New insights about the introduction of the Portuguese oyster, Crassostrea angulata, into the North East Atlantic from Asia based on a highly polymorphic mitochondrial region

It is commonly presumed that the Portuguese oyster Crassostrea angulata was introduced into the North East (NE) Atlantic from Asia. The analysis of the nucleotide sequence of a highly polymorphic non-coding mitochondrial region (major noncoding region - MNR) of C. angulata samples collected in Europe (Portugal), Africa (Morocco) and Asia (Shantou and Taiwan) provided new insight into the introduction of this species into the NE Atlantic. Sixty haplotypes and a nucleotide diversity of 0.0077 were observed in 130 analyzed sequences. Higher nucleotide diversity levels were observed in NE Atlantic sites than in Asian sites and significant genetic differentiation was found between the two. Our results suggest that C. angulata might have been introduced to the NE Atlantic by multiple introductory events, though the exact origins remain unknown since none of the analyzed Asian sites seemed to have been a source of introduction. The nucleotide diversity of C. angulata was higher than that previously reported for Pacific oyster C. gigas in Europe and Asia for the same mitochondrial region. The results obtained in the present study suggest that NE Atlantic C. angulata stocks are a unique genetic resource, which highlights the importance of their conservation

Identification of molecular and physiological responses to chronic environmental challenge in an invasive species: the Pacific oyster, Crassostrea gigas

Understanding the environmental responses of an invasive species is critical in predicting how ecosystem composition may be transformed in the future, especially under climate change. In this study, Crassostrea gigas, a species well adapted to the highly variable intertidal environment, was exposed to the chronic environmental challenges of temperature (19 and 24°C) and pH (ambient seawater and a reduction of 0.4 pH units) in an extended 3-month laboratory-based study. Physiological parameters were measured (condition index, shell growth, respiration, excretion rates, O:N ratios, and ability to repair shell damage) alongside molecular analyses. Temperature was by far the most important stressor, as demonstrated by reduced condition indexes and shell growth at 24°C, with relatively little effect detected for pH. Transcriptional profiling using candidate genes and SOLiD sequencing of mantle tissue revealed that classical “stress” genes, previously reported to be upregulated under acute temperature challenges, were not significantly expressed in any of the treatments, emphasizing the different response between acute and longer term chronic stress. The transcriptional profiling also elaborated on the cellular responses underpinning the physiological results, including the identification of the PI3K/AKT/mTOR pathway as a potentially novel marker for chronic environmental challenge. This study represents a first attempt to understand the energetic consequences of cumulative thermal stress on the intertidal C. gigas which could significantly impact on coastal ecosystem biodiversity and function in the future

Whole Genome Sequencing of Hepatitis A Virus Using a PCR-Free Single-Molecule Nanopore Sequencing Approach

Hepatitis A virus (HAV) is one of the most common causes of acute viral hepatitis in humans. Although HAV has a relatively small genome, there are several factors limiting whole genome sequencing such as PCR amplification artefacts and ambiguities in de novo assembly. The recently developed Oxford Nanopore technologies (ONT) allows single-molecule sequencing of long-size fragments of DNA or RNA using PCR-free strategies. We have sequenced the whole genome of HAV using a PCR-free approach by direct reverse-transcribed sequencing. We were able to sequence HAV cDNA and obtain reads over 7 kilobases in length containing almost the whole genome of the virus. The comparison of these raw long nanopore reads with the HAV reference wild type revealed a nucleotide sequence identity between 81.1 and 96.6%. By de novo assembly of all HAV reads we obtained a consensus sequence of 7362 bases, with a nucleotide sequence identity of 99.0% with the genome of the HAV strain pHM175/18f. When the assembly was performed using as reference the HAV strain pHM175/18f a consensus with a sequence similarity of 99.8 % was obtained. We have also used an ONT amplicon-based assay to sequence two fragments of the VP3 and VP1 regions which showed a sequence similarity of 100% with matching regions of the consensus sequence obtained using the direct cDNA sequencing approach. This study showed the applicability of ONT sequencing technologies to obtain the whole genome of HAV by direct cDNA nanopore sequencing, highlighting the utility of this PCR-free approach for HAV characterization and potentially other viruses of the Picornaviridae family

Comparative study of shell shape and muscle scar pigmentation in the closely related cupped oysters Crassostrea angulata, C-gigas and their reciprocal hybrids

The taxonomic status of the cupped oysters Crassostrea angulata and C. gigas has received considerable attention in the last decades. Based on larval shell morphology, experimental hybridization, allozymes and nuclear DNA studies several authors have considered these two taxa as being synonymous. However, mitochondrial data showed clear genetic differences between the two taxa. In addition, microsatellite- based studies and cytogenetic studies have also provided evidence that supports their differentiation. Considerable differences have also been observed at the phenotypic level in terms of growth rate and ecophysiological parameters. In the present study, C. angulata from Sado estuary ( Portugal) and C. gigas from Seudre estuary ( France) were collected and factorial crosses were performed. Juveniles of the different progenies were reared in Ria Formosa ( Portugal) under common conditions to determine if they exhibited differences in shell shape and in pigmentation of the adductor muscle scar. Significant morphometric differences between C. angulata and C. gigas progenies were indicated by univariate and multivariate analyses. Univariate analysis of size- adjusted shell measurements revealed significant differences between the two taxa for shell depth, muscle scar height, and length of ligamental area. Both reciprocal hybrids showed intermediate morphometric characters between parental lines. In addition, significant differences were also observed between C. angulata and C. gigas progenies in terms of pigmentation of adductor muscle scar. C. angulata and both reciprocal hybrid progenies showed highly pigmented adductor muscle scars whereas in C. gigas progeny the pigmentation was lighter. The differences in shell shape and muscle scar pigmentation observed in the present study support the distinction of the two taxa.info:eu-repo/semantics/publishedVersio

Diagnosis and prevalence of two new species of haplosporidians infecting shore crabs Carcinus maenas: Haplosporidium carcini n. sp., and H. cranc n. sp.

This study provides a morphological and phylogenetic characterization of two novel species of the order Haplosporida (Haplosporidium carcini n. sp., and H. cranc n. sp.) infecting the common shore crab Carcinus maenas collected at one location in Swansea Bay, South Wales, UK. Both parasites were observed in the haemolymph, gills and hepatopancreas. The prevalence of clinical infections (i.e. parasites seen directly in fresh haemolymph preparations) was low, at ~1%, whereas subclinical levels, detected by polymerase chain reaction, were slightly higher at ~2%. Although no spores were found in any of the infected crabs examined histologically (n = 334), the morphology of monokaryotic and dikaryotic unicellular stages of the parasites enabled differentiation between the two new species. Phylogenetic analyses of the new species based on the small subunit (SSU) rDNA gene placed H. cranc in a clade of otherwise uncharacterized environmental sequences from marine samples, and H. carcini in a clade with other crustacean-associated lineages

Setting performance indicators for coastal marine protected areas: An expert-based methodology

Marine Protected Areas (MPAs) require effective indicators to assess their performance, in compliance with the goals of relevant national and international commitments. Achieving and prioritizing shortlists of multidisciplinary indicators demands a significant effort from specialists to depict the multiple conservation and socioeconomic interests, and the large complexity of natural systems. The present paper describes a structured expert-based methodology (process and outputs) to co-define a list of multidisciplinary MPA performance indicators. This work was promoted by the management authority of coastal MPAs in mainland Portugal to gather a consensual and feasible list of indicators that would guide the design of a future national monitoring program. Hence, Portuguese coastal MPAs served as a case study to develop such a process between 2019 and 2020. In the end, participants (1) agreed on a shortlist of prioritized indicators (i.e., environmental, governance, and socioeconomic indicators) and (2) defined minimum monitoring frequencies for the indicators in this list, compatible with the potential replicability of the associated survey methods. The present approach recommends that management plans incorporate monitoring procedures and survey methods, with a validated list of indicators and associated monitoring periodicity, agreed among researchers, MPA managers and governance experts. The proposed methodology, and the lessons learned from it, can support future processes aiming to define and prioritize MPA performance indicatorsFundação para a Ciência e Tecnologia - FCT, European Maritime and Fisheries Fund (EMFF)info:eu-repo/semantics/publishedVersio