SNP discovery and gene annotation in the surf clam Mesodesma donacium

The main objective of this research was to identify single‐nucleotide polymorphisms (SNPs) from an Expressed Sequences Tags (EST) data set generated by 454 pyrosequencing in the soft clam Mesodesma donacium. A total of 180 159 ESTs were yielded from a M. donacium cDNA library. De novo assembly was performed using stringent calling parameters, producing 10 178 contigs and 41 765 singletons. Here, a total of 2594 SNPs were discovered related to 613 consensus sequences, achieving a frequency of 1 SNPs per 260 bp. SNP variants showed that A/G, A/T and C/T were the most abundant among the identified polymorphisms. We validated a total of 12 SNPs loci by HRMA for annotated genes such as heat shock protein‐70 and the translation elongation factor 1‐alpha. The Gene Ontology analysis regarding molecular function level revealed that sequences with SNPs were mainly classified to protein and nucleotide binding, as well hydrolase activity, ion binding and oxidoreductase activity. Further, biological processes like cellular and metabolic process, biogenesis, localization and biological regulation were highly annotated. The most expressed genes were related to the mitochondrial electron transport chain, senescence‐associated protein, ubiquitin and actin. Interestingly, some relevant genes related to immune response and biomineralization showed a high abundance, such as tumor necrosis factor (TNF)‐alpha‐receptor‐like protein, serine protease inhibitor, heat shock protein, aragonite‐binding protein and ferritin. This study contributes to relevant genes associated with functional polymorphisms and gives an overview for future genetic investigations.     

Biophysical models for Japanese scallop, Mizuhopecten yessoensis, aquaculture site selection in Funka Bay, Hokkaido, Japan, using remotely sensed data and geographic information system

Scallop aquaculture has grown to become the most successful marine shellfish farming venture in Japan. To ensure long-term sustainability of scallop production, suitable sites need to be selected. This study was conducted to identify the most suitable sites for hanging culture of Japanese scallop (Mizuhopecten yessoensis) based on measured biophysical parameters and modeled using geographic information system (GIS) models. Four biophysical parameters that influence Japanese scallop aquaculture in Funka Bay were selected: sea temperature, food availability (chlorophyll-a), suspended solid, and bathymetry. The SeaWiFS and MODIS images were used to extract most of these parameters. A series of GIS models was developed to identify suitable sites for scallop culture using a multi-criteria evaluation (MCE) known as weighted linear combination. Suitability scores were ranked on a scale from 1 (least suitable) to 8 (most suitable), and about 83% of the total potential area with bottom depths less than 60 m had scores of 7 and 8. The final model outputs were compared with field verification data and found to be consistent. Local sensitivity analysis was used to test the robustness of the model output. Based on this analysis, the order of importance of the variables affecting the model was as follows: suspended solid > chlorophyll-a > sea temperature.     

Evidence of clandestine harvest and failure of conservation policies for Argopecten purpuratus in the Rinconada Marine Reserve (Chile)

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Divergent selection and realized heritability for growth in the Japanese scallop,Patinopecten yessoensis Jay

In order to improve the production and accurately estimate response to selection, divergent selection for growth in shell height was conducted in a cultured population of the Japanese scallop Patinopecten yessoensis. Applying the same selection intensity ± 1.756 in upward and downward directions, three groups including two selected groups of Fast and Slow and one non‐selected Control group were created, which were reared under the same environmental conditions at any stage. Differences always significantly existed among the three groups (P<0.05), except for larvae at day 1 and at day 5, and in the order of Fast>Control>Slow. The average standardized response to selection (SR), realized heritability (h_R²) and genetic gain (GG) was 0.473%, 0.269% and 7.85% for the Fast group and 0.381%, 0.217% and 6.60% for the Slow group respectively. Moreover, significant differences (P<0.05) were detected between the fast and the slow lines in both SR and h_R², providing evidence for an asymmetric response in two directions. Performance in shell height is improved by 7.85% in the fast line after one generation selection, suggesting that mass selection for faster growth in a cultured population of the Japanese scallop is effective.     

Characterization of expressed sequence tag-derived single-nucleotide polymorphisms in the bay scallop <Emphasis Type="Italic">Argopecten irradians irradians</Emphasis>

We describe herein the discovery of 3905 putative single-nucleotide polymorphisms (SNPs) from the alignment of 4968 sequences in a bay scallop Argopecten irradians irradians expressed sequenced tag (EST) database. The observed frequency of SNPs was estimated at one every 118.2 bp of contig sequences. Thirty of the SNPs were chosen for validation by tetra-primer amplification refractory mutation system-polymerase chain reaction procedure, and 17 of them were polymorphic with minor allele frequency ranging from 0.016 to 0.484. BLASTX gave significant hits for all but one of the 17 genotyped SNP-containing contigs, 11 of which were located in coding regions and all resulted in a synonymous substitution. Parentage simulation demonstrated that SNP markers were more sensitive to the number of parents compared with microsatellites, thus more SNPs were needed to counteract low polymorphism. A table of optimal codons was deduced from the analysis of the A. i. irradians EST dataset to explain the frequency difference for a specific SNP. It seems that the selection of codon usage may be partly responsible for the frequency difference between the two alleles of the coding SNPs. These are the first SNPs developed for the bay scallop and will provide a useful complement to currently available genetic markers.     

Full‐length cloning and phylogenetic analyses of translationally controlled tumour protein and ferritin genes from the Indian white prawn,Fenneropenaeus indicus (H. Milne Edwards)

Elucidation, through molecular analyses, of bacterial afflictions in commercially important aquaculture‐reared shrimps is pivotal for the prevention and/or control of disease outbreaks. In this study, we examined the phylogenetic relatedness and compared the possible immune‐related functional roles of both translationally controlled tumour protein (TCTP) and ferritin genes with previous studies. Both TCTP and ferritin genes were substantially upregulated in the Indian white prawn, Fenneropenaeus indicus (H. Milne Edwards), post‐larvae following bath challenge with the virulent strain of bacteria, Vibrio harveyi D3. Full‐length cloning of these genes by rapid amplification of complementary DNA ends ‐polymerase chain reaction (RACE‐PCR) yielded 727‐base pair (bp)‐long TCTP and 1212‐bp‐long ferritin gene sequences. Their open reading frames (ORFs) were 507 and 510 bp, respectively. The TCTP‐ORF coded for 168 amino acids with three substitutions at positions 37, 141, 155, and the ferritin ORF coded for 170 amino acids with no species‐specific substitutions. Phylogenetic analysis suggested the closest relatedness of both TCTP and ferritin from F. indicus to Chinese white prawn, Fenneropenaeus chinensis (Osbeck). In addition to reporting the full‐length sequences of these immune‐relevant genes, this study highlighted their conserved natures, which perhaps make them important defence‐related proteins in the innate immune system of F. indicus.     

Expression of four muscle proteins at different growth stages of Günther's walking catfish Clarias macrocephalus

The complete cDNA sequences of four contractile muscle genes of walking catfish Clarias macrocephalus were characterized by assembling partial EST sequences from a skeletal muscle cDNA library. The four genes were parvalbumin 4 (PV4) (670 bp), troponin C (TnC) (1065 bp), troponin I (TnI) (843 bp) and myosin light chain 3 (MLC3) (953 bp), leading to deduced amino acid sequences of 109, 160, 176 and 150 residues respectively. During the larval stage, TnC mRNA showed the highest levels of expression with a 1.4‐fold increase from day 1 to day 30 post hatch. At 90 days, the relative expression levels of PV4, TnC and MLC3 were the highest, with similar proportions in the skeletal muscle, corresponding to the highest relative growth rate of walking catfish. Expression of the three calcium‐binding proteins remained high in 6‐month‐old fish, with higher levels of PV4. The different proportions of muscle proteins expressed suggested the significance of their contributions to fish growth and appeared to be correlated with the functional properties of muscle cells, which were observed from changes in the swimming activity of the fish.     

The recruitment of scallops (and beyond) by two different artificial collectors (Gulf of Taranto,Mediterranean Sea)

This study provides for the first time an evaluation of the natural availability of scallop seeds along the coastal area of Taranto (Mediterranean Sea, Southern Italy). To select the best artificial collectors to harvest scallop seeds in this area, cylindrical collectors (Cyl) were compared to traditional ‘Japanese‐style onion bags’ (Bag) across three sites. Scallops represented 26.6% of total bivalve recruitment among all collectors (782 ± 331 ind. m^?2). The most recruited scallops were Flexopecten glaber and Mimachlamys varia. The white (F. glaber) and black scallops (M. varia) were abundant at all three sites, while the queen scallop, Aequipecten opercularis, was only found at one site. Beyond the recruitment of scallops, numerous other potential commercial bivalve species were collected, including Limaria tuberculata, which was the most abundant bivalve species (33.3% of total recruitment). At all sites, Bag collectors had higher recruitment than Cyl collectors. Scallops recruited in artificial collectors exhibited a multicohort size distribution at most sites, suggesting potentially continuous reproductive activity in the scallop populations. F. glaber, M. varia and L. tuberculata could be good candidates to diversify the aquaculture production because of the availability of seed and their growth rate. Moreover, the farming of these bivalves integrates well with other forms of aquaculture (e.g. mussels), thus potentially offering farmers an additional source of income.     

Patterns and impacts of fish bycatch in a scallop dredge fishery

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Stress response and lipid composition in shrimp Litopenaeus vannamei fed diets enriched with squid or scallop viscera meal

We substituted 10% of fishmeal with scallop (Argopecten ventricosus) or squid (Dosidiscus gigas) by‐product meal in a feed for shrimp (Litopenaeus vannamei) growth assay that lasted 60 days and then shrimps were submitted to handling stress. Shrimp fed the control diet had smaller weight gain and lower recovery after stress. Prostaglandins in gills were affected by diet and stress, with significant differences in shrimp fed the control diet before and after the stress. Hydroperoxides were increased after stress, while isoprostanoids were higher in shrimp fed the control diet. The gene expression of Glutathione peroxidase and Catalase was increased on the stressed shrimp fed the scallop diet, with no differences among diets for Superoxide dismutase or Cyclooxygenase. Arachidonic acid in the phospholipid fraction of muscle was the only fatty acid affected by stress and diet. Cholesterol oxidation products were significantly higher in shrimp fed the scallop diet compared to shrimp fed the control diet before applying the stress. A PCA analysis grouped in one‐factor growth variables, as well as Superoxide dismutase, esterified astaxanthin and isoprostane, while another factor was determined by stress, prostaglandin and hydroperoxide levels in gills. The partial substitution of fishmeal with scallop of squid by‐product meals affected growth and stress differently, indicating shrimp that grew up more were not necessarily more tolerant to handling stress, which can have a strong repercussion in aquaculture, particularly during density control, recording of growth and harvest.     

Evaluation of hemato-immunological parameters during the reproductive cycle of the scallop Nodipecten nodosus in association with a carotenoid-enriched diet

The cultivation of scallops Nodipecten nodosus is a promising activity emerging in Brazil. The purpose of this study was to characterize the immune system of N. nodosus and evaluate the modulation of some hemato-immunological parameters during the reproductive cycle, in association with an astaxanthin-enriched diet. It was hypothesized that a supplementation on astaxanthin could enhance scallop immune system and minimize stress of reproduction. Scallops were separated in different groups: juveniles (J), adults (A), sexually mature (M), and recently spawned (S) animals. The last two groups were fed standard (M and S) or astaxanthin-enriched (Ma and Sa) diets. Scallop hemolymph contained two hemocyte populations: hyaline (HH) and granular hemocytes (GH). Antimicrobial peptides, similar to mussel defensins and mytilins, were found by immunodetection within the GH granules, even though the scallop hemolymph did not exhibit significant antimicrobial activity against different bacteria, including marine vibrios. Scallop hemocytes were able to phagocytose zymosan and produce reactive oxygen intermediates (ROI–NBT reduction). The number of circulating hemocytes (Neubauer chamber) varied from 12 to 26.10⁶ cells mL^− 1, and the GH was always the predominant cell type (67–99%). The plasma of N. nodosus contained lectins, specific to galactose and sialoconjugates, and their agglutinating activity was partially calcium-dependent. A phenoloxidase (PO) activity (146–446 U min^− 1 mg^− 1) was observed in the scallop hemolymph. However, this activity was not induced by trypsin or components of microorganism surface, and was strongly enhanced by alkaline pH (≥ 8.5). The total protein concentration of the plasma varied from 240 to 660 μg mL^− 1. In general terms, all examined hemato-immunological parameters (hemograms, superoxide anion production, PO activity, lectin titers and total protein concentration) had a similar profile during all the scallop reproductive cycle. Their levels increased significantly from juveniles to adults (except PO activity), and declined markedly (immune depletion) in the sexually mature scallops. After spawning, the animals had a tendency to recover the standard levels of their immune parameters. Apparently, the astaxanthin-enriched diet had no effect on the tested immune parameters except for a slight influence on the scallop immune-oxidative reactions (ROI production). The results obtained in this study suggested the occurrence of a general immune depletion in the sexually mature scallops, confirming that the reproductive stage is a critical period in scallop life.     

Predation risk management of sea stars (Asterias amurensis and Distolasterias nipon) by adjusting the density and size of seeded scallops (Mizuhopecten yessoensis): an improvement to local mariculture

Optimizing the release density and size of juvenile commercial species for local ecosystems is a critical issue that should be considered when countering predation impacts. To ascertain whether mariculture production of the Japanese scallop (Mizuhopecten yessoensis) could be enhanced by modifying releasing practices, we experimentally investigated the effects of density and size of scallop seeds on their survival in the presence of two sea star species, Asterias amurensis and Distolasterias nipon, with different predation capacities. Under current mariculture practices, the juveniles are briefly exposed to air just before release; therefore, we also examined how air exposure stress increased the predation risk. Scallop survival in the presence of both sea stars increased by?>?20% by increasing scallop size from 30 to 50 mm. Increasing scallop density (range: 5 to 30 scallops m^?2) increased each individual’s survival in the presence of A. amurensis but had no significant effect on predation by D. nipon. Therefore, the release of smaller quantities of large-sized scallops rather than larger quantities of small scallops is recommended to reduce D. nipon predation. In the presence of sea stars, especially by D. nipon, the predation impact on small scallops increased after just a few hours of air exposure, indicating that air exposure of juvenile scallops should be minimized. Our results will mitigate the economic cost of mariculture by facilitating the determination of optimal release conditions for juvenile scallops.

     

Feeding behavior and food consumption of Japanese whelk, <Emphasis Type="Italic">Buccinum isaotakii</Emphasis> (Neogastropoda: Buccinidae)

Abilities of detection and preference of food by a Japanese whelk, Buccinum isaotakii, were investigated under laboratory conditions. Consumption per food item and the corresponding assimilation efficiency were also de termined. Whelks were more sensitive to fresh foods than boiled foods, detecting them in a short period of time varying between 3.8 (±0.8) and 23.1 (±3.1) minutes. B. isaotakii preferred fish, polychate and bivalves and the time spent on these foods was more than 10 min as compared with those of shrimp (2.0 min) and squid (1.0 min). The highest daily consumption rate was observed on sardine, followed by scallop, polychaete and oyster. The relationship between shell length and consumption of different food items was positive. The coefficients of determination were relatively high for sardine, scallop, polychaete and oyster (r ², between 0.46 and 0.51) but not for clam, mussel shrimp and squid (r ², between 0.003 and 0.176), suggesting that the former could be predators of consumption for B. isaotakii. Observed were relatively high assimilation efficiencies on sardine, scallop, polychaete, and oyster, ranging from 30 to 70%. The present results are the intial information pertaining to the feeding biology of this whelk, thus, in-depth studies like an energy budget are recommended to deepen the understanding about seed production and resource management.     

Effect of emerged shipment on the physiological condition of the adductor muscle in adult giant lion's paw scallop Nodipecten subnodosus (Sowerby 1835)

Currently, there is an increasing interest in shipping live adult scallops to markets and broodstock to hatcheries; nevertheless, information about the shipping effect on live scallops physiology is very scarce. In a previous study, a method in emersion was developed to ship scallop seed out of water, but it was not known whether this method is useful to transport adult organisms. As a consequence, the purpose of this study was to evaluate the effects of transportation by the emersion method on the physiological status of the adductor muscle of adult giant lion's paw scallop. Live specimens were packaged in a container and transported in emersion for 11 h. Six scallops were frozen in a farm and a similar number were frozen as soon as they arrived to the laboratory. Physiological indices were determined in each lot and the survival was estimated 24 h after re‐immersion. As a result of the transportation, a significant loss (P<0.05) of total carbohydrates, glycogen, adenosine 5′‐triphosphate and adenilated energetic charge, and a significant increase (P<0.05) in the free amino acids concentration were observed. Eighty‐eight per cent survival was achieved; therefore, we conclude that this method is appropriate for shipping live adult scallops of Nodipecten subnodosus.     

Effects of hydrodynamic factors on Pecten maximus larval development

Hatchery production of great scallop, Pecten maximus, remains unpredictable, notably due to poor larval survival. Large‐scale flow‐through systems up to 3500 L have been developed to avoid the use of antibiotics in static systems. Alternatively, small‐scale flow‐through systems have been successfully applied for oysters but they proved to be unsuitable to rear scallop larvae. By focusing on physical factors presumed to limit P. maximus larval development, this study aimed to optimize great scallop larvae rearing parameters under controlled conditions. First, the influence of aeration on larval performances, energetic metabolism and antioxidant defences were studied both in static and flow‐through systems. Aeration depressed larval food intake, regardless of the intensities of flow tested (100 ml/min, 155 ml/min and 270 ml/min). On the other hand, antioxidant enzyme activities remained constant or decreased, suggesting that antioxidant defences were inactivated. The increase in citrate synthase activity suggested an increase in metabolic rate possibly due to a turbulent stressful environment. All larvae exposed to such turbulence died before reaching metamorphosis, whereas those reared without aeration survived well (≈ 95%). The effects of water renewal were thereafter studied in 50‐L flow‐through flat‐bottomed tanks. No differences in survival (20.4 ± 0.5%), growth (3.8 ± 0.2 μm/d), competence (5.6 ± 0.2%), energetic metabolism level and antioxidant enzyme activities were observed when comparing 12.5 and 25 L/hr water renewal. Whereas air bubbling leads to detrimental effects, flow‐through in small flat‐bottomed tanks appears to be a suitable technique for scallop larvae rearing.