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Fish skeletal development has long been correlated with nutritional factors. Lack of zebrafish nutritional standardization, especially during the early stages, decreases the reproducibility of the conducted research. The present study represents an evaluation of four commercial (A, D, zebrafish specific; B, generic for freshwater larvae; C, specific for marine fish larvae) and one experimental (Ctrl) early diets on zebrafish skeletal development. Skeletal abnormalities rates in the different experimental groups were assessed at the end of the larval period (20 days post-fertilization, dpf) and after a swimming challenge test (SCT, 20–24 dpf). At 20 dpf, results revealed a significant effect of diet on the rate of caudal-peduncle scoliosis and gill-cover abnormalities, which were relatively elevated in B and C groups. SCT results focused on swimming-induced lordosis, which was comparatively elevated in diets C and D (83% ± 7% and 75% ± 10%, respectively, vs. 52% ± 18% in diet A). No significant effects of dry diets were observed on the survival and growth rate of zebrafish. Results are discussed with respect to the deferential diet composition between the groups and the species requirements. A potential nutritional control of haemal lordosis in finfish aquaculture is suggested.

Lordosis of the haemal part of the vertebral column is a frequent abnormality in reared fish. Haemal lordosis develops during the late larval and early juvenile life stages of fish, mainly due to the high swimming activity of the fish in the rearing tanks. In the present study, we have examined whether haemal lordosis recovers during the growth of European seabass. Furthermore, we aimed to develop simple morphometric indices (PrAn1 and PrAn2) that might link the severity of lordosis at the juvenile stage with fish morphological quality at harvesting. At 111 days post-hatching (dph, 53 ± 4 mm standard length, SL), 600 seabass juveniles with lordotic (L, 200 fish) or normal (N, 400 fish) external morphology were selected and introduced in a common tank. At 150 dph (75 ± 7 mm SL), 350 fish were randomly selected, pit-tagged and transferred to a sea cage for on-growing up to 502 dph (234 ± 16 mm SL). The morphological examination of the fish at 150 and 502 dph revealed that 60% (46 out of 77) of L juveniles turned into normal phenotype by the end of on-growing period. Interestingly, 56% of the fish with recovered external morphology (N-Rec) presented either a completely normal vertebral column (31%) or minor abnormalities of individual vertebrae (25%). Following the results of geometric morphometric analysis, the differences in the body shape between N-Rec and N fish were not statistically significant (p > 0.05, canonical variate analysis). The examined morphometric indices were effective in discriminating the normal fish from 58% (PrAn1) to 65% (PrAn2) of lordotic juveniles. Results are discussed with respect to the mechanism of lordosis recovery, and spotlights on their application for quality control and cull out of the abnormal fish in commercial hatcheries.

Targeting in zebrafish fast growth, high survival rates and improved reproductive performance has led over the last years in variable feeding regimes between different facilities. Despite its significance on fish function and welfare, normal skeletal development has rarely been evaluated in establishing the best feeding practices for zebrafish. The aim of this study was to establish a protocol for normal skeletal development, growth and survival of zebrafish larvae through live feed-to-microdiet transition at an appropriate rate. Four feeding regimes including feeding exclusively on Artemia nauplii (A) or dry microdiet (D), and feeding on both Artemia and microdiet at two different transition rates (slow (B) or fast (C)) were applied from 5 to 24 dpf (days post-fertilization). Results demonstrated a significant effect of feeding regimes on the incidence of skeletal abnormalities (gill cover, fins and vertebral column, p

The lordosis of the haemal part of the vertebral column is a frequent abnormality in reared fish. Haemal lordosis develops during the late larval and early juvenile period, mainly due to the high swimming activity of the fish in the rearing tanks. In the present study we examined whether haemal lordosis recovers during the growth of European sea bass. Furthermore, we aimed to develop simple morphometric indices (PrAn1 and PrAn2) that could link the severity of lordosis at the juvenile stage with fish morphological quality at harvesting. At 111 days post-hatching (dph, 53±4 mm standard length, SL), 600 seabass juveniles with lordotic (L, 200 fish) or normal (N, 400 fish) external morphology were selected and introduced in a common tank. At 150 dph (75±7 mm SL), 350 fish were randomly selected, pit-tagged and transferred in a sea cage for on-growing up to 502 dph (234±16 mm SL). The morphological examination of the fish at 150 and 502 dph revealed that the 60% (46 out of 77) of L juveniles turned into normal phenotype by the end of on-growing period. Interestingly, 56% of the fish with recovered external morphology (N-Rec) presented either a completely normal vertebral column (31%) or minor abnormalities of individual vertebrae (25%). Following the results of geometric morphometric analysis, the differences in the body shape between N-Rec and N fish were not significant (p>0.05, canonical variate analysis). The examined morphometric indices were effective in discriminating the normal fish from the 58% (PrAn1) to 65% (PrAn2) of lordotic juveniles. Results are discussed with respect to the mechanism of lordosis recovery, as well as to their application for the quality control and cull out of the abnormal fish in the commercial hatcheries.

Temperatures experienced during early ontogeny significantly influence fish phenotypes, with clear consequences for the wild and reared stocks. We examined the effect of temperature (17, 20, or 23°C) during the short embryonic and yolk-sac larval period, on the swimming performance and skeleton of metamorphosing Gilthead seabream larvae. In the following ontogenetic period, all fish were subjected to common temperature (20°C). The critical swimming speed of metamorphosing larvae was significantly decreased from 9.7 ± 0.6 TL/s (total length per second) at 17°C developmental temperature (DT) to 8.7 ± 0.6 and 8.8 ± 0.7 TL/s at 20 and 23°C DT respectively (p p ,p p&thinsp

Titanium dioxide nanoparticles (TiO2-NPs) are among the most popular manufactured and widely used nanoparticles. They are released into the environment, affecting terrestrial and aquatic ecosystems, with unexpected consequences to organisms and human health. The present study investigates the mediated toxicity imposed to the freshwater fish species, zebrafish (Danio rerio) and the prussian carp (Carassius gibelio), and to the terrestrial land snail Cornu aspersum, after their exposure to sublethal concentrations of TiO2-NPs. Oxidative, proteolytic, genotoxic and apoptotic parameters in fish liver and gills, as well as on snail hemocytes were studied and the swimming performance was estimated in order to (a) estimate and suggest the most susceptible animal, and (b) propose a common battery of biomarkers as the most suitable indicator for biomonitoring studies against TiO2-NPs. Our in vivo experiments demonstrated that NPs induced detrimental effects on animal physiology and swimming behavior, while no general pattern was observed in species and tissues responsiveness. Generally, TiO2-NPs seemed to activate a group of molecules that are common for aquatic as well as terrestrial animals, implying the existence of a conserved mechanism. It seems that after exposure to TiO2-NPs, a common mechanism is activated that involves the stimulation of immune system with the production of ROS, damage of lysosomal membrane, protein carbonylation, lipid peroxidation, DNA damage, following proteolysis by ubiquitin and finally apoptosis. Thus, the simultaneous use of the latter biomarkers could be suggested as a reliable multi parameter approach for biomonitoring of aquatic and terrestrial ecosystems against TiO2-NPs. Keywords: Cornu aspersum, Danio rerio, Carassius gibelio, Oxidative stress biomarkers, Swimming performance, Apoptosis

Titanium dioxide nanoparticles (TiO

The anabolic effect of exercise on muscles and bones is well documented. In teleost fish, exercise has been shown to accelerate skeletogenesis, to increase bone volume, and to change the shape of vertebral bodies. Still, increased swimming has also been reported to induce malformations of the teleost vertebral column, particularly lordosis. This study examines whether zebrafish (Danio rerio) develops lordosis as a result of continuous physical exercise. Zebrafish were subjected, for 1 week, to an increased swimming exercise of 5.0, 6.5 or 8.0 total body lengths (TL) per second. Control and exercise group zebrafish were examined for the presence of vertebral abnormalities, by in vivo examination, whole mount staining for bone and cartilage and histology and micro-computed tomography (CT) scanning. Exercise zebrafish developed a significantly higher rate of lordosis in the haemal part of the vertebral column. At the end of the experiment, the frequency of lordosis in the control groups was 0.5 +/- 1.3% and that in the exercise groups was 7.5 +/- 10.6%, 47.5 +/- 10.6% and 92.5 +/- 6.0% of 5.0, 6.5 and 8.0 TL center dot s(-1), respectively. Histological analysis and CT scanning revealed abnormal vertebrae with dorsal folding of the vertebral body end plates. Possible mechanisms that trigger lordotic spine malformations are discussed. This is the first study to report a quick, reliable and welfare-compatible method of inducing skeletal abnormalities in a vertebrate model during the post-embryonic period.