Coordinated regulation of chromatophore differentiation and melanogenesis during the ontogeny of skin pigmentation of Solea senegalensis (Kaup, 1858)

Abnormal pigmentation of Senegalese sole has been described as one problem facing the full exploitation of its commercial production. To improve our understanding of flatfish pigmentation of this commercially important species we have evaluated eleven genes related to two different processes of pigmentation: melanophore differentiation, and melanin production. The temporal distribution of gene expression peaks corresponds well with changes in pigmentation patterns and the intensity of skin melanization. Several gene ratios were also examined to put in perspective possible genetic markers for the different stages of normal pigmentation development. Further, the phenotypic changes that occur during morphogenesis correspond well with the main transitions in gene expression that occur. Given the dramatic phenotypic alterations which flatfish undergo, including the asymmetric coloration that occurs between the ocular and the blind side, and the synchrony of the two processes of morphogenesis and pigmentation ontogenesis, these species constitute an interesting model for the study of pigmentation. In this study we present a first approximation towards explaining the genetic mechanisms for regulating pigmentation ontogeny in Senegalese sole, Solea senegalensis.Ministry of Science and Innovation (MICIIN) of the Spanish Government [AGL2008-03897-C04-01/ACU]; European Community [FP7/2007-2013-222719-LIFECYCLE]; Spanish Governmentinfo:eu-repo/semantics/publishedVersio

Thermal imprinting modifies adult stress and innate immune responsiveness in the teleost sea bream

The impact of thermal imprinting on the plasticity of the hypothalamic-pituitary-interrenal (HPI) axis and stress response in an adult ectotherm, the gilthead sea bream (Sparus aurata, L.), during its development was assessed. Fish were reared under 4 thermal regimes, and the resulting adults exposed to acute confinement stress and plasma cortisol levels and genes of the HPI axis were monitored. Changes in immune function, a common result of stress, were also evaluated using histomorphometric measurements of melanomacrophages centers (MMCs) in the head kidney and by monitoring macrophage-related transcripts. Thermal history significantly modified the HPI responsiveness in adult sea bream when eggs and larvae were reared at a higher than optimal temperature (HT, 22 degrees C), and they had a reduced amplitude in their cortisol response and significantly upregulated pituitary pomc and head kidney star transcripts. Additionally, after an acute stress challenge, immune function was modified and the head kidney of adult fish reared during development at high temperatures (HT and LHT, 18-22 degrees C) had a decreased number of MMCs and a significant downregulation of dopachrome tautomerase. Thermal imprinting during development influenced adult sea bream physiology and increased plasma levels of glucose and sodium even in the absence of an acute stress in fish reared under a high-low thermal regime (HLT, 22-18 degrees C). Overall, the results demonstrate that temperature during early development influences the adult HPI axis and immune function in a teleost fish.project Lifecycle EU-FP7 [222719]FCT- Foundation for Science and Technology [CCMAR/Multi/04326/2013]info:eu-repo/semantics/publishedVersio

Diets containing shrimp protein hydrolysates provided protection to European sea bass (Dicentrarchus labrax) affected by a Vibrio pelagius natural infection outbreak

This study was conducted to investigate the effects of the dietary supplementation of shrimp protein hydrolysate (SPH) on somatic growth performance, innate immune response in juvenile European sea bass (Dicentrarchus labrax) and their differential cumulative mortality when affected by a Vibrio pelagius natural infection outbreak. A diet containing 20% fish meal (FM) was used as a control, whereas three other diets differing in the level of FM inclusion (75 and 25% FM replacement by plant protein sources) and the inclusion of the additive (5% FM, 5% FM + 5% SPH and 15% FM + 5% SPH) were tested. After 110 days, there were no statistically significant differences in somatic growth parameters nor proximate composition in fish fed different experimental diets (P > 0.05), while the humoral non-specific immune responses (lysozyme, bacteriolytic and complement activities) were significantly enhanced by the inclusion of SPH in diets (P < 0.05). Additionally, an outbreak of the pathogenic bacteria V. pelagius, a bacterial species previously described as producer of the virulence factor hemolysin, occurred in all experimental tanks (4 replicates per diet) due to crowding and repeated handling stress for fish sorting. Survival rates among different experimental groups ten days after the bacterial epizootic differed depending on the diets, with groups containing SPH showing the best results (P < 0.05). In particular, fish fed the 15% FM + 5% SPH diet showed the highest survival rate (96.4 ± 5.0%), followed by those fed the 5% FM5 + 5% SPH5 (61.8 ± 16.3%). In contrast, survival rates in fish fed diets deprived of the additive (20% FM and 5% FM5 diets) were the lowest ones (32.0 ± 6.7% and 38.2 ± 13.5%, respectively). The present study showed that SPH can be incorporated in aquafeeds with high levels of FM substitution by PP sources without detrimental impact on the somatic growth performance of fish. In addition, the non-specific humoral immunity in seabass and their survival when affected by an epizootic outbreak of V. pelagius were positively affected, which showed the immunomodulatory benefits of shrimp protein hydrolysate to promote health and prevent diseases in fish.info:eu-repo/semantics/acceptedVersio

Vibrio mediterranei, a Potential Emerging Pathogen of Marine Fauna: Investigation of pathogenicity using a bacterial challenge in Pinna nobilis and development of a species‐specific PCR

Aims Extreme mortality events affecting Pinna nobilis, some associated to Vibrio mediterranei, have depleted many populations of this bivalve. The objective of this study was to demonstrate pathogenicity of V. mediterranei in the host P. nobilis by performing a bacterial challenge in P. nobilis to understand if V. mediterranei has specific virulence in this host. To assist this objective, a secondary objective was to develop a species‐specific DNA diagnostic test. Methods and Results P. nobilis collected from local bays were used in a challenge experiment with V. mediterranei (strain IRTA18‐108). Virulence in the host background of P. nobilis was demonstrated at doses of 103 CFUs / animal. An alignment of published Vibrio spp. atpA sequences was used to design V. mediterranei ‐specific primers. Further, data mining of published literature and V. mediterranei genomes identified multiple virulence‐related genes (vir genes) from which specific primers were designed for PCR detection of selected genes. Conclusion V. mediterranei strain IRTA18‐108 is pathogenic in the host P. nobilis . The virulence genes sod, rtx , and mshA were identified in this strain. Temperatures of 24ºC or higher appear to trigger onset of virulence. Sensitivity and specificity of the Vm atpA PCR is useful for diagnosis of Vibriosis in shellfish. Significance and Impact of the Study The presence of previously described virulence genes have been confirmed in this strain. The specific Vm atpA PCR assay will aid management of future epizootics of this emerging pathogen of aquatic fauna, and improve surveillance capabilities for mortality events where Vibrios are suspect.info:eu-repo/semantics/acceptedVersio

Rearing European Eel (Anguilla anguilla) Elvers in a Biofloc System

European eel (Anguilla anguilla) elvers (initial body weight (BW) = 3 g) were raised in triplicate for 60 days in a biofloc system (BFT) at 21 °C. Data from the current first study evaluating this farming technology indicated that European eel elvers adapted well to BFT systems as data on growth performance (specific growth rate = 1.48% ± 0.13 BW/day and FCR = 1.05 ± 0.09) indicated, with production costs using BFT being lower than conventional RAS units. The most critical issues associated with this aquaculture system were the maintenance of the biofloc in tanks by the regular addition of refined sugar (46% C) to keep a relationship for C:N of 20:1, and the prevention of emergence of opportunistic pathogens like the monogenean Pseudodactylogyrus sp. The overall results of this study in terms of elvers’ performance and quality and the composition of the biofloc material and its microbial composition indicated that BFT, which is considered to be one of the most cost-effective, sustainable, and environmentally friendly farming systems due to its zero water exchange and improvement of feed conversion ratio by the dietary contribution of bioflocs, may be satisfactorily used for farming European eels elvers at a density of 2 kg/m3. However, further studies are needed to test this technology with older eel stages.This project received partial funding from the European Union’s Horizon 2020 research and innovation program under grant agreement No 862658.info:eu-repo/semantics/publishedVersio

Identificación de las especies europeas del género Maja (Decapoda: Brachyura: Majidae): análisis de PCR-RFLP de una región del mtADN COI y consideraciones morfológicas

Four species of crabs of the genus Maja have been described along the European coast: M. brachydactyla, M. squinado, M. goltziana and M. crispata. The commercially important species M. brachydactyla and M. squinado achieve the largest body sizes and are the most similar in morphology, and are therefore easily confused. The four species of Maja were identified using a novel morphometric index and a polymerase chain reaction followed by restriction fragment length polymorphism analysis (RFLP). The relationship between carapace length and the distance between the tips of antorbital spines was used to distinguish adults of M. brachydactyla and M. squinado. PCR-RFLP analysis of a partial sequence of the mitochondrial cytochrome oxidase type I (COI) revealed that the four species of the genus Maja can be unambiguously discriminated using the combination of restriction endonucleases enzymes HpyCH4V and Ase I. The molecular identification may be particularly useful in larvae, juvenile and young crabs, when the morphological differences found in adults are not applicable.Cuatro especies del género Maja han sido descritas en las costas europeas: M. brachydactyla, M. squinado, M. goltziana y M. crispata. Las especies M. brachydactyla y M. squinado, que tienen importancia comercial, alcanzan los tamaños más grandes y son morfológicamente muy similares, siendo muy fácil confundirlas. La identificación de las cuatro especies se ha realizado utilizando un nuevo índice morfométrico y un análisis de polimorfismos de fragmentos de restricción (RFLP). La relación entre la longitud del cefalotórax y la distancia entre los extremos distales de las espinas antorbitales se ha utilizado para la diferenciación de los adultos de M. brachydactyla y M. squinado. El análisis PCR-RFLP de una secuencia parcial de la citocromo oxidasa tipo I mitocondrial (COI) indica que las cuatro especies del género Maja pueden ser discriminadas usando una combinación de las endonucleasas HpyCH4V y Ase I. La identificación molecular puede ser particularmente útil en las larvas, juveniles y cangrejos jóvenes, cuando las diferencias morfológicas encontradas en los adultos no son aplicables

Do the Escherichia coli European Union shellfish safety standards predict the presence of Arcobacter spp., a potential zoonotic pathogen?

The genus Arcobacter comprises Campylobacter-related species, considered zoonotic emergent pathogens, the presence of which in water has been associated with fecal pollution. Discharges of fecal polluted water into the sea have been considered as one of the main reasons for the presence of Arcobacter in shellfish, and this may represent a risk for public health. In this study, the European Union shellfish food safety criteria based on levels of Escherichia coli were studied in relation to their capacity to predict the presence of Arcobacter species. In addition, the accumulation factor (AF) that measures the concentration ratio between the microbes present in the shellfish and in the water, was also studied for both bacteria. The results show that the presence of E. coli correlated with the presence of the potentially pathogenic species A. butzleri and A. cryaerophilus. However, in 26.1% of the shellfish samples (corresponding to those taken during summer months) E. coli failed to predict the presence of, for instance A. butzleri and A. skirrowii, among other species. In the rest of the samples a significant correlation between the concentration of E. coli and Arcobacter spp. (mussels and oyster; R2 = 0.744) was found. This study indicates that the presence of E. coli can predict the presence of pathogenic Arcobacter species in shellfish samples harvested from water with temperatures lower than 26.2 °C. Consumption of shellfish collected at higher temperatures which may not be permissive to the growth of E. coli but does allow growth of Arcobacter spp., may represent a risk for consumers.info:eu-repo/semantics/acceptedVersio

Dual quantitative PCR assay for identification and enumeration of Karlodinium veneficum and Karlodinium armiger combined with a simple and rapid DNA extraction method

Karlodinium is a dinoflagellate genus responsible for massive fish mortality events worldwide. It is commonly found in Alfacs Bay (NW Mediterranean Sea), where the presence of two Karlodinium species (K. veneficum and K. armiger) with different toxicities has been reported. Microscopy analysis is not able to differentiate between these two species. Therefore, new and rapid methods that accurately and specifically detect and differentiate these two species are crucial to facilitate routine monitoring, to provide early warnings and to study population dynamics. In this work, a quantitative real-time PCR (qPCR) method to detect and enumerate K. veneficum and K. armiger is presented. The ITS1 region of the ribosomal DNA was used to design species-specific primers. The specificity of the primers together with the melting curve profile provided a reliable qualitative identification and discrimination between the two Karlodinium species. Additionally, a simple and rapid DNA extraction method was used. Standard curves were constructed from 10-fold dilutions of cultured microalgae cells. Finally, the applicability of the assay was tested with field samples collected from Alfacs Bay. Results showed a significant correlation between qPCR determinations and light microscopy counts (y = 2.838 x + 564; R2 = 0.936). Overall, the qPCR method developed herein is specific, rapid, accurate, and promising for the detection of these two Karlodinium species in environmental samples.info:eu-repo/semantics/acceptedVersio