Due to the high mortality rates and poor growth generally observed in Octopus vulgaris paralarval rearing experiments, it was decided to organize a working group in order to formulate recommendations to tackle this problem. Over a dozen scientists representing the most active current research groups related to this subject attended the meeting in Vigo, Spain, in November 2005. The aim of this working group was to determine the bottlenecks that prevent success in paralarval rearing, define the most appropriate rearing conditions, and identify required future research. This paper describes rearing techniques for the O. vulgaris paralarvae used by the different research participant teams, with regard to tank systems, feeding environment, and diets (Artemia, crustacean zoeae, sandeel flakes, copepods, etc.). Additionally, it includes other related themes such as the culture of Artemia and copepods, organisms that are commonly used in paralarval rearing. When embarking on O. vulgaris rearing it is advised to use prey rich in DHA (docosaenoic acid, 22:6n-3) and EPA (eicosapentaenoic acid, 20:5n-3), and with high DHA/EPA ratio. Such prey could be enriched Artemia, accompanied or not by crustacean zoeae or any microdiet. It is also recommended that, in future studies, values of growth and survival rates are recorded at the beginning of the benthic phase, in order to compare them to successful previous studies. Dry weight and DHA/EPA ratio of paralarvae may also be good criteria to define paralarval viability and evaluate success of the rearing system.

13 14To increase current knowledge on the nutritional value of natural prey organisms, the 15 biochemical components of mainly three copepods (Acartia grani, Centropages 16 hamatus, and Eurytemora affinis) from a marine pond system were analysed once a 17 week from spring until late fall, over two years. The analysed components were total 18 lipid, lipid class composition, total lipid fatty acid composition, free amino acids, total 19 protein, protein-bound amino acids, pigment (astaxanthin and ß-carotene), and 20 vitamins (A, thiamine, riboflavin, C, D 3 , and E). Copepod dry weight (DW), dry 21 matter (% of wet weight), and ash content (% of DW) were also determined. The data 22 are unique due to the homogenous content of copepods in the samples and the long 23 time span of sampling. The copepods were characterised by moderate levels of lipids 24 (6.9-22.5% of DW), with polar lipids accounting for 37.9 to70.2% of the total lipid. 25The most abundant fatty acids in total lipid (as % of total lipid) were 16:0 (palmitic 26 * Corresponding author. Tel.: +47 56182262; fax: +47 56182222. E-mail address: Terje.van.der.Meeren@imr.no A C C E P T E D M A N U S C R I P T , 8.3-24.6%), and 22:6n-3 (DHA, 13.9-42.3%). The 27 amount of 20:4n-6 (ARA) was generally low (0-2.6%), giving an EPA/ARA range 28 between 7.5and 49.5. The DHA/EPA ratio was between 1.0 and 4.9. ACCEPTED MANUSCRIPT2 acid, 10.8-17.1%), 20:5n-3 (EPA

Cod larvae, Gadus morhua L., were reared in the laboratory and released to a large marine enclosure4 to Sdaysafter hatching(6-8" C). Thedevelopment ofthedigestive system was studied until day 24 after hatching. Morphological investigations of the jaw apparatus and the digestive tract showed that the larvae are able to absorb ingested food well before exhaustion of the yolk sac. The foregut, and especially the midgut, were particularly active in lipid absorption, and the hindgut was characterized by pinocytotic activity. During the first days of feeding, no distinct prey organisms were observed in the gut. and signs of food absorption in the epithelial cells of the gut were sparse. A distinct red fluorescence, restricted to the hindgut, was observed from day I 1 to day 19. On the basis of changes in absorptive pattern in the gut we suggest that changes in digestive and absorptive abilities, as well as in nutritional needs, take place around days 15-17 after hatching.In starved larvae, signs of degeneration of the gut tissue were first visible in the foregut. By day 9 after hatching, microvilli was degenerated to such an extent that the ability to absorb food must have been severely restricted. Iflarvae are starved longer than this, they will probably not survive.

The current commercial production protocols for Atlantic cod depend on enriched rotifers and Artemia during first-feeding, but development and growth remain inferior to fish fed natural zooplankton. Two experiments were conducted in order to identify the underlying factors for this phenomenon. In the first experiment (Exp-1), groups of cod larvae were fed either (a) natural zooplankton, mainly copepods, increasing the size of prey as the larvae grew or (b) enriched rotifers followed by Artemia (the intensive group). In the second experiment (Exp-2), two groups of larvae were fed as in Exp-1, while a third group was fed copepod nauplii (approximately the size of rotifers) throughout the larval stage. In both experiments, growth was not significantly different between the groups during the first three weeks after hatching, but from the last part of the rotifer feeding period and onwards, the growth of the larvae fed copepods was higher than that of the intensive group. In Exp-2, the growth was similar between the two copepod groups during the expeimental period, indicating that nutrient composition, not prey size caused the better growth on copepods. Analyses of the prey showed that total fatty acid composition and the ratio of phospholipids to total lipids was slightly different in the prey organisms, and that protein, taurine, astaxanthin and zinc were lower on a dry weight basis in rotifers than in copepods. Other measured nutrients as DHA, all analysed vitamins, manganese, copper and selenium were similar or higher in the rotifers. When compared to the present knowledge on nutrient requirements, protein and taurine appeared to be the most likely limiting nutrients for growth in cod larvae fed rotifers and Artemia. Larvae fed rotifers/Artemia had a higher whole body lipid content than larvae fed copepods at the end of the experiment (stage 5) after the fish had been fed the same formulated diet for approximately 2 weeks.

The simultaneous effect of temperature (5, 7, 10 and 13°C) and light on the rates of oxygen consumption and ammonia excretion of larval and early juvenile Atlantic cod Gadus morhua was examined. Larvae increased their mean dry body mass by 2000 times within 48 d. Instantaneous growth rate exceeded 30% d -1 towards the end of the study period, and proportionality of growth followed a triphasic pattern, during which body water content significantly declined but no inflection could be detected in the metabolic exponents. Data were rigorously tested via Model-I (least squares) and Model-II (geometric mean) regression techniques, and the aerobic metabolic rate was found to scale allometrically with both dry and wet body mass. The metabolic exponent was not affected by increasing temperature, but was significantly decreased by the presence of light (b = 0.88 to 0.89 for light-adapted larvae; b = 0.90 to 0.91 for dark-adapted larvae). The effect of light on small larvae (4 to 7 mm standard length, SL) caused a 30 to 40% increase in metabolic rate, while no effect was observed in larger juveniles (40 to 60 mm SL). Acute temperature acclimation of Atlantic cod of 4 to 60 mm SL (0.04 to 350 mg dry mass) demonstrated normal thermal sensitivity with Q 10 values of 2.4 for dark-adapted larvae and 2.6 for light-adapted larvae. Rates of ammonia excretion also scaled allometrically with wet and dry body mass and showed greater variability in dark-adapted compared to light-adapted larvae. Comparison of the molar rates of ammonia excretion and oxygen consumption revealed that Atlantic cod larvae have a high reliance on amino acids as fuel for energy dissipation. With lipids as the assumed co-substrate, amino acids were estimated to account for 70 to 95% of total substrate oxidation for larvae up to 7 mm SL (first 3 to 4 wk of post-hatch development). Beyond 7 mm SL, the reliance on amino acids as fuel began to decline, but even in juveniles of 40 to 60 mm SL, amino acids still represented the dominant source of fuel. For juveniles of between 10 and 20 mm SL, both the rates of oxygen consumption and ammonia excretion remained unaffected by the presence of food in the gut. For short-term fasted juveniles (35 to 60 mm SL), however, a substantial decline in the rate of ammonia excretion was observed. This indicates that during short-term fasting (8 to 12 h) early juvenile Atlantic cod conserve amino acids, rather than funneling them into the tricarboxylic acid cycle.KEY WORDS: Scaling · Metabolism · Fuel preference · Free amino acid · Q 10 · Temperature · Cod larvae Resale or republication not permitted without written consent of the publisherMar Ecol Prog Ser 243: [217][218][219][220][221][222][223][224][225][226][227][228][229][230][231][232][233][234] 2002 1975). For the juvenile and adult stages of fishes, however, a general exponent of 0.8 has been proposed (Winberg 1956, Wieser 1995, Clarke & Johnston 1999. Giguère et al. (1988) have proposed that the metabolic rates of larval fishes scale isometrically with incre...

Recent studies have shown that crude oil exposure affects cardiac development in fish by disrupting excitation-contraction (EC) coupling. We previously found that eggs of Atlantic haddock (Melanogrammus aeglefinus) bind dispersed oil droplets, potentially leading to more profound toxic effects from uptake of polycyclic aromatic hydrocarbons (PAHs). Using lower concentrations of dispersed crude oil (0.7–7 μg/L ∑PAH), here we exposed a broader range of developmental stages over both short and prolonged durations. We quantified effects on cardiac function and morphogenesis, characterized novel craniofacial defects, and examined the expression of genes encoding potential targets underlying cardiac and craniofacial defects. Because of oil droplet binding, a 24-hr exposure was sufficient to create severe cardiac and craniofacial abnormalities. The specific nature of the craniofacial abnormalities suggests that crude oil may target common craniofacial and cardiac precursor cells either directly or indirectly by affecting ion channels and intracellular calcium in particular. Furthermore, down-regulation of genes encoding specific components of the EC coupling machinery suggests that crude oil disrupts excitation-transcription coupling or normal feedback regulation of ion channels blocked by PAHs. These data support a unifying hypothesis whereby depletion of intracellular calcium pools by crude oil-derived PAHs disrupts several pathways critical for organogenesis in fish.

The toxicity resulting from exposure to oil droplets in marine fish embryos and larvae is still subject for debate. The most detailed studies have investigated the effects of water-dissolved components of crude oil in water accommodated fractions (WAFs) that lack bulk oil droplets. Although exposure to dissolved petroleum compounds alone is sufficient to cause the characteristic developmental toxicity of crude oil, few studies have addressed whether physical interaction with oil micro-droplets are a relevant exposure pathway for open water marine speices. Here we used controlled delivery of mechanically dispersed crude oil to expose pelagic embryos and larvae of a marine teleost, the Atlantic haddock (Melanogrammus aeglefinus). Haddock embryos were exposed continuously to two different concentrations of dispersed crude oil, high and low, or in pulses. By 24 hours of exposure, micro-droplets of oil were observed adhering and accumulating on the chorion, accompanied by highly elevated levels of cyp1a, a biomarker for exposure to aromatic hydrocarbons. Embryos from all treatment groups showed abnormalities representative of crude oil cardiotoxicity at hatch (5 days of exposure), such as pericardial and yolk sac edema. Compared to other species, the frequency and severity of toxic effects was higher than expected for the waterborne PAH concentrations (e.g., 100% of larvae had edema at the low treatment). These findings suggest an enhanced tissue uptake of PAHs and/or other petroleum compounds from attached oil droplets. These studies highlight a novel property of haddock embryos that leads to greater than expected impact from dispersed crude oil. Given the very limited number of marine species tested in similar exposures, the likelihood of other species with similar properties could be high. This unanticipated result therefore has implications for assessing the ecological impacts of oil spills and the use of methods for dispersing oil in the open sea.

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