Impact of COVID-19 on Croatian mariculture: Findings from the first national surveys.

An overview of the negative effects of the COVID-19 pandemic on the mariculture industry of the republic of Croatia is provided. An initial online survey was circulated early after the onset of the pandemic and a follow-up field survey was performed a year into the pandemic. The surveyed companies varied in size (micro to medium enterprises), location (north, central and southern coast) and cultured organism (European flat oyster, Mediterranean mussel, European sea bass, Gilthead sea bream and/or Bluefin tuna) and were asked questions on the subject of economic and job losses, aquaculture supply chain processes and implemented or proposed measures for mitigation of negative effects. Results from the online survey showed higher economic loss than job loss, but companies reported increased job loss in the period leading to the field survey. Most companies reported reductions in sales and avenues of procurement, which, in addition to direct stressors, indirectly affected business processes. Micro enterprises fared well due to their part-time nature, low capital investments and running costs, while small to medium enterprises were under the most pressure. Large enterprises were barely affected as they had secure local and/or international distribution chains and dominated the market. Producers most affected were those that relied on the HoReCa market for product placement and/or had difficulty coping with existing stressors. Bivalve producers generally experienced a higher drop in sales than finfish farms and companies with specialized production were not able to adapt to market changes to the degree that more versatile businesses seemed capable of.

Orally administered fatty acids enhance anorectic potential but do not activate central fatty acid sensing in Senegalese sole post-larvae.

Studies in fish have reported the presence and function of fatty acid (FA)-sensing systems comparable in many aspects to those known in mammals. Such studies were carried out in juvenile and adult fish, but the presence of FA-sensing systems and control of food intake have never been evaluated in early life stages, despite the importance of establishing when appetite regulation becomes functional in larval fish. In this study, we aimed to elucidate the possible effects of different specific FAs on neural FA-sensing systems and neuropeptides involved in the control of food intake in Senegalese sole post-larvae. To achieve this, we orally administered post-larvae with different solutions containing pure FA - oleate (OA), linoleate (LA), α-linolenate (ALA) or eicosapentaenoate (EPA) - and evaluated changes in mRNA abundance of neuropeptides involved in the control of food intake and of transcripts related to putative FA-sensing systems, 3 and 6 h post-administration. The changes in neuropeptide gene expression were relatively consistent with the activation of anorectic pathways (enhanced cart4 and pomcb) and a decrease in orexigenic factors (npy) following intake of FA. Even though there were a few differences depending on the nature of the FA, the observed changes appear to suggest the existence of a putative anorectic response in post-larvae fish to the ingestion of all four tested FAs. However, changes in neuropeptides cannot be explained by the integration of metabolic information regarding FAs in circulation through FA-sensing mechanisms in the brain. Only the reduction in mRNA levels of the FA metabolism gene acc in OA-treated (6 h), ALA-treated (3 h) and EPA-treated (3 and 6 h) post-larvae could be indicative of the presence of a FA-sensing system, but most genes either were not significantly regulated (fat/cd36-lmp2, acly, kir6.x, srebp1c) or were affected in a way that was inconsistent with FA-sensing mechanisms (fat/cd36-pg4l, fas, cpt1.1, cpt1.2, cpt1.3, sur, pparα and lxrα).

Dietary fatty acid composition affects food intake and gut-brain satiety signaling in Senegalese sole (Solea senegalensis, Kaup 1858) larvae and post-larvae.

Little is known how dietary lipids affect food intake during larval development of fish, especially with regard to fatty acid (FA) composition. In fact, very little work has been done on appetite regulation and food intake in fish larvae in general, due to biological and technical difficulties associated with this type of studies. A new method using fluorescent microspheres as markers was developed in this study to evaluate food intake and prey selectivity of Senegalese sole larvae and post-larvae. Food intake was quantified in fish fed Artemia metanauplii enriched with oils differing in FA profile: cod liver oil (CLO), linseed oil (LSO), soybean oil (SBO) or olive oil (OO). The fish did not preferentially ingest a specific diet when presented with a choice. However, pre-metamorphic larvae from the CLO treatment ingested more metanauplii per g body weight, while differences in post-larvae were not significant. These findings were developed further by analyzing mRNA levels of a range of putative anorexigenic (pyya, pyyb, glp1, cckl, cart1a, cart1b, cart2a, cart4, pomca, pomcb, crf) and orexigenic (gal, npy, agrp2) genes, to identify those which are significantly affected by feeding and/or dietary FA composition. The variety of expression patterns observed highlighted the complexity of appetite regulatory mechanisms. In general, fish fed the CLO diet tended to show gene expression patterns most dissimilar to the remaining treatments. Expression in pre-metamorphic larvae was generally less in accordance with the putative function of the genes than in post-larvae, which could suggest a yet underdeveloped regulatory system.

Dietary Fatty Acid Metabolism is Affected More by Lipid Level than Source in Senegalese Sole Juveniles: Interactions for Optimal Dietary Formulation.

This study analyses the effects of dietary lipid level and source on lipid absorption and metabolism in Senegalese sole (Solea senegalensis). Juvenile fish were fed 4 experimental diets containing either 100 % fish oil (FO) or 25 % FO and 75 % vegetable oil (VO; rapeseed, linseed and soybean oils) at two lipid levels (~8 or ~18 %). Effects were assessed on fish performance, body proximate composition and lipid accumulation, activity of hepatic lipogenic and fatty acid oxidative enzymes and, finally, on the expression of genes related to lipid metabolism in liver and intestine, and to intestinal absorption, both pre- and postprandially. Increased dietary lipid level had no major effects on growth and feeding performance (FCR), although fish fed FO had marginally better growth. Nevertheless, diets induced significant changes in lipid accumulation and metabolism. Hepatic lipid deposits were higher in fish fed VO, associated to increased hepatic ATP citrate lyase activity and up-regulated carnitine palmitoyltransferase 1 (cpt1) mRNA levels post-prandially. However, lipid level had a larger effect on gene expression of metabolic (lipogenesis and ß-oxidation) genes than lipid source, mostly at fasting. High dietary lipid level down-regulated fatty acid synthase expression in liver and intestine, and increased cpt1 mRNA in liver. Large lipid accumulations were observed in the enterocytes of fish fed high lipid diets. This was possibly a result of a poor capacity to adapt to high dietary lipid level, as most genes involved in intestinal absorption were not regulated in response to the diet.

Hypothalamic fatty acid sensing in Senegalese sole (Solea senegalensis): response to long-chain saturated, monounsaturated, and polyunsaturated (n-3) fatty acids.

We assessed the presence of fatty acid (FA)-sensing mechanisms in hypothalamus of Senegalese sole (Solea senegalensis) and investigated their sensitivity to FA chain length and/or level of unsaturation. Stearate (SA, saturated FA), oleate (OA, monounsaturated FA of the same chain length), α-linolenate [ALA, a n-3 polyunsaturated fatty acid (PUFA) of the same chain length], and eicosapentanoate (EPA, a n-3 PUFA of a larger chain length) were injected intraperitoneally. Parameters related to FA sensing and neuropeptide expression in the hypothalamus were assessed after 3 h and changes in accumulated food intake after 4, 24, and 48 h. Three FA sensing systems characterized in rainbow trout were also found in Senegalese sole and were activated by OA in a way similar to that previously characterized in rainbow trout and mammals. These hypothalamic FA sensing systems were also activated by ALA, differing from mammals, where n-3 PUFAs do not seem to activate FA sensors. This might suggest additional roles and highlights the importance of n-3 PUFA in fish diets, especially in marine species. The activation of FA sensing seems to be partially dependent on acyl chain length and degree of saturation, as no major changes were observed after treating fish with SA or EPA. The activation of FA sensing systems by OA and ALA, but not SA or EPA, is further reflected in the expression of hypothalamic neuropeptides involved in the control of food intake. Both OA and ALA enhanced anorexigenic capacity compatible with the activation of FA sensing systems.

Characterization of seven cocaine- and amphetamine-regulated transcripts (CARTs) differentially expressed in the brain and peripheral tissues of Solea senegalensis (Kaup).

CART (cocaine- and amphetamine-regulated transcript) is a peptide with neurotransmitter and neuroendocrine functions with several key roles, both centrally and peripherally. In mammals there is a single gene that produces two alternatively spliced variants in rat and a single transcript in human but in teleosts multiple genes have been found. In the present study we report the existence of seven transcripts in Senegalese sole and characterize their sequences and phylogenetic relationships, as well as their expression patterns in the brain and peripheral tissues, and in response to feeding. Both cart2a and cart4 showed a ubiquitous expression in the brain, while cart1a, cart1b and cart3a were similarly expressed and had higher transcript levels in the mesencephalon, followed by the diencephalon. On the other hand, cart2b showed a main expression in the olfactory bulbs, and cart3b was predominantly expressed in the spinal cord. The expression profile in peripheral tissues differed substantially between cart's, even between more recently duplicated genes. Collectively, all the tissues examined, except the muscle, express at least one of the different cart's, although the highest transcript levels were found in the brain, gonads (ovary and testis) and, in some cases, eye and kidney. Concerning the feeding response, only brain cart1a, cart2a and cart4 showed a significant postprandial regulation, although future studies are necessary to assess potential confounding effects of stress imposed by the force feeding technique employed. Senegalese sole exhibits the highest number of cart genes reported to date in a vertebrate species. Their differential expression patterns and feeding regulation suggest that multiple cart genes, resulting from at least 3 rounds of whole genome duplication, have been retained in fish genomes through subfunctionalization, or possibly even through neofunctionalization.