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The fatty acid (FA) profile of wild Octopus vulgaris paralarvae of estimated age was individually analyzed for the very first time in order to establish a reference for comparison in rearing and nutritional studies. Age of each paralarvae was estimated by analysing daily increments on lateral hood surface of beaks. Wild paralarvae age ranged between 6 and 8 days and their FA composition resembled that from hatchlings produced under culture conditions. However, when compared with the FA composition of up to 20 days old cultured paralarvae described in the bibliography, some striking differences were found. Results showed higher levels of docosahexaenoic acid (22:6n − 3, DHA), lower contents of 18:1n − 9, 18:1n − 7 and 18:2n − 6 and negligible levels of 18:3n − 3 in wild paralarvae, when collated to reared one. These results seem to indicate that preys/diets supplied to cultured paralarvae fail to resemble paralarval natural composition and as a result do not fulfil their FA requirement. The individual applied technique developed in this study will allow to refine the study of wild paralarvae along its development, as well as to compare wild and cultured paralarvae of similar age. Statement of relevance Artemia does not fulfil paralarval fatty acid requirements. ; This study was funded by Spanish Government under Projects OCTOPHYS (Ref. AGL2010-22120-C03), OCTOWELF (Ref. AGL2013-49101-C2-1-R) and LARECO (CTM2011-25929). Most of the authors of this paper participate to the COST network, Action FA1301 (CephsInAction) focussing on Cephalopod welfare. D. Garrido was financed by a Ph.D. grant by Spanish Institute of Oceanography (BOE 3rd November 2011). ; Peer Reviewed

13 pages, 3 tables, 8 figures ; The common octopus Octopus vulgaris is a highly valuable species worldwide, but to understand its population dynamics and requirements under culture conditions, it is crucial to improve our knowledge about its planktonic stages. Previous studies validating daily beak growth increments in these stages allowed age estimation and comparison of wild and cultured paralarvae. We aimed to improve age estimations in captivity, addressing the effect of temperature and diet, to obtain an accurate estimation of age in wild specimens collected from the coast to the open ocean off NW Spain and Morocco. We analysed the beak growth increments of reared paralarvae at 14 and 21°C with 2 different crustacean prey taxa (Artemia and spider crab Maja brachydactyla zoeae) over 30 d. Daily increment deposition at 21°C was confirmed, whereas <1 increment d−1 was recorded at 14°C. The width of the reading area grew accordingly with age; therefore, this beak region may be suitable for age estimation. A general linear model (GLM) analysis showed that temperature and the interaction of age × temperature significantly influenced increment deposition, whereas diet did not. The number of growth rings recorded in wild paralarvae beaks ranged from 0−8 on the coast, 7−11 on the continental shelf and 2−28 in the open ocean. Corrected age estimates of wild paralarvae were obtained with the GLM using the mean temperatures recorded in the wild, supporting the hypothesis that O. vulgaris leave the coastal area and develop in the open ocean transported by upwelling filaments. ; This study was funded by Spanish Government under Projects OCTOWELF (Ref. AGL2013- 49101-C2) and CAIBEX (CTM2007-66408-C02). ; Peer reviewed

Nowadays, the common octopus (Octopus vulgaris) culture is hampered by massive mortalities occurring during early life-cycle stages (paralarvae). Despite the causes of the high paralarvae mortality are not yet well-defined and understood, the nutritional stress caused by inadequate diets is pointed out as one of the main factors. In this study, the effects of diet on paralarvae is analyzed through a proteomic approach, to search for novel biomarkers of nutritional stress. A total of 43 proteins showing differential expression in the different conditions studied have been identified. The analysis highlights proteins related with the carbohydrate metabolism: glyceraldehyde-3-phosphate-dedydrogenase (GAPDH), triosephosphate isomerase; other ways of energetic metabolism: NADP+-specific isocitrate dehydrogenase, arginine kinase; detoxification: glutathione-S-transferase (GST); stress: heat shock proteins (HSP70); structural constituent of eye lens: S-crystallin 3; and cytoskeleton: actin, actin-beta/gamma1, beta actin. These results allow defining characteristic proteomes of paralarvae depending on the diet; as well as the use of several of these proteins as novel biomarkers to evaluate their welfare linked to nutritional stress. Notably, the changes of proteins like S-crystallin 3, arginine kinase and NAD+ specific isocitrate dehydrogenase, may be related to fed vs. starving paralarvae, particularly in the first 4 days of development. ; This study was funded by the "Ministerio de Economía y Competividad (Spanish Government)" under the projects OCTOPHYS (AGL-2010-22120-CO3-02) and OCTOWELF (AGL2013-49101-C2-2-R), and by the "Generalitat Valenciana" under the project PROMETEO II/2014/085. We acknowledge support of the publication fee by the CSIC Open Access Support Initiative through its Unit of Information Resources for Research (URICI) ; Peer reviewed ; Peer Reviewed

Normal development, growth and the maintenance of health and well-being are only possible if all the digestive tract functions (e.g., motility, digestion, and absorption) operate normally and in concert. Understanding the physiological processes and the impact of external factors (e.g., handling, temperature, diet quality including exposure to food toxins, exposure to viral/bacterial infections and parasites) is important for normal laboratory maintenance of the animal in a research setting, as well as for optimizing conditions for aquaculture at each life stage. The study of the physiology of the cephalopod digestive apparatus has mainly focused on Sepia officinalis (Bidder, 1966; Boucaud-Camou and Boucher-Rodoni, 1983; Mangold and Bidder, 1989; Quintela and Andrade, 2002a,b; Sykes et al., 2013; Costa et al., 2014), Octopus vulgaris (Boucher-Rodoni and Mangold, 1977; Boucaud-Camou and Boucher-Rodoni, 1983; Andrews and Tansey, 1983b; Mangold and Bidder, 1989), Octopus maya (Martínez et al., 2011a,b, 2012; Rosas et al., 2013; Linares et al., 2015; Pech-Puch et al., 2016). Few studies have been carried out in Loligo vulgaris and other squid (Bidder, 1950; Mangold and Bidder, 1989). Furthermore, the morphology, motility and absorptive functions of the digestive tract of Nautilus pompilius have been the subject of limited investigation (Westermann and Schipp, 1998a,b, 1999; Ruth et al., 1999; Westermann et al., 2000, 2002). The inclusion of all "live cephalopods," taken to mean all living species (about 700), at all life stages after hatching, in Directive 2010/63/EU (European Parliament and Council of the European Union, 2010) covering the use of animals in scientific research and education poses a number of challenges for research (Smith et al., 2013; Fiorito et al., 2015) including that aimed at optimizing practices in aquaculture (Sykes et al., 2012; Smith et al., 2013; Fiorito et al., 2015). Whilst the Directive regulates studies in the Member States of the European Union, the principles it enshrines and the ...

The effects of culture conditions on growth, survival, predation, and nutritional composition of the mysid Gastrosaccus roscoffensis were studied. Light had a negative effect on the predation rates of G roscoffensis and predator size was important to prey on Artemia nauplii and rotifers (Brachionus plicatilis). Growth rates were higher in the first three weeks and measured individuals increased almost 7 mm in length in six weeks. Mortality rate was not constant, but it peaked during the first 10 days and after the 40th day of culture. Lipid class composition of cultured G. roscoffensis juveniles showed differences compared to wild G roscoffensis and other mysid species, with a high proportion of neutral lipids (72.04% total lipids) mainly triacylglycerol (41.74%). Fatty acid composition was characterized by high levels of 18:3n-3 (23.16% total lipids) and monoenes in cultured G. roscoffensis. The experiments demonstrated the feasibility of the culturing conditions assayed for G roscoffensis, although further experiments should be carried out to test this mysid as a prey for new species of commercial interest, including fish and cephalopods. ; research project PRESAPUL [PI SolsubC200801000162]; Canarian Government (ACIISI); research project NUTRIPULPO; JACUMAR (Spanish Government) ; info:eu-repo/semantics/publishedVersion

The high mortalities observed during Octopus vulgaris paralarvae culture have been associated with a nutritional imbalance, with long-chain polyunsaturated fatty acids (LC-PUFA) appearing to have a critical role. In order to determine the in vivo capability of O. vulgaris hatchlings to incorporate and metabolise unsaturated fatty acids (FA), hatchlings were incubated in flat-bottom 6-well tissue culture plates at a density of 90 hatchlings/well in 10mL of seawater (36‰). Incubations were performed with gentle stirring at 21°C for 6h with 0.2μCi (0.3μM) of [1-14C]-labelled FA including 18:1n-9, 18:2n-6, 18:3n-3, 20:4n-6 (ARA), 20:5n-3 (EPA) or 22:6n-3 (DHA), which were added directly to the seawater as their potassium salts bound to bovine serum albumin (BSA). A control treatment without [1-14C]FA was also assessed. O. vulgaris hatchlings not only possessed the ability to incorporate FA bound to BSA, but also to esterify them into phospholipid, with marked specificity. [1-14C]DHA and [1-14C]C18 FA substrates were mainly esterified into phosphatidylcholine, while [1-14C]ARA and [1-14C]EPA were esterified into phosphatidylethanolamine. The majority of radioactivity from [1-14C]FA incorporated into hatchling total lipid was recovered as unmodified FA with elongation being the only metabolism detected. Of the FA investigated, [1-14C]ARA was the most efficiently incorporated into hatchling lipids, but it was also the least modified FA. The fact that no desaturation activity was recorded towards the FA tested in this experiment may indicate that the nutritional requirements of O. vulgaris hatchlings in terms of FA are highly specific and LC-PUFA must be considered essential dietary nutrients. © 2014 Elsevier B.V. ; This work was partially supported by project OCTOPHYS. Ref. AGL 2010-22120-CO3 funded by the Spanish Government. D.B. Reis (SFRH/BD/76863/2011) and A.V. Sykes (SFRH/BPD/36100/2007) wish to thank Fundação para a Ciência e a Tecnologia for their grants. ; Peer Reviewed

Comunicación presentada en las CephsInAction MC & WGs meetings, celebradas en Barcelona, España, del 14 al 16 de marzo de 2014 ; This work was partially supported by project OCTOPHYS. Ref.AGL2010-22120-CO3 funded by Spanish Government. A.V. Sykes (SFRH/BPD/36100/2007) and D.B. Reis (SFRH/BD/7686/2011) wish to thank Fundação para a Ciência e a Tecnologia for their grants. COST FA1301-Cephs In Action ; Peer Reviewed