Effect of temperature on embryonic development of the marine gastropod Charonia seguenzae (Aradas & Benoit, 1870).

The Mediterranean Triton Charonia seguenzae (Aradas & Benoit, 1870) is an endangered marine gastropod. Re-establishment of C. seguenzae populations in the depleted habitats requires knowledge of its biology and breeding in captivity. Temperature has been recorded to affect the development in marine gastropods. The present study aims to describe the encapsulated development and study the effect of three temperature regimes, that the embryos are exposed to in the wild (17, 20, 23οC), on it. At the stage of morula (7th Day After Deposition - DAD) 180 egg capsules were separated in three groups of 60 capsules. One group remained at 23 °C and the other two were acclimated at 20 and 17 °C. Two capsules were sampled randomly from each temperature setup (every day during the first 5 days, D1-D5, every other day from D7 to D17 and every other two days from D20 to eclosion), opened and the eggs, embryos or larvae were photographed under stereoscope. Stages of development and shape were assessed and dimensions were measured from microphotographs. All developmental stages were described in relation to temperature and time. At 23οC eclosion of free swimming veligers occurred 49 DAD, 17 days faster than the embryonic development at 20οC. Eclosion at 17 οC was not observed up until the 74th DAD when the last sampling was conducted. An increased amount of larval deformities was observed at 17οC reaching 94% during the last sampling (D74), while at eclosion only 4 and 3% of the hatching larvae were deformed at 20 and 23 οC respectively. In this study temperature appears as a key factor during the development of the marine gastropod Charonia seguenzae, affecting the survival and developmental rate. Although temperature affected the size of intermediate stages, the size of free swimming veligers at 20 and 23 οC did not differ.

Effect of temperature on the development of deformities during the embryonic stages of Charonia seguenzae (Aradas & Benoit, 1870).

The Mediterranean Triton Charonia seguenzae (Aradas and Benoit, 1870) is an endangered marine gastropod. Re-establishment of C. seguenzae populations in the depleted habitats requires knowledge of its biology and breeding in captivity. Deformities have a huge impact on offspring survival and quality. Temperature has been recorded to affect the development of deformities in marine gastropods. The present study aims to identify the stage of development at which deformities occur, under four temperature regimes (17, 23, 26 and 29 °C). At the stages of trochophore, veliger and free veliger larvae, three capsules that were acclimated at the examined temperatures at the stage of morula were collected, opened and 50 larvae per capsule sampled. Deformities were observed at every examined developmental stage under all tested temperatures. The lower rate of deformities at every stage occurred at 23 °C. The higher tested temperature (29 °C) was lethal and at the lower tested temperature (17 °C) almost every specimen was deformed (96.66%) at eclosion. The effect of acclimation at four developmental stages (morula, trochophore, shell formation and veliger) on the development of deformities at the free veligers of Charonia seguenzae, was studied under three temperature conditions (17, 26 and 29 °C). At eclosion, three capsules were collected, opened and 50 larvae per capsule were sampled. The acclimation at morula and trochophore larva stages led to the higher rates of deformities at eclosion. The size of the free veliger larvae was also affected by temperature with maximum size at eclosion observed at 23 °C. Charonia seguenzae's embryos tolerate elevated temperatures within environmental limits (26 °C) but near future global warming will probably pose a threat to their survival. The free veliger larvae survival at the environmental minimum is related to the time window of the acclimation, since Triton's embryos are more vulnerable to temperature alterations during the early developmental stages.

Indicator PCBs in farmed and wild fish in Greece - Risk assessment for the Greek population.

Health benefits of fish consumption could be counterbalanced by the intake of contaminants after long term fish consumption, burdened even in trace levels. The presence of the indicator PCBs (NDL-PCBs and PCB 118) in farmed and wild seabream and seabass was evaluated. For the determination of PCB, a GC-MS method was developed and evaluated. The association of PCB accumulation in fish with seasonality, locality, production mode and species was also investigated. A new approach for the risk characterisation after exposure to NDL-PCB through fish consumption in Greece was developed, based on the real exposure and the permitted maximum levels of both aggregated dietary exposure and exposure through fish consumption. PCB levels determined in fish were below established permitted limits (6.24 ng/g 95th percentile), while PCB levels and congener distribution varied significantly between farmed and wild fish (p = 0.001). Seasonality was highlighted as an important factor affecting NDL-PCBs accumulation, with high levels coinciding with the reproduction period of each species. Differences were also depicted for sampling sites, with PCB 118 presenting significantly higher values in open seas while NDL-PCB congeners in closed seas. Risk assessment of NDL-PCB intake through fish consumption corrected for the aggregated exposure revealed no risk for the consumers.

Microbial strains isolated from CO2-venting Kolumbo submarine volcano show enhanced co-tolerance to acidity and antibiotics.

As ocean acidification intensifies, there is growing global concern about the impacts that future pH levels are likely to have on marine life and ecosystems. By analogy, a steep decrease of seawater pH with depth is encountered inside the Kolumbo submarine volcano (northeast Santorini) as a result of natural CO2 venting, making this system ideal for ocean acidification research. Here, we investigated whether the increase of acidity towards deeper layers of Kolumbo crater had any effect on relevant phenotypic traits of bacterial isolates. A total of 31 Pseudomonas strains were isolated from both surface- (SSL) and deep-seawater layers (DSL), with the latter presenting a significantly higher acid tolerance. In particular, the DSL strains were able to cope with H+ levels that were 18 times higher. Similarly, the DSL isolates exhibited a significantly higher tolerance than SSL strains against six commonly used antibiotics and As(III). More importantly, a significant positive correlation was revealed between antibiotics and acid tolerance across the entire set of SSL and DSL isolates. Our findings imply that Pseudomonas species with higher resilience to antibiotics could be favored by the prospect of acidifying oceans. Further studies are required to determine if this feature is universal across marine bacteria and to assess potential ecological impacts.

Cadmium, lead and mercury in muscle tissue of gilthead seabream and seabass: Risk evaluation for consumers.

Cadmium (Cd), lead (Pb) and mercury (Hg) presence was investigated in the muscle tissue of gilthead seabream and seabass, collected from various aquaculture sites of the Aegean and Cretan Sea as well as from the fish market (fisheries). Risk for the Greek population through consumption of these species was estimated using two approaches: Target Hazard Quotient (THQ) and Hazard Index (HI). All heavy metal levels in the fish tissue were below the established safe limits for consumption. Metal accumulation was found to differ amongst mode of production, species, location and seasonality. Seabass demonstrated higher Hg and lower Cd concentrations than seabream, Hg and Pb seem to be more accumulated in closed seas and Pb values displayed a linear increasing trend from warmer to colder periods. Regression analysis revealed that the main contributing factor to Cd accumulation is species (beta: -0.28, 95%CI: -0.48 to -0.09); lead is predominately affected by seasonality (beta: 0.44, 95%CI: 0.29 to 0.59), Hg accumulation is mainly affected by location (beta: -0.32, 95%CI: -0.61 to -0.03) while wild seabream accumulates greater levels for Hg and Pb than farmed. Risk analysis demonstrated that consumption of the studied species, is safe for all metals (HI < 0.460 and TTHQ < 0.299).

Nonlinear responses to waterborne cadmium exposure in zebrafish. An in vivo study.

Cadmium (Cd) has proved to be associated with numerous toxic effects in aquatic organisms via waterborne exposure. With a view to investigate Cd toxicity along a broad spectrum of exposures reaching from environmental to toxic, we employed adult zebrafish (Danio rerio) for an in vivo study. A number of 10 fish per tank were placed in 40L tanks and were exposed for 30 days to 0.0, 5.0, 25, 50, 75, 100 and 1000µgCd per liter. There were 2 tanks for each Cd exposure (duplicate experiment). Mortality was recorded daily, dead fish were collected and tissue samples were obtained for histologic observation, whereas remaining tissues were stored for Cd burden determination. Surviving fish were collected at the end of the experiment. Median overall survival (OS) in days was found to be 9.0, 11.0, 8.0 and 7.0 for 25µg/L, 50µg/L, 75µg/L and 100µg/L respectively, with all of them showing mortality greater than 50%. Remarkably, fish exposed to the highest Cd concentration (1000µg/L) survived the longest exhibiting a mean OS of 29.2 days. Cd determination in fish tissue was conducted with an in house ICP-MS method and levels ranged from 3.1 to 29.1ng/mg. Log Cd tissue levels were significantly correlated with the log Cd exposure levels (r = 0.535, p < 0.001). The highest Cd burden was determined for fish exposed to 1000µgCd /L (mean = 12.2ng/mg). Histopathology supported these results. Our findings disclose a deviation in toxic responses through the range of Cd concentrations, leading to nonlinear responses. These differentiated responses, could be linked to hormesis phenomena.

Effects of graded dietary levels of soy protein concentrate supplemented with methionine and phosphate on the immune and antioxidant responses of gilthead sea bream (Sparus aurata L.).

The effects of a dietary soy protein concentrate (SPC) as a fish meal (FM) substitute, on selected innate immune responses, the oxidative status, hepatic and intestinal morphology of gilthead sea bream, Sparus aurata, were evaluated after a three-month feeding trial. Isonitrogenous (45% crude protein) and isoenergetic (23 kJ/g gross energy) diets with 20% (SPC20), 40% (SPC40) and 60% (SPC60) of SPC inclusion, supplemented with methionine and phosphate, were evaluated against a diet containing FM as the sole protein source. Diets were allocated in triplicate groups of 26-g fish (8 kg m-3/tank) and administered for three months. Immune responses were evaluated by performing immunological assays in blood (respiratory burst activity) and serum (myeloperoxidase content, bacteriolytic and lysozyme activity), as well as by gene expression analysis of immune-associated genes (MHCIIα, ß2m, CSF-1R, NCCRP-1, TGF-ß1, HSP70) in the head kidney and distal intestine. In addition, oxidative stress was evaluated by measuring the activity of liver enzymes associated with the antioxidant system. The respiratory burst activity of blood was significantly decreased in the SPC40 group, while serum myeloperoxidase content and bacteriolytic and lysozyme activities were affected. Significantly higher expression levels of NCCRP-1 and HSP70 were found in SPC60 head kidneys, while increased intestinal MHCIIα and NCCRP-1 transcripts were observed in SPC40. Hepatic antioxidant enzyme activity of glutathione reductase and glutathione-S-transferase was significantly enhanced in the SPC40 and SPC60 groups, while superoxide dismutase activity was increased only in the SPC40 group. Moreover, increased lipid accumulation in the enterocytes of the distal intestine was observed in the SPC60 group. Overall, a three-month feeding period with diets over 40% of dietary SPC inclusion as a FM substitute, indicated increases on immune and antioxidant enzyme responses, suggesting the dietary SPC levels that gilthead sea bream can tolerate.

Temperature sex determination in the European sea bass, Dicentrarchus labrax (L., 1758) (Teleostei, Perciformes, Moronidae): critical sensitive ontogenetic phase.

The temperature sex determination (TSD) mechanism in the European sea bass (Dicentrarchus labrax L.) was studied in respect to: a) the TSD sensitivity during the different developmental stages; and b) the intrapopulation correlation of sex determination with the growth rate up to the end of the TSD-sensitive period. At the stage of half-epiboly, eggs from the same batch were divided into four groups and subjected to different thermal treatments: a) 15 degrees C (G15 group) and b) 20 degrees C (G20 group) up to the middle of metamorphosis stage; c) 15 degrees C up to the end of yolk-sac larval stage and subsequently to 20 degrees C (G15-5 group); and d) 15 degrees C up to the end of the preflexion stage and then to 20 degrees C (G15-10 group). At the end of the treatments, size grading was applied and four additional populations were established from the upper (L) and lower (S) size portions of the G15 and G20 populations: G15L, G15S, G20L, and G20S. During the following growing phase, all populations were subjected to common rearing conditions. The sex ratios of each population were macroscopically determined at 190-210 mm mean total length. Female incidence was significantly affected (P < 0.05) by the different thermal treatments: 66.1% in the G15, 47.1% in the G15-10, 37.6% in the G15-5, and 18.1% in the G20 group. In addition, sex ratio was correlated with the growth rate of the fish up to the end of the TSD-sensitive period, with the larger fish presenting a significantly higher (P < 0.01) female incidence than the smaller fish in both thermal regimes tested: 73.1% in G15L vs. 57% in G15S, and 36.6% in G20L vs. 22.5% in G20S group. Results provide, for the first time, clear evidence that the sea bass is sensitive to TSD during all different ontogenetic stages up to metamorphosis, and that sex ratio is correlated with the growth rate of the fish well before the differentiation and maturation of the gonads.