Impacts of ocean acidification and warming on post-larval growth and metabolism in two populations of the great scallop (Pecten maximus).

Ocean acidification and warming are key stressors for many marine organisms. Some organisms display physiological acclimatization or plasticity, but this may vary across species ranges, especially if populations are adapted to local climatic conditions. Understanding how acclimatization potential varies among populations is therefore important in predicting species responses to climate change. We carried out a common garden experiment to investigate how different populations of the economically important great scallop (Pecten maximus) from France and Norway responded to variation in temperature and PCO2 concentration. After acclimation, post-larval scallops (spat) were reared for 31 days at one of two temperatures (13°C or 19°C) under either ambient or elevated PCO2 (pH 8.0 and pH 7.7). We combined measures of proteomic, metabolic and phenotypic traits to produce an integrative picture of how physiological plasticity varies between the populations. The proteome of French spat showed significant sensitivity to environmental variation, with 12 metabolic, structural and stress-response proteins responding to temperature and/or PCO2. Principal component analysis revealed seven energy metabolism proteins in French spat that were consistent with countering ROS stress under elevated temperature. Oxygen uptake in French spat did not change under elevated temperature but increased under elevated PCO2. In contrast, Norwegian spat reduced oxygen uptake under both elevated temperature and PCO2. Metabolic plasticity allows French scallops to maintain greater energy availability for growth compared with Norwegian spat. However, increased physiological plasticity and growth in French spat may come at a cost, as they showed reduced survival compared with Norwegian scallops under elevated temperature.

Fruit Juices of Etcho (Pachycereus pecten-aboriginum) and Giant Cardon (Pachycereus pringlei) are Sources of Health-Promoting Ingredients with Potential Anticancer Properties.

Mexico is one of the main diversification centers of cacti in the world, with more than 500 endemic species, most of which remain nutritionally and functionally uncharacterized. The columnar cacti of the genus Pachycereus comprise five underutilized endemic Mexican species, whose nutraceutical properties have only been studied in the P. weberi species. Therefore, this study aimed to evaluate the nutritional quality and bioactive properties of etcho (P. pecten-aboriginum) and giant cardon (P. pringlei) fruit. The physical, chemical, and nutritional composition of etcho and giant cardon fruits were characterized, as well as the profile and content of bioactive compounds, antioxidant activity (ABTS•+ and DPPH•), and antiproliferative capacity in cervical (HeLa) and breast cancer (MDA-MB-231, MCF-7, and T-47D) cell lines. Our results suggest that etcho and giant cardon fruits are rich sources of essential nutrients and bioactive phytochemicals (including K, Mg, P, dietary fiber, polyphenolic compounds, vitamin C, betalains, and myo-inositol) with antioxidant and anticancer potential by inhibiting the proliferation of all evaluated cell lines with IC50 values in the range of 198 to 287 µg of gallic acid equivalents/mL. Therefore, etcho and giant cardon fruits could be used for nutraceutical purposes, and their consumption could promote health benefits.

Temperature-associated selection linked to putative chromosomal inversions in king scallop (Pecten maximus).

The genomic landscape of divergence-the distribution of differences among populations or species across the genome-is increasingly characterized to understand the role that microevolutionary forces such as natural selection and recombination play in causing and maintaining genetic divergence. This line of inquiry has also revealed chromosome structure variation to be an important factor shaping the landscape of adaptive genetic variation. Owing to a high prevalence of chromosome structure variation and the strong pressure for local adaptation necessitated by their sessile nature, bivalve molluscs are an ideal taxon for exploring the relationship between chromosome structure variation and local adaptation. Here, we report a population genomic survey of king scallop (Pecten maximus) across its natural range in the northeastern Atlantic Ocean, using a recent chromosome-level genome assembly. We report the presence of at least three large (12-22 Mb), putative chromosomal inversions associated with sea surface temperature and whose frequencies are in contrast to neutral population structure. These results highlight a potentially large role for recombination-suppressing chromosomal inversions in local adaptation and suggest a hypothesis to explain the maintenance of differences in reproductive timing found at relatively small spatial scales across king scallop populations.

Identification of the shell-boring parasite Polydora hoplura (Annelida: Spionidae) on wild stocks of Pecten maximus in Galician waters, NW Spain.

Infestations by shell-boring polychaetes have been gaining attention in recent years due to the harmful effects that their presence can pose to the growth and production of marine bivalves worldwide. The Galician region, located in the north-western corner of the Iberian Peninsula, is one of the major producers of marine bivalves in the word and therefore highly dependent on this industry. The recent finding of an unknown polychaete boring into the shells of the king scallop, Pecten maximus, has raised the attention of producers and authorities due to its potential harm to exploitable populations and the economical losses that could entail. The results from the morphological examination revealed the spionid Polydora hoplura as the only species present. DNA of a specimen was extracted and the mitochondrial 16S rRNA gene was partially sequenced for comparison with published sequences and confirmation of the morphological identification. Consequently, we report for the first time in the area the infestation of wild harvested populations of P. maximus by the mud worm Polydora hoplura. The presence of this shell-boring cosmopolitan invader could pose a threat to king scallop exploitation in Galicia; potential economic impacts need to be evaluated.

The presence of Apicomplexan parasites in king scallops (Pecten maximus) in Scottish waters.

The king scallop (Pecten maximus) is a commercially important species found around the United Kingdom coast. The association of an Apicomplexan-like parasite with mass mortality of Icelandic scallop (Chlamys islandica) in Iceland and the presence of identical parasites in king scallop (Pecten maximus) and queen scallop (Aequipecten opercularis) in Scotland raised serious concerns regarding the health of Scottish king scallops. Marine Scotland Science (MSS) conducted a survey in 2016 to assess the prevalence and the intensity of parasite infection in king scallops. King scallops were collected and sampled during the annual scallop dredge surveys in the Shetland Isles and the east and west coast of Scotland. The king scallop adductor muscle was macroscopically examined and tissue imprints taken to grade the intensity of infection. The parasite was present in the majority of the king scallops sampled in all surveyed areas: Shetland Isles 87.1%, east coast 76.0% and west coast of Scotland 64.1%. However, the parasitic infestations were light in intensity with the majority of the king scallops graded as 1 (≤20 zoites per microscopic field). No macroscopic changes in the adductor muscle were observed and histopathology examination revealed minor localized fiber degeneration of adjacent fibers to parasite clusters. The results suggested the parasite to be widespread around the Scottish coast and it appears to be able to live within the king scallop at low intensity of infection without causing significant downgrade of the adductor muscle (in terms of colour or texture) or mortality. The partial genome sequence of the parasite in king scallops from Scottish waters was identical to the one reported by Kristmundsson and Freeman (2018) in the Icelandic scallop in Icelandic waters.

Intracellular bacteria in New Zealand shellfish are identified as Endozoicomonas species.

Kaimoana (shellfish, seafood) is an important food source and a significant social and cultural component of many New Zealand communities, especially the indigenous Maori. Over the past decade a decline has been detected in shellfish health and an increase in mortality events around New Zealand. Intracellular bacteria termed Rickettsia-like organisms (RLOs) have been observed in New Zealand bivalve molluscs during shellfish mortality events. Affected bivalves include cockles Austrovenus stutchburyi, ringed dosinia Dosinia anus, green-lipped mussels Perna canaliculus, pipi Paphies australis, toheroa Paphies ventricosa, tuatua Paphies subtriangulata, deepwater tuatua Paphies donacina and scallops Pecten novaezelandiae. RLOs are an informal morphology-based classification of intracellular bacteria, with the exact identification often unknown. Using shellfish collected during mortality events from 2014 to 2019 and apparently healthy samples collected in 2018 and 2019, we aimed to identify RLOs in New Zealand shellfish. Bacterial 16S rRNA gene sequences from RLO-infected shellfish showed >95% identity to published Endozoicomonas species. In situ hybridization confirmed the presence of the sequenced gene in the gill epithelium and digestive epithelium of all study species. A genus-specific quantitative PCR, targeting the 16S rRNA gene was developed to detect Endozoicomonas spp. in shellfish tissue. Prevalence of Endozoicomonas spp. in samples from mortality events and healthy shellfish analysed by quantitative PCR was high. Samples collected from mortality events, however, had a significantly higher load of Endozoicomonas spp. than the healthy samples. These results give us a greater understanding of these intracellular bacteria and their presence in populations of New Zealand shellfish.

Exposure to seismic air gun signals causes physiological harm and alters behavior in the scallop Pecten fumatus.

Seismic surveys map the seabed using intense, low-frequency sound signals that penetrate kilometers into the Earth's crust. Little is known regarding how invertebrates, including economically and ecologically important bivalves, are affected by exposure to seismic signals. In a series of field-based experiments, we investigate the impact of exposure to seismic surveys on scallops, using measurements of physiological and behavioral parameters to determine whether exposure may cause mass mortality or result in other sublethal effects. Exposure to seismic signals was found to significantly increase mortality, particularly over a chronic (months postexposure) time scale, though not beyond naturally occurring rates of mortality. Exposure did not elicit energetically expensive behaviors, but scallops showed significant changes in behavioral patterns during exposure, through a reduction in classic behaviors and demonstration of a nonclassic "flinch" response to air gun signals. Furthermore, scallops showed persistent alterations in recessing reflex behavior following exposure, with the rate of recessing increasing with repeated exposure. Hemolymph (blood analog) physiology showed a compromised capacity for homeostasis and potential immunodeficiency, as a range of hemolymph biochemistry parameters were altered and the density of circulating hemocytes (blood cell analog) was significantly reduced, with effects observed over acute (hours to days) and chronic (months) scales. The size of the air gun had no effect, but repeated exposure intensified responses. We postulate that the observed impacts resulted from high seabed ground accelerations driven by the air gun signal. Given the scope of physiological disruption, we conclude that seismic exposure can harm scallops.

Shedding and survival of an intracellular pathogenic Endozoicomonas-like organism infecting king scallop Pecten maximus.

The Lyme Bay marine protected area (MPA) hosts a valuable population of king scallop Pecten maximus L. Recently, an Endozoicomonas-like organism (ELO), infecting host gill epithelial tissue, was associated with king scallop mass mortality events within the Lyme Bay MPA. Currently, very little is known about its transmission and survival outside the host. In this investigation, animals collected outside of reported mortality events showed high levels of ELO infection. Gill tissue disruption and the release of bacteria into the interlamellar space was seen histologically, suggesting shedding of ELO from host animals. To investigate pathogen survival outside the host, infected scallops were maintained in static water for a 24 h period, and then removed. Over the subsequent 8 d, water samples were collected and the quantity of ELO 16S rRNA transcript was measured by TaqManTM quantitative PCR (qPCR). The 16S rRNA transcript quantity was stable outside the host for 6 d before bacteria survival declined 2 logs (7.9 × 108 16S rRNA to 2.3 × 106 transcripts), suggesting that ELO can survive independently outside the host organism. The ELO-specific qPCR probe can therefore be used in future field studies of ELO prevalence within the environment and fauna of the Lyme Bay MPA.

Molecular Characterization of an Endozoicomonas-Like Organism Causing Infection in the King Scallop (Pecten maximus L.).

One of the fastest growing fisheries in the UK is the king scallop (Pecten maximus L.), also currently rated as the second most valuable fishery. Mass mortality events in scallops have been reported worldwide, often with the causative agent(s) remaining uncharacterized. In May 2013 and 2014, two mass mortality events affecting king scallops were recorded in the Lyme Bay marine protected area (MPA) in Southwest England. Histopathological examination showed gill epithelial tissues infected with intracellular microcolonies (IMCs) of bacteria resembling Rickettsia-like organisms (RLOs), often with bacteria released in vascular spaces. Large colonies were associated with cellular and tissue disruption of the gills. Ultrastructural examination confirmed the intracellular location of these organisms in affected epithelial cells. The 16S rRNA gene sequences of the putative IMCs obtained from infected king scallop gill samples, collected from both mortality events, were identical and had a 99.4% identity to 16S rRNA gene sequences obtained from "Candidatus Endonucleobacter bathymodioli" and 95% with Endozoicomonas species. In situ hybridization assays using 16S rRNA gene probes confirmed the presence of the sequenced IMC gene in the gill tissues. Additional DNA sequences of the bacterium were obtained using high-throughput (Illumina) sequencing, and bioinformatic analysis identified over 1,000 genes with high similarity to protein sequences from Endozoicomonas spp. (ranging from 77 to 87% identity). Specific PCR assays were developed and applied to screen for the presence of IMC 16S rRNA gene sequences in king scallop gill tissues collected at the Lyme Bay MPA during 2015 and 2016. There was 100% prevalence of the IMCs in these gill tissues, and the 16S rRNA gene sequences identified were identical to the sequence found during the previous mortality event.IMPORTANCE Molluscan mass mortalities associated with IMCs have been reported worldwide for many years; however, apart from histological and ultrastructural characterization, characterization of the etiological agents is limited. In the present work, we provide detailed molecular characterization of an Endozoicomonas-like organism (ELO) associated with an important commercial scallop species.

Uptake, Whole-Body Distribution, and Depuration of Nanoplastics by the Scallop Pecten maximus at Environmentally Realistic Concentrations.

Previous studies of uptake and effects of nanoplastics by marine organisms have been conducted at what may be unrealistically high concentrations. This is a consequence of the analytical challenges in tracking plastic particles in organisms at environmentally relevant concentrations and highlights the need for new approaches. Here, we present pulse exposures of 14C-radiolabeled nanopolystyrene to a commercially important mollusk, Pecten maximus, at what have been predicted to be environmentally relevant concentrations (<15 µg L-1). Uptake was rapid and was greater for 24 nm than for 250 nm particles. After 6 h, autoradiography showed accumulation of 250 nm nanoplastics in the intestine, while 24 nm particles were dispersed throughout the whole-body, possibly indicating some translocation across epithelial membranes. However, depuration was also relatively rapid for both sizes; 24 nm particles were no longer detectable after 14 days, although some 250 nm particles were still detectable after 48 days. Particle size thus apparently influenced the biokinetics and suggests a need for chronic exposure studies. Modeling extrapolations indicated that it could take 300 days of continued environmental exposure for uptake to reach equilibrium in scallop body tissues although the concentrations would still below 2.7 mg g-1. Comparison with previous work in which scallops were exposed to nonplastic (silver) nanomaterials of similar size (20 nm), suggests that nanoparticle composition may also influence the uptake tissue distributions somewhat.

Arcobacter lekithochrous sp. nov., isolated from a molluscan hatchery.

Four bacterial strains, LFT 1.7T, LT2C 2.5, LT4C 2.8 and TM 4.6, were isolated from great scallop (Pecten maximus) larvae and tank seawater in a Norwegian hatchery and characterized by a polyphasic approach including determination of phenotypic, chemotaxonomic and genomic traits. All were Gram-stain-negative, motile rods, oxidase- and catalase-positive and required sea salts for growth. Major fatty acids present were summed feature 3 (C16 : 1ω7c/C16 : 1ω6c), summed feature 8 (C18 : 1ω7c or C18 : 1ω6c), C16 : 0, C14 : 0, summed feature 2 (C14 : 0 3-OH/iso-C16 : 1 I), C12 : 0 3-OH and C12 : 0. Strain LFT 1.7T contained menaquinone MK-6 as the sole respiratory quinone. Phylogenetic analysis based on 16S rRNA gene sequences indicated that all strains formed a distinct lineage within the genus Arcobacter with a low similarity to known species (94.77-95.32 %). The DNA G+C content was 28.7 mol%. Results of in silico DNA-DNA hybridization and average nucleotide identity confirmed that the isolates constitute a novel species of Arcobacter, for which the name Arcobacter lekithochrous sp. nov. is proposed. The type strain is LFT 1.7T (=CECT 8942T=DSM 100870T).

Stock enhancement or sea ranching? Insights from monitoring the genetic diversity, relatedness and effective population size in a seeded great scallop population (Pecten maximus).

The mass release of hatchery-propagated stocks raises numerous questions concerning its efficiency in terms of local recruitment and effect on the genetic diversity of wild populations. A seeding program, consisting of mass release of hatchery-produced juveniles in the local naturally occurring population of great scallops (Pecten maximus L.), was initiated in the early 1980s in the Bay of Brest (France). The present study aims at evaluating whether this seeding program leads to actual population enhancement, with detectable effects on genetic diversity and effective population size, or consists of sea ranching with limited genetic consequences on the wild stock. To address this question, microsatellite-based genetic monitoring of three hatchery-born and naturally recruited populations was conducted over a 5-year period. Results showed a limited reduction in allelic richness but a strong alteration of allelic frequencies in hatchery populations, while genetic diversity appeared very stable over time in the wild populations. A temporal increase in relatedness was observed in both cultured stock and wild populations. Effective population size (Ne) estimates were low and variable in the wild population. Moreover, the application of the Ryman-Laikre model suggested a high contribution of hatchery-born scallops to the reproductive output of the wild population. Overall, the data suggest that the main objective of the seeding program, which is stock enhancement, is fulfilled. Moreover, gene flow from surrounding populations and/or the reproductive input of undetected sub-populations within the bay may buffer the Ryman-Laikre effect and ensure the retention of the local genetic variability.

The quality of great scallop (Pecten maximus) sperm after thawing.

Most publications devoted to the cryopreservation of mollusc sperm have focused on the definition of technical protocols, avoiding the description of sperm quality after thawing. The present study investigated the effects of cryopreservation on sperm quality in the great scallop. Wild scallop were fished during the natural spawning period and conditioned in the hatchery before use. Sperm samples were obtained after intragonadal injection of serotonin and cryopreserved using a previously published protocol. Sperm quality was assessed using a panel of four parameters: sperm motility characteristics, using a computer assisted sperm analysis plugin with Image J, intracellular ATP content using an ATP-Lite kit, sperm integrity, using flow cytometry and sperm morphology, using transmission electron microscopy. For each parameter, fresh (control) and thawed spermatozoa were compared. A significant decrease of both the percentage of motile spermatozoa (reduction: 75%) and sperm swimming speed (86%) were observed for thawed sperm compared with fresh sperm. The percentage of living spermatozoa, as assessed using flow cytometry, was significantly lower for thawed sperm (72.4±2.5%) compared with fresh sperm (86.4±1.1). However, no significant difference of intracellular sperm ATP content was observed between fresh and thawed sperm. Post thawing, while some spermatozoa showed little or no morphological differences compared with fresh sperm, others had undergone drastic changes, including swelling of the plasma membrane, structural alterations of the chromatin and damage to mitochondria. In conclusion, the descriptive parameters studied in the present work showed that the quality of thawed great scallop sperm was lower than that of fresh cells but was still sufficient for use in aquaculture programs and sperm cryobanking for this species.

Deciphering the molecular adaptation of the king scallop (Pecten maximus) to heat stress using transcriptomics and proteomics.

BACKGROUND: The capacity of marine species to survive chronic heat stress underpins their ability to survive warming oceans as a result of climate change. In this study RNA-Seq and 2-DE proteomics were employed to decipher the molecular response of the sub-tidal bivalve Pecten maximus, to elevated temperatures. RESULTS: Individuals were maintained at three different temperatures (15, 21 and 25 °C) for 56 days, representing control conditions, maximum environmental temperature and extreme warming, with individuals sampled at seven time points. The scallops thrived at 21 °C, but suffered a reduction in condition at 25 °C. RNA-Seq analyses produced 26,064 assembled contigs, of which 531 were differentially expressed, with putative annotation assigned to 177 transcripts. The proteomic approach identified 24 differentially expressed proteins, with nine identified by mass spectrometry. Network analysis of these results indicated a pivotal role for GAPDH and AP-1 signalling pathways. Data also suggested a remodelling of the cell structure, as revealed by the differential expression of genes involved in the cytoskeleton and cell membrane and a reduction in DNA repair. They also indicated the diversion of energetic metabolism towards the mobilization of lipid energy reserves to fuel the increased metabolic rate at the higher temperature. CONCLUSIONS: This work provides preliminary insights into the response of P. maximus to chronic heat stress and provides a basis for future studies examining the tipping points and energetic trade-offs of scallop culture in warming oceans.

A novel satellite DNA isolated in Pecten jacobaeus shows high sequence similarity among molluscs.

The aim of this work is to investigate the sequence conservation and the evolution of repeated DNA in related species. Satellite DNA is a component of eukaryotic genomes and is made up of tandemly repeated sequences. These sequences are affected by high rates of mutation that lead to the occurrence of species-specific satellite DNAs, which are different in terms of both quantity and quality. In this work, a novel repetitive DNA family, named PjHhaI sat, is described in Pecten jacobaeus. The quantitative analyses revealed a different abundance of this element in the molluscan species investigated in agreement with the "library hypothesis" even if, in this case, at a high taxonomic level. In addition, the qualitative analysis demonstrated an astonishing sequence conservation not only among scallops but also in six other molluscan species belonging to three classes. These findings suggest that the PjHhaI sat may be considered as the most ancients of DNA described so far, which remained "frozen" during molluscan evolution. The widespread distribution of this sat DNA in molluscs as well as its long evolutionary preservation open up questions on the functional role of this element. A future challenge might be the identification of proteins or molecules which interact with the PjHhaI sat.

Emerging patterns of genetic variation in the New Zealand endemic scallop Pecten novaezelandiae.

Both historical and contemporary processes influence the genetic structure of species, but the relative roles of such processes are still difficult to access. Population genetic studies of species with recent evolutionary histories such as the New Zealand endemic scallop Pecten novaezelandiae (<1 Ma) permit testing of the effects of recent processes affecting gene flow and shaping genetic structure. In addition, studies encompassing the entire distributional range of species can provide insight into colonization processes. Analyses of genetic variation in P. novaezelandiae (952 individuals from 14 locations, genotyped at 10 microsatellite loci) revealed a weak but significant regional structure across the distributional range of the species, as well as latitudinal gradients of genetic diversity and differentiation: estimates of migration rates supported these patterns. Our results suggest that the observed genetic structure and latitudinal gradients reflect a stepping-stone model of colonization (north to south) and emerging divergence of populations as a result of ongoing limitations to gene flow and insufficient time to reach migration-drift equilibrium. The low levels of interpopulation and interregional genetic differentiation detected over hundreds of kilometres reflect the recent evolutionary history of P. novaezelandiae and stand in contrast to patterns reported for other evolutionary older species at the same spatial scale. The outcomes of this study contribute to a better understanding of evolutionary processes influencing the genetic variation of species and provide vital information on the genetic structure of P. novaezelandiae.

Sinobacterium norvegicum sp. nov., isolated from great scallop (Pecten maximus) broodstock and emended description of Sinobacterium caligoides.

Six isolates were recovered from great scallop (Pecten maximus) broodstock in a hatchery in Bergen, Norway. The strains were thoroughly characterized by a polyphasic approach. Phylogenetic analysis based on 16S rRNA gene sequences indicated that these strains are related to the genus Sinobacterium, showing sequence similarities between 96.97 and 97.63 % with the only species of the genus, Sinobacterium caligoides. Phenotypic characterization showed that the strains are typical marine halophiles, Gram negative, aerobic chemoorganotrophs, and allowed their differentiation from the closely related taxa. The G+C content of the novel strains was 52.2 ± 1 mol% and the predominant fatty acids were C16:0, C16:1 ω7c/C16:1 ω6c and C18:1 ω7c. The value for DNA-DNA relatedness between strain 3CM4(T) and the S. caligoides type strain LMG 25705(T) was 46  %. Hybridization values between strain 3CM4(T) and the other scallop isolates ranged from 82 to 93.6 %. Based on all data collected, the six scallop strains represent a novel species of the genus Sinobacterium, for which the name Sinobacterium norvegicum sp. nov., is proposed with strain 3CM4(T) (=CECT 8267(T); =CAIM 1884(T)) as type strain.

Pectinases from Sphenophorus levis Vaurie, 1978 (Coleoptera: Curculionidae): putative accessory digestive enzymes.

The cell wall in plants offers protection against invading organisms and is mainly composed of the polysaccharides pectin, cellulose, and hemicellulose, which can be degraded by plant cell wall degrading enzymes (PCWDEs). Such enzymes are often synthesized by free living microorganisms or endosymbionts that live in the gut of some animals, including certain phytophagous insects. Thus, the ability of an insect to degrade the cell wall was once thought to be related to endosymbiont enzyme activity. However, recent studies have revealed that some phytophagous insects are able to synthesize their own PCWDEs by endogenous genes, although questions regarding the origin of these genes remain unclear. This study describes two pectinases from the sugarcane weevil, Sphenophorus levis Vaurie, 1978 (Sl-pectinases), which is considered one of the most serious agricultural pests in Brazil. Two cDNA sequences identified in a cDNA library of the insect larvae coding for a pectin methylesterase (PME) and an endo-polygalacturonase (endo-PG)-denominated Sl-PME and Sl-endoPG, respectively-were isolated and characterized. The quantitative real-time reverse transcriptase polymerase chain reaction expression profile for both Sl-pectinases showed mRNA production mainly in the insect feeding stages and exclusively in midgut tissue of the larvae. This analysis, together Western blotting data, suggests that Sl-pectinases have a digestive role. Phylogenetic analyses indicate that Sl-PME and Sl-endoPG sequences are closely related to bacteria and fungi, respectively. Moreover, the partial genomic sequences of the pectinases were amplified from insect fat body DNA, which was certified to be free of endosymbiotic DNA. The analysis of genomic sequences revealed the existence of two small introns with 53 and 166 bp in Sl-endoPG, which is similar to the common pattern in fungal introns. In contrast, no intron was identified in the Sl-PME genomic sequence, as generally observed in bacteria. These data support the theory of horizontal gene transfer proposed for the origin of insect pectinases, reinforcing the acquisition of PME genes from bacteria and endo-PG genes from fungi.

Deep transcriptome sequencing of Pecten maximus hemocytes: a genomic resource for bivalve immunology.

Pecten maximus, the king scallop, is a bivalve species with important commercial value for both fisheries and aquaculture, traditionally consumed in several European countries. Major problems in larval rearing, however, still limit hatchery-based seed production. High mortalities during early larval stages, likely related to bacterial pathogens, represent the most relevant bottleneck. To address this issue, understanding host defense mechanisms against microbes is extremely important. In this study next-generation RNA-sequencing was carried on scallop hemocytes. To enrich for immune-related transcripts, cDNA libraries from hemocytes challenged in vivo with inactivated-Vibrio anguillarum and in vitro with pathogen-associated molecular patterns, as well as unchallenged controls, were sequenced yielding 216,444,674 sequence reads. De novo assembly of the scallop hemocyte transcriptome consisted of 73,732 contigs (31% annotated). A total of 934 contigs encoded proteins with a known immune function, grouped into several functional categories. Particular attention was reserved to Toll-like receptors (TLRs), a family of pattern recognition receptors (PRRs) involved in non-self recognition. Through mining the scallop hemocyte transcriptome, at least four TLRs could be identified. The organization of canonical TLR domains demonstrated that single cysteine cluster and multiple cysteine cluster TLRs co-exist in this species. In addition, preliminary data concerning their mRNA level following bacterial challenge suggested that different members of this family could exhibit opposite responses to pathogenic stimuli. Finally, a global analysis of differential expression comparing gene-expression levels in in vitro and in vivo stimulated hemocytes against controls provided evidence on a large set of transcripts involved in the great scallop immune response.

Respiratory response to combined heat and hypoxia in the marine bivalves Pecten maximus and Mytilus spp.

Coastal ecosystems are increasingly disturbed by the increase of mean sea surface temperature and expansion of hypoxic areas. The objectives of the present work were to describe and compare the respiratory responses to combined heat and hypoxia in two bivalve species (Pecten maximus and Mytilus spp.) living in two contrasted coastal habitats (subtidal and intertidal, respectively). Results were consistent with the vertical zonation of both species. Mytilus spp. seemed to cope better with a temperature increase than P. maximus, which was found to be outside of its optimal thermal window at 25°C. Concerning respiratory responses to hypoxia at a given temperature, P. maximus displayed greater oxyregulation capacity that was maintained over a larger range of O2 levels, as compared to Mytilus spp. When acclimation temperatures increased, both species showed a decrease in their oxyregulation capacities alongside a reduction in aerobic performance, especially in P. maximus. The comparison between species suggests that subtidal species, such as P. maximus, might be more vulnerable to a combination of heat and hypoxia than intertidal species, such as Mytilus spp. Lastly, this study highlighted the utility of segmented linear models to estimate PcO2 and regulation percentages in marine organisms exposed to hypoxia.