, between 620 and 819 m depth. These specimens showed morphometric diagnostic features corresponding to those of Norwegian skates from the Northeast Atlantic and the central western Mediterranean Sea. Moreover, the Alboran individuals were genetically compared to Northeast Atlantic specimens available in GenBank by means of two mitochondrial DNA fragments: cytochrome c oxidase subunit I (COI) and cytochrome b (Cytb). Analyses showed that the Northeast Atlantic Norwegian skate specimens and the Alboran Sea ones were genetically similar and shared haplotypes, corroborating the identification of the Alboran individuals as Dipturus nidarosiensis. However, they were different from other Dipturus species distributed throughout the Northeast Atlantic and the Mediterranean Sea. Our results confirm the occurrence of this deep-sea large skate species in the Mediterranean, although the IUCN Red List of Threatened Species does not consider it possible (Stehmann et al., 2015)Versión del edito

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Diet of the Insular Lizard, Podarcis lilfordi (Günther, 1874): Complementary Morphological and Molecular Approaches

[EN] The diets of insular lizards are extremely varied, depending on the different environmental characteristics of each island population. This is particularly evident in the case of the populations of small coastal islets of the Balearic Islands, where the Balearic lizard, Podarcis lilfordi, is found. The study of trophic ecology carried out by means of traditional tools, such as morphological analysis of feces, has made it possible to detect numerous prey and nutritional elements. However, these methods are clearly insufficient, as some rare groups are not detected. It is also difficult to identify remains of marine subsidies or of foods contributed to these small islands by other predators, such as seabirds. The current study demonstrates the advantages of combining morphological diet analysis with the molecular study of individual feces samples obtained from the same populations. We obtained a greater diversity of prey groups using the combined methodologies, with each method identifying prey items that were not detected using the other method. Particularly, the study of diets at the molecular level identified plant species consumed by lizards that were, occasionally, not identified in morphological analyses. Conversely, the traditional morphological study of an equivalent number of fecal samples allowed for the identification of several prey groups that had not been detected in the molecular study. From this viewpoint, the advantages and disadvantages of each methodology are discussed.Ministerio de Economía y Competitividad; Conselleria d'Educació, Cultura i Universitats (Govern de les Illes Balears); Fondo Europeo de Desarrollo Regional (FEDER

State of the Art Review of Environmental DNA Genomics

In recent years, environmental DNA (eDNA) coupled with metabarcoding methodologies has emerged as a promising tool with the potential to improve biodiversity assessment, diet analysis, detection of rare or invasive species, population genetics, and ecosystem functional analysis (Bohmann et al., 2014; Goldberg et al., 2015). eDNA is a complex mixture of genomic DNA from many organisms found in an environment, wherefore DNA is extracted from an environmental sample without accessing the target organism (Lodge et al., 2012; Taberlet et al., 2012a). In general, the eDNA approach involves a series of steps that include eDNA capture, preservation, extraction, amplification, and sequencing to ensure match to target species. Several types of samples have already been used to recover eDNA, including water, soil, feces, pollen, and air (Taberlet et al., 2012a; Deiner et al., 2015). Nevertheless, marine environments are probably the most challenging and difficult aquatic environments for applying the eDNA method. This is because of the extreme water-volume to biomass ratio, the effects of sea currents and wave action on dispersion and dilution of eDNA, and the impact of salinity on the preservation and extraction of eDNA (Thomsen et al., 2012a). eDNA detection features create uncertainty; hence its characterization and appropriate use requires better understanding of eDNA in four domains: origin, state, transport, and fate (Turner et al., 2015). DNA in environmental samples is typically highly degraded into fragments of often less than 150 base pairs (Deagle et al., 2006) and is not always easy to extract. Degradation of eDNA in the environment limits the scope of eDNA studies, as often only small segments of genetic material remain (Turner et al., 2014). eDNA concentration is dependent on biomass, age, and feeding activity of organisms, as well as physiology, life history, and space use (Barnes et al., 2014; Goldberg et al., 2016). Mitochondrial DNA is typically targeted because there are a great number of copies compared to nuclear DNA, its effectiveness in identifying organism to the species level by means of DNA barcoding, including in fish, and its accessibility via universal sequence databases on public servers (e.g. GENBANK and Bold Systems) (Rees et al., 2014). Amplified mitochondrial eDNA may originate from extracellular DNA fragments, mitochondria, cells, excretion, or eggs, and the amount of eDNA quantified is likely to vary depending on the genetic matter being targeted (Herder et al., 2014; Goldberg et al., 2016). A crucial step in the eDNA workflow is DNA capture. Several studies focus on optimization of sampling design and eDNA capture and extraction methods (e.g. Turner et al., 2014; Deiner et al., 2015, Eichmiller et al., 2016). Collection methods typically seek to identify organisms at low densities and, thus, should be optimized for detection sensitivity. Because of this, multiple protocols have been developed in the literature and may be applied to different types of samples (e.g. Turner et al., 2014; Deiner et al., 2015); however, protocol election needs to be carefully considered depending on the goals of the study and the type of sample being analyzed. eDNA extraction protocols that are being optimized within the frame of the FishGenome project target three main applications: single species detection, estimation of abundance and biomass of target species, and biodiversity assessment. The FishGenome project will use two main methods to analyze eDNA: High-Throughput Sequencing (HTS) for biodiversity assessment, and quantitative Polymerase Chain Reaction (qPCR) for the quantification of a target species. For the HTS method, both universal and species-specific primers may be used, but this will depend on the goal of the study. Power of detection will depend on the affinity to the targeted taxa sequences and the availability of DNA reference collection databases needed for species identification. HTS is mostly used to detect multiple species and for biodiversity assessment. Meanwhile, qPCR is widely used for gene expression analysis due to its large dynamic range, tremendous sensitivity, high sequence specificity, little to no postamplification processing, and sample throughput (Lodge et al., 2012). This method is usually performed for species detection and involves the use of species-specific primer sets; it also allows the quantification of target species DNA, which has been shown to correlate with species abundance and biomass in the environment (Lodge et al., 2012; Thomsen et al., 2012a). Since eDNA is a sensitive method, there are many potential sources of “errors”. Some of these errors, which are associated to collecting, laboratory, and bioinformatic procedures, are: contamination, inhibition, amplification and sequencing errors, computational artifacts, and inaccurate taxonomic assignment (Thomsen et al., 2016; Barnes and Turner 2016). Out of these errors, the most serious is probably the risk of contamination and hence the possibility of false positive results. The use and sensitivity of HTS has further complicated the contamination issue, as a very high throughput of DNA sequences is produced (Ficetola et al. 2016). Thus, understanding potential sources of errors and translating these into methodological protocols and interpretations of the results is crucial for reliable outcomes. Along these lines, eDNA offers a potential method to revolutionize marine biomonitoring by significantly augmenting spatial and temporal biological monitoring in aquatic ecosystems due to the ease of collecting water samples (Thomsen and Willerslev, 2015; Sassoubre et al., 2016). eDNA also has the potential to advance fisheries monitoring and conservation by improving detection-probabilities for rare fishes that often comprise a large proportion of the total species richness found in species assemblages. The non-invasive nature of eDNA analysis may provide advantages over traditional capture-based sampling by making it possible to determine the presence or absence of species without disturbance to the fish or their environment. This approach could be especially beneficial in situations of endangered species, where there is significant risk of injury to fishes or damage to their critical habitat (Evans and Lamberti, 2018). More investigations are required in order to understand how well the eDNA method will work for aquatic species, to evaluate the effect of species abundance on detection efficiency, and to upscale species detection from local water samples to larger spatial areas, such as drainage basins. However, it is challenging to work with such small amounts of DNA.N

Environmental DNA: State-of-the-art of its application for fisheries assessment in marine environments

Fisheries management involves a broad and complex set of tasks that are necessary to prevent overfishing and to help the recovery of overfished stock. Monitoring fishing activities based on two main sources, landings data and scientific surveys, is a challenging task. Fisheries collection data is often limited, which compromises the accuracy of the results obtained. Therefore, several emerging applications of molecular methods have the potential to provide unique understanding of ecological processes in marine environments and to build stronger empirical underpinnings for the Ecosystem-Based Fisheries Management. Environmental DNA (eDNA) is a complex mixture of genetic material shed by those organisms that inhabit a given environment, whereby DNA is extracted from an environmental sample without accessing the target organism. eDNA studies can be categorized into two main approaches, i) eDNA metabarcoding or semi-targeted (community) approaches and ii) species-specific or targeted approaches (single). Although both categories are often discussed, they differ drastically in their methodology, interpretations and accuracy. Both approaches involve a series of steps that include eDNA capture, preservation, extraction and amplification. This detection will depend on the affinity to the targeted taxa sequences and completeness and accuracy of DNA reference collection databases. The eDNA method applied in marine environments are probably the most challenging aquatic environments for applying this technique. This is because of the extreme relationship between water-volume to biomass, dynamics and the physical and chemical properties of seawater that affect dispersion, dilution and preservation. Here, we review the present application of this novel method in fishery assessment in marine environments. To date, many studies suggest that this method offers the potential to revolutionize fisheries monitoring, which will contribute to improving the range of tasks involved in fisheries management. The compelling conclusion is that the methodological steps including in eDNA surveys should be standardized and that research efforts should focus on developing appropriately validated tests to address environmental and sampling factors that may affect eDNA detection in marine environments in order to draw reliable conclusions. This bioassessment tool can assist fisheries professionals in achieve their research, management, and conservation objectives, but not as a replacement for time-proven assessment methods.The information and views set out in this publication are based on scientific data and information collected under Service Contract “Improving cost-efficiency of fisheries research surveys and fish stocks assessments using next-generation genetic sequencing methods [EMFF/2018/015]” signed with the European Climate, Infrastructure and Environment Executive Agency (CINEA) and funded by the European Union. The information and views set out in this publication are those of the author(s) and do not necessarily reflect the official opinion of CINEA or of the European Commission. Neither CINEA nor the European Commission can guarantee the accuracy of the scientific data/information collected under the above Specific Contract or the data/information included in this publication. Neither CINEA nor the European Commission or any person acting on their behalf may be held responsible for the use which may be made of the information contained therein.” SRA (Vicenç Mut Estabilidad) and MB (ForInDoc) were supported by postdoctoral contracts co-funded by the Regional Government of the Balearic Islands and the European Social Fund.Peer reviewe

Faecal Microbiota Divergence in Allopatric Populations of Podarcis lilfordi and P. pityusensis, Two Lizard Species Endemic to the Balearic Islands

[EN] Gut microbial communities provide essential functions to their hosts and are known to influence both their ecology and evolution. However, our knowledge of these complex associations is still very limited in reptiles. Here we report the 16S rRNA gene faecal microbiota profiles of two lizard species endemic to the Balearic archipelago (Podarcis lilfordi and P. pityusensis), encompassing their allopatric range of distribution through a noninvasive sampling, as an alternative to previous studies that implied killing specimens of these IUCN endangered and near-threatened species, respectively. Both lizard species showed a faecal microbiome composition consistent with their omnivorous trophic ecology, with a high representation of cellulolytic bacteria taxa. We also identified species-specific core microbiota signatures and retrieved lizard species, islet ascription, and seasonality as the main factors in explaining bacterial community composition. The different Balearic Podarcis populations are characterised by harbouring a high proportion of unique bacterial taxa, thus reinforcing their view as unique and divergent evolutionary entities.Ministerio de Economía y Competitividad; FEDER; Conselleria d'Educació, Cultura i Universitat