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[Background] The development of a sustainable business model with social acceptance, makes necessary to develop new strategies to guarantee the growth, health, and well-being of farmed animals. Debaryomyces hansenii is a yeast species that can be used as a probiotic in aquaculture due to its capacity to i) promote cell proliferation and differentiation, ii) have immunostimulatory effects, iii) modulate gut microbiota, and/or iv) enhance the digestive function. To provide inside into the effects of D. hansenii on juveniles of gilthead seabream (Sparus aurata) condition, we integrated the evaluation of the main key performance indicators coupled with the integrative analysis of the intestine condition, through histological and microbiota state, and its transcriptomic profiling., [Results] After 70 days of a nutritional trial in which a diet with low levels of fishmeal (7%) was supplemented with 1.1% of D. hansenii (17.2 × 105 CFU), an increase of ca. 12% in somatic growth was observed together with an improvement in feed conversion in fish fed a yeast-supplemented diet. In terms of intestinal condition, this probiotic modulated gut microbiota without affecting the intestine cell organization, whereas an increase in the staining intensity of mucins rich in carboxylated and weakly sulphated glycoconjugates coupled with changes in the affinity for certain lectins were noted in goblet cells. Changes in microbiota were characterized by the reduction in abundance of several groups of Proteobacteria, especially those characterized as opportunistic groups. The microarrays-based transcriptomic analysis found 232 differential expressed genes in the anterior-mid intestine of S. aurata, that were mostly related to metabolic, antioxidant, immune, and symbiotic processes., [Conclusions] Dietary administration of D. hansenii enhanced somatic growth and improved feed efficiency parameters, results that were coupled to an improvement of intestinal condition as histochemical and transcriptomic tools indicated. This probiotic yeast stimulated host-microbiota interactions without altering the intestinal cell organization nor generating dysbiosis, which demonstrated its safety as a feed additive. At the transcriptomic level, D. hansenii promoted metabolic pathways, mainly protein-related, sphingolipid, and thymidylate pathways, in addition to enhance antioxidant-related intestinal mechanisms, and to regulate sentinel immune processes, potentiating the defensive capacity meanwhile maintaining the homeostatic status of the intestine.

Following a request from the European Commission, EFSA was asked to deliver a scientific opinion on the safety and efficacy of l-valine produced by fermentation using a non-genetically modified strain of Corynebacterium glutamicum (CGMCC 18932). l-Valine is intended to be used in feed and water for drinking as a nutritional additive, functional group amino acids, their salts and analogues, for all animal species and categories. The production strain is considered suitable for the qualified presumption of safety (QPS) approach to safety assessment. The FEEDAP Panel concluded that l-valine produced by C. glutamicum CGMCC 18932 is considered safe for the target species when supplemented to the diet in appropriate amounts according to the nutritional needs of the target species. The use of l-valine produced using C. glutamicum CGMCC 18932 in animal nutrition is considered safe for the consumer and for the environment. No conclusion could be drawn on the potential of l-valine produced using C. glutamicum CGMCC 18932 to be irritant to the skin or eyes, or a dermal sensitiser due to the lack of data. The FEEDAP Panel concluded that the l-valine produced by fermentation using C. glutamicum CGMCC 18932 is an efficacious source of the essential amino acid l-valine for non-ruminant nutrition. To be as efficacious in ruminants as in non-ruminants, supplemental l-valine requires protection against ruminal degradation. The FEEDAP Panel expressed concerns on the use of amino acids in water for drinking for hygienic reasons.

Following a request from the European Commission, EFSA was asked to deliver a scientific opinion on the feed additive consisting of Lactiplantibacillus plantarum (formerly Lactobacillus plantarum) CECT 8350 and Limosilactobacillus reuteri (formerly Lactobacillus reuteri) CECT 8700 (AQ02) as a zootechnical feed additive for suckling piglets. In a previous opinion the FEEDAP Panel concluded that the additive is considered safe for the target species, the consumer, and the environment. The Panel concluded that the additive should be considered a respiratory sensitiser but could not conclude on the skin/eye irritation potential or on its skin sensitisation potential. The Panel previously could not conclude on the efficacy of AQ02. The applicant has provided supplementary information to support the efficacy of the additive in suckling piglets. Based on the data provided, the FEEDAP Panel could not conclude on the efficacy of the additive.

Following a request from the European Commission, EFSA was asked to deliver a scientific opinion on the safety and efficacy of l-valine produced by fermentation using a non-genetically modified strain of Corynebacterium glutamicum (CGMCC 18932). l-Valine is intended to be used in feed and water for drinking as a nutritional additive, functional group amino acids, their salts and analogues, for all animal species and categories. The production strain is considered suitable for the qualified presumption of safety (QPS) approach to safety assessment. The FEEDAP Panel concluded that l-valine produced by C. glutamicum CGMCC 18932 is considered safe for the target species when supplemented to the diet in appropriate amounts according to the nutritional needs of the target species. The use of l-valine produced using C. glutamicum CGMCC 18932 in animal nutrition is considered safe for the consumer and for the environment. No conclusion could be drawn on the potential of l-valine produced using C. glutamicum CGMCC 18932 to be irritant to the skin or eyes, or a dermal sensitiser due to the lack of data. The FEEDAP Panel concluded that the l-valine produced by fermentation using C. glutamicum CGMCC 18932 is an efficacious source of the essential amino acid l-valine for non-ruminant nutrition. To be as efficacious in ruminants as in non-ruminants, supplemental l-valine requires protection against ruminal degradation. The FEEDAP Panel expressed concerns on the use of amino acids in water for drinking for hygienic reasons.

Following a request from the European Commission, EFSA was asked to deliver a scientific opinion on the feed additive consisting of Lactiplantibacillus plantarum (formerly Lactobacillus plantarum) CECT 8350 and Limosilactobacillus reuteri (formerly Lactobacillus reuteri) CECT 8700 (AQ02) as a zootechnical feed additive for suckling piglets. In a previous opinion the FEEDAP Panel concluded that the additive is considered safe for the target species, the consumer, and the environment. The Panel concluded that the additive should be considered a respiratory sensitiser but could not conclude on the skin/eye irritation potential or on its skin sensitisation potential. The Panel previously could not conclude on the efficacy of AQ02. The applicant has provided supplementary information to support the efficacy of the additive in suckling piglets. Based on the data provided, the FEEDAP Panel could not conclude on the efficacy of the additive.

The inclusion of a medicinal plant leaf extract (MPLE) from sage (Salvia officinalis) and lemon verbena (Lippia citriodora), rich in verbascoside and triterpenic compounds like ursolic acid, was evaluated in gilthead seabream (Sparus aurata) fed a low fishmeal-based diet (48% crude protein, 17% crude fat, 21.7 MJ kg-1, 7% fishmeal, 15% fish oil) for 92 days. In particular, the study focused on the effect of these phytogenic compounds on the gut condition by analyzing the transcriptomic profiling (microarray analysis) and histological structure of the intestinal mucosa, as well as the histochemical properties of mucins stored in goblet cells. A total number of 506 differentially expressed genes (285 up- and 221 down-regulated) were found when comparing the transcriptomic profiling of the intestine from fish fed the control and MPLE diets. The gut transcripteractome revealed an expression profile that favored biological mechanisms associated to the 1) immune system, particularly involving T cell activation and differentiation, 2) gut integrity (i.e., adherens and tight junctions) and cellular proliferation, and 3) cellular proteolytic pathways. The histological analysis showed that the MPLE dietary supplementation promoted an increase in the number of intestinal goblet cells and modified the composition of mucins’ glycoproteins stored in goblet cells, with an increase in the staining intensity of neutral mucins, as well as in mucins rich in carboxylated and weakly sulfated glycoconjugates, particularly those rich in sialic acid residues. The integration of transcriptomic and histological results showed that the evaluated MPLE from sage and lemon verbena is responsible for the maintenance of intestinal health, supporting gut homeostasis and increasing the integrity of the intestinal epithelium, which suggests that this phytogenic may be considered as a promising sustainable functional additive for aquafeeds.

The inclusion of a medicinal plant leaf extract (MPLE) from sage (Salvia officinalis) and lemon verbena (Lippia citriodora), rich in verbascoside and triterpenic compounds like ursolic acid, was evaluated in gilthead seabream (Sparus aurata) fed a low fishmeal-based diet (48% crude protein, 17% crude fat, 21.7 MJ kg-1, 7% fishmeal, 15% fish oil) for 92 days. In particular, the study focused on the effect of these phytogenic compounds on the gut condition by analyzing the transcriptomic profiling (microarray analysis) and histological structure of the intestinal mucosa, as well as the histochemical properties of mucins stored in goblet cells. A total number of 506 differentially expressed genes (285 up- and 221 down-regulated) were found when comparing the transcriptomic profiling of the intestine from fish fed the control and MPLE diets. The gut transcripteractome revealed an expression profile that favored biological mechanisms associated to the 1) immune system, particularly involving T cell activation and differentiation, 2) gut integrity (i.e., adherens and tight junctions) and cellular proliferation, and 3) cellular proteolytic pathways. The histological analysis showed that the MPLE dietary supplementation promoted an increase in the number of intestinal goblet cells and modified the composition of mucins’ glycoproteins stored in goblet cells, with an increase in the staining intensity of neutral mucins, as well as in mucins rich in carboxylated and weakly sulfated glycoconjugates, particularly those rich in sialic acid residues. The integration of transcriptomic and histological results showed that the evaluated MPLE from sage and lemon verbena is responsible for the maintenance of intestinal health, supporting gut homeostasis and increasing the integrity of the intestinal epithelium, which suggests that this phytogenic may be considered as a promising sustainable functional additive for aquafeeds.

A microencapsulated feed additive composed by garlic, carvacrol and thymol essential oils (EOs) was evaluated regarding its protective effect in gills parasitized by Sparicotyle chrysophrii in Sparus aurata. A nutritional trial (65 days) followed by a cohabitation challenge with parasitized fish (39 days) were performed. Transcriptomic analysis by microarrays of gills of fish fed the EOs diet showed an up-regulation of genes related to biogenesis, vesicular transport and exocytosis, leukocyte-mediated immunity, oxidation–reduction and overall metabolism processes. The functional network obtained indicates a tissue-specific pro-inflammatory immune response arbitrated by degranulating acidophilic granulocytes, sustained by antioxidant and anti-inflammatory responses. The histochemical study of gills also showed an increase of carboxylate glycoproteins containing sialic acid in mucous and epithelial cells of fish fed the EOs diet, suggesting a mucosal defence mechanism through the modulation of mucin secretions. The outcomes of the in vivo challenge supported the transcriptomic results obtained from the nutritional trial, where a significant reduction of 78% in the abundance of S. chrysophrii total parasitation and a decrease in the prevalence of most parasitic developmental stages evaluated were observed in fish fed the EOs diet. These results suggest that the microencapsulation of garlic, carvacrol and thymol EOs could be considered an effective natural dietary strategy with antiparasitic properties against the ectoparasite S. chrysophrii.

Un aditivo comercial compuesto por aceites esenciales de ajo, timol y carvacrol, AROTEC-G®, se usó en la formulación de una dieta para dorada (Sparus aurata) en un ensayo nutricional como estrategia de protección contra el ectoparásito Sparicotyle chrysophrii. Como dieta control se utilizó una dieta standard (46 % de proteína). No se observaron diferencias significativas en el índice de crecimiento entre los peces alimentados con la dieta AROTEC-G® y la dieta control. El perfil transcriptómico de piel obtenido por microarrays demuestra que los genes diferencialmente expresados relacionados con el procesamiento de RNA, transporte vesicular y biogénesis están en su mayoría sobre-expresados en los peces alimentados con AROTEC-G® comparado con los peces alimentados con la dieta control. En el moco de la piel los resultados revelan una reducción significativa de los biomarcadores clásicos asociados a respuesta de estrés. En las branquias, el estudio histopatológico indica una mejora en la condición de las branquias en peces alimentados con AROTEC-G®. Estas alteraciones coinciden con el aumento de la sobre-expresión de genes diferencialmente expresados relacionados con el sistema inmune, status redox, y respuesta metabólica en peces alimentados con el aditivo. Posteriormente, se realizó un desafío por cohabitación con S. chrysophrii, donde se observó una disminución significativa de la prevalencia de parásitos en los peces alimentados con AROTEC-G®. En conjunto, los resultados sugieren que AROTEC-G® es un compuesto natural interesante para la formulación de dietas funcionales destinadas a proteger el estado de salud de la dorada frente a la infección por ectoparásitos como S. chrysophrii.

Vibrio harveyi has been reported as the dominant heterotrophic bacterial species in western Mediterranean coastal areas during warm seasons, and is recognized as an economically significant pathogen for the aquaculture industry. The present work aimed to evaluate the pathogenicity of a V. harveyi strain isolated from ascitic fluid collected from cultured gilthead seabream and then used in a challenge experiment involving the two most important fish species in Mediterranean aquaculture: gilthead seabream, Sparus aurata and European seabass, Dicentrarchus labrax. The ascitic fluid from diseased juvenile seabreams, previously vaccinated against Photobacterium damselae and Vibrio anguillarum, was extracted and bacteria cultivated for isolation and characterization. Additionally, different tissues were sampled for histological evaluation and description. Significant histopathological responses were observed in hepatic and mucosal tissues. One of the strains isolated from ascitic fluid, IRTA 17-43, was selected for a bacterial challenge. Additionally, the attenuation of virulence through sequential passage of the strain on solid media was also assessed. In parallel, a co-habitation trial was performed in order to evaluate the possible transfer of the bacteria between injected and healthy individuals. Pathogenicity trials in gilthead seabream resulted in only 25% mortality when injected with 10 CFU mL, whereas, for European seabass, a mortality of 95% was recorded, with clear signs of vibriosis. When passed sequentially on solid media, the strain IRTA-17-43 showed a decrease of 35% in cumulative mortality for European seabass. No apparent transmission of the pathogen occurred during the co-habitation trial for both species. In conclusion, although few external signs of V. harveyi are observed in vaccinated carriers, internal effects of the infection were clear and severe. Although no horizontal transfer of infection was observed, the risk of occurrence between carriers and